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Context The levels of adrenal androgens are increased through the action of steroidogenic enzymes with morphological changes in the adrenal zona reticularis. Objective We investigated longitudinal changes in androgen levels and steroidogenic enzyme activities during early childhood. Design and Participants From a prospective children’s cohort, the Environment and Development of Children cohort, 114 boys and 86 girls with available blood samples from ages 2, 4, and 6 years were included. Outcome Measurements Serum concentrations of adrenal androgens using liquid chromatography-tandem mass spectrometry and steroidogenic enzyme activity calculated by the precursor/product ratio. Results During ages 2 to 4 years, 17,20-lyase and dehydroepiandrosterone (DHEA) sulfotransferase activities increased (P Conclusion This study supports in vivo human evidence of increased 17,20-lyase and DHEA sulfotransferase activities and decreased 3β-HSD activity during early childhood.

Patients with craniopharyngioma are susceptible to autonomic dysfunction as a result of hypothalamic damage. We evaluated indices of heart rate variability (HRV) in patients with childhood-onset craniopharyngioma to investigate autonomic function and its relationship with components of the metabolic syndrome (MetS). This cross-sectional, case-only study included 53 patients (10–30 years of age). We measured the standard deviation of all normal R-R intervals (SDNN) and total power indicating overall HRV, the root-mean square of the difference of successive R-R intervals (RMSSD) and high frequency indicating parasympathetic modulation, and low frequency. These indices were compared according to the presence of the MetS. During the mean 10.8 years of follow-up, 25% of patients were diagnosed with the MetS. Patients with the MetS showed significantly lower levels of SDNN (29.0 vs. 40.6 ms), total power (416.1 vs. 1129.6 ms2), RMSSD (20.1 vs. 34.5 ms), high frequency (94.7 vs. 338.5 ms2), and low frequency (94.5 vs. 289.4 ms2) than those without (p

Low-iodine diet (LID) is a crucial preparation for radioactive iodine (RAI) treatment or scan in thyroid cancer. The aim of this study is to analyze the influence of thyroid stimulating hormone (TSH) stimulation protocols and other clinical factors on LID adequacy. Thyroid cancer patients who underwent LID for RAI scan or treatment were retrospectively analyzed. Patients were guided to have LID for 2 weeks before RAI administration and urine iodine/creatinine ratio (UICR, μg/g Cr) was measured. TSH stimulation was conducted using either thyroid hormone withdrawal (THW) or recombinant human TSH (rhTSH) injection. Adequacy of LID was classified by UICR as ‘excellent (< 50)’, ‘adequate (50–100)’, ‘inadequate (101–250)’ and ‘poor (> 250)’. A total of 1715 UICR measurements from 1054 patients were analyzed. UICR was significantly higher in case of rhTSH use than THW (72.4 ± 48.1 vs. 29.9 ± 45.8 μg/g Cr, P < 0.001). In patients who underwent LID twice using both TSH stimulation protocols alternately, UICR was higher in case of rhTSH than THW regardless of the order of method. Among clinical factors, female, old-age, and the first LID were significant factors to show higher UICR. Although the adequacy of LID was ‘adequate’ or ‘excellent’ in most patients, multivariate analysis demonstrated that THW method, male, young age, and prior LID-experience were significant determinants for achieving ‘excellent’ adequacy of LID. In conclusion, UICR was higher and the proportion of ‘excellent’ LID adequacy was lower with rhTSH than with THW. UICR was higher also in women, old-age, and LID-naïve patients. Further researches are required to suggest effective methods to reduce body iodine pool in case of rhTSH use and to validate the efficacy of such methods on outcomes of RAI treatment.

Accurate investigations of adrenal hormone levels play a vital role in pediatric endocrinology for the detection of steroid-related disorders. In this study, we developed and validated a simultaneous assay for eight serum steroids, i.e., DHEA, androstenedione, testosterone, progesterone, 17‑hydroxyprogesterone, DHEA‑sulfate, pregnenolone‑sulfate and cholesterol-sulfate, using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Serum samples were prepared by liquid-liquid extraction with methyl t‑butyl ether. Quantitation by LC-MS/MS was performed in multiple reaction monitoring mode with an electrospray ionization source. The run time was 10 min. Analytical performance was evaluated, including imprecision, linearity, ion suppression, carry over and detection capabilities. Serum specimens from 59 children and 120 adults were analyzed to compare the distribution of steroid levels between the groups. All hormones were effectively extracted and separated using our method. The method was essentially free from potential interference and ion suppression. Within-run and between-run imprecision values were20%. The lower limits of quantification varied from 0.025 to 10 ng/mL. The results were generally good and correlated with those obtained using immunoassay techniques. We developed the HPLC-MS/MS method for the simultaneous measurement of free and sulfated steroid hormones. The performance of the developed method was generally acceptable. Thus, this method may provide a novel approach to steroid profiling in children of age before adrenarche.

