RNA-Binding Motif Protein RBM47 Promotes Invasiveness of Glioblastoma Through Activation of Epithelial-to-Mesenchymal Transition Program.

Background: RNA-binding motif proteins (RBMs) have been widely implicated in the tumorigenesis of multiple human cancers but rarely investigated in glioblastoma (GBM). Methods: The expression level of RBM47 and its correlation with prognosis of GBM were examined using bioinformatics, quantitative reverse transcription PCR, and Western blot analysis. The colony formation assay and Cell Counting Kit-8 assay were used to determine the biological role of RBM47 in GBM. To measure invasiveness we used the wound healing assay and transwell assay. The regulatory relationship between RBM47 and the epithelial-to-mesenchymal transition (EMT) was examined by Western blot analysis and bioinformatic analysis. Results: Through integrative analysis of clinical proteomic and genomic tumor datasets, we found that RBM47 is significantly upregulated in GBM mesenchymal subtype, and its high expression is correlated with poor prognosis. In in vitro biological experiments, we observed a significant inhibitory effect of RBM47 knockdown on colony formation and cell growth using GBM cell lines. Conversely, overexpression of RBM47 restored and accelerated these processes. Moreover, in vitro, wound healing assays demonstrated the role of RBM46 in promoting and cell migration and invasion. Mechanistically, RBM47 enhances invasive capacity through the activation of the EMT program. In RBM47-knockdown cells, the expression levels of Vimentin and CD44 were suppressed, and the level of E-cadherin was increased. Conclusions: Taken together our results demonstrate the tumor promoting characteristics of RBM46 and suggest that it could be used both as a therapeutic target and prognostically.

Current status and outlook of potential applications of biodegradable materials in cerebral vascular stents.

The treatment of intracranial aneurysms (IAs) has undergone a very significant transformation in recent decades, and endovascular interventions have gradually become one of the most common treatments. As permanent metal stents can cause some degree of long-term damage to patients, biodegradable stent materials are emerging as attractive potential alternatives. By reviewing the current research status and the advantages and disadvantages of existing biodegradable biomaterials, this review expects to provide a valuable reference for subsequent research on biodegradable biomaterials.

Geometrical and Hemodynamic Characteristic Changes of Small Anterior Communicating Artery Aneurysms during Follow-ups in a Retrospective Analysis.

BACKGROUND: Small intracranial aneurysms have a low risk of rupture. However, ruptured anterior communicating artery (ACoA) aneurysms are usually smaller in clinical practice. The retrospective study aimed to investigate the geometrical and hemodynamic changes of small unruptured ACoA aneurysms during serial follow-ups. MATERIALS AND METHODS: We retrospectively collected patients with small unruptured ACoA aneurysms that were not repaired, who had serial follow-ups from the Electronic Medical Record System in four tertiary hospitals. The geometrical parameters of ACoA aneurysms were measured using a three-dimensional reconstructed model. Intra-aneurysmal hemodynamic parameters were computed using a high-resolution computational fluid dynamics model. Geometrical and hemodynamic changes of the aneurysms were evaluated at each follow-up. RESULTS: Five patients with small unruptured ACoA aneurysms that were not repaired were identified and included in this analysis. Aneurysms rupture occurred in two patients with aneurysm growth. The formation and enlargement of an irregular bleb at the aneurysm neck or dome were observed before the rupture. Ruptured aneurysms showed high wall shear stress (WSS) in the high inflow zone of aneurysm neck while low WSS and high oscillatory shear index (OSI) in the flow-recirculating region of aneurysm dome. Three unruptured aneurysms maintained a stable morphology and a physiological level of WSS. CONCLUSIONS: Aneurysm growth, low WSS, and high OSI at the dome and/or high WSS at the neck potentially contribute to the rupture of small ACoA aneurysms. These aneurysms should be considered for the treatment regardless of the small size.

A multi-dimensional non-uniform corrosion model for bioabsorbable metallic vascular stents.

