人精浆对不同丝裂原诱导淋转反应的影响

本文通过人精浆对PHA、ConA和PWM三种丝裂原诱导淋巴细胞转化反应的影响,间接了解HSP对T辅助、T抑制和B淋巴细胞功能的影响,结果发现不同浓度的精浆对三种丝裂原诱导的淋转反应均有明显抑制作用(P<0.05),且三种浓度HSP的抑制作用在无显著性差异(P>0.05);中浓度的精浆对PHA、ConA和PWM诱导转反应的抑制率分别为69.7、51.7和48.3％,说明精浆对机体的T辅助、T抑制性和B淋巴细胞功能均有明显的抑制作用。此外,还对精浆的免疫抑制作用机理和意义进行了讨论。     

Hepatoprotective Effect of Mixture of Dipropyl Polysulfides in Concanavalin A-Induced Hepatitis

The main biologically active components of plants belonging to the genus Allium, responsible for their biological activities, including anti-inflammatory, antioxidant and immunomodulatory, are organosulfur compounds. The aim of this study was to synthetize the mixture of dipropyl polysulfides (DPPS) and to test their biological activity in acute hepatitis. C57BL/6 mice were administered orally with DPPS 6 h before intravenous injection of Concanavalin A (ConA). Liver inflammation, necrosis and hepatocytes apoptosis were determined by histological analyses. Cytokines in liver tissue were determined by ELISA, expression of adhesive molecules and enzymes by RT PCR, while liver mononuclear cells were analyzed by flow cytometry. DPPS pretreatment significantly attenuated liver inflammation and injury, as evidenced by biochemical and histopathological observations. In DPPS-pretreated mice, messenger RNA levels of adhesion molecules and NADPH oxidase complex were significantly reduced, while the expression of SOD enzymes was enhanced. DPPS pretreatment decreased protein level of inflammatory cytokines and increased percentage of T regulatory cells in the livers of ConA mice. DPPS showed hepatoprotective effects in ConA-induced hepatitis, characterized by attenuation of inflammation and affection of Th17/Treg balance in favor of T regulatory cells and implicating potential therapeutic usage of DPPS mixture in inflammatory liver diseases.     

Hsp70 vaccination-induced primary immune responses in efferent lymph of the draining lymph node

Bovine paratuberculosis is a highly prevalent chronic infection of the small intestine in cattle, caused by Mycobacterium avium subspecies paratuberculosis (MAP). In earlier studies we showed the protective effect of Hsp70/DDA subunit vaccination against paratuberculosis. In the current study we set out to measure primary immune responses generated at the site of Hsp70 vaccination. Lymph vessel cannulation was performed to obtain efferent lymph from the prescapular lymph node draining the neck area where the vaccine was applied. Hsp70 vaccination induced a significant increase of CD21⁺ B cells in efferent lymph, accounting for up to 40% of efferent cells post-vaccination. Proliferation (Ki67⁺) within the CD21⁺ B cell and CD4⁺ T cell populations peaked between day 3 and day 5 post-vaccination. From day 7, Hsp70-specific antibody secreting cells (ASCs) could be detected in efferent lymph. Hsp70-specific antibodies, mainly of the IgG1 isotype, were also detected from this time point onwards. However, post-vaccination IFN-γ production in efferent lymph was non-sustained. In conclusion, Hsp70-vaccination induces only limited Th1 type immune responsiveness as reflected in efferent lymph draining the vaccination site. This is in line with our previous observations in peripheral blood. The main primary immunological outcome of the Hsp70/DDA subunit vaccination is B cell activation and abundant Hsp70-specific IgG1 production. This warrants the question whether Hsp70-specific antibodies contribute to the observed protective effect of Hsp70 vaccination in calves.     

Inhibition of IL-2 inducible T-cell kinase alleviates T-cell activation and murine myocardial inflammation associated with CVB3 infection

Background

Coxsackievirus B3 (CVB3) infection causes myocarditis, pancreatitis, and aseptic meningitis. Targeting antigen-specific T cell reactions might be a promising way to alleviate the inflammatory response induced by CVB3 infection. IL-2-inducible T-cell kinase (ITK), a member of Tec kinase family expressed mainly in T cells, plays an important role in the activation of T cells. The role of ITK in viral myocarditis induced by CVB3 has not been documented.

