The Angio-Fibrotic Switch of VEGF and CTGF in Proliferative Diabetic Retinopathy

BACKGROUND: In proliferative diabetic retinopathy (PDR), vascular endothelial growth factor (VEGF) and connective tissue growth factor (CTGF) cause blindness by neovascularization and subsequent fibrosis, but their relative contribution to both processes is unknown. We hypothesize that the balance between levels of pro-angiogenic VEGF and pro-fibrotic CTGF regulates angiogenesis, the angio-fibrotic switch, and the resulting fibrosis and scarring. METHODS/PRINCIPAL FINDINGS: VEGF and CTGF were measured by ELISA in 68 vitreous samples of patients with proliferative DR (PDR, N = 32), macular hole (N = 13) or macular pucker (N = 23) and were related to clinical data, including degree of intra-ocular neovascularization and fibrosis. In addition, clinical cases of PDR (n = 4) were studied before and after pan-retinal photocoagulation and intra-vitreal injections with bevacizumab, an antibody against VEGF. Neovascularization and fibrosis in various degrees occurred almost exclusively in PDR patients. In PDR patients, vitreous CTGF levels were significantly associated with degree of fibrosis and with VEGF levels, but not with neovascularization, whereas VEGF levels were associated only with neovascularization. The ratio of CTGF and VEGF was the strongest predictor of degree of fibrosis. As predicted by these findings, patients with PDR demonstrated a temporary increase in intra-ocular fibrosis after anti-VEGF treatment or laser treatment. CONCLUSIONS/SIGNIFICANCE: CTGF is primarily a pro-fibrotic factor in the eye, and a shift in the balance between CTGF and VEGF is associated with the switch from angiogenesis to fibrosis in proliferative retinopathy

Regulation of Fibronectin-EDA through CTGF Domain–Specific Interactions with TGFβ2 and Its Receptor TGFβRII

Fibronectin containing extra domain A (FN-EDA) is highly expressed in PVR membranes. The authors show that connective tissue growth factor promotes FN-EDA expression induced by transforming growth factor–β through direct protein interactions and complex regulatory mechanisms

TGF-β–Stimulated CTGF Production Enhanced by Collagen and Associated with Biogenesis of a Novel 31-kDa CTGF Form in Human Corneal Fibroblasts

Evidence for a novel cell-associated 31-kDa CTGF fragment that is produced by human corneal fibroblasts and regulated by collagen

Connective tissue growth factor antagonizes transforming growth factor-β1/Smad signalling in renal mesangial cells

International audienceThe critical involvement of transforming growth factor-β1 (TGF-β1) in diabetic nephropathy (DN) is well established, however the role of connective tissue growth factor (CTGF) in regulating the complex interplay of TGF-β1 signalling networks is poorly understood. The purpose of this study was to investigate co-operative signaling between CTGF and TGF-β1 and its physiological significance. CTGF was determined to bind directly to TβRIII 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-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 crosstalk with CTGF emphasizing the potential utility of targeting CTGF in DN

Connective tissue growth factor antagonizes transforming growth factor-?1/Smad signalling in renal mesangial cells

The critical involvement of TGF-?1 (transforming growth factor-?1) in DN (diabetic nephropathy) is well established. However, the role of CTGF (connective tissue growth factor) in regulating the complex interplay of TGF-?1 signalling networks is poorly understood. The purpose of the present study was to investigate co-operative signalling between CTGF and TGF-?1 and its physiological significance. CTGF was determined to bind directly to the T?RIII (TGF-? type III receptor) and antagonize TGF-?1-induced Smad phosphorylation and transcriptional responses via its N-terminal half. Furthermore, TGF-?1 binding to its receptor was inhibited by CTGF. A consequent shift towards non-canonical TGF-?1 signalling and expression of a unique profile of differentially regulated genes was observed in CTGF/TGF-?1-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 T?RIII restored TGF-?1-mediated Smad signalling and cell contractility, suggesting that T?RIII is key for CTGFmediated regulation of TGF-?1. Comparison of gene expression profiles from CTGF/TGF-?1-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-?1 are regulated by cross-talk with CTGF, emphasizing the potential utility of targeting CTGF in DN

