Immunological adjuvant effect of Boswellia serrata (BOS 2000) on specific antibody and cellular response to ovalbumin in mice

In this study, the biopolymeric fraction BOS 2000 from Boswellia serrata was evaluated for its potential ability as adjuvants on the immune responses to ovalbumin (OVA) in mice. Balb/c mice were immunized subcutaneously with OVA 100 μg alone or with OVA 100 μg dissolved in saline containing alum (200 μg) or BOS 2000 (10, 20, 40 and 80 μg) on Days 1 and 15. Two weeks later, OVA specific antibodies in serum; concanavalin A (Con A), OVA stimulated splenocyte proliferation, CD4/CD8/CD80/CD86 analysis in spleen cells and its estimation of cytokines (IL-2 and IFN gamma) from cell culture supernatant were measured. OVA specific IgG, IgG1 and IgG2a antibody levels in serum were significantly enhanced by BOS 2000 (80 μg) compared with OVA control group. Moreover, the adjuvant effect of BOS 2000 (80 μg) on the OVA-specific IgG, IgG1, and IgG2a antibody responses to OVA in mice were more significant than those of alum. BOS 2000 significantly enhanced the Con A and OVA induced splenocyte proliferation in the OVA immunized mice especially at a dose of 80 μg (p<0.001). However, no significant differences were observed among the OVA group and OVA/alum group. At a dose of 80 μg (p<0.001), there was a significant increase in the CD4/CD8 and CD80/CD86 analysis in spleen cells and cytokine (IL-2 and IFN-gamma) profile in the spleen cell culture supernatant was observed. In conclusion, BOS 2000 seems to be a promising balanced Th1 and Th2 directing immunological adjuvants which can enhance the immunogenicity of vaccine.     

The feeding of beta-carotene down-regulates serum IgE levels and inhibits the type I allergic response in mice

Feed containing beta-carotene was administered orally to BALB/c mice immunized intraperitoneally with ovalbumin (OVA) for approximately 1 month. The titers of OVA-specific IgE, OVA-specific IgG1 and OVA-specific IgG2a in the mouse sera were determined. The OVA-specific IgE titer and OVA-specific IgG1 titer by mice fed beta-carotene were significantly inhibited. On the other hand, the OVA-specific IgG2a titer in mice fed beta-carotene was significantly greater than those of control mice. The OVA-specific IgE suppression of beta-carotene feeding was dose-dependent. We also examined the effect of fed beta-carotene on active systemic anaphylaxis. Feeding beta-carotene to mice immunized with OVA inhibited the immediate reduction of the body temperature induced by antigen stimulation. Furthermore, the increase in serum histamine in the mice fed beta-carotene under active systemic anaphylaxis was lower than in controls. We then examined the pattern of cytokine production by spleen cells from mice followed by restimulation with OVA in vitro. The spleen cells from the mice fed beta-carotene produced more IFN-gamma, IL-12 and IL-2 than those from the control group. In contrast, the spleen cells from the mice fed beta-carotene produced less IL-4, IL-5, IL-6, IL-10 than those from the control group. Furthermore, analysis of IFN-gamma mRNA levels of the splenocytes using the real-time quantitative RT-PCR technique revealed higher levels in the splenocytes from the mice fed beta-carotene. These findings suggest that feeding beta-carotene improves the helper T cell (T(H))1-T(H)2 balance, inhibiting specific IgE and IgG1 production and antigen-induced anaphylactic response.     

枸杞子多糖对小鼠白细胞介素-2活性的增强作用(英文)

本文研究了中药枸杞子的提取物枸杞子多糖(LBP)对成年(2月龄)和老年(16月龄)小鼠脾白细胞介素-2(IL-2)活性的影响。在诱导脾细胞IL-2的培养液(脾细胞5×10~6/ml和ConA 4μg/ml)中加入LBP,所制备的含IL-2上清液使成年小鼠胸腺细胞在体外增殖活性([~3H]TdR参入法)升高。老年小鼠IL-2促进淋巴细胞增殖水平显著低于成年。LBP可使老年小鼠IL-2的这种作用显著提高,达成年水平。这些结果表明,LBP对IL-2的活性有增强作用并使老年小鼠IL-2活性得到恢复。     

Prenatal administration of indomethacin modulates Th2 cytokines in juvenile rats

Indomethacin (IND) suppresses the T-dependent antibody response (TDAR) in juvenile males when it is administered to pregnant rats during late gestation. In this study, the effect of IND on cytokine production in juvenile rats was examined to investigate the mechanism behind the suppression of antibody production. IND was orally administered to pregnant SD rats on days 18–21 of gestation. After parturition, the spleen cells isolated from 3-week-old pups were incubated with concanavalin A (Con A) or lipopolysaccharide (LPS). The level of cytokines in the culture supernatant was measured. IL-10 decreased significantly in the males, and IL-6 and TNF-α tended to decrease in both sexes.     

