长链非编码 RNA PVT1 通过靶向 miR-17-5p 并调控 NF-κB 通路促进脂多糖诱导的脓毒症急性肾损伤。
LncRNA PVT1 accelerates LPS-induced septic acute kidney injury through targeting miR-17-5p and regulating NF-κB pathway.
机构信息
Emergency Department, Jingzhou Central Hospital, The Second Clinical Medical College, Yangtze University, Jingzhou, No. 1 Renmin Road, Jingzhou, 433000, Hubei, China.
ICU Department, Jingzhou Central Hospital, The Second Clinical Medical College, Yangtze University, Jingzhou, Hubei, China.
出版信息
Int Urol Nephrol. 2021 Nov;53(11):2409-2419. doi: 10.1007/s11255-021-02905-8. Epub 2021 Jun 5.
BACKGROUND
Long noncoding RNA PVT1 is associated with diverse human diseases, including acute kidney injury (AKI). However, our understandings of PVT1 on septic AKI are limited.
METHODS
The septic AKI model was constructed through lipopolysaccharide (LPS) treatment. PVT1 and miR-17-5p levels were measured using qRT-PCR analysis. The concentrations of inflammatory cytokines were determined with ELISA kits. Cell viability and apoptosis were assessed using CCK-8 assay and flow-cytometric analysis, respectively. Protein levels were examined using western blot assay. The targeting association between miR-17-5p and PVT1 was verified by dual-luciferase reporter, RIP and RNA pull-down assays.
RESULTS
PVT1 level was elevated and miR-17-5p level was declined in septic AKI patients' serum and LPS-stimulated HK-2 cells. Cell viability was suppressed and cell apoptosis and inflammation were promoted after LPS treatment. PVT1 knockdown or miR-17-5p elevation restored LPS-mediated HK-2 cell injury. MiR-17-5p was sponged by PVT1, and its inhibition weakened the impact of PVT1 deficiency on LPS-mediated injury of HK-2 cells. In addition, PVT1 knockdown inactivated NF-κB pathway mediated by LPS treatment, but miR-17-5p inhibition further reversed this effect.
CONCLUSION
PVT1 knockdown promoted cell viability, suppressed inflammatory response and apoptosis by regulating miR-17-5p expression and NF-κB pathway in LPS-stimulated HK-2 cells.
背景
长链非编码 RNA PVT1 与多种人类疾病相关，包括急性肾损伤（AKI）。然而，我们对 PVT1 在脓毒症 AKI 中的作用了解有限。
方法
通过脂多糖（LPS）处理构建脓毒症 AKI 模型。采用 qRT-PCR 分析检测 PVT1 和 miR-17-5p 的水平。通过 ELISA 试剂盒测定炎症细胞因子的浓度。分别通过 CCK-8 测定和流式细胞术分析评估细胞活力和细胞凋亡。使用 Western blot 检测蛋白水平。通过双荧光素酶报告、RIP 和 RNA 下拉实验验证 miR-17-5p 与 PVT1 之间的靶向关系。
结果
脓毒症 AKI 患者血清和 LPS 刺激的 HK-2 细胞中 PVT1 水平升高，miR-17-5p 水平降低。LPS 处理后细胞活力受到抑制，细胞凋亡和炎症反应增强。LPS 介导的 HK-2 细胞损伤后，PVT1 敲低或 miR-17-5p 上调得到恢复。miR-17-5p 被 PVT1 海绵吸收，其抑制作用削弱了 PVT1 缺乏对 LPS 介导的 HK-2 细胞损伤的影响。此外，PVT1 敲低抑制了 LPS 处理介导的 NF-κB 通路，但 miR-17-5p 抑制进一步逆转了这一作用。
结论
PVT1 敲低通过调节 LPS 刺激的 HK-2 细胞中 miR-17-5p 的表达和 NF-κB 通路，促进细胞活力，抑制炎症反应和细胞凋亡。
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