Chronic intermittent hypoxia-induced BNIP3 expression mitigates contractile dysfunction and myocardial injury in animal and cell model via modulating autophagy.

Obstructive sleep apnea syndrome is generally associated with multiple cardiovascular disorders, such as myocardial hypertrophy. Autophagy is strictly modulated to maintain cardiac homeostasis. Post-injury autophagy is closely associated with pathological cardiac hypertrophy. BCL2 interacting protein 3 (BNIP3) and BNIP3-like protein (BNIP3L) can cause cell death and are important for hypoxia-elicited autophagy. Here, we evaluated whether BNIP3 could mitigate functional remodeling and cardiac hypertrophy through regulation of autophagy. Male WT rats or rats with BNIP3 knockout were subjected to chronic intermittent hypoxia (CIH) for 8 h/day over 5 weeks. Echocardiography and morphology were employed to assess the cardioprotective effects. Autophagy was assessed via transmission electron microscopy and detection of LC3, p62, and Beclin-1. Terminal deoxynucleotidyl transferase dUTP nick end labeling and the Bax/Bcl2 ratio were used to monitor apoptosis. Biochemical evaluations were performed to assess oxidative stress. Additionally, BNIP3-knockdown H9c2 cells that were subjected to CIH were used to examine autophagy and apoptosis to confirm the findings of the animal study. The CIH group showed elevated heart weight/body weight and left ventricle weight/body weight proportions, along with left ventricular remodeling. CIH-exposed rats exhibited dramatically higher fractional shortening and ejection fractions than the controls. In addition, the levels of autophagy markers Beclin-1 and LC3-II/I were increased, whereas the level of p62 was reduced by CIH treatment. The oxidative marker levels and the apoptosis index in the CIH group were markedly increased. Knockout of BNIP3 significantly aggravated the impairment in cardiac function, apoptosis, oxidative stress, and hypertrophy of CIH rats, while significantly reducing autophagy. The autophagy-associated PI3K/Akt/mTOR pathway was also deactivated by BNIP3 knockout. At the cellular level, CIH treatment significantly upregulated autophagy and apoptosis; however, BNIP3 silencing reduced autophagy and promoted apoptosis. CIH treatment-mediated upregulation of BNIP3 expression plays a crucial role in autophagy by targeting the PI3K/Akt/mTOR pathway, alleviating cardiac hypertrophy.

Long noncoding RNA THRIL promotes foam cell formation and inflammation in macrophages.

Tumor necrosis factor-α (TNF-α) and heterogenous nuclear ribonucleoprotein L (hnRNPL)-related immunoregulatory lincRNA (THRIL) is a long noncoding RNA (lncRNA) involved in various inflammatory diseases. However, its role in atherosclerosis is not known. In this study, we aimed to investigate the function of THRIL in mediating macrophage inflammation and foam cell formation. The expression of THRIL was quantified in THP-1 macrophages after treatment with oxidized low-density lipoprotein (oxLDL). The effect of THRIL overexpression and knockdown on oxLDL-induced inflammatory responses and lipid accumulation was determined. THRIL-associated protein partners were identified by RNA pull-down and RNA immunoprecipitation assays. We show that THRIL is upregulated in macrophages after oxLDL treatment. Knockdown of THRIL blocks oxLDL-induced expression of interleukin-1ß (IL-1ß), IL-6, and TNF-α and lipid accumulation. Conversely, ectopic expression of THRIL enhances inflammatory gene expression and lipid deposition in oxLDL-treated macrophages. Moreover, THRIL depletion increases cholesterol efflux from macrophages and the expression of ATP-binding cassette transporter (ABC) A1 and ABCG1. FOXO1 is identified as a protein partner of THRIL and promotes macrophage inflammation and lipid accumulation. Furthermore, overexpression of FOXO1 restores lipid accumulation and inflammatory cytokine production in THRIL-depleted macrophages. In conclusion, our data suggest a model where THRIL interacts with FOXO1 to promote macrophage inflammation and foam cell formation. THRIL may represent a therapeutic target for atherosclerosis.

