OBJECTIVE To investigate the effects and mechanisms of Lycium ruthenicum Murr. in improving sleep. METHODS Network pharmacology was employed to identify the active components of L. ruthenicum and their associated disease targets， followed by enrichment analysis. A caffeine‑induced zebrafish model of sleep deprivation was established ， and the zebrafish were treated with L. ruthenicum Murr. extract （LRME） at concentrations of 0.1， 0.2 and 0.4 mg/mL， respectively； 24 h later， behavioral changes of zebrafish and pathological alterations in brain neurons were subsequently observed. The levels of inflammatory factors [interleukin-6 （IL-6）， IL-1β， IL-10， tumor necrosis factor-α （TNF-α）]， oxidative stress markers [superoxide dismutase （SOD）， malondialdehyde （MDA）， glutathione peroxidase （GSH-Px）， catalase （CAT）]， and neurotransmitters [5- hydroxytryptamine （5-HT）， γ-aminobutyric acid （GABA）， glutamic acid （Glu）， dopamine （DA）， and norepinephrine （NE）] were measured. The protein expression levels of protein kinase B1 （AKT1）， phosphorylated AKT1 （p-AKT1）， epidermal growth factor receptor （EGFR）， B-cell lymphoma 2 （Bcl-2）， sarcoma proto-oncogene，non-receptor tyrosine kinase （SRC）， and heat shock protein 90α family class A member 1 （HSP90AA1） in the zebrafish were also determined. RESULTS A total of 12 active components and 176 intersecting disease targets were identified through network pharmacology analysis. Among these， apigenin， naringenin and others were recognized as core active compounds， while AKT1， EGFR and others served as key targets； EGFR tyrosine kinase inhibitor resistance signaling pathway was identified as the critical pathway. The sleep improvement rates in zebrafish of LRME low-， medium-， and high-dose groups were 54.60%， 69.03% and 77.97%， 开发。E-mail：hjp_yft@163.com respectively， while the inhibition ratios of locomotor distance were 0.57， 0.83 and 0.95， respectively. Compared with the model group， the number of resting counts， resting time and resting distance were significantly increased/extended in LRME medium- and high-dose groups （P＜0.05）. Neuronal damage in the brain was alleviated. Additionally， the levels of IL-6， IL-1β， TNF-α， MDA， Glu， DA and NE， as well as the protein expression levels of AKT1， p-AKT1， EGFR， SRC and HSP90AA1， were markedly reduced （P＜0.05）， while the levels of IL-10， SOD， GSH-Px， CAT， 5-HT and GABA， as well as Bcl-2 protein expression， were significantly elevated （P＜0.05）. CONCLUSIONS L. ruthenicum Murr. demonstrates sleep-improving effects， and its specific mechanism may be related to the regulation of inflammatory responses， oxidative stress， neurotransmitter balance， and the EGFR tyrosine kinase inhibitor resistance signaling pathway.

BACKGROUND:Exercise is an effective way to improve microvascular function in patients with type 2 diabetes.In recent years,exercise has been used as an intervention therapy for microvascular dysfunction in patients with type 2 diabetes.However,few studies have systematically explored the influence of factors,such as"exercise type,exercise intensity and amount of exercise,"on microvascular function in patients.To some extent,this limits the formulation of precise exercise prescriptions to improve microvascular dysfunction in patients with type 2 diabetes and the comparison of study results.OBJECTIVE:To investigate the effects of exercise type,intensity,frequency and amount of exercise on microvascular function in patients with type 2 diabetes mellitus,and to make suggestions on exercise prescription.METHODS:The first author used computer to search the studies on the improvement of microvascular function in type 2 diabetes patients involving exercise in CNKI,WanFang,PubMed and other databases.The search terms were"diabetes mellitus,type 2 diabetes mellitus,microcirculation,microvascular reactivity,microvessels,capillaries,vasodilation,blood perfusion volume,endothelial cells,shear stress,exercise,aerobic exercise,resistance exercise,high-intensity exercise"in Chinese and English.The articles were screened by a quick glance at the article titles and abstracts to exclude those that were not closely related to the topic,and finally 60 articles were included for review.RESULTS AND CONCLUSION:(1)Exercise is an effective way to improve microvascular function in patients with type 2 diabetes.Aerobic exercise lasting 12-24 weeks,3-5 times/week,exercise time＞30 minutes and intensity between 40％and 59％reserve oxygen intake can significantly improve microvascular function in patients with type 2 diabetes.On the basis of aerobic exercise,systemic resistance exercise 2-3 times a week(50％-85％1RM,every other day)or pressure resistance exercise can obtain better intervention effects.(2)In addition,exercise can improve microvascular function in patients with type 2 diabetes in a"dose-effect"manner,and patients can get better results from the intervention by increasing the amount of exercise,while maintaining safety.(3)The mechanism of exercise improving microvascular function in patients with type 2 diabetes is mainly related to promoting the release of nitric oxide and vascular endothelial growth factor from endothelial cells and inhibiting the release of endothelin1.

