The WWOX/HIF1A Axis Downregulation Alters Glucose Metabolism and Predispose to Metabolic Disorders

Recent reports indicate that the hypoxia-induced factor (HIF1α) and the Warburg effect play an initiating role in glucotoxicity, which underlies disorders in metabolic diseases. WWOX has been identified as a HIF1α regulator. WWOX downregulation leads to an increased expression of HIF1α target genes encoding glucose transporters and glycolysis’ enzymes. It has been proven in the normoglycemic mice cells and in gestational diabetes patients. The aim of the study was to determine WWOX’s role in glucose metabolism regulation in hyperglycemia and hypoxia to confirm its importance in the development of metabolic disorders. For this purpose, the WWOX gene was silenced in human normal fibroblasts, and then cells were cultured under different sugar and oxygen levels. Thereafter, it was investigated how WWOX silencing alters the genes and proteins expression profile of glucose transporters and glycolysis pathway enzymes, and their activity. In normoxia normoglycemia, higher glycolysis genes expression, their activity, and the lactate concentration were observed in WWOX KO fibroblasts in comparison to control cells. In normoxia hyperglycemia, it was observed a decrease of insulin-dependent glucose uptake and a further increase of lactate. It likely intensifies hyperglycemia condition, which deepen the glucose toxic effect. Then, in hypoxia hyperglycemia, WWOX KO caused weaker glucose uptake and elevated lactate production. In conclusion, the WWOX/HIF1A axis downregulation alters glucose metabolism and probably predispose to metabolic disorders.     

静电纺再生丝素纳米纤维的结构与性能

静电纺丝获得的丝素纳米级纤维可作为细胞培养支架,用于纺丝工艺及后处理能改变丝素微细结构,影响其水溶性和力学性能。本文采用XRD、FTIR、固态13CNMR和DSC研究了不同工艺下丝素纳米纤维及经甲醇处理后的微细结构,比较了不同微细结构下的水溶性和力学性能。结果表明,电纺丝的微细结构受纺丝工艺影响,高电压、纺丝液中丝素质量分数大时纺得的电纺丝结晶度高,经甲醇处理后,β化程度提高;w(丝素)=11%、15%时制备的电纺丝断裂强度分别为8.5、11.9 cN/mm;w(丝素)=11%、19%,水溶性由51.2%下降到43.3%;w(丝素)=19%、电压32 kV制得的电纺丝甲醇处理前后水溶性从43.3%下降到6.6%,说明丝素纳米纤维结晶度提高,强度增加、水溶性下降,满足了细胞支架的要求。     

Silk Fiber-Reinforced Hyaluronic Acid-Based Hydrogel for Cartilage Tissue Engineering

A continuing challenge in cartilage tissue engineering for cartilage regeneration is the creation of a suitable synthetic microenvironment for chondrocytes and tissue regeneration. The aim of this study was to develop a highly tunable hybrid scaffold based on a silk fibroin matrix (SM) and a hyaluronic acid (HA) hydrogel. Human articular chondrocytes were embedded in a porous 3-dimensional SM, before infiltration with tyramine modified HA hydrogel. Scaffolds were cultured in chondropermissive medium with and without TGF-β1. Cell viability and cell distribution were assessed using CellTiter-Blue assay and Live/Dead staining. Chondrogenic marker expression was detected using qPCR. Biosynthesis of matrix compounds was analyzed by dimethylmethylene blue assay and immuno-histology. Differences in biomaterial stiffness and stress relaxation were characterized using a one-step unconfined compression test. Cell morphology was investigated by scanning electron microscopy. Hybrid scaffold revealed superior chondro-inductive and biomechanical properties compared to sole SM. The presence of HA and TGF-β1 increased chondrogenic marker gene expression and matrix deposition. Hybrid scaffolds offer cytocompatible and highly tunable properties as cell-carrier systems, as well as favorable biomechanical properties.     

Injectable Thixotropic β–Cyclodextrin–Functionalized Hydrogels Based on Guanosine Quartet Assembly

Facile method for the preparation of β–cyclodextrin–functionalized hydrogels based on guanosine quartet assembly was described. A series of seven hydrogels were prepared by linking β–cyclodextrin molecules with guanosine moieties in different ratios through benzene–1,4–diboronic acid linker in the presence of potassium hydroxide. The potassium ions acted as a reticulation agent by forming guanosine quartets, leading to the formation of self–sustained transparent hydrogels. The ratios of the β–cyclodextrin and guanosine components have a significant effect on the internal structuration of the components and, correspondingly, on the mechanical properties of the final gels, offering a tunablity of the system by varying the components ratio. The insights into the hydrogels’ structuration were achieved by circular dichroism, scanning electron microscopy, atomic force microscopy, and X–ray diffraction. Rheological measurements revealed self–healing and thixotropic properties of all the investigated samples, which, in combination with available cyclodextrin cavities for active components loading, make them remarkable candidates for specific applications in biomedical and pharmaceutical fields. Moreover, all the prepared samples displayed selective antimicrobial properties against S. aureus in planktonic and biofilm phase, the activity also depending on the guanosine and cyclodextrin ratio within the hydrogel structure.     

