The reliability of the disabled children's quality-of-life questionnaire in Swedish children with diabetes.

AIM: To determine the reliability of the disabled children's quality-of-life measure (DISABKIDS) chronic generic questionnaire and diabetes module in children. The questionnaire is being evaluated for repeated routine health-related quality-of-life (HrQoL) assessment and in association with the Swedish national paediatric diabetes registry (Swediabkids), which is a tool for regular clinical use. METHODS: Children and parents completed the questionnaire during a routine visit to the diabetes clinic. In total, 120 families completed the test and retest. Split-half reliability correlation and intraclass correlation (ICC) coefficients were calculated. Bland & Altman plots were calculated on the generic HRQoL domain. RESULTS: Both child and parent versions showed good internal consistency. Test-retest ICC coefficients for the generic HrQoL module were 0.913 for the children and 0.820 for the parent version. All generic domains independently showed good reliability. The diabetes module had a score of 0.855 for children and 0.823 for parents. Split-half correlation for generic and diabetes modules was 0.930 and 0.848 for children, 0.953 and 0.903 for parents. Bland and Altman plots showed substantial agreement between the two administrations for both children and parents. CONCLUSION: The DISABKIDS questionnaire is a reliable instrument for the repeated measurements of HrQoL in children with diabetes.

Extended evaluation of the safety and efficacy of GAD treatment of children and adolescents with recent-onset type 1 diabetes: a randomised controlled trial.

AIMS/HYPOTHESIS: The aim of this study was to investigate the safety and efficacy of alum formulated glutamic acid decarboxylase GAD(65) (GAD-alum) treatment of children and adolescents with type 1 diabetes after 4 years of follow-up. METHODS: Seventy children and adolescents aged 10-18 years with recent onset type 1 diabetes participated in a phase II, double-blind, randomised placebo-controlled clinical trial. Patients identified as possible participants attended one of eight clinics in Sweden to receive information about the study and for an eligibility check, including a medical history. Participants were randomised to one of the two treatment groups and received either a subcutaneous injection of 20 µg of GAD-alum or placebo at baseline and 1 month later. The study was blinded to participants and investigators until month 30. The study was unblinded at 15 months to the sponsor and statistician in order to evaluate the data. At follow-up after 30 months there was a significant preservation of residual insulin secretion, as measured by C-peptide, in the group receiving GAD-alum compared with placebo. This was particularly evident in patients with <6 months disease duration at baseline. There were no treatment-related serious adverse events. We have now followed these patients for 4 years. Overall, 59 patients, 29 who had been treated with GAD-alum and 30 who had received placebo, gave their informed consent. RESULTS: One patient in each treatment group experienced an episode of keto-acidosis between months 30 and 48. There were no treatment-related adverse events. The primary efficacy endpoint was the change in fasting C-peptide concentration from baseline to 15 months after the prime injection for all participants per protocol set. In the GAD-alum group fasting C-peptide was 0.332 ± 0.032 nmol/l at day 1 and 0.215 ± 0.031 nmol/l at month 15. The corresponding figures for the placebo group were 0.354 ± 0.039 and 0.184 ± 0.033 nmol/l, respectively. The decline in fasting C-peptide levels between day 1 and month 1, was smaller in the GAD-alum group than the placebo group. The difference between the treatment groups was not statistically significant. In those patients who were treated within 6 months of diabetes diagnosis, fasting C-peptide had decreased significantly less in the GAD-alum group than in the placebo-treated group after 4 years. CONCLUSION/INTERPRETATION: Four years after treatment with GAD-alum, children and adolescents with recent-onset type 1 diabetes continue to show no adverse events and possibly to show clinically relevant preservation of C-peptide. TRIAL REGISTRATION: ClinicalTrials.gov NCT00435981 FUNDING: The study was funded by The Swedish Research Council K2008-55X-20652-01-3, Barndiabetesfonden (The Swedish Child Diabetes Foundation), the Research Council of Southeast Sweden, and an unrestricted grant from Diamyd Medical AB.

