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Decoy receptor‑3 regulates inflammation and apoptosis via PI3K/AKT signaling pathway in coronary heart disease
- Authors:
- Published online on: January 30, 2019 https://doi.org/10.3892/etm.2019.7222
- Pages: 2614-2622
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Copyright: © Chen et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
Abstract
Introduction
Coronary heart disease is a generic term for coronary arteries atherosclerosis lesions caused by a large number of factors, such as vascular cavity stenosis, occlusion, myocardial ischemia, hypoxia, necrosis, inflammation and apoptosis of myocardial cells (1,2). It has been demonstrated that reduced blood pressure and cholesterol are important factors in reducing deaths from coronary heart disease (3). In addition, a large number of proteins are associated with the protective effects for host against myocardial ischemia and reperfusion injury through modulating myocardial apoptosis and inflammation (4,5). Furthermore, inflammation and apoptosis contribute to the initiation and development of coronary heart disease, which have been considered as the prognostic indicators for patients with coronary heart disease following drug treatment in clinical settings (6,7). Therefore, a number of potential strategies to relieve or mitigate the apoptosis of myocardial cells have been proposed to protect the heart against coronary heart disease, such as ameliorating the apoptosis of cardiac cells mediated by endothelial stem cell (8), or activating the signaling pathway to promote the expression of anti-apoptosis proteins and decrease pro-apoptosis proteins to protect the heart (9).
Coronary heart disease is the leading cause of mortality worldwide and is closely associated with metabolism disorders of endogenous substances (10). A growing number of studies have reported that apoptosis of myocardial cells has a crucial role in the progression of cardiovascular diseases (11–13). Apoptosis of leukocytes has been considered as a marker of neutrophil-endotheliocyte interaction in coronary heart disease (14). Expression of apoptosis factors has become an important reference of surgical results following coronary stent implantation in coronary heart disease patients determined by characteristics of neutrophil cluster of differentiation (CD)11b, platelet CD62P, endothelin, monocyte CD11b, and neutrophil CD178 in patients with coronary heart disease prior to and following primary coronary stenting (15).
Inflammation as the cause of coronary heart disease has been reviewed previously (16). Expression levels of inflammation cytokines including interleukin (IL)-6, tumor necrosis factor (TNF)-α and IL-1 in patients with coronary heart disease may also be associated with the progression pathogenesis (17). These reports indicate that inhibition of inflammation contributes to the treatment of coronary heart disease. In the present study, the inhibitory effects of decoy receptor-3 (DCR-3) on inflammation were investigated.
It has been demonstrated that DCR-3 can act as a pleiotropic immunomodulator for enhancing angiogenesis, which may be associated with the progression of coronary heart disease (18). DCR-3 is well known as soluble receptor TR6, which belongs to the TNFR superfamily and is constituted by a 300-aa polypeptide (19,20). DCR-3 has been identified as the most common biomarker of tumor deterioration in cancer patients by a previous meta-analysis (21). However, no previous report has focused on the clinical significance of DCR-3 expression levels in coronary and peripheral blood of coronary heart disease.
In the present study, the DCR-3 concentration levels were investigated in patients with coronary heart disease. The efficacy and molecular mechanism of DCR-3 were evaluated in mice with coronary heart disease. Inflammation and apoptosis of myocardial cells were investigated in mice with coronary heart disease following treatment with DCR-3. The phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway was also analyzed in DCR-3-treated myocardial cells.
Materials and methods
Ethics statement
A total of 242 patients (age, 27–56 years; female:male, 1:1.1) with coronary heart disease and 103 healthy volunteers age, (28–63 years; female:male, 1:1.1) were recruited from Department of Cardiology, Zhujiang Hospital, Southern Medical University (Guangzhou, China) between January and May 2016 for analyzing serum levels of DCR-3. The patients were classified into four groups. Health group contained participants without angina. Primary group participants who only suffered angina during intense, rapid or prolonged physical activity or exercise. Moderate group were slightly limited in daily activities, as angina occurred following brisk walking, going upstairs, walking in cold air or after mood swings. Severe group were under great limitation and suffered angina after even mild exercise. The present study was approved by the Ethics Committee of Southern Medical University (Guangzhou, China). All patients provided written informed consent prior to experiments. Primary, moderate and severe coronary heart disease was diagnosed using 2014 ACC/AHA Recommendations for the management of NSTE ACS (22).
Cell culture
Cardiac fibroblasts were harvested from C57BL/6J mice as described previously (23). A total of 50 female C57BL/6J (age, 8 weeks; weight, 28–32 g) mice were purchased from OrientBio, Inc. (Seongnam, Korea). All mice were given free access to food and water, and housed at 23°C with 50% humidity and a 12-h artificial light/dark cycle. Briefly, mice were sacrificed under 20 mg/kg subcutaneous meperidine anesthesia (24). Harvested hearts were washed with PBS and heart tissues were digested with digestion buffer prepared with 0.05% (w/v) collagenase type II (Worthington Biochemical Corporation, Lakewood, NJ, USA) and 0.06% (w/v) pancreatin (Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) in PBS for 10 min at 37°C. The supernatant was collected and filtrated with a 70-µm filter and collected cells were centrifuged at 100 × g for 4 min at 25°C and resuspended in culture medium: RPMI-1640 medium (Sigma-Aldrich; Merck KGaA) with penicillin-streptomycin (Thermo Fisher Scientific, Inc., Waltham, MA, USA) at 37°C for 12 h. Following washing out digestion buffer with RPMI-1640 medium twice, cells were seeded on cell culture dishes and incubated for 1 h at 37°C. Following incubation, unattached cells were removed and attached cells were cultured with fresh culture medium supplemented with 10% fetal bovine serum (Sigma-Aldrich; Merck KGaA). Cells were cultured in a 5% CO2 incubator with a humidified atmosphere at 37°C.
