| Objective:The experiment aims to analyse the difference of mi RNA expression spectrum in the peripheral blood between the patients with the oral and maxillofacial squamous cell carcinoma and the normal people,screening the differences for serum mi RNA in the oral and maxillofacial squamous cell carcinoma, and verifying optionally mi RNA,looking pecific expression of serum mi RNA for the OSCC.To provide the basis and theoretical foundation for early diagnosis,clinical treatment and prognosis of the disease.Methods:1 According to the standard operating procedures of peripheral blood collection,collecting the 3 blood samples from the patients with the OSCC and normal people.Among them,patients in the experimental group did not receive any special treatment before and all had histopathological diagnosis basis;besides,eliminating the tumour from other physical systems or tumor metastasis to the oral cavity.Needing fasting peripheral venous blood 2m L in the morning,with anticoagulant treatment and preserving at-80℃.Blood samples could not be placed more than 1h at room temperature.2 Using RNA extraction kit to extract total RNA of the samples until diluting the samples to make each RNA concentration equal.3 Measuring concentration and purity of the RNA.4 Diluting the samples to make each RNA concentration equal, processing the RNA reverserse transcription according to the instruction.5 Using Human Serum Plasma Mirna Pcr Array kit(96-well plates),to process q RT-PCR reaction and screen the difference of mi RNA.6 Selecting hsa-mi R-222-3p、hsa-mi R-423-5p and expanding the sample quantity to 30 cases of each group;Extracting total RNA,and verifying concentration and purity of the RNA.7 Diluting the samples to make each RNA concentration equal, processing q RT-PCR reaction.8 Result determination and statistical analysis.2-△△C T represents for the expression difference of mi RNA,to analyse each gene CT value,then,using each of its target gene CT value to minus its internal reference gene CT value,which is named△CT,the formula is △CT=CT(target gene)-CT(internal reference gene);Among them,C.39 is as for internal reference.Next,using the CT of the experimental group to minus the CT of the control group,the formula is △△CT=△CT(experimental group)-△CT(control group).Fold change value is 2-△△C T,calculating the average.All of the above statistical methods were performed by Excel Software.Fold change value >2,which indicates the expression level of mi RNA in the experimental group is up-regulation and significant,compared with the control group.Fold change value <0.5,which indicates the expression level of mi RNA in the experimental group is down-regulation and significant,compared with the control group.Using relative quantitative method to detect hsa-mi R-222-3p, hsa-mi R-423-5p in the specific expression of the OSCC.Level difference for mi RNA expression is RQ = 2-△△C T.For the peripheral blood in the OSCC and normal group,level difference for two kinds of mi RNA expression is analyzed by mean±standard deviation,T test which is appropriate for totally two independent and random samples,P < 0.05 indicates difference is statistically significant.All of the above statistical methods are analysed by SPSS13.0 statistical software.Results:1 It was found that 8 serum mi RNAs in OSCC patients were two fold higher than that in the normal people’s(Fold Change>2).2 It was found that 18 serum mi RNAs in OSCC patients were half fold lower than that in the normal people’s(Fold Change<0.5).3 In 30 cases of the experimental group,the expression average of hsa-mi R-222-3p is 5.587±0.218,while the average of 30 cases is 0.972±0.177 in the control group.According to the T test,for two completely random independent samples,hsa-mi R-222-3p is significant in the peripheral blood of oral and maxillofacial squamous cell carcinoma and normal group(P<0.05).That is,hsa-mi R-222-3p expression of the OSCC would increase significantly compared with the normal group(P<0.05).4 In 30 cases of the experimental group,the expression average of hsa-mi R-423-5p is 0.431±0.066,while the average of 30 cases is 1.206±0.195 in the control group.According to the T test,for two completely random independent samples,hsa-mi R-423-5p is significant in the peripheral blood of the OSCC and the normal group(P<0.05).That is,hsa-mi R-423-5p expression of the OSCC would reduce significantly compared with the normal group(P<0.05).Conclusions:1 There are the differences of mi RNA expression spectrum in the peripheral blood between the OSCC and the normal.So,there may be a relationship between the expression of serum mi RNA and the OSCC.2 Hsa-mi R-222-3p expression of the OSCC would increase significantly compared with the normal group,and hsa-mi R-423-5p expression of the OSCC would reduce significantly compared with the normal group.Hsa-mi R-222-3p,hsa-mi R-423-5p may become a biological index for the diagnosis of the OSCC. |
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