BACKGROUND:Exercise is an effective way to improve microvascular function in patients with type 2 diabetes.In recent years,exercise has been used as an intervention therapy for microvascular dysfunction in patients with type 2 diabetes.However,few studies have systematically explored the influence of factors,such as"exercise type,exercise intensity and amount of exercise,"on microvascular function in patients.To some extent,this limits the formulation of precise exercise prescriptions to improve microvascular dysfunction in patients with type 2 diabetes and the comparison of study results.OBJECTIVE:To investigate the effects of exercise type,intensity,frequency and amount of exercise on microvascular function in patients with type 2 diabetes mellitus,and to make suggestions on exercise prescription.METHODS:The first author used computer to search the studies on the improvement of microvascular function in type 2 diabetes patients involving exercise in CNKI,WanFang,PubMed and other databases.The search terms were"diabetes mellitus,type 2 diabetes mellitus,microcirculation,microvascular reactivity,microvessels,capillaries,vasodilation,blood perfusion volume,endothelial cells,shear stress,exercise,aerobic exercise,resistance exercise,high-intensity exercise"in Chinese and English.The articles were screened by a quick glance at the article titles and abstracts to exclude those that were not closely related to the topic,and finally 60 articles were included for review.RESULTS AND CONCLUSION:(1)Exercise is an effective way to improve microvascular function in patients with type 2 diabetes.Aerobic exercise lasting 12-24 weeks,3-5 times/week,exercise time＞30 minutes and intensity between 40％and 59％reserve oxygen intake can significantly improve microvascular function in patients with type 2 diabetes.On the basis of aerobic exercise,systemic resistance exercise 2-3 times a week(50％-85％1RM,every other day)or pressure resistance exercise can obtain better intervention effects.(2)In addition,exercise can improve microvascular function in patients with type 2 diabetes in a"dose-effect"manner,and patients can get better results from the intervention by increasing the amount of exercise,while maintaining safety.(3)The mechanism of exercise improving microvascular function in patients with type 2 diabetes is mainly related to promoting the release of nitric oxide and vascular endothelial growth factor from endothelial cells and inhibiting the release of endothelin1.

ObjectiveSpinal cord injury (SCI) directly impairs the regulatory function of the autonomic nervous system, induces intestinal dysfunction, and significantly reduces patients’ quality of life. Preclinical studies have shown that electroacupuncture (EA) therapy can regulate the brain-gut axis and is used to treat central nervous system diseases such as major depressive disorder, Alzheimer’s disease and Parkinson’s disease. Recent research has established that fecal microbiota transplantation (FMT) from EA-treated SCI rats restored intestinal motility and colonic morphology. However, it remains unclear whether the regulation of gut microbiota by EA therapy directly contributes to neural repair after SCI. This study aims to explore whether gut microbiota mediates the neuroprotective effect of EA in the treatment of SCI and its possible mechanism. MethodsThe study employed RNA transcriptome analysis of spinal cord tissue to characterize gene expression profiles and to identify key signaling pathways following EA treatment for SCI. Hematoxylin-Eosin (HE) staining and Nissl staining were used to observe the morphological changes in spinal cord tissue. Western blot (WB) and enzyme-linked immunosorbent assay (ELISA) were applied to detect the effects of EA on the expression of proteins related to nucleotide-binding domain leucine-rich repeat and pyrin domain-containing receptor 3 (NLRP3) -dependent pyroptosis. Using 16S rDNA sequencing, the study observed alterations in gut microbiota diversity and community composition in SCI rats. Prior to establishing SCI models, rats were pretreated with an antibiotic cocktail to induce gut dysbiosis, and the effects on intestinal function and spinal cord neural repair were evaluated. FMT was performed to investigate the regulatory effects of post-EA FMT on motor function, general status, liver and spleen indices, and NLRP3-mediated pyroptosis in SCI rats. ResultsEA improved motor function and reduced regulated neuronal cell death in SCI rats. Transcriptomic analysis demonstrated the activation of immune- and inflammation-related pathways post-SCI, including NOD-like receptors, nuclear factor-kappa B(NF-κB), and Toll-like receptor (TLR) pathways. EA primarily influenced intestinal inflammation and autoimmune functions. 