Dissertation > Excellent graduate degree dissertation topics show

Immunosuppressant Tacrolimus on the mechanism of invasion and metastasis of hepatocellular carcinoma

Author: ZhouShaoLai
Tutor: FanJia
School: Fudan University
Course: Biochemistry and Molecular Biology
Keywords: HCC Tacrolimus Liver transplantation VEGF-C Lymphatic metastasis
CLC: R735.7
Type: Master's thesis
Year: 2011
Downloads: 63
Quote: 0
Read: Download Dissertation

Abstract

Hepatocellular Carcinoma (HCC) is one of the most common cancers in the world. The mortality of HCC took the third place among all the malignaant tumors and the second place in China. Five year survival rate has been greatly developed due to the conception of "early diagnosis and early treatment of HCC". Hepatectomy by now is still the first choise for people who desire a long term survival, while only 20% of the patients fit the hepatectomy indication cause of the liver cirrhosis and hypohepatia. Liver transplantation (LT) is ideally the optimal treatment for HCC on cirrhosis as it treats both the tumor and the underlying condition that generated it. But still HCC recurrence after liver transplantation. Immunosuppression increases the risk of recurrence as a independent predictor.The most common immunosuppress therapeutic protocol is FK506/CsA+MMF+glucocorticoid in worldwide, while some transplantation centers replace FK506/CsA of rapamycin. Both FK506 and CsA are calcineurin inhibitors. They affect the T-cell and B-cell response by binding to immunophilins and interferring with the activation of calcineurin. This is the orchestrated suppression of immune responses leads to the therapeutic efficacy. Some in vivo researches and clinical retrospective reviews indicate that calcineruin inhibitors is probably an independent predictor for the HCC recurrence after LT. However, some in vitro experiments show that CsA/FK506 can not inhibit or promote the growth of the hepatoma carcinoma cell. Whether or not calcineurin inhibitors promote the cancer recurrence after LT is still in dispute.This article is to elucidate the effect of tacrolimus treatment on tumor growth and metastasis of hepatocellular carcinoma (HCC). In our results, we found that tacrolimus had no effect on the proliferation of HCC in vitro or in vivo. Treatment with tacrolimus resulted in a dose-dependent increase in the invasion potential of HCC cells in vitro, in the density of peritumoral lymphatic vessels, and in the number and volume of metastatic lymph nodes in ACI rats. qRT-PCR, immunohistochemisty and Western blot revealed that tacrolimus increased the levels of expression of VEGF-C in HCC. Then, we use TMA To explore the expression of VEGF-C protein in HCC tissues and to analyse the relationship between its expression with clinical pathology, invasiveness and prognosis of HCC.Part one Effect of FK506 on growth and metastasis of HCCObjective:To elucidate the effect of tacrolimus treatment on tumor growth and metastasis of hepatocellular carcinoma (HCC).Materials:In vitro CCK8 assays were performed to determine whether tacrolimus affects the growth of HCC cells under various concentration (0ng/ml;10ng/ml;100ng/ml)at different time(24h,48,72h,96h) and 24-well BioCoat Matrigel Invasion Chambers were used to determine the effect of FK506 on the invasion potential of HCC cells under various concentration (0ng/ml;10ng/ml;100ng/ml). In vivo HCC-bearing rats, we tested effect of FK506 on tumor growth and metastasis under low (0.15mg/kg)and high(0.3mg/kg) dosage of FK506.Results:Tacrolimus had no effect on the proliferation of HCC in vitro or in vivo. Treatment with tacrolimus resulted in a dose-dependent increase in the invasion potential of HCC cells in vitro and in the number and volume of metastatic lymph nodes in ACI rats. Conclusion:FK506 increases the invasion potential of HCC cells in vitro and promotes lymphatic metastasis on HCC-bearing ACI rats.Part two The mechanism of HCC lymphatic metastasis induced by FK506Objective:To test the mechanism of HCC lymphatic metastasis induced by FK506.Materials:Levels of VEGF-C expression by HCC cells were measured by qRT-PCR, Western blot under various concentrations (Ong/ml; 10ng/ml; lOOng/ml) of FK506, qRT-PCR, immunohistochemisty and Western blot tested the levels of expression of VEGF-C in HCC tissues, lymphangiogenesis were assessed by VEGFR-3 immunostaining on tumor and peritumor tissues. Results:Tacrolimus enhanced VEGF-C mRNA and protein expression in Hep 3B, MHCC97H and MH3924A cells in a dose-dependent manner. Tumor VEGF-C expression in HCC-bearing rats was significantly higher in the 0.15 mg/kg tacrolimus groups and more in the 0.3 mg/kg tacrolimus groups when compared with controls. The density of peritumor lymphatic vessels was significantly increased in the 0.15 mg/kg tacrolimus group and in the 0.3 mg/kg tacrolimus group when compared with controlsConclusion:Tacrolimus promoted HCC lymphatic metastasis might be duo to increasing VEGF-C expression by HCC cells and facilitates HCC lymphangiogenesis. Part 3 VEGF-C protein in HCC tissues and corresponding clinical significanceObjective:To explore the expression of VEGF-C protein in HCC tissues and to analyse the relationship between its expression with clinical pathology, invasiveness and prognosis of HCC.Materials:We used high-throughput TMA technology and immunochemistry to dectect VEGF-C expression in a group of 323 HCC patients after curative resections and in another group of 159 HCC patients after transplantation, and then made correlation analysis between VEGF-C with other clinical-pathological characters and survival rates. We also compared VEGF-C expression in predicting the reccurence of HCC after curative resections or LT, and evaluated the significance of VEGF-C as a predictive marker for the prognosis of HCC after curative resections or LT.Results:We used high-throughput TMA technology to assess the relationship between VEGF-C and prognosis of HCC in a group of 323 HCC patients who underwent curative resections and another group of 159 HCC patients who underwent LT. In one group of 323 HCC patients who underwent curative resections Patients with negative VEGF-C expression had no significantly prognosis than VEGF-C - positive patients (DFS, P=0.51; OS, P=0.43). The 1-,3- and 5-year DFS and OS rates for VEGF-C- and VEGF-C ’patients were 78% and 86% versus 50% and 64%,41% and 52% versus 70% and 85%, and 50% and 61% versus 39% and 50%, respectively. In another group of 159 HCC patients who underwent LT, Patients with positive VEGF-C expression had a significantly poorer prognosis than VEGF-C-negative patients (DFS, P= 0.009; OS, P= 0.005). The 1-,3-and 5-year DFS and OS rates for VEGF-C" and VEGF-C+patients were 86% and 93% versus 68% and 78%,58% and 68% versus 75% and 86%, and 47% and 60% versus 36% and 48%, respectively. The prognostic value of VEGF-C expression was evaluated in HCC patients using univariate analysis, which showed that TNM stage, vascular invasion, tumor size, GGT, tumor differentiation and tumor envelope were predictors for DFS and OS. Multivariate analysis was conducted with four of the variables (VEGF-C expression, vascular invasion, tumor size, GGT, tumor differentiation and tumor envelope), which was shown to be significant in the univariate analysis and no obvious correlation between each other. VEGF-C expression was the independent variable for predicting poor DFS and OS.Conclusion:VEGF-C expression can not predict DFS and OS on HCC patients after curative resections. But in HCC patients after LT, VEGF-C expression was the independent variable for predicting poor DFS and OS.

Immunosuppressant Tacrolimus on the mechanism of invasion and metastasis of hepatocellular carcinoma

Abstract

Related Dissertations