Progressive changes in cortical state before and after spontaneous arousals from sleep in elderly and middle-aged women.

Arousals are often considered to be events which have an abrupt onset and offset, indicating abrupt changes in the state of the cortex. We hypothesized that cortical state, as reflected in electroencephalograph (EEG) signals, exhibits progressive systematic changes before and after a spontaneous, isolated arousal and that the time courses of the spectral components of the EEG before and after an arousal would differ between healthy middle-aged and elderly subjects. We analyzed the power spectrum and Sample Entropy of the C3A2 EEG before and after isolated arousals from 20 middle-aged (47.2±2.0 years) and 20 elderly (78.4±3.8 years) women using polysomnograms from the Sleep Heart Health Study database. In middle-aged women, all EEG spectral band powers <16 Hz exhibited a significant increase relative to baseline at some time in the 21 s before an arousal, but only low- (0.2-2.0 Hz) and high-frequency (2.0-4.0 Hz) delta increased in elderly and only during the last 7 s pre-arousal. Post-arousal, all frequency bands below 12 Hz transiently fell below pre-arousal baseline in both age groups. Consistent with these findings, Sample Entropy decreased steadily before an arousal, increased markedly during the arousal, and remained above pre-arousal baseline levels for ∼30 s after the arousal. In middle-aged, but not in elderly, women the presence of early pre-arousal low delta power was associated with shorter arousals. We propose that this attenuation of the effect of the arousing stimulus may be related to the slow (<1 Hz) cortical state oscillation, and that prolonged alterations of cortical state due to arousals may contribute to the poor correlation between indices of arousals and indices of sleepiness or impaired cognitive function.

Blowing smoke in Yellowstone: air quality impacts of oversnow motorized recreation in the park.

Snowmobile use in Yellowstone National Park has been shown to impact air quality, with implications for the safety and welfare of Park staff and other Park resource values. Localized impacts have been documented at several high-use sites in the Park, but the broader spatial variability of snowmobile emissions and air quality was not understood. Measurements of 87 volatile organic compounds (VOCs) were made for ambient air sampled across the Park and West Yellowstone, Montana, during 2 days of the 2002-2003 winter use season, 1 year before the implementation of a new snowmobile policy. The data were compared with similar data from pristine West Coast sites at similar latitudes. Backward trajectories of local air masses, alkyl nitrate-parent alkane ratios, and atmospheric soundings were used to identify the VOC sources and assess their impact. Different oversnow vehicle types used in the Park were sampled to determine their relative influence on air mass pollutant composition. VOCs were of local origin and demonstrated strong spatiotemporal variability that is primarily influenced by levels of snowmobile traffic on given road segments at different times of day. High levels of snowmobile traffic in and around West Yellowstone produced consistently high levels of benzene, toluene, and carbon monoxide.

Characterization and application of multiple genetic markers for Plasmodium malariae.

Plasmodium malariae, a protozoan parasite that causes malaria in humans, has a global distribution in tropical and subtropical regions and is commonly found in sympatry with other Plasmodium species of humans. Little is known about the genetics or population structure of P. malariae. In the present study, we describe polymorphic genetic markers for P. malariae and present the first molecular epidemiological data for this parasite. Six microsatellite or minisatellite markers were validated using 76 P. malariae samples from a diverse geographical range. The repeat unit length varied from 2 to 17 bp, and up to 10 different alleles per locus were detected. Multiple genotypes of P. malariae were detected in 33 of 70 samples from humans with naturally acquired infection. Heterozygosity was calculated to be between 0.236 and 0.811. Allelic diversity was reduced for samples from South America and, at some loci, in samples from Thailand compared with those from Malawi. The number of unique multilocus genotypes defined using the 6 markers was significantly greater in Malawi than in Thailand, even when data from single genotype infections were used. There was a significant reduction in the multiplicity of infection in symptomatic infections compared with asymptomatic ones, suggesting that clinical episodes are usually caused by the expansion of a single genotype.

Genotyping of Plasmodium falciparum infections by PCR: a comparative multicentre study.

