二叶式主动脉瓣中微小RNA的改变：狭窄瓣膜与反流瓣膜的比较
Altered microRNAs in bicuspid aortic valve: a comparison between stenotic and insufficient valves.
作者信息
Nigam Vishal, Sievers Hans H, Jensen Brian C, Sier Holger A, Simpson Paul C, Srivastava Deepak, Mohamed Salah A
机构信息
Department of Pediatrics (Cardiology), University of California, San Francisco, USA.
出版信息
J Heart Valve Dis. 2010 Jul;19(4):459-65.
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.
研究背景与目的
二叶式主动脉瓣（BAV）是先天性心脏病最常见的形式，是主动脉狭窄（AS）和主动脉瓣关闭不全（AI）的主要病因。AS通常由钙化性瓣膜病引起。最近，微小RNA（miRNA）已被证明可调节基因表达。本研究的目的是检测与AI患者相比，AS患者主动脉瓣叶中发生改变的miRNA。还进行了体外实验以确定这些miRNA是否能调节钙化相关基因。
方法
从9名接受主动脉瓣置换术（AVR）的男性患者（平均年龄44.9±13.8岁）中收集主动脉瓣样本（融合和未融合的瓣叶）。使用包含1421种miRNA的PIQOR miRXplore微阵列，并与用Cy5标记的融合瓣叶样本杂交；未融合的样本用作对照，并用Cy3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
通过微阵列分析，显示7种miRNA在AS和AI患者之间存在统计学差异。在狭窄样本中，（通过qRT-PCR）显示MiR-26a和miR-195水平分别降低了65%和59%（p<0.05），MiR-30b降低了62%（p<0.06）。用miR-26a或miR-30b模拟物处理的人AVIC显示钙化相关基因的mRNA水平降低。MiR-26a使骨形态发生蛋白2（BMP2）降低36%，碱性磷酸酶（ALPL）降低38%，SMAD1降低26%，而MiR-30b使SMAD1的表达降低18%，SMAD3的表达降低12%。相反，用miR-195处理的AVIC的钙化相关基因的mRNA水平升高，如BMP2升高68%，RUNX2升高11%。
结论
与因AI需要置换的患者相比，因AS需要AVR的患者的主动脉瓣中MiR-26a、miR-30b和miR-195均降低。这些miRNA在体外似乎能调节钙化相关基因。
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