Male reproductive health under threat: Short term exposure to radiofrequency radiations emitted by common mobile jammers.

BACKGROUND: Modern life prompted man to increasingly generate, transmit and use electricity that leads to exposure to different levels of electromagnetic fields (EMFs). Substantial evidence indicates that exposure to common sources of EMF such as mobile phones, laptops or wireless internet-connected laptops decreases human semen quality. In some countries, mobile jammers are occasionally used in offices, shrines, conference rooms and cinemas to block the signal. AIMS: To the best of our knowledge, this is the first study to investigate the effect of short term exposure of human sperm samples to radiofrequency (RF) radiations emitted by common mobile jammers. SUBJECTS AND METHODS: Fresh semen samples were collected by masturbation from 30 healthy donors who had referred to Infertility Treatment Center at the Mother and Child Hospital with their wives. Female problem was diagnosed as the reason for infertility in these couples. STATISTICAL ANALYSIS: T-test and analysis of variance were used to show statistical significance. RESULTS: The motility of sperm samples exposed to jammer RF radiation for 2 or 4 h were significantly lower than those of sham-exposed samples. These findings lead us to the conclusion that mobile jammers may significantly decrease sperm motility and the couples' chances of conception. CONCLUSION: Based on these results, it can be suggested that in countries that have not banned mobile jammer use, legislations should be urgently passed to restrict the use of these signal blocking devices in public or private places.

Randomized phase II trial of gemcitabine-cisplatin with or without trastuzumab in HER2-positive non-small-cell lung cancer.

BACKGROUND: Trastuzumab provides significant clinical benefits in HER2-positive metastatic breast cancer patients when administered in combination with chemotherapy. Chemotherapy has also been shown to be beneficial in some patients with advanced non-small-cell lung cancer (NSCLC). The present randomized phase II trial examined the effect of adding trastuzumab to a standard chemotherapeutic combination (gemcitabine-cisplatin) in patients with HER2-positive NSCLC. PATIENTS AND METHODS: Patients with untreated stage IIIB/IV HER2-positive NSCLC received up to six 21-day cycles of gemcitabine 1250 mg/m(2) (days 1 and 8) and cisplatin 75 mg/m(2) (day 1). Patients in the trastuzumab arm received trastuzumab 4 mg/kg intravenously (i.v.) followed by 2 mg/kg/week i.v. until progression. RESULTS: Of 619 patients screened, 103 were eligible. Fifty-one patients were treated with trastuzumab plus gemcitabine-cisplatin and 50 with gemcitabine-cisplatin alone. Efficacy was similar in the trastuzumab and control arms: response rate 36% versus 41%; median time to progression 6.3 versus 7.2 months; and median progression-free survival (PFS) 6.1 versus 7 months. Response rate (83%) and median PFS (8.5 months) appeared relatively good in the six trastuzumab-treated patients with HER2 3+ or fluorescence in situ hybridization (FISH)-positive NSCLC. Addition of trastuzumab to gemcitabine-cisplatin was well tolerated, side-effects were as expected, and trastuzumab did not exacerbate the known toxicity of gemcitabine and cisplatin. Symptomatic cardiotoxicity was observed in one trastuzumab-treated patient. Serum trastuzumab concentrations in the presence of gemcitabine-cisplatin were comparable to those of trastuzumab alone. CONCLUSIONS: Trastuzumab plus gemcitabine-cisplatin is well tolerated. Clinical benefit was not observed. Although HER2 3+/FISH-positive patients may benefit from trastuzumab, the subgroup is too small to provide definitive information. No significant effect of gemcitabine-cisplatin on trastuzumab pharmacokinetics was observed.

No evidence that panic attacks are associated with the white coat effect in hypertension.

