Platelet Apoptotic Response May Be Associated With the Capacity of Aspirin to Inhibit Platelets.

An inadequate platelet response to aspirin (ASA) has been identified in some patients under chronic ASA treatment. The aim of this study was to analyze if ASA-sensitive and ASA-resistant platelets have differences in their apoptotic capability. Clinically stable ischemic coronary patients who had been taking ASA (100 mg/d) for at least 9 months before inclusion were divided into ASA-resistant (n = 11) and ASA-sensitive (n = 13) groups as defined by the PFA-100 test. Platelets from ASA-sensitive patients showed higher expression of the proapoptotic proteins Bak and Bax than those from ASA-resistant patients, although only Bak protein remained different when the results were adjusted by age. In resting platelets, neither caspase-3 activity nor cytosolic cytochrome C levels were different between both experimental groups. Stimulation of platelets with calcium ionophore (10 nmol/L, A23187) increased caspase-3 activity (1.91-fold higher; P < 0.05) and cytosolic cytochrome C levels (1.84-fold higher; P < 0.05) to a higher degree in ASA-sensitive than in ASA-resistant platelets. In conclusion, ASA-sensitive platelets seem to be better prepared to undergo apoptosis during robust platelet activation.

New circulating biomarkers for predicting cardiovascular death in healthy population.

There is interest to analyse newer biomarkers to identify healthy individuals at risk to develop cardiovascular disease (CVD) incidents and death. To determine in healthy individuals new circulating protein biomarkers, whose systemic levels may be associated with the risk of future development of CVD incidents and death. The study was performed in 82 individuals from the Malmö Diet and Cancer study cohort, free from CVD of whom 41 developed CVD and 41 did not. Plasma proteins related to inflammation and thrombo-coagulating processes were analysed. α1-antitrypsin isotype 3 plasma levels were significantly higher while apolipoprotein J plasma levels were lower in participants that developed CVD incidents than those that did not develop acute cardiovascular episode. Of 82 participants, 17 died by CVD causes. There were proteins whose expression in plasma was significantly higher in participants suffering CVD death as compared with those that did not die by CVD. These proteins included: fibrinogen ß-chain isotypes 1 and 3, fibrinogen-γ-chain isotype 2, vitamin D-binding protein isotypes 1, 2 and 3, α1-antitrypsin isotypes 3 and 6, haptoglobin isotypes 3,4,5 and 5, haemopexin isotypes 1 and 2, and Rho/Rac guanine nucleotide exchange factor 2. Moreover, apolipoprotein J plasma levels were found lower in participants that died by cardiovascular cause. Association between plasma levels of proteins and CVD death was independent of age, gender, conventional risk factors and plasma C-reactive protein levels. Several protein plasma levels and protein isotypes related to inflammation and thrombo-coagulating phenomena were independently associated with the risk of future CVD death.

Nitric oxide from mononuclear cells may be involved in platelet responsiveness to aspirin.

BACKGROUND: Several mechanisms have been proposed to explain why some platelets have a reduced response to aspirin (ASA). Among them, it was reported an increased circulating level of vitamin-D-binding protein (DBP). In addition, nitric oxide (NO) released from mononuclear cells was involved in the antiplatelet effects of ASA. The aim was to analyse the relationship between platelet response to ASA and both NO generation and vitamin-D-binding protein content in mononuclear cells. MATERIALS AND METHODS: Mononuclear cells were obtained from patients with stable coronary artery disease that were divided by a platelet functionality test (PFA-100) as ASA-sensitive (n=23) and ASA resistant (n=27). RESULTS: Both the release of NO (determined by nitrite+nitrate concentration) and the expression of endothelial-type NO synthase (eNOS) were higher in mononuclear cells from ASA sensitive as compared with those from ASA-resistant patients. There was a positive correlation between either the release of NO and the expression of eNOS protein in mononuclear cells with the ability of ASA to inhibit platelet activity. DBP content in mononuclear cells was higher in ASA resistant than in ASA sensitive. The level of DBP content in mononuclear cells was negatively associated with the ability of ASA to inhibit platelets. However, in vitro experiments suggested that there was no association between DBP and NO production by mononuclear cells. CONCLUSIONS: Mononuclear cells from patients with platelets with lower responsiveness to ASA showed a reduced ability to produce NO.

