Assuntos
Pressão Sanguínea/efeitos dos fármacos , Meios de Contraste/efeitos adversos , Inibidores Enzimáticos/farmacologia , Hipotensão/induzido quimicamente , NG-Nitroarginina Metil Éster/farmacologia , Animais , Meios de Contraste/administração & dosagem , Histamina/biossíntese , Injeções Intravenosas , Iopamidol/administração & dosagem , Iopamidol/efeitos adversos , Iotalamato de Meglumina/administração & dosagem , Iotalamato de Meglumina/efeitos adversos , Óxido Nítrico/biossíntese , Ratos , Ratos Sprague-Dawley , Ácidos Tri-Iodobenzoicos/administração & dosagem , Ácidos Tri-Iodobenzoicos/efeitos adversosAssuntos
Anafilaxia/induzido quimicamente , Pressão Sanguínea/efeitos dos fármacos , Meios de Contraste/toxicidade , Hipersensibilidade a Drogas/fisiopatologia , Óxido Nítrico/fisiologia , Anafilaxia/fisiopatologia , Anafilaxia/prevenção & controle , Animais , Hipersensibilidade a Drogas/prevenção & controle , Homeostase/efeitos dos fármacos , Homeostase/fisiologia , Iotalamato de Meglumina/farmacologia , Dose Letal Mediana , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Ratos , Ratos Endogâmicos BN , Ratos Sprague-DawleyRESUMO
RATIONALE AND OBJECTIVES: We assessed the role that nitric oxide (NO) plays in contrast media (CM) toxicity, using 100% lethal dose (LD100) studies in hyperimmune Brown Norway (BN) rats. METHODS: Ninety-two BN rats and 41 Sprague-Dawley (SD) rats underwent CM LD100 tail vein injections with methylglucamine iothalamate or sodium iothalamate to the point of cessation of respiration. Methylglucamine hydrochloride also was injected. The injections were accompanied by L-arginine (L-Arg) or D-arginine (D-Arg) analogues or by an H1 blocker. L-Arg analogues inhibit NO formation, and D-Arg analogues do not. RESULTS: An L-Arg analogue, but not a D-Arg analogue, increased the tolerance of BN rats (p < .005) for methylglucamine iothalamate but not for sodium iothalamate. The L-Arg analogue also protected BN rats against methylglucamine chloride injections (p < .002). H1 blockade protected BN rats against methylglucamine iothalamate (p < .0005) and methylglucamine chloride (p < .005) injections. None of these measures altered the CM tolerance of SD rats. In SD rats, injections of either methylglucamine iothalamate or sodium iothalamate along with a D-Arg analogue or normal saline were better tolerated than similar injections in BN rats (p < .01 and .002 for methylglucamine iothalamate and sodium iothalamate, respectively). In SD rats but not BN rats, sodium iothalamate was better tolerated than was methylglucamine iothalamate (p < .0005). CONCLUSION: NO appears to play a significant role in BN rats LD100 CM toxicity and has been implicated by others in the blood pressure fall characterizing some forms of antigen-induced anaphylaxis [1, 2]. The results of the current study and the literature suggest that methylglucamine-modulated release of histamine from mast cells may underlie the NO production.
