Golgi Phosphoprotein 3 Inhibits the Apoptosis of Human Glioma Cells in Part by Downregulating N-myc Downstream Regulated Gene 1
Xin LiMengyou LiXiuli TianQingzhe LiQingyang LuQingbin JiaLianqun ZhangJinqiang YanXueyuan LiXingang Li
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BACKGROUND Golgi phosphoprotein 3 (GOLPH3) has been reported to be involved in the development of several human cancers. Our previous study showed that GOLPH3 expression in glioma tissues was related to the severity of the malignancy of the cancer. However, the mechanism by which GOLPH3 affects cell apoptosis is largely unknown. The present study was designed to explore the possible mechanism of GOLPH3 in cell apoptosis. MATERIAL AND METHODS To analyze the biological role of GOLPH3 in glioma cells, we used GOLPH3 small interference RNA in apoptosis of glioma cells. The apoptosis of glioma cells was detected by flow cytometry. The expression level of GOLPH3 and NDRG1 protein was determined by Western blot analyses and immunohistochemical staining, respectively, to evaluate their association with glioma. Tumor tissues were collected from patients with glioma. Normal cerebral tissues were acquired from cerebral trauma patients undergoing internal decompression surgery. RESULTS We confirm that the decrease of GOLPH3 that promotes the apoptosis of glioma cells may be regulated by the activation of NDRG1 and cleaved capcase 3. There was a inverse association between GOLPH3 and NDRG1 in glioma samples. CONCLUSIONS Our findings indicate that GOLPH3 and NDRG1 both play an important role in glioma etiology. Either GOLPH3 or NDRG1 might be a potential candidate for malignant glioma therapy.Objective:To explore the methods of using Flow Cytometry to examine the apoptosis.Methods:Flow Cytometry was used to examine the DNA content,the phosphatidyl serine,the mitcchondrial membrane potential and the calcium ion,the apoptosis was analyzed.Results:The DNA content had some deflexion.The phosphatidyl serine examination can reflect the early and advanced stage apoptosis punctually.The mitcchondrial membrane potential and the calcium ion examination can reflect the physiological index of apoptosis cell.But if combined with morphology the analysis will be more objective.Conclusion:Through comparing empirical method,we find more precise methods of using the Flow Cytometry to examine the apoptosis in the future.
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Objective To investigate the possible role of caspase-3 and bcl-2 played in As_4S_4 induced apoptosis of cervical tumor Hela cell line.Method Hela cells were cultured with various concentrations(7.5,15,30 and 60 mg/L)of As_4S_4 for 12,24,36,48 and 60 h respectively,then measuring the situation of cell growth apoptosis with MTT and flow cytometry analysis.The expressions of(bcl-2) and caspase-3 protein were detected by Western blot,caspase-3 activity was evaluated with flow cytometry.Result The results showed that As_4S_4 might time and dose dependently depress the proliferation of cervical tumor Hela cell line significantly,the IC_(50) for 24 h was 30 mg/L.Flow cytometry revealed after 24 h co-culture the apoptotic rates of Hela cells were(8.13±1.13)%、(29.58±2.51)%、(46.24±3.92)% and(62.36±4.42)%,respectively(P0.01),under different concentrations of As_4S_4(7.5,15,30,60 mg/L),while the apoptotic peak did not appear in control group(P0.01).The expression of bcl-2 protein was also down-regulated;while caspase-3 protein was up-regulated with the increase of dose,(2.67±0.22)%,(9.53±0.15)%,(21.28±0.43)%,(39.63±0.80)% and(63.40±0.69)%,respectively(P0.01).Conclusion As_4S_4 has the effect of depressing proliferation and enhancing apoptosis on Hela cell in vitro,its mechanism may related to down-regulating the expression of bcl-2 protein and activating the activity of caspase-3.
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Abstract Background: Glioma is a type of malignant cancer in the central nervous system. New predictive biomarkers have been investigated in recent years, but the clinical prognosis in glioma remains poor. The function of CPLX2 in glioma and the probable molecular mechanism of tumor suppression was the focus of this investigation. Methods: The glioma transcriptome profile is downloaded from The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA) databases were performed to analyze the expression of CPLX2 in glioma. RT-qPCR was performed to detect the expression of CPLX2 in 68 glioma subjects, these patients who have been followed up. Kaplan-Meier survival analyses were done to evaluate the effect of CPLX2 on the prognosis of glioma patients. The CPLX2 knockdown and overexpressed cell lines were constructed to investigate the effect of CPLX2 on glioma. The cell growth, colony formation, and tumor formation in xenograft were performed. Results: The expression of CPLX2 was downregulated in glioma and negatively correlated to the grade of glioma. The higher expression of CPLX2 predicted a longer survival through the analysis of Kaplan-Meier survival curves. Overexpressed CPLX2 impaired tumorigenesis in glioma progression both in vivo and in vitro . Knocking down of CPLX2 promoted the proliferation of the glioma cells. The analysis of GSEA and co-expression analysis revealed that CPLX2 may affect the malignancy of glioma by regulating hypoxia and inflammation pathway. Conclusions: Our data indicated that CPLX2 functioned as a tumor suppressor and could be used as a potential prognostic marker in glioma.
