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Biomedical subjects

D M Ward

Publications and source records attributed to D M Ward.

At least 19 recordsLinked to original sources
PubMed1992

Uncultivated cyanobacteria, Chloroflexus-like inhabitants, and spirochete-like inhabitants of a hot spring microbial mat.

Analysis of 16S rRNA sequences retrieved as cDNA (16S rcDNA) from the Octopus Spring cyanobacterial mat has permitted phylogenetic characterization of some uncultivated community members, expanding our knowledge or diversity within this microbial community. Two new cyanobacterial 16S rRNA sequences were discovered, raising to four the number of cyanobacterial sequence types known to occur in the mat. None of the sequences found is that of the cultivated thermophilic cyanobacterium Synechococcus lividus. A new 16S rRNA sequence characteristic of green nonsulfur bacteria and their relatives was discovered, raising to two the number of such sequences known to exist in the mat. Both are unique among the 16S rRNA sequences of cultivated members of this group, including an Octopus Spring isolate of Chloroflexus aurantiacus and Heliothrix oregonensis, whose sequences we report herein. Two spirochete-like 16S rRNA sequences were discovered. One can be placed in the leptospira subdivision of the spirochete group, but the other has such a loose affiliation with the spirochete group that it might actually belong to an as yet unrecognized subdivision or even to a new eubacterial line of descent.

Bacteria↗
PubMed1992

Sex-associated differences in left ventricular function in aortic stenosis of the elderly.

BACKGROUND: In aortic stenosis, the response of the left ventricle to pressure overload varies from compensated hypertrophy to overt heart failure. The determinants of left ventricular adaptation are poorly understood. METHODS AND RESULTS: Left ventricular function was compared to assess the role of sex in 34 women and 29 men 60 years or older with both hemodynamic and echocardiographic data characteristic of severe aortic stenosis and no important coronary artery disease. Despite a similar degree of left ventricular outflow obstruction in women versus men (aortic valve area 0.54 +/- 0.20 versus 0.59 +/- 0.19 cm2, NS), the left ventricle of women had a greater fractional shortening (37 +/- 12 versus 25 +/- 12%, p = 0.001), achieved a smaller end-systolic chamber size (1.82 +/- 0.64 versus 2.17 +/- 0.65 cm/m2, p = 0.04), and generated more pressure (210 +/- 35 versus 182 +/- 29 mm Hg, p = 0.001) with a greater maximum positive dP/dt (2.153 +/- 794 versus 1,595 +/- 384 mm Hg/sec, p = 0.02). The men had a lower cardiac index (2.12 +/- 0.59 versus 2.49 +/- 0.63 l/min/m2, p = 0.02), higher mean pulmonary artery pressure (35 +/- 13 versus 27 +/- 10 mm Hg, p = 0.01), and shorter ejection period (340 +/- 40 versus 370 +/- 40 msec, p = 0.02). Women and men were equally symptomatic. Supernormal left ventricular ejection performance was present in 41% of the women and only 14% of the men (p = 0.002). This subgroup of women had a small, thick-walled chamber (end-diastolic radius to thickness ratio, 1.58 +/- 0.52 versus 2.45 +/- 0.51 in control women, p = 0.01) with low end-systolic wall stress. Subnormal ejection performance was present in 64% of the men and only 18% of the women (p = 0.002). This subgroup of men had an increased chamber size and high end-systolic wall stress compared with control men. Greater left ventricular mass was present in men compared with women (211 +/- 55 versus 179 +/- 55 g/m2, p = 0.03). CONCLUSIONS: Sex is a factor in left ventricular adaptation to valvular aortic stenosis in adults 60 years or older.

Aged↗
PubMed1992

Sequential cycles of high-dose carboplatin administered with recombinant human granulocyte-macrophage colony-stimulating factor and repeated infusions of autologous peripheral-blood progenitor cells: a novel and effective method for delivering multiple courses of dose-intensive therapy.

