风湿性心脏瓣膜钙化的二尖瓣组织中巨噬细胞和炎性因子表达分析

目的 探讨风湿性心脏瓣膜钙化的二尖瓣组织中巨噬细胞分布规律和相关炎性因子表达情况.方法 收集行二尖瓣置换术的风湿性心脏病患者13例为研究组,同期5例终末期心脏病行心脏移植手术的患者为对照组.收集二尖瓣组织标本,用CD68标记全部巨噬细胞,用iNOS,CD163标记M1,M2型巨噬细胞,观察钙化的二尖瓣组织中巨噬细胞浸润情况,同时观察巨噬细胞相关炎症介质(eNOS,IL-10,Arg-1,巨噬细胞集落刺激因子)在钙化二尖瓣中的表达情况.结果 研究组CD68表达积分高于对照组,差异具有统计学意义(P<0.05),间质和内膜层存在大量巨噬细胞浸润;研究组iNOS阳性的M1型巨噬细胞表达积分高于对照组,差异具有显著统计学意义(P<0.01);研究组CD163阳性的M2型巨噬细胞表达积分低于对照组,差异具有统计学意义(P<0.05);研究组eNOS的表达积分高于对照组,差异具有显著统计学意义(P<0.01);研究组抑炎因子IL-10,Arg-1表达积分低于对照组,差异均具有显著统计学意义(P<0.01);研究组巨噬细胞集落刺激因子的表达量低于对照组,差异具有显著统计学意义(P<0.01).结论 风湿性二尖瓣钙化过程中以M1型巨噬细胞浸润为主,可通过上调eNOS等炎性因子来直接促进炎性反应,通过抑制IL-10等炎性因子来间接促进炎性反应.巨噬细胞集落刺激因子表达下调可减少M1型向M2型转化,并长期维持炎性反应.     

1型糖尿病致病机理的细胞学研究

1型糖尿病是以自身免疫系统对胰岛β细胞进行特异性损伤为特征的自身免疫疾病，其中T淋巴细胞，B淋巴细胞及巨噬细胞对胰岛β细胞的损伤起重要作用。     

紫花地丁提取物对LPS诱导RAW 264.7细胞的体外抗炎作用

观察紫花地丁提取物对脂多糖诱导的RAW 264.7巨噬细胞的抗炎作用并探讨其机制.采用脂多糖(1μg/mL)诱导RAW 264.7巨噬细胞建立体外炎症模型,格里斯试剂法和酶联免疫吸附法分别检测细胞上清液中的NO、TNF-α、IL-6、IL-1β的浓度,凝胶成像法检测细胞中iNOS、COX-2蛋白水平.结果显示,紫花地丁水提物、醇提物剂量依赖地降低脂多糖刺激巨噬细胞分泌NO、TNF-α、IL-6、IL-1β以及其iNOS、COX-2蛋白水平.相同剂量(100μg/mL)的紫花地丁醇提物抗炎活性优于水提物.结果表明,紫花地丁水提物、醇提物可抑制脂多糖刺激的RAW 264.7细胞炎症,其机制可能与抑制iNOS、COX-2蛋白表达,减少NO、TNF-α、IL-6、IL-1β等炎性因子的分泌有关.     

甘精胰岛素联合诺和龙对2型糖尿病患者 胰岛功能及TNF-α,IL-6的影响

目的研究甘精胰岛素联合诺和龙对于高血糖状态的初发2型糖尿病患者胰岛功能、TNF-α及IL-6的作用.方法选取初发2型糖尿病高血糖状态患者120例,随机平均分为观察组和对照组,每组60例.两组患者均给予糖尿病饮食控制、运动训练和健康知识讲座等基本治疗.对照组在此基础上予以诺和灵30R治疗;观察组在此基础上予以甘精胰岛素联合诺和龙治疗.治疗3个月后复查患者TNF-α、IL-6、胰岛素水平和低血糖事件发生率.结果观察组患者空腹胰岛素水平显著低于对照组(P0.05),而餐后2 h胰岛素水平显著高于对照组(P0.05),差异具有统计学意义.观察组患者TNF-α,IL-6水平均显著低于对照组(P0.05),差异具有统计学意义.低血糖事件发生率观察组显著低于对照组(P0.05),差异具有统计学意义.结论甘精胰岛素联合诺和龙控制血糖效果好,能有效改善2型糖尿病高血糖状态患者体内胰岛功能,同时降低TNF-α,IL-6水平.     

