Pharmacological inhibition of DNA methylation attenuates pressure overload-induced cardiac hypertrophy in rats.

BACKGROUND: Heart failure is associated with altered gene expression and DNA methylation. De novo DNA methylation is associated with gene silencing, but its role in cardiac pathology remains incompletely understood. We hypothesized that inhibition of DNA methyltransferases (DNMT) might prevent the deregulation of gene expression and the deterioration of cardiac function under pressure overload (PO). To test this hypothesis, we evaluated a DNMT inhibitor in PO in rats and analysed DNA methylation in cardiomyocytes. METHODS AND RESULTS: Young male Wistar rats were subjected to PO by transverse aortic constriction (TAC) or to sham surgery. Rats from both groups received solvent or 12.5 mg/kg body weight of the non-nucleosidic DNMT inhibitor RG108, initiated on the day of the intervention. After 4 weeks, we analysed cardiac function by MRI, fibrosis with Sirius Red staining, gene expression by RNA sequencing and qPCR, and DNA methylation by reduced representation bisulphite sequencing (RRBS). RG108 attenuated the ~70% increase in heart weight/body weight ratio of TAC over sham to 47% over sham, partially rescued reduced contractility, diminished the fibrotic response and the downregulation of a set of genes including Atp2a2 (SERCA2a) and Adrb1 (beta1-adrenoceptor). RG108 was associated with significantly lower global DNA methylation in cardiomyocytes by ~2%. The differentially methylated pathways were "cardiac hypertrophy", "cell death" and "xenobiotic metabolism signalling". Among these, "cardiac hypertrophy" was associated with significant methylation differences in the group comparison sham vs. TAC, but not significant between sham+RG108 and TAC+RG108 treatment, suggesting that RG108 partially prevented differential methylation. However, when comparing TAC and TAC+RG108, the pathway cardiac hypertrophy was not significantly differentially methylated. CONCLUSIONS: DNMT inhibitor treatment is associated with attenuation of cardiac hypertrophy and moderate changes in cardiomyocyte DNA methylation. The potential mechanistic link between these two effects and the role of non-myocytes need further clarification.

An alternative renin isoform is cardioprotective by modulating mitochondrial metabolism.

The renin-angiotensin system promotes oxidative stress, apoptosis, necrosis, fibrosis, and thus heart failure. Secretory renin plays a central role in these processes, initiating the generation of angiotensins. Nevertheless, alternative renin transcripts exist, which code for a cytosolically localized renin isoform (cyto-renin) that is cardioprotective. We tested the hypothesis that the protective effects are associated with a beneficial switch of metabolic and mitochondrial functions. To assess H9c2 cell mitochondrial parameters, we used the Seahorse XF analyser. Cardiac H9c2 cells overexpressing cyto-renin exhibited enhanced nonmitochondrial oxygen consumption, lactate accumulation, and LDH activity, reflecting a switch to more aerobic glycolysis known as Warburg effect. Additionally, mitochondrial spare capacity and cell respiratory control ratio were enhanced, indicating an increased potential to tolerate stress conditions. Renin knockdown induced opposite effects on mitochondrial functions without influencing metabolic parameters. Thus, the protective effects of cyto-renin are associated with an altered bioenergetic profile and an enhanced stress tolerance, which are favourable under ischaemic conditions. Therefore, cyto-renin is a promising new target for the prevention of ischaemia-induced myocardial damage.

(Pro)renin receptor (ATP6AP2) depletion arrests As4.1 cells in the G0/G1 phase thereby increasing formation of primary cilia.

The (pro)renin receptor [(P)RR, ATP6AP2] is a multifunctional transmembrane protein that activates local renin-angiotensin systems, but also interacts with Wnt pathways and vacuolar H+ -ATPase (V-ATPase) during organogenesis. The aim of this study was to characterize the role of ATP6AP2 in the cell cycle in more detail. ATP6AP2 down-regulation by siRNA in renal As4.1 cells resulted in a reduction in the rate of proliferation and a G0/G1 phase cell cycle arrest. We identified a number of novel target genes downstream of ATP6AP2 knock-down that were related to the primary cilium (Bbs-1, Bbs-3, Bbs-7, Rabl5, Ttc26, Mks-11, Mks-5, Mks-2, Tctn2, Nme7) and the cell cycle (Pierce1, Clock, Ppif). Accordingly, the number of cells expressing the primary cilium was markedly increased. We found no indication that these effects were dependent of V-ATPase activity, as ATP6AP2 knock-down did not affect lysosomal pH and bafilomycin A neither influenced the ciliary expression pattern nor the percentage of ciliated cells. Furthermore, ATP6AP2 appears to be essential for mitosis. ATP6AP2 translocated from the endoplasmatic reticulum to mitotic spindle poles (pro-, meta- and anaphase) and the central spindle bundle (telophase) and ATP6AP2 knock-down results in markedly deformed spindles. We conclude that ATP6AP2 is necessary for cell division, cell cycle progression and mitosis. ATP6AP2 also inhibits ciliogenesis, thus promoting proliferation and preventing differentiation.

