College of Pharmacy, Xinxiang Medical University, No. 601 Jinsui Avenue, Hongqi District, Xinxiang, 453003, Henan, China.
Collaborative Innovation Center of Molecular Diagnosis and Laboratory Medicine in Henan Province, School of Laboratory Medicine, Xinxiang Medical University, Xinxiang, 453003, Henan, China.
Pflugers Arch. 2019 Feb;471(2):347-355. doi: 10.1007/s00424-018-2219-8. Epub 2018 Oct 23.
Our study explored the effects of lncRNA UCA1 on the proliferation and apoptosis in hypoxic human pulmonary artery smooth muscle cells (HPASMCs) and highlighted the endogenous relationship between UCA1, ING5, and hnRNP I in cell proliferation. Hypoxia-induced HPASMCs were used to simulate pulmonary arterial hypertension in vitro. Microarray assay was adopted to screen the dysregulated expressed lncRNAs in HPASMCs to find out the target gene of our study. And RT-qPCR was performed to detect the expression of lncRNA UCA1 under hypoxia and normoxia. After transfection, the relationship between UCA1 and cell proliferation in HPASMCs under hypoxia were determined by cell proliferation assay and relative expression of PCNA. Next, ELISA assays were conducted to measure the protein levels of PCNA and ING5. What's more, flow cytometry was employed to measure the apoptosis rate in differentially UCA1-expressed HPASMCs. RIP assays were conducted to further clarify the endogenous relationship between UCA1 and ING5 in hypoxic HPASMCs. Finally, the effects of ING5 to HPASMCs were detected after transfection of ING5 and UCA1 to figure out the role of ING5 in HPASMCs. Hypoxia was revealed to induce proliferation and inhibited apoptosis in HPASMCs. Besides, UCA1 was confirmed to be highly expressed under hypoxia compared with normoxia. UCA1 boosted cell proliferation under hypoxia in HPASMCs. However, the apoptosis was suppressed in the hypoxic HPASMCs transfected with pcDNA3.1-UCA1. Further, mechanism studies found that UCA1 competed with ING5 for hnRNP I, so that upregulating UCA1 inhibited the protein levels of ING5. And finally we found that ING5 restrained cell viability, but promoted cell apoptosis in hypoxic HPASMCs, which was reversed by UCA1 over-expression. In summary, our findings manifested that UCA1 promoted proliferation and restrained apoptosis by competing with ING5 for hnRNP I in HPASMCs induced by hypoxia, indicating their potential roles for the cure of hypoxic pulmonary hypertension.
我们的研究探讨了长链非编码 RNA UCA1 对低氧人肺动脉平滑肌细胞(HPASMCs)增殖和凋亡的影响,并强调了 UCA1、ING5 和 hnRNP I 在细胞增殖中的内源性关系。缺氧诱导的 HPASMCs 用于体外模拟肺动脉高血压。采用微阵列分析筛选 HPASMCs 中失调表达的 lncRNAs,以找到我们研究的靶基因。然后通过 RT-qPCR 检测低氧和常氧下 lncRNA UCA1 的表达。转染后,通过细胞增殖试验和 PCNA 的相对表达来确定 UCA1 与低氧条件下 HPASMCs 增殖的关系。接下来,通过 ELISA 测定 PCNA 和 ING5 的蛋白水平。此外,通过流式细胞术测定不同 UCA1 表达的 HPASMCs 的凋亡率。RIP 实验进一步阐明了低氧 HPASMCs 中 UCA1 和 ING5 之间的内源性关系。最后,转染 ING5 和 UCA1 后检测 ING5 对 HPASMCs 的影响,以确定 ING5 在 HPASMCs 中的作用。低氧诱导 HPASMCs 增殖并抑制凋亡。此外,与常氧相比,UCA1 在低氧下的表达明显升高。UCA1 促进低氧条件下 HPASMCs 的细胞增殖。然而,pcDNA3.1-UCA1 转染的低氧 HPASMCs 中凋亡受到抑制。进一步的机制研究发现,UCA1 与 ING5 竞争 hnRNP I,从而上调 UCA1 抑制 ING5 的蛋白水平。最后,我们发现 ING5 在低氧 HPASMCs 中抑制细胞活力,但促进细胞凋亡,而 UCA1 过表达则逆转了这一现象。综上所述,我们的研究结果表明,UCA1 通过与 ING5 竞争 hnRNP I,在低氧诱导的 HPASMCs 中促进增殖并抑制凋亡,表明它们在低氧性肺动脉高压的治疗中具有潜在作用。