Abstract P415: ACE N-domain Regulates High-glucose Mediated Interleukin-1 beta Production by Renal Epithelial Cells Independently From Angiotensin II Generation

Hypertension ◽  
2017 ◽  
Vol 70 (suppl_1) ◽  
Author(s):  
Jorge F Giani ◽  
Ellen A Bernstein ◽  
Masahiro Eriguchi ◽  
Romer A Gonzalez-Villalobos ◽  
Kenneth E Bernstein
Keyword(s):  
Diabetic Nephropathy ◽  
Angiotensin Ii ◽  
Epithelial Cells ◽  
High Glucose ◽  
Interleukin 1 ◽  
Kidney Injury ◽  
Fold Increase ◽  
Renal Epithelial Cells ◽  
Proximal Tubular ◽  
Sense And Respond

Research studies demonstrated that interleukin (IL)-1β contributes to the development of diabetic nephropathy and hypertension. However, the origin and regulation of IL-1β synthesis during diabetic kidney injury are still unknown. Here, we hypothesize that renal epithelial cells produce IL-1β in response to a high glucose stress and that angiotensin converting enzyme (ACE) plays a key role in this process. To study this, we isolated proximal tubular (PT) epithelial cells from wild-type (WT) and mice lacking either the ACE N-domain (NKO) or the C-domain (CKO) catalytic activity. These cells were exposed to normal (5 mM) or high (30 mM) glucose for 24 hours. IL-1β produced by PT cells were assessed by ELISA and RT-PCR. High glucose induced WT PT cells to release significant amounts of IL-1β (from 5±1 to 70±6 pg/ml, p<0.001; n=6). When WT PT cells were exposed to a high glucose media in the presence of an ACE inhibitor (lisinopril, 10 mM), IL-1β levels were significantly reduced (from 70±6 to 38±6 pg/ml, p<0.01). In contrast, AT1 receptor blockade by losartan did not change the amount of IL-1β produced by WT PT cells. To determine which ACE domain is associated with IL-1β production, NKO and CKO PT cells were exposed to high glucose. Strikingly, NKO PT cells released lower amounts of IL-1β when exposed to high glucose compared to WT (NKO: 15±7 vs. WT: 79±9 pg/ml, p<0.01, n=4). No differences were observed between WT and CKO PT cells. Since the ACE N-domain degrades the anti-inflammatory tetrapeptide N-acetyl-Ser-Asp-Lys-Pro (AcSDKP), we tested whether the lower IL-1β production in NKO PT cells was due to an accumulation of AcSDKP. For this, we pre-treated NKO PT cells with a prolyl endopeptidase inhibitor (S17092, 50μM) to prevent the production of AcSDKP. Notably, this treatment increased the IL-1β response to high glucose in NKO PT cells (2.1±0.3-fold increase, p<0.01, n=4). Our data indicate that: 1) PT cells can sense and respond to high glucose by secreting IL-1β and 2) the absence of the ACE N-domain blunts the production of IL-1β through a mechanism that involves AcSDKP accumulation. In conclusion, ACE might contribute to the inflammatory response that underlays diabetic nephropathy independently from angiotensin II generation.

Metabolic inhibition-induced transient Ca2+ increase depends on mitochondria in a human proximal renal cell line

2007 ◽  
Vol 293 (2) ◽  
pp. F533-F540 ◽  
Author(s):  
Adrian Caplanusi ◽  
Andrew J. Fuller ◽  
Romer A. Gonzalez-Villalobos ◽  
Timothy G. Hammond ◽  
L. G. Navar
Keyword(s):  
Epithelial Cells ◽  
Mdck Cells ◽  
Fold Increase ◽  
Recovery Phase ◽  
Metabolic Inhibition ◽  
Human Origin ◽  
Renal Epithelial Cells ◽  
Proximal Tubular ◽  
Renal Cell Line ◽  
Signaling Process