Background Pyrethroid pesticides are reported to be the most commonly used residential insecticides worldwide. We aimed to investigate the relationship between prenatal and postnatal 3-phenoxybenzoic acid (3-PBA) concentrations, and growth and adiposity parameters in 4-year-old children. Method We obtained data from 578 children who participated in the prospective Environment and Development of Children (EDC) study at around 4 years of age (45–55 months) between August 2008 and July 2011. Anthropometric measurements were obtained at age 4 years. Prenatal and postnatal urinary 3-PBA concentration was measured in maternal urine samples at around 20 weeks of gestation, and in the 4-year-old children, respectively. Result The detection frequency of urinary 3-PBA (geometric mean concentration) was 98–99% (0.98 μg/g Cr) in maternal urine, and almost 99–100% (1.34 μg/g Cr) in 4-year-old children. Prenatal urinary3-PBA concentration was not associated with height, weight, or body mass index (BMI) z-scores at 4 years of age, regardless of sex. Postnatal urinary3-PBA concentration was not related to height z-scores, but was positively associated with weight z-scores with marginal significance among only girls (p = 0.058). Analyzed by sex, there was a significant relationship between postnatal urinary 3-PBA concentration and BMI z-scores (p = 0.015) among girls, after adjusting for covariates. Conclusion Childhood urinary 3-PBA concentration measured at 4 years of age was positively associated with BMI z-scores in 4-year-old girls, but prenatal urinary 3-PBA concentration at midterm pregnancy exhibited no association.

Background Previous studies have suggested links between exposure to ambient air pollutants and increased risk of congenital heart defects. However, few studies have investigated the association between other congenital diseases and traffic-related air pollution. In this study, we assessed the relationship between prenatal exposure to fine particulate matter (PM2.5) and nitrogen dioxide (NO2) with congenital diseases in South Korea. Methods Patients with one or more congenital diseases and a control group of patients with non-infective gastroenteritis and colitis with a case:control ratio of 1:3 were obtained from the National Health Insurance Service data for 2008–2013 in South Korea. We estimated the associations of PM2.5 and NO2 exposures with congenital diseases using generalized estimation equations after controlling for covariates. Results Maternal PM2.5 exposure during the first and second trimester showed positive associations with overall congenital diseases, with changes of 14.7% (95% confidence intervals (CI), 9.3%, 20.3%) and 16.2% (95% CI, 11.0%, 21.7%), respectively, per 11.1 μg/m3 and 10.2 μg/m3 increase of PM2.5 interquartile range (IQR). Similarly, NO2 exposure during the first and second trimester was associated with increased numbers of overall congenital anomalies, with 8.2% (95% CI, 4.2%, 12.3%) and 15.6% (95% CI, 9.3%, 22.2%) more cases, respectively, per 10.6 ppb increase of NO2. We found that maternal PM2.5 exposure during the first and second trimesters of pregnancy was significantly associated with increased risk of specific congenital diseases, including subtypes affecting the circulatory, genitourinary, and musculoskeletal system. However, no significant associations were observed during the third trimester. Maternal NO2 exposure across the entire pregnancy was associated with malformations of the musculoskeletal system. Conclusions Our study identified significant links between in utero exposure to PM2.5 and NO2 and certain congenital diseases, and suggests that stricter controls on PM2.5 and NO2 concentrations are required.

