AKT regulates IL-1β-induced proliferation and activation of hepatic stellate cells

Background: Activated hepatic stellate cells (HSCs) are closely involved in the initiation, perpetuation, and resolution of liver fibrosis. Pro-inflammatory cytokine levels are positively correlated with the transition from liver injury to fibrogenesis and contribute to HSC pathophysiology in liver fibrosis. Methods: In this study, we investigated the effect of the pro-inflammatory cytokine interleukin (IL)-1β on the proliferation and signaling pathways involved in fibrogenesis in LX-2 cells, an HSC cell line, using western blotting and cell proliferation assays. Results: IL-1β increased the proliferation rate and α-smooth muscle actin (SMA) expression of LX-2 cells in a dose-dependent manner. Within 1 h after IL-1β treatment, c-Jun N-terminal kinase (JNK), p38, and nuclear factor-κB (NF-κB) signaling was activated in LX-2 cells. Subsequently, protein kinase B (AKT) phosphorylation and an increase in α- SMA expression were observed in LX-2 cells. Each inhibitor of JNK, p38, or NF-κB decreased cell proliferation, AKT phosphorylation, and α-SMA expression in IL-1β-treated LX-2 cells. Conclusion: These results indicate that JNK, p38, and NF-κB signals converge at AKT phosphorylation, leading to LX-2 activation by IL-1β. Therefore, the AKT signaling pathway can be used as a target for alleviating liver fibrosis by the inflammatory cytokine IL-1β.     

Ang-(1-7) exerts anti-inflammatory and antioxidant activities on high glucose-induced injury by prohibiting NF-κB-IL-1β and activating HO-1 pathways in HUVECs

Previous reports have suggested that Ang-(1-7) may have a protective effect in endothelial cells against high glucose (HG)-induced cell injury thanks to a modulatory mechanism in the NF-κB signaling pathway. In this study, we have examined whether NF-κB-IL-1β and Heme oxygenase-1 (HO-1) pathways contribute to the protection of Ang-(1-7) against hyperglycemia-induced inflammation and oxidative stress in human umbilical vein endothelial cells (HUVECs). Our results indicate that time-varying exposures of HUVECs, from 1 h to 24 h, to high glucose concentrations result in an increased expression of phosphorylated (p)-p65 and HO-1 in a time-dependent manner. As an inhibitor of NF-κB, pyrrolidinedithiocarbamic acid (PDTC) suppressed IL-1β production induced by HG. Of note, HUVECs previously treated with Ang-(1-7) (2 μM) for 30 min before being exposed to HG concentrations significantly ameliorated the HG-increased in p-p65 and IL-1β expression; whereas obviously up-regulated the level of HO-1, along with inhibition of oxidative stress, inflammation, and the HG-induced cytotoxicity. Importantly, when HUVECs were previously treated either with PDTC or IL-1Ra for 30 min before being exposed to HG, it significantly prevented damages caused by high glucose concentrations mentioned above, while the treatment of HO-1 inhibitor Sn-protoporphyrin (SnPP) before exposure to both HG and Ang-(1-7) significantly blocked the protective effect exerted by Ang-(1-7) on endothelial cells against injuries induced by HG mentioned above. To conclude, the data of this study showed that activation and inhibition of the NF-κB-IL-1β pathway and HO-1 pathway may constitute an important defense mechanism against endothelial cell damage caused by HG concentrations. We additionally gave new evidence showing that exogenous Ang-(1-7) exerts a protective effect on HUVECs against the HG-induced cell injury via the inhibition and the activation of the NF-κB-IL-1β pathway and the HO-1 pathway, respectively.     

Biosynthesis of raw starch degrading β-cyclodextrin glycosyltransferase by immobilized cells of <i>Bacillus licheniformis</i> using potato wastewater

The study was sought to enhance the synthesis of thermal stable β-cyclodextrin glycosyltransferase (β-CGTase) using potato wastewater as a low-cost medium and assess the degree to which it is efficient for industrial production of β-cyclodextrin (β-CD) from raw potato starch. Thermophilic bacteria producing β-CGTase was isolated from Saudi Arabia and the promising strain was identified as Bacillus licheniformis using phylogenetic analysis of the 16S rRNA gene. Alginate-encapsulated cultures exhibited twice-fold of β-CGTase production more than free cells. Scanning electron microscopy (SEM) of polymeric capsules indicated the potential for a longer shelf-life, which promotes the restoration of activity in bacterial cells across semi-continuous fermentation of β-CGTase production for 252 h. The optimal conditions for β-CGTase synthesis using potato wastewater medium were at 36 h, pH of 8.0, and 50°C with 0.4% potato starch and 0.6% yeast extract as carbon and nitrogen sources, respectively. The purified enzyme showed a specific activity of 63.90 U/mg with a molecular weight of ∼84.6 kDa as determined by SDS-PAGE analysis. The high enzyme activity was observed up to 60°C, and complete stability was achieved at 75°C. High levels of activity and stability were shown at pH 8.0, and the pH range from 7.0–10.0, respectively. The enzyme has an appreciable affinity for raw potato starch with a Km of 5.7 × 10⁻⁶ M and a Vmax of 87.71 µmoL/mL/min. β-CD production was effective against 25 U/g of raw potato starch. The outcomes demonstrated its feasibility to develop a fermentation process by integrating the cost-effective production of β-CGTase having distinctive properties for β-CD production with ecofriendly utilization of potato wastewater.     

