CRL4 antagonizes SCFFbxo7-mediated turnover of cereblon and BK channel to regulate learning and memory.

Intellectual disability (ID), one of the most common human developmental disorders, can be caused by genetic mutations in Cullin 4B (Cul4B) and cereblon (CRBN). CRBN is a substrate receptor for the Cul4A/B-DDB1 ubiquitin ligase (CRL4) and can target voltage- and calcium-activated BK channel for ER retention. Here we report that ID-associated CRL4CRBN mutations abolish the interaction of the BK channel with CRL4, and redirect the BK channel to the SCFFbxo7 ubiquitin ligase for proteasomal degradation. Glioma cell lines harbouring CRBN mutations record density-dependent decrease of BK currents, which can be restored by blocking Cullin ubiquitin ligase activity. Importantly, mice with neuron-specific deletion of DDB1 or CRBN express reduced BK protein levels in the brain, and exhibit similar impairment in learning and memory, a deficit that can be partially rescued by activating the BK channel. Our results reveal a competitive targeting of the BK channel by two ubiquitin ligases to achieve exquisite control of its stability, and support changes in neuronal excitability as a common pathogenic mechanism underlying CRL4CRBN-associated ID.

Danshen (Salvia miltiorrhiza) restricts MD2/TLR4-MyD88 complex formation and signalling in acute myocardial infarction-induced heart failure.

Heart failure (HF) represents a major public health burden. Inflammation has been shown to be a critical factor in the progression of HF, regardless of the aetiology. Disappointingly, the majority of clinical trials targeting aspects of inflammation in patients with HF have been largely negative. Many clinical researches demonstrate that danshen has a good efficacy on HF, and however, whether danshen exerts anti-inflammatory effects against HF remains unclear. In our study, the employment of a water extracted and alcohol precipitated of danshen extract attenuated cardiac dysfunction and inflammation response in acute myocardial infarction-induced HF rats. Transcriptome technique and validation results revealed that TLR4 signalling pathway was involved in the anti-inflammation effects of danshen. In vitro, danshen reduced the release of inflammatory mediators in LPS-stimulated RAW264.7 macrophage cells. Besides, the LPS-stimulated macrophage conditioned media was applied to induce cardiac H9C2 cells injury, which could be attenuated by danshen. Furtherly, knock-down and overexpression of TLR4 were utilized to confirm that danshen ameliorated inflammatory injury via MyD88-dependent TLR4-TRAF6-NF-κB signalling pathway in cardiomyocytes. Furthermore, by utilizing co-immunoprecipitation, danshen was proved to suppress MD2/TLR4 complex formation and MyD88 recruitment. In conclusion, our results demonstrated that danshen ameliorates inflammatory injury by controlling MD2/TLR4-MyD88 complex formation and TLR4-TRAF6-NF-κB signalling pathway in acute myocardial infarction-induced HF.

Role of epithelial Na+ channels in endothelial function.

An increasing number of mechano-sensitive ion channels in endothelial cells have been identified in response to blood flow and hydrostatic pressure. However, how these channels respond to flow under different physiological and pathological conditions remains unknown. Our results show that epithelial Na(+) channels (ENaCs) colocalize with hemeoxygenase-1 (HO-1) and hemeoxygenase-2 (HO-2) within the caveolae on the apical membrane of endothelial cells and are sensitive to stretch pressure and shear stress. ENaCs exhibited low levels of activity until their physiological environment was changed; in this case, the upregulation of HO-1, which in turn facilitated heme degradation and hence increased the carbon monoxide (CO) generation. CO potently increased the bioactivity of ENaCs, releasing the channel from inhibition. Endothelial cells responded to shear stress by increasing the Na(+) influx rate. Elevation of intracellular Na(+) concentration hampered the transportation of l-arginine, resulting in impaired nitric oxide (NO) generation. Our data suggest that ENaCs that are endogenous to human endothelial cells are mechano-sensitive. Persistent activation of ENaCs could inevitably lead to endothelium dysfunction and even vascular diseases such as atherosclerosis.