RNA isolation and reverse transcription-quantitative polymerase chain reaction (RT-qPCR)
Total RNA was extracted from cardiac fibroblasts using TRIzol reagent (Life Technologies; Thermo Fisher Scientific, Inc.). Quality of extracted mRNA was identified by Spectroscopy Elemental Isotope Analysis using NanoDrop Lite (Thermo Fisher Scientific, Inc.). According to the manufacturer's descriptions, total RNA (1 µg) was reverse-transcribed to cDNA using the Transcriptor First Strand cDNA Synthesis kit (Invitrogen; Thermo Fisher Scientific, Inc.). The qPCR reaction was performed with a SYBR-Green detection system (Bio SYBR Green Master Mix; Takara Biotechnology Co., Ltd., Dalian, China). The following primers were synthesized by Invitrogen; Thermo Fisher Scientific, Inc.: DCR-3, forward 5′-CTCTTCCTCCCATGACAC-3′ and reverse 5′-CTGGAAAGCCACAAAGTC-3′; and β-actin, forward 5′-CGTGAAAAGATGACCCAGATCA-3′ and reverse 5′-CAGCCTGGATGGCTACGTACA-3′. PCR cycling conditions were performed at 95°C for 30 sec and 45 cycles of 95°C for 5 sec, 56.5°C for 10 sec and 72°C for 10 sec. Relative mRNA expression changes were calculated by the 2−ΔΔCq method (25). The results are expressed as a fold-change compared with the β-actin control.
Small interfering RNA (siRNA) transfection
siRNA to target PI3K (Si-PI3K) and scrambled siRNA (Si-vector) were designed and synthesized by Shanghai GenePharma Co., Ltd. (Shanghai, China). The sequences were as follows: Si-PI3K, forward 5′-CCAACAACAGCAUGAACAAdTdT-3′ and reverse 5′-UUGUUCAUGCUGUUGUUGGdTdT-3′; and Si-vector, forward 5′-AUGAACGUGAAUUGCUCAAdTdT-3′ and reverse 5′-UUGAGCAAUUCACGUUCAUdTdT-3′. Cardiac fibroblasts isolated from experimental mice were seeded in 24-well plates and were cultured to 80% confluence. Then 1.25 µl 25 µM Si-PI3K or Si-vector were transfected to cells using Lipofectamine™ RNAi MAX (Invitrogen; Thermo Fisher Scientific, Inc.) according to the manufacturer's instructions. Si-PI3K and Si-vector were from Shanghai GenePharma Co., Ltd. (Shanghai, China). The interval between transfection and subsequent experiments was 24 h.
Syntax score
Syntax score of patients with coronary heart disease were calculated according to a previous study (26). The DCR-3 expression and syntax score were analyzed in patients with coronary heart disease with healthy volunteers as controls.
ELISA
In the protein detection assay, serum levels of DCR-3 were measured using an ELISA kit (DY142; Bio-Rad Laboratories, Inc., Hercules, CA, USA). The operating steps were conducted according to the manufacturer's instructions. The final results were recorded at 450 nm on an ELISA plate reader.
Western blotting
Myocardial cells harvested from mice were lysed in radioimmunoprecipitation assay buffer containing a phosphatase inhibitor and the protease inhibitor cocktail (Pierce; Thermo Fisher Scientific, Inc.). Protein concentrations were determined via bicinchoninic protein assay (Pierce; Thermo Fisher Scientific, Inc.). Equal amounts of proteins (40 µg/lane) were loaded and separated by 12% SDS-PAGE. The proteins were transferred to polyvinylidene difluoride membranes (EMD Millipore, Billerica, MA, USA). The membranes were then blocked for 1 h at 37°C with 10% bovine serum albumin (cat. no. 10735108001; Roche Applied Science, Penzburg, Germany). The following rabbit anti-mouse antibodies were used to incubate protein for 12 h at 4°C: DCR-3 (1:1,000, cat. no. ab8405), cardiac troponin (cTn)T (1:1,000; cat. no. ab45932), cTn1 (1:1,000; cat. no. ab210798), IL-6 (1:1,000; cat. no. ab83053), C-reactive protein 1 (CRP-1) (1:1,000; cat. no. ab211631), albumin (1:1,000; cat. no. ab207327), intercellular adhesion molecule (ICAM)-1 (1:1,000; cat. no. ab119817), vascular cell adhesion molecule (VCAM)-1 (1:1,000; cat. no. ab134047), PI3K (1:1,000; cat. no. ab40776), AKT (1:1,000; cat. no. ab8805), phosphorylated (p)AKT (1:1,000; cat. no. ab81283) and β-actin (1:2,000; cat. no. ab8226; all Abcam, Cambridge, UK). Horseradish peroxidase (HRP)-conjugated goat anti-rabbit antibody (1:5,000; cat. no. HAF019; Bio-Rad Laboratories, Inc.) was used as a secondary antibody for 2 h at 37°C and protein was detected using a western blotting Luminol reagent (cat. no. 12015218001; Sigma-Aldrich; Merck KGaA) for enhanced chemiluminescence. Lanes were observed using light microscopy (magnification, ×4). The density of the bands was analyzed using Quantity One software, version 4.62 (Bio-Rad Laboratories, Inc.).
