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Tissue inhibitor matrix metalloproteinase 1 (TIMP1) has been reported to act as a tumor oncogene in colon cancer. However, little is known about the biological role of TIMP1 in gastric cancer. In this study, we found that the expression of TIMP1 in GC tissues was upregulated compared with the normal gastric tissues. TIMP1 was confirmed as a direct target of miR-6745 and silencing TIMP1 mimicked the effects of miR-6745 in GC cells. Further mechanism studies have shown that miR-6745 inhibits the Wnt/β-catenin pathway by targeting TIMP1, thereby inhibiting cell proliferation, migration and invasion. In addition, through the analysis of GC tissues, a negative correlation between miR-6745 and TIMP1 was found in 42 GC tissues. Our findings indicate that the miR-6745-TIMP1 axis regulates Wnt/βcatenin signaling and participates in GC tumorigenesis and provide a potential therapeutic target for preventing GC progression.

As gene delivery tools, lentiviral vectors (LV) have broad applications in chimeric antigen receptor therapy (CAR-T). Large-scale production of functional LV is limited by the adherent, serum-dependent nature of HEK293T cells used in the manufacturing. HEK293T adherent cells were adapted to suspension cells in a serum-free medium to establish large-scale processes for functional LV production in a stirred bioreactor without micro-carriers. The results showed that 293 T suspension was successfully cultivated in F media (293 CD05 medium and SMM293-TII with 1:1 volume ratio), and the cells retained the capacity for LV production. After cultivation in a 5.5 L bioreactor for 4 days, the cells produced 1.5 ± 0.3 × 107 TU/mL raw LV, and the lentiviral transduction efficiency was 48.6 ± 2.8% in T Cells. The yield of LV equaled to the previous shake flask. The critical process steps were completed to enable a large-scale LV production process. Besides, a cryopreservation solution was developed to reduce protein involvement, avoid cell grafting and reduce process cost. The process is cost-effective and easy to scale up production, which is expected to be highly competitive.

Liraglutide is an analogue of human glucagon-like peptide-1 (GLP-1), an endogenous intestinal hormone which play essential roles in the regulation of glycolipid metabolism. To investigate the role of lactic acid bacteria (LAB) in the lipid-lowering effect of liraglutide, forty mice were divided into normal saline-treated basal diet (NFD), normal saline-treated high-fat food (HFD), 10.0 mg/kg/d simvastatin-treated HFD (SIM + HFD), 200 and 400 μg/kg/d liraglutide-treated HFD (LL + HFD and HL + HFD) groups for 5 weeks. 16S rDNA sequencing, real-time quantitative PCR and western blot were used to detected changes of intestinal flora, cholesterol 7α-hydroxylase (CYP7A1), LDL-receptor (LDLR) and 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR). Results showed that liraglutide could up-regulate CYP7A1 and LDLR, whereas down-regulate HMGCR. Besides, liraglutide enhance the abundance of lactobacillaceae in gut of hyperlipidemic mice and increase the bile tolerance ability of LAB by up-regulating bile salt hydrolases, and the lysate of liraglutide-sensitive LAB could also directly down-regulate HMGCR, the key enzyme in cholesterol synthesis, and inhibit hepatocyte steatosis. These findings might provide new theoretical guidance for clinical application of liraglutide and threw a light on research and development of anti-obesity, hypolipidemic and cholesterol-lowering drugs or functional foods.

Guaipyridine alkaloids (-)-rupestine D, (-)-guaipyridine, (-)-epiguaipyridine, and (-)-cananodine together with two stereoisomers 8-epi-rupestine D and 5-epi-cananodine were synthesized enantioselectively from readily available citronellol. The key steps in this synthesis are (i) intermolecular opening of a trisubstituted epoxide for the formation of a chiral center at C-8; (ii) ring-closing metathesis for the construction of a seven-membered carbocyclic ring; and (iii) biomimetic cyclization of a 1,5-dicarbonyl compound for the construction of a pyridine-fused bicyclic skeleton.

Context Peucedanol is a major extract of Peucedanum japonicum Thunb. (Apiaceae) roots, which is a commonly used herb in paediatrics. Its interaction with cytochrome P450 enzymes (CYP450s) would lead to adverse effects or even failure of therapy. Objective The interaction between peucedanol and CYP450s was investigated. Materials and methods Peucedanol (0, 2.5, 5, 10, 25, 50, and 100 μM) was incubated with eight human liver CYP isoforms (CYP1A2, 2A6, 3A4, 2C8, 2C9, 2C19, 2D6, and 2E1), in pooled human liver microsomes (HLMs) for 30 min with specific inhibitors as positive controls and untreated HLMs as negative controls. The enzyme kinetics and time-dependent study (0, 5, 10, 15, and 30 min) were performed to obtain corresponding parameters in vitro. Results Peucedanol significantly inhibited the activity of CYP1A2, 2D6, and 3A4 in a dose-dependent manner with IC50 values of 6.03, 13.57, and 7.58 μM, respectively. Peucedanol served as a non-competitive inhibitor of CYP3A4 with a Ki value of 4.07 μM and a competitive inhibitor of CYP1A2 and 2D6 with a Ki values of 3.39 and 6.77 μM, respectively. Moreover, the inhibition of CYP3A4 was time-dependent with the Ki/Kinact value of 5.44/0.046 min/μM. Discussion and conclusions In vitro inhibitory effect of peucedanol on the activity of CYP1A2, 2A6, and 3A4 was reported in this study. As these CYPs are involved in the metabolism of various drugs, these results implied potential drug-drug interactions between peucedanol and drugs metabolized by CYP1A2, 2D6, and 3A4, which needs further in vivo validation.

There is evidence that major depressive disorder (MDD) is related to serum lipid level alterations. However, the neural correlates underlying this association remain poorly understood. Forty-nine patients with MDD and fifty healthy controls (HCs) underwent structural, resting-state functional and diffusion magnetic resonance imaging scans. Voxel-based morphometry, functional connectivity (FC) and tract-based spatial statistics analyses were performed to assess brain structure and function, respectively. Blood samples were collected to measure serum levels of lipid variables including total cholesterol, triglyceride and high density lipoprotein cholesterol (HDL-C). Correlation and mediation analyses were conducted to investigate the associations of serum lipid levels with brain imaging measures in MDD patients and HCs, respectively. We found that the serum HDL-C level in MDD patients was lower than that in HCs. The lower serum HDL-C level was associated with lower gray matter volume (GMV) in ventromedial prefrontal cortex (VMPFC), higher within-network FC of the default mode network, and lower micro-structural integrity in multiple white matter regions in MDD patients. Moreover, the within-default mode network FC mediated the relationship between GMV in VMPFC and serum HDL-C level; white matter integrity in genu of corpus callosum mediated the relationship between serum HDL-C level and depressive symptom severity. However, we did not observe any correlations between serum lipids and brain imaging parameters in HCs. These findings help to identify neural correlates underlying the association between depression and serum HDL-C change, which may provide new insight into intervention, treatment and prevention of depression from the perspective of regulating serum lipids.

