Connexin32 deficiency is associated with liver injury, inflammation and oxidative stress in experimental non-alcoholic steatohepatitis.

Non-alcoholic steatohepatitis is a highly prevalent liver pathology featured by hepatocellular fat deposition and inflammation. Connexin32, which is the major building block of hepatocellular gap junctions, has a protective role in hepatocarcinogenesis and is downregulated in chronic liver diseases. However, the role of connexin32 in non-alcoholic steatohepatitis remains unclear. Connexin32-/- mice and their wild-type littermates were fed a choline-deficient high-fat diet. The manifestation of non-alcoholic steatohepatitis was evaluated based on a battery of clinically relevant read-outs, including histopathological examination, diverse indicators of inflammation and liver damage, in-depth lipid analysis, assessment of oxidative stress, insulin and glucose tolerance, liver regeneration and lipid-related biomarkers. Overall, more pronounced liver damage, inflammation and oxidative stress were observed in connexin32-/- mice compared to wild-type animals. No differences were found in insulin and glucose tolerance measurements and liver regeneration. However, two lipid-related genes, srebf1 and fabp3, were upregulated in Cx32-/- mice in comparison with wild-type animals. These findings suggest that connexin32-based signalling is not directly involved in steatosis as such, but rather in the sequelae of this process, which underlie progression of non-alcoholic steatohepatitis.

Connexin32 deficiency exacerbates carbon tetrachloride-induced hepatocellular injury and liver fibrosis in mice.

OBJECTIVE: Liver fibrosis results from the perpetuation of the normal wound healing response to several types of injury. Despite the wealth of knowledge regarding the involvement of intracellular and extracellular signaling pathways in liver fibrogenesis, information about the role of intercellular communication mediated by gap junctions is scarce. METHODS: In this study, liver fibrosis was chemically induced by carbon tetrachloride in mice lacking connexin32, the major liver gap junction constituent. The manifestation of liver fibrosis was evaluated based on a series of read-outs, including collagen morphometric and mRNA analysis, oxidative stress, apoptotic, proliferative and inflammatory markers. RESULTS: More pronounced liver damage and enhanced collagen deposition were observed in connexin32 knockout mice compared to wild-type animals in experimentally triggered induced liver fibrosis. No differences between both groups were noticed in apoptotic signaling nor in inflammation markers. However, connexin32 deficient mice displayed decreased catalase activity and increased malondialdehyde levels. CONCLUSION: These findings could suggest that connexin32-based signaling mediates tissue resistance against liver damage by the modulation of the antioxidant capacity. In turn, this could point to a role for connexin32 signaling as a therapeutic target in the treatment of liver fibrosis.

Immunohisto- and Cytochemistry Analysis of Connexins.

Immunohistochemistry (IHC) is a ubiquitous used technique to identify and analyze protein expression in the context of tissue and cell morphology. In the connexin research field, IHC is applied to identify the subcellular location of connexin proteins, as this can be directly linked to their functionality. The present chapter describes a protocol for fluorescent IHC to detect connexin proteins in tissues slices and cells, with slight modifications depending on the nature of biological sample, histological processing, and/or protein expression level. Basically, fluorescent IHC is a short, simple, and cost-effective technique, which allows the visualization of proteins based on fluorescent-labeled antibody-antigen recognition.

Connexins, Pannexins, and Their Channels in Fibroproliferative Diseases.

Cellular and molecular mechanisms of wound healing, tissue repair, and fibrogenesis are established in different organs and are essential for the maintenance of function and tissue integrity after cell injury. These mechanisms are also involved in a plethora of fibroproliferative diseases or organ-specific fibrotic disorders, all of which are associated with the excessive deposition of extracellular matrix components. Fibroblasts, which are key cells in tissue repair and fibrogenesis, rely on communicative cellular networks to ensure efficient control of these processes and to prevent abnormal accumulation of extracellular matrix into the tissue. Despite the significant impact on human health, and thus the epidemiologic relevance, there is still no effective treatment for most fibrosis-related diseases. This paper provides an overview of current concepts and mechanisms involved in the participation of cellular communication via connexin-based pores as well as pannexin-based channels in the processes of tissue repair and fibrogenesis in chronic diseases. Understanding these mechanisms may contribute to the development of new therapeutic strategies to clinically manage fibroproliferative diseases and organ-specific fibrotic disorders.