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Biomedical subjects

S L Young

Publications and source records attributed to S L Young.

At least 73 records · Page 4Linked to original sources
PubMed1991

Surfactant apoprotein in adult rat lung compartments is increased by dexamethasone.

The distribution of the major surfactant apoprotein (SP-A) in adult rat lung was determined in order to gain insight into its metabolism, including packaging of SP-A into lamellar bodies. The effect of glucocorticoid treatment on surfactant apoprotein was studied to test whether regulation of surfactant apoprotein genes, which has been described for the fetal lung, can be demonstrated in the adult animal. We measured the amounts of immunoreactive SP-A in several lung tissue compartments and lavage fractions from control animals and from the lungs of rats given dexamethasone for 1 wk. Protein and phospholipids were measured, SP-A was quantitated with a noncompetitive enzyme-linked immunoabsorbent assay (ELISA) and SP-A, SP-B, and SP-C mRNAs were estimated by Northern blotting. We found an 85-fold concentration of SP-A in a lamellar body-rich fraction compared with lung tissue homogenate and we calculated that as much as one-half of all the tissue SP-A might be accounted for by a lamellar body pool. After 1 wk of dexamethasone treatment, there was an increase in adult rat lung SP-A, SP-B, and SP-C mRNA and a substantial increase in tissue and lavage fluid immunoreactive SP-A pools. Lamellar body fraction SP-A content per lung was 1.4-fold higher after dexamethasone, and there was a fivefold increase in the lavage SP-A pool, much of which was inseparable from the alveolar macrophages. We conclude that SP-A is concentrated in the lamellar bodies of type II cells, that dexamethasone treatment increased all surfactant mRNAs, and that it increased SP-A content in adult rat lung.

Animals↗
PubMed1991

Ontogeny of rat lung type II cells correlated with surfactant lipid and surfactant apoprotein expression.

During the last stages of intrauterine growth, remarkable changes occur in the alveolar epithelium that include cellular differentiation and increased production of surfactant lipid and apoprotein. We made morphometric measurements of type II cell characteristics from rats aged gestational day 20 to 14 days postnatal. We also measured the amounts of disaturated phosphatidylcholine (DSPC) and surfactant apoprotein (SP-A) in lung tissue, bronchoalveolar lavage, and a lamellar body-rich fraction, and we estimated the lung content of mRNAs for SP-A, SP-B, and SP-C. Lavage and lamellar body surfactant lipid and apoprotein content per lung showed a pattern of a sharp rise in the early postnatal period, then a substantial decline, and a second increase by day 14. When normalized for dry lung weight, the highest DSPC values were found on postnatal day 1 in all compartments. The fraction of whole lung DSPC found in lamellar body or lavage was greatest in the 48-h period surrounding birth. Lamellar body SP-A was greater than lavage SP-A on gestational day 22, but a day later the lavage SP-A was 16 times greater than the lamellar body SP-A. The lung tissue content of all three apoprotein mRNAs increased sharply before birth, fell during the 1st postnatal wk, and then rose again to adult levels. Type II cell number and lamellar body number per milligram of dry lung tissue was highest on post-natal day 1 and fell by one-half during the 1st postnatal wk. The amount of DSPC per unit of lamellar body volume rose to its greatest value on postnatal day 1 and then decreased more than threefold. These findings indicate a pattern of expansion of surfactant cellular and biochemical pools at the time of birth in the rat.

Aging↗
PubMed1990

The effect of the one-chain to two-chain conversion in tissue plasminogen activator: characterization of mutations at position 275.

