Francesca Scarpini

Constitutive activation of the Ras/MAP kinase pathway and enhanced TCR signaling by targeting the Shc adaptor to membrane rafts.

The Shc adaptor is responsible for coupling receptor tyrosine kinases and tyrosine kinase-associated receptors to the Ras/MAP kinase pathway. Shc is believed to be regulated by a change in subcellular localization from the cytosol to the plasma membrane, where it recruits Grb-2/Sos complexes and hence permits juxtaposition of the guanine nucleotide exchange factor Sos to Ras, resulting in GDP/GTP exchange and Ras activation. Shc has been recently shown to inducibly colocalize in detergent-resistant membrane rafts together with the activated TCR and associated signaling molecules. To understand whether Shc localization in membrane rafts is sufficient to regulate Shc function, we constructed a Shc chimera containing the Ras membrane localization motif at the C-terminus. We show that membrane targeted Shc was constitutively localized in the plasma membrane of T-cells, and was mostly compartmentalized in lipid rafts. Membrane targeted Shc was phosphorylated on tyrosine residues and bound Grb-2/Sos in the absence of TCR engagement. Furthermore, expression of membrane targeted Shc resulted in constitutive downstream signaling, including Erk2 activation and enhancement of TCR dependent activation of the TCR responsive transcription factor NF-AT. Hence localization of Shc in membrane rafts is sufficient for Shc to acquire a signaling competent state. Interestingly, a membrane targeted Shc mutant lacking both Grb-2 binding sites was not only incapable of signaling in the absence of TCR triggering, but transdominantly inhibited endogenous Shc, supporting a non redundant role for Shc in the activation of the Ras/MAP kinase pathway in T-cells.

Role of Genetic Factors in Statins Side-Effects

Statins are relevant drugs involved in the reduction of cardiovascular events both in primary and secondary prevention. Related muscular side-effects are the most common cause of withdrawal and statins discontinuation could induce a negative rebound effect in terms of vascular events. Among factors in association with statins side-effects the combination with other drugs and the female sex are established conditions. However recent data suggest a specific genetic influence in intolerance development, at least for some statins. Indeed a genome-wide study in patients treated with simvastatin found an impressive association between single-nucleotide polymorphisms (SNPs) located within SLCO1B1 gene on chromosome 12 and established myopathy. Furthermore, the association between the SLCO1B1*5 variant and side-effects was found also in patients treated with atorvastatin but not, apparently, with pravastatin and categorized as carriers of mild-myopathy. Recently a similar evidence has been suggested in type 2 diabetic patients treated mainly with simvastatin. However another relevant issue is that, apart from genetic influence in liver transporters influencing drug levels, the complexity of mechanisms involved in the muscular side effects of statins has been addressed by the evidence of other influencing pathways such as the variant within the COQ2 gene involved in Coenzyme Q(10) mild-asymptomatic deficiency and skeletal muscle drug transporters expression. In conclusion, the picture of putative pharmacogenetic modulation of statins safety is reaching a growing body of evidence for translation into clinical practice but more specific studies for each single statin, in different clinical settings, both from genome-wide or competitive candidate genes evaluation, are needed before describing a definitive class-risk profile.

Statins discontinuation in compliant chronic users induces atherothrombotic profile despite baseline clinical setting and treatments.

Statins offer important benefits for the large populations of individuals at high risk for cardiovascular and cerebrovascular events in both primary and secondary preventions [1]. Due to the positive clinical impact, the concern of statins withdrawal on putative rebound vascular events has been suggested and evaluated both in terms of biological mechanisms or clinical settings as extensively reviewed [2– 3]. Furthermore, although from retrospective analysis, several observations have confirmed an increased events rate especially when statins were withdrawn during acute phase of acute vascular stress like coronary syndromes [4–5], major vascular surgery [6–7] and ischemic stroke [8], also in a randomized study [9]. Still unsolved is whether statin withdrawal syndrome does exist directly or could be related mainly to the specific atherothrombotic risk related to the behaviour of baseline disease as suggested by the discordant data from the retrospective analysis of the TNT study in patients with stable coronary disease [10]. The aim of the present study was to evaluate the putative effects of statins withdrawal due to several causes on atherothrombotic and inflammatory activation as well as lipid profile in subjects in primary prevention for cardiovascular risk on established statin therapy for at least one year. The patients population consisted of ninety-six subjects treated for familial dyslipidaemia (group A); one-hundred-forty subjects treated for dyslipidaemia and hypertension (group B) and/or clinical atherosclerosis (including ASA 100 mg/day, group C) [11–12] that were evaluated at seven (T7), fourteen (T14), thirty (T30) and sixty (T60) days after statin withdrawal for lipid profile including oxidized-LDL (ox-LDL); hs-CRP, soluble CD40 ligand (sCD40L), platelet P-selectin (pCD62+) and endogenous thrombin potential (ETP). Patients gave written informed consent to blood collection after ethic committee approval according to the guidelines of ICJME as for a previous protocol [13–14]. Fifty healthy subjects matched for gender and age were evaluated as controls for measurable parameters. Total cholesterol, LDLC, high-density lipoprotein cholesterol (HDL-C), triglycerides, ox-LDL, hsCRP andpCD62+expressionwere evaluated as previously described [14]. Apo-A and Apo-B were determined by immunonephelometry

