RESUMEN
BACKGROUND: Heart failure (HF) patients are at a greater risk for nosocomial infections due to their higher prevalence of comorbidities, readmission rates, and prolonged hospital stays. Clostridioides difficile infection (CDI) remains a common nosocomial infection in hospitalized patients. AIM: To identify patients with a diagnosis of acute HF and stratified based on the presence of CDI, using the National Inpatient Sample (NIS) database from 2016 to 2020. METHODS: Adjusted odds ratios (aOR) of in-hospital outcomes were calculated, and a propensity-matched analysis was performed. FINDINGS: Of 536,595 acute HF hospitalizations in this timeframe, 3030 were also diagnosed with CDI (0.56%). Patients with acute HF and CDI had significantly higher in-hospital mortality (adjusted odds ratio: (aOR): 1.91), cardiac arrest (aOR: 1.66), and use of mechanical circulatory support (MCS) (aOR 2.42). In propensity-matched analysis, in-hospital mortality (13.71 vs 8.44%; P=0.005), septic shock (7.54 vs 3.33%; P=0.002), and use of MCS (7.19 vs 3.68%; P=0.009) were significantly higher in HF patients with CDI than without. Coexisting neurological disease (aOR: 3.74) and liver disease (aOR: 2.97) showed the strongest association as independent predictors of mortality. HF patients with CDI had longer hospital stays (14.45 ± 19.40 vs 5.44 ± 7.10 days; P<0.0001) and higher inflation-adjusted total hospital costs ($186,225 ± 376,485 vs $60,740 ± 153,992; P<0.001) compared to those without CDI. CONCLUSION: The occurrence of concomitant CDI in patients admitted with acute HF exacerbation is associated with worse in-hospital outcomes and deaths as well as longer hospitalizations and greater financial cost.
Asunto(s)
Clostridioides difficile , Infecciones por Clostridium , Insuficiencia Cardíaca , Humanos , Pacientes Internos , Hospitalización , Infecciones por Clostridium/diagnóstico , Insuficiencia Cardíaca/complicaciones , Insuficiencia Cardíaca/epidemiología , Estudios RetrospectivosRESUMEN
Adjunctive use of phosphodiesterase (PDE) inhibitor can enhance antiplatelet and vasoprotective properties in patients with cardiovascular disease. The aim of this study was to evaluate the impact of PDE5 inhibitor dipyridamole on platelet function in stented patients with high platelet reactivity (HPR) during dual antiplatelet therapy (DAPT) with aspirin and clopidogrel. Patients with HPR after 600-mg clopidogrel loading were randomly assigned to adjunctive dipyridamole 75 mg twice daily to standard DAPT (DIP group; n = 45) or double-dose clopidogrel of 150 mg daily (DOUBLE group; n = 46) for 30 days. Platelet function was assessed at baseline and 30-day follow-up with platelet reactivity index (PRI) by vasodilator-stimulated phosphoprotein-phosphorylation (VASP-P) assay and platelet aggregation (PA) by light transmittance aggregometry (LTA). Primary endpoint was PRI at 30-day follow-up. HPR was defined as PRI > 50%. Baseline platelet function did not differ between the groups. Following 30-day therapy, platelet function was significantly reduced in the DIP and DOUBLE groups (all p-values ≤ 0.004 and ≤ 0.068, respectively). PRI values were not significantly different between the two groups (mean difference: 3.1%; 95% confidence interval: -2.8% to 9.0%: p = 0.295). PA values and prevalence of HPR were similar between the groups. However, a significant number of patients still exhibited HPR in the DIP (75.6%) and DOUBLE (67.4%) groups. In conclusion, among stented HPR patients, adding dipyridamole to DAPT does not reduce platelet reactivity and prevalence of HPR compared with double-dose clopidogrel therapy, and therefore both strategies are inadequate to overcome HPR.
