Proliferation Of Human Glioblastoma Cells Research Articles

Ubiquitination of p21 by E3 ligase RNF135 promotes the proliferation of human glioblastoma cells.

Glioblastoma multiforme (GBM) is a highly heterogeneous tumor of the central nervous system with a high mortality rate. The upregulation of RING finger protein 135 (RNF135), an E3 ligase, has been observed in GBM, but the associated mechanisms have not been fully elucidated. The aim of the present study was to identify the substrate of RNF135 and study its functions in GBM. Bioinformatics analyses were performed. In addition, RNF135 was overexpressed or knocked down in human U87 and U251 GBM cells, and the effect on cell proliferation was analyzed using Cell Counting Kit-8 and colony formation assays. Furthermore, the interaction of RNF135 with its potential substrate was analyzed using glutathione S-transferase, yeast two-hybrid, immunoprecipitation (IP), co-IP and immunoblotting assays. Bioinformatics analysis indicated that RNF135 serves as a marker of poor prognosis in GBM. The overexpression of RNF135 was demonstrated to promote the proliferation of GBM cells, while the knockdown of RNF135 inhibited GBM cell growth. In addition, the results of the interaction experiments indicate that RNF135 interacts with p21 and mediates the degradation of p21 by ubiquitination. The major site of RNF135-mediated p21 ubiquitination was identified as K163. Further experiments demonstrated that RNF135 promotes the proliferation of GBM cells mainly via p21. In summary, these findings suggest that RNF135 has potential as a therapeutic target for the treatment of GBM.

Nerolidol inhibited U-251 human glioblastoma cell proliferation and triggered apoptosis via the upregulation of the p38 MAPK signaling pathway.

Glioblastoma multiforme (GBM) is a lethal brain tumor with high mortality and morbidity. Nerolidol (NRD) is a sesquiterpene alcohol sequestered from the essential oils of aromatic florae with potent antioxidant, antiviral, anticancer, cardioprotective, and neuroprotective activity. The aim of the study was to investigate the underlying cell-cycle mechanisms of NRD-mediated antiproliferative and apoptosis activities in GBM using human U-251 cells. The current research investigated the antiproliferative and apoptotic activities of NRD on U-251 cells. The effects of NRD were measured using a Cell Counting Kit-8 (CCK-8) assay, 4',6-diamidino-2-phenylindole (DAPI) staining, messenger ribonucleic acid (mRNA) level assessment, and western blot assay. Nerolidol decreased U-251 viability in a dose-dependent manner, as well as induced apoptotic activity, reduced B-cell lymphoma-2 (BCL-2) levels, and increased mRNA expression of BCL-2-associated X (Bax), caspase-3 and caspase-9. The attenuation of the cyclin-D1, cyclin-dependent kinase 4 (CDK4) and CDK6 mRNA expression confirmed cell cycle regulation. Western blot analysis of CDK1 indicated reductions in cyclin-B1 and p21. Furthermore, NRD prompted apoptosis through p38 amelioration and increased phosphorylated extracellular signal-related kinase 1 (p-ERK1) and phosphorylated c-Jun N-terminal protein kinase 1 (p-JNK1) levels. Nerolidol inhibited GBM cell viability and induced apoptosis through the regulation of cell-cycle proteins via p38 mitogen-activated protein kinase (MAPK) signaling pathways. Thus, NRD could be developed as a potential natural therapeutic agent for GBM.

Abstract 395: An alternating magnetic field suppresses tumor growth

Abstract Application of physical forces, ranging from ultrasound to electric field, is recommended in various clinical practice guidelines, including those for bone fracture or cancer treatment. However, mechanistic information for such treatments has been poorly provided, due to the lack of comprehensive study model. Here, we show an alternating magnetic field (AMF) per se exhibits direct anti-cancer effect through enhancing oxidative phosphorylation (OXPOHS) and thus metabolic reprograming. Proliferation of human glioblastoma cells (U87 and LN229), but not normal human astrocytes (NHA), was inhibited upon exposure to AMF within the narrow frequency range. Extracellular acidification rate was decreased while oxygen consumption was increased. These data indicated that AMF induced metabolic reprograming. This was accompanied by increased reactive oxygen species (ROS) production. Mouse models grafted with human glioblastoma cells showed that AMF exposure, 30 minutes per day for two weeks, suppressed the tumor growth and prolonged the overall survival. Our results suggest that the AMF exposure may be a simple strategy to inhibit cancer cell growth by utilizing oxidative stress via metabolic reprograming. Citation Format: Masanari Umemura, Taisuke Akimoto, Rafikul Islam, Kazuhito Kishi, Akane Nagasako, Rina Nakakaji, Takashi Yamaguchi, Yuto Mizuno, Soichiro Ishikawa, Tetsuya Yamamoto, Yoshihiro Ishikawa. An alternating magnetic field suppresses tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 395.