Crosstalk Between Bone and Fat Tissue: Associations Between Vitamin D, Osteocalcin, Adipokines, and Markers of Glucose Metabolism Among Adolescents.

OBJECTIVES: This study aimed to investigate the relationship between 25-hydroxyvitamin D (25(OH)D), osteocalcin, markers of glucose metabolism, and obesity-related parameters among adolescents. METHODS: This was a cross-sectional study with 198 adolescents age 14-18 years. Weight, height, and waist and hip circumferences were measured, as well as the following biochemical parameters: serum 25(OH)D, parathyroid hormone (PTH), total (tOC) and undercarboxylated (ucOC) osteocalcin, adiponectin, leptin, glucose, and insulin. The homeostasis model of assessment estimate of insulin resistance (HOMA-IR) and ß-cell function (HOMA-ß) and quantitative insulin sensitivity check index (QUICKI) were calculated. Student's t test, analysis of variance (ANOVA), Pearson's correlation, and linear regression models were performed. RESULTS: Overweight was observed in 42.6% of the sample. This group presented significantly higher PTH, leptin, insulin, HOMA-IR, and HOMA-ß and lower 25(OH)D, adiponectin, tOC, ucOC, and QUICKI than normal-weight subjects. 25(OH)D was positively correlated with ucOC and adiponectin and negatively associated with body mass index (BMI), weight, and waist circumference (p < 0.05 for all). The association between 25(OH)D and ucOC was also observed in the multiple regression analysis, adjusted for age, BMI, and season of the year (partial r2 = 0.071, p < 0.0001). 25(OH)D and ucOC were not associated with markers of glucose metabolism. However, leptin was strongly correlated with insulin, HOMA-IR, HOMA-ß, and QUICKI (p < 0.0001 for all). CONCLUSION: The present study demonstrated that undercarboxylated osteocalcin is related to 25(OH)D and adiponectin concentrations. Both ucOC and 25(OH)D were lower in overweight and obese adolescents, reinforcing the importance of fighting obesity. Although a relationship of ucOC and 25(OH)D with markers of glucose metabolism was not observed, leptin has shown to be the hormone most related to energy homeostasis.

Evaluation of a Modified Pamidronate Protocol for the Treatment of Osteogenesis Imperfecta.

Intravenous pamidronate is widely used to treat children with osteogenesis imperfecta (OI). In a well-studied protocol ('standard protocol'), pamidronate is given at a daily dose of 1 mg per kg body weight over 4 h on 3 successive days; infusion cycles are repeated every 4 months. Here, we evaluated renal safety of a simpler protocol for intravenous pamidronate infusions (2 mg per kg body weight given in a single infusion over 2 h, repeated every 4 months; 'modified protocol'). Results of 18 patients with OI types I, III, or IV treated with the modified protocol for 12 months were compared to 18 historic controls, treated with standard protocol. In the modified protocol, mild transient post-infusion increases in serum creatinine were found during each infusion but after 12 months serum creatinine remained similar from baseline [0.40 mg/dl (SD: 0.13)] to the end of the study [0.41 mg/dl (SD: 0.11)] (P = 0.79). The two protocols led to similar changes in serum creatinine during the first pamidronate infusion [modified protocol: +2% (SD: 21%); standard protocol: -3% (SD: 8%); P = 0.32]. Areal lumbar spine bone mineral density Z-scores increased from -2.7 (SD: 1.5) to -1.8 (SD: 1.4) with the modified protocol, and from -4.1 (SD: 1.4) to -3.1 (SD: 1.1) with standard protocol (P = 0.68 for group differences in bone density Z-score changes). The modified pamidronate protocol is safe and may have similar effects on bone density as the standard pamidronate protocol. More studies are needed with longer follow-up to prove anti-fracture efficacy.