IL-17、VEGF与糖尿病微血管病变部位及区域数量的关系

目的:探讨2型糖尿病患者不同部位及区域数量微血管病变外周血IL-17和VEGF水平的变化和意义.方法:检测2型糖尿病患者外周血中IL-17和VEGF水平,按照微血管病变部位,比较糖尿病视网膜病变、糖尿病肾病及糖尿病周围神经病变患者以及不同损伤区域数量微血管病变患者IL-17和VEGF的水平.结果:糖尿病微血管病变患者IL-17、VEGF水平高于无血管病变患者和健康对照组者(p<0.01).糖尿病视网膜病变、糖尿病肾病和糖尿病周围神经病变患者IL-17、VEGF水平比较差异无统计学意义(p<0.01).合并3种微血管病变患者IL-17、VEGF水平高于合并1种微血管病变患者(p<0.01),合并2种微血管病变患者IL-17、VEGF水平高于合并1种微血管病变患者(p<0.01),糖尿病微血管病变组的血清IL-17与VEGF有明显的正相关(r=0.412,p<0.05)结论糖尿病微血管病变患者外周IL-17、VEGF水平升高,IL-17、VEGF水平与微血管病变部位无关,与微血管病变损伤区域数量有关,炎症因子IL-17、VEGF水平及其相互作用与糖尿病微血管病变的广泛性一致.     

小白菊内酯对RAW264.7炎症细胞因子表达的抑制

目的:研究小白菊内酯对细菌脂多糖诱导炎症反应的抑制作用.方法:选用小鼠巨噬细胞RAW264.7,采用实时定量多聚酶链式反应及酶联免疫吸附法,检测不同浓度小白菊内酯(0.1、1、10、50μmol/L)和细菌脂多糖(10μmol/L)作用后,细胞因子的mRNA(IL-6、TNF-α、Ptgs2、IL-1β、IL-10、MMP13与CXCL1)及蛋白(IL-6、TNF-α、IL-1β与IL-10)表达水平的改变.实验同时设立4个对照组,包括DMSO组、脂多糖组、DMSO+脂多糖组及阳性药Bay 11-7082+脂多糖组.结果:浓度分别为10和50μmol/L小白菊内酯稳定有效地抑制脂多糖诱导小鼠巨噬细胞产生细胞因子.在mRNA水平上,IL-6、TNF-α、Ptgs2、IL-1β、IL-10与MMP13的mRNA表达水平明显下降,差异有显著性,CXCL1 mRNA表达水平改变无统计学差异;在蛋白水平上,IL-6、TNF-α、IL-1β与IL-10的表达亦显著性下降,有统计学差异.结论:小白菊内酯有效抑制了细菌脂多糖诱导的小鼠巨噬细胞中炎症细胞因子的表达.     

2型糖尿病患者的社区干预对血清IL-1、TNF-a的影响

探讨社区健康管理干预对2型糖尿病患者血清白细胞介素-1(IL-1)、肿瘤坏死因子-α(TNF-α)的影响。将社区264名2型糖尿病患者随机分为2组,实验组177名给与社区健康管理干预,督促其控制血糖、控制血压等,干预期6月,对照组87名不干预,测定两组干预前后的糖化血红蛋白、血清IL-1、TNF-α水平。实验组干预后的血清IL-1、TNF-α水平较对照组显著下降(P0.05),糖化血红蛋白在两组间无显著差异。结论社区健康管理干预能显著降低2型糖尿病患者的血清IL-1、TNF-α水平,提示其可减轻2型糖尿病患者的血管炎症反应。     

NLRP3炎症小体抑制剂研究进展

NLRP3是一种重要的胞内模式识别受体,可以与ASC及pro-caspase-1形成炎症小体,促进IL-1β和IL-18等炎性介质的成熟和分泌,从而促进炎症反应.NLRP3炎症小体活化失调与多种人类重大疾病密切相关,如痛风、动脉粥样硬化、神经退行性疾病和2型糖尿病等.因此NLRP3炎症小体是上述疾病的潜在干预靶点,许多NLRP3炎症小体抑制剂对相关疾病表现出良好的预防或者治疗效果.本文对NLRP3炎症小体抑制剂最近的研究进展进行简要综述.     

