Dynamic trajectory and cell communication of different cell clusters in malignant progression of glioblastoma / 北京大学学报(医学版)

Objective:To delve deeply into the dynamic trajectories of cell subpopulations and the communication network among immune cell subgroups during the malignant progression of glioblastoma(GBM),and to endeavor to unearth key risk biomarkers in the GBM malignancy progression,so as to provide a more profound understanding for the treatment and prognosis of this disease by integrating tran-scriptomic data and clinical information of the GBM patients.Methods:Utilizing single-cell sequencing data analysis,we constructed a cell subgroup atlas during the malignant progression of GBM.The Mono-cle2 tool was employed to build dynamic progression trajectories of the tumor cell subgroups in GBM.Through gene enrichment analysis,we explored the biological processes enriched in genes that significant-ly changed with the malignancy progression of GBM tumor cell subpopulations.CellChat was used to identify the communication network between the different immune cell subgroups.Survival analysis helped in identifying risk molecular markers that impacted the patient prognosis during the malignant pro-gression of GBM.This methodological approach offered a comprehensive and detailed examination of the cellular and molecular dynamics within GBM,providing a robust framework for understanding the disease's progression and potential therapeutic targets.Results:The analysis of single-cell sequencing data identified 6 different cell types,including lymphocytes,pericytes,oligodendrocytes,macrophages,glioma cells,and microglia.The 27 151 cells in the single-cell dataset included 3 881 cells from the pa-tients with low-grade glioma(LGG),10 166 cells from the patients with newly diagnosed GBM,and 13 104 cells from the patients with recurrent glioma(rGBM).The pseudo-time analysis of the glioma cell subgroups indicated significant cellular heterogeneity during malignant progression.The cell interaction analysis of immune cell subgroups revealed the communication network among the different immune sub-groups in GBM malignancy,identifying 22 biologically significant ligand-receptor pairs across 12 key bio-logical pathways.Survival analysis had identified 8 genes related to the prognosis of the GBM patients,among which SERPINE1,COL6A1,SPP1,LTF,C1S,AEBP1,and SAA1L were high-risk genes in the GBM patients,and ABCC8 was low-risk genes in the GBM patients.These findings not only provided new theoretical bases for the treatment of GBM,but also offered fresh insights for the prognosis assessment and treatment decision-making for the GBM patients.Conclusion:This research comprehensively and pro-foundly reveals the dynamic changes in glioma cell subpopulations and the communication patterns among the immune cell subgroups during the malignant progression of GBM.These findings are of significant im-portance for understanding the complex biological processes of GBM,providing crucial new insights for precision medicine and treatment decisions in GBM.Through these studies,we hope to provide more ef-fective treatment options and more accurate prognostic assessments for the patients with GBM.

Development and validation of a multi-modality fusion deep learning model for differentiating glioblastoma from solitary brain metastases / 中南大学学报(医学版)

Objective:Glioblastoma(GBM)and brain metastases(BMs)are the two most common malignant brain tumors in adults.Magnetic resonance imaging(MRI)is a commonly used method for screening and evaluating the prognosis of brain tumors,but the specificity and sensitivity of conventional MRI sequences in differential diagnosis of GBM and BMs are limited.In recent years,deep neural network has shown great potential in the realization of diagnostic classification and the establishment of clinical decision support system.This study aims to apply the radiomics features extracted by deep learning techniques to explore the feasibility of accurate preoperative classification for newly diagnosed GBM and solitary brain metastases(SBMs),and to further explore the impact of multimodality data fusion on classification tasks. Methods:Standard protocol cranial MRI sequence data from 135 newly diagnosed GBM patients and 73 patients with SBMs confirmed by histopathologic or clinical diagnosis were retrospectively analyzed.First,structural T1-weight,T1C-weight,and T2-weight were selected as 3 inputs to the entire model,regions of interest(ROIs)were manually delineated on the registered three modal MR images,and multimodality radiomics features were obtained,dimensions were reduced using a random forest(RF)-based feature selection method,and the importance of each feature was further analyzed.Secondly,we used the method of contrast disentangled to find the shared features and complementary features between different modal features.Finally,the response of each sample to GBM and SBMs was predicted by fusing 2 features from different modalities. Results:The radiomics features using machine learning and the multi-modal fusion method had a good discriminatory ability for GBM and SBMs.Furthermore,compared with single-modal data,the multimodal fusion models using machine learning algorithms such as support vector machine(SVM),Logistic regression,RF,adaptive boosting(AdaBoost),and gradient boosting decision tree(GBDT)achieved significant improvements,with area under the curve(AUC)values of 0.974,0.978,0.943,0.938,and 0.947,respectively;our comparative disentangled multi-modal MR fusion method performs well,and the results of AUC,accuracy(ACC),sensitivity(SEN)and specificity(SPE)in the test set were 0.985,0.984,0.900,and 0.990,respectively.Compared with other multi-modal fusion methods,AUC,ACC,and SEN in this study all achieved the best performance.In the ablation experiment to verify the effects of each module component in this study,AUC,ACC,and SEN increased by 1.6%,10.9%and 15.0%,respectively after 3 loss functions were used simultaneously. Conclusion:A deep learning-based contrast disentangled multi-modal MR radiomics feature fusion technique helps to improve GBM and SBMs classification accuracy.

