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1.
Yao Xue Xue Bao ; (12): 1256-1266, 2023.
Artigo em Chinês | WPRIM | ID: wpr-978675

RESUMO

Our studies were aimed to explore the effect and mechanism of the inhibition of the formation of vasculogenic mimicry (VM) in human glioblastoma cells by Xihuang pill (XHP) medicated serum through regulating the hypoxia inducible factor-1α (HIF-1α)/vascular endothelial growth factor A (VEGFA)/vascular endothelial growth factor receptor 2 (VEGFR2) signaling pathway. The medicated serum of XHP was prepared by gavage for 7 days to male SD rats (approval number of animal experiment ethics: 202105A051). The hypoxia model of U251 cells was established using 200 μmol·L-1 of CoCl2. After treatment with XHP-medicated serum, cell viability and proliferation of U251 cells were detected by CCK-8 and cell cloning experiment. Cell apoptosis and cell cycle of U251 cells were determined by flow cytometry. Cell migration and invasion were evaluated by wound healing and Transwell invasion assay. The formation of VM was assessed by three-dimensional cell culture of U251 cells. The protein expression levels of HIF-1α, VEGFA, VEGFR2, phosphorylated-VEGFR2 (p-VEGFR2), vascular endothelial-cadherin (VE-cadherin), Eph receptor tyrosine kinases A2 (EphA2), matrix metalloproteinase 2 (MMP2), matrix metalloproteinase 14 (MMP14) and laminin γ2 in U251 cells were detected by Western blot. The results showed that 10% XHP-medicated serum had little effect on the cell viability, proliferation, apoptosis and cell cycle of U251 cells under hypoxia. Compared with the model group, 10% XHP-medicated serum at 1.0, 1.5 and 2.0 h significantly decreased the migration rate (P < 0.01) and the number of invading U251 cells (P < 0.01). 10% XHP-medicated serum at 2.0 h significantly suppressed the formation of VM tubular structures in U251 cells under the condition of hypoxia (P < 0.01). Western blot experiment showed that 10% XHP-medicated serum significantly down-regulated the expression of HIF-1α, VEGFA, phospho-VEGFR2, VE-cadherin, EphA2 and MMP14 proteins (P < 0.05). In conclusion, XHP could inhibit the formation of VM in human glioblastoma U251 cells to suppress the angiogenesis by down-regulating the HIF-1α/VEGFA/VEGFR2 signaling pathway.

2.
Artigo em Chinês | WPRIM | ID: wpr-992079

RESUMO

Objective:To investigate the effects of over-expression of E2F transcription factor 1 (E2F1) on proliferation, invasion, apoptosis and radiosensitivity of glioma cell U251.Methods:Real-time quantitative PCR (qRT-PCR) were used to detect the differential expression of E2F1 mRNA in glioma cells LN18, SW1088, U251 and normal brain glial cells. The stable over-expression of E2F1 plasmid was constructed and transfected into U251 cells. qRT-PCR and Western blot test were used to detect the expression of E2F1, pituitary tumor transforming gene 1(PTTG1), C-Myc, B-cell lymphoma-2 (Bcl-2), Bcl2-associated X (Bax) mRNA and protein expression in the control group and E2F1 over-expression group.U251 cells were divided into control group(no X-ray irradiation), irradiation group(6 Gy dose of X-ray), and irradiation + E2F1 over-expression group(transfected with E2F1 first, then irradiated by 6 Gy of X-ray). Cell proliferation ability was detected by cell counting Kit-8(CCK-8) cell viability detection reagent, and cell invasion and migration ability were detected by Transwell chamber. Apoptosis and cell cycle were detected by flow cytometry.GraphPad Prism 8.0 was used for data analysis.The statistical methods were one-way ANOVA and independent sample t-test. Results:qRT-PCR showed that there was statistical difference in the mRNA levels of E2F1( F=201.92, P<0.05) in different cell lines.The expression levels of E2F1 mRNA in LN18(4.04±0.29), SW1088(3.19±0.16)and U251(4.66±0.20) cells were higher than those in HEB(1.02±0.07)cells ( q=27.00, 19.40, 32.52, all P<0.05). After successfully constructing U251 cells with stable over-expression of E2F1 plasmid, qRT-PCR and Western blot detection results showed that: the mRNA and protein levels of E2F1, PTTG1, C-Myc and Bcl-2 in E2F1 over-expression group were higher than those in control group ( t=77.16, 57.88, 4.63, 51.13, 7.50, 70.85, 8.38, 48.81, all P<0.05). Bax mRNA(0.20±0.01) and protein(0.66±0.01) levels were lower than those in control group((1.00±0.02), (0.94±0.01)), and the differences were statistically significant ( t=1.74, 54.65, both P<0.05). After X-ray irradiation (6 Gy), CCK8 detection results showed: the proliferation ability of the three groups at 24, 48, 72 and 96 h were significantly different ( F=95.41, 187.53, 1 158.49, 7 883.78, all P<0.05). The proliferation capacity of the irradiation group were lower than those of the control group at 24, 48, 72 and 96 h ( q=19.51, 27.20, 66.60, 174.9, all P<0.05). The proliferation capacity of irradiation + E2F1 over-expression group at 24, 48, 72 and 96 h were higher than those of irradiation group ( q=10.63, 10.81, 21.11, 60.90, all P<0.05). Transwell assay results showed that there were significant differences in cell invasion and migration ability among the three groups ( F=315.38, 681.10, both P<0.05). The invasion and migration ability of cells in the irradiation group were lower than those in the control group ( q=35.09, 12.76, both P<0.05), and the invasion and migration ability of cells in the irradiation + E2F1 over-expression group were higher than those in the irradiation group ( q=52.06, 22.81, both P<0.05). Flow cytometry showed that there were significant differences in apoptosis rate and percentage of cells in each cycle among the three groups ( F=667.63, 3 213.30, 3 011.26, 861.98, all P<0.05). The percentage of the apoptosis rate, S phase and G2 phase cells in the irradiation group were higher than those in the control group ( q=51.10, 89.39, 51.82, all P<0.05), while the percentage of G1 phase cells in the irradiation group was lower than that in the control group ( q=141.2, P<0.05). The apoptosis rate and percentage of S phase and G2 phase cells in the irradiation + E2F1 over-expression group were lower than those in the irradiation group ( q=18.87, 41.42, 29.31, all P<0.05), while the number of G1 phase cells in the irradiation + E2F1 over-expression group was lower than that in the irradiation group ( q=70.73, P<0.05). Conclusion:Over-expression of E2F1 can reduce the radiosensitivity of glioma U251 cells by regulating the expression of mRNA and protein of genes related to cell cycle and apoptosis, and E2F1 may be involved in the radioresistance of glioma cells.

