流行性出血热病毒灭活后免疫原性的初步研究

本文用较高感染滴度的Vero-E6细胞和小白鼠乳鼠脑病毒,经56℃1小时加温或1:2,000福尔马林灭活后,免疫成年小白鼠、黑线姬鼠和长爪沙鼠。结果Vero-E6细胞培养的病毒均未使动物产生可测出的抗体,而小白鼠乳鼠脑病毒则获得良好的抗体阳转率(92％)。同时证明,热灭活较福尔马林灭活者好,加佐剂较不加佐剂好。     

单纯疱疹病毒感染细胞膜系统脂质分子流动性的变化

运用荧光偏振技术了解单纯疱疹病毒(HSV)I型Sm44株感染非洲绿猴肾(Vero)细胞后细胞膜系统脂质分子流动性的变化.结果表明,在所观察的感染后的5个时相中(感染后0、7、14、21、28小时),感染早期(感染后0、7小时)细胞膜的流动性变化不大;感染晚期(14至28小时)细胞膜的脂质流动性明显增强,与未感染病毒的对照组细胞有显著差异,这提示HSV感染晚期的装配,成熟与释放是与被感染细胞的生物膜系统密切相关的.本文对运用该生物物理技术研究病毒和宿主细胞相互关系的意义进行了初步讨论.     

单纯疱疹病毒感染细胞膜系统脂质分子流动性的变化

运用荧光偏振技术了解单纯疱疹病毒(HSV)I型Sm44株感染非洲绿猴肾(Vero)细胞后细胞膜系统脂质分子流动性的变化.结果表明,在所观察的感染后的5个时相中(感染后0、7、14、21、28小时),感染早期(感染后0、7小时)细胞膜的流动性变化不大;感染晚期(14至28小时)细胞膜的脂质流动性明显增强,与未感染病毒的对照组细胞有显著差异,这提示HSV感染晚期的装配,成熟与释放是与被感染细胞的生物膜系统密切相关的.本文对运用该生物物理技术研究病毒和宿主细胞相互关系的意义进行了初步讨论.     

四株单纯疱疹病毒(HSV)感染 四种细胞的SDS—PAGE分析

我们用 SDS—PAGE 测定四株 HSV 感染 HEp—2细胞、HeLa 细胞、人胚肺细胞、鸡胚细胞多肽与未感染细胞多肽的电泳图结果表明:四小时感染细胞多肽与未感染细胞多肽相比未见有区别,二十四小时的感染细胞多肽与未感染细胞多肽的电泳带显示有所区别。其中感染 HSV—1和 HSV—2型的鸡胚细胞多肽电泳带亦显示有所区别。说明鸡胚细胞对 HSV—1型和 HSV—2型的敏感性差异,亦可以从电泳带反映出来,说明电泳带可作为 HSV 分型的又一方法。经蔗糖梯度超离心纯化的未标记同位素的地方株 HSV—1CC21株和 HSV—2W 株病毒颗粒用 SDS 裂解,并进行 SDS—PAGE 表明这两株病毒的结构多肽分别为12和15种,显示型的区别。本文还就关于在制备疫苗时以选择何种细胞为宜的问题进行了讨论。     

流行性乙型脑炎病毒的变異II．不同毒株通过鸡胚组织培养后的毒力变異

1．流行性乙型脑炎不同毒栋通过鸡胚悬浮组织块连续传代50一100代后对小白鼠的神经外毒力逐渐下降,但其表现程度不完全一致,原来神经外毒力较低的SA4毒株则下降比较多,直至完全丧失。脑腔毒力的变化在不同毒株之同的差异更为明显;P3及Lm株经培养200代左右其毒力始格保持恒定,而SA4株毒力(LD50)则自181代以后开始下降1一2log。 2．对小白鼠脑腔毒力下降的SA4株对恒河猴脑腔致病力亦有一定的减弱,主耍表现在猴子于感染病毒后的发病溉伏期及病程比原毒株延长约一倍。 3．通过鸡胚组织块连续传代后的病毒对小白鼠的致病力和免疫力之间有一定的关系,SA4株当其脑腔毒力开始下降,腹腔毒力完全丧失后,则几乎完全失去了对小白鼠的 免疫性,而P3及Lm株的神经外毒力虽有下降,但仍然保持一定的致病力,因而仍然具有一定的免疫力。     

丝瓜提取物的抗乙型脑炎病毒感染作用

本文报道以乙型脑炎病毒皮下感染小白鼠为实验模型,观察丝瓜提取物(L 043)的抗感染作用。在感染病毒前腹腔注射L 043时,有很明显的预防作用,其保护率为60—30％;在感染病毒3．5小时后注射L 043时,其保护率(20—27％)较低。实验证明L 043对乙型脑炎病毒无直接灭活作用;对组织培养细胞、小白鼠、家兔,未见有明显的毒性。     

