Osamu Takase

Intrarenal Injection of Bone Marrow-Derived Angiogenic Cells Reduces Endothelial Injury and Mesangial Cell Activation in Experimental Glomerulonephritis

Loss of glomerular endothelial cells has been suggested to contribute to the progression of glomerular injury. Although therapeutic angiogenesis induced by administration of bone marrow-derived endothelial progenitor cells has been observed in disease models of endothelial injury, the effects on renal disease have not been clarified. Whether administration of culture-modified bone marrow mononuclear cells would mitigate the glomerular endothelial injury in anti-Thy1.1 nephritis was investigated. After cultivation under conditions that promote endothelial progenitor cell growth, bone marrow mononuclear cells were labeled with CM-DiI, a fluorescence marker, and injected into the left renal artery of Lewis rats with anti-Thy1.1 glomerulonephritis. The decrease in glomerular endothelial cells was significantly attenuated in the left kidney, as compared with the right, in nephritic rats that received the cell infusion. Glomerular injury score, the area positive for mesangial alpha-smooth muscle actin, and infiltration of macrophages were significantly decreased in the left kidney. CM-DiI-positive cells were distributed in glomeruli of the left kidney but not in those of the right kidney. Among CM-DiI-labeled cells incorporated into glomeruli, 16.5 +/- 1.2% of cells were stained with an endothelial marker, rat endothelial cell antigen-1. Culture-modified mononuclear cells secreted 281.2 +/- 85.0 pg of vascular endothelial growth factor per 10(5) cells per day. In conclusion, intra-arterial administration of culture-modified bone marrow mononuclear cells reduced endothelial injury and mesangial activation in anti-Thy1.1 glomerulonephritis. Incorporation into the glomerular endothelial lining and production of angiogenic factor(s) are likely to contribute to the protective effects of culture-modified mononuclear cells against glomerular injury.

The Role of NF-κB Signaling in the Maintenance of Pluripotency of Human Induced Pluripotent Stem Cells

NF-κB signaling plays an essential role in maintaining the undifferentiated state of embryonic stem (ES) cells. However, opposing roles of NF-κB have been reported in mouse and human ES cells, and the role of NF-κB in human induced pluripotent stem (iPS) cells has not yet been clarified. Here, we report the role of NF-κB signaling in maintaining the undifferentiated state of human iPS cells. Compared with differentiated cells, undifferentiated human iPS cells showed an augmentation of NF-κB activity. During differentiation induced by the removal of feeder cells and FGF2, we observed a reduction in NF-κB activity, the expression of the undifferentiation markers Oct3/4 and Nanog, and the up-regulation of the differentiated markers WT-1 and Pax-2. The specific knockdown of NF-κB signaling using p65 siRNA also reduced the expression of Oct3/4 and Nanog and up-regulated WT-1 and Pax-2 but did not change the ES-like colony formation. Our results show that the augmentation of NF-κB signaling maintains the undifferentiated state of human iPS and suggest the importance of this signaling pathway in maintenance of human iPS cells.

NF-κB-dependent genes induced by proteinuria and identified using DNA microarrays

BackgroundA close correlation has been shown between tubulointerstitial (TI) injury and the outcome of renal dysfunction, and nuclear factor-kappaB (NFκB) has been shown to play a key role in proteinuria-induced TI injury. To explore the molecular mechanisms of the proteinuria-induced TI injury further, we have analyzed renal gene expression with DNA microarrays, with and without specific inhibition of NF-κB in the proximal tubules.MethodsUnilaterally nephrectomized rats loaded with bovine serum albumin (BSA) were used as a model of proteinuric renal injury. Renal NF-κB activation was inhibited by gene transfer of the truncated form of IκBα (inhibitor of NF-κB) via the injection of a recombinant adenovirus vector into the renal artery, an method established in a previous study. Total RNA was extracted from the kidney and analyzed with a DNA microarrays containing 1081 genes.ResultsRenal NF-κB activation and TI injury in BSA-loaded proteinuric rats were inhibited by the gene transfer of the truncated form of IκBα. DNA microarray analysis revealed 45 up-regulated genes and six down-regulated genes in the proteinuric rats, and expression of 23 of these 51 genes was significantly altered by NF-κB inhibition. Among these 23 genes, we focused on clusterin and confirmed the results of microarray analysis by Western blotting and PCR.ConclusionIn this study, 23 genes of 51 proteinuria-related genes were regulated by NF-κB activation, suggesting that some of these genes may serve as target molecules for the treatment of progressive TI injury.