Effects of factor Xa on the expression of proteins in femoral arteries from type 2 diabetic patients.

AIM: Further to its pivotal role in haemostasis, factor Xa (FXa) promotes effects on the vascular wall. The purpose of the study was to evaluate if FXa modifies the expression level of energy metabolism and oxidative stress-related proteins in femoral arteries obtained from type 2 diabetic patients with end-stage vasculopathy. METHODS: Femoral arteries were obtained from 12 type 2 diabetic patients who underwent leg amputation. Segments from the femoral arteries were incubated in vitro alone and in the presence of 25 nmol l(-1) FXa and 25 nmol l(-1) FXa + 50 nmol l(-1) rivaroxaban. RESULTS: In the femoral arteries, FXa increased triosephosphate isomerase and glyceraldehyde-3-phosphate dehydrogenase isotype 1 expression but decreased pyruvate dehydrogenase expression. These facts were accompanied by an increased content of acetyl-CoA. Aconitase activity was reduced in FXa-incubated femoral arteries as compared with control. Moreover, FXa increased the protein expression level of oxidative stress-related proteins which was accompanied by an increased malonyldialdehyde arterial content. The FXa inhibitor, rivaroxaban, failed to prevent the reduced expression of pyruvate dehydrogenase induced by FXa but reduced acetyl-CoA content and reverted the decreased aconitase activity observed with FXa alone. Rivaroxaban + FXa but not FXa alone increased the expression level of carnitine palmitoyltransferase I and II, two mitochondrial long chain fatty acid transporters. Rivaroxaban also prevented the increased expression of oxidative stress-related proteins induced by FXa alone. CONCLUSIONS: In femoral isolated arteries from type 2 diabetic patients with end-stage vasculopathy, FXa promoted disruption of the aerobic mitochondrial metabolism. Rivaroxaban prevented such effects and even seemed to favour long chain fatty acid transport into mitochondria.

Vardenafil improves penile erection in type 2 diabetes mellitus patients with erectile dysfunction: role of tropomyosin.

INTRODUCTION: Evidences have been suggested that phosphodiesterase type 5 (PDE5) inhibition promotes vasculoprotective benefits in patients with cardiovascular diseases. AIM: The aim of this study is to analyze the systemic effect of PDE5 inhibition in type 2 diabetes mellitus patients with erectile dysfunction (ED) determining changes in the expression levels of plasma proteins. METHODS: Seventeen patients with controlled type 2 diabetes mellitus and ED were included in the study. Patients received vardenafil hydrochloride 20 mg on demand during 12 weeks. At the beginning and 12 weeks after vardenafil administration, plasma samples were collected and analyzed using proteomics. MAIN OUTCOME MEASURES: International Index of Erectile Function-Erectile Function Domain (IIEF-EFD) and plasma protein expression before and after vardenafil administration. Nitrate/nitrite release, PDE5, and soluble guanylate cyclase (sGC) expression and cyclic guanosine monophosphate (cGMP) content in cultured bovine aortic endothelial cells (BAECs). RESULTS: The IIEF-EFD score was markedly improved after 12 weeks of vardenafil administration. Plasma levels of alpha 1-antitrypsin isotypes 4 and 6 and ß-tropomyosin were decreased, whereas apolipoprotein AI isoype 5 was increased 12 weeks after vardenafil administration. Only ß-tropomyosin plasma levels were inversely correlated with IIEF-EFD score. Tropomyosin has been added to cultured BAECs and after 24 hours reduced the protein expression level of sGC-ß1 subunit and decreased the cGMP content. Tropomyosin did not modify PDE5 expression and nitric oxide release in BAECs as compared with control BAECs. Vardenafil (10 µg/mL) did not modify sGC-ß1 subunit expression in tropomyosin + vardenafil-incubated BAECs; however, vardenafil significantly reversed the reduction of cGMP content induced by tropomyosin. CONCLUSION: Vardenafil administration improved erectile functionality in controlled type 2 diabetes mellitus patients with ED, which was associated with reduction of circulating plasma ß-tropomyosin levels. Tropomyosin affected by itself the cGMP generating system suggesting a possible new mechanism involved in ED. Vardenafil reversed the reduction effect of cGMP content elicited by tropomyosin in BAECs.

