Rapamycin reduces podocyte adhesion damage caused by sublytic C5b-9 via autophagy activation

Objective To determine the effect of rapamycin on sublytic C5b-9-induced podocyte adhesion damage, and whether autophagy is involved in this progression. Methods Sublytic complement C5b-9 stimulation was used in vitro. Autophagosomes were viewed using electron microscopy. Western blotting was used to measure the change of autophagy-related markers. Attachment assay was used to assess the adhesion of podocyte. Confocal microscopy was used to explore the expression patterns of cytoskeletal protein F-actin. Flow cytometry was used to measure the level of adhesion-associated protein integrin α3. Results (1) For ensuring sublytic complement injury, the maximal amounts of anti-podocyte antiserum and 160×-diluted normal human serum were used without inducing cell lysis (defined as ＞5% LDH release). (2) Sublytic C5b-9 promoted autophagy in podocyte in vitro. The proautophagic effect of sublytic C5b-9 manifested in the form of accumulated autophagosomes and enhanced expression of LC3-Ⅱ. (3) Inhibition of autophagy by 3-methyadenine enhanced the effect of sublytic C5b-9-induced podocyte injury, including serious cytoskeleton damage and markedly reduced adhesion of podocyte. (4) Rapamycin treatment significantly improved the above lesions. (5) Rapamycin enhanced autophagy induced by sublytic C5b-9 in podocyte. Conclusions In summary, rapamycin can improve sublytic C5b-9-induced podocyte adhesion damage by appropriate autophagy activation. These findings provide important information for the development of appropriate protocols for the application of mTOR (mammalian target of rapamycin) inhibitors in podocytopathy.