3.
China Pharmacy ; (12): 2471-2475, 2023.
Artigo em Chinês | WPRIM | ID: wpr-997004

RESUMO

ABSTRACT OBJECTIVE To investigate the effects and mechanism of ginsenoside Rh2 on the proliferation and apoptosis in human glioma U87 and U251 cells. METHODS Using human glioma U87 and U251 cells as subjects, the proliferation and apoptosis, as well as the expression of histone deacetylase 1(HDAC1) protein and apoptosis-related proteins [B cell lymphoma-2(Bcl-2), Bcl-2-associated X protein (Bax) and cleaved caspase-3] were detected after being treated with different concentrations of ginsenoside Rh2. RESULTS The concentrations of 10,20,30,40,50,60,70,80 μmol/L ginsenoside Rh2 could generally significantly increase the proliferation inhibition rate of U87 and U251 cells (P<0.05 or P<0.01), and the half inhibitory concentrations of this component after 48 hours of action were 51.34 and 55.84 μmol/L, respectively;30,50 μmol/L ginsenoside Rh2 could increase the total apoptotic rate of both types of cells, reduced the protein expressions of HDAC1 and Bcl-2, and increased the protein expressions of Bax and cleaved caspase-3 significantly (P<0.05 or P<0.01). CONCLUSIONS Ginsenoside Rh2 has a significant inhibitory effect on the proliferation of glioma cells and promotes the apoptosis of cells, which may be through reducing the expression of HDAC1 protein and activating the Bcl-2 family protein-mediated apoptosis pathway.

4.
Artigo em Chinês | WPRIM | ID: wpr-941019

RESUMO

OBJECTIVE@#To explore the effect of atorvastatin (AVT) on biological behaviors and the miR-146a/PI3K/Akt signaling pathway in human glioma cells.@*METHODS@#Human glioma U251 cells were treated with 8.0 μmol/L AVT or transfected with a miR-146a inhibitor or a negative control fragment (miR-146a NC) prior to AVT treatment. RT-PCR was used to detect miR-146a expression in the cells, and the changes in cell proliferation rate, apoptosis, cell invasion and migration were detected using MTT assay, flow cytometry, and Transwell assay. Western blotting was performed to detect the changes in cellular expressions of proteins in the PI3K/Akt signaling pathway.@*RESULTS@#AVT treatment for 48 h resulted in significantly increased miR-146a expression and cell apoptosis (P < 0.01) and obviously lowered the cell proliferation rate, invasion index, migration index, and expressions of p-PI3K and p-Akt protein in U251 cells (P < 0.01). Compared with AVT treatment alone, transfection with miR-146a inhibitor prior to AVT treatment significantly reduced miR-146a expression and cell apoptosis (P < 0.01), increased the cell proliferation rate, promoted cell invasion and migration, and enhanced the expressions of p-PI3K and p-Akt proteins in the cells (P < 0.01); these effects were not observed following transfection with miR-146a NC group (P>0.05).@*CONCLUSION@#AVT can inhibit the proliferation, invasion and migration and promote apoptosis of human glioma cells possibly by up-regulating miR-146a expression and inhibiting the PI3K/Akt signaling pathway.


