| [Objective]1.Collect the tissue and blood samples of lung adenocarcinoma patients in Xu anwei area,improve the relevant case information of patients,and establish a s pecimen bank.2.To study whether there is difference in the expression of mir-125b-5p in ca ncer tissue,normal tissue and blood of Xuanwei lung adenocarcinoma patients with and without lymph node metastasis,and to determine whether the differen ce is consistent.3.To analyze the difference of mir-125b-5p expression between cancer tissue a nd normal tissue of Xuanwei lung adenocarcinoma patients,blood samples of Xuanwei lung adenocarcinoma patients and healthy control group.4.To analyze whether the expression difference of mir-125b-5p in tissue and b lood samples can help to diagnose lung adenocarcinoma.5.To analyze the relationship between the expression of mir-125b-5p in tissues and blood samples and clinicopathological characteristics.6.Explore the most appropriate experimental blood components and the best m ethod of nucleic acid extraction[Method]1.Select the patients with composite inclusion criteria and collect their intraope rative tissue samples.The tissue samples were collected within 30 minutes afte r the operation.The normal tissue was more than 5 cm away from the cancer t issue,and the size of the tissue block was less than 1cm.The collection proce ss was strictly in accordance with the order of taking the normal tissue first a nd then the cancer tissue.The peripheral blood samples of the patients were c ollected 24 hours before the operation,and then put into anticoagulant tubeand non anticoagulant tube respectively,and then put into nucleic acid protection re agent tube.The healthy family members of the patients were selected as the bl ood samples of the healthy people,and all of them signed the informed conse nt.2.RT-qPCR was used to detect the cancer and normal tissues of Xuanwei lun g adenocarcinoma patients,and the expression of mir-125b-5p was analyzed.3.RT-qPCR was used to detect the expression of mir-125b-5p in the blood sa mples of Xuanwei lung adenocarcinoma patients and healthy people.4.ROC curve was used to analyze whether the expression difference of mir-125b-5p in tissue and blood samples could help the diagnosis of lung adenocarci noma.5.Spss21.0 software package was used to make statistics and analysis of the e xperimental results.The patient's age was displayed by mean±standard deviati on(x±s).The data of multiple groups were compared with each other and o ne-way was selected ANOVA analysis showed that Wilcoxon method was used to count the expression of mir-125b-5p between tissues,Mann Whitney U test was used to count the expression of mir-125b-5p in serum,blood cells and pl asma,and the data of RT-qPCR in tissues and blood samples were displayed b y quartile confidence interval.The expression level of mir-125b-5p was compar ed with the clinicopathological data such as age,gender,tumor size,TNM stag e and lymph node metastasis The connection of materials was measured by tw o-sided ?2test,with a statistical difference of P<0.05.[Result]1.41 specimens of lung adenocarcinoma and peripheral blood were collected,a nd the relevant case information was improved,and the specimen bank was est ablished.2.By comparing the expression of mir-125b-5p in the tissues and serum sampl es of patients with and without lymph node metastasis,we found that the expr ession of mir-125b-5p in the tissues of patients with lymph node metastasis wa s 0.6(0.063-0.789)higher than that in the tissues of patients without lymph n ode metastasis(0.0061(0.0002-0.026),P=0.11<0.05;the expression of mir-125b-5p in the tissues of patients with lymph node metastasis was 6.24(0.027-8.423)higher than that in the tissues of patients without lymph node metastasi s Tissue 0.072(0.0007-0.012),P=0.007<0.05;serum 0.606(0.242-0.466)in patients with lymph node metastasis was higher than serum-0.832(-1.352-0.798)in patients without lymph node metastasis,P=0.017<0.05.The expressio n of mir-125b-5p in patients with lymph node metastasis was higher than thati n patients without lymph node metastasis.The expression level of mir-125b-5p in normal tissues was 1.333(0.023-7.338)higher than that in cancer tissues(0.453(0.004-1.628)P<0.05).4.The expression of mir-125b-5p in patients' serum and healthy control sample s was significantly different.The expression of mir-125b-5p in patients' group was 0.222(-0.446-0.391)significantly higher than that in healthy group(-0.325(-0.879-0.227)P<0.05,and the difference was statistically significant.5.ROC curve to explore mir-125b-5p as a diagnostic indicator of lung adenoc arcinoma.In serum,the relative expression sensitivity of mir-125b-5p is 0.687,t he specificity is 0.818,the area under the curve(AUC)is 0.753;in tissue,the expression sensitivity of mir-125b-5p is 0.517,the specificity is 0.759,and the AUC is 0.638,respectively,which is significant for the diagnosis of lung aden ocarcinoma.6.There was no significant correlation between the expression of mir-125b-5p a ndpatients' age,gender,smoking history,,tumor size and stage(P>0.05).7.Analysis by ultraviolet spectrophotometer:Serum extracted by Biog method(concentration:45.9±20.4;OD value 260/280:1.9±0.1);Od260/230:1.9±0.1 is better than plasma(concentration:35.8±12.9;od260/280:1.9±0.1;od260/230:1.9±0.1)and blood cells(concentration:455.0±192.3;od260/280:1.7±0.1;od260/230:0.8±0.2)and can be applied to clinical practice.[Conclusion]1.The expression of mir-125b-5p in cancer tissue,normal tissue and seru m of lymph node metastasis group was higher than that of non lymph node m etastasis group,which proved that there was a correlation between lymph node metastasis and the expression of mir-125b-5p.Further understanding of the me chanism of lymph node metastasis and mir-125b-5p expression is the key point of studying lung adenocarcinoma in Xuanwei area2.The expression of mir-125b-5p was significantly different between cance r and normal tissues,blood samples and healthy controls.3.The sensitivity and specificity of mir-125b-5p and the area under the c urve were analyzed by ROC curve.It was proved that mir-125b-5p could be u sed as an evaluation index to assist the diagnosis of lung adenocarcinoma.4.There was no significant correlation between the expression of mir-125b-5p in tissue and blood samples and the age,gender,smoking history of patien ts.5.In blood samples,the serum extracted by Biog method is better than ot her methods in terms of concentration and purity(OD value),but there is still a certain gap between the results and the RNA extracted from tissues.6.The method of extraction and preservation of samples in this project ha s good stability of nucleic acid,standardized,rapid and orderly extraction of sa mples,avoided the pollution of RNA enzyme,and guaranteed the follow-up ex periment.The cryopreservation of liquid nitrogen can reduce the cost of sample collection,and the collection method of the subject sample can achieve long-t erm and stable preservation,which is suitable for subsequent experiments. |
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