ObjectiveTo explore the mechanism of action of Paeoniae Radix Rubra and Aconiti Lateralis Radix Praeparata （CSFZ） in the treatment of acute-on-chronic liver failure （ACLF） through network pharmacology， molecular docking， and animal experiments. MethodsNetwork pharmacology was used to identify potential targets and related signaling pathways for the treatment of ACLF with CSFZ. Molecular docking was used to examine the binding activity of the core components with corresponding key targets. An ACLF rat model was established by subcutaneous and tail vein injections of bovine serum albumin combined with lipopolysaccharide （LPS） + D-galactosamine （D-GalN） intraperitoneal injection. A normal control group （NC）， a model group， a CSFZ group （CSFZ， 5.85 g·kg^-1）， and a hepatocyte growth-promoting granule group （HGFG， 4.05 g·kg^-1） were set up in this study. Pathological changes in rat liver tissue were observed using hematoxylin and eosin （HE） and Masson staining. Enzyme-linked immunosorbent assay （ELISA） was used to detect the expression levels of interleukin-6 （IL-6）， B-cell lymphoma-2 （Bcl-2）， Caspase-3， and albumin （ALB）. Real-time quantitative polymerase chain reaction （Real-time PCR） and Western blot were used to measure the mRNA and protein expression levels of phosphoinositide 3-kinase （PI3K）， protein kinase B （Akt）， phosphorylated PI3K （p-PI3K）， and phosphorylated Akt （p-Akt）. ResultsNetwork pharmacology screening identified 49 active ingredients of CSFZ， 103 action targets， and 3 317 targets related to ACLF. Among these， 74 targets overlapped with CSFZ drug targets. Key nodes in the protein-protein interaction （PPI） network included Akt1， tumor necrosis factor （TNF）， IL-6， Bcl-2， and Caspase-3. Gene Ontology （GO） functional analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analysis identified multiple signaling pathways， with the PI3K/Akt signaling pathway being the most frequent. Molecular docking showed that the core components of the drug exhibited good binding activity with the corresponding key targets. Animal experiments confirmed that CSFZ significantly improved liver tissue pathological damage in ACLF rats， reduced the release of inflammatory factors and liver cell apoptosis， and upregulated the expression levels of the PI3K/Akt signaling pathway. ConclusionThrough network pharmacology， molecular docking， and in vivo experiments， this study confirms the effect of CSFZ in reducing liver cell inflammatory damage and inhibiting liver cell apoptosis. The specific mechanism may be related to its involvement in regulating the PI3K/Akt signaling pathway.

ObjectiveTo study the regulatory effects of Yinchenhao Tang combined with Yinchen Zhufu Tang on the expression of regulatory T （Treg）/helper T 17 （Th17） cells cultured in vitro from the patients with hepatitis B virus-related acute-on-chronic liver failure （HBV-ACLF）. MethodsFresh peripheral blood was collected from the patients with HBV-ACLF for the separation of peripheral blood mononuclear cells （PBMCs）. Immunomagnetic beads were used to isolate primary Treg and naive CD4⁺ T cells. After in vitro expansion， naive CD4⁺ T cells were induced to differentiate into Th17 cells. Rats were treated with the clearing method （Yinchenhao Tang）， warming method （Yinchen Zhufu Tang）， and combination of clearing method with warming method （Yinchenhao Tang combined with Yinchen Zhufu Tang， also known as Wenyang Jiedu Huayu Prescription）， respectively， and then the medicated plasma samples were collected. Meanwhile， blank plasma was collected from the rats treated with normal saline. Cells were classified into blank， clearing method （5.04 g·kg^-1）， warming method （6.21 g·kg^-1）， and combination of clearing method with warming method （17.1 g·kg^-1） groups and treated with corresponding plasma. The frequency of Treg/Th17 cells was detected by flow cytometry. The level of transforming growth factor-β （TGF-β） was measured by the enzyme-linked immunosorbent assay. The cytometric bead array （CBA） was employed to measure the levels of interleukin-10 （IL-10）， interleukin-17A （IL-17A）， tumor necrosis factor-α （TNF-α）， and interleukin-23 （IL-23）. The mRNA and protein levels of Forkhead box P3 （FoxP3） and retinoic acid-related orphan receptor-gamma t （ROR-γt） were determined by Real-time PCR and Western blot， respectively. ResultsCompared with the blank group， the combination of clearing method with warming method group showed decreased frequency of Treg and Th17 cells， lowered levels of Treg cytokines （TGF-β and IL-10） and Th17 cytokines （TNF-α， IL-17A， and IL-23）， and down-regulated mRNA and protein levels of FoxP3 and ROR-γt （P<0.01）. Compared with the clearing method group， the combination of clearing method with warming method group showed decreased Treg cell frequency and down-regulated mRNA and protein levels of FoxP3. Meanwhile， the combination group showed decreased Th17 cell frequency， lowered levels of TGF-β， IL-10， TNF-α， IL-17A， and IL-23， and down-regulated mRNA and protein levels of ROR-γt （P<0.05， P<0.01）. Compared with the warming method group， the combination of clearing method with warming method group showed decreased frequency of Treg cells and down-regulated mRNA and protein levels of FoxP3. Meanwhile， the combination group showed decreased Th17 cell frequency， declined levels of TGF-β， IL-10， TNF-α， IL-17A， and IL-23， and down-regulated mRNA and protein levels of ROR-γt （P<0.05）. ConclusionThe combination of clearing method with warming method can down-regulate the expression of specific cytokines of Treg and Th17 cells， inhibit the over activation of Treg and Th17 cells， and reduce the secretion of cytokines such as TGF-β， IL-10， TNF-α， IL-17A， and IL-23， thereby alleviating inflammation and improving the prognosis of the patients with liver failure.

ObjectiveTo investigate the differential expression of intestinal flora in patients with different traditional Chinese medicine （TCM） syndrome types （Yang Huang syndrome， Yin-Yang Huang syndrome， and Yin Huang syndrome） of hepatitis B virus-related acute-on-chronic liver failure （HBV-ACLF） and clarify the biological basis of jaundice and Yin Huang syndrome in liver failure. MethodsA total of 20 cases of HBV-ACLF patients were included in the Yang Huang group， 20 cases in the Yin-Yang Huang group， 16 cases in the Yin Huang group， and 20 healthy adult volunteers. 16S rRNA gene sequencing was used to detect the diversity， species distribution， and differences of the subjects' intestinal flora， and bioinformatics analysis was conducted. ResultsCompared with those in the healthy control group， the species richness and diversity of intestinal flora in the HBV-ACLF Yang Huang group， Yin-Yang Huang group， and Yin Huang group were significantly reduced， and there were significant differences in the composition of intestinal flora compared with healthy volunteers. However， there were no significant differences in the species richness， diversity， and composition of intestinal flora among the three groups. LEfSe analysis showed that compared with the healthy control group， the HBV-ACLF Yang Huang group showed significant enrichment of Staphylococcus aureus（P<0.01）. Yin-Yang Huang group showed significant enrichment of s_Ileibacterium valens（P<0.05，P<0.01）， and the Yin Huang group showed significant enrichment of Enterococcus faecium and Streptococcus sali varius（P<0.05）. These strains may be biomarkers between the three groups of patients and the healthy control group. Compared with that in the Yin-Yang Huang group， Tyzzerella_nexilis was significantly enriched in the Yang-Huang group， and Streptococcus lactiae was significantly enriched in the Yin-Yang Huang group. Compared with that in the Yang-Huang group and the Yin-Yang Huang group， Enterococcus faecalis was significantly enriched in the Yin Huang group. The above strains may be biomarkers among the three groups of patients， and Enterococcus faecium may be a biomarker for the transition from the Yang Huang group to the Yin Huang group. ConclusionsThere are significant differences in the intestinal flora between patients with HBV-ACLF Yang Huang syndrome， Yin-Yang Huang syndrome， and Yin Huang syndrome. Enterococcus faecium is significantly enriched in the Yin Huang syndrome group， suggesting that dysbiosis of the intestinal flora may be the biological basis for jaundice and Yin Huang syndrome in liver failure.

