| Hepatocellular carcinoma (HCC) is one of the major diseases that endanger the health of Chinese people, its incidence rates are ranked the 6th and the mortality rates are the 3rd of the all malignancy.HCC is aprototype of hypervascular tumors. Compared with other solid tumors, it is difficult to diagnose at the early stage. Over 90%patients who are diagnosed are already in the far advanced stages. Notably, HCC generation and progression exhibit high metastatic and invasive characteristics, especially intrahepatic and distal metastasis as well as frequent postoperative recurrence, which are the main cause of high mortality. In order to improve the early diagnosis and prognosis, we urgently need to further explore the mechanism of promoting the development of HCC and the occurrence of early metastasis. At the same time, a large number of clinical data showed that the occurrence and progression of HCC exhibit significant gender disparity. Gradually to get the consensus is that estrogen plays a very important role in the occurrence and development of HCC, however, compared to the other sex hormone sensitive organs, relatively few studies of the role of estrogen in HCC, especially to explore its molecular mechanism is obviously insufficient. Therefore, based on the analysis of a large number of human chronic hepatitis and HCC samples, combined with in vitro experiments and animal models, the role of ER alpha (ERa) and miR-30ein hepatocarcinogenesis have been systematically investigated, and following main results are obtained:1. Based on a large number of human HCC clinical specimens, the transcription and translation of MTA1 gene in HCC tissue, corresponding adjacent tissue, and normal tissue samples were determined by quantity real-time polymerase chain reaction (PCR) and immunohistoehemistry (IHC), and the correlation between MTA1 and survival and recurrence of HCC patients was analyzed by Kaplan-Meier curve followed by log-rank test. The results showed that the transcription and translation of MTA 1 gene were significantly higher in the HCC tissues and the adjacent cirhotic tissues compared to the normal liver tissues (P<0.01). Expression level of MTA1 was related to survival and recurrence rates of the HCC patients. Based on this result, we can conclude that MTA1 was a suitable prognostic index predicting survival and recurrence after surgical resection of HCC.2. Sex disparity in HCC development is a long term hot spot in international HCC research field. In our study, quantitative real-time PCR and western blot analyses were used to measure levels of MTA1. The effecton HCC cell proliferation and invasion was assessed by EdU incorporation assays and Transwell, respectively. ShRNAand dual-luciferase assays were used to investigate the regulatory relationship between MTA1 and ERain cell lines. We found that significant high expression of MTA1 was investigated in male tumor and adjacent tissues compared to normal liver tissue, while it is not the case in female. Increased MTA1 was linear correlated to decreased ERain male tumor and adjacent tissues which is due to transcription suppression by E2/ERαcomplex on MTA1 promoter region confirmedby both in vitro cell model and luciferase reporter gene assay. The MTAl promoter has three functional ER-element half-sites that lead to decreased MTAl transcription and expression.ERaoverexpression suppressed the proliferation and invasion of hepatocellular carcinoma cells (HCC). In addition,overexpression of MTA1 attenuated ERa-mediated suppression of the proliferation and invasion ofHCC cells andtumor formation in vivo. These results suggested feedback regulation between ERaand MTA1. In summary, ourresults demonstrated that ERasuppressed proliferation and invasion of human HCC cells through down-regulationof MTA1 transcription. Thus, a novel cell signaling pathway interpreting sex disparity of HCC incidence was discovered:down-regulated expression of ERain male HCC and adjacent tissues attenuated the protection by E2 in chronic hepatitis and premalignant which led to overexpression of MTA1 and highly incidence rates in male inHCC.3. MTA1 was a classic metastasis related protein which was essential toepithelial-to-mesenchymal transition (EMT); however, more information concerning its up and down-stream regulation was rare. In this study, we sought to investigate its up-stream regulation and down-stream effector in human hepatocellular carcinoma (HCC). Expression of varying genes was determined by both real-time PCR and immunohistochemistry, statistical analysis and bioinformatics was involved to predict potential association and regulations, both in vivo and in vitro studies were applied to verify the regulations. Totally 94 paired HCC and adjacent tissues were involved in the study, and the results indicated that decreased of miR-30e was associated with increased MTA1 in human HCC.MiR-30e exerted its regulation on MTA1 transcription by binding to its 3’UTR region, and was negatively associated with EMT. Furthermore, significantly high expression of MTA1 was associated with overexpression of ErbB2 in human HCC, and MTA1 can promote transcription of ErbB2 acting as a promoter by binding with HDAC2. The EMT promotion effect by MTA1 was largely depended on ErbB2, either knockdown or inhibit the activity of ErbB2 can significantly attenuated EMT promotion by overexpressing MTA1 both in vitro and in vivo. In general, down-regulation of miR-30e can enhance MTA1 in human HCC, and furthermore to promote cell invasion and metastasis by promoting ErbB2.In conclusion,according to our studies,MTAlis not only involved in theHCC development and growth,but also closely associated with the post-operative recurrenceand a poor prognosis or a worse survival rates. On one hand, we uncovered that down-regulation of ERain male HCC and adjacent tissues could up-regulate MTA1 expression as a transcription factor which impact on the biological characteristics of HCC such as proliferation and metastasis by enhancing the proliferation and invasion capacity. On the other hand, down-regulation of miR-30ein human HCC can also regulate MTAl expression at the post-transcription level, increase ErbB2 expression and furthermore enhance the invasion and metastasis of HCC by promoting EMT. In summary, our study is an adding understanding to the gender disparity of HCC progression. Clarification of the mechanism of MTA1 promoted hepatocarcinogenesis is important for the identification of novel therapeutic targets in HCC prevention and treatment. |
PDF Full Text Request