流感通用疫苗的研究现状与展望

流感在全球范围内的不断暴发引起了世界的高度关注。预防流感最有效和最经济的手段是疫苗免疫。流感病毒的多宿主和高度变异性,使得流感疫苗的有效性受到限制。目前使用的疫苗只对与疫苗株相同或高度相近的病毒株具有保护效力。一旦变异较大或新亚型的流感病毒出现,现有疫苗就会失去其保护效力。因此,研制一种可抵御变异较大或不同亚型流感病毒的通用疫苗成为流感疫苗研究的热点。选择流感病毒的保守蛋白做抗原是研制流感通用疫苗的主要方法,已有至少3种流感通用疫苗进入临床研究阶段。改变疫苗免疫途径和免疫程序,也能提高疫苗的交叉免疫效果。本文对流感通用疫苗的研究进展进行了综述与展望。     

通用流感疫苗研究进展

流感是一种对人类危害极大的传染病,接种疫苗被认为是预防流感的最有效手段。目前所用的流感疫苗主要是根据现行流行株的减毒或灭活病毒疫苗及基于流感血凝素和神经氨酸酶设计的重组蛋白质疫苗。但流感病毒变异大,易逃逸机体免疫监视,有效的疫苗须不断分离新流行株和不断更新疫苗免疫原。为解决这一问题,很多科学家一直在研究基于病毒高度保守性蛋白质、能够预防所有流感病毒毒株、可诱导持久保护性免疫的通用流感疫苗。我们对基于基质蛋白M2、核蛋白等的通用流感疫苗做一简要综述。     

Vero细胞培养流行性感冒病毒的研究

探索Vero细胞培养流感病毒和用于研制流感疫苗的可行性。确立Vero细胞培养流感病毒的最适条件,把流感病毒接种于Vero细胞上培养和传代,于不同时间收获病毒液,进行灭活和超滤浓缩实验,检测血凝素滴度(HA)。结果表明胰酶、pH值、残留牛血清是流感病毒在Vero细胞培养的影响因素,最佳收毒时间为72-96小时。Vero细胞上培养流感病毒的4～7代,HA滴度较高。病毒液用0．05％福尔马林4℃,7天即可灭活,用超滤技术能浓缩流感病毒液。Vero细胞可用于流感病毒的培养和疫苗的开发。     

临床试验中的流感通用疫苗研发进展

人类与流感病毒的斗争史已经过去一百多年,在人群抗体选择的压力下,流感病毒不断发生突变、重组等方式的进化和流行,在流感病毒的危险因素下,人类的预防医学、病原生物学和疫苗学也取得了巨大进步。对流感通用疫苗的研发成为流感疫苗研发的下一个征程,同时需要全球范围内的广泛协作和科技发展。为此本文综述了近期流感通用疫苗的研究进展,并对已经进入临床试验阶段的通用疫苗进行了分析,为流感疫苗研究提供参考。     

通用流感疫苗研究进展

流行性感冒病毒(influenza virus,简称流感病毒)是一种严重威胁人类健康的病毒。全世界每年有上万人死于流感病毒引起的流行性感冒(influenza,简称流感)。同时,流感也对全球经济发展产生严重影响。接种流感疫苗(influenza vaccines)是流感防控的重要手段之一。流感病毒亚型众多、抗原变异较快且因其流行趋势难以预测等因素导致传统流感疫苗接种后免疫效果不佳,研发通用流感疫苗成为预防流感的一种理想策略。现对以流感病毒血凝素(hemagglutinin, HA)、神经氨酸酶(neuraminidase, NA)及流感病毒其他结构蛋白保守区域为免疫原的通用流感疫苗的研究进展作一概述。     

甲型流感病毒M2e通用疫苗的研究进展

流感严重地影响人们的身体健康和工作生活,给社会带来巨大经济损失。疫苗接种是预防流感的有效措施之一,市场上的流感疫苗包括流感灭活疫苗、减毒活疫苗和亚单位疫苗等。这些流感疫苗只能预防相同亚型流感病毒的感染,无法预防不同亚型流感病毒引起的季节性流感和流感大流行,因此迫切需要研发广谱的能预防不同亚型的甲型流感病毒感染的通用疫苗。在此简要介绍甲型流感病毒M2e通用疫苗的研究进展。     