Changes in cardiac energy metabolic pathways in overweighed rats fed a high-fat diet.

BACKGROUND: Heart produces ATP through long-chain fatty acids beta oxidation. PURPOSE: To analyze whether in ventricular myocardium, high-fat diet may modify the expression of proteins associated with energy metabolism before myocardial function was affected. METHODS: Wistar Kyoto rats were divided into two groups: (a) rats fed standard diet (control; n = 6) and (b) rats fed high-fat diet (HFD; n = 6). Proteins from left ventricles were analyzed by two-dimensional electrophoresis, mass spectrometry and Western blotting. RESULTS: Rats fed with HFD showed higher body weight, insulin, glucose, leptin and total cholesterol plasma levels as compared with those fed with standard diet. However, myocardial functional parameters were not different between them. The protein expression of 3-ketoacyl-CoA thiolase, acyl-CoA hydrolase mitochondrial precursor and enoyl-CoA hydratase, three long-chain fatty acid ß-oxidation-related enzymes, and carnitine-O-palmitoyltransferase I was significantly higher in left ventricles from HFD rats. Protein expression of triosephosphate isomerase was higher in left ventricles from HFD rats than in those from control. Two α/ß-enolase isotypes and glyceraldehyde-3-phosphate isomerase were significantly increased in HFD rats as compared with control. Pyruvate and lactate contents were similar in HFD and control groups. Expression of proteins associated with Krebs cycle and mitochondrial oxidative phosphorylation was higher in HFD rats. CONCLUSIONS: Expression of proteins involved in left ventricle metabolic energy was enhanced before myocardial functionality was affected in rats fed with HFD. These findings may probably indicate higher cardiac energy requirement due to weight increase by HFD.

Plasma desmoplakin I biomarker of vascular recurrence after ischemic stroke.

Stroke patients have a high risk of vascular recurrence. Biomarkers related to vascular recurrence, however, remain to be identified. The aim of the study was to identify, through proteomic analysis, plasma biomarkers associated with vascular recurrence within one year after the first ischemic stroke. This is a substudy (n = 134) of a large prospective multicenter study of post-stroke patients with an ischemic stroke. Plasma samples were obtained at inclusion. Among the identified proteins, only plasma levels of desmoplakin I were associated with protection against a new vascular event (Odds ratio: 0.64; 95% CI: 0.46-0.89; p = 0.009) after adjustment for hypercholesterolemia, statins and previous atherothrombotic stroke subtype. A greater number of patients without vascular recurrence had been treated with statins within three months of the recent ischemic stroke. Only patients who had been taking statins for 3 months after the ischemic stroke and did not suffer vascular recurrence over a follow-up year, have higher levels of desmoplakin I at the time of inclusion (Odds ratio 0.49; 95% CI: 0.28-0.86; p = 0.013). Increased desmoplakin I levels, determined within 1-3 months of the first ischemic stroke, could be a biomarker for statin responsiveness against a new vascular event in post-ischemic stroke patients taking statins early (1-3 months) after the ischemic stroke.

Expression of cytoskeleton and energetic metabolism-related proteins at human abdominal aortic aneurysm sites.

OBJECTIVE: The purpose of this study was to evaluate the expression of proteins related to cytoskeleton and energetic metabolism at abdominal aortic aneurysm (AAA) sites using proteomics. Several remodeling-related mechanisms have been associated with AAA formation but less is known about the expression of proteins associated with cytoskeleton and energetic metabolism in AAAs. METHODS: AAA samples (6.73 ± 0.40 cm size) were obtained from 13 patients during elective aneurysm repair. Control abdominal aortic samples were obtained from 12 organ donors. Proteins were analyzed using two-dimensional electrophoresis and mass spectrometry. RESULTS: The expression of filamin was increased in the AAA site compared to control abdominal aortic samples while microfibril-associated glycoprotein-4 isotype 1, annexin A5 isotype 1, and annexin A2 were reduced compared with control abdominal aortic samples. Reduction in expression level of energetic metabolism-associated proteins such as triosephosphate isomerase, glyceraldehyde 3-phosphate dehydrogenase, and cytosolic aldehyde dehydrogenase was also observed in AAAs compared to controls. Reduction of triosephosphate isomerase expression was also observed by Western blot, which was accompanied by diminished triosephosphate isomerase activity. At the AAA site, pyruvate dehydrogenase expression was reduced and the content of both lactate and pyruvate was increased with respect to controls without changes in lactate dehydrogenase activity. CONCLUSIONS: The present results suggest that an anaerobic metabolic state may be favored further to reduce the expression of cytoskeleton-related proteins. The better knowledge of molecular mechanism involved in AAAs may favor development of new clinical strategies.

