β2-AR mRNA Research Articles

Effects of Weight Loss and Aerobic Exercise Training on Adi-Pose Tissue Zinc α2-Glycoprotein and Associated Genes in Obesity.

Zinc α2-glycoprotein (ZAG) has been implicated in fatty acid metabolism and utilization and is lower in obese and higher in cachexic adults compared to those of normal weight. Previous studies suggest that ZAG binds to the beta3-adrenergic receptor (β3AR) to influence fatty acid metabolism in adipose tissue by regulating hormone sensitive lipase (HSL). The purpose of this study is to investigate the effects of a six-month weight loss (WL) or aerobic exercise (AEX) intervention on adipose tissue and skeletal muscle ZAG mRNA levels and protein expression, as well as the expression of β3AR, and HSL. Abdominal adipose tissue (AB) and gluteal adipose tissue (Glut) and vastus lateralis muscle biopsies were performed before and after WL (n = 13) or AEX (n = 13). ZAG, HSL, and β3AR expressions were determined by RT-PCR, and ZAG and HSL plasma levels by ELISA. Body weight decreased by 9.69% (p < 0.001) in WL and did not change with AEX. Maximal oxygen consumption (VO2max) increased by 7.1% (p < 0.005) after WL and by 16.69% (p < 0.001) after AEX. WL significantly decreased body weight with a reduction of percentage of fat, fat mass, fat-free mass (FFM). AEX decreased percent fat and increased VO2max, but did not change fat mass and FFM. Abdominal ZAG and HSL mRNA levels did not change significantly after WL or AEX. There were no changes in plasma ZAG, HSL and adipose tissue β3AR mRNA levels after WL and AEX. ZAG, HSL and β3AR mRNA expressions in adipose tissue are positively associated each other. Adipose tissue abdominal and gluteal HSL are negatively associated with HOMA-IR (Homeostatic Model Assessment for Insulin Resistance), and both ZAG and HSL adipose tissue are negatively associated with fasting glucose and the glucose area under the curve. Further work is needed to elucidate the role of ZAG and HSL in the propensity for weight gain and the ability of exercise to mitigate these responses.

Expression pattern and clinical significance of beta 2-adrenergic receptor in oral squamous cell carcinoma: an emerging prognostic indicator and future therapeutic target.

Beta 2-AdrenergicReceptor (β2-AR) is significantly overexpressed in various types of malignancies, which is associated with the worst prognosis. However, the role of β2-AR in oral cancer is not well identified. The present study aimed at investigating the β2-AR gene expression and its significance in relation withthe clinicopathological features and overall survival of oral squamous cell carcinoma (OSCC) patients. Immunohistochemistry, western blot and quantitative real-time PCR techniques were used to analyze β2-AR protein and mRNA levels in a total of 65 histopathologically confirmed OSCC tissues (case group) and 65 normal tissues (control group) from the oral cavity. Out of the total of 65 OSCC tissues, 41 tissues (63.1%) exhibited high expression for β2-AR protein. Percent positivity and relative density (mean ± SD) of protein were higher in the case group ascompared to the control group (positivity 40.31 ± 3.01 vs. 20.46 ± 1.93, p < 0.001; density 2.77 ± 1.17 vs. 1.28 ± 0.37, p < 0.001). In addition, β2-AR mRNA level was also upregulated in patients comparedtothe controls (2.36 ± 1.30 vs. 1.09 ± 0.42, p < 0.001) and showed a positive correlation with immunostaining of protein in OSCC (r = 0.48, p = 0.011). High β2-AR protein expression was significantly associated with multiple risk habits (p = 0.045), histological differentiation (p = 0.013), clinical TNM stages (p = 0.014), and poor survival (p = 0.006) of patients. In the Cox proportional hazards model, β2-AR was identified as a prognostic biomarker of OSCC (p = 0.047). β2-AR protein level is identified as an independent significant prognostic factor in patients with oral carcinoma.

Abstract 9084: Selective Beta3-Adrenergic Receptor Antagonist Therapy Causes Regression of Cardiac Dysfunction in a Rat Model of Progressive Heart Failure: Insights into Molecular and Cellular Mechanisms