Assuntos
Humanos , Apoptose , Atorvastatina/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Glioma/patologia , MicroRNAs/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais
5.
Braz. J. Pharm. Sci. (Online) ; 58: e181116, 2022. graf
Artigo em Inglês | LILACS-Express | LILACS | ID: biblio-1374540

RESUMO

Abstract The aim of this study was to investigate the effect of TiO2/N-succinyl-chitosan composite (TiO2/ NSCS) photodynamic therapy (PDT), while considering the effects of various light sources on the activation of photosensitizer. The methyl thiazolyl tetrazolium assay was used to examine the cell survival rate of the cells. The results showed that glioma cell strain (U251) was the most sensitive cancer cell strain to TiO2/NSCS. When the concentration of TiO2/NSCS was between 0 and 800 μg·mL-1, there was no obvious cytotoxicity to normal liver cells (HL-7702) and U251 cells. During the PDT process, the photokilling effect of TiO2/NSCS on U251 cells under ultraviolet-A (UVA) light irradiation was stronger than that of pure TiO2, and its killing effects were positively correlated with concentration and irradiation time. In addition, both UVA and visible light could excite TiO2/ NSCS, which had significant killing effect on U251 cells. The results of acridine orange/ethidium bromide fluorescent double staining and Annexin V/propidium iodide double staining indicated that TiO2/NSCS under UVA and visible light irradiation could kill U251 cells by inducing apoptosis, and the apoptosis rate of TiO2/NSCS treatment groups was higher than that of TiO2 treatment groups. Therefore, TiO2/NSCS might be used as a potential photosensitizer in PDT.

6.
Artigo em Chinês | WPRIM | ID: wpr-886478

RESUMO

@#[摘 要] 目的:探讨四氧化三铁(Fe3O4)纳米粒子(PION)作为药物载体增强二氢卟吩e6(chlorin e6,Ce6)在胶质瘤中的增效作用。方法:采用高温降解法和相转移法制备PEG-Fe3O4@Ce6复合纳米粒子(PION@E6),用水合粒径分析、透射电镜、胶体稳定性分析、紫外可见光吸收光谱等方法对PION@E6进行鉴定。CCK-8法检测胶质瘤U251细胞的增殖活性,流式细胞术检测细胞的凋亡水平,DCFH-DA探针法检测细胞中活性氧(reactive oxygen species,ROS)的水平。构建BALB/c-nu裸鼠胶质瘤U251细胞移植瘤模型,动物活体荧光成像术及磁共振成像(MRI)观察PION@E6及Ce6在移植瘤中的潴留时间,比较PION@E6声动力治疗组及Ce6声动力治疗组的第28天生存情况及肿瘤体积。结果:PION@E6的核心粒径为10 nm、水合粒径为(37.86±12.90)nm,具有良好的水溶性和稳定性;吸收光谱及XRD图谱显示Ce6已经负载到Fe3O4纳米粒子上。与Ce6声动力组比较,PION@E6声动力组U251细胞的增殖活性显著下降(P<0.05),细胞凋亡率显著升高(均P<0.05),细胞中ROS水平显著升高(P<0.05)。荷瘤裸鼠胶质瘤U251细胞移植瘤治疗实验结果显示,与Ce6声动力治疗组比较,PION@E6声动力治疗组裸鼠移植瘤组织中潴留时间显著延长(P<0.05),存活的裸鼠数显著增多,移植瘤体积显著缩小(P<0.01)。结论:Fe3O4纳米粒子对Ce6介导的胶质瘤U251细胞声动力治疗具有明显的增效作用。

7.
Yao Xue Xue Bao ; (12): 786-792, 2021.
Artigo em Chinês | WPRIM | ID: wpr-876512

RESUMO

Sempervirine, a yohimbane-type alkaloid isolated from Gelsemium elegans, was found to significantly inhibit the cellular proliferation of U251 cells in vitro and in vivo in a dose-dependent manner. U251 cells were treated with 0-16 μmol·L-1 of sempervirine for 24, 48 or 72 h. An MTT assay and clone formation assay were used to investigate cell survival and clone formation. Hoechst staining and Annexin V-FITC/PI staining were used to measure cell apoptosis. The expression of PI3K, AKT, p-AKT, Bax, Bcl-2, caspase-3 and cleaved caspase-3 was determined by Western blot analysis. The antitumor effect of sempervirine in vivo was investigated by inoculating nude mice with U251 cells. All animal experiments were in strict accordance with the regulations of the Biomedical Ethics Committee of Fujian Medical University (Fujian, China). The results show that sempervirine significantly inhibits the proliferation and induces the apoptosis of U251 cells, promotes cleavage of caspase-3, down-regulates the protein expression of PI3K and Bcl-2/Bax, and inhibits phosphorylation of AKT in vitro. Intraperitoneal injection of 4 or 8 mg·kg-1·day-1 of sempervirine inhibits U251 cells tumor growth in the xenograft nude mice, and tumor weight decreased by 44.76% and 61.26%, respectively. Our study shows that sempervirine significantly inhibits the proliferation of U251 cells in vitro and in vivo, laying a foundation for further research and development of its anti-glioma effect.