Inhibition of pyruvate kinase M2 by reactive oxygen species contributes to the development of pulmonary arterial hypertension.

AIMS: Pulmonary arterial hypertension [1] is a proliferative disorder associated with enhanced proliferation and suppressed apoptosis of pulmonary artery smooth muscle cells (PASMCs). Reactive oxygen species (ROS) is implicated in the development of PAH and regulates the vascular tone and functions. However, which cellular signaling mechanisms are triggered by ROS in PAH is still unknown. Hence, here we wished to characterize the signaling mechanisms triggered by ROS. METHODS AND RESULTS: By Western blots, we showed that increased intracellular ROS caused inhibition of the glycolytic pyruvate kinase M2 (PKM2) activity through promoting the phosphorylation of PKM2. Monocrotaline (MCT)-induced rats developed severe PAH and right ventricular hypertrophy, with a significant increase in the P-PKM2 and decrease in pyruvate kinase activity which could be attenuated with the treatments of PKM2 activators, FBP and l-serine. The antioxidant NAC, apocynin and MnTBAP had the similar protective effects in the development of PAH. In vitro assays confirmed that inhibition of PKM2 activity could modulate the flux of glycolytic intermediates in support of cell proliferation through the increased pentose phosphate pathway (PPP). Increased ROS and decreased PKM2 activity also promoted the Cav1.2 expression and intracellular calcium. CONCLUSION: Our data provide new evidence that PKM2 makes a critical regulatory contribution to the PAHs for the first time. Decreased pyruvate kinase M2 activity confers additional advantages to rat PASMCs by allowing them to sustain anti-oxidant responses and thereby support cell survival in PAH. It may become a novel treatment strategy in PAH by using of PKM2 activators.

The ACAT2 expression of human leukocytes is responsible for the excretion of lipoproteins containing cholesteryl/steryl esters.

Acyl-coenzymeA:cholesterol acyltransferase 2 (ACAT2) is abundantly expressed in intestine and fetal liver of healthy human. Our previous studies have shown that in monocytic cells the low-level expression of human ACAT2 gene with specific CpG-hypomethylated promoter is regulated by the CCAAT/enhancer binding protein (C/EBP) transcription factors. In this study, we further report that the ACAT2 gene expression is attributable to the C/EBPs in the human leukocytes and correlated with the excretion of fluorescent lipoproteins containing the ACAT2-catalyzed NBD22-steryl esters. Moreover, this lipoprotein excretion can be inhibited by the ACAT2 isoform-selective inhibitor pyripyropene A (PPPA) in a dose-dependent manner, and employed to determine the half maximum inhibitory concentration (IC50) values of PPPA. Significantly, it is found that the differentiation-inducing factor all-trans retinoic acid, but not the proinflammatory cytokine tumor necrosis factor-α, enhances this ACAT2-dependent lipoprotein excretion. These data demonstrate that the ACAT2 expression of human leukocytes is responsible for the excretion of lipoproteins containing cholesteryl/steryl esters (CE/SE), and suggest that the excretion of lipoproteins containing the ACAT2-catalyzed CS/SE may avoid cytotoxicity through decreasing the excess intracellular cholesterols/sterols (especially various oxysterols), which is essential for the action of the human leukocytes.

Low-level expression of human ACAT2 gene in monocytic cells is regulated by the C/EBP transcription factors.

Acyl-coenzyme A:cholesterol acyltransferases (ACATs) are the exclusive intracellular enzymes that catalyze the formation of cholesteryl/steryl esters (CE/SE). In our previous work, we found that the high-level expression of human ACAT2 gene with the CpG hypomethylation of its whole promoter was synergistically regulated by two transcription factors Cdx2 and HNF1α in the intestine and fetal liver. Here, we first observed that the specific CpG-hypomethylated promoter was correlated with the low expression of human ACAT2 gene in monocytic cell line THP-1. Then, two CCAAT/enhancer binding protein (C/EBP) elements within the activation domain in the specific CpG-hypomethylation promoter region were identified, and the expression of ACAT2 in THP-1 cells was evidently decreased when the C/EBP transcription factors were knock-downed using RNAi technology. Furthermore, ChIP assay confirmed that C/EBPs directly bind to their elements for low-level expression of human ACAT2 gene in THP-1 cells. Significantly, the increased expressions of ACAT2 and C/EBPs were also found in macrophages differentiated from both ATRA-treated THP-1 cells and cultured human blood monocytes. These results demonstrate that the low-level expression of human ACAT2 gene with specific CpG-hypomethylated promoter is regulated by the C/EBP transcription factors in monocytic cells, and imply that the lowly expressed ACAT2 catalyzes the synthesis of certain CE/SE that are assembled into lipoproteins for the secretion.

