Objective To investigate the potential causal associations between 3 common subjective sleep problems,daytime sleepiness,insomnia,and sleep apnea,and epigenetic age acceleration.Methods A two-sample Mendelian randomization(MR)study was conducted the publicly available genome-wide association study(GWAS)data to assess the causal effects of self-reported sleep problems on 4 indicators of epigenetic age acceleration,including intrinsic epigenetic age acceleration(IEAA),GrimAge acceleration,HannumAge acceleration,and PhenoAge acceleration.The primary analytical method was inverse-variance weighting(IVW),supplemented by sensitivity analyses using MR-Egger regression and the weighted median(WM)method to examine the robustness of the findings.Results IVW showed that daytime sleepiness was significantly positively associated with IEAA(β=1.83,95%CI:0.17～2.49,P=0.031)and HannumAge acceleration(β=1.81,95%CI:0.21～2.42,P=0.027),suggesting a potential accelerating effect on epigenetic aging.Insomnia also showed significant positive associations with IEAA(β=1.57,95%CI:0.40～2.73,P=0.009)and GrimAge acceleration(β=1.37,95%CI:0.18～2.56,P=0.024).No significant causal relationship was observed between sleep apnea and any indicator of epigenetic age acceleration(P>0.05).Conclusion Daytime sleepiness and insomnia may accelerate epigenetic aging,indicating their potential biological role in the aging process.The impact of sleep apnea remains unclear and warrants further investigation.

AIM:To elucidate the pharmacody-namic and network pharmacological mechanisms of total flavonoids of Carthamus tinctorius L.,to ex-plore their key targets and related pathways,and to clarify their mechanism of action against hepatic fibrosis.METHODS:The total flavonoids of Cartha-mus tinctorius L.were determined by LC-MS/MS and analysed for their compositions;the active in-gredients were screened by TCMSP database,SWISS ADME database and literature search;the targets related to total flavonoids of Carthamus tinctorius L.were screened by Swiss Target Predic-tion database;and the targets related to hepatic fi-brosis were screened by GeneCards database;the anti-hepatic fibrosis targets of total flavonoids of Carthamus tinctorius L.were obtained by taking the intersection of Venny.2.1.0;the protein interac-tions were analysed by STRING database;the visu-alization analysis was carried out by Cytoscape soft-ware;the GO function and KEGG pathway analysis was carried out by Metascape platform;and molec-ular docking was verified by using AutoDock soft-ware for the core targets and active ingredients.The mechanism of anti-hepatic fibrosis of total fla-vonoids of Carthamus tinctorius L.was verified by animal model and in vitro cell experiments.RE-SULTS:A total of 41 flavonoid components were identified in Carthamus tinctorius L.Through the network pharmacological analysis,149 anti-hepatic fibrosis targets of total flavonoids of Carthamus tinctorius L.were obtained,including 23 core tar-gets.The GO enrichment analyses involved a total of three aspects,namely,biological process(BP),cellular component(CC),and molecular function(MF).KEGG enrichment results showed that PI3K/Akt and MAPK are pathways involved in the devel-opment of hepatic fibrosis.Molecular docking veri-fied that the active ingredients Quercetin,Acacetin and Glabridin were tightly bound to Akt1 and HI-FIA,respectively.In animal model experiments,it was observed by HE and Masson staining that fibro-plasia was reduced,collagen deposition was re-duced,inflammatory cell infiltration was reduced,and fibrotic liver tissues were improved in total fla-vonoids of Carthamus tinctorius L.administration group.In isolated cell experiments:Western blot-ting results suggested that total flavonoids of Car-thamus tinctorius L.could decrease the hepatic fi-brosis marker factor α-SMA,Collagen1(P＜0.01)and PI3K,Akt protein expression(P＜0.01).CONCLU-SION:Total flavonoids of Carthamus tinctorius L.ex-erted anti-hepatic fibrosis effects through multi-components,multi-targets and multi-pathways,and their mechanism of action may be achieved by regulating the PI3K/Akt signalling pathway.

