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目的探讨阻抑骨髓基质细胞衍生因子(SDF1)表达对与其共培养的Jurkat细胞增殖、凋亡的影响。方法脂质体介导SDF1特异性RNA干扰质粒转染培养的急性白血病骨髓基质细胞,G418筛选阳性克隆(A组),采用ELISA法检测SDF1表达变化;与Jurkat细胞共培养,绘制生长曲线并计算倍增时间,用流式细胞术检测细胞周期,末端脱氧核苷酸转移酶介导的dUTP缺失末端标记(TUNEL)法检测细胞凋亡,免疫细胞化学检测增殖细胞核抗原(PCNA)、Bcl2、Bax、Fas、FasL表达。以未转染急性白血病骨髓基质细胞(B组)及正常骨髓基质细胞(C组)作为对照。结果A组骨髓基质细胞培养上清SDF1含量为(384±41)pg/ml,较B组[(2474±271)pg/ml]、C组[(1324±154)pg/ml]明显降低。与之共培养的Jurkat细胞,A组与B组、C组比较,细胞增殖速度减缓,倍增时间延长(A组42h,B组29h,C组33h);细胞周期分析G0/G1期细胞比例增多[A组(28.47±2.39)%,B组(19.43±2.80)%,C组(27.15±2.07)%],S期细胞减少[A组(25.57±1.90)%,B组(74.48±3.23)%,C组(60.99±2.33)%],G2/M期细胞增多[A组(45.96±3.24)%,B组(6.09±1.96)%,C组(11.86±1.98)%];细胞凋亡率增加[A组(15.2±0.8)%,B组(5.4±0.7)%,C组(9.5±0.4)%];PCNA、Bcl2、Fas表达减少,Bax、FasL表达增多。结论阻抑SDF1表达在一定程度上抑制与骨髓基质细胞共培养的Jurkat细胞的增殖活性,并促进其凋亡。
Objective To investigate the effect of inhibiting the expression of bone marrow stromal cell derived factor (SDF1) on the proliferation and apoptosis of Jurkat cells co-cultured with SDF1. Methods SDF1-specific RNA interference plasmids were transfected into acute leukemia bone marrow stromal cells by lipofectamine. The positive clones were screened by G418 (group A), the expression of SDF1 was detected by ELISA. The growth curves were co-cultured with Jurkat cells and calculated Doubling time, cell cycle was detected by flow cytometry, apoptosis was detected by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL), PCNA, Bcl2, Bax, Fas, FasL expression. Untransfected acute leukemia bone marrow stromal cells (B group) and normal bone marrow stromal cells (C group) as a control. Results The level of SDF1 in the supernatant of bone marrow stromal cells in group A was (384 ± 41) pg / ml, which was significantly lower than that in group B [(2474 ± 271) pg / ml] and group C [(1324 ± 154) pg / ml] The Jurkat cells co-cultured with group A showed slower cell proliferation and longer doubling time compared with group B and group C (42 h in group A, 29 h in group B, and 33 h in group C). The proportion of cells in G0 / G1 phase was increased (28.47 ± 2.39% in group A, 19.43 ± 2.80% in group B and 27.15 ± 2.07% in group C). The number of S phase cells in group A (25.57 ± 1.90)% and group B (74.48 ± 3.23) (45.96 ± 3.24)% in group A, (6.09 ± 1.96)% in group B, and (11.86 ± 1.98)% in group C) (15.2 ± 0.8)% in group A, (5.4 ± 0.7)% in group B, and (9.5 ± 0.4)% in group C). The expressions of PCNA, Bcl2 and Fas decreased and the expressions of Bax and FasL increased. Conclusion Inhibition of SDF1 expression inhibits the proliferation of Jurkat cells co-cultured with bone marrow stromal cells to a certain extent and promotes apoptosis.