Brain-derived neurotrophic factor mRNA downregulation produced by social isolation is blocked by intrahippocampal interleukin-1 receptor antagonist.

Manipulations that increase the expression of the pro-inflammatory cytokine interleukin-1beta (IL-1beta) in the hippocampus (e.g. peripheral administration of lipopolysaccharide, i.c.v. glycoprotein 120, social isolation) as well as the intrahippocampal injection of IL-1beta following a learning experience, dramatically impair the memory of that experience if the formation of the memory requires the hippocampus. Here we employed social isolation to further study this phenomenon, as well as its relation to brain-derived neurotrophic factor (BDNF). BDNF was studied because of its well-documented role in the formation of hippocampally based memory. A 6 h period of social isolation immediately after contextual fear conditioning impaired memory for context fear measured 48 h later, and decreased BDNF mRNA in the dentate gyrus and the CA3 region of the hippocampus assessed immediately after the isolation. Moreover, an intrahippocampal injection of the IL-1 receptor antagonist prior to the isolation period prevented both the BDNF downregulation and the memory impairments produced by the isolation. These data suggest that hippocampal-dependent memory impairments induced by elevated levels of brain IL-1beta may occur via an IL-1beta-induced downregulation in hippocampal BDNF.

Characterization of the oligosaccharide structures associated with the cystic fibrosis transmembrane conductance regulator.

The cystic fibrosis transmembrane conductance regulator (CFTR) is a plasma membrane-associated glycoprotein. The protein can exist in three different molecular weight forms of approximately 127, 131, and 160 kDa, representing either nonglycosylated, core glycosylated, or fully mature, complex glycosylated CFTR, respectively. The most common mutation in cystic fibrosis (CF) results in the synthesis of a variant (DeltaF508-CFTR) that is incompletely glycosylated and defective in its trafficking to the cell surface. In this study, we have analyzed the oligosaccharide structures associated with the different forms of recombinant CFTR, by expressing and purifying the channel protein from either mammalian Chinese hamster ovary (CHO) or insect Sf9 cells. Using glycosidases and FACE analysis (fluorophore-assisted carbohydrate electrophoresis) we determined that purified CHO-CFTR contained polylactosaminoglycan (PL) sequences, while Sf9-CFTR had only oligomannosidic saccharides with fucosylation on the innermost GlcNAc. The presence of PL sequences on the recombinant CHO-CFTR is consistent with a normal feature of mammalian processing, since endogenous CFTR isolated from T84 cells displayed a similar pattern of glycosylation. The present study also reports on the use of FACE for the qualitative analysis of small amounts of glycoprotein oligosaccharides released enzymatically.

The Royal London Space Planning: an integration of space analysis and treatment planning: Part I: Assessing the space required to meet treatment objectives.

The Royal London Space Planning process has evolved since 1985 to ensure a disciplined approach to diagnosis and treatment planning and to provide a record to justify treatment decisions for professional accountability. The analysis takes into consideration most aspects of a given malocclusion and aims to quantify the space required in each dental arch to attain the treatment objectives. Space planning also helps determine whether the objectives are likely to be attainable and helps in the planning of treatment mechanics and the control of anchorage. The process of analysis is divided into 2 sections. The first part consists of assessing the original malocclusion according to various component parts, any of which may have an effect on space if altered during treatment. These components are crowding and spacing, occlusal curves, arch width, anteroposterior position of labial segments, mesiodistal angulation, and incisor inclination. The second part of the analysis, which will be published in a separate article, deals with the effect of treatment procedures, such as extractions, tooth-size modifications, distal or mesial molar movements, as well as natural growth, on the space required. Space planning should be regarded only as a useful guide, as many areas of orthodontics-including growth, biological response, and patient compliance-cannot be controlled with total accuracy.

The Royal London Space Planning: an integration of space analysis and treatment planning: Part II: The effect of other treatment procedures on space.

