Journal: Colloids and surfaces. B, Biointerfaces

Article Title: Photothermal induction of pyroptosis in malignant glioma spheroids using (16-mercaptohexadecyl)trimethylammonium bromide-modified cationic gold nanorods.

doi: 10.1016/j.colsurfb.2024.114128

Figure Lengend Snippet: Fig. 2. Optimization of NIR laser irradiation conditions. (A) Scheme of cell spheroid preparation and laser irradiation. (B) Confocal images of cell spheroid Sections (4 µm) prepared from U87 and GL261 cells, which were pre-incubated with different concentrations of MTAB-GNRs for 24 h (0 – 50 µM Au0) before spheroid formation. To visualize GNRs, a back-scattered light from the longitudinal LSPR mode of GNRs was utilized. Nuclei and F-actin were stained by 4′,6-diamidino-2- phenylindole (DAPI) and anti-actin antibody, respectively (Z-stack; scale bar, 10 μm). (C, D) FACS viability evaluation of single cells isolated from GL261 spheroids pre-treated with 50 µM (Au0) MTAB-GNRs (GNRs) or from control nanoparticle-free spheroids (Ctrl). Both types of spheroids were exposed to NIR laser light set to different laser power (0 – 15 W) for 5 min and cultured for another 5 h at 37◦C and 5 % CO2 before analysis. Cell viability was determined as the number of metabolically active (viable), calcein positive cells (C) or as the total number of dead cells positive to 7-AAD, Apopxin (PS), or both (D).

Article Snippet: Human glioblastoma cell line U-87 MG (U87; ATCC, Manassas, USA) and murine malignant glioma cell line GL261 (Leibniz Institute DSMZ, Braunschweig, Germany) were cultured in Dulbecco ̓s modified Eagle ̓s medium (DMEM) containing 4.5 g/L glucose (Biochrom AG, Berlin, Germany) and supplemented with 10 % foetal bovine serum (FBS; Invitrogen/Gibco, Grand Island, USA), penicillin (100 U/mL; Merck KGaA Darmstadt, Germany) and streptomycin (100 μg/mL; Merck KGaA, Darmstadt, Germany).

Techniques: Irradiation, Incubation, Staining, Isolation, Control, Cell Culture, Metabolic Labelling

Journal: Colloids and surfaces. B, Biointerfaces

Article Title: Photothermal induction of pyroptosis in malignant glioma spheroids using (16-mercaptohexadecyl)trimethylammonium bromide-modified cationic gold nanorods.

doi: 10.1016/j.colsurfb.2024.114128

Figure Lengend Snippet: Fig. 3. Photothermal ablation of 3D glioma tumor spheroids. (A, B) 3D tumor spheroids prepared from (A) U87 and (B) GL261 cells pre-incubated with 0 – 10 µM (Au0) MTAB-GNRs for 24 h and then irradiated with NIR laser beam set to laser power of 7 W for 5 min. Cell viability was determined by calcein (green) and 7-AAD (red) staining immediately (0 h) or 24 h after NIR laser light exposure (scale bar, 400 µm). (C) Representative FACS profiles of Apopxin/7-AAD staining of U87 or GL261 cells dissociated from spheroids pre-incubated with different concentrations of MTAB-GNRs (0 – 50 µM Au0) and irradiated with NIR laser light of 7 W laser power for 5 min. Spheroids were analyzed 5 h after irradiation. (D – G) Graphs of FACS analysis of irradiated spheroids expressing the number of calcein positive U87 (D) or GL261 (E) cells and the total number of PS-positive U87 (F) or GL261 (G) cells (Q2 and Q3) determined using Apopxin/7-AAD staining (quadrant Q1, PS-, 7- AAD+; quadrant Q2, PS+, 7-AAD+; quadrant Q3, PSS+, 7-AAD-; quadrant Q4, PSS-, 7-AAD-).

Article Snippet: Human glioblastoma cell line U-87 MG (U87; ATCC, Manassas, USA) and murine malignant glioma cell line GL261 (Leibniz Institute DSMZ, Braunschweig, Germany) were cultured in Dulbecco ̓s modified Eagle ̓s medium (DMEM) containing 4.5 g/L glucose (Biochrom AG, Berlin, Germany) and supplemented with 10 % foetal bovine serum (FBS; Invitrogen/Gibco, Grand Island, USA), penicillin (100 U/mL; Merck KGaA Darmstadt, Germany) and streptomycin (100 μg/mL; Merck KGaA, Darmstadt, Germany).

