Respiratory syncytial virus infection in immunocompetent and immunocompromised murine

The purpose of this study is to distinguish respiratory syncytial virus ( RSV) infection and immunology between immunocompetent and immunocompromised murine and to explore immune mechanism of RSV infection. At various time points after RSV infection of BALB/c mice and nude mice, pulmonary viral titers were assayed, RSV antigen was tested by direct immune-fluorescent assay and immu nohistochemistry. Pulmonary mRNA expressions of Toll like receptor (TLR)2 and TLR4 were assayed by RT-PCR. CD4 cells and CD8 cells in peripheral blood were examined by flow cytometry and plasma total IgE was assayed by ELISA. Leukocytes in bronchoalveolar lavage fluid (BALF) and pulmonary histology were identified to reflect airway inflammation. It was found that RSV titers of both mice peaked on the 3rd day post infection with a much higher level of viral titer in nude mice than in BALB/c mice and a longer viral duration in nude mice (over 9 days post infection) than in BALB/c mice (6 days post infection). RSV infection induced higher viral antigen expression in nude mice (0.267±0.045) than in BALB/c mice (0.168±0.031). RSV infection enhanced pulmonary TLR4 expression of BALB/c mice (51.96%±11.34%) and nude mice (48.96%±12.35%) compared with each control (34.04%±10.06% and 32.37%±9.87% respectively). CD4 peripheral blood cells increased in RSV infected BALB/c mice (66.51%±2.09%) compared with the control BALB/c mice (51.63%±5.90%), and CD4 cells and CD8 cells were deficient in nude mice. RSV infection increased plasma total IgE in both mice, and BALB/c mice had a larger amount of IgE on the 7th day post infection (9.02 ng/ml±2.90 ng/ml) and on the 14th day post infection (12.76 ng/ml±4.15 ng/ml) than corresponding nude mice (3.72 ng/ml±1.06 ng/ml and 7.62 ng/ml±3.08 ng/ml respectively on the 7th and 14th day post infection). RSV infected nude mice had more severe airway inflammation than infected BALB/c mice. It is concluded that BALB/c mice and nude mice presented similar RSV infectious characteristics. However, infection of nude mice showed higher viral titer with longer duration and more severe airway inflammation, lower level of plasm total IgE and CD4 peripheral blood cells, but the similar pulmonary TLR4 expression with BALB/c mice.     

Technetium-99m-labeled annexin V imaging for detecting prosthetic joint infection in a rabbit model

Accurate and timely diagnosis of prosthetic joint infection is essential to initiate early treatment and achieve a favorable outcome.In this study,we used a rabbit model to assess the feasibility of technetium-99m-labeled annexin V for detecting prosthetic joint infection.Right knee arthroplasty was performed on 24 New Zealand rabbits.After surgery,methicillin-susceptible Staphylococcus aureus was intra-articularly injected to create a model of prosthetic joint infection(the infected group,n = 12).Rabbits in the control group were injected with sterile saline(n=12).Seven and 21 days after surgery,technetium-99m-labeled annexin V imaging was performed in 6 rabbits of each group.Images were acquired 1 and 4 hours after injection of technetium-99 mlabeled annexin V(150 MBq).The operated-to-normal-knee activity ratios were calculated for quantitative analysis.Seven days after surgery,increased technetium-99m-labeled annexin V uptake was observed in all cases.However,at 21 days a notable decrease was found in the control group,but not in the infected group.The operated-to-normal-knee activity ratios of the infected group were 1.84 ± 0.29 in the early phase and 2.19 ±0.34 in the delay phase,both of which were significantly higher than those of the control group(P=0.03 and P=0.02).The receiver operator characteristic curve analysis showed that the operated-to-normal-knee activity ratios of the delay phase at 21 days was the best indicator,with an accuracy of 80%.In conclusion,technetium-99m-labeled annexin V imaging could effectively distinguish an infected prosthetic joint from an uninfected prosthetic joint in a rabbit model.     

