Background/Aims: Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation. Methods: The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation. Results: MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs. Conclusions In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.

Background/Aims: Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation. Methods: The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation. Results: MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs. Conclusions In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.

ObjectiveTo study on the different therapeutic effects and potential mechanisms of Rhei Radix et Rhizoma（RH） before and after steaming with wine on constipation model mice. MethodsFifty-four male ICR mice were randomly divided into control group， model group， lactulose group（1.5 mg·kg^-1）， high， medium and low dose groups of RH and RH steaming with wine（PRH）（8， 4， 1 g·kg^-1）. Except for the control group， the constipation model was replicated by gavage of loperamide hydrochloride（6 mg·kg^-1） in the other groups. After 2 weeks of modeling， each administration group was gavaged with the corresponding dose of drug solution， and the control and model groups were given an equal volume of normal saline， 1 time/d for 2 consecutive weeks. After administration， the feces were collected for 16S rRNA sequencing， the levels of gastrin（GAS）， motilin（MTL）， interleukin-6（IL-6）， γ-interferon（IFN-γ） in the colonic tissue were detected by enzyme-linked immunosorbent assay（ELISA）， the histopathological changes of colon were observed by hematoxylin-eosin（HE） staining， flow cytometry was used to detect the proportion changes of CD4⁺， CD8⁺ and regulatory T cell（Treg） in peripheral blood. ResultsCompared with the control group， the model group showed significantly decrease in fecal number in 24 h， fecal quality and fecal water rate（P<0.01）， the colon was seen to have necrotic shedding of mucosal epithelium， localized intestinal glands in the lamina propria were degenerated， necrotic and atrophied， a few lymphocytes were seen to infiltrate in the necrotic area in a scattered manner， the contents of GAS and MTL， the proportions of CD4⁺， CD8⁺ and Treg were significantly reduced（P<0.01）， the contents of IL-6 and IFN-γ were significantly elevated（P<0.05， P<0.01）. Compared with the model group， the fecal number in 24 h， fecal quality and fecal water rate of high-dose groups of RH and PRH were significantly increased（P<0.05， P<0.01）， the pathological damage of the colon was alleviated to varying degrees， the contents of GAS， MTL， IL-6 and IFN-γ were significantly regressed（P<0.05， P<0.01）， and the proportions of CD4⁺ and CD8⁺ were significantly increased（P<0.01）， although the proportion of Treg showed an upward trend， there was no significant difference. In addition， the results of intestinal flora showed that the number of amplicon sequence variant（ASV） and Alpha diversity were decreased in the model group compared with the control group， and there was a significant difference in Beta diversity， with a decrease in the relative abundance of Lactobacillus and an increase in the relative abundances of Bacillus and Helicobacter. Compared with the model group， the ASV number and Alpha diversity were increased in the high-dose groups of RH and PRH， and there was a trend of regression of Beta diversity to the control group， the relative abundance of Lactobacillus increased， and the relative abundances of Bacillus and Helicobacter decreased. ConclusionRH and PRH can improve dysbacteriosis， promote immune system activation， inhibit the release of inflammatory factors for enhancing the gastrointestinal function， which may be one of the potential mechanisms of their therapeutic effect on constipation.

Metabolic-associated fatty liver disease (MAFLD) and osteoporosis (OP) are two very common metabolic diseases. A growing body of experimental evidence supports a pathophysiological link between MAFLD and OP. MAFLD is often associated with the development of OP. Rutaecarpine (RUT) is one of the main active components of Chinese medicine Euodiae Fructus. Our previous studies have demonstrated that RUT has lipid-lowering, anti-inflammatory and anti-atherosclerotic effects, and can improve the OP of rats. However, whether RUT can improve both fatty liver and OP symptoms of MAFLD mice at the same time remains to be investigated. In this study, we used C57BL/6 mice fed a high-fat diet (HFD) for 4 months to construct a MAFLD model, and gave the mice a low dose (5 mg·kg^-1) and a high dose (15 mg·kg^-1) of RUT by gavage for 4 weeks. The effects of RUT on liver steatosis and bone metabolism were then evaluated at the end of the experiment [this experiment was approved by the Experimental Animal Ethics Committee of Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences (approval number: IMB-20190124D₃03)]. The results showed that RUT treatment significantly reduced hepatic steatosis and lipid accumulation, and significantly reduced bone loss and promoted bone formation. In summary, this study shows that RUT has an effect of improving fatty liver and OP in MAFLD mice.

Background/Aims: Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation. Methods: The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation. Results: MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs. Conclusions In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.

