Application of Chitosan-Based Nano-particles for Disease Diagnosis and Gene Transfection
Date Issued
2010
Date
2010
Author(s)
Yang, Shu-Jyuan
Abstract
Colorectal cancer is one of the leading causes of malignant death in Taiwan because it often remains undetected until later stages of the disease. Improved methods of detecting dysplasia and tumors during colonoscopy will improve mortality. In this study, a chitosan based nano-particle was successfully prepared and loaded with 5-aminolaevulinic acid (5-ALA) that could be taken up by colorectal cancer cells (HT29 and Caco-2) but escape from being engulfed by E. coli. that might lead to misinterpretation of endoscopic examination in clinical. The z-average diameter of the chitosan based nano-particles was about 100 nm, and the zeta-potential was higher than 20 mV that provided enough zeta-potential to prevent aggregation. Furthermore, folic acid can be covalently conjugated to chitosan molecules via its γ-carboxyl moiety and thus retain a high affinity for colorectal cancer cells bearing the folate receptor over-expression. The chitosan nano-particle conjugated with folic acid could be significantly taken up by HT29 and Caco-2 cell lines, most likely via receptor-mediated endocytosis, and enhance the protoporphyrin IX (PpIX) accumulation in cytoplasma after a short-term uptake period. When alginate was physically incorporated into the
chitosan based nano-particles and then fed to colorectal cancer cells, the fluorescent intensity of PpIX in cells could be remarkably improved by competing with 5-ALA for
chitosan in cellular lysosomes, resulting in a higher amount of 5-ALA release and PpIX accumulation. Therefore, the chitosan nano-particle conjugated with folic acid and incorporated with alginate appears to be an ideal vector for colorectal-specific drug delivery of 5-ALA for fluorescent endoscopic detection of colorectal cancer.
Moreover, gene therapy has been used to treat a variety of health problems. However, inefficiency of transfection and the lack of safe gene vectors have limited progress. Fabrication of a vector that is safe and has high transfection efficiency is crucial for the development of successful gene therapies. In this study, we also demonstrated the successful preparation of chitosan based nano-particles that were complexed with the pAcGFP1-C1 plasmid. The prepared nano-particles protected the DNA from enzymatic digestion and improved the efficiency of gene transfection of HeLa cells and 293T cells. The efficiency of gene transfection was further enhanced by exposure to an ultrasound (US) regimen either for in vitro or in vivo tests. Although US reduced cell viability, the simultaneous use of gene transfection and tumor destruction by a focused US may have advantages for the gene therapy of cancer. We conclude that the combined use of DNA-complexed chitosan based nano-particles and US provides a high efficiency gene transfection system for human cancer cells and shows promise for use in future clinical gene therapy.
Subjects
Colorectal cancer
chitosan
5-aminolaevulinic acid
protoporphyrin IX
SDGs
Type
thesis
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