| Yes-associated protein (YAP) is the downstream target of the Hippo signaling pathway, which is a transcription co-activator cooperates with TEAD transcription factor. Over-expression of YAP can induce proliferation of cancer cells. A study which was based on a Chinese cohort in Hong Kong showed that YAP was an independent prognostic marker of Hepatocellular carcinoma (HCC) patients.Nowadays, HCC is one of the most common cancers worldwide, especially in Asia. Hepatic resection remains the best choice of HCC therapeutic strategies, but the short-term recurrence is still a problem which leads poor prognosis.Clinical evidence suggests that stimulation of tumor growth may occur after resection of liver tumors, and that liver regeneration could accelerate cancer cell proliferation. So, it is necessary to study YAP in liver regeneration. It is important to confirm whether YAP was associated with HBV correlation factors and determine the mechanism by which YAP is related to prognosis.Purpose:1. The first aim of this study was to assess the relationship between YAP and some pathological features of HCC, especially HBV correlation factors, to search the clue of YAP leading poor prognosis.2. The second aim was to assess the relationship between YAP and tumor short-term recurrence and prognosis of HCC.3. The third aim of this study was to investigate YAP gene expression and protein levels during rat liver regeneration, thereby explore possible relationships between YAP and LATS1 in regenerating liver.4. The fourth aim was to evaluate the influence of cancer cell proliferation activity and the level of LATS1 and TGF-β1 when YAP gene was down regulated in human hepatocellular carcinoma cells.Experimental design or Method:1. A tissue microarray including 80 HCC samples were retrospectively analyzed. The expression of YAP by IHC in a cohort of liver cancer and matched adjacent non-tumor tissue samples from HCC patients after liver resection. The Pearson Chi-Square test, Independent-Sample T Test and Kaplan-Meier survival analysis were used. Besides, we examined YAP expression in the process of growth of rat liver cancer using immunohistochemical method.2. The animal model of 70% rat hepatectomy was made. Samples were taken at designated time intervals: 4 h, 48 h, 104 h, 115 h, and 240 h post-treatment, with five rats per group. We examined YAP expression in rat regenerating liver using real-time PCR, Western blotting, and immunohistochemical methods. Synchronized changes of the protein level of YAP and LATS1 were visualized in rat regenerating tissues using Western blotting method.3. The YAP-shRNA was transduced into 97H cells with lentiviral vectors. We study the proliferation ability of 97H-YAP-RNAi(+) and 97H-YAP-RNAi(–) cells by MTS technology and cell cycle analysis. We investigated the expression of YAP in human hepatocellular carcinoma cells (97H, 97L).Results:1. Results showed YAP was associated with HCC differentiation not HBV correlation factors. The mean onset age of patients with YAP-positive HCC (46.30±9.18) was lower than patients with YAP-negative HCC (51.21±11.63) (P = 0.047). The YAP-positive group had a higher proportion of patients who were dead or had recurrence within two years after partial hepatectomy, and had a lower survival rate.2. Our results showed that YAP protein levels were markedly increased in the regenerating liver (2.5-3 times), while YAP mRNA levels decreased slightly (1.47-2.17 times) (P =0.017). Both YAP mRNA and protein levels were higher in hepatocellular carcinoma (HCC) than in para-cancerous tissue. Besides, LATS1 decreased gradually in the process of liver regeneration.3. Knockdown of YAP by lentivirus-shRNA significantly inhibited 97H cell growth in vitro, at the same time LATS1 protein decreased but TGF-β1 mRNA increased. Besides, the liver cancer cell with high level YAP gene and protein had high proliferation and metastatic potential.Conclusions:1. YAP-positive HCCs usually have poorly-differentiated characteristics, the patients have early onset and poor prognosis. The expression of YAP is not obviously associated with HBV correlation factors.2. Elevated YAP level in liver regeneration is from the reduction of YAP phosphorylation, which is mediated by the LATS1. The abnormally high expression of YAP in liver cancer cell is from high level of YAPmRNA.3. When liver regeneration happened in YAP-positive HCC patients, further elevated YAP protein in residual cancer may accelerate the proliferation of cancer cells.4. Knockdown of YAP is a independent factor which can inhibit hepatocellular carcinoma cell proliferation.
|
PDF Full Text Request