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Lorenzo CORSI

Ricercatore Universitario
Dipartimento di Scienze della Vita sede ex-Scienze Biomediche

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Pubblicazioni

2023 - Cannabidiol-rich non-psychotropic Cannabis sativa L. oils attenuate peripheral neuropathy symptoms by regulation of CB2-mediated microglial neuroinflammation [Articolo su rivista]
Borgonetti, Vittoria; Anceschi, Lisa; Brighenti, Virginia; Corsi, Lorenzo; Governa, Paolo; Manetti, Fabrizio; Pellati, Federica; Galeotti, Nicoletta
abstract

Neuropathic pain (NP) is a chronic disease that affects the normal quality of life of patients. To date, the therapies available are only symptomatic and they are unable to reduce the progression of the disease. Many studies reported the efficacy of Cannabis sativa L. (C. sativa) on NP, but no Δ9-tetrahydrocannabinol (Δ9-THC)-free extracts have been investigated in detail for this activity so far. The principal aim of this work is to investigate the potential pain-relieving effect of innovative cannabidiol-rich non-psychotropic C. sativa oils, with a high content of terpenes (K2), compared to the same extract devoid of terpenes (K1). Oral administration of K2 (25 mg kg−1) induced a rapid and long-lasting relief of pain hypersensitivity in a mice model of peripheral neuropathy. In spinal cord samples, K2 reduced mitogen-activated protein kinase (MAPKs) levels and neuroinflammatory factors. These effects were reverted by the administration of a CB2 antagonist (AM630), but not by a CB1 antagonist (AM251). Conversely, K1 showed a lower efficacy in the absence of CB1/CB2-mediated mechanisms. In LPS-stimulated murine microglial cells (BV2), K2 reduced microglia pro-inflammatory phenotype through the downregulation of histone deacetylase 1 (HDAC-1) and nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor (IKBα) and increased interleukin-10 (IL-10) expression, an important antiinflammatory cytokine. In conclusion, these results suggested that K2 oral administration attenuated NP symptoms by reducing spinal neuroinflammation and underline the important role of the synergism between cannabinoids and terpenes.

2022 - A Proteomic Platform Unveils the Brain Glycogen Phosphorylase as a Potential Therapeutic Target for Glioblastoma Multiforme [Articolo su rivista]
Ferraro, Giusy; Mozzicafreddo, Matteo; Ettari, Roberta; Corsi, Lorenzo; Chiara Monti, Maria
abstract

In the last few years, several efforts have been made to identify original strategies against glioblastoma multiforme (GBM): this requires a more detailed investigation of the molecular mechanism of GBM so that novel targets can be identified for new possible therapeutic agents. Here, using a combined biochemical and proteomic approach, we evaluated the ability of a blood-brain barrier-permeable 2,3-benzodiazepin-4-one, called 1g, to interfere with the activity and the expression of brain glycogen phosphorylase (PYGB) on U87MG cell line in parallel with the capability of this compound to inhibit the cell growth and cycle. Thus, our results highlighted PYGB as a potential therapeutic target in GBM prompting 1g as a capable anticancer drug thanks to its ability to negatively modulate the uptake and metabolism of glucose, the so-called "Warburg effect", whose increase is considered a common feature of cancer cells in respect of their normal counterparts.

2022 - A Three-Case Series of Thrombotic Deaths in Patients over 50 with Comorbidities Temporally after modRNA COVID-19 Vaccination [Articolo su rivista]
Roncati, Luca; Manenti, Antonio; Corsi, Lorenzo
abstract

Coronavirus disease 2019 (COVID-19) is the most dramatic pandemic of the new millennium; to counteract it, specific vaccines have been launched in record time under emergency use authorization or conditional marketing authorization by virtue of a favorable risk/benefit balance. Among the various technological platforms, there is that exploiting a nucleoside-modified messenger RNA (modRNA), such as Comirnaty®, and that which is adenoviral vector-based. In the ongoing pharmacovigilance, the product information of the latter has been updated about the risk of thrombotic thrombocytopenia, venous thromboembolism without thrombocytopenia and immune thrombocytopenia without thrombosis. However, from an in-depth literature review, the same adverse events can rarely occur with modRNA vaccines too. In support of this, we here report a three-case series of thrombotic deaths in patients over 50 with comorbidities temporally after Comirnaty®, investigated by means of post-mortem histopathology and immunohistochemistry. In two out of three cases, the cause of death is traced back to pulmonary microthromboses rich in activated platelets, quite similar morphologically to those described in patients who died from severe COVID-19. Even if remote in the face of millions of administered doses, clinicians should be aware of the possible thrombotic risk also after Comirnaty®, in order to avoid a misdiagnosis with potentially lethal consequences. Since COVID-19 vaccines are inoculated in subjects to be protected, maximum attention must be paid to their safety, and prophylactic measures to increase it are always welcome. In light of the evidence, the product information of modRNA COVID-19 vaccines should be updated about the thrombotic risk, as happened for adenoviral vector-based vaccines.

2022 - Characterization of biocompatible scaffolds manufactured by fused filament fabrication of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate [Articolo su rivista]
Volpini, Valentina; Giubilini, Alberto; Corsi, Lorenzo; Nobili, Andrea; Bondioli, Federica
abstract

We characterize poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBH) scaffolds for tissue repair and regeneration, manufactured by three-dimensional fused filament fabrication (FFF). PHBH belongs to the class of polyhydroxyalkanoates with interesting biodegradable and biocompatible capabilities, especially attractive for tissue engineering. Equally, FFF stands as a promising manufacturing technology for the production of custom-designed scaffolds. We address thermal, rheological and cytotoxicity properties of PHBH, placing special emphasis on the mechanical response of the printed material in a wide deformation range. Indeed, effective mechanical properties are assessed in both the linear and nonlinear regime. To warrant uniqueness of the material parameters, these are measured directly through digital image correlation, both in tension and compression, while experimental data fitting of finite-element analyses is only adopted for the determination of the second invariant coefficient in the nonlinear regime. Mechanical data are clearly porosity dependent, and they are given for both the cubic and the honeycomb infill pattern. Local strain spikes due to the presence of defects are observed and measured: those falling in the range 70–100% lead to macro-crack development and, ultimately, to failure. Results suggest the significant potential attached to FFF printing of PHBH for customizable medical devices which are biocompatible and mechanically resilient.

2022 - Chemical characterization of non-psychoactive Cannabis sativa L. extracts, in vitro antiproliferative activity and induction of apoptosis in chronic myelogenous leukaemia cancer cells [Articolo su rivista]
Anceschi, L.; Codeluppi, A.; Brighenti, V.; Tassinari, R.; Taglioli, V.; Marchetti, L.; Roncati, L.; Alessandrini, A.; Corsi, L.; Pellati, F.
abstract

In this study, extracts from non-psychoactive Cannabis sativa L. varieties were characterized by means of ultra high-performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC-HRMS) and their antiproliferative activity was assessed in vitro. The human chronic myelogenous leukaemia cell line K562 was chosen to investigate the mechanism of cell death. The effect on the cell cycle and cell death was analysed by flow cytometry. Proteins related to apoptosis were studied by western blotting. Mechanical properties of cells were assessed using the Micropipette Aspiration Technique (MAT). The results indicated that the cannabidiol (CBD)-rich extract inhibited cell proliferation of K562 cell line in a dose-dependent manner and induced apoptosis via caspase 3 and 7 activation. A significant decrease in the mitochondrial membrane potential was detected, together with the release of cytochrome c into the cytosol. The main apoptotic markers were not involved in the mechanism of cell death. The extract was also able to modify the mechanical properties of cells. Thus, this hemp extract and its pure component CBD deserve further investigation for a possible application against myeloproliferative diseases, also in association with other anticancer drugs.

2021 - A Fully Integrated Arduino-Based System for the Application of Stretching Stimuli to Living Cells and Their Time-Lapse Observation: A Do-It-Yourself Biology Approach [Articolo su rivista]
Ragazzini, G.; Guerzoni, J.; Mescola, A.; Di Rosa, D.; Corsi, L.; Alessandrini, A.
abstract

Mechanobiology has nowadays acquired the status of a topic of fundamental importance in a degree in Biological Sciences. It is inherently a multidisciplinary topic where biology, physics and engineering competences are required. A course in mechanobiology should include lab experiences where students can appreciate how mechanical stimuli from outside affect living cell behaviour. Here we describe all the steps to build a cell stretcher inside an on-stage cell incubator. This device allows exposing living cells to a periodic mechanical stimulus similar to what happens in physiological conditions such as, for example, in the vascular system or in the lungs. The reaction of the cells to the periodic mechanical stretching represents a prototype of a mechanobiological signal integrated by living cells. We also provide the theoretical and experimental aspects related to the calibration of the stretcher apparatus at a level accessible to researchers not used to dealing with topics like continuum mechanics and analysis of deformations. We tested our device by stretching cells of two different lines, U87-MG and Balb-3T3 cells, and we analysed and discussed the effect of the periodic stimulus on both cell reorientation and migration. We also discuss the basic aspects related to the quantitative analysis of the reorientation process and of cell migration. We think that the device we propose can be easily reproduced at low-cost within a project-oriented course in the fields of biology, biotechnology and medical engineering.

2021 - Abnormal immunothrombosis and lupus anticoagulant in a catastrophic COVID-19 recalling Asherson’s syndrome [Articolo su rivista]
Roncati, L.; Corsi, L.; Barbolini, G.
abstract

Background: Coronavirus disease 2019 (COVID-19) is a complex disease with many clinicopathological aspects, including abnormal immunothrombosis, and the full comprehension of its pathogenetic mechanisms is urgently required. Methods/Results: By means of a multidisciplinary approach, we here report a catastrophic COVID-19 in a 44-year-old Philippine male patient, discovered lupus anticoagulant (LAC)-positive shortly before death, occurred 8 days after hospitalization in a clinical scenario refractory to standard high acuity care recalling Asherson’s syndrome (catastrophic antiphospholipid syndrome). Conclusion: A parallelism between this severe form of COVID-19 and Asherson’s syndrome can be so drawn. Both the diseases in fact exhibit hypercytokinemia, thrombotic microangiopathy, disseminated intravascular coagulation and multiple organ failure, they show a relationship with viral infections, and they are burdened by a high mortality rate. A genetic predisposition to develop these two overlapping conditions may be supposed.

2021 - Micronized / ultramicronized palmitoylethanolamide (PEA) as natural neuroprotector against COVID-19 inflammation [Articolo su rivista]
Roncati, L.; Lusenti, B.; Pellati, F.; Corsi, L.
abstract

Coronavirus Disease 2019 (COVID-19) is upsetting the world and innovative therapeutic solutions are needed in an attempt to counter this new pandemic. Great hope lies in vaccines, but drugs to cure the infected patient are just as necessary. In the most severe forms of the disease, a cytokine storm with neuroinflammation occurs, putting the patient's life at serious risk, with sometimes long-lasting sequelae. Palmitoylethanolamide (PEA) is known to possess anti-inflammatory and neuroprotective properties, which make it an ideal candidate to be assumed in the earliest stage of the disease. Here, we provide a mini-review on the topic, pointing out phospholipids consumption in COVID-19, the possible development of an antiphospholipid syndrome secondary to SARS-CoV-2 infection, and reporting our preliminary single-case experience concerning to a 45-year-old COVID-19 female patient recently treated with success by micronized / ultramicronized PEA.

2021 - Nucleoside-modified messenger RNA COVID-19 vaccine platform [Articolo su rivista]
Roncati, L.; Corsi, L.
abstract

On March 11, 2020, the World Health Organization declared coronavirus disease 2019 (COVID-19) a pandemic; from that date, the vaccine race has begun, and many technology platforms to develop a specific and effective COVID-19 vaccine have been launched in several clinical trials (protein subunit, RNA-based, DNA-based, replicating viral vector, nonreplicating viral vector, inactivated virus, live attenuated virus, and virus-like particle). Among the next-generation strategies, nucleoside-modified messenger RNA vaccines appear the most attractive, not only to counteract emerging pathogens but also for the possible applications in regenerative medicine and cancer therapy. However, exactly as all innovative drugs, they deserve careful pharmacovigilance in the short and long term.

2021 - Possible Association Between DHEA and PKCε in Hepatic Encephalopathy Amelioration: A Pilot Study [Articolo su rivista]
Di Cerbo, A.; Roncati, L.; Marini, C.; Carnevale, G.; Zavatti, M.; Avallone, R.; Corsi, L.
abstract

Objective: Hepatic encephalopathy (HE) is a neuropsychiatric syndrome caused by liver failure and by an impaired neurotransmission and neurological function caused by hyperammonemia (HA). HE, in turn, decreases the phosphorylation of protein kinase C epsilon (PKCε), contributing to the impairment of neuronal functions. Dehydroepiandrosterone (DHEA) exerts a neuroprotective effect by increasing the GABAergic tone through GABAA receptor stimulation. Therefore, we investigated the protective effect of DHEA in an animal model of HE, and the possible modulation of PKCε expression in different brain area. Methods: Fulminant hepatic failure was induced in 18 male, Sprague–Dawley rats by i.p. administration of 3 g/kg D-galactosamine, and after 30 min, a group of animals received a subcutaneous injection of 25 mg/kg (DHEA) repeated twice a day (3 days). Exploratory behavior and general activity were evaluated 24 h and 48 h after the treatments by the open field test. Then, brain cortex and cerebellum were used for immunoblotting analysis of PKCε level. Results: DHEA administration showed a significant improvement of locomotor activity both 24 and 48 h after D-galactosamine treatment (****p < 0.0001) but did not ameliorate liver parenchymal degeneration. Western blot analysis revealed a reduced immunoreactivity of PKCε (*p < 0.05) following D-galactosamine treatment in rat cortex and cerebellum. After the addition of DHEA, PKCε increased in the cortex in comparison with the D-galactosamine-treated (***p < 0.001) and control group (*p < 0.05), but decreased in the cerebellum (*p < 0.05) with respect to the control group. PKCε decreased after treatment with NH4Cl alone and in combination with DHEA in both cerebellum and cortex (****p < 0.0001). MTS assay demonstrated the synergistic neurotoxic action of NH4Cl and glutamate pretreatment in cerebellum and cortex along with an increased cell survival after DHEA pretreatment, which was significant only in the cerebellum (*p < 0.05). Conclusion: An association between the DHEA-mediated increase of PKCε expression and the improvement of comatose symptoms was observed. PKCε activation and expression in the brain could inhibit GABA-ergic tone counteracting HE symptoms. In addition, DHEA seemed to ameliorate the symptoms of HE and to increase the expression of PKCε in cortex and cerebellum.