Chiasma and recombination data in plants: are they compatible?

It has long been considered that the number of chiasmata formed during meiosis corresponds to the number of crossovers indicated by the genetic map. However, recent investigations in plants show an unexpected discrepancy in the results obtained when calculating the total number of crossover events per meiosis by these two methods. Is this discrepancy due to methodological difficulties? Or is there something fundamentally wrong with our understanding of crossovers and chiasmata?

Effects of insulin on lipolysis and lipogenesis in hamster white adipocytes with high sensitivity to hormones.

A modified procedure for preparation of hamster adipocytes by collagenase digestion under carefully controlled conditions has been developed. The adipocytes were 4- to 8-fold more sensitive to catecholamine stimulation of lipolysis than cells prepared by a commonly used method (Hittelman, K.J., Wu, C.F. and Butcher, R.W. (1973) Biochim. Biophys. Acta 304, 188-196) and also more sensitive to the anti-lipolytic action of insulin. The effects of insulin on lipogenesis, measured as [3H]glucose conversion to cell lipids, and on catecholamine-stimulated lipolysis were compared under identical conditions with the same cell batch. Isoprenaline-stimulated lipolysis was found to be half-maximally inhibited by an insulin concentration 8-fold lower than that stimulating lipogenesis to a corresponding extent (half-maximal effects at insulin concentrations of 40 vs. 300 pM). A similar difference was found when cells had been stimulated with adrenaline instead of isoprenaline.

The robustness of recombination frequency estimates in intercrosses with dominant markers.

The robustness of the maximum likelihood estimates of recombination frequencies has been investigated in double intercrosses with complete dominance at both loci. The robustness was investigated with respect to bias in the recombination frequency estimates due to: (1) limited sample sizes, (2) heterogeneity in recombination frequencies between sexes or among meioses and (3) factors that distort the segregation-misclassification or differential viability. In the coupling phase, the recombination frequency estimates are quite robust with respect to most of the investigated factors. Potentially, the most serious cause of a bias is misclassifications, which tend to increase the recombination frequency estimates. In the repulsion phase, misclassifications are particularly serious, leading to extreme discrepancies between true and observed values. In addition, limited sample size and sex differences in recombination can also bias recombination frequency estimates in repulsion. These effects may pose serious problem in genetic mapping with random amplified polymorphic DNA (RAPD) markers.

Positive correlation between recombination rates and levels of genetic variation in natural populations of sea beet (Beta vulgaris subsp. maritima).

The relation between the level of genetic variation and the rate of recombination per physical unit was investigated in sea beet (Beta vulgaris subsp. maritima). The rate of recombination per physical unit was estimated indirectly through marker density in an RFLP linkage map of sugar beet. From this map, we also selected RFLP markers covering two of the nine chromosomes in Beta. The markers were used to estimate the level of genetic variation in three populations of sea beet, two from Italy and one from England. Two estimates of genetic variation were employed, one based on the number of alleles in the sample and the other on heterozygosity. A statistically significant positive correlation was found between recombination rate and genetic variation. Several theoretical explanations for this are discussed, background selection being one. A correlation similar to this has been observed previously in Drosophila, one that was higher than what we obtained for Beta. This is consistent with various biological differences between the two species.

Effects of acetate, acetaldehyde, and ethanol on lipolysis in isolated rat adipocytes.

The effects of different concentrations of acetate, acetaldehyde, and ethanol, alone or in combination, on the lipolysis rate, measured as glycerol release, were studied in isolated adipocytes from fed or fasted rats, in the basal state and at various levels of norepinephrine stimulation. Acetate inhibited the glycerol release in a dose-dependent manner (congruent to 10% inhibition at 2 mM, 25--70% at 10 mM) with the most marked effects at low to moderate norepinephrine concentratione and at maximal (100 ng/ml) hormone stimulation. Ethanol, up to 100 mM concentration, had no effect on the lipolysis rate. Acetaldehyde, up to 1 mM concentration, had no reproducible effect. Ethanol, acetaldehyde, and acetate in combination inhibited glycerol release to an extent similar to that of acetate alone.

