ABSTRACT
Background: Therapy management in patients suffering from mental health disorders is complex and the risks derived from changes or interruptions of treatment should not be ignored. Medication reconciliation in psychiatry may reduce medication errors and promote patient safety during transitions of care. Objective: To identify the influence of complementary information sources in the construction of the best possible medication history, and to ascertain the potential clinical impact of discrepancies identified in a medication reconciliation service. Methods: An observational study was conducted in an acute mental hospital unit, with a further validation in an internal medicine unit. Adult patients taking at least one medicine admitted in the unit were included. Patients/caregivers were interviewed upon admission and the information gathered was compared with hospital medical and shared electronic medical records. Once the best possible medication history was gathered, therapeutic information was reconciled against the prescription on admission to identify discrepancies. Potential clinical impact of medication errors was classified using the International Safety Classification. Results: During the study period, 148 patients were admitted, 50.7% females, mean age 54.6 years (SD=16.3). Collaboration of a caregiver was a needed in 74% of the interviews. In total, 1,147 drugs were considered to obtain patients' best possible medication history. After reconciliation, 560 clinically sound intentional discrepancies were identified and 359 discrepancies required further clarification from prescribers: 84.12% "drug omission", 5.57% "drug substitution", 6.96% "dose change", and 3.34% "dosage frequency change". Potential clinical impact of these medication discrepancies was classified as: 95 mild, 100 moderate, and 29 severe medication errors. Conclusion: About 1 in three intentional discrepancies observed in a pharmacists-led medication reconciliation service required further clarification from prescribers, being 80% of them unintentional discrepancies. Results highlight the importance of the caregiver as source of information for the psychiatric patient, the relevance of analyzing shared electronic health records until 6 months before, and the need to use hospital medical records efficiently. Additionally, 29 discrepancies were classified as errors with potentially severe clinical impact. A medication reconciliation service is concluded to be feasible and necessary in a mental health unit.
ABSTRACT
Background: Therapy management in patients suffering from mental health disorders is complex and the risks derived from changes or interruptions of treatment should not be ignored. Medication reconciliation in psychiatry may reduce medication errors and promote patient safety during transitions of care. Objective: To identify the influence of complementary information sources in the construction of the best possible medication history, and to ascertain the potential clinical impact of discrepancies identified in a medication reconciliation service. Methods: An observational study was conducted in an acute mental hospital unit, with a further validation in an internal medicine unit. Adult patients taking at least one medicine admitted in the unit were included. Patients/caregivers were interviewed upon admission and the information gathered was compared with hospital medical and shared electronic medical records. Once the best possible medication history was gathered, therapeutic information was reconciled against the prescription on admission to identify discrepancies. Potential clinical impact of medication errors was classified using the International Safety Classification. Results: During the study period, 148 patients were admitted, 50.7% females, mean age 54.6 years (SD=16.3). Collaboration of a caregiver was a needed in 74% of the interviews. In total, 1,147 drugs were considered to obtain patients best possible medication history. After reconciliation, 560 clinically sound intentional discrepancies were identified and 359 discrepancies required further clarification from prescribers: 84.12% drug omission, 5.57% drug substitution, 6.96% dose change, and 3.34% dosage frequency change. Potential clinical impact of these medication discrepancies was classified as: 95 mild, 100 moderate, and 29 severe medication errors. (AU)
Subject(s)
Humans , Male , Female , Young Adult , Adult , Middle Aged , Aged , Aged, 80 and over , Medication Reconciliation , Medication Errors , Mental Health , Hospitals, Psychiatric , Caregivers , PharmacistsABSTRACT
BACKGROUND: Beers Criteria are one of the best known explicit criteria to identify inappropriate medication in elderly that can be used in medication review. The access to patients' medical records may be different among healthcare professionals and settings and, subsequently, the identification of patients' diagnoses may be compromised. OBJECTIVE: To assess the consequences of ignoring patient diagnoses when applying 2015 Beers Criteria to identify potentially inappropriate medication (PIM). SETTING: Three nursing homes in Central Portugal. METHOD: Medical records of nursing home residents over 65 years old were appraised to identify medication profile and medical conditions. 2015 Beers Criteria were used with and without considering patients' diagnoses. To compare the number of PIM and PIM-qualifying criteria complied in these two judgements, Wilcoxon signed-rank tests were performed. MAIN OUTCOME MEASURE: Number of PIMs and number of PIM-qualifying criteria. RESULTS: A total of 185 patients with a mean age of 86.7 years (SD = 7.8) with a majority of female (70.3%) were studied. When assessing the patients with full access to the diagnoses, median number of PIMs was 4 (IQR 0-10) and number of PIM-qualifying criteria was 5 (IQR 0-15). When evaluating only patient current medication, median number of PIMs was 4 (IQR 0-10) and PIM-qualifying criteria was 4 (IQR 0-12). Statistical difference was found in the number of PIM-qualifying criteria identified (p < 0.001), but not in the number of PIMs per patient (p = 0.090). In 171 patients (92.4%) PIMs identified were identical when using or ignoring their medical diagnoses. However, in 80 patients (43.2%) the PIM-qualifying criteria complied were different with and without access to patient diagnoses. CONCLUSION: Although restricted access to patients' diagnoses may limit the judgement of Beers PIM-qualifying criteria, this limitation had no effect on the number of PIM identified.