CD59 Silencing via Retrovirus-Mediated RNA Interference Enhanced Complement-Mediated Cell Damage in Ovary Cancer

CD59, belonging to membrane complement regulatory proteins （mCRPs）, inhibits the cytolytic activity of complement and is over-expressed in solid cancers, including ovary cancer. The aim of the present study was to construct recombinant retrovirus encoding shRNA targeted human CD59 and infect A2780 cells in order to investigate the relationship between decreased CD59 expression and tumorigenesis of ovary cancer, siCD59 and siCD59-C were successfully constructed and identified by PCR, restriction endonuclease analyses and DNA sequencing, respectively. The siCD59 was able to efficiently infect A2780 cells, which was confirmed by Western blotting. When incubated with fresh normal human serum （8%, v/v） for 1 h at 37℃, the cell viability was decreased and cell damage was increased in siCD59 infected A2780 cells compared to siCD59-C infected cells. This led to the activation of caspase-3. The apoptosis in siCD59 infected cells was shown with hypercondensed nuclei using Hoechst staining. Meanwhile, the weight of ovary tumor graft in nude mice was significantly decreased in siCD59 group compared to that of siCD59-C group. And the expression of CD59 protein in tumor tissue in siCD59 group was significantly decreased. These results suggested that CD59 silencing in ovary cancer cells v/a retrovirusmediated RNAi can enhance complement-mediated cell damage, inhibiting growth of ovary cancer. CD59 might be a potential target for gene therapy in ovary cancer. Cellular ＆ Molecular Immunology.     

红花多糖对小鼠Lewis肺癌移植瘤生长及转移的影响

Objective To investigate the inhibitory effects of safflower polysaccharide (SPS) on tumor growth and metastasis of T739 mice transplanted with Lewis lung carcinoma cells, then explore the anti-tumor mechanism of action. Methods The xenografts derived from Lewis lung carcinoma cells were established in T739 mice. Inoculated mice were randomly divided into control group, low dose SPS group, moderate dose SPS group and high dose SPS group, daily oral administration, a total of 28 times. Tumor volume and weight were measured and lung metastatic foci of the mice were observed under microscope. MTT assay was used to investigate both the inhibitive effect of SPS on Lewis lung carcinoma cells and the influence on proliferation of T lymphocytes. Results There was no inhibitory effect of SPS on Lewis lung carcinoma cells in vitro. In vivo, SPS could promote proliferation of T lymphocytes and inhibit tumor growth and metastasis. There were significant differences when comparing the SPS groups with control group in terms of the volume, weight and lung metastatic foci of the transplanted tumor ( P ＜ 0.05 ). Conclusion SPS could inhibit the tumor growth and metastasis.The anti-tumormechanism of action was about related to increase the immunological function of mice.[Key words] Safflower Polysaccharide; Lewis lung carcinoma; Transplanted tumor; Lung metastatic     

不同年龄人群感染甲型H1N1流感病毒临床特征分析

Objective To describe the feature of different age patients with A-H1N1.Methods Cross-sectional study was performed in 95 patients who were confirmed to be infected with A-H1N1 from May,2009 to July,2009,in according to their age.Results The average age of patients with A-H1N1 infection was 23.44±14.73.Accumulative prevalence in children and young adult reached 74.7% of total patients.There was a trend that the subclinical infection rate raised gradually from 0-15 years group to over 45 years group.The percent of lymphocyte in 0-15 years group was significantly higher than other age groups,P=0.039.The average time of virus shedding were 6.5±2.10 days (from 2 days to 12 days),and there were no significant difference in diverse age groups,P=0.272.13 out of 95(13.7%)patients presented complications related with A-H1N1 infection,and 4 of 6 patients complicated with pneumonia were in the 0-15 years group.Conclusion The distribution of age in A-H1N1 infection is markedly different from seasonal influenza,with more cases in school children and young adults and fewer cases in older adults. Flu-like symptoms in children were apparent and pneumonia was the major complication in children.     

不同年龄人群感染甲型H1N1流感病毒临床特征分析

Objective To describe the feature of different age patients with A-H1N1.Methods Cross-sectional study was performed in 95 patients who were confirmed to be infected with A-H1N1 from May,2009 to July,2009,in according to their age.Results The average age of patients with A-H1N1 infection was 23.44±14.73.Accumulative prevalence in children and young adult reached 74.7% of total patients.There was a trend that the subclinical infection rate raised gradually from 0-15 years group to over 45 years group.The percent of lymphocyte in 0-15 years group was significantly higher than other age groups,P=0.039.The average time of virus shedding were 6.5±2.10 days (from 2 days to 12 days),and there were no significant difference in diverse age groups,P=0.272.13 out of 95(13.7%)patients presented complications related with A-H1N1 infection,and 4 of 6 patients complicated with pneumonia were in the 0-15 years group.Conclusion The distribution of age in A-H1N1 infection is markedly different from seasonal influenza,with more cases in school children and young adults and fewer cases in older adults. Flu-like symptoms in children were apparent and pneumonia was the major complication in children.     