Background: Heart failure (CHF) causes cardiac β 3 -adrenergic receptor (AR) upregulation with resultant enhanced β 3 -AR-stimulated-G i -coupled negative modulation on cardiac function. This suggests a direct and contributing role of β 3 -AR activation on CHF progression. However, its precise role is unclear. We hypothesize that up-regulation of cardiac β 3 -AR promotes CHF progression. Chronic β 3 -AR antagonist (ANT) may improve Left ventricular (LV) myocyte function, and β-adrenergic reserve, thus preventing the progression of CHF. Methods: LV and myocyte functional response and β 1 - and β 3 -AR expression were compared in 3 groups of SD male rats (8 each) during 3 months: 1) CHF, 3 months after receiving Isoproterenol (ISO, 170 mg/kg, sq, for 2 days); 2) CHF/β 3 -ANT, 2 months after receiving ISO, L-748,337, a selective β 3 -AR antagonist (β 3 -ANT) (10 -7 M/kg/day, mini pump) was initiated and was given for 1 month; and 3) sham controls. Results: Versus controls, ISO-treated rats had CHF onset at 1 month after ISO and progressed to severe CHF at 3 months. Plasma norepinephrine (NE, 1298 vs 261 pg/ml) increased 4-fold; whereas, stroke volume (SV) and ejection fraction (EF, 34 vs 62%) decreased more than 45%. These changes were parallel with significant reductions in cell contraction (dL/dt max , 95 vs 189 μm/s), relaxation (dR/dt max ,75 vs 158 μm/s) and [Ca 2+ ] i transient ([Ca 2+ ] iT ) (0.17 vs 0.26). Acute superfusion of ISO (10 -8 M) caused much less increases in dL/dt max (34 vs 74%), dR/dt max and [Ca 2+ ] IT (14 vs 28%). These changes were associated with significantly decreased β 1 -AR mRNA (52%, 0.30 vs 0.62), but increased β 3 -AR mRNA (119%, 1.25 vs 0.57) expression. CHF with β 3 -ANT treatment, significantly decreased plasma NE (197 pg/ml). EF (59%) and SV returned to normal control values. The signal ratios of β 1 -AR mRNA (0.59) and β 3 -AR mRNA (0.66) recovered to control levels. ISO-induced increases in dL/dt max (75%), dR/dt max and [Ca 2+ ] iT were also significantly augmented. Conclusion: Chronic β 3 -ANT leads to normalization of LV myocyte β 1 - and β 3 -AR gene expression, restores normal LV myocyte β-AR reserve, and leads to regression of LV and myocyte dysfunction in a rat model of progressive CHF. Thus, β 3 -AR blocker may provide a new therapeutic strategy for the treatment of CHF.

Adrenergic receptor behaviors of mesenchymal stem cells obtained from different tissue sources and the effect of the receptor blockade on differentiation

In this study, it was aimed to analyze behavioral changes of adrenergic receptors (ARs) in first three passages and osteogenic/adipogenic differentiation of mesenchymal stem cells (MSCs) derived from placenta fetal membrane (FM) and bone marrow (BM). It was also aimed to evaluate effects of receptor blockade on differentiation. We obtained first three passages of MSCs from placenta and BM samples. For cell identification, the cells were analyzed by flow cytometry using CD34, CD45 and CD3, CD105 antibodies in each passage. The effects of propranolol and phenoxybenzamine at incremental doses were analyzed by MTT. In addition, cell cultures were separately maintained with the blockers or without after second passage. After each passage and differentiation, α1A, α1B, α2A, α2B, β1, β2, β3 AR-mRNA expressions analyzed by RT-qPCR technique. BMP6 and PPARG mRNA expressions only after differentiation and passage 3 were analyzed. A microscopic examination was also performed. Our results showed that AR expression behaviors were different in MSCs obtained from different tissue sources. In particular, α1A-AR and α2A-AR were expressed with considerably high coefficients in differentiation under blocker effect in BM-derived MSCs. No such coefficients were observed in any group of placental MSCs. In addition, it was found that the blockers stimulated adipogenesis in BM-derived MSCs during osteogenic differentiation. MSCs exhibit protein expressions that vary according to source of tissue and differentiation. Given that MSCs from different sources are used for repair and modulation, our study makes implications of this variable expression intriguing in the clinical practice.

Disruption of STAT6 Signal Promotes Cardiac Fibrosis Through the Mobilization and Transformation of CD11b+ Immature Myeloid Cells.

Cardiac fibrosis is an important pathological basis of various cardiovascular diseases. The roles of STAT6 signal in allergy, immune regulation, tumorigenesis, and renal fibrosis have been documented. However, the function and mechanism of STAT6 signal in sympathetic overactivation-induced cardiac fibrosis have not been fully elucidated. This study explores the novel role of STAT6 signal in isoproterenol (ISO)–induced cardiac fibrosis through the regulation of inflammatory response and the differentiation of macrophages from immature myeloid cells. The expression levels of STAT6, β1-adrenergic receptor (β1-AR), and inflammatory factors [interleukin α (IL-1α), IL-6, IL-18, and transforming growth factor β (TGF-β)] in CD11b+ myeloid cells were analyzed with a microarray study. The levels of IL-6 and TGF-β1 in the CD11b+ myeloid cells–derived macrophages were detected with reverse transcriptase–polymerase chain reaction (RT-PCR). STAT6–knockout (KO) and WT mice were used to establish a murine cardiac fibrosis model by ISO injection. Cardiac fibroblasts were isolated from the hearts of newborn STAT6-KO and WT mice, and STAT6 expression was measured by Western blotting and RT-PCR after ISO stimulation, while α-smooth muscle actin (α-SMA) expression was detected by immunofluorescence and immunohistochemistry staining. Cardiac function and pathological characteristics were examined by echocardiography and immunohistochemistry staining, respectively. Immunohistochemistry staining with anti-CD11b was performed to detect the infiltration of CD11b+ myeloid cells in heart tissue. Flow cytometry analysis was used to measure the percentages of CD11b+ myeloid cells and CD11b+Ly6C+ macrophages in the peripheral blood. The results showed that STAT6 was highly expressed in CD11b+ myeloid cells located in injured hearts, and STAT6 expression in cardiac fibroblasts was down-regulated after ISO treatment. STAT6 deficiency further aggravated ISO-induced increased expression of α-SMA in cardiac fibroblasts, myocardial fibrosis, and cardiac dysfunction. The activation of ISO/β1-AR signal aggravated cardiac inflammatory infiltration, promoted CD11b+ myeloid cell mobilization, and enhanced CD11b+Ly6C+/low macrophage differentiation, which was further exacerbated by STAT6 deficiency. Furthermore, β1-AR mRNA expression significantly increased in splenic CD11b+ myeloid cells compared to their bone marrow–derived controls, and STAT6 deficiency promoted β1-AR expression in an MI-induced sensitive cardiac fibrosis mouse model. The spleen-derived CD11b+ myeloid cells of STAT6-KO mice produced more IL-1α, IL-18, and TGF-β than their WT counterparts. Taken together, these results suggest that STAT6 signal plays a critical role in ISO-induced β1-AR overactivation and systemic inflammatory cascades, contributing to cardiac fibrogenesis. STAT6 should be a promising cardioprotective target against myocardial fibrosis and heart failure after β1-AR overactivation–induced myocardial injury.

