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Biomedical subjects

L M Shaw

Publications and source records attributed to L M Shaw.

At least 91 records · Page 5Linked to original sources
PubMed1995

Regulation of alpha 6 beta 1 integrin-mediated migration in macrophages.

Several integrin alpha subunits have structural variants that are identical in their extracellular and transmembrane domains but that differ in their cytoplasmic domains. In the present study, we examined the possibility that the A and B variants of the alpha 6 beta 1 integrin laminin receptor differ in function. P388D1 macrophages that had been transfected with the alpha A integrin subunit were 3-4 fold more migratory than P388D1 macrophages that had been transfected with the alpha 6 B integrin subunit. Deletion of the alpha 6 cytoplasmic domain markedly inhibited the ability of the alpha 6 beta 1 receptor to promote migration.

Calcium↗
PubMed1994

Cannabinoids and appetite stimulation.

Appetite stimulation by cannabinoids is highly variable. Four within-subject design studies explored the effects of age, gender, satiety status, route of drug administration, and dose on intake. One study involved a single oral administration of active drug (15 mg males, 10 mg females) or placebo to an age and gender stratified sample of 57 healthy, adult light marijuana users. Eleven subjects received single doses by oral, sublingual, and inhaled routes in a second study. In the third study, 10 subjects ingested a single oral dose in fasted and fed states. A 2.5 mg dose was administered b.i.d. for 3 days by oral and rectal suppository routes in the fourth study. Mean daily energy intake was significantly elevated following chronic dosing by rectal suppository, but not oral capsule, relative to all acute dosing regimens except inhalation. Total daily energy intake was comparable on fed and fasted days, suggesting satiety mechanisms were not impaired by the drug. Subject age, gender, reported "high," and plasma drug level were not significantly associated with drug effects on food intake.

Adult↗
PubMed1994

Regulation of cellular interactions with laminin by integrin cytoplasmic domains: the A and B structural variants of the alpha 6 beta 1 integrin differentially modulate the adhesive strength, morphology, and migration of macrophages.

Several integrin alpha subunits have structural variants that are identical in their extracellular and transmembrane domains but that differ in their cytoplasmic domains. The functional significance of these variants, however, is unknown. In the present study, we examined the possibility that the A and B variants of the alpha 6 beta 1 integrin laminin receptor differ in function. For this purpose, we expressed the alpha 6A and alpha 6B cDNAs, as well as a truncated alpha 6 cDNA (alpha 6-delta CYT) in which the cytoplasmic domain sequence was deleted after the GFFKR pentapeptide, in P388D1 cells, an alpha 6 deficient macrophage cell line. Populations of stable alpha 6A, alpha 6B, and alpha 6-delta CYT transfectants that expressed equivalent levels of cell surface alpha 6 were obtained by fluorescence-activated cell sorter and shown to form heterodimers with endogenous beta 1 subunits. Upon attachment to laminin, the alpha 6A transfectants extended numerous pseudopodia. In contrast, the alpha 6B transfectants remained rounded and extended few processes. The transfectants were also examined for their ability to migrate toward a laminin substratum using Transwell chambers. The alpha 6A transfectants were three- to fourfold more migratory than the alpha 6B transfectants. The alpha 6-delta CYT transfectants did not attach to laminin in normal culture medium, but they did attach in the presence of Mn2+. The alpha 6-delta CYT transfectants migrated to a lesser extent than either the alpha 6A or alpha 6B transfectants in the presence of Mn2+. The alpha 6 transfectants differed significantly in the concentration of substratum bound laminin required for half-maximal adhesion in the presence of Mn2+:alpha 6A (2.1 micrograms/ml), alpha 6B (6.3 micrograms/ml), and alpha 6-delta CYT (8.8 micrograms/ml). Divalent cation titration studies revealed that these transfectants also differed significantly in both the [Ca2+] and [Mn2+] required to obtain half-maximal adhesion to laminin. These data demonstrate that the A and B variants of the alpha 6 cytoplasmic domain can differentially modulate the function of the alpha 6 beta 1 extracellular domain.

Animals↗
PubMed1994

Effects of cannabinoids (marijuana) on taste intensity and hedonic ratings and salivary flow of adults.

