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Biomedical subjects

D L Dyer

Publications and source records attributed to D L Dyer.

At least 19 recordsLinked to original sources
PubMed2000

Alcohol-free instant hand sanitizer reduces elementary school illness absenteeism.

UNLABELLED: BACKGROUND AND HYPOTHESES: A substantial percentage of school absenteeism among children is related to transmissible infection. Rates of transmission can be reduced by hand washing with soap and water, but such washing occurs infrequently. This study tested whether an alcohol-free instant hand sanitizer (CleanHands) could reduce illness absenteeism in school-age children. METHODS: A 10-week, open-label, crossover study was performed on 420 elementary school-age children (ages 5-12). Students were given a brief orientation immediately prior to the start of the study on the relationship of germs, illness, and hand washing. Each student in the treatment group then received the test product in individual bottles, with instructions to apply one to two sprays to the hands after coming into the classroom, before eating, and after using the restroom, in addition to their normal hand washing with soap and water. The control group was instructed to continue hand washing as normal with non-medicated soap. After 4 weeks of treatment and a 2-week wash-out period, the control and experimental groups were reversed. Data gathered on absenteeism were classified as gastrointestinal or respiratory related and normalized for nonillness-related absenteeism and school holidays. RESULTS: Compared to the hand washing-only control group, students using CleanHands were found to have 41.9% fewer illness-related absence days, representing a 28.9% and a 49.7% drop in gastrointestinal- and respiratory-related illnesses, respectively. Likewise, absence incidence decreased by 31.7%, consisting of a 44.2% and 50.2% decrease in incidence of gastrointestinal- and respiratory-related illnesses, respectively. No adverse events were reported during the study. CONCLUSIONS: Daily use of the instant hand sanitizer was associated with significantly lower rates of illness-related absenteeism.

Absenteeism↗
PubMed1999

Antithrombotic efficacy of thrombin inhibitor L-374,087: intravenous activity in a primate model of venous thrombus extension and oral activity in a canine model of primary venous and coronary artery thrombosis.

The small molecule direct thrombin inhibitor L-374,087 was characterized across species in an in vitro activated partial thromboplastin clotting time (aPTT) assay and in vivo in rhesus monkey and dog thrombosis models. In vitro in rhesus, dog, and human plasma, L-374,087 concentrations eliciting 2-fold increases in aPTT were 0.25, 1.9, and 0.28 microM, respectively. In anesthetized rhesus monkeys, 300 microgram/kg bolus plus 12 microgram/kg/min and 300 microgram/kg bolus plus 30 microgram/kg/min L-374,087 i.v. infusions significantly reduced jugular vein thrombus extension, with both regimens limiting venous thrombus extension to 2-fold that of baseline thrombus mass compared with a 5-fold extension observed in the vehicle control group. Antithrombotic efficacy in the rhesus with the lower-dose regimen was achieved with 2.3- to 2.4-fold increases in aPTT and prothrombin time. In a conscious instrumented dog model of electrolytic vessel injury, the oral administration of two 10 mg/kg L-374,087 doses 12 h apart significantly reduced jugular vein thrombus mass, reduced the incidence of and delayed time to occlusive coronary artery thrombosis, and significantly reduced coronary artery thrombus mass and ensuing posterolateral myocardial infarct size. Antithrombotic efficacy in the dog was achieved with 1.6- to 2.0-fold increases in aPTT at 1 to 6 h after oral dosing with L-374,087. These results indicate significant antithrombotic efficacy against both venous and coronary arterial thrombosis with L-374,087 with only moderate elevations in aPTT or prothrombin time. The oral efficacy of L-374,087 characterizes this compound as a prototype for the further development of orally active direct thrombin inhibitors.

Administration, Oral↗
PubMed1998

Efficacious, orally bioavailable thrombin inhibitors based on 3-aminopyridinone or 3-aminopyrazinone acetamide peptidomimetic templates.

We have addressed the key deficiency of noncovalent pyridinone acetamide thrombin inhibitor L-374,087 (1), namely, its modest half-lives in animals, by making a chemically stable 3-alkylaminopyrazinone bioisostere for its 3-sulfonylaminopyridinone core. Compound 3 (L-375,378), the closest aminopyrazinone analogue of 1, has comparable selectivity and slightly decreased efficacy but significantly improved pharmacokinetics in rats, dogs, and monkeys to 1. We have developed an efficient and versatile synthesis of 3, and this compound has been chosen for further preclinical and clinical development.

