The Influence Of IGF-IR Antisense Oligodeoxynucleotide To The Expression IGF-IR Protein And MRNA In Human Esophageal Carcinoma EC9706 Cells Line

Background and objective:Esophageal carcinoma is a kind of common malignant tumor in our country. The data which being publicized by WHO in 1997 indicates that the 46.6% people died of esophageal carcinoma in the world are Chinese, the esophageal carcinoma mortality in china lies in the first place in the world, especially in Henan province. To the explore about the new therapy method of esophageal carcinoma is of momentous significance.A great deal of studies show that the high expression of IGF-IR lies in malignant nervous system tumor, liver cell carcinoma, breast carcinoma, adrenal carcinoma and lung carcinoma, the expression exists in humanmelanoma cells line, spongiocytoma cells line, breast carcinoma cells line, pancreatic carcinoma cells line and liver carcinoma cells line too. The expression of IGF-IR was closely correlated with the pathogenesis, development and metastasis of tumor, but about the expression of IGF-IR in esophageal carcinoma cells haven't been reported. Because IGF-IR plays an important role in pathogenesis and development of many tumor, IGF-IR has become a key target gene in gene therapy of carcinoma, gene therapy of tumor by IGF-IR antisense oligodeoxynucleotide has become a focus. There are some studies show that IGF-IR antisense oligodeoxynucleotide can inhibit osteosarcoma, spongiocytoma, breast carcinoma, colon carcinoma, pancreatic carcinoma and liver carcinoma in vivo and in vitro, but there is no report about the effects of IGF-IR antisense oligodeoxynucleotide to esophageal carcinoma cells. This experiment, through immunohistochemistry SP method and in situ hybridization, investigates the influence of IGF-IR antisense oligodeoxynucleotide to the expression IGF-IR protein and mRNA in human esophageal carcinoma EC9706 cell line, so as to provide some theory basis for target gene therapy of esophageal carcinoma by IGF-IR antisense oligodeoxynucleotide, and to find a new method for therapy malignant tumor. Materials and methods: 1. Three IGF-IR antisense oligodeoxynucleotides(ASODN-tl ,ASODN-t2,ASODN-t3) and one non-oligodeoxynucleotides (N-ODN)were designed.2. Human esophageal carcinoma EC9706 cells were transfected by ASODNs in three kinds of concentrations (150Hg/ml, 200H/ml, 250Mg/ml) respectively.3. To observe the influence of IGF-IR antisense oligodeoxynucleotide on the expression of IGF-IR protein in human esophageal carcinoma EC9706 cells by using immunohisrochemistry.4. To observe the influence of IGF-IR antisense oligodeoxynucleotide on the expression of IGF-IR mRNA in human esophageal carcinoma EC9706 cells by using in situ hybridization.Results:1. IGF-IR protein lies in the cytoplasm of human esophageal carcinoma EC9706 cell, shows brown-yellow granule, there in no positive signal in negative control group. IGF-IR mRNA lies in the cytoplasm of human esophageal carcinoma EC9706 cell, shows blue-purple granule, and there is no positive signal in the cell samples without probe and RNase pretreatment.2. It was significant inhibition of IGF-IR antisense oligodeoxynucleotids in three kinds of concentration to the expression IGF-IR protein in all EC9706 cells ,the most was 250ug/ml ASODN-t3 ,but it was no significant difference in different concentration and different antisense oligodeoxynucleotides (P>0.05). It were significant difference between control group and ASODN-tl, ASODN-t2, ASODN-t3(P<0.05 or P<0.01).3. The expression of IGF-IR mRNA in EC9706 cells were obviouslydepressed by ASODN-tl , ASODN-t2, ASODN-t3, the most was the group of 250ug/ml ASODN-t3. and no significant difference was found in different concentration and antisense oligodeoxynucleotides (P>0.05); it was the obvious difference between control group and N-ODN, ASODN-tl, ASODN-t2,ASODN-t3, (iM).O5 orP<0.01).4. The expression of IGF-IR protein and mRNA were suppressed significantly by three different concentrations of antisense oligodeoxynucleotides, it had a correlation between the suppressive effection for expression of IGF-IR protein and expression of IGF-IR mRNA (r=10.04, P<0.01). Conclusion:l.The expression IGF-IR protein and mRNA in human esophageal carcinoma EC9706 cells line indicates that the IGF-IR is the effective gene substance of esophageal carcinoma cells.2.The IGF-IR expression was correlated with the pathogenesis, development and metastasis of tumor, The expression of IGF-IR protein and mRNA in EC9706 cells were depressed in a certain degree by three different kinds of concentration ASODNs, it indicated IGF-IR ASODNs could inhibit pathogenesis,development and metastasis of esophageal carcinoma or orther malignant tumors through inhibiting the expression of IGF-IR. 3.While three different kinds of concentration IGF-IR ASODNs could inhibitthe expression IGF-IR protein and mRNA in EC9706 cells ,the most was 250 u g/ml ASODN-t3 ,it indicated that the 250 u g/ml ASODN was the best effective inhibition concentration and it provided a very important reference on inhibiting the expression of IGF-IR protein and mRNA in other malignant tumor cells by using IGF-IR ASODN.4.Three different kinds of concentration IGF-IR ASODNs had the suppressive effection not only to the expression IGF-IR protein in EC9706 cells, but also to the mRNA , it had a correlation through statistical analysize.The results indicated that it was probably the more significance in judging prognosis and treatment of esophageal carcinoma by examing the expression of IGF-IR protein and mRNA in the meantime.