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en: 14-3-3 proteins stabilize actin and vimentin filaments to maintain processe in renal glomerular podocyte

en: 14-3-3 proteins are a ubiquitously expressed family of adaptor proteins. Despite exhibiting high sequence homology, several 14-3-3 isoforms have isoform-specific binding partners and roles. We reported that 14-3-3β interacts with FKBP12 and synaptopodin to maintain the structure of actin fibers in podocytes. However, the precise localization and differential role of 14-3-3 isoforms in kidneys are unclear. Herein, we showed that 14-3-3β in glomeruli was restricted in podocytes, and 14-3-3ς in glomeruli was expressed in podocytes and mesangial cells. Although 14-3-3β was dominantly co-localized with FKBP12 in the foot processes, a part of 14-3-3β was co-localized with Par3 at the slit diaphragm. 14-3-3β interacted with Par3, and FKBP12 bound to 14-3-3β competitively with Par3. Deletion of 14-3-3β enhanced the interaction of Par3 with Par6 in podocytes. Gene silencing for 14-3-3β altered the structure of actin fibers and process formation. 14-3-3β and synaptopodin expression was decreased in podocyte injury models. In contrast, 14-3-3ς in podocytes was expressed in the primary processes. 14-3-3ς interacted with vimentin but not with the actin-associated proteins FKBP12 and synaptopodin. Gene silencing for 14-3-3ς altered the structure of vimentin fibers and process formation. 14-3-3ς and vimentin expression was increased in the early phase of podocyte injury models but was decreased in the late stage. Together, the localization of 14-3-3β at actin cytoskeleton plays a role in maintaining the foot processes and the Par complex in podocytes. In contrast, 14-3-3ς at vimentin cytoskeleton is essential for maintaining primary processes.

en: The FASEB journal

ja: The FASEB journal


en: 14-3-3蛋白質は細胞骨格とPar複合体の調節によって細胞突起を維持する

ja: 14-3-3蛋白質は細胞骨格とPar複合体の調節によって細胞突起を維持する

en: Japanese Journal of Nephrology

ja: 日本腎臓学会誌


en: 腎糸球体における14-3-3 isoforms の機能解析

ja: 腎糸球体における14-3-3 isoforms の機能解析


en: Th17 Cells Participate in Thy1.1 Glomerulonephritis Which Is Ameliorated by Tacrolimus

en: Thy1.1 glomerulonephritis (Thy1.1 GN) in rats is widely used as an experimental model of mesangial proliferative glomerulonephritis (GN). We previously reported that T-helper (Th) cells were accumulated in glomeruli from the early phase of this model and that not Th2 cells but Th1 cells play an important role in the development of glomerular alterations. Although Th17 is reported to be involved in the pathogenesis of several autoimmune diseases, the role of Th17 cells in the pathogenesis of mesangial alterations in Thy1.1 GN remains unclear. The kinetics of the infiltration of subsets of Th cells and the expression of IL-17 in Thy1.1 GN were analyzed. Next, the localization and the cell types of IL-17 receptor (IL-17R)-positive cells and IL-6-positive cells were analyzed. Then, the effect of tacrolimus on the expressions of Th17-related cytokines in Thy1.1 GN was analyzed. Not only Th1 cells but also Th17 cells were recruited into glomeruli from the early phase of the disease. mRNA expression of IL-17 in glomeruli was elevated. The increased positive expression of IL-17R was detected in the mesangial area, and some of IL-17R-positive cells were co-stained with IL-6. Tacrolimus treatment ameliorated mesangial alterations by suppressing the expressions of Th17-related cytokines such as IL-17 and IL-6. Th17 cells participate in the development of Thy1.1 GN, a mimic of mesangial proliferative GN, and Th17 cells and their related cytokines are pertinent therapeutic targets.

