T98G Cells Research Articles

The treatment effects and the underlying mechanism of B cell translocation gene 1 on the oncogenesis of brain glioma.

Glioma is characterized by cell over-proliferation, aggressive phenotype, and angiogenesis. B cell translocation gene 1 (BTG1) works as a tumor suppressor in various cancer types. We aimed to study the functions of BTG1 in glioma development. We tested the BTG1 expressions in various glioma cell lines. T98G and U87 cells were transfected with siBTG1 and BTG1 expression vector, respectively, then assessing cell viability, cell migration, and invasion abilities. Flow cytometry was performed to analyze apoptosis and cell cycle distribution. The status of angiogenesis was assessed by In vitro angiogenesis assay. Quantitative polymerase chain reaction and Western blot analysis were used for expression analyses. The inhibitor FH535 (20 μM) and agonist LiCl (20 mM) of the Wnt/β-catenin pathway were introduced to treat transfected cells. BTG1 was low-expressed in glioma cell lines. In T98G cell transfected with siBTG1, cell proliferation, migration, invasion, and angiogenesis abilities were notably increased, BTG1 silencing promoted the cell survival and cell cycle progression from G0/G1 to the S phase. In BTG overexpressed U87 cell, cell proliferation, migration, invasion, and angiogenesis abilities were significantly inhibited. In addition, cell apoptosis significantly increased and the number of G0/G1 phase cells was also significantly increased. We also found that the activation of Wnt/β-catenin was significantly inhibited in BTG1 group. LiCl and FH535 treatments could partially reverse the effects of BTG1 on glioma cell viabilities. Our data suggested that BTG1 functions as a tumor suppressor in glioma, and the anticancer ability of BTG1 involves the repression of the Wnt/β-catenin pathway.

Design and Synthesis of a New Soluble Natural β-Carboline Derivative for Preclinical Study by Intravenous Injection.

Harmine is a natural β-carboline compound showing several biological activities, including antiproliferative properties, but this soluble natural molecule lacks selectivity. Harmine derivatives were reported to overcome this problem, but they are usually poorly soluble. Here, we designed and synthesized a new 2, 7, 9-trisubstituted molecule (1-methyl-7-(3-methylbutoxy)-9-propyl-2-[(pyridin-2-yl)methyl]-9H-pyrido[3,4-b]indol-2-ium bromide) with a solubility of 1.87 ± 0.07 mg/mL in a simulated injection vehicle. This compound is stable for at least 72 h in acidic and physiological conditions (pH 1.1 and 7.4) as well as in a simulated injection vehicle (physiological liquid + 0.1% Tween80®). Solubility in those media is 1.06 ± 0.08 mg/mL and 1.62 ± 0.13 mg/mL at pH 7.4 and 1. The synthesized molecule displays a significant activity on five different cancer cell lines (IC50 range from 0.2 to 2 µM on A549, MDA-MB-231, PANC-1, T98G and Hs683 cell lines). This compound is also more active on cancer cells (MDA-MB-231) than on normal cells (MCF-10a) at IC50 concentrations. Due to its high activity at low concentration, such solubility values should be sufficient for further in vivo antitumoral activity evaluation via intravenous injection.

Hyperbaric oxygen increases glioma cell sensitivity to antitumor treatment with a novel isothiourea derivative invitro.

