Discovery of novel cholesteryl ester transfer protein (CETP) inhibitors by a multi-stage virtual screening.

Cholesteryl ester transfer protein (CETP) is a promising therapeutic target for cardiovascular diseases. It effectively lowers the low-density lipoprotein cholesterol levels and increases the high-density lipoprotein cholesterol levels in the human plasma. This study identified novel and highly potent CETP inhibitors using virtual screening techniques. Molecular docking and molecular dynamics (MD) simulations revealed the binding patterns of these inhibitors, with the top 50 compounds selected according to their predicted binding affinity. Protein-ligand interaction analyses were performed, leading to the selection of 26 compounds for further evaluation. A CETP inhibition assay confirmed the inhibitory activities of the selected compounds. The results of the MD simulations revealed the structural stability of the protein-ligand complexes, with the binding site remaining significantly unchanged, indicating that the five compounds (AK-968/40709303, AG-690/11820117, AO-081/41378586, AK-968/12713193, and AN-465/14952302) identified have the potential as active CETP inhibitors and are promising leads for drug development.

Attenuation of estrogen and its receptors in the post-menopausal stage exacerbates dyslipidemia and leads to cognitive impairment.

Cognitive dysfunction increases as menopause progresses. We previously found that estrogen receptors (ERs) contribute to dyslipidemia, but the specific relationship between ERs, dyslipidemia and cognitive dysfunction remains poorly understood. In the present study, we analyzed sequencing data from female hippocampus and normal breast aspirate samples from normal and Alzheimer's disease (AD) women, and the results suggest that abnormal ERs signaling is associated with dyslipidemia and cognitive dysfunction. We replicated a mouse model of dyslipidemia and postmenopausal status in LDLR-/- mice and treated them with ß-estradiol or simvastatin, and found that ovariectomy in LDLR-/- mice led to an exacerbation of dyslipidemia and increased hippocampal apoptosis and cognitive impairment, which were associated with reduced estradiol levels and ERα, ERß and GPER expression. In vitro, a lipid overload model of SH-SY-5Y cells was established and treated with inhibitors of ERs. ß-estradiol or simvastatin effectively attenuated dyslipidemia-induced neuronal apoptosis via upregulation of ERs, whereas ERα, ERß and GPER inhibitors together abolished the protective effect of simvastatin on lipid-induced neuronal apoptosis. We conclude that decreased estrogen and its receptor function in the postmenopausal stage promote neuronal damage and cognitive impairment by exacerbating dyslipidemia, and that estrogen supplementation or lipid lowering is an effective way to ameliorate hippocampal damage and cognitive dysfunction via upregulation of ERs.

Effects of Compound Danshen Dripping Pills on Ventricular Remodeling and Cardiac Function after Acute Anterior Wall ST-Segment Elevation Myocardial Infarction (CODE-AAMI): Protocol for a Randomized Placebo-Controlled Trial.

BACKGROUND: Ventricular remodeling after acute anterior wall ST-segment elevation myocardial infarction (AAMI) is an important factor in occurrence of heart failure which additionally results in poor prognosis. Therefore, the treatment of ventricular remodeling needs to be further optimized. Compound Danshen Dripping Pills (CDDP), a traditional Chinese medicine, exerts a protective effect on microcirculatory disturbance caused by ischemia-reperfusion injury and attenuates ventricular remodeling after myocardial infarction. OBJECTIVE: This study is designed to evaluate the efficacy and safety of CDDP in improving ventricular remodeling and cardiac function after AAMI on a larger scale. METHODS: This study is a multi-center, randomized, double-blind, placebo-controlled, parallel-group clinical trial. The total of 268 patients with AAMI after primary percutaneous coronary intervention (pPCI) will be randomly assigned 1:1 to the CDDP group (n=134) and control group (n=134) with a follow-up of 48 weeks. Both groups will be treated with standard therapy of ST-segment elevation myocardial infarction (STEMI), with the CDDP group administrating 20 tablets of CDDP before pPCI and 10 tablets 3 times daily after pPCI, and the control group treated with a placebo simultaneously. The primary endpoint is 48-week echocardiographic outcomes including left ventricular ejection fraction (LVEF), left ventricular end-diastolic volume index (LVEDVI), and left ventricular end-systolic volume index (LVESVI). The secondary endpoint includes the change in N terminal pro-B-type natriuretic peptide (NT-proBNP) level, arrhythmias, and cardiovascular events (death, cardiac arrest, or cardiopulmonary resuscitation, rehospitalization due to heart failure or angina pectoris, deterioration of cardiac function, and stroke). Investigators and patients are both blinded to the allocated treatment. DISCUSSION: This prospective study will investigate the efficacy and safety of CDDP in improving ventricular remodeling and cardiac function in patients undergoing pPCI for a first AAMI. Patients in the CDDP group will be compared with those in the control group. If certified to be effective, CDDP treatment in AAMI will probably be advised on a larger scale. (Trial registration No. NCT05000411).

