[Immunoprotective effect of a plasmid DNA vaccine pCMV-LC3-Ag85B against Mycobaterium tuberculosis].
OBJECTIVE
To construct an autophagy-targeted vaccine harboring the genes encoding Ag85B and microtubule-associated protein light chain-3 (LC3) and to explore its immunoprotection against Mycobacterium tuberculosis (MTB).


METHODS
The pCMV-LC3-Ag85B plasmid was constructed and used to transfect RAW264.7 cells. The level of LC3-Ag85B was detected using Western blotting. Then, BALB/c mice were immunized with pCMV, pCMV-Ag85B and pCMV-LC3-Ag85B plasmid, respectively. In vitro, two weeks after the last immunization, the secretion of IL-2, IFN-γ, IL-4 and IL-10 from Ag85B-stimulated lymphocytes was measured by ELISA. Three months after the last immunization, all mice were challenged with MTB H37Rv via the tail vein and the bacterial loads in their spleens and lungs were determined by colony formation assay.


RESULTS
The LC3-Ag85B fusion protein was expressed in RAW264.7 cells that had been transfected with pCMV-LC3-Ag85B and the expression level was in a dose-dependent manner. Compared with the pCMV-Ag85B treatment group, pCMV-LC3-Ag85B-immunized mice showed a significant increase of IL-2 and IFN-γ levels and the lower loads of MTB in the spleens and lungs.


CONCLUSIONS
pCMV-LC3-Ag85B can enhance a specific Th1-predominant immunity and a superior immunoprotection against MTB, which may provide a new practical strategy for the development of improved vaccines against MTB.
Department of Immunology and Laboratory Medicine, Medical School, Institute of Infection and Immunology, Anhui University of Science and Technology, Huainan 232001, China.
Dong
D
Hu
Department of Immunology and Laboratory Medicine, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
Xiaokang
X
Yang
Department of Immunology and Laboratory Medicine, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
Wan
W
Wang
Affiliated Cancer Hospital, Anhui University of Science and Technology, Huainan 232001, China.
Yingru
Y
Xing
Department of Immunology and Laboratory Medicine, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
Wenyang
W
Wang
Department of Immunology and Laboratory Medicine, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
Zhaohong
Z
DU
Department of Immunology and Laboratory Medicine, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
Qi
Q
Yiu
Department of Immunology and Laboratory Medicine, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
Li
L
Zhang
Department of Immunology and Laboratory Medicine, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
Liping
L
Chen
Department of Immunology and Laboratory Medicine, Medical School, Institute of Infection and Immunology, Anhui University of Science and Technology, Huainan 232001, China.
Jing
J
Wu
Department of Immunology and Laboratory Medicine, Medical School, Institute of Infection and Immunology, Anhui University of Science and Technology, Huainan 232001, China.
Rongbo
R
Zhang
D000217:Acyltransferases / Q000235:genetics* / Q000276:immunology; D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics* / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology; D016207:Cytokines / Q000378:metabolism; D003587:Cytomegalovirus / Q000235:genetics; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D008869:Microtubule-Associated Proteins / Q000235:genetics*; D010957:Plasmids; D014376:Tuberculosis / Q000517:prevention & control; D032581:Tuberculosis Vaccines / Q000276:immunology*; D019444:Vaccines, DNA / Q000276:immunology*
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D016207:Cytokines; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D032581:Tuberculosis Vaccines; D019444:Vaccines, DNA; D000217:Acyltransferases; C121433:antigen 85B, Mycobacterium tuberculosis