| Objective:Lung cancer is currently one of the highest mortality of malignant tumor.Since 2008,lung cancer had replaced liver cancer to become the leading cause of the cancer death.Chronic inflammation is now being recognized as a major driving force in the development of about one-third of all the known cancers,including lung cancer.In addition,chronic inflammation induces the expression of various molecules from the tumors themselves or surrounding cells,and promotes the development of tumor-associated immune-suppressive microenvironment.The tumor-associated inflammatory microenvironment consists of myeloid-derived suppressor cells(MDSCs),regulatory T cells(Tregs),tumor-associated macrophage(TAMs),regulatory dendritic cell(DCregs)as well as stroma cells and cancer cells.In recent years,there has been great focus on determining whether alveolar type ?(AT-?)cells function as antigen presenting cells(APCs),because AT-? cells constitutively express major histocompatibility complex molecules(MHC)and contribute to T cell activation.However,AT-? cells expressing high levels of MHC-?,but lacking of CD80 and CD86 costimulatory molecule,could not activate Th1 CD4+ T cells,rather they induced pulmonary tolerance by triggering the differentiation of regulatory T cells(Tregs),which may contribute to immune-suppressive inflammation microenvironment in lung tissues.Recently,we found increased MHC-? expression in alveolar epithelium was observed associated with enhanced Treg infiltration in mice lung tissues with AFG1-induced inflammation and AFG1-induced adenocarcinoma.Furthermore,we treated human AT-? cell model-A549 cells with AFG1 and TNF-a together to mimic an AFG1-induced inflammatory response in vitro,and found they coordinately enhanced MHC-?,CD74,COX-2,IL-10 and TGF-b expression on A549 cells.The findings support that phenotypic upregualton of MHC-? in AT-? may develop a special antigen-presenting ability capable of activating Tregs,which contributes to lung tumorigenesis.However,it is still unclear whether lung adenocarcinoma cells arising from AT-? express higher level of MHC-?.Whether upregulation of MHC-? in AT-? cells contributes to Tregs proliferation in lung adenocarcinoma needs to be elucidated.In this study,we first detected the expression of human major histocompatibility complex(HLA)class ? molecules(HLA-DR),MHC-? invariant chain CD74,and b2,as well as TNF-a and IL-6 in 68 cases of human lung adenocarcinoma and 30 cases of lung squamous cell carcinoma samples,and also explored the relation between HLA-DR expression and Tregs infiltration in tumor-infiltrating T cells in lung adenocarcinoma samples.Then,we applied a urethane-induced chronic inflammation-associated lung adenocarcinoma mice model.We detected the expression of MHC-?,CD74,Tregs specific marker(FOXP3),TNF-a and p-NF-kB in urethane-induced inflamed lung tissues and lung adenocarcinoma.Methods: The expression of HLA-DR?CD74??2?TNF-??IL-6 and COX-2 in 20 cases of normal lung tissue(NLT),68 cases of lung adenocarcinoma(LA)and 30 cases of lung squamous carcinoma(LSC)was detected by immunohistochemistery.The relationship between their expression and the clinical pathological features,as well as the correlation between HLA-DR expression and Tregs infiltration of lung adenocarcinoma was also analyzed.Then we treated the mice with urethane to induce lung chronic inflammation as well as TNF-a/NF-kB mediated chronic inflammation-associated lung adenocarcinoma mice model.We detected the expression of MHC-?,CD74,Tregs specific marker FOXP3,TNF-? and p-NF-kB in urethane-induced inflamed lung tissues and lung adenocarcinoma.Results: 1 Immunohistochemical staining results of HLA-DR,CD74 and ?2 1.1 Expressions of HLA-DR,CD74 and b2 in LA and LSCThe positive immunoreaction of HLA-DR,CD74 and b2 was all located in cytoplasm,which appearing brown particles.The positive expression rate of HLA-DR in LA,LSC and control was 57.4%,10% and 5% respectively,and the results showed that the expression of HLA-DR in LA was significantly higher than that in LSA(P<0.05).The positive expression rate of CD74 in LA,LSC and control was 85.3%,10% and 10% respectively,and the expression of CD74 in LA was significantly higher than that in LSA(P<0.05).The positive expression rate of b2 in LA,LSC and control was 67.6%,20% and 10% respectively,and the expression of b2 in LA was significantly higher than that in LSA(P<0.05).1.2 The clinical pathological significance of HLA-DR,CD74 and ?2 in LA and LSCThe positive rate of HLA-DR in well/moderate-differentiated group(67.4%)was significantly higher than that in low-differentiated group(36.4%,P<0.05).There was no relationship between the positive expression of HLA-DR in LA and the age,sex,lymph node metastasis(P>0.05).In LSC,the expression of HLA did not have any correlation with the age,sex,lymph node metastasis and tumor differentiation(P>0.05).The results showed the expression of HLA-DR in LA was significantly related with tumor differentiation: the lower of the differentiation,the lower of the expression.But no such relation was seen in LSC.The expression of CD74 and ?2 in LA and LSC showed no relationship with the age,sex,lymph node metastasis,tumor differentiation(P>0.05).2 Correlation between HLA-DR and FOXP3 expression in LAIncreased FOXP3 positive Treg cells were observed in some lung adenocarcinoma samples with higher expression of HLA-DR.We found a positive correlation between