8.
Artigo em Chinês | WPRIM | ID: wpr-988413

RESUMO

Objective To investigate the inhibitory effect and mechanism of temozolomide on migration and invasion of U251 human glioma cells enhanced by plumbagin. Methods CCK-8 method was used to study the effects of plumbagin, temozolomide and plumbagin+temozolomide on the proliferation of glioma U251 cells. Cell scratch test was used to detect the migration of U251 cells in the control (DMSO), plumbagin, temozolomide and plumbagin+temozolomide groups for 48h. Transwell assay was used to detect the effect of the combination therapy on the invasion of U251 cells. Western blot was used to detect the relative expression levels of E-cadherin in three groups. Results CCK-8 showed that the proliferation inhibition rate of U251 cells treated with plumbagin (1.25 μmol/L) combined with temozolomide (200 μmol/L) for 48h was 75.69%, significantly higher than that treated with plumbagin alone (P=0.012) or temozolomide alone (P=0.034). Cell scratch assay showed that the combination of plumbagin and temozolomide could significantly enhance the inhibition effect of temozolomide on the migration of U251 cells (P=0.023). Transwell assay showed that the invasion ability of U251 cells was significantly decreased after the combination therapy (P < 0.05). The protein expression of E-cadherin in the combination group was significantly higher than those in plumbagin or temozolomide groups (P < 0.05). Conclusion Plumbagin combined with temozolomide can inhibit the migration and invasion of glioma cells and enhance the sensitivity of glioma cells to temozolomide. And the effect is achieved by the protein expression of E-cadherin.

9.
Artigo em Chinês | WPRIM | ID: wpr-910458

RESUMO

Objective:To investigate the effect of hydroxyurea (HU) combined with temozolomide (TMZ) and radiotherapy (RT) on the sensitivity of human glioma U251 cells to chemoradiotherapy (CRT).Methods:Human glioma U251 cell line was cultured in vitro. CCK8 cell assay was used to detect the proliferation activity of U251 cells treated with different concentrations of HU/TMZ under different conditions. Flow cytometry was utilized to detect apoptosis rate and cell cycle distribution of U251 cells. Transwell chamber assay and scratch test were performed to evaluate the changes of cell invasion and migration. The expression levels of apoptosis proteins were determined by Western blot. Colony formation assay was adopted to detect the cell survival fraction . Results:HU concentration at 50μmol/L and below did not affect the proliferation of human glioma U251 cells ( P>0.05). Low-dose HU combined with CRT significantly inhibited cell proliferation ( P<0.05), invasion ( P<0.01) and migration (12h P<0.001, 24h P<0.01), and promoted cell apoptosis ( P<0.01) compared with the use of CRT alone. Application of 50μmol/L HU combined with RT increased the radiosensitivity of cells (SER=1.49), significantly prolonged the cell cycle of S phase and G 2 phase (both P<0.05), considerably up-regulated the expression levels of the apoptosis-associated proteins of Caspase-3 and Bax and significantly down-regulated the expression level of anti-apoptosis protein of Bcl-2(all P<0.001). Conclusions:Compared with CRT, HU combined with CRT can further inhibit the proliferation, invasion and migration of human glioma U251 cells, promote cell apoptosis, increase the radiosensitivity and prolong the cell cycle of S and G 2 phases, thereby enhancing the sensitivity of human glioma U251 cells to CRT.

10.
Chinese Pharmacological Bulletin ; (12): 687-692, 2021.
Artigo em Chinês | WPRIM | ID: wpr-1014419

RESUMO

Aim To investigate the effect of miR-199a-5p on the proliferation and migration of human glioma cells. Methods U251 cells were selected as experimental subjects to construct a U251 cell line overexpressing miR-199a-5p. The experiment was divided into; control group (U251 cells without transfection, Control), negative control group (transfected with empty vector plasmid U251 cells, NC) and experimental group (transfected with miR-199a-5p mature mimics, mimics). Real-time fluorescent quantitative PCR was used to detect the expression of miR-199a-5p in each group; CCK-8 was used to detect the proliferation of cells transfected with miR-199a-5p; the cell scratch test and Transwell migration test were used to detect the migration of U251 in each group; Western blot was applied to detect DDR1 expression; a U251 cell line overexpressing DDR1 was constructed to detect the effect of overexpression of DDR1 on the proliferation and migration of U251 cells transfected with miR-199a-5p. Results The level of miR-199a-5p in mimics group was significantly higher than that in control group (P < 0.01), the cell viability was reduced (P < 0.01), and the proliferation ability was weakened (P <0. 01). The expression of DDR1 in miR-199a-5p group cells was significantly reduced (P < 0. 01). Compared with mimincs group, the pcDNA3. 1-DDR1 transfected group could up-regulate DDR1 (P < 0. 01), increase cell viability, and promote cell proliferation (P < 0. 05 or P < 0. 01). Conclusions miR-199a-5p can down-regulate the expression of DDR1 and inhibit the proliferation and migration of human glioma cells.