Apoptotic and Autophagic Cell Death Effects of the Hexane Extract of Tropical Marine Algae Halymenia durvillei against Human Glioblastoma Cells: In vitro and in silico Studies

Glioblastoma (GBM) considered as aggressive brain cancer with high mortality rate in patients even after surgical resection. Resistant to chemotherapy is the major problem in GBM therapy. Discovery of novel bioactive compounds from algae is being investigated as alternative sources for potential treatment as well as prevention in glioblastoma. This study revealed the effects of marine red algae extract from hexane solvent fraction of Halymenia durvillei (HDHE) on proliferation and cell death in A172 human GBM cells. HDHE decreased proliferation and promoted cell cycle arrest at G2/M phase. HDHE induced apoptotic cell death in A172 cells through mitochondrial membrane dysfunction, the decrease of anti-apoptotic Bcl-2 protein expression, and activation of caspase 3/7. Moreover, HDHE increased intracellular reactive oxygen species (ROS) production and accumulation of LC3-II, an autophagic marker. The docked conformation of palmitic acid, a major component of HDHE, showed a high affinity binding to TP53 and Beclin-1 as cell death-related target molecules. This research conclusively demonstrated that HDHE might serve as a potent anticancer agent against glioblastoma by promoting apoptotic and autophagic cell death in A172 human GBM cells. HIGHLIGHTS The palmitic acid-enriched extract of red alga Halymenia durvillei (HDHE) could inhibit proliferation of A172 human glioblastoma cells by arresting cell cycle at G2/M phase HDHE could induce both caspase-dependent apoptotic and autophagic death of A172 cells Palmitic acid, a major component of HDHE, showed a high affinity binding to TP53 and Beclin-1 The extract could be beneficial to develop as chemopreventive agents or food supplements against glioblastoma GRAPHICAL ABSTRACT

Celecoxib Suppresses NF-κB p65 (RelA) and TNFα Expression Signaling in Glioblastoma.

Glioblastoma (GBM) harbors significant genetic heterogeneity, high infiltrative capacity, and patterns of relapse following many therapies. The expression of nuclear factor kappa-B (NF-κB p65 (RelA)) and signaling pathways is constitutively activated in GBM through inflammatory stimulation such as tumor necrosis factor-alpha (TNFα), cell invasion, motility, abnormal physiological stimuli, and inducible chemoresistance. However, the underlying anti-tumor and anti-proliferative mechanisms of NF-κB p65 (RelA) and TNFα are still poorly defined. This study aimed to investigate the expression profiling of NF-κB p65 (RelA) and TNFα as well as the effectiveness of celecoxib along with temozolomide (TMZ) in reducing the growth of the human GBM cell line SF-767. genome-wide expression profiling, enrichment analysis, immune infiltration, quantitative expression, and the Microculture Tetrazolium Test (MTT) proliferation assay were performed to appraise the effects of celecoxib and TMZ. demonstrated the upregulation of NF-κB p65 (RelA) and TNFα and celecoxib reduced the viability of the human glioblastoma cell line SF-767, cell proliferation, and NF-κB p65 (RelA) and TNFα expression in a dose-dependent manner. Overall, these findings demonstrate for the first time how celecoxib therapy could mitigate the invasive characteristics of the human GBM cell line SF-767 by inhibiting the NF-κB mediated stimulation of the inflammatory cascade. based on current findings, we propose that celecoxib as a drug candidate in combination with temozolomide might dampen the transcriptional and enzymatic activities associated with the aggressiveness of GBM and reduce the expression of GBM-associated NF-κB p65 (RelA) and TNFα inflammatory genes expression.

TRAIL and Celastrol Combinational Treatment Suppresses Proliferation, Migration, and Invasion of Human Glioblastoma Cells via Targeting Wnt/β-catenin Signaling Pathway.

