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Effects and Mechanism of lncRNA CRNDE on Sepsis-Induced Acute Kidney Injury.

ABSTRACT: Objective:To investigate the effects of lncRNA CRNDE on sepsis-associated acute kidney injury in the human kidney 2 cell line and explore the potential mechanisms. Methods:HK-2 cells were treated with lipopolysaccharides to induce injuries. The expression of CRNDE and miR-146a in HK-2 cells were altered by a transient transfection assay. Cell apoptosis was detected by a flow cytometry assay, and the levels of inflammatory cytokines including TNF-?, IL-6, IL-8, and IL-1? were assessed by ELISA. Furthermore, western blot analysis was performed to detect the expression levels of TLR4/NF-?B pathway-related proteins. And a luciferase reporter gene assay was used to verify if miR-146a was the target of CRNDE. Results:LPS treatment increased CRNDE expression in HK-2 cells. CRNDE overexpression enhanced cell injuries in HK-2 cells significantly increasing inflammatory cytokine levels, including TNF-?, IL-6, IL-8, and IL-1?, and cell apoptosis. In addition, CRNDE overexpression further activated the TLR4/NF-?B pathways in HK-2 cells. Inversely, opposite results were observed in the miR-146a mimic treatment group, and the miR-146a inhibitor could reverse the protein expression changes of TLR4/NF-?B in the si-CRNDE and LPS treatment group. Conclusion:This study demonstrated that CRNDE overexpression could activate the TLR4/NF-?B signaling pathway by regulating miR-146a, which accelerated LPS-induced inflammation and apoptosis in HK-2 cells.

SUBMITTER: Wu S

PROVIDER: S-EPMC7211233 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Effects and Mechanism of lncRNA CRNDE on Sepsis-Induced Acute Kidney Injury.

Wu Shen S Qiu Hengyi H Wang Qiao Q Cao Zheng Z Wang Jianmin J

Analytical cellular pathology (Amsterdam) 20200501

<h4>Objective</h4>To investigate the effects of lncRNA CRNDE on sepsis-associated acute kidney injury in the human kidney 2 cell line and explore the potential mechanisms.<h4>Methods</h4>HK-2 cells were treated with lipopolysaccharides to induce injuries. The expression of CRNDE and miR-146a in HK-2 cells were altered by a transient transfection assay. Cell apoptosis was detected by a flow cytometry assay, and the levels of inflammatory cytokines including TNF-<i>α</i>, IL-6, IL-8, and IL-1<i>β< ...[more]

PMID: 32399391

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Project description:Recent studies have shown that autophagy upregulation can attenuate sepsis-induced acute kidney injury (SAKI). The tumor suppressor p53 has emerged as an autophagy regulator in various forms of acute kidney injury (AKI). Our previous studies showed that p53 acetylation exacerbated hemorrhagic shock-induced AKI and lipopolysaccharide (LPS)-induced endothelial barrier dysfunction. However, the role of p53-regulated autophagy in SAKI has not been examined and requires clarification. In this study, we observed the dynamic changes of autophagy in renal tubular epithelial cells (RTECs) and verified the protective effects of autophagy activation on SAKI. We also examined the changes in the protein expression, intracellular distribution (nuclear and cytoplasmic), and acetylation/deacetylation levels of p53 during SAKI following cecal ligation and puncture (CLP) or LPS treatment in mice and in a LPS-challenged human RTEC cell line (HK-2 cells). After sepsis stimulation, the autophagy levels of RTECs increased temporarily, followed by a sharp decrease. Autophagy inhibition was accompanied by an increased renal tubular injury score. By contrast, autophagy agonists could reduce renal tubular damage following sepsis. Surprisingly, the expression of p53 protein in both the renal cortex and HK-2 cells did not significantly change following sepsis stimulation. However, the translocation of p53 from the nucleus to the cytoplasm increased, and the acetylation of p53 was enhanced. In the mechanistic study, we found that the induction of p53 deacetylation, due to either the resveratrol/quercetin -induced activation of the deacetylase Sirtuin 1 (Sirt1) or the mutation of the acetylated lysine site in p53, promoted RTEC autophagy and alleviated SAKI. In addition, we found that acetylated p53 was easier to bind with Beclin1 and accelerated its ubiquitination-mediated degradation. Our study underscores the importance of deacetylated p53-mediated RTEC autophagy in future SAKI treatments.

Project description:Sepsis-induced acute kidney injury (AKI) is a frequent complication of critically ill patients and leads to high mortality rates. The specificity of currently available urinary biomarkers for AKI in the context of sepsis is questioned. This study aimed to discover urinary biomarkers for septic AKI by contemporary shotgun proteomics in a mouse model for sepsis and to validate these in individual urine samples of mice and human septic patients with and without AKI. At 48 h after uterine ligation and inoculation of Escherichia coli, aged mice (48 weeks) became septic. A subgroup developed AKI, defined by serum creatinine, blood urea nitrogen, and renal histology. Separate pools of urine from septic mice with and without AKI mice were collected during 12 h before and between 36-48 h after infection, and their proteome compositions were quantitatively compared. Candidate biomarkers were validated by Western blot analysis of urine, plasma, and renal tissue homogenates from individual mice, and a limited number of urine samples from human septic patients with and without AKI. Urinary neutrophil gelatinase-associated lipocalin, thioredoxin, gelsolin, chitinase 3-like protein 1 and -3 (CHI3L3) and acidic mammalian chitinase were the most distinctive candidate biomarkers selected for septic AKI. Both neutrophil gelatinase-associated lipocalin and thioredoxin were detected in urine of septic mice and increased with severity of AKI. Acidic mammalian chitinase was only present in urine of septic mice with AKI. Both urinary chitinase 3-like protein 1 and -3 were only detected in septic mice with severe AKI. The human homologue chitinase 3-like protein 1 was found to be more excreted in urine from septic patients with AKI than without. In summary, urinary chitinase 3-like protein 1 and -3 and acidic mammalian chitinase discriminated sepsis from sepsis-induced AKI in mice. Further studies of human chitinase proteins are likely to lead to additional insights in septic AKI.

Dataset Information

Effects and Mechanism of lncRNA CRNDE on Sepsis-Induced Acute Kidney Injury.

Publications

Effects and Mechanism of lncRNA CRNDE on Sepsis-Induced Acute Kidney Injury.

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