11.
Artigo em Chinês | WPRIM | ID: wpr-825118

RESUMO

@#[Abstract] Objective: To explore whether gambogic acid can enhance the sensitivity of glioma U251 cells to temozolomide and further explore its mechanism. Methods: U251 cells were cultured in vitro and divided into blank control group, gambogic acid treatment group, temozolomide alone treatment group and combined treatment group. The cells survival rates of cells in each group was detected by CCK-8 assay. Flow cytometry was used to detect cell apoptosis and changes in ROS level. Western blotting was used to detect the changes in protein expressions. Results: CCK-8 results showed that the cells survival rates of the four groups after treatment for 24 h were (98.65±3.68)%, (93.58±2.47)%, (66.81±2.39)% and (38.65±4.13)%, respectively. It can be seen that the combined treatment could significantly increase the inhibitory effect of temozolomide on U251 cells. The proportion of apoptotic U251 cells in the combined treatment group was (61.43±2.58)%, which was significantly higher than that of (26.68±1.82)% in the temozolomide-treated group. Combined treatment of gambogic acid and temozolomide could up-regulate ROS level in U251 cells, reduce the expressions of GLUT-3 and p-AKT, and inhibit the GLUT-3/AKT signaling pathway. Conclusion: Gambogic acid combined with temozolomide can enhance the sensitivity of U251 cells to temozolomide by up-regulating ROS level and inhibiting GLUT-3/AKT signaling pathway in U251 cells, and provides a theoretical basis for the application of gambogic acid in the treatment of glioma.

12.
Artigo em Chinês | WPRIM | ID: wpr-829330

RESUMO

@#[Abstract] Objective: To investigate the miR-423-5p expression in brain glioma tissues and cell lines, and its promotive effect on temozolomide (TMZ) chemoresistance by targeting PDCD5 (programmed cell death protein 5). Methods: Tumor tissues and matched peritumoral tissues were collected from 20 brain glioma patients who were surgically treated in the Department of Neurosurgery, Affiliated Hospital of Beihua University between January 2017 and December 2018. Glioblastoma cell lines (U251, U87, SHG-44) and human normal glial cell line HMC-3 were also used in the study. The relative expression of miR-423-5p and PDCD5 in brain glioma and peritumoral tissues and cell lines was detected by qPCR. The synthesized miR-423-5p mimics and miR-NC were respectively transfected into U251 and U87 cells; meanwhile, TMZ at different concentrations (50, 100, 150 and 200 μmol/L) were also used to treat the cells. Then, the chemoresistance of cells to TMZ were determined. MTT assay and colony formation assay were used to examine the proliferation of U251 and U87 cells, andWestern blotting was used to detect the expression of c-caspase 3, Bcl-2 and PDCD5 proteins in U251 and U87 cells. The targeting relationship between PDCD5 and miR-423-5p was validated through Dual luciferase reporter gene assay. Results: miR-423-5p was highly expressed in glioma tissues and glioma cell lines (all P<0.01). As compared with the miR-NC group, the proliferation and TMZ-chemoresistance of U251 and U87 cells in miR-423-5p mimics group significantly increased (all P<0.01). Dual luciferase reporter gene assay validated that miR-423-5p could bind with PDCD5 3' UTR to suppress the expression of PDCD5. Conclusion: High expression of miR-423-5p enhances the chemoresistance of glioma cells to TMZ, and miR-423-5p may serve as a potential therapeutic target in the treatment of brain glioma.

13.
Artigo em Chinês | WPRIM | ID: wpr-793342

RESUMO

@# Objective: To explore the role of tumor suppressor gene programmed cell death 5 gene (PDCD5) in the growth and temozolomide (TMZ) sensitivity of brain glioma cells. Methods:Atotal of 116 patients with cerebral glioma admitted to the Department of Neurosurgery, First Clinical Hospital of Jilin University from January 2009 to December 2014 were enrolled in this study. QPCR, WB and immunohistochemistry method were used to detect the mRNAand protein expressions of PDCD5 in glioma cell lines (U87, U251), U87 cell line with stable PDCD5 expression (U87-PDCD5), glioma cells with si-PDCD5 transfection and primary cerebral glioma tissues, respectively. MTT assay was used to detect the effect of over-expression or knockdown of PDCD5 on the growth and TMZ-sensitivity of glioma cells. The subcutaneous tumor-bearing model of glioma cell line U87 was established in nude mice, and then the experimental mice were randomly divided into control group, TMZ group, PDCD5 group and TMZ+exogenous PDCD5 recombinant expression vector group.After 20 days, the animals were sacrificed by cervical dislocation and the tumor tissue was excised to measure the tumor volume and weigh. The expression of PDCD5 in tumor tissues was detected by qPCR and WB methods, and the effects of PDCD5 combined with TMZ on the growth of gliomas were also analyzed. Results: The relative mRNA and protein expressions of PDCD5 in U87 cells were significantly lower than those in U251 cells (both P<0.05), and the mRNA and protein expressions of PDCD5 in high level glioma tissues were significantly lower than those in low level tissues (all P<0.05). The sensitivity of U87-PDCD5 cells and U251 cells to TMZ was higher than that of U87 cells (all P<0.05). The sensitivity of cells to TMZ in U87-PDCD5-siRNA group and U251siRNA group was significantly lower than that of the control group (both P<0.05). The tumor volume and weigh to fnudemicexenografts were compared,and the results showed control group>TMZ group>PDCD 5group>combined group(allP<0.05);however, the mRNA and protein expressions of PDCD5 in the transplanted tumor tissues of each group showed the opposite trend (all P<0.05). Conclusion: PDCD5 over-expression can enhance the chemosensitivity of braingliomato the chemotherapy drug TMZ, while silencing of PDCD5 expression exertsthe opposite effect.The combination of PDCD5 and TMZ can better inhibit the growth of xenografts in nude mice.