PSXIV-33 α-chaconine induces myogenesis of bovine satellite cells but less affect adipogenesis of adipocytes

Abstract α-chaconine is a steroidal glycoalkaloid compound found in leaves, fruit and tubers of the solanaceae family such as potato, tomato, and eggplant. α-chaconine poisoning was reported in various animal models such as mice, rabbit, and chicken. However, previous study indicated that it is used to induce performance of beef cattle. We hypothesized that α-chaconine may affect myogenesis of bovine satellite cells but less affect adipogenesis of bovine adipocytes. Bovine satellite cells (BSC) were pronase-liberated from semimembranosus (SM) and longissimus dorsi (LD) muscle tissues of three new-born Hanwoo calves. Bovine preadipocytes were isolated from perirenal (PR) adipose tissues. BSC and preadipocytes were incubated with dulbecco modified eagle medium (DMEM) with 10% fetal bovine serum for proliferation. BSC induced differentiation with DMEM with 3% horse serum and preadipocytes induced differentiation with DMEM with insulin, dexamethasone, and ciglitazone. Cells were treated with various levels of α-chaconine (control, 0.001, 0.01, 0.1, 1, and 10μM). mRNA abundance for myosin heavy chain 2X (MHC2X), glucose transportor 4 (GLUT4), β2-adrenergic receptor (β2-AR), peroxisome proliferator-activated receptor γ (PPARγ), stearoyl-CoA desaturase (SCD) were measured by real-time quantitative PCR. Data were analyzed as a completely randomized design using the MIXED model, each treatment performed in triplicated. Means were considered different at P &amp;lt; 0.05. Relative MHC2X mRNA abundance was greater in BSC with dose-titrated chaconine treatments compared to the control (P &amp;lt; 0.05). Relative levels of β2-AR were greater in both BSC and adipocytes based on dose-dependent treatements (P &amp;lt; 0.05). The levels of PPARγ and SCD have no dose-titrated treatments in bovine PR adipocytes (P &amp;gt; 0.05). These result indicated that α-chaconine have a dose-dependent effect on MHC2X and β2-AR mRNA BSC and adipocytes but did not affect to adipogenesis in bovine PR adipocytes.

HuR regulates phospholamban expression in isoproterenol-induced cardiac remodelling.

The elevated expression of phospholamban (PLB) has been observed in heart failure and cardiac remodelling, inhibiting the affinity of Ca2+ pump to Ca2+ thereby impairing heart relaxation. However, the mechanisms underlying the regulation of PLB remains to be further studied. The present study aims to test the role of RNA-binding protein HuR in the regulation of PLB and the impact of this regulatory process in cardiac remodelling. A mouse model specifically deleted HuR in cardiomyocytes were used for testing the role of HuR in regulating PLB during isoproterenol (ISO)-induced cardiac remodelling. HuR deficiency did not significantly influence the phenotype and function of mouse heart under static status. However, deletion of HuR in cardiomyocytes mitigated the effect of ISO in inducing PLB expression and reducing β1-AR expression, in turn aggravating ISO-induced myocardial hypertrophy and cardiac fibrosis. In H9C2 cells, association of HuR with PLB and β1-AR mRNAs stabilized PLB mRNA and destabilized β1-AR mRNA, respectively. HuR stabilizes PLB mRNA and destabilizes β1-AR mRNA. The HuR-PLB and HuR-β1-AR regulatory processes impact on ISO-induced cardiac remodelling.

