BACKGROUND:Studies have shown that metformin has anti-inflammatory,anti-tumor,anti-aging and vasoprotective effects,and can inhibit the progression of osteoarthritis,but its specific mechanism of action remains unclear. OBJECTIVE:To investigate the mechanism of metformin on cartilage protection in a rat model of osteoarthritis. METHODS:Forty male Sprague-Dawley rats were randomly divided into four groups(n=10 per group):blank,control,sham-operated,and metformin groups.The blank group did not undergo any surgery.In the sham-operated group,the joint cavity was exposed.In the model group and the metformin group,the modified Hulth method was used to establish the osteoarthritis model.At 1 day after modeling,the rats in the metformin group were given 200 mg/kg/d metformin by gavage,and the model,blank,and sham-operated groups were given normal saline by gavage.Administration in each group was given for 4 weeks consecutively.Hematoxylin-eosin staining,toluidine blue staining,and safranin O-fast green staining were used to observe the morphological structure of rat knee joints.Immunohistochemical staining and western blot were used to detect the protein expression of SOX9,type Ⅱ collagen,a disintegrin and metalloproteinase with thrombospondin motifs 5(ADAMTS5),Beclin1,P62,phosphatidylinositol 3-kinase(PI3K),p-PI3K,protein kinase B(AKT),p-AKT,mammalian target of rapamycin(Mtor),and p-Mtor in rat cartilage tissue. RESULTS AND CONCLUSION:The results of hematoxylin-eosin,toluidine blue and safranin O-fast green staining showed smooth cartilage surface of the knee joints and normal histomorphology in the blank group and the sham-operated group,while in the model group,there was irregular cartilage surface of the knee joint and cartilage damage,with a decrease in the number of chondrocytes and the content of proteoglycans in the cartilage matrix.In the metformin group,there was a significant improvement in the damage to the structure of the cartilage in the knee joints of the rats,and the cartilage surface tended to be smooth,with an increase in the number of chondrocytes and the content of proteoglycans in the cartilage matrix.Immunohistochemistry staining and western blot results showed that compared with the control and sham-operated groups,the expression of SOX9,type Ⅱ collagen,and Beclin1 proteins in the cartilage tissue of rats in the model group was significantly decreased(P<0.05).Conversely,the expression of ADAMTS5,P62,as well as p-PI3K,p-AKT,and p-Mtor proteins was significantly increased(P<0.05).Furthermore,compared with the model group,the expression of SOX9,type Ⅱ collagen,and Beclin1 proteins in the cartilage tissue of rats in the metformin group was significantly increased(P<0.05),while the expression of ADAMTS5,P62,as well as p-PI3K,p-AKT,and p-Mtor proteins was significantly decreased(P<0.05).To conclude,Metformin can improve the autophagy activity of chondrocytes and reduce the degradation of cartilage matrix in osteoarthritis rats by inhibiting the activation of PI3K/AKT/Mtor signaling pathway,thus exerting a protective effect on articular cartilage.

