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.

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:Intervertebral disk degeneration is a pathological change caused by a series of complex molecular mechanisms that result in the aging and damage of intervertebral discs,ultimately leading to severe clinical symptoms.Traditional Chinese medicine has unique advantages in the treatment of intervertebral disk degeneration due to its low cost,non-addictive nature,multi-target effects,minimally toxic and side effects,and high patient acceptance. OBJECTIVE:To review the latest research results of traditional Chinese medicine monomer intervention-related signaling pathways in the treatment of intervertebral disk degeneration,describe and analyze the action mechanism of traditional Chinese medicine monomer on intervertebral disk degeneration,and provide a new approach and theoretical basis for future basic research and clinical treatment. METHODS:The first author searched for relevant literature from January 2018 to February 2023 in CNKI,PubMed,VIP,and WanFang using the search terms"intervertebral disc,signal pathway".The articles that did not meet the criteria were excluded after preliminary screening of the titles and abstracts.Finally,72 articles were selected for review and analysis. RESULTS AND CONCLUSION:Traditional Chinese medicine monomers can regulate multiple classical signaling pathways such as Wnt/β-catenin,PI3K/Akt,mTOR,NF-κB,and MAPK.They achieve this by regulating oxidative stress,adjusting the expression of pro/anti-apoptotic proteins in cells,stimulating cellular autophagy function,reducing stimulation of cell inflammatory factors,increasing the expression of extracellular matrix markers,reducing the production of matrix-degrading enzymes,maintaining the synthesis and stability of extracellular matrix,inducing differentiation of mesenchymal stem cells in the nucleus pulposus into nucleus pulposus cells,promoting endogenous repair and reconstruction,controlling apoptosis and aging of nucleus pulposus cells,and increasing the activity of nucleus pulposus cells.These actions improve the microenvironment within the intervertebral disc,maintain the normal physiological function of the intervertebral disc,and delay intervertebral disc degeneration.

BACKGROUND:At present,bone marrow mesenchymal stem cells are the main seed cells used in cell therapy for the treatment of intervertebral disc degeneration.However,the use of bone marrow mesenchymal stem cells as seed cells for the regeneration of fibrous rings is at risk of heterotopic ossification and teratoma at the repair site.Therefore,it is of great economic and social significance to find a new kind of seed cells for tissue engineering of annulus fibrosus for the treatment of intervertebral disc degeneration. OBJECTIVE:To isolate and purify rat annulus fibrosus-derived stem cells by adherent method combined with fibronectin differential adhesion screening method,and to observe its purification effect and biological characteristics. METHODS:Annulus fibrosus tissue was obtained from a SD rat intervertebral disc.Primary annulus fibrosus-derived stem cells were obtained by the mechanical-enzymatic digestion method.Annulus fibronectin differential adhesion method was used to purify annulus fibrosus-derived stem cells.Morphological changes and proliferation of cells were observed through a microscope.The expression levels of stem cell markers were detected by immunofluorescence technique and qPCR.The screened cells were subjected to multi-lineage cell differentiation and characteristic gene detection. RESULTS AND CONCLUSION:(1)The purified cells grew well,and most of them were angular and star-shaped multi-process cells,which had good proliferation ability.(2)Cells were positive for cell membrane surface antigens CD73,CD90 and CD105,while negative for CD45 and CD34.(3)After specific induction,cells could successfully differentiate into osteoblasts,chondroblasts and lipoblasts.(4)Collagen-I,Runx-2 after osteogenic induction,Collagen Ⅱ,Sox-9 after chondrogenic induction,and PPAR-γ and LPL after lipogenic induction were highly expressed in cells,and the difference was significant compared with that before induction(P<0.05).(5)These findings confirm that the adherent method combined with fibronectin differential adhesion method is effective enough to screen,isolate and purify rat annulus fibrosus-derived stem cells,and has good cell biological properties,good proliferation ability and multiple differentiation potential.

