human glioblastoma gbm ln229  (ATCC)

Journal: Cancers

Article Title: Tumor-Suppressive microRNA Therapy Inhibits Growth of Glioblastoma Multiforme Xenografts

doi: 10.3390/cancers18091479

Figure Lengend Snippet: FOXM1, eEF2K, and AXL expression in GBM patient tumors and patient survival in combined brain cohort. ( A – F ) The Rembrandt brain tumor patient dataset was evaluated and indicated that FOXM1 ( A ), AXL ( C ), and eEF2K ( E ) are overexpressed in GBM tumor samples compared to non-tumor, mixed glioma, oligodendroglioma and astrocytoma tumor samples. Data are presented as mRNA expression (log2). Kaplan–Meier survival analysis indicated that higher FOXM1 ( B ), eEF2K ( D ), and AXL ( F ) expression levels in combined brain tumor cohort are associated with shorter overall patient survival ( p < 0.0001, p = 0.0032, p < 0.0001). ( G ) Western blot analysis of GBM cell lines (LN229, U87, U373, U118) showed different levels of FOXM1, eEF2K, and AXL expression. Original western blots are presented in .

Article Snippet: The human glioblastoma multiforme cell lines U87, LN229, U373, and U118 were obtained from ATCC.

Techniques: Expressing, Western Blot

Journal: Cancers

Article Title: Tumor-Suppressive microRNA Therapy Inhibits Growth of Glioblastoma Multiforme Xenografts

doi: 10.3390/cancers18091479

Figure Lengend Snippet: miR-449b-5p, miR-329-3p and miR-518c suppress colony formation and spheroid formation. ( A , B ) Analysis of total area of colony formation of LN229, U87, U118 and U373 cells treated with miR-449b-5p, miR-329-3p and miR-518c or control miR. miR-449b-5p, miR-329-3p and miR-518c significantly suppressed colony formation area of LN229 ( p = 0.0087, p = 0.0019, p = 0.0150), U87 ( p < 0.0001, p < 0.0001, p = 0.0002), U118 cells ( p < 0.0001, p < 0.0001, p < 0.0001) and U373 cells ( p = 0.0002, p = 0.0002, p = 0.0021). ( C , D ) Analysis of spheroid formation ability of LN229 and U87 cells in ultra-low attachment plates. The cells were monitored for five days. miR-449b-5p, miR-329-3p and miR-518c significantly suppressed spheroid formation of LN229 ( p = 0.0042, p = 0.0016, p = 0.0290) and U87 cells ( p = 0.0087, p = 0.0235, p = 0.0087). Original western blots are presented in .

Article Snippet: The human glioblastoma multiforme cell lines U87, LN229, U373, and U118 were obtained from ATCC.

Techniques: Control, Western Blot

Journal: Translational Oncology

Article Title: CD38 overexpression drives glioblastoma progression via L1CAM/ICAM1/JAK-STAT-Driven tumor microenvironment rewiring

doi: 10.1016/j.tranon.2026.102758

Figure Lengend Snippet: A. Evaluation of CD38 expression following human and mouse GBM cell lines (A) and (B) overexpression of CD38 in T98G and Ln229 (human GBM cell lines) using western blot and fluorescent microscope imaging. (C) Significant overexpression of CD38 in transduced Gl-261 cells compared to wild type Gl-261 cells. B Kaplan-Meier overall survival analysis indicated significantly decreased in CD38-OE GBM- bearing mice (26.45 days) versus CD38-WT GBM- bearing mice (35.73 days) (p < 0.0001; n = 11 per group).

