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TAT-LHRH conjugated low molecular weight chitosan as a gene carrier specific for hepatocellular carcinoma cells.

Liu L, Dong X, Zhu D, Song L, Zhang H, Leng XG - Int J Nanomedicine (2014)

Bottom Line: To develop a chitosan-based nonviral gene carrier capable of delivering genes specifically into hepatoma cells, a bifunctional peptide composed of the TAT (transactivator of transcription) peptide and luteinizing hormone-releasing hormone (LHRH) was conjugated with low molecular weight chitosan, resulting in a TAT-LHRH-chitosan conjugate (TLC).In vitro targeting specificity and transfection efficiency were analyzed with a GE IN Cell Analyzer 2000 High-Content Cellular Analysis System.The results demonstrated that TLC had stronger DNA condensing power than unmodified chitosan, and that TLCDNPs were of roughly round shape with average diameter of 70-85 nm and zeta potential of +30 mV and were relatively stable in solution.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Bioengineering, Institute of Biomedical Engineering, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin Key Laboratory of Biomedical Materials, Tianjin, People's Republic of China.

ABSTRACT
To develop a chitosan-based nonviral gene carrier capable of delivering genes specifically into hepatoma cells, a bifunctional peptide composed of the TAT (transactivator of transcription) peptide and luteinizing hormone-releasing hormone (LHRH) was conjugated with low molecular weight chitosan, resulting in a TAT-LHRH-chitosan conjugate (TLC). TLC/DNA nanoparticles (TLCDNPs) were characterized by agarose gel retardation, atomic force microscopy, and dynamic light scattering analysis. In vitro targeting specificity and transfection efficiency were analyzed with a GE IN Cell Analyzer 2000 High-Content Cellular Analysis System. The results demonstrated that TLC had stronger DNA condensing power than unmodified chitosan, and that TLCDNPs were of roughly round shape with average diameter of 70-85 nm and zeta potential of +30 mV and were relatively stable in solution. The in vitro study demonstrated TLC was highly selective for hepatoma cells and essentially nontoxic.

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Morphologic observation of TAT-LHRH-chitosan/DNA complexes at different N/P ratios with an atomic force microscope; (A) (N/P) 2:1, (B) 4:1, (C) 10:1.Abbreviations: N/P, free molar ratio of NH2/PO4; TAT-LHRH, transactivator of transcription – luteinizing hormone-releasing hormone.
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f7-ijn-9-2879: Morphologic observation of TAT-LHRH-chitosan/DNA complexes at different N/P ratios with an atomic force microscope; (A) (N/P) 2:1, (B) 4:1, (C) 10:1.Abbreviations: N/P, free molar ratio of NH2/PO4; TAT-LHRH, transactivator of transcription – luteinizing hormone-releasing hormone.

Mentions: Agarose gel retardation was performed to analyze the DNA-entrapping capability of TCL and it showed that the DNA was completely entrapped in TLC at an N/P ratio higher than 1:1 (Figure 5), while the DNA was completely entrapped by the unmodified chitosan only when the N/P ratio reached 8:1 or beyond, indicating the stronger DNA-entrapping capability of TLC in comparison with the unmodified chitosan. As shown in Figures 6 and 7, the average diameter of the TLCDNPs was 70–87 nm, the polydispersity index was about 0.12–0.4, and the zeta potential was about +30 mV as demonstrated by Zetasizer Nano ZS (Malvern Instruments), with roughly round and relatively uniformed shape as characterized with an AFM. The current study also investigated the stability of TLCDNPs in water and it demonstrated that the size and zeta potential of TLCDNPs were relatively stable during the 2 weeks of observation (Figure 8).


TAT-LHRH conjugated low molecular weight chitosan as a gene carrier specific for hepatocellular carcinoma cells.

Liu L, Dong X, Zhu D, Song L, Zhang H, Leng XG - Int J Nanomedicine (2014)

Morphologic observation of TAT-LHRH-chitosan/DNA complexes at different N/P ratios with an atomic force microscope; (A) (N/P) 2:1, (B) 4:1, (C) 10:1.Abbreviations: N/P, free molar ratio of NH2/PO4; TAT-LHRH, transactivator of transcription – luteinizing hormone-releasing hormone.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4061174&req=5

f7-ijn-9-2879: Morphologic observation of TAT-LHRH-chitosan/DNA complexes at different N/P ratios with an atomic force microscope; (A) (N/P) 2:1, (B) 4:1, (C) 10:1.Abbreviations: N/P, free molar ratio of NH2/PO4; TAT-LHRH, transactivator of transcription – luteinizing hormone-releasing hormone.
Mentions: Agarose gel retardation was performed to analyze the DNA-entrapping capability of TCL and it showed that the DNA was completely entrapped in TLC at an N/P ratio higher than 1:1 (Figure 5), while the DNA was completely entrapped by the unmodified chitosan only when the N/P ratio reached 8:1 or beyond, indicating the stronger DNA-entrapping capability of TLC in comparison with the unmodified chitosan. As shown in Figures 6 and 7, the average diameter of the TLCDNPs was 70–87 nm, the polydispersity index was about 0.12–0.4, and the zeta potential was about +30 mV as demonstrated by Zetasizer Nano ZS (Malvern Instruments), with roughly round and relatively uniformed shape as characterized with an AFM. The current study also investigated the stability of TLCDNPs in water and it demonstrated that the size and zeta potential of TLCDNPs were relatively stable during the 2 weeks of observation (Figure 8).

Bottom Line: To develop a chitosan-based nonviral gene carrier capable of delivering genes specifically into hepatoma cells, a bifunctional peptide composed of the TAT (transactivator of transcription) peptide and luteinizing hormone-releasing hormone (LHRH) was conjugated with low molecular weight chitosan, resulting in a TAT-LHRH-chitosan conjugate (TLC).In vitro targeting specificity and transfection efficiency were analyzed with a GE IN Cell Analyzer 2000 High-Content Cellular Analysis System.The results demonstrated that TLC had stronger DNA condensing power than unmodified chitosan, and that TLCDNPs were of roughly round shape with average diameter of 70-85 nm and zeta potential of +30 mV and were relatively stable in solution.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Bioengineering, Institute of Biomedical Engineering, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin Key Laboratory of Biomedical Materials, Tianjin, People's Republic of China.

ABSTRACT
To develop a chitosan-based nonviral gene carrier capable of delivering genes specifically into hepatoma cells, a bifunctional peptide composed of the TAT (transactivator of transcription) peptide and luteinizing hormone-releasing hormone (LHRH) was conjugated with low molecular weight chitosan, resulting in a TAT-LHRH-chitosan conjugate (TLC). TLC/DNA nanoparticles (TLCDNPs) were characterized by agarose gel retardation, atomic force microscopy, and dynamic light scattering analysis. In vitro targeting specificity and transfection efficiency were analyzed with a GE IN Cell Analyzer 2000 High-Content Cellular Analysis System. The results demonstrated that TLC had stronger DNA condensing power than unmodified chitosan, and that TLCDNPs were of roughly round shape with average diameter of 70-85 nm and zeta potential of +30 mV and were relatively stable in solution. The in vitro study demonstrated TLC was highly selective for hepatoma cells and essentially nontoxic.

Show MeSH
Related in: MedlinePlus