Objective To investigate the effects of total flavones from Oxytropis falcata Bunge on hepatic fibrosis(HF)induced by carbon tetrachloride and liver transforming growth factor(TGF-β)/Smad signaling pathway.Methods Forty-eight male rats were randomly divided into normal group(intraperitoneal injection of peanut oil,intragastric administration of 0.9％NaCl),model group(intraperitoneal injection of 40％CC14 peanut oil solution induced HF model,intragastric administration of 0.9％NaCl),positive control group(modeling,intragastric administration of 0.2 mg·kg-1 of colchicine),experimental-L,-M,-H groups(modeling,intragastric administration of 100,200 and 400 mg·kg-1 of total flavonoid extract of Oxytropis falcata Bunge),8 individuals in each group,for 4 consecutive weeks.The histopathological changes were observed by hematoxylin-eosin and Masson staining.Serum liver function and liver fibrosis were measured;erum inflammatory factors were detected;fluorescence quantitative polymerase chain reaction(RT-qPCR)was used to determine gene expression in liver.Results The pathological injury of liver tissue in the model group was serious,and a large number of inflammatory factors and collagen fibers were accumulated,while the rest of the treatment groups had different degrees of remission.In normal group,model group,positive control group,experimental-L,-M,-H groups,glutamic-pyruvic transaminase levels were(49.28±12.44),(5 885.42±948.37),(4 454.60±489.27),(4 650.47±843.53),(3 761.75±887.30)and(3 544.90±1 066.75)μg·L-1;glutamic-oxaloacetic transaminase levels were(186.90±46.89),(5 936.23±793.81),(3 971.37±780.28),(4 360.30±863.35),(3 943.10±439.47)and(3 971.38±631.08)μg·L-1;hyaluronic acid levels were(45.08±17.16),(104.32±36.06),(66.83±20.09),(70.30±21.07),(60.00±9.68)and(59.02±10.73)μg·L-1;laminin levels were(23.13±3.89),(60.85±13.66),(35.67±9.92),(39.98±9.39),(36.55±12.21)and(34.68±24.83)μg·L-1;type Ⅲ procollagen level were(24.98±5.34),(82.58±30.14),(40.70±16.14),(51.08±23.21),(43.60±12.48)and(44.20±11.66)p±g·L-1;interleukin(IL)-1β levels were(37.63±1.24),(46.10±3.23),(39.22±2.36),(41.33±0.93),(40.25±2.04)and(39.18±2.23)pg·mL-1;tumor necrosis factor-α levels were(314.58±20.56),(383.71±16.97),(349.00±7.93),(348.88±25.11),(325.75±27.84)and(335.07±21.33)pg·mL-1;TGF-β1 mRNA expression of relative quantity respectively were 1.00±0.00,60.99±15.70,9.61±1.59,7.37±1.09,6.41±0.64,6.87±1.09;Smad7 mRNA relative expression were 1.00±0.00,0.34±0.05,0.21±0.03,0.35±0.02,0.38±0.02,0.42±0.03.The above indexes in the model group were compared with the normal group,and the above indexes in the experimental-M,-H groups were compared with the model group,and the differences were statistically significant(P＜0.05,P＜0.01,P＜0.001).Conclusion Total flavonoids of Oxytropis falcata Bunge have protective effects on CC14-induced liver fibrosis in rats,and the mechanism may be related to the regulation of TGF-β/Smad pathway.

Objective To observe the intervention effect of hypericin(HYP)on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)-induced Parkinson's disease(PD)mice model and its mechanism.Methods Thirty C57BL/6 mice were randomly divided into normal,model and experimental groups with 10 mice per group.PD mouse model was established after 7 days of intraperitoneal injection of MPTP,and drug intervention was carried out from the first day of modeling.Normal group and model group were intraperitoneally injected with 500 μL·kg·d-1 0.9％NaCl.The experimental group was intraperitoneally injected with 25 mg·kg·d-1 HYP.The three groups of rats were given the drug once each time for 14 days.The expression levels of tyrosine hydroxylase(TH),Nod-like receptor thermal protein domain protein 3(NLRP3)and ionized calcium binding adapter molecule 1(Iba1)in the striatum of nigra were detected by Western blot.Results The climbing time of normal,model and experimental groups was(5.35±0.43),(9.71±1.19)and(8.07±0.34)s;suspension scores were(2.92±0.15),(1.38±0.28)and(1.96±0.28)points;the relative expression levels of TH protein were 1.04±0.06,0.51±0.09 and 0.75±0.07;the relative expression levels of NLRP3 protein were 0.51±0.03,1.00±0.04 and 0.77±0.06;the relative expression levels of Iba1 protein were 0.68±0.10,1.30±0.28 and 0.89±0.05,respectively.The above indexes in the model group were statistically significant compared with the experimental group and the normal group(all P＜0.01).Conclusion HYP plays a therapeutic role in PD by inhibiting the expression of NLRP3 inflammasome in PD mice.

ObjectiveThe human angiotensin converting enzyme2 （hACE2） transgenic mouse model was used to clarify the antiviral efficacy of BD-77 against a novel coronavirus SARS-CoV-2 and explore the action mechanism of BD-77 against SARS-CoV-2. MethodSARS-CoV-2 Omicron and Delta variant strains-infected VeroE6 cell models were established and administered with BD-77 to observe the antiviral effect of BD-77 in vitro. A kit was used to detect the effect of BD-77 in vitro on the binding of spike S protein of SARS-CoV-2 virus （Delta/Omicron） to angiotensin converting enzyme2 （ACE2）. Chromatography was adopted to detect the binding of BD-77 to the S protein and N protein of the novel coronavirus. hACE2 transgenic C57BL/6 mice were divided into a blank control group， SARS-CoV-2 infection group， BD-77 administration groups of 37.5 mg·kg^-1 and 75 mg·kg^-1， with eight mice in each group. The pneumonia model of SARS-CoV-2-infected hACE2 transgenic mice was built to observe the survival of the mice， detect the virus titer of the lung tissue of the mice， and observe the lesions in the lung tissue. ResultBD-77 had a certain inhibitory effect on Omicron and Delta variant strains in vitro， with median inhibitory concentration （IC₅₀） of 526.3 mg·L^-1 and 653.0 mg·L^-1， respectively. BD-77 had no significant inhibitory effect on the binding of the S protein of WT， Omicron， and Delta variant strains of SARS-CoV-2 to ACE2 and had no binding effect with the S protein and N protein of the novel coronavirus. No mice in the blank group died， while the mortality rate of SARS-CoV-2-infected mice was 75%. There was a large amount of virus replication in the lung tissue of the mice and large areas of inflammatory infiltration in the lung tissue and interstitium. Compared with the model group， BD-77 administration groups of 37.5 mg·kg^-1 and 75 mg·kg^-1 could reduce the mortality of mice， significantly lower the virus titer in the lung tissue of mice （P<0.05）， and improve lung lesions. ConclusionBD-77 demonstrated significant inhibitory effects against SARS-CoV-2 virus in vitro and in vivo. However， its mechanism of action did not involve direct inhibition of the virus itself or intervention in the virus-host binding process. This finding suggests that the mechanism of action of BD-77 needs to be thoroughly investigated and elucidated by further experiments.

