MicroRNA-124 alone might represent an indicator signifying cholangiocarcinoma prognosis

MicroRNA-124 alone might represent an indicator signifying cholangiocarcinoma prognosis | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Primary research MicroRNA-124 alone might represent an indicator signifying cholangiocarcinoma prognosis Ning Wang, Yanni Li, Yanfang Zheng, Huoming Chen, Xiaolong Wen, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-33111/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background: Previous studies have demonstrated that microRNAs (miRNAs) played a crucial role in various diseases, including cancers. The aim of the study was to evaluate the clinical significance of miR-124 in patients with cholangiocarcinoma (CCA). Methods: The expression pattern of miR-124 was detected in CCA tissues using quantitative reserve transcription polymerase chain reaction (qRT-PCR). The correlation of miR-124 expression with clinicopathological features and overall survival of patients were explored using chi-square test, Kaplan-Meier methods and Cox regression analyses. Results: The miR-124 expression level was strong down-regulated in CCA tissues compared with normal para-cancerous tissues ( P <0.001). Moreover, aberrant miR-124 expression was significantly associated with differentiation ( P =0.045) and lymph node metastasis ( P =0.040). In addition, Kaplan-Meier method and log-rank test revealed that patients with low miR-124 expression has a poorer overall survival compared with those with high miR-124 expression ( P =0.002). Furthermore, multivariate analysis confirmed that miR-124 expression ( P =0.006; HR=2.006; 95%CI: 1.224-3.289) was an independent prognostic indicator in CCA. Conclusions: Collectively, our results defined miR-124 expression plays important roles in CCA patients. MiR-124 expression might used as a valuable prognostic biomarker for patients with CCA. Cancer Biology MicroRNA-124 Cholangiocarcinoma Survival Prognosis Figures Figure 1 Figure 2 Background Cholangiocarcinoma (CCA) is a devastating malignancy derived from bile duct epithelial cells, and the mortality associated with CCA is currently increasing worldwide [ 1 ]. According to the statistics, the incidence of CCA changes widely among different geographic regions; the highest number of cases occurs in southeast Asia while the lowest number occurs in Australia [ 2 ]. Anatomically, CCA is categorized as intrahepatic cholangiocarcinomas (IHCCs) and extrahepatic cholangiocarcinoma (EHCCs), the latter being further divided into perihilar CCAs and distal CCAs [ 3 ]. CCA is difficult to diagnosis in early-stage due to its lack of early symptoms [ 4 ], and it is featured by high frequency of recurrence and metastasis. What’s more, there is no effective chemoprevention or treatment, leading to an unfavorable prognosis [ 5 ]. MicroRNAs (miRNAs) are a type of endogenous coding small molecular RNAs with 18–25 nucleotides in length that could negatively control the translational inhibition of target mRNAs through base-pairing with their 3’-untranslated region (3’-UTR) [ 6 , 7 ]. MicroRNAs are known to play an important role in diverse biological processes, such as apoptosis, cell proliferation, and differentiation [ 8 ], which displays their functionality in carcinogenesis as tumor suppressor genes or oncogenes [ 9 – 11 ]. miR-124 is one of the cancer-related miRNAs. It is a brain-enriched miRNA and was first discovered to be involved in stem cell regulation and neuro-development [ 12 , 13 ]. Previous studies indicated that miR-124 is down-regulated in various cancers. For example, in colorectal cancer, miR-124 is strongly decreased, and promotes apoptosis of colorectal cells [ 14 ]. In addition, Li et al. revealed that miR-124 is reduced and negatively related to lymph node metastasis in breast cancer [ 15 ]. The above studies indicated miR-124 play a crucial role in tumorigenesis and progression. However, the clinical significance of miR-124 remains unknown in CCA. In the present study, we measured the relative expression level of miR-124 in CCA tissues via qRT-PCR, and analyzed the relationship of miR-124 level with clinicopathological features. We also explored the prognostic performance of miR-124 in CCA. Methods Patients and tissues samples The use of tissues for this study has been approved by the Ethics Committee of the PLA Rocket Force Characteristic Medical Center. At the time of initial diagnosis, all patients had provided consent in the sense that their tumor samples could be used for investigation purposes. Written informed consents were received from all participants involved in the study. In total, 119 frozen cancerous samples from CCA patients undergoing surgery from at our department were collected in this study. In addition, 119 matched para-cancerous samples were collected. All the specimens were diagnosed by two pathologists separately. None of the patients recruited in this study had undergone preoperative chemotherapy or radiotherapy. The clinical features of the patients, including age, gender, tumor size, location, distant metastasis, differentiation, TNM stage, and lymph node metastasis were listed in Table 1 . The overall survival time was computed as the time from the date of first pathologic diagnosis to the date of death or last follow-up. RNA extraction and quantitative reserve transcription polymerase chain reaction (qRT-PCR) Total RNA from fresh tissues was isolated using the mirVana miRNA Isolation kit (Ambion, Austin, TX, USA) according to the manufacturer’s instructions. Single-stranded cDNA for microRNA analysis was also synthesized by RT using the TaqMan MicroRNA Reverse Transcription Kit (Applied Biosystems, Foster City, CA, USA) according to the manufacturer’s instructions. qRT-PCR method was used to assess the expression levels of miR-124 with TaqMan miRNA assay kit (Applied Biosystems, Foster, CA, USA) on a 7500 Fast Real-time PCR System (Applied Biosystems, California, USA). U6 RNA was used as an endogenous reference for normalizing the expression levels of miR-124. The threshold cycle (Ct) was defined as the fractional cycle number at which the fluorescence passed the fixed threshold. Each sample was measured in triplicate, and the relative amount of miR-124 to U6 was calculated using the 2 -△△CT method. Statistical analysis All statistical analyses were preformed with the SPSS 18.0 software (SPSS Inc., Chicago, IL, USA) and graphs were generated via GraphPad Prism 5.0 (GraphPad Software Inc., USA). Data are shown as the mean ± standard deviation (SD). The Independent Student t test was used to compare the differences between groups. The relationship of miR-124 expression with clinicopathological factors was analyzed using Chi-square test. Patients survival and their differences were determined by the Kaplan-Meier method and log-rank test. A Cox’s regression model was used for univariate and multivariate analysis. P <0.05 was considered to indicated a statistically