Expression of intercellular cell adhesion molecule 1 and matrix metalloproteinase 9 in endometrium of pregnant mice with long-term exposure to low concentration of sevoflurane北大核心

BACKGROUND: Epidemiological studies have found that female workers who are exposed to waste sevoflurane for a long time have reduced fertility and increased incidence of abortion and fetal deformity. OBJECTIVE: To imitate the working environment of long-term exposure to waste sevoflurane and investigate the mechanism of embryo implantation disorder induced by low-concentration sevoflurane exposure by observing the expression of intercellular adhesion molecule 1, integrin β1, P-selectin, and matrix metalloproteinase 9 in endometrium of pregnant mice exposed to low concentrations of sevoflurane, attempting to provide a basis for clinical safe drug use and occupational protection and to lay a foundation for further research on the mechanism of inhalation anesthetics on embryo implantation at gene and molecular levels. METHODS: Forty female Kunming mice and sixteen male Kunming mice, aged 6 weeks, weighting (20±2) g, were caged separately. Forty female mice were randomly divided into experimental group and control group. Mice in the experimental group were exposed to 0.1% sevoflurane, 6 hours per day, while those in the control group were exposed to the air. Thirty days later, the female mice in estrus were caged with mature male mice at a rate of 2:1. Whether the female mice were pregnant was observed at 7:00 am on the second day after mating. The pregnant mice were kept independently in the original condition (n ≥ 8 pregnant mice in each group). Mouse uterus on day 4.5 of gestation was removed and sliced for histological observation. The expression of intercellular adhesion molecule 1, integrin β1, P-selectin, and matrix metalloproteinase 9 in endometrial tissue was detected by immunohistochemistry and statistically analyzed. The average integrated absorbance value of positive reactants was calculated. RESULTS AND CONCLUSION: In the experimental and control groups, 17 and 15 female mice were respectively found in estrus; 12 and 10 female mice respectively were found vaginal plugs after mating. Five and two female mice were found pseudopregnant on day 4.5 of gestation in the experimental and control groups, respectively. Therefore, there were 7 and 8 pregnant mice in the experimental and control groups, respectively, and the vacant value in the experimental group was replaced by the average value obtained in the same group, which would be subsequently used in the controlled trial. Immunohistochemical results showed that the endometrium (glandular epithelial cells, luminal epithelial cells and stromal cells) of all pregnant mice were positively stained brownish yellow, but the expressions of intercellular adhesion molecule 1, integrin β1, P-selectin, and matrix metalloproteinase 9 (0.019±0.007, 0.017±0.007, 0.015±0.005, 0.012±0.005) in the experimental group were significantly lower than those in the control group (0.032±0.014, 0.025±0.008, 0.021±0.007, 0.023±0.005) (P < 0.05). All these findings indicate that long-term exposure to 0.1% sevoflurane may affect the adhesion of mouse embryos to endometrium and the embryo implantation, which may be related to the inhibitory effects of sevoflurane on the expressions of intercellular adhesion molecule 1, integrin β1, P-selectin, and matrix metalloproteinase 9, causing the imbalance in endometrial immunoregulation, the invasion of trophoblast cells into the endometrium and the inhibition of endometrial decidualization, and miscarriage and bleeding due to damaged vascular endothelium. © 2023, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