Objective To study the mechanism of Sanhuang Decotion in the treatment of ulcerative colitis(UC)under Candida albicans colonization in mice based on Dectin-1-Syk-CARD9 signaling pathway.Methods Mice model of UC with fungal colonization were established with dextran sodium sulfate free drinking and C.albicans intragastric administration.Mice were divided into normal control group,model group,sulfasalazine group,fluconazole group,and Sanhuang Decotion low-and high-dosage groups,and receive corresponding drug interventions.General state of mice were observed,and the disease activity index(DAI)score of mice were calculated.The load of C.albicans in intestine was detected,the length of the colon was measured,and pathological scoring of the colon tissue was performed.The ultrastructural changes of colon epithelium were observed under transmission electron microscopy.The contents of TNF-α,IL-6 and IL-12 in serum and colon tissues were detected by ELISA.The mRNA and protein expression of Dectin-1,Syk,CARD9,NF-κBp65 and inflammation factors in intestinal epithelial cells and colon tissues were detected by qPCR,Western blot and immunohistochemistry.Results Compared with the normal control group,the model group mice showed reduced activity,decreased food intake,accompanied by loose stools,significantly increased DAI score,increased load of C.albicans in the intestine,shortened colon length,and increased histopathological score,with widening of gap between colon epithelial cells,cytoplasmic dissolution,mitochondrial swelling;TNF-α,IL-6 and IL-12 in serum and colon tissue increased,the expressions of Dectin-1 and CARD9 mRNA and protein in colon epithelial cells increased,p-Syk,p-NF-κBp65,CARD9,TNF-α,IL-1β,IL-6 protein expression in colon tissue increased(P<0.01,P<0.05).Compared with the model group,the Sanhuang Decotion high-dosage group mice showed a significant decrease in DAI score,decreased intestinal C.albicans load,increased colon length,decreased histopathological score,more complete and orderly arrangement of microvilli in colon epithelial cells,mild mitochondrial swelling,TNF-α,IL-6 and IL-12 in serum and colon tissue decreased,and the mRNA and protein expression of Dectin-1 and CARD9 in colon tissue increased,the expression of p-Syk,p-NF-κBp65,CARD9,TNF-α,IL-1β,IL-6 protein in colon tissue decreased(P<0.01,P<0.05).Conclusion Sanhuang Decotion may exert an anti C.albicans colonization UC effect by inhibiting the Dectin-1-Syk-CARD9 signaling pathway and reducing the release of inflammatory factors.

Damarane-type triterpene saponins are the main active ingredients in Gynostemma pentaphyllum (Thunb.) Makino. By using D101 macroporous adsorption resin and silica gel open-columns, C18 medium-pressure column chromatography, and preparative high performance liquid chromatography, we isolated four compounds from the leaves of G. pentaphyllum planted in Guangxi Zhuang Autonomous Region. Their structures were determined by comprehensive analyses of MS, NMR data and circular dichroism spectroscopy and identified as (3β,12β)-dihydroxy-25-peroxyhydrohydrodammarane-20,23-diene-3-O-[β-D-glucopyranosyl(1→2)]-β-D-glucopyranoside (1), (3β,12β,24S)-trihydroxydammarane-20,25-diene-3-O-[β-D-glucopyranosyl(1→2)]-β-D-glucopyranoside (2), (3β,20α)-dihydroxy-24-en-12β,22S-epoxydammarane-3-O-[β-D-glucopyranosyl(1→2)]-β-D-glucopyranoside (3), (3β,12β,20S)-trihydroxydammarane-24-en-3-O-[6-O-acetyl-β-D-glucopyranosyl(1→2)-β-D-glucopyranosyl]-20-O-β-D-xylopyranosyl(1→3)-α-L-rhamnopyranosyl (1→6)-β-D-glucopyranoside (4). Compounds 1-4 are new dammarane-type triterpene saponins.