[Silencing miRNA210 Alleviates Myocardial Cell Damage in Myocarditis by Hypoxia Inducible Factor-vascular Endothelial Growth Factor Pathway].

Objective To investigate the effect of miRNA210 on primary myocardial cells in lipopolysaccharide(LPS)-induced myocarditis.Methods CCK8 method was used to detect the effect of miRNA210 on the viability of primary myocardial cells in normal or LPS-induced myocarditis rats.ELISA was performed to detect the secretion of tumor necrosis factor(TNF)-α and interleukin(IL)-1ß after miRNA210 treatment.Flow cytometry was used to detect the apoptosis of primary myocardial cells before and after the intervention.Western blotting was used to detect the expression of TNF-α and IL-1ß.The expression of apoptosis-related proteins bcl-2,bax,caspase-3,and hypoxia inducible factor 1 (HIF1)-vascular endothelial growth factor(VEGF)were detected by Western blotting.Results CCK8 detection results showed that,compared with the control group,the effect of miRNA210 mimic(t=0.000,P=1.000)and siRNA(t=0.686,P=0.500)interference had no significant difference on primary rat cardiomyocytes.The viability of rat primary cardiomyocytes significantly decreased after LPS treatment(t=8.764,P<0.001);compared with LPS group,the viability of rat primary cardiomyocytes significantly increased after inhibition of miRNA210(t=3.576, P=0.012).ELISA showed that,compared with the control group,the expressions of IL-1 ß(t=4.166,P=0.014)and TNF-α(t=6.309,P=0.003)were significantly up-regulated after LPS induction;compared with the LPS group,the expressions of IL-1 ß(t=4.096,P=0.015)and TNF-α(t=4.424,P=0.011)were significantly increased after application of miRNA210 mimic.After silencing miRNA210,IL-1 ß(t=4.287,P=0.012)and TNF-α(t=3.577,P=0.023)were significantly down-regulated.Flow cytometry showed that,compared with the control group,the apoptosis of primary cardiomyocytes induced by LPS was significantly increased(t=32.780,P<0.001);compared with LPS group,the apoptosis induced by LPS was significantly aggravated by miRNA210 mimic(t=7.412,P= 0.002),and the apoptosis rate of primary cardiomyocytes was significantly decreased after miRNA210 was silenced(t=11.720,P<0.001).Western blot analysis showed that,compared with the control group,LPS significantly down-regulated the expression of bcl-2(t=8.346,P=0.001)and increased the expressions of bax(t=12.890,P<0.001)and caspase-3(t=4.331,P=0.012).Compared with LPS group,the expression of bcl-2(t=6.074,P<0.001)was significantly decreased,and the expressions of bax(t=5.376,P=0.022)and caspase-3(t=5.859,P=0.004)were increased after miRNA210 mimic.After silencing miRNA210,the expression of bcl-2 significantly increased(t=3.873,P=0.017),the expressions of bax(t=5.205,P=0.006)and caspase-3(t=2.800,P=0.040)significantly decreased.Compared with the control group,the expressions of HIF1(t =10.380,P=0.001)and VEGF(t=4.973,P=0.008)were significantly up-regulated in LPS group.Compared with LPS group,the expressions of HIF1(t=8.952,P<0.001)and VEGF(t=11.203,P=0.001)were significantly up-regulated after miRNA210 mimic was applied,while HIF1(t=3.893,P=0.017)and VEGF expression(t=3.181,P=0.033)were decreased after miRNA210 was silenced.Compared with LPS group,the expressions of bax(t= 4.899,P=0.008),HIF1(t=2.833,P=0.047),caspase-3(t=2.877,P=0.045),and VEGF(t= 2.994, P=0.040)were significantly decreased,and the expression of bcl-2 was increased(t=3.392,P=0.017).Conclusion Silencing miRNA210 can attenuate LPS-induced cardiomyocyte injury through HIF1-VEGF-mediated apoptotic pathway.