小鼠沙眼衣原体肺感染过程中Treg细胞与Th17反应关系

[目的]对沙眼衣原体在BALB/c小鼠肺部感染过程中CD4+ CD25+Foxp3+调节性T细胞(regulatory T cells,Treg)与Th17反应关系进行初步探讨.[方法]取6-8周龄的BALB/c小鼠,鼻腔吸入25 μL含5×103 IFU的沙眼衣原体鼠肺炎菌株(Chlamydia muridarum,Cm),建立沙眼衣原体小鼠肺感染模型.监测感染后不同时期小鼠体重变化;检测肺组织衣原体包涵体形成单位( IFU)及肺组织病理改变;利用流式细胞术检测Cm感染后小鼠体内Treg细胞百分率;ELISA检测肺组织上清液IL-6、TGF-β、IL-17、IL-2细胞因子的的表达;qRT-PCR检测KC( keratinocyte derived chemokine) mRNA和MIP-2( macrophage inflammatory protein-2)mRNA的表达差异.[结果]用5xl03 IFU Cm经鼻腔吸人感染后小鼠发生沙眼衣原体肺炎,表现为体重下降、肺组织大量炎症细胞浸润并可检测到衣原体繁殖.Cm感染后第3天,小鼠体内Treg细胞占CD4 +T细胞的百分比明显降,随后开始恢复,第7天恢复原来水平,一直持续到衣原体清除.TGF-β、IL-2的表达与Treg细胞动态变化一致.与Th17相关细胞因子IL-6、IL-17和Th17相关趋化因子KC、MIP-2的表达于第3天开始升高,至第7天达到最高水平,随后逐渐减少.[结论]在衣原体感染BALB/c小鼠过程中,Treg可能通过提供TGF-β并在IL-6帮助下促进Th17应答产生.     

IL-37对粉尘螨诱导气道上皮及固有免疫细胞产生IL-6的作用研究

目的探讨气道上皮细胞及固有免疫细胞经粉尘螨刺激后,白细胞介素37(interleukin 37,IL-37)对其产生细胞因子IL-6的影响。方法体外培养人肺泡上皮细胞系A549、小鼠肺上皮细胞系MLE-12、小鼠巨噬细胞系RAW264.7和原代小鼠固有淋巴样2型细胞(ILC2细胞),当细胞融合度达70%时,用IL-37预处理2 h,再给予粉尘螨粗提物刺激细胞,每种细胞均设PBS、IL-37及粉尘螨对照,并分别于不同时间点收集细胞沉淀和细胞培养液上清;用实时聚合酶链反应(real-time polymerase chain reaction,real-time PCR)法检测细胞因子IL-6在mRNA水平上的变化,用双抗体夹心ELISA检测细胞因子IL-6在蛋白水平的变化,用流式细胞术分选ILC2细胞并检测ILC2细胞表面IL-37受体的表达情况。结果粉尘螨粗提物可促进A549细胞、MLE-12细胞、RAW264.7细胞和ILC2细胞表达I L-6,且呈时间依赖的方式(P0.05)。IL-37可抑制粉尘螨粗提物所诱导的IL-6在A549细胞、MLE-12细胞和RAW264.7细胞表达(P0.05);但IL-37对粉尘螨粗提物刺激ILC2细胞分泌IL-6无明显抑制作用(P0.05)。结论IL-37可抑制粉尘螨诱导的气道上皮细胞和巨噬细胞产生IL-6,并可通过负向调控下调气道炎症反应,为IL-37在哮喘治疗中的潜在应用提供了实验依据。     

肺炎支原体感染小鼠体内IL-6、IFN-γ、IL-5的水平及克拉霉素的影响

目的：研究肺炎支原体感染小鼠血清及支气管肺泡灌洗液中细胞因子的水平,了解克拉霉素治疗对细胞因子的影响。方法：将昆明小鼠分为感染组、正常组及药物治疗组,建立小鼠肺炎支原体感染模型成功后,药物组用克拉霉素（6g·kg^-1·d^-1）进行治疗,连续5天。在肺炎支原体感染模型建立后的第8天,检测三组小鼠的血清及支气管肺泡灌洗液中IL-6、IFN-γ、IL-5水平。结果-相比较于正常组,感染组小鼠支气管灌洗液和血清中的IL-6、IFN-γ、IL-5均显著升高（P〈0．05）。而克拉霉素的使用能使IL-6、IFN-γ降低（P〈0．05）,但对IL-5水平无影响。结论：肺炎支原体感染使小鼠IL-6、IFN-γ、IL-5水平增高,克拉霉素治疗有一定疗效。     

