Journal: Journal of Cerebral Blood Flow & Metabolism

Article Title: Mutant KRAS in brain endothelial cells promotes vascular inflammation and impairs vascular integrity in brain arteriovenous malformation

doi: 10.1177/0271678X261421424

Figure Lengend Snippet: MG corroborate with KRAS-G12V-EC to promote the loss of EC integrity in vitro. (a) A schematic diagram of the experiment. KRAS-G12V-EC-CM (conditioned medium) was added to BV2-MG and incubated for 48 h ( for b ), and then BV2-CM was transferred to control EC for 48 h ( for c ), or KRAS-G12V-EC-CM was added to control EC for 48 hours ( for d ); (b) Bar graphs showing increased mRNA levels in inflammatory cytokines (IL-6, IL-1β), proteolytic enzymes (MMP-9), and the MG-survival factor (CSF1R) in KRAS-G12V-EC-CM treated BV2-MG; (c) Bar graphs showing decreased mRNA levels of junction markers, CDH5 and TJP1, in EC after 2 days of incubation with BV2-CM, while OCLN and CLDN5 did not change; (d) Bar graphs showing non-significant changes in mRNA levels of junction markers in EC after 2 days of incubation with KRAS-G12V-EC-CM. Unpaired t -test. ns indicates no significant differences. All experiments are repeated three times. * p < 0.05. ** p < 0.01.

Article Snippet: BV2 murine MG (CRL-2468; ATCC, Manassas, VA, USA) were cultured at passage four with a medium containing high glucose (CM002-050; GenDEPOT, Katy, TX, USA), 10% fetal bovine serum (F0901-050; GenDEPOT), and penicillin-streptomycin (CA005; GenDEPOT) at 37°C in a 5% CO 2 humidified incubator.

Techniques: In Vitro, Incubation, Control

Journal: Journal of Cerebral Blood Flow & Metabolism

Article Title: Mutant KRAS in brain endothelial cells promotes vascular inflammation and impairs vascular integrity in brain arteriovenous malformation

doi: 10.1177/0271678X261421424

Figure Lengend Snippet: Minocycline attenuates KRAS-G12V-EC-induced MG migration in vitro. (a) BV2-MG transmigration activities were tested using a transwell insert (please see the detailed procedure in the section “Materials and methods”); (b) Immunofluorescence staining showing accelerated BV2-MG migration 12 h later by KRAS mutation in ECs. Scale bars = 20 μm; (c, d) Minocycline (100 μM) or PBS was treated on ECs 1 day post-mutant KRAS (or GFP) transfection to test the effects on BV2-MG migration. Bar graphs showing migrated Iba1 + MG in the bottom well at 4 days post-minocycline treatment, compared to PBS. ANOVA with Turkey’s multiple comparisons test. Scale bars = 100 μm. *** p < 0.001.

Article Snippet: BV2 murine MG (CRL-2468; ATCC, Manassas, VA, USA) were cultured at passage four with a medium containing high glucose (CM002-050; GenDEPOT, Katy, TX, USA), 10% fetal bovine serum (F0901-050; GenDEPOT), and penicillin-streptomycin (CA005; GenDEPOT) at 37°C in a 5% CO 2 humidified incubator.

Techniques: Migration, In Vitro, Transmigration Assay, Immunofluorescence, Staining, Mutagenesis, Transfection

Journal: International Journal of Molecular Sciences

Article Title: A Set of Dysregulated Target Genes to Reduce Neuroinflammation at Molecular Level

doi: 10.3390/ijms23137175

Figure Lengend Snippet: The effects of conditioned medium obtained from BV2 inflamed microglia on HT22 cell viability. Cell viability is expressed as the percentage of untreated cells (vehicle alone = 100% cell viability). Graph shows the mean + SEM of three independent experiments performed in triplicate. The statistically significant differences between experimental conditions and untreated control cells are shown by asterisks (** p < 0.01).

Article Snippet: The murine BV2 microglia cells were obtained from the Interlab Cell Line Collection, Banca Biologicae Cell Factory, Italy, and cultured using Roswell Park Memorial Institute medium 1640 (RPMI) supplemented with 10% FBS (Sigma, St. Louis, MO, USA), 2 mM L-glutamine (Sigma), 100 mM sodium pyruvate (Sigma), 100 U/mL penicillin, and 100 mg/mL streptomycin (Sigma) in a 5% CO 2 incubator at 37 °C.

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: A Set of Dysregulated Target Genes to Reduce Neuroinflammation at Molecular Level

doi: 10.3390/ijms23137175

Figure Lengend Snippet: Networks representing the interactions among the proteins corresponding to the selected genes before (left) and after the enrichment procedure (right) in HT22 ( a ) and BV2 ( b ) cells.

