Search

G-protein-coupled receptor 40 (GPR40) has an anti-apoptotic effect in pancreatic β-cells. However, its role in renal tubular cell apoptosis remains unclear. To explore the role of GPR40 in renal tubular apoptosis, a two-week unilateral ureteral obstruction (UUO) mouse model was used. The protein expression of GPR40 was decreased, while the Bax/Bcl-2 protein expression ratio, the expression of tumor necrosis factor (TNF)-α mRNA, and angiotensin II type 1 receptor (AT1R) protein were increased in mice with UUO. In vitro, pretreatment of rat proximal tubular (NRK52E) cells with GW9508, a GPR40 agonist, attenuated the decreased cell viability, increased the Bax/Bcl-2 protein expression ratio, increased protein expression of cleaved caspase-3 and activated the nuclear translocation of nuclear factor-κB (NF-κB) p65 subunit induced by TNF-α treatment. TNF-α treatment significantly increased the expression of AT1R protein and the generation of reactive oxygen species (ROS), whereas GW9508 treatment markedly reversed these effects. Pretreatment with GW1100, a GPR40 antagonist, or silencing of GPR40 in NRK52E cells promoted the increased expression of the cleaved caspase-3 protein by TNF-α treatment. Our results demonstrate that decreased expression of GPR40 is associated with apoptosis via TNF-α and AT1R in the ureteral obstructed kidney. The activation of GPR40 attenuates TNF-α-induced apoptosis by inhibiting AT1R expression and ROS generation through regulation of the NF-κB signaling pathway.

The p-cresyl sulfate accumulates in kidney disease and may be involved in renal injury. α-Lipoic acid (α-LA) acts as an antioxidant in cell injury. We investigated the effects of α-LA treatment on p-cresyl sulfate-induced renal tubular injury. p-Cresyl sulfate induced cell death, and increased Bax/Bcl-2, cleaved caspase-3, Beclin-1, and LC3BII/LC3BI in human renal proximal tubular epithelial (HK-2) cells, which was counteracted by α-LA treatment. p-Cresyl sulfate-induced apoptosis was reduced by autophagy inhibitor 3-methyladenine, and p-cresyl sulfate induced autophagy was reduced by pan-caspase inhibitor Z-VAD-FMK. Moreover, p-cresyl sulfate treatment increased the expression of ER stress proteins and decreased the expression of baculoviral IAP repeat-containing proteins 6; these effects were prevented by α-LA treatment. Apoptosis and autophagy were associated with the phosphorylation of mitogen-activated protein kinase and nuclear translocation of the nuclear factor-κB p65 subunit. Pretreatment inhibitors of p38 and JNK, and knockdown of ATF4 gene reduced apoptosis- and autophagy-related protein expressions in p-cresyl sulfate treated HK-2 cells. These results demonstrate that α-lipoic acid attenuated p-cresyl sulfate-induced cell death by suppression of apoptosis and autophagy via regulation of ER stress in HK-2 cells., (Copyright © 2019 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Background/aims: Indoxyl sulfate (IS) is a uremic toxin and an important causative factor in the progression of chronic kidney disease. Recently, paricalcitol (19-nor-1,25-dihydroxyvitamin D2) was shown to exhibit protective effects in kidney injury. Here, we investigated the effects of paricalcitol treatment on IS-induced renal tubular injury., Methods: The fluorescent dye 2',7'-dichlorofluorescein diacetate was used to measure intracellular reactive oxygen species (ROS) following IS administration in human renal proximal tubular epithelial (HK-2) cells. The effects of IS on cell viability were determined using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays and levels of apoptosis-related proteins (Bcl-2-associated protein X [Bax] and B-cell lymphoma 2 [Bcl-2]), nuclear factor-κB (NF- κB) p65, and phosphorylation of mitogen-activated protein kinase (MAPK) and protein kinase B (Akt) were determined by semiquantitative immunoblotting. The promoter activity of NF-κB was measured by luciferase assays and apoptosis was determined by f low cytometry of cells stained with f luorescein isothiocyanate-conjugated Annexin V protein., Results: IS treatment increased ROS production, decreased cell viability and induced apoptosis in HK-2 cells. IS treatment increased the expression of apoptosis-related protein Bax, decreased Bcl-2 expression, and activated phosphorylation of MAPK, NF-κB p65, and Akt. In contrast, paricalcitol treatment decreased Bax expression, increased Bcl-2 expression, and inhibited phosphorylation of MAPK, NF-κB p65, and Akt in HK-2 cells. NF-κB promoter activity was increased following IS, administration and was counteracted by pretreatment with paricalcitol. Additionally, flow cytometry analysis revealed that IS-induced apoptosis was attenuated by paricalcitol treatment, which resulted in decreased numbers of fluorescein isothiocyanate-conjugated Annexin V positive cells., Conclusion: Treatment with paricalcitol inhibited IS-induced apoptosis by regulating MAPK, NF-κB, and Akt signaling pathway in HK-2 cells.

