Systemic lupus erythematosus (SLE) is an autoimmune disease with female predominance that involves multiple factors, including sex hormones and their receptor isoforms/variants, such as estrogen receptor alpha (ERα). The mechanisms of nuclear hormone receptor variants in immune cell activation and in SLE pathogenesis remain poorly understood. This study aimed to investigate the effect of two ERα variants, the classic full-length ERα66 and a short variant ERα46, in modulating proliferation and immune responses in in vitro models of lupus-like inflammation.

The male-derived human microglia HMC3 cells and murine macrophage RAW 264.7 cells and primary human renal mesangial cells (HRMC) from male or female donors were transfected with plasmid control (PC), ERα66, ERα46 plasmid, or a 1:1 ratio of both ERα variants. At 24-72 h post transfection, cell proliferation and viability were assessed by EdU and CCK-8 assays. Cells were further stimulated by a Toll-like receptor 7 (TLR7) agonist, IFNα, IFNγ, or LPS, and then the production of inflammatory cytokines was examined by ELISA and RT-qPCR.

In human microglial cells, known to play a critical role in CNS lupus, overexpressing ERα46 promoted cell proliferation. In a second myeloid cell line, murine RAW 264.7 macrophages, overexpressing ERα46 promoted cell viability but not proliferation. Both ERα66 and ERα46 promoted inflammatory cytokine expression in RAW macrophages in response to TLR7 stimulation. Lastly, we utilized mesangial cells HRMC, which have innate immune-like activities in the renal glomerulus during injury and can modulate inflammation in nephritis. Overexpressing ERα66 or ERα46 exerted distinct effects on proliferation and inflammation that were sexually dimorphic. Specifically, both ERα variants inhibited proliferation but minimally influenced the inflammatory phenotype of male HRMC. In female HRMC, ERα66 and ERα46 modulated the expression of multiple inflammatory cytokines, suggesting sex-specific functions of ERα variants.

Together, these findings demonstrate that ERα66 and ERα46 differentially influence cell proliferation as well as inflammatory responses to lupus-relevant stimulation that is cell type- and sex-specific.