Microbiology Department, College of Medicine and Medical Research Institute, Chungbuk National University, 12 Gaeshin-Dong Heungduk-Ku, Cheongju 361-763, Republic of Korea.
Virol J. 2013 Apr 3;10:104. doi: 10.1186/1743-422X-10-104.
Influenza vaccines are prepared annually based on global epidemiological surveillance data. However, since there is no method by which to predict the influenza strain that will cause the next pandemic, the demand to develop new vaccination strategies with broad cross-reactivity against influenza viruses are clearly important. The ectodomain of the influenza M2 protein (M2e) is an attractive target for developing a vaccine with broad cross-reactivity. For these reasons, we investigated the efficacy of an inactivated H9N2 virus vaccine (a-H9N2) mixed with M2e (1xM2e or 4xM2e) proteins expressed in Escherichia coli, which contains the consensus of sequence the extracellular domain of matrix 2 (M2e) of A/chicken/Vietnam/27262/09 (H5N1) avian influenza virus, and investigated its humoral immune response and cross-protection against influenza A viruses.
Mice were intramuscularly immunized with a-H9N2, 1xM2e alone, 4xM2e alone, a-H9N2/1xM2e, or a-H9N2/4xM2e. Three weeks post-vaccination, mice were challenged with lethal homologous (A/ chicken /Korea/ma163/04, H9N2) or heterosubtypic virus (A/Philippines/2/82, H3N2 and A/aquatic bird/Korea/maW81/05, H5N2). Our studies demonstrate that the survival of mice immunized with a-H9N2/1xM2e or with a-H9N2/4xM2e (100% survival) was significantly higher than that of mouse-adapted H9N2 virus-infected mice vaccinated with 1xM2e alone or with 4xM2e alone (0% survival). We also evaluated the protective efficacy of the M2e + vaccine against infection with mouse-adapted H5N2 influenza virus. Protection from death in the control group (0% survival) was similar to that of the 1×M2e alone and 4xM2e alone-vaccinated groups (0% survival). Only 40% of mice vaccinated with vaccine alone survived challenge with H5N2, while the a-H9N2/1×M2e and a-H9N2/4×M2e groups showed 80% and 100% survival following mouse-adapted H5N2 challenge, respectively. We also examined cross-protection against human H3N2 virus and found that the a-H9N2/1×M2e group displayed partial cross-protection against H3N2 (40% survival), whereas vaccine alone, 1×M2e alone, 4×M2e alone, or H9N2/1×M2e groups showed incomplete protection (0% survival) in response to challenge with a lethal dose of human H3N2 virus.
Taken together, these results suggest that prokaryote-expressed M2e protein improved inactivated H9N2 virus vaccine efficacy and achieved cross-protection against lethal influenza A virus infection in mice.
流感疫苗是基于全球流行病学监测数据每年制备的。然而，由于没有方法预测下一次大流行将导致哪种流感毒株，因此显然需要开发针对流感病毒具有广泛交叉反应性的新疫苗接种策略。流感 M2 蛋白（M2e）的外域是开发具有广泛交叉反应性的疫苗的有吸引力的靶标。出于这些原因，我们研究了含有 A/鸡/越南/27262/09（H5N1）禽流感病毒基质 2（M2）的外域的共识序列的大肠杆菌中表达的灭活 H9N2 病毒疫苗（a-H9N2）与 M2e（1xM2e 或 4xM2e）蛋白混合的功效，以及其针对甲型流感病毒的体液免疫反应和交叉保护作用。
小鼠经肌肉内免疫接种 a-H9N2、1xM2e 单独、4xM2e 单独、a-H9N2/1xM2e 或 a-H9N2/4xM2e。免疫接种后 3 周，用致死同源（A/鸡/韩国/ma163/04，H9N2）或异源病毒（A/菲律宾/2/82，H3N2 和 A/水禽/韩国/maW81/05，H5N2）对小鼠进行攻毒。我们的研究表明，用 a-H9N2/1xM2e 或 a-H9N2/4xM2e（100%存活）免疫接种的小鼠的存活率明显高于仅用 1xM2e 单独或仅用 4xM2e 单独接种的经适应小鼠的 H9N2 病毒感染的小鼠（0%存活）。我们还评估了 M2e+疫苗对感染适应小鼠的 H5N2 流感病毒的保护作用。对照组（0%存活）的死亡保护作用与仅用 1xM2e 单独和 4xM2e 单独接种的组（0%存活）相似。仅用疫苗单独接种的小鼠中有 40%在接受 H5N2 攻毒后存活下来，而用 a-H9N2/1xM2e 和 a-H9N2/4xM2e 攻毒的组在接受适应小鼠的 H5N2 攻毒后分别显示出 80%和 100%的存活。我们还检查了对人 H3N2 病毒的交叉保护作用，发现 a-H9N2/1xM2e 组对 H3N2 显示出部分交叉保护作用（40%存活），而仅用疫苗、1xM2e 单独、4xM2e 单独或 H9N2/1xM2e 组在接受致死剂量的人 H3N2 病毒攻毒时则显示出不完全保护作用（0%存活）。
综上所述，这些结果表明，原核表达的 M2e 蛋白提高了灭活的 H9N2 病毒疫苗的功效，并在小鼠中实现了针对致死性甲型流感病毒感染的交叉保护作用。
