Objective:To investigate the mechanisms of S100 calcium binding protein A10 (S100A10) regulating the proliferation and inflammation of psoriatic keratinocytes by targeting nuclear factor-κB (NF-κB) and signal transduction and activator of transcription 3 (STAT3).Methods:Skin lesion tissues from psoriasis patients (10 cases) and normal skin tissues from healthy control (10 cases) who underwent plastic surgery at the Department of Dermatology and Medical Cosmetology, Honghui Hospital Affiliated to Xi′an Jiaotong University from January to June 2024 were collected. The pCMV plasmid and small interfering RNA plasmid were transfected into human immortalized epidermal HaCaT cells to establish the S100A10 overexpression group and the S100A10 knockdown group, respectively. Untreated HaCaT cells were used as the control group. NF-κB and STAT3 pathway inhibitors were added to the cells in the S100A10 overexpression group to create two subgroups: the S100A10 overexpression+NF-κB inhibitor group and the S100A10 overexpression+STAT3 inhibitor group. The effect of S100A10 on HaCaT cell proliferation was determined by cell counting kit-8 assay. The effect of S100A10 on HaCaT cell apoptosis was detected by extracellular phosphatidylserine binding protein V-fluorescein isothiocyanate/ propidium iodide double staining. The expression of S100A10 in normal skin tissue and skin lesion tissues of psoriasis patients was detected by Western blotting. The relative expression of apoptosis-related protein cysteine aspartic acid specific protease-3 (Caspase-3) and NF-κB and STAT3 pathway-related proteins phosphorylated NF-κB p65 (p-NF-κB p65) and phosphorylated STAT3 (p-STAT3) was detected. The effects of S100A10 on the relative expression of interleukin (IL)-6, tumor necrosis factor-α ( TNF-α), IL-1β, and CXC chemokine ligand 8 ( CXCL8) mRNA were detected by real-time reverse transcription-PCR. The effects of S100A10 on the levels of IL-6, TNF-α, IL-1β and CXCL8 in cell supernatant were detected by enzyme-linked immunosorbent assay. One-way analysis of variance and independent t test were used for data analysis. Results:The expression of S100A10 protein in the skin lesion tissues of psoriasis patients was higher than that in normal skin tissues (1.58±0.13 vs 1.00±0.09, P<0.05). Compared with the control group, the cell survival rates of the S100A10 knockdown group [(99.89±1.03)% vs (82.24±6.03)%], the relative expression of p-NF-κB p65 (0.47±0.06 vs 0.21±0.03) and p-STAT3 protein (0.59±0.12 vs 0.17±0.03), the relative expression of IL-6 (0.98±0.12 vs 0.63±0.07), TNF-α (0.97±0.13 vs 0.71±0.09), IL-1β (1.02±0.14 vs 0.51±0.09) and CXCL8 mRNA (1.01±0.16 vs 0.59±0.11), the levels of IL-6 [(27.69±3.47) pg/ml vs (13.65±2.11) pg/ml], TNF-α [(19.21±2.16) pg/ml vs (10.06±1.44) pg/ml], IL-1β [(15.52±1.03) pg/ml vs (5.17±0.96) pg/ml] and CXCL8 [(62.87±8.48) pg/ml vs (47.11±6.35) pg/ml] in the cell supernatant were lower (all P<0.05), and the cell apoptosis rate [(15.41±2.37)% vs (26.38±4.16)%] and Caspase-3 protein relative expression (0.31±0.04 vs 0.74±0.12) were higher (both P<0.05). The cell survival rates [(145.24±6.03)%], the relative expression of p-NF-κB p65 and p-STAT3 protein (2.37±0.25, 3.98±0.47), the relative expression of IL-6, TNF-α, IL-1β and CXCL8 mRNA (2.27±0.64, 3.31±0.61, 2.74±0.43, 3.11±0.48), the levels of IL-6, TNF-α, IL-1β and CXCL8 in the cell supernatant [(51.17±7.95), (33.65±4.19), (29.95±4.07), (79.97±10.32) pg/ml] in the S100A10 overexpression group were higher than those in the control group (all P<0.05), and the cell apoptosis rate [(5.09±0.73)%] and Caspase-3 protein relative expression (0.09±0.01) were lower than those in the control group (both P<0.05). The cell survival rates [(123.65±9.42)%, (122.94±8.14)%], the relative expression of p-NF-κB p65 (1.51±0.19, 1.49±0.21) and p-STAT3 protein (1.67±0.29, 1.69±0.31), the relative expression of IL-6 (1.69±0.14, 1.73±0.15), TNF-α (1.92±0.27, 1.94±0.25), IL-1β (1.85±0.16, 1.87±0.21) and CXCL8 mRNA (2.02±0.34, 2.01±0.39), the levels of IL-6 [(35.55±5.12), (36.18±5.24) pg/ml], TNF-α [(25.17±3.08), (25.23±3.17) pg/ml], IL-1β [(22.08±3.11), (22.11±3.24) pg/ml] and CXCL8 [(70.04±9.31), (70.11±10.29) pg/ml] in the cell supernatant in the S100A10 overexpression+NF-κB inhibitor group and the S100A10 overexpression+STAT3 inhibitor group