Bioabsorbable metallic vascular stents (BMVSs) are an innovative technological advancement in the medical engineering field of vascular implants. BMVSs have great potential to revolutionize vascular intervention, but the lack of understanding of the construction material's natural corrosion within the body inhibits the use in clinical medicine. In this study, a corrosion function concept for in vivo implants was created to develop a multi-dimensional, non-uniform corrosion model with a larger goal of simulating the mechanical integrity of BMVSs. This proposed corrosion model simulates the corrosion rate and its effects on magnesium (Mg) alloy AZ31 based on continuum damage mechanics. The model was calibrated using three degradation experiments on Mg alloy specimens. These experiments focused on multi-dimensional corrosion, mass loss rate, and mechanical integrity during the corrosion process. Lastly, to verify the applicability of the proposed model, the resulting corrosion behaviors and mechanical characteristics of the BMVSs were implemented into a finite element framework to produce an overarching simulation of the BMVS's degradation in vivo. The results of the experiments and simulations revealed a proportional link between the corrosion of BMVSs and the number of exposed surfaces. A non-linear decline in mechanical integrity with increasing mass loss was also discovered through experimentation and modeling. Furthermore, the model and simulation can provide some details about changes in morphology and mechanics during BMVS corrosion. This work gives new insights into accurately modeling for BMVS degradation and can be used to optimize product development of BMVSs. STATEMENT OF SIGNIFICANCE: Bioabsorbable metallic vascular stents (BMVSs) are an innovative technological advancement in the medical engineering field of vascular implants. Despite BMVSs have great potential to revolutionize vascular intervention, the lack of understanding of the construction material's natural corrosion within the body inhibits their use in clinical medicine. In this study, a novel multi-dimensional non-uniform corrosion model was proposed to unveil the mechanisms during the in vivo degradation of bioabsorbable metallic implants, which can accurately capture the overlooked changes in morphology and mechanics during BMVS corrosion. This work provides a technical solution to enhance the modeling accuracy in BMVS degradation and can be further used to optimize the design of BMVSs in the future.

Study on the bending behavior of biodegradable metal cerebral vascular stents using finite element analysis.

Excellent bending behavior is evaluated as the primary factor during the design of biodegradable metal cerebral vascular stents (BMCVSs), which enables vascular stents to be successfully delivered to the targeted location and avoids unnecessary damage to blood vessels. Unfortunately, this bending behavior has been barely investigated which limits the design of BMCVSs with optimal structures. Herein, six BMCVSs were designed and their bending process were simulated using finite element analysis (FEA). Then, the effects of the stent bridge connection type and structure on the bending behavior were systematically analyzed and an universal mathematical model was further established, in which the influence of the structure parameters of the stent bridge on the flexibility of stents was considered. After that, the bending mechanism of the high-stress zone of the bridge was investigated. Finally, the causes and effects of the self-contacting phenomenon as well as the inner-stent protrusion phenomenon in the bending state were analyzed theoretically, and corresponding solutions were proposed to optimize the design of stents. The numerical results show that the stents with the dislocation-line W-shaped unit have better flexibility than the other stents. The flexibility is positively correlated to the cube of the length of linear part and to the square of the curvature of curved part. The self-contacting phenomenon of the bridge during bending can constrain the formation of inner-stent protrusion, which can eliminate the negative effects of the implanted stents on the hemodynamics in blood vessels. This study is expected to provide practical guidance for the structural design of BMCVSs for clinical applications.

Isoalantolactone inhibits IKKß kinase activity to interrupt the NF-κB/COX-2-mediated signaling cascade and induces apoptosis regulated by the mitochondrial translocation of cofilin in glioblastoma.

Isoalantolactone (IATL), a sesquiterpene lactone compound, possesses many pharmacological and biological activities, but its role in glioblastoma (GBM) treatment is still unknown. The aim of the current study was to investigate the antiglioma effects of IATL and to explore the underlying molecular mechanisms. In the current study, the biological functions of IATL were examined by MTT, cell migration, colony formation, and cell apoptosis assays. Confocal immunofluorescence techniques, chromatin immunoprecipitation, and pull-down assays were used to explore the precise underlying molecular mechanisms. To examine IATL activity and the molecular mechanisms by which it inhibits glioma growth in vivo, we used a xenograft tumor mouse model. Furthermore, Western blotting was used to confirm the changes in protein expression after IATL treatment. According to the results, IATL inhibited IKKß phosphorylation, thus inhibiting both the binding of NF-κB to the cyclooxygenase 2 (COX-2) promoter and the recruitment of p300 and eventually inhibiting COX-2 expression. In addition, IATL induced glioma cell apoptosis by promoting the conversion of F-actin to G-actin, which in turn activates the cytochrome c (Cyt c) and caspase-dependent apoptotic pathways. In the animal experiments, IATL reduced the size and weight of glioma tumors in xenograft mice and inhibited the expression of COX-2 and phosphorylated NF-κB p65 in the transplanted tumors. In conclusion, the current study indicated that IATL inhibited the expression of COX-2 through the NF-κB signaling pathway and induced the apoptosis of glioma cells by increasing actin transformation. These results suggested that IATL could be greatly effective in GBM treatment.