Asunto(s)
Aspirina/uso terapéutico , Plaquetas/efectos de los fármacos , Dipiridamol/uso terapéutico , Resistencia a Medicamentos , Intervención Coronaria Percutánea , Inhibidores de Fosfodiesterasa 5/uso terapéutico , Inhibidores de Agregación Plaquetaria/uso terapéutico , Ticlopidina/análogos & derivados , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Anciano , Aspirina/efectos adversos , Biomarcadores/sangre , Plaquetas/metabolismo , Moléculas de Adhesión Celular/sangre , Clopidogrel , Citocromo P-450 CYP2C19/genética , Citocromo P-450 CYP3A/genética , Dipiridamol/efectos adversos , Quimioterapia Combinada , Femenino , Genotipo , Humanos , Masculino , Proteínas de Microfilamentos/sangre , Persona de Mediana Edad , Intervención Coronaria Percutánea/efectos adversos , Intervención Coronaria Percutánea/instrumentación , Inhibidores de Fosfodiesterasa 5/efectos adversos , Fosfoproteínas/sangre , Agregación Plaquetaria/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/efectos adversos , Pruebas de Función Plaquetaria , Estudios Prospectivos , República de Corea , Stents , Ticlopidina/efectos adversos , Ticlopidina/uso terapéutico , Factores de Tiempo , Resultado del TratamientoRESUMEN
CYP2C19 genotype has been shown to impact response to clopidogrel 75-mg but not prasugrel 10-mg. Here, we assessed effects of CYP2C19 metaboliser status on pharmacokinetics (PK) and pharmacodynamic (PD) responses to prasugrel 5-mg and 10-mg and clopidogrel 75-mg using data from two PK/PD studies in stable coronary artery disease (CAD) patients (GENERATIONS and FEATHER). Active metabolite concentrations (area under the curve, AUC[0-tlast]), maximum platelet aggregation (MPA) measured by light transmission aggregometry, vasodilator-stimulated phosphoprotein platelet reactivity index, and VerifyNow P2Y12-platelet reaction units (VN-PRU) were analysed by CYP2C19-predicted phenotype (extensive metaboliser [EM; N=154], *2-*8 non-carriers, vs reduced metaboliser [RM; N=41],*2-*8 carriers/*17 non-carriers). AUC(0-tlast) was unaffected by metaboliser status for prasugrel 5-mg and 10-mg (geometric mean EM/RM ratios 1.00, 95% confidence interval [CI]: 0.86,1.17, p>0.99; and 0.97, 95% CI:0.85,1.12, p=0.71, respectively), but was lower among RMs receiving clopidogrel 75-mg (1.37, 95% CI:1.14,1.65, p<0.001). Platelet reactivity was not significantly affected by CYP2C19 metaboliser status for prasugrel 5-mg, or for prasugrel 10-mg by MPA and VN-PRU, but for clopidogrel 75-mg was significantly higher in reduced metabolisers (all measures p<0.01). Prasugrel 10-mg showed greater antiplatelet effects vs clopidogrel 75-mg (all comparisons p<0.001). Prasugrel 5-mg showed greater antiplatelet effects vs clopidogrel 75-mg in RMs (all p<0.001), and comparable effects in EMs (all p≥0.37). In contrast to clopidogrel, prasugrel active metabolite PK was not influenced by CYP2C19 genotype. Antiplatelet effect for prasugrel 10-mg was greater irrespective of metaboliser status and for prasugrel 5-mg was greater for RMs and comparable for EMs as compared to clopidogrel 75-mg.
Asunto(s)
Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Citocromo P-450 CYP2C19/genética , Piperazinas/farmacocinética , Inhibidores de Agregación Plaquetaria/farmacocinética , Tiofenos/farmacocinética , Ticlopidina/análogos & derivados , Anciano , Protocolos Clínicos , Clopidogrel , Enfermedad de la Arteria Coronaria/genética , Femenino , Genotipo , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Piperazinas/administración & dosificación , Agregación Plaquetaria/efectos de los fármacos , Agregación Plaquetaria/genética , Inhibidores de Agregación Plaquetaria/administración & dosificación , Polimorfismo Genético , Clorhidrato de Prasugrel , Tiofenos/administración & dosificación , Ticlopidina/administración & dosificación , Ticlopidina/farmacocinética , Resultado del TratamientoRESUMEN
Aspirin and statin therapy are mainstay treatments in patients with coronary artery disease (CAD). The relation between statin therapy, in vivo thromboxane (Tx) generation; a marker of inflammation, and blood thrombogenicity has never been explored. Urinary 11-dehydro (dh) TxB2 was determined in patients with suspected CAD on 325 mg daily aspirin therapy prior to undergoing cardiac catheterisation (n=281). Thrombogenicity was estimated by thrombelastographic measurement of thrombin-induced platelet-fibrin clot strength (TIP-FCS) and lipids/lipoproteins were determined by vertical density gradient ultracentrifugation/ELISA. The influence of statin therapy and dose was analysed by the atorvastatin equivalent dose (5-10 mg, 20-40 mg, or 80 mg daily). Statin therapy (n=186) was associated with a dose-dependent reduction in urinary 11-dh TxB2 (p=0.046) that was independent of LDL and apo B100 levels but was strongly related to TIP-FCS (p=0.006). By multivariate analysis, no statin therapy (n=95) and female gender were independently associated with high urinary 11-dh TxB2 [OR=2.95 (0.1.57-5.50, p=0.0007); OR=2.25 (1.24-4.05, p=0.007)], respectively. In aspirin-treated patients, statin therapy was independently and inversely associated with inflammation in a dose-dependent manner. Elevated 11-dh TxB2 was associated with a prothrombotic state indicated by high TIP-FCS. Our data suggest that measurement of urinary 11-dTxB2 may be a useful method to optimise statin dosing in order to reduce thrombotic risk.