Excess TNF-α in the blood activates monocytes with the potential to directly form cholesteryl ester-laden cells.

The tumor necrosis factor-α (TNF-α) and monocytic cells play a critical role in the development of atherosclerosis, which is the major cause of coronary heart disease (CHD). In this work, we investigated the effect of excess TNF-α on monocytes in the blood and found that blood monocytes from the CHD patients had the potential to directly form cholesteryl ester (CE)-laden cells under the in vitro incubation with oxLDL. The plasma levels of proinflammatory cytokines, such as TNF-α, interleukin 6 (IL-6), and C reactive protein (CRP), in the CHD patients were significantly higher than those in the control healthy volunteers. However, only the plasma level of TNF-α, but not of IL-6 or CRP, is positively correlated with the potential of blood monocytes to directly form CE-laden cells. By using human blood monocytes and monocytic THP-1 cells, the activating effect of TNF-α on the formation of the CE-laden cells was demonstrated, which could be specifically blocked by the anti-TNF-α antibody. Furthermore, it was also revealed that TNF-α could boost adhesion and oxLDL uptake of the monocytes by enhancing the expression of the functional adhesion molecules and scavenger receptors, respectively. Finally, the results of in vivo and in vitro experiments with a mouse model confirmed that excess TNF-α in the blood activates monocytes with the potential to directly form CE-laden cells. These data demonstrate that excess TNF-α in the blood is the primary trigger for the development of atherosclerosis and CHD.

ACAT1 regulates the dynamics of free cholesterols in plasma membrane which leads to the APP-α-processing alteration.

Acyl-CoA:cholesterol acyltransferase 1 (ACAT1) is a key enzyme exclusively using free cholesterols as the substrates in cell and is involved in the cellular cholesterol homeostasis. In this study, we used human neuroblastoma cell line SK-N-SH as a model and first observed that inhibiting ACAT1 can decrease the amyloid precursor protein (APP)-α-processing. Meanwhile, the transfection experiments using the small interfering RNA and expression plasmid of ACAT1 indicated that ACAT1 can dependently affect the APP-α-processing. Furthermore, inhibiting ACAT1 was found to increase the free cholesterols in plasma membrane (PM-FC), and the increased PM-FC caused by inhibiting ACAT1 can lead to the decrease of the APP-α-processing, indicating that ACAT1 regulates the dynamics of PM-FC, which leads to the alteration of the APP-α-processing. More importantly, further results showed that under the ACAT1 inhibition, the alterations of the PM-FC and the subsequent APP-α-processing are not dependent on the cellular total cholesterol level, confirming that ACAT1 regulates the dynamics of PM-FC. Finally, we revealed that even when the Niemann-Pick-Type C-dependent pathway is blocked, the ACAT1 inhibition still obviously results in the PM-FC increase, suggesting that the ACAT1-dependent pathway is responsible for the shuttling of PM-FC to the intracellular pool. Our data provide a novel insight that ACAT1 which enzymatically regulates the dynamics of PM-FC may play important roles in the human neuronal cells.

One-step electrodeposition of MWCNTs-Cu MOF films for the ratiometric electrochemical analysis of glyphosate.

