1.Hepatic nontargeted lipidomics study for mechanism of Shaqi concen-trated pills attenuating high-fat diet-induced metabolic dysfunction-asso-ciated fatty liver disease in mice
Shuyin BAO ; Xuan WANG ; Pengju BAI ; Qiong WU ; Qianqian MA
Chinese Journal of Pathophysiology 2025;41(8):1504-1513
AIM:This study aimed to investigate the differences in hepatic lipid metabolites in ICR mice in-duced by a high-fat diet and treated with Shaqi concentrated pills(SQ).METHODS:Thirty 8-week-old SPF-grade ICR mice were randomly assigned to five groups:the normal(CON,n=6)group,the high-fat diet model(HFD,n=6)group,the low-dose SQ administration(SQL,n=6)group,the high-dose SQ administration(SQH,n=6)group,and the liver-protecting tablets positive control(PLT,n=6)group.The HFD group was fed a diet consisting of 60%fat for 8 weeks to es-tablish a metabolic-dysfunction-associated fatty liver disease model.Upon successful model establishment,the SQL group received a daily gavage of 395 mg/kg for 4 weeks,while the SQH group received 790 mg·kg-1·d-1.The PLT group was ad-ministered liver-protecting tablets at a dosage of 0.655 g/kg via gavage.Body weight and food intake were monitored week-ly.Liver indices,including Lee's index,triglyceride(TG),total cholesterol(TC),high-density lipoprotein cholesterol(HDL-C),low-density lipoprotein cholesterol(LDL-C),alanine aminotransferase(ALT),and aspartate transaminase(AST)levels,were measured in each group.Hematoxylin-eosin(HE)staining and oil red O staining were performed to assess the extent of pathological damage in liver tissues.Western blot analysis was conducted to evaluate the protein expres-sion levels of choline/ethanolamine phosphotransferase-1(CEPT1),adipose triglyceride lipase(ATGL),and diacylglycerol acyltransferase(DGAT)in the liver.A non-targeted lipidomic analysis using LC-MS was employed to detect changes in hepatic lipid content,and multivariate statistical analyses(principal component analysis and orthogonal partial least squares discriminant analysis)were utilized to compare lipid metabolic profiles among the groups and identify differential lipid metabolites.RESULTS:Compared to the CON group,mice in the HFD group exhibited significantly increased body weight,blood glucose levels,serum TG,TC,LDL-C,ALT,and AST levels,accompanied by a marked decrease in HDL-C levels.HE and oil red O staining results revealed significant lipid droplet accumulation in the liver tissues of HFD mice.In contrast,mice in the SQL and SQH groups showed significant reductions in body weight,blood glucose,serum TG,TC,LDL-C,ALT,and AST levels,along with increased HDL-C levels and less lipid accumulation in liver tissues compared to the HFD group.Staining of liver sections confirmed that SQ treatment mitigated the abnormal accumulation of lipid droplets.Lipidomic analysis indicated that SQ treatment normalized 25 aberrantly expressed lipid metabolites to lev-els comparable to the CON group and identified nine representative differential lipid metabolites.Western blot results dem-onstrated that SQ treatment reduced the protein expression levels of ATGL and DGAT while increasing the expression of CEPT1.CONCLUSION:Treatment with SQ can alleviate metabolic dysfunction-associated fatty liver disease(MAFLD)by modulating triglyceride metabolism,phosphatidylcholine metabolism,and dimethylphosphatidylethanolamine lipid me-tabolism,thereby altering the hepatic lipid profile in MAFLD mice.