Objectives Vitamin D is essential for bone health. Not only total but also free 25-hydroxyvitamin D (25OHD) may contribute to bone mass. We sought to determine which vitamin D measure best reflected clinical and bone parameters in healthy children. Methods A cross-sectional study including 146 healthy children (71 boys, 9.5 ± 1.9 years) conducted at a tertiary medical center. We used a multiplex liquid chromatography-tandem mass spectrometry-based assay to simultaneously measure vitamin D metabolites. The bioavailable and free 25OHD (25OHDBioA and 25OHDFree) levels were calculated using the genotype-specific or genotype-constant affinity coefficients of vitamin D-binding proteins (yielding spe-25OHDBioA, spe-25OHDFree and con-25OHDBioA, con-25OHDFree respectively). The 25OHDFree level was directly measured (m-25OHDFree). Bone mineral content (BMC) and bone mineral density (BMD) were assessed via dual-energy X-ray absorptiometry. Results The total 25OHD (25OHDTotal), the two forms of 25OHDBioA, the three forms of 25OHDFree, and 24,25-dihydroxyvitamin D3 levels correlated with parathyroid hormone level (all p < 0.01). Serum 25OHDTotal and m-25OHDFree levels were influenced by age, pubertal status, season, body mass index (BMI), daylight hours, and vitamin D intake (all p < 0.05). The con-25OHDBioA and con-25OHDFree levels better reflected pubertal status and daylight hours than did the spe-25OHDBioA and spe-25OHDFree levels (both p < 0.01). The association between the 25OHDTotal level and bone parameters varied according to the BMI (interaction p < 0.05). In 109 normal-weight children, the con-25OHDBioA and con-25OHDFree levels correlated with total body BMC and BMD (both p < 0.05), whereas the 25OHDTotal and 24,25-dihydroxyvitamin D3 levels were associated with total body BMC (both p < 0.05). No such association was found in overweight or obese children. Conclusions In healthy children, total, bioavailable, and free 25OHD levels comparably reflected lifestyle factors. In normal-weight children, the con-25OHDBioA and con-25OHDFree, but not m-25OHDFree levels, reflected bone mass, as did the 25OHDTotal level.

Background/Aim Phthalate is a well-known endocrine-disrupting chemical that has anti-androgenic effects. Although there are several studies on the relationship between body composition and phthalate, studies that investigated the effects of phthalate on skeletal muscle during childhood are lacking. Methods We analyzed data from 481 mother-and-child pairs enrolled in the Environment and Development of Children cohort in South Korea. We examined the association between phthalate metabolites (mono [2-ethyl-5-hydroxyhexyl] phthalate [MEHHP], mono [2-ethyl-5-oxohexyl] phthalate [MEOHP], molar sum of MEHHP and MEOHP [Σ DEHP], and mono-n-butyl phthalate [MnBP]) in prenatal maternal urine and children's urine at the age of 6, and body composition indices (body mass index [BMI] z-score, percentage of fat mass, fat mass index, percentage of skeletal muscle, and the skeletal muscle index [SMI]) measured when the child was 6 years using a bioelectrical impedance analyzer. Results A 2-fold increase in Σ DEHP and MnBP in the prenatal maternal urine was significantly associated with a −0.07 unit (95% CI: −0.11, −0.03) and −0.09 unit (95% CI: −0.14, −0.03) change in SMI, respectively, in 6-year old girls alone. BMI z-score was also negatively associated with a 2-fold increase in MEHHP and MnBP in prenatal maternal urine as −0.11 unit (95% CI: −0.22, −0.01) and −0.15 unit (95% CI: −0.28, −0.02) change, respectively, only among girls. Among boys, phthalate metabolites in the prenatal and children's urine were not significantly associated with any body composition indices. Conclusions Our longitudinal study shows that high levels of prenatal exposure to phthalates are significantly associated with decreased SMI among girls. We can postulate that anti-androgenic effects of phthalates during pregnancy may affect girl offspring's muscle growth.

Previous animal studies have reported that pyrethroids can cause dopamine system abnormalities and attention-deficit/hyperactivity disorder (ADHD) phenotypes. However, epidemiological studies investigating the associations between pyrethroid exposure and ADHD are limited. We aimed to investigate the association between pyrethroid exposure and ADHD-like symptoms among preschool-age children. We used data from 385 children at 4 years of age participating in the Environment and Development of Children (EDC) study. We evaluated pyrethroid exposure through questionnaires and urinary 3-phenoxybenzoic acid (3-PBA) concentrations. We assessed ADHD-like symptoms using the Korean ADHD rating scale (K-ARS). We conducted negative binomial regressions to evaluate the associations between pyrethroid exposure and ADHD-like symptoms. Residential use of insecticide adhesive (β = 0.42, 95% CI: 0.11, 0.74) and insecticide spray (β = 0.33, 95% CI: 0.08, 0.59) was associated with an increase in log-transformed creatinine-adjusted urinary 3-PBA concentrations. Residential insecticide adhesive use was associated with a 51.6% increase in K-ARS scores (95% confidence interval [CI]: 6.3, 116.1) among boys, when compared with non-users. When compared with creatinine-adjusted 3-PBA levels0.50 μg/g creatinine, creatinine-adjusted 3-PBA levels ≥3.80 μg/g creatinine were associated with a 58% increase in K-ARS scores (95% CI: 0.1, 150.5) among boys. We found associations of residential pyrethroid insecticide use and urinary 3-PBA concentrations with K-ARS scores among preschool-age boys. Since the present study explored cross-sectional associations in preschool-age children, the possibility of reverse causality cannot be dismissed. Further studies implementing a cohort study design are warranted.

