Four-Parameter FluoroSpot Assay Reveals That the Varicella Zoster Virus Elicits a Robust Memory T Cell IL-10 Response throughout Childhood.

During childhood, the composition and function of the T cell compartment undergoes significant changes. In healthy individuals, primary infection with herpesviruses is followed by latency, and occasional subclinical reactivation ensures transmission and contributes to an emerging pool of memory T cells. In immunocompromised individuals, herpesviruses can be life threatening. However, knowledge about the spectrum of virus-specific cytokine responses is limited. Here, we investigated peripheral blood mononuclear cells (PBMCs) from children with differential carrier statuses for cytomegalovirus (CMV), Epstein-Barr virus (EBV), and varicella zoster virus (VZV) (n = 32, age 1 to 17 years). We examined memory T cell subsets as well as IFN-γ-, IL-10-, IL-17A-, and IL-22-producing T cells after polyclonal activation or stimulation with viral peptides using flow cytometry and a 4-parameter FluoroSpot assay. Age and herpesvirus carriage influenced the size of the memory T cell subsets. A positive association between age and the number of IFN-γ-, IL-17A- and IL-22-producing T cells was found following polyclonal activation. For CMV, age was positively associated with IL-17A spot-forming cells (SFC), while for VZV, age was negatively associated with IL-22 and positively associated with IFN-γ SFC. Upon activation with CMV, VZV, and EBV peptides, IFN-γ SFCs dominated. Notably, VZV responses were characterized by a higher IL-10 SFC population compared to both CMV and EBV. Our findings suggest that cytokine responses vary across herpesvirus-type-specific memory T cells and may more adequately reflect their composition. An observed deviation between polyclonal and herpesvirus-specific T cell cytokine responses in children needs to be considered when interpreting the associations between herpesvirus carrier status and bulk T cell reactivity. In summary, these findings may have implications for the treatment of immunocompromised patients. IMPORTANCE Infection with herpesviruses accounts for 35 to 40 billion human cases worldwide. Despite this, little is known about how herpesviruses shape the immune system in the asymptomatic carrier. Particularly in children, primary infection is connected to no or mild symptoms ahead of latency for life. Most research on cellular responses against herpesviruses focuses on inflammatory cytokines associated with antiproliferative and antitumor mechanisms and not the spectrum of cytokine responses in healthy humans. This study investigated four divergent cytokine-producing T cell responses to herpesviruses, reflecting different immunological functions. Three common childhood herpesviruses were selected: Epstein-Barr virus, cytomegalovirus, and varicella-zoster virus. Curiously, not all viruses induced the same pattern of cytokines. Varicella-zoster responses were characterized by IL-10, which is considered regulatory. Besides broadening understanding of responses to herpesviruses, our results raise the possibility that reactivation of varicella-zoster may be counterproductive in cancer treatment through the action of IL-10-producing T-cells.

T Cell Subsets During Early Life and Their Implication in the Treatment of Childhood Acute Lymphoblastic Leukemia.

The immune system plays a major role in recognizing and eliminating malignant cells, and this has been exploited in the development of immunotherapies aimed at either activating or reactivating the anti-tumor activity of a patient's immune system. A wide range of therapeutic approaches involving T lymphocytes, such as programmed cell death protein ligand-1 (PDL-1) inhibitors, cytotoxic T-lymphocyte-associated protein-4 (CTLA-4) blockers, and CD19-targeted T-cell therapy through chimeric antigen receptor (CAR)-T cells or CD19/CD3 bi-specific T-cell engagers, have been introduced to the field of oncology, leading to significant improvements in overall survival of adult cancer patients. During the past few years, the availability and approval of T-cell based immunotherapies have become a reality also for the treatment of childhood cancers. However, the distribution, ratio of regulatory to effector cells and the quality of T-cell responses early in life are distinct from those during adolescence and adulthood, raising the possibility that these differences impact the efficacy of immunotherapy. Herein we provide a brief overview of the properties of conventional T cell subsets during early life. Focusing on the most common cancer type during childhood, acute lymphoblastic leukemia (ALL), we describe how current conventional therapies used against ALL influence the T-cell compartment of small children. We describe early life T-cell responses in relation to immunotherapies engaging T-cell anticancer reactivity and present our opinion that it is not only immaturity of the adaptive immune system, but also the impact of an immunosuppressive environment that may prove disadvantageous in the setting of immunotherapies targeting pediatric cancer cells.

