Promoted Glioma Cell Apoptosis Research Articles

Let-7b-5p promotes triptolide-induced growth-inhibiting effects in glioma by targeting IGF1R.

Glioma is one of the most common malignancies of the central nervous system. The therapeutic effect has not been satisfactory despite advances in comprehensive treatment techniques. Our previous studies have found that triptolide inhibits glioma proliferation through the ROS/JNK pathway, but in-depth mechanisms need to be explored. Recent studies have confirmed that miRNAs may function as tumor suppressor genes or oncogenes and be involved in cancer development and progression. In this study, we found that let-7b-5p expression levels closely correlated with WHO grades and overall survival in patients in tumor glioma-CGGA-mRNAseq-325, and the upregulation of let-7b-5p can inhibit the proliferation and induce apoptosis of glioma cells. Functionally, upregulation of let-7b-5p increased the inhibitory effect on cell viability and colony formation caused by triptolide and promoted the apoptosis rate of triptolide-treated U251 cells. Conversely, downregulation of let-7b-5p had the opposite effect, indicating that let-7b-5p is a tumor suppressor miRNA in glioma cells. Moreover, target prediction, luciferase reporter assays and functional experiments revealed that IGF1R was a direct target of let-7b-5p. In addition, upregulation of IGF1R reversed the triptolide-regulated inhibition of cell viability but promoted glioma cell apoptosis and activated the ROS/JNK signaling pathway induced by triptolide. The results obtained in vivo experiments substantiated those from the in vitro experiments. In summary, the current study provides evidence that triptolide inhibits the growth of glioma cells by regulating the let-7b-5p-IGF1R-ROS/JNK axis in vitro and in vivo. These findings may provide new ideas and potential targets for molecularly targeted therapies for comprehensive glioma treatment.

USP7 Enables Immune Escape of Glioma Cells by Regulating PD-L1 Expression

ABSTRACT Background Glioma is the most common primary intracranial tumor. This study investigated the mechanism of ubiquitin-specific processing protease 7 (USP7) on the immune escape of glioma cells via the regulation of programmed cell death ligand-1 (PD-L1). Methods USP7 and PD-L1 expressions in glioma and normal brain tissues were detected using quantitative reverse transcriptase polymerase chain reaction and Western blot. The glioma cells U-251 MG were transfected with si-USP7 and pcDNA3.1-PD-L1, or treated with anti-PD-L1, after which the cell viability, colony-forming ability, and apoptosis rate were evaluated using cell counting kit-8, colony formation, and TdT-mediated dUTP nick-end labeling assays. Then, CD8+ T cells were purified, extracted, then co-cultured with U-251 MG cells. CD8+ T cell proliferation and the concentrations of interferon (IFN-γ), tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β, interleukin (IL)-1β, and IL-10 were verified using carboxyfluorescein succinimidyl amino ester proliferation assay and enzyme-linked immunosorbent assay. Afterwards, the interaction between USP7 and PD-L1 and the ubiquitination level of PD-L1 were also assessed. Results USP7 was highly-expressed and PD-L1 mRNA levels did not change, while PD-L1 protein levels were up-regulated in the glioma cells. Silencing USP7 in U-251 MG cells limited the growth of the glioma cells, promoted glioma cell apoptosis, and facilitated the proliferation of CD8+ T cells, thus inhibiting immune escape. USP7 stabilized PD-L1 expression through deubiquitination. PD-L1 overexpression reversed the inhibitory effect of silencing USP7 on the immune escape of glioma cells. Conclusion USP7 stabilized PD-L1 through deubiquitination and accelerated the immune escape of glioma cells.

CPVL promotes glioma progression via STAT1 pathway inhibition through interactions with the BTK/p300 axis.

CPVL (carboxypeptidase, vitellogenic-like) is a serine carboxypeptidase that was first characterized in human macrophages. However, the function of CPVL remains unclear in a variety of tumors. The quantitative PCR (qPCR), Western blotting, and IHC assays were utilized to measure the CPVL expression. CPVL was significantly upregulated in glioma cells and tissues compared with normal cells and tissues, respectively. Moreover, high CPVL expression was correlated with advanced clinical grade and poor prognosis. Silencing of CPVL promoted glioma cell apoptosis, and it inhibited cell proliferation and tumorigenicity in vitro and in vivo. Ingenuity Pathway Analysis (IPA) demonstrated that CPVL silencing activated the IFN-γ/STAT1 signaling pathway, thereby inducing glioma cell apoptosis. Mechanistically, immunopurification, mass spectrometry, IP, and glutathione S-transferase (GST) pull-down experiments elucidated that CPVL physically interacts with Bruton’s tyrosine kinase (BTK) and downregulates the STAT1 phosphorylation through promoting p300-mediated STAT1 acetylation. Our findings reveal the crucial role of CPVL in promoting the progression of glioma through suppressing STAT1 phosphorylation. CPVL might serve as a potential prognostic biomarker and therapeutic target for the treatment of glioma.

