Coronary Artery In Mice Research Articles

Y4 RNA fragments from cardiosphere-derived cells ameliorate diabetic myocardial ischemia‒reperfusion injury by inhibiting protein kinase C β-mediated macrophage polarization

The specific pathophysiological pathways through which diabetes exacerbates myocardial ischemia/reperfusion (I/R) injury remain unclear; however, dysregulation of immune and inflammatory cells, potentially driven by abnormalities in their number and function due to diabetes, may play a significant role. In the present investigation, we simulated myocardial I/R injury by inducing ischemia through ligation of the left anterior descending coronary artery in mice for 40 min, followed by reperfusion for 24 h. Previous studies have indicated that protein kinase Cβ (PKCβ) is upregulated under hyperglycemic conditions and is implicated in the development of various diabetic complications. The Y4 RNA fragment is identified as the predominant small RNA component present in the extracellular vesicles of cardio sphere-derived cells (CDCs), exhibiting notable anti-inflammatory properties in the contexts of myocardial infarction and cardiac hypertrophy. Our investigation revealed that the administration of Y4 RNA into the ventricular cavity of db/db mice following myocardial I/R injury markedly enhanced cardiac function. Furthermore, Y4 RNA was observed to facilitate M2 macrophage polarization and interleukin-10 secretion through the suppression of PKCβ activation. The mechanism by which Y4 RNA affects PKCβ by regulating macrophage activation within the inflammatory environment involves the inhibition of ERK1/2 phosphorylation In our study, the role of PKCβ in regulating macrophage polarization during myocardial I/R injury was investigated through the use of PKCβ knockout mice. Our findings indicate that PKCβ plays a crucial role in modulating the inflammatory response associated with macrophage activation in db/db mice experiencing myocardial I/R, with a notable exacerbation of this response observed upon significant upregulation of PKCβ expression. In vitro studies further elucidated the protective mechanism by which Y4 RNA modulates the PKCβ/ERK1/2 signaling pathway to induce M2 macrophage activation. Overall, our findings suggest that Y4 RNA plays an anti-inflammatory role in diabetic I/R injury, suggesting a novel therapeutic approach for managing myocardial I/R injury in diabetic individuals.

Rosuvastatin Enhances Lymphangiogenesis after Myocardial Infarction by Regulating the miRNAs/Vascular Endothelial Growth Factor Receptor 3 (miRNAs/VEGFR3) Pathway.

Several clinical studies have suggested that the early administration of statins could reduce the risk of in-hospital mortality in acute myocardial infarction (AMI) patients. Recently, some studies have identified that stimulating lymphangiogenesis after AMI could improve cardiac function by reducing myocardial edema and inflammation. This study aimed to identify the effect of rosuvastatin on postinfarct lymphangiogenesis and to identify the underlying mechanism of this effect. Myocardial infarction (MI) was induced by ligation of the left anterior descending coronary artery in mice orally administered rosuvastatin for 7 days. The changes in cardiac function, pathology, and lymphangiogenesis following MI were measured by echocardiography and immunostaining. EdU, Matrigel tube formation, and scratch wound assays were used to evaluate the effect of rosuvastatin on the proliferation, tube formation, and migration of the lymphatic endothelial cell line SVEC4-10. The expression of miR-107-3p, miR-491-5p, and VEGFR3 was measured by polymerase chain reaction (PCR) and Western blotting. A gain-of-function study was performed using miR-107-3p and miR-491-5p mimics. The rosuvastatin-treated mice had a significantly improved ejection fraction and increased lymphatic plexus density 7 days after MI. Rosuvastatin also reduced myocardial edema and inflammatory response after MI. We used a VEGFR3 inhibitor to partially reverse these effects. Rosuvastatin promoted the proliferation, migration, and tube formation of SVEC4-10 cells. PCR and Western blot analyses revealed that rosuvastatin intervention downregulated miR-107-3p and miR-491-5p and promoted VEGFR3 expression. The gain-of-function study showed that miR-107-3p and miR-491-5p could inhibit the proliferation, migration, and tube formation of SVEC4-10 cells. Rosuvastatin could improve heart function by promoting lymphangiogenesis after MI by regulating the miRNAs/VEGFR3 pathway.

