The Expression And Clinical Significance Of MicroRNA-150 In Intrahepatic Cholangiocarcinoma Patients

Intrahepatic cholangiocarcinoma(ICC) is the second most common type of primary hepatic malignancy second to hepatocellular carcinoma(HCC) among all liver malignancies. Although ICC accounts for 10~15% of liver cancers, its incidence and mortality has increased drastically over the past several decades in C hina and even worldwide. ICC has become one of the major malignant tumor which threat to human life and health. In most cases, there is no particular clinical symptom for the early onset in ICC, and no specific or practical laboratory markers for early diagnosis. And rapid progress of the condition caused the majority of patients be firstly diagnosed at advanced stage. Thus, they lost the best opportunity to accept radical surgery, and the prognosis is poor.Conversely, the ICC patients who received diagnosis and treatment at early stage, could always benefit with a longer survival time. To explore effective methods of the early clinical diagnosis and treatment of ICC is crucial for patient’ survival and prognosis. To date, limited serum markers for ICC have been reported to be useful in diagnostic procedures, but the sensitivity and specificity of these markers remain far lower than expected. Therefore, the identification of new tumor biomarker with high sensitivity and specificity for ICC is the urgent problem need to solve.Micro RNAs are small non-coding RNA molecules, which degrade or negatively regulate the translation of target m RNA according to the degree of base pairing to complementary sites within its 3’-untranslated region(3’UTR). mi RNAs are widely involved in the growth and development of the human body. Dysregulation of mi RN As is closely related to a variety of disease including tumor. Additionally, mi RN As could play a role as tumor suppressor gene or oncogene in the formation of malignancies, regulating the occurrence and development of tumor. There are a lot of researches suggest that mi RN A expression profile has obvious differences in different type of tumors, and can be steady monitored, which suggest micro RNA could be applied as a new biomarker to the diagnosis of neoplasms.In this study, in order to seek a mi RNA as a high-efficient tumor biomarker, micro RNA microarray、RT-q PCR and statistical analysis were used. The study include the following four parts.Part I: collecting、sorting medical records and s pecimens and selecting enrolled patientsMethod: Software Epidata(version 3.1) was used to improve the database of prime liver cancer partents. According to the the inclusion criteria, the clinical information of intrahepatic cholangiocarcinoma patients and controls was collected. The comparability between the two groups analysis was performed using the software SPSS 21.0.Result: Compared with the group of normal controls, the average serum albumin(ALB) value was lower than in the group of intrahepatic cholangiocarcinoma patients, with an average value of 40.9 g/L ± 4.30 g/L vs. 44.9 g/L ± 3.84 g/L(P =0.010). However, there were no markedly differences in other clinicopathological features between the two groups.Conclusion: Albumin, synthesized in the liver, is the important index for evaluation of nutritional status. C hronic wasting diseases such as malignant tumor, serum albumin will be reduced. Other clinical information with the exception of serum albumin value between the two groups are similar. So, the group of intrahepatic cholangiocarcinoma patients and the group of controls are entirely comparable.Part II: setting up the mi RNA expression profile of ICC, selecting mi RNA-150 as a target geneMethod: mi RN A microarray was used to screen mi RN A expression profile of ICC. Then, micro RNA-150 was selected as target gene in this study after reviewing literatures and analyzing the microarray data.Result: We found that 10 mi RNAs were dysregulated in the ICC cancerous tissues. The most outstanding under-expression values were found for mi R-150、mi R-638、mi R-378c、mi R-4530 and mi R-4459. In addition, mi RN A-93、mi RNA-425、mi RN A-660、mi R-494 and mi RNA-574 were found over expressed in the study. Then, mi RNA-150 was selected as the target gene in further study among the 10 mi RNAs.Conclusion: There are a little difference in the micro RN A expression profile between our study and previous studies. The main reason for this phenomenon may be the heterogeneities between different specimens. It also shows that the development of tumor may be regulated by diversity genes. The expression level of mi RNA-150 is abnormal in a lot of tumors. However, there are few clinical research about the relationship between mi RN A-150 and ICC. Therefore, it is feasible to reasch the mi RN A-150 as a target gene in the future.Part III: To detect the expression level of mi RNA-150 in tissue and plasma specimens, RT-q PCR was carried out.Method: mi RNeasy® Mini kit and RN24-Bl OODmisi were used to extracte total RNAs which included mi RN As from tissues and plasma samples, respectively. Then, Nano Drop? lite spectrophotometer was used to detecte the purity of the RNA solution. Also, the expression level of mi RN A-150 in tissue and plasma specimens was detected using RT-q PCR.Result: The expression level mi RNA-150 was lower in cancerous tissue in comparison with peritumoral noncancerous tissue,while it was higher in plasma of ICC patient in comparison with plasma of controls.Conclusion: We found contrary expression profiles of mi R-150 between the tumor tissues and blood samples. In other tumors, the expression level of mi RNA-150 are high or/and low, we think that mi RN A-150 has different expression patterns in different tumors and its abnormal expression might be a common phenomenon in malignancies. For the contrary expression profiles of mi R-150 between the tumor tissues and blood samples, we think that mi RNA-150 may play the role of tumor suppressor gene and adjust the expression level of mi RN A-150 in the blood through negative feedback in patients with ICC.Part IV: relevance between plasma mi R-150 expression level and clinical features for ICC patients.Method: The relevance between plasma mi R-150 expression level and clinical features were analyzed using software SPSS 21.0 according to clinicopathological features of 15 ICC patients and the expression level of plasma mi RNA-150. A value of P<0.05 was considered to indicate a statistically significant difference.Result: The expression level of mi RNA-150 in plasma is a diagnostic biomarker for ICC. When combined mi RNA-150 with CA19-9, the sensitivity、specificity and AUC went up to 80% 、100% and 0.920, respectively. On the base of the mean expression level of mi R-150, 15 ICC patients were divided into 2 groups: low expression group(n=8), containing the ones with a plasma mi R-150 level less than the mean value of-0.507, and a high expression group(n=7) with a plasma mi R-150 level higher than the mean value of-0.507. No significant differences were noted between the 2 groups.Conclusion: When diagnosing ICC, the sensitivity of plasma mi RN A-150 was 93.3%, and the specificity was 53.3%. However, while combined it with serum CA19-9, the indexes of sensitivity、specificity and AUC were improved observably. In addition, there were no significant differences between the low expression group and high expression group. This observation suggests that the mi R-150 expression level was not affected by albumin(ALB), total bilirubin(TB), alanine(ALT), aspartate aminotransferase(AST) and other clinical indices. Therefore, mi R-150 exists stably in blood samples from ICC patients. It might be a specificity biomarker for ICC as expected.