巨噬细胞集落刺激因子对RAW264.7细胞谷胱甘肽过氧化物酶活性及其基因表达的影响

为探讨巨噬细胞集落刺激因子抗单核巨噬细胞过氧化损伤的机理 ,以富含巨噬细胞集落刺激因子的L92 9细胞条件培养基作为细胞因子来源 ,采用酶活性测定、反转录聚合酶链反应等技术 ,研究了巨噬细胞集落刺激因子对小鼠巨噬细胞系RAW2 6 4.7细胞谷胱甘肽过氧化物酶活性及mRNA表达的影响。结果发现 ,巨噬细胞集落刺激因子可以提高RAW2 6 4.7细胞硒谷胱甘肽过氧化物酶及非硒谷胱甘肽过氧化物酶活性 ,并增强RAW2 6 4.7细胞血浆谷胱甘肽过氧化物酶mRNA的表达 ,而对RAW2 6 4.7细胞磷脂氢过氧化物谷胱甘肽过氧化物酶的表达无诱导作用     

巨噬细胞集落刺激因子对小鼠腹腔巨噬细胞抗氧化能力的影响

越来越多的实验结果证实巨噬细胞集落刺激因子在动脉粥样硬化发生过程中的重要作用，为了探讨巨噬细胞集落刺激因子和活性氧与动脉粥样硬化之间的相互关系，观察了巨噬细胞集落刺激因子对培养的小鼠腹腔巨噬细胞抗氧化酶活性的影响。结果发现巨噬细胞集落刺激因子能使小鼠腹腔巨噬细胞谷胱甘肽过氧化物酶活性提高３２％，超氧化物歧化酶的活性提高１２０％，并减轻叔丁基氢过氧化物促巨噬细胞源性泡沫细胞形成，以及提高巨噬细胞的生存数。此结果提示巨噬细胞集落刺激因子提高杭氧化酶活性的作用可能是其虽增加对氧化修饰低密度脂蛋白和胆固醇的摄入却能阻止泡沫细胞形成及动脉粥样硬化发生发展的原因之一。     

巨噬细胞集落刺激因子对小鼠腹腔巨噬细胞抗氧化能力的影响

越来越多的实验结果证实巨噬细胞集落刺激因子在动脉粥样硬化发生过程中的重要作用，为了探讨巨噬细胞集落刺激因子和活性氧与动脉粥样硬化之间的相互关系，观察了巨噬细胞集落刺激因子对培养的小鼠腹巨噬细胞抗氧化酶活性的影响。结果发现巨噬细胞集落刺激因子能使小鼠腔巨噬细胞谷胱甘肽过氧化物酶活性提高３２％，超氧化歧化酶的活性提高１２０％，并减轻叔丁基氢过氧化物促巨噬细胞源性泡沫细胞形成，以及提高巨噬细胞的生存数。     

巨噬细胞集落刺激因子与脑缺血

在中枢神经系统内 ,巨噬细胞集落刺激因子主要由星形细胞产生。发育期的大脑皮质神经元和小胶质细胞表达巨噬细胞集落刺激因子的受体。在脑缺血时 ,中枢神经系统的巨噬细胞集落刺激因子及其受体的表达均上调 ,补充外源性巨噬细胞集落刺激因子可明显减轻缺血性脑损伤 ,推测这种缺血保护作用是通过巨噬细胞集落刺激因子与受体介导的信号传导途径以及与其他因子的相互作用实现的。     

巨噬细胞集落刺激因子对小鼠腹腔巨噬细胞清道夫受体途径和细胞内胆固醇酯积聚的影响

为探索巨噬细胞集落刺激因子、清道夫受体、氧化型低密度脂蛋白与动脉粥样硬化的关系，观察了重组人巨噬细胞集落刺激因子对小鼠腹腔巨噬细胞清道夫受体途径的影响以及重组人巨噬细胞集落刺激因子对氧化型低密度脂蛋白所致细胞内胆固醇酯积聚的影响．结果发现重组人巨噬细胞集落刺激因子能增加培养的小鼠腹腔巨噬细胞表面的清道夫受体数目，使之对氧化型低密度脂蛋白的结合和降解呈现剂量和时间依赖性增加，并使细胞内胆固醇酯积聚增多．表明巨噬细胞集落刺激因子可通过增加清道夫受体数目使小鼠腹腔巨噬细胞对氧化型低密度脂蛋白的结合和降解增多，从而增加细胞内胆固醇酯含量，促进动脉壁内泡沫细胞形成．     

