Identification of the novel protein QQS as a component of the starch metabolic network in Arabidopsis leaves.

Little is known about the role of proteins that lack primary sequence homology with any known motifs (proteins with unknown functions, PUFs); these comprise more than 10% of all proteins. This paper offers a generalized experimental strategy for identifying the functions of such proteins, particularly in relation to metabolism. Using this strategy, we have identified a novel regulatory function for Arabidopsis locus At3g30720 (which we term QQS for qua-quine starch). QQS expression, revealed through global mRNA profiling, is up-regulated in an Arabidopsis Atss3 mutant that lacks starch synthase III and has increased leaf starch content. Analysis of public microarray data using MetaOmGraph (metnetdb.org), in combination with transgenic Arabidopsis lines containing QQS promoter-GUS transgenes, indicated that QQS expression responds to a variety of developmental/genetic/environmental perturbations. In addition to the increase in the Atss3 mutant, QQS is up-regulated in the carbohydrate mutants mex1 and sis8. A 586 nt sequence for the QQS mRNA was identified by 5' and 3' RACE experiments. The QQS transcript is predicted to encode a protein of 59 amino acids, whose expression was confirmed by immunological Western blot analysis. The QQS gene is recognizable in sequenced Arabidopsis ecotypes, but is not identifiable in any other sequenced species, including the closely related Brassica napus. Transgenic RNA interference lines in which QQS expression is reduced show excess leaf starch content at the end of the illumination phase of a diurnal cycle. Taken together, the data identify QQS as a potential novel regulator of starch biosynthesis.

Ghrelin concentrations reflect body mass index rather than feeding status in obese girls.

Ghrelin stimulates both appetite and secretion of growth hormone (GH). We hypothesized that fasting should increase ghrelin, thereby increasing GH concentrations in obesity. Eight obese girls underwent a 48-h fast, receiving 25% of calories for ideal body weight. Blood was obtained every 15 min for the last 24 h of the fast. Four months later, six obese girls had blood obtained in the fed state. Two additional obese and five lean girls had blood obtained in the fed state. Ghrelin was determined in 3-h pools. Mean ghrelin concentrations were 0.41 +/- 0.03 ng/mL for lean girls and 0.16 +/- 0.01 ng/mL in obese fed girls (p < 0.0001). Lean fed girls had diurnal variation of ghrelin whereas obese fed girls did not. Fasting neither increased ghrelin (0.18 +/- 0.01 ng/mL) nor restored diurnal variation. Ghrelin concentrations were related to the body mass index (BMI) SD score (SDS) (r = 0.45, p = 0.005). For the six obese girls who participated in both fasting and fed studies, change in mean ghrelin concentration between studies was related to change in BMI SDS but not fed or fasting state. Ghrelin concentrations are, thus, a function of BMI rather than feeding status in obese girls.

Relationships among body mass index, parental perceptions, birthweight and parental weight after referral to a weight clinic.

OBJECTIVE: We sought to determine whether, in a specialty referral clinic, parental perceptions of their child's obesity were commensurate with the child's body mass index z score. Secondarily, we examined the impact of birth weight and parental body mass index on their child's body mass index z score and review results of an intake questionnaire. DESIGN: Cross-sectional study SETTING: University of Michigan from March 21, 2003 through June 30, 2004 PARTICIPANTS: Eighty-two children ages 1-20.2 years of age INTERVENTION: Body mass index z score for all participants was calculated. An intake questionnaire was completed by caregivers in which they were asked to describe their child as little overweight, overweight, very overweight or obese. OUTCOME MEASURES: Mean body mass index z score was compared to each parental descriptor. Regression analysis related body mass index z score to birthweight and parental body mass index. RESULTS: Body mass index z score was not related to parental descriptors. Maternal body mass index and child birthweight were predictors of the child's body mass index z score (r2=0.15, p<0.05; and r2=0.11, p<0.05, respectively). Both together produced a better model than either alone (r2=0.23, p<0.05). There was no relationship between paternal and child body mass index z score (p>0.05). CONCLUSIONS: There is a divergence between the parental perception of childhood obesity and the clinical definition that persists even in the context of an explicit referral. Given the significant impact of maternal weight on childhood overweight, education for prevention of overweight youth should encompass prenatal, early childhood and adolescent health maintenance.

Obesity and sex steroid changes across puberty: evidence for marked hyperandrogenemia in pre- and early pubertal obese girls.