Cannabinoids purportedly improve taste responsiveness and enhance the sensory appeal of foods. These properties and a commonly cited oral drying effect were evaluated in a series of studies with 'light' marijuana users. The first was a double-blind, placebo-controlled, acute oral dosing trial, involving an age and gender stratified sample of 57 adults. An influence of route of drug delivery was explored in another 11 individuals who were administered a single dose orally, sublingually and via cigarette. To explore effects following chronic administration, six additional individuals were dosed twice per day for 3 days orally and by rectal suppository. Taste intensity and hedonic responses for sweet, sour, salty and bitter food stimuli were monitored at baseline, 2, 4 and 6 hours post-dosing in the acute studies, and daily in the chronic study. Stimulated saliva samples were collected at these same times. Salivary flow rate was significantly negatively correlated with plasma drug levels, and reported 'high' 2 and 4 h post-dosing. No effects of the drug were observed on taste responses. Self-reported shifts in taste responsiveness and hedonics may be related to alterations of memory and cognition, rather than gustatory function.

Administration, Inhalation↗
PubMed1994

Utility of pretransplantation cyclosporine pharmacokinetic studies.

Pretransplant cyclosporine (CsA) pharmacokinetic analysis of an individual patient is advocated as a more accurate method of determining the optimal dose schedule of CsA for immediate posttransplant patients than traditional mg/kg dosing methods. Eight adult renal transplant candidates (age range 28-69 years) were studied. CsA whole blood analysis was done with a monoclonal fluorescence polarization immunoassay (mFPIA) and high-performance liquid chromatography HPLC. Noncompartmental modeling methods were used to derive CsA pharmacokinetic values. At 1 and 3 months posttransplant CsA pharmacokinetic analyses were completed on five subjects and the average steady state CsA concentration for the dosing interval, Cav, was compared to predicted values calculated from pretransplant pharmacokinetic parameters for each subject. At 6 months posttransplant, actual and predicted Cav were compared in three subjects. Correlation between predicted and actual Cav at 1 month posttransplant was poor (mean actual Cav = 365 ng/ml versus mean predicted Cav = 238 ng/ml; r2 = 0.361). At 3 months posttransplant, the discrepancy between predicted and actual Cav was greater for all five subjects (r2 = 0.039) and this trend persisted for three subjects at 6 months posttransplantation. The mFPIA analysis overestimated the parent CsA concentration when compared to HPLC analysis; the degree of overestimation ranged from 118 to 180%. The mFPIA assay variability may have contributed to the poor correlation between pre- and posttransplant Cav values. There appears to be little or no basis for subjecting transplant candidates to sophisticated pharmacokinetic tests in order to develop specific CsA dosing guidelines for the posttransplant phase.

Adult↗
PubMed1994

Diabetic nephropathy and microalbuminuria in the community. The South Auckland Diabetes Survey.

OBJECTIVE: To compare the clinical, anthropometric, and metabolic characteristics of New Zealand Europeans, Maori, and Pacific Islanders with non-insulin-dependent diabetes mellitus (NIDDM) with emphasis on risk factors for the development of diabetic nephropathy. RESEARCH DESIGN AND METHODS: A cross-sectional survey of 555 (74% of 750 available) diabetic patients attending diabetes clinics and randomly selected primary care centers was conducted in Auckland, New Zealand. RESULTS: Among those with NIDDM, Maori and Pacific Islanders were younger at diagnosis, more obese, and had poorer glucose control when compared with the Europeans (fructosamine in mumol/l: Maori 335 +/- 78, Pacific Islanders 367 +/- 90, Europeans 318 +/- 55; overall P < 0.001). Systolic blood pressure (sBP) was higher in Maori (145 +/- 31 mmHg) and lower in Pacific Islanders (135 +/- 25 mmHg) when compared with Europeans (141 +/- 25 mmHg; overall P < 0.005). Mean estimated daily urinary albumin excretion (UAE) was 18.2 (15.5-1.3) mg/day in Europeans, 94.8 (60.5-148.7) mg/day in Maori, and 44.2 (32.3-60.3) mg/day in Pacific Islanders. The prevalence of proteinuria and end-stage renal failure were also higher in Maori and Pacific Islanders. The excess prevalence of microalbuminuria and proteinuria in Maori was present within 5 years of diagnosis. Europeans with impaired renal function were least likely to have associated proteinuria or microalbuminuria. Microalbuminuria and nephropathy were not consistently associated with either higher blood pressure or worse glucose control. CONCLUSIONS: NIDDM in Maori and Pacific Islanders is associated with a greater degree of proteinuria and end-stage renal failure than that in Europeans. This observation is not explained by conventional risk factors.

