A positive FOXP3/lncRNA SNHG1 feedback axis ameliorates cardiomyocytes hypertrophy by negatively regulating Parkin-mediated mitophagy.

In this study, we identified FOXP3 as a transcription factor for lncRNA SNHG1, which exerts a significant protective role against cardiomyocyte hypertrophy. Through DNA-pull down experiments and ChIP analysis, we confirmed that FOXP3 could bind to the promoter of SNHG1. Luciferase reporter and RT-qPCR experiments validated that FOXP3 overexpression promoted SNHG1 expression in cardiomyocytes. Furthermore, in a model of cardiomyocyte hypertrophy, FOXP3 expression was upregulated, particularly in cardiomyocytes. Functional assays demonstrated that FOXP3 overexpression inhibited cardiomyocyte hypertrophy, while FOXP3 knockdown held the opposite effect. Additionally, we revealed that lncRNA SNHG1 acted as a sponge for miR-182, miR-326, and miR-3918, thereby stabilizing FOXP3 mRNA in cardiomyocytes. The protective role of SNHG1 against cardiomyocyte hypertrophy was found to depend on the presence of FOXP3, forming a positive FOXP3/SNHG1 feedback axis. Moreover, we unveiled this positive FOXP3/SNHG1 feedback axis suppressed cardiomyocyte hypertrophy by negatively regulating Parkin-mediated mitophagy. These findings provide novel insights into the molecular mechanisms underlying cardiomyocyte hypertrophy and offer potential therapeutic targets for related interventions.

MicroRNA-194 inhibits isoproterenol-induced chronic cardiac hypertrophy via targeting CnA/NFATc2 signaling in H9c2 cells.

Background: This study explored the effects of microRNA(miR)-194 on chronic cardiac hypertrophy (CH) induced by isoproterenol (ISO). The potential mechanism through regulation of the calcineurin A (CnA)/nuclear factor of activated T cells (NFAT) c2 pathway was investigated in the rat cardiomyoblast cell line H9c2. Methods: H9c2 cells were treated with ISO to induce cardiomyocyte hypertrophy to simulate CH in vitro. The cell surface area was assessed by phalloidin staining. The expression of miR-194, CnA mRNA, and CnA protein were assessed. Furthermore, the cellular localization of the NFATc2 protein after induction of CH was detected. The relationship between miR-194 and the CnA mRNA 3'-untranslated region (UTR) was verified by dual luciferase report assays. By constructing cardiomyocyte cell models with low expression of miR-194 and/or CnA, the effects of miR-194 and CnA on the localization of the NFATc2 protein and cell hypertrophy was investigated. Rescue experiments were conducted to analyze whether overexpression of miR-194 could alleviate the cell hypertrophy induced by ISO. Results: The results demonstrated that induction with ISO significantly increased the surface area of H9c2 cells. After induction, the expression of miR-194 decreased, while both CnA mRNA and protein expression increased. Furthermore, the nuclear translocation of NFATc2 was obvious. MiR-194 bound to the 3'-UTR of CnA mRNA and inhibited CnA protein expression. Inhibition of miR-194 expression activated NFATc2 protein expression and increased the H9c2 cell surface area. After CnA expression was disturbed, hypertrophy induced by miR-194 down-regulation was blocked. In addition, overexpression of miR-194 significantly alleviated cell hypertrophy and activation of the CnA/NFATc2 pathway caused by ISO. Conclusions: In conclusion, increasing the expression of miR-194 can alleviate CH by targeting can and inhibiting the CnA/NFATc2 pathway.

Cilostazol alleviate nicotine induced cardiomyocytes hypertrophy through modulation of autophagy by CTSB/ROS/p38MAPK/JNK feedback loop.