Expression and methylation status of LAMA2 are associated with the invasiveness of nonfunctioning PitNET.

The laminin subunit alpha 2 (LAMA2) gene encodes an alpha 2 chain, which constitutes one of the subunits of laminin 2 (merosin) and laminin 4 (s-merosin). In the current study, we investigated the relationship between LAMA2 promoter methylation status and the invasiveness of clinically nonfunctioning pituitary adenomas (PitNETs). Specimens from patients with nonfunctioning PitNET were classified into three groups according to preoperative computed tomography (CT)/magnetic resonance imaging findings: a normal group (n = 6), non-invasive group (n = 11) and invasive group (n = 6). LAMA2 expression was assessed using quantitative real-time polymerase chain reaction (RT-qPCR) and western blotting, and the methylation status of the LAMA2 promoter region was observed using sodium bisulfite sequencing. Furthermore, 5-aza-2-deoxycytidine was used to explore the relationship between decreased LAMA expression and methylation in PitNET cells. According to the RT-qPCR and western blotting results, LAMA2 expression was downregulated in invasive PitNET, while the methylation of the LAMA2 promoter was increased. Methylation of the LAMA2 promoter decreased the expression of LAMA2. Thus, changes in LAMA2 expression due to promoter methylation were inversely correlated with the invasiveness of PitNET and the protein functions as a tumor suppressor. In addition, overexpression and demethylation of LAMA2 suppressed the invasion of PitNET cells, partially by exerting effects on the PTEN-PI3K/AKT signaling pathway and matrix metalloproteinase-9 (MMP-9). Furthermore, a xenograft model was also generated, and LAMA2 overexpression significantly suppressed tumor growth in vivo. Thus, LAMA2 expression and methylation patterns might be used as biomarkers to predict the prognosis of patients with PitNET.

Silencing NUDT21 Attenuates the Mesenchymal Identity of Glioblastoma Cells via the NF-κB Pathway.

The proneural (PN) and mesenchymal (MES) subtypes of glioblastoma multiforme (GBM) are robust and generally consistent with classification schemes. GBMs in the MES subclass are predominantly primary tumors that, compared to PN tumors, exhibit a worse prognosis; thus, understanding the mechanism of MES differentiation may be of great benefit for the treatment of GBM. Nuclear factor kappa B (NF-κB) signaling is critically important in GBM, and activation of NF-κB could induce MES transdifferentiation in GBM, which warrants additional research. NUDT21 is a newly discovered tumor-associated gene according to our current research. The exact roles of NUDT21 in cancer incidence have not been elucidated. Here, we report that NUDT21 expression was upregulated in human glioma tissues and that NUDT21 promoted glioma cell proliferation, likely through the NF-κB signaling pathway. Gene set enrichment analysis, western blotting, and quantitative real-time reverse transcription polymerase chain reaction confirmed that NF-κB inhibitor zeta (NFKBIZ) was a downstream target affected by NUDT21 and that the MES identity genes in glioblastoma cells, CHI3L1 and FN1, were also differentially regulated. Our results suggest that NUDT21 is an upstream regulator of the NF-κB pathway and a potential molecular target for the MES subtype of GBM.

Microsurgical treatment of large and giant paraclinoid carotid aneurysms using a revised endovascular suction decompression technique with Invatec Mo.Ma device.

Endovascular retrograde suction decompression (RSD) with balloon occlusion of the internal carotid artery is helpful to facilitate clipping large and giant paraclinoid carotid aneurysms. The authors reported a revised endovascular technique without internal carotid access using Mo.Ma device and analyzed its feasibility. In the series, 15 consecutive patients harboring 15 large and giant paraclinoid carotid aneurysms were clipped with assistance of this revised RSD technique. The technical feasibility of the procedure, procedure-related complications, angiographic results, and clinical outcome were evaluated. Technical success was achieved in 14 patients with aneurysm neck clipping and internal carotid artery (ICA) patent. No complication related to this endovascular technique occurred. At follow-up (mean time 15.3months), the modified Rankin Scale score was excellent in 11 patients, good in two patients and poor in one patient. Their preliminary experience indicates that revised retrograde suction decompression technique with Mo.Ma device seems effective and safe in the surgical treatment of large and giant paraclinoid ICA aneurysms.