To investigate the mechanistic basis for the anti-proliferation and anti-invasion effect of tumor necrosis factor-related apoptosis-induced ligand (TRAIL) and celastrol combination treatment (TCCT) in glioblastoma cells. Cell counting kit-8 was used to detect the effects of different concentrations of celastrol (0-16 µmol/L) and TRAIL (0-500 ng/mL) on the cell viability of glioblastoma cells. U87 cells were randomly divided into 4 groups, namely control, TRAIL (TRAIL 100 ng/mL), Cel (celastrol 0.5 µmol/L) and TCCT (TRAIL 100 ng/mL+ celastrol 0.5 µmol/L). Cell proliferation, migration, and invasion were detected by colony formation, wound healing, and Transwell assays, respectively. Quantitative reverse transcription polymerase chain reaction and Western blotting were performed to assess the levels of epithelial-mesenchymal transition (EMT) markers (zona occludens, N-cadherin, vimentin, zinc finger E-box-binding homeobox, Slug, and β-catenin). Wnt pathway was activated by lithium chloride (LiCl, 20 mol/L) and the mechanism for action of TCCT was explored. Celastrol and TRAIL synergistically inhibited the proliferation, migration, invasion, and EMT of U87 cells (P<0.01). TCCT up-regulated the expression of GSK-3β and down-regulated the expression of β-catenin and its associated proteins (P<0.05 or P<0.01), including c-Myc, Cyclin-D1, and matrix metalloproteinase (MMP)-2. In addition, LiCl, an activator of the Wnt signaling pathway, restored the inhibitory effects of TCCT on the expression of β-catenin and its downstream genes, as well as the migration and invasion of glioblastoma cells (P<0.05 or P<0.01). Celastrol and TRAIL can synergistically suppress glioblastoma cell migration, invasion, and EMT, potentially through inhibition of Wnt/β-catenin pathway. This underlies a novel mechanism of action for TCCT as an effective therapy for glioblastoma.

Nanosecond pulsed cold atmospheric plasma jet suppresses proliferation and migration of human glioblastoma cells via apoptosis promotion and EMT inhibition

Glioblastoma (GBM) is one of the most aggressive and challenging cancers to treat. Despite extensive research on dozens of cancer cells, including GBM, the effect of cold atmospheric plasma (CAP) on the invasive migration of GBM cells has received limited attention, and the underlying mechanisms remain poorly understood. This study aims to investigate the potential molecular mechanism of ns-CAPJ in inhibiting the invasive migration of human GBM cells. The findings indicate that ns-CAPJ significantly reduces GBM cell invasion and migration, and induces apoptosis in GBM cells. Further mechanistic studies demonstrate a direct correlation between the suppression of the epithelial-mesenchymal transition (EMT) signaling pathway and ns-CAPJ’s inhibitory effect on GBM cell invasion and migration. Additionally, combined with the N-acetyl cysteine (NAC, a ROS inhibitor) assay, we found that the ROS stimulated by the ns-CAPJ plays an important role in suppressing the EMT process. This work is expected to provide new insight into understanding the molecular mechanisms of how ns-CAPJ inhibits the proliferation and migration of human GBM cells.

P10.30.B COMBINATION TREATMENT WITH ERIBULIN AND TTFIELDS INDUCES APOPTOSIS AND G2/M CELL CYCLE ARREST IN HUMAN GLIOBLASTOMA CELLS

