Downregulation of TRPC6 protein expression by high glucose, a possible mechanism for the impaired Ca2+ signaling in glomerular mesangial cells in diabetes
AJP - Renal Physiology
The present study was performed to investigate whether transient receptor potential (TRPC)6 participated in Ca 2ϩ signaling of glomerular mesangial cells (MCs) and expression of this protein was altered in diabetes. Western blots and real-time PCR were used to evaluate the expression level of TRPC6 protein and mRNA, respectively. Cell-attached patch-clamp and fura-2 fluorescence measurements were utilized to assess angiotensin II (ANG II)-stimulated membrane currents and Ca 2ϩ responses in MCs.
... ϩ responses in MCs. In cultured human MCs, high glucose significantly reduced expression of TRPC6 protein, but there was no effect on either TRPC1 or TRPC3. The high glucose-induced effect on TRPC6 was time and dose dependent with the maximum effect observed on day 7 and at 30 mM glucose, respectively. In glomeruli isolated from streptozotocin-induced diabetic rats, TRPC6, but not TRPC1, was markedly reduced compared with the glomeruli of control rats. Furthermore, TRPC6 mRNA in MCs was also significantly decreased by high glucose as early as 1 day after treatment with maximal reduction on day 4. Patch-clamp experiments showed that ANG II-stimulated membrane currents in MCs were significantly attenuated or enhanced by knockdown or overexpression of TRPC6, respectively. Fura-2 fluorescence measurements revealed that the ANG II-induced Ca 2ϩ influxes were markedly inhibited in MCs with TRPC6 knockdown, reminiscent of the impaired Ca 2ϩ entry in response to ANG II in high glucose-treated MCs. These results suggest that the TRPC6 protein expression in MCs was downregulated by high glucose and the deficiency of TRPC6 protein might contribute to the impaired Ca 2ϩ signaling of MCs seen in diabetes. Ca 2ϩ influxes; glomeruli; diabetic nephropathy; hyperfiltration DIABETIC NEPHROPATHY (DN) is a major and devastating complication of diabetes. At the very onset of diabetes, a dominant pathophysiological characteristic is the development of renal glomerular hyperfiltration (2, 39). This early hemodynamic phenotype provokes the subsequent demise of a diabetic kidney. Diabetic hyperfiltration is derived from a combination of decreased responsiveness of both the renal afferent arterioles and glomerular mesangial cells (MCs) to vasoconstrictors (2, 3, 20, 29, 42) . MCs, located within glomerular capillary loops, physiologically regulate glomerular hemodynamics (26, 37). In * S. Graham and M. Ding contributed equally to the work.