Protocol to study microcircuits in the medial entorhinal cortex in mice using multiple patch-clamp recordings and morphological reconstruction.

Multiple patch-clamp recordings and morphological reconstruction are powerful approaches for neuronal microcircuitry dissection and cell type classification but are challenging due to the sophisticated expertise needed. Here, we present a protocol for applying these techniques to neurons in the medial entorhinal cortex (MEC) of mice. We detail steps to prepare brain slices containing MEC and perform simultaneous multiple whole-cell recordings, followed by procedures of histological staining and neuronal reconstruction. We then describe how we analyze morphological and electrophysiological features. For complete details on the use and execution of this protocol, please refer to Shi et al.1.

Enhancing the Interaction of Carbon Nanotubes by Metal-Organic Decomposition with Improved Mechanical Strength and Ultra-Broadband EMI Shielding Performance.

The remarkable properties of carbon nanotubes (CNTs) have led to promising applications in the field of electromagnetic interference (EMI) shielding. However, for macroscopic CNT assemblies, such as CNT film, achieving high electrical and mechanical properties remains challenging, which heavily depends on the tube-tube interactions of CNTs. Herein, we develop a novel strategy based on metal-organic decomposition (MOD) to fabricate a flexible silver-carbon nanotube (Ag-CNT) film. The Ag particles are introduced in situ into the CNT film through annealing of MOD, leading to enhanced tube-tube interactions. As a result, the electrical conductivity of Ag-CNT film is up to 6.82 × 105 S m-1, and the EMI shielding effectiveness of Ag-CNT film with a thickness of ~ 7.8 µm exceeds 66 dB in the ultra-broad frequency range (3-40 GHz). The tensile strength and Young's modulus of Ag-CNT film increase from 30.09 ± 3.14 to 76.06 ± 6.20 MPa (~ 253%) and from 1.12 ± 0.33 to 8.90 ± 0.97 GPa (~ 795%), respectively. Moreover, the Ag-CNT film exhibits excellent near-field shielding performance, which can effectively block wireless transmission. This innovative approach provides an effective route to further apply macroscopic CNT assemblies to future portable and wearable electronic devices.

Natural polyphenols for drug delivery and tissue engineering construction: A review.

Polyphenols, natural compounds rich in phenolic structures, are gaining prominence due to their antioxidant, anti-inflammatory, antibacterial, and anticancer properties, making them valuable in biomedical applications. Through covalent and noncovalent interactions, polyphenols can bind to biomaterials, enhancing their performance and compensating for their shortcomings. Such polyphenol-based biomaterials not only increase the efficacy of polyphenols but also improve drug stability, control release kinetics, and boost the therapeutic effects of drugs. They offer the potential for targeted drug delivery, reducing off-target impacts and enhancing therapeutic outcomes. In tissue engineering, polyphenols promote cell adhesion, proliferation, and differentiation, thus aiding in the formation of functional tissues. Additionally, they offer excellent biocompatibility and mechanical strength, essential in designing scaffolds. This review explores the significant roles of polyphenols in tissue engineering and drug delivery, emphasizing their potential in advancing biomedical research and healthcare.

Seasonal changes of soil microbiota and its association with environmental factors in coal mining subsidence area.

BACKGROUND: As a special type of wetland, the new wetland in the coal mining subsidence area is highly sensitive to environmental changes. In recent years, more and more attention has been paid to the studies of soil microbial diversity in newly born wetlands in coal mining subsidence areas. However, there are few reports on the seasonal variation of soil microbial diversity and its relationship with soil physical and chemical properties. METHODS: In this study, 16S rRNA gene sequencing technology was used to analyze the seasonal changes of soil microbial composition and functional diversity in newly formed wetlands in coal mining subsidence areas, and to determine the seasonal changes of soil nutrient elements and physical and chemical properties in coal mining subsidence areas, so as to analyze the correlation between soil microbial diversity and soil nutrient elements and physical and chemical properties in newly formed wetlands in coal mining subsidence areas. RESULTS: A total of 16,050 OTUs were obtained after sample gene noise reduction. Proteobacteria, Acidobacteriota and Bacteroidota were the highest abundance in the coal mining subsidence area of Jining. The two seasons gathered separately, and temperature (Temp), total phosphorus (TP), available phosphorus (AP), total organic carbon (TOC) and dry matter content (DMC) were the key factors for the seasonal change of soil microbial community in the wetland of the coal mining subsidence area of Jining. The contents of Temp, AP and TP were significantly correlated with the abundance of soil microorganisms in summer subsidence area, while the contents of DMC and TOC were significantly correlated with the abundance of soil microorganisms in winter subsidence area. CONCLUSION: Soil microbial diversity in coal mining subsidence area was correlated with the seasons. Temp, TP, AP, TOC and DMC were the key factors for the seasonal change of soil microbial community in the wetland of the coal mining subsidence area of Jining.

