银屑病患者皮肤组织一氧化氮的检测分析

目的:研究皮肤组织一氧化氮(NO)水平与银屑病之间的关系。方法:选择寻常型银屑病患者54例,利用Griss化学比色法检测患者皮损组织及非皮损组织中NO水平。结果:银屑病患者皮损组织及非皮损组织中NO水平明显高于正常人且与疾病严重程度有相关性(P＜0.01)。结论:银屑病患者皮肤组织中存在诱导型一氧化氮合成酶(iNOS)的异常表达,与疾病的严重程度呈一定正相关。     

黄芩甙影响人成纤维细胞诱导型一氧化氮合酶蛋白表达研究

目的 研究成纤维细胞诱导型一氧化氮合酶(iNOS)蛋白表达情况，以及细胞因子和黄芩甙对其表达的影响，探讨黄芩甙治疗银屑病机制。方法 体外培养的成纤维细胞，通过TNF-α、IFN-γ和IL-8刺激，用免疫组化方法研究成纤维细胞表达iNOS情况，并观察黄芩甙对其表达的影响。结果 未受到细胞因子刺激时成纤维细胞不表达iNOS；TNF-α(终浓度1000U／mL)、IFN-γ(200U／mL)、IL-8(200pg/mL)单独或分别组合后刺激成纤维细胞24h后的蛋白分析显示：单独TNF-α能够刺激成纤维细胞表达较弱iNOS，而IFN-γ或IL-8处理未显示该效应，联合TNF-α、IFN-γ和IL-8显示增强的诱导iNOS表达作用，免疫组化有强染色区，位于胞浆近胞核处；50μg／mL黄芩甙能够抑制细胞因子诱导成纤维细胞表达iNOS。结论黄芩甙能够抑制细胞因子刺激成纤维细胞的iNOS蛋白表达，减少NO的产生，发挥抗炎和治疗银屑病的作用。     

皮肤血管瘤中NOS和VEGF的表达及其与血管增生的关系

目的研究内皮型一氧化氮合酶(eNOS)、诱生型一氧化氮合酶(iNOS)及血管内皮生长因子(VEGF)在增生期皮肤血管瘤组织中的表达及关系,探讨它们在皮肤血管瘤血管生成中的作用。方法应用免疫组化法检测51例增生期皮肤血管瘤标本中eNOS,iNOS和VEGF的表达,第Ⅷ因子相关抗原(FⅧRAg)血管内皮细胞特异性染色计数肿瘤微血管密度(MVD)。结果42例血管瘤组织表达VEGF,32例表达iNOS,37例表达eNOS,血管畸形表达较弱或不表达eNOS,iNOS和VEGF;eNOS表达阳性者MVD与表达阴性者差异无显著性(P>0.05),iNOS表达阳性者MVD与表达阴性者差异有显著性(P<0.05),VEGF表达阳性者MVD与表达阳性者差异有极显著性(P<0.01)。结论iNOS对VEGF的表达和在调节血管生成过程中可能起着重要作用;VEGF和iNOS两者对血管瘤血管生成具有促进作用。     

TNF-α和IL-1β对HaCaT细胞诱生型一氧化氮合酶表达的影响

目的　探讨炎症性细胞因子肿瘤坏死因子 α(tumornecrosisfactor α ,TNF α)协同白细胞介素 1β(inter leukin 1β ,IL 1β)对体外培养角质形成细胞 (keratinocye ,KC)株HaCaT细胞诱生型一氧化氮合酶 (induciblenitricoxidesyn thase ,iNOS)mRNA和蛋白表达的调节作用 ,以及地塞米松对TNF α和IL 1β作用的影响。 方法　用RT PCR、Westernblotting和免疫组织化学 (SP)方法检测HaCaT细胞iNOSmRNA和蛋白表达情况。结果　正常培养HaCaT细胞iNOS微弱表达或不表达 ,TNF α协同IL 1β显著上调HaCaT细胞iNOSmRNA和蛋白表达 ,地塞米松可显著抑制TNF α和IL 1β的作用。结论　推测TNF α和IL 1β可能通过上调角质形成细胞表达iNOS合成释放的一氧化氮 (niricoxide ,NO)参与皮肤免疫和炎症反应 ;地塞米松的治疗效应可能部分与其能抑制iNOS表达有关。     