流感疫苗接种引起发作性睡病

在2009年全球A(H1N1)流感病毒大流行期间,多种A(H1N1)大流行流感疫苗被批准应用。一种以AS03-为佐剂的A(H1N1)大流行流感疫苗(Pandemrix)被应用于47个国家,达3 100万剂。在应用一年后,开始出现接种Pandemrix疫苗所引起的发作性睡病的报道。发作性睡病是一种尚无治疗方法的疾病,主要症状表现是患者在白天常发生不可自控的睡眠。报道首先来自于瑞士,然后丹麦,继而欧洲的许多国家。迄今为止,已有1 300多人因接种Pandemrix疫苗发生了发作性睡病,生产Pandemrix疫苗的葛兰素史克(Glaxo Smith-Kline,GSK)公司已承认了两者间的关联。2015年7月1日,科学转化医学杂志的一篇文章揭示Pandemrix疫苗中的流感病毒核蛋白刺激所产生的抗体可以和人下丘脑泌素受体2发生交叉反应,表明Pandemrix疫苗激发的自身免疫反应是其引起发作性睡病的主因,提示有必要显著减少或去除流感病毒疫苗中的流感病毒核蛋白。     

流行性感冒病毒鸡胚高产株的遗传特性分析

流行性感冒（流感）病毒的基因组由分节段的单股负链RNA组成,其中A、B型流感病毒含8个基因节段［1］。它的第4和第6节段分别编码病毒的血凝素（HA）和神经氨酸酶（NA）,决定病毒的抗原性,其它6个节段与病毒的生长特性有关［2］。在流感疫苗生产中,为了提高产量,利用高产毒株与流行毒株基因重配获得重组病毒,它含有流行毒株的第4、第6节段和高产毒株的其它6个节段,这样既具有流行毒株的抗原性又具有高产特性,可以用来降低疫苗的生产成本。     

一株2011年出现的季节性甲型H1N1流行性感冒病毒血凝素基因特性的研究

本文通过比较2011年分离培养的1株季节性甲型H1N1流行性感冒(简称流感)病毒(A/Shanghai/1167/2011(H1N1))与历年季节性甲型H1N1流感病毒的血凝素(HA)基因,追溯该病毒的基因变异与来源,探讨该毒株的出现对流感防控工作的意义.采用反转录-聚合酶链反应(RT-PCR)方法扩增病毒的HA和神经氨酸酶(NA)片段,并进行测序;应用分子生物学软件对获得的序列进行分析,绘制基因进化树;同时,通过血凝抑制试验检测2011年下半年健康人群中该流感病毒的抗体水平.结果显示,A/Shanghai/1167/2011(H1N1)的HA基因序列与世界卫生组织(WHO)2007～2008年季节性甲型H1N1流感病毒疫苗株A/Brisbane/59/2007(H1N1)最接近,同源性达99.2%,与新型甲型H1N1流感病毒A/California/07/2009疫苗株同源性仅为72.4%.其HA基因裂解位点为PSIQSR↓GLF,尚未出现高致病性的分子特征.HA片段共编码557个氨基酸,有9个潜在的糖基化位点,序列与2009年前WHO疫苗株A/NewCaledonia/20/1999(H1N1)、A/SolomonIslands/3/2006(H1N1)和/Brisbane/59/2007(H1N1)相比,分别有15、12和4处不同,这些差异分布在Sa、Sb、Ca1、Ca2、Cb 5个抗原决定簇的氨基酸差异分别有5、5和2处.该毒株在健康人群血清的抗体阳性率为34.33%,几何平均效价(GMT)为10.38.A/Shanghai/1167/2011(H1N1)是2011年出现在上海地区的一个季节性甲型H1N1流感病毒毒株,其抗原变异与既往季节性甲型H1N1流感病毒相比不大,但在以A(H1N1)pdm09为主要流行株的年份检测到散在发生的既往季节性甲型H1N1流感病毒毒株应当引起重视,其在人群中的抗体水平较低,易引起流行,需要提高对类流感人群中此种毒株的持续监测.     

广谱流感疫苗的研究进展

广谱流感疫苗是指能够诱导针对流感病毒的广谱中和抗体或广谱的细胞免疫反应,从而保护动物或人类免受多数流感病毒毒株感染的疫苗。流感病毒广谱中和抗体的发现及机制研究为广谱流感疫苗的研发提供了新的思路。同时,流感病毒细胞免疫方面的研究进展、佐剂以及免疫策略等的研究进展都极大促进了广谱流感疫苗的研发。本文从以上几个方面介绍广谱流感疫苗的研究进展,并对未来的应用进行评价及展望。     

Avian influenza virus

Recent outbreaks of highly pathogenic avian influenza in chickens and ducks that occurred in 9 Asian countries including Japan alarmed to realize that there is no border for infections and gave a rise to great concern for human health as well as for agriculture. This H5N1 virus jumped the species barrier and caused severe disease with high mortality in humans in Viet Nam and Thailand; 15 deaths of 22 cases and 8 of 12, respectively. A second concern was the possibility that the situation could give rise to another influenza pandemic in humans since genetic reassortment may occur between avian and human influenza viruses when a person is concurrently infected with viruses from both species. This process of gene swapping inside the human body can give rise to a new subtype of the influenza virus to which humans would not have immunity. The outbreaks also emphasized the need to continue active surveillance on avian influenza throughout the year to undertake aggressive emergency control measures as soon as an infection is detected.     