Glioblastoma (GBM) is the most common primary brain tumor. Tumor hypoxia is a pivotal factor responsible for the progression of this malignant glioma, and its resistance to radiation and chemotherapy. Thus, improved tumor tissue oxygenation may promote greater sensitivity to anticancer treatment. Protein kinase D1 (PKD1) protects cells from oxidative stress, and its abnormal activity serves an important role in multiple malignancies. The present study examined the effects of various oxygen conditions on the cytotoxic potential of the novel isothiourea derivate N,N′-dimethyl-S-(2,3,4,5,6-pentabromobenzyl)- isothiouronium bromide (ZKK-3) against the T98G GBM cell line. ZKK-3 was applied at concentrations of 10, 25 and 50 µM, and cells were maintained under conditions of normoxia, anoxia, hypoxia, hyperbaric oxygen (HBO), hypoxia/hypoxia and hypoxia/HBO. The proliferation and viability of neoplastic cells, and protein expression levels of hypoxia-inducible factor 1α (HIF-1α), PKD1, phosphorylated (p)PKD1 (Ser 916) and pPKD1 (Ser 744/748) kinases were evaluated. Oxygen deficiency, particularly regarding hypoxia, could diminish the cytotoxic effect of ZKK-3 at 25 and 50 µM and improve T98G cell survival compared with normoxia. HBO significantly reduced cell proliferation and increased T98G cell sensitivity to ZKK-3 when compared with normoxia. HIF-1α expression levels were increased under hypoxia compared with normoxia and decreased under HBO compared with hypoxia/hypoxia at 0, 10 and 50 µM ZKK-3, suggesting that HBO improved oxygenation of the cells. ZKK-3 exhibited inhibitory activity against pPKD1 (Ser 916) kinase; however, the examined oxygen conditions did not appear to significantly influence the expression of this phosphorylated form in cells treated with the tested compound. Regarding pPKD1 (Ser 744/748), a significant difference in expression was observed only for cells treated with 10 µM ZKK-3 and hypoxia/hypoxia compared with normoxia. However, there were significant differences in the expression levels of both phosphorylated forms of PKD1 under different oxygen conditions in the controls. In conclusion, the combination of isothiourea derivatives and hyperbaric oxygenation appears to be a promising therapeutic approach for malignant glioma treatment.

Ubiquitin-specific protease 4 promotes glioblastoma multiforme via activating ERK pathway.

BackgroundGlioblastoma multiforme (GBM) is one of the most common brain tumors in adults. Current treatments cannot increase survival to a large extent, as the glioblastoma development mechanisms remain unknown. It has been well documented that ubiquitination contributes to tumor initiation and/or progression in many kinds of cancer. Ubiquitin-specific protease 4 (USP4), a member of deubiquitinating enzymes (DUBs) family, can remove ubiquitin residues and play a role in cancer development.MethodsIn the current study, lentiviruses were used to manipulate the expression of USP4. Real-time PCR and Western blot were used to measure the expression level of USP4. Then, CCK-8 and annexin-V staining were used to detect cell proliferation and cell apoptosis, respectively.ResultsFirst, we found that USP4 was highly upregulated in GBM tissues in comparison with that in normal tissues and high level of USP4 correlated with poor prognosis. Moreover, knockdown of USP4 could significantly inhibit cell proliferation and increase cell apoptosis in U87 and T98G cells. Cells with stable USP4 reduction exhibited slower tumor growth rate and smaller tumor size than the control group cells in a xenograft mouse model. Inhibition of USP4 downregulated the expression of PCNA, Bcl-2 and p-ERK1/2, but upregulated the expression of Bax both in vitro and in vivo. Inversely, USP4 overexpression could attenuate the effects contributed by ERK inhibitor. TGF-βR inhibition reduced level of TGF-βR1, p-smad2 and p-ERK1/2 which can partially be rescued by USP4 overexpression.ConclusionUSP4, as a potential novel oncogene, promotes GBM by activation of ERK pathway through regulating TGF-β.

The radiobiological effects of He, C and Ne ions as a function of LET on various glioblastoma cell lines.