The Royal London Space Planning process is carried out in 2 stages. The first stage, assessing the space required to attain the treatment objectives, was described in Part I of this report, published earlier. In Part II, the process of integrating space analysis with treatment planning continues with consideration of the effects other treatment procedures have on space. These procedures include tooth enlargement or reduction, tooth extraction, the creation of space for prosthetic replacement, and mesial and distal molar movement. The effects of favorable and unfavorable growth are also considered. A brief case report is presented to demonstrate use of the Royal London Space Planning.

How much space is created from expansion or premolar extraction?

The aim of this study was first to investigate the relationship between maxillary arch expansion and change in arch depth (overjet), and secondly to quantify the reduction in maxillary arch depth following extraction of 4\4 with complete space closure. A model of maxillary typodont teeth was constructed to allow expansion and premolar removal. Arch dimensions were recorded using a reflex microscope. A linear relationship was found between arch expansion and reduction of the arch depth. When the premolars were removed, there was a greater reduction in arch depth than the mesio-distal width of these teeth.

The influence of maxillary incisor inclination on arch length.

This ex vivo study was designed to investigate Andrews' hypothesis that there is a space implication when incisors are torqued correctly. A working model was constructed to allow acrylic typodont incisors of varying known values of inclination to be substituted into the model. The arch lengths of the various 'set-ups' were measured using a reflex microscope linked to a PC. In order to quantify the space requirement of clinical relevance for adequate incisor torque, the method was repeated by substituting replicas of patients' 'natural' incisors. For both acrylic and natural incisors it was found that, as the inclination of the teeth increased, there was an increase in all arch lengths, this being greater for the natural incisors. This larger increase for the natural incisors was related not only to their increased size, but was also dependent on the morphology of the incisor. Those incisors which were parallel-sided showed the greatest increase in arch length, whereas the incisors that were relatively triangular in shape showed the smallest increase. When the inclination of an 'average' set of 21/12 is increased by 5 degrees, an increase in the arch length of approximately 1 mm may be expected.

Early Root Response to Meloidogyne incognita in Resistant and Susceptible Alfalfa Cultivars.

The early events of Meloidogyne incognita behavior and associated host responses following root penetration were studied in resistant (cv. Moapa 69) and susceptible (cv. Lahontan) alfalfa. Ten-day-old seedlings of alfalfa cultivars were inoculated with second-stage juveniles (J2) and harvested 12, 24, 48, and 72 hours and 7, 14, and 21 days later. Both cultivars supported similar root penetration and initial J2 migration. By 72 hours after inoculation the majority of J2 were amassed inside the vascular cylinder in roots of susceptible Lahontan, while J2 had not entered the vascular cylinder of resistant Moapa 69 and remained clumped at the root apex. Nematode development progressed normally in Lahontan, but J2 were not observed in Moapa 69 after day 7. The greatest differences between RNA translation products isolated from inoculated and uninoculated roots of Lahanton occurred 72 hours after inoculation. Only minor differences in gene expression were observed between inoculated and uninoculated Moapa 69 roots at 72 hours. Comparison of translation products from inoculated versus mechanically wounded Lahontan roots revealed products that were specific to or enhanced in nematode-infected plants. Moapa 69 appears to possess a type of resistance to M. incognita that does not depend on a conventional hypersensitive response.

Studies on the microheterogeneity and in vitro activity of glycosylated and nonglycosylated recombinant human prolactin separated using a novel purification process.