Galectin-1 binds to influenza virus and ameliorates influenza virus pathogenesis

Innate immune response is important for viral clearance during influenza virus infection. Galectin-1, which belongs to S-type lectins, contains a conserved carbohydrate recognition domain that recognizes galactose-containing oligosaccharides. Since the envelope proteins of influenza virus are highly glycosylated, we studied the role of galectin-1 in influenza virus infection in vitro and in mice. We found that galectin-1 was upregulated in the lungs of mice during influenza virus infection. There was a positive correlation between galectin-1 levels and viral loads during the acute phase of viral infection. Cells treated with recombinant human galectin-1 generated lower viral yields after influenza virus infection. Galectin-1 could directly bind to the envelope glycoproteins of influenza A/WSN/33 virus and inhibit its hemagglutination activity and infectivity. It also bound to different subtypes of influenza A virus with micromolar dissociation constant (K(d)) values and protected cells against influenza virus-induced cell death. We used nanoparticle, surface plasmon resonance analysis and transmission electron microscopy to further demonstrate the direct binding of galectin-1 to influenza virus. More importantly, we show for the first time that intranasal treatment of galectin-1 could enhance survival of mice against lethal challenge with influenza virus by reducing viral load, inflammation, and apoptosis in the lung. Furthermore, galectin-1 knockout mice were more susceptible to influenza virus infection than wild-type mice. Collectively, our results indicate that galectin-1 has anti-influenza virus activity by binding to viral surface and inhibiting its infectivity. Thus, galectin-1 may be further explored as a novel therapeutic agent for influenza.     

Protection of inactivated influenza virus vaccine against lethal influenza virus infection in diabetic mice

Influenza virus infection frequently causes complications and some excess mortality in the patients with diabetes. Vaccination is an effective measure to prevent influenza virus infection. In this paper, antibody response and protection against influenza virus infection induced by vaccination were studied in mouse model of diabetes. Healthy and diabetic BALB/c mice were immunized once or twice with inactivated influenza virus vaccine at various dosages. Four weeks after the first immunization or 1 week after the second immunization, the mice were challenged with influenza virus at a lethal dose. The result showed that the antibody responses in diabetic mice were inhibited. Immunization once with high dose or twice with low dose of vaccine provided full protection against lethal influenza virus challenge in diabetic mice, however, in healthy mice, immunization only once with low dose provided a full protection.     

A seven-segmented influenza A virus expressing the influenza C virus glycoprotein HEF

Influenza viruses are classified into three types: A, B, and C. The genomes of A- and B-type influenza viruses consist of eight RNA segments, whereas influenza C viruses only have seven RNAs. Both A and B influenza viruses contain two major surface glycoproteins: the hemagglutinin (HA) and the neuraminidase (NA). Influenza C viruses have only one major surface glycoprotein, HEF (hemagglutinin-esterase fusion). By using reverse genetics, we generated two seven-segmented chimeric influenza viruses. Each possesses six RNA segments from influenza virus A/Puerto Rico/8/34 (PB2, PB1, PA, NP, M, and NS); the seventh RNA segment encodes either the influenza virus C/Johannesburg/1/66 HEF full-length protein or a chimeric protein HEF-Ecto, which consists of the HEF ectodomain and the HA transmembrane and cytoplasmic regions. To facilitate packaging of the heterologous segment, both the HEF and HEF-Ecto coding regions are flanked by HA packaging sequences. When introduced as an eighth segment with the NA packaging sequences, both viruses are able to stably express a green fluorescent protein (GFP) gene, indicating a potential use for these viruses as vaccine vectors to carry foreign antigens. Finally, we show that incorporation of a GFP RNA segment enhances the growth of seven-segmented viruses, indicating that efficient influenza A viral RNA packaging requires the presence of eight RNA segments. These results support a selective mechanism of viral RNA recruitment to the budding site.     

Highly pathogenic avian influenza virus inducing influenza pneumonia in humans

The human disease caused by avian influenza virus in South Asia is a typical example of emerging infection resulting from transmission of a known causative agent to a new host. The first cases with a comparatively high level of lethality rates among patients were registered in 1997 and 1999. The situation is a special phenomenon in epidemiology which requires deep evolutionary and ecological analysis, as well as theoretical interpretation. With the example of avian influenza virus in Western Europe and South Asia in 2003-2004 the practical expediency of modern concepts "foci versus epidemics" and "eradication versus vaccination" is now evident.