KCNH2 gene mutation: a potential link between epilepsy and long QT-2 syndrome.

Long QT syndrome (LQTS) is closely associated with syncope, seizure, and sudden death but LQTS is frequently misdiagnosed as epilepsy. LQTS and epilepsy both belong to the group of ion channelopathies that manifest in the heart and brain. Therefore, genetic analysis of genes associated with potassium and sodium homeostasis and electrical disorders may reveal a link between epilepsy and lethal cardiac arrhythmia. Here, the authors report a young woman who suffered recurrent seizure episodes and syncopes that occurred while walking and also during rest. She showed electroencephalogram abnormalities and a pathological prolonged QTc interval in electrocardiogram. The patient and the patient's asymptomatic family members underwent genetic screening of the three genes most frequently associated with LQTS: KCNQ1, KCNH2, and SCN5A. The patient and the family members did not show DNA alterations in the genes KCNQ1 and SCN5A associated with LQT-1 and LQT-3, respectively. However, the patient showed a de novo mutation 2587T→C in exon 10 of KCNH2 gene associated with LQT-2. The mutation caused a stop codon substitution (R863X) in the HERG channel, leading to a 296-amino acid deletion. The patient's asymptomatic relatives did not show the KCNH2 gene mutation. R863X alteration in HERG channel may be involved in both prolonged QTc interval and epilepsy. This fact raises the possibility that R863X alteration in KCNH2-encoded potassium channel may confer susceptibility for epilepsy and cardiac LQT-2 arrhythmia.

Spike Protein Subunits of SARS-CoV-2 Alter Mitochondrial Metabolism in Human Pulmonary Microvascular Endothelial Cells: Involvement of Factor Xa.

Background: Mitochondria have been involved in host defense upon viral infections. Factor Xa (FXa), a coagulating factor, may also have influence on mitochondrial functionalities. The aim was to analyze if in human pulmonary microvascular endothelial cells (HPMEC), the SARS-CoV-2 (COVID-19) spike protein subunits, S1 and S2 (S1+S2), could alter mitochondrial metabolism and what is the role of FXA. Methods: HPMEC were incubated with and without recombinants S1+S2 (10 nmol/L each). Results: In control conditions, S1+S2 failed to modify FXa expression. However, in LPS (1 µg/mL)-incubated HPMEC, S1+S2 significantly increased FXa production. LPS tended to reduce mitochondrial membrane potential with respect to control, but in higher and significant degree, it was reduced when S1+S2 were present. LPS did not significantly modify cytochrome c oxidase activity as compared with control. Addition of S1+S2 spike subunits to LPS-incubated HPMEC significantly increased cytochrome c oxidase activity with respect to control. Lactate dehydrogenase activity was also increased by S1+S2 with respect to control and LPS alone. Protein expression level of uncoupled protein-2 (UCP-2) was markedly expressed when S1+S2 were added together to LPS. Rivaroxaban (50 nmol/L), a specific FXa inhibitor, significantly reduced all the above-mentioned alterations induced by S1+S2 including UCP-2 expression. Conclusions: In HPMEC undergoing to preinflammatory condition, COVID-19 S1+S2 spike subunits promoted alterations in mitochondria metabolism suggesting a shift from aerobic towards anaerobic metabolism that was accompanied of high FXa production. Rivaroxaban prevented all the mitochondrial metabolic changes mediated by the present COVID-19 S1 and S2 spike subunits suggesting the involvement of endogenous FXa.

Proteomic changes related to "bewildered" circulating platelets in the acute coronary syndrome.

