Department of Microbiology and Pathogen Biology, Air Force Medical University, Xi'an, China.
Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, China.
Front Immunol. 2019 Jul 3;10:1519. doi: 10.3389/fimmu.2019.01519. eCollection 2019.
Bacillus Calmette-Guerin (BCG) is a live attenuated vaccine against tuberculosis (TB) and remains the most commonly used vaccine worldwide. However, BCG has varied protective efficiency in adults and has safety concerns in immunocompromised population. Thus, effective vaccines are necessary for preventing the prevalence of TB. Cyclic di-AMP (c-di-AMP) is a bacterial second messenger which regulates various cellular processes and host immune response. Previous work found that c-di-AMP regulates bacterial physiological function, pathogenicity and host type I IFN response. In this study, we constructed a recombinant BCG (rBCG) by overexpressing DisA, the diadenylate cyclase of (Mtb), and observed the physiological changes of rBCG-DisA. The immunological characteristics of rBCG-DisA were investigated on humoral and cellar immune responses in a mice infection model. Our study demonstrated that overexpression of DisA in BCG does not affect the growth but reduces the length of BCG. rBCG-DisA-immunized mice show similar humoral and cellar immune responses in BCG-immunized mice. After Mtb infection, the splenic lymphocytes from both BCG and rBCG-DisA-immunized mice produced more IFN-γ, IL-2, and IL-10 than the un-immunized (UN) mice, while the cytokine levels of the rBCG-DisA group increased significantly than those of the BCG group. The transcription of IFN-β, IL-1β and autophagy related genes (Atgs) were up-regulated in macrophages after treated with c-di-AMP or bacterial infection. The productions of IL-6 were increased after Mtb challenge, especially in the rBCG-DisA-immunized mice. Strikingly, H3K4me3, the epigenetic marker of innate immune memory, was found in both two immunized groups, and the rBCG-DisA group showed stronger expression of H3K4me3 than that of BCG. In addition, the pathological changes of rBCG-DisA immunized mice were similar to that of BCG-immunized mice. The bacterial burdens in the lungs and spleens of BCG- and rBCG-DisA-immunized mice were significantly decreased, but there was no significant difference between the two immunized groups. Together, these results suggested that compared to BCG, rBCG-DisA vaccination, induces stronger immune responses but did not provided additional protection against Mtb infection in this study, which may be related to the innate immunity memory. Hence, c-di-AMP is a promising immunomodulator for a further developed BCG as a better vaccine.
卡介苗（BCG）是一种预防结核病（TB）的减毒活疫苗，仍是目前全球应用最广泛的疫苗。然而，BCG 在成人中的保护效果存在差异，且在免疫功能低下人群中存在安全性问题。因此，需要有效的疫苗来预防结核病的流行。环二核苷酸（c-di-AMP）是一种细菌第二信使，可调节多种细胞过程和宿主免疫反应。先前的研究发现 c-di-AMP 可调节细菌的生理功能、致病性和宿主 I 型干扰素反应。在这项研究中，我们通过过表达分枝杆菌的二腺苷酸环化酶 DisA 构建了重组卡介苗（rBCG），并观察了 rBCG-DisA 的生理变化。我们在小鼠感染模型中研究了 rBCG-DisA 的免疫学特征，包括体液和细胞免疫反应。我们的研究表明，BCG 中 DisA 的过表达不影响其生长，但会降低 BCG 的长度。rBCG-DisA 免疫的小鼠在体液和细胞免疫反应方面与 BCG 免疫的小鼠相似。在 Mtb 感染后，rBCG-DisA 免疫的小鼠的脾淋巴细胞比未免疫（UN）小鼠产生更多的 IFN-γ、IL-2 和 IL-10，而 rBCG-DisA 组的细胞因子水平明显高于 BCG 组。用 c-di-AMP 或细菌感染处理巨噬细胞后，IFN-β、IL-1β 和自噬相关基因（Atgs）的转录上调。在 Mtb 挑战后，IL-6 的产生增加，尤其是在 rBCG-DisA 免疫的小鼠中。值得注意的是，在两种免疫组中均发现了组蛋白 H3K4me3，这是先天免疫记忆的表观遗传标记，而 rBCG-DisA 组的 H3K4me3 表达强于 BCG 组。此外，rBCG-DisA 免疫的小鼠的病理变化与 BCG 免疫的小鼠相似。BCG 和 rBCG-DisA 免疫的小鼠的肺部和脾脏中的细菌负荷明显降低，但两组之间没有显著差异。综上所述，与 BCG 相比，rBCG-DisA 疫苗接种可诱导更强的免疫反应，但在本研究中并未提供针对 Mtb 感染的额外保护，这可能与先天免疫记忆有关。因此，c-di-AMP 是一种有前途的免疫调节剂，可进一步开发作为更好的疫苗的 BCG。