Relationship Between Breast Cancer Risk and Polymorphisms in CLOCK Gene: A Systematic Review and Meta-Analysis.

Previous studies found that the circadian clock gene participated in the genesis and development of breast cancer. However, research findings on the relationship between polymorphisms in the CLOCK gene and breast cancer risk were inconsistent. This study performed a meta-analysis of the association between CLOCK gene polymorphisms and breast cancer risk. PubMed, Cochrane Library, and Embase databases were electronically searched to collect studies on the association between CLOCK gene polymorphisms and breast cancer risk from inception to February 14, 2022. The quality of the included literature was assessed using the Newcastle-Ottawa Scale. For statistical analysis, odds ratio (OR) and 95% confidence intervals (CIs) were calculated using STATA 14.0. In addition, publication bias was performed by the funnel diagram and the Harbord's regression test. And sensitivity analysis was assessed by the trim and fill method. A total of 6 eligible studies, including 10,164 subjects (5488 breast cancer cases and 4676 controls), were screened in this meta-analysis. Though we did not find a significant association between the polymorphisms in the overall CLOCK gene with breast cancer risk [OR (95%CI) = 0.98 (0.96, 1.01), P = 0.148], we found that compared with T/T types of rs3749474 in CLOCK, T/C and C/C types of rs3749474 were associated with lower risk of breast cancer [OR (95%CI) = 0.93 (0.88, 0.98), P = 0.003]. The sensitivity analysis confirmed the robustness of the results. The funnel plot showed no significant publication bias. Polymorphisms in the CLOCK gene might be associated with breast cancer risk. More studies are needed to confirm the conclusion.

Activation of estrogen receptor α inhibits TLR4 signaling in macrophages and alleviates the instability of atherosclerotic plaques in the postmenopausal stage.

Acute cardiovascular events increase significantly in postmenopausal women. The relationship between estrogen receptor (ER) and plaque stability in the postmenopausal stage remains to be elucidated. We aimed to explore whether ERα activation improves plaque instability in the postmenopausal stage. Here, we report that postmenopausal women showed increased macrophage activation and plaque instability with increased MCP-1, MMP9, TLR4, MYD88 and NF-κB p65 and decreased ERα and TIMP1 expression in the vascular endothelium. Moreover, ovariectomy in LDLR-/- mice resulted in a significant increase in plaque area and necrotic core area, as well as a significant decrease in collagen content and an increase in macrophage accumulation in the artery. Ovariectomy also reduced serum estrogen levels and ERα expression and upregulated TLR4 and MMP9 expression in arteries in LDLR-/- mice. Estrogen or phytoestrogen therapy upregulated the expression level of ERα in ovariectomized mice and increased plaque stability by inhibiting macrophage accumulation and TLR4 signaling. In vitro, LPS incubation of RAW264.7 cells resulted in a significant decrease in ERα and TIMP1 expression and an increase in TLR4 activation, and estrogen or phytoestrogen treatment increased ERα and TIMP1 expression and inhibited TLR4 activation and MMP9 expression in LPS-treated RAW264.7 cells. Compared to control siRNA transfected RAW264.7 cells, TLR4 siRNA promoted TIMP1 expression in RAW264.7 cells with LPS incubation, but did not affect ERα expression in RAW264.7 cells with or without LPS treatment. The ERα inhibitor MPP abolished the regulatory effect of estrogen or phytoestrogen on LPS-induced RAW264.7 cells. In conclusion, the present study demonstrates that decreased ERα expression promotes macrophage infiltration and plaque instability in the postmenopausal stage, and activation of ERα in the postmenopausal stage alleviates atherosclerotic plaque instability by inhibiting TLR4 signaling and macrophage-related inflammation.

