哮喘儿童细胞因子信号转导抑制因子mRNA表达与Th1/Th2的关系

目的：细胞因子信号转导抑制因子(SOCS)对JAK-STAT途径的细胞因子如白介素、干扰素等的调节起重要作用,目前SOCS与哮喘的关系仍在研究中。本研究观察SOCS1和SOCS3 mRNA在哮喘儿童外周血单个核细胞（PBMC）中的表达水平与CD4+ T细胞IFN-γ/IL-4平衡及特异性IgE（sIgE）的关系。方法：采集44例4~14岁过敏性哮喘患儿及30例健康儿童PBMC,分别用流式细胞仪分析CD4+ T细胞IFN-γ/IL-4比值,另提取总RNA,采用SYBR Green I逆转录荧光定量PCR的方法检测每组SOCS1和SOCS3 mRNA的表达。结果：哮喘组患儿外周血IFN-γ阳性的CD4+T细胞百分比［（15.7±2.0）%］及IFN-γ/IL-4比值（3.4±1.5）均低于对照组［分别为（19.1±2.7)%、4.8±2.9］;而SOCS1 mRNA（⊿Ct值11.1±1.9）表达显著高于对照组（⊿Ct值12.6±2.8）。两组儿童SOCS1 mRNA表达均与外周血分泌IFN-γ的CD4+ T细胞百分比呈负相关（P<0.05）。SOCS1和SOCS3与sIgE均无相关性。结论：SOCS1 mRNA在哮喘组患儿外周血中高表达,并与Th2占优势的免疫失衡有关。     

Changes in humoral responses to β-lactoglobulin in tolerant patients suggest a particular role for IgG4 in delayed, non-IgE-mediated cow''s milk allergy

The major cow's milk allergen beta-lactoglobulin (beta-LG) is relatively resistant to enzymatic degradation and may therefore be involved in non-immunoglobulin (Ig)E-mediated cow's milk allergy (CMA) with delayed gastrointestinal symptoms. Serum levels of beta-LG-specific IgG(1), IgG(4), IgE, and IgA were compared in clinically reactive and tolerized IgE-mediated and non-IgE-mediated CMA with delayed gastrointestinal symptoms (n = 29) and controls (n = 10). Tolerance was associated with decreased beta-LG-specific IgE, IgG(1), and IgG(4) levels in both patient groups. However, the significantly increased beta-LG-specific IgG(4) levels in clinically reactive non-IgE-mediated CMA patients, and its median 36-fold reduction in tolerant patients, suggested a possible immunopathological role for IgG(4) in delayed CMA. Similarly, the significantly increased beta-LG-specific IgE levels in IgE-mediated CMA patients were decreased 44-fold in tolerant patients. The tolerant patients had apparently shifted the humoral immune response from a beta-LG-specific IgE- and/or IgG(4)-dominated immune response to an IgA-dominated immune response as the IgA/IgE or IgA/IgG(4) ratios increased 90- and 15-fold in tolerant IgE-mediated-, and non-IgE-mediated CMA patients, respectively. Thus, the marked difference in beta-LG-specific Ig ratios suggested a tolerance-induced inhibition of a Th(2)-type of immune response with significantly increased IgA dominance in both CMA patient groups.     

Responses of CD4(+) CD25(+) Foxp3(+) and IL-10-secreting type I T regulatory cells to cluster-specific immunotherapy for allergic rhinitis in children

