Heart Tissue Levels Research Articles

Association between Diastolic Dysfunction with Inflammation and Oxidative Stress in Females ob/ob Mice.

Objective: To evaluate autonomic and cardiovascular function, as well as inflammatory and oxidative stress markers in ob/ob female mice.Methods: Metabolic parameters, cardiac function, arterial pressure (AP), autonomic, hormonal, inflammatory, and oxidative stress markers were evaluated in 12-weeks female wild-type (WT group) and ob/ob mice (OB group).Results: OB animals showed increased body weight, blood glucose, and triglyceride levels, along with glucose intolerance, when compared to WT animals. Ejection fraction (EF) and AP were similar between groups; however, the OB group presented diastolic dysfunction, as well as an impairment on myocardial performance index. Moreover, the OB group exhibited important autonomic dysfunction and baroreflex sensitivity impairment, when compared to WT group. OB group showed increased Angiotensin II levels in heart and renal tissues; decreased adiponectin and increased inflammatory markers in adipose tissue and spleen. Additionally, OB mice presented a higher damage to proteins and lipoperoxidation and lower activity of antioxidant enzymes in kidney and heart. Correlations were found between autonomic dysfunction with angiotensin II and inflammatory mediators, as well as between inflammation and oxidative stress.Conclusions: Our results showed that female adult ob/ob mice presented discrete diastolic dysfunction accompanied by autonomic disorder, which is associated with inflammation and oxidative stress in these animals.

Zebra blenny protein hydrolysates as a source of bioactive peptides with prevention effect against oxidative dysfunctions and DNA damage in heart tissues of rats fed a cholesterol-rich diet

High-cholesterol diet (HCD) is suspected to produce in excess free radicals having adverse effects on human health and causing atherosclerosis damage in heart tissues. In our study, the effects of zebra blenny protein hydrolysates (ZBPHs) were investigated on cardiac oxidant/antioxidant status as well as DNA damage and histopathological disorders in rats, fed with a hypercholesterolemic diet. The molecular weight distribution of the hydrolysates was determined by size exclusion chromatography, which analyzed a representative hydrolysate type with a weight range of 3–20kDa. ZBPHs effectively protected heart genomic DNA against oxidative damage induced by Fenton's reagent. HCD promoted oxidative stress with a rise in serum aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) activities and thiobarbituric acid reactive substance (TBARS), advanced oxidation protein product (AOPP) and hydrogen peroxide (H2O2) levels in heart tissues. An increase in glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT) activities as well as a fall in ATPase activities and glutathione (GSH) level was also noted in heart of hypercholesterolemic rats. Treatment with ZBPHs ameliorated the biochemical parameters cited above. In addition, pre-treatment with ZBPHs prevented DNA fragmentation. The histopathological findings confirmed the biochemical results and the potential antioxidant activities of ZBPHs which can help the cure and management of cardiovascular diseases induced by high-cholesterol levels.

Effect of Androgen on Normal Biodistribution of [18F]-2'-Fluoro-5-methyl-1-beta-D-arabinofuranosyluracil (18F-FMAU) in Athymic Non-tumor-bearing Male Mice.

We assessed the association between the presence and absence of androgen on the normal biodistribution of the positron emission tomography (PET) cellular proliferation imaging biomarker, [18F]-2'-Fluoro-5-methyl-1-beta-D-arabinofuranosyluracil (18F-FMAU), in mice. Non-castrated (n=4) and castrated (n=4) athymic non-tumor-bearing male mice served as models for presence and absence, respectively, of androgen. MicroPET-CT scans were performed 1 h following tail vein administration of 200 uCi of 18F-FMAU. Imaging was performed at baseline and then at 7-day intervals longitudinally for 35 days only in castrated mice following subcutaneous introduction of a 12.5 mg, 21-day release, dihydrotestosterone pellet. Mean standardized uptake values (SUVmean) were obtained for liver, heart, and muscle. Several two-group comparisons of average of SUVmean were performed. Pre-pellet baseline average SUVmean (±s.d.) values in castrated mice were significantly lower than baseline non-castrated values, increased on day 15 and reached peak values on day 28, at which time they were significantly higher than corresponding baseline levels in both non-castrated and pre-pellet castrated mice. The peak values decreased significantly following dihydrotestosterone withdrawal. There is a significant modulatory effect of androgen on normal 18F-FMAU uptake levels in mice liver, heart and muscle tissues.