PURPOSE: The trial was undertaken to study the effect of administering granulocyte-macrophage colony-stimulating factor (GM-CSF) with and without peripheral-blood progenitor cells (PBPC) on the hematologic and nonhematologic toxicity observed with multiple cycles of high-dose carboplatin chemotherapy. PATIENTS AND METHODS: Eighteen patients with a variety of solid tumors received a total of 40 cycles of carboplatin, 1,200 mg/m2 per cycle, administered by continuous infusion over 96 hours. All 40 courses were administered with a daily 4-hour intravenous (IV) infusion of either 5 or 10 micrograms/kg/d of recombinant human Escherichia coli-derived GM-CSF. The first 20 courses were administered without PBPC support (treatment A). Because of severe neutropenia and thrombocytopenia, the next 20 courses of therapy were administered with GM-CSF, PBPC, and oral antibiotic prophylaxis (treatment B). RESULTS: The addition of PBPC support led to a significant reduction in the duration of neutropenia (10.5 v 7.5 days; P = .027) and thrombocytopenia (12.4 v 5.2 days; P = .001), number of RBC transfusions (six v three; P = .01) and platelet transfusions (10.3 v 3.7; P = .013), number of hospital days (12.6 v 2.9; P = .01), and days of IV antibiotics (11.8 v 2.4; P = .007) per cycle. Significant increases in the weekly dose intensity (206 v 285 mg/m2/wk; P = .014) and total dose (2,287 v 3,600 mg/m2; P = .018) of carboplatin delivered were also observed with treatment B. The overall response rate in this study was 70%, with 11 of 16 assessable patients achieving either a complete (three patients) or partial (eight patients) remission. CONCLUSION: This combination of GM-CSF and PBPC infusion represents an effective method for delivering multiple cycles of high-dose carboplatin chemotherapy and may serve as a model for the administration of high-dose chemotherapy in future trials.

Adult↗
PubMed1991

16S rRNA sequences of uncultivated hot spring cyanobacterial mat inhabitants retrieved as randomly primed cDNA.

Cloning and analysis of cDNAs synthesized from rRNAs is one approach to assess the species composition of natural microbial communities. In some earlier attempts to synthesize cDNA from 16S rRNA (16S rcDNA) from the Octopus Spring cyanobacterial mat, a dominance of short 16S rcDNAs was observed, which appear to have originated only from certain organisms. Priming of cDNA synthesis from small ribosomal subunit RNA with random deoxyhexanucleotides can retrieve longer sequences, more suitable for phylogenetic analysis. Here we report the retrieval of 16S rRNA sequences from three formerly uncultured community members. One sequence type, which was retrieved three times from a total of five sequences analyzed, can be placed in the cyanobacterial phylum. A second sequence type is related to 16S rRNAs from green nonsulfur bacteria. The third sequence type may represent a novel phylogenetic type.

Base Sequence↗
PubMed1991

Recent advances in continuous renal replacement therapy: citrate anticoagulated continuous arteriovenous hemodialysis.

Trisodium citrate was used as a regional anticoagulant on 24 patients on continuous arteriovenous hemodialysis (CAVHD), obviating the need for systemic heparinization. Principles of CAVHD, potential complications, and nursing responsibilities are addressed. Clearances, blood flow rate, ultrafiltration and filter patency compare favorably with heparin CAVHD. Citrate anticoagulated CAVHD avoids heparin-associated complications in the critically ill uremic patient.

Acute Kidney Injury↗
PubMed1990

The rate of internalization of different receptor-ligand complexes in alveolar macrophages is receptor-specific.

To probe the mechanisms of endocytosis in alveolar macrophages, we examined the internalization rates of three different receptors. Initial rates of internalization for mannosylated BSA, diferric transferrin and alpha-macroglobulin-proteinase complexes were all different. Although the absolute rates of internalization varied depending on the cell preparation, transferrin was internalized at 10-20% and alpha-macroglobulin-proteinase complex at 40-60% of the rate of manosylated-BSA. Incubation of cells with transferrin did not affect the rate of internalization of mannosylated BSA or alpha-macroglobulin-proteinase complexes, and the rates of internalization were independent of receptor occupancy. These different internalization rates could not be ascribed to different rates of diacytosis. Altering the distribution of unoccupied surface receptors by either trypsin treatment of cells at 0 degree C or exposure to hyperosmotic solutions resulted in the absolute internalization rates being affected by the experimental condition, but the hierarchy in receptor internalization rates was maintained. The fact that a variety of conditions affect receptor internalization rates to the same degree implies the existence of co-ordinate regulation at a single rate-limiting step. Based on these results, we suggest that differences in internalization rate reflect the ability of ligand-receptor complexes to be captured by coated pits.

