中华肿瘤杂志
中華腫瘤雜誌
중화종류잡지
CHINESE JOURNAL OF ONCOLOGY
2013年
5期
331-336
,共6页
刘诗权%苏颖洁%黄杰安%覃蒙斌%唐国都
劉詩權%囌穎潔%黃傑安%覃矇斌%唐國都
류시권%소영길%황걸안%담몽빈%당국도
结肠肿瘤%鞘氨醇激酶1%黏着斑激酶%细胞黏附分子%细胞增殖%细胞迁移%细胞侵袭
結腸腫瘤%鞘氨醇激酶1%黏著斑激酶%細胞黏附分子%細胞增殖%細胞遷移%細胞侵襲
결장종류%초안순격매1%점착반격매%세포점부분자%세포증식%세포천이%세포침습
Colonic neoplasms%Sphingosine kinase 1%Focal adhesion kinase%Cell adhesion molecule%Cell proliferation%Cell migration%Cell invasion
目的 探讨鞘氨醇激酶l(SphKl)对人结肠癌LoVo细胞增殖、侵袭、迁移、黏着斑激酶(FAK)通路以及细胞间黏附分子1(ICAM-1)和血管细胞黏附分子1(VCAM-1)的影响.方法 采用12-十四酸佛波酯-13-乙酸盐(PMA)和N,N-二甲基鞘氨醇(DMS)诱导和抑制人结肠癌LoVo细胞SphK1的活性,放射自显影法检测SphK1的活性,四甲基偶氮唑蓝(MTT)法检测细胞的增殖活性,Boyden小室观察细胞迁移和侵袭能力的变化,酶联免疫吸附试验(ELISA)测定可溶性ICAM-1(sICAM-1)和可溶性VCAM-1(sVCAM-1)浓度,逆转录聚合酶链反应(RT-PCR)检测FAK、ICAM-1和VCAM-1 mRNA的表达,Western blot法检测FAK和磷酸化FAK(p-FAK)蛋白的表达.结果 PMA和DMS能分别显著地诱导和抑制SphK1活性,PMA呈时间-剂量依赖性促进细胞的增殖,而DMS则呈时间-剂量依赖性抑制细胞的增殖.PMA和DMS处理24 h后,对照组、PMA组和DMS组迁移细胞数分别为(75.48±6.12)个、(143.36±8.73)个和(38.57 ±3.24)个,侵袭细胞数分别为(64.19±5.36)个、(118.46±6.25)个和(32.48±4.27)个,与对照组比较,PMA组迁移和侵袭细胞数明显增多,DMS组则显著减少(均P＜0.01).对照组、PMA组和DMS组的FAK mRNA相对表达水平分别为0.42±0.04、0.82±0.06和0.23±0.02,ICAM-1 mRNA的相对表达水平分别为0.49±0.06、0.74±0.05和0.26±0.03,VCAM-1 mRNA的相对表达水平分别为0.69±0.04、0.89±0.09和0.37±0.04,与对照组比较,差异均有统计学意义(均P＜0.01).对照组、PMA组和DMS组的FAK蛋白相对表达水平分别为0.34±0.04、0.52±0.06和0.20±0.03,p-FAK蛋白相对表达水平分别为0.37 ±0.05、0.51±0.06和0.09±0.02,与对照组比较,差异均有统计学意义(均P＜0.01).slCAM-1和sVCAM-1浓度随着SphK1活性的增强而升高,随着SphK1活性的抑制而降低,与对照组比较,差异均有统计学意义(均P＜0.01).结论 SphK1可能通过激活FAK通路上调ICAM-1和VCAM-1的表达,从而促进LoVo细胞的增殖、迁移和侵袭.
目的 探討鞘氨醇激酶l(SphKl)對人結腸癌LoVo細胞增殖、侵襲、遷移、黏著斑激酶(FAK)通路以及細胞間黏附分子1(ICAM-1)和血管細胞黏附分子1(VCAM-1)的影響.方法 採用12-十四痠彿波酯-13-乙痠鹽(PMA)和N,N-二甲基鞘氨醇(DMS)誘導和抑製人結腸癌LoVo細胞SphK1的活性,放射自顯影法檢測SphK1的活性,四甲基偶氮唑藍(MTT)法檢測細胞的增殖活性,Boyden小室觀察細胞遷移和侵襲能力的變化,酶聯免疫吸附試驗(ELISA)測定可溶性ICAM-1(sICAM-1)和可溶性VCAM-1(sVCAM-1)濃度,逆轉錄聚閤酶鏈反應(RT-PCR)檢測FAK、ICAM-1和VCAM-1 mRNA的錶達,Western blot法檢測FAK和燐痠化FAK(p-FAK)蛋白的錶達.結果 PMA和DMS能分彆顯著地誘導和抑製SphK1活性,PMA呈時間-劑量依賴性促進細胞的增殖,而DMS則呈時間-劑量依賴性抑製細胞的增殖.PMA和DMS處理24 h後,對照組、PMA組和DMS組遷移細胞數分彆為(75.48±6.12)箇、(143.36±8.73)箇和(38.57 ±3.24)箇,侵襲細胞數分彆為(64.19±5.36)箇、(118.46±6.25)箇和(32.48±4.27)箇,與對照組比較,PMA組遷移和侵襲細胞數明顯增多,DMS組則顯著減少(均P＜0.01).對照組、PMA組和DMS組的FAK mRNA相對錶達水平分彆為0.42±0.04、0.82±0.06和0.23±0.02,ICAM-1 mRNA的相對錶達水平分彆為0.49±0.06、0.74±0.05和0.26±0.03,VCAM-1 mRNA的相對錶達水平分彆為0.69±0.04、0.89±0.09和0.37±0.04,與對照組比較,差異均有統計學意義(均P＜0.01).對照組、PMA組和DMS組的FAK蛋白相對錶達水平分彆為0.34±0.04、0.52±0.06和0.20±0.03,p-FAK蛋白相對錶達水平分彆為0.37 ±0.05、0.51±0.06和0.09±0.02,與對照組比較,差異均有統計學意義(均P＜0.01).slCAM-1和sVCAM-1濃度隨著SphK1活性的增彊而升高,隨著SphK1活性的抑製而降低,與對照組比較,差異均有統計學意義(均P＜0.01).結論 SphK1可能通過激活FAK通路上調ICAM-1和VCAM-1的錶達,從而促進LoVo細胞的增殖、遷移和侵襲.
