Human milk contains large amounts of free oligosaccharides (HMOs). HMOs have been shown to exert antiinflammatory properties, and evidence for their immunomodulatory effects is increasing. The purpose of this study was to evaluate influences of two human breast milk-derived oligosaccharide samples (neutral and acidic oligosaccharides), and of a low-molecularweight fucoidan on cytokine production and activation of cord blood mononuclear cells. Cord blood mononuclear cells from randomly chosen healthy newborns were co-cultured with the oligosaccharide samples. By means of flow cytometry, intracellular cytokine production (d 20) and surface marker expression of T cells (d 5) were measured. In vitro-induced Ig levels were quantified nephelometrically (total IgG1) and by ELISA (total IgE) in the supernatant of cell cultures. The acidic oligosaccharide fraction increased the percentage of interferon-␥ producing CD3ϩCD4ϩ and CD3ϩCD8ϩ cells (p Ͻ 0.05) and the IL-13 production in CD3ϩCD8ϩ cells (p Ͻ 0.05). In acidic oligosaccharide cultures, CD25ϩ expression on CD3ϩCD4ϩ cells was significantly elevated (p Ͻ 0.05). Low-molecular-weight fucoidan induced IL-4 production in CD3ϩCD4ϩ T cells (p Ͻ 0.05) and IL-13 production in CD3ϩCD8ϩ T cells (p Ͻ 0.05), whereas interferon-␥ production remained unaffected in both T-cell populations. Ig production (total IgE and total IgG1) remained unaffected. Human milk-derived oligosaccharides and plant-derived oligosaccharides affect the cytokine production and activation of cord blood derived T cells in vitro. Therefore, oligosaccharides and, in particular, acidic oligosaccharides may influence lymphocyte maturation in breast-fed newborns. The postnatal period is a crucial time for the maturation of the immune system. At birth, T-lymphocytes exhibit a Th2-profile, characterized by a limited ability to produce cytokines. Throughout the first months after birth these Th2-skewed responses are modified into low-level immunity, predominantly expressing Th1-cytokines and IgG-antibodies, particularly of the IgG1 subclass (1,2).Human milk is a biologic fluid containing large amounts of free oligosaccharides. HMOs represent the third largest solid component (after lactose and lipids) in breast milk, occurring at a concentration of 20 -23 g/L in colostrum and 12-14 g/L in mature milk (3). HMOs are very resistant to enzymatic hydrolysis (4,5), indicating that these oligosaccharides must display essential nonnutritive functions.Throughout the last decade, numerous studies demonstrated the ability of HMOs to inhibit pathogens acting as receptors for microbes (6,7). Only a few reports on direct effects on immune function have been published so far. Human milk oligosaccharide structures like lacto-N-fucopentaose III (LNFPIII) and lacto-N-neotetraose (LNneoT) showed an effect on murine IL-10 production (8). It is further discussed that human milk is involved in the generation of antiinflammatory mediators that suppress Th1-type and inflammatory responses in mice (9). 0031-3998/04/5604-0536 PEDIATRIC RES...