We investigated the effects of cluster specific immunotherapy (SIT) with Dermatophagoides pteronyssinus (Der p) on CD4(+) CD25(+) Foxp3(+) Treg cells and IL-10-secreting type I T regulatory (Tr1) cells in Der p-sensitized children with allergic rhinitis (AR). We performed a prospective randomized study involving 46 children (aged 8-13 yr), of whom 25 children received Der p-SIT + pharmacotherapy and 21 received only pharmacotherapy, over a period of 1 yr. Prior to and at end of treatment, CD4(+) CD25(+) Foxp3(+) Treg cells and allergen-specific IL-10(+) IL-4(-) , IFN-γ(+) IL-4(-) , and IL-4(+) IFN-γ-CD4(+) T cells were measured by flow cytometry. Similarly, IL-4, IFN-γ, and IL-10 in supernatants from allergen-stimulated peripheral blood mononuclear cell (PBMC) cultures were measured by ELISA, and the suppressive effect of CD4(+) CD25(high) T cells on cell proliferation and cytokine release was estimated from both groups. Allergen-specific serum IgE and IgG4 were also assessed at the beginning and end of treatment by RAST and ELISA, respectively. The levels of allergen-specific Tr1 cells, IgG4, and allergen-induced IL-10 synthesis from PBMC cultures were significantly increased after SIT for 1 yr compared with baseline levels (p < 0.001 for all), with significant correlation between increased levels of Tr1 cells and improvements in nasal symptoms (r = 0.48, p < 0.05). In contrast, the levels of CD4(+) CD25(+) Foxp3(+) T cells, allergen-specific Th1 and Th2 cells, the production of IL-4 and IFN-γ, and the function of CD4(+) CD25(high) T cells were not altered in either group at the end of treatment. These data suggest that the up-regulation of Tr1 cells may play an important role in SIT and be a useful marker of successful SIT in AR patients.     

不同浓度地塞米松和甲泼尼龙对支气管哮喘患儿外周血单个核细胞免疫功能的影响

目的 观察不同浓度地塞米松(DEX)和甲泼尼龙(MP)对支气管哮喘患儿外周血单个核细胞(PBMC)中CD4+T淋巴细胞中2个功能性亚群(Th1/Th2)功能状态的影响.方法 选择2001年6月-2002年6月在重庆医科大学附属儿童医院就诊的15例哮喘患儿和14例健康体检儿童为研究对象.分为哮喘对照组、10-7mol·L-1DEX干预组、10-8mol·L-1DEX干预组、10-9mol·L-1DEX干预组、10-7mol·L-1MP干预组、10-8mol·L-1MP干预组、10-9mol·L-1MP干预组.清晨取其空腹静脉血5 mL,肝素抗凝,采用密度梯度离心法分离PBMC,加植物血凝素进行刺激培养,分别用10-7mol·L-1、10-8mol·L-1及10-9mol·L-1DEX或MP体外干预培养48 h.同期采集健康儿童空腹静脉血,同法分离培养.用ELISA法测定培养上清中γ干扰素(IFN-γ)、IL-4、IL-10及IL-12水平,并计算不同浓度DEX或MP对PBMC分泌细胞因子的抑制率.结果 1.哮喘对照组PBMC分泌IL-4水平显著高于健康对照组(P<0.05),IFN-γ/IL-4比值较健康对照组显著降低(P<0.05),2组间IFN-γ、IL-10及IL-12水平比较差异均无统计学意义(Pa>0.05).2.DEX和MP均可明显抑制哮喘患儿PBMC分泌IFN-γ、IL-4及IL-12,但对IL-10抑制作用差;与健康对照组比较,哮喘组DEX和MP抑制PBMC分泌IL-10的作用弱,差异有统计学意义(Pa<0.05).3.DEX和MP均以浓度依赖方式抑制哮喘患儿PBMC分泌IL-4,DEX在10-9mol·L-1时有促进IL-4分泌的效应,MP在10-9mol·L-1时可抑制IL-4的分泌,二组比较差异有统计学意义(P<0.05). 若以IFN-γ/IL-4表示Th1/Th2间的平衡,则MP可恢复Th1/Th2平衡(P<0.05).结论 DEX和MP均可抑制Th1/Th2类细胞因子分泌,但MP有助于恢复Th1/Th2平衡,提示临床选择MP治疗支气管哮喘更有利.     