Eicosapentaenoic acid regulates IκBα and prevents tubulointerstitial injury in kidney.

Fish oil containing n-3 polyunsaturated fatty acids such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) is well known to prevent the progression of IgA nephropathy. However, the mechanism through which fish oil prevents the progression of renal injury remains uncertain. We tried to clarify the effects of EPA on tubulointerstitial injury in the kidney both in vivo and in vitro. We examined the effects of EPA, especially to focus on nuclear factor kappa B (NF-κB), using Thy-1 nephritis models. Also the mechanism of EPA was investigated using small-interfering RNA (siRNA) in lipopolysaccharide (LPS)-stimulated proximal tubular epithelial cells (PTECs). In Thy-1 nephritis models, EPA significantly inhibited tubulointerstitial injury and the infiltration of macrophages into tubulointerstitial lesions except severe glomerular injury at early stage. Compared with control animals, NF-κB activation was significantly augmented in the Thy-1 nephritic kidney. However, treatment with EPA significantly reduced NF-κB activation, down-regulated the expressions of NF-κB-dependent molecules. Also in LPS-stimulated PTECs, LPS augmented NF-κB activation and the expression of NF-κB-dependent molecules. As in the case with the Thy-1 nephritis models, treatment with EPA inhibited them, prevented the degradation of IκBα in LPS-stimulated PTECs. Pre-treatment with siRNA for IκBα abolished the inhibitory effect of EPA on LPS-induced NF-κB activation, suggesting that EPA inhibited NF-κB activation by regulating IκBα. Our results indicate that EPA prevents the early progression of tubulointerstitial injury in Thy-1 nephritis models, and the inhibitory effect of EPA on the expression of inflammatory molecules via the regulation of IκBα in cultured cells may explain this mechanism.

Comparison of antioxidant activity of cilnidipine and amlodipine

To the Editor: Priority claims are often difficult to substantiate. Such is the case with the Canadian school of nocturnal hemodialysis, which has continuously refused to recognize that they were not the first to carry out overnight hemodialysis. Thus, the statement ‘nocturnal hemodialysis, a technique first developed in the 1970s’ published in Kidney International, cannot be allowed to go unchallenged. On numerous occasions, I have pointed out to the Canadian nocturnal hemodialysis school that frequency of hemodialysis does not permit claims to originality in the use of the night for hemodialysis. We were the first to report the successful use of unattended overnight hemodialysis in 1963 and this was recognized by Scribner in 1966 when he stated ‘Shaldon has taken a big step forward in this respect by demonstrating the feasibility of unattended nighttime hemodialysis’. The development of the high low venous pressure monitor, which was the key to safe overnight hemodialysis was also reported in the Lancet in 1963. As regards frequency, we started in 1961 with two dialyses per week, but very soon increased the frequency to three, four, and even five dialyses per week. The system was reported in detail at the first meeting of the European Dialysis and Transplant Association in Amsterdam in 1964.

Immunomodulation with eicosapentaenoic acid supports the treatment of autoimmune small-vessel vasculitis

Small-vessel vasculitis is a life-threatening autoimmune disease that is frequently associated with anti-neutrophil cytoplasmic antibodies (ANCAs). Conventional immunotherapy including steroids and cyclophosphamide can cause serious adverse events, limiting the efficacy and safety of treatment. Eicosapentaenoic acid (EPA), a key component of fish oil, is an omega-3 polyunsaturated fatty acid widely known to be cardioprotective and beneficial for vascular function. We report two elderly patients with systemic ANCA-associated vasculitis (AAV) in whom the administration of EPA in concert with steroids safely induced and maintained remission, without the use of additioal immunosuppressants. To explore the mechanisms by which EPA enhances the treatment of AAV, we employed SCG/Kj mice as a spontaneous murine model of AAV. Dietary enrichment with EPA significantly delayed the onset of crescentic glomerulonephritis and prolonged the overall survival. EPA-derived anti-inflammatory lipid mediators and their precursors were present in the kidney, plasma, spleen, and lungs in the EPA-treated mice. Furthermore, a decrease in ANCA production and CD4/CD8-double negative T cells, and an increase in Foxp3+ regulatory T cells in the lymph nodes of the kidney were observed in the EPA-treated mice. These clinical and experimental observations suggest that EPA can safely support and augment conventional therapy for treating autoimmune small-vessel vasculitis.