The alkaloids of Corydalis hendersonii Hemsl. contribute to the cardioprotective effect against ischemic injury in mice by attenuating cardiomyocyte apoptosis via p38 MAPK signaling pathway

BackgroundThere is a characteristic Tibetan folk medicine in China named Corydalis hendersonii Hemsl. (CH) has been used for treatment of cardiovascular related diseases, called “plethora” in Tibetan medicine. Previous studies demonstrated that ethanol extract of CH showed anti-acute myocardial infarction (AMI) effect through inhibiting fibrosis and inflammation. Rich alkaloids fraction (RAF) is isolated from CH, but whether RAF possessing an equivalent effect with the CH ethanol extract and by which mechanism it protects against AMI has not yet reported. The paper aimed to study the potential role of RAF on myocardial injured mice and its underlying mechanism.Materials and methodsLiquid chromatography mass spectrometry-ion trap-time of flight (LCMS-IT-TOF) was used to analyze the chemical profile and isolate pure compounds. The ligation of left anterior descending (LAD) of coronary artery in mice was used to evaluate the in vivo anti-AMI effect, by dividing into eight groups: Sham, Model, Fosinopril (10 mg/kg, i.g.), total extract (TE, 400 mg/kg, i.g.), poor alkaloids fraction (PAF, 300 mg/kg, i.g.), and RAF (25, 50, and 100 mg/kg, respectively, i.g.) groups. Echocardiography was used to evaluate mice heart function through the index of left ventricular end-systolic diameter (LVEDs), left ventricular end-diastolic diameter (LVEDd), fractional shortening (FS) and ejection fraction (EF). We detected the lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) in the serum and the plasma level of angiotensin II (AngII). The apoptosis of mice myocardial tissue was verified by TUNEL assay. The expression of p38 mitogen-activated protein kinases (p38 MAPK), Bcl-2 and Bcl-2-associated X protein (Bax) were detected through immunofluorescence staining, qRT-PCR and western blot in mice heart tissue and H9c2 cells.ResultsEchocardiography data indicated that the values of LVEDd and LVEDs were reduced and the values of FS and EF were improved by TE and RAF significantly. RAF also decreased the levels of LDH, CK-MB and AngII and significantly inhibited inflammatory cells in the marginal zone of myocardial infarction. The TUNEL assay results showed that RAF significantly attenuated cell apoptosis. Immunofluorescence and qRT-PCR assay showed that RAF inhibited p38 MAPK, Bax, and Bcl-2 proteins in mice myocardium. Western blot results validated that the expressions of key proteins were inhibited by RAF. Also, the apoptotic cells and apoptosis-related proteins were dramatically reduced by RAF in vivo and in vitro. Besides, RAF and PAF were analyzed by LCMS-IT-TOF to identify the main compounds and to demonstrate the difference between them. The results showed that a total of 14 alkaloids were identified, which indicated that the isoquinoline alkaloids were the main ingredients in RAF may contributing to the cardioprotective effect in mice.ConclusionsRAF improves cardiac function by inhibiting apoptosis via p38 MAPK signaling pathway, and RAF contributes to the effect against myocardial ischemic injury of TE in mice, which provides a substantial reference for the clinical application against ischemia heart disease and quality control of CH.Graphical

Dexmedetomidine reduces myocardial ischemia-reperfusion injury in young mice through MIF/AMPK/GLUT4 axis

BackgroundReperfusion of ischemic tissue has adverse impact on the myocardium. Dexmedetomidine (Dex) is a α2-adrenergic receptor (α2-AR) agonist with sedative and analgesic effects. Macrophage migration inhibition factor (MIF) is a pressure-regulating cytokine and is responsible for inflammatory and immune diseases. This study aims to reveal the consequences of Dex on myocardial ischemia-reperfusion injury (IRI) in young mice.MethodsFifty mice were raised and examined. At the end of the experiment, all mice were euthanized. The anterior descending department of the left coronary artery in mice was under ischemia for 60 min, then the ligation line was released and reperfused for 120 min to establish the IRI model. Mice were randomly divided into Sham, control, treatment using 4,5-dihydro-3-(4-hydroxyphenyl)-5-isoxazoleacetic acid (ISO-1), Dex treatment, and Dex combined ISO-1 treatment groups. Interleukin (IL)-6, IL-10 and tumor necrosis factor (TNF-α) were determined by enzyme-linked immunosorbent assay (ELISA). Reactive oxygen species (ROS) and ATP levels were recorded. The expressions of MIF, P-adenosine monophosphate-activated kinase α (AMPKα), glucose transporter (GLUT)4, Bax and Bcl-2 were detected by Western Blot (WB). Hematoxylin and Eosin (H&E) staining was used to study cell morphology. Apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay. Echocardiography was carried out at the end of reperfusion, and the infarct size was calculated by Electron microscopy.ResultsI/R + Dex group showed significantly increased IL-6 and TNF-α levels and reduced myocardial cell necrosis and apoptosis. H&E staining showed alleviated myocardial disorder, myocardial cell swelling, myocardial fiber fracture, and inflammatory cell infiltration in I/R + Dex group. Myocardial cell necrosis and apoptosis were significantly reduced in I/R + Dex group. ATP level in myocardial tissue of mice in I/R group was substantially decreased, while that in Dex group was increased. WB results showed that MIF, P-AMPK α, GLUT4 and Bcl-2 levels were increased and Bax levels were decreased in I/R + Dex group.ConclusionDex may exert myocardial protection in young mice through MIF/AMPK/GLUT4 axis.