A model of chiasma reduction of closely formed crossovers.

A model of chiasma reduction is developed, evaluated by computer simulation and discussed in relation to the evolution of interference. The model assumes that adjacently formed crossovers can interact, if there is incomplete sister chromatid cohesion between them, and give rise to a reduced number of chiasmata. In the absence of crossover position interference this leads to a considerable risk of non-disjunction for an average sized chromosome. It is suggested that an important function of crossover interference is to reduce this effect. The question is raised whether chiasma reduction takes place today. Some available cytological data can be interpreted as showing that chiasma reduction occurs in normal meiosis. The possibility of chiasma reduction therefore needs further attention.

Crossover distribution in barley analysed through RFLP linkage data.

The pattern of recombination in barley with regard to (1) the distribution of crossover points among whole gametes, (2) the distribution of crossover points among individual chromosomes and (3) the distribution of crossover points within chromosomes has been analysed using data sets underlying two recently published restriction fragment length polymorphism (RFLP) linkage maps representing male and female meiosis, respectively. The data indicated that the process of recombination had been random with no interference. The two data sets gave similar results, indicating that male and female meiosis in barley do no differ significantly. The possibility of using RFLP data in studies of crossover distribution is stressed.

Continuous monitoring of free fatty acid release from adipocytes by pH-stat titration.

A methods for direct and continuous monitoring of free fatty acid release in adipocyte suspensions is described. Using a pH-stat apparatus the protons from the released free fatty acids are continuously titrated and the accumulated amount of OH- added is monitored on a recorder against time, the slope thus indicating the rate of free fatty acid release. Sinc pH is kept constant, an incubation medium with a low buffering capacity can be used, which gives the method a high sensitivity. Under the conditions described, free fatty acid release from 5% of maximal norepinephrine stumulation of rat adipocytes can be accurately measured and the kinetics can be followed over extended periods of time.

Regulation of adipose-tissue lipolysis by phosphorylation of hormone-sensitive lipase.

Hormone-sensitive lipase has been purified from rat adipose tissue to almost 50 per cent protein purity and partially characterized. The isolated enzyme can be phosphorylated by ATP-Mg2+ in the presence of the catalytic subunit of cyclic AMP-dependent protein kinase from the same tissue. Its activity towards emulsified triglyceride is thereby increased two-fold. The enzyme is phosphorylated also in the intact adipocyte, verifying the physiological relevance of the findings with the isolated enzyme. Noradrenaline causes a rapid increase in phosphorylation of the enzyme in intact adipocytes, immediately followed by a marked increase of its activity. Addition of dibutyryl-cyclic AMP to the adipocytes causes the same effects. The extent of phosphorylation of the enzyme after maximal noradrenaline stimulation of the adipocytes is rapidly decreased by insulin addition in close association with inhibition of the lipase activity. The results demonstrate that these hormones regulate the activity of the hormone-sensitive lipase, ie the rate of lipolysis in the adipocytes, by changes of the degree of phosphorylation of the enzyme.

Effects of distance and pollen competition on gene flow in the wind-pollinated grass Festuca pratensis Huds.

Pollen dispersal and gene flow in the grass meadow fescue (Festuca pratensis Huds.) were studied using two populations which were homozygous for different allozymes at the Gpi-2 locus. The populations were established in a concentric donor-acceptor field experiment. Gene flow was found mainly to be affected by the distance between the donor and acceptor plants. Analysing 21 132 acceptor plant progenies, gene flow was shown to decrease rapidly with distance to the donor field up to 75 m, and beyond this distance much more slowly. The ability of donor pollen to fertilize acceptor plants depended very much on the density of the acceptor plants. Pairs of acceptor plants produced more compatible pollen locally, and captured significantly less donor pollen than single-plants. Despite the higher seed production of acceptor plants planted in pairs, the absolute number of heterozygous seeds carrying the donor allele was always lower than for single-plants. Wind direction had only a slight effect upon the type of pollen captured. Because of pollen production within the two plant populations being continuous and overlapping, the time when anthesis occurred had little effect on gene flow between the populations. Vigorous and tall acceptor plants with many panicles, high seed yield and high 1000-seed weight were able to capture more donor pollen than shorter plants. The results may be used to assess the risk of gene flow and to develop strategies for monitoring the spread of transgenes from genetically modified grasses.