Subject(s)
Inappropriate Prescribing/prevention & control , Potentially Inappropriate Medication List/standards , Aged , Aged, 80 and over , Cohort Studies , Cross-Sectional Studies , Female , Humans , Inappropriate Prescribing/trends , Male , Polypharmacy , Portugal , Potentially Inappropriate Medication List/trendsABSTRACT
The aim of this study was to determine the correlation between total nitrite/nitrate concentrations (NOx) and the kinetic parameters of monoamine oxidase enzymes (MAO-A and MAO-B) and semicarbazide-sensitive amine oxidase (SSAO) in human mesenteric arteries. Arteries were from non-diabetic and type 2 diabetic patients with sigmoid or rectum carcinoma for whom surgery was the first option and who were not exposed to neo-adjuvant therapy. Segments of human inferior mesenteric arteries from non-diabetic (61.1 ± 8.9 years old, 7 males and 5 females, N = 12) and type 2 diabetic patients (65.8 ± 6.2 years old, 8 males and 4 females, N = 12) were used to determine NOx concentrations and the kinetic parameters of MAO-A, MAO-B and SSAO by the Griess reaction and by radiochemical assay, respectively. The NOx concentrations in arteries from diabetic patients did not differ significantly from those of the non-diabetic group (10.28 ± 4.61 vs 10.71 ± 4.32 nmol/mg protein, respectively). In the non-diabetic group, there was a positive correlation between NOx concentrations and MAO-B parameters: Km (r = 0.612, P = 0.034) and Vmax (r = 0.593, P = 0.042), and a negative correlation with the SSAO parameters: Km (r = -0.625, P = 0.029) and Vmax (r = -0.754, P = 0.005). However, in the diabetic group no correlation was found between NOx concentrations and the three kinetic parameters of the enzymes. These results suggest an important function of sympathetic nerves and vascular NOx concentrations in arteries of non-diabetic patients. Thus, these results confirm the importance of a balance between oxidants and antioxidants in the maintenance of vascular homeostasis to prevent oxidative stress.
Subject(s)
Aged , Female , Humans , Middle Aged , Amine Oxidase (Copper-Containing)/metabolism , /metabolism , Mesenteric Arteries/chemistry , Monoamine Oxidase/metabolism , Nitrates/analysis , Nitrites/analysis , Case-Control Studies , /enzymology , Mesenteric Arteries/enzymology , Rectal Neoplasms/enzymology , Sigmoid Neoplasms/enzymologyABSTRACT
The aim of this study was to determine the correlation between total nitrite/nitrate concentrations (NOx) and the kinetic parameters of monoamine oxidase enzymes (MAO-A and MAO-B) and semicarbazide-sensitive amine oxidase (SSAO) in human mesenteric arteries. Arteries were from non-diabetic and type 2 diabetic patients with sigmoid or rectum carcinoma for whom surgery was the first option and who were not exposed to neo-adjuvant therapy. Segments of human inferior mesenteric arteries from non-diabetic (61.1 ± 8.9 years old, 7 males and 5 females, N = 12) and type 2 diabetic patients (65.8 ± 6.2 years old, 8 males and 4 females, N = 12) were used to determine NOx concentrations and the kinetic parameters of MAO-A, MAO-B and SSAO by the Griess reaction and by radiochemical assay, respectively. The NOx concentrations in arteries from diabetic patients did not differ significantly from those of the non-diabetic group (10.28 ± 4.61 vs 10.71 ± 4.32 nmol/mg protein, respectively). In the non-diabetic group, there was a positive correlation between NOx concentrations and MAO-B parameters: Km (r = 0.612, P = 0.034) and Vmax (r = 0.593, P = 0.042), and a negative correlation with the SSAO parameters: Km (r = -0.625, P = 0.029) and Vmax (r = -0.754, P = 0.005). However, in the diabetic group no correlation was found between NOx concentrations and the three kinetic parameters of the enzymes. These results suggest an important function of sympathetic nerves and vascular NOx concentrations in arteries of non-diabetic patients. Thus, these results confirm the importance of a balance between oxidants and antioxidants in the maintenance of vascular homeostasis to prevent oxidative stress.