2021 - Ruolo di Rhodiola rosea L. in un modello in vitro di stress indotto da CRH [Relazione in Atti di Convegno]
Corsi, L
abstract

2020 - A novel benzodiazepine derivative that suppresses microtubules dynamics and impairs mitotic progression [Articolo su rivista]
Pirani, V; Métivier, M; Gallaud, E; Thomas, A; Ku, S; Chretien, D; Ettari, R; Giet, R; Corsi, L; Benaud, C4.
abstract

A novel 2,3-benzodiazepine-4 derivative, named 1g, has recently been shown to function as an anti-proliferative compound. We now show that it perturbs the formation of a functional mitotic spindle, inducing a spindle assembly checkpoint (SAC)-dependent arrest in human cells. Live analysis of individual microtubules indicates that 1g promotes a rapid and reversible reduction in microtubule growth. Unlike most anti-mitotic compounds, 1g does not interfere directly with tubulin, nor perturbs microtubules assembly in vitro The observation that 1g also triggers a SAC-dependent mitotic delay associated with chromosome segregation in Drosophila neural stem cells, suggests it targets a conserved microtubules regulation module in human and flies. Altogether, our results indicate that 1g is a novel promising antimitotic drug with the unique properties altering microtubules growth and mitotic spindle organization.

2020 - Hippocampal synaptic and membrane function in the DBA/2J-mdx mouse model of Duchenne muscular dystrophy [Articolo su rivista]
Bianchi, Riccardo; Eilers, Wouter; Pellati, Federica; Corsi, Lorenzo; Foster, Helen; Foster, Keith; Tamagnini, Francesco
abstract

Dystrophin deficiency is associated with alterations in cell physiology. The functional consequences of dystrophin deficiency are particularly severe for muscle physiology, as observed in Duchenne muscle dystrophy (DMD). DMD is caused by the absence of a 427 kDa isoform of dystrophin. However, in addition to muscular dystrophy symptoms, DMD is frequently associated with memory and attention deficits and epilepsy. While this may be associated with a role for dystrophin in neuronal physiology, it is not clear what neuronal alterations are linked with DMD. Our work shows that CA1 pyramidal neurons from DBA/2J-mdx mice have increased afterhyperpolarization compared to WT controls. All the other electrotonic and electrogenic membrane properties were unaffected by this genotype. Finally, basal synaptic transmission, short-term and long-term synaptic plasticity at Schaffer collateral to CA1 glutamatergic synapses were unchanged between mdx and WT controls. These data show that the excitatory component of hippocampal activity is largely preserved in DBA/2J-mdx mice. Further studies, extending the investigation to the inhibitory GABAergic function, may provide a more complete picture of the functional, network alterations underlying impaired cognition in DMD. In addition, the investigation of changes in neuronal single conductance biophysical properties associated with this genotype, is required to identify the functional alterations associated with dystrophin deficiency and clarify its role in neuronal function.

2020 - Protective Effects of Borago officinalis (Borago) on Cold Restraint Stress-Induced Gastric Ulcers in Rats: A Pilot Study [Articolo su rivista]
Di Cerbo, Alessandro; Carnevale, Gianluca; Avallone, Rossella; Zavatti, Manuela; Corsi, Lorenzo
abstract

Stress is a typical body's natural defense to a generic physical or psychic change. A specific linking mechanism between ulcer onset and psycho-physical stress prolonged exposure has been reported. We decided to investigate the possible effects of Borago officinalis L. (Borago) in preventing physical (stress)-induced gastric ulcers in a rat model. Eighty male Sprague-Dawley rats were randomly divided into 16 groups, pretreated with a control solution, omeprazole (20 mg/kg), Borago methanolic extract (25, 50, 100, 250, and 500 mg/kg), Borago organic extract (50, 100, 250, and 500 mg/kg), Borago aqueous extract (5, 10, 20, 30, and 40 mg/kg), and D(-)-2-Amino-5-phosphonovaleric acid (AP5) (25 mg/kg) and kept in stressful conditions such as water immersion and restraint-induced stress ulcers. The animals were sacrificed and their stomach scored for the severity and the number of gastric ulcers. Methanolic extract (500 mg/kg) significantly reduced both ulcer parameters (***p < 0.001 and **p < 0.01, respectively). Aqueous and organic extract significantly decreased severity score at 5 and 10 mg/kg (**p < 0.01 and ***p < 0.001, respectively), and at 250 and 500 mg/kg (***p < 0.001), respectively, while gastric ulcers' resulted number significantly reduced only at 10 mg/kg (*p < 0.05) and at 500 mg/kg (**p < 0.01), respectively. On the other hand, aqueous extract significantly increased the mucosal gastric content of cAMP (*p < 0.05) and NR2A and NR2B subunits (*p < 0.05 and **p < 0.01, respectively) at 5 mg/kg. Organic extract showed also a significant cytotoxic effect at 500 and 1,000 mg/kg with a 3T3 cell viability reduction of 43.6% (**p < 0.01) and 92.1% (***p < 0.001), respectively. Borago aqueous extract at 10 mg/kg could be considered as a potential protective agent against stress-induced ulcers, and it is reasonable to possibly ascribe such protective activity to a modulation of the NR2A and NR2B subunit expression.

2020 - Rhodiola rosea L. modulates inflammatory processes in a CRH-activated BV2 cell model [Articolo su rivista]
Borgonetti, Vittoria; Governa, Paolo; Biagi, Marco; Dalia, Pasquale; Corsi, Lorenzo
abstract

Background: Rhodiola rosea L. (Crassulaceae) has been used for years in the traditional medicine of several countries as an adaptogen drug, able to preserve homeostasis in response to stress stimuli. Currently R. rosea roots and rhizome are classified as a traditional herbal medicinal product for temporary relief of symptoms of stress, such as fatigue and sensation of weakness by the European Medicines Agency. Hypothesis/Purpose: Increasing evidences suggest the involvement of neuroinflammation in response to stress. However, whether the modulation of neuroinflammatory parameters could be involved in the antistress effect of R. rosea has been barely studied. Thus, the aim of this work is to investigate the possible modulation of molecular inflammatory processes elicited by a R. rosea roots and rhizome ethanolic extract in an in vitro model of corticotropin releasing hormone (CRH)-stimulated BV2 microglial cells. Methods: BV2 cells were stimulated with CRH 100 nM and changes in cell viability, cytokines production and heat shock protein 70 (HSP70) levels were evaluated. Intracellular pathways related to inflammation, such as nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB) nuclear translocation and mitogen-activated protein kinases (MAPK) activation were also analyzed. Results: We found that R. rosea extract (2.7% m/m rosavin and 1% m/m salidroside) 20 µg/ml was able to (MKK2), extracellular signal-regulated kinase 1/2 (ERK 1/2) and c-Jun n-terminal kinase (JNK), resulting in a reduction of HSP70 expression. Conclusion: This work expands the knowledge of the intracellular mechanisms involved in R. rosea antistress activity and may be useful for the study of other adaptogen drugs. counteract the neuroinflammatory effect of CRH by inhibiting NF-κB nuclear translocation with a mechanism of action involving the modulation of mitogen-activated protein kinase-activated protein kinase 2

2019 - Chemical Composition and In Vitro Neuroprotective Activity of Fibre-Type Cannabis sativa L. (Hemp) [Articolo su rivista]
Corsi, Lorenzo; Pellati, Federica; Brighenti, Virginia; Plessi, Nicolò; Benvenuti, Stefania
abstract

Background: Fibre-type Cannabis sativa L. (hemp) usually contains cannabidiolic acid and cannabidiol as the main non-psychoactive cannabinoids. Even though there is evidence of the neuroprotective activity of pure cannabidiol, no in vitro studies have reported so far the role of hemp extracts on neuroprotection. The objective of this study was to evaluate the neuroprotective effect of hemp extracts in in vitro cellular models of neurotoxicity. Methods: One extract was obtained from raw hemp inflorescences, while the other was prepared from the same plant material submitted to a decarboxylation process. The composition of both these extracts was evaluated by HPLC-UV/DAD. Human neuroblastoma SH-SY5Y and microglial BV-2 cell lines treated with rotenone were selected as the model of neurodegeneration. The neuroprotection of hemp extracts was assessed also in serum-free conditions both in the presence and in the absence of rotenone as the toxic agent by using the same cell lines. The neuroprotective potential of cannabidiol was tested in parallel. Results: The decarboxylated hemp extract possesses a mild neuroprotective activity on BV-2 cells treated with rotenone, higher than that of pure cannabidiol. As regards serum-free experiments, the nondecarboxylated hemp extract was the most effective neuroprotective agent toward SH-SY5Y cells, while BV-2 cells were better protected from the toxic insult by the decarboxylated extract and cannabidiol. Conclusion: Both hemp extracts and pure cannabidiol displayed a moderate neuroprotective activity in the neurotoxicity models considered in this study; in addition, they showed a trophic effect on SHSY5Y cells.

2019 - Composti antitumorali [Brevetto]
Corsi, Lorenzo; Alessandrini, Andrea; Ettari, Roberta; Bennaud, Christelle
abstract

L’invenzione riguarda l’uso di derivati derivato dell’1-(4-amino-3,5-dimetilfenil)-3,5-diidro-7,8-etilenediossi-4h-2,3-benzodiazepin-4-one come agenti antitumorali.

2019 - Effetti del concentrato, liofilizzato e alcohol free, da uva rossa fermentata liofenol in modelli in vitro di infiammazione intestinale e neurotossicità [Abstract in Atti di Convegno]
Cocetta, Veronica; Corsi, Lorenzo; Montopoli, Monica
abstract

Liofenol è un liofilizzato di vino rosso biologico piemontese messo a punto dall’esperienza dell’azienda Ca’ Novella. Liofenol ha ricevuto molta attenzione da parte della comunità scientifica per la sua attività antiproliferativa in vitro (Signorelli et al., 2015) ed antiossidante su modelli animali (Rossi et al., 2016). Negli ultimi anni Liofenol è stato ottimizzato dal punto di vista fitochimico e standardizzato in polifenoli totali (7,5-10% m/m) con alto tenore in resveratrolo, >10 mg/Kg. Sulla base delle indicazioni salutistiche e farmacologiche riguardanti il possibile uso dei polifenoli del vino sui disordini infiammatori e ossidativi, in questo lavoro Liofenol è stato testato in due modelli cellulari: linea epiteliale intestinale Caco-2 e linea neuronale SHSY-5Y. Liofenol, in maniera non concentrazione dipendente, protegge le cellule SHSY-5Y dalla neurotossicità indotta da rotenone. Nella linea Caco-2, Liofenol ha dimostrato possedere un’azione antiossidante riducendo la produzione di radicali liberi dell’ossigeno (ROS) e contrastando la riduzione di resistenza trans-epiteliale del monostrato cellulare indotta da stress ossidativo. Non ha invece mostrato effetti positivi nel contrastare l’azione data da stimolo infiammatorio. In conclusione, Liofenol può essere considerata una innovativa fonte alcohol-free di polifenoli da vino, con un’interessante prospettiva di sviluppo come alimento salutistico pensato per contrastare alterazioni ossidative ed infiammatorie, con particolare riferimento alle problematiche intestinali.

2019 - Fabrication of a low-cost on-stage cell incubator with full automation [Articolo su rivista]
Regazzini, Gregorio; Mercola, Andrea; Corsi, Lorenzo; Alessandrini, Andrea
abstract

In recent years the possibility of observing by microscopy the dynamic activity of living cells has been largely pursued. We have developed a low-cost (~ 260 euros) on-stage cell incubator for inverted optical microscopes. This device allows to keep cells in good conditions for their survival and proliferation. The device is based on the use of the Arduino microprocessor interfaced with LabView. It can be connected to a computer via USB port allowing to monitor and register all the useful parameters of the measurements: temperature, CO2 concentration and relative humidity. It consists of a closed metallic and plastic (PMMA) chassis which provides optical transparency to the petri dish in order to use interference contrast imaging techniques. The system exploits also a second Arduino microprocessor to perform autofocus of the images and to automatically acquire images at defined time intervals. Cell biology laboratories could easily construct this device to allow also students to follow dynamic processes of living cells and to practice with the DIY (Do-It-Yourself) approach to biology. At the same time, students could become familiar with the use of low-cost microprocessors like Arduino.

2019 - Glutamate Receptors and Glioblastoma Multiforme: An Old "Route" for New Perspectives [Articolo su rivista]
Corsi, L; Mescola, A; Alessandrini, A
abstract

Glioblastoma multiforme (GBM) is the most aggressive malignant tumor of the central nervous system, with poor survival in both treated and untreated patients. Recent studies began to explain the molecular pathway, comprising the dynamic structural and mechanical changes involved in GBM. In this context, some studies showed that the human glioblastoma cells release high levels of glutamate, which regulates the proliferation and survival of neuronal progenitor cells. Considering that cancer cells possess properties in common with neural progenitor cells, it is likely that the functions of glutamate receptors may affect the growth of cancer cells and, therefore, open the road to new and more targeted therapies.

