Altered microRNAs in bicuspid aortic valve: a comparison between stenotic and insufficient valves.
Altered microRNAs in bicuspid aortic valve: a comparison between stenotic and insufficient valves.
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发表时间：
2010-07
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通讯作者：
Vishal Nigam;H. Sievers;B. Jensen;H. Sier;P. Simpson;D. Srivastava;S. A. Mohamed
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作者：
Vishal Nigam;H. Sievers;B. Jensen;H. Sier;P. Simpson;D. Srivastava;S. A. Mohamed
二叶式主动脉瓣（BAV）是最常见的先天性心脏病，是主动脉瓣狭窄（AS）和主动脉瓣关闭不全（AI）的主要原因。AS通常由钙化性瓣膜疾病引起。最近，microRNAs（miRNAs）已被证明可以调节基因表达。该研究的目的是检查与AI患者相比，AS患者主动脉瓣小叶中发生改变的miRNA。还进行了体外实验以确定这些miRNAs是否可以调节钙化相关基因。方法：从9例接受主动脉瓣置换术（AVR）的男性患者（平均年龄44.9 ± 13.8岁）中采集主动脉瓣样本（融合和未融合瓣叶）。使用含有1，421个miRNA的PIQOR miRXplore微阵列，并与用Cy 5标记的融合小叶样品杂交;未融合的样品用作对照，并用Cy 3标记。进行定量逆转录-聚合酶链反应（qRT-PCR）以验证miRNA阵列结果。用miRNA模拟物处理培养的人主动脉瓣间质细胞（AVIC），并进行qRT-PCR以确定mRNA的任何变化。结果基因芯片分析显示，有7种miRNAs在AS和AI患者中存在显著差异。在狭窄样品中，显示MiR-26 a和miR-195水平（通过qRT-PCR）分别降低65%和59%（p < 0.05），并且MiR-30 b降低62%（p < 0.06）。用miR-26 a或miR-30 b模拟物处理的人AVIC显示钙化相关基因的mRNA水平降低。MiR-26 a抑制BMP 2 36%，碱性磷酸酶（ALPL）38%，SMAD 1 26%，而MiR-30 b降低SMAD 1 18%和SMAD 3 12%的表达。相比之下，miR-195处理的AVIC增加了钙化相关基因的mRNA水平，如BMP 2增加了68%，RUNX 2增加了11%。结论：与因AI而需要置换的患者相比，因AS而需要AVR的患者主动脉瓣中的miR-26 a、miR-30 b和miR-195均降低。这些miRNAs似乎在体外调节钙化相关基因。
BACKGROUND AND AIM OF THE STUDY Bicuspid aortic valve (BAV), the most common form of congenital heart disease, is a leading cause of aortic stenosis (AS) and aortic insufficiency (AI). AS is typically caused by calcific valve disease. Recently, microRNAs (miRNAs) have been shown to modulate gene expression. The study aim was to examine the miRNAs that were altered in the aortic valve leaflets of patients with AS compared to those in patients with AI. In-vitro experiments were also carried out to determine if these miRNAs could modulate calcification-related genes. METHODS Aortic valve samples (fused and unfused leaflets) were collected from nine male patients (mean age 44.9 +/- 13.8 years) undergoing aortic valve replacement (AVR). PIQOR miRXplore Microarrays containing 1,421 miRNAs were used and hybridized to fused leaflet samples labeled with Cy5; unfused samples were used as controls and labeled with Cy3. A quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to validate the miRNA array results. Cultured human aortic valve interstitial cells (AVICs) were treated with miRNA mimics, and qRT-PCR was carried out to determine any changes in mRNAs. RESULTS By microarray analysis, seven miRNAs were shown to be statistically different between the AS and AI patients. In the stenotic samples, the MiR-26a and miR-195 levels were shown (by qRT-PCR) to be reduced by 65% and 59%, respectively (p < 0.05), and MiR-30b to be reduced by 62% (p < 0.06). Human AVICs treated with miR-26a or miR-30b mimics showed decreased mRNA levels of calcification-related genes. MiR-26a repressed BMP2 by 36%, alkaline phosphatase (ALPL) by 38%, and SMAD1 by 26%, while MiR-30b reduced the expression of SMAD1 by 18% and of SMAD3 by 12%. In contrast, miR-195-treated AVICs had increased mRNA levels of calcification-related genes, such as BMP2 by 68% and RUNX2 by 11%. CONCLUSION MiR-26a, miR-30b, and miR-195 were each decreased in the aortic valves of patients requiring AVR due to AS, compared to those requiring replacement due to AI. These miRNAs appear to modulate calcification-related genes in vitro.