Objective·To identify and evaluate novel and reliable non-invasive serological biomarkers for detecting pancreatic ductal adenocarcinoma(PDAC).Methods·Sixty-seven PDAC patients(Ruijin cohort Ⅰ)were recruited at Pancreatic Center,Ruijin Hospital,Shanghai Jiao Tong University School of Medicine,from May 2018 to December 2019.Global proteome profiling of 67 PDAC tumor tissues and 67 matched adjacent normal tissues was performed using mass spectrum.Bioinformatics analysis on the proteomics data was conducted to identify new biomarkers,and receiver operating characteristic(ROC)curves and the area under the curve(AUC)were used to evaluate their value of detecting PDAC.The proteomic and mRNA sequencing data were further downloaded and analysed from the Clinical Proteomic Tumor Analysis Consortium(CPTAC)cohort for validation.In addition,the Ruijin Cohort Ⅱ,consisting of 47 PDAC patients and 75 healthy individuals,was recruited for a case-control study from June 2021 to June 2022.Enzyme-linked immunosorbent assay(ELISA)was used to detect the expression level of new biomarkers in the serum of patients and healthy individuals to evaluate the serological diagnostic values of them.Results·Collagen type Ⅻ α1 chain(COL12A1)was identified as a candidate biomarker for PDAC diagnosis based on differential expression analysis on the proteomic data and was validated to be higher in tumor tissues than in adjacent normal tissues in the CPTAC cohort.In addition,COL12A1 protein levels were significantly higher in the sera of PDAC patients than in those of healthy controls,showing good diagnostic performance with an AUC of 0.82,a sensitivity of 81%,and a specificity of 83%.ROC analysis revealed that COL12A1 improved the performance of carbohydrate antigen 199(CA199)in distinguishing PDAC patients from healthy individuals(AUCCA199=0.91 vs AUCCA199+COL12A1=0.95,P<0.05).Furthermore,COL12A1 also showed excellent ability to distinguish early-stage PDAC patients(stage Ⅰ?Ⅱ)from healthy individuals(AUCCOL12A1=0.83),and significantly improved the AUC of CA199 in early-stage PDAC patients(AUCCA199=0.92 vs AUCCA199+COL12A1=0.97,P<0.05).Conclusion·COL12A1 is a potential serological diagnostic marker that complements CA199 in detecting early-stage PDAC.

OBJECTIVE: To explore the peripheral neural mechanism underlying representation along the distribution of stomach meridian induced by intestinal inflammatory reaction using diarrhea predominant-irritable bowel syndrome (IBS-D) mice. METHODS: Among 62 healthy male C57BL/6 mice of clean grade, 12 mice were randomly selected and divided into a control group and a model group, 6 mice in each group, additionally, 12 mice were randomly selected and divided into a Tianshu group, a Liangqiu group and a Zusanli group, 4 mice in each group. In the model group, citrobacter was administered orally to establish IBS-D model. In the control group and the model group, the visceral pain threshold was observed using fecal colorectal distension (fCRD) induced electromyography of external oblique muscle, the positive cell number of neutrophil in the colonic muscularis was detected by myeloperoxidase (MPO) staining, the number, location and distribution rule of Evans blue (EB) extravasation points were observed by injection of EB staining solution into the tail vein. In the Tianshu group, the Liangqiu group and the Zusanli group, fluorescent dye Dil was injected at bilateral "Tianshu" (ST25), "Liangqiu" (ST34) and "Zusanli" (ST36) respectively, to observe the dye-positive cell number in different dorsal root ganglion (DRG) segments. In the control group and the model group, the activation of satellite glial cells (SGCs) in different DRG segments was observed by immunofluorescence. RESULTS: Compared with the control group, in the model group, the area under curve of electromyography of external oblique muscle was increased at fCRD of 25, 50 and 75 μL distilled water (P<0.001, P<0.01); the MPO-positive cell number of neutrophil in the colonic muscularis was increased (P<0.01). Few EB extravasation points could be found in the control group, while there were much more EB extravasation points observed in the model group, which was specially distribution in the area of stomach meridian, from "Huaroumen" (ST24) to "Zusanli" (ST36), as well as the surface area dominated by L₂-L₅ segment of the spinal cord. The Dil-positive cells were mainly exhibited in the DRG of T₁₁, L₅ and L₄ segments in the Tianshu group, the Liangqiu group and the Zusanli group, respectively. Compared with the control group, the ratio of glial fibrillary acidic protein (GFAP)/glutamine synthetase (GS) co-expression was increased in the DRG of T₁₁, L₄ and L₅ segments in the model group (P<0.05, P<0.01). CONCLUSION The activation of SGCs within DRG of T₁₁, L₄ and L₅ segments may relate closely to the occurrence of the representation along the stomach meridian distribution in IBS-D mice.
Animals ; Male ; Mice ; Irritable Bowel Syndrome/therapy* ; Mice, Inbred C57BL ; Meridians ; Stomach/physiopathology* ; Humans ; Acupuncture Points ; Disease Models, Animal