Background: Nicotine is, to a large extent, responsible for smoking-mediated renal dysfunction. This study investigated nicotine's effects on renal tubular epithelial cell apoptosis in vitro and it explored the mechanisms underlying its effects., Methods: Human proximal tubular epithelial (HK-2) cells were treated with nicotine. Cell viability was examined by using the WST-1 assay. Intracellular levels of reactive oxygen species (ROS) and the expression of mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) proteins were determined. The messenger ribonucleic acid and the protein expression associated with the nicotine acetylcholine receptors (nAChRs) in HK-2 cells was examined, and apoptosis was detected using flow cytometry, cell cycle analysis, and immunoblot analysis., Results: The HK-2 cells were endowed with nAChRs. Nicotine treatment reduced cell viability dose dependently, increased ROS levels, and increased extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK expression. Nicotine increased NF-κB activation, which was attenuated by N-acetyl-L-cysteine, and ERK and JNK inhibitors, but was not affected by a p38 MAPK inhibitor. Nicotine increased the Bax/Bcl-2 ratio, which was attenuated by N-acetyl-L-cysteine, the NF-κB inhibitor, Bay 11-7082, and hexamethonium, a non-specific nAChR blocker. Flow cytometry revealed nicotine-induced G2/M phase arrest. While nicotine treatment increased the expression of phosphorylated cdc2 and histone H3, a marker of G2/M phase arrest, hexamethonium and Bay 11-7082 pretreatment reduced their expression., Conclusions: Nicotine caused apoptosis in HK-2 cells by inducing ROS generation that activated the NF-κB signaling pathway via the MAPK pathway and it arrested the cell cycle at the G2/M phase. Nicotine-induced apoptosis in HK-2 cells involves the nAChRs.

G-protein-coupled receptor 40 (GPR40) is known to play a role in the regulation of fatty acids, insulin secretion and inflammation. However, the pathophysiological roles of GPR40 in kidney disease have not yet been identified. In the present study, we investigated the expression of GPR40 during cisplatin-induced kidney injury using male Sprague-Dawley rats that were treated with 8 mg/kg cisplatin. Control rats were treated with saline. Following treatment with cisplatin, the protein expression of GPR40 in the kidneys was decreased in association with an increase in serum creatinine levels and the Bax/Bcl-2 expression ratio. To further investigate the function of GPR40, the human renal proximal tubule epithelial cell line (HK-2) was cultured with cisplatin in the absence or presence of GW9508, a selective GPR40 agonist. Pre-treatment of the HK-2 cells with GW9508 attenuated the decrease in cell viability induced by treatment with cisplatin. Treatment with cisplatin increased the number of cells with condensed nuclei, which was ameliorated by GW9508 pre-treatment. TUNEL assay also revealed that pre-treatment with GW9508 ameliorated cisplatin-induced apoptosis. Treatment with cisplatin increased the Bax/Bcl-2 expression ratio and cleaved caspase-3 expression, and promoted the activation of nuclear factor-κB (NF-κB). These changes were attenuated by pre-treatment with GW9508. The cisplatin-induced generation of reactive oxygen species (ROS) and the activation of the Src/epidermal growth factor receptor (EGFR)/extracellular signal-regulated kinase (ERK) pathway were also counteracted by pre-treatment with GW9508. Thus, the activation of GPR40 attenuates cisplatin-induced apoptosis by inhibiting the generation of ROS, the activation of the Src/EGFR/ERK signaling pathway and the nuclear activation of NF-κB and pro-apoptotic factors.