Sensitive and accurate determination of glyphosate (GLYP) is vital for food safety and environmental protection. Herein, a novel electrochemical ratiometric biosensor was designed for the accurate quantification of GLYP through one-step electrodeposition of MWCNTs-Cu MOF films. MWCNTs-Cu MOF nanostructures were directly electro-synthesized in situ on the electrode from the precursor solution. The combination of Cu MOFs with MWCNTs not merely improved the conductivity of MOFs, but also enhanced the sensitivity of the biosensor. Furthermore, Cu sites within Cu MOFs were turned into CuCl to further amplify the current signal and enable the specific recognition of GLYP through competing reactions with the transformation of CuCl into non-electroactive Cu-GLYP. Meanwhile, internal reference molecules of methylene blue (MB) were incorporated to improve the measurement accuracy of GLYP for reducing unpredictable measurement errors aroused by environmental deviations. The ratiometric electrochemical sensor exhibited a high linearity with the logarithmic value of GLYP concentration from 0.5 nM to 400 nM. The detection limit was estimated to be as low as 0.014 nM. Finally, the present sensor with ratiometric signal export was applied for GLYP analysis in real samples with high sensitivity and accuracy. The simplicity and reliability of the ratiometric sensor make it a worthy and powerful tool for food and environmental monitoring. This design strategy also provides an avenue for the development of simple and efficient biosensors for other substances.

In vivo monitoring of glutathione in a live rat brain based on the ratiometric signal output of 2D Cu-TCPP(Fe) nanosheets.

Herein, a novel ratiometric electrochemical platform was developed for in vivo analysis of GSH based on the dual signal output of 2D Cu-TCPP(Fe) nanosheets. Our method with high selectivity and high accuracy enabled GSH monitoring in a live rat brain, and accurate GSH concentrations were firstly reported in different brain regions upon global cerebral ischemia.

An electrochemical microsensor based on a specific recognition element for the simultaneous detection of hydrogen peroxide and ascorbic acid in the live rat brain.

A novel electrochemical microsensor was developed for the ratiometric and simultaneous determination of hydrogen peroxide (H2O2) and ascorbic acid (AA) based on the borate-phenol "switch" recognition mechanism and carbon nanotube (CNT) catalytic characteristics. First of all, a carbon fiber microelectrode (CFME) was coated with CNTs. Then, a specific probe, 9-anthraceneboronic acid pinacol ester (9-AP), was screened and decorated on CNTs through π-π stacking for the recognition of H2O2 based on the transformation of boric acid ester into electroactive phenols. CNTs not only served as the amplifiers of current signals, but also as catalysts facilitating AA oxidation. Meanwhile, ferrocenecarboxylic acid (Fc), inert to H2O2 and AA, was modified on another amino-functionalized CNT microelectrode via an amide bond as an internal reference channel for avoiding errors caused by environmental discrepancies. The two-channel ratiometric microsensor enabled the sensitive and accurate detection of H2O2 and AA simultaneously, and the detection limits were estimated to be 0.09 µM and 4.12 µM, respectively. The developed microsensor with remarkable analytical performance was finally applied for the simultaneous detection of H2O2 and AA in the live rat brain.

Transcriptomic Study on the Lungs of Broilers with Ascites Syndrome.

Although broiler ascites syndrome (AS) has been extensively studied, its pathogenesis remains unclear. The lack of cardiopulmonary function in broilers causes relative hypoxia in the body; hence, the lung is the main target organ of AS. However, the transcriptome of AS lung tissue in broilers has not been studied. In this study, an AS model was successfully constructed, and lung tissues of three AS broilers and three healthy broilers were obtained for RNA sequencing (RNA-seq) and pathological observation. The results showed that 614 genes were up-regulated and 828 genes were down-regulated in the AS group compared with the normal group. Gene Ontology (GO) functional annotation revealed the following up-regulated genes: FABP4, APLN, EIF2AK4, HMOX1, MMP9, THBS1, TLR4, BCL2; and down-regulated genes: APELA, FGF7, WNT5A, CDK6, IL7, IL7R, APLNR. These genes have attracted much attention in cardiovascular diseases such as pulmonary hypertension. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that multiple metabolic processes were enriched, indicating abnormal lung metabolism of AS in broilers. These findings elucidate the potential genes and signal pathways in the lungs of broilers with AS and provide a potential target for studying the pathogenesis and preventing AS.

