Alcoholic cerebellar degeneration in bile-duct ligated rats.
M.Sc., Clinical Pharmacology, Université de Montréal
Direction:
- Dr Christopher Rose
2017 - 2018
Related Publications
Farzaneh Tamnanloo, Xiaoru Chen, Mariana Oliveira, Mélanie Tremblay, Christopher F. Rose.
Background: Hepatic encephalopathy (HE) is a debilitating neurological complication of chronic liver disease. Alcohol is a major etiological factor known to induce liver injury and disease. However, excessive alcohol consumption has been shown to also induce atrophy of the cerebellum and cerebellar degeneration. To date, the role of alcohol in the development of HE remains unclear. Here we examined the effects of constant alcohol consumption on the neurological decline in rats with chronic liver disease induced following bile duct ligation (BDL). Method: 5-week BDL rats and Sham-operated controls (Sham) were used. Starting day 7 after surgery, rats were gavaged twice a day (3 hours apart) with alcohol at a dose of 3g/kg, 5 days per week, for 4 weeks. Motor coordination was assessed using Rotarod every week until week 5. At the end of the model (day 40), anxiety-like behavior was assessed using the open field (OF) and elevated plus maze (EPM). Upon sacrifice, brains were collected, and western blot and immunohistochemical (IHC) analyses were used to investigate the neuronal integrity as well as assess apoptosis and necroptosis pathways in the cerebellum. Results: Alcohol worsened motor coordination performance in weeks 2, 3, 4, and 5 in BDL-alcohol rats (p<0.01 vs respective shams). Anxiety-like behavior significantly increased in BDL-alcohol rats, with an increase in time spent in the closed arms of EPM and a decrease in time spent in the center of the open field (p<0.05 vs respective shams). These impairments were associated with decreased neuronal markers of NeuN and SMI311 (p<0.01 and p<0.05, respectively), increased apoptotic markers of cleaved/pro-Caspase3 and Bax/Bcl2 ratio (p<0.001 and p<0.01 respectively), increased necroptosis markers of pRIP3 and pMLKL (p<0.01 and p<0.001, respectively), decreased total antioxidant capacity (p<0.001) and increased oxidative stress marker of 4-HNE (p<0.05) in the cerebellum of BDL-alcohol rats when compared to respective controls. IHC results confirmed the colocalization of apoptotic marker (cleaved Caspase3) and necroptosis marker (pMLKL) in the granular and Purkinje layer neurons of the cerebellum of BDL-alcohol rats. Conclusion: Constant alcohol consumption exacerbates HE by worsening motor coordination impairment and increasing anxiety in BDL rats. Furthermore, our results show neuronal loss through apoptosis and necroptosis in the cerebellum of BDL-alcohol rats. Additionally, higher levels of oxidative stress marker of 4-HNE and decreased total antioxidant capacity in the cerebellum of BDL-alcohol rats suggest that oxidative stress is the triggering factor of apoptosis and necroptosis pathway leading to neuronal loss/injury. These results demonstrate the adverse effect of constant alcohol consumption on the development of HE and neuronal integrity in chronic liver disease.
Farzaneh Tamnanloo, Xiaoru Chen, Mélanie Tremblay, Christopher F. Rose.
Problématique: L'encéphalopathie hépatique (EH) est un syndrome neuropsychiatrique résultant d'une maladie du foie. Il a été démontré que le microbiote intestinal influence le cerveau et l'association entre l'altération du microbiote et les maladies hépatiques ressort dans de nombreuses études. Objectif : Explorer l'impact de la transplantation de microbiote fécal (FMT) sur le développement de l’EH de rats ayant subi une ligature du canal biliaire (BDL). Méthodologie: Des rats mâles ont été randomisés en trois groupes : SHAM, BDL-VEH et BDL-FMT recevant quotidiennement la FMT provenant des rats SHAM. Après cinq semaines, des tests comportementaux sont effectués pour évaluer la mémoire à court/long terme, l'anxiété et la coordination motrice. Les fèces et plasma ont été collectés pour séquençage bactérien et analyses. Résultats: Les BDL-VEH ont développé une perte de mémoire à court/long terme et une perte de coordination motrice comparée aux rats SHAM. Cependant, les altérations neurologiques sont prévenues dans le groupe BDL-FMT. Une modulation du microbiote a été constatée pour les rats BDL-FMT comparé aux BDL-VEH. Les cytokines (TNF-α et IL-1β) ne varient entre les groupes BDL. L'analyse des acides gras à chaîne courte dans les fèces et le plasma a montré une variation du propionate et du butyrate entre les groupes BDL. Le propionate plasmatique est positivement corrélé aux scores de comportement. Discussion: Nos résultats démontrent que la FMT améliore la mémoire et la coordination motrice chez les rats BDL. La FMT conduit à un nouveau profil spécifique du microbiote avec la présence du propionate comme métabolite à explorer.
