IL-6协同M-CSF体外诱导CD14~+单核细胞向M2样巨噬细胞分化

目的探讨IL-6协同M-CSF体外诱导CD14~+单核细胞向M2样巨噬细胞分化的作用机制,并检测诱导而来的M2样巨噬细胞的表型特点以及对肿瘤细胞的作用。方法体外构建IL-6诱导单核细胞分化模型,q RT-PCR检测M2表型分子(IL-10、TGF-β)在不同质量浓度IL-6刺激下的表达水平;Western blot检测STAT3信号通路的活化水平;采用细胞迁移、增殖实验验证IL-6诱导的M2样巨噬细胞对肿瘤细胞增殖、迁移的作用。结果在IL-6诱导分化的巨噬细胞中,IL-10、TGF-β表达增高呈IL-6剂量依赖性,且IL-6大量激活JAK/STAT3信号通路中的STAT3蛋白,使其磷酸化,si RNA沉默STAT3后,IL-10、TGF-β表达水平相应降低;此外,M2样巨噬细胞上清处理的实验组比对照组胃癌细胞增殖速度和迁移能力显著增强。结论 IL-6可以通过激活STAT3信号通路诱导单核细胞高表达IL-10、TGF-β,低表达IL-12的M2样巨噬细胞分化,并且M2样巨噬细胞可以促进肿瘤细胞的增殖和迁移。     

嗜酸性粒细胞通过调节巨噬细胞极化减轻脂多糖诱导的急性肺损伤

目的：观察体外嗜酸性粒细胞（Eos）对巨噬细胞极化的影响,以及体内注射嗜酸性粒细胞对脂多糖（LPS） 诱导的急性肺损伤（ALI）的作用及其相关机制。方法：原代培养小鼠巨噬细胞,在嗜酸性粒细胞存在或不存 在的情况下,将细胞暴露于脂多糖和干扰素-γ（IFN-γ）以模拟ALI,采用荧光定量PCR 检测各组巨噬细胞中促 炎及抗炎因子的表达,采用免疫印迹法检测巨噬细胞过氧化物酶增殖物激活受体γ（PPARγ）蛋白的表达;将 PPARγ siRNA（si-PPARγ）转染小鼠巨噬细胞,采用免疫印迹法检测巨噬细胞的极化情况;Balb/c 小鼠随机分为 对照组、ALI 组和ALI+Eos 组,ALI 组为通过气管内注射LPS 诱导ALI,ALI+Eos 组给予尾静脉注射嗜酸性粒细 胞,对照组尾静脉注射生理盐水。监测动物7 d 的存活情况。在LPS 注射后3 h 和24 h 处死动物,通过组织学评估 左肺损伤程度,荧光定量PCR 检测右肺促炎和抗炎细胞因子mRNA水平,采用免疫印迹法检测右肺组织PPARγ 蛋白的表达,流式细胞术检测肺泡巨噬细胞诱导型一氧化氮合酶（iNOS）和CD206 的百分比。结果：与嗜酸性 粒细胞共培养后,用LPS+IFN-γ 刺激巨噬细胞,其肿瘤坏死因子-α（TNF-α）和白细胞介素（IL）-6 mRNA水平 呈剂量依赖性下降;而IL-10 mRNA 水平和PPARγ 活性呈剂量依赖性增加。巨噬细胞与嗜酸性粒细胞共培养后, 其iNOS 表达增加和重组人精氨酸酶1（Arg1）表达减少,呈剂量依赖性逆转。转染si-PPARγ 抑制了嗜酸性粒细 胞促进巨噬细胞向M2表型分化的作用。静脉注射嗜酸性粒细胞显著提高了ALI 小鼠的存活率,且显著改善小鼠 的肺组织损伤,降低肺组织IL-6 和TNF-α mRNA水平,增加IL-10 mRNA 水平和PPARγ 活性。此外,嗜酸性粒 细胞干预增加了肺泡巨噬细胞CD206 的百分比,而iNOS 的百分比明显降低。结论：嗜酸性粒细胞通过促进巨噬 细胞M2型极化改善LPS 诱导的ALI。     

