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ORIGINAL RESEARCH ARTICLE

Renal Activity of Akt Kinase in Obese Zucker Rats

Jana dychová^*, Ludmila Kazdová^*, Terezie Pelikanová^*, Jessie N. Lindsley, Sharon Anderson^, and Radko Komers^*,,1

^* Diabetes Center, Institute for Clinical and Experimental Medicine, Prague, Czech Republic; Division of Nephrology and Hypertension, Department of Medicine, Oregon Health and Science University, Portland, Oregon 97239; and Portland VA Medical Center, Portland, Oregon 97239

To whom requests for reprints should be addressed at ¹ Division of Nephrology and Hypertension, Oregon Health & Science University, PP262, 3314 SW US Veterans Hospital Road, Portland, OR 97239-2940. E-mail:komersr{at}ohsu.edu

Insulin resistance (IR) and consequent hyperinsulinemia are hallmarks of Type 2 diabetes (DM2). Akt kinase (Akt) is an important molecule in insulin signaling, implicated in regulation of glucose uptake, cell growth, cell survival, protein synthesis, and endothelial nitric oxide (NO) production. Impaired Akt activation in insulin-sensitive tissues contributes to IR. However, Akt activity in other tissues, particularly those affected by complications of DM2, has been less studied. We hypothesized that hyperinsulinemia could have an impact on activity of Akt and its effectors involved in regulation of renal morphology and function in DM2. To address this issue, renal cortical Akt was determined in obese Zucker rats (ZO), a model of DM2, and lean controls (ZL). We also studied expression and phosphorylation of the mammalian target of rapamycin (mTOR) and endothelial NO synthase (eNOS), molecules downstream of Akt in the insulin signaling cascade, and documented modulators of renal injury. Akt activity was measured by a kinase assay with GSK-3 as a substrate. Expression of phosphorylated (active) and total proteins was measured by immunoblotting and immunohistochemistry. Renal Akt activity was increased in ZO as compared to ZL rats, in parallel with progressive hyperinsulinemia. No differences in Akt were observed in the skeletal muscle. Corresponding to increases in Akt activity, ZO rats demonstrated enhanced phosphorylation of renal mTOR. Acute PI3K inhibition with wortmannin (100 µg/kg) attenuated renal Akt and mTOR activities in ZO, but not in ZL rats. In contrast to mTOR, eNOS phosphorylation was similar in ZO and ZL rats, despite higher total eNOS expression. In conclusion, ZO rats demonstrated increases in renal Akt and mTOR activity and expression. However, eNOS phosphorylation did not follow this pattern. These data suggest that DM2 is associated with selective IR in the kidney, allowing pro-growth signaling via mTOR, whereas potentially protective effects mediated by eNOS are blunted.

Key Words: Akt kinase • phosphatidylinositol 3-kinase • Zucker rat • mammalian target of rapamycin • mTOR • endothelial nitric oxide synthase