Aim To investigate the synergistic sensiti-zation effect of saikosaponin D(SSD)combined with temozolomide(TMZ)on glioblastoma cells(GBM)and its molecular mechanism.Methods The sensitiv-ity of RG-2,U251 and LN-428 GBM cell lines to SSD and TMZ was analyzed by CCK-8 method combined with HE staining,and the optimal compatible concen-tration was screened.The effect of HE staining com-bined with Hoechst fluorescence staining on the prolif-eration of GBM cell line was detected by clonal forma-tion experiment.The autophagosome formation of GBM cells was observed by monodansylcadaverine(MDC)staining.The expression and distribution of endoplas-mic reticulum stress-related factors and apoptosis and autophagy proteins were detected by Western blot and ICC.Results The sensitivity order of GBM cells to TMZ was RG-2＞U251＞LN-428.The results of com-bined administration showed the synergistic inhibitory effect of SSD combined with TMZ on proliferation of GBM cell lines,which was confirmed by cell cloning formation experiment.Compared with the TMZ group,Hoechst fluorescence staining showed a significant in-crease in the number of nuclear bright staining in the combined administration group.MDC fluorescence staining showed that there were more dense green parti-cles in the cytoplasm of SSD/TMZ plus group than that of TMZ group.Western blot results showed that com-pared with TMZ group,the expression of ER stress markers GRP78,CHOP,p-PERK and ATF6 signifi-cantly increased in SSD/TMZ group(P＜0.05).The expressions of apoptosis proteins caspase-12,caspase-9,caspase-3,cleaved caspase-3,Bax and autophagy proteins LC3 and Beclin-1 significantly increased(P＜0.05),which were verified by ICC test.Conclusions SSD can cooperate with TMZ to inhibit the prolifera-tion of GBM cells and induce apoptosis and autophagy,and enhance the sensitivity of GBM cells to TMZ by ac-tivating endoplasmic reticulum stress pathway.

Aim To investigate the expression level of CBX4 in esophageal squamous cell carcinoma(ESCC)and the effect of CBX4 on ESCC proliferation and un-derlying molecular mechanisms.Methods The ex-pression of CBX4 in different cancers was analyzed in Pan-cancers.The expression level of CBX4 in ESCC was analyzed by t-test based on Gene Expression Omni-bus(GEO)data.The viability of CBX4-overex-pressed/knockdown ESCC cells was detected by MTT assay,colony formation assay and flow cytometry assay.Furthermore,the tumor volumn,tumor weight and Ki67 expression were measured by mouse xenograft assay and immunohistochemistry.The mRNA and protein ex-pression levels of apoptosis-related genes PARP、Bcl-2、Bax were determined by qRT-PCR and Western blot,respectively.In addition,the underlying molecular mechanism of CBX4 in ESCC was revealed by qRT-PCR and Western blot.Results CBX4 was upregulat-ed in various cancers.The expression level of CBX4 in ESCC was higher than that in normal tissues(P＜0.05)based on Gene Expression Omnibus(GEO)da-ta.Compared with the normal group,the proliferation of CBX4 knockdown ESCC cells was significantly in-hibited and the apoptosis was promoted(P＜0.05).Meanwhile,the mRNA and protein expression levels of cleaved PARP and Bax were upregulated while that of Bcl-2 was downregulated.In CBX4 overexpression group,tumor volume in vivo increased(P＜0.05).Immunohistochemical results also showed an increase in Ki67 expression.Furthermore,the results of RNA-seq,bioinformatics analysis and qRT-PCR experiments indicated that CBX4 probably regulated the prolifera-tion and apoptosis of ESCC through p38 MAPK signa-ling pathway.Conclusion CBX4 is highly expressed in ESCC and plays as an oncogene role,which might regulate cell proliferation through the p38 MAPK signa-ling pathway.

Objective:To better evaluate the prevalence and epidemiological characteristics of metabolic-associated fatty liver disease (MAFLD) based on liver transient elastography.Methods:It was a cross-sectional study. A total of 6 961 patients without hepatitis, who underwent liver transient elastography examination at the Department of Hepatology, Zhongnan Hospital of Wuhan University from November, 2021 to April, 2022 were included. The patients were categorized into normal and mild, moderate, severe MAFLD groups according to FibroTouch controlled attenuation parameters (CAP). The CAP values among different body mass index (BMI) groups were compared using analysis of variance. The distribution characteristics and the incidence of MAFLD in different age, gender, body mass index(BMI), blood glucose and lipids groups using the chi-square test.Results:The total detection rate of MAFLD and severe MAFLD in the population with a BMI≥28.0 kg/m 2 was 99.6% and 71.8%, respectively. The detection rate of MAFLD in people with normal BMI was 28.4%. The detection rate of MAFLD in women of childbearing age or in the perimenopausal period were both significantly lower than that in men of the same age (40.3% vs 54.9%, χ 2=20.78, P<0.001; 43.1% vs 58.4%, χ 2=27.43, P<0.001), but there was no statistically significant difference in MAFLD detection rate between postmenopausal women and men of the same age. The detection rates of MAFLD in the group with abnormal blood glucose and lipids were both significantly higher than those in the group with normal blood glucose and lipids [69.7%(196/281) vs 35.2%(2 354/6 680), χ 2=138.36, P<0.001; 54.3%(1 696/3 124) vs 37.1%(1 420/3 837), χ 2=207.99, P<0.001]. Conclusion:Non-hepatitis patients had a higher prevalence of MAFLD. The BMI, gender, age, blood glucose, and lipids levels are all strongly associated with MAFLD.

