De novo and inherited mutations in the X-linked gene CLCN4 are associated with syndromic intellectual disability and behavior and seizure disorders in males and females.

Variants in CLCN4, which encodes the chloride/hydrogen ion exchanger CIC-4 prominently expressed in brain, were recently described to cause X-linked intellectual disability and epilepsy. We present detailed phenotypic information on 52 individuals from 16 families with CLCN4-related disorder: 5 affected females and 2 affected males with a de novo variant in CLCN4 (6 individuals previously unreported) and 27 affected males, 3 affected females and 15 asymptomatic female carriers from 9 families with inherited CLCN4 variants (4 families previously unreported). Intellectual disability ranged from borderline to profound. Behavioral and psychiatric disorders were common in both child- and adulthood, and included autistic features, mood disorders, obsessive-compulsive behaviors and hetero- and autoaggression. Epilepsy was common, with severity ranging from epileptic encephalopathy to well-controlled seizures. Several affected individuals showed white matter changes on cerebral neuroimaging and progressive neurological symptoms, including movement disorders and spasticity. Heterozygous females can be as severely affected as males. The variability of symptoms in females is not correlated with the X inactivation pattern studied in their blood. The mutation spectrum includes frameshift, missense and splice site variants and one single-exon deletion. All missense variants were predicted to affect CLCN4's function based on in silico tools and either segregated with the phenotype in the family or were de novo. Pathogenicity of all previously unreported missense variants was further supported by electrophysiological studies in Xenopus laevis oocytes. We compare CLCN4-related disorder with conditions related to dysfunction of other members of the CLC family.

In utero gene therapy rescues microcephaly caused by Pqbp1-hypofunction in neural stem progenitor cells.

Human mutations in PQBP1, a molecule involved in transcription and splicing, result in a reduced but architecturally normal brain. Examination of a conditional Pqbp1-knockout (cKO) mouse with microcephaly failed to reveal either abnormal centrosomes or mitotic spindles, increased neurogenesis from the neural stem progenitor cell (NSPC) pool or increased cell death in vivo. Instead, we observed an increase in the length of the cell cycle, particularly for the M phase in NSPCs. Corresponding to the developmental expression of Pqbp1, the stem cell pool in vivo was decreased at E10 and remained at a low level during neurogenesis (E15) in Pqbp1-cKO mice. The expression profiles of NSPCs derived from the cKO mouse revealed significant changes in gene groups that control the M phase, including anaphase-promoting complex genes, via aberrant transcription and RNA splicing. Exogenous Apc4, a hub protein in the network of affected genes, recovered the cell cycle, proliferation, and cell phenotypes of NSPCs caused by Pqbp1-cKO. These data reveal a mechanism of brain size control based on the simple reduction of the NSPC pool by cell cycle time elongation. Finally, we demonstrated that in utero gene therapy for Pqbp1-cKO mice by intraperitoneal injection of the PQBP1-AAV vector at E10 successfully rescued microcephaly with preserved cortical structures and improved behavioral abnormalities in Pqbp1-cKO mice, opening a new strategy for treating this intractable developmental disorder.

The X-chromosome-linked intellectual disability protein PQBP1 is a component of neuronal RNA granules and regulates the appearance of stress granules.

The polyglutamine-binding protein 1 (PQBP1) has been linked to several X-linked intellectual disability disorders and progressive neurodegenerative diseases. While it is currently known that PQBP1 localizes in nuclear speckles and is engaged in transcription and splicing, we have now identified a cytoplasmic pool of PQBP1. Analysis of PQBP1 complexes revealed six novel interacting proteins, namely the RNA-binding proteins KSRP, SFPQ/PSF, DDX1 and Caprin-1, and two subunits of the intracellular transport-related dynactin complex, p150(Glued) and p27. PQBP1 protein complex formation is dependent on the presence of RNA. Immunofluorescence studies revealed that in primary neurons, PQBP1 co-localizes with its interaction partners in specific cytoplasmic granules, which stained positive for RNA. Our results suggest that PQBP1 plays a role in cytoplasmic mRNA metabolism. This is further supported by the partial co-localization and interaction of PQBP1 with the fragile X mental retardation protein (FMRP), which is one of the best-studied proteins found in RNA granules. In further studies, we show that arsenite-induced oxidative stress caused relocalization of PQBP1 to stress granules (SGs), where PQBP1 co-localizes with the new binding partners as well as with FMRP. Additional results indicated that the cellular distribution of PQBP1 plays a role in SG assembly. Together these data demonstrate a role for PQBP1 in the modulation of SGs and suggest its involvement in the transport of neuronal RNA granules, which are of critical importance for the development and maintenance of neuronal networks, thus illuminating a route by which PQBP1 aberrations might influence cognitive function.

