Vascular Electrical Stimulation with Wireless, Battery-Free, and Fully Implantable Features Reduces Atherosclerotic Plaque Formation Through Sirt1-Mediated Autophagy.

Electrical stimulation (ES) is a safe and effective procedure in clinical rehabilitation with few adverse effects. However, studies on ES for atherosclerosis (AS) are scarce because ES does not provide a long-term intervention for chronic disease processes. Battery-free implants and surgically mounted them in the abdominal aorta of high-fat-fed Apolipoprotein E (ApoE-/- ) mice are used, which are electrically stimulated for four weeks using a wireless ES device to observe changes in atherosclerotic plaques. Results showed that there is almost no growth of atherosclerotic plaque at the stimulated site in AopE-/- mice after ES. RNA-sequencing (RNA-seq) analysis of Thp-1 macrophages reveal that the transcriptional activity of autophagy-related genes increase substantially after ES. Additionally, ES reduces lipid accumulation in macrophages by restoring ABCA1- and ABCG1-mediated cholesterol efflux. Mechanistically, it is demonstrated that ES reduced lipid accumulation through Sirtuin 1 (Sirt1)/Autophagy related 5 (Atg5) pathway-mediated autophagy. Furthermore, ES reverse autophagic dysfunction in macrophages of AopE-/- mouse plaques by restoring Sirt1, blunting P62 accumulation, and inhibiting the secretion of interleukin (IL)-6, resulting in the alleviation of atherosclerotic lesion formation. Here, a novel approach is shown in which ES can be used as a promising therapeutic strategy for AS treatment through Sirt1/Atg5 pathway-mediated autophagy.

Combined effects of irbesartan and carvedilol on expression of tissue factor and tissue factor pathway inhibitor in rats after myocardial infarction.

The objective of this study was to investigate the effects of irbesartan, carvedilol, and irbesartan plus carvedilol on the expression of tissue factor (TF) and tissue factor pathway inhibitor (TFPI) mRNA and protein in rat myocardium after myocardial infarction (MI). MI was induced in male Wistar rats by ligation of the anterior descending branch of the left coronary artery. Irbesartan at 50 mg/kg/day, carvedilol at 1 mg/kg/day, irbesartan plus carvedilol, or placebo was administered intragastrically; expression of TF and TFPI mRNA and protein was determined by RT-PCR and Western blot analysis. The relative left ventricle weights were lower in all three treatment groups than in the placebo group, with the lowest relative weight in the irbesartan plus carvedilol group (P < 0.001). The size of the infarcted area was lower in the carvedilol and the combined groups than in the placebo group (P < 0.001). The levels of expression of TF and TFPI mRNA and protein were lower in the combined group than in the placebo group or the carvedilol group (P < 0.001). Treatment with irbesartan plus carvedilol reduced the expression of TF and TFPI mRNA and protein after MI in rats, and combined treatment with both agents had greater effects than the single agents alone. These findings suggest that the beneficial effects of these drugs may include anticoagulation and that combined therapy with both agents is an option that should be evaluated further.

Surface PEGylated Cancer Cell Membrane-Coated Nanoparticles for Codelivery of Curcumin and Doxorubicin for the Treatment of Multidrug Resistant Esophageal Carcinoma.

OBJECTIVE: The emergence of multi-drug resistance (MDR) in esophageal carcinoma has severely affected the effect of chemotherapy and shortened the survival of patients. To this end, we intend to develop a biomimetic nano-targeting drug modified by cancer cell membrane, and investigate its therapeutic effect. METHODS: The degradable poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) co-loaded with doxorubicin (DOX) and curcumin (Cur) were prepared by solvent evaporation method. TE10 cell membrane and Distearoyl phosphatidylethanolamine-polyethylene glycol (DSPE-PEG) were then coated on the PLGA NPs by membrane extrusion to prepare the PEG-TE10@PLGA@DOX-Cur NPs (PMPNs). Size and zeta potential of the PMPNs were analyzed by lazer particle analyzer, and the morphology of PMPNs was observed by transmission electron microscope. The TE10 cell membrane protein on PMPNs was analyzed by gel electrophoresis. The DOX-resistant esophageal cancer cell model TE10/DOX was established through high-dose induction. The In vitro homologous targeting ability of PMPNs was evaluated by cell uptake assay, and the in vitro anti-tumor effect of PMPNs was assessed through CCK-8, clone formation and flow cytometry. A Balb/c mouse model of TE10/DOX xenograft was constructed to evaluate the anti-tumor effect in vivo and the bio-safety of PMPNs. RESULTS: The prepared cell membrane coated PMPNs had a regular spherical structure with an average diameter of 177 nm. PMPNs could directly target TE10 and TE10/DOX cells or TE10/DOX xenografted tumor and effectively inhibit the growth of DOX-resistant esophageal carcinoma. Besides, the PMPNs was confirmed to have high biosafety. CONCLUSION: In this study, a targeted biomimetic nano-drug delivery system PMPNs was successfully prepared, which overcome the MDR of esophageal carcinoma by co-delivering DOX and sensitizer curcumin.