BACKGROUND:Metformin is currently considered the first-line medication for the treatment of type 2 diabetes.Metformin may delay the progression of osteoarthritis,but its specific mechanism of action remains unclear.OBJECTIVE:To evaluate the therapeutic effects and the related action mechanisms of metformin on osteoarthritis in rats.METHODS:(1)Network pharmacology:Potential common targets for metformin,osteoarthritis,and ferroptosis were screened using the CTD,SwissTargetPrediction,GeneCards,and OMIM databases.After importing the targets into the STRING database,protein-protein interaction analysis was conducted to identify the key targets for metformin,osteoarthritis,and ferroptosis.(2)Molecular docking:P53 and its downstream factor SLC7A11 protein structures in PDB format were downloaded from the PDB database.The 2D structure of metformin was converted to a 3D structure,and molecular docking of metformin with the proteins was performed using Discovery Studio 2019 Client.(3)In vivo experiments:Thirty male SD rats were randomly divided into three groups(n=10).The blank group did not receive surgery.The osteoarthritis model was established using the modified Hulth method for the model and metformin groups.One day after the surgery,rats in the metformin group were gavaged with metformin 200 mg/kg per day,while the blank and model groups were gavaged with physiological saline.Treatment continued for 4 weeks.Hematoxylin-eosin staining and Safranin O-fast green staining were used to observe the pathological morphology and structure of the knee cartilage,and Mankin scoring was performed.ELISA was used to measure the levels of tumor necrosis factor-α and interleukin-6 in the serum.The microplate method was used to measure serum ferroptosis-related indicators,including glutathione,malondialdehyde,and Fe2+.Immunofluorescence staining,western blot assay,and real-time qPCR were used to detect the protein and mRNA expression of P53,SLC7A11,glutathione peroxidase 4,proteoglycans,and matrix metalloproteinase 13 in the cartilage tissue of the rats.RESULTS AND CONCLUSION:(1)A total of 96 intersecting targets among metformin,osteoarthritis,and ferroptosis were identified.After protein-protein interaction analysis,77 potential targets were found.Further screening identified the core targets as TP53,AKT1,JUN,interleukin-6,MYC,interleukin-1β,and tumor necrosis factor-α,among others.(2)Docking analysis results showed that metformin bound strongly and stably with P53 and its downstream factor SLC7A11.(3)In the model group,the knee cartilage surface was irregular,with cartilage tissue defects and reduced chondrocyte numbers.Compared to the model group,the knee cartilage structure damage in the metformin group was significantly improved,with a smoother cartilage surface and increased chondrocyte numbers.The Mankin score in the model group was significantly higher than that in the blank group,while the Mankin score in the intervention group was significantly lower than that in the model group.(4)Compared with the model group,the metformin group had significantly lower levels of tumor necrosis factor-α,interleukin-6,malondialdehyde,and Fe2+,and significantly higher glutathione levels.(5)Compared to the model group,the metformin group had significantly increased protein and mRNA expression of SLC7A11,glutathione peroxidase 4,and proteoglycans,and significantly decreased protein and mRNA expression of P53 and matrix metalloproteinase 13 in their cartilage tissue.(6)The results indicate that metformin can effectively improve cartilage damage in osteoarthritis rats and alleviate chondrocyte ferroptosis by inhibiting the aberrantly activated P53/SLC7A11/glutathione peroxidase 4 signaling pathway.This improvement in chondrocyte iron metabolism and lipid peroxidation response further reduces cartilage matrix degradation and prevents further cartilage damage and inflammatory response.

AIM:To investigate the therapeutic effect of recombinant human interleukin-37(rhIL-37)inter-vention on extracellular matrix deposition in human bronchial epithelial cells exposed to fine particulate matter(PM2.5),and to explore the underlying mechanisms.METHODS:An airway injury cell model was established using normal human bronchial epithelial 16HBE cells treated with various concentrations(6.25,12.5,25 and 50 mg/L)of PM2.5.Western blot and immunofluorescence were employed to assess the expression level of Wnt family member 5a(Wnt5a).The con-centration of PM2.5 that maximized Wnt5a expression was identified as the optimal concentration for further analysis.The cells were divided into 3 groups,PM2.5+siRNA,PM2.5+Wnt5a siRNA and control+Wnt5a siRNA,to evaluate the expres-sion of fibronectin,the secretion of IL-6 and IL-1β,and the production of reactive oxygen species(ROS).A subsequent set of experiments was organized into PM2.5+rhIL-37,PM2.5 and control groups to determine whether rhIL-37 down-regu-lates the expression of Wnt5a and fibronectin,and reduces the secretion of IL-6 and IL-1β.RESULTS:(1)After 24 h of exposure to various concentrations of PM2.5,the expression of Wnt5a in 16HBE cells was significantly elevated,except at 50 mg/L,compared with control group(P＜0.05).The highest Wnt5a expression was observed at 12.5 mg/L PM2.5(P＜0.01).(2)Fibronectin expression peaked after 24 h of exposure to PM2.5 at 12.5 mg/L(P＜0.01).(3)Successful knock-down of Wnt5a resulted in decreased levels of fibronectin and nuclear factor E2-related factor(Nrf2).Additionally,the production of ROS and the secretion of IL-6 and IL-1β were significantly reduced compared with control group(P＜0.01).(4)The administration of rhIL-37 before PM2.5 exposure led to decreased levels of Wnt5a,fibronectin and Nrf2 compared with PM2.5 group(P＜0.05).Furthermore,rhIL-37 treatment reduced the ROS production and the secretion of IL-6 and IL-1β(P＜0.05).CONCLUSION:In vitro cell models demonstrate that PM2.5 can induce extracellular matrix deposition in bronchial epithelial cells,potentially through the modulation of inflammatory response and oxidative stress influenced by Wnt5a.Additionally,rhIL-37 appears to mitigate inflammatory response and oxidative stress by down-regulating Wnt5a.