16S rDNA sequencing illustrated that EA did not alter the diversity of gut microbiota. However, EA altered the gut microbiota composition in SCI rats, increasing Lactobacillus and Akkermansia genera while rebalancing the Firmicutes/Bacteroidetes ratio. Furthermore, depletion of gut microbiota by antibiotics disrupted the intestinal barrier, reduced the expression of intestinal barrier proteins Zonula Occludens-1 (ZO-1) and Occludin, elevated serum lipopolysaccharide-binding protein (LBP) levels, exacerbated spinal cord tissue damage, and hindered motor function recovery in SCI rats. FMT from donors treated with EA reduced LBP levels in the intestine, blood, and spinal cord of rats, inhibited the TLR4 myeloid differentiation primary response protein 88 (MyD88)-NF‑κB pathway and NLRP3-dependent pyroptosis, and improved motor function. On the other hand, FMT treatment resulted in decreased body weight and food intake, whereas FMT using EA-treated donors effectively alleviated these alterations. ConclusionEA effectively alleviated neuroinflammatory responses in rats with SCI, primarily through regulating the gut microbiota and suppressing the NLRP3-dependent pyroptosis signaling pathway.

ObjectiveSpinal cord injury (SCI) directly impairs the regulatory function of the autonomic nervous system, induces intestinal dysfunction, and significantly reduces patients’ quality of life. Preclinical studies have shown that electroacupuncture (EA) therapy can regulate the brain-gut axis and is used to treat central nervous system diseases such as major depressive disorder, Alzheimer’s disease and Parkinson’s disease. Recent research has established that fecal microbiota transplantation (FMT) from EA-treated SCI rats restored intestinal motility and colonic morphology. However, it remains unclear whether the regulation of gut microbiota by EA therapy directly contributes to neural repair after SCI. This study aims to explore whether gut microbiota mediates the neuroprotective effect of EA in the treatment of SCI and its possible mechanism. MethodsThe study employed RNA transcriptome analysis of spinal cord tissue to characterize gene expression profiles and to identify key signaling pathways following EA treatment for SCI. Hematoxylin-Eosin (HE) staining and Nissl staining were used to observe the morphological changes in spinal cord tissue. Western blot (WB) and enzyme-linked immunosorbent assay (ELISA) were applied to detect the effects of EA on the expression of proteins related to nucleotide-binding domain leucine-rich repeat and pyrin domain-containing receptor 3 (NLRP3) -dependent pyroptosis. Using 16S rDNA sequencing, the study observed alterations in gut microbiota diversity and community composition in SCI rats. Prior to establishing SCI models, rats were pretreated with an antibiotic cocktail to induce gut dysbiosis, and the effects on intestinal function and spinal cord neural repair were evaluated. FMT was performed to investigate the regulatory effects of post-EA FMT on motor function, general status, liver and spleen indices, and NLRP3-mediated pyroptosis in SCI rats. ResultsEA improved motor function and reduced regulated neuronal cell death in SCI rats. Transcriptomic analysis demonstrated the activation of immune- and inflammation-related pathways post-SCI, including NOD-like receptors, nuclear factor-kappa B(NF-κB), and Toll-like receptor (TLR) pathways. EA primarily influenced intestinal inflammation and autoimmune functions. 16S rDNA sequencing illustrated that EA did not alter the diversity of gut microbiota. However, EA altered the gut microbiota composition in SCI rats, increasing Lactobacillus and Akkermansia genera while rebalancing the Firmicutes/Bacteroidetes ratio. Furthermore, depletion of gut microbiota by antibiotics disrupted the intestinal barrier, reduced the expression of intestinal barrier proteins Zonula Occludens-1 (ZO-1) and Occludin, elevated serum lipopolysaccharide-binding protein (LBP) levels, exacerbated spinal cord tissue damage, and hindered motor function recovery in SCI rats. FMT from donors treated with EA reduced LBP levels in the intestine, blood, and spinal cord of rats, inhibited the TLR4 myeloid differentiation primary response protein 88 (MyD88)-NF‑κB pathway and NLRP3-dependent pyroptosis, and improved motor function. On the other hand, FMT treatment resulted in decreased body weight and food intake, whereas FMT using EA-treated donors effectively alleviated these alterations. ConclusionEA effectively alleviated neuroinflammatory responses in rats with SCI, primarily through regulating the gut microbiota and suppressing the NLRP3-dependent pyroptosis signaling pathway.