IL-35 interferes with splenic T cells in a clinical and experimental model of acute respiratory distress syndrome

Acute respiratory distress syndrome (ARDS) is a life-threatening critical care syndrome with uncontrolled inflammation that is a central issue. Its main characteristic is inflammatory mediators and cytokines as well as agglutinating chemokines that injure target cells. Interleukin (IL)-35 is a newly identified IL-12 cytokine family member with structural similarities to other IL-12, IL-23, and IL-27 cytokines but unique immunological functions. How IL-35 functions in ARDS is unclear. The purpose of our study was to determine what role IL-35 played in the development of ARDS. Here we found serum IL-35 concentrations were significantly elevated in patients with ARDS relative to healthy people. Moreover, we established a mouse model of lipopolysaccharide- and cecal ligation and puncture-induced ARDS treated with neutralizing antibodies (anti-IL-35 Ebi3 or anti-IL-35 P35); the results showed that lung injury occurred more often than in untreated models and the inflammatory cytokines CXCL-1, tumor necrosis factor alpha, IL-6, and IL-17A increased significantly after neutralizing antibody treatment in bronchoalveolar lavage fluid and serum. Therefore IL-35 can protect against the development of ARDS. Even more interesting in our study was that we discovered IL-35 expression differed between lung and spleen across different ARDS models, which further demonstrated that the spleen likely has an important role in extrapulmonary ARDS model only, improving the ratio of CD4⁺/CD4⁺CD25⁺Foxp3⁺(Tregs). Meanwhile in our clinical work, we also found that the concentration of IL-35 and the ratio of CD4⁺/Treg in the serum are higher and the mortality is lower than those with the spleen deficiency in patients with extrapulmonary ARDS. Therefore, IL-35 is protective in ARDS by promoting the ratio of splenic CD4⁺/Tregs in extrapulmonary ARDS, and as such, may be a therapeutic target.     

小鼠黑素瘤细胞培养上清对同型小鼠淋巴细胞转铁蛋白受体CD71的抑制作用分析

目的通过检测小鼠黑色素瘤细胞(B16F10)培养上清对同型小鼠淋巴细胞活化分子CD71表达的抑制,探讨黑素瘤细胞抑制淋巴细胞活化、介导肿瘤细胞免疫逃逸的机制。方法用B16F10培养上清培养同型小鼠脾淋巴细胞(上清组),经植物血凝素活化,用流式细胞仪检测小鼠脾淋巴细胞转铁蛋白受体CD71表达水平,用RPMI1640完全培养液代替B16F10培养上清作为对照组。结果上清组培养上清培养的淋巴细胞CD71表达率为(55.41±3.70)%低于对照组的(60.92±2.02)%,差异有统计学意义(P<0.05)。结论 B16F10培养上清对淋巴细胞表达CD71具有抑制作用,可抑制淋巴细胞活化,成为免疫逃逸机制之一。     

Expression of IL-23, VEGF and TLR2/TLR4 on mononuclear cells after exposure to Pseudomonas aeruginosa

Pseudomonas aeruginosa is a Gram-negative, aerobic bacillus causing infections of the respiratory and other organ systems in susceptible hosts. Although it does not cause pulmonary infections in immunocompetent individuals, P. aeruginosa causes chronic lung infection in individuals with cystic fibrosis and nosocomial pneumonia resulting in significant morbidity and mortality. Exogenous administration of an important P. aeruginosa virulence factor, lipase, present in P. aeruginosa culture supernatant, induces potent mononuclear cell activation leading to the production of numerous proinflammatory cytokines. In particular, P. aeruginosa culture supernatant stimulated increased proliferation of THP-1 cells and monocytes (MN). The addition of culture supernatant to THP-1 cells and MN also induced Interleukin (IL)-23 and vascular endothelial growth factor (VEGF) release in a time-dependent manner. To investigate whether any compounds present in the supernatant lipase contributed to releasing IL-23 and VEGF, the culture supernatant from P. aeruginosa containing lipase was treated with hexadecylsulfonylfluoride (AMSF). The AMSF-treated culture supernatant (CS) did not show any induction on the IL-23 and VEGF release compared to the cells treated with CS without AMSF. We also showed that Toll-like receptors (TLR)2/TLR4 are expressed in THP-1 cells and MN treated with P. aeruginosa CS in a time-dependent fashion. Flow cytometry analysis revealed a higher TLR4 and a lower TLR2 expression at 48 and 72 h of treatment. The treatment of cells with TLR4 neutralizing antibody, and to a lesser extent with TLR2 neutralizing antibody, resulted in a decrease in P. aeruginosa CS-induced IL-23 and VEGF production.     