Aminopyridines↗
PubMed1998

L-374,087, an efficacious, orally bioavailable, pyridinone acetamide thrombin inhibitor.

Replacement of the amidinopiperidine P1 group of 3-benzylsulfonylamino-6-methyl-2-pyridinone acetamide thrombin inhibitor L-373,890 (2) with a mildly basic 5-linked 2-amino-6-methylpyridine results in an equipotent compound L-374,087 (5, Ki = 0.5 nM). Compound 5 is highly selective for thrombin over trypsin, is efficacious in the rat ferric chloride model of arterial thrombosis and is orally bioavailable in dogs and cynomolgus monkeys. The structural basis for the critical importance of both methyl groups in 5 was confirmed by X-ray crystallography.

Administration, Oral↗
PubMed1998

Testing a new alcohol-free hand sanitizer to combat infection.

Universal precautions require that perioperative health care personnel wash their hand before and after all patient contact. Time constraints, however, can make adhering to universal precautions, including proper hand washing, difficult. Some perioperative health care workers, therefore, routinely use rise-free hand sanitizers to supplement normal hand washing. This study evaluated immediate and persistent antimicrobial effectiveness of two alcohol--containing hand sanitizers and a novel surfactant, allantoin, benzalkonium chloride (SAB) hand sanitizer using a federally approved effectiveness protocol. Results indicate that all three products were equally effective after a single application. After repeated use, the alcohol-containing sanitizers did not meet federal performance standards, and the alcohol-free sanitizer did. These properties and others illustrated in this article indicate that the nonflammable, alcohol-free SAB hand sanitizer is the most favorable of the rise-free hand sanitizer formulas for normal hand washing.

Adolescent↗
PubMed1997

Human intestinal folate transport: cloning, expression, and distribution of complementary RNA.

BACKGROUND & AIMS: Despite intensive investigations, very little is known about the molecular identity(ies) of the intestinal folate transport system(s), especially in humans. The aim of this study was to isolate a functional human intestinal folate carrier complementary DNA (cDNA) clone and determine the distribution of complementary RNA at the tissue and cellular levels. METHODS: Hybridization screening, modified Marathon cDNA amplification, expression in Xenopus oocytes, Northern analysis, and in situ hybridization were used. RESULTS: The hIFC-1 cDNA contains an open reading frame for 591 amino acids (relative molecular mass = 64,826, pI = 9.4, 12 transmembrane domains, three protein kinase C phosphorylation sites, and one N-glycosylation site) with 74% DNA and 66% amino acid sequence homologies with the mouse cDNA counterpart. Xenopus oocytes injected with hIFC-1 cRNA show induced folate uptake that was (1) saturable with substrate concentration (apparent Michaelis constant = 0.71 +/- 0.06 micromol/L; maximum velocity = 128 +/- 3 fmol x h(-1) x oocyte(-1)), (2) inhibited by methotrexate, folinic acid, and folic acid (Ki = 0.84 micromol/L, 0.71 micromol/L, and 10 micromol/L, respectively), and (3) sensitive to 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (Ki = 0.29 mmol/L). Northern analysis showed wide distribution of hIFC1-complementary messenger RNA species in various human tissues. In situ hybridization on sections of human jejunum showed preferential hIFC-1 expression in epithelial cells, especially in the upper half of the villi. CONCLUSIONS: These results represent the first molecular characterization of a human small intestinal folate carrier.

Amino Acid Sequence↗
PubMed1996

Intestinal folate transport: identification of a cDNA involved in folate transport and the functional expression and distribution of its mRNA.

