Preparation Of Cd11c Monoclonal Antibodies Modified Immunoliposomes Loading OVA And Its Effects On Dendritic Cells

Dendritic cells (DCs) are professional antigen presenting cells, they are living in the center position for the process of capture antigen, processing, presenting in the immune system, and they can also stimulate the initial and acquired the immune response. Dendritic cells play an important role in anti-infection immunity and anti-tumor immune response, which has become a hot spot of immunology research in recent years. The CD11c is one identified specific surface markers of DCs in the current, it has higher expression in immature DCs(iDCs) and mature DCs(mDCs). Large numbers of studies also confirmed that CD11c can be used as a valid target, mediated endocytosis of antigen, promote DCs specific immune response.As the drug carriers, liposomes are mainly composed of phospholipids. It has a class of cell membrane structure, so integration with cell phospholipid bilayer is easy. Denpending on the structural characteristics of the cell phospholipid bilayer, it has a lot of characteristics, such as carrying a large quantity, protecting the drugs from degradation, easy biofilm fusion, sustained release, reducing the dosage and so on. It is possible to improve the therapeutic index of the drug, changing the distribution of encapsulating drugs in vivo. In many diseases, the treatment shows a significant superiority, so we select the liposomes as the drug carrier. In order to overcome the problems of low transfection efficiency, poor targeting, we have constructed CD11c monoclonal antibody modified immunoliposomes parcel of OVA, CD11c-OVA-imliposomes were constructed with CD11c monoclonal antibody covalently linked to OVA-liposomes. Incorporation of Malermide-PEG2000-distearoylphosphatidylethanolamine into the liposomes material, CD11c monoclonal antibody and2-Iminothiolane hydrochloride(2-IT) combine together make the CD11c mAb thiolysis at the periphery, followed efficient attachement of maleimide-derivatized liposomes with the SH-C11c monoclonal antibody took place under mild conditions.On the base of CD11c-OVA-imliposomes constructed by us, we evaluated the pharmaceutical characterization of liposomes, such as particle size, Zeta potential, encapsulation efficiency and CD11c mAb conjugation efficiency. Particle size, Zeta potential and morphology of OVA-liposomes and CD11c-OVA-imliposomes were observed by dynamic light-scattering detector(DLS) and transmission electron microscope(TEM). The results showed that both liposomes were quiet round, smooth surface, glossy and no adhesion, homogenous with a145～160nm average size distribution. The Zeta potential of OVA-liposomes were (-33.6±1.1)mv, CD11c-OVA-imliposmes were (-31.2±1.7)mv. It could be speculated that CD11c mAb had presented on the OVA-liposomes surface due to the increase in particle size (146nmâ†'159nm) and descend in Zeta potential. High performance liquid chromatography(HPLC) was used to determine encapsulation efficiency of OVA-liposomes and CDllc conjugation efficiency on the OVA-liposomes. The encapsulation efficiency of OVA-liposmes were (32.5±1.9)%and the CD11c conjugation efficiency was about (77.6±3.2)%.In vitro experiment, CD11c-OVA-imliposomes targeting to DCs were studied. The CD11c-OVA-imliposomes targeting ability was quantitative and qualitative detected by flow cytometry(FCM) and laser scanning cofocal microscope(LSCM). The results show that the CD11c-OVA-imliposomes to DCs having a specificity targeting. Measured by flow cytometry of the OVA-liposomes and CD11c-OVA-imliposomes to DCs expression affect through I-Ab, CD40, CD80and CD86. We found that both of them improved expression in varying degrees. CD11c-OVA-imliposmes to promote a higher degree compared to the OVA-liposomes. It shows that CD11c-OVA-imliposomes can be more effective promotion of the phenotype of DCs and promoting it can better play its antigen presenting function.This study successfully prepared and characterization the CD11c-OVA-imliposomes, validation its specific targeting to DCs in vitro. It has proved that CD11c-OVA-imliposomes can effectively promote the maturation of DCs surface(I-Ab, CD40, CD80and CD86) expression and promote mature phenotype of DCs.