A new transgenic rat model overexpressing the angiotensin II type 2 receptor provides evidence for inhibition of cell proliferation in the outer adrenal cortex.

This study aimed to elucidate the role of the AT(2) receptor (AT(2)R), which is expressed and upregulated in the adrenal zona glomerulosa (ZG) under conditions of increased aldosterone production. We developed a novel transgenic rat (TGR; TGRCXmAT(2)R) that overexpresses the AT(2)R in the adrenal gland, heart, kidney, brain, skeletal muscle, testes, lung, spleen, aorta, and vein. As a consequence the total angiotensin II (Ang II) binding sites increased 7.8-fold in the kidney, 25-fold in the heart, and twofold in the adrenals. The AT(2)R number amounted to 82-98% of total Ang II binding sites. In the ZG of TGRCXmAT(2)R, the AT(2)R density was elevated threefold relative to wild-type (WT) littermates, whereas AT(1)R density remained unchanged. TGRCXmAT(2)R rats were viable and exhibited normal reproduction, blood pressure, and kidney function. Notably, a slightly but significantly reduced body weight and a moderate increase in plasma urea were observed. With respect to adrenal function, 24-h urinary and plasma aldosterone concentrations were unaffected in TGRCXmAT(2)R at baseline. Three and 14 days of Ang II infusion (300 ng·min(-1)·kg(-1)) increased plasma aldosterone levels in WT and in TGR. These changes were completely abolished by the AT(1)R blocker losartan. Of note, glomerulosa cell proliferation, as indicated by the number of Ki-67-positive glomerulosa cells, was stimulated by Ang II in TGR and WT rats; however, this increase was significantly attenuated in TGR overexpressing the AT(2)R. In conclusion, AT(2)R in the adrenal ZG inhibits Ang II-induced cell proliferation but has no obvious lasting effect on the regulation of the aldosterone production at the investigated stages.

Regression of cardiac hypertrophy in cyp1a1ren-2 transgenic rats.

PURPOSE: To evaluate the usefulness of the cyp1a1ren-2 transgenic rat model of inducible hypertension for studies of the development and regression of cardiac hypertrophy. MATERIALS AND METHODS: Cyp1a1ren-2 rats received a diet containing 0% or 0.167% indole-3-carbinonl (I3C) for 4 weeks to induce hypertension. Cardiac magnetic resonance imaging (MRI) at 7 T was performed every second week for 10 weeks to measure left ventricular mass and the ejection fraction. Concomitantly, in six cyp1a1ren-2 rats blood pressure was recorded telemetrically. RESULTS: Plasma prorenin concentrations rose from 138 ± 38 to 15,490 ± 3990 ng/angiotensin I/mL/h (P < 0.001) in I3C-treated transgenic rats and returned to basal levels after cessation of I3C. Mean blood pressure increased to a plateau of 169 ± 11 mmHg by the second week of induction. After cessation of I3C (day 28), arterial pressure dropped to values slightly below those prior to induction within 4 days (basal: 106 ± 7 mmHg, day 32: 103 ± 21 mmHg; NS). At day 28, left ventricular mass was increased by 39% vs. 4% in controls (P < 0.001) without changes of the ejection fraction. Cardiac hypertrophy was completely reversed at day 70, as evaluated by MRI. CONCLUSION: The cyp1a1ren-2 transgenic rat is a useful model to study reversal and healing in the absence of surgical interventions.

Official position of the Brazilian Association of Bone Assessment and Metabolism (ABRASSO) on the evaluation of body composition by densitometry: part I (technical aspects)-general concepts, indications, acquisition, and analysis.

OBJECTIVE: To review the technical aspects of body composition assessment by dual-energy X-ray absorptiometry (DXA) and other methods based on the most recent scientific evidence. MATERIALS AND METHODS: This Official Position is a result of efforts by the Scientific Committee of the Brazilian Association of Bone Assessment and Metabolism (Associação Brasileira de Avaliação Óssea e Osteometabolismo, ABRASSO) and health care professionals with expertise in body composition assessment who were invited to contribute to the preparation of this document. The authors searched current databases for relevant publications. In this first part of the Official Position, the authors discuss the different methods and parameters used for body composition assessment, general principles of DXA, and aspects of the acquisition and analysis of DXA scans. CONCLUSION: Considering aspects of accuracy, precision, cost, duration, and ability to evaluate all three compartments, DXA is considered the gold-standard method for body composition assessment, particularly for the evaluation of fat mass. In order to ensure reliable, adequate, and reproducible DXA reports, great attention is required regarding quality control procedures, preparation, removal of external artifacts, imaging acquisition, and data analysis and interpretation.