Dysregulated megakaryocyte distribution associated with nestin+ mesenchymal stem cells in immune thrombocytopenia

AbstractImpaired megakaryocyte (MK) maturation and reduced platelet production are important causes of immune thrombocytopenia (ITP). However, MK distribution and bone marrow (BM) niche alteration in ITP are unclear. To investigate the maturation and distribution of MKs in the BM niche and examine the components of BM niche regulation of MK migration, BM and peripheral blood were obtained from 30 ITP patients and 28 healthy donors. Nestin+ mesenchymal stem cells (MSCs) and CD41+ MKs were sorted by fluorescence-activated cell sorting. The components of the BM niche and related signaling were analyzed via immunofluorescence, flow cytometry, enzyme-linked immunosorbent assay, reverse transcription polymerase chain reaction, and western blot analysis. The number of MKs in the BM vascular niche was reduced in ITP. Moreover, the concentrations of CXCL12 and CXCR4+ MKs in the BM were decreased in ITP. Further investigation demonstrated that nestin+ MSCs and CXCL12 messenger RNA (mRNA) in nestin+ MSCs were both reduced whereas the apoptosis of nestin+ MSCs was significantly increased in ITP. Sympathetic nerves, Schwann cells, the proportion of β3-adrenoreceptor (β3-AR)+ nestin+ MSCs, and β3-AR mRNA in nestin+ MSCs were all markedly reduced in ITP. Moreover, matrix metalloproteinase 9, vascular endothelial growth factor (VEGF), and VEGF receptor 1 were significantly reduced in ITP. Our data show that impaired MK distribution mediated by an abnormal CXCL12/CXCR4 axis is partially involved in reduced platelet production in ITP. Moreover, sympathetic neuropathy and nestin+ MSC apoptosis may have an effect on the alterations of BM CXCL12 in ITP.

Cardiovascular variability and β-ARs gene expression at two stages of doxorubicin – Induced cardiomyopathy

Using comprehensive analysis of heart rate (HRV) and blood pressure (BPV) short-term variability we estimated the time course of changes of autonomic nervous system remodeling in two stages of doxorubicin-induced cardiomyopathy (DCM). We also investigated the level of gene expression of cardiac β-1 (β-1AR) and β-2 (β-2AR) adrenoceptors. Experiments were performed in adult male Wistar rats equipped with indwelling catheters for BP recording and blood withdrawal. A 15 mg/kg total cumulative dose of doxorubicin was injected i.p. to rats to induce DCM or saline for control (n=18). Rats were assessed for general toxicity, cardiovascular hemodynamic and echocardiography before treatment (n=6), 35 days (DOX35; n=6) and 70 days (DOX70; n=6) post-treatment. HRV was evaluated by spectral analysis, Poincaré plots, sample and approximate entropy. Expression of β-1AR and β-2AR mRNA was evaluated by RT-qPCR. Doxorubicin-treated rats exhibited poor general condition and lower survival than saline-treated rats. In DOX35 rats, there were no echocardiography signs of decompensation, no increase in serum cardiac troponins, but there was an increase of HRV and decrease of HR complexity. In these rats typical microscopic signs of cardiotoxicity were seen along with over-expression of β-1AR mRNA. 70 days post-treatment echocardiography revealed signs of decompensation and serum cardiac troponin T was increased. At this stage BPV decreased. In conclusion, HRV increase matches transient over-expression of cardiac β-1AR mRNA in compensate stage of DCM while decompensate stage of DCM is characterized by a decrease of BPV and no changes in β-1AR and β-2AR gene expression.

A3707 Myocardial microRNA-101 downregulates classical B1-adrenoreceptor signing cascade resulting in improving the cardiac function

Objectives: We tested the hypothesis that the miR-101 down-regulates β1-AR expression and signaling resulting in improving the cardiac function. Methods: Bioinformatic analysis andhe dual luciferase assays was used to determine whether miR-101 target β1-AR mRNA. Through miR-101 mimic or inhibitor in H9C2 cell, the expression of β1-AR and protein kinase A (PKA), PKA phosphorylation (p-PKA) were tested by western blot. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to determine miR-101 expression profiles in the heart of spontaneously hypertensive rats (SHR) and age-matched Wistar-Kyoto (WKY) rats (n = 6, respectively). Rats were treated with miR-101 agomiR or antagomiR to build a miR-101-upregulated/ downregulated animal model. Cardiac function and HR were assessed by ultrasound diagnostic instrument, systolic blood pressure (SBP) was determined by non-invasive blood pressure analyzer, the expression of β1-AR, PKA, p-PKA protein were detected. Results: I The dual luciferase assays confomed that β1-AR is a direct target gene of miR-101. In vitro study, the expression level of β1-AR, PKA, p-PKA protein were decreased in H9C2 cell transfected with miR-101 mimic (p < 0.05). Meanwhile the expression level of β1-AR expression, PKA protein, p-PKA protein elevated in both H9C2 cell transfected with miR-101 inhibitor; In vivo study, the level of myocardial miR-101 expression were decreased in SHR rats compared to β1-AR (−/ − ) rats and WKY rats (p < 0.05). Elevated of β1-AR protein, PKA protein, p-PKA protein, cAMP level as well as BP, EF, FS in SHR rats was descresed by miR-101 mimic transfection. (p < 0.05). Conclusion: Myocardial microRNA-101 downregulates classical β1-adrenoreceptor signing cascade resulting in improving the cardiac function