2019 - Tetracyclines: insights and updates of their use in human and animal pathology and their potential toxicity [Articolo su rivista]
DI CERBO, Alessandro; Pezzuto, Federica; Guidetti, Gianandrea; Canello, Sergio; Corsi, Lorenzo
abstract

etracycline antibiotics (TCs) have been widely employed to treat bacterial infections and other pathologic conditions in humans and pets.Although most of TCs have been almost ruled out from the human clinical practice they are still used as growth promoters and to treat promiscuityand overcrowding pathologies in the intensive animal farming. As a consequence, TCs are commonly found in all ecological compartments withpotential direct or indirect toxicological effects on animals and, generally, on all living organisms. Moreover, clinical and in vitro observationsraised the hypothesis that the widespread of some adverse food reactions and, to a less extent, antibiotic resistance phenomena could be ascribed tothe presence of TCs residues in edible and non-edible tissues of intensive animal farming intended for animal and human consumption. Suchresidues may pose serious health threat, depending on the type of food and the amount of residue present.The aim of this review is to provide new insights about the clinical uses of TCs in humans and animals and their potential toxic effects as residuesin the environment or as food components. (16) (PDF) Tetracyclines: insights and updates of their use in human and animal pathology and their potential toxicity. Available from: https://www.researchgate.net/publication/330857709_Tetracyclines_insights_and_updates_of_their_use_in_human_and_animal_pathology_and_their_potential_toxicity [accessed Mar 06 2019].

2018 - Adverse food reactions in dogs due to antibiotic residues in pet food: a preliminary study [Articolo su rivista]
Di Cerbo, Alessandro; Canello, Sergio; Guidetti, Gianandrea; Fiore, Filippo; Corsi, Lorenzo; Rubattu, Nicola; Testa, Cecilia; Cocco, Raffaella
abstract

In the last decades, adverse food reactions have increased considerably in dogs and cats. In this study we report on the possible onset of food intolerances symptoms, including otitis, diarrhoea, generalised anxiety, and dermatitis in a cohort of 8 dogs consuming commercial diets. All dogs received an organic chicken-based diet for 15 days. We performed analysis of blood biochemical parameters, kibble composition, and oxytetracycline (OTC) serum concentration before and after 15 days of organic chicken-based diet supplementation. We hypothesised that a chronic intake of contaminated food enhanced by the presence of nanoparticle aggregates might be at the base of the onset of pharmacologic or idiopathic food intolerances. At the end of the evaluation period, an overall significant reduction of otitis, diarrhoea, generalised anxiety, and dermatitis was observed. Biochemical analyses indicate a significant increase in the alkaline phosphatase, from 41 to 52.5 U/L, after 15 days (••p <0.01), while a significant decrease in Gamma-glutamyl transferase and urea, from 9.37 to 6.25 U/L and from 32.13 ± 8.72 to 22.13 ± 7.8 mg/dL, respectively, was observed (•p <0.05). A significant decrease, from 0.22 to 0.02 μg/mL, in mean OTC serum concentration was also observed (••p <0.01). Composition analysis revealed the presence of OTC, calcium, aluminium, silicon, and phosphorous nanoparticle aggregates. Further research on a wider sample size would help to confirm the hypothesis proposed here.

2018 - Cannabis sativa L. and Nonpsychoactive Cannabinoids: Their Chemistry and Role against Oxidative Stress, Inflammation, and Cancer [Articolo su rivista]
Pellati, Federica; Borgonetti, Vittoria; Brighenti, Virginia; Biagi, Marco; Benvenuti, Stefania; Corsi, Lorenzo
abstract

In the last decades, a lot of attention has been paid to the compounds present in medicinal Cannabis sativa L., such as Δ9-Tetrahydrocannabinol (Δ9-THC) and cannabidiol (CBD), and their effects on inflammation and cancer-related pain. The National Cancer Institute (NCI) currently recognizes medicinal C. sativa as an effective treatment for providing relief in a number of symptoms associated with cancer, including pain, loss of appetite, nausea and vomiting, and anxiety. Several studies have described CBD as a multitarget molecule, acting as an adaptogen, and as a modulator, in different ways, depending on the type and location of disequilibrium both in the brain and in the body, mainly interacting with specific receptor proteins CB1 and CB2. CBD is present in both medicinal and fibre-Type C. sativa plants, but, unlike Δ9-THC, it is completely nonpsychoactive. Fibre-Type C. sativa (hemp) differs from medicinal C. sativa, since it contains only few levels of Δ9-THC and high levels of CBD and related nonpsychoactive compounds. In recent years, a number of preclinical researches have been focused on the role of CBD as an anticancer molecule, suggesting CBD (and CBD-like molecules present in the hemp extract) as a possible candidate for future clinical trials. CBD has been found to possess antioxidant activity in many studies, thus suggesting a possible role in the prevention of both neurodegenerative and cardiovascular diseases. In animal models, CBD has been shown to inhibit the progression of several cancer types. Moreover, it has been found that coadministration of CBD and Δ9-THC, followed by radiation therapy, causes an increase of autophagy and apoptosis in cancer cells. In addition, CBD is able to inhibit cell proliferation and to increase apoptosis in different types of cancer models. These activities seem to involve also alternative pathways, such as the interactions with TRPV and GRP55 receptor complexes. Moreover, the finding that the acidic precursor of CBD (cannabidiolic acid, CBDA) is able to inhibit the migration of breast cancer cells and to downregulate the proto-oncogene c-fos and the cyclooxygenase-2 (COX-2) highlights the possibility that CBDA might act on a common pathway of inflammation and cancer mechanisms, which might be responsible for its anticancer activity. In the light of all these findings, in this review we explore the effects and the molecular mechanisms of CBD on inflammation and cancer processes, highlighting also the role of minor cannabinoids and noncannabinoids constituents of Δ9-THC deprived hemp.

2018 - Oxytetracycline-Protein Complex: The Dark Side of Pet Food [Articolo su rivista]
DI CERBO, Alessandro; Scarano, Antonio; Pezzuto, Federica; Guidetti, Gianandrea; Canello, Sergio; Pinetti, Diego; Genovese, Filippo; Corsi, Lorenzo
abstract

Background: Worldwide antibiotic abuse represents a huge burden, which can have a deep impact on pet and human health through nutrition and medicalization representing another way of antibiotic resistance transmission. Objective: We aimed our research to determine a possible complex formation between biological bone substrates, such as proteins, and Oxytetracycline (OTC), an approved antibiotic for use in zootechny, which might determine a toxic effect on K562 cells. Method: Cell viability and HPLC-ESI/QqToF assays were used to assess potential toxicity of bone extract derived from OTC-treated chickens according to standard withdrawal times and from untreated chickens at 24, 48 and 72h of incubation. Results: Cell culture medium with ground bone from chickens reared in the presence of OTC (OTC-CCM) resulted significantly cytotoxic at every incubation time regardless of the bone concentration while cell culture medium with ground bone from chickens reared without OTC (BIO-CCM) resulted significantly cytotoxic only after 72h of incubation. HPLC-ESI/QqToF assay ruled out the possible presence of OTC main derivatives possibly released by bone within culture medium until 1 μg/mL. Conclusion: The presence of a protein complex with OTC is able to exert a cytotoxic effect once released in the medium after 24-48h of incubation.

2017 - AFM investigation of mechanical properties of glioblastoma multiforme cells and their relation to motility [Abstract in Atti di Convegno]
Mescola, A; Alessandrini, Andrea; Corsi, Lorenzo
abstract

Glial tumors are clinically classified in 4 groups according to their malignancy level. Glial tumors belonging to the IV group are called Glioblastoma Multiforme (GBM) and they are among the most aggressive brain tumors. In the recent years the mechanical phenotype of cells has been recognized as a valuable marker of their malignancy level [1-3]. Here we studied by AFM the mechanical behavior of U87mg cells when exposed to a drug which interferes with their cytoskeleton affecting also their migration ability. We found that U87mg cells exposed to the tested drug presented a decreased migration potential which is correlated with an increased stiffness of the cells and with a loss of polarity. By exploiting AFM Dynamic Mechanical Analysis we also characterized the behavior of the cells for different probing frequencies. By exploiting immunofluorescence microscopy we also investigated the effect of the tested drug on the reorganization of the cell cytoskeleton finding a strong increase of the presence of stress fibers.

2016 - A novel 2,3-benzodiazepine-4-one derivative AMPA antagonist inhibits G2/M transition and induces apoptosis in human leukemia Jurkat T cell line [Articolo su rivista]
Parenti, Sandra; Casagrande, Giacomo; Montanari, Monica; Espahbodinia, M.; Ettari, R.; Grande, Alexis; Corsi, Lorenzo
abstract

It has been shown that the antagonism of glutamate receptors activity was able inhibit proliferation and induce apoptosis in several neuronal and non-neuronal cancer cell lines. In addition, it has been shown that glutamate might facilitate the spread and growth of leukemia T cells through interactions with AMPA receptors. The aim of the present study was to investigate the modulation of cell cycle elicited by a novel 2,3-benzodiazepine-4- one non-competitive AMPA antagonist derivative in the human leukemia Jurkat T cells. Our results indicated that the 1-(4-amino-3,5-dimethylphenyl)-3,5-dihydro-7,8-ethylenedioxy-4 h-2,3- benzodiazepin-4-one, named 1 g, exerted a significant growth inhibition of leukemia Jurkat T cells in a time and dose dependent manner, arresting the transition of G2/M phase through activation of Myt-1. The molecule also induced apoptosis through the enhanced expression of the pro-apoptotic p53, and the inhibition of Bcl-2, and Bcl-xl, followed by the activation of caspase-3. The results suggested that compound 1 g might act mostly as a cytostatic rather than cytotoxic compound. Al- though further studies are necessary, in order to identify others specific pathways involved in the activity of the present molecule, the presented results identified a novel molecule acting on specific G2/M checkpoint reg- ulation pathway. Finally, our data suggest that compound 1 g might be a good molecule for future development in the cancer research

2016 - Carcinogenic potential of metal nanoparticles in BALB/3T3 cell transformation assay. [Articolo su rivista]
Sighinolfi, Gl; Artoni, Erica; E., Gatti Am; Corsi, L
abstract

Metal-based nanoparticles (NPs), are currently used in many application fields including consumer products, pharmaceuticals, and biomedical treatments. In spite to their wide applications, an in-depth study of their potential toxic effects is still lacking. The aim of the present research was to investigate the potential initiator or promoter-like activity of different metallic NPs such as gold, iron, cobalt, and cerium using the Balb/3T3 two-stage transformation assay. The results indicated that all the selected metallic NPs, except for cobalt, when used as initiators did not induce any transformation in Balb/3T3 cell line. Moreover, Au and Fe3 O4 NPs, when used in place of the tumor promoter treatment TPA, increased significantly the number of Foci/dish as compared to the MCA treatment alone. The number of Foci/dish was 2.6 for Au NPs and 2.13 for Fe3 O4 ones, similar to those obtained by the positive control treatment (MCA + TPA), whereas 1.27 for MCA treatment alone. On the contrary, CeO2 NPs did not show any difference in the number of Foci/dish, as compared to MCA alone, but it decreased the number of foci by 65% in comparison to the positive control (MCA + TPA). As expected, cobalt NPs showed an increased cytotoxicity and only a few surviving cells were found at the time of analysis showing a number of Foci/dish of 0.13. For the first time, our data clearly showed that Au and Fe3 O4 NPs act as promoters in the two stage transformational assay, suggesting the importance to fully investigate the NPs carcinogenic potential with different models.

2015 - Supplementation of omega 3 fatty acids improves oxidative stress in activated BV2 microglial cell line [Articolo su rivista]
Corsi, Lorenzo; MOMO DONGMO, BENDJ EDITH; Avallone, Rossella
abstract

Abstract Many reports have shown promising beneficial effects of long-chain polyunsaturated fatty acids (L-PUFAs) of the omega 3 series in several brain diseases. In the present study, we tested the hypothesis that omega 3 fatty acids supplement reduced pro-inflammatory functions in vitro and in vivo. We demonstrated that a supplement rich in PUFAs (SRP) increased cell viability in a dose-dependent manner suggesting its protective role against lipopolysaccharide (LPS)-induced cell death in BV2 microglial cell line. In the same cultures, the supplement rich in PUFAs reduced the reactive oxygen species (ROS) and nitric oxide (NO) production. A most prominent target for ROS management is the family of peroxisome proliferator-activated receptors (PPARs). The co-treatment with SRP and LPS increased significantly the nuclear immunoreactivity of PPAR-γwhen compared the LPS treatment alone. Moreover, the chronic administration of the SRP in rats, increased the immunoreactivity of the PPAR-γ1 protein confirming its potential neuroprotective effect.

2014 - Cytotoxic effect of hemin in colonic epithelial cell line: Involvement of 18kDa translocator protein (TSPO). [Articolo su rivista]
Gemelli, Claudia; Dongmo, Bm; Ferrarini, F; Grande, Alexis; Corsi, Lorenzo
abstract

Aims The aim of this study is to investigate the effect of hemin in colonic epithelial cells (Caco-2) cell proliferation and if this effect was due to a direct modulation of 18-kDa translocator protein (TSPO) and/or heme oxygenase type 1 (HO-1). Main methods The main methods are as follow: cell proliferation and cell cytotoxic assays on Caco-2 cell lines treated with hemin in the presence or not of 1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide (PK 11195) and Sn-protoporphyrin IX (Sn-PPIX), and immunoblotting for TSPO and HO-1 protein analysis, siRNA directed against TSPO. Key findings Hemin was shown to be toxic for the Caco-2 cell line in a concentration and time dependent manner. Although hemin was able to induce HO-1 in a dose dependent manner, a specific HO-1 inhibitor, Sn-PPIX, was unable to interfere with the effect of hemin on Caco-2 cells. Instead, PK 11195, a specific TSPO ligand, was able to counteract the effect of hemin, suggesting an important role of TSPO in the hemin activity. Cell viability assay further confirms the high cytotoxic effects exerted by hemin on Caco-2 cells expressing TSPO compared to the siRNA-TSPO targeted cells. In addition, hemin was able to decrease significantly the TSPO protein density in a dose dependent manner after 24 h of incubation. Significance The interaction and the consecutive down regulation of TSPO by hemin played an important role in the control of Caco-2 cell viability. The presented data suggest that TSPO might contribute to protect cells from potential toxic compounds such as free tetrapyrroles, candidating this receptor as a survival receptor protein.