Immunofluorescence
Myocardial cells were fixed with formaldehyde solution (10%) for 2 h at 37°C and processed according to standard procedures. The sections are blocked with 5% bovine serum albumin (Sigma-Aldrich; Merck KGaA) at 37°C for 15 min. Cells were incubated with primary antibody against DCR-3 (1:1,000, cat. no. ab8405) for 12 h at 4°C. Cells were then incubated with HRP-conjugated anti-rabbit antibodies (1:200; cat. no. 71623, Bio-Rad Laboratories, Inc.) for 2 h at 37°C. Cells were washed and mounted with VectaShield mounting media with DAPI (Vector Laboratories, Inc., Burlingame, CA, USA) and kept in the dark at 4°C prior to microscopic analysis. All images (magnification, ×10) were captured with a confocal microscope (Fluoview1000; Olympus Corporation, Tokyo, Japan).
Animal experiments
A total of 50 female C57BL/6J (age, 8 weeks; weight, 28–32 g) mice were purchased from OrientBio, Inc. (Seongnam, Korea). All mice were given free access to food and water, and housed at 23°C with 50% humidity with a 12-h artificial light/dark cycle. A total of 45 mice were fed with a high-fat diet to establish coronary heart disease model according to a previous study (27); 5 healthy mice and 5 model mice were used to demonstrated the changes in DCR-3 level due to coronary heart disease. The remaining 40 model mice were divided into two groups (n=20/group) and received an intravenous injection of DCR-3 (10 mg/kg; Sigma-Aldrich; Merck KGaA) or PBS (Control). The treatments were repeated once daily for 10 days. Following 3 weeks, animals were sacrificed by cervical dislocation under anesthesia with 3% isoflurane (Sigma-Aldrich; Merck KGaA). The present study has been approved by the Committee for Experimental Animal Studies of Southern Medical University.
Histological assay
Cardiac slices isolated from mice with cardiac fibrosis were prepared and fixed in 4% paraformaldehyde for 30 min at 37°C. Cardiac slices were performed using an avidin-biotin-peroxidase technique. Paraffin-embedded tissue sections were prepared, heated at 65°C to melt the paraffin and dewaxed to water with xylene, ethyl alcohol (100, 95, 85 and 75%) and water successively. Epitope retrieval was performed for further analysis. The paraffin sections (4 µm) were incubated with hydrogen peroxide (3%) for 10 min, and were blocked with 5% bovine serum albumin (Sigma-Aldrich; Merck KGaA) for 10 min at 37°C. Finally, the sections were incubated with rabbit anti-mouse anti-DCR-3 (cat. no. ab8405; Abcam) diluted with PBS (1:1,000) or anti-major histocompatibility complex І antibodies (cat. no. ab185706; Abcam) diluted with PBS (1:1,000), for 12 h at 4°C following blocking with 5% bovine serum albumin (Sigma-Aldrich; Merck KGaA). Following rinsing, sections were incubated in the presence of a biotinylated horse anti-rabbit antibody (cat. no. a0545; 1:500; Chemicon; Merck KGaA) for 2 h at 37°C. Sections were washed and observed (magnification, ×10) using fluorescent video microscopy (BZ-9000; Keyence Corporation, Osaka, Japan).
Terminal deoxynucleotidyl-transferase-mediated dUTP nick end labeling (TUNEL) assay
The TUNEL assay was used for the analysis of apoptosis of myocardial tissue in experimental mice at 37°C following simvastatin treatment (10 mg/kg/day; Sigma-Aldrich; Merck KGaA) or an equivalent dose of PBS. Procedures were performed as previously described (27). Briefly, 4-µm tissue sections were fixed with 4% paraformaldehyde at 37°C followed by permeabilization with 0.1% Triton X-100 for 1 h at 37°C. Subsequently, tissue sections or myocardial cells (1×106) were stained with TUNEL reaction mixture (Sigma-Aldrich; Merck KGaA) at 37°C for 2 h. Tissue sections and myocardial cells were washed 3 times in TBS-Tween-20. TUNEL assays were conducted using a TUNEL fluorescence FITC kit (Roche Diagnostics, Indianapolis, IN, USA) according to the manufacturer's instructions. Samples were mounted with neutral gum. Images in 6 fields of view (magnification, ×400) were captured using a Zeiss LSM 510 confocal microscope (Zeiss AG, Oberkochen, Germany) at a wavelength of 488 nm.
Statistical analysis
Results are expressed as the mean + standard deviation. All data were analyzed using SPSS software (version 19.0; IBM Corp., Armonk, NY, USA). Comparisons between two groups and among multiple groups were conducted by Student's t-test and one-way analysis of variance followed by Tukey's post hoc test, respectively. P<0.05 was considered to indicate a statistically significant difference.
Results
DCR-3 expression levels in myocardial cells of patients with coronary heart disease
In the present study, the plasma concentration levels of DCR-3 in patients with coronary heart disease were analyzed. As presented in Fig. 1A, it was observed that plasma concentration levels of DCR-3 were downregulated in peripheral blood with the severity of the disease increasing, as determined by ELISA. In addition, it was also demonstrated that mRNA and protein expression in myocardial cells were decreased with increasing disease severity, as determined by RT-qPCR and western blotting, respectively (Fig. 1B and C). Furthermore, circulating DCR-3 levels were associated with the Syntax score and severity of coronary heart disease, which was demonstrated by the increase in SXscore along with more serious disease condition (Fig. 1D). Taken together, these results suggest DCR-3 expression levels can predict the events and severity of coronary heart disease.