Arterial marker genes EphrinB2 and HEY2 are essential for cardiovascular development and postnatal neovascularization. Our previous study confirmed that E2F1 could activate the transcription of EphrinB2 and HEY2 in human mesenchymal stem cells; however, the detailed mechanism has not been resolved yet. In this study, we focused on the interaction between E2F1 and DNMT3A, a de novo DNA methyltransferase, on regulating the expression of EphrinB2 and HEY2, and explored the potential mechanisms. Gain- and loss-of-function experiments implicated the positive effect of E2F1 on the expression of EphrinB2 and HEY2 and tube formation in human umbilical artery endothelial cells. Accumulation of DNMT3A decreased the levels of EphrinB2 and HEY2, and impaired tube formation induced by E2F1, while inhibiting DNMT3A by RNA interference augmented their expression and angiogenesis in E2F1-trasfected cells. We then asked whether the low expressions of EphrinB2 and HEY2 induced by DNMT3A are related to the methylation status of their promoters. Surprisingly, the methylation status of the CpG islands in the promoter region was not significantly affected by overexpression of exogenous DNMT3A. Furthermore, the interaction between E2F1 and DNMT3A was confirmed by co-immunoprecipitation. DNMT3A could inhibit the transcription of EphrinB2 and HEY2 promoters by affecting the binding of E2F1 to its recognition sequences as revealed by luciferase reporter assay and chromatin immunoprecipitation. These results identified a novel mechanism underlying the cooperation of DNMT3A with E2F1 on regulating target gene expression, and revealed their roles in the angiogenic process.

STING is essential for control of infections and for tumor immunosurveillance, but it can also drive pathological inflammation. STING resides on the endoplasmic reticulum (ER) and traffics following stimulation to the ERGIC/Golgi, where signaling occurs. Although STING ER exit is the rate-limiting step in STING signaling, the mechanism that drives this process is not understood. Here we identify STEEP as a positive regulator of STING signaling. STEEP was associated with STING and promoted trafficking from the ER. This was mediated through stimulation of phosphatidylinositol-3-phosphate (PtdIns(3)P) production and ER membrane curvature formation, thus inducing COPII-mediated ER-to-Golgi trafficking of STING. Depletion of STEEP impaired STING-driven gene expression in response to virus infection in brain tissue and in cells from patients with STING-associated diseases. Interestingly, STING gain-of-function mutants from patients interacted strongly with STEEP, leading to increased ER PtdIns(3)P levels and membrane curvature. Thus, STEEP enables STING signaling by promoting ER exit. STING ER exit is the rate-limiting step in STING signaling, but the mechanism that drives this process is not understood. Paludan and colleagues identify CxORF56, called STEEP here, as a positive regulator of STING signaling.

Background:Scutellariae Radix (Huangqin) is commonly processed into 3 products for different clinical applications. However, a simple analytical method for quality control has rarely been reported to quickly estimate the degree of processing Huangqin or distinguish differently processed products or unqualified Huangqin products.Objective:To study a new strategy for quality control in the processing practice of Huangqin.Methods:Seven kinds of flavonoids that mainly exist in Huangqin were determined by HPLC-DAD. Chromatographic fingerprints were established to study the variation and discipline of the 3 processed products of Huangqin. PCA and OPLS-DA were used to classify differently processed products of Huangqin.Results:The results showed that baicalin and wogonoside were the main components in the crude and the alcohol Huangqin herb while baicalein and wogonin mainly existed in carbonized Huangqin. The results of mathematical statistics revealed that the processing techniques can make the quality of medicinal materials more uniform.Conclusion:This multivariate monitoring strategy is suitable for quality control in the processing of Huangqin.

Cisplatin resistance has been a major factor limiting its clinical use as a chemotherapy drug. The present study aimed to investigate whether SET and MYND domain-containing protein 3 (SMYD3), a histone methyltransferase closely associated with tumors can affect the sensitivity of tumors to cisplatin chemotherapy. Real time-qPCR, western blotting, the luciferase reporter, MTT and clonogenic assays were performed to detect the effects of SMYD3 on the chemotherapy capacity of cisplatin. In the present study, SMYD3 exhibited different expression patterns in MCF-7 and T47D breast cancer cells. In addition, this differential expression was associated with tumor cell resistance to cisplatin. Furthermore, SMYD3 knockdown following small interfering RNA transfection increased cisplatin sensitivity, whereas SMYD3 overexpression decreased cisplatin sensitivity. In addition, SMYD3 knockdown synergistically enhanced cisplatin-induced cell apoptosis. SMYD3 expression was downregulated during cisplatin treatment. In addition, transcriptional regulatory activities of SMYD3 3'-untranslated region were also downregulated. These results suggested that SMYD3 may affect cell sensitivity to cisplatin and participate in the development of cisplatin resistance, which is a process that may involve microRNA-124-mediated regulation.

The development of an effective pharmacological countermeasure is needed to reduce the morbidity and mortality in high-dose ionizing radiation-induced acute damage. Genistein has shown bioactivity in alleviating radiation damage and is currently synthesized by chemosynthetic methods. Due to concerns about chemical residues and high costs, the clinical application of genistein is still a major challenge. In this study, we aimed to establish an efficient method for the extraction of genistein from Fructus sophorae. The effects of extracted genistein (FSGen) on preventing intestinal injury from radiation were further investigated in this study. C57/BL mice were exposed to 7.5 Gy whole body irradiation with and without FSGen treatments. Histological analysis demonstrated significant structural and functional restitution of the intestine and bone marrow in FSGen-pretreated cohorts after irradiation. Through mRNA expression, protein expression, and small interfering RNA analyses, we demonstrated that FSGen protects IEC-6 cells against radiation damage by upregulating the Rassf1a and Ercc1 genes to effectively attenuate DNA irradiation damage. Together, our data established an effective method to extract genistein from the Fructus sophorae plant with high purity, and validated the beneficial roles of the FSGen in protecting the radiation damage. These results promise the future applications of Fructus sophorae extracted genistein in the protection of radiation related damages.

Gene therapy is considered a potential treatment for Duchenne muscular dystrophy (DMD). Researchers have been working on this for many years to find effective therapeutic targets. Here, we found that MRTF-A (myocardin-related transcription factor A) could activate the transcription of L-type Ca2+-channel-related protein CACNA1S (calcium voltage-gated channel subunit alpha1 S) by binding to the CarG box in the promoter of CACNA1S. However, increased phosphorylation and decreased expression of MRTF-A were observed, along with the expression of CACNA1S reduced in mdx mice. Further, the decreased expression and increased phosphorylation of MRTF-A could inhibit the release of Ca2+ via CACNA1S. Therefore, MRTF-A may be a potential molecular target for the diagnosis and treatment of DMD.

