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目的探讨自噬在塞来昔布对脑胶质瘤SHG-44细胞放射增敏中的作用。方法选择脑胶质瘤细胞株SHG-44作为实验对象,并分成对照组(C组)、塞来昔布组(D组)、辐射组(R组)和联合组(D+R组)。集落形成法检测塞来昔布对SHG-44细胞的放射增敏作用。吖啶橙染色、FITC标记LC3-Ⅱ抗体和电镜共同检测细胞的自噬水平。流式细胞仪检测细胞周期分布和凋亡情况。结果塞来昔布(30μmol/L)增加SHG-44细胞的放射敏感性,诱导肿瘤细胞自噬和G2/M期阻滞。电镜观察发现塞来昔布、辐射及联合作用后胞质内可见自噬体,而无明显的核浓缩和核碎片。吖啶橙染色和FITC-MAP1-LC3-Ⅱ标记自噬体发现D+R组细胞自噬水平明显高于R组(P<0.05)。D+R组G2/M期细胞比例为(34.26±2.20)%,R组为(29.15±1.99)%,D+R组明显多于R组(P<0.05),而相应的凋亡比例分别为(9.7±1.24)%和(8.2±0.93)%,两组差异无统计学意义(P>0.05)。随着辐射剂量的增加,细胞自噬水平增高,而细胞存活率呈指数性递减(决定指数R=0.98,P<0.05)。结论塞来昔布增强辐射诱导SHG-44细胞G2/M期阻滞、促进细胞自噬、诱导细胞自噬性死亡,可能是其增加放射敏感性的机制之一。
Objective To investigate the role of autophagy in celecoxib radiosensitization of SHG-44 glioma cells. Methods The glioma cell line SHG-44 was selected as experimental group and divided into control group (C group), celecoxib group (D group), radiation group (R group) and combined group (D + R group). The colony formation assay was used to detect the radiosensitization effect of celecoxib on SHG-44 cells. Acridine orange staining, FITC labeled LC3-Ⅱ antibody and electron microscopy were used to detect the level of autophagy. Flow cytometry was used to detect cell cycle distribution and apoptosis. Results Celecoxib (30μmol / L) increased the radiosensitivity of SHG-44 cells and induced autophagy and G2 / M arrest. Electron microscopy showed that celecoxib displayed autophagosomes in the cytoplasm after irradiation and combined effects without obvious nuclear enrichment and nuclear fragmentation. Acridine orange staining and FITC-MAP1-LC3-Ⅱ labeled autophagosome showed that autophagy in D + R group was significantly higher than that in R group (P <0.05). The proportion of cells in G2 / M phase in D + R group was (34.26 ± 2.20)%, in R group (29.15 ± 1.99)%, and in D + R group was significantly higher than that in R group (P <0.05) (9.7 ± 1.24)% and (8.2 ± 0.93)%, respectively. There was no significant difference between the two groups (P> 0.05). With the increase of radiation dose, the level of autophagy increased while the cell survival rate decreased exponentially (decision index R = 0.98, P <0.05). Conclusion Celecoxib may increase G2 / M phase arrest and promote autophagy in SHG-44 cells induced by radiation, which may be one of the mechanisms by which celecoxib increases the radiosensitivity.