Techniques: Incubation, Irradiation, Staining, Expressing

Journal: Colloids and surfaces. B, Biointerfaces

Article Title: Photothermal induction of pyroptosis in malignant glioma spheroids using (16-mercaptohexadecyl)trimethylammonium bromide-modified cationic gold nanorods.

doi: 10.1016/j.colsurfb.2024.114128

Figure Lengend Snippet: Fig. 4. Assessment of cell recovery from photothermally induced stress. (A) Crystal violet staining of U87 and GL261 cell colonies formed from single cells dissociated from spheroids pre-treated with 0 – 5 µM (Au0) MTABGNRs and irradiated with NIR laser light set to 7 W for 5 min (day 13) and (B, C) calculation of the total area of cell colonies per dish in (B) U87 and (C) GL261 cells. Colony formation was calculated on day 9 – 13 after spheroids irradiation. Single cells from non- irradiated nanoparticle-free spheroids were used as controls (Ctrl).

Article Snippet: Human glioblastoma cell line U-87 MG (U87; ATCC, Manassas, USA) and murine malignant glioma cell line GL261 (Leibniz Institute DSMZ, Braunschweig, Germany) were cultured in Dulbecco ̓s modified Eagle ̓s medium (DMEM) containing 4.5 g/L glucose (Biochrom AG, Berlin, Germany) and supplemented with 10 % foetal bovine serum (FBS; Invitrogen/Gibco, Grand Island, USA), penicillin (100 U/mL; Merck KGaA Darmstadt, Germany) and streptomycin (100 μg/mL; Merck KGaA, Darmstadt, Germany).

Techniques: Cell Recovery, Staining, Irradiation

Journal: Colloids and surfaces. B, Biointerfaces

Article Title: Photothermal induction of pyroptosis in malignant glioma spheroids using (16-mercaptohexadecyl)trimethylammonium bromide-modified cationic gold nanorods.

doi: 10.1016/j.colsurfb.2024.114128

Figure Lengend Snippet: Fig. 5. Immunofluorescence-based analysis of proteins related to regulated cell death signaling. (A) Detection of the apoptosis marker cleaved caspase 3 on paraffin sections (4 µm) prepared from U87 cell spheroids that were pre-treated with 5 µM (Au0) MTAB-GNRs for 24 h, irradiated with 7 W of NIR laser light for 5 min, and subsequently cultured for different time (1 – 15 h). Sections from non-irradiated spheroids were used as controls (Ctrl; scale bar, 25 µm). (B) Immunofluorescence staining of cleaved caspase 1 on identically prepared sections from U87 and GL261 cell spheroids determined 1 and 3 h after laser irradiation (scale bar, 30 µm). (C) Protein expression level of the inflammasome sensor NLRP3 visualized 1 h post laser treatment of U87 cell spheroids containing MTAB-GNRs compared to non- irradiated control (scale bar, 25 µm). (D, E) Immunofluorescence detection of cleaved GSDMD on sections prepared from non-irradiated GL261 cell spheroids and from laser-exposed spheroids cultured after irradiation for an additional 5 h scanned by (D) fluorescence (scale bar, 25 µm) and (E) confocal microscope (Z-stack; scale bar, 7.5 µm). Back-scattered light was utilized for GNRs visualization. Nuclei were counter-stained with DAPI.

Article Snippet: Human glioblastoma cell line U-87 MG (U87; ATCC, Manassas, USA) and murine malignant glioma cell line GL261 (Leibniz Institute DSMZ, Braunschweig, Germany) were cultured in Dulbecco ̓s modified Eagle ̓s medium (DMEM) containing 4.5 g/L glucose (Biochrom AG, Berlin, Germany) and supplemented with 10 % foetal bovine serum (FBS; Invitrogen/Gibco, Grand Island, USA), penicillin (100 U/mL; Merck KGaA Darmstadt, Germany) and streptomycin (100 μg/mL; Merck KGaA, Darmstadt, Germany).

