Ectopic Liver Tissue Formation in Rats with Induced Liver Fibrosis

Introduction: The possible alternative approach to whole-organ transplantation is a cell-based therapy, which can also be used as a "bridge" to liver transplantation.  However, morphological and functional changes in the liver of patients suffering from chronic liver fibrosis and cirrhosis restrict the effectiveness of direct cell transplantation. Therefore, extra hepatic sites for cell transplantation, including the spleen, pancreas, peritoneal cavity, and subrenal capsule, could be a useful therapeutic approach for compensation of liver functions. However, a method of transplantation of hepatocytes into ectopic sites is needed to improve hepatocyte engraftment. Previously published data has demonstrated that mouse lymph nodes can support the engraftment and proliferation of hepatocytes as ES and rescue Fah mice from lethal liver failure. Thus, the aim of the study was to evaluate the engraftment of i.p. injected allogeneic hepatocytes into extra hepatic sites in albino rats with chemically induced liver fibrosis (LF). Materials and methods: Albino rats were randomly divided into 4 groups: (1) intact group (n = 18); (2) rats with induced LF (n = 18); (3) rats with induced LF and transplanted with hepatocytes (n = 18); (4) as a control, rats were treated with cyclosporine A only (n = 18). In order to prevent an immune response, groups 2 and 3 were subjected to immunosuppression by cyclosporine A (25 mg/kg per day). LF was induced using N-nitrosodimethylamine (NDMA), i.p., 10 mg/kg, three times a week for 4 weeks and confirmed by histological analysis of the liver samples. Hepatocytes transplantation (HT) was performed two days after NDMA exposure cessation by i.p. injection of 5×106 freshly isolated allogeneic hepatocytes. Liver function was assessed by quantifying blood biochemical parameters (ALT, AST, GGT, total protein, bilirubin, and albumin) at 1 week, 1 month, and 2 months after hepatocytes transplantation (HT). To confirm a hepatocytes’ engraftment, we conducted immunohistochemical staining against HepPar1.Results: We observed a 30% mortality rate among rats with LF within 1 week after NDMA exposure cessation, while 100% of animals with HT survived. ALT, AST, and GGT activities and bilirubin levels were markedly elevated in blood samples of LF rats compared to the control animals. However, HT significantly improved ALT, AST, and GGT activity as well as bilirubin levels. We also observed decreased levels of total protein and albumin in the blood serum of rats with LF, while HT normalized these parameters. At the same time, we have not detected any statistical differences of the studied parameters in the group 4, which was treated with Cyclosporine A only, compared with the intact animals. HepPar1 immunohistochemical staining of the different tissue sections demonstrated the presence of engrafted hepatocytes, mainly within enlarged Peyer's patches (aggregated lymphoid nodules in the lowest portion of the small intestine).Conclusion: The results of our study provide evidence that HT improves animal survival and liver functions. One potential reason for these results is that ectopic hepatic mass inside the Peyer's patches can rescue rats from liver failure

Isolation of primary human hepatocytes from cirrhotic liver

Chronic degenerative liver diseases are among most complex social, clinical and epidemiological health problems worldwide. This is due to the steady increase in the incidence and mortality of patients with this pathology. Orthotopic liver transplantation is the only way to save the lives of patients with decompensated diffuse and focal lesions of the liver. One-year survival after liver transplantation reaches 60-80%, but more than half of the patients on the waiting list do not survive until operation. In this regard, hepatocyte transplantation could be an option for the patients who are on the waiting list for organ transplantation

Isolation of primary human hepatocytes from cirrhotic liver

Chronic degenerative liver diseases are among most complex social, clinical and epidemiological health problems worldwide. This is due to the steady increase in the incidence and mortality of patients with this pathology. Orthotopic liver transplantation is the only way to save the lives of patients with decompensated diffuse and focal lesions of the liver. One-year survival after liver transplantation reaches 60-80%, but more than half of the patients on the waiting list do not survive until operation. In this regard, hepatocyte transplantation could be an option for the patients who are on the waiting list for organ transplantation

Differentiation of mesenchymal stem cells into hepatocytes

Acute and chronic liver diseases are common in Kazakhstan and other countries. These diseases are known to cause significant disability and death. In many cases, liver transplantation is the last resort for patients with end stage liver disease, but it is an extremely expensive procedure and is associated with many risks. The most important among them is an immune rejection. Autologous cell transplantation is a potential therapeutic approach for liver regeneration and could become an alternative to organ transplantation. In this regard, mesenchymal stem cells (MSCs) are a very attractive source for differentiation into hepatocytes. These cells can be isolated from bone marrow and adipose tissue of the patient and exponentially expanded in vitro. Transplantation of hepatocytes differentiated from MSCs could become a new promising approach in treatment of the patients with chronic liver conditions