Objective To analyze the PLCβ4 gene mRNA expression and its clinical significance in hepatocellular carcinoma (HCC) based on TCGA database. Methods Based on the data on 424 clinical samples (including 374 cases of HCC tissues and 50 cases of nontumor liver tissues) in the TCGA database, Kaplan–Meier method, Cox regression analysis, and immune infiltration analysis were performed to evaluate the relationship between PLCβ4 gene and the clinical characteristics and survival prognosis of HCC patients. Correlation analysis between PLCβ4 gene and 24 types of immune cells was applied to investigate the relationship between PLCβ4 gene and immune cell infiltration and mRNA expression level of TP53 gene, a high-frequency mutation gene in HCC. In addition, paraffin sections of highly, moderately, and poorly differentiated tumor tissues and normal liver tissues from HCC patients were collected. The histopathological observation was carried out via HE staining method, and the expression levels of PLCβ4 and Ki-67 proteins in each clinical sample were verified through the immunohistochemical method. Results The expression level of PLCβ4 gene in HCC was significantly higher than that in normal tissues (P<0.01), and all patients in the PLCβ4 high-expression group had a significantly longer overall survival than those in the low-expression group (P<0.05), which suggested that PLCβ4 substantially affected the prognosis of HCC patients. Correlation analysis showed that the expression level of PLCβ4 gene was highly correlated with immune cell infiltration and the expression level of TP53 gene. As verified by clinical sample experiments, HE staining experiments and immunohistochemical results revealed that PLCβ4 gene expression in HCC tissue samples was significantly higher than that in normal tissues (P<0.001), and it was negatively correlated with the degree of differentiation. Conclusion PLCβ4 may serve as an independent prognostic factor in HCC and is expected to be a novel molecular target for HCC treatment.

In this study, RAW264.7 cells were employed to investigate the effects of honey-processed Astragalus on their energy metabolism and polarization, and explore the scientific connotation of the enhanced efficacy of honey-processed Astragalus on invigorating spleen-stomach and replenishing Qi. The medicated sera were prepared by intragastric administration of rats with water extracts of crude and honey-processed Astragalus, and the composition changes of medicated sera of crude and honey-processed Astragalus were analyzed by using LC-MS technology. The cell survival rates were detected and the concentrations of medicated sera were screened through CCK-8 assay. The differences of cell phagocytic rates, ATP energy metabolism, and NO secretion between crude and honey-processed Astragalus were evaluated by using neutral red phagocytosis assay, ATP detection kit, and NO detection kit. The effects of crude and honey-processed Astragalus on TNF-α secretion of RAW264.7 cells were detected by employing ELISA kit. The effects of crude and honey-processed Astragalus on polarization of RAW264.7 cells were evaluated by utilizing flow cytometry. The differential metabolites related to glycolysis in cell lysates and culture media were screened by using LC-MS technology. The experiment was approved by the experimental animal ethics committee from Shanxi University of Chinese Medicine (No. AWE202407352). The results showed that the contents of betaine, amino acids, and ononin in the prepared medicated sera of rats treated by intragastric administration with water extracts of honey-processed Astragalus increased compared to those in crude one. The results of CCK-8 experiment showed that there were no cytotoxic effects on RAW264.7 cells in the medicated sera of crude and honey-processd Astragalus at different concentration. The phagocytic index and ATP yield both increased to varying degrees after administration of the medicated sera to cells. The secretion of NO in normal and inflammatory cells increased and decreased respectively, and the effect of honey-processed Astragalus was better than that of crude one. The results of ELISA kit showed that the medicated sera of both crude and honey-processed Astragalus could promote the secretion of TNF-α in a concentration-dependent manner, and the promoting effect of honey-processed Astragalus was stronger than that of crude one. The results of flow cytometry showed that the medicated sera of both crude and honey-processed Astragalus could promote M1-type polarization and inhibit M2-type polarization of RAW264.7 cells, and the effect of honey-processed Astragalus was better than that of crude one. Compared to crude Astragalus, the metabolites related to glycolysis in the cell lysates and culture media of the medicated sera of honey-processed Astragalus were generally on the rise, indicating that the effect on promoting glycolysis of honey-processed Astragalus was better than that of crude one. In summary, honey-processed Astragalus can promote the polarization and energy metabolism of RAW264.7 cells, and participate in positive immune regulation, which is correlated with its enhanced effect of invigorating spleen-stomach and replenishing Qi.