Piperine modulates isoproterenol induced myocardial ischemia through antioxidant and anti-dyslipidemic effect in male Wistar rats

Myocardial infarction due to ischemia accounts for majority of deaths among cardiovascular disorders. Isoproterenol (ISO) induced myocardial infarction and the protection offered by piperine was investigated in the present report. Lipid profile analysis by determining the levels of cholesterol, phospholipids, triglycerides and lipoproteins in serum and heart tissues showed anti-dyslipidemic action of piperine against ISO induced myocardial injury by modulating the ISO induced altered lipid profiles, maintaining to near control values. ISO treatment increased TBARS levels, PCC, serum markers of heart, depleted antioxidant status (GSH, SOD, CAT, GPx and GST) in tissues and, total, protein- and non-protein-sulfhydryl levels in serum and heart tissues. Piperine pre-treatment decreased the levels of serum markers, lipid peroxidation and PCC with increased antioxidant status in the heart tissues of ISO administered rats. Increased levels of the glycoprotein components in serum and decreased levels in heart tissues upon ISO administration were restored to near normal levels by piperine pre-treatment. Our present reports also showed the modulatory effect of piperine on membrane bound ATPase's showing protection against ISO induced changes in membrane fluidity. The present study proved piperine as a potent therapeutic agent with its antioxidant and anti-dyslipidemic action against ISO induced myocardial infarction.

Antioxidant effects of melatonin in heart tissue after induction of experimental periodontitis in rats.

The aim of this study was to evaluate the effects of melatonin on the oxidative stress in heart tissues after induction of experimental periodontitis in rats. Thirty Wistar Albino male rats were divided into four groups as follows: healthy + saline solution (Hs, n = 7), healthy + melatonin (Hm, n = 7), periodontitis + saline solution (Ps, n = 8), and periodontitis + melatonin (Pm, n = 8). Experimental periodontitis was induced using a ligature placed at the gingival margin of the maxillary second molars. Melatonin was applied intraperitoneally (10 mg/kg) every day for 2 weeks. After sacrificing the rats, serum levels of malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) levels, and melatonin levels were evaluated. The Pm group exhibited lower alveolar bone loss than the Ps group. Melatonin levels increased in the periodontitis groups, and the Pm group had lower MDA levels and higher GSH-Px levels than the Ps group. These findings suggest that melatonin administration reduces MDA and increases GSH-Px levels in heart tissue, and these effects may be due to its antioxidant properties. Further studies are needed to understand the effects of melatonin on the association between periodontitis and cardiovascular disease.

Anti-Interleukin-22-Neutralizing Antibody Attenuates Angiotensin II-Induced Cardiac Hypertrophy in Mice.

Background Interleukin- (IL-) 22 is considered a proinflammatory cytokine. Recent evidence has demonstrated that it plays a role in cardiovascular diseases. In the recent study, we investigate whether IL-22 is involved in cardiac hypertrophy. Methods Angiotensin II was used to build hypertrophy model and the IL-22 and IL-22 receptor 1 (IL-22R1) levels in heart tissue were measured. In addition, angiotensin II-treated mice received an injection of anti-IL-22-neutralizing antibody (nAb) to investigate the effects of IL-22 nAb on myocardial hypertrophy, cardiac function, and cardiac fibrosis; the activation of the signaling pathway and the prohypertrophic inflammatory cytokine mRNA levels was detected. Furthermore, the effect of IL-22 nAb on angiotensin II-induced hypertrophy in vitro was also determined. Results IL-22 and IL-22R1 levels were significantly increased after angiotensin II infusion. Anti-IL-22 nAb significantly alleviated the severity of hypertrophy, prevented systolic and diastolic abnormalities, reduced cardiac fibrosis, STAT3 and ERK phosphorylation, and downregulated the mRNA expression of IL-17, IL-6, IL-1β, IFN-γ, and TNF-α. In addition, IL-22 nAb attenuated angiotensin II-induced hypertrophy in H9C2 cells. Conclusion Our data demonstrated that neutralization of IL-22 alleviated angiotensin II-induced cardiac hypertrophy. The downregulation of IL-22 may be a novel therapeutic strategy to prevent cardiac hypertrophy.