CTRP12 alleviates cardiomyocyte ischemia‑reperfusion injury via regulation of KLF15.

Myocardial ischemia-reperfusion (I/R) serves a crucial role in myocardial infarction. C1q/TNF-related protein 12 (CTRP12) is a secretory protein involved in metabolism. It has been reported that CTRP12 participates in the regulation of numerous cardiovascular diseases. However, its role in myocardial I/R injury remains unclear. In the present study, the left anterior descending coronary artery in mice was ligated to establish a mouse I/R model. A myocardial hypoxia-reoxygenation (H/R) cell model was also established. Cardiomyocyte injury was evaluated using hematoxylin and eosin staining, Cell Counting Kit-8 and a lactate dehydrogenase (LDH) kit. The expression levels of CTRP12 and Krueppel-like factor 15 (KLF15) in murine myocardial tissues and H9c2 cells were determined using reverse transcription-quantitative PCR and western blotting, as KLF15 was previously reported to protect against I/R-induced cardiomyocyte damage. Furthermore, inflammatory factors TNF-α, IL-1β and IL-6 were analyzed using ELISA while apoptosis was assessed using TUNEL assays and western blotting. Moreover, the activity of the CTRP12 promoter was determined using a dual-luciferase reporter assay. The results demonstrated that I/R surgery markedly exacerbated myocardial tissue damage, whereas H/R treatment significantly reduced cell viability and significantly increased LDH activity as well as the release of inflammatory factors and apoptosis. I/R and H/R induction significantly reduced the expression levels of CTRP12 and KLF15. CTRP12 overexpression significantly alleviated H/R-induced cell injury and significantly inhibited inflammation and apoptosis. Further analysis demonstrated that KLF15 could significantly promote the activity of the CTRP12 promoter. However, following CTRP12 knockdown, KLF15 overexpression exacerbated cell injury, inflammation and apoptosis. In conclusion, the present study demonstrated that CTRP12 may mitigate inflammation and apoptosis in H/R-induced cardiomyocytes, possibly via the regulation of KLF15, which provided a theoretical basis for the potential treatment of I/R-induced myocardial infarction.

Role of tissue resident VSIG4+ macrophages in cardiac remodeling after acute myocardial infarction in mice

Adaptive and innate immune cells have been shown to govern cardiac injury and remodeling following acute myocardial infarction (AMI). A detailed understanding of the interactions between inflammation and the structural and functional alterations of the infarcted heart could pave the way for the development of innovative therapeutic strategies. In the infarcted heart, macrophages constitute a heterogeneous population including monocyte-derived macrophages (MDM) emerging from splenic and medullary reservoirs and tissue-resident macrophages (TRM). MDM rather exert pro-inflammatory deleterious effects whereas TRM are more likely to perform anti-inflammatory protective functions in the infarcted heart. Using single cell RNA sequencing, we revealed that VSIG4 (B7 family-related protein V-set and Ig domain-containing 4) characterized a specific subset of TRM in the cardiac tissue. In the present work, we therefore aim to evaluate the role of VSIG4 + TRM after AMI. Acute MI was induced by permanent ligation of the left anterior descending coronary artery in mice. Cardiac function and remodeling were assessed using echocardiography and immunohistochemistry. Vsig4 depletion impaired cardiac function and promoted adverse cardiac remodeling after experimental AMI in mice. WT mice, transplanted with bone marrow (BM) derived cells isolated from Vsig4-deficient mice (Vsig4-/-) or WT mice, exhibited comparable cardiac function and remodeling after AMI. Conversely, Vsig4 deficient mice transplanted with WT BM-derived cells, displayed worse cardiac function and adverse ventricular remodeling after AMI, when compared to WT mice re-supplemented with WT BM-derived cells. Moreover, FACS-sorted Vsig4+ macrophages injected into infarcted hearts 2 weeks post-AMI, improved cardiac function in both WT and Vsig4-/- mice, supporting the beneficial role of VSIG4+ TRM in the context of AMI. We have also demonstrated that IL4 and IL13, through IL4 receptor alpha and IL13 receptor alpha1, fostered Vsig4 expression in peritoneal macrophages (PEM), a prototypical example of TRM. Meanwhile, IL4 and IL13 elicited the release of CCL3, CCL4 and CCL5, chemokine ligands of CCR5, in cultured WT PEM but not in Vsig4-deficient PEM. Moreover, induction of AMI in Vsig4-/- mice hampered the number of CCR5+ regulatory T lymphocytes in infarcted hearts. VSIG4+ TRM are key regulators of cardiac healing after AMI likely through their ability to foster the infiltration of anti-inflammatory regulatory T cells in the infarcted heart.