The authors aimed to determine whether hypertensive patients with panic attacks or panic disorder have a larger white coat effect (difference between clinic blood pressure measured under standard conditions and mean daytime ambulatory blood pressure) than hypertensive patients without panic attacks. White coat effect was compared in a hospital hypertension clinic between 24 patients with panic attacks in the previous 6 months (12 with panic disorder) and 23 hypertensive controls. There were no significant differences between cases and controls in clinic blood pressure, mean daytime ambulatory blood pressure, or white coat effect (18/3 vs. 19/6 mm Hg; difference for systolic, -1.9 mm Hg; 95% confidence interval, -15.8 to +12.0; difference for diastolic, -3.0 mm Hg; 95% confidence interval, -10.2 to +4.3). Comparing only patients with panic disorder with controls, there were again no significant differences in clinic blood pressure, mean daytime ambulatory blood pressure, or white coat effect. This study provides no evidence for an exaggerated white coat effect in hypertensive patients who have experienced panic attacks or panic disorder. However, only larger studies could exclude differences in white coat effect <12/4 mm Hg, or an exaggerated white coat effect in a minority of patients with panic attacks.

Pharmacokinetics and safety of voriconazole following intravenous- to oral-dose escalation regimens.

In this study, the safety, tolerability, and pharmacokinetics of intravenous (i.v.)- to oral-dose regimens of voriconazole were evaluated with a group of 42 healthy men, 41 of whom completed the study. Two cohorts of subjects participated in the study. Cohort 1 (n = 28) took part in two study periods, each consisting of 14 days separated by a minimum 7-day washout. In one of the periods, 14 subjects received 6 mg/kg i.v. twice a day (b.i.d.) on day 1 followed by 3 mg/kg i.v. b.i.d. on days 2 to 7 and were then switched to 200 mg orally b.i.d. for days 8 to 14. In the other period, subjects received 6 mg/kg i.v. b.i.d. on day 1 followed by 5 mg/kg i.v. b.i.d. on days 2 to 7 and were then switched to 400 mg orally b.i.d. for days 8 to 14. The remaining 14 subjects in cohort 1 received a matching placebo throughout the study. In cohort 2 (n = 14), 7 subjects received 6 mg/kg i.v. b.i.d. on day 1 followed by 4 mg/kg i.v. b.i.d. on days 2 to 7 and were then switched to 300 mg orally b.i.d. for days 8 to 14. The remaining seven subjects in cohort 2 received a matching placebo. Blood samples were taken prior to dosing on days 1 to 6 and on days 8 to 13. Blood samples were drawn prior to dosing and at frequent intervals up to 12 h following the morning dose on days 7 and 14 of each study period. The samples were assayed for voriconazole by a high-performance liquid chromatography method. The maximum concentration in plasma (C(max)) occurred at the end of the 1-h i.v. infusion and between 1.4 and 1.8 h after oral administration. Voriconazole exhibited nonlinear pharmacokinetics, possibly due to saturable metabolism. For cohort 1, both C(max) and the area under the concentration-time curve within a dosage interval (AUC(tau)) increased disproportionately with dose for both i.v. and oral dosing. For i.v. dosing, a 1.7-fold increase in dose resulted in 2.4- and 3.1-fold increases in C(max) and AUC(tau), respectively. Similarly, a 2-fold increase in oral dosing resulted in 2.8- and 3.9-fold increases in C(max) and AUC(tau), respectively. The mean values for C(max) observed following oral dosing were lower than those obtained after i.v. administration, ranging from 62.7 to 89.6% of the i.v. value. After the switch from i.v. to oral dosing, most subjects achieved steady state by day 4, and mean minimum concentrations in plasma remained above clinically important MICs. The pharmacokinetic profiles for saliva followed a pattern similar to those observed for plasma; there was a highly significant correlation between plasma and saliva voriconazole concentrations (P < 0.0001). Voriconazole was well tolerated; the most commonly reported adverse events in voriconazole-treated subjects were mild to moderate headache, rash, and abnormal vision. Visual function tests detected no further abnormalities during voriconazole treatment.

