The Clinical Significance Of HLA-G And Its Mechanism In Gastric Cancer

BACKGROUND AND OBJECTIVES: Gastric cancer(GC)is a common malignant tumor,with high morbidity and mortality rate.The immunoediting and tumor escape of cancer cells play a vital role in carcinogenes is.Human leukocyte antigen-G(HLA-G)is a kind of non-classical major histocompatibility complex antigen.Accumulating data showed that aberrant expression of HLA-G plays an important role in carcinogenes is and is associated with prognosis.However,the c linical and pathogenic role of HLA-G in GC is still not well understood.This study was to investigate the clinical significance and potential mechanism of HLA-G about tumor escape in GC,and validate the impact of HLA-G-effected NK cells in GC progression in vivo.METHODS:(1)Expression of HLA-G in GC tissue from forty-nine patients was analyzed by immunohistochemistry and Western-blot.Tumor-infiltrating NK cells were analyzed by immunohistochemistry,and the relevance between HLA-G and infiltrating NK cells was also evaluated.(2)Clinical and laboratory data were collected and the clinical significance of HLA-G and tumor-infiltrating NK cells was evaluated.Survival analys is was also analyzed based on HLA-G.(3)Tumor-infiltrating NK cells was isolated and the expression of surface receptors on NK cells was analyzed by flow cytometry.(4)The expression of HLA-G on GC cell lines(including MKN-28,MKN-45,SGC-7901,AZ-521,MGC-803 and BGC-823)was evaluated by Western-blot and q PCR.The expression of immunoglobulin-like transcript 2(ILT2)and killer cell immunoglobulin-like receptor 2DL4(KIR2DL4)on NK-92 MI cell lines was evaluated by Western-blot.(5)MGC-803 cell lines with HLA-G overexpression and NK-92 MI cell lines with ILT2 overexpression were constructed by lentivirus transfection.(6)NK-92 MI cell lines with different expression of ILT2 and GC cells lines with different expression of HLA-G were co-cultured.The effect of HLA-G on NK cells proliferation was examined by Cell Counting Kit-8(CCK8)assay.LDH release assay was used to evaluate the effect of HLA-G on the cytotoxic activity of NK cells,the levels of IFN-? and TNF-? in co-cultured supernatant were detected by ELISA.(7)Mice bearing a xenograft tumor model were used to examine the effect of HLA-G on the anti-tumor effect of NK cells.RESULTS:(1)HLA-G positive expression was detected in most of the GC tissues(61.22%),and GC patients with HLA-G positive had the characters of preoperative anemia(P=0.021),severer T stage(P=0.012),severer N stage(P=0.015)and severer AJCC stage(P=0.001).HLA-G was also correlated with poor prognosis in GC patients.(2)The expression of HLA-G was negatively associated with the number of tumor-infiltrating NK cells in GC tissues.ILT2 was the major kind of receptors expressed on tumor-infiltrating NK cells.GC patients with less tumor-infiltrating NK cells showed severer N stage and AJCC stage.(3)GC cell lines,inc luding MKN-28,MKN-45,SGC-7901,AZ-521,MGC-803 and BGC-823,expressed a relatively lower level of HLA-G.Both ILT2 and KIR2 DL were lower expressed on NK-92 MI.In co-culture experiments,GC cell lines with overexpression of HLA-G showed the ability to inhibit the cell proliferation and cytotoxic activity of NK-92 MI cells,as well as reduce the secretion of IFN-? and TNF-? through ILT2.(4)In vivo experiment showed that GC cells lines with HLA-G overexpression can resist the anti-tumor effect of NK cells with ILT2 overexpression incompletely,with a larger tumor size.CONCLUSIONS:(1)HLA-G positive expression was seen in most GC patients,and the expression of HLA-G was strongly correlated with the disease severity and adverse prognosis of GC.(2)The amount of tumor-infiltrating NK cells was negatively correlated with HLA-G expression in GC tissue,and associated with N stage and AJCC stage of GC.(3)GC cells with HLA-G overexpression might inhibit the proliferation,cytotoxic activity and secretion of IFN-? and TNF-? of NK cells through ILT2.(4)In vivo experiments indicated that GC cells can inhibit the antitumor-effect of NK cells though HLA-G and ILT2 interaction.