M2巨噬细胞分泌的IL-10通过JAK2/STAT3信号通路调节乳腺癌细胞的增殖、侵袭与凋亡

目的探讨M2巨噬细胞分泌的IL-10通过JAK2/STAT3信号通路对乳腺癌细胞的增殖、侵袭迁移与凋亡的影响。方法THP-1细胞诱导为M2巨噬细胞,并对标志基因(CD206、ARG1、IL-6和IFN-β)进行检测;ELISA检测细胞上清液中IL-10的表达;细胞集落形成实验和EdU实验检测细胞的增殖能力;细胞侵袭与迁移实验检测细胞的侵袭与迁移能力;细胞流式术检测细胞的凋亡率和细胞周期;Western blot检测JAK2/STAT3通路相关蛋白的表达。结果 M2巨噬细胞在体外被成功诱导并进行了鉴定;与未经处理的THP-1细胞相比,M2巨噬细胞上清液中IL-10的明显增加;M2巨噬细胞分泌的IL-10可促进MDA-MB-231细胞的增殖、侵袭与迁移(均P0.001),抑制MDA-MB-231细胞的凋亡(P0.001);抑制IL-10的表达可抑制MDA-MB-231细胞的增殖、侵袭与迁移(均P0.05),促进MDA-MB-231细胞的凋亡(P0.05);IL-10可激活JAK2/STAT3信号通路,而抑制IL-10的表达可抑制JAK2/STAT3信号通路的激活。结论 M2巨噬细胞分泌的IL-10可促进乳腺癌细胞的增殖、侵袭与迁移,抑制乳腺癌细胞凋亡,其机制可能与JAK2/STAT3信号通路激活有关。     

IL-37通过调控M-CSF和抑制IL-6-JAK2/STAT3信号通路抑制骨质疏松的机制研究

目的:探讨IL-37 在抑制骨质疏松过程中的作用机制。方法:选取本院2013 年1 月至2015 年12 月收治的97例骨质疏松患者及在本院行骨折手术的81 例无骨质疏松患者(对照组)为研究对象,检测两组血清中IL-37 及IL-6 的水平。构建IL-37 转基因小鼠,将C57BL/6J 小鼠、IL-37 转基因小鼠分别设置假手术(Sham)组,手术组(卵巢切除术,OVX 组)。8 周后,取小鼠血清,检测血清中雌激素水平、碱性磷酸酶水平(ALP)、血钙和血磷水平;同时取小鼠的双侧股骨、脊柱,病理切片分析股骨组织形态结构,骨密度仪检测脊柱骨密度变化。分离培养各组小鼠骨髓基质细胞(Bone marrow stromal cells,BMSCs), 检测BMSCs 的体外增殖能力,M-CSF 及IL-6 的表达及STAT3 的激活。IL-37 转染小鼠成骨细胞MC3T3-E1,转染后72 h,ELISA 检测上清中M-CSF 及IL-6,流式细胞术检测MC3T3-E1 细胞的凋亡,Western blot 检测STAT3 的激活。结果:骨质疏松患者血清中IL-37 水平显著低于对照组(P<0.05),而IL-6 则显著高于对照组;C57BL/6J 小鼠、IL-37 转基因小鼠OVX 组血清中雌激素、血钙和血磷显著低于假手术组,而ALP 水平显著高于假手术组(P<0.05),但IL-37 转基因小鼠OVX 组血钙和血磷则显著高于C57BL/6J 小鼠OVX 组(P<0.05)。股骨病理切片及脊柱骨密度结果显示,C57BL/6J 小鼠、IL-37 转基因小鼠OVX组均出现组织形态结构的破坏和骨密度下降,但IL-37 转基因小鼠明显优于C57BL/6J 小鼠(P<0.05)。IL-37 转基因小鼠OVX 组BMSCs 增殖能力显著高于C57BL/6J 小鼠OVX 组,而STAT3 的激活和M-CSF 的表达则显著低于C57BL/6J 小鼠OVX组(P<0.05)。MC3T3-E1 细胞转染IL-37 后能明显抑制M-CSF 及IL-6 的表达,而STAT3 的激活也明显被抑制,流式细胞检测显示转染IL-37 后能显著抑制MC3T3-E1 细胞的凋亡。结论:骨质疏松患者血清IL-37 水平显著降低,IL-37 可能是通过调控M-CSF 及IL-6-JAK2/ STAT3 信号通路促进BMSCs 增殖和抑制成骨细胞的凋亡从而抑制骨质疏松的进展。     

