单核-巨噬细胞的异质性及其对创面愈合的调控研究进展

单核吞噬细胞系统(MPS)主要由骨髓中单核细胞前体、外周血中的单核细胞、组织中的巨噬细胞和树突状细胞(DC)组成。其中,巨噬细胞和DC分布在机体的多个组织器官,发挥免疫监控和抵御病原体入侵的作用。然而,不同部位的这些细胞在组织形态和功能上存在一定差异,即使是同一器官中的巨噬细胞也不尽相同。皮肤中的单核-巨噬细胞主要包括表皮中的朗格汉斯细胞以及真皮中的巨噬细胞和真皮树突状细胞(dDC),它们共同参与调控创面愈合过程中的炎症反应、肉芽组织生成和组织重塑,达到促进创面愈合的作用。     

肉芽肿性松弛皮肤病--一种罕见皮肤T细胞淋巴瘤类型

目的：报道肉芽肿性松弛皮肤病临床病理学特征。方法：对其临床、组织病理学、免疫组化及分子生物学行为进行研究。结果；临床表现为腋窝及腹股沟巨大垂吊皮肤肿块伴四肢腹部红色斑丘疹。组织学呈弥漫性淋巴细胞、多核巨细胞及组织细胞浸润真皮及皮下组织、弹性纤维消失。免疫组化小淋巴细胞呈CD3＋、CD4＋、CD45RO＋、CD8－、CD15－、CD20－、CD30－、CD56－、TIA－1－和GranzymeB-；多核巨细胞及单核细胞呈lysozyme 、CD68 和Mac387-;S－100蛋白阳性细胞散在分布。 分子生物学检查示T细胞受体γ链基因重排。EBWER1／2原位杂交呈 阴性反应。结论：肉芽肿性松弛皮肤病是CD4＋辅助性T细胞淋巴瘤的罕见类型。     

弥漫性掌跖角皮症一家系四代13例

先证者（Ⅲ4）男，32岁，出生后1个月左右即发现双侧手掌足底侧缘发黄、增厚，以后逐渐加重弥漫至整个掌跖，皮损边缘有一线状红斑，皮损持续存在，皮肤增厚即粗糙愈来愈明显，有时出现裂口，对日常的生活及劳作有一定的影响，冬天较重，夏季稍有减轻，与外伤及日晒无关，不伴有明显的瘙痒。体格检查：双侧掌跖可见弥漫性角化增厚，对称分布，一些部位呈黄色，掌跖侧缘与正常皮肤有明显的分界线，指（趾）甲甲板稍增厚，四肢及躯干未见异常皮损，头发及肘膝部位皮肤均未见异常。足侧缘组织病理学检查：表皮明显角化过度，呈疣状增生，     

胎儿皮肤中CK20的表达及与Merkel细胞分布的相关性研究

目的研究不同发育时期胎儿皮肤表皮基底层中细胞角蛋白20（CK20）的分布、数量,以揭示Merkel细胞分布的规律。方法CK20是Merkel细胞特异而敏感的标记物,分别取42例不同胎龄（16—30周）胎儿掌、跖、胸、背部的全层皮肤,采用免疫组化SP法观察CK20。结果掌、跖部位CK20阳性细胞的数量显著高于胸、背部;胚胎16～20和21—25周显著高于26—30周。结论（1）表皮Merkel细胞的数量存在解剖部位的差异。（2）胚胎发育中期Merkel细胞的数量显著多于胚胎发育的晚期。（3）Merkel细胞在表皮突内的高表达可能与表皮突结构的形成有关。     