中波紫外线对人角质形成细胞iNOS的作用

目的观察中波紫外线(UVB)照射对体外培养正常人角质形成细胞(HKC)中诱导型一氧化氮合成酶(iNOS)的作用,探讨紫外线引起皮肤细胞急性损伤的发病机制。方法体外培养HKC,用免疫荧光染色和蛋白印迹技术检测照射后不同时间细胞内iNOS的水平。结果HKC经UVB照射后,免疫荧光法检测到细胞膜胞浆侧有iNOS,蛋白印迹法检测到细胞提取总蛋白中分子量为130kDa的蛋白条带可以被抗iNOS多克隆抗体所识别,二者结果相符合。结论UVB照射可诱导体外培养的HKC在胞浆内产生iNOS,iNOS蛋白的表达可能与紫外线引起的皮肤急性炎症反应有关。     

CD44 Expression in Normal Human Skin and Skin Tumors

CD44 is thought to be a principal cell surface receptor for hyaluronic acid. Although the distribution of hyalulonic acid has been studied, little is known about the distribution of the CD44 molecule in the human skin and skin tumors. This study was undertaken to investigate the distribution of the CD44 molecule in normal human skin as well as in benign and malignant skin tumors. In normal skin, CD44 was expressed on 1) keratinocyte cell surfaces throughout the epidermis except for the granular and horny layers, 2) hair follicular cells, 3) eccrine sweat gland cells, and 4) cell surfaces of dendritic cells in the dermis. In skin tumors, although CD44 was expressed on the tumor cell surface of seborreic keratosis, Bowen's disease, and squamous cell carcinoma as in normal skin, we could not detect any CD44 expression on the cell surface of the tumor cells of basal cell carcinoma. However, CD44 positive dendritic cells were observed in the tumor islands of basal cell carcinoma. Phenotypic analysis suggested that these CD44 positive cells were melanocytes.     

降钙素基因相关肽对HaCaT细胞一氧化氮合酶表达的影响

目的 研究降钙素基因相关肽对体外培养角质形成细胞HaCaT株神经型一氧化氮合酶(NOS)mRNA、蛋白表达和NO释放的调节作用。方法 应用NO试剂盒(酶法)检测培养细胞上清液中NO水平,RT-PCR和免疫组化(SP)方法检测HaCaT细胞神经型NOSmRNA和蛋白的表达水平。结果 体外正常培养的HaCaT细胞表达和释放基础水平的神经型NOS和NO,降钙素基因相关肽上调HaCaT细胞神经型NOS表达和NO释放,降钙素基因相关肽受体抑制剂CGRP-8-37抑制降钙素基因相关肽的作用。结论 降钙素基因相关肽诱导角质形成细胞表达神经型NOS并促进神经型NO释放。     

Macrophage Markers 25F9 and 27E10 on Human Keratinocytes in Normal and Diseased Skin

In the present study, an immunohistochemical analysis was made of the expression pattern of different macrophage markers such as 25F9, 27E10 and RM 3/1 on the surface of human keratinocytes (HK) in biopsies obtained from healthy volunteers and from patients with lichen planus, chronic cutaneous graft-versus-host reaction, mycosis fungoides, and purpura pigmentosa chronica. In biopsies from the healthy volunteers and from both clinically involved and uninvolved skin of the patients, the HK of the basal cell layer exhibited a specific peroxidase-positive reaction when the monoclonal antibody against 25F9 was used. Lesional HK from all patients studied displayed 25F9 and 27E10 in nearly the entire epidermis. The present findings provide further evidence that HK and macrophages share a number of common cell surface moieties.     

