ObjectiveTo explore the impact of exogenous chitosan on the growth and metabolism of Glycyrrhiza uralensis Fisch. (G. uralensis) and to improve the quality of cultivated G. uralensis for both medicine and food and aid in the increase in the content of effective components in G. uralensis.MethodsIn this study, whole G. uralensis plants were treated with exogenous chitosan, and comprehensive analyses of secondary metabolites and proteins were conducted using liquid chromatography with tandem mass spectrometry and isobaric tag for relative and absolute quantitation, respectively. Effects of chitosan induction on endogenous hormones of G. uralensis were analyzed using an enzyme-linked immunosorbent assay. Gene ontology function annotation and Kyoto Encyclopedia of Genes and Genomes pathway annotation were conducted to study the effect of chitosan induction on the proteome.ResultsChitosan induction significantly increased the levels of flavonoids in G. uralensis; however, the variation in triterpenoids was not substantial. Biological processes, including photosynthesis, secondary metabolism, and abiotic stress responses, were significantly enriched. Additionally, the photosynthetic pathway, photosynthesis-antenna protein pathway, and plant hormone signal transduction pathway were significantly enriched. In the flavonoid biosynthesis pathway, the upstream-related enzyme phenylalanine ammonia-lyase (PAL) and the downstream-related enzymes chalcone synthase (CHS), polyketide reductase (PKR), chalcone isomerase (CHI), and vestitone reductase (VR) were significantly upregulated.ConclusionsOur findings suggest that chitosan induction may promote the tricarboxylic acid (TCA) cycle, and the TCA cycle enhancement significantly upregulated PAL, CHS, PKR, CHI, and VR, the five key enzymes involved in flavonoid synthesis of G. uralensis, indicating that chitosan induction activated the entire metabolic pathway associated with flavonoids in G. uralensis. Our findings provide a reference for improving the quality of cultivated G. uralensis from the perspective of pharmacodynamic components.

In modern society, people are increasingly exposed to chronic stress, leading to various mental disorders. However, the activities of brain regions, especially neural firing patterns related to specific behaviors, remain unclear. In this study, we introduce a novel approach, NeuroSync, which integrates open-field behavioral testing with electrophysiological recordings from emotion-related brain regions, specifically the central amygdala and the paraventricular nucleus of the hypothalamus, to explore the mechanisms of negative emotions induced by chronic stress in mice. By applying machine vision techniques, we quantified behaviors in the open field, and signal processing algorithms elucidated the neural underpinnings of the observed behaviors. Synchronizing behavioral and electrophysiological data revealed significant correlations between neural firing patterns and stress-related behaviors, providing insights into real-time brain activity underlying behavioral responses. This research combines deep learning and machine learning to synchronize high-resolution video and electrophysiological data, offering new insights into neural-behavioral dynamics under chronic stress conditions.
Animals ; Mice ; Male ; Emotions/physiology* ; Stress, Psychological/physiopathology* ; Action Potentials/physiology* ; Mice, Inbred C57BL ; Behavior, Animal/physiology* ; Machine Learning ; Amygdala/physiopathology* ; Neurons/physiology* ; Paraventricular Hypothalamic Nucleus/physiopathology* ; Brain/physiology*

