CTGF and lymphangiogenesis in renal and peritoneal fibrosis

Long term peritoneal dialysis (PD) treatment results in submesothelial fibrosis, neoangiogenesis, and ultrafiltration failure (UFF). Peritoneal lymphangiogenesis with increased lymphatic absorption is very likely to contribute to UFF in peritoneal fibrosis.

We generated two non-bacterial rat models, a scraping model and a zymosan injection model, which are useful for investigating the pathophysiology of fibrosis, neoangiogenesis, and lymphangiogenesis triggered by peritonitis-induced inflammation.

We found that transforming growth factor-b (TGF-b) increased expression of an important lymphangiogenic growth factor, vascular endothelial growth factor (VEGF)-C in peritoneal mesothelial cells. Inhibition of TGF-b signaling reduced VEGF-C expression and lymphangiogenesis in parallel with reduction of fibrosis and inflammation, and that treatment with soluble VEGF receptor (VEGFR)-3 specifically suppressed lymphangiogenesis and improved impaired ultrafiltration in a peritoneal fibrosis model. Blocking of VEGFR-3 signaling might be a useful strategy to improve lymphangiogenesis-dependent UFF.

Connective tissue growth factor (CTGF/CCN2), an important determinant of fibrotic tissue remodeling, was also associated with VEGF-C expression and lymphangiogenesis in human PD effluents, human peritoneal biopsies, and in a rat peritoneal fibrosis model, and a positive correlation between CTGF and VEGF-C enhancement was observed in peritoneal mesothelial cells treated with TGF-b1, implying a possible novel role for CTGF in PD-related peritoneal lymphangiogenesis.

Lymphangiogenesis is also associated with tubulointerstitial fibrosis in kidney diseases. We observed that TGF-b, VEGF-C, and lymphatic vessels were time-dependently increased in a rat unilateral ureteral obstruction (UUO) model, and that TGF-b increased VEGF-C production in cultured renal tubular cells and macrophages, while inhibition of TGF-b signaling reduced VEGF-C expression and lymphatic vessel formation in the UUO model, similar to our findings in the peritoneal fibrosis model. Furthermore, CTGF knockdown reduced VEGF-C expression and decreased lymphangiogenesis in parallel with reduction of fibrosis in the mouse UUO model, while CTGF increased VEGF-C production in renal tubular cells. On the other hand, CTGF could directly bind to VEGF-C, and thereby suppress VEGF-C-induced lymphatic endothelial cells growth, but this direct inhibitory effect of CTGF on VEGF-C was abrogated by cleavage of CTGF. Thus, we identified a role for CTGF in renal lymphangiogenesis both through the regulation of VEGF-C production and activity.

We also evaluated the effect of aging on tissue response to kidney injury, and found that UUO kidneys of old mice showed less interstitial fibrosis compared to young mice. This was associated with increased bone morphogenetic protein (BMP) 6 and decreased CTGF expression. Our further observations that CTGF directly bound to BMP6 and suppressed BMP6 downstream signaling, suggested that the renoprotective effects of preserved BMP6 expression might be further upregulated by the lower CTGF expression in old UUO kidneys might be responsible for the less fibrotic response in older kidneys.

Finally, CTGF is also an important mediator of renal allograft fibrosis and we identified tubulointerstitial CTGF [removed]CTGFti) at 3 months as an independent predictor of interstitial fibrosis (IF) and tubular atrophy (TA) at 5 years after transplantation in stable renal transplant recipients, and that CTGFti and urinary CTGF levels at 3 months when added to donor age improved prediction of later IF/TA development.