Forensic investigation on a combined death by food aspiration and acute escitalopram intoxication occurred to a psychiatric subject in a nursing home.

Food aspiration is one of the major health risks for elderly people in nursing homes which could lead to death. Moreover, misconducts in pharmacotherapy may represent a potential risk of adverse drug reactions. It is reported here the toxicological evaluation of a combined death by food aspiration and acute escitalopram intoxication of a psychiatric subject, occurred in a nursing home. An 89-year-old man, suffering from dysphagia and Alzheimer's, was resident in a nursing home. He was fed with a liquid diet administered directly in mouth using a syringe. The man was also being treated with escitalopram 10 mg tablet. One evening, after receiving the meal in the usual way, the man complained of sudden illness. Carried to the emergency room, the man died about 3 h later with a diagnosis of cardiogenic shock subsequentially to ab ingestis. The histological findings revealed the presence of exogenous material, probably food, up to the finest bronchial branches. The toxicological examination revealed the presence of escitalopram and its main metabolite, desmethylcitalopram: in the blood 1972 ng/ml and 285 ng/ml, in the brain 4657 ng/g and 1025 ng/g, in the gastric content 2317 ng/g and 423 ng/g, in the lung 21,771 ng/g and 468 ng/g, respectively. The bad practice of the nurses to dissolve the escitalopram tablet in the liquefied food and to administer the therapy with a syringe directly into the mouth emerged thanks this investigation. Following food aspiration, escitalopram was absorbed by inhalation route, reaching high concentrations in blood and tissues. The death occurred due to a combined mechanism between food aspiration and the escitalopram toxic action.

Lethal vortioxetine poisoning? A forensic investigation.

Vortioxetine is an antidepressant recently licensed in the US and EU for the treatment of major depressive disorder. No fatalities from vortioxetine overdose have been reported, yet. Two cases of attempted suicide are described in the literature, although no toxicological analyzes were conducted. Vortioxetine concentrations found in blood and organs in a case of a probable acute lethal intoxication are reported here. A 65-year-old woman was found on the floor behind her bed with no vital signs. The woman was recently on vortioxetine 10 mg/day for major depression, anxiety, and psychotic attacks. Vortioxetine was quantified in blood, brain, liver, kidney, and lung samples by LC-MS/MS. Vortioxetine concentrations were: 1.197 ng/ml in the blood, 804 ng/g in the brain, 8.992 ng/g in the lung, 1.389 ng/g in the liver, 292 in the kidney. No other substance was found. In the case reported here, the blood concentration was approximately 35-135 times higher than the antemortem therapeutic value. Histological examination showed signs of a probable sudden cardiac death following to arrhythmia, with no evidence of myocardial infarction. The present case indicate that blood concentrations close to 1,000 ng/mL could lead to death, involving probably to a cardiac toxicity.

Uniparental disomy causes deficiencies of vitamin K-dependent proteins.

Essentials Vitamin K-dependent coagulant factor deficiency (VKCFD) is a rare autosomal recessive disorder. We describe a case of inherited VKCFD due to uniparental disomy. The homozygous mutation caused the absence of GGCX isoform 1 and overexpression of Δ2GGCX. Hepatic and non-hepatic vitamin K-dependent proteins must be assayed to monitor VKCFD treatment. SUMMARY: Background Inherited deficiency of all vitamin K-dependent coagulant factors (VKCFD) is a rare autosomal recessive disorder caused by mutations in the γ-glutamyl carboxylase gene (GGCX) or the vitamin K epoxide reductase gene (VKORC1), with great heterogeneity in terms of both clinical presentation and response to treatment. Objective To characterize the molecular basis of VKCFD in a Spanish family. Methods and Results Sequencing of candidate genes, comparative genomic hybridization and massive sequencing identified a new mechanism causing VKCFD in the proband. Uniparental disomy (UPD) of chromosome 2 caused homozygosity of a mutation (c.44-1G>A) resulting in aberrant GGCX splicing. This change contributed to absent expression of the mRNA coding for the full-length protein, and to four-fold overexpression of the smaller mRNA isoform lacking exon 2 (Δ2GGCX). Δ2GGCX might be responsible for two unexpected clinical observations in the patient: (i) increased plasma osteocalcin levels following vitamin K1 supplementation; and (ii) a mild non-bleeding phenotype. Conclusions Our study identifies a new autosomal disease, VKCFD1, caused by UPD. These data suggest that the Δ2GGCX isoform may retain enzymatic activity, and strongly encourage the evaluation of both hepatic and non-hepatic vitamin K-dependent proteins to assess differing responses to vitamin K supplementation in VKCFD patients.

