Discovery of novel alkaline-tolerant xylanases from fecal microbiota of dairy cows.

Xylo-oligosaccharides (XOS) are considered as a promising type of prebiotics that can be used in foods, feeds, and healthcare products. Xylanases play a key role in the production of XOS from xylan. In this study, we conducted a metagenomic analysis of the fecal microbiota from dairy cows fed with different types of fodders. Despite the diversity in their diets, the main phyla observed in all fecal microbiota were Firmicutes and Bacteroidetes. At the genus level, one group of dairy cows that were fed probiotic fermented herbal mixture-containing fodders displayed decreased abundance of Methanobrevibacter and increased growth of beneficial Akkermansia bacteria. Additionally, this group exhibited a high microbial richness and diversity. Through our analysis, we obtained a comprehensive dataset comprising over 280,000 carbohydrate-active enzyme genes. Among these, we identified a total of 163 potential xylanase genes and subsequently expressed 34 of them in Escherichia coli. Out of the 34 expressed genes, two alkaline xylanases with excellent temperature stability and pH tolerance were obtained. Notably, CDW-xyl-8 exhibited xylanase activity of 96.1 ± 7.5 U/mg protein, with an optimal working temperature of 55 â„ƒ and optimal pH of 8.0. CDW-xyl-16 displayed an activity of 427.3 ± 9.1 U/mg protein with an optimal pH of 8.5 and an optimal temperature at 40 â„ƒ. Bioinformatic analyses and structural modeling suggest that CDW-xyl-8 belongs to GH10 family xylanase, and CDW-xyl-16 is a GH11 family xylanase. Both enzymes have the ability to hydrolyze beechwood xylan and produce XOS. In conclusion, this metagenomic study provides valuable insights into the fecal microbiota composition of dairy cows fed different fodder types, revealing main microbial groups and demonstrating the abundance of xylanases. Furthermore, the characterization of two novel xylanases highlights their potential application in XOS production.

Preparation of imidazole acids grafted chitosan with enhanced antioxidant, antibacterial and antitumor activities.

Herein, imidazole acids grafted chitosan derivatives were synthesized, including HACC, HACC derivatives, TMC, TMC derivatives, amidated chitosan and amidated chitosan bearing imidazolium salts. The prepared chitosan derivatives were characterized by FT-IR and 1H NMR. The tests evaluated the biological antioxidant, antibacterial, and cytotoxic activities of chitosan derivatives. The antioxidant capacity (DPPH radical, superoxide anion radical and hydroxyl radical) of chitosan derivatives was 2.4-8.3 times higher than that of chitosan. The antibacterial capacity against E. coli and S. aureus of cationic derivatives (HACC derivatives, TMC derivatives, and amidated chitosan bearing imidazolium salts) was more active than only imidazole-chitosan (amidated chitosan). In particular, the inhibition effect of HACC derivatives on E. coli was 15.625 µg/mL. Moreover, the series of chitosan derivatives bearing imidazole acids showed certain activity against MCF-7 and A549 cells. The present results suggest that the chitosan derivatives in this paper seem to be promising carrier materials for use in drug delivery systems.

Serum metabolomics analysis in patients with alcohol dependence.

Objective: Alcohol dependence (AD) is a chronic recurrent mental disease caused by long-term drinking. It is one of the most prevalent public health problems. However, AD diagnosis lacks objective biomarkers. This study was aimed to shed some light on potential biomarkers of AD patients by investigating the serum metabolomics profiles of AD patients and the controls. Methods: Liquid chromatography-mass spectrometry (LC-MS) was used to detect the serum metabolites of 29 AD patients (AD) and 28 controls. Six samples were set aside as the validation set (Control: n = 3; AD group: n = 3), and the remaining were used as the training set (Control: n = 26; AD group: n = 25). Principal component analysis (PCA) and partial least squares discriminant analysis (PCA-DA) were performed to analyze the training set samples. The metabolic pathways were analyzed using the MetPA database. The signal pathways with pathway impact >0.2, value of p <0.05, and FDR < 0.05 were selected. From the screened pathways, the metabolites whose levels changed by at least 3-fold were screened. The metabolites with no numerical overlap in their concentrations in the AD and the control groups were screened out and verified with the validation set. Results: The serum metabolomic profiles of the control and the AD groups were significantly different. We identified six significantly altered metabolic signal pathways, including protein digestion and absorption; alanine, aspartate, and glutamate metabolism; arginine biosynthesis; linoleic acid metabolism; butanoate metabolism; and GABAergic synapse. In these six signal pathways, the levels of 28 metabolites were found to be significantly altered. Of these, the alterations of 11 metabolites changed by at least 3-fold compared to the control group. Of these 11 metabolites, those with no numerical overlap in their concentrations between the AD and the control groups were GABA, 4-hydroxybutanoic acid, L-glutamic acid, citric acid and L-glutamine. Conclusion: The metabolite profile of the AD group was significantly different from that of the control group. GABA, 4-hydroxybutanoic acid, L-glutamic acid, citric acid, and L-glutamine could be used as potential diagnostic markers for AD.