HBV基因型与HBV感染慢性化、重症化的关系

Objective To explore the association between HBV genotype and chronic/severe liver disease with HBV infection in Chinese patients.Methods Serum samples were collected from 2922 patients with HBV infection.HBV genotyping was performed with type-specific primers polymerase chain reaction,and the virological and biochemical markers were detected,which differences in the genotypes between various clinical types of HBV infection and liver function and virological markers between various HBV genotyping were analyzed.Results The genotype B,C,BC combinations,D of 2922 patients with HBV infection accounted for 15.9%,83.5%,0.41%,0.21% respectively.The ratio of genotype B in acute hepatitis group was higher(P=0.003),which the ratio of genotype C in the cirrhosis group and the hepatocellular carcinoma group was higher(P=0.000,0.000).The difference in ratio of genotype C was not statistically significant between acute-on-chronic liver failure group and chronic hepatitis group.HBeAg-positive rate,viral load and liver function markers of B,C genotype group in acute hepatitis group and chronic hepatitis group were not significant different.HBeAg-positive rates of genotype C in acute-on-chronic liver failure group,cirrhosis group,hepatoeellular carcinoma group were higher than that of genotype B(P=0.000,0.024,0.003).Viral load of genotype C in hepatocellular carcinoma group was higher than that of genotype B(P=0.025).Cholinesterase levels of genotype C in the acute-on-chronic liver failure group and the hepatocellular carcinoma group was lower than that of genotype B(P=0.0004、0.02).Conclusion There were HBV genotype B,C,B/C combinations and D in Chinese patients with HBV infection,with genotype B and C being the major ones.Compared with HBV genotype B,genotype C in Chinese patients with HBV infection was more likely to chronic infection,evolved to cirrhosis and hepatocellular carcinoma, but genotype difference was not observed in occurrence of acute-on-chronic liver failure.Genotype was not significant effect in acute and chronic hepatitis B,but HBeAg-positive rate/viral load was higher and liver damage was more severe in severe and end-stage genotype C HBV infection patients.     

肺炎链球菌菌体蛋白LytA和CbpA对感染小鼠的诊断价值

目的 通过基因工程技术获取肺炎链球菌自溶素(autolysin,LytA)和胆碱结合蛋白A(choline binding protein A,CbpA),探讨其对肺炎链球菌感染小鼠的血清学诊断价值.方法 根据GenBank中lytA和cbpA基因保守序列设计合成特异性引物,采用PCR技术从肺炎链球菌基因组中扩增lytA和cbpA保守序列片段;经由IPTG诱导并通过等电点洗脱方法获取纯化的重组蛋白LytA和CbpA;Western blot测定表达蛋白的抗体结合活性.以LytA和CbpA为抗原,建市ELISA反应模式测定感染小鼠血清中相应的IgG和IgM抗体,评价诊断价值.结果 成功构建了原核表达载体pET-32a(+)/lytA和pET-32a(+)/cbpA,表达的重组蛋白LytA和CbpA具有较好的抗体结合活性.实验组小鼠(肺炎链球菌感染组)IgM和IgG类抗LytA抗体滴度高于对照组(止常对照组和乙型链球菌感染对照组),差异有统计学意义(P<0.05),CbpA抗体与正常对照组差异有统计学意义,与乙型链球菌感染组差异无统计学意义(P>0.05).CbpA蛋白对感染小鼠诊断的敏感性较高(IgG:83.3%;lgM:75.0%),而LytA特异性较高(IgG:100%;IgM:100%).结论 协同利用重组蛋白CbpA诊断敏感性及LytA的特异性,对肺炎链球菌感染小鼠的血清学诊断具有一定价值,为进一步用于临床检验奠定基础.

Abstract：

Objective To obtain the pneumococcal autolysin(LytA)and choline binding protein A(CbpA)by prokaryotic expression system and investigate their diagnosis for infection caused by Streptococcus pneumoniae.Methods The specific primers were designed according to lytA and cbpA of Streptococcus pneumoniae gene sequence.lytA and cbpA were amplified by PCR form the pneumococcus genome.After IPTG inducing,the recombinant proteins were purified by electroeluting of bag filter,detected by SDS-PAGE and Western blot.Serum lgG and IgM antibodies accordingly of BALB/c mice infected with Streptococcus pneurnoniae were detected by ELISA.Results The recombinant plasmid pET-32a(+)/lytA and pET-32a (+)/cbpA were constructed successfully.Fusion proteins LytA and CbpA were expressed and displayed expected antigenicity.IgM and IgG antibodies level anti LytA were significantly higher than the control group (infections with B Streptococcus group and healthy mice),(P<0.05),but antibodies level anti CbpA did not increase as compared with group infected with B Streptococcus(P>0.05).Diagnostic sensitivity of CbpA was 83.3%(IgG)and 75.0%(IgM).Diagnostic specificity of LytA was 100%(IgG and IgM).Conclusion The synergistic use of specificity of LytA and sensitivity of CbpA may be worthy of serological diagnosis for Streptococcus pneumoniae infection,and may be used for further clinical test.     

