Recombinant Mycobacterium smegmatis expressing Hsp65-hIL-2 fusion protein and its influence on lymphocyte function in mice
Asian Pacific Journal of Tropical Medicine
;
(12): 347-351, 2012.
Article
Dans Anglais
| WPRIM
| ID: wpr-819770
ABSTRACT
OBJECTIVE@#To construct a strain of recombinant Mycobacterium smegmatis expressing the heat shock protein 65 (Hsp65) and human interleukin 2 (IL-2) fusion protein (rMS-Hsp65/IL-2) and to explore the effect of this construct on lymphocyte function in mice.@*METHODS@#The fusion gene encoding Hsp65-hIL-2 was cloned into shuttle vector pSMT3. The recombinant plasmid pSMT3-Hsp65-hIL-2 was transferred to Mycobacterium smegmatis by electroporation. Positive clones were selected by hygromycin and identified by PCR. The expression of fusion protein Hsp65-hIL-2 was verified using indirect immunofluorescence staining. Mice were immunized for two times by subcutaneously injection with 1×10(6) CFU rMS-Hsp65/IL-2 at a three-week interval. Two weeks after the second immunization, mice were sacrificed and the serum samples were collected for determination of anti-Hsp65 specific IgG. Splenic lymphocytes were isolated and treated with the rMS-Hsp65/IL-2 to determine lymphocytic proliferation activity by MTT assay. IFN-γ- and IL-2 in the medium of the treated cells were also determined by ELISA.@*RESULTS@#Successful construction of rMS-Hsp65/IL-2 was verified by PCR and immunofluorescence staining. Compared to the splenic lymphocytes isolated from mice immunized with Bacille Calmette-Guerin or mice immunized with Mycobacterium smegmatis alone, the splenic lymphocytes isolated from mice immunized with rMS-Hsp65/IL-2 showed a marked increase in the proliferation of lymphocytes, together with an increased production of important cytokines such as IFN-γ-and IL-2.@*CONCLUSIONS@#rMS-Hsp65/IL-2 markedly enhances lymphocyte function. Therefore, the fusion protein generated by rMS-Hsp65/IL-2 may be of potential value in generating an effective vaccine against tuberculosis.
Texte intégral:
Disponible
Indice:
WPRIM (Pacifique occidental)
Sujet Principal:
Rate
/
Protéines bactériennes
/
Protéines recombinantes
/
Immunoglobuline G
/
Lymphocytes
/
Cytokines
/
Interleukine-2
/
Chaperonine-60
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Mycobacterium smegmatis
/
Prolifération cellulaire
Type d'étude:
Étude pronostique
Limites du sujet:
Animaux
langue:
Anglais
Texte intégral:
Asian Pacific Journal of Tropical Medicine
Année:
2012
Type:
Article
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