巨噬细胞集落刺激因子与脑缺血

在中枢神经系统内，巨噬细胞集落刺激因子主要由星形细胞产生，发育期的大脑皮质神经元和小脑质细胞表达巨噬细胞集落刺激因子的受体，在脑缺血时，中枢神经系统的巨噬细胞集落刺激因子及其受体的表达均上调，补充外源性巨噬细胞集落刺激因子可明显减轻缺血性脑损伤，推测这种缺血保护作用是通过巨噬细胞集落刺激因子与受体介导的信号传导途径以及与其他因子的相互作用实现的。     

巨噬细胞集落刺激因子在动脉粥样硬化发生中的作用

动脉粥样硬化斑块损伤处可见巨噬细胞集落刺激因子表达增强，巨噬细胞集落刺激因子能通过调节核－巨噬细胞的一系列功能，影响胆固醇代谢，抑制高胆固醇血症家兔泡沫细胞形成及减少主动脉斑块面积。巨噬细胞集落刺激因子与氧化修饰低密度脂蛋白关系密切，二者可以相互作用，共同影响着动脉粥样硬化的进程。     

其它

其它２６４ＩｇＡ类风湿因子亚型的临床意义［英］／ＪｏｎｓｓｏｎＴ…∥ＡｎｎＲｈｅｕｍＤｉｓ．－１９９５，５４（７）．－５７８～５８１本文评价了ＩｇＡ类风湿因子（ＲＦ）亚型在类风湿性关节炎（ＲＡ）诊断及发病机制中的意义。方法用酶联免疫吸附法（ＥＬＩＳＡ...     

哮喘患者淋巴细胞粒细胞—巨噬细胞集落刺激因子mRNA的表达

哮喘患者淋巴细胞粒细胞－巨噬细胞集落刺激因子ｍＲＮＡ的表达金远林王长征王春霞钱桂生活化的Ｔ淋巴细胞产生的细胞因子白细胞介素（ＩＬ）－５、粒细胞－巨噬细胞集落刺激因子（ＧＭ－ＣＳＦ）及ＩＬ－３，在以嗜酸细胞（Ｅｏｓ）为主的哮喘非特异性气道炎症产生中起重...     

巨噬细胞集落刺激因子对DEL细胞增殖作用的研究

目的 研究巨噬细胞集落刺激因子对动脉粥样硬化病变处伴随血管平滑肌细胞共同积聚的巨噬细胞的增殖作用及机制。方法 以能分化为巨噬细胞表型的ＤＥＬ细胞为离体细胞实验模型。将富含巨噬细胞集落刺激因子的１９２９细胞条件培养液或重组的人巨噬细胞集落刺激因子加入体外培养的处于静止期的ＤＥＬ细胞,以氚标胸腺嘧啶核苷或５－溴－悄嘧啶 核苷掺入法分别检测ＤＥＬ细胞ＤＮＡ的合成,另以Ｎｏｒｔｈｅｒｎ ｂｌｏｔ法检测ＤＥ     

Macrophage colony-stimulating factor reduces tert-butyl hydroperoxide induced oxidative injury to monocytes/macrophages.

The transformation of macrophages into foam cells is an important event in the development of atherosclerosis, and the oxidative injury caused by oxidized low density lipoprotein (Ox-LDL) plays an essential role in that process. It has been proved that macrophage colony-stimulating factor (M-CSF) could prevent the progression of atherosclerosis in Watanabe heritable hypercholesterolemic (WHHL) rabbits. We proposed that the anti-atherogenic effect of M-CSF was partly associated with its protective effect on monocyte-derived macrophages from Ox-LDL induced oxidative injury. In order to prove this, we investigated the effect of M-CSF on the oxidative injury caused by tert-butyl hydroperoxide (tbOOH) to mouse peritoneal macrophages and U937/J774 cell lines. The results showed that M-CSF could protect mouse peritoneal macrophages from oxidative injury (presented by cell morphology and cell survival rate); L929 cell-conditioned medium (L929-CM) had the same effect as M-CSF; and anti-M-CSF monoclonal antibody could mostly block the protective effect of L929-CM on macrophages. L929-CM was proved to be also able to decrease the impact of plasma membrane fluidity in U937 and J774 cells treated with tbOOH. Incubation with tbOOH caused DNA fragmentation in U937 cells. The presence of L929-CM greatly reduced the number of apoptotic U937 cells characterized by DNA fragmentation. From these results, we concluded that M-CSF could protect monocytes/macrophages from oxidative injury. It may be one of the mechanisms which explain the anti-atherogenic effect of exogenous M-CSF in WHHL rabbits.     