2.Hepatic nontargeted lipidomics study for mechanism of Shaqi concen-trated pills attenuating high-fat diet-induced metabolic dysfunction-asso-ciated fatty liver disease in mice
Shuyin BAO ; Xuan WANG ; Pengju BAI ; Qiong WU ; Qianqian MA
Chinese Journal of Pathophysiology 2025;41(8):1504-1513
AIM:This study aimed to investigate the differences in hepatic lipid metabolites in ICR mice in-duced by a high-fat diet and treated with Shaqi concentrated pills(SQ).METHODS:Thirty 8-week-old SPF-grade ICR mice were randomly assigned to five groups:the normal(CON,n=6)group,the high-fat diet model(HFD,n=6)group,the low-dose SQ administration(SQL,n=6)group,the high-dose SQ administration(SQH,n=6)group,and the liver-protecting tablets positive control(PLT,n=6)group.The HFD group was fed a diet consisting of 60%fat for 8 weeks to es-tablish a metabolic-dysfunction-associated fatty liver disease model.Upon successful model establishment,the SQL group received a daily gavage of 395 mg/kg for 4 weeks,while the SQH group received 790 mg·kg-1·d-1.The PLT group was ad-ministered liver-protecting tablets at a dosage of 0.655 g/kg via gavage.Body weight and food intake were monitored week-ly.Liver indices,including Lee's index,triglyceride(TG),total cholesterol(TC),high-density lipoprotein cholesterol(HDL-C),low-density lipoprotein cholesterol(LDL-C),alanine aminotransferase(ALT),and aspartate transaminase(AST)levels,were measured in each group.Hematoxylin-eosin(HE)staining and oil red O staining were performed to assess the extent of pathological damage in liver tissues.Western blot analysis was conducted to evaluate the protein expres-sion levels of choline/ethanolamine phosphotransferase-1(CEPT1),adipose triglyceride lipase(ATGL),and diacylglycerol acyltransferase(DGAT)in the liver.A non-targeted lipidomic analysis using LC-MS was employed to detect changes in hepatic lipid content,and multivariate statistical analyses(principal component analysis and orthogonal partial least squares discriminant analysis)were utilized to compare lipid metabolic profiles among the groups and identify differential lipid metabolites.RESULTS:Compared to the CON group,mice in the HFD group exhibited significantly increased body weight,blood glucose levels,serum TG,TC,LDL-C,ALT,and AST levels,accompanied by a marked decrease in HDL-C levels.HE and oil red O staining results revealed significant lipid droplet accumulation in the liver tissues of HFD mice.In contrast,mice in the SQL and SQH groups showed significant reductions in body weight,blood glucose,serum TG,TC,LDL-C,ALT,and AST levels,along with increased HDL-C levels and less lipid accumulation in liver tissues compared to the HFD group.Staining of liver sections confirmed that SQ treatment mitigated the abnormal accumulation of lipid droplets.Lipidomic analysis indicated that SQ treatment normalized 25 aberrantly expressed lipid metabolites to lev-els comparable to the CON group and identified nine representative differential lipid metabolites.Western blot results dem-onstrated that SQ treatment reduced the protein expression levels of ATGL and DGAT while increasing the expression of CEPT1.CONCLUSION:Treatment with SQ can alleviate metabolic dysfunction-associated fatty liver disease(MAFLD)by modulating triglyceride metabolism,phosphatidylcholine metabolism,and dimethylphosphatidylethanolamine lipid me-tabolism,thereby altering the hepatic lipid profile in MAFLD mice.
3.Application of remote fundus image reading training in improving the diagnostic ability of ophthalmologists in Xinjiang Uygur Autonomous Region
Xufeng ZHAO ; Xiuyan MA ; Xuejing LI ; Li QIN ; Lina SUO ; Wenping MA ; Shihao LI ; Jiaxing CHEN ; Yaxin YANG ; Xinxia LI ; Tao XIE ; Juan LI ; Yuanshan XIONG ; Zhiyong WU ; Zhihuai CHANG ; Mowen FANG ; Ting WANG ; Hong LIU ; Pengju MA ; Pengpeng LIU ; Jing RAN ; Di YANG ; Changle XUE ; Zhen XIE ; Zhao FAN ; Yang WANG ; Peng LEI ; Qingjiang HUANG ; Bing LI ; Shi FENG ; Zhangwanyu WEI ; Yishuang MAO ; Weihong YU ; Linjun ZHANG
Chinese Journal of Ocular Fundus Diseases 2024;40(12):941-946
Objective:To explore the feasibility and effect of remote medical education model using online film reading training to improve the ability of ophthalmologists in the Xinjiang Uygur Autonomous Region (hereinafter referred to as "Xinjiang Region" ) in diagnosing fundus diseases.Methods:The three-level film reading training system of Xinjiang Production and Construction Corps system division hospital-Corps Hospital-Peking Union Medical College Hospital was established. From June 2022 to January 2023, 4 159 posterior color fundus images were continuously collected from Department of Ophthalmology of Xinjiang Corps Hospital and 4 divisional hospitals in the Corps medical system. Among them, hypertensive retinopathy, diabetic retinopathy, exudative age-related macular degeneration (AMD), atrophic AMD and retinal vein occlusion were 3 073, 651, 43, 186 and 206 cases, respectively. The images were divided into 3 rounds (first, second and last) according to the proportion of diseases. The doctors who participated in the training (hereinafter referred