Recombinant human PRL was produced in a murine C127 cell expression system and purified to greater than 97% homogeneity using anion and cation exchange chromatography. This material was biologically equivalent to pituitary-derived PRL in both an enzyme-linked immunosorbent assay and the Nb2 lymphoma cell proliferation assay. The predominant PRL forms were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting as being 23 and 25 kilodaltons (kDa). These mass values were confirmed by electrospray mass spectroscopy. Glycosidase digestions indicated that the 25-kDa PRL is N-glycosylated and sialylated, whereas 23-kDa PRL is nonglycosylated. Glycosylated and nonglycosylated forms of the hormone were individually purified to greater than 95% homogeneity using novel cation exchange chromatography. Isoelectric focusing demonstrated that both forms consist of multiple charge isomers, with the charge heterogeneity of the glycosylated form primarily due to differences in sialylation. Monosaccharide analysis of the glycosylated form suggested a minimal complex oligosaccharide chain that may be fucosylated and partially sialylated. Oligosaccharide mol wt were determined by electrospray ionization mass spectroscopy. Analysis of the oligosaccharides by fluorophore-assisted carbohydrate electrophoresis indicated that bi- and triantennary oligosaccharide forms are predominant and have multiple combinations of terminal sialylation. Both forms of PRL were active in the Nb2 lymphoma cell proliferation assay; however, the 23-kDa nonglycosylated form was 3-4 times more active in this assay than the 25-kDa glycosylated form.

A method for monitoring the glycosylation of recombinant glycoproteins from conditioned medium, using fluorophore-assisted carbohydrate electrophoresis.

We have developed a method for monitoring the N-glycosylation of recombinant glycoproteins directly from conditioned medium samples. Proteins in the conditioned medium are separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electroblotted onto polyvinylidene fluoride membranes. After staining the membranes with Coomassie blue, the protein(s) of interest is excised. Oligosaccharides are released from the membrane-bound glycoprotein by digesting with peptide N4-(acetyl-beta-glucosaminyl) asparagine amidase and labeled with the fluorophore 8-aminonaphthalene-1,3,6-trisulfonate (ANTS). Labeled oligosaccharides are then separated on polyacrylamide gels which allow for the direct comparison of samples. We have shown that recombinant human lysosomal hydrolase alpha-galactosidase A is N-glycosylated with both sialylated and phosphorylated oligosaccharides. ANTS-labeled oligosaccharide bands from alpha-galactosidase A were isolated from polyacrylamide gels. Sialylated and phosphorylated bands were identified by shifts in their electrophoretic mobility after digesting with neuraminidase or alkaline phosphatase to remove sialic acid or phosphate groups, respectively. Using the ANTS-labeled oligosaccharides from alpha-galactosidase A, we have shown that polyacrylamide gels can be used to resolve sialylated and phosphorylated oligosaccharide structures.

Defective expression of CD23 and autocrine growth-stimulation in Epstein-Barr virus (EBV)-transformed B cells from patients with Wiskott-Aldrich syndrome (WAS).

WAS is an X-linked, recessive, immune deficiency syndrome, characteristically associated with lymphocyte and platelet dysfunction. Peripheral B lymphocytes from WAS patients are nonresponsive to polysaccharide antigens and show reduced numbers of cells expressing the integral membrane glycoprotein, CD23. The release of CD23 proteolytic fragments, so-called soluble CD23 (sCD23), by B lymphoblasts and EBV-transformed B cell lines has previously been described, and these fragments have been shown to stimulate autocrine growth of these cells. We have found that the surface expression of CD23 is reduced on WAS compared with control EBV-B cells. Surface CD23 levels were reduced two-fold in four WAS cell lines (group I) and nine-fold in four other lines (group II). Group II WAS cell lines also showed reduced growth rates in serum-free medium when compared with group I cell lines and EBV-B cell lines from eight normal subjects. In contrast to the group II WAS lines, group I and EBV-B cells from normal individuals produced an autocrine-growth factor activity which could be absorbed by anti-CD23 antibodies. Immunoprecipitation of sCD23 from culture supernatants confirmed that group I WAS cell lines produced less sCD23, particularly the 37K fragment which was prevalent in control EBV-B cells. Northern analysis showed that CD23 mRNA levels were increased three-fold in group I and unchanged in group II WAS compared with normal EBV-B cell lines, suggesting that decreased surface expression in WAS EBV-B cells reflects post-transcriptional events. Together these results suggest that reduced cell surface expression and aberrant proteolysis of CD23 occurs in WAS patients' B lymphocytes and may contribute to impaired immune function in these patients.

Measurement of 226Ra and 228Ra in water by gamma-ray counting after preconcentration on ion-exchange resin.

