Sequential contribution of L- and P-selectin to leukocyte rolling in vivo.

Leukocyte recruitment into inflammatory sites is initiated by a reversible transient adhesive contact with the endothelium called leukocyte rolling, which is thought to be mediated by the selectin family of adhesion molecules. Selectin-mediated rolling precedes inflammatory cell emigration, which is significantly impaired in both P- and L-selectin gene-deficient mice. We report here that approximately 13% of all leukocytes passing venules of the cremaster muscle of wild-type mice roll along the endothelium at < 20 min after surgical dissection. Rolling leukocyte flux fraction reaches a maximum of 28% at 40-60 min and returns to 13% at 80-120 min. In P-selectin-deficient mice, rolling is absent initially and reaches 5% at 80-120 min. Rolling flux fraction in L-selectin-deficient mice is similar to wild type initially and declines to 5% at 80-120 min. In both wild-type and L-selectin-deficient mice, initial leukocyte rolling (0-60 min) is completely blocked by the P-selectin monoclonal antibody (mAb) RB40.34, but unaffected by L-selectin mAb MEL-14. Conversely, rolling at later time points (60-120 min) is inhibited by mAb MEL-14 but not by mAb RB40.34. After treatment with tumor necrosis factor (TNF)-alpha for 2 h, approximately 24% of all passing leukocytes roll in cremaster venules of wild-type and P-selectin gene-deficient mice. Rolling in TNF-alpha-treated mice is unaffected by P-selectin mAb or E-selectin mAb 10E9.6. By contrast, rolling in TNF-alpha-treated P-selectin-deficient mice is completely blocked by L-selectin mAb. These data show that P-selectin is important during the initial induction of leukocyte rolling after tissue trauma. At later time points and in TNF-alpha-treated preparations, rolling is largely L-selectin dependent. Under the conditions tested, we are unable to find evidence for involvement of E-selectin in leukocyte rolling in mice.

Anterior pituitary hypoplasia and dwarfism in mice lacking the dopamine transporter.

Deletion of the dopamine transporter (DAT) results in increased dopaminergic tone, anterior pituitary hypoplasia, dwarfism, and an inability to lactate. DAT elimination alters the spatial distribution and dramatically reduces the numbers of lactotrophs and somatotrophs in the pituitary. Despite having normal circulating levels of growth hormone and prolactin in blood, hypoplastic glands from DAT-/- mice fail to respond to secretagog stimulation. The effects of DAT deletion on pituitary function result from elevated DA levels that down-regulate the lactotroph D2 DA receptors and depress hypothalamic growth hormone-releasing hormone content. These results reveal an unexpected and important role or DA in the control of developmental events in the pituitary gland and assign a critical role for hypothalamic DA reuptake in regulating these events.

Regulation of membrane trafficking by a novel Cdc42-related protein in Caenorhabditis elegans epithelial cells.

Rho GTPases are mainly known for their implication in cytoskeleton remodeling. They have also been recently shown to regulate various aspects of membrane trafficking. Here, we report the identification and the characterization of a novel Caenorhabditis elegans Cdc42-related protein, CRP-1, that shows atypical enzymatic characteristics in vitro. Expression in mouse fibroblasts revealed that, in contrast with CDC-42, CRP-1 was unable to reorganize the actin cytoskeleton and mainly localized to trans-Golgi network and recycling endosomes. This subcellular localization, as well as its expression profile restricted to a subset of epithelial-like cells in C. elegans, suggested a potential function for this protein in polarized membrane trafficking. Consistent with this hypothesis, alteration of CRP-1 expression affected the apical trafficking of CHE-14 in vulval and rectal epithelial cells and sphingolipids (C(6)-NBD-ceramide) uptake and/or trafficking in intestinal cells. However, it did not affect basolateral trafficking of myotactin in the pharynx and the targeting of IFB-2 and AJM-1, two cytosolic apical markers of intestine epithelial cells. Hence, our data demonstrate a function for CRP-1 in the regulation of membrane trafficking in a subset of cells with epithelial characteristics.

P-selectin/ICAM-1 double mutant mice: acute emigration of neutrophils into the peritoneum is completely absent but is normal into pulmonary alveoli.