AIM:To investigate the expression profile and biological significance of long noncoding RNA(lncRNA)zinc finger antisense 1(ZFAS1)in the brains of mice with diabetic encephalopathy(DE).METHODS:Ten db/m mice served as the normal control group,while twenty 22-week-old db/db mice were used to establish the DE model and randomly divided into two subgroups:ten as the db/db model control and the remaining ten receiving ZFAS1 gene knockdown(db/db+sh-ZFAS1)via lentiviral transfection.Weekly measurements of body weight and blood glucose levels were performed.Brain tissues were collected for Nissl staining to evaluate neuronal damage,TUNEL assay to detect apop-tosis,and immunofluorescence staining to examine neural biomarker expression.Serum levels of tumor necrosis factor-α(TNF-α)and oxidative stress markers,including reactive oxygen species(ROS),malondialdehyde(MDA),superoxide dismutase(SOD),catalase(CAT)and glutathione peroxidase(GSH-Px),were determined.Western blot was conducted to quantify the protein expression levels of B-cell lymphoma-2(Bcl-2),Bcl-2-associated X protein(Bax),p38 mitogen-activated protein kinase and phosphorylated p38(p-p38)in brain tissues.The expression levels of ZFAS1 and caspase-3 mRNA were determined by RT-qPCR.RESULTS:Knockdown of ZFAS1 in db/db mice significantly improved cognitive function,alleviated hippocampal neuronal damage,and reduced body weight and blood glucose levels(P<0.01).More-over,oxidative stress was mitigated,as evidenced by decreased MDA and ROS levels(P<0.01)and increased activity of antioxidant enzymes,GSH-Px,SOD and CAT(P<0.01 or P<0.05).Meanwhile,ZFAS1 silencing down-regulated Bax and p-p38/p38 protein expression(P<0.01 or P<0.05)while up-regulating Bcl-2(P<0.01).Consistently,RT-qPCR confirmed significant down-regulation of ZFAS1 and caspase-3 mRNA levels(P<0.01).CONCLUSION:lncRNA ZFAS1 is highly expressed in the hippocampus of DE mice.Down-regulation of ZFAS1 expression enhances cognitive func-tion,suppresses oxidative stress,and inhibits neuronal apoptosis,thereby attenuating neural damage in DE.

OBJECTIVE To study and analyse the main components of Tongluo Muzu Powder transdermal absorption solution,as well as to investigate its mechanism of action in the treatment of osteoarthritis of the knee.METHODS The transdermal absorption solution of Tongluo Muzu Powder was extracted by Franz vertical diffusion cell,and its components were analyzed by UHPLC-MS/MS.Then TCMSP,Swiss target prediction,Gene Cards and other databases were introduced to predict the possible targets of the effec-tive components of transdermal absorption solution for the prevention and treatment of knee osteoarthritis,and Cytoscape software was used to construct the"drug-component-disease-target"visual network,and then the protein interaction network was obtained through the string database to find the core targets.Finally,AutoDock and PyMOL were used to verify the molecular docking of main active in-gredients and targets,and the gene ontology(GO)function analysis of core genes and the enrichment analysis of Kyoto Encyclopedia of genes and genomes(KEGG)pathway were conducted.Primary mouse chondrocytes were extracted,and the predicted targets of net-work pharmacology were verified by ELISA,Western blot,qPCR,etc.RESULTS A total of 48 effective components of the transder-mal absorption solution of Tongluo Muzu Powder and 88 possible targets for the treatment of knee osteoarthritis were screened out.The results of enrichment analysis showed that it was mainly involved in the process of apoptosis and oxidative stress.Molecular docking suggested that the main active ingredients and targets had good binding ability.The results showed that the lyophilized powder of the transdermal absorption solution of Tongluo Muzu Powder could reduce the concentration of inflammatory factors in the supernatant of chondrocytes after LPS intervention in a dose-dependent manner(P<0.05,P<0.01).The protein expression of p-PI3K/PI3K,p-AKT/AKT,MMP13 and p53 was decreased,and the protein expression of collagen Ⅱ was increased(P<0.05,P<0.01).At the same time,the mRNA expression of MMP13 and p53 was decreased and the mRNA expression of collagen Ⅱ was increased(P<0.05,P<0.01).In addition,it reduced the fluorescence intensity of TUNEL staining in cells(P<0.01).CONCLUSION By combining UH-PLC-MS/MS technology with network pharmacology,the main active ingredients of Tongluo Muzu Powder transdermal absorption solu-tion are preliminarily understood,and its potential mechanism of action in the treatment of knee osteoarthritis are predicted.