Subject(s)
Amine Oxidase (Copper-Containing)/metabolism , Diabetes Mellitus, Type 2/metabolism , Mesenteric Arteries/chemistry , Monoamine Oxidase/metabolism , Nitrates/analysis , Nitrites/analysis , Aged , Case-Control Studies , Diabetes Mellitus, Type 2/enzymology , Female , Humans , Male , Mesenteric Arteries/enzymology , Middle Aged , Rectal Neoplasms/enzymology , Sigmoid Neoplasms/enzymologyABSTRACT
Monoamine oxidase (MAO, type A and B) and semicarbazide-sensitive amine oxidase (SSAO) metabolize biogenic amines, however, the impact of these enzymes in arteries from patients with type 2 diabetes remains poorly understood. We investigated the kinetic parameters of the enzymes to establish putative correlations with noradrenaline (NA) content and patient age in human mesenteric arteries from type 2 diabetic patients. The kinetic parameters were evaluated by radiochemical assay and NA content by high-performance liquid chromatography (HPLC). The activity of MAO-A and SSAO in type 2 diabetic vascular tissues was significantly lower compared to the activity obtained in non-diabetic tissues. In the correlation between MAO-A (K(m)) and NA content, we found a positive correlation for both the diabetic and non-diabetic group, but no correlation was established for patient age. In both groups, MAO-B (V(max)) showed a negative correlation with age. The results show that MAO-A and SSAO activities and NA content of type 2 diabetic tissues are lower compared to the non-diabetic tissues, while MAO-B activity remained unchanged. These remarks suggest that MAO-A and SSAO may play an important role in vascular tissue as well as in the vascular pathophysiology of type 2 diabetes.
Subject(s)
Amine Oxidase (Copper-Containing)/metabolism , Diabetes Mellitus, Type 2/enzymology , Mesenteric Arteries/enzymology , Monoamine Oxidase/metabolism , Aged , Amine Oxidase (Copper-Containing)/analysis , Female , Humans , Kinetics , Male , Middle Aged , Monoamine Oxidase/analysis , Norepinephrine/metabolismABSTRACT
Indomethacin (IM) is a non-steroidal anti-inflammatory drug which inhibits prostaglandin biosynthesis. It is practically insoluble in water and has the capacity to induce gastric injury. Hydroxypropyl-beta-cyclodextrin (HP-beta-CD) is an alkylated derivative of beta-CD with the capacity to form inclusion complexes with suitable molecules. IM is considered to form partial inclusion complexes with HP-beta-CD by enclosure of the p-chlorobenzoic part of the molecule in the cyclodextrin channel, reducing the adverse effects. The aim of this paper is to evaluate the gastric damage induced by the IM inclusion complex prepared by freeze-drying and spray-drying. A total of 135 Wistar rats weighing 224.4 +/- 62.5 g were put into 10 groups. They were allowed free access to water but were maintained fasted for 18 h before the first administration until the end of the experiment. IM acid-form, IM trihydrated-sodium-salt and IM-HP-beta-CD spray and freeze-dried, at normal and toxic doses, were administered through gastric cannula once/day for 3 days. Seventy-two hours after the first administration, the animals were sacrificed and the stomachs collected and prepared for morphological study by using the haematoxylin-eosin technique. Lesion indexes (rated 0/4) were developed and the type of injury was scored according to the severity of damage and the incidence of microscopic evidence of harm. Microscopic assessment demonstrated levels of injury with index one on 10-25%. The type of complexation method had different incidence but the same degree. The results show that IM inclusion complexation protects against gastric injury, reducing the incidence and the maximum degree of severity from 4 to 1, with a better performance of the spray-dried complex.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/toxicity , Gastric Mucosa/drug effects , Indomethacin/toxicity , Stomach Ulcer/pathology , beta-Cyclodextrins/chemistry , 2-Hydroxypropyl-beta-cyclodextrin , Acids , Animals , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Drug Compounding , Drug Evaluation, Preclinical , Female , Freeze Drying , Gastric Mucosa/pathology , Indomethacin/chemistry , Male , Rats , Rats, Wistar , Salts , Solubility , Stomach Ulcer/chemically inducedABSTRACT
The choice of appropriate animal models for the initial in vivo testing of potential anticonvulsant compounds is one of the most important steps in the successful search for new antiepileptic drugs. The purpose of this paper is to describe the most important aspects to take into account when performing the maximal electroshock seizure (MES) test in the routine laboratory screening of new antiepileptics: the conventional and threshold MES test experimental procedures, the factors affecting experimental data (laboratory conditions, administration vehicles and drug formulations, time after drug administration, and stimulus duration and site of stimulation) and the assessment of anticonvulsant activity are discussed.
