Acidic Tumor Microenvironment Activates Hepatic Stellate Cells And Promotes Hepatocellular Carcinoma Metastasis

Besides tumor cells,cancer associated stromal cells are also exposed to the acidic tumor microenvironment.But the impact of acidic tumor microenvironment on cancer associated stromal cells was unclear.To investigate potential mechanisms for adapting acidosis,we examined the effect of acidic culture condition(p H6.2)on human hepatic stellate cells(HSC),which frequently infiltrate in the tissue of hepatocellular carcinoma(HCC).Acidic p H exposure enhanced the activation of human hepatic stellate cell line LX-2,which was characterized by elevated expression levels of ?-SMA(actin,alpha 2,smooth muscle,aorta),rearranged cell skeleton,and increased chemotaxis.Meanwhile,the growth of HSC was inhibited significantly because of apoptosis and G0/G1 cycle block induced by the acidic culture condition.An integrative genomic analysis revealed that the canonical HSC activation pathway was stimulated and Erk1/2 might play a key role in the transforming of quiescent HSC to the activated status under acidity stress.The increased phosphorylation level of Erk1/2 under acidity was further confirmed by immunoblotting analysis and the elevated expression level of ?-SMA could be reduced by U0126,an inhibitor of Erk1/2 phosphorylation.Esomeprazole(ESOM),a proton pump inhibitor(PPI),was applied on the mouse xenograft model in which LX-2 cells were co-implanted with HCC cells.Once the acidity of tumor tissue was impaired by ESOM,the activation of HSC and the phosphorylation of Erk1/2 in LX-2 cells both were attenuated.These results indicated that HSC adapted to the acidic microenvironment via Erk1/2-dependent activation which could be attenuated when the acidic tumor microenvironment was impaired by proton pump inhibitor.The migration of HCC cells could also be promoted by the condition medium(CM)collected from the acidity-activated LX-2 cells in vitro.Similarly,the in vivo experiment showed that more distant metastasis occurred when HCC and HSC were co-implanted orthotopically in nude mouse liver.Analysis for CM by cytokine array and enzyme-linked immuno sorbent assay(ELISA)suggested the increase of OPN(Secreted phosphoprotein 1,osteopotin)secretory level might account for the metastasis-promoting effect on HCC from acidity-activated LX-2.When neutralizating antibody of OPN was added into the CM collected from LX-2,promotion of HCC migration was reduced.The above results suggested that OPN derived from the acidity-activated HSC might contributed to HCC metastasis.The analysis of cancerous tissue samples of 49 HCC patients indicated the expression levels of ?-SMA and OPN was positive correlated,and the combination of ?-SMA and OPN was a powerful predictor for poor prognosis of HCC patients.The above results indicated that HSC survived via activation under acidic tumor environment could perform metastasis-promoting function in HCC.Conclusion: In this project,acidic tumor microenvironment was found to enhance HSC activation,which depended on the phosphorylation of Erk1/2.Acidity-activated HSC contributed to HCC metastasis through OPN.Combination of HSC activation and OPN up-regulation indicated poor prognosis for HCC.These results provide a novel clue for understanding the mechanism of acidic tumor environment in the development and progression of HCC.Acidic tumor microenvironment can aggravate hepatic stellate cells activation and promote HCC metastasis,which may be a potential therapeutic strategy for HCC.