The function and affinity maturation of HIV-1 gp120-specific monoclonal antibodies derived from colostral B cells.

Despite the risk of transmitting HIV-1, mothers in resource-poor areas are encouraged to breastfeed their infants because of beneficial immunologic and nutritional factors in milk. Interestingly, in the absence of antiretroviral prophylaxis, the overwhelming majority of HIV-1-exposed, breastfeeding infants are naturally protected from infection. To understand the role of HIV-1 envelope (Env)-specific antibodies in breast milk in natural protection against infant virus transmission, we produced 19 HIV-1 Env-specific monoclonal antibodies (mAbs) isolated from colostrum B cells of HIV-1-infected mothers and investigated their specificity, evolution, and anti-HIV-1 functions. Despite the previously reported genetic compartmentalization and gp120-specific bias of colostrum HIV Env-specific B cells, the colostrum Env-specific mAbs described here demonstrated a broad range of gp120 epitope specificities and functions, including inhibition of epithelial cell binding and dendritic cell-mediated virus transfer, neutralization, and antibody-dependent cellular cytotoxicity. We also identified divergent patterns of colostrum Env-specific B-cell lineage evolution with respect to crossreactivity to gastrointestinal commensal bacteria, indicating that commensal bacterial antigens play a role in shaping the local breast milk immunoglobulin G (IgG) repertoire. Maternal vaccine strategies to specifically target this breast milk B-cell population may be necessary to achieve safe breastfeeding for all HIV-1-exposed infants.

Are field measures of adiposity sufficient to establish fatness-related linkages with metabolic outcomes in adolescents?

OBJECTIVE: To examine the associations between the adiposity-related information conveyed by field fatness measures: body mass index (BMI), waist circumference (WC) and sum of triceps and subscapular skinfolds (SUM SF) relative to dual-energy X-ray absorptiometry (DXA), beyond their common intercorrelations, with three important metabolic variables in US adolescents. METHODS: We analyzed data on adiposity and insulin resistance (HOMA-IR), serum triglycerides (TGs) and total cholesterol (TC) from three US national surveys. In two-stage least-square modeling, we first calculated the common adiposity variance, and then used multivariate linear and quantile regressions to access residual associations with each measure. RESULTS: Basic associations for each of the adiposity measures were similar but differences emerged in residual adiposity analyses scaled by s.d. units. While a 1 s.d. change in residual variance in DXA total fat beyond that accounted for by BMI (DXA|BMI) was strongly and significantly associated with all outcomes, associations with DXA accounting for SUM SF (DXA|SUM SF) and WC (DXA|WC) were weak or nonsignificant. Contrasted amongst themselves, the residual score association between BMI|SUM SF (ß=0.06, P<0.0001) and HOMA-IR was weaker, and half as strong as that for the converse, SUM SF|BMI (ß=0.13, P=0.020). SUM SF|WC was stronger than WC|SUM SF (ß=0.08, P<0.0001 vs SUM SF|WC ß=0.13, P<0.0001). Associations were similar for TGs and TC. CONCLUSIONS: Laboratory methods like DXA offer minimal explanatory advantage over field methods in assessing adiposity-related contributions to metabolic outcomes in adolescents. Among the simple fatness measures, skinfolds convey additional information beyond BMI and WC when estimating associations both at the population mean and at the upper extremes of metabolic factors.

Comparability of skinfold thickness to DXA whole-body total fat in their associations with serum triglycerides in youths.

