Dexmedetomidine Attenuates Hypoxia/Reoxygenation Injury of H9C2 Myocardial Cells by Upregulating miR-146a Expression via the MAPK Signal Pathway.

INTRODUCTION AND OBJECTIVE: Dexmedetomidine (Dex) and a number of miRNAs contribute to ischemia/reperfusion injury. We aimed to explore the role of Dex and miR-146a on myocardial cells injured by hypoxia/reoxygenation (H/R). METHOD: H9C2 cells were injured by H/R. Cell viability was tested using the cell counting kit-8. Lactate dehydrogenase (LDH) activity, superoxide dismutase (SOD) activity, and malondialdehyde (MDA) levels were determined using commercial kits. Flow cytometry was performed to determine apoptosis rate and reactive oxygen species (ROS) level. Protein and mRNA levels were assessed using Western blot and qPCR. RESULTS: miR-146a expression and cell viability of H9C2 cells were downregulated under the circumstance of H/R injury. The tendency could be reversed by Dex, which could also upregulate SOD activity and decrease apoptosis, LDH activity, MDA, 78-kDa glucose-regulated protein (GRP78), and C/EBP homologous protein (CHOP) levels of H9C2 cells. GRP78, CHOP levels, and cell viability were negatively modulated by miR-146a. Dex elevated cell viability, catalase, MnSOD, and NAD(P)H dehydrogenase (NQO1) levels but suppressed apoptosis rate, GRP78, and CHOP levels by increasing miR-146a expression and downregulating ROS, phosphorylation of p38, and extracellular signal-regulated kinases 1/2 levels. By using SB203580 (SB), the p38 mitogen-activated protein kinase (MAPK) inhibitor, Dex or the inhibition of miR-146 upregulated cell viability but downregulated GRP78 and CHOP levels. CONCLUSION: Dex might regulate miR-146a expression, which could further modulate the endoplasmic reticulum stress and oxidative stress and eventually affect the cell viability and apoptosis of myocardial cells injured by H/R via the MAPK signal pathway.

Bakuchiol Alleviates Hyperglycemia-Induced Diabetic Cardiomyopathy by Reducing Myocardial Oxidative Stress via Activating the SIRT1/Nrf2 Signaling Pathway.

Bakuchiol (BAK), a monoterpene phenol reported to have exerted a variety of pharmacological effects, has been related to multiple diseases, including myocardial ischemia reperfusion injury, pressure overload-induced cardiac hypertrophy, diabetes, liver fibrosis, and cancer. However, the effects of BAK on hyperglycemia-caused diabetic cardiomyopathy and its underlying mechanisms remain unclear. In this study, streptozotocin-induced mouse model and high-glucose-treated cell model were conducted to investigate the protective roles of BAK on diabetic cardiomyopathy, in either the presence or absence of SIRT1-specific inhibitor EX527, SIRT1 siRNA, or Nrf2 siRNA. Our data demonstrated for the first time that BAK could significantly abate diabetic cardiomyopathy by alleviating the cardiac dysfunction, ameliorating the myocardial fibrosis, mitigating the cardiac hypertrophy, and reducing the cardiomyocyte apoptosis. Furthermore, BAK achieved its antifibrotic and antihypertrophic actions by inhibiting the TGF-ß1/Smad3 pathway, as well as decreasing the expressions of fibrosis- and hypertrophy-related markers. Intriguingly, these above effects of BAK were largely attributed to the remarkable activation of SIRT1/Nrf2 signaling, which eventually strengthened cardiac antioxidative capacity by elevating the antioxidant production and reducing the reactive oxygen species generation. However, all the beneficial results were markedly abolished with the administration of EX527, SIRT1 siRNA, or Nrf2 siRNA. In summary, these novel findings indicate that BAK exhibits its therapeutic properties against hyperglycemia-caused diabetic cardiomyopathy by attenuating myocardial oxidative damage via activating the SIRT1/Nrf2 signaling.

Downregulated microRNA­133a induces HUVECs injury: Potential role of the (pro) renin receptor in angiotensin II­dependent hypertension.

The renin­angiotensin system (RAS) serves an essential role in hypertension. MicroRNAs (miRs) have been reported to be important regulators in angiotensin (Ang) II­dependent hypertension. We aimed to explore the roles of Ang II and miR­133a in the mechanism underlying hypertension. Human umbilical vein endothelial cells (HUVECs) were identified by immunofluorescence staining. Cell viability and miR­133a expression under the inhibition of Ang II of various concentrations were determined by an MTT assay and reverse transcription­quantitative polymerase chain reaction (RT­qPCR), respectively. The effects of HUVECs transfected with miR­133a mimic or inhibitor on Ang II­induced apoptosis were measured using flow cytometry. The potential targeting of miR­133a to the 3' untranslated region of (pro) renin receptor (PRR) was assessed using TargetScan and a dual­luciferase assay. The effects of PRR interference using small interfering (si)RNA on PRR expression and the rate of apoptosis were determined by RT­qPCR, western blotting and flow cytometry, respectively. Ang II at a concentration of 10­5 M significantly inhibited the cell viability (P<0.05) and miR­133a expression (P<0.01); Downregulation of miR­133a suppressed cell viability. HUVECs transfected with miR­133a mimic reduced the rate of Ang II­induced apoptosis from 21.99 to 12.38%, but miR­133a inhibitor promoted Ang II­induced apoptosis (apoptosis rate, 28.9%). PRR was predicted to be a target gene of miR­133a. Transfection with siPRR decreased the apoptotic rate in Ang II + negative control and Ang II + miR­133a inhibitor group to 11.39 and 12.94%, respectively. Our findings also suggested that Ang II promoted PRR expression to enhance the apoptotic rate of HUVECs via the suppression of miR­133a. Furthermore, siPRR efficiently decreased the Ang II­induced apoptosis.

