KIR2DL3 and KIR2DL1 show similar impact on licensing of human NK cells

Killer cell immunoglobulin‐like receptor/HLA class I (KIR/HLA‐I) combinations are associated with disease risk, implicating functional roles for NK cells (NKCs) or KIR⁺ T cells. KIR/HLA‐I interactions can act through inhibition of NKC activation by target cells and NKC licensing for greater intrinsic responsiveness. We compared licensing conferred by the weaker, HLA‐C group 1/KIR2DL3, and the stronger, HLA‐C group 2/KIR2DL1, inhibitory combinations. The “rheostat model” predicts weaker licensing by HLA‐C1/KIR2DL3 interactions than HLA‐C2/KIR2DL1. We analyzed degranulation in NKC subsets expressing single and multiple receptors for HLA‐I. NKG2A had the strongest licensing impact, while KIR2DL3, KIR2DL1, and KIR3DL1 were weaker, and not significantly different to each other. Presence of one or two matched HLA‐C allotypes did not alter licensing of KIR2DL3⁺ and KIR2DL1⁺ NKC. Coexpression of activating KIR2DS1 disarmed KIR2DL3⁺ and KIR2DL1⁺ NKC to a similar extent. KIR3DL1 and NKG2A combined for more enhanced licensing of double‐positive NKC than the combination of KIR2DL3 and KIR2DL1. Thus, KIR2DL3 and KIR2DL1 have similar capacity to license NKC, suggesting that inhibitory signal strength and amount of available HLA‐C ligands do not correlate with NKC licensing. Altogether, our results show that the basis for disease associations of HLA‐C and KIR2DL likely encompasses factors other than licensing.     

Pharmacokinetics of oral mycophenolate mofetil in combination with CsA in dogs after nonmyeloablative allogeneic hematopoietic stem cell transplantation

Mycophenolate mofetil (MMF) has been used successfully in solid organ transplantation (SOT) and more recently in nonmyeloablative hematopoietic stem cell transplantation (HSCT) for prophylaxis of graft rejection and acute graft-versus-host disease. However, the pharmacokinetics of MMF seem to differ when applied in HSCT compared to SOT. Here, we analyzed pharmacokinetics of mycophenolic acid (MPA), the active metabolite of MMF, in a nonmyeloablative canine HSCT model. Dogs received nonmyeloablative TBI for conditioning followed by leukocyte antigen-identical littermate HSCT and immunosuppression containing cyclosporin A (CsA) and different doses of MMF. Pharmacokinetics were performed on days 2, 14 and 27. Dose escalation of MMF from 10 to 30 mg/kg tended to increase area under the curve (AUC) and the apparent oral clearance by 45 and 110%, respectively. Doses applied had no linear association with MPA concentration or blood trough level. No significant drug accumulation occurred over time. Using a twice daily MMF regimen, we conclude that an AUC of 30-60 mug/ml h as recommended for SOT cannot be reached in HSCT. Toxicities did not permit single doses higher than 30 mg/kg. Thus, if larger AUCs are desired in order to assure sufficient immunosuppression in HSCT, MMF might have to be administered at least three times daily.     

Cost-effective production of a vaginal protein microbicide to prevent HIV transmission

A series of small-molecule microbicides has been developed for vaginal delivery to prevent heterosexual HIV transmission, but results from human clinical trials have been disappointing. Protein-based microbicides, such as HIV-specific monoclonal antibodies, have been considered as an alternative approach. Despite their promising safety profile and efficacy, the major drawback of such molecules is the economy of large-scale production in mammalian cells, the current system of choice. Here, we show that an alternative biomanufacturing platform is now available for one of the most promising anti-HIV antibodies (2G12). Our data show that the HIV-neutralization capability of the antibody is equal to or superior to that of the same antibody produced in CHO cells. We conclude that this protein production system may provide a means to achieve microbicide ingredient manufacture at costs that would allow product introduction and manufacture in the developing world.     