Adolescent↗
PubMed1993

Inside-out integrin signaling in macrophages. Analysis of the role of the alpha 6A beta 1 and alpha 6B beta 1 integrin variants in laminin adhesion by cDNA expression in an alpha 6 integrin-deficient macrophage cell line.

Leukocytes use the alpha 6 beta 1 integrin to adhere to laminin based on mAb inhibition and affinity chromatography studies. This adhesion requires leukocyte stimulation with either PMA or specific cytokines, a process that has been termed "inside-out" integrin signaling. In the present study, the involvement of alpha 6 integrin structural variants in this regulated adhesion was examined using mouse macrophages. The two known alpha 6 structural variants, alpha 6A and alpha 6B, differ only in their cytoplasmic domain sequences. Using reverse transcriptase-polymerase chain reaction, we observed that macrophages express only the alpha 6A structural variant, in contrast to most cell types which express both alpha 6A and alpha 6B variants. The role of this integrin subunit in macrophage adhesion was assessed by cDNA transfection of P388D1 cells. We found that this mouse macrophage cell line does not adhere to laminin even in response to phorbol 12-myristate 13-acetate (PMA) stimulation, though it does adhere normally to fibronectin and tissue culture plastic. Subsequent analysis employing reverse transcriptase-polymerase chain reaction and immunoprecipitation of surface labeled cells revealed that this cell line expresses neither the alpha 6A nor alpha 6B integrin subunits. Stable transfection of either the chick or human alpha 6A cDNAs into P388D1 cells resulted in chimeric alpha 6A beta 1 surface expression. The alpha 6A transfectants exhibited inside-out integrin signaling because PMA stimulation markedly increased their ability to adhere to laminin but it did not increase alpha 6A beta 1 surface expression. Similar results were obtained after transfection of the human alpha 6B cDNA. Analysis of the human transfectants was facilitated by the generation of a monoclonal antibody, 2B7, that is specific for the human alpha 6 integrin subunit. These observations demonstrate that both alpha 6A beta 1 and alpha 6B beta 1 can be regulated by inside-out signaling pathways in macrophages, even though this cell type expresses only alpha 6A beta 1. The data presented also demonstrate clearly that the alpha 6A and alpha 6B cytoplasmic domains do not differ in their ability to be regulated by PMA.

Amino Acid Sequence↗
PubMed1993

Fimbrin localized to an insoluble cytoskeletal fraction is constitutively phosphorylated on its headpiece domain in adherent macrophages.

The actin-bundling protein fimbrin is homologous to 1-plastin, a 65kD phosphoprotein expressed in leukocytes and transformed cells [de Arruda et al., J. Cell Biol. 111, 1069-1080]. Because fimbrin is present in cell adhesion sites, we studied the phosphorylation state of fimbrin and its distribution in macrophages sequentially extracted with Triton-X-100 (soluble fraction), Tween 40-deoxy-cholate (cytoskeletal fraction), and SDS (insoluble cytoskeletal fraction). The approximate distribution of fimbrin and actin among these fractions was found to be: 65% fimbrin/55% actin in the soluble fraction, 30% fimbrin/20% actin in the cytoskeletal fraction, and 5% fimbrin/25% actin in the insoluble cytoskeletal fraction. PMA did not alter this distribution. Fluorescence microscopy of acetone-extracted macrophages showed that actin is concentrated in podosomes at the substratum interface and is diffusely distributed throughout the remainder of the cell. Fimbrin colocalizes with actin in podosomes and also exhibits a punctate distribution in the cytoplasm that overlaps with actin. In Tween 40/DOC-extracted cells, podosomes remain, and fimbrin also exhibits a punctate distribution along actin filaments. Metabolic 32PO4 labeling revealed that fimbrin is constitutively phosphorylated and that phosphorylated fimbrin is concentrated in the insoluble cytoskeletal fraction. PMA increased the relative levels of fimbrin phosphorylation twofold but did not alter the pattern of fimbrin fluorescence or the distribution of phosphorylated fimbrin. Limited trypsin digestion and phosphoamino acid analysis demonstrated that phosphorylation occurs specifically on serine residues within the 10kD headpiece domain of fimbrin. Phosphorylation of the headpiece domain could regulate the actin binding and bundling properties of fimbrin, or it could regulate the interaction of fimbrin with other proteins.

Actins↗
PubMed1993

Bypassing the first-pass effect for the therapeutic use of cannabinoids.

