Pam3CSK4诱导肥大细胞产生IL-6的效应研究

目的探讨Pam3CSK4对肥大细胞产生IL-6的影响及其涉及的机制。方法体外诱导培养小鼠骨髓来源的肥大细胞(bone marrow-derived mast cells,BMMCs),通过RT-PCR、Western blot检测BMMCs Toll样受体2(toll-like receptor 2,TLR2)基因和蛋白的表达;在有或无SB203580预处理的情况下,Pam3CSK4刺激BMMCs 24 h后,ELISA检测上清中IL-6的含量;同时应用Western blot对Pam3CSK4作用后BMMCs p38MAPK磷酸化的情况进行检测。结果 BMMCs存在TLR2的表达;BMMCs受Pam3CSK4(10~100 ng/ml)刺激后,IL-6的分泌明显增加(P0.01),并且出现了明显的p38MAPK磷酸化;与Pam3CSK4单刺激组相比,SB203580与Pam3CSK4联合刺激组IL-6的含量明显降低(P0.01)。结论 Pam3CSK4诱导肥大细胞产生IL-6,这一效应涉及p38MAPK信号途径。     

Pam3CSK4对血小板活化及其TLR2表达的影响

Objective To investgate role of TLR2 in the activation, the innate immune and inflammation of human platelets. Methods Human washed platelets were separated from healthy people(n=5) and were stimulated with different concentrations(1μg/ml, 5μg/ml, 10μg/ml) of TLR2 agonistPam3CSK4(a synthetic bacterial lipoproteins). Then the platelet aggregation rate, the expression of CD62p and TLR2 on the platelet surface were measured. Results The platelet aggregation rate were (28.32±5.67)%, (52.56±8.54)% and (76.24±11.23)%, respectively, at concentration of 1μg/ml, 5μg/mland 10μg/ml of Pam3CSK4, more than (12.83±2.43)% at 0μg/ml of it. In addition, the expression of CD62p were (18.45±2.66)%, (22.45±2.04)%, (29.53±4.08)%, respectively at above concentration of Pam3 CSK4, more than (11.20±1.67)% of CD62p at control group(P<0.01). The expression of TLR2 was not significantly increased at a lower concentration of Pam3CSK4(1μg/ml) with (16.85±6.10)% compared with(10.81±3.99)% at the control group. However, it were (21.15±9.90)% and (22.52±9.26)%, respectively, at a higher concentration(5μg/ml, 10μg/ml)more than(10.81±3.99)% at the control group(P<0.05). Conclusion Pam3CSK4 induce aggregation, activation and the up-regulation of TLR2 of platelet by stimulating TLR2 receptor of it. Thereafter, TLR2 play an important role in the innate immune of platelet.     

Pam3CSK4对血小板活化及其TLR2表达的影响

Objective To investgate role of TLR2 in the activation, the innate immune and inflammation of human platelets. Methods Human washed platelets were separated from healthy people(n=5) and were stimulated with different concentrations(1μg/ml, 5μg/ml, 10μg/ml) of TLR2 agonistPam3CSK4(a synthetic bacterial lipoproteins). Then the platelet aggregation rate, the expression of CD62p and TLR2 on the platelet surface were measured. Results The platelet aggregation rate were (28.32±5.67)%, (52.56±8.54)% and (76.24±11.23)%, respectively, at concentration of 1μg/ml, 5μg/mland 10μg/ml of Pam3CSK4, more than (12.83±2.43)% at 0μg/ml of it. In addition, the expression of CD62p were (18.45±2.66)%, (22.45±2.04)%, (29.53±4.08)%, respectively at above concentration of Pam3 CSK4, more than (11.20±1.67)% of CD62p at control group(P<0.01). The expression of TLR2 was not significantly increased at a lower concentration of Pam3CSK4(1μg/ml) with (16.85±6.10)% compared with(10.81±3.99)% at the control group. However, it were (21.15±9.90)% and (22.52±9.26)%, respectively, at a higher concentration(5μg/ml, 10μg/ml)more than(10.81±3.99)% at the control group(P<0.05). Conclusion Pam3CSK4 induce aggregation, activation and the up-regulation of TLR2 of platelet by stimulating TLR2 receptor of it. Thereafter, TLR2 play an important role in the innate immune of platelet.     