Transfusion-associated Serratia marcescens infection: studies of the mechanism of action

The growth of two strains of Serratia marcescens in blood components was tested in this study. One of the strains had been implicated in the epidemic of transfusion-associated sepsis experienced in Denmark and Sweden in 1991. In whole blood with a final concentration of 100 colony- forming units per mL of S. marcescens, there was an immediate reduction of more than 95 percent of colony-forming units, but no reduction of the bacterial concentration if the blood had been white cell-reduced before inoculation. This is interpreted as an effect due to phagocytosis by white cells and as a lack of bactericidal effect of the plasma. A reduction to 10 percent of the original concentration, observed if the blood had a nominal content of white cells, was most likely due to phagocytosis. White cell reduction by filtration after inoculation further reduced the bacterial concentration of one of the strains tested, but, after a 1-week lag phase, growth accelerated to high concentrations by 6 weeks. In platelet-rich plasma prepared from S. marcescens-inoculated units, abundant growth was found after 24 hours, increasing to very high concentrations (10(12) colony-forming units/mL) during 10-day storage at 22 +/− 2 degrees C. Keeping the whole blood at ambient temperature for 20 hours before preparation of platelet-rich plasma caused only temporary reduction of bacterial concentration in the S. marcescens experiments, but resulted in a complete absence of bacteria in the platelet-rich plasma for 10 days in control experiments performed with Staphylococcus epidermidis.(ABSTRACT TRUNCATED AT 250 WORDS)     

食欲素和食欲素受体与运动

目的:食欲素是一种重要的神经肽,通过了解食欲素的相关作用及其表达的调节因素等,可以为肥胖的发生机制及其防治工作,以及在运动中的研究开辟新的途径。资料来源:应用计算机检索1998/2006Medline的相关文章,检索词"orexin""hypocretin",并限定文章语言种类为英文。同时计算机检索1998/2006中国期刊全文数据库(http://www.cnki.net)的相关文章,检索项"关键词",检索词"食欲素""食欲肽""增食因子""胖素""阿立新素",限定文章语言种类为中文。共检索到200余篇文献。资料选择:对资料进行初审,纳入标准:①食欲素及其受体研究。②影响食欲素表达因素的研究。③食欲素与肥胖关系的研究。④食欲素与运动行为关系的研究。排除标准:重复性研究。资料提炼:共收集到56篇相关文献,中文文献均为全文,大部分外文文献为全文,其他为摘要。在这些文献中18篇为重复或类似的同一研究,38篇文献纳入分析。资料综合:食欲素是orphanG蛋白质共轭受体的配体,是下丘脑重要的神经肽。可分为食欲素A和食欲素B两种,它们来源于同一前体,其神经纤维贯穿脑和脊髓。食欲素受体分为OX1R和OX2R两型,OX1RmRNA和OX2RmRNA在脑内的分布不同。食欲素有调节摄食和能量平衡、调节睡眠觉醒状态、调节饮水等功能;其表达量受睡眠、营养状况、环境、性别等因素的影响。国内对食欲素的研究不多,主要是肥胖、低氧以及低糖状态对食欲素影响的研究;国外可见通过行为干预对食欲素的研究,以及其他因子对食欲素的影响。结论:食欲素作为一种重要的神经肽,除具有调节食欲和平衡能量的作用外,还具有保持觉醒,参与免疫应答等功能;食欲素抗体或受体拮抗剂还可能治疗肥胖、糖尿病等能量代谢失衡性疾病。但国内外对食欲素在运动训练及减肥中的相关研究还较少。     