Lewis base-catalyzed cascade [4 + 2]-annulation reaction of N-alkoxy acrylamides and acyl isothiocyanates: facile access to 2-imino-1,3-thiazinone derivatives.

A formal [4 + 2]-cycloaddition reaction of N-alkoxy acrylamides and acyl isothiocyanates was developed via a Lewis base-catalyzed cascade aza-nucleophilic addition/thio-Michael addition process under mild conditions. This study provides a facile approach for preparing 2-imino-1,3-thiazinone derivatives in moderate to excellent yields and enriches the field of heterocyclic acrylamide chemistry. The reported method features metal-free reaction conditions, high atom economy, and easy operation. Moreover, the reaction was successfully scaled up and derivatization reactions were successfully performed.

A cyclic peptide-based PROTAC induces intracellular degradation of palmitoyltransferase and potently decreases PD-L1 expression in human cervical cancer cells.

Introduction: Our previous research has found that degradation of palmitoyltransferase in tumor cells using a linear peptide PROTAC leads to a significant decrease in PD-L1 expression in tumors. However, this degradation is not a sustained and efficient process. Therefore, we designed a cyclic peptide PROTAC to achieve this efficient anti-PD-L1 effect. Methods: We designed and synthesized an improvement in linear peptide PROTAC targeting palmitoyltransferase DHHC3, and used disulfide bonds to stabilize the continuous N- and C-termini of the peptides to maintain their structure. Cellular and molecular biology techniques were used to test the effect of this cyclic peptide on PD-L1. Results: In human cervical cancer cells, our cyclic peptide PROTAC can significantly downregulate palmitoyl transferase DHHC3 and PD-L1 expressions. This targeted degradation effect is enhanced with increasing doses and treatment duration, with a DC50 value much lower than that of linear peptides. Additionally, flow cytometry analysis of fluorescence intensity shows an increase in the amount of cyclic peptide entering the cell membrane with prolonged treatment time and higher concentrations. The Cellular Thermal Shift Assay (CETSA) method used in this study indicates effective binding between our novel cyclic peptide and DHHC3 protein, leading to a change in the thermal stability of the latter. The degradation of PD-L1 can be effectively blocked by the proteasome inhibitor MG132. Results from clone formation experiments illustrate that our cyclic peptide can enhance the proliferative inhibition effect of cisplatin on the C33A cell line. Furthermore, in the T cell-C33A co-culture system, cyclic peptides target the degradation of PD-L1, thereby blocking the interaction between PD-L1 and PD-1, and promoting the secretion of IFN-γ and TNF-α in the co-culture system supernatant. Conclusion: Our results demonstrate that a disulfide-bridged cyclic peptide PROTAC targeting palmitoyltransferase can provide a stable and improved anti-PD-L1 activity in human tumor cells.

Dual Signal-Enhanced Electrochemiluminescence Strategy Based on Functionalized Biochar for Detecting Aflatoxin B1.

Metal-organic frameworks (MOFs) are often used as carriers in the preparation of electrochemiluminescent (ECL) materials, and ECL materials stabilized in the aqueous phase can be prepared by encapsulating chromophores inside MOFs by an in situ growth method. In this study, nanocomposites MIL-88B(Fe)-NH2@Ru(py)32+ with excellent ECL response were prepared by encapsulating Tris(2,2'-bipyridine)ruthenium dichloride (Ru(py)32+) inside MIL-88B(Fe)-NH2 using the one-step hydrothermal method. MIL-88B(Fe)-NH2 possesses abundant amino groups, which can accelerate the catalytic activation process of K2S2O8, and its abundant pores are also conducive to the enhancement of the transmission rate of co-reactant agents, ions, and electrons, which effectively improves the ECL efficiency. In order to obtain more excellent ECL signals, we prepared aminated biochar (NH2-biochar) using Pu-erh tea dregs as precursor and loaded gold nanoparticles (Au NPs) on its surface as substrate material for modified electrodes. Both NH2-biochar and Au NPs can also be used as a co-reactant promoter to catalyze the activation process of co-reactant K2S2O8. Therefore, a sandwich-type ECL immunosensor was prepared based on a dual signal-enhanced strategy for the highly sensitive and selective detection of aflatoxin B1 (AFB1). Under the optimal experimental conditions, the sensitive detection of AFB1 was achieved in the range of 1 pg·mL-1~100 ng·mL-1 with a detection limit of 209 fg·mL-1. The proposed dual signal-enhanced ECL immunosensor can provide a simple, convenient, and efficient method for the sensitive detection of AFB1 in food and agricultural products.

