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Proliferation of interstitial fibroblasts is a hallmark of progressive renal fibrosis commonly resulting in chronic kidney failure. The intermediate-conductance [Ca.sup.2+]-activated [K.sup.+] channel ([K.sub.Ca]3.1) has been proposed to promote mitogenesis in several cell types and contribute to disease states characterized by excessive proliferation. Here, we hypothesized that [K.sub.Ca]3.1 activity is pivotal for renal fibroblast proliferation and that deficiency or pharmacological blockade of [K.sub.Ca]3.1 suppresses development of renal fibrosis. We found that mitogenic stimulation up-regulated [K.sub.Ca]3.1 in murine renal fibroblasts via a MEK-dependent mechanism and that selective blockade of [K.sub.Ca]3.1 functions potently inhibited fibroblast proliferation by [G.sub.0]/[G.sub.1] arrest. Renal fibrosis induced by unilateral ureteral obstruction (UUO) in mice was paralleled by a robust up-regulation of [K.sub.Ca]3.1 in affected kidneys. Mice lacking [K.sub.Ca]3.1 ([K.sub.Ca][3.1.sup.-/-]) showed a significant reduction in fibrotic marker expression, chronic tubulointerstitial damage, collagen deposition and [alpha][SMA.sup.+] cells in kidneys after UUO, whereas functional renal parenchyma was better preserved. Pharmacological treatment with the selective [K.sub.Ca]3.1 blocker TRAM-34 similarly attenuated progression of UUO-induced renal fibrosis in wild-type mice and rats. In conclusion, our data demonstrate that [K.sub.Ca]3.1 is involved in renal fibroblast proliferation and fibrogenesis and suggest that [K.sub.Ca]3.1 may represent a therapeutic target for the treatment of fibrotic kidney disease. ion channels | fibroblasts | kidney | organ fibrosis

Basic fibroblast growth factor (FGF-2) plays a role in renal fibrogenesis, although its potential implications for tubulointerstitial involvement in diabetic nephropathy are unknown. We evaluated the expression of FGF-2 in kidney biopsies from patients with diabetic nephropathy and studied the mechanisms of its induction in human renal fibroblasts under hyperglycemia. Tubulointerstitial expression of FGF-2 was significantly upregulated in diabetic nephropathy compared with control kidneys with a good correlation to the degree of the injury. Fibroblasts cultivated in high glucose displayed increased FGF-2 mRNA as well as protein synthesis and secretion compared with normal glucose. Proliferation rates under hyperglycemia were significantly higher and could be almost completely inhibited by addition of a neutralizing FGF-2 antibody. Alterations in proliferation were associated with changes in [p27.sup.kip1] expression. Hyperglycemia induced the expression of PKC-[beta]1 and PKC-[beta]2; however, only inhibition of PKC-[beta]1 but not PKC-[beta]2 led to a significant decrease of FGF-2 levels. Relevance of the culture findings and functional association was corroborated by colocalization of FGF-2 and PKC-[beta] in human diabetic kidneys in vivo. High glucose stimulated fibronectin synthesis and secretion, which could be substantially prevented by inhibition of PKC-[beta]1 and to a lesser extent by inhibiting the FGF-2. Expression of active phosphorylated form of p38 mitogen-activated protein kinase was upregulated under hyperglycemia; however, its inhibition had no effects on FGF-2 synthesis. Our results implicate a role of FGF-2 in high glucose-altered molecular signaling in pathogenesis of diabetic renal disease. tubulointerstitial involvement; fibronectin synthesis; protein kinase C; renal biopsy

Growing evidence suggests that a proportion of interstitial myofibroblasts detected during renal tubulointerstitial fibrosis originates from tubular epithelial cells by a process called epithelial-mesenchymal transition (EMT). The IL-6-type cytokine oncostatin M (OSM) has been recently implicated in the induction of EMT. We investigated OSM effects on the expression of both cell-cell contact proteins and mesenchymal markers and studied OSM-induced intracellular signaling mechanisms associated with these events in human proximal tubular cells. Human recombinant OSM attenuated the expression of N-cadherin, E-cadherin, and claudin-2 in human kidney-2 (HK-2) ceils associated with the induction of HK-2 cell scattering in 3D collagen matrices. Conversely, expression of collagen type I, vimentin, and S100A4 was induced by OSM. OSM-stimulated cell scattering was inhibited by antibodies against gp130. Besides inducing phosphorylation of Stat1 and Stat3, OSM led to a strong concentration--and time-dependent phosphorylation of the mitogen-activated protein kinases ERK1, ERK2, and ERK5. MEK1/2 inhibitor U0126 (10 [micro]M) blocked basal and OSM-induced ERK1/2 phosphorylation but not phosphorylation of either ERK5 or Stat1/3. Both synthetic MEK1/2 inhibitors U0126 and C1-1040, when used at concentrations which inhibit ERK1/2 phosphorylation but not ERK5 phosphorylation, restored N-cadherin expression in the presence of OSM, inhibited basal claudin-2 expression, but did not affect either basal or OSM-inhibited E-cadherin expression or OSM-induced expression of collagen type I and vimentin. These results suggest that in human proximal tubular cells ERK1/2 signaling represents an important component of OSM's inhibitory effect on N-cadherin expression. Furthermore, functional ERK1/2 signaling is necessary for basal claudin-2 expression.

Tubulointerstitial fibrosis is a hallmark feature of chronic renal injury. Specific therapies to control the progression of renal fibrosis toward end-stage renal failure are limited. Previous studies have demonstrated that expression of endogenous bone morphogenic protein-7 (BMP-7) is reduced in the kidneys of several inducible mouse models of acute and chronic renal disease and that administration of exogenous recombinant human BMP-7 (rhBMP-7) has a beneficial effect on kidney function. Here we report that treatment with rhBMP-7 leads to improved renal function, histology, and survival in mice deficient in the ([[alpha].sub.3]-chain of type IV collagen and MRL/Mp[J.sup.lpr/lpr] lupus mice, two genetic models for chronic renal injury and fibrosis. Such therapeutic benefit is also associated with a significant decrease in the expression of profibrotic molecules, such as type I collagen and fibronectin, in renal fibroblasts. Additionally, rhBMP-7 induces expression of active matrix metalloproteinase-2, which is potentially important for removal of fibrotic matrix. Collectively, these studies provide further evidence for rhBMP-7 as an important bone-associated protein with protective function against renal pathology. tubulointerstitial fibrosis; fibroblasts; osteogenic protein-1; matrix metalloproteinases

Author(s): Michael Zeisberg [1, 2]; Jun-ichi Hanai [1, 2]; Hikaru Sugimoto [1]; Tadanori Mammoto [1]; David Charytan [1]; Frank Strutz [1]; Raghu Kalluri (corresponding author) [1] Bone morphogenic protein (BMP)-7 [...]

A study on cis-acting elements present in the the fibroblast-specific protein (FSP1) promoter that are involved in nuclear transcription of fibroblasts is presented. FSP1 belongs to the S100 family of calcium-binding proteins that regulate microtubule dynamics, cytoskeletal-membrane interactions and calcium signal transduction. The first exon of FSP1 is noncoding and a proximal element between -187 and -88 bp has been detected to be active in fibroblasts.