源于细菌CpG基序的寡核苷酸激活单核/巨噬细胞抗白血病效应的研究

目的： 研究源于细菌CpG基序的寡核苷酸激活单核/巨噬细胞抗白血病细胞的作用。方法: 用血细胞分离机从健康人外周血分离并诱导出单核-巨噬细胞，流式细胞仪检测细胞表面CD14分子和CD16分子表达状况。设计合成含CpG基序的寡核苷酸(CpG-ODN)和不含CpG基序的寡核苷酸（nonCpG-ODN）分别作用于单核/巨噬细胞, MTT法检测经寡核苷酸作用后,单核/巨噬细胞抗白血病K562细胞的效应, 用ELISA法检测其分泌细胞因子IL-12、TNF-α的表达。结果: 从健康人外周血分离并成功诱导出单核/巨噬细胞，证实CpG-ODN作用于单核/巨噬细胞,可显著增强单核/巨噬细胞体外抗白血病细胞的作用，同时能促进单核/巨噬细胞分泌细胞因子IL-12、TNF-α。结论: 源于细菌CpG-ODN可增强单核/巨噬细胞介导的抗白血病细胞作用，此为白血病免疫治疗提供了新的途径。     

血管树突状细胞在人主动脉粥样硬化早期病变中的分布

目的：探讨血管树突状细胞在人早期动脉粥样硬化(AS)病变中的分布模式。方法：人主动脉标本15例主要取自尸检和外科手术，常规连续切片，分别行HE及S100／CD1a免疫细胞化学染色，光镜下观察S100／CD1a阳性细胞分布情况。结果：15例HE染色标本中，2例正常，13例人动脉血管可见内膜的增厚及泡沫细胞等AS早期病理表现。9例S100／CD1a染色阳性，阳性率为69．2％。S100／CD1a阳性细胞分布在病变的内膜和外膜，外膜的S100／CD1a阳性细胞主要分布在滋养血管的周围。结论：在AS早期病变部位有血管树突状细胞的聚集，主要分布在病变血管的内膜和外膜，提示血管树突状细胞可能参与了AS早期的免疫反应。     

糖尿病模型皮肤创面中人真皮多能干细胞向表皮细胞的分化

背景：研究表明,干细胞分化为何种细胞与其所处的微环境密切相关。因此设想：人真皮多能干细胞处于皮肤创面的微环境中,可能具有向人皮肤细胞转化的潜能。 目的：探讨糖尿病皮肤创面微环境中的人真皮多能干细胞分化为表皮细胞的可能性。 方法：分离培养人真皮多能干细胞并制作糖尿病裸鼠模型皮肤创面,将标记5-BrdU的第3~5代人真皮多能干细胞以注射方式回植入糖尿病裸鼠创面组织周围,分别于注射后2,3周取材,常规石蜡包埋、连续切片,行BrdU和角蛋白免疫组织化学染色。 结果与结论：BrdU阳性细胞出现在表皮中,且连续切片中部分BrdU阳性细胞也同时表达角蛋白。说明在糖尿病创面愈合过程中,人真皮多能干细胞具有向表皮细胞分化的潜能。     

寻常型银屑病皮肤真表皮T细胞分类比较

目的：探讨寻常型银屑病（PV）的免疫发病机制。方法：应用链霉卵白素－过氧化物酶法（SP法），比较不同病期寻常银屑病皮肤真皮T细胞表型表达及真皮微血管E－选择素表达变化等情况。结果：（1）PV进展期皮损表皮CD4^ 细胞、CD8^ 细胞及CD45RO^ 细胞，真皮CD3^ 细胞、CD4^ 细胞、CD45RO^ 细胞及CLA^ 细胞高于静止期（P均＜0．05）；（2）静止期皮损表皮CD3^ 细胞，真皮CD3^ 细胞、CD4^ 细胞、CD8^ 细胞、CD45RO^ 细胞及CLA^ 细胞高于消退期皮损，消退期皮损表皮CD8^ 细胞高于静止期（P均＜0．05）；（3）进展期皮损周边无损害皮肤真皮CD4^ 细胞高于静止期皮周（P＜0．05）；（4）PV正常皮肤与正常皮肤与正常人皮肤的T细胞各严型间差异无显著性（P＞0．05）；（5）E－选择素表达强度与PV皮损及皮损周边无损害皮肤中浸润的CLA^ 细胞数量间的关联具有显著性（P均＜0．05）。结论：PV皮损中浸润的T细胞主要表达CLA、CD45RO，CD4^ 细胞可能在PV的发生及维持中起一定作用，但不能排除CD8^ 细胞的作用，E－选择素与CLA间的相互作用在介导T细胞的皮肤归巢中发挥重要作用。     