Purpose Monogenic diabetes is a specific type of diabetes in which precision medicine could be applied. In this study, we used targeted panel sequencing to investigate pathogenic variants in Korean patients with clinically suspected monogenic diabetes. Methods The eligibility criteria for inclusion were patients with nontype 1 diabetes with age at onset ≤30 years and body mass index (BMI) ≤30 kg/m2. Among the 2090 patients with nontype 1 diabetes, 109 had suspected monogenic diabetes and underwent genetic testing. We analyzed 30 monogenic diabetes genes using targeted panel sequencing. The pathogenicity of the genetic variants was evaluated according to the American College of Medical Genetics and Genomics and Association for Molecular Pathology guidelines. Results Among the 109 patients with suspected monogenic diabetes, 23 patients (21.1%) harbored pathogenic/likely pathogenic variants. A total of 14 pathogenic/likely pathogenic variants of common maturity-onset diabetes of the young (MODY) genes were identified in GCK, HNF1A, HNF4A, and HNF1B. Other pathogenic/likely pathogenic variants were identified in WFS1, INS, ABCC8, and FOXP3. The mitochondrial DNA 3243A>G variant was identified in five participants. Patients with pathogenic/likely pathogenic variants had a significantly higher MODY probability, a lower BMI, and a lower C-peptide level than those without pathogenic/likely pathogenic variants (P = 0.007, P = 0.001, and P = 0.012, respectively). Conclusions Using targeted panel sequencing followed by pathogenicity evaluation, we were able to make molecular genetic diagnoses for 23 patients (21.1%) with suspected monogenic diabetes. Lower BMI, higher MODY probability, and lower C-peptide level were characteristics of these participants.

Context:Large-sample studies with long-term follow-up data are limited for pediatric patients with thyroid cancer.Objective:Secular changes in clinicopathological characteristics and outcomes in pediatric patients with thyroid cancer were investigated and compared with those of adults.Design and Patients:A retrospective review of 150 pediatric patients with thyroid cancer managed between 1980 and 2013 was conducted. The long-term outcomes of 124 patients followed up for 12 months or longer were evaluated. Predictors of recurrence-free survival (RFS) in pediatric patients with papillary thyroid cancer (ped-PTC group) were compared with those of 3071 adult patients.Results:The proportion of small tumors (Conclusions:The pathological characteristics and recurrence rates of pediatric thyroid cancer have not changed over 33 years. Although younger patients present with more advanced disease, multifocality rather than age at diagnosis predicted recurrence. Recurrence was higher in pediatric than adult patients with multifocal papillary thyroid cancer.

The obligatory intracellular protozoan parasite Entamoeba histolytica causes amebic dysentery and liver abscess. E. histolytica adheres to the host tissues in a contact-dependent manner. E. histolytica excretory-secretory products (ESP) might play critical roles during invasion. We comparatively analyzed the secretome profile of E. histolytica pathogenic HM-1:IMSS and non-pathogenic Rahman strains. The two ESP revealed similar but distinct spotting patterns. In both ESP, alcohol dehydrogenase, enolase 1, and transketolase, which control classical carbohydrate metabolism and other moonlighting effects, constituted the most abundant fractions. We recognized differently secreted molecules. Secretion of cytoskeletal organization proteins (actin, actin binding protein, and EHI_068510), protein remodeling amino peptidase, and multifunctional elongation factor 1-α were increased in Rahman. Conversely, carbohydrate metabolizing enolase 1, alcohol dehydrogenase, transketolase, calponin, phosphoglucose mutase, malic enzyme and EHI_156420, xenobiotic scavenging superoxide dismutase and EHI_140740, and pyruvate:ferredoxin oxidoreductase and coronin (carbohydrate metabolism/detoxification) showed reduced secretion. Transcription levels of some genes involved in these processes also decreased. Changes of secretory behavior, especially decreased secretion of multifunctional carbohydrate metabolizing enzymes and detoxifying proteins that importantly participated in amoeba pathogenesis might reflect avirulent nature of Rahman strain in the host.