IL-13对小鼠AGR2表达的影响及其在哮喘气道黏液过度分泌中的作用

目的研究白细胞介素-13(IL-13)对AGR2mRNA和蛋白在哮喘小鼠肺组织中表达的影响,探讨AGR2在哮喘气道黏液过度分泌中的作用。方法 18只雌性小鼠随机分为哮喘组、对照组和IL-13组,IL-13组于26d-28d激发前经鼻滴入100μg重组小鼠IL-13。收集支气管肺泡灌洗液(BALF)计嗜酸性细胞分类计数。Real time-PCR方法检测肺组织AGR2mRNA表达。免疫组化法分别检测AGR2蛋白和Muc5ac蛋白在小鼠肺组织的表达。结果哮喘组BALF中嗜酸性细胞分类计数(19.1±6.34)%较正常组(0.28±0.29)%明显增多(P<0.01);IL-13组BALF中嗜酸性细胞分类计数(30.05±9.32)%较哮喘组明显升高(P<0.01)。IL-13组小鼠肺组织中AGR2mRNA(1.702±0.046)和蛋白(0.617±0.028)的表达较哮喘组(1.52±0.071,P<0.01;0.505±0.078,P<0.05)升高,IL-13组AGR2mRNA与Muc5ac蛋白的表达水平呈直线正相关(r=0.862,P<0.05);AGR2蛋白与Muc5ac蛋白水平呈直线正相关(r=0.847,P<0.05)。结论 AGR2可能参与了哮喘气道黏液过度分泌发病机制,IL-13可通过上调其表达,进一步促进黏蛋白Muc5ac表达。     

肺炎支原体感染小鼠体内IL-6、IFN-γ、IL-5的水平 及克拉霉素的影响

目的：研究肺炎支原体感染小鼠血清及支气管肺泡灌洗液中细胞因子的水平,了解克拉霉素治疗对细胞因子的影响。方法： 将昆明小鼠分为感染组、正常组及药物治疗组,建立小鼠肺炎支原体感染模型成功后,药物组用克拉霉素（6 g·kg-1·d-1）进行治疗, 连续5 天。在肺炎支原体感染模型建立后的第8 天,检测三组小鼠的血清及支气管肺泡灌洗液中IL-6、IFN-γ、IL-5 水平。结果： 相比较于正常组,感染组小鼠支气管灌洗液和血清中的IL-6、IFN-γ、IL-5 均显著升高(P ＜0.05)。而克拉霉素的使用能使IL-6、 IFN-γ降低(P ＜0.05),但对IL-5 水平无影响。结论：肺炎支原体感染使小鼠IL-6、IFN-γ、IL-5 水平增高,克拉霉素治疗有一定疗 效。     

Ghrelin抑制H202诱导的肺泡上皮A549细胞中IL-8的产生

目的：通过细胞培养的方法,初步研究了生长激素释放肽Ghrelin在氧化应激相关的肺泡上皮细胞炎症反应中的作用。方法：首先是用双氧水（H202）刺激A549细胞建立肺泡上皮细胞炎症反应模型,分别加入不同浓度的Ghrelin及普通培养基培养A549细胞。用酶联免疫吸附技术（ELISA）从蛋白水平检测上清液中IL-8的含量,采用逆转录聚合酶链反应技术（RT—PCR）检测炎性细胞因子IL．8mRNA的表达。结果：H2O2可以使A549细胞IL-8蛋白的释放及IL-8mRNA的表达明显升高（P〈0．05）,而用Gllrelin干预后IL．8的释放及IL-8mRNA的表达被抑制,明显低于单纯H2O2刺激的模型组（P〈0．05）,且随着浓度的增加Ghrelin的这种抑制作用逐渐增强。结论：分别从蛋白水平和基因水平证明了Ghrelin能够抑制H202诱导的肺泡上皮A549细胞中IL-8的产生,由此推测Ghrelin可能能够抑制以COPD、支气管哮喘为代表的氧化应激相关的肺部炎症反应。     