Safety profile of 0.0015% tafluprost eye drops in China: a post-marketing observational study.

AIM: To investigate the treatment pattern and safety of tafluprost for glaucoma and ocular hypertension (OH) in clinical practice in China. METHODS: This post-marketing observational study included patients who received tafluprost to lower intraocular pressure (IOP) within 30d between September 2017 and March 2020 in 20 hospitals in China. Adverse drug reactions (ADRs) during tafluprost treatment and within 30d after the treatment were collected. RESULTS: A total of 2544 patients were included in this study, of them 58.5% (1488/2544) had primary open angle glaucoma (POAG), 21.9% (556/2544) had OH and 19.7% (500/2544) used tafluprost for other reasons. Of 359 ADRs occurred in 10.1% (258/2544) patients, and no serious adverse event occurred. The most common ADR was conjunctival hyperemia (128 ADRs in 124 patients, 4.9%). Totally 1670 participants (65.6%) combined tafluprost with carbonic anhydrase inhibitors (CAIs; 37.1%, 620/1670), sympathomimetics (33.5%, 559/1670), ß-blockers (33.2%, 555/1670), other prostaglandin analogs (PGAs; 15.6%, 260/1670) and other eye drops (15.1%, 253/1670). The highest incidence of conjunctival hyperemia was noted in patients who received tafluprost in combination with other PGAs (23 ADRs in 23 patients, 8.8%, 23/260) and the lowest was in combination with CAIs (16 ADRs in 16 patients, 2.6%, 16/620). Tafluprost was applied in primary angle-closure glaucoma (41.6%, 208/500), after glaucoma surgery (17.8%, 89/500) and after non-glaucoma surgery (15.8%, 79/500). CONCLUSION: Tafluprost is safe for POAG and OH, and tolerable when combined with other eye drops and under various clinical circumstances.

The protective effects and mechanisms of melatonin on white matter damage in neonatal rats / 中国新生儿科杂志

Objective:To study the protective effects and mechanisms of melatonin (MTn) on lipopolysaccharide (LPS) and hypoxic-ischemic(HI) induced white matter damage (WMD) in neonatal rats.Methods:Seventy-two 3-day-old newborn Sprague-Dawley (SD) rats were randomly assigned into sham operation group (the sham group), model group (the HI group) and MTn intervention group (the HI+MTn group) ( n=24 for each group). For the sham group, only dissection of the right common carotid artery was performed without ligation. Animal models of WMD were established using LPS pretreatment and HI method in both the HI group and HI+MTn group. The HI+MTn group received MTn intraperitoneal injection (15 mg/kg, 1 h before LPS injection and then once daily). The HI group and the sham group received equal volume of normal saline containing 1% ethanol intraperitoneal injection. The rats were sacrificed on d7 of experiment and periventricular white matter (PVWM) was collected for hematoxylin-eosin (HE) and TUNEL staining to determine WMD and apoptosis. The distribution and morphology of microglial cells in the PVWM were studied using IBA1 immunofluorescence staining. Reactive oxygen species (ROS) kit was used to detect ROS. The expression of nucleotide-binding domain-like receptor protein 3 (NLRP3) inflammasomes, interleukin (IL)-1β, IL-18 and mitochondrial autophagy markers (pink1 and parkin) were determined using real-time quantitative PCR. Results:Compared with the sham group, the HI group showed WMD, cell degeneration and necrosis,increased cell apoptosis and increased expressions of NLRP3 inflammasomes and downstream inflammatory factors (IL-1β and IL-18) in PVWM. Compared with the HI group,the HI+MTn group showed reduced WMD, cell apoptosis, microglia infiltration and inflammatory factors expression. MTn increased pink1 and parkin expression and reduced ROS production in PVWM.Conclusions:MTn reduces ROS production by enhancing mitochondrial autophagy and inhibits NLRP3 inflammasomes hyperactivation to alleviate endotoxin- and HI-induced WMD in neonatal rats.