Identification of miRNAs as potential modulators of tissue factor expression in patients with systemic lupus erythematosus and antiphospholipid syndrome.

BACKGROUND: Tissue factor (TF) is the main initiator of the coagulation cascade and elements that may upregulate its expression might provoke thrombotic events. Systemic lupus erythematosus (SLE) and antiphospholipid syndrome (APS) are autoimmune diseases characterized by a high TF expression in monocytes. OBJECTIVES: To examine the role of microRNAs (miRNAs) in TF expression and to evaluate their levels in SLE and APS patients. METHODS: An in silico search was performed to find potential putative binding sites of miRNAs in TF mRNA. In vitro validation was performed transfecting cells expressing TF (THP-1 and MDA-MB-231) with oligonucleotide miRNA precursors and inhibitors. Additionally, reporter assays were performed to test for the binding of miR-20a to TF mRNA. Levels of miRNAs and TF were measured by quantitative (qRT-PCR) in patients with APS and SLE. RESULTS: Overexpression of miRNA precursors, but not inhibitors, of two of the members of cluster miR-17∼92, for example miR-19b and miR-20a, in cells expressing TF decreased TF mRNA, protein levels, and procoagulant activity between 30% and 60%. Reporter assays showed that miR-20a binds to TF mRNA. Finally, we measured levels of miR-19b and miR-20a in monocytes from patients with APS and SLE and observed significantly lower miRNAs levels in comparison with healthy subjects inversely correlated with the levels of TF. CONCLUSIONS: Down-regulation of miR-19b and miR-20a observed in patients with SLE and APS could contribute to increased TF expression and thus provoke the hypercoagulable state characteristic of these patients.

Pharmacogenetics of acenocoumarol in patients with extreme dose requirements.

SUMMARY BACKGROUND: There is currently intense debate as to whether pharmacogenetic algorithms for estimating the initial dose of coumarins provide a more accurate dose than the fixed-dose approach. Recently, it has been suggested that the greatest benefit of pharmacogenetic algorithms is observed in patients with extreme dose requirements. OBJECTIVES: To identify clinical and genetic factors that better characterize patients who need extreme acenocoumarol doses for steady anticoagulation state. PATIENTS/METHODS: We reviewed 9538 patients with a steady acenocoumarol dose from three Spanish hospitals, selecting 83 who took or= 30.00 mg week(-1) (p95). We also selected patients matched by gender and age taking 13.50-14.00 mg week(-1) (p50). We genotyped VKORC1 (rs9923231), CALU (rs1043550), GGCX (rs699664), CYP2C9 (rs1799853; rs1057910), CYP4F2 (rs2108622) and F7 (rs5742910) single-nucleotide polymorphisms (SNPs). RESULTS: Comparison between p5 and p95 revealed five parameters with significant differences: body surface area (BSA) (P = 0.006), age, VKORC1, CYP2C9 and CYP4F2 genotypes (all P < 0.001). First VKORC1, and second, CYP2C9 SNPs played a strong effect by determining extreme doses, particularly in p95. Only one out of 203 p95 had the VKORC1 A-1639A genotype, but this subject was CYP2C9*1/*1. In contrast, nine out of 83 p5 carried the VKORC1 G-1639G genotype, although six of them were CYP2C9*3 homozygotes and another two were heterozygotes. Surprisingly, CYP4F2 V433M SNP displayed prevalences that suggest that its influence might only be evident when patients are treated with high doses. CONCLUSION: Two clinical data, age and BSA, and three SNPs in the VKORC1, CYP2C9 and CYP4F2 genes strongly predict outlier patients treated with acenocoumarol.

