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Migraine has been associated with an increased risk for stroke, especially in young women.1 Among the pathophysiologic mechanisms linking migraine and stroke, coagulation abnormalities have been regarded as a logical link.2

Background. Normalization of arterial pressure occurs in just a few patients with hypertensive chronic kidney disease undergoing kidney transplantation. Hypertension in kidney transplant recipients may be related to multiple factors. We aimed to assess whether hypertension in kidney-transplanted patients may be linked to reinnervation of renal arteries of the transplanted kidney. Methods. We investigated renal arteries innervation from native and transplanted kidneys in three patients 5 months, 2 years and 11 years after transplantation, respectively. Four transplanted kidneys from non-hypertensive patients on immunosuppressive treatment without evidence of hypertensive arteriolar damage were used as controls. Results . Evidence of nerve sprouting was observed as early as 5 months following transplantation, probably originated from ganglions of recipient patient located near the arterial anastomosis and was associated with mild hypertensive arteriolar damage. Regeneration of periadventitial nerves was already complete 2 years after transplantation. Nerve density tended to reach values observed in native kidney arteries and was associated with hypertension-related arteriolar lesions in transplanted kidneys. Control kidneys, albeit on an immunosuppressive regimen, presented only a modest regeneration of sympathetic nerves. Conclusions . Our results suggest that the considerable increase in sympathetic nerves, as found in patients with severe arterial damage, may be correlated to hypertension rather than to immunosuppressive therapy, thus providing a morphological basis for hypertension recurrence despite renal denervation.

Atherosclerosis is an inflammatory chronic disease affecting arterial vessels and leading to vascular diseases, such as stroke and myocardial infarction. The relationship between atherosclerosis and risk of neurodegeneration has been established, in particular with vascular cognitive impairment and dementia (VCID). Systemic atherosclerosis increases the risk of VCID by inducing cerebral infarction, or through systemic or local inflammatory factors that underlie both atherosclerosis and cognition. Omega-3 and omega-6 polyunsaturated fatty acids (PUFAs) are involved in inflammatory processes, but with opposite roles. Specifically, omega-3 PUFAs exert anti-inflammatory properties by competing with omega-6 PUFAs and displacing arachidonic acid in membrane phospholipids, decreasing the production of pro-inflammatory eicosanoids. Experimental studies and some clinical trials have demonstrated that omega-3 PUFA supplementation may reduce the risk of different phenotypes of atherosclerosis and cardiovascular disease. This review describes the link between atherosclerosis, VCID and inflammation, as well as how omega-3 PUFA supplementation may be useful to prevent and treat inflammatory-related diseases.

Metabolic diseases are chronic disorders correlated to a greater risk of cardiovascular event and death. Recently, many data have sustained the biological link between microvascular dysfunction, oxidative stress, vascular inflammation, and metabolic diseases. The determination of new and specific blood biomarkers of vascular inflammation associated with obesity-related metabolic syndrome (MetS) and diabetes such as lipoprotein-associated phospholipase A2 (Lp-PLA2) could be useful to identify subject with high risk of cardiovascular events. Lp-PLA2 participates by a crucial role in microvascular dysfunction and oxidative stress showing positive association with metabolic disorders. In this review, we will argue the evolving role of Lp-PLA2 in predicting cardiovascular events in metabolic disease patients.

Sympathetic nervous system activity is increased after cardiopulmonary arrest, resulting in vasoconstrictor release from the perivascular sympathetic nerves of cerebral arteries. However, the pathophysiological function of the perivascular sympathetic nerves in the ischemic brain remains unclear. A rat model of global cerebral ischemia (asphyxial cardiac arrest, ACA) was used to investigate perivascular sympathetic nerves of cerebral arteries via bilateral decentralization (preganglionic lesion) of the superior cervical ganglion (SCG). Decentralization of the SCG 5 days before ACA alleviated hypoperfusion and afforded hippocampal neuroprotection and improved functional outcomes. These studies can provide further insights into the functional mechanism(s) of the sympathetic nervous system during ischemia. NEW & NOTEWORTHY Interruption of the perivascular sympathetic nerves can alleviate CA-induced hypoperfusion and neuronal cell death in the CA1 region of the hippocampus to enhance functional learning and memory.

BACKGROUND AND AIM: Several measurements were taken for frailty classification in geriatric population. "Frailty index" is based on "deficits in health," but it is still not available in Italian version. Thus, the aim of the present work was to validate a version of "frailty index" for the Italian geriatric community. METHODS: The validation of Italian frailty index (IFi) is based on a cohort study that enrolled 1077 non-disabled outpatients aged 65 years or older (81.3 ± 6.5 years) in Naples (Italy). IFi has been expressed as a ratio of deficits present/deficits considered after a comprehensive geriatric assessment. IFi was stratified in light, moderate and severe frailty. Mortality, disability (considering an increase in ADL lost ≥1 from the baseline) and hospitalization were considered at 3, 6, 12, 18 and 24 months of follow-up. Area under curve (AUC) was evaluated for both Fried's and IFi frailty index. RESULT: At the end of follow-up, mortality increased from 1.0 to 30.3%, disability from 40.9 to 92.3% and hospitalization from 0.0 to 59.0% (p < 0.001 for trend). Multivariate analysis shows that the relative risk for unit increase in IFi is 1.09 (95% CI = 1.01-1.17, p = 0.013) for mortality, 1.04 (95% CI = 1.01-1.06, p = 0.024) for disability and 1.03 (95% CI = 1.01-1.07, p = 0.041) for hospitalization. AUC is higher in IFi with respect to Fried's frailty index when considering mortality (0.809 vs. 0.658, respectively), disability (0.800 vs. 0.729, respectively) and hospitalization (0.707 vs. 0.646, respectively). CONCLUSIONS: IFi is a valid measure of frailty after the comprehensive geriatric assessment in an Italian cohort of non-institutionalized patients.

