Circ_0000064 promotes high glucose-induced renal tubular epithelial cells injury to facilitate diabetic nephropathy progression through miR-532-3p/ROCK1 axis

Abstract

Background Circular RNA (circRNA) has been shown to mediate diabetic nephropathy (DN) development by regulating renal tubular epithelial cells (RTECs) injury. However, the role and mechanism of circ_0000064 in high glucose (HG)-induced RTECs injury have not been fully elucidated. Methods Human RTECs (HK-2) were exposed to HG to induce cell injury. Cell oxidative stress was assessed by detecting the levels of oxidative stress-markers. Moreover, cell proliferation and apoptosis were determined by CCK8 assay, EDU assay and flow cytometry. The protein levels of proliferation markers, apoptosis markers and Rho-associated coiled-coil-containing kinase 1 (ROCK1) were measured using western blot analysis. Furthermore, quantitative real-time PCR was performed to assess the expression of circ_0000064, microRNA (miR)-532-3p and ROCK1. The interaction between miR-532-3p and circ_0000064 or ROCK1 was confirmed by dual-luciferase reporter assay and RNA pull-down assay. Results Our results revealed that HG treatment could promote HK-2 cells oxidative stress, apoptosis, fibrosis, and inhibit proliferation. Circ_0000064 expression was increased in the serum of DN patients and HG-induced HK-2 cells, and silenced circ_0000064 could relieve HG-induced HK-2 cells injury. MiR-532-3p could be sponged by circ_0000064, and its overexpression also alleviated HG-induced HK-2 cells injury. Besides, the regulation of circ_0000064 knockdown on HG-induced HK-2 cells injury could be reversed by miR-532-3p inhibitor. Additionally, ROCK1 was a target of miR-532-3p, and its expression was inhibited by circ_0000064 knockdown. The inhibition effect of circ_0000064 knockdown on HG-induced HK-2 cells injury also could be reversed by overexpressing ROCK1. Conclusion In summary, circ_0000064 knockdown might alleviate HG-induced HK-2 cells injury via regulating the miR-532-3p/ROCK1 axis, which provided a new perspective for DN treatment.

The role of circular RNA in Diabetic Nephropathy

“…It also inhibits hyperglycemia-induced oxidative stress, inflammation, apoptosis and extracellular matrix accumulation in mesangial cells, thereby inhibiting mesangial cell proliferation and renal fibrosis. Therefore, CIRC_0000064 may be a key regulatory factor in promoting the progression of diabetic nephropathy and provide a new target for the treatment of DKD [42,43].…”

CircRNAs: versatile players and new targets in organ fibrosis

“…circ_0000064 expression is also increased in the serum of patients with diabetic nephropathy and HG-induced HK-2 cells. Silencing circ_0000064 attenuates HG-induced HK-2 cell injury by targeting the miR-532-3p/ROCK1 axis [ 80 ]. Overexpression of circHIPK3 aggravates folic acid-induced renal tubulointerstitial fibrosis by sponging miR-30a, subsequently activating TGF-β1 signaling in HK-2 cells [ 81 ].…”