OBJECTIVE: To observe the effects of acupoint catgut embedding (ACE) on gut microbiota and fecal short-chain fatty acids (SCFAs) levels in patients with Parkinson's disease (PD) with constipation. METHODS: A total of 80 PD patients with constipation were randomly divided into an observation group and a control group, 40 cases in each group. Additionally, 40 healthy individuals were recruited as a healthy control group. The control group received conventional Western medical treatment for PD combined with polyethylene glycol (PEG), once daily for eight weeks. The observation group received additional ACE treatment at bilateral Tianshu (ST25), Zusanli (ST36), and Shangjuxu (ST37), once every two weeks for eight weeks. The healthy control group received no intervention. The spontaneous bowel movements (SBMs) per week and patient assessment of constipation quality of life (PAC-QOL) scores were assessed at baseline and after treatment in the two groups. Fecal samples were collected at the end of treatment for the observation and the control groups and at baseline for the healthy control group. Gut microbiota composition and diversity were analyzed using 16S rRNA method, and SCFA levels were measured using high-performance liquid chromatography (HPLC). RESULTS: Compared before treatment, the observation group showed a significant increase in SBMs (P<0.01), and PAC-QOL scores including physical discomfort, psychosocial discomfort, worry and concern, and total score were significantly reduced (P<0.01) after treatment; the control group also showed a reduction in PAC-QOL total score after treatment (P<0.01). After treatment, the observation group had significantly more SBMs (P<0.01), and lower PAC-QOL physical discomfort, psychosocial discomfort, worry and concern scores, and total score (P<0.01), and higher PAC-QOL satisfaction score (P<0.01) than the control group. Compared with the healthy control group, the control group showed decreased Chao1 and Ace indices (P<0.01). Compared with the healthy control group, the relative abundance of Prevotella and Roseburia was increased (P<0.05), while that of Enterobacter and Ruminococcus torques (six species in total) was decreased (P<0.05) in the control group. Compared with the control group, the observation group had increased relative abundance of Dialister, Parabacteroides, and Ruminococcus torques (P<0.05), and decreased relative abundance of Prevotella and Eubacterium ruminantium (P<0.05). Compared with the healthy control group, the control group had increased fecal SCFA levels (P<0.05); compared with the control group, the observation group had reduced fecal SCFA levels (P<0.05). Compared with the healthy control group, acetic acid, propionic acid, and butyric acid levels were elevated in the control group (P<0.05); compared with the control group, acetic acid, propionic acid, and butyric acid levels were decreased in the observation group (P<0.05). CONCLUSION ACE could increase spontaneous bowel movements and improve the quality of life in PD patients with constipation, which may be related to the regulation of gut microbiota composition and SCFA levels.
Humans ; Constipation/metabolism* ; Male ; Gastrointestinal Microbiome ; Acupuncture Points ; Female ; Middle Aged ; Parkinson Disease/complications* ; Aged ; Fatty Acids, Volatile/metabolism* ; Catgut ; Feces/microbiology* ; Acupuncture Therapy ; Quality of Life ; Adult

Objective To explore the relationship between the inhibitory effect of quercetin on AC16 cardiomyocyte proliferation and the Wnt/β-catenin signaling pathway,the relationship between quercetin(quercetin,QCT)and the proliferation of AC16 cardiomyocytes through in vitro was investigated.Methods AC16 cells were stimulated with different concentrations of QCT.The effects of QCT on AC16 cell proliferation were detected by inverted microscope photography,trypan blue counting,and CCK-8 assay.Western blot was used to detect the effects of QCT on the expression of phosphorylated β-catenin(p-β-catenin),c-Myc,β-catenin,low density lipoprotein receptor-related protein 5(LRP5),and LRP6.The effect of quercetin on cell proliferation was detected after overexpressing β-catenin ΔN,LRP5,and LRP6 genes,and after silencing LRP5 and LRP6 genes by trypan blue counting.Results Compared with the control group,QCT could decrease the number of AC16 cells and inhibit the proliferation rate,which was concentration-dependent.At the protein expression level,10 and 20 μmol/L QCT led to an significant upregulated modification of p-β-catenin protein(P<0.05)and significant downregulation of c-Myc,β-catenin,LRP5,and LRP6 protein expression(P<0.05)in AC16 cells.Therefore,10 μmol/L QCT was chosen as the intervention concentration.Overexpression of β-catenin ΔN,LRP5,and LRP6 genes in AC16 cells significantly rescued the cell proliferation inhibition caused by 10 μmol/L QCT compared to the drug-only group(P<0.05).Conversely,silencing LRP5 and LRP6 genes led to inhibition of AC16 cell proliferation,and the combination with 10 μmol/L QCT did not exacerbate the inhibition(P>0.05).Conclusion The quercetin could inhibit the Wnt/β-catenin signaling pathway via significant downregulation of LRP5/6,thereby attenuate cell proliferation of AC16 cardiomyocytes.