Decreased plasma endogenous soluble RAGE, and enhanced adipokine secretion, oxidative stress and platelet/coagulative activation identify non-alcoholic fatty liver disease among patients with familial combined hyperlipidemia and/or metabolic syndrome

OBJECTIVE In patients with familial combined hyperlipidemia (FCHL), without metabolic syndrome (MS), occurrence of non-alcoholic fatty liver disease (NAFLD) is related to a specific pro-inflammatory profile, influenced by genetic traits, involved in oxidative stress and adipokine secretion. Among FCHL or MS patients, hyperactivity of the ligand-receptor for advanced glycation-end-products (RAGE) pathway, as reflected by inadequate protective response by the endogenous secretory (es)RAGE, in concert with genetic predisposition, may identify those with NAFLD even before and regardless of MS. METHODS We cross-sectionally compared 60 patients with vs. 50 without NAFLD. Each group included patients with FCHL alone, MS alone, and FCHL plus MS. RESULTS NAFLD patients had significantly lower plasma esRAGE, IL-10 and adiponectin, and higher CD40 ligand, endogenous thrombin potential and oxidized LDL. The effects of MS plus FCHL were additive. The genotypic cluster including LOX-1 IVS4-14A plus ADIPO 45GG and 256 GT/GG plus IL-10 10-1082G, together with higher esRAGE levels highly discriminate FCHL and MS patients not developing NAFLD. CONCLUSIONS Among FCHL or MS patients, noncarriers of the protective genotypic cluster, with lower esRAGE and higher degree of atherothrombotic abnormalities coincide with the diagnosis of NAFLD. This suggests an interplay between genotype, adipokine secretion, oxidative stress and platelet/coagulative activation, accelerating NAFLD occurrence as a proxy for cardiovascular disease.

Neurohumoral improvement and torsional dynamics in patients with heart failure after treatment with levosimendan

Background Levosimendan improves clinical and hemodynamic parameters exerting an anti-inflammatory and antiapoptotic effect in decompensated heart failure. The aim of this study was to evaluate the effects of levosimendan on LV torsion, plasma levels of NT-proBNP and on the balance between pro-inflammatory (TNF-α, IL-6) and anti-inflammatory cytokines (IL-10). Methods We enrolled 24 patients (age 62 ± 7 years) with acute HF, NYHA class IV and severe LV dysfunction. All patients underwent transthoracic echocardiography using two-dimensional speckle tracking analysis to detect LV twist angle (LVTA), at baseline and 1 week after treatment with levosimendan infusion. Biochemical parameters (pro-BNP, IL-6, IL-10, TNF-α) were determined by enzyme-linked immunosorbent (ELISA). Results After one week, we observed an improvement in LV function especially in LVTA (4.15 ± 2.54 vs 2.9 ± 2.1 p < 0.01), in LV ejection fraction (27.3 ± 8.04 vs 21.6 ± 6.8 p = 0.03) and also a significant reduction in BNP levels (1844 ± 560 vs 4713 ± 1050, p = 0.03). The multiple linear regression analysis showed a significant relation between a reduction of TNF α/IL-10 ratio (Δ > 20%) and BNP (Δ > 40%), LVEF (Δ > 10%) and LVTA (Δ > 20%) (O.R. 1.77, 95% C.I. 1.11–2.83; O.R. 1.49, 95% C.I. 1.08–2.67; O.R. 1.66, 95% C.I. 1.10–2.74, respectively, confirmed p, all < 0.01 by Hosmer–Lemeshov confirmation and the formal test for interaction). Conclusions Levosimendan exerts a hemodynamic effect by improving EF and LV torsion in patients with acute HF in association with a positive effect on the balance between pro and anti-inflammatory cytokines.