The effects of the charged ion species 4He, 12C and 20Ne on glioblastoma multiforme (GBM) T98G, U87 and LN18 cell lines were compared with the effects of 200 kVp X-rays (1.7 keV/μm). These cell lines have different genetic profiles. Individual GBM relative biological effectiveness (RBE) was estimated in two ways: the RBE10 at 10% survival fraction and the RBE2Gy after 2 Gy doses. The linear quadratic model radiosensitivity parameters α and β and the α/β ratio of each ion type were determined as a function of LET. Mono-energetic 4He, 12C and 20Ne ions were generated by the Heavy Ion Medical Accelerator at the National Institute of Radiological Sciences in Chiba, Japan. Colony-formation assays were used to evaluate the survival fractions. The LET of the various ions used ranged from 2.3 to 100 keV/μm (covering the depth–dose plateau region to clinically relevant LET at the Bragg peak). For U87 and LN18, the RBE10 increased with LET and peaked at 85 keV/μm, whereas T98G peaked at 100 keV/μm. All three GBM α parameters peaked at 100 keV/μm. There is a statistically significant difference between the three GBM RBE10 values, except at 100 keV/μm (P < 0.01), and a statistically significant difference between the α values of the GBM cell lines, except at 85 and 100 keV/μm. The biological response varied depending on the GBM cell lines and on the ions used.

Design, Synthesis, and Evaluation of Lipopeptide Conjugates of Mercaptoundecahydrododecaborate for Boron Neutron Capture Therapy.

We developed new 10 B carriers for boron neutron capture therapy (BNCT) that can effectively transport and accumulate boron clusters into cells. These carriers consist of a lipopeptide, mercaptoundecahydrododecaborate (BSH), and a disulfide linker. The carriers were conceived according to the structure of pepducin, a membrane-penetrating lipopeptide targeting protease-activated receptor 1 (PAR1). To improve the membrane permeability of BSH, the structure was optimized using various lipopeptides possessing different peptides and lipid moieties. These synthesized lipopeptides were conjugated with BSH and evaluated for intracellular uptake using T98G glioblastoma cells. Among them, the most effectively incorporated and accumulated in the cells was compound 5 a, which contains a peptide of 13 residues derived from the intracellular third loop of PAR1 and a palmitoyl group. For further improvement of 10 B accumulation in cells, the introduction of an amine linker was investigated; intracellular uptake similar to that of 5 a was observed for compound 14, which has a piperazine linker. Both compounds 5 a and 14 showed a stronger radiosensitizing effect than BSH along on T98G cells under mixed-neutron beam irradiation. The results demonstrate that lipopeptide conjugation is effective for enhancing intracellular delivery and accumulation of BSH and improving the cytotoxic effect of BNCT.

The knocking down of the oncoprotein Golgi phosphoprotein 3 in T98G cells of glioblastoma multiforme disrupts cell migration by affecting focal adhesion dynamics in a focal adhesion kinase-dependent manner.

Golgi phosphoprotein 3 (GOLPH3) is a conserved protein of the Golgi apparatus that in humans has been implicated in tumorigenesis. However, the precise function of GOLPH3 in malignant transformation is still unknown. Nevertheless, clinicopathological data shows that in more than a dozen kinds of cancer, including gliomas, GOLPH3 could be found overexpressed, which correlates with poor prognosis. Experimental data shows that overexpression of GOLPH3 leads to transformation of primary cells and to tumor growth enhancement. Conversely, the knocking down of GOLPH3 in GOLPH3-overexpressing tumor cells reduces tumorigenic features, such as cell proliferation and cell migration and invasion. The cumulative evidence indicate that GOLPH3 is an oncoprotein that promotes tumorigenicity by a mechanism that impact at different levels in different types of cells, including the sorting of Golgi glycosyltransferases, signaling pathways, and the actin cytoskeleton. How GOLPH3 connects mechanistically these processes has not been determined yet. Further studies are important to have a more complete understanding of the role of GOLPH3 as oncoprotein. Given the genetic diversity in cancer, a still outstanding aspect is how in this inherent heterogeneity GOLPH3 could possibly exert its oncogenic function. We have aimed to evaluate the contribution of GOLPH3 overexpression in the malignant phenotype of different types of tumor cells. Here, we analyzed the effect on cell migration that resulted from stable, RNAi-mediated knocking down of GOLPH3 in T98G cells of glioblastoma multiforme, a human glioma cell line with unique features. We found that the reduction of GOLPH3 levels produced dramatic changes in cell morphology, involving rearrangements of the actin cytoskeleton and reduction in the number and dynamics of focal adhesions. These effects correlated with decreased cell migration and invasion due to affected persistence and directionality of cell motility. Moreover, the knocking down of GOLPH3 also caused a reduction in autoactivation of focal adhesion kinase (FAK), a cytoplasmic tyrosine kinase that regulates focal adhesions. Our data support a model in which GOLPH3 in T98G cells promotes cell migration by stimulating the activity of FAK.