SummaryThe effect of gut microbiota on obesity and insulin resistance is now recognized, but the underlying host-dependent mechanisms remain poorly undefined. We find that tissue inhibitor of metalloproteinase 3 knockout (Timp3−/−) mice fed a high-fat diet exhibit gut microbiota dysbiosis, an increase in branched chain and aromatic (BCAA) metabolites, liver steatosis, and an increase in circulating soluble IL-6 receptors (sIL6Rs). sIL6Rs can then activate inflammatory cells, such as CD11c+ cells, which drive metabolic inflammation. Depleting the microbiota through antibiotic treatment significantly improves glucose tolerance, hepatic steatosis, and systemic inflammation, and neutralizing sIL6R signaling reduces inflammation, but only mildly impacts glucose tolerance. Collectively, our results suggest that gut microbiota is the primary driver of the observed metabolic dysfunction, which is mediated, in part, through IL-6 signaling. Our findings also identify an important role for Timp3 in mediating the effect of the microbiota in metabolic diseases.

Permanent AF is characterized by an increased mortality in elderly subjects with CHF. Moreover, AF increased the risk of mortality also in elderly subjects without CHF. Thus, we examined long-term mortality in community-dwelling elderly people with and without CHF. A total of 1332 subjects aged 65 and older were selected from the electoral rolls of Campania, a region of southern Italy. The relationship between AF and mortality during a 12-year follow-up in 125 subjects with CHF and in 1.143 subjects without CHF were studied. Elderly subjects showed a higher mortality in those with respect to those without AF (72.1% vs. 51.8%; p< 0.01). Similarly, elderly subjects without CHF showed a higher mortality in those with respect to those without AF (61.8% vs. 49.8%; p< 0.05). In contrast, elderly subjects with CHF showed a similar mortality in those with respect to those without AF (74.7% vs. 82.4%; p=. 0.234). Multivariate analysis shows that AF was predictive of mortality in all elderly subjects (Hazard Risk = HR = 1.39, 95% confidence interval (CI) = 1.25-2.82; p< 0.001). When the analysis was conducted considering the presence and the absence of CHF, AF was strongly predictive of mortality in elderly subjects without CHF (HR = 1.95, 95%CI = 1.25-4.51; p< 0.001) but not in those with CHF (HR = 1.12, 95%CI = 0.97-3.69; p=. 0.321). We concluded that AF is able to predict long-term mortality in elderly subjects. Moreover, AF is strongly predictive of long-term mortality in the absence but not in the presence of CHF. © 2011 Elsevier Ireland Ltd.

Background and Purpose— Homocysteine levels are determined by genetic and environmental factors. Several studies have linked high plasma levels of total homocysteine to the increased risk of cardiovascular disease, stroke, and many other conditions. However, the exact mechanism of documented and novel total homocysteine quantitative trait loci to that risk is unknown. Methods— We have performed linkage analysis in 100 high-risk Dominican families with 1362 members. Probands were selected from the population-based Northern Manhattan Study. A set of 405 microsatellite markers was used to screen the whole genome. Variance components analysis was used to detect evidence for linkage after adjusting for stroke risk factors. Ordered-subset analysis based on Dominican Republic enrollment was conducted. Results— Total homocysteine levels had a heritability of 0.44 ( P P =0.0005) with a peak at D17S2193 and was significantly increased in a subset of families with a high proportion of Dominican Republic enrollment (MLOD=3.92, P =0.0022). Additionally, modest evidence for linkage was found at chromosome 2p21 (MLOD=1.77, P =0.0033) with a peak at D2S1356 and was significantly increased in a subset of families with a low proportion of Dominican Republic enrollment (MLOD=2.82, P =0.0097). Conclusions— We found a strong evidence for novel quantitative trait loci on chromosomes 2 and 17 for total homocysteine plasma levels in Dominican families. Our family study provides essential data for a better understanding of the genetic mechanisms associated with elevated total homocysteine levels leading to cardiovascular disease after accounting for environmental risk factors.

Background Several studies suggest transient ischemic attack (TIA) may be neuroprotective against ischemic stroke analogous to preinfarction angina's protection against acute myocardial infarction. However, this protective ischemic preconditioning-like effect may not be present in all ages, especially among the elderly. The purpose of this study was to determine the neuroprotective effect of TIAs (clinical equivalent of cerebral ischemic preconditioning) to neurologic damage after cerebral ischemic injury in patients over 65 years of age. Methods We reviewed the medical charts of patients with ischemic stroke for presence of TIAs within 72 hours before stroke onset. Stroke severity was evaluated by the National Institutes of Health Stroke Scale and disability by a modified Rankin scale. Results We evaluated 203 patients (≥65 years) with diagnosis of acute ischemic stroke and categorized them according to the presence (n = 42, 21%) or absence (n = 161, 79%) of TIAs within 72 hours of stroke onset. Patients were monitored until discharged from the hospital (length of hospital stay 14.5 ± 4.8 days). No significant differences in the National Institutes of Health Stroke Scale and modified Rankin scale scores were observed between those patients with TIAs and those without TIAs present before stroke onset at admission or discharge. Conclusion These results suggest that the neuroprotective mechanism of cerebral ischemic preconditioning may not be present or functional in the elderly.