CONTEXT: Peripubertal obesity is associated with abnormal sex steroid concentrations, but the timing of onset and degree of these abnormalities remain unclear. OBJECTIVE: The objective of the study was to assess the degree of hyperandrogenemia across puberty in obese girls and assess overnight sex steroid changes in Tanner stage 1-3 girls. DESIGN: This was a cross-sectional analysis. SETTING: The study was conducted at general clinical research centers. SUBJECTS: Thirty normal-weight (body mass index for age < 85%) and 74 obese (body mass index for age >or= 95%) peripubertal girls. INTERVENTION: Blood samples (circa 0500-0700 h) were taken while fasting. Samples from the preceding evening (circa 2300 h) were obtained in 23 Tanner 1-3 girls. MAIN OUTCOME MEASURES: Hormone concentrations stratified by Tanner stage were measured. RESULTS: Compared with normal-weight girls, mean free testosterone (T) was elevated 2- to 9-fold across puberty in obese girls, whereas fasting insulin was 3-fold elevated in obese Tanner 1-3 girls (P < 0.05). Mean LH was lower in obese Tanner 1 and 2 girls (P < 0.05) but not in more mature girls. In a subgroup of normal-weight Tanner 1-3 girls (n = 17), mean progesterone (P) and T increased overnight 2.3- and 2.4-fold, respectively (P

The association of obesity and hyperandrogenemia during the pubertal transition in girls: obesity as a potential factor in the genesis of postpubertal hyperandrogenism.

CONTEXT: Adolescent hyperandrogenemia is considered a forerunner of adult polycystic ovary syndrome, but its etiology remains uncertain. OBJECTIVE: Our objective was to explore the hypothesis that peripubertal obesity is associated with hyperandrogenemia. DESIGN AND SETTING: We performed a cross-sectional analysis of data obtained at General Clinical Research Centers. SUBJECTS: Subjects were 41 obese [body mass index (BMI) for age, >or=95%] and 35 normal-weight (BMI for age, <95%) peripubertal girls. INTERVENTION: We used pooled blood samples (approximately 0500-0700 h; n = 64) while fasting or single morning (fasting) samples (n = 12). MAIN OUTCOME MEASURES: We assessed adiposity and androgen concentrations. RESULTS: BMI correlated with total testosterone (T) (r(s) = 0.59), SHBG (r(s) = -0.69), and free T (r(s) = 0.69); free T was three times as great in obese girls compared with normal-weight girls (P < 0.0001 for all). BMI correlated with insulin (r(s) = 0.52); both insulin and LH correlated with free T (r(s) = 0.45 and 0.44, respectively; P < 0.001 for all). When analyzing early pubertal girls (pubertal stages 1-3; n = 36) alone, BMI correlated with total T (r(s) = 0.65), SHBG (r(s) = -0.74), and free T (r(s) = 0.75); free T was five times as great in obese early-pubertal girls (P < 0.001 for all). BMI correlated with insulin (r(s) = 0.65), and insulin correlated with free T (r(s) = 0.63, P < 0.01 for both). BMI correlated with free T while simultaneously adjusting for age, pubertal stage, insulin, LH, and dehydroepiandrosterone sulfate. CONCLUSION: Peripubertal obesity is associated with marked hyperandrogenemia, which is especially pronounced in early puberty.

Diurnal changes in FSH-regulatory peptides and their relationship to gonadotrophins in pubertal girls.

BACKGROUND: FSH-regulatory peptides participate with GnRH and sex steroids to regulate serum FSH concentrations. We hypothesized that day/night variations in FSH serum concentrations would be associated with diurnal variation in FSH-regulatory peptides. METHODS: Blood was obtained every 15 min for 24 h beginning at 08:00 h in eight girls [seven with variations in growth or puberty and one with idiopathic hypogonadotrophic hypogonadism (IHH)] and for 12 h beginning at 20:00 h in 12 additional girls with variant puberty, eight with gonadal dysgenesis or ovarian failure (GD/OF) and one with IHH. Samples across 3 h blocks were pooled for determination of LH, FSH, activin-A, inhibin-B and follistatin 288. RESULTS: LH and FSH concentrations increased from 23:00 to 08:00 h with respect to daytime concentrations in pubertal girls (P<0.005) but only LH increased (P=0.002) in girls with GD/OF. In pubertal girls, inhibin-B declined during the day (P=0.019), reaching a nadir between 17:00 and 22:45 h just prior to the night-time increase in FSH. Follistatin concentrations exhibited diurnal variation (P=0.028), with the greatest concentrations occurring between 05:00 and 11:00 h. Activin-A concentrations declined coincident with the night-time increase in FSH in pubertal girls (P<0.0001) but not in girls with GD/OF. CONCLUSIONS: The directionality of changes in FSH-regulatory proteins supports the notion that FSH-regulatory peptides may contribute to the night-time augmentation of circulating FSH during puberty in girls.