Background Heparinase I from Pedobacter heparinus (Ph-HepI), which specifically cleaves heparin and heparan sulfate, is one of the most extensively studied glycosaminoglycan lyases. Enzymatic degradation of heparin by heparin lyases not only largely facilitates heparin structural analysis but also showed great potential to produce low-molecular-weight heparin (LMWH) in an environmentally friendly way. However, industrial applications of Ph-HepI have been limited by their poor yield and enzyme activity. In this work, we improve the specific enzyme activity of Ph-HepI based on homology modeling, multiple sequence alignment, molecular docking and site-directed mutagenesis. Results Three mutations (S169D, A259D, S169D/A259D) exhibited a 50.18, 40.43, and 122.05% increase in the specific enzyme activity and a 91.67, 108.33, and 75% increase in the yield, respectively. The catalytic efficiencies (kcat/Km) of the mutanted enzymes S169D, A259D, and S169D/A259D were higher than those of the wild-type enzyme by 275, 164, and 406%, respectively. Mass spectrometry and activity detection showed the enzyme degradation products were in line with the standards of the European Pharmacopoeia. Protein structure analysis showed that hydrogen bonds and ionic bonds were important factors for improving specific enzyme activity and yield. Conclusions We found that the mutant S169D/A259D had more industrial application value than the wild-type enzyme due to molecular modifications. Our results provide a new strategy to increase the catalytic efficiency of other heparinases. Electronic supplementary material The online version of this article (10.1186/s12896-019-0553-3) contains supplementary material, which is available to authorized users.

Thermostable and halotolerant GH11 xylanases are highly desirable in various industrial applications but they were rarely characterized. In this study, we firstly revealed the genetic diversity of GH11 xylanases from the metagenomic DNA of a saline-alkaline soil by constructing a gene fragment library. Twelve distinct xylanase fragment sequences were identified, which shared 62–96% identity to known GH11 xylanases. Among them, Xyn22 with the highest relative abundance was cloned and found to exhibit excellent halotolerance. Xyn22 retained 80 and 58% relative activity in the presence of 3 and 5 M NaCl, respectively, and had no loss of activity after incubation in 3 M NaCl at 37 °C for 1 h. Additionally, the acidic amino acid residues E137 and E139 on the protein's surface were revealed to cooperatively play important roles in its excellent halotolerance and halostabilty. Furthermore, the thermostability of Xyn22 was significantly improved without compromising halotolerance by introducing an aromatic interaction between Y48 and F53. In conclusion, this study not only provides a thermostable and halotolerant GH11 xylanase, but also reveals the key roles of surface-exposed amino acids in its thermostability and halotolerance, which could be used for further molecular modification of other xylanases and similar enzymes.

Tumor targeting agents are being developed for early tumor detection and therapeutics. We previously identified the peptide SNFYMPL (SNF*) and demonstrated its specific binding to human esophageal specimens of high-grade dysplasia (HGD) and adenocarcinoma with imaging ex vivo. Here, we aim to identify the target for this peptide and investigate its potential applications in imaging and drug delivery. With SNF* conjugated affinity chromatography, mass spectrum, Western blot, enzyme-linked immunosorbent assay (ELISA), and molecular docking, we found that the epithelial cell adhesion molecule (EpCAM) was the potential target of SNF*. Next, we showed that FITC-labeled SNF* (SNF*-FITC) colocalized with EpCAM antibody on the surface of esophageal adenocarcinoma cells OE33, and SNF*-FITC binding patterns significantly changed after EpCAM knockdown or exogenous EpCAM transfection. With the data from TCGA, we demonstrated that EpCAM was overexpressed in 17 types of cancers. Using colon and gastric adenocarcinoma cells and tissues as examples, we found that SNF*-FITC bound in a pattern was colocalized with EpCAM antibody, and the SNF* binding did not upregulate the EpCAM downstream Wnt signals. Subsequently, we conjugated SNF* with our previously constructed poly(histidine)-PEG/DSPE copolymer micelles. SNF* labeling significantly improved the micelle binding with colon and gastric adenocarcinoma cells in vitro, and enhanced the antitumor effects and decreased the toxicities of the micelles in vivo. In conclusion, we identified and validated SNF* as a specific peptide for EpCAM. The future potential use of SNF* peptide in multiple tumor surveillance and tumor-targeted therapeutics was demonstrated.

N-Acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) is a critical negative regulator of fibrosis development in the liver. However, its extremely short half-life in vivo greatly compromises its potential applications. Here, we report an Ac-SDKP analog peptide with d-amino acid replacement (Ac-SDD KD P). The stability of Ac-SDD KD P and its prevention of liver fibrosis were investigated in vitro and in vivo. The stabilities of Ac-SDKP and Ac-SDD KD P exposed to angiotensin-1-converting enzyme (ACE) and their half-lives in rats and human sera were determined by high-performance liquid chromatography. The inhibitory effects of Ac-SDKP and Ac-SDD KD P on the proliferation and activation of hepatic stellate cells (HSC-T6) were evaluated using the Cell Counting Kit-8, Western blotting, reverse transcription quantitative polymerase chain reaction, and immunofluorescence assays. Finally, the protective effects of Ac-SDKP and Ac-SDD KD P on carbon tetrachloride (CCl4 )-induced liver fibrosis in rats were compared. d-Amino acid replacement significantly enhanced the stability of the peptide to ACE and prolonged the half-life of Ac-SDKP in rats and human sera. The Ac-SDKP-mediated inhibition of HSC-T6 cell proliferation was well preserved, and Ac-SDD KD P exerted inhibitory effects comparable to Ac-SDKP on α-smooth muscle actin (α-SMA), collagen I and III expression, and phosphorylated-Smad-2 expression. After intraperitoneal (i.p.) administration, Ac-SDD KD P exhibited significantly greater protection than Ac-SDKP against CCl4 -induced liver fibrosis in rats. The serum alanine aminotransferase, aspartate aminotransferase, albumin, and total protein levels of the Ac-SDD KD P-treated rats were significantly lower than those of the Ac-SDKP-treated rats. α-SMA, CD45, and collagen I and III expression, as well as Smad-2 phosphorylation were significantly attenuated in the livers of the Ac-SDD KD P-treated rats compared to those of the Ac-SDKP-treated rats. Furthermore, we showed that the Ac-SDD KD P concentration in the rat liver increased to a physiological level of 60 min after i.p. administration, although i.p. administration of Ac-SDKP failed to enhance the peptide concentration in the rat liver. Our findings indicate that d-amino acid replacement is a simple and effective method to enhance the stability of Ac-SDKP. Ac-SDD KD P represents potential application of Ac-SDKP in fibrosis treatment and provides a new potential treatment strategy for liver fibrosis. © 2019 IUBMB Life, 71(9):1302-1312, 2019.