Genetic diversity of malaria parasites represents a major issue in understanding several aspects of malaria infection and disease. Genotyping of Plasmodium falciparum infections with polymerase chain reaction (PCR)-based methods has therefore been introduced in epidemiological studies. Polymorphic regions of the msp1, msp2 and glurp genes are the most frequently used markers for genotyping, but methods may differ. A multicentre study was therefore conducted to evaluate the comparability of results from different laboratories when the same samples were analysed. Analyses of laboratory-cloned lines revealed high specificity but varying sensitivity. Detection of low-density clones was hampered in multiclonal infections. Analyses of isolates from Tanzania and Papua New Guinea revealed similar positivity rates with the same allelic types identified. The number of alleles detected per isolate, however, varied systematically between the laboratories especially at high parasite densities. When the analyses were repeated within the laboratories, high agreement was found in getting positive or negative results but with a random variation in the number of alleles detected. The msp2 locus appeared to be the most informative single marker for analyses of multiplicity of infection. Genotyping by PCR is a powerful tool for studies on genetic diversity of P. falciparum but this study has revealed limitations in comparing results on multiplicity of infection derived from different laboratories and emphasizes the need for highly standardized laboratory protocols.

Age- and species-specific duration of infection in asymptomatic malaria infections in Papua New Guinea.

The burden and duration of asymptomatic malaria infections were measured in residents of the malaria endemic village of Gonoa, Madang Province, Papua New Guinea. Plasmodium falciparum, P. vivax and P. malariae infections in people aged 4 years to adulthood were compared. Frequent sampling at 3-day intervals for up to 61 days allowed assessment of individual episodes of infection. Statistical assessment of P. falciparum detection revealed a periodicity consistent with synchronous replication of this species over periods up to 27 days. The duration of P. falciparum episodes was longer across all age groups than that of P. vivax and P. malariae. A trend for decreasing duration with age was also noted in data from each species. This was most prominent in P. falciparum infections: median duration in 4-year-olds was > 48 days compared with a median between 9 and 15 days in older children and adults. The results are consistent with the slow acquisition of immunity to antigenically diverse Plasmodium populations and suggest a faster rate of acquisition to P. vivax and P. malariae than to P. falciparum.

Genetic diversity and dynamics of plasmodium falciparum and P. vivax populations in multiply infected children with asymptomatic malaria infections in Papua New Guinea.

We describe the dynamics of co-infections of Plasmodium falciparum and P. vivax in 28 asymptomatic children by genotyping these species using the polymorphic loci Msp2 and Msp3alpha, respectively. The total number of Plasmodium spp. infections detected using 3 day sampling over 61 days varied between 1 and 14 (mean 6.6). The dynamics of P. falciparum and P. vivax genotypes varied greatly both within and amongst children. Periodicity in the detection of P. falciparum infections is consistent with the synchronous replication of individual genotypes. Replication synchrony of multiple co-infecting genotypes was not detected. In 4-year-old children P. falciparum genotype complexity was reduced and episodes lasted significantly longer (median duration > 60 days) when compared to children aged 5-14 years (median duration 9 days). P. vivax genotype complexity was not correlated with age but the episode duration was also longer for this species in 4-year-olds than in older children but was not as long as P. falciparum episodes. Recurrence of P. falciparum and P. vivax genotypes over weeks was observed. We interpret these major fluctuations in the density of genotypes over time as the result of the mechanism of antigenic variation thought to be present in these Plasmodium species.

Tuberculosis control and molecular epidemiology in a South African gold-mining community.

BACKGROUND: Gold miners have very high rates of tuberculosis. The contribution of infections imported into mining communities versus transmission within them is not known and has implications for control strategies. METHODS: We did a prospective, population-based molecular and conventional epidemiological study of pulmonary tuberculosis in a group of goldminers. Clusters were defined as groups of patients with Mycobacterium tuberculosis isolates with identical IS6110 DNA fingerprints. We compared the frequency of possible risk factors in the clustered and non-clustered patients whose isolates had fingerprints with more than four bands, and re-interviewed members of 45 clusters. FINDINGS: Of 448 patients, ten were excluded because they had false-positive cultures. Fingerprints were made in 419 of 438, of which 371 had more than four bands. 248 of 371 were categorised into 62 clusters. At least 50% of tuberculosis cases were due to transmission within the community. Patients who had failed treatment at entry to the study were more likely to be in clusters (adjusted odds ratio 3.41 [95% CI 1.25-9.27]). Patients with multidrug-resistant isolates were more likely to have failed treatment but were less likely to be clustered than those with a sensitive strain (0.27 [0.09-0.83]). HIV infection was common (177 of 370 tested) but not associated with clustering. INTERPRETATION: Despite a control programme that cures 86% of new cases, most tuberculosis in this mining community is due to ongoing transmission. Persistently infectious individuals who have previously failed treatment may be responsible for one third of tuberculosis cases. WHO targets for cure rates are not sufficient to interrupt transmission of tuberculosis in this setting. Indicators that are more closely linked to the rate of ongoing transmission are needed.

