The Reliability of Arousal Threshold During Sleep

Thirty-five subjects from two independent studies were awakened at EEG-defined periods during the night with 1000 Hz ascending tone series. Awakenings were made five to eight times per night during stage 2, stage 4, or REM sleep over a series of nights in good and poor sleepers. Reliability was assessed within stage, within night, between stages, and between nights. Good and poor sleepers did not differ in either depth of sleep or reliability of arousal threshold and were thus pooled in the analyses. From night to night, the most consistency was seen in stage 4 (r=.74), although REM reliability (r?₁= .49) and stage 2 reliability (r?₁= .50 and r?₁= .69 in the two respective studies) estimates were also greater than zero. Early sleep onset and morning arousals were more variable. Reliability estimates on arousal thresholds taken within the same night for stage 2 were r= .64 and r?₁= .77 for the two studies and r= .96 for REM. The depth of sleep was not correlated with awake auditory threshold. It was concluded that five or six carefully placed arousals could give a good estimate of an individual's usual arousal threshold.     

Evaluation of Streptococcus pneumoniae Type XIV Opsonins by Phagocytosis-Associated Chemiluminescence and a Bactericidal Assay

The relative roles of serum factors required for opsonization of type XIV Streptococcus pneumoniae were investigated by means of luminol-enhanced chemiluminescence (CL), bactericidal, and immunofluorescence assays employing adult sera containing high (>1,000 ng of antibody nitrogen per ml) or low (<200 ng of antibody nitrogen per ml) antibody concentrations as determined by radioimmunoassay. Specific antibody concentration correlated directly with both total and heat-labile CL activity (P < 0.005) and with the bactericidal index (P < 0.05) at a serum concentration of 10%. The importance of specific antibody as an opsonin was confirmed by the abolition of CL activity and immunoglobulin immunofluorescence observed after absorption of heated sera with type XIV pneumococcal cells and by the dose response in CL and bactericidal activity observed with the addition of immunoglobulin G to hypogammaglobulinemic serum. A role for the classical complement pathway in opsonization was indicated by significantly greater CL integrals for high-antibody sera than for low-antibody sera depleted of factor D and by the bactericidal activity noted for untreated, but not magnesium ethylene glycol-bis(β-aminoethyl ether)-N,N-tetraacetic acid-chelated low-antibody sera. The alternative pathway contributed more than half of the CL activity of both high- and low-antibody sera. However, after magnesium ethylene glycol-bis(β-aminoethyl ether)-N,N-tetraacetic acid chelation, only sera with high antibody concentrations or agammaglobulinemic serum reconstituted with immunoglobulin G with high specific antibody levels supported significant bactericidal activity. Therefore, type-specific antibody and complement promote opsonization of type XIV S. pneumoniae, and this may occur via either complement pathway. These results suggest that CL is a suitable tool to delineate serum factors and their contribution to opsonization, but results must be related to other functional assays.     

The expression patterns of mRNA-encoding stem cell factor, internal stem cell factor and c-kit in the prepubertal and adult porcine ovary

The receptor, c-kit, and its ligand, stem cell factor (SCF), are important regulators of ovarian follicle growth and development. The aim of this study was to identify the sites of expression of mRNA for c-kit and SCF in prepubertal and mature (pregnant and non-pregnant) animals. Ovaries were recovered from prepubertal animals, non-pregnant sows and five sows at approximately 3 months of gestation. Ovine SCF and c-kit DNA were cloned into plasmid vectors to produce RNA probes. Expression of mRNA encoding SCF and c-kit were detected via in situ hybridization. Both mRNA were detected throughout ovaries from all animals. This study provides evidence that the growth-factor complex is required throughout follicle development, and also for continued maintenance of the corpus luteum (CL) in the mature animal. SCF mRNA was localized to the granulosa cell layer and was also extensively expressed in endothelial tissue and throughout the CL. c-kit mRNA was detected in the theca layer, oocytes and also in CL. In conclusion, expression of SCF and c-kit mRNA in granulosa and theca cells, respectively, indicate an important interaction between somatic cells throughout follicle development and that in the mature animal, SCF and c-kit potentially have a role in maintaining progesterone secretion by the CL. The observations of continued expression of SCF and c-kit throughout development suggest that there may be differences in the role of this receptor-ligand complex between large mono- vs. poly ovulatory species, such as the pig.     