IL-17A对慢性阻塞性肺疾病的干预作用及其机制*

目的:探讨白细胞介素-17A(IL-17A)对慢性阻塞性肺疾病(COPD)的干预作用及其机制。方法:C57BL/6小鼠随机分为野生型空白对照组、野生型COPD组和IL-7A敲除COPD组,每组20只。野生型空白对照组小鼠不做任何处理,其余两组小鼠暴露于香烟烟雾(1支/次,4次/日,每次45 min,每次间隔时间为1 h,总干预时间为90 d)制作COPD模型。干预结束24 h后,利用动物肺功能检测系统测定小鼠肺功能。收集小鼠支气管肺泡灌洗液(BALF),测定BALF细胞计数和分类。收集小鼠肺组织,采用流式细胞法测定气道上皮IL-17A表达水平,采用酶联免疫吸附法测定肺组织炎症因子水平。采用蛋白免疫印迹法测定小鼠肺组织JNK/AP1信号通路蛋白表达水平。结果:与野生型空白对照组小鼠比较,野生型COPD组小鼠气道上皮IL-17A表达水平明显升高,吸气峰流速(PIF)和呼气峰流速(PEF)明显降低,BALF中性粒细胞、嗜酸性粒细胞、淋巴细胞和巨噬细胞数明显升高,肺组织CXC类趋化因子1(CXCL1)、CXC类趋化因子2(CXCL2)、白细胞介素-1β(IL-1β)和白细胞介素-6(IL-6)表达水平明显升高,JNK、cJun和cFos磷酸化水平及AP1表达水平明显升高(P＜0.05);与野生型COPD组小鼠比较,IL-7A敲除COPD组小鼠气道上皮IL-17A表达水平明显降低,PIF和PEF明显升高,BALF中性粒细胞、嗜酸性粒细胞、淋巴细胞和巨噬细胞数明显降低,肺组织CXCL1、CXCL2、IL-1β和IL-6表达水平明显降低,JNK、cJun和cFos磷酸化水平及AP1表达水平明显降低(P＜0.05)。结论:香烟烟雾可诱导小鼠气道上皮产生IL-17A,降低(或抑制)IL-17A的产生(或表达或分泌),通过抑制JNK/AP1信号通路,减轻COPD气道炎症反应,改善COPD小鼠肺功能。     

Ghrelin抑制H2O2诱导的肺泡上皮A549细胞中IL-8的产生

目的:通过细胞培养的方法,初步研究了生长激素释放肽Ghrelin在氧化应激相关的肺泡上皮细胞炎症反应中的作用.方法:首先是用双氧水(H2O2)刺激A549细胞建立肺泡上皮细胞炎症反应模型,分别加入不同浓度的Ghrelin及普通培养基培养A549细胞,用酶联免疫吸附技术(ELISA)从蛋白水平检测上清液中IL-8的含量,采用逆转录聚合酶链反应技术(RT-PCR)检测炎性细胞因子IL-8mRNA的表达.结果:H2O2可以使A549细胞IL-8蛋白的释放及IL-8mRNA的表达明显升高(P＜0.05),而用Ghrelin干预后IL-8的释放及IL-8mRNA的表达被抑制,明显低于单纯H2O2刺激的模型组(P＜0.05),且随着浓度的增加Ghrelin的这种抑制作用逐渐增强.结论:分别从蛋白水平和基因水平证明了Ghrelin能够抑制H2O2诱导的肺泡上皮A549细胞中IL-8的产生,由此推测Ghrelin可能能够抑制以COPD、支气管哮喘为代表的氧化应激相关的肺部炎症反应.     

敌草快中毒诱导小鼠急性肺损伤对IL-17表达的影响

目的观察敌草快中毒诱导小鼠急性肺损伤对IL17表达的影响。方法取健康雄性SD大鼠120只,按随机数字表法分为空白组、假手术组、中毒组,各40只。中毒组给予一次性腹腔注射20 mg/kg敌草快制备小鼠中毒模型,假手术组给予腹腔注射等剂量磷酸盐缓冲液造模,空白组未作任何处理。观察3组小鼠造模后6 h、24 h及72 h的死亡情况,并检测其肺组织干湿比重及肺泡灌洗液中IL-17表达情况。结果假手术组、空白组6 h、24 h及72 h均无小鼠死亡现象,而中毒组6 h、24 h及72 h小鼠死亡分别为2只、5只、2只,在24 h时死亡率达到高峰。假手术组、空白组6 h、24 h及72 h的小鼠肺组织干湿比重及肺泡灌洗液中IL-17表达均无明显变化;而中毒组小鼠各时段的肺组织干湿比重及肺泡灌洗液中IL-17表达明显增多,均高于空白组、假手术组(均P0.05),且在24 h时达到高峰。结论敌草快诱导小鼠急性肺损伤可促使IL-17分泌增多,因此推测下调IL-17的表达可以减少肺损伤。     

