Cytokine secretion by peripheral blood mononuclear cells in myasthenia gravis.

We studied spontaneous secretion of anti-acetylcholine receptor antibody (AChRAb), IgG and cytokines by peripheral blood mononuclear cells (PBMC) from 19 MG patients without therapy and 10 normal controls. IgG secretion was higher in the culture medium of MG than in that of normal controls. AChRAb secretion was correlated with IgG secretion in MG. Interleukin (IL)-2, IL-4, IL-5, IL-6, IL-10, IL-13, interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha secreted by PBMC from MG patients were not different from those produced by those from normal controls. IgG secretion was, however, correlated with the secretions of IL-5 and IL-6 in MG. Spontaneous B cell activation was suspected in patients with MG.     

体外诱导人外周血单个核细胞向肝样细胞的分化

背景：外周血具有收集方便、供者不需麻醉、无骨髓穿刺痛苦等优点,利用人外周血干细胞移植的方法来获得转分化后的肝样细胞临床应用前景较好。 目的：探讨人外周血单个核细胞在肝细胞生长因子及成纤维细胞生长因子4诱导下分化为肝样细胞的可行性。 设计、时间及地点：细胞学体外对照观察,于2007-05/10在中国科学院大连化物所1806组细胞培养室和大连医科大学附属第一医院中心实验室完成。 材料：外周血由大连市红十字血液中心11名健康志愿献血者提供。肝细胞生长因子、巨噬细胞集落刺激因子、成纤维细胞生长因子4为Peprotech公司产品。 方法：采集志愿者外周血,密度梯度离心法分离外周血单个核细胞,加入RPMI1640液培养2 h后,去除未贴壁细胞。将贴壁细胞分为4组：对照1组加入含140 μmol/L β-巯基乙醇、体积分数为0.1胎牛血清的RPMI1640液培养6 d;对照2组、肝细胞生长因子组、成纤维细胞生长因子组均在其基础上向RPMI1640液中加入5 μg/L巨噬细胞集落刺激因子、0.4 μg/L白细胞介素3。洗涤后,对照组换用含胎牛血清的RPMI1640培养液继续培养20 d,肝细胞生长因子组在其基础上加入20 μg/L肝细胞生长因子,成纤维细胞生长因子组加入10 μg/L成纤维细胞生长因子4。 主要观察指标：诱导分化过程中细胞形态学变化,免疫荧光检测诱导后甲胎蛋白及白蛋白的表达。 结果：刚分离的单个核细胞形态呈较均一的圆形,活细胞率>95%,培养2 h后贴壁细胞呈毛糙的圆形,6 d后肝细胞生长因子组、成纤维细胞生长因子组细胞呈集落样生长,细胞趋向融合,对照组细胞未形成集落,呈单个细胞生长。经诱导培养后,4 d时部分细胞体积变大,向类圆形细胞转变,随培养时间延长,类圆形细胞比例增加,至20 d后细胞数量逐渐减少;对照组培养20 d后多数细胞呈梭形或纤维样,少部分细胞呈圆形。诱导培养后第6,10,14,20天,肝细胞生长因子组、成纤维细胞生长因子组甲胎蛋白、白蛋白均呈阳性表达,对照组各时间点均呈阴性表达。 结论：在肝细胞生长因子及成纤维细胞生长因子4诱导条件下,人外周血单个核细胞具有向肝样细胞分化的潜能。 关键词：外周血单个核细胞;肝细胞生长因子;成纤维细胞生长因子;肝细胞;分化     

Immunomodulatory effect of peripheral benzodiazepine receptor ligands on human mononuclear cells.

Immunomodulatory effect of ligands active at the peripheral benzodiazepine receptor (PBR) was examined in human peripheral blood mononuclear cells (PBMC). Ro5-4864, PK11195 and diazepam suppressed phytohemagglutinin (PHA) and concanavalin A (ConA) induced proliferation of PBMC. All three ligands inhibited interleukin-3-like activity (IL-3-LA) secretion, while the production of interleukin-2 (IL-2) was inhibited by Ro5-4864 and diazepam only. The selective central benzodiazepine ligand clonazepam did not affect the cellular immune functions examined. Our results indicate an in-vitro immuno-suppressive activity of peripheral and mixed, but not central type benzodiazepine ligands.     

Surface phenotypes of peripheral blood mononuclear cells in multiple sclerosis

Using monoclonal antibodies the subpopulations of mononuclear cells were studied in peripheral blood in patients with multiple sclerosis in active phase of the disease. The results suggested the following conclusions: 1. In the active phase of multiple sclerosis (during exacerbations and slowly progressive course) a rise occurs in the proportions of Ia-positive and M1-positive cells without changes in the proportions of T4 and T8 lymphocytes and in their mutual ratio. 2. No differences were observed in the percent of peripheral blood mononuclear cells between all patients in the active phase of the disease and those with slowly progressing disease. 3. During immunomodulatory treatment (alternating administration of prednisone and levamisole) a rise was observed in the percent of suppressor cells T8/Ia-positive, and a fall in the T4/T8 suppressor cell ratio.     