Biliary duct injury,congenital biliary atresia,biliary tract tumors,primary sclerosing cholangitis,etc.are common and refractory diseases in the digestive system in clinical practice.The existing surgical operations and drug treatments demonstrate limited effects.Organoids,as an emerging technology,have attracted much attention in recent years for deeply understanding the pathogenesis and development of these diseases and seeking more effective treatment approaches.An organoid,a three-dimensional complex derived from stem/progenitor cells,can simulate the complex structure and physiological function of tissues or organs in vitro.It provides an important platform for studying the pathogenesis of biliary tract diseases and brings new hope for the repair and regeneration of biliary tract injury.The seed cells for constructing biliary organoids are mainly biliary tract epithelial cells,pluripotent stem cells,etc.The conventional technologies for constructing biliary organoids mainly include embedding,rotary culture,and hanging drop culture.In recent years,new culture technologies such as organ chip and three-dimensional and four-dimensional printing are emerging.This article reviews the construction methods of biliary organoids,discusses the application of these organoids in disease model construction,disease mechanism research,drug screening,and tissue/organ repair,and proposes the current problems and future research directions of biliary organoids,which will provide reference for treating common refractory digestive system diseases in clinical practice.
Organoids ; Humans ; Tissue Engineering/methods* ; Biliary Tract/cytology*

Objective: To investigate the changing trends for associations of anxiety and depressive symptoms with smartphone addiction among middle school students, so as to provide a scientific basis for preventing smartphone addiction in middle school students. Methods: From 2022 to 2023, a method of combining convenient sampling with cluster sampling was used to select 8 923 middle school students from 27 junior high schools and 3 senior high schools in a district of Shenzhen City between September 2022 (baseline, T1) and September 2023 (follow up, T2). The Smartphone Addiction Scale-Short Version (SAS-SV), Patients Health Questionnaire-9 Item (PHQ-9), and Generalized Anxiety Disorder 7-item Scale (GAD-7) were administered to assess smartphone addiction, anxiety and depressive symptoms. Mixed effects models were used to analyze the association of anxiety and depressive symptoms with smartphone addiction among middle school students. Results: From September 2022 to September 2023, the reported prevalence of smartphone addiction increased from 24.22% to 25.25% ( χ 2=45.71); and smartphone addiction scores [ 24.00 (16.00, 32.00),25.00(16.00, 33.00)], anxiety symptom scores [2.00(0.00, 7.00),3.00(0.00, 7.00)] and depressive symptom scores[3.00(0.00, 8.00),5.00(0.00, 9.00)] all significantly increased ( Z =-17.43, -42.38, -41.57) (all P <0.05). There were statistically significant difference in the distribution of anxiety and depression symptom levels among middle school students in 2022 and 2023 ( χ 2=85.15, 106.85, both P <0.05). After adjusting for covariates such as age, gender and family background, mixed effects models revealed dose response associations of anxiety and depressive symptoms with smartphone addiction among middle school students:mild anxiety symptom( OR =3.22), moderate to severe anxiety symptom ( OR =5.36), mild depressive symptom ( OR =3.32) and moderate to severe depressive symptom ( OR =6.13) were significantly associated with higher risks of smartphone addiction (all P <0.05). Interaction effect analysis found that co existing anxiety and depressive symptoms synergistically increased addiction risk by 5.60 times ( OR =5.60) compared to the asymptomatic group, with 32% of the combined risk attributable to their interaction ( S=1.64, AP =0.32)(both P < 0.05 ). Conclusions Anxiety and depressive symptoms are significantly associated with smartphone addiction, exhibiting a synergistic effect. Attention should be paid to emotional issues and smartphone addiction among middle school students.