Inflammatory biomarkers predict higher risk of hyperglycemic crises but not outcomes in diabetic patients with COVID-19.

Background: Inflammation is a predictor of severe complications in patients with COVID-19 infection under a variety of clinical settings. A few studies suggested that COVID-19 infection was a trigger of hyperglycemic crises including diabetic ketoacidosis (DKA) and/or hyperglycemic hyperosmolar state (HHS). However, the association between inflammation and hyperglycemic crises in diabetic patients with COVID-19 infection is unclear. Methods: One hundred and twenty-four patients with type 2 diabetes mellitus (T2DM) and COVID-19 infection from January 2023 to March 2023 were retrospectively analyzed. Demographic, clinical, and laboratory data, especially inflammatory markers including white blood cell (WBC), neutrophils, neutrophil-to-lymphocyte ratio (NLR), c-reactive protein (CRP) and procalcitonin (PCT) were collected and compared between patients with or without DKA and/or HHS. Multivariable logistic regression analysis was conducted to explore the association between inflammatory biomarkers and the prevalence of hyperglycemic crises. Patients were followed up 6 months for outcomes. Results: Among 124 diabetic patients with COVID-19, 9 were diagnosed with DKA or HHS. Comparing COVID-19 without acute diabetic complications (ADC), patients with DKA or HHS showed elevated levels of c-reactive protein (CRP, P=0.0312) and procalcitonin (PCT, P=0.0270). The power of CRP and PCT to discriminate DKA or HHS with the area under the receiver operating characteristics curve (AUROC) were 0.723 and 0.794, respectively. Multivariate logistic regression indicated 1.95-fold and 1.97-fold increased risk of DKA or HHS with 1-unit increment of CRP and PCT, respectively. However, neither CRP nor PCT could predict poor outcomes in diabetic patients with COVID-19. Conclusion: In this small sample size study, we firstly found that elevated serum CRP and PCT levels increased the risk of hyperglycemic crises in T2DM patients with COVID-19 infection. More study is needed to confirm our findings.

Long noncoding RNA lncPostn links TGF-ß and p53 signaling pathways to transcriptional regulation of cardiac fibrosis.

Cardiac fibroblasts are essential for the homeostasis of the extracellular matrix, whose remodeling in many cardiovascular diseases leads to fibrosis. Long noncoding RNAs (lncRNAs) are associated with cardiac pathologies, but their functions in cardiac fibroblasts and contributions to cardiac fibrosis remain unclear. Here, we aimed to identify fibroblast-enriched lncRNAs essential in myocardial infarction (MI)-induced fibrosis and explore the molecular mechanisms responsible for their functions. Global lncRNA profiling was performed in post-MI mouse heart ventricles and transforming growth factor-ß (TGF-ß)-treated primary cardiac fibroblasts and confirmed in published data sets. We identified the cardiac fibroblast-enriched lncPostn, whose expression is stimulated in cardiac fibrosis induced by MI and the extracellular growth factor TGF-ß. The promoter of lncPostn contains a functional TGF-ß response element, and lncPostn knockdown suppresses TGF-ß-stimulated cardiac fibroblast activation and improves cardiac functions post-MI. LncPostn stabilizes and recruits EP300 to the profibrotic periostin's promoter, representing a major mechanism for its transcriptional activation. Moreover, both MI and TGF-ß enhance lncPostn expression while suppressing the cellular growth gatekeeper p53. TGF-ß and p53 knockdown-induced profibrotic gene expression and fibrosis occur mainly through lncPostn and show additive effects. Finally, levels of serum lncPostn are significantly increased in patients' postacute MI and show a strong correlation with fibrosis markers, revealing a potential biomarker of cardiac fibrosis. Our findings identify the fibroblast-enriched lncPostn as a potent profibrotic factor, providing a transcriptional link between TGF-ß and p53 signaling pathways to regulate fibrosis in cardiac fibroblasts.NEW & NOTEWORTHY Cardiac fibroblasts are essential for the homeostasis of the extracellular matrix, whose remodeling in many cardiovascular diseases leads to fibrosis. Long noncoding RNAs are functional and contribute to the biological processes of cardiovascular development and disorders. Our findings identify the fibroblast-enriched lncPostn as a potent profibrotic factor and demonstrate that serum lncPostn level may serve as a potential biomarker of human cardiac fibrosis postacute myocardial infarction.