The β2-adrenergic receptor is involved in differences in the protein degradation level of the pectoral muscle between fast- and slow-growing chicks during the neonatal period

The aim of this study was to investigate whether β2-AR mRNA expression is involved in either atrogin-1/MAFbx mRNA expression or protein degradation in chicken skeletal muscle by comparing fast- and slow-growing chicks during the neonatal period. Based on their body weight gain from 1 to 5 days of age, 5-day-old chicks (Gallus gallus domestics) were divided into a slow-growing and a fast-growing group, the mean weight gains of which were 6.3 ± 1.3 g/day and 11.3 ± 0.9 g/day, respectively. The ratio of pectoral muscle weight to total body weight was higher in the fast-growing group of chicks than in the slow-growing group. In addition, the plasma 3-methylhistidine concentration, an index of protein degradation in skeletal muscle, was significantly lower in the fast-growing than in the slow-growing chicks. The mRNA expression of β2-AR, which we previously found is involved in decreasing muscle protein degradation by suppression atrogin-1/MAFbx mRNA expression, was significantly higher in the pectoral muscle of the fast-growing group compared with that of the slow-growing group. Concordantly, lower mRNA expression of atrogin-1/MAFbx was observed in the pectoral muscle of the fast-growing chicks. However, in the sartorius muscle, which is a muscle in the thigh, the ratio of the muscle weight to total body weight was not significantly different between the two groups of chicks at 5 days of age. In addition, there was no significant difference in the mRNA expressions of β2-AR and atrogin-1/MAFbx in the sartorius muscle between these two groups. These results suggest that β2-AR expression levels might be physiologically significant in the control of protein degradation in the pectoral muscle of neonatal chicks.

Characterization of a β2 adrenergic receptor protein precursor in the European eel (Anguilla anguilla) and its tissue distribution across silvering

This study provides the characterization and tissue distribution of a β2-AR in the female European eel during silvering, aiming to better understand the adrenergic system involvement in this critical maturation event. A putative β2-AR (ADRB2) mRNA was cloned and sequenced. Amino acid residues and motifs important for ligand binding are generally conserved across fish and between fish and mammals, although the occurrence of some sequence variabilities may explain the noted peculiarities of eel AR interaction with pharmacological ligands. The tissue distribution of the ADRB2 gene product was analyzed in five tissues of the eel at different silvering stages and compared with that of the ADRA1 mRNA encoding an α1-AR subtype. On the whole, data suggested that relative ADRA1/ADRB2 tissue expression across silvering is part of the preparatory (molecular) adjustments required to face changes in habitats and migration efforts.

Effects of mild hypothermia on β-adrenergic signaling pathway in a cardiac arrest swine model

To observe the effect of mild hypothermia on myocardial β-adrenergic receptor (β-AR) signal pathway after cardiopulmonary resuscitation (CPR) in pigs with cardiac arrest (CA) and explore the mechanism of myocardial protection. Healthy male Landraces were collected for reproducing the CA-CPR model (after 8-minute untreated ventricular fibrillation, CPR was implemented). The animals were divided into two groups according to random number table (n = 8). In the mild hypothermia group, the blood temperature of the animals was induced to 33 centigrade and maintained for 6 hours within 20 minutes after return of spontaneous circulation (ROSC) by using a hypothermia therapeutic apparatus. In the control group, the body temperature of the animals was maintained at (38.0±0.5)centigrade with cold and warm blankets. The heart rate (HR), mean arterial pressure (MAP), the maximum rate of increase or decrease in left rentricular pressure (+dp/dt max) were measured during the course of the experiment. The cardiac output (CO) was measured by heat dilution methods before CA (baseline), and 0.5, 1, 3, 6 hours after ROSC respectively, the venous blood was collected to detect the concentration of cTnI. Left ventricular ejection fraction (LVEF) was measured with cardiac ultrasound before CA and 6 hours after ROSC. Animals were sacrificed at 6 hours after ROSC and the myocardial tissue was harvested quickly, the mRNA expression of β1-AR in myocardium was detected by reverse transcription-polymerase chain reaction (RT-PCR), the contents of adenylate cyclase (AC) and cyclic adenosine monophosphate (cAMP) were detected by enzyme linked immunosorbent assay (ELISA), the protein content of G protein-coupled receptor kinase 2 (GRK2) was detected by Western Blot. After successful resuscitation, the HR of both groups were significantly higher than the baseline values, CO, ±dp/dt max were significantly decreased, MAP were not significantly changed, serum cTnI levels were significantly increased. Compared with the control group, HR at 0.5, 1, 3 hours after ROSC were significantly decreased in mild hypothermia group (bpm: 142.80±12.83 vs. 176.88±15.14, 115.80±11.48 vs. 147.88±18.53, 112.60±7.40 vs. 138.50±12.02, all P < 0.01), CO was significantly increased at 1 hours and 3 hours after ROSC (L/min: 3.97±0.40 vs. 3.02±0.32, 4.00±0.11 vs. 3.11±0.59, both P < 0.01), +dp/dt max at 3 hours and 6 hours was also significantly increased after ROSC [+dp/dt max (mmHg/s): 3 402.5±612.7 vs. 2 130.0±450.6, 3 857.5±510.4 vs. 2 562.5±633.9; -dp/dt max (mmHg/s): 2 935.0±753.2 vs. 1 732.5±513.6, 3 520.0±563.6 vs. 2 510.0±554.3, all P < 0.05], the cTnI was significantly decreased at 3 hours and 6 hours afher ROSC (μg/L: 1.39±0.40 vs. 3.24±0.78, 1.46±0.35 vs. 3.78±0.93, both P < 0.01). The left at 6 hours after ROSC in both groups was decreased as compared with that before CA. The LVEF in the mild hypothermia group was higher than that in the control group (0.52±0.04 vs. 0.40±0.05, P < 0.05). The mRNA expression of β1-AR, and concentrations of AC and cAMP in hypothermia group were significantly higher than those in control group [β1-AR mRNA (2-ΔΔCT): 1.18±0.39 vs. 0.55±0.17, AC (ng/L): 197.0±10.5 vs. 162.0±6.3, cAMP (nmol/L): 1 310.58±48.82 vs. 891.25±64.95, all P < 0.05], GRK2 was lower than that in the control group (GRK2/GAPDH: 0.45±0.05 vs. 0.80±0.08, P < 0.05). Mild hypothermia can reduce the degree of cardiac function injury after CPR, and its mechanism may be related to the reduction of impaired myocardial β-AR signaling after CPR.

