小鼠腹腔巨噬细胞非特异性阳离子外向电流的基本特性

小鼠腹腔巨噬细胞非特异性阳离子外向电流的基本特性张晓峰丰美福（中国科学院动物研究所生物膜与膜生物工程国家重点实验室北京１０００８０）吴才宏周培爱（北京大学生命科学学院北京１００８７１）近十几年来,在免疫细胞上发现了几种受体门控的非特异性阳离子通道,它...     

双歧杆菌对裸鼠腹腔巨噬细胞NO形成的调节作用

给裸小鼠腹腔注射青春型双歧杆菌,每天一次,连续５天,以Ｇｒｉｅｓ试剂测定了裸鼠腹腔巨噬细胞分泌ＮＯ的含量。结果表明：双歧杆菌注射组其腹腔巨噬细胞产生ＮＯ的量显著高于对照组,具有显著的统计学意义（Ｐ＜００１）。提示青春型双歧杆菌可激活巨噬细胞,使之产生一定量的ＮＯ,ＮＯ在介导双歧杆菌的多种生理功能方面起重要作用     

冬虫夏草粗多糖诱导小鼠腹腔巨噬细胞产生TNF-α的作用

研究了 3种冬虫夏草提取的粗多糖对小鼠腹腔巨噬细胞产生肿瘤坏死因子的作用。结果表明野生冬虫夏草、冬虫夏草菌丝体和拟青霉子实体的粗多糖在体外都能诱导小鼠腹腔巨噬细胞产生TNF α ,但野生冬虫夏草在体外诱导小鼠腹腔巨噬细胞产生TNF α的强度比冬虫夏草菌丝体和拟青霉子实体高 ,拟青霉子实体又比冬虫夏草菌丝体强     

乳酸杆菌对小鼠腹腔巨噬细胞活化作用及细胞毒作用的初步研究

本文研究了乳酸杆菌对小鼠腹腔巨噬细胞（ＰＥＭ）的活化作用及其细胞毒作用。实验结果表明：腹腔注射乳酸杆菌后,ＰＥＭ体外细胞毒作用及体内细胞毒作用增强,酸性磷酸酶含量及非特异性酯酶含量增加、ＴＮＦ—α释放量增加。电镜显示：ＰＥＭ体积增大、伪足增多、线粒体、溶酶体等细胞器数量增多。本研究表明：乳酸杆菌具有较强的活化ＰＥＭ的作用。     

长爪沙鼠腹腔巨噬细胞对流行性出血热病毒的敏感性

本文就长爪沙鼠腹腔巨噬细胞(MΦ)对流行性出血热病毒(EHFV)的敏感性进行了试验,用3株野鼠型(A9、R3、76-118)和1株家鼠型EHFV(R22),并用对EHFV敏感的Vero-E6细胞作对照,结果沙鼠腹腔巨噬细胞感染EHFV后,第1代第8天前即可查见明显的特异性荧光,第16天左右达高峰,病毒滴度≥10~(-7.5),与Vero-E6细胞比较,培养上清液的病毒滴度,沙鼠MΦ较Vero-E6细胞高1～4个对数,当感染病毒量很低时,在Vero-E6细胞内测不出特异性抗原,而在沙鼠MΦ内病毒仍可繁殖,滴度达10~(-5.(?)),中和试验、间接免疫荧光检测和荧光阻断试验均证明,沙鼠MΦ内繁殖的病毒确实为EHFV。     

红毛五加多糖不同组分对小鼠腹腔巨噬细胞免疫功能的研究

本文研究红毛五加多糖不同组分(AHP-I、AHP-II、AHP-III)对小鼠腹腔巨噬细胞免疫调节功能的影响,为进一步阐明红毛五加多糖对小鼠免疫调节作用机制奠定基础。采用不同浓度的3种多糖组分作用于小鼠腹腔巨噬细胞,测定其对巨噬细胞吞噬中性红、释放NO能力、分泌IL-6、TNF-α、IL-1β水平的影响。最后结果是红毛五加多糖的3种不同组分对小鼠免疫细胞有不同的刺激能力。其中,AHP-II可极其显著地增强吞噬细胞的吞噬功能,促进其合成NO,促进巨噬细胞细胞因子的分泌。因此红毛五加多糖能激活小鼠腹腔巨噬细胞,其中,AHP-II是最重要的作用组分。     

