蚕蚀性角膜溃疡患者外周血淋巴细胞免疫表型和T细胞活化的研究

目的：观察蚕蚀性角膜溃疡患者外周血淋巴细胞免疫表到和T细胞活化的变化,进一步探讨本病患者的免疫功能。方法：用流式细胞术检测8例活动性蚕蚀性角膜溃病患者外周血淋巴细胞对项免疫表型变化,用植物凝集素和沸被醉酯作为刺激剂,以HLA－DR,CD69、CD71、CD25的表达为T细胞活化的指标,用流式细胞术分析。结果：蚕蚀性角膜溃疡患者外周血CD4、CD5、CD8、CD10、CD11b、CD15、CD16、CD19、CD23、CD25、CD41、CD69、HLA-DP、HLADR抗原表达阳性淋巴细胞以及HLA－DR、CD69、CD71、CD25在CD3和CD8细胞的表达均明显高于正常对照组。结论：蚕蚀性角膜溃疡患者具有增强的细胞免疫和体液免疫,外周血T细胞及其亚群出现异常活化,反映患者体内存在自身免疫反应而支持该病发病之自身免疫学说。     

The Role of Innate Immune Cells in Nonalcoholic Fatty Liver Disease

Nonalcoholic fatty liver disease (NAFLD) is a very common hepatic pathology featuring steatosis and is linked to obesity and related conditions, such as the metabolic syndrome. When hepatic steatosis is accompanied by inflammation, the disorder is defined as nonalcoholic steatohepatitis (NASH), which in turn can progress toward fibrosis development that can ultimately result in cirrhosis. Cells of innate immunity, such as neutrophils or macrophages, are central regulators of NASH-related inflammation. Recent studies utilizing new experimental technologies, such as single-cell RNA sequencing, have revealed substantial heterogeneity within the macrophage populations of the liver, suggesting distinct functions of liver-resident Kupffer cells and recruited monocyte-derived macrophages with regards to regulation of liver inflammation and progression of NASH pathogenesis. Herein, we discuss recent developments concerning the function of innate immune cell subsets in NAFLD and NASH.     

Role of flow cytometry in diagnostics of myelodysplastic syndromes--beyond the WHO 2008 classification

Multiparameter flow cytometry (FCM) is an excellent method to follow the expression patterns of differentiation antigens using monoclonal antibodies to surface and cytoplasmic proteins. Although several authors described various aberrant immunophenotypic features in the bone marrow of patients with myelodysplastic syndromes (MDS), the World Health Organization 2008 classification recommended that, only if 3 or more phenotypic abnormalities are found involving 1 or more of the myeloid lineages can the aberrant FCM findings be considered suggestive of MDS. In the absence of conclusive morphologic and/or cytogenetic features, FCM abnormalities alone were considered not sufficient to establish MDS diagnosis and further follow-up of the patients was recommended. Review of the literature gives accumulating evidence that FCM has become an important part of the integrated diagnostic work-up of patients with suspected MDS. Several studies have also reported FCM findings significant for prognosis and therapy choice in MDS patients. Technical progress in multicolor FCM and new analysis programs, together with ongoing efforts to standardize the methodology, will make it possible to apply FCM in individual risk assessment and choice of best therapy for MDS patients.     

Innate Immune Cells in the Adipose Tissue in Health and Metabolic Disease

Metabolic disorders, such as obesity, type 2 diabetes mellitus, and nonalcoholic fatty liver disease, are characterized by chronic low-grade tissue and systemic inflammation. During obesity, the adipose tissue undergoes immunometabolic and functional transformation. Adipose tissue inflammation is driven by innate and adaptive immune cells and instigates insulin resistance. Here, we discuss the role of innate immune cells, that is, macrophages, neutrophils, eosinophils, natural killer cells, innate lymphoid type 2 cells, dendritic cells, and mast cells, in the adipose tissue in the healthy (lean) and diseased (obese) state and describe how their function is shaped by the obesogenic microenvironment, and humoral, paracrine, and cellular interactions. Moreover, we particularly outline the role of hypoxia as a central regulator in adipose tissue inflammation. Finally, we discuss the long-lasting effects of adipose tissue inflammation and its potential reversibility through drugs, caloric restriction, or exercise training.     