OBJECTIVE: To determine the comparability of triceps and subscapular skinfold thicknesses with dual X-ray absorptiometry (DXA) whole-body total fat (kg) in relation to serum triglyceride (TG) levels and increased risk of elevated TG levels, and identified optimum skinfold cutoffs for screening purposes in US adolescents. SUBJECTS/METHODS: Data from triceps and subscapular skinfold thickness, DXA whole-body total fat and serum TGs were obtained from 1505 US adolescents ages 12.00-17.99 years, who participated in two continuous National Health and Nutrition Examination Survey (NHANES) cycles 2001-2004. Study associations were examined with linear and logistic models, and ROC (receiver operating characteristic) analyses were used to derive skinfold cutoffs for identifying the risk of elevated TG levels. RESULTS: Using area under the curves (AUCs) as metrics of prediction accuracy (with bootstrapped 95% CIs), no significant differences were found between skinfolds and DXA logistic models for predicting elevated TG levels. Similarly, skinfold and DXA models had comparable precision in predicting continuous serum TG from bootstrapped root mean squared errors for both sexes. Population-adjusted marginal mean estimates indicated that youths whose skinfolds are in the top quartile had TG levels within 83-108 mg/dl. Skinfold cutoffs for predicting elevated estimated TG using ROC analyses showed that cutoffs decreased with age and ranged from 13 to 30 mm for ages 12-17, in yearly intervals. CONCLUSION: Skinfold thicknesses were comparable to DXA whole-body total fat in predicting serum TG levels. These skinfold cutoffs could be used in practical settings as a first pass screener for identifying US adolescents at risk of elevated serum TGs.

A two-part model for reference curve estimation subject to a limit of detection.

Reference curves are commonly used to identify individuals with extreme values of clinically relevant variables or stages of progression which depend naturally on age or maturation. Estimation of reference curves can be complicated by a technical limit of detection (LOD) that censors the measurement from the left, as is the case in our study of reproductive hormone levels in boys around the time of the onset of puberty. We discuss issues with common approaches to the LOD problem in the context of our pubertal hormone study, and propose a two-part model that addresses these issues. One part of the proposed model specifies the probability of a measurement exceeding the LOD as a function of age. The other part of the model specifies the conditional distribution of a measurement given that it exceeds the LOD, again as a function of age. Information from the two parts can be combined to estimate the identifiable portion (i.e. above the LOD) of a reference curve and to calculate the relative standing of a given measurement above the LOD. Unlike some common approaches to LOD problems, the two-part model is free of untestable assumptions involving unobservable quantities, flexible for modeling the observable data, and easy to implement with existing software. The method is illustrated with hormone data from the Third National Health and Nutrition Examination Survey.

Otitis media and associations with overweight status in toddlers.

INTRODUCTION: Otitis media (OM) is a significant disease that affects nearly all children early in life. Recently, childhood overweight has become an epidemic. Past research has demonstrated that a history of OM is related to food preferences and overweight through proposed physiological mechanisms. The purpose of this study was to explore the relationship between recurrent OM (ROM)/tympanostomy tube treatment and overweight status. METHODS: Data were analyzed from a prospective cohort of mothers and children recruited from 1991-1996 from a local health maintenance organization. ROM and tympanostomy tube status were obtained through a combination of physical exam and medical record abstraction. ROM and tympanostomy tube status were analyzed as categorical variables with weight-for-length (WFL) data from well child checks. Chi-square and logistic regression for univariate and multivariate analyses were performed. RESULTS: 11.4% of children had a WFL measure at two years of age ≥ 95 th percentile. Those children with a history of tympanostomy tube treatment had a significantly increased risk of having a WFL ≥ 95 th percentile after controlling for birth weight, maternal prenatal smoking, maternal education, and family income (OR 3.32, 95% CI 1.43-7.72). The alternative hypothesis that children with larger WFL at two month of age would have a greater number of OM episodes by two years of age was not significant. CONCLUSION: The findings of this study are consistent with the hypothesis and prior research that OM treated with tympanostomy tubes is associated with overweight status.

Association of inflammation with worsening HOMA-insulin resistance.

