Shin Yamashita

Identification of Renal Progenitor-Like Tubular Cells that Participate in the Regeneration Processes of the Kidney

The present study was conducted to explore renal progenitor-like cells that are actively engaged in tubular regeneration after injury. For addressing this issue, the existence of label-retaining cells (LRC; slow-cycling cells) in normal rat kidneys by in vivo bromodeoxyuridine (BrdU) labeling was examined. LRC were scattering among renal epithelial tubular cells of normal rat kidneys. During the recovery after renal ischemia, LRC underwent cell division and most of them became positive for proliferating cell nuclear antigen. In contrast, proliferating cell nuclear antigen-positive but BrdU-negative tubular cells were rarely observed, suggesting that cells proliferating during tubular regeneration are essentially derived from LRC. At an early phase of tubular regeneration, descendants of LRC expressed a mesenchymal marker, vimentin, and eventually became positive for an epithelial marker, E-cadherin, after multiple cell divisions. These findings suggested that LRC function as a source of regenerating cells to replace injured cells. Collectively, it was concluded that LRC are renal progenitor-like tubular cells that provide regenerating cells, which actively proliferate and eventually differentiate into epithelial cell, during tubular regeneration. It may be possible to regenerate renal tubules in vivo through the activation of LRC.

Activin A Is a Potent Activator of Renal Interstitial Fibroblasts

The present study was conducted to examine the involvement of the activin-follistatin system in the fibrotic process of the kidney. Immunoreactive activin A was upregulated in tubular cells in the kidneys with unilateral ureteral obstruction but not in normal and contralateral kidneys. Activin A promoted cell proliferation, enhanced the expression of type I collagen mRNA, and induced the production of alpha-smooth muscle actin in a rat kidney fibroblast cell line (NRK-49F cells) as well as in primary cultured renal interstitial fibroblasts. In contrast, activin A did not affect the expressions of alpha-smooth muscle actin and type I collagen in renal epithelial tubular cell lines LLC-PK1, and MDCK. Follistatin, an antagonist of activin A, significantly inhibited cell proliferation in NRK-49F cells. Blockade of activin signaling by overexpression of truncated type II activin receptor, which lacked the intracellular kinase domain, decreased cell proliferation and reduced the expression level of type I collagen mRNA in NRK-49F cells. The expression of activin A was induced by TGF-beta 1 or activin A itself. Induction of type I collagen expression by TGF-beta 1 was reduced by follistatin or by overexpression of truncated type II activin receptor. These results suggest that activin A produced by tubular cells acts as a paracrine factor that activates renal interstitial fibroblasts during the fibrotic processes of the kidney.

Involvement of renal progenitor tubular cells in epithelial- to-mesenchymal transition in fibrotic rat kidneys

Renal progenitor tubular cells (label-retaining cells [LRC]) were recently identified in normal kidneys by in vivo bromodeoxyuridine (BrdU) labeling. This study was conducted to examine the behavior of LRC in renal fibrosis. BrdU was injected intraperitoneally into normal rats daily for 7 d. After a 2-wk chase period, unilateral ureteral obstruction (UUO) was induced in these rats. In normal and contralateral kidneys, LRC were observed scattering among tubular epithelial cells. After UUO, the number of the LRC significantly increased, and most of them were positive for proliferating cell nuclear antigen (PCNA). In contrast, PCNA+ cells lacking BrdU label were rarely observed. It is interesting that LRC were detected not only in tubules but also in the interstitium after UUO. Laminin staining showed that a number of the LRC were adjacent to the destroyed tubular basement membrane. Some tubules, including LRC, lost the expression of E-cadherin after UUO. A large number of cell populations expressed vimentin, heat shock protein 47, or alpha-smooth muscle actin in the UUO kidneys, and each population contained LRC. None of the LRC was positive for these fibroblastic markers in contralateral kidneys. When renal tubules from BrdU-treated rats were cultured in the gel, some cells protruded from the periphery of the tubules and migrated into the gel. Most of these cells were BrdU+. Neither the total content of BrdU in the kidneys nor the number of LRC in bone marrow significantly changed after UUO. Collectively, these results suggest that LRC is a cell population that proliferates, migrates, and transdifferentiates into fibroblast-like cells during renal fibrosis.

Increased levels of serum sulfite in patients with acute pneumonia.

Sulfite, a common air pollutant, is toxic for humans, causing hypersensitivity or chronic airway diseases. We previously reported that sulfite is actively produced from neutrophils by stimulation with bacterial endotoxin, lipopolysaccharide (LPS). We also found that the serum sulfite concentration is increased in a rat model of sepsis induced by systemic injection of LPS. However, information on sulfite metabolism in human inflammatory conditions is limited. In the current study, the serum concentration of sulfite was determined in 25 patients with acute pneumonia. Serum sulfite concentration in pneumonia patients was significantly higher than that in control subjects (3.75 ± 0.88 vs. 1.23 ± 0.48 &mgr;M, respectively, P < 0.05). Among 20 patients, serum sulfite was serially determined before and after antibiotic therapy. The levels of serum sulfite were significantly reduced during the recovery phase compared with those during the acute phase (1.34 ± 0.56 vs. 3.65 ± 0.92 &mgr;M, respectively, P < 0.05). Moreover, neutrophils obtained from three patients during the acute phase of pneumonia spontaneously produced higher amounts of sulfite in vitro than those obtained after recovery. There was a close positive correlation (r = 0.71, P < 0.05) between serum sulfite and C-reactive protein (CRP) in patients with pneumonia. Taken together, the current findings suggest that serum sulfite increases during systemic inflammation in humans. Activated neutrophils might be responsible, at least in part, for the up-regulation of sulfite. Given various biological effects reported previously, sulfite may act as a mediator in inflammation.

