Different cytokine patterns induced by Helicobacter pylori and Lactobacillus acidophilus extracts in PBMCs of patients with abdominal aortic aneurysm

Abdominal aortic aneurysm (AAA) is a degenerative inflammatory disease with unknown etiology. AAA is characterized by abdominal aortic dilatation more than 3 cm and is often asymptomatic, but the rupture of aneurysm can lead to death. Age, smoking and male sex are major predisposing factors of AAA.This study compares the effect of Helicobacter (H.) pylori and Lactobacillus (L.) acidophilus on the cytokine profile of PBMCs of 5 men with abdominal aortic aneurysm (AAA) and 5 men with normal/insignificant angiography, CT-Scan and ultrasonography results in the single-culture and in the co-culture with HUVECs. IL-2, IL-4, IL-5, IL-6, IL-9, IL-10, IL-13, IL-17A, IL-17 F, IL-21, IL-22, IFN-γ and TNF-α were measured in culture supernatants using a commercial fluorescent-labeled-bead assay.In general, CagA+ H. pylori-extract induced higher production of IFN-γ, IL-13 and IL-21 by PBMCs. Treatment of patients’ PBMCs with CagA⁺ H. pylori-extract induced Th2 cytokines while treatment of controls’ PBMCs with CagA⁺ H. pylori-extract increased Th1 cytokines. In the co-culture, however, patients’ PBMCs produced Th1 cytokines irrespective of extract treatment, while controls’ PBMCs produced Th2 cytokines and decreased IL-10. CagA+ H. pylori- as well as L. acidophilus-extract induced higher levels of IL-9 by controls’ PBMCs in co-culture with HUVECs than patients (P = 0.05 and P = 0.01).The cytokine pattern of PBMCs induced by CagA+ H. pylori- and L. acidophilus-extracts in the co-culture with HUVECs shows differences in AAA patients and in comparison to controls. Decreased secretion of IL-9, IL-21 and IL-22 by PBMCs of patients treated with CagA+ H. pylori extract in co-culture, as opposed to non-AAA controls may indicate the active role ECs play in AAA. Simultaneous production of IL-10 and Th1 cytokines in patients and pronounced Th2 cytokines in controls in response to both bacteria may point to the inherent differences between patients and controls, which need further investigation.     

SOCS2真核表达载体的构建及在3T3 L1前体脂肪细胞中的表达

细胞因子信号抑制蛋白(SOCS)是细胞因子信号通路的重要负调控因子.构建SOCS2真核表达载体并转染3T3-L1前体脂肪细胞,为研究SOCS2对不同细胞因子调控脂代谢奠定基础.取小鼠皮下脂肪提总RNA进行RT-PCR,克隆获得pMDl8-SOCS2质粒载体,双酶切目的基因,并克隆至质粒载体pcDNA3.1,成功构建了重组质粒载体pcDNA3.1-SOCS2;重组质粒瞬时转染3T3-L1前体脂肪细胞系,以未转染和空载体转染作为对照,RT-PCR和Westem blotting检测重组质粒转染组SOCS2核酸和蛋白表达,表达水平均显著高于对照组,为进一步研究SOCS2基因对不同细胞因子调控脂肪代谢提供基础.     

Evaluation of efficacy of a new live Salmonella Typhimurium vaccine candidate in a murine model

Efficacy of a new live Salmonella Typhimurium vaccine candidate attenuated by two virulence gene deletions was evaluated in this study. Live form of the vaccine was orally administered and formalin-inactivated form was intramuscularly (IM) administered. 105 female BALB/c mice were used and divided into 5 groups, A to E, containing 21 mice per group. Serum IgG and secretory IgA titers and levels of serum IFN-γ, IL-4, TNF-α, IL-12 of the immunized groups, especially group C (oral prime-oral booster) significantly increased. In addition, all animals in groups C showed no clinical symptoms and survived the virulent challenges, whereas all or some of mice from the other groups died of the challenge. These indicate that the vaccine candidate can be an effective tool for prevention of Salmonella infections by inducing robustly protective immune responses and leading to the production of inflammatory cytokines. Booster immunization, especially via oral administration, is necessary to optimize its protection.     

补饲开食料对断奶前后牦牛犊牛生长性能和免疫器官发育的影响

旨在探讨补饲开食料对牦牛犊牛生长性能和免疫器官发育的影响。选取出生日龄、体质量相近、健康的大通牦牛犊牛（公）30头,随机等分为两组,对照组犊牛饲喂代乳粉和苜蓿干草,补饲组犊牛在对照组基础上补饲开食料,两组犊牛的干物质饲喂质量保持一致。分别在犊牛断奶前（160 日龄）和断奶后（230 日龄）进行屠宰,每组各屠宰5头,采取肝脏、胸腺和脾脏,观察各器官组织形态,测定分泌激素和细胞因子含量。结果表明：补饲开食料显著提高160日龄和230日龄犊牛平均日增质量。补饲开食料促进160日龄和230日龄犊牛肝细胞的发育;160日龄犊牛肝脏细胞因子含量显著高于对照组,230日龄对照组犊牛肝脏细胞因子含量显著高于试验组。160日龄和230日龄犊牛胸腺皮质厚度显著高于对照组,胸腺细胞因子含量差异不显著。160日龄试验组犊牛脾脏脾小结半径显著大于对照组,脾脏细胞因子含量显著高于对照组;230日龄两组犊牛脾脏细胞因子含量差异不显著。综上,补饲开食料提高160日龄犊牛生长性能,促进免疫器官发育,有效提高犊牛系统免疫力;对230日龄犊牛免疫器官形态发育和功能也具有潜在正向调控作用。     

