Ox-LDL Causes Endothelial Cell Injury Through ASK1/NLRP3-Mediated Inflammasome Activation via Endoplasmic Reticulum Stress.

OBJECTIVE: This study was to investigate the mechanism of inflammatory pathology modification induced by ox-LDL in endothelial cells. METHODOLOGY: In this study, we firstly investigated the efflux of cholesterol of endothelial cells under the treatment of ox-LDL, and cell proliferation, ROS production, cell apoptosis was measured. Further, proteins of ASK1, NLRP3 inflammasomes and endoplasmic reticulum stress response were detected. Afterwards, ASK1 inhibitor (GS-4997) or endoplasmic reticulum stress (ERS) inhibitor (4-PBA) was used to measure the performance of endothelial cells. RESULTS: In this study, endothelial cells were treated with ox-LDLs alone or in combination with a GS-4997 or 4-PBA. Results showed that ox-LDLs attenuated the efflux of cholesterol from endothelial cells in a dose-dependent manner. Ox-LDLs inhibited the proliferation of endothelial cells, and induced their apoptosis and production of reactive oxygen species (ROS). Additionally, ox-LDLs upregulated the levels of phosphorylated ASK1, ERS-related proteins (chop, p-PERK, GRP78, and p-IRE-1), and inflammation-associated proteins (NLRP3, IL-1ß, and caspase 1) in endothelial cells. Moreover, we proved that GS-4997 could partly reverse ox-LDL-mediated cell proliferation, apoptosis, ROS production, and inflammation in endothelial cells, and increase cholesterol efflux. We also found that 4-PBA could attenuate the effects of ox-LDLs on endothelial cell cholesterol efflux, proliferation, apoptosis, ROS production, and inflammation. CONCLUSION: Our results suggest that cholesterol efflux from endothelial cells is reduced by ox-LDLs, and these reductions in cholesterol efflux are accompanied by increased NLRP3 inflammasome signaling, ASK1 and higher levels of endoplasmic reticulum stress. Our results suggest this axis as potential targets for treating atherosclerosis.

Higenamine Improves Cardiac and Renal Fibrosis in Rats With Cardiorenal Syndrome via ASK1 Signaling Pathway.

The pathogenesis of cardiorenal syndrome (CRS) is very complex, and currently there is no effective treatment for CRS. Higenamine (HI) has been shown to improve cardiac function in rats with heart failure. However, the role of higenamine in CRS remains unknown. Here, in vitro, higenamine treatment markedly reduced neonatal rat cardiac fibroblast collagen synthesis and inhibited neonatal rat cardiac myocyte hypertrophy. In our study, a rat model of type 2 CRS was induced by left anterior descending coronary artery ligation combined with 5/6 subtotal nephrectomy (STNx). Higenamine treatment decreased serum creatinine (Scr), blood urea nitrogen, and brain natriuretic peptide levels and was capable of improving left ventricular remodeling and systolic function in CRS rats, accompanied with decreased expression of transforming growth factor-ß1 (TGF-ß1), α-smooth muscle actin (α-SMA) and collagen I (Col1A1). Moreover, higenamine significantly inhibited the protein expression of phosphorylated apoptosis signal-regulated kinase 1 (p-ASK1) and downstream mitogen-activated protein kinases (MAPK) (ERK, P38)/NF-κB in cardiorenal tissues of CRS rats and neonatal rat cardiac fibroblast/neonatal rat cardiac myocyte cells. Our study demonstrated that higenamine improved cardiorenal function in CRS rats and attenuated heart and kidney fibrosis possibly via targeting ASK1/MAPK (ERK, P38)/NF-κB signaling pathway. This finding extends our knowledge on the role of higenamine in cardiorenal fibrosis, providing a potential target to prevent the progression of CRS.

[Zhenwu Decoction delays ventricular hypertrophy in rats with uremic cardiomyopathy].

OBJECTIVE: To investigate the inhibitory effect of Zhenwu Decoction on ventricular hypertrophy in rats with uremic cardiomyopathy and explore the mechanism. METHODS: Cardiocytes isolated from suckling rats were divided into control group and indoxyl sulfate (IS) group, and the protein synthesis was assayed with [3H]- leucine incorporation and cellular protein expressions were detected using Western blotting. Fifty SD rats were randomly divided into sham operation group, model group, and low- and high-dose Zhenwu Decoction treatment groups, and except for those in the sham operation group, all the rats underwent 5/6 nephrectomy. Four weeks after the operation, the rats in low- and high-dose treatment groups were given Zhenwu Decoction via gavage at the dose of 4.5 g/kg and 13.5 g/kg, respectively; the rats in the sham-operated and model groups were given an equal volume of distilled water. After 4 weeks of treatment, serum levels of IS were determined, and cardiac and ventricular mass indexes were measured in the rats; cardiac ultrasound was performed and Western blotting was used to measure the expressions of BNP, p-ERK1/2, p-p38 and p-JNK in the myocardium. RESULTS: Rat cardiomyocytes treated with IS showed significantly enhanced protein synthesis and increased expression levels of BNP, p-erk1/2, and p-p38 as compared with the control cells (P < 0.01), but the expression of p-jnk was comparable between the two groups. In the animal experiment, the rats in the model group showed significantly increased serum creatinine (SCr) and urea nitrogen (BUN) levels, 24-h urine protein (24 hUpro), plasma IS level, left ventricular mass index (LVMI) and whole heart mass index (HMI) compared with those in the sham group (P < 0.01); Both LVESD and LVEDD were significantly reduced and LVAWS, LVAWD, LVPWS and LVPWD were significantly increased in the model rat, which also presented with obvious cardiomyocyte hypertrophy and increased myocardial expressions of BNP, p-ERK1/2, p-p38 and p-jnk (P < 0.01). Compared with the rats in the model group, the rats treated with low-dose and high-dose Zhenwu Decoction had significantly lowered levels of SCr, BUN, 24 hUpro and IS (P < 0.05) and decreased LVMI and HMI; LVESD, LVEDD, LVPWS, LVAWS, and LVAWD were improved more obviously in the high-dose group, and the myocardial expressions of BNP, p-ERK1/2, p-p38 and p-JNK was significantly downregulated after the treatment. CONCLUSIONS: Zhenwu Decoctin can reduce plasma IS levels and inhibit ventricular hypertrophy to delay ventricular remodeling in rats with uremic cardiomyopathy.