Culture of glioblastoma U87 stem-like cells and identification of its metabolic phenotype and tumorigenic ability / 基础医学与临床

Objective To cultivate glioblastoma U87 stem-like cells(SLCs)and to detect the level of stemness bio-markers,mitochondrial respiratory capacity and the capacity of in vivo tumorigenesis.Methods B-27,growth factors EGF and bFGF was added into DMEM/F-12 culture in serum-free stem cell culture medium for U87 SLCs.Suspended culture of U87 SLCs was suspended using the neuro-sphere formation assay,while adherent culture of U87 SLCs was achieved by coating Matrigel matrix on the culture surface.The mRNA and protein level of stemness biomarkers in culture were detected using real-time quantitative PCR and Western blot.The proportion of CD133+cells in culture was detected by flow cytometry.The changes of cell oxygen consumption rate were detected by Seahorse cell metabo-lism analysis.Cell tumorigenesis ability was verified by subcutaneous tumor transplantation in animals.Results U87 SLCs in stem cell culture medium would grow into typical sphere morphology within one week,and the spheres would continue to grow as the culture process prolongs.At the appropriate concentration of adhesive,U87 SLCs adhered to and grow well in stem cell culture medium.The mRNA transcription of stemness biomarkers such as CD133,nes-tin,OLIG2,CD44,CD15,and integrin α6(ITGA6)was significantly increased as found in both culture methods,and the protein levels of CD133 and nestin were also increased under both methods(P＜0.05).U87 SLCs showed higher mitochondrial reserve respiratory capacity(P＜0.05).U87 SLCs could form larger subcutaneous tumors with fewer inoculated cells(P＜0.05),and grew faster in vivo with stronger tumorigenic ability.Conclusions U87 SLCs have typical stemness characteristics and may function as tumor cell model with higher stemness properties.

Research progress of CD73/NT5E in glioblastoma / 实用医学杂志

Glioma is the most common primary central nervous system tumor,mainly derived from glial cells,with strong invasiveness,easy recurrence,and poor prognosis.Glioblastoma is a high-grade glioma with the highest degree of malignancy.The clinical treatment method is mainly surgical resection,supplemented by compre-hensive treatment such as radiotherapy,chemotherapy,and electric field therapy,but the treatment effect is not satisfactory.In recent years,with the rapid development of the field of tumor immunotherapy,CD73 is a novel immune checkpoint related to adenosine metabolism,which can promote tumor progression by inhibiting anti-tumor immune responses and promoting angiogenesis.This article systematically reviews the mechanism of action of CD73 and discusses its biological role and application in glioma,aiming to provide potential treatment options for glioma patients.

NUDT5 promotes the growth, metastasis, and Warburg effect of IDH wild-type glioblastoma multiforme cells by upregulating TRIM47

@#Objective: To explore the regulatory mechanism of NUDT5 in glioblastoma multiforme (GBM). Methods: GEPIA database was used to predict the expressions of NUDT5 and tripartite motif family proteins 47 (TRIM47) in GBM patients. RT-qPCR and Western blot analyses were performed to examine NUDT5 expression in GBM cells. LN-229 cell proliferation, migration as well as invasion were estimated by CCK- 8, colony formation, wound healing, and Transwell assays following interference with NUDT5. ECAR assay, L-lactic acid kit, glucose detection kit, and ATP detection kit were applied for the detection of glycolysis-related indexes. Co-immunoprecipitation experiment was carried out to verify the relationship between NUDT5 and TRIM47. Results: GEPIA database showed that NUDT5 expression was significantly increased in GBM patients. Inhibiting the expression of NUDT5 in GBM cells significantly suppressed the viability, proliferation, invasion, migration, and glycolysis of GBM cells. Moreover, TRIM47 was highly expressed in GBM cells and interacted with NUDT5. Overexpression of TRIM47 partially reversed the inhibitory effect of NUDT5 downregulation on the proliferation, metastasis, and glycolysis of GBM cells. Conclusions: NUDT5 promotes the growth, metastasis, and Warburg effect of GBM cells by upregulating TRIM47. Both NUDT5 and TRIM47 can be used as targets for GMB treatment.

