Search PubMed⌕ Search

Biomedical subjects

R B Low

Publications and source records attributed to R B Low.

At least 91 records · Page 5Linked to original sources
PubMed1982

Effect of phalloidin on liver actin distribution, content, and turnover.

Phalloidin increases F-actin microfilament content and actin-directed immunofluorescence in hepatocytes in vivo and also increases actin polymerization and the stability of F-actin in vitro. We studied the sensitivity of immunofluorescent staining of actin to an actin depolymerizing factor (ADF) as well as actin content, degree of polymerization, and turnover in livers of in vivo phalloidin-treated rats. Pretreatment with ADF abolished anti-actin antibody (AAA) staining of normal liver but did not modify staining of livers from phalloidin-treated animals. Planimetric analyses of SDS-polyacrylamide gels showed the percent actin of total protein was increased by approximately 40% and the absolute amount of actin by approximately 43%, ten days after daily phalloidin treatment (50 micrograms/100 gm body weight). Similar but smaller changes could be seen after one day of treatment. Ultracentrifugational analyses of liver extracts indicated no change in the amount or proportion of G-actin but a 194% increase in the proportion of F-actin in ten-day treated animals, changes also apparent in one day animals. Neither the relative fractional rate of actin synthesis nor its synthesis as a percent of total protein synthesis was altered either at one-day or ten-day post-phalloidin treatment. Dualisotope experiments indicated that the rate of actin degradation was decreased selectively in the one- to three-day period following drug treatment. Thus, phalloidin appears to stabilize actin against the depolymerizing actions of ADF, increases the proportion of F-actin without altering the size of the G-actin pool, and causes accumulation of actin by decreasing its relative rate of degradation.

Actin Depolymerizing Factors↗
PubMed1982

Marked anterior displacement of the trachea and larynx from an esophageal obturator airway (EOA).

A 67-year-old man suffering cardiopulmonary arrest had an apparently uneventful placement of an esophageal obturator airway (EOA) by a well-trained ambulance crew. Subsequent clinical evaluation showed ventilation to be marginal. Attempts to insert an endotracheal tube were unsuccessful because the larynx could not be visualized. A subsequent postmortem examination showed that the EOA tube was kinked and bent back on itself so that the distal (balloon) end lay underneath the larynx, displacing it anteriorly.

Aged↗
PubMed1982

Macrophage phagocytosis: analysis of particle binding and internalization.

An improved radioassay for analysis of phagocytosis has been used to quantitate and characterize the binding and internalization of zymosan by monolayers of rabbit pulmonary alveolar macrophages. This method distinguishes zymosan particles reversibly bound to the cell surface from those internalized. The zymosan was radiolabeled with technetium 99m(99mTc), a gamma-emitter with a 6-h half-life. Use of 99mTc as the radiolabel also permitted simultaneous determinations of pinocytosis and cellular protein content using [14C]sucrose and [3H]amino acids, respectively. All endocytic data were normalized per adherent cell based on this latter measurement. A significant fraction of the cell-associated particles was bound to the cell surface but not internalized. Failure to correct for this compartment would have resulted in overestimation of phagocytic rate and total cellular capacity. Both binding and ingestion of functionally unopsonized zymosan were found to be saturable, temperature sensitive, dependent on glycolytic energy, dependent on a trypsin-sensitive membrane component, described by a maximal rate, and limited by a finite capacity. The time courses of both processes were found to be similar. These results led us to conclude that, in our system, both binding and internalization were active processes and that the limited capacity to ingest zymosan was not explained by a concomitant reduction in binding of the particle to the cell membrane. Furthermore, it was found that phagocytosis did not change the rate of fluid-phase pinocytosis, consistent with the concept that the cell membrane is a functional mosaic as has been previously found by others for phagocytic and transport sites in this cell type.

Amino Acids↗
PubMed1982

Insulin stimulation of protein synthesis in cultured skeletal and cardiac muscle cells.

The effects of acute exposure to insulin on protein synthesis were examined in primary, differentiated cultures of embryonic chick heart and skeletal muscle cells. Synthetic rates were calculated using the specific activity of tRNA-bound leucine as precursor, a specific activity that was significantly less than that of extracellular leucine but greater than that of free, intracellular leucine at 0.2 mM external leucine. Insulin did not alter these relationships. Doses of insulin in the physiological range produced significant stimulation of protein synthesis in both cell types. Maximal responses, involving approximately 30% increases in both absolute and fractional rates, were observed at higher insulin concentrations. Significant stimulation by insulin was seen in cardiac cells after only 1 h of insulin treatment, and the effects of the hormone were observed both in the presence and absence of serum in the culture medium.