Real world effectiveness of PCSK-9 inhibitors combined with statins versus statins-based therapy among patients with very high risk of atherosclerotic cardiovascular disease in China (RWE-PCSK study).

BACKGROUND: The efficacy and safety of proprotein convertase subtilisin/kexin type 9 (PCSK-9) inhibitors were confirmed by several clinical trials, but its effectiveness in routine clinical practice in China has not been evaluated. This study aims to describe the real world effectiveness of PCSK-9 inhibitors combined with statins compared with statins-based therapy among patients with very high risk of atherosclerotic cardiovascular disease (ASCVD). METHODS: This is a multi-center observational study, enrolled patients from 32 hospitals who underwent percutaneous coronary intervention (PCI) from January to June in 2019. There are 453 patients treated with PCSK-9 inhibitors combined with statins in PCSK-9 inhibitor group and 2,610 patients treated with statins-based lipid lowering therapies in statins-based group. The lipid control rate and incidence of major adverse cardiovascular events (MACE) over six months were compared between two groups. A propensity score-matched (PSM) analysis was used to balance two groups on confounding factors. Survival analysis using Kaplan-Meier methods was applied for MACE. RESULTS: In a total of 3,063 patients, 89.91% of patients had received moderate or high-intensity statins-based therapy before PCI, but only 9.47% of patients had low-density lipoprotein cholesterol (LDL-C) levels below 1.4 mmol/L at baseline. In the PSM selected patients, LDL-C level was reduced by 42.57% in PCSK-9 inhibitor group and 30.81% (P < 0.001) in statins-based group after six months. The proportion of LDL-C ≤ 1.0 mmol/L increased from 5.29% to 29.26% in PCSK-9 inhibitor group and 0.23% to 6.11% in statins-based group, and the proportion of LDL-C ≤ 1.4 mmol/L increased from 10.36% to 47.69% in PCSK-9 inhibitor group and 2.99% to 18.43% in statins-based group ( P < 0.001 for both). There was no significant difference between PCSK-9 inhibitor and statins-based treatment in reducing the risk of MACE (hazard ratio = 2.52, 95% CI: 0.49-12.97, P = 0.250). CONCLUSIONS: In the real world, PCSK-9 inhibitors combined with statins could significantly reduce LDL-C levels among patients with very high risk of ASCVD in China. The long-term clinical benefits for patients received PCSK-9 inhibitor to reduce the risk of MACE is still unclear and requires further study.

NLK interacts with 14­3­3ζ to restore the expression of E­cadherin.

The Nemo­like kinase (NLK), a conserved serine/threonine kinase, plays a critical role in the regulation of a variety of transcription factors, with important roles in determining cell fate. Although recent studies have demonstrated decreased expression patterns of NLK in various types of human cancer, the functional mechanism of NLK in cancer development has not been elucidated. Here, in the present study overexpression of NLK was found to inhibit the growth and migration of the non­small cell lung cancer A549 cell line. NLK was subsequently found to interact with 14­3­3ζ (also known as YWHAZ), which is responsible for E­cadherin silencing during epithelial­mesenchymal transition (EMT). Furthermore, NLK overexpression was able to restore the expression of E­cadherin inhibited by 14­3­3ζ. Notably, NLK interacts with 14­3­3ζ and prevents its dimerization, which is essential for 14­3­3ζ stability and function. By fusing two copies of the 14­3­3ζ gene, via a Gly­rich linker, a non­dissociable dimer of 14­3­3ζ was formed. It was found that NLK was unable to restore the expression of E­cadherin inhibited by the overexpression of the fused dimer of 14­3­3ζ. In addition, the increased ability of migration induced by the overexpression of fused 14­3­3ζ dimer could not be altered by NLK overexpression. The results from the present study indicate that NLK is a negative regulator of 14­3­3ζ and plays a tumor suppressive role in the inhibition of cancer cell migration.

Valsartan inhibits RhoA-ROCK2-MYL pathway in rat model of alcoholic cardiomyopathy.