Effects of prolonged food restriction on some aspects of lipid metabolism in Norwegian and Svalbard reindeer.

The high-arctic Svalbard reindeer (SR) deposit great amounts of body fat in autumn for subsequent use during winter when food is often in short supply. Captive SR and, for comparative reasons, the sub-arctic Norwegian reindeer (NR) were offered 15% of their ad libitum food intake during a 21-day period in September/October and its effect on fat metabolism was investigated. Plasma free fatty acids (FFA), glycerol, glucose, insulin and urea as well as lipogenic and lipolytic activity of isolated adipocytes were determined. Levels of FFA and glycerol increased immediately when food intake was restricted, reaching the highest levels in SR. Plasma glucose was fairly constant in NR, but decreased in SR. Plasma insulin decreased in both species. Plasma urea increased steadily from day 5 in NR and from day 11 in SR, after a transient rise on day 1 in both. Lipogenic activity had vanished completely in both NR and SR adipocytes when tested after 13 days of food restriction, while lipolytic activity was initially increased, after which it decreased in adipocytes from both species. After 21 days of food restriction SR adipocytes exhibited another marked increase in lipolytic activity, while the fat deposits of NR at that time were too small to allow examination. Thus, reindeer do not differ from other species in their lipogenic responses, although they show some hitherto undescribed lipolytic responses to prolonged food restriction.

Seasonal changes in lipogenesis and lipolysis in isolated adipocytes from Svalbard and Norwegian reindeer.

Arctic reindeer exhibit marked seasonal changes in fat deposition and mobilization. At intervals throughout the year, therefore, we have measured feed intake of both Svalbard (SR) and Norwegian reindeer (NR) together with the seasonal changes in size, lipogenic and lipolytic capacity of isolated adipocytes from both sub-species. Feed intake of both NR and SR was maximal in August, but declined thereafter, reaching minimum values in January (NR) and March (SR), 55 and 69% below the August value, respectively. NR and SR adipocyte volume changed in parallel and were reduced to the same extent (69%) from their maximum in August to their minimum in May. Adipocyte lipogenic capacity, measured as acetate incorporation into cellular lipid at saturated acetate concentrations, was lowest in January (NR adipocytes) and March (SR adipocytes), 92 and 90%, respectively, below the maximum values, which were obtained in August. Lipolytic capacity, measured as maximum adrenaline-stimulated glycerol release, was high in SR adipocytes from March through to October and in NR adipocytes from July through to January. Minimum lipolytic capacity, on the other hand, was found in January (SR adipocytes) and March (NR adipocytes). The present findings may be explained by alterations in lipogenic enzyme activity and in the lipolytic activation system.

Effects of volatile fatty acids and ketone bodies on lipolysis in isolated adipocytes from Norwegian reindeer (Rangifer tarandus).

The effect of volatile fatty acids (acetate and propionate) and ketone bodies (acetoacetate and DL-3-hydroxybutyrate) on adrenaline stimulated lipolysis have been studied in isolated adipocytes from Norwegian reindeer. All four substances caused a strong (63-90%) inhibition of lipolysis, measured as glycerol release, at low concentrations of adrenaline (5 nM). At higher adrenaline concentrations (50 nM) inhibition only took place in the presence of propionate (30% inhibition) or DL-3-hydroxybutyrate (43% inhibition). DL-3-hydroxybutyrate caused a marked reduction in adrenaline stimulated glycerol release, even at very low concentrations. The effective dose causing 50% inhibition (ED50) was 1.5 mM. It is suggested that both volatile fatty acids and ketone bodies can act as important volatile regulators of lipopolysis in this high arctic ruminant undergoing great seasonal changes in volatile fatty acid and supposedly ketone body production.