Cardiovascular pathologies in the French are not prevalent despite high dietary saturated fat consumption. This is commonly referred to as the "French Paradox" attributing its anti-lipidemic effects to moderate consumption of red wine. Resveratrol, a phytoalexin found in red wine, is currently the focus of intense research both in the cardiovascular system and the brain. Current research suggests resveratrol may enhance prognosis of neurological disorders such as, Parkinson's, Huntington's, Alzheimer's diseases and stroke. The beneficial effects of resveratrol include: antioxidation, free radical scavenger, and modulation of neuronal energy homeostasis and glutamatergic receptors/ion channels. Resveratrol directly increases sirtuin 1 (SIRT1) activity, a NAD(+) (oxidized form of nicotinamide adenine dinucleotide)-dependent histone deacetylase related to increased lifespan in various species similar to calorie restriction. We recently demonstrated that brief resveratrol pretreatment conferred neuroprotection against cerebral ischemia via SIRT1 activation. This neuroprotective effect produced by resveratrol was similar to ischemic preconditioning-induced neuroprotection, which protects against lethal ischemic insults in the brain and other organ systems. Inhibition of SIRT1 abolished ischemic preconditioning-induced neuroprotection in CA1 region of the hippocampus. Since resveratrol and ischemic preconditioning-induced neuroprotection require activation of SIRT1, this common signaling pathway may provide targeted therapeutic treatment modalities as it relates to stroke and other brain pathologies. In this review, we will examine common signaling pathways, cellular targets of resveratrol, and ischemic preconditioning-induced neuroprotection as it relates to the brain.

Mortality related to heart rate (HR) increase in the elderly has not yet been well established. To ascertain the relationships among cognitive impairment (CI), mortality, and HR increase, the authors prospectively studied a random sample of elderly subjects stratified according to presence or absence of CI. Elderly subjects randomly selected in 1991 (n = 1332) were followed up for 12 years. Mortality was established in 98.1% of the subjects. When HR was stratified in quartiles (69, 70-75, 76-80, and80 bpm), mortality was linearly associated with increased HR in all (from 47.7 to 57.0; r2 = .43, p = .019) and in subjects without (from 41.7 to 51.1%; r2 = .50, p = .043) but not in those with CI (from 57.5 to 66.1; r2 = .20, p = .363). Cox regression analysis, adjusted for several variables, shows that HR doesn't predict mortality in all subjects (RR 0.69; 95% CI = 0.27-1.73) or in those with CI (RR 0.91; 95% CI = 0.81-1.02). In contrast, HR predicts mortality in subjects without CI (RR 1.10; 95% CI = 1.00-1.22). Hence, HR increase is a predictor of mortality in elderly subjects without CI. However, when considering all elderly subjects and those with CI, HR increase seems to have no effect on mortality. Thus, CI should be considered when focusing on HR increase as risk factor for mortality in the elderly.

Atherosclerosis is a chronic inflammatory disease characterized by the infiltration of pro-inflammatory macrophages into a lipid-laden plaque. ITCH is an E3 ubiquitin ligase that has been shown to polarize macrophages to an anti-inflammatory phenotype. We therefore investigated the effect of ITCH deficiency on the development of atherosclerosis. ApoE−/−ITCH−/− mice fed a western diet for 12 weeks showed increased circulating M2 macrophages together with a reduction in plaque formation. Bone marrow transplantation recreated the haemopoietic phenotype of increased circulating M2 macrophages but failed to affect plaque development. Intriguingly, the loss of ITCH lead to a reduction in circulating cholesterol levels through interference with nuclear SREBP2 clearance. This resulted in increased LDL reuptake through upregulation of LDL receptor expression. Furthermore, ApoE−/−ITCH−/− mice exhibit reduced hepatic steatosis, increased mitochondrial oxidative capacity and an increased reliance on fatty acids as energy source. We found that ITCH ubiquitinates SIRT6, leading to its breakdown and thus promoting hepatic lipid infiltration through reduced fatty acid oxidation. The E3 Ubiquitin Ligase ITCH modulates lipid metabolism impacting on atherosclerosis progression independently from effects on myeloid cells polarization through control of SIRT6 and SREBP2 ubiquitination. Thus, modulation of ITCH may provide a target for the treatment of hypercholesterolemia and hyperlipidemia.

Background and aim Molecular adsorbent recycling system (MARS) treatment is able to remove both hydrosoluble and small- and medium-sized lipophilic toxins. MARS plays an important role in modifying liver failure complications, such as hepatorenal syndrome and hepatic encephalopathy. We sought to evaluate the clinical efficacy and safety of a MARS device in a consecutive series of hepatic failure patients. Materials Twenty patients with acute liver failure, transplantation failure, or acute on chronic liver failure fulfilled the inclusion criteria of total bilirubin ≥10 mg/dL and at least one of the following: hepatic encephalopathy (HE) ≥II grade, hepatorenal syndrome (HRS) for chronic patients or total bilirubin ≥5 mg/dL and HE ≥I grade for acute patients. Results MARS was able to reduce cholestatic parameters and improve neurologic status and renal function parameters in all treated patients. We also observed an improvement in the 3-month survival rate compared to the expected outcome in patients with MELD scores between 20 and 29, as well as 30 and 39. Conclusions Based on these results, we confirm the safety and clinical efficacy of MARS treatment, with the best results in patients with MELD score of 20 to 29. Further studies are necessary to confirm whether this treatment is able to modify patient outcomes and prognosis.