Establishment and validation of a method for determination of interesting protein expression level in cells infected with recombinant adeno-associated virus / 中国生物制品学杂志

@#Objective To establish and validate a method for the determination of the interesting protein expression level of recombinant adeno-associated virus(rAAV)infected cells,so as to monitor the product quality in different stages of rAAV9production process.Methods After incubation of serial diluted rAAV samples with infection enhancer Envirus-AAV,the human malignant glioblastoma cells(U87-MG)pretreated with hydroxyurea(HU)were infected.Using rAAV9 reference as the standard,the expression level of glutaryl-CoA dehydrogenase(GCDH)was detected by ELISA,and the specificity,accuracy,precision,linear range,limit of quantitation(LOQ)and durability of the method were verified.Eight batches of rAAV9 samples were detected by the established method.Results The A_(450)-A_(630) value of the sample buffer was 0.3,which was slightly lower than the lowest dilution point(1 ng/mL)of the four-parameter standard curve for protein quantification.The average recoveries of samples with 150%,100% and 50% theoretical relative titer levels were in the range of 100.0%-107.3%.The RSDs of the target protein expression level of the samples with three theoretical relative titer levels detected by the same experimenter three times and different experimenters were all less than 25%.There was a good linear relationship between rAAV9 samples and the target protein expression levels in the range of 50%-150% theoretical relative titer levels,and the linear regression equation was y = 1.077 x-0.022,R~2= 0.984.The LOQ of the method was 0.59,namely 6.0×10~(12) vg/mL.After U87-MG cells were incubated with HU for different time(18,21,24 h),and the culture supernatant was stored under different conditions(room temperature for 0.5 h,below-60 ℃ for 12 h,below-60 ℃ for 24 h).The RSDs of target protein expression levels were all less than 25%.The target protein expression levels of 1-8 batches of rAAV9 samples were 111%,121%,72%,65%,86%,75%,102% and 91%,respectively.Conclusion The established method for the determination of the target protein expression level after rAAV infection has good specificity,accuracy,precision and durability,and can be used for the quality control of products in different stages of rAAV9 production.

Screening ferroptosis associated with glioblastoma prognosis and immunity based on bioinformatics / 安徽医科大学学报

Objective @#To investigate the molecular mechanism of ferroptosis in glioblastoma (GBM) and to provide insights for identifying new therapeutic targets . @*Methods @#GSE108474 was selected from gene expression omnibus (GEO) database and differentially expressed genes (DEGs ) in GBM were obtained by using GEO2R , compared with the gene set in the Ferroptosis database (FerrDb) to identify ferroptosis related gene . GO and KEGG enrich ment analyses were conducted using DAVID database . A protein-protein interaction network was created using String web site . Hub genes with high connectivity were confirmed using Cytoscape software . Prognostic and immune infiltration analyses were performed using TIMER web site . RNA expression levels and gene correlation analyses were carried out using GEPIA web site . Differential expression of hub gene proteins was analyzed by using the HPA database. Tumor immune characteristic correlations were examined using TISIDB database . Differences in mRNA expression of hub genes between tumor cells A172 and U251MG and normal astrocytes HA1800 were compared u sing the quantitative real time PCR @*Results @#Out of 5 331 differentially expressed genes , 114 were related to fer roptosis . GO and KEGG enrichment analysis suggested that these 114 genes might play roles in positive regulation of gene expression , and affect tumor progression through ferroptosis and autophagy pathways . 10 hub genes were i dentified in the protein protein interaction network , among which cluster of differentiation 44 ( CD44 ) , murine double minute 2 (MDM2) and signal transducer and activator of transcription 3 (STAT3) were found to be highly expressed in tumors with lower survival rates . CD44 , MDM2 and STAT3 mRNA expression were higher in GBM cells compared to normal cells . Protein expression of CD44 , MDM2 and STAT3 was higher in high grade glioma tis sues than that in normal tissues . The expression of three genes in the tumor was negatively correlated with ferropto sis . Immune infiltration analysis revealed that CD44 , MDM2 and STAT3 in the tumor were related to the infiltration of neutrophils , CD4 + T cells , and dendritic cells , and the expression of three genes was related to various chemo kines and their receptors .@*Conclusion @#CD44 , MDM2 and STAT3 may play a role in tumor ferroptosis and immune regulation , which have the potential to become a therapeutic target for GBM .