3H-spiroperidol binding to human peripheral mononuclear cells: methodological aspects.

3H-spiroperidol binding to lymphocytes has been proposed as a vulnerability marker for schizophrenia. However, the biological significance and even existence of this "binding site" are still in controversy. Therefore, the present study reevaluated methodological details using a filtration binding assay. The results indicated that some well-known, but obviously uncontrolled pitfalls might contribute to this controversy [e.g., unspecific filter binding, which increased in the presence of (+)-butaclamol, or a variable amount of contaminating granulocytes). Moreover, due to an atypically shaped saturation curve, different mathematical methods to analyze the data were used and compared. The present data should help us to understand the biological relevance of this marker, as viewed in different laboratories.     

阿尔茨海默病患者外周血OSTM1表达的变化

目的观察阿尔茨海默病(AD)患者与健康对照组外周血单核粒细胞中骨硬化症相关蛋白1(OSTM1)表达的差异并探讨其意义。方法 AD患者与对照组各30例,采用Ficoll-Hypaque密度梯度离心法分离人外周血单核粒细胞,培养24h收集细胞。细胞爬片后,采用免疫荧光细胞化学技术,观察OSTM1在单核粒细胞中分布并比较两组荧光信号强弱;蛋白印迹法(Western blotting)检测OSTM1蛋白表达水平;实时定量多聚链式反应(RT-PCR)法检测OSTM1mRNA表达水平。结果 1OSTM1表达情况:人外周血中表达OSTM1主要集中在胞浆,与对照组比较,AD组细胞荧光信号明显增强且分布范围扩大;2两组OSTM1在蛋白表达水上的平差异:AD组OSTM1表达(0.53!0.04)较对照组(0.36!0.03)比较差异有统计学意义(P<0.05);3两组在OSTM1基因水平表达情况:AD组与对照组OSTM1 mRNA分别为0.75!0.13和0.32!0.11,AD组OSTM1 mRNA表达水平与对照组相比,差异有统计学意义(P<0.05)。结论 AD组OSTM1在蛋白、基因水平较对照组均明显升高,推测OSTM1在AD发病机制中具有关键作用。     

Pharmacological modulation of the pontine micturition center.

Previous studies have indicated that an area of the rostral pontine tegmentum known as the 'pontine micturition center' (PMC) plays an essential role in the regulation of lower urinary tract function. The present pharmacologic experiments were conducted on either decerebrate unanesthetized or chloralose anesthetized cats to identify the location of the PMC and to examine the neurotransmitter mechanisms controlling micturition. Microinjections of excitatory and inhibitory amino acids were made at stereotaxic coordinates P1 to P3, L2 to L3, H0 to H-4 where electrical stimulation with trains of pulses (2-30 V, 80-120 Hz and 50-300 ms train duration) elicited short latency (less than 2 s) bladder contractions or voiding. Injections of L-glutamate (L-GLUT) (20-130 nmol) or DL-homocysteic acid (DLH) (20-100 nmol) into the region of the locus coeruleus or parabrachial nucleus elicited voiding as well as an increase in the frequency or amplitude of isovolumetric bladder contractions. In some anesthetized animals, L-GLUT and DLH also had mixed excitatory-inhibitory or pure inhibitory effects. Injections of muscimol (9-70 nmol) depressed rhythmic bladder activity, increased the bladder volume for inducing micturition or completely abolished the voiding induced by bladder filling. The inhibitory effects of muscimol were reversed by microinjections of bicuculline methiodide (BCMI) (3-22 nmol). Injections of BCMI (1-1.5 nmol) into untreated cats stimulated bladder activity and lowered the bladder volume for inducing micturition. It is concluded that: (1) neurons in the rostral pons are an essential component of the micturition reflex pathway, (2) several populations of neurons located in the region of the locus coeruleus complex and parabrachial nucleus contribute to the functions of the PMC, and (3) PMC neurons are under a tonic GABAergic inhibitory control which regulates the micturition threshold and in turn regulates bladder capacity.     

