The Expression And Clinical Value Of B7-H4in Renal Clear Cell Carcinoma

Objective：1. To investigate the expression and clinical value of B7-H4in thetumor tissue and the blood samples of clear cell renal cell carcinoma;2. To investigate the influences of IL-2, IFN-α and IFN-γ uponB7-H4expression and proliferation activity in clear cell renal cellcarcinoma cells(786-0cells).Methods:1. The B7-H4protein expression in clear cell renal cell carcinomaand the nearby renal tissues was detected by immunohistochemistry in154patients, in order to investigate the relationship between the B7-H4expression level and the clinicopathological characteristics;2. The sB7-H4(secretory B7-H4) expression in the blood of43ccRCCpatients was detected by ELISA to investigate the relationship betweenthe sB7-H4expression and the clinicopathological characteristics;3.786-0cells were cultured in cell culture fluid with IL-2、IFN-αor IFN-γ(100to8000U/ml), CCK-8was carryed out to investigate therelationship between the proliferation activity and the drugconcentration;4.786-0cells were cultured in cell culture fluid with IL-2、IFN-αor IFN-γ(1000U/ml), RT-PCR was carryed out to investigate the expressionof B7-H4mRNA;5.786-0cells were cultured in cell culture fluid with IL-2、IFN-αor IFN-γ(1000U/ml), immunocytochemistry was carryed out to investigatethe expression of B7-H4membrane protein; 6.786-0cells were cultured in cell culture fluid with IL-2、IFN-αor IFN-γ(1000U/ml), ELISA was carryed out to investigate the expressionof sB7-H4;7.786-0cells were cultured in cell culture fluid with IL-2、IFN-αor IFN-γ(1000U/ml), flow cytometry was carryed out to investigate theexpression of B7-H4membrane protein;Results:1.The B7-H4expression was significantly increased in ccRCC tissuescompared to the nearby renal tissues; the positive rate of B7-H4in ccRCCtissues was59.09%(91/154), which is related to clinicopathologicalcharacteristics(TNM stages and clinical stages)（P﹤0.05）,but it isunrelated to the general characyers(gender and age);2. The positive rate of sB7-H4in the blood of ccRCC patients was53.49%（23/43）, which is related to clinicopathologicalcharacteristics(TNM stages and clinical stages)（P﹤0.05）,but it isunrelated to the general characyers(gender and age);3.The proliferation activity of the786-0cells was suppressed moreand more by the ascending concentration of the IL-2、IFN-α or IFN-γ,andthe maximum drug concentration of1%suppression ratio is1000U/ml;4. RT-PCR test showed that the B7-H4mRNA expression was up-regulatedafter stimulated by IL-2、IFN-α and IFN-γ;5. Immunocytochemistry showed that the expression of B7-H4was mainlylocated in the cell membrane, the B7-H4protein expression wasup-regulated after stimulated by IL-2、IFN-α and IFN-γ;6. ELISA showed that each IL-2(44.89±0.97ng/ml)、 IFN-α(46.74±2.25ng/ml) and IFN-γ group (47.31±1.12ng/ml) of the sB7-H4expression was higher than unstimμlated group（34.42±1.69ng/ml）（P﹤0.05）;7. FCS assay showed that each IL-2(44.89±0.94)%、IFN-α (46.41 ±0.55)%, and IFN-γ group (54.18±1.42)%of B7-H4protein expression washigher than unstimμlated group (30.45±0.96)%(P﹤0.05).Conclusion:1. There is an obvious negative correlation between B7-H4expressionand clinical and pathogenic indicators(TNM staging). B7-H4could be usedas a new molecular biology marker to diagnose the ccRCC on early stageand it also can predict the prognosis of the ccRCC patients in the furture;2. IL-2、IFN-α and IFN-γ could up-regulate the expression of B7-H4at the both level of mRNA and protein，the immune escape pathway inducedby B7-H4maybe one of the most important reason why the efficacy of IL-2and IFN-α is low and the efficacy of IFN-γ could not be observed inmRCC.B7-H4could be used as a new molecular biology marker to selecttreatment options of ccRCC patients.