Deficits in the IgG+ memory B-cell recovery after anthracycline treatment is confined to the spleen of rhesus macaques.

OBJECTIVES: Loss of vaccine-induced antibodies (Abs) after chemotherapy against paediatric acute lymphoblastic leukaemia (ALL) is common and often necessitates re-immunisation after cessation of treatment. Even so, some ALL survivors fail to mount or to maintain protective Abs. Germinal centres (GCs) are clusters of proliferating B cells in follicles of secondary lymphoid tissues (SLTs) formed during adaptive immune responses and the origins of long-lived memory B and plasma cells that are the source of Abs. Furthermore, productive GC reactions depend on T follicular helper (TFH) cells. To understand why chemotherapy induces deficits in Ab responses, we examined how SLTs were affected by chemotherapy. METHODS: Rhesus macaques were infused with either three cycles of the anthracycline doxorubicin or saline, followed by immunisation with a de novo and booster antigen. Spleen and lymph nodes were removed, and memory B, bulk T and TFH cells were examined. RESULTS: Despite adequate GC morphology, a diminished memory and IgG+ B-cell population along with diminished total and booster vaccine-specific IgG-producing memory B cells were noted in the spleens of macaques with past doxorubicin exposure compared to the saline-treated controls (P < 0.05). Intact bulk T and TFH cells were found in the SLTs of treated macaques, which displayed higher CD40L upregulation capacity by their splenic CXCR5+ helper T cells (P < 0.01). In contrast to the spleen, the immune cell populations studied were comparable between the lymph nodes of both saline- and doxorubicin-treated macaques. CONCLUSION: Our findings suggest that the splenic memory B-cell subset, compared to its lymph node counterpart, is more severely altered by anthracycline treatment.

The Feasibility of Host Transcriptome Profiling as a Diagnostic Tool for Microbial Etiology in Childhood Cancer Patients with Febrile Neutropenia.

Infection is a common and serious complication of cancer treatment in children that often presents as febrile neutropenia (FN). Gene-expression profiling techniques can reveal transcriptional signatures that discriminate between viral, bacterial and asymptomatic infections in otherwise healthy children. Here, we examined whether gene-expression profiling was feasible in children with FN who were undergoing cancer treatment. The blood transcriptome of the children (n = 63) was investigated at time of FN diagnosed as viral, bacterial, co-infection or unknown etiology, respectively, and compared to control samples derived from 12 of the patients following the FN episode. RNA sequencing was successful in 43 (68%) of the FN episodes. Only two genes were significantly differentially expressed in the bacterial versus the control group. Significantly up-regulated genes in patients with the other three etiologies versus the control group were enriched with cellular processes related to proliferation and cellular stress response, with no clear enrichment with innate responses to pathogens. Among the significantly down-regulated genes, a few clustered into pathways connected to responses to infection. In the present study of children during cancer treatment, the blood transcriptome was not suitable for determining the etiology of FN because of too few circulating immune cells for reliable gene expression analysis.

Altered proportions of circulating CXCR5+ helper T cells do not dampen influenza vaccine responses in children with rheumatic disease.

Biological therapy options for the treatment of rheumatic disease target molecules that can affect the cross-talk between innate and adaptive immune responses upon vaccination. Influenza vaccination in children with rheumatic disease has been recommended, but there are only sparse data on the quality of vaccine responses from pediatric patients treated with biological therapy. We conducted an influenza vaccine study over 3 consecutive seasons where the antibody response to TIV was evaluated in children with PRD (n = 78), including both non-treated (n = 17) and treated (with methotrexate, TNF-inhibitors with or without methotrexate, or IL-inhibitors, n = 61) children as well as healthy age-matched controls (n = 24). Peripheral B cells, T and NK cell populations, as well as CXCR5+ (follicular) helper T cells (TFH) and chemokines involved in antibody responses were assessed prior to immunization in the same cohort. Data on disease duration, therapy and data on previous influenza vaccinations were retrieved. The proportion of circulating TFH cells were significantly lower in non-treated children with PRD compared to treated patients and healthy controls. The significantly lower proportion of TFH cells was mirrored by a marked significant increase in CXCL13 serum level, the ligand for CXCR5, with higher levels in non-treated children with PRD compared to treated patients and healthy controls. However, the proportion of TFH cells or CXCL13 level at the time of vaccination was not a predictor of the antibody response to TIV in this cohort of children. Children with PRD had an overall similar response to TIV as healthy children. Although not significant, children treated with TNF-inhibitors differed as a few children remained seronegative towards H3N2- and influenza B viruses after immunization. Our data show that children with PRD respond to TIV as healthy children. Furthermore, plasma CXCL13 levels did not correlate to the proportion of TFH cells in blood prior to immunisation, or to antibody responses following immunization.