MV3和HaCaT细胞接种去表皮真皮组织上构建人工皮肤黑素瘤模型

背景：课题组曾成功的用MV3黑素瘤细胞和人角质形成细胞体外重建黑素瘤三维模型。 目的：利用MV3黑素瘤细胞和HaCaT细胞结合去表皮的真皮体外重建黑素瘤模型。 方法：MV3黑素瘤细胞和HaCaT细胞按照不同比例混合接种于人去表皮的真皮组织上,采用液下培养和空气-液面培养相结合技术进行培养,体外构建组织工程皮肤模型。对所构建的皮肤黑素瘤模型进行常规切片免疫组化观察。 结果与结论：苏木精-伊红染色显示MV3黑素瘤细胞在去表皮真皮表面成层分布或形成瘤灶,HaCaT细胞和瘤细胞混合生长,形成典型的表皮样结构。部分瘤细胞浸润到去表皮真皮浅层或内部,成瘤灶分布。CK10、CK-pan和S-100免疫组化染色显示阳性。随着MV3∶HaCaT细胞比例的增高,CK10,CK-pan由表层逐渐下移,由层状分布变为团块状分布,S-100蛋白染色则分层逐渐明显,部分区域成瘤状分布。结果可见利用MV3黑素瘤细胞和HaCaT细胞结合去表皮真皮体外可以构建皮肤黑素瘤模型。     

CypA/CD147及MMP-2在人慢性牙周炎牙龈组织中CD68+细胞上表达的研究

目的：观察亲环素A(CypA)、CD147及基质金属蛋白酶2(MMP-2)在人慢性牙周炎患者牙龈组织中CD68+细胞（单核细胞/巨噬细胞）的表达情况,探讨CD68+细胞中CypA、CD147及MMP-2在不同病变程度牙周炎牙龈组织中的表达及作用机制。方法：将60例受试者分为4组,每组15例,分别为：健康组及慢性牙周炎（轻、中、重）组。免疫荧光双染色（DIF）后荧光显微镜下分别观察牙龈组织中CD68-CypA、CD68-CD147及CD68-MMP-2双阳性细胞表达情况并计算单位面积双阳性细胞密度。结果：（1）各组牙龈组织中CD68-CypA、CD68-CD147及CD68-MMP-2双阳性细胞密度差异显著（P<0.01）;(2)各组牙龈组织中CD68-CypA双阳性细胞（r=0.959,P=0.000）、CD68-CD147双阳性细胞（r=0.948,P=0.000）及CD68-MMP-2双阳性细胞密度（r=0.945,P=0.000）与牙周炎的严重程度呈正相关;（3）各组牙龈组织中CD68-CypA与CD68-MMP-2双阳性表达正向相关（r=0.933,P=0.000）;CD...     

Human macrophage lectin specific for galactose/N-acetylgalactosamine is a marker for cells at an intermediate stage in their differentiation from monocytes into macrophages