Assuntos
Anafilaxia/fisiopatologia , Meios de Contraste/toxicidade , Pulmão/fisiopatologia , Óxido Nítrico/fisiologia , Anafilaxia/induzido quimicamente , Anafilaxia/patologia , Animais , Meios de Contraste/administração & dosagem , Interações Medicamentosas , Inibidores Enzimáticos/farmacologia , Antagonistas dos Receptores Histamínicos H1/farmacologia , Injeções Intravenosas , Iotalamato de Meglumina/administração & dosagem , Iotalamato de Meglumina/toxicidade , Ácido Iotalâmico/administração & dosagem , Ácido Iotalâmico/toxicidade , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Ratos , Ratos Endogâmicos BN , Ratos Sprague-Dawley , ômega-N-Metilarginina/farmacologiaAssuntos
Iotalamato de Meglumina/toxicidade , Óxido Nítrico/fisiologia , Animais , Arginina/administração & dosagem , Arginina/análogos & derivados , Imunização , Imunoglobulina E/análise , NG-Nitroarginina Metil Éster , Óxido Nítrico/antagonistas & inibidores , Nitroarginina , Ovalbumina/imunologia , Ratos , Ratos Endogâmicos BN , Ratos Sprague-Dawley , Ratos Wistar , ômega-N-MetilargininaRESUMO
Interleukin 1 (IL-1), bacterial lipopolysaccharide (LPS) and tumor necrosis factor (TNF alpha) enhance the adherence properties of endothelial cells (EC) for neutrophils (PMN). This is mediated in part by the up-regulation of Intercellular Adhesion Molecule 1 (ICAM-1) on EC. Phorbol esters, which activate protein kinase c (PKC) and enhance the adherence properties of EC for PMN also up-regulate the ICAM-1 expression on EC. We investigated the effect of PKC inhibitors on ICAM-1 expression of human umbilical vein EC (HUVEC). Staurosporine (STS) and 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7) prevented inflammatory mediator-dependent stimulation of both ICAM-1 expression and PMN adherence by HUVEC (ID50 for STS = 2.7-2.9 microM; for H-7 = 7.6-8.8 microM). Inhibition was dose and time-dependent and was not due to HUVEC toxicity. The STS analog K252a and the H-7 analog W-7 were less potent inhibitors of ICAM-1 up-regulation and adherence promotion. Prolonged exposure of HUVEC to phorbol myristate acetate down-regulated PKC activity and inhibited subsequent ICAM-1 up-regulation by this agent and by IL-1. We conclude that inflammatory mediator induced stimulation of HUVEC expression of ICAM-1 and promotion of adherence properties are mediated in part by activation of PKC.
Assuntos
Moléculas de Adesão Celular/biossíntese , Endotélio Vascular/metabolismo , Interleucina-1/farmacologia , Lipopolissacarídeos/farmacologia , Proteína Quinase C/antagonistas & inibidores , Fator de Necrose Tumoral alfa/farmacologia , Adesão Celular , Células Cultivadas , Relação Dose-Resposta a Droga , Regulação para Baixo , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/enzimologia , Humanos , Cinética , Neutrófilos/fisiologia , Veias Umbilicais/efeitos dos fármacos , Veias Umbilicais/enzimologia , Veias Umbilicais/metabolismoRESUMO
Inflammatory stimuli enhance the adherence properties of human umbilical vein endothelial cells (HUVEC) for neutrophils (PMN). This is mediated in part by the up-regulation on HUVEC of Intercellular Adhesion Molecule 1 (ICAM 1). Phorbol esters, which activate protein kinase c (PKC), have also been reported to enhance the adherence properties of HUVEC for PMN. We investigated the effect of agents which activate PKC on the expression of ICAM 1 by HUVEC. Both phorbol myristate acetate (PMA) and Mezerein, a non-phorbol which also stimulates PKC, enhanced both the expression of ICAM 1 on HUVEC and the adherence of HUVEC for PMN. The PKC inhibitors staurosporine and H-7 prevented both PMA and Mezerein-induced stimulation of HUVEC expression of ICAM 1 and adherence for PMN. We conclude that activation of PKC in HUVEC is associated with increased expression of ICAM 1 on HUVEC. PKC-mediated up-regulation of ICAM 1 may be responsible, in part, for the promotion of endothelial cell adherence properties toward PMN.