运动疲劳大鼠能量代谢与红景天苷的影响

目的:已有研究表明,红景天具有抗缺氧、抗疲劳、抗血乳酸含量增加的作用。实验观察红景天苷对运动疲劳大鼠能量代谢的影响。方法:实验于2006-11/12在哈尔滨商业大学药物研究所药理实验室完成。①实验材料:高山红景天,红景天苷含量为3.065%;清洁级成年雄性SD大鼠40只,体质量160～180g。②实验分组:将大鼠随机分为4组(10只/组):即空白对照组、低、中、高剂量组。③实验过程:经口灌胃,空白对照组给予生理盐水,低、中、高剂量组分别给予红景天苷100,200,300mg/kg,连续灌胃14d。末次灌胃30min后,在大鼠尾根部负重5%体质量的铅皮,进行力竭性游泳。④实验评估:比较对照组与给药组大鼠游泳存活时间的长短;观察力竭游泳后各组大鼠血乳酸、血清尿素氮、血清肌酸激酶和肝糖元的变化。结果:每组取8只,共32只进入结果分析。①给药组大鼠游泳存活时间较对照组明显延长,差异有非常显著性意义(P<0.01),且高剂量组大鼠游泳存活时间最长。②与对照组相比,给药组对大鼠运动后的血乳酸、血清尿素氮、血清肌酸激酶有降低作用(P<0.05,P<0.01),肝糖元含量有所增加(P<0.01)。结论:红景天苷可以改善大鼠运动后的整体状况,可以通过调节能量代谢平衡提高大鼠的运动能力,有一定的抗疲劳作用。     

Expression and characterization of an iron-regulated hemin-binding protein, HbpA, from Leptospira interrogans serovar Lai

In an earlier study, based on the ferric enterobactin receptor FepA of Escherichia coli, we identified and modeled a TonB-dependent outer membrane receptor protein (LB191) from the genome of Leptospira interrogans serovar Lai. Based on in silico analysis, we hypothesized that this protein was an iron-dependent hemin-binding protein. In this study, we provide experimental evidence to prove that this protein, termed HbpA (hemin-binding protein A), is indeed an iron-regulated hemin-binding protein. We cloned and expressed the full-length 81-kDa recombinant rHbpA protein and a truncated 55-kDa protein from L. interrogans serovar Lai, both of which bind hemin-agarose. Assay of hemin-associated peroxidase activity and spectrofluorimetric analysis provided confirmatory evidence of hemin binding by HbpA. Immunofluorescence studies by confocal microscopy and the microscopic agglutination test demonstrated the surface localization and the iron-regulated expression of HbpA in L. interrogans. Southern blot analysis confirmed our earlier observation that the hbpA gene was present only in some of the pathogenic serovars and was absent in Leptospira biflexa. Hemin-agarose affinity studies showed another hemin-binding protein with a molecular mass of approximately 44 kDa, whose expression was independent of iron levels. This protein was seen in several serovars, including nonpathogenic L. biflexa. Sequence analysis and immunoreactivity with specific antibodies showed this protein to be LipL41.     

The role of protein glycosylation in allergy

The asparagine-linked carbohydrate moieties of plant and insect glycoproteins are the most abundant environmental immune determinants. They are the structural basis of what is known as cross-reactive carbohydrate determinants (CCDs). Despite some structural variation, the two main motifs are the xylose and the core-3-linked fucose, which form the essential part of two independent epitopes. Plants contain both epitopes, insect glycoproteins only fucose. These epitopes and other fucosylated determinants are also found in helminth parasites where they exert remarkable immunomodulatory effects. About 20% or more of allergic patients generate specific anti-glycan IgE, which is often accompanied by IgG. Even though antibody-binding glycoproteins are widespread in pollens, foods and insect venoms, CCDs do not appear to cause clinical symptoms in most, if not all patients. When IgE binding is solely due to CCDs, a glycoprotein allergen thus can be rated as clinical irrelevant allergen. Low binding affinity between IgE and plant N-glycans now drops out as a plausible explanation for the benign nature of CCDs. This rather may result from blocking antibodies induced by an incidental 'immune therapy' ('glyco-specific immune therapy') exerted by everyday contact with plant materials, e.g. fruits or vegetables. The need to detect and suppress anti-CCD IgE without interference from peptide epitopes can be best met by artificial glycoprotein allergens. Hydroxyproline-linked arabinose (single beta-arabinofuranosyl residues) has been identified as a new IgE-binding carbohydrate epitope in the major mugwort allergen. However, currently the occurrence of this O-glycan determinant appears to be rather restricted.     

