Persistently low natural killer cell activity in normal adults: immunological, hormonal and mood correlates

This current study is concerned with the identification of the incidence of persistently low natural killer (NK) cell activity in an essentially healthy population of young adults, and the characterization of the psychological profile associated with this pattern of NK activity. Eighty-eight individuals, between the ages of 18 and 44 years, were accrued to this project. Subjects were tested serially, at baseline induction into the study, and at 2 and 4 weeks follow-up. Baseline assessment included a complete physical examination and laboratory work-up in order to exclude anyone ill at the time of enrollment, and to have complete physical status information on all subjects at the time of accrual. NK activity, urinary catecholamine levels and psychological status were also assessed at baseline, and at the specified follow-up intervals. Individuals with low NK activity were operationally defined as those having NK function either below the group mean, or below the group lowest quartile, at baseline and at two follow-up assessments. Results showed that in both univariate analyses, as well as in logistic regression models, age and the perception of environmental stressors or 'hassles' predicted persistently low NK activity. Younger subjects, who perceived environmental events to which they were exposed as more serious in nature, were more likely to exhibit a persistently low NK profile over time than older individuals who perceived daily events as less important to them.     

The hypogonadotropic state of the prepubertal male rhesus monkey (Macaca mulatta) is not associated with a decrease in hypothalamic gonadotropin-releasing hormone content

Hypothalamic contents of gonadotropin-releasing hormone (GnRH) in neonatally orchidectomized infant, juvenile, and adult monkeys were measured by a radioimmunoassay (RIA) and by an in vivo bioassay that utilized luteinizing hormone (LH) secretion in estrogen- and progesterone-treated ovariectomized rats. The results of the bioassay provided no evidence to suggest that hypothalamic GnRH content in juvenile monkeys (mean = 83 ng/hypothalamus; n = 3) was less than that in infants (mean = 54 ng/hypothalamus; n = 4) and adults (mean = 36 ng/hypothalamus; n = 3). A similar developmental pattern in hypothalamic GnRH content was also observed when the decapeptide was measured by RIA. In striking contrast to the maintenance of hypothalamic GnRH content throughout postnatal development, pituitary gonadotropin contents and serum gonadotropin concentrations were markedly reduced in juvenile monkeys.     

Cellular senescence involves stochastic processes causing loss of expression of differentiated function genes: visualization by in situ hybridization for steroid 17 alpha-hydroxylase in bovine adrenocortical cells

When grown for long periods in culture, bovine adrenocortical cells lose the expression of a differentiated function gene, steroid 17 alpha-hydroxylase. Previously, we documented a decline in 17 alpha-hydroxylase mRNA with increasing culture passage level after induction with cyclic AMP (P. J. Hornsby et al., 1987, Proc. Natl. Acad. Sci. USA 84, 1580). We used in situ hybridization to investigate the loss of expression of this gene during cellular senescence at an individual cell level. In primary cultures, cells were uniformly positive for hybridization with cDNA for 17 alpha-hydroxylase after cyclic AMP induction. After two passages, cultures comprised a mixture of hybridizing and nonhybridizing cells. Cells appeared either to hybridize at a level comparable to that in primary cultures or to be nonhybridizing. When in situ hybridization was combined with immunofluorescence, cells positive for immunofluorescence were also positive for hybridization. Senescing mass cultures showed decreasing numbers of positive cells, and after 30 passages cultures comprised entirely nonhybridizing cells. Thus, the previously observed decline in overall 17 alpha-hydroxylase mRNA levels results from a decline in the fraction of expressing cells in the culture, and the rate of loss of expressing cells is in agreement with the rate of loss of total 17 alpha-hydroxylase mRNA. Primary clones, even when isolated at an early stage of clonal expansion, had mixtures of subclones of hybridizing and nonhybridizing cells. On recloning, hybridizing subclones usually produced uniformly nonhybridizing sub-subclones. Some subclones within primary clones had a morphology associated with replicative senescence (flattened cells with sparse intercellular contacts), yet had high numbers of hybridizing cells. We conclude that, in both mass and clonal populations, cells initially expressing 17 alpha-hydroxylase rapidly give rise to clones of nonexpressing cells. Such cells are continually derived by a stochastic process from cells originally expressing the gene.     

Correlates of male mating success in a lekking bird with male-male cooperation

Correlates of male mating success were examined in a population of long-tailed manakins, Chiroxiphia linearis, that included 270 colour-banded individuals. Long-tailed manakins have a lek mating system and male-male cooperation in courtship display. Multivariate analysis of behavioural variables indicated that female visitation correlated with the number of unison ‘toledo’ calls given by male partners. Given a female vist, copulatory success was correlated with the ‘butterfly’ display component of the dual-male dance. Both ‘toledo’ output and dance display differed significantly between perch-zones. Only six to eight partnerships in a local population of as many as 55 males per season performed call displays at a level (75–335 toledos per h) that was correlated with any female visitation. Data on crown plumage of female visitors suggested that younger females may have been less discriminating than were older females. The relationship between variance in mating success and the evolution of cooperative male display is discussed.     