丝素/羧甲基壳聚糖共混膜的结构与性能

利用丝素(SF)与羧甲基壳聚糖(CMCS)共混制取不同比例的SF/CMCS共混膜。研究了CMCS诱导的丝素构象转变行为,测试了共混膜的吸湿性、透湿性和保水性。当CMCS的质量分数为5%时,共混膜中丝素的构象以β-折叠为主;当CMCS的质量分数为10%时,共混膜中丝素的构象由β-折叠向α-螺旋发生转变;当CMCS的质量分数达到15%时,共混膜中丝素的构象向无规卷曲发生转变。当CMCS质量分数小于15%时,共混膜中SF与CMCS具有良好的相容性,溶胀度较小,吸湿性随CMCS含量的增加而迅速降低。     

Recombinant Silk Proteins with Additional Polyalanine Have Excellent Mechanical Properties

This paper explores the structures of exogenous protein molecules that can effectively improve the mechanical properties of silkworm silk. Several transgenic vectors fused with the silkworm fibroin light chain and type 3 repeats in different multiples of the ampullate dragline silk protein 1 (MaSp1) from black widow spider with different lengths of the polyalanine motifs were constructed for this study. Transgenic silkworms were successfully obtained by piggyBac-mediated microinjection. Molecular detection showed that foreign proteins were successfully secreted and contained within the cocoon shells. According to the prediction of PONDR^® VSL2 and PONDR^® VL-XT, the type 3 repeats and the polyalanine motif of the MaSp1 protein were amorphous. The results of FTIR analysis showed that the content of β-sheets in the silk of transgenic silkworms engineered with transgenic vectors with additional polyalanine was significantly higher than that of wild-type silkworm silk. Additionally, silk with a higher β-sheet content had better fracture strength and Young’s modulus. The mechanical properties of silk with longer chains of exogenous proteins were improved. In general, our results provide theoretical guidance and technical support for the large-scale production of excellent bionic silk.     

蚕丝蛋白开发利用的研究进展

较为全面地综述了最近几十年国内外关于蚕丝蛋白的组成、性能,介绍了蚕丝新用途和利用原理,蚕丝脱胶精炼方法,以及蚕丝在化妆品、医药、固定化酶载体材料、食品、再生丝和涂膜化纤表面等方面的应用和研究进展。     

Synthesis and Characterization of New Biodegradable Injectable Thermosensitive Smart Hydrogels for 5-Fluorouracil Delivery

In this paper, injectable, thermosensitive smart hydrogel local drug delivery systems (LDDSs) releasing the model antitumour drug 5-fluorouracil (5-FU) were developed. The systems were based on biodegradable triblock copolymers synthesized via ring opening polymerization (ROP) of ε-caprolactone (CL) in the presence of poly(ethylene glycol) (PEG) and zirconium(IV) acetylacetonate (Zr(acac)₄), as co-initiator and catalyst, respectively. The structure, molecular weight (M_n) and molecular weight distribution (Đ) of the synthesized materials was studied in detail using nuclear magnetic resonance (NMR) and gel permeation chromatography (GPC) techniques; the optimal synthesis conditions were determined. The structure corresponded well to the theoretical assumptions. The produced hydrogels demonstrated a sharp sol–gel transition at temperature close to physiological value, forming a stable gel with good mechanical properties at 37 °C. The kinetics and mechanism of in vitro 5-FU release were characterized by zero order, first order, Higuchi and Korsmeyer–Peppas mathematical models. The obtained results indicate good release control; the kinetics were generally defined as first order according to the predominant diffusion mechanism; and the total drug release time was approximately 12 h. The copolymers were considered to be biodegradable and non-toxic; the resulting hydrogels appear to be promising as short-term LDDSs, potentially useful in antitumor therapy.     