布鲁氏菌感染诱发巨噬细胞炎症反应的初步研究

为研究NLRP3炎症小体在布鲁氏菌侵染小鼠巨噬细胞过程中活化介导的炎症反应,建立牛种布鲁氏菌2308株侵染小鼠巨噬细胞RAW264.7的模型,侵染比例100∶1(细菌∶细胞),RT-PCR技术检测细菌侵染过程中NLRP3炎症小体相关分子NLRP3和Caspase-1 m RNA的变化水平;Western blot分析NLRP3和Caspase-1在蛋白水平的变化;ELISA检测感染细胞上清中炎症因子IL-18和IL-1β的释放量。结果表明:与对照组相比布鲁氏菌感染小鼠巨噬细胞后NLRP3和Caspase-1 m RNA和蛋白表达水平显著增加(P0.05),炎性因子IL-1β和IL-18的含量也显著升高(P0.05)。结论:布鲁氏菌感染可激活NLRP3炎症小体,为进一步研究布鲁氏菌感染引发致病机制奠定理论基础。     

2型糖尿病对动脉粥样硬化的影响机制

2型糖尿病的主要特征是胰岛素抵抗和胰岛素分泌相对不足,研究表明胰岛素具有抗炎的功能,胰岛素抵抗或胰岛素分泌相对不足会促进炎症反应。动脉粥样硬化是一种动脉壁炎症性疾病,包括内皮功能异常,单核细胞募集,白细胞粘附及迁移,氧化型低密度脂蛋白(ox-LDL)被巨噬细胞摄取,泡沫细胞形成,最终形成粥样斑块。2型糖尿病对动脉粥样硬化的影响是多方面的,包括加剧炎症反应、糖基化终末产物的增加和糖脂代谢紊乱等。本文主要从炎症角度分析,阐明炎症反应与胰岛素抵抗互作的机理,对于2型糖尿病和动脉粥样硬化的预防及治疗具有十分重要的意义。     

自拟方“通痹祛风汤”对大鼠急性痛风性关节炎抗炎作用研究

目的:观察"通痹祛风汤"对急性痛风性关节炎模型大鼠不同时相的影响,探讨其治疗急性痛风性关节炎的作用机制.方法:注射尿酸钠建立大鼠痛风性关节炎模型,大鼠灌胃给药,观察对大鼠关节肿胀率及关节滑膜囊表达IL-1 mRNA和TNF-mRNA(RT-PCR)的影响.结果:"通痹祛风汤"可有效抑制大鼠踝关节肿胀程度;关节滑膜囊TNF-mRNA和IL-1 mRNAr的表达明显减低.结论:"通痹祛风汤"可抑制痛风性关节炎TNF-和IL-1的产生而发挥抗炎作用,明显减轻关节的病理性损伤,有效抑制关节炎的临床进程.     

高血糖条件下神经降压素调节巨噬细胞炎症反应的实验研究

目的观察神经降压素在高血糖条件下对巨噬细胞的影响。方法取昆明小鼠脾巨噬细胞,高糖培养液培养15d,细胞分PBS对照组和神经降压索干预组。ELISA检测法检测TNF-、IL-1β及IL-6的释放;RT—PCR检测TNF-、IL-1β及IL-6mRNA的表达。结果高糖条件下神经降压素能降低巨噬细胞TNF-、IL-1β及IL-6的释放和mRNA的表达（P〈0．05）。结论在高糖条件下,神经降压素可以通过抑制炎性细胞因子的释放影响巨噬细胞的反应。     

Macrophage-specific PPARgamma controls alternative activation and improves insulin resistance