Techniques: Immunofluorescence, Marker, Irradiation, Cell Culture, Staining, Expressing, Control, Fluorescence, Microscopy

Journal: Journal for Immunotherapy of Cancer

Article Title: NKG2C/KLRC2 tumor cell expression enhances immunotherapeutic efficacy against glioblastoma

doi: 10.1136/jitc-2024-009210

Figure Lengend Snippet: Effect of KLRC2 overexpression in murine and human glioma. GFP cDNA-expressing GL261 or KLRC2 cDNA-expressing GL261 cells were injected into the striatum of C57BL/6 mice. ( A ) Western blot analysis quantified NKG2C expression in GL261 brain tumors with Rho as the normalization control (GFP, n=6; NKG2C, n=5). ( B ) Representative H&E staining of tumor sections. KLRC2 cDNA-expressing GL261 brain tumors show microcystic formations (arrow). ( C ) Representative immunohistochemical staining of tumor sections using a CD68 or a CD206-specific antibody. Quantification of each staining is shown above (n=5). ( D–F ) qRT-PCR analysis of the expression for several macrophage-related ( D ) immune-related ( E ) or KLRC2- associated ( F ) gene markers in GFP versus KLRC2 cDNA-expressing GL261 brain tumors (n=7). ( G–H ) Flow cytometry assessment of immune cell population percentages in human glioma categorized by KLRC2 expression levels (n=8). *p≤0.05, **p≤0.01, ***p≤0.001, ****p≤0.0001. Scale: 2.5 mm and 100 µm ( B ) 50 µm ( C ). cDNA, complementary DNA; GFP, green fluorescent protein; qRT-PCR, quantitative real time-polymerase chain reaction.

Article Snippet: The GL261 murine glioma cell line was purchased from Creative Biolabs and grown in Dulbecco's Modified Eagle Medium (DMEM) with 10% fetal bovine serum (FBS), 2 mM L-glutamine, 0.1% penicillin (100 U/mL), and streptomycin (100 μg/mL).

Techniques: Over Expression, Expressing, Injection, Western Blot, Control, Staining, Immunohistochemical staining, Quantitative RT-PCR, Flow Cytometry, Real-time Polymerase Chain Reaction

Journal: Journal for Immunotherapy of Cancer

Article Title: NKG2C/KLRC2 tumor cell expression enhances immunotherapeutic efficacy against glioblastoma

doi: 10.1136/jitc-2024-009210

Figure Lengend Snippet: KLRC2 cDNA expression enhances the antitumor immune response against a brain tumor after treatment with PD-1 mAb. ( A ) qRT-PCR-based KLRC2 expression in tumor tissues comparing GFP control with KLRC2 in the PD-1 mAb-treated group (n=6). ( B ) Mouse survival rates post-injection of GFP (n=7) or KLRC2 (n=9) GL261 cells and treatment that is depicted by Kaplan-Meier curves (p value=0.0145). ( C–D ) Representative immunohistochemical staining of tumor sections using CD68 and CD206 (n=5) ( C ) or CD3 and CD8 (GFP, n=4; NKG2C, n=7) ( D ) antibodies. Quantification of each staining is shown on the top. (E) Heatmap of normalized expression levels of genes associated with the immune environment. The heatmap’s color gradation corresponds to the relative expression levels of targeted markers. *p<0.05, **p<0.01. Scale: 50 µm ( C–D ). GFP, Green Fluorescent Protein; mAb, monoclonal antibodies; PD-1, programmed cell death protein-1; qRT-PCR, quantitative real time-polymerase chain reaction.

Article Snippet: The GL261 murine glioma cell line was purchased from Creative Biolabs and grown in Dulbecco's Modified Eagle Medium (DMEM) with 10% fetal bovine serum (FBS), 2 mM L-glutamine, 0.1% penicillin (100 U/mL), and streptomycin (100 μg/mL).

Techniques: Expressing, Quantitative RT-PCR, Control, Injection, Immunohistochemical staining, Staining, Bioprocessing, Real-time Polymerase Chain Reaction

Journal: Cancer Research Communications

Article Title: CAR T-cell Design-dependent Remodeling of the Brain Tumor Immune Microenvironment Modulates Tumor-associated Macrophages and Anti-glioma Activity

doi: 10.1158/2767-9764.crc-23-0424

Figure Lengend Snippet: FIGURE 2 Surface expression of 41BBL on CD28-based mB7-H3-CAR T-cells enhances effector cytokines release in repeat stimulation assay. Culture supernatants were collected at 24 hours after repeated stimulation with GL261 tumor cells at 2:1 ratio and analyzed using Multiplex assay. Summary plots of cytokines and chemokines produced by CAR T-cells after first stimulation (A) and fourth stimulation (B) against GL261 tumor cells (n = 4,

Article Snippet: The murine GL261 glioma cell line was procured from the Leibniz Institute (DSMZ-GermanCollection ofMicroorganisms andCell Cultures).