Cross-species interactions between malaria parasites in humans.

The dynamics of multiple Plasmodium infections in asymptomatic children living under intense malaria transmission pressure provide evidence for a density-dependent regulation that transcends species as well as genotype. This regulation, in combination with species- and genotype-specific immune responses, results in nonindependent, sequential episodes of infection with each species.

Polymorphism at the merozoite surface protein-3alpha locus of Plasmodium vivax: global and local diversity.

Allelic diversity at the Plasmodium vivax merozoite surface protein-3alpha (PvMsp-3alpha) locus was investigated using a combined polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) protocol. Symptomatic patient isolates from global geographic origins showed a high level of polymorphism at the nucleotide level. These samples were used to validate the sensitivity, specificity, and reproducibility of the PCR/RFLP method. It was then used to investigate PvMsp3alpha diversity in field samples from children living in a single village in a malaria-endemic region of Papua New Guinea, with the aim of assessing the usefulness of this locus as an epidemiologic marker of P. vivax infections. Eleven PvMsp-3alpha alleles were distinguishable in 16 samples with single infections, revealing extensive parasite polymorphism within this restricted area. Multiple infections were easily detected and accounted for 5 (23%) of 22 positive samples. Pairs of samples from individual children provided preliminary evidence for high turnover of P. vivax populations.

Developmental shift in the relative percentages of lung fibroblast subsets: role of apoptosis postseptation.

We have used the lipophilic, fluorescent dye Nile red and flow cytometry to identify and isolate two rat lung fibroblast subsets, lipid-containing interstitial cells (LICs) and non-LICs (NLICs) and to quantitate developmental changes in the relative percentages of these subsets. A significant decrease was observed in the percentage of LICs (from 79.0 +/- 3.8% on postnatal day 4 to 28.6 +/- 4.2% on day 30; P < 0.0001). To determine whether one or both subsets undergo apoptosis postseptation, fibroblasts from 16- to 18-day rats were treated with BODIPY-conjugated dUTP to label DNA strand breaks, which were then quantitated by flow cytometry. Apoptotic cells were judged to be predominantly LICs based on flow cytometric estimates of cell size and granularity and on light-microscopic colocalization of intracellular lipid and Hoechst-positive apoptotic bodies. Cell proliferation was compared in LICs and NLICs with both an in vitro [(3)H]thymidine incorporation assay and cell cycle analysis of propidium iodide-stained cells. Results of both assays indicated that on days 4-5, LICs proliferated more rapidly than NLICs. Tropoelastin and fibronectin mRNA expression, evaluated by RT-PCR, indicated that although tropoelastin mRNA levels did not differ, fibronectin mRNA levels were approximately ninefold greater in LICs. These results demonstrate the feasibility of a flow cytometric assay for the analysis of size, granularity, and intracellular lipid content of neonatal rat lung fibroblast subsets. Subsets differed substantially with respect to proliferative capacity, fibronectin mRNA expression, and incidence of apoptosis postseptation. Together with the observed changes in relative percentages of fibroblast subsets with age, these data suggest that the ratio of LICs to NLICs could be a critical determinant of fibroblast function during lung development.

The postnatal age of rat lung fibroblasts influences G1/S phase transition in vitro.