Asunto(s)
Aspirina/administración & dosificación , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Inhibidores de Hidroximetilglutaril-CoA Reductasas/uso terapéutico , Mediadores de Inflamación/orina , Inhibidores de Agregación Plaquetaria/administración & dosificación , Tromboxano B2/análogos & derivados , Anciano , Biomarcadores/orina , Coagulación Sanguínea/efectos de los fármacos , Distribución de Chi-Cuadrado , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/diagnóstico , Enfermedad de la Arteria Coronaria/orina , Relación Dosis-Respuesta a Droga , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Lípidos/sangre , Masculino , Persona de Mediana Edad , Análisis Multivariante , Oportunidad Relativa , Modelos de Riesgos Proporcionales , Factores de Riesgo , Factores Sexuales , Tromboelastografía , Tromboxano B2/orina , Resultado del Tratamiento , UltracentrifugaciónAsunto(s)
Hidrocarburo de Aril Hidroxilasas/genética , Intervención Coronaria Percutánea , Farmacogenética , Ticlopidina/análogos & derivados , Adenosina Difosfato/química , Adulto , Anciano , Alelos , Pueblo Asiatico , Clopidogrel , Estudios de Cohortes , Citocromo P-450 CYP2C19 , Femenino , Genotipo , Heterocigoto , Humanos , Masculino , Persona de Mediana Edad , Análisis Multivariante , Oportunidad Relativa , Proyectos Piloto , Inhibidores de Agregación Plaquetaria/uso terapéutico , Pruebas de Función Plaquetaria , Polimorfismo de Nucleótido Simple , Ticlopidina/uso terapéuticoAsunto(s)
Plaquetas/efectos de los fármacos , Moléculas de Adhesión Celular/sangre , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Proteínas de Microfilamentos/sangre , Fosfoproteínas/sangre , Inhibidores de Agregación Plaquetaria/uso terapéutico , Agregación Plaquetaria/efectos de los fármacos , Pruebas de Función Plaquetaria , Antagonistas del Receptor Purinérgico P2Y/uso terapéutico , Receptores Purinérgicos P2Y12/efectos de los fármacos , Biomarcadores/sangre , Plaquetas/metabolismo , Enfermedad de la Arteria Coronaria/sangre , Quimioterapia Combinada , Humanos , Valor Predictivo de las Pruebas , Curva ROC , Ensayos Clínicos Controlados Aleatorios como Asunto , Receptores Purinérgicos P2Y12/sangre , Resultado del TratamientoRESUMEN
PA32540 combines 325 mg enteric-coated (EC) aspirin (ASA) with 40 mg immediate-release omeprazole; its influence on the antiplatelet effect of clopidogrel (C) is unknown. In this randomized, open-label study, subjects (n = 30) were treated with (i) 300 mg C + 325 mg ECASA followed by 75 mg C + 325 mg ECASA on days 2-7, (ii) 300 mg C + PA32540 followed by 75 mg C + PA32540 on days 2-7, or (iii) PA32540 in the morning + 300 mg C 10 h later on day 1 and PA32540 in the morning + 75 mg C 10 h later on days 2-7. We analyzed the noninferiority of PA32540 relative to ECASA, as defined by the upper bound of the 95% confidence interval ≤10% for the difference in least-square means of platelet inhibition between the treatments. As compared to ECASA+C, synchronous treatment of PA32540+C was not noninferior, whereas the spacing strategy of PA32540+C was noninferior. Spacing the administration of PA32540 and clopidogrel lessens the interaction observed with synchronous administration; PA32540 administration with clopidogrel may be associated with a different antiplatelet profile.