An oral formulation of delta-9-tetrahydrocannabinol (THC) in sesame oil (Marinol) is at present used for the management of chemotherapy-related nausea and emesis. However, due partly to poor bioavailability, its efficacy is variable. To circumvent possible metabolism in the gut and a first-pass effect by the liver, a suppository formulation of THC hemisuccinate ester was prepared. Administration of the suppository containing 11.8 mg of the hemisuccinate ester (equivalent to 9 mg THC) to three adult females (two of whom had previously exhibited low plasma drug levels following a 10-mg dose of the oral formulation) led to a marked and sustained elevation of plasma drug levels. Areas under the curves for plasma THC were more than 30-fold higher than after oral dosing. The suppository was well tolerated. The higher and more sustained plasma drug level achieved with this new formulation should enhance its antiemetic efficacy.

Adult↗
PubMed1993

Regulation of alpha 6 beta 1 integrin laminin receptor function by the cytoplasmic domain of the alpha 6 subunit.

The alpha 6 beta 1 integrin is expressed on the macrophage surface in an inactive state and requires cellular activation with PMA or cytokines to function as a laminin receptor (Shaw, L. M., J. M. Messier, and A. M. Mercurio. 1990. J. Cell Biol. 110:2167-2174). In the present study, the role of the alpha 6 subunit cytoplasmic domain in alpha 6 beta 1 integrin activation was examined. The use of P388D1 cells, an alpha 6-integrin deficient macrophage cell line, facilitated this analysis because expression of either the alpha 6A or alpha 6B subunit cDNAs restores their activation responsive laminin adhesion (Shaw, L. S., M. Lotz, and A. M. Mercurio. 1993. J. Biol. Chem. 268:11401-11408). A truncated alpha 6 cDNA, alpha 6-delta CYT, was constructed in which the human cytoplasmic domain sequence was deleted after the GFFKR pentapeptide. Expression of this cDNA in P388D1 cells resulted in the surface expression of a chimeric alpha 6-delta CYT beta 1 integrin that was unable to mediate laminin adhesion or increase this adhesion in response to PMA under normal conditions, i.e., in medium that contained physiological concentrations of Ca++ and Mg++. The alpha 6A-delta CYT transfectants adhered to laminin, however, when Ca++/Mg++ was replaced with 150 microM Mn++. We also assessed the role of serine phosphorylation in the regulation of alpha 6A beta 1 integrin function by site-directed mutagenesis of the two serine residues present in the alpha 6A cytoplasmic domain because this domain is phosphorylated on serine residues in response to stimuli that activate the laminin receptor function of alpha 6 A beta 1. Point mutations were introduced in the alpha 6A cDNA that changed either serine residue #1064 (M1) or serine residue #1071 (M2) to alanine residues. In addition, a double mutant (M3) was constructed in which both serine residues were changed to alanine residues. P388D1 transfectants which expressed these serine mutations adhered to laminin in response to PMA to the same extent as cells transfected with wild-type alpha 6A cDNA. These findings provide evidence for a novel mode of integrin regulation that is distinct from that reported for other regulated integrins (O'Toole, T. E., D. Mandelman, J. Forsyth, S. J. Shattil, E. F. Plow, and M. H. Ginsberg. 1991. Science (Wash. DC). 254:845-847. Hibbs, M. L., H. Xu, S. A. Stacker, and T. A. Springer. 1991. Science (Wash. DC). 251:1611-1613), and they demonstrate that serine phosphorylation of the alpha 6A cytoplasmic domain is not involved in this regulation.

Amino Acid Sequence↗
PubMed1993

The use of concentration measurements of parent drug and metabolites during clinical trials.

The need for well-designed pharmacokinetic (PK) and pharmacodynamic (PD) studies early in the development of new drugs is described. In this review we illustrate the application and cost-effectiveness of optimal sampling theory in PK study design for ongoing clinical trial studies of ethyol, a chemoprotector drug. The importance of careful selection of the appropriate biological fluid in which to measure drug concentration at the earliest possible stage of new drug development is described in the context of the development of new immunosuppressive drugs. We focus on the requirement for well-validated analytical methodology in PK-PD studies, described in a discussion of the analytical methodology in use in clinical trials of two immunosuppressive agents, cyclosporin G and RS-61443 (mycophenolate mofetil).

Clinical Trials as Topic↗
PubMed1993

Pharmacokinetics and effect of cocaine on cerebral blood flow in the newborn.