Insight into the role of PCSK9 in glucose metabolism.

Diabetes mellitus (DM) is strongly associated with an increased risk of atherosclerotic cardiovascular disease (ASCVD). Proprotein convertase subtilisin/kexin type 9 (PCSK9) was recently identified as an important regulator of circulating low-density lipoprotein-cholesterol (LDL-C) levels via degradation of the LDL receptor, proving to be a valid target to improve lipoprotein profiles and cardiovascular outcomes in patients with ASCVD. Beyond LDL receptor processing and cholesterol homeostasis, the PCSK9 protein has recently been verified to be associated with glucose metabolism. Importantly, clinical trials suggest that treatment with PCSK9 inhibitors for patients with DM is more effective. Hence, in this review, we summarize the current findings derived from experimental, preclinical, and clinical studies regarding the association between PCSK9 and glucose metabolism, including the relationship of PCSK9 genetic mutations to glucose metabolism and diabetes, the link between plasma PCSK9 concentrations and glucose metabolic parameters, the effects of glucose-lowering drugs on plasma PCSK9 levels and the impacts of PCSK9 inhibitors on cardiovascular outcomes of patients with DM. Clinically, exploring this field may improve our understanding regarding the roles of PCSK9 in glucose metabolism and may offer an in-depth interpretation of how PCSK9 inhibitors exert effects on the treatment of patients with DM.

Triglyceride-glucose index for the detection of subclinical heart failure with preserved ejection fraction in patients with type 2 diabetes.

Objectives: The triglyceride-glucose (TyG) index has been identified as a reliable and simple surrogate of insulin resistance. In this study, we sought to determine the association between TyG index and cardiac function among asymptomatic individuals with type 2 diabetes (T2DM) without history of any cardiovascular disease. Materials and methods: The cross-sectional study enrolled 180 T2DM patients without cardiac symptoms. Heart failure with preserved ejection fraction (HFpEF) was defined as Heart Failure Association (HFA)-PEFF score ≥ 5 points. Results: A total of 38 (21.1%) diabetic patients were identified with HFpEF. Compared with the low-TyG group (TyG index <9.47), patients in high-TyG group (TyG index ≥9.47) showed increased risk of metabolic syndrome and diastolic dysfunction (p < 0.05 for each). Furthermore, after adjustment of confounding variables, the TyG index showed positive correlation with risk factors of metabolic syndrome (including BMI, waist circumference, blood pressure, HbA1c, TG, TC, non-HDL-C, and fasting blood glucose, p < 0.05 for each) and parameters of diastolic dysfunction (E/e' ratio, p < 0.0001) in patients with T2DM. Moreover, Receiver Operating Characteristic curve analysis showed that the TyG index could be better to predict the risk of suspected HFpEF than other indicators (AUC: 0.706, 95% CI: 0.612-0.801). According, on multiple regression analysis, TyG index was independently correlated with the incidence of HFpEF (odds ratio: 0.786, p = 0.0019), indicating that TyG index could be a reliable biomarker to predict the risk of HFpEF. Conclusion: The TyG index showed a positive correlation with the risk of subclinical HFpEF in patients with T2DM, providing a new marker to predict and treat HFpEF in diabetes.

Corrigendum: Reconstruction and Analysis of the lncRNA-miRNA-mRNA Network Based on Competitive Endogenous RNA Reveal Functional lncRNAs in Dilated Cardiomyopathy.

[This corrects the article DOI: 10.3389/fgene.2019.01149.].