Assuntos
Antígenos de Superfície/análise , Diterpenos , Endotélio Vascular/metabolismo , Glicoproteínas de Membrana/biossíntese , Proteína Quinase C/metabolismo , 1-(5-Isoquinolinasulfonil)-2-Metilpiperazina , Alcaloides/farmacologia , Carcinógenos/farmacologia , Adesão Celular , Moléculas de Adesão Celular , Endotélio Vascular/efeitos dos fármacos , Ativação Enzimática , Humanos , Isoquinolinas/farmacologia , Cinética , Ésteres de Forbol/farmacologia , Piperazinas/farmacologia , Estaurosporina , Terpenos/farmacologia , Acetato de Tetradecanoilforbol/farmacologiaRESUMO
The authors reported previously that stored granulocytes (PMN) had decreased receptor affinity (Kd) for and chemotaxis (CTX) to the chemotactic peptide F-Met-Leu-Phe (FMLP), but the evidence did not favor a significant role for altered FMLP receptor affinity in causing diminished CTX of stored PMN. Since recruitment and/or recycling of FMLP receptors is required for normal CTX, the hypothesis that stored PMN might have abnormal FMLP receptor cycling was tested. The effect of storage on the proportion of high- and low-affinity FMLP receptors was also investigated. Units of PMN were tested within 4 hours of collection and after 24 and 48 hours of storage at 22 degrees C, unagitated, in 150-ml transfer packs. In comparison to fresh PMN, there was no alteration in the Kd of the high-affinity FMLP receptor of PMN stored for 24 to 48 hours; however, the Kd of the low-affinity receptor increased (fresh PMN = 36 +/- 5 nM; 24 hours = 107 +/- 19; 48 hours = 121 +/- 22; p less than 0.01 for both 24 and 48 h versus fresh PMN). Likewise, while the number of high-affinity receptors increased (fresh PMN = 25,000 +/- 6,000 receptors/PMN; 24 hours = 95,000 +/- 21,000; 48 hours = 161,000 +/- 40,000; p less than 0.01 for both 24 and 48 hours versus fresh PMN). No abnormality was found in the ability of stored PMN to down-regulate FMLP receptors in the presence of ligand or to reexpress FMLP receptors after a 15-minute incubation in the absence of ligand.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Granulócitos/ultraestrutura , Receptores Imunológicos/metabolismo , Preservação de Sangue , Humanos , N-Formilmetionina Leucil-Fenilalanina , Receptores de Formil Peptídeo , Receptores Imunológicos/fisiologiaRESUMO
Granulocytes stored in the blood bank prior to transfusion undergo progressive decrements in their ability to circulate and migrate in vivo and to migrate in vitro (chemotaxis). The pathogenesis of granulocyte (PMN) chemotaxis (CTX) dysfunction after room-temperature storage of PMN is unclear. Previous work in the authors' laboratory and others led to the hypothesis that intracellular transmission of chemotactic signals, referred to as stimulus-response coupling (SRC), might be abnormal in stored PMN. This report presents an investigation of the ability of fresh and stored PMN to generate and respond to leukotriene-B4 (LTB4), the chief intracellular amplifier of SRC for CTX. PMN were sampled from concentrates within 4 hours of collection and after 24 and 48 hours of storage in transfer packs at room temperature (RT). Fresh, stimulated PMN synthesized 202 +/- 51 ng of LTB4 and 110 +/- 11 ng of 5-hydroxyeicosatetraenoic acid (HETE) per 10(7) PMN. Synthesis of LTB4, but not HETE was significantly decreased after 24 hours' storage, and LTB4 and HETE synthesis decreased after 48 hours. The incubation of stored PMN with arachidonic acid (AA) maintained levels of LTB4 synthesis in PMN stored for 24 but not 48 hours. Also, the CTX defect of stored PMN to F-Met-Leu-Phe (FMLP) was not improved by the supplementation of PMN with exogenous sources of LTB4 or AA.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Quimiotaxia de Leucócito , Granulócitos/citologia , Ácido Araquidônico , Ácidos Araquidônicos/farmacologia , Coleta de Amostras Sanguíneas/métodos , Granulócitos/efeitos dos fármacos , Humanos , Ácidos Hidroxieicosatetraenoicos/biossíntese , Leucotrieno B4/farmacologia , TemperaturaRESUMO
Granulocyte (PMN) concentrates collected for transfusion to septic, neutropenic patients are stored in the blood bank for various periods of time before they are given. Current methods of blood bank storage of PMN concentrates are associated with impaired in vitro PMN chemotaxis (CTX) and in vivo recovery and circulation kinetics after 24 hours of storage. This suggested the possibility that PMN may become hyperadherent during storage. To test this hypothesis, PMN concentrates were harvested and stored at both 22 and 6 degrees C and their adherence properties to relevant biologic surfaces, endothelial cell (EC) monolayers, and extracellular matrix (ECM) derived from endothelium were measured. Adherence was measured within 4 hours of collection and after 24 and 48 hours of storage. The aggregation properties of fresh and stored PMN were also studied. The adherence of fresh, unstimulated PMN to EC and ECM (31 +/- 5% and 34 +/- 4%, respectively) increased significantly after storage for 24 hours (EC = 41 +/- 8%; ECM = 43 +/- 4%) at 22 degrees C. F-Met-Leu-Phe (FMLP) stimulated the adherence of fresh PMN (EC = 37 +/- 4%; ECM = 42 +/- 4%; p less than 0.05). The adherence of PMN stored at 22 degrees C was further stimulated by FMLP (EC = 46 +/- 6%; ECM = 50 +/- 4%). PMN stored at 6 degrees C had significantly higher adherence than PMN stored at 22 degrees C, and the percentage of increase in adherence induced by FMLP was attenuated in PMN stored at 6 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Adesão Celular/efeitos dos fármacos , Endotélio/citologia , Granulócitos/citologia , Temperatura , Coleta de Amostras Sanguíneas , Quimiotaxia de Leucócito , Matriz Extracelular/ultraestrutura , Humanos , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Fatores de TempoAssuntos
