The effects of megadose methylprednisolone therapy on the immune system in childhood immune thrombocytopenia

Immune thrombocytopenia (ITP) is a frequently encountered disease in childhood. Recent reports pointed to the benefit of high-dose steroid in ITP treatment since it resulted in a better outcome in a shorter time than IV immunoglobulin therapy. In the authors' clinic, mainly after 1984, megadose methyl prednisolone (MDMP) has been used for ITP treatment. There is no report that includes an extensive immune system examination of megadose steroid effect in childhood ITP. The purpose of this study is to determine the effects of MDMP therapy on the immune system in childhood ITP for serum immunoglobulins, absolute lymphocyte and lymphocyte subclass counts, and in vitro blastogenic responses to mitogens. The authors have demonstrated that lymphocyte subtypes (CD3, CD4, CD8, CD19) and serum immunoglobulin G, A, M increased after a short-course MDMP therapy in comparison to pretreatment values in mixed group (acute + chronic) of childhood ITP patients. Also blastic transformation function of lymphocytes with Conca-A and phytohemaglutinin showed an upward trend in 6 of the 9 patients. Steroid are thought to have a suppressive effect on the immune system but this study suggests that short-course MDMP may not be hazardous to the immune system.     

特发性血小板减少性紫癜患者血小板抗体及淋巴细胞亚群的研究

目的探讨血小板相关抗体(PAIgG)和T淋巴细胞亚群的变化,在特发性血小板减少性紫癜(ITP)免疫发病机制中的作用、临床意义。方法采用间接免疫荧光法测定30例ITP患者及20例正常对照组的PAIgG,20例ITP患儿治疗后复查PAIgG。同时采用流式细胞仪直接免疫荧光法检测外周T血淋巴细胞亚群。结果ITP组PAIgG阳性率为80%,正常对照组为20%(P<0.001),ITP组PAIgG明显高于正常对照组(P<0.001),20例ITP患儿治疗后复查PAIgG,其数值明显下降,差异有显著性(P<0.001)。T淋巴细胞亚群中,ITP组CD3、CD4、CD4/CD8显著低于正常对照组(P<0.01),CD8则显著高于正常对照组(P<0.01)。结论抗血小板相关抗体对提高ITP的诊断、疗效及预后的判断有一定的实用价值,T淋巴细胞亚群的变化能较好的反映ITP的病理机制。     

Immunological profile in children with minimal change nephrotic syndrome

Peripheral blood from patients with active stage of minimal change nephrotic syndrome (MCNS) was examined for concanavalin A (ConA)-inducible suppressor T cell activity, proliferative response to phytohemagglutinin (PHA) and in the autologous (AMLR) and allogeneic (MLR) mixed lymphocyte reaction, proportions of T cells with receptors for IgM (Tu) or IgG (T gamma) and the levels of serum immunoglobulin M, G and A. Six of 9 patients with MCNS studies showed deficiency of ConA-induced suppressor cell activity. In the AMLR, only one of 9 patients with MCNS demonstrated depressed proliferative response (p less than 0.05). In the allogeneic MLR, T cells from 5 of 9 patients with MCNS demonstrated poor proliferative response when stimulated with normal control non-T cells. Five of 9 patients with MCNS had depressed proliferative response to PHA. The proportion of total T cells, Tu cells and T gamma cells in the patient group were comparable to healthy control group. Serum IgG was significantly decreased in 7 of 11 patients. This study demonstrates multiple immunological abnormalities in patients with MCNS that might play a role in its pathogenesis.     