significant result. Results Aberrant expression of miR-124 in CCA tissues Firstly, we compared the miR-124 expression in 119 CCA tissues and para-cancerous tissues with qRT-PCR assay. As indicated in Figure 1 , the miR-124 level was significantly lower in CCA tissues than para-cancerous samples ( P <0.001), suggesting that expression of miR-124 was down-regulated in CCA tissues. Association of miR-124 expression with clinicopathological characteristics in CCA The correlation between miR-124 expression and clinicopathological parameters was examined in 119 cases ( Table 2 ). Based on the median value (1.48) of miR-124 expression, all patients were divided into two groups as followings: low miR-124 expression group (n=63) and high miR-124 expression group (n=56). Among the parameters listed, a significant correlation was observed between low miR-124 expression and differentiation ( P =0.045), lymph node metastasis ( P =0.040). However, there were no relationships between miR-124 expression and other clinicopathological factors, including age, gender, tumor size, location, distant metastasis and TNM stage (all P >0.05)( Figure 2 ). Relationship between miR-124 expression and overall survival Next, we analyzed the correlation of miR-124 expression with patients clinical outcome. Kaplan-Meier analysis showed that miR-124 expression was significantly associated with poor overall survival in CCA patients. According to log-rank test, the overall survival rate of patients with low miR-124 expression was lower than that of patients with high miR-124 expression ( P =0.001). In univariate Cox regression analysis, miR-124 expression ( P =0.003, HR=2.107, 95%CI: 1.287-3.448) was a prognostic factor for cancer mortality in CCA. Additionally, lymph node metastasis ( P =0.001, HR=2.269, 95%CI: 1.416-3.638) was also significantly related to overall survival. In a multivariate analysis, miR-124 expression was an unfavorable independent prognostic indicator in patients with CCA ( P =0.006, HR=2.006, 95%CI: 1.224-3.289). Discussion CCA is the most common primary hepatic malignancy worldwide. Despite tremendous improvements in developing multimodal treatments, the clinical outcome of CCA patients remains unfavorable. Thus, it is of great significance to identify powerful prognostic indicator for CCA. The discovery that non-coding components of the genome, including miRNAs, can conduct to the pathogenesis of cancer has led researchers to contemplate using these molecules to guide clinical strategies custom [16]. So far, a large number of studies suggest that miRNAs play a key role in tumor development. Increasing evidence suggest that deregulated expression of miRNAs participates in the initiation of CCA progression, where they act as either suppressors or promoters at the post-transcriptional regulation stage. For instance, Yang et al. reported that miR-144 was significantly down-regulated in CCA tissues and might be a tumor suppressor of CCA cell proliferation and invasion through targeting LIS1 [17]. Zhang et al. showed that miR-26a was increased in human CCA tissues and cell lines and overexpression of miR-26a increased proliferation of CCA cells and colony formation in vitro [18]. Wang et al. revealed that miR-138 was underexpressed in CCA tissues, and downregulation of miR-138 promoted the proliferation, migration and invasion of CCA cells [19]. Liu et al. confirmed that over-expression of miR-21 significantly promoted cell migration, invasion, and xenografts growth [20]. Previous research confirmed that miR-124 was dysregulated in various cancers, and aberrant miR-124 expression was involved in the prognosis of tumors. For example, Dong et al. showed that miR-124 had lower expression in breast cancer specimens and low miR-124 expression was a significant independent predictor of poor survival in breast cancer [21]. Gilje et al. showed that miR-124 is a potential new biomarker for prediction of neurological prognosis following cardiac arrest [22]. Han et al. demonstrated that miR-124 had a low expression in osteosarcoma tissues and low miR-124 expression was an unfavorable prognostic factor for overall survival [23]. Zhang et al. proved that miR-124 expression level was significantly decreased in lung cancer tissues and miR-124 was an independent prognostic factor for overall survival in lung cancer [24]. However, to our knowledge, the clinical significance of miR-124 in CCA has not been reported. Therefore, we investigated the feasibility of miR-124 as a new potential biomarker of prognosis for CCA. In the present study, our data showed that miR-124 expression was significantly decreased in CCA tissues in comparisons with normal para-cancerous tissues. The relationship between miR-124 expression and various clinicopathological characteristics of CCA patients was investigated. The results indicated that abnormal miR-124 expression was dramatically correlated with differentiation and lymph node metastasis. It demonstrated that miR-124 might be involved in tumor metastasis and progression of CCA. Furthermore, the overall survival rate of low miR-124 group was significantly shorter than that of high miR-124 expression group. Finally, in a multivariate Cox model, we found that miR-124 expression was an independent poor prognostic indicator for overall survival of CCA patients. Our results were in line with previous studies. For example, Tian et al. showed that miR-124 was reduced in CCA tissues and negatively related to lymph node involvement and distant metastasis [25]. Conclusion In conclusion, our results showed that miR-124 expression was remarkably reduced in CCA tissues. The expression of miR-124 was closely associated with the progression of CCA patients. More importantly, the down-regulation of miR-124 is an independent poor indicator of prognosis in CCA patients. We believed that it is just the beginning of a long and still unexplored avenue of a highly promising investigation of miR-124. A more in-depth and larger-scale study remains to identify the role of miR-124 in CCA. Abbreviations microRNAs (miRNAs) cholangiocarcinoma (CCA) quantitative reserve transcription polymerase chain reaction (qRT-PCR) intrahepatic cholangiocarcinomas (IHCCs) extrahepatic cholangiocarcinoma (EHCCs) 3’-untranslated region (3’-UTR) Declarations Ethics approval and consent to participate This study was supported by the Ethics Committee of the PLA Rocket Force Characteristic Medical Center and also has been carried out in accordance with the World Medical Association Declaration of Helsinki. Consent for publication The subjects had been informed the objective. Certainly, written consents were signed by every subject in this study. Data availability The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request. Funding Funding information is not applicable. Competing interests The authors declare that they have no competing interests. Authors’ contributions N.W. design of the work; Y.L. the acquisition, analysis, Y.Z. interpretation