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Proximal Tubular Epithelial Cells (proximal + tubular_epithelial_cell)
Selected AbstractsOverexpression of RGPR-p117 enhances regucalcin gene promoter activity in cloned normal rat kidney proximal tubular epithelial cells: Involvement of TTGGC motifJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2006Natsumi Sawada Abstract A novel protein RGPR-p117 was discovered as regucalcin gene promoter region-related protein that binds to the TTGGC motif using a yeast one-hybrid system. RGPR-p117 is localized in the nucleus of kidney cells, and overexpression of RGPR-p117 can modulate regucalcin protein and its mRNA expression in the cloned normal rat kidney proximal tubular epithelial NRK52E cells. This study was undertaken to determine whether overexpression of RGPR-p117 enhances the regucalcin promoter activity using the ,710/+18 LUC construct (wild-type) or ,710/+18 LUC construct (mutant) with deletion of ,523/,435 including TTGGC motif. NRK52E cells (wild-type) or stable HA-RGPR-p117/phCMV2-transfected cells (transfectant) were cultured in Dulbecco's minimum essential medium (DMEM) containing 5% bovine serum (BS). Wild-type cells or transfectants were transfected with the ,710/+18 LUC construct vector or the ,710/+18 LUC construct with deletion of ,523/,435. Wild-type cells or transfectants with subconfluency were cultured for 48 h in a DMEM medium containing either vehicle, BS (5%), or parathyroid hormone (1,34) (PTH; 10,7 M). Luciferase activity in wild-type cells was significantly increased with culture of BS or PTH. This increase was significantly blocked in the presence of various protein kinase inhibitors (staurosporine and PD 98059). Luciferase activity in transfectants was significantly increased as compared with that of wild-type cells in the absence of BS or PTH. The increase in luciferase activity in transfectants was completely decreased in mutant with deletion of ,523/,435 sequence of regucalcin promoter. This was also seen using the ,710/+18 LUC construct with deletion of ,523/,503 sequence containing TTGGC motif. The increase in luciferase activity in transfectants was not significantly enhanced with culture of BS (5%), PTH (10,7 M), Bay K 8644 (10,6 M), phorbol 12-myristate 13-acetate (PMA; 10,6 M), or N6, 2,-dibutyryl cyclic adenosine 3,, 5,-monophosphate (DcAMP; 10,4 M). The increase in luciferase activity in transfectants was completely inhibited with culture of dibucaine (10,6 M), staurosporine (10,9 M), PD 98059 (10,8 M), wortmannin (10,8 M), genistein (10,6 M), vanadate (10,6 M), or okadaic acid (10,6 M) which are inhibitors of various kinases and protein phosphatases. This study demonstrates that RGPR-p117 can enhance the regucalcin promoter activity which is related to the NF-1 consensus sequences including TTGGC motif, and that its enhancing effect is partly mediated through phosphorylation and dephosphorylation in NRK52E cells. J. Cell. Biochem. 99: 589,597, 2006. © 2006 Wiley-Liss, Inc. [source] miR-17, miR-19b, miR-20a, and miR-106a are down-regulated in human agingAGING CELL, Issue 2 2010Matthias Hackl Summary Aging is a multifactorial process where deterioration of body functions is driven by stochastic damage while counteracted by distinct genetically encoded repair systems. To better understand the genetic component of aging, many studies have addressed the gene and protein expression profiles of various aging model systems engaging different organisms from yeast to human. The recently identified small non-coding miRNAs are potent post-transcriptional regulators that can modify the expression of up to several hundred target genes per single miRNA, similar to transcription factors. Increasing evidence shows that miRNAs contribute to the regulation of most if not all important physiological processes, including aging. However, so far the contribution of miRNAs to age-related and senescence-related changes in gene expression remains elusive. To address this question, we have selected four replicative cell aging models including endothelial cells, replicated CD8+ T cells, renal proximal tubular epithelial cells, and skin