to as the "training") were 15 ophthalmologists from the Corps Hospital of Xinjiang Region and the division hospital of the Corps system. There were 7 male and 8 female. Age was (38.1±4.0) years. The titles of senior, deputy senior, intermediate and junior are 1, 6, 5 and 3 respectively; Bachelor's degree and master's degree are 13 and 2 respectively. The working time of fundus disease specialty was (9.6±3.3) years. The film reading system training was conducted before the first round of labeling, and after each round of film reading, the doctors of Peking Union Medical College Hospital gave feedback and explanation on the film reading results. The diagnostic consistency, sensitivity and specificity were compared by paired sample t test. Spearman or Pearson correlation analysis was conducted between the improvement of diagnostic level and professional title, education, age and working hours of ocular fundus disease. Results:All the participating doctors completed the first, second and last reading. After each round of film reading, the film reading summary was carried out for 2 hours. The average diagnostic agreement rates of participating physicians were 53.0%, 67.0% and 75.0%, respectively. The sensitivity and specificity were 0.38, 0.69, 054 and 0.66, 0.85, 0.96, respectively. There was significant difference between the first and last examination ( P<0.001). The sensitivity of the second reading was significantly higher than that of the first reading, and the sensitivity of the last reading was significantly lower than that of the second reading, with statistical significance ( P<0.05). The specificity of the second reading was significantly higher than that of the first reading, and the last reading was significantly higher than that of the second reading, with statistical significance ( P<0.05). There was no significant correlation ( P>0.05) between the improvement of diagnostic level of participating physicians and educational background ( Rho=0.07), professional title ( Rho=0.13), age ( r=0.20), and working time of ophthalmofundus disease specialty ( r=0.26). Conclusions:Relying on the three-level online telemedicine training, it can improve the ability of ophthalmologists in Xinjiang region to diagnose fundus diseases. The preliminary telemedicine education model has demonstrated potential for feasibility and effectiveness in remote areas with inadequate medical resources.
4.Application of remote fundus image reading training in improving the diagnostic ability of ophthalmologists in Xinjiang Uygur Autonomous Region
Xufeng ZHAO ; Xiuyan MA ; Xuejing LI ; Li QIN ; Lina SUO ; Wenping MA ; Shihao LI ; Jiaxing CHEN ; Yaxin YANG ; Xinxia LI ; Tao XIE ; Juan LI ; Yuanshan XIONG ; Zhiyong WU ; Zhihuai CHANG ; Mowen FANG ; Ting WANG ; Hong LIU ; Pengju MA ; Pengpeng LIU ; Jing RAN ; Di YANG ; Changle XUE ; Zhen XIE ; Zhao FAN ; Yang WANG ; Peng LEI ; Qingjiang HUANG ; Bing LI ; Shi FENG ; Zhangwanyu WEI ; Yishuang MAO ; Weihong YU ; Linjun ZHANG
Chinese Journal of Ocular Fundus Diseases 2024;40(12):941-946
Objective:To explore the feasibility and effect of remote medical education model using online film reading training to improve the ability of ophthalmologists in the Xinjiang Uygur Autonomous Region (hereinafter referred to as "Xinjiang Region" ) in diagnosing fundus diseases.Methods:The three-level film reading training system of Xinjiang Production and Construction Corps system division hospital-Corps Hospital-Peking Union Medical College Hospital was established. From June 2022 to January 2023, 4 159 posterior color fundus images were continuously collected from Department of Ophthalmology of Xinjiang Corps Hospital and 4 divisional hospitals in the Corps medical system. Among them, hypertensive retinopathy, diabetic retinopathy, exudative age-related macular degeneration (AMD), atrophic AMD and retinal vein occlusion were 3 073, 651, 43, 186 and 206 cases, respectively. The images were divided into 3 rounds (first, second and last) according to the proportion of diseases. The doctors who participated in the training (hereinafter referred to as the "training") were 15 ophthalmologists from the Corps Hospital of Xinjiang Region and the division hospital of the Corps system. There were 7 male and 8 female. Age was (38.1±4.0) years. The titles of senior, deputy senior, intermediate and junior are 1, 6, 5 and 3 respectively; Bachelor's degree and master's degree are 13 and 2 respectively. The working time of fundus disease specialty was (9.6±3.3) years. The film reading system training was conducted before the first round of labeling, and after each round of film reading, the doctors of Peking Union Medical College Hospital gave feedback and explanation on the film reading results. The diagnostic consistency, sensitivity and specificity were compared by paired sample t test. Spearman or Pearson correlation analysis was conducted between the improvement of diagnostic level and professional title, education, age and working hours of ocular fundus disease. Results:All the participating doctors completed the first, second and last reading. After