Dhaq-gm-csf, Immunoliposome Studies, The Killing Effect Of Hl60 Cells

Objective To prepare DHAQ-GM-CSF immunoliposomes and observe the cytotoxicity of HL60 cells treated with the immunoliposomes for providing the preclinical data for AML therapy. Method DHAQ liposomes were prepared by reverse-phase evaporation(REV), high speed centrifugation was applied to separate the DHAQ liposomes and dissociative DHAQ, colorimetry was employed to determine encapsulation efficiency, the liposomes' structure and particle size were determine by transmission electron microscopy. GM-CSF was coupled to the DHAQ liposomes by glutaraldhyde method. Gradient centrifugation was employed to separate the DHAQ-GM-CSF immunoliposomes, UV-spectrophotometric analysis was applied to mensurate the coupled efficiency, flow cytometry analysis was applied to determine the immunoconjugate retained efficiency. The cytotoxicity of HL60 cells and targetability of GM-CSF were investigated by tetrazolium microculture(MTT) cellular cytotoxicity test. The rest data were collected from transmission electron microscopy, flow cytometric analysis. Results 1: The encapsulation efficiency of DHAQ liposomes was 81%; the liposomes were monolayer liposomes in the most, and the particle size was 170-220nm, the coupled efficiency was 41.7%, the immunoconjugate retained efficiency was 75.1%. 2: DHAQ-GM-CSF immunoliposomes, DHAQ liposomes and DHAQ all puted up cytotoxicity of HL60, the cytotoxicity of DHAQ-GM-CSF immunoliposomes and DHAQ liposomes were all better than DHAQ (p<0.05) , and the cytotoxicity of DHAQ-GM-CSF immunoliposomes was better than DHAQ liposomes(p<0.05). After treated with DHAQ-GM-CSF immunoliposomes, DHAQ liposomes and DHAQ for 24h, the IC50 values were3.04μg/ml,9.33μg/ml,18.15μg/ml and after 48h the IC50 were 0.87μg/ml,3.02μg/ml,6.69μg/ml. The inhibition rate increased in the dose-dependent manner. 3: The cytotoxicity of DHAQ-GM-CSF immunoliposomes could be weakened by GM-CSF. 4: The apoptotic rate of HL60 cells of DHAQ-GM-CSF immunoliposomes group was higher than that of DHAQ liposomes and DHAQ groups. 5: The cellar ultrastructure was markedly destroyed. Conclusions 1: The methods applied in this study were equal to the preparation of DHAQ-GM-CSF immunoliposomes. the encapsulation efficiency, the coupled efficiency and the immunoconjugate retained efficiency were so satisfying. 2: In vitro, DHAQ-GM-CSF immunoliposomes have cytotoxicity of HL60 cells and could induce HL60 apoptosis. 3: Because the IC50 of DHAQ-GM-CSF immunoliposomes was lower than that of DHAQ, so DHAQ-GM-CSF immunoliposomes could improve the curative effect and reduce the toxicity of DHAQ. 4: The cytotoxicity of DHAQ-GM-CSF immunoliposomes have tagetability compared with DHAQ liposomes.