Ischemic preconditioning (IP) has been proposed as an endogenous form of protection against ischemia reperfusion injury. IP, however, does not prevent post-ischemic dysfunction in the aging heart but may be partially corrected by exercise training and food restriction. We investigated the role of exercise training combined with food restriction on restoring IP in the aging heart. Effects of IP against ischemia-reperfusion injury in isolated hearts from adult (A, 6 months old), sedentary 'ad libitum' fed (SL), trained ad libitum fed (TL), sedentary food-restricted (SR), trained- and food-restricted senescent rats (TR) (24 months old) were investigated. Norepinephrine release in coronary effluent was determined by high performance liquid cromatography. IP significantly improved final recovery of percent developed pressure in hearts from A (p

The interleukin 7 receptor alpha-chain (IL-7Ralpha) is essential for T cell development in both humans and mice and for B cell development in mice. Whereas the transcription factor PU.1 regulates IL-7Ralpha expression in mouse pro-B cells via a GGAA motif, we demonstrate here that GA binding protein (GABP) bound to this site and was essential in the regulation of IL-7Ralpha expression in T cells, where PU.1 is not expressed. Moreover, IL-7Ralpha expression was diminished substantially in thymocytes but was normal on B220(+) fetal liver cells from mouse embryos with diminished expression of GABPalpha. Thus, GABP is essential for the regulation of IL-7Ralpha expression in T cells, and the differential regulation of IL-7Ralpha in distinct lymphoid lineages is achieved at least in part by differential recruitment of factors to the same GGAA motif.

Sirtuins are NAD⁺-dependent deacetylases, lipoamidases, and ADP-ribosyltransferases that link cellular metabolism to multiple intracellular pathways that influence processes as diverse as cell survival, longevity, and cancer growth. Sirtuins influence the extent of neuronal death in stroke. However, different sirtuins appear to have opposite roles in neuronal protection. In Caenorhabditis elegans, we found that knock-out of mitochondrial sirtuin sir-2.3, homologous to mammalian SIRT4, is protective in both chemical ischemia and hyperactive channel induced necrosis. Furthermore, the protective effect of sir-2.3 knock-out is enhanced by block of glycolysis and eliminated by a null mutation in daf-16/FOXO transcription factor, supporting the involvement of the insulin/IGF pathway. However, data in Caenorhabditis elegans cell culture suggest that the effects of sir-2.3 knock-out act downstream of the DAF-2/IGF-1 receptor. Analysis of ROS in sir-2.3 knock-out reveals that ROS become elevated in this mutant under ischemic conditions in dietary deprivation (DD), but to a lesser extent than in wild type, suggesting more robust activation of a ROS scavenging system in this mutant in the absence of food. This work suggests a deleterious role of SIRT4 during ischemic processes in mammals that must be further investigated and reveals a novel pathway that can be targeted for the design of therapies aimed at protecting neurons from death in ischemic conditions., Author summary Sirtuins are NAD+-dependent enzymes that play key roles in neuronal death. However, the contribution of sirtuins to this cellular process is controversial with results supporting that activation or inhibition are protective depending on the cellular context and type of sirtuin. In this manuscript we report that knock-out of Caenorhabditis elegans mitochondrial sirtuin 2.3, homolog of mammalian SIRT4, spares neurons from ischemic neuronal death and that this protective effect is enhanced by block of glycolysis. Furthermore, we report that ROS is eliminated more efficiently in sir-2.3 knock-out as compared to wild type, under dietary deprivation. We conclude that inhibition of mammalian SIRT4 combined with dietary deprivation must be explored as a novel therapeutic approach to prevent ischemic damage.

Background and Purpose— Few studies have examined the early effects of statins on carotid artery elasticity, a potential surrogate marker of cardiovascular risk. This study examined the short-term effects of atorvastatin 80 mg daily on carotid elasticity measured by high-resolution B-mode ultrasound. Methods— The study included 40 stroke-free and statin-naive subjects older than age 45 (mean age, 70±7 years; 55% men; 64% Caribbean-Hispanic). Outcome measures included carotid stiffness indices at 14 and 30 days after initiation of treatment. The systolic and diastolic diameters of the right common carotid artery were averaged from multiple B-mode imaging frames. Absolute and relative changes of strain [(systolic diameter−diastolic diameter)/diastolic diameter], stiffness (β) [ln (systolic/diastolic blood pressure)/strain], and distensibility (1/β adjusted for wall thickness) from baseline were compared by the repeated measures t test and were considered significant at α=0.05. Results— Baseline mean stiffness was 0.08 (95% confidence interval [CI], 0.06–0.10). It significantly decreased at day 30 to 0.05 (CI, 0.04–0.06; P P Conclusions— Short-term treatment with high-dose atorvastatin was associated with improvement in the carotid elasticity metrics. Carotid artery elasticity measured by B-mode ultrasound is a simple noninvasive measure of arterial wall function and may be a useful surrogate end point in clinical trials targeting individuals at increased risk for atherosclerosis.

