Differential gene expression profiles of peripheral blood mononuclear cells in childhood asthma

Objective: Asthma is a common childhood disease with strong genetic components. This study compared whole-genome expression differences between asthmatic young children and healthy controls to identify gene signatures of childhood asthma. Methods: Total RNA extracted from peripheral blood mononuclear cells (PBMC) was subjected to microarray analysis. QRT-PCR was performed to verify the microarray results. Classification and functional characterization of differential genes were illustrated by hierarchical clustering and gene ontology analysis. Multiple logistic regression (MLR) analysis, receiver operating characteristic (ROC) curve analysis, and discriminate power were used to scan asthma-specific diagnostic markers. Results: For fold-change>2 and p?<?0.05, there were 758 named differential genes. The results of QRT-PCR confirmed successfully the array data. Hierarchical clustering divided 29 highly possible genes into seven categories and the genes in the same cluster were likely to possess similar expression patterns or functions. Gene ontology analysis presented that differential genes primarily enriched in immune response, response to stress or stimulus, and regulation of apoptosis in biological process. MLR and ROC curve analysis revealed that the combination of ADAM33, Smad7, and LIGHT possessed excellent discriminating power. Conclusions: The combination of ADAM33, Smad7, and LIGHT would be a reliable and useful childhood asthma model for prediction and diagnosis.     

应用基因芯片技术研究原发性胆汁性肝硬化患者外周血单个核细胞基因表达谱特征

目的 应用基因芯片技术分析原发性胆汁性肝硬化患者外周血单个核细胞的基因表达谱特征.方法 选取9例原发性胆汁性肝硬化患者、9名健康对照为研究对象,提取其外周血单个核细胞总RNA,进行人类全基因组寡核苷酸芯片(约22000个基因)检测,筛选出差异表达基因及相关信号通路.结果 原发性胆汁性肝硬化患者外周血单个核细胞中差异表达的基因共有79个,其中21个表达上调,58个表达下调.这些基因对应着27条信号通路,其中有6条通路参与了免疫调控及细胞凋亡过程,分别是自然杀伤细胞介导的细胞毒通路、Toll样受体信号通路、抗原加工提呈通路、细胞因子相互作用通路、T细胞受体信号通路、细胞凋亡通路.结论 原发性胆汁性肝硬化患者外周血单个核细胞中存在特有的差异表达基因,针对这些基因及相关信号通路的进一步研究,将为探索发病机制和寻找生物标志物提供新的方向.     

运用基因芯片初步研究类风湿关节炎患者外周血单个核细胞基因表达谱特征

目的运用基因芯片技术研究类风湿关节炎(RA)基因表达谱特征,从全基因组角度探讨RA相关致病基因。方法提取5例活动期RA患者和3名健康对照者外周血单个核细胞(PBMC)中总RNA,Illumina寡核苷酸基因芯片分析每个样本的48 000个基因变化。反转录-聚合酶链反应(RT-PCR)验证芯片结果。结果RA患者与健康对照者相比2倍上调基因122个,2倍下调基因29个。分子及生物功能分析显示上调表达的基因涉及传递蛋白、蛋白翻译合成、代谢、转录调控、信号转导、趋化因子、细胞周期和凋亡相关基因等种类。结论RA涉及多环节的病理过程,基因芯片技术有利于进一步揭示RA分子机制,发现新的治疗靶标。     

Reduced expression of cellularfos gene in peripheral blood mononuclear cells of patients with primary biliary cirrhosis

Abnormal expression of certain cellular oncogenes in peripheral blood mononuclear cells (PBMC) is associated with several autoimmune disorders and is thought to reflect a pathologically activated state in various lymphocytic subpopulations. Using the Northern blot hybridization technique, we studied the expression of cellular (c-)fos, c-myc and N-ras genes in PBMC isolated from patients with primary biliary cirrhosis (PBC) and normal control subjects. The expression of c-fos gene was reduced significantly in patients with PBC, while there was no significant difference between the two groups in the expression of c-myc and N-ras genes. Stimulation of PBMC with mitogens in vitro increased c-fos gene expression markedly to a similar extent in both normal subjects and in patients with PBC. Furthermore, the reduced expression of c-fos gene recovered significantly in all the patients examined after treatment with ursodeoxycholic acid (UDCA). These results suggest that the reduced expression of c-fos gene in PBMC is a reversible functional manifestation of PBC and that UDCA therapy for PBC may improve abnormal lymphocyte function.     

