Expression of the T-cell markers CD2 and CD28 in healthy and atopic children during the first 18 months of life

Atopy may be associated with a reduced T-cell function early in life, particularly regarding maturation of Th1 responses. The T-cell surface molecules CD2 and CD28 are involved in important T-cell activation pathways. Stimulation via the CD2 receptor increases the responsiveness to interleukin (IL)-12, which is a potent inducer of Th1 responses, whereas CD28 stimulation is critical for Th2 differentiation. Our aim was to prospectively study the expression of the cell-surface markers CD2 and CD28 on T-cells in relation to development of atopic disease. Children (n = 172) were followed from birth to 18 months and the cumulative history of atopic disease was recorded. Blood samples were obtained at birth and at 18 months, and in a subgroup of 78 infants also at 3, 6 and 12 months. Flow cytometry was used to analyze the T-cell markers CD2 and CD28, the latter also within the subsets of T-helper (CD4+) and T-cytotoxic (CD8+) cells. At 18 months, 31 children had and 118 did not have atopic symptoms. At this age, skin prick test (SPT) positive children with atopic symptoms with or without an atopic family history (AFH) showed a lower expression of CD2 mode fluorescence intensity (FI) as well as a lower proportion of CD2+ cells, as compared with non-sensitized children with neither atopic symptoms nor AFH. This was accompanied by a higher expression of CD28 FI on CD2+CD8+CD28+ cells. No significant differences were seen at time points before 18 months, although the proportion of CD2+ tended to be low also earlier in life. In conclusion, the observed reduced expression of CD2 in atopic infants may support previous findings that atopy is associated with a reduced CD2 function. The high CD28 FI in SPT positive children with atopic symptoms may possibly be a consequence of a TH2-skewed immune system.     

Enhanced production of IL4 but not of IFNλ and IL10 by peripheral blood mononuclear cells from atopic children in response to CD2 plus CD28 stimulation

Peripheral blood mononuclear cells from 16 children with atopic disease (range of IgE levels: 33 - 2892 kU/l) and 12 age matched controls were stimulated either with mAbs specific for CD3, CD2, CD3 plus CD28, CD2 plus CD28, with Tetanus Toxoid, SEA, or PHA plus PMA and their cell proliferation was determined. In addition, their cytokine production (IL2, IL4, ILIO, IFNλ) following selected stimuli was measured. We found that the cells from atopies proliferated significantly better in response to CD2 stimulation than control cells, with no difference in response to CD3 or SEA stimulation. Furthermore, cells from atopies produced significantly higher amounts of IL4 than cells from controls, a difference most pronounced following CD2 plus CD28 stimulation. No differential production was found for IL 10 and IFNλ. We conclude that in atopic children with moderately elevated IgE a hyperreactivity of the CD2 pathway of stimulation and a clear elevation of IL4 but not of IL 10 or IENλ production can be demonstrated.     

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.     

Determination of T‐cell subpopulations and intracellular cytokine production (interleukin‐2, interleukin‐4, and interferon‐γ) by cord blood T‐lymphocytes of neonates from atopic and non‐atopic parents

This report describes the results of a prospective study on immunological markers in cord blood for the prediction of allergic diseases in children. First we evaluated methodological aspects of the flow cytometric technique on cord blood cytokine measurements. Subsequently, the T‐cell subsets and percentage of cytokine‐producing cord blood T‐helper (Th) and T‐suppressor/cytotoxic lymphocytes of neonates from atopic and non‐atopic parents were compared. A group of 33 healthy, full‐term newborn infants of whom 23/33 were at risk for atopy (i.e. having at least one parent with one or more atopic symptoms and positive specific immunoglobulin E [IgE] to at least one common inhalant allergen) was studied. A flow cytometric technique was used to analyze cord blood T‐cell subsets and to determine the percentage of interleukin (IL)‐2‐, IL‐4‐, and interferon‐γ (IFN‐γ)‐producing cord blood Th and T‐suppressor/cytotoxic lymphocytes following stimulation with phorbol 12‐myristate 13‐acetate (PMA) and ionomycin. The percentage of CD3 (T lymphocytes), CD3⁺ CD4⁺ (Th lymphocytes), CD3⁺ CD8⁺ (T‐suppressor/cytotoxic lymphocytes), CD19⁺ (B lymphocytes), CD3⁺ CD4⁺ CD45RO⁺ (memory Th lymphocytes), and CD3⁺ CD4⁺ CD45RA⁺ (naive Th lymphocytes) cells was unrelated to parental atopic status. PMA stimulation augmented the percentage of IL‐2‐ and IFN‐γ‐producing Th and T‐suppressor/cytotoxic lymphocytes, whereas the number of IL‐4‐producing T lymphocytes remained very low or undetectable. No differences in the percentage of IL‐2‐, IL‐4‐ and IFN‐γ‐producing Th and T‐suppressor/cytotoxic lymphocytes were found between neonates from atopic and non‐atopic parents. These results will be re‐evaluated when the atopic status of the children at the age of 1 and 2 years can be assessed.     