Huangqin decoction regulates autophagy to intervene with intestinal acute graft-versus-host disease in mice北大核心

BACKGROUND: How to use traditional Chinese medicine to intervene the imbalance of autophagy after intestinal mucosal barrier injury, so as to ultimately intervene the occurrence of gastrointestinal acute graft-versus-host disease, is an urgent problem to be solved after hematopoietic stem cell transplantation. OBJECTIVE: To verify the precise mechanism by which Huangqin Decoction interferes with acute intestinal graft-versus-host disease. METHODS: CB6F1 mice were randomly divided into normal control group, model control group, low-dose Huangqin Decoction group, medium-does Huangqin Decoction group and high-does Huangqin Decoction group, with 16 mice per group. CB6F1 mice in the model control group, low-dose Huangqin Decoction group, medium-does Huangqin Decoction group and high-does Huangqin Decoction group were infused with mononuclear cell suspension (bone marrow cell 8×107 + spleen cell 8×107) obtained from Balb/c mice via caudal vein within 4 hours after60Co whole body irradiation (radiation dose was 8 Gy). Different concentrations of Huangqin Decoction were given by gavage on the same day after modeling. The rats in the model control group and the normal control group were given the same volume of normal saline by gavage for 15 days. Eight hours after the last gavage, the small intestine tissues of six mice in each group were collected. PCR and western blot assay were used to detect the expression levels of LC3II/I, Beclin1 and P62. The pathological grading of small intestinal mucosa was scored by hematoxylin-eosin staining. The autophagic vesicle structure of small intestinal mucosal epithelial cells was observed by transmission electron microscope. The remaining 10 rats in each group (except the normal control group) were used to observe the clinical grading of acute graft-versus-host disease and record the survival time. RESULTS AND CONCLUSION: (1) After the application of Huangqin Decoction, the survival time of mice was significantly prolonged; the clinical acute graft-versus-host disease score was significantly decreased, and the pathological grading score of small intestinal mucosa was significantly decreased. The score of medium-does Huangqin Decoction group and high-does Huangqin Decoction group was significantly lower than that of model control group, but there was no significant difference between medium-does Huangqin Decoction group and high-does Huangqin Decoction group. (2) The LC3II/I and Beclin1 expression was significantly lower in the model control group than that in the normal control group (P < 0.01), and P62 expression was significantly higher than that in the normal control group (P < 0.01). Huangqin Decoction could promote the recovery of LC3II/I and Beclin1 levels and downregulate p62 levels (P < 0.01). (3) Under transmission electron microscope, the number of autophagic vesicles in the treatment group was significantly higher than that in the model control group, accompanied by the recovery of important organelles such as mitochondria. (4) The results confirm that by interfering autophagy related proteins, Huangqin Decoction can promote the recovery of autophagy in acute graft-versus-host disease, protect intestinal mucosal barrier and reduce intestinal rejection after transplantation and has promise as a new treatment for acute graft-versus-host disease. © 2022, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

Characterization of glycosylphosphatidylinositol-anchored decay accelerating factor (GPI-DAF) and transmembrane DAF gene expression in wild-type and GPI-DAF gene knockout mice using polyclonal and monoclonal antibodies with dual or single specificity

Decay-accelerating factor (DAF, CD55) is a glycosylphosphatidylinositol (GPI)-linked membrane inhibitor of complement activation. While human and other mammalian species contain only one DAF gene, two distinct DAF genes, referred to as GPI-DAF and transmembrane (TM)-DAF, respectively, have been identified in the mouse. Using several independently generated monoclonal and polyclonal antibodies, either with dual or single specificity for GPI-DAF and TM-DAF gene products, we have examined the expression of the two DAF genes in tissues of the wild-type and a strain of knockout mouse whose GPI-DAF gene has been inactivated. By fluorescence-activated cell sorting (FACS) analysis, we found that DAF protein is present on the wild-type mouse erythrocytes and lymphocytes but no signal was detectable on the same cells of GPI-DAF gene knockout mice. Both T and B lymphocytes and splenic macrophages express the GPI-DAF gene but the expression level is higher on B lymphocytes than on T lymphocytes. Within the T cell population, both CD4+ and CD8+ T cells are positive. DAF protein was detected by immunohistochemistry at high levels on wild-type mouse spermatids and mature sperm. In contrast, only mature sperm stained positive in the GPI-DAF gene knockout mouse testis, suggesting that GPI-DAF but not the TM-DAF gene is expressed on spermatids. Examination of the fetoplacental unit at the day 7.5 stage revealed that GPI-DAF but not the TM-DAF gene is expressed in the maternal decidua cells surrounding the trophoectoderm of the embryo. No DAF expression was detected on trophoblast or the embryo proper. These findings suggest that although the TM-DAF gene is irrelevant on mouse blood cells, the two DAF genes may have different roles in germ cell development and/or mature sperm function. Because complement receptor 1-related gene/protein y (Crry) has been shown to be expressed on early mouse embryos, the complete lack of GPI-DAF and TM-DAF gene expression in early mouse development may explain the observed sensitivity of Crry-deficient embryos to maternal complement attack.     