Background Chronic heart failure (CHF) is characterized by a high mortality in the elderly. Moreover, depression status is also related to poor prognosis in advancing age. Thus, we sought to determine whether depressive status predicts long-term mortality in subject with or without CHF. Methods and results Long-term mortality after 12-year follow-up in 125 elderly subjects with CHF and 1143 elderly subjects without CHF was studied. Depression was evaluated using the Geriatric Depression Scale (GDS), and all subjects were stratified in tertiles according to GDS score (0–10, 11–20 and 21–30). With increasing GDS score, long-term mortality at the end of follow-up increased from 43Æ4% to 72Æ0% in subjects without (P < 0Æ001) and from 52Æ6% to 83Æ9% in subjects with (P < 0Æ007) CHF. In multivariate analysis, GDS appeared to be predictive of long-term mortality in the absence (Hazard ratio = 1Æ01; confidence interval 95% 1Æ00–1Æ05; P = 0Æ04) and, even more, in the presence of CHF (Hazard ratio = 1Æ08; confidence interval 95% 1Æ01–1Æ15; P < 0Æ01). Conclusions Depression symptoms predict long-term mortality in elderly subjects without and, even more, with CHF. Thus, depression can be considered a strong predictor of death in CHF elderly subjects.

Background and aims: Elderly subjects are characterized by a high prevalence of chronic obstructive pulmonary disease (COPD) and frailty. This study examined the predictive role of frailty on long-term mortality in elderly subjects with and without COPD. Methods: The study assessed mortality after a 12-year follow-up in 489 subjects with COPD and 799 subjects without COPD, selected in 1992. Frailty was assessed according to the Frailty Staging System scores ranging from 1 to 7. Results: After 12 years’ follow-up, mortality was 48.1% in subjects without and 60.7% in subjects with COPD (p

Race/ethnic differences in carotid arterial function and structure exist among those with cerebrovascular disease, but whether differences persist among healthy populations is unknown. Our objective was to investigate differences in carotid artery diameter and stiffness between race/ethnic groups, and examine whether these race/ethnic differences were age-dependent.Carotid diameters were assessed by B-mode ultrasound among 1536 participants from the Northern Manhattan Study (NOMAS), and carotid stiffness metrics were calculated. We used multivariable linear regression models to determine the relationship between race/ethnicity and both carotid arterial stiffness and carotid diastolic diameter.Mean participant age was 70 ± 9 years (Hispanics = 68 ± 8, blacks = 72 ± 9, and whites = 74 ± 9, p0.0001). Mean DDIAM was 6.2 ± 1.0mm (Hispanics = 6.2 ± 0.9 mm, blacks = 6.3 ± 1.0 mm, and whites = 6.3 ± 1.0 mm, p0.005) and mean STIFF was 8.7 ± 6.3 (Hispanics = 8.5 ± 5.7, blacks = 9.2 ± 6.2 and whites = 8.9 ± 6.9, p0.02). In a model that adjusted for sociodemographics and vascular risk factors including hypertension, diabetes, dislipidemia, renal function, physical acticity and a history of known coronary artery diseases; age was positively associated with greater DDIAM in Hispanics (p0.0001) but not among blacks or whites. Older age was associated with greater stiffness among Hispanics (p0.0001) and blacks (p0.003), but not among whites.We found race/ethnic differences in the association between age and arterial stiffness and diameter, including age-dependent arterial dilation observed in Hispanics that was not observed among blacks or whites.

Preanalytical variables, including the anticoagulants and stabilizing agents, time, storage temperature, and methods of DNA extraction applied to blood samples, may affect quality and quantity of isolated nucleic acids for future genomic applications. Considering the large number of collected samples, standard operating procedure optimization for whole blood preservation before DNA extraction is a crucial step in a biological repository. Moreover, the future application of the biological material may not be known subsequent to its extraction. To define standard operating procedures for whole blood preservation before DNA extraction, we aimed to determine whether different combinations of anticoagulants, blood storage temperatures, and time intervals before storage at -80°C might have an impact on quality and quantity of subsequent extracted DNA. After spectrophotometer quantification, the quality and integrity of DNA were assessed by agarose gel electrophoresis, polymerase chain reaction, and real-time polymerase chain reaction methods. We observed that decrease in DNA recovery during blood storage time was more pronounced at room temperature than at 4°C. Based on our experience, we recommend as anticoagulants of choice sodium citrate and ethylenediaminetetraacetate, whereas sodium citrate theophylline adenosine dipyridamole could represent an alternative choice, excluding a priori lithium heparin and Fluoride-Oxalate. Based on the overall evaluation criteria, we conclude that the procedures necessary to preserve the whole blood before the DNA extraction may have a significant impact on downstream molecular biological applications.

Background Left ventricular mass (LVM) is an important risk factor for cardiovascular disease. Previously we found evidence for linkage to chromosome 12p11 in Dominican families, with a significant increase in a subset of families with high average waist circumference (WC). In the present study, we use association analysis to further study the genetic effect on LVM. Methods Association analysis with LVM was done in the one LOD critical region of the linkage peak in an independent sample of 897 Caribbean Hispanics. Genotype data were available on 7085 SNPs from 23 to 53 MB on chromosome 12p11. Adjustment was made for vascular risk factors and population substructure using an additive genetic model. Subset analysis by WC was performed to test for a difference in genetic effects between the high and low WC subsets. Results In the overall analysis, the most significant association was found to rs10743465, downstream of the SOX5 gene (p = 1.27E-05). Also, 19 additional SNPs had nominal p < 0.001. In the subset analysis, the most significant difference in genetic effect between those with high and low WC occurred with rs1157480 (p = 1.37E-04 for the difference in β coefficients), located upstream of TMTC1. Twelve additional SNPs in or near 6 genes had p < 0.001. Conclusions The current study supports previously identified evidence by linkage for a genetic effect on LVM on chromosome 12p11 using association analysis in population-based Caribbean Hispanic cohort. SOX5 may play an important role in the regulation of LVM. An interaction of TMTC1 with abdominal obesity may contribute to phenotypic variation of LVM.

