Biomedical subjects

S L Young

Publications and source records attributed to S L Young.

At least 55 records · Page 3Linked to original sources

PubMed1994

Statistical methodology for the concurrent assessment of interrater and intrarater reliability: using goniometric measurements as an example.

Statistical methodology for the concurrent assessment of interrater and intrarater reliability is presented. Application of the methodology is illustrated with an example of one therapist using two goniometers repeatedly to measure knee joint angles. Methods for estimating the coefficients, testing hypotheses, constructing confidence intervals, and computing sample size requirements are provided. In addition, the calculation and clinical interpretation of the standard error of measurement (SEM) are discussed. It is recommended that (1) when both interrater and intrarater reliability are being assessed, a repeated-measures design be used to take advantage of the increased precision gained by using all observations in the statistical analysis, and (2) appropriate statistical tests, confidence intervals, and SEMs always be used in conjunction with the estimated reliability coefficients.

Bias↗

PubMed1994

Molecular and cellular processing of lung surfactant.

Pulmonary surfactant, a complex material that lines the alveolar surface of the lung, is synthesized in the type II pneumocyte. Surfactant consists largely of phospholipids, of which phosphatidylcholine is by far the most abundant component, and is mainly responsible for surface activity. Surfactant also contains four unique proteins, surfactant protein (SP)-A, SP-B, SP-C, and SP-D, which are synthesized in a lung-specific manner. SP-A and SP-D are glycoproteins (M(r) approximately 30,000-40,000) whereas SP-B and SP-C are small (M(r) approximately 5,000-18,000), extremely hydrophobic proteolipids released from large precursors by proteolysis. Synthesis of surfactant lipids and proteins is developmentally regulated in fetal lung and can be accelerated by glucocorticoids and other hormones. Developing fetal lung in vivo and in organ culture has been used extensively to study regulation of surfactant synthesis and gene expression. Glucocorticoids stimulate the rate of fetal lung phosphatidylcholine biosynthesis and the activity of the rate-regulatory enzyme, cholinephosphate cytidylyltransferase (CYT). The hormone, however, does not increase the amount of CYT; there is evidence that the increase in activity is mediated by increased fatty biosynthesis due to enhanced expression of the fatty acid synthase gene. Glucocorticoids also regulate expression of the SP-A, SP-B, and SP-C genes in the late gestation fetal lung. Hormone response elements and other cis-acting regulatory elements have been identified in the 5'-flanking regions of the SP-A, SP-B, and SP-C genes. Surfactant phospholipids are stored in lamellar bodies, secretory granules in the type II cell, and secreted by exocytosis. Lamellar bodies are also rich in SP-B and SP-C but there are conflicting data on the cellular distribution of SP-A. Secretion of SP-A may be constitutive and occur independently of lamellar bodies. Phosphatidylcholine secretion is a regulated process, and in isolated type II cells it can be stimulated by physiological and other agents that act via at least three signal-transduction mechanisms. After secretion, surfactant is transformed into tubular myelin, and the lipid and protein components are separated as the lipid is inserted into a monolayer at the air-liquid interface. The majority of surfactant is removed from the alveolar space by reuptake into the type II cell by mechanisms that may include receptor-mediated endocytosis. Some components of surfactant are directly recycled into new surfactant whereas other components are degraded.

Animals↗

PubMed1994

Natural surfactant and hyperoxic lung injury in primates. II. Morphometric analyses.