Two-finger digital rectal examination for the diagnosis of anal fistula: protocol for a randomized controlled trial.

Background: Anal fistula is an anorectal infectious disease caused by a perianal abscess or perianal disease. Accurate anorectal examinations are of great significance. The two-finger digital rectal examination (TF-DRE) has been used in clinical practice, with a lack of comprehensive research on the value of the TF-DRE in the diagnosis of anal fistula. This study will compare the difference in the diagnostic value of the TF-DRE, traditional digital rectal examination (DRE), and anorectal ultrasonography in the diagnosis of anal fistula. Methods: For patients who meet the inclusion criteria, a TF-DRE will be performed to explore the number and location of the external and internal orifices, the number of fistulas, and the relationship between the fistula and the perianal sphincter. A DRE and anorectal ultrasonography will also be performed, and the same data will be recorded. To make a comparison, the final diagnosis results of the clinicians during the operation will be taken as the gold standard, the accuracy of the TF-DRE in diagnosing anal fistula will be calculated, and the significance of the TF-DRE in the preoperative diagnosis of anal fistula will be studied and analyzed. All the statistical results will be analyzed using SPSS22.0 (IBM, USA), and a P value <0.05 will be considered statistically significant. Discussion: The research protocol details the advantages of the TF-DRE compared to the DRE and anorectal ultrasonography in the diagnosis of anal fistula. This study will provide clinical evidence of the diagnostic value of the TF-DRE in the diagnosis of anal fistula. Currently, there is a lack of high-quality research using scientific methods on this innovative anorectal examination method. This study will provide rigorously designed clinical evidence on the TF-DRE. Registration: Chinese Clinical Trials Registry ChiCTR2100045450.

Temperature and phosphorus: the main environmental factors affecting the seasonal variation of soil bacterial diversity in Nansi Lake Wetland.

Introduction: The soil bacteria promote the circulation conversion of lake nutrients and play an important role in maintaining the balance of the lake ecosystem. Few studies have investigated the association of seasonal variation in bacteria and environmental factors in inland freshwater lake wetlands. Nansi Lake is a large shallow freshwater lake in northern China. It is an important hub of the eastern route of the South-to-North Water Diversion Project. Methods: In this study, bacterial 16S rRNA genes were used to analyze the variation of soil bacterial community diversity in Nansi Lake Wetland and its influencing factors in different seasons. Results: It is showed that the phylum, family, and genus with the largest relative abundance in the soil of Nansi Lake Wetland are Proteobacteria, Nitrosomonadaceae, and MND1, respectively. There were significant seasonal differences in soil bacterial diversity in Nansi Lake Wetland, which was significantly higher in summer than in winter. Seasonal variation in environmental factors was significantly correlated with the variation in bacterial communities. Temperature and the content of available phosphorus may be the key factors influencing seasonal variation in bacterial diversity. Discussion: The results of this study further enhance our understanding of the relationship between bacterial community diversity and environmental factors in the lake wetland ecosystem, which can provide scientific data for the conservation of Nansi Lake Wetland.

Laminar and dorsoventral organization of layer 1 interneuronal microcircuitry in superficial layers of the medial entorhinal cortex.