Immunological Profile in Children with Minimal Change Nephrotic Syndrome

ABSTRACT. Peripheral blood from patients with active stage of minimal change nephrotic syndrome (MCNS) was examined for concanavalin A (ConA)-inducible suppressor T cell activity, proliferative response to phytohemagglutinin (PHA) and in the autologous (AMLR) and allogeneic (MLR) mixed lymphocyte reaction, proportions of T cells with receptors for IgM (Tu) or IgG (Tγ) and the levels of serum immunoglobulin M, G and A. Six of 9 patients with MCNS studies showed deficiency of ConA-induced suppressor cell activity. In the AMLR, only one of 9 patients with MCNS demonstrated depressed proliferative response (p<0.05). In the allogeneic MLR, T cells from 5 of 9 patients with MCNS demonstrated poor proliferative response when stimulated with normal control non-T cells. Five of 9 patients with MCNS had depressed proliferative response to PHA. The proportion of total T cells, Tu cells and Ty cells in the patient group were comparable to healthy control group. Serum IgG was significantly decreased in 7 of 11 patients. This study demonstrates multiple immunological abnormalities in patients with MCNS that might play a role in its pathogenesis.     

静注免疫球蛋白及地塞米松对特发性血小板减少性紫癜患儿免疫功能的影响

探讨大剂量地塞米松 (DEX)及静注免疫球蛋白 (IVIG)治疗 ,对特发性血小板减少性紫癜 (ITP)患儿外周血 T淋巴细胞亚群及免疫球蛋白的影响 ,在以DEX、IVIG治疗 ITP患儿 ,治疗前后各抽血一次 ;以 APAAP法测定 T淋巴细胞亚群 ,以单向琼脂免疫扩散法测定免疫球蛋白。结果表明 1.ITP患儿外周血 CD4+ 降低 ,CD8+增高 CD4+ /CD8+ ,显著降低。单纯 DEX组治疗后 ,CD4+、CD8+均显著降低 ,CD4+ /CD8+升高 ,Ig A、Ig G、Ig M降低 ;单纯 IVIG组治疗后 ,CD8+显著降低 ,CD4+ /CD8+升高 ,Ig G显著升高。2 .单纯 DEX组治疗后 ITP患儿外周血白细胞计数显著高于治疗前 ,单纯 IVIG组与 IVIG加 DEX组治疗前后无显著差异。治疗过程中院内交叉感染率单纯 DEX组为 31.43% ,单纯 IVIG治疗组为 2 5 % ,IVIG加 DEX组为2 8.5 7%。因此 ,本文认为 ITP患儿外周血 T淋巴细胞亚群表达异常 ,IVIG及 DEX治疗均干扰了 ITP患儿机体的免疫状态     

急慢性ITP患儿细胞免疫功能变化及临床意义

目的 　观察ITP患儿细胞免疫功能的变化及临床意义。方法　 采用流式细胞仪单克隆抗体免疫荧光法动态检测 3 5例急慢性ITP患儿的T淋巴细胞亚群、NK细胞。结果 　急慢性组均有CD4+、CD4+ CD8+显著降低 ,CD8+显著升高 ,P <0 0 1;慢性组CD3+、CD4+ CD8+显著低于急性组 ,P <0 0 5 ,NK细胞在慢性组中显著降低P <0 0 1。激素不降低急性组CD4+,丙球升高慢性组的CD4+、NK细胞两者分别能降低急慢性组的CD8+。 结论 　急慢性组均有T细胞亚群比例失调 ,慢性型更为明显 ,CD4+ CD8+、NK细胞对判断病程、预后有一定意义 ,激素不增加感染机会。     

Cell-mediated cytotoxicity in down syndrome: impairment of allogeneic mixed lymphocyte reaction,NK and NK-like activities

Peripheral blood mononuclear cells (PBMC) from 16 non-institutionalized patients with Down syndrome (DS) were studied with various monoclonal antibodies and analysed for natural killer (NK), and NK-like activity. Lymphocyte proliferation and cytotoxic T-lymphocyte (CTL) cytotoxicity generated in mixed lymphocyte culture (MLC) were also evaluated in 11 DS patients. Phenotypic characterization of PBMC from DS subjects confirms our previous findings of high numbers of CD8+ lymphocytes and HNK-1+, and CD16+ cells. Lymphocyte proliferation and CTL cytotoxicity generated in MLC were low or absent in most patients. NK activity was low in almost all DS patients, while NK-like cytotoxicity generated in MLC was normal in the majority and did not correlate with NK activity from unstimulated PBMC.Abbreviations BSA bovine serum albumin - DS Down syndrome - CTL cytotoxic T lymphocytes - FCS fetal calf serum - MLC mixed lymphocyte culture - NK natural killer - PBMC peripheral blood mononuclear cells - PBS phosphate-buffered saline - PHA phytohaemagglutinin     