Farzaneh Tamnanloo, Xiaoru Chen, Mélanie Tremblay, Christopher F. Rose.
Contexte : L'encéphalopathie hépatique (EH) est une complication neurologique débilitante de la maladie hépatique chronique (MHC). L'alcool est un facteur étiologique majeur connu pour induire des dommages hépatiques, mais son rôle dans le développement de l'EH reste flou. Nous avons examiné les effets de la consommation constante d'alcool sur le déclin neurologique chez les rats atteints de MHC. Méthode : Des rats opérés avec une ligation de voie biliaire (BDL) et des témoins (Sham) après 5 semaines ont été utilisés. Au jour 7, les rats ont été gavés deux fois par jour avec de l'éthanol (3 g/kg, 5 j/sem, pendant 4 sem). La coordination motrice a été évaluée par Rotarod et le comportement anxieux par champ ouvert et labyrinthe surélevé. Nous avons effectué des immunobuvardages afin d'étudier l'intégrité neuronale dans le cervelet. Résultats : L'alcool a aggravé la coordination motrice et augmenté le comportement de type anxieux chez les rats BDL-alcool. Les déficiences étaient associées à une diminution des marqueurs neuronaux de NeuN et SMI311, une augmentation des marqueurs apoptotiques pro-Caspase3/clivée et Bax/Bcl2 et de nécroptose de pRIP3 et pMLKL, une diminution de la capacité antioxydante totale et une augmentation d’un marqueur de stress oxydatif, le 4-HNE, dans le cervelet des rats BDL-alcool par rapport aux témoins respectifs. Conclusion : Les niveaux plus élevés de 4-HNE et la diminution de la capacité antioxydante dans le cervelet des rats BDL-alcool suggèrent que le stress oxydatif est le facteur déclenchant de l'apoptose et de la nécroptose entraînant une perte/lésion neuronale.
Farzaneh Tamnanloo, Xiaoru Chen, Mélanie Tremblay, Christopher F. Rose.
Background: Hepatic encephalopathy (HE) is a debilitating neurological complication of chronic liver disease with alcohol being a common etiological factor. However, excessive alcohol consumption has been shown to impact neurological integrity. To date, the influence of alcohol in the development of HE remains unclear. Therefore, we examined the effect of constant alcohol consumption on neurological decline in rats with chronic liver disease induced via bile-duct ligation (BDL). Method: 5-week BDL rats and Sham-operated controls were used. Day 7 after surgery, rats were administered alcohol twice a day (dose of 3g/kg, via gavage) for 4 weeks. Motor coordination (rotarod) and anxiety-like behavior (open field (OF) and elevated plus maze (EPM)) were assessed. Upon sacrifice, brains were collected, and western blot and immunohistochemical (IHC) analyses were used to investigate neuronal integrity in frontal cortex and cerebellum. Results: Alcohol (BDL-alcohol rats) further impaired motor coordination at weeks 2, 3, 4, and 5 compared to SHAM-alcohol (p<0.01). Furthermore, BDL-alcohol rats demonstrated an increase in anxiety-like behavior; increase in time spent in the closed arms of EPM and decrease in time spent in the center of the OF (p<0.05 vs SHAM-alcohol). BDL-alcohol rats demonstrated a decrease in neuronal markers of NeuN and SMI311 (p<0.01 and p<0.05, respectively), an increase in apoptotic markers of cleaved/pro-caspase3 (p<0.001), an increase in necroptosis markers of pRIP3 and pMLKL (p<0.01 and p<0.001, respectively), a decrease in total antioxidant capacity (p<0.001) and an increase in oxidative stress marker of 4-HNE (p<0.05) in the cerebellum (not found in frontal cortex) compared to all groups. IHC results confirmed the colocalization of apoptotic marker (cleaved Caspase3) and necroptosis marker (pMLKL) in the granular and Purkinje layer neurons of the cerebellum of BDL-alcohol rats. Conclusion: Constant alcohol consumption exacerbates HE and leads to neuronal loss via apoptosis and necroptosis in the cerebellum. Additionally, higher levels of oxidative stress marker of 4-HNE and decreased total antioxidant capacity in the cerebellum of BDL-alcohol rats suggest that oxidative stress is a triggering factor leading to neuronal loss/injury. These results demonstrate an adverse effect of constant alcohol consumption on the development of HE and neuronal integrity in chronic liver disease.
Farzaneh Tamnanloo, Xiaoru Chen, Mélanie Tremblay, Christopher F. Rose.