Synthesis of Sucrose-HDI Cooligomers: New Polyols for Novel Polyurethane Networks

Sucrose-1,6-hexamethylene diisocyanate (HDI) cooligomers were synthesized and used as new polyols for poly(ε-caprolactone) (PCL)-based polyurethanes. The polyaddition reaction of sucrose and HDI was monitored by MALDI-TOF MS. It was found that by selecting appropriate reaction conditions, mostly linear oligomer chains containing 16 sucrose units could be obtained. For the synthesis of polyurethane networks, prepolymers were prepared by the reaction of poly(ε-caprolactone) (PCL, 10 kg/mol) with HDI or 4,4′-methylene diphenyl diisocyanate (MDI) and were reacted with sucrose-HDI cooligomers. The so-obtained sucrose-containing polyurethanes were characterized by means of attenuated total reflectance Fourier-transform infrared spectroscopy (ATR-FT IR), swelling, mechanical (uniaxial tensile tests) and differential scanning calorimetry (DSC).     

TCR Recognition of Peptide–MHC-I: Rule Makers and Breakers

T cells are a critical part of the adaptive immune system that are able to distinguish between healthy and unhealthy cells. Upon recognition of protein fragments (peptides), activated T cells will contribute to the immune response and help clear infection. The major histocompatibility complex (MHC) molecules, or human leukocyte antigens (HLA) in humans, bind these peptides to present them to T cells that recognise them with their surface T cell receptors (TCR). This recognition event is the first step that leads to T cell activation, and in turn can dictate disease outcomes. The visualisation of TCR interaction with pMHC using structural biology has been crucial in understanding this key event, unravelling the parameters that drive this interaction and their impact on the immune response. The last five years has been the most productive within the field, wherein half of current unique TCR–pMHC-I structures to date were determined within this time. Here, we review the new insights learned from these recent TCR–pMHC-I structures and their impact on T cell activation.     

温度和pH双重敏感性水凝胶的制备及表征

在5种不同温度下聚合交联,制备了一系列温度和pH双重敏感性聚(N-异丙基丙烯酰胺-co-衣康酸)水凝胶。实验发现,15、25℃得到的凝胶是透明的,45、55℃得到的凝胶是白色不透明的,而在相转变温度附近(35℃)得到的凝胶则是半透明的。FTIR测定表明,它们具有相似的化学组成和结构。SEM观察证实,它们具有不同的表面形态。测定了不同温度和pH下达到平衡时水凝胶的溶胀比,考察了水凝胶在水和强酸性溶液中的去溶胀动力学。结果表明,与15℃或25℃制备的水凝胶相比,45℃和55℃制备的水凝胶的性能有显著提高:(1)溶胀比大为增加。15℃或25℃制备的水凝胶在25℃时溶胀比分别为65.3和68.1,而45℃和55℃制备的水凝胶此时溶胀比分别高达105.7和110.1;(2)45℃和55℃制备的水凝胶在极端环境下对温度的变化仍具有较快的响应速率。例如在温度为60℃,pH=1.67的强酸条件下,45℃和55℃制备的水凝胶在60 min内皆可失去95%的水,而15℃或25℃制备的水凝胶在120 min内只能失去42%左右的水。     

β-Hexachlorocyclohexane Drives Carcinogenesis in the Human Normal Bronchial Epithelium Cell Line BEAS-2B

Organochlorine pesticides constitute the majority of the total environmental pollutants, and a wide range of compounds have been found to be carcinogenic to humans. Among all, growing interest has been focused on β-hexachlorocyclohexane (β-HCH), virtually the most hazardous and, at the same time, the most poorly investigated member of the hexachlorocyclohexane family. Considering the multifaceted biochemical activities of β-HCH, already established in our previous studies, the aim of this work is to assess whether β-HCH could also trigger cellular malignant transformation toward cancer development. For this purpose, experiments were performed on the human normal bronchial epithelium cell line BEAS-2B exposed to 10 µM β-HCH. The obtained results strongly support the carcinogenic potential of β-HCH, which is achieved through both non-genotoxic (activation of oncogenic signaling pathways and proliferative activity) and indirect genotoxic (ROS production and DNA damage) mechanisms that significantly affect cellular macroscopic characteristics and functions such as cell morphology, cell cycle profile, and apoptosis. Taking all these elements into account, the presented study provides important elements to further characterize β-HCH, which appears to be a full-fledged carcinogenic agent.     