尘螨变应性哮喘患儿外周血T淋巴细胞中协同刺激分子和细胞因子的异常表达及其意义

目的分析变应原刺激前后变应性哮喘患儿外周血T淋巴细胞表面协同刺激分子及胞内细胞因子表达的变化,探讨CD28家族不同协同刺激信号在变应性哮喘免疫病理机制中的作用。方法选取尘螨变应性哮喘患儿(哮喘组)和健康儿童(健康对照组)各30例,密度梯度离心法分离其外周血单个核细胞,应用免疫荧光标记和流式细胞术检测尘螨刺激前后体外培养的CD4+T淋巴细胞表面协同刺激分子CD28、可诱导共刺激分子(ICOS)和细胞毒T淋巴细胞相关抗原4(CTLA-4)的表达,运用细胞内染色技术检测CD4+T淋巴细胞内细胞因子γ干扰素(IFN-γ)、IL-4和IL-13的表达。并运用统计学方法比较哮喘组和健康对照组之间的差异。结果哮喘组患儿外周血CD4+T淋巴细胞表面CD28和ICOS的表达与健康对照组比较差异均无统计学意义(Pa>0.05),而CTLA-4的表达显著降低(P<0.01);细胞内IFN-γ表达水平显著升高(P<0.000 1),而IL-4和IL-13表达水平无明显变化(Pa>0.05)。经尘螨刺激后,体外培养的哮喘患儿外周血CD4+T淋巴细胞表面ICOS的表达较健康对照组儿童显著上调(P<0.000 1),CD28和CTLA-4的表达则无明显变化(Pa>0.05);细胞内细胞因子IL-4和IL-13的表达显著上调(Pa<0.000 1),而IFN-γ的表达则无明显差异(P>0.05)。结论变应性哮喘患儿外周血存在组成性的CTLA-4的表达下调和细胞因子IFN-γ的表达上调,介导Th1型细胞的异常活化;而变应原尘螨的刺激又介导了ICOS依赖的Th2型细胞的分化,导致Th1/Th2失衡。     

连续性血液净化对多器官功能障碍幼猪T辅助细胞免疫功能影响的机制研究

目的 研究连续性血液净化(CBP)对多器官功能障碍(MODS)幼猪在疾病不同时期外周血Th1、Th2细胞数量及其细胞培养上清液中细胞因子(IFN-γ、TNF-α、IL-12,IL-10、IL-4)水平的影响.方法 24只幼猪随机分为对照组和CBP干预组,每组12只.内毒素诱导多器官功能障碍.CBP主要采用连续性静脉静脉血液透析滤过(continuous venovenous hemodiafiltration,CVVHDF).造模完成后为0 h,MODS对照组在造模后0、2、4、6 h进行检测;血液净化干预组造模后即开始CVVHDF,分别在0 h及CVVHDF干预2 h、4 h、6 h进行检测.在各时点采用免疫磁珠分离两组幼猪外周血T辅助细胞,采用流式细胞仪检测Th1、Th2细胞数目,ELISA检测各时点T细胞培养上清液中IFN-γ、TNF-α、IL-12、IL-10、IL-4的表达水平.结果 幼猪诱导MODS模型后,辅助性T细胞分泌炎症因子TNF-α明显上升,而促进Th1细胞分化的因子IL-12和Th1细胞因子IFN-γ和调节因子IL-10呈下降趋势,Th2细胞因子IL-4则没有明显的变化,Th1/Th2比值下降;经过CVVHDF治疗,TNF-α和IL-4减少,IL-12、IFN-γ和IL-10在CVVHDF6 h后明显上升,Th1/Th2比值上升.结论 MODS幼猪模型经过CVVHDF治疗,有助于炎症因子的清除,增加机体的细胞免疫功能,恢复Th1/Th2的平衡.     

支气管哮喘患儿血清白细胞介素-4、白细胞介素-5和γ干扰素水平检测的意义

目的 探讨支气管哮喘患儿血清IL-4、IL-5和γ干扰素(IFN-γ)水平及其在支气管哮喘发病中的作用.方法 选取30例门诊随诊的哮喘缓解期患儿作为哮喘组,30例同期健康体检儿童作为健康对照组,2组儿童均于清晨空腹取外周静脉血1.0 mL制备血清,应用ELISA法对2组儿童血清IL-4、IL-5和IFN-γ水平进行测定.结果 哮喘组患儿血清IL-4、IL-5水平均较健康对照组升高,差异有统计学意义(t= 2.48、2.63,Pa<0.05),但其血清IFN-γ水平较健康对照组显著降低(t= 3.02 ,P<0.01).结论 哮喘患儿存在Th1/Th2细胞免疫失衡,Th1免疫功能低下,IFN-γ下调可能是哮喘发病的重要原因,对哮喘有负向调节作用. 哮喘患儿同时存在Th2细胞免疫功能亢进,以IL-4、IL-5为代表的Th2免疫功能在儿童哮喘的发病中发挥正性促炎作用.     