Preparation of l-Arginine Schiff Bases Modified Chitosan Derivatives and Their Antimicrobial and Antioxidant Properties.

We successfully prepared a series of l-arginine Schiff bases acylated chitosan derivatives, aiming to improve the antioxidant activity and antimicrobial activity of chitosan by introducing a furan ring, pyridine ring, and l-arginine structure. The accuracy of the structures of ten compounds was characterized by FT-IR and 1H NMR. In terms of DPPH radical scavenging activity, except for compound CR3PCA, the scavenging rate of other compounds was higher than chitosan, especially CRCF and CRBF had strong scavenging abilities. At the same time, in the superoxide-radical scavenging activity assay, CRCF, CRBF, CR3PCA, CR2C3PCA, and CR2B3PCA were comparable to positive control at 1.60 mg/mL. Simultaneously, CRFF, CRCF, and CRBF had a certain inhibitory effect on Botrytis cinerea. Furthermore, the inhibitory effect of CRFF, CRCF, and CR3PCA on Staphylococcus aureus was very well, close to the positive control at 1.00 mg/mL. CRCF and CR2B3PCA showed better inhibitory effects on Escherichia coli than other compounds. The MTT assay was used to determine the cytotoxicity of the chitosan derivatives, which proved their safety to fibroblast cells. In summary, the study indicated that some of these compounds have the potential for further development and utilization in the preparation of antioxidants and antimicrobial agents.

Characterization of Novel Pectinolytic Enzymes Derived from the Efficient Lignocellulose Degradation Microbiota.

Diverse pectinolytic enzymes are widely applied in the food, papermaking, and other industries, and they account for more than 25% of the global industrial enzyme demands. Efficient lignocellulose degradation microbiota are reservoirs of pectinolytic enzymes and other lignocellulose-degrading genes. Metagenomics has been widely used to discover new pectinolytic enzymes. Here, we used a metagenomic strategy to characterize pectinolytic genes from one efficient lignocellulose-degrading microbiota derived from pulp and paper wastewater treatment microbiota. A total of 23 predicted full-length GH28 and PL1 family pectinolytic genes were selectively cloned and expressed in Escherichia coli, and 5 of the expressed proteins had pectinolytic activities. Among them, the characterization of one pectinolytic enzyme, PW-pGH28-3, which has a 58.4% identity with an exo-polygalacturonase gene of Aquipluma nitroreducens, was further investigated. The optimal pH and optimal temperature of PW-pGH28-3 were 8.0 and 40 °C, respectively, and its pectinolytic activity at the optimal condition was 13.5 ± 1.1 U/mg protein. Bioinformatics analyses and structural modeling suggest that PW-pGH28-3 is a novel secretory exo-polygalacturonase, which is confirmed by its hydrolysates of polygalacturonic acid. The detection of PW-pGH28-3 and other pectinolytic genes showed that efficient lignocellulose degradation microbiota could provide potential efficient pectinolytic enzymes for industrial application. In the future, improving metagenomic screening efficiency would discover efficient lignocellulose-degrading enzymes and lead to the sustainable and green utilization of lignocellulose.

Preparation, Anticoagulant and Antioxidant Properties of Glucosamine-Heparin Salt.

Excessive inorganic ions in vivo may lead to electrolyte disorders and induce damage to the human body. Therefore, preparation of enhanced bioactivity compounds, composed of activated organic cations and organic anions, is of great interest among researchers. In this work, glucosamine-heparin salt (GHS) was primarily synthesized with positively charged glucosamine hydrochloride (GAH) and negatively charged heparin sodium (Heps) by ion exchange method. Then, the detailed structural information of the GHS was characterized by FTIR, 1H NMR spectroscopy and ICP-MS. In addition, its anticoagulant potency and antioxidant properties were evaluated, respectively. The results demonstrated that GHS salt achieved enhanced antioxidant activities, including 98.78% of O2•- radical scavenging activity, 91.23% of •OH radical scavenging rate and 66.49% of DPPH radical scavenging capacity at 1.6 mg/mL, severally. Meanwhile, anticoagulant potency (ATTP) of GHS strengthened from 153.10 ± 17.14 to 180.03 ± 6.02 at 0.75 µmol/L. Thus, introducing cationic glucosamine residues into GHS could improve its anticoagulant activity. The findings suggest that GHS product with a small amount of inorganic ions can greatly abate the prime cost of antioxidants and anticoagulants, and has significant economic benefits and practical significance.