Role of ROS in A?42 Mediated Activation of Cerebral Endothelial Cells

Introduction. There is substantial evidence that the deposition of aggregated amyloid-beta peptide (A?) in brain parenchyma and brain vessels is the main cause of neuronal dysfunction and death in Alzheimer’s disease (AD). A? exhibits multiple cytotoxic effects on neurons and glial cells and causes dysfunction of the blood brain barrier (BBB). In AD brains, an increased deposition of A? in the cerebral vasculature has been found to be correlated with increased transmigration of blood-borne inflammatory cells and neurovascular inflammation. However, regulatory mediators of these processes remain to be elucidated. In this study, we examined the role of ROS in actin polymerization and expression of adhesion molecules (P-selectin) on the surface of the cerebral endothelial cells (CECs) that are activated by A?42.Materials and methods. Mouse BEnd3 line (ATCC) was used in this research. BEnd3 cells respond to A? treatment similarly to human primary CECs and are a common model to investigate CECs’ function. We used immortalized bEnd3 cells as the following: controls; cells incubated with A?42 for 10, 30, and 60 minutes; cells incubated with 30 mM of antioxidant N-acetylcysteine (NAC) for 1 hr; and, cells pre-treated with NAC followed by A?42 exposure. We measured DHE fluorescence to investigate intracellular ROS production. Immunofluorescent microscopy of anti-P-selectin and oregon green phalloidin was used to quantify the surface P-selectin expression and actin polymerization, and Western blot analysis was used to analyze total P-selectin expression.Results. The results of this study have demonstrated a significant time-dependent ROS accumulation after 10 minutes, 30 minutes, and 60 minutes of A?42 treatment, while A?42 stimulated ROS production in CECs was attenuated by pre-treatment with the NAC antioxidant. We also found that A?42 increased P-selectin fluorescence at the surface of bEnd3 cells in a time dependent manner in parallel to ROS elevation. However, total expression levels of P-selectin were not changed following exposure to A?42. Pre-treatment with NAC attenuated A?42 induced P-selectin localization, while NAC alone did not significantly affect P selectin localization. As a positive control, H2O2 also increased P-selectin expression on the cell surface, which peaked after 30 minutes of H2O2 treatment. Exposure of CECs with A?42 promoted actin polymerization, which peaked after 10 minutes of A?42 treatment, while no significant increase of F-actin intensity was observed when cells were pre-treated with NAC. H2O2 was able to mimic A?42 induced oxidative stress, causing increased actin polymerization with similar timing.Conclusions. The results of our study have indicated that A?42 induced accumulation of P-selectin on the surface of bEnd3 cells and promoted actin polymerization, and all these events were correlated with ROS generation. The rapid post-translational cell signaling response mediated by ROS may well represent an important physiological trigger of the microvascular inflammatory responses in AD and requires further investigations

Differentiation of mesenchymal stem cells into hepatocytes

Acute and chronic liver diseases are common in Kazakhstan and other countries. These diseases are known to cause significant disability and death. In many cases, liver transplantation is the last resort for patients with end stage liver disease, but it is an extremely expensive procedure and is associated with many risks. The most important among them is an immune rejection. Autologous cell transplantation is a potential therapeutic approach for liver regeneration and could become an alternative to organ transplantation. In this regard, mesenchymal stem cells (MSCs) are a very attractive source for differentiation into hepatocytes. These cells can be isolated from bone marrow and adipose tissue of the patient and exponentially expanded in vitro. Transplantation of hepatocytes differentiated from MSCs could become a new promising approach in treatment of the patients with chronic liver conditions

Research on the bio-distribution of interferon gamma-containing pharmacocytes

Viral hepatitis B and C- is a widespread infectious disease of the liver and is a potential threat to human life. Treatment of HCV and HBV requires prolonged (parenteral or oral) administration of antiviral and immune-stimulating agents, which often have serious side effects that lead to abrupt termination of the therapy and the development of viral resistance. Targeted delivery of drugs directly to the affected organ is one of the most promising areas that seek to improve the outcome of treatment of patients with chronic diseases. Among the various drug delivery systems, erythrocytic pharmacocytes are of special interest. They are a shade of red blood cells, that could be loaded with antiviral drugs and drugs that boost the immune system. Owing to the fact that erythrocytes are destroyed by mononuclear phagocytes in the liver, it is assumable that the EP serve as a promising and direct means of delivering antiviral drugs and immune-modulators into the liver parenchyma to treat viral hepatitis

Research on the bio-distribution of interferon gamma-containing pharmacocytes

Viral hepatitis B and C- is a widespread infectious disease of the liver and is a potential threat to human life. Treatment of HCV and HBV requires prolonged (parenteral or oral) administration of antiviral and immune-stimulating agents, which often have serious side effects that lead to abrupt termination of the therapy and the development of viral resistance. Targeted delivery of drugs directly to the affected organ is one of the most promising areas that seek to improve the outcome of treatment of patients with chronic diseases. Among the various drug delivery systems, erythrocytic pharmacocytes are of special interest. They are a shade of red blood cells, that could be loaded with antiviral drugs and drugs that boost the immune system. Owing to the fact that erythrocytes are destroyed by mononuclear phagocytes in the liver, it is assumable that the EP serve as a promising and direct means of delivering antiviral drugs and immune-modulators into the liver parenchyma to treat viral hepatitis