BACKGROUND:Metformin is currently considered the first-line medication for the treatment of type 2 diabetes.Metformin may delay the progression of osteoarthritis,but its specific mechanism of action remains unclear.OBJECTIVE:To evaluate the therapeutic effects and the related action mechanisms of metformin on osteoarthritis in rats.METHODS:(1)Network pharmacology:Potential common targets for metformin,osteoarthritis,and ferroptosis were screened using the CTD,SwissTargetPrediction,GeneCards,and OMIM databases.After importing the targets into the STRING database,protein-protein interaction analysis was conducted to identify the key targets for metformin,osteoarthritis,and ferroptosis.(2)Molecular docking:P53 and its downstream factor SLC7A11 protein structures in PDB format were downloaded from the PDB database.The 2D structure of metformin was converted to a 3D structure,and molecular docking of metformin with the proteins was performed using Discovery Studio 2019 Client.(3)In vivo experiments:Thirty male SD rats were randomly divided into three groups(n=10).The blank group did not receive surgery.The osteoarthritis model was established using the modified Hulth method for the model and metformin groups.One day after the surgery,rats in the metformin group were gavaged with metformin 200 mg/kg per day,while the blank and model groups were gavaged with physiological saline.Treatment continued for 4 weeks.Hematoxylin-eosin staining and Safranin O-fast green staining were used to observe the pathological morphology and structure of the knee cartilage,and Mankin scoring was performed.ELISA was used to measure the levels of tumor necrosis factor-α and interleukin-6 in the serum.The microplate method was used to measure serum ferroptosis-related indicators,including glutathione,malondialdehyde,and Fe2+.Immunofluorescence staining,western blot assay,and real-time qPCR were used to detect the protein and mRNA expression of P53,SLC7A11,glutathione peroxidase 4,proteoglycans,and matrix metalloproteinase 13 in the cartilage tissue of the rats.RESULTS AND CONCLUSION:(1)A total of 96 intersecting targets among metformin,osteoarthritis,and ferroptosis were identified.After protein-protein interaction analysis,77 potential targets were found.Further screening identified the core targets as TP53,AKT1,JUN,interleukin-6,MYC,interleukin-1β,and tumor necrosis factor-α,among others.(2)Docking analysis results showed that metformin bound strongly and stably with P53 and its downstream factor SLC7A11.(3)In the model group,the knee cartilage surface was irregular,with cartilage tissue defects and reduced chondrocyte numbers.Compared to the model group,the knee cartilage structure damage in the metformin group was significantly improved,with a smoother cartilage surface and increased chondrocyte numbers.The Mankin score in the model group was significantly higher than that in the blank group,while the Mankin score in the intervention group was significantly lower than that in the model group.(4)Compared with the model group,the metformin group had significantly lower levels of tumor necrosis factor-α,interleukin-6,malondialdehyde,and Fe2+,and significantly higher glutathione levels.(5)Compared to the model group,the metformin group had significantly increased protein and mRNA expression of SLC7A11,glutathione peroxidase 4,and proteoglycans,and significantly decreased protein and mRNA expression of P53 and matrix metalloproteinase 13 in their cartilage tissue.(6)The results indicate that metformin can effectively improve cartilage damage in osteoarthritis rats and alleviate chondrocyte ferroptosis by inhibiting the aberrantly activated P53/SLC7A11/glutathione peroxidase 4 signaling pathway.This improvement in chondrocyte iron metabolism and lipid peroxidation response further reduces cartilage matrix degradation and prevents further cartilage damage and inflammatory response.