Hipercolesterolemia/tratamento farmacológico , Lovastatina/uso terapêutico , Adulto , Idoso , Resinas de Troca Aniônica/uso terapêutico , HDL-Colesterol/sangue , LDL-Colesterol/sangue , Quimioterapia Combinada , Feminino , Humanos , Hiperlipoproteinemia Tipo II/tratamento farmacológico , Lovastatina/efeitos adversos , Masculino , Pessoa de Meia-Idade , Niacina/uso terapêutico , Cooperação do PacienteRESUMO
STUDY OBJECTIVE: To evaluate the efficacy and tolerability of lovastatin under controlled conditions in heterozygous familial hypercholesterolemia. DESIGN: Randomized, double-blind, placebo-controlled, multicenter trial. SETTING: Five lipid clinics with a central laboratory and coordinating center. PATIENTS: 101 adult patients with heterozygous familial hypercholesterolemia. INTERVENTIONS: Patients were on a lipid-lowering diet throughout the study. After a 4-week placebo baseline period, patients were randomized to five equal treatment groups. Each group received a different sequence of placebo or lovastatin 5 to 40 mg twice daily or 20 to 40 mg once daily in the evening, during three consecutive 6-week periods. MEASUREMENTS AND MAIN RESULTS: The mean reductions in total plasma cholesterol and low-density lipoprotein cholesterol across the dosage ranges were 14% to 34% and 17% to 39%, respectively (p compared with zero and placebo less than 0.01). High-density lipoprotein cholesterol and apolipoproteins AI and AII rose slightly. Apolipoprotein B fell substantially at the higher dosage levels (-23% at 40 mg twice daily, p less than 0.01), indicating a reduction in the concentration of circulating low-density lipoprotein particles. Maximum response was achieved in 4 to 6 weeks. Twice-daily dosing was slightly more efficient than once-daily dosing. Of those patients receiving 40 mg twice a day, 89% had a fall in low-density lipoprotein cholesterol of at least 20%, and 61% had a fall of at least 40%. Adverse effects attributable to lovastatin were minimal, and no patient was withdrawn from the study. CONCLUSION: Lovastatin was well tolerated and effective in the treatment of familial hypercholesterolemia.
Assuntos
Hiperlipoproteinemia Tipo II/tratamento farmacológico , Lovastatina/uso terapêutico , Adulto , Alanina Transaminase/sangue , Apolipoproteínas B/sangue , Catarata/induzido quimicamente , Colesterol/sangue , Método Duplo-Cego , Esquema de Medicação , Avaliação de Medicamentos , Feminino , Heterozigoto , Humanos , Hiperlipoproteinemia Tipo II/genética , Lovastatina/administração & dosagem , Lovastatina/efeitos adversos , Masculino , Distribuição AleatóriaRESUMO
We previously showed that an artificial blood substitute containing perfluorocarbons, Fluosol-DA, inhibited both neutrophil migration and adherence, due to its detergent component, Pluronic F-68. The purpose of the studies we report here was to determine if Fluosol or Pluronic might also reduce in vivo neutrophil migration and impair host resistance to bacterial infection. We studied in vivo PMN migration by injecting mice intraperitoneally (IP) with glycogen, followed by intravenous (IV) infusion of saline, Fluosol, or Pluronic. Peritoneal lavage after eight hours showed a significant decrease in the accumulation of PMN in lavage fluids of animals given either Fluosol or Pluronic (control--.19 +/- .03 X 10(6) PMN/mL, glycogen--1.35 +/- .14; glycogen/Fluosol--0.63 +/- .12; glycogen/Pluronic--0.69 +/- .07). We ascertained the effect of Fluosol and Pluronic on infection mortality by injecting mice IV with saline, Fluosol, or Pluronic, followed by a quantity of E coli (0.6 X 10(7] IP shown in preliminary studies to kill 20% to 50% of the mice in 24 hours. The 24-hour mortality was 14/45-saline, 24/32-Fluosol (chi 2 = 17.1; P less than .001) and 17/23 - Pluronic (chi = 11.2; P less than .001). Neither Fluosol nor Pluronic caused mortality without E coli. The increase in infection mortality occurred when Fluosol was given either two hours before, or simultaneously with E coli, but only with the simultaneous administration of bacteria and Pluronic. Pluronic did not alter reticuloendothelial system (RES) clearance function. These studies indicate that, in an animal model, Fluosol-DA, due to its detergent component Pluronic F-68, impaired neutrophil delivery to an inflammatory locus, and resulted in an increased rate of infection mortality. Since Pluronic did not result in RES blockade, but did impair the delivery of PMN to an inflammatory locus, our results suggest that the latter effect is responsible for the increase in infection mortality.