were higher than those in S100A10 knockdown group (all P<0.05), and the cell apoptosis rates [(20.13±4.62)%, (20.21±4.91)%] and Caspase-3 protein relative expression (0.15±0.03, 0.16±0.04) were lower than those in S100A10 knockdown group (all P<0.05).The cell survival rates, the relative expression of p-NF-κB p65 and p-STAT3 protein, the relative expression of IL-6, TNF-α, IL-1β and CXCL8 mRNA, the levels of IL-6, TNF-α, IL-1β and CXCL8 in the cell supernatant in the S100A10 overexpression+NF-κB inhibitor group and the S100A10 overexpression+STAT3 inhibitor group were lower than those in S100A10 overexpression group (all P<0.05), and the cell apoptosis rates and Caspase-3 protein relative expression were higher than those in S100A10 overexpression group (all P<0.05). Conclusions:S100A10 may regulate the proliferation and inflammation of keratinocytes in psoriasis by targeting NF-κB and STAT3.

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.

ObjectiveThis paper aims to verify the therapeutic effect of Cranial Painkiller pills' extract powder prepared by the new process on the rat's trigeminal neuralgia model caused by infraorbital injection of Talci Pulvis， evaluate its potential clinical application value， and compare the therapeutic effect with that of Cranial Painkiller granules， so as to provide data support for the application of the Cranial Painkiller pills' extract powder and precise treatment. MethodsThe rat's trigeminal neuralgia model was constructed by infraorbital injection of Talci Pulvis， and the rats were randomly divided into the normal group， model group， carbamazepine group （60 mg·kg^-1）， Cranial Painkiller granules group （2.70 g·kg^-1）， and low， medium， and high dosage groups of Cranial Painkiller pills' extract powder （1.35， 2.70， 5.40 g·kg^-1） according to the basal mechanical pain thresholds， and there were 10 rats in each group. The drug was administered by gavage to each group 2 h after modeling， and distilled water was given by gavage to the normal and model groups under the same conditions once a day for 10 d. Von Frey brushes were used to measure mechanical pain thresholds in rats. Hematoxylin-eosin （HE） staining was used to detect pathological changes in the trigeminal ganglion， and enzyme-linked immunosorbent assay （ELISA） was used to detect the inflammatory factors interleukin-1 （IL-1）， interleukin-6 （IL-6）， interleukin-8 （IL-8）， and tumor necrosis factor-α （TNF-α） levels in rat serum， as well as neuropeptide substance P （SP） and β-endorphin （β-EP） levels in rat brain tissue. Western blot technique was used to detect the levels of NLRP3， ASC， Caspase-1， and OTULIN proteins in rat brain tissue. ResultsCompared with the normal group， the pain threshold of rats in the model group showed a continuous significant decrease （P<0.01）. The pathological damage of brain tissue was significant （P<0.01）， and the inflammatory levels of IL-1， IL-6， IL-8， and TNF-α in serum were significantly elevated （P<0.01）. The level of the SP in the brain tissue was significantly elevated （P<0.01）， and the level of β-EP was significantly reduced （P<0.01）， while the level of OTULIN was significantly reduced， and NLRP3， ASC， and Caspase-1 protein levels were significantly elevated （P<0.01）. After administration of the drug， compared with the model group， the pain threshold of each dose group of the Cranial Painkiller pills' extract powder and the Cranial Painkiller granules group significantly increased （P<0.01）. The inflammatory levels of IL-1， IL-6， IL-8， and TNF-α and SP levels significantly decreased （P<0.01）， and the β-EP levels were significantly elevated （P<0.01）， while the levels of OTULIN protein were significantly elevated （P<0.05， P<0.01）， and the levels of NLRP3， ASC proteins were decreased （P<0.01）in high dose Cranial Painkiller pills' extract powder. Meanwhile， compared with those in the model group， the trigeminal ganglion lesions of rats in the Cranial Painkiller pills' extract powder and Cranial Painkiller granules groups showed different degrees of improvement （P<0.05， P<0.01）. ConclusionThe Cranial Painkiller pills' extract powder has significant therapeutic effects on the rat model of trigeminal neuralgia induced by infraorbital injection of Talci Pulvis， and its mechanism is related to the improvement of OTULIN-regulated neuroinflammation.

