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SCF-Skp2 E3 ubiquitin ligase (Skp2 hereafter) targets several cell cycle regulatory proteins for degradation via the ubiquitindependent pathway. However, the target-specific physiological functions of Skp2 have not been fully elucidated in kidney diseases. We previously reported an increase in Skp2 in progressive nephropathy and amelioration of unilateral ureteral obstruction (UUO) renal injury associated with renal accumulation of p27 in Skp2-/- mice. However, it remains unclear whether the amelioration of renal injury in Skp2-/- mice is solely caused by p27 accumulation, since Skp2 targets several other proteins. Using Skp2-/-p27-/- mice, we investigated whether Skp2 specifically targets p27 in the progressive nephropathy mediated by UUO. In contrast to the marked suppression of UUO renal injury in Skp2-/- mice, progression of tubular dilatation associated with tubular epithelial cell proliferation and tubulointerstitial fibrosis with increased expression of collagen and a-smooth muscle actin were observed in the obstructed kidneys in Skp2-/-p27-/- mice. No significant increases in other Skp2 target proteins including p57, p130, TOB1, cyclin A and cyclin D1 were noted in the UUO kidney in Skp2-/- mice, while p21, c-Myc, b-Myb and cyclin E were slightly increased. Contrary to the ameliorated UUO renal injure by Skp2-deficiency, the amelioration was canceled by the additional p27-deficiency in Skp2-/-p27-/- mice. These findings suggest a pathogenic role of the reduction in p27 targeted by Skp2 in the progression of nephropathy in UUO mice. [ABSTRACT FROM AUTHOR]

We report on a patient with membranoproliferative glomerulonephritis (MPGN) type III, whose repeated renal biopsies show evolution from a type I-like pattern to a type III pattern of immune complex formation over a 1-year period. Based on the development of experimental in situ immune complex glomerulonephritis, we discuss possible mechanisms for the formation of subepithelial deposits in type III MPGN with reference to an experimental in situ immune complex model of glomerulonephritis. [ABSTRACT FROM AUTHOR]

Hepatocyte growth factor and its receptor, Met, activate biological pathways necessary for repair and regeneration following kidney injury. The Met receptor is expressed in multiple cell types within the kidney, each of which is capable of regulating fibrotic responses. To specifically address the role of the Met receptor in the adult collecting duct during renal injury, a conditional knockout mouse (Metfl/fl;HoxB7-Cre) was generated and tested using unilateral ureteral obstruction, a model of nephron injury, fibrosis, and repair. Following obstruction in these mice there was increased expression of collagens I and IV along with plasminogen activator inhibitor 1, a known regulator of matrix degradation, compared to ureteral obstructed non-flox littermates. There were trends toward increased interstitial fibrosis, infiltration of the interstitium, and acute tubular necrosis in the knockout mice despite similar degrees of hydronephrosis to the control littermates. The Metfl/fl;HoxB7-Cre mice; however, had reduced tubular cell proliferation and kidney regenerative capacity after release of the obstruction, thus leading to diminished functional recovery. We suggest that Met receptor signaling in the collecting duct acts as a major regulator of cell survival and propagation of the repair process with a possible secondary role to diminish inflammatory and fibrotic responses. [ABSTRACT FROM AUTHOR]

Treatment with anti-TGF-β antibody ameliorates chronic progressive nephritis by inhibiting Smad/TGF-β signaling. Background. Although short-term treatment with anti-transforming growth factor-β (TGF-β) antibody (αT) has been shown to prevent early glomerular lesions, its long-term effects and molecular mechanisms, including intracellular signaling, remain poorly understood. We examined whether αT treatment induces prevention of renal insufficiency and fibrosis, and affects the TGF-β/Smad signaling pathway in rats with chronic progressive anti-thymocyte serum (ATS) nephritis induced by repeated ATS injections on days 0 and 7. Methods. Nephritic and non-nephritic rats were treated with either αT or control immunoglobulin (Ig)G twice weekly for 4 weeks from days 7 to 35 (each group, N= 21). Renal lesions and cortical expression of TGF-β1, TGF-β2, TGF-β3, type II TGF-β receptor (TβRII), Smads, type I collagen, and plasminogen activator inhibitor-1 were examined by immunohistochemistry, Western blot, and/or real-time reverse transcription polymerase chain reaction (RT-PCR). The binding of Smad3 in renal cortical cell nuclei to the Smad-binding element (SBE) was investigated by the electrophoretic mobility shift assay. Results. Nephritic rats developed heavy proteinuria, renal insufficiency, and increased extracellular matrix deposition resulting in renal fibrosis. Cortical expression levels of TGF-β1, TGF-β2, TβRII, and Smad2, but not TGF-β3, Smad3, and Smad4 were increased. Expression and preferential localization of phosphorylated Smad2/3 in the glomerular and tubular cell nuclei, and Smad3-SBE complex-forming activity were also increased. Four-week αT treatment resulted in marked amelioration of chronic progressive ATS nephritis at 8 weeks. Conclusion. In chronic progressive ATS nephritis, the TGF-β/Smad signaling was up-regulated. TGF-β blockade by αT suppressed the progression of renal scarring, at least in part, via inhibition of activated TGF-β/Smad signaling. [ABSTRACT FROM AUTHOR]