Nicotine is proved to be an important factor for cardiac hypertrophy. Autophagy is important cell recycling system involved in the regulation of cardiac hypertrophy. Cilostazol, which is often used in the management of peripheral vascular disease. However, the effects of cilostazol on nicotine induced autophagy and cardiac hypertrophy are unclear. Here, we aim to determine the role and molecular mechanism of cilostazol in alleviating nicotine-induced cardiomyocytes hypertrophy through modulating autophagy and the underlying mechanisms. Our results clarified that nicotine stimulation caused cardiomyocytes hypertrophy and autophagy flux impairment significantly in neonatal rat ventricular myocytes (NRVMs), which were evidenced by augments of LC3-II and p62 levels, and impaired autophagosomes clearance. Interestingly, cathepsin B (CTSB) activity decreased dramatically after stimulation with nicotine in NRVMs, which was crucial for substrate degradation in the late stage of autophagy process, and cilostazol could reverse this effect dramatically. Intracellular ROS levels were increased significantly after nicotine exposure. Meanwhile, p38MAPK and JNK were activated after nicotine treatment. By using ROS scavenger N-acetyl-cysteine (NAC) could reverse the effects of nicotine by down-regulation the phosphorylation of p38MAPK and JNK pathways, and pretreatment of specific inhibitors of p38MAPK and JNK could restore the autophagy impairment and cardiomyocytes hypertrophy induced by nicotine. Moreover, CTSB activity of lysosome regained after the treatment with cilostazol. Cilostazol also inhibited the ROS accumulation and the activation of p38MAPK and JNK, which providing novel connection between lysosome CTSB and ROS/p38MAPK/JNK related oxidative stress pathway. This is the first demonstration that cilostazol could alleviate nicotine induced cardiomyocytes hypertrophy through restoration of autophagy flux by activation of CTSB and inhibiting ROS/p38/JNK pathway, exhibiting a feedback loop on regulation of autophagy and cardiomyocytes hypertrophy.

Regulation of the Golgi apparatus via GOLPH3-mediated new selective autophagy.

AIMS: Although previous studies elaborated that selective autophagy was involved in quality control of some organelles, including nucleus, mitochondria, the endoplasmic reticulum and peroxisomes, it remained unclear whether the selective autophagy of the Golgi apparatus (Golgiphagy) existed or not. MAIN METHODS: In this study, H9c2 cells, HUVECs, HA-VSMCs and HEK293T cells were treated with autophagy inducers, Golgi stress inducers and cardiomyocytes hypertrophy stimulators. The Golgiphagy was evaluated by analysing the co-localization of Golgi markers and LC3B. Furthermore, the transmission electron microscope was used to observe the occurrence of Golgiphagy. The co-immunoprecipitation assay was used to evaluate the interaction of GOLPH3 and LC3B. KEY FINDINGS: Results showed that starvation promoted the co-localization of both GM130-positive and TGN46-positive Golgi fragments with LC3B-positive autophagosomes in H9c2 cells, HUVECs, HA-VSMCs and HEK293T cells. Transmission electron microscopy images showed that Golgi apparatus was sequestered into the autophagosomes in the starvation group. Moreover, Golgi stress inducers also facilitated the co-localization of Golgi markers and LC3B in H9c2 cells, HUVECs, HA-VSMCs and HEK293T cells. Furthermore, cardiomyocyte hypertrophy stimulators also triggered the appearance of Golgiphagy in H9c2 cells. Importantly, the co-immunoprecipitation assay indicated endogenous GOLPH3 interacted with LC3B in H9c2 cells, HUVECs, HA-VSMCs. However, knocking down GOLPH3 inhibited the Golgiphagy. SIGNIFICANCE: This study unveiled a new selective autophagy of the Golgi apparatus (Golgiphagy). In addition, GOLPH3 might act as a novel cargo receptor to regulate Golgiphagy. Maintaining homeostasis of the Golgi apparatus via GOLPH3-mediated autophagy was indispensable for cell survival.

Role and molecular mechanism of RGS6 in regulation of cardiomyocyte hypertrophy / 中国药理学通报

Aim To explore the regulatory effect and mechanism of Regulators of G-protein Signaling 6 (RGS6) in Ang H-induced hypertrophy. Methods RGS6 expression was up-and down-regulated respectively by AdRGS6 and AdshRGSó lentivirus. Each group was treated with Ang II /PBS. Cell size and changes of hypertrophic markers(ANP and BNP) were evaluated. Then ROS levels and the total protein and phosphorylated protein of E R K 1 / 2, p38, JNK1/2 in each group were detected. When the altered signaling was clarified, a reversal experiment of hypertrophic cardiomyocytes was conducted to further verify the signaling pathway. The RGS6 up-regulated cardiomyocytes of hypertrophy treated with ROS inhibitor (DPI) were experimental group. Results (Î) After stimulated with Ang II for 48 h, the size of AdRGS6-infected cell increased compared with AdGFP infection group. In addition, the mRNA levels of ANP and BNP also i n - creased. After stimulated with Ang II , the ROS levels and p - J N K l / 2 proteins of AdRGS6-infected cells all increased compared with AdGFP infection group. (2) After stimulated with Ang II for 48 h, the size of A d - shRGS6-infected cell decreased compared with AdshRNA infection group and the mRNA levels of ANP and BNP also decreased. The ROS levels and p - J N K l / 2 proteins of AdshRGS6-infected cell all decreased compared with AdshRNA infection group. (3) In the reversal experiments, compared with AdRGS6 + Ang 1 1 / DMSO group, the size of AdRGS6 + Ang H / D P I cardiomyocytes significantly decreased, and the downstream signaling of p - J N K l / 2 was improved. Conclusions RGS6 may be a key factor to cardiac hypertrophy. RGS6 aggravates Ang II -induced cardiomyocyte hypertrophy via activating R 0 S - K N K 1 / 2 signaling pathway.

