Chronic hyperplastic anemia as an independent risk factor for atherosclerotic lesions: a lesson from thalassemia intermedia

Introduction. Cardiovascular involvement represents a well-known complication and the primary cause of mortality, both in transfusion-dependent beta thalassemia major (β-TM) and in transfusion-independent beta thalassemia intermedia (β-TI). In β-TM, heart iron overload is considered the main cause of this complication. This is likely due to poor adherence to iron-chelating therapy, resulting in the inability of the body to efficiently remove iron excess derived from transfused red blood cell breakdown. Different clinical pictures may instead be evoked in cardiovascular involvement occurring in β-TI; however, until now, no factor has emerged as the major one responsible for these complications. 
Design and Methods. In the present study, iron status, and lipid profiles in serum, as well as lipid content in peripheral blood mononuclear cells (PBMCs) were evaluated in 70 adult β-TM and in 22 adult β-TI patients. Ninety-two age-matched blood donors, free from any form of thalassemia, were utilized as controls. The mRNA levels of genes involved in the regulation of iron metabolism, such as interleukine 1 alfa (IL1α), tumor necrosis factor alfa (TNFα), as well as those involved in cholesterol homeostasis, such as acetyl-coenzymeA: cholesterol acyltransferase (ACAT-1), neutral cholesterol ester hydrolase (nCEH), and ATP binding cassette-A (ABCA1), were also evaluated in PBMCs from the above subjects.
Results. In β-TI patients, serum iron, transferrin saturation and erythropoietin levels were higher, while transferrin and hepcidin were lower, compared to both β-TM and controls. Hepcidin and ILα mRNA levels were found to be reduced in β-TI-PBMCs, while those of TNFα were increased. A reduction in total and high density lipoprotein cholesterol (TC and HDL-C) in serum, and an accumulation of neutral lipids (NL), coupled with increased mRNA levels of ACAT-1 and decreased nCEH in PBMCs were also observed in β-TI. 
Conclusions. Since most of the parameters found to be altered in β-TI patients have a key role in the initiation and progression of atherosclerosis, we suggest that cardiovascular complications in these patients may be, at least partially, dependent on the occurrence of premature atherosclerotic lesions. 
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Metabolomic patterns associated to QTc interval in shiftworkers: an explorative analysis

Objectives: 1H NMR-metabolomic approach was used to investigate QTc interval correlation with plasma metabolic profiles in shiftworkers. Methods: Socio-demographic data, electrocardiographic QTc interval and plasma metabolic profiles from 32 male shiftworkers, were correlated by multivariate regression analysis. Results: We found a positive correlation between QTc interval values, body mass index, glycemia and lactate level and a negative correlation between QTc interval and both pyroglutamate and 3-hydroxybutyrate plasma level. Conclusions: Our analysis provides evidence of the association between clinical, metabolic profiles and QTc interval values. This could be used to identify markers of early effects and/or susceptibility in shiftworker

Interaction between dietary and lifestyle risk factors and N-Acetyltransferase polymorphisms in B-Cell lymphoma etiology

Background: Gene-environment interactions are suggested to play a role in lymphomagenesis. Methods: We tested the interaction between the NAT1/NAT2 phenotype, as inferred by the respective genotypes, and exposure to dietary and lifestyle risk factors, in 199 incident lymphoma cases and 188 population controls. We used unconditional logistic regression to calculate the odds ratio (OR) and its 95% confidence interval for lymphoma (all subtypes combined) and B cell lymphoma, associated to the rapid NAT1 phenotype and to the intermediate and slow NAT2 phenotype, and to the estimated dietary intake of heterocyclic amines and folate, current smoking, coffee, and use of permanent hair dyes, as well as the respective interaction terms. We adjusted the ORs by age, gender, and education, and we used the likelihood ratio test to test the interaction between the NAT1, NAT2 phenotype and the dietary and lifestyle variables. Results: We observed an increase in risk of lymphoma (all subtypes combined) and B-cell lymphoma in particular associated with the estimated above median dietary intake of heterocyclic amines (OR = 4.2, 95%CI 1.2 – 14.8) and folate (OR = 4.1, 95%CI 0.7 – 22.4) among subjects with the NAT1 rapid acetylator phenotype, but not independent on the NAT1 phenotype. The test for interaction was significant for heterocyclic amines, but not for folate (p for interaction = 0.026 and 0.076 respectively). Ever use of permanent hair dyes was associated with an elevated risk independent on the NAT1, NAT2 phenotypes; risk decreased to null among intermediate and slow NAT1 acetylators (p for interaction = 0.010). Conclusions: Our results suggest that NAT1, NAT2 polymorphisms interact with dietary and lifestyle exposures in modulating risk of lymphoma and particularly B-cell lymphoma