DCR-3 has benefits for the treatment of mice with coronary heart disease by improvement of coronary lesions
To investigate the efficacy of DCR-3 for coronary heart disease, the therapeutic effects of DCR-3 were analyzed in mice with coronary heart disease. By analyzing the DCR-3 expression in myocardial cells, it was observed that DCR-3 expression in myocardial cells was analyzed. It was observed that DCR-3 expression levels were downregulated in myocardial cells isolated from mice with coronary heart disease (Fig. 2A). It was also observed that plasma concentration levels were downregulated in the coronary heart disease mice (Fig. 2B). In addition, DCR-3 treatment affected the lesions and inhibited the apoptosis of arterial vascular smooth muscle, as determined by histological analysis (Fig. 2C). Furthermore, results indicated that myocardial expression levels of injury-associated proteins cTnT and cTn І were upregulated in myocardium after treatment with DCR-3 (Fig. 2D). Taken together, these results suggest that DCR-3 is beneficial for the remission of coronary heart disease induced by TGF-β1 through improvement of coronary lesions.
DCR-3 inhibits expression levels of inflammatory factors in myocardial cells
A previous study indicated that inflammatory responses are associated with the progression of coronary heart disease (28). Therefore, the inflammatory factor expression was analyzed in myocardial cells and tissues. As presented in Fig. 3A-E, following treatment with DCR-3, IL-6, CRP-1, serum albumin, ICAM-1 and VCAM-1 were all decreased in myocardial cells in mice with coronary heart disease, which indicated that DCR-3 suppressed the expression levels of these inflammatory factors. Furthermore, DCR-3 could significantly ameliorate the myocardial inflammation of lymphocytes as well (Fig. 3F). In addition, the increased expression of DCR-3 was confirmed by western blotting in myocardial cells treated with DCR-3 (Fig. 3G). Collectively, these results suggest that DCR-3 can inhibit expression levels of inflammatory factors in myocardial cells in experimental mice with coronary heart disease.
DCR-3 improves inflammatory and apoptosis in myocardial cells through the PI3K/AKT signaling pathway
It has been demonstrated previously that pretreatment of inflammatory factors by activating the PI3K/AKT signaling pathway contributes to recovery of coronary heart disease-induced myocardial ischemia and injury (29). In the present study, the PI3K/AKT signaling pathway in myocardial cells was analyzed in experimental mice with coronary heart disease treated by DCR-3. As presented in Fig. 4A and B, DCR-3 in myocardial cells upregulated the expression levels of PI3K and p-AKT/AKT, compared with the control group. The results in Fig. 4C demonstrated that endogenous inhibition of PI3K expression by Si-PI3K was successfully achieved in myocardial cells. In vitro assays demonstrated that endogenous inhibition of PI3K expression by Si-PI3K suppressed AKT expression in myocardial cells (Fig. 4D). Furthermore, representative histological images revealed that Si-PI3K also markedly inhibited improvement of DCR-3-mediated apoptosis in myocardial cells, as determined by TUNEL and immunofluorescence (Fig. 4E and F). These findings suggest that PI3K/AKT signaling pathway involves the anti-inflammatory and anti-apoptosis potential of DCR-3 in myocardial cells in the progression of coronary heart disease induced by TGF-β1.
DCR-3 prolongs survival of mice with coronary heart disease
The efficacy for the survival of mice with coronary heart disease following treatment with DCR-3 was then determined. As presented in Fig. 5, DCR-3 significantly prolonged the survival of mice with coronary heart disease induced by TGF-β1 in a 90-day observation (n=20 in each group). Survival analysis suggests that DCR-3 may be an efficient agent for the treatment of coronary heart disease.
Discussion
DCR-3 acts as a soluble receptor of Fas ligand (FasL), TNF superfamily member 14 (LIGHT) and TNF superfamily member 15 that is reported highly expressed in cancer cells, but is expressed less in myocardial cells (30). A previous study has indicated that expression of DCR-3 is notable in chronic liver disease (31). However, the significance of expression of DCR-3 in myocardial cells in progression of coronary heart disease has not been reported in a previous study (32). The present study investigated the molecular mechanism of DCR-3 in myocardial cells in the progression of coronary heart disease. Although previous reports have reported that plasma DCR-3 levels may indicate the severity of coronary heart disease (33), the potential mechanism mediated by DCR-3 has not been reported in myocardial cells. The present results demonstrated that DCR-3 is downregulated in patients with coronary heart disease and increasing DCR-3 expression can prevent inflammation and apoptosis in myocardial cells induced by coronary heart disease. DCR-3 also mediate anti-inflammatory and anti-apoptosis potential through the PI3K/AKT signaling pathway in myocardial cells in the progression of coronary heart disease induced by TGF-β1. These findings may elucidate the function of DCR-3 in processes of apoptosis and inflammation for patients with coronary heart disease.