During ischemia or hypoxia an increase in intracellular cytosolic Ca2+ induces deleterious events but is also implicated in signaling processes triggered in such conditions. In MDCK cells (distal tubular origin), it was shown that mitochondria confer protection during metabolic inhibition (MI), by buffering the Ca2+ overload via mitochondrial Na+-Ca2+ exchanger (NCX). To further assess this process in cells of human origin, human cortical renal epithelial cells (proximal tubular origin) were subjected to MI and changes in cytosolic Ca2+ ([Ca2+]i), Na+, and ATP concentrations were monitored. MI was accomplished with both antimycin A and 2-deoxyglucose and induced a 3.5-fold increase in [Ca2+]i, reaching 136.5 ± 15.8 nM in the first 3.45 min. Subsequently [Ca2+]i dropped and stabilized to 62.7 ± 7.3 nM by 30 min. The first phase of the transient increase was La3+ sensitive, not influenced by diltiazem, and abolished when mitochondria were deenergized with the protonophore carbonylcyanide p-trifluoromethoxyphenylhydrazone. The subsequent recovery phase was impaired in a Na+-free medium and weakened when the mitochondrial NCX was blocked with 7-chloro-5-(2-chlorophenyl)-1,5-dihydro-4,1-benzothiazepin-2(3H)-one (CGP-37157). Thus Ca2+ entry is likely mediated by store-operated Ca2+ channels and depends on energized mitochondria, whereas [Ca2+]i recovery relied partially on the activity of mitochondrial NCX. These results indicate a possible mitochondrial-mediated signaling process triggered by MI, support the hypothesis that mitochondrial NCX has an important role in the Ca2+ clearance, and overall suggest that mitochondria play a preponderant role in the regulation of responses to MI in human renal epithelial cells.

scholarly journals LncRNA TUG1/miR-29c-3p/SIRT1 axis regulates endoplasmic reticulum stress-mediated renal epithelial cells injury in diabetic nephropathy model in vitro

PLoS ONE ◽  
2021 ◽  
Vol 16 (6) ◽  
pp. e0252761
Author(s):  
Shaoqiang Wang ◽  
Pengfei Yi ◽  
Na Wang ◽  
Min Song ◽  
Wenhui Li ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Diabetic Nephropathy ◽  
Epithelial Cells ◽  
Endoplasmic Reticulum Stress ◽  
High Glucose ◽  
Potential Role ◽  
Renal Epithelial Cells ◽  
Renal Tubular ◽  
Induced Apoptosis ◽  
Lncrna Tug1

Long non-coding RNAs (lncRNAs) are important regulators in diabetic nephropathy. In this study, we investigated the potential role of lncRNA TUG1 in regulating endoplasmic reticulum stress (ERS)-mediated apoptosis in high glucose induced renal tubular epithelial cells. Human renal tubular epithelial cell line HK-2 was challenged with high glucose following transfection with lncRNA TUG1, miR-29c-3p mimics or inhibitor expression plasmid, either alone or in combination, for different experimental purposes. Potential binding effects between TUG1 and miR-29c-3p, as well as between miR-29c-3p and SIRT1 were verified. High glucose induced apoptosis and ERS in HK-2 cells, and significantly decreased TUG1 expression. Overexpressed TUG1 could prevent high glucose-induced apoptosis and alleviated ERS via negatively regulating miR-29c-3p. In contrast, miR-29c-3p increased HK-2 cells apoptosis and ERS upon high glucose-challenge. SIRT1 was a direct target gene of miR-29c-3p in HK-2 cells, which participated in the effects of miR-29c-3p on HK-2 cells. Mechanistically, TUG1 suppressed the expression of miR-29c-3p, thus counteracting its function in downregulating the level of SIRT1. TUG1 regulates miR-29c-3p/SIRT1 and subsequent ERS to relieve high glucose induced renal epithelial cells injury, and suggests a potential role for TUG1 as a promising diagnostic marker of diabetic nephropathy.

scholarly journals High Glucose Increases Metallothionein Expression in Renal Proximal Tubular Epithelial Cells

2011 ◽  
Vol 2011 ◽  
pp. 1-8 ◽  
Author(s):  
Daisuke Ogawa ◽  
Masato Asanuma ◽  
Ikuko Miyazaki ◽  
Hiromi Tachibana ◽  
Jun Wada ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Diabetic Nephropathy ◽  
Vitamin E ◽  
Epithelial Cells ◽  
High Glucose ◽  
Male Rats ◽  
Tubular Epithelial Cells ◽  
Proximal Tubular Epithelial Cells ◽  
Proximal Tubular ◽  
Renal Proximal Tubular

Metallothionein (MT) is an intracellular metal-binding, cysteine-rich protein, and is a potent antioxidant that protects cells and tissues from oxidative stress. Although the major isoforms MT-1 and -2 (MT-1/-2) are highly inducible in many tissues, the distribution and role of MT-1/-2 in diabetic nephropathy are poorly understood. In this study, diabetes was induced in adult male rats by streptozotocin, and renal tissues were stained with antibodies for MT-1/-2. MT-1/-2 expression was also evaluated in mProx24 cells, a mouse renal proximal tubular epithelial cell line, stimulated with high glucose medium and pretreated with the antioxidant vitamin E. MT-1/-2 expression was gradually and dramatically increased, mainly in the proximal tubular epithelial cells and to a lesser extent in the podocytes in diabetic rats, but was hardly observed in control rats. MT-1/-2 expression was also increased by high glucose stimulation in mProx24 cells. Because the induction of MT was suppressed by pretreatment with vitamin E, the expression of MT-1/-2 is induced, at least in part, by high glucose-induced oxidative stress. These observations suggest that MT-1/-2 is induced in renal proximal tubular epithelial cells as an antioxidant to protect the kidney from oxidative stress, and may offer a novel therapeutic target against diabetic nephropathy.

