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Connective tissue growth factor (CTGF) plays a key role in renal fibrosis. Urinary CTGF is elevated in various renal diseases and may have biomarker potential. However, it is unknown which processes contribute to elevated urinary CTGF levels. Thus far, urinary CTGF was considered to reflect renal expression. We investigated how tubular dysfunction affects urinary CTGF levels. To study this, we administered recombinant CTGF intravenously to rodents. We used both full-length CTGF and the N[H.sub.2]-terminal fragment, since the N[H.sub.2]-fragment is the predominant form detected in urine. Renal CTGF extraction, determined by simultaneous arterial and renal vein sampling, was 18 [+ or -] 3% for full-length CTGF and 21 [+ or -] 1% for the N[H.sub.2]-fragment. Fractional excretion was very low for both CTGFs (0.02 [+ or -] 0.006% and 0.10 [+ or -] 0.02%, respectively), indicating that >99% of the extracted CTGF was metabolized by the kidney. Immunohistochemistry revealed extensive proximal tubular uptake of CTGF in apical endocytic vesicles and colocalization with megalin. Urinary CTGF was elevated in megalin-and cubilin-deficient mice but not in cubilin-deficient mice. Inhibition of tubular reabsorption by Gelofusine reduced renal uptake of CTGF and increased urinary CTGF. In healthy volunteers, Gelofusine also induced an increase of urinary CTGF excretion, comparable to the increase of [[beta].sub.2]-microglobulin excretion (r = 0.99). Furthermore, urinary CTGF correlated with [[beta].sub.2]-microglobulin (r = 0.85) in renal disease patients (n = 108), and only [[beta].sub.2]-microglobulin emerged as an independent determinant of urinary CTGF. Thus filtered CTGF is normally reabsorbed almost completely in proximal tubules via megalin, and elevated urinary CTGF may largely reflect proximal tubular dysfunction. proximal tubular reabsorption; biomarker; megalin doi: 10.1152/ajprenal.00694.2009. more...

Peritoneal fibrosis (PF) is an important complication of peritoneal dialysis (PD) therapy that often occurs in association with peritoneal high transport rate and ultrafiltration failure (UFF). To study the possible pathogenic role of connective tissue growth factor (CTGF) in the relationship of PF and UFF, dialysate CTGF contents (n = 178) and tissue CTGF expression (n = 61) were investigated by ELISA, real-time PCR, immunohistochemistry, and in situ hybridization. CTGF production with and without TGF-[[beta].sub.1] stimulation in human peritoneal mesothelial cells (HPMC) from the spent patients' peritoneal dialysate (n = 32) was studied in vitro. The dialysate-to-plasma ratio for creatinine (D/P Cr) was positively conelated to dialysate CTGF concentration and estimated local peritoneal production of CTGF. CTGF mRNA expression was 11.4-fold higher in peritoneal membranes with UFF than in pre-PD renal failure peritoneum and was correlated with thickness of the peritoneum. CTGF protein and mRNA were detected in mesothelium and in fibroblast-like cells. In cultured HPMC, TGF-[[beta].sub.1]-induced expression of CTGF mRNA was increased at 12 and 24 h and was correlated with D/P Cr. In contrast, bone morphogenic protein-4 mRNA expression was inversely correlated with D/P Cr. Our results suggest that high peritoneal transport state is associated with fibrosis and increased peritoneal CTGF expression and production by mesothelial ceils, which can be stimulated by TGF-[[beta].sub.1]. Dialysate CTGF concentration could be a biomarker for both peritoneal fibrosis and membrane function. Functional alteration of mesothelial cells may be involved in progression of peritoneal fibrosis in high transport state. TGF-[beta]; BMP-4; BMP-7; fibrosis; ultrafiltratiou failure doi:10.1152/ajprenal.00368.2009 more...

Connective tissue growth factor (CTGF) is overexpressed in diabetic nephropathy (DN) and has therefore been implicated in its pathogenesis. The objective of the present study was to determine the tissue distribution of increased CTGF expression and the relationship of plasma, urinary, and renal CTGF levels to the development and severity of DN. We studied the relationship between CTGF and renal pathology in streptozotocin (STZ)-induced diabetes in C57BL/6J mice. Diabetic and age-matched control mice were killed after 1,2, 4, and 9 wk of diabetes. In addition, key paraineters of diabetes and DN were analyzed in 10-mo-old diabetic ob/oh mice and their ob/+ littermates. STZ-induced diabetic mice showed ti significantly increased urinary albumin excretion after 1 wk and increased mesangial matrix score after 2 wk. Increased renal fibronectin, fibronectin ED-A, and collagen IV[[alpha].sub.1] expression, as well as elevated plasma creatinine levels, were observed after 9 wk. After 2 wk, CTGF mRNA was upregulated threefold in the renal cortex. By 9 wk, CTGF inRNA was also increased in the heart and liver. In contrast, transforming growth factor-[[beta].sub.1] mRNA content was significantly increased only in the kidney by 9 wk. Renal CTGF expression was mainly localized in podocytes and parietal glomerular epithelial cells, and less prominent in mesangial cells. In addition, plasma CTGF levels and urinary CTGF excretion were increased in diabetic mice. Moreover, albuminuria strongly correlated with urinary CTGF excretion (R = 0.83, P < 0.0001). increased CTGF expression was also demonstrated in type 2 diabetic ob/ob mice, which points to a causal relationship between diabetes and CTGF and thus argues against a role of STZ in this process. The observed relationship of podocyte and urinary CTGF to markers of DN suggests a pathogenic role of CTGF in the development of DN. diabetes: podocyte: fibrosis: kidney more...