Proapoptotic Activity and Chemosensitizing Effect of the Novel Akt Inhibitor (2S)-1-(1H-Indol-3-yl)-3-[5-(3-methyl-2H-indazol-5-yl)pyridin-3-yl]oxypropan2-amine (A443654) in T-Cell Acute Lymphoblastic Leukemia

Constitutively activated AKT kinase is a common feature of T-cell acute lymphoblastic leukemia (T-ALL). Here, we report that the novel AKT inhibitor (2S)-1-(1H-indol-3-yl)-3-[5-(3-methyl-2Hindazol-5-yl)pyridin-3-yl]oxypropan2-amine (A443654) leads to rapid cell death of T-ALL lines and patient samples. Treatment of CEM, Jurkat, and MOLT-4 cells with nanomolar doses of the inhibitor led to AKT phosphorylation accompanied by dephosphorylation and activation of the downstream target, glycogen synthase kinase-3â. Effects were time- and dose-dependent, resulting in apoptotic cell death. Treatment of Jurkat cells with A443654 resulted in activation of caspase-2, -3, -6, -8, and -9. Apoptotic cell death was mostly dependent on caspase-2 activation, as demonstrated by preincubation with a selective pharmacological inhibitor. It is remarkable that A443654 was highly effective against the drug-resistant cell line CEMVBL100, which expresses 170-kDa P-glycoprotein. Moreover, A443654 synergized with the DNA-damaging agent etoposide in both drug-sensitive and drug-resistant cell lines when coadministered [combination index (CI) = 0.39] or when pretreated with etoposide followed by A443654 (CI = 0.689). The efficacy of A443654 was confirmed using blasts from six patients with T-ALL, all of whom displayed low levels of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and constitutive phosphorylation of Akt on Ser473. At 1 ìM, the inhibitor was able to induce apoptotic cell death of T-ALL blast cells, as indicated by flow cytometric analysis of samples immunostained for active (cleaved) caspase-3. Because activated AKT is seen in a large percentage of patients with T-ALL, A443654, either alone or in combination with existing drugs, may be a useful therapy for primary and drug-resistant T-ALL. Originally published Molecular Pharmacology, Vol. 74, No. 3, Sep 200

In vitro synergistic anti-prion effect of cholesterol ester modulators

Background. Our studies on the role of cholesterol in prion infection/replication showed that brains and peripheral cells of sheep susceptible to or suffering from Scrapie were characterized by an altered cholesterol homeostasis compared to animals with a scrapie-resistant genotype, and that drugs influencing cholesterol esterification were endowed with selective anti-prion activity in N2a cell lines infected with the 22L and RML prion strains. Results. In prion-infected N2a cell lines we now report increased anti-prion activity of dual-drug combinations consisting of cholesterol ester modulators associated with prion inhibitors Synergism was obtained with the cholesterol ester modulators everolimus, pioglitazone, progesterone, and verapamil associated with the anti-prion chlorpromazine, and with everolimus and pioglitazone associated with the anti-prion quinacrine. Comparative lipid analyses in prion-infected and non-infected N2a cells by colorimetric, enzymatic, and chemical means, clearly demonstrated a derangement of type and distribution of cholesterol esters, free cholesterol, and triglycerides in the infected N2a cells. Although single-drug treatments influenced lipid syntheses, only the combined-drug treatments appeared to restore a lipid profile similar to that of untreated-uninfected cells. Conclusions. We conclude that the anti-prion synergistic effect of cholesterol ester modulators with the cholesterol metabolism interfering anti-prion drugs chlorpromazine and quinacrine may arise from the ability of combined drugs to re-establish the intracellular lipid profile of untreated-uninfected cells. Overall, these data suggest that inhibition of prion replication can be readily potentiated by combinatorial drug treatments, and that steps of cholesterol/cholesterol ester metabolism may represent suitable targets