Subject(s)
Anticonvulsants/pharmacology , Disease Models, Animal , Seizures/drug therapy , Animals , Anticonvulsants/administration & dosage , Drug Evaluation, Preclinical/methods , Electroshock , Humans , Mice , Pharmaceutical Vehicles/chemistry , Rats , Seizures/physiopathology , Time FactorsABSTRACT
OBJECTIVE: The aim of the present study was to evaluate the pharmacokinetic profile of lamotrigine (LTG) in epileptic patients submitted to video-electroencephalography (VEEG) monitoring and, in addition, to investigate the influence of concomitant antiepileptic drugs (AEDs) on the kinetics of LTG. METHODS: The analysis assumed a one-compartment open model with first-order absorption and elimination. The kinetic estimates obtained in this population were validated by using the Prediction-Error approach. The influence of medication was also assessed by the calculation of the LTG concentration-to-dose ratio. Patients (n=135) were divided into four groups according to the co-medication: Group 1, patients taking LTG with enzyme-inducer agents; Group 2, patients receiving LTG with valproic acid; Group 3, patients receiving both inducers and inhibitors of LTG metabolism; Group 4, patients under AEDs not known to alter LTG metabolism. RESULTS: The obtained estimates for clearance (CL) (L/h/kg) [0.075+/-0.029 (Group 1), 0.014+/-0.005 (Group 2), 0.025+/-0.008 (Group 3) and 0.044+/-0.011 (Group 4)] appear to be the most appropriate set to be implemented in clinical practice as prior information, as demonstrated by the accuracy and precision of the measurements. In addition, the influence of co-medication on the LTG profile was further confirmed by the basal LTG concentration-to-dose ratio. CONCLUSION: The results of the present investigation may contribute to achieving the goal of optimizing patients' clinical outcomes by managing their medication regimen through measured drug concentrations. Patients submitted to VEEG monitoring may benefit from this study, as the results may be used to provide better drug management in this medical setting.
Subject(s)
Anticonvulsants/pharmacokinetics , Electroencephalography , Epilepsy/drug therapy , Triazines/pharmacokinetics , Adolescent , Adult , Epilepsy/metabolism , Epilepsy/physiopathology , Female , Half-Life , Humans , Lamotrigine , Male , Middle Aged , Monitoring, Physiologic , Video RecordingABSTRACT
The purpose of this study is to characterize the neuropharmacokinetics of lamotrigine following a single intraperitoneal dose. Adult male Wistar rats were given lamotrigine dose of 5, 10, or 20 mg/kg. Blood and brain samples were obtained at predetermined times over 120 h and analyzed by HPLC. The overall characteristics of plasma curves were determined by noncompartmental analysis with WINNONLIN. The kinetic characterization of lamotrigine distribution between plasma and brain was performed by indirect numerical deconvolution with MULTI(FILT). A linear disposition kinetics was observed within 5-20 mg/kg. The lamotrigine concentrations in brain homogenate were approx. twofold higher than in plasma. The following pharmacokinetic parameters were obtained for lamotrigine 5, 10, and 20 mg/kg, respectively: clearance of distribution from plasma to brain normalized with the volume of the brain, CL/V(h(-1)) = 4.64, 2.47, 2.40; brain-to-plasma partition coefficient, P = 0.40, 0.37, 0.34; first-order transfer rate constant from the brain to the plasma, K(h(-1)) = 11.68, 6.68, 5.96; single-pass mean transit time in the brain, MTT(h) = 0.086, 0.150, 0.168. These results indicate that lamotrigine plasma levels may be good indicators of lamotrigine levels in the brain and that higher response intensities could be expected with higher doses of lamotrigine, since efficacious concentrations are maintained for a longer period.
Subject(s)
Brain/metabolism , Triazines/administration & dosage , Triazines/pharmacokinetics , Animals , Area Under Curve , Biological Availability , Brain/drug effects , Brain/pathology , Chromatography, High Pressure Liquid , Dose-Response Relationship, Drug , Drug Administration Schedule , Injections, Intraperitoneal , Lamotrigine , Male , Rats , Rats, Wistar , Time Factors , Triazines/bloodABSTRACT
The aim of this study was to perform a pharmacokinetic/pharmacodynamic (PK/PD) modelling of lamotrigine following its acute administration to rats. Adult male Wistar rats were given 10 mg/kg of lamotrigine intraperitoneally. Plasma and brain samples were obtained at predetermined times over 120 h post-dose and analysed by liquid chromatography. The anticonvulsant profile against maximal electroshock seizure stimulation was determined over 48 h after dosing. As a linear relationship between lamotrigine plasma and brain profiles was observed, only the plasma data set was used to establish the PK/PD relationship. To fit the effect-time course of lamotrigine, the PK/PD simultaneous fitting link model was used: the pharmacokinetic parameters and dosing information were used in the one-compartment first-order model to predict concentrations, which were then used to model the pharmacodynamic data with the sigmoid Emax model, in order to estimate all the parameters simultaneously. The following parameters were obtained: Vd = 2.00 L/kg, k(abs) = 8.50 h(-1), k(el) = 0.025 h(-1), k(e0) = 3.75 h(-1), Emax = 100.0% (fixed), EC50 = 3.44 mg/L and gamma = 8.64. From these results, it can be stated that lamotrigine is extensively distributed through the body, its plasma elimination half-life is around 28 h and a lamotrigine plasma concentration of 3.44 mg/L is enough to protect 50% of the animals. When compared with humans, the plasma concentrations achieved with this dose were within the therapeutic concentration range that had been proposed for epileptic patients. With the present PK/PD modelling it was possible to fit simultaneously the time-courses of the plasma levels and the anticonvulsant effect of lamotrigine, providing information not only about the pharmacokinetics of lamotrigine in the rat but also about its anticonvulsant response over time. As this approach can be easily applied to other drugs, it becomes a useful tool for an explanatory comparison between lamotrigine and other antiepileptic drugs.