Targeting Ferroptosis to Enhance Radiosensitivity of Glioblastoma / 生物化学与生物物理进展

Glioblastoma (GBM), one of the most common malignant tumors in the central nervous system (CNS), is characterized by diffuse and invasive growth as well as resistance to various combination therapies. GBM is the most prevalent type with the highest degree of malignancy and the worst prognosis. While current clinical treatments include surgical resection, radiotherapy, temozolomide chemotherapy, novel molecular targeted therapy, and immunotherapy, the median survival time of GBM patients is only about one year. Radiotherapy is one of the important treatment modalities for GBM, which relies on ionizing radiation to eradicate tumor cells. Approximately 60% to 70% of patients need to receive radiotherapy as postoperative radiotherapy or neoadjuvant radiotherapy during the treatment process. However, during radiotherapy, the radioresistant effect caused by DNA repair activation and cell apoptosis inhibition impedes the therapeutic effect of malignant glioblastoma.Ferroptosis was first proposed by Dr. Brent R. Stockwell in 2012. It is an iron-dependent mode of cell death induced by excessive lipid peroxidation. Although the application of ferroptosis in tumor therapy is still in the exploratory stage, it provides a completely new idea for tumor therapy as a novel form of cell death. Ferroptosis has played a significant role in the treatment of GBM. Specifically, research has revealed the key processes of ferroptosis occurrence, including intracellular iron accumulation, reactive oxygen species (ROS) generation, lipid peroxidation, and a decrease in the activity of the antioxidant system. Among them, glutathione peroxidase 4(GPX4) in the cytoplasm and mitochondria, ferroptosis suppressor protein 1 (FSP1) on the plasma membrane, and dihydroorotate dehydrogenase (DHODH) in the mitochondria constitute an antioxidant protection system against ferroptosis. In iron metabolism, nuclear receptor coactivator 4 (NCOA4) can mediate ferritin autophagy to regulate intracellular iron balance based on intracellular iron content. Heme oxygenase1 (HMOX1) catalyzes heme degradation to release iron and regulate ferroptosis. Radiation can trigger ferroptosis by generating ROS, inhibiting the signaling axis of the antioxidant system, depleting glutathione, upregulating acyl-CoA synthase long chain family member 4 (ACSL4), and inducing autophagy. Interestingly, some articles has documented that exposure to low doses of radiation (6 Gy for 24 h or 8 Gy for 4-12 h) can induce the expression of SLC7A11 and GPX4 in breast cancer and lung cancer cells, leading to radiation resistance, while radiation-induced ferroptosis occurs after 48 h. In contrast, high doses of ionizing radiation (20 Gy and 50 Gy) increase lipid peroxidation after 24 h. This suggests that radiation-induced oxidative stress is a double-edged sword that can regulate ferroptosis in both directions, and the ultimate fate of cells after radiation exposure——developing resistance and achieving homeostasis or undergoing ferroptosis——depends on the degree and duration of membrane lipid damage caused by the radiation dose. In addition, during the process of radiotherapy, methods such as inducing iron overload, damaging the antioxidant system, and disrupting mitochondrial function are used to target ferroptosis, thereby enhancing the radiosensitivity of glioblastoma. By promoting the occurrence of ferroptosis in tumor cells as a strategy to improve radiotherapy sensitivity, we can enhance the killing effect of ionizing radiation on tumor cells, thus providing more treatment options for patients with glioblastoma. In this paper, we reviewed ferroptosis and its mechanism, analyzed the molecular mechanism of radiation-induced ferroptosis, and discussed the effective strategies to regulate ferroptosis in enhancing the sensitivity of radiotherapy, with a view to providing an important reference value for improving the current status of glioblastoma treatment.

HDG inhibits the proliferation and migration of GBM cells and induces their apoptosis / 安徽医科大学学报