Relation between stressful life events,neuropeptides and cytokines: results from the LISA birth cohort study

Stressful life events evidently have an impact on development of allergic diseases, but the mechanism linking stress to pathological changes of immune system function is still not fully understood. The aim of our study was to investigate the relationship between stressful life events, neuropeptide and cytokine concentrations in children. Within the LISAplus (Life style-Immune system-Allergy) study, blood samples from children of 6 yr of age were analysed for concentration of the neuropeptides vasoactive intestinal peptide (VIP), somatostatin (SOM), substance P (SP) and the Th1/Th2 cytokines interferon-γ (IFN-γ) and interleukin (IL)-4. Life events such as severe disease or death of a family member, unemployment or divorce of the parents were assessed with a questionnaire filled in by the parents. For 234 children, blood analysis and questionnaire data regarding life events were available. Children with separated/divorced parents showed high VIP levels and high concentrations of the Th2 cytokine IL-4 in their blood. Severe diseases and death of a family member were neither associated with neuropeptide levels nor with cytokine concentrations. Unemployment of the parents was associated with decreased IFN-γ concentrations in children’s blood but not with neuropeptide levels, whereas children experiencing concomitant severe disease and death of a family member had reduced SP blood levels. The neuropeptide VIP might be a mediator between stressful life events and immune regulation contributing to the Th2 shifted immune response in children with separated/divorced parents. Unemployment of the parents was associated with immune regulation in children on the basis of a still unknown mechanism whereas reduced SP levels seem to have no effect on immune regulation.     

头孢呋辛对哮喘儿童外周血单个核细胞Th1/Th2平衡的影响

目的研究头孢呋辛对哮喘儿童外周血单个核细胞Th1/Th2平衡的影响。方法采用流式细胞术检测哮喘和健康儿童外周血单个核细胞IFN-γ和IL-4水平,以及哮喘儿童外周血单个核细胞经头孢呋辛体外干预后的IFN-γ和IL-4水平。结果与健康儿童相比,哮喘患儿外周血单个核细胞的IFN-γ和IFN-γ/IL-4比值降低,差异有统计学意义(P<0.05);哮喘患儿外周血单个核细胞在体外与头孢呋辛(100 mg/L)孵育48 h后,IL-4水平升高,差异有统计学意义(P<0.05),而IFN-γ的变化无统计学意义(P>0.05);IFN-γ/IL-4比值则降低,差异有统计学意义(P<0.01)。结论哮喘患儿外周血单个核细胞以Th2(IL-4)占优势,Th1/Th2比值平衡失调;而头孢呋辛更加剧这一倾斜,不利于哮喘治疗。     

Defective tumor necrosis factor-α production in infants with cow’s milk allergy

As an aid to clarifying the role of immune mechanisms in the development of cow’s milk allergy (CMA) in suckling infants, we studied the capacity of peripheral blood mononuclear cells (PBMC) to produce tumor necrosis factor-α (TNF-α) in vitro. The study population consisted of 43 infants, aged 0.12–11.2 months; of these, 31 had challenge-proven cow’s milk allergy manifested with either skin or gastrointestinal symptoms or both. In addition, 12 healthy infants were studied as controls. The spontaneous, unstimulated and mitogen-induced production of TNF-α and interferon-γ (IFN-γ) by isolated peripheral blood leukocytes was evaluated. TNF-α and IFN-γ production of PBMC was significantly lower in infants with cow’s milk allergy than in healthy children. Our results indicate that, in infants with CMA, the function of TNF-α-producing cells is defective. This might disturb the development of oral tolerance and thereby lead to cow’s milk allergy. These results may help to clarify the etiopathology of CMA.     