The effect of telemedicine on secondary prevention of atherosclerotic cardiovascular disease: A systematic review and meta-analysis.

Aim: The purpose of this systematic review was to evaluate the efficiency of telemedicine on the secondary level of prevention of patients with arteriosclerotic cardiovascular disease (ASCVD), provide evidence for the application of telemedicine in secondary prevention and promote the development of telemedicine in secondary prevention. Methods: A computer-based search was conducted in MEDLINE, Embase, Pubmed, EBSCO, CINAHL, the Cochrane Library, and Web of Science. Randomized controlled trials regarding the effect of telemedicine on secondary prevention of ASCVD were included from inception to May, 2022. Meta-analysis was used to compare the results of the included studies by RevMan5.4 software. The Cochrane Collaboration bias risk tool was used to perform risk of bias assessment in this study. Outcomes included risk factors, physical activity and exercise, muscle function, exercise compliance, medication adherence, healthy diet, depression and anxiety, self-efficacy, knowledge score, economy, and safety endpoints. Subgroup analysis was carried out for different main intervention measures included in the literature. Results: A total of 32 randomized clinical studies (n = 10 997 participants) were included in the meta-analysis. Compared with usual secondary prevention (USP) group, participants in telemedicine of secondary prevention (TOSP) group showed significant improvement in some risk factors including BMI (MD -0.87, p = 0.002), SBP (MD -4.09, p = 0.007) and DBP (MD -2.91, p = 0.0002) when they use the telephone as the intervention. In physical activity and exercise, Patients in TOSP showed an improvement in VO2 Peak (mL⋅kg-1⋅min-1) (OR 1.58, p = 0.02), 6MWT (MD 21.41, p = 0.001), GSLTPA score (MD 2.89, p = 0.005). Effects on medication adherence, exercise compliance, muscle function, healthy diet, economy and self-efficacy were synthesized narratively. Patients in TOSP did not show a reduction in knowledge score, depression, anxiety and safety endpoints. Conclusion: There is a net benefit of secondary prevention supported by telemedicine (especially when using the telephone as an intervention) in patients with ASCVD in the terms of some risk factors, physical activity and exercise. There are still controversies in the improvement of medication adherence, exercise compliance, muscle function, healthy diet, knowledge score, self-efficacy and economy via telemedicine, which is worth exploring. Larger samples size and longer-term follow-ups are needed in future studies. Systematic review registration: [https://www.crd.york.ac.uk/PROSPERO/display_record.php?RecordID=330478], identifier [CRD42022330478].

Effects of Shenfu Qiangxin Drink on H2O2-induced oxidative stress, inflammation and apoptosis in neonatal rat cardiomyocytes and possible underlying mechanisms.

The aim of the present study was to investigate the effects of Shenfu Qiangxin Drink (SFQXD) on acute myocardial infarction (AMI) and identify the possible underlying mechanisms. Levels of reactive oxygen species (ROS) and inflammatory factors, including interleukin (IL)-6, IL-1ß and tumor necrosis factor-α (TNF-α) in the blood samples of patients with AMI were measured using commercially available kits by visible spectrophotometry after SFQXD administration. The contents of phosphorylated (p-) forkhead box O3a (FOXO3a) was examined using an ELISA kit. In addition, a hydrogen peroxide (H2O2)-induced myocardial injury model was established in vitro using neonatal rat cardiomyocytes. Following treatment with SFQXD, the levels of intracellular ROS, cell apoptosis, oxidative stress- and inflammation-related markers were measured using commercially available kits by visible spectrophotometry. Additionally, western blot analysis was used to measure the expression of sirtuin-4 (SIRT4), p-FOXO3a, acetylated FOXO3a (ace-FOXO3a) and apoptosis-related genes (Bcl-2, Bax, BIM and cleaved caspase-3). Subsequently, to investigate the possible underlying regulatory mechanisms, SIRT4 expression was silenced by transfection with small hairpin RNA against SIRT4, following which changes in the extent of oxidative stress, inflammation and apoptosis were assessed. The levels of ROS and interleukin (IL)-1ß were found to be significantly reduced, whilst FOXO3a phosphorylation was markedly increased following administration with SFQXD. In vitro, SFQXD dose-dependently inhibited H2O2-induced oxidative stress, inflammation and apoptosis in neonatal rat cardiomyocytes. In addition, FOXO3a phosphorylation was markedly upregulated whilst FOXO3a acetylation was downregulated following treatment of H2O2-induced primary neonatal cardiomyocytes with SFQXD. SIRT4 knockdown also markedly reversed the effects of SFQXD on oxidative stress, inflammation and apoptosis in neonatal rat cardiomyocytes. In conclusion, these findings demonstrated that SFQXD may alleviate oxidative stress-induced myocardial injury by potentially regulating SIRT4/FOXO3a signaling, suggesting that SFQXD may be of clinical value for the treatment of AMI.