GnRH agonist stimulation of the pituitary-gonadal axis in children: age and sex differences in circulating inhibin-B and activin-A.

BACKGROUND: Inhibin-B decreases and activin increases FSH secretion in adults. We investigated whether an FSH-inhibin/activin feedback loop exists before or during puberty. METHODS: FSH secretion was stimulated with 10 microg/kg leuprolide acetate (GnRH agonist) in 18 girls, ages 1.0-13.2 years, and 11 boys, ages 8.9-15.2 years, with variations in pubertal development, and in five normal 9- to 10-year-old girls. Blood, obtained at 0, 0.5, 1, 2, 4, 8, 12, 16, 20 and 24 h after GnRH agonist, was analysed for LH, FSH, activin-A, inhibin-A, inhibin-B, follistatin 288 and estradiol/testosterone. RESULTS: FSH increased within 30 min of GnRH agonist administration with a peak greater in girls than boys (P=0.0006). Baseline inhibin-B was greater in boys than girls (P=0.01), while baseline activin-A concentrations were greater in girls. GnRH agonist-stimulated FSH increased inhibin-B in girls by 8 h and in boys by 20 h (P<0.05), but did not affect activin-A. Inhibin-B increases were seen only in girls older than 5 years. CONCLUSIONS: An inhibin-B-FSH feedback loop exists prior to the onset of puberty in girls older than 5 years. Sex differences in activin-A and inhibin-B concentrations may be responsible for sex differences in serum FSH concentrations.

Augmentation of growth hormone secretion after testosterone treatment in boys with constitutional delay of growth and adolescence: evidence against an increase in hypothalamic secretion of growth hormone-releasing hormone.

The increase in pituitary GH secretion that occurs during mid-late puberty in boys follows an increase in circulating testosterone (T) concentration; the direct mechanism by which this occurs is unknown. We hypothesized that T increases GH secretion during puberty by augmenting hypothalamic output of GHRH. Using constant infusions of a GHRH antagonist, we tested this hypothesis in six early pubertal boys with constitutional delay of growth and adolescence who had a mean chronological age of 14.0 +/- 0.3 yr and mean bone age of 11.4 +/- 0.2 yr. Blood samples were obtained from subjects every 15 min for 24 h during the overnight infusion of normal saline (2000-0600 h) and again during the overnight infusion of GHRH antagonist (0.33 microg/kg/h) the following night. Subjects then received transdermal T (5-mg patch) for 12 h nightly and were studied again after 4 wk of treatment. Serum samples were assayed for GH and total ghrelin; the percent suppression of GH during GHRH antagonist infusion was calculated. Morning serum T rose from 0.44 +/- 0.09 to 4.43 +/- 0.74 microg/liter (P = 0.005). T treatment was associated with a 92.6% increase in mean nocturnal GH secretion area under the curve (830 +/- 177 to 1599 +/- 340 microg/24 h.liter). Infusion of GHRH-antagonist suppressed mean nocturnal GH area under the curve by 29.1% before T treatment (830 +/- 177 to 621 +/- 168 microg/24 h.liter), and by 29.4% after T treatment (1599 +/- 340 to 1182 +/- 249 microg/24 h.liter; P = 0.99). Somatotroph sensitivity to GHRH was tested with 0.1- and 1.0-microg/kg doses of GHRH-44 iv; GH response did not change with regard to T treatment. The mean 24-h concentration of total ghrelin was unchanged with regard to T treatment. In summary, nightly transdermal T administration in six boys with constitutional delay of growth and adolescence increased GH output almost 2-fold, whereas the degree of GH suppressibility by GHRH antagonist remained unchanged. We conclude that the T-associated augmentation of GH secretion during early puberty in boys is unlikely to involve an absolute increase in hypothalamic GHRH output.

Sex differences in FSH-regulatory peptides in pubertal age boys and girls and effects of sex steroid treatment.