Objective·To identify and evaluate novel and reliable non-invasive serological biomarkers for detecting pancreatic ductal adenocarcinoma(PDAC).Methods·Sixty-seven PDAC patients(Ruijin cohort Ⅰ)were recruited at Pancreatic Center,Ruijin Hospital,Shanghai Jiao Tong University School of Medicine,from May 2018 to December 2019.Global proteome profiling of 67 PDAC tumor tissues and 67 matched adjacent normal tissues was performed using mass spectrum.Bioinformatics analysis on the proteomics data was conducted to identify new biomarkers,and receiver operating characteristic(ROC)curves and the area under the curve(AUC)were used to evaluate their value of detecting PDAC.The proteomic and mRNA sequencing data were further downloaded and analysed from the Clinical Proteomic Tumor Analysis Consortium(CPTAC)cohort for validation.In addition,the Ruijin Cohort Ⅱ,consisting of 47 PDAC patients and 75 healthy individuals,was recruited for a case-control study from June 2021 to June 2022.Enzyme-linked immunosorbent assay(ELISA)was used to detect the expression level of new biomarkers in the serum of patients and healthy individuals to evaluate the serological diagnostic values of them.Results·Collagen type Ⅻ α1 chain(COL12A1)was identified as a candidate biomarker for PDAC diagnosis based on differential expression analysis on the proteomic data and was validated to be higher in tumor tissues than in adjacent normal tissues in the CPTAC cohort.In addition,COL12A1 protein levels were significantly higher in the sera of PDAC patients than in those of healthy controls,showing good diagnostic performance with an AUC of 0.82,a sensitivity of 81%,and a specificity of 83%.ROC analysis revealed that COL12A1 improved the performance of carbohydrate antigen 199(CA199)in distinguishing PDAC patients from healthy individuals(AUCCA199=0.91 vs AUCCA199+COL12A1=0.95,P<0.05).Furthermore,COL12A1 also showed excellent ability to distinguish early-stage PDAC patients(stage Ⅰ?Ⅱ)from healthy individuals(AUCCOL12A1=0.83),and significantly improved the AUC of CA199 in early-stage PDAC patients(AUCCA199=0.92 vs AUCCA199+COL12A1=0.97,P<0.05).Conclusion·COL12A1 is a potential serological diagnostic marker that complements CA199 in detecting early-stage PDAC.

AIM:To elucidate the pharmacody-namic and network pharmacological mechanisms of total flavonoids of Carthamus tinctorius L.,to ex-plore their key targets and related pathways,and to clarify their mechanism of action against hepatic fibrosis.METHODS:The total flavonoids of Cartha-mus tinctorius L.were determined by LC-MS/MS and analysed for their compositions;the active in-gredients were screened by TCMSP database,SWISS ADME database and literature search;the targets related to total flavonoids of Carthamus tinctorius L.were screened by Swiss Target Predic-tion database;and the targets related to hepatic fi-brosis were screened by GeneCards database;the anti-hepatic fibrosis targets of total flavonoids of Carthamus tinctorius L.were obtained by taking the intersection of Venny.2.1.0;the protein interac-tions were analysed by STRING database;the visu-alization analysis was carried out by Cytoscape soft-ware;the GO function and KEGG pathway analysis was carried out by Metascape platform;and molec-ular docking was verified by using AutoDock soft-ware for the core targets and active ingredients.The mechanism of anti-hepatic fibrosis of total fla-vonoids of Carthamus tinctorius L.was verified by animal model and in vitro cell experiments.RE-SULTS:A total of 41 flavonoid components were identified in Carthamus tinctorius L.Through the network pharmacological analysis,149 anti-hepatic fibrosis targets of total flavonoids of Carthamus tinctorius L.were obtained,including 23 core tar-gets.The GO enrichment analyses involved a total of three aspects,namely,biological process(BP),cellular component(CC),and molecular function(MF).KEGG enrichment results showed that PI3K/Akt and MAPK are pathways involved in the devel-opment of hepatic fibrosis.Molecular docking veri-fied that the active ingredients Quercetin,Acacetin and Glabridin were tightly bound to Akt1 and HI-FIA,respectively.In animal model experiments,it was observed by HE and Masson staining that fibro-plasia was reduced,collagen deposition was re-duced,inflammatory cell infiltration was reduced,and fibrotic liver tissues were improved in total fla-vonoids of Carthamus tinctorius L.administration group.In isolated cell experiments:Western blot-ting results suggested that total flavonoids of Car-thamus tinctorius L.could decrease the hepatic fi-brosis marker factor α-SMA,Collagen1(P＜0.01)and PI3K,Akt protein expression(P＜0.01).CONCLU-SION:Total flavonoids of Carthamus tinctorius L.ex-erted anti-hepatic fibrosis effects through multi-components,multi-targets and multi-pathways,and their mechanism of action may be achieved by regulating the PI3K/Akt signalling pathway.