Assuntos
Substitutos Sanguíneos/farmacologia , Movimento Celular/efeitos dos fármacos , Neutrófilos/citologia , Peritonite/mortalidade , Animais , Quimiotaxia de Leucócito/efeitos dos fármacos , Modelos Animais de Doenças , Combinação de Medicamentos/farmacologia , Infecções por Escherichia coli , Fluorocarbonos/farmacologia , Derivados de Hidroxietil Amido , Camundongos , Camundongos Endogâmicos , Cavidade Peritoneal/citologia , Peritonite/sangue , Peritonite/etiologiaRESUMO
Optimal function of transfused granulocytes (PMNs) requires adequate glycogen metabolism. Previous studies in our laboratory suggested that stored PMNs had decreased glycogen. We report here the glycogen content and chemotaxis of stored PMNs, and the ability of fresh and stored PMNs to use glycogen as the fuel source for chemotaxis. PMNs were prepared from 8 fresh units of blood drawn into citrate-phosphate-dextrose-adenine, suspended at 2 or 8 X 10(7) PMN per ml in autologous plasma with or without 15 mM sodium bicarbonate, and stored at 22 to 24 degrees C in transfer packs for 48 hours. Glycogen was measured on resting PMNs, and after challenge with opsonized zymosan and F-Met-Leu-Phe (FMLP). The chemotaxis of fresh and stored PMNs was measured in the presence or absence of extracellular glucose. Fresh PMNs contained 10.3 +/- 0.5 (mean +/- SEM) micrograms of glycogen per 10(6) PMN. Glycogen decreased by 4.2 +/- 0.9 micrograms per 10(6) PMN after challenge with opsonized zymosan and by 1.1 +/- 0.6 micrograms per 10(6) PMN after FMLP. After 48 hours of storage, neutrophil glycogen increased by 18 percent, except in units stored at a concentration of PMN of 8 X 10(7) per ml without sodium bicarbonate. In PMNs from these units stored without bicarbonate, glycogen decreased by 9 percent (p less than .05), and there was a 19 and 55 percent decrease in the ability of PMN from these units to metabolize glycogen after exposure to opsonized zymosan and FMLP, respectively (p less than 0.05). In addition, in PMNs from units stored at a concentration of PMN of 8 X 10(7) per ml without bicarbonate, there was a 47 and 70 percent decrease in chemotaxis at 24 and 48 hours, respectively (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Preservação de Sangue , Glicogênio/sangue , Neutrófilos/metabolismo , Glicemia/metabolismo , Sobrevivência Celular , Quimiotaxia de Leucócito , Humanos , Lactatos/sangue , Ácido Láctico , Neutrófilos/fisiologiaRESUMO
We investigated the possibility that the functional impairment in neutrophil (PMN) chemotaxis which occurs during granulocyte concentrate storage might be due to autotoxicity from the release of neutrophil granule contents during storage. Preliminary experiments confirmed that the exposure of fresh PMNs to the intracellular contents of disrupted PMNs, decreased the subsequent chemotaxis of the fresh PMNs by 63 +/- 5 percent compared to control PMNs (p less than .01). Freshly harvested neutrophils were stored at low (2 X 10(7) PMN/ml) or high cell concentration (8 X 10(7) PMN/ml) with or without 15 mM sodium bicarbonate (in order to maintain pH). Prior to storage, and 24 and 48 hours after storage at 22 to 24 degrees C, we measured the cell and unit plasma content of lactate dehydrogenase (LDH), beta-glucuronidase, and lysozyme. These enzymes served as markers for cell lysis, and primary and specific neutrophil granule contents, respectively. We also measured the effect