Abstract BACKGROUND TTFields has emerged to a first line treatment in newly diagnosed glioblastoma (GBM) patients. Several preclinical studies showed that the alternating electric fields inhibit proliferation and survival of human glioblastoma cells by disruption of the spindle apparatus and impairment of mitotic segregation. Eribulin is an antimitotic drug, that is currently used in the third line treatment of breast cancer. As it is also able to inhibit the pro-mitotic function of TERT, which is mutated in 80% of human glioblastoma, the drug was also shown to have a radiosensitizing and antiproliferative effect on human glioblastoma and ovarian cancer cell lines. This study aims to evaluate the effect of the combination of Eribulin and TTFields on the survival and cell cycle of glioblastoma cells. MATERIAL AND METHODS U87, A172 and four patient derived glioblastoma cell lines were cultured and plated to 80% confluence. Cells were then treated with TTFields at a frequency of 200kHz for 24 hours and/or received the ED50 and 0.5-fold ED50 of Eribulin. Controls remained untreated respectively. After 24 hours cells were harvested and counted by automatic cell count. Survival and cell cycle status were evaluated by FACS analysis. RESULTS The rate of early apoptotic cells was 82.28% in the group treated with TTFields and Eribulin. With TTFields treatment only 50.99% and with Eribulin alone 14.38% of cells were in an early apoptotic state. Untreated control cells had an apoptosis rate of 4.25%. The combination of TTFields and Eribulin was significantly more effective than the single treatment modalities (p&amp;gt;0.05). Treatment with TTFields increased the rate of cells an G2/M status. This effect was even stronger in the group with combination treatment. CONCLUSION Combination treatment of the antimitotic drug Eribulin with TTFields is able to improve the inhibitory effect of the drugs on cell survival. Furthermore the combination therapy effects the cell cycle of glioblastoma cells inducing G2/M arrest. Future preclinical studies should focus on the antimitotic effect and the influence of the treatment on genome integrity for a more detailed insight into the mechanisms of action.

Platycodin D suppresses proliferation, migration, and invasion of human glioblastoma cells through regulation of Skp2

BackgroundPlatycodin D (PD) is a major bioactive component of Platycodon grandiflorum, a medicinal herb that is widely used in China, and is effective against various human cancers, including glioblastoma multiforme (GBM). S phase kinase-related protein 2 (Skp2) is oncogenic and overexpressed in various human tumors. It is highly expressed in GBM and its expression is correlated with tumor growth, drug resistance and poor prognosis. In this study, we investigated whether inhibition of glioma progression by PD is mediated by decreasing expression of Skp2. MethodsCell Counting Kit-8 (CCK-8) and Transwell assays were used to determine the effects of PD on GBM cell proliferation, migration, and invasion in vitro. mRNA and protein expression were determined by real time polymerase chain reaction (RT-PCR) and western blotting, respectively. The U87 xenograft model was used to verify the anti-glioma effect of PD in vivo. Expression levels of Skp2 protein were analyzed by immunofluorescence staining. ResultsPD suppressed proliferation and motility of GBM cells in vitro. The expression of Skp2 in U87 and U251 cells was significantly reduced by PD. PD mainly decreased the cytoplasmic expression of Skp2 in glioma cells. Skp2 protein expression was downregulated by PD, resulting in upregulation of its downstream targets, p21and p27. The inhibitory effect of PD was enhanced by Skp2 knockdown in GBM cells and reversed in cells with Skp2 overexpression. ConclusionPD suppresses glioma development by regulation of Skp2 in GBM cells.

Secretome of Dental Pulp-Derived Stem Cells Reduces Inflammation and Proliferation of Glioblastoma Cells by Deactivating Mapk-Akt Pathway.

Dental pulp-derived stem cells (DPSC) is a promising therapy as they modulate the immune response, so we evaluated the inhibitory effect of DPSC secretome (DPSC℗) on the proliferation and inflammation in human glioblastoma (GBM) cells (U-87 MG) and elucidated the concomitant mechanisms involved. The U87-MG cells were cultured with DPSC℗ for 24 h and assessed the expression of inflammatory molecules using quantitative reverse transcription-polymerase chain reaction (qRT-PCR), generation of reactive oxygen species (ROS), and mitochondrial functionality using a seahorse flux analyzer. MTT (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide) assay and cell cycle analysis were performed to evaluate the proliferation and cell cycle. Finally, the protein levels were determined by western blot. DPSC℗ reduced the inflammation and proliferation of U-87 MG cells by down-regulating the pro-inflammatory markers and up-regulating anti-inflammatory markers expressions through ROS-mediated signaling. Moreover, DPSC℗ significantly reduced the mitochondrial membrane potential (MMP) in the cells. The cellular bioenergetics revealed that all the parameters of oxygen consumption rate (OCAR) and the extracellular acidification rate (ECAR) were significantly decreased in the GBM cells after the addition of DPSC℗. Additionally, DPSC℗ decreased the GBM cell proliferation by arresting the cell cycle at the G1 phase through activation (phosphorylation) of checkpoint molecule CHK1. Furthermore, mechanistically, we found that the DPSC℗ impedes the phosphorylation of the mitogen-activated protein kinases (P38 MAPK) and protein kinase B (AKT) pathway. Our findings lend the first evidence of the inhibitory effects of DPSC℗ on proliferation and inflammation in GBM cells by altering the P38 MAPK-AKT pathway.