Associations between markers of glucose metabolism and bone measures among diabetic and non-diabetic adults.

PURPOSE: To investigate the relationships between bone measures, vitamin D status and markers of glucose metabolism among diabetic and non-diabetic adults. METHODS: Cross sectional study with 298 adults (mean age 57.5 years, SD = 14.8; 44.3% male, 16.9% diabetic) participants of the Health Survey-São Paulo (ISA-Capital) 2014-2015. Blood samples were collected to assess serum glucose, insulin and 25 hydroxyvitamin D [25(OH)D] concentrations. Dual-energy x-ray absorptiometry (DXA) was performed to determine total body fat; total lean mass; full body bone mineral density (BMD); lumbar spine BMD and bone mineral content (BMC); and femur BMD and BMC. Fat mass index (FMI), lean mass index (LMI), quantitative insulin sensitivity check index (QUICKI), homeostasis model assessment of insulin resistance (HOMA-IR) and of ß-pancreatic cell function (HOMA-ß) were calculated. Linear regression analysis were performed. RESULTS: Multiple bone measures were associated with markers of glucose metabolism in analyses adjusted by age and sex. However, after additional adjustments by LMI, FMI and serum 25(OH)D, only associations of lumbar spine BMC with HOMA-IR (ß = 0.167; p = 0.035) and QUICKI (ß = -1.879; p = 0.027) persisted, in the subgroup of diabetic participants. Analysis restricted to diabetic subjects revealed stronger correlations between bone parameters and markers of glucose metabolism. CONCLUSIONS: Our study observed positive associations between BMD and markers of insulin resistance among a sample of adults. Correlations were stronger among diabetic subjects, and some associations between bone and glucose metabolism were independent of adiposity. Findings reinforce the need of further research for better understanding the bidirectional and multifactorial crosstalk between glucose homeostasis and bone metabolism.

The renin-angiotensin system as a primary cause of polyarteritis nodosa in rats.

Polyarteritis nodosa is a necrotizing vasculitis of medium-sized arteries of unknown origin. Hypertension is present in 30% of patients with polyarteritis nodosa. In those cases, high renin levels are thought to be secondary to renal involvement. The present study was performed to identify causal factors of polyarteritis nodosa. In cyp1a1ren-2 transgenic rats, vasculitis of medium-sized arteries resembling classical polyarteritis nodosa can be induced. In this model, oral administration of indole-3-carbinol (I3C) activates the liver-specific cyp1a1 promoter, leading to prorenin expression in a dose-dependent manner. After the first 6 weeks of chronic induction with 0.125% I3C, the mean arterial pressure reached a plateau of about 170 mmHg. Ten out of 11 I3C-treated rats, which were chronically instrumented with a telemetric device to measure blood pressure, developed polyarteritis nodosa within 10 weeks of I3C treatment. I3C alone or instrumentation alone did not cause polyarteritis nodosa. The angiotensin-converting enzyme inhibitor captopril completely prevented the development of polyarteritis nodosa, indicating that local angiotensin II generation is a pathogenetic factor in this model. The renin-angiotensin system can play a primary role in the development of polyarteritis nodosa in rats.

Correlations among vitamin K intake, body fat, lipid profile and glucose homeostasis in adults and the elderly.