BACKGROUND:Enhancing the differentiation of induced pluripotent stem cells into lung stem cells is crucial for repairing lung injuries.NKX2.1 is the earliest marker of lung epithelial differentiation and plays a significant regulatory role in lung development.However,the impact of its expression on the differentiation of induced pluripotent stem cells into lung stem cells remains inadequately understood.OBJECTIVE:To investigate the effect of NKX2.1 on the differentiation of induced pluripotent stem cells into lung stem cells.METHODS:Induced pluripotent stem cells were cultured in vitro.The expression of specific pluripotent stem cell genes was assessed using real-time fluorescence quantitative PCR.NKX2.1 was overexpressed in induced pluripotent stem cells,which were then induced to differentiate into lung stem cells.The expression of FoxA2,SOX9,and P63 was determined via quantitative PCR and immunofluorescence on day 7 of induction of differentiation.The expression of the alveolar marker SPB and SPC was evaluated through immunofluorescence staining on day 7 of induction of differentiation.RESULTS AND CONCLUSION:(1)Induced pluripotent stem cells in vitro were tightly packed and showed typical clonoid growth and significantly expressed stem cell-specific genes OCT-4,SOX2,and NANOG.(2)Compared with the non-transfected control group,the expression of NKX2.1 in human induced pluripotent stem cells was significantly increased in the NKX2.1 overexpression group(P＜0.000 1).(3)Seven days after induction of differentiation,compared with the non-transfected control group,the expression of lung stem cell-related markers FoxA2,SOX9,and P63 was significantly increased in the NKX2.1 overexpression group(P＜0.000 1).(4)Thirteen days after induction of differentiation,compared with the non-transfected control group,the fluorescence intensity of alveolar cell marker molecules SPB and SPC increased significantly in the overexpression NKX2.1 group.The results show that NKX2.1 can promote the differentiation of induced pluripotent stem cells into lung stem cells.

Transmissible gastroenteritis(TGE)is a highly contagious gastrointestinal disease of pigs caused by the transmissible gastroenteritis virus(TGEV).It results in severe diarrhea and high mortality in suckling piglets.As no specific treatment available,vaccination is considered to be one of the most cost-effective measures for preventing and controlling TGE.However,the immune protection provided by vaccines based on traditional TGEV strains is becoming inadequate due to the continuous emergence of strong and new strains of the virus,which poses a serious threat to the pig industry's health and development.Therefore,the development of new vaccines with im-proved efficiency and broad-spectrum coverage is urgently needed.This paper aims to summarize the pathogenic structural characteristics of TGEV,discuss the latest advancements in TGEV vac-cine research and development,and propose future strategies for the development of highly effec-tive TGEV vaccines.The findings of this paper can serve as a valuable reference for effectively pre-venting and controlling TGE in clinical practice.

OBJECTIVE To study and analyse the main components of Tongluo Muzu Powder transdermal absorption solution,as well as to investigate its mechanism of action in the treatment of osteoarthritis of the knee.METHODS The transdermal absorption solution of Tongluo Muzu Powder was extracted by Franz vertical diffusion cell,and its components were analyzed by UHPLC-MS/MS.Then TCMSP,Swiss target prediction,Gene Cards and other databases were introduced to predict the possible targets of the effec-tive components of transdermal absorption solution for the prevention and treatment of knee osteoarthritis,and Cytoscape software was used to construct the"drug-component-disease-target"visual network,and then the protein interaction network was obtained through the string database to find the core targets.Finally,AutoDock and PyMOL were used to verify the molecular docking of main active in-gredients and targets,and the gene ontology(GO)function analysis of core genes and the enrichment analysis of Kyoto Encyclopedia of genes and genomes(KEGG)pathway were conducted.Primary mouse chondrocytes were extracted,and the predicted targets of net-work pharmacology were verified by ELISA,Western blot,qPCR,etc.RESULTS A total of 48 effective components of the transder-mal absorption solution of Tongluo Muzu Powder and 88 possible targets for the treatment of knee osteoarthritis were screened out.The results of enrichment analysis showed that it was mainly involved in the process of apoptosis and oxidative stress.Molecular docking suggested that the main active ingredients and targets had good binding ability.The results showed that the lyophilized powder of the transdermal absorption solution of Tongluo Muzu Powder could reduce the concentration of inflammatory factors in the supernatant of chondrocytes after LPS intervention in a dose-dependent manner(P<0.05,P<0.01).The protein expression of p-PI3K/PI3K,p-AKT/AKT,MMP13 and p53 was decreased,and the protein expression of collagen Ⅱ was increased(P<0.05,P<0.01).At the same time,the mRNA expression of MMP13 and p53 was decreased and the mRNA expression of collagen Ⅱ was increased(P<0.05,P<0.01).In addition,it reduced the fluorescence intensity of TUNEL staining in cells(P<0.01).CONCLUSION By combining UH-PLC-MS/MS technology with network pharmacology,the main active ingredients of Tongluo Muzu Powder transdermal absorption solu-tion are preliminarily understood,and its potential mechanism of action in the treatment of knee osteoarthritis are predicted.