ObjectiveWe evaluated frequency and risk factors of delayed TSH elevation (dTSH) and investigated follow-up outcomes in the dTSH group with venous TSH (v-TSH) levels of 6-20 mU/L according to whether late preterm infants born at gestational age (GA) 35-36 weeks had risk factors.MethodsThe medical records of 810 neonates (414 boys) born at Seoul National University Hospital who had a normal neonatal screening test (NST) and underwent the first repeat venous blood test at 10-21 days post birth were reviewed.ResultsSeventy-three (9.0%) neonates showed dTSH, defined as a v-TSH level ≥6.0 mU/L, 12 of whom (1.5%) were started on levothyroxine medication. A multivariate-adjusted model indicated that a low birth weight (LBW ConclusionsdTSH was detected in 9.0% and levothyroxine was indicated in 1.5% of infants born at GA 35-36 weeks, particularly those with a LBW, a congenital anomaly, or history of ICM exposure. Either levothyroxine or retesting is indicated for late preterm neonates with TSH levels ≥10 mU/L regardless of risk factors. If healthy preterm neonates show v-TSH levels of 6-10 mU/L, a second repeat test may not be necessary; however, further studies are required to set a threshold for retesting.

The primary purpose of this study was to investigate the effects of phthalate exposure on the intelligence and attentional performance of 6-year old children when adjusting each other as covariates. We also investigated the differential effects of phthalate exposure on the intelligence and attention according to exposure period (maternal or children). Urine concentrations of mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP), and mono-n-butyl phthalate (MBP) were analyzed. Multivariable linear regression models were used to investigate the relationship between exposure to various phthalates with IQ scores and continuous performance test (CPT) variables. There were robust associations between child MEHHP and MEOHP levels with full scale IQ (FSIQ) even after adjusting for demographic variables and CPT scores (MEHHP -9.27, 95% CI: -17.25, -1.29; MEOHP -9.83, 95% CI: -17.44, -2.21). Child MEHHP and MEHOP levels negatively affected omission errors (MEHHP -20.36, 95% CI: -34.17, -6.55; MEOHP -18.93, 95% CI: -32.58, -5.28) and the response time variability (MEHHP -21.07, 95% CI: -39.04, -3.10; MEOHP -20.41, 95% CI -38.14, -2.69) of the CPT after adjusting for demographic variables and IQ. Maternal phthalate exposure had no effects on IQ or CPT variables. These results suggest that children phthalate exposure, but not maternal exposure, has an adverse effect on IQ and attentional performance, and these associations were found to be independent of each other.

Entamoeba histolytica is an extracellular tissue parasite causing colitis and occasional liver abscess in humans. E. histolytica-derived secretory products (SPs) contain large amounts of cysteine proteases (CPs), one of the important amoebic virulence factors. Although tissue-residing mast cells play an important role in the mucosal inflammatory response to this pathogen, it is not known whether the SPs induce mast cell activation. In this study, when human mast cells (HMC-1 cells) were stimulated with SPs collected from pathogenic wild-type amoebae, interleukin IL-8 mRNA expression and production were significantly increased compared with cells incubated with medium alone. Inhibition of CP activity in the SPs with heat or the CP inhibitor E64 resulted in significant reduction of IL-8 production. Moreover, SPs obtained from inhibitors of cysteine protease (ICP)-overexpressing amoebae with low CP activity showed weaker stimulatory effects on IL-8 production than the wild-type control. Preincubation of HMC-1 cells with antibodies to human protease-activated receptor 2 (PAR2) did not affect the SP-induced IL-8 production. These results suggest that cysteine proteases in E. histolytica-derived secretory products stimulate mast cells to produce IL-8 via a PAR2-independent mechanism, which contributes to IL-8-mediated tissue inflammatory responses during the early phase of human amoebiasis.

