[创伤对大鼠C6胶质瘤肿瘤干细胞体内致瘤性的影响]

Liu Wen-ke, Ma Lu, Liu Yan-hui, Wang Xiu-jie, Jiang Shu

Department of Neurosurgery, West China Hospital, Sichuan University, Chengdu 610041, China.

Sichuan Da Xue Xue Bao Yi Xue Ban. 2011 Mar;42(2):161-5, 169.

OBJECTIVE

To investigate the influence of trauma on the tumorigenicity of rat glial tumor stem cells (C6-side population cell, C6-SP) in vivo.

METHODS

Rat glial tumor stem cells C6-SP were isolated by flowcytometry. The biological behavior of C6-SP cells were observed by means of MTT experiment, single cell cloning, and cell cycle study with FCM CD133 expression was measured by immunofluorescence and FCM.. The tumorigenicity of C6-SP cells in vivo was evaluated by in situ tumor growth after intracranial implantation. The rat model was established by intracranial implantation of C6-SP cells. 10 days later, the rats in experimental groups were subjected to orthotopic or ectopic trauma. 24 days later, brain specimen was retrieved, gross tumor volume was measured, and Ki67 was evaluated by immunochemistry. The migration of stem cell was studied by the method to observe the relocation of C6-SP cells.

RESULTS

Clustrus tumor growth was seen when C6-SP cell was cultured in serum-free medium. The doubling time of C6-SP cell was shorter than ordinary C6 cell. Single stem cell cloning efficiency of C6-SP cell was 77% whereas that of ordinary C6 cell was 16.4%. Among cloned C6-SP cell and ordinary C6 cell, 49.7% +/- 5.2% and 35.2 +/- 4.3% were at G0/G1 phase respectively. CD133 expression of C6-SP cells was positively shown by immunofluorescence. Tumorigenesis was 100% 2 weeks after in situ implantation of C6-SP cells. Gross tumor volume and Ki67 expression of orthotopic trauma group were larger and higher than those of ectopic trauma group or blank control group (P < 0.05) whereas difference was insignificant in the later two groups (P > 0.05). Red stained cells relocation was seen in both traumatized groups and absent in controls.

CONCLUSION

C6-SP cells are the tumor stem cells (TSCs) for rat glioma. Trauma at the lesional site could increase tumorigenicity of the C6-SP cells. Moreover, trauma could induce migration of C6-SP cells in brain.

目的

探讨创伤对大鼠胶质瘤干细胞（C6侧群细胞，C6-SP）体内致瘤性的影响。

方法

采用流式细胞术分离大鼠胶质瘤干细胞C6-SP。通过MTT实验、单细胞克隆及FCM进行细胞周期研究观察C6-SP细胞的生物学行为，采用免疫荧光和FCM检测CD133表达。通过颅内植入后原位肿瘤生长评估C6-SP细胞在体内的致瘤性。建立C6-SP细胞颅内植入大鼠模型。10天后，对实验组大鼠进行原位或异位创伤。24天后，取出脑标本，测量肿瘤总体积，采用免疫化学法评估Ki67。通过观察C6-SP细胞重定位的方法研究干细胞的迁移。

结果

C6-SP细胞在无血清培养基中培养时可见成簇肿瘤生长。C6-SP细胞的倍增时间短于普通C6细胞。C6-SP细胞的单干细胞克隆效率为77%，而普通C6细胞为16.4%。在克隆的C6-SP细胞和普通C6细胞中，分别有49.7%±5.2%和35.2±4.3%处于G0/G1期。免疫荧光显示C6-SP细胞CD133表达呈阳性。C6-SP细胞原位植入2周后致瘤率为100%。原位创伤组的肿瘤总体积和Ki67表达高于异位创伤组或空白对照组（P<0.05），而后两组差异不显著（P>0.05）。创伤组均可见红色染色细胞重定位，对照组未见。