The impact of apelin-36 on isolated rat hearts as a member of apelin family.

Apelinis an endogenousadipocytokine thatplays an important role in the regulation of cardiovascular function. Apelin-36 is a member of apelin family. However cardiac reports related to apelin-36 are very rare. The purpose of this study is to investigate the impact of apelin-36 onhemodynamic variables of the isolated rat hearts. Twenty-eight ratswere equally divided into four groups: control, apelin-36 at the following concentrations: 1 nM, 10 nM and 100 nM. Theisolatedhearts were perfused with modified Krebs-Henseleit solution (mK-Hs) by using theLangendorffmethod. Cardiac parameters, including left ventricular developed pressure (LVDP),maximal rate of pressure development (+dP/dtmax), heartrate (HR) and coronary flow (CF) were measured. Gene expressions relevant to cardiomyocyte contractility were determined by real-time PCR. 10 and 100 nM doses of apelin-36 perfusion led topositive inotropy with an increase of LVDP and +dP/dtmax, which are the indexes of cardiac contractility. Furthermore both doses of apelin-36 increased endothelial nitric oxide synthase (eNOS), sarco/endoplasmic reticulum Ca2+-ATPase (Serca2a) andβ2-Adrenergic receptors (β2-AR) mRNA. These results showed that apelin-36 had a positive inotropic effect on the isolated rat heart and can induceeNOS,Serca2aandβ2-ARgenes activation that enhancescontractilityof theheart (Tab. 1, Fig. 2, Ref. 23).

(-)-Epigallocatechin-3-gallate, the major green tea catechin, regulates the desensitization of β1 adrenoceptor via GRK2 in experimental heart failure.

We investigated the effect of (-)-epigallocatechin-3-gallate (EGCG) on cardiac function and its mechanism, which focused on the desensitization of β1-AR and GRK2 in heart failure (HF) rats. HF was induced by abdominal aortic coarctation. Four weeks after HF induction, the rats were given EGCG (25, 50, 100mg/kg/day). Cardiac function was assessed by measuring haemodynamic parameters. Histological changes were analyzed by HE and Masson's trichrome staining. The expression of β1-AR was detected by immunohistochemistry and immunofluorescence. The membrane expression of β1-AR and GRK2 was detected by western blot. The expression levels of β1-AR mRNA and GRK2 mRNA were evaluated by Q-PCR. Compared to the control group, the left ventricular end diastolic pressure, mean blood pressure, heart weight/body weight, and posterior wall thickness in the HF group were significantly increased, whereas the left ventricular systolic pressure, maximum rate of left ventricular pressure rise (+dP/dt max) and maximum rate of left ventricular pressure fall (-dP/dt max) were clearly decreased. EGCG could improve cardiac function by regulating these parameters. Inflammatory cell infiltration, irregularly arranged cardiomyocytes, swelling of cardiomyocytes and myocardial fibrosis were observed in HF rats' myocardial morphology, and EGCG obviously improved the morphological signs. The expression of β1-AR was significantly decreased in the left ventricle tissue of HF rats by immunohistochemistry and immunofluorescence. The membrane expression of β1-AR decreased, whereas GRK2 increased in vivo and in vitro by western blot. EGCG could down-regulate the membrane expression of GRK2 and up-regulate the expression of β1-AR. There were no significant differences in the total expression of β1-AR mRNA and GRK2 mRNA. EGCG has therapeutic effects on the heart function of HF rats. The mechanism might be related to the inhibition of the transfer membrane of GRK2 and to the reduction of the desensitization of β1-AR.