en: American Journal of Nephrology

ja: American Journal of Nephrology


en: Tacrolimus ameliorates podocyte injury by restoring FK506 binding protein 12 (FKBP12) at actin cytoskeleton


en: Synbindin Downregulation Participates in Slit Diaphragm Dysfunction

en: Synbindin, originally identified as a neuronal cytoplasmic molecule, was found in glomeruli. The cDNA subtractive hybridization technique showed the mRNA expression of synbindin in glomeruli was downregulated in puromycin aminonucleoside (PAN) nephropathy, a mimic of minimal-change nephrotic syndrome. The expression of synbindin in podocytes was analyzed in normal rats and 2 types of rat nephrotic models, anti-nephrin antibody-induced nephropathy, a pure slit diaphragm injury model, and PAN nephropathy, by immunohistochemical analysis and RT-PCR techniques. To elucidate the function of synbindin, a gene silencing study with human cultured podocytes was performed. Synbindin was mainly expressed at the slit diaphragm area of glomerular epithelial cells (podocytes). In both nephrotic models, decreased mRNA expression and the altered staining of synbindin were already detected at the early phase when proteinuria and the altered staining of nephrin, a key molecule of slit diaphragm, were not detected yet. Synbindin staining was clearly reduced when severe proteinuria was observed. When the cultured podocytes were treated with siRNA for synbindin, the cell changed to a round shape, and filamentous actin structure was clearly altered. The expression of ephrin-B1, a transmembrane protein at slit diaphragm, was clearly lowered, and synaptic vesicle-associated protein 2B (SV2B) was upregulated in the synbindin knockdown cells. Synbindin participates in maintaining foot processes and slit diaphragm as a downstream molecule of SV2B-mediated vesicle transport. Synbindin downregulation participates in slit diaphragm dysfunction. Synbindin can be an early marker to detect podocyte injury.


en: NephrinEphrin-B1Na+/H+ Exchanger Regulatory Factor 2EzrinActin Axis Is Critical in Podocyte Injury

en: Ephrin-B1 is one of the critical components of the slit diaphragm of kidney glomerular podocyte. However, the precise function of ephrin-B1 is unclear. To clarify the function of ephrin-B1, ephrin-B1-associated molecules were studied. RNA-sequencing analysis suggested that Na/H exchanger regulatory factor 2 (NHERF2), a scaffolding protein, is associated with ephrin-B1. NHERF2 was expressed at the apical area and the slit diaphragm, and interacted with the nephrin-ephrin-B1 complex at the slit diaphragm. The nephrin-ephrin-B1-NHERF2 complex interacted with ezrin bound to F-actin. NHERF2 bound ephrin-B1 via its first postsynaptic density protein-95/disks large/zonula occludens-1 domain, and podocalyxin via its second postsynaptic density protein-95/disks large/zonula occludens-1 domain. Both in vitro analyses with human embryonic kidney 293 cells and in vivo study with rat nephrotic model showed that stimulaiton of the slit diaphragm, phosphorylation of nephrin and ephrin-B1, and dephosphorylation of NHERF2 and ezrin, disrupted the linkages of ephrin-B1-NHERF2 and NHERF2-ezrin. It is conceivable that the linkage of nephrin-ephrin-B1-NHERF2-ezrin-actin is a novel critical axis in the podocytes. Ephrin-B1 phosphorylation also disrupted the linkage of an apical transmembrane protein, podocalyxin, with NHERF2-ezrin-actin. The phosphorylation of ephrin-B1 and the consequent dephosphorylation of NHERF2 are critical initiation events leading to podocyte injury.