Influence of Hypoxia Inducible Factor-1α of Endothelial Progenitor Cells on Left Ventricular Function in Experimental Myocardial Infarction

Based on the promotion of myocardial activity via endothelial progenitor cells (EPCs) subsequent to acute myocardial infarction (AMI), our research was designed to explore the influence of excessive HIF-1α expression in expanded EPCs (eEPCs) on promotion of the activity of left ventricle subsequent to MI. Isolation of EPCs from cord blood was performed before transduction with the help of retroviral vector with or without HIF-1α expression. Transplantation was performed subsequent to ligation of the left anterior descending coronary artery in mice. Ejection fraction (EF) of left ventricle was promoted via transplantation after 2 weeks. Excessive HIF-1α expression enhanced EF of left ventricle and decreased the extent of MI. It was revealed via functional studies that excessive HIF-1α expression enhanced proliferation of EPCs triggered by low oxygen concentration and suppressed cell death in the region of infarction. Moreover, markers of endothelium CD31, VEGF, and eNOS were upregulated. Transplantation of eEPCs with excessive HIF-1α expression in AMI can promote myocardial activities by increasing differentiation, generation of vessels, proliferation of eEPCs, and suppressing cell death. The above findings propose that regulation of EPCs via HIF-1α enhances the activity as well as mobilization of EPCs, indicating that reinforcement of expression of HIF-1α is beneficial for coronary heart disease.

Fibroblast growth factor 12 attenuated cardiac remodeling via suppressing oxidative stress

Fibroblast growth factors (FGFs) mediate key cardiac functions from development to homeostasis and disease. The current research was to explore the effects of FGF12 in the fibrosis of cardiac function and heart failure, and whether FGF12 alleviated cardiac fibrosis via inhibition of oxidative stress. Ligation of left coronary artery in mice induced heart failure and myocardial infarction (MI). Angiotensin II (Ang II) was administered to cardiac fibroblasts (CFs). FGF12 upregulation or FGF12 transgenic (Tg) mice could improve cardiac dysfunction of MI mice, and attenuated cardiac fibrosis of heart failure induced by MI in mice. FGF12 overexpression suppressed the increases of collagen I, collagen III and fibronectin which was induced by Ang II in CFs. The oxidative stress was enhanced in the heart of MI mice and CFs treated with Ang II, and these enhances were attenuated via FGF12 overexpression. Superoxide dismutase (SOD) overexpression inhibited the enhancements of collagen I, collagen III and fibronectin in the heart of MI mice, and in the CFs treated with Ang II. Overexpression of nicotinamide adenine dinucleotide phosphate oxidases (Nox1) reversed the attenuating influences of FGF12 on the enhancements of collagen I, collagen III and fibronectin in the CFs induced by Ang II. These outcomes showed that FGF12 upregulation can improve cardiac dysfunction and heart fibrosis of heart failure. FGF12 attenuates oxidative stress to suppress the cardiac fibrosis.

The Therapeutic Effect of miRNA-19a/19b on Heart Failure in Mice and the Mechanism of Myocardial Regeneration and Repair.

Heart failure is a common cardiovascular disease. The elderly have a high risk of illness. Many patients die of the disease every year. It has become a killer for all humans. Cardiovascular disease often triggers myocardial ischemia and myocardial cell apoptosis, resulting in myocardial damage and even heart failure in patients with cardiovascular disease. Related studies have shown that miRNA-19a/19b plays an important role in myocardial damage and heart failure character. The purpose of this article is to further explore the therapeutic effect of miRNA-19a/19b on heart failure and the regulation mechanism of myocardial regeneration and repair. In this paper, 40 male mice were used as experimental objects to perform ligation surgery on the coronary arteries of mice. To establish an animal model of myocardial infarction and heart failure, then inject miRNA-19a/19b reagent through the tail vein of the mouse, observe the changes of the myocardial cell and the area of the myocardial infarction by immunofluorescence PCR technology, and detect the myocardial tissue in the mouse. Specific changes in the expression of miRNA-19a/19b. The results of the study showed that miRNA-19a/19b can improve the cardiac function of mice. The cardiac function index of mice increased from the original (263.13±5.26) to (385.48±6.92), the area of myocardial infarction decreased by 27.5%, and the proliferation rate of myocardial cells increased by 18.6%, the rate of myocardial regeneration and repair increased by 20.4%. Therefore, it can be seen that miRNA-19a/19b has a significant effect on the treatment of heart failure in mice, can effectively improve heart function, and can promote the regeneration and repair of damaged myocardium, proving the feasibility of miRNA-19a/19b for cardiovascular disease.