Effect of electroacpuncture of Neiguan (PC 6) and Tianquan (PC 2) on skin temperature, blood perfusion, and adrenoceptor response in rats with acute myocardial ischemia

ObjectiveTo investigate the effect of electroacupuncture (EA) of acupoints Neiguan (PC 6) and Tianquan (PC 2) on the skin temperature, blood perfusion, and alpha1- and beta2-adrenoreceptor (α1-AR and β2-AR) protein and mRNA level in rats with acute myocardial ischemia. MethodsThirty male adult Wistar rats [(230 ± 20) g] were randomly assigned into five groups (n = 6 each): a control group, sham operation group, model group, model group treated with EA at low frequency (L-EA, 2 Hz, 1 mA) and model group treated with EA at high frequency (H-EA, 100 Hz, 1 mA). The rat model was prepared by ligating the left anterior descending coronary artery. Electroacupuncture was performed at the left Neiguan (PC 6) for 20min daily for 3 d. After the 3rd time of the treatment, measurements of skin blood perfusion and temperature in Neiguan (PC 6) and Tianquan (PC 2) in all groups were made by laser speckle contrast imaging and infrared thermal image instrument. Real-time PCR and ELISA were used to measure mRNA level and protein level of α1-AR and β2-AR in the skin tissues of Neiguan (PC 6) and Tianquan (PC 2), respectively. Serum levels of cTnT and electrocardiogram were used to identify the state of myocardium. ResultsIn the group receiving electroacupunture at Neiguan (PC 6), compared with control group, the skin temperature, blood perfusion, and β2-AR mRNA and protein level of model group significantly decreased (P < 0.05), α1-AR mRNA and protein level of model group significantly increased (P < 0.05); compared with model group, the skin temperature, blood perfusion, and β2-AR mRNA and protein level of L-EA significantly increased (P < 0.05), α1-AR mRNA and protein level of L-EA and H-EA significantly decreased (P < 0.05). The same trend has taken place in the former four groups of the Tianquan (PC 2). ConclusionLow-or high-frequency electroacupuncture can improve the skin temperature and blood perfusion which may be induced by decreasing the level of α1-AR and increasing the level of β2-AR of the Neiguan (PC 6) and Tianquan (PC 2) in the rat with acute myocaidial ischemia.

Responses of PKCε to cardiac overloads on myocardial sympathetic innervation and NET expression.

Protein kinase C (PKC) is a key mediator of many diverse physiological and pathological responses. PKC activation play an important regulatory role of cardiac function. The present study was performed to investigate whether there were differential activations of the PKCε and how the activation coupled with norepinephrine transporter (NET) surface expression, sympathetic innervation pattern and extracellular matrix remodeling in different cardiac hemodynamic overloads induced by abdominal aortic constriction or aortocaval fistula. At 8weeks after the operations, heart failure were induced, accompanied with myocardial hypertrophy, which was more pronounced in pressure overload (POL) than that of volume overload (VOL) rats, left ventricular dysfunction and increased plasma norepinephrine (NE). In POL rats there was an increase in myocardial collagen deposition, in contrast, the amount decreased in VOL as compared with the sham rats. POL remarkably upregulated PKCε membrane-cytosol ratio and downregulated NET membrane fraction, whereas, in VOL induced opposite changes. Accompanied with the PKCε activation, nerve sprouting, evidenced by myocardial GAP43 protein increased, and different nerve phenotypes were found, in POL tyrosine hydroxylase (TH) positive nerve density increased with NET and choline acetyltransferase (ChAT) immunoreactivity density decreased, in contrast, in VOL NET and ChAT increased, TH did not change. The overloads did not induce alteration of NET mRNA expression, but resulted in different myocardial β1-AR mRNA expression, in POL β1-AR mRNAwas significantly downregulated, while in VOL rats unaltered. Conclusion, the present results suggested that the different cardiac hemodynamic overload could differentially activate a common signaling, PKCε intermediate and thereby generate biological diversity.

Estrogen replacement attenuates stress-induced pressor responses through vasorelaxation via β2-adrenoceptors in peripheral arteries of ovariectomized rats.

We examined whether chronic estrogen replacement has an inhibitory effect on stress-induced pressor responses via activation of β2-adrenoceptor (AR) in peripheral arteries of ovariectomized rats. Female Wistar rats aged 9 wk were ovariectomized. After 4 wk, pellets containing either 17β-estradiol (E2) or placebo (Pla) were subcutaneously implanted into the rats. After 4 wk of treatment, rats underwent cage-switch stress, and, in a separate experiment, a subset received an infusion of isoproterenol (ISO) with or without pretreatment with the β1-AR blocker atenolol or the β2-AR blocker butoxamine. In addition, the isolated mesenteric artery was used to assess the concentration-related relaxing responses to ISO and the β1- or β2-AR mRNA level. The cage-switch stress-induced pressor response was significantly attenuated in the E2-treated group compared with the Pla-treated group. Pretreatment with atenolol reduced blood pressure responses in both groups. However, butoxamine enhanced the pressor response only in the E2-treated group, resulting in no difference between the two groups. In addition, the intravenous ISO-induced depressor response was significantly enhanced in the E2-treated group compared with the Pla-treated group. Furthermore, the difference in the depressor response was abolished by pretreatment with butoxamine but not by atenolol. In the isolated mesenteric artery, butoxamine caused a rightward shift in ISO-induced concentration-related relaxation in the E2-treated group. The β2-AR mRNA level in the mesenteric artery was higher in the E2-treated group than in the Pla-treated group. These results suggest that estrogen replacement attenuated the stress-induced pressor response probably by suppressing vasoconstriction via activation of β2-ARs in peripheral arteries of ovariectomized rats. NEW & NOTEWORTHY In this study, we show, for the first time, that estrogen replacement has an inhibitory effect on the psychological stress-induced pressor response through vasorelaxation via β2-adrenoceptors, probably due to overexpression of β2-adrenoceptor mRNA, in peripheral arteries of ovariectomized rats.