Camboginol and Morelloflavone from Garcinia dulcis (Roxb.) Kurz Flower Extract Promote Autophagic Cell Death against Human Glioblastoma Cells through Endoplasmic Reticulum Stress.

Garcinia dulcis is a tropical plant native to Southeast Asia that is traditionally used as a folk remedy to cure several pathological symptoms. Camboginol and morelloflavone have been revealed by previous studies as the principal bioactive compounds from the flower extract of G. dulcis. The disease-preventing properties of camboginol or morelloflavone, including anti-cancer, from various parts of G. dulcis have been revealed by recent studies. Glioblastoma is the aggressive malignant stage of brain cancer and suffers from chemotherapeutic resistance. This study aimed to test the anti-cancer effect of G. dulcis flower extract against the proliferation of A172 human glioblastoma cells. The extract had cytotoxic activity and promoted cell cycle arrest at the S and G2/M phases. Autophagic cell death was promoted by cytotoxic concentrations of the extract, as observed by enhancing autophagic flux and the expression of autophagic markers. Autophagic cell death induced by the extract might be associated with endoplasmic reticulum (ER) stress. Conclusively, it was indicated by this study that the extract from the flower of G. dulcis had a protective effect against the proliferation of A172 human glioblastoma cells through the induction of ER stress-mediated cytotoxic autophagy.

Role of the Ca2+ channel α2δ-1 auxiliary subunit in proliferation and migration of human glioblastoma cells.

The overexpression of α2δ-1 is related to the development and degree of malignancy of diverse types of cancer. This protein is an auxiliary subunit of voltage-gated Ca2+ (CaV) channels, whose expression favors the trafficking of the main pore-forming subunit of the channel complex (α1) to the plasma membrane, thereby generating an increase in Ca2+ entry. Interestingly, TLR-4, a protein belonging to the family of toll-like receptors that participate in the inflammatory response and the transcription factor Sp1, have been linked to the progression of glioblastoma multiforme (GBM). Therefore, this report aimed to evaluate the role of the α2δ-1 subunit in the progression of GBM and investigate whether Sp1 regulates its expression after the activation of TLR-4. To this end, the expression of α2δ-1, TLR-4, and Sp1 was assessed in the U87 human glioblastoma cell line, and proliferation and migration assays were conducted using different agonists and antagonists. The actions of α2δ-1 were also investigated using overexpression and knockdown strategies. Initial luciferase assays and Western blot analyses showed that the activation of TLR-4 favors the transcription and expression of α2δ-1, which promoted the proliferation and migration of the U87 cells. Consistent with this, overexpression of α2δ-1, Sp1, and TLR-4 increased cell proliferation and migration, while their knockdown with specific siRNAs abrogated these actions. Our data also suggest that TLR-4-mediated regulation of α2δ-1 expression occurs through the NF-kB signaling pathway. Together, these findings strongly suggest that the activation of TLR-4 increases the expression of α2δ-1 in U87 cells, favoring their proliferative and migratory potential, which might eventually provide a theoretical basis to examine novel biomarkers and molecular targets for the diagnosis and treatment of GBM.

Echinacoside (ECH) suppresses proliferation, migration, and invasion of human glioblastoma cells by inhibiting Skp2-triggered epithelial-mesenchymal transition (EMT)

BackgroundEchinacoside (ECH) is a phenylethanoid extracted from the stems of Cistanches salsa, an herb used in Chinese medicine formulations, and is effective against glioblastoma multiforme (GBM). Epithelial-mesenchymal transition (EMT) is the cornerstone of tumorigenesis and metastasis, and increases the malignant behavior of GBM cells. The S phase kinase-related protein 2 (skp2), an oncoprotein associated with EMT, is highly expressed in GBM and significantly associated with drug resistance, tumor grade and dismal prognosis. The aim of this study was to explore the inhibitory effects of ECH against GBM development and skp2-induced EMT. MethodsCCK-8, EdU incorporation, transwell, colony formation and sphere formation assays were used to determine the effects of ECH on GBM cell viability, proliferation, migration and invasion in vitro. The in vivo anti-glioma effects of ECH were examined using a U87 xenograft model. The expression levels of skp2 protein, EMT-associated markers (vimentin and snail) and stemness markers (Nestin and sox2) were analyzed by immunofluorescence staining and western blotting experiments. ResultsECH suppressed the proliferation, invasiveness and migration of GBM cells in vitro, as well as the growth of U87 xenograft in vivo. In addition, ECH downregulated the skp2 protein, EMT-related markers (vimentin and snail) and stemness markers (sox2 and Nestin). The inhibitory effects of ECH were augmented in the skp2-knockdown GBM cells, and reversed in cells with ectopic expression of skp2. ConclusionECH inhibits glioma development by suppressing skp2-induced EMT of GBM cells.