Blunted beta-adrenergic responsivity of peripheral blood mononuclear cells in endogenous depression. Isoproterenol dose-response studies

Previous studies of peripheral blood mononuclear cells isolated from drug-free, hospitalized patients with endogenous major depression have demonstrated a diminished adenosine 3',5'-monophosphate (cyclic AMP) response to single concentrations of isoproterenol as compared with that obtained from normal control subjects. We now report results of isoproterenol dose-response studies that indicate lower basal levels of cyclic AMP as well as diminished cyclic AMP levels in response to isoproterenol stimulation at concentrations ranging from 10(-10) to 10(-5) mol/L in drug-free, hospitalized patients with endogenous depression. The major factor responsible for the diminished cyclic AMP production in the depressed patients was a loss of receptor sites capable of cyclic AMP production. Taken together with our previously reported finding that beta-adrenergic antagonist binding was normal in peripheral blood mononuclear cells obtained from depressed patients, the results of the dose-response studies suggest a loss of receptor function (desensitization) rather than a diminished number of receptor binding sites (down-regulation) as the underlying mechanism. Potential explanations for beta-adrenergic desensitization and its implications for the catecholamine hypothesis of depressive disorders are discussed.     

重症肌无力患者外周血单个核细胞白细胞介素6活性变化

探讨白细胞介素 6(IL -6)在重症肌无力 (MG)发病机理中的作用及其临床意义。　　方法　采用3H -TdR掺入法检测 40例正常对照及 42例MG患者外周血单个核细胞IL -6分泌活性。　　结果　MG患者组IL -6活性显著高于正常对照组 ,且IL -6活性变化与乙酰胆碱受体抗体产生以及与MG临床类型、病情、预后密切相关。　　结论　IL -6在MG发病机理中起着重要作用 ,检测IL -6活性对区分MG临床类型、判断病情、推测预后、指导治疗有重要的参考价值。     

Increased interleukin-1 production by peripheral blood mononuclear cells in patients with multiple sclerosis.

The production of interleukin-1 (IL-1) by peripheral blood mononuclear cells (MNC) was assessed in patients with relapsing multiple sclerosis (MS) in both the active and inactive phase, in chronic progressive MS patients, in other neurological diseases, and in healthy subjects. Production was determined by measuring the IL-1 concentration in cultures with MNC supernatants using enzyme-linked immunosorbent assay (ELISA). IL-1 in sera of MS patients and healthy subjects also was investigated. MNC IL-1 alpha production was significantly higher in MS patients (180.2 +/- 177.5 pg/ml) than in healthy subjects (66.2 +/- 66.0 pg/ml) (P less than 0.05). Relapsing MS patients in the active phase had significantly higher MNC IL-1 alpha concentrations (360.1 +/- 130.0 pg/ml) than normal subjects (P less than 0.001), but MNC IL-1 alpha production in patients with relapsing MS in the inactive phase (65.3 +/- 52.8 pg/ml) or chronic progressive MS (80.9 +/- 71.9 pg/ml) was not increased significantly. MNC IL-1 beta production in MS patients was not elevated significantly. IL-1 alpha and -1 beta were not detected in sera of MS patients. The correlation between increased IL-1 alpha production and the clinical course of MS suggests that activated MNC may play a role in the pathogenesis of MS.     

Expression of costimulatory molecules on peripheral blood mononuclear cells in multiple sclerosis

OBJECTIVE: To determine whether there was aberrant expression of costimulatory molecules and their receptors on peripheral blood mononuclear cells of MS patients and healthy individuals and whether expression correlated with disease status. METHODS: Forty-six patients with MS and 29 healthy individuals were analyzed by direct 2-color immunofluorescence flow cytometry for expression of CD80, CD86, CD28 and CTLA-4 on T and B cells and monocytes. RESULTS: Expression of CD80 on CD4+ T cells was upregulated in progressing MS patients compared to stable MS patients and controls. Marked increase in the expression of CD80 and CD86 was seen on both CD4+ and CD8+ T cells from an MS patient with rapidly progressing disease. No difference in the expression of the costimulatory molecules or their ligands was seen between IFN-beta treated and non-treated MS patients. CONCLUSIONS: Aberrant expression of costimulatory molecules occurs on T lymphocytes in MS patients with progressing disease.     

Dopamine transporter immunoreactivity in peripheral blood mononuclear cells in amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a chronic progressive neuromuscular disorder of unknown etiology, characterized by weakness, muscle wasting, fasciculations, and increased reflexes, with conserved intellect and higher functions. The neuropathology of ALS is mostly confined to damage of the motor neurons in the cerebral cortex, some motor nuclei of the brainstem, and anterior horns of the spinal cord. However, there is evidence for the involvement of other neuronal systems in the disease. In particular, damage of the dopamine neurons has been shown by neurochemical and imaging studies in the brain and spinal cord of ALS patients. Recent reports suggest that peripheral blood mononuclear cells (PBMC) may represent a useful in vivo model to study neurochemical alterations that occur in neurodegenerative disorders. Here we demonstrate the significant reduction of dopamine transporter immunoreactivity in PBMC of patients affected by ALS with respect to healthy subjects. These results extend our knowledge of damage of the dopamine system in ALS to peripheral cells. Thus, the original concept of ALS as an isolated degeneration of motor neurons seems to extend to a more widespread understanding of the disease with involvement of other neuronal systems in the central as well as peripheral nervous system.