Inonotus obliquus alleviates proteinuria of MRL/lpr mice by inhibiting ROS-NLRP3-mediated podocyte pyroptosis via facilitating Nrf2 translocation

Abstract

Proteinuria is a main feature of lupus nephritis (LN) in which NLRP3-mediated podocyte pyroptosis further exacerbates proteinuria. Our previous study confirmed the proteinuria-lowering, podocyte-protecting and anti-inflammatory effects of Inonotus obliquus (chaga) in diabetic kidney disease rats. Our in vivo and in vitro studies were conducted in female MRL/lpr mice and human glomerular podocyte cell line treated with IgG from LN patients' serum. Proteinuria changes were detected by urine protein test kit, C3 and ds-DNA by ELISA, renal histopathology and podocyte morphology were studied also. Western blot and RT-PCR were carried out to detect protein and mRNA levels of WT-1, Nephrin, NLRP3, GSDMD-N, Caspase-1 and IL-1β. ROS in podocytes was detected by fluorescent probes and MDA, LDH levels were measured by ELISA, and Nrf2 translocation was detected by immunofluorescence. In vivo study, chaga significantly decreased proteinuria, improved podocyte morphology and kidney pathological changes, meanwhile increased WT-1 and Nephrin expressions and suppressed NLRP3, GSDMD-N, Caspase-1 and IL-1β expressions when compared to the LN model. In vitro study, similar to Nrf2 activator, chaga facilitated Nrf2 translocate from cytosol into nucleus, suppressed ROS, MDA and LDH generation, then mitigated podocyte pyroptosis. The results indicated that the proteinuria-reducing effect of chaga on LN mice may be related to its promotion of Nrf2 translocation, thereby inhibiting NLRP3/Caspase-1/GSDMD-N-mediated podocyte pyroptosis.