human glioma cell u87 mg  (ATCC)

Journal: Cell Communication and Signaling : CCS

Article Title: EF24 targets METTL3 to reprogram m6A methylation and induce ferroptosis: an epitranscriptomic mechanism with therapeutical potential for glioma

doi: 10.1186/s12964-025-02583-4

Figure Lengend Snippet: EF24 inhibits glioma cell proliferation. A CCK-8 assay showing the inhibitory effect of EF24 on glioma cell viability and the IC50 values for LN229 and A172 cells. B Plate colony formation assay assessing the effect of EF24 on glioma cell proliferation. C Immunofluorescence analysis of the effect of EF24 on glioma cell proliferation (magnification ×200, scale bar: 50 μm). Note: * P < 0.05, ** P < 0.01, *** P < 0.001 compared to the control group

Article Snippet: Human glioma cell lines LN229 (RRID: CVCL-0393) and A172 (RRID: CVCL-0131) were procured from the American Type Culture Collection (ATCC; Manassas, VA, USA), cultured in Dulbecco’s Modified Eagle Medium (DMEM, Gibco, Thermo Fisher Scientific) supplemented with 10% heat-inactivated fetal bovine serum (FBS, Gibco) and 1% penicillin-streptomycin antibiotic cocktail (10,000 U/mL penicillin, 10 mg/mL streptomycin).

Techniques: CCK-8 Assay, Colony Assay, Immunofluorescence, Control

Journal: Cell Communication and Signaling : CCS

Article Title: EF24 targets METTL3 to reprogram m6A methylation and induce ferroptosis: an epitranscriptomic mechanism with therapeutical potential for glioma

doi: 10.1186/s12964-025-02583-4

Figure Lengend Snippet: METTL3 inhibits the ferroptosis in glioma cells. A Western blotting analysis of NRF2 and GPX4 in LN229 and A172 cells in the negative control group and the METTL3 knockdown group. B Correlation analysis of METTL3 expression with NRF2 and GPX4 mRNA levels in GBM tissue specimens. C Detection of intracellular malondialdehyde (MDA) levels after METTL3 knockdown. D Intracellular glutathione (GSH) levels following METTL3 knockdown. E TEM images showing mitochondrial morphological changes in METTL3-knockdown cells (magnification: ×7000, scale bar = 2.0 μm; magnification: ×20000, scale bar = 50 μm). F Detection of glioma proliferation following METTL3 knockdown by colony formation assay. Note: * P < 0.05, ** P < 0.01, *** P < 0.001 versus the control group

Article Snippet: Human glioma cell lines LN229 (RRID: CVCL-0393) and A172 (RRID: CVCL-0131) were procured from the American Type Culture Collection (ATCC; Manassas, VA, USA), cultured in Dulbecco’s Modified Eagle Medium (DMEM, Gibco, Thermo Fisher Scientific) supplemented with 10% heat-inactivated fetal bovine serum (FBS, Gibco) and 1% penicillin-streptomycin antibiotic cocktail (10,000 U/mL penicillin, 10 mg/mL streptomycin).

Techniques: Western Blot, Negative Control, Knockdown, Expressing, Colony Assay, Control

Journal: Cell Communication and Signaling : CCS

Article Title: EF24 targets METTL3 to reprogram m6A methylation and induce ferroptosis: an epitranscriptomic mechanism with therapeutical potential for glioma

doi: 10.1186/s12964-025-02583-4

Figure Lengend Snippet: METTL3 recruits YTHDF1 to regulate NRF2 mRNA stability. A Actinomycin assay showing the effect of YTHDF1 on NRF2 stability. B qRT-PCR showing the effect of knockdown of YTHDF1 on NRF2 expression. C Correlation analysis of YTHDF1 and NRF2 expression in GBM tissue specimens. D Western blotting showing the effect of knockdown of YTHDF1 on NRF2 expression. E RIP-qPCR revealing the binding of YTHDF1 with NRF2 in stable METTL3 knockdown and negative control LN229 and A172 cells. Note: * P < 0.05, ** P < 0.01, *** P < 0.001 compared to control

Article Snippet: Human glioma cell lines LN229 (RRID: CVCL-0393) and A172 (RRID: CVCL-0131) were procured from the American Type Culture Collection (ATCC; Manassas, VA, USA), cultured in Dulbecco’s Modified Eagle Medium (DMEM, Gibco, Thermo Fisher Scientific) supplemented with 10% heat-inactivated fetal bovine serum (FBS, Gibco) and 1% penicillin-streptomycin antibiotic cocktail (10,000 U/mL penicillin, 10 mg/mL streptomycin).