Article Snippet: Both T98G and LN229 human glioblastoma cell lines were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Expressing, Over Expression, Western Blot, Microscopy, Imaging

Journal: Translational Oncology

Article Title: CD38 overexpression drives glioblastoma progression via L1CAM/ICAM1/JAK-STAT-Driven tumor microenvironment rewiring

doi: 10.1016/j.tranon.2026.102758

Figure Lengend Snippet: A. Comparison of doubling time among CD38-OE, CD38-WT, and CD38-Knockdown human GBM cells (T98G and LN229) and CD38-OE, CD38-WT GL-261 cells displayed non-significant changes. (n = 3, triplicate). One-way ANOVA, no significant differences. B Western blot quantification of CKIs (p16, p21, p27) in GL-261 cells (n = 3). One-way ANOVA, no significant differences.

Article Snippet: Both T98G and LN229 human glioblastoma cell lines were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Comparison, Knockdown, Western Blot

Journal: Redox Biology

Article Title: SMURF2 attenuates NRF2-driven tumor progression by acting as a nuclear brake on NRF2 during cellular stress

doi: 10.1016/j.redox.2026.104102

Figure Lengend Snippet: SMURF2 prohibits the stress-mediated formation of ub + /p62 + aggresomes. (A and B) LN229 cells were transfected with HA or HA-SMURF2 and treated with DMSO; MG132 (10 μM, 12 h); H 2 O 2 (200 μM, 2 h); or LPS (100 ng/mL, 12 h). Representative immunofluorescence (IF) images of the colocalization of ub and p62. Nuclei stained with DAPI (A). Quantification the percentage of cells with ub + /p62 + puncta (B). (C – E) HEK293T cells were transfected with HA or HA-SMURF2 and subsequently treated with MG132 (10 μM, 12 h) or H 2 O 2 (200 μM, 2 h). Detergent-soluble and detergent-insoluble fractions were analyzed by western blotting with indicated antibodies (C and D). Quantification of the relative intensity of ub and p62 levels in the detergent-insoluble fractions shown in C and D (E). (F) LN229 cells were transfected with HA or HA-SMURF2 and treated with DMSO; MG132 (10 μM, 12 h); H 2 O 2 (200 μM, 2 h); or LPS (100 ng/mL, 12 h). Representative IF images of the colocalization of Proteostat and p62. Nuclei stained with DAPI. (G) The proposed model suggests that SMURF2 inhibits the formation of ub + /p62 + aggresomes/ALIS under stress conditions. Data were presented as the mean ± SD from three independent experiments. NS: not significant, ∗∗ p < 0.01, ∗∗∗ p < 0.001. Scale bar: 10 μm. Short Exp: short exposure; Long Exp: long exposure.

Article Snippet: The human glioblastoma cell lines LN229 and human embryonic kidney 293T (HEK293T) cell lines were purchased from the American Type Culture Collection (ATCC).

Techniques: Transfection, Immunofluorescence, Staining, Western Blot

Journal: Redox Biology

Article Title: SMURF2 attenuates NRF2-driven tumor progression by acting as a nuclear brake on NRF2 during cellular stress