Characterization of a murine gene homologous to the bovine CaCC chloride channel.

The bovine CaCC protein is a putative Ca2+-dependent Cl- channel of airway epithelial cells. Therefore, CaCC proteins could contribute to transepithelial Cl- transport and accordingly modify the phenotype of cystic fibrosis (CF) patients. We have identified a murine EST containing a full-length cDNA coding for a 902-amino-acid protein highly homologous to bovine CaCC. The murine gene (mCaCC) maps to chromosome 3 at the H2-H3 band and is expressed, as indicated by Northern blot analysis, in mouse skin and kidney but not in brain, heart, lung or testis. RT-PCR indicates a low expression in tracheal epithelial cells. Heterologous expression of mCaCC in Xenopus oocytes elicits membrane currents that are anion-selective and inhibited by DIDS and by niflumic acid, a blocker of the endogenous chloride current in oocytes. The identification of genes belonging to the CaCC family will help to evaluate their role as ion channels or channel regulators and their actual contribution to epithelial chloride transport.

Characterization of the human gene coding for the swelling-dependent chloride channel ICln at position 11q13.5-14.1 (CLNS1A) and further characterization of the chromosome 6 (CLNS1B) localization.

Expression cloning revealed a chloride channel (ICln) that we found to be fundamental for the regulatory volume decrease in a variety of cells. The chromosomal localization of the human ICln-gene showed two loci, one at chromosome 11 in position q13.5-q14.1, termed CLNS1A, and a second one at chromosome 6 at position p12.1-q13, termed CLNS1B. In this study, we offer a detailed characterization of the CLNS1A gene and provide the exact position (6p12) and sequence data of CLNS1B, an intronless gene 91.3% homologous to the coding region of CLNS1A.

Measurements of (Na+,K+)ATPase after in vitro hypoxia and reoxygenation are affected by methods of membrane preparation.

(Na+,K+ )ATPase activity was evaluated in membranes from rat hippocampal slices after in vitro hypoxia and reoxygenation. Membranes were prepared with two different methods, one using an isotonic medium and another using a hypotonic one. The changes that were found after hypoxia went into opposite directions in the two cases. Membranes prepared in a hypotonic medium are probably more suitable for these measurements. Using these membranes, hypoxia results in a slight decrease of (Na+,K+)ATPase activity and in a further decrease after reoxygenation. We also found that expressing (Na+,K+)ATPase activity as a percent of total ATPase activity is appropriate for membranes prepared under hypotonic conditions and can unveil (by reducing variability between experiments) significant changes that may be masked in small samples like ours.

Desirability of control versus locus of control: relationship to paralinguistics in the type A interview.

David Glass has suggested that need for control is an important dimension underlying the Type A coronary-prone behavior pattern. However, this hypothesis has never been directly tested. Findings from the present study supported the hypothesis by showing a significant relationship between measures of desire for control and the Type A pattern as assessed by both the structured interview and questionnaire methods, and with Type A voice stylistics. In contrast, locus of control scores correlated only with the questionnaire-based measure of Type A, and were unrelated to desire for control scores, interview-defined Type A, and Type A noncontent speech characteristics. Some evidence suggests the hypothesis that autonomic nervous system arousal can create psychological discomfort that may serve to increase both need for control and Type A vocal stylistics. In addition, such vocal mannerisms may actually facilitate control of the social environment. Evidence implicating genetic factors in these processes is discussed.

Smoking, caffeine, and stress: effects on blood pressure and heart rate in male and female college students.

This study examined the independent and joint effects of cigarette smoking and caffeine consumption on blood pressure (BP) and heart rate (HR) reactions to stress in male and female college students. Following an initial physiological baseline, participants received one of four experimental treatments (paced smoking, caffeine, smoking plus caffeine, or neither) and had BP and HR measured before, during, and after two stressful tasks. The results revealed that, compared with the control condition, caffeine ingestion enhanced the magnitude of stress-induced systolic BP and HR reactions. Smoking combined additively with stress, and the joint effect of smoking and caffeine was no greater than either taken alone. Males and females were generally similar in their BP and HR responses to smoking, caffeine, and stress. Inconsistencies with previous research and possible physiological mechanisms underlying the observed effects are discussed.

Potential for hostility and dimensions of anger.

