2.80 胃粘膜肥大细胞和5-HT阳性细胞在感觉过敏的功能性消化不良病人中的改变

目的功能性消化不良(FD)病人除了有胃动力障碍外,近来还发现存在着内脏感觉过敏.肥大细胞在胃肠道含量丰富,我们的初步研究已证实其在FD的发病中有一定作用.本研究的目的是探讨在FD病人中胃粘膜肥大细胞与胃感觉过敏之间的关系.方法 23例FD患者(男9例,女14例,年龄32±8.8岁)和15例健康自愿者(男6 例,女9例,年龄29.6±7.9岁)为受检者.运用电子恒压器检测近端胃感觉阈值,在扩张刺激胃前后分别取近端胃粘膜组织活检.使用SABC方法染色肥大细胞及5-HT阳性细胞,采用病理图像分析系统获得肥大细胞和5-HT阳性细胞的平均光密度值(OD).结果 1. FD患者近端胃机械感觉阈值明显低于正常人.有11例(47.8%)的F D患者胃机械感觉阈值低于正常人的最低值,我们称之为感觉过敏组.2.感觉过敏组FD患者近端胃粘膜肥大细胞数(19.5±5.6/HSP)明显高于感觉正常组FD患者(16.3±5.5/HSP, P＜0.05)和正常人(15.9±5.6/HSP,P＜0.05).3.扩张刺激前肥大细胞和5- HT阳性细胞的OD值在正常人、感觉正常组FD患者、感觉过敏组FD患者之间无差异;扩张刺激后感觉过敏组FD患者的肥大细胞OD值减少百分比与感觉正常组FD患者和正常人相比明显增加 (感觉过敏组FD患者∶感觉正常组FD患者∶正常人为16.2%±3.87%∶7.41±2.23%∶7.2 1%±3.25%,ANOVA,P＜0.05),感觉过敏组FD患者5-HT阳性细胞的OD值减少百分比与感觉正常组FD患者和正常人相比亦明显增加(感觉过敏组FD患者∶感觉正常组FD患者∶正常人为22.6%±3.24%∶7.72%±3.15%∶5.12%±2.11%,ANOVA, P＜0.05).结论 1.感觉正常组FD患者近端胃粘膜肥大细胞和5-HT阳性细胞无改变.2 .感觉过敏组FD患者近端胃粘膜肥大细胞数量增多,扩张刺激后肥大细胞和5-HT阳性细胞脱颗粒增多,提示它们可能与FD患者胃感觉异常有关.     

成骨细胞响应机械刺激的力化学转导机理

应力环境可调节骨组织的生长、吸收和重建。在细胞水平上,机械刺激可以影响成骨细胞的生理活性,如增殖、碱性磷酸酶活性、骨钙素合成等。力转导是将生物物理力转化为细胞的生化响应的过程,它是许多生理功能的基础,早期响应基因(c- fos,c- jun) ,第二信使系统(Ca2 ,NO,c AMP) ,力敏感阳离子通道等都参与了成骨细胞响应机械刺激的力化学转导过程     

一种大鼠腹腔肥大细胞分离方法

目的介绍一种密度梯度离心分离大鼠腹腔肥大细胞(RPMC)的技术。方法比较改良方法和原方法2种分离的细胞量、活性及纯度。台盼蓝染色检测细胞存活率;甲苯胺蓝染色检测肥大细胞纯度。将弃去密度梯度离心上液2 m(lA方法)与2.5 m(lB方法)比较,以分析弃去的细胞液的量与肥大细胞纯度的关系。用C48/80刺激分离的RPMC,检测细胞释放的β-氨基己糖苷酶和组胺以确定分离的RPMC脱颗粒活性;透视电镜扫描观察细胞。结果改良方法分离的细胞显著多于原方法分离的细胞数;两种方法分离的细胞存活率都在99%以上;肥大细胞纯度都在90%以上。C48/80刺激RPMC,分离的细胞释放大量的β-氨基己糖苷酶和组胺,高于空白对照组。电镜观察细胞具有明显的肥大细胞形态。结论改良的Percoll密度梯度离心分离大鼠腹腔肥大细胞的方法可行。     