Reversion of malignant phenotypes of human glioblastoma cells by ß-elemene through ß-catenin-mediated regulation of stemness-, differentiation- and epithelial-to-mesenchymal transition-related molecules.

BACKGROUND: Glioblastoma is the most common and lethal type of primary brain tumor. ß-Elemene, a natural plant drug extracted from Curcuma wenyujin, has shown strong anti-tumor effects in various tumors with low toxicity. However, the effects of ß-elemene on malignant phenotypes of human glioblastoma cells remain to be elucidated. Here we evaluated the effects of ß-elemene on cell proliferation, survival, stemness, differentiation and the epithelial-to-mesenchymal transition (EMT) in vitro and in vivo, and investigated the mechanisms underlying these effects. METHODS: Human primary and U87 glioblastoma cells were treated with ß-elemene, cell viability was measured using a cell counting kit-8 assay, and treated cells were evaluated by flow cytometry. Western blot analysis was carried out to determine the expression levels of stemness markers, differentiation-related molecules and EMT-related effectors. Transwell assays were performed to further determine EMT of glioblastoma cells. To evaluate the effect of ß-elemene on glioblastoma in vivo, we subcutaneously injected glioblastoma cells into the flank of nude mice and then intraperitoneally injected NaCl or ß-elemene. The tumor xenograft volumes were measured every 3 days and the expression of stemness-, differentiation- and EMT-related effectors was determined by Western blot assays in xenografts. RESULTS: ß-Elemene inhibited proliferation, promoted apoptosis, impaired invasiveness in glioblastoma cells and suppressed the growth of animal xenografts. The expression levels of the stemness markers CD133 and ATP-binding cassette subfamily G member 2 as well as the mesenchymal markers N-cadherin and ß-catenin were significantly downregulated, whereas the expression levels of the differentiation-related effectors glial fibrillary acidic protein, Notch1, and sonic hedgehog as well as the epithelial marker E-cadherin were upregulated by ß-elemene in vitro and in vivo. Interestingly, the expression of vimentin was increased by ß-elemene in vitro; this result was opposite that for the in vivo procedure. Inhibiting ß-catenin enhanced the anti-proliferative, EMT-inhibitory and specific marker expression-regulatory effects of ß-elemene. CONCLUSIONS: ß-Elemene reversed malignant phenotypes of human glioblastoma cells through ß-catenin-involved regulation of stemness-, differentiation- and EMT-related molecules. ß-Elemene represents a potentially valuable agent for glioblastoma therapy.

Ursolic acid alleviates early brain injury after experimental subarachnoid hemorrhage by suppressing TLR4-mediated inflammatory pathway.

Our previous studies proved that ursolic acid (UA) protected against early brain injury (EBI) by modulating oxidative stress after experimental subarachnoid hemorrhage (SAH), but it has not been evaluated yet about its effects on an inflammatory pathway in a SAH model. This study was undertaken to evaluate the influence of UA on the toll-like receptor 4 (TLR4) signaling pathway after SAH. Adult male SD rats were divided into vehicle-treated sham, vehicle-treated SAH, and UA-treated SAH groups. The endovascular puncture model was used to induce SAH and all the rats were subsequently sacrificed at 48 h after SAH. UA administration markedly decreased the expressions of TLR4 pathway-related agents, such as intercellular adhesion molecule-1 (ICAM-1), TLR4, nuclear factor-κB (NF-κB) P65, interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), inducible nitric oxide synthase (iNOS) and matrix metalloproteinase (MMP)-9. Apoptosis detected by TUNEL indicated that fewer positive cells appeared in UA administration SAH groups than the control group. In conclusion, UA may attenuate EBI after SAH in rats by suppressing the TLR4-mediated inflammatory pathway.

Autophagy activation contributes to the neuroprotection of remote ischemic perconditioning against focal cerebral ischemia in rats.