Tissue plasminogen activator (t-PA) is homologous to other serine proteases and contains an apparent activation cleavage site at arginine 275. It has been demonstrated that this arginine-275 can be replaced with either glutamic acid (Tate, K. M., Higgins, D. L., Holmes, W. E., Winkler, M. E., Heyneker, H. L., and Vehar, G. A. Biochemistry 26, 338-343, 1987) or glycine (Peterson, L. C., Johannessen, M., Foster, D., Kumar, A., and Mulvihill, E. Biochim. Biophys. Acta 952, 245-254, 1988; Boose, J. A., Kuismanen, E., Gerard, R., Sambrook, J. and Gething, M.-J. Biochemistry 28, 635-643, 1989) so that the product of the plasminogen activation reaction, plasmin, can no longer hydrolyze the one-chain form of t-PA to the two-chain form. These "one-chain" t-PA variants had diminished activity, compared to wild-type t-PA, in the absence of a cofactor, but in the presence of the fibrin(ogen) cofactor the two variants had activity similar to wild-type t-PA. In order to compare the effects of all possible substitutions, t-PA variants with each of the other nineteen amino acids besides arginine at position 275 were produced by site-directed mutagenesis. All were recovered from cell culture supernatants completely in the one-chain form, except for R275 (wild-type) and R275K, which were partially converted to the two-chain form. These latter two species could be completely converted to the two-chain form by plasmin. In addition, these two forms showed significantly more plasminogen activating activity in the absence of a fibrin(ogen) cofactor, compared to the other 18 variants. In the presence of a cofactor, all of the t-PA mutants had plasminogen activating activity equivalent to wild-type t-PA, except for R275C. The R275C t-PA had comparatively less clot lysis and fibrin binding activity as well. Presumably the new cysteine in this variant was involved in a mixed disulfide or caused misfolding of the molecule resulting in decreased activity. The difference in the plasminogen activating activity of one- and two-chain forms of t-PA was investigated by determining the apparent Michaelis constants and the apparent turnover numbers for R275E t-PA, which remains in the one-chain form throughout the assay, and two-chain R275 t-PA. The kinetic constants were measured in both the presence and the absence of plasmin-digested fibrinogen.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1990

The regulation of proenkephalin expression in a distinct population of glial cells.

The expression of opioid genes was examined in isolated populations of glial cells in primary culture. Northern blot analysis of purified type I astrocytes, oligodendrocytes and mixed oligodendrocyte-type-2-astrocyte lineage cells derived from cerebral cortex demonstrated robust expression of proenkephalin mRNA exclusively in type I astrocytes. The expression of proenkephalin mRNA was stimulated by the beta-adrenergic agonist isoproterenol, and 8-(4-chlorophenyl thio)adenosine 3'-5'-cyclic monophosphate (cpt-cAMP). Both of these compounds regulated a proenkephalin-chloramphenicol acetyltransferase fusion gene transiently transfected into type I astrocytes. HPLC and immunoassay of the cell culture media revealed significant levels of unprocessed proenkephalin secreted by the cell and this secretion was stimulated by isoproterenol and cpt-cAMP. The relatively high levels of proenkephalin expressed suggest that enhanced expression in astrocytes may be important during neural development, in trauma-induced gliosis and in neuroimmune interactions.

Animals↗
PubMed1990

Measurement of boundaries using a digitizer tablet.

The perimeter is the most error prone of the primary measurements (length, perimeter and area) made when using a device such as a digitizer tablet to trace profiles on micrographs. To allow for minimization of this error an expression is developed relating the error in the perimeter to the digitizer resolution plus other errors in the entry process. The predictions of this expression are shown to be in good agreement with the results of computer simulation of the tracing process, as well as results from manual tracing using a digitizer tablet. This analysis of the digitizer entry process also provides a method by which an optimal choice of the sample spacing between digitizer coordinates can be made.

Computer Simulation↗
PubMed1990

Dentists' and physicians' attitudes on the role of the dental health care team in a cardiovascular risk factor reduction program.

This study examines the attitudes of both dentists and physicians on the role of the dental health care team in a cardiovascular risk factor reduction program directed at a general "well" population. A questionnaire was sent to a stratified sample of primary care dentists and physicians in the Augusta, Georgia, metropolitan area. The questionnaire was divided into four parts: the demographic component and three attitude components (the affective component, the cognitive component, and the action component). Results showed that dentists tended to have a positive attitude about performing cholesterol screening, nutrition counseling, and blood pressure screening in the dental office. Physicians tended to be negative about the idea of dentists performing cholesterol screening and nutrition counseling and were undecided about this role for dentists in a cardiovascular risk factor reduction program. Both dentists and physicians had positive attitudes about the idea of dentists referring high cardiovascular risk patients to physicians, as well as about the role of dentists in blood pressure screening. Overall, although dentists and physicians expressed concerns about the role of dentists in a cardiovascular risk factor reduction program, responses of both groups demonstrated a willingness to explore this concept further.