Effects of age, gender, BMI, settlement and smoking on lead and cadmium accumulation in heart tissue

Cadmium (Cd) and lead (Pb) are well-known risk factors for peripheral arterial and some other cardiovascular diseases. The aim of the study was to determine the level of both toxic metals, in the human heart tissue and compare the accumulation to their age, gender, body mass index, living environment and smoking habit. Heart tissues of the 112 autopsy cases, representing a sample set of population living in capital city of Turkey: Ankara Province (both in city center and suburban), were collected from Turkey Ministry of Justice, Council of Forensic Medicine in Ankara Branch. Lead and cadmium analysis was carried out by Graphite Furnace Atomic Spectrometry technique. Cd/Pb results and individuals’ information were assessed by SPSS statistically. Average value of Pb was found as 1374.99±237.01ppb and Cd was found as 287.54±66.43ppb in all heart tissue samples. Consideration of metal levels in autopsy heart tissues by age, statistically significant positive correlation in Cd was observed (p

Evaluation of oxidative stress markers in the heart and liver of rainbow trout (Oncorhynchus mykiss walbaum) exposed to the formalin.

The aim of this study was to examine change in lipid and protein oxidation biomarkers, transamination enzymes and lactate dehydrogenase activities, lactate and pyruvate levels in liver and heart tissue of rainbow trout (Oncorhynchus mykiss Walbaum) that was exposed to formalin baths. Increase of 2-thiobarbituric acid reactive substances and carbonyl derivatives of protein oxidative destruction was noticed only in cardiac tissue of formalin-exposed fish. Activity of lactate dehydrogenase and lactate level in the cardiac tissue were elevated, indicating active glycolysis. Effects of formalin disinfection were different in both tissues. Aldehydic and ketonic derivatives of oxidatively modified proteins in liver were consistently reduced upon exposure to the formalin. In support of this, decrease in alanine and aspartate aminotransferases was noticed. Formalin disinfection of rainbow trout results in metabolic plasticity, predominantly in liver with decreased levels of oxidative stress biomarkers and aminotransferases activity. Formalin-induced oxidative stress in the cardiac tissue was more considerable.

Testosterone and Voluntary Exercise Promote Angiogenesis in Hearts of Rats with Diabetes by Enhancing Expression of VEGF-A and SDF-1a.

Impaired angiogenesis in cardiac tissue is a major complication of diabetes. This study was aimed to evaluate the effects of testosterone and voluntary exercise on vascular endothelial growth factor-A (VEGF-A), stromal cell-derived factor 1a (SDF-1a) and myocardial capillary density in heart of rats with diabetes. Type 1 diabetes was induced by intraperitoneal injection of 55 mg/kg of streptozotocin in 80 male Wistar rats. After 42 days of treatment with testosterone (2 mg/kg/day) or voluntary exercise alone or in combination, angiogenesis was determined in the hearts by immunostaining for PECAM-1/CD31. The expressions of VEGF-A and SDF-1a levels in heart were also determined by the ELISA method. Our results showed that capillary density, VEGF-A and SDF-1a levels in the heart were significantly decreased in castrated rats with diabetes, whereas these effects were reversed by testosterone and exercise. Furthermore, simultaneous treatment of castrated rats with diabetes with testosterone and exercise had a synergistic effect on capillary density, VEGF-A and SDF-1a levels in the heart. In the group with diabetes, either testosterone or exercise increased capillary density, VEGF-A and SDF-1a protein levels in heart tissue. However, the effects of combination therapy in rats with diabetes with testosterone and exercise on capillary density, VEGF-A and SDF-1a levels in the heart was synergistic. Our findings suggest that testosterone and exercise can promote neoangiogenesis in rats with diabetes and in castrated rats with diabetes. The proangiogenesis effect of testosterone and exercise is associated with the enhanced expression of VEGF-A and SDF-1a in heart tissue.

Protective Effect of Tert Butyl Hydroquinone on Diazinon-Induced Oxidative Stress in Brain and Heart of Male Rats