Techniques: Quantitative RT-PCR, Knockdown, Expressing, Western Blot, Binding Assay, Negative Control, Control

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Histone demethylase KDM9 activates the CCND1/AKT pathway to promote the malignant progression of gliomas through interaction with BRD2

doi: 10.1007/s00018-025-05900-9

Figure Lengend Snippet: KDM9 promotes malignant progression and reduces TMZ sensitivity in glioma. After using LN229 and SW1088 cells with knocked down KDM9 (LN229-shNC, LN229-shKDM9#1, LN229-shKDM9#2, SW1088-shNC, SW1088-shKDM9#1, SW1088-shKDM9#2) or the transfection of pcDNA3.1-KDM9 plasmid (KDM9-OE) in U251 cells for 48 h, ( A ) cell proliferation was measured via the CCK-8 assay. ( B ) Cell invasion was evaluated via the transwell assay. ( C ) The sphere-formation assay was conducted to assess the stem-like characteristics of glioma cells. ( D ) A Western blot experiment was performed to examine the expression of the stemness markers Nestin and CD133. ( E ) TMZ sensitivity was assessed via the colony formation assay after treatment with TMZ (100 μM) in glioma cells. ( F – H ) LN229-shKDM9 cells (1 × 10 7 ) were injected subcutaneously into nude mice to establish xenograft models ( N = 9). Tumor size ( F ), tumor volume ( G ), and tumor weight ( H ) were measured. ( I ) H&E and IHC staining were performed to detect the protein expression of KDM9 and Ki67 in the tumor tissues of nude mice. NC: negative control of shRNA, Vec: blank plasmid. Data are shown as the mean ± SD. ** P < 0.01 and *** P < 0.001

Article Snippet: The normal human brain glial cells HEB; the human glioma cells LN229 (ATCC: CRL-2611), SW1088 (ATCC: HTB-12), and T98G (ATCC: CRL-1690), U118 (ATCC: HTB-15), U251, and human embryonic kidney cells 293 T (ATCC: CRL-11268) were obtained from the Cell Center of Central South University.

Techniques: Transfection, Plasmid Preparation, CCK-8 Assay, Transwell Assay, Tube Formation Assay, Western Blot, Expressing, Colony Assay, Injection, Immunohistochemistry, Negative Control, shRNA

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Histone demethylase KDM9 activates the CCND1/AKT pathway to promote the malignant progression of gliomas through interaction with BRD2

doi: 10.1007/s00018-025-05900-9

Figure Lengend Snippet: H4K20me2 demethylation was mediated by KDM9 in the CCND1 promotor region to promote its transcription. RNA-Seq was performed on LN229 cells with knocked down KDM9, ( A ) Volcano plots of differential genes are shown, along with ( B ) bubble plots of enriched signaling pathways for differentially expressed genes. ( C ) ChIP-Seq was conducted using the H4K20me2 antibody in LN229 cells, and the distribution of reads around the transcription start site (TSS) was displayed. ( D ) The intersection between genes enriched in the PI3K/AKT pathway from RNA-Seq and genes obtained from ChIP-Seq is shown by a Venn diagram. ( E ) The intersecting genes are shown on a clustering heatmap. ( F , G ) qPCR was performed to detect mRNA expression of BCL2L11, BCL2, CCND1, and CREB3L2 in glioma cells after the knockdown or overexpression of KDM9. ( H ) Western blot experiment was performed to detect the protein expression of CCND1 in glioma cells with the knockdown or overexpression of KDM9. ( I ) Dual-luciferase reporter assay was performed in 293 T cells with KDM9 knockdown to detect CCND1 transcription activity. ( J ) Schematic diagram of the ChIP-qPCR primer (designed around the transcription start site of CCND1 from −2000 to + 300 bp). ( K ) ChIP-qPCR was used to measure the enrichment level of H4K20me2 at the CCND1 P4 promotor region (IgG was used as a negative control). NC: negative control for shRNA; Vec: blank plasmid. Data are shown as the mean ± SD. * P < 0.05, ** P < 0.01, and *** P < 0.001

Article Snippet: The normal human brain glial cells HEB; the human glioma cells LN229 (ATCC: CRL-2611), SW1088 (ATCC: HTB-12), and T98G (ATCC: CRL-1690), U118 (ATCC: HTB-15), U251, and human embryonic kidney cells 293 T (ATCC: CRL-11268) were obtained from the Cell Center of Central South University.