Generation and Characterization of Right Ventricular Myocardial Infarction Induced by Permanent Ligation of the Right Coronary Artery in Mice

Right ventricular infarction (RVI) is a common presentation in clinical practice. Severe RVI can lead to fatal hemodynamic dysfunction and arrhythmia. In contrast to the extensively used mouse myocardial infarction (MI) model generated by left coronary artery ligation, the RVI mouse model is rarely employed due to the difficulty associated with model generation. Research on the mechanisms and treatment of RVI-induced RV remodeling and dysfunction requires animal models to mimic the pathophysiology of RVI in patients. This study introduces a feasible procedure for RVI model generation in C57BL/6J mice. Further, this model was characterized based on the following: infarct size evaluation at 24 h after MI, assessment of cardiac remodeling and function with echocardiography, RV hemodynamics assessment, and histology of the infarct zone at 4 weeks after RVI. In addition, a coronary vasculature cast was performed to observe the coronary arterial arrangement in RV. This mouse model of RVI would facilitate the research on mechanisms of right heart failure and seek new therapeutic targets of RV remodeling.

Generation and Characterization of Right Ventricular Myocardial Infarction Induced by Permanent Ligation of the Right Coronary Artery in Mice

Generation and Characterization of Right Ventricular Myocardial Infarction Induced by Permanent Ligation of the Right Coronary Artery in Mice

Abstract P184: Novel Model Of Ionizing Radiation-induced Mouse Coronary Arteriosclerosis, Which Was Attenuated By Precise Time Treatment Of Poly (adp-ribose) Polymerase (parp) Inhibitor

Background: It is well known that radiation therapy (RT) induces coronary artery disease (CAD). However, since we don’t have an adequate mouse model to evaluate CAD after RT, it has been difficult to perform pre-clinical study to detect the effective treatment. The inhibition of PARPs can prevent apoptosis and inflammation. Although PARP inhibitors have already been used as anti-tumor agents, their long-term use in CAD is not recommended because the inhibition of DNA damage response can cause DNA instability and eventually cancer. In this study, we restricted the use of PARP inhibitors to the time of IR exposure only and detected IR-induced atherosclerosis (AS). Methods and Results: Marino et al. reported that thoracic aortic constriction (TAC) can exacerbate coronary artery stenosis and myocardial infarction in ApoE -/- mice. We tested this model in LDLR -/- mice that were fed a high fat (HFD) with or without IR (3, 5, and 10 Gy). We observed cardiac hypertrophy and dose-dependent reduction of fractional shortening (FS) after 4-5 weeks of TAC (FS%, 33.74 ± 7.35 (Non-IR, n=9), 26.59 ± 9.19 (3Gy, n=6), 28.49 ± 2.49 (5Gy, n=5), 22.62 ± 6.16 (10Gy, n=6), mean ± SD, p <0.05). Most importantly, we observed localized cardiac dysfunction and infarct only in mice exposed to 10 Gy (n = 2 out of 6), detected by transmural strain analysis with echocardiography. Next, the whole heart was sectioned, with sets of 11 consecutive sections of 5 μm collected every 450 μm interval. We also found the diffuse increase of vascular wall thickness at left anterior descending coronary artery in mice exposed to IR. We fed a HFD on LDLR -/- mice with Olaparib (10 mg/kg/day) or vehicle one day before & after, and the day of IR (5 Gy twice, total 6 days only), then performed TAC. The reduction of FS induced by IR (10 Gy) was significantly improved by the precise time treatment of Olaparib against IR. Conclusion: These data suggested the usefulness of TAC-induced coronary AS mouse model to develop medical countermeasures against RT-induced CAD as pre-clinical study.

Elabela gene therapy promotes angiogenesis after myocardial infarction.

This study was aimed at investigating whether Elabela (ELA) gene therapy can promote angiogenesis in the treatment of myocardial infarction (MI). The fusion expression plasmid pAAV‐3 × Flag/ELA‐32 was successfully constructed using molecular cloning technique. The model of acute MI was established by ligating the left anterior descending coronary artery in mice. Adeno‐associated virus serotype 9 (AAV9) was injected into the surrounding myocardium and tail vein immediately after the model was established. AAV was injected again from the tail vein one week later. Compared with the MI+PBS (control) group, the serum N‐terminal pro‐brain natriuretic peptide (NT‐proBNP) concentration, and the values of left ventricular end‐diastolic diameter (LVDd) and left ventricular end‐systolic diameter (LVDs) of the MI+AAV‐ELA (gene therapy) group were significantly decreased, while the value of left ventricular ejection fraction was significantly increased at 2 and 4 weeks after operation. Compared with the control group, the expression of CD105 and vWF and the percentage of CD31‐ and Ki67–co‐positive cells were significantly increased in the gene therapy group. Moreover, the expressions of apelin peptide jejunum (APJ) receptor, vascular endothelial growth factor (VEGF), VEGFR2, Jagged1 and Notch3 in the heart tissue around the infarction were up‐regulated in mice with gene therapy. The results suggest that ELA activates VEFG/VEGFR2 and Jagged1/Notch3 pathways through APJ to promote angiogenesis after myocardial infarction. ELA gene therapy may be used in the treatment of ischaemic cardiomyopathy in future.

Dexmedetomidine improves cardiac function and protects against maladaptive remodeling following myocardial infarction.