Expression of human plasminogen cDNA in a baculovirus vector-infected insect cell system

A cDNA that encodes the human plasminogen (HPg) amino acid sequence has been inserted adjacent to the polyhedrin promoter in the genome of the baculovirus, Autographa californica nuclear polyhedrosis virus, which was then used to infect cultured cells of the farm armyworm, Spodoptera frugiperda. Under the conditions of cell growth employed, recombinant (rec)-HPg was secreted into the medium after 24 h postinfection (p.i.), at which point virtually no rec-HPg antigen remained inside the cells. At 48 h p.i., a maximal level of intact rec-HPg was present in the medium, which underwent substantial proteolytic digestion after that time. The rec-HPg produced by this expression system possessed a molecular weight equivalent to that of plasma [Glu1]-plasminogen. In addition, the rec-HPg adsorbed to Sepharose-lysine, and was eluted with epsilon-aminocaproic acid (EACA). The recombinant protein also interacted with polyclonal antibodies generated to plasma HPg, as well as with a monoclonal antibody directed against a distinct region (kringle 1-3) of the plasma HPg molecule. Finally, the insect-expressed rec-HPg was activatable to plasmin (HPm) by urokinase. The results demonstrate that this expression system produces a full-length functional single-chain rec-HPg, which can be isolated intact from the culture medium, with some consideration for the temporal events that occur in secretion and longer-term degradation of the protein. The fact that this rec-HPg was converted to HPm with a plasminogen activator, and that it interacted with anti-plasma HPg polyclonal and monoclonal antibodies, as well as with the ligand, EACA, indicates that the molecule retains many of its important functional properties and is folded in an integral manner.     

Endo-exonuclease of Aspergillus nidulans

Endo-exonuclease (EE) has been found in both active and inactive, but trypsin-activatable, forms in Aspergillus nidulans. Active EE was present mainly in nuclei, mitochondria, and vacuoles, while trypsin-activatable EE was mainly in the cytosol. The active form accounts for over 90% of the neutral deoxyribonuclease activity extracted from mycelia. A single strand (ss) DNA-binding EE associated with a 28 kilodalton (kDa) polypeptide was partially purified and characterized. It was found to closely resemble, in size and enzymological properties, the ss-DNA-binding EE previously purified from Neurospora crassa. Aspergillus nidulans EE was also found to be immunochemically related to the N. crassa EE and, like that enzyme, was probably derived from a polypeptide of 90 kDa or larger through proteolysis during extraction and purification. It had divalent metal ion-dependent (Mg2+, Mn2+, or Zn2+) activity on both DNA and RNA, which ultimately produced small 5'-P-terminated oligonucleotides. The nuclease activity was mixed endo- and exo-nucleolytic with ss-DNA as substrate, but largely exonucleolytic with double strand (ds) DNA. Superhelical phi X-174 DNA was nicked by EE to form relaxed circular and then linear ds-DNA, which was rapidly degraded to shorter fragments. Linearized pBR322 DNA was extensively nicked internally under conditions where there was relatively low exonuclease activity, but this nicking required that 5'-P-termini be present on the linear ds-DNA. The levels of active EE found in extracts of two recombination-deficient mutants of A. nidulans, uvsC and uvsE, dit not differ significantly from those in extracts of the wild type.     

Differential methylation of the hypervariable locus DXS255 on active and inactive X chromosomes correlates with the expression of a human X-linked gene

Consistent differences in methylation of particular cytosine residues in the DNA of active and inactive X chromosomes can be used for rapid, direct analysis of X-inactivation patterns in different female tissues. We have studied methylation of the highly polymorphic DXS255 locus in tissues from patients with deficiency of the E1 alpha subunit of the pyruvate dehydrogenase complex in whom the results can be correlated directly with total enzyme activity, levels of immunoreactive protein, and patterns of cell mosaicism. The results confirm that methylation of the DXS255 locus correlates with X-chromosome expression. In patients and normal controls, the pattern of X inactivation varied widely from tissue to tissue and often deviated markedly from a 50:50 proportion. These deviations are likely to reflect small numbers of tissue-specific stem cells at the time of random X inactivation and cannot be taken alone as evidence for selection or "nonrandom" inactivation.     

Dynamic changes in optic fiber terminal arbors lead to retinotopic map formation: an in vivo confocal microscopic study

Dynamic remodeling of retinal ganglion cell terminal arbors has been proposed to contribute to formation of the topographically ordered retinotectal projection. To test this directly, the growth of individual terminal arbors was observed in live X. laevis tadpoles using a confocal microscope to visualize their complex three-dimensional structure. During initial development, nasal and temporal retinal arbors covered overlapping tectal areas. Despite subsequent remodeling, the dimensions and positions of the temporal arbors remained relatively stable. In contrast, the nasal arbors grew caudally, as they extended caudal branches and retracted rostral branches. These results suggest that differences in the remodeling of the nasal and temporal arbors lead to the emergence of retino-topography along the rostrocaudal axis of the tectum. All the terminal arbors were dynamic, including those with stable dimensions, suggesting that continual remodeling of arbors may be a universal feature of neuronal projections.     

Tired,weak, or in need of rest: fatigue among general practice attenders.

OBJECTIVES--To determine the prevalence and associations of symptoms of fatigue. DESIGN--Questionnaire survey. SETTING--London general practice. PARTICIPANTS--611 General practice attenders. MAIN OUTCOME MEASURES--Scores on a fatigue questionnaire and reasons given for fatigue. RESULTS--10.2% Of men (17/167) and 10.6% of women (47/444) had substantial fatigue for one month or more. Age, occupation, and marital status exerted minor effects. Subjects attributed fatigue equally to physical and non-physical causes. Physical ill health, including viral infection, was associated with more severe fatigue. Women rather than men blamed family responsibilities for their fatigue. The profile of persistent fatigue did not differ from that of short duration. Only one person met criteria for the chronic fatigue syndrome. CONCLUSIONS--Fatigue is a common complaint among general practice attenders and can be severe. Patients may attribute this to physical, psychological, and social stress.