Adult↗
PubMed1990

Alveolar type II cell responses to chronic inhalation of chrysotile asbestos in rats.

The effects of chronic exposure to chrysotile asbestos on alveolar type II cells were examined in the lungs of Fischer 344 rats. Morphometric and three-dimensional analyses were used to characterize the alveolar type II cell and to determine the relationship of asbestos fiber localization to ultrastructural change in these cells. During the 2-yr period of study, type II cell number and volume increased to values more than 4 times those seen in controls. Ultrastructurally, cisternal dilations of the rough endoplasmic reticulum (RER) composed 12% of the total cell volume after 12 mo of exposure to asbestos and was still 15% of the total cell volume 1 yr after fiber exposure had ended compared to less than 1% in control cells. Asbestos fiber density surrounding these cells was directly proportional to the degree of cisternal dilatation in the cell; however, lamellar body volume and number in these cells were not different from that found in control type II cells. The incidence of a subset of type II cells with large lamellated inclusions was 10-fold greater in regions near bronchiolar-alveolar duct junctions, compared to more distal gas exchange regions of the lungs. Normal-sized lamellar bodies were fused to these large lamellated inclusions. These cells also contained significantly greater numbers of lamellar bodies and multivesicular bodies than those type II cells in more distal lung regions. These ultrastructural changes observed in type II cells may be a simple dose response to inhaled asbestos or the manifestation of two distinct populations of cells in the lungs that respond to asbestos in different ways. Asbestos fiber dose, cellular microenvironment, and aberrations of the cell plasma membrane and/or cell cytoskeleton (i.e., microtubules and filaments) are discussed as potential factors in the changes noted in type II cells.

Administration, Inhalation↗
PubMed1990

Neonatal hyperoxia alters the pulmonary alveolar and capillary structure of 40-day-old rats.

High inspired oxygen concentrations during the neonatal period profoundly inhibit rat lung development, an effect that is partly reversed during recovery in air. Persistent effects of neonatal hyperoxia on the size and number of alveoli or the structure of pulmonary capillaries have not been well defined. Using light and electron microscopic morphometry plus quantitative three-dimensional reconstructions of alveoli, we examined the lungs of 40-day-old rats that were exposed to more than 95% oxygen for the first 7 days after birth. Neonatal hyperoxia administered to rats resulted in abnormally enlarged air spaces at age 40 days. The fraction of the lung consisting of parenchyma was significantly increased and alveolar surface area was 13% lower than controls. There was an abnormal enlargement of alveolar ducts, which reduced by 24% the relative amount of air in the alveoli, compared to that in the alveolar ducts. The number of alveoli per lung and the mean volume of an alveolus were not different between the groups, but alveolar size class distributions were different, with significantly more very small and very large alveoli in 40-day-old rats after neonatal hyperoxia. By scanning electron microscopy, the alveolar surface of the exposed animals had a corrugated appearance, which was especially evident along alveolar ducts. Transmission electron microscopy revealed a greater density of capillaries, particularly in the alveolar regions close to terminal airways. Based on a random sample of the entire parenchymal region, capillary blood volume per cm2 of alveolar basal lamina was 18% greater. The results demonstrate that neonatal exposure to hyperoxia can cause abnormalities in the pulmonary alveolar and capillary structure of 40-day-old rats, and that these changes are similar to some features of broncho-pulmonary dysplasia.

Animals↗
PubMed1989

Postnatal growth of pulmonary acini and alveoli in normal and oxygen-exposed rats studied by serial section reconstructions.