Background: The present study was designed to investigate the chronic effects of diazinon on oxidative stress markers in brain and heart of rats and the possible protective effects of tert butyl hydroquinone (TBHQ), as an antioxidant. Objectives: Therefore, the present study was undertaken to evaluate the protective role of TBHQ on oxidative stress induced by diazinon in brain and heart tissues of rats after 7 weeks exposure to sub-lethal dose of diazinon. Materials and Methods: In this experimental study, 28 male Wistar rats which were randomly divided into four groups: diazinon group (10 mg/kg, once a day), TBHQ group (0.028 g/kg of diet, once a day), diazinon + TBHQ group (diazinon, 10 mg/kg, once a day + TBHQ, 0.028 g/kg of diet, once a day) and control group (corn oil, as vehicle of diazinon and TBHQ). The animals were treated with diazinon, TBHQ or corn oil orally using a stomach tube for 7 weeks. Results: At the end of 7th week, total thiol groups, ferric reducing antioxidant power (FRAP) and malondialdehyde (MDA) levels in brain and heart tissues were investigated. A significant increase in MDA levels (P = 0.01) in heart tissue was evident in diazinon group, when compared to control group. Rats of the TBHQ and diazinon + TBHQ groups presented a significant increase in thiol groups (P = 0.01), when compared to control and diazinon groups. In addition, TBHQ administration significantly increased the FRAP level of brain tissue in TBHQ and TBHQ + diazinon groups, when compared with control and diazinon groups, respectively (P = 0.001). Conclusions: The results of the present study showed that TBHQ treatment could improve antioxidant status in brain and heart tissues of rats with chronic toxicity of diazinon. However, it could not ameliorate the lipid peroxidation sufficiently.

Effects of aminoguanidine treatment on endothelial nitric oxide synthase expression and nitric oxide levels in streptozotocin induced diabetic rats

Objective: Formation of advanced glycation end products (AGE) in Diabetes Mellitus contribute to endothelial dysfunction by permanent modifications of proteins and decrease NO concentration which regulate vascular tonicity and thus endothelial functions. We investigated the effects of AGE inhibitor aminoguanidine (AG) treatment on endothelial nitric oxide synthase (eNOS) expression and NO levels in streptozotocin induced diabetic rats. Material and method: 38 male Sprague-Dawley rats were separated into 4 groups. Group 1 was healthy controls without any intervention (HC group). Group 2 received AG in drinking water (HC+AG group). Third and fourth groups received 65 mg/kg streptozotocin intraperitoneally to induce diabetes. Group 3 was followed without further intervention (DM group). Group 4 received aminoguanidine in drinking water (DM+AG group). After 12 weeks, glucose, HbA 1c , NO levels an eNOS expressions were determined. Results: NO levels were significantly lower in DM and HC+AG groups compared to HC group. There was no significant difference between the NO levels of DM and DM+AG groups. eNOS protein levels in heart tissues were significantly lower in DM group than the HC group. eNOS levels were significantly higher in DM+AG than DM group. There was no significant difference in eNOS levels in kidney tissues among the groups. Conclusion: Decreased NO levels in DM group was consistent with the literature. eNOS expression in heart tissues were decreased in DM group, their concentration were increased with AG therapy. Further studies are needed to elaboate the effects of AG in tissues and the side effects observed in some studies.

Tetrahydrobiopterin Concentrations in Normal and Coronary Artery Diseased Heart Tissue

Background: Tetrahydrobiopterin (BH4) is a cofactor that plays a major role in cardiovascular health and disease. BH4 levels in the human heart have not been previously reported. Objective and Methods: Using a novel LC-MS/MS method we measured BH4 and BH2 levels in human heart tissue from subjects with (n=19) and without (n=19) coronary artery disease (CAD). Results: The concentration of BH4 was significantly lower in CAD subjects compared to controls (p=0.0006). There was a trend for a decrease in BH4/BH2 ratio (p=0.09). mRNA expression of GCH1, NOS3 and was not significantly different between CAD and controls. Conclusions: We conclude that the decrease in BH4 concentration in the left ventricular wall of subjects with CAD may be a causative factor or consequence of the failing heart.

Pretreatment with a combination of ligustrazine and berberine improves cardiac function in rats with coronary microembolization.

We have shown that a combination of ligustrazine and berberine produces more effective inhibition on platelet activation and inflammatory reactions in rat acute myocardial infarction compared with either agent alone. In this study we evaluated the beneficial effects of a combination of ligustrazine and berberine in a rat model of coronary microembolization (CME). SD rats were treated with ligustrazine, berberine, ligustrazine+berberine, or clopidogrel for 2 weeks. When the treatment completed, CME was induced by injection of sodium laurate into the left ventricular, while obstructing the ascending aorta. All rats were intubated for hemodynamic measurements. Blood samples were collected for biochemical analyses, flow cytometry, and ELISAs. Heart tissues were isolated for histopathology and subsequent protein analyses. Pretreatment with the combination of ligustrazine (27 mg·kg(-1)·d(-1)) and berberine (90 mg·kg(-1)·d(-1)) significantly improved cardiac function, and decreased myocardial necrosis, inflammatory cell infiltration, microthrombosis and serum CK-MB levels in CME rats. In addition, this combination significantly decreased plasma ET-1 levels and von Willebrand factor, inhibited ADP-induced platelet activation, and reduced TNFα, IL-1β, ICAM-1 and RANTES levels in serum and heart tissues. The protective effects of this combination were more prominent than those of ligustrazine or berberine alone, but comparable to those of a positive control clopidogrel (6.75 mg·kg(-1)·d(-1)). The combination of ligustrazine and berberine significantly improved cardiac function in rat CME model via a mechanism involving antiplatelet and anti-inflammatory effects.