Seminal plasma did not influence the presence of transforming growth factor-β1, interleukine-10 and interleukin-6 in porcine follicles shortly after insemination

Background

The effects of seminal plasma on the presence of the cytokines transforming growth factor (TGF)-β1, interleukin (IL)-10 and IL-6 in ovarian follicles and follicular fluid were studied shortly after insemination in gilts.Ovaries from gilts were sampled 5–6 h after insemination with either seminal plasma (SP), fresh semen in extender (Beltsville thawing solution, BTS), spermatozoa in extender (Spz), or only BTS (control).

Results

Immunohistochemical (IHC) labeling of TGF-β1, IL-10 and IL-6 was evident in the ovarian oocytes and granulosa cells independent of stage of follicular development (antral follicles). Theca interna cells were labeled to a high degree in mature follicles. No consistent differences between treatment groups could be observed for any of the cytokines.In follicular fluid, high concentrations of TGF-β1 were found while the levels of IL-10 and IL-6 were low. There were no differences between treatment groups.

Conclusions

Our results show a presence of the cytokines TGF-β1, IL-6 and IL-10 in oocytes, granulosa and theca cells, as well as in the fluid of mature follicles suggesting a role of these cytokines in intra-ovarian cell communication. However, treatment (SP, fresh semen in BTS, spermatozoa in BTS or BTS) did not influence the IHC-labeling pattern or the levels of these cytokines in follicular fluid shortly after insemination.     

共轭亚油酸免疫学研究进展

CLA是一类含十八碳二烯酸次生衍生的共轭多烯的多种位置与空间异构体的总称,仅“c9,t11”、“t10,c12”、“t9,t11”、“t10,t12”异构体具有较强的生物学功能。免疫学方面已研究表明:能够增加外周血液和肠系膜淋巴结中的IgGI、gA和IgM的含量,降低IgE的水平;增加促有丝分裂原和促进T、B淋巴细胞增殖调控;提高T细胞CD 特异效应物的功能和淋巴细胞的细胞毒性;以及激活巨噬细胞表达iNOS-mRNA、IL-1I、L-6I、L-4和TNF-α等细胞因子能力等的参与对机体免疫系统的调节作用。这些缓解动物免疫应激功能表达的调控机理,可能与激活巨噬细胞核转录因子———过氧化物酶增殖体激活受体-γ(PPAR-γ)的反应元件,降低干扰素γ诱导的炎性介质mRNA表达有关。     

Immune dysregulation in flea allergy dermatitis--a model for the immunopathogenesis of allergic dermatitis

BACKGROUND: Flea allergy dermatitis (FAD) is a common skin disease in dogs and can be induced experimentally. It often coexists with other allergic conditions. So far no studies have investigated the quantitative production of cytokine mRNA in skin biopsies and peripheral blood mononuclear cells (PBMC) in flea allergic dogs. OBJECTIVE: The aim of our study was to improve the understanding of the immunopathogenesis of allergic dermatitis as a response to fleabites. MATERIAL AND METHODS: Allergic and non-allergic dogs were exposed to fleas. Before and after 4 days of flea exposure mRNA was isolated from biopsies and PBMC. Production of chymase, tryptase, IL-4, IL-5, IL-13, TNF-alpha and IFN-gamma mRNA was measured by real-time RT-PCR. The inflammatory infiltrate in the skin was scored semi-quantitatively. The number of eosinophils, mast cells (MC) and IgE+ cells/mm2 was evaluated to complete the picture. RESULTS: FAD was associated with a higher number of MC before flea exposure and with a significant increase of eosinophils after flea exposure as compared to non-allergic dogs. The number of IgE+ cells was higher in allergic dogs before and after flea exposure. In allergic dogs mRNA for most cytokines and proteases tested was higher before flea exposure than after flea exposure. After exposure to fleas an increased mRNA production was only observed in non-allergic dogs. In vitro stimulation with flea antigen resulted in a decreased expression of most cytokines in allergic dogs before flea exposure. In contrast, in PBMC, only increased levels of IL-4 and IL-5 mRNA were observed in allergic dogs before flea exposure. However, after flea exposure and additional stimulation with flea antigen the production of mRNA for all cytokines tested was significantly increased in allergic dogs. CONCLUSION: We demonstrated that the response in biopsies and PBMC is different and that FAD is associated with a TH2 response.     

