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2015年30卷3期

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Review

Emerging complexity and new roles for the RIG-I-like receptors in innate antiviral immunity

John S. Errett, Michael Gale

2015, 30(3): 163 doi: 10.1007/s12250-015-3604-5

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Innate immunity is critical for the control of virus infection and operates to restrict viral susceptibility and direct antiviral immunity for protection from acute or chronic viral-associated diseases including cancer. RIG-I like receptors (RLRs) are cytosolic RNA helicases that function as pathogen recognition receptors to detect RNA pathogen associated molecular patterns (PAMPs) of virus infection. The RLRs include RIG-I, MDA5, and LGP2. They function to recognize and bind to PAMP motifs within viral RNA in a process that directs the RLR to trigger downstream signaling cascades that induce innate immunity that controls viral replication and spread. Products of RLR signaling also serve to modulate the adaptive immune response to infection. Recent studies have additionally connected RLRs to signaling cascades that impart inflammatory and apoptotic responses to virus infection. Viral evasion of RLR signaling supports viral outgrowth and pathogenesis, including the onset of viral-associated cancer.
Research Article

mTOR通过转录因子c-Fos调控TLR诱导的抗HBV和HCV的Th1反应

何丽, 臧爱萍, 杜旻, 马大鹏, 袁川评, 周纯, 穆静, 施焕静, 李大鹏, 黄栩林, 邓强, 肖建华, 鄢慧民, 惠利健, 蓝柯, 熊思东, 李晓霞, 黄忠, 肖晖

2015, 30(3): 174 doi: 10.1007/s12250-015-3606-3

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尽管IL-12在诱导Th1和CTL细胞免疫应答中起着非常关键的作用,但单独用TLR配体刺激树突状细胞和巨噬细胞时,只能诱导少量的IL-12表达,表明其表达水平受着严格的调控。通过本项研究,我们发现并阐述了TLR诱导IL-12表达,促进Th1应答的新机制。通过筛选蛋白磷酸激酶的小分子抑制剂库,我们发现了一些参与调节LPS诱导IL-12表达的蛋白激酶。通过进一步的研究,我们发现PI3K,ERK和mTOR是在天然免疫细胞中调节IL-12和IL-10的表达,从而调控机体的Th1免疫反应的关键激酶。此外,我们发现这些激酶都调控了TLR诱导转录因子c-Fos的表达,而且c-Fos通过与NF-κB相互作用,调节NF-κB与IL-12和IL-10启动子的结合,在转录水平上调控了IL-12和IL-10等基因的表达。此外,用mTOR的抑制剂rapamycin作为佐剂,可以有效地提升乙肝和丙肝疫苗的效果。综上所述,这些结果表明mTOR通过调控c-Fos,进而调控TLR诱导的IL-12和IL-10表达,影响了疫苗诱导的Th1免疫反应。

SOCS1在呼吸道合胞病毒持续感染中的表达抑制细胞固有免疫反应

郑君文, 杨璞, 唐燕, 赵东赤

2015, 30(3): 190 doi: 10.1007/s12250-015-3597-0

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呼吸道合胞病毒(RSV) 持续感染的HEp-2细胞是由不同生物学特性的细胞构成,包括了病毒抗原阳性和阴性的细胞亚群。目前对RSV持续感染时病毒复制和逃逸宿主免疫的机制尚不清楚。本研究对RSV持续感染细胞生物学特点和固有免疫反应的调节的机制进行了初步分析。首先分离克隆RSV持续感染的HEp-2细胞,并传代培养。采用siRNA沉默RIG-I或TLR3基因,检测SOCS1和STAT1/2在病毒持续感染的细胞中表达,ELISA检测培养上清细胞因子浓度,RT-PCR检测抗病毒基因的mRNA转录水平。共分离8个RSV持续感染的细胞克隆,在连续传代(克隆)过程中,每个克隆均会分化成病毒抗原阳性和阴性的两种截然不同的细胞群体。前者细胞内可检测病毒低水平复制,抵抗野生型RSV再次感染,分泌高水平的干扰素(IFN)-β,巨噬细胞炎性蛋白(Mip)-1α,白介素 (IL)-8。此外持续感染细胞表达高水平的TLR3, RIG-I和SOCS1。抑制TLR3 mRNA的表达可降低SOCS1蛋白质表达和细胞因子的分泌。结果提示,RSV持续细胞是一种炎症状态细胞群,而SOCS1高表达抑制了经TLR3激活的干扰素途径的抗病毒基因表达,与病毒的持久或潜伏感染密切相关,是流行爆发潜在的传染源。

Design of a heterosubtypic epitope-based peptide vaccine fused with hemokinin-1 against influenza viruses