An Immunohistochemical Study of BCA-225 in Various Skin Cancers

We performed an immunohistochemical study of BCA-225, which is a glycoprotein secreted by the T47D breast carcinoma cell line and recognized by monoclonal antibody BRST-1 (clone name: CU-18), in normal skin and various skin cancers. In normal skin, BCA-225 was positive only in the secretory portion of both eccrine and apocrine glands and in mature cells of the sebaceous gland. We observed 10 cases of squamous cell carcinoma of the skin, 10 cases of basal cell carcinoma without sebaceous differentiation, 3 cases of basal cell carcinoma with sebaceous differentiation, 6 cases of malignant trichilemmoma, 8 cases of eccrine porocarcinoma, 3 cases of ductal carcinoma, 1 case of malignant clear cell hidradenoma, 1 case of apocrine adenocarcinoma, 6 cases of extra-ocular sebaceous carcinoma, 5 cases of extramammary Paget's disease with underlying adenocarcinoma, and 11 cases of extramammary Paget's disease without underlying adenocarcinoma. Most of the cases of sweat gland carcinoma, basal cell carcinoma with sebaceous differentiation, sebaceous carcinoma, and extramammary Paget's disease were positive for BCA-225, while none of the cases of squamous cell carcinoma, basal cell carcinoma without sebaceous differentiation, or malignant trichilemoma were positive. Based on these findings, we believe that BCA-225 is useful in distinguishing tumors with sweat gland and sebaceous differentiation and extramammary Paget's disease from tumors without such differentiation.     

Immunohistochemical study of angiotensin receptors in normal human sweat glands and eccrine poroma

BACKGROUND: Angiotensin II exerts its actions through its specific receptors. However, expression of these receptors has not been determined in sweat glands. OBJECTIVES: To clarify the expression and localization of the angiotensin receptors in normal human sweat glands and eccrine poroma. METHODS: Expression of angiotensin type 1 (AT1) and type 2 (AT2) receptors in normal human eccrine and apocrine sweat glands and 12 cases of eccrine poroma was studied using immunohistochemistry. RESULTS: In eccrine sweat glands, the acrosyringium and the inner surfaces and luminal cells of the intradermal duct showed positive staining with AT1. In apocrine sweat glands, the intraepithelial duct and luminal cells of the intradermal duct showed positive staining with AT1. In 12 cases of eccrine poroma, some of the tumour cells in the tumour strands and cells surrounding the luminal structures stained positively. There were no positive findings with AT2. CONCLUSIONS: Studying AT1 distribution may be useful in understanding the pathophysiology of sweat glands and sweat gland tumours.     

恶性黑素瘤内皮型一氧化氮合酶和血管内皮生长因子的表达

目的 探讨内皮型一氧化氮合酶(eNOS)和血管内皮生长因子(VEGF)与人恶性黑素瘤血管生成的关系。方法 免疫组化法检测31例人恶性黑素瘤标本eNOS、VEGF和第Ⅷ因子相关抗原(FⅧRAg)血管内皮细胞特异性染色,计数肿瘤微血管密度(MVD)。结果 24例恶性黑素瘤组织表达eNOS,26例表达VEGF,色素痣组织不表达eNOS和VEGF;VEGF与eNOS的表达呈正相关;eNOS、VEGF的表达与恶性黑素瘤MVD呈正相关,恶性黑素瘤MVD明显高于色素痣;eNOS、VEGF表达与淋巴结转移无关。结论 MVD随着eNOS和VEGF表达的增强而增加。     

Thrombospondin Receptor (CD36) Expression of Human Keratinocytes during Wound Healing in a SCID Mouse/Human Skin Repair Model

Using a human skin/severe combined immunodeficient (SCID) chimeric mouse model, we examined the keratinocyte expression of the thrombospondin receptor (CD36) and its ligand thrombospondin-1 (TSP1) in acute uninflamed wounds. Positive suprabasal keratinocyte expression of CD36 was observed as early as 30 minutes after wounding in the adjacent, intact epidermis; it disappeared 4 days later. Keratinocytes of the freshly re-epithelised wounds and those of the surrounding epidermis remained TSP1-negative throughout the whole observation period of 7 days. Our results indicate that CD36-positive keratinocytes, probably in connection with activated, TSP1-positive thrombocytes, may play an important role in the early phase of wound healing.     