Background: Viral infections cause significant morbidity & mortality in patients undergoing unrelated allogeneic BMT transplantation before immune reconstitution is completed. It poses a greater risk for recipients of unrelated umbilical cord blood (uUCB) transplants as there is no established antiviral immunity in naive UCB lymphocytes available for adoptive transfer. UCB T cells also lack Th1/Tc1 cytokines, Granzymes & Perforin which are prerequisites to control viral pathogens. Another major limitation of uUCBT is the lack of donor cells available for post-transplant donor leukocyte infusions (DLI) to boost immunity or induce GVL. However, a fraction of the uCB graft could be available for T cell expansion. In this study we evaluated the feasibility of ex vivo expansion of UCB T cells. We postulated that following expansion naive T cells may mature & acquire a phenotype compatible with effector function as assessed by expression of essential cytokines & de novo expression of members of the granzyme-perforin pathway. Methods: Thawed UBC research samples with a leukocyte content

Unrelated cord blood transplantation (UCBT) is a viable option for patients who lack HLA-matched sibling donors. However, opportunistic infections (OI), occurring in the first 100 days, remain the major cause of morbidity and mortality. Viral infections are the primary cause of OI death. Despite the large number of T cells in UCB grafts there is no established antiviral immunity in nave UCB lymphocytes available for adoptive transfer. UCB T cells also lack expression of Th1/Tc1 cytokines, Granzymes A, B, and Perforin which are prerequisite cytotoxic effector molecules for controlling viral pathogens. We postulated that following transplant the infused naïve T cells will be critically influenced by the lymphopenic environment and emerging de novo acquired or latent viruses. Phenotypic alterations induced by these pathogens may identify those at risk for OI even before virological/clinical diagnosis is evident. Since our previous analysis of d+ 50 immune profile could not fully separate true risk factors from immunological correlates/consequences of OI we designed a new study. Methods: 70 pediatric recipients of HLA-mismatched UCBT were studied prospectively as soon as the WBC exceeded 0.3/ul, at a median of day+18, typically before myeloid engraftment and prior to the onset of clinical infections. Unprocessed whole blood was stained in “Trucount” (B-D) tubes for absolute T cell and dendritic cell (DC1, DC2) counts, while T cells subsets were similarly analyzed by 4 color FACS from Ficolled specimens for lymphocyte subsets, their activation state, tissue homing molecules. Intracellular (ic) staining identified cytotoxic effectors and inducible cytokine secretion besides characterizing homeostatic proliferation and apoptosis. 2-tailed student’s t-test identified significant variables (p Results: 39/70 patients developed OI at a median of day +35 (37 viral cases, 1 case of Legionella and 1 Candida), in average 2weeks after immune studies. While only 2/31 non-OI patients died (relapse), 18/39 from the OI group died at a median of 82 days post UCBT, p=0.001. Median f/up of survivors is 28 ± 17 months. Age, malignancy, cell dose had no impact on OI presence. The median values for major leukocytes were; WBC: 0.7×10e3/ul, T cells: 66/ul, NK cells: 46/ul, B cells: 0/ul, CD4+T cells: 42/ul, %CD4+ T cells: 64%. Both the OI+ and OI negative group had comparable WBC, CD3+, CD4+ T cells, NK lymphocytes, or DC1, DC2 subsets. Strikingly, 44% of circulating T cells were in cell cycle (KI-67+) regardless of OI status reflecting vigorous homeostatic expansion with only 16% preserving the CD45RA+/CD62L+ phenotype. Notably, ~9% of the circulating T cells were entering apoptosis (activated Caspase-3+), regardless of OI status. In those who eventually developed OI significantly more CD8+ cells circulated (40% vs 28%, p=0.03), more expressed CCR-5 (82% vs 55%, p=0.009), and more T cells were secreting IFNγ (35% vs 12%, p=0.006). In patients who developed OI significantly more Perforin+/CD8+ T cells circulated (49% vs 26%, p=0.01) and more of them expressed Granzyme B (p=0.05). There was higher BCL-2 expression in T cells from patients developing OI (p=0.04), and greater presence of CD28−/CD57+/CD8+ ‘effector’ CTLs (p=0.014). We conclude that in lymphopenic UCBT recipients subclinical viral infections may skew T cell maturation allowing early identification those at risk for clinical OI.