IL-31在哮喘小鼠中的动态表达及对肺泡上皮细胞表达CCL11和CCL22的影响

目的通过观察IL-31在OVA诱导小鼠哮喘模型中的动态表达及IL-31对肺泡上皮细胞表达趋化因子CCL11和CCL22的影响,探讨IL-31在哮喘气道炎症中的作用。方法常规OVA法建立小鼠哮喘模型,于末次激发后取肺组织HE染色及AB-PAS染色,收集支气管肺泡灌洗液(BALF)进行白细胞和嗜酸性粒细胞(EOS)计数,ELISA法检测血浆中IgE、IL-4、IFN-γ、IL-31水平,荧光定量PCR检测肺组织IL-31R mRNA表达水平,同时体外培养小鼠肺泡上皮细胞,用IL-31处理24 h后检测CCL11和CCL22 mRNA的表达水平。结果成功构建哮喘小鼠模型,哮喘小鼠BALF中白细胞总数、嗜酸性粒细胞百分比和血浆中IgE水平明显增多。哮喘小鼠肺组织病理切片见中性粒细胞、EOS浸润。哮喘小鼠血浆中Th2类细胞因子IL-4水平明显高于对照组,Th1类细胞因子IFN-γ明显低于对照组。哮喘小鼠血浆IL-31水平和肺组织IL-31R mRNA表达水平明显增高,第2周至第8周虽略有降低但仍明显高于对照组。IL-31作用小鼠肺泡上皮细胞24h后,趋化因子CCL11、CCL22mRNA表达增高。结论IL-31通过刺激趋化因子表达募集炎性细胞,促进气道炎症的发生发展。     

长链非编码RNA肺腺癌转移相关转录本1对巨噬细胞极化影响的研究

目的研究长链非编码RNA(lnc RNA)肺腺癌转移相关转录本1(MALAT1)对巨噬细胞极化的影响。方法采用佛波酯诱导人THP-1细胞分化为成熟的巨噬细胞,IFN-γ/LPS刺激诱导M1型极化,IL-4刺激诱导M2型极化,采用实时荧光定量PCR(RT-q PCR)检测各组细胞中MALAT1的表达。si RNA干扰巨噬细胞MALAT1表达,加入IL-4继续诱导M2型极化,RT-q PCR检测极化相关基因表达;酶联免疫吸附试验(ELISA)检测TNF-α、IL-12、CCL22、IL-10的水平。si RNA干扰M2型巨噬细胞MALAT1表达,研究其对M2极化表型的影响。结果 MALAT1在M2型巨噬细胞中表达显著升高,M1向M2极化改变MALAT1表达升高,而M2向M1极化改变MALAT1表达降低。MALAT1干扰后,IL-4诱导的M2型巨噬细胞TNF-α、IL-12分泌升高,CCL22、IL-10分泌降低;CXCL10、CXCL11、HLA-DR表达升高,CCL17、CCL18、CD163表达降低。M2型巨噬细胞MALAT1干扰后,TNF-α、IL-12分泌升高,CCL22、IL-10分泌降低。结论 MALAT1参与调控巨噬细胞极化,干扰MALAT1表达有效抑制M2型巨噬细胞极化,促进M1型巨噬细胞极化。     

