AIM Degenerative calcific aortic valve disease (DCAVD) is a common valve disease characterized by massive calcium deposits in the aortic valve. Osteoblast differentiation of valve interstitial cells (VICs) is responsible for the formation of calcific nodules. This study aims to explore the function and underlying mechanism of long non-coding RNA (lncRNA) AFAP1-AS1 (actin filament-associated protein 1 antisense RNA 1) in the pathogenesis of DCAVD. METHODS AFAP1-AS1, miR-155 and mRNA levels were detected by qRT-PCR. Protein levels were measured by Western blot. Calcification deposition was examined by Alizarin Red staining. The interaction between AFAP1-AS1 and miR-155, as well as miR-155 and SMAD5 was evaluated using luciferase reporter assay. RESULTS AFAP1-AS1 expression was increased both in calcified aortic valves from DCAVD patients and after osteogenic induction in human VICs. Furthermore, AFAP1-AS1 overexpression promoted osteogenic differentiation of VICs, whereas AFAP1-AS1 knockdown inhibited osteogenic differentiation. Mechanistically, AFAP1-AS1 acted as a sponge for miR-155 to elevate SMAD5 expression. Further functional assays revealed that miR-155 mimic and SMAD5 silencing effectively reversed AFAP1-AS1-promoted osteogenic differentiation of VICs. CONCLUSION Collectively, AFAP1-AS1 promotes osteogenic differentiation of VICs, at least in part, by sponging miR-155 to upregulate SMAD5. This study sheds new light on lncRNA-directed therapeutics in DCAVD.

中文翻译：

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LncRNA AFAP1-AS1通过调节miR-155 / SMAD5轴促进人主动脉瓣间质细胞的成骨细胞分化。

目的变性钙化性主动脉瓣膜疾病（DCAVD）是一种常见的瓣膜疾病，其特征是主动脉瓣中大量钙沉积。瓣膜间质细胞（VIC）的成骨细胞分化是钙化结节形成的原因。本研究旨在探讨长非编码RNA（lncRNA）AFAP1-AS1（肌动蛋白丝相关蛋白1反义RNA 1）在DCAVD发病机理中的功能和潜在机制。方法采用qRT-PCR检测AFAP1-AS1，miR-155和mRNA水平。通过蛋白质印迹法测量蛋白质水平。通过茜素红染色检查钙化沉积。使用荧光素酶报告基因分析评估了AFAP1-AS1和miR-155以及miR-155和SMAD5之间的相互作用。结果DCAPD患者的钙化主动脉瓣以及人VIC的成骨诱导后，AFAP1-AS1表达均增加。此外，AFAP1-AS1的过表达促进了VIC的成骨分化，而AFAP1-AS1的抑制则抑制了成骨分化。从机理上讲，AFAP1-AS1充当miR-155的海绵，从而提高了SMAD5的表达。进一步的功能分析表明，miR-155模拟和SMAD5沉默有效逆转了AFAP1-AS1促进的VIC成骨分化。结论AFAP1-AS1总的来说通过使miR-155变海绵状上调SMAD5，至少部分地促进了VIC的成骨分化。这项研究为DCAVD中lncRNA指导的治疗方法提供了新的思路。而AFAP1-AS1组合物抑制成骨分化。从机理上讲，AFAP1-AS1充当miR-155的海绵，从而提高了SMAD5的表达。进一步的功能分析表明，miR-155模拟和SMAD5沉默有效逆转了AFAP1-AS1促进的VIC成骨分化。结论AFAP1-AS1总的来说通过使miR-155变海绵状上调SMAD5，至少部分地促进了VIC的成骨分化。这项研究为DCAVD中lncRNA指导的治疗方法提供了新的思路。而AFAP1-AS1组合物抑制成骨分化。从机理上讲，AFAP1-AS1充当miR-155的海绵，从而提高了SMAD5的表达。进一步的功能分析表明，miR-155模拟和SMAD5沉默有效逆转了AFAP1-AS1促进的VIC成骨分化。结论AFAP1-AS1总的来说通过使miR-155变海绵状上调SMAD5，至少部分地促进了VIC的成骨分化。这项研究为DCAVD中lncRNA指导的治疗方法提供了新的思路。AFAP1-AS1至少可以通过使miR-155海绵化以上调SMAD5来促进VIC的成骨分化。这项研究为DCAVD中lncRNA指导的治疗方法提供了新的思路。AFAP1-AS1至少可以通过使miR-155海绵化以上调SMAD5来促进VIC的成骨分化。这项研究为DCAVD中lncRNA指导的治疗方法提供了新的思路。