Paternal Tobacco Smoke Correlated to Offspring Asthma and Prenatal Epigenetic Programming.

Rationale: Little is known about effects of paternal tobacco smoke (PTS) on the offspring's asthma and its prenatal epigenetic programming. Objective: To investigate whether PTS exposure was associated with the offspring's asthma and correlated to epigenetic CG methylation of potential tobacco-related immune genes: LMO2, GSTM1 or/and IL-10 genes. Measurements and Main Results: In a birth cohort of 1,629 newborns, we measured exposure rates of PTS (23%) and maternal tobacco smoke (MTS, 0.2%), cord blood DNA methylation, infant respiratory tract infection, childhood DNA methylation, and childhood allergic diseases. Infants with prenatal PTS exposure had a significantly higher risk of asthma by the age of 6 than those without (p = 0.026). The PTS exposure doses at 0, <20, and ≧20 cigarettes per day were significantly associated with the trend of childhood asthma and the increase of LMO2-E148 (p = 0.006), and IL10_P325 (p = 0.008) CG methylation. The combination of higher CG methylation levels of LMO2_E148, IL10_P325, and GSTM1_P266 corresponded to the highest risk of asthma by 43.48%, compared to other combinations (16.67-23.08%) in the 3-way multi-factor dimensionality reduction (MDR) analysis. The LMO2_P794 and GSTM1_P266 CG methylation levels at age 0 were significantly correlated to those at age of 6. Conclusions: Prenatal PTS exposure increases CG methylation contents of immune genes, such as LMO2 and IL-10, which significantly retained from newborn stage to 6 years of age and correlated to development of childhood asthma. Modulation of the LMO2 and IL-10 CG methylation and/or their gene expression may provide a regimen for early prevention of PTS-associated childhood asthma. Descriptor number: 1.10 Asthma Mediators. Scientific Knowledge on the Subject: It has been better known that maternal tobacco smoke (MTS) has an impact on the offspring's asthma via epigenetic modification. Little is known about effects of paternal tobacco smoke (PTS) on the offspring's asthma and its prenatal epigenetic programming. What This Study Adds to the Field: Prenatal tobacco smoke (PTS) can program epigenetic modifications in certain genes, such as LMO2 and IL-10, and that these modifications are correlated to childhood asthma development. The higher the PTS exposure dose the higher the CG methylation levels are found. The combination of higher CG methylation levels of LMO2_E148, IL10_P325 and GSTM1_P266 corresponded to the highest risk of asthma. Measuring the DNA methylation levels of certain genes might help to predict high-risk populations for childhood asthma and provide a potential target to prevent the development of childhood asthma.

Prevalence of infant sneezing without colds and prediction of childhood allergy diseases in a prospective cohort study.

BACKGROUND: Allergy sensitization may begin during the perinatal period, but predicting allergic diseases in infancy remains difficult. This study attempted to identify early predictors of childhood allergy diseases in a prospective cohort study. MATERIALS AND METHODS: In a prospective birth cohort study at southern Taiwan locating in a subtropical region, questionnaire surveys of sneezing or cough without colds at 6 and 18 months of age were recorded, and the correlation with allergy diseases was assessed at 3 and 6 years of age. RESULTS: A total of 1812 pregnant women and 1848 newborn infants were prenatally enrolled, and 1543, 1344, 1236, and 756 children completed the follow-up at ages 6 months, 18 months, 3 years and 6 years, respectively. The prevalence of infant sneezing without colds at 6 and 18 months of age was 30.3% and 19.2%, respectively. The prevalence of infant cough without colds at 6 and 18 months of age was 10.6% and 5.7%, respectively. Infant sneezing without colds at 18 months of age was significantly correlated with atopic dermatitis, allergic rhinitis and asthma at 6 years of age. Infant cough without colds at 18 months of age significantly predicted asthma but not atopic dermatitis or allergic rhinitis at 6 years of age. CONCLUSIONS: Infant sneezing without colds predicted all allergy diseases at 6 years of age in a subtropical country. This highlights a potential non-invasive clue in a subtropical region for the early prediction, treatment and prevention of childhood allergy diseases in infancy.

A prospective birth cohort study of different risk factors for development of allergic diseases in offspring of non-atopic parents.