Pam3CSK4对血小板活化及其TLR2表达的影响

目的 探讨Toll样受体2(TLR2)在血小板激活及免疫方面的作用.方法 取健康人(n=5)全血6 ml,以密度梯度离心法制备洗涤血小板.用1、5、10μg/ml的TLR2激动剂Pam3CSK4(一种合成的细菌脂蛋白)刺激人洗涤血小板,然后测定血小板聚集率,血小板表面蛋白CD62p和TLR2表达的变化.结果 Pam3CSK4以浓度0、1、5和10μg/ml激活血小板:聚集率增加分别为(12.83±2.43)%、(28.32±5.67)%、(52.56±8.54)%、(76.24±11.23)%,P<0.01;血小板表面CD62p表达量增加分别为(11.20±1.67)%、(18.45±2.66)%、(22.45±2.04)%、(29.53±4.08)%,P<0.01.Pam3CSK4在1μg/ml时TLR2表达为(16.85±6.10)%,与对照组(10.81±3.99)%相比差异无统计学意义(P>0.05),在5 μg/ml、10μg/ml时TLR2表达量分别为(21.15±9.90)%和(22.52±9.26)%,与对照组比较差异有统计学意义(P<0.05).结论 细菌脂蛋白Pam3CSK4通过激活TLR2引起血小板聚集、活化,是血小板参与抑制革兰阳性细菌感染的免疫应答机制之一.     

The Toll-like receptor 1/2 agonists Pam(3) CSK(4) and human β-defensin-3 differentially induce interleukin-10 and nuclear factor-κB signalling patterns in human monocytes

Human β-defensin 3 (hBD-3) activates antigen-presenting cells through Toll-like receptors (TLRs) 1/2. Several TLR1/2 agonists have been identified but little is known about how they might differentially affect cellular activation. We compared the effects of hBD-3 with those of another TLR1/2 agonist, Pam(3) CSK(4) , in human monocytes. Monocytes incubated with hBD-3 or Pam(3) CSK(4) produced interleukin-6 (IL-6), IL-8 and IL-1β, but only Pam(3) CSK(4) induced IL-10. The IL-10 induction by Pam(3) CSK(4) caused down-modulation of the co-stimulatory molecule, CD86, whereas CD86 expression was increased in monocytes exposed to hBD-3. Assessment of signalling pathways linked to IL-10 induction indicated that mitogen-activated protein kinases were activated similarly by hBD-3 or Pam(3) CSK(4) , whereas the non-canonical nuclear factor-κB pathway was only induced by Pam(3) CSK(4) . Our data suggest that the lack of non-canonical nuclear factor-κB signalling by hBD-3 could contribute to the failure of this TLR agonist to induce production of the anti-inflammatory cytokine, IL-10, in human monocytes.     

在MRSA系统性感染模型中Pam3CSK4预处理降低小鼠肾组织的炎症反应

目的:在耐甲氧西林金葡菌(MRSA)系统性感染模型中,观察低剂量TLR2激动剂Pam3CSK4预处理对小鼠肾组织中炎症反应的影响并初步探讨其机制。方法:于感染前48 h、24 h对BALB/c小鼠行尾静脉注射Pam3CSK4(10μg/100μl/只);2×10~7CFU/只MRSA经静脉感染小鼠,ELISA和荧光实时定量PCR(Q-PCR)检测细胞因子水平,Q-PCR检测TLR2、IRAKs等相对表达量,Western blot检测NF-κB p65磷酸化、IRAK-M及A20表达。结果:与对照组相比,预处理组在感染6 h后肾组织中TNF-α、IL-6、IL-1β、CCL3和IFN-γ含量显著减少,i NOS表达量降低,IL-10和TGF-β表达量增高,TLR2表达下降;处理组肾组织在感染12 h后IRAK-1表达无明显增加,而IRAK-M表达量显著增加;Western blot结果显示Pam3CSK4预处理组NF-κBp65磷酸化降低,IRAK-M表达明显增高。结论:Pam3CSK4预处理降低MRSA系统性感染小鼠肾组织的炎症反应,这可能与诱导IRAK-M表达相关。     

