摘要目的 探讨机械力对人肺泡Ⅱ型上皮细胞A549细胞黏附分子-1(ICAM-1)表达的影响.方法 (1)将接种在加力板上的A549细胞分为张力组、压力组和对照组3组,用四点弯曲加力仪对细胞进行机械力刺激:实验组以1000 μstrain强度的张力、压力分别刺激细胞6 h,通过Western blot和半定量逆转录.聚合酶链反应(RT-PCR)检测ICAM-1蛋白及其mRNA的表达,对照组行伪暴露.(2)用特异性细胞外信号调节激酶1/2(ERK1/2)的阻断剂PD98059(浓度为25 μmol/L)预处理3组细胞60 min后,再用1000 μstrain强度的张力、压力分别刺激细胞6 h,以Western blot和RT-PCR法榆测ICAM-1蛋白及其mRNA的表达,对照组在阻断剂处理后行伪暴露.采用单因素方差分析(ANOWA)对数据进行分析,组间两两比较用SNK法.结果 (1)张力组、压力组ICAM-1蛋白吸光度值分别为1.16±0.07、1.05±0.02,高于对照组的0.78±0.07(F值为3.31,P<0.05);张力、压力组ICAM-1 mRNA分别表达为1.42±0.05、1.27 4±0.05,亦高于对照组的0.13 4±0.04(F值为23.1,P<0.01).(2)PD98059预处理细胞后张力组和对照组ICAM-1蛋白吸光度值分别为1.62±0.10和0.50±0.08,差异有统计学意义(q=3.75,P<0.05);压力组ICAM-1蛋白表达水平为0.60±0.03,与对照组相比差异无统计学意义(q=0.32,P>0.05);张力组和对照组ICAM-1 mRNA表达为1.57 ±0.03和0.35±0.29.差异有统计学意义(q=3.51,P<0.05);压力组ICAM-1 mRNA表达为0.46±0.03,与对照组相比差异无统计学意义(q=0.32,P>0.05).结论 机械力可增加A549细胞间黏附分子ICAM-1的表达;阻断剂PD98059可部分抑制机械力诱导的ICAM-1的表达,提示ERK1/2通路可能部分参与了机械力诱导A549细胞ICAM-1表达的信号传导.

abstractsObjective To explore the expression of intercellular adhesion molecule-1 (ICAM-1) in cultured human alveolar type 2 cells (A549) stimulated by mechanical force in vitro. Methods Cells were divided into 3 groups: a tensile stress group, a compressive stress group and a control group. The four-point bending system was used to stimulate A549 cells. The cells were stimulated by tensile stress or compressive stress respectively at the same magnitude of 1000 μstrain for 6 h. Sham cells in control group were not subjected to mechanical loading. The protein level and mRNA level of ICAM-1 were measured by Western blot and RT-PCR. Then an inhibitor was added to further explore the possible mechanism. The cells were divided into a tensile stress + inhibitor group, a compressive stress + inhibitor group and a control group. The cells were pretreated with PD98059, a specific inhibitor of extracellular signal-regulated kinase ( ERK) for 60 min, and then stimulated respectively by tensile stress or compressive stress at the same magnitude of 1000 μstrain for 6 h or were not subjected to mechanical loading. ICAM-1 protein and mRNA concentrations were determined by Western blot and RT-PCR, respectively. The data were analyzed by one-way ANOVA and Student-Newman-Keuls were used to compare 2 means. Results The expression of ICAM-1 protein in the tensile stress group ( 1. 16 ± 0.07) or the compressive stress group (1. 05 ± 0. 02) were significantly higher than that of the control group (0. 78 ± 0. 07, F = 3. 31, P < 0. 05 ) , and the expression of ICAM-1 mRNA in the tensile stress group (1.42 ±0. 05) or the compressive stress group (1. 27 ±0. 05) were also significantly higher than that of the control group ( 0. 13 ± 0. 04, F = 23. 1, P < 0. 01). After pretreated with PD98059 for 60 min, the expression of ICAM-1 protein in the tensile stress group (1.62 ±0.10) was significantly higher than that of the control group ( 0. 50 ± 0. 03, q = 3. 75, P < 0. 05 ) , while there was no significant difference between the compressive stress group (0. 60 ±0. 03, q =0. 32, P >0. 05) and the control group. At the transcription level, the expression of ICAM-1 in the tensile stress group (1.57 ±0.03) was significantly higher than that of the control group ( 0. 35 ± 0. 29, q = 3. 51, P < 0.05 ) , while there was no significant difference between the compressive stress group (0. 46 ±0. 03 , q =0. 32,P >0. 05) and the control group. Conclusions Mechanical forces upregulate the expression of ICAM-1 in A549 cells. PD98059 partly inhibits the upregulation of ICAM-1 induced by mechanical forces. ERK pathway may be partly involved in signal transduction of mechanical force induced expression of ICAM-1 in A549 cells.