Triglyceride-glucose index as a marker in cardiovascular diseases: landscape and limitations.

The triglyceride-glucose (TyG) index has been identified as a reliable alternative biomarker of insulin resistance (IR). Recently, a considerable number of studies have provided robust statistical evidence suggesting that the TyG index is associated with the development and prognosis of cardiovascular disease (CVD). Nevertheless, the application of the TyG index as a marker of CVD has not systemically been evaluated, and even less information exists regarding the underlying mechanisms associated with CVD. To this end, in this review, we summarize the history of the use of the TyG index as a surrogate marker for IR. We aimed to highlight the application value of the TyG index for a variety of CVD types and to explore the potential limitations of using this index as a predictor for cardiovascular events to improve its application value for CVD and provide more extensive and precise supporting evidence.

The Role of Mitochondrial Biogenesis Dysfunction in Diabetic Cardiomyopathy.

Diabetic cardiomyopathy (DCM) is described as abnormalities of myocardial structure and function in diabetic patients without other well-established cardiovascular factors. Although multiple pathological mechanisms involving in this unique myocardial disorder, mitochondrial dysfunction may play an important role in its development of DCM. Recently, considerable progresses have suggested that mitochondrial biogenesis is a tightly controlled process initiating mitochondrial generation and maintaining mitochondrial function, appears to be associated with DCM. Nonetheless, an outlook on the mechanisms and clinical relevance of dysfunction in mitochondrial biogenesis among patients with DCM is not completely understood. In this review, hence, we will summarize the role of mitochondrial biogenesis dysfunction in the development of DCM, especially the molecular underlying mechanism concerning the signaling pathways beyond the stimulation and inhibition of mitochondrial biogenesis. Additionally, the evaluations and potential therapeutic strategies regarding mitochondrial biogenesis dysfunction in DCM is also presented.

Erratum: Crucial Role of miR-433 in Regulating Cardiac Fibrosis: Erratum.

[This corrects the article DOI: 10.7150/thno.15007.].

Long Noncoding RNA Cardiac Physiological Hypertrophy-Associated Regulator Induces Cardiac Physiological Hypertrophy and Promotes Functional Recovery After Myocardial Ischemia-Reperfusion Injury.

BACKGROUND: The benefits of exercise training in the cardiovascular system have been well accepted; however, the underlying mechanism remains to be explored. Here, we report the initial functional characterization of an exercise-induced cardiac physiological hypertrophy-associated novel long noncoding RNA (lncRNA). METHODS: Using lncRNA microarray profiling, we identified lncRNAs in contributing the modulation of exercise-induced cardiac growth that we termed cardiac physiological hypertrophy-associated regulator (CPhar). Mice with adeno-associated virus serotype 9 driving CPhar overexpression and knockdown were used in in vivo experiments. Swim training was used to induce physiological cardiac hypertrophy in mice, and ischemia reperfusion injury surgery was conducted to investigate the protective effects of CPhar in mice. To investigate the mechanisms of CPhar's function, we performed various analyses including quantitative reverse transcription polymerase chain reaction, Western blot, histology, cardiac function (by echocardiography), functional rescue experiments, mass spectrometry, in vitro RNA transcription, RNA pulldown, RNA immunoprecipitation, chromatin immunoprecipitation assay, luciferase reporter assay, and coimmunoprecipitation assays. RESULTS: We screened the lncRNAs in contributing the modulation of exercise-induced cardiac growth through lncRNA microarray profiling and found that CPhar was increased with exercise and was necessary for exercise-induced physiological cardiac growth. The gain and loss of function of CPhar regulated the expression of proliferation markers, hypertrophy, and apoptosis in cultured neonatal mouse cardiomyocytes. Overexpression of CPhar prevented myocardial ischemia reperfusion injury and cardiac dysfunction in vivo. We identified DDX17 (DEAD-Box Helicase 17) as a binding partner of CPhar in regulating CPhar downstream factor ATF7 (activating transcription factor 7) by sequestering C/EBPß (CCAAT/enhancer binding protein beta). CONCLUSIONS: Our study of this lncRNA CPhar provides new insights into the regulation of exercise-induced cardiac physiological growth, demonstrating the cardioprotective role of CPhar in the heart, and expanding our mechanistic understanding of lncRNA function, as well.