2012 - IN VITRO TOXICITY OF METAL NANOPARTICLES IN NEURONAL AND GLIAL CELLS [Poster]
Corsi, Lorenzo; Puja, Giulia; Artoni, Erica
abstract

Several in vitro studies have indicated the potential toxicity of NPs to various types of neuronal and glial cells. SH-SY5Y neuroblastoma cell line has already been used to assess NP-induced neurotoxicity. Exposure of SH-SY5Y cells to Fe2O3, CuO and ZnO NPs was found to decrease cell viability showing a dose-dependent toxic response [3]. Similarly human glioblastoma U87MG cell line was studied to evaluate the effect of silicon dioxide nanoparticles [4]. The results showed a decreased mitochondrial energy production and an altered cell survival/proliferation signaling. At present little is known concerning the potential adverse effects of the NPs on neuronal cells, thus it seems urgent to investigate a possible NPs role in neurotoxicity. U87MG cells showed a decrease in cell viability following treatment with NPs of iron, cobalt oxide and cerium oxide at both 48 and 72h. Cobalt NPs reduced the cell viability of approximately 21% after 48 hours of exposure and the effect remained the same even after 72h. NPs of cerium oxide showed a decrease in cell viability comparable to that of cobalt NPs (21%) but only at the longer incubation time (72h). On the contrary, gold particles did not show any modulation of cell viability. In SH-SY5Y cells all the NPs tested showed a decrease of cell viability after 48 and 72 hours of exposure. Indeed, after 72h the NPs of gold and iron oxide showed a decrease of cell viability by over 16% relative to the control, whereas cobalt and of cerium oxide NPs reduced the cell viability by approximately the 32%. Rat primary cells showed no significant decline in cell viability after 72h of exposure to gold and iron oxide NPs in both sub population (glial cells and neurons). Instead cobalt and cerium oxide NPs led to a decrease of cell viability in glial cells and neurons of 56% and 23% respectively after 72h. Interestingly glial cells, as verified by ESEM imaging, internalized most of the particles but the cell mitochondrial metabolism seemed to be less affected than the neuronal one.

2011 - Anti-inflammatory activity of the non-peptidyl low molecular weight radical scavenger IAC in carrageenan-induced oedema in rats [Articolo su rivista]
Corsi, Lorenzo; Zavatti, Manuela; Geminiani, Elisa; Zanoli, Paola; Baraldi, Mario
abstract

Objective  In this research we investigated the anti-inflammatory activity of a non-peptidyl low molecular weight radical scavenger (IAC) in an acute and chronic animal model of inflammation.Methods  For this purpose the effect of IAC (10, 25, 50 mg/kg) was tested in rats on the associated behavioral responses to subsequent inflammatory and noxious challenges, such as hind paw oedema induced by intra-plantar injection of carrageenan and granuloma induced by subcutaneous implant of a cotton pellet, using indometacin (2.5 mg/kg) as reference drug. Moreover, the serum level of several cytokines was tested in the animal treated (or not) with IAC (50 mg/kg) both in the absence and presence of carrageenan-induced inflammation.Key findings  IAC showed a significant anti-inflammatory activity in both in acute and chronic models of inflammation. In addition IAC down regulated significantly the serum levels of interleukin (IL) 2 and IL6 whereas it increased the serum concentration of IL1α and glutathione.Conclusion  Although it remains to be elucidated whether or not the antioxidant property of IAC is directly responsible for the modulation of the tested cytokines, these results suggest IAC to be a possible candidate for a novel anti-inflammatory compound

2011 - In-vitro assessment of new methods for cell massive uptake based on nanoparticles transfection [Poster]
Corsi, Lorenzo; Artoni, Erica
abstract

New nanotechnological products containing nano-sized particles (NPs) are already present in the market, but the results of nanotoxicological tests are not exhaustive and sometimes contradictory. The cytotoxicity tests ruled by chemical standards appear inadequate for the NPs very peculiar adhesive superficial properties that make their handling difficult and not immediately controllable. The present study highlights a new strategy to set up meaningful in-vitro tests to assess cells massive NPs uptake. A new approach was developed to obtain homogenous dispersion of NPs in-vitro. A lipid-based transfection technique (lipofection) was evaluated in two different cell lines: a mouse fibroblast embryonic cell line (NIH/3T3) and a human hepatoblastoma derived cells (HEP-G2), using gold (Au), iron (Fe3O4), cobalt oxide (Co) and cerium oxide (CeO2) NPs. The results indicate that after massive entrance of NPs due the lipofection technique, cells showed a stimulus in the proliferation, except for Co NPs. The technique may represent an innovative approach to perform toxicity tests, inducing a homogeneous NPs exposure. Further studies should be carried out to validate and improve this method.

2010 - Effects of the Novel Non-Peptidyl Low Molecular Weight Radical Scavenger IAC in Different Models of Inflammation: A New Perspective in Anti- Inflammatory Therapy [Articolo su rivista]
Corsi, Lorenzo
abstract

The bis (1-hydroxy-2,2,6,6-tetramethyl-4-piperidinyl)-decandioate called IAC, is a new non-peptidyl low molecular weight radical scavenger able to give a fast reaction with the majority of radical species involved in the oxidative stress. This intrinsic property might be of particular interest in all the processes where it presents an over production of reactive oxygen/nitrogen species (ROS/RNS) such as inflammation. Indeed, it is well known that systemic inflammatory response is associated with the production of ROS, nitric oxide (NO), which in turn deplete the endogenous GSH, mediating cytotoxicity. It has been shown that IAC through its antioxidant activity, exerted a protective effect in vitro in islets isolated from type-2 diabetic patients, and in vivo in a non-obese diabetic mouse model and in DNBS-induced colitis in rats. The ability of IAC to protect brain from ischemia, suggests a possible use of the compound in broad range of inflammatory- related diseases. It is well known that the use of non steroidal anti-inflammatory drugs (NSAIDs) is associated with a broad spectrum of untoward side-effects such as gastrointestinal ulceration. The major pathogenetic element in the development of these effects is the depletion of prostaglandins (PGs) through inhibition of cyclooxygenase. The evidence that IAC protects gastric mucosa in an animal model of indomethacin-induced ulcer, through local increase of PGE2 levels and antioxidant activity, candidates this compound as a novel, promising, anti-inflammatory compound avoiding the major common untoward side-effects elicited by NSAID's

2009 - Anti-ulcer activity of IAC, a novel free-radical scavenger, in rats. [Articolo su rivista]
Zavatti, Manuela; Corsi, Lorenzo; Zanoli, Paola; Baraldi, Mario
abstract

Objectives We investigated the ability of a novel low-molecular-weight free-radical scavenger, bis(1-hydroxy-2,2,6,6-tetramethyl-4-piperidinyl)-decandioate (IAC), to protect the gastric mucosa from indometacin-induced ulceration.Methods The pharmacological effects of IAC, administered orally or by intraperitoneal injection, on the gastric mucosa were assessed in a rat model of gastric ulceration induced by indometacin. The effect of IAC on the level of prostaglandin PGE2 in the gastric mucosa was also investigated. Results IAC, which has no ulcerative activity per se, had a preventive and protective activity against indometacin-induced gastric ulceration. This effect could be only partially attributed to a modulatory effect of IAC on PGE2 levels; it is more likely to be due to the antioxidant activity of the compound. Conclusions Taking into account the properties of IAC and the mechanisms underlying gastric inflammation elicited by non-steroidal anti-inflammatory drugs, IAC may represent a novel anti-ulcer agent.

2009 - Natural endogenous ligands for benzodiazepine receptors in hepatic encephalopathy [Articolo su rivista]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; Venturini, I; Baraldi, Claudia; Zeneroli, Maria Luisa
abstract

Benzodiazepines of natural origin (NBZDs) have been found in human blood and brains as well as in medicinal plants and foods. In plasma and brain tissue there are i.e. diazepam and nordiazepam equal to commercial drugs but there are also other benzodiazepine-like compounds termed “endozepines”, which act as agonists at the benzodiazepine receptors of central type (CBR). A synthetic pathway for the production of NBZDs has not yet been found, but it has been suggested that micro-organisms may synthesize molecules with benzodiazepine-like structures. Hence NBZDs could be of both endogenous and exogenous source and be considered as natural anxyolitic and sedative. Interestingly there are also natural compounds, such as the polypeptide Diazepam Binding Inhibitor (DBI) acting as an “inversive agonist” implicated in fair and panic disorders. It has been suggested that NBZDs may play a role in the pathogenesis of hepatic encephalopathy (HE). Multidirectional studies evaluated NBZDs levels (1) in the blood of normal subjects, of cirrhotic with or without HE and in commercial benzodiazepine consumers; (2) in the blood of cirrhotic treated or not with a non-absorbable antibiotic; (3) in several constituents of our diet. In conclusion, NBZDs increase sometime in cirrhotics with or without HE but they reach concentrations not higher than those found in commercial benzodiazepines consumers. Hence NBZDs must be considered as occasional precipitating factor of HE and benzodiazepine antagonists only symptomatic drugs. The finding that NBZDs may be in part synthesized by intestinal bacterial flora and in part constituent of our diet underlines the importance to feed cirrhotic patients with selected food.

2009 - The phytoestrogen ferutinin affects female sexual behavior modulating ERα expression in the hypothalamus [Articolo su rivista]
Zanoli, Paola; Zavatti, Manuela; Geminiani, Elisa; Corsi, Lorenzo; Baraldi, Mario
abstract

This study was designed to assess the effect of the phytoestrogenic compound ferutinin, chronically administered in ovariectomized progesterone primed rats, alone or in combination with estradiol benzoate. After 2, 3 and 4 weeks of treatments, female rats were tested for receptive (lordosis) and proceptive behaviors (hops, darts and ear wigglings). Ferutinin given alone markedly increased the intensity of the lordotic response in ovariectomized rats but failed to significantly affect proceptivity. On the other hand estradiol benzoate significantly increased both receptive and proceptive behaviors. When administered in combination with estradiol, ferutinin reduced the increase in receptivity and proceptivity due to estrogen effects, acting as an antiestrogen. At the end of the behavioral experiments, animals were sacrificed and Western blot analysis of estrogen receptor α (ERα) levels was performed in the dissected hypothalami. Ferutinin increased ERα expression when administered alone, as estradiol did, but decreased the response to estradiol when administered in combination. These results suggest that ferutinin displays estrogenic or antiestrogenic activity through ERα in the hypothalamus, depending on the absence or the presence of estrogen priming

2008 - Peripheral benzodiazepine receptor (PBR) new insight in cell proliferation and cell differentiation review [Articolo su rivista]
Corsi, Lorenzo; Geminiani, Elisa; Baraldi, Mario
abstract

The peripheral benzodiazepine receptor (PBR), is an 18 kDa protein of the mammalian mitochondrial membrane and is a highly conserved protein among the mammalian. PBR is involved in numerous biological functions, including steroid biosynthesis, mitochondrial oxidative phosporylation and cell proliferation. The presence of PBR at the nuclear subcellular level has been demonstrated in aggressive breast cancer cell line and human glioma cells, where it seems to be involved in cell proliferation. In our previous studies we investigated the presence of nuclear PBR in different hepatic tumour cell lines with regard to binding to [3H] PK 11195 and protein analysis. The results obtained by saturation binding experiments and Scatchard analysis of nuclear PBR density in parallel with the results on the growth curves of the cell lines tested, indicate that the nuclear PBR density correlates inversely with cell doubling time. Moreover, the cell line with high nuclear PBR proliferates in response to PBR ligands, whereas that with low nuclear PBR does not. All these findings support the idea that PBR could play a pivotal role in cell proliferation and this receptor protein could be potentially important either in early diagnosis or chemopreventive strategies against degenerative disease.

2005 - Endozepines in recurrent stupor [Articolo su rivista]
Cortelli, P.; Avallone, Rossella; Baraldi, Mario; Zeneroli, Maria Luisa; Mandrioli, J.; Corsi, Lorenzo; Riva, R.; Tuniper, P.; Lugaresi, E.; Baruzzi, A.; Montagna, P.
abstract

Stupor is a condition from which the subject can be aroused only by vigorous stimuli. Most patients with stupor have a diffuse organic cerebral dysfunction. Rarely stupor is recurrent and no specific causes can be found. Patients with idiopathic recurrent stupor were awakened by i.v. administration of an antagonist (flumazenil) of the benzodiazepine recognition site located in the GABA(A) receptor. Since no exogenous benzodiazepines were detected in plasma and cerebrospinal fluid by high performance liquid chromatography, an excess of endogenous benzodiazepine-like compounds (endozepines) was proposed as the cause of stupor. The existence of endozepines, their widespread distribution in the CNS and their involvement in hepatic encephalopathy are established. However, the origin of these compounds, how biosynthesis occurs and the mechanisms and causes through which they alter brain functions are poorly understood. The fact that a number of synthetic benzodiazepines are difficult to detect using conventional techniques and the discovery that some cases of recurrent stupor were caused by fraudulent administration of lorazepam question whether the concept of endozepine recurrent stupor can be sustained. This review summarizes the state of endozepine physiology and pharmacology and the clinical syndromes attributed to their involvement. A diagnostic work-up to define endozepine-induced recurrent stupor is suggested.