重组人白介素2对小鼠弓形虫垂直传播的影响

目的　研究rhuIL - 2对弓形虫垂直传播的影响。方法　用不同剂量rhuIL - 2处理感染弓形虫孕鼠 ,于妊娠第12天取孕鼠子宫 ,观察记录平均活胎率的变化 ,并将胚胎及胎盘固定 ,做免疫组化染色 ,观察胎盘胎鼠感染情况 ,计算孕鼠的垂直传播率。结果　高剂量组与感染组相比活胎率明显升高 (P <0 0 5 ) ;两处理组的垂直传播率均较感染组明显下降 (P<0 0 5 ) ;免疫组化结果显示 ,胎盘滋养层细胞、绒毛间隙、蜕膜细胞均有大量弓形虫抗原表达 ;部分孕鼠的胎盘有弓形虫抗原表达 ,但其胚胎组织弓形虫抗原的表达却呈阴性。结论　rhuIL - 2一定程度上可以降低弓形虫的垂直传播率 ,对感染孕鼠及胚胎起到一定保护作用。胎盘作为局部免疫屏障具有重要的研究价值。     

紫外线减毒弓形虫免疫对小鼠旋毛虫感染肌肉组织病理的影响

目的探讨紫外线减毒弓形虫免疫对小鼠旋毛虫感染肌肉期组织病理的影响。方法ICR小鼠分为两组，实验组为紫外线减毒弓形虫免疫3周后再感染旋毛虫，对照组为旋毛虫单独感染。结果与旋毛虫单感染相比，小鼠预先经紫外线减毒弓形虫免疫再感染旋毛虫后23d，其肌肉期幼虫周围的炎症细胞浸润明显减轻，肌肉组织的病理损伤也明显减轻。结论紫外线减毒弓形虫免疫对小鼠旋毛虫感染肌肉期的组织病理有一定的免疫调节作用。     

CD8+ T cells are the major lymphocyte subpopulation involved in the protective immune response to Toxoplasma gondii in mice.

The ability of the major T cell subsets to adoptively transfer resistance to T. gondii infection was studied. Spleen cells harvested from mice with a 3-month T. gondii infection and cells from uninfected mice were enriched for T cells by nylon/wool purification. Adoptive transfer of these cells from both groups of donor mice led to a significant increase in the survival of syngeneic recipient mice infected intraperitoneally with 20 T. gondii cysts. Increased survival was mediated particularly by CD4-depleted but also, to a lesser extent, CD8-depleted subpopulations. These results were confirmed in T cell reconstituted athymic nude mice. Unfractionated T cells from chronically infected donors produced a significant inhibition of cyst formation in the brains of recipient mice measured 10 weeks after infection compared with control mice. The inhibition of cyst formation was ablated by pretreating T cells with anti-CD8 antibody and complement, but not anti-CD4 antibody and complement. Mice receiving cells from infected donors produced an early increase in their IgG1 and IgG2a antibody titres compared with mice given cells from uninfected animals. The depletion of either CD8+ or CD4+ immune cells appeared to have little effect on the antibody responses in recipient mice and there was no correlation between antibody levels and immunity. The results indicate that CD8+ T lymphocytes from convalescent T. gondii-infected BALB/c mice are the principal mediators of resistance to T. gondii, although CD4+ T cells appear to be involved during the acute phase of infection.     

Opportunistic infections and retrovirus-induced immunodeficiency: studies of acute and chronic infections with Toxoplasma gondii in mice infected with LP-BM5 murine leukemia viruses.