Objective Recent research has investigated the possible inverse relationship between vitamin K intake and body fat. In addition, an increasing number of studies are supporting a key role for this vitamin in improving lipid profile and insulin sensitivity and reducing the risk of type 2 diabetes mellitus, but little is known about what mechanisms would be involved. Thus, the objective of this study was to investigate the relationship between vitamin K intake (in the form of phylloquinone - PK), body fat, lipid profile and markers of glucose homeostasis in adults and the elderly. Subjects and methods A cross-sectional study with 298 participants (46% men) in the São Paulo Health Survey 2014-2015. Spearman correlations were performed to evaluate the associations between vitamin K intake and the biochemical and body composition measures. Results Among normal-weight male adults (n = 15), PK intake presented a positive correlation with the quantitative insulin sensitivity check index (QUICKI) (r = 0.525; p = 0.045). Among men with high fat mass index (FMI) (n = 101), PK intake had a negative correlation with homeostasis model assessment estimate for ß-cell function (HOMA-ß) (r = -0.227; p = 0.022). In women with high FMI (n = 122), PK intake had a negative correlation with HOMA-ß (r = -0.199, p = 0.032) and insulin (r = -0.207, p = 0.026). No correlations were found between PK intake and lipid profile. Conclusions Our findings support a potential relationship among PK intake, body fat and markers of glucose homeostasis in adults and the elderly.

Cytosolic renin is targeted to mitochondria and induces apoptosis in H9c2 rat cardiomyoblasts.

One important goal in cardiology is to prevent necrotic cell death in the heart. Necrotic cell death attracts neutrophils and monocytes into the injured myocardium. The consequences are fibrosis, remodelling and cardiac failure. The renin-angiotensin system promotes the development of cardiac failure. Recently, alternative renin transcripts have been identified lacking the signal sequence for a cotranslational transport to the endoplasmatic reticulum. These transcripts encode for a cytosolic renin with unknown functions. The expression of this alternative transcript increases after myocardial infarction. We hypothesized that cytosolic renin plays a role in survival and death of cardiomyocytes. To test this hypothesis, we overexpressed secretory or cytosolic renin in H9c2 cardiomyblasts and determined the rate of proliferation, necrosis and apoptosis. Proliferation rate, as indicated by BrdU incorporation into DNA, was reduced by secretory and cytosolic renin (cells transfected with control vector: 0.33 +/- 0.06; secretory renin: 0.12 +/- 0.02; P < 0.05; cytosolic renin: 0.15 +/- 0.03; P < 0.05). Necrosis was increased by secretory renin but decreased by cytosolic renin (LDH release after 10 days from cells transfected with control vector: 68.5 +/- 14.9; secretory renin: 100.0 +/- 0; cytosolic renin: 25.5 +/- 5.3% of content, each P < 0.05). Mitochondrial apoptosis, as indicated by phosphatidylserin translocation to the outer membrane, was unaffected by secretory renin but increased by cytosolic renin (controls: 23.8 +/- 3.9%; secretory renin: 22.1 +/- 4.7%; cytoplasmatic renin: 41.2 +/- 3.8%; P < 0.05). The data demonstrate that a cytosolic renin exists in cardiomyocytes, which in contradiction to secretory renin protects from necrosis but increases apoptosis. Non-secretory cytosolic renin can be considered as a new target for cardiac failure.

Lack of cardiac fibrosis in a new model of high prorenin hyperaldosteronism.

The aim of the present study was to test the hypothesis that elevation of prorenin in plasma is sufficient to induce cardiac fibrosis. Normotensive cyp1a1ren-2 transgenic rats with normal plasma prorenin and aldosterone levels were given 0.125% indole-3-carbinol (I3C) orally for a period of 12 wk. Plasma prorenin and aldosterone levels were determined in 4-wk intervals, and cardiac marker enzymes for hypertrophy, fibrosis, and oxidative stress as well as cardiac pathology were investigated. In I3C-treated cyp1a1 ren-2 transgenic rats, plasma prorenin concentrations were >100-fold elevated (> or = 7.1 + or - 2.6 microg ANG I.ml(-1).h(-1) vs. < or = 0.07 + or - 0.1; P < 0.001), whereas active renin levels were suppressed (0.09 + or - 0.02 vs. 0.2 + or - 0.1; P < 0.05). Aldosterone concentrations were elevated three- to fourfold for a period of >4 wk (574 + or - 51 vs. 160 + or - 68 pg/ml; P < 0.01). After 12 wk of I3C, rats exhibited moderate cardiac hypertrophy (heart weight/body weight 2.5 + or - 0.04 vs. 3.1 + or - 0.1 mg/g; P < 0.01). There was a slight increase in mRNA contents of endothelin 1 (1.21 + or - 0.08 vs. 0.75 + or - 0.007; P < 0.001), NADP oxidase-2 (1.03 + or - 0.006 vs. 0.76 + or - 0.04; P < 0.001), transforming growth factor-beta (0.99 + or - 0.06 vs. 0.84 + or - 0.04; P < 0.05), collagen type I (1.32 + or - 0.32 vs. 0.94 + or - 0.18; P < 0.05), and intercellular adhesion molecule-1 (1.12 + or - 0.12 vs. 0.84 + or - 0.08; P < 0.05). These genes are known to be stimulated by the renin-angiotensin system. There were no histological signs of fibrosis in the heart. We found that prorenin and aldosterone alone are not sufficient to induce considerable cardiac fibrosis in the absence of sodium load.