Qishen granule (QSG) exerts cardioprotective effects by inhibiting NLRP3 inflammasome and pyroptosis in myocardial infarction rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Qishen granule (QSG) is a traditional Chinese medicine formulation that is widely used in clinical practice for the treatment of myocardial infarction (MI), and its efficacy and safety have been well approved. However, the underlying mechanism by which QSG alleviates inflammation and cell pyroptosis remains unknown. AIM OF THE STUDY: The aim of this study was to clarify whether QSG ameliorated MI by inhibiting inflammasome activation and cell pyroptosis. MATERIALS AND METHODS: In vivo, SD male rats were subjected to the left anterior ascending branch (LAD) ligation to construct MI model. And in vitro, OGD/R, ISO, Ang II and LPS-ATP were used to induce H9C2 cell injury. Cell viability and ROS were detected by CCK8 and DCFH-DA dye respectively. Western blots were applied to detect the expression of inflammasome-related proteins. Cell pyroptosis was evaluated by Calcein-AM/PI staining, Hoechst/PI staining and NT-GSDMD expression. RESULTS: QSG administration improved the cardiac function, as well as reduced inflammatory cell infiltration and collagen deposition. In H9C2 cells, OGD/R failed to induce inflammasome activation, while ISO, Ang II and LPS-ATP successfully induced inflammasome activation and cell pyroptosis, as evidenced by increased Caspase-1(P20) and NT-GSDMD. In LPS-ATP induced H9C2 model, ROS production and cell pyroptosis were suppressed when treated with QSG. Furthermore, QSG significantly decreased the protein levels of P65-NF-κB, NLRP3, ASC, Caspase-1 (P20), Cleaved IL-18, Cleaved IL-1ß and NT-GSDMD. CONCLUSION: This study is the first to demonstrate that QSG has cardioprotective effects by inhibiting inflammasome activation and pyroptosis, which are considered as promising therapeutic targets for MI.

Shexiang Tongxin Dropping Pills Promote Macrophage Polarization-Induced Angiogenesis Against Coronary Microvascular Dysfunction via PI3K/Akt/mTORC1 Pathway.

Background: Accumulating evidence suggests that coronary microvascular dysfunction (CMD) is one of the important causes of coronary artery diseases. Angiogenesis can effectively improve CMD by increasing blood supply capacity, recovering cardiac function and poor hemodynamics. Clinical studies have approved Shexiang Tongxin dropping pill (STDP), which has exerted remarkable roles on ameliorating CMD, but the effects and mechanisms of STDPs on angiogenesis have not been clarified. Purpose: The purpose of this study was to elucidate the effects and potential mechanisms of STDPs on macrophage polarization-induced angiogenesis against CMD. Methods: Echocardiography, optical microangiography (OMAG), and histological examination were applied to evaluate cardioprotection and proangiogenic effects of STDPs on left anterior descending (LAD) ligation-induced CMD rats. In vitro, oxygen-glucose deprivation-reperfusion (OGD/R)-induced HUVEC model and LPS-stimulated bone marrow-derived macrophage (BMDM) model were established to observe the effects of STDPs on angiogenesis and M2 macrophage polarization. Results: STDPs improved cardiac function, increased microvascular density, and the number of M2 macrophages in the heart of CMD rats. In vitro, STDPs accelerated the proliferation, migration, and tube formation in OGD/R-induced HUVECs similar to the effects of VEGF-A. Furthermore, in LPS-stimulated BMDMs model, STDPs modulated M2 macrophage polarization and increased VEGF-A release via the PI3K/AKT/mTORC1 pathway. Conclusion: STDPs promoted macrophage polarization-induced angiogenesis against CMD via the PI3K/Akt/mTORC1 pathway. Our results demonstrated that the phenotype transformation of macrophages and stimulating the secretion of VEGF-A may be applied as novel cardioprotective targets for the treatment of CMD.

Tanshinone I inhibits doxorubicin-induced cardiotoxicity by regulating Nrf2 signaling pathway.

