Estrogen-induced Expression Of TWEAK Accelerates The Progression Of Lupus Nephritis

Objectives:Estrogens and tumor necrosis factor-like weak inducer of apoptosis(TWEAK) have been shown to play key roles in lupus nephritis(LN). However, the interaction between estrogens and TWEAK remains unclear in LN. The aim of this study was to investigate the roles and mechanisms of 17β-estradiol(E2) in TWEAK expression in LN.Methods:1. In vitro study, peripheral blood mononuclear cells(PBMCs) were isolated from the peripheral blood of LN patients.The cells were divided equally into four groups and treated for 24 hours with either one of the followings: 1 × 10-5 mmol/L E2;1 × 10-5 mmol/L E2 combined with 1 × 10-4 mmol/L ICI 182,780(ER-selective antagonist);1 × 10-5 mmol/L E2 with 1 × 10-2 mmol/L methyl-piperidino-pyrazole(MPP, ERα-selective modulator) or DMSO vehicle alone. QPCR was used to analyse the expression levels of TWEAK mRNA.2. Female MRL/lpr and MRL/MpJ mice were used for in vivo studies. Ovariectomy was performed on 12-week-old MRL/lpr female mice and sham-operation group were just removed the fat issues around the ovary. After one week, the MRL/lpr mice were divided into seven groups. In sham-operaion group, only DMSO was intraperitoneally injected twice per week. In the six ovariectomy groups, four groups were intraperitoneally injected twice per week with the followings: DMSO;5 ug E2; 5 ug E2 combined with 50 ug ICI 182,780 and 5 ug E2 with 50 ug MPP,respectively. In other two groups, E2 combined with 2 × 107 transducing units(TU) lentivirus(LV)-TWEAK-short hairpin RNA(TWEAK-shRNA group) or 2 × 107 TU LV-control-shRNA(control-shRNA group) were injected through the tail vain. The mice was sacrificed on day 8, 15 and 29 after the treatments. Real-time PCR and Western blot analyses were used to assess the mRNA and protein levels of renal TWEAK. Enzyme-linked immunosorbent assay(ELISA) was used to detedemine the serum IL-6 and anti-dsDNA levels. Hematoxylin and eosin(H&E), periodic acid-Schiff(PAS) and Masson’s trichrome staining were performed on renal tissue to assess renal damage.Results:1. In vitro study, we found that TWEAK mRNA expression in PBMCs was significantly up-regulated after E2 treatment and down-regulated after co-treatment with ICI 182,780 or MPP.2. In vivo study, compared with sham-operated MRL/lpr mice, ovariectomized mice treated with DMSO vehicle alone, showed lower expression levels of renal TWEAK mRNA and protein. The expression of both mRNA and protein in ovariectomized mice were upregulated after E2 treatment and downregulated after ICI 182,780 or MPP co-treatment. Moreover, severe renal damage and higher expression levels of renal TWEAK mRNA was observed in E2-treated ovariectomized mice, accompanied by higher serum levels of interleukin(IL)-6, compared with DMSO vehicle-treated ovariectomized mice. Co-treatment with LV-TWEAK-shRNA reversed these changes.Conclusion:1.E2 plays an important role in the upregulation of TWEAK expression in LN through an ER-dependent pathway and might aggravate the kidney damage in LN.2.TWEAK pathway may be a potential approach to the treatment of LN.