In the neonatal rat lung, alveolar development occurs from postnatal Days 4-13, during which time there is a fourfold increase in interstitial fibroblasts. Factors influencing emergence of new septa and cell proliferation associated with septal elongation have yet to be identified, in part because of difficulties inherent in studying this process in vivo. Using flow cytometric analysis of the DNA content of freshly isolated lung fibroblasts, we found that proliferation, as indicated by the percentage of cells in S plus G2/M phases, peaked on postnatal Day 4 (P < 0.04). By Days 9-10 the proliferation rate was lower than on Days 3, 4, 5, or 6 (P < 0.005). We then evaluated rates of in vitro proliferation as a function of postnatal age in first passage fibroblasts and found that the proliferative phenotype expressed in vivo persists in vitro. Fibroblasts from 4-5-d-old pups increased in number and incorporated 3H-thymidine at a faster rate than did fibroblasts obtained from pups at other postnatal ages (P < 0.0001). Age-dependent differences in cell cycle transit time were compared in fibroblasts synchronized by serum starvation and analyzed by flow cytometry at 2-h intervals from 13-21 h after release from serum starvation. A greater percentage of cells from 5-d-old pups entered S phase during this period than was seen for cells obtained from 2-, 9-, 13-, or 23-d-old rat pups (P = 0.0001). Cells from 5-, 9-, and 13-d-old pups reentered G0/G1 by 21 h after release from serum starvation, in contrast to fibroblasts from 2- and 23-d-old rats which did not. Throughout the 15-h period after release from serum starvation, levels of cyclin E, which peaks at the G1/S border, were highest in the 5-d-old cells (P < 0.025). Synchronization with 2.5 mM hydroxyurea which inhibits DNA synthesis completely abolished age-related differences in cell cycle transit time, implying that age-dependent differences in lung fibroblast proliferation rates are the result of events occurring before S-phase entry.

Lung fibroblasts undergo apoptosis following alveolarization.

In the rat lung, primary saccules are transformed into alveoli from postnatal Days 4 to 13, after which time there is a 20% reduction in the number of lung fibroblasts as the interstitial volume of the alveolar walls decreases. Our objective was to determine whether apoptosis is a factor in the observed decrease in the number of interstitial lung fibroblasts beyond Day 13. We used both histologic and flow cytometric assays to detect in lung fibroblasts the DNA fragmentation and condensation that are characteristic of apoptosis. In addition, we evaluated levels of bcl-2 and BAX messenger RNAs (mRNAs) using a reverse transcriptase-polymerase chain reaction (RT-PCR) assay. Apoptotic cells were quantitated in glycol methacrylate-embedded sections of neonatal rat lungs using the terminal transferase dUTP-digoxygenin nick end-labeling (TUNEL) method. Although TUNEL-positive interstitial cells were observed in the lungs of rats ranging in age from 10 to 16 d, a dramatic increase in apoptotic cells was seen on Day 17. Although diminished in number, TUNEL-positive cells were still present on Day 28. Hoechst-stained apoptotic bodies were observed in isolated lung cells that were vimentin-positive and factor VIII-negative, which identified the apoptotic cells as fibroblasts as opposed to endothelial cells. Flow cytometric analysis of freshly isolated lung fibroblasts stained with Hoechst 33342 indicated a 24% increase in chromatin condensation in cells from 17-d versus 16-d rats. DNA fragmentation was also quantitated by flow cytometry in freshly isolated fibroblasts labeled with BODIPY-conjugated dUTP in the presence of terminal deoxynucleotidyl transferase. The percentage of lung fibroblasts containing fragmented DNA was 51.4 +/- 13.4 in 17-d, 36.9 +/- 8.6 in 18-d, and 13.8 +/- 5.4 in 19-d rat pups. Finally, evaluation by RT-PCR indicated that on postnatal Day 17, mRNA for bcl-2, which inhibits apoptosis, was decreased to 73.5 +/- 11.4% (P < 0.001) of Day 5 controls; whereas mRNA for BAX, which enhances apoptosis, was increased to 243.0 +/- 102.0% (P < 0.001) of Day 5 values. These results demonstrate that rat lung fibroblasts undergo apoptosis after the completion of alveolarization, and suggest that this decrease in fibroblast number plays an important role in the thinning and remodeling of the alveolar walls of the lung.

Transcriptional regulation of tropoelastin expression in rat lung fibroblasts: changes with age and hyperoxia.

