Pre-diagnostic genotyping identifies T1D subjects with impaired Treg IL-2 signaling and an elevated proportion of FOXP3+IL-17+ cells.

T-regulatory cells (Tregs) are essential for immune tolerance, and animal studies implicate their dysfunction in type 1 diabetes (T1D) pathogenesis. Tregs require interleukin-2 (IL-2) for their suppressive function, and variants in IL-2/IL-2R pathway genes have been associated with T1D. We previously reported that recent-onset T1D subjects have an increased population of FOXP3lo Tregs that secrete the pro-inflammatory cytokine, interleukin-17 (IL-17). We hypothesize that IL-2 signaling defects may drive T1D development by skewing protective Tregs towards an inflammatory Th17 phenotype. Overall, we found that the proportion of FOXP3+IL-17+ cells in T1D subjects pre-diagnosis was unchanged compared with healthy controls. However, stratification by IL2RA single-nucleotide polymorphisms revealed that T1D subjects with the rs3118470 CC risk variant have Tregs with IL-2 signaling defects and an increased proportion of FOXP3+IL-17+ cells before diagnosis. These data suggest a potential mechanism for genetically controlled loss of Treg function via dysfunctional IL-2 signaling in T1D.

Interaction between the Val158Met catechol-O-methyltransferase gene variant and second-generation antipsychotic treatment on blood pressure in children.

Second-generation antipsychotic (SGA) medications are associated with cardiometabolic risk factors such as obesity and elevated blood pressure (BP) in some individuals. The goal of this study is to determine whether the Val158Met variant (rs4680) in the catechol-O-methyltransferase (COMT) gene, associated with BP in adults, is associated with elevated BP in SGA-treated children. A cross-sectional population of SGA-treated (n=134) and SGA-naive (n=168) children, ⩽18 years of age, were genotyped and assessed for markers of cardiometabolic health. An interaction was found between SGA treatment and COMT genotype for BP. After adjusting for covariates, SGA-treated children with the Met allele had higher systolic and diastolic BP (P=0.014 and P=0.034, respectively), and higher fasting glucose concentrations (P=0.030) compared with children with the Val/Val genotype. This was not observed in SGA-naive children. The Met allele of the COMT Val158Met variant may identify SGA-treated children at risk for elevated BP and fasting blood glucose concentrations.

Cardiometabolic risk and the MTHFR C677T variant in children treated with second-generation antipsychotics.

Second-generation antipsychotics (SGAs) are increasingly being used to treat children with a variety of psychiatric illnesses. Metabolic syndrome (MetS), a risk factor for cardiovascular disease, is a side-effect of SGA-treatment. We conducted a cross-sectional study and assessed the association of the methylenetetrahydrofolate reductase (MTHFR) C677T variant with features of MetS in SGA-treated (n=105) and SGA-naïve (n=112) children. We targeted the MTHFR C677T variant, because it is associated with risk for cardiovascular disease, and features of MetS in adults without psychiatric illness. MetS in children is based on the presence of any three of the following: waist circumference ≥ 90th percentile for age and sex; plasma triglyceride ≥ 1.24 mmol l(-1); plasma high-density lipoprotein-cholesterol ≤ 1.03 mmol l(-1); systolic or diastolic blood pressure ≥ 90th percentile for age, sex, and height; and fasting glucose ≥ 5.6 mmol l(-1). We found that 15% of SGA-treated children had MetS compared with 2% of SGA-naïve children (OR 8.113, P<0.05). No effect of the MTHFR C677T variant on psychiatric diagnosis was observed. The MTHFR 677T allele was associated (P<0.05) with MetS (OR 5.75, 95% CI= 1.18-28.12) in SGA-treated children. Models adjusted for duration of SGA treatment, ethnicity, sex, age and use of other medications revealed a positive relationship between the MTHFR 677T allele and diastolic blood pressure Z-scores (P=0.001) and fasting plasma glucose (P<0.05) in SGA-treated children. These findings illustrate the high prevalence of MetS in SGA-treated children and suggest metabolic alterations associated with the MTHFR C677T variant may have a role in the development of MetS features in SGA-treated children.

Impaired proinsulin processing is a characteristic of transplanted islets.