Diffuse lung injury is accompanied by low compliance and hypoxemia with histological evidence of endothelial and alveolar epithelial cell disruption. The histological effects of treatment of an acute diffuse lung injury with a natural surfactant product were evaluated in a primate model because surfactant function and content have been shown to be abnormal in diffuse lung injury in both animals and humans. Ten baboons were ventilated with 100% O2 for 96 h, and 5 were given an aerosol of natural porcine surfactant. Physiological and biochemical measurements of the effects of hyperoxia and surfactant treatment are presented in a companion paper. After O2 exposure, lungs were fixed and processed for quantitative electron microscopy. The responses to O2 included epithelial and endothelial cell injuries, interstitial edema, and inflammation. The hyperoxic animals treated with surfactant were compared with the untreated animals; the treatments altered neutrophil distribution, fibroblast proliferation, and changes in the volumes of type I epithelial cells and endothelial cells. Surfactant-treated animals also had decreased lamellar body volume density in type II epithelial cells and preservation of endothelial cell integrity. These changes suggest complex effects of natural surfactant on the pulmonary response to hyperoxia, including protection against epithelial and endothelial cell destruction as well as significant interstitial inflammation and fibroblast proliferation. We conclude that natural surfactant treatment of hyperoxic lung injury in primates resulted in partial protection of epithelial and endothelial cells but also increased the accumulation of fibroblasts in the lung.

Aerosols↗

PubMed1994

Natural surfactant and hyperoxic lung injury in primates. I. Physiology and biochemistry.

Surfactant dysfunction contributes to the pathophysiology of adult respiratory distress syndrome (ARDS), and we hypothesized that surfactant treatment would improve experimental ARDS produced by continuous exposure to hyperoxia. Twelve healthy male baboons (10-15 kg) were anesthetized, paralyzed, and mechanically ventilated with 2.5 cmH2O positive end-expiratory pressure (PEEP) for 96 h. Baboons were divided into three groups: 1) the O2 group (n = 5) received 100% O2, 2) the surfactant group (n = 5) received 100% O2 and aerosolized porcine surfactant, and 3) a control group (n = 2) was ventilated at fractional concentration of inspired O2 of 0.21 for 96 h to control for effects of anesthesia and mechanical ventilation. Hemodynamic parameters were obtained every 12 h, and ventilation-perfusion (VA/Q) distribution was measured daily by multiple inert gas elimination technique. PEEP was increased once or twice daily to 10 cmH2O for 30 min to study its effects on measurements of VA/Q. At the end of experiments, lungs were obtained for biochemical analysis. Prolonged hyperoxia resulted in progressive worsening in VA/Q, hemodynamic deterioration, severe lung edema, and altered surfactant metabolism. Surfactant administration increased disaturated phosphatidylcholine in lavage fluid but did not improve lung edema or gas exchange. In the surfactant group, however, the addition of 10 cmH2O PEEP resulted in a greater degree of shunt reduction than did 2.5 cmH2O PEEP (47 vs. 31% in the O2 group, P < 0.05). We conclude that aerosolized porcine surfactant did not prevent pulmonary O2 injury in baboons, but it potentiated the shunt-reducing effect of PEEP.

Aerosols↗

PubMed1994

Synthetic surfactant scavenges oxidants and protects against hyperoxic lung injury.

Injury and mortality after exposure to 100% oxygen can be diminished by surfactants that may operate by mechanisms other than those responsible for surface tension effects. We tested the hypotheses that 1) synthetic surfactant and its components function as antioxidants in vitro and 2) decrements in hyperoxic injury after treatment with a surfactant and its components are associated with decreases in oxidative stress to the lung. A synthetic surfactant (Exosurf) and its non-surface-active components tyloxapol and cetyl alcohol were incubated in an iron-containing hydroxyl radical-generating system to determine their abilities to prevent oxidation of deoxyribose. Doses of tyloxapol, cetyl alcohol, and artificial surfactant diminished the absorbance of thiobarbituric acid-reactive products of deoxyribose. Similarly, tyloxapol, cetyl alcohol, and the surfactant decreased hydroxylated products of salicylate in the same system. Rats were instilled intratracheally with saline, tyloxapol, tyloxapol plus cetyl alcohol, or artificial surfactant and immediately exposed to air or 100% oxygen. After 61 h of oxygen exposure, pleural fluid volume and wet-to-dry lung weight ratios were decreased in animals treated with surfactant and/or its components. There were also decrements in thiobarbituric acid-reactive products of lung tissue. In separate experiments, mean survival of saline-treated rats exposed to 100% oxygen was 67.3 +/- 8.1 h and > 96 h for rats given the surfactant or its components. We conclude that tyloxapol, cetyl alcohol, and Exosurf can function as antioxidants in vitro and their in vivo instillation is associated with reduction in measures of hyperoxic injury, oxidized tissue products, and mortality.