ObjectiveTo investigate the protective effect and mechanism of verbenalin on mouse mononuclear macrophage leukemia cells （RAW264.7） damaged by human coronavirus （HCoV）-229E infection， thereby providing experimental evidence for its development and application. MethodsRAW264.7 macrophages were infected with different concentrations of HCoV-229E to establish a coronavirus-induced macrophage injury model using the cell counting kit-8 （CCK-8） assay for assessing cell proliferation and viability. Cells were randomly divided into four groups： normal control， verbenalin group （125 μmol·L^-1）， model group （HCoV-229E）， and HCoV-229E + verbenalin group （HCoV-229E + 125 μmol·L^-1 verbenalin）. Cell viability was measured using the CCK-8 assay， and the maximum non-toxic concentration （CC₀）， half-maximal cytotoxic concentration （CC₅₀）， half-maximal effective concentration （EC₅₀）， and selectivity index （SI） of verbenalin were calculated. Calcein/PI double staining was used to assess cell viability and cytotoxicity， and JC-1 staining was applied to evaluate changes in mitochondrial membrane potential （MMP）. mito-Keima adenovirus labeling was used to assess mitophagy levels in each group. ResultsA macrophage infection model was successfully established by infecting RAW264.7 cells with the original concentration of HCoV-229E for 36 h. The CC₀ of verbenalin was 125 μmol·L^-1. The CC₅₀ was 448.25 μmol·L^-1. The EC₅₀ against HCoV-229E-infected cells was 46.28 μmol·L^-1， and the SI was 9.68. Compared with the normal group， the model group showed significantly reduced cell survival rate （P<0.01）， increased cell death rate （P<0.01）， decreased MMP （P<0.01）， and suppressed mitophagy （P<0.01）. In contrast， verbenalin treatment significantly improved cell survival rate （P<0.01）， reduced cell death rate （P<0.01）， alleviated MMP loss （P<0.01）， and enhanced mitophagy levels （P<0.01） compared with the model group. ConclusionVerbenalin can enhance the survival rate of macrophages following HCoV-229E infection. The underlying mechanism may be associated with the activation of mitophagy， maintenance of MMP stability， and alleviation of mitochondrial damage.

Objective To explore the correlation between the expression level of secreted curl associated protein 5(SFRP5)and the condition and prognosis of patients with severe pneumonia.Methods A total of 168 patients with pneumonia admitted to the Department of Respiratory and Critical Care Medicine of this hospital from June 2022 to June 2024 were selected as the research subjects,and 78 healthy individuals who underwent physical examinations in this hospital during the same period were selected as the control group.According to the acute physiology and chronic health evaluation Ⅱ(APACHEⅡ)score,the patients were di-vided into the common pneumonia group(APACHE Ⅱ score＜15,n=82)and the severe pneumonia group(APACHE Ⅱ score≥ 15,n=86).Peripheral venous blood was collected from the three groups after fasting for 8 to 12 hours and peripheral blood mononuclear cells were extracted by centrifuging.Real-time fluores-cence quantitative PCR was used to detect the expression level of SFRP5 mRNA in peripheral blood mononu-clear cells,which was Used to determine the expression level of SFRP5.Pearson analysis was used to analyze the correlations between the expression level of SFRP5 and the levels of inflammatory factors,pulmonary function indices and the condition in patients with severe pneumonia.The receiver operating characteristic(ROC)curve was used to analyze the diagnostic efficiency of the expression level of SFRP5 for the prognosis of patients with severe pneumonia.Results There were also significant differences in the levels of WBC,C re-active protein(CRP),procalcitonin(PCT),interleukin(IL)-6,and IL-8 among the three groups(P＜0.05),with the highest levels observed in the severe pneumonia group and the lowest in the control group.There were significant differences in the levels of forced expiratory volume in the first second(FEV1),forced vital capacity(FVC),and FEV1/FVC among the three groups(P＜0.05),with the highest levels observed in the severe pneumonia group and the lowest in the control group.The expression level of SFRP5 in the severe pneumonia group was the lowest(0.46±0.13),followed by the common pneumonia group(0.78±0.15),and the control group was the highest level(1.23±0.22),with statistically significant differences(P＜0.05).The results of Pearson correlation analysis showed that the expression level of SFRP5 in patients with severe pneu-monia was negatively correlated with the levels of WBC,CRP,PCT,IL-6,IL-8,and the APACHE Ⅱ score(P＜0.05),and positively correlated with the levels of FEV1,FVC,and FEV1/FVC(P＜0.05).The area un-der the curve(AUC)of the expression level of SFRP5 for predicting poor prognosis in patients with severe pneumonia was 0.843(95％CI:0.748-0.912),with a sensitivity of 68.18％and a specificity of 89.06％.Conclusion The expression level of SFRP5 is related to the condition of patients with severe pneumonia,and SFRP5 can predict the prognosis of patients with severe pneumonia.

