| As a major disease threatening human health,tumors have high morbidity and mortality.It is very important to study the effective early diagnosis,ideal therapeutic target and reasonable prognosis detection based on the mechanism of tumorigenesis.Tumor growth,invasion,and metastasis depend on the interaction of cancer cells and/or mesenchymal cells in the tumor microenvironment.The extracellular matrix components,active factors and extracellular vesicles secreted by these cells are involved in the process of inflammation,vascular lymphogenesis,cell proliferation,cell metastasis,immunosuppression,epithelial-mesenchymal transition,and are potential markers of tumors,it is also a rich source of potential tumor markers and therapeutic drug targets.Hepatocellular carcinoma is the final result of interaction of chronic inflammation,angiogenesis and microenvironment.It is of scientific significance and clinical value to explore the role of secretory components in microenvironment in the development of hepatocellular carcinomaThe role of liver interstitial cells-hepatic stellate cells,and liver parenchymal tumor cells in the microenvironment,that is,the effects of secreted proteins,exosomes released by these cells on tumors are not completely clear.Based on the proteomic technology to determine the composition of its secreted products,this study explored the effects of secreted proteins and exosomes on tumor cell proliferation and migration,and its role in tumor chemotherapy drug resistance,and to characterize the difference in protein expression after induction.Annotate and analyze differential proteins through bioinformatics and online databases,provides basic research data and references for revealing the dynamic composition of tumor microenvironment and its molecular mechanisms in tumorigenesis.In this study,human hepatoma cell line Huh7,SMMC-7721 cells and activated hepatic stellate cell line LX-2 were used to extract secreted proteins by ultrafiltration,ultracentrifugation and kits to extract exosomes;Based on the identification of exosomes by electron microscope and Western blot,the secretory protein and protein composition of exosomes were characterized by two-dimensional electrophoresis.The cell viability was observed by CCK-8 method,the cell migration ability was observed by scratch test and Transwell chamber method,and the effects of secreted proteins and exosomes on tumor cells and the role of sorafenib resistance were explored.2D was used to characterize the differentially expressed proteins of secreted proteins and exosomes affecting tumor cells,and identified by MALDI-TOF/TOF mass spectrometry.Finally,the relationship between secretory proteins,exosome proteins and tumor differential proteins was analyzed by bioinformatics and online database TCGA and GEO.The results showed that secreted proteins and exosomes derived from Huh7,SMMC-7721 and LX-2 cells had effects on the proliferation and migration of tumor cells(P<0.05),and the exosomes from tumor cells promoted the drug resistance of Huh7 and SMMC-7721 cells to sorafenil(P<0.05).By proteomic characterization,the secretory proteins and exosomes derived from Huh7,SMMC-7721 and LX-2 had different protein expression profiles,and most of the secretory proteins originated from exosomes.Different methods for extracting exosome expression profiles have differences.The LX-2 secreted protein participates in the stress response together with the differential protein of tumor cell expression caused by the secreted protein.Some proteins were involved in NF-kB pathway and endoplasmic reticulum protein process and correlated with tumorigenesis and poor prognosis(P<0.05).Exosome in microenvironment and its intervention in differentially expressed proteins of tumor cells participate in a variety of biological processes,and some proteins are significantly associated with tumorigenesis and poor prognosis(P<0.01),and involved in tumor cell resistance to sorafenib.The conclusions of this study are as follows: the expression profiles of secretory proteins and exosomes of activated hepatic stellate cells in tumor microenvironment are different from those of tumor cells,and most of the secreted proteins are derived from exosomes;Secreted proteins and exosomes in tumor microenvironment have a some influence on the proliferation and migration of tumors,as well as the resistance of tumor-derived exosomes to tumors,and can change the expression level of tumor cell proteins.Differential proteins are involved in some stress pathways,such as endoplasmic reticulum protein imbalance,glycolysis,and PI3K-Akt signaling pathway,Rho-dependent cellular motor signaling pathways to promote tumor proliferation and migration;RAN,CAP1,HSPA5,ANXA5,DNM1 L PSMA6,PYGB,and SGTA proteins were significantly associated with the occurrence and poor prognosis of hepatocellular carcinoma;ANXA5,YWHAZ,and proteasome were associated with sorafenib resistance. |
PDF Full Text Request