几种细胞因子对肥大细胞生长的影响

应用体外培养的小鼠骨髓肥大细胞作为实验模型，观察了几种细胞因子对肥大细胞生长的作用，证明小鼠造血干细胞因子（ｍＳＣＦ）可支持经培养已生长为近乎纯一的肥大细胞，但不能刺激原代培养的肥大细胞启动生长。白介素－３（ＩＬ－３）和促红细胞生成素（Ｅｐｏ）也能支持肥大细胞的生长。     

组胺对肥大细胞激活的研究

组胺是肥大细胞和嗜碱性粒细胞内组氨酸脱羧基后产生的一种胺类物质 ,是肥大细胞和嗜碱性粒细胞脱颗粒的标志物 ,本研究通过组胺对人大肠肥大细胞进行体外干预 ,探讨组胺对人肥大细胞类胰蛋白酶释放的调节和可能的机制。结果发现 :1.组胺可诱导人大肠肥大细胞以非剂量依赖性的方式释放类胰蛋白酶 ,引起最大释放量的组胺浓度为 10 0ng/ml,比基础分泌量多出 3.5倍。组胺浓度高至 10 0 0ng/ml和 10 0 0 0ng/ml时 ,其诱导类胰蛋白酶的释放量介于组胺浓度为 10ng/ml与 10 0ng/ml之间。浓度为 10ng/ml的组胺对人肥大细胞的刺激强度与 10 μg/ml的…     

IL-12调节蛋白酶激活受体在肥大细胞的表达

目的:探讨IL-12对肥大细胞蛋白酶激活受体(PARs)表达的影响.方法:不同浓度IL-12刺激肥大细胞后, 在不同时间点, 用实时定量逆转录聚合酶链式反应(q-RT-PCR)和流式细胞术(FCM), 在mRNA水平和蛋白水平检测PAR-1、 PAR-2、 PAR-3和PAR-4在肥大细胞P815表面和胞内的表达情况.结果:IL-12下调肥大细胞膜表面PAR-2蛋白的表达, 上调肥大细胞膜表面和胞质内PAR-4蛋白的表达, IL-12抗体的应用可阻断IL-12对肥大细胞PARs蛋白表达的影响;IL-12上调肥大细胞PAR-1、 3、 4 mRNA的表达, 下调PAR-2 mRNA的表达.结论:IL-12在过敏性炎症反应中的调节作用可能与IL-12调节肥大细胞PARs表达有关.     

肥大细胞

近年由于IgE的发现,对肥大细胞(mastcell)的机能有所了解。IgE是引起肥大细胞脱颗粒的刺激之一,仅在肥大细胞和嗜碱性粒细胞表面存在IgE受体,机体的IgE大部分固着于肥大细和嗜碱性粒细胞的IgE的FC受体上。此类细胞是即刻型变态反应的细胞。由于肥大细胞含有组胺,故在炎症时起重要作用。对甲苯胺兰染色呈异染现象是由于肥大细胞含肝素所致,体内肝索的大部分存在于肥大详胞。     

IL-9基因定位于小鼠第13号染色体和人第5号染色体上

肥大细胞生长因子P40是一种32000～39000糖蛋白。它能刺激体外辅助性T细胞生长,增强肥大细胞活性,并具有巨核母细胞白血病生长因子活性,被命名为IL-9。     

蛋白酶激活受体2介导肥大细胞IL-4分泌

目的:探讨蛋白酶激活受体2(Proteinase activated receptor-2,PAR-2)的激活对肥大细胞P815介质分泌的影响.方法:肥大细胞培养后用PAR-2激动肽,胰蛋白酶、类胰蛋白酶、弹性蛋白酶结合PAR-2拮抗肽激发肥大细胞,收集上清并用ELISA方法检测组胺、IL-4和IL-6的水平.结果:PAR-2激动肽、胰蛋白酶、类胰蛋白酶以浓度依赖的方式促进肥大细胞P815分泌IL-4.PAR-2拮抗肽能阻断胰蛋白酶和类胰蛋白酶引起的肥大细胞IL-4分泌(P<0.05).胰蛋白酶、类胰蛋白酶以及PAR-2激动肽对肥大细胞IL-6的分泌和组胺释放无明显影响.弹性蛋白酶对肥大细胞IL-4、IL-6分泌及组胺释放功能无影响.结论:PAR-2的激活介导肥大细胞P815分泌IL-4;胰蛋白酶和类胰蛋白酶刺激肥大细胞IL-4分泌的发现为肥大细胞激活的自我放大机制提供了新的理论依据.     