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.

Objective To investigate the expression of small nuclear ribonucleoprotein D1 (SNRPD1) in the gastric cancer tissue and evaluate the predictive value of SNRPD1 expression level for the long-term prognosis of gastric cancer patients and the possible functioning mechanism of SNRPD1. Methods The UALCAN and Gene Expression Profiling Interactive Analysis (GEPIA) were employed to analyze the expression level of SNRPD1 in pan-cancer and its relationship with the prognosis of gastric cancer.The clinical data of 109 patients who underwent radical surgery for gastric cancer from January 2014 to January 2017 in the First Affiliated Hospital of Bengbu Medical University were retrospectively analyzed.Gastric cancer and paracancerous tissue samples were collected,and the expression of SNRPD1 was detected by immunohistochemical staining.Lentiviral transfection was employed to construct the BGC-823 gastric cancer cell models with stable high and low expression of SNRPD1,respectively.The CCK-8 assay and colony formation assay were employed to measure the proliferation of gastric cancer cells,and flow cytometry was used to analyze the cell cycle.Western blotting was employed to determine the expression levels of proteins in the signaling pathway. Results The data from UALCAN and GEPIA showed that SNRPD1 was highly expressed in the tissue of malignant tumors including gastric cancer (P<0.001).The expression level of SNRPD1 in the gastric cancer tissue was higher than that in the paracancerous tissue (P<0.001).Moreover,the expression level of SNRPD1 was positively correlated with the levels of carcinoembryonic antigen (P<0.001),carbohydrate antigen 19-9 (P<0.001),G stage (P=0.042),T stage (P=0.002),and N stage (P=0.027) in the patients with gastric cancer.The high expression of SNRPD1 had a predictive value for the long-term prognosis of gastric cancer (P<0.001),and it was an independent risk factor for the death of gastric cancer patients (P=0.003).The results of gene ontology and kyoto encyclopedia of genes and Genomes enrichment analyses showed that SNRPD1 was involved in the regulation of the cell cycle.The results of CCK-8 and colony formation assays showed that up-regulation of SNRPD1 promoted the proliferation of gastric cancer cells (P<0.001,P<0.001).The up-regulation of SNRPD1 up-regulated the expression of cyclin-dependent kinase 6 and G₁/S-specific cyclin-D1 (P<0.001,P=0.002),whereas the interference in SNRPD1 led to opposite results (P=0.004,P<0.001).SNRPD1 accelerated the G₁/S phase transition of gastric cancer cells (P<0.001).The overexpression of SNRPD1 promoted the expression of phosphorylated phosphatidylinositol 3-kinase (PI3K) and phosphorylated protein kinase B (Akt) in gastric cancer cells (P=0.043,P<0.001),whereas disruption of SNRPD1 inhibited their expression (both P<0.001).Insulin-like growth factor 1,an agonist of the PI3K/Akt signaling pathway,promoted the proliferation of gastric cancer cells with SNRPD1 disturbed (P=0.002). Conclusion High expression of SNRPD1 in the gastric cancer tissues is associated with poor prognosis,and it may promote tumor cell proliferation and regulate the cell cycle by activating the PI3K/Akt signaling pathway.