Inflammation is one of the most common symptoms of patients with acute coronary heart disease and the marked inflammation during acute coronary syndrome contributes to later depression in a subset of patients (34). Inflammatory response may be used as a prognostic indicator for borderline lesion coronary heart disease patients, who were treated with naoxintong (35). In addition, El-Mesallamy et al (36) have indicated that inflammation is one of factors in the imitation of coronary heart disease. Myocardial tissue has a higher level of inflammatory factors and lymphocytes. However, a stable DCR-3 analogue can reduce FasL-induced murine pulmonary inflammation (37). At the same time, DCR-3 may improve experimental autoimmune encephalomyelitis by directly counteracting local inflammation, suggesting that DCR-3 is the potential agent for treating human multiple sclerosis (38). All of these reports suggest that DCR-3 is associated with inflammatory responses in patients with cardiovascular disease. Additionally, although previous data demonstrated that slightly elevated cTnS levels may not be a sensitive prognostic marker for the Chinese population (39), a number of studies have indicated that serum cTnT levels are also regarded as an indicator of myocardial injury in ischemic and hemorrhagic stroke patients (40–42). The present results demonstrated that DCR-3 increased serum levels of cTnT and cTn1 in mice with coronary heart disease and that DCR-3 can inhibit inflammation responses and expression levels of inflammatory responses in myocardial cells in mice with coronary heart disease.
Apoptosis of myocardial cells is another crucial factor in initiating coronary heart disease (7). Considerable evidence has indicated that apoptosis has an important role in hepatocyte death in chronic liver disease (43). Research also demonstrated that apoptosis of leukocytes acts as a disease marker of neutrophil-endotheliocyte interaction in patients with coronary heart disease (14). Liu et al (15) have previously investigated the expression levels of inflammatory and apoptosis factors in coronary stent implantation in coronary heart disease patients, and demonstrated that downregulation of inflammatory and apoptosis factors is beneficial to the recovery of coronary heart disease. DCR-3, as a pleiotropic immunomodulator for enhancing angiogenesis, has been observed to downregulate cell apoptosis by binding with its receptor of FasL and LIGHT, and subsequently improved survival of many types of cell, such as human IPF fibroblasts (44) and mouse lymphocytes (45). Furthermore, DCR-3 also suppressed FasL-induced apoptosis via extracellular signal-regulated kinase 1/2 activation in various cancer cells (46–48). Furthermore, a number of studies have reported DCR-3 function in lymphocytes and dendritic cells (45,49). The present study indicated that DCR-3 could inhibit apoptosis of myocardial cells in mice with coronary heart disease, which contributed to protect myocardial cells against injury in the process of coronary heart disease. Notably, injury-associated protein levels of cTnT and cTn І were markedly upregulated in myocardium by the treatment of DCR-3. These results suggest that inhibition of apoptosis is the essential function of DCR-3 for the treatment of coronary heart disease. However, the present study did not use a healthy or sham control to identify the role of DCR-3 for the treatment of coronary heart disease, which is a limitation of this research.
The PI3K/AKT signaling pathway is involved in the progression of myocardial infarction in the process of coronary heart disease (50). Hu et al (51) presented the protective effect of proanthocyanidins on anoxia-reoxygenation injury of myocardial cells and results have indicated that the PI3K/AKT/glycogen synthase kinase-3β pathway mediates mitochondrial ATP-sensitive potassium channel to regulate oxidative stress. It has also been indicated that the PI3K/AKT pathway-regulated recruiting T cells could attenuate myocardial ischemia-reperfusion injury in a mice model (52). In the present study, by investigating the association between DCR-3 and the PI3K/AKT signal pathway in myocardial cells in mice with coronary heart disease, it was observed that DCR-3 mediated the improvement of myocardial ischemia via the PI3K/AKT signaling pathway. Notably, it also demonstrated that endogenous inhibition of PI3K expression by Si-PI3K abolished DCR-3-mediated anti-inflammation and anti-apoptosis in myocardial cells in mice with coronary heart disease.
In conclusion, the present study investigated the efficacy of DCR-3 in the progression of coronary heart disease and the molecular mechanism of DCR-3-mediated anti-inflammation and anti-apoptosis in myocardial cells in mice with coronary heart disease. Results have demonstrated that patients and mice with coronary heart disease have lower DCR-3 expression levels in peripheral blood. DCR-3 can inhibit inflammatory factor expression and apoptosis of myocardial cells and that the PI3K/AKT signaling pathway is associated with anti-inflammation and anti-apoptosis in the process of coronary heart disease. These results confirm that DCR-3 treatment may be beneficial for the recovery of coronary heart disease by anti-inflammation and anti-apoptosis in myocardial cells through the PI3K/AKT signaling pathway, suggesting that DCR-3 may be able to predict the extent and severity of cardiovascular diseases.
Acknowledgements
Not applicable.
Funding
The present study was supported by grants from Fujian Provincial Health and Family Planning Young and Middle-aged Key Members Talent Training Project (grant no. 2017-ZQN-9), Fujian Provincial Natural Science Funding Project (grant no. 2018J01244), The Key Clinical Specialty Discipline Construction Program of Fujian (Program of Vasculocardiology) and Fujian Provincial Youth Scientific Research Project of Health Department (grant no. 2012-2-10).
Availability of data and materials
The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.
Authors' contributions
XJC, RHW and WC designed the experiments. XJC and RHW performed the experiments and analysis. RHW and LL helped analyze the data. RHW, WC and ZLL wrote the manuscript. All authors read and approved the final manuscript.
Ethics approval and consent to participate
The present study was approved by the Ethics Committee and the Committee for Experimental Animal Studies of Southern Medical University (Guangzhou, China). All patients provided written informed consent.
Patient consent for publication
All patients provided written informed consent.
Competing interests
The authors declare that they have no competing interests.