组胺对人结肠肥大细胞类胰蛋白酶释放的调节作用

探讨组胺对人结肠肥大细胞类胰蛋白酶释放的调节作用。经酶消化后获取人结肠组织肥大细胞,激发后行多种干预实验。用酶联免疫吸附试验法测定类胰蛋白酶。结果发现组胺可诱导人结肠肥大细胞释放类胰蛋白酶,浓度为100μg/L时组胺释放量最大,为基础值的3．5倍。浓度为10μg/L时对人肥大细胞的刺激强度与10mg/L的抗IgE抗体相似。组胺的作用从加样后10s开始,5min后完成。百日咳毒素和抗霉素A联合2．脱氧-D-葡萄糖可显著抑制组胺诱导人结肠肥大细胞释放类胰蛋白酶。100及1000μg/L的组胺与抗IgE抗体或离子载体钙同时加入细胞中,诱导类胰蛋白酶释放的能力低于组胺单独作用组。结论为组胺可激活人结肠肥大细胞,还以自身放大机制调节肥大细胞的脱颗粒过程。     

Radiation-resistant IgE-secreting cells in the mouse: susceptibility to suppressor T cells

Mast cells can be dispersed from human tonsils by a mechanical procedure or by the use of proteolytic enzymes. The latter procedure does not always increase cell yield and may adversely affect the mast cell response to secretory stimuli. Mechanically dispersed tonsillar mast cells secrete histamine in response to stimulation with anti-IgE and calcium ionophore A23187 by a calcium-dependent and energy-dependent mechanism. With compound 48/80 and basic polyamines, histamine release only occurs at cytotoxic concentrations. Functionally, human tonsillar mast cells are similar to adenoidal and lung mast cells.     

Spatiotemporally and mechanically controlled triggering of mast cells using atomic force microscopy

Mast cells are thought to be sensitive to mechanical forces, for example, coughing in asthma or pressure in “physical urticarias.” Conversion of mechanical forces to biochemical signals could potentially augment antigenic signaling. Studying the combined effects of mechanical and antigenic cues on mast cells and other hematopoietic cells has proven difficult. Here, we present an approach using a modified atomic force microscope cantilever to deliver antigenic signals to mast cells while simultaneously applying mechanical forces. We developed a strategy to concurrently record degranulation events by fluorescence microscopy during antigenic triggering. Finally, we also measured the mechanical forces generated by mast cells while antigen receptors are ligated. We showed that mast cells respond to antigen delivered by the atomic force microscopy cantilever with prompt degranulation and the generation of strong pushing and pulling forces. We did not discern any relationship between applied mechanical forces and the kinetics of degranulation. These experiments present a new method for dissecting the interactions of mechanical and biochemical cues in the signaling responses of immune cells.     

Histamine release from dispersed human intestinal mast cells

To study the human intestinal mast cell of children and adults, we combined a sensitive glassfibre-based histamine assay with the enzymatic and mechanical dispersion of surgical specimens or mucosal biopsies. The method yields between 1.2 x 10(3) to 4.6 x 10(3) mast cells/mg tissue constituting 1.2% to 5.3% of total cell count. The mast cell yield, however, depends on the intestinal tissue specimen used for dispersion. Aliquots containing 1500 mast cells per sample are sufficient for measuring significant amounts of histamine (greater than or equal to 0.15 ng histamine per sample), thus making it possible, to carry out approximately 75 tests for four mucosal biopsies of 10 mg each. The intestinal mast cell releases histamine in a dose-dependent manner on challenge with anti-IgE (6-600 U/ml), ionophore A23187 (0.25-1.0 microM), and Concanavalin A (0.7-25.0 micrograms/ml). The histamine release shows interindividual variation with a net histamine release between 0 to 2.5 ng/samples dependent on the secretatogue. In general, it is not necessary to passively sensitize the mast cells to obtain a sufficient histamine release response to anti-IgE challenge, indicating the presence of intact and functional cell-bound IgE. However, it is shown that four of 10 non-atopic intestinal mast cell samples could be passively sensitized with human plasma containing either mite- or grass-specific IgE without stripping off the IgE first. This indicates the presence of free and preserved Fc-receptors on the dispersed mast cells in some subjects. In addition, it is found that the phorbolester TPA increases the histamine release response to A23187 and turns anti-IgE non-responding mast cells into responding mast cells, but TPA alone at 2 to 16 ng/ml has no histamine releasing effect. In patients with anti-IgE responding mast cells no additional effect of TPA is seen. Finally, no substantial differences between mast cells of children and adults are demonstrated.     