Neutrophil emigration during an inflammatory response is mediated through interactions between adhesion molecules on endothelial cells and neutrophils. P-Selectin mediates rolling or slowing of neutrophils, while intercellular adhesion molecule-1 (ICAM-1) contributes to the firm adhesion and emigration of neutrophils. Removing the function of either molecule partially prevents neutrophil emigration. To analyze further the role of P-selectin and ICAM-1, we have generated a line of mice with mutations in both of these molecules. While mice with either mutation alone show a 60-70% reduction in acute neutrophil emigration into the peritoneum during Streptococcus pneumoniae-induced peritonitis, double mutant mice show a complete loss of neutrophil emigration. In contrast, neutrophil emigration into the alveolar spaces during acute S. pneumoniae-induced pneumonia is normal in double mutant mice. These data demonstrate organ-specific differences, since emigration into the peritoneum requires both adhesion molecules while emigration into the lung requires neither. In the peritoneum, P-selectin-independent and ICAM-1-independent adhesive mechanisms permit reduced emigration when one of these molecules is deficient, but P-selectin-independent mechanisms cannot lead to ICAM-1-independent firm adhesion and emigration.

The influence of age, alcohol consumption, and body build on gonadal function in men.

Basal plasma levels of testosterone, dihydrotestosterone, estradiol, and gonadotropins and testosterone-binding capacity (percent radioactive testosterone bound to protein) were measured in health carefully screened young (31-44 yr old; n = 44) and older (64-88 yr old; n = 42) male participants in the Normative Aging Study of the V.A. There was no statistically significant effect of age on testosterone [younger group, 4.16 +/- 0.27 (SEM) ng/ml; older group, 4.62 +/- 0.32 (SEM) ng/ml] or the free testosterone index [younger group, 2.05 +/- 0.14 (SEM) ng/ml; older group, 1.76 +/- 0.11 (SEM) ng/ml]. The testosterone-binding capacity was higher in the older group (younger group, 50.10 +/- 1.18% (SEM); older group, 60.10 +/- 1.18% (SEM); P less than 0.001). Of the two products of testosterone metabolism studied, estradiol did not change with age, while dihydrotestosterone was lower [young group, 0.25 +/- 0.02 (SEM) ng/ml; older group, 0.20 +/- 0.01 (SEM) ng/ml; P = 0.03] in the older group. FSH levels were increased among the older men [older group, 92.9 +/- 6.0 (SEM) ng/ml; younger group, 61.1 +/- 5.0 (SEM) ng/ml; P less than 0.001]. LH levels were not significantly influenced by age. There was no effect of level of chronic stable alcohol intake on gonadal function, as estimated by testosterone levels and the free testosterone index. Analysis of the relationship between body build and hormone levels indicated that estradiol levels were highest in gynandromorphic men and lowest in mesomorphic men.

Gonadal hormones modulate 5-hydroxytryptamine2A receptors: emphasis on the rat frontal cortex.

The effect of short-term (Sprague-Dawley rats, two weeks) and long-term ovariectomy (Sprague-Dawley and Fischer rats, three months) on serotonin 5-hydroxytryptamine2A receptors in different regions of the brain and its possible correction with an 17 beta-estradiol treatment (10 micrograms, b.i.d., two weeks) were studied in comparison to intact rats. Saturation binding assays were performed using [3H]ketanserin to estimate 5-hydroxytryptamine2A receptor density and affinity in tissue homogenates of frontal cortex of Fischer rats and quantitative autoradiography was performed to evaluate receptor specific binding in frontoparietal cortex, nucleus accumbens, striatum and dorsal raphe nucleus of Fischer rats, and in frontal cortex of the two strains of rats. Messenger RNA levels of 5-hydroxytryptamine2A receptors were measured by in situ hybridization in frontal cortex of the two strains of rats. An overall decrease of 5-hydroxytryptamine2A receptor densities was found in all the brain regions of ovariectomized Fischer rats assayed, and this could be restored towards control levels by estradiol treatment. No change in the 5-hydroxytryptamine2A receptor affinity was measured in the frontal cortex. A similar pattern of changes was observed for the messenger RNA levels encoding the 5-hydroxytryptamine2A receptors and receptor density, suggesting the implication of a genomic mechanism. Experiments in Sprague-Dawley rats confirmed and extended the results obtained with Fischer rats. By analogy, in humans, this 5-hydroxytryptamine2A receptor modulation may underlie the mood and movement disorders associated with menopause.

