Department of Cardiovasology, the First Hospital of Hebei Medical University, No. 89 Donggang Road, Yuhua District, Shijiazhuang, Hebei, 050031, China.
Department of Cardiovasology, Han Dan First Hospital, No. 24, Congtai Road, Congtai District, Handan, Hebei, 056001, China.
Tissue Eng Regen Med. 2020 Oct;17(5):659-670. doi: 10.1007/s13770-020-00270-8. Epub 2020 Aug 6.
The cardiomyocyte apoptosis is considered as one of major contributions to cardiac remodeling after myocardial infarction (MI). Numerous studies find that circular RNAs (circRNAs) play pivotal roles in a variety of biological functions. However, the role of circ_0068655 in MI and human induced pluripotent stem-derived cardiomyocytes (HCMs) remains unknown.
The expression of circ_0068655, miR-498, and PRKC apoptosis WT1 regulator (PAWR) in human MI heart tissues and hypoxia subjected HCMs was evaluated with qRT-PCR and Western blot. The effects of circ_0068655 on hypoxia-induced apoptotic death and cell migration in HCMs were evaluated with qRT-PCR, cell viability, cell death ELISA (POD), and Caspase-3 activity assay, and Trans-well assay, respectively. Furthermore, luciferase assay, qRT-PCR, biotin-labeled miRNA pulldown assay, and Western blot were employed in the functional studies.
We found that the expression of circ_0068655 and PAWR was enhanced in MI patients and hypoxia subjected HCMs; by contrast, the expression of miR-498 decreased. Inhibited expression of circ_0068655 in HMCs counteracted hypoxia-induced apoptotic death and impaired cell migration, in sharp contrast to circ_0068655 knockdown. We identified that circ_0068655 sponged an endogenous miR-498 to sequester and inhibit its activity, leading to the increased PAWR expression.
Our findings reveal that the expression of circ_0068655 can promote cardiomyocyte apoptosis through the modulation of miR-498-PAWR axis in vitro, which highlights the diagnostic and therapeutic value of circ_0068655 in patients with MI.
心肌细胞凋亡被认为是心肌梗死后心脏重构的主要原因之一。大量研究发现，环状 RNA（circRNAs）在多种生物学功能中发挥着关键作用。然而，circ_0068655 在心肌梗死和人诱导多能干细胞衍生的心肌细胞（HCMs）中的作用尚不清楚。
采用 qRT-PCR 和 Western blot 检测人心肌梗死心脏组织和缺氧处理的 HCMs 中 circ_0068655、miR-498 和 PRKC 凋亡 WT1 调节剂（PAWR）的表达。采用 qRT-PCR、细胞活力测定、细胞死亡 ELISA（POD）和 Caspase-3 活性测定以及 Trans-well 测定分别评估 circ_0068655 对 HCMs 缺氧诱导的凋亡死亡和细胞迁移的影响。此外，还进行了荧光素酶报告基因检测、qRT-PCR、生物素标记 miRNA 下拉测定和 Western blot 等功能研究。
我们发现，circ_0068655 和 PAWR 的表达在心肌梗死患者和缺氧处理的 HCMs 中增加，而 miR-498 的表达减少。HCMs 中 circ_0068655 的表达抑制逆转了缺氧诱导的凋亡死亡和细胞迁移受损，与 circ_0068655 敲低形成鲜明对比。我们发现，circ_0068655 作为内源性 miR-498 的海绵，通过与 miR-498 结合并抑制其活性，导致 PAWR 表达增加。
我们的研究结果表明，circ_0068655 在体外通过调节 miR-498-PAWR 轴促进心肌细胞凋亡，这凸显了 circ_0068655 在心肌梗死患者中的诊断和治疗价值。