scholarly journals Metallothionein deficiency exacerbates diabetic nephropathy in streptozotocin-induced diabetic mice

2014 ◽  
Vol 306 (1) ◽  
pp. F105-F115 ◽  
Author(s):  
Hiromi Tachibana ◽  
Daisuke Ogawa ◽  
Norio Sogawa ◽  
Masato Asanuma ◽  
Ikuko Miyazaki ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Diabetic Nephropathy ◽  
Epithelial Cells ◽  
High Glucose ◽  
Tubular Epithelial Cells ◽  
Diabetic Kidney ◽  
Proximal Tubular Epithelial Cells ◽  
Proximal Tubular ◽  
In The Diabetic Kidney

Oxidative stress and inflammation play important roles in diabetic complications, including diabetic nephropathy. Metallothionein (MT) is induced in proximal tubular epithelial cells as an antioxidant in the diabetic kidney; however, the role of MT in renal function remains unclear. We therefore investigated whether MT deficiency accelerates diabetic nephropathy through oxidative stress and inflammation. Diabetes was induced by streptozotocin injection in MT-deficient (MT−/−) and MT+/+ mice. Urinary albumin excretion, histological changes, markers for reactive oxygen species (ROS), and kidney inflammation were measured. Murine proximal tubular epithelial (mProx24) cells were used to further elucidate the role of MT under high-glucose conditions. Parameters of diabetic nephropathy and markers of ROS and inflammation were accelerated in diabetic MT−/− mice compared with diabetic MT+/+ mice, despite equivalent levels of hyperglycemia. MT deficiency accelerated interstitial fibrosis and macrophage infiltration into the interstitium in the diabetic kidney. Electron microscopy revealed abnormal mitochondrial morphology in proximal tubular epithelial cells in diabetic MT−/− mice. In vitro studies demonstrated that knockdown of MT by small interfering RNA enhanced mitochondrial ROS generation and inflammation-related gene expression in mProx24 cells cultured under high-glucose conditions. The results of this study suggest that MT may play a key role in protecting the kidney against high glucose-induced ROS and subsequent inflammation in diabetic nephropathy.

scholarly journals LincRNA-p21 Inhibits Cisplatin-Induced Apoptosis of Human Renal Proximal Tubular Epithelial Cells by Sponging miR-449a

2021 ◽  
pp. 1-7
Author(s):  
Zhen Li ◽  
Gang Hou
Keyword(s):  
Breast Cancer ◽  
Epithelial Cells ◽  
Kidney Injury ◽  
Protective Role ◽  
Tubular Epithelial Cells ◽  
Breast Cancer Patients ◽  
Proximal Tubular Epithelial Cells ◽  
Proximal Tubular ◽  
Renal Proximal Tubular ◽  
Induced Apoptosis

<b><i>Introduction:</i></b> LincRNA-p21 is predicted to interact with miR-449a, which plays a protective role in cisplatin-induced acute kidney injury (CIA). <b><i>Objective:</i></b> This study aimed to analyze the involvement of lincRNA-p21 in breast cancer patients with CIA. <b><i>Methods:</i></b> Levels of lincRNA-p21 in plasma from CIA, triple negative breast cancer, and control groups were measured by performing RT-qPCR. The potential interaction between lincRNA-p21 and miR-449a was first predicted by RT-qPCR. The relationship between lincRNA-p21 and miR-449a was analyzed by overexpression experiment. <b><i>Results:</i></b> We found that lincRNA-p21 is downregulated in CIA. Dual luciferase activity assay showed that lincRNA-p21 and miR-449a can interact with each other, while overexpression of lincRNA-p21 and miR-449a failed to affect the expression of each other. In human renal proximal tubular epithelial cells (HRPTEpCs), cisplatin led to the upregulated miR-449a but downregulated lincRNA-p21. Interestingly, lincRNA-p21 overexpression led to reduced enhancing effects of miR-449a on the cisplatin-induced apoptosis of HRPTEpCs. <b><i>Conclusion:</i></b> Therefore, lincRNA-p21 is downregulated in CIA and may sponge miR-449a to inhibit cisplatin-induced apoptosis of HRPTEpCs.