Background: Connective tissue growth factor (CTGF) is widely thought to promote the development of fibrosis in collaboration with transforming growth factor (TGF)-β; however, most of the evidence for its involvement comes from correlative and culture-based studies. In this study, the importance of CTGF in tissue fibrosis was directly examined in three murine models of fibrotic disease: a novel model of multiorgan fibrosis induced by repeated intraperitoneal injections of CTGF and TGF-β2; the unilateral ureteral obstruction (UUO) renal fibrosis model; and an intratracheal bleomycin instillation model of pulmonary fibrosis. Results: Intraperitoneal coadministration of CTGF and TGF-β2 elicited a profound fibrotic response that was inhibited by the human anti-CTGF antibody FG-3019, as indicated by the ability of FG-3019 to ameliorate the histologic signs of fibrosis and reduce the otherwise increased hydroxyproline:proline (Hyp:Pro) ratios by 25% in kidney (P < 0.05), 30% in liver (P < 0.01) and 63% in lung (P < 0.05). Moreover, administration of either cytokine alone failed to elicit a fibrotic response, thus demonstrating that CTGF is both necessary and sufficient to initiate fibrosis in the presence of TGF-β and vice versa. In keeping with this requirement for CTGF function in fibrosis, FG- 3019 also reduced the renal Hyp:Pro response up to 20% after UUO (P < 0.05). In bleomycin-injured animals, a similar trend towards a FG-3019 treatment effect was observed (38% reduction in total lung Hyp, P = 0.056). Thus, FG-3019 antibody treatment consistently reduced excessive collagen deposition and the pathologic severity of fibrosis in all models. Conclusion: Cooperative interactions between CTGF and TGF-β signaling are required to elicit overt tissue fibrosis. This interdependence and the observed anti-fibrotic effects of FG-3019 indicate that anti-CTGF therapy may provide therapeutic benefit in different forms of fibroproliferative disease. [ABSTRACT FROM AUTHOR] more...

Klotho is a type-1 membrane protein predominantly produced in the kidney, the extracellular domain of which is secreted into the systemic circulation. Membranous and secreted Klotho protect organs, including the kidney, but whether and how Klotho directly protects the glomerular filter is unknown. Here, we report that secreted Klotho suppressed transient receptor potential channel 6 (TRPC6)-mediated Ca 2+ influx in cultured mouse podocytes by inhibiting phosphoinositide 3-kinase-dependent exocytosis of the channel. Furthermore, soluble Klotho reduced ATP-stimulated actin cytoskeletal remodeling and transepithelial albumin leakage in these cells. Overexpression of TRPC6 by gene delivery in mice induced albuminuria, and exogenous administration of Klotho ameliorated the albuminuria. Notably, immunofluorescence and in situ hybridization revealed Klotho expression in podocytes of mouse and human kidney. Heterozygous Klotho-deficient CKD mice had aggravated albuminuria compared with that in wild-type CKD mice with a similar degree of hypertension and reduced clearance function. Finally, disrupting the integrity of glomerular filter by saline infusion-mediated extracellular fluid volume expansion increased urinary Klotho excretion. These results reveal a potential novel function of Klotho in protecting the glomerular filter, and may offer a new therapeutic strategy for treatment of proteinuria., (Copyright © 2016 by the American Society of Nephrology.) more...

The critical involvement of TGF-β1 (transforming growth factor-β1) in DN (diabetic nephropathy) is well established. However, the role of CTGF (connective tissue growth factor) in regulating the complex interplay of TGF-β1 signalling networks is poorly understood. The purpose of the present study was to investigate co-operative signalling between CTGF and TGF-β1 and its physiological significance. CTGF was determined to bind directly to the TβRIII (TGF-β type III receptor) and antagonize TGF-β1-induced Smad phosphorylation and transcriptional responses via its N-terminal half. Furthermore, TGF-β1 binding to its receptor was inhibited by CTGF. A consequent shift towards non-canonical TGF-β1 signalling and expression of a unique profile of differentially regulated genes was observed in CTGF/TGF-β1-treated mesangial cells. Decreased levels of Smad2/3 phosphorylation were evident in STZ (streptozotocin)-induced diabetic mice, concomitant with increased levels of CTGF. Knockdown of TβRIII restored TGF-β1-mediated Smad signalling and cell contractility, suggesting that TβRIII is key for CTGF-mediated regulation of TGF-β1. Comparison of gene expression profiles from CTGF/TGF-β1-treated mesangial cells and human renal biopsy material with histological diagnosis of DN revealed significant correlation among gene clusters. In summary, mesangial cell responses to TGF-β1 are regulated by cross-talk with CTGF, emphasizing the potential utility of targeting CTGF in DN. more...