The function and mechanism of the JARID2/CCND1 axis in modulating glioma cell growth and sensitivity to temozolomide (TMZ)

ABSTRACT A maximal surgical resection followed by radiotherapy and chemotherapy with temozolomide (TMZ) as the representative agent is the standard therapy for gliomas. However, tumor cell resistance to radiotherapy and chemotherapy leads to poor prognosis and high mortality in patients with glioma. In the present study, we demonstrated that JARID2 was downregulated and CCND1 was upregulated within glioma tissues of different grades and glioma cells. In tissue samples, JARID2 was negatively correlated with CCND1. JARID2 overexpression significantly inhibited glioma cell viability, promoted glioma cell apoptosis upon TMZ treatment, and increased p21, cleaved-PARP, and cleaved-caspase3 in TMZ-treated glioma cells. JASPAR tool predicted the possible binding sites between JARID2 and CCND1 promoter regions; through direct binding to CCND1 promoter region, JARID2 negatively regulated CCND1 expression. Under TMZ treatment, JARID2 overexpression inhibited CCND1 expression, promoted glioma cell apoptosis, and increased p21, cleaved-PARP, and cleaved-caspase3 in glioma cells treated with TMZ; meanwhile, CCND1 overexpression exerted opposite effects on glioma cells treated with TMZ and partially reversed the effects of JARID2 overexpression. In conclusion, JARID2 targets and inhibits CCND1. The JARID2/CCND1 axis modulates glioma cell growth and glioma cell sensitivity to TMZ.

Hsa_circ_0001017 inhibits proliferation and metastasis via regulating the let-7g-3p/NDST3 axis in glioma

Glioma is one of the common types of intracranial malignancies, which seriously threatens human health. Circular RNAs (circRNAs) play critical roles in various tumours. This study aims to observe the functions and mechanism of circ_0001017 in glioma progression. Hsa_circ_0001017 expression was analysed in glioma tissue and cells using qRT-PCR assay. Flow cytometer, Cell Counting Kit-8 (CCK-8), colony formation, wound healing, and Transwell were used to analyse glioma cell apoptosis, viability, colony-forming ability, migration, and invasion. The expression of N-deacetylase and N-sulfotransferase 3 (NDST3) and epithelial-mesenchymal transition (EMT)-related proteins was measured using western blot analysis. Luciferase reporter and RNA immunoprecipitation (RIP) assay were performed to determine the target relationship. In glioma tissues and cells, hsa_circ_0001017 expression was decreased. The overexpression of hsa_circ_0001017 inhibited glioma cell proliferation, EMT, migration, and invasion and promoted glioma cell apoptosis, while the knockdown of hsa_circ_0001017 caused the opposite results. Mechanistically, hsa_circ_0001017 sponged hsa-let-7g-3p and NDST3 was the target gene of hsa-let-7g-3p. Moreover, the tumour-suppressive role of circ_0001017 was associated with hsa-let-7g-3p and NDST3. Hsa_circ_0001017 suppressed the growth and metastasis and increased cell apoptosis of glioma, and this effect was associated with hsa-let-7g-3p/NDST3 axis.

Inhibition of Glioma Cell Growth and Apoptosis Induction through Targeting Wnt10B Expression by Pyrazolo[4,3-c]pyridine-4-one.