Elastic fibers are thought to provide structural support for secondary septa as the lung undergoes the transition from the saccular to the alveolar stage. The synthesis of the soluble precursor of elastin, tropoelastin, occurs during a finite developmental period. We have investigated the developmental regulation of tropoelastin gene transcription and mRNA expression in fetal and postnatal rat lung fibroblasts and have assessed the changes in tropoelastin gene expression caused by hyperoxic exposure during secondary septal development. With the use of an RT-PCR assay and intron-specific primers to detect heterogeneous nuclear RNA (hnRNA) and intron-spanning primers to detect mRNA in freshly isolated rat lung fibroblasts, tropoelastin gene expression was found to be upregulated late in gestation. From days 18 to 21 of gestation, there was a 4.5-fold increase in tropoelastin hnRNA (P < 0.0001) and a 6-fold increase in mRNA (P = 0.002). After birth, tropoelastin expression was downregulated. Signals decreased from fetal day 21 to postnatal day 2 for both tropoelastin hnRNA (P = 0. 021) and mRNA (P = 0.043). Tropoelastin hnRNA decreased further from days 2 to 6 (P = 0.04). Both tropoelastin hnRNA and mRNA were again upregulated during alveolarization from days 9 to 11, indicating that, once upregulated, transcription of the tropoelastin gene is not constant but varies with fetal and postnatal age. Exposure to >95% oxygen, when initiated on postnatal day 2 or 3 and continued until day 11, significantly diminished the developmental increase in tropoelastin hnRNA (P < 0.005) and mRNA (P < 0.05) normally seen on days 9-11, indicating that the postnatal upregulation of tropoelastin gene expression is inhibited by hyperoxic exposure in the early postnatal period.

Elevated laminin concentrations in lung secretions of preterm infants supported by mechanical ventilation are correlated with radiographic abnormalities.

OBJECTIVE: The objective of this study was to test the hypothesis that the presence of laminin in neonatal tracheal aspirates would be indicative of damage to the structural integrity of the basal laminae of the lung caused by barotrauma and hyperoxia. We predicted that disruption of the basal laminae would be a critical determinant of lung injury and fibrotic repair in the preterm infant whose lungs were ventilated with supplemental oxygen. STUDY DESIGN: The study group consisted of 23 premature infants in the neonatal intensive care unit whose lungs were ventilated by supplemental oxygen. We quantitated concentrations of laminin and fibronectin from sequential tracheal aspirates by enzyme-linked immunosorbent assays. A two-way analysis of variance was used to compare laminin and fibronectin concentrations in infants with and without radiographic evidence of coarse pulmonary markings indicative of fibrotic repair of lung injury. RESULTS: The concentrations of laminin, but not fibronectin, were significantly higher throughout the first 5 weeks of life in infants with abnormal chest radiographs at 36 weeks after conception. The concentrations of laminin in infant serum were approximately 1/30 that of tracheal aspirate laminin concentrations, suggesting that little if any of the laminin detected in the tracheal aspirates was derived from the serum. CONCLUSIONS: Increased concentrations of laminin in tracheal aspirates may be an indication of lung injury and fibrotic repair in the preterm infant.

Recent transmission of tuberculosis in a cohort of HIV-1-infected female sex workers in Nairobi, Kenya.

OBJECTIVES: To describe the epidemiological and clinical characteristics of HIV-related tuberculosis in a female cohort, and to investigate the relative importance of recently transmitted infection and reactivation in the pathogenesis of adult HIV-related tuberculosis. DESIGN: Members of an established cohort of female sex workers in Nairobi were enrolled in a prospective study. Women were followed up regularly and seen on demand when sick. METHODS: Between October 1989 and September 1992 we followed 587 HIV-infected and 132 HIV-seronegative women. Standard protocols were used to investigate common presentations. Cases of tuberculosis were identified clinically or by culture. All available Mycobacterium tuberculosis strains underwent DNA fingerprint analysis. RESULTS: Forty-nine incident and four recurrent episodes of tuberculosis were seen in HIV-infected women; no disease was seen in seronegative sex workers (P = 0.0003). The overall incidence rate of tuberculosis was 34.5 per 1000 person-years amongst HIV-infected participants. In purified protein derivative (PPD) skin test-positive women the rate was 66.7 per 1000 person-years versus 18.1 per 1000 person-years in PPD-negative women. Twenty incident cases (41%) were clinically compatible with primary disease. DNA fingerprint analysis of strains from 32 incident cases identified two clusters comprising two and nine patients; allowing for index cases, 10 patients (28%) may have had recently transmitted disease. Three out of 10 (30%) patients who were initially PPD skin test-negative became PPD-positive. Taken together, 26 incident cases (53%) may have been recently infected. DNA fingerprint analysis also identified two (50%) of the four recurrent tuberculosis episodes as reinfection. CONCLUSIONS: Substantial recent transmission of tuberculosis appears to be occurring in Nairobi amongst HIV-infected sex workers. It may be incorrect to assume in other regions of high tuberculosis transmission that active HIV-related tuberculosis usually represents reactivation of latent infection.