on neutrophil chemotaxis of adding a protease inhibitor, phenylmethylsulfonyl fluoride (PMSF), to the storage medium. In addition, we measured the ability of PMNs to degranulate in response to an inflammatory stimulus before and after storage. The cell content of granule markers was largely unchanged during storage, except in the case of the units at a concentration of 8 X 10(7) PMN per ml stored without bicarbonate. In these units, lysozyme activity decreased by 15 +/- 7 percent after 48 hours of storage (p less than 0.02 vs. fresh PMNs). Likewise, the plasma content of LDH, beta-glucuronidase, and lysozyme increased significantly during storage, especially in units of high cell concentration stored without bicarbonate.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Preservação de Sangue , Grânulos Citoplasmáticos/enzimologia , Neutrófilos , Quimiotaxia de Leucócito/efeitos dos fármacos , Glucuronidase/sangue , Humanos , L-Lactato Desidrogenase/sangue , Muramidase/sangue , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Neutrófilos/enzimologia , Neutrófilos/fisiologia , Fluoreto de Fenilmetilsulfonil/farmacologia , Fatores de TempoRESUMO
Our aim was to determine the effects of the substitution of sucrose polyester (SPE) for dietary fat in a 16-week outpatient study in 36 obese subjects with primary hypercholesterolemia. The subjects were randomized into three groups who followed a 16-week treatment period where all subjects received hypocaloric diets which provided approximately 7 kcal/lb body weight, a polyunsaturated/saturated (P/S) fat ratio of 0.9, and 180 mg cholesterol/day. The percentages of calories as fat in the 3 groups were as follows: a low fat diet group (n = 12) received 27% of dietary calories as fat, a low fat plus SPE group (n = 13) received 25% of calories as fat plus 27 g SPE/day as a bread spread and salad dressing, and a third group (placebo, n = 11) received 37% of calories as fat with a 27 g/day conventional fat placebo (bread spread and salad dressing). Mean weight loss from baseline in the 16 week treatment period was 2.6, 3.9, and 3.4% respectively in the placebo, diet, and SPE groups, p less than .05 for each group, without significant differences between the groups. There was a mean reduction of low density lipoprotein cholesterol (LDL-C) of 16% in the SPE group (p less than .05), more than twice the reductions in the placebo and diet groups, 5% and 6%, respectively. There was a mean 20% reduction in the SPE group in triglyceride and very low density lipoprotein cholesterol (p less than .05), compared to 7 and 10% reductions in the placebo and diet groups respectively. The degree of weight loss was correlated with the degree of reduction in LDL-C in the low fat diet group, and in the low fat diet group plus SPE (r = 0.59 for both groups). Without confounding by different levels of dietary cholesterol or P/S, SPE induced significant reductions in LDL-C in hypercholesterolemic obese subjects beyond the effects of weight loss alone. The effects of SPE were significantly greater than those achieved by the use of a diet which severely limited conventional dietary fat intake (to 40 g/day). SPE in the form of a bread spread and a salad dressing is a practical formulation for outpatient hypocholesterolemic low fat diets and provides the lubricity and organoleptic benefits of authentic foods without the dense caloric content of digestible fats.