Prevalence of vitamin D insufficiency in Brazilian adolescents.

BACKGROUND/AIMS: Cutaneous sun exposure and dietary vitamin D intake are important determinants of vitamin D status. The objective of the present study was to evaluate the vitamin D status of a group of healthy adolescent students living in Brazil. METHODS: One hundred and thirty-six adolescents, 64 boys and 72 girls, aged 16-20 years old, living in a rural town in the state of São Paulo, Brazil, participated in this study. RESULTS: The mean dietary vitamin D intake was 140 (120-156) IU/day [3.5 (3.0-3.9) microg/day]. Only 14.9% of the students met the daily adequate intake recommendation of vitamin D. Only 27.9% practice physical activity outdoors and 17.6% of the adolescents apply sunscreen daily. The mean 25(OH)D concentration was 73.0 (22.0) nmol/l [29.2 (8.8) ng/ml]. Vitamin D insufficiency was observed in 60% of adolescents. CONCLUSIONS: The present study suggests that even in a sunny climate like Brazil the prevalence of vitamin D insufficiency in adolescents is high. Most likely this is due to low intakes of vitamin D in this group. Due to the limited extent of natural dietary sources of vitamin D, a policy of vitamin D food fortification should be considered in the future, and in the meantime greater use of vitamin D supplements in this population group should be encouraged to provide the increased amounts of this essential nutrient for optimal health.

Dose-dependent titration of prorenin and blood pressure in Cyp1a1ren-2 transgenic rats: absence of prorenin-induced glomerulosclerosis.

OBJECTIVE: Prorenin has been associated with cardiovascular disease and the development of glomerulosclerosis via a renin/prorenin receptor. In cyp1a1ren-2 transgenic rats, prorenin levels and arterial pressure can be increased by oral administration of indole-3-carbinol (I3C). The transgenic strain has been used as a model of malignant hypertension. METHODS: The present study was designed to test the hypotheses that (i) low doses of I3C would result in dose-dependent sustained increases in arterial pressure without signs of malignancy, making cyp1a1ren-2 transgenic rats a useful model to study nonmalignant hypertension, and (ii) cyp1a1ren-2 transgenic rats would develop glomerulosclerosis when they were chronically exposed to 0.125% I3C in their diet. RESULTS: I3C treatment for 2 weeks resulted in increases of plasma prorenin concentrations and arterial pressure in a dose-dependent manner. Rats thrived well over a period of 12 weeks on dietary I3C concentrations (wt/wt) of 0.125%. Plasma prorenin concentration rose from 0.1 +/- 0.1 microg to 17.9 +/- 5.0 mug angiotensin I/ml per h (P < 0.01) and mean arterial pressure increased to a plateau of 170 +/- 5 mmHg (P < 0.001) between weeks 6 and 12. After 12 weeks of 0.125% I3C, rats exhibited moderate hypertensive renal vasculopathy, but no histological signs of glomerulosclerosis. CONCLUSIONS: The cyp1a1ren-2 transgenic rat model allows for chronic dose-dependent titration of arterial pressure by a simple and non-invasive intervention, making this strain a useful model to study malignant and nonmalignant arterial hypertension. High circulating prorenin levels per se do not cause glomerulosclerosis.

Integration of "omics" techniques: Dronedarone affects cardiac remodeling in the infarction border zone.

