ABSTRACT
Aim: To prepare chitosan nanoparticlescarrying gene and study its characteristics in vitro and in vivo.Methods:12-ACSs was dissolved in 0.05mol/L sodium acetate buffer to form a solution of 1mg/ml and a DNA(plasmid-encoding antisense ECE ) solution of 0.1mg/ml was dissolved in 25mmol/L Na2SO4.The charge ratio of components is selected as 2/1 for 12-ACSs /DNA complex.The complex was prepared by mixing 12-ACSs solution with DNA solution prior to observation by using transmission electron microscopy.Using Electrophoretic Retardation of DNA Migration detection,DNA precipitation,Binding Equilibration and DNase Resistivity Assay,the formation of 12-ACSs /DNA complex was determined and its stability was simultaneously evaluated.MTT Cell Proliferation Assays was performed on investigation of the cytotoxicity of 12-ACSs /DNA nanoparticles in bronchial epithelial cells (16HBE).The transfection efficiency of 12-ACSs /DNA nanoparticles in vitro and in vivo was investigated in 16HBE cells and Balb/c mice.Results: 12-ACSs /DNA nanoparticles (100~150nm) were observed by transmission electron microscopy.12-ACSs can protect the plasmid encoding antisence-ECE from degradation by DNase I.12-ACSs can transfer plasmid-encoding antisense ECE into 16HBE cells and into the airway epithelium in mice.As shown by fluorescent microscopy,green fluorescent protein reporter can be observed in the transfected cells as well as in the airway epithelium of the treated mice.And it showed a lower cell cytotocixity in cultured 16HBE cells and in mice treated with12-ACSs /DNA nanoparticles.Conclusion: In summery,the chitosan can be used as an effective protectant for DNA as well as an enhancer for in vitro gene transfection.
ABSTRACT
<p><b>OBJECTIVE</b>To observe the therapeutic effect and mechanism of Naohuandan (NHD) in treating senile dementia (SD).</p><p><b>METHODS</b>Clinical study: Fifty-eight patients with SD, whose diagnosis conforms to the Diagnostic Standard of DSM-IV issued by American Association of Psychiatry, were enrolled and randomly assigned into two groups. The 30 patients in the treated group were treated with NHD, 4 capsules each time, 3 times daily. The 28 patients in the control group were treated with Piracetam, 1.6 g each time, 3 times daily. The therapeutic course for both groups was 3 months. The therapeutic efficacy was estimated and compared by comprehensive scores of memory and cognition, scores of Mini-mental State Examination (MMSE) and Activities of Daily Living (ADL). Experimental study: Rats were divided into the control group, the model group and the high-dosage and low-dosage NHD treated groups. The protective effect of NHD on the per-oxidative damage of hippocampal neurons in beta-amyloid protein induced SD model was observed and the related criteria were determined.</p><p><b>RESULTS</b>Clinical study showed that both NHD and Piracetam could improve the clinical symptoms of patients, the two medicines showing insignificant difference in total effective rate. But NHD was better in elevating MMSE score and lowering ADL score in patients than Piracetam (P < 0.05 and P < 0.01). Experimental study showed that (1) 24 and 72 hrs after modeling, the activity of SOD and GSH were lower and the level of MDA higher in the model group than those in the control group (P < 0.05 or P < 0.01). Compared with the model group at the corresponding time points, in the high-dosage NHD group, SOD and GSH were higher, MDA was lower (P < 0.05 or P < 0.01); but in the low-dosage NHD group, SOD at the 72nd hr was higher (P < 0.05) and MDA at 24th and 72nd hrs was lower (P < 0.01). And most of the criteria in the high-dosage NHD group was improved better than that in the low-dosage NHD group. (2) The survival rates of neurons in various groups were not different significantly (P > 0.05) 24 hrs after modeling, but that in the high-dosage NHD group was significantly higher than that in the model group (P < 0.01) and in the low-dosage NHD group 72 hrs after modeling (P < 0.05).</p><p><b>CONCLUSION</b>NHD is an effective Chinese herbal preparation for treatment of SD, and its mechanism is related with its inhibition on peroxidative injury and protection on neurons.</p>