Guochun Chen

Rapamycin Ameliorates Kidney Fibrosis by Inhibiting the Activation of mTOR Signaling in Interstitial Macrophages and Myofibroblasts

Interstitial fibrosis is an inevitable outcome of all kinds of progressive chronic kidney disease (CKD). Emerging data indicate that rapamycin can ameliorate kidney fibrosis by reducing the interstitial infiltrates and accumulation of extra cellular matrix (ECM). However, the cellular mechanism that regulates those changes has not been well understood yet. In this study, we revealed the persistent activation of mammalian target of rapamycin (mTOR) signaling in the interstitial macrophages and myofibroblasts, but rarely in injured proximal epithelial cells, CD4+ T cells, neutrophils, or endothelial cells, during the development of kidney fibrosis. Administration of rapamycin to unilateral ureteral obstruction (UUO) mice significantly suppressed the immunoreactivity of mTOR signaling, which decreased the inflammatory responses and ECM accumulation in the obstructed kidneys. Isolated macrophages from rapamycin-treated obstructed kidneys presented less inflammatory activity than vehicle groups. In vitro study confirmed that rapamycin significantly inhibited the fibrogenic activation of cultured fibroblasts (NIH3T3 cells), which was induced by the stimulation of TGF-β1. Further experiment revealed that rapamycin did not directly inhibit the fibrogenesis of HK2 cells with aristolochic acid treatment. Our findings clarified that rapamycin can ameliorate kidney fibrosis by blocking the mTOR signaling in interstitial macrophages and myofibroblasts.

A glimpse of matrix metalloproteinases in diabetic nephropathy.

Matrix metalloproteinases (MMPs) are proteolytic enzymes belonging to the family of zinc-dependent endopeptidases that are capable of degrading almost all the proteinaceous components of the extracellular matrix (ECM). It is known that MMPs play a role in a number of renal diseases, such as, various forms of glomerulonephritis and tubular diseases, including some of the inherited kidney diseases. In this regard, ECM accumulation is considered to be a hallmark morphologic finding of diabetic nephropathy, which not only is related to the excessive synthesis of matrix proteins, but also to their decreased degradation by the MMPs. In recent years, increasing evidence suggest that there is a good correlation between the activity or expression of MMPs and progression of renal disease in patients with diabetic nephropathy and in various experimental animal models. In such a diabetic milieu, the expression of MMPs is modulated by high glucose, advanced glycation end products (AGEs), TGF-β, reactive oxygen species (ROS), transcription factors and some of the microRNAs. In this review, we focused on the structure and functions of MMPs, and their role in the pathogenesis of diabetic nephropathy.

Transition of Mesothelial Cell to Fibroblast in Peritoneal Dialysis: EMT, Stem Cell or Bystander?

Long-term peritoneal dialysis (PD) can lead to fibrotic changes in the peritoneum, characterized by loss of mesothelial cells (MCs) and thickening of the submesothelial area with an accumulation of collagen and myofibroblasts. The origin of myofibroblasts is a central question in peritoneal fibrosis that remains unanswered at present. Numerous clinical and experimental studies have suggested that MCs, through epithelial-mesenchymal transition (EMT), contribute to the pool of peritoneal myofibroblasts. However, recent work has placed significant doubts on the paradigm of EMT in organ fibrogenesis (in the kidney particularly), highlighting the need to reconsider the role of EMT in the generation of myofibroblasts in peritoneal fibrosis. In particular, selective cell isolation and lineage-tracing experiments have suggested the existence of progenitor cells in the peritoneum, which are able to switch to fibroblast-like cells when stimulated by the local environment. These findings highlight the plastic nature of MCs and its contribution to peritoneal fibrogenesis. In this review, we summarize the key findings and caveats of EMT in organ fibrogenesis, with a focus on PD-related peritoneal fibrosis, and discuss the potential of peritoneal MCs as a source of myofibroblasts.

Lipopolysaccharide Induces Chronic Kidney Injury and Fibrosis through Activation of mTOR Signaling in Macrophages.