腹腔巨噬细胞促进传代K562细胞系的体外增殖

在实验过程中经常遇到所要应用的靶细胞或效应细胞生长状态欠佳,如果使用这样的细胞作实验,其结果不准确,不可靠。在制备鼠-鼠杂交瘤的研究中,腹腔巨噬细胞可以支持1—10个抗体形成细胞在体少生长并分泌单克隆抗体。故我们将生长状态较差的K_(562)细胞(图1)与昆明小鼠腹腔巨噬细胞共培养7天(图2)。图1显示细胞折光性不良、形态不整存     

小鼠腹腔巨噬细胞的分离与培养

目的:建立一种小鼠腹腔巨噬细胞分离的简便方法,为低强度激光照射对巨噬细胞功能的影响研究提供实验细胞.方法:以无血清的RPMI-1640培养液灌洗小鼠腹腔,分离获取小鼠腹腔巨噬细胞,在含10%小牛血清的RPMI-1640培养液中培养.采用倒置显微镜观察细胞形态,台盼蓝染色计算存活率,瑞氏染色计算纯度.结果:获得高纯度的巨噬细胞,具备巨噬细胞的形态特征.结论:本法是一种简便实用的分离小鼠腹腔巨噬细胞的方法.     

异质性荧光染色的巨噬细胞功能研究──Ⅱ．异质性荧光染色的巨噬细胞过氧化物酶活性

细胞化学研究文献报导,巨噬细胞中过氧化物酶的活性有明显的差异,只有部分巨噬细胞过氧化物酶呈阳性反应。本文用淀粉、白喉类毒素和卡介苗分别诱导和活化小鼠腹腔巨噬细胞,进行过氧化物酶反应,并以７６９０－Ｘｕ荧光染液复染后证明,酶反应阳性细胞呈蓝色荧光,而酶反应阴性细胞为淡蓝绿色和黄色荧光。实验表明,过氧化物酶阳性的巨噬细胞是分化程度低的幼稚细胞,因此,过氧化物酶的活性可作为低分化的巨噬细胞的一种标志酶。同时,本文用免疫荧光单克隆抗体间接染色法观察了三种物质诱导和活化的异质性荧光染色的巨噬细胞的分泌功能。     

M-CSF与L929细胞培养上清诱导分化骨髓巨噬细胞的差异分析

[目的]通过比较常用的M-CSF与L929细胞培养上清两种诱导方式获得骨髓巨噬细胞(BMDM)的形态及基本生物学功能,为选择制备BMDM的方法提供实验依据。[方法]用两种方式诱导小鼠骨髓获取BMDM,然后分别利用Zeiss荧光显微镜和流式细胞仪比较BMDM的细胞形态及纯度;通过吞噬实验、杀菌实验以及液相芯片技术(liquid chip)比较两组BMDM的生物学功能;同时将上述结果与体内分化成熟的腹腔巨噬细胞相比较。[结果]两组诱导分化成熟的BMDM均呈现不规则形或梭形;BMDM的纯度分别为M-CSF组98.6%,L929上清组99.6%;腹腔细胞贴壁后巨噬细胞的纯度是98.8%;吞噬实验、杀菌实验两组之间及其与腹腔巨噬细胞相比无显著差异;炎症因子释放水平检测中3组细胞存在IL-6或TNF-α某些时间点释放水平差异。[结论]M-CS与L929上清诱导方式所获得BMDM的形态相似,纯度相近(98.6%比99.6%),吞噬细菌能力无显著统计学差异(21.31±5.83比26.10±6.11,t=-0.745,P0.05),杀菌能力无显著差异(杀菌30min:36.41%±4.21%比39.53%±6.75%;60min:44.35%±6.75%比45.27%±1.96%,两者均P0.05),但是炎症因子释放的某些时间点有差异(TNF-α的释放在LPS刺激后24 h M-CSF组高于L929组:549.92±412比271.47±432,t=-0.34,P0.05),应根据实验目的选择获取BMDM的诱导方式。     

氧化修饰低密度脂蛋白对巨噬细胞一氧化氮合酶基因表达的影响

氧化修饰LDL(OX-LDL)可抑制脂多糖(LPS)诱导的巨噬细胞NO释放, 而正常(N-LDL)和乙酰化LDL(AC-LDL)则没有抑制作用.OX-LDL对NO释放的抑制作用随LDL修饰程度的升高而增强,且具有浓度和时间效应.狭缝杂交结果显示OX-LDL处理可使LPS诱导的巨噬细胞NOS mRNA含量下降,提示OX-LDL对NO释放的抑制作用可能发生在转录水平.     

