The Role And Prognosis Research Of Inflammation Regulatory Factors GSK-3β In The Occur And Development Of Esophageal Cancer

Objective Our present studies investigated the expression of GSK3β and STAT3 in esophageal squamous cell carcinoma(ESCC), and investigate their relationship with clinical pathological characteristics and prognosis; and investigate the influence of GSK3β and STAT3 inhibitor to esophageal cancer cell line growth、proliferation、migration ability and nude mice tumor, Analysis the regulating relations of GSK3β and STAT3, Discuss their mechanism in the occurrence and development of esophageal cancer.Methods Through RT-PCR and Western blotting assays to detect the expression of mRNA and protein of GSK3β, the expression of GSK3β and STAT3 proteins in esophageal cancer and adjacent tissue were also detected by immunohistochemistry, and analyzed for their correlation with clinicopathological features and prognostic. The expression of GSK3β mRNA in esophageal cancer cell line of different differentiation was also detected by RT-PCR. We also used MTT and Wound-healing Assays to examine the influence of GSK3β and STAT3 inhibitor to esophageal cancer cell line growth、proliferation and migration ability, We injected KYSE70 cells into nude mice subcutaneously, then injected GSK-3β and STAT3 inhibitor into nude mice’s abdominal cavity, observed the change of the time of tumor formation and the size of the tumor. Through immunohistochemistry to detect the expression of GSK3β、STAT3 and P-STAT3 in nude mice’s tumor which have treated with GSK-3β and STAT3 inhibitor. SPSS17.0 software was used for statistical analysis. P values less than 0.05 were considered statistically significant.Results 1. RT-PCR showed that the mRNA expression of GSK3β is significant higher in esophageal cancer tissue, as compared with the adjacent tissues(P<0.05). Furthermore, RT-PCR analyses were also conducted to determine the levels of GSK3β mRNA in four ESC cell lines. The results revealed that the expression of GSK3β was significantly higher in the poor differentiated esophageal cancer(KYSE70) compared with the cell lines which were well and moderately differentiated esophageal cancers(KYSE30、 EC9706、 KYSE140). Western blotting showed that the protein expression of GSK3β is significant higher in esophageal cancer tissue, as compared with the adjacent tissues(P<0.05). As compared with cancerous tissue, phosphorylation of GSK3β(P-GSK3βSer9) substantially increased in para-cancerous tissue. the difference was statistically significant(P<0.05). immunohistochemistry showed that the positive expression of GSK3β and STAT3 showed yellow or brown particles, and mainly located in the cytoplasm, some nuclei were also visible staining. In the immunohistochemistry of 50 ESCC, 64% and 18% of the cases were GSK3β positive in cancer tissue and the adjacent tissues. the difference was statistically significant(P<0.01), High expression of GSK3β in ESCC patients was significant difference with Gender, the degree of differentiation and lymph node metastasis(P<0.05). In the immunohistochemistry of 50 ESCC, 66% and 42% of the cases were STAT3 positive in cancer tissue and the adjacent tissues. The difference was statistically significant(P<0.05), High expression of STAT3 in ESCC patients was significant difference with lymph node metastasis, depth of invasion and TNM stage(P<0.05). In addition to, GSK3β and STAT3 expression were positively correlated. Then we analyze the expression of RT-PCR 、 Western blotting and immunohistochemistry, the results show that the m RNA and protein expression of GSK3β is consistent.2. We follow up 50 patients with esophageal cancer. The results showed that the 30 months survival rates of GSK3β positive group and negative group were 56.3% and 83.3%, respectively, the 30-months survival rate of STAT3 positive group and negative group were were 51.4% and 86.6%, respectively. The Kaplan-Meier survival analysis indicated compared with negative group, the positive expression group have a poor prognosis.3. MTT、Wound Healing assays and animal experiments found that after the KYSE30 and KYSE70 cell line and the rodents were treated with GSK3β and STAT3 inhibitors, The cell and tumor growth is restrained obviously, if the cell were treated with LiCl and WP-1066 at the same time, the inhibition effect were further enhanced. the immunohistochemistry result show that the esophageal cancer cell were treated with GSK3β and STAT3 inhibitor,the expression of GSK3β、STAT3 and P-STAT3 were changed. The results suggested GSK3β may potentially regulate STAT3 to affect the occurrence and development of esophageal cancer.Conclusions 1. The expression of GSK3β and STAT3 in esophageal cancer is significantly higher than in the adjacent tissue, which suggested GSK3β and STAT3 may be participated in the occurrence and development of esophageal cancer. There may be a synergistic effect exist in the procession of the occurrence and development of ESCC. GSK3β and STAT3 may be prognostic marker of ESCC patients.2. GSK3β and STAT3 inhibitors may suppressed the ability of proliferation、migration of esophageal cancer cells and nude mice tumor; GSK3β may potentially regulate STAT3 to affect the occurrence and development of esophageal cancer. GSK3β and STAT3 can be the molecular targets of esophageal cancer treatment.