支气管哮喘患者肺泡巨噬细胞释放内皮素-1的研究

为研究支气管哮喘患者肺泡巨噬细胞（Ａｍ）所释放的内皮素１（ＥＴ１）在哮喘发病中的作用,对发作期哮喘患者２０例,对照组８例,经纤支镜行支气管肺泡灌洗术获得Ａｍ,调整Ａｍ至１×１０６／ｍｌ后分３孔体外培养（１ｍｌ／孔）;生理盐水对照孔、氟美松孔（１μｇ／ｍｌ）、氨茶碱孔（１５μｇ／ｍｌ）,培养６ｈ后取上清液,采用放免法测定ＥＴ１。显示哮喘组血浆ＥＴ１明显高于对照组,并于第１秒用力肺活量（ＦＥＶ１．０）呈显著负相关。哮喘组Ａｍ体外释放ＥＴ１与对照组无显著差异,与ＦＥＶ１．０无相关性。氟美松孔Ａｍ释放ＥＴ１明显低于生理盐水对照孔;氨茶碱孔与生理盐水孔或氟美松孔无差异。提示哮喘Ａｍ所释放ＥＴ１在哮喘发病中意义不大     

The role of VCAM-l/VLA-4 in the activation of allogenic T cells by murine macrophages

Vascular cell adhesion molecule 1 (VCAM-1) is a member of immunoglobulin superfamily. The principal ligand for VCAM-1 is integrin α4β/VLA-4 (very late antigen 4). It was reported that VCAM-1 was expressed on macrophages and dendritic cells, but little is known about its function on these professional antigen presenting cells (APC). The present study was performed to investigate the expression of VCAM-1 on macrophages and the role of VCAM-l/VLA-4 in the activation of allogenic T cells by murine macrophages. We analyzed VCAM-1 expression on peritoneal macrophages and macrophage cell line J774A.1 by fluorescence-activated cell sorting (FACS). Using neutralizing antibodies, we further analyzed the role of VCAM-l/VLA-4 interaction in macrophage and allogenic T cell mixed lymphocyte reaction (MLR). We found that VCAM-1 was constitutively expressed on macrophages and its expression level was upregulated by soluble tumor associated antigen (freeze-thaw lysates of FBL-3 leukemia cells) and TNF-a. In MLR assays, we observed that blocking VCAM-l/VLA-4 interaction with anti-VCAM-1 or anti-VLA-4 mAbs caused significant inhibition of the proliferative response and IL-2 production. These results suggest that VCAM-lon macrophages not only facilitates the cell-to-cell contact through adhesive interaction but also plays a role in the costimulation of T cells via its interaction with VLA-4 on the T cells. This work was supported by grants from the National Natural Science Foundation of China.No. (39730420). This is one of papers of the special issue on gene therapy research (Chin J Cancer Res Vol. 9 No. 4 December, 1997).     

Microcystin-LR promote intestinal secretion of water and electrolytes in rats.

We showed previously that exposure to microcystin-LR causes renal toxic effects in isolated perfused rat kidney, and that inflammatory mediators from supernatants of macrophages stimulated by microcystin-LR are involved in this process. The aim of this research was to examine water and electrolytes secretion in vivo, induced by microcystin-LR and supernatant of macrophages stimulated for this toxin (SUP.MphiS + MCLR), using perfused rat ileal segment and ligated intestinal loop models. We found microcystin-LR at 1 microg/ml (0.09 +/- 0.003* vs. control 0.07 +/- 0.001 g of secretion/2 cm of loop; P < 0.05*) and the SUP.MphiS + MCLR after 18 h postinoculation (0.10 +/- 0.003 vs. control 0.03 +/- 0.002 g/cm) caused intestinal secretion. In addition, microcystin-LR caused significant sodium secretion (-2.18 +/- 0.72* vs. control 2.18 +/- 0.50 microEq g(-1) min(-1)), potassium (-0.26 +/- 0.04* vs. control 0.32 +/- 0.03 microEq g(-1) min(-1)), chloride (MCLR = -3.29 +/- 1.93* vs. control 0.88 +/- 1.25 microEq g(-1) min(-1)) and water (-0.012 +/- 0.004* vs. control 0.002 +/- 0.002 ml g(-1) min(-1)). We also demonstrated SUP.MphiS + MCLR to induce intestinal secretion of electrolytes (sodium, potassium, chloride) and water. These findings suggested that microcystin-LR and lamina propria macrophages-derived mediators are able to induce intestinal secretion in vivo, probably via inhibition of protein phosphatase.     