Insights into defective serological memory after acute lymphoblastic leukaemia treatment: The role of the plasma cell survival niche, memory B-cells and gut microbiota in vaccine responses.

Acute lymphoblastic leukaemia (ALL) is the most common type of cancer in children, accounting for approximately 25% of childhood cancer cases. As a result of effective treatments over the past decades, paediatric ALL mortality has been greatly reduced. Chemotherapy, however, has a range of harmful side effects including the loss of protective antibodies against vaccine-preventable diseases. Since ALL survivors have an increased risk of health problems including organ insufficiencies, acquired vaccine-preventable infections subsequent to clinical remission could become life threatening to these individuals. This review will summarize clinical findings regarding defective humoral immunity in ALL survivors, identify current knowledge gaps and highlight mechanisms related to deficiencies in the B-cell compartment important for serological memory. Further, we illuminate the emerging evidence for a relationship between chemotherapy and gut microbiota, which could play an important role in vaccine responses and the shaping of a young immune system subjected to maturation and recovery.

Assessment of Cytokine and Chemokine Signatures as Potential Biomarkers of Childhood Community-acquired Pneumonia Severity: A Nested Cohort Study in India.

BACKGROUND: Pediatric community-acquired pneumonia (CAP) is a leading cause of childhood mortality in developing countries. In resource-poor settings, pneumonia diagnosis is commonly made clinically, based on World Health Organization guidelines, where breathing difficulty or cough and age-adjusted tachypnea suffice to establish diagnosis. Also, the severity of CAP is generally based on clinical features and existing biomarkers do not reliably correlate to either clinical severity or outcome. Here, we asked whether systemic immune and inflammatory mediators could act as biomarkers predicting CAP severity or outcome. METHODS: Serum from a subset of a CAP cohort (n = 196), enrolled in India, classified according to World Health Organization criteria as having pneumonia or severe pneumonia, was used for simultaneous measurement of 21 systemic cytokines and chemokines. RESULTS: We found significantly higher IL-6, IL-8, IL-13, IFN-γ and lower CCL22 concentrations in patients with severe compared with mild CAP (P values: 0.019, 0.036, 0.006, 0.016 and 0.003, respectively). Based on higher MIP-1α, IL-8, IL-17 or lower CCL22 response pattern at the time of enrolment, children with fatal outcome showed markedly different pattern of inflammatory response compared with children classified with the same disease severity, but with nonfatal outcome (P values: 0.043, 0.017, 0.008 and 0.020, respectively). CONCLUSIONS: Our results suggest a relation between an elevated mixed cytokine response and CAP severity on one hand, and a bias toward uncontrolled neutrophilic inflammation in subjects with fatal outcome on the other. Collectively our findings contribute to increased knowledge on new biomarkers that can potentially predict severity and outcome of childhood CAP in the future.

Epstein-Barr Virus, but Not Cytomegalovirus, Latency Accelerates the Decay of Childhood Measles and Rubella Vaccine Responses-A 10-Year Follow-up of a Swedish Birth Cohort.

