The aim of the present study was to investigate the effects of various decellularization methods on the histological and biomechanical properties of rabbit tendons. In total, six chemical reagents, including 1% t-octyl-phenoxypolyethoxyethanol (Triton-X 100), 0.5% sodium dodecyl sulfate (SDS), 1% tri-n-butyl phosphate (TnBP), 1% Triton-X 100 + 0.5% SDS, 1% TnBP + 0.5% SDS and 1% TnBP + 1% Triton-X 100, were used on rabbit semitendinosus muscles and flexor digitorum tendons for 24 h to remove cells. Hematoxylin and eosin staining was applied for histological observation, while tension testing was used for biomechanical studies. The effects of the various decellularization methods on the histological structure and biomechanical properties of rabbit tendons were evaluated. A group of fresh tendons treated with phosphate-buffered saline served as controls. The various decellularization methods resulted in different effects on the tendons. All the treatment groups exhibited a decrease in tendon biomechanical properties, but no statistically significant differences were observed among the experimental groups. The extensibility of the 1% TnBP-treated group was found to be greater than that of the other groups; however, the difference was not statistically significant. Histologically, the 1% TnBP + 0.5% SDS treatment was shown to have the least impact on the rabbit tendon structure, with good decellularization and no clear cellular remnants observed. The 1% Triton-X 100 + 0.5% SDS treatment had a pronounced effect on the tendon collagen structure and a number of collagen ruptures were observed. Overall, 1% TnBP + 0.5% SDS was found to be the most effective compared with the other treatments, as this treatment preserved the tendon collagen structure while completely removing the cells. Tendons treated with 1% TnBP + 0.5% SDS were histologically similar to normal tendon tissue and biomechanically similar to the tendons in the control group.

Percutaneous immune method is becoming an attractive alternative for DNA vaccine as a lot of antigen presenting cells are existed in the viable epidermis. However, due to the barrier function of stratum corneum, it would be hard for DNA vaccine to reach the viable epidermis of the skin. In order to deliver the DNA vaccine successfully cross the stratum corneum, pentagram silicon microneedle array was prepared in this study, and fluorescently labeled nanoparticle was taken as the model to observe the situation inside the skin processed by microneedle. Via microneedle nanoparticles could enter the skin through the micro-channel (diameter about 20-30 µm) and its amount is greatly larger than that enter though the hair follicle of intact skin. A new type of gene vector Pluronic P123-modified polyethyleneimine (P123-PEI) was synthesized by high molecular weight polyethylenimine and Pluronic P123 with the molar ratio of 1 : 1 to take the advantage of P123-PEI as low cytotoxicity and high transfection efficiency. Mice were immunized percutaneously with Hepatitis B DNA vaccine/P123-PEI nano-complexes by microneedle. The humoral and cellular immunity generated in percutaneously immunized mice through microneedle array by Hepatitis B DNA vaccine/P123-PEI nano-complex was significantly higher than that of DNA vaccine intramuscular administration.Key words hepatitis B DNA vaccine; microneedle; percutaneous immune; nano-complex; Pluronic P123-modified polyethyleneimine DNA vaccine that the eukaryon expression plasmid encoded by antigen gene was inoculated in vivo and can express corresponding antigen to stimulate the organism to generate immune response to the antigen and protective immunity was the third generation vaccine on the basis of gene therapy and transgenic technology, which marked a vaccine revolution and has a wide application prospect. 1) However, DNA vaccines had its own disadvantage. Numerous experiments, particularly big animal experiments and human clinical trials, demonstrated that currently DNA vaccine had the disadvantages such as large dose, low bioavailability, great variation in immunity effect among individuals and low level of humoral immunity induced etc. The disadvantage of DNA vaccine enormously affected the research progress and the wide clinical application.2) According to in vivo immune mechanism of DNA vaccine, antigen presenting cells (APCs) like phagocytes and dendritic cells was playing an important role in the process of transfection, expression and presentation of DNA vaccine. 3,4)How to improve the cell transfection efficiency of DNA vaccine, particularly the transfection efficiency of APCs, becomes one of the key approaches for improvement of immunity effect of DNA vaccines. Currently, the dosage forms of DNA vaccine are mainly water solution or lyophilized powder injection and intramuscular injection is the most common route of immune administration. DNA vaccine is usually consisted of 2-10 kbp plasmid DNA and his molecular weight reached million with a strong hydrophilicity and low oi...