Recent reviews have linked Potential for Hostility derived from the Structured Interview (SI) to coronary artery disease, independent of the global Type A pattern. The present study examined the construct validity of Potential for Hostility ratings by correlating Potential for Hostility with 21 scales from four widely used anger/hostility measures: 7 scales from the Anger Self-Report, 8 scales from the Buss-Durkee Hostility Inventory, the total score from the Novaco Anger Inventory, and 5 scales from the Multidimensional Anger Inventory. The pattern of correlations revealed that Potential for Hostility was significantly related to scales reflecting awareness and arousal of anger, particularly the verbal expression of anger. To identify underlying anger dimensions, the 21 scales were factor-analyzed. Examination of two and three rotated principal components confirmed previous solutions. The first component, representing anger-arousing and -eliciting situations and anger awareness, was labeled Experience of Anger. The second component, consisting of scales dealing with either physical assault or verbal expression of anger, was labeled Expression of Anger. When a third factor was retained, it contained scales of suspicion, mistrust-suspicion, and guilt: It was therefore labeled Suspicion-Guilt. Potential for Hostility was correlated only with the Expression of Anger factor in the two-factor solution; in the three-factor solution, Potential for Hostility was correlated equally with the Experience of Anger and Expression of Anger factors but was not correlated with the Suspicion-Guilt factor. The implications of these results for the assessment of hostility are discussed.

Ethnicity, gender, family history of myocardial infarction, and hemodynamic responses to laboratory stressors in children.

Relationship among ethnicity, gender, grandparents' histories of early myocardial infarction, and hemodynamic responses to forehead cold and treadmill exercise were examined in 87 6-to-8-year-olds (57 White, 30 Black). Boys had greater increases in systemic vascular resistance and decreases in cardiac index to forehead cold. Girls had greater increases and quicker recovery in heart rate to dynamic exercise. Blacks had greater increases and slower recovery in diastolic pressure to exercise and forehead cold. Blacks showed greater increases and slower recovery in systemic vascular resistance to forehead cold. Positive-family-history children had greater increases in diastolic pressure and systemic vascular resistance to forehead cold and greater diastolic pressure increases to exercise. Positive-family-history Blacks had greater increases in systolic pressure to exercise and slower recovery than all other groups.

Component analysis of the type A coronary-prone behavior pattern in male and female college students.

Two studies investigated the psychological dimensions encompassed by the Rosenman Structured Interview (SI) method of assessing the Type A behavior pattern. In Study 1, female students completed the SI and Jenkins Activity Survey (JAS). Factor analysis of the verbal stylistic and answer content components of the SI yielded a 5-factor solution, with one factor--Clinical Rating--accounting for most nonerror variance in Type A ratings. The remaining factors were derived from answer content and correlated more substantially with JAS Type A ratings than with SI Type A ratings. In Study 2, male and female college students completed the SI and JAS and a battery of questionnaires that tap trait dimensions implied by the conceptual definition of Type A. For both males and females, SI ratings of Type A could almost be completely predicted by scores on the Clinical Rating factor. For both sexes, content dimensions of the SI related to other measures of Type A and to Type A-consistent traits, whereas the Clinical Rating factor was only modestly associated with such traits. Moreover, sex differences were observed in the trait constellations composing SI and JAS definitions of Type A. These results suggest that a discrepancy exists between conceptual and operational definitions of the Type A pattern.

Dietary assessment instruments for preschool children: reliability of parental responses to the 24-hour recall and a food frequency questionnaire.

Nutrient intakes of preschool-age children were assessed with a 24-hour dietary recall and a 3-month food frequency questionnaire (FFQ). Parents of 55 preschoolers (mean age = 4.25 +/- 0.59 years) completed the recall and FFQ on two occasions 1 week apart. The recalls and FFQs were analyzed for energy, cholesterol, protein, total carbohydrate, calcium, sodium, potassium, and saturated, polyunsaturated, and monounsaturated fats; each nutrient was expressed as raw value, value per kg body weight, and value per 1,000 kcal. Test-retest reliability estimates for the 24-hour recall indicated significant variability in reported total energy intake, but stable reports of intake were observed for one or all units of expression for polyunsaturated fats, cholesterol, protein, total carbohydrate, calcium, and potassium. The FFQ showed significant positive test-retest reliability estimates for all nutrients for all units of expression. Comparison of the recall and FFQ data showed similar percentages of intakes of energy from fat, carbohydrate, and protein and significant correlations for reported intakes of cholesterol, protein, calcium, and potassium. Comparison of the recall and FFQ data with recall data from a comparable cohort showed lower reported intakes for our sample, with the exception of protein, carbohydrate, calcium, and potassium. If validation studies are successful, the FFQ may be useful in epidemiological studies of preschoolers' intakes over extended periods. The recall may prove to be a useful tool in the assessment of day-to-day variations in macronutrient intakes.