We studied the expression of a human macrophage lectin specific for galactose/N-acetylgalactosamine (hMGL) during macrophage differentiation. The expression of hMGL during the in vitro differentiation induced by human serum was examined by immunostaining and Western blotting with a specific mAb, MLD-1, as well as with RT-PCR analysis. hMGL was detected on cells at an intermediate stage of differentiation. These cells were round, slightly larger in size (12.7 +/- 0.2 microm) than monocytes (9.8 +/- 0.1 microm) and expressed the macrophage marker CD14, but lacked the dendritic cell marker CD1a. The highest levels of expression occurred after 2-4 days of culture. At this time point, MLD-1 prominently stained 20-40% of the cells. Monocytes cultured for 16 h or fully differentiated monocyte-derived macrophages were negative or weak for hMGL expression. Similar transient expression was also observed during granulocyte macrophage colony stimulating factor- or macrophage colony stimulating factor-dependent macrophage differentiation. The lectin was characterized as a functional endocytic receptor for glycosylated macromolecules, since the uptake of carbohydrate polymers was partially inhibited by the addition of MLD-1. The distribution of hMGL(+) cells in normal human skin was found by immunostaining to be mainly in the upper dermis distant from vascular structures. More than 90% of the hMGL(+) cells were double stained with anti-CD68 mAb and constituted approximately 20% of the CD68(+) cells. We suggest that the dermal hMGL(+) cells are a subset of differentiated cells derived from monocytes and that hMGL is a unique marker for cells at an intermediate stage of macrophage differentiation.     

Recently infiltrating MAC387(+) monocytes/macrophages a third macrophage population involved in SIV and HIV encephalitic lesion formation

Monocytes/macrophages are critical components of HIV and SIV encephalitic lesions. We used in vivo BrdU labeling and markers specific to stages of macrophage differentiation or inflammation to define macrophage heterogeneity and to better define the role of macrophage populations in lesion formation and productive infection. Lesions were heterogeneously composed of resident macrophages (CD68(+)HAM56(+)), perivascular macrophages (CD163(+) CD68(+)MAC387(-)), and recently infiltrated MAC387(+) CD68(-)CD163(-) monocytes/macrophages. At 24 and 48 hours after BrdU inoculation, 30% of MAC387(+) monocytes/macrophages were BrdU(+), consistent with their being recently infiltrated. In perivascular cuffs with low-level SIV replication, MAC387(+) monocytes/macrophages outnumbered CD68(+) macrophages. Conversely, lesions with numerous SIV-p28(+) macrophages and multinucleated giant cells had fewer MAC387(+) monocytes/macrophages. The MAC387(+) cells were not productively infected nor did they express detectable CCR2, unlike perivascular macrophages. Overall, we found that the proportion of MAC387(+) cells tends to be higher than the proportion of CD68(+) macrophages in the brain of animals with mild encephalitis; the ratio was reversed with more severe encephalitis. These results suggest that development of SIV and HIV encephalitis is an active and ongoing process that involves the recruitment and accumulation of: i) nonproductively infected MAC387(+) monocytes/macrophages that are present with inflammation (potentially M1-like macrophages), ii) CD163(+) perivascular macrophages (consistent with M2-like macrophages), and iii) CD68(+) or HAM56(+) resident macrophages. The latter two populations are cellular reservoirs for productive infection.     

Immunohistochemical analysis of small plaque parapsoriasis: involvement of dendritic cells

Small plaque parapsoriasis (SPP) is one of the cutaneous T-cell lymphoproliferative disorders. The aim of the present study was to show the antigenic profile of a subset of dendritic cells and lymphocytes in SPP in comparison with normal cells to provide data on the role of these two cell types in the pathogenesis of SPP. Skin biopsy specimens of lesions were obtained from 8 patients with SPP. Biopsies of the healthy skin from 9 control individuals were also analyzed. Immunohistochemistry was performed on the frozen tissue sections to reveal binding of anti-HLA Class II, anti-CD1a, anti-CD4, anti-CD8, anti-CD44, anti-CD45, and anti-CD68 monoclonal antibodies. There was a statistically significant increase in the number of CD1a(+), Langerhans cells (LCs), HLA-DR-immunoreactive and, CD1a-positive dermal dendritic cells and CD68(+) macrophages in the SPP group (p=0.008, 0.008, 0.002 and <0.0009, respectively). The number of lymphocytes positive for CD4, CD8 and CD45 was significantly higher than normal in the SPP group (p=0.015, <0.0009 and <0.0009, respectively). Our study demonstrates that both peptide- and lipid-based antigens are involved in the persistent antigenic exposure in SPP. Dendritic cells play a pivotal role in SPP by presenting antigens by both LC and dermal dendritic cells via MHC Class II and CD1a molecules. The CD68(+) macrophages are thought to be involved in the immune response in this pathology as an antigen-presenting cell.     