Synergistic Proapoptotic Activity of Recombinant Trail Plus the AKT Inhibitor Perifosine in Acute Myelogenous Leukemia Cells

To potentiate the response of acute myelogenous leukemia (AML) cells to TNF-Related Apoptosis- Inducing Ligand (TRAIL) cytotoxicity, we have examined the efficacy of a combination with perifosine, a novel phosphatidylinositol 3-kinase (PI3K)/Akt signaling inhibitor. The rationale for using such a combination is that perifosine was recently described to increase TRAIL-R2 receptor expression and decrease the cellular FLICE-Inhibitory Protein (cFLIP) in human lung cancer cell lines. Perifosine and TRAIL both induced cell death by apoptosis in the THP-1 AML cell line, which is characterized by constitutive PI3K/Akt activation, but lacks functional p53. Perifosine, at concentrations below IC50, dephosphorylated Akt and increased TRAIL-R2 levels, as demonstrated by western blot, RT-PCR, and flow cytometric analysis. Perifosine also decreased the long isoform of cFLIP (cFLIP-L) and the X-linked Inhibitor of Apoptosis Protein (XIAP) expression. Perifosine and TRAIL synergized to activate caspase-8 and induce apoptosis, which was blocked by a caspase- 8 selective inhibitor. Upregulation of TRAIL-R2 expression was dependent on a protein kinase Cα/ c-Jun-NH2-kinase 2/c-Jun signaling pathway activated by perifosine through reactive oxygen species production. Perifosine synergized with TRAIL also in primary AML cells displaying constitutive activation of the Akt pathway, by inducing apoptosis, Akt dephosphorylation, TRAIL-R2 upregulation, cFLIP-L and XIAP downregulation, and c-Jun phosphorylation. The combined treatment negatively affected the clonogenic activity of CD34+ cells from AML patients. In contrast, CD34+ cells from healthy donors were resistant to perifosine and TRAIL treatment. Our findings suggest that the combination perifosine and TRAIL might offer a novel therapeutic strategy for AML. Originally published Cancer Research, Vol. 68, No. 22, Nov 200

Altered cholesterol ester cycle in ex vivo skin fibroblasts from Alzheimer patients

Recent studies in both animal and cell models of Alzheimer's disease (AD) indicated that sub-cellular cholesterol distribution seems to regulate amyloid-beta (A[beta]) generation in the brain. In particular, cholesterol-esters (CE), rather than total cholesterol levels, appear directly correlated with A[beta] production. Here we observed that, similarly to brain cells, skin fibroblasts obtained from AD patients produce and accumulate more CE than skin fibroblasts from age-matched healthy controls do. AD fibroblasts also exhibited a 2 fold increase in the expression of ACAT1, in addition to lower levels of SREBP2, nCEH, Caveolin-1 and ABCA1 mRNA levels, all of which are involved in the CE cycle. HMGCoA-reductase and LDL-receptor mRNAs levels did not show statistically significant changes in AD, compared to non-AD, cells. Furthermore, although APP mRNA did not significantly vary, neprilysin (NEP), the most important enzyme in the proteolysis of A[beta], was expressed at very low levels in skin fibroblasts of sporadic AD patients. Our results contribute to the concept that AD may be the consequence of a basic and systemic defect in the CE cycle. Moreover, our results identify new possible targets for the diagnosis, prevention, and cure or, at least, amelioration of the symptoms of AD