Obesity and insulin resistance, the cardinal features of metabolic syndrome, are closely associated with a state of low-grade inflammation. In adipose tissue chronic overnutrition leads to macrophage infiltration, resulting in local inflammation that potentiates insulin resistance. For instance, transgenic expression of Mcp1 (also known as chemokine ligand 2, Ccl2) in adipose tissue increases macrophage infiltration, inflammation and insulin resistance. Conversely, disruption of Mcp1 or its receptor Ccr2 impairs migration of macrophages into adipose tissue, thereby lowering adipose tissue inflammation and improving insulin sensitivity. These findings together suggest a correlation between macrophage content in adipose tissue and insulin resistance. However, resident macrophages in tissues display tremendous heterogeneity in their activities and functions, primarily reflecting their local metabolic and immune microenvironment. While Mcp1 directs recruitment of pro-inflammatory classically activated macrophages to sites of tissue damage, resident macrophages, such as those present in the adipose tissue of lean mice, display the alternatively activated phenotype. Despite their higher capacity to repair tissue, the precise role of alternatively activated macrophages in obesity-induced insulin resistance remains unknown. Using mice with macrophage-specific deletion of the peroxisome proliferator activated receptor-gamma (PPARgamma), we show here that PPARgamma is required for maturation of alternatively activated macrophages. Disruption of PPARgamma in myeloid cells impairs alternative macrophage activation, and predisposes these animals to development of diet-induced obesity, insulin resistance, and glucose intolerance. Furthermore, gene expression profiling revealed that downregulation of oxidative phosphorylation gene expression in skeletal muscle and liver leads to decreased insulin sensitivity in these tissues. Together, our findings suggest that resident alternatively activated macrophages have a beneficial role in regulating nutrient homeostasis and suggest that macrophage polarization towards the alternative state might be a useful strategy for treating type 2 diabetes.     

beta-Cell death during progression to diabetes.

The hallmark of type 1 diabetes is specific destruction of pancreatic islet beta-cells. Apoptosis of beta-cells may be crucial at several points during disease progression, initiating leukocyte invasion of the islets and terminating the production of insulin in islet cells. beta-Cell apoptosis may also be involved in the occasional evolution of type 2 into type 1 diabetes.     

山香圆总黄酮对慢性咽炎模型大鼠 NF-κB、IκBα表达的影响

以2.5%氨水喷咽部建立大鼠慢性咽炎模型,用HE染色法观察咽部病理形态学改变,用ELISA法检测血清炎症因子TNF-α、IL-1β、IL-6,用RT-qPCR测定咽部组织NF-κBp65mRNA的表达,用Western blot检测咽部组织NF-κBp65和IκBα蛋白的表达,探讨山香圆总黄酮对慢性咽炎模型大鼠 NF-κB、IκBα表达的影响.结果显示:山香圆总黄酮能明显改善模型大鼠咽部病理形态学; 与模型组比较,山香圆总黄酮各组能不同程度地下调TNF-α、IL-1β、IL-6和NF-κBp65的表达并上调IκBα的表达,其中高剂量组最显著.这表明山香圆总黄酮能通过抑制NF-κB的表达和IκBα的解离下调炎症因子TNF-α、IL-1β、IL-6的表达,从而减轻炎症,发挥抗慢性咽炎作用.     

Non-canonical inflammasome activation targets caspase-11

Caspase-1 activation by inflammasome scaffolds comprised of intracellular nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) and the adaptor ASC is believed to be essential for production of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 during the innate immune response. Here we show, with C57BL/6 Casp11 gene-targeted mice, that caspase-11 (also known as caspase-4) is critical for caspase-1 activation and IL-1β production in macrophages infected with Escherichia coli, Citrobacter rodentium or Vibrio cholerae. Strain 129 mice, like Casp11(-/-) mice, exhibited defects in IL-1β production and harboured a mutation in the Casp11 locus that attenuated caspase-11 expression. This finding is important because published targeting of the Casp1 gene was done using strain 129 embryonic stem cells. Casp1 and Casp11 are too close in the genome to be segregated by recombination; consequently, the published Casp1(-/-) mice lack both caspase-11 and caspase-1. Interestingly, Casp11(-/-) macrophages secreted IL-1β normally in response to ATP and monosodium urate, indicating that caspase-11 is engaged by a non-canonical inflammasome. Casp1(-/-)Casp11(129mt/129mt) macrophages expressing caspase-11 from a C57BL/6 bacterial artificial chromosome transgene failed to secrete IL-1β regardless of stimulus, confirming an essential role for caspase-1 in IL-1β production. Caspase-11 rather than caspase-1, however, was required for non-canonical inflammasome-triggered macrophage cell death, indicating that caspase-11 orchestrates both caspase-1-dependent and -independent outputs. Caspase-1 activation by non-canonical stimuli required NLRP3 and ASC, but caspase-11 processing and cell death did not, implying that there is a distinct activator of caspase-11. Lastly, loss of caspase-11 rather than caspase-1 protected mice from a lethal dose of lipopolysaccharide. These data highlight a unique pro-inflammatory role for caspase-11 in the innate immune response to clinically significant bacterial infections.     