A rugged resin concentration method followed by gamma counting was developed for measuring 226Ra and 228Ra in drinking water. Using typical distribution system pressure, 310 kPa (45 psig), radium in a 20-L sample was concentrated 100-fold onto 200 mL of ion-exchange resin beads that were then sealed in a glass jar for gamma activity measurement. Parameters and materials affecting the concentration step were studied with the objective of maximizing radium recovery in a short time. Empty bed contact times were varied from 0.1-3.0 min (2.0-0.67 mL min-1) and input concentrations of radium were varied from 370-3700 Bq m-3 (10-100 pCi L-1). Varying these parameters did not affect the percentage recovery of radium by the resin. At each empty bed contact time after all adsorbent media were tested, more than 99% of the influent radium was adsorbed. In these experiments, activities were counted for 100 min using a gamma-ray spectrometer equipped with a 3 x 3-inch sodium iodide (thalium-activated) [NaI(Tl)] crystal detector. Activity calculations were based on resin loaded with traceable standard solutions of 226Ra and 286Ra from the National Institute of Standards and Technology. Analyzing entire spectra with a least-squares analysis program resulted in limits of detection for 226Ra and 228Ra of 7.4 Bq m-3 (0.2 pCi L-1) and 33.3 Bq m-3 (0.9 pCi L-1), respectively.

Aberrant O-linked oligosaccharide biosynthesis in lymphocytes and platelets from patients with the Wiskott-Aldrich syndrome.

The Wiskott-Aldrich syndrome (WAS) is an X-linked recessive immunodeficiency affecting B lymphocytes, T lymphocytes, and platelets. Previous studies on lymphocytes from WAS patients have revealed that leu-kosialin (CD43), a cell-surface glycoprotein bearing approximately 90 O-linked oligosaccharide chains, shows an aberrant electrophoretic mobility. To determine whether this finding reflects a different pattern of O-linked glycosylation in WAS cells, we have compared healthy individuals and WAS patients with respect to glycosyltransferase activities in T lymphocytes, platelets, and Epstein-Barr virus (EBV)-immortalized B cell lines. Stimulation of peripheral T cells from normal individuals in vitro with anti-CD3 antibodies and interleukin-2 was associated with a 3-fold increase in UDP-GlcNAc:Gal beta 3GalNAc-R (GlcNAc to GalNAc) beta 6-N-acetylglucosaminyltransferase (core 2 GlcNAc-T) from 0.8 to 2.2 nmol/mg/h. In contrast, peripheral T lymphocytes from WAS patients showed an inversion of this phenotype with high core 2 GlcNAc-T activity in unstimulated cells (2.3 nmol/mg/h) and a 2-3-fold decrease in activity following stimulation. Core 2 GlcNAc-T activity was also three times higher in platelets from WAS patients than in normal platelets. Glycosyltransferase activities were measured in immortalized B cell lines established from WAS and normal subjects by infection with EBV. Core 2 GlcNAc-T was less than 0.4 nmol/mg/h in WAS EBV-B cell lines compared to 2.4 nmol/mg/h in EBV-B cell lines from healthy individuals, In contrast, CMP-SA:SA alpha 2-3Gal beta 1-3GalNAc-R (where SA represents sialyl (sialic acid to GalNAc) alpha 6-sialyltransferase II activity was 2.0 nmol/mg/h in the WAS EBV-B cell and less than .01 nmol/mg/h in EBV-B cell lines derived from normal subjects. Eleven other glycosyltransferase activities were measured and found to be similar in EBV-B cell lines from WAS and normal individuals. Polylactosamine sequences were much reduced in the O-linked oligosaccharides of CD43 from WAS EBV-B cells consistent with decreased core 2 GlcNAc-T activity and expression of core 1 oligosaccharides in the cells. In conclusion, B cells, T cells, and platelets in WAS patients show abnormal expression of two developmentally regulated glycosyltransferases, consistent with the idea that the WAS immunodeficiency is due to a failure of normal lymphocyte maturation.