Preparation of Doxorubicin-Loaded Carboxymethyl-ß-Cyclodextrin/Chitosan Nanoparticles with Antioxidant, Antitumor Activities and pH-Sensitive Release.

In this study, chitosan nanoparticles (HF-CD NPs) were synthesized by an ionic gelation method using negatively charged carboxymethyl-ß-cyclodextrin and positively charged 2-hydroxypropyltrimethyl ammonium chloride chitosan bearing folic acid. The surface morphology of HF-CD NPs was spherical or oval, and they possessed relatively small particle size (192 ± 8 nm) and positive zeta potential (+20 ± 2 mV). Meanwhile, doxorubicin (Dox) was selected as model drug to investigate the prepared nanoparticles' potential to serve as a drug delivery carrier. The drug loading efficiency of drug-loaded nanoparticles (HF-Dox-CD NPs) was 31.25%. In vitro release profiles showed that Dox release of nanoparticles represented a pH-sensitive sustained and controlled release characteristic. At the same time, the antioxidant activity of nanoparticles was measured, and chitosan nanoparticles possessed good antioxidant activity and could inhibit the lipid peroxidation inside the cell and avoid material infection. Notably, CCK-8 assay testified that the nanoparticles were safe drug carriers and significantly enhanced the antitumor activity of Dox. The nanoparticles possessed good antioxidant activity, pH-sensitive sustained controlled release, enhanced antitumor activity, and could be expected to serve as a drug carrier in future with broad application prospects.

Enhanced antifungal and antioxidant activities of new chitosan derivatives modified with Schiff base bearing benzenoid/heterocyclic moieties.

In this work, chitosan derivatives modified with Schiff base bearing benzenoid/heterocyclic moieties were successfully prepared via amidation reaction. Specific structural characterization was implemented using FTIR and 1H NMR, and the DS of chitosan derivatives were quantitatively calculated by ratio of hydrogen proton integral. Meanwhile, the antifungal activity against two common plant pathogenic fungi (Fusarium oxysporum f. sp. cubense and Glomerella cingulata) was assayed in vitro by hyphal measurement, and data proved that the introduction of functional groups including benzene/heterocyclic compounds and Schiff base groups greatly enhanced the antifungal activity. Besides, the antioxidant efficiency was investigated in vitro, and all chitosan derivatives exhibited significantly increased antioxidant activity. Specially, the scavenging effect of 2SATCS was 96.62% at 1.6 mg/mL, which was close to the positive control VC (98.84%). These results indicated that chitosan derivatives with enhanced antifungal and antioxidant activities could serve as potential biomaterial for antifungal and antioxidant applications.

Antimicrobial and Antioxidant Activities of N-2-Hydroxypropyltrimethyl Ammonium Chitosan Derivatives Bearing Amino Acid Schiff Bases.

N-2-hydroxypropyltrimethyl ammonium chloride chitosan (HACC), a cationic quaternary ammonium salt polymer exhibiting good solubility in water, is widely used because of its low toxicity and good biocompatibility. Herein, through ion exchange reaction, we prepared N-2-hydroxypropyltrimethyl ammonium chitosan derivatives bearing amino acid Schiff bases with good biological activities. The accuracy of the structures was verified by FT-IR and 1H NMR. The antibacterial activity, antifungal activity, and scavenging ability of DPPH radical and superoxide radical of HACC derivatives were significantly improved compared with that of HACC. In particular, HACGM (HACC-potassium 2-((2-hydroxy-3-methoxybenzylidene)amino)acetate) and HACGB (HACC-potassium 2-((5-bromo-2-hydroxybenzylidene)amino)acetate) showed good inhibitory effect on bacteria and fungi, including Staphylococcus aureus, Escherichia coli, Botrytis cinerea, and Fusarium oxysporum f. sp. cubense. The inhibition rate of HACGB on Staphylococcus aureus and Escherichia coli could reach 100% at the concentration of 0.1 mg/mL, and the inhibition rate of HACGM and HACGB on Botrytis cinerea and Fusarium oxysporum f. sp. cubense could also reach 100% at the concentration of 0.5 mg/mL. Improving antimicrobial and antioxidant activities of HACC could provide ideas and experiences for the development and utilization of chitosan derivatives.