Allergen-induced cytokine secretion in atopic and non-atopic asthmatic children

Atopic asthma is characterized by excessive T helper 2 (Th2)-like immunity to allergens in the bronchial mucosa. The Th2-cytokine interleukin (IL)-4 induces IgE production, while the Th2-cytokine IL-5 promotes eosinophilic inflammation in the airways of asthmatics. Most asthmatics are atopic, but a subgroup is non-atopic. We hypothesize that allergen-induced Th2, particularly IL-5, responses can be observed in peripheral blood in both atopic and non-atopic asthmatic children but not in healthy control children. The aim of the present study was to determine IL-4, IL-5, IL-9, IL-10, IL-13 and IFN-γ secretion induced from peripheral blood mononuclear cells (PBMC) by a broad panel of inhalant allergens (timothy, cat, birch, dog and house dust mite) in asthmatic children with and without sensitization. The study included 13 atopic asthmatic, 5 non-atopic asthmatic, and 12 non-atopic non-asthmatic children. PBMC were stimulated with allergens and cytokine production was measured with enzyme-linked immunosorbent assay (ELISA). Higher levels of cat and dog antigen-induced IL-5 release were more commonly observed in both atopic and non-atopic asthmatics than in controls. Children with atopic, but not non-atopic, asthma produced higher levels of allergen-induced IL-4 and IL-9 than controls. Non-atopic asthmatics produced more IL-10 than atopic asthmatics after cat stimulation. High levels of eosinophilia-associated IL-5 responses are induced by cat and dog allergen in both atopic and non-atopic asthmatic children. The Th2 cytokines IL-4 and IL-9 were associated only with atopic asthma, probably due to their IgE-inducing properties.     

Casein-specific immunoglobulins in cow''s milk allergic patient subgroups reveal a shift to IgA dominance in tolerant patients

Differences in casein-specific immunoglobulin (Ig) G-subclass and IgA serum levels between reactive and tolerant patients may hint at the immunopathogenesis during tolerance development in cow's milk allergy (CMA). alpha-, beta- and kappa-casein-specific IgG(1), IgG(4), IgE and IgA serum levels were compared in clinically reactive and tolerized IgE-mediated (n = 15) and non-IgE-mediated (n = 14) CMA with delayed gastrointestinal symptoms, using enzyme-linked immunosorbent assay (ELISA) and immunoblot techniques. The median anti-casein IgE levels in clinically reactive IgE-mediated CMA patients (n = 9) were 140- to 180-fold higher than in tolerized patients (n = 6) and 160- to 200-fold higher than in controls (n = 10). Median alpha-, beta- and kappa-casein-specific IgG(1) and IgG(4) levels were nine- to 60-fold higher in reactive patients and five- to 60-fold in tolerized patients. Clinical tolerance in IgE-mediated CMA was thus associated with decreased casein-specific IgE, IgG(4) and IgG(1), whereas serum IgA anti-alpha -, beta- and kappa-casein remained practically unaltered. Tolerized cow's milk protein (CMP)-sensitive atopic dermatitis had, in particular, decreased kappa-casein-specific IgG(1) levels, compared with clinically reactive patients. The ELISA levels to immunoblot correlation profile for the alpha-, beta- and kappa-casein-specific IgE suggested that the IgE-mediated CMA patients predominantly reacted to tertiary alpha- and beta-casein epitopes whereas the IgE in non-IgE-mediated patients reacted to linearized alpha-, beta- and kappa-casein epitopes. Clinical tolerance in non-IgE-mediated CMA patients (n = 9) was associated with a four- to 10-fold decrease in casein-specific IgE levels, accompanied by a five- to eightfold decrease in IgG(1) and five- to 60-fold decrease in IgG(4) levels, whereas casein-specific IgA levels remained unaltered. Thus, tolerance in both patient groups was characterized by a generalized decreased humoral immune response to caseins, which induced a functional shift to IgA dominance.     

T-cell signal transduction in children with cow's milk allergy – increased MAP kinase activation in patients with acute symptoms of cow's milk allergy

The precise immune mechanisms behind cow's milk allergy (CMA) are still unknown. Previously, the production of the cytokines TNF-α and IFN-γ in T cells from children with CMA has been shown to be decreased, and the production of IL-4 has been shown to be increased when compared to healthy children. As these aberrations in cytokine production may be associated with disturbances in cellular function, we investigated whether T-cell signal transduction is abnormal in children with CMA. For this purpose we evaluated the activation of the MAP kinase Erk2. Thirty-nine infants were included in the study. Of those with CMA, 13 had acute symptoms and 9 were free of symptoms due to a successful elimination diet at the time of the study. To activate T cells and to stimulate MAP kinase phosphorylation, peripheral blood mononuclear cells (PBMC) were incubated with Concanavalin A (ConA). The change in MAP kinase phosphorylation was measured by Western blotting. The increase in MAP kinase phosphorylation after stimulation with ConA for 5 min was significantly higher in cells from patients with acute symptoms of CMA than in cells from CMA patients free of symptoms or cells from healthy children. A time-course experiment showed that the change in MAP kinase phosphorylation was still increasing after 10 min incubation in cells from patients with acute symptoms of CMA. The increased MAP kinase activation was found to correlate positively with non-IgE mediated CMA in patients with acute symptoms of CMA.     