of data; H.C. the creation of new software used in the work; X.W., Z.L. have drafted the work or substantively revised it. All authors read and approved the final manuscript. Acknowledgements Not applicable. References Patel T: Increasing incidence and mortality of primary intrahepatic cholangiocarcinoma in the United States. Hepatology 2001, 33(6):1353-1357. Bergquist A, von Seth E: Epidemiology of cholangiocarcinoma. Best practice & research Clinical gastroenterology 2015, 29(2):221-232. Gatto M, Bragazzi MC, Semeraro R, Napoli C, Gentile R, Torrice A, Gaudio E, Alvaro D: Cholangiocarcinoma: update and future perspectives. Digestive and liver disease : official journal of the Italian Society of Gastroenterology and the Italian Association for the Study of the Liver 2010, 42(4):253-260. Blechacz B, Gores GJ: Cholangiocarcinoma: advances in pathogenesis, diagnosis, and treatment. Hepatology 2008, 48(1):308-321. Patel T: Cholangiocarcinoma--controversies and challenges. Nature reviews Gastroenterology & hepatology 2011, 8(4):189-200. He L, Hannon GJ: MicroRNAs: small RNAs with a big role in gene regulation. Nature reviews Genetics 2004, 5(7):522-531. Bartel DP: MicroRNAs: target recognition and regulatory functions. Cell 2009, 136(2):215-233. Bartel DP: MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 2004, 116(2):281-297. Hezova R, Kovarikova A, Srovnal J, Zemanova M, Harustiak T, Ehrmann J, Hajduch M, Sachlova M, Svoboda M, Slaby O: MiR-205 functions as a tumor suppressor in adenocarcinoma and an oncogene in squamous cell carcinoma of esophagus. Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine 2016, 37(6):8007-8018. Shenouda SK, Alahari SK: MicroRNA function in cancer: oncogene or a tumor suppressor? Cancer metastasis reviews 2009, 28(3-4):369-378. Wang W, Luo YP: MicroRNAs in breast cancer: oncogene and tumor suppressors with clinical potential. Journal of Zhejiang University Science B 2015, 16(1):18-31. Lee MR, Kim JS, Kim KS: miR-124a is important for migratory cell fate transition during gastrulation of human embryonic stem cells. Stem Cells 2010, 28(9):1550-1559. Cheng LC, Pastrana E, Tavazoie M, Doetsch F: miR-124 regulates adult neurogenesis in the subventricular zone stem cell niche. Nature neuroscience 2009, 12(4):399-408. Zhang J, Lu Y, Yue X, Li H, Luo X, Wang Y, Wang K, Wan J: MiR-124 suppresses growth of human colorectal cancer by inhibiting STAT3. PloS one 2013, 8(8):e70300. Li L, Luo J, Wang B, Wang D, Xie X, Yuan L, Guo J, Xi S, Gao J, Lin X et al : Microrna-124 targets flotillin-1 to regulate proliferation and migration in breast cancer. Molecular cancer 2013, 12:163. Nana-Sinkam SP, Croce CM: Clinical applications for microRNAs in cancer. Clinical pharmacology and therapeutics 2013, 93(1):98-104. Yang R, Chen Y, Tang C, Li H, Wang B, Yan Q, Hu J, Zou S: MicroRNA-144 suppresses cholangiocarcinoma cell proliferation and invasion through targeting platelet activating factor acetylhydrolase isoform 1b. BMC cancer 2014, 14:917. Zhang J, Han C, Wu T: MicroRNA-26a promotes cholangiocarcinoma growth by activating beta-catenin. Gastroenterology 2012, 143(1):246-256 e248. Wang Q, Tang H, Yin S, Dong C: Downregulation of microRNA-138 enhances the proliferation, migration and invasion of cholangiocarcinoma cells through the upregulation of RhoC/p-ERK/MMP-2/MMP-9. Oncology reports 2013, 29(5):2046-2052. Liu Z, Jin ZY, Liu CH, Xie F, Lin XS, Huang Q: MicroRNA-21 regulates biological behavior by inducing EMT in human cholangiocarcinoma. International journal of clinical and experimental pathology 2015, 8(5):4684-4694. Dong LL, Chen LM, Wang WM, Zhang LM: Decreased expression of microRNA-124 is an independent unfavorable prognostic factor for patients with breast cancer. Diagnostic pathology 2015, 10:45. Gilje P, Gidlof O, Rundgren M, Cronberg T, Al-Mashat M, Olde B, Friberg H, Erlinge D: The brain-enriched microRNA miR-124 in plasma predicts neurological outcome after cardiac arrest. Crit Care 2014, 18(2):R40. Han G, Wang Y, Bi W, Jia J, Wang W: MicroRNA-124 functions as a tumor suppressor and indicates prognosis in human osteosarcoma. Experimental and therapeutic medicine 2015, 9(3):679-684. Zhang Y, Li H, Han J: Down-regulation of microRNA-124 is correlated with tumor metastasis and poor prognosis in patients with lung cancer. International journal of clinical and experimental pathology 2015, 8(2):1967-1972. Tian F, Chen J, Zheng S, Li D, Zhao X, Jiang P, Li J, Wang S: miR-124 targets GATA6 to suppress cholangiocarcinoma cell invasion and metastasis. BMC cancer 2017, 17(1):175. Tables Table 1. The relationship between miR-124 expression and clinicopathological features Features Cases (n=119) MiR-124 expression χ 2 P -value Low (n=63) High (n=56) Age (years) <68 58 26 32 2.990 0.084 ≥68 61 37 24 Gender Male 73 43 30 2.695 0.101 Female 46 20 26 Tumor size (cm) <5 63 33 32 0.271 0.603 ≥5 54 30 24 Location Intrahepatic 24 14 10 0.351 0.554 Extrahepatic 95 49 46 Distant metastasis Negative 88 42 46 3.686 0.055 Positive 31 21 10 Differentiation Well-moderate 76 35 41 4.006 0.045 Poor 43 28 15 TNM stage Ⅰ-Ⅱ 61 28 33 2.489 0.115 Ⅲ-Ⅳ 58 35 23 Lymph node metastasis Absent 69 31 38 4.233 0.040 Present 50 32 18 Table 2. The univariate and multivariate analyses of clinical factors for overall survival of CCA Factors Univariate analysis Multivariate analysis HR (95%CI) P -value HR (95%CI) P -value MiR-124 expression 2.107 (1.287-3.448) 0.003 2.006 (1.224-3.289) 0.006 Age 0.901 (0.565-1.438) 0.663 - - Gender 1.504 (0.924-2.448) 0.100 - - Tumor size (cm) 1.353 (0.848-2.160) 0.204 - - Location 0.814 (0.466-1.422) 0.470 - - Distant metastasis 1.397 (0.839-2.327) 0.199 - - Differentiation 1.153 (0.711-1.870) 0.563 - - TNM stage 1.446 (0.905-2.310) 0.122 - - Lymph node metastasis 2.269 (1.416-3.638) 0.001 2.175 (1.354-3.492) 0.001 Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-33111","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Primary research","associatedPublications":[],"authors":[{"id":641679,"identity":"cd75db7e-cf9f-495f-b291-6428f147bf67","order_by":0,"name":"Ning Wang","email":"","orcid":"","institution":"the PLA Rocket Force Characteristic Medical Center","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Ning","middleName":"","lastName":"Wang","suffix":""},{"id":641680,"identity":"31cdb7cf-14c9-4c62-88d0-f47c4c9c5cdb","order_by":1,"name":"Yanni Li","email":"","orcid":"","institution":"the PLA Rocket Force Characteristic Medical Center","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Yanni","middleName":"","lastName":"Li","suffix":""},{"id":641681,"identity":"6e907660-7112-4cf3-a182-02e11d51d672","order_by":2,"name":"Yanfang Zheng","email":"","orcid":"","institution":"the PLA Rocket Force Characteristic Medical Center","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Yanfang","middleName":"","lastName":"Zheng","suffix":""},{"id":641682,"identity":"326db660-1f2b-4cf1-87e3-60e5ea42acb4","order_by":3,"name":"Huoming