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Objective: To study the role of p38MAPK in mediating TNF-α-induced apoptosis of rat glioma cell line C6. Methods: Effect of TNF-α on the proliferation of C6 cells was determined by MTT assay. The TNF-α induced apoptosis was detected by transmission electron microscopy and flow cytometry. The expression of p38MAPK was detected by SABC method and Western-blot. The effect of SB202190, a specific inhibitor of p38MAPK, on TNF-α-induced apoptosis was observed by flow cytometry and SABC method. Results: Inhibitory rate of TNF-α(2×105 U/L) on C6 cells was 43. 75% . In the TNF-α treated group, apoptotic cells were observed by transmission electron microscopy and the apoptotic rate was 37. 5% by flow cytometry. p38MAPK positive signals were detected by SABC method and Western-blot. In the SB202190 treated group, the apoptotic rate was 7. 0% and no p38MAPK signals were found. Conclusion: Apoptosis of C6 cells and expression of p38MAPK can be induced by TNF-α. The activation of p38MAPK promotes the apoptosis of C6 cells.
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Purpose:To study the role of p38MAPK in mediating TNF α induced apoptosis in rat glioma cells C6.Methods:The proliferation activity of C6 cells after the treatment by TNF α was observed by MTT assay. The TNF α induced apoptosis was detected by transmission electron microscopy and flow cytometry. The expression of p38MAPK was detected by SABC method and Westernblot. The effect of SB202190, a specific inhibitor of p38MAPK on TNF α induced apoptosis was observed by flow cytometry and SABC method. Results:The inhibitory rate of TNF α (2×10 5 U/L) on C6 cells was 43.75%. In the TNF α treated group, apoptotic cells were observed by transmission electron microscopy and the apoptotic rate was 37.5% by flow cytometry, p38MAPK positive signals were found by SABC method and Westernblot. In the SB202190 treated group, the apoptotic rate was 7.0% and no p38MAPK signals were found.Conclusions:The apoptosis of C6 cells and expression of p38MAPK could be induced by TNF α. The activation of p38MAPK promoted the apoptosis of C6 cells. [
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Objective: To study the role of p38MAPK in mediating TNF-α-induced apoptosis of rat glioma cell line C6. Methods: Effect of TNF-α on the proliferation of C6 cells was determined by MTT assay. The TNF-α induced apoptosis was detected by transmission electron microscopy and flow cytometry. The expression of p38MAPK was detected by SABC method and Western-blot. The effect of SB202190, a specific inhibitor of p38MAPK, on TNF-α-induced apoptosis was observed by flow cytometry and SABC method. Results: Inhibitory rate of TNF-α(2×105 U/L) on C6 cells was 43.75%. In the TNF-α treated group, apoptotic cells were observed by transmission electron microscopy and the apoptotic rate was 37.5% by flow cytometry. p38MAPK positive signals were detected by SABC method and Western-blot. In the SB202190 treated group, the apoptotic rate was 7.0% and no p38MAPK signals were found. Conclusion: Apoptosis of C6 cells and expression of p38MAPK can be induced by TNF-α. The activation of p38MAPK promotes the apoptosis of C6 cells.
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This thesis studied the impact of GPC with different concentrations over the vascular endothelial cells apoptosis by flow cytometry and analyzed the influence of GPC on oxygen free radicals in initiating cell apoptosis. Make use of flow cytometry instrument to detect the apoptosis rate of vascular endothelial cell by adding different dose of GPC separately. Through experiments, the apoptosis rates of normal control group, positive control group, GPC (fifty) group, GPC (one hundred) group, GPC (one hundred and fifty) group, GPC (two hundred) group, separately were 11.89, 12.63, 10.17, 5.45, 4.79 and 4.53%, respectively. The experiment results indicated that GPC possessed distinct inhibitory effect on vascular endothelial cells apoptosis evoked by hydroxyl radical.
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Glioma is a tumor of the brain. Although the clinical regimens and surgical techniques for glioma have improved, therapies of advanced glioma remain challenging, carrying dismal overall survival and therapeutic success rates. Evidence has shown that miRNAs played important roles in glioma development. The current study aimed at investigating the function of a novel cancerogenic miRNA, miR-93, in glioma progression by investigating the expression and mechanism of it.qRT-PCR was conducted to assess the miR-93 expression and the mRNA expression of target gene in glioma tissues and cells. The invasion and migration abilities of the glioma cells were determined by transwell assays. Luciferase reporter assay was performed to confirm the target of miR-93.The results indicated that miR-93 expression in glioma tissues and cells was increased significantly than that in normal brain tissues and cells. Furthermore, miR-93 promoted glioma cell migration and invasion. RBL2 was recognized as a direct target of miR-93 in glioma cells, and overexpression of RBL2 could reverse the stimulative effect of miR-93 in glioma cell.The above findings suggested that miR-93 together with RBL2 could be diagnostic targets and novel prognostic markers for glioma.
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