Animals↗

PubMed1993

Recycling of surfactant lipid and apoprotein-A studied by electron microscopic autoradiography.

Recycling of lipid and apoprotein components by lung epithelium appears to be a key part of the metabolism of surfactant, but many details of this process are poorly understood, including its efficiency and which intracellular pathways participate. We chose electron microscopic autoradiography to track time-dependent changes in the intracellular location of [methyl-3H]-choline dipalmitoyl phosphatidylcholine or 125I-labeled apoprotein-A that were incorporated into liposomes and instilled intratracheally into rats or added to cultures of freshly isolated rat lung type II cells. At times from 2 to 120 min after the start of labeling, lungs or cells were fixed and processed for autoradiography. We found a time-dependent uptake of lipid and apoprotein by type II cells and by macrophages but not by other lung cell types. There was a time-dependent incorporation of lipid and apoprotein label into lamellar bodies, but the labels appeared in other intracellular compartments without a convincing time-dependent relationship to lamellar bodies. Although all type II cell organelles received some lipid-bound label, the apoprotein-bound label was not found within electron-dense multivesicular bodies (d-MVB). Selective lack of apoprotein label in d-MVB indicated a segregation of lipid and apoprotein labels and suggested a unique role for the d-MVB, possibly as a degradation pathway. Our results also provided conclusive morphological evidence of surfactant lipid and apoprotein recycling by type II cells.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1993

Mechanisms responsible for reduced contextual conditioning with massed unsignaled unconditional stimuli.

Massed presentation of unsignaled shock results in less conditional freezing to contextual cues than do distributed presentations. Consistent with an account of the learning deficit based on the perceptual-defensive-recuperative theory, the massed-shock deficit was attenuated by preexposure to shock or the conditioning context. This formulation was also successfully applied to the deficit in conditioning that occurs when a single shock is given immediately after placement in a context. Opponent-process theory was not supported by 2 findings: (a) The deficit was neither enhanced by shock preexposure nor reduced by an opioid antagonist, and (b) unconditional reactions were greater with massed shock. Inconsistent with the suggestion that the effect is a performance artifact specific to freezing, the massed-shock deficit was apparent for a 2nd measure of conditioning.

Animals↗

PubMed1992

Conformational similarities between one-chain and two-chain tissue plasminogen activator (t-PA): implications to the activation mechanism on one-chain t-PA.

Tissue plasminogen activator (t-PA) is an exceptional serine protease, because unlike most other serine protease zymogens single-chain tissue plasminogen activator (sct-PA) possesses a substantial amount of proteolytic activity. The unusual reaction of sct-PA afforded the opportunity to directly compare the active site environment of sct-PA and two-chain tissue plasminogen activator (tct-PA) in solution through the application of a series of nitroxide spin labels and fluorophores. These labels, which have been previously shown to covalently label the catalytic serine of other serine proteases, inactivated both sct-PA and tct-PA. The labels can be divided into two classes: those which form tetrahedral complexes (sulfonates) and those which form trigonal complexes (anthranilates). Those which formed tetrahedral complexes were found to be insensitive to structural differences between sct-PA and tct-PA at the active site. In contrast, those which formed trigonal complexes could differentiate and monitor the sct-PA to tct-PA conversion by fluorescence spectroscopy. Models of the structure of sct-PA and tct-PA were constructed on the basis of the known X-ray structures of other serine protease zymogen and active enzyme forms. One of the nitroxide spin labels was modeled into the sct-PA and tct-PA structures in two possible orientations, both of which could be sensitive to structural differences between sct-PA and tct-PA. These models formed the structural rationale used to explain the results obtained with the "tetrahedral" and "trigonal" probes, as well as to offer a possible explanation for the unique reactivity of sct-PA.