Layer 1 (L1) interneurons (INs) participate in various brain functions by gating information flow in the neocortex, but their role in the medial entorhinal cortex (MEC) is still unknown, largely due to scant knowledge of MEC L1 microcircuitry. Using simultaneous triple-octuple whole-cell recordings and morphological reconstructions, we comprehensively depict L1IN networks in the MEC. We identify three morphologically distinct types of L1INs with characteristic electrophysiological properties. We dissect intra- and inter-laminar cell-type-specific microcircuits of L1INs, showing connectivity patterns different from those in the neocortex. Remarkably, motif analysis reveals transitive and clustered features of L1 networks, as well as over-represented trans-laminar motifs. Finally, we demonstrate the dorsoventral gradient of L1IN microcircuits, with dorsal L1 neurogliaform cells receiving fewer intra-laminar inputs but exerting more inhibition on L2 principal neurons. These results thus present a more comprehensive picture of L1IN microcircuitry, which is indispensable for deciphering the function of L1INs in the MEC.

Stapled peptide PROTAC induced significantly greater anti-PD-L1 effects than inhibitor in human cervical cancer cells.

Introduction: Immune checkpoint inhibitors (ICIs) are monoclonal antibodies that target immune checkpoints that suppress immune cell activity. Low efficiency and high resistance are currently the main barriers to their clinical application. As a representative technology of targeted protein degradation, proteolysis-targeting chimeras (PROTACs) are considered to have potential for addressing these limitations. Methods: We synthesized a stapled peptide-based PROTAC (SP-PROTAC) that specifically targeted palmitoyltransferase ZDHHC3 and resulted in the decrease of PD-L1 in human cervical cancer cell lines. Flow cytometry, confocal microscopy, protein immunoblotting, Cellular Thermal Shift Assay (CETSA), and MTT assay analyses were conducted to evaluate the effects of the designed peptide and verify its safety in human cells. Results: In cervical cancer celllines C33A and HeLa, the stapled peptide strongly downregulated PD-L1 to < 50% of baseline level at 0.1 µM. DHHC3 expression decreased in both dosedependentand time-dependent manners. MG132, the proteasome inhibitor, can alleviate the SP-PROTAC mediated degradation of PD-L1 in human cancer cells. In a co-culture model of C33A and T cells, treatment with the peptide induced IFN-γ and TNF-α release in a dose-dependent manner by degrading PD-L1. These effects were more significant than that of the PD-L1 inhibitor, BMS-8. Conclusions: Cells treated with 0.1 µM of SP-PROTAC or BMS-8 for 4 h revealed that the stapled peptide decreased PD-L1 more effectively than BMS-8. DHHC3-targeting SP-PROTAC decreased PD-L1 in human cervical cancer more effectively than the inhibitor BMS-8.

MPI-VGAE: protein-metabolite enzymatic reaction link learning by variational graph autoencoders.

Enzymatic reactions are crucial to explore the mechanistic function of metabolites and proteins in cellular processes and to understand the etiology of diseases. The increasing number of interconnected metabolic reactions allows the development of in silico deep learning-based methods to discover new enzymatic reaction links between metabolites and proteins to further expand the landscape of existing metabolite-protein interactome. Computational approaches to predict the enzymatic reaction link by metabolite-protein interaction (MPI) prediction are still very limited. In this study, we developed a Variational Graph Autoencoders (VGAE)-based framework to predict MPI in genome-scale heterogeneous enzymatic reaction networks across ten organisms. By incorporating molecular features of metabolites and proteins as well as neighboring information in the MPI networks, our MPI-VGAE predictor achieved the best predictive performance compared to other machine learning methods. Moreover, when applying the MPI-VGAE framework to reconstruct hundreds of metabolic pathways, functional enzymatic reaction networks and a metabolite-metabolite interaction network, our method showed the most robust performance among all scenarios. To the best of our knowledge, this is the first MPI predictor by VGAE for enzymatic reaction link prediction. Furthermore, we implemented the MPI-VGAE framework to reconstruct the disease-specific MPI network based on the disrupted metabolites and proteins in Alzheimer's disease and colorectal cancer, respectively. A substantial number of novel enzymatic reaction links were identified. We further validated and explored the interactions of these enzymatic reactions using molecular docking. These results highlight the potential of the MPI-VGAE framework for the discovery of novel disease-related enzymatic reactions and facilitate the study of the disrupted metabolisms in diseases.

Genome-scale enzymatic reaction prediction by variational graph autoencoders.