rhFGF-7对体外培养人皮肤角质形成细胞增殖的影响

目的建立一种人皮肤角质形成细胞分离和培养的方法,研究重组人成纤维细胞生长因子7(rhFGF-7)对人皮肤角质形成细胞增殖的影响。方法取环切术后包皮,分别用胰酶和dispaseⅡ酶消化法分离表皮,用无血清培养基进行无滋养层培养,采用免疫细胞化学方法对培养的细胞进行鉴定,并用细胞计数法测定细胞的生长曲线,MTT法检测rhFGF-7对人皮肤角质形成细胞增殖的影响。结果与胰酶消化分离表皮相比,dispaseⅡ酶消化获得的表皮,其细胞贴壁率高,增殖速度快,且成纤维细胞污染少;荧光显微镜下细胞胞浆呈黄绿色,为角蛋白阳性染色;培养的角质形成细胞可持续增殖至第8代,10ng/mL的rhFGF-7促角质形成细胞增殖作用最显著。结论所建立的人皮肤角质形成细胞分离和培养方法,可获得高纯度的人皮肤角质形成细胞,rhFGF-7可促进体外培养人皮肤角质形成细胞的增殖。     

一氧化氮诱导正常人皮肤炎症反应的实验研究

目的探讨一氧化氮(NO)在皮肤炎症反应中的作用。方法于健康人皮肤外用两种NO供体,通过真皮血流变化曲线及化学发光法确定与5%亚硝酸盐产生相近的NO生理效应及NO浓度的沸石一氧化氮霜浓度,应用免疫组化及ELISA法测定二者外用后炎症反应及浸润T淋巴细胞种类。结果①33%沸石一氧化氮霜与5%亚硝酸盐霜外用于皮肤后,二者血流曲线相近,前者真皮浅层NO水平略高于后者,二者比较无明显差异(P>0.05)。②二者外用于皮肤后,免疫组化结果示亚硝酸盐组表皮朗格汉斯细胞较正常对照组明显减少(P<0.05),真皮CD3、CD4、CD68及中性粒细胞弹性蛋白酶阳性细胞较正常对照明显升高。而外用沸石一氧化氮霜组真皮中示CD4阳性细胞增高(P<0.05)。③ELISA法测定外用33%沸石一氧化氮霜后表皮负压吸引疱疱液中IFN-γ水平较正常对照组显著增高,IL-4水平未测出。结论NO对正常人皮肤无明显致炎症作用,仅可引起真皮内Th1细胞浸润。     

Role of Integrins in Cell Adhesion and Polarity in Normal Keratinocytes and Human Skin Pathologies

In vitro, normal human keratinocytes reconstitute a differentiated stratified epidermis, maintaining the same gene expression pattern as its in vivo counterpart and are suitable for permanent grafting onto patients. Keratinocyte adhesion to basal lamina and lateral interactions among basal epidermal cells are also mediated by integrin receptors that are sorted to defined plasma membrane domains. The hemidesmosome-associated integrin α₆β₄ is sharply localized at the basal surface of basal cells and codistributes with laminin and nicein/kalinin; the α₂β₁ and α₃β₁ integrins are enriched laterally and play crucial roles in cell-cell interaction and proper colony morphology. During wound healing, proliferating and migrating keratinocytes express on their plasma membrane α_vβ₅ and α₅β₁, which allow keratinocyte attachment and migration over the provisional matrix present in the wound. TGFβ, which is an autocrine and paracrine mediator in wound healing, specifically increases the synthesis and expression of α_vβ₅ and α₅β₁, induces the de novo expression of α_vβ₆, and determines the loss of integrin polarization. In hyperproliferative skin diseases, such as skin cancer or psoriasis vulgaris, and in normal keratinocytes forced into more frequent cell cycles, the polarized expression of integrins is lost, and α₅β₁ becomes costitutively expressed on the plasma membrane. In addition, the α₆β₄ integrin becomes associated with focal contacts. Nerve growth factor (NGF) is a potent autocrine stimulator of keratinocyte growth and induces melanocyte migration toward the leading edge of a healing wound. We are currently investigating the NGF-dependent modulation of integrin expression and function in keratinocytes and melanocytes in both normal epidermis and several hyperproliferative skin diseases.     

