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Objective To construct a PAEH/PEG DA hydrogel sustained-release system containing combined Pseudomonas aeruginosa (PA) outer membrane protein F (OprF) and Pseudomonas V antigen (PcrV) gene DNA vaccine, and evaluate the in vivo immune efficacy of the system. Methods The PAEH/PEG DA hydrogel containing combined PA DNA vaccine was prepared with a simple mixing method. The gelation time was tested, and the cytotoxicity (DC 2.4), degradation and cumulative release rate in vitro of the hydrogel were evaluated. Nine mice were randomly divided into 3 groups (3 each): 30 min group, 2 d group and 7 d group, the in vivo degradability and security of the hydrogel were evaluated by gelation volume changes at the injection site and histopathological sections of the skin, respectively. Eighteen mice were randomly divided into 6 groups (3 each): control group (PBS),hydrogel group (Gel), En/pVAX1-OprF group (O), En/pVAX1-PcrV group (P), En/(pVAX1-OprF+pVAX1-PcrV) group (OP), and hydrogel + En/(pVAX1-OprF+pVAX1-PcrV) group (GOP). Mice were immunized 3 times with a 10-day interval, then sacrificed 2 weeks after last immunization. The levels of serum specific IgG antibody, splenic lymphocyte stimulation index (SI) and interferon-γ(IFN-γ) in the supernatant of splenic lymphocytes were detected. Results The PAEH/PEG DA hydrogel sustained-release system required only about 30 min to form a gelation state. There was no significant toxicity to DC 2.4 cells in vitro, and approximately 85%of plasmid DNA was released after 36 hours of in vitro release. The degradation time of the hydrogel was nearly the same in vitro and in vivo. It could be almost completely degraded in about 7 days, and had good in vivo biodegradability and biosafety. The results of in vivo immune test showed that, compared with PBS group, no significant changes existed in the levels of specific IgG antibodies,splenic lymphocyte SI and IFN-γ in the Gel group (P>0.05). Compared with the corresponding DNA vaccine groups (O group or P group) and PBS group, the specific IgG antibody levels, spleen lymphocyte SI and IFN-γ level increased obviously in OP group and GOP group (P<0.05, P<0.01). While compared with OP group, the levels of specific IgG antibodies, splenic lymphocyte SI and IFN-γ increased markedly in GOP group (P<0.05, P<0.01). Conclusion The PAEH/PEG DA hydrogel sustained-release systems containing PA OprE and PcrF gene combined DNA vaccines, which can slowly release the combined DNA vaccine and further enhance the immune efficacy of combined DNA vaccine, are one of the promising strategies for the development of PA vaccines.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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