Amino Acid Sequence↗

PubMed1992

Bile acid transport in the anhepatic rat.

Hepatocyte dysfunction eventually results in the loss of canalicular bile formation. Without canalicular flow, intestinal bile acid may originate from plasma by reverse transport. Anhepatic rats with preserved intestinal function permit evaluation of such transport. In the present study, plasma taurocholate clearance was markedly decreased in anhepatic rats. The relative proportion of free cholate increased with time. Peripheral tissues contained virtually only cleared taurocholate, but the intestinal contents were mainly free cholate. This indicates the intestinal contents as the source of the plasma cholate and shows an equilibrium between intestinal and plasma bile acid even without bile flow. The enteral administration of an anion exchange resin to anhepatic rats increased intestinal bile acid recovery and decreased the bile acid recovery in tissue. Plasma bile acid concentration was decreased and fractional loss increased threefold, confirming the anhepatic plasma-intestine bile acid equilibrium. However, the enhanced plasma clearance produced by the resin was less than 1% of the fractional loss found in the intact rat. These data show a very limited bile acid flux between intestine and plasma without bile flow, which could be modestly influenced by an intestinal bile acid sequestrant.

Animals↗

PubMed1992

Associative regulation of Pavlovian fear conditioning: unconditional stimulus intensity, incentive shifts, and latent inhibition.

Conditional stimuli (CS) associated with painful unconditional stimuli (US) produce a naloxone-reversible analgesia. The analgesia serves as a negative-feedback regulation of fear conditioning that can account for the impact of US intensity and CS predictiveness on Pavlovian fear conditioning. In Experiment 1 training under naloxone produced learning curves that approached the same high asymptote despite US intensity. Shifting drug treatment during acquisition had effects that paralleled US intensity shifts. In Experiment 3 naloxone reversed Hall-Pearce (1979) negative transfer using a contextual CS, indicating that conditional analgesia acquired during the CS-weak-footshock phase retards acquisition in the CS-strong-footshock phase. Experiment 5 used a tone CS in both a latent-inhibition and a negative-transfer procedure. Only negative transfer was blocked by naloxone. Therefore, negative transfer but not latent inhibition is mediated by a reduction of US processing.

Animals↗

PubMed1992

Cocaine: its effects on maternal and child health.

Cocaine abuse has become one of America's leading public health problems. Its use throughout pregnancy is associated with an increased risk of abruptio placentae, stillbirth, and preterm labor. Cocaine-associated neonatal complications include congenital malformations, decreased fetal growth, seizures, cerebral infarction and hemorrhage, auditory system deficits, sudden infant death syndrome, cardiac arrhythmias, necrotizing enterocolitis, and behavioral changes. Children followed throughout the first year of life continue to show developmental delay. Infants and children growing up in cocaine-abusing families are at risk for drug-related injuries. Accidental and intentional intoxication has occurred in infants and children from the smoke of freebase cocaine. The drug has also caused intoxication in breast-feeding infants. Adolescents experimenting with cocaine are at risk, with an apparently high frequency of seizures and loss of consciousness, as well as behavioral changes and psychosocial dysfunction.

Abnormalities, Drug-Induced↗

PubMed1991

Triple helix formation inhibits transcription elongation in vitro.

We have identified a 15-nucleotide site within a G-free transcription cassette that forms triple helix with sequence-specific oligodeoxyribonucleotides. When oligodeoxynucleotides were added to template DNA prior to in vitro transcription, a significant fraction of transcripts were truncated at a site corresponding to the region of triple helix formation. Kinetic analysis of the transcription products demonstrated that these truncated transcripts could be elongated to full length upon prolonged incubation. When an alkylating base was incorporated into the oligodeoxynucleotide to form covalent triple helix, most of the transcripts remained truncated. We conclude that triple helix formation can stall or, in the case of covalent crosslinking, can block RNA polymerase II and thus may provide a method for the specific inhibition of gene expression.