Low-dose cyclosporin A microemulsion in children with severe atopic dermatitis: Clinical and immunological effects

Cyclosporin A (CsA) is an effective and well-tolerated treatment for severe childhood atopic dermatitis (AD). By starting at a low dose, the therapeutic safety should be further increased. The aim of this study was to evaluate low-dose CsA in childhood AD with respect to clinical outcome and modulation of T-cell dysregulation. In an open prospective study, 10 children (age: 22–106 months) with severe AD (mean objective SCORAD score > 40 on two baseline measurements at a minimum interval of 2 weeks) were treated with CsA solution for 8 weeks. All patients received a starting dose of 2.5 mg/kg/day, which was increased stepwise in non-responders to a maximum of dose of 5 mg/kg/day. Disease activity was monitored using the SCORAD index. The frequency of cytokine-producing peripheral blood T lymphocytes was analyzed by intracellular cytokine staining, and T-cell numbers were measured by fluorescence-activated cell sorter (FACS) analysis. Twenty healthy age-matched children were included as controls for the immunological data. Nine of the 10 patients had a SCORAD reduction of at least 35%. In seven patients this was achieved with low-dose CsA at 2.5 mg/kg/day (n = 4) and 3.5 mg kg/day (n = 3). Seven of the nine responders experienced no relapse within the 4-week follow-up period. At baseline the percentage of interleukin-4 (IL-4), IL-13, and human leucocyte antigen (HLA)-DR-positive CD3⁺ cells was higher in the patient group than in the controls. After CsA treatment there was a significant reduction in interferon-γ (IFN-γ), IL-2, IL-4, IL-13, and HLA-DR-positive CD3⁺ cells. Hence, in severe pediatric AD, CsA microemulsion, when started at a low dose (2.5 mg/kg/day), improves clinical measures of disease, reduces T-lymphocyte cytokine production, and regulates T-cell activation.     

Proliferative responses of CD4+ T-cell population to ovalbumin in patients with atopic dermatitis who are sensitive to hen eggs

Proliferative responses of peripheral blood mononuclear cells and T cells with monocytes to ovalbumin were significantly higher than those of B cells with monocytes to ovalbumin in patients with atopic dermatitis (AD) who were sensitive to hen eggs. The CD4+ T-cell/CD8+ T-cell ratios (the values obtained by dividing the maximum stimulation index of CD4+ T cells with monocytes to ovalbumin by the maximum stimulation index of CD8+ T cells with monocytes to ovalbumin) were significantly higher in AD patients sensitive to hen eggs than in non-atopic healthy controls. The proliferative responses of CD4+ T cells with monocytes to ovalbumin were more intensive than those of CD8+ T cells with monocytes in patients with AD sensitive to hen eggs compared with non-atopic healthy controls. These results suggest that the cells responding to ovalbumin are predominantly CD4+ T cells. However, there was no relationship between the stimulation index of proliferative responses of peripheral blood mononuclear cells to ovalbumin and RAST scores for hen eggs. Thus it is possible that the majority of the CD4+ T cells which respond to ovalbumin are not CD4+ helper T cells for IgE production.     

Association between asymptomatic infections and linear growth in 18–24‐month‐old Malawian children

Inadequate diet and frequent symptomatic infections are considered major causes of growth stunting in low‐income countries, but interventions targeting these risk factors have achieved limited success. Asymptomatic infections can restrict growth, but little is known about their role in global stunting prevalence. We investigated factors related to length‐for‐age Z‐score (LAZ) at 24 months by constructing an interconnected network of various infections, biomarkers of inflammation (as assessed by alpha‐1‐acid glycoprotein [AGP]), and growth (insulin‐like growth factor 1 [IGF‐1] and collagen X biomarker [CXM]) at 18 months, as well as other children, maternal, and household level factors. Among 604 children, there was a continuous decline in mean LAZ and increased mean length deficit from birth to 24 months. At 18 months of age, the percentage of asymptomatic children who carried each pathogen was: 84.5% enterovirus, 15.5% parechovirus, 7.7% norovirus, 4.6% rhinovirus, 0.6% rotavirus, 69.6% Campylobacter, 53.8% Giardia lamblia, 11.9% malaria parasites, 10.2% Shigella, and 2.7% Cryptosporidium. The mean plasma IGF‐1 concentration was 12.5 ng/ml and 68% of the children had systemic inflammation (plasma AGP concentration >1 g/L). Shigella infection was associated with lower LAZ at 24 months through both direct and indirect pathways, whereas enterovirus, norovirus, Campylobacter, Cryptosporidium, and malaria infections were associated with lower LAZ at 24 months indirectly, predominantly through increased systemic inflammation and reduced plasma IGF‐1 and CXM concentration at 18 months.     