Inhibition of TRPC6 channels ameliorates renal fibrosis and contributes to renal protection by soluble klotho

Fibrosis is an exaggerated form of tissue repair that occurs with serious damage or repetitive injury and ultimately leads to organ failure due to the excessive scarring. Increased calcium ion entry through the TRPC6 channel has been associated with the pathogenesis of heart and glomerular diseases, but its role in renal interstitial fibrosis is unknown. We studied this by deletion of Trpc6 in mice and found it decreased unilateral ureteral obstruction-induced interstitial fibrosis and blunted increased mRNA expression of fibrosis-related genes in the ureteral obstructed kidney relative to that in the kidney of wild-type mice. Administration of BTP2, a pyrazol derivative known to inhibit function of several TRPC channels, also ameliorated obstruction-induced renal fibrosis and gene expression in wild-type mice. BTP2 inhibited carbachol-activated TRPC3 and TRPC6 channel activities in HEK293 cells. Ureteral obstruction caused over a 10-fold increase in mRNA expression for TRPC3 as well as TRPC6 in the kidneys of obstructed relative to the sham-operated mice. The magnitude of protection against obstruction-induced fibrosis in Trpc3 and Trpc6 double knockout mice was not different from that in Trpc6 knockout mice. Klotho, a membrane and soluble protein predominantly produced in the kidney, is known to confer protection against renal fibrosis. Administration of soluble klotho significantly reduced obstruction-induced renal fibrosis in wild-type mice, but not in Trpc6 knockout mice, indicating that klotho and TRPC6 inhibition act in the same pathway to protect against obstruction-induced renal fibrosis. Thus klotho and TRPC6 may be pharmacologic targets for treating renal fibrosis.Fil: Wu, Yueh-Lin. University of Texas Southwestern Medical Center; Estados Unidos. Taipei Medical University Hospital; China. Taipei Medical University; ChinaFil: Xie, Jian. University of Texas Southwestern Medical Center; Estados UnidosFil: An, Sung-Wan. University of Texas Southwestern Medical Center; Estados UnidosFil: Oliver, Noelynn. Boehringer Ingelheim Pharmaceuticals Inc.; Estados UnidosFil: Barrezueta, Nestor X.. Boehringer Ingelheim Pharmaceuticals Inc.; Estados UnidosFil: Lin, Mei-Hsiang. Taipei Medical University; ChinaFil: Birnbaumer, Lutz. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Pontificia Universidad Católica Argentina ; Argentina. Research Triangle Park; Estados UnidosFil: Huang, Chou-Long. University of Texas Southwestern Medical Center; Estados Unido

Temporal expression profile and distribution pattern indicate a role of connective tissue growth factor (CTGF/CCN-2) in diabetic nephropathy in mice

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 parameters of diabetes and DN were analyzed in 10-mo-old diabetic ob/ob mice and their ob/+ littermates. STZ-induced diabetic mice showed a significantly increased urinary albumin excretion after 1 wk and increased mesangial matrix score after 2 wk. Increased renal fibronectin, fibronectin ED-A, and collagen IValpha1 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 mRNA was also increased in the heart and liver. In contrast, transforming growth factor-beta1 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 D

Renal proximal tubular dysfunction is a major determinant of urinary connective tissue growth factor excretion.

Contains fulltext : 89432.pdf (publisher's version ) (Open Access)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 NH(2)-terminal fragment, since the NH(2)-fragment is the predominant form detected in urine. Renal CTGF extraction, determined by simultaneous arterial and renal vein sampling, was 18 +/- 3% for full-length CTGF and 21 +/- 1% for the NH(2)-fragment. Fractional excretion was very low for both CTGFs (0.02 +/- 0.006% and 0.10 +/- 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(2)-microglobulin excretion (r = 0.99). Furthermore, urinary CTGF correlated with beta(2)-microglobulin (r = 0.85) in renal disease patients (n = 108), and only beta(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.1 juni 201