新疆乌鲁木齐地区142例急性白血病流式细胞术免疫分型研究

目的:研究新疆乌鲁木齐地区急性白血病(AL)患者免疫表型特征及分布特点。方法:选用细胞表面分子CD20、CD14、CD3、CD2、CD33、HLA-DR、CD15、CD10、CD5、CD22、CD7、CD13、CD34、CD11b、CD19、CD117等的单克隆抗体,采用流式细胞仪CD45/SSC双参数散点图设门法对142例AL患者进行免疫表型分析。结果:35例急性淋巴细胞白血病(ALL),其中6例伴有髓系抗原的表达(14.3%)且表达最频繁的为CD13;96例急性髓系白血病(AML),其中21例伴有淋系抗原的表达(21.9%)且表达最频繁的为CD7;11例为FAB难以分类的急性白血病(UAL),兼有淋系和髓系抗原的表达。ALL免疫分型特点在新疆汉族和维吾尔族(简称维族)中差异无统计学意义(P>0.05),在AML中,汉族髓系抗原的表达率依次为CD13>CD33>CD15,维族髓系抗原的表达率依次为CD13>CD15>CD14,且维族患者多伴有淋系抗原CD7的表达。结论:FCM免疫分型是在细胞形态学和细胞染色基础上对AL诊断与分型的重要补充,免疫表型的检测对AL的诊断和治疗有重要意义。     

白血病免疫分型诊断技术的进展

白血病是造血系统的恶性疾病,俗称“血癌”,是国内十大高发恶性肿瘤之一.目前,免疫分型已是当今白血病诊断分型、预后判断和残留病检测的一个有力手段.免疫分型诊断的主要方法有免疫细胞化学技术、流式细胞术、细胞芯片技术,也有个别学者在尝试压电免疫传感器.临床上仍以流式细胞术(FCM)方法为主,目前一次最多能检测到15种抗原,且昂贵.而细胞免疫芯片检测技术,一次可以检测数十种乃至上百种细胞抗原,如能应用于临床,将是一种方便,快速,简单,经济实用的白血病免疫分型检测手段.     

Flow cytometry and its application in small animal oncology

Flow cytometry measures multiple characteristics of single cells. The use of flow cytometry in the veterinary clinical laboratory has increased considerably during the past decade. The most common applications of flow cytometry in small animal oncology are measurement of DNA content in tumours and immunophenotyping of haematopoietic malignancies. DNA ploidy and S-phase rate of various tumours in dogs have been found to be independent predictor of patient outcome. In dogs with lymphomas immunophenotyping is recommended as a part of the patient work-up. Flow cytometry has shown to be a suitable method for immunophenotyping of canine lymphomas. However, it has not become a routine technique in small animal oncology yet. This report reviews the applications of flow cytometry in small animal oncology. Besides basic principles and technical aspects, the clinical relevance of DNA-analysis and immunophenotyping are discussed.     

多参数流式细胞术在非霍奇金淋巴瘤诊断中的应用

目的探讨流式细胞术在非霍奇金淋巴瘤（NHL）诊断中的应用意义。方法用多参数流式细胞术对40例淋巴组织增生性疾病的细针穿刺或手术切除淋巴结新鲜标本的活细胞表面抗原进行检测,同时结合细胞涂片的形态学观察,分析淋巴细胞的免疫表型特征,并与病理诊断及免疫组织化学结果进行比较。结果40例诊断为NHL的标本,经过流式细胞免疫分型联合细胞涂片观察细胞形态进行分析,37例（92．5％）符合NHL,其中病理组织学诊断为B—NHL的20例,经过流式细胞仪检测均为B—NHL,符合率为100％;病理组织学诊断为T—NHL的17例中,经过流式细胞仪检测符合T—NHL的12例（70．6％）;2例（11．8％）修正为B—NHL;3例（17．6％）未能明确诊断。结论流式细胞免疫分型有助于提高NHL诊断的正确性和亚型分类的准确性。     