Remote ischemic perconditioning (RIPer) has been proved to provide potent cardioprotection. However, there are few studies on neuroprotection of RIPer. This study aims to clarify the neuroprotective effect of RIPer and the role of autophagy induced by RIPer against cerebral ischemia reperfusion injury in rats. Using a transient middle cerebral artery occlusion (MCAO) model in rats to imitate focal cerebral ischemia. RIPer was carried out 4 cycles of 10 min ischemia and 10 min reperfusion, with a thin elastic band tourniquet encircled on the bilateral femoral arteries at the start of 10 min after MCAO. Autophagy inhibitor 3-methyladenine (3-MA) and autophagy inducer rapamycin were administered respectively to determine the contribution of autophagy in RIPer. Neurologic deficit scores, infarct volume, brain edema, Nissl staining, TUNEL assay, immunohistochemistry and western blot was performed to analyze the neuroprotection of RIPer and the contribution of autophagy in RIPer. RIPer significantly exerted neuroprotective effects against cerebral ischemia reperfusion injury in rats, and the autophagy-lysosome pathway was activated by RIPer treatment. 3-MA reversed the neuroprotective effects induced by RIPer, whereas rapamycin ameliorated the brain ischemic injury. Autophagy activation contributes to the neuroprotection by RIPer against focal cerebral ischemia in rats.

Ursolic acid reduces oxidative stress to alleviate early brain injury following experimental subarachnoid hemorrhage.

Ursolic acid (UA), a well-known anti-oxidative reagent, has been reported to protect the brain against ischemic stoke. However, the potential role of UA in protecting against early brain injury (EBI) after subarachnoid hemorrhage (SAH) remains unclear. The present study aimed to examine the effect of UA against EBI following SAH, and to demonstrate whether the effect is associated with its powerful antioxidant property. Male SD rats were divided into vehicle-treated sham, vehicle-treated SAH, and UA-treated SAH groups. The endovascular puncture model was used to induce SAH and all the rats were subsequently sacrificed at 48h after SAH. The results show that UA administration could significantly attenuate EBI (including brain edema, blood-brain barrier disruption, neural cell apoptosis, and neurological deficient) after SAH in rats and up-regulate the antioxidative levels in the rat cerebral cortex, suggesting that administration of UA in experimental SAH rats could alleviate brain injury symptom, potentially through its powerful antioxidant property. Hence, we concluded that UA might be a novel therapeutic agent for EBI following SAH.

ß-Elemene inhibits proliferation through crosstalk between glia maturation factor ß and extracellular signal­regulated kinase 1/2 and impairs drug resistance to temozolomide in glioblastoma cells.

ß-elemene, a plant-derived drug extracted from Curcuma wenyujin, has demonstrated marked antiproliferative effects on glioblastoma, while toxicity remains low. However, the underlying molecular mechanisms of the antitumor activity of ß-elemene remain to be elucidated. Previously, it was identified that the glia maturation factor ß (GMFß)/mitogen-activated protein kinase kinase (MAPK) 3/6/p38 pathway participates in the antiproliferative activity of ß-elemene on glioblastoma. In the present study, in order to illustrate the association of GMFß and the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway, U87 and U251 cells were treated with ß-elemene at various doses and for different durations, and the expression of phosphorylated ERK1/2 (p-ERK1/2), ERK1/2, B-cell lymphoma 2 (Bcl-2), Bcl2-associated X and survivin was examined by western blot analysis. Following treatment with ß-elemene and the ERK1/2 inhibitor PD98059, U87 cell viability was evaluated using a Cell Counting Kit-8 (CCK-8) assay, and the expression levels of Bcl-2 and survivin were examined by western blot analysis. GMFß was then downregulated by RNA interference in ß-elemene-treated U87 cells, and the effect of this on the expression of ERK1/2 and p-ERK1/2 was determined by western blot analysis. Finally, the chemosensitisation of U87 cells to temozolomide (TMZ) through ß-elemene was examined using the CCK-8 assay. The results demonstrated that ß-elemene inhibited the proliferation of U87 glioblastoma cells through the GMFß­dependent inactivation of the ERK1/2-Bcl-2/survivin pathway. Furthermore, inhibition of ERK1/2 by PD98059 enhanced the antitumor effect of ß-elemene and impaired the expression levels of Bcl-2 and survivin. ß-elemene also increased the sensitivity of U87 glioblastoma cells to the chemotherapeutic TMZ, which was synergistically enhanced by PD98059. In conclusion, these results suggested that GMFß-dependent inactivation of the ERK1/2-Bcl-2/survivin pathway mediated the antiproliferative effect of ß-elemene on glioblastoma. Therefore, ß-elemene is a promising chemosensitizer or adjuvant therapeutic for TMZ against glioblastoma brain tumors.

ß-elemene inhibits stemness, promotes differentiation and impairs chemoresistance to temozolomide in glioblastoma stem-like cells.