BACKGROUND: Epstein-Barr virus (EBV) and cytomegalovirus (CMV) are ubiquitous and persistent herpesviruses commonly acquired during childhood. Both viruses have a significant impact on the immune system, especially through mediating the establishment of cellular immunity, which keeps these viruses under control for life. Far less is known about how these viruses influence B-cell responses. OBJECTIVES: To evaluate the impact of latent EBV and CMV infection on rubella- and measles-specific antibody responses as well as on the B-cell compartment in a prospective birth cohort followed during the first 10 years of life. METHODS: IgG titers against rubella and measles vaccines were measured in plasma obtained from the same donors at 2, 5, and 10 years of age. Peripheral B-cell subsets were evaluated ex vivo at 2 and 5 years of age. Factors related to optimal B-cell responses including IL-21 and CXCL13 levels in plasma were measured at all-time points. RESULTS: EBV carriage in the absence of CMV associated with an accelerated decline of rubella and measles-specific IgG levels (p = 0.003 and p = 0.019, respectively, linear mixed model analysis), while CMV carriage in the absence of EBV associated with delayed IgG decay over time for rubella (p = 0.034). At 5 years of age, EBV but not CMV latency associated with a lower percentage of plasmablasts, but higher IL-21 levels in the circulation. CONCLUSION: Our findings suggest that EBV carriage in the absence of CMV influences the B-cell compartment and the dynamics of antibody responses over time during steady state in the otherwise healthy host.

Associations between EBV and CMV Seropositivity, Early Exposures, and Gut Microbiota in a Prospective Birth Cohort: A 10-Year Follow-up.

Early-life infections with persistent Epstein-Barr virus (EBV) and cytomegalovirus (CMV) are delayed in affluent countries, probably due to alterations in early environmental exposures, such as maternal age, siblings, and day-care attendance. We have previously reported that the timing of EBV and CMV contraction is related both to allergic sensitization and changes in functional competence of immune cells, while the presence/absence of lactobacilli [Lactobacillus (L.) casei, L. paracasei, and L. rhamnosus] or Staphylococcus (S.) aureus in feces is related to the risk for allergy. Here, we used the same prospective longitudinal birth cohort of children to investigate early-life environmental exposures and their influence on EBV and CMV contraction over time. Since gut microbes also belong to this category of early exposures, we investigated their association with herpesvirus contraction. Our results show that these two viruses are acquired with different kinetics and that EBV and CMV seroprevalence at 10 years of age was 47 and 57%, respectively. We also observed that a delayed EBV or CMV infection was associated with older maternal age [time ratio (TR) 1.14, 95% confidence interval (CI) 1.07-1.21, P adj < 0.001 and TR 1.09, CI 1.03-1.16, P adj = 0.008, respectively]. Further, we present the novel finding that S. aureus colonization reduced the time to CMV acquisition (TR 0.21, CI 0.06-0.78, P adj = 0.02). Together, these findings suggest that there is a relationship between timing of herpesvirus acquisition and early-life immune modulating exposures, which interestingly also includes the early infant gut microbiota.

Etiology of community acquired pneumonia among children in India: prospective, cohort study.

BACKGROUND: Childhood community acquired pneumonia (CAP) is a significant problem in developing countries, and confirmation of microbial etiology is important for individual, as well as public health. However, there is paucity of data from a large cohort, examining multiple biological specimens for diverse pathogens (bacteria and viruses). The Community Acquired Pneumonia Etiology Study (CAPES) was designed to address this knowledge gap. METHODS: We enrolled children with CAP (based on WHO IMCI criteria of tachypnea with cough or breathing difficulty) over 24 consecutive months, and recorded presenting symptoms, risk factors, clinical signs, and chest radiography. We performed blood and nasopharyngeal aspirate (NPA) bacterial cultures, and serology (Mycoplasma pneumoniae, Chlamydophila pneumoniae). We also performed multiplex PCR for 25 bacterial/viral species in a subgroup representing 20% of the cohort. Children requiring endotracheal intubation underwent culture and PCR of bronchoalveolar lavage (BAL) specimens. FINDINGS: We enrolled 2345 children. NPA and blood cultures yielded bacteria in only 322 (13.7%) and 49 (2.1%) children respectively. In NPA, Streptococcus pneumoniae (79.1%) predominated, followed by Haemophilus influenzae (9.6%) and Staphylococcus aureus (6.8%). In blood, S. aureus (30.6%) dominated, followed by S. pneumoniae (20.4%) and Klebsiella pneumoniae (12.2%). M. pneumoniae and C. pneumoniae serology were positive in 4.3% and 1.1% respectively. Multiplex PCR in 428 NPA specimens identified organisms in 422 (98.6%); of these 352 (82.2%) had multiple organisms and only 70 (16.4%) had a single organism viz. S. pneumoniae: 35 (50%), Cytomegalovirus (CMV): 13 (18.6%), Respiratory Syncytial Virus (RSV): 9 (12.9%), other viruses: 6 (8.7%), S. aureus: 5 (7.1%), and H. influenzae: 2 (2.9%). BAL PCR (n = 30) identified single pathogens in 10 (S. pneumoniae-3, CMV-3, S. aureus-2, H. influenzae-2) and multiple pathogens in 18 children. There were 108 (4.6%) deaths. The pattern of pathogens identified did not correlate with pneumonia severity or mortality. CONCLUSIONS: The majority of children with CAP have multiple pathogens (bacteria and viruses). S. pneumoniae and S. aureus predominate in NPA and blood respectively. CMV and RSV were the dominant respiratory viruses in NPA and BAL. The presence of multiple pathogens, especially organisms associated with nasopharyngeal carriage, precludes confirmation of a causal relationship in most cases.