2005 - Evaluation of rifaximin, placebo and lactulose in reducing the levels of benzodiazepine-like compounds in patients with liver cirrhosis: a pilot study [Articolo su rivista]
I., Venturini; A., Ferrieri; Farina, Franco; Cosenza, Francesca; Avallone, Rossella; Corsi, Lorenzo; Baraldi, Mario; Zeneroli, Maria Luisa
abstract

Benzodiazepine-like compounds (BZDs), either taken with the diet or synthesized by intestinal bacterial flora, may represent a precipitating factor for hepatic encephalopathy (HE) in cirrhotic patients. We evaluated whether a diet and/or treatment with rifaximin or lactulose can reduce serum concentrations of BZDs in 18 cirrhotic patients without HE. Patients were given a standard diet for 7 days to keep the dietary intake of BZDs constant and were then randomized to a 7-day treatment with rifaximin 1,200 mg/day, lactulose 10-20 g three times daily, or placebo. Blood samples were collected at enrollment, at the end of the diet and drug treatment periods, and 7 days after the drug was stopped (follow-up). Serum concentrations of BZDs were measured by a radioligand binding technique after high-performance liquid chromatography extraction and purification and were expressed as diazepam equivalents (DE). No change in serum BZD concentrations was observed during the diet, while a statistically significant decrease from 105.6 +/- 66.5 to 63.5 +/- 49.5 pmol DE/ml was achieved in rifaximin-treated patients (p < 0.05) but not in patients treated with lactulose or placebo. During the followup, serum BZD concentrations returned to 104.5 +/- 74.0 pmol DE/ml in rifaximin-treated patients (p < 0.05 vs. end-treatment values), while no significant change was observed in the lactulose- and placebo-treated patients. These data indicate that control of bacterial flora with cyclic administration of rifaximin plays a pivotal role in avoiding increased plasma concentrations of BZDs, which represent a precipitating factor for HE inpatients with severe liver disease.

2005 - Management of hepatic encephalopathy: Role of rifaximin [Articolo su rivista]
Zeneroli, Maria Luisa; Avallone, Rossella; Corsi, Lorenzo; I., Venturini; Baraldi, Claudia; Baraldi, Mario
abstract

Hepatic encephalopathy (HE) is a neuropsychiatric syndrome, which develops in patients with acute or chronic liver failure. It is widely accepted to be due to impairment of hepatic clearance of toxic products from the gut such as ammonia. Accumulation of ammonia induces a glutamate neurotoxicity leading to an increased tone of the gamma-aminobutyric acid A (GABA-A) receptor system in the brain which results in HE. Factors either increasing the ammonia levels (protein load, constipation, sepsis, or gastrointestinal bleeding) or potentiating the functional activity of the GABAergic system [natural benzodiazepine-like compounds (NBZDs) or exogenous benzodiazepines] may act as precipitating factors of HE. NBZDs are present in trace amounts in the blood of normal subjects and have been found to be increased in the blood of patients with liver cirrhosis, with or without HE. These compounds may derive either from the diet since they have been found in plants, vegetables and animals or from gut bacteria. The observation that intestinal bacterial flora is involved in the production of both primary agent of HE (ammonia) and precipitating factors (NBZDs) suggests that the use of nonabsorbable antibiotics such as rifaximin may be useful in preventing episodes of HE in patients with liver cirrhosis. Copyright

2005 - Nuclear location-dependent role of peripheral benzodiazepine receptor (PBR) in hepatic tumoral cell lines proliferation [Abstract in Atti di Convegno]
Geminiani, Elisa; Corsi, Lorenzo; Avallone, Rossella; Baraldi, Mario
abstract

The peripharal typo benzodiazepine receptor (PBR) was described as another class of binding sites for benzodiazepines distinct from the central benzodiazepine receptor (CBR). In our study we investigated the presence of nuclear PBR in different hepatic tumor cell lines with ragards to binding to [3H]PK11195 and protein analysis. The results obtained indicated that the nuclear PBR density correlates inversely with cell doubling time.

2005 - Nuclear location-dependent role of peripheral benzodiazepine receptor (PBR) in hepatic tumoral cell lines proliferation [Articolo su rivista]
Corsi, Lorenzo; Geminiani, Elisa; Avallone, Rossella; Baraldi, Mario
abstract

PBR is involved in numerous biological functions, including steroid biosynthesis, mitochondrial oxidative phosphorylation and cell proliferation. The presence of PBR at the perinuclear/nuclear subcellular level has been demonstrated in aggressive breast cancer cell lines and human glioma cells where it seems to be involved in cell proliferation. In our study we investigated the presence of perinuclear/nuclear PBR in different hepatic tumor cell lines with regard to binding to [H-3] PK 11195 and protein analysis. The results obtained by saturation binding experiments and scatchard analysis of perinuclear/nuclear PBR density in parallel with the results on the growth curves of the cell lines tested, indicate that the perinuclear/nuclear PBR density correlates inversely with cell doubling time. Moreover, the cell line with high perinuclear/nuclear PBR proliferated in response to PBR ligand, whereas that with low perinuclear/nuclear PBR did not. Our results reinforce the idea that the subcellular localisation of PBR defines its function and that this receptor could be a possible target for new strategies against cancer.

2004 - Peripheral benzodiazepine receptors in potatoes (Solanum tuberosum) [Articolo su rivista]
Corsi, Lorenzo; Avallone, Rossella; Geminiani, Elisa; Cosenza, Francesca; I., Venturini; Baraldi, Mario
abstract

The peripheral benzodiazepine receptor (PBR), an internal protein of the mammalian mitochondrial membrane, is involved in several metabolic functions such as steroidogenesis, oxidative phosphorylation, and regulation of cell proliferation. Here we report the presence of PBRs in parenchymal and meristematic tissues of potato (Solanum tuberosum). PBRs are heterogeneously distributed in potato and are highly expressed in meristematic cells. In particular the receptor protein is mainly localised in the meristematic nuclear subcellular preparation. This 30-36 kDa protein, which corresponds to PBR, is increased, indeed, in meristematic compared to the parenchymal tissue. This suggests an involvement of this receptor in the regulation of cell plant growth. In addition, the demonstration that PBRs are also present in vegetables supports the hypothesis of a highly conserved receptor system during phylogenesis. Published by Elsevier Inc.

2003 - Presenza di composti ad attivita’ ansiolitico-sedativa nella filera di produzione del parmigiano reggiano. [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; Puja, Giulia; Losi, Gabriele; Baraldi, Mario
abstract

Da tempo è stata descritta la presenza di sostanze ad attività sedativa nel latte tuttavia la loro struttura chimica è ancora oggetto di studio. Recentemente è stato isolato dall’s1-caseina un peptide denominato s1-CN-(f91-100) o s1-casozepina [1] mentre il nostro gruppo ha identificato nel latte altre sostanze non peptidiche ad attività benzodiazepino-simile [2]. Non è ancora noto, comunque, quali e quante sostanze presenti nel latte si ritrovano nei vari formaggi dopo i processi di produzione. Nel presente lavoro è stata valutata la presenza di composti ad attività benzodiazepino-simile nel parmigiano reggiano. In particolare è stata analizzata la filiera di produzione del parmigiano reggiano prelevando i seguenti campioni presso un’unica azienda in modo da escludere variabilità legate a fattori di alimentazione, ambiente, procedimento di produzione: fieno, latte intero, latte magro, siero, siero innesto, panna, cagliata, tosone e parmigiano reggiano. I campioni estratti con metanolo sono stati purificati mediante analisi HPLC a 0.8 ml/min utilizzando una colonna LiChrospher 100 RP-18 column (250x4.0 mm; 5 m) equilibrata con 80% di acqua/0.1% TFA e 20% di acetonitrile. I campioni sono stati analizzati a gradiente (0.5%) di acqua/0.1% TFA and acetonitrile dal 20 al 58% di acetonitrile. Per ogni campione sono state raccolte 75 frazioni, liofilizzate e testate per valutare la loro capacità di legarsi specificatamente al sito di riconoscimento centrale benzodiazepinico mediante dosaggio radioisotopici utilizzando come ligando marcato il [3H]RO 15-1788, antagonista benzodiazepinco. La concentrazione ottenuta per estrapolazione da una curva di competizione generata con autentico diazepam, viene espressa in ng diazepam equivalenti /g di campione. Nel tosone e nel parmigiano reggiano sono state rilevate piccole quantità di sostanze BDZ-simili (0,78 ± 0,12 e 0,86 ± 0,17 ng DE/g rispettivamente) mentre nel latte e nel siero la loro concentrazione risulta significativamente maggiore (117,75 ± 9,2 e 215 ± 15,7 ng DE/g rispettivamente). E’ interessante sottolineare che la concentrazione di tali sostanze nel fieno (14,8 ± 1,64 DE ng/g) è inferiore rispetto al latte, considerando che l’alimentazione della mucca è esclusivamente basata sul fieno si può ipotizzare che il contenuto che si ritrova nel latte sia anche il contributo di una sintesi endogena. L’attività ansiolitico-sedativa delle frazione isolate dal siero di latte è stata valutata sia in vitro su neuroni corticali con la tecnica elettrofisiologia del patch-clamp che in vivo.Bibliografia1. L. Miclo, E. Perrin, A. Driou, V. Papadopoulos et al. Characterization of a-casozepina, a tryptic peptide from bovine as1-caseina with benzodiazepine-like activity. FASEB J: 15, 1780-1782, 20012. R. Avallone, L. Corsi, M.L. Zeneroli and M. Baraldi. Presence of benzodiazepine-like molecules in food and their implication in the nutrition of cirrhotic patients. Innovative Food Science and Technologies: 2/3, 193-198, 2001

2003 - Presenza di sostanze Benzodiazepino-simili in estratti di foglie di tabacco (Nicotiana tabacum) [Relazione in Atti di Convegno]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; Geminiani, Elisa
abstract

Le finalità della ricerca sono rappresentate dal desiderio di chiarire le possibili utilizzazioni a fini salutistici dei singoli principi attivi presenti nelle foglie di tabacco avendo ben presenti gli efftti prodotti dai composti benzodiazepino-simili a secondo della loro capacità di interferire sui recettori centrali (attività anziolitico-sedativa e miorilassante) e su quelli periferici (steroidogenesi, proliferazione cellulare ed attività sui processi infiammatori).

2003 - Recettore periferico delle benzodiazpine e diazeoma binding inhibitor nel carcinoma epatocellulare [Abstract in Atti di Convegno]
Corsi, Lorenzo; I., Venturini; Avallone, Rossella; Cosenza, Francesca; Baraldi, Mario; Zeneroli, Maria Luisa
abstract

L'elevata espressione del recettore periferico delle benzodiazepine e del suo ligando endogeno DBI in tessuti tumorali potrebbe fortemente indicare un'implicazione di questo soistema proteico nel metabolismo delle cellule neoplastiche.

2002 - Antiproliferative effects of Ceratonia siliqua L. on mouse hepatocellular carcinoma cell line [Articolo su rivista]
Corsi, Lorenzo; Avallone, Rossella; F., Cosenza; F., Farina; Baraldi, Claudia; Baraldi, Mario
abstract

Extracts from pods and leaves of carob (Ceratonia siliqua L.) were tested for their ability to inhibit cell proliferation of mouse hepatocellular carcinoma cell line (T1). The two extracts showed a marked alteration of T1 cell proliferation in a dose-related fashion reaching the maximal effect at 1 mg/ml. Moreover, we demonstrated that leaf and pod extracts were able to induce apoptosis in T1 cell lines after 24-h treatment mediating a direct activation of the caspase 3 pathway. HPLC analysis revealed the presence of gallic acid, epigallocatechin-3-gallate and (-) epi catechin - 3 -gallate in pod and leaf extracts, compounds well known to exert antiproliferative effects. Their concentration reached 6.28 mg/g in carob leaves and 1.36 mg/g in carob pods extract. The discovery that carob pod and leaf extracts contained antiproliferative agents could be of practical importance in the development of functional foods and/or chemopreventive drugs.

2002 - Vegetable and mammalian milk as a source of benzodiazepine-like molecules [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; Baraldi, Mario
abstract

One of the challenging questions till facing benzodiazepine (BDZ) researchers is the identification of endogenous BDZ ligands in the brain, which could modulate -aminobutyric acid (GABAA) neurotrasmission. Several substances with benzodiazepine-like activity were found in food. In particular their amount was particularly high in milk. Different type of milk of animal or vegetal origin were analysed by HPLC analysis. The serum obtained by acid precipitation of milk was chromatographed at 0.8 ml/min on a LiChrospher 100 RP-18 column (250x4.0 mm; 5 m) equilibrated with 80% water/0.1% TFA and 20% acetonitrile. The sample was analysed using a water/0.1% TFA and acetonitrile gradient at 0.5% from 20 to 58% acetonitrile. 75 fraction were collected, lyophilised and tested for their ability to inhibit [3H]RO 15-1788 specific binding to central BDZ binding site or [3H]PK 11195 specific binding to peripheral BDZ binding site (PBR). The results are reported in figure. Thermic treatments, like UHT, or the biological procedures do not change the BDZ-like molecules concentration. Since the PBR is involved in cell proliferation the fractions active on PBR were tested on the ability to modulate cell proliferation of hepatocellular carcinoma cell lines (T1). The chronic treatment with the active fraction showed a significant dose-dependent inhibition of T1 cell proliferation indicating a potential use of milk as functional food.

2001 - Ammonia and endogenous benzodiazepine-like compounds in the pathogenesis of hepatic encephalopathy [Articolo su rivista]
Venturini, I.; Corsi, L.; Avallone, R.; Farina, F.; Bedogni, G.; Baraldi, C.; Baraldi, M.; Zeneroli, M. L.
abstract

Background: Ammonia and endogenous benzodiazepines (BDZs) are two of the most important agents among those taken into consideration in the pathogenesis of hepatic encephalopathy (HE). Methods: Venous ammonia and endogenous BDZs sera levels were assayed in 58 liver cirrhosis patients (34 male, 24 female) free of commercial BDZs. Endogenous BDZs were measured by binding assay after highperformance liquid chromatography purification. Ammonia was assessed by colorimetric test. Results: Endogenous BDZs and ammonia were significantly higher in Child-Pugh class C than in class B and class A (P &lt; 0.05), correlating to the severity of the liver dysfunction but not with the degree of HE. A significant difference, in fact, was noted between degree 0 (no HE) versus III-IV of HE (P &lt; 0.05), but not between degrees I-II versus III-IV. Regression analysis performed to find a correlation between the ammonia and BDZ levels in HE resulted negative. Conclusion: Clinical evidence is provided in cirrhotic patients that ammonia and endogenous BDZ levels do not correlate with each other in the outcome of HE.

