MicroRNA-5010-5p 通过调节 PPP2R2D 的表达改善高葡萄糖诱导的肾小管上皮细胞炎症

IF 3.7 2区 医学 Q2 ENDOCRINOLOGY & METABOLISM BMJ Open Diabetes Research & Care Pub Date : 2024-03-01 DOI:10.1136/bmjdrc-2023-003784
Sunghee Choi, Mithun Kumer Sarker, Mi Ra Yu, Haekyung Lee, Soon Hyo Kwon, Jin Seok Jeon, Hyunjin Noh, Hyoungnae Kim
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引用次数: 0

摘要

引言 我们以前曾报道过,在糖尿病肾病(DKD)患者体内,有八种循环外泌体微RNA(miRNA)明显上调。然而,它们在肾脏中的具体作用和分子机制仍然未知。在这八个 miRNAs 中,我们评估了 miR-5010-5p 在糖尿病条件下对肾小管上皮细胞的影响。研究设计与方法 我们利用重组质粒将 miR-5010-5p 模拟物转染肾小管上皮细胞系 HK-2。使用双荧光素酶检测法确定了 hsa-miR-5010-5p 的靶基因。细胞活力通过 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基溴化四唑测定法进行评估。此外,还分别通过实时 PCR 和 Western 印迹法测定了 mRNA 和蛋白质的表达水平。结果 高血糖水平对 HK-2 细胞中 miR-5010-5p 的胞内表达无明显影响。转染 miR-5010-5p mimic 不会改变细胞的活力。然而,在高糖条件下,转染了 miR-5010-5p 的 HK-2 细胞的炎症细胞因子(如单核细胞趋化蛋白-1、白细胞介素-1β 和肿瘤坏死因子-ɑ)表达水平明显下降。这些变化伴随着磷酸化 AMP 激活蛋白激酶(AMPK)表达的恢复和核因子-kappa B 磷酸化的降低。双荧光素酶测定显示,miR-5010-5p 靶向蛋白磷酸酶 2 调节亚基 B delta(PPP2R2D)基因,该基因是蛋白磷酸酶 2A 的一个亚基,可调节 AMPK 磷酸化。结论 我们的研究结果表明,miR-5010-5p 表达量的增加可通过调节靶基因 PPP2R2D(它可调节 AMPK 磷酸化)来减轻高血糖诱导的肾小管上皮细胞炎症反应。因此,miR-5010-5p 可能是治疗 DKD 的一个有前景的靶点。如有合理要求,可提供相关数据。
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MicroRNA-5010-5p ameliorates high-glucose induced inflammation in renal tubular epithelial cells by modulating the expression of PPP2R2D
Introduction We previously reported the significant upregulation of eight circulating exosomal microRNAs (miRNAs) in patients with diabetic kidney disease (DKD). However, their specific roles and molecular mechanisms in the kidney remain unknown. Among the eight miRNAs, we evaluated the effects of miR-5010-5p on renal tubular epithelial cells under diabetic conditions in this study. Research design and methods We transfected the renal tubular epithelial cell line, HK-2, with an miR-5010-5p mimic using recombinant plasmids. The target gene of hsa-miR-5010-5p was identified using a dual-luciferase assay. Cell viability was assessed via the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. Moreover, mRNA and protein expression levels were determined via real-time PCR and western blotting, respectively. Results High glucose levels did not significantly affect the intracellular expression of miR-5010-5p in HK-2 cells. Transfection of the miR-5010-5p mimic caused no change in cell viability. However, miR-5010-5p-transfected HK-2 cells exhibited significantly decreased expression levels of inflammatory cytokines, such as the monocyte chemoattractant protein-1, interleukin-1β, and tumor necrosis factor-ɑ, under high-glucose conditions. These changes were accompanied by the restored expression of phosphorylated AMP-activated protein kinase (AMPK) and decreased phosphorylation of nuclear factor-kappa B. Dual-luciferase assay revealed that miR-5010-5p targeted the gene, protein phosphatase 2 regulatory subunit B delta (PPP2R2D), a subunit of protein phosphatase 2A, which modulates AMPK phosphorylation. Conclusions Our findings suggest that increased miR-5010-5p expression reduces high glucose-induced inflammatory responses in renal tubular epithelial cells via the regulation of the target gene, PPP2R2D, which modulates AMPK phosphorylation. Therefore, miR-5010-5p may be a promising therapeutic target for DKD. Data are available upon reasonable request.
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来源期刊
BMJ Open Diabetes Research & Care
BMJ Open Diabetes Research & Care Medicine-Endocrinology, Diabetes and Metabolism
CiteScore
9.30
自引率
2.40%
发文量
123
审稿时长
18 weeks
期刊介绍: BMJ Open Diabetes Research & Care is an open access journal committed to publishing high-quality, basic and clinical research articles regarding type 1 and type 2 diabetes, and associated complications. Only original content will be accepted, and submissions are subject to rigorous peer review to ensure the publication of high-quality — and evidence-based — original research articles.
期刊最新文献
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