Analysis of proliferating lymphocyte subpopulations in newborns and adults

Surface markers of peripheral lymphocyte subpopulations were determined before and after in vitro stimulation of neonatal and adult mononuclear cells with phytohaemagglutinin (PHA) and pokeweed mitogen (PWM). The type of cell responding to each mitogen was identified by a method combining autoradiography and the peroxidase-antiperoxidase (PAP) staining method. In comparison with adults, there is a lower proportion of OKT3-positive lymphocytes in neonates (43% vs. 72%). Analysis of lymphocyte subpopulations after stimulating mononuclear cells with PHA and PWM showed that in neonates nearly the same percentages of OKT4-and OKT8-positive proliferating cells (43% and 40%, respectively) could be observed, whereas in the proliferating cells from the adults, the OKT8 surface marker predominated (41% vs. 18%).Abbreviations PHA phytohaemagglutinin - PWM pokeweed mitogen - PAP peroxidase-antiperoxidase - MEM minimal essential medium - FCS fetal calf serum - MIF migration inhibition factor - LIF leukocyte migration inhibition factor - ConA concanavalin A     

Analysis of lymphocyte proliferative response subpopulations in very low birth weight infants and during the first 8 weeks of life

Cell-mediated immunity is not well characterized in very low birth weight infants, and abnormalities may represent a significant vulnerability to infection. This report describes 165 serial studies in 58 infants between 700 and 1300 g birth weight during the first 8 wk of life. Two ml of blood were drawn at 2-wk intervals to measure T cell numbers and subsets and response to phytohemagglutinin (PHA). Overall, lymphocyte proliferation to PHA averaged 17,264 cpm, significantly less than the adult control (23,566 cpm). T cell numbers and subsets were CD3 62% (adult controls 75%), CD4 45% (49%), and CD8 18.6% (27%). Values at birth were lower as all parameters increased for at least the first 4 wk of life: PHA at birth was 15,464 cpm, CD3 48%, CD4 37%, and CD8 13%. Because of the lymphocytosis of premature infants, the absolute numbers of total T cells and subsets were within the normal adult range despite less than 50% of the mononuclear cells at birth being T cells. A study of five infants demonstrated an average of 52% B7+ cells at birth showing that the number of B cells at birth was increased approximately 10-fold over the control number in adults. Clinical correlation showed that the increases in both the % CD8 and the absolute number of CD8+ lymphocytes after birth were correlated with both the occurrence of sepsis and the assessed lymphocyte subsets in a sizeable number of very low birth weight infants serially during the first 8 wk of life including lymphocyte function using isolated mononuclear cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

急性特发性血小板减少性紫癜患儿免疫功能变化及其临床意义

目的检测特发性血小板减少性紫癜(ITP)患儿的免疫功能并探讨ITP的发病机制。方法抗血小板抗体(PAIgG)测定采用放射免疫法,IgA、IgG、IgM测定采用WL-快速免疫消浊比浊法,T淋巴细胞亚群测定采用APAAP法,血清IL-2、sIL-2R和IL-6测定采用ELISA法。结果 ITP患儿的IL-2、IL-6,IgA、IgM、CD8及IL-2R表达均明显升高(P<0.01),CD4+、CD4+/CD8+细胞均明显减少(P<0.01)。结论细胞免疫及体液免疫异常共同参与ITP致病机制,调节淋巴细胞亚群平衡有助于寻找ITP治疗的新途径。     

