Novel Effects of Combination Therapy Through Inhibition of Caspase-1/Gasdermin D Induced-Pyroptosis in Lupus Nephritis

Abstract

ObjectivesCombination therapy with mycophenolate mofetil, tacrolimus and steroids are effective in achieving complete remission in lupus nephritis (LN). Combination therapy uniquely downregulated caspase-1 compared with monotherapies, which can cleave gasdermin D (GSDMD) and was recently identified as the pyroptosis executioner. We therefore investigated whether combination therapy enabled the suppression of caspase-1/GSDMD-mediated pyroptosis in LN.MethodsExpression and activation of GSDMD were detected in kidney specimens of the human and mouse with LN using immunohistochemical staining and immunoblotting. Primary podocytes isolated from MRL/lpr mice were incubated with LPS+ATP, and pretreated with monotherapy or combination therapy. Inhibition of caspase-1/GSDMD-induced pyroptosis by combination therapy were assessed in MRL/lpr mice and human specimens. Pyroptosis was examined using a FAM caspase-1 kit and flow cytometry. The correlation between pyroptosis in peripheral blood and the systemic lupus erythematosus disease activity index (SLEDAI) was analyzed.ResultsKidney tissue specimens from LN patients and mice exhibited greatly increased expression levels and cleavage of GSDMD. In cultured podocytes, combination treatment significantly suppressed the activation of NLRP3 and caspase-1 and reduced GSDMD N-terminal levels. Combination therapy repressed disease progression through inhibition of caspase-1/GSDMD-mediated pyroptosis in both humans and MRL/lpr mice. Caspase-1/PI positive cell numbers in peripheral blood were positively correlated with SLE-DAI. LN patients with complete remission and partial remission had remarkably reduced caspase-1/PI positive cell numbers compared to baseline. Ac-FLTD-CMK, a GSDMD-derived inhibitor, prevented the development of LN.ConclusionCombination therapy suppressed caspase-1/GSDMD-mediated pyroptosis in vitro and in vivo and reduced disease progression.