each round of film reading, the film reading summary was carried out for 2 hours. The average diagnostic agreement rates of participating physicians were 53.0%, 67.0% and 75.0%, respectively. The sensitivity and specificity were 0.38, 0.69, 054 and 0.66, 0.85, 0.96, respectively. There was significant difference between the first and last examination ( P<0.001). The sensitivity of the second reading was significantly higher than that of the first reading, and the sensitivity of the last reading was significantly lower than that of the second reading, with statistical significance ( P<0.05). The specificity of the second reading was significantly higher than that of the first reading, and the last reading was significantly higher than that of the second reading, with statistical significance ( P<0.05). There was no significant correlation ( P>0.05) between the improvement of diagnostic level of participating physicians and educational background ( Rho=0.07), professional title ( Rho=0.13), age ( r=0.20), and working time of ophthalmofundus disease specialty ( r=0.26). Conclusions:Relying on the three-level online telemedicine training, it can improve the ability of ophthalmologists in Xinjiang region to diagnose fundus diseases. The preliminary telemedicine education model has demonstrated potential for feasibility and effectiveness in remote areas with inadequate medical resources.
5.Complement C3a receptor 1 high-expression progresses malignancy in glioma through nuclear factor-kappa B signaling pathway
Xiansheng LI ; Yanjuan MA ; Pengju MA ; Zhijie YIN ; Fuguang LI ; Weilong YANG ; Lei HUI
Chinese Journal of Neuromedicine 2022;21(12):1195-1202
Objective:To investigate the complement C3a receptor 1 (C3AR1) expression in glioma and its mechanism in progressing malignancy.Methods:(1) The C3AR1 mRNA expression data and clinical information were obtained in 607 glioma patients from The Cancer Genome Atlas (TCGA) database and 656 glioma patients from Chinese Glioma Genome Atlas (CGGA) database; the differences in C3AR1 mRNA expression were analyzed among gliomas with different World Health Organization (WHO) grading. The overall survival and disease-free survival were compared between high and low C3AR1 mRNA expression patients obtained from TCGA database by Gene expression profiling interactive analysis (GEPIA). Gene body (GO) function analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of C3AR1 related differentially expressed genes were performed by DAVID database. Correlation of C3AR1 mRNA expression with immune cell infiltration was analyzed using TIMER online website. (2) The brain tissues from 3 non-tumor patients and 9 glioma patients surgically resected in Department of Neurosurgery, First Affiliated Hospital of Xinxiang Medical University from January 2019 to September 2021 were collected; the C3AR1 protein expression was detected by Western blotting. (3) The in vitro cultured U87 and U251 cells were divided into negative control group and C3AR1 knockdown group ( C3AR1 being knocked down by lentivirus transfection); and CCK-8 assay, plate cloning assay and Transwell assay were used to detect the proliferation rate, number of colony formation and number of membrane penetrating cells. Western blotting was used to detect the nuclear factor-κB (NF-κB) signaling pathway protein expressions. Results:(1) In TCGA database, the C3AR1 mRNA expression in gliomas of WHO grading II, grading III and grading IV increased sequentially, with significant differences ( P<0.05). In CGGA database, the C3AR1 mRNA expression in glioma of WHO grading IV was statistically higher than that in gliomas of WHO grading II and grading III ( P<0.05). GEPIA showed that the overall survival and disease-free survival in the low C3AR1 mRNA expression group were statistically higher than those in the high C3AR1 mRNA expression group ( P<0.05). GO function analysis and KEGG pathway enrichment analysis revealed that C3AR1 related differentially expressed genes were more enriched in such biological processes and signaling pathways as calcium homeostasis, membrane structural valves, proton transmembrane transporter protein activity, chemokine signaling pathway and NF-κB signaling pathway. TIMER showed that C3AR1 mRNA expression in glioblastoma and low-grade glioma was positively correlated with infiltration degrees of B cells, CD4 + T cells, neutrophils, macrophages and dendritic cells, and C3AR1 mRNA expression in glioblastoma was negatively correlated with infiltration degree of CD8 + T cells ( P<0.05). (2) C3AR1 protein expression in glioma tissues was significantly higher than that in non-tumor tissues. (3) Compared with the negative control group, the C3AR1 knockdown group group had significantly lower proliferation rate, smaller numbers of colony formation and membrane penetrating cells, and lower expressions of NF-κB, phosphorylated (p)-NF-κB, p-NF-κB inhibitory protein (IκB)α, p-I-κB kinase (IKK)α and N-cadherin, and significantly higher E-cadherin expression. Conclusion:C3AR1 is highly expressed in glioma and progresses malignancy through NF-κB signaling pathway.