BACKGROUND: Allergic diseases are thought to be inherited. Prevalence of allergic diseases has, however, increased dramatically in last decades, suggesting environmental causes for the development of allergic diseases. OBJECTIVE: We studied risk factors associated with the development of atopic dermatitis (AD), allergic rhinitis (AR) and asthma (AS) in children of non-atopic parents in a subtropical country. METHODS: In a birth cohort of 1,497 newborns, parents were prenatally enrolled and validated for allergic diseases by questionnaire, physician-verified and total or specific Immunoglobulin E (IgE) levels; 1,236 and 756 children, respectively, completed their 3-year and 6-year follow-up. Clinical examination, questionnaire, and blood samples for total and specific IgE of the children were collected at each follow-up visit. RESULTS: Prevalence of AD, AR and AS was, respectively, 8.2%, 30.8% and 12.4% in children of non-atopic parents. Prevalence of AR (p<.001) and AS (p=.018) was significantly higher in children of parents who were both atopic. A combination of Cesarean section (C/S) and breastfeeding for more than 1 month showed the highest risk for AD (OR=3.111, p=.006). Infants living in homes with curtains and no air filters had the highest risk for AR (OR=2.647, p<.001), and male infants of non-atopic parents living in homes without air filters had the highest risk for AS (OR=1.930, p=.039). CONCLUSIONS: Breastfeeding and C/S affect development of AD. Gender, use of curtains and/or air filters affect AR and AS, suggesting that control of the perinatal environment is necessary for the prevention of atopic diseases in children of non-atopic parents.

Differentiation of type 2 diabetes mellitus with different complications by proteomic analysis of plasma low abundance proteins.

BACKGROUND: Few biomarkers of type 2 diabetes mellitus (T2DM) are replicable in the differentiation of T2DM with different complications. We aimed to identify proteomic biomarkers among T2DM patients with nephropathy or retinopathy. METHODS: Plasma low abundance proteins were enriched by depletion of 14 high abundance proteins using an affinity removal system, and subjected to nanoflow liquid chromatography electrospray ionization (nano LC-ESI) mass spectrometry after a gel electrophoresis with in-gel digestion. The plasma differential proteomes between normal adults and diabetic patients were validated by another cohort of 149 T2DM patients. RESULTS: A total of 826 proteins in plasma were consistently identified from 8 plasma samples of normal adults, and 817 proteins were consistently identified in 8 plasma samples of T2DM patients. Using the MetaCore analysis, low abundance proteins in plasma between normal adults and T2DM patients were significantly different in 5 functional pathways. Moreover, plasma prolactin-induced protein (PIP), thrombospondin-2 (THBS2), L1 cell adhesion molecule (L1CAM) and neutrophil gelatinase-associated lipocalin (NGAL) levels were higher in T2DM patients. Further, PIP, THBS2 and NGAL were significantly higher in T2DM patients with nephropathy (albuminuria) but not in those with retinopathy, while L1CAM levels were higher in T2DM patients with retinopathy. CONCLUSIONS: This study identified that higher PIP, THBS2 and/or NGAL levels were significantly associated with nephropathy of T2DM, and higher L1CAM but normal PIP, THBS2 or NGAL was significantly associated with retinopathy of T2DM.

Gender-dependent effect of GSTM1 genotype on childhood asthma associated with prenatal tobacco smoke exposure.

It remains unclear whether the GSTM1 genotype interacts with tobacco smoke exposure (TSE) in asthma development. This study aimed to investigate the interactions among GSTM1 genotype, gender, and prenatal TSE with regard to childhood asthma development. In a longitudinal birth cohort in Taiwan, 756 newborns completed a 6-year follow-up, and 591 children with DNA samples available for GSTM1 genotyping were included in the study, and the interactive influences of gender-GSTM1 genotyping-prenatal TSE on childhood asthma development were analyzed. Among these 591 children, 138 (23.4%) had physician-diagnosed asthma at 6 years of age, and 347 (58.7%) were null-GSTM1. Prenatal TSE significantly increased the prevalence of childhood asthma in null-GSTM1 children relative to those with positive GSTM1. Further analysis showed that prenatal TSE significantly increased the risk of childhood asthma in girls with null-GSTM1. Furthermore, among the children without prenatal TSE, girls with null-GSTM1 had a significantly lower risk of developing childhood asthma and a lower total IgE level at 6 years of age than those with positive GSTM1. This study demonstrates that the GSTM1 null genotype presents a protective effect against asthma development in girls, but the risk of asthma development increases significantly under prenatal TSE.