斑秃患者血浆内皮素及血清一氧化氮水平测定及其意义

目的 :探讨血浆内皮素 (ET)和血清一氧化氮 (NO)在斑秃发病中的作用。方法 :应用放射免疫法和Greiss法测定 2 3例斑秃活动期患者、1 1例斑秃稳定期患者以及 2 5例正常人血浆ET和血清NO值。结果 :斑秃活动期患者ET水平和ET/NO比值显著高于正常人 (P <0 0 5 ) ,但其NO水平与正常人无显著性差异 (P >0 0 5 ) ;斑秃稳定期ET和NO水平以及ET/NO比值与正常人无显著性差异(P >0 0 5 )。结论 :斑秃活动期患者ET水平和ET/NO比值可能与斑秃的活动性有关     

大黄素次酸对豚鼠皮肤黑素细胞一氧化氮合酶的调控作用

目的:探讨大黄有效成分大黄素次酸对豚鼠皮肤黑素细胞的一氧化氮合酶(Nitric Oxide Synthase;NOS)表达的影响,探索大黄在活体皮肤中对黑素细胞的黑素合成的有效作用浓度和作用机制.方法:将21只雄性豚鼠随机分成对照组及5个实验组,用5种浓度大黄素次酸对局部皮肤进行皮下注射处理,48小时后取材,以免疫组织化学方法(SABC)显示NOS的表达,用光学显微镜和图象分析仪获得实验结果,进行统计分析.结果:在大黄素次酸作用下,表皮黑素细胞NOS表达明显减少,光密度明显下降(P＜0.05);不同浓度药物作用之间无显著差异(P＞0.05).结论:大黄素次酸对黑色素细胞NOS的表达具有调节作用,提示大黄对黑素细胞的调节是经NO信号介导途径,为大黄的临床应用提供实验依据.     

The distribution of choline acetyltransferase- and acetylcholinesterase-like immunoreactivity in the palmar skin of patients with palmoplantar pustulosis

The distribution of choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) in involved skin in patients with palmoplantar pustulosis (PPP) and in normal palmar skin in healthy non-smokers and smokers has been studied by immunohistochemistry, especially in relation to the sweat gland apparatus. The sweat gland and its duct showed ChAT- and AChE-like immunoreactivity (LI) of varying intensity in all three groups and with stronger reactivity than in the epidermis. ChAT-LI was present in the coil and in the duct except in the corneal layer. Smokers and patients with PPP displayed significantly fewer ChAT+ acrosyringia than non-smokers. In the patients with PPP, the granulocytes in the pustules and in the papillary dermis displayed ChAT-LI. Western blot analysis of granulocytes from peripheral blood from healthy donors confirmed the presence of ChAT-like proteins in large amounts in neutrophils and small amounts in eosinophils. AChE-LI of varying intensity was found in all parts of the sweat gland apparatus in all three groups. The strongest AChE-LI in the acrosyringia was seen in the lowest part of the stratum corneum, where the PPP pustules are located. No significant differences in staining pattern or intensity were found between the coils, nerve fibres surrounding the coils or ducts. The number of mast cells in the papillary dermis was about four times larger in the patients with PPP than in the control subjects. AChE-LI was observed in about 25% of the mast cells in non-smoking control subjects and in patients with PPP, but only in 10% of those in the smoking control subjects. Our findings indicate that the (non-neuronal) cholinergic system may be involved in cutaneous inflammatory processes.