Current treatment for lupus nephritis : direct cellular effects that impact on kidney inflammation fibrosis

Abstract

Lupus nephritis (LN) is an important cause of acute kidney injury and chronic kidney disease. Reducing acute immune-mediated kidney injury and preventing kidney fibrosis are important in ensuring optimal long-term kidney and patient survival. Treatment of LN is challenging since there is considerable individual variation in clinical presentation and response to treatment. The current standard-of-care treatment for LN patients includes combined use of an immunosuppressive agent and glucocorticoid. Mycophenolate (MPA), azathioprine (AZA), rapamycin (RAPA) and tacrolimus (TAC) are effective medications. Animal and cell studies have also shown that these agents have anti-inflammatory and/or anti-fibrotic effects on resident kidney cells, independent of their immunosuppressive properties. Details of cellular pathogenic pathways in LN progression, and the effects of MPA, AZA, RAPA and TAC on these pathways, remain obscure. Experiments in this project showed that in NZB/W F1 mice, LN progression was characterized by histopathological features including mesangial expansion, glomerulosclerosis, tubular dilation and atrophy, protein cast formation and mononuclear cell infiltration, and increased expression of mediators of kidney injury, inflammation and fibrosis including havcr1, lcn2, il1b, il6, ccl2, acta2, fn1, col1a1 and col3a1. RNA-Seq analysis of kidney cortical tissue from NZB/W F1 mice showed that active nephritis, compared with pre-disease mice, was associated with downregulation of mitochondrial-mediated energy metabolism-related pathways including fatty acid oxidation, ATP metabolic process and TCA cycle. Active LN was also accompanied by enrichment in gene expression encoding for cellular senescence, PI3K-Akt, Hippo and proximal tubule bicarbonate reclamation pathways. MPA-treated mice showed a significant reduction in gene expression profile related to kidney injury, inflammation, and fibrosis including the Hippo pathway. In contrast, RAPA-, AZA- and TAC-treated mice showed only modest reduction in the expression of these genes. Also, MPA and RAPA, but not AZA or TAC, suppressed cellular senescence-related genes in NZB/W F1 mice. All four immunosuppressive agents upregulated genes involved in cell metabolism and bicarbonate reclamation pathways, and downregulated genes involved in PI3K/Akt signaling. In proximal tubular epithelial cells (PTEC), TGF-β1, a key fibrotic mediator, induced actin polymerization, and activation of Hippo and SMAD2/3 signaling with increased α-smooth muscle actin, fibronectin and collagen expression. RNA-Seq analysis demonstrated that TGF-β1 induced genes enriched for ECM assembly, ECM-receptor interaction, focal adhesion, PI3K/Akt and Hippo pathways, suggesting that alteration in these pathways detected in the kidney cortex of NZB/W F1 mice was attributed, at least in part, to PTEC injury. MPA significantly downregulated genes enriched in these pathways that were induced by TGF-β1, while RAPA, AZA and TAC showed minimal effect. Experiments using immunofluorescence staining, luciferase assays and subcellular fractionation showed that MPA exerted its anti-fibrotic effect on PTEC through inhibition of TGF-β1-induced actin cytoskeleton remodeling, inactivation of the Hippo pathway through cytosolic YAP retention, and decreased SMAD3 phosphorylation. In summary, the results from this study demonstrated that active LN is associated with alterations in pathways regulating renal tubular cell metabolism, ECM organization, cellular senescence and bicarbonate reclamation, and MPA, in contrast to other immunosuppressive agents such as AZA, TAC and RAPA, has a more pronounced effect in suppressing these pathogenic processes.