Objective @#To investigate the effects of hederagenin (HDG) on proliferation , migration , invasion and apoptosis of glioblastoma (GBM) cells and involved mechanism.@*Methods @#Human GBM cell lines U87 , U251 and human brain glial cell line (HEB) were selected as the study subjects , and HDG 0 μmol/L ( or 0 mg/kg) was used as the control group. MTT , EdU staining and cell plate cloning were used to detect the effect of HDG on the proliferation of GBM cells. Trypan blue staining was used to detect GBM cell death affected by HDG. The effects of HDG on migration and invasion of GBM cells were detected by cell scratch and Transwell assay. To analyze the effects of HDG on apoptosis of GBM cells , apoptosis⁃related proteins Bcl⁃2 , Bax , p53 and cleaved caspase⁃3 were detected by Western blot. Mitochondrial potential change was detected by JC⁃10 staining , and apoptotic cell count was displayed by Annexin V ⁃FITC staining. The effect of HDG on tumor bearing in GBM was analyzed by xeno transplantation in BALB/C mice. @*Results @#Compared with the control group (HDG 0 μmol/L) , HDG significantly inhibited the proliferation , migration and invasion of U87 and U251 cells , and they were dependent on the use dose of HDG. Trypan blue staining showed that HDG obviously increased death number of GBM cells. The mitochondrial potential of GBM cells was remarkedly decreased , the number of apoptotic GBM cells obviously increased , the expressions of apoptosis⁃related proteins p53 , Bax , cleaved⁃caspase3 were up⁃regulated and Bcl⁃2 was down⁃regulated by HDG in U87 and U251 cells. HDG significantly inhibited the size of subcutaneous GBM , the Ki67 positive rate of GBM cells and caused a large number of GBM cells to die in BALB/C mice. HDG had no obvious toxic effect on human HEB cells and the liver of tumor⁃bearing mice. @*Conclusion @#HDG can significantly inhibit the proliferation , migration and invasion of GBM cells and induce the apoptosis of them. The mechanism of HDG induced apoptosis of GBM cells may be through mitochondrial damage and regulation of p53 and Bcl⁃2/Bax expression.

nteraction between FAK/αB-crystalline is important for viability of the glioblastoma cells

Abstract Glioblastoma multiforme is a tumor of the central nervous system. Focal Adhesion Kinase (FAK) and αB-crystalline are two proteins involved in glioblastoma development. In this study, we investigated whether the FAK/αB-crystalline interaction is important for glioblastoma cells, we aimed to investigate the interaction of these two proteins in the glioblastoma multiforme cell line U87-MG. Two peptides named FP01 peptide (derived from αB-crystalline) and FP02 peptide (derived from FAK) were synthesized for this study. Treatment of U87-MG with the peptides FP01 and FP02 in the concentration at 50 µM reduced the viability cellular to around 41% and 51%, respectively. Morphological alterations in the cells treated with the peptides when compared to the control were observed. This study suggests that the interaction between FAK and αB-crystalline is important for the viability of glioblastoma cells

Effects of Apatinib combined with Temozolomide on levels of sPD-1 and sPD-L1 in patients with drug-resistant recurrent glioblastoma

Abstract Objective This study aimed to explore the effects of Apatinib combined with Temozolomide (TMZ) on the levels of Soluble PD-1 (sPD-1) and Soluble Programmed Death-1 Ligand (sPD-L1) in patients with drug-resistant recurrent Glioblastoma (GB). Study design A total of 69 patients with recurrent GB from September 2020 to March 2022 were recruited and assigned to the control group (n = 34) and observation group (n = 35) according to different treatment options after tumor recurrence. The control group was treated with TMZ, and the observation group was treated with Apatinib combined with TMZ. Levels of sPD-1 and spd-l1, clinical efficacy, survival time and adverse reactions were observed and compared between the two groups. Results General data including gender, age, body mass index, and combined diseases indicated no statistical significance between groups (p > 0.05). Before the intervention, sPD-1 and sPD-L1 levels were not significantly different in the two groups (p > 0.05). After interventions, levels of PD-1 and sPD-L1 levels decreased significantly (p < 0.05). The objective remission rate and clinical benefit rate of the observation group were higher and overall survival and progression-free survival were longer than those of the control group (p < 0.05). No significant difference was observed in major adverse reactions among patients (p > 0.05). Conclusions Apatinib combined with TMZ is safe and effective in the treatment of recurrent GB. The combined application of the two can reduce the levels of sPD-1 and sPD-L1, which has important clinical application value.

Role of cyclin D1 in glial tumors—A retrospective and observational study

Aim: High-grade glial tumors remain as one of the most lethal malignancies. Cyclin D1 is expressed in some human malignancies and is the potential target of intervention. The present study aims to determine the relationship of cyclin D1 expression with other clinicopathological parameters. Materials and Methods: A cross-sectional study was carried out in a tertiary care center. Biopsy proven 66 cases of glial tumor patients were included in the study. The patients with incomplete clinical details were excluded from the study. Immunohistochemistry using antibodies for IDH 1 and cyclin d1 was done in all the cases. Glial tumors were reclassified according to WHO 2016 classification. Data analysis was performed using SPSS 26.0 for the windows. Result: Among 66 patients, 49 (74.3%) were males and 17 (25.7%) were females. The age of the patients ranged from 20 years to 70 years. Overall, 6.02% were of grade I Glial tumors, 22.7% were of grade II Glial tumors, 19.6% patients were of grade III Glial tumors, and 51.6% patients were of grade IV Glial tumors. Of 66 samples tested cyclin D1 was positive in 25 (37.87%) as high expressers and 7 (10.60%) were low expressers. Our study showed a significant correlation between the expression of cyclin D1 with grade and IDH mutation status, No significant correlation of cyclin D1 was noted with age or sex of the patient. Conclusion: Cyclin D1 was associated with a higher grade of the glial tumor. It can be a potential marker both for prognosis and treatment of glial tumors.

