RESUMO
50 kDa chitosan was conjugated with folate, a specific tissue-targeting ligand. Nanoparticles such as chitosan-DNA and folate-chitosan-DNA were prepared by coacervation process. The hydrodynamic intravenous injection of nanoparticles was performed in the right posterior paw in normal and arthritic rats. Our results demonstrated that the fluorescence intensity of DsRed detected was 5 to 12 times more in the right soleus muscle and in the right gastro muscle than other tissue sections. ß-galactosidase gene expression with X-gal substrate and folate-chitosan-plasmid nanoparticles showed best coloration in the soleus muscle. Treated arthritic animals also showed a significant decrease in paw swelling and IL-1ß and PGE2 concentration in serum compared to untreated rats. This study demonstrated that a nonviral gene therapeutic approach using hydrodynamic delivery could help transfect more efficiently folate-chitosan-DNA nanoparticles in vitro/in vivo and could decrease inflammation in arthritic rats.
Assuntos
Artrite Experimental/terapia , Quitosana/administração & dosagem , DNA/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Ácido Fólico/administração & dosagem , Nanopartículas/administração & dosagem , Análise de Variância , Animais , Artrite Experimental/genética , Artrite Experimental/metabolismo , DNA/genética , Dinoprostona/metabolismo , Modelos Animais de Doenças , Feminino , Adjuvante de Freund/administração & dosagem , Histocitoquímica , Humanos , Injeções Intravenosas , Proteína Antagonista do Receptor de Interleucina 1/genética , Proteína Antagonista do Receptor de Interleucina 1/metabolismo , Interleucina-1beta/metabolismo , Músculo Esquelético/metabolismo , Nanoconjugados/administração & dosagem , Nanoconjugados/química , Nanopartículas/química , Ratos , Ratos Endogâmicos Lew , Tarso Animal/patologia , Distribuição Tecidual , beta-Galactosidase/biossíntese , beta-Galactosidase/genéticaRESUMO
Interleukin-1 receptor antagonist (IL-1Ra), is a natural blocker of the inflammatory cytokine interleukin-1. Using a rat adjuvant-induced arthritis (AIA) model of rheumatoid arthritis (RA), we examined the protective effects of IL-1Ra in bone metabolism in vivo after folate-mediated nonviral gene delivery. We detected secreted human IL-1Ra protein in serum and cultured primary osteoblasts of rats that were treated with chitosan-IL-1Ra and folate-IL-1Ra-chitosan nanoparticles, respectively. In vivo, IL-1Ra gene delivery significantly reverted alterations in bone turnover observed in arthritic animals by modulating the level of osteocalcin (OC) as well as the activities of alkaline phosphatase and tartrate-resistant acid phosphatase. The protective effects of these nanoparticles were evident from the decrease in the expression levels of interleukine-1beta and prostaglandin E(2) as well as osteoclast number and other histopathological findings. Compared to naked DNA and chitosan-DNA, folate-chitosan-DNA nanoparticles were less cytotoxic and enhanced IL-1Ra protein synthesis in vitro and offered a better protection against inflammation and abnormal bone metabolism in vivo. Nonviral gene therapy with folate-chitosan-DNA nanoparticles containing the IL-1 Ra gene seemed to protect against bone damage and inflammation in rat adjuvant-induced arthritis model.
Assuntos
Artrite Experimental/terapia , Osso e Ossos/metabolismo , Terapia Genética/métodos , Proteína Antagonista do Receptor de Interleucina 1/genética , Nanopartículas , Fosfatase Ácida/sangue , Fosfatase Alcalina/sangue , Animais , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Osso e Ossos/patologia , Quitosana/química , DNA/química , Dinoprostona/sangue , Ácido Fólico/química , Técnicas de Transferência de Genes , Humanos , Interleucina-1beta/sangue , Isoenzimas/sangue , Nanopartículas/química , Osteoblastos/metabolismo , Ratos , Fosfatase Ácida Resistente a Tartarato , Fator de Necrose Tumoral alfa/sangueRESUMO
Quantum dots (QDs) are emerging as alternative or complementary tools to the organic fluorescent dyes currently used in bioimaging. QDs hold several advantages over conventional fluorescent dyes including greater photostability and a wider range of excitation/emission wavelengths. However, recent work suggests that QDs exert deleterious effects on cellular processes. This study examined the subcellular localization and toxicity of cadmium telluride (CdTe) QDs and pharmacological means of preventing QD-induced cell death. The localization of CdTe QDs was found to depend upon QD size. CdTe QDs exhibited marked cytotoxicity in PC12 and N9 cells at concentrations as low as 10 microg/ml in chronic treatment paradigms. QD-induced cell death was characterized by chromatin condensation and membrane blebbing and was more pronounced with small (2r=2.2+/-0.1 nm), green emitting positively charged QDs than large (2r=5.2+/-0.1 nm), equally charged red emitting QDs. Pretreatment of cells with the antioxidant N-acetylcysteine and with bovine serum albumin, but not Trolox, significantly reduced the QD-induced cell death. These findings suggest that the size of QDs contributes to their subcellular distribution and that drugs can alter QD-induced cytotoxicity.