人子宫内膜干细胞的分离、培养及鉴定

目的 分离、培养子宫内膜干细胞并进行鉴定.方法 采用酶消化和机械方法相结合分离人子宫内膜细胞,两次滤网过滤分离基质细胞和上皮细胞.采用有限稀释法培养基质细胞和上皮细胞,观察细胞的形态及生长特性,免疫荧光检测波形蛋白和细胞角蛋白表达情况.原代培养15 d后,计算细胞克隆形成率,流式细胞术检测抗原CD133、CD34、CD45、CD90、CD73及CD29表达.结果 基质细胞呈纤维样细胞形态,排列呈辐射状,免疫荧光显示其波形蛋白表达阳性;原代培养15 d后,细胞克隆形成率达(1.34±0.44)％.上皮细胞呈多角形或椭圆形,免疫荧光显示其细胞角蛋白表达阳性;原代培养15d后,未发现克隆形成.流式细胞术检测克隆形成的基质细胞CD29、CD90、CD73表达阳性,CD34、CD45、CD133表达阴性.结论 人体存在少量具有克隆形成能力和高度增生潜能的子宫内膜干细胞,这些干细胞也可能来源于骨髓间充质干细胞.     

The Toxicity of Nonsteroidal Anti-inflammatory Eye Drops against Human Corneal Epithelial Cells in Vitro

This study investigated the toxicity of commercial non-steroid anti-inflammatory drug (NSAID) eye solutions against corneal epithelial cells in vitro. The biologic effects of 1/100-, 1/50-, and 1/10-diluted bromfenac sodium, pranoprofen, diclofenac sodium, and the fluorometholone on corneal epithelial cells were evaluated after 1-, 4-, 12-, and 24-hr of exposure compared to corneal epithelial cell treated with balanced salt solution as control. Cellular metabolic activity, cellular damage, and morphology were assessed. Corneal epithelial cell migration was quantified by the scratch-wound assay. Compared to bromfenac and pranoprofen, the cellular metabolic activity of diclofenac and fluorometholone significantly decreased after 12-hr exposure, which was maintained for 24-hr compared to control. Especially, at 1/10-diluted eye solution for 24-hr exposure, the LDH titers of fluorometholone and diclofenac sodium markedly increased more than those of bromfenac and pranoprofen. In diclofenac sodium, the Na⁺ concentration was lower and amount of preservatives was higher than other NSAIDs eye solutions tested. However, the K⁺ and Cl^- concentration, pH, and osmolarity were similar for all NSAIDs eye solutions. Bromfenac and pranoprofen significantly promoted cell migration, and restored wound gap after 48-hr exposure, compared with that of diclofenac or fluorometholone. At 1/50-diluted eye solution for 48-hr exposure, the corneal epithelial cellular morphology of diclofenac and fluorometholone induced more damage than that of bromfenac or pranoprofen. Overall, the corneal epithelial cells in bromfenac and pranoprofen NSAID eye solutions are less damaged compared to those in diclofenac, included fluorometholone as steroid eye solution.     