Roles and regulation of bone morphogenetic protein-7 in kidney development and diseases

The gene encoding bone morphogenetic protein-7 (Bmp7) is expressed in the developing kidney in embryos and also in the mature organ in adults. During kidney development, expression of Bmp7 is essential to determine the final number of nephrons in and proper size of the organ. The secreted BMP7 acts on the nephron progenitor cells to exert its dual functions: To maintain and expand the progenitor population and to provide them with competence to respond to differentiation cues, each relying on distinct signaling pathways. Intriguingly, in the adult organ, BMP7 has been implicated in protection against and regeneration from injury. Exogenous administration of recombinant BMP7 to animal models of kidney diseases has shown promising effects in counteracting inflammation, apoptosis and fibrosis evoked upon injury. Although the expression pattern of Bmp7 has been well described, the mechanisms by which it is regulated have remained elusive and the processes by which the secretion sites of BMP7 impinge upon its functions in kidney development and diseases have not yet been assessed. Understanding the regulatory mechanisms will pave the way towards gaining better insight into the roles of BMP7, and to achieving desired control of the gene expression as a therapeutic strategy for kidney diseases.

Adult stem-like cells in kidney.

Human pluripotent cells are promising for treatment for kidney diseases, but the protocols for derivation of kidney cell types are still controversial. Kidney tissue regeneration is well confirmed in several lower vertebrates such as fish, and the repair of nephrons after tubular damages is commonly observed after renal injury. Even in adult mammal kidney, renal progenitor cell or system is reportedly presents suggesting that adult stem-like cells in kidney can be practical clinical targets for kidney diseases. However, it is still unclear if kidney stem cells or stem-like cells exist or not. In general, stemness is defined by several factors such as self-renewal capacity, multi-lineage potency and characteristic gene expression profiles. The definite use of stemness may be obstacle to understand kidney regeneration, and here we describe the recent broad findings of kidney regeneration and the cells that contribute regeneration.

Basic helix-loop-helix transcriptional factor MyoR regulates BMP-7 in acute kidney injury

MyoR was originally identified as a transcriptional repressor in embryonic skeletal muscle precursors, but its function in adult kidney has not been clarified. In this study, we tried to clarify the functional role of MyoR using MyoR(-/-) mice. Cisplatin induced a significantly higher degree of severe renal dysfunction, tubular injury, and mortality in MyoR(-/-) mice than in wild-type mice. The injection of cisplatin significantly increased the number of apoptotic cells in the kidney tissues of MyoR(-/-) mice, compared with that in wild-type mice. To clarify the mechanism of severe cisplatin-induced damage and apoptosis in MyoR(-/-) mice, we focused on the p53 signaling pathway and bone morphogenic protein-7 (BMP-7). Treatment with cisplatin significantly activated p53 signaling in cultured renal proximal tubular epithelial cells (RTECs) in both wild-type and MyoR(-/-) mice, but no significant difference between the groups was observed. The injection of cisplatin significantly increased the expression of BMP-7 in the kidney tissues of wild-type mice, but no increase was observed in the MyoR(-/-) mice. Treatment with cisplatin significantly increased the expression of BMP-7 in cultured RTECs from wild-type mice but not in those from MyoR(-/-) mice. Moreover, treatment with recombinant BMP-7 rescued the cisplatin-induced apoptosis in RTECs from MyoR(-/-) mice. Taken together, our results demonstrate a new protective role of MyoR in adult kidneys that acts through the regulation of BMP-7.

