Inhibition of Endocytosis of Clathrin-Mediated Angiotensin II Receptor Type 1 in Podocytes Augments Glomerular Injury
Authors: Kazunori Inoue, Xuefei Tian, Heino Velazquez, Keita Soda, Zhen Wang, Christopher E. Pedigo, Ying Wang, Elizabeth Cross, Marwin Groener, Jee-Won Shin, Wei Li, Hossam Hassan, Koichi Yamamoto, Peter Mundel, Shuta Ishibe
Journal: Journal of the American Society of Nephrology (2019)

Significance Statement The amelioration of proteinuria resulting from inhibition of the renin-angiotensin pathway is thought to be predominantly caused by reduction in intraglomerular pressure. However, because studies have produced conflicting findings, whether podocyte-associated angiotensin II receptor signaling directly contributes to podocyte injury remains unclear. Angiotensin II receptor type 1 (AT1R) is internalized by clathrin- and dynamin-mediated endocytosis, and in this study the authors used podocyte-specific Dynamin 1 and 2 double-knockout mice to examine the effect of angiotensin II stimulation on AT1R in these double-knockout mice. Loss of AT1R internalization accentuated Rac1 activation and membrane ruffling in Dnm double-knockout podocytes. Podocyte-specific deletion of the receptor in Dnm double-knockout mice demonstrated improved albuminuria and kidney function and attenuation of membrane abnormalities—findings suggesting that podocyte-associated AT1R signaling augments podocyte injury. Background Inhibition of the renin-angiotensin system remains a cornerstone in reducing proteinuria and progression of kidney failure, effects believed to be the result of reduction in BP and glomerular hyperfiltration. However, studies have yielded conflicting results on whether podocyte-specific angiotensin II (AngII) signaling directly induces podocyte injury. Previous research has found that after AngII stimulation, β -arrestin–bound angiotensin II receptor type 1 (AT1R) is internalized in a clathrin- and dynamin-dependent manner, and that Dynamin1 and Dynamin2 double-knockout mice exhibit impaired clathrin-mediated endocytosis. Methods We used podocyte-specific Dyn double-knockout mice to examine AngII-stimulated AT1R internalization and signaling in primary podocytes and controls. We also examined the in vivo effect of AngII in these double-knockout mice through renin-angiotensin system blockers and through deletion of Agtr1a (which encodes the predominant AT1R isoform expressed in kidney, AT1aR). We tested calcium influx, Rac1 activation, and lamellipodial extension in control and primary podocytes of Dnm double-knockout mice treated with AngII. Results We confirmed augmented AngII-stimulated AT1R signaling in primary Dnm double-knockout podocytes resulting from arrest of clathrin-coated pit turnover. Genetic ablation of podocyte Agtr1a in Dnm double-knockout mice demonstrated improved albuminuria and kidney function compared with the double-knockout mice. Isolation of podocytes from Dnm double-knockout mice revealed abnormal membrane dynamics, with increased Rac1 activation and lamellipodial extension, which was attenuated in Dnm double-knockout podocytes lacking AT1aR. Conclusions Our results indicate that inhibiting aberrant podocyte-associated AT1aR signaling pathways has a protective effect in maintaining the integrity of the glomerular filtration barrier.