We sought to determine whether recipients of islet transplants have defective proinsulin processing. Individuals who had islet allo- or autotransplantation were compared to healthy nondiabetic subjects. Insulin (I), total proinsulin (TP), intact proinsulin and C-peptide (CP) were measured in samples of fasting serum by immunoassay, and the ratios of TP/TP+I and TP/CP were calculated. Islet allotransplant recipients had elevated TP levels relative to nondiabetic controls (16.8 [5.5-28.8] vs. 8.4 [4.0-21.8] pmol/L; p < 0.05) and autologous transplant recipients (7.3 [0.3-82.3] pmol/L; p < 0.05). Islet autotransplant recipients had significantly higher TP/TP+I ratios relative to nondiabetic controls (35.9 +/- 6.4 vs. 13.9 +/- 1.4%; p < 0.001). Islet allotransplant recipients, some of whom were on insulin, tended to have higher TP/TP+I ratios. The TP/CP ratio was significantly higher in both islet autotransplant (8.9 [0.6-105.2]; p < 0.05) and allotransplant recipients (2.4 [0.8-8.8]; p < 0.001) relative to nondiabetic controls (1.4 [0.5-2.6]%). Consistent with these findings, TP/TP+I and TP/CP values in islet autotransplant recipients increased significantly by 1-year posttransplant compared to preoperative levels (TP/CP: 3.8 +/- 0.6 vs. 23.3 +/- 7.9%; p < 0.05). Both allo- and autotransplant subjects who received <10,000 IE/kg had higher TP/CP ratios than those who received >10,000 IE/kg. Islet transplant recipients exhibit defects in the processing of proinsulin similar to that observed in subjects with type 2 diabetes manifest as higher levels of total proinsulin and increased TP/TP+I and TP/CP ratios.

Diabetes screening of children in a remote First Nations community on the west coast of Canada: challenges and solutions.

Type 2 diabetes (T2D) and its precursor, impaired glucose tolerance (IGT), are now reaching epidemic proportions among Aboriginal Canadians. Of particular concern is the appearance and increasing prevalence of T2D and IGT among Aboriginal youth. At the request of three communities in the Tsimshian nation on the northern coast of British Columbia (with which the Department of Pediatrics, University of British Columbia, had a pre-existing partnership) a screening program was undertaken to determine the prevalence of T2D and IGT among the children. The long-term goal was the collaborative development of intervention programs for each community. The challenges of meeting this request included the sociological and ethical issues associated with research in First Nations communities, as well as the pragmatic issues of conducting complex research in remote communities. Three separate visits were undertaken to respect the cultural dynamics and capacity of the community to accommodate a project of this magnitude. The process began with dialogue, listening and presentations to the community. Only then began the planning of logistics and application for funding. Next, the team visited the communities to ensure understanding of exactly what was involved for the community, each child and family, and to be certain that consent was fully informed. For the diabetes screening visit, special arrangements including chartering a Beaver float plane were needed for the transport of the five-member team with all the necessary equipment, including a -20(o)C freezer to safeguard the integrity of blood samples. The 100% consent rate, successful conduct of study, and retention of community support achieved by the process, indicate that population-based clinical research is possible in remote First Nations communities. This is best achieved with appropriate dialogue, care, respect and planning to overcome the sociological, ethical and practical challenges.

Sub-ambient temperature effects on the separation of monosaccharides by high-performance anion-exchange chromatography with pulse amperometric detection. Application to marine chemistry.

The effects of column temperature in the range 10-45 degrees C using high-performance anion-exchange chromatography (HPAEC) and pulse amperometric detection are described for the determination of monosaccharides. The influence of temperature was tested with an isocratic elution of NaOH at concentrations varying from 2.5 to 20 mM and with a post-column addition of 1 M NaOH. The results showed that small changes of temperature greatly affect retention times and resolution (Rs) of monosaccharides and particularly those of the both pairs xylose-mannose and rhamnose-arabinose which cannot be simultaneously detected at usual room temperature (approximately 25 degrees C). Our results suggest that a subambient temperature of 17 degrees C and an eluent concentration of 19 mM are the more appropriate conditions for an acceptable separation (R(s rha/ara) = 1.02, R(s man/xyl) = 0.70) in a short analytical run time (35 min). The results showed that within the range of temperatures studied, enthalpy and entropy are invariant of temperature indicating that changes in the retention processes are mainly due to temperature than other associated changes in the system. This study demonstrated the importance of controlling temperature during HPAEC of monosaccharides, both to accomplish highly reproducible retention times and to achieve optimal separation of sugars. This method gave acceptable results for detection of marine sugars.

Electrocardiographic findings in adolescents with eating disorders.