AIMS/HYPOTHESIS: We examined the cross-sectional and longitudinal relationships between C-reactive protein (CRP), a marker of low-grade inflammation, and insulin resistance and whether the association was independent of obesity and oxidative stress. METHODS: CRP and insulin resistance (homeostasis model assessment of insulin resistance [HOMA-IR]) data were obtained in a population-based, prospective observational study, Coronary Artery Risk Development in Young Adults (CARDIA), during 1992-2006. RESULTS: CRP showed a significant positive association with insulin resistance, both cross-sectionally and longitudinally (5 year follow-up). The estimated increment in HOMA-IR was 0.34 log(e)(pmol/l x [mmol/l]/156.25) (p value for trend <0.0001) in the highest vs lowest CRP quartiles in cross-sectional analysis, whereas the corresponding estimate was 0.12 (p trend <0.0001) in the highest vs lowest CRP quartiles longitudinally over 5 years. The gradient of HOMA-IR across CRP was attenuated but remained statistically significant after controlling for body fat measurements (0.06 in the highest vs lowest CRP in both cross-sectional [p value for trend = 0.001] and longitudinal analyses [p value for trend = 0.01]), and was little changed by further adjustment for oxidative stress markers (F(2)-isoprostanes and oxidised LDL). There were consistent increments in the levels of HOMA-IR with increasing concentrations of CRP over time. In contrast, higher HOMA-IR did not predict future increases in CRP. Findings were similar using fibrinogen as the predictor variable. CONCLUSIONS/INTERPRETATION: Although a substantial portion of this association was explained by obesity, CRP was independently related to concurrent and future insulin resistance.

Growth and endocrine function in patients with Hurler syndrome after hematopoietic stem cell transplantation.

Short stature is characteristic of Hurler syndrome, or mucopolysaccharidosis type IH (MPS IH). Hematopoietic stem cell transplantation (HSCT) is used to treat children with MPS IH. While HSCT corrects some of the metabolic features of MPS IH, its effects on growth are not well delineated. We investigated growth in patients with MPS IH after HSCT and described accompanying endocrine abnormalities. A cohort of 48 patients with MPS IH who had received HSCT between 1983 and 2005 were included. The prevalence of short stature (height <-2 s.d. score, SDS) before HSCT was 9%, and increased to 71% at last follow-up (6.9+/-5.1 years after HSCT). Short stature was positively associated with increased age at HSCT (P=0.002) and TBI (P=0.009). In total, 23% had growth hormone deficiency and/or low insulin-like growth factor-1, one female patient had premature adrenarche, one precocious puberty and 27% had clinical or subclinical hypothyroidism. Growth failure is highly prevalent in children with MPS IH after HSCT. Children who had no TBI exposure and were younger at the time of HSCT had a better height outcome.

Diurnal cycles of salivary cortisol in older adults.

Cortisol has a well-documented circadian pattern. However, recent studies have demonstrated that individual variation in diurnal cortisol patterns occurs in young adult populations. Since older adults experience altered sleep-wake cycles and changes in circadian rhythmicity, we may see even greater variations in diurnal cortisol patterns in older adults. This study examined salivary cortisol patterns in 48 community dwelling older adults. Participants (mean age 76+/-6) collected saliva every 2 h over a three-day period. Cortisol was assayed by using RIA. Cortisol cycles were defined as inconsistent, typical or flat based on the slopes of two sequential daily cortisol patterns. Demographic, physical, psychological and behavioral measures were tested for group differences using t-tests and chi-square analyses. Forty-eight percent of the sample had inconsistent cycles, 50% had typical cycles and 2% had flat cycles. This sample had a higher percentage of inconsistent cycles and fewer flat cycles than reported for young adults (p=0.008) (Psychoneuroendocrinology 22 (1997) 89). Those with inconsistent cycles were younger and reported higher caffeine and food intake than those with typical cycles. This study demonstrates that normal diurnal rhythms of cortisol can be maintained in older adults, while day-to-day variation may increase.

A randomized trial of a brief intervention to increase fruit and vegetable intake: a replication study among callers to the CIS.