BackgroundGliomas are commonly diagnosed tumors in the central nervous system that have an elevated mortality rate. The present study evaluated pyrazolo[4,3-c]pyridine-4-one (PP-4-one) as an anti-proliferative agent against glioma cells and investigated the associated mechanism.Material/MethodsThe changes in cell growth were analyzed by Cell Counting Kit-8 (CCK-8) and apoptosis by flow cytometry using Annexin V-FITC staining kit. The FACSCalibur flow cytometer was used for analysis of DNA content and western blotting for protein expression.ResultsThe PP-4-one treatment suppressed viability of U251, C6, and U87 cells significantly at a concentration of 0.25 μM. At a concentration of 16 μM, PP-4-one treatment for 72 hours suppressed viability of U251, C6, and U87 cells to 24%, 21%, and 20%, respectively. Treatment with PP-4-one suppressed cyclic 3′,5′-adenosine monophosphate (cAMP) levels in U251 and C6 cells significantly (P<0.05) depending on the concentration. The apoptotic cells were increased significantly (P<0.05) by PP-4-one treatment in U251 and C6 cell cultures. A considerable enhancement in the proportion of U251 and C6 cells in the G0/G1 phase was recorded on incubation with PP-4-one. Treatment of U251 and C6 cells with PP-4-one markedly enhanced p21 expression relative to the control. The B-cell lymphoma (Bcl-2) level in PP-4-one treated U251 and C6 cells was markedly lower relative to the control cells. The Bax, caspase-3, and caspase-9 levels were elevated markedly by PP-4-one treatment in U251 and C6 cells.ConclusionsThis study demonstrated that PP-4-one has anti-proliferative potential for glioma cells via targeting cAMP and Bcl-2 levels. It also promoted glioma cell apoptosis through caspase activation and arrest of the cell cycle. Thus, PP-4-one may be used to develop drug candidates for the glioma treatment.

MiR-483 Targeted SOX3 to Suppress Glioma Cell Migration, Invasion and Promote Cell Apoptosis.

ObjectiveGlioma is the most common malignant brain tumor that has high aggressiveness. The aim of this study was to investigate the potential therapeutic targets for gliomas.Materials and MethodsReal-time quantitative polymerase chain reaction (RT-qPCR) was employed to calculate the expression of miRNA and genes. The connection between the expression of miR-483 and patients’ overall survival rate was evaluated using Kaplan–Meier analysis. In addition, the underlying mechanism was detected using luciferase assay.ResultsThe expression level of miR-483 was significantly decreased in glioma tissue samples and cell lines, compared to the adjacent tissues and normal cell lines. Downregulation of miR-483 or upregulation of SOX3 was associated with overall survival of glioma patients. Additionally, overexpression of miR-483 promotes cell invasion and migration and inhibits apoptosis. In addition, miR-483 directly targeted to SOX3, and the expression of miR-483 has a negative correlation with SOX3 in glioma tissues. SOX3 reversed partial functions of miR-483 on cell migration, invasion, and promoted cell apoptosis in glioma.ConclusionMiR-483 inhibited glioma cell migration, invasion, and promoted glioma cell apoptosis by targeting SOX3. MiR-483 maybe acted as a potential target for the treatment of glioma.

LncRNA-135528 inhibits tumor progression by up-regulating CXCL10 through the JAK/STAT pathway.

Spontaneous tumor regression can be observed in many tumors, however, studies related to the altered expression of lncRNA in spontaneous glioma regression are limited, and the potential contributions of lncRNAs to spontaneous glioma regression remain unknown. To investigate the biological roles of lncRNA-135528 in spontaneous glioma regression. The cDNA fragment of lncRNA-135528 was obtained by rapid-amplification of cDNA ends (RACE) technology and cloned into the plvx-mcmv-zsgreen-puro vector. Additionally, we stably silenced or overexpressed lncRNA-135528 in G422 cells by transfecting with siRNA against lncRNA-135528 or lncRNA-135528 overexpression plasmid. Then, we examined lncRNA-135528 overexpressing and lncRNA-135528 silencing on glioma cells and its effects on CXCL10 and JAK/STAT pathways. The main findings indicated that lncRNA-135528 promoted glioma cell apoptosis, inhibited cell proliferation and arrested cell cycle progression; the up-regulation of lncRNA135528 led to significantly increased CXCL10 levels and the differential expression of mRNA associated with JAK/STAT pathway in glioma cells. lncRNA-135528 can inhibit tumor progression by up-regulating CXCL10 through the JAK/STAT pathway.

MiR-152 regulated glioma cell proliferation and apoptosis via Runx2 mediated by DNMT1

BackgroundAberrant DNA methylation is associated with tumor onset and progression. Study has verified that the DNA methylation of miR-152 was mediated in many tumors, but whether it involved in glioblastomas was still unclear. MethodsThis study enrolled 20 patients with glioma to analyze the expression pattern of miR-152. Real-time PCR and western blot were used to detect the mRNA or protein expression level, respectively. The relationship between miR-152 and runx2 was detected by Luciferase reporter assay. The methylation level of miR-152 was determined by methylation-specific PCR. Cell proliferation and apoptosis were detected by MTT and Annexin-FITC/PI assay. ResultsThe expression of miR-152 was down-regulated while the expression of DNMT1 was up-regulated in both glioma tissue and cell lines. MiR-152 was hypermethylated and its expression was negatively correlated with DNMT in glioma cell lines. DNMT1 knockdown promoted the expression of miR-152, however, DNMT1 overexpression suppressed the expression of miR-152. MiR-152 overexpression promoted glioma cell apoptosis while miR-152 knockdown promoted cell proliferation. MiR-152 targets Runx2 to regulate its expression, Runx2 overexpression abolished the effects of miR-152 overexpression. ConclusionMiR-152 regulated cell proliferation and apoptosis of glioma mediated by Runx2, while the mechanism of down regulated miR-152 in glioma tissues and cells was its hypermethylation.