Dexmedetomidine (DEX), a highly specific and selective α2 adrenergic receptor agonist, has been demonstrated to possess potential cardioprotective effects. However, the mechanisms underlying this process remain to be fully illuminated. In the present study, a myocardial infarction (MI) animal model was generated by permanently ligating the left anterior descending coronary artery in mice. Cardiac function and collagen content were evaluated by transthoracic echocardiography and picrosirius red staining, respectively. Apoptosis was determined by the relative expression levels of Bax and Bcl-2 and the myocardial caspase-3 activity. Additionally, nicotinamide adenine dinucleotide phosphate oxidase (NOX)-derived oxidative stress was evaluated by the relative expression of Nox2 and Nox4, along with the myocardial contents of malondialdehyde (MDA) and superoxide dismutase (SOD) activity. It was demonstrated that intraperitoneal DEX treatment (20 µg/kg/day) improved the systolic function of the left ventricle, and decreased the fibrotic changes in post-myocardial infarction mice, which was paralleled by a decrease in the levels of apoptosis. Subsequent experiments indicated that the restoration of redox signaling was achieved by DEX administration, and the over-activation of NOXs, including Nox2 and Nox4, was markedly inhibited. In conclusion, this present study suggested that DEX was cardioprotective and limited the excess production of NOX-derived ROS in ischemic heart disease, implying that DEX is a promising novel drug, especially for patients who have suffered MI.

P6292Role of CXCL12gamma isoform and its interactions with heparan-sulfates in post-ischemic cardiac remodeling

Abstract Introduction Myocardial infarction (MI) is a severe ischemic disease precipitating long-term adverse remodeling and heart failure. The chemokine CXCL12/SDF-1 is essential for cardiovascular system development and plays a prominent role in physio-pathological processes such as inflammation, angiogenesis and tissue fibrosis. In addition to the binding to its cognate receptors CXCR4 and CXCR7, CXCL12 interacts with heparan-sulfates (HS) which coordinate its biological activity. We have previously highlighted the essential role of CXCL12/HS interactions in vascular growth and remodeling in the setting of critical limb ischemia. In addition, studies in experimental model of MI revealed a protective role for the CXCL12α isoform, through the regulation of cardiomyocyte survival and recruitment of inflammatory cells. However, in mice, three CXCL12 isoforms (α, β and γ) have been identified and, among them, the CXCL12γ isoform shows an unchallenged ability to cooperate with HS, suggesting a putative pivotal role in tissue repair. Objectives The aim of the study is to analyze the role of CXCL12γ isoform and the importance of CXCL12/HS interactions in post-ischemic cardiac remodeling in an acute model of MI. Methods MI was induced by permanent ligation of the left ascending coronary artery in mice carrying a Cxcl12 gene mutation that precludes interactions with HS (Cxcl12Gagtm) and in Cxcl12γ knock-in animals (Cxcl12γ-KI) harboring CXCL12γ deficiency. Alternatively, the impact of CXCL12γ overexpression and the importance of its interactions with HS was also evaluated in wild-type (WT) mice receiving transcutaneous echo-guided injections of adenovirus encoding WT Cxcl12γ or HS-binding-disabled Cxcl12γ in cardiac tissue. Cardiac function and remodeling have been assessed through echocardiography analysis, evaluation of infarct size, interstitial fibrosis, vascular growth (capillary and arteriole densities) and inflammatory cell infiltration into the cardiac tissue. Results After MI, Cxcl12Gagtm and Cxcl12γ-KI animals exhibit reduction in cardiac function and adverse left ventricular remodeling when compared to their respective WT littermates. Interestingly, overexpression of CXCL12γ in WT mice cardiac restored cardiac function by reducing the size of the infarcted area, interstitial fibrosis and promoting vascular growth. In sharp contrast, HS–binding disabled CXCL12gamma mutants failed to improve cardiac function and to abrogate adverse left ventricular remodeling. Conclusion We show that CXCL12γ isoform plays an important role in the regulation of post-ischemic cardiac function and remodeling and that its interactions with HS are essential for adequate cardiac repair in the setting of acute MI.

Evaluation of coronary artery hemodynamics and its relationship with atherosclerosis in west diet fed LDL-R knockout mice by ultrasound bio-microscopy