T-lymphocyte subpopulations in tuberculosis

BACKGROUND: Tuberculosis is associated with both qualitative and quantitative defects in the cell mediated immune response. The changes that occur in the lymphocyte profile in blood in children with tuberculosis are not well understood. DESIGN: Prospective study. SETTING: Referral hospitals. METHODS: Lymphocyte subpopulations were determined by flow cytometry in 17 healthy tuberculin positive children, in 22 children with newly diagnosed pulmonary tuberculosis and in 8 of these children after antituberculosis therapy. RESULTS: Absolute numbers and percentages of CD3+ and CD4+ T cells were reduced in children with tuberculosis, compared to controls. CD4+ counts increased significantly following antituberculosis treatment, compared to baseline values. In contrast, the proportion of T cells expressing the gdT cell receptor was similar in tuberculosis patients and controls. CONCLUSION: Children with tuberculosis have a systemic decrease in the proportion and number of CD3+ and CD4+ T cells which reverses during therapy.     

Elevated common acute lymphoblastic leukemia antigen expression in pediatric immune thrombocytopenic purpura.

Bone marrow examination is often performed in thrombocytopenic children to distinguish immune thrombocytopenic purpura (ITP) from acute leukemia. We describe a patient with thrombocytopenia and 50% common acute lymphoblastic leukemia antigen (CALLA) positivity in his marrow who was subsequently shown to have ITP. CALLA (CD10) is a surface antigen found in early B-lymphocytes and is elevated in most cases of childhood acute lymphoblastic leukemia (ALL). This case prompted us to prospectively study the frequency of immature lymphocyte populations in children with ITP. Fourteen patients with acute ITP and five with other conditions were studied. The two groups were comparable with respect to age: ITP mean, 4.3 (range 0.3-15.5) years; control mean, 5.8 (0.6-13.8) years. The ITP group had a significantly higher percentage of CD10 positive bone marrow lymphocytes (p = 0.007). Five of the 10 patients younger than 4 years of age in the ITP group had CD10 levels of greater than 30%, which is in the leukemic range, whereas none of the control patients had a CD10 levels of greater than 17% (p = 0.003). There was good correlation between CD10 positivity and B4 positivity indicating that both of these markers arise from the same population of immature B-lymphocytes. None of the ITP patients who were older than 4 years had a CD10 level of greater than 30%. We conclude that it is common to have an increase in the proportion of immature lymphocytes in the marrow of young children with ITP. The cause of this increase in CD10 positive cells is unknown.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of short-term administration of human chorionic gonadotropin on immune functions in cryptorchid children

To delineate the effects of human chorionic gonadotropin (hCG) administration on immune responsiveness, immunological parameters including serum immunoglobulins, total and differential white blood cell count T and B lymphocyte membrane phenotype, in vitro, proliferative response to phytohaemagglutinin, Concanavalin A (ConA) and pokeweed mitogen were studied in 13 prepubertal cryptorchid boys before, during, and 3 months after hCG therapy. Before treatment, all the immunological parameters were normal except for an unexpected high percentage of T suppressor-cytotoxic cells (CD8+). During therapy, the absolute number of total peripheral blood lymphocytes, and that of total T-cells, T helper-inducer cells and of CD8+ subsets were diminished. The percentage of CD8+ cells and lymphocyte response to ConA decreased significantly and returned to normal after hCG withdrawal. The possible effects of long-term hCG treatment remain to be determined.     

Immune evaluation of 50 children with neuroblastoma at onset

Several studies document the importance of immunity in the host-tumor relationship in neuroblastoma patients, evidentiating a correlation between clinical compromission and immune impairment. In this study we evaluated some aspects of cellular and humoral immune capacity in 50 neuroblastoma patients at onset. The aim of this investigation was to define, if any, the common immune pattern of these patients, and to evidentiate a possible correlation with prognosis. Immune tests performed were serum immunoglobulin quantitation, absolute value of total T and B lymphocytes, and lymphocyte reactivity to phytohemagglutinin (PHA), Concanavalin A (ConA), and Pokeweed mitogen (PWM). In patients with localized or regional disease, diminished values of lymphocytes were observed in 4/16 cases, a datum highly correlated to a poor final outcome (P <0.01). Mitogen response and serum immunoglobulin levels were frequently altered, but no prognostic value was evidentiated. Thirty-one children with disseminated disease presented more frequent and extended abnormalities: No parameter was found to correlate with prognosis, except for an impaired PHA response that paradoxically assumed a favourable prognostic meaning. Our results suggest that only the total lymphocyte number can contribute to predict the survival ratio in patients with regional disease.     