Background: Enzymatic reaction networks are crucial to explore the mechanistic function of metabolites and proteins in biological systems and understanding the etiology of diseases and potential target for drug discovery. The increasing number of metabolic reactions allows the development of deep learning-based methods to discover new enzymatic reactions, which will expand the landscape of existing enzymatic reaction networks to investigate the disrupted metabolisms in diseases. Results: In this study, we propose the MPI-VGAE framework to predict metabolite-protein interactions (MPI) in a genome-scale heterogeneous enzymatic reaction network across ten organisms with thousands of enzymatic reactions. We improved the Variational Graph Autoencoders (VGAE) model to incorporate both molecular features of metabolites and proteins as well as neighboring features to achieve the best predictive performance of MPI. The MPI-VGAE framework showed robust performance in the reconstruction of hundreds of metabolic pathways and five functional enzymatic reaction networks. The MPI-VGAE framework was also applied to a homogenous metabolic reaction network and achieved as high performance as other state-of-art methods. Furthermore, the MPI-VGAE framework could be implemented to reconstruct the disease-specific MPI network based on hundreds of disrupted metabolites and proteins in Alzheimer's disease and colorectal cancer, respectively. A substantial number of new potential enzymatic reactions were predicted and validated by molecular docking. These results highlight the potential of the MPI-VGAE framework for the discovery of novel disease-related enzymatic reactions and drug targets in real-world applications. Data availability and implementation: The MPI-VGAE framework and datasets are publicly accessible on GitHub https://github.com/mmetalab/mpi-vgae . Author Biographies: Cheng Wang received his Ph.D. in Chemistry from The Ohio State Univesity, USA. He is currently a Assistant Professor in School of Public Health at Shandong University, China. His research interests include bioinformatics, machine learning-based approach with applications to biomedical networks. Chuang Yuan is a research assistant at Shandong University. He obtained the MS degree in Biology at the University of Science and Technology of China. His research interests include biochemistry & molecular biology, cell biology, biomedicine, bioinformatics, and computational biology. Yahui Wang is a PhD student in Department of Chemistry at Washington University in St. Louis. Her research interests include biochemistry, mass spectrometry-based metabolomics, and cancer metabolism. Ranran Chen is a master graduate student in School of Public Health at University of Shandong, China. Yuying Shi is a master graduate student in School of Public Health at University of Shandong, China. Gary J. Patti is the Michael and Tana Powell Professor at Washington University in St. Louis, where he holds appointments in the Department of Chemisrty and the Department of Medicine. He is also the Senior Director of the Center for Metabolomics and Isotope Tracing at Washington University. His research interests include metabolomics, bioinformatics, high-throughput mass spectrometry, environmental health, cancer, and aging. Leyi Wei received his Ph.D. in Computer Science from Xiamen University, China. He is currently a Professor in School of Software at Shandong University, China. His research interests include machine learning and its applications to bioinformatics. Qingzhen Hou received his Ph.D. in the Centre for Integrative Bioinformatics VU (IBIVU) from Vrije Universiteit Amsterdam, the Netherlands. Since 2020, He has serveved as the head of Bioinformatics Center in National Institute of Health Data Science of China and Assistant Professor in School of Public Health, Shandong University, China. His areas of research are bioinformatics and computational biophysics. Key points: Genome-scale heterogeneous networks of metabolite-protein interaction (MPI) based on thousands of enzymatic reactions across ten organisms were constructed semi-automatically.An enzymatic reaction prediction method called Metabolite-Protein Interaction Variational Graph Autoencoders (MPI-VGAE) was developed and optimized to achieve higher performance compared with existing machine learning methods by using both molecular features of metabolites and proteins.MPI-VGAE is broadly useful for applications involving the reconstruction of metabolic pathways, functional enzymatic reaction networks, and homogenous networks (e.g., metabolic reaction networks).By implementing MPI-VGAE to Alzheimer's disease and colorectal cancer, we obtained several novel disease-related protein-metabolite reactions with biological meanings. Moreover, we further investigated the reasonable binding details of protein-metabolite interactions using molecular docking approaches which provided useful information for disease mechanism and drug design.

Enhancing the Chemical Stability of MXene Through Synergy of Hydrogen Bond and Coordination Bond in Aqueous Solution.