RA患者易感基因肽酰基精氨酸脱亚氨酶4mRNA的表达及其意义

目的检测酰基精氨酸脱亚氨酶4（PAD14）在类风湿关节炎（RA）患者外周血单个核细胞（PBMC）mRNA的表达，分析其与RA临床相关指标的相关性，探讨PAD14在RA发病机制中的作用。方法采用反转录聚合酶链反应（RT—PCR）实时荧光定量法对RA组（53例）、骨关节炎（OA）组（27例）及正常对照组（30名）PBMC中PAD14 mRNA表达进行检测，并分析其与抗环瓜氨酸（CCP）抗体、疾病活动关节评分28（DAS28）、患者病程的关系。结果RA组PAD14 mRNA表达明显高于OA组和正常对照组，差异有统计学意义（P〈0．05）；PAD14 mRNA的表达在OA组、正常对照组差异无统计学意义（P〉0．05）。RA组中PAD14 mRNA表达与抗CCP抗体和DAS28评分水平呈明显正相关（r=0．452．0．653，P均〈0．05），与病程呈负相关（r=-0．68，P〈0．05）。结论RA组PAD14 mRNA表达显著增高，并与DAS28评分和抗CCP抗体水平呈明显正相关，与病程呈负相关。提示RA患者PAD14 mRNA表达明显增高，与疾病活动程度和自身免疫功能异常相关。     

Toll-like receptor expression in lupus peripheral blood mononuclear cells

OBJECTIVE: To investigate expression of members of the Toll-like receptor (TLR) family in peripheral blood mononuclear cells (PBMC) in patients with systemic lupus erythematosus (SLE). METHODS: We analyzed PBMC from 14 patients with SLE and 15 healthy subjects. The surface expressions of TLR2 and TLR4 and intracellular expression of TLR9 on PBMC were analyzed by flow cytometry. RESULTS: Although TLR4 expressions on CD14+ monocytes were not significantly different between healthy subjects and patients with SLE, TLR2 expressions on monocytes were reduced in patients with SLE compared to healthy subjects. Intracellular TLR9 expression levels of CD19+ B lymphocytes were significantly elevated in patients with SLE. However, the TLR9 expression levels of plasmacytoid dendritic cells were not significantly different between these patients and healthy subjects. CONCLUSION: Our results show that human peripheral blood B cells express TLR9 and that its expression is increased in patients with SLE. This upregulated expression of TLR9 in B cells may be related to the abnormal B cell hyperactivity in patients with SLE.     

A gene expression signature for recent onset rheumatoid arthritis in peripheral blood mononuclear cells

BACKGROUND: In previous studies the presence of a distinct gene expression pattern has been shown in peripheral blood cells from patients with autoimmune disease. OBJECTIVE: To determine whether other specific signatures might be used to identify subsets of these autoimmune diseases and whether gene expression patterns in early disease might identify pathogenetic factors. METHODS: Peripheral blood mononuclear cells were acquired from patients with rheumatoid arthritis (RA) and analysed by microarrays containing over 4300 named human genes. Patients with RA for <2 years were compared with subjects with longstanding RA (average duration 10 years) and with patients with other immune or autoimmune diagnoses. RESULTS: Cluster analyses permitted separation of the patients with early RA (ERA) from those with longstanding disease. Comparison with other patient groups suggested that the ERA signature showed some overlap with that seen in the normal immune response to viral antigen as well as with a subset of patients with systemic lupus erythematosus. CONCLUSIONS: The ERA signature may reflect, in part, a response to an unknown infectious agent. Furthermore, shared features with some lupus patients suggest that common aetiological factors and pathogenetic pathways may be involved in these two autoimmune disorders.     

Blood-sample processing for the study of age-dependent gene expression in peripheral blood mononuclear cells

Although most new biogerontological studies seeking to identify longevity candidate genes and factors involved in successful human aging are population based, and likely to involve the collection of blood from extremely old individuals, to our knowledge no unified protocols have yet been published to describe a methodology permitting the simultaneous generation of different kinds of biological specimens derived from a single source of a very small volume of peripheral blood. Here we describe a method permitting the simultaneous generation of plasma, RNA, DNA, protein, fixed lymphocytes, and frozen blood aliquots from a single 10- to 30-ml blood sample obtained from donors of any age (10-102 years old), and we show that the quality and quantity of DNA, RNA, protein, and fixed lymphocytes obtained do not vary significantly with age. As is frequently observed, the older individuals have higher plasma proportions.     