OBJECTIVES: We sought to determine the prevalence and clinical correlates of electrocardiographic findings in adolescents with eating disorders. STUDY DESIGN: We undertook matched case-control study of electrocardiographic findings at initial assessment in 62 adolescents with anorexia nervosa, 9 with bulimia nervosa, and 26 with eating disorder not otherwise specified presenting from March 1995 to September 1996. RESULTS: Mean (+/- 1 standard deviation) age was 15.0 +/- 1.4 years (95% were female). Patients with anorexia nervosa had significantly lower heart rates (mean case-control difference: -20 +/- 17 beats per minute), lower R in V(6) (-2.6 +/- 5.5 mm), longer QRS interval (+.004 +/-.010 seconds), shorter mean QTc (-.0136 +/-.033 seconds) and lesser QTc dispersion (-.010 +/-. 031 seconds). The bulimia nervosa group had slightly longer mean QTc (.019 +/-.020 seconds), with no significant case-control differences in the eating disorder not otherwise specified group. CONCLUSION: Electrocardiographic findings are abnormal in adolescents with anorexia nervosa but not in adolescents with bulimia nervosa or eating disorder not otherwise specified.

Validity of newborn oscillometric blood pressure.

The objective of this study was to determine the validity of oscillometric blood pressure in relation to arterial blood pressure. Thirty-one newborns were studied. Clinical characteristics, complications, and treatment interventions were documented. Arterial pressure (every 2 sec) and oscillometric pressure (every 3 min) were concurrently recorded for 1-2 h. Serial observations of oscillometric pressure followed the trend of arterial pressure in the individual newborn. However, the study averaged oscillometric pressures were lower than the arterial pressures: systolic, by 1 mmHg; mean, by 5.3 mmHg (p < 0.0001); and diastolic, by 4.6 mmHg (p < 0.0001). The variance of individual, 15-min averaged, and 1-h averaged observations of oscillometric pressure in relation to arterial pressure was examined. Variance for individual observations may be large. The least variance of oscillometric pressure was in the 1-h averaged mean pressure, in which the difference was +/- 2 mmHg in 77% and +/- 4 mmHg in 95% of observations. Clinical characteristics, with the exception of birth weight and treatment interventions, did not affect the variance of oscillometric pressure. This study implies that the offset in relation to arterial pressure should be established for each oscillometric pressure monitoring system. Hourly averaged mean oscillometric pressure is satisfactory for many newborn assessments and management circumstances. However, arterial pressure may be necessary to accurately document transient hypertension or hypotension or an unstable blood pressure.

Newborn complications after intrapartum asphyxia with metabolic acidosis in the preterm fetus.

OBJECTIVE: Our purpose was to determine the nature of the complications in preterm newborns after intrapartum fetal asphyxia with metabolic acidosis at delivery. STUDY DESIGN: Thirty-seven preterm fetuses with metabolic acidosis were matched with 37 preterm fetuses with normal blood gas measurements at delivery. A complication score expressed the magnitude of newborn complications during the 10 days after delivery. RESULTS: The mean complication score for the preterm newborns at 32 to 36 weeks in the asphyxia group, 9.6, was significantly greater than that for the control group, 3.1. Fetal asphyxia was associated with severe complications in all systems. The complication scores for the preterm newborns < 32 weeks in the asphyxia group were of the same order as the control group. This may be related in part to a short duration of the asphyxial insult. The Apgar score at 1 minute was a valuable predictor of newborn complications in both the asphyxia and control groups. CONCLUSIONS: Intrapartum fetal asphyxia with metabolic acidosis at delivery is an important factor in the occurrence of severe complications, particularly in the central nervous system, respiratory system, and kidney, of preterm newborns.

Newborn complications after intrapartum asphyxia with metabolic acidosis in the term fetus.

OBJECTIVES: Our purpose was to determine the newborn complications after respiratory or metabolic acidosis at delivery and to demonstrate the characteristics of an asphyxial insult predictive of these complications. STUDY DESIGN: Fifty-nine term fetuses with metabolic acidosis were matched with 59 fetuses with normal blood gas measures at delivery. Fifty-one fetuses with respiratory acidosis were also examined. A complication score expressed the magnitude of newborn complications during the 10 days after delivery. RESULTS: Newborn complications were not increased after respiratory acidosis. Newborn complications after metabolic acidosis increase in frequency and severity with the increasing severity and duration of the metabolic acidosis. Thirty-two of the 59 newborns in the metabolic acidosis group had a high complication score. The index values predictive of high scores were the duration of the metabolic acidosis and the Apgar score at 1 minute. CONCLUSIONS: Intrapartum fetal asphyxia with a severe metabolic acidosis accounts for complications in all newborn systems. The probability of a high complication score increases from 14% with favorable index values to 85% with unfavorable index values.