MXenes with unique physicochemical properties have shown substantial potential in electromagnetic interference (EMI) shielding. However, the chemical instability and mechanical fragility of MXenes has become a major hurdle for their application. Abundant strategies have been dedicated to improving the oxidation stability of colloidal solution or mechanical properties of films, which always come at the expense of electrical conductivity and chemical compatibility. Here, hydrogen bond (H-bond) and coordination bond are employed to achieve chemical and colloidal stability of MXenes (0.1 mg mL-1 ) by occupying the reaction sites of Ti3 C2 Tx attacking of water and oxygen molecules. Compared to the Ti3 C2 Tx , the Ti3 C2 Tx modified with alanine via H-bond shows significantly improved oxidation stability (at room temperature over 35 days), while the Ti3 C2 Tx modified with cysteine by synergy of H-bond and coordination bond can be maintained even after 120 days. Simulation and experimental results verify the formation of H-bond and Ti-S bond by a Lewis acid-base interaction between Ti3 C2 Tx and cysteine. Furthermore, the synergy strategy significantly improves the mechanical strength of the assembled film (up to 78.1 ± 7.9 MPa), corresponding the increment of 203% compared to untreated one, almost without compromising the electrical conductivity and EMI shielding performance.

PROTACs in Epigenetic Cancer Therapy: Current Status and Future Opportunities.

The epigenetic regulation of gene functions has been proven to be strongly associated with the development and progression of cancer. Reprogramming the cancer epigenome landscape is one of the most promising target therapies in both treatments and in reversing drug resistance. Proteolytic targeted chimeras (PROTACs) are an emerging therapeutic modality for selective degradation via the native ubiquitin-proteasome system. Rapid advances in PROTACs have facilitated the exploration of targeting epigenetic proteins, a lot of PROTAC degraders have already been designed in the field of epigenetic cancer therapy, and PROTACs targeting epigenetic proteins can better exploit target druggability and improve the mechanistic understanding of the epigenetic regulation of cancer. Thus, this review focuses on the progress made in the development of PROTAC degraders and PROTAC drugs targeting epigenetics in cancer and discusses challenges and future opportunities for the field.

Restricted cubic spline model analysis of the association between anal fistula and anorectal abscess incidence and body mass index.

Objective: The epidemiological profile of anal fistula and anorectal abscess has not been well studied. Based on the results of a retrospective cross-sectional survey, we aimed to investigate the potential influential factors associated with anal fistula and anorectal abscess. Methods: We conducted a retrospective analysis of outpatients who visited the proctology department at China-Japan Friendship Hospital between January 2017 and May 2022. A comprehensive questionnaire was designed to collect potential influential factors, and according to formal anorectal examination and the corresponding diagnostic criteria, all the participants were divided into patients with anal fistula or perianal abscess and healthy control group. Multiple logistic regression was used to identify factors in significant association with anal fistula and perianal abscess. Additionally, we combined restricted cubic spline regression to examine the dose-response relationship between factors and the risk of developing anal fistula or anorectal abscess. Results: The present study included 1,223 participants, including 1,018 males and 206 females, with 275 anal fistulas, 184 anorectal abscesses, and 765 healthy controls. We found no statistically significant differences between patients and controls in basic information and preoperative assessment of life factors, except for body mass index. It was indicated that people with overweight or obesity were more prone to anal fistula (OR overweight = 1.35, 95% CI: 1.00-1.82, P = 0.047; OR obesity = 3.44, 95% CI: 2.26-5.26, P < 0.001) or anorectal abscess (OR overweight = 1.41, 95% CI: 1.00-1.99, P = 0.05; OR obesity: 2.24, 95% CI: 1.37-3.67, P = 0.001) than normal-weight individuals. The dose-response research indicated the J-shaped trend between the ascending BMI levels and the higher risk of suffering from anal fistula and anorectal abscess. Conclusions: Our findings indicate that overweight and obesity are risk factors for anal fistula and anorectal abscess, which plays a role in the prevention of anorectal diseases. This provides some theoretical basis for clinicians to provide health education to their patients.

Gut microbiome of captive wolves is more similar to domestic dogs than wild wolves indicated by metagenomics study.