BACKGROUND: Doxorubicin (DOX) is a powerful anti-tumor anthracycline drug. However, its clinical use is limited due to the side effect of cardiotoxicity. Tanshinone I (Tan I) is one of the major tanshinones isolated from Salvia miltiorrhiza. Studies have shown that Tan I is effective in the treatment of cardiovascular diseases. However, the potential effects of Tan I against DOX-induced cardiotoxicity (DIC) have yet to be explored. PURPOSE: This study aimed to explore whether Tan I can protect against DIC and to reveal whether Tan I can exert anti-oxidative effect by regulating nuclear erythroid factor 2-related factor 2 (Nrf2) pathway. METHODS: DIC models were established in vivo by intravenous injection of DOX. Echocardiography was used to monitor the cardiac function of mice. Transmission electron microscopy was used to assess mitochondrial damage. Oxidative stress was measured by dihydroethidium (DHE) staining and western blotting. The accumulation and nuclear translocation of Nrf2 was detected by immunofluorescence. H9C2 cellular DIC model was established in vitro to explore the pharmacological mechanism. Nrf2 small interfering (si)-RNA was applied to H9C2 cells to explore whether Tan I exerted protective effect against DIC through Nrf2 signaling pathway. The protective effects of Tan I on mitochondrial function and mitochondrial membrane permeability were measured by MitoSOX™ Red and JC-1 staining assays, respectively. RESULTS: In vivo experiments revealed that Tan I could improve cardiac function and protect against DOX-induced myocardial structural damages in mice models. The oxidative stress induced by DOX was suppressed and apoptosis was mitigated by Tan I treatment. Tan I protected against DOX-induced mitochondrial structural damage. Meanwhile, key proteins in Nrf2 pathways were upregulated by Tan I treatment. In vitro studies showed that Tan I attenuated DOX-induced generation of reactive oxygen species (ROS) in cultured H9C2 cells, reduced apoptotic rates, protected mitochondrial functions and up-regulated Nrf2 signaling pathway. Tan I promoted accumulation and nuclear translocation of Nrf2 protein. In addition, interference of Nrf2 abrogated the anti-oxidative effects of Tan I and reversed the expressions of key proteins in Nrf2 pathway. The protective effects of Tan I on mitochondrial integrity was also mitigated by Nrf2 interference. CONCLUSION: Tan I could reduce oxidative stress and protect against DIC through regulating Nrf2 signaling pathway. Nrf2 is a potential target and Tan I is a novel candidate agent for the treatment of DIC.

Exploring the protective effects of PNS on acute myocardial ischaemia-induced heart failure by Transcriptome analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax notoginseng saponins (PNS) were extracted from Panax notoginseng (Burkill) F.H. Chen, a natural product often used as a therapeutic agent in China. PNS has showed obvious therapeutic effect in heart failure (HF) treatment. However, its targets and pharmacological mechanisms remain elusive. AIM OF THE STUDY: This research attempted to determine both the effects and mechanisms of PNS involved in AMI treatment, namely, acute myocardial infarction-induced HF. MATERIALS AND METHODS: An AMI-induced HF model was generated by left anterior descending (LAD) ligation in rats. Transcriptome analyses were performed to identify differentially expressed genes (DEGs) and pathway enrichment. Real-time quantitative PCR (RT-qPCR) verified the HF-related genes differentially expressed after PNS treatment. Finally, a model of H9C2 cells subjected to OGD/R, which is equivalent to oxygen-glucose deprivation/reperfusion, was established to identify the potential mechanism of PNS in the treatment of HF. RESULTS: PNS ameliorated cardiac function and protected against structural alterations of the myocardium in HF rats. Transcriptome analysis showed that PNS upregulated 1749 genes and downregulated 1069 genes in the heart. Functional enrichment analysis demonstrated that the metabolic process was enriched among the DEGs. KEGG pathway analysis revealed that the PPAR signalling pathway was particularly involved in the protective function of PNS. The effects of PNS on the PPAR pathway were validated in vivo; PNS treatment effectively increased the expression of PPARα, RXRα, and PGC1α in rats with AMI-induced HF. In addition, PNS was shown to regulate the expression of downstream energy metabolism-related proteins. Interestingly, the addition of the PPARα inhibitor GW6471 abolished the beneficial effects of PNS. CONCLUSIONS: PNS exerts a cardioprotective function in a multicomponent and multitarget manner. The PPAR signalling pathway is one of the key pathways by which PNS protects against HF, and PPARα is a possible target for HF treatment.

