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Abstract：
Vascular calcification (VC) increases the future risk of cardiovascular events in uraemic patients, but effective therapies are still unavailable. Accurate identification of those at risk of developing VC using pathogenesis-based biomarkers is of particular interest and may facilitate individualized risk stratification. We aimed to uncover microRNA (miRNA)-target protein-based biomarker panels for evaluating uraemic VC probability and severity.
We created a three-tiered in vitro VC model and an in vivo uraemic rat model receiving high phosphate diet to mimic uraemic VC. RNAs from the three-tiered in vitro and in vivo uraemic VC models underwent miRNA and mRNA microarray, with results screened for differentially expressed miRNAs and their target genes as biomarkers. Findings were validated in original models and additionally in an ex vivo VC model and human cells, followed by functional assays of identified miRNAs and target proteins, and tests of sera from end-stage renal disease (ESRD) and non-dialysis-dependent chronic kidney disease (CKD) patients without and with VC. Totally 122 down-regulated and 119 up-regulated miRNAs during calcification progression were identified initially; further list narrowing based on miRNA-mRNA pairing, anti-correlation, and functional enrichment left 16 and 14 differentially expressed miRNAs and mRNAs. Levels of four miRNAs (miR-10b-5p, miR-195, miR-125b-2-3p, and miR-378a-3p) were shown to decrease throughout all models tested, while one mRNA (SULF1, a potential target of miR-378a-3p) exhibited the opposite trend concurrently. Among 96 ESRD (70.8% with VC) and 59 CKD patients (61% with VC), serum miR-125b2-3p and miR-378a-3p decreased with greater VC severity, while serum SULF1 levels increased. Adding serum miR-125b-2-3p, miR-378a-3p, and SULF1 into regression models for VC substantially improved performance compared to using clinical variables alone.
Using a translational approach, we discovered a novel panel of biomarkers for gauging the probability/severity of uraemic VC based on miRNAs/target proteins, which improved the diagnostic accuracy.
We created a three-tiered in vitro VC model and an in vivo uraemic rat model receiving high phosphate diet to mimic uraemic VC. RNAs from the three-tiered in vitro and in vivo uraemic VC models underwent miRNA and mRNA microarray, with results screened for differentially expressed miRNAs and their target genes as biomarkers. Findings were validated in original models and additionally in an ex vivo VC model and human cells, followed by functional assays of identified miRNAs and target proteins, and tests of sera from end-stage renal disease (ESRD) and non-dialysis-dependent chronic kidney disease (CKD) patients without and with VC. Totally 122 down-regulated and 119 up-regulated miRNAs during calcification progression were identified initially; further list narrowing based on miRNA-mRNA pairing, anti-correlation, and functional enrichment left 16 and 14 differentially expressed miRNAs and mRNAs. Levels of four miRNAs (miR-10b-5p, miR-195, miR-125b-2-3p, and miR-378a-3p) were shown to decrease throughout all models tested, while one mRNA (SULF1, a potential target of miR-378a-3p) exhibited the opposite trend concurrently. Among 96 ESRD (70.8% with VC) and 59 CKD patients (61% with VC), serum miR-125b2-3p and miR-378a-3p decreased with greater VC severity, while serum SULF1 levels increased. Adding serum miR-125b-2-3p, miR-378a-3p, and SULF1 into regression models for VC substantially improved performance compared to using clinical variables alone.
Using a translational approach, we discovered a novel panel of biomarkers for gauging the probability/severity of uraemic VC based on miRNAs/target proteins, which improved the diagnostic accuracy.
摘要：
血管钙化（VC）会增加尿毒症患者未来发生心血管事件的风险，但有效的治疗方法仍然不可用。使用基于发病机理的生物标志物准确识别有发展VC风险的人是特别感兴趣的，并且可以促进个性化风险分层。我们旨在揭示microRNA（miRNA）-基于靶蛋白的生物标志物小组，用于评估尿毒症VC的概率和严重程度。
我们创建了三层体外VC模型和接受高磷酸盐饮食以模拟尿毒症VC的体内尿毒症大鼠模型。来自三层体外和体内尿毒症VC模型的RNA进行了miRNA和mRNA微阵列，结果筛选了差异表达的miRNA及其靶基因作为生物标志物。在原始模型以及离体VC模型和人细胞中验证了研究结果，然后是鉴定的miRNA和靶蛋白的功能测定，和来自终末期肾病（ESRD）和无VC和有VC的非透析依赖性慢性肾病（CKD）患者的血清测试。在钙化进程中总共鉴定出122个下调和119个上调的miRNA;基于miRNA-mRNA配对进一步列表缩小,反相关,和功能富集剩下16和14个差异表达的miRNA和mRNA。四种miRNA的水平（miR-10b-5p，miR-195,miR-125b-2-3p,和miR-378a-3p）显示在所有测试的模型中降低，而一个mRNA（SULF1，miR-378a-3p的潜在靶标）同时表现出相反的趋势。在96例ESRD（VC占70.8％）和59例CKD患者（VC占61％）中，血清miR-125b2-3p和miR-378a-3p随着VC严重程度的增加而降低，而血清SULF1水平升高。添加血清miR-125b-2-3p，miR-378a-3p,与仅使用临床变量相比，将SULF1纳入VC的回归模型大大改善了性能。
使用平移方法，我们发现了一组新的生物标志物，用于基于miRNA/靶蛋白来衡量尿毒症VC的概率/严重程度，提高了诊断的准确性。
我们创建了三层体外VC模型和接受高磷酸盐饮食以模拟尿毒症VC的体内尿毒症大鼠模型。来自三层体外和体内尿毒症VC模型的RNA进行了miRNA和mRNA微阵列，结果筛选了差异表达的miRNA及其靶基因作为生物标志物。在原始模型以及离体VC模型和人细胞中验证了研究结果，然后是鉴定的miRNA和靶蛋白的功能测定，和来自终末期肾病（ESRD）和无VC和有VC的非透析依赖性慢性肾病（CKD）患者的血清测试。在钙化进程中总共鉴定出122个下调和119个上调的miRNA;基于miRNA-mRNA配对进一步列表缩小,反相关,和功能富集剩下16和14个差异表达的miRNA和mRNA。四种miRNA的水平（miR-10b-5p，miR-195,miR-125b-2-3p,和miR-378a-3p）显示在所有测试的模型中降低，而一个mRNA（SULF1，miR-378a-3p的潜在靶标）同时表现出相反的趋势。在96例ESRD（VC占70.8％）和59例CKD患者（VC占61％）中，血清miR-125b2-3p和miR-378a-3p随着VC严重程度的增加而降低，而血清SULF1水平升高。添加血清miR-125b-2-3p，miR-378a-3p,与仅使用临床变量相比，将SULF1纳入VC的回归模型大大改善了性能。
使用平移方法，我们发现了一组新的生物标志物，用于基于miRNA/靶蛋白来衡量尿毒症VC的概率/严重程度，提高了诊断的准确性。