Transforming growth factor-β (TGFβ) inhibition of epstein-barr virus (EBV)- and interleukin-4 (IL-4)-induced immunoglobulin production in human B lymphocytes

This study reports on the effects of TGF on the secretion of Ig isotypes by highly purified (>99% CD20-positive) human peripheral blood B cells. Stimulation of these B cell preparations with EBV resulted in the secretion of IgM, IgG, and IgA and the addition of IL-4 induced readily detectable levels (>100 ng/ml) of IgE between 10 and 25 days of culture. TGF1 and TGF2 showed similar dose-dependent suppression of IgM, IgG, and IgA, and the relative proportion of IgG and IgA remained unchanged in the presence of TGF. IgE production induced by EBV and IL-4 was significantly inhibited by TGF. TGF effects on Ig secretion were not related to inhibition of B cell proliferation by this cytokine. In contrast to these TGF effects on EBV activation of primary B cells, the constitutive Ig secretion by EBV-transformed B cells was resistant to TGF, while the increase in Ig secretion induced by IL-6 was inhibited by TGF. Thus, TGF inhibits the EBV-induced secretion of the major Ig isotypes in peripheral blood B cells and has differential effects on Ig secretion by transformed B cells.     

共刺激分子4-1BB及其配体4-1BB Ligand(4-1BBL)与肿瘤免疫

1970年Bretscher和Cohn首先提出“T细胞激活双信号”假说以来 ,细胞免疫“双信号”学说得到大量的体外实验的证实[1] 。介导共刺激信号的分子有多种 ,4 1BB与 4 1BBL是其中非常重要的一个成员。4 1BB 4 1BBL介导的共刺激信号可以诱导T细胞的活化、增殖及抗凋亡 ,维持T细胞的长期生存及提高细胞因子的分泌 ,4 1BB还能通过CD2 8非常依赖途径刺激T细胞的增殖并在树突状细胞和单核细胞介导的细胞免疫中起作用。因此 ,4 1BB和其配体 4 1BBL介导的共刺激信号是激发有效的抗肿瘤细胞免疫应答尤其是长期抗肿瘤效应所必需的 ,有望为…     

《中国组织工程研究与临床康复》(CRTER)杂志执行编委名单(2010-4/2011-4)

<正>经过审稿工作的考核及编委会讨论,聘任下述专家为本刊执行编委,聘任时间为2010年4月至2011年4月,任期1年。感谢这些执行编委和审稿专家为保证本刊客观、公正、及时、规范的审稿工作所做出的贡献。     

2-[(4-甲基-反-丙烯酸酯)苯基]-4,5-二-(4-N,N-甲基异丙基-氨基苯基)-1(H)-咪唑克服肿瘤多药耐药性作用及其机制

目的 :研究一种新化合物 2 [(4 甲基 反 丙烯酸酯 ) ] 4,5 二 (4 N ,N 甲基异丙基 氨基苯基 ) 1 (H) 咪唑 (FG0 2 0 3 2 6)对人乳腺癌细胞阿霉素耐药株 (MCF 7/adr)多药耐药性 (Multidrugresistance ,MDR)的逆转作用及其机制。方法 :采用MTT法检测药物细胞毒性作用 ,荧光分光光度计法检测该化合物对MCF 7/adr细胞及MCF 7细胞内ADM积累的影响。结果 :MCF F/adr较其相应敏感株对ADM耐药 99倍FG0 2 0 3 2 6对MCF 7/adr细胞及MCF 7细胞的IC50均大于 5 0 μm ,在≤ 1 0 μM浓度时对上述细胞无细胞毒作用 ,细胞存活大于…     