Endolysosomal two‐pore channels regulate autophagy in cardiomyocytes

Two-pore channels (TPCs) were identified as a novel family of endolysosome-targeted calcium release channels gated by nicotinic acid adenine dinucleotide phosphate, as also as intracellular Na(+) channels able to control endolysosomal fusion, a key process in autophagic flux. Autophagy, an evolutionarily ancient response to cellular stress, has been implicated in the pathogenesis of a wide range of cardiovascular pathologies, including heart failure. We report direct evidence indicating that TPCs are involved in regulating autophagy in cardiomyocytes, and that TPC knockout mice show alterations in the cardiac lysosomal system. TPC downregulation implies a decrease in the viability of cardiomyocytes under starvation conditions. In cardiac tissues from both humans and rats, TPC transcripts and protein levels were higher in females than in males, and correlated negatively with markers of autophagy. We conclude that the endolysosomal channels TPC1 and TPC2 are essential for appropriate basal and induced autophagic flux in cardiomyocytes, and also that they are differentially expressed in male and female hearts. Autophagy participates in physiological and pathological remodelling of the heart. The endolysosomal two-pore channels (TPCs), TPC1 and TPC2, have been implicated in the regulation of autophagy. The present study aimed to investigate the role of TPC1 and TPC2 in basal and induced cardiac autophagic activity. In cultured cardiomyocytes, starvation induced a significant increase in TPC1 and TPC2 transcripts and protein levels that paralleled the increase in autophagy identified by increased LC3-II and decreased p62 levels. Small interfering RNA depletion of TPC2 alone or together with TPC1 increased both LC3II and p62 levels under basal conditions and in response to serum starvation, suggesting that, under conditions of severe energy depletion (serum plus glucose starvation), changes in the autophagic flux (as assessed by use of bafilomycin A1) occurred either when TPC1 or TPC2 were downregulated. The knockdown of TPCs diminished cardiomyocyte viability under starvation and simulated ischaemia. Electron micrographs of hearts from TPC1/2 double knockout mice showed that cardiomyocytes contained large numbers of immature lysosomes with diameters significantly smaller than those of wild-type mice. In cardiac tissues from humans and rats, TPC1 and TPC2 transcripts and protein levels were higher in females than in males. Furthermore, transcript levels of TPCs correlated negatively with p62 levels in heart tissues. TPC1 and TPC2 are essential for appropriate basal and induced autophagic flux in cardiomyocytes (i.e. there is a negative effect on cell viability under stress conditions in their absence) and they are differentially expressed in male and female human and murine hearts, where they correlate with markers of autophagy.

Myocardial Delivery of Lipidoid Nanoparticle Carrying modRNA Induces Rapid and Transient Expression.

Nanoparticle-based delivery of nucleotides offers an alternative to viral vectors for gene therapy. We report highly efficient in vivo delivery of modified mRNA (modRNA) to rat and pig myocardium using formulated lipidoid nanoparticles (FLNP). Direct myocardial injection of FLNP containing 1-10 μg eGFPmodRNA in the rat (n = 3 per group) showed dose-dependent enhanced green fluorescent protein (eGFP) mRNA levels in heart tissue 20 hours after injection, over 60-fold higher than for naked modRNA. Off-target expression, including lung, liver, and spleen, was <10% of that in heart. Expression kinetics after injecting 5 μg FLNP/eGFPmodRNA showed robust expression at 6 hours that reduced by half at 48 hours and was barely detectable at 2 weeks. Intracoronary administration of 10 μg FLNP/eGFPmodRNA also proved successful, although cardiac expression of eGFP mRNA at 20 hours was lower than direct injection, and off-target expression was correspondingly higher. Findings were confirmed in a pilot study in pigs using direct myocardial injection as well as percutaneous intracoronary delivery, in healthy and myocardial infarction models, achieving expression throughout the ventricular wall. Fluorescence microscopy revealed GFP-positive cardiomyocytes in treated hearts. This nanoparticle-enabled approach for highly efficient, rapid and short-term mRNA expression in the heart offers new opportunities to optimize gene therapies for enhancing cardiac function and regeneration.