Monoglycerides induce apoptosis in human leukemic cells while sparing normal peripheral blood mononuclear cells

A major drawback of the current antineoplastic treatments is their lack of specificity toward cancer cells, because they are most often cytotoxic to normal cells, thus creating related side effects. Hence, the identification of new apoptosis-inducing agents, specifically targeting malignant cells while sparing their normal counterparts, is of crucial interest. We show here that monoglycerides, a family of lipids consisting of a single fatty acid attached to a glycerol backbone, induce cell death in several human leukemic cell lines. Importantly, treatment of primary leukemic cells, obtained from B-cell chronic lymphocytic leukemia patients, resulted in rapid apoptosis. In striking contrast, resting or activated human peripheral blood mononuclear cells from healthy individuals were resistant to the same treatment. Therefore, these compounds could represent potential antileukemic drugs or could allow for the design of novel therapeutic agents applied to leukemia.     

Cytokine and antioxidant gene profiles from peripheral blood mononuclear cells of Pelibuey lambs after Haemonchus contortus infection

The expression profiles of cytokines and antioxidant genes were determined from an experimental infection with H. contortus in Pelibuey lambs. The infection was followed for 34 days (d) to determine the number of eggs per gram (epg) and the packed cell volume (PCV). Differential white cell counts and expression profile estimations of IL‐2, IL‐4, IL‐5, IL‐6, IL‐8, IL‐10, IL‐13, FCεR1A, GPX and SOD1 were determined at 0 hour, 4 hours, 2 days and 14 days post‐infection (PI) in infected and control groups. Comparison of the fold change between 0 and 4‐hours, 4‐hours and 2‐days and 2‐ and 14‐days periods was performed. Significant differences (P<.05) between epg (>2000) and PCV (>30%) were determined after 21 days and were also observed with regard to monocyte and lymphocyte cells after 2 and 7 days PI. At 0 hour and 14 days PI, the GPX and IL‐2 genes showed a 0.37‐ and 0.49‐fold decrease in expression, respectively. In contrast, upregulation was observed at 4 hours of IL‐8 (2.58) and FCεR1A (2.71), at 2 days for IL‐4 (2.14) and IL‐8 (4.02) and at 14 days for IL‐2 (0.41), IL‐10 (2.35) and FCεR1A (2.28). The comparison between the intervals of infection showed high expression values against H. contortus infection in Pelibuey sheep after the 2nd period of PI involving a dichotomy T cells.     

Chronic stress down-regulates growth hormone gene expression in peripheral blood mononuclear cells of older adults

"Pituitary" peptides are produced in both endocrine and immune cells. Acute and chronic stress can alter pituitary peptide secretion and might also influence neuroendocrine gene expression in human immune cells. We reasoned that, in Alzheimer caregivers, the chronic stress of caregiving would impact on the sympathetic-adrenal-medullary and hypothalamicpituitary-adrenal axis possibly leading to alterations in GH mRNA in their peripheral blood mononuclear cells (PBMCs). Therefore, we evaluated 10 caregivers and 10 controls subjects using a math and speech stress protocol to determine their neuroendocrine profile and to evaluate any relationship with mononuclear cell GH mRNA levels simultaneously acquired and then evaluated by a quantitative competitive RT-PCR technique. We found a significant (p<.0001) decrease 50% in GH mRNA levels in cells from caregivers. Plasma ACTH and norepinephrine levels were negatively correlated with GH mRNA levels, suggesting their possible role in the down-regulation of mononuclear cell GH gene expression. These observations support the hypothesis that experiences associated with caregiving alter the brain's autonomic nervous system and neuroendocrine control of the hypothalamic-pituitary axis. These and perhaps other influences may then produce altered GH gene expression in mononuclear cells of chronically stressed individuals. It is tempting to speculate that the decreased GH mRNA that we found in these chronically stressed caregivers was partially responsible for their poor response to influenza vaccine and their delayed wound healing.