Accumulating evidence indicates that glioblastoma stem-like cells (GSCs) are key factors in tumour development, recurrence and chemoresistance. The impairment of stemness and the enhancement of differentiation contributes to the weakening of radiation and chemotherapy resistance of GSCs. We previously found that ß-elemene was an effective anti-glioblastoma agent and chemosensitizer. In this study, we examined the distribution of CD133(+) cells in human glioblastoma tissues by immunohistochemistry. Following treatment with ß-elemene, the formation of GSC spheres was investigated by manual counting, the proliferation of GSCs was measured with a Cell Counting Kit-8 (CCK-8) assay, and the dispersion of GSC spheres was observed with an inverted microscope. GSC spheres were treated with ß-elemene, and the expression levels of CD133, ATP-binding cassette subfamily G member 2 (ABCG2) and glial fibrillary acidic protein (GFAP) were examined by western blotting. After treatment with ß-elemene, the volumes and weights of GSC xenografts were measured, and the expression of CD133, ABCG2 and GFAP was evaluated through immunohistochemistry analysis. After treatment with ß-elemene and temozolomide (TMZ), GSC viability was examined by the CCK-8 assay, and the volumes and weights of xenografts were measured. We found that CD133(+) cells were assembled in some vascular walls and also sparsely distributed in other parts of glioblastoma tissues. ß-elemene decreased the formation of GSC spheres, dispersed GSC spheres and inhibited the proliferation of GSCs in vitro and in vivo. In the GSC spheres and xenografts treated with ß-elemene, the expression of CD133 and ABCG2 was significantly downregulated, and the expression of GFAP increased. Furthermore, the sensitivity of GSCs to TMZ was enhanced in vitro and in vivo. These results suggest that ß-elemene impaired the stemness of GSC spheres, promoted their differentiation and sensitized GSCs to TMZ. ß-elemene will hopefully become a valuable agent to enhance the effects of radiotherapy and chemotherapy.

Does closure of acid-sensing ion channels reduce ischemia/reperfusion injury in the rat brain?

Acidosis is a common characteristic of brain damage. Because studies have shown that permeable Ca(2+)-acid-sensing ion channels can mediate the toxic effects of calcium ions, they have become new targets against pain and various intracranial diseases. However, the mechanism associated with expression of these channels remains unclear. This study sought to observe the expression characteristics of permeable Ca(2+)-acid-sensing ion channels during different reperfusion inflows in rats after cerebral ischemia. The rat models were randomly divided into three groups: adaptive ischemia/reperfusion group, one-time ischemia/reperfusion group, and severe cerebral ischemic injury group. Western blot assays and immunofluorescence staining results exhibited that when compared with the one-time ischemia/reperfusion group, acid-sensing ion channel 3 and Bcl-x/l expression decreased in the adaptive ischemia/reperfusion group. Calmodulin expression was lowest in the adaptive ischemia/reperfusion group. Following adaptive reperfusion, common carotid artery flow was close to normal, and the pH value improved. Results verified that adaptive reperfusion following cerebral ischemia can suppress acid-sensing ion channel 3 expression, significantly reduce Ca(2+) influx, inhibit calcium overload, and diminish Ca(2+) toxicity. The effects of adaptive ischemia/reperfusion on suppressing cell apoptosis and relieving brain damage were better than that of one-time ischemia/reperfusion.

Β-elemene inhibits Hsp90/Raf-1 molecular complex inducing apoptosis of glioblastoma cells.

ß-Elemene, an active component of herb medicine Curcuma wenyujin, has been shown to antagonize glioblastoma cells by inducing apoptosis. However, how ß-elemene induces apoptosis of these cells remains unclear. In this study, we report that ß-elemene disrupted the formation of the Hsp90/Raf-1 complex, a key step in maintaining the conformation stability of Raf-1, and caused deactivation of Raf-1 and inhibition of the ERK pathway, thereby leading to apoptosis of glioblastoma cells. Specifically, treatment of glioblastoma cell lines with ß-elemene attenuated phosphorylation of multiple members of the kinase families in the Ras/Raf/MEK/ERK cascade, including Raf-1 and ERK as well as downstream signaling targets such as Bcl-2. These results suggest that the Hsp90/Raf-1 complex could be a promising molecular target for new drug development for the treatment of glioblastoma.

Lysosomal membrane permeabilization contributes to elemene emulsion-induced apoptosis in A549 cells.