非对称性二甲基精氨酸促进大鼠腹腔巨噬细胞诱导型一氧化氮合酶表达

目的观察非对称性二甲基精氨酸(asymmetric dimethylarginine,ADMA)对培养的大鼠腹腔巨噬细胞诱导型一氧化氮合酶(iNOS)表达的影响,探讨其在脂质沉积中的作用及机制。方法分组①正常对照组以PBS缓冲液与巨噬细胞共孵育,②氧化型低密度脂蛋白(oxLDL)组在巨噬细胞培养液中加oxLDL(50mg/L),③O A组在巨噬细胞培养液中加oxLDL(50mg/L)和ADMA(20μmol/L),④O A L组在培养液中加ox-LDL(50mg/L)、ADMA(20μmol/L)、L-Arg(1.2mmol/L)。以硝酸还原酶法和化学比色法分别测定培养液中一氧化氮(NO)含量与iNOS活力,以RT-PCR和Western Blotting分别检测iNOSmRNA和蛋白表达。结果①OxLDL组与O A组NO含量降低,iNOS活力升高,且以O A组明显,与正常对照组比较差异均有统计学意义(均P<0.05);NO与iNOS呈负相关(r=-0.697,P<0.05)。②O A组iNOS mRNA和蛋白表达增强,与正常对照组相比差异有统计学意义(P<0.05或P<0.01)。结论ADMA抑制大鼠腹腔巨噬细胞合成NO,增强iNOS基因与蛋白表达,可能是ADMA促使巨噬细胞转变为泡沫细胞、促进动脉粥样硬化发生发展的机制之一。     

神经胶质瘤中肿瘤相关巨噬细胞研究进展

肿瘤相关巨噬细胞（TAM）是异质性细胞群体,是肿瘤微环境中炎性细胞的主要成分。这些细胞在神经胶质瘤中部分源自中枢神经系统小胶质细胞和循环单核细胞,并与神经胶质瘤的血管生成、免疫抑制、肿瘤进展和侵袭有关。文章综述了TAM通过各种途径促进神经胶质瘤发展的潜在机制,为神经胶质瘤的靶向治疗提供了新的可能性。     

靶向肿瘤相关巨噬细胞治疗卵巢癌的研究进展

卵巢癌是致死率最高的妇科恶性肿瘤,免疫因素在卵巢癌的发生、发展中起重要作用。肿瘤相关巨噬细胞（tumor-associated macrophages,TAMs）是卵巢癌肿瘤微环境中重要的免疫细胞,可分泌多种细胞因子促进癌细胞增殖及血管生成并抑制肿瘤免疫,在卵巢癌进展中发挥重要作用。因此,靶向TAMs治疗卵巢癌成为目前的研究热点之一。其具体治疗策略包括：抑制TAMs的募集、增强TAMs的吞噬能力、消耗TAMs、将M2型TAMs去极化为M1型TAMs或抑制TAMs向M2型极化以及阻断TAMs与肿瘤细胞的相互作用。目前,已有诸多基础研究证实化疗药物、免疫检查点阻断剂、纳米药物和中药调节TAMs治疗或协同治疗对卵巢癌具有一定疗效。综述靶向TAMs治疗卵巢癌的原理及相关进展,以期为其进一步研究及临床应用提供参考。     

巨噬细胞诱发的实验性增生性玻璃体视网膜病变中的几种早期炎性细胞因子

目的观察在增生性玻璃体视网膜病变（ｐｒｏｌｉｆｅｒａｔｉｖｅｖｉｔｒｅｏｒｅｔｉｎｏｐａｔｈｙ,ＰＶＲ）模型中几种早期炎性细胞因子的含量变化及其与ＰＶＲ形成时相的关系。方法采集巨噬细胞诱发的兔眼ＰＶＲ模型的玻璃体液用双抗体夹心法酶联免疫吸附测试（ｅｎｚｙｍｅｌｉｎｋｅｄｉｍｍｕｎｏｓｏｒｂｅｎｔａｓａｙ,ＥＬＩＳＡ）试剂盒,检测样本中肿瘤坏死因子α（ｔｕｍｏｒｎｅｃｒｏｓｉｓｆａｃｔｏｒα,ＴＮＦα）、白细胞介素８（ｉｎｔｅｒｌｅｕｋｉｎ８,ＩＬ８）和白细胞介素６（ｉｎｔｅｒｌｅｕｋｉｎ６,ＩＬ６）的含量,并进行多元回归拟合曲线分析。结果ＴＮＦα在巨噬细胞注入后７天上升,２１天达高峰值,为３０９ｐｇ／ｍｌ;ＩＬ８与ＩＬ６在７～２１天也维持高水平,在１４天分别达高峰值１３２５ｐｇ／ｍｌ和９９８ｐｇ／ｍｌ。与对照组比较差异均有非常显著性。结论３种早期炎性细胞因子ＴＮＦα、ＩＬ８和ＩＬ６在本模型的炎症期和细胞增生期均呈明显的高水平,在瘢痕期水平降低,提示它们对眼内细胞增生具有重要的启动和调控作用。     