人巨细胞病毒AD169株感染家兔致病机理的初步研究

本文试用家兔作HCMV感染致病机制模型研究,用家兔荧光法及病毒再分离技术考证了感染期间的病毒血症动态。观察到兔在原发性病毒感染后的第13无病毒首先在单核细胞(MC)、淋巴细胞(LC)中显现,并向血浆排放病毒,进而随血道播散至全身组织,引起相应靶器官感染致病。     

乙型脑炎14-2株冻干活疫苗的生产研究

乙型脑炎病毒14-2株经地鼠肾细胞连续传23代,各代次的病毒滴度和回传乳鼠后的毒力都较稳定,与5-3株无差异。各代次的神经毒力和中枢神经外毒力也与5-3株相同。脑内感染12～14克小白鼠均不致死,皮下感染也不致病。用地鼠肾细胞经36℃培养4天,病毒增殖达到高峰,滴度为8.0～8.5logTCID50/0.2ml。病毒培养液的pH值为7.4～7.6时,病毒增殖高峰可持续3天。以1％明胶、5％蔗糖为保护剂,在冻干后无真空、不充氮的条件下。14—2株活疫苗在37℃可保存10天,室温(16～31℃)可保存4个月,5～8℃可保存一年,病毒滴度均无明显下降.冻干后充氮或不充氮病毒的滴度及稳定性看不出差异。冻干14—2株活疫苗融化后,用Eagle’s液稀释,置22～23℃8小时,滴度不变,若以生理盐水稀释,则可保持2～4小时;以蒸馏水稀释只能稳定2小时。     

单纯疱疹病毒与天然免疫系统的相互作用的研究进展

单纯疱疹病毒(Herpes simplex virus,HSV)包括1型和2型,属于人类疱疹病毒家族的α亚族,也是人类感染最常见的病原体之一,感染可导致多种疱疹性疾病,包括疱疹性眼炎及可引起失明的疱疹性角膜炎。HSV的致病机制与病毒和宿主天然免疫系统的相互作用密切相关,涉及到病毒逃逸干扰素反应,核转录因子κB(Nuclear factorκB,NF-κB)信号通路以及病毒与免疫细胞相互作用等多个环节。本文就近年来单纯疱疹病毒与天然免疫系统的相互作用的研究进展进行综述。     

蛇瓜提取物的抗病毒感染作用和诱生干扰素的活性

本实验证明,在流行性乙型脑炎病毒皮下感染前,给小白鼠注射蛇瓜提取物(简称E.T.a.)有明显的保护作用;在感染后给药,则小白鼠存活天数可明显延长,但其保护作用低。给家兔静脉注射E.T.a.,血清中可测到Ⅰ型干扰素,高峰出现在注射后2小时。经初步鉴定,蛇瓜提取物的有效成份似为某种双链RNA,但需进一步确证。     

禽流感H5N1病毒不同途径感染BALB/c小鼠的临床和病理比较

我们将禽流感H5N1病毒通过鼻腔、尾静脉和脑内接种方式接种BALB/c小鼠,观察小鼠在感染过程中临床症状和各个组织器官的病理变化。感染1天后,小鼠打堆、猥琐、毛色混乱、眼角有分泌物;感染第2天的小鼠均出现死亡;感染4天后,小鼠从临床症状上表现恢复。收集各种方式感染的小鼠的主要脏器进行组织病理学检查,结果显示,小鼠不同接种途径感染H5N1流感病毒后,所产生的病理变化相似;从各个脏器的病变程度来看,病毒最先到达的器官损伤最严重。感染第2天各组织器官的病变最严重,随着感染时间增长,肺脏和肾脏等器官有不同程度的恢复,心脏和肝脏等器官的病变在感染过程中未见明显恢复。     

Prolonged exercise suppresses antigen-specific cytokine response to upper respiratory infection.

Fatiguing exercise has been associated with an increased susceptibility to infection. This study examined the antigen-specific T-helper (Th) type 1 and Th type 2 cytokine response to herpes simplex virus (HSV) infection after an acute bout of fatiguing exercise. Male BALB/cJ mice ran on a treadmill (Ex) until voluntary fatigue (approximately 2.5 h), and control mice were handled and remained next to the treadmill. Mice were infected with HSV 20 min after exercise. Mice were killed 2 or 7 days postinfection, and sera and spleens were taken for the determination of HSV-specific serum IgM, splenocyte cytokine production during culture with HSV, and splenocyte natural killer cell cytotoxicity. Both Th type 1 [interleukin (IL)-2, interferon-gamma, IL-12] and Th type 2 (IL-10) cytokine production in spleen cell cultures, as well as natural killer cell cytotoxicity, decreased in Ex on day 2 postinfection. On day 7 postinfection, there was no difference in HSV-specific serum IgM or cytokine production by cells from control and Ex mice, with the exception of decreased IL-12 in Ex mice. These findings suggest that fatiguing exercise may alter the kinetics of antigen-specific cytokine production.     