Background: Septic kidney injury is one of the most common complications in critically ill patients with a high risk of developing chronic kidney disease (CKD). Emerging data indicate that mammalian target of rapamyci (mTOR) signaling plays a major role in septic inflammation by regulating the immune response of macrophage. This study was designed to evaluate the role of mTOR signaling in kidney macrophages during endotoxemia-induced chronic kidney injury and subsequent fibrogenesis. Methods: Male C57BL/6 mice were used for all animal studies (n = 9 for each group). Lipopolysaccharide (LPS) was injected intraperitoneally (1 mg/kg) every 2 days to induce persistent endotoxemia. Rapamycin (1 mg/kg·day) was administered to a subgroup of mice 1 day prior to LPS treatment and continued to termination of the experiment. In ex-vivo experiment, RAW264.7 cells were cultured and treated with LPS (2 µg/ml) for 48 h while a subgroup of cells were incubated in the presence of rapamycin (50 nmol) for 2 h. Results: Continuous administration of LPS resulted in progressive macrophage infiltration, tubular injury and collagen deposition in mice kidneys. Rapamycin markedly ameliorated LPS-induced kidney pathological changes. Expression of pS6K was rarely observed in normal kidney macrophages, but significantly increased with time by LPS treatment. In ex-vivo study, LPS induced prominent production of IL-1β and MCP-1 in cultured RAW264.7 cells, which was significantly suppressed by rapamycin. Conclusion: Taken together, our findings show that endotoxemia results in activation of mTOR signaling in macrophages, leading to progressive kidney inflammatory injuries and subsequent fibrosis. Our study may reveal a mechanism involved in the development of sepsis-associated CKD and kidney fibrosis.

Changes in expression of four molecular marker proteins and one microRNA in mesothelial cells of the peritoneal dialysate effluent fluid of peritoneal dialysis patients

The aim of this study was to detect the expression of microRNA-200c and epithelial-mesenchymal transition (EMT) in the mesothelial cells of the peritoneal dialysate effluent fluid of peritoneal dialysis (PD) patients, and to investigate the association between microRNA-200c and peritoneal mesothelial cell EMT. Twelve patients who had recently started continuous ambulatory peritoneal dialysis (PD start group) and 16 patients who had been undergoing peritoneal dialysis for >6 months (PD >6 months group) were randomly chosen for the isolation, culture and identification of effluent cells. qPCR and western blot analysis were used to detect the expression levels of microRNA-200c and the levels of four cellular marker proteins, E-cadherin, vimentin, fibronectin (FN) and COL-1, in effluent cells. The results showed that the effluent cells in peritoneal dialysis were peritoneal mesothelial cells. The level of E-cadherin protein expression was significantly lower in the PD >6 months group than in the PD start group, while vimentin, FN and COL-1 protein expression levels were significantly increased in the PD >6 months group. microRNA-200c in the PD >6 months group was significantly downregulated. The E-cadherin protein expression level was significantly decreased and vimentin, FN and COL-1 protein expression levels were significantly increased in the PD >6 months group. The level of microRNA-200c was significantly reduced in the PD > 6 months group, suggesting that microRNA-200c may be associated with EMT.

TLR9 and BAFF: their expression in patients with IgA nephropathy.

Since it was first described in 1968, immunoglobulin (Ig)A nephropathy (IgAN) has become the most commonly diagnosed form of primary glomerular disease worldwide. A number of reports have shown that toll‑like receptor 9 (TLR9) and B‑cell activating factor (BAFF) may be associated with IgAN; however, sufficient evidence has not yet to be delivered. In the present study, serum levels of BAFF as well as TLR9 mRNA and protein levels in peripheral blood mononuclear cells (PBMCs) were assessed. Expression of TLR9 mRNA in PBMCs was examined by quantitative polymerase chain reaction and the TLR9 protein was determined by western blot analysis. The levels of serum BAFF and IgA1 were determined by specific ELISA. Serum levels of BAFF and IgA1 as well as levels of TLR9 mRNA and protein in PMBCs were significantly higher in patients with IgAN compared with patients with minimal glomerular abnormalities (P<0.05, P<0.01, P<0.01 and P<0.01, respectively) and normal controls (P<0.01, P<0.01, P<0.05 and P<0.01, respectively). A correlation and regression analysis was performed to determine the pathogenesis of IgAN. In patients with IgAN, serum levels of BAFF were positively correlated with IgA1 levels (rp, 0.515; P<0.01) and mesangial IgA deposition density (rp, 0.746; P<0.01). Expression levels of TLR9 protein in PBMCs of IgAN patients were positively correlated with levels of serum BAFF (rp, 0.444; P<0.05) and IgA1 (rp, 0.633; P<0.01). These results suggested that overexpression of TLR9 mRNA and protein in PBMCs and elevated levels of serum BAFF may be associated with overexpression of serum IgA1, and, furthermore, may have a role in the development of IgAN.

Aged rats are susceptible to nephrotoxicity induced by iodinated contrast media.

