| The induction of immune tolerance after liver transplantation was contributed to high expression of regulatory T cells which can increase the risk of tumor recurrence and reduce the long-term survival rate for patients with hepatocellular carcinoma (HCC) after liver transplantation(LT). So, the immune tolerance induction and tumor recurrence prevention for these patients postoperative was mutually contradictory. Pervasive application of Tacrolimus (FK506) can promote the risk of tumor recurrence, while the mTOR inhibitors (Sirolimus, SRL) can replace this adverse reaction. Facing enormous pressure of tumor recurrence for HCC patients after LT, there is also no effective control strategy.As a result, seeking the immune suppression program which has the anti-tumor effect meanwhile was so important. Therefore, the purpose of this research is to analyze the function of Treg on the immunoregulation and anti-tumor immunity for HCC liver transplantation recipients with Sirolimus Combined Therapeutic Regimen (SCTR) also called Triple Anti-tumor Therapy (TATT) and to further explore the survival value of this scheme for HCC liver transplantation recipients. Based on this, the research thesis is divided into three parts to discuss.Part I:Treg expression difference between different liver transplantation groupsThe FoxP3+Treg expression level of patients with HCC and tumor relapse after HCC liver transplantation was significantly higher than that in normal group (P> 0.05). The FoxP3+Treg of patients with cirrhosis liver transplantation was maintained at near normal levels (P> 0.05), which was kept in a lower level of patients with no tumor relapse and long-term survival after HCC liver transplantation, the rejection rate of two groups was doesn’t increased. The variation tendency of FoxP3+Treg for HCC liver transplantation patients was decreased to the bottom level immediately after LT, then gradually increased to a higher level at early stage (with no significant difference compared with normal and cirrhotic liver transplantation recipients), began to gradually reduce after 6 months, was maintained at a low level after 1 year postoperative similarly with long-term survival patients intervened by SCTR (P< 0.05). These suggested that the relatively high level of FoxP3+Treg nearing the normal level for liver cirrhosis after LT helps to maintain status for long-term immunosuppression, and has little effect on the cellular immune function with no increasing the incidence of rejection; compared with this, the FoxP3+Treg level of PB for patients after LT was effectively down-regulated by SCTR intervention program which can generate in comparable strength of immune inhibition, at the same time, maintain a sufficient number of CD8+T effector cells to play anti-tumor cellular immune function, achieving the anti-tumor relapse effect. Therefore, we believed that this low level of FoxP3+Treg contributed to the long-term survival for liver transplantation patients, meanwhile reduced the risk of tumor recurrence.Part Ⅱ:Relevance analysis between immunoregulation and anti-tumor immune on animal model which was simulated the characteristics of tumor recurrence after HCC liver transplantationTo establish the animal model which was simulated the characteristics of tumor recurrence after HCC liver transplantation with the chemical-induced cancer method. Taking the peripheral blood (PB) and then killing the rats after the ending of experiment. The macroscopic liver granular nodules in model group, the max was almost 10mm, HE staining was showed HCC. On the contrary, hepatic pathological changes were obviously alleviated by drug intervention which most remarkable in TATT group.FoxP3+Treg of PB in model control group was significantly higher than that in normal group and intervention group (P< 0.05), however, TATT intervention can significantly reduce the levels of FoxP3+Treg (P< 0.05). For the single drug intervention, although the level of FoxP3+Treg in PB was decreased, there were no significant differences between groups (P> 0.05). The FoxP3+Treg in liver and spleen were changed similarly as the PB level except the model control group which was significantly lower than that of peripheral blood levels (P< 0.05). At the inhibitory effect aspect of Treg, the number of CD4+and CD8+T cells of PB, liver and spleen was decreased most obviously in the model control group (P< 0.05), nevertheless, the number was significantly increased by TATT intervention which closed to the normal level (P> 0.05) and also significantly higher than the single drug group and model control group (P< 0.05). Furthermore, negative correlation with the expression of CD4+T, CD8+T and CD4+/CD8+T.At the antigen Presentation and recognition aspect, dendritic cells (Dendritic Cell, DCs) expression levels were significantly improve by the TATT method which had significantly difference in statistic compared with model control group and single drug intervention groups (P< 0.05) and in contrast with the expression of FoxP3+Treg cells.For the HCC associated index, AFP level of model control group was significantly higher than that of the normal level (P< 0.05), were decreased by drug intervention which also most remarkable in TATT group (P< 0.05). The level of Vascular Endothelial Growth Factor (VEGF) in PB of model control group was significantly increased (P< 0.05) in contrast with normal level and the VEGF expression in HCC tissue with immumohistochemical staining was also increased significantly (P< 0.05). Similarly, both in the PB and HCC tissue, VEGF expression of TATT group was significantly lower than normal, model control group and single drug groups (P< 0.05).To sum up, in the simulation immunosuppressive environment, we find that TATT intervention can significantly inhibit the growth of hepatocarcinoma cells or canceration process, significantly prolong the time of death, decrease mortality(P< 0.05), this can attribute to TATT treatment which can mainly decrease the expression of FoxP3+Treg effectively and maintain at a relatively low state, weaken the direct and indirect inhibitory effect on effect T cells, increasing the expression of antigen presenting cell CD11c+DCs to inhibition the proliferation of FoxP3+Treg, all these can further enhance the cellular immune response through increasing the presentation, recognition, damage and monitoring role of tumor antigens, finally play the role of inhibit growth and proliferation of cancer cell.Part III:To research the long-term survival value of SCTR program in graft after liver transplantation for HCCWith the retrospective cohort studies, the patients with HCC beyond UCSF criterion after LT who accepted the SCTR treatment were followed up to 2014 December, finding that SCTR treatment can significantly prolong the survival time (1,3 year overall survival rate and disease-free survival rate; 100%vs.94.4%) and delay the tumor relapse, and significantly reduce the level of FoxP3+Treg cells, and the maintenance of AFP in normal the level of HCC, suggesting that SCTR/TATT method can be used as a effective choice for prevention and treatment of tumor recurrence after LT, the FoxP3+Treg level increasing abnormally postoperative which was obviously earlier than the AFP changes helped to indicate the tumor recurrence, suggesting that early detection of FoxP3+Treg cells postoperative can be as one of the monitoring index of tumor relapse. |
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