Some effects of alcohol and simulated altitude on complex performance scores and breathalyzer readings.

Each of 17 men was trained on 7 tasks in the Multiple Task Performance Battery and then performed over a 2-week period in four experimental sessions: ground level with and without alcohol, and simulated altitude (12,500 ft), with and without alcohol. Subjects breathed appropriate gas mixtures through oxygen masks at both ground level and altitude. Alcohol doses of 100-proof vodka mixed with juice 2.2 ml X kg-1 of body weight. Results showed no differential effect of simulated altitude on breathalyzer readings (peaks averaged 78 mg % at 12,500 ft and 77 mg % at ground level). The best performance occurred at ground level under placebo conditions; the 12,500-ft simulated altitude produced some decrement for placebo scores. Alcohol at ground level resulted in significantly impaired performance during the morning sessions; the addition of altitude to the alcohol condition further depressed performance scores, but to about the same extent that placebo scores were depressed by altitude. Thus, there was no synergistic interactive effect of alcohol and altitude on either breathalyzer readings or performance scores.

Physiological biochemical, and performance responses to a 24-hour crash diet.

Twelve overweight male subjects were evaluated on both a normal diet and a 24-h crash diet. During approximately 2 1/4-h complex performance tests subjects breathed an O2/N2 mixture equivalent to 3810 m. (12,500 ft). There were no significant findings due to diet for heart rate, blood pressure, serum electrolytes, subjective fatigue and urinary excretion of K+, epinephrine and norepinephrine. body temperatures were lower (p less than 0.05) for the crash diet than for the normal diet. Serum glucose levels increased during the normal diet and decreased during the crash diet. Hematocrit increased more for the crash diet (p less than 0.05) than for the normal diet. Urinary excretion of 17-ketogenic steroids was less (p less than 0.001) during sleep for the crash diet than for the normal diet. Complex performance showed no significant differences when subjects were tested under low workloads. Performance was enhanced during the crash diet when subjects were tested under the medium and high workload conditions.

Pulmonary and symptom threshold effects of ozone in airline passenger and cockpit crew surrogates.

Previous studies showed that the ozone concentration for pulmonary and symptom threshold effects in flight attendant surrogates lies between 0.20 and 0.30 ppmv for a 3-h exposure with intermittent treadmill exercise at 1829 m (MSL) stimulated cabin altitude. In the present study of sedentary occupants of the in-flight airline cabin, the same protocol was used except for omitting all treadmill exercise. Symptoms were assessed with a standardized questionnaire. Pulmonary function was assessed using standardized quantitative spirometry. Male smoker and nonsmoker airline passanger and cockpit crew surrogates 40-59 years of age were used. Small but statistically significant displacements occurred in symptoms and in some spirometry parameters. In general, the younger subjects appeared more sensitive to ozone than the older subjects. No significant differences appeared between smokers' and nonsmokers' responses to ozone exposure. It is concluded that the ozone threshold of these sedentary surrogates under these experimental conditions is right at 0.30 ppmv.

Effects of ozone on symptoms and cardiopulmonary function in a flight attendant surrogate population.

Two studies were conducted to ascertain the lowest ozone concentration threshold for statistically significant adverse effects on cardiopulmonary function and symptoms in male and female surrogates of the flight attendant population. Both studies simulated in-flight environmental conditions at 1,829 m mean sea level (MSL) chamber altitude. The ozone exposures were 0.2 parts per million by volume (ppmv) for 4 h in the first study and 0.3 ppmv for 3 h in the second study. Each subject was similarly exposed to air only (no ozone) on another occasion. Treadmill exercise, equivalent to workloads of on-duty flight attendants, was performed at altitude. Cardiorespiratory measurements were made during ozone exposure, whereas spirometry and symptom assessments were made immediately before and after exposure. The ozone threshold for reversible adverse effects on symptoms and spirometry function was reached by a 3-h exposure to 0.3 ppmv. The data suggest a greater symptomatic sensitivity to ozone in females.