Methodology

In this study, we inhibited the ITK expression in Jurkat cells, primary human peripheral blood mononuclear cells (PBMC), and mouse splenocytes by ITK-specific siRNA. The inhibition efficiently suppressed cell proliferation (P < 0.05) and T-cell related cytokine secretion (P < 0.05). In order to inhibit ITK in vivo, the pGCSIL plasmid containing short hairpin RNAs targeting ITK was constructed and transduced into mice infected with CVB3. ITK-inhibited mice showed reduced cell proliferation (3, 5, and 7 days post-challenge, P < 0.05) as well as CD4+ and CD8+ T cells (5 days post-challenge, P < 0.05). The altered production of inflammatory cytokines alleviated pathologic heart damage and improved mice survival rate (P < 0.05).

Conclusion

ITK played an important role in the T cell development and represented a new target for the modulation of T-cell-mediated inflammatory response by CVB3 infection.     

PHA、ConA及CD3ε单克隆抗体对小鼠淋巴细胞迁移及分泌趋化因子的影响

目的探讨不同浓度的PHA、ConA及CD3ε单克隆抗体对Balb/c小鼠淋巴细胞迁移及分泌趋化因子的影响,初步了解这3种刺激剂与小鼠淋巴细胞迁移功能之间的关系。方法通过研磨法分离Balb/c小鼠脾脏淋巴细胞,用不同浓度的PHA、ConA及CD3ε单克隆抗体分别刺激培养淋巴细胞,收集细胞上清,用Traswell法测定细胞上清对淋巴细胞迁移率的影响;ELISA法检测细胞上清中趋化因子XCL1、CCL4、CCL5、CCL3、IL-16、IL-8及CCL20的水平。结果 PHA组各个浓度刺激下淋巴细胞的迁移率和对照组无明显差异;ConA组的刺激浓度为20μg/ml时,其对应的淋巴细胞迁移率明显高于对照组;CD3ε单抗组,其浓度分别为2.5、10μg/ml时,对应的淋巴细胞迁移率明显高于对照组。在各个刺激组中,ELISA测得的多种趋化因子水平在总体上存在明显差异。结论 ConA及CD3ε单克隆抗体分别刺激小鼠淋巴细胞后,其分泌趋化因子的功能发生了明显的改变,这些趋化因子影响着小鼠淋巴细胞的迁移功能。趋化因子XCL1、CCL4、CCL5、CCL3的水平明显高于对照,说明这几个因子在淋巴细胞迁移中起到重要的作用。根据实验结果初步推荐CD3ε单抗可以作为小鼠淋巴细胞迁移试验中的首选阳性对照,次选为ConA。     

Immunosuppressive Effect of Cyclosporin A in Two Lymphocyte Transfer Models in Rats: Comparison of in Vivo and in Vitro Treatment

The immunosuppressive effect of CS-A was first studied in the assay for local GvHR. 3 x 10⁶ viable spleen cells from LEW rats were injected into one foot pad of LEW x BN hybrids and both PLN were weighed 7 days later. Treatment of the recipients with 3 doses of 50 mg/kg/day of CS-A suppressed this local GvHR. The effect was more pronounced when treatment started at the time of cell transfer rather than a few days before peak response. In-vitro incubation of the cellular inoculum with CS-A also prevented local GvHR. Histology of the PLN confirmed the quantitative results expressed by the PLN index. CS-A was further investigated in the EAE model in LEW rats. It protected rats sensitised with spinal cord emulsified in complete Freund's adjuvant for as long as they were treated with CS-A. Treatment delayed until after the appearance of EAE also markedly improved their condition. Oral treatment of recipients with 50 mg/kg/day CS-A prevented the development of adoptive EAE following the transfer of lymphoid cells conditioned in vitro. The presence of 0.1-1.0 μg CS-A in the medium of the sensitised lymphoid cells also inhibited the adoptive transfer of EAE. Finally, if the cells for the adoptive transfer were derived from CS-A-treated sensitised donors, they failed to induce EAE. Histological examination supported the symptomatic findings     

ConA对红细胞膜力学性质的影响

本文研究了伴刀豆蛋白ＣｏｎＡ（Ｃｏｎｃａｎａｖａｌｉｎ　Ａ）以受体配体的作用方 式对红细胞膜力学性质的影响，结果显示出：（１）经ＣｏｎＡ处理后的血影红细胞膜 （ｒｅｓｅａｌ ｇｏａｓｔ）蛋白构象发生改变，蛋白运动性下降；（２）红细胞变形恢复半时间缩 短．表明ＣｏｎＡ作用后的红细胞膜切变弹性模量可能增大；（３）红细胞机械脆性随 ＣｏｎＡ浓度呈上升趋势，说明膜稳定性受到影响；（４）红细咆膜脂流动性下降．     