14.
Chinese Pharmacological Bulletin ; (12): 1399-1405, 2019.
Artigo em Chinês | WPRIM | ID: wpr-857126

RESUMO

Aim To prepare curcumin-PLGA nanoparticles [curcumin poly(lactic-co-glycolic acid) nanoparticles, Cur-PLGA-NPs ] and evaluate their effects on glioma cells in vitro. Methods C6 cells and U251 cells were subcultured and randomly divided into control group, curcumin group and curcumin-PLGA group. The morphology of Cur-PLGA-NPs was observed by inverted fluorescence microscope and transmission electron microscopy. The particle size of Cur-PLGA-NPs was detected by Malvem particle size potentiometer. The encapsulation efficiency and drug loading rate of Cur-PLGA-NPs were detected by a multi-function microplate reader. The uptake of Cur-PLGA-NPs by C6 cells and U251 cells was observed by fluorescence microscopy. The effect of Cur-PLGA-NPs on the viability of C6 cells and U251 cells was detected by CCK-8. The effect of Cur-PLGA-NPs on apoptosis was assessed by flow cytometry. Apoptosis was detected by Hoechst staining. Results The Cur-PLGA-NPs had an average particle size of (284. 6 ± 9. 0) nm and were spherical. The encapsulation efficiency was (70.712 ±2.615)%, and the drug loading rate was (2.828 ±0.105) %. Compared with control group and curcumin group, C6 cells and U251 cells significantly increased the uptake of Cur-PLGA-NPs (P < 0. 05). Compared with control group and curcumin group, the cell viability of C6 cells and U251 cells in curcumin-PLGA group significantly decreased (P < 0. 05). Compared with control group and curcumin group, the Cur-PLGA-NPs induced apoptosis significantly in C6 cells and U251 cells(P <0. 05). Conclusion Compared with curcumin, Cur-PLGA-NPs can enhance the uptake of drugs by tumor cells, promoting tumor cell apoptosis.

15.
Chinese Journal of Neuromedicine ; (12): 885-895, 2019.
Artigo em Chinês | WPRIM | ID: wpr-1035086

RESUMO

Objective To study the influence of micro (miR)-325 in progression of glioma and its molecular mechanism by regulating transferrin receptor (TFRC) gene expression in glioma cells. Methods (1) Thirty-five glioma tissues and paired adjacent normal tissues were collected during surgical excision performed in our hospital from January 2015 to January 2018. The miR-325 and TFRC mRNA expression levels in the glioma tissues and paired adjacent normal tissues were detected by inverse transcription-quantitative PCR (RT-qPCR); the expression of miR-325 in glioma tissues of patients with different clinical characteristics and the survival curves of patients with low or high miR-325 expressions were compared. (2) RT-qPCR was used to examine the miR-325 expression in HA, U251, and U87 cell lines in vitro; the regulatory relations between miR-325 and its potential target gene TFRC in U251, and U87 cell lines were measured by luciferase report assay; miR-325 mimic and its negative control were transfected into U251 and U87 cell lines for 48 h, and then, the mRNA and protein expressions of TFRC were detected by RT-qPCR and Western blotting, respectively; control small interfering RNA (siRNA)+nonsense inhibitor, TFRC siRNA+nonsense inhibitor, and siTFRC+miR-325 inhibitor were transfected into U251 and U87 cell lines for 48 h, respectively, Western blotting was employed to detect the TFRC protein expression, cell proliferation was detected by CCK-8 assay, and cell invasion was detected by Transwell assay; pcDNA3.1 empty vector+nonsense sequence, TFRC pcDNA3. 1+nonsense sequence, TFRC pcDNA3.1+miR-325 mimic were transfected into U251 and U87 cell lines for 48 h, respectively, TFRC protein expression was detected by Western blotting, cell proliferation was detected by CCK-8 assay, and cell invasion was detected by Transwell assay. Results (1) As compared with those in the adjacent tissues, the miR-325 expression was significantly decreased and the TFRC mRNA expression was statistically increased in glioma tissues (P<0.05); the TFRC mRNA expression and miR-325 expression were negatively correlated in glioma tissues (P<0.05); as compared with patients with Karnofsky functional status scores≥80, patients with scores<80 had significantly decreased miR-325 expression; as compared with glioma tissues of WHO grading I-II, glioma tissues of grading III-IV had significantly decreased miR-325 expression (P<0.05); the survival rate of patients with low miR-325 expression was statistically lower than that of patients with high miR-325 expression (P< 0.05). (2) As compared with that in HA cells, the miR-325 expression was statistically down-regulated in U87 and U251 cells (P<0.05); in TFRC wild-type (TFRC WT) transfected cells, the miR-325 mimic group had significantly lower luciferase activity than the nonsense sequence group, while the miR-325 inhibitor group had significantly higher luciferase activity than the nonsense inhibitor group (P<0.05); as compared with those in the nonsense sequence group, the TFRC mRNA and protein expressions were statistically decreased in U87 and U251 cells of miR-325 mimic group; as compared with those in the control siRNA+nonsense inhibitor group, the TFRC protein expression and absorbance value were significantly decreased, and number of invasive cells was significantly smaller in the siTFRC+nonsense inhibitor group; and as compared with those in the siTFRC+nonsense inhibitor group, the TFRC protein expression and absorbance value were significantly increased, and number of invasive cells was significantly larger in the siTFRC+miR-325 inhibitor group (P<0.05); as compared with the pcDNA3.1 empty vector+nonsense sequence group, the TFRC protein expression and absorbance value were significantly increased, and number of invasive cells was significantly larger in the TFRC pcDNA3.1 +nonsense sequence group, and as compared with the TFRC pcDNA3.1+nonsense sequence group, the TFRC protein expression and absorbance value were significantly decreased, and number of invasive cells was significantly smaller in the TFRC pcDNA3.1+miR-325 mimic group (P<0.05). Conclusion The miR-325 expression is decreased in glioma cells and has a tumor suppressor effect; patients with low miR-325 expression have poor prognosis; miR-325 inhibits cancer cell progression by inhibiting the expression of the target gene TFRC.