Is coronary risk an accurate surrogate for cardiovascular risk for treatment decisions in mild hypertension? A population validation.

OBJECTIVE: To examine the relationship between coronary (CHD) and cardiovascular (CVD) risk in patients with uncomplicated mild hypertension and to determine the accuracy of using CHD risk > or = 15% over 10 years to identify for antihypertensive treatment those patients with CVD risk > or = 20% over 10 years as advised in recent British guidelines. DESIGN: Comparison of decisions made using CHD risk > or = 15% over 10 years calculated by the Framingham risk function and estimated using a simple table with CVD risk > or = 20% over 10 years. SETTING: British population. SUBJECTS: People aged 35-64 years with uncomplicated mild systolic hypertension (SBP 140-159 mmHg, n = 624) from the 1995 Scottish Health Survey. MAIN OUTCOME MEASURES: Relationship between CHD and CVD risk. Sensitivity, specificity, positive and negative predictive values (PPV and NPV). RESULTS: CHD risk 15% over 10 years was equivalent to CVD risk 21% over 10 years. Exact CHD risk > or = 15% over 10 years had sensitivity 79%, specificity 98%, PPV 94% and NPV 93% in detecting CVD risk > or = 20% over 10 years. Use of the table to estimate CHD risk > or = 15% over 10 years gave sensitivity 88%, specificity 90%, PPV 76% and NPV 95%. CONCLUSION: CHD risk appears acceptably accurate for targeting treatment in mild hypertension. The risk assessment table, which slightly overestimates CHD risk, was more sensitive in identifying patients with CVD risk > or = 20% over 10 years and may be preferable to using exact CHD risk. European guidelines which suggest targeting treatment for mild hypertension at CHD risk > or = 20% over 10 years are over-conservative compared with British guidelines.

Aspirin for primary prevention of coronary heart disease: safety and absolute benefit related to coronary risk derived from meta-analysis of randomised trials.

OBJECTIVE: To determine the cardiovascular and coronary risk thresholds at which aspirin for primary prevention of coronary heart disease is safe and worthwhile. DESIGN: Meta-analysis of four randomised controlled trials of aspirin for primary prevention. The benefit and harm from aspirin treatment were examined to determine: (1) the cardiovascular and coronary risk threshold at which benefit in prevention of myocardial infarction exceeds harm from significant bleeding; and (2) the absolute benefit expressed as number needed to treat (NNT) for aspirin net of cerebral haemorrhage and other bleeding complications at different levels of coronary risk. MAIN OUTCOME MEASURES: Benefit from aspirin, expressed as reduction in cardiovascular events, myocardial infarctions, strokes, and total mortality; harm caused by aspirin in relation to significant bleeds and major haemorrhages. RESULTS: Aspirin for primary prevention significantly reduced all cardiovascular events by 15% (95% confidence interval (CI) 6% to 22%) and myocardial infarctions by 30% (95% CI 21% to 38%), and non-significantly reduced all deaths by 6% (95% CI -4% to 15%). Aspirin non-significantly increased strokes by 6% (95% CI -24% to 9%) and significantly increased bleeding complications by 69% (95% CI 38% to 107%). The risk of major bleeding balanced the reduction in cardiovascular events when cardiovascular event risk was 0.22%/year. The upper 95% CI for this estimate suggests that harm from aspirin is unlikely to outweigh benefit provided the cardiovascular event risk is 0.8%/year, equivalent to a coronary risk of 0.6%/year. At coronary event risk 1.5%/year, the five year NNT was 44 to prevent a myocardial infarction, and 77 to prevent a myocardial infarction net of any important bleeding complication. At coronary event risk 1%/year the NNT was 67 to prevent a myocardial infarction, and 182 to prevent a myocardial infarction net of important bleeding. CONCLUSIONS: Aspirin treatment for primary prevention is safe and worthwhile at coronary event risk >/= 1.5%/year; safe but of limited value at coronary risk 1%/year; and unsafe at coronary event risk 0.5%/year. Advice on aspirin for primary prevention requires formal accurate estimation of absolute coronary event risk.