Multiple articles have confirmed that an imbalance of the intestinal microbiota is closely related to aberrant immune responses of the intestines and to the pathogenesis of inflammatory bowel diseases (IBDs). Probiotic strains have been identified for the treatment and prevention of IBDs. The aim of this study was to screen a new probiotic strain with anti-inflammatory activity and investigate the potential mechanisms underlying its activity. We identified a new probiotic strain, L. lactis ML2018, that has anti-inflammatory properties and was isolated from traditional fermented food. In an in vitro experiment, L. lactis ML2018 prevented the release of nitric oxide (NO) and the production of inflammatory factors induced by lipopolysaccharides (LPS) in RAW264.7 cells. The in vivo anti-inflammatory effects of L. lactis ML2018 were evaluated using a dextran sulfate sodium (DSS)-induced animal model of colitis. Oral administration of L. lactis ML2018 significantly ameliorated colitis induced by DSS, which included preventing a decrease in body weight, shortening of the colon length and apoptosis of epithelial cells. L. lactis ML2018 could inhibit DSS-induced intestinal inflammation by preventing the overproduction of proinflammatory factors, suppressing the infiltration of macrophages, controlling the fibrosis, improving the integrity of the intestinal epithelial barrier and upregulating the concentrations of short-chain fatty acids (SCFAs). Moreover, L. lactis ML2018 could prevent inflammation by inhibiting the activation of the NF-κB and MAPK signaling pathways. These data suggest that L. lactis ML2018 could have therapeutic potential for treating IBDs.

Myocardin is an important factor that regulates cardiac hypertrophy, and its activity can be regulated by GATA4. However, the molecular mechanism of the above process remains unclear. This paper presents three kinds of possible molecular mechanisms of GATA4 inhibiting myocardin activity in the process of cardiac hypertrophy. First, a competitive combination of GATA4 and SRF with myocardin could reduce the formation of the myocardin-SRF-CarG box complex when GATA4 was overexpressed. Second, overexpression of GATA4 could inhibit the combination of myocardin and p300 and downregulate acetylated myocardin levels. Finally, GATA4 could upregulate the phosphorylation of myocardin protein upon activation of the ERK pathway. These findings may provide insight into the function of GATA4 and myocardin in the occurrence and development of cardiac hypertrophy.

Lung cancer remains one of the most aggressive human malignancies with a low survival rate. Hyperoside (quercetin-3-O-β- d -galactopyranoside) is a flavonol glycoside with an anti-cancer activity. The microRNA-let-7 was widely regarded as a tumor suppressor in human tumors. Here, we investigated the role of hyperoside and let-7a-5p on the lung cancer cell proliferation, cell cycle and apoptosis in A549 cells in vitro. Our results showed that hyperoside could inhibit the proliferation of A549 cells through inducing apoptosis and G1/S phase arrest. Let-7a-5p could inhibit the proliferation of A549 cells via inhibiting the process of G1/S phase. Additionally, hyperoside and let-7a-5p had a synergetic effect on suppressing the proliferation of A549 cells; microRNA-let-7a-5p directly regulated the expression of CCND1 in A549 cells. Our study illustrated that hyperoside and microRNA-let7a-5p might provide a synergistic effect on anti-cancer, which may provide a new idea for lung cancer treatment.

Two glycoside hydrolases encoded by the mannan utilization gene cluster of alkaliphilic Bacillus sp. N16-5 were studied. The recombinant Gal27A (rGal27A) hydrolyzed both galactomannans and oligo-galactomannans to release galactose, while the recombinant Man113A (rMan113A) showed poor activity toward galactomannans, but it hydrolyzed manno-oligosaccharides to release mannose and mannobiose. rGal27A showed synergistic interactions with rMan113A and recombinant β-mannanase ManA (rManA), which is also from Bacillus sp. N16-5, in galactomannan degradation. The synergy degree of rGal27A and rManA on hydrolysis of locust bean gum and guar gum was 1.13 and 2.21, respectively, and that of rGal27A and rMan113A reached 2.00 and 2.68. The main products of galactomannan hydrolyzed by rGal27A and rManA simultaneously were galactose, mannose, mannobiose, and mannotriose, while those of galactomannan hydrolyzed by rGal27A and rMan113A were galactose and mannose. The yields of mannose, mannobiose, and mannotriose dramatically increased compared with the hydrolysis in the presence of rManA or rMan113A alone.

Crocins, as a kind of water-soluble carotenoid pigment, are a series of ester compounds formed from crocetin and gentibiose or glucose, and mainly distributed among Crocus sativus L. (CSL), Gardenia jasminoides Ellis. (GJE). Crocins exhibit a wide range of pharmacological effects on neurodegeneration, cardiovascular disease, cerebrovascular disease, depression, liver disease, arthritis, tumor, diabetes, etc. This review systematically discussed the pharmacologic study of crocins in the aspect of structural characteristic and pharmacokinetics, and summarized the mechanism of treating disease. It summarized the abundant research of crocins from 1984 to 2020 based on the above aspects, which provide a reference for the deeply development and application of crocins.

Mannan is an important renewable resource whose backbone can be hydrolyzed by β-mannanases to generate manno-oligosaccharides of various sizes. Only a few glycoside hydrolase (GH) 113 family β-mannanases have been functionally and structurally characterize. Here, we report the function and structure of a novel GH113 β-mannanase from Bacillus sp. N16-5 (BaMan113A). BaMan113A exhibits a substrate preference toward manno-oligosaccharides and releases mannose and mannobiose as main hydrolytic products. The crystal structure of BaMan113A suggest that the enzyme shows a semi-enclosed substrate-binding cleft and the amino acids surrounding the +2 subsite form a steric barrier to terminate the substrate-binding tunnel. Based on these structural features, we conducted mutagenesis to engineer BaMan113A to remove the steric hindrance of the substrate-binding tunnel. We found that F101E and N236Y variants exhibit increased specific activity toward mannans comparing to the wild-type enzyme. Meanwhile, the product profiles of these two variants toward polysaccharides changed from mannose to a series of manno-oligosaccharides. The crystal structure of variant N236Y was also determined to illustrate the molecular basis underlying the mutation. In conclusion, we report the functional and structural features of a novel GH113 β-mannanase, and successfully improved its endo-acting activity by using structure-based engineering.