The present study aimed to investigate variations in the Ras homolog gene family, member A (RhoA)-Rho-associated protein kinase 2 (ROCK2)-myosin light chain (MYL) pathway in a rat model of alcoholic cardiomyopathy (ACM) and the role of angiotensin-converting enzyme inhibitor drugs. Rat models of ACM were established via alcoholic gavage + free access to alcohol. The structural and functional changes of the heart were analyzed by hematoxylin-eosin staining, Masson's trichrome staining, immunohistochemistry staining, western blotting and fluorescence quantitative polymerase chain reaction. A total of 16 weeks later, a decreased ejection fraction and left ventricular fractional shortening in the alcohol group compared with the control group were demonstrated resulting in an increased left ventricular end diastolic diameter. These adverse effects were ameliorated following treatment with valsartan. In addition, the alcohol group revealed a disorganized arrangement of myocardial filaments, which was improved upon treatment with valsartan. RhoA and ROCK2 protein expression significantly increased in myocardial cells in the alcohol compared with the control group. Following drug intervention with valsartan, expression of RhoA and ROCK2 proteins were inhibited in the alcohol group. Furthermore, significantly elevated RhoA and ROCK2 and decreased MYL protein and mRNA expression in the alcohol group was demonstrated compared with the control group. Administration of valsartan reversed the expression profile of RhoA, ROCK and MYL in ACM. Expression of RhoA and ROCK were elevated with downregulation of MYL resulting in heart failure. However, the angiotensin receptor antagonist diminished the expression of RhoA and ROCK and enhanced the expression of MYL. The results of the present study suggest a curative effect of valsartan in ACM.

Neutrophil extracellular traps induced by IL-8 aggravate atherosclerosis via activation NF-κB signaling in macrophages.

Here, we sought to explore the underlying role of interleukin (IL)-8 in neutrophil extracellular traps (NETs) formation during atherosclerosis (AS). The concentration of pro-inflammatory cytokines IL-8, IL-6 and IL-1ß was determined by enzyme-linked immunosorbent assay (ELISA). NETs formation was evaluated by immunofluorescence and myeloperoxidase (MPO)-DNA complex ELISA. The mRNA levels of IL-8 and Toll-like receptor 9 (TLR9) were measured by quantitative real-time PCR (qRT-PCR). The phosphorylation levels of NF-κB p65 were detected by western blotting. The hematoxylin and eosin (H&E) staining of atherosclerotic lesion areas was performed in ApoE-deficiency mice. Results showed that patients with AS showed higher serum levels of IL-8, a pro-inflammatory cytokine and NETs. IL-8 interacted with its receptor CXC chemokine receptor 2 (CXCR2) on neutrophils, leading to the formation of NETs via Src and extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinases (MAPK) signaling to aggravate AS progression in vivo. PMA-induced NETosis directly upregulated the TLR9/NF-κB pathway in macrophages and subsequently initiated the release of IL-8. Our data reveal a neutrophil-macrophage interaction in AS progression, and indicate that NETs represent as a novel therapeutic target in treatment of AS and other cardiovascular diseases (CVD).

miR-130b-5p promotes proliferation, migration and invasion of gastric cancer cells via targeting RASAL1.

The aim of the present study was to investigate the targeted interaction between microRNA (miR)-130b-5p and RAS protein activator like 1 (RASAL1) gene and elucidate the function of miR-130b-5p in cell proliferation, migration and invasion in gastric cancer. Expression of miR-130b-5p and RASAL1 in seven gastric cell lines was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). MGC803 cells were selected for further study since they exhibited a marked increase in expression of miR-130b-5p accompanied by decreased expression of RASAL1. MGC803 cells were transfected with miR-130b-5p mimics and miR-130b-5p inhibitor using Lipofectamine 2000 for over- and underexpression, respectively, with cells transfected with negative control (NC) sequence as the control. In addition, a luciferase reporter gene assay was performed to evaluate the targeted interaction between miR-130b-5p and RASAL1. Then, alterations in RASAL1 expression were detected by RT-qPCR and western blot analysis following transfection with miR-130b-5p mimics and miR-130b-5p inhibitor. Cell proliferation, colony formation, and migration and invasion ability were detected by MTT, colony formation and Transwell assays, respectively. RASAL1 was demonstrated to be a target gene of miR-130b-5p by luciferase reporter gene assay. In addition, the expression of RASAL1 was significantly lower in MGC803 cells that were transfected with miR-130b-5p mimics and significantly higher in cells transfected with miR-130b-5p inhibitor in comparison with cells transfected with NC (P<0.05). Furthermore, the experimental group transfected with miR-130b-5p mimics manifested significantly higher cell proliferation, increased colony formation and increased migratory and invasive capacities (P<0.05). By contrast, cells transfected with miR-130b-5p inhibitor exhibited significantly lower cell proliferation, decreased colony formation and decreased migratory and invasive capacities, compared with cells transfected with NC (P<0.05). In conclusion, RASAL1 was demonstrated to be a target gene of miR-130b-5p. Overexpression of miR-130b-5p results in promoted proliferation, colony formation and migration and invasion abilities through targeted modulation of RASAL1.

TAB3 promotes human esophageal squamous cell carcinoma proliferation and invasion via the NF­κB pathway.