Influence of genetic polymorphisms in the beta2-adrenoceptor on desensitization in human lung mast cells.

The beta-adrenoceptor agonist, isoprenaline, inhibited the immunoglobulin E-mediated release of histamine from human lung mast cells (HLMC). Long-term (24 h) exposure of HLMC to isoprenaline reduced the subsequent effectiveness of isoprenaline to inhibit histamine release. The extent of this functional desensitization was variable with some HLMC preparations resistant and others highly susceptible. We sought to determine whether the variability in the degree of functional desensitization was influenced by genetic polymorphisms in the beta2-adrenoceptor. HLMC preparations were genotyped at two polymorphic loci, positions 16 (arg to gly) and 27 (gln to glu), and the effect of desensitizing conditions (24 h with 10(-6) M isoprenaline) on the subsequent ability of isoprenaline (10(-7) M) to inhibit histamine release from HLMC was determined (n = 72). In HLMC preparations expressing beta2-adrenoceptors with arg (wild-type) or gly (mutant) at position 16, desensitization was 71 +/- 5% (n = 18) or 43 +/- 5%, (n = 26), respectively, whereas the desensitization was 59 +/- 6% (n = 28) for heterozygotes at this position. In HLMC preparations expressing beta2-adrenoceptors with gln (wild-type) or glu (mutant) at position 27, desensitization was 65 +/- 5% (n = 25) or 28 +/- 7% (n = 17), respectively, whereas the desensitization was 61 +/- 5% (n = 30) for heterozygotes at this position. These data suggest that mutant (gly16 and glu27) forms of the receptor are resistant to desensitization compared to wild-type (arg16 and gln27) forms. However, analyses to determine the relative contributions of positions 16 and 27 suggest that position 27 is more important in influencing the degree of functional desensitization.

Coronary and cardiovascular risk estimation for primary prevention: validation of a new Sheffield table in the 1995 Scottish health survey population.

OBJECTIVE: To examine the accuracy of a new version of the Sheffield table designed to aid decisions on lipids screening and detect thresholds for risk of coronary heart disease needed to implement current guidelines for primary prevention of cardiovascular disease. DESIGN: Comparison of decisions made on the basis of the table with absolute risk of coronary heart disease or cardiovascular disease calculated by the Framingham risk function. The decisions related to statin treatment when coronary risk is >/=30% over 10 years; aspirin treatment when the risk is >/=15% over 10 years; and the treatment of mild hypertension when the cardiovascular risk is >/=20% over 10 years. SETTING: The table is designed for use in general practice. SUBJECTS: Random sample of 1000 people aged 35-64 years from the 1995 Scottish health survey. MAIN OUTCOME MEASURES: Sensitivity, specificity, and positive and negative predictive values of the table. RESULTS: 13% of people had a coronary risk of >/=15%, and 2. 2% a risk of >/=30%, over 10 years. 22% had mild hypertension (systolic blood pressure 140-159 mm Hg). The table indicated lipids screening for everyone with a coronary risk of >/=15% over 10 years, for 95% of people with a ratio of total cholesterol to high density lipoprotein cholesterol of >/=8.0, but for <50% with a coronary risk of <5% over 10 years. Sensitivity and specificity were 97% and 95% respectively for a coronary risk of >/=15% over 10 years; 82% and 99% for a coronary risk of >/=30% over 10 years; and 88% and 90% for a cardiovascular risk of >/=20% over 10 years in mild hypertension. CONCLUSION: The table identifies all high risk people for lipids screening, reduces screening of low risk people by more than half, and ensures that treatments are prescribed appropriately to those at high risk, while avoiding inappropriate treatment of people at low risk.