Background: Codonopsis Radix (CR) as the tonic Chinese herbis is good at benefiting Qi and promoting saliva, while rice-processed CR (RCR) can alleviate the original dryness and more specialize in invigorating the spleen and stopping diarrhea. Despite CR and RCR are long-standing medicines commonly used in clinic, information on their differences in substance basis is limited, let alone dynamic changes of their main chemical components. Given this, a more comprehensive understanding of processing principles in CR is needed.Methods: In this study, a strategy for rapid separation and identification of chemical constituents in crude CR and RCR was developed by HPLC-Q-TOF-MS/MS. Moreover, chemical constituents (lobetyolin and atractylenolide III) with significant content change were quantitatively determined by HPLC-QQQ-MS, 5-hydroxymethylfurfural (5-HMF) was quantified by HPLC, and codonopsis pilosula polysaccharides (CPPS) was measured by phenol-sulphuric acid method.Results: A total of 28 compounds were identified from CR and RCR, including 12 organic acids, 6 phenylpropanoids, 5 hexanol glycosides and hexylene glycosides, 2 polyacetylenes, 1 alcohol compound, 1 sesquiterpene and 1 aldehyde. In order to explore the dynamic changes from raw CR to processed RCR, the content of representative components in several samples collected during processing was determined. The quantitative results indicated that the contents of lobetyolin, atractylenolide III and CPPS were lower in RCR than these in CR, and 5-HMF was newly formed in processed products, which manifested chemical composition change before and after processing. Conclusions: Processing principles and dynamic changes of CR were preliminarily elucidated. Lobetyolin, atractylenolide III, 5-HMF and CPPS were recommended as quality markers to distinguish CR from RCR. Since the established method is rapid, accurate and readily applicable, it will be helpful for quality control and identification of crude CR and RCR.

Inhibition of Poly(ADP-ribose) polymerase (PARP) is effective for breast cancer susceptibility genes 1 (BRCA1)-deficient breast cancers. Although hormones play critical roles on the occurrence as well as being used in conventional therapies of breast cancer, their impacts on PARP-targeted therapy have been poorly addressed. Here, we showed that addition of estrogen could enhance the cytotoxicity of PARP inhibitors on estrogen receptor (ER)-positive breast cancer cells, causing significant suppression of cell growth. Further analysis revealed that the impact was due to estrogen's stimulating the production of nitric oxide (NO), which could be abrogated when blocking NO formation. Moreover, the effect of estrogen can be resembled by two exogenous nitric oxide donors (SNAP and GSNO). Using ER-negative cell line MDA-MB231, estrogen could not enhance the cell killing of PARP inhibitors any more, but addition of NO donors re-established the enhancing effects. The increased NO level led to accumulation of DNA double strand breaks (DSBs) based on the formation of H2AX foci. Consistent with earlier studies, we demonstrated that NO suppressed the expression of BRCA1, a key player involved in DSB recombination repair. Taken together, these data reveal an important role of estrogen on the treatment of PARP inhibitors, which may affect its clinical treatment and should be considered in precision therapies for ER-positive and negative cancers.

BackgroundIn order to explore the molecular mechanism of cardiomyocyte-dependent myocardial gene expression and cardiomyocyte differentiation in cardiac hypertrophy, and to provide new insights for cardiac hypertrophy.MethodsCardiac myocytes were isolated from day 1-3 Sprague-Dawley rat pups. Real time quantitative PCR, western blot and immunocytochemistry Assay were used to detect the expression and localization of related genes. CO-IP was used to detect direct protein interactions between Myocardin and STAT3. Luciferase reporter assay and chromatin immunoprecipitation were used to detect the binding of Myocardin to the promoter of a downstream target gene. Microinjection of zebrafish embryos was used to examine the effects of STAT3 and Myocardin interactions on cardiac development in vivoResultsThe N-terminus of STAT3 directly binds to the basic domain of myocardin and inhibits the transcriptional activity of Myocardin-mediated cardiac-specific genes ANF and α-actinin, thereby inhibiting their expression, and further inhibit myocardin-mediated cardiac hypertrophy in vivo.ConclusionsIn summary, our report states that signal transduction and transcriptional activation factor 3 (STAT3) are inhibitors of the major cardiac hypertrophic transcription factor Myocardiin, which is required for cardiomyocyte differentiation. The STAT3-cardiacin interaction identified nuclear hormone receptor-mediated and cardiac-specific gene-regulated convergence sites and suggested a possible mechanism for cardioprotective effects.

Antimicrobial peptides have been attracting increasing attention for their multiple beneficial effects. In present study, a novel AMP with a molecular weight of 1875.5 Da, was identified from the genome of Lactobacillus casei HZ1. The peptide, which was named as LHH1 was comprised of 16 amino acid residues, and its α-helix content was 95.34% when dissolved in 30 mM SDS. LHH1 exhibited a broad range of antimicrobial activities against Gram-positive bacteria and fungus. It could effectively inhibit Staphylococcus aureus with a minimum inhibitory concentration of 3.5 μM and showed a low hemolytic activity. The scanning electron microscope, confocal laser scanning microscope and flow cytometry results showed that LHH1 exerted its antibacterial activity by damaging the cell membrane of Staphylococcus aureus. Meanwhile, LHH1 also exhibited anti-cancer cell activities against several cancer cells via breaking the cell membrane of MGC803, HCT116 and C666-1 cancer cells.

Background Zhi-Zi-Da-Huang decoction (ZZDHD) is a famous Traditional Chinese Medicine decoction due to its therapeutic effects on clinical hepatobiliary disorders. ZZDHD is composed of Gardeniae Fructus, Rhei Radix et Rhizoma, Fructus Aurantii Immaturus, and Sojae Semen Praeparatum. With the development of current technology, dispensing granules have been widely used for convenience. However, limited research has been conducted to determine differences in the chemical compounds between dispensing granules and traditional decoction. Methods A strategy based on UPLC-qTOF-MS/MS and UPLC-DAD was established to quantitatively and qualitatively analyze the chemical compounds present in single- and co-boiled ZZDHD. First, we utilized UPLC-qTOF-MS/MS to identify the compounds in single- and co-boiled ZZDHD. Then, 15 compounds were quantitatively analyzed in ZZDHD by UPLC-DAD. Finally, fingerprint and chemometric analyses were adopted to evaluate the difference between single- and co-boiled ZZDHD. Results One hundred and forty-seven compounds were authenticated or determined according to their retention behaviors, mass mensuration, and characteristic fragment ions. In addition, 15 analytes of single- and co-boiled ZZDHD indicated a significant difference in the content of many compounds, including geniposide (G2), emodin, chrysophanol, daidzin, and crocin-I. The similarities of single and co-boiled ZZDHD were higher than 0.99, as evaluated by fingerprinting; 30 common peaks were evaluated by the chemometric analysis, which found 13 important variable values (VIP > 1) including C3, A3, crocetin, daidzin, C5, physcion, G2, genipin-1-O-β-D-gentiobioside (G1), chrysophanol, emodin, A2, A5, and C6. Conclusions This study provided an experimental basis for explaining the rationality of single-and co-boiled ZZDHD, and a study model for comparing the chemical substances between traditional decoctions and dispensing granules.