ConA诱导肝细胞损伤机理及CsA对其损伤的影响

目的　探讨NO、脂质过氧化反应及蛋白巯基状态改变在ConA诱导肝组织细胞损伤过程中的作用以及CsA对其损伤的影响。方法 :尾静脉注射ConA( 2 0mg·kg-1)于雄性BALB/c小鼠作为试验组 ;提前半小时予以CsA( 2 5mg·kg-1)后再按试验组处理作为CsA组。观察血清中ALT、AST、NO-2 及肝组织MDA、TSH、PSH含量在 3h、6h、9h和 12h的动态变化。结果　试验组血清中ALT、AST进行性升高 ,在 12h时与对照组、CsA组比较均P <0 .0 1。试验组动物体内有大量的NO合成 ,血清中NO-2 随时间呈明显上升趋势 ,在 12h时与对照组、CsA组比较均P <0 .0 1。试验组动物肝组织MDA含量逐步升高 ,在 12h与对照组、CsA组比较P <0 .0 1。试验组动物肝组织TSH含量不断下降在 12h与对照组、CsA组比较P <0 .0 5 ;PSH含量亦不断下降在 12h与对照组、CsA组比较P <0 .0 1。对照组和CsA组血清中ALT、AST及NO-2 前后变化不大 ,肝组织MDA、TSH、PSH含量变化亦不明显。结论　在ConA所致肝损伤中 ,动物体内NO生物合成机制被激活 ,是肝细胞损伤的重要介质之一。同时脂质过氧化的发生以及蛋白巯基持续消耗致肝组织细胞的抗氧化能力和解毒能力下降也均是ConA性肝损伤机制之一。免疫抑制剂CsA预处理能阻断ConA性肝损伤的发生 ,免疫介导机制亦可能是其损伤     

Evaluation of immunization with gut membrane glycoproteins of Ostertagia ostertagi against homologous challenge in calves and against Haemonchus contortus in sheep

Peanut and ConA lectins were used as ligands to isolate glycoproteins from detergent extracts of adult Ostertagia ostertagi membranes. As judged by their profiles following SDS-PAGE, these fractions closely resembled the equivalents from Haemonchus contortus which are derived from the nematode intestinal cell microvillar membranes and which are highly protective when used as antigens. Groups of calves were immunized with the peanut and ConA binding fractions of Ostertagia, either as separate or pooled antigens mixed with QuilA as adjuvant. All calves, including controls immunized with adjuvant only, were challenged with a single dose of infective Ostertagia larvae and faecal egg counts were monitored for 5 weeks. In two experiments where the antigen fractions were pooled, moderate (30-50%), but statistically significant reductions in egg output were observed, but the number of worms was not diminished. No significant protection was observed in a third trial where groups of calves were immunized with peanut or ConA binding proteins given separately. Two further trials were conducted in sheep immunized with the same Ostertagia fractions but challenged with Haemonchus. Irrespective of whether they were administered separately or together, the Ostertagia antigens cross protected efficiently against Haemonchus reducing egg counts by between 81% and 97% and worm numbers by between 57% and 84%.     

小鼠胸膜间皮细胞培养上清对脾细胞增殖反应的影响

目的研究小鼠胸膜间皮细胞培养上清(MPMS)对脾细胞生长与功能的影响。方法取BALB/c小鼠胸膜组织块直接培养和用胰蛋白酶消化后培养胸膜间皮细胞。于培养的不同时间,取MPMS对ConA刺激脾细胞的增殖,采用3H-TdR掺入法测定。结果直接培养法收集的MPMS-A分别以1∶1及1∶10的比例稀释时,对脾细胞自发掺入3H-TdR的增殖均具有明显地抑制作用,与对照组(培基 脾细胞,MS)相比较差异显著(P<0.05-0.001);用胰蛋白酶消化收集的培养3 d的MPMS-B,在1∶1稀释度时,对小鼠脾细胞的增殖具有促进作用(P<0.05-0.01)。而培养3 d收集的MPMS-B,作1:10的比例稀释时,其对ConA刺激的脾细胞增殖反应具有促进作用,与对照组(MS ConA)相比较差异显著(P<0.01)。结论胸膜细胞培养上清对脾细胞增殖呈双向作用影响,而抑制作用大于增强作用。可为研究中医藏像理论与免疫的关系提供基础研究依据。