Differential flow cytometric detection of intracellular cytokines in peripheral and peritoneal mononuclear cells of women with endometriosis

OBJECTIVE: The pathogenesis of endometriosis is related to functional changes in CD3+ and CD14+ cells observed both at the local and systemic level. Here we investigated whether, and if so, how the body compartment influences cytokine expression in stimulated peritoneal and peripheral CD3+ and CD14+ cells of women with endometriosis. STUDY DESIGN: Isolated peripheral blood (PB) and peritoneal fluid (PF) mononuclear cells from women with endometriosis were cultured under non-adherent conditions and stimulated with PMA and ionomycin for 6h to induce intracellular cytokine synthesis of TNF-alpha, IFN-gamma, and IL-8 by CD3+ cells or with LPS for 9h to produce TNF-alpha, IL-6, IL-10, MCP-1, and IL-8 by CD14+ cells. RESULTS: The percentages of positive CD3+ cells stained for TNF-alpha and IFN-gamma were significantly higher and those stained for IL-8 were significantly lower in PF compared with PB, this being independent of the stage of endometriosis. In contrast, the percentages of CD14+ cells producing TNF-alpha, IL-6, IL-10, MCP-1, and IL-8 were significantly higher in PB than PF of women with endometriosis. CONCLUSIONS: Monocytes/macrophages and lymphocytes derived from the peripheral and peritoneal compartments of women with endometriosis differentially respond to stimulated cytokine synthesis induction. However, it is difficult to state whether the observed phenomenon is more related to body compartment influence per se or to the presence of endometriosis.     

高通量基因芯片初步筛选ITP脾和正常脾巨噬细胞的差异表达基因

目的研究免疫性血小板减少性紫癜(ITP)患者脾和正常脾巨噬细胞的差异表达基因。方法提取9例ITP患者及9例正常人脾巨噬细胞的总RNA,反转录制备含荧光分子Cy5标记的cDNA探针,与含有30968点cDNA的表达谱芯片杂交后扫描荧光强度,筛选出差异表达的基因并进行分析。结果基因芯片共筛选出1545个差异表达基因,其中上调基因718个,下调基因827个。差异表达基因涉及免疫反应、细胞黏附及受体调节、细胞信号转导、细胞骨架和运动代谢、细胞凋亡、酶调活性等方面。差异表达基因生物路径涉及Toll样受体信号通路,Fcγ受体介导吞噬通路,促分裂原活化蛋白激酶信号通路、细胞内吞通路等。结论基因芯片筛选ITP患者脾巨噬细胞的差异表达基因是有效的。差异表达基因可能为ITP发病机制的研究提供新证据和治疗靶标。     

采用流式细胞分选术区分中枢神经系统内小胶质细胞和浸润巨噬细胞

目的 建立通过流式细胞术区分中枢神经系统内小胶质细胞和浸润巨噬细胞的方法。方法 成年C57BL/6小鼠建立双侧颈总动脉狭窄模型,分别采用PLX5622或氯磷酸盐脂质体给药处理。分离、匀浆、重悬后的脑和脊髓组织,通过Percoll密度梯度离心,得到单个核细胞悬液。进一步采用抗CD45、抗CD11b和抗Ly6C抗体,流式分选获得小胶质细胞(CD11b+ CD45low Ly-6C-^细胞群)和浸润巨噬细胞(CD11b+ CD45high Ly-6C+^细胞群),并分析和验证PLX5622和氯磷酸盐脂质体两种给药范式获得的处理效果。结果 通过抗CD45、抗CD11b和抗Ly6C三种抗体可以有效区分中枢神经系统内小胶质细胞和浸润巨噬细胞。PLX5622给药组的小胶质细胞显著减少,而氯磷酸盐脂质体给药组的浸润巨噬细胞显著减少。结论 本方法利用流式分选术有效区分中枢神经系统内小胶质细胞和浸润巨噬细胞;PLX5622和氯磷酸盐脂质体两种给药范式可分别选择性清除中枢神经系统内的小胶质细胞和浸润的巨噬细胞。