Probiotics are thought to have immunomodulatory functions, improve inflammatory disorders and treat inflammatory bowel disease (IBD). Here, we screened a new probiotic strain with anti-inflammatory activity and investigated its effect on the immune cell response and histone acetylation. Lactobacillus casei (L. casei) LH23 inhibited the production of nitric oxide and inflammatory factors induced by lipopolysaccharides in RAW264.7 cells, which was associated with inhibiting the over-activation of the JNK/p38 signaling pathway. Furthermore, L. casei LH23 can significantly ameliorate dextran sulfate sodium (DSS)-induced mouse colitis in vivo by reducing numbers of macrophages (CD11b+F4/80+) and their secreted inflammatory cytokines. Myeloperoxidase activity was also decreased in mice treated with LH23. The administration of L. casei LH23 induced the increase of CD3+CD4+CD25+ regulatory T cells among mesenteric lymph nodes. Meanwhile, LH23 treatment could augment short chain fatty acid contents. Importantly, we reported here for the first time that DSS treatment significantly decreased the level of histone H3K9 acetylation, while supplementation of L. casei LH23 restored the level of histone H3K9 acetylation in colon tissues. These data suggest that L. casei LH23 may have been beneficial for preventing and treating IBD.

Fumonisins have posed hazardous threat to human and animal health worldwide. Enzymatic degradation is a desirable detoxification approach but is severely hindered by serious shortage of detoxification enzymes. After mining enzymes by bioinformatics analysis, a novel carboxylesterase FumDSB from Sphingomonadales bacterium was expressed in Escherichia coli, and confirmed to catalyze fumonisin B1 to produce hydrolyzed fumonisin B1 by liquid chromatography mass spectrometry for the first time. FumDSB showed high sequence novelty, sharing only ~34% sequence identity with three reported fumonisin detoxification carboxylesterases. Besides, FumDSB displayed its high degrading activity at 30–40 °C within a broad pH range from 6.0 to 9.0, which is perfectly suitable to be used in animal physiological condition. It also exhibited excellent pH stability and moderate thermostability. This study provides a FB1 detoxification carboxylesterase which could be further used as a potential food and feed additive.

Intestinal mucosal barriers help the body resist many intestinal inflammatory diseases, such as inflammatory bowel disease (IBD). In this study, we identified a novel bacterium promoting the repair of intestinal mucosa and investigated the potential mechanisms underlying its activity. Culture supernatant of Bacillus subtilis RZ001 upregulated the expression of mucin 2 (MUC2) and tight junction (TJ) proteins in HT-29 cells in vitro. Oral administration of B. subtilis RZ001 may have significantly reduced symptoms such as the dextran sulfate sodium (DSS)-induced decrease in body weight, shortening of colon length and overproduction of proinflammatory factors. The number of goblet cells and levels of MUC2 and TJ proteins were significantly increased in adult mice fed with B. subtilis RZ001. B. subtilis RZ001 cells upregulated the levels of MUC2 in the intestinal organoids. Furthermore, culture supernatant of B. subtilis RZ001 could suppress the Notch signalling pathway and activate the expression of atonal homolog 1 (Atoh1). The transcription factor Atoh1 is required for intestinal secretory cell differentiation and activates transcription of MUC2 via binding to E-boxes on the MUC2 promoter. Taken together, B. subtilis strain RZ001 has the potential for treating IBD. The present study is helpful to elucidate the mechanisms of B. subtilis action.

Chitooligosaccharides (COS) have a variety of biological activities due to their positively charged amino groups. Studies have shown that COS have antidiabetic effects, but their molecular mechanism has not been fully elucidated. The present study confirmed that COS can reduce hyperglycemia and hyperlipidemia, prevent obesity, and enhance histological changes in the livers of mice with type 2 diabetes mellitus (T2DM). Additionally, treatment with COS can modulate the composition of the gut microbiota in the colon by altering the abundance of Firmicutes, Bacteroidetes, and Proteobacteria. Furthermore, in T2DM mice, treatment with COS can upregulate the cholesterol-degrading enzymes cholesterol 7-alpha-hydroxylase (CYP7A1) and incretin glucagon-like peptide 1 (GLP-1) while specifically inhibiting the transcription and expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), the key enzyme in cholesterol synthesis. Furthermore, using an oleic acid-induced hepatocyte steatosis model, we found that HMGCR can be directly transactivated by SET and MYND domain containing 3 (SMYD3), a transcriptional regulator, via 5&prime, CCCTCC-3&prime, element in the promoter. Overexpression of SMYD3 can suppress the inhibitory effect of COS on HMGCR, and COS might regulate HMGCR by inhibiting SMYD3, thereby exerting hypolipidemic functions. To the best of our knowledge, this study is the first to illustrate that COS mediate glucose and lipid metabolism disorders by regulating gut microbiota and SMYD3-mediated signaling pathways.

Treatments on neoplastic diseases and cancer using genotoxic drugs often cause long-term health problems related to premature aging. The underlying mechanism is poorly understood. Based on the study of a long-lasting senescence-like growth arrest (10-12 weeks) of human dermal fibroblasts induced by psoralen plus UVA (PUVA) treatment, we here revealed that slowly repaired bulky DNA damages can serve as a “molecular scar” leading to reduced cell proliferation through persistent endogenous production of reactive oxygen species (ROS) that caused accelerated telomere erosion. The elevated levels of ROS were the results of mitochondrial dysfunction and the activation of NADPH oxidase (NOX). A combined inhibition of DNA-PK and PARP1 could suppress the level of ROS. Together with a reduced expression level of BRCA1 as well as the upregulation of PP2A and 53BP1, these data suggest that the NHEJ repair of DNA double-strand breaks may be the initial trigger of metabolic changes leading to ROS production. Further study showed that stimulation of the pentose phosphate pathway played an important role for NOX activation, and ROS could be efficiently suppressed by modulating the NADP/NADPH ratio. Interestingly, feeding cells with ribose-5-phosphate, a precursor for nucleotide biosynthesis that produced through the PPP, could evidently suppress the ROS level and prevent the cell enlargement related to mitochondrial biogenesis. Taken together, these results revealed an important signaling pathway between DNA damage repair and the cell metabolism, which contributed to the premature aging effects of PUVA, and may be generally applicable for a large category of chemotherapeutic reagents including many cancer drugs.

Deposition of type I collage in ECM is an important property of various fibrotic diseases including breast cancer. The excessive expression of type I collagen contributes to the rigidity of cancer tissue and increases the mechanical stresses which facilitate metastasis and proliferation of cancer cells via the activation of TGF-β signaling pathway. The increased mechanical stresses also cause the compression of blood vessels and result in hypoperfusion and impaired drug delivery in cancer tissue. Additionally, type I collage functions as the ligand of α2β1-integrin and DDR1/2 receptors on the membrane of cancer cells to initiate signal transduction leading to metastasis. The expression of type I collage in cancer cells is previously shown to be inducible by TGF-β however the detailed mechanism by which the synthesis of type I collagen is regulated in breast cancer cells remains unclear. Herein, we report that MRTF-A, a co-activator of SRF, is important for the regulation of type I collagen gene COL1A1 in breast cancer cells. MRTF-A physically interacted with the promoter of COL1A1 to facilitate histone acetylation and RNA polymerase II recruitment. The RhoC-ROCK signaling pathway which controls the nuclear localization of MRTF-A regulated the transcription of COL1A1 in human breast cancer cells. TGF-β and Wnt signaling increased the expression of both MRTF-A and COL1A1. Furthermore, depletion of MRTF-A abolished the upregulation of COL1A1 in response to the TGF-β or Wnt signaling, indicating the importance of MRTF-A in the synthesis of type I collagen in breast cancer. Given the crucial roles of type I collagen in the formation of metastasis-prone and hypoperfusion microenvironment, MRTF-A would be a potential target for the development of anti-breast cancer activities.