Identification of a lncRNA-miRNA-mRNA network based on competitive endogenous RNA theory reveals functional lncRNAs in hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is an autosomal dominant disease that affects 1 in every 200 people in the general population, leading to cardiac ischemia, heart failure and increased risk of sudden death. Recently, accumulating evidence has suggested that long noncoding RNAs (lncRNAs) may serve specific roles in various biological processes and participate in the pathology of various diseases, including HCM. Although a large number of lncRNAs have been detected, the functions of lncRNAs in HCM are still unknown. In the present study, a global triple network based on competitive endogenous RNA (ceRNA) theory was constructed using data from the National Center for Biotechnology Information Gene Expression Omnibus. Furthermore, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses of mRNAs in the lncRNA-microRNA (miRNA)-mRNA network were performed using the Cytoscape plugins, BiNGO and Database. The lncRNA-miRNA-mRNA network was composed of 30 lncRNA nodes, 94 mRNA nodes and 8 miRNA nodes. Subsequently, hub nodes and the number of relationship pairs were analyzed and showed that 5 lncRNAs (ENST00000597346.1, ENST00000458178.1, ENST00000544461.1, ENST00000567093.1 and ENST00000571219.1) were closely related to HCM. Cluster module analysis and Random Walk with Restart of the ceRNA network further confirmed the potential role of two lncRNAs (ENST00000458178.1 and ENST00000567093.1) in HCM. The present study provides a new strategy for identifying potential pathways associated with HCM or other diseases. Furthermore, lncRNA-miRNA pairs may be regarded as candidate diagnostic biomarkers or potential therapeutic targets for HCM.

Value of circulating miRNA-21 in the diagnosis of subclinical diabetic cardiomyopathy.

BACKGROUND: Diabetic cardiomyopathy (DCM) is a type of cardiac dysfunction that affects approximately 12% of diabetic patients, ultimately leading to heart failure or even death. However, there is currently no efficient or specific biomarker for DCM diagnosis. METHODS: A total of 266 subjects with type II diabetes (T2DM) were enrolled in this study and were divided into the T2DM with cardiac dysfunction (DCM) group and T2DM without cardiac dysfunction (non-DCM) group. The diagnostic efficacy of miR-21 was determined and compared with that of serum hemoglobin A1c% (HbA1c%). Db/db mice and H9c2 cells stimulated with high glucose (HG)/high fatty acid (PA) were used as in vivo and in vitro models of DCM, respectively. RESULTS: Through echocardiography and gated-myocardial perfusion imaging (gated-MPI), 49 patients were selected to be enrolled in the DCM group, with 49 matched controls in the non-DCM group. The circulating miR-21 levels were significantly decreased in the DCM group compared to the non-DCM group (P < 0.001). The diagnostic efficiency of miR-21 (area under the curve AUC = 0.899) was higher than that of other parameters, including HbA1c%. Moreover, when miR-21 was combined with the duration of diabetes, HbA1c%, and lipid profiles, the AUC was the highest (AUC = 0.939) and had the highest diagnostic efficiency. Furthermore, overexpression of miR-21 improved the impaired mitochondrial biogenesis and decreased the cardiomyocyte apoptosis induced by HG/PA, while inhibition of miR-21 exerted the opposite effects. CONCLUSIONS: Our findings identify circulating miR-21 as a novel biomarker in the diagnosis of DCM and provide an underlying mechanism for miRNA-based therapy for the treatment of DCM. TRIAL REGISTRATION: The study was approved by the Ethics Committee of the Third Affiliated Hospital of Soochow University and has been registered in the Chinese Clinical Trial Registry (ChiCTR1900027080).