References
Tully PJ and Baumeister H: Collaborative care for comorbid depression and coronary heart disease: A systematic review and meta-analysis of randomised controlled trials. BMJ Open. 5:e0091282015. View Article : Google Scholar : PubMed/NCBI | |
Heikkilä K, Koskinen OA, Agarwal A, Tikkinen KA, Mäki M and Kaukinen K: Associations of coeliac disease with coronary heart disease and cerebrovascular disease: A systematic review and meta-analysis. Nutr Metab Cardiovasc Dis. 25:816–831. 2015. View Article : Google Scholar : PubMed/NCBI | |
Mayor S: Reduced blood pressure and cholesterol are main factors in fall in deaths from coronary heart disease. BMJ. 350:h4152015. View Article : Google Scholar : PubMed/NCBI | |
Niermann C, Gorressen S, Klier M, Gowert NS, Billuart P, Kelm M, Merx MW and Elvers M: Oligophrenin1 protects mice against myocardial ischemia and reperfusion injury by modulating inflammation and myocardial apoptosis. Cell Signal. 28:967–978. 2016. View Article : Google Scholar : PubMed/NCBI | |
Guo CX, Jiang X, Zeng XJ, Wang HX, Li HH, Du FH and Chen BX: Soluble receptor for advanced glycation end-products protects against ischemia/reperfusion-induced myocardial apoptosis via regulating the ubiquitin proteasome system. Free Radic Biol Med. 94:17–26. 2016. View Article : Google Scholar : PubMed/NCBI | |
O'Neal WT, Soliman EZ, Howard G, Howard VJ, Safford MM, Cushman M and Zakai NA: Inflammation and hemostasis in atrial fibrillation and coronary heart disease: The REasons for geographic and racial differences in stroke study. Atherosclerosis. 243:192–197. 2015. View Article : Google Scholar : PubMed/NCBI | |
Geng YJ: Molecular mechanisms for cardiovascular stem cell apoptosis and growth in the hearts with atherosclerotic coronary disease and ischemic heart failure. Ann N Y Acad Sci. 1010:687–697. 2003. View Article : Google Scholar : PubMed/NCBI | |
Fang Y, Chen S, Liu Z, Ai W, He X, Wang L, Xie P, Jiang B and Fang H: Endothelial stem cells attenuate cardiac apoptosis via downregulating cardiac microRNA-146a in a rat model of coronary heart disease. Exp Ther Med. 16:4246–4252. 2018.PubMed/NCBI | |
Xu L, Jiang X, Wei F and Zhu H: Leonurine protects cardiac function following acute myocardial infarction through anti-apoptosis by the PI3K/AKT/GSK3β signaling pathway. Mol Med Rep. 18:1582–1590. 2018.PubMed/NCBI | |
Wang Z, Zhang J, Ren T and Dong Z: Targeted metabolomic profiling of cardioprotective effect of Ginkgo biloba L.extract on myocardial ischemia in rats. Phytomedicine. international journal of phytotherapy and phytopharmacology. 23:621–631. 2016. View Article : Google Scholar | |
Wang L, Niu X, Hu J, Xing H, Sun M, Wang J, Jian Q and Yang H: After myocardial ischemia-reperfusion, mir-29a, and let7 could affect apoptosis through regulating IGF-1. BioMed Research International. 2015:2454122015. View Article : Google Scholar : PubMed/NCBI | |
Wakiyama H, Cowan DB, Toyoda Y, Federman M, Levitsky S and McCully JD: Selective opening of mitochondrial ATP-sensitive potassium channels during surgically induced myocardial ischemia decreases necrosis and apoptosis. Eur J Cardiothorac Surg. 21:424–433. 2002. View Article : Google Scholar : PubMed/NCBI | |
Elsasser A, Suzuki K, Lorenz-Meyer S, Bode C and Schaper J: The role of apoptosis in myocardial ischemia: A critical appraisal. Basic Res Cardiol. 96:219–226. 2001. View Article : Google Scholar : PubMed/NCBI | |
Salmina AB, Shul'man VA, Nikulina SY, Trufanova LV, Fursov AA, But'yanov PA, Kuskaev AP, Bol'shakova EV and Kotlovskii MY: Apoptosis of leukocytes as a marker of neutrophil-endotheliocyte interaction in coronary heart disease. Bull Exp Biol Med. 144:39–41. 2007.(In English, Russian). View Article : Google Scholar : PubMed/NCBI | |
Liu LL, Lin LR, Lu CX, Fu JG, Chao PL, Jin HW, Zhang ZY and Yang TC: Expression of inflammatory and apoptosis factors following coronary stent implantation in coronary heart disease patients. Int Immunopharmacol. 11:1850–1854. 2011. View Article : Google Scholar : PubMed/NCBI | |
Azambuja MI: Inflammation as the cause of coronary heart disease. Lancet Infect Dis. 10:142–143. 2010. View Article : Google Scholar : PubMed/NCBI | |
Wu T, Sun QF, Yang PS, Feng W and Liu XL: Levels of inflammation cytokines in patients with coronary heart disease and periodontal disease. Zhonghua Kou Qiang Yi Xue Za Zhi. 45:265–268. 2010.(In Chinese). PubMed/NCBI | |
Yan Y, Song D, Liu L, Meng X, Qi C and Wang J: The relationship of plasma decoy receptor 3 and coronary collateral circulation in patients with coronary artery disease. Life Sci. 189:84–88. 2017. View Article : Google Scholar : PubMed/NCBI | |