Protective effect of esmolol on peritoneal macrophages in rats with sepsis

Objective To investigate the protective effects of esmolol on peritoneal macrophages in rats with sepsis. Methods Rat peritoneal macrophages were deprived of serum and divided into four groups, namely normal group, LPS stimulated group, esmolol group and LPS plus esmolol group. Quantitative real-time PCR and Western blot were performed to assay the expressions and protein levels of TNF-α, β1-AR and p65. The activation of NF-κB was determined by electrophoretic mobility shift assay. Results The expression of TNF-α mRNA and protein level was significantly increased in LPS-treated group than those in normal group (P<0.01), and those in LPS plus esmolol group was significantly lower than those in LPS stimulated group (P<0.01). The expression of β1-AR mRNA and protein level were significantly higher in LPS-treated group than those in normal group (P<0.01), The expression of β1-AR mRNA and protein level in the esmolol group was lower than those in normal group (P<0.01), and those in LPS plus esmolol group was lower than those in LPS stimulated group (P<0.01). Besides, compared with the normal group, the expression of p65 mRNA in the LPS stimulated group was noticeably higher (P<0.01). The expression of P65 mRNA in LPS plus esmolol group was significantly lower than that in LPS-treated group (P<0.01). The NF-κB/DNA binding force in the LPS stimulated group was significantly higher than that in normal group (P<0.01). Compared with the LPS stimulated group, the NF-κB/DNA binding force in the LPS plus esmolol group was significantly lower (P<0.01). Conclusions The expression of TNF-α mRNA induced by LPS was decreased by esmolol in rat peritoneal macrophages which might be achieved via inhibiting NF-κB pathway by the modulation role of β1 adrenergic receptor. Key words: Esmolol; NF-κB; β1-AR; TNF-α

Zilpaterol Hydrochloride affects Cellular Muscle Metabolism and Lipid Components of 10 Different Muscles in Feedlot Heifers

This study determined if zilpaterol hydrochloride (ZH) altered muscle metabolism and lipid components of 10 muscles. Crossbred heifers were either supplemented with ZH (n = 9) or not (Control; n = 10). Muscle tissue was collected (adductor femoris, biceps femoris, gluteus medius, infraspinatus, latissimus dorsi, longissimus dorsi, pectoralis profundi, semitendinosus, subscapularis, trapezius) immediately following carcass splitting. The mRNA abundance of AMPkɑ, IGF-I, MHC-I, IIA and IIX, β1-adrenergic receptor (βAR) and β2AR was determined, as well as, cross-sectional area and proportion of myosin isoforms, β1AR, β2AR, β3AR, nuclei, and satellite cell density. Furthermore, neutral (NL) and polar lipid (PL) fatty acids (FA) were quantified (mg/g). Zilpaterol hydrochloride decreased MHC-IIA mRNA (P = 0.007). In addition, ZH decreased total nuclei and β1AR and increased MHC-IIX cross-sectional area (P ≤ 0.021). Quantity of NL FA were not affected by ZH (P ≥ 0.173). However, among PL FA the ratio of PUFA:SFA was greater with ZH (P = 0.048). Muscle type impacted mRNA concentration of AMPkɑ, IGF-I, MHC-I, IIA, IIX, and β1AR mRNA concentration (P ≤ 0.037). Furthermore, the fiber type proportion, fiber cross-sectional area, and the densities of nuclei, β1AR, β2AR, β3AR, and satellite cells were influenced by muscle type (P ≤ 0.030). Total NL FA were affected by muscle (P ≤ 0.046). Meanwhile, total PL FA did not differ due to muscle (P = 0.242). However, prominent PL FA,18:0, 18:1 trans, and 18:2 n-6 were each greater (P &lt; 0.05) among the oxidative subscapularis compared with glycolytic semitendinosus and adductor femoris. Overall, these data reveal that ZH impacts muscle metabolism and myogenic activity that establishes protein deposition. Meanwhile, ZH did not alter triglyceride content (NL), but cell membrane saturation (PL) was influenced, in accordance with alterations to muscle fiber type. Muscle also influenced muscle fiber type and lipid components. Therefore, muscle biology is greatly influenced by muscle but also through dietary inclusion of ZH.