Ferritinophagy activation and sideroflexin1-dependent mitochondria iron overload is involved in apelin-13-induced cardiomyocytes hypertrophy.

Excess iron accumulation and cardiac oxidative stress have been shown as important mediators of cardiac hypertrophy, whereas it remains largely elusive about the occurrence of mitochondrial iron overload and its significance during cardiac hypertrophy. In the present study, we aim to investigate the role of NCOA4-mediated ferritinophagy and SFXN1-dependent mitochondria iron overload in apelin-13-induced cardiomyocytes hypertrophy. Apelin-13 significantly promotes ferric citrate (FAC)-induced total cellular and mitochondria ion production, as well as mitochondria ROS contents. Mechanistically, apelin-13 effectively induces the expression of SFXN1, a mitochondria iron transporting protein and NCOA4, a cargo receptor of ferritinophagy in dose and time-dependent manner. Conversely, blockade of APJ by F13A abolishes these stimulatory effects. In addition, apelin-13-triggered mitochondria iron overload is reversed by the genetic inhibition of SFXN1 and NCOA4. NCOA4 deficiency via its silencing also interferes with the enhanced expression of SFXN1 evoked by apelin-13. In apelin-13-treated H9c2 cells, the promotion in cell diameter, volume as well as protein contents are obviously suppressed by the knockdown of NCOA4 and SFXN1 with their corresponding siRNAs. Remarkably, the human and murine hypertrophic hearts models, as well as apelin-13-injected mice models, present evident cardiac mitochondrial iron deposition and raised expressions of NCOA4 and SFXN1. Taken together, these results provide experimental evidences that NCOA4-mediated ferritinophagy might be defined as an essential mechanism leading to apelin-13-cardiomyocytes hypertrophy in SFXN1-dependent mitochondria iron overload manners.

miR-200a-5p augments cardiomyocyte hypertrophy induced by glucose metabolism disorder via the regulation of selenoproteins.

Selenium and selenoproteins are identified as potential determinants in pathological cellular hypertrophy. Cardiomyocytes hypertrophy is a compensatory form of heart disease characterized by increased size of cardiomyocytes. However, the link between cardiac hypertrophy and Se-specific microRNA (miRNA) remains to be characterized. In the current study, we established a miR-200a-5p mimic and an inhibitor cardiomyocytes model. Cardiomyocytes hypertrophy was induced in the miR-200a-5p mimic group. Hence, we detected the glucose level of cardiomyocytes to estimate the cellular glucose uptake. The effect of miR-200a-5p overexpression and the low expression on 25 selenoproteins mRNA levels was further explored using reverse transcription polymerase chain reaction. Overexpression of miR-200a-5p elevated glucose uptake and Txnrd2, 3 expression and reduced Sepp1, Seln, Selt, and Sep15 expression in cardiomyocytes. Contrary results were observed in cardiomyocytes with the knockdown of miR-200a-5p. We next assessed glucose metabolism-related genes in cardiomyocytes. The results showed that miR-200a-5p had a negative correlation with insulin-like growth factor gene-1, insulin-like growth factor binding protein (IGFBP)1, IGFBP2, IGFBP3, IGFBP4, and IGFBP5 and had a positive correlation with Akt, glucose transporter family (GLUT)2, GLUT3, and GLUT4. These results support the involvement of selenoproteins and glucose metabolism in the control of cardiomyocytes hypertrophy by Se-specific miRNA, suggesting that miR-200a-5p inhibited the expression of stress-related selenoproteins to alter glucose transport leading to glucose metabolism disorder, eventually inducing cardiomyocytes hypertrophy. Our finding highlights a pivotal role of Se-specific miRNA and selenoproteins in cardiac hypertrophy.