Association of panic disorder and panic attacks with hypertension.

PURPOSE: Previous studies of the association between hypertension and panic disorder were uncontrolled or involved small numbers of patients. PATIENTS AND METHODS: We compared the prevalence of panic disorder and panic attacks in 351 patients with documented hypertension who were randomly selected from all hypertensive patients registered in one primary care practice with age- and gender-matched normotensive patients from the same practice and with hypertensive patients attending a hospital clinic. All three groups completed questionnaires for panic disorder based on standard criteria, as well as the Hospital Anxiety and Depression scale. RESULTS: The prevalence of current (previous 6 months) panic attacks was significantly greater in primary care patients with hypertension (17%, P <0.05) and hospital-based hypertensive patients (19%, P <0.01) than in normotensive patients (11%). Similar results were seen for lifetime panic attacks (35% versus 39% versus 22%; both P for comparisons with normotensive patients <0.001). The prevalence of panic disorder was significantly greater in primary care patients with hypertension (13%) than normotensive patients (8%, P <0.05). Anxiety scores were significantly higher in both hypertensive groups than in normotensive patients. Depression scores were significantly higher in hospital-based hypertensive patients than in the other two groups. The reported diagnosis of hypertension antedated the onset of panic attacks in a large majority of patients (P <0.01). CONCLUSIONS: Physicians caring for patients with hypertension should be aware of the significantly greater prevalence of panic attacks in these patients.

Influence of receptor reserve on beta-adrenoceptor-mediated responses in human lung mast cells.

1. The effects of the beta-adrenoceptor agonists isoprenaline and salbutamol on IgE-mediated histamine release from human lung mast cells (HLMC) were evaluated. Both agonists (10(-10)-10(-5) M) inhibited histamine release in a dose-dependent manner and isoprenaline (pD2, 8.3+/-0.1, mean+/-s.e.mean) was more potent than salbutamol (7.3+/-0.1). Moreover, the mean data indicated that salbutamol was a partial agonist when compared with isoprenaline. However, there was a large degree of interexperimental variability because, in 11 of 32 experiments, salbutamol was a full agonist and, in 21 of 32 experiments, a partial agonist relative to isoprenaline. These data suggest that different HLMC preparations possess variable receptor reserves. 2. The effect of the irreversible beta-adrenoceptor antagonist, bromoacetylalprenolol menthane (BAAM), on the inhibition of IgE-mediated histamine release by both isoprenaline and prostaglandin E2 (PGE2) was assessed. Whereas BAAM (100 nM) antagonized the isoprenaline inhibition of histamine release from activated HLMC, BAAM had no effect on the PGE2 inhibition. Pretreatment of HLMC with the beta2-selective competitive antagonist, ICI 118551 (100 nM), protected against the loss in responsiveness to isoprenaline following treatment with BAAM. 3. Concentrations of 1, 10 and 100 nM of BAAM caused dose-dependent rightward shifts in the dose-response curve for the isoprenaline inhibition of histamine release. Furthermore, there was a dose-dependent reduction in the maximal inhibitory response obtained with isoprenaline following treatments with increasing concentrations of BAAM. Although the rightward shifts in the isoprenaline dose-response curves, with a given concentration of BAAM, were similar in all experiments, there was some variability in the depression of the maximal response in individual experiments. Thus, in 6 of 16 experiments, BAAM (1 nM) did not depress the maximal response to isoprenaline, whereas in 10 of 16 experiments there was a depression (7 to 49% reduction) in the maximal response. These data suggest that different HLMC preparations possess variable receptor reserves. 4. Isoprenaline was more potent as an inhibitor in those HLMC preparations in which there was a larger receptor reserve (i.e. preparations in which the maximal inhibitory response to isoprenaline was unaffected by pretreatment with 1 nM BAAM). 5. The influence of receptor reserve on the inhibition by salbutamol of histamine release from HLMC was evaluated. There was a good correlation (r=0.77) between receptor reserve and the maximal response (relative to isoprenaline) obtained with salbutamol. Thus, HLMC preparations with larger receptor reserves were more responsive to salbutamol. 6. Receptor reserve influenced the desensitization of beta-adrenoceptor-mediated responses in HLMC. Cells were incubated (24 h) with isoprenaline (1 microM), washed and then the ability of a second isoprenaline (10(-10)-10(-5) M) exposure to inhibit histamine release was assessed. The pretreatment caused a reduction in the isoprenaline inhibition of histamine release although the extent of desensitization was highly variable, ranging from essentially negligible levels in some preparations to substantial reductions (93% desensitization) in the ability of isoprenaline to inhibit histamine release. There was a reasonable correlation (r=0.59) between receptor reserve and desensitization. Preparations that possessed a larger receptor reserve were more resistant to desensitization. 7. Collectively, these data suggest that a receptor reserve exists for the beta-adrenoceptor-mediated inhibition of histamine release from HLMC but that the size of this reserve varies between HLMC preparations. Moreover, the size of this receptor reserve may influence the sensitivity of HLMC to beta-adrenoceptor agonists and the susceptibility of individual HLMC preparations to desensitization.