Contribution of the Testosterone Androgen Receptor-PARD3B Signaling Axis to Tumorigenesis and Malignance of Glioblastoma Multiforme through Stimulating Cell Proliferation and Colony Formation.

Background: Glioblastoma multiforme (GBM) is the most common and malignant brain tumor with very poor prognoses. After surgical resection of the primary tumor, rapid proliferation of residual glioblastoma cells is a critical cause explaining tumor malignance and recurrence. In this study, we evaluated de novo roles of the testosterone androgen receptor (AR)–PARD3B signaling axis in the tumorigenesis and malignance of human GBM and the possible related mechanisms. Methods: AR and PARD3B gene expressions and their correlations were mined from The Cancer Genome Atlas (TCGA) database and analyzed using the UALCAN system. Analyses using a real-time PCR, cell proliferation, and colony formation and a loss-of-function strategy by suppressing AR activity with its specific inhibitor, enzalutamide, were then carried out to determine roles of the testosterone AR–PARD3B signaling axis in tumor malignance. Results: Expressions of AR, PARD3B mRNA, and proteins in human GBM tissues were upregulated compared to normal human brain tissues. In contrast, levels of AR and PARD3B mRNA in most TCGA pan-cancer types were downregulated compared to their respective normal tissues. Interestingly, a highly positive correlation between AR and PARD3B gene expressions in human GBM was identified. The results of a bioinformatics search further showed that there were five AR-specific DNA-binding elements predicted in the 5′ promoter of the PARD3B gene. Regarding the mechanisms, exposure of human glioblastoma cells to testosterone induced AR and PARD3B gene expressions and successively stimulated cell proliferation and colony formation. Suppressing AR activity concurrently resulted in significant attenuations of testosterone-induced PARD3B gene expression, cell proliferation, and colony formation in human glioblastoma cells. Conclusions: This study showed the contribution of the testosterone AR–PARD3B signaling axis to the tumorigenesis and malignance of human GBM through stimulating cell proliferation and colony formation. Therefore, the AR-PARD3B signaling axis could be targeted for potential therapy for human GBM.

Under lithium carbonate administration, nicotine triggers cell dysfunction in human glioblastoma U-251MG cells, which is distinct from cotinine.

Nicotine is an alkaloid found in tobacco leaves. Smoking prevention has been a neglected issue in psychiatry; nicotine intake in conjunction with the administration of the mood stabilizer, lithium carbonate (Li2CO3), may negatively affect brain cells. The present study investigated the combined effects of nicotine and its metabolite, cotinine, and Li2CO3 compared to acetylcholine and dopamine in U-251MG human glioblastoma cells. Cell proliferation was found to be decreased by nicotine and to be further suppressed following treatment with Li2CO3, accompanied by mitotic catastrophe and increased levels of superoxide anion radicals. By contrast, cotinine did not exert such detrimental effects. It was also found that acetylcholine did not suppress cell proliferation, whereas dopamine in conjunction with Li2CO3 decreased cell proliferation in a concentration-dependent manner. The nicotine-induced cell growth inhibition was restored by mecamylamine, a non-competitive antagonist of nicotinic acetylcholine receptors. On the whole, the findings of the present study suggest that nicotine combined with Li2CO3 leads to the suppression of the proliferation of human glioblastoma cells accompanied by mitotic catastrophe and superoxide anion radical generation. These findings may provide further cellular biological insight into the risks associated with smoking under Li2CO3 administration.

Tizoxanide Promotes Apoptosis in Glioblastoma by Inhibiting CDK1 Activity.