Heat shock protein 22 reduces phenylephrine-induced cardiomyocytes hypertrophy / 中华急诊医学杂志

Objective To investigate the effect of Hsp22 on phenylephrine-induced cardiomyocytes hypertrophy.Methods Primary rat myocardial cells were isolated and cultured in Department of Cardiology,the First Affiliated Hospital of Zhengzhou University.Cells were divided into four groups randomly:Control group,model group,treatment group with 1 μg/mL Hsp22,and treatment group with 10 μg/mL Hsp22.Phenylephrine stimuli was used to induce cardiomyocytes hypertrophy model.Cell viability was measured by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.Cardiomyocytes surface area was evaluated by α-actin immunofluorescence staining.Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the transcription level of hypertrophic markers.Reactive oxygen species level was detected by 2',7'-Dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescent probe.Apoptosis was detected by TUNEL staining.Signal pathway protein expression was detected by Western blot.SPSS 13.0 was used for statistical analysis.Data were expressed as mean + standard deviation.All data were analyzed by one-way ANOVA between groups.Comparisons between two groups were performed using LSD-t test.A P＜0.05 was considered statistically significant.Results Different concentrations of Hsp22 had no effect on cardiomyocytes viability (F=6.622;P＞0.05).Phenylephrine stimulation significantly increased cardiomyocytes area (t=10.80;P＜0.05),increased the transcription level of hypertrophy markers atrial natriuretic peptide (t=37.72;P＜0.05),type B natriuretic peptide (t=16.85;P＜0.05),and myosin heavy chain beta (t=41.53;P＜0.05).Different concentrations of Hsp22 significantly reduced cardiomyocytes area (PE+ 1 μg/mL Hsp22 t=4.018;P＜0.05;PE+10 μg/mL Hsp22 t=10.80;P＜0.05),reduced the transcription level of hypertrophic markers atrial natriuretic peptide (PE+1 μg/mL Hsp22 t=27.12,P＜0.05;PE+10 μg/mL Hsp22 t=37.72,P＜0.05),type B natriuretic peptide (PE+1 μg/mL Hsp22 t=4.82,P＜0.05;PE+10 μg/mL Hsp22 t=12.74,P＜0.05),and myosin heavy chain beta (PE+1 μg/mL Hsp22 t=23.68,P＜0.05;PE+10 μg/mL Hsp22 t=30.54,P＜0.05).Westem blot showed that Hsp22 increased the activation of AMP-activated protein kinase α (PE+1 μg/mL Hsp22 t=5.89,P＜0.05;PE+10 μg/mL Hsp22 t=5.88,P＜0.05),reduced mTOR phosphorylation level (PE+1 μg/mL Hsp22 t=16.80,P＜0.05;PE+10.μg/mL Hsp22 t=20.46,P＜0.05).Conclusions Hsp22 inhibits cardiomyocytes hypertrophy by activating AMP-activated protein kinase α.Hsp22 may become a potential anti-hypertrophic drug.

Curcumin attenuates cardiomyocyte hypertrophy induced by high glucose and insulin via the PPARγ/Akt/NO signaling pathway.

AIM: To investigate the potential effect of curcumin on cardiomyocyte hypertrophy and a possible mechanism involving the PPARγ/Akt/NO signaling pathway in diabetes. METHODS: The cardiomyocyte hypertrophy induced by high glucose (25.5mmol/L) and insulin (0.1µmol/L) (HGI) and the antihypertrophic effect of curcumin were evaluated in primary culture by measuring the cell surface area, protein content and atrial natriuretic factor (ANF) mRNA expression. The mRNA and protein expressions were assayed by reverse transcription PCR and Western blotting, whereas the NO concentration and endothelial NO synthase (eNOS) activity were determined using nitrate reduction and ELISA methods, respectively. RESULTS: The cardiomyocyte hypertrophy induced by HGI was characterized by increasing ANF mRNA expression, total protein content, and cell surface area, with downregulated mRNA and protein expressions of both PPARγ and Akt, which paralleled the declining eNOS mRNA expression, eNOS content, and NO concentration. The effects of HGI were inhibited by curcumin (1, 3, 10µmol/L) in a concentration-dependent manner. GW9662 (10µmol/L), a selective PPARγ antagonist, could abolish the effects of curcumin. LY294002 (20µmol/L), an Akt blocker, and N(G)-nitro-l-arginine-methyl ester (100µmol/L), a NOS inhibitor, could also diminish the effects of curcumin. CONCLUSIONS: The results suggested that curcumin supplementation can improve HGI-induced cardiomyocytes hypertrophy in vitro through the activation of PPARγ/Akt/NO signaling pathway.