Role of monocytes in anti-CD3-induced T-cell DNA synthesis: Effect of chloroquine and monensin on anti-CD3-induced human T-cell activation

Anti-CD3 monoclonal antibody (MoAb) induces proliferation of freshly isolated peripheral blood T cells only in the presence of monocytes/macrophages and requires binding of the Fc portion of antibody to monocytes/macrophages. In this investigation, we examined whether monocytes process anti-CD3 similar to any soluble antigen and present to T cells in context with HLA-DR to induce maximal DNA synthesis. Adherent monocytes were pulsed with anti-CD3 MoAb in the presence or absence of the lysozomotropic agents chloroquine and monensin, which are known to inhibit processing of soluble antigens, washed extensively, and then incubated with autologous T cells in the absence of soluble anti-CD3, and³H-thymidine incorporation and CD25 expression were measured. Both monensin and chloroquine inhibited anti-CD3-pulsed monocyte-induced T-cell DNA synthesis and CD25 expression in a dose-dependent manner. This inhibitory effect was not due to any loss in cell viability or the effect on the expression of HLA-DR on monocytes. Paraformaldehyde-fixed monocytes pulsed with anti-CD3 MoAb induced significantly less DNA synthesis, HLA-DR expression, and CD25 antigen expression on autologous T cells as compared to responses induced by unfixed anti-CD3-pulsed monocytes. The treatment of anti-CD3-pulsed monocytes with frame-work-specific anti-HLA-DR MoAb inhibited their capacity to induce T-cell DNA synthesis. These data suggest that monocytes, in addition to serving as the matrix for cross-linking, also process anti-CD3 MoAb and present to the T cells in the context of HLA-DR antigens to induce optimal DNA synthesis.     

严重急性呼吸综合征患者肾内免疫细胞的免疫组织化学观察

目的探讨严重急性呼吸综合征(SARS)患者肾内免疫细胞的免疫组织化学变化并对其作用进行分析。方法应用免疫组织化学方法显示6例SARS死亡患者和3例意外死亡者肾内免疫细胞,进行其形态观察,并采用图像分析系统进行定量分析。结果6例SARS患者肾内CD68 单核/巨噬细胞显著增多(P<0.05),CD3 T细胞、CD20 B细胞和S-100 树突状细胞与对照组无差别(P>0.05)。结论SARS患者肾内单核/巨噬细胞为主要免疫应答细胞,提示单核/巨噬细胞在SARS肾病变中起着重要作用。而T细胞、B细胞以及树突状细胞可能受到SARS冠状病毒的攻击,其功能可能遭到破坏。     

A unique patient with an Ullrich-Turner syndrome variant and mosaicism for a tiny r(X) and a partial proximal duplication 1q

We report on an apparently previously undescribed neonatal diffuse congenital hyperkeratosis with spontaneous improvement. The child, born to consanguinous parents, presented at birth with a verrucous hyperkeratosis involving face, trunk, and limbs, but sparing palms and soles. No visceral or skeletal abnormality was associated and neurosensory status was normal. The skin condition improved dramatically during the first month of life. At age 7 years, the child was healthy with normal psychomotor development and growth. He had an abnormal curvature of nose, ulerythema ophryogenes, and large ears. The skin was moderately dry. This favorable clinical outcome led us to propose the term "regressive congenital hyperkeratosis" until further molecular characterization of this new phenotype.     