Immunoregulation of Inflammation in Chronic Kidney Disease

Chronic kidney disease (CKD) is a commonly encountered condition in clinical practice. Up to 15% of the adult population worldwide is affected by CKD. Patients with CKD are at a high risk of developing hypertension and other cardiovascular diseases, with associated morbidity and mortality. Stage V is end-stage renal disease (ESRD), for which patients require renal replacement therapy with either dialysis or renal transplantation. A patient with ESRD on dialysis has a 50% chance of surviving three years. Overall, CKD is a disease with a tenfold increased incidence over three decades in which affected patients can lose 70% of their life span. There are many underlying etiologies of CKD, including diabetes, hypertension, primary and secondary glomerulonephritis/vasculitis, and tubulointerstitial nephritis. Irrespective of initiator, there appears to be an important role for inflammation in CKD. In fact, patients with CKD are frequently treated nonspecifically with immunosuppressive and/or antihypertensive agents. Unfortunately, there have not been new treatment regimens for CKD induced by various underlying causes. This special issue covers the broad topic of immunoregulation of inflammation in CKD. Four review articles and two research articles discuss the mechanism of inflammation in animal models of lupus nephritis and obstructive uropathy, as well as effects on resident renal cells. It is our hope that these contribute to a better understanding of immunoregulation of inflammation in CKD, which can stimulate development of better therapeutic approaches and provision of optimal care to patients. In the first paper, “Mediators of inflammation and their effect on resident renal cells: implications in lupus nephritis,” S. Yung et al. review immunoregulation in lupus nephritis, an important cause of CKD. They concentrate on interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), type I interferons (IFNs) and hyaluronan, as among the most important mediators in this disease. They provide background of the fundamental biology of IL-6, TNF-α, type I IFNs, and hyaluronan, followed by their roles in both experimental and human lupus nephritis. In the second paper, “LMW heparin prevents increased kidney expression of proinflammatory mediators in (NZB×NZW)F1 mice,” A. Hedberg et al. have studied effects of low molecular weight (LMW) heparin on lupus nephritis occurring in (NZB×NZW)F1 (B/W) mice. Like in human lupus nephritis, these mice progressively develop CKD. As this occurs, there is increased expression of a diversity of proinflammatory mediators. In their study, they show that LMW heparin specifically lowers CCR2, IL-1β, and TLR7 expression. This may be attributable to the ability of LMW heparin to enhance nucleosomal degradation and/or binding to glomerular sites as components of immune complexes. In the third paper, “Interactions between cytokines, congenital anomalies of kidney and urinary tract and chronic kidney disease,” A. C. S. Silva et al. reviewed the relative roles for cytokines and chemokines in the pathophysiology of congenital anomalies of the kidney and urinary tract and how they can affect progression of CKD. They include experimental and clinical evidence to show that urine measurements of cytokines could prove useful as predictors of urinary tract obstruction and renal scarring. In the fourth paper, “MicroRNAs implicated in the immunopathogenesis of lupus nephritis,” C. B. Chafin and C. M. Reilly provide a review that addresses a current and important topic: the role of microRNAs in the pathogenesis of lupus nephritis. The authors collated a large amount of available data regarding the potential role of microRNAs as therapeutic targets and how these might underlie potential treatment strategies for this important disease. In the fifth paper, “Inflammatory chemokine expression via toll-like receptor 3 signaling in normal human mesangial cells,” H. Tanaka and T. Imaizumi summarize their experimental results regarding signaling pathways in human mesangial cells activated upon treatment with a synthetic analogue of viral dsRNA. The signaling pathways activated through TLR3 in mesangial cells may be proinflammatory. The relevance includes effects of viral and “pseudoviral” infections on existing CKD, as well as pathogenic mechanisms that may underlie primary glomerulonephritis. In the sixth paper, “Contrasting effects of systemic monocyte/macrophage and CD4+ T cell depletion in a reversible ureteral obstruction mouse model of chronic kidney disease,” L. Chaves et al. show that depleting macrophage and CD4+ T cells had distinct effects on manifestations of CKD in a reversible model of unilateral ureteral obstruction (rUUO). Based on these results, the authors concluded that modulation of immune cells during injury and repair altered the development of CKD in the rUUO model. Their rUUO model is unique, and findings in this study provide interesting clues to the mechanisms of CKD progression. We are certain that the readers of this special issue will find several interesting points of discussion in the published papers. We hope these papers can stimulate further experimentation to dissect the immunoregulation of inflammation in CKD and allow development of new therapeutic strategies for CKD.