沙眼衣原体感染诱导小鼠生殖道局部促炎性细胞因子的表达情况

建立小鼠生殖道沙眼衣原体感染模型,观察小鼠生殖道局部促炎性细胞因子的表达。将小鼠生物型沙眼衣原体C. muridarum 1＆#215;104 IFU阴道接种于C57B6背景雌性小鼠,取感染后阴道拭子做沙眼衣原体培养,计算IFU,监测小鼠感染和病原体清除情况;80 d后处死小鼠,检测子宫输卵管病理改变;ELISA检测感染过程中小鼠生殖道促炎性细胞因子IL-1α、IL-6、MIP-2和TNF-α产生情况。小鼠感染在第3至第15天维持较高水平,然后病原体被逐渐清除,整个病程约3~5周;病理检测显示子宫输卵有严重炎症、管腔扩张积水,狭窄等;于感染后第3天检测到局部IL-1α、IL-6、MIP-2分泌,第7天达高峰,然后逐渐下降至正常水平（ IL-6于11 d恢复正常,IL-1α和 MIP-2于15 d恢复正常）。 TNF-α仅在第7天检测到高水平表达。相对于TNF-α和IL-6,IL-1α和MIP-2维持时间较长。成功建立沙眼衣原体感染小鼠生殖道模型,沙眼衣原体急性感染可诱导小鼠生殖道局部分泌IL-1α、IL-6、MIP-2和TNF-α。     

Endogenous IL-17 as a mediator of neutrophil recruitment caused by endotoxin exposure in mouse airways

We have previously demonstrated that administration of the recently described cytokine IL-17 in rat airways in vivo recruits and activates neutrophils locally. In the current study, we examined whether endogenous IL-17 is involved in mediating neutrophil recruitment caused by endotoxin exposure in mouse airways. Our in vivo data show that local endotoxin exposure causes the release of free, soluble IL-17 protein 6 h later. Systemic pretreatment with a neutralizing anti-IL-17 Ab almost completely inhibits neutrophil recruitment 24 h, but not 6 h, after endotoxin exposure in the airways. Pretreatment with neutralizing anti-IL-6 and anti-macrophage inflammatory protein (MIP)-2 Abs inhibits neutrophil recruitment caused by local endotoxin exposure and IL-17, respectively. Our in vitro data show that endotoxin exposure stimulates the release of soluble IL-17 protein in T lymphocytes harvested from lung and spleen, respectively, and that this cytokine release requires coculture with airway macrophages. Intracellular IL-17 protein is detected in T lymphocytes from spleen but not in airway macrophages after coculture and stimulation of these two cell types. Finally, anti-IL-17 does not alter endotoxin-induced release of IL-6 and MIP-2 from T lymphocytes and airway macrophages in coculture. In conclusion, our results indicate that endotoxin exposure causes the release of IL-17 from T lymphocytes and that this cytokine release requires the presence of macrophages. Once released, endogenous IL-17 acts in part by inducing local release of neutrophil-mobilizing cytokines such as IL-6 and MIP-2, from nonlymphocyte, nonmacrophage cells, and this contributes to recruitment of neutrophils in the airways. These IL-17-related mechanisms constitute potential targets for pharmacotherapy against exaggerated neutrophil recruitment in airway disease.     

IL-17, produced by lymphocytes and neutrophils,is necessary for lipopolysaccharide-induced airway neutrophilia: IL-15 as a possible trigger