Arnebiae Radix prevents atrial fibrillation in rats by ameliorating atrial remodeling and cardiac function.

ETHNOPHARMACOLOGICAL RELEVANCE: Arnebiae Radix, a common herbal medicine in China, is often utilized to treat blood-heat syndrome and has been reported to exert an effect on the heart. AIM OF THE STUDY: The combination of acetylcholine (Ach) and CaCl2 has been widely used to induce atrial fibrillation (AF) in animals. However, whether Arnebiae Radix displays any preventive action on Ach-CaCl2 induced AF in rats remains uncertain. In our study, we attempted to investigate the protective effects of Arnebiae Radix on Ach-CaCl2 induced AF compared to amiodarone, which was employed as the positive control. MATERIALS AND METHODS: To establish the AF model, SD rats were treated with a mixture of 0.1 mL/100 g Ach-CaCl2 (60 µg/mL Ach and 10 mg/mL CaCl2) by tail vein injection for 7 days. Rats were also given a gavage of Arnebiae Radix (0.18 g/mL) one week before or concurrently with the establishment of the AF model. At the end of the experimental period, the induction, duration and timing of AF were monitored using electrocardiogram recordings. Left atrial tissues were stained to observe the level of fibrosis. Electrophysiological measurements were used to examine atrial size and function. RESULTS: In Ach-CaCl2-induced AF rats, Arnebiae Radix decreased AF induction, duration and susceptibility to AF. In addition, Arnebiae Radix significantly reduced atrial fibrosis and inhibited atrial enlargement induced by Ach-CaCl2. Moreover, there was an apparent improvement in cardiac function in the Arnebiae Radix-treated group. CONCLUSIONS: Our findings indicate that Arnebiae Radix treatment can attenuate Ach-CaCl2-induced atrial injury and serve as an effective therapeutic strategy for the treatment of AF in the future.

Hydroxytyrosol and olive leaf extract exert cardioprotective effects by inhibiting GRP78 and CHOP expression.

Myocardial infarction triggers massive biochemical changes, even cardiac cell death. Endoplasmic reticulum stress is involved in the pathology of myocardial infarction-mediated apoptosis. In the present study, myocardial cell line H9c2 cells were treated with cobalt chloride (CoCl2) to induce hypoxia. Isoproterenol was used for two successive days to induce myocardial infarction in SD rats. The cardioprotective effect of olive leaf extract (OLE) and its main constituent hydroxytyrosol and the underlying mechanisms were evaluated. The results showed that hydroxytyrosol markedly protected H9c2 cells against CoCl2-induced apoptosis. Hydroxytyrosol could reduce the mRNA and protein expression of GRP78 and CHOP induced by CoCl2 in vitro. In vivo, the decreased ejection fraction and fractional shortening, increased heart weight/body ratio, the formation of infarction, disordered cardiac muscle fibers and infiltration of inflammatory cells induced by isoproterenol could be significantly ameliorated by pretreatment with OLE for a month. Similarly, OLE could also reverse the increase of GRP78 and CHOP expression induced by isoproterenol. Therefore, OLE and hydroxytyrosol exert a cardioprotective effect through endoplasmic reticulum stress, which could be a new target for the prevention and treatment of cardiovascular diseases.

