Effect of mycophenolate and rapamycin on renal fibrosis in lupus nephritis.

Lupus nephritis (LN) leads to chronic kidney disease (CKD) through progressive fibrosis. Mycophenolate inhibits inosine monophosphate dehydrogenase and is a standard treatment for LN. The mammalian or mechanistic target of rapamycin (mTOR) pathway is activated in LN. Rapamycin inhibits mTOR and is effective in preventing kidney transplant rejection, with the additional merits of reduced incidence of malignancies and viral infections. The effect of mycophenolate or rapamycin on kidney fibrosis in LN has not been investigated. We investigated the effects of mycophenolate and rapamycin in New Zealand Black and White first generation (NZB/W F1) murine LN and human mesangial cells (HMCs), focusing on mechanisms leading to kidney fibrosis. Treatment of mice with mycophenolate or rapamycin improved nephritis manifestations, decreased anti-double stranded (ds) DNA antibody titer and reduced immunoglobulin G (IgG) deposition in the kidney. Both mycophenolate and rapamycin, especially the latter, decreased glomerular mTOR Ser2448 phosphorylation. Renal histology in untreated mice showed mesangial proliferation and progressive glomerulosclerosis with tubular atrophy, and increased expression of transforming growth factor ß1 (TGF-ß1), monocyte chemoattractant protein-1 (MCP-1), α-smooth muscle actin (α-SMA), fibronectin (FN) and collagen. Both mycophenolate and rapamycin ameliorated the histopathological changes. Results from in vitro experiments showed that both mycophenolate and rapamycin decreased mesangial cell proliferation and their binding with anti-dsDNA antibodies. Mycophenolate and rapamycin also down-regulated mTOR and extracellular signal-regulated kinase (ERK) phosphorylation and inhibited fibrotic responses in mesangial cells that were induced by anti-dsDNA antibodies or TGF-ß1. Our findings suggest that, in addition to immunosuppression, mycophenolate and rapamycin may reduce fibrosis in LN, which has important implications in preventing CKD in patients with LN.

Binding of anti-dsDNA antibodies to proximal tubular epithelial cells contributes to renal tubulointerstitial inflammation.

Immune deposits are often observed along the tubular basement membrane in patients with lupus nephritis, but the role of anti-dsDNA antibody (Ab) deposition on tubulointerstitial inflammation remains to be investigated. We examined the effect of human polyclonal anti-dsDNA Abs on inflammatory processes in cultured proximal renal tubular epithelial cells (PTEC, HK-2 cells) and their association with serum levels of interleukin (IL)-6, IL-8 and monocyte chemoattractant protein-1 (MCP-1) in patients. Binding of anti-dsDNA Abs to HK-2 cells was investigated by cellular ELISA, flow cytometry and immunohistochemistry. IL-6, IL-8 and MCP-1 secretion, mitogen-activated protein kinase (MAPK) activation and the effect of mycophenolic acid (MPA) were investigated by ELISAs and Western blot analysis. NZBWF1/J mice with active nephritis were randomized to receive either mycophenolate mofetil (MMF) (100 mg/kg per day) or vehicle for up to 12 weeks to study renal histopathology focusing on tubulointerstitial changes. Our results demonstrated that anti-dsDNA Abs bound to HK-2 cell surface and induced IL-6, IL-8 and MCP-1 secretion through distinct MAPK pathways. MPA inhibited anti-dsDNA Ab binding to HK-2 cells and suppressed apical and basolateral IL-6 and IL-8, but not MCP-1, secretion. Anti-dsDNA Ab level correlated with serum and tubulointerstitial expression of IL-6, IL-8 and MCP-1. MMF treatment in NZBWF1/J mice reduced anti-dsDNA Ab production and MAPK activation in the renal tubulointerstitium, together with decreased IL-6 and MCP-1 expression. Our data demonstrate that anti-dsDNA Abs contribute to inflammatory processes in the tubulointerstitium in lupus nephritis through their binding to proximal renal tubular epithelial cells and induction of pro-inflammatory mediators, and MPA ameliorates anti-dsDNA Ab induced IL-6 and IL-8 secretion in these cells.

Effect of Combined Mycophenolate and Rapamycin Treatment on Kidney Fibrosis in Murine Lupus Nephritis.

Background: A significant proportion of lupus nephritis patients develop chronic kidney disease (CKD) and progressive kidney fibrosis, for which there is no specific treatment. We previously reported that mycophenolate or rapamycin monotherapy showed comparable efficacy in suppressing kidney fibrosis in a murine model of lupus nephritis through their direct action on mesangial cells. We extended our study to investigate the effect of combined mycophenolate and rapamycin treatment (MR) on kidney fibrosis in NZBWF1/J mice. Methods: Female NZBWF1/J mice with active nephritis were randomized to receive vehicle or treatment with mycophenolate (50 mg/kg/day) and rapamycin (1.5 mg/kg/day) (MR) for up to 12 weeks, and the effect of treatment on clinical parameters, kidney histology, and fibrotic processes was investigated. Results: Progression of nephritis in untreated mice was accompanied by mesangial proliferation, glomerulosclerosis, tubular atrophy, protein cast formation, increased mTOR and ERK phosphorylation, and induction of TGF-ß1, IL-6, α-smooth muscle actin, fibronectin, and collagen expression. Combined MR treatment prolonged survival, improved kidney function, decreased anti-dsDNA antibody level, and ameliorated histopathological changes. The effect of combined MR treatment on kidney histology and function was comparable to that of mycophenolate or rapamycin monotherapy. In vitro studies in human mesangial cells showed that exogenous TGF-ß1 and IL-6 both induced mTOR and ERK phosphorylation and downstream fibrotic processes. Both mycophenolic acid and rapamycin inhibited inflammatory and fibrotic processes induced by TGF-ß1 or IL-6 by downregulating mTOR and ERK phosphorylation. Conclusions: Our findings indicate that combined mycophenolate and rapamycin, at reduced dose, improves kidney fibrosis in murine lupus nephritis through their distinct effect on mTOR and ERK signaling in mesangial cells.