Although the mechanism of folate intestinal transport has been the subject of intensive studies, very little is known about the molecular identity of the transport system(s) involved. In this investigation, we screened a mouse intestinal cDNA library using as probe the cDNA clone of a reduced folate carrier (RFC1) of mouse leukemia L1210 cells, and identified a positive clone, IFC1(RFC1). The cloned cDNA consisted of 2274 base pairs with an open reading frame that encodes a putative polypeptide of 512 amino acids with a predicted molecular mass of 58,112 daltons and 12 putative transmembrane domains. The polypeptide appears to carry a net positive charge (pI = 8.6) which may be important for its interaction with the negatively charged substrate. Functional identity of the IFC1(RFC1) clone was established by expression in Xenopus oocytes. An 11-fold increase in 5-methyltetrahydrofolate (5-MTHF) uptake was observed in oocytes injected with 10 ng IFC1(RFC1) cRNA compared to water-injected controls. The expressed folate uptake in the cRNA injected oocyte was (1) 4,4'-diisothiocyanatosilbene-2,2'-disulfonic acid (DIDS)-sensitive; and (2) saturable with an apparent Km of 1.99 +/- 0.32 micrometers and a V(max) of 3782 +/- 188 fmol/oocyte per h. The distribution of mRNA species complementary to IFC1(RFC1) in different mouse tissues was examined by Northern blot analysis. In addition to the small intestine, expression of such mRNA species were also found in the kidney, large intestine, brain, heart and liver. Furthermore, mRNA species complementary to IFC1(RFC1) were also detected by Northern blot analysis in the small intestine of human and other animal species (rat and rabbit). Expression of mRNA complementary to IFC1(RFC1) was markedly higher in rat intestinal villus cells than in crypt cells. These results represent the first identification of a folate transporter in mammalian intestine.

Animals↗
PubMed1996

Lysophosphatidic acid-induced neurite retraction in PC12 cells: control by phosphoinositide-Ca2+ signaling and Rho.

The endogenous phospholipid mediator lysophosphatidic acid (LPA) caused growth cone collapse, neurite retraction, and cell flattening in differentiated PC12 cells. Neurite retraction was blocked by cytochalasin B and ADP-ribosylation of the small-molecular-weight G protein Rho by the Clostridium botulinum C-3 toxin. LPA induced a transient rise in the level of inositol 1,4,5-trisphosphate, and retraction was blocked by inhibitors of phospholipase beta. Repeated application of LPA elicited homologous desensitization of the Ca2+ mobilization response. The activation of the phosphoinositide (PIP)-Ca2+ second messenger system played a permissive role in the morphoregulatory response. Blockers of protein kinase C--chelerythrine, a myristoylated pseudosubstrate peptide, staurosporine, and depletion of protein kinase C from the cells by long-term phorbol ester treatment--all diminished neurite retraction by interfering with LPA-induced Ca2+ mobilization, which was required for the withdrawal of neurites. A brief 15-min treatment with 4 beta-phorbol 12-myristate 13-acetate also blocked retraction and Ca2+ mobilization, by inactivating the LPA receptor. Inhibition of protein tyrosine phosphorylation by herbimycin diminished retraction. Although activation of the PIP-Ca2+ second messenger system appears necessary for the Rho-mediated rearrangements of the actin cytoskeleton, bradykinin, which activates similar signaling events, failed to cause retraction, indicating that a yet unidentified novel mechanism is also involved in the LPA-induced morphoregulatory response.

Actins↗
PubMed1996

Lysophosphatidic acid-induced neurite retraction in PC12 cells: neurite-protective effects of cyclic AMP signaling.

Effects of the cyclic AMP second messenger system were studied on the retraction of neurites elicited by the phospholipid mediator lysophosphatidic acid (LPA) in PC12 cells. LPA stimulation inhibited adenylyl cyclase, indicating that the LPA receptor couples to the heterotrimeric Gi proteins. However, pertussis toxin or expression of dominant negative Ras did not prevent neurite retraction. In contrast, cholera toxin, forskolin, and application of dibutyryl-cyclic AMP prevented neurite retraction. The neurite-protective effect of forskolin was blocked by Rp-adenosine 3',5'-phosphorothioate. Forskolin and dibutyryl-cyclic AMP both failed to protect neurites in A126-1B2 and 123.7 cells, which lack cyclic AMP-activated protein kinase. Data indicate that elevation of cyclic AMP levels triggers a cyclic AMP-activated protein kinase-dependent mechanism that opposes the functioning of the morphoregulatory signaling activated by LPA. ADP-ribosylation of Rho by the Clostridium botulinum C-3 toxin in 123.7 cells caused neuronal differentiation, indicated by neurite extension, and blocked LPA-induced neurite retraction. LPA activates Gq- and Gi-linked signaling in parallel; therefore, a morphoregulatory signaling network hypothesis is proposed versus the simplistic approach of a signaling pathway. The signaling network integrates the receptor-activated individual, sequential, and parallel signaling events into an interactive network whose individual components may fulfill required and permissive functions encoding the cellular response.