Ovariectomy and estradiol treatment affect the dopamine transporter and its gene expression in the rat brain.

The impact of gonadal hormone withdrawal and estrogen therapy was investigated on the rat dopamine transporter (DAT). Short-term ovariectomized (ST-OVX, 2 weeks) and long-term ovariectomized (LT-OVX, 3 months) rats were treated or not with 17beta-estradiol (E2) for 2 weeks. DAT mRNA expression was measured by in situ hybridization in the substantia nigra pars compacta (SNc) for the nigrostriatal pathway and the ventral tegmental area (VTA) for the mesolimbic pathway whereas DAT levels were assessed by [3H]GBR-12935 autoradiography, respectively, in the striatum and the nucleus accumbens. Ovariectomy produced a time-dependent decrease of the DAT density in the striatum and the nucleus accumbens and the E2 treatment did not significantly restore these DAT levels. Neither ST-OVX nor E2 treatment of the ST-OVX animals altered the DAT mRNA expression in the SNc and the VTA. However, LT-OVX animals showed increased DAT mRNA levels in these regions. E2 treatment of LT-OVX animals partially restored DAT mRNA levels in the SNc and left these levels unchanged in the VTA. These opposite variations induced by OVX on the DAT density and their mRNA levels suggest the involvement of non-genomic mechanisms, such as post-transcriptional events and/or membrane effects. Altered neurotransmission following gonadal hormone withdrawal may contribute to CNS disorders occurring at menopause in predisposed women. Ovariectomized rats constitute a useful model to study the changes in neurotransmitters balance occurring after menopause.

Sar1-p-benzoylphenylalanine-angiotensin, a new photoaffinity probe for selective labeling of the type 2 angiotensin receptor.

Previous photoaffinity labeling of angiotensin II (Ang) receptors with azidophenylalanine containing Ang analogs produced high yield labeling of a 60 kDa protein on bovine adrenocortical membranes. This preparation is mostly enriched in the type 1 Ang receptor (AT1) and AT1 selective ligands (L158,809) totally prevented labeling, therefore confirming the AT1 nature of the labeled protein. Our attempt to photolabel the type 2 Ang receptor (AT2) of human myometrium with [Sar1,D-Phe(N3)8]Ang was unsuccessful, revealing a high degree of photolabeling selectivity. An Ang analog, [Sar1,Bpa8]Ang (or BpaAng) was prepared containing the photosensitive amino acid p-benzoylphenylalanine (Bpa). This compound was a specific but non-competitive Ang antagonist on rabbit aorta with a pA2 of 8.5. It displayed good binding affinities for bovine adrenocortical membranes (Kd = 6.5 nM), a predominantly AT1 preparation, and for human myometrium membranes (Kd = 0.39 nM), a predominantly AT2 preparation. Photolabeling experiments with iodinated BpaAng showed that AT1 was not covalently labeled whereas AT2 was covalently labeled with high yield. Labeling specificity was verified with the AT2-selective ligand PD123319 and with the AT1-selective antagonist L158,809. Our results indicate that 125I-BpaAng is exclusively labeling AT2 sites. This compound should be a useful tool for further biochemical characterization of the AT2 binding site.

Construct validation of optimism and pessimism in older men: findings from the normative aging study.

Validation of Scheier and Carver's (1985) Life Orientation Test (LOT) has identified associations between bipolar optimism and several external constructs. However, optimism and pessimism may be not bipolar, but rather separate constructs. Furthermore, these constructs may be indistinguishable from personality traits, such as neuroticism and extraversion. This study examined the associations of separate optimism and pessimism measures with self-reports of hassles, psychological symptoms, and illness severity, controlling for personality. Ss were 1,192 men from the Normative Aging Study. Findings suggest that optimism and pessimism are separate and that their relations to external criteria remain, although attenuated, when neuroticism and extraversion are controlled.