Human and yeast zeta-crystallins bind AU-rich elements in RNA.

Zeta-crystallins constitute a family of proteins with NADPH:quinone reductase activity found initially in mammalian lenses but now known to be present in many other organisms and tissues. Few proteins from this family have been characterized, and their function remains unclear. In the present work, zeta-crystallins from human and yeast (Zta1p) were expressed, purified and characterized. Both enzymes are able to reduce ortho-quinones in the presence of NADPH but are not active with 2-alkenals. Deletion of the ZTA1 gene makes yeast more sensitive to menadione and hydrogen peroxide, suggesting a role in the oxidative stress response. The human and yeast enzymes specifically bind to adenine-uracil rich elements (ARE) in RNA, indicating that both enzymes are ARE-binding proteins and that this property has been conserved in zeta-crystallins throughout evolution. This supports a role for zeta-crystallins as trans-acting factors that could regulate the turnover of certain mRNAs.

Hemostasis and fibrinolysis factors in first-degree relatives of patients with Type 2 diabetes without hypertension.

First-degree relatives of type 2 diabetic patients with or without a family history of hypertension are at increased risk for cardiovascular diseases. The aim of this study was to verify some possible hemostatic alterations in first-degree relatives of type 2 diabetic, normotensive and hypertensive patients. In 78 non-diabetic, normotensive first-degree relatives of type 2 diabetic patients (47 without a family history of hypertension and 31 with a family history of hypertension) and in 36 normoglycemic, normotensive subjects with no family history of hypertension, we evaluated plasma levels of fasting glucose and insulin, tissue-type plasminogen activator (t-PA), plasminogen activator-inhibitor (PAI-1), D-dimer (DD) and prothrombin fragment 1 + 2 (F1+2). Insulin resistance, calculated by the HOMA model, and plasma levels of t-PA and PAI-1 were significantly higher in relatives of diabetics compared to controls. As far as the thrombin activation indexes are concerned, we detected a significant increase in DD and F1+2 in relatives of diabetics with hypertension compared to other study subjects. In conclusion, our data indicate that familial predisposition may influence the hemostatic system in first-degree relatives of diabetic and/or hypertensive patients.

Treatment with methylprednisolone in relapses of multiple sclerosis patients: immunological evidence of immediate and short-term but not long-lasting effects.

Relapses of multiple sclerosis (MS) are treated commonly with high-dose intravenous methylprednisolone (MP) given over a period of 3-5 days. The mechanisms responsible for the beneficial effects of MP in attacks are not clearly established. It is also controversial whether this treatment may have a long-term effect. Here, peripheral blood samples from relapsing--remitting MS patients in acute relapse were analysed by flow cytometry just before steroid treatment and at different time points after initiation of the therapy. We observed an immediate (day 3) decrease in the percentage of CD4+ lymphocytes, with a relative increase in the memory (CD4+CD45R0+) subpopulation. A longer standing effect of MP on IFN-gamma production, CD54, CCR5, CXCR3 and CD95 (Fas) expression was also observed on CD4+ cells after 1 month of treatment initiation. Six months after the therapy, during clinical remission, no changes due to ivMP therapy were detected. These results support that MP treatment of relapses induces immediate post-treatment and short-term effects on the immune system that could partly account for the clinical and radiological improvement observed in MS patients. However, no conclusion can be drawn as to a possible long-term or even intermediate influence of ivMP treatment on the course of the disease.

Hair testing for drugs of abuse: evaluation of external cocaine contamination and risk of false positives.