Asunto(s)
Aspirina/farmacología , Omeprazol/farmacología , Inhibidores de Agregación Plaquetaria/farmacología , Ticlopidina/análogos & derivados , Adulto , Antiulcerosos/administración & dosificación , Antiulcerosos/farmacología , Aspirina/administración & dosificación , Clopidogrel , Estudios Cruzados , Relación Dosis-Respuesta a Droga , Esquema de Medicación , Combinación de Medicamentos , Interacciones Farmacológicas , Femenino , Humanos , Análisis de los Mínimos Cuadrados , Masculino , Omeprazol/administración & dosificación , Inhibidores de Agregación Plaquetaria/administración & dosificación , Comprimidos Recubiertos , Ticlopidina/administración & dosificación , Ticlopidina/farmacologíaRESUMEN
A common functional variant in paraoxonase 1 (PON1), Q192R, was recently reported to be a major determinant of clopidogrel response. This variant was genotyped in 566 participants of the Amish Pharmacogenomics of Anti-Platelet Intervention (PAPI) study and in 227 percutaneous coronary intervention (PCI) patients. Serum paraoxonase activity was measured in a subset of 79 PAPI participants. PON1 Q192R was not associated with pre- or post-clopidogrel platelet aggregation in the PAPI study (P = 0.16 and P = 0.21, respectively) or the PCI cohort (P = 0.47 and P = 0.91, respectively). The Q192 allele was not associated with cardiovascular events (hazard ratio (HR) 0.46, 95% confidence interval (CI) 0.20-1.06; P = 0.07). No correlation was observed between paraoxonase activity and post-clopidogrel platelet aggregation (r(2) < 0.01, P = 0.78). None of 49 additional PON1 variants evaluated was associated with post-clopidogrel platelet aggregation. These findings do not support a role for PON1 as a determinant of clopidogrel response.
Asunto(s)
Arildialquilfosfatasa/genética , Enfermedades Cardiovasculares/genética , Enfermedades Cardiovasculares/mortalidad , Estudios de Asociación Genética/métodos , Variación Genética/genética , Ticlopidina/análogos & derivados , Adulto , Anciano , Enfermedades Cardiovasculares/tratamiento farmacológico , Clopidogrel , Estudios de Cohortes , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Tasa de Supervivencia/tendencias , Ticlopidina/uso terapéutico , Resultado del TratamientoRESUMEN
BACKGROUND: The rate of recovery of platelet function after discontinuation of P2Y(12) inhibitors depends on the reversibility of the antiplatelet effect and the extent of the on-treatment response. P2Y(12) inhibition increases the bleeding risk in patients requiring surgery. OBJECTIVES: To evaluate recovery of platelet function after discontinuation of ticagrelor vs. clopidogrel in stable coronary artery disease (CAD) patients with high levels of platelet inhibition (HPI) during the ONSET/OFFSET study. METHODS: Patients received aspirin 75-100 mg per day and either ticagrelor 90 mg twice-daily or clopidogrel 75 mg daily for 6 weeks. This subanalysis included patients with HPI after the last dose of maintenance therapy, defined as: inhibition of platelet aggregation (IPA) > 75% 4 h post-dose (ADP 20 µm, final extent); < 120 P2Y(12) reaction units 8 h post-dose (VerifyNow P2Y(12) assay); or platelet reactivity index < 50% 8 h post-dose (VASP-P assay). RESULTS: IPA > 75% was observed in 39 out of 47 ticagrelor-treated and 17 out of 44 clopidogrel-treated patients. The rate of offset of IPA over 4-72 h was greater with ticagrelor (IPA %/hour slope: -1.11 vs. -0.67 for clopidogrel; P < 0.0001). Mean IPA was significantly lower with ticagrelor than clopidogrel between 48 and 168 h post-dose (P < 0.01). Similar findings were observed with the other assays. The average time for IPA to decline from 30% to 10% was 50.8 h with ticagrelor vs. 110.4 h with clopidogrel. CONCLUSIONS: In patients with HPI, recovery of platelet function was more rapid after discontinuation of ticagrelor than clopidogrel leading to significantly greater platelet reactivity by 48 h after the last dose in the ticagrelor group.
Asunto(s)
Adenosina/análogos & derivados , Plaquetas/efectos de los fármacos , Plaquetas/fisiología , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Inhibidores de Agregación Plaquetaria/administración & dosificación , Antagonistas del Receptor Purinérgico P2Y/administración & dosificación , Ticlopidina/análogos & derivados , Adenosina/administración & dosificación , Adulto , Anciano , Anciano de 80 o más Años , Clopidogrel , Método Doble Ciego , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pruebas de Función Plaquetaria , Ticagrelor , Ticlopidina/administración & dosificación , Factores de TiempoRESUMEN
UNLABELLED: To study the effect of a new direct acting reversible P2Y(12) inhibitor, elinogrel (PRT060128), and the relation to cytochrome P450 (CYP) polymorphisms in patients with high platelet reactivity (HPR) on standard dual antiplatelet therapy. METHODS AND RESULTS: We studied the pharmacodynamic and pharmacokinetic effects of a single 60-mg oral dose of elinogrel in 20 of 45 previously stented stable patients with HPR. We also genotyped for CYP2C19*2,3,5,17 and CYP3A5*3. Platelet reactivity fell within 4 h of dosing, the earliest time point evaluated as measured by the following assays: maximum 5 and 10 microM ADP LTA (P < 0.001 for both vs. predosing); maximum 20 microM ADP LTA (P < 0.05); VerifyNow (P < 0.001); thrombelastography (P < 0.05); VASP phosphorylation (P < 0.01); and perfusion chamber assay (P < 0.05); this was reversible within 24 h in these same assays (P = ns vs. predosing for all assays). CYP2C19*2 was present in 44% of all patients but was more frequent in HPR patients (77% vs. 16%, P = 0.0004). CONCLUSIONS: HPR is reversibly overcome by a single 60-mg oral dose of elinogrel, a drug now being investigated in a phase 2 trial. CYP2C19*2 was associated with HPR during conventional dual antiplatelet therapy.