Vitamin B3 inhibits apoptosis and promotes autophagy of islet β cells under high glucose stress

Background: Hyperglycemia is a typical symptom of diabetes. High glucose induces apoptosis of islet β cells. While autophagy functions in cytoprotection and autophagic cell death. The interaction between autophagy and apoptosis is important in the modulation of the function of islet β cells. Vitamin B3 can induce autophagy and inhibit islet β apoptosis.Method: The mechanism of vitamin B3-mediated protective effect on the function of islet β cells was explored by the method of western blot, immunofluorescence and flow cytometry.Results: In the present study, high glucose stress increased the apoptosis rate, while vitamin B3 reduced the apoptosis rate. The effect of vitamin B3 on autophagy flux under normal and high glucose stress was also investigated. Vitamin B3 increased the number of autophagosomes and increased the light chain (LC)3-II/LC3-I ratio. In contrast, vitamin B3 decreased sequestosome 1 (SQSTM1)/p62 protein expression and inhibited the phosphorylation of mammalian ribosomal protein S6 kinase β-1 (p70S6K/S6K1), which was a substrate of mammalian target of rapamycin (mTOR) under normal and high glucose stress. To further verify the protective effect of vitamin B3 on apoptosis, we treated islet β cell RIN-m5F with autophagy inhibitor 3-methyladenine (3-MA). Vitamin B3 decreased the apoptosis rate under high glucose stress, while the inhibition of apoptosis by vitamin B3 was blocked after adding 3-MA.Conclusion: Our data suggested that vitamin B3 reduced the apoptosis rate of β cells, possibly through inducing autophagy under high glucose stress.     

β-内酰胺类抗生素质谱裂解机理研究

采用电喷雾-四极杆-飞行时间串联质谱(ESI-Q-TOF-MS/MS)对3种头孢类和2种青霉素类药物的[M+Na]⁺ 裂解规律进行研究。头孢类药物[M+Na]⁺裂解可产生许多互补的碎片离子,1,3-H重排和retro-Diels-Alder (RDA)是其重要的裂解方式;青霉素类药物[M+Na]⁺裂解除了一些常规的中性分子丢失外,在氘代实验的基础上提出了比以往更合理的H₂S丢失途径,即1,5-H重排。本实验丰富了β-内酰胺类药物的质谱裂解途径,对应用质谱技术快速鉴定复杂体系中β-内酰胺类抗生素具有重要作用。     

Interleukin-1β regulates metalloproteinase activity and leptin secretion in a cytotrophoblast model

Implantation is one of the most regulated processes in human reproduction, by endocrine and immunological systems. Cytokines are involved in embryo-maternal communication and an impaired balance could result in pregnancy loss. Here we investigated the effect of interleukin 1-β on the activity of two important metalloproteinases (MMP-2 and MMP-9) that are involved in extracellular matrix remodeling as well as the secretion of leptin, one of the reproductive hormones actively regulating their activity and secretion. We found that IL-1β activates matrix metalloproteinase activity as well as increases leptin secretion. We propose that this interleukin, through the regulation of leptin, in turn activates matrix metalloproteinases which results in an increased cytotrophoblast invasion.     