Neutrophil: lymphocyte ratio is positively associated with subclinical diabetic cardiomyopathy.

BACKGROUND: Subclinical diabetic cardiomyopathy (DCM) occurs frequently in asymptomatic subjects with Type 2 diabetes mellitus (T2DM). The direct association between the immune system and DCM with effective biomarkers has been demonstrated in previous studies. METHODS: Five hundred seven subjects with T2DM were recruited from April 2018 to October 2019 and divided into T2DM with cardiac dysfunction (DCM) group and T2DM without cardiac dysfunction (non-DCM) group. The relationship between the quartiles of Neutrophil: lymphocyte ratio (NLR) and subclinical DCM was evaluated by using adjusted logistic regression models.(covariates: age, sex, BMI, duration of diabetes, and hyperlipidemia). RESULTS: Blood NLR was significantly upregulated in DCM group compared to non-DCM group (P = 0.05). Then the adjusted odds ratio (95% CI) of the highest NLR quartile was 14.32 (2.92-70.31) compared with the lowest quartile of NLR after multiple adjusted (P < 0.001). However, there was no significant relation between neutrophil and lymphocyte counts and the occurrence of DCM in T2DM patients. CONCLUSIONS: This study demonstrated that NLR was associated with the occurrence of subclinical DCM, suggesting that NLR may be a biomarker for predicting DCM with effectiveness and accuracy. TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR1900027080) . Registered 30 October 2019. Retrospectively registered: www.medresman.org.

MiR-144 protects the heart from hyperglycemia-induced injury by regulating mitochondrial biogenesis and cardiomyocyte apoptosis.

Several lines of evidence have revealed the potential of microRNAs (miRNAs, miRs) as biomarkers for detecting diabetic cardiomyopathy, although their functions in hyperglycemic cardiac dysfunction are still lacking. In this study, mitochondrial biogenesis was markedly impaired induced by high glucose (HG), as evidenced by dysregulated mitochondrial structure, reduced mitochondrial DNA contents, and biogenesis-related mRNA levels, accompanied by increased cell apoptosis. MiR-144 was identified to be decreased in HG-induced cardiomyocytes and in streptozotocin (STZ)-challenged heart samples. Forced miR-144 expression enhanced mitochondrial biogenesis and suppressed cell apoptosis, while miR-144 inhibition exhibited the opposite results. Rac-1 was identified as a target gene of miR-144. Decreased Rac-1 levels activated AMPK phosphorylation and PGC-1α deacetylation, leading to increased mitochondrial biogenesis and reduced cell apoptosis. Importantly, the systemic neutralization of miR-144 attenuated mitochondrial disorder and ventricular dysfunction following STZ treatment. Additionally, plasma miR-144 decreased markedly in diabetic patients with cardiac dysfunction. The receiver-operator characteristic curve showed that plasma miR-144 could specifically predict diabetic patients developing cardiac dysfunction. In conclusion, this study provides strong evidence suggesting that miR-144 protects heart from hyperglycemia-induced injury by improving mitochondrial biogenesis and decreasing cell apoptosis via targeting Rac-1. Forced miR-144 expression might, thus, be a protective strategy for treating hyperglycemia-induced cardiac dysfunction.

Long Noncoding RNA FTX Reduces Hypertrophy of Neonatal Mouse Cardiac Myocytes and Regulates the PTEN/PI3K/Akt Signaling Pathway by Sponging MicroRNA-22.