Limitations of a simple technique for movement compensation via movement-modified fluence profiles

The delivery of intensity-modulated radiation therapy (IMRT) through the dynamic multileaf collimator (dMLC) technique is well known and recently it has been shown how this can be modified to deliver fluence to a body which is moving in a regular and totally predictable manner. This involves making the leaves 'breath' in tandem with the body motion. This note presents an entirely alternative suggestion in which the leaves do not 'breath' at all but the fluence profile to be mapped to the unbreathing leaf motions is modified so that, when sampled by the specified motion, the actual fluence delivered is close to the desired fluence. Limitations of the concept are discussed.     

Regaining chondrocyte phenotype in thermosensitive gel culture

Chondrocyte tissue engineering continues to be a challenging problem. When chondrocytes are duplicated in vitro, it is imperative to obtain an adequate number of cells of optimal phenotype. A temperature-sensitive polymer gel, a copolymer of poly(N-isopropylacrylamide) and acrylic acid (PNiPAAm-co-Aac), has the ability of gelling at 37 degrees C (the lower critical solution temperature, LCST) or above and liquefying below that temperature (Vernon and Gutowska, Macromol. Symp. 1996;109:155-167). The hypothesis of this study was that chondrocytes could (1) duplicate in the copolymer gel; (2) regain their chondrocyte phenotype; and (3) be easily recovered from the gel by simply lowering the temperature below 37 degrees C. Chondrocytes from adult rabbit scapular cartilage were harvested and cultured in a monolayer culture until confluency (approximately 2 weeks). Next, the cells were harvested and seeded into the copolymer gel and cultured for 2-4 weeks. The phenotype of the cultured cells was then characterized. Two groups of control cultures, monolayer and agarose gel, were used to compare their ability to maintain chondrocyte phenotype. The results showed that chondrocytes isolated from rabbit scapula can re-express chondrocyte phenotype in agarose culture and polymer gel culture but not in monolayer culture. Also, cultured chondrocytes can be easily recovered from polymer gel culture by simply lowering the temperature. This new in vitro method of chondrocyte culture is recommended for chondrocyte propagation and regaining chondrocyte phenotype before cell seeding or transplantation.     

Lassa fever. Effective therapy with ribavirin

In a study of Lassa fever in Sierra Leone, West Africa, we identified two variables associated with a high risk of death, and we evaluated the efficacy of ribavirin and Lassa virus-convalescent plasma for the treatment of Lassa fever. A serum aspartate aminotransferase level greater than or equal to 150 IU per liter at the time of hospital admission was associated with a case-fatality rate of 55 percent (33 of 60). Patients with the same risk factor who were treated for 10 days with intravenous ribavirin, begun within the first 6 days after the onset of fever, had a case-fatality rate of 5 percent (1 of 20) (P = 0.0002 by Fisher's exact test). Patients whose treatment began seven or more days after the onset of fever had a case-fatality rate of 26 percent (11 of 43) (P = 0.01). Viremia with levels greater than or equal to 10(3.6) TCID50 per milliliter on admission was associated with a case-fatality rate of 76 percent (35 of 46). Patients with this risk factor who were treated with intravenous ribavirin within the first six days after onset of fever had a case-fatality rate of 9 percent (1 of 11) (P = 0.006), whereas those treated after seven days or more of illness had a fatality rate of 47 percent (9 of 19) (P = 0.035). Oral ribavirin was also effective in patients at high risk of death. Lassa-convalescent plasma did not significantly reduce mortality in any of the high-risk groups. We conclude that ribavirin is effective in the treatment of Lassa fever and that it should be used at any point in the illness, as well as for postexposure prophylaxis.     