Asunto(s)
Hidrocarburo de Aril Hidroxilasas/genética , Aspirina/uso terapéutico , Plaquetas/efectos de los fármacos , Enfermedad de la Arteria Coronaria/terapia , Inhibidores de Agregación Plaquetaria/uso terapéutico , Polimorfismo Genético , Antagonistas del Receptor Purinérgico P2 , Quinazolinonas/uso terapéutico , Sulfonamidas/uso terapéutico , Ticlopidina/análogos & derivados , Adenosina Difosfato , Administración Oral , Anciano , Angioplastia Coronaria con Balón/instrumentación , Hidrocarburo de Aril Hidroxilasas/metabolismo , Coagulación Sanguínea/efectos de los fármacos , Plaquetas/metabolismo , Moléculas de Adhesión Celular/sangre , Clopidogrel , Colágeno/uso terapéutico , Citocromo P-450 CYP2C19 , Citocromo P-450 CYP3A/genética , Quimioterapia Combinada , Femenino , Frecuencia de los Genes , Genotipo , Humanos , Masculino , Proteínas de Microfilamentos/sangre , Persona de Mediana Edad , Fenotipo , Fosfoproteínas/sangre , Fosforilación , Agregación Plaquetaria/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/administración & dosificación , Inhibidores de Agregación Plaquetaria/farmacocinética , Quinazolinonas/administración & dosificación , Quinazolinonas/farmacocinética , Receptores Purinérgicos P2/metabolismo , Receptores Purinérgicos P2Y12 , Stents , Sulfonamidas/administración & dosificación , Sulfonamidas/farmacocinética , Tromboelastografía , Ticlopidina/uso terapéuticoRESUMEN
Percutaneous coronary intervention (PCI) has significantly improved clinical outcomes in coronary artery disease patients. Since PCI is associated with platelet activation, antiplatelet therapy with aspirin, clopidogrel and GPIIb/IIIa inhibitors comprise the cornerstone strategy during and following PCI. The latter agents are arguably the most important drugs we administer to the patient with established coronary artery disease since they are specifically given to prevent the most catastrophic event, the formation of an occlusive arterial thrombus. Numerous clinical trials have confirmed the efficacy of antiplatelet therapy in attenuating recurrent ischaemic event occurrence. Despite the extensive use of antiplatelet therapies, ischaemic event occurrence such as post-procedural myocardial infarction and stent thrombosis still remains an important concern and highlights the need for improved treatment strategies. A major limitation of current treatment is the application of a "one size fits all" strategy advocated by the guidelines that completely ignores the evaluation of the individual antiplatelet response. Pharmacodynamic studies have revealed the limitations of aspirin and clopidogrel treatment that include response variability, and a high prevalence of antiplatelet non-responsiveness associated with significant risk for recurrent ischemic event occurrence. Thus, two major paradoxes in cardiovascular medicine today are: 1) despite the overwhelming evidence that platelet reactivity strongly influences the development of potentially catastrophic events including myocardial infarction and stent thrombosis in the PCI patient, no measurement is made in clinical practice to assess the presence of blood vulnerability (platelet reactivity) and 2) despite the overwhelming evidence that the effect of dual antiplatelet therapy with aspirin and P2Y12 receptor blockers is variable, the guidelines largely recommend a uniform, "one size fits all" dosing of these agents in the PCI patient without any confirmation of an adequate antiplatelet effect.