Xylosidases with tolerance to high concentration of salts, organic solvents, and enzyme hydrolytic products are preferential for industrial application but were rarely reported. In this study, a novel xylosidase XYL21 belong to glycoside hydrolase 39 was characterized with optimal temperature of 45 °C and optimal pH of 5.50. Different to other GH39 xylosidases, XYL21 had excellent tolerance to salts, the activity of which is not inhibited but slightly increased in 0.50–1.50 M NaCl. It is also tolerant to organic solvents, especially retaining 105.18% relative activity even in the presence of 15.00% (v/v) ethanol. Moreover, XYL21 was insensitive to the final lignocellulose hydrolysis products including glucose, xylose, arabinose, mannose and galactose, which retains 111.36% and 53.49% relative activity in 0.30 and 0.90 M xylose, respectively. Further structural modeling analysis indicated that its excellent tolerance may be attributed to its high structural flexibility caused by the high proportion of random coils. Furthermore, XYL21 had a wide substrate specificity to catalyze xylan and xylo-oligosaccharides, and it significantly cooperated with xylanase to improve the hydrolysis efficiency with 1.52-fold. Considering these unique properties, XYL21 is a good candidate for both basic research and various potential industrial applications such as seafood processing and bioethanol production.

Moisture content in coal is an important factor affecting the coal seam gas extraction. It directly affects the storage and flow of gas in bituminous coal. In this paper, the cylindrical bituminous coal cores of Xutuan coal mine in Huaibei coal mine group were studied as experimental objects, using the laboratory self-designed experimental device Gas Adsorption and Strain Testing Apparatus system. The influence of the bituminous coal moisture content on gas adsorption characteristics was studied. Drying experiments of coal samples showed that they lose the original moisture content following the exponential decay function of time. At wetting, the saturated moisture content in coal samples increased following the Exponential Association function of time. The experimental results show that the average original moisture content and average saturated moisture content of raw coal samples are 1.3 and 2.4%, respectively. On this basis, the gas adsorption experiments on samples with different moisture contents under different gas pressures were carried out. With the moisture content increase, the gas adsorption capacity and saturation value decreased and the decrease rate gradually reduced. The single exponential decay function describes the gas adsorption capacity dependence on moisture content. Moisture content also affects the adsorption deformation of bituminous coal. At high moisture content, the adsorption deformation of bituminous coal is less.

To establish stable infliximab-expressing Chinese hamster ovary (CHO) cells with high tolerance to serum-free culture. Bcl-2 antagonist/killer 1 (BAK1), which is a key mediator of the apoptosis pathway, was disrupted, and infliximab, which is a broadly used monoclonal antibody for the treatment of rheumatoid arthritis and other autoimmune diseases, was incorporated into the BAK1 locus of the CHO chromosome using the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas genome-editing technique. The activating effects of serum starvation on BAK1 and cytochrome C (CytC) were suppressed in the genome-edited cells, and the ability of the cells to resist the serum starvation-induced loss of mitochondrial membrane potential and apoptosis was increased, as indicated by the results of polymerase chain reaction (PCR), flow cytometry, enzyme-linked immunosorbent assay (ELISA) and high-performance liquid chromatography (HPLC) analysis. In addition, during subsequent passages, infliximab could be stably produced in the genome-edited CHO cells, and the recombinant antibody could effectively antagonize the cytotoxic effect of tumor necrosis factor α (TNFα). A CHO cell line capable of stably expressing infliximab and adapting to serum-free culture was constructed. This work lays the foundation for the development of infliximab biosimilars.

To achieve biosynthesis of caffeoylmalic acid from glucose in engineered Escherichia coli. We constructed the biosynthetic pathway of caffeoylmalic acid in E. coli by co-expression of heterologous genes RgTAL, HpaBC, At4CL2 and HCT2. To enhance the production of caffeoylmalic acid, we optimized the tyrosine metabolic pathway of E. coli to increase the supply of the substrate caffeic acid. Consequently, an E. coli–E. coli co-culture system was used for the efficient production of caffeoylmalic acid. The final titer of caffeoylmalic acid reached 570.1 mg/L. Microbial production of caffeoylmalic acid using glucose has application potential. In addition, microbial co-culture is an efficient tool for producing caffeic acid esters.

Abstract The retinoid X receptor alpha (RXRα) is an important therapeutic target impacting diverse biological processes. Activation of RXRα is known to suppress cancer cell growth. However, the cellular mechanism has been elusive. In the present study, we addressed its role during stem cell differentiation and the underlying connections with carcinogenesis. RXRα was significantly upregulated following the differentiation of human mesenchymal stem cell (hMSC) toward the formation of endothelial cell (EC). However, overexpression of RXRα in hMSC provoked a senescence-like phenotype accompanied by the elevation of tumor suppressor p53, p21, and p16. Consistently, RXRα level was suppressed in cancer cells (~five times lower compared to differentiated hMSC), and its elevation could inhibit the proliferation, migration, and angiogenesis of cancer cells. We further demonstrated that these inhibitory effects were related to RXRα’s interaction with estrogen receptor α (ERα) as well as EGF and ANGPTL3 through modulating PI3K/AKT signaling pathway by AKT and FAK phosphorylation. Moreover, RXRα inhibited glycolytic metabolism in cancer cells, which might be underlying its inhibition of differentiation and carcinogenic features. These data suggest that RXRα acts as a suppressor rather than a driving force during stem cell differentiation, and unbalanced RXRα can trigger multiple yet connected signaling pathways in preventing carcinogenesis.

Sulforaphane (SFN), a natural compound derived from cruciferous vegetables, has been proved to possess potent anti-cancer activity. SMYD3 is a histone methyltransferase which is closely related to the proliferation and migration of cancer cells. This study showed that SFN could dose-dependently induce cell cycle arrest, stimulate apoptosis, and inhibit proliferation and migration of gastric carcinoma cells. Accompanied with these anti-cancer effects, SMYD3 and its downstream genes, myosin regulatory light chain 9, and cysteine-rich angiogenic inducer 61, was downregulated by SFN. Furthermore, overexpression of SMYD3 via transfection could abolish the effects of SFN, suggesting that SMYD3 might be an important mediator of SFN. To the best of our knowledge, this is the first report describing the role of SMYD3 in the anti-cancer of SFN. These findings might throw light on the development of novel anti-cancer drugs and functional food using SFN-rich cruciferous vegetables.