Acute coronary syndromes (ACS) are associated with platelet activation. The aim of the present study was to study the protein expression level associated with glycolysis, oxidative stress, cytoskeleton and cell survival in platelets obtained during an ACS. Platelets from 42 coronary ischemic patients, divided into patients admitted within 24 h after the onset of chest pain (ACS group; n=16) and patients with stable coronary ischemic disease (CAD, n=26), were analyzed using proteomics. The expression levels of proteins involved in cellular cytoskeleton (F-actin capping, ß-tubulin, α-tubulin isotypes 1 and 2, vinculin, vimentin and two Ras-related protein Rab-7b isotypes), glycolysis pathway (glyceraldehyde-3-phosphate dehydrogenase, lactate dehydrogenase and two pyruvate kinase isotypes) and cellular-related antioxidant system (manganese superoxide dismutase) and even the expression and activity of glutathione-S-transferase were significantly reduced in platelets from ACS patients compared to CAD patients. Moreover, reduction in the expression of proteins associated with cell survival such as proteasome subunit ß type 1 was also observed in ACS platelets compared with CAD platelets. Principal component and logistic regression analysis suggested the existence of factors (proteins) expressed in the platelets inversely associated with acute coronary ischemia. In summary, these results suggest the existence of circulating antioxidant, cytoskeleton and glycolytic-"bewildered" platelets during the acute phase of a coronary event.

Functional status and inflammation after preseason training program in professional and recreational soccer players: a proteomic approach.

The purpose of the study was to determine if an intensive pre- season training program modifies the inflammatory status in professional soccer players and if this inflammatory profile may be associated with the physical state. We compared plasma protein biomarkers, using proteomics, and the physiological state and cardiac function in 12 professional soccer players and 9 recreational soccer players. Reduced cardiac low frequency [LF] after the pre- season training program previous competition with respect to recreational soccer players was found. No differences were found in cardiac high frequency, cardiac high frequency/low frequency ratio, tension index and oxygen volume consumption. Alpha-1-antitrypsin isotype-3, fibrinogen-gamma isotypes-1, 2 and 3 and vitamin-D-binding protein isotype-1 were reduced in professionals players compared with those in recreational players. However, an increased content of alpha-1-antitrypsin isotype-6 and alpha-1-antichymotrypsin 1 and 4 were found in professional soccer players. Spearman's analysis showed a positive correlation between LF and fibrinogen-gamma chain isotype 3; but LF was negatively correlated with alpha-antichymotrypsin isotype 4. Professional soccer players submitted to an intensive training showed differences in the content of plasma proteins associated with inflammatory/oxidative stress and thrombosis with respect to recreational soccer players. Proteomics analysis in combination with the analysis of cardiac function assessment may be useful to know more in depth molecular processes associated with sport and intensive exercise. Key pointsProteomics allow us to find differences in the plasma protein content in sportsmen.Just after pre-season training program, professional soccer players showed lower content of circulating proteins associated with inflammation compared to recreational soccer players.Proteomic analysis in combination with the analysis of cardiac function may be useful to know more in depth molecular inflammatory and oxidative processes associated with the sport.

Clinical and Surgical Outcomes in Extensive Scalp Reconstruction after Oncologic Resection: A Comparison of Anterolateral Thigh, Latissimus Dorsi and Omental Free Flaps.

Microsurgical scalp reconstruction is indicated in patients with large scalp defects. The aim of this study was to compare the outcomes of scalp reconstruction in oncologic patients reconstructed with latissimus dorsi (LD), anterolateral thigh (ALT), and omental (OM) free flaps. Thirty oncologic patients underwent scalp reconstruction with LD (10), ALT (11), and OM (9) flaps. The length of the vascular pedicle, the operation time, the possibility of a two-team approach, the length of hospital stays, the complications, and the aesthetic results were evaluated. The OM flap was the flap with the shortest vascular pedicle length with a mean of 6.26 ± 0.16 cm, compared to the LD flap, which was 12.34 ± 0.55 cm and the ALT flap with 13.20 ± 0.26 cm (p < 0.05). The average time of surgery was 6.6 ± 0.14 h in patients reconstructed with OM, compared to the LD flap, which was 8.91 ± 0.32 h and the ALT flap with 7.53 ± 0.22 h (p < 0.05). A two-team approach was performed in all patients for OM flaps and ALT flaps, but only in two patients reconstructed with the LD flap (p < 0.001). In patients reconstructed with the OM flap, a very satisfactory or satisfactory result was reported in seven patients (77.8%). Eight patients reported a very unsatisfactory or unsatisfactory result with LD flap (80%) and 10 patients with ALT flap (90.9%) (p = 0.002). The mean hospital stay after surgery was not statistically significant (p > 0.05). As for complications, two patients reconstructed with OM flap, five LT flaps, and two ALT flaps developed complications, not statistically significant (p = 0.235). Omental flap, latissimus dorsi flap, and anterolateral thigh flap fulfill most of the characteristics for complex scalp reconstruction. The decision on which flap to use should be based on clinical aspects of the patients taking into account that the three flaps show similar rates of complications and length of hospital stay. Regarding the aesthetic outcome, OM flap or LD flap should be considered for reconstruction of extensive scalp defects.