2001 - Effect of epicatechin-3-gallate (ECG) and epigallocatechin-3-gallate (EGCG) on cel proliferation, role of peripheral benzodiazepine receptor (PBR) [Abstract in Atti di Convegno]
Corsi, Lorenzo; R., Guagliumi; Avallone, Rossella; Baraldi, Mario
abstract

The polyphenolic compounds showed cancer chemopreventive effect in animal tumor model. Peripheral benzodiazepine receptor i involved in cell proliferation and malignancy. We investigated and compared the effect og epigallocatechin-3-gallate and epicatechin-3-gallate on the proliferation of mouse hepatocellular carcinoma cel line.

2001 - Natural Modulators of Anxiety and Sleep Disturbances in Food [Relazione in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; G., Lugli; Baraldi, Mario
abstract

HPLC analysis of foos extracts revealed the presence of several fractions that exhibited diaplacing activity on benzodiazepine-binding assay to rat cerebellar membranes.

2001 - Presence of benzodiazepine-like compound molecules in food and their implication in the nutrition of cirrhotic patients [Articolo su rivista]
Avallone, Rossella; Corsi, Lorenzo; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

Benzodiazepine (BDZ)-like compounds, present in trace amounts in normal subjects increase in the blood of liver cirrhotic patients. The origin of these compounds is still unknown but they are present in medicinal plants and foods. Herein we report the detection of BDZ-like molecules in fruits, vegetables, cereals, meat, milk and cheeses and in different cultivars of potatoes, tomatoes and carrots. The extracted food was separated by HPLC purification and the collected fractions were tested by radioreceptor binding assay in order to evaluate their ability to selectively bind the central benzodiazepine receptors. The mean value was 14.80 ng of diazepam equivalent (DE)/g in fruits, 4.34 ng DE/g in vegetables, 6.35 in cereals and 4.09 in meat. BDZ-like compounds are poorly present in cheeses and completely absent in olive and seeds oil. From these findings it is possible to select food with low amount of BDZ-like molecules useful for cirrhotic diet in order to prevent hepatic encephalopathy.

2001 - Rybozyme-mediated reduction of the GABAA a1 subunit. [Articolo su rivista]
Subramanian, J; Corsi, Lorenzo; Vicini, S; Whiting, ; P. J., AND NEALE JH
abstract

As an approach to understanding the role of the α1 subunit of the GABAA receptor, ribozymes were designed to reduce expression of this subunit protein by hydrolysis of α1 subunit message and antisense inactivation. The ribozyme cleavage sites were selected through homology comparison of all known murine GABAA receptor subunits at the amino acid and nucleotide sequence level. Two ribozymes were designed and synthesized: one against the extracellular domain and the other against the cytoplasmic domain. These ribozymes were cloned in a mammalian expression plasmid, pZeoSV2 (+). Cleavage of both extracellular and cytoplasmic domain transcripts by the respective ribozymes was observed when each ribozyme was tested against in vitro transcribed mRNA. The stable cell line, 122, expressing recombinant human GABAA α1, β2 and γ2S subunits of receptor was stably transfected with the cytoplasmic domain ribozyme (cy) alone and with both the cytoplasmic (cy) and extracellular domain (ex) ribozyme expression plasmids. Northern analysis showed a 55–60% reduction of α1 mRNA in clones of cells transfected with either the single ribozyme (Cy) or with both ribozymes (EC). The α1 protein level was reduced 75% in a stable Cy clone and more than 90% in a stable EC clone when compared with α1 expression in 122 cells and the vector transfected (Zeo) cells. Electrophysiological analysis revealed that the GABAA receptor properties were very similar in 122 cells and in stable clones in which the subunit protein expression had been greatly reduced. No significant difference was detected in the potentiation of the receptor response by either bretazenil or zolpidem. These data demonstrate the efficacy of the ribozyme approach in dramatically reducing GABAA subunit protein levels in transfected cells and identify those elements that will be important to the application of similar ribozymes to knock-down transmitter receptor subunit proteins under inducible promoters in transgenic mice.

2000 - Benzodiazepine esogene ed endogene: iplicazioni in taluni stati patologici [Capitolo/Saggio]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; Zeneroli, Maria Luisa
abstract

Le benzodiazepine che si trovano nel plasma dell'uomo potrebbero derivare dagli alimenti, dall'azione della flora batterica intestinale e probabilmente da una sintesi endogena ancora sconosciuta.

2000 - Endogenous and exogenous benzodiazepines [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; I., Venturini; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

It is clear that there is a level of benzodiazepines in the serum healthy man free of benzodiazepines medication and that alteration of these levels could influence the state of vigilance and anxiety.

2000 - Endogenous and exogenous benzodiazepines [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; I., Venturini; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

Herein we report the detection of benzodiazepine-like molecules in food suc as carrot, maize, soy-bean, peam wheat, salad and different cultivars of potatoes and tomatoes. The extracted food exhibit after HPLC separation a series of substances able to inhibit the binding of [3H]RO 15-1788 to rat cerebellar membranes hence indicating a benzodiazepine-like activity.

2000 - Endogenous benzodiazepines [Articolo su rivista]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; I., Venturini; C., Baraldi; Zeneroli, Maria Luisa
abstract

The existence of endogenous benzodiazepines such as diazepam and nordiazepam has been provided in human blood and brains as well as in medicinal plants and foods. It must be stressed, however, that in plasma and brain tissue there are also other benzodiazepine-like compounds termed 'endozepines' which are not halogenated. A synthetic pathway for the production of benzodiazepine-like compounds and endozepines has not yet been found, hence it may be surmised that these compounds could be of exogenous source. Changes in the level of endogenous circulating benzodiazepines due to food or drug ingestion could be responsible for pathological conditions. Clinical experiments were designed in order to study the levels of the endogenous benzodiazepines in vegetables and in the blood of control subjects and of cirrhotic patients. These patients accumulate benzodiazepines because of decreased liver metabolization capacity and impaired renal secretion, reaching plasma concentrations similar to those recorded in commercial benzodiazepine consumers.

2000 - Endogenous benzodiazepines: from physiology to patology [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; I., Venturini; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

Endogenous benzodiazepines were measured by the radioligand binding technique after high performance liquidchromatography (HPLC) purification. The presence of diazepam andN-desmethyldiazepam was assayed by HPLC-electrospray tandem mass spectrometry. Diazepam binding inhibitor was studied in serum by radioimmunoassay. Results—Endogenous benzodiazepineswere below the limit of detection in 7% of patients with encephalopathy. When detectable, their levels were at least comparable with those of benzodiazepine consumers and correlated with the liver dysfunction but not the stage of encephalopathy.Serum levels of diazepam binding inhibitor tended to decrease when endogenous benzodiazepines levels increased. Conclusions—Endogenous benzodiazepines may accumulate in patients withliver cirrhosis during the course of the disease, and the phenomenon appears to be independent of the presence or absenceof encephalopathy.

2000 - Le benzodiazepine dalla molecola alla pratica clinica [Monografia/Trattato scientifico]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; Baraldi, Claudia; Zeneroli, Maria Luisa
abstract

La ricerca di molecola endogene ad attività benzodiazepino-simile ha portato alla scoperta di molecole sia negli alimenti che nell'uomo.

2000 - Le benzodiazepine naturali [Capitolo/Saggio]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; Baraldi, Claudia; Ml, Zeneroli
abstract

Sull’origine delle benzodiazepine naturali trovate nei tessuti e nel sangue di mammiferi sono state formulate due ipotesi: a) origine esogena; b) origine endogena. Entrambe sono state prese in considerazione.Per quanto riguarda l’origine esogena, un nostro studio sistematico sul contenuto di sostanze benzodiazepino-simili negli alimenti (verdure, cereali, frutta, carne, latte, formaggio e condimenti) utilizzati nella dieta mediterranea, ha messo in evidenza una differenza significativa nel contenuto di tali composti fra le varietà dei vegetali diverse sia per genotipo sia per fenotipo [2]. La nozione che le benzodiazepine naturali sono presenti in molti alimenti ha suggerito la possibilità di interferire sullo stato di vigilanza dell’uomo, attraverso la somministrazione di diete particolari capaci di modificarne l’omeostasi nel sangue. E’ possibile pertanto mettere a punto diete a basso contenuto di tali sostanze nei pazienti a rischio di sviluppare disturbi neurologici come nel caso della cirrosi epatica o utilizzare diete ricche di benzodiazepine naturali quali il latte nei soggetti ansiosi o con disturbi del sonno.Per quanto riguarda l’origine endogena, pazienti affetti da cirrosi epatica di media gravità sono stati in primo luogo sottoposti ad una dieta in modo da controllare l’apporto benzodiazepinico alimentare [3]. Dopo 3 giorni di dieta essi sono stati trattati per via orale con rifaximina (800 mg/dì per os per 7 giorni), antibiotico non assorbibile capace di modificare la flora batterica intestinale. L’azione antibatterica della rifaximina ha prodotto in questo breve periodo di terapia una riduzione media del 40 % dei livelli di sostanze benzodiazepino-simili nel sangue. Questa osservazione sembra indicare che la flora batterica intestinale sia capace di influenzare i livelli di sostanze benzodiazepino-simili nel sangue e suggerisce la possibilità di utilizzare nella dieta probiotici e prebiotici e relative associazioni nel tentativo di aumentare la flora batterica intestinale utile alla produzione di benzodiazepine endogene capaci di prevenire eccessive lesioni psichiche e disturbi del sonno.

2000 - Presence of benzodiazepines-like molecules in food and theri implication in the nutrition of cirrhotic patients. [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; I., Ventu; Farina, Franco; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

The data presented indicate that it is possible to select vegetables with low amount of benzodiazepines-like molecules which could be more appropriate for cirrhotic diet in order to avoid the occurence of episodes of encephalopathy in patients with severe liver disease.

1999 - Endogenous benzodiazepines [Abstract in Atti di Convegno]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; I., Venturini; Zeneroli, Maria Luisa
abstract

The presence of specific receptors for benzodiazepines (BDZ) suggested that these recognition sites could subserve a physiological role in the regulation of anxiety and sleep only if stimulated by endoigenous ligands. A polypeptide nmed DBI could be one putative endogenous ligands with diazepma binding inhibitory properties and with anxiogenic activity. Clinical experiments were designed in order to study the levels of the endogenous BDZs and in particular the relationship between BDZs and DBI in control.

1999 - Increased expression of peripheral benzodiazepine receptors and diazepam binding inhibitor in human tumors sited in the liver [Articolo su rivista]
I., Venturini; H., Alho; I., Podkletnova; Corsi, Lorenzo; E., Rybnikova; R., Pellicci; Baraldi, Mario; M., Pelto Huikko; P., Helen; Zeneroli, Maria Luisa
abstract

The peripheral benzodiazepine receptor system triggers intracellular metabolic events and has been associated with cell proliferation. Its endogenous ligand, the diazepam binding inhibitor, contributes to steroidogenesis by promoting cholesterol delivery to the inner mitochondrial membrane. The present study was undertaken to verify whether this system is altered in tumors sited in the liver. Peripheral benzodiazepine receptors and diazepam binding inhibitor were studied using immunocytochemistry and in situ hybridization in 9 human tumors sited in the liver, in liver hyperplasia, cirrhotic nodular regeneration, intestinal adenocarcinoma and in surrounding non-tumoral tissue. Immunocytochemical staining and in situ hybridization demonstrated that peripheral benzodiazepine receptors and diazepam binding inhibitor were more prominently expressed in neoplastic cells than in non-tumoral tissue. They were present in the same cells, suggesting that diazepam binding inhibitor may act in an intracrine manner in these cells. Higher peripheral benzodiazepine receptors and diazepam binding inhibitor expression in tumor cells suggest an implication of this system in the metabolism of neoplastic cells. Furthermore the evaluation of peripheral benzodiazepine receptor and diazepam binding inhibitor expression might be useful in evaluating malignancy and in diagnostic approaches of tumors in liver tissue.

1998 - A possible mechanism of resistance to death during cell proliferation in hepatocellular carcinoma: the mitochondrial benzodiazepine receptor system. [Abstract in Atti di Convegno]
I., Venturini; Zeneroli, Maria Luisa; Corsi, Lorenzo; R., Pellicci; G., Ardizzone; A., Arrigo; Farina, Franco; C., Ferrarese; N., Pecora; M., Frigo; Avallone, Rossella; Baraldi, Mario
abstract

The aims of the present research w to detremine the role of the mitocondrial benzodiazepine receptors (MBRs) in hepatocellular carcinoma (HCC). The densities of MBRs and the level of DBI in PPIK were tested in tumoral and non tumoral liver tissue of patients with HCC.

1998 - Ammonia and 1,4-benzodiazepines in encephalopathic patients with fulminatn hepatic failure [Abstract in Atti di Convegno]
Zeneroli, Maria Luisa; I., Venturini; Corsi, Lorenzo; Avallone, Rossella; Farina, Franco; G., Ardizzone; M., Centenaro; A., Arrigo; L., Miglioli; P., Schreier; M., Kleinschnitz; Baraldi, Mario
abstract

Recently it has been surmised that both ammonia and benzodiazepine-like compounds (BDZs) may sinergistically contribute to hepatic encephalopathy (HE). In order to clarify this hypothesis, the levels of venous ammonia and BDZs were assayed in patients wuth HE due to fulminant hepatic failure.

1998 - Ammonia and 1,4-benzodiazepines in patients with fulminat hepatic failure [Abstract in Atti di Convegno]
Zeneroli, Maria Luisa; I., Venturini; Corsi, Lorenzo; Avallone, Rossella; Farina, Franco; G., Ardizzone; M., Centenaro; A., Arrigo; A., Caronna; P., Schreier; M., Kleinschnitz; Baraldi, Mario
abstract

The inconsistent presence of benzodiazepines in hepatic encephalopathy (HE) due to fulminat hepatic failure and the presence at the awakening from coma, indicate that HE i note strictly depoendent on the levels of these compounds.