Marriott HM, Daigneault M, Thompson AA, Walmsley SR, Gill SK, Witcher DR, Wroblewski VJ, Hellewell PG, Whyte MK and Dockrell DH: A decoy receptor 3 analogue reduces localised defects in phagocyte function in pneumococcal pneumonia. Thorax. 67:985–992. 2012. View Article : Google Scholar : PubMed/NCBI | |
Wu SF, Liu TM, Lin YC, Sytwu HK, Juan HF, Chen ST, Shen KL, his SC and Hsieh SL: Immunomodulatory effect of decoy receptor 3 on the differentiation and function of bone marrow-derived dendritic cells in nonobese diabetic mice: From regulatory mechanism to clinical implication. J Leukoc Biol. 75:293–306. 2004. View Article : Google Scholar : PubMed/NCBI | |
Jiang M, Lin X, He R, Lin X, Liang L, Tang R, Xiong D, Wei K, Dang Y, Feng Z and Chen G: Decoy receptor 3 (DcR3) as a biomarker of tumor deterioration in female reproductive cancers: A meta-analysis. Med Sci Monit. 22:1850–1857. 2016. View Article : Google Scholar : PubMed/NCBI | |
Dietz R and Rauch B: German Society of Cardiology-Heart Circulation Research; German Society for Prevention Rehabilitation of Cardiac Diseases; German Society for Thoracic CardiovascularSurgery: Guidelines for diagnosis and treatment of chronic coronary heart disease. Issued by the executive committee of the German Society of Cardiology - Heart Circulation Research in cooperation with the German Society for Prevention and Rehabilitation of Cardiac Diseases and the German Society for Thoracic and Cardiovascular Surgery. Z Kardiol. 92:501–521. 2003.(In German). PubMed/NCBI | |
Golden HB, Gollapudi D, Gerilechaogetu F, Li J, Cristales RJ, Peng X and Dostal DE: Isolation of cardiac myocytes and fibroblasts from neonatal rat pups. Methods Mol Biol. 843:205–214. 2012. View Article : Google Scholar : PubMed/NCBI | |
Lee I, Chalon J, Ramanathan S, Gross S and Turndorf H: Analgesic properties of meperidine, amitriptyline and phenelzine in mice. Can Anaesth Soc J. 30:501–505. 1983. View Article : Google Scholar : PubMed/NCBI | |
Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI | |
Börekçi A, Gür M, Şeker T, Baykan AO, Özaltun B, Karakoyun S, Karakurt A, Türkoğlu C, Makça I and Çaylı M: Coronary collateral circulation in patients with chronic coronary total occlusion; its relationship with cardiac risk markers and SYNTAX score. Perfusion. 30:457–464. 2015. View Article : Google Scholar : PubMed/NCBI | |
Wu ZW, Liu YF, Wang S and Li B: miRNA-146a induces vascular smooth muscle cell apoptosis in a rat model of coronary heart disease via NF-κB pathway. Genet Mol Res. 14:18703–18712. 2015. View Article : Google Scholar : PubMed/NCBI | |
Werba JP, Veglia F, Amato M, Baldassarre D, Massironi P, Meroni PL, Riboldi P, Tremoli E and Camera M: Patients with a history of stable or unstable coronary heart disease have different acute phase responses to an inflammatory stimulus. Atherosclerosis. 196:835–840. 2008. View Article : Google Scholar : PubMed/NCBI | |
Rong R and Xijun X: Erythropoietin pretreatment suppresses inflammation by activating the PI3K/Akt signaling pathway in myocardial ischemia-reperfusion injury. Exp Ther Med. 10:413–418. 2015. View Article : Google Scholar : PubMed/NCBI | |
Funke B, Autschbach F, Kim S, Lasitschka F, Strauch U, Rogler G, Gdynia G, Li L, Gretz N, Macher-Goeppinger S, et al: Functional characterisation of decoy receptor 3 in Crohn's disease. Gut. 58:483–491. 2009. View Article : Google Scholar : PubMed/NCBI | |
Kim S, Kotoula V, Hytiroglou P, Zardavas D and Zhang L: Significance of increased expression of decoy receptor 3 in chronic liver disease. Dig Liver Dis. 41:591–598. 2009. View Article : Google Scholar : PubMed/NCBI | |
Chang TY, Hsu CY, Huang PH, Chiang CH, Leu HB, Huang CC, Chen JW and Lin SJ: Usefulness of circulating decoy receptor 3 in predicting coronary artery disease severity and future major adverse cardiovascular events in patients with multivessel coronary artery disease. Am J Cardiol. 116:1028–1033. 2015. View Article : Google Scholar : PubMed/NCBI | |
Kim S, McAuliffe WJ, Zaritskaya LS, Moore PA, Zhang L and Nardelli B: Selective induction of tumor necrosis receptor factor 6/decoy receptor 3 release by bacterial antigens in human monocytes and myeloid dendritic cells. Infect Immun. 72:89–93. 2004. View Article : Google Scholar : PubMed/NCBI | |
Steptoe A, Wikman A, Molloy GJ, Messerli-Bürgy N and Kaski JC: Inflammation and symptoms of depression and anxiety in patients with acute coronary heart disease. Brain Behav Immun. 31:183–188. 2013. View Article : Google Scholar : PubMed/NCBI | |
Li SR, Wang TH and Zhang BJ: Effects of naoxintong capsule on the inflammation and prognosis in borderline lesion coronary heart disease patients. Zhongguo Zhong Xi Yi Jie He Za Zhi. 32:607–611. 2012.(In Chinese). PubMed/NCBI | |