Protective Effects of Olive Leaf Extract on Acrolein-Exacerbated Myocardial Infarction via an Endoplasmic Reticulum Stress Pathway.

Many studies reported that air pollution particulate matter (PM) exposure was associated with myocardial infarction (MI). Acrolein representing the unsaturated aldehydes, the main component of PM, derives from the incomplete combustion of wood, plastic, fossil fuels and the main constitute of cigarette smoking. However, the effect of acrolein on MI remains not that clear. In the current study, the effect of acrolein-exacerbated MI was investigated. In vivo, male Sprague-Dawley rats received olive leaf extract (OLE) followed by acrolein, then isoprenaline (ISO) was received by subcutaneous injection to induce MI. Results showed that the expression levels of GRP78 and CHOP, two major components of endoplasmic reticulum (ER) stress were higher in the combination of acrolein and ISO than those in ISO treatment. The apoptosis marker, Bax, was also higher while the anti-apoptosis indicator, Bcl2 expression was lower both at protein and mRNA levels in the combination group. Also, the acrolein-protein adducts and myocardial pathological damage increased in the combination of acrolein and ISO relative to the ISO treatment. Besides, cardiac parameters, ejection fraction (EF) and fractional shortening (FS) were reduced more significantly when acrolein was added than in ISO treatment. Interestingly, all the changes were able to be ameliorated by OLE. Since hydroxytyrosol (HT) and oleuropein (OP) were the main components in OLE, we next investigated the effect of HT and OP on cardiomyocyte H9c2 cell apoptosis induced by acrolein through ER stress and Bax pathway. Results showed that GRP78, CHOP and Bax expression were upregulated, while Bcl2 expression was downregulated both at the protein and mRNA levels, when the H9c2 cells were treated with acrolein. In addition, pretreatment with HT can reverse the expression of GRP78, CHOP, Bax and Bcl2 on the protein and mRNA levels, while there was no effect of OP on the expression of GRP78 and CHOP on the mRNA levels. Overall, all these results demonstrated that OLE and the main components (HT and OP) could prevent the negative effects of acrolein on myocardium and cardiomyocytes.

The novel bis-benzylisoquinoline PY35 reverses P-glycoprotein-mediated multidrug resistance.

Multidrug resistance (MDR) to chemotherapeutic drugs is the main cause of chemotherapy failure in cancer treatment, and it generally results from expression of ATP-dependent efflux pump P-glycoprotein (P-gp). MDR reversal agents typically act by inhibiting the drug efflux activity of P-gp, thereby increasing intracellular drug levels. PY35 is a novel 5-substituted tetrandrine (Tet) derivative (CN Application No. 201210238709.6). The present study was performed to investigate the ability of PY35 to reverse P-gp-mediated MDR and its mechanism in resistant K562/Adriamycin (ADM), MCF-7/ADM cells and their sensitive cell lines K562 and MCF-7. The ability of PY35 to reverse drug resistance was evaluated by MTT assay. The results showed that PY35 can reverse MDR more effectively than the drug prototype­Tet. The P-gp function was assessed by the Rhodamine 123 (Rho-123; a P-gp substrate) uptake assay with flow cytometry (FCM) and laser scanning confocal microscopes (LSCM); it showed that the MDR cells pumped Rho-123 out the cells, while their sensitive cells scarcely showed efflux. The presence of PY35 efficiently decreased the efflux of the Rho-123, showing that PY35 can reverse P-gp-mediated MDR by increasing the intracellular concentration of Rho-123. The intracellular accumulation of ADM was analyzed by FCM and showed that the coadministration of PY35 and ADM had clearer accumulation than the treatment of Tet and ADM, and was also more evident than treatment with only ADM. The effect of PY35 on the expression of P-gp was assessed by western blotting. The results indicated that PY35 does not inhibit the expression level of the P-gp. This study indicated that PY35 can effectively reverse P-gp-mediated MDR, not by inhibiting the expression of P-gp, but by the coadministration of PY35 and ADM that could increase the intracellular accumulation of drugs. Thus, PY35 may be a potential inhibitor to overcome drug resistance.