| Objective:In order to detect the expression of SOX9 in lung cancer tissues and analyze the relationship between the expression of SOX9 in non-small cell lung cancer and clinical pathological factors and prognosis of patients.To further explore the clinical value of SOX9 for the clinical diagnosis and treatment of lung cancer.Methods:The SOX9 gene with significant differential expression in lung cancer tumor stem cells was screened by bioinformatics analysis technology in advance.First,40 patients with non-small cell lung cancer in our hospital were collected for postoperative lung cancer tissue,paracancerous tissue and clinical data.Real-time fluorescence quantitative PCR(qRT-PCR)was used to detect SOX9 mRNA expression level in specimen tissues.The expression level of SOX9 protein in lung cancer tissues was detected by Western blotting and immunohistochemistry to verify and analyze whether SOX9 is overexpressed in non-small cell lung cancer tissues.Another 87 samples of human lung cancer tissue chips(HLugA180Su06)collected from June 2004 to June 2009 were purchased to further expand the tissue sample size,including 87 paraffin blocks of lung cancer tissue samples and patient details,followed up for 10 years.Hematoxylin-eosin(HE)staining method was used to determine the pathological diagnosis of paraffin blocks in each tissue specimen.The expression level of SOX9 protein in lung adenocarcinoma patients was analyzed by immunohistochemistry,and the relationship between SOX9 and clinicopathological features and prognosis were analyzed.The relationship between SOX9 and survival prognosis of patients was verified by Kaplan-Meier plot online database.Results:1.Compared with the control cells,many genes(including SOX9 gene)are highly expressed in lung cancer globular(dry)cells.The overexpression of SOX9 gene was significantly different.2.Of the 40 patients,31(77.5%)were lung adenocarcinoma,7(17.5%)were lung squamous cell carcinoma,and 2(5.0%)were other types.And the expression of SOX9 mRNA and protein in cancer tissues was significantly higher than that in the matched adjacent tissues.(P<0.05).3.Expanded sample size experiment verification:(1)HE staining results showed that 87 cases of lung cancer tissues were diagnosed as lung adenocarcinoma;(2)According to the clinical data of the patients,the clinical stages were 28 cases in stage I,25 cases in stage II,32 cases in stage III and 1 case in stage IV.Lung adenocarcinoma mainly occurred in the elderly(50,57.5%),male patients 47(54.1%)were slightly higher than that of female patients(40,45.9%),33(45.9%)of 87 patients had different degrees of lymph node metastasis,39(54.1%)had no lymph node metastasis.The overexpression of SOX9 was detected in 87 lung adenocarcinoma tissues.There were 45 cases(51.7%)in the high expression group and 42 cases(48.3%)in the low expression group.Twenty eight patients survived in 10 years of follow-up,the survival rate was 32.1%,59 patients died,the mortality rate was 67.8%.(3)The results of SOX9 staining in cytoplasm showed that there were 41 cases with high expression rate(52.3%),while the expression of SOX9 protein in paracancerous tissues was lower(78.100%)(χ2=55.617,P<0.001).The expression of Sox9 in the nucleus of lung cancer was also significantly higher than that in the adjacent tissues(χ2=55.841,P<0.001).All of the above showed that the expression of Sox9 protein in lung adenocarcinoma was significantly higher than that in adjacent tissues.4.Correlation between SOX9 Expression in Cytoplasm and Clinicopathological Factors : Compared with patients without lymph node metastasis(N0),the expression level of SOX9 in lung adenocarcinoma patients with lymph node metastasis(N1~N3)was significantly increased(χ2=6.998,P = 0.008);Compared with T1/T2 patients,SOX9 expression in lung cancer tissues of patients with T stage T3/T4 was significantly increased(χ2=5.202,P=0.023);Compared with the patients with clinical stage I/II,the higher the clinical stage,the more obvious the high expression of SOX9 is(χ2=7.362,P=0.007).5.The correlation between the expression of SOX9 protein in the cytoplasm and the degree of pathological malignancy and clinical stage:(1)The results showed that the expression of SOX9 in cytoplasm of high,medium and low differentiated tissues increased obviously with the decrease of differentiation degree.At the same time,the positive expression rate of SOX9 in the cytoplasm of the patients was counted.It was found that the expression of SOX9 increased with the decrease of differentiation.But there was no significant difference between the low differentiation group and the middle differentiation group(P>0.05).