Protein kinase C (PKC) is a family of serine/threonine-isozymes that are involved in many signaling events in normal and disease states. Previous studies from our lab have demonstrated that ɛPKC plays a pivotal role in neuroprotection induced by ischemic preconditioning. However, the role of ɛPKC during and after brain ischemia is not clearly defined. Therefore, in the present study, we tested the hypothesis that activation of ɛPKC during an ischemic event is neuroprotective. Furthermore, other studies have demonstrated that ɛPKC mediates cerebral ischemic tolerance in the rat brain by decreasing vascular tone. Thus, we also tested the effects of ɛPKC activation during ischemia on cerebral blood flow (CBF). We found that ψɛ-Receptors for Activated C Kinase (RACK), a ɛPKC-selective peptide activator, injected intravenously 30 min before induction of global cerebral ischemia conferred neuroprotection in the CA1 region of the rat hippocampus. Moreover, measurements of CBF before, during, and after cerebral ischemia revealed a significant reduction in the reperfusion phase of rats pretreated with ψɛRACK as compared to Tat peptide (vehicle). Our results suggest that ɛPKC can protect the rat brain against ischemic damage by regulating CBF. Thus, ɛPKC may be one of the treatment modalities against ischemic injury.

International audience; Cell-based therapies for global cerebral ischemia represent promising approaches for neuronal damage prevention and tissue repair promotion. We examined the potential of marrow-isolated adult multilineage-inducible (MIAMI) cells, a homogeneous subpopulation of immature human mesenchymal stromal cell, injected into the hippocampus to prevent neuronal damage induced by global ischemia using rat organotypic hippocampal slices exposed to oxygen-glucose deprivation and rats subjected to asphyxial cardiac arrest. We next examined the value of combining fibronectin-coated biomimetic microcarriers (FN-BMMs) with epidermal growth factor (EGF)/basic fibroblast growth factor (bFGF) pre-treated MIAMI compared to EGF/bFGF pre-treated MIAMI cells alone, for their in vitro and in vivo neuroprotective capacity. Naive and EGF/bFGF pre-treated MIAMI cells significantly protected the Cornu Ammonis layer 1 (CA1) against ischemic death in hippocampal slices and increased CA1 survival in rats. MIAMI cells therapeutic value was significantly increased when delivering the cells complexed with FN-BMMs, probably by increasing stem cell survival and paracrine secretion of pro-survival and/or anti-inflammatory molecules as concluded from survival, differentiation and gene expression analysis. Four days after oxygen and glucose deprivation and asphyxial cardiac arrest, few transplanted cells administered alone survived in the brain whereas stem cell survival improved when injected complexed with FN-BMMs. Interestingly, a large fraction of the transplanted cells administered alone or in complexes expressed betaIII-tubulin suggesting that partial neuronal transdifferentiation may be a contributing factor to the neuroprotective mechanism of MIAMI cells.

Background aims. Procedures for cardiomyocyte differentiation of mouse embryonic stem cells (mESCs) utilize different amounts of serum. Because the serum composition is unknown, unambiguous characterization of the differentiation process is biased. All reported serum-free protocols used for compound testing provide serum throughout the differentiation process. We report on an embryoid body (EB)-based procedure for cardiomyocyte differentiation of mESCs in which serum is provided only in the earliest step (hanging drop, 0--2 days). Methods. To assess cardiomyocyte differentiation, we generated an mESCs clone that expressed green fluorescence protein (GFP) under the control of the myosin light chain 2v (MLC2v) promoter. To define the lowest serum concentration required for efficient induction of cardiomyocyte differentiation, EBs were formed in presence of 5% (S5), 10% (S10) and 15% (S15) serum until day 2, then switched to a serum-free medium. Results. Analysis of cardiac-specific transcripts on day 6 of differentiation showed that 10% (S10) was the minimum amount of serum for efficient continuation of cultures under serum-free conditions. Spontaneously beating foci were detected in 90.0 +/-+/- 5.5% of S10 EBs on day 7 of differentiation, and cardiomyocyte markers were expressed from day 8 of differentiation (MLC2v-driven GFP; alpha alpha-myosin heavy chain). Dose--response curves to isoproterenol showed that the beating rate increased by 113.0 +/-+/- 39.4%, with a concentration for half-maximal effect (EC(50)) of 25.7 nM. Conclusions. The development of functional cardiomyocytes from mESCs is not affected by serum withdrawal after EBs formation. This culture system represents a new model for cardiomyocyte differentiation of mESCs to assess the effects of compounds on the process of cardiomyogenesis.