Subject(s)
Anticonvulsants/pharmacology , Anticonvulsants/pharmacokinetics , Triazines/pharmacology , Triazines/pharmacokinetics , Animals , Anticonvulsants/administration & dosage , Injections, Intraperitoneal , Lamotrigine , Male , Rats , Rats, Wistar , Triazines/administration & dosageABSTRACT
OBJECTIVE: The aim of the present study was to investigate the usefulness of the CA-125 area under the curve (AUC) as a new kinetic parameter for predicting overall survival in patients with ovarian cancer. In addition, the relationship of CA-125 AUC with other prognostic factors of ovarian cancer was evaluated. METHODS: Ninety-two patients that underwent primary line chemotherapy within 4 months after submission to cytoreductive surgery were included. For each patient, CA-125 AUC was calculated and a statistical analysis was conducted to compare CA-125 AUC behavior among patients according to several covariates. RESULTS: The mean age at diagnostic time was found to be 55.5 (16.1-82.4) years with a mean survival of 39.2 (3.5-100.1; SE = 2.6) months. Across FIGO stage I, II, III, and IV patients had a mean CA-125 AUC of 18.2, 24.6, 147.8, and 574.6 IU/ml*days, respectively (P < 0.05). At the evaluation date, living patients had a mean CA-125 AUC of 40.1 in contrast to 234.1 IU/ml*days (P < 0.05) for deceased ones. Patients with a complete response to primary chemotherapy had a mean CA-125 AUC of 48.8, while patients with a partial response had a mean of 251.7 IU/ml*days, and patients with no response or disease progression had a mean of 316.5 IU/ml*days (P < 0.05). The best CA-125 AUC performance is in predicting patient complete response to chemotherapy with a cut-off of 100 IU/ml*days and an accuracy of 82%. CONCLUSIONS: Despite CA-125 AUC high correlation with the FIGO stage, residual disease, and patient final outcome, the main interest of CA-125 AUC calculation is to evaluate the treatment efficacy and to foresee a full chemotherapy response. Further studies should be carried out before extrapolating these results to other data sets.
Subject(s)
CA-125 Antigen/blood , Ovarian Neoplasms/blood , Adolescent , Adult , Aged , Aged, 80 and over , Area Under Curve , Combined Modality Therapy , Female , Humans , Middle Aged , Neoplasm Staging , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/pathology , Ovarian Neoplasms/surgery , Prognosis , Retrospective StudiesABSTRACT
As it has been previously shown that lamotrigine (LTG) accumulates in the kidney of male rats, the purpose of the present investigation was to characterize the kidney profiles of LTG and its kidney distribution pattern in male rats, in order to confirm if a preferential distribution exists and to analyse if it does or does not affect the LTG systemic pharmacokinetics. Adult male Wistar rats were intraperitoneally injected with 5, 10 and 20 mg/kg of LTG. The concentration-time profiles of LTG in plasma and whole kidney were determined over 120 h postdose. The distribution of LTG in the rat kidney was investigated in another group of rats by measuring LTG levels in the renal cortex and medulla. The LTG plasma concentration-time profiles revealed a linear relationship with dose. However, a slight increase in the LTG elimination half-life with dose was observed. In contrast, a nonlinear relationship was established between LTG kidney levels and the dose administered. Consequently, nonparallel patterns were observed between LTG plasma and kidney profiles. The LTG kidney distribution pattern revealed an accumulation of LTG in the renal cortex. The present study demonstrated that LTG distributes preferentially to the kidneys of the male rat in a dose-dependent manner and suggests that such distribution may slightly affect the systemic kinetics of the drug.