doi: 10.1016/j.redox.2026.104102

Figure Lengend Snippet: SMURF2 nuclear translocation facilitates the degradation of NRF2 within the nucleus. (A and B) HEK293T cells were first transfected with HA-NRF2-WT. Subsequently, cells were transfected either with Flag or Flag-SMURF2 (A), or with SMURF2 siRNA or scrambled siRNA oligos for 48 h (B). Cells were then treated with H 2 O 2 (200 μM, 2 h) or LPS (100 ng/mL, 12 h). Following treatment, subcellular fractionation was performed to isolate total, nuclear, and cytoplasmic proteins, followed by western blot analysis with indicated antibodies. (C and D) LN229 cells were transfected with HA-SMURF2, then treated with PBS, H 2 O 2 (200 μM, 2 h) (C) or LPS (100 ng/mL, 12 h) (D). Representative IF images showing the subcellular localization of HA-SMURF2 are presented; nuclei were stained with DAPI. (E and F) HEK293T cells were treated with PBS, H 2 O 2 (200 μM, 2 h) (E), or LPS (100 ng/mL, 12 h) (F). Subcellular fractionation was performed to isolate total, nuclear, and cytoplasmic proteins, followed by western blot analysis with indicated antibodies. (G) Schematic depiction of NRF2 NLS1/2−Mut , NRF2 NES1/2−Mut , and SMURF2 NLS−Mut . (H–K) HEK293T cells were first transfected with HA-NRF2 NLS2−Mut (H and I) or HA-NRF2 NES2−Mut (J and K). Subsequently, cells were transfected either with Flag or Flag-SMURF2, then treated with H 2 O 2 (200 μM, 2 h) (H and J) or LPS (100 ng/mL, 12 h) (I and K). Following treatment, subcellular fractionation was performed to isolate total, nuclear, and cytoplasmic proteins, followed by western blot analysis with indicated antibodies. (L) Schematic of SMURF2 domains with NLS indicated (single-letter code). SMURF2 NLS−Mut denotes deletion of NLS residues (Top). LN229 cells transfected with Flag-SMURF2-WT or Flag-SMURF2 NLS−Mut were treated with DMSO, MG132 (10 μM, 12 h), H 2 O 2 (200 μM, 2 h), or LPS (100 ng/mL, 12 h). Representative IF images showing subcellular localization of Flag-SMURF2 or Flag-SMURF2 NLS−Mut ; nuclei were stained with DAPI (Bottom). (M) HEK293T cells were first transfected with HA-NRF2-WT. Subsequently, cells were transfected either with Flag or Flag-SMURF2 NLS−Mut , then treated with H 2 O 2 (200 μM, 2 h) or LPS (100 ng/mL, 12 h). Following treatment, subcellular fractionation was performed to isolate total, nuclear, and cytoplasmic proteins, followed by western blot analysis with indicated antibodies. (N) The proposed model indicates that SMURF2 specifically degrades NRF2 in the nucleus. Scale bar: 5 μm, Scale bar: 10 μm. Data were presented in three independent experiments.

Article Snippet: The human glioblastoma cell lines LN229 and human embryonic kidney 293T (HEK293T) cell lines were purchased from the American Type Culture Collection (ATCC).

Techniques: Translocation Assay, Transfection, Fractionation, Western Blot, Staining

Journal: Redox Biology

Article Title: SMURF2 attenuates NRF2-driven tumor progression by acting as a nuclear brake on NRF2 during cellular stress

