GDF11 Attenuated ANG II-Induced Hypertrophic Cardiomyopathy and Expression of ANP, BNP and Beta-MHC Through Down- Regulating CCL11 in Mice.
Growth differentiation factor 11 (GDF11) decreases with age, and increased C-C motif chemokine 11 (CCL11) is involved in aging. However, the effects of GDF11 on Angiotensin II (ANG II)-induced hypertrophic cardiomyopathy and expression of markers for volume overload and hypertrophy such as ANP, BNP and beta-MHC, as well as the relationship between GDF11 and CCL11 in hypertrophic cardiomyopathy are unclear. Therefore, the current study aimed to examine the effects of GDF11 on ANG II-induced hypertrophic cardiomyopathy and expression of ANP, BNP and beta-MHC in mice, and explore possible molecular mechanisms.
Vectors were constructed and viruses were packaged. Mouse cardiomyocytes were treated with ANG II for 24 h. Meanwhile, mouse cardiomyocytes were divided into 4 groups: (1) control; (2) ANG II; (3) ANG II+GDF11; and (4) ANG II+CCL11. Furthermore, mouse cardiomyocytes were treated with GDF11 and CCL11 proteins for 48 h, respectively. The thickness of IVS and LVPS during systole and diastole were measured by cardiac ultrasound in the mouse model of hypertrophic cardiomyopathy. The relative expression of ANP, BNP, beta-MHC, CCL11 and GDF11 in cardiomyocytes or heart tissue of mice was detected by qPCR or Western blot. 3'- UTR luciferase reporter assay was utilized to examine the relationship between GDF11 and the expression of CCL11.
The expression of ANP, BNP, and beta-MHC in mouse cardiomyocytes was significantly increased after the cells were treated with 800 nM ANG II, which was utilized in the following cell experiments. After ANG II treatment, 0.2 ng/ml GDF11 group displayed the highest inhibition of expression of ANP, BNP and beta-MHC in mouse cardiomyocytes, whereas 50 ng/ml CCL11 group displayed the highest stimulation of the expression. GDF11 at 10 ng/ml significantly decreased the expression of CCL11 in mouse cardiomyocytes as compared to the control group. Mice treated with ANG II had increased thickness of IVS and LVPS during both systole and diastole, which was significantly attenuated by GDF11 overexpression. GDF11 overexpression attenuated the increase in expression of ANP, BNP and beta-MHC in the mice model of hypertrophic cardiomyopathy. The relative serum concentration of GDF11 was markedly decreased, and CCL11 was dramatically increased in mice with hypertrophic cardiomyopathy. GDF11 overexpression restored the serum concentration of GDF11 and CCL11 in the mice model of hypertrophic cardiomyopathy. In addition, GDF11 interference group had markedly increased expression of CCL11, whereas GDF11 overexpression group had significantly decreased expression of CCL11 in luciferase reporter assay.
GDF11 attenuated ANG II-induced hypertrophic cardiomyopathy and expression of ANP, BNP and beta-MHC through down-regulating CCL11 in mice.
Zhang C
，Wang Y
，Ge Z
，Lin J
，Liu J
，Yuan X
，Lin Z
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Overexpression of miR-142-3p improves mitochondrial function in cardiac hypertrophy.
Our previous studies have shown that Src homology 2 (SH2) B adaptor protein 1 (SH2B1) plays an important role in cardiac hypertrophy, but the specific mechanism remains to be studied. Through bioinformatics and related research, it is found that miR-14 2-3 p is closely related to SH2B1. Exploring the relationship between miR-14 2-3 p and gene SH2B1 expression is beneficial for the treatment of cardiac hypertrophy. SH2B1 is a key factor regulating energy metabolism, mitochondria are the main organelles of energy metabolism and cardiac hypertrophy are closely related to mitochondrial dysfunction. So it is particularly important to explore the relationship between miR-14 2-3 p and SH2B1 and myocardial mitochondrial function. In this study, we investigated whether overexpression of miR-14 2-3 p can inhibit the expression of gene SH2B1, ameliorate cardiac mitochondrial dysfunction and cardiac hypertrophy.