Objective Sirtuins (SIRTs) and mitochondrial uncoupling proteins (UCPs) have been implicated in cardiovascular diseases through the control of reactive oxygen species production. This study sought to investigate the association between genetic variants in the SIRT and UCP genes and carotid plaque. Methods In a group of 1018 stroke-free subjects from the Northern Manhattan Study with high-definition carotid ultrasonography and genotyping, we investigated the associations of 85 single nucleotide polymorphisms (SNPs) in the 11 SIRT and UCP genes with the presence and number of carotid plaques, and evaluated interactions of SNPs with sex, smoking, diabetes and hypertension as well as interactions between SNPs significantly associated with carotid plaque. Results Overall, 60% of subjects had carotid plaques. After adjustment for demographic and vascular risk factors, T-carriers of the SIRT6 SNP rs107251 had an increased risk for carotid plaque (odds ratio, OR = 1.71, 95% CI = 1.23–2.37, Bonferroni-corrected p = 0.03) and for a number of plaques (rate ratio, RR = 1.31, 1.18–1.45, Bonferroni-corrected p = 1.4×10−5), whereas T-carriers of the UCP5 SNP rs5977238 had an decreased risk for carotid plaque (OR = 0.49, 95% CI = 0.32–0.74, Bonferroni-corrected p = 0.02) and plaque number (RR = 0.64, 95% CI = 0.52–0.78, Bonferroni-corrected p = 4.9×10−4). Some interactions with a nominal p≤0.01 were found between sex and SNPs in the UCP1 and UCP3 gene; between smoking, diabetes, hypertension and SNPs in UCP5 and SIRT5; and between SNPs in the UCP5 gene and the UCP1, SIRT1, SIRT3, SIRT5, and SIRT6 genes in association with plaque phenotypes. Conclusion We observed significant associations between genetic variants in the SIRT6 and UCP5 genes and atherosclerotic plaque. We also found potential effect modifications by sex, smoking and vascular risk factors of the SIRT/UCP genes in the associations with atherosclerotic plaque. Further studies are needed to validate our observations.

Cerebral ischemia causes blood flow derangements characterized by hyperemia (increased cerebral blood flow, CBF) and subsequent hypoperfusion (decreased CBF). We previously demonstrated that protein kinase C delta (δPKC) plays an important role in hippocampal neuronal death after ischemia. However, whether part of this protection is due to the role of δPKC on CBF following cerebral ischemia remains poorly understood. We hypothesized that δPKC exacerbates hyperemia and subsequent hypoperfusion resulting in CBF derangements following ischemia. Sprague-Dawley (SD) rats pretreated with a δPKC specific inhibitor (δV1-1, 0.5 mg/kg) exhibited attenuation of hyperemia and latent hypoperfusion characterized by vasoconstriction followed by vasodilation of microvessels after 2-vessel occlusion plus hypotension measured by 2-photon microscopy. In an asphyxial cardiac arrest model (ACA), SD rats treated with δV1-1 (pre- and post-ischemia) exhibited improved perfusion after 24 hrs and less hippocampal CA1 neuronal death 7 days after ACA. These results suggest possible therapeutic potential of δPKC in modulating CBF and neuronal damage after cerebral ischemia.

Background Arterial stiffness, an intermediate pre-clinical marker of atherosclerosis, has been associated with an increased risk of stroke and cardiovascular disease. The metabolic syndrome and its components are established cardiovascular disease risk factors and may also increase arterial stiffness; however, data regarding this are limited. Aim The goal of this study was to determine the association between the metabolic syndrome and carotid artery stiffness in an elderly multi-ethnic cohort. Methods Carotid artery stiffness was assessed by carotid ultrasound as part of the Northern Manhattan Study, a prospective population-based cohort of stroke-free individuals. Carotid artery stiffness was calculated as [In (systolic BP/diastolic BP)/strain], where strain was [(systolic diameter diastolic diameter)/diastolic diameter]. Metabolic syndrome was defined by the National Cholesterol Education Program: Adult Treatment Panel III criteria. LogSTIFF was analysed as the dependent variable in linear regression models, adjusting for demographics, education, current smoking, presence of carotid plaque and intima-media thickness. Results Carotid artery stiffness was analysed in 1133 Northern Manhattan Study subjects (mean age 65 ± 9 years; 61% women; 58% Hispanic, 22% Black and 20% Caucasian). The prevalence of the metabolic syndrome was 49%. The mean LogSTIFF was 2·01±0·61 among those with the metabolic syndrome and 1·90 ± 0·59 among those without the metabolic syndrome ( P = 0·003). The metabolic syndrome was significantly associated with increased logSTIFF in the final adjusted model (parameter estimate β = 0·100, P = 0·01). Among individual metabolic syndrome components, waist circumference and elevated blood pressure were most significantly associated with a mean increase in logSTIFF ( P < 0·01). Conclusion The metabolic syndrome is significantly associated with increased carotid artery stiffness in a multiethnic population. Increased carotid artery stiffness may, in part, explain a high risk of stroke among individuals with the metabolic syndrome.

Large doses of omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are used to treat several diseases including hypertriglyceridemia in humans. Modest levels of EPA and DHA may be obtained from food, particularly from fatty fish. This review presents the literature examining the differences between omega-3 fatty acid dietary supplementation and prescribed omega-3-acid ethyl esters (P-OM3). Reports published between 1995 and 2007 containing sources, recommended intake, and differences in the various formulations of omega-3 fatty acids were sought in PubMed and the Food and Drug Administration (FDA) Websites. However, lack of head-to-head clinical trials using both P-OM3 and dietary-supplement omega-3 fatty acids is the greatest limitation of this review. Although many kinds of omega-3 fatty acid dietary supplements are available, the efficacy, quality, and safety of these products are questionable because they are beyond any pharmaceutical control. Thus, P-OM3 is the only FDA approved omega-3 fatty acid product which is available in the United States as an adjunct to diet to improve human health.