Humans ; Stomach Neoplasms/pathology* ; Prognosis ; Cell Line, Tumor ; Cell Proliferation ; Retrospective Studies ; Cell Cycle ; Male ; Female

Objective To investigate the role and mechanism of afzelin(AFZ)in treating Crohn's disease-like colitis.Methods A mouse model of 2,4,6-trinitrobenzene sulfonic acid-induced colitis was established to assess the effect of AFZ on experimental colitis in vivo.A Caco-2 cell model of tumor necrosis factor(TNF)-α-induced inflammation was established to evaluate the effects of AFZ on the intestinal barrier function,intestinal epithelial cell apoptosis,and mitochondrial function in vitro.The animal and cell experiments were performed to validate the regulatory role of the adenosine monophosphate-activated protein kinase(AMPK)/silent information regulater 1(SIRT1)/peroxisome proliferator-activated receptor gamma coactivator(PGC)-1α pathway in the treatment of colitis with AFZ.Results AFZ reduced the disease activity index(P=0.003),weight loss(P<0.001),colon shortening(P<0.001),inflammation score(P=0.002),pro-inflammatory cytokine release(interleukin-6:P<0.001;TNF-α:P=0.010),and intestinal barrier permeability(fluorescein isothiocyanate dextran 4:P<0.001;intestinal-type fatty acid-binding protein:P=0.013).Meanwhile,AFZ increased the colonic transepithelial electric resistance(P=0.001),reduced bacterial translocation(P<0.001),and promoted the localization and up-regulated the expression of tight junction proteins [zonula occluden-1(P=0.005) and Claudin-1(P=0.024)].AFZ exerted a protective effect on the Caco-2 cells exposed to TNF-α in terms of intestinal epithelial cell permeability(P=0.017),transepithelial electric resistance(P=0.014),and tight junction protein[zonula occluden-1(P=0.014) and Claudin-1(P=0.006)] localization and expression.Furthermore,the cell and animal experiments confirmed that AFZ reduced the percentage of apoptosis(P<0.001,P=0.013)and the expression of cleaved-caspase 3(P=0.028,P=0.004)and Bax(P=0.004,P=0.020),and upregulated the Bcl2(P=0.020,P=0.006)level in intestinal epithelial cells.Additionally,AFZ increased the number of mitochondria,mitochondrial membrane potential,and copy number of mitochondrial DNA(P=0.007)in intestinal epithelial cells,while enhancing the activities of mitochondrial respiratory chain complex Ⅰ(P=0.005)and complex Ⅳ(P=0.001).The activation of the AMPK/SIRT1/PGC-1α pathway was involved in the protective effects of AFZ on mitochondrial function and apoptosis in intestinal epithelial cells.Conclusion AFZ alleviates mitochondrial dysfunction and apoptosis in intestinal epithelial cells by activating the AMPK/SIRT1/PGC-1α pathway,thereby ameliorating intestinal barrier dysfunction and experimental colitis.
Animals ; Colitis/drug therapy* ; Humans ; Caco-2 Cells ; Mice ; Trinitrobenzenesulfonic Acid ; Apoptosis/drug effects* ; Disease Models, Animal ; AMP-Activated Protein Kinases/metabolism* ; Sirtuin 1/metabolism*

Background: Intradermal microdroplet injections of botulinum toxin type-A (BoNT/A) effectively ameliorate rosacea-related angiogenesis, but the mechanism remains unclear. Objective: To explore the anti-angiogenesis of BoNT/A in the rosacea-like mouse model and to measure the secretion of vascular endothelial growth factor (VEGF) by mast cells. Methods: A rosacea-like mouse model was induced by LL37 in both Mas-related G-proteincoupled receptor B2 conditional knockout (MrgprB2 −/− ) mice and wild-type (WT) mice, then treated with BoNT/A and/or Apatinib. The abundance of endothelial cells and mast cells in mouse skin was determined using dual immunofluorescence staining. The VEGF levels in supernatants and cell lysates of laboratory of allergic disease 2 (LAD2) mast cells were assessed using reverse transcription quantitative polymerase chain reaction, western blots, and enzyme-linked immunosorbent assay. The effect of conditioned medium (CM) collected from LAD2 on human umbilical vein endothelial cells (HUVECs) was determined using tube formation assays. The number of proliferative cells was confirmed using the 5-ethynyl-2’-deoxyuridine incorporation assays.The effect of BoNT/A on the internalization of Mas-related G-protein-coupled receptor X2 (MRGPRX2) was detected using flow cytometry and immunofluorescence staining. Results: LL37-induced rosacea-like skin manifestations were significantly alleviated in MrgprB2 −/− mice compared to WT controls. BoNT/A mitigated the LL37-induced secretion of VEGF by LAD2. The CM from BoNT/A-treated LAD2 inhibited HUVEC proliferation and tube formation. The LAD2 cells co-treated with LL37 and BoNT/A exhibited dramatically enhanced MRGPRX2 internalization. Conclusion BoNT/A enhances LL37-mediated MRGPRX2 internalization in mast cells, thereby reducing VEGF secretion and neovascularization and improving facial flushing symptom in rosacea.