Mice infected with LP-BM5 murine leukemia viruses develop a syndrome, termed mouse AIDS (MAIDS), characterized by increasingly severe immunodeficiency and progressive lymphoproliferation. Virus-infected mice were examined for the ability to resist acute infection and to control chronic infection with the protozoan Toxoplasma gondii, a major opportunistic pathogen of individuals infected with human immunodeficiency virus. Mice infected with the retroviruses for 2 or 4 weeks responded normally to challenge with the parasite, but mice inoculated with the protozoan 8 or 12 weeks after viral infection died with acute disease due to T. gondii. Increased sensitivity to acute infection was associated with a reduced ability to produce gamma interferon (IFN-gamma) and with established changes in CD4+ T-cell function. Mice latently infected with T. gondii and then inoculated with the retrovirus mixture were found to reactivate the parasite infection, with 30 to 40% of dually infected animals dying between 5 and 16 weeks after viral infection. Reactivation was associated with reduced proliferation and impaired production of IFN-gamma in response to stimulation with soluble T. gondii antigens or to concanavalin A. Continuing resistance to lethal reactivation in the remaining mice was shown to require CD8+ T cells and expression of IFN-gamma. In addition, it was found that chronic infection with T. gondii altered the course of MAIDS by inhibiting the progression of splenomegaly and immunodeficiency and reducing the expression of both the helper and etiologic defective viruses. These results support previous studies which indicate that infection with T. gondii is controlled by synergistic interactions between CD4+ and CD8+ T cells, the functions of which are progressively impaired during the course of MAIDS.     

紫外线减毒弓形虫对小鼠旋毛虫感染的影响

目的探讨紫外线减毒弓形虫免疫对小鼠旋毛虫感染的影响。方法ICR小鼠分为两组，实验组为紫外线减毒弓形虫免疫3周后再感染旋毛虫，对照组为旋毛虫单独感染。结果与旋毛虫单感染相比，小鼠预先经紫外线减毒弓形虫免疫再感染旋毛虫，在感染后5～14d其小肠内的成虫数明显增加（肠道排虫明显延迟），而感染后23-90d其肌肉中的幼虫数明显下降。结论紫外线减毒弓形虫免疫对小鼠旋毛虫感染具有一定的免疫调节和异源性保护作用。     

Effect of administration of a recombinant adenovirus expressing the genes for IFN-gamma and interleukin-12 on acute murine toxoplasmosis.

The effect of recombinant murine interferon-gamma (rMuIFN-gamma) produced from an adenovirus construct on Toxoplasma gondii in tissue culture and on the outcome of a T. gondii infection in mice was determined. Supernatants from AdCMVMuIFN-gamma-infected mouse lung epithelial (MuLE) cells were evaluated for the ability to produce biologically active IFN-gamma by measuring the capacity of the supernatants to activate peritoneal macrophages for killing of T. gondii. The bioactivity of IFN-gamma in supernatants increased with increasing multiplicity of infection (moi). Replication was inhibited 43%, 67%, and 70% by supernatants from MuLE cells infected with AdCMVMuIFN-gamma moi 5, 10, and 50, respectively, (p < 0.01 compared with controls). Bioactivity of IFN-gamma also increased as the length of time after infection increased. T. gondii replication was inhibited 28% and 36%, respectively, by AdCMVMuIFN-gamma-infected MuLE cell supernatants recovered at 24 and 48 h (p < 0.01 compared with control). In vivo administration of AdCMVMuIFN-gamma exhibited 33% mortality by day 9 in mice acutely infected with T. gondii compared with 100% mortality in control mice (p = 0.045). Administration of AdCMVIL-12 reduced mortality to 40% compared with control mice. However, this reduction was not significant (p = 0.08). Overall survival was extended 2 days with AdCMVMuINF-gamma administration and 5 days with AdCMVIL-12. AdCMVMuIFN-gamma in vitro inhibits T. gondii, and in vivo AdCMVMuIFN-gamma and AdCMVIL-12 lead to increased survival in mice.     