BACKGROUND: Results are reported from a large randomized trial designed to increase fruit and vegetable consumption among callers to the National Cancer Institute's Cancer Information Service (CIS) (n = 1,717). METHODS: CIS callers assigned to the intervention group (n = 861) received a brief proactive educational intervention over the telephone at the end of usual service, with two follow-up mailouts. Key educational messages and print material derived from the NCI 5 A Day for Better Health program were provided to intervention participants. Participants were interviewed by telephone at 4 weeks (n = 1,307), 4 months (n = 1,180), and 12 months for follow-up (n = 1,016). RESULTS: Results obtained from a single-item measure of fruit and vegetable consumption indicate a significant intervention effect of 0.88 servings per day at 4 weeks follow-up (P < 0.001), 0.63 servings per day at 4 months follow-up (P < 0.001), and 0.43 servings per day at 12 months follow-up (P < 0.001). Using a 7-item food frequency measure, an intervention effect of 0.63 servings per day was obtained at 4 weeks follow-up (P < 0.001), compared with 0.39 servings per day at 4 months follow-up (P = 0.002) and 0.44 servings per day at 12 months follow-up (P = 0.002). A 24-h recall assessment included in the 4-month interviews also yielded a significant intervention effect of 0.67 servings per day (P = 0.015). The vast majority of callers (90%) endorsed the strategy of providing 5 A Day information proactively within the CIS. CONCLUSIONS: This brief educational intervention was associated with higher levels of self-reported fruit and vegetable intake at both short- and long-term follow-up. Additional research is recommended to test this or a similar intervention in diverse populations.

C/EBP epsilon mediates myeloid differentiation and is regulated by the CCAAT displacement protein (CDP/cut).

Neutrophils from CCAAT enhancer binding protein epsilon (C/EBP epsilon) knockout mice have morphological and biochemical features similar to those observed in patients with an extremely rare congenital disorder called neutrophil-specific secondary granule deficiency (SGD). SGD is characterized by frequent bacterial infections attributed, in part, to the lack of neutrophil secondary granule proteins (SGP). A mutation that results in loss of functional C/EBP epsilon activity has recently been described in an SGD patient, and has been postulated to be the cause of the disease in this patient. We have previously demonstrated that overexpression of CCAAT displacement protein (CDP/cut), a highly conserved transcriptional repressor of developmentally regulated genes, suppresses expression of SGP genes in 32Dcl3 cells. This phenotype resembles that observed in both C/EBP epsilon(-/-) mice and in SGD patients. Based on these observations we investigated potential interactions between C/EBP epsilon and CDP/cut during neutrophil maturation. In this study, we demonstrate that inducible expression of C/EBP epsilon in 32Dcl3/tet cells results in granulocytic differentiation. Furthermore, Northern blot analysis of G-CSF-induced CDP/cut overexpressing 32Dcl3 cells revealed absence of C/EBP epsilon mRNA. We therefore hypothesize that C/EBP epsilon positively regulates SGP gene expression, and that C/EBP epsilon is itself negatively regulated by CDP/cut during neutrophil maturation. We further demonstrate that the C/EBP epsilon promoter is regulated by CDP/cut during myeloid differentiation.

Reliability and validity of parental measurements of infant size.

This study evaluates the reliability and validity of parental measurements of infant size, using illustrated instructions and simple measuring tools. Following pilot tests, final methods were evaluated on a sample of 28 parents (26 mothers and 2 fathers) of infants from 1 to 6 weeks of age. Parents independently measured twice the infant's head circumference (HC), mid-upper arm circumference (MUAC), abdominal circumference (AC), and recumbent length (RL). Infants were also measured twice by a trained observer. Mean parental measures were correlated (intraclass R) with the observer criterion measures at the levels of 0.81 for RL, 0.70 for AC, 0.80 for MUAC, and 0.94 for HC. Relative differences in the means for measurements obtained by parents and the trained observer were small, ranging from 0.3% for HC and RL, to 3.8% for AC. Intraclass correlations of reliability between the two parental measurements ranged from R = 0.84 for MUAC to 0.96 for RL. Given the reliability and validity of the results, the methods tested yield measurements that are suitable for ranking individuals and for use in group-level analyses, and at least in the case of head circumference, for individual-level analyses.

Validity and reliability of self-reported stature and weight of US adolescents.