Mast cell tryptase as a mediator of hyperresponsiveness in human isolated bronchi

BACKGROUND: Although the role of mediators and cytokines produced by mast cells is well established in asthmatic bronchial inflammation, the contribution of mast cell-derived proteases to the development of hyperresponsiveness remains unclear. There have been reports indicating that tryptase alters the mechanical activity of animal airway smooth muscle or spontaneously sensitized human isolated airways. OBJECTIVE: The aim of this study was to analyse the effect of purified mast cell tryptase on non-sensitized human isolated bronchi. METHODS: Both central and peripheral bronchi, dissected from lung specimens obtained at thoracotomy, were studied in terms of both mechanical activity i.e. isometric contraction in response to a variety of agonists and distribution of inflammatory cells i.e. immunohistochemistry. RESULTS: In both proximal and distal bronchi, the reactivity to histamine was significantly increased by a previous incubation in the presence of 1 microg/mL of tryptase (increase in maximal force, DeltaFmax was 12.1 +/- 3.8%, and 8.8 +/- 3.1%, respectively). This effect of tryptase on histamine-induced contraction was completely abrogated in the presence of the protease inhibitor benzamidine (100 micromol/L). Histological examination of specimens exposed to tryptase demonstrated an increase in mast cell number within the subepithelial tissue whereas mast cell numbers in the epithelial layer concomittently decreased. CONCLUSION: These results indicate that human mast cell tryptase alters the contractile response of non-sensitized human isolated bronchi and that this alteration is accompanied by a change in the mast cell distribution within the airway wall.     

Mucosal mast cells as a component of the inflammatory response to lower-urinary tract infection

Globule cells have been observed in mucosae for many years. Recently, a subpopulation of these globule cells in the intestinal mucosa of man and rodents have been identified as unique mast cells. In this communication, intraepithelial globule cells and some lamina-propria mast cells found in the normal rat urinary-bladder wall have been characterized as mucosal mast cells, similar to intestinal mast cells, and differentiated morphologically and histochemically from rat peritoneal mast cells. The number of mast cells in the bladder wall increased significantly during various bladder manipulations, including mechanical trauma, parasitic infestation and bacterial infection. The origin and function of the mucosal mast cells remains unknown.     

Mast cells in allergy: innate instructors of adaptive responses

The function of mast cells as effector cells in allergy has been extensively studied. However, increasing insight into mast cell physiology has revealed new mast cell functions and has introduced mast cells as key players in the regulation of innate as well as adaptive immunity. For example, mast cells have recently been found to express Toll-like receptors (TLRs), which enable them to participate in the innate immune response against pathogens. Furthermore, mast cells have been reported to interact with B cells, dendritic cells and T cells and thereby modulate the direction of an adaptive immune response. Finally, recent documentation that mast cells express functional MHC class II and costimulatory molecules and release immunologically active exosomes, has raised the possibility that mast cells also engage in (as yet) poorly understood antigen presentation functions. In this review, we explore the hypothesis that mast cells serve as central mediators between innate and adaptive immunity, rather as pure effector cells, during allergic innate responses.     

Mast cells in innate immunity

Summary: Mast cells are known to be the main effector cells in the elicitation of the IgE-mediated allergic response. The specific location of mast cells within tissues that interface the external environment, and the extent of their functional capacity, including the ability to phagocytose and to produce and secrete a wide spectrum of mediators, have led investigators to propose a potential role for mast cells in innate immune responses. Certain microorganisms have been found to interact either directly or indirectly with mast cells. This interaction results in mast cell activation and mediator release which elicit an inflammatory response or direct killing leading to bacterial clearance. The in vivo relevance of these in vitro observations has been demonstrated by the use of complement-deficient and/or mast cell-deficient and mast cell-reconstituted mice. It thus has been shown that both C3 and mast cell- and tumor necrosis factor‐a-dependent recruitment of circulating leukocytes with bactericidal properties are crucial to a full response in certain models of acute infection. Modulation of mast cell numbers in vivo was also found to affect the host response against bacterial infection. Thus, mast cells do have a role in innate immunity in defined animal models of bacterial infection. Whether mast cells participate in innate immune responses in the protection of the human host against bacteria remains to be determined.     