Protein binding of aspirin and salicylate measured by in vivo ultrafiltration.

OBJECTIVE: Methods for measuring protein binding of drugs generally require direct measurement of the concentration of unbound drug and thus may require a highly sensitive assay. In vivo ultrafiltration has been used to determine protein binding of endogenous substances. We have examined its value for measuring protein binding of drugs because it requires measurement of only the concentration of total drug, not unbound drug, in plasma. METHODS: The protein binding of aspirin and its metabolite salicylate was measured in 29 healthy subjects 20 minutes after a single oral dose of 600 mg soluble aspirin, by the new method, in vivo ultrafiltration, as well as by a standard method, in vitro ultracentrifugation. RESULTS: The data for salicylate were examined systematically to determine the optimal method of determining estimates of protein binding by in vivo ultrafiltration. Estimates of protein binding of salicylate were 81.7% +/- 10.1% (mean +/- SD) by the in vivo method and 81.6% +/- 11.3% by in vitro ultracentrifugation. Bland-Altman analysis of agreement showed that within-individual differences in percentage of protein binding determined by the two methods did not differ significantly from zero (mean difference, 0.07%; 95% confidence interval, -2.33 to +2.46). There was a highly significant correlation between estimates of protein binding by the two methods (r = 0.82; p = 0.001). Protein binding of aspirin was estimated of protein binding by the two methods (r = 0.82; p = 0.001). Protein binding of aspirin was estimated at 58.3% +/- 9.6% by in vivo ultrafiltration and could not be estimated by in vitro ultracentrifugation because the concentration of unbound aspirin in plasma was below the limit of detection for the assay. CONCLUSION: In vivo ultrafiltration can be used to measure protein binding of drugs and has potential advantages over conventional methods. A sensitive assay may not be required because the unbound drug need not be measured, measurement in vivo may maintain more physiologic conditions, and it may be useful in measuring protein binding of drugs that are degraded rapidly in vitro.

Panic disorder, anxiety and depression in resistant hypertension--a case-control study.