儿童1型糖尿病TH1/TH2免疫应答状况研究

目的研究儿童1型糖尿病T     

急性期川崎病γ干扰素组蛋白乙酰化改变及意义初探

目的探讨急性期川崎病(KD)患儿IFN-γ组蛋白乙酰化改变及其在KD免疫发病机制中的作用。方法以2015年2月至2016年6月深圳市儿童医院诊断并住院治疗的KD患儿为KD组,经IVIG治疗后为KD-IVIG组,KD组依冠状动脉有无损伤分为冠状动脉损伤(KD-CAL+)亚组和无冠状动脉损伤(KD-CAL-)亚组,于IVIG治疗前、后取血备检;同期体检的健康儿童为对照组。采用染色质免疫共沉淀法检测外周血CD4+T细胞IFN-γ组蛋白H3乙酰化水平及组蛋白乙酰化酶p300和去乙酰化酶HDAC1/2结合水平;流式细胞术检测外周血CD4+IFN-γ+细胞(Th1)比例及IFN-γ、p STAT4、p STAT5和T-bet蛋白表达水平;实时荧光定量PCR检测CD4+T细胞IFN-γ、IL-2Rα/β、IL-12Rβ1/2、IL-18Rα/β和ITLR4m RNA表达;ELISA检测外周血浆中IL-12、IL-2和IL-18浓度。结果 1KD患儿38例(男20例),年龄1~5.2岁;KD-CAL+亚组16例,KD-CAL-亚组22例。对照组32例(男17例),年龄1.2~4.9岁。KD组和对照组年龄、性别差异无统计学意义。2急性期KD患儿Th1细胞比例、IFN-γm RNA和蛋白表达及其组蛋白乙酰化水平显著高于对照组(P0.05),且KDCAL+亚组高于KD-CAL-亚组(P0.05),经IVIG治疗后明显恢复(P0.05)。3急性期KD患儿CD4+T细胞IFN-γ组蛋白乙酰化酶p300结合水平显著增加(P0.05),HDAC1/2结合水平明显降低(P0.05),p300/HDAC1/2明显高于对照组且与IFN-γ组蛋白H3乙酰化水平呈正相关(r=0.52;P0.05),经IVIG治疗后均有所恢复(P0.05)。其中KD-CAL+亚组p300结合水平及p300/HDAC1/2比值均高于KD-CAL-亚组(P0.05),而HDAC1/2结合水平则低于后者(P0.05)。4急性期KD患儿血浆IL-2、IL-12和IL-18浓度显著增高(P0.05),CD4+T细胞表面受体IL-2Rα/β、IL-12Rβ1/2、IL-18Rα/β和TLR4及其下游信号分子p STAT5、p STAT4、T-bet和My D88表达明显上调(P0.05),且KD-CAL+亚组前述各项均高于KDCAL-亚组(P0.05),经IVIG治疗后均有不同程度恢复(P0.05)。结论 IFN-γ组蛋白过度乙酰化可能是导致急性期KD患儿免疫功能紊乱的重要因素之一。     