(E)-苯乙基-3-(3,5-二羟基-4-异丙基苯基)丙烯酸酯的抗炎作用及抗炎初步机制研究

目的研究(E)-苯乙基-3-(3,5-二羟基-4-异丙基苯基)丙烯酸酯(THCA354)化合物的抗炎作用及抗炎初步机制。方法通过二甲苯和佛波酯诱导的小鼠耳肿胀模型来评价THCA354的抗炎作用;进一步通过佛波酯诱导的小鼠耳肿胀模型来评价该化合物对小鼠耳组织中髓过氧化酶(MPO)的活性和IL-1β、IL-6、TNF-α炎症细胞因子的抑制作用。结果与模型组相比,局部涂抹THCA354化合物的小鼠耳朵厚度,淋巴细胞的浸润,耳组织中髓过氧化物酶活性和IL-1β、IL-6、TNF-α炎症细胞因子的表达均显著降低。结论 THCA354化合物可能通过抑制中性粒细胞的活化和IL-1β、IL-6、TNF-α炎症细胞因子的释放从而有效地缓解小鼠耳组织炎症水肿和淋巴细胞的浸润,发挥抗炎作用。     

(E)-苯乙基-3-(3,5-二羟基-4-异丙基苯基)丙烯酸酯对小鼠皮炎湿疹的治疗作用初步研究

目的研究(E)-苯乙基-3-(3,5-二羟基-4-异丙基苯基)丙烯酸酯(THCA354)凝胶剂对小鼠变应性接触性皮炎(ACD)的治疗作用及机制。方法通过对小鼠耳廓涂抹2,4-二硝基氟苯(DNFB)诱导建立ACD模型,测定给予DNFB刺激后24 h的耳肿胀厚度和对小鼠耳组织进行病理组织形态学分析来共同评价THCA354凝胶剂的抗过敏作用效果;采用ELISA法和RT-PCR定量检测小鼠耳组织中炎症细胞因子含量及m RNA表达水平,初步阐明其作用机制。结果与阴性对照组相比,局部涂抹THCA354的小鼠耳肿胀程度、淋巴细胞浸润程度、耳组织匀浆中促炎细胞因子含量和m RNA表达水平均显著降低,抗炎细胞因子的含量和m RNA表达水平均显著升高。结论 THCA354可能是通过改变炎症细胞因子的分泌和m RNA表达水平,调节T细胞亚群的平衡从而发挥抗过敏作用。     

Sensitization to airborne and food allergens in Reykjavík (Iceland) and Uppsala (Sweden) - a comparative study

BACKGROUND: The aim of this study was to compare the prevalence of atopic sensitization and possible risk factors for allergies in two ethnically similar but geographically widely separated urban populations. METHODS: Data from two centers of the European Community Respiratory Health Survey, Reykjavik, Iceland, and Uppsala, Sweden, were utilized. This included a structured interview, skin prick tests, and blood samples for total and specific IgE for common aeroallergens. Additional measurements of specific IgE antibodies to common food antigens were performed. Furthermore, data on social environment, lifestyle, air pollution, and meteorologic variables were compared. RESULTS: Skin prick tests were done on 540 individuals in Reykjavik and 527 in Uppsala. The overall prevalence of at least one positive prick test was 20.5% in Reykjavik and 34.2% in Uppsala (P<0.001). Total and specific IgE were measured in serum from 521 subjects in Reykjavik and 472 in Uppsala. The geometric mean value for total IgE was significantly lower in Reykjavik (13.4 kU/l) than in Uppsala (24.7 kU/l) (P<0.001). Similarly, the overall prevalence of at least one specific IgE to airborne allergens was 23.6% in Reykjavik and 32.3% in Uppsala (P<0.01). Specific IgE to a food panel (fx5) was measured in 502 subjects in Reykjavik, and 434 in Uppsala. In Reykjavik, 20 individuals (4.0%) were positive to one or more of the allergens in the food panel compared to 27 (6.0%) in Uppsala. When the single allergens present in the food panel were measured, altogether 16 positive reactions were found in Reykjavik compared to 47 in Uppsala (P<0.05). CONCLUSIONS: The prevalence of sensitization to both airborne and food allergens was lower in Reykjavik than in Uppsala. The difference may be due to environmental and/or dietary differences or to some yet undefined factor.     