Radioimmunoassay of detirelix ([ N-Ac-D-Nal(2)1,D-p-Cl-Phe2,D-Trp3, D-hArg(Et)6(2),D-Ala10]-luteinizing hormone-releasing hormone) in plasma or serum

A procedure for the radioimmunoassay (RIA) of detirelix in plasma or serum at concentrations as low as 0.15 ng/ml is described. Antiserum was produced by deacetylation of the N-terminus amino groups of detirelix and coupling this analog to bovine serum albumin with a carbodiimide and immunizing rabbits with the resultant conjugate. For RIA, 125I-labeled detirelix was used as the tracer and a double antibody procedure was used to separate the free and bound fractions. No purification of samples was required prior to RIA. Accuracy of the method was assessed by adding known quantities of detirelix to detirelix-free plasma and determining the ratio of measured to added analyte. Linear regression analysis for the concentration range 0.15-150.0 ng/ml yielded a regression equation of y = 0.88 X +1.46 and a correlation coefficient of 0.996. Additional validation was obtained from an in vivo study in which [14C]detirelix was administered to monkeys and plasma clearance profiles were determined by RIA and an HPLC-radiochemical method. The RIA results were in good agreement with those obtained by the HPLC method.     

Evidence for a calcium regulated, bidirectional intronic promoter in the murine TCR V{alpha}1 gene

Previous studies of the TCR chain gene have located promoterelements 5' to the start of the various V genes. The only fullycharacterized enhancer for the entire chain gene (V, J andC genes) has been located {small tilde}3 kb from the 3' endof C. We now report the existence of additional regulatory elementslocated in the introns of several murine V genes (V1, V3 andVB6.2.16). In the case of V1, this element appears to be a promoterwith bidirectional activity that is not T cell specific. Interestingly,upstream of the promoter in the antisense strand, an open readingframe has been found that codes for a small molecular weightprotein ({small tilde}60 amino acids) that contains a prollne-richregion and a tyrosine-isoleucine motif that has homology toIgß (the B29 gene product). A rabbit antiserum madeagainst this sequence has confirmed its existence by Westernblot and immunoprecipitation. Thus this V1 intronic promoterhas the potential not only to induce the formation of a truncatedV1 gene product, but also regulates the expression of a smallmolecular weight protein that may be involved in lymphocyteantigen receptor signaling. The activity of this promoter isregulated by changes in intracellular calcium. In the presenceof ionomycin the promoter is down-regulated in the sense directionand its activity is enhanced in the antisense direction. Thisresult suggests that this promoter can act differentially toproduce two very different gene products. The bidirectionalV1 promoter appears to be the first in the Ig superfamily toinduce potentially functional proteins in both directions.     

Relationship of glucocorticoid suppression of arachidonic acid metabolism to alteration of neutrophil function

The role of arachidonic acid metabolism in glucocorticoid-induced suppression of neutrophil function was investigated. In vivo treatment of cattle with dexamethasone decreased the production of lipoxygenase products of arachidonic acid metabolism (leukotriene B4 [LTB4] and H-5-hydroxy-6,8,11,14-eicosatetraenoic acid [5-HETE]) in neutrophils. We previously reported that in vivo dexamethasone treatment resulted in alteration of in vitro neutrophil oxidative metabolism, iodination, antibody-dependent cell-mediated cytotoxicity (ADCC), and random migration. To determine if the decrease in production of LTB4 and 5-HETE were responsible for the changes in neutrophil function, neutrophils were treated in vitro with inhibitors of the lipoxygenase enzyme (BW755c and nordihydroguaiaretic acid), and their function was evaluated. A decrease in neutrophil oxidative metabolism and iodination was observed, but there was no effect on neutrophil random migration or ADCC. The results suggest that in vivo treatment with glucocorticoids inhibit neutrophil oxidative metabolism and iodination by decreasing the formation of lipoxygenase products of arachidonic acid metabolism but alter neutrophil random migration and ADCC by a mechanism independent of arachidonic acid metabolism.