The Effect Of Platelet In The Formation Of Hepatocellular Carcinoma With Portal Vein Tumor Thrombosis

BackgroundHepatocellular carcinoma(HCC),the third most fatal cancer,has high mortality and poor prognosis around the world.Although numerous aggressive or palliative therapies were reported,the prognosis of HCC patients still remains unsatisfactory.One of the important reason is the high rate of metastasis and portal vein invasion(PVTT)in them.Growing evidences showed that platelet(PLT)plays an important role in Oncogenesis and tumor metastasis.However,the relationship between PLT and PVTT is still remain unknown.PurposeThis study to evaluate the relationship between PLT count and the prognosis of HCC patients with PVTT and to investigate the mechanisms of platelet involved in the process of PVTT formation.Methods1.Survival analysis of HCC patients with PVTT with different platelet countsA total of 694 HCC patients with PVTT,divided into three independent cohorts according to platelet counts,were analysed to test the correlation between platelet counts and the prognosis of PVTT patients.Preoperative PLT level were measured at baseline and ranged from 100 to 300×10^9/L was used as normal standard.A PLT count of <100×10^9/L or >300×10^9/L was defined as thrombocytopenia or thrombocytosis,respectively.2.Detection of Platelet activation of HCC paitents with PVTT and gene expression profiling among different tissues of themPlatelet activation of HCC patients with or without PVTT was tested by platelet aggregation assay and flow cytometry.cDNA microarray analyses were used to investigate the specially highly expressed mRNAs in PVTT tissues and the corresponding tumor tissues.According to the results of the cDNA microarray,GO and KEGG pathway analysis was used to analyse the abnormal signal pathways and reveal the molecular mechanisms of PVTT.3.Evaluating the mechanisms of platelet in PVTTCandidate mRNA was selected on the basis of the results of GO and KEGG pathway analysis of differential expressed mRNAs in the PVTT tissues and the expression of it was validated in more PVTT tissues and cell lines.Thrombogenesis experiment was performed through co-culture between HCC cell lines and platelet in vitro.Results1.Of the all study population,thrombocytopenia was more prevalent in PVTT patients than in those with thrombocytosis(20.9%).Survival analysis comparison among the three group patients showed that PVTT patients with thrombocytopenia has longer RFS and OS than those with normal PLT count(both p<0.001)or thrombocytosis(p=0.008 and p=0.046).However,there were no significant difference in prognosis between patients with normal platelet counts and thrombocytosis(p=0.578 and p=0.877).After one-to-one PSM,PVTT subgroup analysis was performed and the results showed thrombocytopenia is only associated with better RFS in patients with type II PVTT(p=0.001),and better OS in those with type I PVTT(p=0.021)or type II PVTT(p=0.029).Moreover,univariate and multivariate analysis showed that PLT level were the independent prognostic factors for overall survival.2.The results of platelet aggregation assay and flow cytometry showed platelet activation in HCC patients with PVTT is higher than it in those without PVTT.And cDNA microarray revealed that different tissues with different expression of mRNA and specific and different-expressed mRNA was detected in PVTT tissues.By the analysis of GO enrichment and KEGG pathways enrichment,the different-expressed genes were involved in series of biology process which include cytolysis,monocarboxylic acid metabolic process,integrin pathway and HNF3 B pathway.3.KITLG(also called stem cell factor,SCF)was selected as candidate mRNA which was specific higher expression in PVTT and CSQT-2 cell lines.The expression of cancer stem cell marker,EpCAM,CD44,CD90 and CD133,are found to agree with the KITLG level.Thrombogenesis experiment in vitro showed that CSQT-2 promotes thrombosis via high expression of KITLG,which can be blocked by platelet antagonist.Conclusions1.Preoperative thrombocytopenia is a positive predictor of survival in HCC patients with PVTT and patients with thrombocytopenia have a longer time of survival than others.2.Platelet activation in HCC patients with PVTT is higher than it in those without PVTT.And there were numerous specific different-expression mRNA in PVTT tissues.3.KITLG and markers of cancer stem cell,EpCAM,CD44 and CD90 was higher in PVTT,and KITLG is a vital mRNA in maintain characters of cancer stem cells.Thrombogenesis experiment in vitro showed that CSQT-2 promotes thrombosis via high expression of KITLG,which can be blocked by platelet antagonist.Potential applications and noveltyThese results showed the close relationship between platelet count and the prognosis of HCC patients with PVTT.High tumour KITLG expression was associated with formation of PVTT by interaction with platelet,which is a new mechanism for PVTT formation.PLT count is a prognostic predictor to identify high risk patients and plays a important role in evaluating the therapeutic strategy for PVTT.