再生丝素/壳聚糖共混纳米纤维的结构与性能

以98%的甲酸为溶剂,不同质量分数的再生丝素溶液和3.5%的壳聚糖溶液以质量比70∶30共混静电纺丝。测定了壳聚糖的含量对共混膜的结构及力学、溶解等性能的影响。结果表明:随着壳聚糖相对含量的增加,丝素β化程度提高,纤维结晶度增大,丝素与壳聚糖以70∶30共混时的溶失率最小;甲醇处理后,溶失率明显降低;共混纳米纤维的断裂强度随着壳聚糖相对含量的增大而增加,柔软性也逐渐提高;共混纤维膜具有优异的抗菌性。     

An Exploratory Pilot Study of Genetic Marker for IgE-Mediated Allergic Diseases with Expressions of FcεR1α and Cε

The high affinity immunoglobulin E (IgE) receptor-FcεR1 is mainly expressed on the surface of effector cells. Cross-linking of IgE Abs bound to FcεR1 by multi-valent antigens can induce the activation of these cells and the secretion of inflammatory mediators. Since FcεR1 plays a central role in the induction and maintenance of allergic responses, this study aimed to investigate the association of FcεR1 with the allergic phenotype of Cε expression and cytokine and histamine release from peripheral leukocytes. Peripheral leukocytes from 67 allergic and 50 non-allergic subjects were used for genotyping analysis. Peripheral mononuclear cells (PBMCs) were used for Cε expression and ELISpot analysis, while polymorphonuclear cells (PMNs) were used for histamine release. The association between genotype polymorphism of the FcεR1α promoter region (rs2427827 and rs2251746) and allergic features of Cε expression and histamine were analyzed, and their effects on leukocytes function were compared with wild type. The genotype polymorphisms of FcεR1α promoter region with CT and TT in rs2427827 and TC in rs2251746 were significantly higher in allergic patients than in non-allergic controls. Patients with single nucleotide polymorphism (SNP) of FcεR1α promoter region had high levels of total IgE, mite-specific Der p 2 (Group 2 allergen of Dermatophagoides pteronyssinus)-specific IgE and IgE secretion B cells. The mRNA expression of FcεR1α was significantly increased after Der p2 stimulation in PBMCs with SNPs of the FcεR1α promoter region. Despite the increased Cε mRNA expression in PBMCs and histamine release from PMNs and the up-regulated mRNA expression of interleukin (IL)-6 and IL-8 secretions after Der p2 stimulation, there was no statistically significant difference between SNPs of the FcεR1α promoter region and the wild type. SNPs of FcεR1α promoter region were associated with IgE expression, IgE producing B cells, and increased Der p2-induced FcεR1α mRNA expression. These SNPs may be used as a disease marker for IgE-mediated allergic inflammation caused by Dermatophagoides pteronyssinus.     

静电纺丝工艺参数对丝素/壳聚糖纳米纤维的形貌及直径的影响

以98%的甲酸为溶剂,不同质量分数的再生丝素溶液和3.5%的壳聚糖溶液以质量比70:30共混静电纺丝。用扫描电子显微镜(SEM)观察了丝素质量分数、电压和极距(喷丝口到收集装置的距离)对丝素/壳聚糖纳米纤维的形貌及直径的影响。正交试验结果表明:在丝素/壳聚糖溶液静电纺丝的工艺参数中,对纤维平均直径的影响因素由大到小依次为丝素质量分数、电压、极距。单因素试验表明:丝素/壳聚糖纳米纤维的平均直径及其分布范围随丝素质量分数的增加而增大;在15￣30kV范围内纤维的平均直径随电压增大而减小;当极距大于12cm时,对纤维直径影响不大。最佳工艺条件为:丝素质量分数13%,电压30kV,极距为12cm,制得的纳米纤维平均直径104nm。     

凝固条件对纤维素/丝素蛋白纤维相形态及性能的影响

蛋白质纤维具有光滑柔顺、透气吸湿等优点,然而天然蛋白纤维产量有限。再生蛋白纤维的制备通常采用与其它成纤高分子接枝或共混的方法,有利于提高再生蛋白纤维的断裂强度。选用同为天然高分子的纤维素为基体,以共溶剂溶解纤维素与蛋白质,进而纺丝成形制得力学性能满足要求的纤维素/丝素蛋白共混纤维。为了探究凝固剂组成对纤维素/丝素蛋白共混纤维相形态及性能的影响,选用水、乙醇、乙醇/1-丁基-3-甲基咪唑氯盐([BMIM]Cl)等作为凝固剂。研究发现:乙醇作为凝固剂时,纤维素与丝素蛋白能很好地同时凝固;而当在乙醇凝固浴中加入适量的[BMIM]Cl径向均匀分散。通过对凝固剂组成的调控能有效提升纤维的力学强度。     