Effects of platelets on the protein expression in aortic segments: A proteomic approach.

It is well known the effects of the vascular wall on platelet activity but little is known about the effects of platelets on the proteins expression in the vascular wall. We analyzed whether platelets may modify the protein expression in the vascular wall. We used an in vitro model coincubating human platelet rich plasma (PRP) with control and 10 ng/ml tumor necrosis factor-α (TNF-α)-preincubated bovine aortic segments. 2DE, mass spectrometry and Western blot analysis were used to determine changes in the expression of proteins associated with the cytoskeleton and energetic metabolism in the aortic segments. In control healthy vascular wall, only the cytoskeleton-related proteins expression was modified by PRP. However, when PRP was coincubated with TNF-α pre-stimulated aortic segments lesser number of cytoskeleton-related proteins were modified. With respect to energetic metabolism, in control segments, PRP failed to modify any of the analyzed energetic-related proteins. However, in TNF-α-preincubated segments the presence of PRP upexpressed glyceraldehyde-3-phosphate dehydrogenase. Moreover, by western blot experiments it was observed that in TNF-α-preincubated segments the expression of fructose 1,6-bisphosphate aldolase was downregulated by platelets. However, no differences were found in the expression of triosephosphate isomerase and ATP synthase α-chain. In addition, the activity of fructose 1,6-bisphosphate aldolase and piruvate content was significantly reduced without modification on triosephosphate isomerase activity. In conclusion, the crosstalk between platelets and vascular wall is bidirectional and platelets regulated in the vascular wall the expression of proteins associated with the cytoskeleton and energetic metabolism, particularly in the healthy vascular wall.

A proteomic approach to determine changes in proteins involved in the myocardial metabolism in left ventricles of spontaneously hypertensive rats.

BACKGROUND: Different works have suggested that in the hypertrophied heart the energy metabolic pathway shifts to glycolysis. Our aim was to evaluate using proteomics the expression of proteins associated with different energetic metabolic pathways in hypertrophied left ventricles of spontaneously hypertensive rats (SHR). METHODS: 24-weeks-old SHR with stable hypertension and established left ventricle hypertrophy were used. Normotensive Wistar Kyoto rats were used as control. Proteins from left ventricles were analyzed by 2-dimensional electrophoresis and identified by comparison with a virtual rat heart proteomic map and mass spectrometry. RESULTS: Enoyl-CoA hydratase expression, an enzyme involved in fatty acid beta-oxidation, was reduced whereas the expression of other beta-oxidation enzymes, 3-ketoacyl-CoA thiolase and the mitochondrial precursor of acyl-CoA thioester hydrolase, was increased in the hypertrophied left ventricles. The expression of two enzymes involved in the first steps of glycolysis, fructose bisphosphate aldolase and triosephosphate isomerase, was reduced in the left ventricle of SHR. Pyruvate dehydrogenase expression, enzyme involved in glucose oxidation, was enhanced in the hypertrophied ventricles whereas proteins of the tricarboxylic acid cycle were not modified. Proteins involved in the mitochondrial oxidative phosphorylation were overexpressed whereas the alpha-subunit of the mitochondrial precursor of ATP synthase was downexpressed. CONCLUSIONS: Several proteins involved in the main energy metabolic pathways were up and downexpressed. Moreover, our results seem to suggest that probably neither fatty acid beta-oxidation nor glycolysis are the only sources for energy in the hypertrophied left ventricle.