T98G Cell Death Induced by Photothermal Treatment with Hollow Gold Nanoshell-Coupled Silica Microrods Prepared from Escherichia Coli.

As an alternative to traditional cancer treatment, photothermal therapy is a promising method with advantages such as noninvasiveness and high efficiency. Herein, we synthesized armored golden Escherichia coli (AGE) microrods as photothermal agents to evaluate the viability of cancer cell. The hollow gold nanoshell (HAuNS) was synthesized for photothermal effects under the near-infrared (NIR) region using unicellular E. coli as a framework. Coupling HAuNS onto the surface of E. coli@SiO2 enhanced temperature elevation and resulted in high conversion efficiency. The synthesized AGE microrods had excellent photothermal stability under NIR laser irradiation in the five-times recycling experiment. The temperature elevation of AGE microrod solution reached 43.7 °C, which induced hyperthermia-mediated killing of tumor cells. The results of the cytotoxicity test revealed the AGE microrod-induced T98G cell death mediated via apoptosis.

CircRNA hsa-circ-0014359 promotes glioma progression by regulating miR-153/PI3K signaling

RNA back splicing produces circRNA, a new type of non-coding RNA. Studies have indicated that circRNAs play important roles in malignant tumors of the central nervous system. Here, we aimed to evaluate the expression of the circRNA, hsa-circ-0014359 (circ-0014359) in human glioma cell lines to assess its function in glioma progression and prognosis. The expression of circ-0014359 was increased in T98G and SHG44 cancer cell lines and glioma tissues from patients, when compared with control cells and tissue. SiRNA-mediated silencing of circ-0014359 potently inhibited cell viability, migration, invasion, and apoptosis of glioma cells. Further, our observations indicated that circ-0014359 acted as a miRNA-153 (miR-153) sponge in glioma cells. Transfection of miR-153 inhibitor significantly suppressed si-circ-0014359-induced inhibition of cell viability, cell migration, and invasion. The increased expression of circ-0014359 levels in glioma cells was correlated with downregulated expression of miR-153. Overexpression of miR-153 reduced p-AKTser473 (a PI3K pathway indicator) and the rescue experiment showed enhanced p-AKTser473 expression. Together, our study suggests that circ-0014359 promotes glioma progression via targeting miR-153/PI3K signaling pathway. Thus, our study provides insights into glioma progression and reveals potential new targets for treatment of glioma.

Targeted and Synergic Glioblastoma Treatment: Multifunctional Nanoparticles Delivering Verteporfin as Adjuvant Therapy for Temozolomide Chemotherapy.