Chen","email":"","orcid":"","institution":"the PLA Rocket Force Characteristic Medical Center","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Huoming","middleName":"","lastName":"Chen","suffix":""},{"id":641683,"identity":"32144532-8769-4250-a84e-63ed19603dd7","order_by":4,"name":"Xiaolong Wen","email":"","orcid":"","institution":"University of Science and Technology Beijing","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Xiaolong","middleName":"","lastName":"Wen","suffix":""},{"id":641684,"identity":"70420787-b218-4786-b87a-4e2b45b2f0d7","order_by":5,"name":"Zhaoxia Li","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAuUlEQVRIiWNgGAWjYDACCQaGD0BKjo29+QDRWhhnACljPp5jCaRpSZwnkaNAnA752b0HGz7uqE1vY8hhYPhRsY2wFoM75xIbZ545ntvGcPYAY8+Z20Rokcgxf8zbdiy3jbEvgZmxjQgt8jNyDJuBWtLZmHkMiNPCcAOspSaBjY1YLQZALY0z2w4YtvGwJRwkyi8ghzV8bKuTl5//+OCDHxXEOAwCDoPJA0SrB4I6UhSPglEwCkbBSAMAJV89E5G+fGkAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0001-6993-5507","institution":"the PLA Rocket Force Characteristic Medical Center","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Zhaoxia","middleName":"","lastName":"Li","suffix":""}],"badges":[],"createdAt":"2020-06-02 04:19:28","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-33111/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-33111/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":1665348,"identity":"016e337a-f0ba-46d0-ad32-e239d61c59c6","added_by":"auto","created_at":"2020-07-23 19:51:27","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":796556,"visible":true,"origin":"","legend":"The relative expression of miR-124 was examined in CCA tissues via qRT-PCR assay. Results showed that miR-124 level was significantly lower in CCA tissues compared with normal para-carcinoma tissues (P\u003c0.001).","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-33111/v1/floatimage2.png"},{"id":1665349,"identity":"f5b8d7e9-14fb-4b0f-ba1f-2861d37d1bd6","added_by":"auto","created_at":"2020-07-23 19:51:27","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":727694,"visible":true,"origin":"","legend":"Kaplan-meier analysis of the impact of miR-124 expression on overall survival. Patients with low miR-124 expression had a shorter overall survival rate than those with high miR-124 expression (P=0.002).","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-33111/v1/floatimage1.png"},{"id":13538013,"identity":"8271b472-e639-4f30-bb72-4123d4326b54","added_by":"auto","created_at":"2021-09-17 01:37:53","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":674414,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-33111/v1/bd862a0d-f86a-4788-9f3a-721c3c415668.pdf"}],"financialInterests":"","formattedTitle":"MicroRNA-124 alone might represent an indicator signifying cholangiocarcinoma prognosis","fulltext":[{"header":"Background","content":" \u003cp\u003eCholangiocarcinoma (CCA) is a devastating malignancy derived from bile duct epithelial cells, and the mortality associated with CCA is currently increasing worldwide [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. According to the statistics, the incidence of CCA changes widely among different geographic regions; the highest number of cases occurs in southeast Asia while the lowest number occurs in Australia [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Anatomically, CCA is categorized as intrahepatic cholangiocarcinomas (IHCCs) and extrahepatic cholangiocarcinoma (EHCCs), the latter being further divided into perihilar CCAs and distal CCAs [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. CCA is difficult to diagnosis in early-stage due to its lack of early symptoms [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e], and it is featured by high frequency of recurrence and metastasis. What\u0026rsquo;s more, there is no effective chemoprevention or treatment, leading to an unfavorable prognosis [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eMicroRNAs (miRNAs) are a type of endogenous coding small molecular RNAs with 18\u0026ndash;25 nucleotides in length that could negatively control the translational inhibition of target mRNAs through base-pairing with their 3\u0026rsquo;-untranslated region (3\u0026rsquo;-UTR) [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. MicroRNAs are known to play an important role in diverse biological processes, such as apoptosis, cell proliferation, and differentiation [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e], which displays their functionality in carcinogenesis as tumor suppressor genes or oncogenes [\u003cspan additionalcitationids=\"CR10\" citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e].\u003c/p\u003e \u003cp\u003emiR-124 is one of the cancer-related miRNAs. It is a brain-enriched miRNA and was first discovered to be involved in stem cell regulation and neuro-development [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Previous studies indicated that miR-124 is down-regulated in various cancers. For example, in colorectal cancer, miR-124 is strongly decreased, and promotes apoptosis of colorectal cells [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. In addition, Li \u003cem\u003eet al.\u003c/em\u003e revealed that miR-124 is reduced and negatively related to lymph node metastasis in breast cancer [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. The above studies indicated miR-124 play a crucial role in tumorigenesis and progression. However, the clinical significance of miR-124 remains unknown in CCA.\u003c/p\u003e \u003cp\u003eIn the present study, we measured the relative expression level of miR-124 in CCA tissues via qRT-PCR, and analyzed the relationship of miR-124 level with clinicopathological features. We also explored the prognostic performance of miR-124 in CCA.\u003c/p\u003e "},{"header":"Methods","content":"\u003cp\u003e\u003cem\u003ePatients and tissues samples\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe use of tissues for this study has been approved by the Ethics Committee of the PLA Rocket Force Characteristic Medical Center. At the time of initial diagnosis, all patients had provided consent in the sense that their tumor samples could be used for investigation purposes. Written informed consents were received from all participants involved in the study.\u003c/p\u003e\n\u003cp\u003eIn total, 119 frozen cancerous samples from CCA patients undergoing surgery from at our department were collected in this study. In addition, 119 matched para-cancerous samples were collected. All the specimens were diagnosed by two pathologists separately. None of the patients recruited in this study had undergone preoperative chemotherapy or radiotherapy. The clinical features of the patients, including age, gender, tumor size, location, distant metastasis, differentiation, TNM stage, and lymph node metastasis were listed in \u003cstrong\u003eTable 1\u003c/strong\u003e. The overall survival time was computed as the time from the date of first pathologic diagnosis to the date of death or last follow-up.