Spatiotemporal Evolution of the Global Species Diversity of Rhododendron.

Evolutionary radiation is a widely recognized mode of species diversification, but its underlying mechanisms have not been unambiguously resolved for species-rich cosmopolitan plant genera. In particular, it remains largely unknown how biological and environmental factors have jointly driven its occurrence in specific regions. Here, we use Rhododendron, the largest genus of woody plants in the Northern Hemisphere, to investigate how geographic and climatic factors, as well as functional traits, worked together to trigger plant evolutionary radiations and shape the global patterns of species richness based on a solid species phylogeny. Using 3,437 orthologous nuclear genes, we reconstructed the first highly supported and dated backbone phylogeny of Rhododendron comprising 200 species that represent all subgenera, sections, and nearly all multispecies subsections, and found that most extant species originated by evolutionary radiations when the genus migrated southward from circumboreal areas to tropical/subtropical mountains, showing rapid increases of both net diversification rate and evolutionary rate of environmental factors in the Miocene. We also found that the geographically uneven diversification of Rhododendron led to a much higher diversity in Asia than in other continents, which was mainly driven by two environmental variables, that is, elevation range and annual precipitation, and were further strengthened by the adaptation of leaf functional traits. Our study provides a good example of integrating phylogenomic and ecological analyses in deciphering the mechanisms of plant evolutionary radiations, and sheds new light on how the intensification of the Asian monsoon has driven evolutionary radiations in large plant genera of the Himalaya-Hengduan Mountains.

Developments in Fatty Acid-Derived Insect Pheromone Production Using Engineered Yeasts.

The use of traditional chemical insecticides for pest control often leads to environmental pollution and a decrease in biodiversity. Recently, insect sex pheromones were applied for sustainable biocontrol of pests in fields, due to their limited adverse impacts on biodiversity and food safety compared to that of other conventional insecticides. However, the structures of insect pheromones are complex, and their chemical synthesis is not commercially feasible. As yeasts have been widely used for fatty acid-derived pheromone production in the past few years, using engineered yeasts may be promising and sustainable for the low-cost production of fatty acid-derived pheromones. The primary fatty acids produced by Saccharomyces cerevisiae and other yeasts are C16 and C18, and it is also possible to rewire/reprogram the metabolic flux for other fatty acids or fatty acid derivatives. This review summarizes the fatty acid biosynthetic pathway in S. cerevisiae and recent progress in yeast engineering in terms of metabolic engineering and synthetic biology strategies to produce insect pheromones. In the future, insect pheromones produced by yeasts might provide an eco-friendly pest control method in agricultural fields.

New synthetic adriamycin-incorporated chitosan nanoparticles with enhanced antioxidant, antitumor activities and pH-sensitive drug release.

In this paper, adriamycin-incorporated chitosan nanoparticles were synthesized by ionic gelation using negatively charged carboxymethyl chitosan and positively charged 2-hydroxypropyltrimethyl ammonium chloride chitosan. The method was efficient to obtain nanoparticles with low polydispersity index and small hydrodynamic diameter. And high zeta potential value indicated that nanoparticles had good stability. The adriamycin release of nanoparticles represented a significant response to pH, with the fastest release in phosphate buffer solution at pH 6.8. Meanwhile, the antioxidant efficiency of nanoparticles was assayed, and nanoparticles represented significant enhancement in radicals scavenging activity. The assay of cell viability by CCK-8 test exhibited that nanoparticles led to statistically significant decrease in cell viability for four kinds of cancer cells (HEPG-2, A549, MCF-7, and BGC-823). It was indicated that the nanoparticles with enhanced biological activity, reduced cytotoxicity, and pH-sensitive release could be served as potential drug carrier in drug delivery system.

Novel 2-Hydroxypropyltrimethyl Ammonium Chitosan Derivatives: Synthesis, Characterization, Moisture Absorption and Retention Properties.