The 2021 World Health Organization classification of gliomas: an imaging approach / Nova classificação das neoplasias gliais segundo a Organização Mundial da Saúde 2021, com enfoque radiológico

Abstract The purpose of this pictorial essay is to describe the recommendations of the 2021 World Health Organization classification for adult-type and pediatric-type gliomas and to discuss the main modifications in relation to the previous (2016) classification, exemplified by imaging, histological, and molecular findings in nine patients followed at our institutions. In recent years, molecular biomarkers have gained importance in the diagnosis and classification of gliomas, mainly because they have been shown to correlate with the biological behavior and prognosis of such tumors. It is important for neuroradiologists to familiarize themselves with this new classification of central nervous system tumors, so that they can use this knowledge in evaluating and reporting the imaging examinations of patients with glioma.

Resumo O propósito deste ensaio iconográfico é descrever e discutir as novas recomendações da Organização Mundial da Saúde de 2021, referente aos gliomas dos tipos adulto e infantil, e suas principais diferenças com a classificação anterior (2016), exemplificadas com imagens de nove casos de pacientes atendidos nas nossas instituições. Recentemente, há uma crescente significância dos marcadores moleculares no diagnóstico e classificação dos gliomas e tumores do sistema nervoso central, principalmente pela correlação com o comportamento biológico e o prognóstico. É importante que os neurorradiologistas estejam familiarizados com a nova classificação dos tumores do sistema nervoso central para a prática clínica, na avaliação e emissão de laudos e opiniões nas imagens dos pacientes com gliomas.

WHO CNS5 2021 incluye mutaciones específicas en gliomas que pueden ser identificadas con biomarcadores cuantitativos de resonancia magnética / WHO CNS5 2021 includes specific mutations in gliomas that can be identified with MRI quantitative biomarkers

Resumen En 2021 se publicó la última versión de la clasificación de tumores del sistema nervioso central de la Organización Mundial de la Salud (WHO CNS5 por sus siglas en inglés), considerada un estándar internacional. Las primeras ediciones se basaron en características histológicas y posteriormente se incorporaron aspectos relacionados con nuevos conocimientos. En la revisión de 2016 se implementaron características moleculares para la clasificación y estadificación de los gliomas, como la presencia de mutaciones en IDH1 y IDH2. Actualmente, las técnicas de resonancia magnética avanzada permiten valorar la presencia de 2-HG (oncometabolito incrementado ante mutaciones en IDH), de forma que indirectamente y sin procedimientos invasivos pueden identificarse las mutaciones en IDH. La resonancia magnética avanzada es un procedimiento aún en desarrollo, de gran utilidad para el diagnóstico y manejo de distintas patologías. En el presente documento se abordan las implicaciones de la WHO CNS5 en la evaluación de gliomas, así como aspectos históricos, las bases de la resonancia magnética convencional y secuencias de resonancia magnética avanzada útiles en la clasificación actual.

Abstract In 2021, the latest version of the World Health Organization classification of central nervous system tumors (WHO CNS5) was published, which is considered an international standard. The first editions of this classification were based on histological characteristics and, subsequently, aspects related to new knowledge were incorporated. In the 2016 revision, molecular characteristics were implemented for the classification and staging of gliomas, such as the presence of mutations in IDH1 or IDH2. Currently, advanced magnetic resonance imaging (MRI) techniques allow assessing for the presence of 2-HG (increased oncometabolite that precedes IDH mutations), whereby IDH mutations can be indirectly identified, without invasive procedures being required. Advanced MRI is a growing field, highly useful for diagnosis and management of different pathologies. This document addresses the implications of WHO CNS5 classification in the evaluation of gliomas, as well as historical aspects, the bases of conventional MRI, and advanced MRI sequences useful in current classification.