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eRNA extraction and quantitative reserve transcription polymerase chain reaction (qRT-PCR)\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eTotal RNA from fresh tissues was isolated using the mirVana miRNA Isolation kit (Ambion, Austin, TX, USA) according to the manufacturer\u0026rsquo;s instructions. Single-stranded cDNA for microRNA analysis was also synthesized by RT using the TaqMan MicroRNA Reverse Transcription Kit (Applied Biosystems, Foster City, CA, USA) according to the manufacturer\u0026rsquo;s instructions. qRT-PCR method was used to assess the expression levels of miR-124 with\u0026nbsp; TaqMan miRNA assay kit (Applied Biosystems, Foster, CA, USA) on a 7500 Fast Real-time PCR System (Applied Biosystems, California, USA). U6 RNA was used as an endogenous reference for normalizing the expression levels of miR-124. The threshold cycle (Ct) was defined as the fractional cycle number at which the fluorescence passed the fixed threshold. Each sample was measured in triplicate, and the relative amount of miR-124 to U6 was calculated using the 2\u003csup\u003e-△△CT\u003c/sup\u003e method.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eStatistical analysis\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eAll statistical analyses were preformed with the SPSS 18.0 software (SPSS Inc., Chicago, IL, USA) and graphs were generated via GraphPad Prism 5.0 (GraphPad Software Inc., USA). Data are shown as the mean \u0026plusmn; standard deviation (SD). The Independent Student \u003cem\u003et\u003c/em\u003e test was used to compare the differences between groups. The relationship of miR-124 expression with clinicopathological factors was analyzed using Chi-square test. Patients survival and their differences were determined by the Kaplan-Meier method and log-rank test. A Cox\u0026rsquo;s regression model was used for univariate and multivariate analysis. \u003cem\u003eP\u003c/em\u003e\u0026lt;0.05 was considered to indicated a statistically significant result.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cem\u003eAberrant expression of miR-124 in CCA tissues\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eFirstly, we compared the miR-124 expression in 119 CCA tissues and \u0026nbsp;para-cancerous tissues with qRT-PCR assay. As indicated in \u003cstrong\u003eFigure 1\u003c/strong\u003e, the miR-124 level was significantly lower in CCA tissues than para-cancerous samples (\u003cem\u003eP\u003c/em\u003e\u0026lt;0.001), suggesting that expression of miR-124 was down-regulated in CCA tissues.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAssociation of miR-124 expression with clinicopathological characteristics in CCA\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe correlation between miR-124 expression and clinicopathological parameters was examined in 119 cases (\u003cstrong\u003eTable 2\u003c/strong\u003e). Based on the median value (1.48) of miR-124 expression, all patients were divided into two groups as followings: low miR-124 expression group (n=63) and high miR-124 expression group (n=56). Among the parameters listed, a significant correlation was observed between low miR-124 expression and differentiation (\u003cem\u003eP\u003c/em\u003e=0.045), lymph node metastasis (\u003cem\u003eP\u003c/em\u003e=0.040). However, there were no relationships between miR-124 expression and other clinicopathological factors, including age, gender, tumor size, location, distant metastasis and TNM stage (all \u003cem\u003eP\u003c/em\u003e\u0026gt;0.05)(\u003cstrong\u003eFigure 2\u003c/strong\u003e).\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eRelationship between miR-124 expression and overall survival\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eNext, we analyzed the correlation of miR-124 expression with patients clinical outcome. Kaplan-Meier analysis showed that miR-124 expression was significantly associated with poor overall survival in CCA patients. According to log-rank test, the overall survival rate of patients with low miR-124 expression was lower than that of patients with high miR-124 expression (\u003cem\u003eP\u003c/em\u003e=0.001).\u003c/p\u003e\n\u003cp\u003eIn univariate Cox regression analysis, miR-124 expression (\u003cem\u003eP\u003c/em\u003e=0.003, HR=2.107, 95%CI: 1.287-3.448) was a prognostic factor for cancer mortality in CCA. Additionally, lymph node metastasis (\u003cem\u003eP\u003c/em\u003e=0.001, HR=2.269, 95%CI: 1.416-3.638) was also significantly related to overall survival. In a multivariate analysis, miR-124 expression was an unfavorable independent prognostic indicator in patients with CCA (\u003cem\u003eP\u003c/em\u003e=0.006, HR=2.006, 95%CI: 1.224-3.289).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eCCA is the most common primary hepatic malignancy worldwide. Despite tremendous improvements in developing multimodal treatments, the clinical outcome of CCA patients remains unfavorable. Thus, it is of great significance to identify powerful prognostic indicator for CCA.\u003c/p\u003e\n\u003cp\u003eThe discovery that non-coding components of the genome, including miRNAs, can conduct to the pathogenesis of cancer has led researchers to contemplate using these molecules to guide clinical strategies custom [16]. So far, a large number of studies suggest that miRNAs play a key role in tumor development. Increasing evidence suggest that deregulated expression of miRNAs participates in the initiation of CCA progression, where they act as either suppressors or promoters at the post-transcriptional regulation stage. For instance, Yang \u003cem\u003eet al.\u003c/em\u003e reported that miR-144 was significantly down-regulated in CCA tissues and might be a tumor suppressor of CCA cell proliferation and invasion through targeting LIS1 [17]. Zhang \u003cem\u003eet al.\u003c/em\u003e showed that miR-26a was increased in human CCA tissues and cell lines and overexpression of miR-26a increased proliferation of CCA cells and colony formation in \u003cem\u003evitro\u003c/em\u003e [18]. Wang \u003cem\u003eet al.\u003c/em\u003e revealed that miR-138 was underexpressed in CCA tissues, and downregulation of miR-138 promoted the proliferation, migration and invasion of CCA cells [19]. Liu \u003cem\u003eet al.\u003c/em\u003e confirmed that over-expression of miR-21 significantly promoted cell migration, invasion, and xenografts growth [20].\u003c/p\u003e\n\u003cp\u003ePrevious research confirmed that miR-124 was dysregulated in various cancers, and aberrant miR-124 expression was involved in the prognosis of tumors. For example, Dong \u003cem\u003eet al.\u003c/em\u003e showed that miR-124 had lower expression in breast cancer specimens and low miR-124 expression was a significant independent predictor of poor survival in breast cancer [21]. Gilje \u003cem\u003eet al.