In some laboratories hair testing may be the main method for the evaluation of individual's drug history, however, compelling evidence supports the possibility that the presence of a small amount of drug in hair can derive from external contamination. The aim of the present study is to verify if a single external contamination with a small amount of cocaine will last sufficiently long to make a contaminated subject indistinguishable from active users, and if normal washing practices together with the decontamination procedures are sufficient to completely remove the external contamination. The results obtained using the decontamination methods suggested in literature demonstrate that significant concentrations of cocaine (>1 ng/mg) and moderate quantities of benzoylecgonine (generally <0.5 ng/mg) are still detectable up to 10 weeks after contamination. These results question the reliability of hair testing. In fact, even using the most sophisticated decontamination procedures it is not possible to distinguish a drug-contaminated subject from an active user. Thus, while a negative result excludes both chronic use and "contact" with drugs, a positive result cannot and must not be interpreted as a sure sign of drug addiction, but should be further confirmed by urine analysis.

Increased indexes of thrombin activation in advanced stages of hypertension.

BACKGROUND: The hemostatic system plays an important role in thrombotic lesions, which can complicate the clinical course of hypertensive patients. The aim of this study was to verify a possible activation of the blood clotting processes, the evaluation of two markers of thrombin activation in 62 hypertensive patients, with and without vascular complications, compared with a control group. METHODS AND RESULTS: In 22 patients with newly diagnosed uncomplicated essential hypertension, in 40 hypertensive patients with clinically evident vascular complications (20 patients with controlled blood pressure and 20 with uncontrolled and high blood pressure) and in 20 normotensive sex- and age-matched subjects, two indexes of thrombin generation and action, namely prothrombin fragment 1 + 2 (F1 + 2) and fibrinopeptide A (FPA) were evaluated. The observed values show an increase of the F1 + 2 levels in patients with overt vascular complications; those with higher blood pressure, moreover, showed FPA levels higher than those with controlled blood pressure. CONCLUSIONS: These results seem to indicate that plasma F1 + 2 levels are significantly elevated, as a marker of a thrombosis-prone status, in patients with organic damage. Successively, with progress of hypertension and increasing blood pressure, the evidence of elevated FPA levels seems to indicate a clear prethrombotic situation which could turn into a thrombotic state.

CD4(+)CD45RO(+)CD49d(high) cells are involved in the pathogenesis of relapsing-remitting multiple sclerosis.

In the animal model of multiple sclerosis, experimental autoimmune encephalomyelitis, encephalitogenic T cells differ from the non-encephalitogenic ones in their expression of CD49d. The disease-inducing CD49d(high) and not the CD49d(low) cells enter the brain parenchyma. In this context, we characterized CD4(+)(CD45RO(+))CD49d(high) cells in relapsing-remitting multiple sclerosis (RR-MS) patients. These cells, showing characteristics of activated cells able to produce pro-inflammatory cytokines, were found to be increased in peripheral blood during relapses and present in high numbers in cerebrospinal fluid. These results suggest that the CD4(+)CD45RO(+)CD49d(high) subpopulation in RR-MS patients includes autoreactive cells and may be target for immunotherapy.

Evaluation of haemostatic parameters and circadian variations of the haemostatic system in patients with systemic sclerosis and Raynaud's phenomenon.

BACKGROUND: Systemic sclerosis (SSc) is a multisystemic disease characterized by proliferation and swelling of endothelial cells and other disorders. Raynaud's phenomenon (RP) is a disturbance, with unknown pathogenesis, that may be a precursor to SSc. The aim of this study was to investigate possible alterations in the haemostatic system and to examine whether there is a circadian variation in haemostatic variables at the initial stage of SSc. METHODS: In 20 patients with RP (in all patients secondary to SSc) and in 10 controls the levels of thrombomodulin (TM), beta-thromboglobulin (beta-TG), D-dimer (DD), tissue-type plasminogen activator (t-PA) and plasminogen activator-inhibitor (PAI-1) were measured in venous plasma samples taken at 9.00 and 14.00. RESULTS: Only TM levels were found to be higher in patients than in controls. Moreover the PAI-I levels, in the patient group, showed a significant circadian rhythm (with peak values at 9.00). No significant circadian variations for the other parameters were detected. CONCLUSIONS: These data seem to indicate that in patients with RP there is an endothelial damage reflected by a significant elevation of the TM plasma level and a circadian variation in plasma PAI-1, which was higher in the morning. This observation may be an area worth exploring for its importance potential in the knowledge of Raynaud's phenomenon.