BACKGROUND Cardiac myocyte hypertrophy results from clinical conditions that include hypertension and valvular heart disease, and can result in heart failure. This study aimed to investigate the expression and role of the long noncoding RNA FTX (lnc-FTX), an X-inactive-specific transcript (XIST) regulator transcribed from the X chromosome, in hypertrophy of neonatal mouse cardiac myocytes induced by angiotensin II (Ang II) in vitro. MATERIAL AND METHODS Cardiac myocytes were isolated from neonatal mice and cultured with and without Ang II. Immunofluorescence, with localization of an antibody to alpha-smooth muscle actin (alpha-SMA), was used to identify the neonatal mouse cardiac myocytes. Quantitative real-time polymerase chain reaction (qRT-PCR) measured gene expression levels. The cell counting kit-8 (CCK-8) assay was used to determine cell viability, and Western blot measured protein expression. StarBase v2.0 bioinformatics software was used for target gene prediction and was confirmed with the luciferase reporter assay. RESULTS The expression of lnc-FTX was reduced in mouse cardiac myocytes treated with Ang II. Overexpression of lnc-FTX reduced cell apoptosis, cardiomyocyte contractility, and the expression of c-Jun, A-type natriuretic peptide (ANP), and B-type natriuretic peptide (BNP) induced by Ang II. The target of lnc-FTX was micro-RNA 22 (miRNA-22). The mechanism of action of lnc-FTX in neonatal mouse cardiac myocytes was through suppression of the PI3K/Akt signaling pathway by promoting the release of PTEN by sponging miRNA-22. CONCLUSIONS The expression of lnc-FTX was associated with reduced hypertrophy of neonatal mouse cardiac myocytes and regulated the PTEN/PI3K/Akt signaling pathway by sponging miRNA-22.

Reconstruction and Analysis of the lncRNA-miRNA-mRNA Network Based on Competitive Endogenous RNA Reveal Functional lncRNAs in Dilated Cardiomyopathy.

Dilated cardiomyopathy (DCM) is an important cause of sudden death and heart failure with an unknown etiology. Recent studies have suggested that long non-coding RNA (lncRNA) can interact with microRNA (miRNA) and indirectly interact with mRNA through competitive endogenous RNA (ceRNA) activities. However, the mechanism of ceRNA in DCM remains unclear. In this study, a miRNA array was first performed using heart samples from DCM patients and healthy controls. For further validation, we conducted real-time quantitative reverse transcription (RT)-PCR using samples from DCM patients and a doxorubicin-induced rodent model of cardiomyopathy, revealing that miR-144-3p and miR-451a were down-regulated, and miR-21-5p was up-regulated. Based on the ceRNA theory, we constructed a global triple network using data from the National Center for Biotechnology Information Gene Expression Omnibus (NCBI-GEO) and our miRNA array. The lncRNA-miRNA-mRNA network comprised 22 lncRNA nodes, 32 mRNA nodes, and 11 miRNA nodes. Hub nodes and the number of relationship pairs were then analyzed, and the results showed that two lncRNAs (NONHSAT001691 and NONHSAT006358) targeting miR-144/451 were highly related to DCM. Then, cluster module and random walk with restart for the ceRNA network were analyzed and identified four lncRNAs (NONHSAT026953/NONHSAT006250/NONHSAT133928/NONHSAT041662) targeting miR-21 that were significantly related to DCM. This study provides a new strategy for research on DCM or other diseases. Furthermore, lncRNA-miRNA pairs may be regarded as candidate diagnostic biomarkers or potential therapeutic targets of DCM.

Upregulation of Circular RNA CircNFIB Attenuates Cardiac Fibrosis by Sponging miR-433.

Cardiac fibrosis is the pathological consequence of fibroblast proliferation and fibroblast-to-myofibroblast transition. As a new class of endogenous non-coding RNAs, circular RNAs (circRNAs) have been identified in many cardiovascular diseases including fibrosis, generally acting as microRNA (miRNA) sponges. Here, we report that the expression of circRNA-circNFIB was decreased in mice post-myocardial infarction heart samples, as well as in primary adult cardiac fibroblasts treated with TGF-ß. Forced expression of circNFIB decreased cell proliferation in both NIH/3T3 cells and primary adult fibroblasts as evidenced by EdU incorporation. Conversely, inhibition of circNFIB promoted adult fibroblast proliferation. Furthermore, circNFIB was identified as a miR-433 endogenous sponge. Overexpression of circNFIB could attenuate pro-proliferative effects induced by the miR-433 mimic while inhibition of circNFIB exhibited opposite results. Finally, upregulation of circNFIB also reversed the expression levels of target genes and downstream signaling pathways of miR-433. In conclusion, circNFIB is critical for protection against cardiac fibrosis. The circNFIB-miR-433 axis may represent a novel therapeutic approach for treatment of fibrotic diseases.