Dronedarone improves microvascular flow during atrial fibrillation and reduces the infarct size in acute models of myocardial infarction. However, dronedarone might be harmful in patients with recent decompensated heart failure and increases mortality in patients with permanent atrial fibrillation. A pathophysiological explanation for these discrepant data is lacking. This study investigated the effects of dronedarone on gene and protein expression in the infarcted area and border zone in pigs subjected to anterior ischemia/reperfusion myocardial infarction. The ischemia/reperfusion myocardial infarction was induced in 16 pigs. Eight pigs were treated with dronedarone for 28 days after myocardial infarction, the remaining pigs served as control. Microarray-based transcriptome profiling and 2D-DIGE-based proteome analysis were used to assess the effects of dronedarone on left ventricular gene expression in healthy (LV), infarcted (MI), and border zone tissue. Selected targets were validated by RT-qPCR or immunoblot analyses, with special emphasize given to the transcriptome/proteome overlap. Combined "omics" analysis was performed to identify most significant disease and function charts affected by dronedarone and to establish an integrated network. The levels of 879 (BZ) or 7 (MI) transcripts and 51 (LV) or 15 (BZ) proteins were significantly altered by dronedarone, pointing to a substantial efficacy of dronedarone in the border zone. Transcriptome and proteome data indicate that dronedarone influences post-infarction remodeling processes and identify matricellular proteins as major targets of dronedarone in this setting. This finding is fully supported by the disease and function charts as well as by the integrated network established by combined "omics". Dronedarone therapy alters myocardial gene expression after acute myocardial infarction with pronounced effects in the border zone. Dronedarone promotes infarct healing via regulation of periostin and might contribute to the limitation of its expansion as well as cardiac rupture. Thus, there are no experimental hints that dronedarone per se has direct harmful effects after MI in ventricular tissue. Impact statement Dronedarone reduced the infarct size in models of acute myocardial infarction (MI). Here, we show that dronedarone attenuates many of the substantial changes in gene expression that are provoked by acute myocardial infarction (AMI) in pigs. Dronedarone modifies the expression of gene panels related to post-infarction cardiac healing and remodeling processes and, most remarkably, this occurs predominantly in the infarction border-zone and much less so in the vital or infarcted myocardium. Combined "omics" identified matricellular proteins and ECM as major dronedarone-regulated targets and emphasizes their relevance for Disease Charts and Tox Function Charts associated with tissue remodeling and cellular movement. The results demonstrate dronedarone's capability of regulating cardiac repair and remodeling processes specifically in the infarction border zone and identify underlying mechanisms and pathways that might be employed in future therapeutic strategies to improve long-term cardiac tissue function and stability.

Effects of parathyroidectomy on bone remodeling markers and vitamin D status in patients with chronic kidney disease-mineral and bone disorder.

BACKGROUND/AIMS: Chronic renal failure (CRF) is often associated with bone disorders including chronic kidney disease-mineral and bone disorder (CKD-MBD). Parathyroid hormone (PTH) has a relationship to bone remodeling, and so this study was undertaken to evaluate changes in bone remodeling markers after parathyroidectomy (PTX). METHODS: Twelve adult patients, mean age 43.4 +/- 12.7 years, of both genders, were evaluated, prior to and six months after PTX. Analysis of biochemical markers of bone metabolism, such as total and ionized calcium, phosphorus, 25(OH)D(3), total alkaline phosphatase (TAP), bone-specific alkaline phosphatase (BAP), intact PTH, osteoprotegerin (OPG), and tartrate-resistant acid phosphatase isoform 5b (TRAP), were measured. RESULTS: No changes were observed after PTX in the serum total and ionized calcium, TAP, BAP, and 25(OH)D(3). After surgery there was a significant decrease in serum phosphorus, iPTH, and TRAP (P < 0.001). No significant change was observed in OPG; however there was a positive correlation between OPG and 25(OH)D(3) before and after surgery (r = 0.774, P = 0.014; and r = 0.706, P = 0.01, respectively). The percentage of patients with vitamin D deficiency decreased from 16.7% to 8.3%, while those with sufficient levels increased from 41.7% to 58.3%. CONCLUSION: The small number of patients in the study notwithstanding, the present study is unique because it provides information on bone metabolism and vitamin D status six months after PTX. The removal of parathyroid glands significantly decreased bone resorption and indicated a tendency of 25(OH)D(3) concentration to increase. However, the precise role of OPG and BAP in the improvement in bone remodeling in patients with CKD-MBD requires further study.

Anti-necrotic and cardioprotective effects of a cytosolic renin isoform under ischemia-related conditions.