The present study investigated the effect of cocaine (COC) on cerebral circulation (CBF) and oxidative metabolism (CMRO2) in the newborn piglet and aimed to relate pharmacokinetics of cocaine to cerebrovascular effects. COC decreased CBF and CMRO2 from 75 to 64 and 4.27 to 3.91 ml/min/100 g, respectively, at 4 min with reduced flow to all brain regions (p < 0.05) which returned to baseline by 10 min. COC was rapidly metabolized with a t1/2 of 43 min and peak plasma concentration of 1,172 ng/ml. Norcocaine (NOR) appeared in plasma and CSF within 3 min of cocaine administration and remained elevated for the duration of the study along with COC in the CSF. These data show that the timing of the peak plasma COC level is associated with maximal decreased CBF. Further, the stable elevated level of COC and NOR in the CSF suggests that biotransformation does not occur in the brain. As a result, accumulation of these drugs may occur in the brain with successive COC use and affect the developing CNS in a deleterious manner.

Animals↗
PubMed1993

Effects of cyclosporine and its metabolites in the isolated perfused rat kidney.

The isolated perfused rat kidney (IPK) was used to study the acute effects of cyclosporin A (CsA) and its metabolites (M1, M17, M18, M21 and M-COOH). GFR, renal vascular resistance, and sodium, potassium and water reabsorption were measured before and after the addition of CsA/metabolites/vehicles. There was no difference in CsA effect (mild decrease in GFR and increase in renal vascular resistance with the inclusion of plasma (10 mL) or whole blood (20 mL) in the albumin perfusate (120 mL). Intralipid was used as the vehicle for CsA and the metabolites because methanol, ethanol, and Cremophor had significant effects on GFR. Intralipid enhanced the effect of CsA 25-fold, giving CsA dose responses comparable to those of human kidneys. This enhanced effect with intralipid was due to vasoconstriction, not vascular obstruction, and was apparently specific to CsA (no enhancement of norepinephrine with Intralipid). The primary metabolites (M1, M17, and M21) caused decreases in GFR comparable to or slightly less than those caused by CsA. The secondary metabolites (M18 and M-COOH) caused more modest declines in GFR. Cyclosporine metabolite levels in patient blood often greatly exceed levels of the parent drug; these studies suggest that the metabolites may contribute significantly to CsA nephrotoxicity in patients.

Animals↗
PubMed1993

The underreporting of cocaine-related trauma: drug abuse warning network reports vs hospital toxicology tests.

OBJECTIVE: The purpose of this study was to assess whether cocaine-related trauma is underreported to the US Federal Drug Abuse Warning Network (DAWN). METHODS: We compared DAWN reports filed by an urban emergency department with the department's toxicology results for patients treated for major trauma. DAWN regulations in effect during the study period required the reporting of all patients treated for injury who used drugs or who tested positive for drugs of abuse. RESULTS: Of 520 patients treated for major trauma, 217 (42%) were tested for a variety of drugs. Of these, 82 (38%) tested positive for cocaine. Of the 102 patients injured in motor vehicle accidents, 20 (20%) tested positive for cocaine. Of the 59 patients injured in motor vehicle accidents who were under age 40, 18 (30%) tested positive for cocaine. Of 100 victims of violent assault, 57 tested positive for cocaine. During the time period studied, DAWN recorded 48 hospital visits associated with cocaine, none involving trauma or injury. CONCLUSIONS: Cocaine-related trauma was unreported to DAWN despite the hospital's compliance with the system's guidelines. The pattern of DAWN reports from other institutions suggests that underreporting of cocaine-related injury is widespread.

Accidents↗
PubMed1993

Unregulated self-medication in an ambulatory surgery population.

The objective of this study was to determine whether unregulated self-medication in an ambulatory surgery population was a significant problem in terms of the incidence and nature of drugs detected. We asked 67 consecutive patients who presented for elective ambulatory surgery at a university medical center to volunteer for serum and urine testing with the assurance of confidentiality. All patients denied nonprescription drug use. Initial testing was for amphetamines, barbiturates, benzodiazepines, cocaine, opiates, phencyclidine (PCP), tetrahydrocannabinol (THC) and alcohol. Of the 64 patients who agreed to be tested, 16 patients tested positive for at least one of the above substances; 4 patients demonstrated residua of 2 or 3 substances. No distinctive characteristic of those with evidence of drug use could be identified. We believe that the 25% of patients who tested positive in this group is a conservative estimate of possible unrecognized drug use. We have demonstrated a significant incidence of substance abuse in the ambulatory surgery environment. These findings have serious safety implications, and a higher awareness of possible nonprescription drug use in this environment is needed in medical and support personnel.

Adult↗