scholarly journals Proximal tubule-specific overexpression of netrin-1 suppresses acute kidney injury-induced interstitial fibrosis and glomerulosclerosis through suppression of IL-6/STAT3 signaling

2013 ◽  
Vol 304 (8) ◽  
pp. F1054-F1065 ◽  
Author(s):  
Punithavathi Ranganathan ◽  
Calpurnia Jayakumar ◽  
Ganesan Ramesh
Keyword(s):  
Acute Kidney Injury ◽  
Epithelial Cells ◽  
Interstitial Fibrosis ◽  
Kidney Injury ◽  
Transgenic Animals ◽  
Acute Injury ◽  
Tubular Epithelial Cells ◽  
Wild Type ◽  
Proximal Tubular Epithelial Cells ◽  
Proximal Tubular

Acute kidney injury-induced organ fibrosis is recognized as a major risk factor for the development of chronic kidney disease, which remains one of the leading causes of death in the developed world. However, knowledge on molecules that may suppress the fibrogenic response after injury is lacking. In ischemic models of acute kidney injury, we demonstrate a new function of netrin-1 in regulating interstitial fibrosis. Acute injury was promptly followed by a rise in serum creatinine in both wild-type and netrin-1 transgenic animals. However, the wild-type showed a slow recovery of kidney function compared with netrin-1 transgenic animals and reached baseline by 3 wk. Histological examination showed increased infiltration of interstitial macrophages, extensive fibrosis, reduction of capillary density, and glomerulosclerosis. Collagen IV and α-smooth muscle actin expression was absent in sham-operated kidneys; however, their expression was significantly increased at 2 wk and peaked at 3 wk after reperfusion. These changes were reduced in the transgenic mouse kidney, which overexpresses netrin-1 in proximal tubular epithelial cells. Fibrosis was associated with increased expression of IL-6 and extensive and chronic activation of STAT3. Administration of IL-6 exacerbated fibrosis in vivo in wild-type, but not in netrin-1 transgenic mice kidney and increased collagen I expression and STAT3 activation in vitro in renal epithelial cells subjected to hypoxia-reoxygenation, which was suppressed by netrin-1. Our data suggest that proximal tubular epithelial cells may play a prominent role in interstitial fibrosis and that netrin-1 could be a useful therapeutic agent for treating kidney fibrosis.

scholarly journals P2X7receptors mediate deleterious renal epithelial-fibroblast cross talk

2011 ◽  
Vol 300 (1) ◽  
pp. F62-F70 ◽  
Author(s):  
Murugavel Ponnusamy ◽  
Li Ma ◽  
Rujun Gong ◽  
Maoyin Pang ◽  
Y. Eugene Chin ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Pyridoxal Phosphate ◽  
Purinergic Receptor ◽  
Purinergic Signaling ◽  
Kidney Injury ◽  
Fibroblast Cell ◽  
Underlying Mechanism ◽  
Disulfonic Acid ◽  
Small Interference Rna ◽  
Renal Epithelial Cells

Peritubular fibroblasts in the kidney are the major erythropoietin-producing cells and also contribute to renal repair following acute kidney injury (AKI). Although few fibroblasts were observed in the interstitium adjacent to damaged tubular epithelium in the early phase of AKI, the underlying mechanism by which their numbers were reduced remains unknown. In this study, we tested the hypothesis that damaged renal epithelial cells directly induce renal interstitial fibroblast death by releasing intracellular ATP and activating purinergic signaling. Exposure of a cultured rat renal interstitial fibroblast cell line (NRK-49F) to necrotic renal proximal tubular cells (RPTC) lysate or supernatant induced NRK-49F cell death by apoptosis and necrosis. Depletion of ATP with apyrase or inhibition of the P2X purinergic receptor with pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid blocked the deleterious effect of necrotic RPTC supernatant. The P2X7receptor, an ATP-sensitive purinergic receptor, was not detected in cultured NRK-49F cells but was inducible by necrotic RPTC supernatant. Treatment with A438079, a highly selective P2X7receptor inhibitor, or knockdown of the P2X7receptor with small interference RNA diminished renal fibroblast death induced by necrotic RPTC supernatant. Conversely, overexpression of the P2X7receptor potentiated this response. Collectively, these findings provide strong evidence that damaged renal epithelial cells can directly induce the death of renal interstitial fibroblasts by ATP activation of the P2X7receptor.

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