Antitumor effects of hydrogen peroxide in vivo

Glucose oxidase, covalently coupled to polystyrene microspheres (GOL), produced H(2)0(2) at an average rate of 3.6 nmol/min per 10(9) beads under standard assay conditions. Injection of 1.3 × 10(10) to 1.1 × 10(11) GOL i.p. prolonged the survival of mice by 27 percent after injection of 10(6) P388 lymphoma cells in the same site, consistent with destruction of 97.6 percent of the tumor cells. Placing mice for several hours in 100 percent O(2), the probable rate-limiting substrate for GOL, afforded a 42 percent prolongation of survival from P388 lymphoma, consistent with destruction of 99.6 percent of the tumor cells. When the P388 inoculum was 10(5), 10(4), or 10(3) cells, GOL led to long-term survival (presumed cure) of 23 percent, 77 percent, and 92 percent of the mice, respectively, consistent with reduction of the injected tumor dose to less than 10 cells. Subcutaneous growth of 10(5) P388 cells (approximately 300 lethal dose to 50 percent of mice) was suppressed in 83 percent of mice by admixture of GOL with the tumor cell inoculum. GOL alone had no effect against a more peroxide-resistant tumor, P815 mastocytoma. However, P815 cell glutathione reductase could be inhibited in vivo by well-tolerated doses of the antitumor agent, 1,3-bis(2-chloroethyl)- 1-nitrosourea (BCNU). BCNU alone cured few mice with P815. Together, BCNU and GOL apparently cured 86 percent of mice injected with 10(6) P815 cells i.p. The protective effect of GOL was abolished by boiling it to inactivate the enzyme, by co-injection of catalase coupled to latex beads, or by delaying the injection of tumor cells for 3 h, by which time the beads had formed aggregates. Soluble glucose oxidase, in doses threefold higher than that bound to GOL, had no detectable antitumor effect. A single injection of preformed H(2)0(2) readily killed P388 cells in the peritoneal cavity, but only at doses nearly lethal to the mice. In contrast, GOL had very little toxicity, as judged by the normal appearance of the mice for over 400 d, gross and microscopic findings at autopsy, and various blood tests. GOL injected i.p. remained in the peritoneal cavity, where it was gradually organized into granulomata by macrophages, without generalized inflammation. Thus, an H(2)0(2)-generating system confined to the tumor bed exerted clear- cut antitumor effects with little toxicity to the host.     

Exposure to anthrax toxin alters human leucocyte expression of anthrax toxin receptor 1

Anthrax is a toxin‐mediated disease, the lethal effects of which are initiated by the binding of protective antigen (PA) with one of three reported cell surface toxin receptors (ANTXR). Receptor binding has been shown to influence host susceptibility to the toxins. Despite this crucial role for ANTXR in the outcome of disease, and the reported immunomodulatory consequence of the anthrax toxins during infection, little is known about ANTXR expression on human leucocytes. We characterized the expression levels of ANTXR1 (TEM8) on human leucocytes using flow cytometry. In order to assess the effect of prior toxin exposure on ANTXR1 expression levels, leucocytes from individuals with no known exposure, those exposed to toxin through vaccination and convalescent individuals were analysed. Donors could be defined as either ‘low’ or ‘high’ expressers based on the percentage of ANTXR1‐positive monocytes detected. Previous exposure to toxins appears to modulate ANTXR1 expression, exposure through active infection being associated with lower receptor expression. A significant correlation between low receptor expression and high anthrax toxin‐specific interferon (IFN)‐γ responses was observed in previously infected individuals. We propose that there is an attenuation of ANTXR1 expression post‐infection which may be a protective mechanism that has evolved to prevent reinfection.