Introduction: Unrelated umbilical cord blood transplantation (UUCBT) is a viable option for those who lack HLA-matched sibling donors. However, opportunistic infections (OI) occurring in the first 100 days, remain the major cause of morbidity and mortality. Viral infections are the primary cause of OI death. As previously shown, cord blood T cells have significantly less preformed effector molecules available intracellularly to kill virally infected cells via the perforin-granzyme pathway than adult PB T cells. Since several virally infected patients control their infections in the absence of specific antiviral therapy ( e.g adenoviral enteris, polyoma cystitis) we postulated that the T cell compartment of those UUCBT recipients who experience early viral infections maty upregulate expression of the perforin exocytosis pathway. In parallel the impact of viral infection on T cell turnover would also be appreciable. Here we report on 19 prospectively studied pediatric patients, all full donor chimera, following myeloablative therapy. Methods: On day +50 we determined by 4-color FACS the expression of intracellular Granzyme, A, B, along with perforin. To monitor T cell turnover proliferating cells were identified by monitoring for the KI-67 nuclear antigen. The expression of the antiapoptosis gene BCL-2 was similarly monitored in both CD4+ and CD8+ T cells. We analyzed their association with the development of de novo OI up to day +100 employing Student’s t-test. Results: Mean age of patients was 6.2 years. 10 of 19 patients developed OI (adenovirus x 4, CMV x 7, EBVx1, parainfluenza x 1) with 5/10 patients experiencing more than one viral infections simultaneously) at a median of 29 days after UUCBT. Of those with OI 6/10 died due to their infections while 8/9 without OI are alive at a median of 15.8 months after UUCBT with one death due to leukemic relapse. Table I presents the correlation between the tested parameters with the development of OI. Patients experiencing viral infections had significantly higher % of their T cells in particular CD8+ T cells equipped with effectors of cytotoxicity and were proliferating in higher percentage compared to those with no active infections. However, the anti-apoptotic protein BCL-2 expression was significantly lower in patients experiencing OI that may lead to their shorter life span and overall T cell lymphopenia observed in OI patietns that we have previously detected in a larger cohort of 102 patients (ASBMT 2004 abstract#48). Conclusion: Correlating with active viral infections significant maturation of cord blood T cells is evident as early as 50 days after UUCBT towards acquiring effector molecules of the perforin pathway. Enhanced T cell proliferation is counteracted by reduced expression of BCL-2 that may lead to the lymphopenia in patients with OI. Future strategies aiming to enhance the longevity of antiviral T cells may protect from death due to viral infections. Univariate analysis VARIABLE MEDIAN VALUE FOR PATIENTS WITH OI MEDIAN VALUE FOR PATIENTS WITHOUT OI t-Test p value % Granzyme A+ T cells 52% 9% 0.006 % Granzyme A+ CD8+ T cells 91% 47%

HLA-mismatched unrelated umbilical cord blood transplantation (uUCBT) is a suitable option for those who lack HLA-matched sibling donors. Although UCB T cells mount significantly less cytotoxicity and cytokine response than adult T cells uUCBT can lead to significant aGVHD. We hypothesized that select features of T cell and dendritic cell (DC) reconstitution will correlate with the development of aGVHD. Here we report on 106 prospectively studied pediatric patients, all full donor chimera. Methods: On day +50 we determined by 4-color surface and ic. FACS the reconstitution of T cell subsets, lymphoid, and myeloid DC, and measured their homeostatic proliferation, activation state, and inducible cytokine secretion to correlate with the risk for developing aGVHD. Logistic regression models were created on variables with a univariate P value 1Ag and select patterns of T cell subset and DC reconstitution may identify those at risk for GVHD. UNIVARIATE AND MULTIVARIATE ANALYSIS OF DAY+50 IMMUNE PROFILE AND THE RISK FOR DEVELOPING GRADE II–IV ACUTE GVHD VARIABLE Median Values for patients with GVHD Median Values for patients without GVHD Rank Sum Test p-value Logistic regression p-value Confounders tested; age, gender, weight, CMV serology, HLA mismatch, malignancy, TBI, presence of OI, infused total cell dose/kg, CD34+ progenitor/kg, CD3+ T cell dose/kg % TNF secreting T cells 18% 2% 0.0001 0.001 %CD8+ T cells secreting TNF 5.8% 0.6% 0.008 ≤ 0.03 %CD4+ T cells secreting TNF 22% 4% 0.0002 ≤ 0.02 % IL-2 secreting T cells 17% 8% 0.0009 ≤ 0.005 %CD4+ T cells secreting IL-2 31% 14% 0.007 ≤ 0.017

Effect of Rutin on Brain Cell Membrane Function and Lipid Peroxidation During Cerebral Hypoxia in Newborn Piglets † 933