Effect Of Echinococcus Granulosus Cyst Fluid On Host Liver Cell Of Mitogen-activated Protein Kinases And Cell Cycle In Vitro

Objective: To discuss the effect on Mitogen-activated protein kinases and PCNA，cyclin-D1，cyclin-A，cyclin-B1，cyclin-E of Echinococcus granulosus cyst fluid infectedhost cell. Methods: Conventional culture cells, The hydatid cyst fluid taken from cysticechinococcosis of sheep liver at Urumqi slaughterhouse. Collect the hydatid cyst fluidfrom the sheep liver of cystic echinococcosis and filter by filter of0.22um and Stored at-80°C. Conventional culture HepG2cells for24h, When the cells adherent solid changeno FCS culture medium for24h, Cells were divided into three groups after no FCSculture medium: the control group(no FCS group), the text group (containing hydatidcystic fluid without FCS culture medium3.75%HCF group、7.5%HCF group、15%HCFgroup、30%HCF group、60%HCF group）, the normal group (containing10%FCS culturemedium),toxie and proliferation effect on cell by hydatid cyst fluid were asseyed withMTT method after culture cell24h,48h,72h and96h. Cell cycle was detected by Flowcytometry at24h,48h and72h. The same method cultured cells and collected cells at24h,48h,72h and96h. Extract of HepG2cells protein and stored at-80°C, Western-Blot testthe expression of p-ERK, PCNA, cyclin-A, cyclin-B1, cyclin-D1, cyclin-E. Results:MTT text results showed that:15%,30%and60%HCF group promote the proliferationof HepG2cells at48h72h and96h compared to no FCS group, MTT test show15%,30%and60%HCF group compared with the no FCS group at48h:(0.317±0.057the VS0.229±0.066)(P=0.003P <0.01),(0.532±0.041the VS0.229±0.066)(P=0.003P<0.01),(0.600±0.043the VS0.229±0.066)(P=0.004P <0.01), MTT test show15%,30%and60%HCF group compared with the no FCS group at72h:（0.436±0.009VS0.236±0.034）(P=0.039P <0.05)、（0.673±0.070VS0.236±0.034）(P=0.008P <0.05)、 （0.834±0.082VS0.236±0.034）(P=0.001P <0.05)，MTT test show15%,30%and60%HCF group compared with the no FCS group at96h:（0.473±0.003VS0.182±0.045）(P=0.014P <0.05)、（0.771±0.030VS0.182±0.045）(P=0.006P <0.05)、（0.834±0.082VS1.008±0.027）(P=0.001P <0.05)，the difference was statisticallysignificant (P <0.05). Flow cytometry show the group15%HCF S phase of content are1.64times，0.81times and0.83times than no FCS in24h48h and72h. The group30%HCFS phase of content are1.64times,0.81times and0.38times than no FCS at24h,48h, and72h. he western-Blot show the group15%HCF and30%HCF have high expression ofp-ERK in48h and24h (P=0.0003、0.0159，P <0.05), the group15%HCF and30%HCFhave high expression of PCNA in48h and72h(P=0.0002、0.0010，P <0.01), the group15%HCF and30%HCF have high expression of cyclin-D1in24h(P=0.0004、0.0024，P<0.01), the group15%HCF and30%HCF have high expression of cyclin-A in48h and72h(P=0.0150、0.0001，P <0.05), the group15%HCF have high expression of cyclin-B1in48h and72h(P=0.0009、0.0025，P <0.01), the group15%HCF have high expressionof cyclin-E in72h(P=0.0470，P <0.05). Conclusion: The hydatid cyst fluid had noinhibition effect on primary cultured host cell, but it could promote the proliferation ofhost cell.