Study On The Effect Of TAMs On Activation Of STAT3 And Inhibition Of Apoptosis In Non-small Cell Lung Cancer Cells

Background:Lung cancer has always been one of the cancers with the highest morbidity and mortality in the world.Among them,non-small cell lung cancer accounts for about 80% of the incidence.About 50-70% of the patients are diagnosed the advanced stage with a extremely low five-year survival rate.Tumormicroenvironment(TME),also known as "tumor stroma",is the living space of tumor cells.Tumor associated macrophage(TAM),which exist widely,can secrete a variety of cytokines and activate specific cellular pathways,which play an indispensable role in the occurrence and development of tumors.Objective:To verify the reliability of labeling TAMs with CD206 and CD163 in non-small cell lung cancer(NSCLC),and to observe the expression of TAMs in non-small cell lung cancer and its clinicopathological features,and to explore the relationship between them.To investigate the effect of TAMs cells on STAT3 and apoptosis of non-small cell lung cancer cell lines A549 and PC-9.Methods:1.60 cases of NSCLC and their surrounding normal lung tissues were treated by immunohistochemical method.The expressions of TAMs surface markers CD206,CD163,STAT3 and p-STAT3 were observed,and the correlation between them was explored.2.THP-1 was induced into TAMs,by PMA combined with IL-4,and the expression of CD206 and CD163 was detected by PCR to determine whether it was successfully induced into TAMs.3.The supernatant of successfully induced TAMs cells was used as conditioned medium(CM),to co-culture non-small cell lung cancer cell line A549 and PC-9,with AG490,an inhibitor of JAK2/STAT3 pathway,to observe the effect of CM on apoptosis of non-small cell lung cancer cells A549 and PC-9.4.Western Blot method was used to detect the expression level of JAK2,STAT3 and p-STAT3 in the two kinds of cells to determine whether to activate the corresponding pathway,and to detect the expression level of anti-apoptosis protein BCL-2 and apoptosis protein Bax.Finally,the relationship between apoptosis and pathways was analyzed and its mechanism was discussed.Results:1.The number of TAMs infiltration in the stroma of non-small cell lung cancer(squamous cell carcinoma and adenocarcinoma)was significantly higher than that in normal tissue stroma,and the expression of STAT3 and p-STAT3 protein in non-small cell lung cancer tissues was significantly higher than that in adjacent normal tissues.There was no significant correlation between the expression of TAMs,STAT3 and p-STAT3 and the sex,age,depth of invasion,lymph node metastasis,histological grade and TNM clinical grade in patients with non-small cell lung cancer.There was a significant positive correlation between CD206(+),CD163(+)positive cells and p-STAT3 expression in the stroma of non-small cell lung cancer(NSCLC),and the consistent rate was 86.6%.2.THP-1 can be successfully induced into TAMs in the co-culture of PMA and IL-4.3.The supernatant of TAMs cells can increase the malignant degree of non-small cell lung cancer cells.4.The supernatant of TAMs cells could activate JAK2/STAT3 pathway in non-small cell lung cancer cells,which significantly decreased the expression of anti-apoptotic protein Bcl-2,while significantly increased the expression of apoptosis-related protein Bax.Conclusions:TAMs of tumor-associated macrophages in the stroma of non-small cell lung cancer can activate STAT3 signal pathway.Phorbol ester PMA combined with IL-4 can induce human monocyte THP-1 to differentiate into TAMs,which is positively correlated with the concentration of IL-4.Cytokines,that were secreted by TAMs,can activate JAK2/STAT3 signal pathway,and inhibiting the apoptosis of NSCLC.