\u003c/em\u003e showed that miR-124 is a potential new biomarker for prediction of neurological prognosis following cardiac arrest [22]. Han \u003cem\u003eet al. \u003c/em\u003edemonstrated that miR-124 had a low expression in osteosarcoma tissues and low miR-124 expression was an unfavorable prognostic factor for overall survival [23]. Zhang \u003cem\u003eet al.\u003c/em\u003e proved that miR-124 expression level was significantly decreased in lung cancer tissues and miR-124 was an independent prognostic factor for overall survival in lung cancer [24]. However, to our knowledge, the clinical significance of miR-124 in CCA has not been reported. Therefore, we investigated the feasibility of miR-124 as a new potential biomarker of prognosis for CCA.\u003c/p\u003e\n\u003cp\u003eIn the present study, our data showed that miR-124 expression was significantly decreased in CCA tissues in comparisons with normal para-cancerous tissues. The relationship between miR-124 expression and various clinicopathological characteristics of CCA patients was investigated. The results indicated that abnormal miR-124 expression was dramatically correlated with differentiation and lymph node metastasis. It demonstrated that miR-124 might be involved in tumor metastasis and progression of CCA. Furthermore, the overall survival rate of low miR-124 group was significantly shorter than that of high miR-124 expression group. Finally, in a multivariate Cox model, we found that miR-124 expression was an independent poor prognostic indicator for overall survival of CCA patients. Our results were in line with previous studies. For example, Tian \u003cem\u003eet al.\u003c/em\u003e showed that miR-124 was reduced in CCA tissues and negatively related to lymph node involvement and distant metastasis [25].\u003c/p\u003e"},{"header":"Conclusion","content":" \u003cp\u003eIn conclusion, our results showed that miR-124 expression was remarkably reduced in CCA tissues. The expression of miR-124 was closely associated with the progression of CCA patients. More importantly, the down-regulation of miR-124 is an independent poor indicator of prognosis in CCA patients. We believed that it is just the beginning of a long and still unexplored avenue of a highly promising investigation of miR-124. A more in-depth and larger-scale study remains to identify the role of miR-124 in CCA.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003emicroRNAs (miRNAs)\u003c/p\u003e \u003cp\u003echolangiocarcinoma (CCA)\u003c/p\u003e \u003cp\u003equantitative reserve transcription polymerase chain reaction (qRT-PCR)\u003c/p\u003e \u003cp\u003eintrahepatic cholangiocarcinomas (IHCCs)\u003c/p\u003e \u003cp\u003eextrahepatic cholangiocarcinoma (EHCCs)\u003c/p\u003e \u003cp\u003e3\u0026rsquo;-untranslated region (3\u0026rsquo;-UTR)\u003c/p\u003e "},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was supported by the Ethics Committee of the PLA Rocket Force Characteristic Medical Center and also has been carried out in accordance with the World Medical Association Declaration of Helsinki.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe subjects had been informed the objective. Certainly, written consents were signed by every subject in this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFunding information is not applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026rsquo; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eN.W. design of the work; Y.L. the acquisition, analysis, Y.Z. interpretation of data; H.C. the creation of new software used in the work; X.W., Z.L. have drafted the work or substantively revised it. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003ePatel T: Increasing incidence and mortality of primary intrahepatic cholangiocarcinoma in the United States. \u003cem\u003eHepatology \u003c/em\u003e2001, 33(6):1353-1357.\u003c/li\u003e\n\u003cli\u003eBergquist A, von Seth E: Epidemiology of cholangiocarcinoma. \u003cem\u003eBest practice \u0026amp; research Clinical gastroenterology \u003c/em\u003e2015, 29(2):221-232.\u003c/li\u003e\n\u003cli\u003eGatto M, Bragazzi MC, Semeraro R, Napoli C, Gentile R, Torrice A, Gaudio E, Alvaro D: Cholangiocarcinoma: update and future perspectives. \u003cem\u003eDigestive and liver disease : official journal of the Italian Society of Gastroenterology and the Italian Association for the Study of the Liver \u003c/em\u003e2010, 42(4):253-260.\u003c/li\u003e\n\u003cli\u003eBlechacz B, Gores GJ: Cholangiocarcinoma: advances in pathogenesis, diagnosis, and treatment. \u003cem\u003eHepatology \u003c/em\u003e2008, 48(1):308-321.\u003c/li\u003e\n\u003cli\u003ePatel T: Cholangiocarcinoma--controversies and challenges. \u003cem\u003eNature reviews Gastroenterology \u0026amp; hepatology \u003c/em\u003e2011, 8(4):189-200.\u003c/li\u003e\n\u003cli\u003eHe L, Hannon GJ: MicroRNAs: small RNAs with a big role in gene regulation. \u003cem\u003eNature reviews Genetics \u003c/em\u003e2004, 5(7):522-531.\u003c/li\u003e\n\u003cli\u003eBartel DP: MicroRNAs: target recognition and regulatory functions. \u003cem\u003eCell \u003c/em\u003e2009, 136(2):215-233.\u003c/li\u003e\n\u003cli\u003eBartel DP: MicroRNAs: genomics, biogenesis, mechanism, and function. \u003cem\u003eCell \u003c/em\u003e2004, 116(2):281-297.\u003c/li\u003e\n\u003cli\u003eHezova R, Kovarikova A, Srovnal J, Zemanova M, Harustiak T, Ehrmann J, Hajduch M, Sachlova 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F: miR-124 regulates adult neurogenesis in the subventricular zone stem cell niche. \u003cem\u003eNature neuroscience \u003c/em\u003e2009, 12(4):399-408.\u003c/li\u003e\n\u003cli\u003eZhang J, Lu Y, Yue X, Li H, Luo X, Wang Y, Wang K, Wan J: MiR-124 suppresses growth of human colorectal cancer by inhibiting STAT3. \u003cem\u003ePloS one \u003c/em\u003e2013, 8(8):e70300.\u003c/li\u003e\n\u003cli\u003eLi L, Luo J, Wang B, Wang D, Xie X, Yuan L, Guo J, Xi S, Gao J, Lin X\u003cem\u003e et al\u003c/em\u003e: Microrna-124 targets flotillin-1 to regulate proliferation and migration in breast cancer. \u003cem\u003eMolecular cancer \u003c/em\u003e2013, 12:163.\u003c/li\u003e\n\u003cli\u003eNana-Sinkam SP, Croce CM: Clinical applications for microRNAs in cancer. \u003cem\u003eClinical pharmacology and therapeutics \u003c/em\u003e2013, 93(1):98-104.\u003c/li\u003e\n\u003cli\u003eYang R, Chen Y, Tang C, Li H, Wang B, Yan Q, Hu J, Zou S: MicroRNA-144 suppresses cholangiocarcinoma cell proliferation and invasion through targeting platelet activating factor acetylhydrolase isoform 1b. \u003cem\u003eBMC cancer \u003c/em\u003e2014, 14:917.\u003c/li\u003e\n\u003cli\u003eZhang J, Han C, Wu T: MicroRNA-26a promotes cholangiocarcinoma growth by activating beta-catenin. \u003cem\u003eGastroenterology \u003c/em\u003e2012, 143(1):246-256 e248.\u003c/li\u003e\n\u003cli\u003eWang Q, Tang H, Yin S, Dong C: Downregulation of microRNA-138 enhances the proliferation, migration and invasion of cholangiocarcinoma cells through the upregulation of RhoC/p-ERK/MMP-2/MMP-9. \u003cem\u003eOncology reports \u003c/em\u003e2013, 29(5):2046-2052.