Does emotionality predict stress? Findings from the normative aging study.

Investigated whether emotionality, assessed in 1975, predicted the reporting of both objective stress (life events) and subjective stress (hassles) 10 years later, and how emotionality affected the relation between both objective and subjective stress and mental health. The sample consisted of 1,159 older men, participants in the Normative Aging Study. Path analysis revealed that the reporting of stress was confounded with personality: Individuals higher in emotionality reported both more life events and more hassles. Furthermore, individuals higher in emotionality exhibited slightly higher levels of symptoms under stress than did individuals lower in emotionality. Nonetheless, both stress measures contributed independent variance to the prediction of psychological symptoms, even controlling for prior levels of emotionality. Implications for the assessment of stress are discussed.

Mental health differences among retirees and workers: findings from the Normative Aging Study.

Researchers during the past decade have found little effect of retirement on physical health. However, retirement entails a number of losses, and its effect on mental health, as measured by the prevalence of psychological symptoms, is unclear. We examined psychological symptoms in a sample of 1,513 older men, participants in the Normative Aging Study, using the SCL-90-R (Derogatis, 1983). Analyses of variance indicated that retirees reported more psychological symptoms than did workers, even after controlling for physical health status. Exploratory analyses examining the circumstances of retirement found no effects for length of retirement or part-time employment, but did find effects for the timing of retirement. Both early and late retirees reported more psychological symptoms. Late workers (aged 66 and older) reported the fewest symptoms. Reasons for these findings are discussed.

Longitudinal findings from the normative aging study: 1. Does mental health change with age?

Longitudinal changes in self-reported symptoms were investigated using the Cornell Medical Index (CMI) in a sample of 2,041 men. The average man completed 5 CMIs (range = 2-8) over 17 (range = 2-25) years. Using a two-stage growth model, we first regressed symptoms against time on study for each man. The average slope of physical symptoms showed a moderate increase over time, but the average slope of psychological symptoms showed little change. Next, individual differences in change were examined using age at entry as an explanatory variable. Age accounted for 50% of the variance in physical symptoms at entry and 7% of the variance in slopes, but explained neither baseline level nor change in psychological symptoms. Rather, a U-shaped curve described the relation between age and psychological symptom change. These results, in concert with a developmental perspective, may help reconcile conflicting findings on aging and mental health.

Differences in social support among retirees and workers: findings from the Normative Aging Study.

Cross-sectional differences between retirees and workers in the importance of coworkers as a source of support, as well as in general quantitative support (social networks and frequency of interaction) and qualitative support (confidants and the perceived reliability of support) were examined. The sample consisted of 1,513 older men (mean age = 61), participants in the Normative Aging Study. Half (56%) were working, and the rest were retired. Slightly fewer retirees than workers reported coworker friends, especially among those who were long-term retirees or who did not work at all in retirement. Whereas similar findings were seen with quantitative support, workers and retirees reported nearly identical levels of qualitative support. However, retirees almost never discussed personal problems with former coworkers. The relevance of these findings for the convoy construct is discussed.

Personality and aging: a study of the MMPI-2 among older men.

We examined whether separate norms for older men are necessary for the revised Minnesota Multiphasic Personality Inventory (MMPI-2). Scores from 1,459 men in the Normative Aging Study (NAS) (age: M = 61.27, SD = 8.37) were contrasted with those from 1,138 men from the MMPI Restandardization Study (age: M = 41.71, SD = 15.32). Results showed that scores on the MMPI-2 validity, clinical, and content scales for the NAS men were highly similar to those from the MMPI-2 Restandardization sample. There were also few differences between the two groups at the item level. Within-sample analyses revealed some differences between age groups. However, the magnitudes of these differences were small and may represent the single or combined effects of cohort factors and age-related changes in physical health status rather than age-related changes in psychopathology per se. We concluded that special, age-related norms for the MMPI-2 are not needed for older men.

The effect of retirement on somatic complaints.