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Plaquetas/fisiología , Coagulación Sanguínea/fisiología , Plaquetas/efectos de los fármacos , Clopidogrel , Enfermedad Coronaria/tratamiento farmacológico , Enfermedad Coronaria/genética , Humanos , Monitoreo Fisiológico/métodos , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/genética , Activación Plaquetaria/efectos de los fármacos , Adhesividad Plaquetaria , Inhibidores de Agregación Plaquetaria/uso terapéutico , Medicina de Precisión/métodos , Ticlopidina/análogos & derivados , Ticlopidina/uso terapéuticoAsunto(s)
Activación Plaquetaria , Infecciones del Sistema Respiratorio/sangre , Infecciones del Sistema Respiratorio/complicaciones , Adulto , Estudios de Casos y Controles , Femenino , Humanos , Masculino , Persona de Mediana Edad , Infarto del Miocardio/etiología , Pruebas de Función Plaquetaria , Infecciones del Sistema Respiratorio/virología , Accidente Cerebrovascular/etiología , Virosis/sangre , Virosis/complicacionesRESUMEN
INTRODUCTION: We studied the time course of nitric oxide expression in the healing wound and the cell populations responsible for its synthesis. METHODS: Twenty four Lewis rats underwent subcutaneous implantation of polyvinyl alcohol sponges. Rats were sacrificed in groups of three on days 1, 3, 5, 7, 10, 14, and 35 after wounding. The conversion of 3H-labeled arginine to 3H-labeled citrulline, with or without N(G)-L-monomethyl-arginine (L-NMMA) in harvested sponges, was measured. Nitrate/nitrite (NOx) in plasma and wound fluid was quantified by Greiss reaction. Inducible nitric oxide synthase (iNOS) gene expression was determined by Northern analysis and reverse transcriptase-polymerase chain reaction (RT-PCR). Inducible NOS was identified in specific wound cell populations by dual-label flow cytometry. RESULTS: Nitric oxide synthase (NOS) activity peaked at 24 h after wounding (37.7 +/- 0.9 micromol citrulline per milligram sponge), with a steady decline thereafter. Percentage inhibition of NOS activity by l-NMMA was highest on days 1-7 (70-80%). This declined to 50% by day 10 and to 25% by days 14-35. The iNOS gene expression paralleled NOS biochemical activity. RT-PCR confirmed low-level expression up to 10 days after wounding. Plasma NOx levels remained within a narrow range of 22.6 +/- 1.3 to 29.3 +/- 1.5 microM throughout the postwounding period, while corresponding levels in wound fluid (microM) increased steadily from 27 +/- 3.8 on day 1 to 107.2 +/- 10.0 on day 14. Inducible NOS expression was detectable by fluorescence-activated cell sorting in wound macrophages on days 1 and 3 after wounding. CONCLUSIONS: Our findings suggest maximal NOS activity early in cutaneous wound healing, with sustained production up to 10 days after wounding. NOS biochemical activity was paralleled by iNOS gene expression. Plasma NOx remained constant, while wound fluid NOx increased steadily to peak at day 14. Wound macrophages appear to be a source of nitric oxide production in the early phase of wound healing.
Asunto(s)
Óxido Nítrico/metabolismo , Piel/lesiones , Piel/fisiopatología , Cicatrización de Heridas/fisiología , Heridas y Lesiones/fisiopatología , Animales , Inhibidores Enzimáticos/farmacología , Masculino , Nitratos/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico Sintasa de Tipo II , Nitritos/metabolismo , Ratas , Ratas Endogámicas Lew , Factores de Tiempo , omega-N-Metilarginina/farmacologíaRESUMEN
Inducible nitric oxide synthase (iNOS) and its product, nitric oxide, have been shown to play important roles in wound biology. The present study was performed to investigate the role of iNOS in modulating the cytokine cascade during the complex process of skin graft wound healing.Fifteen iNOS-knockout mice and 15 wild-type C57BL/6J mice were subjected to autogenous 1-cm2 intrascapular full-thickness skin grafts. Three animals in each group were killed on postoperative days 3, 5, 7, 10, and 14. Specimens were then analyzed using nonisotopic in situ hybridization versus mRNA of tumor growth factor-beta1, vascular endothelial growth factor, iNOS, endothelial nitric oxide synthase (eNOS), tumor necrosis factor-alpha, and basic fibroblast growth factor, as well as positive and negative control probes. Positive cells in both grafts and wound beds were counted using a Leica microgrid. Scar thickness was measured with a Leica micrometer. Data were analyzed using the unpaired Student's t test. Expression of iNOS was 2- to 4-fold higher in knockout mice than in wild-type mice on postoperative days 5, 7, and 14. Expression of eNOS was 2- to 2.5-fold higher in knockout mice than in wild-type mice on postoperative days 5 and 7. Tumor necrosis factor-alpha expression was 2- to 7-fold higher in knockout mice than in wild-type mice on all postoperative days. In contrast, expression levels of angiogenic/fibrogenic cytokines (vascular endothelial growth factor, basis fibroblast growth factor, and tumor growth factor-beta1) were 2.5- to 4-fold higher in wild-type mice than in knockout mice. Scars were 1.5- to 2.5-fold thicker in knockout mice than in wild-type mice at all time points. All of the above results represent statistically significant differences (p < 0.05). Significantly different patterns of cytokine expression were seen in knockout and wild-type mice. Although the scar layer was thicker in knockout mice, it showed much greater infiltration with inflammatory cells. These data further delineate the modulatory effect of iNOS and nitric oxide in healing skin grafts.