Comparison of interferon-γ-, interleukin (IL)-17- and IL-22-expressing CD4 T cells, IL-22-expressing granulocytes and proinflammatory cytokines during latent and active tuberculosis infection

In this study, we investigated the role and expression of T helper type 17 (Th17) cells and Th17 cytokines in human tuberculosis. We show that the basal proportion of interferon (IFN)-γ-, interleukin (IL)-17- and IL-22-expressing CD4(+) T cells and IL-22-expressing granulocytes in peripheral blood were significantly lower in latently infected healthy individuals and active tuberculosis patients compared to healthy controls. In contrast, CD4(+) T cells expressing IL-17, IL-22 and IFN-γ were increased significantly following mycobacterial antigens stimulation in both latent and actively infected patients. Interestingly, proinflammatory IFN-γ and tumour necrosis factor (TNF)-α were increased following antigen stimulation in latent infection. Similarly, IL-1β, IL-4, IL-8, IL-22 and TNF-α were increased in the serum of latently infected individuals, whereas IL-6 and TNF-α were increased significantly in actively infected patients. Overall, we observed differential induction of IL-17-, IL-22- and IFN-γ-expressing CD4(+) T cells, IL-22-expressing granulocytes and proinflammatory cytokines in circulation and following antigenic stimulation in latent and active tuberculosis.     

Pam3Csk4 预处理增强巨噬细胞对耐甲氧西林金黄色葡萄球菌杀菌作用

目的:初步探讨TLR2激动剂Pam3Csk4预处理后,小鼠腹腔巨噬细胞对耐甲氧西林金葡菌(Methicillin-resistant S.aureus,MRSA)的免疫反应性。方法:1μg/ml Pam3Csk4作用于鼠腹腔巨噬细胞,12 h后以热灭活耐甲氧西林金葡菌刺激细胞,ELISA和荧光定量PCR(Q-PCR)法分别检测培养细胞中TNF-α、IL-6和IL-1及mRNA含量,流式检测小鼠腹腔巨噬细胞对热灭活金葡菌的吞噬能力,平板计数法检测Pam3Csk4预处理巨噬细胞对金葡菌杀菌能力;Q-PCR法检测Pam3Csk4预处理巨噬细胞6 h和12 h后吞噬相关受体与补体受体、一氧化氮诱导合成酶(i NOS)及抗菌肽LL37基因表达。结果:金葡菌刺激后,Pam3Csk4预处理组TNF-α、IL-6、IL-1蛋白和基因水平均显著低于未处理组(P0.05),但Pam3Csk4预处理组细胞对金葡菌吞噬和杀菌能力均显著增强(P0.05),对于MRSA菌株,增强的杀菌能力在补体和抗体参与。进一步Q-PCR结果显示Pam3Csk4预处理巨噬细胞6 h和12 h后调理吞噬相关受体FCγRⅠ/Ⅲ与补体受体CR1/3表达均显著增强,i NOS和LL37基因表达也显著增加。结论:Pam3Csk4预处理小鼠腹腔巨噬细胞能增强其对金黄色葡萄球菌甲氧西林敏感和耐药菌株杀菌或抑菌能力,并降低其相应炎症反应,该现象可能与Pam3Csk4激活巨噬细胞吞噬相关受体以及i NOS和抗菌肽表达有关。     