PIP: A 3-year (1989-92) prospective study of 587 HIV-positive and 132 HIV-negative commercial sex workers in Nairobi, Kenya, revealed substantial recent transmission of tuberculosis in the HIV-infected group. The cohort was enrolled at a community clinic that provides counseling, sexually transmitted disease services, and free condoms. In HIV-positive women, 49 incident and 4 recurrent episodes of tuberculosis were diagnosed during the study period; there were no tuberculosis cases among HIV-negative women. The overall incidence rate of tuberculosis was 34.5/1000 person-years among HIV-positive women. 20 incident cases (41%) met the clinical case definition of primary disease. DNA fingerprint analysis of strains from 32 incident cases suggested 10 women (28%) may have had recently transmitted disease. 3 of 10 women who were initially purified protein derivative (PPD) skin test-negative became PPD-positive. Clinical presentation, tuberculin skin testing, and strain clustering data all independently suggested that substantial Mycobacterium tuberculosis transmission was occurring in HIV-infected prostitutes during the study period. As many as 26 (53%) of the 49 patients with incident disease may have recently acquired tuberculosis and DNA fingerprint analysis identified 2 (50%) of the 4 recurrent tuberculosis episodes as reinfection. These findings challenge the assumption that tuberculosis in HIV-infected individuals represents reactivation of latent endogenous infection.

Postnatal age at onset of hyperoxic exposure influences developmentally regulated tropoelastin gene expression in the neonatal rat lung.

Upregulation of tropoelastin (TE) gene expression in rat lung interstitial fibroblasts normally occurs during alveolar septation. TE message increases at the end of the first week of life, peaks on days 9-11, and returns to barely detectable levels over the next 7-10 days. Our previous in situ hybridization studies indicated that exposure of pups to > 95% oxygen from 3 to 13 days of age interfered with the increased in TE gene expression in interstitial fibroblasts normally seen during septation. However, when the pups were returned to room air, lung fibroblast TE message levels increased, exceeding levels seen in control lungs during the exposure. In addition, TE message levels remained elevated for a week after levels in control lungs had returned to background. A possible interpretation of these results was that the developmentally regulated increase in TE messenger RNA (mRNA) was downregulated by the hyperoxic exposure but resumed when the pups were returned to a normoxic environment. We report herein the results of a subsequent study conducted to determine whether continued hyperoxic exposure beyond day 13 would further delay the peak in TE mRNA. Rat pups were exposed to 95% O2 from 5 to 17 days of age. TE and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) message levels in lung interstitial fibroblasts were assessed by in situ hybridization. As observed in pups exposed from 3 to 13 days, hyperoxic exposure from days 5 to 17 also extended the period during which TE mRNA levels were elevated. After exposure, TE message levels were 99%, 262%, and 223% of controls on days 19, 21, and 23 respectively. In addition, delaying the exposure 2 days until the pups were 5 days old resulted in an upregulation of TE message, relative to control values, during the hyperoxic exposure. In hyperoxic pups, values for TE message expression were 105%, 152%, 168%, and 144% of control pups on days 9, 11, 13, and 16 respectively. The influence on peak TE message expression of postnatal age at the time of exposure was further explored to verify the results of the 3-13 and 5-17 day exposures. When pups were exposed continuously from 2, 3, 4, 5, or 6 days until 11 days of age, the results of both in situ hybridization and Northern blot analysis confirmed our previous observations, demonstrating that the postnatal age at which hyperoxic exposure is initiated influences TE message expression in the developing lung.

Comparison of secretory component for immunoglobulin A with albumin as reference proteins in tracheal aspirate from preterm infants.

