[The role of the analytical laboratory in the monitoring of clinical studies].
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Clinical Trials as Topic↗
Biomedical subjects
R Pacifici
Publications and source records attributed to R Pacifici.
[Pharmacokinetics in a "special population": the aged].
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Aged↗
Household and community determinants of exposure to involuntary smoking: a study of urinary cotinine in children and adolescents.
The study examines the role of several potential predictors of urinary cotinine levels in a cross-sectional sample of 1,072 nonsmoking children and adolescents in Latium, Italy, during 1990-1991. As expected, there was a strong relation between passive exposure to smoking and the amount of maternal and paternal self-reported smoking. The urinary cotinine level increased with a decreasing level of paternal education and with an increasing index of household crowding; self-report of recent exposure to smoking outside the home was a strong predictor of the biologic marker. The analysis was then restricted to 346 subjects whose parents claimed that they were nonsmokers and that there were no smokers at home. In this group, however, 57 children reported some active smoking at home by their parents. Those with parents suspected to be "deceivers" had higher level of urinary cotinine than did those truly not exposed. In addition, urinary cotinine in this group was clearly associated with duration of exposure to smoking outside home. The study indicates that both factors related to family circumstances and exposure outside the household setting are strong determinants of urinary cotinine levels. The finding may be considered a direct confirmation that passive smoking among children should be viewed as a specific community responsibility.
Adolescent↗
Improved clean-up procedure for the high-performance liquid chromatographic assay of clomipramine and its demethylated metabolite in human plasma.
A rapid and selective assay of clomipramine and its metabolite desmethylclomipramine in human plasma, based on high-performance liquid chromatography with UV detection has been developed. The compounds were subjected to solid-phase extraction, using Extrelut 1 cartridges. Recoveries ranged between 88-95% for clomipramine, and 75-80% for desmethylclomipramine. This method has been used for therapeutic monitoring of clomipramine and its metabolite in individuals treated with this drug.
Antidepressive Agents, Tricyclic↗
Solid-phase extraction of nicotine and its metabolites for high-performance liquid chromatographic determination in urine.
A solid-phase extraction, using Extrelut-1 glass columns, has been applied to urine samples of both passive and active smokers for high-performance liquid chromatographic determination of nicotine and its metabolites cotinine and trans-3'-hydroxycotinine. Chromatography was performed using a reversed-phase LC8DB column and a mobile phase consisting of water-acetonitrile (80:9, v/v) containing 5 ml triethylamine, 670 mg/l sodium heptanesulphonate, and 0.034 M each of K2HPO4 and citric acid (pH 4.4), at a flow-rate of 1.6 ml/min. The results obtained indicate that solid-phase extraction is a reliable and quick procedure which can be applied also to other nicotine metabolites.
Chromatography, High Pressure Liquid↗
High-performance liquid chromatographic-electrospray mass spectrometric determination of morphine and its 3- and 6-glucuronides: application to pharmacokinetic studies.
A rapid and selective assay of morphine and its 3- and 6-glucuronides in serum, based on high-performance liquid chromatography-electrospray mass spectrometry has been developed. The analytes and the internal standard, codeine or naltrexone, were subjected to solid-phase extraction, using ethyl solid-phase extraction columns, prior to chromatography. A reversed-phase column and a gradient mobile phase consisting of water and methanol were used. The mass spectrometer was operated in the selected-ion monitoring mode. The following ions were used: m/z 286 for morphine, m/z 300 for codeine, m/z 342 for naltrexone, and m/z 462 for morphine 3- and 6-glucuronides. The limit of quantitation observed with this method was 10 ng/ml morphine, 50 ng/ml morphine-6-glucuronide and 100 ng/ml morphine-3-glucuronide. The present method proved useful for the determination of serum levels of the parent drug and its metabolites in pain patients, heroin addicts and in morphine-treated mice.
Animals↗
Environmental tobacco smoke: nicotine and cotinine concentration in semen.