Branched Poly(ε-caprolactone)-Based Copolyesters of Different Architectures and Their Use in the Preparation of Anticancer Drug-Loaded Nanoparticles

Limitations associated with the use of linear biodegradable polyesters in the preparation of anticancer nano-based drug delivery systems (nanoDDS) have turned scientific attention to the utilization of branched-chain (co-)polymers. In this context, the present study evaluates the use of novel branched poly(ε-caprolactone) (PCL)-based copolymers of different architectures for the preparation of anticancer nanoparticle (NP)-based formulations, using paclitaxel (PTX) as a model drug. Specifically, three PCL-polyol branched polyesters, namely, a three-arm copolymer based on glycerol (PCL-GLY), a four-arm copolymer based on pentaerythritol (PCL-PE), and a five-arm copolymer based on xylitol (PCL-XYL), were synthesized via ring-opening polymerization and characterized by proton nuclear magnetic resonance (¹H-NMR), gel permeation chromatography (GPC), intrinsic viscosity, differential scanning calorimetry (DSC), X-ray diffraction (XRD), and Fourier-transform infrared (FT-IR) spectroscopy and cytotoxicity. Then, PTX-loaded NPs were prepared by an oil-in-water emulsion. The size of the obtained NPs varied from 200 to 300 nm, while the drug was dispersed in crystalline form in all formulations. High encapsulation efficiency and high yields were obtained in all cases, while FTIR analysis showed no molecular drug polymer. Finally, in vitro drug release studies showed that the studied nanocarriers significantly enhanced the dissolution rate and extent of the drug.     

Isolation and Self-Association Studies of Beta-Lactoglobulin

The aim of this study was to investigate isolated β-lactoglobulin (β-LG) from the whey protein isolate (WPI) solution using the column chromatography with SP Sephadex. The physicochemical characterization (self-association, the pH stability in various salt solutions, the identification of oligomeric forms) of the protein obtained have been carried out. The electrophoretically pure β-LG fraction was obtained at pH 4.8. The fraction was characterized by the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/TOF MS) technique. The use of the HCCA matrix indicated the presence of oligomeric β-LG forms, while the SA and DHB matrices enabled the differentiation of A and B isoforms in the sample. The impact of sodium chloride, potassium chloride, ammonium sulfate, and sodium citrate in dispersion medium on β-LG electrophoretic stability in solution was also studied. Type of the dispersion medium led to the changes in the isoelectric point of protein. Sodium citrate stabilizes protein in comparison to ammonium sulfate. Additionally, the potential of capillary electrophoresis (CE) with UV detection using bare fused capillary to monitor β-LG oligomerization was discussed. Obtained CE data were further compared by the asymmetric flow field flow fractionation coupled with the multi-angle light scattering detector (AF4-MALS). It was shown that the β-LG is a monomer at pH 3.0, dimer at pH 7.0. At pH 5.0 (near the isoelectric point), oligomers with structures from dimeric to octameric are formed. However, the appearance of the oligomers equilibrium is dependent on the concentration of protein. The higher quantity of protein leads to the formation of the octamer. The far UV circular dichroism (CD) spectra carried out at pH 3.0, 5.0, and 7.0 confirmed that β-sheet conformation is dominant at pH 3.0, 5.0, while at pH 7.0, this conformation is approximately in the same quantity as α-helix and random structures.     

Irisin,a Link among Fatty Liver Disease,Physical Inactivity and Insulin Resistance

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease in industrialized countries. The increasing prevalence of NAFLD mirrors the outbreak of obesity in western countries, highlighting the connection between these two conditions. Nevertheless, there is currently no specific pharmacotherapy for its treatment. Accepted management begins with weight loss and exercise. Moreover, exercise can provide metabolic benefits independently of weight loss. It is known how long-term aerobic training produces improvements in hepatic triglycerides, visceral adipose tissue and free fatty acids, even if there is no weight reduction. A recent study from Boström et al. unravels a potential molecular mechanism that may explain how exercise, independently of weight loss, can potentially improve metabolic parameters through a new messenger system (irisin) linking muscle and fat tissue. Irisin has been proposed to act as a hormone on subcutaneous white fat cells increasing energy expenditure by means of a program of brown-fat-like development. Moreover, it was also shown that irisin plasma concentration was higher in people who exercise, suggesting a molecular mechanism by which exercise may improve metabolism. The present systematic review is based on the possibility that irisin might represent a hypothetical connection between NAFLD pathogenesis and disease progression.