Scutellarin alleviates complete freund’s adjuvant-induced rheumatoid arthritis in mice by regulating the Keap1/Nrf2/HO-1 pathway

Scutellarin (SCU) is a herbal flavonoid glucuronide with multiple pharmacological activities, including anti-oxidant, anti-inflammation, vascular relaxation, anti-platelet, and myocardial protection. However, the effect of SCU on complete Freund’s adjuvant (CFA)-induced rheumatoid arthritis (RA) had not been studied. In this study, we investigated the beneficial effects of SCU in the CFA-induced RA mice model and the anti-arthritic activity was evaluated by paw edema. Enzyme-linked immunosorbent assay (ELISA) was carried out to evaluate the plasma levels of immunoglobulin (Ig)G, IgE, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, receptor activator of nuclear factor-κB ligand (RANKL), and osteoprotegerin (OPG). Histological slides were prepared from the harvested paws of mice to determine the pathological changes in the joints. The proportions of T helper type 1 (Th1) and T helper type 2 (Th2) cells of CD4+ T lymphocyte subsets were analyzed by flow cytometry. The expression of Kelch-like ECH-associated protein 1 (Keap1), nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1) was analyzed using real-time quantitative PCR (RT-qPCR) and western blotting assays. The present study demonstrated that SCU prevented CFA-induced RA, and inhibited the expression of inflammation factors, IgG, IgE, TNF-α, IL-1β, and IL-6. While SCU also reduced the RANKL level, it increased OPG expression in RA mice. The Th1/Th2 ratio was significantly lower in mice treated with SCU. Additionally, HO-1 expression was reduced while the expression of Keap1 and Nrf2 was elevated following SCU treatment. Results provide preliminary evidence to employ SCU in arthritis treatment which might be related to the regulation of Th1/Th2 balance and the Keap1/Nrf2/HO-1 pathway.     

Suspension state promotes extravasation of breast tumor cells by increasing integrin β1 expression

Mechanical microenvironment can strongly affect the metastatic efficiency of circulating tumor cells. However, the effect of suspension state on their extravasation and the mechanisms involved are still unclear. To explore the influence of suspension state on extravasation (including adhesion, spreading and transendothelial migration) of breast tumor cells and its relevant molecular mechanism, MDA-MB-231 cells were cultured on poly (2-hydroxyethyl methacrylate) coated 6-well plates to minic the suspension state. Suspension state promoted adhesion, spreading and transendothelial migration of MDA-MB-231 cells to EAhy926 endothelial cells (ECs) monolayer under both the static condition and 0.5 dyne/cm² flow shear stress (FSS). The number of cells adhered to ECs monolayer reached 2.15 (static condition, 3 d) and 1.67 (FSS, 3 d) times, and the number of migration reached 10.60 times, respectively, as compared to that in adhesion state. Moreover, MDA-MB-231 cells knockdown of integrin β1 provoked poor adhesion and transendothelial migration, as compared with MDA-MB-231 cells. But it made no difference in cell spreading. Our results implied the increasing expression of integrin β1 induced by suspension culture promoted the adhesion and transendothelial migration of MDA-MB-231 cells, but had no significant influence on their spreading.     

Semaphorin 7A promotes human vascular smooth muscle cell proliferation and migration through the β-catenin signaling pathway

Background: Vascular smooth muscle cells (VSMCs) undergo a conversion from a contractile phenotype to a proliferative synthetic phenotype, contributing to the pathogenesis of cardiovascular diseases. Semaphorin 7A (SEMA7A) is a glycosylphosphatidylinositol-anchored membrane protein that plays an important role in vascular homeostasis by regulating endothelial cell behaviors. However, the expression and role of SEMA7A in VSMCs remain unclear.Methods: In this study, we screened for VSMC-regulating genes in publicly available datasets and analyzed the expression of SEMA7A in human coronary artery smooth muscle cells (hCASMCs) treated with platelet-derived growth factor-BB (PDGF-BB). The effects of SEMA7A overexpression and knockdown on hCASMC proliferation and migration were examined. The signaling pathways involved in the action of SEMA7A in hCASMCs were determined.Results: Bioinformatic analysis showed that SEMA7A was significantly dysregulated in VSMCs treated with oxidized low-density lipoprotein or overexpressing progerin, a pro-atherogenic gene. The PDGF-BB stimulation led to a concentration- and time-dependent induction of SEMA7A. Depletion of SEMA7A attenuated PDGF-BB-induced hCASMC proliferation and migration. Conversely, overexpression of SEMA7A enhanced hCASMC proliferation and migration. Mechanistically, SEMA7A stimulated the activation of the β-catenin pathway and upregulated c-Myc, CCND1, and MMP7. Knockdown of β-catenin impaired SEMA7A-induced hCASMC proliferation and migration.Conclusions: SEMA7A triggers phenotype switching in VSMCs through the β-catenin signaling pathway and may serve as a potential therapeutic target for cardiovascular diseases.     