Recent years have seen a steady increase in interest and demand for the use of humectants based on biodegradable natural polymers in many fields. The aim of this paper is to investigate the moisture absorption and retention properties of 2-hydroxypropyltrimethyl ammonium chitosan derivatives which were modified by anionic compounds via ion exchange. FTIR, 1H NMR, and 13C NMR spectroscopy were used to demonstrate the specific structures of chitosan derivatives. The degrees of substitution for objective products were calculated by the integral ratio of hydrogen atoms according to 1H NMR spectroscopy. Meanwhile, moisture absorption of specimens was assayed in a desiccator at different relative humidity (RH: 43% and 81%), and all target products exhibited enhanced moisture absorption. Furthermore, moisture retention measurement at different relative humidity (RH: 43%, 81%, and drier silica gel) was estimated, and all target products possessed obviously improved moisture retention property. Specifically, after 48 h later, the moisture retention property of HACBA at 81% RH was 372.34%, which was much higher than HA (180.04%). The present study provided a novel method to synthesize chitosan derivatives with significantly improved moisture absorption and retention properties that would serve as potential humectants in biomedical, food, medicine, and cosmetics fields.

Determination of chitosan content with Schiff base method and HPLC.

Tremendous awareness of determination of chitosan content accurately is increasing, due to it has great significance to the quality control of chitosan. In this article, two kinds of chitosan-Schiff base derivatives (BCSB and PCSB) were synthesized by the different average degrees of deacetylation (DD) of chitosan with benzaldehyde or propanal, respectively. The total mass of Schiff base derivative product was dried and obtained without washing and loss. Then, a certain amount of the prepared Schiff base compound was taken to hydrolyze into glucosamine hydrochloride (GAH) in strong hydrochloric acidic environment, whose concentration was quantified by HPLC, and the mass of GAH contained in hydrolysis solution could be calculated. Subsequently, the total quality of GAH obtained by hydrolysis of all of the Schiff base product was calculated and obtained, and then the theoretical mass of chitosan could be deduced and calculated by further converse calculation. Finally, the chitosan content was obtained by combining the sample mass used in Schiff base reaction and the theoretical mass of chitosan. This method was accurate and convenient, providing a preeminent idea and method for the determination of chitosan content.

Modification of carboxymethyl inulin with heterocyclic compounds: Synthesis, characterization, antioxidant and antifungal activities.

A series of novel inulin derivatives were designed and synthesized by the introduction of amino heterocyclic moieties onto carboxymethyl inulin with the aid of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-hydroxysuccinimide. The target products were prepared via three - step chemical synthesis, and structures were identified by FTIR and 1H NMR spectroscopy. Antioxidant activities of inulin derivatives including DPPH - radical scavenging assay, superoxide - radical scavenging assay, hydroxyl - radical scavenging assay, and reducing power were estimated. Meanwhile, their antifungal activities, including Colletotrichum lagenarium and Botrytis cinerea, were also explored by hyphal measurement. In particular, inulin derivatives bearing heterocyclic moieties exhibited a remarkable improvement over inulin on antioxidant and antifungal activities, and their bioactivities decreased roughly in the order of 2ATCMI > 4APCMI > 3APCMI > 2APCMI > 3ATCMI > CMI > inulin. Furthermore, the cytotoxicities of inulin derivatives against L929 cells were evaluated by CCK-8 in vitro, and all samples showed weak cytotoxicities. In a nutshell, the paper provides a practical approach to synthesize novel inulin derivatives with dramatically enhanced bioactivity and good biocompatibility. The product described in paper might serve as a new leading structure for further design of antioxidants or antifungal agents in biomedicine, cosmetics, and other fields.

Nuclear factor Y participates in alcoholic liver disease by activating SREBP1 expression in mice.

Chronic and excessive alcohol consumption leads to alcoholic liver disease (ALD). However, the molecular mechanisms in the regulation of ALD have not been fully deciphered. Liver lipid accumulation is an important research direction in ALD. In this study, the physiological role of nuclear factor Y (NF-Y) in ALD and the related mechanisms were investigated using murine hepatocytes and an ethanol-induced liver injury mouse model. In this study, ethanol promoted hepatic NF-Y expression in a mouse model and Hepa1-6 mouse hepatocytes. Lentivirus-mediated NF-Y overexpression in Hepa1-6 cells markedly increased sterol regulatory element binding protein 1 (SREBP1) and fatty acid synthase (FASN) expression compared with empty vector control cells. Conversely, CRISPR/Cas9-mediated knockdown of NF-Y subunit A (NF-YA) attenuated FASN and SREBP1 expression. Mechanistically, luciferase reporter gene assays and chromatin immunoprecipitation (ChIP) analysis indicated that NF-Y activates the transcription of SREBP1 by directly binding to the CCAAT regulatory sequence motif in the promoter. Overall, our results reveal a previously unrecognized physiological function of NF-Y in ALD by activating sterol regulatory element-binding protein 1 (SREBP1). Modulation of hepatic NF-Y expression may therefore offer an attractive therapeutic approach to manage ALD.