2015, 30(3): 200 doi: 10.1007/s12250-014-3504-0

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Influenza viruses continue to emerge and re-emerge, posing new threats for public health. Control and treatment of influenza depends mainly on vaccination and chemoprophylaxis with approved antiviral drugs. Identification of specific epitopes derived from influenza viruses has significantly advanced the development of epitope-based vaccines. Here, we explore the idea of using HLA binding data to design an epitope-based vaccine that can elicit heterosubtypic T-cell responses against circulating H7N9, H5N1, and H9N2 subtypes. The hemokinin-1 (HK-1) peptide sequence was used to induce immune responses against the influenza viruses. Five conserved high score cytotoxic T lymphocyte (CTL) epitopes restricted to HLA-A*0201-binding peptides within the hemagglutinin (HA) protein of the viruses were chosen, and two HA CTL/HK-1 chimera protein models designed. Using in silico analysis, which involves interferon epitope scanning, protein structure prediction, antigenic epitope determination, and model quality evaluation, chimeric proteins were designed. The applicability of one of these proteins as a heterosubtypic epitopebased vaccine candidate was analyzed.

利用病毒样颗粒制备抗EV71的单克隆抗体

林涛, 鲜于灵芝, 吕颂雅*

2015, 30(3): 208 doi: 10.1007/s12250-015-3573-8

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手足口病是一种流行于五岁以下儿童的流行性疾病, 某些情况下,会导致患者死亡。其中肠道病毒71型C4株是手足口病的主要致病原。尽管目前没有治疗EV71的药物,但是一些研究表明,候选疫苗或病毒的衣壳蛋白可以引起抗EV71的免疫应答。本项研究中,我们利用了从酵母中得到的EV71的病毒样颗粒免疫雌性的BALB/c小鼠(6-8周)来筛选抗EV71的抗体。我们得到了2株能够分泌抗EV71的中和抗体的杂交瘤细胞。在100TCID50病毒的感染情况下,25ug/ml(最低中和浓度)的纯化后的单克隆抗体(D4和G12)能够保护95%的宿主细胞不被病毒感染,同时这些抗体被证实能够结合于EV71的衣壳蛋白VP1。目前的研究确定了这些中和抗体可以用来发展治疗EV71的人源化抗体,以及诊断试验和病毒学研究的理想试剂。
Letter

Cellular signaling in sinecatechins-treated external genital and perianal warts: unraveling the mechanism of action of a botanical therapy

2015, 30(3): 214 doi: 10.1007/s12250-014-3520-0

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Taken together with our previous report, current results provide cursory evidence that the sinecatechins’ mechanism of action likely entails some degree of modulation of inflammatory and apoptotic processes, in particular, the NF-κB-pathway. It is possible that sinecatechins also upregulate the host immune response, since the only gene to be significantly upregulated in VR is the pro-lymphocytic IL2; however, further studies are needed to examine the exact pattern of immune regulation. Moreover, since no DEGs were identified in VNR in the prior report, the present study provides putative insight into the expression patterns of patients who are not responsive to sinecatechins.

New prevalence estimate of Torque Teno virus (TTV) infection in healthy population and patients with chronic viral hepatitis in Jiujiang, China

彭进, 方越越, 赵学森, 彭宜红

2015, 30(3): 218 doi: 10.1007/s12250-014-3531-x

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Our results suggest that TTV is extremely common in the general Chinese population, including infants, providing further evidence that TTV is highly prevalent in the general population worldwide(Mancuso et al., 2013). Furthermore, the overall prevalence of TTV infection based on UTR-PCR in healthy infants, healthy adults, and patients with liver disease was almost 2–3 times higher than that obtained by N22-PCR(p < 0.01, Table 1), which highlights both the considerable influence of the PCR primers on the detection of TTV DNA and the benefits of using UTR-PCR to establish the true overall prevalence of TTV infection. On the other h and , our results also indicate the lack of a clear association of TTV infection with chronic viral hepatitis, which strongly suggests that TTV has little potential for causing hepatitis. However, the virulence and pathogenesis of the different genotypes or strains of TTV remain unclear and require further investigation.

Cats as a potential source of emerging influenza virus infections

2015, 30(3): 221 doi: 10.1007/s12250-015-3580-9

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Based on the findings of the present study, we conclude that cats can be infected with human influenza viruses as well as avian influenza viruses. Actually, the recent study has shown that both human-type(α2, 6-linked sialic acid) and avian-type(α2, 3-linked sialic acid)influenza virus receptors were extensively detected in the respiratory organs such as trachea, bronchus, and lung of the domestic cats(Wang et al., 2013). Therefore, cats may act as a vector for human influenza virus transmission within households, posing a potential public health concern. Furthermore, we detected both H5N1 and human virus-seropositive cats in neighboring areas at similar sampling times, suggesting that cats can be simultaneously infected with both avian and human viruses in H5N1 virus-endemic areas. Thus, cats, like pigs, may act as an intermediate host for the emergence of new, potentially p and emic viruses.