Animals↗
PubMed1982

Altered myosin isozyme patterns from pressure-overloaded and thyrotoxic hypertrophied rabbit hearts.

Cardiac hypertrophy, induced by pressure overload, leads to a depression in the rate of force development, velocity of shortening, tension-dependent heat generation, and myosin ATPase activity, whereas cardiac hypertrophy, induced by thyroxine administration, leads to an increase in these parameters. These changes have been attributed, in part, to structural changes in myosin. In this study, we have investigated changes in the relative content of myosin isozymes and differences in primary structure of the isozymes in pressure-overloaded and thyrotoxic cardiac hypertrophy in the rabbit. Three myosin isozymic forms (V1 = fastest, V2 = intermediate, V3 = slowest mobility) were observed in pyrophosphate polyacrylamide gels from normal hearts with the V3 component being the predominant species. In the pressure-overloaded model, the V1 and V2 components disappeared or were present in reduced amounts leaving the V3 more predominant. The most striking difference was the isozymic profile produced in thyrotoxic hearts where the V1 became the predominant component and V2 and V3 the minor components. alpha-Chymotryptic digestion of myosin heavy chains produced characteristic, reproducible peptide patterns for each of the animal models, as did fluorographic analyses of alpha-chymotryptic digests of 14C-iodoacetamide (IAA)-labeled SH1 peptides of myosin. Our results suggest that altered proportions of myosin isozymes may be responsible for altered cardiac performance.

Adenosine Triphosphatases↗
PubMed1982

Increased contractility of isolated lung parenchyma in an animal model of pulmonary fibrosis induced by bleomycin.

The contractile capability of lung parenchymal strips isolated from normal rats was compared with that of strips isolated from rats with pulmonary fibrosis induced by intratracheal instillation of bleomycin sulfate. Subsequently, the population of contractile cells, both muscle and nonmuscle, was analyzed in each strip by histochemical, immunocytochemical, and ultrastructural means. The force (g force/g tissue wet weight) generated by fibrotic strips was approximately double that of the control strips in response to acetylcholine, epinephrine, and potassium depolarizing solution. The content of smooth muscle in fibrotic and control strips was not significantly different. Immunofluorescent examination indicated increased contractile proteins (actin and myosin) in the thickened alveolar walls of the fibrotic strips in areas devoid of histologically demonstrable smooth muscle. Ultrastructural examination of the fibrotic interstitium revealed an increased population of filament-laden cells that appeared related to the contractile interstitial cell, or myofibroblast. The results indicate that parenchymal strips isolated from fibrotic rat lung can generate increased force in vitro. These responses may be related to increased nonmuscle contractile cells in the fibrotic interstitium.

Animals↗
PubMed1982

Analyses of sequential bronchoalveolar lavage samples from healthy human volunteers.

Subsegmental bronchoalveolar lavage of human subjects provides an accessible sample of cells and secretions from the lower respiratory tract for clinical and research study. Technical factors that may modify lavage results have received little attention. The effects of lavage volume on the patterns of recovered cells and biochemical components were studied by separately analyzing each of 4 sequential 60-ml syringes of saline instilled into a subsegment of the right middle lobe. Fourteen healthy normal subjects volunteered for lavage. Recovery of cells from both nonsmokers and smokers increased and then remained essentially constant in subsequent syringes, suggesting elution of adherent macrophages from the alveolar surface. Protein, carbohydrate, lipid, and potassium appeared in decreasing concentrations in sequential syringes, but different constituents followed separate patterns. Simple mixing models do not explain the observed results. Measured values for the components in a "small" 120-ml lavage may be substantially different from the values in a "large" 240-ml lavage. Further study of the kinetics of recovery of selected lavage components may provide better understanding of the origins and functions of these substances in the lung.

Adolescent↗
PubMed1982

Use of radioisotopes in quantitative studies of lung metabolism.

Quantitatively accurate studies of macromolecule and lipid synthesis in lung and other tissues by using radioactive substrates require detailed knowledge of the specific radioactivity of the appropriate pool of precursor molecules serving the synthetic pathway. A brief summary is provided of how considerations of precursor availability, metabolism, and compartmentation, as well as product remodeling, may affect the accuracy with which rates of protein, DNA, RNA, and lipid synthesis can be measured. Where possible, the application of this material to studies of lung metabolism is discussed, along with approaches that may minimize experimental uncertainties.

Amino Acids↗
PubMed1981

Prolyl-tRNA-based rates of protein and collagen synthesis in human lung fibroblasts.