Validity and reliability of self-reported stature and weight were investigated for U.S. adolescents (12.0-17.0 years) who were participants in the Third National Health and Nutrition Examination Survey (NHANES III). Data were collected on 1,635 youth and were statistically weighted to represent the national population. Self-reported weights are missing from 40% of 12 year olds and 25% of 13 year olds. Those who refused or were unable to provide self-reported weights were younger, shorter, and lighter than those who did. Among those who provided self-reports, the average bias and random error in reporting were largest for the youngest youth. Biases in reporting stature and weight were consistently negative following the NHANES III protocol. The intraclass coefficients between measured and self-reported dimensions within age and gender groups ranged from 0.57 to 0.91 and from 0.85 to 0.98, for stature and weight, respectively. Self-reported stature and weight are not recommended as proxies for measured dimensions for youth less than 14 years of age.

The role of C/EBP(epsilon) in the terminal stages of granulocyte differentiation.

As a consequence of its characterization using both in vitro and knockout mouse models, the myeloid-specific transcription factor, CCAAT/enhancer binding protein (C/EBP)epsilon, has been identified as a critical regulator of terminal granulopoiesis and one of the causative mutations in the human disease, neutrophil-specific granule deficiency. C/EBPs are a family of transcription factors sharing numerous structural and functional features and to date include C/EBPalpha, -beta, -gamma, -delta, -epsilon, and -zeta. C/EBPalpha was the first family member isolated and characterized, its essential role in hepatocyte and adipocyte differentiation demonstrated in knockout mouse models. Subsequent analysis of the hematopoietic elements in fetal mouse liver revealed its critical role in myelopoiesis. Understanding the role of C/EBPepsilon in terminal granulopoiesis in the context of other known transcription factors is ongoing with analysis of deficient and conditionally expressing cell lines and knockout models. Mouse models with targeted gene disruptions have contributed greatly to our understanding of the transcriptional regulation of granulopoiesis. Further manipulation of these models and other conditional expression systems have bypassed some of the limitations of knockout models and helped delineate the interactions of different transcription factors in affecting granulocyte development. Phenotypic expression of the loss of C/EBPepsilon in mice is extreme, resembling absolute neutropenia with systemic infection with P. aeruginosa. Future work will need to explore the regulation of C/EBPepsilon expression, its functional interactions with other transcriptional regulators such as PU.1, and its role in monocyte differentiation and function in the mouse.

C/EBPepsilon -/- mice: increased rate of myeloid proliferation and apoptosis.

CCAAT/enhancer binding protein epsilon (C/EBPepsilon) is essential for terminal granulocytic differentiation. Its expression begins at the transition between the proliferative and non-proliferative compartments of myelopoiesis. We studied the effect of targeted disruption of the C/EBPepsilon gene on murine myeloid proliferation and apoptosis. Bone marrow cellularity of C/EBPepsilon -/- and wild-type mice was 95% and 65%, respectively. The C/EBPepsilon -/- mice had an expansion in the number of their CFU-GM/femur. The number of myeloid committed progenitor cells in the peripheral blood and the spleen of these mice was also increased. Bromodeoxyuridine (BrDU) pulse labeling studies demonstrated that the fraction of actively proliferating cells was two-fold higher in the bone marrow of C/EBPepsilon -/- mice. However, the number of myeloid colonies arising from purified Sca-1+/lin- early hematopoietic progenitor cells and from bone marrow mononuclear cells grown in different cytokine combinations was not significantly different between wild-type and knock-out mice. Also, long-term marrow growth, and CFU were not different between the wild-type and C/EBPepsilon -/- mice. The sensitivity to induction of apoptosis in the committed progenitor cell compartment after either withdrawal of growth factor or brief exposure to etoposide was normal. However, Gr-1 antigen-positive C/EBPepsilon -/- granulocytic cells showed an increased rate of apoptosis in comparison to their wild-type counterparts. In summary, the myeloid compartment appears to be expanded in mice lacking C/EBPepsilon. However, this is not the consequence of an intrinsic myeloproliferation but due to an indirect, possibly cytokine-mediated stimulation of myelopoiesis in vivo. C/EBPepsilon may have a role in the inhibition of apoptosis in maturing granulocytic cells.

Molecular analysis of the mouse S100A9 gene and evidence that the myeloid specific transcription factor C/EBPepsilon is not required for the regulation of the S100A9/A8 gene expression in neutrophils.