The H2S Donor NaHS Changes the Expression Pattern of H2S-Producing Enzymes after Myocardial Infarction.

Aims. To examine the expression patterns of hydrogen sulphide- (H2S-) producing enzymes in ischaemic heart tissue and plasma levels of H2S after 2 weeks of NaHS treatment after myocardial infarction (MI) and to clarify the role of endogenous H2S in the MI process. Results. After MI surgery, 2 weeks of treatment with the H2S donor NaHS alleviated ischaemic injury. Meanwhile, in ischemia myocardium, three H2S-producing enzymes, cystathionine γ-lyase (CSE), cystathionine-β-synthase (CBS), and 3-mercaptopyruvate sulfurtransferase (3-MST) significantly increased. Plasma H2S levels were also elevated. In vitro, NaHS treatment protected cardiomyocytes from hypoxic injury and raised CBS levels in a concentration-dependent manner. Different from in vivo results, however, CSE or 3-MST expression did not change. NaHS treatment increased the activity of CSE/CBS but not of 3-MST. When CSE was either knocked down (in vitro) or knocked out (in vivo), H2S levels significantly decreased, which subsequently exacerbated the ischaemic injury. Meanwhile, the expressions of CBS and 3-MST increased due to compensation. Conclusions. Exogenous H2S treatment changed the expressions of three H2S-producing enzymes and H2S levels after MI, suggesting a new and indirect regulatory mechanism for H2S production and its contribution to cardiac protection. Endogenous H2S plays an important role in protecting ischaemic tissue after MI.

Chronic Aerobic Exercise Decreases Lectin-Like Low Density Lipoprotein (LOX-1) Receptor Expression in Heart of Diabetic Rat.

Background:Overexpression of lectin-like low density lipoprotein (LOX-1) receptor plays an important role in hyperglycemia-induced vascular complications such as atherosclerosis. Based on the beneficial effects of exercise on preventing cardiovascular complications of diabetes, we aimed to examine the protective effects of aerobic exercise on expression of LOX-1 receptor and production of free radicals in the heart of diabetic rats. Methods:Four groups of rats were used: (n = 5 per group): sedentary normal, trained normal, sedentary diabetes and trained diabetes. Diabetes was induced by a single intraperitoneal injection of streptozotocin (50 mg/kg). The exercise protocol was consisted of swimming 30 min/day, 5 days/week for eight weeks. Plasma glucose was evaluated at initiation, weeks 4 and 8 of experiment. At the end of experiment, rats were sacrificed and the heart was removed for determination of nitrate, malondialdehyde, and LOX-1 gene expression. Results:In normal non-diabetic rats, the blood glucose level was <150 mg/dl; however, the induction of diabetes resulted in levels more than >400 mg/dl. Gene expression of LOX-1 was increased in the heart of diabetic rats. Exercise reduced the gene expression of this protein in diabetic states without reducing the blood glucose. Finally, swimming exercise decreased the malondialdehyde and nitrate levels in heart tissue both in control and diabetic rats. Conclusion:Swimming exercise reduces heart expression of the LOX-1 receptor in accompany with reduction of free radicals production. Since these parameters are important in generation of diabetic complications, swimming exercise is a good candidate for reducing these complications.

Comparison of the acute effects of benzo-a-pyrene on adult zebrafish (Danio rerio) cardiorespiratory function following intraperitoneal injection versus aqueous exposure