IL-17 is a cytokine implicated in the regulation of inflammation. We investigated the role of this cytokine in neutrophil recruitment using a model of LPS-induced lung inflammation in mice. In the bronchoalveolar lavage, LPS induced a first influx of neutrophils peaking at day 1, followed by a second wave, peaking at day 2. IL-17 levels were increased during the late phase neutrophilia (day 2), and this was concomitant with an increased number of T cells and macrophages, together with an increase of KC and macrophage-inflammatory protein-2 levels in the lung tissue. Intranasal treatment with a neutralizing murine anti-IL-17 Ab inhibited the late phase neutrophilia. In the bronchoalveolar lavage cells, IL-17 mRNA was detected at days 1, 2, and 3 postchallenge, with a strong expression at day 2. This expression was associated with CD4(+) and CD8(+) cells, but also with neutrophils. When challenged with LPS, despite the absence of T cells, SCID mice also developed a neutrophilic response associated with IL-17 production. In BALB/c mice, IL-15 mRNA, associated mainly with neutrophils, was evidenced 1 day after LPS challenge. In vitro, IL-15 was able to induce IL-17 release from purified spleen CD4(+) cells, but not spleen CD8(+) or airway neutrophils. We have shown that IL-17, produced mainly by CD4(+) cells, but also by neutrophils, plays a role in the mobilization of lung neutrophils following bacterial challenge. In addition, our results suggest that IL-15 could represent a physiological trigger that leads to IL-17 production following bacterial infection.     

IL-17A synergizes with IFN-γ to upregulate iNOS and NO production and inhibit chlamydial growth

IFN-γ-mediated inducible nitric oxide synthase (iNOS) expression is critical for controlling chlamydial infection through microbicidal nitric oxide (NO) production. Interleukin-17A (IL-17A), as a new proinflammatory cytokine, has been shown to play a protective role in host defense against Chlamydia muridarum (Cm) infection. To define the related mechanism, we investigated, in the present study, the effect of IL-17A on IFN-γ induced iNOS expression and NO production during Cm infection in vitro and in vivo. Our data showed that IL-17A significantly enhanced IFN-γ-induced iNOS expression and NO production and inhibited Cm growth in Cm-infected murine lung epithelial (TC-1) cells. The synergistic effect of IL-17A and IFN-γ on Chlamydia clearance from TC-1 cells correlated with iNOS induction. Since one of the main antimicrobial mechanisms of activated macrophages is the release of NO, we also examined the inhibitory effect of IL-17A and IFN-γ on Cm growth in peritoneal macrophages. IL-17A (10 ng/ml) synergizes with IFN-γ (200 U/ml) in macrophages to inhibit Cm growth. This effect was largely reversed by aminoguanidine (AG), an iNOS inhibitor. Finally, neutralization of IL-17A in Cm infected mice resulted in reduced iNOS expression in the lung and higher Cm growth. Taken together, the results indicate that IL-17A and IFN-γ play a synergistic role in inhibiting chlamydial lung infection, at least partially through enhancing iNOS expression and NO production in epithelial cells and macrophages.     

Regulation of chemokine expression by IL-10 in lung inflammation

We have been interested in understanding the mechanisms regulating the inflammatory process underlying acute lung injury. The current studies have employed a model of acute lung inflammation in mice triggered by bacterial lipopolysaccharide. The development of this injury was associated with increased expression of the chemokines, MIP-1alpha and MIP-2, that coordinate recruitment of neutrophils to the lung. IL-10 is a potent, endogenous anti-inflammatory molecule that has been shown to decrease lung inflammation partly on the basis of TNF-alpha and IL-1beta inhibition. In these studies we tested the hypothesis that endogenous IL-10 modulates chemokine expression using the IL-10 knock-out mouse, and then explored the molecular mechanisms by which IL-10 might do so. The results demonstrate that significant elevations in both chemokines were observed in the absence of IL-10 and that these findings were associated with significant increases in lung neutrophil accumulation. In vitro studies defined two, gene-specific, mechanisms by which IL-10 regulated chemokine expression: mRNA destabilization and NF-kappaB inhibition. These results suggested that IL-10 is an important, endogenous regulator of chemokine expression in acute lung inflammation.     

Involvement of vitronectin in lipopolysaccaride-induced acute lung injury

Vitronectin is present in large concentrations in serum and participates in regulation of humoral responses, including coagulation, fibrinolysis, and complement activation. Because alterations in coagulation and fibrinolysis are common in acute lung injury, we examined the role of vitronectin in LPS-induced pulmonary inflammation. Vitronectin concentrations were significantly increased in the lungs after LPS administration. Neutrophil numbers and proinflammatory cytokine levels, including IL-1beta, MIP-2, KC, and IL-6, were significantly reduced in bronchoalveolar lavage fluid from vitronectin-deficient (vitronectin(-/-)) mice, as compared with vitronectin(+/+) mice, after LPS exposure. Similarly, LPS induced increases in lung edema, myeloperoxidase-concentrations, and pulmonary proinflammatory cytokine concentrations were significantly lower in vitronectin(-/-) mice. Vitronectin(-/-) neutrophils demonstrated decreased KC-induced chemotaxis as compared with neutrophils from vitronectin(+/+) mice, and incubation of vitronectin(+/+) neutrophils with vitronectin was associated with increased chemotaxis. Vitronectin(-/-) neutrophils consistently produced more TNF-alpha, MIP-2, and IL-1beta after LPS exposure than did vitronectin(+/+) neutrophils and also showed greater degradation of IkappaB-alpha and increased LPS-induced nuclear accumulation of NF-kappaB compared with vitronectin(+/+) neutrophils. These findings provide a novel vitronectin-dependent mechanism contributing to the development of acute lung injury.     