The high expression of miR-31 in lung adenocarcinoma inhibits the malignancy of lung adenocarcinoma tumor stem cells.

Therapies for lung adenocarcinoma (LUAD) are mainly limited by drug resistance, metastasis or recurrence related to cancer stem cells (CSCs) with high proliferation and self-renewing. This research validated that miR-31 was over-expressed in LUAD by the analysis of generous clinical samples data. And the results of clinical data analysis showed that high expression of miR-31 was more common in patients with worse prognosis. The genes differentially expressed in LUAD tissues compared with normal tissues and A549CD133+ cells (LUAD CSCs) compared with A549 cells were separately screened from Gene Expression Profiling Interactive Analysis and GEO datasets. The target genes that may play a role in the regulation of lung adenocarcinoma was screened by comparison between the differential genes and the target genes of miR-31. The functional enrichment analysis of GO Biological Processes showed that the expression of target genes related to cell proliferation was increased, while the expression of target genes related to cell invasion and metastasis was decreased in LUAD tissues and A549CD133+ cells. The results suggested that miR-31 may have a significant inhibitory effect on the differentiation, invasion, metastasis and adhesion of LUAD CSCs, which was verified in vivo and in vitro experiments. Knock down of miR-31 accelerated xenograft tumor growth and liver metastasis in vivo. Likewise, the carcinogenicity, invasion and metastasis of A549CD133+ CSCs were promoted after miR-31 knockdown. The study validated that miR-31 was up regulated in LUAD and its expression may affect the survival time of patients with lung adenocarcinoma, which indicated that miR-31 may have potential value for diagnosis and prognosis of LUAD. However, the inhibitory effect of miR-31 on tumorigenesis, invasion and metastasis of lung adenocarcinoma CSCs suggested its complexity in the regulation of lung adenocarcinoma, which may be related to its extensive regulation of various target genes.

The multi-faceted role of retinoid X receptor in cardiovascular diseases.

Retinoid X receptors (RXRs) are members of ligand-dependent transcription factors whose effects on a diversity of cellular processes, including cellular proliferation, the immune response, and lipid and glucose metabolism. Knock out of RXRα causes a hypoplasia of the myocardium which is lethal during fetal life. In addition, the heart maintains a well-orchestrated balances in utilizing fatty acids (FAs) and other substrates to meet the high energy requirements. As the master transcriptional regulators of lipid metabolism, RXRs become particularly important for the energy needs of the heart. Accumulating evidence suggested that RXRs may exert direct beneficial effects in the heart both through heterodimerization with other nuclear receptors (NRs) and homodimerization, thus standing as suitable targets for treating in cardiovascular diseases. Although compounds that target RXRs are promising drugs, their use is limited by toxicity. A better understanding of the structural biology of RXRs in cardiovascular disease should enable the rational design of more selective nuclear receptor modulators to overcome these problems. Here, this review summarizes a brief overview of RXRs structure and versatility of RXR action in the control of cardiovascular diseases. And we also discussed the therapeutic potential of RXR ligand.

Autophagy as a novel insight into mechanism of Danqi pill against post-acute myocardial infarction heart failure.