Development and validation of cytokine quantitative, real time RT-PCR assays for characterization of Asian elephant immune responses

Infectious disease is an important factor in Asian elephant health and long-term species survival. In studying disease pathogenesis, it is important to consider not only the pathogen, but also the effectiveness of the host immune response. Currently, there is a paucity of information available on elephant immune function. Measurement of cytokine levels within clinical samples can provide valuable information regarding immune function during health and disease that may elucidate disease susceptibility. To develop tools for assessment of elephant immune function, Asian elephant partial mRNA sequences for interleukin (IL)-2, IL-4, IL-10, IL-12, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and β-actin were determined. Sequence information was then utilized to design elephant-specific primers and probes for quantitative, real time, RT-PCR assays for the measurement of cytokine mRNA. Greater than 300 bps of Asian elephant mRNA sequence were determined for each cytokine of interest. Consistent and reproducible, real time, RT-PCR assays with efficiencies of greater than 93% were also developed. Assay sensitivities ranged from less than 1 to 5000 DNA copies with the exception of IL-12, which had a sensitivity of 42,200 copies. Employment of molecular techniques utilizing mRNA-based detection systems, such as real time, RT-PCR, facilitate sensitive and specific cytokine detection and measurement in samples from species for which commercial reagents are not available. Future studies utilizing these techniques to compare elephant immune function during health and in the face of infection will be useful for characterizing the contribution of the elephant immune system to disease.     

Use of serum concentrations of interleukin-18 and monocyte chemoattractant protein-1 as prognostic indicators in primary immune-mediated hemolytic anemia in dogs

Background: The cytokine response in immune‐mediated hemolytic anemia (IMHA) is poorly characterized and correlation with outcome is unknown. Hypothesis/Objectives: To determine if cytokine activity is correlated with outcome in dogs with IMHA. Animals: Twenty dogs with primary IMHA and 6 control dogs. Methods: Prospective study on dogs with IMHA with blood sampling at admission. Serum activity of interleukin‐2 (IL‐2), IL‐4, IL‐6, IL‐7, IL‐8, IL‐10, IL‐15, IL‐18, monocyte chemoattractant protein‐1 (MCP‐1), granulocyte‐macrophage colony stimulating factor (GM‐CSF), interferon‐inducible protein‐10, interferon‐gamma, and keratinocyte chemoattractant (KC) was assessed. Results: Thirty‐day case fatality rate was 25% (5/20 dogs). Increased concentrations (median [range]) of IL‐2 (45.5 ng/L [0;830] versus 0 ng/L [0;46.8]), IL‐10 (8.2 ng/L [0;60.6] versus 0 ng/L [0;88.2]), KC (1.7 μg/L [0.3;4.7] versus 0.5 μg/L [0.2;1.1]), and MCP‐1 (162 ng/L [97.6;438] versus 124 ng/L [90.2;168]) were observed in dogs with IMHA compared with controls. The cytokine profile was indicative of a mixture of pro‐ and anti‐inflammatory cytokines of various cellular origins. Cytokines/chemokines strongly associated with macrophage/monocyte activation and recruitment were significantly increased in nonsurvivors compared with survivors; IL‐15 (179 ng/L [48.0;570] versus 21.3 ng/L [0;193]), IL‐18 (199 ng/L [58.7;915] versus 37.4 ng/L [0;128]), GM‐CSF (134 ng/L [70.0;863] versus 57.6 ng/L [0;164]), and MCP‐1 (219 ng/L [135;438] versus 159 ng/L [97.6;274]), respectively. Logistic regression suggested increased IL‐18 and MCP‐1 concentrations were independently associated with mortality in this population (P<.05, Wald's type 3). Conclusions and Clinical Importance: A mixed cytokine response is present in dogs with IMHA and mediators of macrophage activation and recruitment might serve as prognostic indicators.     

Inflammatory Cytokine Gene Expression in Blood During the Development of Oligofructose-Induced Laminitis in Horses

Systemic inflammation is a risk factor for laminitis in horses and precedes the onset of lameness in experimental models. We therefore hypothesized that whole-blood inflammatory cytokine expression would increase during the development of laminitis in a carbohydrate overload model. Blood samples were obtained from 14 horses undergoing laminitis induction with 10 g/kg oligofructose as part of another study. Samples were collected at 0, 8, 12, 16, 20, and 24 hours, and lameness evaluations were performed every 4 hours. Expression levels of interleukin-1β (IL-1β), IL-6, IL-8, IL-10, and tumor necrosis factor-α were measured in whole blood by using real-time PCR. IL-1β, IL-8, and IL-10 expression increased above baseline from 8 to 24 hours (P < .001), and IL-6 expression increased at 16 and 20 hours (P = .005). Expression of tumor necrosis factor-α did not change over time. All horses developed clinical laminitis between 12 and 24 hours. Increased mean IL-1β, IL-8, and IL-10 expression detected at 8 hours therefore preceded the onset of lameness. We conclude that peripheral leukocyte cytokine expression increases as systemic inflammation develops in an alimentary carbohydrate overload model of laminitis, and this precedes detection of lameness. Results support current recommendations to control the systemic inflammatory response in order to lower the risk of laminitis in horses.