Knowledge of the dynamics of collagen turnover requires information regarding rates of synthesis of this group of connective-tissue proteins. The relationship of various amino acid pools to the tRNA precursor pool used for protein synthesis is known to vary between different cell types and tissues, even for essential amino acids. We studied extracellular, intracellular and tRNA-proline pools in cultured human lung IMR-90 fibroblasts to determine the relationship between them as candidate proline precursor pools for total protein and collagen synthesis. Time-course experiments showed that the three proline pools attained distinctly different steady-state specific radioactivities (extracellular greater than intracellular greater than tRNA) at the extracellular proline concentration of 0.2 mM. The kinetics of radioisotope incorporation into cell protein and collagenase-digestible protein indicated that the intracellular free proline pool could not be used reliably as a precursor for calculating synthetic rates. However, tRNA-proline behaved isotopically as if it were the precursor and provided synthesis rates 2-3-fold higher than those calculated by using either free proline pool. The incorporation of labelled lysine and leucine was constant over a wide range of extracellular proline concentrations. Fractional rates of protein synthesis based on tRNA-amino acid were the same with [3H]phenylalanine as with [3H]proline. The specific radioactivity of cell-associated hydroxyproline reached a steady-state value 8-10h after radioisotope administration which matched the mean tRNA-proline specific radioactivity, suggesting that tRNA-proline is not isotopically compartmentalized. A model of cellular proline-pool relationship is presented and discussed.

Amino Acids↗
PubMed1981

Protein synthesis in pulmonary alveolar macrophages. Source of amino acids for leucyl-tRNA.

Extracellular, intracellular and tRNA-bound leucine pools of the adherent pulmonary alveolar macrophage were examined to determine the relationships between them and the precursor for protein synthesis. When cells were cultured in media of various leucine concentrations, the patterns of isotope distribution in intracellular and extracellular leucine did not correlate with the patterns seen in protein-bound leucine. hence, the free leucine pools cannot be used reliably as precursors for calculating rates of protein synthesis. tRNA-bound leucine, however, behaved isotopically as if it were the precursor. Constant synthetic rates were calculated using the tRNA specific activity over a wide range of leucine concentrations. In addition, by measuring the tRNA-bound specific activities of three different amino acids, leucine, valine and phenylalanine, and their respective specific activities in protein, we were able to calculate independently three separate but identical synthetic rates. At physiological amino acid concentrations, the macrophage intracellular leucine pool and the tRNA-bound leucine pool received less than half of their amino acids from extracellular sources. At 5 mM external leucine, the intracellular specific activity was indistinguishable from that of the medium leucine, but the specific activity of the tRNA-bound leucine pool remained only about 50% that of the extracellular value. The most straightforward interpretation of why the tRNA-bound leucine did not flood with external label under conditions where the intracellular pool has reached equilibrium is to propose that some portion of the leucine for protein synthesis is derived directly from protein turnover before the degradation products have mixed with the common amino acid pool.

Animals↗
PubMed1981

Exocytosis of pinocytosed fluid in cultured cells: kinetic evidence for rapid turnover and compartmentation.

The uptake and fate of pinocytosed fluid were investigated in monolayers of pulmonary alveolar macrophages and fetal lung fibroblasts using the fluid-phase marker, [14C]sucrose. Initial experiments revealed that cellular accumulation of chromatographically repurified [14C]sucrose was not linear with incubation time. Deviation from linearity was shown to be due to constant exocytosis of accumulating marker. Chromatographic analysis revealed that the cells were unable to metabolize sucrose and were releasing it intact by a process that was temperature-sensitive but not dependent on extracellular calcium and magnesium. A detailed analysis of the kinetics of exocytosis was undertaken by preloading cells with [14C]sucrose for various lengths of time and then monitoring the appearance of radioactivity into isotope-free medium. Results indicated that modeling the process of fluid-phase pinocytosis and subsequent exocytosis required at least two intracellular compartments in series, one compartment being of small size and turning over very rapidly (t1/2 = 5 min in macrophages, 6--8 min in fibroblasts) and the other compartment being apparently larger in size and turning over very slowly (t1/2 = 180 min in macrophages, 430--620 min in fibroblasts). Computer-simulation based on this model confirmed that the kinetics of efflux faithfully reflected the kinetics of influx and that the rate of efflux completely accounted for the deviation from linearity of accumulation kinetics. Moreover, the sizes of the compartments and magnitude of the intercompartment fluxes were such that the majority of fluid internalized in pinocytic vesicles was rapidly returned to the extracellular space via exocytosis. This result provides direct experimental evidence for a process previously thought necessary based solely on morphological and theoretical considerations. Furthermore, the turnover of pinocytosed fluid was so dynamic that accumulation deviated from linearity even within the first few minutes of incubation. We were able to show that the kinetics of exocytosis allowed calculation of the actual pinocytic rate, a rate that was nearly 50% greater than the apparent initial rate obtained from the slope of the uptake curve over the first 10 min.