expression of HLA and Treg marker FOXP3 in LA(r=0.396,P=0.001).The results indicated that higher expression of MHC-? in cancer cells may contribute to Treg difference and infiltration.3 Immunohistochemical staining results of TNF-?,IL-6 and COX-2 3.1 Expressions of TNF-?,IL-6 and COX-2 in LA and LSCThe positive immunoreaction of TNF-?,IL-6 and COX-2 was all located in cytoplasm,which appearing brown particles.The positive expression rate of TNF-? in LA and LSC was 82.4% and 60% respectively,which were all significantly higher than that in normal lung tissue(P<0.05).The expression of TNF-? in LA was significantly higher than that in LSA(P<0.05).The positive expression rate of IL-6 in LA and LSC was 39.7% and 33.3% respectively,which were all significantly higher than that in normal lung tissue(5%,P<0.05).But there were no differences in LA compared with that in LSC(P>0.05).The positive expression rate of COX-2 in LA and LSC was 92.6% and 86.7% respectively,which were all significantly higher than that in normal lung tissue(10%,P<0.05).The COX-2 expression was showed no difference compared with that in LSC(P>0.05).4 Correlation between TNF-? and HLA-DR expression in LAFig3.B showed positive expression of AT-? marker SP-C,TNF-?,HLA-DR and CD74 in the same lung adenocarcinoma.The data supported that lung adenocarcinoma cells arised from AT-? cells,which expressed higher level of TNF-? and HLA-DR.Furthermore,we found a positive correlation between expression of TNF-? and HLA-DR(r=0.459,P=0.000).The results indicated that TNF-? may regulate the MHC-? expression in lung adenocarcinoma.5 The expression of MHC-? and FOXP3 in urethane-induced chronic inflamed lung tissuesThe HE staining results showed that intraperitoneal injection of urethane could induce chronic inflammatory response in alveolar walls since thickened alveolar septum as well as increased inflammatory cells infiltration was observed.Furthermore,immunohistochemical staining showed that there were more CD68,TNF-? and NF-?B positive cells in urethane-treated lung tissues than those in the control mice.The results supported that intraperitoneal injection of urethane induced TNF-?/NF-?B mediated chronic inflammatory response in lung tissues.In the inflammatory environment,we also found higher expression of MHC-? in alveolar epithelial associated with increased the infiltration of FOXP3 positive cells.Thus,the finding indicated that increased expression of MHC-? in alveolar epithelial cells may contribute to Treg infiltration in chronic lung inflammatory environment.6 The expression of MHC-? and FOXP3 in urethane-induced chronic inflammation-associated lung adenocarcinomaSix months later,we found urethane induced lung adenocarcinoma in all urethane-treated mice.We also found higher level expression of TNF-?,p-NF-?B and COX-2 in all lung adenocarcinoma,which suggested that urethane induced TNF-?/NF-?B mediated chronic inflammation-associated lung adenocarcinoma.We also found all lung adenocarcinoma cancer cells expressed SP-C associated with higher level expression of MHC-? and CD74.The results suggested that urethane-induced lung adenocarcinoma originated from AT-? cells which expressed higher level MHC-?.Thus,chronic inflammation may upregulate MHC-? expression in AT-? cells to contribute to lung tumorigenesis.We also detected Tregs marker FOXP3 expression in lymphocytes surrounding the tumor.Increased FOXP3 positive cells were observed in the urethane-induced lung adenocarcinoma.The finding indicated that chronic inflammation-upregulated MHC-? in AT-? cells may contribute to Tregs infiltration in lung adenocarcinoma.Conclusions:1 The expression of HLA-DR?CD74 and ?2 in lung adenocarcinoma was significantly higher than that in squamous carcinoma.2 There was a positive correlation between the expression of HLA-DR and Treg marker FOXP3 in LA.The results indicated that higher expression of MHC-? in cancer cells may contribute to Treg infiltration.3 Lung adenocarcinoma cells arised from AT-? cells,which expressed higher level of TNF-? and HLA-DR.There was a positive correlation between the expression of TNF-? and HLA-DR in cancer cells.The results indicated that TNF-? may regulate the MHC-? expression in lung adenocarcinoma.4 Intraperitoneal injection of urethane induced TNF-?/NF-?B mediated chronic inflammatory response in lung tissues evidenced by increasing expression of TNF-? and NF-?B as well as macrophage infiltration.In the inflammatory environment,higher expression of MHC-? in alveolar epithelial was associated with increased infiltration of FOXP3 positive cells.The finding indicated that increased expression of MHC-? in alveolar epithelial cells may contribute to Treg infiltration in chronic lung inflammatory environment.5 Higher level expression of TNF-?,NF-?B and COX-2 in all urethane-induced lung adenocarcinoma,which suggested that urethane induced TNF-?/NF-?B mediated chronic inflammation-associated lung adenocarcinoma.Lung adenocarcinoma cells originated from AT-? cells overexpressed MHC-? and CD74,which suggested chronic inflammation may upregulate MHC-? expression in AT-? cells to contribute to lung tumorigenesis.Increased FOXP3 positive cells were observed in the urethane-induced lung adenocarcinoma.The finding indicated that chronic inflammation-upregulated MHC-? in AT-? cells may contribute to Treg infiltration in lung adenocarcinoma. |
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