Different genetic associations of the IgE production among fetus, infancy and childhood.

Elevation of serum IgE levels has long been associated with allergic diseases. Many genes have been linked to IgE production, but few have been linked to the developmental aspects of genetic association with IgE production. To clarify developmental genetic association, we investigated what genes and gene-gene interactions affect IgE levels among fetus, infancy and childhood in Taiwan individuals. A birth cohort of 571 children with completion of IgE measurements from newborn to 1.5, 3, and 6 years of age was subject to genetic association analysis on the 384-customized SNPs of 159 allergy candidate genes. Fifty-three SNPs in 37 genes on innate and adaptive immunity, and stress and response were associated with IgE production. Polymorphisms of the IL13, and the HLA-DPA1 and HLA-DQA1 were, respectively, the most significantly associated with the IgE production at newborn and 6 years of age. Analyses of gene-gene interactions indentified that the combination of NPSR1, rs324981 TT with FGF1, rs2282797 CC had the highest risk (85.7%) of IgE elevation at 1.5 years of age (P=1.46 × 10(-4)). The combination of IL13, CYFIP2 and PDE2A was significantly associated with IgE elevation at 3 years of age (P=5.98 × 10(-7)), and the combination of CLEC2D, COLEC11 and CCL2 was significantly associated with IgE elevation at 6 years of age (P=6.65 × 10(-7)). Our study showed that the genetic association profiles of the IgE production among fetus, infancy and childhood are different. Genetic markers for early prediction and prevention of allergic sensitization may rely on age-based genetic association profiles.

Partial protein-hydrolyzed infant formula decreased food sensitization but not allergic diseases in a prospective birth cohort study.

BACKGROUND: Exposure to cow's milk protein in early infancy could lead to increased rates of allergic diseases later in life. We investigated whether feeding a protein-hydrolyzed formula (HF) in the first 6 months of life decreased allergic diseases up to 36 months later. METHODS: Newborns who had at least 1 first-degree family member with a history of atopy and could not breast-feed were enrolled. They were fed with HF or cow's milk infant formula (CM) for at least 6 months via an open-label protocol and were monitored prospectively at 6, 18 and 36 months of age to assess allergy sensitization and allergic diseases. RESULTS: A total of 1,002 infants were enrolled and 679 infants were consistently fed the same formula for the first 6 months of life (345 HF and 334 CM). The percentage of food sensitization (especially to milk protein) was significantly lower in the HF group than in the CM group at 36 months (12.7 vs. 23.4%, p = 0.048). There was no significant difference in the prevalence of aeroallergen sensitization between the groups. Occurrence of allergic diseases during the first 3 years of life was significantly correlated with aeroallergen sensitization, but not to food allergen sensitization, parental atopy or feeding types. CONCLUSIONS: Infants fed with HF during the first 6 months of life had a significantly lower percentage of sensitization to milk protein allergens, but not allergic diseases during the first 3 years of life. Avoidance of cow's milk protein alone in infancy is not enough to decrease rates of allergic diseases.

Increased complement factor H with decreased factor B determined by proteomic differential displays as a biomarker of tai chi chuan exercise.

BACKGROUND: Exhaustive exercise can be associated with short-term immune suppression, but moderate exercise such as tai chi chuan (TCC) has been shown to have beneficial effects on immunity. The mechanisms for the health benefits of exercise remain to be determined, and no potential biomarkers for these beneficial health effects have been identified. This study investigated serum proteomic markers in individuals participating in TCC exercise. METHODS: Two-dimensional fluorescence difference gel electrophoresis was used to compare proteomic markers in 3 individuals before and after 12 weeks of TCC exercise. The different protein spots were identified by mass spectrometry and validated in an additional 20 individuals by western blot analysis. RESULTS: We identified 39 protein spots for 18 proteins with a noticeable increase or decrease after TCC exercise. Validation of the differentially displayed proteins with 20 paired pre- and postexercise samples revealed a significant increase in complement factor H (P = 0.0034) associated with decreases in C1 esterase inhibitor (P = 0.0038) and complement factor B (P = 0.0029). CONCLUSIONS: In this first study of proteomic biomarkers of TCC exercise, we found an increase in complement factor H associated with a decrease in complement factor B. Complement factor H is involved in protection from microangiopathy and macular degeneration and may represent a useful marker of the health effects of exercise.

