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Abstract: FR-PO410

RNA Binding Proteins Tristetraprolin and Human Antigen R Mediate Diabetic Tubulopathy via Modulation of Inflammation and Apoptosis of Renal Tubular Epithelial Cells

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Guo, Jia, The university of Toledo, medical center, Toledo, Ohio, United States
  • Cheng, Fei, The First Affiliated Hospital of Zhengzhou University, ZhengZhou, China
  • Gong, Rujun, University of Toledo Medical Center, Toledo, Ohio, United States
  • Liu, Zhangsuo, The first affiliated hospital of zhengzhou university, Zhengzhou, China
Background

Inflammatory components have been involved in diabetic tubulopathy, a key constituent of diabetic nephropathy (DN) pathology, with undefined molecular mechanisms. RNA binding proteins Tristetraprolin (TTP) and Human antigen R (HuR) have been implicated in inflammatory response in a variety of diseases and exert opposite modulatory effects on a number of inflammatory mediators. Nevertheless, their role in diabetic tubulopathy remains unknown. Evidence suggests that the expression and activity of TTP and HuR are regulated by phosphorylation that is catalyzed by a number of kinases, including GSK3β, a cell signaling transducer with a nephropathic action. The aim of this study was to examine the role of GSK3β -regulated TTP and HuR in diabetic tubulopathy by using an in vitro model of renal tubular epithelial cells (HK-2).

Methods

The mRNA or protein expressions were measured by qRT-PCR or immunoblotting. Apoptosis was analyzed by TUNEL staining. The subcellular localization of TTP, HuR and GSK3β was determined by immunofluorescence staining. GSK3β was silenced by RNA interference to examine the modulatory effect on TTP and HuR expression.

Results

After exposure of HK-2 cells to high ambient glucose (HG, 30Mm glucose), both mRNA and protein expressions of TTP were decreased, while those of HuR were increased, associated with an up-regulation of KIM-1 and TNF-α, loss of E-cadherin and augmented cellular apoptosis, implying that TTP and HuR were oppositely regulated upon high glucose-induced renal tubular cell injury. Tubular cell injury coincided with GSK3β hyperactivity, marked by GSK3β overexpression and suppressed phosphorylation of GSK3β at serine 9. Dual color fluorescent immunocytochemistry staining demonstrated that GSK3β is co-localized with TTP and HuR respectively in different subcellular compartments of HK-2 cells. Moreover, GSK3β seems to be essential for high glucose-induced TTP and HuR dysregulation, because silencing of GSK3β substantially reinstated TTP expression and offset HuR overexpression, concomitant with an improved cellular injury and apoptosis.

Conclusion

The GSK3β regulated RNA binding proteins TTP and HuR play a crucial role in mediating inflammation and apoptosis renal tubular cell injury in diabetes .

Funding

  • Government Support - Non-U.S.