Immunomodulatory properties of Mycoplasma pulmonis. III. Lymphocyte stimulation and cytokine production by Mycoplasma pulmonis products

Experiments are presented that were performed in order to understand the mechanisms causing these effects on the immune system. Mitogenic effects of Mycoplasma membranes on mouse spleen cells were shown using M. capricolum. The observed mitogenic activity is proportional to the amount of membranes used, as measured by protein content. Separation of T and B cells was performed by two techniques, the anti-Thyl.2 plus complement method and the Dynabead technique. Using the former technique, it was shown that removal of T cells markedly reduced effects of stimulation by mycoplasma membranes, but did not abolish it. The separated cells were still stimulated by PHA, indicating that the preparation still contained T cells. Furthermore, removal of T cells preferentially reduced the PHA response over that of mycoplasma membranes, indicating that mycoplasma membranes stimulate both B and T lymphocytes. The Dynabead system was found to be the more efficient separation technique, and by using it we were able to make the following observations. Inactivated Mp, membranes and culture supernatant stimulated B cells, whereas T cells were only slightly stimulated by inactivated Mp and membranes. There was an increase in proliferation when T cells were incubated with adherent cells from peripheral blood. Finally, we showed that spleen cells from infected animals produce more IL-4 and less IFN-gamma than cells from non-infected animals when stimulated with membranes, inactivated Mp, culture supernatant or phytohemagglutinin. Altogether, these results show that lymphocytes from Mycoplasma-infected animals are directly affected and this effect is probably due to superantigen-like molecules from M. pulmonis.     

Enhancement of the immune responses of mice to <Emphasis Type="Italic">Bacillus anthracis</Emphasis> protective antigen by CIA07 combined with alum

Anthrax is an acute zoonotic disease caused by infection with Bacillus anthracis. B. anthracis spores are highly resistant to environmental degradation and are used as a biological weapon. In this study, we investigated the adjuvant activity of CIA07 to anthrax protective antigen (PA). A/J mice were immunized intraperitoneally once, or twice with a 4-week interval, with recombinant PA alone or combined with alum, CpG1826, or CIA07 as adjuvant, and serum anti-PA IgG antibody responses were measured 4 weeks after each immunization. All three adjuvants significantly enhanced anti-PA IgG antibody titer 4 weeks after the priming and boosting immunizations, and alum gave the highest titer. In order to evaluate the adjuvant activity of CIA07 in the presence of alum, Balb/c mice were immunized 3 times at 1-week intervals with PA in combination with alum, CIA07 or alum plus CIA07, and the immune responses were assessed 2 weeks after the third immunization. The serum anti-PA IgG antibody titer of the CIA07-treated group was 14-fold higher than the group given PA alone, and the coadministration of CIA07 with alum further increased the titer 3.5-fold (P < 0.05). The toxin neutralizing activity of the sera from the mice given the combination of CIA07 and alum was 109-times higher than the animals given PA alone. The mice given CIA07 plus alum also showed a marked increase in the number of IFN-γ-, IL-2-, and IL-4-producing CD4⁺ T cells among their splenocytes. These data suggest the potential of CIA07 in combination with alum as an adjuvant for the development of a potent anthrax vaccine.     

scBsAb1/17双特异性抗体对小鼠类风湿性关节炎模型的治疗效果

目的比较不同剂量scBsAb1/17对Ⅱ型胶原(CII)诱导类风湿性关节炎(CIA)模型小鼠的治疗效果,及scBsAb1/17双特异性抗体与单价抗体(anti-IL-1βscFv和anti-IL-17scFv)在CIA模型小鼠中的治疗效果。方法利用CII建立小鼠类风湿性关节炎模型,小鼠成模后开始治疗,每2 d给药1次,治疗29 d。治疗结束后对小鼠关节炎指数进行临床评分,检测各组小鼠血清中CII抗体、CII特异性刺激的脾细胞增殖指数和脾脏中IL-2、IL-1β、IFN-γ、IL-6和TNF-α等细胞因子的表达水平。结果与CIA模型对照组相比,所有治疗组都能明显减轻CIA小鼠的临床症状,并明显降低血清中CII抗体水平、脾细胞增殖指数及脾脏中TNF-α、IL-6、IL-2、IL-1β和IFN-γ的表达量。相同剂量下scBsAb1/17治疗组的脾细胞增殖指数明显低于anti-IL-1βscFv治疗组(P<0.05);并且scBsAb1/17治疗组小鼠脾脏中IL-6、IL-2、IL-1β和IFN-γ的表达量明显低于anti-IL-1βscFv和anti-IL-17AscFv单独治疗组(P<0.01或P<0.05)。结论 scBsAb1/17及单价抗体对CIA模型小鼠都有治疗效果;不同剂量scBsAb1/17对CIA模型鼠的治疗效果呈剂量依赖性;相同剂量条件下,scBsAb1/17的治疗效果要优于单价抗体。     