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental contaminants. PAH exposure causes developmental toxicity in multiple fish species, while acute adult fish toxicity is thought to be minimal. The literature increasingly suggests sublethal PAH effects may occur, but differences in exposure route may confound conclusions. We hypothesized that acute PAH exposure in adult fish will cause cardiorespiratory impairment that will not differ with exposure route. In order to investigate this hypothesis, adult zebrafish (Danio rerio) were injected intraperitoneal (i.p.) twice with increasing concentrations of the prototypical PAH, benzo-a-pyrene (BaP; 0.1, 10, and 1000μg/kg) or exposed aqueously (static, renewal at 24h; 16.2 and 162μg/L) for 48h and compared to corresponding dimethylsulfoxide controls. No mortalities or significant effects on weight of the fish were noted at any exposure concentration or route. At 48h, fish were subjected to swimming tests with concurrent oxygen consumption measurement (n=10 fish/treatment) or echocardiography (n=12 fish/treatment). Oxygen consumption (MO2) was increased at three swimming speeds in BaP-injected groups compared to control (p<0.01 in Fisher’s LSD tests after two-way ANOVA). In contrast, aqueously BaP-exposed fish showed increased MO2 under only basal conditions. Despite increased oxygen demand, ventricular heart rate was significantly decreased in BaP-exposed fish, both injected and aqueously-exposed. Analysis of BaP body burdens in fish tissue allowed for identification of an overlapping dose group between exposure routes, through which comparisons of cardiorespiratory toxicity were then made. This comparison revealed most effects were similar between the two exposures routes, although minor differences were noted. At similar BaP body burdens, injected fish suffered from more severe bradycardia than aqueously exposed fish and had greater levels of increases in cytochrome P4501A (CYP1A) mRNA levels in liver and heart tissue compared to aqueous exposed fish. In conclusion, acute BaP exposure in adult zebrafish had negative effects on cardiorespiratory function. Differences in effect between exposure routes were attributed primarily to differences in bioavailability, since overall, similar effects were noted between the two exposure routes when similar BaP body burdens were achieved.

Abstract 608: A locus on Chromosome 6 in Diversity Outbred Mice Suggests Osteogenic Regulation of Dystrophic Cardiac Calcinosis

Introduction: Dystrophic cardiac calcinosis (DCC) is an age-related cardiomyopathy that involves myocardial injury, necrosis, and calcification. The genetic factors contributing to myocardial calcification are complex, and several loci for DCC have been identified. However, only 1 gene, Abcc6, has been cloned and confirmed to regulate DCC. Our current studies were designed to study the genetic architecture of DCC. Methods and Results: Recently, the Diversity Outbred (DO) population was developed from 8 inbred strains of mice. The DO mice are mosaics of 8 progenitor strains: C57BL6/J, A/J, NOD/ShiLtJ, NZO/HiLtJ, WSB/EiJ, CAST/EiJ, PWK/PhJ and 129S1/SvImJ. Our initial studies examined the effects of diet and genetic background on the development of DCC in the 8 progenitor strains. At 24 weeks of age, the hearts of mice from these 8 strains were collected, and a colorimetric assay was used to measure calcium levels in heart tissue. ANOVA tests indicated a significant effect (p &lt; 0.05) of the underlying genetics on DCC. To identify genes and pathways contributing to DCC, we next performed QTL mapping using 300 DO mice. Our QTL mapping analysis was carried out using a genetic model that incorporates reconstructed haplotypes and accounts for population structure. We identify a significant (LOD 7.10) QTL on chromosome 6 (94.7 Mb - 97.3 Mb) associated with DCC. Detailed analysis identified the CAST/EiJ and PWK/PhJ founder alleles as the greatest genetic contributors to the chromosome 6 peak. Myocardial calcification involves a highly regulated process of mineralization similar to osteogenesis, and several of the positional candidates mediate calcification, including: Fam19a4, Fam19a1, Eogt, and Arl6ip5. Conclusions: This data identifies genes that may regulate the mineralization underlying both DCC and osteogenesis. They may yield new therapeutic targets for DCC and serve as indicators of atherosclerosis.

Cardiac H2S Generation Is Reduced in Ageing Diabetic Mice.

Aims. To examine whether hydrogen sulfide (H2S) generation changed in ageing diabetic mouse hearts. Results. Compared to mice that were fed tap water only, mice that were fed 30% fructose solution for 15 months exhibited typical characteristics of a severe diabetic phenotype with cardiac hypertrophy, fibrosis, and dysfunction. H2S levels in plasma, heart tissues, and urine were significantly reduced in these mice as compared to those in controls. The expression of the H2S-generating enzymes, cystathionine γ-lyase and 3-mercaptopyruvate sulfurtransferase, was significantly decreased in the hearts of fructose-fed mice, whereas cystathionine-β-synthase levels were significantly increased. Conclusion. Our results suggest that this ageing diabetic mouse model developed diabetic cardiomyopathy and that H2S levels were reduced in the diabetic heart due to alterations in three H2S-producing enzymes, which may be involved in the pathogenesis of diabetic cardiomyopathy.