1998 - Benzodiazepine-like compound in foods and plants [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; Baraldi, Mario
abstract

Extracts of potatoes and tomatoes exhibited after HPLC separation a series of substances able to inhibit the binding of [3H]RO 15-1788 to rat cerebellar membranes. The level ranged from 0.03 ng diazepam equivalent (DE)/g to 3.56 ng DE/g inthe potatoes and from 0.01 to 1.55 DE/g in the tomatoes depending from the selected cultivars.

1998 - Benzodiazepine-like compounds in plasma of patients with fulminant hepatic failure [Articolo su rivista]
Zeneroli, Maria Luisa; I., Venturinii; Corsi, Lorenzo; Avallone, Rossella; Farina, Franco; G., Ardizzone; M., Centanaro; A., Arrigo; P., Schreier; M., Kleinschnitz; Baraldi, Mario
abstract

Background: Benzodiazepine-like compounds have been implicated in the pathogenesis of encephalopathy after fulminant hepatic failure. Methods: The levels and the nature of benzodiazepine-like compounds were determined in six cases of fulminant hepatic failure during the course of the disease. Blood samples were collected on admission and a few days later, when the neurologic status had improved in five cases and immediately before death in one case. The compounds were measured in sera with a binding technique after high-performance liquid chromatography purification and analyzed with mass spectrometry;, Results: Their levels were highly variable in those with severe encephalopathy and were still Increased on awakening in some cases. Diazepam and N-desmethyldiazepam were inconsistently present. Conclusions: The inconsistent presence of benzodiazepine-like compounds in encephalopathy after fulminant hepatic failure and their persistence, in some cases, at high levels on awakening from coma seem to Indicate that the encephalopathy is not strictly dependent on the levels of these compounds.

1998 - Chronic dizolcipine (MK 801) reversibly delays GabaA receptor maturation in cerebellar granule neurons in vitro. [Articolo su rivista]
Wang, X. H.; Zhu, W. J.; Corsi, L.; Ikonomovic, S.; Paljung, W. R.; Vicini, S.; Grayson, D. R.
abstract

We investigated the effect of chronically blocking NMDA receptor stimulation to examine changes in GABAA receptor expression and pharmacology in cerebellar granule cells at different stages of maturation. We have previously shown that NMDA-selective glutamate receptor stimulation alters GABAA receptor pharmacology in cerebellar granule neurons in vitro by altering the levels of selective subunits. When NMDA receptor stimulation is blocked with MK-801 during the first week in vitro, a decrease in the α1, γ2S, and γ2L receptor subunit mRNAs occurred. When present only during the second week, changes were limited to the α1 and γ2L mRNAs. Finally, if MK-801 was present during the first week and removed during the second week, these changes reversed. Whole-cell voltage-clamp recordings showed that treatment with MK-801 during either the first or second week increased the EC50 of the receptors for GABA and attenuated the potentiation mediated by flunitrazepam. Last, these properties were reversed if MK-801 was removed after the first week in vitro. Our results suggest that MK-801 reversibly inhibits GABAA receptor maturation by modulating receptor subunit expression and that the altered pharmacological responses appear to be dominated by changes in the levels of allosteric modulation mediated by the γ2 receptor subunit.

1998 - Developmental changes in localization of NMDA receptor subunits in primary cultures of cortical neurons [Articolo su rivista]
LI J., H; Y. H., Wang; B. B., Wolfe; K. E., Krueger; Corsi, Lorenzo; G., Stocca; S., Vicini
abstract

Immunoblot analysis, using antibodies against distinct N‐methyl‐d‐aspartic acid (NMDA) receptor subunits, illustrated that the NR2A and NR2B subunit proteins have developmental profiles in cultured cortical neurons similar to those seen in vivo. NR1 and NR2B subunits display high levels of expression within the first week. In contrast, the NR2A subunit is barely detectable at 7 days in vitro (DIV) and then gradually increased to mature levels at DIV21. Immunocytochemical analysis indicated that NMDA receptor subunits cluster in the dendrites and soma of cortical neurons. Clusters of NR1 and NR2B subunits were observed as early as DIV3, while NR2A clusters were rarely observed before DIV10. At DIV18, NR2B clusters partially co‐localize with those of NR2A subunits, but NR2B clusters always co‐localize with those of NR1 subunits. Synapse formation, as indicated by the presence of presynaptic synaptophysin staining, was observed as early as 48–72 h after plating. However, in several neurons at ages less than DIV5 where synapses were scarce, NR2B and NR1 clusters were abundant. Furthermore, while NR2B subunit clusters were seen both at synaptic and extrasynaptic sites, NR2A clusters occurred almost exclusively in front of synaptophysin‐labelled boutons. This result was supported by electrophysiological recording of NMDA‐mediated synaptic activity [NMDA‐excitatory postsynaptic currents (EPSCs)] in developing neurons. At DIV6, but not at DIV12, CP101, 606, a NR1/NR2B receptor antagonist, antagonized spontaneously occurring NMDA‐EPSCs. Our data indicate that excitatory synapse formation occurs when NMDA receptors comprise NR1 and NR2B subunits, and that NR2A subunits cluster preferentially at synaptic sites

1998 - Diazepam binding inhibitor and total cholesterol plasma levels in cirrhosis and hepatocellular carcinoma [Articolo su rivista]
Venturini, ; Zeneroli, Maria Luisa; Corsi, Lorenzo; Baraldi, Claudia; C., Ferrarese; N., Pecora; M., Frigo; H., Alho; Farina, Franco; Baraldi, Mario
abstract

Cholesterol is used by cells for biosynthetic processes and for steroid synthesis. Although the role of cholesterol in tumorigenesis is not clear it is known that steroids are important factors in human carcinogenesis. A polypeptide, diazepam binding inhibitor (DBI), which is an endogenous ligand for peripheral benzodiazepine receptors enhances steroidigenesis by promoting cholesterol delivery to the inner mitochondrial membrane which represents the rate-limiting step of steroid biosynthesis. We have assayed the total cholesterol (TC) and the DBI plasma concentrations in patients with liver cirrhosis complicated by hepatocellular carcinoma (HCC) in comparison with those of uncomplicated liver cirrhosis. TC and DBI levels have been studied in 73 cirrhotic patients and in 23 patients with HCC. Both TC and DBI levels were higher in HCC patients when compared with age, sex and Child–Pugh class matched cirrhotic controls. The values (mean±S.D.) in patients in Child–Pugh class B and C with and without HCC were respectively 128±30 mg/dl vs. 106±27 mg/dl (P<0.01) and 2.05±0.78 pmol/ml vs. 0.78±0.84 pmol/ml (P<0.0001). The data may be the result of the metabolic influence of tumors that enhances steroid biosynthesis during tumor proliferation.

1998 - Endogenous benzodiazepine-like compounds and diazepam binding inhibitor in serum of patients with liver cirrhosis with and without overt encephalopathy [Articolo su rivista]
Avallone, Rossella; Zeneroli, Maria Luisa; I., Venturini; Corsi, Lorenzo; P., Schreier; M., Kleinschnitz; C., Ferrarese; Farina, Franco; Baraldi, Claudia; N., Pecora; M., Frigo; Baraldi, Mario
abstract

Background/Aim—Despite some controversy, it has been suggested that endogenous benzodiazepine plays a role in the pathogenesis of hepatic encephalopathy. The aim of the present study was to evaluatethe concentrations of endogenous benzodiazepines and the peptide, diazepam binding inhibitor, in the blood of patients with liver cirrhosis with and without overt encephalopathy, and to compare theselevels with those of consumers of commercial benzodiazepines.Subjects—Normal subjects (90), benzodiazepine consumers (14), and cirrhotic patients (113) were studied. Methods—Endogenous benzodiazepines were measured by the radioligand bindingtechnique after high performance liquid chromatography (HPLC) purification. The presence of diazepam and N-desmethyldiazepam was assayed by HPLC-electrospray tandem mass spectrometry.Diazepam binding inhibitor was studied in serum by radioimmunoassay. Results—Endogenous benzodiazepineswere below the limit of detection in 7% of patients with encephalopathy. When detectable, their levels were at least comparable with those of benzodiazepine consumers and correlated with the liver dysfunction but not the stage of encephalopathy.Serum levels of diazepam binding inhibitor tended to decrease when endogenous benzodiazepines levels increased. Conclusions—Endogenous benzodiazepines may accumulate in patients withliver cirrhosis during the course of the disease, and the phenomenon appears to be independent of the presence or absence of encephalopathy.

1998 - Endogenous benzodiazepine-like compounds in serum of normal subjects and in liver cirrhosis patients. [Abstract in Atti di Convegno]
I., Venturini; Avallone, Rossella; Corsi, Lorenzo; Zeneroli, Maria Luisa; P., Schreier; M., Kleinschnitz; Farina, Franco; Baraldi, Mario
abstract

The concentration of benzodiazepines (BDZs) in serum from normal subject resulted under the limit of detection (lower tha 2 nmol DE/l) in 46 cases (51%). In the remaining 44 normal subjects the total amount binding ranged between 6 and 20 nmol DE/l. In the liver cirrhosis BDZs were under the detection limit in 7% of patients withencephalopathy. When detectable, tehi levels were at least comparable with those of benzodiazepines consumers and correlated with the liver dysfuntion but not with the stages of encephlopathy.

1998 - Endogenous benzodiazepines sources and role in liver cirrhosis and in fulminat hepatic failure [Abstract in Atti di Convegno]
Baraldi, Mario; Zeneroli, Maria Luisa; I., Venturini; Corsi, Lorenzo; Avallone, Rossella; Farina, Franco
abstract

The source and the role on endogenous benzodiazepines (BZDs)in the induction of hepatic encephalopathy (HE) are still unknown. BDZs are measured by radioligand binding thecnique after HPLC purificationin normnal subjects (n.90), benzodiazepine consumers (BZC) (n.14), cirrhotic patients (LC) (n.133), fluminat hepatic failure (FHF) (n.6).

1998 - Evidence that plasma levels of natural benzodiazepines are regulated by intestinal bacterial flora. [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; I., Venturini; Farina, Franco; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

The source od natural benzodiazepines is still unknown but they have been described in medicinal plants, vegetables and milk. In order to establish the content of BDZs in our diet, we measured natural BDZs by HPLC followed by radioligand binding assay in potatoes and tomatoes.

1998 - Food and bacterial flora are source of benzodiazepine-like compounds [Abstract in Atti di Convegno]
Zeneroli, Maria Luisa; I., Venturini; S., Stefanelli; Farina, Franco; A., Ferrieri; Corsi, Lorenzo; Avallone, Rossella; R., Cosenza; Baraldi, Mario
abstract

The content of natural benzodiazepines (BDZs) in diet was assayed by HPLC followed by radioligand binding technique in potatoes and tomatoes. We assayed the levels of BDZs in the plasma of 10 liver cirrhosis patients in child B class free of commercial benzodiazepines medication and under a constant diet. The patients were treated with rifaximin (800 mg/die x 7 days) in order to modify the bacterial flora.

1998 - Lanthanum - mediated modification of GABAA receptor deactivation, desensitization and inhibitory synaptic currents [Articolo su rivista]
ZHU W., J; J. F., Wang; Corsi, Lorenzo; S., Vicini
abstract

We investigated La3+ effects on recombinant and native γ-aminobutyric acid A (GABAA) receptors using rapid agonist applications and on inhibitory synaptic currents (IPSCs) in granule and stellate neurons of rat cerebellar slices.Rapid desensitization of currents elicited by 200 ms pulses of 1 mM GABA to small lifted cells transfected with α1β3γ2 cDNAs was greatly decreased by the coapplication of 100 μm LaCl3.GABA responses were unaffected when coapplication lasted only 2 ms. In contrast, with LaCl3 pre-perfusion, a significant slowing of deactivation in response to 2 ms applications was observed. LaCl3 pre-perfusion also prolonged the duration of responses to 20 mM taurine.Outside-out patches excised from cells transfected with α1β3γ2 subunit cDNAs were briefly exposed to a saturating concentration of GABA, eliciting a transient activation of single channel currents with a main conductance of 30 pS. Opening and burst durations increased by pre-equilibration of patches with LaCl3.LaCl3 depressed the peak amplitude without affecting the slow deactivation and desensitization of GABA responses in cells transfected with α6β3γ2 and α6β3δ cDNAs. No significant difference in La3+ modulation of GABA-gated currents was observed between α1β3γ2 and α1β3δ receptors.The effects of LaCl3 on deactivation and desensitization of GABA responses observed in nucleated patches excised from rat cerebellar granule and stellate neurons were comparable to those in the cells transfected with α1β3γ2 cDNAs. In addition, La3+ clearly prolonged the spontaneous IPSC time course without changing the amplitude.Our results indicate that La3+ has a dual action on GABA-gated currents: it decreases desensitization and increases channel opening duration. These actions depend on receptor subunit composition and contribute to the prolongation of IPSCs.

1998 - Mitochondrial benzodiazepine receptor system in hepatocellular carcinoma: a possible mechanims of resistance to death during cell proliferation. [Abstract in Atti di Convegno]
I., Venturini; Zeneroli, Maria Luisa; Corsi, Lorenzo; R., Pellicci; G., Ardizzone; A., Arrigo; Farina, Franco; C., Ferrarese; N., Pecora; M., Frigo; Avallone, Rossella; Baraldi, Mario
abstract

These data demonstrated that in HCC patients there is an increased availability of the substrate for steroid synthesis, associated with an increase functional status of MBRs in HCC tissue. This suggest that in HCC tissue there is an incresed steroidogenesis and a resistance of tumoral against radical damage.

1998 - Peripheral benzodiazepine receptors in liver tumors and presence of thei ligands in vegetbale and officinal plants. [Abstract in Atti di Convegno]
Corsi, Lorenzo; Avallone, Rossella; I., Venturini; Zeneroli, Maria Luisa; H., Alho; Baraldi, Mario
abstract

in this study we provided evidence that during degenerative liver disease as hepatocellular carcinoma (HCC) the peripheral benzodiazepine receptor (PBR) binding sites increse drammatically. Extract of tomatoes, mentha piperita, lavadula officinalis, citrus aurantium and melissa officinalis after HPLC separation gave rise to a series of compounds able to inhibit the binding of [3H]PK 11195, a specific binding for PBRs to rat adrenal glands.

