| Objective: Colon cancer is common malignancy, the incidence rate is increasing, and the prognosis is poor,about 90% of patients died of distant metastasis or recurrence. The process of tumor invasion and metastasis include: tumor cells escape from primary lesion,across the basement membrane and extracellular matrix, migrate into the surrounding blood vessels or lymphatic vessels to reach distant organs , adhere with endothelial cell, leakage outside the vascular and formate metastases. In the process, a large number of cytokines play an important role at various stages, affect tumor cell invasion and metastasis. Among them, the epidermal growth factor / receptor (EGF / EGFR) related closely to the development of colon cancer ,and it is a research hotspot in recent years. EGFR are transmembrane receptor tyrosine protein kinase, it can activate multiple intracellular signaling pathways, including the Ras / Raf / MAPK, PI3K/Akt, JAK / STAT ,et al. JAK2-STAT3 (Janus kinase 2-signal transducer and activator of transcription3) is a recent flurry of signal transduction. JAK2 binding to corresponding membrane receptors to activate receptor tyrosine and expose STAT3 binding sites, then JAK2 phosphorylate tyrosine of STAT3. phosphorylation STAT3 (P-STAT3) form a dimer into the nucleus, combine with the target gene, regulate cell proliferation, apoptosis, angiogenesis and tumor cell immune evasion and so on. STAT3 can regulate the expression of E-cadherin and Vimentin in cells, and these two proteins have important role in the adhesion and migration of tumor cells. The abnormal expression and activation of STAT3 were found in breast cancer, prostate cancer, lung cancer and other malignant tumors,promote significantly tumor development.In view of these observations, we speculated that epidermal growth factor through activating of JAK2-STAT3 pathway regulate the expression of E-cadherin and Vimentin, promote migration and invasion of colon cancer cells .To this end, this study firstly detected the expression of STAT3, E-cadherin and Vimentin in the colon cancer ,revealed the correlation between these proteins and clinical pathology factors in colon cancer. Then LoVo colon cancer cell groups were treated with EGF and JAK2 kinase inhibitor AG490,to study the effects of JAK2/STAT3 pathway on the expression of E-cadherin and Vimentin and the capability of invasion and migration in LoVo colon cancer cells.To investigate the role and mechanism of JAK2/STAT3 pathway in migration and invasion of colon cancer cells, provide some experimental evidence of prevention and treatment for colon cancer metastasis .Methods:1. Collected 70 cases of colon cancer and corresponding adjacent noncancerous tissues to detect STAT3, E-cadherin, Vimentin expression by immunohistochemical staining, compared the expression of the proteins in serosal invasion, lymph node metastasis, pathological stage (TNM), pathological differentiation type of pathology related factors;2. LoVo Colon cancer cells were treated with the specific JAK2 inhibitor AG490 in different concentrations(10,25,50,100,150μmol/L)and different times(6,12,24,72,48 hours).Cell proliferation was detected using MTT assay. LoVo were treated separately in the control group, EGF group, EGF + AG490 group, AG490 group , cell cycle was detected using flow cytometry , P-STAT3 protein was detected using western blot;3. LoVo Colon cancer cells in the control group, EGF group, EGF + AG490 group, AG490 group were measured the capability of migration and invasion by scratch assay and transwell, detected the location and expression of E-cadherin, Vimentin by immunofluorescence and Western blot.Results:1. The positive rates in immunohistochemical detection of STAT3, E-cadherin, Vimentin were 74.29%,32.86%,78.57% in colon cancer tissues, 15.71% ,82.86% ,12.86% in normal colon tissue, correlated with tumor differentiation, depth of invasion , lymph node metastasis and TNM staging (P<0.05), unconcerned with gender,age,location and tumor size (P > 0.05); Colon cancer tissues STAT3 expression was evident positive correlation with the Vimentin (r = 0.616, P<0.01). negative correlated with E-cadherin (r =-0.702, P<0.01).2. AG490 inhibited proliferation of LoVo colon cancer cells in concentration- time-dependence, the best concentration and time was 50μmol / L and 24 hours. Compared with the control group, EGF group P-STAT3 were significantly increased, cell proliferation index (63.17%±4.25)also increased significantly (P <0.01); EGF + AG490 group and AG490 group P-STAT3 was decreased, cell proliferation index (27.91%±3.16,23.85%±1.07)decreased significantly (P < 0.01).3.①Cell scratch assay: LoVo Colon cancer cells in the control group, EGF group, EGF + AG490 group, AG490 group, the healing scratch area was 38%, 80%, 40%, 24%. Compared with the control group, EGF significantly increased the healing area, the difference was statistically significant (P <0.01); EGF + AG490 and AG490 group decreased significantly, the difference was statistically significant (P <0.01);②Transwell: the number of invasive LoVo cell cells in the control group, EGF group, EGF + AG490 group, AG490 group was 35.24±2.65,55.08±2.14,20.19±3.50,16.52±2.02,compared with the control group, EGF group significantly increased the number of invasive cells, the difference was statistically significant, the difference was statistically significant (P <0.01), EGF + AG490 and AG490 group significantly reduced the number of invasive cells, the difference was statistically significant (P <0.05).③Western blot: Compared with the control group, EGF group E-cadherin was significantly reduced and Vimentin was increased significantly, the difference was statistically significant (P <0.01), fluorescence signal of E-cadherin translocation to the cytoplasm, Vimentin in in the cytoplasm; EGF + AG490 and AG490 group E-cadherin was significantly increased, while the Vimentin was decreased significantly, the difference was statistically significant (P <0.05).Conclusions:1. STAT3, Vimentinand E-cadherin could be used as reference indicators of the malignant behavior in colon cancer, It is helpful to evaluate the invasion and metastasis of colon cancer and prognosis of patients through detecting these protein.2. EGF regulated E-cadherin, Vimentin expression through JAK2/STAT3 pathway, and promoted migration and invasion of LoVo colon cancer cells.3. JAK2 kinase inhibitor AG490 can block JAK2/STAT3 pathway, reducing invasive and migration of LoVo colon cancer cell, thus it may have a role in preventing invasion and metastasis of colon cancer.
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