Objective: The objective of this study is to evaluate the effect and mechanism of aging on iodinated-contrast-media-induced nephropathy in male rats. Methods: Twenty-four healthy male rats were initially divided into 12-month-old and 24-month-old age groups (adult and older age groups, respectively; n = 12/group); subsequently, each age group was randomly divided into saline control (NS) and contrast media (CM) groups (n = 6/group). CM (76% diatrizoate, 10 mL/kg b.w.) was given through the caudal vein. Urinary creatinine (Ucr) and serum creatinine (Scr) were detected by an automatic biochemical analyzer. The activities of renal malondialdehyde (MDA), superoxide dismutase (SOD), angiotensin-converting enzyme (ACE), angiotensin II (Ang II), and reduced form of nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase) were determined by spectrophotometric assays with commercially available kits according to the manufacturers’ protocols. Renal histological changes were observed by hematoxylin and eosin staining and scored semiquantitatively. Results: In diatrizoate-injected aged rats, Scr, the activities of ACE, Ang II, MDA, and NADPH oxidase in renal tissues were significantly increased (p < 0.01). The histologic scores were higher in the aged animals with CM treatment than those of control or adult rats (p < 0.01). There was an increasing trend but no significant statistical difference in renal ACE, Ang II, MDA, and NADPH oxidase or histologic scores in adult CM-injected rats compared with control animals (p > 0.05). Conclusions: Older age is an aggravating factor of iodinated-contrast-media-induced nephropathy in male rats. Oxidative stress and the renin–angiotensin system (RAS) may play an important role in nephrotoxicity induced by iodinated contrast media, especially in aged male rats.

A glimpse of the glomerular milieu: From endothelial cell to thrombotic disease in nephrotic syndrome☆

Patients with nephrotic syndrome (NS) carry a high risk of venous thromboembolism (VTE) due to the abnormalities in coagulation and fibrinolysis. Although massive urine protein loss is considered to trigger the cascade of hypercoagulation, the exact nature of VTE in NS patients still remains obscure, especially in some cases when VTE occurs far before the presence of nephrotic proteinuria. Recent findings illustrate that loss of local glomerular homeostasis, like disturbance of cytokine profiles in endothelial cells or aberrant cellular crosstalks in glomerulus, is sufficient to initiate the development of thrombotic disease in glomerulonephropathy. Emerging data have highlighted the glomerular endothelial cell as a key regulator of local homeostasis, which might mediate the haemostatic derangement in the beginning of glomerular disease by expression of numerous prothrombotic factors and result in the subsequent predilection of VTE in NS. As the glomerulus-derived circulating factors are all collected and flushed into the renal vein directly, it is reasonable to suggest that increased release of glomerulus-derived thrombotic regulators, particularly from endothelial cells, may play a significant role in the highest proclivity for the renal vein as the site of thrombosis in NS. In this review, we thus discuss the current understandings of thromboembolism in NS with focus on how the glomerular endothelial cell involves in the pathogenesis of VTE, which may help to increase our understandings in the anti-thrombotic therapy for patients with NS.

New insights into the pathogenesis and treatment of peritoneal fibrosis: A potential role of Wnt/β-catenin induced epithelial to mesenchymal transition and stem cells for therapy

Peritoneal fibrosis is a chronic, progressive progress, which is associated with ultrafiltration failure. In the development of peritoneal fibrosis, Epithelial to mesenchymal transition is an important cellular process whereby epithelial cells transform into mesenchymal cells under physiology and pathology conditions, along with change of cell morphology and expression of related genes. It plays an important role in embryogenesis and development of tissues and organs, as well as organ fibrosis and tumorigenesis. Several intracellular signal transduction pathways induce the process of Epithelial to mesenchymal transition. In recent researches, Wnt/β-catenin induced epithelial to mesenchymal transition was suggested to be an important reason for tissues and organs fibrosis. The following paper reviews the potential role of Wnt/β-catenin induced epithelial to mesenchymal transition in peritoneal fibrosis. New potential therapeutic interventions of peritoneal fibrosis are discussed.

High glucose-induced Galectin-1 in human podocytes implicates the involvement of Galectin-1 in diabetic nephropathy

The diabetic milieu is believed to change the activity, or result in damage of podocytes—a key component of the glomerular filtration barrier and known to secrete matrix for glomerular basement membrane. This in turn contributes to diabetic nephropathy. However, how podocyte dysfunction is triggered in diabetic nephropathy remains ambiguous. Galectin‐1 belongs to Galectin family that bind to β‐galactoside residues of glycosylated proteins. We explored whether Galectin‐1 is dysregulated in diabetic nephropathy using three different techniques, namely real‐time polymerase chain reaction, western blotting, and immunofluorescent staining, to follow the expression of Galectin‐1 under high glucose levels in podocytes. High glucose consistently induced Galectin‐1 expression. Immunohistochemistry using a Galectin‐1‐specific antibody also showed elevated Galectin‐1 in renal tissues of diabetic patients with manifestation of nephropathy, indicating a correlation of Galectin‐1 overexpression with diabetic nephropathy. Upregulation of Galectin‐1 is associated with loss of podocin, which is important for the physiological function of podocytes and decreases in the renal tissues of diabetic nephropathy. Increased Galectin‐1 is a causal event for the high glucose‐induced loss of podocin, since silencing Galectin‐1 in podocytes increased podocin expression in the presence of 25 mM glucose. Thus expression of Galectin‐1 in diabetic nephropathy may serve as a marker and contribute to disease progression by interfering with podocin expression.