[Neutralizing antibodies in patients with multiple sclerosis treated with interferon beta-1b]. / Anticuerpos neutralizantes en pacientes con esclerosis múltiple tratados con interferón beta-1b.

BACKGROUND: Neutralising antibodies (NABs) against interferon beta have been described in one third of patients with multiple sclerosis treated with interferon beta. We have analysed the frequency of NABs and their clinical consequences. PATIENTS AND METHODS: We have studied 68 patients. NABs were determined by protein A Myxovirus assay. RESULTS: Positive NABs were detected in 13% of the patients after 2 years of treatment. CONCLUSIONS: It does not seem to exist a relationship between presence of NABs and a poor evolution of the disease in our patients with multiple sclerosis treated with beta interferon.

Immunological profile of patients with primary progressive multiple sclerosis. Expression of adhesion molecules.

Adhesion molecules are important in the trafficking of peripheral leucocytes into the central nervous system, a major event in the pathogenesis of multiple sclerosis, which is an inflammatory and demyelinating disease. The latest MRI evidence supports clinical divergence between forms of multiple sclerosis with relapses and the primary progressive form without relapses, which shows fewer and smaller inflammatory lesions. With the aim of elucidating whether different pathogenic mechanisms are involved in primary progressive multiple sclerosis, we compared membrane expression of the adhesion molecules ICAM-1 (CD54), LFA-1alpha (CD11a), VLA-4 [alpha(4)/beta(1) integrin (CD49d/CD29)], L-selectin (CD62L) and ICAM-3 (CD50) in peripheral blood and the serum-soluble forms ICAM-1, L-selectin, VCAM-1 and ICAM-3 in 89 patients (39 with the primary progressive form, 25 with the secondary progressive form and 25 with the relapsing-remitting form) and 38 healthy controls. We found a significant decrease in leucocyte surface expression of most of the adhesion molecules tested and an increase in soluble ICAM-1 and L-selectin levels in secondary progressive and relapsing-remitting multiple sclerosis compared with primary progressive multiple sclerosis, which gave results similar to those in controls. These results, which are supported by MRI evidence, show that trafficking of autoreactive leucocytes through the blood-brain barrier is crucial to the pathogenesis of secondary progressive and relapsing-remitting forms of multiple sclerosis, whereas other mechanisms leading to progressive axonal damage would account for primary progressive forms of the disease.

Efficacy evaluation of prostaglandin E1 against placebo in patients with progressive systemic sclerosis and significant Raynaud's phenomenon.

BACKGROUND: Raynaud's phenomenon, due to connective tissue diseases, is difficult to treat successfully. Symptomatic improvement has been reported using nifedipine or iloprost, but adverse side effects may limit their use. The purpose of this study was to examine the effects of PGE1 (Alprostadil) in patients with scleroderma and severe Raynaud's disease. METHODS: Twelve females, aged 50-67 years, were included in the study with six of them receiving a 3-hour infusion of alprostadil at the standard dosage of 60 micrograms in 250 cc of physiological infusion for six consecutive days and the remaining six receiving placebo (250 cc of physiological infusion administered in the same manner). RESULTS: After infusion, blood flow, digitally measured by telethermography was increased only in patients treated with alprostadil. The number, frequency and severity of attacks recorded were reduced only in patients treated with alprostadil. No side effects were recorded during and after the infusion. CONCLUSION: In conclusion, alprostadil is effective in the management of Raynaud's phenomenon, due to scleroderma.