BACKGROUND: It has been suggested that panic disorder can cause or contribute to hypertension or resistance to antihypertensive drugs. OBJECTIVE: To compare the prevalences of panic disorder, panic attacks, anxiety and depression between patients with resistant hypertension and age- and sex-matched patients with non-resistant hypertension. DESIGN: A case-control study of patients attending the Sheffield Hypertension Clinic, using self-completed postal questionnaires to assess panic disorder, anxiety and depression. PATIENTS CASES: With resistant hypertension were defined as patients who presently or previously had systolic blood pressure above 160 mmHg or diastolic blood pressure above 90 mmHg despite the use of three or more antihypertensive agents at full dose. For each of 136 cases, one control with non-resistant hypertension, defined as controlled to < or = 160/90 mmHg by one or two antihypertensive agents, was identified by a bias-free method. Cases and controls were matched for age and sex. MAIN OUTCOME MEASURES: Lifetime and current prevalence of panic attacks, the prevalences of panic disorder, anxiety and depression by Hospital Anxiety and Depression Scale scores, and the severity and frequency of panic attacks. RESULTS: Of the resistant hypertensive patients, 33% had experienced a panic attack compared with 39% of the control non-resistant hypertensives (resistant-non-resistant -6%, 95% confidence interval -19 to +7%). Twelve per cent of the resistant patients and 14% of controls fulfilled the criteria for a current or previous diagnosis of panic disorder (resistant-non-resistant -2%, 95% confidence interval -11% to +7%). There were also no significant differences between the groups in the prevalences of current panic attacks, panic attacks rated as moderate or worse, spontaneous panic attacks and in the frequency of panic attacks. There remained no significant difference between the groups for panic attacks and panic disorder when the analysis was limited to those patients who had idiopathic hypertension. The two groups did not differ significantly in scores for anxiety and depression measured by the Hospital Anxiety and Depression Scale. CONCLUSION: We observed no differences in the prevalences of panic, anxiety and depression between patients with resistant hypertension and non-resistant controls. These factors are probably not implicated in resistance to drug treatment. However, the prevalences of panic disorder and panic attacks were remarkably high in both groups of patients attending a hospital hypertension clinic. The relationship between panic disorder and hypertension deserves further study in a general hypertensive population.

Protection by dexamethasone of the functional desensitization to beta 2-adrenoceptor-mediated responses in human lung mast cells.

1. The beta-adrenoceptor agonist, isoprenaline, inhibited the IgE-mediated release of histamine from human lung mast cells (HLMC) in a dose-dependent manner. Maximal inhibitory effects were obtained with 0.1 microM isoprenaline. However, the inhibition of histamine release from HLMC by isoprenaline (0.1 microM) was highly variable ranging from 33 to 97% inhibition (mean, 59 +/- 3%, n = 27). 2. Long-term (24 h) incubation of HLMC with isoprenaline led to a subsequent reduction in the ability of a second exposure of isoprenaline to inhibit IgE-mediated histamine release from HLMC. The impairment in the ability of isoprenaline (0.1 microM) to inhibit histamine release following desensitizing conditions (1 microM isoprenaline for 24 h) was highly variable amongst HLMC preparations ranging from essentially negligible levels of desensitization in some preparations to complete abrogation of the inhibitory response in others (mean, 65 +/- 6% desensitization, n = 27). 3. The ability of HLMC to recover from desensitization was investigated. Following desensitizing conditions (1 microM isoprenaline for 24 h), HLMC were washed and incubated for 24 h in buffer and the effectiveness of isoprenaline (0.1 microM) to inhibit IgE-mediated histamine release from HLMC was assessed. The extent of recovery was highly variable with some HLMC preparations failing to recover and others displaying a complete restoration of responsiveness to isoprenaline (mean, 40 +/- 6% recovery, n = 23). 4. The effects of the glucocorticoid, dexamethasone, were also investigated. Long-term (24-72 h) treatments with dexamethasone (0.1 microM) had no effect on IgE-mediated histamine release from HLMC. Additionally, long-term (24-72 h) treatments with dexamethasone (0.1 microM) had no effect on the effectiveness of isoprenaline to inhibit histamine release. However, long-term (24-72 h) treatments with dexamethasone (0.1 microM) protected against the functional desensitization induced by incubation (24 h) of HLMC with isoprenaline (1 microM). The protective effect was time-dependent and pretreatment of HLMC with dexamethasone for either 24, 48 or 72 h prevented desensitization by either 15 +/- 7, 19 +/- 5 or 51 +/- 10%, respectively (n = 5-7). 5. HLMC preparations which were relatively refractory to isoprenaline even after withdrawal (24 h) from desensitizing conditions responded more effectively to isoprenaline (0.1 microM) if dexamethasone (0.1 microM) was also included during the recovery period (19 +/- 9% recovery after 24 h in buffer; 50 +/- 8% recovery after 24 h with dexamethasone, n = 5). 6. These data indicate that the responses of different HLMC preparations to isoprenaline, the susceptibility of HLMC to desensitization and the ability of HLMC to recover from desensitizing conditions varies markedly. Dexamethasone, which itself has no direct effects on IgE-mediated histamine release from HLMC, protected HLMC from the functional desensitization to beta-adrenoceptor agonists. Because beta 2-adrenoceptor agonists and glucocorticoids are important in the therapeutic management of asthma and as the HLMC is probably important in certain types of asthma, these findings may have wider clinical implications.