A mild stress such as brief ischemic episodes may protect the heart from a successive and more prolonged myocardial ischemia (ischemic preconditioning). This phenomenon is considered a typical “hormetic mechanism” by which the heart is immunized from pathological insults such as myocardial ischemia. This mechanism is reduced with aging and it may be restored and/or preserved by drugs such as adenosine or nicorandil, a mitochondrial KATP channels, and lifestyle interventions such as physical activity and/or hypocaloric diet. Moreover, since the mechanisms involved in cardiac ischemic preconditioning have been established basic and clinical investigators are encouraged to test several drug in well-controlled animal and human studies in order to prevent and/or restore the age-related reduction of ischemic preconditioning.

Delayed neuroprotection against ischemic challenges is conferred by both ischemic preconditioning (IPC) and preconditioning by activation of the ε-isoform of protein kinase C (εPKC-PC). In vivo, ischemic preconditioning enhances GABA release and ameliorates glutamate release during lethal cerebral ischemia. We tested the hypothesis that IPC and εPKC-PC confer neuroprotection by GABA synapses in rat organotypic hippocampal slices. Ischemic preconditioning or εPKC-PC was induced with 15 mins oxygen-glucose deprivation (OGD) or ψεRACK, a selective εPKC activator; and test ischemia consisted of 40 mins OGD. At the time of peak neuroprotection (48 h after preconditioning), we recorded GABAA receptor-mediated miniature postsynaptic currents (GABA mPSCs) in vulnerable CA1 pyramidal neurons using whole-cell voltage clamp techniques. The frequency and amplitude of GABA mPSCs significantly increased 48 h after IPC. In contrast, εPKC-PC enhanced only the amplitude of GABA mPSCs with no effect on frequency. We next asked if neuroprotection depended on these changes in GABA synapses. Weak antagonism of the GABAA receptor with bicuculline (100 nmol/L) decreased the amplitude of GABA mPSCs by 20.9±6.1%. When applied during test ischemia, 100nmol/L bicuculline abolished neuroprotection conferred by either IPC or εPKC-PC. We conclude that neuroprotection conferred by preconditioning depends on functional modifications of GABA synapses.

Background Preinfarction angina (PrA), clinical equivalent of ischemic preconditioning, confers protection against inhospital mortality for acute myocardial infarction (AMI) in adult but not in elderly patients. This study aims to examine the interaction between physical activity and body mass index (BMI) in preserving the cardioprotective effect of PrA in elderly patients with AMI. Design Elderly patients (Z65 years old) with AMI admitted to Coronary Care Unit. Methods Elderly patients with AMI were retrospectively stratified for the presence and absence of PrA, and for quartiles of BMI and physical activity. In-hospital outcomes (death, cardiogenic shock, and reinfarction and creatine kinase-MB peak) were evaluated. Results In-hospital mortality of 1014 elderly patients with AMI was 19.2% in those with PrA and 22.7% in those without (P = 0.18, NS). Mortality further decreased with increased physical activity and reduced BMI, a trend that was not observed in patients without PrA. When physical activity and BMI were considered together, lowest in-hospital mortality was observed in patients with highest physical activity and normal BMI (from 18.2 to 9.6%; P < 0.01) with the greatest reduction observed in patients with PrA (from 18.3 to 5.1%; P =0.02). Multivariate analysis showed that PrA did not exert a protective effect in all patients irrespective of physical activity and BMI. A protective role was, however, observed in patients with highest physical activity or normal BMI and reached a maximum protective role in patients who showed both highest physical activity and normal BMI [odds ratio =0.08; 95% confidence interval = 0.02–0.72; P < 0.01]. Conclusion The cardioprotective effect of PrA was preserved in elderly patients who showed the highest physical activity and a normal BMI.

Background: comorbidity plays a critical role in the high mortality for chronic heart failure (CHF) in the elderly. Charlson Comorbidity Index (CCI) is the most extensively studied comorbidity index. No studies are available on the ability of CCI to predict mortality in CHF elderly subjects. The aim of the present study was to assess if CCI was able to predict long-term mortality in a random sample of elderly CHF subjects. Methods: long-term mortality after 12-year follow-up in 125 subjects with CHF and 1,143 subjects without CHF was studied. Comorbidity was evaluated using CCI. Findings: in elderly subjects stratified for CCI (1‐3 and ≥4), mortality was higher in non-CHF subjects with CCI ≥4 (52.4% versus 70%, P < 0.002) but not in those with CHF (75.9% versus 77.6%, P = 0.498, NS). Cox regression analysis on 12 years mortality indicated that both CCI (HR = 1.15; 95% CI = 1.01‐1.31; P = 0.035) and CHF (HR = 1.27; 95% CI = 1.04‐8.83; P = 0.003) were predictive of mortality. When Cox analysis was performed by selecting the presence and the absence of CHF, CCI was predictive of mortality in the absence but not in the presence of CHF. Conclusion: CCI does not predict long-term mortality in elderly subjects with CHF.

Gaucher disease is the most common lysosomal storage disease. It is caused by the defective activity of acid β-glucosidase, which results in the accumulation of lipid glucocerebroside in macrophages throughout the body. In this case report we describe the case of a young adult woman with splenomegaly as the primary manifestation of this pathology. This is a case of type 1 Gaucher disease because there is a lack of primary neurological involvement but we have, instead, an age-independent involvement of the visceral organs. It is very important to classify or characterize these patients in a precise manner and to make a complete diagnosis with the help of the many diagnostic resources now at our disposal, especially with genetics, radiology and new techniques of advanced microscopy, also because Gaucher disease requires a long and complex management from early life to adulthood.