fibroblasts. Further included were three organismal aging models including foreskin, mesenchymal stem cells, and CD8+ T cell populations from old and young donors. Using locked nucleic acid-based miRNA microarrays, we identified four commonly regulated miRNAs, miR-17 down-regulated in all seven; miR-19b and miR-20a, down-regulated in six models; and miR-106a down-regulated in five models. Decrease in these miRNAs correlated with increased transcript levels of some established target genes, especially the cdk inhibitor p21/CDKN1A. These results establish miRNAs as novel markers of cell aging in humans. [source] Functional analysis of polyomavirus BK non-coding control region quasispecies from kidney transplant recipientsJOURNAL OF MEDICAL VIROLOGY, Issue 11 2009Gunn-Hege Olsen Abstract Replication of the human polyomavirus BK (BKV) in renal tubular epithelial cells causes viruria and BKV-nephropathy in kidney transplant recipients. Following prolonged high-level BKV replication, rearrangement of the archetype non-coding control region (NCCR) leads to a mixture of BKV variants. The aim of this study was to compare potential functional differences of 12 rearranged (rr)-NCCR variants with the archetype (ww)-NCCR (WWT) found in allograft biopsies or urine from three kidney transplant recipients including two with BKV-nephropathy. Twelve different rr-NCCRs and one archetype ww-NCCR were inserted between the early and late protein coding region of BKV(Dunlop) to make recombinant BKV genomes for transfection into Vero cells. Immunoblotting, immunofluorescence staining, and quantitative PCR demonstrated that viral protein expression and extracellular BKV loads of 10 rr-NCCR variants were similar or higher than observed for the ww-NCCR BKV. Two rr-NCCR variants (RH-2 and RH-19) were non-functional. The functional rr-NCCRs produced infectious progeny successfully infecting primary renal proximal tubular epithelial cells. The number of infected cells and extracellular BKV loads corresponded to the activity seen in Vero cells. Three rr-NCCR variants (RH-1, RH-10, RH-13) only gave rise to a few infected cells similar to ww-NCCR, whereas seven variants had intermediate activity (RH-5, RH-6, RH-8, RH-9, RH-11) or high replication activity (RH-7 and RH-18) with several hundred infected cells per well. The results indicate that both functional and non-functional BKV rr-NCCR variants arise during BKV replication in kidney transplant recipients and that most functional rr-NCCR variants confer a higher replication capacity than archetype ww-NCCR. J. Med. Virol. 81:1959,1967, 2009. © 2009 Wiley-Liss, Inc. [source] Aldosterone induces collagen synthesis via activation of extracellular signal-regulated kinase 1 and 2 in renal proximal tubulesNEPHROLOGY, Issue 8 2008GUOSHUANG XU SUMMARY: Aim: Aldosterone plays a crucial role in renal fibrosis by inducing mesangial cell proliferation and promoting collagen synthesis in renal fibroblasts. However, renal proximal tubule involvement in aldosterone-induced collagen synthesis has not yet been identified. The aim of this study was to examine the potential role of aldosterone in collagen expression and its possible mineralocorticoid receptor (MR)-dependent pathway, mediated by activation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) in cultured human renal proximal tubular epithelial (HKC) cells. Methods: After HKC cells were stimulated by aldosterone with different concentrations for various time and periods, the gene expression and protein synthesis of collagen I, II, III and IV were measured by real-time polymerase chain reaction and western blot, respectively. ERK1/2 activation, ,-smooth muscle actin (,-SMA), and E-cadherin were also detected by western blot. Results: Aldosterone can increase ERK1/2 phosphorylation of human renal proximal tubular epithelial cells in a time- and dose-dependent manner. Although aldosterone had no effect on collagen I and II expression, it increased expression of ,-SMA and collagen III and IV and decreased that of E-cadherin in HKC cells after 48 h. These effects could be prevented by a ERK pathway inhibitor, U0126, or by a selective MR