6.Role of human umbilical cord mesenchymal stem cells in the maturation and differentiation of dendritic cells in patients with severe acute pancreatitis and its mechanism in the inflammation modulation
Make LIANG ; Xuemin LI ; Lei SUN ; Baodong MA ; Pengju LYU ; Han YUE
Chinese Journal of Pancreatology 2021;21(3):201-207
Objective:To investigate the effect of human umbilical cord mesenchymal stem cells (hUC-MSCs) on the maturation and differentiation of dendritic cells (DCs) and the mechanism involved in the regulation of inflammation in patients with severe acute pancreatitis (SAP).Methods:The full-term fetal umbilical cords(about 4-5 cm) were collected from Zhengzhou Central Hospital Affiliated to Zhengzhou University after cesarean section. hUC-MSCs were isolated and cultured in primary culture. Flow cytometry was used for phenotype identification, adipogenic and osteogenic staining. 20 ml peripheral blood samples from 5 SAP patients were collected, and monocytes were isolated using lymphocyte separation solution and then induced by adding granulocyte macrophage colony-stimulating factor (GM-CSF), IL-4 and tumor necrosis factor (TNF)-α and cultured as DCs. According to different culture methods, DCs were divided into DCs group, hUC-MSCs+ DCs group and hUC-MSCs+ DCs+ NS398 group (NS398 was a specific inhibitor of COX-2, a downstream regulatory gene of NF-κB). The phenotype of DCs was detected by flow cytometry, and the levels of IL-1β, IL-lα, IL-2, IL-6 and IL-10 in the supernatant of cell culture for 24 hours were determined. The expression of toll like receptor (TLR)-4, IKKα and NF-κB-p65 were detected by Western blot.Results:The hUC-MSCs were successfully cultured, and their surface markers CD 90, CD 105 and CD 73 were positively expressed, and they could differentiate into adipocytes and bone cells. With the prolongation of culture time, DCs differentiated from immature to mature cells. Compared with the DCs group, the proportion of regulatory DCs (regDCs) was increased in the hUC-MSCs+ DCs group, and the marker CD 11b was significantly up regulated [(14.26±1.25)% vs (4. 87±0.58)%], CD 1a and CD 11c were significantly down regulated [(2.81±0.34)% vs (13.62±1.52)%, (3.88±0.5)% vs (11. 8±1.22)%]. All the difference were statistically significant ( P<0.05). The expression of IL-1β, INF-γ and IL-6 in culture supernatant were down regulated, but the difference was not statistically significant; The pro-inflammatory factor IL-1α was significantly decreased [(14.91±2.58)ng/L vs (30.19±7.75)ng/L], and the anti-inflammatory factor IL-10 was significantly increased [ (17.03±4.69)ng/L vs (1.83±0.14)ng/L]. The expression levels of NF-κB-p65 and TLR4 were significantly down regulated (0.74±0.02 vs 0.97±0.01, 0.89±0.01 vs 1.72±0.01), and the expression of IKKα protein was significantly up regulated (1.12±0.01 vs 0.21±0.01) in hUC-MSCs-DCs group. All the differences were statistically significant (all P value<0.05). Compared with DCs group and hUC-MSCs+ DCs group, the expression levels of NF-κB-p65 and TLR4 were significantly down regulated (0.34±0.01 vs 0.97±0.01, 0.74±0.02 and 0.14±0.01 vs 1.72±0.01, 0.89±0.01), while the expression of IKKα protein was significantly up regulated (1.68±0.01 vs 0.21±0.01, 1.12±0.01) in hUC-MSCs+ DCS+ NS398 group. All the differences were statistically significant (all P value<0.05). Conclusions:In SAP patients, hUC-MSCs can inhibit the maturation and differentiation of DCs, and induce CD 11bhigh CD 1alow CD 11clowrregDCs to participate in immune regulation, which may play an anti-inflammatory role by inhibiting the inflammatory cascade through TLR4/IKKα/NF-κB/COX-2 pathway.