Comparative expression of proteins in left and right atrial appendages from patients with mitral valve disease at sinus rhythm and atrial fibrillation.

INTRODUCTION: The objective was to compare by proteomics the expression of proteins associated with the cytoskeleton, energetic metabolism, and cardiac cytoprotection between left atrial appendages (LAA) and right atrial appendages (RAA) obtained from patients with mitral valve disease both in sinus rhythm (SR, n = 6) and in permanent atrial fibrillation (AF, n = 11). METHODS AND RESULTS: Samples from RAA and LAA were obtained from the same patient. Proteins were separated in 2-dimensional electrophoresis and identified by mass spectrometry. LAA from SR patients upexpressed alpha-actin isotype 1 and desmin isotypes 3 and 5 with respect to RAA. In LAA from AF patients were upexpressed cardiac alpha-actin isotypes 1 and 2, tropomyosin alpha- and beta-chains, and myosin light chain embryonic muscle/atrial isoform with respect to LAA from SR patients. In RAA from AF patients also upexpressed different cytoskeleton associated proteins with respect to RAA from SR patients. Different energetic metabolism-associated proteins were upexpressed in LAA and RAA from AF with respect those from SR patients. In AF patients, the expression of proteins associated with cardiac cytoprotection such as gluthatione-S-transferase, heat shock protein (Hsp) 27, and different Hsp60 isotypes, were higher in RAA but not in LAA with respect to the corresponding appendages in SR patients. CONCLUSIONS: For each individual patient RAA and LAA showed a similar level of proteins expressed associated with cytoskeleton, energetic metabolism, and cardiac cytoprotection. There were more differences in the level of proteins associated with the above-mentioned mechanisms between the atrial appendages from AF with respect to SR patients, which may open new targets for drugs.

Different protein expression in normal and dysfunctional platelets from uremic patients.

Although many uremic patients show platelet dysfunctionality, there are others with normal platelet functionality and even with thrombotic tendencies. Our aim was to evaluate changes in the expression of proteins in functional and dysfunctional uremic platelets. Using the platelet function analyzer (PFA-100) assay, uremic patients were divided according to their platelet functionality into normal (n=7) and dysfunctional (n=8). There were no significant differences in the number of circulating platelets and hematocrit and hemoglobin levels. Two-dimensional electrophoresis and mass spectrometry were used to determine and identify changes in protein expression. The closure time (CT) in the PFA-100 assay was significantly prolonged in the dysfunctional uremic platelets. In the dysfunctional platelets, actin-interacting protein-1 isotype 1 was down-regulated, while integrin IIb was up-regulated. Glutathione-S-transferase isotypes 1 and 2 and peroxiredoxin VI were up-regulated in the dysfunctional platelets. Pearson analysis showed a negative correlation between the platelet expression of integrin IIb and creatinine clearance. A positive correlation was found between creatinine clearance and glutathione-S-transferase isotype 2. Serum uric acid concentration was positively correlated with CT values and glutathione-S-transferase isotype 1. In conclusion, the analysis of the protein expression in uremic platelets with normal and dysfunctional activity revealed differences which may occur at the megakaryocyte level.

Reduction from 2011 to 2017 in adherence to breast cancer screening and non-improvement in the uptake of cervical cancer screening among women living in Spain.

OBJECTIVES: To analyze the uptake of breast and cervical cancer screening according to the 2017 Spanish National Health Survey (SNHS), to compare uptake rates with those obtained in the previous SNHS 2011 and to identify predictors for the uptake for these two screening tests. STUDY DESIGN: Cross-sectional study. MAIN OUTCOME MEASURES: Uptake rates of breast cancer and cervical cancer screening were analyzed for women aged 40-69 and aged 25-65 years, respectively. Independent variables included sociodemographic characteristics and factors related to health status and lifestyle. RESULTS: We found that 66.8 % of women aged 40-69 years had undergone mammography in the previous two years. Positive predictors for mammography uptake were age (50-69 years); marital status (married); Spanish nationality; university education; one or more chronic diseases; no alcohol consumption; physical activity; body mass index <30 kg/m2; and not smoking. We observed that 73.0 % of women aged 25-65 years had undergone cervical cytology screening in the previous three years. Positive predictors for uptake were age (25-52 years); marital status (married); Spanish nationality; middle-high educational level; no chronic diseases; no alcohol consumption; physical activity; body mass index <30 kg/m2; and not smoking. There was a significant decrease in the uptake rate for breast cancer screening from the previous SNHS 2011 (OR 0.89; 95 % CI 0.83-0.94). CONCLUSIONS: The adherence rate for mammography in Spain in 2017 was below the recommended 70 % and was significantly lower than in 2011. The figures for cervical cancer screening were over 70 % and stable over time.

