發(fā)布時間：2018-04-15 23:35

  本文選題：MiR- + 骨肉瘤　； 參考：《中華腫瘤防治雜志》2017年16期

【摘要】：目的骨肉瘤具有局部高度侵襲能力以及快速轉(zhuǎn)移潛能,因而致死率較高。研究指出,miR-300的表達異常與多種腫瘤的發(fā)病相關(guān)。miR-300對骨肉瘤細胞是否存在調(diào)控作用卻屬未知。本研究探討miR-300對骨肉瘤細胞Saos-2增殖與凋亡調(diào)控的作用。方法將miR-300轉(zhuǎn)染進骨肉瘤細胞Saos-2中,實現(xiàn)miR-300在Saos-2細胞內(nèi)的高表達。通過熒光定量PCR測定miR-300在細胞內(nèi)的表達水平;分別采用MTT活細胞測試,細胞周期實驗和克隆形成實驗檢測骨髓瘤細胞的增殖情況;通過蛋白質(zhì)印跡法測定Bcl-2、Bax和Bak,裂解型Caspase-3蛋白水平來鑒定細胞凋亡。結(jié)果轉(zhuǎn)染miR-300的Saos-2細胞,其miR-300表達量為對照組的(10.23±1.04)倍,t=3.613 8,P=0.000 6。MTT實驗結(jié)果顯示,在轉(zhuǎn)染后24和48h,miR-300組的Saos-2相對活細胞百分比相對對照組分別為[(174.35±28.46)%,t=2.219,P=0.03]和[(225.73±24.62)%,t=2.738,P=0.008]。miR-300組Saos-2細胞處在G1期的百分比為39.42%,低于對照組的47.58%,t=2.366,P=0.021;而處在G2期的細胞百分比為19.12%,顯著高于對照組的10.55%,t=2.987,P=0.004。miR-300組Saos-2細胞克隆形成率為(52.53±30.21)%,顯著高于對照組的(24.67±2.05)%,t=2.711,P=0.008 6。miR-300組Saos-2細胞中Bax、Bak和裂解型Caspase-3表達水平分別為對照組的(0.25±0.02)倍(t=2.785,P=0.007)、(0.31±0.03)倍(t=3.223,P=0.002)和(0.36±0.03)倍(t=3.006,P=0.003 8),差異有統(tǒng)計學(xué)意義。Bcl-2蛋白水平為對照組的(6.32±0.57)倍,t=3.218,P=0.002。轉(zhuǎn)染miRZip lent-shMiR-300的Saos-2細胞,其miR-300數(shù)量為對照組的(0.28±0.03)倍,t=3.531,P=0.000 78。轉(zhuǎn)染24h后,miR-300組Saos-2相對活細胞數(shù)為對照組的(64.46±5.32)%,t=2.324,P=0.024;48h后為對照組的(48.14±4.38)%,t=3.009,P=0.004。miR-300組Saos-2細胞處在G1期的百分比為55.71%,高于對照組的46.78%,t=2.108,P=0.039;處在G2期的細胞百分比為2.81%,低于對照組的11.46%,t=3.224,P=0.002。miR-300組的Saos-2細胞的克隆形成率為(17.63±3.89)%,顯著低于對照組的(26.84±2.94)%,t=2.989,P=0.004。miR-300組Saos-2細胞中Bax表達水平為對照組的(6.25±4.23)倍,t=3.138,P=0.002 6;Bak表達水平為對照組的(6.03±4.27)倍,t=3.395,P=0.001 2;裂解型Capase-3表達水平為對照組的(5.35±0.37)倍,t=3.017,P=0.003 7;而Bcl-2蛋白水平為對照組的(0.36±0.02)倍,t=2.769,P=0.007 4。結(jié)論 miR-300在骨肉瘤細胞的增殖與凋亡調(diào)控中具有重要作用,抑制miR-300表達可以一定程度上減少骨髓瘤細胞的增殖,促進其凋亡,為骨肉瘤的病理機制研究提供了新研究方向。
[Abstract]:Objective Osteosarcoma has high local invasion and rapid metastasis potential, so the fatality rate is high.It is suggested that the abnormal expression of miR-300 is related to the pathogenesis of many kinds of tumors. Whether miR-300 can regulate osteosarcoma cells is unknown.To investigate the effect of miR-300 on proliferation and apoptosis of osteosarcoma cell line Saos-2.Methods miR-300 was transfected into Saos-2 of osteosarcoma cells, and the high expression of miR-300 in Saos-2 cells was realized.The expression of miR-300 was measured by fluorescence quantitative PCR, and the proliferation of myeloma cells was detected by MTT living cell test, cell cycle assay and clone formation assay.Apoptotic cells were identified by Western blotting assay of Bcl-2P Bax and Bak-Bax and cleavage Caspase-3 protein levels.Results the expression of miR-300 in Saos-2 cells transfected with miR-300 was 10.23 鹵1.04 times as high as that in the control group. The results of the experiment showed that the expression of miR-300 was 3.6138Pu 0.000 6.MTT.鐨，

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