Assuntos
Anticolesterolemiantes/uso terapêutico , Gorduras na Dieta/administração & dosagem , Ácidos Graxos , Hipercolesterolemia/dietoterapia , Obesidade/dietoterapia , Sacarose/análogos & derivados , Adulto , Idoso , Ingestão de Energia , Estudos de Avaliação como Assunto , Ácidos Graxos Insaturados/administração & dosagem , Feminino , Humanos , Hipercolesterolemia/sangue , Lipídeos/sangue , Lipoproteínas/sangue , Masculino , Pessoa de Meia-Idade , Obesidade/sangue , Cooperação do Paciente , Distribuição Aleatória , Sacarose/uso terapêutico , Vitaminas/sangueRESUMO
In view of previous studies demonstrating a significant correlation between adenosine triphosphate (ATP) depletion and impairment of chemotaxis (CTX) during granulocyte (PMN) storage, we sought to quantitate the relationship between CTX and PMN energy metabolism. We incubated PMNs at 37 degrees C with 2-deoxyglucose (2-dg) in the presence of 5 mmol/L glucose. As expected, ATP inhibition by 2-dg was time-dependent (T 1/2, 18 minutes) and dose-dependent, with half-maximal inhibition of ATP (ID50) with 1.3 +/- .3 mmol/L 2-dg. Similar concentrations of 2-dg inhibited lactate generation, phagocytosis, superoxide anion generation, and degranulation. The random migration of PMNs was inhibited by somewhat higher concentrations of 2-dg (ID50, 12 mmol/L). In contrast, up to 40 mmol/L 2-dg did not inhibit CTX toward synthetic peptides or activated serum. In fact, 2-dg consistently increased the CTX of PMNs toward 10(-8) mol/L f-Met-Leu-Phe (fMLP), to a maximum of 450% of control CTX using 15 mmol/L 2-dg. Half-maximal stimulation (ED50) of CTX occurred at 6.3 +/- 1.0 mmol/L 2-dg. Although maximal CTX toward optimal concentrations of fMLP was consistently increased with 2-dg, the ED50 of CTX to fMLP was unchanged (ED50 with glucose, 2.0 +/- 0.6 nmol/L fMLP; ED50 with 2-dg 2.2 +/- 0.7 nmol/L fMLP), and 2-dg did not increase fMLP receptors. In the absence of glucose, 2-dg exerted similar effects on ATP and CTX, but at doses 30- to 50-fold lower than in the presence of glucose. Other glycolytic inhibitors (iodoacetamide and sodium fluoride) exerted similar effects. Additional studies indicated that CTX enhancement by 2-dg (a) required Mg++ but not Ca++, (b) occurred with PMNs from a patient with chronic granulomatous disease, (c) was unaltered in the presence of inhibitors of proteolysis, (d) was not due to generation of a soluble agent, (e) was not due to alterations in PMN adherence, and (f) was not due to inhibition of glycosylation. We conclude that the chemotaxis, but not the random migration, of PMNs is surprisingly resistant to inhibition of energy metabolism and depletion of ATP, since concentrations of 2-dg that decreased ATP and other cell functions by more than 50% not only did not inhibit, but actually stimulated, CTX. These studies also indicate that the previously reported correlation between ATP depletion and CTX impairment observed in stored PMNs are not causally related.
Assuntos
Trifosfato de Adenosina/metabolismo , Quimiotaxia de Leucócito , Metabolismo Energético , Neutrófilos/fisiologia , Quimiotaxia de Leucócito/efeitos dos fármacos , Desoxiglucose/farmacologia , Humanos , Neutrófilos/efeitos dos fármacosRESUMO
We investigated the effect of a candidate artificial blood substitute, Fluosol-DA (FDA), on human neutrophil function in a serum-free medium. In a 50% (vol/vol) mixture with polymorphonuclear cells (PMN), FDA had no effect on PMN viability, phagocytosis, superoxide anion generation, degranulation, or bactericidal activity. In striking contrast, the random migration and chemotaxis of PMN to both f-Met-Leu-Phe (fMLP) and activated serum were inhibited by 98% +/- 2%, 95% +/- 2%, and 88% +/- 6%, respectively. Inhibition of chemotaxis by FDA required no preincubation, was dose-dependent (50% inhibition [ID50] with a 14% vol/vol mixture with FDA), and was fully reversible by washing PMN free of FDA after one hour but not after 18 hours of incubation (32% +/- 11% inhibition of chemotaxis). FDA itself was not chemotactic and did not impair either the chemotactic activity or binding of fMLP to PMN. FDA also inhibited PMN adhesion (ID50, 9 +/- 1 vol/vol%). The inhibitory component of FDA was found to be its detergent additive, Pluronic F-68, which inhibited random migration, chemotaxis, and adhesion with ID50s of 1.4, 2.4, and 2.9 mg/mL, respectively (equivalent to FDA concentrations of 5, 9, and 11 vol/vol%, respectively). All the other components of FDA were noninhibitory. Plasma samples from humans injected with 8 mL/kg FDA and plasma samples from rabbits injected with 16 mL/kg FDA or an equivalent concentration of Pluronic F-68, when mixed with autologous PMN, also severely inhibited PMN chemotaxis. We conclude that exposure of PMN to clinically relevant concentrations of FDA inhibits PMN migration, presumably due to inhibition of adhesion. The inhibitory effect is entirely due to the detergent, Pluronic F-68. Artificial blood substitutes containing Pluronic F-68 may compromise the ability of PMN to prevent or effectively control microbial infections.