Interleukin-23 rather than interleukin-12 is the critical cytokine for autoimmune inflammation of the brain

Interleukin-12 (IL-12) is a heterodimeric molecule composed of p35 and p40 subunits. Analyses in vitro have defined IL-12 as an important factor for the differentiation of naive T cells into T-helper type 1 CD4+ lymphocytes secreting interferon-gamma (refs 1, 2). Similarly, numerous studies have concluded that IL-12 is essential for T-cell-dependent immune and inflammatory responses in vivo, primarily through the use of IL-12 p40 gene-targeted mice and neutralizing antibodies against p40. The cytokine IL-23, which comprises the p40 subunit of IL-12 but a different p19 subunit, is produced predominantly by macrophages and dendritic cells, and shows activity on memory T cells. Evidence from studies of IL-23 receptor expression and IL-23 overexpression in transgenic mice suggest, however, that IL-23 may also affect macrophage function directly. Here we show, by using gene-targeted mice lacking only IL-23 and cytokine replacement studies, that the perceived central role for IL-12 in autoimmune inflammation, specifically in the brain, has been misinterpreted and that IL-23, and not IL-12, is the critical factor in this response. In addition, we show that IL-23, unlike IL-12, acts more broadly as an end-stage effector cytokine through direct actions on macrophages.     

兔膝骨关节炎模型的建立及评价

目的 通过兔软骨缺损制备兔膝骨关节炎模型并进行评价。方法 将动物按体质量和性别随机分成3组：假手术组,模型组和阳性对照氨基葡萄糖组,每组12只。造模4周后开始灌胃给药,每天1次,连续给药4周。末次给药后,将动物处死,ELISA试剂盒检测血清及关节液中IL-1β和TNF-α水平,对关节软骨和滑膜进行HE染色,并对关节软骨进行Mankin’s评分以评价其损伤情况。结果 与假手术组相比,模型组血清、关节液中IL-1β、TNF-α水平明显升高,而给予氨基葡萄糖可降低血清、关节液中IL-1β、TNF-α水平。HE染色见模型组软骨结构改变、缺损,滑膜有炎细胞浸润;软骨评分明显高于假手术组。给予氨基葡萄糖可以减轻软骨缺损,减少滑膜炎细胞浸润,降低软骨评分。结论 关节软骨钻孔可建立兔膝骨关节炎模型,反映其主要特征,并能通过给予阳性对照药减轻损伤和炎症反应。该模型可用于治疗关节炎药物的评价和筛选。     

蜜环菌β-1,6-葡聚糖通过调节人源巨噬细胞极化发挥抑制结肠癌作用

以人源巨噬细胞THP-1诱导的M2型巨噬细胞为模型,研究从蜜环菌中分离纯化出的β-1,6-葡聚糖(AAMP-A70)对其极化影响,并分析AAMP-A70通过调节巨噬细胞极化抑制结肠癌细胞DLD-1增殖的作用及机制.采用酶联免疫吸附测定(ELISA)和实时荧光定量PCR(qRT-PCR)检测AAMP-A70处理前后THP-1诱导的M2型巨噬细胞中M1型和M2型巨噬细胞标志物表达情况;将各处理组的细胞培养液处理DLD-1细胞,通过MTT和结晶紫染色检测DLD-1细胞的增殖情况;并通过Western-blot实验检测DLD-1细胞的凋亡蛋白表达情况.实验结果表明：AAMP-A70可显著下调M2型巨噬细胞标志物VEGF,Arg-1,TGF-β的表达,促进M1型巨噬细胞标志物TNF-α,IL-1β,CCR7的表达;AAMP-A70处理组细胞的培养液可显著抑制DLD-1的增殖,且呈浓度依赖趋势;AAMP-A70处理组的细胞上清液可诱导DLD-1细胞中表达的cleaved-PARP和cleaved-Caspase-3上调.