Sialylated glycans as receptor and inhibitor of enterovirus 71 infection to DLD-1 intestinal cells.

BACKGROUND: Many viruses recognize specific sugar residues, particularly sulfated or sialylated glycans, as the infection receptors. A change of sialic acid (2-6)-linked galactose (SA-alpha2,6Gal) to SA-alpha2,3Gal determines the receptor for avian flu infection. The receptor for enterovirus 71 (EV71) infection that frequently causes fatal encephalitis in Asian children remains unclear. Currently, there is no effective vaccine or anti-virus agent for EV71 infection. Using DLD-1 intestinal cells, this study investigated whether SA-linked glycan on DLD-1 intestinal cells was a receptor for EV71, and whether natural SA-linked sugars from human milk could block EV71 infection. RESULTS: EV71 specifically infected DLD-1 intestinal cells but not K562 myeloid cells. Depletion of O-linked glycans or glycolipids, but not N-linked glycans, significantly decreased EV71 infection of DLD-1 cells. Pretreatment of DLD-1 cells with sialidase (10 mU, 2 hours) significantly reduced 20-fold EV71 replication (p < 0.01). Taken together, these results suggest that SA-linked O-glycans and glycolipids, but not N-glycans, on DLD-1 cells were responsible for EV71 infection. Purified SA-alpha2,3Gal and SA-alpha2,6Gal from human milk significantly inhibited EV71 infection of DLD-1 cells, indicating terminal SA-linked glycans could be receptors and inhibitors of EV71 infection. CONCLUSION: This is the first in the literature to demonstrate that EV71 uses SA-linked glycans as receptors for infection, and natural SA-linked glycans from human milk can protect intestinal cells from EV71 infection. Further studies will test how a SA-containing glycan can prevent EV71 in the future.

Combination of CTLA-4 and TGFbeta1 gene polymorphisms associated with dengue hemorrhagic fever and virus load in a dengue-2 outbreak.

The pathogenesis of dengue hemorrhagic fever (DHF) has been considered to be massive immune activation of T cells. Abnormal expression of the immune regulatory molecules, CTLA-4 and TGFbeta1, leads to disturbances of regulatory T cell immune response. We investigate the contribution of CTLA-4 and TGFbeta1 in DHF by analyzing them for association with virus load in blood and polymorphisms of CTLA-4 +49A/G, and TGFbeta1 -509C/T in a DEN-2 outbreak. The increased frequency of the TGFbeta1 -509 CC genotype in patients with DHF was compared to those with dengue fever (OR=1.9, p=0.034). Moreover, the presence of the CTLA-4 +49 G allele and TGFbeta1 -509 CC genotype increased the susceptibility to risk of DHF (OR=2.1, p=0.028) and significantly higher virus load (p=0.013). This finding suggests that a combination of CTLA-4 and TGFbeta1 polymorphisms is associated with the susceptibility of DHF and higher virus load.

Higher spontaneous and TNFalpha-induced apoptosis of neonatal blood granulocytes.