Vasoactive Intestinal Peptide (VIP) Inhibits Substrate Adherence Capacity of Rat Peritoneal Macrophages by a Mechanism That Involves cAMP

In this study, vasoactive intestinal peptide (VIP) is shown to inhibit substrate adherence capacity of rat peritoneal macrophages. The inhibitory response occurred in the 0.1-1, 000 nM range of VIP concentrations and it was a time-dependent process. At 15 min, half maximal inhibition (ICw) was obtained at 0.37 ± 0.26 nM and maximal inhibition (53.8%) at 10^?6 M VIP. The inhibitory effect of VIP was correlated with the stimulation by this peptide of cyclic AMP (cAMP) production in rat peritoneal macrophages. Moreover, agents that inhibited VIP-stimulated cAMP production, such as the VIP-antagonist [4-Cl-D-Phe₆ Leu₁₇]-VIP and somatostatin, also decreased the inhibitory effect of VIP on substrate adherence capacity of macrophages. On the contrary, the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX) and the lipid-soluble derivative of cAMP N⁶ 2′-O-dibutyryl cAMP (Bu-cAMP) inhibited the adherence of macrophages to substrate and potentiated the inhibitory action of VIP. These results demonstrate that VIP inhibits substrate adherence capacity of rat peritoneal macrophages by a mechanism that involves cAMP, and show, for the first time, an action of VIP on the function of peritoneal macrophages.     

Vasoactive Intestinal Peptide (VIP) Inhibits Substrate Adherence Capacity of Rat Peritoneal Macrophages by a Mechanism That Involves cAMP

In this study, vasoactive intestinal peptide (VIP) is shown to inhibit substrate adherence capacity of rat peritoneal macrophages. The inhibitory response occurred in the 0.1-1, 000 nM range of VIP concentrations and it was a time-dependent process. At 15 min, half maximal inhibition (ICw) was obtained at 0.37 ± 0.26 nM and maximal inhibition (53.8%) at 10^-6 M VIP. The inhibitory effect of VIP was correlated with the stimulation by this peptide of cyclic AMP (cAMP) production in rat peritoneal macrophages. Moreover, agents that inhibited VIP-stimulated cAMP production, such as the VIP-antagonist [4-Cl-D-Phe₆ Leu₁₇]-VIP and somatostatin, also decreased the inhibitory effect of VIP on substrate adherence capacity of macrophages. On the contrary, the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX) and the lipid-soluble derivative of cAMP N⁶ 2'-O-dibutyryl cAMP (Bu-cAMP) inhibited the adherence of macrophages to substrate and potentiated the inhibitory action of VIP. These results demonstrate that VIP inhibits substrate adherence capacity of rat peritoneal macrophages by a mechanism that involves cAMP, and show, for the first time, an action of VIP on the function of peritoneal macrophages.     

Sporozoites of Mammalian Malaria: Attachment To, Interiorization and Fate Within Macrophages

Sporozoites of Plasmodium berghei and Plasmodium knowlesi, incubated in normal serum readily interact with peritoneal macrophages of mice or rhesus monkeys, respectively. Interiorization of the sporozoite requires that both serum and macrophages be obtained from an animal susceptible to infection by the malaria parasite. Serum requirements for sporozoite attachment to the macrophage are less specific. Phagocytosis is not essential for the parasites to become intracellular. Our findings indicate that active penetration of the sporozoites into the macrophages does occur.     