OBJECTIVES: To determine whether the concentration of secretory component (SC) in tracheal aspirate samples is less altered by changes in alveolar-capillary permeability and thus is a more reliable reference standard than albumin for the measurement of other components obtained by saline lavage in preterm infants. METHODS: A total of 1229 tracheal aspirate and 1530 blood samples were collected from 195 neonates to evaluate the effects of advancing postnatal and gestational age, resolution of acute respiratory distress syndrome (RDS), steroid therapy for chronic lung disease, and acute sepsis on tracheal aspirate SC and albumin levels. The tracheal aspirate and blood samples were analyzed by enzyme-linked immunosorbent assay techniques for SC and albumin concentrations. RESULTS: The mean values for the concentrations of aspirate and plasma SC did not vary significantly during an 8-week study period (n = 100) and did not vary with either gestational age (23 to 36 weeks) or postnatal age. Albumin concentration significantly decreased in aspirate samples from 1.67 +/- 0.77 mg/dl at week 1 to 0.41 +/- 0.21 mg/dl at week 8 (p < 0.001), whereas serum levels increased from 2.65 +/- 0.36 to 2.99 +/- 0.54 gm/dl (p < 0.001), suggesting a decrease in alveolar-capillary leakage with advancing postnatal age. The concentration of SC in aspirate samples from 51 infants who received dexamethasone remained constant during the first week of therapy, whereas the concentration of albumin decreased from 1.33 +/- 0.91 mg/dl at the initiation of therapy to 0.51 +/- 0.34 mg/dl on treatment day 7 (p < 0.001). The onset of sepsis (n = 40) was not accompanied by a significant change in either aspirate SC or albumin levels. However, in infants who had a deterioration in respiratory status concomitant with the onset of sepsis (n = 10), the levels of aspirate albumin increased whereas serum levels decreased (p < 0.001), suggesting an increase in alveolar-capillary leakage; the levels of aspirate SC remained unaltered. CONCLUSIONS: Secretory component may serve as a more valid reference protein for the standardization of tracheal aspirate collection in preterm infants during evaluation of changes in inflammatory mediators in disease states and therapeutic interventions that alter alveolar-capillary integrity.

Cellular location and temporal expression of the Plasmodium falciparum sexual stage antigen Pfs16.

The temporal expression during gametogenesis and the cellular location of the sexual stage specific protein Pfs16, a putative integral membrane protein of Plasmodium falciparum, was investigated using two monoclonal antibodies, 2G7 and 93A3A2. Using sorbitol synchronised, in vitro gametocyte cultures along with immunofluorescence assays, the time at which Pfs16 is first expressed during gametogenesis has been estimated to 35 hours post merozoite invasion. By immunofluorescence assays on thin blood smears monoclonal antibodies specific for Pfs16 react strongly with the gametocyte and also with vesicles within the red blood cell cytoplasm, many of which connect with the gametocyte cell. Purification of parasitophorous vacuole membranes from mature and immature gametocytes and immunoelectron microscopy on gametocytes during gametogenesis have allowed us to locate Pfs16 to the parasitophorous vacuole membrane. During gametogenesis this membrane is shed along with the red blood cell membrane. Immunofluorescence assays and immunoelectron microscopy studies of emerged gametes indicate that in a minority of cases the parasitophorous vacuole membrane along with Pfs16 can be retained to some extent on the gamete surface.

Strain increases airway smooth muscle cell proliferation.

Airway smooth muscle (ASM) cell hyperplasia is a common finding in lung diseases in which increased transpulmonary pressure is necessary for adequate ventilation. The extent to which mechanical deformation of ASM cells contributes to the observed hyperplastic changes has not been addressed. To determine if cyclic stretch alters ASM cell proliferation and protein content in vitro, canine ASM cells were cultured on collagen-coated silastic membranes, subjected to a stretch-relaxation regimen, and compared with cultured cells on a rocking platform or with control (unmanipulated) cells. During the log phase of cell growth, cyclic stretch resulted in increased cell numbers (P < 0.001). Incorporation of [3H]thymidine ([3H]TdR) was assessed after 5 or 14 days in culture during 12-h pulse in stretched, rocked, and control cells. Incorporation of [3H]TdR per cell was elevated in stretched cells when compared with either rocked or control cells at 5 days but not 14 days. Total cellular protein content per well determined after cells had reached confluence was greater in stretched cells than in the controls (74.2 +/- 7.9 stretched versus 49.1 +/- 5.6 control versus 51.1 +/- 6.5 rocker; microgram/10(5) cells; P < 0.05). Transmission electron microscopic observations of stretched cells suggested increased concentrations of myofilaments compared with control cells. These observations suggest that stretch may contribute to increased ASM hyperplasia and airway resistance.