Intranasal Administration of Recombinant Mycobacterium smegmatis Inducing IL-17A Autoantibody Attenuates Airway Inflammation in a Murine Model of Allergic Asthma

Asthma is a chronic inflammatory disorder, previous studies have shown that IL-17A contributes to the development of asthma, and there is a positive correlation between the level of IL-17A and the severity of disease. Here, we constructed recombinant Mycobacterium smegmatis expressing fusion protein Ag85A-IL-17A (rMS-Ag85a-IL-17a) and evaluated whether it could attenuate allergic airway inflammation, and further investigated the underlying mechanism. In this work, the murine model of asthma was established with ovalbumin, and mice were intranasally vaccinated with rMS-Ag85a-IL-17a. Autoantibody of IL-17A in sera was detected, and the airway inflammatory cells infiltration, the local cytokines and chemokines production and the histopathological changes of lung tissue were investigated. We found that the administration of rMS-Ag85a-IL-17a induced the autoantibody of IL-17A in sera. The vaccination of rMS-Ag85a-IL-17a remarkably reduced the infiltration of inflammatory cells and the secretion of mucus in lung tissue and significantly decreased the numbers of the total cells, eosinophils and neutrophils in BALF. Th1 cells count in spleen, Th1 cytokine levels in BALF and supernatant of splenocytes and mediastinal lymph nodes, and T-bet mRNA in lung tissue were significantly increased with rMS-Ag85a-IL-17a administration. Meanwhile, rMS-Ag85a-IL-17a vaccination markedly decreased Th2 cells count, Th2 cytokine and Th17 cytokine levels in BALF and supernatant of splenocytes and mediastinal lymph nodes, and chemokines mRNA expression in lung tissue. These data confirmed that recombinant Mycobacterium smegmatis in vivo could induce autoantibody of IL-17A, which attenuated asthmatic airway inflammation.     

Chlamydia trachomatis mouse pneumonitis lung infection in IL-18 and IL-12 knockout mice: IL-12 is dominant over IL-18 for protective immunity

BACKGROUND: Interferon (IFN)-gamma is a key to protective immunity against a variety of intracellular bacterial infections, including Chlamydia trachomatis. Interleukin (IL)-18, a recently identified Th1 cytokine, together with IL-12 is a strong stimulator for IFN-gamma production. We investigated the relative roles of IL-18 and IL- 12 in protective immunity to C. trachomatis mouse pneumonitis (MoPn) infection using gene knockout (KO) and wild-type (WT) mice. MATERIALS AND METHODS: Mice were intranasally infected with C. trachomatis MoPn and protective immunity was assessed among groups of mice by daily body weight changes, lung growth of MoPn, and histopathological appearances at day 10 postinfection. The corresponding immune responses for each group of mice at the same postinfection time point were evaluated by measuring antigen-specific antibody isotype responses and cytokine profiles. RESULTS: Our results showed that IL-18 deficiency had little or no influence on clearance of MoPn from the lung, although KO mice exhibited slightly more severe inflammatory reactions in lung tissues, as well as reduced systemic and local IFN-gamma production, compared with WT mice. Results with IL-18 KO mice were in sharp contrast to those observed with IL-12 KO mice that showed substantially reduced clearance of MoPn from the lungs, substantial reductions of antigen-specific systemic and lung IFN-gamma production, decreased ratio of MoPn-specific immunoglobulin G (IgG)2a/IgG1, and severe pathological changes in the lung with extensive polymorphonuclear, instead of mononuclear, cell infiltration. Exogenous IL-12 or IL-18 was able to increase IFN-gamma production in IL-18 KO mice; whereas, only exogenous IL-12, but not IL-18, enhanced IFN-gamma production in IL-12 KO mice. Caspase-1 is the key protease for activation of IL-18 precursor into the bioactive form, and caspase-1 KO mice also displayed similar bacterial clearance and body weight loss to that in WT mice at early stages of MoPn infection. This further confirmed that IL-18 was not essential for host defense against chlamydia infection. CONCLUSIONS: These results suggest that IL-12, rather than IL-18, plays the dominant role in the development of protective immunity against chlamydia lung infection, although both cytokines are involved in the in vivo regulation of IFN-gamma production.     