MicroRNA-140-5p inhibits cell proliferation and invasion by regulating VEGFA/MMP2 signaling in glioma

Glioma is the most common primary malignant tumor of the central nervous system, which results in both a poor prognosis and outcome because of the aggressive progression of disease, growth and resistance to surgery, chemotherapy, and radiotherapy. MiR-140-5p is a small, non-coding single-stranded RNA molecule, which was previously studied in the settings of human tongue cancer, hepatocellular carcinoma, and colorectal cancer. However, detailed data that formally demonstrate the contribution of miR-140-5p to glioma development are missing. Similarly, relatively little is known about the relationship of miR-140-5p, vascular endothelial growth factor A, and matrix metalloproteinase-2 in glioma progression. In this study, we found that miR-140-5p expression was significantly decreased in glioma tissues and in the glioma cell-lines U87 and U251 as compared with non-cancerous brain tissues by quantitative real-time polymerase chain reaction. In addition, miR-140-5p inhibited glioma cell proliferation and invasion and promoted glioma cell apoptosis both in vivo and in vitro. Interestingly, while the expression levels of miR-140-5p were higher in glioma cells, the messenger RNA or protein expression levels of vascular endothelial growth factor A and matrix metalloproteinase-2 were lower in glioma cells as determined by quantitative real-time polymerase chain reaction, western blot assay, and immunohistochemistry. By contrast, downregulation in the expression levels of miR-140-5p augmented the messenger RNA and protein expression levels of both vascular endothelial growth factor A and matrix metalloproteinase-2. These findings suggested that miR-140-5p inhibited glioma proliferation and invasion by regulating the vascular endothelial growth factor A/matrix metalloproteinase-2 signaling pathway both in vitro and in vivo.

54 Anti-glioma activity of Rana temporaria chensinensis egg protein hydrolysate

Objectives Rana temporaria chensinensis only live in the Changbai mountain area in the northeast of China. The eggs of R.temporaria chensinensis contain many special ingredients that modulate incretion. This study aimed to investigate the effects of R.temporaria chensinensis egg protein hydrolysate on human glioma C6 cell proliferation and apoptosis. Methods We extracted R.temporaria chensinensis egg protein (500 mg/mL) and investigated its effects on human glioma C6 cell cultures, and subjected it to an MTT assay, colony forming assay, Western blot assay and flow cytometry analysis of apoptosis. We further investigated the effects of R. temporaria chensinensis egg protein hydrolysate (1.5 g/kg) on glioma development and progression in vivo using a mouse model of glioma. Results R.temporaria chensinensis egg protein hydrolysate inhibited the proliferation of glioma cells; these effects were mediated by the phosphoinositide 3-kinase (PI3K)/AKT signalling pathway. Conclusions This study suggested that R.temporaria chensinensis egg protein hydrolysate promotes apoptosis of glioma cells both in vitro and in vivo. Acknowledgments This research was financially supported by Jilin Province Science and Technology Development Program (Grant Nos. 20150309008YY and 20150309009YY).

The diagnostic value and functional roles of phosphoglycerate mutase 1 in glioma.

Previous studies indicated that phosphoglycerate mutase 1 (PGAM1) is involved in many cancer types and promotes breast cancer progression. However, the role of PGAM1 in glioma remains unclear. The present study aimed to investigate the association of PGAM1 expression with glioma grade and the role of PGAM1 in proliferation, apoptosis, migration and invasion of glioma cells. The mRNA and protein expression of PGAM1 was analysed in glioma tissues and normal brain tissues. The expression of PGAM1 was examined further by immunohistochemical analysis. In addition, we inhibited the expression of PGAM1 in glioma cell line by siRNA to evaluate its role in glioma proliferation, apoptosis, migration and invasion. The mRNA and protein expression of PGAM1 was significantly greater in glioma than normal brain tissues. PGAM1 expression was associated with the WHO grade of glioma. siRNA knockdown of PGAM1 significantly inhibited glioma cell proliferation, promoted glioma cell apoptosis, induced S phase cell cycle arrest and inhibited glioma cell migration and invasion invitro. PGAM1 may be associated with the grade of glioma and be involved in the biological behavior of glioma cells. PGAM1 might be a novel therapeutic target in glioma.

