Evaluation of immunogenicity and protective efficacy against Mycobacterium tuberculosis infection elicited by recombinant Mycobacterium bovis BCG expressing human Interleukin-12p70 and Early Secretory Antigen Target-6 fusion protein

Abstract

ESAT-6 protein of Mycobacterium tuberculosis is absent in Mycobacterium bovis BCG and Mycobacterium microti and has been demonstrated to stimulate strong cell-mediated immunity. IL-12 can play crucial roles in regulating IFN-γ production and Th1 effectors production. In this study, we constructed three rBCG vaccines that could express proteins of human IL-12p70 and/or ESAT-6 and evaluated their immunogenicity and protective efficacy in mice. Our experiments illustrated that the rBCG-IE (expressing a fusion protein of human IL-12p70 and ESAT-6) was capable of inducing stronger Th1 type cell-mediated immune responses than conventional BCG, or rBCG-I (expressing human IL-12p70), or rBCG-E (expressing ESAT-6). However, the results of protective experiments showed that rBCG-IE could only confer similar and even lower protective efficacy against M. tuberculosis H37Rv infection compared with BCG vaccine.

Role of Fused Mycobacterium tuberculosis Immunogens and Adjuvants in Modern Tuberculosis Vaccines

“…However, none of the recombinant constructs surpassed the protection induced by wild-type BCG [Table 1; Ref. (25)].…”

Recombinant Mycobacterium smegmatis with a pMyong2 vector expressing Human Immunodeficiency Virus Type I Gag can induce enhanced virus-specific immune responses

“…Thereafter, the plates were washed again and incubated with HRP conjugated streptavidin for 30 min at RT, and reacted with BD OptEIA substrate (BD Biosciences) for 10 min before stopping the reaction with 1N H 2 SO 4 . Optical density (OD) was determined by spectrophotometry at 450 nm50.…”

AMycobacterium bovisBCG-Naked DNA Prime-Boost Vaccination Strategy Induced CD4+and CD8+T-Cell Response againstMycobacterium tuberculosisImmunogens

“…The proliferation of lymphocyte were tested by MTT assay [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazoliumbromide]. The method was described as previous [22, 27]. The results are expressed as the value of stimulation index (SI).…”

“…The splenocytes were prepared and cultured as previously described [27, 28], and the splenocytes were plated in 6-well flat-bottom plates (5∗10 6 cells in 2 mL of cRPMI per well) with 100  μ L TB-PPD (1  μ g/mL; XiangRui Biotech, Ltd., Beijing, China) in each well and incubated for 72 h (37°C, 5%CO 2 ). The cells were collected and washed three times with 0.1 M PBS (PH = 7.2), and then rabbit anti-Mouse CD4 + -PE and anti-Mouse CD8 + -FITC (eBioscience, USA) were added into EP tube for a 30 min incubation in an ice-bath keep out of the sun.…”