过敏性紫癜患者血液嗜酸性粒细胞IL-18和IL-18受体表达升高

目的探讨过敏性紫癜患者血液嗜酸性粒细胞中IL-18、IL-18受体(IL-18 receptor,IL-18R)和IL-18结合蛋白(IL-18 binding protein,IL-18BP)的表达,并分析其相关性。方法收集过敏性紫癜患者的外周静脉血,用蒿草花粉、尘螨和梧桐花粉过敏原粗提液刺激,流式细胞术检测血液中嗜酸性粒细胞IL-18、IL-18R、IL-18BP的表达,并用SPSS分析其相关性。结果过敏性紫癜患者IL-18+和IL-18R+细胞的比例分别升高了1.83倍(P=0.048)和1.12倍(P=0.025),而IL-18BP+嗜酸性粒细胞的平均荧光强度(mean fluorescence intensity,MFI)降低了30%(P=0.005)。过敏性紫癜患者IL-18+和IL-18R+嗜酸性粒细胞的比例呈中度相关(相关系数r=0.559,P=0.000)。此外,梧桐花粉过敏原粗提液诱导过敏性紫癜患者血液IL-18+嗜酸性粒细胞的MFI升高了37%(P=0.027)。结论过敏性紫癜患者嗜酸性粒细胞中IL-18和IL-18R表达上调而IL-18BP表达下调,提示嗜酸性粒细胞表达的IL-18、IL-18BP和IL-18R可能在过敏性紫癜中起重要作用。     

气道内应用IL-12抑制抗原诱导的过敏性气道反应

目的探讨气道内应用白细胞介素 12 (IL- 12 )对抗原诱导的过敏性反应的影响。方法 C5 7BL/ 6小鼠经 OVA免疫建立哮喘模型 ,在主动免疫及抗原激发阶段气道内应用 IL- 12 ,观察肺泡灌洗液细胞成份、肺部淋巴细胞产生细胞因子、外周血 Ig E水平变化。结果 1致敏阶段应用 IL- 12可明显抑制嗜酸性粒细胞浸润、肺淋巴细胞对 IL- 5的分泌以及血浆总 Ig E和抗原特异性 Ig E的水平 ;2激发阶段应用 IL- 12可明显抑制嗜酸性粒细胞的浸润 ,抑制肺淋巴细胞产生 IL- 4、IL- 5 ,增加其产生 IFN- γ,但对抗原特异性 Ig E无明显影响 ;3如致敏和激发阶段均应用 IL- 12 ,则明显抑制肺淋巴细胞产生 IL- 4、IL- 5 ,增强 IFN- γ产生 ,抑制气道内嗜酸性粒细胞的浸润及血浆 Ig E的升高。结论气道应用 IL- 12对抗原诱导的气道过敏性炎症有明显调节作用 ,且与应用时机有关 ,为 IL- 12治疗哮喘提供依据     

IL-4和IL-10调节嗜碱性粒细胞CXCR4表达及功能

目的 研究IL-4和IL-10对人嗜碱性粒细胞上CXC趋化性细胞因子受体-4（CXCR4）表达和配体SDF-1α（Chemokine stromal cell-derived factor-1 alpha)功能的调节。方法 嗜碱性粒细胞的纯化技术,流式细胞术,实时定量逆转录PCR（RT-PCR）,胞内游离Ca^2 的变化。趋 化性技术和组胺释放等方法进行测定与分析。结果 CXCR4大量表达在人外周血静息嗜碱性粒细胞上,IL-4可显著上调CXCR4蛋白和mRNA的表达,而IL-10则明显下调其表达,SDF-1α可通过CXCR4诱导嗜碱性粒细胞中游离Ca^2 增加,激活嗜碱性粒细胞使之产生趋化性游走并释放组胺,此活性可被抗CXCR4单抗所阻断。结论 IL-4和IL-10是CXCR4表达和功能重要的调节细胞因子,CXCR4-SDF-1α复合物相互作用对嗜碱性粒细胞的聚集和活化起着重要作用。     