Effect of macrophage migration inhibitory factor (MIF) in human placental explants infected with Toxoplasma gondii depends on gestational age

Because macrophage migration inhibitory factor (MIF) is a key cytokine in pregnancy and has a role in inflammatory response and pathogen defense, the objective of the present study was to investigate the effects of MIF in first- and third-trimester human placental explants infected with Toxoplasma gondii. Explants were treated with recombinant MIF, IL-12, interferon-γ, transforming growth factor-β1, or IL-10, followed by infection with T. gondii RH strain tachyzoites. Supernatants of cultured explants were assessed for MIF production. Explants were processed for morphologic analysis, immunohistochemistry, and real-time PCR analysis. Comparison of infected and stimulated explants versus noninfected control explants demonstrated a significant increase in MIF release in first-trimester but not third-trimester explants. Tissue parasitism was higher in third- than in first-trimester explants. Moreover, T. gondii DNA content was lower in first-trimester explants treated with MIF compared with untreated explants. However, in third-trimester explants, MIF stimulus decreased T. gondii DNA content only at the highest concentration of the cytokine. In addition, high expression of MIF receptor was observed in first-trimester placental explants, whereas MIF receptor expression was low in third-trimester explants. In conclusion, MIF was up-regulated and demonstrated to be important for control of T. gondii infection in first-trimester explants, whereas lack of MIF up-regulation in third-trimester placentas may be involved in higher susceptibility to infection at this gestational age.     

Rat T cells express neither CD55 nor CD59 and are dependent on Crry for protection from homologous complement

All human blood cells express decay-accelerating factor (DAF, CD55), CD59, and, with the exception of erythrocytes, membrane cofactor protein (MCP, CD46) to protect themselves from damage by the constant low-level activation of complement in serum. In rats and mice MCP is expressed only in testis, whereas DAF and CD59 are broadly distributed. Rats and mice also express a unique complement regulator, Crry. Previously we have shown that DAF was absent from at least 75% of rat T cells. To further investigate this surprising finding, we assessed the expression levels of DAF, CD59 and Crry on all blood cell types in the rat. We found that Crry was abundantly expressed on all blood cells. CD59 was expressed abundantly on erythrocytes and granulocytes but was absent from all T cellsand platelets and a minority of B cells and NK cells. Double staining and depletion studies showed that T cells in all rat strains tested were DAF-CD59-. Neutralization of Crry using a blocking monoclonal antibody rendered T cells susceptible to lysis by homologous complement, indicating that Crry was solely responsible for protecting DAF-CD59- T cells from complement damage in the rat.     

早孕期弓形虫感染对大鼠胎盘RT.BM1及RT1-E基因转录的影响

通过检测正常妊娠和早孕期弓形虫感染的Wister大鼠胎盘RT.BM1和RT1-E mRNA的表达水平,发现二者在正常妊娠中的规律,进而探索早孕期弓形虫感染对胎盘RT.BM1和RT1-E mRNA表达的影响,以阐明弓形虫感染致不良妊娠结局的分子免疫机制。将48只孕鼠随机分为感染组和正常组,每组24只。感染组于妊娠第5天腹腔注射1×105个弓形虫强毒株(RH株)速殖子/只,正常组不作任何处理。两组于孕第9、13、15、17天分别处死6只孕鼠,无菌剖腹,观察胎鼠情况,计数死胎及活胎数。采用实时荧光定量聚合酶链式反应(real-time PCR)检测胎盘RT.BM1和RT1-E mRNA表达水平。结果:①正常组活胎数204只,死胎3只,死胎率1.4%,感染组活胎数133只,死胎27只,死胎率16.9%,死胎率明显高于正常组(P<0.05);②正常组RT.BM1 mRNA在孕第9、13、15、17天的相对表达水平为(3.21±1.32)、(4.35±1.29)、(3.96±1.49)、(11.1±2.15),前三个时间点表达水平基本一致(P>0.05),第17天则明显增高(P<0.01);RT1-E mRNA在各时间点相对表达水平为(0.79±0.45)、(2.15±1.03)、(2.13±1.74)、(2.39±1.85),第9天明显低于后三个时间点(P<0.05);③感染组各时间点RT.BM1 mRNA相对表达水平为(12.4±2.86)、(11.5±3.67)、(6.89±1.59)、(17.3±3.06),第9、13、15天与正常组相比有显著差异(P<0.01);而RT1-E mRNA分别为(1.83±0.886)、(4.72±1.59)、(4.24±2.26)、(3.83±2.26),第9、13、15天与正常组相比均有显著差异(P<0.01)、(P<0.05)、(P<0.01)。结果显示弓形虫感染量合适,动物模型成功建立;RT.BM1和RT1-E mRNA在大鼠胎盘的表达水平与妊娠进展密切相关;大鼠早孕期弓形虫感染可致胎盘局部RT.BM1和RT1-E表达增高,从而打破正常妊娠所需的胎盘局部免疫平衡状态,可能是早孕期弓形虫感染致不良妊娠结局的分子机制之一。