Effect Of Antibacterial-immuiiomodulating Fusion Peptide On Proliferation And Apoptosis

Objective To study the effect of antibacterial-immunomodulating fusion peptide(AIFP) on human ovarian cancer cell proliferation and apoptosis. And research onits antibacterial activity in vitro.Methods The purity of primary cultured cells by Immunofluorescence staining.The ovarian cancer cell proliferation of both the SKOV3and primary treadingwith AIFP in vitro were assayed by MTT method. Apoptotic ratios and cell cycleof these cells, were measured by flow cytometry(FCM) in vitro. Expression ofmRNA and protein in the detection of Akt, CDC25C, cyclinB1bcl-xl and baxgene of PCR and Western blot. Antibacterial activity of AIFP were tested by paperagar diffusion method.Results The result of immunofluorescence showed that the purity of primaryovarian cancer cells reached to84.61%, can be subjected to further analysis.Compared with the control group, MTT assay showed that the ID50were2.75×10-4mg/ml,3.96×10-5mg/ml and3.96×10-6mg/ml after24,48and72htreated with AIFP. Flow cytometry results showed that AIFP at the concentrationof5×10-3mg/ml, after treated with24,48and72hours, the apoptosis rate ofprimary ovarian cancer cells were6.10%,19.31%and40.63%.The same time,the apoptosis rate of SKOV3were2.82%for7.82%, and29.81%, compared withthe negative control group with statistically significant differences (p<0.05).When the concentration of AIFP is5×10-3mg/ml,G2/M phase wereextended（7.15%、8.81%、12.00%）compared with blank control group（2.32%、 2.93%、3.02%）. When the concentration of AIFP is5×10-6mg/ml,G2/M phasewere extended（5.25%、6.90%、8.14%）compared with blank control group（2.32%、2.93%、3.02%）.When the concentration of AIFP were5×10-3mg/ml、5×10-6mg/ml and5×10-9mg/ml, the expression levels of Akt, CDC25C, cyclinB1,BcL-xl were reduced (p <0.05) after treated with AIFP24,48and72hours.Andthe expression of Bax were induced. In western blot experiment,expression ofAkt, CDC25C, cyclinB1, BcL-xl were reduced (p <0.05) after treated with AIFP24,48and72hours. But Bax has the opposite result.The growth curve of E. Coliin AIFP treated group located below the control group, which suggested that thegrowth curve of E. Coli in AIFP treated group were inhibition. Paper agardiffusion experiments suggested that AIFP group showed activity.and thediameter of antibacterial ring were less than ampicillin group(p <0.05), butsignificantly higher than the control group(p <0.05).Conclusion1、we have cultured primary ovarian cancer cell sccessfully, itspurity can be used to the next experiment after being identified byImmunofluorescence staining.2、AIFP able to inhibit the proliferation of primary ovarian cancer cell andSKOV3cell.3、AIFP can promote apoptosis in ovarian cancer cells.4、AIFP can prevent the cycle of ovarian cancer cell in the G2/M phase.5、AIFP can down-regulate the expressions of Akt，CDC25C, cyclinB1andBcl-xl, and up-regulation the expression of bax,which may affect ovarian cancercell proliferation and apoptosis.6、 AIFP have antibacterial activity to E. Coli in vitro.