Distinct Features of Brain-Resident Macrophages: Microglia and Non-Parenchymal Brain Macrophages

Tissue-resident macrophages play an important role in maintaining tissue homeostasis and innate immune defense against invading microbial pathogens. Brain-resident macrophages can be classified into microglia in the brain parenchyma and non-parenchymal brain macrophages, also known as central nervous system-associated or border-associated macrophages, in the brain-circulation interface. Microglia and non-parenchymal brain macrophages, including meningeal, perivascular, and choroid plexus macrophages, are mostly produced during embryonic development, and maintained their population by self-renewal. Microglia have gained much attention for their dual roles in the maintenance of brain homeostasis and the induction of neuroinflammation. In particular, diverse phenotypes of microglia have been increasingly identified under pathological conditions. Single-cell phenotypic analysis revealed that microglia are highly heterogenous and plastic, thus it is difficult to define the status of microglia as M1/M2 or resting/activated state due to complex nature of microglia. Meanwhile, physiological function of non-parenchymal brain macrophages remain to be fully demonstrated. In this review, we have summarized the origin and signatures of brain-resident macrophages and discussed the unique features of microglia, particularly, their phenotypic polarization, diversity of subtypes, and inflammasome responses related to neurodegenerative diseases.     

大剂量地塞米松快速高效诱导巨噬细胞凋亡

运用透射电镜、DNA琼脂糖凝胶电泳、流式细胞术、原位末端标记法(TUNEL)染色等技术,检测了大剂量地塞米松处理小鼠腹腔巨噬细胞的各种变化.结果显示,大剂量地塞米松处理的巨噬细胞发生胞体皱缩、染色质凝聚、胞质浓缩;TUNEL染色呈阳性;0.5 h后DNA凝胶电泳即呈梯状条带;流式细胞术作周期分析出现凋亡峰等明显的凋亡特征.并且随处理时间延长凋亡率升高.结果表明,大剂量地塞米松快速、高效诱导小鼠腹腔巨噬细胞凋亡.     

Effect of Polysaccharides and Human Plasma Lipoproteins on the Secretion of Cystatin C by Peritoneal Macrophages from Normal and Tumor Bearing Mice

Intact peritoneal macrophages in vitro secreted the cysteine proteinase inhibitor cystatin C. Polysaccharides stimulated cystatin C secretion: lipopolysaccharide < carboxymethylated beta-D-glucan < sulfoethylated beta-D-glucan. Human plasma low-density- (LDL) and high-density lipoproteins (HDL) are still more potent inducers of cystatin C secretion by macrophages. Peritoneal macrophages from mice with experimental HA-1 hepatoma compared to those from intact mice secreted more cystatin C with maximum polysaccharide-stimulated secretion after 30 min of incubation. LDL and HDL induced cystatin C secretion by tumor macrophages also.     

异丙肾上腺素对小鼠腹膜巨噬细胞分泌及吞噬功能影响的研究

目的:研究异丙肾上腺素对脂多糖诱导BALB/C小鼠腹膜巨噬细胞分泌TNF-α,IL-10及吞噬功能的影响.方法:分别以10μM、100μM和300μM异丙肾上腺素加入BALB/C小鼠腹膜巨噬细胞培养液,2h后用脂多糖(LPS,10μg/mL)刺激,孵育6h后以ELISA法测定上清中TNF-α水平,24h后测定上清中IL-10的含量,并观察腹膜巨噬细胞时中性红吞噬能力的变化.结果:异丙肾上腺素使BALB/C小鼠腹膜巨噬细胞分泌TNF-α降低,IL-10分泌增加,增加巨噬细胞对中性红的吞噬能力.结论:异丙肾上腺素能调节巨噬细胞分泌功能及吞噬功能,使炎性因子分泌减少,抑炎因子分泌增多,增强巨噬细胞吞噬中性红的能力.     

A Flow Cytometric Assay Reveals a Suppression of Phagocytosis by Rabbit Defensin NP-3A in Mouse Peritoneal Macrophages

Phagocytosis of fluorescein isothiocyanate (FITC)-labeled polystyrene microparticles by peritonea] macrophages from thioglycollate-elicited mice was examined by means of flow cytometry (FCM). This assay revealed that rabbit defensin NP-3A suppressed the phagocytosis in a dose-dependent manner. The present results suggest that NP-3A released from neutrophils is one of the mediators which modulates the activity of macrophages in response to infection.