The genomic locus of the mouse S100A9 (MRP14) gene, a myeloid expressed gene belonging to the S100 family, is split in three exons and two introns. Insertions of B1 like and LINE elements as well as several sequence repeat structures are scattered over the gene suggesting that this region of the S100 gene cluster has been the subject of a high mutational activity in mouse evolution. The insertions may represent molecular footprints of a recently postulated inversion event, which resulted in a rearrangement of the S100 gene cluster in mouse compared to man. Deletion analysis of the promoter reveals, that a 1200 bp fragment is able to direct a cell type-specific expression of a reporter gene in granulocytic 32D cells. Unexpectedly, the myeloid-specific transcription factor C/EBPepsilon is not needed for the transcriptional upregulation of the S100A9 and S100A8 genes in neutrophils. The data described here provide further insights into the evolution of the S100 gene cluster and into the myeloid-specific regulation of the murine S100A9 gene expression.

Associations of periodontal disease with femoral bone mineral density and estrogen replacement therapy: cross-sectional evaluation of US adults from NHANES III.

The objectives of this study were to evaluate the possible association of periodontal disease with (1) femoral bone mineral density (BMD), and (2) estrogen replacement therapy in a large sample of US adults (N= 11,655). The mean clinical attachment loss (CAL) per person was the main outcome variable. Based on the total BMD of the proximal femur and using the WHO diagnostic criteria, subjects were classified as having osteoporosis, osteopenia, or normal BMD. After adjusting for confounders, females with high calculus scores and low BMD had significantly more CAL than females with normal BMD and similar calculus scores (p<0.0001). No association was observed among women with low and intermediate levels of calculus. The greater CAL present among women with low BMD was associated with gingival recession. Patterns of findings were similar but equivocal among men, of whom only 66 were osteoporotic. After adjustment for possible confounders, postmenopausal women who reported having used estrogen replacement therapy presented significantly less mean CAL than those who never used estrogen. These findings indicate that in the presence of high calculus scores, females with osteoporosis are at increased risk for attachment loss and that this risk may be attenuated by the use of estrogen replacement therapy.

Differences and trends in antioxidant dietary intake in smokers and non-smokers, 1980-1992: the Minnesota Heart Survey.

PURPOSE: Differences and secular trends in dietary antioxidant vitamin intake (vitamins E, C, and beta-carotene) in current non-smokers, light smokers, and heavy smokers were examined as part of the Minnesota Heart Survey. METHODS: Three cross-sectional surveys were conducted in adults ages 25-74 years in 1980-82 (N = 1682), 1985-87 (N = 2326), and 1990-92 (N = 2487). Dietary information was obtained from a 24-hour dietary recall. Smoking was assessed through self-report. Intakes were adjusted for age, energy intake, body mass index, education level, and exercise level (vitamins E, C and beta-carotene). RESULTS: Antioxidant vitamin intakes were significantly higher in non-smokers than in light (1-20 cig/day) and heavy smokers (>20 cig/day) when all three survey periods were combined. In men, mean vitamin E intake was 9.2 mg, 8.6 mg, and 7.8 mg for non-smokers, light smokers, and heavy smokers, respectively. Results were similar in men for beta-carotene (non-smokers 1408 microg, light smokers 1287 microg, and heavy smokers 1064 microg), and vitamin C (non-smokers 81 mg, light smokers 67 mg, and heavy smokers 56 mg). Women had results of similar magnitude and direction. From 1980-92, secular trends in men showed non-significant increases from 1980-82 to 1990-92 in beta-carotene (+6.1%), while decreases were observed in vitamins E (-1.1%) and C (-2.6%). In contrast, women had large decreases in all antioxidant vitamin intakes: vitamin E (-13%), vitamin C (-18.6%), and beta-carotene (-16.2%). CONCLUSIONS: Light and heavy smokers had a significantly lower overall mean dietary antioxidant vitamin intake than non-smokers. Over the decade, antioxidant dietary intake remained relatively stable in men and decreased in women in Minneapolis-St. Paul, despite improvements in access to antioxidant rich fruits and vegetables.