We first constructed a pressure overload myocardial hypertrophy model by ligation of the abdominal aorta(AB) of rats. After 4 weeks of modeling, echocardiographic examination showed that the heart volume of the model group became larger, and Hematoxylin and Eosin Staining Kit (HE) staining showed that the cross-sectional area of the heart tissue became larger. The expression of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), β-Myosin Heavy Chain (β-MHC) messenger RNA (mRNA) increased by real‑time polymerase chain reaction (PCR), which proved that the model of cardiac hypertrophy was successfully constructed. Then, miR-14 2-3 p agomir was injected into the tail vein of rats 2 weeks and 4 weeks respectively. The expression of miR-4 2-3 p mRNA was increased by PCR, suggesting that the miR-14 2-3 p plasmid was successfully transfected. At 4 weeks of pressure overload myocardial hypertrophy model, echocardiography was used to detect cardiac function. HE staining of heart tissue and the expression of ANP, BNP, β-MHC mRNA were used to detect cardiac hypertrophy. Flow cytometry was used to detect changes in mitochondrial membrane potential. Secondly, we observed the effect of miR-14 2-3 p on cardiomyocyte hypertrophy and mitochondrial function in vitro by culture neonatal rat cardiomyocytes. Afterwards, using angiotensin (Ang)II-, miRNA mimic- and miRNA mimic nc- treated cardiomyocytes for a given time. α-actin staining found that the myocardial cells became larger, The expression of ANP, BNP, β-MHC mRNA increased by PCR, which proved that AngII-induced cardiac hypertrophy was successfully constructed. Then, the mitochondrial density was measured using mitochondrial Mito-Red staining by Confocal microscope, the mitochondrial membrane potential was evaluated using flow cytometry, Mitochondrial respiration oxygen consumption rate (OCR) was measured by a Seahorse Extracellular Flux Analyzer XF96, and the expression levels of miR-14 2-3 p, ANP, BNP, β-MHC mRNA, SH2B1 in the cardiomyocytes of different groups were measured by RT-PCR and Western blotting. Finally, we used luciferase assay and transfected miR-14 2-3 p agomir in rats, transfected miR-14 2-3 p mimic in Cardiomyocytes, it is found that myocardial SH2B1 mRNA and protein expression both were reduced.
When the pressure overload myocardial hypertrophy model was constructed for four weeks, echocardiography revealed that the heart volume, Left ventricular end diastolic diameter(LVIDd), Left ventricular end systolic diameter (LVIDs), Left ventricular posterior wall thickness (LVPWd), Systolic left ventricular posterior wall (LVPWs), Left ventricle (LV) Mass increased, Ejection fraction (EF) % decreased of AB group increased, but transfected with miR-14 2-3 p agomir of AB, these increase was not significant, EF% reduction was not obvious. HE staining showed that the myocardial cross-sectional area of AB group increased significantly, but the miR-14 2-3 p agomir treatment of AB group did not increase significantly. PCR analysis showed that the expression of ANP, BNP,β-MHC mRNA was significantly increased in AB group, but the miR-14 2-3 p agomir treatment of AB group was not significantly increased. Flow cytometry showed that the mitochondrial membrane potential of AB group was significantly reduced, and the miR-14 2-3 p agomir treatment of AB group was not significantly decreased. During AngII-induced cardiomyocyte hypertrophy, ANP, BNP,β-MHC mRNA expression was increased, while these factors was not significantly increased in miR-14 2-3 p mimic treatment group; mitochondrial membrane potential, mitochondrial density and OCR was significantly decreased in AngII treated group, and these were not significantly reduced in miR-14 2-3 p mimic treatment group; CONCLUSIONS: miR-14 2-3 p not only mitigate cardiac hypertrophy by directly inhibit the expression of gene SH2B1, but also can protect mitochondrial function in cardiac hypertrophy of vitro and vivo.
Liu BL
，Cheng M
，Hu S
，Wang S
，Wang L
，Tu X
，Huang CX
，Jiang H
，Wu G
... -
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MicroRNA-22 downregulation by atorvastatin in a mouse model of cardiac hypertrophy: a new mechanism for antihypertrophic intervention.
Growing evidence shows that microRNAs (miRNAs) are involved in various cardiac processes including cardiac hypertrophy. However, the modulation of miRNA by pharmacological intervention in cardiomyocyte hypertrophy has not been disclosed yet. methods: We constructed neonatal rat cardiomyocyte hypertrophy induced by angiotensin II stimulation and subjected to cardiomyocyte immunochemistry, qRT-PCR and immunoblotting analysis. In addition, we constructed the mouse cardiac hypertrophy using angomir-22 stimulation and demonstrated the potential antihypertrophic mechnism of atorvastatin.
The results showed that a collection of miRNAs were aberrantly expressed in hypertrophic cardiomyocytes induced by angiotensin II stimulation. In addition, overexpression of miR-22 was found in angiotensin II-induced hypertrophic cardiomyocytes, whereas administration of atorvastatin could reverse the upregulation of miRNA-22 nearly back to the control level. Furthermore, up-regulation of miRNA-22 in cardiomyocytes in vitro and in vivo could induce cardiac hypertrophy, which could suppress the protein level of phosphatase and tensin homolog deleted on chromosome ten (PTEN). Concomitantly, the production of ANP, BNP and β-MHC was enhanced and cardiomyocyte size was increased. However, atorvastatin could markedly knockdown miRNA-22 expression and reverse these changes in cardiomyocytes. These results suggest that the contribution of atrovastatin to cardiomyocyte hypertrophy may be involved in downregulation of miRNA-22 expression, which modulates the activity of PTEN in cardiomyocyte hypertrophy. conclusion: This study demonstrates for the first time the modulation of miRNA-22 can be achieved by pharmacological intervention. The data generated from this study provides a rationale for the development of miRNA-based strategies for antihypertrophic treatment.
Tu Y
，Wan L
，Bu L
，Zhao D
，Dong D
，Huang T
，Cheng Z
，Shen B
... -
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