\u003c/li\u003e\n\u003cli\u003eLiu Z, Jin ZY, Liu CH, Xie F, Lin XS, Huang Q: MicroRNA-21 regulates biological behavior by inducing EMT in human cholangiocarcinoma. \u003cem\u003eInternational journal of clinical and experimental pathology \u003c/em\u003e2015, 8(5):4684-4694.\u003c/li\u003e\n\u003cli\u003eDong LL, Chen LM, Wang WM, Zhang LM: Decreased expression of microRNA-124 is an independent unfavorable prognostic factor for patients with breast cancer. \u003cem\u003eDiagnostic pathology \u003c/em\u003e2015, 10:45.\u003c/li\u003e\n\u003cli\u003eGilje P, Gidlof O, Rundgren M, Cronberg T, Al-Mashat M, Olde B, Friberg H, Erlinge D: The brain-enriched microRNA miR-124 in plasma predicts neurological outcome after cardiac arrest. \u003cem\u003eCrit Care \u003c/em\u003e2014, 18(2):R40.\u003c/li\u003e\n\u003cli\u003eHan G, Wang Y, Bi W, Jia J, Wang W: MicroRNA-124 functions as a tumor suppressor and indicates prognosis in human osteosarcoma. \u003cem\u003eExperimental and therapeutic medicine \u003c/em\u003e2015, 9(3):679-684.\u003c/li\u003e\n\u003cli\u003eZhang Y, Li H, Han J: Down-regulation of microRNA-124 is correlated with tumor metastasis and poor prognosis in patients with lung cancer. \u003cem\u003eInternational journal of clinical and experimental pathology \u003c/em\u003e2015, 8(2):1967-1972.\u003c/li\u003e\n\u003cli\u003eTian F, Chen J, Zheng S, Li D, Zhao X, Jiang P, Li J, Wang S: miR-124 targets GATA6 to suppress cholangiocarcinoma cell invasion and metastasis. \u003cem\u003eBMC cancer \u003c/em\u003e2017, 17(1):175.\u003c/li\u003e\n\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003e\u003cstrong\u003eTable 1. \u003c/strong\u003eThe relationship between miR-124 expression and clinicopathological features\u003c/p\u003e\n\u003ctable\u003e\n\u003ctbody\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 66px;\" rowspan=\"2\" width=\"168\"\u003e\n\u003cp\u003eFeatures\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 66px;\" rowspan=\"2\" width=\"119\"\u003e\n\u003cp\u003eCases (n=119)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" colspan=\"2\" width=\"204\"\u003e\n\u003cp\u003eMiR-124 expression\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 66px;\" rowspan=\"2\" width=\"83\"\u003e\n\u003cp\u003e\u003cem\u003e\u0026chi;\u003csup\u003e2\u003c/sup\u003e\u003c/em\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 66px;\" rowspan=\"2\" width=\"83\"\u003e\n\u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003eLow (n=63)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003eHigh (n=56)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eAge (years)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003e\u0026lt;68\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e58\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e26\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e32\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e2.990\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.084\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003e\u0026ge;68\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e61\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e37\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e24\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eGender\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eMale\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e73\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e43\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e30\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e2.695\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.101\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eFemale\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e46\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e20\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e26\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eTumor size (cm)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003e\u0026lt;5\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e63\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e33\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e32\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.271\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.603\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003e\u0026ge;5\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e54\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e30\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e24\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eLocation\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eIntrahepatic\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e24\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e14\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.351\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.554\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eExtrahepatic\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e95\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e49\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e46\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eDistant metastasis\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eNegative\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e88\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e42\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e46\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e3.686\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.055\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003ePositive\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e31\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e21\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eDifferentiation\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eWell-moderate\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e76\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e35\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e41\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e4.006\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.045\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003ePoor\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e43\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e28\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33.1875px;\"\u003e\n\u003ctd style=\"height: 33.1875px;\" width=\"168\"\u003e\n\u003cp\u003eTNM stage\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33.1875px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33.1875px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33.1875px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33.1875px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33.1875px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eⅠ-Ⅱ\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e61\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e28\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e33\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e2.489\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.115\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eⅢ-Ⅳ\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e58\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e35\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e23\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eLymph