16.
Artigo em Chinês | WPRIM | ID: wpr-700765

RESUMO

Objective Cell cycle-associated protein 1 (Caprin-1) is closely related to the development and progression of cancer. This study aimed to explore the expression of Caprin-1 in the clinical glioma specimen and its influence on the biological char-acteristics of the glioma cell line. Methods Brain tissue specimens were collected from 29 glioma patients and 2 normal humans that died of accidental trauma. A stably transfected U251 cell line with overex-pressed Caprin-1 was established,and the U251 cells were transfected with the pEGFP-C1 plasmid (the negative control group), or the pEGFP-C1-Caprin-1 plasmid (the experimental group), or left un-transfected (the blank control group). The expressions of Caprin-1 mRNA and protein in the cells were determined by RT-PCR and Western blot, and the proliferation and migration of the cells exam-ined by scratch test and Transwell assay,respectively. Results The expression of Caprin-1 was upregulated with the increased grade of glioma,145.9±22.0,444.4±110.0,and 1661.0±54.5 in WHO gradeⅡ,Ⅲ,andⅣglioma,respectively,significantly higher than in the normal brain tissue (P<0.05). Both the mRNA and protein expressions of Caprin-1 were remarkably higher in the experimental group (1.70±0.19 and 1.07±0.09) than in the blank control(0.89±0.10 and 0.52±0.04) and negative control(0.98±0.08 and 0.58± 0.03) (P<0.05).The A value was also markedly higher in the former group(2.55±0.14) than in the latter two(1.40±0.06 and 1.35± 0.04) (P<0.01),and so were the count of migrated cells(526.00±42.19 vs 289.00±29.24 and 279.00±32.48,P<0.01) and the ex-pression of CyclinD1 (0.60±0.05 vs 0.13±0.03 and 0.15±0.05, P<0.01). Conclusion The expression of Caprin-1 in the U251 cells was upregulated with the increased WHO grade of glioma,and the overexpression of Caprin-1 accelerated the proliferation and mi-gration of the U251 cells.

17.
Artigo em Chinês | WPRIM | ID: wpr-704086

RESUMO

Objective To clarify the expression of leptin and leptin receptor in normal brain tissues and gliomas and investigate the effect of exogenous leptin on the proliferation,migration and invasion of human glioma U251 cell line.Methods Immunohistochemical staining was used to detect the expression of leptin and leptin receptor in 50 cases of different grades of glioma tissues and 20 cases of normal brain tissues.The effects of exogenous leptin on proliferation,migration and invasion of U251 cells were detected by MTT assay,cell scratch assay and Transwell invasion assay.Results (1) The positive expression rates of leptin and leptin receptors in glioma tissues were 50.0% and 92.0%,respectively.(2)Proliferation activity:leptin concentrations of 0 ng/ml,10 ng/ml,and 50 ng/ml had no significant difference in the proliferation of U251 cells (absorbance:0.263±0.015,0.273±0.017 and 0.277±0.006,respectively),and the leptin concentration of 100 ng/ml had a significant effect on the proliferation of U251 cells (absorbance:0.315±0.005,P<0.05).(3)Migration ability:the migration rate of U251 cells treated with different concentrations of leptin increased significantly with the passage of time,and the migration rate was most significant at the concentration of 100 ng/ml ((93.313±3.080) %),and the difference was statistically significant (P<0.05).(4)Invasive ability:with the increase of leptin concentration and the prolongation of the action time,the invasive ability of U251 cells was enhanced.When leptin was used at a concentration of 100 ng/ml,the number of penetrating cells were the biggest(135±2).Conclusion Leptin and leptin receptors are involved in the occurrence of gliomas;and exogenous leptin promotes the proliferation of U251 cells and has time and dose dependability on the migration and invasion of U251 cells.

