摘要
目的:观察转化生长因子-β(TGF-β)和干扰素-γ(IFN-γ)对黑色素瘤细胞的增殖、迁移和侵袭能力的影响。方法:体外培养黑色素瘤细胞,待细胞长到80%融合度时,加入TGF-β(浓度为5ng/mL)和IFN-γ(浓度为10ng/mL)因子,将黑色素瘤细胞分为对照组、TGF-β组、IFN-γ组和TGF-β+IFN-γ组,然后各组分别采用磺酰罗丹明B(SRB)增殖测定法观察黑色素瘤细胞培养后8h、16h、24h、32h、40h和48h时间点的增殖情况,于540nm吸光度下比色读取吸光度(A540)来表示细胞的增殖变化。采用200μL吸头对各组细胞的表面进行划痕处理,随后于12h观察划痕之间距离(即代表肿瘤细胞迁移的能力)的变化。采用测Transwell系统孔外膜上细胞的595nm吸光度(A595)来表示肿瘤细胞的侵袭能力。采用明胶酶谱法观察各组肿瘤细胞的MMP-2、MMP-9活性水平,用图像分析软件测定聚丙烯酰胺凝胶中MMP-2、MMP-9的条带灰度(代表MMPs的活性水平)。结果:TGF-β可以增强黑色素瘤细胞的增殖、迁移和侵袭能力(P<0.05),IFN-γ则抑制黑色素瘤细胞的增殖、迁移和侵袭能力(P<0.05),二者同时处理则对黑色素瘤细胞的影响作用微弱或消失(P>0.05)。结论:体外实验中,TGF-β可以干扰IFN-γ对黑色素瘤细胞的增殖、迁移和侵袭能力的抑制作用,本研究可为炎症和肿瘤的相互关系研究奠定实验基础。
Objective： To investigate the influence of TGF-β and IFN-γ on the proliferation, migration and invasion of melanoma cells. Methods： Melanoma cells were cultured in vitro. When tumor cells were confluent about 80% degree, cytokines were added into cell culture media. The concentration of TGF-β and IFN-β was 5ng/mL and 10ng/mL, respectively. Melanoma cells were divided into free-cytokine group, TGF-β group, IFN-γ group, TGF-β and IFN-γ group. Tumor cells in each group were then incubated for 8h, 16h, 24h, 32h, 40h and 48h, respectively. After incubation, fixing and staining with SRB, the optical densities and percentage viability were then determined by absorption at 540 nm （A 540）. The scarification of tumor cells in each group on the surface was created by a 200μ/L pipette tube. The motility of tumor cells in each group was assessed by measuring the distance between scarifications. The speed of the scuffing closure was monitored after 12h. The invasive ability of melanoma cells was observed by transwell cultivation. The tumor cells that invaded through the Matrigel and adhered to the bottom of the outside membrane were determined by absorption at 595 nm （A595）. Gelatin zymography assay was used to examine the levels of matrix metalloproteinases-2 （MMP-2） and matrix metalloproteinases-9 （MMP-9） activity when the tumor cells were treated with cytokines after 24h. MMP-2 and MMP-9 activity was demonstrated by gradation in the sodium dodecyl sulfate polyacrylamide gel electrophoresis （SDS-PAGE） gelatin. MMP-2 and MMP-9 activity was determined by Image analy-sis Software. Results： TGF-β promoted the proliferation, migration and invasion of melanoma cells （P〈0.05）. However, IFN-γ inhibited the proliferation, migration and invasion of melanoma cells （P〈0.05）. The effect weakened or disappeared when both of them were used （P〉0.05）. Conclusion： In vitro, TGF-β may affect the inhibitory effect of IFN-γ on the proliferation, migration and invasion of melanoma cells.
出处
《中国肿瘤临床》
CAS
CSCD
北大核心
2010年第3期134-137,共4页
Chinese Journal of Clinical Oncology
基金
国家自然科学基金(编号:30830049)
中瑞国际合作项目基金资助(编号:09ZCZDSF04400)~~