抗TNF-α和IL-1βIgY抗体治疗豚鼠过敏性鼻炎机理研究

目的：探讨抗TNF-α和IL-1βIgY治疗豚鼠过敏性鼻炎的作用及机制。方法：随机分正常对照组（ C组,17只）、模型组（ M组,27只）、0.1％抗TNF-α和IL-1βIgY治疗组（ Z1组,21只）、丙酸氟替卡松治疗组（ Z2组,21只）;应用卵清白蛋白建立豚鼠过敏性鼻炎模型;分别在治疗后2、4、8 h进行鼻灌洗和支气管肺灌洗,收集鼻灌洗液（ NLF）和支气管肺泡灌洗液（BALF）,观察炎症细胞;HE染色观察鼻黏膜和肺组织病理改变;免疫组化分析其炎症因子原位表达情况。结果：M组鼻黏膜可见大量嗜酸性粒细胞浸润和炎症改变,肺间质水肿、肺间隔和支气管平滑肌增厚,嗜酸性粒细胞浸润, Z1组和Z2组炎症病理改变明显减轻。 NLF和BALF中嗜酸性粒细胞、淋巴细胞、中性粒细胞在Z1和Z2组显著少于M组（ P＜0.05）。 Z1组鼻黏膜IL-1β和肺组织IL-1β和TNF-α从2 h开始,而IL-5和IL-33从4 h开始表达显著低于M组（ P＜0.05）。结论：抗TNF-α和IL-1βIgY滴鼻治疗显著减轻了豚鼠过敏性鼻炎伴过敏性支气管哮喘的炎症病理反应,抑制了嗜酸性粒细胞浸润及炎症细胞因子表达。     

Enhanced innate immune responsiveness to pulmonary Cryptococcus neoformans infection is associated with resistance to progressive infection

Genetically regulated mechanisms of host defense against Cryptococcus neoformans infection are not well understood. In this study, pulmonary infection with the moderately virulent C. neoformans strain 24067 was used to compare the host resistance phenotype of C57BL/6J with that of inbred mouse strain SJL/J. At 7 days or later after infection, C57BL/6J mice exhibited a significantly greater fungal burden in the lungs than SJL/J mice. Characterization of the pulmonary innate immune response at 3 h after cryptococcal infection revealed that resistant SJL/J mice exhibited significantly higher neutrophilia, with elevated levels of inflammatory cytokine tumor necrosis factor alpha (TNF-α) and keratinocyte-derived chemokine (KC)/CXCL1 in the airways, as well as increased whole-lung mRNA expression of chemokines KC/CXCL1, MIP-1α/CCL3, MIP-1β/CCL4, MIP-2/CXCL2, and MCP-1/CCL2 and cytokines interleukin 1β (IL-1β) and IL-1Ra. At 7 and 14 days after infection, SJL/J mice maintained significantly higher levels of TNF-α and KC/CXCL1 in the airways and exhibited a Th1 response characterized by elevated levels of lung gamma interferon (IFN-γ) and IL-12/IL-23p40, while C57BL/6J mice exhibited Th2 immunity as defined by eosinophilia and IL-4 production. Alveolar and resident peritoneal macrophages from SJL/J mice also secreted significantly greater amounts of TNF-α and KC/CXCL1 following in vitro stimulation with C. neoformans. Intracellular signaling analysis demonstrated that TNF-α and KC/CXCL1 production was regulated by NF-κB and phosphatidylinositol 3 kinase in both strains; however, SJL/J macrophages exhibited heightened and prolonged activation in response to C. neoformans infection compared to that of C57BL/6J. Taken together, these data demonstrate that an enhanced innate immune response against pulmonary C. neoformans infection in SJL/J mice is associated with natural resistance to progressive infection.     

Modulation of the expression of M150 on macrophages by Th1/Th2 cytokines and co-stimulatory molecules CD40, B7-1, B7-2 and ICAM-1

M150 is an 150-kDa protein associated with the surface of macrophages and is responsible chiefly for the activation of Th1 cells. It is a unique subset of the lysosome-associated membrane protein-1 glycoprotein and its co-stimulatory activity depends on its post-translational modification, which has a distinct glycosylation pattern restricted to macrophages. In the present study, we have observed that M150 is expressed constitutively on peritoneal but not splenic macrophages isolated from mice of different genetic backgrounds: Balb/c, C57BL/6 and C3He. However, M150 was expressed not only on peritoneal but also on splenic macrophages of non-obese diabetic (NOD) mice. Expression on splenic macrophages was induced by culture with lipopolysaccharide (LPS). Expression could also be significantly up-regulated by interferon (IFN)-gamma and granulocyte-macrophage colony stimulating factor (GM-CSF) but was inhibited by interleukin (IL)-10; IL-4 exhibited no effect. Further, cross-linking of B7-2, CD40, ICAM-1 but not B7-1 enhanced the level of M150 significantly. IFN-gamma and GM-CSF acted synergistically with CD40. The significance of these findings is that cytokines IFN-gamma, GM-CSF and IL-10 and the co-stimulatory molecules B7-2, CD40 and ICAM-1 can regulate the expression of M150 on macrophages.     