BACKGROUND: FSH concentrations are higher in girls than in boys before puberty. We hypothesized that steroid-mediated changes in FSH-regulatory proteins underlie the sex differences in FSH secretion and pubertal timing. METHODS: FSH-regulatory proteins, LH, FSH and sex steroids were measured in five boys, 10 girls, and five girls with Turner syndrome before and during sex steroid treatment (girls, 0.05 mg/day estradiol; boys, 5 mg/day testosterone) for up to 4 weeks. Blood was obtained every 15 min from 20.00 to 08.00 h before and during sex steroid treatment. RESULTS: The mean FSH concentration was higher in girls than in boys (P = 0.0044). Activin-A concentrations were greater (P < 0.0001) and inhibin-B concentrations lower (P < 0.0001) in girls compared with boys. Steroid treatment (i) suppressed LH/FSH concentrations in all subjects; (ii) increased the mean activin-A concentration in all but the Turner girls (P = 0.001); and (iii) decreased inhibin-B concentrations in boys (P = 0.005) but not in girls. Total follistatin and follistatin 288 concentrations did not differ by sex. CONCLUSIONS: Sex steroids regulate circulating activin-A and inhibin-B concentrations in children. The lower inhibin-B and higher activin-A concentrations may explain the higher FSH and earlier onset of puberty in girls.

MetNet: Software to Build and Model the Biogenetic Lattice of Arabidopsis.

MetNet (http://www.botany.iastate.edu/ approximately mash/metnetex/metabolicnetex.html) is publicly available software in development for analysis of genome-wide RNA, protein and metabolite profiling data. The software is designed to enable the biologist to visualize, statistically analyse and model a metabolic and regulatory network map of Arabidopsis, combined with gene expression profiling data. It contains a JAVA interface to an interactions database (MetNetDB) containing information on regulatory and metabolic interactions derived from a combination of web databases (TAIR, KEGG, BRENDA) and input from biologists in their area of expertise. FCModeler captures input from MetNetDB in a graphical form. Sub-networks can be identified and interpreted using simple fuzzy cognitive maps. FCModeler is intended to develop and evaluate hypotheses, and provide a modelling framework for assessing the large amounts of data captured by high-throughput gene expression experiments. FCModeler and MetNetDB are currently being extended to three-dimensional virtual reality display. The MetNet map, together with gene expression data, can be viewed using multivariate graphics tools in GGobi linked with the data analytic tools in R. Users can highlight different parts of the metabolic network and see the relevant expression data highlighted in other data plots. Multi-dimensional expression data can be rotated through different dimensions. Statistical analysis can be computed alongside the visual. MetNet is designed to provide a framework for the formulation of testable hypotheses regarding the function of specific genes, and in the long term provide the basis for identification of metabolic and regulatory networks that control plant composition and development.

Incomplete modified fast in obese early pubertal girls leads to an increase in 24-hour growth hormone concentration and a lessening of the circadian pattern in leptin.

We studied nutrition and GH in eight obese girls, aged 6-11 yr. Blood was sampled every 15 min for 24 h. A 48-h diet providing 25% of assumed caloric needs was imposed, with repeat sampling during the last 24 h. Six nonfasting lean girls were also studied, and their mean GH was 3 times that of the obese girls in the fed state (P = 0.024). Dieting increased mean GH by 60% (P = 0.0028). There was no difference in pulse number for either group, but total secretion for lean girls was 3.9 times greater than that in obese girls during the fed state. With dieting, obese girls increased their total GH secretion by 60% (P = 0.010), but maintained lower total secretion, approximately 40% that of lean girls (P = 0.014). Mean leptin in obese girls in the fed state was 6.2 times greater than mean leptin in lean girls (P = 0.0001), with higher concentrations at night (P < 0.05) and lowering of total mean leptin while dieting. We conclude that in early pubertal obese girls, short-term caloric restriction partially reverses the low GH state that is characteristic of obesity. The change is concomitant with a decrease in leptin and a lessening of circadian differences.

Social factors associated with prolonged hospitalization among diabetic children.

OBJECTIVE: To determine social factors associated with increased risk of hospital admission from diabetic ketoacidosis (DKA) or diabetic coma as well as risk of prolonged hospital stay. METHODS: A cohort of all children (/=7 days. RESULTS: A total of 8443 children with a primary hospital diagnosis of DKA and 123 children with type 1 DM and coma were identified; 55% of the children were girls, 32% were nonwhite, 29% received Medicaid insurance, and 33% resided in areas of poverty. Children with prolonged hospital stay were significantly more likely to be of nonwhite race (odds ratio [OR]: 2.0; 95% confidence interval [CI]: 1.6-2.5), to receive Medicaid insurance (OR: 1.4; 95% CI: 1.1-1.7), to live in areas of poverty (OR: 1.3; 95% CI: 1.1-1.7), and to be of younger age. CONCLUSIONS: When compared with state census data, nonwhite and poor children were more likely to be admitted with complications of DM and to have significantly prolonged and expensive hospital stays. These children should be targeted for intensive diabetes education and outpatient medical support both to improve their health and potentially to decrease total health care costs.