Adaptation during the domestication from wolves (Canis lupus) to dogs (Canis lupus familiaris) is a debated ecological topic. Changes in food and environment are major divergences in the domestication of dogs. Gut microbes play an important role in animal adaptation to the food and environmental changes. In this study, shotgun sequencing was performed to compare the species diversity and functional diversity of gut microbes in wild wolves (group CLW, n = 3), captive wolves (group CLC, n = 4), and domestic dogs (group CLF, n = 4). The results found that Bacteroidetes, Firmicutes, Fusobacteria, Proteobacteria and Actinobacteria were the most abundant phyla and Bacteroides, Fusobacterium, Prevotella, Megamonas, Paraprevotella, Faecalibacterium, Clostridium were the most abundant genera in the gut of wolves and dogs. Groups CLW, CLC and CLF have shown significant difference in gut microbial species diversity and functional diversity. Bacteroides, Fusobacterium and Faecalibacterium were most abundant genera in groups CLW, CLC and CLF, respectively. Their abundance varied significantly among groups. Compared to the wild wolves, the intestinal microbiol genes of domestic dogs were significantly enriched in the carbohydrate metabolism pathway of KEGG database. One hundred and seventy-seven enzymes were detected with significantly higher abundance in group CLF than that in group CLW, and 49 enzymes showed extremely significant higher abundance in group CLF than that in group CLW (q < 0.01) base on the function abundance annotated in CAZy database. It is noteworthy that there were also significant differences in the abundance of 140 enzymes between groups CLC and CLW (q < 0.05). Clustering analysis based on both the species and the function abundance of intestinal microbiota all found that groups CLC and CLF clustered into one branch, while samples from group CLW clustered into the other branch. This result suggests that captive wolves are more similar to domestic dogs than wild wolves in both species composition and function composition of intestinal microbiota.

Icaritin, a metabolite of Icarrin, Alleviates non-alcoholic fatty liver disease via inhibition of lipogenesis and ER stress.

Non-alcoholic fatty liver disease (NAFLD) is one of the most serious global public health concerns. However, there are currently no effective drugs for treatment of this disease. Icariin (ICA), a small-molecule natural product extracted from Epimedium brevicornu Maxim, offers various pharmacological activities. In the present work, we wondered whether ICA can attenuate NAFLD in db/db mice treated with ICA for 8 weeks and how ICA exerts an influence on NAFLD. In db/db mice, ICA treatment had a robust effect on inhibition of lipogenesis associated with NAFLD amelioration by decreasing liver lipid deposition, together with ameliorating insulin sensitivity, glucose tolerance, and fasting serum glucose. Of note, ICA-treated rats showed a much higher concentration of icaritin (ICT) in plasma, a major metabolite of ICA, about 2000 times higher than that of ICA by liquid chromatography mass spectrometry (LC-MS). Interestingly, ICT, rather than ICA, can dramatically decrease hepatic lipogenesis-related markers in oleate acid/palmitate acid (OA/PA)-induced steatosis in primary hepatocytes (PH) and HepG2 cells, and hepatic lipid accumulation in db/db mice, demonstrating the inhibitory effect of ICT on lipogenesis. Mechanistically, we found that anti-lipogenic activities of ICT were related to reducing endoplasmic reticulum (ER) stress as evidenced by Western blot, qPCR, and other assays in thapsigargin (THP) induced-ER stress models. To our knowledge, this is the first report showing the unexpected and key role for ICT on the prevention of NAFLD in db/db mice through an ER stress mechanism.

Preparation of hydrophobic composite membranes based on carboxymethyl cellulose and modified pectin: Effects of grafting a long-chain saturated fatty acid.

In this study, docosanoic acid, a very long chain fatty acid, was used to modify pectin, and the products were incorporated with carboxylmethyl cellulose (CMC) to prepare a hydrophobic composite. Results of structural characterisation showed that docosanoic acid was grafted to pectin through the esterification reaction, and the highest grafting ratio was 7.89 %. After grafting with docosanoic acid, the emulsifying activity and stability of pectin were significantly enhanced from 1.23 × 10-2 and 9.27 % to 4.78 × 10-2 and 26.73 %. Moreover, when modified pectin was blended with CMC instead of native pectin, the hydrophobicity of the composite membranes increased significantly. In detail, the highest contact angle of the composite membrane incorporated with modified pectin was 97.6°, which was much higher than that with native pectin (68.9°). As the grafting ratio of pectin increased, the water vapor permeability of the composite membranes significantly increased, while the water absorption decreased. Furthermore, the mechanical properties and transparency of the composite membranes could be improved by grafting docosanoic acid into pectin. All the results indicated that incorporating docosanoic acid possibly helped improve the comprehensive properties of the composite membranes based on polysaccharides and expand their application in food packaging.