Despite advances in cancer therapies, glioblastoma multiforme treatment remains inefficient due to the brain-blood barrier (BBB) inhibitory activity and to the low temozolomide (TMZ) chemotherapeutic selectivity. To improve therapeutic outcomes, in this work we propose two strategies, (i) photodynamic therapy (PDT) as adjuvant treatment and (ii) engineering of multifunctional theranostic/targeted nanoparticles ( m-NPs) that integrate biotin as a targeting moiety with rhodamine-B as a theranostic agent in pluronic P85/F127 copolymers. These smart m-NPs can surmount the BBB and coencapsulate multiple cargoes under optimized conditions. Overall, the present study conducts a rational m-NP design, characterization, and optimizes the formulation conditions. Confocal microscopy studies on T98-G, U87-MG, and U343 glioblastoma cells and on NIH-3T3 normal fibroblast cells show that the m-NPs and the encapsulated drugs are selectively taken up by tumor cells presenting a broad intracellular distribution. The formulations display no toxicity in the absence of light and are not toxic to healthy cells, but they exert a robust synergic action in cancer cells in the case of concomitant PDT/TMZ treatment, especially at low TMZ concentrations and higher light doses, as demonstrated by nonlinear dose-effect curves based on the Chou-Talalay method. The results evidenced different mechanisms of action related to the disjoint cell cycle phases at the optimal PDT/TMZ ratio. This effect favors synergism between the PDT and the chemotherapy with TMZ, enhances the antiproliferative effect, and overcomes cross-resistance mechanisms. These results point out that m-NP-based PDT adjuvant therapy is a promising strategy to improve TMZ-based glioblastoma multiforme treatments.

MIX2: A Novel Natural Multi-Component Modulator of Multidrug-Resistance and Hallmarks of Cancer Cells

Multidrug resistance is one of the key barriers suppressing the effectiveness of drug therapies of malignant tumors. Here, we report a study on the effect of a mix of natural extracts (MIX2) prepared from fresh fruits of Prunus spinosa, Crataegus monogyna, Sorbus aucuparia, and Euonymus europaeus on the classic hallmarks of cancer cells and the expression of multidrug resistance proteins. In the studies, HeLa and T98G cell lines, and classic methods of molecular biology, including RT-qPCR, Western blot, flow cytometry, and confocal imaging, were used. Additionally, migration, adhesion, and proliferation assays were performed. The obtained results indicate that the MIX2 cocktail presents strong anti-cancer properties. MIX2 is not toxic, but at the same time significantly alters the migration, proliferation, and adhesion of tumor cells. Furthermore, it was found that cells exposed to the mixture presented a significantly reduced expression level of genes associated with MDR, including ABCB1, which encodes for glycoprotein P. In vitro data showed that MIX2 effectively sensitizes tumor cells to doxorubicin. We postulate that modulation of the multidrug resistance phenotype of tumors with the use of MIX2 may be considered as a safe and applicable tool in sustaining drug delivery therapies of malignancies.

Transfection with GLS2 Glutaminase (GAB) Sensitizes Human Glioblastoma Cell Lines to Oxidative Stress by a Common Mechanism Involving Suppression of the PI3K/AKT Pathway

GLS-encoded glutaminase promotes tumorigenesis, while GLS2-encoded glutaminase displays tumor-suppressive properties. In glioblastoma (GBM), the most aggressive brain tumor, GLS is highly expressed and in most cases GLS2 is silenced. Previously, it was shown that transfection with a sequence encoding GAB, the main GLS2 isoform, decreased the survival, growth, and ability to migrate of human GBM cells T98G and increased their sensitivity towards an alkylating agent temozolomide (TMZ) and oxidative stress compared to the controls, by a not well-defined mechanism. In this study we report that GAB transfection inhibits growth and increases susceptibility towards TMZ and H2O2-mediated oxidative stress of two other GBM cell lines, U87MG and LN229. We also show that in GAB-transfected cells treated with H2O2, the PI3K/AKT pathway is less induced compared to the pcDNA-transfected counterparts and that pretreatment with PDGF-BB, an activator of AKT, protects GAB-transfected cells from death caused by the H2O2 treatment. In conclusion, our results show that (i) GAB suppresses the malignant phenotype of the GBM cells of different tumorigenic potentials and genetic backgrounds and (ii) the GAB-mediated increase of sensitivity to oxidative stress is causally related to the inhibition of the PI3K/AKT pathway. The upregulation of the GLS2 expression and the inhibition of the PI3K/AKT pathway may become a novel combined therapeutic strategy for anti-glioma preclinical investigations.