Background: Hepatic encephalopathy (HE) is a debilitating neurological complication of chronic liver disease with alcohol being a common etiological factor. However, excessive alcohol consumption has been shown to impact neurological integrity. To date, the influence of alcohol in the development of HE remains unclear. Therefore, we examined the effect of constant alcohol consumption on neurological decline in rats with chronic liver disease induced via bile-duct ligation (BDL). Method: 5-week BDL rats and Sham-operated controls were used. Day 7 after surgery, rats were administered alcohol twice a day (dose of 3g/kg, via gavage) for 4 weeks. Motor coordination (rotarod) and anxiety-like behavior (open field (OF) and elevated plus maze (EPM)) were assessed. Upon sacrifice, brains were collected, and western blot and immunohistochemical (IHC) analyses were used to investigate neuronal integrity in frontal cortex and cerebellum. Results: Alcohol (BDL-alcohol rats) further impaired motor coordination at weeks 2, 3, 4, and 5 compared to SHAM-alcohol (p<0.01). Furthermore, BDL-alcohol rats demonstrated an increase in anxiety-like behavior; increase in time spent in the closed arms of EPM and decrease in time spent in the center of the OF (p<0.05 vs SHAM-alcohol). BDL-alcohol rats demonstrated a decrease in neuronal markers of NeuN and SMI311 (p<0.01 and p<0.05, respectively), an increase in apoptotic markers of cleaved/pro-caspase3 (p<0.001), an increase in necroptosis markers of pRIP3 and pMLKL (p<0.01 and p<0.001, respectively), a decrease in total antioxidant capacity (p<0.001) and an increase in oxidative stress marker of 4-HNE (p<0.05) in the cerebellum (not found in frontal cortex) compared to all groups. IHC results confirmed the colocalization of apoptotic marker (cleaved Caspase3) and necroptosis marker (pMLKL) in the granular and Purkinje layer neurons of the cerebellum of BDL-alcohol rats. Conclusion: Constant alcohol consumption exacerbates HE and leads to neuronal loss via apoptosis and necroptosis in the cerebellum. Additionally, higher levels of oxidative stress marker of 4-HNE and decreased total antioxidant capacity in the cerebellum of BDL-alcohol rats suggest that oxidative stress is a triggering factor leading to neuronal loss/injury. These results demonstrate an adverse effect of constant alcohol consumption on the development of HE and neuronal integrity in chronic liver disease.
Alcohol accelerates hepatic encephalopathy in a rat bile duct ligation model.
Xiaoru Chen, Mariana Oliveira, Grégory Petrazzo, Rafael Ochoa-Sanchez, Mélanie Tremblay, Christopher Rose.
Hepatic encephalopathy (HE) is a common and debilitating neuropsychiatric complication of liver disease characterized by a constellation of symptoms, including cognitive, psychiatric and motor disturbances. One of the causes of liver disease is alcoholic cirrhosis, which can induce acquired cerebellar degeneration syndrome, atrophy of the cerebellum producing symptoms of ataxia and motor difficulties. In the literature, very few experimental studies concern the role of alcohol on the development of HE. Here we examine the effects of ethanol on bile duct ligation (BDL) rats, a HE model, using a variety of behavioral tasks on motor coordination, open field behavior and memory. BDL rats were subjected to double ligation on the common bile duct with dissection between the ligatures, sham-operated rats underwent the same surgery except for ligation. We first effectuated a dose-response study (N=4-5) in BDL rats to determine the optimal dosage regimen of ethanol. 7 days after surgery, BDL rats were given ethanol by intragastric gavage with 1, 2, 4, 6 and 8 g/kg or saline over a 4-weeks period. Crews Scale and blood ethanol concentration were checked weekly. The accelerating rotarod was used to assess motor coordination. Then the dose regimen of intermittent exposure of 2X3g/kg 3h apart 5 days per week during 4 weeks was chosen to further elucidate the evolution of neurological deficits in both BDL rats and their sham operated controls. The behavioral assessments were performed at 7, 14, 21, 28 and 35 days, one day after blood ethanol concentration dropped to zero of each treatment cycle. Rats were assigned to the following groups (N=9-10): Sham + saline, Sham + ethanol, BDL + saline, BDL + ethanol. The baseline of rotarod performance and gait analysis parameters (at 7 days) showed there were no difference among groups before ethanol gavage. However, BDL + ethanol group rats had gradually impaired coordination performance and motor activity, contrary to those of Sham+ ethanol group who improved their performance gradually by learning. No significant differencet was observed in gait analysis. In addition, blood ethanol concentration over time showed a different metabolism mode in BDL compared with that of Sham + ethanol rats. The changes in coordination performance of BDL+ethanol rats might be associated with some biomolecular changes in the brain. The fact that the front cortex water content in these rats slightly increased as well as the weight of cerebellum reduced suggests that neuroinflammation and cerebellar atrophy might be involved in. Results of the current study indicate heavy alcohol ingestion impairs gradually motor coordination during cirrhosis. BDL rats treated with alcohol allowed perform studies on motor alterations in less than 5 weeks, will be an efficient animal model for the study of HE induced by ethanol and for the search of new treatment strategies.
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