Allergies are increasing, and despite deeper insights into the immunologic basis of these diseases, preventive measures are not yet efficient. As the induction of allergic diseases is often triggered in early childhood, perinatal or prenatal preventive strategies would be beneficial. We investigated the transfer of inhalant and nutritive allergens across the human placenta. For this purpose, the maternal side of a placental cotyledon was perfused in vitro with an allergen-containing medium, and a specific ELISA was used to detect the allergens on the fetal side. Both allergens evaluated, birch pollen major allergen Bet v1 and the milk allergen beta-lactoglobulin, could be shown to cross the placenta. The nutritive allergen beta-lactoglobulin was not only transferred across the placenta in all eight experiments, but was also detectable within the first minutes of perfusion. The peak allergen concentration on the fetal side could be increased by addition of human immunoglobulin. For the inhalant allergen Bet v1, transfer was observed in two of 10 placental experiments, and only if human immunoglobulin was added. A pulsatility wave with a frequency of 30 -35 min suggested an active transfer mechanism. We conclude that allergens are actively and selectively transferred across the placenta. Therefore, controlled maternal allergen exposure might offer new ways to induce tolerance to specific allergens in the fetus. There is increasing evidence that the prevalence of allergic diseases is currently rising at an unprecedented rate (1-3). It is also becoming clear that this rise is largely restricted to developed countries, a phenomenon that is widely believed to be linked to a western lifestyle (4). Eliciting agents, preferentially environmental allergens, are predominantly found in indoor environments. Recently, concern has been raised as to any priming effects of exposure to environmental influences even before birth; that is, through the placenta.Allergen-specific T cell proliferation in cord blood cells has been demonstrated by many groups and for many different allergens (5-15). Inhalant and nutritive allergens in the form of crude extracts [birch pollen (5, 7), house dust mite (8 -10, 15), timothy grass pollen (5, 7), cat fur (5, 7), BLG (5, 7, 13, 16), alpha-casein (13), beta-casein (13), kappa-casein (13), BSA (5-7, 13), and ovalbumin (5, 7, 11, 15, 16)] and in the form of recombinant allergens [Lol p1 (9), Der p1 (9), Bet v1 (14), and Phl p1 (14)] has been studied. A line of evidence now supports the hypothesis that fetal T cells are exposed during gestation to maternally derived allergens. These allergens may be ingested or inhaled by the mother (14). The time at which maternofetal allergen exposure takes place in the course of a pregnancy is less clear. The current knowledge suggests early times during gestation, presumably around wk 20 of gestation (5, 14), Szépfalusi et al. unpublished experiments).The occurrence of prenatal T cell priming and its possible impact on later allergic status has been the subject...

Pollen contact in early infancy may enhance the risk for subsequent pollen allergy. In this study likelihood of a prenatal antigen contact, as a result of inhalation of pollen allergens by the mother, was investigated. Due to the seasonal occurrence of allergens studied, the date of priming can be estimated, and this can supply data about the maturation of the fetal immune system. Proliferative responses of umbilical cord blood mononuclear cells (UCB MNCs) to the recombinant major allergens of birch (rBet v 1) and timothy grass (rPhl p 1) were analyzed throughout the whole year. A positive proliferative response was regarded as the criterion for a prenatal contact of the immune system with the allergen. Prenatal priming with both allergens was observed. Timothy grass pollen displayed considerably higher antigenicity than did birch pollen. The susceptibility of the fetal immune system to be primed by these allergens varies during the gestation period. The majority of positive responses to rPhl p 1 and rBet v 1 were found in UCB samples in which antigen contact (the respective pollen season) took place in the first 6 mo of pregnancy. Our results offer indirect evidence that, shortly after migration of T cell precursors to the epithelial thymus, T cells are mature enough for priming with antigens. No relationship was found between the susceptibility of the fetal immune system to be primed by these allergens and the clinical history of the family concerning type I allergy.

(M.P., M.W.) Human naive CD4 ϩ T helper (Th) and CD8 ϩ cytotoxic (Tc) T cells, which only produce IL-2, may differentiate into Th1/Tc1-or Th2/Tc2-like lymphocytes, characterized by their cytokine production profile. 1␣,25-dihydroxyvitamin D 3 (1␣,25(OH) 2 D 3 ) has been reported to inhibit Th1/Tc1-related, but increase Th2/ Tc2-associated cytokines in T cells from adults. In industrialized countries, vitamin D supplementation for prevention of rickets is initiated within the first days of life and continued throughout the entire first year. Epidemiologic studies suggest an association of vitamin D exposure in newborns with the incidence of allergic diseases in later life. This study addresses the effects of 1␣,25(OH) 2 D 3 on Th1/Tc1 versus Th2/Tc2 differentiation in long term cell cultures of (naive) cord blood T lymphocytes. Our results show that in CD4 ϩ as well as CD8 ϩ cord blood cells, 1␣,25(OH) 2 D 3 inhibits not only IL-12-generated IFN-␥ production, but also suppresses IL-4 and IL Human naive T cells, which produce only IL-2, can differentiate into at least two functionally distinct subsets of memory or effector T cells, which can be distinguished by their cytokine profile (1, 2): CD4 ϩ Th1 cells on one hand producing predominantly IFN-␥, IL-2, and lymphotoxin, and CD4 ϩ Th2 cells on the other hand, which are the main source of IL-4 and IL-5, represent two extremely polarized phenotypes of a continuous spectrum of cytokine-producing T lymphocytes. Similar subpopulations (i.e. Tc1 and Tc2) exist in the CD8 ϩ T cell population (3, 4). Th1-and Th2-associated immune responses have been implicated in the pathogenesis of infectious, allergic, and autoimmune diseases (3).The local cytokine environment plays an important role in the differentiation of naive T cells along the Th1/Tc1 or Th2/Tc2 pathway (5, 6). For example, in vitro priming of naive T cells in the presence of IL-4 generates IL-4-producing Th2 effector cells (7-10). Similarly, IL-12, secreted by monocytes and dendritic cells, induces the capacity to produce IFN-␥ in neonatal T cells (11).Furthermore, the arachidonic acid metabolite prostaglandin E2 (12, 13), and several members of the steroid hormone family, such as progesterone (14, 15), glucocorticoids (16), and 1␣,25-dihydroxyvitamin D 3 (1␣,25(OH) 2 D 3 ) (17) have been reported to influence Th and Tc subset generation. 1␣,25(OH) 2 D 3 is not only required for normal mineral homeostasis but also regulates differentiation, growth, and function of different cell types such as hematopoietic and immune cells as well as a variety of normal and malignant epithelial cells (18 -20). The discovery of the high affinity receptor for 1␣,25(OH) 2 D 3 in monocytes and activated lymphocytes (21) suggested that this steroid acts as an immunoregulatory hormone (22,23). Interference with cytokine production of monocytes and lymphocytes seems to be a key mechanism by which 1␣,25(OH) 2 D 3 interacts with the immune system. Suppression of mitogen-induced proliferation (24) and inhibition of secretion of IL-1, ...