ADP Ribose Transferases↗
PubMed1995

Riboflavin uptake by native Xenopus laevis oocytes.

The existence of a membrane-associated uptake carrier for riboflavin (RF) is demonstrated in Xenopus oocytes. Uptake of low (0.017 microM) and high (3 microM) concentrations of RF was linear with time for up to 2 hours, and occurred with little initial binding to oocytes, and little metabolism. Uptake of RF was found to be independent of extracellular pH and Na+. The initial rate of RF uptake was saturable as a function of concentration with an apparent Km of 0.41 +/- 0.02 microM and a Vmax of 2.86 +/- 0.04 fmol/oocyte per h. Uptake of 3H-RF was inhibited by unlabeled RF and by the structural analogs lumiflavin, isoriboflavin (iso-RF), 8-aminoriboflavin (8-NH2-RF), 8-hydroxyriboflavin (8-OH-RF), and lumichrome, but was not affected by flavin adenine dinucleotide (FAD), D-ribose or lumazine. Uptake of RF was significantly retarded by the metabolic inhibitor 2,4-dinitrophenol. The sulfhydryl group-modifying reagents p-chloromercuriphenylsulfonate (pCMPS), p-chloromercuribenzoate (pCMB), N-ethylmaleimide and 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl) all caused significant inhibition in RF uptake. The inhibitory effect of pCMPS was completely reversed by treatment of pCMPS-pretreated cells with reducing agents. While the transmembrane transport inhibitors 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS), 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) and furosemide had no effect on RF uptake, amiloride and probenecid suppressed RF uptake in a dose-dependent fashion. Closer examination of the inhibition mediated by amiloride showed that it was competitive in nature with an apparent Ki of approximately 1.8 mM, whereas the inhibition induced by probenecid was nonspecific. Together, these findings indicate that Xenopus oocytes possess an endogenous, specific, membrane-associated carrier-mediated uptake system for RF. The results also demonstrate the usefulness of Xenopus oocytes as a model system with which to study the RF transport event across biological membranes, which should further out present understanding of RF uptake by various vertebrate cells.

Amiloride↗
PubMed1994

Lysophosphatidic acid possesses dual action in cell proliferation.

Lysophosphatidic acid (LPA) induces mitogenic responses in cultured fibroblasts through a pertussis toxin-sensitive signaling pathway. In contrast, we have shown that LPA inhibits the proliferation of Sp2/0-Ag14 myeloma cells. To resolve this apparent controversy, LPA-elicited responses in cell proliferation and the underlying second messenger mechanisms were compared in Sp2/0-Ag14 myeloma and NIH 3T3 fibroblast cells. The antimitogenic response was not elicited by micromolar concentrations of phosphatidic acid, phosphatidylglycerol, or diacylglycerol. In NIH 3T3 and Sp2 cells, LPA elicited an increase in inositol trisphosphate and a subsequent transient increase in free cytoplasmic Ca2+. Unlike the mitogenic response in NIH 3T3 cells, the antimitogenic effect was not affected by pertussis toxin; on the contrary, it was accompanied by an increase in cAMP. In Sp2 cells, cAMP analogs, forskolin, and isobutylmethylxanthine inhibited cell proliferation and enhanced LPA action in an additive manner, suggesting that an LPA-elicited increase in cAMP-mediated signaling was responsible for the antimitogenic response. In addition to the mitogenic response in fibroblasts and the antimitogenic response in tumor cell lines, there are some cell types (Jurkat T-cell lymphoma and primary astrocytes) in which LPA is ineffective in altering cell proliferation. The cell-type-specific dual action of LPA suggests that this endogenous lipid mediator when released from activated cells might play an important role as a regulator, rather than a ubiquitous inducer, of cell proliferation.

3T3 Cells↗
PubMed1994

Human intestinal cell line Caco-2: a useful model for studying cellular and molecular regulation of biotin uptake.

