CAJ | 학술논문

目的探讨人髓系白血病细胞株HL-60在诱导分化过程中血管内皮生长因子(VEGF)表达及分泌水平变化的分子机制.方法四甲基偶氮唑蓝(MTT)法检测HL-60细胞被全反式维甲酸(ATRA)诱导分化后的增殖水平变化,流式细胞术测定HL-60细胞CD11b表达和细胞周期的改变,半定量逆转录聚合酶链式反应(RT-PCR)检测HL-60细胞VEGF、c-myc、缺氧诱导因子(HIF)-1α、基质金属蛋白酶(MMP)-9和MMP-2 mRNA表达水平,应用酶联免疫吸附试验(ELISA)法检测药物作用前后HL-60细胞培养上清液中VEGF蛋白的含量.结果经ATRA诱导分化后,HL-60细胞增殖水平明显降低,CD11b表达水平明显升高,出现粒系定向分化趋势,且分化程度明显升高(P<0.05),c-myc与VEGFmRNA表达水平均明显下调(P<0.05),HIF-1α mRNA表达水平则明显升高(P<0.05).用MMP-2及MMP-9抑制因子Ⅰ阻断HL-60细胞MMP-9和MMP-2表达后,细胞培养上清中VEGF蛋白分泌量明显降低(P<0.05).结论 HL-60细胞被诱导分化过程中VEGF的表达与c-myc表达正相关,与HIF-1α表达负相关.MMP-9、MMP-2可能是调控HL-60细胞分泌VEGF的主要原因之一.
목적 탐토인수계백혈병세포주HL-60재유도분화과정중혈관내피생장인자(VEGF)표체급분비수평변화적분자궤제.방법 사갑기우담서람(MTT)법검측HL-60세포피전반식유갑산(ATRA)유도분화후적증식수평변화,류식세포술측정HL-60세포CD11b표체화세포주기적개변,반정량역전록취합매련식반응(RT-PCR)검측HL-60세포VEGF、c-myc、결양유도인자(HIF)-1α、기질금속단백매(MMP)-9화MMP-2 mRNA표체수평,응용매련면역흡부시험(ELISA)법검측약물작용전후HL-60세포배양상청액중VEGF단백적함량.결과 경ATRA유도분화후,HL-60세포증식수평명현강저,CD11b표체수평명현승고,출현립계정향분화추세,차분화정도명현승고(P<0.05),c-myc여VEGFmRNA표체수평균명현하조(P<0.05),HIF-1α mRNA표체수평칙명현승고(P<0.05).용MMP-2급MMP-9억제인자Ⅰ조단HL-60세포MMP-9화MMP-2표체후,세포배양상청중VEGF단백분비량명현강저(P<0.05).결론 HL-60세포피유도분화과정중VEGF적표체여c-myc표체정상관,여HIF-1α표체부상관.MMP-9、MMP-2가능시조공HL-60세포분비VEGF적주요원인지일.
Objective To explore molecular mechanism of expression of vascular endothelial growth factor (VEGF) mRNA and secretion of VEGF protein in HL-60 cells induced by all-trans refinoic acid (ATRA). Methods MTr method was used to detect the proliferation of HL-60 cells induced by ATRA,cell cycle and CD11b expression in HL-60 cells were detected by flow cytometry. Expression of VEGF, c-myc, by-poxia-inducible factor(HIF)-lα, matrix metalloproteinase (MMP)-9 and MMP-2 mRNA were detected by semi-quantitative RT-PCR. VEGF protein in HL-60 cells culture supernatant was measured by ELISA before and after being induced by ATRA. Results After treatment with ATRA,the proliferation of HL-60 cells was obviously inhibited, CD11b expression increased, trend of granulocyte directional differentiation emerged, and differentiation degree was increasd(P <0. 05) ;expression level of c-my and VEGF mRNA was down-regulated (P < 0. 05), but expression level of HIF-1α mRNA was up-regulated (P < 0. 05). VEGF protein level in HL-60 cells culture supernatant was decreased by blocking the expression of MMP-9 or MMP-2(P <0. 05).Conclusion VEGF expression has positive correlation with c-myc expression,but has negative correlation with HIF-1α expression. MMP-9 and MMP-2 may be the main factors regulating VEGF secretion in HL-60 cells.