Forty-three nonsmokers attending an infertility clinic reported their degree of passive exposure to environmental tobacco smoke (ETS) and provided samples of blood and seminal plasma. Levels of nicotine and cotinine in biological fluid were evaluated by a radioimmunoassay. The data indicate that exposure to ETS results in measurable nicotine and cotinine levels in seminal plasma. Seminal plasma cotinine concentration also shows a significant positive correlation with degree of reported exposure.
Adult↗
Appendicular cortical bone loss after age 65: sex-dependent event?
Distal radius photodensitometric and second metacarpal radiogrammetric measurements were obtained from computerized analyses of standard hand X-Ray films of 296 Caucasian subjects (189 women and 107 men). This sample included 134 subjects > or = 65 years old (75 women and 59 men). Distal radius bone density and metacarpal index showed a significant linear decrease with age in both sexes. Rates of bone loss, calculated from the regression curves, were -0.7% per year in women and -0.5% per year in men by distal radial photodensitometry, and -0.49% per year in women and -0.33% per year in men by metacarpal radiogrammetry. In the elderly subgroup, women > or = 65 years of age showed an even faster bone loss, with an annual decrease of -1.4% by distal radial photodensitometry. Conversely, men > or = 65 years of age had no significant bone loss, not even by metacarpal radiogrammetry. In conclusion, these data suggest that appendicular cortical bone loss occurs at a higher rate in elderly females than in the elderly males, both at the distal radial and at the metacarpal site.
Absorptiometry, Photon↗
Morphine affects cytostatic activity of macrophages by the modulation of nitric oxide release.
The serum levels of morphine and its glucuronide metabolites were quantitated in C57BL/6 mice at various intervals following subcutaneous administration of morphine. Since one of the major mechanisms of killing by macrophages is the production of nitric oxide, pharmacokinetics data were correlated with cytostatic activity and the release of NO2- (stable end product of NO metabolism). Morphine and its 3-glucuronide metabolite appear in serum of treated mice, reaching a peak of concentration at 20 min. However, morphine 3'-glucuronide levels were much higher than those of the drug itself, even when the morphine concentration levelled off. Both cytostasis and NO2- production of L1210-activated macrophages were significantly enhanced by opioid treatment immediately after drug injection (peaking after 40 min). In contrast, morphine induced a strong inhibition of both cytostasis and NO2- production 24 h after treatment. The modulation of both cytostasis and NO2- production induced by morphine was completely antagonized by pretreatment of mice with the opioid antagonist naltrexone. The involvement of an inducible isoform of NO synthase was suggested by the inhibitory effects of dexamethasone on NO2- production. These data indicate that in vivo administration of morphine can induce a modulation of the NO biosynthesis of peritoneal macrophages.
Analgesics, Opioid↗
Cytokine production in blood mononuclear cells from epileptic patients.
Although convulsive disorders have been associated with immune abnormalities, little is known about cytokine production in epileptic patients. We evaluated the concentrations of interleukin-1 alpha (IL-1 alpha), interleukin-1 beta (IL-1 beta), interleukin-2 (IL-2), and interleukin-6 (IL-6) secreted by peripheral blood mononuclear cells (PBMC) from epileptic patients. The PBMC collected from epileptic patients, as compared with those of a control group, showed greater production of IL-1 alpha, IL-1 beta, and IL-6 in response to in vitro stimulation with mitogen. There was no statistical difference among patients treated with different antiepileptic drugs (AEDs). Significantly greater IL-2 production was observed in PBMC from carbamazepine (CBZ)-treated patients as compared with both the control group or with valproate and phenobarbital (PB)-treated patients.
Adolescent↗
Immunologic aspects of vigabatrin treatment in epileptic children.