AIM:To investigate the therapeutic effect of recombinant human interleukin-37(rhIL-37)inter-vention on extracellular matrix deposition in human bronchial epithelial cells exposed to fine particulate matter(PM2.5),and to explore the underlying mechanisms.METHODS:An airway injury cell model was established using normal human bronchial epithelial 16HBE cells treated with various concentrations(6.25,12.5,25 and 50 mg/L)of PM2.5.Western blot and immunofluorescence were employed to assess the expression level of Wnt family member 5a(Wnt5a).The con-centration of PM2.5 that maximized Wnt5a expression was identified as the optimal concentration for further analysis.The cells were divided into 3 groups,PM2.5+siRNA,PM2.5+Wnt5a siRNA and control+Wnt5a siRNA,to evaluate the expres-sion of fibronectin,the secretion of IL-6 and IL-1β,and the production of reactive oxygen species(ROS).A subsequent set of experiments was organized into PM2.5+rhIL-37,PM2.5 and control groups to determine whether rhIL-37 down-regu-lates the expression of Wnt5a and fibronectin,and reduces the secretion of IL-6 and IL-1β.RESULTS:(1)After 24 h of exposure to various concentrations of PM2.5,the expression of Wnt5a in 16HBE cells was significantly elevated,except at 50 mg/L,compared with control group(P＜0.05).The highest Wnt5a expression was observed at 12.5 mg/L PM2.5(P＜0.01).(2)Fibronectin expression peaked after 24 h of exposure to PM2.5 at 12.5 mg/L(P＜0.01).(3)Successful knock-down of Wnt5a resulted in decreased levels of fibronectin and nuclear factor E2-related factor(Nrf2).Additionally,the production of ROS and the secretion of IL-6 and IL-1β were significantly reduced compared with control group(P＜0.01).(4)The administration of rhIL-37 before PM2.5 exposure led to decreased levels of Wnt5a,fibronectin and Nrf2 compared with PM2.5 group(P＜0.05).Furthermore,rhIL-37 treatment reduced the ROS production and the secretion of IL-6 and IL-1β(P＜0.05).CONCLUSION:In vitro cell models demonstrate that PM2.5 can induce extracellular matrix deposition in bronchial epithelial cells,potentially through the modulation of inflammatory response and oxidative stress influenced by Wnt5a.Additionally,rhIL-37 appears to mitigate inflammatory response and oxidative stress by down-regulating Wnt5a.

BACKGROUND:Previous research by the research team found that domestically produced porous tantalum is beneficial for early adhesion and proliferation of MG63 cells,and can be used as a scaffold material for bone tissue engineering. OBJECTIVE:To investigate the effect of domestic porous tantalum modified by osteogenic induction factor slow-release system on the adhesion,proliferation,and differentiation of MG63 cells. METHODS:Osteogenic induction factor slow-release system was constructed by adding 15%volume fraction of osteogenic factor solution to poly(lactic-co-glycolic-acid)gel.The passage 3 MG63 cells were inoculated on a porous tantalum surface(control group),porous tantalum surface coated with poly(lactic-co-glycolic-acid)copolymer gel(gel group),and porous tantalum surface coated with osteoblastic induction factor slow-release system(slow-release system group),and co-cultured for 5 days.The surface cytoskeleton of the material was observed by phalloidine staining.Cell proliferation was detected by flow cytometry.Western blot assay and RT-qPCR were used to detect the protein and mRNA expressions of type Ⅰ collagen,osteopontin,and RUNX-2 on the surface cells of the material. RESULTS AND CONCLUSION:(1)Phalloidine staining showed that MG63 cells adhered to and grew on the surface and inside of the three groups of porous tantalum,and the matrix secreted by the cells covered the surface of the material.(2)Flow cytometry showed that the cell proliferation in the slow-release system group was faster than that in the control group and the gel group(P<0.05).(3)Western blot assay and RT-qPCR showed that the protein and mRNA expressions of type Ⅰ collagen,osteopontin,and RUNX-2 in the slow-release system group were higher than those in the control group and gel group(P<0.05).(4)The results showed that the domestic porous tantalum modified by the osteogenic induction factor slow-release system was beneficial to the adhesion,proliferation,and differentiation of MG63 osteoblasts.