Nonspecific suppressor cell activity and lymphocyte response to beta-lactoglobulin in cow's milk protein hypersensitivity

beta-Lactoglobulin (BLG) is clinically the most frequent allergen in cow's milk protein hypersensitivity (CMPH). We therefore assessed its in vitro blastogenic effect on lymphocytes of patients suffering from CMPH. Twenty-two patients aged from 1 to 20 months were investigated. beta-Lactoglobulin produced a significantly (p less than 0.00003) higher blastogenic response in lymphocytes of patients with CMPH (mean stimulation index 7.7 +/- 0.7 SEM) than in 26 age-matched controls (2.7 +/- 0.4). This response was age related, being most marked in infants up to 5 months of age. Lymphocyte proliferation following stimulation with BLG can be a useful in vitro test in the diagnosis of CMPH. Because of this augmented lymphocyte response, we considered the possibility of a related defect in immune suppression. Nonspecific concanavalin A-induced suppressor cell activity was assessed in the above patient and control populations. Concanavalin A-induced suppressor cell activity was significantly (p less than 0.05) reduced in patients with CMPH. This too was most marked in the first few months of life. We suggest that the development of CMPH may be due to delayed maturation of a suppressor cell population.     

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目的通过测定Tourette综合征(TS)患儿血清抗链球菌溶血素O(ASO)、T淋巴细胞亚群和自然杀伤细胞(NKC),观察链球菌感染与TS的关系、TS细胞免疫功能状态及免疫调节剂的治疗作用。方法2005-05—2005-10对首都儿研所神经科门诊就诊的58例TS患儿分别采用透射比浊法检测ASO,双色免疫荧光法检测T淋巴细胞亚群及NKC。以同期健康体检儿童为对照组。对细胞免疫异常者随机分为两组,分别予免疫调节剂或免疫调节剂 多巴胺受体阻滞剂治疗。结果(1)TS患儿ASO阳性率较对照组显著增高(P<0.05);(2)TS患儿中,ASO阳性组CD4 显著低于ASO阴性组(P<0.05);TS患儿CD3 、CD4 、CD4 /CD8 显著低于对照组(P均<0.05),NKC显著高于对照组(P<0.05);(3)细胞免疫异常的TS患儿,单纯采用免疫调节剂治疗或免疫调节剂 多巴胺受体阻滞剂治疗,两组间疗效差异无显著性(P>0.05)。结论部分TS患儿发病与链球菌感染有关;TS患儿存在细胞免疫功能改变,表现为T淋巴细胞亚群平衡失调和NKC升高,链球菌感染可能与之有关;单纯免疫调节剂治疗可能对链球菌感染触发的TS有效。     

T cell response to anti-CD3 antibody in Down's syndrome.

The non-specific mitogen phytohaemagglutinin (PHA) and an anti-CD3 (OKT3) monoclonal antibody were used to measure the lymphocyte proliferative response in blood samples from 15 subjects with Down''s syndrome. Blood from 15 healthy controls closely matched for age and sex was also assayed. The mean blastogenic value in PHA stimulated patient lymphocyte cultures was similar to that calculated in the controls. In contrast, the mitogenic response of lymphocytes from patients with Down''s syndrome to anti-CD3 stimulation was on average significantly reduced. Immunofluorescence studies and additional experiments carried out by using semiallogeneic (maternal) monocytes as a source of antigen presenting cells showed that the impaired anti-CD3 induced mitogenesis in Down''s syndrome could not be ascribed either to a lack of binding of the antibody to the trisomic cells, or to a defective monocyte-T cell interaction. These findings help to explain the cellular basis of the immune defect in Down''s syndrome.     