Potential role of macrophage colony-stimulating factor in the proliferation of DEL cells.

The replication and activation of both vascular smooth muscle cells and macrophages, which have previously entered the arterial wall, are key events in the atherosclerotic process. The importance of macrophage colony-stimulating factor (MCSF) in control of the growth/proliferation of both cell types confers to this compound a central role in the development of vascular lesions. In order to gain insight into the mechanisms of macrophage proliferation, we investigated the effect of MCSF upon the proliferation of DEL cells. DEL cells constitute a monocyte/histiocytic cell line that differentiates along a macrophage lineage following exposure to phorbol ester. DEL cells constitutively express MCSF, and its receptor MCSFR is encoded by c-fms. We examined whether MCSF might play a role in the proliferation of cultured DEL cells. [3H]Thymidine or 5-bromo-2-deoxyuridine (BrdU) incorporation was measured following the addition of recombinant MCSF or L929 cell supernatant (as a source of MCSF) to quiescent DEL cells. In DEL cells, serum-free L929 cell supernatant induced DNA synthesis in a dose-dependent manner, and such an effect could be blunted by pretreatment of L929 cell supernatant with anti-mouse MCSF antibody. In these cells, DNA synthesis could also be triggered in a dose-dependent manner by the addition of recombinant human MCSF (rh MCSF) or thrombin. These findings clearly show that MCSF influences DEL cell proliferation and suggest an autocrine loop activation. They indicate that MCSF plays an important role in the development of vascular lesions, which occur during atherosclerotic progression.     

巨噬细胞集落刺激因子与血管平滑肌细胞增殖

目的一些以血管平滑肌细胞（ＶＳＭＣ）增殖为特点的血管疾病，在病变部位常有巨噬细胞浸润。本研究巨噬细胞集落刺激因子（ＭＣＳＦ）在ＶＳＭＣ生长调节中的作用。方法实验采用培养大鼠主动脉ＶＳＭＣ，细胞增殖观察指标采用氚标胸腺嘧啶核苷掺入法，并用Ｎｏｒｔｈｅｒｎｂｌｏｔ技术测定原癌基因表达。结果（１）Ｌ９２９细胞上清液（富含ＭＣＳＦ）及重组ＭＣＳＦ以剂量依赖关系刺激氚标胸腺嘧啶核苷掺入；（２）ＶＳＭＣ在接受刺激后表达某些原癌基因，如ｃｆｏｓ、ｃｍｙｃ、ｅｒｇ１和ＪｕｎＢ；（３）凝血酶、ＰＤＧＦ、ｂＦＧＦ与ＭＣＳＦ在促增殖作用上具有协同作用。结论ＭＣＳＦ与其它生长因子协同作用，通过自分泌／旁分泌机制调控ＶＳＭＣ增殖，从而可能在血管病变的形成和进展中起重要作用     

热休克预处理对脂多糖所致RAW 264.7细胞迁移的影响

目的 观察热休克预处理对脂多糖所致巨噬细胞迁移的影响.方法 采用ELISA观察脂多糖预处理对RAW264.7巨噬细胞中肿瘤坏死因子α和白细胞介素1β释放的影响;采用趋化实验观察脂多糖处理对RAW264.7巨噬细胞迁移的影响;采用趋化实验观察热休克预处理对RAW264.7巨噬细胞迁移的影响.结果 用500μg/L脂多糖作用RAW264.7巨噬细胞1 h,肿瘤坏死因子α和白细胞介素1β的释放较对照组明显增加(P<0.05);用500μg/L脂多糖处理细胞24 h, RAW264.7巨噬细胞的迁移较对照组明显增加(P<0.05);给予细胞42.5℃热休克预处理1 h后,再予500μg/L脂多糖处理细胞,RAW264.7巨噬细胞迁移较单纯脂多糖处理组明显减少(P<0.05).结论 热休克预处理能抑制脂多糖所致RAW264.7巨噬细胞迁移.     