同位素稀释-气相色谱-串联质谱法测定猪肉中残留的4种β-受体激动剂

采用同位素稀释质谱法,结合固相萃取技术,建立了猪肉中克伦特罗、妥布特罗、溴布特罗、沙丁胺醇等4种β-受体激动剂残留量的气相色谱-串联质谱（GC-MS/MS）检测方法。向绞碎的猪肉中加入同位素内标克伦特罗-D₉、妥布特罗-D₉、溴布特罗-D₉,经β-盐酸葡萄糖醛苷酶/芳基硫酸酯酶酶解,醋酸铵缓冲溶液提取,高氯酸调pH,HLB-MCX固相萃取柱串联净化,用-N,O-双三甲基硅基三氟乙酰胺(BSTFA)+1%三甲基氯硅烷（TMCS）衍生化后,进行GC-MS/MS测定,使用内标法定量。结果表明,猪肉中添加2.5~35.0 μg/kg水平的β-受体激动剂,回收率为92.2%~112.8%,批内变异系数（CV）为1.0%~12.6%,批间变异系数（CV）为1.9%~13.2%。4种β-受体激动剂的线性范围为1.35~158.00 μg/L,相关系数R²≥0.999 7,方法最低检测限为0.13~0.40 μg/kg,最低定量限为0.40~1.27 μg/kg。该方法精密度好、灵敏度高,可简便、准确地测定猪肉中4种β-受体激动剂的残留量。     

Anti-inflammatory and antioxidant potential capacities of AD-MSCs and BM-MSCs in suppressing pancreatic β-cells auto-immunity and apoptosis in rats with T1DM induced model

Since Type 1 diabetes (T1DM) occurs when β-cells mass is reduced to less than 20% of the normal level due to autoimmune destruction of cells resulting in the inability to secrete insulin, preservation or replenishment of the functional β-cells mass has become a major therapeutic focus for this diabetic type treatment. Thus, this 4-week work plan was designed to determine which mesenchymal stem cells (MSCs) type is more appropriate to alleviate pancreatic hazards resulting from diabetes induction; via tracking a comparative study between MSCs derived from adipose tissue (AD-MSCs) and from bone marrow (BM-MSCs) in management of T1DM considering their immunomodulatory, anti-apoptotic and antioxidative roles. Rats were divided randomly into 4 groups; control, STZ-diabetic (D), D+AD-MSCs, and D+BM-MSCs groups. Both stem cells types in this study were allogenic. Herein, both oxidative stress and antioxidant markers were evaluated using colorimetric analysis, while inflammatory, immune and apoptotic markers were assessed through flow cytometric analysis. Results showed that diabetic rats treated with either AD-MSCs or BM-MSCs exhibited marked pancreatic antioxidant and anti-inflammatory activities that were able to initiate pancreatic immunomodulation and reducing β-cells apoptotic death, thus, help to restore their normal insulin secretion and hypoglycemic abilities. However, AD-MSCs injection was shown to be superior as a pancreatic regenerative tool in overcoming diabetes; owing to their marked antioxidant, anti-inflammatory, immunomodulatory, and anti-apoptotic characteristics over BM-MSCs treatment.     

Minireview: C2- and C4-position 17β-estradiol metabolites and their relation to breast cancer

C2- and C4-position 17β-estradiol metabolites play an important role in breast carcinogenesis. 2-Hydroxyestradiol and 4-hydroxyestradiol are implicated in tumorigenesis via two pathways. These pathways entail increased cell proliferation and the formation of reactive oxygen species that trigger an increase in the likelihood of deoxyribonucleic acid mutations.
2-Methoxyestradiol, a 17β-estradiol metabolite, however, causes induction of apoptosis in transformed and tumor cells; thus exhibiting an antiproliferative effect on tumor growth. The 4-hydroxyestradiol:2- methoxyestradiol and 2-hydroxyestradiol:2-methoxyestradiolratios therefore ought to be taken into account as possible indicators of carcinogenesis.     

Interleukin-1 receptor antagonist: From synthesis to therapeutic applications

The cytokine channel’s mechanism for self-regulation involves the application of antagonistic cytokines that are synthesized to connect to the receptors and release soluble cytokine receptors. The very first receptor antagonist of cytokine that was naturally present was interleukin-1 receptor antagonist (IL-1Ra). The IL-1Ra protein forms are disinfected from supernatants of cultured monocytes on stacked IgG. The family of IL-1 consists of IL-1α, IL-1β and IL-1Ra. Human monocytes regulate the production of IL-Ra. IL-Ra takes part in normal physiological functions by using specific antibodies, and acts as an anti-inflammatory agent. IL-Ra is synthesized in the tissues during the period of active disease and can be systematically measured and/or estimated. Maintenance of the levels of IL-Ra and IL-1 is the main factor for host resistance in patients during diseased conditions, as IL-Ra acts as an inherent regulator of various inflammatory responses. In this article, we focuse on how IL-Ra is synthesized and performs its functions once the inflammatory responses are activated.     