鲨肝活性肽的免疫调节和抗细胞凋亡作用

目的　为了选择鲨肝活性肽 (SHAP)临床应用的适应症并了解其作用机制。方法　分离培养人外周血单核细胞 (PBMC) ,检测SHAP对其分泌IFN α和IFN γ的影响 ;用环磷酰胺 (Cy)建立免疫低下模型 ,检测SHAP对免疫低下小鼠血清溶血素的生成及血清IL 2水平的影响 ;用流式细胞仪分析SHAP对对乙酰氨基酚 (AAP)诱导小鼠肝损伤模型中细胞凋亡的影响 :用Fas单克隆抗体诱导小鼠暴发性肝炎和抑制肝癌细胞株SMMC772 1的增殖 ,用SHAP处理后 ,分析小鼠血清谷丙转氨酶 (GPT)的水平及SMMC772 1增殖的变化情况。结果　SHAP能够有效诱导PBMC分泌IFN α和IFN γ ,促进免疫低下小鼠血清溶血素的生成和提高血清IL 2的水平 ,显著降低AAP诱导的肝细胞凋亡。 5 0mg·L- 1的SHAP可消除 5mg·L- 1Fas单克隆抗体对SMMC772 1增殖的抑制作用。 3mg·kg- 1的SHAP能显著降低Fas单克隆抗体诱导的暴发性肝炎小鼠的血清GPT水平。结论　SHAP具有免疫调节作用和抗细胞凋亡的作用 ,可能是SHAP防治肝炎的作用机制之一。     

The role of macrophages in stimulation of immune induction and myelopoiesis. I: Comparison of activity of macrophage-derived factors in granulopoiesis and immunostimulation.

Hemopoietic growth and immunological inductive factors from human peripheral leukocyte conditioned medium and mouse macrophage culture supernatant fluids were compared. Factors derived from human peripheral leukocyte conditioned medium were found to substitute for those of mouse macrophage derivation in immunological assays, namely, induction of a cytotoxic T-lymphocyte response in macrophage-depleted mouse spleen cultures. Association of macrophage-derived factors with B2-microglobulin (B2m) antigen determinants was observed by inhibition with great antihuman B2m, affinity chromatography, and direct replacement of mouse macrophage-derived factors with human urinary B2m.     

Stimulation by a hydroxythiazolobenzimidazole of enhanced formation of antibodies to sheep erythrocytes in vitro and in vivo.

A hydroxythiazolobenzimidazole, a low molecular weight compound, was found to have an immunoenhancing effect on both the in vivo and in vitro antibody response of mouse spleen cells to an optimum immunizing dose of sheep red blood cells. At noncytotoxic concentrations the optimum range of 25 to 50 micrograms per 5 x 19(6) spleen cells was most effective in vitro. Concentrations greater than 100 micrograms per culture were toxic in vitro and reduced cell viability as well as antibody responsiveness. The compound enhanced to an even greater degree the antibody response of spleen cell cultures immunized with suboptimum doses of SRBC. The background PFC response, in the absence of SRBC, also was stimulated by the benzimidazole. These immunoenhancing responses were not related to mitogenic effects, since increased thymidine uptake did not occur when normal mouse spleen cells were incubated with graded doses of the compound. Therefore, the immunostimulatory properties of the compound, both in vivo and in vitro, were not due to mitogenic stimulation of lymphoid cells.     

