本文選題：人牙周膜細胞 + 成骨分化��； 參考：《口腔醫(yī)學研究》2017年09期

【摘要】：目的:探討微小RNA-34a在牙周膜細胞成骨向分化中的作用。方法:體外分離培養(yǎng)人牙周膜細胞(hPDLCs),取第3~6代用于實驗。首先,對hPDLCs進行成骨誘導液處理,在3、7、14d后利用實時熒光定量PCR(qRT-PCR)檢測miR-34a基因的表達變化。然后,構(gòu)建慢病毒載體pCDH-pre-miR-34a和pCDH,將慢病毒載體感染hPDLCs,構(gòu)建pre-miR-34a過表達細胞模型(hPDLCs/34a)和空載體對照細胞模型(hPDLCs/pCDH),并誘導hPDLCs成骨分化3、7、14和21d。分析成骨標志基因堿性磷酸酶(ALP)、Runt相關轉(zhuǎn)錄因子2(Runx2)、骨鈣素(OCN)及骨涎蛋白(BSP)表達變化,ALP活性及鈣化結(jié)節(jié)形成的茜素紅染色情況。結(jié)果:hPDLCs經(jīng)成骨分化誘導后,miR-34a基因表達在第3天開始明顯升高,并呈逐漸增高趨勢,差異均有統(tǒng)計學意義(P0.001)。與hPDLCs/pCDH組相比,hPDLCs/34a組的ALP活性和茜素紅染色均有明顯減弱。qRT-PCR結(jié)果顯示:hPDLCs/34a組的ALP表達量在成骨誘導7d時較hPDLCs/pCDH組降低(P0.05);Runx2、OCN及BSP表達量的差異在各時間點基本無統(tǒng)計學意義。結(jié)論:在體外條件下,miR-34a抑制人牙周膜細胞成骨向分化。
[Abstract]:Objective: to investigate the role of small RNA-34a in osteogenic differentiation of periodontal ligament cells. Methods: human periodontal ligament cells were isolated and cultured in vitro. Firstly, hPDLCs was treated with osteoblast inducer and the expression of miR-34a gene was detected by real-time fluorescence quantitative PCR qRT-PCRat 14 days later. Then, lentivirus vectors pCDH-pre-miR-34a and pCDH were constructed, lentivirus vectors were infected with hPDLCs, and pre-miR-34a overexpression cell models were constructed. The changes of ALP activity and alizarin red staining in calcified nodules were analyzed by analyzing the expression of osteoblastic marker gene ALPT-Runt related transcription factor 2runx2, osteocalcin OCNand bone sialoprotein (BSPP). Results the expression of miR-34a gene began to increase significantly on the 3rd day after osteogenic differentiation induced by% hPDLCs, and the expression of miR-34a gene increased gradually. The difference was statistically significant (P 0.001). Compared with hPDLCs/pCDH group, ALP activity and alizarin red staining in hPDLC / 34a group were significantly decreased. The results of qRT-PCR showed that the expression of ALP in 10 hPDLC / 34a group was significantly lower than that in hPDLCs/pCDH group on the 7th day after osteogenesis. There was no significant difference in the expression of ALP and BSP between the two groups at all time points. Conclusion: in vitro, miR-34a inhibits osteogenic differentiation of human periodontal ligament cells.
【作者單位】： 武漢大學口腔醫(yī)學院口腔基礎醫(yī)學省部共建國家重點實驗室培訓基地和口腔生物醫(yī)學教育部重點實驗室;武漢大學口腔醫(yī)院修復科;
【基金】：國家自然科學基金(編號:81200812) 武漢市應用基礎研究計劃項目(編號:2016060101010043)
【分類號】：R781.4

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