Animals↗
PubMed1990

16S rRNA sequences reveal numerous uncultured microorganisms in a natural community.

Microbiologists have been constrained in their efforts to describe the compositions of natural microbial communities using traditional methods. Few microorganisms have sufficiently distinctive morphology to be recognized by microscopy. Culture-dependent methods are biased, as a microorganism can be cultivated only after its physiological niche is perceived and duplicated experimentally. It is therefore widely believed that fewer than 20% of the extant microorganisms have been discovered, and that culture methods are inadequate for studying microbial community composition. In view of the physiological and phylogenetic diversity among microorganisms, speculation that 80% or more of microbes remain undiscovered raises the question of how well we know the Earth's biota and its biochemical potential. We have performed a culture-independent analysis of the composition of a well-studied hot spring microbial community, using a common but distinctive cellular component, 16S ribosomal RNA. Our results confirm speculations about the diversity of uncultured microorganisms it contains.

Base Sequence↗
PubMed1990

Inhibition of late endosome-lysosome fusion: studies on the mechanism by which isotonic-K+ buffers alter intracellular ligand movement.

Incubation of alveolar macrophages or hepatocytes in media in which Na+ is replaced by K+ ("isotonic-K buffer") inhibited the movement of internalized ligand from late endosomes to lysosomes (Ward et al.: Journal of Cell Biology 110:1013-1022, 1990). In this study we investigate the mechanism responsible for the isotonic-K+ block in movement of ligand from late endosomes to lysosomes. We observed that iso-K+ inhibition of endosome-lysosome fusion is not unique to alveolar macrophages or hepatocytes but can be seen in a variety of cell types including J774 and Hela cells. The inhibition in intracellular ligand movement was time dependent with the maximum change occurring after 60 minutes. Once established the inhibition resulted in a prolonged and apparently permanent decrease in vesicle movement. Cells were able to recover from the effects of iso-K+ buffers over a time course of 5-10 minutes when placed back in Na(+)-containing media. The effect of iso-K+ buffers was independent of intracellular pH changes and appeared to involve cell swelling. When cells were incubated in iso-K+ buffers under conditions in which cell volume changes were reduced, intracellular ligand movement approached normal levels. Such conditions included replacing Cl- with the less permeant anion gluconate, and by addition of sucrose to isotonic-K+ buffers. Analysis of the mechanism by which changes in cell volume could alter intracellular movement ruled out changes in cyclic nucleotides, Ca2+, or microtubules. These results suggest that changes in cell shape or volume can alter intracellular transport systems by novel routes.

Animals↗
PubMed1990

Temporomandibular synovial fluid pressure response to altered mandibular positions.

Hydrostatic synovial fluid pressure within the superior aspect of the temporomandibular joint space of the growing pig, Sus scrofa domesticus, was examined in response to various acute and chronic alterations of mandibular position. Bilateral measurements of pressure were recorded with chronically implanted wick catheters in three 8-week-old pigs before and at the time of appliance placement and then at 2-day intervals until the animals were 20 weeks old. Besides confirming the observations of a previous study, we noted that forward positioning of the mandible caused an increase in synovial fluid pressure that decayed to baseline levels within 2 hours. Posterior positioning of the mandible effected a larger increase in pressure that partially decayed over 2 hours but did not return to baseline levels over the entire course of the experiment. These results may have significance in the understanding of the physiology of the temporomandibular joint, local control mechanisms involved in the normal and treatment-aided growth of the mandible, and disorders of the temporomandibular joint.

Animals↗
PubMed1990

Fusion of sequentially internalized vesicles in alveolar macrophages.