Efficacy and safety of human chorionic gonadotropin combined with human menopausal gonadotropin and a gonadotropin-releasing hormone pump for male adolescents with congenital hypogonadotropic hypogonadism / 中华医学杂志(英文版)

BACKGROUND@#Compared to adult studies, studies which involve the treatment of pediatric congenital hypogonadotropic hypogonadism (CHH) are limited and no universal treatment regimen is available. The aim of this study was to evaluate the feasibility of human chorionic gonadotropin (hCG)/human menopausal gonadotropin (hMG) therapy for treating male adolescents with CHH.@*METHODS@#Male adolescent CHH patients were treated with hCG/hMG (n = 20) or a gonadotropin-releasing hormone (GnRH) pump (n = 21). The treatment was divided into a study phase (0-3 months) and a follow-up phase (3-12 months). The testicular volume (TV), penile length (PL), penis diameter (PD), and sex hormone levels were compared between the two groups. The TV and other indicators between the groups were analyzed using a t-test (equal variance) or a rank sum test (unequal variance).@*RESULTS@#Before treatment, there was no statistical difference between the two groups in terms of the biochemistry, hormones, and other demographic indicators. After 3 months of treatment, the TV of the hCG/hMG and GnRH groups increased to 5.1 ± 2.3 mL and 4.1 ± 1.8 mL, respectively; however, the difference was not statistically significant (P > 0.05, t = 1.394). The PL reached 6.9 ± 1.8 cm and 5.1 ± 1.6 cm (P  0.05, t = 0.314). After 9 to 12 months of treatment, the T level was higher in the hCG/hMG group. Other parameters did not exhibit a statistical difference.@*CONCLUSIONS@#The hCG/hMG regimen is feasible and effective for treating male adolescents with CHH. The initial 3 months of treatment may be a window to optimally observe the strongest effects of therapy. Furthermore, results from the extended time-period showed positive outcomes at the 1-year mark; however, the long-term effectiveness, strengths, and weaknesses of the hCG/hMG regimen require further research.@*TRIAL REGISTRATION@#ClinicalTrials.gov, NCT02880280; https://clinicaltrials.gov/ct2/show/NCT02880280.

Construction of Rho GDIa-sgRNAs plasmids by clustered regularly interspaced short palindromic repeats/associated protein 9 and the effect on migration of Hepa 1-6 cell line in mouse / 解剖学报

Objective To construct the recombinant plasmids of knocking down Rho guanine dissociation inhibitor α (GDIα ) gene by using clustered regularly interspaced short palindromic repeats/associated protein 9 (CRISPR/Cas9) technique, and investigate the effect of Rho GDIα interference on the migration of Hepa 1-6 cells of mouse in order to provide the method of prevention and treatment of liver cancer. Methods To construct and identify the PX458-Rho GDIα-single guide (sg) RNAs by using CRISPR/Cas9 technique. And the Hepa 1-6 cells were transfected by liposomes with PX458-Rho GDIα-sgRNAs for 48 hours respectively, and cells treated with PX458 plasmids were used as control. The migration ability of Hepa 1-6 was checked by wound healing assay and Transwell assay, respectively. Results The expression of Rho GDIα was depressed in group of PX458-Rho GDIα-sgRNAl transfection which was detected by using RT-PCR. The migration distance of Hepa 1-6 in PX458-Rho GDIα-sgRNAl transfection group was significantly promoted comparing with the control group which was transfected with PX458 only, and the cell number of PX458-Rho GDIα-sgRNAl group was more than that in control group by using transwell assay, indicating concluded that knocking down of Rho GDIα promoted the migration ability of Hepal-6 cells. Conclusion The result is explicit that in vivo, Rho GDIα may inhibit the migration of Hepal-6 partially. Overexpression of Rho GDIα might be used as an important method to prevent the metastasize of carcinoma.