Asunto(s)
Citocinas/biosíntesis , Óxido Nítrico Sintasa/biosíntesis , Trasplante de Piel/fisiología , Cicatrización de Heridas/fisiología , Animales , Hibridación in Situ , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Óxido Nítrico Sintasa de Tipo II , ARN Mensajero/genéticaRESUMEN
BACKGROUND: We have previously shown that the blockade of nitric oxide (NO) synthesis impairs wound healing, in particular collagen synthesis. Conversely, impaired wound healing is accompanied by decreased wound NO synthesis. Fibroblast collagen synthesis, proliferation, and fibroblast-mediated matrix contraction are critical to wound healing. We examined the wound healing-related phenotypic changes that are induced by the loss of inducible nitric oxide synthase (iNOS) gene function in fibroblasts. METHODS: Dermal fibroblasts were obtained from 8- to 12-week-old iNOS--knock out (KO; C57BL/Ai-[KO] Nos2 N5) and wild type mice by an explant technique and used after 1 to 3 passages. Proliferation ([(3)H]-thymidine incorporation) and collagen synthesis ([(3)H]-proline incorporation into collagenase-sensitive protein) were studied after stimulation with 10% fetal bovine serum. Matrix remodeling was assessed by the measurement of the contraction of fibroblast-populated collagen lattices. RESULTS: iNOS-KO fibroblasts proliferated more slowly, synthesized less collagen, and contracted fibroblast-populated collagen lattices more slowly than wild-type fibroblast. Collagen synthesis was restored to normal in KO fibroblasts in response to NO donors (s-nitroso-N-acetylpenicillamine). CONCLUSIONS: iNOS deficiency causes significant impairment in wound healing-related properties of fibroblasts, which suggests that NO plays an important role in wound healing.
Asunto(s)
Óxido Nítrico Sintasa/genética , Óxido Nítrico Sintasa/metabolismo , Cicatrización de Heridas/fisiología , Animales , División Celular/fisiología , Células Cultivadas , Colágeno/biosíntesis , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/enzimología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Donantes de Óxido Nítrico , Óxido Nítrico Sintasa de Tipo II , Penicilamina/análogos & derivados , Penicilamina/farmacología , Piel/citologíaRESUMEN
In order to study wound healing, it is often necessary to administer various wound-active substances by the systemic route. It is unclear whether the observed effects are the result of local or systemic influence of the agent administered. Furthermore, high systemic doses are often required to achieve activity at the wound level. Direct intrawound administration of substances is traumatic and disruptive to the fragile wound environment and increases the risk of infection. We devised a system for continuous atraumatic delivery of substances directly to subcutaneously implanted polyvinyl alcohol sponges, an adaptation of a well-established model of wound healing. Sponge-catheter constructs were fashioned by feeding identical lengths of silicone catheters through two 40-mg sponge disks (on edge). The distal sponge was fixed 0.5 cm from the distal, ligated end of the catheter and centered over two 1-mm holes in the catheter tubing. The proximal sponge was fixed over nonperforated catheter with its edge 2 cm proximal from the close edge of the distal sponge. Each construct was connected to a mini-osmotic pump (infusion rate 1 microl/h) loaded with an appropriate infusate and inserted subcutaneously on the dorsum of anesthetized male Sprague-Dawley rats. Hydroxyproline (OHP) content of sponges, a measure of collagen deposition, was determined at 7 days postwounding. Infusion of India ink confirmed selective delivery to the distal sponge. Saline infusion alone significantly elevated OHP content compared to noninfused sponges (450 +/- 43 vs 328 +/- 36 microg OHP/100 mg sponge, P < 0.05). Infusion of S-methylisothiourea (a selective iNOS inhibitor, 84 microg/sponge/24 h) successfully inhibited NO production (35.9 +/- 3.1 vs 49.6 +/- 3.6 microM, P < 0.05) and decreased sponge OHP content (385 +/- 60 vs 568 +/- 70 microg OHP/100 mg sponge, P < 0.05) without the toxic side effect (i.e., weight loss) seen with systemic administration. Infusion of an adenoviral solution containing mouse iNOS cDNA resulted in successful transduction of wound cells demonstrating the ability to deliver genes to a healing wound model. The data demonstrate that manipulation of wound physiology is possible by local delivery of low doses of wound-active compounds to the wound site. This promises to be a powerful tool for the study of both normal and impaired wound healing.