Objective To evaluate the relationship between atherosclerosis and hemodynamic of coronary artery in mice detecting by ultrasound bio-microscopy flow imaging. Methods Double 14 20-week-old LDL-R-/- and C57BL/6 male mice were selected, and randomly divided into two groups in each genotype according to weight. Each two groups were fed to 28 weeks or 36 weeks age respectively with west diet. Coronary artery hemodynamics in these mice were assessed in vivo by Vevo®2100 ultrasound imaging system, then the intima-media thickness(IMT) of aorta in histopathology were analyzed. The differences of coronary artery hemodynamic parameters such as maximum velocity (Vmax), mean velocity (Vmean) and velocity time integral (VTI) were compared between mice of different genotypes of the same week and mice of different weeks of the same genotype. And the relationship between coronary artery hemodynamic in ultrasound and aortic IMT in histopathology were analyzed. Results ①All coronary hemodynamic parameters in LDL-R-/- mice were significantly lower than those of wild-type mice except the Vmax between two 28-week-old genotypes group at the same weeks of age of different genotypes (all P 0.05). ②The histopathological measurements of aortic IMT in LDL-R-/- mice were significantly higher than those of wild type mice(all P<0.05), and those of 36-week-old mice were significantly higher than those of 28-week-old mice(all P<0.05). ③All coronary hemodynamic parameters such as Vmax, Vmean and VTI were negatively correlated with pathological measurements of aortic IMT (r=-0.532, -0.423, -0.524; all P<0.05). Conclusions The parameters of coronary artery hemodynamics obtained by ultrasound bio-microscopy are well correlated with the pathological results of atherosclerosis. Ultrasound bio-microscopic flow imaging can be used as a new method to evaluate the degree of atherosclerosis in mice by detecting the hemodynamic parameters of coronary artery. Key words: Ultrasound bio-microscopy; Mice; Coronary artery; Hemodynamics; Atherosclerosis

Dectin-2-induced CCL2 production in tissue-resident macrophages ignites cardiac arteritis.

Environmental triggers, including those from pathogens, are thought to play an important role in triggering autoimmune diseases, such as vasculitis, in genetically susceptible individuals. The mechanism by which activation of the innate immune system contributes to vessel-specific autoimmunity in vasculitis is not known. Systemic administration of Candida albicans water-soluble extract (CAWS) induces vasculitis in the aortic root and coronary arteries of mice that mimics human Kawasaki disease. We found that Dectin-2 signaling in macrophages resident in the aortic root of the heart induced early CCL2 production and the initial recruitment of CCR2+ inflammatory monocytes (iMo) into the aortic root and coronary arteries. iMo differentiated into monocyte-derived dendritic cells (Mo-DC) in the vessel wall and were induced to release IL-1β in a Dectin-2-Syk-NLRP3 inflammasome dependent pathway. IL-1β then activated cardiac endothelial cells to express CXCL1 and CCL2 and adhesion molecules that induced neutrophil and further iMo recruitment and accumulation in the aortic root and coronary arteries. Our findings demonstrate that Dectin-2-mediated induction of CCL2 production by macrophages resident in the aortic root and coronary arteries initiates vascular inflammation in a model of Kawasaki disease, suggesting an important role for the innate immune system in initiating vasculitis.

481-P: KCNMA1 Transcriptional Regulation by Nrf2 in Diabetic Vessels

KCNMA1 encodes the large conductance Ca2+-activated K+ channel pore-forming subunits (BK-α), a major ionic mechanism that regulates vascular smooth muscle physiology and is impaired in high fat diet-induced (HFD) diabetic mice. The nuclear factor erythroid 2-related factor 2 (Nrf2) is a master transcription factor that regulates a wide variety of cellular proteins by binding to the antioxidant response elements (AREs). We found that the promoter region of KCNMA1 contains multiple AREs, but whether Nrf2 regulates KCNMA1 expression is unknown. In this study, we hypothesize that Nrf2 is a transcriptional regulator of KCNMA1 and the reduced Nrf2 activity in diabetes contributes to BK channelopathy. Using videomicroscopy, patch clamp recordings, molecular and cellular approaches, we found that BK channel activity and BK channel-mediated vasodilation were diminished in the coronary arteries of mice after 4 months on HFD. This was accompanied by significant reductions in BK-α and Nrf2 protein expression. In cultured coronary artery smooth muscle cells (CASMCs), adenoviral transduction with Nrf2 (50 MOI) resulted in an increase of BK-α mRNA by 10-fold and BK-α protein expression by 2-fold, while a 70% knockdown of Nrf2 by shRNA produced a 30% downregulation of BK-α protein levels. Luciferase reporter assays showed robust luciferase signals in cells 48 h after co-transfection with Nrf2 vectors and reporter vectors containing a segment of the KCNMA1 promoter region with a predicted ARE-Nrf2 binding site, but not those with empty vectors or KCNMA1 reporter vectors with the ARE-Nrf2 binding site deleted. Most importantly, treatment with dimethyl fumarate (a Nrf2 activator) increased BK-α mRNA and protein expression in cultured CASMCs, and rescued BK channel function and BK channel-mediated vasodilation in coronary arteries of HFD mice. Our results indicate that Nrf2 transcriptionally regulates KCNMA1 expression in CASMCs and has a therapeutic potential in the treatment of BK channelopathy and vasculopathy in diabetes. Disclosure H. Lee: None. X. Sun: None. X. Wang: None. T. Lu: None. Funding American Diabetes Association (1-18-IBS-210 to H-C.L.); National Institutes of Health (R01HL74180)