Elemene is a broad-spectrum antitumor agent. In the present study, lysosomal membrane permeabilization (LMP) was detected after short elemene emulsion--exposure (12 h) that preceded a decrease of the mitochondrial membrane potential and DNA damage (24 h) in A549 cells. At later time points (36 h) elemene emulsion caused the appearance of A549 cells with apoptotic features, including apoptotic morphology, phosphatidylserine exposure, and caspase-3 activation. A significant increase in protein expression for cathepsin D was also observed utilizing Western blot analysis after exposure to elemene emulsion for 12 h. The present study showed that elemene emulsion induced the increased levels of reactive oxygen species (ROS) and depletion of glutathione (GSH) in A549 cells. Cells treated with pepstatin A, an inhibitor for cathepsin D, showed a significant inhibition in DNA damage, mitochondrial membrane permeabilization, caspase-3 activation, and phosphatidylserine exposure. These results demonstrated that apoptosis induced by elemene emulsion in A549 cells is mediated in part through LMP and lysosomal protease cathepsin D.

ß-elemene inhibits proliferation of human glioblastoma cells through the activation of glia maturation factor ß and induces sensitization to cisplatin.

ß-elemene, a natural drug extracted from Curcuma wenyujin, strongly inhibits glioblastoma growth. However, the mechanism of ß-elemene antitumor action remains unclear. Glia maturation factor ß (GMFß) regulates cellular growth, fission, differentiation and apoptosis. It has been reported that overexpression of GMFß inhibits the growth of glioblastoma cells and decreases tumor volume. To illustrate the role of GMFß in the anti-proliferative effect of ß-elemene in glioblastoma, U87 cells were treated with ß-elemene at various doses and for different periods of time, and levels of phospho-GMFß (p-GMFß) and total GMFß were determined by immunoprecipitation and western blot analysis. Upon GMFß silencing using RNA interference, the antitumor action of ß-elemene was evaluated in a methyl thiazolyl tetrazolium assay and by semi-quantitative western blot analysis of MKK3/6 and p-MKK3/6 expression. Finally, chemosensitization to cisplatin by ß-elemene was examined using a cell counting array, and the cell growth inhibitory rate was calculated. The results showed that ß-elemene inhibits U87 cell viability through the activation of the GMFß signaling pathway. Conversely, silencing the expression of GMFß reversed the antitumor effect of ß-elemene and impaired the phosphorylation of MKK3/6. Furthermore, ß-elemene increased the sensitivity of U87 glioblastoma cells to the chemotherapeutic agent cisplatin. Taken together, these results suggest that activation of the GMFß pathway mediates the antitumor effect of ß-elemene in glioblastoma. GMFß is a putative molecular target for glioblastoma therapy.

ß-Elemene inhibits proliferation of human glioblastoma cells and causes cell-cycle G0/G1 arrest via mutually compensatory activation of MKK3 and MKK6.

ß-elemene, a natural plant drug extracted from Curcuma wenyujin, has shown a strong anti-glioblastoma effect. However, the antitumor mechanism of ß-elemene remains unclear. Mitogen-activated protein kinase kinase-3 (MKK3) and -6 (MKK6) can regulate cellular growth, fission, differentiation and apoptosis. To illustrate the role of MKK3 and MKK6 in the anti-glioblastoma proliferation effect of ß-elemene, U87 cells were treated with ß-elemene at various doses or for different times, and then phosphorylated MKK3 (p-MKK3), phosphorylated MKK6 (p-MKK6), MKK3 and MKK6 were detected by Western blot assay. After transient transfection with dominant-negative mutant plasmids of MKK3 and MKK6, cell viability and cell cycle stage were determined by methyl thiazolyl tetrazolium assay and flow cytometry, respectively. Results showed that ß-elemene inhibited the proliferation of U87 glioblastoma cells and arrested them in G0/G1 phase through up-regulating p-MKK3 and p-MKK6 levels. In contrast, inhibition of MKK3 and MKK6 reversed the antitumor effect of ß-elemene. Furthermore, when either MKK3 or MKK6 was inhibited by a dominant-negative plasmid, the other was compensatorily activated in the presence of ß-elemene. Taken together, our findings indicate that mutually compensatory activation of MKK3 and MKK6 mediates the anti-glioblastoma effect of ß-elemene. MKK3 and MKK6 might be two putative targets for molecular therapy against glioblastoma.