El-Mesallamy HO, Hamdy NM, Salman TM and Ibrahim SM: Adiponectin and sE-selectin concentrations in relation to inflammation in obese type 2 diabetic patients with coronary heart disease. Angiology. 63:96–102. 2012. View Article : Google Scholar : PubMed/NCBI | |
Wortinger MA, Foley JW, Larocque P, Witcher DR, Lahn M, Jakubowski JA, Glasebrook A and Song HY: Fas ligand-induced murine pulmonary inflammation is reduced by a stable decoy receptor 3 analogue. Immunology. 110:225–233. 2003. View Article : Google Scholar : PubMed/NCBI | |
Chen SJ, Wang YL, Kao JH, Wu SF, Lo WT, Wu CC, Tao PL, Wang CC, Chang DM and Sytwu HK: Decoy receptor 3 ameliorates experimental autoimmune encephalomyelitis by directly counteracting local inflammation and downregulating Th17 cells. Mol Immunol. 47:567–574. 2009. View Article : Google Scholar : PubMed/NCBI | |
Zhang M, He H, Wang ZM, Xu Z, Zhou N, Tao Z, Chen B, Li C, Zhu T, Yang D, et al: Diagnostic and prognostic value of minor elevated cardiac troponin levels for percutaneous coronary intervention-related myocardial injury: A prospective, single-center and double-blind study. J Biomed Res. 28:98–107. 2014.PubMed/NCBI | |
Apak I, Iltumur K, Tamam Y and Kaya N: Serum cardiac troponin T levels as an indicator of myocardial injury in ischemic and hemorrhagic stroke patients. Tohoku J Exp Med. 205:93–101. 2005. View Article : Google Scholar : PubMed/NCBI | |
Nesher N, Zisman E, Wolf T, Sharony R, Bolotin G, David M, Uretzky G and Pizov R: Strict thermoregulation attenuates myocardial injury during coronary artery bypass graft surgery as reflected by reduced levels of cardiac-specific troponin I. Anesth Analg. 96:328–335. 2003. View Article : Google Scholar : PubMed/NCBI | |
Boriani G, Biffi M, Cervi V, Bronzetti G, Magagnoli G, Zannoli R and Branzi A: Evaluation of myocardial injury following repeated internal atrial shocks by monitoring serum cardiac troponin I levels. Chest. 118:342–347. 2000. View Article : Google Scholar : PubMed/NCBI | |
Wu ZW, Liu YF, Wang S and Li B: Corrigendum miRNA-146a induces vascular smooth muscle cell apoptosis in a rat model of coronary heart disease via NF-kappaB pathway - Genet. Mol. Res. 14(4): 18703–18712, Genet Mol Res 15. 2016. View Article : Google Scholar | |
Im J, Kim K, Hergert P and Nho RS: Idiopathic pulmonary fibrosis fibroblasts become resistant to Fas ligand-dependent apoptosis via the alteration of decoy receptor 3. J Pathol. 240:25–37. 2016. View Article : Google Scholar : PubMed/NCBI | |
Liang D, Hou Y, Lou X and Chen H: Decoy receptor 3 improves survival in experimental sepsis by suppressing the inflammatory response and lymphocyte apoptosis. PLoS One. 10:e01316802015. View Article : Google Scholar : PubMed/NCBI | |
Zhang Y, Li D, Zhao X, Song S, Zhang L, Zhu D, Wang Z, Chen X and Zhou J: Decoy receptor 3 suppresses FasL-induced apoptosis via ERK1/2 activation in pancreatic cancer cells. Biochem Biophys Res Commun. 463:1144–1151. 2015. View Article : Google Scholar : PubMed/NCBI | |
Koksal IT, Sanlioglu AD, Karacay B, Griffith TS and Sanlioglu S: Tumor necrosis factor-related apoptosis inducing ligand-R4 decoy receptor expression is correlated with high Gleason scores, prostate-specific antigen recurrence, and decreased survival in patients with prostate carcinoma. Urol Oncol. 26:158–165. 2008. View Article : Google Scholar : PubMed/NCBI | |
Zhou J, Song S, He S, Wang Z, Zhang B, Li D and Zhu D: Silencing of decoy receptor 3 (DcR3) expression by siRNA in pancreatic carcinoma cells induces Fas ligand-mediated apoptosis in vitro and in vivo. Int J Mol Med. 32:653–660. 2013. View Article : Google Scholar : PubMed/NCBI | |
You RI, Chang YC, Chen PM, Wang WS, Hsu TL, Yang CY, Lee CT and Hsieh SL: Apoptosis of dendritic cells induced by decoy receptor 3 (DcR3). Blood. 111:1480–1488. 2008. View Article : Google Scholar : PubMed/NCBI | |
Zhang Z, Li S, Cui M, Gao X, Sun D, Qin X, Narsinh K, Li C, Jia H, Li C, et al: Rosuvastatin enhances the therapeutic efficacy of adipose-derived mesenchymal stem cells for myocardial infarction via PI3K/Akt and MEK/ERK pathways. Basic Res Cardiol. 108:3332013. View Article : Google Scholar : PubMed/NCBI | |
Hu Y, Li L, Yin W, Shen L, You B and Gao H: Protective effect of proanthocyanidins on anoxia-reoxygenation injury of myocardial cells mediated by the PI3K/Akt/GSK-3beta pathway and mitochondrial ATP-sensitive potassium channel. Mol Med Rep. 10:2051–2058. 2014. View Article : Google Scholar : PubMed/NCBI | |
Fang J, Hu F, Ke D, Yan Y, Liao Z, Yuan X, Wu L, Jiang Q and Chen L: N,N-dimethylsphingosine attenuates myocardial ischemia-reperfusion injury by recruiting regulatory T cells through PI3K/Akt pathway in mice. Basic Res Cardiol. 111:322016. View Article : Google Scholar : PubMed/NCBI |