IL-3 is an important differentiation factor for the development of prostaglandin E2-producing macrophages between C57BL/6 and BALB/c mice

We have previously reported that peritoneal and splenic macrophages from Th2-dominant BALB/c mice produced higher amounts of prostaglandin (PG) E2 than cells from C57BL/6 mice. In this study, we investigated how macrophages from BALB/c mice acquire the ability of enhanced PGE2 production, using bone marrow-derived macrophages differentiated by M-CSF, GM-CSF or IL-3. There is no strain difference in PGE2 production by GM-CSF- and M-CSF-differentiated macrophages; however, IL-3-differentiated macrophages from BALB/c mice produced higher amounts of PGE2 and lower amounts of type I cytokines than cells from C57BL/6 mice. IL-3-differentiated macrophages from BALB/c mice expressed larger amounts of mRNA of membrane-bound (microsomal) PGE synthase-1 (mPGES-1). The amounts of PGE2 produced by macrophages were significantly reduced in mPGES-1-deficient mice, and these mice displayed enhanced Th1 responses after Propionibacterium acnes treatment compared with wild-type mice. Microarray analysis revealed 63 genes that are differentially expressed more than fivefold in macrophages between C57BL/6 and BALB/c mice. These results indicate that mPGES-1-mediated PGE2 produced by macrophages regulates immune responses, and IL-3 is an important factor for the differentiation of macrophages that produce higher amounts of PGE2 through mPGES-1 activity in BALB/c mice.     

NOD mice have a severely impaired ability to recruit leukocytes into sites of inflammation

The accumulation of macrophages (M Phi) and dendritic cells (DC) in the pancreas plays a crucial role in the pathogenesis of autoimmune diabetes. We studied the recruitment of monocytes, M Phi and DC to sites of inflammation, i.e. the peritoneal cavity and a subcutaneously elicited air pouch in the NOD mouse model of autoimmune diabetes. The leukocyte recruitment was studied from 1 to 7 days after injection of thioglycollate (peritoneum), C5a (peritoneum, air pouch), CCL2 and CCL3 (air pouch). C57BL/6 and BALB/c mice served as controls. Morphological and flow cytometric analysis of the recruited cells was performed, IL-1 beta, TNF-alpha, IL-6, IL-12 and IL-10 in exudates measured, and in vitro CCL2-chemotaxis of exudate M Phi (Boyden chamber) determined. NOD mice were strongly impaired in the recruitment of M Phi, DC, monocytes, and granulocytes. Chemokine-injected air pouches of NOD mice showed an increased IL-10 and a decreased IL-1 beta level, while the other cytokines were normally or very lowly expressed. In addition, NOD exudate M Phi displayed an impaired in vitro CCL2-induced migration. Our data show that NOD mice have an impaired ability to recruit leukocytes into sites of inflammation elicited in the peritoneum and the air pouch. A raised IL-10/IL-1 beta ratio at these sites and a deficient migratory capacity of NOD monocytes are important determinants in this impairment.     

Sephadex induces eosinophil migration to the rat and mouse peritoneal cavity: involvement of mast cells,LTB4, TNF-α, IL-8 and PAF