Techniques: Expressing, Multiplex Assay, Produced

Journal: Cancer Research Communications

Article Title: CAR T-cell Design-dependent Remodeling of the Brain Tumor Immune Microenvironment Modulates Tumor-associated Macrophages and Anti-glioma Activity

doi: 10.1158/2767-9764.crc-23-0424

Figure Lengend Snippet: FIGURE 3 CAR structural design significantly impacts anti-glioma efficacy of mB7-H3 CAR T-cells in the GL261 immunocompetent model. Albino C57BL/6 mice were transplanted with 1 × 105 GL261 cells orthotopically, followed 7 days later by intratumoral injection of 3 × 106 mB7-H3-CAR T-cells

Article Snippet: The murine GL261 glioma cell line was procured from the Leibniz Institute (DSMZ-GermanCollection ofMicroorganisms andCell Cultures).

Techniques: Injection

Journal: Cancer Research Communications

Article Title: CAR T-cell Design-dependent Remodeling of the Brain Tumor Immune Microenvironment Modulates Tumor-associated Macrophages and Anti-glioma Activity

doi: 10.1158/2767-9764.crc-23-0424

Figure Lengend Snippet: FIGURE 4 TIME heterogeneity after CAR T-cell treatment. A, Experimental scheme. Albino C57BL/6 mice were transplanted with 1 × 105 GL261 cells orthotopically, followed 25 days later by intratumoral injection of 3 × 106 mB7-H3-CAR T-cells (28.mζ, BBL-28.mζ,CD8tmBB.ζ, or Ctrl). Tumors were collected at 4 days after treatment and processed for scRNA-seq. Scheme created with BioRender.com. B, UMAP with major cell subsets in all tumor

Article Snippet: The murine GL261 glioma cell line was procured from the Leibniz Institute (DSMZ-GermanCollection ofMicroorganisms andCell Cultures).

Techniques: Injection

Journal: Cancer Research Communications

Article Title: CAR T-cell Design-dependent Remodeling of the Brain Tumor Immune Microenvironment Modulates Tumor-associated Macrophages and Anti-glioma Activity

doi: 10.1158/2767-9764.crc-23-0424

Figure Lengend Snippet: FIGURE 7 Global Mac/MG depletion abrogates effective CAR T-cell responses. GL261 glioma-bearing mice were treated with BLZ945 at 200 mg/kg starting 5 days after tumor implantation. A, Experimental scheme of BLZ945 macrophage depletion kinetics experiment. Daily drug dosing via oral gavage was for 2 weeks and tumors were harvested for FACS analysis at days 9, 16, and 20 after tumor implantation. B, Summary plot showing

Article Snippet: The murine GL261 glioma cell line was procured from the Leibniz Institute (DSMZ-GermanCollection ofMicroorganisms andCell Cultures).

Techniques: Tumor Implantation

Journal: Drug Delivery and Translational Research

Article Title: Local delivery of doxorubicin prodrug via lipid nanocapsule–based hydrogel for the treatment of glioblastoma

doi: 10.1007/s13346-023-01456-y

Figure Lengend Snippet: Proof-of-concept studies to evaluate the anti-tumor efficacy of prodrug DOXC 12 in orthotopic GL261-bearing mice. A Schematic representation of the therapeutic regimens tested in an orthotopic GL261 mouse model. DOXC 12 was intratumorally administered by CED at a dose of 2.5 mg/kg (50 μg/mouse) on day 15 or at a dose of 3.75 mg/kg (75 μg/mouse) on day 11. B Kaplan–Meier survival curves of mice after intratumoral administration of low dose/late treatments or high-dose/early treatment regimen with DOXC 12 ( n = 8–17) (MS median survival; * p < 0.05)

Article Snippet: Murine glioma GL261 cell line (DSMZ, Germany) was cultured in Eagle’s Minimum Essential Medium (EMEM; ATTC, USA) supplemented with penicillin G sodium (100 U/mL) and streptomycin sulfate (100 μg/mL) (Gibco, USA) and 10% Bovine Fetal Serum (Gibco, USA).