Activin A Produced by Ureteric Bud Is a Differentiation Factor For Metanephric Mesenchyme

The present study was conducted to investigate the role of the activin-follistatin system in the development of metanephros. Organ culture system and cultured metanephric mesenchymal cells were used to address this issue. Activin A was localized in ureteric bud. Activin type II receptor was localized in ureteric bud as well as metanephric mesenchyme. In an organ culture system, exogenous activin A reduced the size of cultured metanephroi, delayed ureteric bud branching, and enlarged the tips of ureteric bud. Follistatin, an antagonist of activin A was used to clarify the role of endogenous activin A. Exogenous follistatin enlarged the size of cultured metanephroi, increased ureteric bud branching, and promoted cell growth in ureteric bud. Blockade of activin signaling by adenoviral transfection of dominantly negative activin mutant receptor mimics the effect of follistatin. In cultured metanephric mesenchymal cells, activin A promoted cell growth; conversely, follistatin induced apoptosis. Furthermore, activin A induced the expressions of epithelial differentiation markers in these cells. These results suggest that activin A produced by ureteric bud is not only an important regulator of ureteric bud branching, but also a differentiation factor for metanephric mesenchyme during kidney development.

Fluvastatin Reduces Renal Fibroblast Proliferation and Production of Type III Collagen: Therapeutic Implications for Tubulointerstitial Fibrosis

Background: Accumulating evidence suggests that hydroxymethylglutaryl-CoA reductase inhibitors have many biological effects beyond reducing cholesterol synthesis. In a mouse model of renal interstitial fibrosis induced by unilateral ureteral obstruction, fluvastatin, one of the lipophilic hydroxymethylglutaryl-CoA reductase inhibitors, was shown to ameliorate fibrosis. Methods: In the present study, we examined the direct effects of fluvastatin on proliferation, matrix and growth factor production by rat kidney fibroblasts (NRK-49F cells). Results: Treatment with fluvastatin reduced proliferation of NRK-49F cells in a dose-dependent manner. The addition of mevalonate or geranylgeranyl pyrophosphate but not farnesyl pyrophosphate to the culture medium almost completely abolished the effect of fluvastatin. Moreover, fluvastatin treatment decreased the expression of activated Rho in NRK-49F cells suggesting that fluvastatin may decrease cell growth through blocking the activation of Rho. The majority of fluvastatin-treated cells were arrested at the G1 phase, associated with down-regulation of cyclin A and up-regulation of cyclin-dependent kinase inhibitor p27kip1, indicating that cell cycle modulation is an important mechanism. Fluvastatin significantly decreased messenger RNA expression of type III collagen and connective tissue growth factor. Conclusions: Taken together, it is suggested that fluvastatin may prevent tubulointerstitial fibrosis in a variety of progressive renal diseases by inhibiting proliferation of interstitial fibroblasts and their matrix synthesis.

Follistatin, an Activin Antagonist, Ameliorates Renal Interstitial Fibrosis in a Rat Model of Unilateral Ureteral Obstruction

Activin, a member of the TGF-β superfamily, regulates cell growth and differentiation in various cell types. Activin A acts as a negative regulator of renal development as well as tubular regeneration after renal injury. However, it remains unknown whether activin A is involved in renal fibrosis. To clarify this issue, we utilized a rat model of unilateral ureteral obstruction (UUO). The expression of activin A was significantly increased in the UUO kidneys compared to that in contralateral kidneys. Activin A was detected in glomerular mesangial cells and interstitial fibroblasts in normal kidneys. In UUO kidneys, activin A was abundantly expressed by interstitial α-SMA-positive myofibroblasts. Administration of recombinant follistatin, an activin antagonist, reduced the fibrotic area in the UUO kidneys. The number of proliferating cells in the interstitium, but not in the tubules, was significantly lower in the follistatin-treated kidneys. Expression of α-SMA, deposition of type I collagen and fibronectin, and CD68-positive macrophage infiltration were significantly suppressed in the follistatin-treated kidneys. These data suggest that activin A produced by interstitial fibroblasts acts as a potent profibrotic factor during renal fibrosis. Blockade of activin A action may be a novel approach for the prevention of renal fibrosis progression.

Efficacy of tocilizumab, a humanized neutralizing antibody against interleukin-6 receptor, in progressive renal injury associated with Castleman's disease

Castlemans disease is a benign lymphoproliferative disorder in which interleukin-6 (IL-6), a pleiotropic proinflammatory cytokine, is thought to play a pathogenetic role. Presented is the case of a 72-year-old man with Castlemans disease who exhibited progressive renal dysfunction with proteinuria. Renal biopsy revealed mesangial hypercellularity and matrix expansion in most glomeruli and peritubular inflammatory cell infiltration. Immunofluorescence studies showed intense deposition of IgG in a granular pattern along the glomerular basement membrane. Histological features were compatible with membranoproliferative glomerulonephritis accompanied by interstitial inflammatory cell infiltration. Immunohistological analysis showed that IL-6 was abundantly expressed by tubular cells and interstitial macrophages, suggesting involvement of IL-6 in the renal injury. As a result of administration of tocilizumab, a humanized anti-IL-6 receptor antibody, the patient experienced clinical and biochemical improvement of Castlemans disease, including marked reduction of proteinuria and stabilization of renal function. These findings suggest the efficacy of tocilizumab against Castlemans disease and its renal complications.