ObjectiveTo study the expression differences of regulatory T cells （Treg） and T helper 17 cells （Th17） in the peripheral blood of patients with hepatitis B virus-related acute-on-chronic liver failure （HBV-ACLF） in five types of traditional Chinese medicine （TCM） syndrome. MethodsA total of 144 patients with HBV-ACLF were included and divided into five types of TCM syndrome， including 34 cases of heat-toxin amassment syndrome， 44 cases of dampness-heat amassment syndrome， 27 cases of Qi-deficiency and stasis jaundice syndrome， 21 cases of spleen-kidney Yang deficiency syndrome， and 18 cases of liver-kidney Yin deficiency syndrome. Meanwhile， 30 healthy volunteers were included as controls. The frequency of Treg and Th17 cells in the peripheral blood of subjects in each group was detected by flow cytometry， and the Treg/Th17 ratio was calculated. Cytometric bead array （CBA） was used to detect the levels of cytokines interleukin （IL）-10， transforming growth factor-β （TGF-β）， tumor necrosis factor-α （TNF-α）， IL-17A， and IL-23. Real-time fluorescence quantitative PCR （Real-time PCR） detected the mRNA expression of forkhead box P3 （FoxP3） and retinoic acid-related orphan receptor gamma t （ROR-γt）. Results（1） Compared with that in the healthy control group， the frequency of Treg and Th17 cells in patients with various TCM syndrome types of HBV-ACLF increased （P<0.05）. Compared with that in the heat-toxin amassment syndrome group， the frequency of Treg and Th17 cells decreased in the dampness-heat amassment syndrome group （P<0.05）， while the frequency of Treg and Th17 cells increased in the Qi-deficiency and stasis jaundice syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with that in the dampness-heat amassment syndrome group， the frequency of Treg and Th17 cells increased in the dampness-heat amassment syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with that in the Qi-deficiency and stasis jaundice syndrome group， the frequency of Treg and Th17 cells increased in the spleen-kidney Yang deficiency syndrome group （P<0.05）， while the frequency of Treg and Th17 cells decreased in the liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with the spleen-kidney Yang deficiency syndrome group， the frequency of Treg and Th17 cells decreased in the liver-kidney Yin deficiency syndrome group （P<0.05）. （2） Compared with that in the healthy control group， the Treg/Th17 cell ratio in patients with various TCM syndromes of HBV-ACLF decreased （P<0.05）. Compared with that in the heat-toxin amassment syndrome group， the Treg/Th17 cell ratio increased in the dampness-heat amassment syndrome group （P<0.05）， while it decreased in the Qi-deficiency and stasis jaundice syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with that in the dampness-heat amassment syndrome group， the Treg/Th17 cell ratio decreased in the Qi-deficiency and stasis jaundice syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with that in the Qi-deficiency and stasis jaundice syndrome group， the Treg/Th17 cell ratio increased in the spleen-kidney Yang deficiency syndrome group and liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with the spleen-kidney Yang deficiency syndrome group， the Treg/Th17 cell ratio in the liver-kidney Yin deficiency syndrome group increased （P<0.05）. （3） Compared with those in the healthy control group， the levels of Treg-related cytokines IL-10 and TGF-β， as well as Th17-related cytokines TNF-α， IL-17A， and IL-23， were elevated in patients with various TCM syndrome types of HBV-ACLF （P<0.05）. There was no significant difference in TNF-α levels among different TCM syndrome types. Compared with those in the heat-toxin amassment syndrome group， the levels of IL-10， TNF-β， IL-17A， and IL-23 in the dampness-heat amassment syndrome group， Qi-deficiency and stasis jaundice syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome groups increased （P<0.05）. Compared with those in the dampness-heat amassment syndrome group， the levels of IL-10， TGF-β， IL-17A， and IL-23 increased in the Qi-deficiency and stasis jaundice syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with those in the Qi-deficiency and stasis jaundice syndrome group， the levels of IL-10， TGF-β， IL-17A， and IL-23 in the spleen-kidney Yang deficiency syndrome group increased （P<0.05）， while those in the liver-kidney Yin deficiency syndrome group decreased （P<0.05）. Compared with those in the spleen-kidney Yang deficiency syndrome， the levels of IL-10， TGF-β， IL-17A， and IL-23 in the liver-kidney Yin deficiency syndrome group decreased （P<0.05）. （4） Compared with that in the healthy control group， the mRNA of Treg/Th17 cell specific transcription factors FoxP3 and ROR-γt were elevated in patients with various TCM syndrome types of HBV-ACLF （P<0.05）. Compared with that in the heat-toxin amassment syndrome group， the mRNA of FoxP3 and ROR-γt increased in the Qi-deficiency and stasis jaundice syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with that in the dampness-heat amassment syndrome group， the mRNA of FoxP3 and ROR-γt increased in the Qi-deficiency and stasis jaundice syndrome group， spleen-kidney Yang deficiency syndrome group， and liver-kidney Yin deficiency syndrome （P<0.05）. Compared with that in the Qi-deficiency and stasis jaundice syndrome group， the mRNA of FoxP3 and ROR-γt in the spleen-kidney Yang deficiency syndrome group increased （P<0.05）， and it decreased in the liver-kidney Yin deficiency syndrome group （P<0.05）. Compared with that in the spleen-kidney Yang deficiency syndrome group， the mRNA of FoxP3 and ROR-γt decreased in the liver-kidney Yin deficiency syndrome group （P<0.05）. ConclusionThe frequency and ratio of Treg/Th17 cells， as well as the expression of related cytokines and specific receptors in peripheral blood of patients with HBV-ACLF in five types of TCM syndromes are different， which has certain reference value for TCM syndrome differentiation and treatment of patients with HBV-ACLF.

Objective:To investigate the effects and molecular mechanisms of Wilms tumor 1-associated proteins(WTAP)on the cell biological properties of esophageal squamous cell carcinoma(ESCC)cells.Methods:31 pairs of ESCC tissues and their paired paracancerous tissues that were surgically resected at the Second Clinical Medical College of Chuanbei Medical College between September 2019 and April 2021 were collected.The esophageal cancer cells KYSE30,KYSE410,KYSE150,KYSE510,TE-1,and normal human esophageal epithelial cells HET-1A were routinely cultured.Transfection reagents were used to transfect si-NC,si-WTAP#1 and si-WTAP#2 nucleic acids into KYSE150 and KYSE510 cells.The cells were divided into si-NC,si-WTAP#1 and si-WTAP#2 groups.The expressions of WTAP and MAP3K9 mRNA were detected in the cells of each group by qPCR assay.CCK-8 assay,clone formation assay,and scratch healing assay,Transwell assay were employed to detect the effects of knockdown of WTAP expression on ESCC cell proliferation,migration,invasion and apoptosis.WB assay was used to detect the expressions of WTAP,MAP3K9,EMT and MAPK pathway-related proteins in ESCC cells of each group knocked down of WTAP;immunohistochemistry to detect the expression of WTAP proteins in ESCC tissues,immunoprecipitation of methylated RNA(MeRIP)-qPCR assay to detect the level of MAP3K9 m6A in ESCC cells,actinomycin D assay to detect the stability of mRNA of MAP3K9,and database data to analyze the expression,target genes,functional enrichment,and interacting RNA of WTAP.Results:WTAP was highly expressed in ESCC tissues and cells(P<0.05 or P<0.01 or P<0.001)and correlated with the degree of differentiation(P<0.01);the expression of WTAP mRNA and its protein were successfully knocked down in KYSE150 and KYSE510 cells(P<0.01 or P<0.001);the knockdown of WTAP significantly inhibited the proliferation,migration and invasion of KYSE150 and KYSE510 cells(P<0.05 or P<0.01 or P<0.001),and promoted the apoptosis of KYSE150 and KYSE510 cells(P<0.05 or P<0.01).Knockdown of WTAP resulted in a significant decrease in the m6A level of MAP3K9(P<0.05),and its mRNA expression level and mRNA stability were both significantly reduced(P<0.05).Database data analysis showed that WTAP target genes clustered in the MAPK signaling pathway;the expression levels of MAP3K9,p-ERK,N-cadherin,and MMP9 were significantly reduced(P<0.05 or P<0.01),and the expression level of E-cadherin was significantly elevated(P<0.05 or P<0.01)in the KYSE150 and KYSE510 cells after knockdown of WTAP.Conclusions:WTAP is highly expressed in ESCC tissues and cells and correlates with their differentiation.It promotes the stability of MAP3K9 mRNA through m6A modification,activates the MAPK pathway and thus promotes the malignant biological behaviors of ESCC cells.