短发夹RNA沉默Tim3分子表达对哮喘小鼠外周血Th1和Th17细胞的影响

目的：建立小鼠支气管哮喘模型,采用T 细胞免疫球蛋白黏蛋白分子-3（Tim-3）特异短发夹RNA（shRNA）沉默外周血单个核细胞（PBMCs）中Tim-3的表达,探讨Tim-3对辅助性T细胞1 (Th1)和Th17细胞分化的影响。方法：用卵白蛋白(OVA)致敏并激发建立哮喘小鼠模型,分离哮喘小鼠PBMCs,用 Tim-3特异的shRNA 片段沉默哮喘小鼠PBMCs中高表达的Tim-3 基因。Real-time PCR 和 Western blot 检测 Tim-3 的表达,流式细胞分析技术（FACS）检测Th1和Th17的比例;ELISA 检测细胞培养上清液中干扰素 γ（IFN-γ）,白介素-4（IL-4）,和 白介素-17（IL-17）的水平。结果：哮喘组小鼠PBMCs中Tim-3 mRNA表达明显升高; 使用特异Tim-3 shRNA沉默哮喘小鼠PBMCs后,沉默组PBMCs中Th1细胞比例显著升高,Th17细胞比例显著下降,与阴性对照组相比差异有统计学意义（P<0.01）;沉默组PBMCs培养上清液中IFN-γ明显升高,IL-17明显降低,与阴性对照组相比差异有统计学意义（P<0.05）, IL- 4水平无明显变化。结论：特异Tim-3 shRNA有效地沉默了Tim-3的表达,Tim-3表达的改变影响了T 细胞的分化。     

儿童重型再生障碍性贫血Th1/Th2细胞相关因子表达研究

目的利用流式细胞术探讨儿童重型再生障碍性贫血(简称"再障")患儿体内Th1和Th2细胞相关因子白细胞介素-2(IL-2)、白细胞介素4(IL-4)、白细胞介素-10(IL-10)和干扰素-γ(IFN-γ)的变化特点。方法 2014年11月至2015年8月,在郑州大学第一附属医院诊治的30例初诊重型再障患儿,同时选择正常体检儿童20名为对照组,采取其静脉血,分别用流式细胞术测定所取血样标本中IL-2、IL-4、IL-10和IFN-γ的含量。并用独立样本t检验的统计分析方法分析两组儿重外周血中上述4种细胞因子的差异是否有统计学意义,用ROC曲线评价上述4种细胞因子对于诊断重型再障的临床意义。结果(1)重型再障患儿外周血中IL-2、IFN-γ浓度分别为7.56±5.93(pg/mL)、8.67±3.72(pg/mL),均高于对照组的4.45±2.94(pg/mL)、4.89±2.86(pg/mL),差异具有显著性(P0.05);IL-4、IL-10浓度分别为4.48±3.28(pg/mL)、9.00±3.60(pg/mL).均低于对照组的7.36±7.02(pg/mL)、10.93±5.13(pg/mL),但差异无显著性(P0.05)。(2)重型再障患儿外周血中IL-2 ROC曲线下面积(AUC)为0.746(95%CI 0.697~0.816),具有中度诊断价值;IL-4的AUC为0.662(95%CI 0.597~0.716),为低诊断价值;IL-10的AUC为0.616(95%CI 0.458~0.774),为低诊断价值;IFN-γ的AUC为0.817(95%CI 0.697~0.978),具中度诊断价值。结论 IL-2、IFN-γ为负调控造血因子,参与再障发病机制,但是Th1和Th2细胞相关因子(IL-2、IFN-γ和IL-4、IL-10)的表达水平高低,对于儿童获得性再障的诊断价值比较有限,     

Levels of Th1 and Th2 cytokines in children with post-infectious bronchiolitis obliterans

BACKGROUND: Post-infectious bronchiolitis obliterans (BO) is a chronic obstructive airway disease associated with inflammation and fibrosis of the small airways; it is more common in children who have had acute viral bronchiolitis. No previous studies have reported the immune response of BO. Unbalanced Th1/Th2 immune response might be one of the risk factors for developing this illness. AIM: To compare the production of interferon (IFN)-gamma, interleukin (IL)-4 and IL-10 in peripheral blood mononuclear cell cultures in children with BO and in healthy children. METHODS: From March 2003 to October 2003, children with BO and healthy children were selected from the paediatric outpatient clinics in our centre. Peripheral blood was collected and mononuclear cells were separated and cultured (96 hours) with 1% phytohaemagglutinin stimulation. The supernatant was stored and cytokine levels were measured through ELISA. RESULTS: IFN-gamma, IL-4 and IL-10 levels were not significantly different between the groups studied. Family history of atopy was significantly associated with subjects with BO (p=0.02). CONCLUSIONS: Our results suggest that unbalanced peripheral blood Th1/Th2 immune response of children with post-infectious BO might not be associated with its pathophysiology. Further studies are required to better understand the role of risk factors, including viral genotype, viral load or tissue repair abnormalities in the development of post-infectious BO.