Objective:The incidence of early-onset colorectal cancer (EOCRC) is increasing globally; however, the molecular characteristics and prognosis of sporadic EOCRC are unclear. In this systematic review and meta-analysis, we aimed to investigate the incidence of gene mutations and their association with cancer survival in sporadic EOCRC, focusing on six common gene mutations ( TP53, BRAF, KRAS, NRAS, PTEN, and APC). Methods:Ovid Embase and Ovid Medline electronic databases were searched for studies involving patients with sporadic EOCRC (i.e., diagnosed with colorectal cancer before the age of 50 years and with no evidence of hereditary syndromes predisposing to colorectal cancer). The included articles were evaluated using quality assessment tools. Meta-analysis was performed using random-effects and fixed-effects models. Cochran's Q statistic and the I2 index were used to assess heterogeneity. The incidence of the six common gene mutations listed above in sporadic EOCRC and their association with cancer survival were evaluated.Results:(1) Incidence of specific gene mutations in sporadic EOCRC. A total of 34 articles were included in this meta-analysis. The incidence of APC gene mutation was 36% (from 13 articles, 95%CI: 19%-55%, P=0.043); of KRAS gene mutation 30% (from 26 articles, 95%CI: 24%-35%, P=0.190); of BRAF gene mutation 7% (from 18 articles, 95%CI: 5%-11%, P=0.422); of NRAS gene mutation 4% (from five articles, 95%CI: 3%-5%, P=0.586); of PTEN gene mutation 6% (from six articles, 95%CI: 4%-10%, P=0.968); and of TP53 gene mutation 59% (from 13 articles, 95%CI: 49%-68%, P=0.164). (2) Association between gene mutations and survival in sporadic EOCRC . A total of six articles were included in this meta-analysis. Compared with wild-type BRAF, mutant BRAF was significantly associated with increased overall mortality risk in patients with EOCRC (pooled HR=2.85, 95%CI: 1.45-5.60, P=0.002). Subgroup analysis showed that the incidence of BRAF gene mutation was higher in Eastern than in Western countries, whereas the incidence of TP53, KRAS, NRAS, and APC gene mutations was lower. There was no significant difference in the incidence of PTEN gene mutation between different regions. Conclusion:Compared with colorectal cancer occurring in the general population, the incidence of APC and KRAS mutations is lower in EOCRC, whereas the incidence of TP53 mutation remains consistent. BRAF mutation is associated with increased overall mortality risk in patients with EOCRC.

Objective:The incidence of early-onset colorectal cancer (EOCRC) is increasing globally; however, the molecular characteristics and prognosis of sporadic EOCRC are unclear. In this systematic review and meta-analysis, we aimed to investigate the incidence of gene mutations and their association with cancer survival in sporadic EOCRC, focusing on six common gene mutations ( TP53, BRAF, KRAS, NRAS, PTEN, and APC). Methods:Ovid Embase and Ovid Medline electronic databases were searched for studies involving patients with sporadic EOCRC (i.e., diagnosed with colorectal cancer before the age of 50 years and with no evidence of hereditary syndromes predisposing to colorectal cancer). The included articles were evaluated using quality assessment tools. Meta-analysis was performed using random-effects and fixed-effects models. Cochran's Q statistic and the I2 index were used to assess heterogeneity. The incidence of the six common gene mutations listed above in sporadic EOCRC and their association with cancer survival were evaluated.Results:(1) Incidence of specific gene mutations in sporadic EOCRC. A total of 34 articles were included in this meta-analysis. The incidence of APC gene mutation was 36% (from 13 articles, 95%CI: 19%-55%, P=0.043); of KRAS gene mutation 30% (from 26 articles, 95%CI: 24%-35%, P=0.190); of BRAF gene mutation 7% (from 18 articles, 95%CI: 5%-11%, P=0.422); of NRAS gene mutation 4% (from five articles, 95%CI: 3%-5%, P=0.586); of PTEN gene mutation 6% (from six articles, 95%CI: 4%-10%, P=0.968); and of TP53 gene mutation 59% (from 13 articles, 95%CI: 49%-68%, P=0.164). (2) Association between gene mutations and survival in sporadic EOCRC . A total of six articles were included in this meta-analysis. Compared with wild-type BRAF, mutant BRAF was significantly associated with increased overall mortality risk in patients with EOCRC (pooled HR=2.85, 95%CI: 1.45-5.60, P=0.002). Subgroup analysis showed that the incidence of BRAF gene mutation was higher in Eastern than in Western countries, whereas the incidence of TP53, KRAS, NRAS, and APC gene mutations was lower. There was no significant difference in the incidence of PTEN gene mutation between different regions. Conclusion:Compared with colorectal cancer occurring in the general population, the incidence of APC and KRAS mutations is lower in EOCRC, whereas the incidence of TP53 mutation remains consistent. BRAF mutation is associated with increased overall mortality risk in patients with EOCRC.