The expression and clinical significance of forkhead box protein K1 in glioma tissues

Objective Explore of the expression and significance of forkhead box protein K1 (FOXK1) in glioma. Methods Real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) and Western blotting analysis of mRNA and protein expression levels of FOXK1 in glioma tissues; Kaplan-Meier analysis of the relationship between FOXK1 and prognosis parameters; RNA interference on the expression of FOXK1 in glioblastoma cell. Transwell migration assay for the migration of LN18 cells. Cell counting kit-8 (CCK-8) counting for T98G cell proliferation. Results RT-qPCR and Western blotting showed that FOXK1 mRNA and protein is highly expressed in glioma tissues. The expression of FOXK1 was not significantly correlated with gender (P>0.05) and age of glioma patients (P>0.05), while FOXK1 expression was significantly with WHO classification (P<0.01). Kaplan-Meier analysis found that patients with high expression of FOXK1 had a poor prognosis (P<0.05). Transwell migration assay results indicate that FOXO1 promotes cancer cell metastasis. The CCK-8 cell count results indicate that FOXK1 promotes glioma cell proliferation. Conclusion FOXK1 is highly expressed in glioma patients and is associated with glioma development, progression, and metastasis. Key words: Forkhead box protein K1; Glioblastoma; Metastasis

A microRNA profile of pediatric glioblastoma: The role of NUCKS1 upregulation.

MicroRNAs (miRNAs/miRs) are a novel class of gene regulators that may be involved in tumor chemoresistance. Recently, specific miRNA expression profiles have been identified in adult glioblastoma (aGBM), but there are only limited data available on the role of miRNAs in pediatric GBM (pGBM). In the present study, the expression profile of miRNAs was examined in seven pGBMs and three human GBM cell lines (U87MG, A172 and T98G), compared with a non-tumoral pool of pediatric cerebral cortex samples by microarray analysis. A set of differentially expressed miRNAs was identified, including miR-490, miR-876-3p, miR-876-5p, miR-448 and miR-137 (downregulated), as well as miR-501-3p (upregulated). Through bioinformatics analysis, a series of target genes was predicted. In addition, similar gene expression patterns in pGBMs and cell lines was confirmed. Of note, drug resistant T98G cells had upregulated nuclear casein kinase and cyclin-dependent kinase substrate 1 (NUCKS1) expression, a protein overexpressed in many tumors that serves an important role in cell proliferation and progression. On the basis of the present preliminary report, it could be intriguing to further investigate the relationship between each of the identified differentially expressed miRNAs and NUCKS1, in order to clarify their involvement in the multi-drug resistance mechanism of pGBMs.

Gene Expression of Molecules Regulating Apoptotic Pathways in Glioblastoma Multiforme Treated with Umbilical Cord Stem Cell Conditioned Medium.

BackgroundGlioblastoma multiforme (GBM) is the most malignant primary brain tumour and there is no definite cure. It has been suggested that there are significant interactions among mesenchymal stem cells (MSCs), their released factors and tumour cells that ultimately determine GBM’s growth pattern. This study aims to analyse the expression of molecules involved in GBM cell apoptotic pathways following treatment with the MSC secretome.MethodsA conditioned medium of umbilical cord-derived MSCs (UCMSC-CM) was generated by culturing the cells on serum-free αMEM for 24 h. Following this, human GBM T98G cells were treated with UCMSC-CM for 24 h. Quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) was then performed to measure the mRNA expression of survivin, caspase-9, TNF-related apoptosis-inducing ligand (TRAIL), DR4 and DcR1.ResultsmRNA expression of caspase-9 in CM-treated T98G cells increased 1.6-fold (P = 0.017), whereas mRNA expression of survivin increased 3.5-fold (P = 0.002). On the other hand, TRAIL protein expression was upregulated (1.2-fold), whereas mRNA expression was downregulated (0.4-fold), in CM-treated cells. Moreover, there was an increase in the mRNA expression of both DR4 (3.5-fold) and DcR1 (1,368.5-fold) in CM-treated cells.ConclusionThe UCMSC-CM was able to regulate the expression of molecules involved in GBM cell apoptotic pathways. However, the expression of anti-apoptotic molecules was more upregulated than that of pro-apoptotic molecules.