Assuntos
Substitutos Sanguíneos/farmacologia , Fluorocarbonos/farmacologia , Fagócitos/efeitos dos fármacos , Animais , Inibição de Migração Celular , Movimento Celular/efeitos dos fármacos , Quimiotaxia de Leucócito/efeitos dos fármacos , Combinação de Medicamentos/farmacologia , Humanos , Derivados de Hidroxietil Amido , Técnicas In Vitro , Cinética , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Fagócitos/fisiologia , CoelhosRESUMO
Through technological advances in granulocyte collection, it has become possible to collect neutrophils (PMNs) routinely in high concentration (greater than 5 X 10(7) PMN/ml) for transfusion. Previous studies in this laboratory suggested that storage of neutrophils for transfusion at high PMN concentrations resulted in impaired adenosine triphosphate (ATP) and hydrogen ion maintenance. The studies we report here were designed to assess the effect of PMN storage at concentrations which are usual (2 X 10(7) PMN/ml), intermediate (5 X 10(7) PMN/ml), and high (8 X 10(7) PMN/ml) on chemotactic responses, and to identify variables which are easily measured and might predict the chemotactic function of stored PMNs. Granulocyte concentrates were stored in plastic bags at 2,5, and 8 X 10(7) PMN per ml, with or without 15 mM bicarbonate (HCO3). The random migration (RM) chemotaxis (CTX), ATP, and relative cell size (VOL) of the fresh and stored cells and the pH, glucose, and lactate concentrations in the supernatant medium were measured in the freshly prepared units after 24 and 48 hours storage at room temperature. We found that RM, CTX, ATP, glucose, and pH decreased significantly (p less than .02) following storage for 24 and 48 hours, particularly in units stored at the higher cell concentrations. Cell volume and lactate increased significantly with storage for 24 and 48 hours, and these values were also greater in units stored at the higher cell concentration (p less than .02).(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Granulócitos/imunologia , Concentração de Íons de Hidrogênio , Trifosfato de Adenosina/análise , Bicarbonatos/farmacologia , Preservação de Sangue , Quimiotaxia , HumanosRESUMO
Methionine-containing chemotactic peptides, such as formyl-methionyl-leucyl-phenylalanine (FMLP), are inactivated via a neutrophil-derived, myeloperoxidase-mediated oxidation of the methionine residue. We report that extracellular inactivation of FMLP by myeloperoxidase modulates the apparent binding of methionine-containing chemotactic peptides to their surface receptors. Inhibitors of myeloperoxidase enhanced FMLP binding. At subsaturating concentrations of 3H-FMLP (20 nM), 1 mM cyanide (KCN) increased the binding of 3H-FMLP to human neutrophils (PMN) by 51% +/- 12%. Similar increases occurred with 0.1 mM azide and 10 mM aminotriazole (ATZ). KCN had little effect on maximal 3H-FMLP binding to PMN at saturation (control-17,040 +/- 910 receptors/PMN; KCN-16,820 +/- 1,940 receptors/PMN), but decreased the concentration of 3H-FMLP required to half-saturate the PMN receptors (control-39 +/- 3 nM; KCN-17 +/- 1 nM). ATZ gave similar results. The binding to PMN of the non-methionine-containing chemotactic peptide 125I-formyl-norleucyl-leucyl-phenylalanyl-norleucyl-tyrosyl-lysine (125I-FNLPNTL) was unaltered by KCN. Also, the binding of 3H-FMLP to myeloperoxidase-deficient PMN was unaltered by KCN. Both KCN and ATZ decreased the oxidation of FMLP by PMN. Finally, ATZ (but not KCN) enhanced the chemotactic migration of PMN in response to submaximal concentrations of FMLP. These studies show that intact PMN inactivate methionine-containing chemotactic peptides by a pathway that is sensitive to myeloperoxidase inhibitors and is absent in myeloperoxidase-deficient PMN. This action results in an apparent decrease in the affinity of the chemotactic peptide receptor for methionine-containing chemotactic peptides, which may modulate chemotatic events in inflammatory loci.