Relationship Between Erectile Dysfunction, Diabetes and Dyslipidemia in Hypertensive-Treated Men.

PURPOSE: PRESIDEN study is a large study to analyze the erectile dysfunction (ED) incidence in Spanish population. The present study is a pilot sub-analysis from PRESIDEN to determine if ED or plasma testosterone (TST) level in controlled hypertensive patients may be associated with comorbidities and/or plasma nitrite+nitrate and antioxidant capacity. MATERIALS AND METHODS: Forty-four hypertensive individuals were aleatory selected from PRESIDEN study, matching by age (28 showing ED and 16 without ED). RESULT: Diabetes was present in 28.57% of ED patients and in 18.75% of patients without ED. In patients with and without ED, increasing age showed tendency of higher frequency of an additional comorbidity (diabetes or dyslipemia) (P = .09). Apparently, plasma TST levels were lower in older ED patients compared to younger patients with and without ED, although it did not reach statistical significance (P = .69). Older ED patients also showed lower TST levels than older patients without ED, although it was not statistical significant (16.15 ± 2.84 vs 13.91± 2.77; P = .69). Dyslipidemia was showed by 52.17% with lower TST (? nmol/L) while 23.80% of patients with plasma TST levels > 15 nmol/L had dyslipidemia. The percentage of ED patients was similar between patients with low and high TST levels. CONCLUSION: More ED hypertensive patients seem to show two comorbidities (diabetes and dyslipidemia) than hypertensivepatients without ED. Younger patients with ED tended to show more commonly diabetes than older ED patients. Plasma TST levels were not associated with more prevalence of ED but lower plasma TST levels showed tendency to higher prevalence of dyslipidemia.

Effect of parathyroid-hormone-related protein on human platelet activation.

Evidence suggests that PTHrP [PTH (parathyroid hormone)-related protein] can act as an inflammatory mediator in several pathological settings including cardiovascular disease. The aim of the present study was to determine whether PTHrP might be involved in human platelet activation. We used a turbidimetric method to determine platelet aggregation. The expression of PTH1R (PTH type 1 receptor) in human platelets was analysed by Western blot and flow cytometry analyses. PTHrP-(1-36) (10(-7) mol/l) by itself failed to modify the activation of platelets. However, it significantly enhanced ADP-induced platelet activation, and also increased the ability of other agonists (thrombin, collagen and arachidonic acid) to induce platelet aggregation. H89 (10(-6) mol/l) and 25 x 10(-6) mol/l Rp-cAMPS (adenosine 3',5'-cyclic monophosphorothioate Rp-isomer), two protein kinase A inhibitors, and 25 x 10(-9) mol/l bisindolylmaleimide I, a protein kinase C inhibitor, partially decreased the enhancing effect of PTHrP-(1-36) on ADP-induced platelet activation. Meanwhile, 10(-6) mol/l PTHrP-(7-34), a PTH1R antagonist, as well as 10(-5) mol/l PD098059, a MAPK (mitogen-activated protein kinase) inhibitor, or a farnesyltransferase inhibitor abolished this effect of PTHrP-(1-36). Moreover, 10(-7) mol/l PTHrP-(1-36) increased (2-fold over control) MAPK activation in human platelets. PTH1R was detected in platelets, and the number of platelets expressing it on their surface in patients during AMI (acute myocardial infarction) was not different from that in a group of patients with similar cardiovascular risk factors without AMI. Western blot analysis showed that total PTH1R protein levels were markedly higher in platelets from control than those from AMI patients. PTH1R was found in plasma, where its levels were increased in AMI patients compared with controls. In conclusion, human platelets express the PTH1R. PTHrP can interact with this receptor to enhance human platelet activation induced by several agonists through a MAPK-dependent mechanism.