No abstract

Allergen-specific cord blood reactivity may be attributed to low levels of allergens crossing the human placenta and providing the fetus with the necessary stimulus for T cell priming.

Our data suggests that maternal sensitization to allergens is associated with the reduced maternal production of the Th2 antagonist IFN-gamma and elevated production of the Th2 cytokine IL-13 in the offspring.

Raised T-cell proliferation of cord blood mononuclear cells (CBMC) in response to various ingestant and inhalant allergens has been reported in newborns, suggesting a prenatal allergen contact. In general, for in vitro proliferation assays a concentration of 50 x 10(3) or 100 x 10(3) cells/well are used. The aim of this study was to analyze whether cell concentration influences T-cell reactivity in cord blood cells and to study differences of T-cell reactivity triggered by inhalant and ingestant allergens. CBMC from 51 neonates (34 females: 22 with and 29 without a family history of allergy, i.e. FH+ or FH-) were incubated with interleukin-2 (IL-2), beta-lactoglobulin (beta-LG), ovalbumin (OVA), house dust mite allergen Dermatophagoides pteronyssinus (Der p 1), and timothy grass allergen Phleum pratense (Ph1 p 1) for 7 days. The cell concentration ranged from 6.25 x 10(3) to 100 x 10(3) cells/well. Proliferation was assessed by incorporation of [3H]-thymidine and was expressed as counts per minute (c.p.m.). In unstimulated cells, a decreasing cell concentration paralleled a steep drop of background activity. In response to IL-2, a decreasing cell concentration led to a slow decrease of c.p.m. The corresponding mean stimulation indices (SI) were 9, 32, 77, 47, and 21 for 100 x 10(3), 50 x 10(3), 25 x 10(3), 12.5 x 10(3), and 6.25 x 10(3) cells/well, respectively. In addition, the highest number of positive proliferative responses to specific allergens were obscured at lower cell concentrations. For beta-LG, the maximal number of positive responses were obtained between 25 x 10(3) (n = 44) and 12.5 x 10(3) (n = 46) cells/well, for OVA at 25 x 10(3) (n = 3) cells/well, for Der p 1 at 50 x 10(3) (n = 5) cells/well, and for Ph1 p 1 between 25 x 10(3) and 12.5 x 10(3) (n = 5) cells/well. Positive proliferation in at least one of the tested assays was observed in 100% of samples in response to beta-LG, in 22% in response to Ph1 p 1, and in 14% in response to OVA and Der p 1. T-cell reactivity did not differ between samples of newborns with or without a family history of atopy. Therefore, sensitivity of T-cell proliferation measurement is highly influenced by background proliferation of unstimulated cells. Hence, proliferation assays with lower cell numbers unmask T-cell reactivity in response to ingestant and inhalant allergens. We suggest the use of concentrations of 12.5 x 10(3)-50 x 10(3) cells/well in proliferation experiments.