The effect and mechanism of Xihuang pill inhibits the formation of vasculogenic mimicry in human glioblastoma cells by down-regulating HIF-1α/VEGFA/VEGFR2 signaling pathway / 药学学报

Our studies were aimed to explore the effect and mechanism of the inhibition of the formation of vasculogenic mimicry (VM) in human glioblastoma cells by Xihuang pill (XHP) medicated serum through regulating the hypoxia inducible factor-1α (HIF-1α)/vascular endothelial growth factor A (VEGFA)/vascular endothelial growth factor receptor 2 (VEGFR2) signaling pathway. The medicated serum of XHP was prepared by gavage for 7 days to male SD rats (approval number of animal experiment ethics: 202105A051). The hypoxia model of U251 cells was established using 200 μmol·L-1 of CoCl2. After treatment with XHP-medicated serum, cell viability and proliferation of U251 cells were detected by CCK-8 and cell cloning experiment. Cell apoptosis and cell cycle of U251 cells were determined by flow cytometry. Cell migration and invasion were evaluated by wound healing and Transwell invasion assay. The formation of VM was assessed by three-dimensional cell culture of U251 cells. The protein expression levels of HIF-1α, VEGFA, VEGFR2, phosphorylated-VEGFR2 (p-VEGFR2), vascular endothelial-cadherin (VE-cadherin), Eph receptor tyrosine kinases A2 (EphA2), matrix metalloproteinase 2 (MMP2), matrix metalloproteinase 14 (MMP14) and laminin γ2 in U251 cells were detected by Western blot. The results showed that 10% XHP-medicated serum had little effect on the cell viability, proliferation, apoptosis and cell cycle of U251 cells under hypoxia. Compared with the model group, 10% XHP-medicated serum at 1.0, 1.5 and 2.0 h significantly decreased the migration rate (P < 0.01) and the number of invading U251 cells (P < 0.01). 10% XHP-medicated serum at 2.0 h significantly suppressed the formation of VM tubular structures in U251 cells under the condition of hypoxia (P < 0.01). Western blot experiment showed that 10% XHP-medicated serum significantly down-regulated the expression of HIF-1α, VEGFA, phospho-VEGFR2, VE-cadherin, EphA2 and MMP14 proteins (P < 0.05). In conclusion, XHP could inhibit the formation of VM in human glioblastoma U251 cells to suppress the angiogenesis by down-regulating the HIF-1α/VEGFA/VEGFR2 signaling pathway.

The action mechanism of glioblastoma cell-derived exosome: a review / 生物工程学报

Patients with glioblastoma (GBM) generally have a bad prognosis and short overall survival after being treated with surgery, chemotherapy or radiotherapy due to the histological heterogeneity, strong invasive ability and rapid postoperative recurrence of GBM. The components of GBM cell-derived exosome (GBM-exo) can regulate the proliferation and migration of GBM cell via cytokines, miRNAs, DNA molecules and proteins, promote the angiogenesis via angiogenic proteins and non-coding RNAs, mediate tumor immune evasion by targeting immune checkpoints with regulatory factors, proteins and drugs, and reduce drug resistance of GBM cells through non-coding RNAs. GBM-exo is expected to be an important target for the personalized treatment of GBM and a marker for diagnosis and prognosis of this kind of disease. This review summarizes the preparation methods, biological characteristics, functions and molecular mechanisms of GBM-exo on cell proliferation, angiogenesis, immune evasion and drug resistance of GBM to facilitate developing new strategies for the diagnosis and treatment of GBM.

Effect of Morus alba extract sanggenon C on growth and proliferation of glioblastoma cells / 中国中药杂志

Glioblastoma is the most common primary cranial malignancy, and chemotherapy remains an important tool for its treatment. Sanggenon C(San C), a class of natural flavonoids extracted from Morus plants, is a potential antitumor herbal monomer. In this study, the effect of San C on the growth and proliferation of glioblastoma cells was examined by methyl thiazolyl tetrazolium(MTT) assay and 5-bromodeoxyuridinc(BrdU) labeling assay. The effect of San C on the tumor cell cycle was examined by flow cytometry, and the effect of San C on clone formation and self-renewal ability of tumor cells was examined by soft agar assay. Western blot and bioinformatics analysis were used to investigate the mechanism of the antitumor activity of San C. In the presence of San C, the MTT assay showed that San C significantly inhibited the growth and proliferation of tumor cells in a dose and time-dependent manner. BrdU labeling assay showed that San C significantly attenuated the DNA replication activity in the nucleus of tumor cells. Flow cytometry confirmed that San C blocked the cell cycle of tumor cells in G_0/G_1 phase. The soft agar clone formation assay revealed that San C significantly attenuated the clone formation and self-renewal ability of tumor cells. The gene set enrichment analysis(GSEA) implied that San C inhibited the tumor cell division cycle by affecting the myelocytomatosis viral oncogene(MYC) signaling pathway. Western blot assay revealed that San C inhibited the expression of cyclin through the regulation of the MYC signaling pathway by lysine demethylase 4B(KDM4B), which ultimately inhibited the growth and proliferation of glioblastoma cells and self-renewal. In conclusion, San C exhibits the potential antitumor activity by targeting the KDM4B-MYC axis to inhibit glioblastoma cell growth, proliferation, and self-renewal.

