B7-H1在结直肠癌血管内皮细胞与内皮细胞中的差异性表达及机制

Differential expression of B7-H1 between colorectal carcinoma vascular endothelial cells and endo-thelial cells and the mechanism

目的探讨共刺激分子B7-H1在结直肠癌血管内皮细胞（CCVEC）与内皮细胞（EC）中的差异性表达及机制。方法应用免疫组织化学、实时定量聚合酶链反应（ReM—timePCR）及Westernblot等检测CCVEC与EC的B7-H1差异性表达，并通过白细胞介素（IL）．10、干扰素（IFN）-^y和双因子联合（BFC）的阳性与阴性干预来探讨其机制。差异性表达3组（CCVEC、EC和共培养成纤维细胞组），阳性干预4组（IL-10、IFN-％BFC、CCVEC组），阴性干预4组（IL-10中和组、IFN-1中和组、BFC中和组、CCVEC组），干预前后分别检测EC的B7．Ⅲ表达。结果与EC的阴性表达不同，CCVECB7-H1阳性细胞表达率为（75．00±17．41）％，中位染色评分为3．13；阳性干预IL．10、IFN-1与BFC刺激Ec后，其B7-H1表达率分别为（29．58±4．86）％、（32．08±5．08）％和（34．79±4．40）％，评分为I．46、1．58和1．71（P〉0．05），但与CCVEC差异有统计学意义（P〈0．05）；阴性干预IL．10、IFN-1与BFC中和后，ECB7-H1表达率分别为（36．21±4。05）％、（31．04±3．56）％和（7．02±2．31）％，评分为1．89、1．61、0．62（P〈0．05）。且B7·H1mRNA与蛋白表达比较也证实了该结果。结论B7-Hl为区别EC与CCVEC的分子标志物之-，其表达与肿瘤微环境相关，IL．10和IFN．1为微环境中的重要相关作用因子。

Objective To investigate the differential expression of costimulatory molecules BT-H1 between colorectal carcinoma vascular endothelial cells （CCVEC） and endothelial cells （EC） and the mechanism. Methods Immunohistochemistry, real-time quantitative polymerase chain reaction （ Real-time PCR） and Western blotting were applied to detect the differential expression of B7-H1 between CCVEC and human umbilical vein endothelial cells （HUVEC）. Intervened positively with interleukin （IL）-10 or inter- feron （IFN）-＇y or Bi-factor combined （BFC） and negative handle with anti-IL-10 neutralization antibody Or anti-IFN-~ neutralization antibody or BFC neutralization antibodies to investigate the differential expression and the mechanism of B7-H1 between CCVEC and HUVEC. Firstly, the cells of differential expression of B7-H1 were divided into CCVEC group, HUVEC group, Co-culture HUVEC with Fibroblast group;Then, the cells of positive handle were divided into IL-10 group, IFN-group, Bifactor combined （BFC） group and CCVEC group; And the cells of negative handle were divided into anti-IL-10 neutralization antibodies group, anti-IFN-7 Neutralization antibodies group, Bifactor combined（BFC） group, CCVEC group. Results Different with B7-H1 expression negatively in HUVEC cultured alone,the expression of BT-H1 in CCVEC is strongly positive,and the ratio of B7-H1 positive expression was （75.00±17. 41 ）% ,and the median level score of staining was 3.13;In positive handle group, Stimulated with IL-10 or IFN or BFC, the ratio of positive expression rate in EC were （ 29. 58±4. 86 ） %, （ 32.08±5.08 ） % , （ 34. 79 ±4. 40 ） %, and the median level score of staining were 1.46,1.58,1.71, respectively. There was no significant difference in the median level score among IL-10 group and IFN-3, group and BFC group （P 〉 O. 05 ）, but there was remark- able difference of median staining score with CCVEC （ P 〈 O. 05 ）. In negative handle group, Neutralized with anti-IL-lO neutralization antibodies or anti-lFN neutralization antibodies or bifactor combined neu- tralization antibodies, the ratio of positive expression rate in EC were （36. 21±4. 05 ）%, （31.04±3.56） %, （7.02±2. 31 ） %, and the median level score of staining were 1.89,1.61, O. 62, respectively,which were lower than those in CCVEC group （ P 〈 0.05 ）. Observed the expression of BT-H1 mRNA and the protein which confirmed by the results of cellular immunohistochemistry. Conclusion The cosignalling molecular B7-H1 was one of mark for distinguishing CCVEC from EC. That expression was associated with colorectal carcinoma microenvironment. And cytokines IL-10 and IFN-7 play an important role in colorectal carcinoma microenvironment.