Inflammation and apoptosis are the major contributors to the mechanisms of acute kidney injury (AKI) due to renal ischemia-reperfusion injury (IRI). Maslinic acid (MA), a pentacyclic triterpene acid mostly found in dietary plants, the current study was to demonstrate the renoprotective effect of MA on IRI-induced AKI, and to investigate the role of inflammation and apoptosis-related signaling pathways as a molecular mechanism. C57BL/6J mice were subjected to IRI for 72 h, and MA was daily administered by intraperitoneal injection during this period. In parallel, rat renal proximal tubule cells (NRK52E) were prophylactically treated with MA and then exposed to hydrogen peroxide (H2O2). MA treatment significantly inhibited the mRNA expression of interleukin (IL-1β), tumor necrosis factor-α (TGF-α), monocyte chemoattractant protein-1 (MCP-1), and intercellular adhesion molecule-1(ICAM-1). Also, MA reduced the expression of Bax/Bcl2 ratio and cleaved caspase-3. In NRK52 cells, MA inhibited the IκBα degradation, blocked NF-κB/p65 phosphorylation, and nuclear translocation. The phosphorylation of ERK, JNK, and p38 was attenuated by MA in IRI-induced kidney injury and H2O2-stimulated NRK52 cells. The expression levels of IL-1β, MCP-1, and ICAM-1 were upregulated in H2O2-stimulated NRK52E cells, which was attenuated by NF-κB inhibitor. H2O2 treatment increased the Bax/Bcl2 ratio and cleaved caspase-3 in NRK52E cells, which was counteracted by MAPK inhibitors. Together, our data demonstrate that MA suppresses IR-induced AKI injury through NF-κB and MAPK signaling pathways and that MA is a promising agent in the treatment of kidney diseases. [ABSTRACT FROM AUTHOR]

This study aimed to examine the protective effects of aliskiren on gentamicin-induced nephropathy. Rats were injected with gentamicin (100 mg/kg per day) for 14 days. Aliskiren was infused for two weeks. Human proximal tubular epithelial cell lines (HK-2) were cultured with gentamicin in the absence or presence of aliskiren. Inflammatory profibrotic and apoptotic markers were evaluated in vivo and in vitro. Aliskiren treatment attenuated the decreased creatinine clearance, increased fractional sodium excretion, glomerulosclerosis and tubulointerstitial fibrosis and counteracted the increased ED-1 expression in gentamicin-treated rats. The levels of inflammatory cytokines (TNF-α, IL-1β and IFN-γ) and adhesion molecules (MCP-1, ICAM-1 and VCAM-1) increased in the gentamicin-treated kidneys. These changes were restored by aliskiren co-treatment. Aliskiren effectively reversed transforming growth factor-β-induced fibrotic responses such as induction of α-smooth muscle actin in gentamicin-treated rat kidneys. Along with these changes, aliskiren also attenuated the increase in nuclear factor κB and phosphorylated extracellular signal-regulated kinase (pERK 1/2) levels in HK-2 cells cultured with gentamicin. In addition, aliskiren decreased the number of TUNEL-positive nuclei and reduced the expression of proapoptotic markers in gentamicin-treated HK-2 cells. These findings suggest that aliskiren attenuates gentamicin-induced nephropathy by suppression of inflammatory, profibrotic and apoptotic factors through inhibition of the nuclear factor κB, Smads and mitogen-activated protein kinase signaling pathways. [ABSTRACT FROM PUBLISHER]