Mechanistic Study Of The Role Of Protein Tyrosine Phosphatase Receptor-type O Involved In The Inflammatory Microenvironment Of Hepatocellular Carcinoma

1.Estrogen sensitive PTPRO expression represses hepatocellular carcinoma progression by control of STAT3Protein tyrosine phosphatase receptor-type O(PTPRO)was recently described as a tumor suppressor in various types of cancers,but the detailed function and mechanism remained unknown.Notably,hepatocellular carcinoma(HCC)generation and progression exhibit significant gender disparity and inflammatory characteristics,which gravely threat the health of Chinese population.In Study I,we aimed to clarify the role of PTPRO in HCC parenchyma.It has been demonstrated in 180 pairs(120 male and 60 female)of clinical HCC specimens that PTPRO level was significantly reduced as compared to adjacent tissues,and PTPRO level in male adjacent tissue was lower than female.Given the fact of pathological deficiency and gender bias of PTPRO expression in HCC,we further uncovered that estrogen receptor(ER?)could up-regulate PTPRO expression as a transcription factor.Moreover,in vitro study showed that cell proliferation was inhibited and apoptosis was promoted in PTPRO transduced HCC cell lines.In an inflammation-driven HCC model,ptpro-/-mice represented obviously increased tumor number and size.As was responsive for the tumor suppressive position,PTPRO was proved to down-regulate(signal transducer and activator of transcription 3)STAT3 activity dependent of(Janus kinase 2)JAK2 and(phosphoinositide 3-kinase)PI3K dephosphorylation.Interestingly,PTPRO mechanistically contributed to c-Src activation as a result of its dephosphorylation.Thus,the suppressive role of PTPRO in HCC could be ascribed to inactivation of STAT3,which dominates the tumor-promoting inflammatory signaling.2.PTPRO plays a dual role in hepatic ischemia reperfusion injury through feedback activation of NF-?BSince PTPRO were proved to suppress HCC progression,we got to be curious about its role in benign liver tissue.It is critical if it also inhibits the survival of normal hepatocytes,which may contribute to liver failure undergoing liver injury.Partial liver resection is the principle therapy for early-stage human HCC,during which hepatic ischemia reperfusion(IR)injury is frequent and inevitable.Nuclear factor-?B(NF-?B)activation in hepatocytes and macrophages appears as a double-edged-sword in IR injury,which is also important in inflammation and tumorigenesis.Based on our recent data,PTPRO was a potential activator of c-Src,which can in turn activate NF-?B pathway.In Study II,we aimed to determine the change and function of PTPRO in the acute liver injury during IR.Clinical patients with benign liver condition undergoing liver surgery were recruited in our study.Wild type(WT)and ptpro-/-C57BL/6 mice were processed to construct hepatic IR models.Isolated mouse hepatocytes and macrophages were treated with peroxide or TNF? in vitro.In human and mouse IR models,PTPRO level was decreased in the early phase but reversed in the late phase.In vitro studies demonstrated that NF-?B up-regulated PTPRO transcription.Using ptpro-/-mice and primary cells,we found that PTPRO deficiency resulted in reduction of NF-?B activation in both hepatocytes and macrophages and was correlated to c-Src phosphorylation;precisely,PTPRO in hepatocytes alleviated,but PTPROt in macrophages exacerbated IR injury.Thus,it has been demonstrated PTPRO activates NF-?B in a positive feedback manner,and plays a dual role in hepatic IR injury.3.PTPROt maintains T cell immunity in the microenvironment of hepatocellular carcinomaIntratumoral T cells play a central role in anti-tumor immunity,and the balance between T effector cells(Teff)and regulatory T cells(Treg)affects the prognosis of cancer patients.However,educated by tumor microenvironment,T cells frequently fail in their responsibility.Since we knew truncated isoform of PTPRO(PTPROt)was expressed in immune cells and contributed to acute liver inflammation,we therefore turned to investigate its role in T cell mediated anti-tumor immunity.In Study III,we recruited 70 HCC patients and 30 healthy volunteers for clinical investigation,and analyzed cellular tumor immunity by using ptpro-/-C57BL/6 mice and NOD/SCID mice.As it turned out,PTPROt expression was significantly down-regulated in human HCC infiltrating T cells due to the hypoxia microenvironment and inhibition by hypoxia-inducible factor-1?(HIF-1?);PTPROt expression highly correlated with the intratumoral Teff/Treg ratio and clinicopathologic characteristics.Moreover,PTPROt deficiency attenuated T cell mediated anti-tumor immunity and remarkably promoted mouse HCC growth.Mechanistically,deletion of PTPROt decreased Teff quantity and quality through phosphorylation of lymphocyte specific tyrosine kinase(Lck),but increased Treg differentiation through phosphorylation of signal transducer and activator of transcription(STAT)5.In support of the Teff/Treg homeostasis,PTPROt serves as an important tumor suppressor in HCC microenvironment.Conclusions: we have identified PTPRO as a multi-pronged tumor suppressor in HCC,whose expression not only inhibits HCC progression in both tumor cells and tumor microenvironment,but also protects the normal liver function against inflammatory injury or stress.