목적 탐토초안순격매l(SphKl)대인결장암LoVo세포증식、침습、천이、점착반격매(FAK)통로이급세포간점부분자1(ICAM-1)화혈관세포점부분자1(VCAM-1)적영향.방법 채용12-십사산불파지-13-을산염(PMA)화N,N-이갑기초안순(DMS)유도화억제인결장암LoVo세포SphK1적활성,방사자현영법검측SphK1적활성,사갑기우담서람(MTT)법검측세포적증식활성,Boyden소실관찰세포천이화침습능력적변화,매련면역흡부시험(ELISA)측정가용성ICAM-1(sICAM-1)화가용성VCAM-1(sVCAM-1)농도,역전록취합매련반응(RT-PCR)검측FAK、ICAM-1화VCAM-1 mRNA적표체,Western blot법검측FAK화린산화FAK(p-FAK)단백적표체.결과 PMA화DMS능분별현저지유도화억제SphK1활성,PMA정시간-제량의뢰성촉진세포적증식,이DMS칙정시간-제량의뢰성억제세포적증식.PMA화DMS처리24 h후,대조조、PMA조화DMS조천이세포수분별위(75.48±6.12)개、(143.36±8.73)개화(38.57 ±3.24)개,침습세포수분별위(64.19±5.36)개、(118.46±6.25)개화(32.48±4.27)개,여대조조비교,PMA조천이화침습세포수명현증다,DMS조칙현저감소(균P＜0.01).대조조、PMA조화DMS조적FAK mRNA상대표체수평분별위0.42±0.04、0.82±0.06화0.23±0.02,ICAM-1 mRNA적상대표체수평분별위0.49±0.06、0.74±0.05화0.26±0.03,VCAM-1 mRNA적상대표체수평분별위0.69±0.04、0.89±0.09화0.37±0.04,여대조조비교,차이균유통계학의의(균P＜0.01).대조조、PMA조화DMS조적FAK단백상대표체수평분별위0.34±0.04、0.52±0.06화0.20±0.03,p-FAK단백상대표체수평분별위0.37 ±0.05、0.51±0.06화0.09±0.02,여대조조비교,차이균유통계학의의(균P＜0.01).slCAM-1화sVCAM-1농도수착SphK1활성적증강이승고,수착SphK1활성적억제이강저,여대조조비교,차이균유통계학의의(균P＜0.01).결론 SphK1가능통과격활FAK통로상조ICAM-1화VCAM-1적표체,종이촉진LoVo세포적증식、천이화침습.
Objective To investigate the effects of sphingosine kinase I (SphKl) on the proliferation,migration and invasion of human colon cancer LoVo cells,and to explore the related mechanisms.Methods Human colon cancer LoVo cells were divided into three groups:phorbol 12myristate 13-acetate (PMA) was used to induce the activation of SphKl in the PMA group,N,Ndimethylsphingosine (DMS) used to suppress the activity of SphKl in DMS group,and the cells treated with equal amount of 0.9 % NaCI instead of drugs served as the control group.The activity of SphKI was assayed by autoradiography,the cell proliferation was assessed by MIT assay,cell migration and invasion were examined by Boyden chamber assay,concentrations of sICAM-1 and sVCAM-1 were assayed by EUSA,and activity of SphK1 was efficiently induced by PMA and significantly suppressed by DMS.PMA induced cell proliferation in a time-and dose-dependent manner.On the contrast,DMS suppressed cell proliferation in a time-and dose-dependent manner.After treating with PMA,the number of migrating and invasing cells were increased to 143.36 ± 8.73 and 118.46 ± 6.25,significantly higher than those of the control group (75.48 ±6.12 and 64.19 ± 5.36).After treating with DMS,the number of migrating and invasing cells were decreased to 38.57 ± 3.24 and 32.48 ± 4.27,significantly lower than those of the control group (P ＜0.01).The relative expression levels of FAK,ICAM-1 and VCAM-1 mRNA in the PMA group were 0.82 ±0.06,0.74 ± 0.05 and 0.89 ± 0.09,and those in the DMS group were 0.23 ± 0.02,0.26 ± 0.03 and 0.37 ± 0.04,with significant differences between the PMA,DMS and control groups (P ＜ 0.01).Compared with the control group,the relative expression levels of FAK and p-FAK proteins in the PMA group (0.52 ± 0.06 and 0.51 ± 0.06) were significantly elevated,and those of the DMS group (0.20 ±0.03 and 0.09 ± 0.02) were significantly decreased.In addition,the concentrations of slCAM-1 and sVCAM-1 were significantly elevated with the activation of SphK1.On the contrary,those of the DMS group were significantly reduced with the suppression of SphK1 (Both P ＜ 0.01).Conclusions SphK1 may enhance the proliferation,migration and invasion of colon cancer LoVo cells through activating FAK pathway and up-regulating the expression of ICAM-1 and VCAM-1.