The mechanisms of enterocyte and molecular regulation of biotin uptake are poorly understood. An intestinal cell line processing the transport characteristics of native intestinal cells is highly desirable to investigate the finer details of the cellular processing and molecular regulation of biotin transport. In the present study, we investigated the uptake of the water-soluble vitamin biotin by a human intestinal cell line Caco-2. Uptake of both low (4 nM) and high (20 microM) concentrations of biotin by confluent monolayers of Caco-2 cells was appreciable and linear for up to 10 min of incubation. Replacement of Na+ in the incubation medium with other monovalent cations--K+, choline, Li+ and NH4(+)--caused a significant inhibition of biotin uptake; a relatively lesser inhibition was seen with Li+. Initial rate of uptake of biotin was temperature-dependent and saturable as a function of concentration at 37 degrees C but not at 4 degrees C. The Vmax and apparent Km of the temperature-dependent saturable process were 520 pmol/mg protein per min and 9.5 microM, respectively. The addition of unlabeled biotin and the structural analogue desthiobiotin to the incubation media caused a significant inhibition of the uptake of [3H]biotin. The inhibitory effect of desthiobiotin was competitive in nature with an inhibition constant (Ki) of 41 microM. Biocytin, on the other hand, was a weak inhibitor and biotin methyl ester and diaminobiotin did not have any effect. Pretreatment of Caco-2 cells with the monovalent cation ionophore gramicidin and the Na+, K+(-)ATPase inhibitor ouabain caused significant inhibition of biotin uptake. Pretreatment with the K+ ionophore valinomycin did not affect biotin uptake. Using the 'Activation Method', the stoichiometric ratio of biotin- to Na+ coupling was found to be 1:1. Growing confluent Caco-2 cells in a biotin-deficient environment resulted in rapid up-regulation of biotin transport with a marked increase (258%) in the Vmax of biotin uptake. These findings demonstrate that biotin uptake by Caco-2 cells is via a carrier-mediated system. This system is temperature-dependent, driven by Na(+)-gradient and is regulated by the substrate level. These in-vitro findings are very similar to and further confirm previous findings in human and animal studies and dispute other findings previously reported for Caco-2 cells; the present study also demonstrates the suitability of this system for further characterization of the cellular and molecular regulation of biotin uptake.

Biological Transport↗
PubMed1994

Expression of a rabbit renal ascorbic acid transporter in Xenopus laevis oocytes.

We examined the expression of renal ascorbic acid transporter(s) in Xenopus laevis oocytes after microinjection of cells with poly(A)+ RNA extracted from rabbit kidney cortex. Concomitant expression of the Na+-glucose cotransporter served as a control in these studies. Injection of poly(A)+ RNA into oocytes produced over a fivefold increase in the uptake of [14C]ascorbic acid (570 microM) compared with water-injected cells. Size fractionation of the kidney cortex mRNA by sucrose gradient revealed that the mRNA species that induced ascorbic acid transporter expression in oocytes was present in a fraction centered around 2.0 kilobases (kb) and had a size range of 1.8-3.1 kb. Injection of the active fraction into oocytes produced a > 40-fold increase in ascorbic acid uptake compared with water-injected controls. Expression of ascorbic acid transporter(s) was noticeable as early as 2 days after injection and was maximal after 7 days; it was also dependent on the amount of mRNA injected into oocytes. The induced uptake of [14C]ascorbic acid after injection of mRNA into oocytes was 1) Na+ dependent, as indicated by the almost complete lack of transport on removal of Na+ from the incubation medium; 2) significantly inhibited by unlabeled ascorbic acid and its structural analogue isoascorbic acid but not by D-glucose; and 3) saturable as a function of increasing the substrate concentration in the incubation medium (100-1,000 microM), with an apparent Km of 258 +/- 72.5 microM and a maximum velocity of 29.6 +/- 2.8 pmol.oocyte-1.2 h-1. These data demonstrate that X. laevis oocytes are a suitable system to functionally express the mammalian renal ascorbic acid transporter.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1969

Fluorescent europium chelate sta.

The europium chelate of 4,4,4-trifluoro-1-(2-thienyl)-1,3-butanedione (thenoyl-trifluoroacetone; TTA) is firmly bound to microorganisms. It fluoresces brightly at 613 nm with activation at 340 nm. Cells may be stained with 10(-3)m chelate in 50% ethyl alcohol, followed by washing with 50% ethyl alcohol. Equal or better stains are produced with 10(-3)m aqueous europium salt, water wash, and 10(-2)m aqueous TTA. A noncomplexing buffer should be used to maintain the pH at 6.5 to 6.8.

Bacillus megaterium↗