Background/Purpose: We made the novel observation that glycosphingolipid (GSL) levels and neuraminidase (NEU) (an enzyme that mediates GSL catabolism) activity/expression are altered in the kidneys and/or urine of lupus mice and human patients with proliferative nephritis compared to their non-nephritic counterparts and healthy controls. Specifically, elevated GSL levels were observed in the mesangial region of glomeruli. We hypothesize that activation of mesangial cells (MCs) in the progression of lupus nephritis is mediated in part by NEU1-mediated LacCer elevation, contributing to renal inflammation in lupus nephritis.

Methods: NEU1 expression and the GSL Lactosylceramide (LacCer) levels in kidney sections were analyzed by immunohistochemistry and immunofluorescence. MES13 mouse MCs were: 1) incubated with the GSLs LacCer and glucosylceramide (GlcCer) over time and cytokine gene expression measured by real-time RTPCR; and 2) transfected with increasing amounts of an expression vector for NEU1, and IL-6 production was measured in the media by ELISA, while cellular LacCer levels were measured by Supercritical Fluid Chromatography coupled with tandem mass spectrometry. Primary MCs were grown from glomeruli isolated from kidneys of pre-nephritic MRL/lpr mice. Once established and verified to be MCs, the primary MCs were analyzed between passages 6 and 10 following activation with heat aggregated IgG (HA-IgG, a mimic of immune complex deposition) for IL-6 and MCP-1 production by ELISA, Neu1 message levels by real-time RTPCR, and NEU1 expression and cellular localization by immunofluorescence.

Results: NEU1 and LacCer were observed to be highly expressed in MCs on renal sections of nephritic MRL/lpr mice by immunohistochemistry. Using the mouse MES13 MC line, we demonstrate that exogenous addition of LacCer and GlcCer increases the expression of several cytokines, including IL-6. Over-expression of NEU1 increases LacCer levels and IL-6 production. In primary MCs from pre-nephritic lupus mice, Neu1 expression, IL-6 production and MCP-1 production are significantly increased following addition of HA-IgG in a dose-dependent manner. IL-6 production is significantly increased within 6 hours following addition of HA-IgG stimulation. Importantly, addition of an FDA-approved inhibitor of NEU activity significantly and dose-dependently inhibited HA-IgG-induced IL-6, but not MCP-1 production. NEU1, typically located in the lysosomes, is translocated to the plasma membrane in some cell types upon activation. Analyses of changes in cellular localiation of NEU1 in HA-IgG-activated lupus prone MCs is pending.

Conclusion: Together these results suggest that immune complex activated IL-6 production of MRL/lpr lupus prone MCs is mediated by NEU-mediated GSL catabolism. Targeting NEU activity, or NEU1 specifically, may reduce MC cytokine production and thus renal inflammation in lupus nephritis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/immune-complex-induced-il-6-production-by-lupus-prone-mesangial-cells-is-mediated-by-neuraminidase-activity/