Decreased serum HMGB1 associated with M2 macrophage polarization and patients with calcific aortic valve disease

Except for the standard aortic valve replacement, no effective medical treatment is available to prevent or delay calcific aortic valve disease (CAVD) progression. Recently, macrophages and high-mobility group box 1 (HMGB1) are the most intriguing candidates in various inflammatory disorders. However, the association between serum HMGB1, CAVD, and macrophage polarization remains unclear. Therefore, we examined whether the level of serum HMGB1 is clinically associated with aortic valve calcification and whether HMGB1 treatment can promote macrophage differentiation toward M1 or M2 phenotype. This experimental study included 19 CAVD patients and 20 healthy controls whose serum HMGB1 levels were examined by ELISA assay. THP-1 macrophage polarization system was established to test the polarization capability of HMGB1 treatment. The results showed that serum levels of HMGB1 were significantly reduced in patients with CAVD. HMGB1 treatment promoted M2 macrophage polarization but not M1 phenotype with increased IL-10 expression and reduced inducible nitric oxide synthase (iNOS) expression. Our findings suggest that serum HMGB1 is negatively associated with the development of aortic valve calcification, and HMGB1 treatment may facilitate M2 macrophage polarization for reducing aortic valve calcification.     

Association of TRIM22 with the type 1 interferon response during primary human cytomegalovirus infection in THP-1 macrophages

As a response factor of interferon, tripartite motif (TRIM) 22 was reported to exert antiviral activity against viruses. In this study, THP-1 macrophages were infected with human cytomegalovirus (HCMV) to establish the HCMV lytic infection model. The mRNA levels of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α) and interferonbeta (IFN-β) were significantly up-regulated in THP-1 macrophages at different infection time and titers. Moreover, for the first time, upregulation of TRIM22 expression was found during HCMV infection at both mRNA and protein levels in THP-1 macrophages. Furthermore, IFN-β could induce TRIM22 expression in THP-1 macrophages or HCMV infected THP-1 macrophages. Depletion of TRIM22 increased replication activity of HCMV with increasing of HCMV titers and HCMV proteins. In conclusion, it is the first report that HCMV can induce TRIM22 activation through IFN-β signaling and TRIM22 can suppress replication of HCMV in THP-1 macrophages.     

MiR-4262 regulates differentiation and osteogenesis of human periodontal stem cells by targeting suppressor of cytokine signaling 4

Periodontitis, as a chronic inflammatory disease, remains unsolved, and the pathogenesis of this disease has not been fully elucidated. In this study, the effect of miR-4262 was investigated in tumor necrosis factor-α (TNF-α) induced human periodontal stem cells (hPDLSCs) for the first time. The gene expression involved in this study was determined using polymerase chain reaction (PCR), the expressions of relevant proteins were determined by western blot analysis, and the levels of IL-1β, IL-6, and MCP-1 were estimated by enzyme-linked immunosorbent assay (ELISA) assay. The luciferase reporter assay was performed for verification of the target gene, alkaline phosphatase (ALP) activity detection was used for differentiation capacity, and alizarin red staining assay was used for mineralization capacity. The inhibition of miRNA-4262, which resulted in the upregulation of suppressor of cytokine signaling 4 (SOCS4), showed protective effects, including anti-inflammation and promotional effects on osteogenesis as well as differentiation in TNF-α induced hPDLSCs. These results provided insights into the roles of miRNAs in regulating the inflammatory response, differentiation, and osteogenesis in hPDLSCs, which may promote the understanding of the mechanisms of periodontitis and find out a better therapeutic application.     

Long non-coding RNA MIR22HG inhibits the adipogenesis of human bone marrow mesenchymal stem cells with the involvement of Wnt/β-catenin pathway

Osteoporosis is a frequently occurring bone remodeling disorder worldwide with one characteristic being decreasing bone mineral density and a predisposition to bone fracture, which diminishes patients’ quality of life. Several studies showed that imbalance between the osteogenesis and adipogenesis of bone marrow mesenchymal stem cells (BMSCs) took part in the development of osteoporosis. In previous study, we found MIR22HG regulated the osteogenesis of human BMSCs positively. In this study, we found that MIR22HG was decreased during the adipogenesis of human BMSCs and exerted negative effects on adipogenesis with the involvement of Wnt/β-catenin signaling pathway both in vitro and in vivo. Nitazoxanide could inhibit Wnt signaling and relieve MIR22HG’s suppression on adipogenesis. These findings indicated that MIR22HG had great potential in clinical application for osteoporosis treatment and prevention.