Three-dimensional reconstructions from serial sections were used to examine postnatal lung development of rats reared in air (control) or oxygen. From birth to age 21 days, control lung volume increased ninefold, and the average volume of each ventilatory unit (all airspaces distal to a single respiratory bronchiole) increased seven times. There were approximately 5,000 ventilatory units at birth and on day 21, indicating that the lung grew by enlargement and subdivision of ventilatory units and not by their multiplication. Growth in hyperoxia (greater than 97%) for 7 days had no effect on the number of ventilatory units but, compared to controls, total lung volume and ventilatory unit volume were reduced 32% and 16%, respectively. At birth there were 0.6 x 10(6) alveoli, and at age 7 days in controls alveolar number increased 16-fold while the average volume of a single alveolus fell to one-sixth that at birth. Exposure to hyperoxia for 7 days stopped alveolarization; the surface area to volume ratio (Sa/V) of the ventilatory unit was lower, alveolar number was the same as at birth, and the alveoli present were large. At age 21 days, after 14 days of recovery in air, lung volume and ventilatory unit volume were greater than in controls but the Sa/V of the ventilatory unit was still depressed 20%. Alveoli from oxygen-exposed lungs were larger than in controls, and a greater size distribution coefficient showed them to be more variable. A shape coefficient for alveoli did not change as a function of the animal's age or oxygen treatment; it demonstrated proportional growth of alveolar height and diameter.

Animals↗
PubMed1989

Androgens amplify beta-adrenergic and FSH stimulation of granulosa cells.

The effects of androgens on granulosa cell stimulation by isoproterenol and follicle-stimulating hormone (FSH) were determined. Two functional parameters of granulosa cell stimulation were monitored: (a) activity of a transfected proopiomelanocortin (POMC) promoter and (b) production of progesterone. Treatment with the beta-adrenergic agonist, isoproterenol, stimulated steroidogenesis, and both isoproterenol and FSH appeared to enhance POMC promoter activity. The non-aromatizable androgen, 5 alpha-dihydrotestosterone (DHT), produced no effect on either parameter, but it potentiated the steroidogenic response to isoproterenol. Preliminary data also indicated a potentiation by DHT of the FSH-mediated increase in POMC promoter activity; results with a combination of DHT and isoproterenol were suggestive of potentiation. A possible role for androgen amplification of adrenergic stimulation in polycystic ovarian syndrome is discussed.

Androgens↗
PubMed1989

Cellular uptake and processing of surfactant lipids and apoprotein SP-A by rat lung.

The intracellular pathways and the kinetics of metabolism of surfactant apoprotein and lipid, which may be recycled from the alveolar space, are largely unknown. We used a lipid-apoprotein complex made from liposomes of pure lipids in a ratio found in mammalian pulmonary surfactant plus surfactant apoprotein (SP-A, Mr = 26,000-36,000) to test some possible relationships in the recycling of these major surfactant components between intrapulmonary compartments. After intratracheal instillation of 80 microliters of an apoprotein-liposome mixture with separate radiolabels in the lipid and the apoprotein, rats were killed at times from 8 min to 4 h later. The lungs were lavaged with saline, and subcellular fractions were isolated on discontinuous sucrose density gradients. Both the [14C]lipid radiolabel and the 125I-apoprotein radiolabel demonstrated a time-dependent increase in radioactivity recovered in a lamellar body-enriched fraction. Uptake of the radiolabels into other subcellular fractions did not exhibit a clear-cut time dependence; more of the protein than the lipid radiolabel was found in the Golgi-rich and microsomal fractions. We conclude that both the lipid and apoprotein portions of lung surfactant are taken up by lung cells and are incorporated into secretory granules of the cells.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1989

Gonadotropin regulation of the rat proopiomelanocortin promoter: characterization by transfection of primary ovarian granulosa cells.