7.Effect of soluble amyloid precursor protein α on nerve cell apoptosis and neurological function in subarachnoid hemorrhage rats
Haigang CHANG ; Lei HUI ; Pengju MA ; Xiangsheng LI ; Ruihua LIU ; Baozhe JIN
Chinese Journal of Geriatric Heart Brain and Vessel Diseases 2018;20(6):639-641
Objective To study the effect of soluble amyloid precursor protein α (sAPPα) on nerve cell apoptosis and neurological function in subarachnoid hemorrhage (SAH) rats.Methods Sixty male SD rats were randomly divided into control group (n=20),SAH+saline group (n=20) and SAH+sAPPα group (n=20).A SAH model was established by injecting autologous blood into cistern magna in rats.After a SAH model was established for SAH + saline group and SAH + sAPPα group by injecting saline and sAPPα respectively into the cistern magna of rats,the apoptotic cells were detected by immunofluorescene with TUNEL staining and the neurological function was scored in 10 rats from each group on day 3 after injection of sAPPα and saline.Results The number of apoptotic cells in brain tissue was significantly greater in SAH+saline group than in control group (P<0.05) and was significantly smaller in SAH+sAPPα group than in SAH+ saline group (P<0.05).The neurological function score was 26.7±0.5,13.9±0.7 and 23.0±0.8 respectively in control group,SAH + saline group and SAH + sAPPα group.Conclusion sAPPα alleviates secondary damage of neurological function by inhibiting the apoptosis of nerve cells in rats after SAH and can thus improve their neurological function.
8.Epidural fluid collection secondary to cranioplasty in 47 patients with traumatic brain injury after decompressive craniectomy
Haigang CHANG ; Yaxiao WANG ; Pengju MA ; Fazheng SHEN ; Zhijie YIN ; Ruihua LIU ; Yangyang WANG ; Xiang ZHOU ; Baozhe JIN
Chinese Journal of Neuromedicine 2018;17(3):248-253
Objective To identify the reasons and treatment strategies of epidural fluid collection (EFC) secondary to cranioplasty in patients with traumatic brain injury after decompressive craniectomy.Methods From June 2013 to July 2017,a retrospective analysis was performed on clinical data of 150 patients with traumatic brain injury after decompressive craniectomy in our hospital.A total of 47 patients experienced EFC following cranioplasty and 103 not.Risk factors of EFC after cranioplasty were analyzed by multiple factor Logistic regression.Results For the 47 EFC patients,32 patients had no obvious clinical symptoms and EFC was absorbed gradually through conservative therapy;15 patients had clinical symptoms,such as mental deterioration,headache,or limb weakness.EFC disappeared through vacuation in 4 patients and subcutaneous drainage in 11.The proportions of patients with skull defect>80 cm2,dural defect and dural calcification in patients with EFC were significantly higher as compared with those without EFC (P<0.05).Multiple factor Logistic regression analysis showed that skull defect>80 cm2 and dural mater calcification were independent risk factors for EFC after cranioplasty.Conclusions Patients with large skull defect>80 cm2 and dural calcification are prone to have EFC after cranioplasty.Careful evaluation of imaging data,good surgical skills and strengthening postoperative management can reduce incidence of EFC after cranioplasty.