Genetic Influence in Statin Intolerance

To the Editor: We read with great interest the report titled “Common Nonsynonymous Substitutions in SLCO1B1 Predispose to Statin Intolerance in Routinely Treated Individuals With Type 2 Diabetes,” by Donnelly and co-workers, which describes the association between single-nucleotide polymorphisms located within the SLCO1B1 gene and predisposition to statin intolerance in subjects with diabetes.1 The relevance of SLCO1B1 polymorphisms in statin-induced myopathy was described in the genomewide SEARCH study in the context of severe myopathy, especially in subjects treated with simvastatin 80 mg.2 There is partially confirmed evidence of the role of the SLCO1B1*5 variant in inducing statin intolerance in patients treated with atorvastatin, whereas there is no such confirmatory evidence with respect to the role of the SLCO1B1*5 variant in patients treated with pravastatin who have mild myopathy.3 The data presented by Donnelly et al.1 have relevance in the context of the dosage of simvastatin used2 and the criteria applied in defining intolerance resulting in myopathy. These criteria provide a better fit with the conditions in actual clinical practice as compared to the criteria proposed in SEARCH. However, in our opinion, the apparent role of the SLCO1B1 rs4149056 polymorphism in statin-induced myopathy should be evaluated cautiously for two main reasons. First, individuals with diabetes are at risk of statin-induced myopathy regardless of genetic factors.4 Second, as pointed out by the authors, the greater prevalence of simvastatintreated subjects enables the collection of sufficient data to suggest specific roles for the studied genetic traits in inducing intolerance to simvastatin.1 However, different patterns could occur with respect to other molecules, as has been observed previously.3 We are currently conducting a study aimed at assessing the potential role of specific genetic traits and other environmental variables in muscle and liver intolerance to statins, as defined in accordance with the description presented by Donnelly and co-workers.1,5 In particular, we have observed that patients treated with rosuvastatin were more likely to show statin-induced myopathy if they were carriers of polymorphisms within the COQ2 gene (rs4693075), which induces CoQ10 pathway modifications (another putative mechanism of statin-induced myopathy), regardless of the presence of polymorphisms in SLCO1B1.5 SLCO1B1 variants were also confirmed to have a role in atorvastatin-related myopathy;3,5 however, evidence of even more severe intolerance was observed in the simultaneous presence of SLCO1B1 and COQ2 polymorphisms.5 In conclusion, we find a growing body of evidence for the putative pharmacogenetic modulations that impact the safety and efficacy of statins. Some of the findings show promise of being translatable into clinical practice. However, further specific studies for each statin in different clinical settings are needed, involving both genomewide and competitive candidate gene evaluation, before one can describe a reliable drug class–intolerance risk profile.

Genetic influence in liver steatosis prevalence and proatherothrombotic/inflammatory profile in familial combined hyperlipoproteinemia

presentation data could potentially have affected our results. Our results were also limited to in-hospital outcomes, but benefits of reduced DTB may emerge later, with a reduction in heart failure and a later mortality. In conclusion, our study found no correlation between short DTB or time to presentation and in-hospital mortality. Further investigation is needed to prove that short DTB does not have an impact onmortality in Japanese STEMI patients.

Safety of long-term simvastatin discontinuation

1 WHO. Recommendations for the management of common childhood conditions with limited resources: newborn conditions, dysentery, pneumonia, oxygen use and delivery, common causes of fever, severe acute malnutrition and supportive care. http://www.who.int/ maternal_child_adolescent/documents/ management_childhood_conditions/en/ index.html (accessed April 24, 2012). 2 Kallander K, Hildenwall H, Waiswa P, Galiwango E, Peterson S, Pariyo G. Delayed care seeking for fatal pneumonia in children aged under fi ve years in Uganda: a case-series study. Bull World Health Organ 2008; 86: 332–38. 3 Sazawal S, Black RE. Eff ect of pneumonia case management on mortality in neonates, infants, and preschool children: a meta-analysis of community-based trials. Lancet Infect Dis 2003; 3: 547–56. 4 UNICEF, WHO. WHO/UNICEF joint statement on management of pneumonia in community settings. http://www.who.int/maternal_child_ adolescent/documents/fch_cah_04_06/en/ index.html (accessed April 17, 2012). 5 Marsh DR, Gilroy KE, Van de Weerdt R, Wansi E, Qazi S. Community case management of pneumonia: at a tipping point? Bull World Health Organ 2008; 86: 381–89. contradicted by various observations in real-world clinical settings. Furthermore, statin with drawal could induce a proathero thrombotic state even in patients treated with other antiathero throm botic agents and cause a related increase in LDL cholesterol. What we cannot tell from the paper is how many patients, if any, have stopped simvastatin, and what their characteristics are in terms of cardio vascular risk profi le, LDL cholesterol target, and associated treatments for the observational period. This information is essential because the present message could induce some readers to consider that dis continuation of simvastatin treatment after about 5 years gives vascular protection even in the subsequent years in all patients, and this is probably not the case. The HPS Collaborative Group should at least clearly state that this observation fi ts with the studied population of patients who were probably very close to the LDL target for their cardiovascular risk even after simvastatin discontinuation.