Ontogeny and characterization of Factor XIIIa+ cells in developing human skin

Factor XIIIa (FXIIIa), a coagulation transglutaminase, is a cytoplasmic marker for dermal dendritic cells reported to be bone marrow-derived, phagocytic and antigen-presenting. In non-inflamed skin, these cells populate the papillary dermis in a perivascular distribution. They are increased in dermatoproliferative disorders and have been implicated as dermal stimulants for psoriatic hyperkeratosis. Since developing skin provides an example of dermal influence on the epidermis, we evaluated the presence of FXIIIa⁺ cells in human fetal skin to determine whether their location would suggest a role in morphogenetic events in the skin. Embryonic and fetal skin of progressive estimated gestational ages (EGA) was examined using immunocytochemistry with a polyclonal antibody to FXIIIa. At 6 weeks EGA, globular FXIIIa⁺ cells were present in the hypodermis. By 7–8 weeks, a compact sub-epidermal network of fusiform FXIIIa⁺ cells was also evident. By 11–12 weeks, the sub-epidermal cellular network was no longer FXIIIa⁺, but discrete FXIIIa⁺ dendritic cells were present in the reticular dermis. With advancing gestational age, FXIIIa⁺ dendritic cells populated the papillary dermis in a perivascular distribution. This adult-like distribution persisted through 22 weeks EGA, the oldest specimen examined. Because FXIIIa⁺ cells were evident in embryonic skin before the onset of bone marrow hematopoietic function, the skin was double-labeled with the FXIIIa antibody and with monoclonal antibodies to CD45 (marker for bone marrow-derived cells), CD68 (marker for macrophages) and HLA-DR (class II major histocompatibility antigen). Most of the FXIIIa⁺ dendritic cells did not colocalize CD45, but were CD68⁺; some cells did react with the HLA-DR antibody. Notably, the FXIIIa⁺ cells of the sub-epidermal network in the 7 weeks EGA specimens did not react with the other antibodies. We conclude that FXIIIa⁺ cells are first present in embryonic hypodermis and sub-epidermal dermis and later they are distributed in the papillary dermis in a perivascular pattern. In embryonic skin FXIIIa⁺ cells are not exclusively dendritic. Our data support the idea that cells that express FXIIIa do not constitute a unique bone marrow-derived cell type, but that multiple cell types produce FXIIIa.     

Presence of T cells and macrophages in inflammatory vitiligo skin parallels melanocyte disappearance.

Evidence for the involvement of cellular immunity in the etiopathogenesis of the hypopigmentary disorder vitiligo is provided by rare cases of inflammatory vitiligo. Nonlesional, perilesional, and lesional skin biopsies from three inflammatory vitiligo patients were immunohistochemically analyzed. The composition of inflammatory infiltrates present in perilesional skin was analyzed by antibodies to T cells (CD2, CD3, CD4, and CD8), Langerhans cells (CD1a), and macrophages (CD36 and CD68). The presence of activation markers on inflammatory cells was evaluated by analysis of HLA-DR, interleukin-2 receptor, and HECA452 expression. The presence or absence of melanocytes was determined by the antibody NKI-beteb. Moreover, the abundance of matrix molecule tenascin was semi-quantified using T2H5. Results indicate that within perilesional skin, epidermis-infiltrating T cells exhibit an increased CD8/CD4 ratio and increased cutaneous lymphocyte antigen and interleukin-2 receptor expression. These cells are frequently juxtapositionally apposed to remaining melanocytes. In perilesional dermis, CD68+OKM5- macrophages were more numerous than in lesional or nonlesional skin. Keratinocytes as well as melanocytes consistently express major histocompatibility complex class II antigens along stretches of basal and suprabasal layers in perilesional epidermis. Moreover, inflammation is accompanied by increased tenascin content. Although these observations do not permit differentiation between the immune infiltrates being a result as opposed to the cause of the disease process, results presented in this study are very suggestive of involvement of local immune reactivity in melanocyte destruction.