牛免疫缺陷病毒R29-127毒株中存在反义转录的证据

刘彬, 赵学潮, 沈文远, 孔晓红*

2015, 30(3): 224 doi: 10.1007/s12250-015-3559-6

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反转录病毒家族的多个成员被发现存在反义转录的现象,包括人T细胞白血病病(human T cell leukemia virus, HTLV)、人免疫缺陷病毒 (human immunodeficiency virus, HIV) 以及猫免疫缺陷病毒 (feline immunodeficiency virus ,FIV) 等。然而在其他种属的反转录病毒中反义转录是否普遍存在目前并不清楚。在本研究中,我们检测了牛免疫缺陷病毒 (bovine immunodeficiency virus ,BIV) 的反义转录。对BIV的R29-127毒株前病毒的反义序列进行分析,通过软件预测发现在3' LTR和跨膜糖蛋白区域各存在一个潜在的转录起始位点。转录其实位点下游可以预测到一个poly A信号。在转录起始位点和poly A信号之间存在一个长度为408 bp开放阅读框(Open Reading Frame, ORF)。然后通过单链特异性反转录PCR检测BIV的反义转录本。结果表明在BIV127前病毒转染后的293T细胞中能够检测到反义转录本的存在。为了研究BIV感染的细胞是否能够发生反义转录,我们在293T细胞中转染pBIV127感染克隆进行BIV病毒的包装,然后对BIV许可细胞系BIVE进行感染。结果发现在被感染的BIVE细胞中也可以检测到反义转录本的存在。此外,我们将反义ORF克隆到pEGFP-C1载体上的EGFP基因下游进行融合表达,并在HeLa细胞中观察融合蛋白的分布。结果发现融合蛋白主要分布在细胞质中,并以非均匀的特征分布在细胞核周围。本研究为BIV反义转录的存在提供了证据。

兔病毒性出血热病毒SYBR实时定量PCR检测方法的建立及病毒在实验感染兔组织的分布

刘文凯, 党如意, 王兴龙*

2015, 30(3): 228 doi: 10.1007/s12250-015-3560-0

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In this study, we developed a real-time PCR assay for RHDV detection and quantitation. With this method, the distribution of RHDV in the internal organs and body fluids of infected rabbits was analyzed. The highest viral RNA load was found in the spleen, followed by that in the liver. The virus is shed mainly through the oral, nasal, and urethral routes. Through testing of clinical samples, we found that most farmed rabbits that had been immunized had low levels of RHDV in their tissues. These findings may be useful for further research on the pathogenic mechanism of RHDV and potential RHDV receptors in organs.

Characterization of an infectious pancreatic necrosis virus from rainbow trout fry (Onhorhynchus mykiss) in West Ukraine

2015, 30(3): 231 doi: 10.1007/s12250-014-3513-z

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In this report, an aquatic birnavirus was isolated from rainbow trout fry, Onhorhynchus mykiss, during a fish health inspection in fish-farms in the west region of Ukraine near the rivers Siret and Cheremosh(Chernivtsi region). Preliminary examination of infected fish revealed a range of lesions, particularly in pancreatic tissue. Morphological changes, such as vacuole enlargement and cell rounding, were caused by viruses in appropriate cell lines. Investigation by electron microscopy demonstrated that the isolated virus was ultrastructurally similar to infectious pancreatic necrosis virus(IPNV). In addition, the nucleotide sequences of the 1120 bp VP2 gene fragments were analyzed and the identity of the isolated IPNV to strain Sp was revealed. The identity of nucleotide sequences was 97%–99%; the isolates of Sp strains most closely related to the Ukrainian IPNV isolate were found in Norway.
PERSPECTIVE

Are developing countries prepared to face Ebola-like outbreaks?

2015, 30(3): 234 doi: 10.1007/s12250-015-3564-9

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Many developing countries lack a functional health care system, especially African countries. Such a system is not only important to monitor emergencies but also to develop an integrated response; otherwise, any epidemic can spread rapidly(as was observed in the case of Ebola) and pose huge problems worldwide. According to WHO reports, the basic health care system in most countries affected by Ebola is very fragile(http://www.who.int/healthsystems); substantial investments are needed to improve the infrastructure and bring reforms. This is not only important for African countries; in fact, most developing countries need additional investments to improve the healthcare sector.