Animals↗
PubMed1981

Organization of actin in epithelial cells during regenerative and neoplastic conditions. Correlation of morphologic, immunofluorescent, and biochemical findings.

To study the distribution and the degree of polymerization of actin in regenerating and neoplastic cells we have examined: (1) the immunofluorescent staining of these cells with human antiactin antibodies (AAA), (2) the sensitivity of cellular actin to AAA staining after treatment with the actin-depolymerizing factor present in plasma or serum of several animal species, and (3) the total and relative amounts of F- and G-actin in tissue preparations, determined by means of differential centrifugation and densitometric analysis of stained sodium dodecyl sulfate-polyacrylamide gels. There was little or no difference in staining intensity between normal and regenerating or tumoral tissues when treated with AAA, but AAA staining of normal tissues was abolished, whereas staining of regenerating and tumoral tissues remained intense after incubation with actin-depolymerizing factor. Gel analysis showed that normal, regenerating, and neoplastic epidermal cells contained similar amounts of actin. Compared to controls, regenerating liver tissue contained slightly more total actin, nearly the same amount of G-actin, but a substantially increased amount of F-actin. These results suggest that: (1) compared to controls, a greater proportion of regenerating and neoplastic tissue actin is stabilized against the action of actin-depolymerizing factor; (2) changes involving actin in regenerating and neoplastic epithelial cells reflect changes in its degree of polymerization rather than its total amount.

Actins↗
PubMed1979

Isolation and characterization of fibroblasts obtained by pulmonary lavage of human subjects.

Cells that possess the morphology and collagen synthetic capacity of fibroblasts were recovered by bronchofiberscopic subsegmental pulmonary lavage from patients with pulmonary fibrosis, from patients with miscellaneous nonfibrotic lung diseases and from healthy volunteers. Lavage cells were placed in tissue culture, observed for 2 to 6 weeks, and compared with human lavage pulmonary alveolar macrophages (PAM), WI-38 and IMR-90 human fetal lung fibroblasts, and adult lung tissue fibroblasts (CLAC-76). Lavage fibroblsts (LF) were identified as proliferating clones in monolayers of nonproliferating PAM and could be subcultured repeatedly. Fibroglasts were propagated from 28 of the 92 lavage specimens cultured. Time-lapse cinematography showed similar distributions of interdivision times for LF, CLAC-76 and WI-38, but the LF and CLAC-76 lines had slower mean migration rates than the fetal line. Light, scanning, and transmission electron microscopy of LF showed attenuated spindle-shaped cells with interdigitating filopodia, flat surfaces with few microvilli, and containing numerous cytoplasmic polyribosomes and rough endoplasmic reticulum. Extracellular fibrils with the appearance of collagen were seen. Collagen synthesis by LF was measured as 3.9% to 4.9% of the cell-associated protein sensitive to bacterial collagenase. This protein was rich in hydroxyproline, and had an electrophoretic migration pattern identical to known collagen. LF did not contain lysozyme although this enzyme was abundant in fresh and 1-week cultured PAM. Thus LF were similar to human fetal and adult lung tissue fibroblasts in their morphology, tissue culture characteristics, constitutive enzymes and collagen synthetic properties but were distinctly different from PAM.

Cell Line↗
PubMed1978

The relationship of mechanical Vmax to myosin ATPase activity in rabbit and marmot ventricular muscle.

Papillary muscle mechanics and ventricular myosin calcium-activated ATPase activity were measured in the same heart as a function of temperature (8--28 degrees) in rabbits and marmots, in order to examine further the hypothesis that the velocity of cardiac muscle shortening at zero load (Vmax) is correlated with myosin ATPase activity. There was a similar Q10 for Vmax in each muscle type, as measured with isotonic afterloaded quick-releases at 30--33% time-to-peak tension; the calcium activated ATPase of myosin in the two muscle types also was similar. The least squares linear regression of rabbit Vmax on calcium-activated myosin ATPase activity was the same as in the marmot, so all the data were pooled to yield a linear regression (Y = 0.47 +/- 3.82X) with a high correlation between the two variables [r = 0.95, P less than 0.01 (ANOV)]. Furthermore, the correlation proved to be predictive of cardiac Vmax and myosin ATPase activity levels in other experiments where these two measurements decreased below normal as a result of hypertrophic growth. Consequently, the quantitative relationship between Vmax and myosin ATPase defined here may prove to be predictive of the ability of cardiac muscle to release bond energy.

Animals↗