GSDMD-MEDIATED PYROPTOSIS IN MACROPHAGES IS INVOLVED IN THE PATHOGENESIS OF ENDOMETRIOSIS

Abstract

Gasdermin D (GSDMD)-executed pyroptosis is a crucial event involved in inflammatory diseases. But its role in endometriosis remains largely unknown. This study aims to determine the role of GSDMD -mediated pyroptosis in endometriosis. Endometriosis was induced by injection of syngeneic endometrial tissue fragments into C57/BL6 female wild-type (WT) mice. The mice were euthanized and endometriotic tissues were collected two weeks after modeling. Infiltration of inflammatory cells was detected by immunostaining. Expression of pyroptosis-related proteins were detected by western blotting. We further generated GSDMD knock-out (KO) mice. Endometriosis was induced in GSDMD KO mice and WT controls. The average number, weight and size of the lesions were compared between two groups. Samples of endometriotic lesions were collected and single-cell RNA sequencing (sc-RNA seq) was performed using the 10x genomics chromium single cell controller. The establishment of endometriosis model was confirmed by histology. Immunostaining showed extensive macrophage infiltration in the lesions. Western blot demonstrated up-regulated expression of full-length and cleaved (activated form) GSDMD. We further induced endometriosis in eight GSDMD KO mice and seven WT controls. The average number, weight and maximum diameter of the lesions in KO mice were all markedly less than those in WT mice. Results of sc-RNA seq revealed that GSDMD was predominantly expressed in monocytes. The proportion of monocytes was 20.4% in WT lesions, but dropped to 9.6% in KO lesions, while the proportion in peripheral blood remained constant. Cell-cell interaction analysis suggested that monocytes regulated fibroblasts through interleukin-1β (IL-1β)/interleukin-1 receptor (IL1r) pathway. Differential expression analysis showed that both IL-1β in monocytes and IL1r in fibroblasts were down-regulated in lesions from KO mice. Our study indicated that activation of GSDMD-mediated pyroptosis in macrophages is involved in the pathogenesis of endometriosis. Knockout of GSDMD significantly suppressed the formation of endometriotic lesions, possibly through down-regulation of IL-1β/IL1r pathway.