Inhibitory effects of κ-opioid receptor stimulation on ne-induced cardiac hypertrophy of the cultured myocardial cells / 中国药理学通报

AIM: To study the effects of κ-opioid receptor activation on the cardiac hypertrophy, cultured cardiomyocytes were to used study the inhibitory effects of U58,488H on cellular volume and norepinephrine(NE)-induced hypertrophy in the presence or absence of norbinaltorphimine(nor-BNI). METHODS: The protein content was assayed with Lowry's method. The cardiomyocytes' volumes was measured by an eyepiece graticule of inverted microscope. The contracting frequency of cultured myocytes was counted by the inverted microscope. RESULTS: A κ-opioid receptor agonist U50, 488H at (0.1 ∼ 10 μmol·L-1) inhibited the protein content of cultured myocardial cells in normal serum medium in a dose-dependent manner. The inhibitory effects of U50,488H were completely blocked by pretreatment with nor-BNI, a specific κ-opioid receptor antagonist at 1 μmol·L-1. U50,488H (0.1 ∼ 10 μmol·L-1) also inhibited NE-induced cellular hypertrophy in low serum medium, which also were abolished by nor-BNI (1 μmol· L-1). CONCLUSIONS: U50,488H inhibited NE-induced hypertrophy. The inhibitory effects of U50,488H are involved in mediating the action of NE-induced cardiac hypertrophy.

Effects of Ca~(2+) /CaM-dependent calcineurin signaling pathway on cardiomyocytes hypertrophy of rats induced by neuropeptide Y / 中国临床药理学与治疗学

AIM : To investigate the effects of Ca 2+ /CaM dependent calcineurin(CaN) signaling pathway on cardiomyocytes hypertrophy of rat induced by neuropeptide Y(NPY). METHODS : Cardiomyocytes of neonatal Wistar rats were cultured with NPY of various concentrations (10,100 nmol?L -1 ). Cyclosporine A (CsA) was used to inhibit the activity of CaN. The methods of 3H Leu incorporation was used to assess protein synthesis rate in cardiomyocytes. Western blot and histochemistry were used to measure CaN protein expression and CaN activity in cardiomyocytes. RESULTS : 3 H Leu incorporation of cardiomyocytes were increased significantly by 100 nmol?L -1 NPY ( P

Inhibitory effects of ?-opioid receptor stimulation on NE-induced cardiac hypertrophy of the cultured myocardial cells / 中国药理学通报

AIM To study the effects of ?-opioid receptor activation on the cardiac hypertrophy, cultured cardiomyocytes were to used study the inhibitory effects of U58,488H on cellular volume and norepinephrine(NE)-induced hypertrophy in the presence or absence of norbinaltorphimine(nor-BNI). METHODS The protein content was assayed with Lowry's method. The cardiomyocytes'volumes was measured by an eyepiece graticule of inverted microscope. The contracting frequency of cultured myocytes was counted by the inverted microscope. RESULTS A ?-opioid receptor agonist U50,488H at (0.1～10 ?mol?L -1 ) inhibited the protein content of cultured myocardial cells in normal serum medium in a dose-dependent manner. The inhibitory effects of U50,488H were completely blocked by pretreatment with nor-BNI, a specific ?-opioid receptor antagonist at 1 ?mol?L -1 . U50,488H (0.1～10 ?mol?L -1 ) also inhibited NE-induced cellular hypertrophy in low serum medium, which also were abolished by nor-BNI(1 ?mol?L -1 ). CONCLUSIONS U50,488H inhibited NE-induced hypertrophy. The inhibitory effects of U50,488H are involved in mediating the action of NE-induced cardiac hypertrophy.

Effect of iptakalim on prevention isoprenaline induced cardiomyocytes hypertrophy / 中国药理学通报

Aim To investigate the effect of iptakalim on neonatal cardiomyocytes hypertrophy induced by isoprenaline(ISO).Methods Hypertrophy in neonatal rat cardiac myocytes was established via culture with ISO.Total protein content was measured by Lowrys method.The fluorescence intensity of intracellular Ca2+ was detected respectively with Fluo-3/AM loading by the laser confocal microscopy.Results ① 1?10-5mol?L-1 ISO can activated ?-AR completely;② the total protein of cardiomyocytes and intracellular i was markedly decreased by treatment with IPT and represented a dose-dependent manner.Conclusion IPT could inhibit the increase of protein content of cardiomyocytes induced by ISO,which may be contributed to the decease of intracellular Ca2+ concentration.