To characterize the transcriptional effects of human (h)FSH and hCG on the POMC gene, primary rat granulosa cells were transiently transfected with a chloramphenicol acetyltransferase (CAT) reporter plasmid under the control of the POMC promoter and 5' region. POMC-CAT contains a fragment of the rat POMC gene, extending from nucleotide -704 to nucleotide +63, fused to the CAT gene. Treatment of POMC-CAT-transfected cells with either hFSH (20 ng/ml) or hCG (10 ng/ml) significantly increased CAT enzyme activity; however, neither hCG nor hFSH increased CAT enzyme activity in cells transfected with pSV2-CAT, a reporter plasmid under the control of the SV40 virus promoter and 5' region. The phosphodiesterase inhibitor isobutylmethylxanthine or the nonhydrolyzable cAMP analog cAMP-chlorothiophenyl significantly increased CAT activity in POMC-CAT-transfected granulosa cells. Human FSH stimulated transcription 10, 20, and 40 h after treatment, but FSH stimulation at the two earlier time points was 2.5- to 5.5-fold greater than that at 40 h. Gonadotropin-stimulated steroidogenesis was equivalent in POMC-CAT-transfected granulosa cells, untransfected, and mock-transfected cells. This indicates that transfection left the physiological hormone response intact. These data demonstrate the following. 1) 767 basepairs of the rat POMC gene are enough to confer gonadotropin stimulation on the CAT marker gene in granulosa cells. 2) Although the POMC promotor lacks a well conserved cAMP response element, either of two different pharmacological manipulations of granulosa cells that raise intracellular cAMP can also stimulate POMC-driven CAT expression. 3) Transfected primary cultures of granulosa cells provide a nontransformed, physiologically relevant model with which to study hormone-regulated gene expression.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-3-isobutylxanthine↗
PubMed1987

Alterations of surfactant pools in fetal and newborn rat lungs.

Preparation by the developing alveolar epithelium for the transition to air breathing and surfactant secretion at birth are critical components of neonatal survival. We combined morphometric analysis and biochemical assays of lung phospholipids to measure the amount and redistribution of lung surfactant during the perinatal period of rats. Within 10 min of the start of air breathing, there was a small increase in type II cell lamellar body content by morphometric and biochemical estimates. By 24 h, the whole lung and alveolar extracellular pool surfactant lipid had substantially increased. Subfractionation of the alveolar surfactant pool obtained at four times, from birth to 24 h of life, demonstrated a 20-fold increase in the ratio of phospholipid in a tubular myelin-rich fraction compared to a unilamellar vesicle-rich fraction. We conclude that packaging of surfactant may be very active immediately postbirth. Our results also indicate a major shift in the physical forms of extracellular surfactant during the first hours of air breathing.

Animals↗
PubMed1986

Dexamethasone increases adult rat lung surfactant lipids.

Prenatal administration of glucocorticoids stimulates epithelial cell maturation and induces a precocious development of pulmonary surfactant. The response of the adult lung to steroid administration is less well understood. We administered dexamethasone (2 mg X kg-1 X day-1) to adult male rats for 1 wk by daily subcutaneous injection. After pentobarbital anesthesia we lavaged the lungs and also isolated lamellar bodies from the tissue. Lipid analyses of the extracellular and intracellular surfactant compartments showed two- to fourfold greater amounts of total phospholipids and disaturated phosphatidylcholine compared with control. These changes were not found in kidney nor liver and were not present in plasma membrane, mitochondrial, or microsomal fractions from lungs. Morphometric analyses of the type II cells showed that anatomic measures of the lamellar body pool did not increase. We conclude that glucocorticoids have a significant effect to increase lung surfactant lipid pools of adult rat lungs by changing the phospholipid content of lamellar bodies, without changing lamellar body volume.

Animals↗
PubMed1986

The regulation of granulosa cell proopiomelanocortin messenger ribonucleic acid by androgens and gonadotropins.

We have previously demonstrated the presence of proopiomelanocortin (POMC) messenger RNA (mRNA) in rat granulosa cells. This study examines the unique, tissue-specific regulation of granulosa cell POMC mRNA levels by hormones with established regulatory effects on these cells. Northern blot analysis of immature rat ovarian RNA revealed the presence of a single mRNA of approximately 900 base pairs in length. The levels of ovarian POMC mRNA increased approximately 6-fold 48 h after priming with PMSG when expressed per microgram of total RNA. Granulosa cells were isolated from immature PMSG-primed rats and cultured under serum-free conditions in the presence and absence of various hormones to determine their effect on POMC mRNA levels. Treatment of the cells for 48 h with LH (100 ng/ml), androstenedione (10(-7) M), or LH plus androstenedione elevated POMC mRNA levels. LH alone elicited a 5-fold increase in POMC mRNA, and androstenedione elicited a 10-fold increase; the combination treatment led to a 47-fold increase. The effects of the nonaromatizable androgen dihydrotestosterone (DHT) were also evaluated. Treatment for 48 h with DHT (10(-7) M) elicited a 40-fold increase in POMC mRNA levels. In vivo experiments measuring ovarian POMC mRNA from immature female rats corroborated the in vitro results. Animals injected sc with DHT (500 micrograms) demonstrated 5-fold increases in ovarian POMC mRNA when expressed per microgram of total RNA. These studies provide both in vitro and in vivo evidence that granulosa cell POMC mRNA is under unique hormonal regulation by both androgens and gonadotropins.