儿童急性ITP Th1/Th2细胞功能状态初步研究

目的探讨急性儿童特发性血小板减少性紫癜(ITP)Th1/Th2细胞功能状态及其在ITP免疫发病机制中的作用。方法采用流式细胞术胞内标记法检测外周血CD4+T细胞IL-4和IFN-γ阳性表达率;逆转录-聚合酶链反应(RT-PCR)及荧光定量PCR检测外周血CD4+T细胞中IFN-γ、IL-4、IL-5、IL-13、T-bet、GATA-3、SOCS-1、SOCS-3和TIM-3等Th相关的细胞因子及转录因子mRNA表达。结果①急性ITP患儿Th1细胞阳性率明显低于正常同年龄对照组(3.36%±1.25%vs12.71%±2.29%,P<0.01),Th2细胞比例明显升高(2.63%±1.35%vs0.46%±0.17%,P<0.01),Th1/Th2比值显著降低(1.45±0.57vs34.13±5.76,P<0.01);②急性ITP患儿GATA-3、SOCS-3mRNA表达明显增高,T-bet及SOCS-5表达与同年龄对照组无显著差异(P>0.05),TIM-3表达明显增高(P<0.01);③急性ITP患儿CD4+T细胞高表达Th2类细胞因子IL-4、IL-5、IL-13(P<0.01),Th1类细胞因子IFN-γ与正常对照比较无显著性差异(P>0.05)。结论急性ITP患儿Th2细胞过度活化,可能与ITP免疫功能紊乱有关。     

儿童免疫相关性疾病的常规免疫学指标研究

目的比较特发性血小板减少性紫癜(ITP)、过敏性紫癜(HSP)及皮肤黏膜淋巴结综合征(KD)的常规免疫学指标变化及静脉注射免疫球蛋白(IVIG)的应用。方法用流式细胞仪测T细胞亚群,免疫比浊法测免疫球蛋白(Ig)及补体。常规加IVIG治疗。结果ITP患儿CD3 CD4 T细胞降低并IgG升高;HSP患儿IgA升高伴CD3 T细胞增高。KD患儿CD3 CD8 T细胞降低。IVIG可明显改善3种疾病临床症状,尤其有助KD患儿冠状动脉损害发生率减少。结论T细胞亚群、lg 应作为ITP、HSP及KD的常规检查项目。早期应用IVIG对改善预后及缩短病程有重要意义。     

卡介苗体外对急性淋巴细胞性白血病患儿细胞毒性T淋巴细胞杀伤HL-60细胞效应的影响

目的：探讨卡介苗（BCG）体外对急性淋巴细胞性白血病(ALL)患儿外周血细胞毒性T淋巴细胞（CTL）杀伤HL-60细胞效应的影响。方法：应用Ficoll-Hypaque法分离ALL患儿（白血病组）和健康儿童（对照组）的外周血单个核细胞,在白介素-2（IL-2）、植物血凝素（PHA）和BCG作用下诱导成CTL细胞;应用流式细胞仪测定CD3、CD3+CD4+、CD3+CD8+比例变化;应用MTT法检测CTL对HL-60细胞的杀伤力。结果：培养前2 d,对照组和白血病组CTL细胞数量及体积均无明显变化,第3天开始两组细胞均开始增殖,体积变大,6～10 d达高峰;培养至第10天时,加入BCG的白血病组和对照组细胞数量分别较无BCG的白血病组和对照组增多;白血病组CD3+CD8+比例明显高于对照组,经BCG作用后两组CD3+CD8+比例均明显升高;白血病组CTL对HL-60细胞的杀伤力明显低于对照组。结论：BCG可协同IL-2、PHA体外促进CTL细胞增殖并提高其对HL-60细胞的杀伤力。