Granulocytes play an important role in inflammatory diseases. Neonates tend to develop granulocytopenia under sepsis and stress. It remains unclear whether apoptosis of neonatal granulocytes is different from that of adult granulocytes. In this study, we analyzed the discrepancy of granulocyte apoptosis between cord blood (CB) and adult blood (AB). We found that spontaneous and tumor necrosis factor-alpha (TNFalpha)-induced granulocyte apoptosis as determined by phosphatidylserine expression and DNA fragmentation were more prominent in CB granulocytes than AB granulocytes. CD95 ligand and TNFalpha levels were significantly higher in CB than in AB (p = 0.001 and p < 0.001, respectively). TNF receptor-2 and CD95 expression on CB granulocytes were not different from those on AB granulocytes. However, the TNF receptor-1 (TNFR1) expression was lower on CB granulocytes than that on AB granulocytes (69.98 +/- 7.32 versus 89.04 +/- 3.73%, p = 0.029). This decrease of TNFR1 expression on neonatal granulocytes might be related to a higher plasma TNFalpha level associated with an intrinsic defect on the dynamic change of membrane TNFR1 expression in neonatal granulocytes. The expression of anti-apoptotic molecule Bcl-2 in neonatal granulocytes was also lower than that in adult granulocytes (4.64 +/- 0.51 versus 7.24 +/- 1.17 unit/mg protein, p = 0.032). Moreover, another anti-apoptotic signal, nuclear factor-kappaB nuclear translocation, was also lower in CB than AB granulocytes. Results from this study suggest that higher plasma death ligands associated with lower anti-apoptotic molecules in granulocytes may act an important role in triggering neonatal granulocyte apoptosis.

Gender-limited association of cytotoxic T-lymphocyte antigen-4 (CTLA-4) polymorphism with cord blood IgE levels.

Allergic mechanism has long been attributed to IgE-mediated reaction. The relationship between gene polymorphism and cord blood IgE (CB IgE) is unclear. We investigated whether elevation of CB IgE levels was associated with polymorphisms of cytotoxic T-lymphocyte antigen 4 (CTLA-4) at (-318) CT and (+49) AG positions in a gender-limited fashion. CB IgE levels were determined by Pharmacia CAP system and the CTLA-4 polymorphisms at (-318) and (+49) were determined by restriction fragment length polymorphism (RFLP). A total of 644 consecutive umbilical cord bloods were collected for this study. 32.9% of newborn infants had detectable IgE levels (> or =0.35 kU/l). 25.6% of the male newborns had elevated CB IgE levels (> or =0.5 kU/l) similar to those of the female newborns (22.7%). The CTLA-4 polymorphism at (+49) but not (-318) was significantly associated with elevated CB IgE levels (p = 0.004). The association of CTLA-4 (+49) A allele with elevated CB IgE levels was found only in females. Both male and female infants with different CTLA-4 (-318) genotypes had no difference in the rates of elevated CB IgE levels. A linkage disequilibrium between CTLA-4 (+49) G and (-318) C allele was found in this Chinese population. Subjects with the (+49, GG and -318, CC) genotype had a significantly lower rate of elevated CB IgE levels. Association of the CTLA-4 (+49) polymorphism with elevated CB IgE levels was found only in female infants. Newborn infants with the (+49, GG and -318, CC) genotype tended to have a low rate of elevated CB IgE.

Association of cord blood cytokines with prematurity and cerebral palsy.

BACKGROUND: Many studies have shown that certain cytokines in amniotic fluids are correlated to premature labor and neonatal brain insults. AIMS: We investigated whether different fetal phagocyte and vascular mediators including IL-8, myeloperoxidase (MPO), PGE(2) and soluble vascular cell adhesion molecule-1 (sVCAM-1) levels were correlated to prematurity and cerebral palsy (CP) of premature infants. SUBJECTS: Umbilical cord blood samples from 96 preterm babies from 2250 cord blood collections were studied. METHOD: The enzyme-linked immunoassay (ELISA) technique was used to determine concentrations of cord blood IL-8, MPO, PGE(2) and sVCAM-1. RESULTS: Preterm babies with gestational age (GA) < or =32 weeks had significant higher cord blood IL-8, MPO and PGE(2) levels than full-term babies. These mediators, however, were not correlated to gestational age, suggesting that increases of these mediators are more related to preterm delivery but not fetal maturation. Further analysis showed that IL-8, a mediator mainly from monocytes, but not MPO, another mediator mostly from granulocytes, was correlated to cerebral palsy. CONCLUSIONS: Taken together, these results suggest that both premature monocytes and granulocytes activation are involved in preterm delivery, but maybe only monocytes activation is correlated to premature infants' cerebral palsy. Selective manipulation of monocytes activation may be useful to prevent premature-related cerebral palsy.

Prenatal prediction of infant atopy by maternal but not paternal total IgE levels.