Both allergen-specific CD4 and CD8 Type 2 T cells decreased in asthmatic children with immunotherapy

Allergen-specific immunotherapy (IT) has been used for the treatment of atopic diseases since the turn of this century. The precise working mechanisms, however, remain to be clarified. The aim of this study was to investigate the role of particular subsets of allergen-specific T cells in the non-atopic individuals, untreated asthmatic children and the asthmatic children receiving immunotherapy. We collected peripheral blood from 16 untreated asthmatic children and 17 asthmatic children receiving immunotherapy over one and half years. All the patients were sensitive to mite allergen. Peripheral blood mononuclear cells (PBMC) were isolated and, in vitro , stimulated with crude mite extract to enrich the mite-specific T-cell population. After 14 days, the enriched mite-specific T cells were stimulated with phorbol-12-myristate-13-acetate (PMA) and ionomycin for intracellular detection of cytokines such as IFN-γ, IL-4 in CD4⁺ and CD8⁺ T lymphocytes. The data here demonstrated that the levels of mite-specific IgG4 and IgA increased significantly in asthmatic children after immunotherapy. In addition, both IL-4 expressing CD4⁺ and CD8⁺ T cells were significantly lower in asthmatic children after immunotherapy compared with those of before treatment and the normal control (p < 0.05). In contrast, the frequency of IFN-γ expressing CD4⁺ and CD8⁺ T cells did not significantly differ between untreated and SIT-treated groups. All these data suggested that decreased Type 2 CD4⁺ and CD8⁺ T cells might be closely correlated with the regulatory mechanisms of immunotherapy.     

Collagenous sprue in a 3‐month‐old infant

Collagenous sprue (CS) is a severe malabsorption disorder, the etiology of which has not been well defined. Herein, we report the case of a 3‐month‐old infant with CS who responded to steroid and immunomodulator treatment and presented a thick subepithelial collagen band. A 3‐month‐old Japanese girl presented with severe watery diarrhea that lasted for 2 weeks. She was admitted to the referring hospital, but symptomatic improvement was not achieved with fasting and rehydration. Gastroduodenal endoscopy showed an edematous duodenal mucosal surface. Duodenal biopsy indicated severe villous atrophy with infiltration of mostly CD8‐positive T cells; and deposition of subepithelial collagen was confirmed. The subepithelial collagen deposits, however, had disappeared after treatment. Historically, child‐onset CS is extremely rare and this case is likely to be the youngest case of infantile CS. The present case suggests that CS should be considered as a differential diagnosis for intractable diarrhea, even in infants.     

The C−159T polymorphism in the CD14 promoter is associated with serum total IgE concentration in atopic Chinese children

Activation of macrophages through CD14 by microbes is crucial in inducing immunity by type 1 T helper cells. A C-to-T polymorphism at position −159 of CD14 was associated with serum total IgE level in Caucasians but not in Japanese subjects. The objective of this study is to determine whether this polymorphic marker is associated with atopy and asthma phenotypes in Chinese children. Restriction fragment length polymorphism was used to characterize CD14/−159 genotypes. Microparticle immunoassay was used to measure serum total IgE level; fluorescent enzyme immunoassay was performed to measure serum concentrations of specific IgE to aeroallergens; and enzyme-linked immunosorbent assay was used to measure serum levels of soluble CD14 (sCD14). Lung function in asthmatics was assessed by spirometry. Two hundred and fifty-eight patients and 92 control children were recruited. Their mean serum total IgE concentrations were 331 and 74 kIU/l, respectively (p < 0.0001). Atopy, defined as the presence of at least one allergen-specific IgE in serum, was found in 220 (85%) patients and in 41 (45%) controls (p < 0.0001). Serum sCD14 levels were significantly associated with CD14/−159 genotypes (p = 0.004). Atopic subjects with CC genotype in CD14/−159 had the highest serum total IgE levels compared with CT and TT genotypes, with the respective mean values being 661, 427 and 380 kIU/l (p = 0.015). Similarly, a higher proportion of subjects with CC genotype had increased serum total IgE concentration (p = 0.039). This polymorphic marker was not associated with asthma or aeroallergen sensitization in our cohort. Our results suggest that the C−159T of CD14 was associated with serum total IgE concentration in atopic Chinese children.     