OBJECTIVE AND DESIGN: In this study we investigated the chemotactic mediators involved in the Sephadex-induced eosinophil migration into the peritoneal cavities of rats and mice, and which resident peritoneal cells release these mediators. MATERIALS AND METHODS: Sephadex suspension was injected into the peritoneal cavities of rats or mice which were pretreated, or not, with specific drugs that inhibit synthesis or production of the inflammatory mediators and eosinophil chemotactic activities were observed. To investigate the role of resident peritoneal cells as a source of these chemotactic factors, the macrophage population was enhanced or the mast cell population was depleted. The resident cells were also stimulated, in vitro, with Sephadex and the chemotactic activity of the supernatants was determined. RESULTS: Sephadex induced dose and time dependent eosinophil migration in rats and mouse, which were inhibited by dexamethasone and MK 886. BN 52021 only affected the eosinophil migration into the mouse peritoneal cavity. An increase in the macrophage population did not alter the eosinophil migration induced by Sephadex in rat or mouse. However, mast cell population depletion reduced eosinophil migration in rats, but did not alter the migration in mice. Sephadex-stimulated rat mast cells released an eosinophil chemotactic factor whose release was inhibited by dexamethasone and MK 886. Anti-TNF-alpha and anti-IL-8 Abs inhibited the chemotactic activity of the mast cell supernatant. CONCLUSION: Sephadex-induced eosinophil migration into the rat peritoneal cavity is dependent on mast cells, which release LTB4, TNF-alpha and CINC-1. Conversely, Sephadex-induced eosinophil migration into the mouse peritoneal cavity is mediated by PAF and LTB4, which are not released from resident macrophages or mast cells.     

Macrophage activation during Plasmodium chabaudi AS infection in resistant C57BL/6 and susceptible A/J mice.

Macrophage activation was examined in resistant C57BL/6 and susceptible A/J mice during the course of blood-stage infection with Plasmodium chabaudi AS. Three parameters of macrophage activation (lipopolysaccharide [LPS]- and malaria antigen-induced tumor necrosis factor [TNF] production in vitro, phorbol myristate acetate [PMA]-induced production of oxygen metabolites in vitro, and Ia antigen expression) were assessed during infection in populations of peritoneal and splenic macrophages recovered from infected mice of the two strains. The peak level of LPS-induced TNF production in vitro by splenic macrophages from both infected C57BL/6 and infected A/J mice occurred on day 7, which was 3 days before the peak of parasitemia. Although the kinetics of TNF production in vitro in response to either LPS, soluble malaria antigen, or intact parasitized erythrocytes varied in some of the other macrophage populations during infection, there was no significant difference in the peak level of production. Peritoneal and splenic macrophages from infected C57BL/6 mice exhibited significantly increased PMA-induced production of H2O2 in vitro on day 7. Peritoneal macrophages from infected A/J mice also exhibited significant PMA-induced H2O2 production on day 7, while production by splenic macrophages from these hosts was not increased in comparison with production by cells from normal animals. Only peritoneal macrophages from infected C57BL/6 mice produced significantly increased levels of O2-, and this occurred on day 7 postinfection. Ia antigen expression by both peritoneal and splenic macrophages from resistant C57BL/6 and susceptible A/J mice was significantly increased during P. chabaudi AS infection. However, the percentage of Ia+ peritoneal macrophages on days 8 and 10 postinfection and Ia+ splenic macrophages on day 3 postinfection was significantly higher in C57BL/6 than in A/J mice. Thus, these results demonstrate that macrophages from P. chabaudi AS-infected A/J mice exhibit defects in oxygen metabolism and Ia antigen expression which may contribute to the susceptibility of these hosts to this intraerythrocytic parasite. The cause-and-effect relationship between these defects and the susceptibility of A/J mice to P. chabaudi AS is unknown.     

Evidence of a role for CD200 in regulation of immune rejection of leukaemic tumour cells in C57BL/6 mice.

Increased expression of the molecule CD200 in mice receiving renal allografts is associated with immunosuppression leading to increased graft survival, and altered cytokine production in lymphocytes harvested from the transplanted animals. Preferential production of IL-4, IL-10 and TGFbeta occurs on donor-specific restimulation in vitro, with decreased production of IL-2, IFNgamma and TNFalpha. These effects are enhanced by simultaneous infusion of CD200 immunoadhesin (CD200Fc) and donor CD200 receptor (CD200r) bearing macrophages to transplanted mice. C57BL/6 mice do not normally resist growth of EL4 or C1498 leukaemia tumour cells. Following transplantation of cyclophosphamide-treated C57BL/6 with T-depleted C3H bone marrow cells, or for the EL4 tumour, immunization of C57BL/6 mice with tumour cells transfected with a vector encoding the co-stimulatory molecule CD80 (EL4-CD80), mice resist growth of tumour challenge. Immunization of C57BL/6 mice with EL4 cells overexpressing CD86 (EL4-CD86) is ineffective. Protection from tumour growth in either model is suppressed by infusion of CD200Fc, an effect enhanced by co-infusion of CD200r+ macrophages. CD200Fc acts on both CD4+ and CD8+ cells to produce this suppression. These data are consistent with the hypothesis that immunosuppression following CD200-CD200r interaction can regulate a functionally important tumour growth inhibition response in mice.     