Subject(s)
Anticonvulsants/pharmacokinetics , Kidney/metabolism , Triazines/pharmacokinetics , Animals , Anticonvulsants/administration & dosage , Anticonvulsants/blood , Area Under Curve , Chromatography, High Pressure Liquid , Half-Life , Injections, Intraperitoneal , Kidney Cortex/metabolism , Kidney Medulla/metabolism , Lamotrigine , Male , Rats , Rats, Wistar , Triazines/administration & dosage , Triazines/bloodABSTRACT
The pro-inflammatory cytokine interleukin-1beta (IL-1) induces articular chondrocytes to produce reactive oxygen species (ROS), including hydrogen peroxide (H2O2), which mediate some IL-1-induced responses. This study aimed at elucidating the role of ROS, particularly H2O2, in mediating IL-1-induced activation of the transcription factor activator protein-1 (AP-1) in primary cultures of articular chondrocytes. AP-1 may function either as an inducer or as a repressor of the inducible nitric oxide synthase (iNOS) gene promoter. Since we observed that AP-1 is not required for iNOS expression in chondrocytes, we also investigated whether it is a repressor of this gene. The results of electrophoretic mobility shift assays showed that both IL-1 and H2O2 activated AP-1 and that inhibition of IL-1-induced ROS production abrogated AP-1 activation. The AP-1 complexes, induced by either IL-1 or H2O2, contained c-Fos/c-Jun and c-Fos/JunD heterodimers, but IL-1 activated AP-1 with a kinetics slower than that observed with H2O2. Pre-activation of AP-1, before stimulation of the cells with IL-1, did not inhibit iNOS mRNA and protein synthesis, relative to cells treated with IL-1 alone. These results indicate that H2O2 is a major mediator of IL-1-induced AP-1 activation in articular chondrocytes and that inhibition of ROS production is an effective strategy to block this IL-1-induced response. This study also identifies c-Fos/c-Jun and c-Fos/JunD heterodimers as the AP-1 transcription factors induced by IL-1, which, although not involved in the transcriptional regulation of the iNOS gene, may be important for the regulation of other genes also relevant in arthritic diseases, namely the collagenase-1 and IL-8 genes.
Subject(s)
Chondrocytes/enzymology , Hydrogen Peroxide/pharmacology , Interleukin-1/metabolism , Nitric Oxide Synthase/biosynthesis , Transcription Factor AP-1/metabolism , Animals , Blotting, Northern , Blotting, Western , Cattle , Cell Nucleus/metabolism , Cells, Cultured , Chondrocytes/metabolism , Collagenases/biosynthesis , Cytoplasm/metabolism , Dimerization , Dose-Response Relationship, Drug , Enzyme Activation , Inflammation , Interleukin-8/biosynthesis , Kinetics , Nitric Oxide Synthase Type II , Promoter Regions, Genetic , Proto-Oncogene Proteins c-fos/metabolism , Proto-Oncogene Proteins c-jun/metabolism , RNA, Messenger/metabolism , Reactive Oxygen Species , Recombinant Proteins/metabolismABSTRACT
Our previous studies showed that reactive oxygen species (ROS) are required for the pro-inflammatory cytokine interleukin-1 beta (IL-1) to induce the activity of the Nuclear transcription Factor-kappa B (NF-kappa B) and the expression of the inducible isoform of the nitric oxide synthase (iNOS) in bovine articular chondrocytes. This study aimed at elucidating the role of hydrogen peroxide (H(2)O(2)) and the superoxide radical, two major ROS, in mediating those IL-1-induced responses. The results obtained show that chondrocytes produce both H(2)O(2) and superoxide radical in response to IL-1. Treatment of the chondrocyte cultures with H(2)O(2) alone did not induce NF-kappa B activation or iNOS expression. Addition of H(2)O(2) simultaneously with IL-1 did neither enhance nor inhibit NF-kappa B activation and iNOS expression, relatively to treatment with IL-1 alone. Accordingly, treatment with catalase did not inhibit those IL-1-induced responses. Treatment with superoxide dismutase, however, effectively prevented IL-1-induced I kappa B-alpha degradation and iNOS expression. Taken together, the results obtained indicate that superoxide mediates IL-1-induced I kappa B-alpha degradation and the consequent NF-kappa B activation and iNOS expression in chondrocytes, whereas H(2)O(2) does not seem to participate in those IL-1-induced responses. In conclusion, the present study identifies the superoxide radical as the ROS involved in mediating the IL-1-induced signaling pathway that leads to NF-kappa B activation and to the expression of NF-kappa B-dependent genes in bovine articular chondrocytes.
