Major vault protein in the pathogenesis of tubulo-interstitial inflammation and fibrosis in chronic kidney disease

Abstract

Chronic kidney disease (CKD) is a major global health challenge that results in considerable morbidity and mortality. Irrespective of original aetiology, progressive tubulo-interstitial fibrosis drives CKD progression to end-stage kidney disease. There is no effective treatment for kidney fibrosis. The mechanisms that mediate tubulo-interstitial fibrosis are complex and incompletely understood. Understanding the mechanisms is a prerequisite for the development of novel therapeutic options. Major vault protein (MVP) is a key component of an intracellular multi-subunit structure called vaults, which are comprised of ribonucleoprotein particles. MVP has been implicated in various cellular processes such as innate immunity, cell differentiation and signal transduction. The role of MVP in organ fibrosis has not been investigated. In this study, MVP expression was found to be markedly increased in kidney biopsies from patients with acute kidney injury or CKD due to various aetiologies; and the increased MVP expression was predominantly localized to proximal tubular epithelial cells (PTEC). Results from experiments using murine models of tubulo-interstitial fibrosis induced by unilateral ureteral obstruction (UUO), or CKD induced by adenine-induced nephropathy, showed that kidney injuries thus induced were associated with increased MVP expression and histopathologic features of tubular atrophy, macrophage infiltration, and expression of mediators of inflammation and fibrosis including PI3K/AKT, CCL2, CCL5, TNF-α, IL-6, VCAM-1, α-smooth muscle actin, fibronectin, and collagen I. Some macrophages in the murine CKD model also co-expressed collagen I, fibronectin, and α-smooth muscle actin, suggesting macrophage-to-myofibroblast transition. Ultrastructural studies also demonstrated podocyte injury, which is a key feature in progressive CKD. MVP knockout attenuated the abnormalities induced by UUO or adenine in the murine models, including the severity of podocyte injury. In cultured HK-2 cells, addition of TNF-α, but not IL-1β, IL-6, IL-8, or TGF-β1, induced MVP expression, which was mediated in part through increased AKT and JNK phosphorylation. TNF-α also increased CCL2, CCL5, IL-6 and IL-8 secretion and fibronectin expression in HK-2 cells, mediated in part through increased AKT phosphorylation. In contrast, HK-2 cells deficient in MVP, generated by CRISPR/Cas9, showed significantly less AKT phosphorylation and associated chemokine and cytokine secretion and fibronectin expression, upon exposure to TNF-α. Also, when exposed to TGF-β1 which is a key mediator of kidney fibrosis, MVP-deficient HK-2 cells showed markedly reduced fibronectin deposition induced by TGF-β1 compared with non-transfected cells, while HK-2 cells over-expressing MVP showed markedly increased fibronectin deposition in the extracellular matrix. In summary, the results from this project showed that MVP expression is increased in patients, or mice, with CKD irrespective of aetiologies, and suggested that MVP is involved in the pathogenic mechanisms leading to tubulo-interstitial inflammation and fibrosis, mediated through increased PI3K/AKT phosphorylation, increased secretion of pro-fibrotic chemokines, macrophage recruitment and its transition to a pro-fibrotic phenotype, and increased fibronectin and collagen deposition.