Theophylline inhibits TNF-alpha-induced CD4 expression on human eosinophils and CD4+ eosinophil migration

BACKGROUND: Increasing evidence regarding asthma suggests that CD4+ cells are preferentially recruited to sites of bronchial inflammation. Interleukin (IL)-16 has been reported as playing an important role in the accumulation of CD4+ cells. We have shown that the CD4 molecule is expressed on normal human eosinophils by tumor necrosis factor (TNF)-alpha stimulation. METHODS: We evaluated the effects of theophylline, KF19514 [a selective phosphodiesterase (PDE) IV inhibitor] and dexamethasone on CD4 expression on eosinophils and eosinophil migration in response to IL-16, a natural soluble ligand of the CD4 molecule. RESULTS: The maximum eosinophil migration was observed when eosinophils were cultured with TNF-alpha at 10 ng/ml for 18 h and the concentration of IL-16 was 10 pg/ml. CD4+ eosinophil migration in response to IL-16 was mostly, if not fully, chemokinetic and this migration was significantly inhibited by Fab of anti-CD4 monoclonal antibody. Theophylline (10(-4)-10(-3) M), KF19514 (10(-7)-10(-6) M) and dexamethasone (10(-8)- 10(-6) M) significantly inhibited CD4 expression on eosinophils induced by TNF-alpha. Theophylline (10(-3) M) and KF19514 (10(-6) M) inhibited CD4+ eosinophil migratory responses induced by IL-16, but 10(-6) M dexamethasone did not. Theophylline and KF19514 augmented the intracellular adenosine-3',5'-cyclic monophosphate (cAMP) concentration in eosinophils, suggesting modulation by cAMP of CD4 expression and eosinophil migration. CONCLUSIONS: These data suggest that TNF-alpha-induced CD4+ eosinophils may contribute to eosinophil migratory responses induced by IL-16. Theophylline and selective PDE IV inhibitor may prevent airway inflammation by downregulating CD4 expression on eosinophils and inhibiting eosinophil migration through CD4 and IL-16 interaction.     

Dynamic changes of peritoneal macrophages and subpopulations during ulcerative colitis to metastasis of colorectal carcinoma in a mouse model

Objective and design

Patients with ulcerative colitis have increased risk of colorectal carcinoma, but little is known about how peritoneal macrophages are involved in ulcerative colitis-associated carcinogenesis. We investigated the alteration of peritoneal macrophages and M1/M2 subpopulations during ulcerative colitis-associated carcinogenesis.

Materials and methods

Expression and functional changes in peritoneal macrophages and M1/M2 subpopulations were investigated by histopathology, flow cytometry, immunofluorescence, cytokines expression by ELISA and QRT-PCR in an azoxymethane (AOM)- and dextran sodium sulfate (DSS)-induced chemical colitis-associated carcinoma mouse model using male Crj:CD-1 (ICR) mice.

Results

Striking evidence observed in histopathology, flow cytometry, cytokine detection, and gene expression analysis all revealed that inflammation-associated cytokines (IL-1β, IL-10, IL-12, IL-6, TNF-α) and migration/invasion-associated factors (G-CSF, GM-CSF, CXCR4, VEGF, TGF-β, ICAM-1) induced by peritoneal M2 macrophages increased significantly during the progression from inflammatory hyperplasia to carcinoma and metastasis. Similar functional changes occurred during peritoneal metastasis in M1 macrophages without changed polarization.

Conclusions

These results suggested that peritoneal M2 macrophages played a critical role in ulcerative colitis-associated carcinogenesis, including unbalanced pro-inflammatory and anti-inflammatory axis and enhanced expression of migration/invasion-associated factors. Furthermore, functional changes of M1 macrophages occurred without changed polarization during carcinogenesis and metastasis.