Study of thrombinic activation indexes, in patients with lower limb critic ischaemia, before and after prostaglandin E1 therapy.

BACKGROUND: In this study the action of a prostaglandin, PGE1, was studied in a group of patients with peripheral arterial occlusive disease (PAOD). METHODS: In 16 patients (14 men and 2 women, aged 47-70 years, mean 57 +/- 7) with PAOD, Fontaine stage IIb and III in critical ischemia, the effects on two indexes of thrombin generation and action of the endovenous administration (2 hours) of 60 micrograms of Alprostadil-PGE1 for four weeks were evaluated. In all artheriopathic patients, before and after pharmacological treatment, the following haemostatic parameters were evaluated: the prothrombin fragment 1 + 2 (F1 + 2) and the fibrinopeptide A(FPA). RESULTS: The patients showed plasma levels of FPA significantly decreased at the end of the treatment. On the other hand, no significant difference in plasma F1 + 2 levels was observed after treatment. CONCLUSIONS: These results seem to indicate that plasma F1 + 2 levels are significantly elevated, as a marker of thrombosis status, in patients with PAOD before and after treatment with PGE1.

Clinical and haemostatic effects of intravenous prostaglandin E1 therapy in patients with peripheral arterial occlusive disease.

BACKGROUND: In this study the action of an antiaggregatory prostaglandin, PGE1, was studied in a group of patients with peripheral arterial occlusive disease (PAOD). METHODS: In 16 patients with PAOD Fontaine stage IIb and III the clinical and haemostatic effects of the endovenous administration of 60 micrograms/die of alprostadil-PGE1 for four weeks, were evaluated. Before and after pharmacological treatment, were evaluated the clinical symptoms (claudicatio intermittens and rest pain) and the following haemostatic parameters: plasma thrombomodulin (TM), beta-thromboglobulin (beta-TG), D-dimer (DD), tissue-type plasminogen activator (t-PA) and plasminogen activator-inhibitor (PAI-1). RESULTS: No significant difference in plasma TM, t-PA and PAI-1 levels was observed after the treatment. On the other hand the patients showed plasma levels of beta-TG and DD significantly decreased at the end of the treatment. From the clinical point of view both claudicatio intermittens and rest pain satisfactorily improved in all patients. CONCLUSIONS: This data confirmed the therapeutic efficacy of PGE1 in the treatment of PAOD patients.

A fatal case of benzene poisoning.

Chronic effects following repeated exposure to low doses of benzene have been well assessed, whereas few data are available about acute exposure to benzene. We report a case of fatal acute intoxication which occurred aboard a chemical cargo ship. Autopsy findings included blood clots inside the heart and main vessels, multi-organ congestion, pulmonary edema and the presence of many vibices in the hypostatic areas. Toxicological analysis of blood and urine showed a benzene concentration of 31.67 and 2.26 micrograms/mL, respectively; high concentrations of benzene (microgram/g) were also found in the lungs (22.23), liver (378.60), brain (178.66), heart (182.57) and kidneys (75.15). The above data provide evidence for benzene distribution in various organs.

Haemostatic effects of iloprost in patients with lower limb ischemia.

The aim of this study was to investigate the haemostatic effects of iloprost, a stable analogue of prostacyclin, in patients with peripheral arterial disease. In a group of 13 patients with obliterative arteriopathies of the lower limbs the plasma levels of thrombomodulin (TM), betathromboglobulin (beta-TG), D-dimer (DD) and plasminogen activator-inhibitor (pAI-1) were measured, and compared to the values obtained from 10 healthy volunteers. All the parameters were found to be significantly higher in vasculopathic patients. These haemostatic evaluations were carried out after 4 weeks of treatment with iloprost up to 2 ng/kg/min, 6 hours infusion per day. During and at the end of treatment a clinical improvement was recorded. The patients also showed a significant decrease in plasma beta-TG and DD at the end of treatment. These data suggest that iloprost exerts clinical improvement, in who may have a part the decrease of platelet activation and of fibrin turnover.