Cytochromes P450 mediating the N-demethylation of amitriptyline.

AIMS: Using human liver microsomes and heterologously expressed human enzymes, we have investigated the involvement of CYPs 1A2, 2C9, 2C19, 2D6 and 3A4 in the N-demethylation of amitriptyline (AMI), with a view to defining likely influences on its clinical pharmacokinetics. METHODS: The kinetics of formation of nortriptyline (NT) from AMI were measured over the substrate concentration range 1-500 microM, using liver microsomes from four extensive metabolisers (EM) and one poor metaboliser (PM) with respect to CYP2D6 activity. RESULTS: The data were best described by a two-site model comprising a Michaelis-Menten function for a high affinity site and a Hill function for a low affinity site. The activity at the low affinity site was eliminated by triacetyloleandomycin and ketoconazole, selective inhibitors of CYP3A4, such that the kinetics were then described by a two-site model comprising two Michaelis-Menten functions. A further decrease in activity was associated with the addition of the CYP2C9 inhibitor sulphaphenazole such that the residual kinetics were best described by a single Michaelis-Menten function. The addition of quinidine, a selective inhibitor of CYP2D6, along with triacetyloleandomycin and sulphaphenazole produced an additional decrease in the rate of NT formation in all but the PM liver, but did not completely eliminate the reaction. The remaining activity was best described by a single Michaelis-Menten function. Inhibitors of CYP1A2 (furafylline) and CYP2C19 (mephenytoin) did not impair NT formation. Microsomes from yeast cells expressing CYP2D6 and from human lymphoblastoid cells expressing CYP3A4 or CYP2C9-Arg N-demethylated AMI, but those from cells expressing CYPs 1A2 and 2C19 did not. CONCLUSIONS: We conclude that CYPs 3A4, 2C9 and 2D6 together with an unidentified enzyme, but not CYPs 1A2 and 2C19, mediate the N-demethylation of AMI. Thus, the clinical pharmacokinetics of AMI would be expected to depend upon the net activities of all of these enzymes. However, the quantitative importance of each isoform is difficult to predict without knowledge of the exposure of the enzymes in vivo to AMI.

Quantitative analysis of amitriptyline and nortriptyline in human plasma and liver microsomal preparations by high-performance liquid chromatography.

A simple, rapid, highly selective and sensitive method for the analysis of amitriptyline and nortriptyline in plasma and human liver microsomes is described. It is suitable for the routine analysis of large numbers of samples using readily available instrumentation and low cost consumables. The detection limit was 2 ng/ml for both compounds and calibration curves were linear over a wide range of concentrations and passed through the origin. The within-batch and between-batch coefficients of variation for amitriptyline and nortriptyline were less than 7.4% and 12.8%, respectively. A series of compounds, including inhibitors used for probing cytochrome P450 activity in vitro, were tested for interference in the assay. Only ketoconazole caused interference and the assay was modified to allow samples containing ketoconazole to be analysed.