Expressions Of E-cad,CD44S And PCNA In The Process Of Tongue Carcinogenesis In Rats Induced By 4NQO

Objective:Oral cancer is a malignant tumor that seriously affects the quality of human life,accounting for an significant proportion in malignant tumors happened in whole body,in head and neck.Under the complex of human living environment,the effects of adverse factors in the environment on human health have increased.In recent years,oral cancer is on a trend of gradually rising.Correlative researchs of demographic and clinical discribed that smoking and drinking man around average age of 58 were easily subject to such malignant tumor.According to statistics,there were about 170,000 new cases of male oral cancer worldwide annually.Therefore,a large number of scientific research personnel devote themselves into the study of its pathogenesis.Tongue squamous cell carcinoma is the most common type of oral cancer and more and more study in-depth of related factors in pathogenesis are in progress.Its carcinogenesis is a complex process incuding many phases.When normal cells changes into cancer cells by relevant functions,they needs to integrate large amounts of gene expression links.Cell adhesion molecule,tumor stem cell markers,cell proliferation markers have important and indicative functions in these process.Qualitative and quantitative research on the changes of the expressions of these proteins which occur in the malignant tumor development process are of great advantage to further clarify the mechanism of carcinogenesis,to the prevention and control of the occurrence and development of malignant tumor.The study will help patients in greatest degree.It has been proved that the positive expression of cell adhesion molecule E-cad on the epithelium tissue is closely related to oral squamous cell carcinoma by a large number of domestic and foreign researchs.the positive expression of it can be used as an important index to judge the prognosis of patients.CD44 is the most common cancer stem cell surface marker.Its positive expression has been identified in breast cancer,gastric cancer and other malignant tumors.PCNA is one kind of proteins which are generated and exist in the nucleus.It has been widely recognized that PCNA can be used as an index to evaluate the state of cell proliferation.This study will explore the expressions of these proteins in the process of oral carcinogenesis in rats.Methods:The experiment was divided into two parts.The first part was to establish the tongue squamous cell carcinoma model in SD rats under closed barrier environment.The mass fraction of 4NQO 0.004%aqueous solution was fed to 90 SD male rats as drinking water.They were drinking freely and the changes of rats’ oral mucosa were observed.The 90 experimental rats were killed at the end of the 10th,14th,18th,22nd and 24th weeks after the start of the experiment respectively.Extract the oral mucosa separately and all the specimens were fixed in 4%mass fraction of poly formaldehyde solution.Paraffin embed the specimens within 24 hours,slice and use HE staining to determine pathological grade and the nature of the lesions.The immunohistochemical streptavidin-perosidase(SP)method was used in the second phase for all specimens to examinate the expressions of CD44S,E-CAD,PCNA to explore the relationship between the expressions and the formation of the tongue squamous cell carcinoma.The results were statistically analyzed by the SPSS 20.0 software.Results:Part one,8 rats died in the process and finally got 82 effective subjects.After pathological grading for the collected samples of rats’ oral mucosa,epithelial rat tongue mucosa,specimens in the phases of normal mucosa,simple hyperplasia,mild dysplasia,moderate dysplasia,severe dysplasia,and squamous cell carcinoma were obtained(Table 1).Tissue samples from the normal tongue mucosa to tongue carcinoma,totally six stages,were obtained.About the sample size differences between different pathological grading,we use multiple independent sample grades data of the Kruskal-Wallis rank sum test.There was statistical Significa--nce of the sample size difference between the pathological grades of tongue mucosal(x2=24.267,P=0.000,P<0.05).There was no statistical significance of the sample size difference between the pathol--ogical grades of buccal mucosal(x2=3.683,P=0.451,P>0.05).There was statistical significance of the sample size difference between the pathological grades of palatal mucosal(x2=14.03,P=0.015,P<0.05).Part Two,the immunohistochemistry test(SP method)was used to detect the expressions of E-cad,CD44S,PCNA in the lingual mucosa epithelium under in normal,hyperplasia,mild dysplasia,severe dysplasia,and squamous cell carcinoma stages.The positive rate of the three proteins respectively are follwed.For the expression of E-cad,the normal mucosa 100%(21/21)is positive.Epithelial hyperplasia 95.24%(20/21),mild epithelial dysplasia 92.86%(13/14),moderate and severe epithelial dysplasia 80%(8/10)as well as tongue squamous cell carcinoma 68.75%(11/16)are all positive.For the expression of CD44S,the normal mucosa71.43%(15/21)is positive.Epithelial hyperplasia 76.19%(16/21),mild epithelial dysplasia 85.71%(12/14),moderate and severe epithelial dyspl--asia 90%(9/10)as well as tongue squamous cell carcinoma 93.75%(15/16)are all positive.For the expression of PCNA,the normal mucosa 9.52%(2/21)is positive.Epithelial hyperplasia 14.29%(3/21),mild epithelial dysplasia 35.71%(5/14),moderate and severe epithelial dysplasia 50%(5/10)as well as tongue squamous cell carcinoma 56.25%(9/16)are all positive.The positive expression of E-cad in tongue squamous cell carcinoma was significantly lower than that of in the abnormal proliferation of lingual mucosa and normal mucosa.The difference were statistically significant(χ2= 15.308,p=0.004,P<0.05).The positive expression of CD44S in tongue squamous cell carcinoma was significantly higher than that of in the abnormal proliferation of lingual mucosa and normal mucosa.The difference were statistically significant(χ2=10.143,p=0.038,P<0.05).The positive expression of PCNA in tongue squamous cell carcinoma was significantly higher than that of in the abnormal proliferation of lingual mucosa and normal mucosa.The positive expression was gradully incresed to higher level from normal mucosa dysplasia to abnormal proliferation of mucosa and further to mucosa squamous cell carcinoma.The difference were statistically significant(χ2=16.676,p=0.002,P<0.05).In the process of tongue carcinogenesis,there was a negative correlation between the change trend of E-cad and the positive expression rate of CD44S.The correlation was statistical significance(rs=-1,p=0.000,P<0.05).There was a negative correlation between the positive rate of E-cad and PCNA.The correlation was statistical significance(rs=-1,P=0.000,P<0.05).There was no correlation between the positive rate of CD44S and PCNA(rs=-0.197,P=0.586,P>0.05).Simply,in the stage of Tongue squamous cell carcinoma,there was no significant correlation between E-cad and CD44S expression(rp=0.383,P=0.143,P>0.05).Conclusions:1.4NQO is still a kind of ideal inducer in the establishment of rat OSCC model even in barrier environment.Rat OSCC mod el can be successfully established.2.In the process of inducing the formation of tongue squamous cell carcinoma on SD rat under barrier environment of 4NQO,the positive expression rate of E-cad decreased gradually from the normal mucosa stage todysplasia stage to squamous cell carcinoma stage.Further discovered that expressions of cell membrane are developed from simple cell membrane to the cell membrane cytoplasm together.The positive expression rate of CD44S was gradually increased and so was PCNA.The positive expression rate of CD44S was gradually increased.The positive expression rates of PCNA also showes an upward trend.The expression rates of E-cad and CD44S in the process of tongue carcinogenesis were in negative correlation relationship.The expression rates of E-cad and PCNA in the process of tongue carcinogenesis are in negative correlation relationship.