This study investigated whether the level of somatic complaint increases as men make the supposedly stressful passage from work to retirement. For 171 retirees, pre- to post-retirement changes in the sum of selected somatic complaints from the Cornell Medical Index were compared with changes over a comparable time interval (3-5 yr) in 386 age peers who continued to work. Although there was slightly increased frequency of somatic complaint over time, eventual retirees did not differ from continuing workers in the amount and direction of change. Among retirees only, change from pre-retirement complaint levels did not vary by retirement age, length of time retired, part-time work in retirement, former occupational level or marital status, nor was it affected by the circumstance of compulsory retirement. These results, which are based on a continuous measure of health status, confirm previous findings from categorical data which conclude that retirement's effect on health is negligible.

Social contexts and motives for drinking in men.

Factor analysis identified nine distinct contextual-motivational factors for drinking in 1517 healthy men. These factors were significant predictors of level of alcohol consumption and discriminated subjects reporting drinking problems from those reporting no problems.

The MAC scale in a normal population: the meaning of "false positives".

The MAC scale has been very successful in identifying alcoholics and, in studies of clinical populations, is often considered a test for predisposition to alcoholism. MacAndrew, however, holds that the MAC scale assesses a more general personality trait characterized by sociability, boldness, rebelliousness and pleasure seeking. The present study examines the distribution of MAC scale scores in a normal population and tests for correlates of high MAC scores other than alcohol-related problems (e.g., arrest history). The sample consisted of 1,117 men, participants in the Normative Aging Study (mean age = 61.6). As expected, heavier drinkers and problem drinkers reported significantly higher MAC scale scores than did lighter and nonproblem drinkers. However, arrestees without drinking problems had MAC scale scores nearly identical to those of problem drinkers without arrest histories (23.19 and 23.42, respectively). Further, 36% of the sample without problem drinking or arrest histories had MAC scale scores of 24 or above, the clinical indicator of alcoholism, and more than 32% of these had scores above 27. In the entire sample, of the 152 men who had MAC scores above 27, 71% had no problems, either with arrest or drinking. Results are interpreted as supporting MacAndrew's interpretation of the meaning of the MAC scale as a general personality measure rather than a specific alcoholism instrument.

Retrospective analysis in smoking cessation research.

The use of retrospective data in smoking research represents possible sources of bias resulting from: (1) baseline incomparability of current, former, and non-smokers, (2) instability of parameters under investigation, and (3) difficulties in recall. A self-report measure of smoking motives was employed in a longitudinal design to examine bias associated with retrospective data analysis. Thirty-six pairs of former smokers and recidivists (smokers who had tried to quit) were matched on age and amount smoked. Although initial responses for both groups were comparable, former smokers were significantly higher on smoking motive factors after quitting compared to recidivists who stayed essentially the same. The findings suggest that mean differences in smoking motives between recidivists and former smokers assessed cross-sectionally may not accurately reflect the magnitude of the differences that existed when both groups were smoking.

Does the source of support matter for different health outcomes? Findings from the Normative Aging Study.

OBJECTIVES: This study investigated the differential relationships between different types and sources of social support and physical and mental health. METHODS: Using data from the Normative Aging Study, 1,386 older men (median age = 62.7 years) were categorized into four groups separately for frequency of interaction with networks and perceived support. RESULTS: More than half the sample reported high levels of support from both sources. One-way ANOVAs revealed that those with high perceived support from both sources reported better physical health and fewer depressive symptoms than those with low support from both sources or high support from family alone. Similarly, those with high perceived support from both sources had lower levels of depressive symptoms than those with low support from both sources, but frequency of contact was unrelated to physical health. DISCUSSION: In general, those with high support from both family and friends reported the highest level of well-being.

Age, smoking inhalation, and pulmonary function.

This study examined the relative effects of age and smoking on pulmonary function. Smoking was measured by six smoking variables, taken singly and as a composite. Subjects were 1,516 male participants in the Normative Aging Study. A stepwise multiple regression with vital capacity (VC) and forced expiratory volume at one second (FEV1.0) as the criteria accounted for 24.4% and 28.3% of the variance, respectively. Two-way analyses of variance showed that the age decline in pulmonary function was substantially greater for high inhalers than it was for low inhalers or nonsmokers. Age and the inhalation index were also noticeably and independently related to a decline in pulmonary function.