Despite suppression of the circulating renin-angiotensin system (RAS), high salt intake (HSI) aggravates kidney injury in chronic kidney disease. To elucidate the effect of HSI on intrarenal RAS, we investigated the levels of intrarenal prorenin, renin, (pro)renin receptor (PRR), receptor-mediated prorenin activation, and ANG II in chronic anti-thymocyte serum (ATS) nephritic rats on HSI. Kidney fibrosis grew more severe in the nephritic rats on HSI than normal salt intake. Despite suppression of plasma renin and ANG II, marked increases in tubular prorenin and renin proteins without concomitant rises in renin mRNA, non-proteolytically activated prorenin, and ANG II were noted in the nephritic rats on HSI. Redistribution of PRR from the cytoplasm to the apical membrane, along with elevated non-proteolytically activated prorenin and ANG II, was observed in the collecting ducts and connecting tubules in the nephritic rats on HSI. Olmesartan decreased cortical prorenin, non-proteolytically activated prorenin and ANG II, and apical membranous PRR in the collecting ducts and connecting tubules, and attenuated the renal lesions. Cell surface trafficking of PRR was enhanced by ANG II and was suppressed by olmesartan in Madin-Darby canine kidney cells. These data suggest the involvement of the ANG II-dependent increase in apical membrane PRR in the augmentation of intrarenal binding of prorenin and renin, followed by nonproteolytic activation of prorenin, enhancement of renin catalytic activity, ANG II generation, and progression of kidney fibrosis in the nephritic rat kidneys on HSI. The origin of the increased tubular prorenin and renin remains to be clarified. Further studies measuring the urinary prorenin and renin are needed.

Increased expression levels of tumor necrosis factor-alpha (TNFalpha) is involved in tubulointerstitial cell proliferation and apoptosis in obstructive renal injury. Two TNFalpha receptors (TNFRs), TNFR1 and TNFR2, are known to exist. On TNFalpha binding, TNFR1 recruits TNFR-associated death domain (TRADD), an assembly platform to mediate TNFR1 signaling. We investigated postreceptor TRADD regulation in rat kidneys with unilateral ureteral obstruction (UUO). Whereas UUO was associated with increased expression levels of TNFalpha, TNFR1, TNFR2, and TRADD mRNAs, it resulted in the marked decrease of TRADD protein levels (which appeared at day 1 and persisted thereafter) and a slight decrease in TNFR1 protein levels at days 7 and 14. Both ubiquitination and degradation of TRADD were increased in UUO kidneys, degradation of TRADD was stimulated by TNFalpha in HK-2 cells, and TRADD degradation was suppressed by proteasome inhibitor. Inhibition of TNFalpha by soluble TNFR2, etanercept, reduced significantly, although transiently, tubular and interstitial cell proliferation, fibronectin expression, and apoptosis in UUO kidneys, and also suppressed TRADD degradation. These data suggest that the decrease in TRADD resulting from enhanced ubiquitin-dependent degradation is involved in obstructive renal injury. Since TRADD is not incorporated into TNFR2-mediated TNFalpha signaling, the persistent decrease in TRADD, associated with a mild decrease in TNFR1 levels, may function, at least in part, to divert TNFalpha signals toward a TNFR2-mediated pathway in UUO kidneys.