Copy number variation (CNV) has emerged as another important genetic marker in addition to SNP for understanding etiology of complex disease. Kv channel interacting protein 1 (KCNIP1) is a Ca2+-dependent transcriptional modulator that contributes to the regulation of insulin secretion. Previous genome-wide CNV assay identified the KCNIP1 gene encompassing a CNV region, however, its further effect and risk rate on type 2 diabetes (T2D) have rarely been addressed, especially in Chinese population. The current study aims to detect and excavate genetic distribution profile of KCNIP1 CNV in Chinese T2D and control populations, and further to investigate the associations with clinical characteristics. Divergent patterns of the KCNIP1 CNV were identified (p < 0.01), in which the copy number gain was predominant in T2D, while the copy number normal accounted for the most in control group. Consistently, the individuals with copy number gain showed significant risk on T2D (OR = 4.550, p < 0.01). The KCNIP1 copy numbers presented significantly positive correlations with fasting plasma glucose and glycated hemoglobin in T2D. For OGTT test, the T2D patients with copy number gain had remarkably elevated glucose contents (60, 120, 180-min, p < 0.05 or p < 0.01) and diminished insulin levels (60, 120-min, p < 0.05) than those with copy number loss and normal, which suggested that the KCNIP1 CNV was correlated with the glucose and insulin action. This is the first CNV association study of the KCNIP1 gene in Chinese population, and these data indicated that KCNIP1 might function as a T2D-susceptibility gene whose dysregulation alters insulin production.

Twelve new compounds including four isoflavonolignans (1a/1b and 2a/2b), two neolignans (3a/3b), and six isoflavonoids (5–10), together with seven known compounds (4 and 11–16) were isolated from the stems of Pisonia umbellifera. The structures were elucidated on the basis of comprehensive spectroscopic analyses and ECD calculation methods. Compounds 1–3 were present as enantiomers that were successfully separated by chiral HPLC. Compounds 1a/1b and 2a/2b are the first examples of isoflavonolignans with a pyranoid ring linking up the isoflavonoid and the monolignol from nature. A putative biosynthetic pathway for the isoflavonolignans was deduced. The anti-inflammatory and cytotoxic activities for all compounds were evaluated.

As a proinflammatory cytokine, tumor necrosis factor-alpha (TNF-α) plays a pivotal role in various autoimmune diseases such as rheumatoid arthritis (RA). Thus, TNF-α has been defined as a therapeutic target for RA. Although some TNF-α antagonists including neutralizing monoclonal antibodies and soluble receptors have been approved to be successful in attenuating symptoms in patients suffering from RA, the long-term use of these passive immunization reagents could cause some problems like a variable degree of immunogenicity. In the present study, in order to wake up active immune responses of RA patients, we developed a recombinant TNF-α therapeutic vaccine (named mrTNF-PADRE) by coupling a 12-amino acid universal Pan HLA-DR Epitope (PADRE) to the protein. Codon optimization was performed to improve the secondary structure of mrTNF-PADRE mRNA to ensure its heterologous expression. As a result, a single codon synonymous mutation greatly elevated recombinant protein expression (about 30% of the total bacteria proteins) in E. coli as compared with the undetectable expression of the unoptimized gene. Although expressed as insoluble inclusion bodies (IBs), the vaccine can be effectively prepared with a purity of over 95% by IBs washing and one-step gel-infiltration chromatography. By this strategy, a stable yield of 5.2 mg purified mrTNF-PADRE per gram of cell paste could be obtained.

In this study, the κ-carrageenan supplementation significantly reduced the gains in weight, fat mass, adipocyte size, fasting blood glucose as well as blood lipid in mice with high-fat diet. The investigation of the gene expression levels revealed that κ-carrageenan could improve high-fat diet induced obesity by the AMPK and PPARγ signaling pathway. The level of TNF-α and LPS in the LCGN group was significantly lower than that in the high-fat diet (HFD) group, while there were no significant difference in the HCGN group. The κ-carrageenan treatment was beneficial to the recovery of intestinal homeostasis in mice with high-fat diet, expecially LCGN. The abundance of Firmicutes, Proteobacteria, Lachnospiraceae, and Desulfovibrionaceae were positively correlated with body weight, fasting blood glucose (FBG), serum low density lipoprotein cholesterol (LDL-c), total cholesterol (TC), and triglyceride (TG) levels. By contrast, Bacteroidetes were negatively correlated with LDL-c and TC levels, while positively with FBG.

As a kind of associated geological gas, coalbed methane (CBM) is mainly adsorbed in the coal seam. The coal-methane adsorption phenomenon can be described by Langmuir monolayer adsorptio n model, BET multilayer adsorption model and the Theory of Volume Filling of Micropore (TVFM), whereas the binary gas adsorption phenomenon can be described by the extended Langmuir Model. For the CBM in the low permeability coal seam, the amount of gas released by direct drainage is relatively limited, which cannot eliminate the gas explosion and outburst hazards. Gas injection is an effective method to promote methane drainage. In this paper, the free desorption and nitrogen injection displacement experiments are comparatively analyzed, which allows verifying the effectiveness of nitrogen injection’s enhancement to gas drainage. The experiment of injecting nitrogen gas into the coal body shows that the coal fracture can be maintained or expanded by the injected gas pressure so that more methane can be released. The nitrogen injection has a higher time efficiency than that of free desorption as well. The displacement ratio of N2/CH4 is in the range of 1-3. Both the injection pressure and confining pressure affect the displacement ratio. The analysis of the desorbed gas components shows that the relationship between the methane component and gas flooding time is an “inverted S” shape curve, and the appropriate time for the methane collection can be inferred by the time interval of the rapid decline of the curve.

Vascular smooth muscle cells (VSMCs), switching from a differentiated to a proliferative phenotype, contribute to various vascular diseases. However, the role of long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 MALAT1 in the phenotype switching of VSMCs remains unclear. Here, we report that the knockdown of MALAT1 promotes the transformation of smooth muscle cells from a proliferative phenotype to a differentiated phenotype. MALAT1 knockdown inhibited cellular proliferation and migration, leading to significant cell cycle arrest in the G2 phase. MALAT1 was downregulated in bone morphogenetic protein-7 (BMP-7)-induced cellular differentiation, while MALAT1 was upregulated in platelet-derived growth factor-BB (PDGF-BB)-induced cellular proliferation. PDGF induced the transformation of smooth muscle cells into a proliferative phenotype accompanied by an increase in autophagy. The downregulation of MALAT1 attenuated PDGF-BB-induced proliferation and migration by inhibiting autophagy. MALAT1 could act as a competing endogenous RNA (ceRNA) to regulate autophagy-related 7 (ATG7) gene expression by sponging miR142-3p. The present study reveals a novel mechanism by which MALAT1 promotes the transformation of smooth muscle cells from contraction to synthetic phenotypes.