Toll样受体2(TLR2)激动剂Pam3Csk4预处理对耐甲氧西林金黄色葡萄球菌攻击小鼠的保护作用

目的耐甲氧西林金黄色葡萄球菌(MRSA)感染日趋严重,免疫防治被寄予厚望。本研究旨在探索Toll样受体2(TLR2)激动剂Pam3Csk4预处理小鼠对MRSA攻击的保护作用。方法每只昆明小鼠用(25、50、100)μg Pam3Csk4尾静脉预处理12、24 h,小鼠尾静脉注射7×1010集落形成单位(CFU)/kg的MRSA(ATCC43300)。小鼠生存率按前24 h每2 h观察1次,后续每6 h记录小鼠存活情况,共观察1周。感染MRSA(3×108CFU/只)6 h后,菌落计数法观察肝、脾和肾等器官对MRSA的清除能力;3×108CFU/只MRSA攻击6、12、24 h后,ELISA检测小鼠血清中肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)、γ干扰素(IFN-γ)和IL-10含量,实时荧光定量PCR(q PCR)检测MRSA攻击后小鼠脾脏TNF-α、IL-6、IFN-γ和IL-10的mRNA水平,以及Pam3Csk4预处理小鼠24 h后,CXC趋化因子配体1(CXCL1)和Fcγ受体Ⅲ(FcγRⅢ)表达量。结果 100μg Pam3Csk4预处理能显著提高MRSA攻击小鼠的生存率,与Pam3Csk4预处理时间呈时间依赖关系;MRSA攻击小鼠的生存时间与Pam3Csk4预处理剂量相关,50μg以上剂量Pam3Csk4预处理能显著提高MRSA攻击小鼠生存率至70%以上;与对照组比较,Pam3Csk4预处理小鼠血清中TNF-α在6 h和12 h均显著降低,24 h后2组间无显著差别;IL-6在处理6 h内显著降低,12 h和24 h后无显著差异,IFN-γ在24 h内均显著降低,而IL-10则无显著变化;Pam3Csk4预处理24 h后,脾脏中CXCL1和FcγRⅢ表达量均显著高于生理盐水组。结论 Pam3Csk4预处理对MRSA攻击小鼠具有保护作用。     

(Z)-1-溴-2-(对氯苯基)-1,2-双(对甲氧苯基)—乙烯晶体结构测定

题目化合物(C_(22)H_(18)O_2Cl Br)经X-光衍射研究,确定了它的分子结构。晶体属单斜晶系,空间群P21/n,晶胞参数a=19.502,b=9.118,c=11.233;结构由MuLTAN-80(P·MAIN等1980)确定。从E—图上确定了溴原子位置。以溴原子为起始相角计算加权Fourier,在Fourier图上定出了全部26个非氢原子坐标·原子坐     

α- and β-Poly(Vinylidene Fluoride) Evoke Different Cellular Behaviours

α-Phase poly(vinylidene fluoride) (PVDF) has chains of zero dipole moments and is, therefore, nonpiezoelectric, while β-phase PVDF has the most significant piezoelectric properties among the polymorphs due to its polar chains. Although many reports describe PVDF as a suitable biomaterial due to its stability and biocompatibility, few considered the specific effects that the different polymorphs exert on cellular behaviour. We hypothesized that α- and β-phase PVDF will exert direct but different influences on cell attachment and metabolic activity. PVDF films were fabricated using N,N-dimethylformamide (DMF) and hexamethylphosphoramide (HMPA) by solvent casting. Samples were characterized by differential scanning calorimetry, Fourier transform infrared spectroscopy and X-ray diffraction. Films containing 83.5% α-phase PVDF (DMF–PVDFα) and 91.4% of β-phase PVDF (HMPA–PVDFβ within the crystalline regions were produced and used to evaluate in vitro attachment and metabolic activity of L929 cells. Cell metabolic activity on both PVDF conformations increased 3-fold over the 1-week culture period, with higher cell metabolic activity observed on DMF–PVDFα on day 5 of culture, compared to HMPA–PVDFβ. Cells grown on DMF–PVDFα were well-spread, flat and expressed spotted paxillin in focal adhesions that were mainly localized to perinuclear regions of the cells, while a high proportion of cells on HMPA–PVDFβ were bulging, round and expressed relatively fewer paxillin spots. Our results suggest that α-phase PVDF supports higher cell metabolic activity and better cell spreading compared to β-phase PVDF. Such variations can potentially be exploited for different biomedical applications.