Disparity in FcεRI-induced degranulation of primary human lung and skin mast cells exposed to adenosine

Inhaled and intravenously administered adenosine induces mast cell-mediated (histamine-dependent) bronchospasm in asthmatics without causing urticaria. A differential response to adenosine by human lung and skin mast cells is shown: low concentrations potentiate FcεRI-induced degranulation of human lung mast cells but not that of skin mast cells. Human lung mast cells were found to express ∼3-fold more A3AR messenger RNA (mRNA) than skin mast cells, suggesting the involvement of the G_i-linked A3AR. Indeed, the adenosine-induced potentiation was sensitive to inhibition by pertussis toxin and, furthermore, could be induced with an A3AR-specific agonist. This study reveals a previously unrecognized disparity in the response to adenosine by primary human mast cells from lung and skin that might explain why adenosine induces a pulmonary but not dermatologic allergy-like response in vivo. In addition, we identify the A3AR as a potentiating receptor of FcεRI-induced degranulation, thereby implicating it in the in vivo bronchoconstrictive response to adenosine in asthmatics.     

SDF-1 induces IL-8 production and transendothelial migration of human cord blood-derived mast cells

BACKGROUND: Mast cell numbers and expression of chemokines are known to increase in the context of angiogenesis and inflammation, but the mechanisms by which this occurs are not understood. Stromal-derived factor-1 (SDF-1) is an important chemokine in angiogenesis and cell migration. The effects of SDF-1 on human mast cells were examined. METHODS: Expression of the SDF-1 receptor CXC chemokine receptor 4 (CXCR4) on mast cells was examined by RT-PCR and flow cytometry. The ability of labeled cord blood-derived mast cells to migrate across HUVEC monolayers in response to SDF-1 was determined. The cytokine and chemokine responses of cord blood-derived mast cells to SDF-1 treatment over 24 h were examined by ELISA. RESULTS: Cord blood-derived human mast cells expressed the CXCR4 receptor for SDF-1 and migrated across HUVEC monolayers in response to this chemokine. Treatment of cord blood-derived mast cells with SDF-1 did not induce degranulation or the production of several cytokines but did induce a highly selective IL-8 response. CONCLUSION: Human mast cells can both migrate across vascular endothelium and produce the pro-angiogenic chemokine IL-8 in response to SDF-1. These responses may be important in angiogenic processes.     

Quantitative changes in mast cell populations after left ventricular assist device implantation

Mast cells have been implicated as important in tissue remodeling and fibrosis. We investigated the effect of mechanical ventricular unloading upon myocardial fibrosis and cardiac mast cell density in patients undergoing left ventricular assist device (LVAD) implantation. Paired myocardial tissue samples were obtained from 30 patients with end-stage cardiomyopathy at the time of LVAD implantation and at the time of removal and were compared with samples taken from donor hearts. Tissue sections were stained and quantitated for mast cells and myocardial fibrosis. Mast cell density (tryptase positive cells) in cardiomyopathy was higher than that in donor hearts (33.5 +/- 3.6 SEM cells/10 fields vs.15.2 +/- 2.0 SEM cells/10 fields respectively, p = 0.04) and was lower than LVAD supported hearts (33.5 +/- 3.6 SEM cells/10 fields vs. 49.8 +/- 5.7 SEM cells/10 fields respectively, p = 0.01). Mast cells are primarily localized in areas of increased interstitial fibrosis adjacent to myocardial cells and not vessels. There was statistically significant correlation between mast cells and interstitial collagen (p = 0.03) in patients before LVAD implantation that did not persist after mechanical support (p = 0.18). These results suggest that mechanical support with left ventricular assist devices induces an increase in mast cell number in the myocardium and an associated decrease in myocardial fibrosis. We believe these data demonstrate a dual role for cardiac mast cells in the increase in fibrosis in heart failure and the decrease after LVAD and its associated cardiac improvement.