Evaluation of canine bone marrow proliferative disorders by use of flow cytometric analysis of CD45 expression and intracytoplasmic complexity

Bone marrow from 14 dogs with proliferative disorders was evaluated by labeling cells with anti-CD45 and analyzing them by use of flow cytometry. Bone marrow from five healthy dogs was also evaluated to identify normal cell labeling patterns. The cells were displayed as the logarithm of red fluorescence intensity vs side-angle light scatter plots (CD45 scatter plots). For healthy dogs, the CD45 scatter plot technique identified four discrete cell populations. The immunophenotype of these cells identified these populations as granulocytes, erythroid cells, lymphocytes, and monocyte/macrophages. The CD45 scatter plot technique identified discrete populations of atypical cells in all dogs with proliferative disorders. Immunophenotyping of the atypical populations resulted in diagnoses of myelodysplastic syndrome, acute myeloid leukemia, acute lymphocytic leukemia, chronic lymphocytic leukemia, metastatic lymphosarcoma, megakaryoblastic leukemia, and metastatic mast cell tumor. Possible clinical applications of the CD45 labeling technique include detection of malignant cell populations for immunophenotyping, early detection of metastasis of lymphosarcoma to bone marrow, detection of residual malignant cells after chemotherapy, and early detection of recurrence of leukemias.     

流式细胞术在我院急性白血病免疫分型中的应用

目的探讨本实验室所用13种单抗对白血病免疫分型的应用价值。方法应用流式细胞术（FCM）使用13种单抗对99例急性白血病进行免疫分型,与形态学分型相比较。结果使用13种单抗从99例急性白血病中分出髓系白血病（AML）59例、淋系白血病（ALL）27例、混合型（MAL）5例、未分化型（AUL）2例,诊断率达93.9%（93/99）。AML中各抗原表达情况：CD13〉CD33〉CD14、CD7〉CD2、CD19〉CD10〉CD5〉CD20,其中多表达CD13、CD33;ALL中各抗原表达情况：CD19〉CD13〉CD10〉CD20〉CD33〉CD5〉CD7〉CD2、CD14,其中CD10、CD13、CD19、CD20表达率较高。免疫分型与形态学分型结果相比较,AML二者的吻合率为90.5%（57/63）、ALL二者的吻合率为76.5%（26/34）;其中1例M0免疫分型为B/M混合、1例M1免疫分型为T/M混合、1例M2a免疫分型为T/M混合、1例L1免疫分型为B/M混合、1例L2免疫分型为T/B混合、1例L2免疫分型为Ly＋-AML、2例L2免疫分型为AUL;1例形态学为浆细胞白血病的免疫分型为AML、1例淋单混合型白血病CD7、CD13、CD14阳性。结论白血病细胞、免疫分型各具特征,形态学相同的白血病免疫分型未必相同,形态学不同的白血病免疫分型未必不同;本实验室所选单抗组合基本能满足白血病的免疫分型,对于一些混合型和未分化型白血病的诊断亦能明确。     

异体角膜刺激引起的人外周血T细胞及其亚群CD25分子表达

目的：观察异体角膜体外对人外周血T细胞及其亚群CD25分子表达的影响。方法： 异体角膜与外周血淋巴细胞体外共同培养后，进行单克隆抗体标记和流式细胞分析。结果： 对照组T细胞CD25表达为25.2％；受角膜或沸波醇酯（PDB）激活后T细胞CD25表达分别为56.8％和80.9％；受角膜和沸波醇酯共同激活后T细胞CD25表达为70.2％；受异体角膜激活后，CD4和CD8 T淋巴细胞CD25表达分别为67.3％和52.3％。结论： 异体角膜组织体外能够刺激人外周T细胞CD25表达和T细胞活化，CD4 T细胞比CD8 T细胞活化更明显。     

流式细胞仪三标记法检测白细胞介素-10对人树突状细胞表型的影响

目的观察白细胞介素(IL)10对体外培养人树突状细胞(DC)表型的影响,探讨流式细胞术三色荧光标记抗体检测细胞表面抗原的方法及意义。方法通过SCF、GMCSF、TGFβ1、Flt3和TNFα体外培养体系,将脐血CD34+造血干细胞诱导扩增获得DC,并于成熟过程中用重组人IL10进行干预。采用流式细胞术的三色荧光标记(FITC、PE、CY)单克隆抗体直接检测技术,分析细胞表型CD1a、CD11c、CD83、CD80、CD86和HLADR。结果IL10可下调成熟中DC表面CD11c、CD83、CD80和CD86的表达。结论IL10通过抑制DC表面黏附共刺激分子表达,可调节DC的递呈抗原功能;此外,采用流式细胞仪的三色荧光标记检测方法,不仅可以节约DC细胞用量,而且具有快速和准确的优点,值得推广应用。     