Role of herpes simplex virus 1 Us3 in viral neuroinvasiveness

Us3 is a serine–threonine protein kinase that is encoded by herpes simplex virus 1 (HSV‐1). In experimental animal models of HSV infection, peripheral and intracranial inoculations can be used to study viral pathogenicity in peripheral sites (e.g., eyes and vagina) and central nervous systems (CNSs), respectively. In addition, peripheral inoculation can be used to investigate this virus' ability to invade the CNS (neuroinvasiveness) from peripheral sites. HSV‐1 Us3 has previously been shown to be critical for viral pathogenicity in both peripheral sites and CNSs of mice. However, the role of HSV‐1 Us3 in viral neuroinvasiveness has not yet been elucidated. In the present study, the yields of a Us3 null mutant virus and its repaired virus in the eyes, trigeminal ganglia, and brains of mice following ocular inoculation were examined. It was found that, although the repaired virus appeared in the brains of mice 3 days after infection, peak replication occurring 7 days after infection, no viral replication of the Us3 null mutant virus was detectable. These findings indicate that HSV‐1 Us3 plays a crucial role in the ability of the virus to invade the brain from the eyes. Thus, HSV‐1 Us3 is a significant neuroinvasiveness factor in vivo.     

Passive immunization of mice with monoclonal antibodies to glycoprotein gB of herpes simplex virus

To investigate the protective ability of monoclonal antibodies (MCAs) to viral glycoprotein in herpes simplex virus (HSV) infection, athymic nude mice were inoculated intracutaneously with HSV type-1 (HSV-1) in the midflank. Three hours after inoculation, one group of mice was passively immunized with one of a series of MCAs to glycoprotein gB of HSV-1, and a control group of mice was given phosphate buffered saline alone. The control mice died within 16 days after infection, whereas the mice passively immunized with any of the MCA showed suppressed development of skin lesions. Three of six mice given MCA failed to develop any visible lesions and no HSV could be isolated from the lumbar dorsal root ganglia of these mice 60 days after the challenge. BALB/c mice were also protected from infection with HSV type 2 by passive immunization with MCA to HSV-1 gB.     

Antiidiotype modulation of herpes simplex virus infection leading to increased pathogenicity.

Antiidiotype reagents that recognize idiotypic determinants associated with the combining site of monoclonal antibodies to herpes simplex virus type 2 ( HSV2 ) were used to manipulate the immune response to HSV2 in BALB/c mice. The injection of antiidiotype antibodies into mice before challenge with a 50% lethal dose of HSV2 resulted in a shorter survival time than that of mice receiving either preimmune rabbit immunoglobulin G or antiidiotype reagents against hepatitis B surface antigen before HSV2 challenge. These findings indicate that the immune response to HSV2 in mice can be modulated through idiotype- antiidiotype networks, thereby increasing the pathogenicity of HSV2 infections.     

CXCR2-/- mice show enhanced susceptibility to herpetic stromal keratitis: a role for IL-6-induced neovascularization

Ocular infection with HSV results in a blinding immunoinflammatory lesion known as herpetic stromal keratitis (HSK). Early preclinical events include inflammatory cell, mainly neutrophils, infiltration of the stroma, and neovascularization. To further evaluate the role of neutrophils in pathogenesis, HSV infection was compared in BALB/c and mice of the same background, but lacking CXCR2, the receptor for chemokines involved in neutrophil recruitment. Our results show clear differences in the outcome of ocular HSV infection in CXCR2-/- compared with control BALB/c mice. Thus, CXCR2-/- animals had minimal PMN influx during the first 7 days postinfection, and this correlated with a longer duration of virus infection in the eye compared with BALB/c mice. The CXCR2-/- mice were also more susceptible to HSV-induced lesions and developed HSK upon exposure to a dose of HSV that was minimally pathogenic to BALB/c mice. The basis for the greater HSK lesion susceptibility of CXCR2-/- mice was associated with an elevated IL-6 response, which appeared in turn to induce the angiogenic factor, vascular endothelial growth factor. Our results serve to further demonstrate the critical role of angiogenesis in the pathogenesis of ocular lesions.     

