Hepatic Expression Profiling Shows Involvement of PKC Epsilon, DGK eta, Tnfaip, and Rho kinase in type 2 Diabetic Nephropathy Rats Jeena Gupta, Anil Bhanudas Gaikwad, and Kulbhushan Tikoo * Laboratory of Chromatin Biology, Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research (NIPER), S.A.S. Nagar, Punjab, India ABSTRACT Type 2 diabetes is associated with an increased risk for developing fatty liver disease, which results in an increased incidence of diabetic nephropathy. Hence, the present study was conceived to identify transcriptional changes in the liver that can provide molecular mediators for increased risk of developing nephropathy associated with type 2 diabetes. Type 2 diabetes was rendered in male SD rats using both high-fat diet and low dose of streptozotocin (35 mg/kg, intraperitonially, i.p.). Hepatic gene expression profiling was performed in animals after development of diabetic nephropathy. The gene expression data were validated by RT-PCR, protein expression, and immunohistochemistry. Gene expression profiling data revealed dramatic increase in expression of PKC epsilon, TNF-alpha-induced protein (four- to seven-folds), and decrease in the expression of DGK eta in the liver of diabetic nephropathic rats. Furthermore, there was an increase in expression of genes regulating Rho signaling pathway, which was further confirmed by increase in Rho kinase activity. To the best of our knowledge, this is the first report which shows the involvement of PKC epsilon, DGK eta, Tnfaip, and Rho kinase in the liver of type 2 diabetic rats and its association with diabetic nephropathy. J. Cell. Biochem. 111: 944–954, 2010. ß 2010 Wiley-Liss, Inc. KEY WORDS: LIVER; TYPE 2 DIABETES; PKC EPSILON; DIABETIC NEPHROPATHY; RHO KINASE L iver is the main organ of glucose disposal and also the foremost organ to be affected in diabetes. The excess of free fatty acids and hyperinsulinemia found in the insulin-resistant state is known to be directly toxic to hepatocytes [Lewis et al., 2002]. Fatty liver is known to be associated with chronic kidney disease and is also an early marker for the progression of diabetic nephropathy [Targher et al., 2008a,b]. Patients with type 2 diabetes seem to be at increased risk for developing fatty liver disease and certainly have a higher risk for developing fibrosis and cirrhosis [Day, 2006; Tolman et al., 2007]. However, the underlying mechanisms by which prevalence of fatty liver in type 2 diabetes increase the risk for developing diabetic nephropathy are poorly understood. In liver, in addition to pathways that are known to be involved in maintaining glucose homeostasis, several other pathways are also affected, finally leading to type 2 diabetes [Saltiel and Kahn, 2001]. These effects are achieved not only by altering activities of key enzymes involved in various signaling cascades via phosphorylation but also by altering expression of certain genes via phosphorylation of certain transcription factors. Therefore, a global analysis of hepatic gene expression appears to be essential for understanding the molecular mechanisms underlying the development of type 2 diabetes and its complications. DNA microarray technology has been widely used to gain insight into the overall effect on gene expression under various pathological conditions and provides a global view of the molecular events, which led to the development of these pathologies. This high Journal of Cellular Biochemistry ARTICLE Journal of Cellular Biochemistry 111:944–954 (2010) 944 Abbreviations used: STZ, streptozotocin; NPD, normal pellet diet; HFD, high-fat diet; HFD/STZ, high-fat diet þ streptozotocin; PGL, plasma glucose; PI, plasma insulin; PTCHL, plasma total cholesterol; PTG, plasma triglycerides; BUN, blood urea nitrogen and PCR, plasma creatinine levels; PKC, protein kinase C; TNF, tumor necrosis factor; DGK, diacylglycerol kinase. Additional Supporting Information may be found in the online version of this article. Grant sponsor: Council of Scientific and Industrial Research (CSIR); Grant number: 09/727(0041)/2005-EMR-I. *Correspondence to: Dr. Kulbhushan Tikoo, Associate professor, Laboratory of Chromatin Biology, Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research (NIPER), Sector 67, S.A.S. Nagar, Punjab-160 062, India. E-mail: tikoo.k@gmail.com Received 17 June 2010; Accepted 15 July 2010  DOI 10.1002/jcb.22783  ß 2010 Wiley-Liss, Inc. Published online 27 July 2010 in Wiley Online Library (wileyonlinelibrary.com).