Role of TLR-2 and Fungal Surface Antigens on Innate Immune Response Against Sporothrix schenckii

Sporotrichosis is an infection caused by the dimorphic fungus Sporothrix schenckii. Toll-like receptors (TLRs) play an important role in immunity, since they bind to pathogen surface antigens and initiate the immune response. However, little is known about the role of TLR-2 and fungal surface antigens in the recognition of S. schenckii and in the subsequent immune response. This study aimed to evaluate the involvement of TLR-2 and fungal surface soluble (SolAg) and lipidic (LipAg) antigens in phagocytosis of S. schenckii and production of immune mediators by macrophages obtained from WT and TLR-2^-/- animals. The results showed that TLR-2^-/- animals had had statistical lower percentage of macrophages with internalized yeasts compared to WT. SolAg and LipAg impaired phagocytosis and immunological mediator production for both WT and TLR-2^-/-. The absence of TLR-2 led to lower production of the cytokines TNF-α, IL-1β, IL-12 and IL-10 compared to WT animals. These results suggest a new insight in relation to how the immune system, through TLR-2, recognizes and induces the production of mediators in response to the fungus S. schenckii.     

呼吸道疾病患儿免疫功能分析

目的 探讨呼吸道感染儿童免疫功能指标的变化.方法 将2012年1月至2012年9月在大连儿童医院住院的呼吸道疾病患儿按临床诊断结果分为支气管炎组、咽炎组、肺炎组.另选取40名正常儿童作为对照组.运用流式细胞术分别检测全血T淋巴细胞分化抗原三项CD3、CD4、CD8及CD4/CD8的比值;同时利用散射比浊终点法检测血清中免疫球蛋白（IgG、IgM、IgA）、补体（C3、C4）.结果 ①支气管炎组、咽炎组、肺炎组检验结果显示,CD3、CD4、CD8、CD4/CD8低于对照组.②以上3组的血清免疫球蛋白与C3水平均低于对照组,差异有统计学意义,C4水平与对照组检测结果基本一致,差异无统计学意义.③不同年龄组结果显示：0～3、4～6年龄段的患儿细胞免疫功能和体液免疫功能均低于对照组,C4与对照组检测结果基本一致,差异无统计学意义.7～12岁患儿体液免疫和细胞免疫功能与对照组比较无显著性差异.④呼吸道感染在患儿性别间无统计学差异.结论 呼吸道疾病患儿免疫功能低下并紊乱;6岁以下患儿年龄越低,免疫功能越低,易引起呼吸道疾病;患儿主要集中在6岁以下;患病儿童与性别无关.     

The Role of Pancreatic Infiltrating Innate Immune Cells in Acute Pancreatitis

Acute pancreatitis (AP) is a leading cause of gastrointestinal-related hospital admissions with significant morbidity and mortality. Although the underlying pathophysiology of AP is rather complex, which greatly limits the treatment options, more and more studies have revealed that infiltrating immune cells play a critical role in the pathogenesis of AP and determine disease severity. Thus, immunomodulatory therapy targeting immune cells and related inflammatory mediators is expected to be a novel treatment modality for AP which may improve the prognosis of patients. Cells of the innate immune system, including macrophages, neutrophils, dendritic cells, and mast cells, represent the majority of infiltrating cells during AP. In this review, an overview of different populations of innate immune cells and their role during AP will be discussed, with a special focus on neutrophils and macrophages.