CAJ | 학술논문

目的观察CD44单克隆抗体A3D8对急性单核细胞白血病细胞系THP-1细胞的增殖抑制作用,并探讨其作用途径.方法采用MTT法检测A3D8对THP-1细胞的增殖抑制效应;流式细胞术分析THP-1细胞CD33、CD15、CD11b、CD14、Annexin-V、caspase-3、细胞周期;Western blot法分析p-Akt、磷酸化细胞外调节蛋白激酶(P-ERK)、bcl-2和p27kip1蛋白的表达.结果 A3D8能显著抑制THP-1 细胞增殖,呈剂量和时间的量效关系.2.0 μg/ml A3D8作用THP-1细胞1～6 d后细胞明显分化.A3D8处理4 d,THP-1细胞CD33、CD15、CD11b、CD14表达水平与对照组比较[平均荧光强度(MFI)]分别为68.9±2.0对39.3±1.5、61.7±5.5对12.9±2.6、67.3±3.8对14.0±2.0、83.0±5.7对8.0±1.0(P均<0.01).细胞周期阻滞在G0/G1期.处理4 d实验组和对照组p-Akt、p-ERK、bcl-2蛋白分别为0.24±0.06对1.20±0.15、0.32±0,05对1.24±0.09、0.11±0.05对0.65 ±0.07,实验组较对照组显著减少(P均<0.01);处理4 d实验组和对照组p27kipl.蛋白分别为1.08±0.09对0.10±0.02,实验组较对照组显著增加(P<0.05).A3D8处理5 d THP-1细胞Annexin-V、caspase-3阳性率与对照组比较,分别为(32.5±2.5)%对(2.4±0.3)%、(33.3±2.5)%对(3.6±0.3)%(P均<0.01).结论 CD44单克隆抗体A3D8可能通过抑制PI3K/Akt和ERK1/2信号通路诱导THP-1细胞分化和凋亡.
목적 관찰CD44단극륭항체A3D8대급성단핵세포백혈병세포계THP-1세포적증식억제작용,병탐토기작용도경.방법 채용MTT법검측A3D8대THP-1세포적증식억제효응;류식세포술분석THP-1세포CD33、CD15、CD11b、CD14、Annexin-V、caspase-3、세포주기;Western blot법분석p-Akt、린산화세포외조절단백격매(P-ERK)、bcl-2화p27kip1단백적표체.결과 A3D8능현저억제THP-1 세포증식,정제량화시간적량효관계.2.0 μg/ml A3D8작용THP-1세포1～6 d후세포명현분화.A3D8처리4 d,THP-1세포CD33、CD15、CD11b、CD14표체수평여대조조비교[평균형광강도(MFI)]분별위68.9±2.0대39.3±1.5、61.7±5.5대12.9±2.6、67.3±3.8대14.0±2.0、83.0±5.7대8.0±1.0(P균<0.01).세포주기조체재G0/G1기.처리4 d실험조화대조조p-Akt、p-ERK、bcl-2단백분별위0.24±0.06대1.20±0.15、0.32±0,05대1.24±0.09、0.11±0.05대0.65 ±0.07,실험조교대조조현저감소(P균<0.01);처리4 d실험조화대조조p27kipl.단백분별위1.08±0.09대0.10±0.02,실험조교대조조현저증가(P<0.05).A3D8처리5 d THP-1세포Annexin-V、caspase-3양성솔여대조조비교,분별위(32.5±2.5)%대(2.4±0.3)%、(33.3±2.5)%대(3.6±0.3)%(P균<0.01).결론 CD44단극륭항체A3D8가능통과억제PI3K/Akt화ERK1/2신호통로유도THP-1세포분화화조망.
Objective To investigate the effects of anti-CD44 mAb A3D8 on the cell proliferation of human acute monocytic leukemia cell line THP-1 and its mechanism. Methods Cell proliferation was assayed with MTT method, the expression of CD33, CD15, CD11b, CD14, Annexin-V, caspase-3 and cell cycle with flow cytometry, and the expression of p-Akt, p-ERK, bcl-2 and p27kipl with Western blot. Results A3D8 could remarkably inhibit the proliferation capacity of the THP-1 cells in a dosage- and time-dependent manner. THP-1 differentiation was observed when treated with A3D8 (2.0 μg/ml) for one to six days. Expression of CD33 (68. 9 ±2.0 vs 39.3 ± 1.5), CD15(61. 7 ±5.5 vs 12.9 ±2.6), CD11b (67.3 ± 3.8vs 14.0±2.0) and CD14 (83.0 ±5.7 vs 8.0 ± 1.0) was significantly increased at day 4 compared with the control group ( all P < 0.01). Cell cycle of the THP-1 cells was arrested in G1/G1. Expression of the Annexin-V [(32.5±2.5)% vs (2.4±0.3)%] and caspase-3 [(33.3 ±2.5)% vs (3.6±0.3)%] was much higher than that in normal controls (all P <0. 01) , and apoptosis was observed in THP-1 cells at day 5. Expression of p-Akt (0.24 ±0.06 vs 1.20 ±0.15), p-ERK (0. 32 ±0.05 vs 1. 24 ±0. 09), and bcl-2 (0. 11 ±0.05 vs 0. 65 ± 0. 07) was much lower than that of the controls ( all P < 0. 01), while p27kipl (1.08 ± 0.09 vs 0. 10 ± 0.02) was significantly increased at day 4 (P < 0.05). Conclusion Anti-CD44 antibody can induce the differentiation and apoptosis of THP-1 cell through inhibiting PI3K/Akt and ERK1/2 signaling pathway.