Визначення понять «резидент» і «нерезидент»: проблеми теорії і практики

Досліджено практику застосування понять «резидент» та «нерезидент» у як в Україні так і окремих зарубіжних країнах, а також надано авторське бачення щодо визначення даних термінів. Автори статті визначають умови оподаткування операцій з нерезидентами з огляду на існуючі особливості правозастосовної діяльності і вно­сять окремі пропозиції щодо удосконалення процедури адміністрування податків, що стягуються з нерезидентів.Исследована практика применения понятий «резидент» и «нерезидент» как в Украине, так и отдельных зарубежных странах, а также предоставлено авторское виденье относительно определения данных терминов. Авторы статьи определяют условия налогообложения операций с нерезидентами, учитывая существующие особен­ности правоприменительной деятельности, вносят отдельные предложения относи­тельно усовершенствования процедуры администрирования налогов, которые удержи­ваются из нерезидентов.Іn this article practice of application of concepts «resident» and «non­resident» are explored both in Ukraine and some foreign countries, and granted the author’s view on the definition of these terms. The authors of the article determine the term of taxation of operations with non­residents, taking into account the existing features of legal activity and make some suggestions in relation to the improvement of procedure of administration of taxes which are tightened from non­residents

Accumulation of neutral lipids in peripheral blood mononuclear cells as a distinctive trait of Alzheimer patients and asymptomatic subjects at risk of disease

<p>Abstract</p> <p>Background</p> <p>Alzheimer's disease is the most common progressive neurodegenerative disease. In recent years, numerous progresses in the discovery of novel Alzheimer's disease molecular biomarkers in brain as well as in biological fluids have been made. Among them, those involving lipid metabolism are emerging as potential candidates. In particular, an accumulation of neutral lipids was recently found by us in skin fibroblasts from Alzheimer's disease patients. Therefore, with the aim to assess whether peripheral alterations in cholesterol homeostasis might be relevant in Alzheimer's disease development and progression, in the present study we analyzed lipid metabolism in plasma and peripheral blood mononuclear cells from Alzheimer's disease patients and from their first-degree relatives.</p> <p>Methods</p> <p>Blood samples were obtained from 93 patients with probable Alzheimer's disease and from 91 of their first-degree relatives. As controls we utilized 57, cognitively normal, over-65 year-old volunteers and 113 blood donors aged 21-66 years, respectively. Data are reported as mean ± standard error. Statistical calculations were performed using the statistical analysis software Origin 8.0 version. Data analysis was done using the Student t-test and the Pearson test.</p> <p>Results</p> <p>Data reported here show high neutral lipid levels and increased ACAT-1 protein in about 85% of peripheral blood mononuclear cells freshly isolated (<it>ex vivo</it>) from patients with probable sporadic Alzheimer's disease compared to about 7% of cognitively normal age-matched controls. A significant reduction in high density lipoprotein-cholesterol levels in plasma from Alzheimer's disease blood samples was also observed. Additionally, correlation analyses reveal a negative correlation between high density lipoprotein-cholesterol and cognitive capacity, as determined by Mini Mental State Examination, as well as between high density lipoprotein-cholesterol and neutral lipid accumulation. We observed great variability in the neutral lipid-peripheral blood mononuclear cells data and in plasma lipid analysis of the subjects enrolled as Alzheimer's disease-first-degree relatives. However, about 30% of them tend to display a peripheral metabolic cholesterol pattern similar to that exhibited by Alzheimer's disease patients.</p> <p>Conclusion</p> <p>We suggest that neutral lipid-peripheral blood mononuclear cells and plasma high density lipoprotein-cholesterol determinations might be of interest to outline a distinctive metabolic profile applying to both Alzheimer's disease patients and asymptomatic subjects at higher risk of disease.</p