Subject(s)
Chondrocytes/drug effects , Hydrogen Peroxide/metabolism , Interleukin-1/metabolism , NF-kappa B/metabolism , Superoxides/metabolism , Animals , Cartilage, Articular , Catalase/pharmacology , Cattle , Cells, Cultured , Chondrocytes/metabolism , Hydrogen Peroxide/pharmacology , Interleukin-1/pharmacology , NF-kappa B/analysis , NF-kappa B/genetics , Nitric Oxide Synthase/biosynthesis , Nitric Oxide Synthase Type II , Signal Transduction , Superoxide Dismutase/pharmacology , Superoxides/pharmacologyABSTRACT
OBJECTIVE AND DESIGN: Determine the sources of nitric oxide (NO) and evaluate its role in the activation of nuclear Factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) and in the expression of NO synthase II (NOS II), induced by interleukin-1beta (IL-1). MATERIAL OR SUBJECTS: Primary cultures of bovine articular chondrocytes. TREATMENT: The cells were treated with IL-1, 5 ng/ml with or without the NO donor S-nitroso-N-acetylpenicillamine (SNAP), in concentrations ranging from 10 to 300 microM. METHODS: NF-kappaB and AP-1 activation were evaluated by electrophoretic mobility shift assay. Northern blot was used to detect NOS II mRNA levels and western blot to evaluate IkappaB-alpha, NOS I and NOS II protein levels. RESULTS: Under basal conditions, chondrocytes expressed NOS I, which was lost upon IL-I treatment. SNAP inhibited IL-I-induced NF-kappaB activation and NOS II expression. When added alone, SNAP induced AP-1 activation to approximately the same extent as IL-I. CONCLUSIONS: These results suggest that, in chondrocytes, NO is a key regulator of the signaling pathways leading from IL-I to NF-kappaB and AP-1 activation and to the expression of genes that are involved in the pathophysiology of arthritic diseases.
Subject(s)
Cartilage, Articular/metabolism , Chondrocytes/metabolism , NF-kappa B/biosynthesis , Nitric Oxide Synthase/biosynthesis , Nitric Oxide/physiology , Transcription Factor AP-1/biosynthesis , Animals , Blotting, Northern , Blotting, Western , Cartilage, Articular/cytology , Cattle , Cell Nucleus/enzymology , Cell Nucleus/metabolism , Cells, Cultured , Cytoplasm/enzymology , Cytoplasm/metabolism , Dose-Response Relationship, Drug , Electrophoretic Mobility Shift Assay , In Vitro Techniques , Interleukin-1/pharmacology , Nitric Oxide Donors/pharmacology , Nitric Oxide Synthase Type II , S-Nitroso-N-Acetylpenicillamine/pharmacology , Signal Transduction/drug effectsABSTRACT
Nitric oxide (NO), produced by the inducible isoform of the NO synthase (iNOS), plays an important role in the pathophysiology of arthritic diseases. This work aimed at elucidating the role of the mitogen-activated protein kinases (MAPK), p38MAPK and p42/44MAPK, and of protein tyrosine kinases (PTK) on interleukin-1beta (IL-1)-induced iNOS expression in bovine articular chondrocytes. The specific inhibitor of the p38MAPK, SB 203580, effectively inhibited IL-1-induced iNOS mRNA and protein synthesis, as well as NO production, while the specific inhibitor of the p42/44MAPK, PD 98059, had no effect. These responses to IL-1 were also inhibited by treatment of the cells with the tyrosine kinase inhibitors, genistein and tyrphostin B42, which also prevented IL-1-induced NF-kappaB activation. The p38MAPK inhibitor, SB 203580, had no effect on IL-1-induced NF-kappaB activation. Finally, the p42/44MAPK inhibitor, PD 98059, prevented IL-1-induced AP-1 activation in a concentration that did not inhibit iNOS expression. In conclusion, this study shows that (1) PTK are part of the signaling pathway that leads to IL-1-induced NF-kappaB activation and iNOS expression; (2) the p38MAPK cascade is required for IL-1-induced iNOS expression; (3) the p42/44MAPK and AP-1 are not involved in IL-1-induced iNOS expression; and (4) NF-kappaB and the p38MAPK lie on two distinct pathways that seem to be independently required for IL-1-induced iNOS expression. Hence, inhibition of any of these two signaling cascades is sufficient to prevent iNOS expression and the subsequent production of NO in articular chondrocytes.