Effects of altitude and two decongestant-antihistamine preparations on physiological functions and performance.

Fourteen men were studied to determine the combined effects of two altitudes--388 and 3,810 m or 1,274 and 12,500 ft--and three preparations--lactose placebo, Compound A (Actified, and Compound B (Dristan). Subjects reported least attentiveness with A and greatest with placebo. Fatigue increased significantly with time while energy, interest, and attentiveness decreased. The Multiple Task Performance Battery (MTPB) showed no effects of altitude, drugs, or time on overall performance; however, performance declined with time in several tasks, while problem solving improved. Subjects enjoyed the problem-solving tasks and may have given them preference as levels of interest declined. Though the MTPB overall composite scores did not change significantly, physiological parameters and subjective evaluations indicate that type of compound and time after ingestion are important. Declines in energy and attentiveness 2.5 h after ingestion could result in neglect of important--although routine--tasks. Hypoxia might enhance this effect and consequences might be worse in subjects whose medical conditions require these drugs.

Biological aspects of suicide: circadian disorganization.

Disturbances in the circadian rhythmicity of biological functions have been reported in various mental disorders. Four lines of research--hormonal, electroencephalographic, cerebral spinal fluid, and circadian rhythmicity--suggest possible changes in suicidal individuals. During a study investigating the effect of a photoperiod shift on circadian rhythms, 15 male, healthy, normal subjects were used. Following a 5-day baseline period a 12-hour photoperiod shift took place and was followed by 10 days of recovery period. Multiple parameters were monitored. Two weeks following completion of the study one subject suicided. The data were examined to determine whether the suicided subject differed, rhythmically, from other subjects. Summation dials describing phase changes and vector difference dials describing dynamic phase relationships of rhythm pairs showed that the rhythms of this subject were poorly synchronized internally during baseline. Total urinary output of all parameters was lower than all other subjects during baseline and more of his urinary parameters rephased incompletely during recovery. The results suggest that circadian asynchrony and an inability to respond effectively to a phase shift may characterize a presuicidal state. These results are discussed in terms of the four lines of research involving biological aspects of suicide and suggest some intriguing interactions.

Transient and permanent effects of hyperthermia in dogs: a study of a simulated air transport environmental stress.

Heat-induced hyperthermia can be a major problem in dogs shipped during summer months. Dogs shipped by air transport can encounter temperatures as high as 54.4 C. Usually, little concern is given to effects produced by hyperthermia. To assess the heat stress problem, 20 dogs were exposed to a temperature of 54.4 C for 30 minutes--10 dogs at 15% relative humidity and 10 dogs at 35%. Dogs did not die as a result of exposure, but certain transient and permanent changes occurred. All dogs had increased heart rate, rectal temperature, blood pH, hemoglobin concentration, packed cell volume, and erythrocyte count. Body weight and PVCO2, decreased. Differences also were shown between the 2 humidity group for blood PH PVCO2, rectal temperature, and weight loss. The major tissue changes attributed to hyperthermia were fragmentation of the myocardium, acute cortical necrosis in kidney, and marked degenerative changes in the cerebellum and cerebral cortex. Changes in the cerebellum and cerebral cortex were considered severe and permanent.

Protection of airline flight attendants from hypoxia following rapid decompression.

To determine the maximum time for working flight attendants to effectively initiate airline passenger mask donning after onset of a rapid, severe decompression, we exposed 10 subjects in two series of tests to a decompression from 6,500 to 34,000 ft (2,000 to 10,400 m) in 26 s, followed by descent at 5,000 ft/min (1,500 m/min) while subjects performed a light-to-moderate workload. Supplemental oxygen was provided in one series from a compressed oxygen system, and in the other series from a chemical oxygen generator system. With delays to mask doning of 10 and 15 s, no hypoxic effects occurred. With delays of 20 and 25 s, increasing hypoxic effects, similar in frequency for the two systems, occurred. Some technical problems in mask donning contributed to losses of consciousness with the latter two delays.