The antiparasitic drug nitazoxanide (NTZ) has received considerable attention for its potential in cancer therapy. In this study, we demonstrate that tizoxanide (TIZ), an active metabolite of NTZ, exhibits antiglioma activity in vitro and in vivo by inducing G2/M cell cycle arrest and apoptosis. In vitro, TIZ dose-dependently inhibited the proliferation of U87, U118, and A172 human glioblastoma (GBM) cells at 48 h with IC50 values of 1.10, 2.31, and 0.73 µM, respectively. Treatment with TIZ (1 and 10 µM) also dose-dependently inhibited the colony formation of these GBM cells and accumulated ROS damage in the nucleus. In silico target fishing combined with network pharmacological disease spectrum analyses of GBM revealed that cycle-dependent kinase 1 (CDK1) is the most compatible target for TIZ and molecular docking by Molecule Operating Environment (MOE) software confirmed it. Mechanistically, TIZ inhibited the phosphorylation of CDK1 at Thr161 and decreased the activity of the CDK1/cyclin B1 complex, arresting the cell cycle at the G2/M phase. TIZ may induce apoptosis via the ROS-mediated apoptotic pathway. In vivo, TIZ suppressed the growth of established subcutaneous and intracranial orthotopic xenograft models of GBM without causing obvious side effects and prolonged the survival of nude mice bearing glioma. Taken together, our results demonstrated that TIZ might be a promising chemotherapy drug in the treatment of GBM.

Green Synthesized Bismuth Oxide Nanoparticles Using Aqueous Rhizome Extract ofCurcuma longaMitigate the Proliferation of Human U87 Glioblastoma Cells by Regulation of the Wnt/β-Catenin Signaling Pathway

The low biocompatibility of inorganic nanoparticles (NPs) is a main concern in their wide applications in the biomedical field. Therefore, the green synthesis of NPs from plant extracts can provide safe NPs for biomedical applications. The present study was aimed to assess the anticancer activity of bismuth oxide (Bi2O3) NPs fabricated using aqueous plant extracts from the rhizome ofCurcuma longa(C. longa). Characterization of green Bi2O3NPs was done using TEM, DLS, and X-ray diffraction analyses. Selective anticancer activity of green Bi2O3NPs against human glioblastoma (U87) cells was assessed using MTT, GSH, MDA, ROS, apoptosis, and caspase-3 assays. Also, qPCR analysis was done to explore the expression ofβ-catenin, cyclin D1, and c-myc at mRNA level as the important genes of the Wnt/β-catenin signaling pathway. The results showed that the green Bi2O3NPs have a crystalline nature with a size of around 30 nm with good colloidal stability attributed to potential bio-fabrication of Bi2O3NPs. Cellular study indicated that green Bi2O3NPs triggered selective anticancer activity against U87 cells through reduction of GSH level and increase of MDA level, ROS level, Annexin+cells, and caspase-3 activity. Also, it was found that IC50concentration of biosynthesized Bi2O3NPs (20μg/mL) resulted in a significant downregulation in the expression ofβ-catenin, cyclin D1, and c-myc genes involved in the Wnt/β-catenin signaling pathway. This study concludes that green Bi2O3NPs bio-fabricated from rhizome ofC. longashow potential selective anticancer activity.

Effects of Vitexin, a Natural Flavonoid Glycoside, on the Proliferation, Invasion, and Apoptosis of Human U251 Glioblastoma Cells.

Glioblastoma is a highly aggressive brain tumor characterized by high recurrence and poor prognosis. Vitexin has shown activities against esophageal, liver, lung, colorectal, and ovarian cancers; however, there is little knowledge on the activity of vitexin against glioblastoma. This study was therefore designed with aims to examine the effects of vitexin on proliferation, invasion, and apoptosis of human U251 glioblastoma cells and explore the underlying molecular mechanisms using mRNA sequencing and molecular docking. Vitexin was found to inhibit cell proliferation, colony formation, and invasion and promote apoptosis in U251 cells. mRNA sequencing identified 499 differentially expressed genes in vitexin-treated U251 cells relative to controls, including 154 upregulated genes and 345 downregulated genes. Gene ontology (GO) term enrichment analysis revealed that the upregulated genes were most significantly enriched in intrinsic apoptotic signaling pathway and the downregulated genes were most significantly enriched in positive regulation of cell development and positive regulation of locomotion relating to biological processes, endoplasmic reticulum lumen and side of membrane relating to cellular components, and receptor ligand activity and receptor regulator activity relating to molecular functions. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis revealed that the upregulated genes were involved in the pathways of transcriptional misregulation in cancer and the downregulated genes were involved in FoxO and JAK/STAT signaling pathways. Western blotting assay revealed that vitexin treatment resulted in reduced p-JAK1, p-JAK3, and p-STAT3 protein expression in U251 cells relative to untreated controls, and molecular docking predicted that vitexin had docking scores of –8.8, –10.8, and –10.5 kJ/mol with STAT3, JAK1, and JAK2, respectively. The results of the present study demonstrate that vitexin inhibits the proliferation and invasion and induces the apoptosis of glioblastoma U251 cells through suppressing the JAK/STAT3 signaling pathway, and vitexin may be a promising potential agent for the chemotherapy of glioblastoma.