Pentraxin-3 in coronary artery disease: A meta-analysis.

AIMS: Studies on the prognostic significance of circulating pentraxin-3 level in patients with coronary artery disease (CAD) have yielded conflicting results. The aim of this meta-analysis was to evaluate the prognostic value of circulating pentraxin-3 level in CAD patients. MATERIALS/METHODS: We made a systematic literature search in Pubmed, Embae, CNKI, Wanfang, and VIP database from their inception to January 10, 2019 for prospective cohort studies that investigated the association between pentraxin-3 level and adverse outcomes in patients with CAD. The outcome measures were all-cause mortality, cardiac death, and cardiac events (cardiac death, nonfatal myocardial infarction, heart failure or coronary revascularization). Multivariable-adjusted risk ratio (RR) with 95% confidence intervals (CI) was pooled for the highest versus the lowest pentraxin-3 group to summarize the predictive value. RESULTS: Nine studies were included, enrolling 5,174 CAD patients. Overall, CAD patients with the highest pentraxin-3 level had an increased risk of all-cause mortality (RR 1.81; 95% CI 1.43-2.28), cardiac death (RR 1.77; 95% CI 1.38-2.26), and cardiac events (RR 1.61; 95% CI 1.16-2.25). However, elevated pentraxin-3 level appeared to not significantly increase the risk of cardiac events (RR 1.63; 95% CI 0.71-3.72) in stable CAD subgroup. CONCLUSIONS: In CAD patients, elevated circulating pentraxin-3 level is possibly an independent predictor of all-cause mortality, cardiac death, and cardiac events. However, interpretation of these findings should be with caution due to the small number of studies analyzed.

The long-term differentiation of embryonic stem cells into cardiomyocytes: an indirect co-culture model.

BACKGROUND: Embryonic Stem Cells (ESCs) can differentiate into cardiomyocytes (CMs) in vitro but the differentiation level from ESCs is low. Here we describe a simple co-culture model by commercially available Millicell™ hanging cell culture inserts to control the long-term differentiation of ESCs into CMs. METHODOLOGY/PRINCIPAL FINDINGS: Mouse ESCs were cultured in hanging drops to form embryoid bodies (EBs) and treated with 0.1 mmol/L ascorbic acid to induce the differentiation of ESCs into CMs. In the indirect co-culture system, EBs were co-cultured with epidermal keratinocytes (EKs) or neonatal CMs (NCMs) by the hanging cell culture inserts (PET membranes with 1 µm pores). The molecular expressions and functional properties of ESC-derived CMs in prolonged culture course were evaluated. During time course of ESC differentiation, the percentages of EBs with contracting areas in NCMs co-culture were significantly higher than that without co-culture or in EKs co-culture. The functional maintenance of ESC-derived CMs were more prominent in NCMs co-culture model. CONCLUSIONS/SIGNIFICANCE: These results indicate that NCMs co-culture promote ESC differentiation and has a further effect on cell growth and differentiation. We assume that the improvement of the differentiating efficiency of ESCs into CMs in the co-culture system do not result from the effect of co-culture directly on cell differentiation, but rather by signaling effects that influence the cells in proliferation and long-term function maintenance.

Three-dimensional co-culture facilitates the differentiation of embryonic stem cells into mature cardiomyocytes.

The cardiomyocyte (CM) differentiation of embryonic stem cells (ESCs) is routinely cultured as two-dimensional (2D) monolayer, which doesn't mimic in vivo physiological environment and may lead to low differentiated level of ESCs. Here, we develop a novel strategy that enhances CM differentiation of ESCs in collagen matrix three-dimensional (3D) culture combined with indirect cardiac fibroblasts co-culture. ESCs were cultured in hanging drops to form embryoid bodies (EBs) and then applied on collagen matrix. The EBs were indirectly co-cultured with cardiac fibroblasts by the hanging cell culture inserts (PET 1 µm). The molecular expressions and ultrastructural characteristics of ESC-derived CMs (ESCMs) were analyzed by real time RT-PCR, immunocytochemistry, and Transmission Electron Microscopy (TEM). We found that the percentage of beating EBs with cardiac fibroblasts co-culture was significantly higher than that without co-culture after differentiation period of 8 days. Type I collagen used as 3D substrates enhanced the late-stage CM differentiation of ESCs and had effect on ultrastructural mature of ESCMs in late-stage development. The combined effects of 3D and co-culture that mimic in vivo physiological environment further improved the efficiency of CM differentiation from ESCs, resulting in fiber-like structures of cardiac cells with organized sarcomeric structure in ESCMs. This novel 3D co-culture system emphasizes the fact that the ESC differentiation is actively responding to cues from their environment and those cues can drive phenotypic control, which provides a useful in vitro model to investigate CM differentiation of stem cells.