(2)Compared with the clinical stage,the higher the clinical stage,the higher the expression of SOX9 in the cytoplasm of the lung adenocarcinoma.In contrast,SOX9 is almost undetectable in normal lung tissue.6.The relationship between the expression level of SOX9 in plasma and the prognosis of patients with lung adenocarcinoma: the median survival time of patients with high expression of SOX9 in plasma was 30 months(95% CI,24.367-35.633 months),while that of patients with low expression was 56 months.The median survival time of the group with high SOX9 expression was significantly lower than that of the group with low SOX9 expression,and the 10-year survival rate was 32.1%,the difference was statistically significant(P<0.001).The results of keplan meierplotter database were consistent with the above prognosis.7.The expression level of SOX9 in the nucleus of female patients with lung adenocarcinoma was significantly higher than that of male patients(χ2=7.878,P=0.005);compared with patients with lower TNM stage,the expression of SOX9 in the nucleus of patients with higher TNM stage was significantly higher(χ2=4.185,P=0.041).However,there was no significant relationship between the expression of SOX9 and the age,pathological grade,T stage,N stage and distant metastasis(P>0.05).8.It was found that the median survival time of patients with high expression and low expression of SOX9 in nucleus was 39 months.There was no difference in the median survival time between the high expression group and the low expression group(P=0.154).9.The univariate Cox analysis of the case characteristics of patients with high and low expression of SOX9 in the plasma showed that the differences of SOX9 expression level(HR=2.703,P=0.001),pathological grade(HR=1.965,P=0.007),lymph node metastasis(HR=1.748,P<0.001)and TNM stage(HR=1.701,P=0.001)were significant affect the survival time of patients.Cox multivariate survival regression analysis showed that SOX9 expression level(HR=2.414,P=0.022)and N stage(HR=1.451,P=0.044)were independent risk factors for survival and prognosis of lung cancer patients.Conclusion:1.Combined with biological information and clinical analysis,it is suggested that SOX9 is highly expressed in lung cancer,suggesting that SOX9 may be a potential functional gene to promote the development of non-small cell lung cancer,and providing a theoretical basis for further study of SOX9 function in non-small cell lung cancer.2.With the decrease of differentiation degree of lung adenocarcinoma,the expression of SOX9 in cytoplasm tends to increase.We believe that the malignant degree of lung adenocarcinoma cells may be related to SOX9 expression in cytoplasm.3.The higher the clinical stage of lung adenocarcinoma patients is,the higher the SOX9 expression level in cancer tissues is both in cytoplasm and nucleus.And the overall survival time of lung adenocarcinoma patients with high expression of SOX9 in cytoplasm is obviously shortened.This suggests that SOX9 can be used as an important index for prognosis of lung adenocarcinoma patients at various stages.The analysis of risk factors indicates that the high expression of SOX9 in cytoplasm is an independent risk factor affecting the prognosis of lung adenocarcinoma patients.4.The expression of SOX9 in cytoplasm is correlated with T stage,lymph node metastasis,TNM stage and survival status of lung adenocarcinoma patients.Although the expression level of SOX9 in nucleus is also related to TNM stage of patients,it is not related to survival time of patients,suggesting that the expression level of SOX9 in cytoplasm may be the main factor to promote the development of lung adenocarcinoma patients.5.The results of SOX9 protein expression in cytoplasm and nucleus are different,that is to say,the prognosis of patients with high SOX9 expression in cytoplasm is significantly shortened,while the survival time of patients with high SOX9 expression in nucleus is not affected,suggesting that SOX9 may play a role mainly in cytoplasm.6.The expression level of SOX9 was different in different genders. |
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