IL-17RA signaling amplifies antibody-induced arthritis

Objective

To investigate the role of IL-17RA signaling in the effector phase of inflammatory arthritis using the K/BxN serum-transfer model.

Methods

Wild-type and Il17ra^−/− mice were injected with serum isolated from arthritic K/BxN mice and their clinical score was recorded daily. Mice were also harvested on days 12 and 21 and ankles were analyzed for cytokine and chemokine mRNA expression by qPCR on day 12 and for bone and cartilage erosions by histology on day 21, respectively. The induction of cytokine and chemokine expression levels by IL-17A in synovial-like fibroblasts was also analyzed using qPCR.

Results

Il17ra^−/− mice were partially protected from clinical signs of arthritis and had markedly fewer cartilage and bone erosions. The expression of several pro-inflammatory mediators, including the chemokines KC/CXCL1, MIP-2/CXCL2, LIX/CXCL5 MIP-1γ/CCL9, MCP-3/CCL7, MIP-3α/CCL20, the cytokines IL-1β, IL-6, RANKL and the matrix metalloproteinases MMP2, MMP3, and MMP13 were decreased in the ankles of Il17ra^−/− mice compared to wild-type mice. Many of these proinflammatory genes attenuated in the ankles of Il17ra^−/− mice were shown to be directly induced by IL-17A in synovial fibroblasts in vitro.

Conclusions

IL-17RA signaling plays a role as an amplifier of the effector phase of inflammatory arthritis. This effect is likely mediated by direct activation of synovial fibroblasts by IL-17RA to produce multiple inflammatory mediators, including chemokines active on neutrophils. Therefore, interrupting IL-17RA signaling maybe a promising pharmacological target for the treatment of inflammatory arthritis.     

Potentiation of endogenous fibrinolysis and rescue from lung ischemia/reperfusion injury in interleukin (IL)-10-reconstituted IL-10 null mice

Little is known about interactions between endogenous anti-inflammatory paradigms and microvascular thrombosis in lung ischemia/reperfusion (I/R) injury. Interleukin (IL)-10 suppresses macrophage activation and down-regulates proinflammatory cytokine production, but there are no available data to suggest a link between IL-10, thrombosis, and fibrinolysis in the setting of I/R. We hypothesized that hypoxia/ischemia triggers IL-10 production, to dampen proinflammatory cytokine and adhesion receptor cascades and to restore vascular patency by fibrinolytic potentiation. Studies were performed in a mouse lung I/R model. IL-10 mRNA levels in lung were increased 43-fold over base line by 1 h of ischemia/2 h of reperfusion, with a corresponding increase in plasma IL-10. Expression was prominently localized in bronchial epithelial cells and mononuclear phagocytes. To study the link between IL-10 and fibrinolysis in vivo, the induction of plasminogen activator inhibitor-1 (PAI-1) was evaluated. Northern analysis demonstrated exaggerated pulmonary PAI-1 expression in IL-10 (-/-) mice after I/R, with a corresponding increase in plasma PAI/tissue-type plasminogen activator activity. In vivo, IL-10 (-/-) mice showed poor postischemic lung function and survival after I/R compared with IL-10 (+/+) mice. Despite a decrease in infiltration of mononuclear phagocytes in I/R lungs of IL-10 (-/-) mice, an increased intravascular pulmonary fibrin deposition was observed by immunohistochemistry and Western blotting, along with increased IL-1 expression. Recombinant IL-10 given to IL-10 (-/-) mice normalized the PAI/tissue-type plasminogen activator ratio, reduced pulmonary vascular fibrin deposition, and rescued mice from lung injury. Since recombinant hirudin (direct thrombin inhibitor) also sufficed to rescue IL-10 (-/-) mice, these data suggest a preeminent role for microvascular thrombosis in I/R lung injury. Ischemia-driven IL-10 expression confers postischemic pulmonary protection by augmenting endogenous fibrinolytic mechanisms.