BACKGROUND: The atopic history of parents has long been used to predict infant atopy. However, bias from questionnaires of allergic history are also frequently suspected, because a large number of vasomotor rhinitis, intrinsic asthma, and seborrheic dermatitis cases are probably misinterpreted to be atopic diseases. OBJECTIVE: We attempted to identify a risk factor other than parental atopic history to predict elevated infant IgE levels and infant atopy. METHODS: A total of 655 core families were prenatally recruited, and finally 545 families completed the study for the prospective analysis of infant atopy at 6 months of age. Atopic history and blood samples of parents were collected in the third trimester during pregnancy. Cord blood (CB) was collected immediately after birth. Infant blood samples and history of infant eczema were collected in the 6-month physical checkup clinic. Blood total IgE and specific IgE levels were determined by use of the Pharmacia CAP system. RESULTS: In univariate analysis, maternal, but not paternal, atopic history correlated with elevated CB IgE levels and the occurrence of infant eczema. Elevated maternal, but not paternal, total IgE levels (>150 KU/L) significantly correlated with increases of CB IgE levels (median, 0.54 vs 0.17 KU/L, P <.001), infant IgE levels (log-transformed mean values, 1.32 +/- 0.51 vs 1.13 +/- 0.51 KU/L, P <.001), and infant eczema (P =.008). Multivariate logistical regression analysis, however, showed that only maternal total IgE levels correlated with CB and infant IgE levels and the development of infant eczema. CONCLUSIONS: The maternal, but not paternal, total IgE level correlates with elevated infant IgE levels and infant atopy. This provides a high specificity (83%) and a sensitivity of 34% for prediction of infant atopy. This suggests that maternal factors, placental factors, or both have an impact on perinatal allergic sensitization.

Prediction of elevated cord blood IgE levels by maternal IgE levels, and the neonate's gender and gestational age.

BACKGROUND: Elevation of cord blood immunoglobulin E (IgE) is used to predict childhood atopy. In an effort to catch such problems at an earlier time, we sought to identify the impact of parental atopy as well as gestational age, the baby's gender, and the season of delivery on cord blood IgE levels. METHODS: The allergic history of parents was collected during pregnancy. Blood samples from parents were collected in the third trimester. Cord blood was collected immediately after birth. Total and specific IgE levels were determined using the Pharmacia CAP system. RESULTS: In total, 437 core family blood samples were collected. Male babies had a significantly higher IgE level (0.535 +/- 0.898 vs. 0.369 +/- 0.565 KU/L, p = 0.021) and a higher frequency of IgE > or = 1.0 KU/L (14.6% vs. 7.5%, p = 0.018) compared to female babies. A cyclic trend in higher cord blood IgE levels was found in babies born in early summer and early winter. Multiple logistic regression analyses revealed that elevation of cord blood IgE levels could be predicted by higher maternal IgE levels (odds ratio [OR] = 6.35; p = 0.000), male baby gender (OR = 2.31; p = 0.021), and increases in gestational age by 1 week (OR = 1.34; p = 0.039). In contrast, neither the allergic history of parents nor elevation of paternal IgE levels could be correlated with elevated cord blood IgE levels of neonates. CONCLUSION: The baby's gender and gestational age, and maternal IgE levels influence cord blood IgE levels. Avoiding allergens and decreasing allergic activities during pregnancy may be the most important means of preventing the fetus from having allergic sensitization.

Different antigens trigger different Th1/Th2 reactions in neonatal mononuclear cells (MNCs) relating to T-bet/GATA-3 expression.