Subject(s)
Chondrocytes/metabolism , Interleukin-1/pharmacology , Mitogen-Activated Protein Kinases/physiology , NF-kappa B/metabolism , Nitric Oxide Synthase/metabolism , Protein-Tyrosine Kinases/physiology , Animals , Cartilage, Articular/cytology , Cattle , Chondrocytes/enzymology , Enzyme Inhibitors/pharmacology , Genistein/pharmacology , Humans , Imidazoles/pharmacology , Mitogen-Activated Protein Kinase 1/antagonists & inhibitors , Mitogen-Activated Protein Kinase 1/physiology , Mitogen-Activated Protein Kinase 3 , Mitogen-Activated Protein Kinases/antagonists & inhibitors , NF-kappa B/drug effects , Nitric Oxide/biosynthesis , Nitric Oxide Synthase/drug effects , Nitric Oxide Synthase Type II , Protein-Tyrosine Kinases/antagonists & inhibitors , Pyridines/pharmacology , Signal Transduction , Transcription Factor AP-1/drug effects , Transcription Factor AP-1/metabolism , Tyrphostins/pharmacology , p38 Mitogen-Activated Protein KinasesABSTRACT
Given that administration vehicles and drug formulations can affect drug bioavailability, their influence on the pharmacokinetic profile of lamotrigine (LTG), a new-generation anti-epileptic drug, was studied in rats. Three different formulations administered intraperitoneally at a dose of 10 mg/kg were used: (1) LTG suspended in a 0.25% methylcelulose solution, (2) LTG dissolved in a 50% propylene glycol solution, and (3) LTG isethionate dissolved in distilled water. Plasma and brain homogenate levels were determined in order to evaluate vehicle-dependent drug absorption. The results demonstrated rapid absorption of LTG when it was administered as an aqueous solution, in contrast to a slower and more erratic absorption after the injection of either the lipophilic solution or the suspension. A plasma peak was achieved 15 min post-dose with the aqueous solution, with a brain peak being achieved 15 min later, while with the other formulations both plasma and brain homogenate peaks were reached 2 h after LTG administration. This study suggests that LTG isethionate dissolved in distilled water is the most suitable formulation for successful LTG pharmacokinetic studies in rats.
Subject(s)
Anticonvulsants/pharmacokinetics , Triazines/pharmacokinetics , Animals , Anticonvulsants/administration & dosage , Anticonvulsants/blood , Area Under Curve , Biological Availability , Brain/metabolism , Chemistry, Pharmaceutical , Chromatography, High Pressure Liquid , Injections, Intraperitoneal , Lamotrigine , Male , Pharmaceutical Vehicles , Rats , Rats, Wistar , Solutions , Suspensions , Triazines/administration & dosage , Triazines/blood , WaterABSTRACT
AIMS: In this work, we studied the mechanisms by which diphenyleneiodonium chloride (DPI) inhibits nitric oxide (NO) synthesis induced by the proinflammatory cytokine interleukin-1beta (IL-1) in bovine articular chondrocytes. To achieve this, we evaluated the ability of DPI to inhibit the expression and activity of the inducible isoform of the NO synthase (iNOS) induced by IL-1. We also studied the ability of DPI to prevent IL-1-induced NF-kappaB activation and reactive oxygen species (ROS) production. RESULTS: Northern and Western blot analysis, respectively, showed that DPI dose-dependently inhibited IL-1-induced iNOS mRNA and protein synthesis in primary cultures of bovine articular chondrocytes. DPI effectively inhibited NO production (IC50=0.03+/-0.004 microM), as evaluated by the method of Griess. Nuclear factor-kappa B (NF-kappaB) activation, as evaluated by electrophoretic mobility shift assay, was inhibited by DPI (1-10 microM) in a dose-dependent manner. IL-1-induced ROS production, as evaluated by measurement of dichlorofluorescein fluorescence, was inhibited by DPI at concentrations that also prevented NF-kappaB activation and iNOS expression. CONCLUSIONS: DPI inhibits IL-1-induced NO production in chondrocytes by two distinct mechanisms: (i) by inhibiting NOS activity, and (ii) by preventing iNOS expression through the blockade of NF-kappaB activation. These results also support the involvement of reactive oxygen species in IL-1-induced NF-kappaB activation and expression of NF-kappaB-dependent genes, such as iNOS.
Subject(s)
Chondrocytes/drug effects , Interleukin-1/metabolism , NF-kappa B/metabolism , Nitric Oxide Synthase/metabolism , Onium Compounds/pharmacology , Reactive Oxygen Species/metabolism , Animals , Cattle , Cells, Cultured , Chondrocytes/metabolism , Dose-Response Relationship, Drug , Enzyme Inhibitors/pharmacology , Nitric Oxide Synthase/antagonists & inhibitors , Nitric Oxide Synthase/genetics , Nitric Oxide Synthase Type IIABSTRACT
A reversed-phase high-performance liquid chromatography assay was developed and validated to determine plasma and brain lamotrigine concentrations allowing pharmacokinetic-pharmacodynamic studies of this new antiepileptic drug in patients and laboratory animals. Lamotrigine and its internal standard were extracted, under alkaline conditions, from plasma and brain homogenate, into ethyl acetate; brain proteins were previously precipitated with trichloroacetic acid. The method was linear between 0.1 and 15.0 mg/l for plasma, with a detection limit of 0.008 mg/l, and between 0.1 and 5.0 mg/l for brain homogenate, with a detection limit of 0.023 mg/l. The method proved to be simple, useful and appropriate, not only for clinical and experimental research, but also for routine monitoring of lamotrigine concentrations in patients.