Abstract：
Polyunsaturated fatty acids (PUFAs) can decrease the risk of calcium oxalate stone formation, which accounts for 80% of all renal stones. This study aimed to investigate the protective mechanisms of PUFAs against renal stones.
Urine samples of patients with renal stones and biopsy tissue samples from patients with nephrocalcinosis were tested for miR-93-5p expression. A renal stone mouse model was established with intraperitoneal injection of glyoxylic acid, during which mice were treated with PUFAs and/or an miR-93-5p inhibitor adenovirus. Periodic acid-Schiff staining, terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling staining, oil red O staining, triacylglycerol assay, and colorimetry testing were performed to assess glycogen deposition, apoptosis, lipid accumulation, blood urea nitrogen, and serum creatinine levels, respectively. Renal proximal tubular epithelial cells (human kidney 2 [HK-2]) were subjected to gain- and loss-of-function assays before calcium-oxalate monohydrate (COM) induction and PUFA treatment. Cell counting kit 8, flow cytometry, and lactate dehydrogenase activity assays were used to examine cell viability, apoptosis, and damage. A luciferase reporter gene assay verified the interaction between miR-93-5p and Pknox1, and miR-93-5p and Pknox1 levels were assessed using a reverse transcription-quantitative polymerase chain reaction and Western blot analysis.
miR-93-5p was downregulated in clinical samples with renal stones and negatively targeted Pknox1. PUFAs increased miR-93-5p expression and reduced apoptosis, glycogen deposition, and lipid accumulation in mice with renal stones, which were annulled by miR-93-5p downregulation. PUFAs increased proliferation and diminished apoptosis, lipid accumulation, and lactate dehydrogenase activity in COM-induced HK-2 cells, which were negated by miR-93-5p inhibition. Pknox1 overexpression reversed the effect of miR-93-5p upregulation on COM-induced HK-2 cells.
PUFAs repressed renal stone-induced renal tubular damage via the miR-93-5p/Pknox1 axis.
Urine samples of patients with renal stones and biopsy tissue samples from patients with nephrocalcinosis were tested for miR-93-5p expression. A renal stone mouse model was established with intraperitoneal injection of glyoxylic acid, during which mice were treated with PUFAs and/or an miR-93-5p inhibitor adenovirus. Periodic acid-Schiff staining, terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling staining, oil red O staining, triacylglycerol assay, and colorimetry testing were performed to assess glycogen deposition, apoptosis, lipid accumulation, blood urea nitrogen, and serum creatinine levels, respectively. Renal proximal tubular epithelial cells (human kidney 2 [HK-2]) were subjected to gain- and loss-of-function assays before calcium-oxalate monohydrate (COM) induction and PUFA treatment. Cell counting kit 8, flow cytometry, and lactate dehydrogenase activity assays were used to examine cell viability, apoptosis, and damage. A luciferase reporter gene assay verified the interaction between miR-93-5p and Pknox1, and miR-93-5p and Pknox1 levels were assessed using a reverse transcription-quantitative polymerase chain reaction and Western blot analysis.
miR-93-5p was downregulated in clinical samples with renal stones and negatively targeted Pknox1. PUFAs increased miR-93-5p expression and reduced apoptosis, glycogen deposition, and lipid accumulation in mice with renal stones, which were annulled by miR-93-5p downregulation. PUFAs increased proliferation and diminished apoptosis, lipid accumulation, and lactate dehydrogenase activity in COM-induced HK-2 cells, which were negated by miR-93-5p inhibition. Pknox1 overexpression reversed the effect of miR-93-5p upregulation on COM-induced HK-2 cells.
PUFAs repressed renal stone-induced renal tubular damage via the miR-93-5p/Pknox1 axis.
摘要：
目的：多不饱和脂肪酸（PUFA）可以降低草酸钙结石形成的风险，占所有肾结石的80%。本研究旨在探讨PUFAs对肾结石的保护机制。
方法：检测肾结石患者的尿液样本和肾钙质沉着患者的活检组织样本的miR-93-5p表达。腹腔注射乙醛酸建立肾结石小鼠模型，在此期间用PUFAs和/或miR-93-5p抑制剂腺病毒治疗小鼠。周期性酸性希夫染色,末端脱氧核苷酸转移酶脱氧尿苷三磷酸缺口末端标记染色，油红O染色，三酰甘油测定，进行比色法测试以评估糖原沉积，凋亡，脂质积累，血尿素氮,和血清肌酐水平，分别。在草酸钙一水合物（COM）诱导和PUFA处理之前，对肾近端肾小管上皮细胞（人肾2[HK-2]）进行功能增益和功能丧失测定。细胞计数试剂盒8，流式细胞术，和乳酸脱氢酶活性测定用于检查细胞活力，凋亡，和损坏。荧光素酶报告基因测定验证了miR-93-5p和Pknoxl之间的相互作用，并且使用逆转录定量聚合酶链反应和Western印迹分析评估了miR-93-5p和Pknoxl水平。
结果：miR-93-5p在肾结石和阴性靶向Pknox1的临床样本中下调。PUFAs增加miR-93-5p表达并减少细胞凋亡，糖原沉积,和肾结石小鼠的脂质积累，通过miR-93-5p下调而无效。PUFA增加增殖和减少凋亡，脂质积累，和COM诱导的HK-2细胞中的乳酸脱氢酶活性，通过miR-93-5p抑制而否定。Pknox1过表达逆转了miR-93-5p上调对COM诱导的HK-2细胞的影响。
结论：PUFA通过miR-93-5p/Pknox1轴抑制肾结石诱导的肾小管损伤。
方法：检测肾结石患者的尿液样本和肾钙质沉着患者的活检组织样本的miR-93-5p表达。腹腔注射乙醛酸建立肾结石小鼠模型，在此期间用PUFAs和/或miR-93-5p抑制剂腺病毒治疗小鼠。周期性酸性希夫染色,末端脱氧核苷酸转移酶脱氧尿苷三磷酸缺口末端标记染色，油红O染色，三酰甘油测定，进行比色法测试以评估糖原沉积，凋亡，脂质积累，血尿素氮,和血清肌酐水平，分别。在草酸钙一水合物（COM）诱导和PUFA处理之前，对肾近端肾小管上皮细胞（人肾2[HK-2]）进行功能增益和功能丧失测定。细胞计数试剂盒8，流式细胞术，和乳酸脱氢酶活性测定用于检查细胞活力，凋亡，和损坏。荧光素酶报告基因测定验证了miR-93-5p和Pknoxl之间的相互作用，并且使用逆转录定量聚合酶链反应和Western印迹分析评估了miR-93-5p和Pknoxl水平。
结果：miR-93-5p在肾结石和阴性靶向Pknox1的临床样本中下调。PUFAs增加miR-93-5p表达并减少细胞凋亡，糖原沉积,和肾结石小鼠的脂质积累，通过miR-93-5p下调而无效。PUFA增加增殖和减少凋亡，脂质积累，和COM诱导的HK-2细胞中的乳酸脱氢酶活性，通过miR-93-5p抑制而否定。Pknox1过表达逆转了miR-93-5p上调对COM诱导的HK-2细胞的影响。
结论：PUFA通过miR-93-5p/Pknox1轴抑制肾结石诱导的肾小管损伤。