Neonates are known to have poor cellular immunity, especially poor Th1 response. We investigated how neonatal mononuclear cells raised different Th1/Th2 reactions in response to different antigens. Employing Dermatophagoides pteronyssinus (Der p) extract and varicella zoster virus (VZV) as antigens, we assessed Th1/Th2 reactions as demonstrated by IL-4/IFNgamma production and mRNA expression, and transcriptional factors T-bet/GATA-3 mRNA expression in mononuclear cells from human umbilical cord blood (CBMC). Results showed that VZV induced a dramatic increase of IFNgamma production by adult peripheral blood mononuclear cells (PBMC), whereas VZV did not drive CBMC to release significant IFNgamma production (1614.7+/-362.0 vs. 49.0+/-29.3,p<0.005). However, Der p induced higher IFNgamma production by CBMC than VZV (298.1+/-171.8 vs. 49.0+/-29.3, P=0.047). In contrast, VZV did not induce significant IL-4 production either by CBMC or by PBMC. Der p induced a comparative IL-4 production by CBMC and PBMC (2.58+/-0.84 vs. 2.04+/-0.37, p>0.05). A real-time RT-PCR analysis of IL-4 and IFNgamma mRNA expression showed that VZV induced a significantly higher IFNgamma, but not IL-4, mRNA expression in PBMC than CBMC. Der p did not induce significant difference of IFNgamma or IL-4 mRNA expression in PBMC and CBMC. VZV enhanced Th1-related transcription factor T-bet mRNA expression, in association with later down-regulation of Th2-related GATA-3 mRNA expression in PBMC. However, VZV did not up-regulate T-bet or down-regulate GATA-3 expression significantly in CBMC. In contrast, Der p induced an early GATA-3 expression and later T-bet expression in CBMC. These results suggest that different antigens trigger various Th1/Th2 reactions in PBMC and CBMC resulting from kinetic changes of T-bet/GATA-3 expression.

Glutamine protects activated human T cells from apoptosis by up-regulating glutathione and Bcl-2 levels.

Glutamine is the most abundant amino acid in the body. A decrease of plasma glutamine concentrations is found in catabolic stress and is related to susceptibility to infections. Glutamine is known to modulate lymphocyte activation; however, little is known about glutamine modulation of cell death of activated human T cells. Using Jurkat T cells, we investigated glutamine modulation of T-cell apoptosis activated by PMA plus ionomycin. We found that glutamine at various concentrations significantly enhanced IL-2 production, cell proliferation, and cell viability of Jurkat T cells. Glutamine also decreased the number of apoptotic cells stimulated with PMA plus ionomycin as demonstrated by flow cytometry. Meanwhile, glutamine down-regulated CD95 and CD95L expression, but up-regulated CD45RO and Bcl-2 expression in activated T cells. Further investigation of CD95-mediated caspase activities revealed that supplementation of glutamine significantly decreased caspase-3 and caspase-8 activities in activated T cells. Since oxidative stress is closely associated with induction of lymphocyte apoptosis, we found that glutamine significantly increased glutathione (GSH), but decreased reactive oxygen species levels in activated T cells. Blockade of intracellular GSH formation enhanced, but exogenous GSH supplementation decreased, activated T-cell apoptosis. Studying normal peripheral lymphoproliferation, we also found that the presence of glutamine increased lymphoproliferation as well as Bcl-2 and CD95 expression; but decreased CD95L and activation-induced T-cell death. Taken together, glutamine appeared to augment lymphoproliferation but suppressed activation-induced T-cell death in both Jurkat T cells and human peripheral T lymphocytes.

Reversible phosphatidylserine expression on blood granulocytes related to membrane perturbation but not DNA strand breaks.

Exposure of phosphatidylserine (PS) on the outer leaflet of the cell membrane is recognized as an early indicator of programmed cell death (apoptosis) in plant and mammalian cells. Currently, there is no literature describing that PS expression on the surface of white blood cells is reversible. We found that a hypotonic 0.2% NaCl or NH(4)Cl lysing solution used to separate white blood cells from red blood cells induced a reversible PS expression on the cell surface of granulocytes and monocytes but not lymphocytes. This reversible PS expression was associated with change of plasma membrane potential but not degranulation-associated membrane mobilization or DNA fragmentation. In contrast, TNF-alpha induced an irreversible PS expression, associated with apoptotic DNA fragmentation shown on gel electrophoresis. The fact that hypotonic shock induced a reversible PS expression on granulocytes, and TNF-alpha induced an irreversible PS expression associated with apoptotic DNA fragmentation indicate the new insight that expression of PS on the outer cell surface does not always represent cell apoptosis. Also, the reversible PS expression was associated with altered plasma-membrane potential but not DNA strand breaks, indicating that early PS expression may be related to the membrane perturbation but not directly related to DNA fragmentation in certain types of cells.