Mechanisms of co-modified liver-targeting liposomes as gene delivery carriers based on cellular uptake and antigens inhibition effect.
In order to deliver antisense oligonucleotides (asODN) into hepatocytes orientedly in the treatment of hepatitis B virus (HBV) infection, the liver-targeting cationic liposomes was developed as a gene carrier, which was co-modified with the ligand of the asialoglycoprotein receptor (ASGPR), beta-sitosterol-beta-d-glucoside (sito-G) and the nonionic surfactant, Brij 35. Flow cytometry (FCM) analysis and enzyme-linked immunosorbent assay (ELISA) showed that the asODN-encapsulating cationic liposomes exhibited high transfection efficiency and strong antigens inhibition effect in primary rat hepatocytes and HepG2.2.15 cells, respectively. With the help of several inhibitors acting on different steps during the targeting lipofection, the cellular uptake mechanisms of the co-modified liver-targeting cationic liposomes were investigated through antigens inhibition effect assay and confocal laser scanning microscopy (CLSM) analysis. The cellular uptake with high transfection efficiency seemed to involve both endocytosis and membrane fusion. The ligand sito-G was confirmed to be able to enhance ASGPR-mediated endocytosis, the nonionic surfactant Brij 35 seemed to be able to facilitate membrane fusion, and the co-modification resulted in the most efficient transfection but no enhanced cytotoxicity. These results suggested that the co-modified liver-targeting cationic liposomes would be a specific and effective carrier to transfer asODN into hepatocytes infected with HBV orientedly.
Zhang
Yuan
Y
School of Pharmaceutical Sciences, Peking University, Beijing, China.
Rong Qi
Xian
X
Gao
Yan
Y
Wei
Lai
L
Maitani
Yoshie
Y
Nagai
Tsuneji
T
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001212:Asialoglycoproteins / Q000494:pharmacology; D001692:Biological Transport; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004705:Endocytosis / Q000187:drug effects; D011992:Endosomes / Q000187:drug effects; D060748:Fetuins; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry; D006509:Hepatitis B / Q000628:therapy; D006511:Hepatitis B Antigens / Q000378:metabolism*; D006515:Hepatitis B virus / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism / Q000821:virology; D006801:Humans; D008081:Liposomes / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D008099:Liver / Q000378:metabolism* / Q000473:pathology / Q000821:virology; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D009550:Nigericin / Q000494:pharmacology; D020319:Oligodeoxyribonucleotides, Antisense / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D011092:Polyethylene Glycols / Q000737:chemistry; D051381:Rats; D014162:Transfection / Q000379:methods*; D000077191:Wortmannin; D000509:alpha-Fetoproteins / Q000494:pharmacology
D000730:Androstadienes; D001212:Asialoglycoproteins; D060748:Fetuins; D006511:Hepatitis B Antigens; D008081:Liposomes; D020319:Oligodeoxyribonucleotides, Antisense; D000509:alpha-Fetoproteins; C017029:asialofetuin; D011092:Polyethylene Glycols; D016650:Fluorescein-5-isothiocyanate; C515901:Brij 35; D009550:Nigericin; D000077191:Wortmannin