Techniques:

Journal: Drug Delivery and Translational Research

Article Title: Local delivery of doxorubicin prodrug via lipid nanocapsule–based hydrogel for the treatment of glioblastoma

doi: 10.1007/s13346-023-01456-y

Figure Lengend Snippet: Cytotoxicity of DOXC 12 and DOXC 12 -LNC CL in GL261 cells for 72 h. The cytotoxic effect of the treatments was assessed by crystal violet assay and presented in curve graph ( A ) and bar graph ( B ). The percentage of cell survival is compared to untreated cells (assumed as 100%) (mean ± SD, n = 3). Statistical analyses were performed using unpaired t -test for the IC 50 values ( A ) and two-way ANOVA with Bonferroni’s multiple comparisons test for bar graph ( B ) (** p < 0.01, **** p < 0.0001)

Article Snippet: Murine glioma GL261 cell line (DSMZ, Germany) was cultured in Eagle’s Minimum Essential Medium (EMEM; ATTC, USA) supplemented with penicillin G sodium (100 U/mL) and streptomycin sulfate (100 μg/mL) (Gibco, USA) and 10% Bovine Fetal Serum (Gibco, USA).

Techniques: Crystal Violet Assay

Journal: Drug Delivery and Translational Research

Article Title: Local delivery of doxorubicin prodrug via lipid nanocapsule–based hydrogel for the treatment of glioblastoma

doi: 10.1007/s13346-023-01456-y

Figure Lengend Snippet: Ex vivo anticancer efficacy of DOXC 12 -LNC CL in mouse organotypic brain slice model bearing GL261-GFP spheroids. A Images of mouse organotypic brain slices bearing GL261-GFP spheroids captured through fluorescent microscopy over time. B Fluorescence intensity of GL261-GFP the spheroids measured by fluorescent microscopy at different time points. Data were normalized to the initial intensity (time zero) and were reported as mean ± SEM using ImageJ software ( n = 4). Statistical analyses were performed using one-way ANOVA with Tukey’s multiple comparisons test (* p < 0.05)

Article Snippet: Murine glioma GL261 cell line (DSMZ, Germany) was cultured in Eagle’s Minimum Essential Medium (EMEM; ATTC, USA) supplemented with penicillin G sodium (100 U/mL) and streptomycin sulfate (100 μg/mL) (Gibco, USA) and 10% Bovine Fetal Serum (Gibco, USA).

Techniques: Ex Vivo, Slice Preparation, Microscopy, Fluorescence, Software

Journal: Drug Delivery and Translational Research

Article Title: Local delivery of doxorubicin prodrug via lipid nanocapsule–based hydrogel for the treatment of glioblastoma

doi: 10.1007/s13346-023-01456-y

Figure Lengend Snippet: In vivo efficacy studies in a GL261 GBM-resected mouse model. A Schematic diagram of the timeline of orthotopic tumor inoculation, resection, and treatment administration. DOXC 12 was locally administered at a dose of 5 mg/kg (5 μL of DOXC 12 -LNC CL hydrogel, 20 mg/mL). Ibuprofen was systemically administered post-surgery and every 24 h for 3 days at a dose of 30 mg/kg. B The Kaplan–Meier survival curves of mice treated with different interventions

Article Snippet: Murine glioma GL261 cell line (DSMZ, Germany) was cultured in Eagle’s Minimum Essential Medium (EMEM; ATTC, USA) supplemented with penicillin G sodium (100 U/mL) and streptomycin sulfate (100 μg/mL) (Gibco, USA) and 10% Bovine Fetal Serum (Gibco, USA).

Techniques: In Vivo

Journal: Drug Delivery and Translational Research

Article Title: Local delivery of doxorubicin prodrug via lipid nanocapsule–based hydrogel for the treatment of glioblastoma

doi: 10.1007/s13346-023-01456-y

Figure Lengend Snippet: In vivo efficacy studies in a GL261 resected mouse model: median survival (days), numbers of long-term survivors in each group, and statistical analysis

Article Snippet: Murine glioma GL261 cell line (DSMZ, Germany) was cultured in Eagle’s Minimum Essential Medium (EMEM; ATTC, USA) supplemented with penicillin G sodium (100 U/mL) and streptomycin sulfate (100 μg/mL) (Gibco, USA) and 10% Bovine Fetal Serum (Gibco, USA).

Techniques: In Vivo