The cellular changes in primary and recurrent infection with herpes simplex virus type 2 in an in vitro model

The cellular changes of primary and recurrent herpes simplex virus (HSV) infection were investigated in in vitro models. In the primary infection model, nuclear changes were characterized by clumping and margination of the nuclear chromatin, a homogeneous ground-glass nuclear appearance, multinucleation and the appearance of two different types of intranuclear inclusions. One of the two types of inclusions appeared as early as six hours postinfection, reached a maximum at nine hours postinfection and gradually decreased thereafter. This early inclusion, relatively small in size as compared to the other type of inclusion, gave a basophilic staining when the Papanicolaou staining method was used. The other type of inclusion was the typical Cowdry type A inclusion, which appeared as early as 12 hours postinfection. Both types of inclusions contained clear perinuclear halo. In the recurrent infection model, the appearance of all the nuclear changes was delayed, the appearance of early inclusions was infrequent, and the Cowdry type A inclusions were observed more frequently than in the primary infection model. These results may indicate that the early inclusion is a sign of rapid virus replication while the Cowdry type A inclusion is one form of the remains of an HSV infection.     

Comparison of the Pathogenicity for Mice of Mycobacterium fortuitum and Mycobacterium abscessus

The pathogenicity for mice of 12 strains of Mycobacterium abscessus was compared with that for 8 strains of M. fortuitum. Both species caused lesions in kidneys and produced "spinning disease" resulting from inner ear infections. No major differences in pathogenicity of these two species were demonstrated. Strain to strain variation was marked, especially with M. abscessus. For example, 1.6 x 10(6) organisms of strain 11188 of M. abscessus produced death in four of five animals within 42 days, whereas strain 380 of M. abscessus failed to produce any deaths within 42 days. In the case of M. fortuitum, the greatest mortality observed was one of five animals, yet the incidence of spinning disease and kidney disease occurred earlier postinfection than in mice infected with M. abscessus. Histologically, abscess formation by a strain of M. abscessus was greater than by a strain of M. fortuitum, but this difference cannot be interpreted as a species difference.     

T regulatory cells contribute to the attenuated primary CD8+ and CD4+ T cell responses to herpes simplex virus type 2 in neonatal mice

The first weeks of life are characterized by immune tolerance and increased susceptibility to intracellular pathogens. The neonatal adaptive response to HSV is attenuated compared with adult control models in humans and mice. T Regulatory cells (Tregs) control autoimmunity and excessive immune responses to infection. We therefore compared Treg responses in the draining lymph nodes (LN) of HSV-infected neonatal and adult C57BL/6 mice with the effect of Treg depletion/inactivation by anti-CD25 (PC61) treatment before infection on Ag-specific T cell effector responses at this site. There was a small, but significant increase in the frequency of CD4(+)Foxp3(+) Tregs at day 3 postinfection (p.i.) in the LN of neonatal and adult mice, compared with age-matched mock-infected controls. Depletion of Tregs before HSV infection significantly enhanced HSV-specific CD8(+) T cell cytotoxicity in vivo, cell number, activation, and granzyme B expression 4 days p.i. only in neonatal mice, and significantly enhanced CD8(+) and CD4(+) T cell IFN-gamma responses in both infected adults and neonates. Treg depletion also reduced the titer of infectious virus in the draining LN and nervous system of infected neonates on days 2 and 3 p.i. Treg suppression of the neonatal CTL response p.i. with HSV was associated with increased expression of TGF-beta in the draining LN at day 4 p.i. compared with uninfected neonates, but IL-10 was increased in infected adults alone. These experiments support the notion that the newborn primary T cell effector responses to HSV are suppressed by Tregs.     

Modulation by Polypodium leucotomos extract of cytokine patterns in experimental trichomoniasis model

The immunomodulating effects of Anapsos, an aqueous hydrosoluble extract obtained from the rhizomes of the fern Polypodium leucotomos, on both pathogenicity and cytokine levels in serum (IFN-gamma/IL-4) were assayed in a Trichomonas vaginalis experimental model (BALB/c mice infected with 10(7) trichomonads and examined at day 15 after infection). Doses of 20 mg/kg/day administered for 10 days before the infection with the parasite induced a decrease of the experimental pathogenicity approximately 10-20% compared to controls. Gross histopathologic changes at abdominal organs and mortality rate, as a consequence of pathogenicity of the protozoa and the immune response of the host, were evaluated. IFN-gamma and IL-4 cytokines were determined on days -5, 0, 5, 10, and 15 postinfection by indirect ELISA. Treatment with PAL before infection modulates and downregulates the IFN-gamma concentration, while anticipates and upregulates the IL-4 level. The assays performed have showed the utility of the murine model of experimental trichomoniasis for the evaluation of immunomodulatory activity of synthetic or natural products.