Pre-eclampsia affects cord blood NK cell expression of activation receptors and serum cytokine levels but not CB monocyte characteristics.

PROBLEM: Maternal immunopathology in pre-eclampsia is well studied; however, less is known regarding the immunological effects on the newborns. Increased inflammation and activation of immune cells at the fetal-maternal interface in pre-eclampsia could influence the neonatal immune compartment. METHOD OF STUDY: Monocytes and natural killer (NK) cells from cord blood (CB) of children with pre-eclamptic or healthy mothers were analyzed by flow cytometry for surface markers and intracellular cytokines. In addition, serum cytokine profiles were investigated using ELISA or cytometric bead array. RESULTS: Neonates born to pre-eclamptic mothers had an inflammatory serum cytokine profile. While CB monocyte characteristics seemed unaffected, CB NK cells from pre-eclamptic pregnancies had higher NKp30, but borderline lower NKG2D expression. CONCLUSION: In utero inflammatory priming of neonatal innate immunity taking place in pre-eclamptic pregnancies might influence specific NK cell functions in newborns.

A glycosylation-dependent CD45RB epitope defines previously unacknowledged CD27⁻IgM(high) B cell subpopulations enriched in young children and after hematopoietic stem cell transplantation.

The immune system is dysfunctional for years after hematopoietic stem cell transplantation (HSCT). A potential cause is an intrinsic B cell deficiency. In a cohort of pediatric HSCT patients few CD27(+) B cells formed after transplantation with the number of CD27(+)IgM(high) cells more affected than class-switched ones. A previously unacknowledged population of CD27(-)IgM(high) cells made up the majority of B cells and this population was also enlarged in healthy children compared to adults. Only a minority of these CD27(-)IgM(high) B cells expressed markers typical for transitional B cells, and the non-transitional CD27(-)IgM(high) cells could be further divided into subpopulations based on their ability to extrude the dye Rhodamine 123 and their expression of CD45RB(MEM55), a glycosylation-dependent epitope. Thus, we define several novel human CD27(-)IgM(high) B cell subpopulations in blood, all of which are present in higher frequencies and numbers in young children and after HSCT than in adults.

Epstein-Barr virus coinfection in children boosts cytomegalovirus-induced differentiation of natural killer cells.

During childhood, infections with cytomegalovirus (CMV) and Epstein-Barr virus (EBV) can occur in close temporal proximity. Active, as well as latent, CMV infection is associated with enlarged subsets of differentiated natural killer (NK) and cytotoxic T cells. How EBV infection may influence CMV-driven immune differentiation is not known. We found that EBV coinfection selectively influenced the NK cell compartment of CMV-seropositive (CMV(+)) children. Coinfected children had significantly higher proportions of peripheral-blood NKG2C(+) NK cells than CMV(+) EBV(-) children. Ex vivo NK cell degranulation after target cell stimulation and plasma IL-15 levels were significantly higher in CMV(+) children. EBV coinfection was related to the highest levels of plasma interleukin-15 (IL-15) and IL-12p70. Remarkably, in vitro EBV infection of peripheral blood mononuclear cells (PBMC) from EBV(-) CMV(+) children increased NKG2C(+) NK cell proportions. A similar tendency was seen in cocultures of PBMC with EBV(+) lymphoblastoid B-cell lines (LCL) and IL-15. After K562 challenge, NKG2C(+) NK cells excelled in regard to degranulation and production of gamma interferon, regardless of whether there was previous coculture with LCL. Taken together, our data suggest that dual latency with these herpesviruses during childhood could contribute to an in vivo environment supporting differentiation and maintenance of distinct NK cell populations. This viral imprint may affect subsequent immune responses through altered distributions of effector cells.