doi: 10.1016/j.redox.2026.104102

Figure Lengend Snippet: SMURF2 promotes cell apoptosis through NRF2 inactivation. (A) LN229 cells were transfected with HA, HA-SMURF2, Myc-NRF2 or HA-SMURF2 + Myc-NRF2 and subsequent treatment with MG132 (10 μM, 12 h), H 2 O 2 (200 μM, 2 h), or LPS (100 ng/mL, 12 h). Representative IF images of the colocalization of ub and p62. Nuclei stained with DAPI. (B and C) HEK293T cells were transfected with empty vector or Myc-NRF2, followed by Flag or Flag-SMURF2 expression. Western blotting was performed using indicated antibodies (B). qRT-PCR was performed using primers specific for indicated genes (C). The fold change in expression in Flag-SMURF2 overexpressing samples was calculated relative to control samples. (D) HEK293T cells were first transfected with NRF2 siRNA for 48 h, followed by transfection with either Myc-NRF2-WT or Myc-NRF2-K555R, and subsequently transfected with Flag or Flag-SMURF2.Western blotting was performed using indicated antibodies. (E) LN229 cells were transfected with SMURF2 siRNA or scramble siRNA oligos for 48 h, followed by treatment with PBS or H 2 O 2 (200 μM, 2 h). Cells were then stained with 2,7-Dichlorodihydrofluorescein diacetate (DCFH-DA, 10 μM) and the ROS level was detected by flow cytometry. (F) Quantification of relative ROS fluorescence intensity in LN229 cells with SMURF2 knockdown or HA-SMURF2 overexpression. (G and H) LN229 cells were transfected with SMURF2 siRNA or scramble siRNA oligos for 48 h or transfected with HA or HA-SMURF2, followed by treatment with PBS or H 2 O 2 (200 μM, 2 h). Cells were then stained with Annexin-V/propidium iodide (PI), and apoptotic cells were detected by flow cytometry (G). Quantification of apoptosis in LN229 cells with SMURF2 knockdown or HA-SMURF2 overexpression (H). (I) HEK293T cells were transfection with Flag or Flag-SMURF2, and subsequent treatment with or without MG132 (10 μM, 12 h), H 2 O 2 (200 μM, 2 h), or LPS (100 ng/ml, 12 h). Western blotting was performed using indicated antibodies. (J and K) HEK293T cells were transfected with empty vector and Myc-NRF2 (J) or Myc-NRF2-WT, Myc-NRF2-K555R (K), followed by Flag or Flag-SMURF2 expression. Cells were subsequently treated with MG132 (10 μM, 12 h), H 2 O 2 (200 μM, 2 h), or LPS (100 ng/mL, 12 h). Western blotting was performed using indicated antibodies. (L) The shSMURF2 and shPLKO cells were treated with or without LPS (100 ng/mL, 12 h) and cultured for 14 days. Colony formation assay was performed for cells. (M) The representative images of shSMURF2 and shPLKO patient-derived cells formed tumors in nude mice with or without LPS (10 mg/kg). (N) The graph showed the quantified data of tumor weight. (O) Immunohistochemistry (IHC) analysis of tumor tissue slides with antibodies against Ki67. Nucleus was stained by hematoxylin. Scale bar, 50 μm (P) The proposed model indicates that SMURF2 promotes cell apoptosis through NRF2 inactivation. Data were presented as the mean ± SD from three independent experiments. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. Scale bar: 10 μm, Scale bar, 50 μm.

Article Snippet: The human glioblastoma cell lines LN229 and human embryonic kidney 293T (HEK293T) cell lines were purchased from the American Type Culture Collection (ATCC).

Techniques: Transfection, Staining, Plasmid Preparation, Expressing, Western Blot, Quantitative RT-PCR, Control, Flow Cytometry, Fluorescence, Knockdown, Over Expression, Cell Culture, Colony Assay, Derivative Assay, Immunohistochemistry

Journal: Cancers

Article Title: Piezo1-ATF3-PPP1r15a Axis Transduces Mechanical Stress into Apoptosis in Glioma Under Low-Intensity Focused Ultrasound

doi: 10.3390/cancers18091445

Figure Lengend Snippet: ATF3 is involved in the ERS of GBM cells treated with LIFU. ( A ) Annexin-V/PI bound U251 or LN229 cells were counted. ( B ) The corresponding statistical results of apoptosis rate. ( C ) The expression levels of Bax, Bcl-2 and caspase3 were determined by Western blot. ( D ) The expression levels of ATF3, ATF4 and CHOP were determined by Western blot. ( E ) U251 and LN229 cells were subjected to reverse transcription-quantitative PCR analysis for ATF4. ( F ) U251 and LN229 cells were subjected to reverse transcription-quantitative PCR analysis for CHOP. ( G ) Representative images of JC-1 staining in U251 with different treatments. ( H ) Representative images of JC-1 staining in LN229 with different treatments. The data are presented as the means ± SEM from at least three independent biological replicates. Statistical comparisons among multiple groups (control, LIFU, siATF3, LIFU + siATF3) were performed using two-way ANOVA followed by Šídák’s multiple comparisons test. *** p < 0.001; **** p < 0.0001. ER, endoplasmic reticulum; GBM, glioblastoma; LIFU, low-intensity focused ultrasound. The original Western blot figures can be found in .

Article Snippet: The American Type Culture Collection provided GBM cell lines (U251 and LN229) and the 293T cell line.

Techniques: Expressing, Western Blot, Reverse Transcription, Real-time Polymerase Chain Reaction, Staining, Control