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Abstract: PO0670

Store-Operated Ca2+ Entry Contributed to High Glucose-Induced Podocyte Injury

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Tao, Yu, University of North Texas Health Science Center, Fort Worth, Texas, United States
  • Chaudhari, Sarika, University of North Texas Health Science Center, Fort Worth, Texas, United States
  • Yazdizadeh Shotorbani, Parisa, University of North Texas Health Science Center, Fort Worth, Texas, United States
  • Ma, Rong, University of North Texas Health Science Center, Fort Worth, Texas, United States
Background

Diabetic Nephropathy is one of the major microvascular complications of diabetes and the most common cause of end stage renal disease. Hyperglycemia is a known pathogenic stimulus for onset and progression of diabetic nephropathy. Podocyte injury is one of early features of the disease. However, the mechanism of the diabetes-induced podocyte injury is not fully understood. Store operated Ca2+ entry (SOCE) has multiple functions in both excitable and non-excitable cells. This ubiquitous Ca2+ signaling includes two key components, Orai1 (a plasma membrane protein mediating SOCE) and STIM1 (an ER membrane protein sensing Ca2+ level in the ER lumen). Previous studies have demonstrated that alterations in SOCE are involved in cell dysfunction in many cell types. However, whether and how SOCE contributes to podocyte injury in diabetes settings are not known. The present study was aimed to determine that enhanced SOCE mediated high glucose (HG)-induced podocyte injury by upregulating calpain activity.

Methods

All experiments were performed using cultured human podocytes. Western blot was conducted to estimate Orai1, STIM1 and nephrin protein abundance. Ca2+ imaging was used to analyze SOCE. Confocal microscopy was used to visualize podocyte actin arrangement. Calpain activity was determined by calpain activity assay kits.

Results

HG (25mM) treatment significantly increased Orai1, but not STIM1 protein abundance for time periods ranging from 2 to 12 hours. The HG-induced Orai1 response was dose dependent. Ca2+ imaging experiment showed that HG treatment for 12 hours significantly increased SOCE. In addition, HG treatment significantly decreased nephrin (a podocyte marker) protein abundance and resulted in cytoskeleton rearrangement by formation of cortical F-actin. Both HG responses were significantly blunted by BTP2 (4 µM), an SOCE inhibitor. Furthermore, we found that activation of SOCE by thapsigargin (1 μM) increased calpain activity which was abolished by BTP2. In addition, BTP2 blunted the increased calpain activity induced by HG treatment. Moreover, calpeptin (a calpain inhibitor) attenuated the HG-induced reduction of nephrin protein abundance.

Conclusion

The present study suggests that enhanced SOCE contributes to HG-induced podocyte injury by increasing calpain activity.

Funding

  • NIDDK Support