Base Sequence↗

PubMed1991

Analgesia for formalin-induced pain by lateral hypothalamic stimulation.

Few studies of analgesia induced by electrical brain stimulation have examined the effects of brain stimulation on responses to tonic pain stimuli. Recent evidence suggests that analgesia for tonic and phasic pain may involve different neural substrates. The present study examined the effects of lateral hypothalamic (LH) stimulation on responses to tonic pain induced by subcutaneous formalin. Our findings demonstrate that LH stimulation produced analgesia for tonic pain and that the effect is primary and not related to stress-induced analgesia. The results are discussed in relation to hypotheses regarding the different neural substrates involved in analgesia for tonic and phasic pain.

Analgesia↗

PubMed1991

A nonradioactive assay for transfected chloramphenicol acetyltransferase activity using fluorescent substrates.

Studies of the transcriptional activity of gene promoters have been greatly assisted by the widespread use of the chloramphenicol acetyltransferase (CAT) gene as a reporter gene. Previous techniques for assaying CAT enzymatic activity have utilized radioactive substrates or cofactors with the resulting complications of handling radioactive materials. We report here the development of fluorescent substrates for the CAT enzyme which form the basis of a CAT enzyme assay of enzyme kinetic parameters (Km and Vmax) and sensitivity similar to those based on radioactive substrates. Fluorescent substrates were designed as analogs of chloramphenicol and were based on the structure-function requirements of the enzyme. Several fluorophores were used to derivatize chloramphenicol base; one of the most effective was the borondipyrromethene difluoride (BODIPY) fluorophore. One BODIPY-chloramphenicol analog was found to have a Km for the purified CAT enzyme of 2 microM (compared to 12 microM for 14C-labeled chloramphenicol) and a Vmax of 120 pmole/min (compared to 180 pmol/min for the radioactive substrate). To verify its usefulness, a BODIPY--chloramphenicol-based CAT assay was used to measure transient transfection of primary cultures of ovarian granulosa cells in serum-free medium. This experimental system requires a highly sensitive assay for detecting transfected CAT gene activity. Robust expression of CAT activity was easily detected in crude cellular extracts using FluoReporter FAST CAT, a kit containing the BODIPY-chloramphenicol analog. The expression was precisely quantified by methanol extraction of the substrate and products from TLC plates and subsequent measurement of fluorescence using excitation-emission spectroscopy.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

Specific inhibition of protein kinase A in granulosa cells abolishes gonadotropin regulation of the proopiomelanocortin promoter.

Gonadotropins (follicle-stimulating hormone (FSH), luteinizing hormone, and human chorionic gonadotropin) and beta-adrenergic agonists have been shown to stimulate expression of the proopiomelanocortin (POMC) gene in ovarian granulosa cells. The current studies investigate the intracellular mechanisms by which gonadotropins regulate gene expression. Primary cultures of rat granulosa cells were transfected with the plasmid POMC-CAT-150, which expresses the chloramphenicol acetyltransferase (CAT) reporter gene under the regulation of the rat POMC 5'-flanking region. CAT activity was stimulated by treatment of the cells with either 20 ng/ml FSH or 1 microM isoproterenol. To assess the role of protein kinase A (ATP:protein phosphotransferase; EC 2.7.1.37) in the gonadotropin and adrenergic response, an expression vector, MtR-AB, encoding a mutant RI regulatory subunit was cotransfected with POMC-CAT-150. The mutant protein kinase A regulatory subunit encoded by MtR-AB lacks functional cAMP-binding sites but effectively binds and specifically inhibits the catalytic activity of protein kinase A. The results of this analysis demonstrated that gonadotropin and adrenergic agonist stimulation of the POMC-CAT reporter construct in primary cultures of rat granulosa cells were abolished by cotransfection with MtR-AB; whereas a control SV40-promoter construct was unaffected by either gonadotropin treatment or cotransfection with MtR-AB. Basal expression directed by the POMC promoter was also decreased by cotransfection with the MtR-AB, implying that basal expression from the POMC promoter may also depend on protein kinase A. Deletion analysis of the POMC sequence indicated regions (-40 to -33 and +4 to +63) important for basal and FSH-stimulated expression. These studies suggest that both gonadotropin and adrenergic stimulation of the POMC promoter are mediated by protein kinase A and that regions proximal to the promoter are essential for gonadotropin-regulated expression from the promoter.