antagonist, spironolactone. Conclusion: The results suggest that aldosterone plays a pivotal role in tubulointerstitial fibrosis by promoting tubular epithelial,mesenchymal transition and collagen synthesis in proximal tubular cells. The process is MR-dependent, and mediated by ERK1/2 mitogen-activated protein kinase pathway. [source] Effects of interleukin 18 on injury and activation of human proximal tubular epithelial cellsNEPHROLOGY, Issue 1 2007DONG LIANG SUMMARY: Background/Aims: Injury and activation of tubular proximal epithelial cells (TEC) play central roles in renal tubulointerstitial fibrosis (TIF), but its mechanisms remain obscure. Interleukin 18 (IL-18) is overproduced during chronic kidney diseases (CKD), but how IL-18 affects the biological behaviour of TEC is not clear. The aim of the present study is to reveal the role of IL-18 in renal TIF. Methods: The expressions of IL-18 and IL-18 receptor in TEC were detected by immunohistochemical staining in vivo and by reverse transcriptase polymerase chain reaction (RT-PCR) in vitro. TEC line (HK-2 cells) were incubated without or with IL-18. Cell proliferation and cell cycle were evaluated by methyl thiazolyl tetrazolium assay and flow cytometric analysis, respectively. Cell apoptosis was assessed by Hoechst 33258 staining. Expression of ,-smooth muscle actin was evaluated by RT-PCR, immunocytochemical staining and flow cytometric analysis, respectively. Type I collagen, fibronectin, MCP-1 and RANTES in cultured supernatants were measured by enzyme-linked immunosorbent assay. Results: IL-18 expression in TEC increased significantly in CKD state. IL-18 receptor was constitutively expressed in normal proximal TEC, and its expression increased strongly in CKD state. Proliferation and cell cycle of HK-2 cells were not affected by IL-18. Cell apoptosis, ,-smooth muscle actin expression, type I collagen and fibronectin production as well as MCP-1 secretion were promoted by IL-18 in dosage- and/or time-dependent manners, but RANTES secretion was not affected. Conclusion: IL-18 may play a crucial role in the process of TIF by promoting TEC injury and activation, and could be a target of the therapeutic approaches against TIF. [source] Thymoquinone decreases AGE-induced NF- ,B activation in proximal tubular epithelial cellsPHYTOTHERAPY RESEARCH, Issue 9 2007Ahmed Amir Radwan Sayed Abstract The inhibitory effects of thymoquinone (TQ) on activation of the redox-sensitive transcription factor nuclear factor kappa B (NF- ,B) and interleukin-6 (IL-6) were studied in vitro. Human proximal tubular epithelial cells (pTECs) were cultivated and stimulated with advanced glycation end products (AGEs) and the effects of TQ were studied. A significant reduction of AGE-induced NF- ,B-activation and Il-6 expression was observed. This points to potential antioxidative qualities of TQ. Copyright © 2007 John Wiley & Sons, Ltd. [source] Thymoquinone protects renal tubular cells against tubular injuryCELL BIOCHEMISTRY AND FUNCTION, Issue 3 2008Ahmed Amir Radwan Sayed Abstract In this work the effect of angiotensin II (AT II) on proximal tubular epithelial cells (pTECs) in vitro was studied. AT II was found to activate the nuclear factor ,B (NF- ,B) and its controlled genes, for example, interleukin 6 (IL-6) of pTECs in a time-dependent manner. Two points with maximum NF- ,B activation were found, the first after 12,h and the second after 3.5 days. The first point may be due to activation of NF- ,B in pTECs in response to AT II while the second may be due to activation of the advanced glycation end product (AGE)/receptor of the AGE (RAGE) system. Thymoquinone (TQ) was found to decrease NF- ,B activation in a dose-dependant manner with maximum inhibitory effect at a concentration of 500,nM. Also, pre-incubation of pTECs with TQ leads to disappearance of the second peak of NF- ,B. These data are consistent with results obtained from IL-6 enzyme-linked immunosorbent assay (ELISA) and transient transfection experiments. The results explain the therapeutic value of TQ which can be used to delay end stage renal diseases in diabetics. Copyright © 2008 John Wiley & Sons, Ltd. [source] | |||||||||||||