The Experimental Study About Human Hepatoma Cell Proliferation, Invasionand Metastasis And Apoptosis Effects And Significance Which Bortezomib Play On

Hepatocellular carcinoma (primary liver cancer,referred to as liver cancer) is theworld’s fifth largest solid tumors and the third leading cause of cancer deaths.so far,surgical resection and liver transplantation still is the only treatment for early HCC.Surgeryis ineffective for the most advanced liver cancer due to metastasis.Liver cancer patientswho is in stage C or D of BCLC staging often use chemotherapy treatment.However, theresults of these traditional chemotherapy for patients who have been advanced is poor,easily prone to chemotherapeutic drugs multidrug resistance (MDR) decreasing thetherapeutic effect. Therefore seeking new molecular targeted therapies become imperativefor HCC patients.Molecular targeted drugs Bortezomib are peptide boronic acid compound,which has high selectivity, small side effects and is the only protease inhibitor applying incancer clinical therapy.Bortezomib which is formerly known as Bortezomib bind toproteasome20S subunit directly and block the enzyme activity,influencing the degradationof intracellular proteins in a plurality of cycles and inducing apoptosis.Bortezomib is thefirst proteasome inhibitor using in clinical,which has been successfully used in thetreatment of relapsed and refractory multiple myeloma and was proved to be effective inthe treatment of I and Phase II clinical trials colorectal cancer,etc. tumors. In the livercancer treatment study, it’s reported that large doses of proteasome inhibitor Bortezomibcan directly induce apoptosis of hepatoma cells.However,the research of these specificmechanisms is still little. This study about culturing five kinds of human hepatoma cell linethat detecte the roles of proliferation and apoptosis which bortezomib play on and itsmechanism. Part1: The roles of proliferation, invasion and metastasis inhibitionwhich bortezomib play on human hepatoma cellsObjective: By detecting the proliferation,invasion and metastasis of Bortezomib infive different kinds of liver cancer cells,which is to explore the correlation betweenubiquitin-proteasome inhibitor Bortezomib and biological behavior for instance cellproliferation, invasion and metastasis of liver cancer cells which bortezomib play on invitro.Methods: By culturing high metastatic liver culture MHCC97-H and M3cell lines,low metastatic HepG2, MHCC97-L and Huh7cell lines.MTT assay was used to detect cellinhibition rate under different concentrations of proteasome inhibitor Bortezomib’s action.Scratch Transwell experiments and experimental methods was used to explore five kindsof liver cancer cell’s growth, adhesion, invasion and migration capabilities.Results: Five hepatoma cell lines which are in logarithmic growth phase were treatedwith different concentrations of Bortezomib for24h,48h,72h after detectionrespectively.MTT results are showed: Bortezomib can inhibit the proliferation of fivehepatoma cell lines compared with the control group, the difference is significantstatistically(P<0.05);The inhibition rate is enhanced with increasing dose and duration ofinhibition. Scratch experiments and Transwell experiments are showed:the invasion andmigration rate of the experimental group cell was significantly lower(P<0.01.)comparedwith the control group;Bortezomib inhibite the proliferation and migration of five humanhepatoma cell lines whch is concentration-dependent.Conclusions:Bortezomib inhibits the proliferation of five hepatoma cell lines andnegatively regulate the invasion and metastasis of hepatocellular carcinoma cell lines,showing a certain time and dose-dependent manner. Part2: Explore the mechanism of Bortezomib how to inhibite theproliferation and promote apoptosis of the hepatocellular carcinomacellsObjective: By studying apoptosis states and the expression of XIAP and Survivinprotein of liver cancer cell lines under the action of Bortezomib cells, which is to explorethe mechanism that Bortezomib how to inhibits the proliferation and promote apoptosis ofhepatoma cells.Methods:Annexin V-FITC and PI double staining is used to detect cells’s apoptosis,which is to explore the effects of Bortezomib.Western blotting is used to detect theexpression of apoptosis-related proteins XIAP and Survivin and to explore howBortezomib play a pro-apoptotic role.Results: Streaming result suggests that: Bortezomib can increase the apoptosis rate ofliver cancer significantly,compared with the control group and the difference wassignificant statistically (P <0.05). Western Blotting result suggests that Bortezomib reducesthe expression of inhibitor of apoptosis protein XIAP, Survivin expression in hepatoma celllines.The difference was significant statistically compared with the control group(P <0.05).Conclusions:Bortezomib exert anti-tumor effect by regulating the channel ofapoptosis, however the main anti-tumor function which is achieved through inhibiting theexpression of pro-apoptotic protein,may be related to the ubiquitin-proteasome pathway.