Previously we reported that internalized ligand-receptor complexes are transported within the alveolar macrophage at a rate that is independent of the ligand and/or receptor but is dependent on the endocytic apparatus (Ward, D. M., R. S. Ajioka, and J. Kaplan. 1989. J. Biol. Chem. 264:8164-8170). To probe the mechanism of intracellular vesicle transport, we examined the ability of vesicles internalized at different times to fuse. The mixing of ligands internalized at different times was studied using the 3,3'-diaminobenzidine/horseradish peroxidase density shift technique. The ability of internalized vesicles to fuse was dependent upon their location in the endocytic pathway. When ligands were administered as tandem pulses a significant amount of mixing (20-40%) of vesicular contents was observed. The pattern of mixing was independent of the ligands employed (transferrin, mannosylated BSA, or alpha macroglobulin), the order of ligand addition, and temperature (37 degrees C or 28 degrees C). Fusion was restricted to a brief period immediately after internalization. The amount of fusion in early endosomes did not increase when cells, given tandem pulses, were chased such that the ligands further traversed the early endocytic pathway. Little fusion, also, was seen when a chase was interposed between the two ligand pulses. The temporal segregation of vesicle contents seen in early endosomes was lost within late endosomes. Extensive mixing of vesicle contents was observed in the later portion of the endocytic pathway. This portion of the pathway is defined by the absence of internalized transferrin and is composed of ligands en route to lysosomes. Incubation of cells in iso-osmotic medium in which Na+ was replaced by K+ inhibited movement of internalized ligands to the lysosome, resulting in ligand accumulation within the late endocytic pathway. The accumulation of ligand was correlated with extensive mixing of sequentially internalized ligands. Although significant amounts of ligand degradation were observed, this compartment was devoid of conventional lysosomal markers such as acid glycosidases. These results indicate changing patterns of vesicle fusion within the endocytic pathway, with a complete loss of temporal ligand segregation in a prelysosomal compartment.

Animals↗
PubMed1990

16S rRNA sequences reveal uncultured inhabitants of a well-studied thermal community.

Molecular methods are beginning to reveal inhabitants of natural microbial communities which have never before been cultured. Our approach involves selective cloning of naturally occurring 16S rRNA sequences as cDNA, and comparison of these sequences to a database which includes 16S rRNA sequences of isolated community members. We provide here an overview of the method and its potential for community analysis. A 16S rRNA sequence retrieved from the well-studied hot spring cyanobacterial mat in Octopus Spring (Yellowstone National Park) is shown as an example of one contributed by an uncultured member of the community.

Base Sequence↗
PubMed1990

Movement of receptors and ligands through the endocytic apparatus in alveolar macrophages.

The ability of alveolar macrophages to remove potentially injurious agents is essential for normal lung function. One mechanism by which macrophages affect this function is by the internalization and clearance of noxious agents from lung fluid. Extracellular macromolecules may be removed as a result of nonspecific uptake via fluid-phase pinocytosis or as a result of binding to surface receptors followed by a highly efficient internalization of the receptor-ligand complex. The ability of macrophages to bind, internalize, and thus sequester extracellular macromolecules is facilitated by the fact that membrane components involved in this process may be reutilized. That is, once internalized, membrane components are not invariably degraded but can be recycled for further use. In this commentary we examine some of the features known about the recycling process in both macrophages and other cell types. We demonstrate that although the rate of internalization of a membrane component is unique to that component, the rate of movement of internalized components within cells is a function of the endocytic apparatus and is independent of the ligands or receptors contained in the internalized vesicle. We also demonstrate how alterations in the rates of recycling may result in changes in the distribution of membrane proteins expressed on the cell surface and affect the ability of a cell to respond to the environment.

Animals↗
PubMed1989

Cohort movement of different ligands and receptors in the intracellular endocytic pathway of alveolar macrophages.