儿童重症化脓性脑膜炎CD3＋ CD8＋ T细胞与炎症指标、体液免疫的变化

目的：通过观察儿童重症化脓性脑膜炎早期血CD3＋CD8＋T细胞的变化,以及与炎症指标、体液免疫指标之间的关系,探讨其在儿童重症化脓性脑膜炎发生发展中的临床意义。方法回顾性分析中国医科大学附属盛京医院PICU 2014年8月1日至2015年12月31日收治的39例1个月~14岁的重症化脓性脑膜炎患儿,血CD3＋CD8＋T细胞计数正常或升高(≥190个/mm3)为A组( n＝22),降低(<190个/mm3)为B组(n＝17),分析患儿的一般资料、血液炎症指标、体液免疫、脑脊液改变在两组患儿中的分布和差异。结果17例(43.6％)患儿CD3＋CD8＋T细胞明显下降;所有4例死亡均为B组患儿;虽然没有统计学差异,但 B 组 Glasgow 昏迷评分<8分者比例(58.8％)高于 A 组(31.8％)。B组C-反应蛋白、降钙素原中位数(最小值-最大值)分别为251.0(26.2-417.0)mg/L、32.7(0.9-100.0)ng/L,远远高于A组的106.5(12.0-458.0)mg/L、4.5(0.1-200.0)ng/L,差异有统计学意义(P<0.05);B组中6例(35.3％)外周血WBC<4×109/L,而 A组为1例(4.6％),中性粒细胞>80％的比例A组为7例(31.8％),而B组为12例(70.6％),两组比较差异有统计学意义(P<0.05)。 B组14例(82.3％)患儿脑脊液中糖含量<2.0 mmol/L,高于A组[11例(50.0％)],两组比较差异有统计学意义(P<0.05)。结论儿童重症化脓性脑膜炎CD3＋CD8＋T细胞可能受到抑制,其与患儿脑功能损伤程度、炎症反应以及预后相关。可能对指导临床免疫制剂的应用有一定帮助。     

Adrenocortical tumor with precocious puberty in a 2‐month‐old girl

Adrenocortical tumor is a rare childhood tumor with a median age at onset of 3.2 years. Virilization is the most common sign. Laparotomy is the reference treatment and has a favorable course. The diagnosis of adrenal tumor can be difficult. The main parameters of malignant tumors are size and metastasis. Analysis of TP53 mutation can facilitate final diagnosis. We report a case of virilizing adrenal tumor that developed in a 2‐month‐old girl, and which was treated with laparoscopic adrenalectomy.     

Munc13‐4 deficiency with CD5 downregulation on activated CD8+ T cells

Familial hemophagocytic lymphohistiocytosis (FHL) is characterized by uncontrolled activation of T cells and macrophages and hypercytokinemia. We have recently described a significant increase in a subpopulation of CD8⁺ T cells with downregulation of CD5 during the acute phase of FHL type2 (FHL2; perforin deficiency), which declines after successful treatment, with a concomitant reduction in serum cytokine level. This unusual subset of CD8⁺ T cells, however, has not been characterized in patients with other subtypes of FHL. Herein, we describe a patient with FHL3 (Munc13‐4 deficiency) carrying compound heterozygous mutations in the UNC13D gene. He had high serum levels of pro‐inflammatory cytokines and significantly increased activated CD8⁺ T cells with downregulation of CD5 during the acute phase, similar to that found in FHL2. This immunophenotypic feature may serve as a useful marker of immune dysregulation in FHL3 in addition to FHL2.     

Ectopic intracranial retinoblastoma in a 3.5‐month‐old infant without eye involvement and without evidence of heritability

Heritable retinoblastoma can rarely be associated with a midline intracranial neuroblastic tumor, referred to as trilateral retinoblastoma. We present an unusual midline brain tumor in an infant that was identified as ectopic retinoblastoma by histopathology, DNA methylation analysis, and molecular genetic detection of biallelic somatic inactivation of the RB1 gene. There was no ocular involvement, and germline mutation was excluded. In this nonresectable tumor, treatment with systemic chemotherapy including high‐dose therapy with autologous stem cell transplantation, but without definite local therapy, resulted in long‐lasting tumor control.