9.Down-regulation of miRNA-99b expression in mTOR/4E-BP1 signal pathway and invasion in glioma U251 cells
Qiankun JI ; Ke REN ; Jianhua ZHAO ; Zhijie YIN ; Pengju MA ; Ruihua LIU ; Baozhe JIN
Chinese Journal of Neuromedicine 2018;17(9):886-891
Objective To detect the down-regulation ofmiRNA-99b expression in cell invasion and its mechanism in human glioma cell line U251.Methods Glioma cell line U251 were routinely cultured in vitro.(1) U251 cells were divided into blank control group,negative control group and miRNA-99b inhibitor group;cells in the latter two groups were transfected with negative control sequences and miRNA-99b inhibitors,respectively;and cells in the blank control group did not give any treatment;mRNA expressions of miRNA-99b and mammalian target of rapamycin (mTOR) in U251 cells were measured by reverse transcription (RT)-PCR;the changes of mTOR,eIF4E-binding protein 1 (4E-BP1) andphosphorylated (p)-4E-BPl protein expressions in U251 cells were detected by Westem blotting;cell invasion was evaluated by Transwell assay.(2) U251 cells were divided into negative control group Ⅰ and mTOR siRNA group,and cells in the two groups were transfected with negative control sequences and mTOR siRNA,respectively;the miRNA-99b and mTOR mRNA expressions in U251 cells were measured by RT-PCR;the mTOR and p-4E-BP1 protein expressions in U251 cells were measured by Western blotting.(3) U251 cells were divided into miRNA-99b inhibitor+negative control group and miRNA-99b inhibitor+mTOR siRNA group,and cells in the two groups were transfected with miRNA-99b inhibitor+negative control sequences and miRNA-99b inhibitor+mTOR siRNA,respectively;the p-4E-BP1 protein expression in U251 cells was measured by Western blotting;cell invasion was evaluated by Transwell assay.Results (1) As compared with those in the blank control group and negative control group,the miRNA-99b rnRNA expression was significantly decreased,the mTOR mRNA and protein expressions and p-4E-BP1 protein expression were significantly increased,and the number of transmembrane cells was significantly larger in U251 cells of miRNA-99b inhibitor group (P<0.05);there were no significant differences in 4E-BP1 protein expression among the three groups (P>0.05).(2) As compared with those in the negative control group Ⅰ,the mTOR mRNA and protein expressions and p-4E-BP1 protein expression were significantly decreased in U251 cells of mTOR siRNA group (P<0.05);there was no significant difference in miRNA-99b mRNA expression between the two groups (P>0.05).(3) As compared with those in the miRNA-99b inhibitor+negative control group,the p-4E-BP1 protein expression and number of transmembrane cells were significantly decreased/smaller in U251 cells ofmiRNA-99b inhibitor+mTOR siRNA group (P<0.05).Conclusions Down-regulation ofmiRNA-99b expression promotes glioma cell invasion,and its mechanism is related to the regulation of mTOR/4E-BP1 signaling pathway.
10.Effects of three kinds of nanoparticles on proliferation and apoptosis of esophageal squamous carcinoma cells
Pengli HAN ; Lei SUN ; Pengju LYU ; Fenfen GONG ; Chao MA ; Guo CHEN ; Yiran ZHU ; Tian XIA ; Wei CAO
Chinese Pharmacological Bulletin 2016;32(6):789-794
Aim TostudytheeffectsofCuO,ZnOand TiO2 nanoparticles on the viability and metastatic po-tential of EC-9706 and EC-109 esophageal squamous carcinomacelllineinvitro.Methods Characteristics of CuO,ZnO and TiO2 nanoparticles were detected u-sing transmission electron microscope (TEM)and dy-namic light scattering (DLS ).EC-9706 and EC-109 cells were treated with different concentrations of CuO, ZnO and TiO2 (5 ~80 mg · L-1 ).The cell prolifera-tion was analyzed by MTT assay.The cell cycle and apoptotic rates were determined by flow cytometry (FCM).The cell invasion was assayed in Transwell chambers.The expression of Bcl-2 and caspase-3 pro-tein in cells was detected by Western blot method.Re-sults CuO,ZnOandTiO2nanoparticleswerespheri-cal with primary particle size 12,20. 6,12 nm.The particles were agglomerated in water and cell culture medium with negative charge.CuO and ZnO nanoparti-cles induced decreases in EC-9706 and EC-109 cell vi-ability dose-dependently.After exposed to increasing concentrations of CuO and ZnO nanoparticles,the cell cycle analysis revealed a decreasing proportion of cells in G2/Mand S phase,and up-regulation of the cells in G0/G1 phase.Apoptotic cells also increased along with decreased cell invasion upon CuO and ZnO treatment. Nanoparticles treatment after 48 h, the activated caspase-3 expression quantity increased significantly and the Bcl-2 expression quantity decreased obviously (P<0. 05 )compared with control group.TiO2 nanop-articles had no obvious effect on the EC-9706 and EC-109 cell proliferation,cell cycle,apoptosis and inva-sion.Conclusion ComparedwithTiO2,CuOand ZnO nanoparticles can inhibit EC-9706 and EC-109 cell viability and metastatic potential,the mechanism of action involves cell cycle arrest in G0/G1 phase and apoptosis.These findings can help the development of nanoparticles as anti-cancer therapeutics for esophageal cancer.

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