Silibinin-induced glioma cell apoptosis by PI3K-mediated but Akt-independent downregulation of FoxM1 expression

The oncogenic transcription factor Forkhead box M1 (FoxM1) is overexpressed in many human tumors, including glioma. As a critical regulator of the cell cycle and apoptosis-related genes, FoxM1 is a potential therapeutic target against human malignant glioma. Silibinin, a flavonoid isolated from Silybum marianum, dose-dependently reduced glioma cell proliferation, promoted apoptosis, and downregulated FoxM1 expression. Knockdown of FoxM1 by small hairpin RNA (shRNA) transfection also promoted glioma cell apoptosis and augmented the antiproliferative and pro-apoptotic properties of silibinin. Moreover, silibinin increased caspase-3 activation, upregulated pro-apoptotic Bax, and suppressed anti-apoptotic Bcl-2 expression, effects enhanced by FoxM1 knockdown. Silibinin treatment suppressed U87 cell PI3K phospho-activation, and simultaneous silibinin exposure, FoxM1 knockdown, and PI3K inhibition additively increased U87 cell apoptosis. Furthermore, PI3K inhibition reduced FoxM1 expression. Akt activity was also suppressed by FoxM1 downregulation but Akt inhibition did not alter FoxM1 expression. Thus, silibinin likely inhibited glioma cell proliferation and induced apoptosis through inactivation of PI3K and FoxM1, leading to activation of the mitochondrial apoptotic pathway. FoxM1 may be a novel target for chemotherapy against human glioma.

MiR-92b inhibitor promoted glioma cell apoptosis via targeting DKK3 and blocking the Wnt/beta-catenin signaling pathway

BackgroundMiR-92b was upregulated in gliomas. However, the association of miR-92b with glioma cell apoptosis and survival remains unknown.MethodsProliferation capability of glioma cells upon tranfection with miR-92b mimics or inhibitors was detected by mutiple analyses, including MTT assays, colony formation assay. Apoptosis abilities of glioma cells were detected by flow cytometric analysis. The target of miR-92b was determined by luciferase reporter and western blot. The association of miR-92b with outcome was examined in twenty glioma patients.ResultsMiR-92b expression was significantly increased in high-grade gliomas compared with low-grade gliomas, and positively correlated with the degree of glioma infiltration. Over-expression of miR-92b increased cell proliferation, whereas knockdown of miR-92b decreased cell proliferation via modulating the levels of the target, Target prediction analysis and a dual luciferase reporting assay confirmed that the inhibitory protein-coding Dickkopf-3 gene (DKK3) was a direct target of miR-92b. Furthermore, miR-92b could regulate the expression of downstream genes of the Wnt/beta-catenin signaling pathway, such as Bcl2, c-myc and p-c-Jun, in glioma cells. Finally, the increased level of miR-92b expression in high-grade gliomas confers poorer overall survival.ConclusionsThe present data indicates that miR-92b directly regulate cell proliferation and apoptosis by targeting DKK3 and act as prognostic factors for glioma patients.

The repressive effect of microRNA-25 inhibitor on human glioma cell growth

Objective To observe the effect of microRNA (miR-25) inhibitor on glioma cell growth.Methods The miR-25 inhibitor was transfected by liposome. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect miR-25 expression after transfection.The cell cycle kinetics and cell proliferation rate were examined by using flow cytometry and methyl thiazol tetrazolium (MTT) assay,the anchorage-independent cell proliferation was evaluated by soft agar assay,the apoptosis rate was tested by Annexin V staining,and the invasive ability was measured by Transwell.Results The antisense oligonucleotide effectively down-regulated the miR-25 expression in glioma cells.Meanwhile,the proportion of cells in S phase was decreased by about 6％-10％,and the proliferation activity was significantly suppressed (P＜0.05 ).Also,the down-regulation of miR-25 promoted glioma cell apoptosis ( P ＜0.05).However,no significant changes were observed in the number of cells getting through matrigel.Conclusion Suppression of miR-25 expression could inhibit the proliferative ability and induce apoptosis of glioma cells,but had no effect on the invasive ability in glioma cells.This preliminary finding suggests that miR-25 may play an oncogenic role in glioma cell growth,but not invasion. Key words: Glioma; MicroRNA-25; Cell growth; Invasion