Biomedical subjects

Nicholas A Shackel

Publications and source records attributed to Nicholas A Shackel.

3 recordsLinked to original sources

PubMed2003

Novel differential gene expression in human cirrhosis detected by suppression subtractive hybridization.

Pathogenic molecular pathways in cirrhotic liver diseases such as hepatitis C virus (HCV), autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are poorly characterized. Differentially expressed genes are often important in disease pathogenesis. Suppression subtractive hybridization (SSH) is a genome-wide approach that enriches for differentially expressed mRNA transcripts. We aimed to make novel observations of differential gene expression in cirrhosis using SSH combined with quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR). Liver transcriptomes in HCV cirrhosis, AIH cirrhosis, PBC, and nondiseased liver tissue were examined by SSH. Resulting complementary DNA (cDNA) clones were rescreened for differential expression by dot-blot hybridization and then sequenced. Selected gene expression was quantified by real-time RT-PCR. Following SSH, 694 clones were rescreened for differential gene expression, of which 145 were sequenced and found to derive from 89 different genes. Seven clones were homologous only with expressed sequence tag (EST) sequences encoding genes having no known function. Up-regulated expression of four genes was confirmed by real-time RT-PCR: transmembrane 4 superfamily member 3 (tetraspanin CO-029) in all forms of cirrhosis, hedgehog interacting protein (HIP) in AIH cirrhosis and chitinase 3-like-1 (HC gp-39 or ykl-40) and arginine-glutamic acid repeat (RERE) in HCV cirrhosis. RERE gene polymorphisms and splice variants were observed in all tissues examined. Tetraspanin CO-029 up-regulation was primarily localized to bile ductular cells. In conclusion, novel observations of differential gene expression in human cirrhosis were made using SSH as the primary discovery tool. In particular, further studies of the RERE gene and its products in HCV associated liver disease are warranted.

Adipokines↗

PubMed2002

Insights into the pathobiology of hepatitis C virus-associated cirrhosis: analysis of intrahepatic differential gene expression.

The pathogenesis of hepatitis C virus (HCV)-associated liver injury involves many genes from multiple pathogenic pathways. cDNA array analysis, which examines the expression of many genes simultaneously, was used to achieve new insights into HCV liver injury. Membrane-based cDNA arrays of 874 genes compared HCV-associated cirrhosis with autoimmune hepatitis-associated cirrhosis as an inflammatory and cirrhotic control, and with nondiseased liver tissue. Array analysis identified many differentially expressed genes that are important in inflammation, fibrosis, proliferation, signaling, apoptosis, and oxidative stress. Genes up-regulated in HCV-associated cirrhosis were predominantly associated with a Th1 immune response, fibrosis, cellular proliferation, and apoptosis. Novel observations of differential gene expression included increased expression of secreted apoptosis-related protein 3, a Wnt pathway gene possibly involved in cellular apoptosis. EMMPRIN (CD147) and discoidin domain receptor 1 (CD167) were also shown to be increased and are likely to play a role in liver fibrosis. Real-time quantitative reverse transcriptase-polymerase chain reaction confirmed the increased expression of 15 genes. The comparison of HCV cirrhosis with autoimmune hepatitis cirrhosis showed a marked difference in the apoptosis-associated gene profile with HCV cirrhosis characterized by increased proapoptotic gene expression whereas autoimmune hepatitis was characterized by increased expression of both antiapoptotic and proapoptotic genes. Furthermore, expression of beta-catenin and the fibrosis-associated protein EMMPRIN were localized by immunohistochemistry to the plasma membranes of hepatocytes and biliary epithelium. In conclusion, HCV-associated cirrhosis was characterized by a proinflammatory, profibrotic, and proapoptotic gene expression profile.

Adult↗

PubMed2002

Gene array analysis and the liver.

Functional genomics methods promise a previously unparalleled high-throughput examination of intrahepatic gene expression. Profiling transcriptomes as well as examining the coordinate expression of many genes in diverse pathobiologic pathways is now possible with techniques such as gene array analysis. However, the nature of the hepatic transcriptome, limitations of the functional genomics methodologies used, and analysis of the data generated are often poorly understood. Further, the use of replicates during analysis and the downstream confirmation of results using a supplemental methodology are important but frequently neglected aspects of experimental design. This review presents an overview of functional genomics methodologies in the organ-specific context of studying intrahepatic gene expression. The focus of the review is the most widely used functional genomic methodology, gene array analysis. The nature of the hepatic transcriptome is discussed, together with a summary of gene array analysis methodologies. Analysis of data generated by gene array analysis, determination of intrahepatic cellular origin of transcript expression, and studies of intrahepatic gene expression using gene array analysis are also reviewed. Finally, the importance of a coordinate experimental approach examining both liver protein and mRNA expression is presented.

Humans↗