Objective To explore the relationship between the inhibitory effect of quercetin on AC16 cardiomyocyte proliferation and the Wnt/β-catenin signaling pathway,the relationship between quercetin(quercetin,QCT)and the proliferation of AC16 cardiomyocytes through in vitro was investigated.Methods AC16 cells were stimulated with different concentrations of QCT.The effects of QCT on AC16 cell proliferation were detected by inverted microscope photography,trypan blue counting,and CCK-8 assay.Western blot was used to detect the effects of QCT on the expression of phosphorylated β-catenin(p-β-catenin),c-Myc,β-catenin,low density lipoprotein receptor-related protein 5(LRP5),and LRP6.The effect of quercetin on cell proliferation was detected after overexpressing β-catenin ΔN,LRP5,and LRP6 genes,and after silencing LRP5 and LRP6 genes by trypan blue counting.Results Compared with the control group,QCT could decrease the number of AC16 cells and inhibit the proliferation rate,which was concentration-dependent.At the protein expression level,10 and 20 μmol/L QCT led to an significant upregulated modification of p-β-catenin protein(P<0.05)and significant downregulation of c-Myc,β-catenin,LRP5,and LRP6 protein expression(P<0.05)in AC16 cells.Therefore,10 μmol/L QCT was chosen as the intervention concentration.Overexpression of β-catenin ΔN,LRP5,and LRP6 genes in AC16 cells significantly rescued the cell proliferation inhibition caused by 10 μmol/L QCT compared to the drug-only group(P<0.05).Conversely,silencing LRP5 and LRP6 genes led to inhibition of AC16 cell proliferation,and the combination with 10 μmol/L QCT did not exacerbate the inhibition(P>0.05).Conclusion The quercetin could inhibit the Wnt/β-catenin signaling pathway via significant downregulation of LRP5/6,thereby attenuate cell proliferation of AC16 cardiomyocytes.

OBJECTIVE To excavate the adverse drug event （ADE） signals of three third-generation tetracycline antibiotics （tigecycline， omadacycline， eravacycline） based on FDA adverse event reporting system （FAERS）， and to provide reference for the safe use of them. METHODS The ADE reports of tigecycline， omadacycline and eravacycline from the first quarter of 2005 to the second quarter of 2023 were retrieved from FAERS database. The ADE signals of 3 kinds of drugs were mined by the method of reporting odds ratio method and the proportional reporting ratio method. RESULTS Totally 2 538 ADE reports with tigecycline， omadacycline and eravacycline as the primary suspected drugs were obtained， including 2 135 tigecycline ADE reports， 349 omadacycline ADE reports and 54 eravacycline ADE reports. A total of 131 ADE positive signals of tigecycline were mined， involving 19 system organ classes （SOCs）， mainly concentrated in investigations， hepatobiliary system， blood and lymphatic system， and gastrointestinal system， etc； the preferred terminologies （PT） with intense signal were hypofibrinogenaemia and blood fibrinogen decreased. Fourteen ADE signals were not mentioned in the drug instruction， such as renal failure， acute kidney injury and hemorrhage. Totally 24 ADE positive signals of omadacycline were mined， involving 6 SOCs， mainly concentrated in the gastrointestinal system and various examinations； the PTs with intense signals were tooth discoloration， jet-like vomiting and loose feces， etc. ADE signals were not mentioned in the drug instructions， included lip swelling， gastroesophageal reflux disease， eosinophilia， skin discoloration， feces softening， and night sweats. Five ADE positive signals of eravacycline were mined， involving 4 SOCs， mainly concentrated in various examinations， gastrointestinal system， etc. The most intense ADE signals were blood fibrinogen decreased and hypofibrinogenaemia. CONCLUSIONS ADE of the gastrointestinal system are mostly identified in the three third-generation tetracycline antibiotics， especially pancreatitis caused by tigecycline and gastroesophageal reflux disease caused by oral administration of omadacycline. The liver function， renal function （for tigecycline） and coagulation function （for tigecycline， eravacycline） should be monitored biyiliang@hotmail.com regularly during medication， so as to prevent the occurrence of serious ADE.