Objective:To obtain a prevalent respiratory syncytial virus (RSV) clinical isolate in Beijing and analyze the genotype and biological characteristics of the strain.Methods:A nasopharyngeal secretion specimen was collected from a child with RSV infection in Beijing in 2023 and used for viral isolation. Viral nucleic acid was amplified using qRT-PCR. The isolated virus was identified by transmission electron microscopy, indirect immunofluorescence assay, and plaque formation assay. A phylogenetic analysis was conducted based on the whole-genome sequencing results. Virus titers were determined, and replication characteristics were analyzed. The efficacy of the isolated strain for in vitro screening of antiviral drugs was validated. Results:A clinical RSV isolate, named hRSV/C-Tan/BJ 202301, was successfully isolated, which could form syncytia in Hep-2 cells. Spherical, filamentous, and irregular virus particles were observed by electron microscopy. Immunofluorescence detection showed green fluorescence in Hep-2 cells, and plaque assay showed round plaques, which were similar to the Long strain in morphology. Genomic sequence analysis showed that it belonged to ON1 genotype. It exhibited similar cell growth kinetics characteristics with the Long strain and could be used for antiviral drug screening in vitro. Conclusions:In this study, one RSV strain is successfully isolated and identified. The biological characteristics and the phylogenetic relationship of this strain reflect the characteristics of the circulating strains in Beijing, which provides experimental material for RSV vaccine development and antiviral drug screening in China.

BACKGROUND: The mechanisms distinguishing metabolically healthy from unhealthy phenotypes within the same BMI categories remain unclear. This study aimed to investigate the associations between regional fat distribution and metabolically unhealthy phenotypes in Chinese adults across different BMI categories. METHODS: This cross-sectional study involving 11833 Chinese adults aged 20 years and older. Covariance analysis, adjusted for age, compared the percentage of regional fat (trunk, leg, or arm fat divided by whole-body fat) between metabolically healthy and unhealthy participants. Trends in regional fat percentage with the number of metabolic abnormalities were assessed by the Jonckheere-Terpstra test. Odds ratios (ORs) and their 95% confidence intervals (CIs) were estimated by logistic regression models. All analyses were performed separately by sex. RESULTS: In non-obese individuals, metabolically unhealthy participants exhibited higher percent trunk fat and lower percent leg fat compared to healthy participants. Additionally, percent trunk fat increased and percent leg fat decreased with the number of metabolic abnormalities. After adjustment for demographic and lifestyle factors, as well as BMI, higher percent trunk fat was associated with increased odds of being metabolically unhealthy [highest vs. lowest quartile: ORs (95%CI) of 1.64 (1.35, 2.00) for men and 2.00 (1.63, 2.46) for women]. Conversely, compared with the lowest quartile, the ORs (95%CI) of metabolically unhealthy phenotype in the highest quartile for percent arm and leg fat were 0.64 (0.53, 0.78) and 0.60 (0.49, 0.74) for men, and 0.72 (0.56, 0.93) and 0.46 (0.36, 0.59) for women, respectively. Significant interactions between BMI and percentage of trunk and leg fat were observed in both sexes, with stronger associations found in individuals with normal weight and overweight. CONCLUSIONS Trunk fat is associated with a higher risk of metabolically unhealthy phenotype, while leg and arm fat are protective factors. Regional fat distribution assessments are crucial for identifying metabolically unhealthy phenotypes, particularly in non-obese individuals.
Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; Young Adult ; Adipose Tissue ; Body Fat Distribution ; Body Mass Index ; China/epidemiology* ; Cross-Sectional Studies ; Health Surveys ; Phenotype

OBJECTIVE: To evaluate the efficacy and safety of a hospital-made resuscitation pack, a Chinese medicinal herbal compound formula designed to enhance recovery in post-bronchoscopy patients. METHODS: In this randomized, single-blind, placebo-controlled clinical trial, eligible patients were randomly assigned 1:1 to either the treatment or control groups. The patients in the treatment group applied the resuscitation pack, which contained aromatic compounded Chinese herbs. The patients in the control group applied a hospital-made, single herb placebo pack. Packs were placed on the Tiantu (CV 22) acupuncture point for 4 h as soon as the bronchoscopy finished. Efficacy indicators, such as recovery time, patients' symptoms including nausea and dizziness, and adverse events (AEs) were observed and compared. The outcome indices were evaluated at baseline, 1 and 24 h after the bronchoscopy. Subgroup analysis was further performed by patients' age and depth of sedation. RESULTS: When applying generalized estimating equations (GEE) to evaluate the intensity of post-bronchoscopy nausea and vomiting, the intensity was lower in the treatment group (163 cases) compared with the control group (162 cases; 95% CI: 0.004, 0.099, P=0.03]. Also, significantly lower intensity of nausea was observed in the 60-70 years of age subgroup (95% CI: 0.029, 0.169, P=0.006) and deep sedation subgroup (95% CI: 0.002, 0.124; P=0.04). There was no significant difference in dizziness between two groups by GEE (95% CI: -0.134, 0.297; P=0.459). In addition, no serious AEs were observed in either group. CONCLUSIONS Our study found that the resuscitation pack markedly improved patients' symptoms by reducing nausea and vomiting after bronchoscopy without AEs, compared with placebo in the perioperative period. (Trial registration No. ChiCTR2000038299).
Humans ; Male ; Middle Aged ; Female ; Bronchoscopy/adverse effects* ; Single-Blind Method ; Aged ; Drugs, Chinese Herbal/adverse effects* ; Treatment Outcome ; Resuscitation ; Adult ; Medicine, Chinese Traditional

Objective To compare the efficacy of 2 L and 3 L polyethylene glycol(PEG)electrolyte solution for bowel preparation in a real-world setting.Methods A real-world,single-center cohort study was conducted on the individuals undergoing colonoscopy in Department of Gastroenterology of Chengdu Third People's Hospital between May and October 2023.Based on our inclusion and exclusion criteria,they were given 2 L(n=4 684)and 3 L(n=3 700)PEG electrolyte solution for bowel preparation.The primary outcome indicator was the adequacy of bowel preparation by Boston bowel preparation score(BBPS).Secondary outcome indicators included the BBPS score,polyp detection rate(PDR),tolerability,compliance,and incidence of adverse events.Results The adequacy rate of bowel preparation was 94.35％in the 3 L group,significantly higher than that of the 2 L group(91.29％,P＜0.001).The 3 L group obtained a higher BBPS score then the 2 L group(6.92±1.06 vs 6.81±1.14,P＜0.001).But there was no statistical difference in the PDR between the 2 groups(P=0.073).And the rate of PEG completion(P=0.810),administration of low residue diet as required(P=0.094)or use of dimethicone(P=0.072)were comparable between the 2 groups.However,the incidences of vomiting(4.5％vs 3.2％,P=0.002),abdominal discomfort(5.0％vs 3.9％,P=0.011)and sleep disturbance(18.0％vs 14.6％,P＜0.001)were obviously higher in the 3 L group than the 2 L group.Conclusion In a real-world setting,2 L PEG is a considerably safe and effective regimen for bowel preparation.