18.
Artigo em Chinês | WPRIM | ID: wpr-732754

RESUMO

Objective To investigate the autophagy and related mechanisms of human glioma U251 cells induced by temozolomide (TMZ) combined with radiotherapy.Methods MTT assay was used to detect the inhibitory effect of TMZ treatments with different concentrations (0~64 μ mol/L) on U251 cells for 24 h and 48 h respectively.The clone formation assay was used to detect the survival fraction of U251 cells under different concentrations of TMZ combined with radiotherapy,and the radiosensitization effect of TMZ was evaluated.The effect of the combined treatment on the apoptosis of U251 cells was detected by flow cytometry.The expression of microtubule-associated protein 1 light chain 3 (LC3)-Ⅰ,LC3-Ⅱ,Beclin-1 and phosphated protein kinase B (p-Akt) was detected by Western Blot.Results The inhibitory effect of TMZ on the proliferation of U251 cells was concentration-and time-dependent,and the IC50 values were 42.25 μmol/L and 25.13 μmol/L,respectively for the 24 h and 48 h treatment.TMZ has a radiosensitizing effect on U251 cells.TMZ combined with radiotherapy can induced apoptosis of U251 cells,and significantly up-regulate the expression levels of LC3-Ⅰ,LC3-Ⅰ and Beclin-1,and the differences were statistically significant (all P<0.01).This combined treatment can down-regulate the expression of p-Akt protein,and the difference was statistically significant (P<0.05).Conclusions TMZ combined with radiotherapy can activate autophagy on U251 cells.The mechanism may involve the inhibition of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway by inhibiting Akt phosphorylation,the promotion of the expression of autophagy marker proteins (LC3-Ⅱ,LC3-Ⅰ and Beclin-1),and subsequent autophagy activation playing an anti-tumor effects.

19.
Zhongcaoyao ; Zhongcaoyao;(24): 2081-2086, 2017.
Artigo em Chinês | WPRIM | ID: wpr-852787

RESUMO

Objective: To investigate the effect of solanine on growth and proliferation of U251 cells. Methods: U251 cells were cultured with different concentration of solanine together. CCK-8 method, wound healing assay, Transwell method, Hochest assay, flow cytometry screening and Western blotting were used to detect inhibitory rate, migration and invasive rates, apoptosis rate and expression level of cell apoptosis-related proteins. Results: CCK-8 assay showed the median inhibition concentration (IC50) of 48 h was 20.05 μg/μL. It was effective that solanine in the concentration range of 5-35 μg/μL could inhibit the growth and proliferation of U251 cells, and played a role in a dose-dependent manner (P < 0.05). Compared with control group, experimental groups can not only inhibit U251 cell migration and invasion in a dose-dependent manner (P < 0.05), but also appear some significant apoptosis characteristics with a concentration dependence. Western blotting assayed that the expression of Bax protein was upregulated, whereas Bcl-2 protein expression was downregulated in experimental groups compared with control group and these changes were dose-dependent. Conclusion: Solanine can inhibit growth and proliferation of U251 cells in effective concentration and in a dose-dependent manner. Solanine can inhibit the invasion and migration of U251 cells. Solanine can induce the apoptosis of U251 cells, and inhibit the proliferation and growth of U251 cells via regulating the expression of Bax and Bcl-2 proteins and affecting Bcl-2/Bax ratio.

20.
China Pharmacy ; (12): 2176-2179, 2017.
Artigo em Chinês | WPRIM | ID: wpr-612348

RESUMO

OBJECTIVE:To evaluate the effects of 3 kinds of serum containing blood-activating and stasis-eliminating TCM compound formulas on aggressive behavior,Janus kinase (JAK)/signal transduction and transcriptional activator (STAT) pathway of glioma U251 cells. METHODS:Rats were randomly divided into normal saline group (5 mL/kg),Taohong Siwu decoction group(5.7 g/kg),Xuefu Zhuyu decoction(8.5 g/kg)and Didang decoction(2.8 g/kg),calculated by crude drug,intragastrically administrated once a day,for 10 d. 10% drug-containing serum culture medium was prepared after 2 h of last administration. After 10% drug-containing serum culture medium intervening U251 cells for 1 week,Transwell method was conducted to detect the cell invasion rate, Western blot was adopted to detect the metal matrix protease 2 (MMP-2), MMP-9, phosphorylated JAK2 (p-JAK2),phosphorylated STAT3(p-STAT3)protein expression;and real-time fluorescence quantitative polymerase chain reaction method was used to detect MMP-2,MMP-9 mRNA expression. RESULTS:Compared with blank serum,Xuefu Zhuyu decoction drug-containing serum can reduce cell invasion rate (P0.05). CON-CLUSIONS:In the 3 kinds of blood-activating and stasis-eliminating TCM compound formulas,Xuefu Zhuyu decoction shows sig-nificant invasive effect on inhibiting U251 cells;the mechanism may be related to inhibiting the activation of JAK2/STAT3 signal pathway and decreasing MMP-2,MMP-9 gene and protein expressions.

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