Techniques: RNA Sequencing, Protein-Protein interactions, ChIP-sequencing, Expressing, Knockdown, Over Expression, Western Blot, Luciferase, Reporter Assay, Activity Assay, ChIP-qPCR, Negative Control, shRNA, Plasmid Preparation

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Histone demethylase KDM9 activates the CCND1/AKT pathway to promote the malignant progression of gliomas through interaction with BRD2

doi: 10.1007/s00018-025-05900-9

Figure Lengend Snippet: KDM9 regulates CCND1 transcription through interaction with BRD2. ( A ) Protein silver staining of IP via KDM9 antibody enrichment in LN229 cells. ( B ) Transcription factors in the protein mass spectrometry data are shown. Exogenous and endogenous IP experiments were performed to analyze the interaction between KDM9 and BRD4 ( C ) or BRD2 ( D ). ( E ) The IF assay was used to detect the expression and co-localization of KDM9 and BRD2 in LN229 cells. ( F ) After the transfection of pcDNA3.1-BRD2-His into LN229-shKDM9 cells for 48 h, ChIP-qPCR was performed to detect the enrichment of H4K20me2 at the CCND1 P4 promoter region (with IgG as negative control). ( G ) After the transfection of wild-type or mutant KDM9 plasmids into U251 cell lines for 48 h, luciferase reporter assays were conducted to detect CCND1 transcription activity (Left). After the transfection of pcDNA3.1-BRD2-His into LN229-shKDM9 cells for 48 h, luciferase reporter assays were performed to detect CCND1 transcription activity (Right). ( H ) After the transfection of shBRD2 plasmids into U251-KDM9-WT cell lines for 48 h, luciferase reporter assays were conducted to detect CCND1 transcription activity. ( I ) After the transfection of pcDNA3.1-BRD2-His into shKDM9 glioma cells for 48 h, a Western blot experiment was conducted to detect the protein expression of KDM9, BRD2, CCND1, AKT/P-AKT, mTOR/P-mTOR, and CD133. Data are shown as the mean ± SD. * P < 0.05, ** P < 0.01, and *** P < 0.001

Article Snippet: The normal human brain glial cells HEB; the human glioma cells LN229 (ATCC: CRL-2611), SW1088 (ATCC: HTB-12), and T98G (ATCC: CRL-1690), U118 (ATCC: HTB-15), U251, and human embryonic kidney cells 293 T (ATCC: CRL-11268) were obtained from the Cell Center of Central South University.

Techniques: Silver Staining, Mass Spectrometry, Expressing, Transfection, ChIP-qPCR, Negative Control, Mutagenesis, Luciferase, Activity Assay, Western Blot

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Histone demethylase KDM9 activates the CCND1/AKT pathway to promote the malignant progression of gliomas through interaction with BRD2

doi: 10.1007/s00018-025-05900-9

Figure Lengend Snippet: KDM9 promotes glioma growth and reduces TMZ sensitivity in vivo. ( A - C ) LN229-shKDM9 or LN229-shNC cells (1 × 10 7 ) were injected subcutaneously into nude mice to establish xenograft models ( N = 6); after the tumor grew to 60-80 mm 3 , TMZ (40 mg/kg) was injected intraperitoneally for 5 consecutive days, and the tumor size ( A ), tumor volume ( B ) and tumor weight ( C ) were measured. ( D ) IHC staining was performed to detect the protein expression of KDM9, H4K20me2, CCND1, P-AKT, and CD133 in nude mouse tumor tissues. ( E ) The bioluminescence images of intracranial injection of LN229-luc-shKDM9 or LN229-luc-shNC cells or/and TMZ treatment in mice ( N = 6). The scale bar for bioluminescence intensity is shown on the right. ( F ) The quantification of the bioluminescence intensity of tumors. ( G ) Representative images of mouse brains stained with H&E at the termination of the experiment. ( H ) Kaplan–Meier survival curves of mice. Data are shown as the mean ± SD. * P < 0.05, ** P < 0.01, and *** P < 0.001

Article Snippet: The normal human brain glial cells HEB; the human glioma cells LN229 (ATCC: CRL-2611), SW1088 (ATCC: HTB-12), and T98G (ATCC: CRL-1690), U118 (ATCC: HTB-15), U251, and human embryonic kidney cells 293 T (ATCC: CRL-11268) were obtained from the Cell Center of Central South University.

Techniques: In Vivo, Injection, Immunohistochemistry, Expressing, Staining