Rheumatoid arthritis serum or synovial fluid and interleukin 2 abnormally expand natural killer-like cells that are potent stimulators of IgM rheumatoid factor.

We show that rheumatoid arthritis (RA) serum or synovial fluid (SF) increases the growth capacity of normal, interleukin 2 (IL-2) driven cell preparations, compared to normal human serum (NHS). Proliferation in RA serum and SF cultures was primarily associated with expansion of natural killer (NK)-like cells (CD16+, CD57+), and in NHS cultures, with T cell (CD3+ CD4+ CD8+) growth. The capacity of RA serum to promote NK cell growth was related to patient global clinical activity and rheumatoid factor (RF) titers. The NK-like cells, but not the T-like cells, induced high levels of IgM RF synthesis in autologous B cells. Thus, alteration in NK cell growth may disrupt NK-B cell circuits in RA and contribute to B cell dysfunction (RF synthesis).

Distinct characteristics of lymphokine-activated killer (LAK) cells derived from patients with B-cell chronic lymphocytic leukemia (B-CLL). A factor in B-CLL serum promotes natural killer cell-like LAK cell growth.

We show that lymphokine-activated killer (LAK) cell precursors derived from patients with B-cell chronic lymphocytic leukemia (B-CLL) and cultured in the presence of recombinant interleukin-2 and normal human serum (NHS), develop into primarily NK cell-like (CD 57+) LAK cells, whereas identically prepared LAK cell precursors from normal subjects develop into mainly T cell-like (CD 3+, CD 8+) LAK cells. B-CLL LAK cells exhibited greater proliferative capacity than did normal LAK cells. When normal LAK cells were grown in B-CLL serum instead of NHS, their proliferation increased; NK cell levels also increased to those found in B-CLL LAK cells, suggesting that B-CLL serum contains a factor that promotes NK cell-like growth, LAK cells derived from normal or B-CLL patients demonstrated similar lytic activity toward K562 and Raji cells. Growth in B-CLL serum did not reduce their lytic potential. Thus, the altered phenotype and growth exhibited by B-CLL LAK cells and normal LAK cells grown in B-CLL serum does not lead to abnormalities in their cytolytic functions. We propose instead that the predominance of NK-like cells in B-CLL LAK cell populations and the presence of an NK cell-like growth factor in B-CLL serum reflect abnormalities related to NK cell-mediated B-cell regulation; ie, either inhibition of normal B-cell growth and/or growth stimulation of the leukemic clone in B-CLL.

Mitogenesis of 3T3 fibroblasts induced by endogenous ganglioside is not mediated by cAMP, protein kinase C, or phosphoinositides turnover.

The B subunit of cholera toxin, which binds specifically to ganglioside GM1, stimulates DNA synthesis in quiescent Swiss 3T3 fibroblasts grown in chemically defined medium. The mitogenic response to the B subunit was potentiated by insulin and other growth factors. To elucidate the mechanism by which the B subunit stimulates cell growth , its effects on several transmembrane signaling systems which have been suggested to play a vital role in cell growth regulation were examined. The B subunit did not increase cAMP levels nor activate adenylate cyclase. The B subunit induced a rapid and profound increase in intracellular free Ca2+ as measured with the fluorescent Ca2+-sensitive dye quin 2/AM. Removal of external Ca2+ completely inhibited the signal, thus suggesting that the B subunit elevates intracellular Ca2+ through a net influx of extracellular Ca2+ rather than by causing the release of Ca2+ from intracellular stores. These findings are consistent with the observations that the B subunit induced reinitiation of DNA synthesis without activation of phospholipase C. There was no increase in the formation of inositol trisphosphate, the second messenger that mediates release of Ca2+ from intracellular stores. In addition, the B subunit still stimulated DNA synthesis in Swiss 3T3 cells pretreated with phorbol ester to down-regulate protein kinase C. These results suggest that the mitogenic effects of the B subunit are mediated mainly by facilitation of Ca2+ influx and that activations of adenylate cyclase, phospholipase C, or protein kinase C are not obligatory steps in the initiation of cell growth by the B subunit. Furthermore, the observation that Ca2+ ionophores, such as ionomycin and A23187, are not mitogenic implies that additional undefined growth signaling pathways may exist in this system.