title: Synthesis and efficient hepatocyte targeting of galactosylated chitosan as a gene carrier in vitro and in vivo.
abstract: While chitosan (CS) has been researched widely as a non-viral vector, its usefulness has been limited by its low cell specificity and transfection efficiency. Therefore, we successfully synthesized galactosylated chitosan (GC) and complexed it with an enhanced green fluorescent protein plasmid (pIRES-EGFP) for transfection into cultured H22 cells (murine hepatic cancer cell line) using various GC/EGFP (N/P) charge ratios. Maximal gene transfection rates detected by flow cytometry occurred at an N/P ratio 5:1. Compared with those of lipofectin/EGFP and naked pIRES-EGFP, GC/EGFP complexes show a very efficient cell-selective transfection to hepatocytes. The MTT assay detected relatively low cytotoxicity in cells transfected with GC. A recombinant plasmid granulocyte-macrophage colony-stimulating factor (GM-SCF) and interleukin (IL) 21 (pIRES/GM-CSF-IL21) was successfully constructed and GC/GM-CSF-IL21 nanoparticles (average diameter, 82.1 nm) were administered via the tail vein of mice with liver metastasis of colon cancer model, for 5 consecutive days. The GC/GM-CSF-IL21 nanoparticles exhibited hepatocyte and passive tumor specificity, increased therapeutic efficacy compared to control groups, promoted leukocytes to aggregate in tumor tissues, and activated the cytotoxicity of natural killer (NK) cells and cytolytic T lymphocyte (CTL). Our results indicate that GC can be used in gene therapy to improve transfection efficiency and can be used as an immunological stimulant in vivo.
authors.affiliation: Department of General Surgery, Shanghai Fifth People's Hospital, Fudan University, Shanghai, China. cmrlq@126.com
authors.forename: Mingrong
authors.initials: M
authors.lastname: Cheng
authors.forename: Qing
authors.initials: Q
authors.lastname: Li
authors.forename: Tao
authors.initials: T
authors.lastname: Wan
authors.forename: Xiaowu
authors.initials: X
authors.lastname: Hong
authors.forename: Houxiang
authors.initials: H
authors.lastname: Chen
authors.forename: Bing
authors.initials: B
authors.lastname: He
authors.forename: Zhijian
authors.initials: Z
authors.lastname: Cheng
authors.forename: Hongzhi
authors.initials: H
authors.lastname: Xu
authors.forename: Tao
authors.initials: T
authors.lastname: Ye
authors.forename: Bingbing
authors.initials: B
authors.lastname: Zha
authors.forename: Jingbo
authors.initials: J
authors.lastname: Wu
authors.forename: Runjiao
authors.initials: R
authors.lastname: Zhou
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D048271:Chitosan / Q000737:chemistry* / Q000378:metabolism*; D003110:Colonic Neoplasms / Q000473:pathology; D005260:Female; D005690:Galactose / Q000737:chemistry*; D005822:Genetic Vectors / Q000737:chemistry* / Q000378:metabolism*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism*; D007378:Interleukins / Q000235:genetics / Q000378:metabolism; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism / Q000556:secondary / Q000628:therapy; D008422:Materials Testing; D051379:Mice; D008807:Mice, Inbred BALB C; D015394:Molecular Structure; D053758:Nanoparticles; D011897:Random Allocation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017550:Spectroscopy, Fourier Transform Infrared; D014018:Tissue Distribution; D014162:Transfection / Q000379:methods*; D000098546:Interleukin-21
chemical_list: D007378:Interleukins; D011993:Recombinant Fusion Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D048271:Chitosan; D000098546:Interleukin-21; D005690:Galactose