Chemotherapeutic Effect of SR9009, a REV-ERB Agonist, on the Human Glioblastoma T98G Cells.

Glioblastoma multiforme is the most aggressive brain tumor, and human T98G cells constitute a useful glioblastoma multiforme model to evaluate the chemotherapeutic agents. Modern life (shiftwork, jetlag, etc.) may cause circadian disorganization promoting higher cancer risk and metabolic disorders. Although little is known about the tumor-intrinsic circadian clock function, pharmacological modulation of circadian components may offer selective anticancer strategies. REV-ERBs are heme-binding circadian clock components acting as repressors of processes involved in tumorigenesis such as metabolism, proliferation, and inflammation. A synthetic pyrrole derivative (SR9009) that acts as REV-ERBs-specific agonists exhibits potent in vivo activity on metabolism and tumor cell viability. Here, we investigated SR9009 effects on T98G cell viability, differential chemotherapy time responses, and underlying metabolic processes (reactive oxygen species [ROS] and lipid droplets [LDs]) and compared it with the proteasome inhibitor Bortezomib treatment. SR9009-treated cells exhibited significant reduction in cell viability with consequences on cell cycle progression. Dexamethasone synchronized cells displayed differential time responses to SR9009 treatment with highest responses 18 to 30 h after synchronization. SR9009 treatment decreased ROS levels while Bortezomib increased them. However, both treatments significantly increased LD levels, whereas the combined treatment showed additive or synergistic effects between both drugs. In addition, we extended these studies to HepG2 cells which also showed a significant decrease in cell viability and ROS levels and the increase in LD levels after SR9009 treatment. Our results suggest that the pharmacological modulation of the tumor-intrinsic clock by REV-ERB agonists severely affects cell metabolism and promotes cytotoxic effects on cancer cells.

The impact of conditioned medium of umbilical cord-derived mesenchymal stem cells toward apoptosis and proliferation of glioblastoma multiforme cells

Glioblastoma multiforme (GBM) is the human malignant and highly invasive brain tumor. Tumor growth and invasive property are affected by mesenchymal stem cells (MSCs) as a part of tumor microenvironment. However, accumulating evidences describe that MSCs could act as pro and anti tumorigenic. The different action of MSCs in regulating tumor growth is mediated by secreted factor. This research purpose was to analyze the impact of MSCs secreted factor in conditioned medium (CM) on apoptosis and proliferation of GBM cells. Primary culture of umbilical cord-derived MSCs (UCSCs) was conducted in this research. CM-UCSCs was prepared by culturing the UCSCs in serum free aMEM for 24 hours. Those CM-UCSCs was 2-fold diluted by Dulbecco’s Modified Eagle Medium (DMEM) high glucose (50% concentration) and used to treat human GBM T98G cells for 24 hours. Following this treatment, apoptosis was detected using annexin V-FITC and cells proliferation was analyzed using trypan blue exclusion test. The results showed that early apoptosis occurred in 14.4 % of control cells and 14.8 % of CM-treated T98G cells, as well as late apoptosis occurred in 7.8 % of control cells and 7.2 % of CM-treated T98G cells. Whereas, GBM cells proliferation was tend to higher in the CM treated cells (3.85×105 cells) compared to the control (2.97×105 cells). In conclusion, CM-UCSCs does not seems to affect apoptosis, but tend to increase cells proliferation. Further research is needed to elaborate the mechanism of UCSCs secretome in stimulating cells proliferation.

Novel small molecule protein kinase CK2 inhibitors exert potent antitumor effects on T98G and SEGA cells in vitro.