UNLABELLED: In the heart, secretory renin promotes hypertrophy, apoptosis, necrosis, fibrosis, and cardiac failure through angiotensin generation from angiotensinogen. Thus, inhibitors of the renin-angiotensin system are among the most potent drugs in the treatment of cardiac failure. Renin transcripts have been identified encoding a renin isoform with unknown targets and unknown functions that are localized to the cytosol and mitochondria. We hypothesize that this isoform, in contrast to secretory renin, exerts cardioprotective effects in an angiotensin-independent manner. Cells overexpressing cytosolic renin were generated by transfection or obtained from CX(exon2-9)renin transgenic rats. Overexpression of cytosolic renin reduced the rate of necrosis in H9c2 cardiomyoblasts and in primary cardiomyocytes after glucose depletion. These effects were not mediated by angiotensin generation since an inhibitor of renin activity did not influence the in vitro effects. siRNA-mediated knockdown of endogenous cytosolic renin increased the rate of necrosis and aggravated the pro-necrotic effects of glucose depletion. Isolated perfused hearts obtained from transgenic rats overexpressing cytosolic renin exhibited a 50% reduction of infarct size after ischemia-reperfusion injury. Cytosolic renin is essential for survival, both under basal conditions and during glucose starvation. The protective effects are angiotensin-independent and contrary to the known actions of secretory renin. KEY MESSAGES: A cytosolic isoform of renin with unknown functions is expressed in the heart. Cytosolic renin diminishes ischemia induced damage to the heart. The protective effects of cytosolic renin contradict the known function of secretory renin. The effects of cytosolic renin are not mediated via angiotensin generation. Renin-binding protein is a potential target for cytosolic renin.

Digital vertebral morphometry performed by DXA: a valuable opportunity for identifying fractures during bone mass assessment.

OBJECTIVES: To evaluate the usefulness of vertebral morphometry in identifying unreferred vertebral fractures and correlate potential risk factors. SUBJECTS AND METHODS: Female patients above 45 years, postmenopausal for at least 2 years, diagnosed with osteoporosis and undergoing treatment for at least three months were considered eligible. All of them underwent bone densitometry and vertebral morphometry performed by concomitant DXA. The presence of fractures was defined between T7 and L4; only moderate and severe fractures were considered for analysis. All volunteers were submitted to laboratory tests, anthropometry and responded a questionnaire on their lifestyle habits and medical history. RESULTS: Thirty two (17%) out of the 188 female patients presented with at least one vertebral fracture, among whom only 4 (12.5%) were previously aware of the fracture. The fractures were mainly located on the thoracic spine. Nine patients had severe fractures (28.1%), whereas 23 had moderate fractures (71.9%). On average, patients with fractures were 5 years older and weighed 5 kilograms less than those without fractures. The creatinine clearance was on average 9 mL/min less in patients with vertebral fracture. The assessment of vertebral fractures by morphometry is a fast, accurate and complementary method associated with low radiation exposure for identifying moderate and severe vertebral fractures. Predisposition to vertebral fractures does not depend solely on BMD.

Dronedarone does not affect infarct volume as assessed by magnetic resonance imaging in a porcine model of myocardial infarction.

Dronedarone has been demonstrated to be harmful in patients with recent decompensated heart failure. Furthermore, a PALLAS study reported that dronedarone therapy increases mortality rates in patients with permanent atrial fibrillation. Although a pathophysiological explanation for these finding remains to be elucidated, the long term effects of dronedarone on myocardial structure and stability have been suggested. The aim of the present study was to determine whether dronedarone therapy affects left ventricular (LV) function in a chronic model of myocardial infarction (MI). An anterior MI was induced in 16 pigs. Of these animals, eight pigs were then treated with dronedarone for 1 week prior to, and 4 weeks following MI, the remaining pigs served as controls. LV angiography was performed 4 weeks after MI to determine the LV ejection fraction (LVEF). A post­mortem magnetic resonance imaging scan of the LV was then performed on the two groups (n=6) to determine the volume and size of the induced MI. Dronedarone therapy did not affect systemic and intracardiac hemodynamic parameters or LVEF during the follow­up assessment. Of note, dronedarone had no negative effect on the total infarct volume and size and did not induce lethal proarrhythmic effects following the induced anterior MI. Therefore, the results suggested that dronedarone did not increase the volume or size of induced anterior MI and did not affect LV performance. Thus, dronedarone therapy was observed to be safe in a porcine model of anterior MI.