The Exosome: a New Player in Diabetic Cardiomyopathy.

Diabetic cardiomyopathy (DCM) or diabetes-induced cardiac dysfunction is a direct consequence of uncontrolled metabolic syndrome and occurs worldwide. However, the underlying cellular and molecular mechanisms remain poorly understood. Recently, exosomes have attracted considerable interest for their use as efficient, targeted, and non-immunogenic delivery systems for biological molecules or pharmacotherapies. This review will summarize the fast-developing field of the regulation and function of exosomes in DCM, affording valuable insights and therapeutic opportunities in combatting diabetes-related cardiac disorder for modern human health.

Recombinant human glucagon-like peptide-1 protects against chronic intermittent hypoxia by improving myocardial energy metabolism and mitochondrial biogenesis.

BACKGROUND AND AIMS: Obstructive sleep apnea syndrome is a chronic disease associated with intermittent hypoxia (IH) and is an important risk factor for cardiovascular disease. Glucagon-like peptide (GLP-1) is a naturally occurring incretin used as a promising therapeutic agent in the treatment of acute myocardial infarction, dilated cardiomyopathy, and advanced heart failure. However, whether GLP-1 can protect against IH-induced cardiac injury is still unclear. Accordingly, in this study, we evaluated the effects of recombinant human GLP-1 (rhGLP-1) on cardiac health in mice. METHODS: Mice were subjected to repetitive 5% O2 for 30 s and 21% O2 for 30 s, for a total of 8 h/day for 4 weeks. Subsequently, mice received subcutaneous injection of saline or rhGLP-1 (100 µg/kg, three times per day). Cardiac function, myocardial apoptosis and fibrosis, energy metabolism, and mitochondrial biogenesis were examined for evaluation of cardiac injury. RESULTS: A reduction in diastolic function (E/A ratio) in mice exposed to IH was significantly reversed by rhGLP-1. IH induced marked cardiomyocyte apoptosis and myocardial fibrosis. Additionally, IH resulted in a shift from fatty acid to glucose metabolism in the myocardium with downregulation of peroxisome proliferator-activated receptor (PPAR) α and PPARγ. Moreover, IH caused a reduction in mitochondrial DNA (mtDNA) replication and transcription, together with reduced mtDNA content and impaired mitochondrial ultrastructure. These changes were abolished by rhGLP-1 via activation of PGC-1α and Akt signaling. CONCLUSIONS: rhGLP-1 protects against IH-induced cardiac injury by improving myocardial energy metabolism and enhancing the early adaptive changes of mitochondrial biogenesis.

Neonatal Rat Cardiomyocytes Isolation, Culture, and Determination of MicroRNAs' Effects in Proliferation.

Cardiomyocytes loss is a major contributor for many cardiovascular diseases including heart failure and myocardial infarction. Although extremely limited, adult cardiomyocytes are able to proliferate. Understanding the molecular mechanisms controlling cardiomyocytes proliferation is extremely important for enhancing cardiomyocyte proliferation to promote cardiac regeneration and repair. MicroRNAs (miRNAs, miRs) are powerful controllers of many essential biological processes including cell proliferation. Here, we described in detail a protocol for isolation and culture of neonatal rat cardiomyocytes and the determination of miRNAs' effects in proliferation based on two well-established methods including EdU and Ki67 immunofluorescent stainings.