Tumours of astroglial origin, both malignant glioblastoma (GBM) and benign subependymal giant cell astrocytoma (SEGA), pose a serious medical problem. Casein kinase 2 (CK2), a member of the serine/threonine kinase family, has antiapoptotic properties and plays a vital role in glial tumour cell survival. It contributes to invasive cell growth and is often upregulated in malignant neoplastic cells; however, its role in benign tumours of astrocytic origin is less understood. In the present study we investigated the effects of small molecule CK2 inhibitors on proliferation and viability of glioma cells in vitro. The experiments were conducted on commercial T98G malignant glioma cell line and the SEGA cell line, derived from a paediatric case of tuberous sclerosis complex (TSC). Cell cultures were incubated with selected CK2 inhibitors: 4,5,6,7-tetrabromo-1H-benzimidazole (TBI), 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole (DMAT) and 4,5,6,7-tetrabromo-1H-benzotriazole (TBB) at 0.1, 1, 10, 25, 50, 75 and 100 µM concentrations for 24 and 48 hours. Cell proliferation was assessed using a cell counter and cell viability was evaluated by MTT assay. TBB at 75 µM and 100 µM, and TBI starting from 25 µM, both reduced T98G cell proliferation after 24 hours, while DMAT was ineffective. All tested small molecule CK2 inhibitors appear to reduce T98G cell growth and viability after 48 hours, although TBI appeared to be the most effective and reduced cell growth in the 50-100 µM dose range. TBI also showed potential efficacy in reducing the number and viability of SEGA cells after 48 hours. Proliferation and viability of SEGA cells have proven resistant to TBB treatment. DMAT only reduced the viability of SEGA cells at 24 (at 100 µM) and 48 hours (10-100 µM). Importantly, normal human astrocyte cells were found to be moderately resistant to TBB, while their viability was mildly reduced at higher doses of DMAT and TBI. In conclusion, CK2 appears to play a role not only in malignant glioma cells but it can also sustain the viability and proliferation of benign astrocytoma cells. The obtained antitumor effects of CK2 inhibitors significantly exceeded their mild or no effect on normal astrocytes in control, which supports the therapeutic potential of these compounds against gliomas.

Proapoptotic effects of novel thiazole derivative on human glioma cells.

The aim of the present study was to investigate the antiproliferative and proapoptotic actions of N-(5-benzyl-1,3-thiazol-2-yl)-3,5-dimethyl-1-benzofuran-2-carboxamide derivative (compound 5) in glioma cells in comparison with the actions of temozolomide (TMZ) and doxorubicin (Dox), used as positive controls. The antiproliferative activity of the compound 5, TMZ, and Dox on human glioblastoma U251 and human glioblastoma multiform T98G cells was measured using the MTT test. Western blot analysis, fluorescent microscopy, agarose gel retardation assay, flow cytometric analysis, and the DNA comet assay under alkaline conditions were carried out to study the effect of compound 5 on U251 cells. This compound showed ~20 times higher cytotoxicity toward U251 and T98G cells compared with the effects of TMZ and approximately two times higher activity than that of the Dox. Compound 5 induced apoptosis in U251 cells by PARP1 and caspase 3 cleavage mechanisms, also inducing an increase in the level of Bax and Bim proapoptotic proteins and a decrease in the level of phosho-ERK1/2 kinase. The cytotoxicity of compound 5 was associated with an increase in the production of the hydrogen peroxide and the formation of DNA single-strand breaks. This compound 5 did not intercalate into a DNA molecule. Thus, the novel thiazole derivative (compound 5) proved to be a potential antiglioma drug that showed much higher cytotoxic action on human glioma cells compared with the effects of TMZ and Dox. Its cytotoxicity is associated with apoptosis induction, production of the reactive oxygen species, and formation of DNA single-strand breaks without significant DNA intercalation.

Inhibition of cell proliferation by wortmannin in T98G cells involved induced inhibition of NF-kB transcriptional activity

Inhibition of cell proliferation by wortmannin in T98G cells involved induced inhibition of NF-kB transcriptional activity