A comparison of oral and parenteral routes for therapeutic vaccination with HSV-2 ISCOMs in mice; cytokine profiles, antibody responses and protection

It is likely that recurrent infections with HSV-2 (or HSV-1) are influenced by local levels of immunity at mucosal surfaces, when virus reactivated from the latent state is infecting mucosal epithelial cells. Increasing the levels of cellular and humoral immunity through immunisation and maintaining such increased levels, may reduce establishment and spread of reactivated virus at the local site, thereby ameliorating recurrent disease symptoms. The use of HSV-2 antigens incorporated into immunostimulating complexes (ISCOMs) for immunisation of mice previously infected with HSV-2 was investigated in the present study. Prophylactic administration of HSV-2 ISCOM vaccine to mice elicits local antibody detectable in nasal washings, serum antibody and the presence of cytokines IL-2, IFN-gamma and IL-4 in supernatants from spleen cell cultures stimulated in vitro with HSV-2 antigens. Use of the same vaccine in mice infected previously with HSV-2, results in increased levels of total and subclass serum ELISA antibody and also increased levels of serum neutralising antibody. Treatment of HSV-2 infected mice with the HSV-2 ISCOM vaccine also induces higher levels of the cytokines IL-2, IFN-gamma and IL-4, in in vitro stimulated spleen cell cultures. Challenge with a lethal dose of HSV-1 showed that mice previously infected with HSV-2 and subsequently given two doses of HSV-2 ISCOMs vaccine were protected.     

Specific polyclonal and monoclonal antibody prevents paraquat accumulation into rat lung slices

Sheep polyclonal and mouse monoclonal antibodies have been produced that bind to the bipyridyl herbicide, paraquat. The binding capacities and affinities of the various antibody solutions (serum, ascites, purified tissue culture supernatant) to paraquat were determined using a radioimmunoassay. All antibody solutions bound paraquat with high affinity (Ka = 10(9)-10(10) l/mol). The sheep polyclonal antisera, the mouse ascites fluid, and the purified culture supernatant had mean binding capacities of 8, 1 and 22 micrograms paraquat/ml respectively. All the antibody preparations were able to prevent the in vitro accumulation of paraquat into rat lung tissue. The amount of antibody to achieve this was dependent upon the binding capacity of the antibody solution, i.e. when the binding capacity of the antibody was equal to the amount of paraquat present in the incubation medium a total blockade of uptake was achieved. When antibody was added to lung tissue that had been accumulating paraquat for 1 hr, the inhibition of uptake was immediate and was complete for at least 2 hr. Both the radioimmunoassay and lung slice experiments indicate that an equivalent of 1 mg of IgG is required to bind 2.5 micrograms of paraquat ion. Preincubation of lung tissue with antibody did not affect the subsequent accumulation of paraquat, nor did it result in a detectable degree of intracellular neutralisation of paraquat as measured by paraquat's ability to stimulate the pentose phosphate pathway. The rate of efflux of paraquat from lung slices prepared from rats dosed intravenously with paraquat was not increased by the presence of antibody in the incubation medium. In conclusion, neutralising antibodies to paraquat have been produced. They bind to paraquat in solution with high affinity and render the paraquat unavailable for its in vitro accumulation into lung cells.     

淫羊藿多糖促进免疫功能的实验研究

中药淫羊藿多糖成分50mg/kg以上剂量连续皮下给药,7天,可显著促进小鼠免疫功能。表现如下:(1)增加脾脏的PFC数,溶血空斑也显著增大。(2)提高血清溶血素水平。(3)增加外周血白细胞数、淋巴细胞及脾有核细胞数,并促进PHA刺激的淋巴细胞转化反应。(4)增强腹腔Mφ吞噬功能。(5)使脾脏重量增加,胸腺减轻。     

Platonin, a cyanine photosensitizing dye, inhibits pyrogen release and results in antipyresis

Intravenous injection of the supernatant fluids from human peripheral blood mononuclear cells (PBMC) incubated with lipopolysaccharide (LPS) caused fever in rabbits. The fever was in parallel with the levels of either interleukin-1 beta (IL-1 beta), IL-6, or tumor necrosis factor-alpha (TNF-alpha) in supernatant fluids. When incubating the platonin with the LPS-human PBMC, both the levels of IL-1 beta, IL-6, or TNF-alpha in supernatant fluids and the pyrogenicity of supernatant fluids were significantly suppressed. The febrile response to supernatant fluids from the LPS-stimulated PBMC was attenuated almost completely by adding anti-IL-1 beta, but not anti-IL-6 or anti-TNF-alpha, monoclonal antibody to supernatant fluids. In addition, both the fever and the increased levels of either IL-1 beta, IL-6, or TNF-alpha in rabbit serum following an intravenous administration of LPS were significantly attenuated by pretreatment with an intravenous dose of platonin. Furthermore, the fever induced by intravenous injection of IL-1 beta was reduced by pretreatment of rabbits with intravenous injection of platonin. The data indicate that platonin inhibits production of pyrogenic cytokines (in particular, IL-1 beta) from PBMC and results in antipyresis.