Asunto(s)
Técnicas Histológicas , Cicatrización de Heridas/fisiología , Adenoviridae/genética , Animales , Hidroxiprolina/antagonistas & inhibidores , Hidroxiprolina/metabolismo , Inyecciones Subcutáneas , Isotiuronio/administración & dosificación , Isotiuronio/análogos & derivados , Isotiuronio/farmacología , Masculino , Nitratos/metabolismo , Óxido Nítrico Sintasa/genética , Óxido Nítrico Sintasa de Tipo II , Nitritos/metabolismo , Alcohol Polivinílico , Poríferos , ARN Mensajero/metabolismo , ARN Viral/metabolismo , Ratas , Ratas Sprague-Dawley , Cloruro de Sodio/administración & dosificación , Cloruro de Sodio/farmacología , Transcripción GenéticaRESUMEN
Following severe trauma and surgical injury, nutritional support via the enteral route has been shown to lead to increased survival and decreased complications when compared to the parenteral route. We hypothesized that the route of nutrient delivery may affect cutaneous wound healing following severe traumatic insult. Forty-six Sprague Dawley rats underwent bilateral closed femoral fractures, central venous catheterization, gastrostomy placement and dorsal skin incision with placement of polyvinyl alcohol sponges into subcutaneous pockets. Identical nutritional infusates of 25% dextrose, 4.25% amino acids, and vitamins were given, half the animals receiving the infusion via the gastrostomy (ENT) and the other half via the venous catheter (TPN). Animals were sacrificed on post-operative days 5, 7, or 10. Wound breaking strength (WBS, g) and sponge granuloma hydroxyproline content (OHP- a measure of wound collagen deposition, microg/ 100mg sponge) were measured. There were no significant nutritional differences between the two feeding groups. On days 5 and 7, WBS was significantly higher in the ENT group (58.0 +/- 3.1 g vs 48.9 +/- 2.6 g, p < 0.05, and 123 +/- 19 g vs 87.6 +/- 4.2 g, p < 0.05 vs TPN respectively). Sponge OHP content on day 5 was significantly higher in the ENT group (101 +/- 3 vs 86.7+/-5.8 microg/100 sponge, p < 0.05). These data demonstrate that the enteral feeding route imparts a benefit to early post-traumatic wound healing s compared to parenteral feeding.
Asunto(s)
Apoyo Nutricional , Cicatrización de Heridas/fisiología , Heridas y Lesiones/terapia , Análisis de Varianza , Animales , Northern Blotting , Intervalos de Confianza , Modelos Animales de Enfermedad , Nutrición Enteral/métodos , Masculino , Nutrición Parenteral/métodos , Periodo Posoperatorio , Probabilidad , Ratas , Ratas Sprague-Dawley , Valores de Referencia , Sensibilidad y Especificidad , Procedimientos Quirúrgicos Operativos , Heridas y Lesiones/fisiopatologíaRESUMEN
BACKGROUND: Although generation of nitric oxide (NO) from inducible nitric oxide synthase (iNOS) has been shown to be required for cutaneous wound healing, no differences have been noted in incisional healing between iNOS knockout (iNOS-KO) and wild type (WT) mice. Because supplemental dietary arginine enhances cutaneous healing in normal rodents and is the sole substrate for NO synthesis, we studied whether arginine can enhance cutaneous wound healing in iNOS-KO mice. METHODS: Twenty iNOS-KO and 20 WT mice, all on a C57BL/6 background, were divided into 4 groups of 10 animals each. Ten animals with each trait were randomized to receive either normal food and tap water or food and water each supplemented with 0.5% arginine (w/w). All animals underwent a 2.5-cm dorsal skin incision with implantation of four 20-mg polyvinyl alcohol sponges into subcutaneous pockets. On postoperative day 14 the animals were killed. The dorsal wound was harvested for breaking strength determination and the wound sponges were assayed for hydroxyproline content and total wound fluid nitrite/nitrate concentration. RESULTS: Dietary arginine supplementation enhanced both wound breaking strength and collagen deposition in WT but not iNOS-KO mice. Wound fluid nitrite/nitrate levels were higher in WT than iNOS-KO animals but were not significantly influenced by additional arginine. CONCLUSIONS: These data demonstrate that supplemental dietary arginine enhances wound healing in normal mice. The loss of a functional iNOS gene abrogates the beneficial effect of arginine in wound healing. This suggests that the metabolism of arginine via the NO pathway is one mechanism by which arginine enhances wound healing.