Hypoxia-induced ROS aggravate tumor progression through HIF-1α-SERPINE1 signaling in glioblastoma / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Hypoxia, as an important hallmark of the tumor microenvironment, is a major cause of oxidative stress and plays a central role in various malignant tumors, including glioblastoma. Elevated reactive oxygen species (ROS) in a hypoxic microenvironment promote glioblastoma progression; however, the underlying mechanism has not been clarified. Herein, we found that hypoxia promoted ROS production, and the proliferation, migration, and invasion of glioblastoma cells, while this promotion was restrained by ROS scavengers N-acetyl-L-cysteine (NAC) and diphenyleneiodonium chloride (DPI). Hypoxia-induced ROS activated hypoxia-inducible factor-1α (HIF-1α) signaling, which enhanced cell migration and invasion by epithelial-mesenchymal transition (EMT). Furthermore, the induction of serine protease inhibitor family E member 1 (SERPINE1) was ROS-dependent under hypoxia, and HIF-1α mediated SERPINE1 increase induced by ROS via binding to the SERPINE1 promoter region, thereby facilitating glioblastoma migration and invasion. Taken together, our data revealed that hypoxia-induced ROS reinforce the hypoxic adaptation of glioblastoma by driving the HIF-1α-SERPINE1 signaling pathway, and that targeting ROS may be a promising therapeutic strategy for glioblastoma.

Resolving the lineage relationship between malignant cells and vascular cells in glioblastomas

Glioblastoma multiforme (GBM), a highly malignant and heterogeneous brain tumor, contains various types of tumor and non-tumor cells. Whether GBM cells can trans-differentiate into non-neural cell types, including mural cells or endothelial cells (ECs), to support tumor growth and invasion remains controversial. Here we generated two genetic GBM models de novo in immunocompetent mouse brains, mimicking essential pathological and molecular features of human GBMs. Lineage-tracing and transplantation studies demonstrated that, although blood vessels in GBM brains underwent drastic remodeling, evidence of trans-differentiation of GBM cells into vascular cells was barely detected. Intriguingly, GBM cells could promiscuously express markers for mural cells during gliomagenesis. Furthermore, single-cell RNA sequencing showed that patterns of copy number variations (CNVs) of mural cells and ECs were distinct from those of GBM cells, indicating discrete origins of GBM cells and vascular components. Importantly, single-cell CNV analysis of human GBM specimens also suggested that GBM cells and vascular cells are likely separate lineages. Rather than expansion owing to trans-differentiation, vascular cell expanded by proliferation during tumorigenesis. Therefore, cross-lineage trans-differentiation of GBM cells is very unlikely to occur during gliomagenesis. Our findings advance understanding of cell lineage dynamics during gliomagenesis, and have implications for targeted treatment of GBMs.

Endocannabinoids are potential inhibitors of glioblastoma multiforme proliferation / 中西医结合学报

Globally, it is evident that glioblastoma multiforme (GBM) is an aggressive malignant cancer with a high mortality rate and no effective treatment options. Glioblastoma is classified as the stage-four progression of a glioma tumor, and its diagnosis results in a shortened life expectancy. Treatment options for GBM include chemotherapy, immunotherapy, surgical intervention, and conventional pharmacotherapy; however, at best, they extend the patient's life by a maximum of 5 years. GBMs are considered incurable due to their high recurrence rate, despite various aggressive therapeutic approaches which can have many serious adverse effects. Ceramides, classified as endocannabinoids, offer a promising novel therapeutic approach for GBM. Endocannabinoids may enhance the apoptosis of GBM cells but have no effect on normal healthy neural cells. Cannabinoids promote atypical protein kinase C, deactivate fatty acid amide hydrolase enzymes, and activate transient receptor potential vanilloid 1 (TRPV1) and TRPV2 to induce pro-apoptotic signaling pathways without increasing endogenous cannabinoids. In previous in vivo studies, endocannabinoids, chemically classified as amide formations of oleic and palmitic acids, have been shown to increase the pro-apoptotic activity of human cancer cells and inhibit cell migration and angiogenesis. This review focuses on the biological synthesis and pharmacology of endogenous cannabinoids for the enhancement of cancer cell apoptosis, which have potential as a novel therapy for GBM. Please cite this article as: Duzan A, Reinken D, McGomery TL, Ferencz N, Plummer JM, Basti MM. Endocannabinoids are potential inhibitors of glioblastoma multiforme proliferation. J Integr Med. 2023; 21(2): 120-128.