Key Laboratory of Oral Diseases of Jiangsu Province and Stomatological Institute of Nanjing Medical University, 140 Hanzhong Road, Nanjing, Jiangsu, 210029, China.
Endodontic Department, School of Stomatology, Nanjing Medical University, 136 Hanzhong Road, Nanjing, Jiangsu, 210029, China.
Cell Death Dis. 2019 Feb 12;10(2):130. doi: 10.1038/s41419-019-1337-3.
Long noncoding RNAs (lncRNAs) exert significant roles at transcriptional and post-transcriptional levels. Stem cells from apical papilla (SCAPs) differentiate into dentin/bone-like tissues under certain conditions. So far, whether lncRNA-H19 can affect the proliferative behaviors and osteo/odontogenesis of SCAPs, as well as its specific mechanism remain to be elucidated. Here, SCAPs were isolated and transfected with the lentiviruses or packaging vectors. Our results showed that lncRNA-H19 had no significant effect on the proliferative behaviors of SCAPs, as presented by CCK-8 assay, EdU assay and flow cytometry (FCM). Furthermore, alkaline phosphatase (ALP) activity, alizarin red staining, Western blot assay (WB), quantitative real-time polymerase chain reaction (qRT-PCR) and in vivo bone formation assay were conducted to verify the biological influences of H19 on SCAPs. Overexpression of H19 led to the enhanced osteo/odontogenesis of SCAPs, whereas knockdown of H19 inhibited these effects. Mechanistically, H19 competitively bound to miR-141 and prevented SPAG9 from miRNA-mediated degradation, thus significantly elevating phosphorylated levels of p38 and JNK and facilitating the committed differentiation of SCAPs. Taken together, the osteo/odontogenesis of SCAPs was upregulated by overexpression of H19 via miR-141/SPAG9 pathway.
长链非编码 RNA(lncRNA)在转录和转录后水平发挥重要作用。根尖乳头干细胞(SCAP)在特定条件下分化为牙本质/骨样组织。到目前为止,lncRNA-H19 是否能影响 SCAP 的增殖行为和成牙/成骨作用,以及其具体机制仍有待阐明。本研究中,分离并转染 SCAP 慢病毒或包装载体。结果显示,CCK-8 检测、EdU 检测和流式细胞术(FCM)结果表明,lncRNA-H19 对 SCAP 的增殖行为没有显著影响。此外,通过碱性磷酸酶(ALP)活性、茜素红染色、Western blot 检测(WB)、实时定量聚合酶链反应(qRT-PCR)和体内骨形成实验验证了 H19 对 SCAP 生物学影响。H19 的过表达促进了 SCAP 的成牙/成骨作用,而 H19 的敲低则抑制了这些作用。机制上,H19 竞争性结合 miR-141 并防止 SPAG9 被 miRNA 介导的降解,从而显著提高 p38 和 JNK 的磷酸化水平,并促进 SCAP 的定向分化。综上所述,通过 miR-141/SPAG9 通路的过表达上调了 SCAP 的成牙/成骨作用。
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