Vigabatrin (VGB) is an antiepileptic drug (AED) that acts by irreversibly inhibiting gamma-aminobutyric acid transaminase (GABA-T). To evaluate immune responses to GVG, we studied 29 idiopathic or symptomatic epileptic children and also examined a control group (n = 15). Epileptic children were tested before and after 1 and 3 months of VGB treatment. Whole blood was used to connect subsets with commercial monoclonal antibodies. Peripheral blood mononuclear cells (PBMC) were used to assess natural killer (NK) cell activity and lymphocyte response to phytohemagglutinin (PHA) and concavalin A (Con A). Immunoglobulin (Ig) levels were tested in serum. At baseline, no immunologic abnormalities were observed in either control or treated patients. During treatment, the percentage and absolute number of B lymphocytes, serum concentration of Ig, number of T total mature lymphocytes (CD3), T-rosetting lymphocytes (CD11), T-helper cells (CD4), and mitogenic response of lymphocytes remained unchanged. Several other immunologic responses showed a statistically significant increase after 1 and 3 months of VGB treatment, however, including the percentage and absolute number of T-suppressor cells (CD8) and NK cells and NK cell activity. The correlation between number of NK cells and NK cell activity was significant. Data obtained demonstrated that VGB may interfere with the modulation of the immune system, especially cytotoxic cell populations.
Anticonvulsants↗
Simultaneous block of interleukin-1 and tumor necrosis factor is required to completely prevent bone loss in the early postovariectomy period.
Considerable evidence supports the hypothesis that estrogen prevents bone loss by blocking the production of cytokines in bone or bone marrow. However, controversy remains on the role of candidate factors, such as interleukin-1 (IL-1), IL-6, and tumor necrosis factor (TNF). As IL-1 and TNF have many additive and/or synergistic effects in bone, we tested the hypothesis that the simultaneous block of IL-1 and TNF is required to prevent the initial phase of rapid bone loss that follows ovariectomy (ovx). To this aim, rats were ovariectomized and treated for 2 weeks with either IL-1 receptor antagonist (IL-1ra), an inhibitor of IL-1, or TNF-binding protein (TNFbp), an inhibitor of TNF. Ovx increased bone marrow cell secretion of IL-1 and TNF and decreased the bone density of the distal femur, as measured by dual energy x-ray absorptiometry. Ovx-induced bone loss was decreased by both IL-1ra and TNFbp and completely prevented by simultaneous treatment with IL-1ra and TNFbp. Combined treatment with IL-1ra and TNFbp decreased urinary pyridinoline cross-links, a marker of bone resorption that reflects osteoclast number and osteoclast activity, whereas treatment with either inhibitor alone was less effective. Both IL-1ra and TNFbp decreased the number of osteoclasts on the endocortical surfaces and stimulated bone formation, but the two inhibitors had no additive effects on these indexes, suggesting that inhibition of osteoclastogenesis and stimulation of bone formation do not account for the additive bone-sparing effects of IL-1ra and TNFbp. These inhibitors had no effect in sham-operated rats, indicating that they specifically blocked estrogen-dependent events. In conclusion, these data indicate that in the early post-ovx period, IL-1 and TNF play a critical causal role in inducing bone loss and do so by stimulating bone resorption and inhibiting bone formation.
Absorptiometry, Photon↗
Production of interleukin-6 in human osteoblasts and human bone marrow stromal cells: evidence that induction by interleukin-1 and tumor necrosis factor-alpha is not regulated by ovarian steroids.
Studies in murine models of osteoporosis have suggested the hypothesis that ovarian steroids may control osteoclastic bone remodeling by limiting the production of interleukin-6 (IL-6) from osteoblasts and bone marrow stromal cells. To investigate this hypothesis in a human model, we have examined 12 separate strains of normal human osteoblasts (HOB) and 11 separate strains of human bone marrow stromal cells (HBMSC) and determined whether ovarian steroids regulate the induction of IL-6 by interleukin-1 beta (IL-1 beta), tumor necrosis factor-alpha (TNF-alpha) or IL-1 + TNF. Treatment with IL-1, TNF or IL-1 + TNF resulted in the induction of IL-6 from both cell types with IL-1 + TNF inducing a synergistic induction of IL-6 in HOB (24- to 324-fold) and HBMSC (35-288 fold). Addition of 17 beta-estradiol or progesterone did not significantly alter IL-6 messenger RNA or protein levels in either HOB or HBMSC cultures stimulated with IL-1, TNF or IL-1 + TNF. Cultures incubated up to 96 h with the steroids did not affect IL-6 expression. Furthermore ovarian steroids did not affect IL-6 production in either HBMSC cultures representative of preosteoblasts or HOB cultures representative of highly differentiated osteoblasts. Specific chloramphenicol acetyl transferase assays and reverse transcriptase-polymerase chain reaction studies also demonstrated that the lack of an estrogen effect was not due to the failure of HOB to express functional estrogen receptors. Therefore, we conclude that the regulation of human osteoclastic bone remodeling by ovarian steroids does not occur through the direct regulation of IL-6 gene transcription or protein secretion in either early stages of osteoblast differentiation or the differentiated osteoblast.