The rate of movement of different receptors and ligands through the intracellular endocytic apparatus was studied in alveolar macrophages. Cells were exposed to iodinated alpha-macroglobulin-protease complexes, mannose terminal glycoproteins, diferric transferrin, and maleylated proteins. By use of the diaminobenzidine density shift procedure, we demonstrated that these ligands were internalized into the same endocytic vesicle. We then compared the rates of transfer to the lysosome or recycling to the cell surface of different ligands/receptors contained in the same endosome. We found that although the rate constant for degradation was ligand specific, the lag time prior to the initiation of degradation was the same for all three ligands. We also found that molecules taken up nonspecifically by fluid-phase pinocytosis had the same lag time prior to degradation as ligands internalized via receptor-mediated endocytosis. These data suggest that different molecules within the same endocytic compartment are transferred to the lysosome (or degradative compartment) at the same rate. We measured the rate of return of receptors to the cell surface by either inactivating surface receptors by protease treatment at 0 degrees C, or by incubating cells with saturating amounts of nonradioactive ligand at 37 degrees C. We then measured the rate of appearance of "new" receptors on the cell surface. Using these approaches, we found that three different receptors were transferred from internal pools to the cell surface at the same rate. The rate of transfer was independent of whether receptors were initially occupied or unoccupied. Our observations indicate that receptor/ligands, once inside alveolar macrophages, are transported by vesicles which transfer their contents as a cohort from one compartment to another. The rate of movement of these receptors is determined by the movement of vesicles and is independent of their content.

3,3'-Diaminobenzidine↗
PubMed1988

Effect of hypo-osmotic incubation on membrane recycling.

Incubation of alveolar macrophages in hypo-osmotic media causes a time-and temperature-dependent increase in the number of surface receptors for three different ligands. Exposure of cells to solutions of 210 mOsM or less, at 37 degrees C but not at 0 degree C, resulted in an increase in the number of surface receptors for diferric transferrin, alpha-macroglobulin-protease complexes, and mannose-terminated glycoproteins. Upon media dilution at 37 degrees C, surface receptor number reached a maximum within 5 min and returned to near-normal values by 30 min. The increase in surface receptor number was the result of a decrease in the rate of internalization of receptors, either occupied or unoccupied. The rate of receptor exteriorization was unaltered by hypo-osmotic incubation of cells. The rate of fluid-phase pinocytosis was also inhibited upon incubation in hypo-osmotic solution. In experiments in which both receptor-mediated endocytosis and fluid phase pinocytosis were measured on the same samples, inhibition of both processes occurred with the same kinetics and to a similar extent. The rate of receptor-mediated endocytosis recovered to normal rates after 60 min in hypo-osmotic solutions, whereas the rate of fluid phase pinocytosis did not recover to the same extent.

Animals↗
PubMed1988

Regulatory volume decrease in alveolar macrophages: cation loss is not correlated with changes in membrane recycling.

Alveolar macrophages regain their normal volume after swelling in hypo-osmotic solutions. This process, termed regulatory volume decrease (RVD), is initiated 3-5 minutes after exposure of cells to hypo-osmotic solutions, and by 30 min, near-normal volumes are attained. Volume decrease does not occur at 0 degrees C or in solutions in which Na+ has been replaced by K+, or Cl- by the impermeant anion gluconate. These results, as well as direct measurement of intracellular cations, indicate that decreases in cell volume result primarily from the loss of K+ and Cl- and are similar to RVD in lymphocytes. Kinetic analysis of cation loss, both by directly measuring changes in intracellular cation content and by assaying rubidium efflux, showed that cation loss occurred immediately upon media dilution. The rate of cation loss fit first-order kinetics and preceded both the initiation of volume decrease and the maximum increase in surface receptor number. These results suggest that the cation transporters responsible for RVD are located at the cell surface and that regulation of activity is not dependent on alterations in membrane movement.

Animals↗
PubMed1988

Atrial natriuretic factor in renal failure and posthemodialytic postural hypotension.

Atrial natriuretic factor (ANF) levels were ten times normal in hemodialysis patients before dialysis. ANF was not cleared by the dialyzer membrane but plasma levels decreased 47% by the end of dialysis. Patients undergoing peritoneal dialysis had plasma ANF levels four times normal and had detectable ANF in their dialysate. Hemodialysis patients with a marked fall in BP after dialysis had higher ANF levels (P less than 0.05) and lower norepinephrine (NE) levels (P less than 0.05) associated with a failure to increase NE in response to dialysis. Elevated ANF levels are associated with postdialysis hypotension in hemodialysis patients.

Adult↗