Cytomegalovirus-seropositive children show inhibition of in vitro EBV infection that is associated with CD8+CD57+ T cell enrichment and IFN-γ.

EBV, a human herpesvirus, is commonly acquired during childhood and persists latently in B cells. EBV seropositivity has been connected to immunomodulatory effects such as altered T and NK cell functional responses as well as protection against early IgE sensitization; however, owing to the asymptomatic presentation during childhood little is known regarding the infection process in children of different ages. In this study, we used mononuclear cells from cord blood and from 2- and 5-y-old EBV-naive children for in vitro EBV infection. We show that the degree of EBV-induced B cell activation and expansion differs between age groups and in particular in relationship to IFN-γ production capacity. EBV infection induced redistribution between B cell subsets with enrichment of IgD(+)CD27(+) cells (commonly referred to as non-switched memory) in infected cord blood cell cultures, and of IgD(-)CD27(+) cells (switched memory) in cell cultures from older children. We also related results to serostatus to CMV, a persistent herpesvirus that can affect differentiation status of T and NK cells. As compared with CMV(-) children, the EBV-induced enrichment of IgD(-)CD27(+) B cells was significantly reduced in infected cell cultures from CMV(+) children. This effect was associated with high levels of IFN-γ and frequencies of highly mature CD8(+)CD57(+) T cells in CMV(+) children. Our results demonstrate that both a child's age and serostatus to CMV will have an impact on EBV-induced B cell activation and expansion, and they point to the ability of viruses with immunomodulatory functions, such as CMV, to affect immune responses within the host system.

Early-life gut bacteria associate with IL-4-, IL-10- and IFN-γ production at two years of age.

Microbial exposure early in life influences immune maturation and potentially also the development of immune-mediated disease. Here we studied early-life gut colonization in relation to cytokine responses at two years of age. Fecal samples were collected from infants during the first two months of life. DNA was extracted from the fecal samples and Bifidobacterium (B.) adolescentis, B. breve, B. bifidum, a group of lactobacilli (L. casei, L. paracasei and L. rhamnosus) as well as Staphylococcus (S.) aureus were detected with real time PCR. Peripheral mononuclear cells were stimulated with phytohaemagglutinin (PHA) and numbers of IL-4-, IL-10- and IFN-γ secreting cells were evaluated using ELISpot. We further stimulated peripheral blood mononuclear cells with bacterial supernatants in vitro and assessed the IL-4-, IL-10- and IFN-γ inducing capacity by flow cytometry and ELISA. Early S. aureus colonization associated with higher numbers of IL-4- (p = 0.022) and IL-10 (p = 0.016) producing cells at two years of age. In contrast to colonization with S. aureus alone, co-colonization with lactobacilli associated with suppression of IL-4- (p = 0.004), IL-10- (p = 0.004) and IFN-γ (p = 0.034) secreting cells. In vitro stimulations of mononuclear cells with bacterial supernatants supported a suppressive role of L. rhamnosus GG on S. aureus-induced cytokine responses. We demonstrate that the early gut colonization pattern associates with the PHA-induced cytokine profile at two years of age and our in vitro findings support that specific bacterial species influence the T helper cell subsets. This suggests that dysbiosis in the early microbiota may modulate the risk of developing inflammatory conditions like allergy.

Age at Epstein-Barr virus infection and Epstein-Barr virus nuclear antigen-1 antibodies in Swedish children.

There is strong evidence supporting a role for Epstein-Barr virus (EBV) in the etiopathogenesis of multiple sclerosis (MS). Because of the strong association between antibody (Ab) titer against EBV nuclear antigen 1 (EBNA1) and late age at EBV infection, manifested as infectious mononucleosis (IM), and MS risk, we sought to determine whether age at primary EBV infection was associated with subsequent anti-EBNA1 Ab titer in a longitudinal study of Swedish children with EBV seropositivity and anti-EBNA1 Ab titers measured at ages 2, 5, and 10.

Cord blood monocyte subsets are similar to adult and show potent peptidoglycan-stimulated cytokine responses.