Validation of a heart rate monitor with children in laboratory and field settings.

The validity of the Sport Tester PE 3000 portable heart rate monitor for use with children was assessed by comparing readings with simultaneously recorded ECG heart rates in three studies, two using laboratory tasks and one in a field setting. Study 1 examined the validity of the Sport Tester with a sample of ten 10-yr-olds performing a cycle ergometer exercise task. Sport Tester readings taken during three 3-min exercise loads were correlated from 0.97 to 0.99 with simultaneously recorded ECG heart rates. In study 2, 23 children from 4 to 6 yr of age performed treadmill exercise for three 1-min intervals while Sport Tester and ECG heart rate data were obtained. Correlations for the three intervals ranged from 0.94 to 0.99. Study 3 obtained ECG and Sport Tester heart rates in 14 7- to 9-yr-olds who engaged in 3-min periods of standing, walking, jogging, throwing a ball, batting a ball, and playing on a jungle gym. Significant correlations of at least 0.98 between Sport Tester and ECG heart rates were obtained during all six activities. For all three studies, standard errors of estimate were low, ranging from 1.1 to 3.7 beats.min-1. For individuals, maximum differences ranged from 0 to 12.4 beats.min-1. Collectively, these findings indicate that the Sport Tester provides valid readings of young children's heart rates across a wide range of exercise involving upper- and lower-body movements.

Family history of hypertension and cardiovascular reactivity to forehead cold stimulation in black male children.

The effects of family history of hypertension on cardiovascular reactivity to forehead cold stimulation was examined in 16 black males from 11 to 14 years of age. Measures of blood pressure, heart rate, cardiac output, stroke volume and systemic vascular resistance were obtained during baseline, forehead cold stimulation and recovery phases. Diastolic blood pressure and systemic vascular resistance increases to the forehead cold stimulation were significantly greater in subjects with positive family histories of hypertension than in those with negative family histories. These findings are compared with the results of adult studies and discussed in terms of racial differences in alpha and beta-adrenergically mediated cardiovascular reactivity.

Individual and cross-spouse correlations of perceptions of family functioning, blood pressure and dimensions of anger.

Individual and cross-spouse correlations of perceptions of family cohesiveness, emotional expressiveness and conflict with blood pressure and five dimensions of anger were examined in 85 couples. Wives' perceptions of family cohesiveness were negatively related to their diastolic blood pressure, whereas husbands' perceptions of all three dimensions of family functioning were unrelated to their blood pressure. Cross-spouse correlations showed that both husbands' and wives' perceptions of emotional expressiveness were negatively related to their spouses' blood pressure, and husbands' perceived family cohesiveness was negatively related to wives' diastolic blood pressure. Individual and cross-spouse correlations with dimensions of anger were also obtained. These findings are discussed in terms of prior cross-spouse correlational research, and gender differences in the pattern of relationships are discussed.

Humoral permeability factors in the nephrotic syndrome: a compendium and prospectus.

The concept that increased glomerular albumin permeability in steroid-resistant nephrotic syndrome is induced by circulating humoral factors is not new. Zimmermann (1) was among the first to demonstrate that serum from a renal transplant patient with recurrent focal segmental glomerulosclerosis (FSGS) could provoke increased albumin excretion when infused in the aorta of intact rats. Unfortunately, the experiment was not easily reproducible, and the possibility that human serum could induce serum sickness in rats was a serious limitation of the original experiment. We now know that inhibitors of permeability activity are present in both normal human and rat serum (see below), which explains the difficulty in replicating the disease in intact animals. In 1974 Shalhoub (2) theorized that a disordered clone of T lymphocytes, present in both minimal change disease and FSGS, secreted a circulating lymphokine "toxic" to the glomerular barrier. In support of this hypothesis, Koyama et al (3) formed hybridomas from T cells from four patients with minimal change disease and three control subjects. The hybridomas of the patients produced a substance that induced proteinuria when injected intravenously into normal rats. However, the study utilized stimulated and not quiescent T cells, and therefore the relevance to the pathogenesis of FSGS is unknown. Hoyer and colleagues first described recurrence of idiopathic nephrotic syndrome after renal transplantation in 1972 (4). Numerous subsequent reports have established the rate of recurrence as being about 30%. Timely plasmapheresis associated with aggressive immunosuppression resolves the proteinuria and disease progression in a large proportion of cases (5). FSGS not only recurs after renal transplantation, but the diseased kidney can also recover when kept protected from the pathological milieu. Rea et al (6) demonstrated that kidneys from a donor with FSGS transplanted into two uremic recipients were free from proteinuria, and that renal function was normal after one year. Ethical and legal considerations aside, recurrence of FSGS after transplantation is strong evidence supporting the role of a humoral factor in the pathogenesis of the disease.

