The Effection Of Ultrasound Mediated LHRHa-targeting Paclitaxel Loaded Lipid Microbubble On The Proliferation And Apoptosis Of Ovarian Cancer A2780/DDP Cells

PREPARATION AND OBSERVATION OFTARGETING ABILITY OF LHRHa-TARGETEDPACLITAXEL LOADED LIPID MICROBUBBLESObjective To investigate the reliability of coupling paclitaxel loadedmicrobubble with LHRHa ligand by biotin-streptavidin bridge methodand its targeting ability in vitro.Methods The biotinylated luteinizing hormone releasing hormoneanalog (LHRHa) was attached to the surface of biotinylated paclitaxelloaded microbubble via streptavidin. Paclitaxel lipid microbubbles coatedwith LHRHa was detected by immunofluorescent staining, the bindingefficiency of the ligands was quantified via flow cytometry, the targetingability of the microbubbles was observed by light microscopy.Results LHRHa targeted paclitaxel lipid microbubbles were positive inimmunofluorescent staining assay.（99.12±1.45）%of PLM-B wascombinated with FITC-streptavidin, the binding efficiency of PLMT was(87.33±2.19)%, and it was much higher than that of PLM-BS andPLM-B (P<0.05); In vitro research of the targeting ability indicated that theLHRHa paclitaxel lipid microbubbles could adhere to the A2780/DDP cellsspecifically.Conclusion The LHRHa peptide was successfully linked to thepaclitaxel lipid microbubble surface through biotin-avidin linkage, and thistargeted microbubble could bind to human ovarian cancer cells speciallyand effectively in vitro. THE EFFECTION OF ULTRASOUND MEDIATEDLHRHa-TARGETING PACLITAXEL LOADED LIPIDMICROBUBBLE ON THE PROLIFERATION ANDAPOPTOSIS OF OVARIAN CANCER A2780/DDP CELLSObjective To investigate the effection of ultrasound mediatedLHRHa-targeting paclitexal loaded microbubbles on the proliferation andapoptosis of A2780/DDP ovarian cancer cells.Methods Cultured A2780/DDP cells were divided into seven groups,which include paclitaxel, paclitaxel plus ultround irradiation, paclitaxelloaded microbubbles plus ultrasound irradition, paclitaxel loaded microbubbles, LHRHa-targeting paclitaxel loaded microbubbles,LHRHa-targeting paclitaxel loaded microbubbles plus ultrasoundirradiation and blank control groups. After treatment, the ratio ofproliferation inhibition was detected by MTT. The cell apoptosis and cellcycle was determined by flow cytometry. Cell ultrastructure was observedby transmission electron microscope and the expression of cyclinB1andcaspase-3was analysed by western blot.Results: The treatment of paclitaxel alone, paclitaxel plus US,PLMplus US, PLMT plus US had significant effects on A2780/DDP cellproliferation and apoptosis. However, among the four groups, the effects ofUS combined PLMT group was the strongest(P＜0.05). And PLM andPLMT alone had no significant effects on cells. The cell inhibitory rate inthe PLMT plus US at24h,48h,72h was (41.30±3.93)%，（67.76±2.45）%and (75.93±2.81）%respectively，which was significantly higher thanother groups at the same time(P＜0.05). And the apoptosis rate of thisgroup determined by flow cytomety was（32.62±0.79）%, which wassignificantly higher than other groups (P＜0.05). The results of flowcytomety showed that the proportion of G2/M phase was significantlyhigher and S phase was lower in paclitaxel, paclitaxel plus US, PLM plusUS, PLMT plus US groups than control group24h after treatment, and theproportion of G2/M phase in the group of PLMT plus US was the most significant(P＜0.05). Apoptosis bodies were observed by transmissionelectron microscope in the PLMT plus US group. The group of PLM plusUS irradiation have the multinucleated cells.The cell cytoplasm appearedmuch of the microfilament and microtubules in the taxol group, and mitosisblock cells were obvious in the paclitaxel combined with US.The cells didnot have morphologic change in PLM and PLMT groups.The results ofwestern blot showed that the expression of cyclinB1and caspase-3proteinwas significantly higher in paclitaxel, paclitaxel plus US, PLM plus US,PLMT plus US groups than control group24h after treatment, and theexpression of cyclinB1and caspase-3protein was more significantly higherin the PLMT plus US group than that in other groups (P＜0.05).Conclusion: Our data provide evidence that PLMT combined with USinhibite proliferation and induce apoptosis of A2780/DDP ovarian cancercells. The mechanism may be related to paclitaxel up-regulation of cyclinB1and caspase-3protein expression, block the cell cycle in the G2/M period.