From October 2021 to February 2023, we retrospectively analyzed the clinical and laboratory data of six patients (three male and three female, median age: 54 years, age range: 29-73 years) with mast cell leukemia (MCL) diagnosed in the First Affiliated Hospital of Soochow University (The Mastocytosis Collaborative Network of China). All patients had acute MCL, with at least one C-finding present. The main clinical presentations were hypoalbuminemia ( n=4), fatigue ( n=3), fever ( n=2), abdominal discomfort ( n=2), osteolytic lesions ( n=2), dizziness ( n=1), skin flushing ( n=1), and weight loss ( n=1). Splenomegaly and lymphadenopathy were noted in six and three patients, respectively. Six patients were strongly positive for CD117, five were positive for CD30 and CD25, and four were positive for CD2. Four patients had a normal karyotype and two patients had an abnormal karyotype. Gene mutations were detected in 4/6 cases. The median serum tryptase level was 24.9 (range: 20.1-171.9) μg/L. Two patients were treated with venetoclax and azacitidine for induction (one patient achieved partial remission by combination with afatinib, while there was no remission after combination with dasatinib in the other patient). Two patients did not achieve complete remission despite treatment with cladribine and imatinib, respectively. One patient treated with interferon combined with glucocorticoids was lost to follow-up, and one patient abandoned treatment. The follow-up time ranged from 1.1 to 21.7 months. Three patients died and two survived. Overall, MCL is a rare subtype of systemic mastocytosis with heterogeneous clinical course, and these patients have poor outcome. A better understanding of the clinical characteristics, treatment, and prognosis of MCL is urgently needed.

Objective:To explore the mechanism of herb-partitioned moxibustion in relieving rat intestinal inflammation by focusing on the neutrophil extracellular traps(NETs)in Crohn disease(CD)development. Methods:Rats were randomly divided into a normal group,a model group,a herb-partitioned moxibustion group,and a mesalazine group.The CD rat model was prepared with 2,4,6-trinitrobenzene sulfonic acid except for rats in the normal group.Rats in the normal group and model group did not receive any treatment but had the same fixation as the other groups.Rats in the herb-partitioned moxibustion group received herb-partitioned moxibustion at Qihai(CV6)and bilateral Tianshu(ST25).Rats in the mesalazine group received intragastric administration of mesalazine enteric-coated tablets.The general situation of rats in each group was recorded,and the histopathological changes in the colon were observed and scored by hematoxylin-eosin staining.The serum concentrations of NETs DNA(NETs-DNA),neutrophil elastase(NE)-DNA,and myeloperoxidase(MPO)-DNA were detected by ABC enzyme-linked immunosorbent assay,and the citrullinated histone 3(citH3),MPO,and NE protein and mRNA expression levels in rat colon tissue were observed by immunofluorescence and real-time quantitative polymerase chain reaction. Results:Compared with the normal group,the mucosal ulcer reached the muscularis,the epithelium was incomplete,the goblet cells decreased obviously with significant inflammatory cell infiltration in the colon;the colonic mucosa damage index(CMDI)score increased significantly(P<0.01);the serum NETs-DNA,NE-DNA,and MPO-DNA concentrations increased(P<0.05);the NE,citH3,and MPO protein and mRNA expression in the colonic tissue increased significantly in the model group(P<0.01 or P<0.05).Compared with the model group,the mucosal epithelium in the herb-partitioned moxibustion group and the mesalazine group was repaired and the goblet cells increased with a few infiltrating inflammatory cells in the colon;the CMDI score decreased(P<0.01);the serum NETs-DNA,NE-DNA,and MPO-DNA concentrations decreased(P<0.05);the NE,citH3,and MPO protein and mRNA expression in the colonic tissue was down-regulated(P<0.01 or P<0.05). Conclusion:Herb-partitioned moxibustion reduced the serum NETs complex and inhibited the protein and mRNA expression of NETs complex in the colon tissue,which may be one mechanism of herb-partitioned moxibustion in relieving colon mucosal inflammation in CD.