en: The American Journal of Pathology

ja: The American Journal of Pathology


en: FKBP12 は14-3-3，synaptopodin と結合し，ポドサイトのアクチン細胞骨格と細胞突起の維持に関与する


en: タクロリムスはアクチン細胞骨格のFKBP12 を保持する事でポドサイト傷害を軽減する


en: 糸球体におけるタクロリムス，シクロスポリンA の結合蛋白質の発現解析

ja: 糸球体におけるタクロリムス，シクロスポリンA の結合蛋白質の発現解析


en: ポドサイト傷害モデルの糸球体におけるFKBP12発現

ja: ポドサイト傷害モデルの糸球体におけるFKBP12発現


en: Progression of renal fibrosis in congenital CKD model rats with reduced number of nephrons

en: A congenital reduction in the number of nephrons is a critical risk factor for both onset of chronic kidney disease (CKD) and its progression to end-stage kidney disease (ESKD). Hypoplastic kidney (HPK) rats have only about 20% of the normal number of nephrons and show progressive CKD. This study used an immunohistological method to assess glomerular and interstitial pathogenesis in male HPK rats aged 35-210days. CD68 positive-macrophages were found to infiltrate into glomeruli in HPK rats aged 35 and 70days and to infiltrate into interstitial tissue in rats aged 140 and 210days. HPK rats aged 35 and 70days showed glomerular hypertrophy, loss of normal linear immunostaining of podocine, and increased expression of PDGFr-β, TGF-β, collagens, and fibronectin, with all of these alterations gradually deteriorating with age. α-SMA-positive myofibroblasts were rarely detected in glomerular tufts, whereas α-SMA-positive glomerular parietal epithelium (GPE) cells were frequently observed along Bowman's capsular walls. The numbers of PDGFr-β-positive fibroblasts in interstitial tissue were increased in rats aged 35days and older, whereas interstitial fibrosis, characterized by the increased expression of tubular PDGF-BB, the appearance of myofibroblasts doubly positive for PDGFr-β and α-SMA, and increased expression of collagens and fibronectin, were observed in rats aged 70 and older. These results clearly indicate that congenital CKD with only 20% of nephrons cause renal fibrosis in rats.

en: Experimental and Toxicologic Pathology

ja: Experimental and Toxicologic Pathology


en: Critical roles of Astrin in the mitosis of immature rat Sertoli cells

en: Male hypogonadism (hgn/hgn) rats show testicular hypoplasia accompanied by dysplastic development of seminiferous tubules due to loss-of-function mutation of the gene encoding Astrin, which is required for mitotic progression in the division cycle of HeLa cells. In the present study, we examined the cytological base leading to the decrease of Sertoli cells in hgn/hgn testes. In hgn/hgn testes on postnatal day 3, anti-phospho-histone H3 (Ser10) (pH3)-positive mitotic phase and TUNEL-positive apoptosis increased in GATA4-positive Sertoli cells. Isolated immature Sertoli cells from hgn/hgn testes showed increased pH3-assessed mitotic index accompanied by decreased 5-bromo-2'-deoxyuridine-incorporation and increased TUNEL-positive apoptosis, suggesting mitotic delay and cell death. In the visualization of mitotic progression by nocodazole (NOC)-mediated cell cycle arrest and subsequent release, hgn/hgn rat-derived Sertoli cells failed to make the transition from prometaphase to metaphase, and the cells with micronuclei and TUNEL-positive cells gradually increased in a time-dependent manner. Western blot analysis detected ≈142 kDa protein expected as Astrin in extracts of +/+ and +/hgn testes and cultured normal Sertoli cells but not in extracts of hgn/hgn testes. CLASP1 was detected in extracts of both normal and hgn/hgn testes, whereas it was localized in kinetochore of normal mitotic Sertoli cells but diffused in cytoplasm of hgn/hgn Sertoli cells. These results indicate that Astrin is required for normal mitotic progression in immature Sertoli cells and that the most severe type of testicullar dysplasia in hgn/hgn rats is caused by mitotic cell death of immature Sertoli cells due to lack of Astrin.

en: Biochemical and Biophysical Research Communications


en: Renal Function and Hematology in Rats with Congenital Renal Hypoplasia

en: Comparative Medicine

ja: Comparative Medicine


en: The microtubule-associated protein astrin transgenesis rescues spermatogenesis and renal function in hypogonadic (hgn/hgn) rats

en: Andrology

ja: Andrology