Session Title: T-cell Biology and Targets in Autoimmune Disease
Session Type: Abstract Submissions (ACR)
Background/Purpose: The Ets factor Fli1 is implicated as a key modulator of lupus disease expression. Over-expressing Fli1 in healthy mice, results in the development of an autoimmune kidney disease similar to that observed in lupus. Lowering the global levels of Fli1 in two lupus mouse models significantly improved kidney disease and prolonged survival. Lowering the levels of Fli1 in hematopoietic cells in MRL/lpr lupus mice resulted in significantly improved kidney disease. The mechanism(s) by which Fli1 exerts this protective effect is unknown. The glycosphingolipid lactosylceramide (LacCer) is a ganglioside precursor to which sialic acid (SA) residues are added by ganglioside synthases or removed by sialidases. Loss of SA residues from gangliosides on the surface of podocytes is linked to proteinuria in glomerulonephritis and lipids with distinct chain lengths are thought to possess distinct biological activities. We demonstrate that lowering Fli1 levels decreases the levels of LacCer and the sialidase Neu1 in the kidney cortex and in T cells of MRL/lpr lupus prone mice. We present additional data demonstrating that Fli1 regulates Neu1 promoter activity in T cells.
Methods: Kidney and/or spleen were harvested from 17-19 week-old MRL/MpJ mice and MRL/lpr Fli1+/+ and Fli1+/- mice. T cells were isolated by negative selection from spleen and left unstimulated or stimulated with anti-CD3/CD28. Supercritical Fluid Chromatography coupled with tandem mass spectrometry was performed on kidney cortex homogenates and isolated T cells to quantify LacCer . Gene expression was analyzed by real-time RTPCR on RNA isolated from kidney cortex and T cells. Immunohistochemistry for LacCer was performed on frozen kidney sections. Neu1 promoter activity was examined by co-transfection of Neu1 promoter/reporter constructs and a Fli1 expression construct in human and mouse T cell lines.
Results: Diseased MRL/lpr mice express a significant 2.5-fold increase in the major LacCer species C16 in the kidney compared to age-matched MRL/MpJ, which do not exhibit kidney disease. MRL/lpr mice that are heterozygote for Fli1 (Fli1+/-) express a significant 2-fold decrease in LacCer C16 in the kidney compared to wild-type age-matched MRL/lpr mice. Similarly, a significant reduction in LacCer staining in cells of the glomeruli is observed by immunohistochemistry. Neu1 expression is significantly elevated 40-fold in the MRL/lpr compared to the MRL/MpJ kidney cortex, but is not significantly different in the MRL/lpr Fli1+/- compared to the wild-type MRL/lpr kidney cortex. Interestingly, Neu1 is reduced 3-4-fold and LacCer levels are significantly reduced 2.5-fold in T cells isolated from Fli1+/- compared to Fli1+/+ MRL/lpr mice. Over-expression of Fli1 results in a dose-dependent increase in Neu1 promoter activity in activated T cells.
Conclusion: Our results demonstrate that one mechanism by which reducing Fli1 levels may be protective in lupus kidney disease is to decrease LacCer levels through the regulation of Neu1 expression. We speculate that Fli1 regulates Neu1 expression in T cells that may act on gangliosides intra- (within the T cell) and inter- (on neighboring kidney cells upon infiltration in the kidney) ceullarly.
J. Berglind Schepp,
T. K. Nowling,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/lowering-fli1-levels-decreases-the-levels-of-lipid-mediators-in-the-kidneys-and-t-cells-of-mrllpr-lupus-prone-mice/