巨噬细胞内表达的人颗粒溶素对胞内结核分枝杆菌的杀菌活性

目的 构建携带人颗粒溶素(GLS)的真核表达质粒并探讨其表达后对巨噬细胞RAW264.7内结核分枝杆菌的杀菌能力.方法利用套式PCR从同种异型抗原激活的人CTL中扩增出GLS基因,并克隆入pBudCE4.1载体,构建重组质粒.将其转染至感染结核分枝杆菌H37Rv的巨噬细胞株RAW264.7中,套式PCR、免疫荧光检测GLS表达;转染96 h后裂解细胞作抗酸染色及菌落计数,检测GLS细胞内直接杀菌活性.数据行t或t'检验.结果成功构建携带GLS的真核表达质粒pBudCE4.1/GLS;在转染的已感染H37Rv巨噬细胞株RAW264.7中,从转录和翻译水平都检测到目标基因GLS的表达产物;转染96 h,佛波酯+pBudCF4.1/GLS组、佛波酯+pBudCE4.1组与未激活初始巨噬细胞荷菌量分别为(1.44±1.25)、(3.16±0.20)和(3.59±0.21)个,两两比较,差异均有统计学意义(t=2.403,t=2.854,均P＜0.05).结论携带人GLS的真核表达质粒在巨噬细胞表达后具有明显的杀菌活性,为进一步作为结核基因治疗疫苗的应用提供了实验依据.     

弓形虫P~(30)的克隆表达及其对巨噬细胞凋亡的影响

目的　获得能表达弓形虫P3 0 蛋白的小鼠巨噬细胞克隆并观察内源性表达的P3 0 蛋白对小鼠巨噬细胞凋亡的影响。方法　通过PCR扩增获得P3 0 基因片段 ,定向克隆到真核表达载体pcDNA3.1/Hygro(- )中 ,利用酶切、DNA序列分析鉴定阳性克隆 ,采用脂质体将重组质粒转染到RAW 2 6 4 .7巨噬细胞中 ,通过Hygromycin筛选和PCR、免疫组化鉴定 ,用流式细胞术DNA倍体分析计算稳定转染和瞬时转染P3 0 基因的巨噬细胞的凋亡率。结果　 1.PCR、酶切、连接的产物经电泳鉴定 ,均与预期设计相符合 ,DNA序列分析发现重组质粒中的目的基因序列与文献报道相符。2 .PCR和免疫组化鉴定发现转染P3 0 重组质粒的巨噬细胞能稳定地复制质粒并表达弓形虫P3 0 蛋白。 3.P3 0 稳定转染的细胞凋亡率均在 2 %左右 ,不同细胞之间没有区别。 4 .瞬时转染空载体和P3 0 重组质粒的巨噬细胞其凋亡率均在 6 %左右 ,没有区别。结论　 1.获得了含P3 0 基因的重组质粒以及能稳定表达弓形虫P3 0 蛋白的小鼠巨噬细胞克隆。2 .无论是稳定转染还是瞬时转染 ,均不能引起巨噬细胞的凋亡 ,说明内源性表达的弓形虫P3 0 蛋白对小鼠巨噬细胞的凋亡没有影响。     

Lipopolysaccharide (LPS)-reactive monoclonal antibodies fail to inhibit LPS-induced tumor necrosis factor secretion by mouse-derived macrophages

Murine monoclonal antibodies (MAbs) reactive with epitopes on the O-side chain, core oligosaccharide, or lipid A of Escherichia coli and Salmonella minnesota lipopolysaccharide (LPS) were evaluated for their ability to inhibit LPS-induced tumor necrosis factor (TNF) secretion by mouse-derived RAW 264.7 macrophages. As little as 50 ng of purified LPS or lipid A stimulated macrophages to produce TNF detectable as cytotoxic activity in an L-929 fibroblast assay. None of 13 MAbs (concentration range, 0.1-1,000 micrograms/mL) blocked LPS- or lipid A (0.025-0.1 micrograms/mL)-induced TNF secretion by RAW 264.7 cells. Rabbit antiserum to synthetic lipid A also failed to block lipid A-induced TNF activity. Similar negative results were obtained when intact bacteria or membrane vesicles were used as TNF inducers. In contrast, polymyxin B, but not the less hydrophobic polymyxin B nonapeptide, produced almost complete inhibition of macrophage TNF secretion induced by LPS, lipid A, membrane vesicles, and intact bacteria. Thus, antibody reactivity with predominantly hydrophilic elements of LPS or lipid A may not affect hydrophobic interactions between lipid A and target cell membranes necessary and sufficient for the induction of TNF. These findings raise doubts concerning the existence of true endotoxin-neutralizing antibodies.