Cardioprotective effects of the total flavonoids of &lt;i&gt;Polygonum cuspidatum&lt;/i&gt; Sieb. et Zucc. Root extract on experimental myocardial infarction in mice

Purpose: To study the cardioprotective effects of the total flavonoids from the roots of Polygonum cuspidatum Sieb. et Zucc. (FHZ) on experimental myocardial infarction in mice. Methods: Ultrasonic-assisted extraction of FHZ was optimized by response surface methodology (RSM) to obtain a higher extraction yield. Myocardial infarction (MI) was established by ligation of the left anterior descending (LAD) branch of the coronary artery in mice. Cardiac troponin T (cTnT), creatine phosphokinase (CPK), lactate dehydrogenase (LDH), lipid peroxide (LPO), malondialdehyde (MDA) and superoxide dismutase (SOD) levels in the serum of mice were assessed by enzyme-linked immunosorbent assay. Furthermore, myocardial infarction size (MIS) was examined by Masson’s Trichrome staining on heart tissues. Results: Optimum extraction conditions of FHZ were as follows: an ethanol concentration of 69.03 %, a liquid-solid ratio of 27.14 g/mL and an extraction time of 30.30 min. The obtained extraction conditions were proven to be accurate and reliable. After treatment with FHZ for 3 and 7 days, the serum level of cTnT in MI mice decreased significantly. Also, the serum levels of CPK, LPO, MDA, and LDH were significantly decreased while SOD level increased in MI mice treated with FHZ. Furthermore, after treatment with FHZ for two weeks, the MIS of MI mice decreased ( p < 0.01). Conclusion: RSM is a useful tool to optimize the ultrasonic-assisted extraction conditions for FHZ. Furthermore, FHZ possesses significant cardioprotective effects on experimental myocardial infarction in mice and thus may find application in the clinical management of myocardial infarction. Keywords: Polygonum cuspidatum, Flavonoids, Response surface methodology, Myocardial infarction, Cardioprotective

Linggui Zhugan Decoction (苓桂术甘汤) Inhibits Ventricular Remodeling after Acute Myocardial Infarction in Rats by Suppressing TGF-β1/Smad Signaling Pathway

To investigate the inhibitory effect of Linggui Zhugan Decoction (LZD, ) on the ventricular remodeling (VR) after acute myocardial infarction (AMI) and related mRNA and proteins expression in transforming growth factor-beta 1 (TGF-β1)/Smad signaling pathway, and explain its putative mechanism. A VR model was generated by ligation of coronary artery in mice. Two weeks after surgery, 60 mice were randomly divided into the model group, the sham-operation group (distilled water), the positive control group (2.4 mg/kg simvastatin), and the low-, medium- and high-dose LZD groups (2.1, 4.2, 8.4 g crude drug/kg, respectively) by a random number table, 10 mice in each group. Mice in each group was treated for 4 weeks. Changes of hemodynamics indices and cardiac weight index were detected by the PowerLab data acquisition and analysis recording instrument. Morphology changes of myocardial tissue were observed by hematoxylin-eosin and Masson staining. The expressions of TGF-β1, Smad2, Smad3, p-Smad2 and p-Smad3 in myocardial tissue were detected by Western blotting. The mRNA expressions of TGF-β1, Smad2 and Smad3 were detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The expressions of matrix metalloprotein 2 (MMP2), MMP9, collagen I and collagen III were observed by immunohistochemical methods. VR mice showed significant dysfunction in hemodynamic indices and cardiac structure and function. Compared with the shamoperation group, myocardial tissue damage, interstitial fibrosis occurred in the model mice, left ventricular systolic pressure (LVSP), left ventricular pressure maximum contraction rate (+dp/dtmax) and left ventricular pressure maximum relaxation rate (-dp/dtmax) decreased significantly (all P<0.01), while left ventricular end-diastolic pressure (LVEDP), cardiac weight index and left ventricular weight index elevated significantly, meanwhile TGF-β1, p-Smad2, p-Smad3, Smad2, Smad3, MMP2, MMP9, collagen I, collagen III protein expressions in myocardial tissue and TGF-β1, Smad2 and Smad3 mRNA expressions increased significantly (all P<0.01). Compared with the model group, LZD could significantly improve the pathological changes of myocardial tissue, increase LVSP, +dp/dtmax and -dp/dtmax, lower LVEDP, reduce the whole heart weight index and left ventricular weight index and inhibit the over-expressions of TGF-β1, p-Smad2, p-Smad3, Smad2, Smad3, MMP2, MMP9, collagen I and collagen III proteins in myocardial tissue and mRNA expressions of TGF-β1, Smad2 and Smad3 (P<0.05 or P<0.01). LZD can significantly suppress VR induced by AMI, and its underlying mechanism may be associated with its inhibitory effect on the TGF-β1/Smad signaling pathway.