1998 - Presence o fbenzodiazepine-like molecules in vegetables and thei implication in the nutrition of cirrhotic pateints. [Abstract in Atti di Convegno]
Avallone, Rossella; Corsi, Lorenzo; I., Venturini; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

The findings indicate that it is possible to select vgetables with low amount of benzodiazepine-like molecules which appear to be more appropriate in order to avoid the occurence of episodes of encephalopathy in patients with severe liver disease.

1998 - Sostanze ad attività benzodiazepino-simile in alimenti e droghe vegetali [Abstract in Atti di Convegno]
Baraldi, Mario; Avallone, Rossella; Corsi, Lorenzo; I., Venturini; Zanoli, Paola; M. L., Zeneroli
abstract

Per chierire l'otigine delle benzodiazepine trovate nei tessuti, sono stati analizzati diversi alimenti fra cui patata e pomodoro. Mediante analisi HPLC sono state separate da estratti di patata e pomodoro diverse sostanze capaci di inibire il legame del [3H]RO 15-1788 alle membrane cerebellari e del [3H]PK 11195 alle membrane surrenali di ratto. La quantità di queste sostanze varia da 0,03 ng/g diazepam equivalente (DE) a 3,56 ng/g di patata e da 0,01 a 1,55 ng/g di pomodoro a seocnda del cultivar analizzato.

1998 - Sources od benzodiazepine-like compounds: food and bacterial flora [Abstract in Atti di Convegno]
Avallone, Rossella; I., Venturini; Corsi, Lorenzo; Farina, Franco; DI BELLA, Maria; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

The observation indicates that intestinal bacterila flora is involved in the production of these compounds. In conclusion, even if cannot excludean endogenous synthesis, these observations seems to prove that the sources of benzodiazepine-like compounds in human could be the result of a combination of the food ingestion and bacterial flora production.

1998 - Upregulation of NR2B subunit of NMDA receptors in cerebellar granule neurons by CaM kinase inhibitor KN93 [Articolo su rivista]
Corsi, Lorenzo; J. H., Li; K. E., Krueger; Y. H., Wang; B. B. WOLFE AND S., Vicini
abstract

Recordings of NMDA-activated currents from cerebellar granule neurons in culture revealed a developmental increase in current density accompanied by a slight decrease of the half-maximal effective concentration. At the same time, a decrease of NMDA receptors comprising NR2B subunits was demonstrated by the reduction in the antagonism of NMDA currents by ifenprodil. Ifenprodil antagonism increased after treatment for 24 h with KN93- and KN62-selective inhibitors of the Ca2+/calmodulin-dependent protein kinases (CaM kinases), indicating a selective increase of receptor containing NR2B subunit. This increase was observed at all ages tested: 4 days in vitro (DIV4), DIV6, and DIV13. Western blot analysis with specific NMDA receptor antibodies performed at DIV6 confirmed the electrophysiological data. At this age, the negative control KN92 was ineffective. The increasing ifenprodil antagonism after KN93 treatment was proportionally greater in cells at DIV13 than at DIV4. Treatment with NMDA (100 µM) of cerebellar cultures for 24 h produced a decrease in the NMDA-induced current density by almost 50% at all ages tested. Ifenprodil antagonism, however, was unchanged. We propose that the expression of NR2B subunits in cerebellar granule cells is selectively stimulated by the inhibition of CaM kinases.

1998 - Up-regulation of peripheral benzodiazepine receptors system in hepatocellular carcinoma. [Articolo su rivista]
Venturini, I; Zeneroli, Maria Luisa; Corsi, Lorenzo; Avallone, Rossella; Farina, Franco; Alho, H; Baraldi, C; Pecora, N; Frigo, M; Ardizzone, G; Arrigo, A; Pellicci, R; Baraldi, Mario
abstract

Increased number of peripheral benzodiazepine receptors (PBRs) have been found in some tumors outside the liver. The present study was to verify whether the PBR system is altered in hepatocellular carcinoma (HCC). The levels of endogenous benzodiazepine-like compounds (BZDs). measured by radioreceptor binding technique after HPLC purification and the endogenous ligand for PBRs. termed diazepam binding inhibitor (DBI). measured by radioimmunoassay utilizing a specific antibody for human DBI, were studied in the blood of 15 normal subjects, 12 liver cirrhosis and 10 patients with HCC. The levels of BZDs in serum were increased hundred fold in liver cirrhosis patients and slightly elevated in HCC patients. DBI was found to be increased in HCC patients. The binding recognition sites for PBRs (Bmax) were increased 4 to 7 fold in HCC tissue in comparison with that found in non-tumoral liver tissue (NTLT). On the contrary the concentrations of DBI were found to be significantly decreased in HCC tissue in comparison with the respective NTLT. These results seem to suggest an implication of PBRs and of their putative endogenous ligands in the metabolism of these neoplastic cells and possibly in their proliferation. The up-regulation of PBRs found in HCC tissue seems to indicate an increased functional activity of these receptors and opens up the possibility of new pharmacological and diagnostic approaches while the changes in the circulating endogenous ligands for the above receptors might be envisaged as early markers of tumorigenesis in liver cirrhosis.

1997 - Benzodiazepine-like compounds in plasma of patients with fulminant hepatic failure. [Abstract in Atti di Convegno]
I., Venturini; R., Cosenza; L., Miglioli; Farina, Franco; Corsi, Lorenzo; Avallone, Rossella; R., Amdedei; G., Ardizzone; E., Minelli; S., Stefanelli; Baraldi, Mario; P., Schreier; M., Kleinschnitz; Zeneroli, Maria Luisa
abstract

In conclusion the inconsistent presence od benzodizepines in hepatic encephalopathy (HE) due to fulminat hepatic failure and the persistence at high levels at the awakening from coma indicate that HE is not strickly dependent on the level of these compounds

1997 - Endogenous benzodiazepine-like compounds in a patients with fulminatn hepatic failure: influence of a decreased renal excretion [Abstract in Atti di Convegno]
R., Cosenza; I., Venturini; L., Miglioli; Farina, Franco; Corsi, Lorenzo; Avallone, Rossella; R., Amedei; G., Ardizzone; A., Arrigo; E., Minelli; S., Stefanelli; Baraldi, Mario; Zeneroli, Maria Luisa
abstract

These observations suggest that encephalopathy due to fulminant hepatic faliure (FHF) may precipitate in the presence of low level of endogenous benzodiazepine and that these compounds may accumulate in the blood during the course of FHF without inducing encephalopath. One of the mechanisms of the increse of these compounds in the blood seems to be decreased urinary excretion due to diuresis.

1997 - N-acetylaspartylglutamate stimulates metabotropic glutamate receptor 3 to regulate expression of the GABA(A) alpha6 subunit in cerebellar granule cells [Articolo su rivista]
Ghose, S; B., Wroblewska; Corsi, Lorenzo; D. R., Grayson; A. L., DE BLAS; S., Vicini; JOSEPH H., Neale
abstract

We have shown that the vertebrate neuropeptide N-acetylaspartylglutamate (NAAG) meets the criteria for a neurotransmitter, including function as a selective metabotropic glutamate receptor (mGluR) 3 agonist. Short-term treatment of cerebellar granule cells with NAAG (30 microM) results in the transient increase in content of GABA(A) alpha6 subunit mRNA. Using quantitative PCR, this increase was determined to be up to 170% of control values. Similar effects are seen following treatment with trans-1-aminocyclopentane-1,3-dicarboxylate and glutamate and are blocked by the mGluR antagonists (2S,3S,4S)-2-methyl-2-(carboxycyclopropyl) glycine and (2S)-alpha-ethylglutamic acid. The effect is pertussis toxin-sensitive. The increase in alpha6 subunit mRNA level can be simulated by activation of other receptors negatively linked to adenylate cyclase activity, such as adenosine A1, alpha2-adrenergic, muscarinic, and GABA(B) receptors. Forskolin stimulation of cyclic AMP (cAMP) levels abolished the effect of NAAG. The change in alpha6 levels induced by 30 microM NAAG can be inhibited in a dose-dependent manner by simultaneous application of increasing doses of the beta-adrenergic receptor agonist isoproterenol. The increase in alpha6 mRNA content is followed by a fourfold increase in alpha6 protein level 6 h posttreatment. Under voltage-clamped conditions, NAAG-treated granule cells demonstrate an increase in the furosemide-induced inhibition of GABA-gated currents in a concentration-dependent manner, indicating an increase in functional alpha6-containing GABA(A) receptors. These data support the hypothesis that NAAG, acting through mGluR3, regulates expression of the GABA(A) alpha6 subunit via a cAMP-mediated pathway and that cAMP-coupled receptors for other neurotransmitters may similarly influence GABA(A) receptor subunit composition

1997 - Peripheral benzodiazepine receptors in hepatocellular carcinoma. [Capitolo/Saggio]
M. L., Zeneroli; R., Pellicci; I., Venturini; G., Ardizzone; A., Arrigo; Corsi, Lorenzo; Avallone, Rossella; Baraldi, Mario
abstract

Peripheral benzodiazepine receptor system (PBRs) triggers intracellular metabolic events and has been associated with cell proliferation. Its endogenous ligand, the diazepam binding inhibitor (DBI), contributes to steroidogenesis by promoting cholesterol delivery to the inner mitochondrial membrane. Since an increased number of PBRs have been found in several tumour tissues and cell lines, the present study was performed in order to verify if the expression of PBRs and DBI were altered in liver tumours. Using immunocytochemestry and in situ hybridisation we studied PBRs end DBI expression in 6 human tumours sited in the liver, in 1 liver hyperplasia, in 1 cirrhotic nodular regeneration, in 1 intestinal adenocarcionma and in correspondent surrounding non tumoral liver tissue (NTLT). Immunocytochemical study and in situ hybridisation revealed that PBRs and DBI were prominently expressed in neoplastic tissues than in NTLT and that they were colocalised in the same cells.

1997 - Sostanze ad attività benzodiazepino-simile in alimenti e in droghe vegetali: implicazioni fisiologiche e patologiche [Articolo su rivista]
Avallone, Rossella; Corsi, Lorenzo; Zanoli, Paola; DI BELLA, Maria; Monzani, Agar; I., Venturini; Zeneroli, Maria Luisa; Baraldi, Mario
abstract

Nel presente lavoro sono state estratte, purificate cromatograficamente e catratterizzate mediante dosaggi radioisotopici sostanze ad attività benzodiazepino-simile in 32 varietà di papata (Solanum tuberosum L.), 12 varietà di pomodoro (Solanum lycopersium L.) e nella camomilla (Matricaria chamomilla L.).

1996 - Benzodiazepine-like compounds and GABA in flower heads of Matricaria Chamomilla [Relazione in Atti di Convegno]
Avallone, Rossella; Zanoli, Paola; Corsi, Lorenzo; Cannazza, Giuseppe; Baraldi, Mario
abstract

Extracted and purified benzodiazepine like compounds from dried flwer heads of Matricaria chamomilla was investigated through radioligand binding assay on rat cerebellar membrane. Moreover intracerbroventrivular injection of purified active fraction produced a significant decrease of locomotor activity in rats.

1996 - Hepatic Encephalopathy in Liver Transplant Recipients Precipitated By Benzodiazepines Present in Transfused Blood [Articolo su rivista]
Zeneroli, Maria Luisa; I., Venturini; Avallone, Rossella; Farina, Franco; Corsi, Lorenzo; Baraldi, Claudia; G., Ardizzone; M., Centanaro; A., Arrigo; Baraldi, Mario
abstract

The observation that there are episodes of encephalopathy in liver cirrhosis patients after orthotopic liver transplantation, despite a well functioning graft and despite the lack of cerebral complications, prompted us to investigate the potential role of circulating benzodiazepine-like compounds in these episodes. The plasma levels of benzodiazepines were examined in 14 liver cirrhotic patients before and after transplantation. The benzodiazepines in the fluids infused during surgery and in individual bags of blood administered after surgery to 4 of these patients were also assayed. Herein we report that benzodiazepines accumulating in the blood of some transplanted patients appear to derive from blood transfusions utilized during surgery. The analysis of the types of benzodiazepines present in the blood utilized for transfusions suggests the use of commercial benzodiazepines by the donors. These compounds seem to be able to precipitate hepatic encephalopathy in patients with preexisting encephalopathy. Hence we suggest not using benzodiazepine consumers as blood donors, at least for patients with encephalopathy undergoing to liver transplantation.

1996 - Hepatic encephalopathy in patients submitted to liver transplantation precipitated by benzodiazepines present in transfused blood. [Articolo su rivista]
Zeneroli, Maria Luisa; I., Venturini; Avallone, Rossella; Farina, Franco; Corsi, Lorenzo; G., Ardizzone; M., Centanaro; A., Arrigo; Baraldi, Mario
abstract

Abstract: The observation that there are episodes of encephalopathy in liver cirrhosis patients after orthotopic liver transplantation, despite a well functioning graft and despite the lack of cerebral complications, prompted us to investigate the potential role of circulating benzodiazepine-like compounds in these episodes. The plasma levels of benzodiazepines were examined in 14 liver cirrhotic patients before and after transplantation. The benzodiazepines in the fluids infused during surgery and in individual bags of blood administered after surgery to 4 of these patients were also assayed. Herein we report that benzodiazepines accumulating in the blood of some transplanted patients appear to derive from blood transfusions utilized during surgery. The analysis of the types of benzodiazepines present in the blood utilized for transfusions suggests the use of commercial benzodiazepines by the donors. These compounds seem to be able to precipitate hepatic encephalopathy in patients with preexisting encephalopathy. Hence we suggest not using benzodiazepine consumers as blood donors, at least for patients with encephalopathy undergoing to liver transplantation.