node metastasis\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003eAbsent\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e69\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e31\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e38\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e4.233\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e0.040\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr style=\"height: 33px;\"\u003e\n\u003ctd style=\"height: 33px;\" width=\"168\"\u003e\n\u003cp\u003ePresent\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"119\"\u003e\n\u003cp\u003e50\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"104\"\u003e\n\u003cp\u003e32\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"100\"\u003e\n\u003cp\u003e18\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd style=\"height: 33px;\" width=\"83\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2. \u003c/strong\u003eThe univariate and multivariate analyses of clinical factors for overall survival of CCA\u003c/p\u003e\n\u003ctable\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"2\" width=\"166\"\u003e\n\u003cp\u003eFactors\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd colspan=\"2\" width=\"265\"\u003e\n\u003cp\u003eUnivariate analysis\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd colspan=\"2\" width=\"226\"\u003e\n\u003cp\u003eMultivariate analysis\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003eHR (95%CI)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003eHR (95%CI)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eMiR-124 expression\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e2.107 (1.287-3.448)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.003\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e2.006 (1.224-3.289)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e0.006\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eAge\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e0.901 (0.565-1.438)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.663\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eGender\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e1.504 (0.924-2.448)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.100\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eTumor size (cm)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e1.353 (0.848-2.160)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.204\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eLocation\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e0.814 (0.466-1.422)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.470\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eDistant metastasis\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e1.397 (0.839-2.327)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.199\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eDifferentiation\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e1.153 (0.711-1.870)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.563\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eTNM stage\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e1.446 (0.905-2.310)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.122\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"166\"\u003e\n\u003cp\u003eLymph node metastasis\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"155\"\u003e\n\u003cp\u003e2.269 (1.416-3.638)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"110\"\u003e\n\u003cp\u003e0.001\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"142\"\u003e\n\u003cp\u003e2.175 (1.354-3.492)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"85\"\u003e\n\u003cp\u003e0.001\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"MicroRNA-124, Cholangiocarcinoma, Survival, Prognosis","lastPublishedDoi":"10.21203/rs.3.rs-33111/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-33111/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground:\u003c/strong\u003e Previous studies have demonstrated that microRNAs (miRNAs) played a crucial role in various diseases, including cancers. The aim of the study was to evaluate the clinical significance of miR-124 in patients with cholangiocarcinoma (CCA).\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eMethods:\u003c/strong\u003e The expression pattern of miR-124 was detected in CCA tissues using quantitative reserve transcription polymerase chain reaction (qRT-PCR). The correlation of miR-124 expression with clinicopathological features and overall survival of patients were explored using chi-square test, Kaplan-Meier methods and Cox regression analyses.\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eResults:\u003c/strong\u003e The miR-124 expression level was strong down-regulated in CCA tissues compared with normal para-cancerous tissues (\u003cem\u003eP\u003c/em\u003e\u0026lt;0.001). Moreover, aberrant miR-124 expression was significantly associated with differentiation (\u003cem\u003eP\u003c/em\u003e=0.045) and lymph node metastasis (\u003cem\u003eP\u003c/em\u003e=0.040). In addition, Kaplan-Meier method and log-rank test revealed that patients with low miR-124 expression has a poorer overall survival compared with those with high miR-124 expression (\u003cem\u003eP\u003c/em\u003e=0.002). Furthermore, multivariate analysis confirmed that miR-124 expression (\u003cem\u003eP\u003c/em\u003e=0.006; HR=2.006; 95%CI: 1.224-3.289) was an independent prognostic indicator in CCA.\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eConclusions:\u003c/strong\u003e Collectively, our results defined miR-124 expression plays important roles in CCA patients. MiR-124 expression might used as a valuable prognostic biomarker for patients with CCA.\u003c/p\u003e","manuscriptTitle":"MicroRNA-124 alone might represent an indicator signifying cholangiocarcinoma prognosis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2020-06-08 21:05:18","doi":"10.21203/rs.3.rs-33111/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"3ad863bd-b8c8-4ee8-8552-7cbe3fb2cf2e","owner":[],"postedDate":"June 8th, 2020","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":115583,"name":"Cancer Biology"}],"tags":[],"updatedAt":"2020-07-23T19:51:26+00:00","versionOfRecord":[],"versionCreatedAt":"2020-06-08 21:05:18","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-33111","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-33111","identity":"rs-33111","version":["v1"]},"buildId":"WrCJVZZCHTDjtuVLN7oU0","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}