Animals↗

PubMed1991

The pulmonary surfactant system.

Surfactant's importance is underscored by the severe morbidity of deficiency states--primary, as in neonatal respiratory distress syndrome, or secondary, as in ARDS. Current knowledge of surfactant synthesis and metabolism is reviewed. Clinically, the effectiveness of exogenous surfactant in RDS has prompted investigation of its administration in ARDS.

Adult↗

PubMed1991

Intracellular mechanisms of gonadotropin-stimulated gene expression in granulosa cells.

Previous studies have shown that the gonadotropins follicle-stimulating hormone and luteinizing hormone stimulate proopiomelanocortin (POMC) promoter activity and mRNA levels in ovarian granulosa cells. The objective of these studies was to determine the role of cAMP-dependent protein kinases (pKA) in gonadotropin-stimulated gene expression. Primary cultures of rat granulosa cells were transfected with a gene construct consisting of the POMC promoter (-150 to +63; designated pOMC-CAT) fused to the chloramphenicol acetyltransferase (CAT) reporter gene either alone or cotransfected with an expression plasmid (designated mutant RI), which overexpresses a mutant form of the murine RI subunit incapable of binding cAMP and serving as an irreversible inhibitor of the catalytic subunit of pKA. Follicle-stimulating hormone or isoproterenol caused a significant stimulation of pOMC-CAT activity in transfected cells. Cotransfection of pOMC-CAT with mutant RI caused a significant inhibition of basal pOMC-CAT activity and abolished the gonadotropin stimulation. As a control, transfection of the SV-40 viral enhancer-promoter fused to CAT (pSV2-CAT) was unresponsive to follicle-stimulating hormone stimulation and cotransfection with mutant RI had no significant effect on pSV2-CAT activity. These studies suggest that gonadotropin regulation of the POMC promoter is mediated by pKA and that promoter activity is stringently controlled by pKA.

Animals↗

PubMed1991

Development of type II pneumocytes in rat lung.

At a late stage of fetal development, the mammalian alveolar epithelium undergoes an abrupt differentiation as a part of the preparation of the lung for the postnatal demands of gas exchange. Some of the most striking changes occur in the type II pneumocytes as they lose their glycogen and start to produce the lamellated inclusion granules that contain pulmonary surfactant. Premature birth before adequate type II cell maturation results in the neonatal respiratory distress syndrome, which is frequently fatal. We have used serial ultrathin sectioning, electron microscopy, and three-dimensional reconstructions to study the ultrastructural features of maturation of rat type II cells from a single rat each at age gestational day 20 through adult stages. We found evidence over this time span for compartmentation of several secretory granule precursors within type II cells. Changes in the polarization of lamellar bodies were observed over the time period studied. We also found marked gestational changes in the number and morphology of type II cell cytoplasmic processes that perforate the basement membrane. Type II cell mitochondria changed in shape during postnatal development from single, spherical to complex, branched structures. Volume composition obtained from serial sections of a small number of type II cells agreed closely with published morphometric data, indicating that throughout the animal's lifespan, type II cells are a homogenous population.

Aging↗