Human monocytes can be divided into two major subpopulations, CD14(++) CD16(-) and CD14(+) CD16(+) cells, which are suggested to play different roles in antimicrobial responses. In neonates, characteristics and functional responses of monocyte subsets have not previously been explored, and might contribute to the qualitative difference between neonatal and adult cytokine profiles. We report that at baseline, monocyte subsets in cord blood and adult peripheral blood are present in similar frequencies, and show similar expression of CD11c, CD80/CD86, CD163 and HLA-DR. In response to the bacterial ligand peptidoglycan, cord blood monocytes had high inherent capacity for production of the early-response cytokines with levels of tumour necrosis factor and interleukin-12p70 exceeding adult levels, and also a higher phosphorylation of p38-mitogen-activated protein kinase. The CD14(+) CD16(+) cells expressed more interleukin-12p70 than CD14(++) CD16(-) cells and were present in a higher frequency in peptidoglycan-stimulated cord blood mononuclear cell cultures. Together, the behaviour of cord blood CD14(+) CD16(+) cells following peptidoglycan stimulation might indicate a qualitative difference between the neonatal antimicrobial response and that of the adult. In addition we found that serum factors in cord blood and adult sera affected cytokine production similarly, with the exception of tumour necrosis factor, regardless of the source of serum or cells. Overall, our data provide new insights into monocyte heterogeneity in cord blood and monocyte subset responses to a bacterial ligand at birth.

Early-life EBV infection protects against persistent IgE sensitization.

BACKGROUND: Infection with EBV has previously been implicated in influencing allergic disorders, but its precise role remains contradictory. The timing of primary infection may contribute to the discrepancies. OBJECTIVE: This study aimed at investigating whether the time-point of primary EBV infection during childhood could be of importance in modulating the risk of developing IgE sensitization. METHODS: A total of 219 Swedish infants were followed prospectively to 5 years of age with clinical examinations, skin prick testing, specific IgE analyses, and determination of serostatus against EBV. RESULTS: After analysis of the children's EBV serostatus, we found that 5-year-olds who were infected with EBV before the age of 2 years were at a significantly lower risk of being persistently IgE-sensitized-that is, sensitized at both 2 and 5 years of age (adjusted odds ratio, 0.34; 95% CI, 0.12-0.94). In contrast, contraction of EBV after 2 years of age was highly associated with late-onset IgE sensitization (adjusted odds ratio, 4.64; 95% CI, 1.57-13.69). Persistently sensitized 5-year-olds had higher specific-IgE levels than children with late-onset IgE sensitization (P < .01). CONCLUSION: Our data support the value of early-life microbial exposure for protection against the development of IgE sensitization and underscore the proximate postnatal years as an important period during which EBV could contribute to an allergo-protective immune profile.

Herpesvirus seropositivity in childhood associates with decreased monocyte-induced NK cell IFN-gamma production.

EBV infection is inversely associated with IgE sensitization in children, and this association is further enhanced by CMV coinfection. In mice, herpesvirus latency causes systemic innate activation and protection from bacterial coinfection, implying the importance of herpesviruses in skewing immune responses during latent infection. Early control of viral infections depends on IFN-gamma release by NK cells, which generally requires the presence of accessory cells. We investigated IFN-gamma production by NK cells in PBMCs from children seropositive (SP) for EBV alone, for both EBV and CMV, or seronegative for both viruses. The ability of classical (CD14(++)CD16(-)) and proinflammatory (CD14(+)CD16(+)) monocytes to induce autologous NK cell IFN-gamma was studied by coculture experiments with enriched CD3(-)CD56(+) cells. Transwell experiments were used to evaluate how monocytes interact with NK cells to induce IFN-gamma synthesis. SP children had a significantly reduced proportion of IFN-gamma(+) NK cells and cognate intracellular IFN-gamma levels, which was more pronounced in CMV-coinfected subjects. Also, resting PBMCs of SP children displayed lower proportions of proinflammatory monocytes. IFN-gamma production by NK cells was dependent on interactions with monocytes, with the proinflammatory subset inducing the highest IFN-gamma. Finally, SP children had markedly lower levels of plasma IFN-gamma, concurrent with in vitro findings. Herpesvirus infections could be one contributing factor for maturation toward balanced Th1-Th2 responses. Our data indicate that early infection by herpesviruses may affect NK cell and monocyte interactions and thereby also influence the development of allergies.