Androgens↗
PubMed1986

Quantitative three-dimensional reconstruction and carbohydrate cytochemistry of rat nonciliated bronchiolar (Clara) cells.

We have employed three-dimensional reconstructions and cytochemistry to analyze nonciliated bronchiolar epithelial (Clara) cells from rat lungs. We identified a previously unreported ovoid granule type, which was characterized by its relative electron-lucency and eccentric electron-dense core. These electron-lucent granules were less common than electron-dense granules and were polarized toward the Golgi. They occupied 0.51% of the cell volume. Quantitative analysis of cell volume fractions showed the more numerous electron-dense granules to be polarized toward the air border; they occupied 2.5% of the cell volume. Periodic acid-thiocarbohydrazide-silver protein staining for hexose sugars indicated no differences between the granule types with regard to their carbohydrate content. Multivesicular bodies occupied 0.42% of total cell volume and were distinctly polarized toward the surface of the Golgi. Rod-shaped granules were present in 4 of the 6 reconstructed cells, ranging in number from 0 to 9 per cell. Nine apparent secretory events were seen in 6 cells and all appeared as exocytosis. We conclude that rat Clara cells contain at least 3 morphologically distinct granule types located in different volumes of the cell, and that exocytosis is the mode of secretion. The functional relationship between the ovoid granule types could include precursor-product lineage or different secretory products. We suggest that multivesicular bodies and light granules are stages in dark granule genesis.

Animals↗
PubMed1985

Arachidonic acid metabolism in cultured aortic endothelial cells. Effect of cAMP and 3-isobutyl-1-methylxanthine.

To investigate the hypothesis that cyclic AMP (cAMP) regulates arachidonic acid metabolism in vascular tissue, we have studied the effects of forskolin (FSK), an activator of adenylate cyclase, and 3-isobutyl-1-methylxanthine (IBMX), a phosphodiesterase inhibitor, on hormone-stimulated prostacyclin (PGI2) synthesis in porcine aortic endothelial cells grown in culture. In these experiments, bradykinin (1 microgram/ml) and A23187 (0.2 microM) potently stimulated PGI2 biosynthesis (9- and 10-fold respectively). However, prostaglandin synthesis in response to either of these agents was not affected by FSK even though FSK elevated intracellular levels of cAMP 10-fold. IBMX failed to elevate basal cAMP levels when incubated with unstimulated cells. Stimulation of IBMX-treated (0.1 but not 1.0 or 4.0 mM) cells with bradykinin, however, did result in increased cAMP levels, presumably due to PGI2 formation and subsequent activation of adenylate cyclase. In addition to phosphodiesterase inhibition, IBMX inhibited PGI2 formation (72% at 1 mM) in a dose-dependent manner so that, at higher doses of IBMX, cAMP levels returned to baseline. Thus, prostacyclin synthesis inhibition by IBMX could not be attributed to elevated cAMP. In other experiments, IBMX (1 mM) was found to directly inhibit arachidonic acid release (32%) and arachidonic acid metabolism (65%) in endothelial cells and to inhibit arachidonic acid conversion to PGE2 by sheep seminal vesicle microsomes (65%). These data suggest that IBMX directly inhibits both phospholipase and cyclooxygenase activities. These experiments do not support the contention that cAMP regulates these enzymes in cultured aortic endothelial cells.

1-Methyl-3-isobutylxanthine↗