BACKGROUND:Enhancing the differentiation of induced pluripotent stem cells into lung stem cells is crucial for repairing lung injuries.NKX2.1 is the earliest marker of lung epithelial differentiation and plays a significant regulatory role in lung development.However,the impact of its expression on the differentiation of induced pluripotent stem cells into lung stem cells remains inadequately understood.OBJECTIVE:To investigate the effect of NKX2.1 on the differentiation of induced pluripotent stem cells into lung stem cells.METHODS:Induced pluripotent stem cells were cultured in vitro.The expression of specific pluripotent stem cell genes was assessed using real-time fluorescence quantitative PCR.NKX2.1 was overexpressed in induced pluripotent stem cells,which were then induced to differentiate into lung stem cells.The expression of FoxA2,SOX9,and P63 was determined via quantitative PCR and immunofluorescence on day 7 of induction of differentiation.The expression of the alveolar marker SPB and SPC was evaluated through immunofluorescence staining on day 7 of induction of differentiation.RESULTS AND CONCLUSION:(1)Induced pluripotent stem cells in vitro were tightly packed and showed typical clonoid growth and significantly expressed stem cell-specific genes OCT-4,SOX2,and NANOG.(2)Compared with the non-transfected control group,the expression of NKX2.1 in human induced pluripotent stem cells was significantly increased in the NKX2.1 overexpression group(P＜0.000 1).(3)Seven days after induction of differentiation,compared with the non-transfected control group,the expression of lung stem cell-related markers FoxA2,SOX9,and P63 was significantly increased in the NKX2.1 overexpression group(P＜0.000 1).(4)Thirteen days after induction of differentiation,compared with the non-transfected control group,the fluorescence intensity of alveolar cell marker molecules SPB and SPC increased significantly in the overexpression NKX2.1 group.The results show that NKX2.1 can promote the differentiation of induced pluripotent stem cells into lung stem cells.

Transmissible gastroenteritis(TGE)is a highly contagious gastrointestinal disease of pigs caused by the transmissible gastroenteritis virus(TGEV).It results in severe diarrhea and high mortality in suckling piglets.As no specific treatment available,vaccination is considered to be one of the most cost-effective measures for preventing and controlling TGE.However,the immune protection provided by vaccines based on traditional TGEV strains is becoming inadequate due to the continuous emergence of strong and new strains of the virus,which poses a serious threat to the pig industry's health and development.Therefore,the development of new vaccines with im-proved efficiency and broad-spectrum coverage is urgently needed.This paper aims to summarize the pathogenic structural characteristics of TGEV,discuss the latest advancements in TGEV vac-cine research and development,and propose future strategies for the development of highly effec-tive TGEV vaccines.The findings of this paper can serve as a valuable reference for effectively pre-venting and controlling TGE in clinical practice.