The relationship between anger-coping styles and lifestyle behaviors in teenagers.

PURPOSE: To investigate the relationship between anger-coping styles (anger expression and anger suppression) and lifestyle behaviors (physical activity and consumption of alcohol, cigarettes, and caffeine) in adolescents. METHODS: A sample of 411 adolescents (198 males: 101 white, 97 black; 213 females: 101 white, 112 black) aged 13-20 years (mean age 15.6 years) completed the Anger Expression Scale and brief self-report questionnaires assessing physical activity (weekly amount, comparison with peers) and consumption of alcohol (frequency and amount over the past 2 weeks), cigarettes (amount over past 2 weeks), and caffeine (from coffee and soda over past week). RESULTS: Correlational and Chi-square analyses showed teenagers high in anger suppression reported consuming alcohol more frequently, spending fewer hours per week in aerobic activity, and being less physically active than their peers. Teenagers high in anger expression reported consuming more caffeinated soda and coffee. CONCLUSIONS: Results suggest that excessive anger suppression or expression may be associated with an imprudent lifestyle relatively early in life.

Consistency of children's hemodynamic responses to laboratory stressors.

Hemodynamic responses to three standard laboratory stressors (i.e., postural change, video game, forehead cold) were evaluated in 341 children (170 males) who had a mean age of 11.2 +/- 2.6 years. Inter-task consistency was evaluated for five physiological parameters for all stressor pairings. Evidence of consistency in systolic and diastolic blood pressure, heart rate, cardiac output, and total peripheral resistance responses to all three stressors was observed. The highest and most consistent correlations across stressors were observed for cardiac output and total peripheral resistance responses. The need for clarification in the categorization of laboratory stressors used to measure reactivity is discussed.

Cardiovascular (CV) responsivity and recovery to acute stress and future CV functioning in youth with family histories of CV disease: a 4-year longitudinal study.

Blood pressure (BP) and heart rate (HR) data obtained during supine rest, in response to and recovery from four laboratory stressors in a baseline year were used to predict supine resting BP and HR values obtained during each of four consecutive annual follow-up evaluations. Subjects were 385 normotensive youth [95 African American (AA) males, 106 AA females, 92 European American (EA) males, 92 EA females] (mean age 12.7+/-2.6 at baseline year) with a positive family history of cardiovascular disease (CVD). During the baseline evaluation subjects were presented with four laboratory stressors (namely, postural change, video game challenge, social competence interview, and parent--child conflict discussion). The BP and HR values taken during each of the laboratory stressors and during the post stressor recovery periods were converted to z-scores which were averaged to yield aggregate measures for systolic and diastolic BP and HR responsivity and recovery. The data obtained during the baseline evaluation were subsequently used to predict the follow-up values of supine resting BP and HR. The prediction models were fairly consistent across each of the 4 follow-up years. Responsivity or recovery accounted for up to 6% of the total variance after accounting for baseline values. Within the prediction models responsivity or recovery accounted for 4--56% of the variance. The predictive value of the derived models did not decline from one annual evaluation to the next over the length of the study. CV recovery may supplement resting and responsivity in the prediction of future development of CVD

Temporal stability of children's cardiovascular (CV) reactivity: role of ethnicity, gender and family history of myocardial infarction.

Cardiovascular responses to three laboratory stressors (i.e., postural change, treadmill exercise, forehead cold) were evaluated in 106 children (72 Whites, 34 Blacks) who varied in family history (FH) of early myocardial infarction (53 positive FH, 53 negative FH). Subjects were evaluated on two occasions separated by one year. In general, regardless of ethnicity, gender or FH, stability of resting blood pressures (BP) and heart rates were comparable to existing data. Resting cardiac output (CO) and total peripheral resistance (TPR) stability estimates were comparable or higher than the BP estimates across groups except for the +FH subjects and males who showed poor stability to one or both parameters. Moderate stability was observed for all parameters in response to forehead cold and low to moderate stability was observed in HR, CO, and TPR postural change reactivity. The only consistent pattern of significant differences in stability estimates involved ethnicity. African-Americans exhibited significantly higher coefficients compared to Whites in TPR at rest and during postural change and forehead cold. Reasons for the low to moderate resting and reactivity stability estimates are discussed.