Adolescent↗
Ligand binding to monocyte alpha 5 beta 1 integrin activates the alpha 2 beta 1 receptor via the alpha 5 subunit cytoplasmic domain and protein kinase C.
Regulation of the functional status of integrin receptors plays a critical role in inflammation and tissue remodeling, as it affects cell adherence and cytokine secretion. We have previously shown that in monocytes the binding of collagen to the alpha 2 beta 1 integrin induces the release of IL-1, an event that is potentiated by binding of fibronectin (Fn) to the alpha 5 beta 1 integrin. In this study, we have investigated the mechanisms leading to this phenomenon. Fn binding to alpha 5 beta 1 induced intracellular signals which increased the alpha 2 beta 1-dependent adhesiveness of monocytes to collagen without modifications of alpha 2 beta 1 expression. By using Abs against the intracellular region of the alpha 5 subunit of the alpha 5 beta 1 receptor, and specific inhibitors of protein kinase C (PKC), we found that the potentiation effect of Fn on monocyte IL-1 production and their adherence to collagen was dependent on an intact alpha 5 subunit cytoplasmic domain, and required PKC activation. Although the alpha 2 beta 1 could be activated by several intracellular second messengers, including protein kinase A and intracellular calcium, the potentiating effect of Fn was mediated only by PKC. These data provide an example of a novel regulatory mechanism: potentiation of beta 1 integrin-mediated events as a result of ligand binding to another integrin of the same class. They also show that the intracellular region of alpha 5 beta 1 plays a critical role in transducing signals generated by ligand binding to alpha 5 beta 1.
Calcium↗
Persistent bone-sparing effect of interleukin-1 receptor antagonist: a hypothesis on the role of IL-1 in ovariectomy-induced bone loss.
The recent finding that treatment with the interleukin-1 (IL-1) inhibitor, interleukin-1 receptor antagonist (IL-1ra) decreases bone loss and bone resorption in ovariectomized rats, strongly suggested that IL-1 mediates, at least in part, the effects of estrogen deficiency on bone resorption. Although in vitro studies have shown that IL-1 activates mature osteoclasts and stimulates osteoclastogenesis, the two main mechanisms by which estrogen deficiency stimulates bone resorption, it is still unclear whether IL-1 mediates both effects of estrogen deficiency in vivo. To investigate this matter, we have examined the changes in bone mineral density (BMD) which occur in ovariectomized rats after completion of 1 month of estrogen or IL-1ra treatment begun at the time of ovariectomy. Ovariectomy caused a marked decreased in BMD which was blocked by 17 beta estradiol and decreased by IL-1ra. Cessation of estrogen therapy was followed by a rapid induction of bone loss, indicating that estrogen blocks the activation and utilization of mature osteoclasts without depleting the bone microenvironment of osteoclast precursors and mature, inactive osteoclasts. In contrast, ovariectomized rats treated with IL-1ra maintained a stable bone density for the first 4 weeks after completion of the treatment. In these rats, bone loss resumed not earlier than 6 weeks after discontinuation of the IL-1ra treatment. Estrogen deficiency was necessary to unveil the bone-sparing effect of IL-1ra because in a control experiment in which rats were treated with IL-1ra for the 4 weeks before ovariectomy, BMD began to decrease immediately after ovariectomy.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
CYP2D6-related oxidation polymorphism in Italy.