Objectives:To evaluate the coronary microcirculatory function in patients with hypertrophic cardiomyopathy(HCM). Methods:Patients who diagnosed with HCM and underwent the measurement of index of microcirculatory resistance(IMR)using pressure-sensing guide wire from November 2021 to April 2023 were prospectively included.Coronary microcirculatory dysfunction(CMD)was defined as IMR≥25 U and patients were grouped accordingly to compare the clinical characteristics. Results:A total of 25 HCM patients were included.Mean age was(58.4±13.3)years,18 were men and mean body mass index was(26.7±3.6)kg/m2.Coronary microcirculatory function was successfully evaluated in all patients and the mean value of IMR was(30.5±15.3)U.There were 15 patients with CMD.Baseline clinical characteristics,laboratory examinations and medications were simialr between patients with and without CMD.The maximal left ventricular wall was significant thicker in patients with CMD compared with that in patients without CMD([20.2±2.8]mm vs.[16.9±2.3]mm,P=0.005).There was no significant difference in other echocardiographic parameters between two groups(all P>0.05).In the range of IMR value less than 50 U(n=22),there was a significant linear positive correlation between maximal left ventricular wall thickness and IMR(r=0.423,P=0.049).There was no significant difference in coronary flow reserve and fractional flow reserve between two groups. Conclusions:The severity of CMD is positively correlated with left ventricular wall thickness in HCM patients.

Objective To investigate the underlying mechanism of metformin's protective effect on articular cartilage in rats afflicted with osteoarthritis.Methods Thirty male SD rats were randomly divided into three groups(n=10 per group)to establish a rat model of knee osteoarthritis.The metformin group received metformin via gavage[200 mg/(kg·d)],while the control and model groups received saline as a control.After 4 weeks,morphological staining was used to observe articular cartilage morphology,and immunohistochemical staining,immunofluores-cence staining,and Western blot were employed to detect the expression of factors related to the SIRT1/p53 signaling pathway,inflammation,and apoptosis.Results Compared to the model group,the metformin group exhibited significantly reduced cartilage structural damage,characterized by a smoother cartilage surface,increased chondrocyte population,and enhanced proteoglycan content.Immunohistochemical staining,immunofluorescence staining,and Western blot analysis revealed significantly higher expression levels of SOX9,Aggrecan,Bcl-2,and SIRT1 proteins in the metformin-treated cartilage tissue compared to the model group.Conversely,lower expression levels of IL-6 TNF-α BAX Caspase-9 and p53 proteins were observed in the metformin group compared to the model group.TUNEL staining results demonstrated a significant reduction in apoptotic chondrocytes within the metformin-treated group when compared with the model group.Conclusion Metformin exerts a protective effect on articular cartilage in SD rat models of osteoarthritis by activating the SIRT1/p53 signaling pathway,leading to decreased chondrocyte apoptosis and inhibition of extracellular matrix degradation.