Following organ injury, morphogenic epithelial responses can vary depending on local cell density. In the present study, the role of cell confluence in determining the responsiveness of renal epithelial cells to the dedifferentiating morphogenic signals of hepatocyte growth factor (HGF) was examined. Increasing confluence resulted in a greater tendency of cells to organize into epithelial tubes and a significant decrease in migratory responsiveness to HGF. Analysis of downstream signaling revealed that the HGF receptor c-Met was equally activated in confluent and nonconfluent cells following HGF stimulation but that phosphoinositide 3-kinase-dependent activation of Akt and Rac were selectively diminished in confluent cells. In nonconfluent cells treated with HGF, the high level of Akt activation resulted in inhibitory phosphorylation of glycogen synthase kinase 3beta (GSK-3beta) and increased beta-catenin nuclear signaling. In contrast, confluent cells, in which HGF-stimulated Akt activation was diminished, displayed less inhibitory phosphorylation of GSK-3beta and less nuclear signaling by beta-catenin. Overexpression of beta-catenin (SA), which cannot be phosphorylated by GSK-3beta and targeted for ubiquitination, significantly increased migration in fully confluent cells. Thus, cells maintained at high confluence selectively downregulate signaling events such as Rac activation and beta-catenin-dependent transcription that would otherwise promote cell dedifferentiation and migration.

Smad ubiquitination regulatory factor 2 (Smurf2), a ubiquitin ligase for Smads, plays critical roles in the regulation of transforming growth factor-beta (TGF-beta)-Smad signaling via ubiquitin-dependent degradation of Smad2 and Smad7. We found that TGF-beta stimulates Smurf2 expression. TGF-beta activated the Smurf2 promoter in a TGF-beta responsive cell lines, whereas IL-1alpha, PDGF and epidermal growth factor did not. TGF-beta-mediated Smurf2 promoter activation was inhibited by Smad7 or an activin receptor-like kinase 5 inhibitor but not by dominant negative Smad or disruption of Smad-binding elements in the promoter. Moreover, inhibition of the phosphatidil inositol 3 kinase (PI3K)/Akt pathway suppressed TGF-beta-mediated Smurf2 induction. These results suggest that TGF-beta stimulates Smurf2 expression by Smad-independent pathway such as PI3K/Akt pathway via TGF-beta receptor.

The nephrotic syndrome is defined by heavy proteinuria due to abnormal increase of glomerular permeability and following hypoalbuminemia, hyperlipidemia and edema. Disorders of size selective barrier, charge selective barrier, slit diaphragm and circulating permeability factors are thought to be the causes of proteinuria. Most patients with nephrotic edema have primary salt retention. Overproduction and impaired catabolism of lipoproteins are the causes of hyperlipidemia. Abnormality of coagulation factors is also associated with nephrotic syndrome. Nephrotic syndrome may be primary or secondary to systemic disorders. Once diagnosed, the cause for the nephrotic syndrome must be examined.

Increases in transforming growth factor-beta (TGF-beta) expression and extracellular matrix accumulation are transient in acute self-limited mesangial proliferative glomerulonephritis induced by a single injection of anti-thymocyte serum (ATS), while these increases persist following repeated injections that produce chronic progressive sclerosing glomerulonephritis with tubulointerstitial lesions. However, little is known about the expression of TGF-beta receptors (TbetaRs) in cells involved in the proliferative and sclerosing renal lesions. A study of protein and mRNA expression for type I (TbetaRI), type II (TbetaRII), and type III (TbetaRIII) TbetaR in both forms of nephritis was therefore carried out by immunohistochemistry and in situ hybridization. Inhibition of cell proliferation and stimulation of matrix production by TGF-beta1 were assessed in isolated glomeruli using [(3)H]thymidine incorporation and [(3)H]proline metabolic labelling, respectively. In acute self-limited nephritis, expression of TbetaRI, TbetaRII, and TbetaRIII increased in the glomerular and Bowman's capsular epithelial cells comprising the glomerular tuft adhesions to Bowman's capsules. However, TbetaRII expression was not prominent in proliferating mesangial cells. Glomeruli isolated from rats with acute self-limited nephritis at day 7, when mesangial cell proliferation was maximal, were partially resistant to the mitoinhibitory effects of TGF-beta1. In contrast, expression of all three TbetaRs was elevated in glomerular and tubulointerstitial lesions in chronic progressive nephritis, and glomeruli isolated from rats with chronic progressive nephritis 7 days after the second ATS injection were sensitive to TGF-beta1. These data suggest that distinct cellular responses to TGF-beta1 resulting from differential expression of TbetaR underlie the difference between acute self-limited mesangial proliferative and chronic progressive sclerosing ATS nephritis in the development of proliferative and sclerotic renal lesions., (Copyright 2002 John Wiley & Sons, Ltd.)