The distribution of the oxidation polymorphism related to cytochrome CYP2D6 (debrisoquine type) was determined in 246 healthy Italian volunteers. Phenotyping was based on HPLC determination of the dextrometorphan/dextrorphan concentration ratio (metabolic ratio) in urine samples collected over an 8 h interval following a single oral 30 mg dose of dextromethorphan hydrobromide. Urinary excretion of dextromethorphan showed a wide interindividual variability, ranging from < or = 0.04 to 3.9% and from 0.5 to 79.6% of the dose, respectively. Metabolic ratios ranged from < or = 0.001 to 6.6. Eleven of the 246 subjects showed a metabolic ratio greater than 0.30, indicating that 4.5% of the population could be ascribed to the poor metabolizer status. The frequency of the poor metabolizer phenotype in this population is within the range described for other Caucasian ethnic groups.
Cytochrome P-450 CYP2D6↗
Pregnancy-associated osteoporosis: report of two cases with long-term bone density follow-up.
Pregnancy-associated osteoporosis is a rare and poorly characterized clinical entity. We describe two women, aged 28 and 31 yr, who developed vertebral osteopaenia with non-traumatic compression fractures within a few months after delivery. In both women secondary causes of osteoporosis were excluded and routine laboratory data including indices of calcium metabolism were normal. In one patient (Patient 1) spontaneous secretion of IL-1 by cultured blood monocytes was elevated 6 months after delivery and a bone biopsy revealed histomorphometric indices consistent with accelerated bone remodelling. BMD was measured by quantitative computed tomography (QCT) and/or dual energy X-ray absorptiometry (DXA). In Patient 1, BMD, measured with QCT, 6 months after delivery was 3.32 Z-score units below the predicted value for age and sex-matched subjects. At that time treatment (phosphate followed by sodium etidronate) was instituted, and QCT measurements obtained 24 and 30 months after delivery revealed a BMD of -2.98 and -2.67 Z-score units, respectively. The treatment was discontinued just before she became pregnant again, and 3 months after the second delivery QCT showed a BMD of -3.06 Z-score units. Further BMD assessments were obtained with DXA at 15 and 18 months after the second delivery, and the BMD values were -1.41 and -1.08, respectively. In Patient 2, spine BMD, measured with QCT, 4 and 10 months after delivery were -1.68 and -1.59 Z-score units, respectively. At 10, 16 and 22 months after delivery, BMD was assessed with DXA. The initial measurement with this technique disclosed a BMD of 0.52 Z-score units.(ABSTRACT TRUNCATED AT 250 WORDS)
Absorptiometry, Photon↗
Osteoporosis: diagnosis with lateral and posteroanterior dual x-ray absorptiometry compared with quantitative CT.
PURPOSE: To compare the diagnostic sensitivity of posteroanterior and lateral dual x-ray absorptiometry (PA-DXA, L-DXA, respectively) and quantitative computed tomography (CT). MATERIALS AND METHODS: Among 108 women undergoing lumbar spine bone mineral density assessment, 66 were healthy (mean age, 52.9 years +/- 1.2 [standard error of mean]) and 42 had osteoporosis (mean age, 66.9 years +/- 1.2). RESULTS: Although both L-DXA and PA-DXA correlated well with quantitative CT (r = .73 and .72, respectively; P < .0001), L-DXA correlated better than PA-DXA with age (r = -.69 and -.50, respectively; P < .0001). Women with osteoporosis showed higher bone loss with quantitative CT (1.33% per year) and L-DXA (0.3% per year) than with PA-DXA (0.07% per year). Logistic regression analysis indicated that quantitative CT and L-DXA but not PA-DXA are significant predictors of osteoporotic fractures. Receiver-operating-characteristic curve analyses showed L-DXA to have a sensitivity and specificity closer to those of quantitative CT than did PA-DXA. CONCLUSION: Performance of L-DXA helped discriminate better than PA-DXA between healthy subjects and those with osteoporosis.
Absorptiometry, Photon↗