Esophageal squamous cell carcinoma (ESCC) has become one of the most common causes of cancer­associated mortality worldwide. Transforming growth factor­activated kinase (TAK1)­binding protein 3 (TAB3) is essential for activation of the NF (NF)­κB pathway in response to TAK1 activation. The NF­κB pathway serves important roles in tumor cell proliferation and migration; however, the clinical relevance of TAB3 and its biological function in ESCC progression remain elusive. The present study investigated the expression and function of TAB3 in ESCC tissues, and its association with the clinical prognosis of patients. The results demonstrated that TAB3 expression was significantly increased in human ESCC cell lines and tissue samples, and the expression of TAB3 was associated with ESCC lymph node metastasis, T stage, pathological grade and Ki­67 expression in 80 ESCC samples, as determined by immunohistochemistry. Patients with ESCC and high TAB3 expression exhibited worse overall survival. Furthermore, knockdown of TAB3 by small interfering RNA inhibited the proliferation of ESCC cells, and reduced the migration and invasion of ESCC cells. In addition, knockdown of TAB3 decreased the expression of the NF­κB pathway in TE­1 cells. Taken together, these results demonstrated that TAB3 may be a promising therapeutic target for the treatment of ESCC.

ATP citrate lyase expression is associated with advanced stage and prognosis in gastric adenocarcinoma.

Gastric adenocarcinoma (GA) is one of the most common cancer worldwide. ATP citrate lyase (ACLY) is generally recognized as a key enzyme of de novo fatty acid synthesis responsible for generation of oxaloacetate and cytosolic acetyl-CoA. This study aimed to investigate the expression level of ACLY in GA and evaluate the relationship between ACLY expression and the prognosis of GA patients. Paraffin archived samples from 83 GA patients were used to analyze ACLY expression by immunohistochemistry. ACLY was significantly upregulated in GA tissues compared with adjacent normal tissues (P < 0.001). High ACLY expression was correlated with advanced stages (P = 0.007) and lymph node metastasis (P = 0.022). Furthermore, patients with low ACLY expression had longer survival time than those with high ACLY expression (P = 0.031). In conclusion, these results indicate that ACLY might serve as a biomarker to predict the progression and prognosis of GA patients.

MG132 proteasome inhibitor upregulates the expression of connexin 43 in rats with adriamycin-induced heart failure.

The connexin 43 (Cx43) gap junction protein is important in the synchronization of contraction of cardiac myocytes. Abnormal expression of Cx43 contributes to ventricular arrhythmia, which is the major cause of sudden death in heart failure (HF). Cx43 is known to interact with zonula occludens (ZO)­1, and the proteasome is involved in the regulation of Cx43 degradation. Although Cx43 is downregulated in heart failure, the underlying mechanisms remain to be elucidated. The present study aimed to investigate the effect of the MG132 proteasome inhibitor on the expression levels of Cx43, ZO­1, 20S proteasome and ubiquitin in a rat model of HF, induced by adriamycin. MG132 reduced adriamycin­induced injury in the failing heart. In addition, MG132 inhibited the expression of 20S proteasome and ubiquitin, accompanied by an upregulation in the expression of Cx43 and ZO­1. These findings suggested that inhibition of the ubiquitin­proteasome system upregulated the expression of Cx43. Therefore, the proteasome inhibitor may be used to prevent degradation of Cx43 in HF, and thus may prevent Cx43-mediated arrhythmia in HF.

RASAL1 attenuates gastric carcinogenesis in nude mice by blocking RAS/ERK signaling.

Recent studies have suggested that the RAS protein activator like-1 (RASAL1) functions as a tumor suppressor in vitro and may play an important role in the development of gastric cancer. However, whether or not RASAL1 suppresses tumor growth in vivo remains to be determined. In the present study, we investigated the role of RASAL1 in gastric carcinogenesis using an in vivo xenograft model. A lentiviral RASAL1 expression vector was constructed and utilized to transfect the human poorly differentiated gastric adenocarcinoma cell line, BGC-823. RASAL1 expression levels were verified by quantitative real-time RT-PCR and Western blotting analysis. Then, we established the nude mice xenograft model using BGC-823 cells either over-expressing RASAL1 or normal. After three weeks, the results showed that the over-expression of RASAL1 led to a significant reduction in both tumor volume and weight compared with the other two control groups. Furthermore, in xenograft tissues the increased expression of RASAL1 in BGC-823 cells caused decreased expression of p-ERK1/2, a downstream moleculein the RAS/RAF/MEK/ERK signal pathway. These findings demonstrated that the over-expression of RASAL1 could inhibit the growth of gastric cancer by inactivation of the RAS/RAF/MEK/ERK pathway in vivo. This study indicates that RASAL1 may attenuate gastric carcinogenesis.