ETHNOPHARMACOLOGICAL RELEVANCE: Danqi Pill, composed of the root of Salvia miltiorrhiza Bunge and the root of Panax notoginseng, is effective in the clinical treatment of myocardial ischemia in coronary heart diseases. A number of studies have shown that autophagy plays an essential role in cardiac function and energy metabolism, and disordered autophagy is associated with the progression of heart failure. However, the effect and mechanism of Danqi pill on autophagy have not been reported yet. AIM OF THE STUDY: This study aims to elucidate whether Danqi pill restores autophagy to protect against HF and its potential mechanism. MATERIALS AND METHODS: Left anterior descending ligation was established to induce an HF rat model, H2O2-stimulated H9C2 cells model was conducted to clarify the effects and potential mechanism of Danqi pill. In vivo, Danqi pill (1.5 g/kg) were orally administered for four weeks and Fenofibrate (10 mg/kg) was selected as a positive group. In vitro, Danqi pill (10-200 µg/mL) was pre-cultured for 24 h and co-cultured with H2O2 stimulation for 4 h. Importantly, transmission electron microscopy and fluorescence GPF-mRFP-LC3 reporter system were combined to monitor autophagy flux. Furtherly, we utilized Compound C, a specific AMPK inhibitor, to validate whether the autophagy was mediated by AMPK-TSC2-mTOR pathway. RESULTS: Danqi pill significantly improved cardiac function and myocardial injury in HF rats. Intriguingly, Danqi pill potently regulated autophagy mainly by promoting the formation of autophagosomes in vivo. Further results demonstrated that expressions of p-AMPK (P < 0.001) and p-TSC2 (P < 0.001) in cardiac tissue were upregulated by Danqi pill, accompanied with downregulation of p-mTOR (P < 0.01) and p-ULK1(P < 0.01). In parallel with the vivo experiment, in vitro study indicated that Danqi pill dramatically restored autophagy flux and regulated expressions of critical autophagy-related molecules. Finally, utilization of Compound C abrogated the effects of Danqi pill on autophagy flux and the expressions of p-TSC2 (P < 0.05), p-mTOR (P < 0.01) and p-ULK1 (P < 0.05). CONCLUSION: Danqi pill could improve cardiac function and protect against cardiomyocytes injury by restoring autophagy via regulating the AMPK-TSC2-mTOR signaling pathway.

TMT-based proteomics analysis of the anti-hepatocellular carcinoma effect of combined dihydroartemisinin and sorafenib.

Hepatocellular carcinoma (HCC), as the major primary liver cancer, is one of the most prevalent malignant diseases with a high mortality rate worldwide. Prior studies have demonstrated that dihydroartemisinin (DHA), the semisynthetic derivative of artemisinin, possesses anti-HCC activity. The multikinase inhibitor sorafenib has been approved for the treatment of HCC. However, the anti-HCC efficacy of DHA combined with sorafenib has not been reported. In this study, we confirmed the significantly enhanced anti-HCC efficacy of DHA in combination with sorafenib compared with that of each agent alone. Tandem Mass Tag (TMT) peptide labeling coupled with LC-MS/MS was used to quantify the proteins from the control, DHA, sorafenib, and DHA + sorafenib groups. In total, 532, 426, 628 differentially expressed proteins (fold change >1.20 or <0.83 and P-value <0.05) were determined by comparing DHA versus control, sorafenib versus control and DHA + sorafenib versus control groups, respectively. Moreover, optimized screening was performed, and 101 optimized differentially expressed proteins were identified. The results of functional analysis of the optimized differentially expressed proteins suggested that they were enriched in cell components such as membrane-bound vesicles, extracellular vesicles, and organelle lumens, and they were mainly involved in biological processes such as cellular component organization, response to stress, and response to chemicals; in addition, they were related to various molecular functions such as protein binding, chromatin binding and enzyme binding. KEGG pathway analysis showed that the optimized differentially expressed proteins were enriched in pyrimidine metabolism, RNA polymerase, base excision repair, and osteoclast differentiation. Protein-protein interaction (PPI) networks of some of the optimized upregulated proteins suggested that they might not only affect vitamin and fat digestion and absorption but may also be involved in tight junctions. In the PPI network, some of the optimized downregulated proteins were enriched in base excision repair, RNA polymerase, purine metabolism, pyrimidine metabolism and mucin type O-glycan biosynthesis. Overall, this research explored the anti-HCC efficacy of DHA combined with sorafenib by using the TMT-based quantitative proteomics technique and might facilitate the understanding of the related anti-HCC molecular mechanism.