Tunisian Native Mentha pulegium L. Extracts: Phytochemical Composition and Biological Activities.

Mint species (Lamiaceae family) have been used as traditional remedies for the treatment of several diseases. In this work, we aimed to characterize the biological activities of the total phenolic and flavonoid contents of Mentha pulegium L. extracts collected from two different regions of Tunisia. The highest amounts of total phenols (74.45 ± 0.01 mg GAE/g DW), flavonoids (28.87 ± 0.02 mg RE/g DW), and condensed tannins (4.35 ± 0.02 mg CE/g DW) were found in the Bizerte locality. Methanolic leaf extracts were subjected to HPLC-UV analysis in order to identify and quantify the phenolic composition. This technique allowed us to identify seven phenolic compounds: two phenolic acids and five flavonoid compounds, such as eriocitrin, hesperidin, narirutin, luteolin, and isorhoifolin, which were found in both extracts with significant differences between samples collected from the different regions (p < 0.05). Furthermore, our results showed that the methanolic extract from leaves collected from Bizerte had the highest antioxidant activities (DPPH IC50 value of 16.31 μg/mL and 570.08 μmol Fe2+/g, respectively). Both extracts showed high radical-scavenging activity as well as significant antimicrobial activity against eight tested bacteria. The highest antimicrobial activities were observed against Gram-positive bacteria with inhibition zone diameters and MIC values ranging between 19 and 32 mm and 40 and 160 µg/mL, respectively. Interestingly, at 10 μg/mL, the extract had a significant effect on cell proliferation of U87 human glioblastoma cells. These findings open perspectives for the use of Mentha pulegium L. extract in green pharmacy, alternative/complementary medicine, and natural preventive therapies for the development of effective antioxidant, antibacterial, and/or antitumoral drugs.

Nuclear receptor NR5A2 negatively regulates cell proliferation and tumor growth in nervous system malignancies

Nervous system malignancies are characterized by rapid progression and poor survival rates. These clinical observations underscore the need for novel therapeutic insights and pharmacological targets. To this end, here, we identify the orphan nuclear receptor NR5A2/LRH1 as a negative regulator of cancer cell proliferation and promising pharmacological target for nervous system-related tumors. In particular, clinical data from publicly available databases suggest that high expression levels of NR5A2 are associated with favorable prognosis in patients with glioblastoma and neuroblastoma tumors. Consistently, we experimentally show that NR5A2 is sufficient to strongly suppress proliferation of both human and mouse glioblastoma and neuroblastoma cells without inducing apoptosis. Moreover, short hairpin RNA-mediated knockdown of the basal expression levels of NR5A2 in glioblastoma cells promotes their cell cycle progression. The antiproliferative effect of NR5A2 is mediated by the transcriptional induction of negative regulators of the cell cycle, CDKN1A (encoding for p21cip1), CDKN1B (encoding for p27kip1) and Prox1 Interestingly, two well-established agonists of NR5A2, dilauroyl phosphatidylcholine (DLPC) and diundecanoyl phosphatidylcholine, are able to mimic the antiproliferative action of NR5A2 in human glioblastoma cells via the induction of the same critical genes. Most importantly, treatment with DLPC inhibits glioblastoma tumor growth in vivo in heterotopic and orthotopic xenograft mouse models. These data indicate a tumor suppressor role of NR5A2 in the nervous system and render this nuclear receptor a potential pharmacological target for the treatment of nervous tissue-related tumors.