Article Snippet: The murine BV2 microglia cells were obtained from the Interlab Cell Line Collection, Banca Biologicae Cell Factory, Italy, and cultured using Roswell Park Memorial Institute medium 1640 (RPMI) supplemented with 10% FBS (Sigma, St. Louis, MO, USA), 2 mM L-glutamine (Sigma), 100 mM sodium pyruvate (Sigma), 100 U/mL penicillin, and 100 mg/mL streptomycin (Sigma) in a 5% CO 2 incubator at 37 °C.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: A Set of Dysregulated Target Genes to Reduce Neuroinflammation at Molecular Level

doi: 10.3390/ijms23137175

Figure Lengend Snippet: Venn diagrams showing the number of genes downregulated ( a ) or upregulated ( b ) in the two cellular systems; HT22 cells in blue and BV2 in orange.

Article Snippet: The murine BV2 microglia cells were obtained from the Interlab Cell Line Collection, Banca Biologicae Cell Factory, Italy, and cultured using Roswell Park Memorial Institute medium 1640 (RPMI) supplemented with 10% FBS (Sigma, St. Louis, MO, USA), 2 mM L-glutamine (Sigma), 100 mM sodium pyruvate (Sigma), 100 U/mL penicillin, and 100 mg/mL streptomycin (Sigma) in a 5% CO 2 incubator at 37 °C.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: A Set of Dysregulated Target Genes to Reduce Neuroinflammation at Molecular Level

doi: 10.3390/ijms23137175

Figure Lengend Snippet: The effects of active principles on viability of HT22 and BV2 cells. HT22 ( a , c , e ) and HT22 and BV2 cells ( b , d , f ) were treated with increasing concentrations of α-lipoic acid ( a , b : 0.1, 0.5, 1, 2), N-acetyl-cysteine ( c , d : 0.1, 0.5, 1, 2) and propentofylline ( e , f : 0.1, 1, 10, 100). All the active priciples concentrations are expressed in mM. The graphs show the mean + SEM of three independent experiments performed in triplicate. The statistically significant differences between experimental conditions and untreated control cells are shown by asterisks (* p < 0.01; ** p < 0.01; *** p < 0.001).

Article Snippet: The murine BV2 microglia cells were obtained from the Interlab Cell Line Collection, Banca Biologicae Cell Factory, Italy, and cultured using Roswell Park Memorial Institute medium 1640 (RPMI) supplemented with 10% FBS (Sigma, St. Louis, MO, USA), 2 mM L-glutamine (Sigma), 100 mM sodium pyruvate (Sigma), 100 U/mL penicillin, and 100 mg/mL streptomycin (Sigma) in a 5% CO 2 incubator at 37 °C.

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: A Set of Dysregulated Target Genes to Reduce Neuroinflammation at Molecular Level

doi: 10.3390/ijms23137175

Figure Lengend Snippet: Fold change ± SEM of inflammatory marker genes in BV2 compared with non-inflamed cells. The statistically significant differences between treated and untreated control cells are shown by asterisks (* p < 0.05, ** p < 0.01) and bold type.

Article Snippet: The murine BV2 microglia cells were obtained from the Interlab Cell Line Collection, Banca Biologicae Cell Factory, Italy, and cultured using Roswell Park Memorial Institute medium 1640 (RPMI) supplemented with 10% FBS (Sigma, St. Louis, MO, USA), 2 mM L-glutamine (Sigma), 100 mM sodium pyruvate (Sigma), 100 U/mL penicillin, and 100 mg/mL streptomycin (Sigma) in a 5% CO 2 incubator at 37 °C.

Techniques: Marker, Control

Journal: International Journal of Molecular Sciences

Article Title: A Set of Dysregulated Target Genes to Reduce Neuroinflammation at Molecular Level

doi: 10.3390/ijms23137175

Figure Lengend Snippet: Representation of the distribution of samples in the new coordinate system obtained through the PCA technique for BV2 cell line. Correlation plot shows how much a gene contributes to each dimension of the new coordinate system. alpha_lip: α-lipoic acid; nac: N-acetyl-cystein; ppf: propentofylline; yes/no: presence/absence of active principle treatment; 1/2/3: replicates.

Article Snippet: The murine BV2 microglia cells were obtained from the Interlab Cell Line Collection, Banca Biologicae Cell Factory, Italy, and cultured using Roswell Park Memorial Institute medium 1640 (RPMI) supplemented with 10% FBS (Sigma, St. Louis, MO, USA), 2 mM L-glutamine (Sigma), 100 mM sodium pyruvate (Sigma), 100 U/mL penicillin, and 100 mg/mL streptomycin (Sigma) in a 5% CO 2 incubator at 37 °C.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: A Set of Dysregulated Target Genes to Reduce Neuroinflammation at Molecular Level

doi: 10.3390/ijms23137175

Figure Lengend Snippet: Set of dysregulated target genes. Selection criteria: involvement in neurodegenerative disease progression in mouse model (in black) and human (in red), as reported in references, and high rate of prediction in our in vitro model highlighted by PCA analysis of treatment-affected genes.

Article Snippet: The murine BV2 microglia cells were obtained from the Interlab Cell Line Collection, Banca Biologicae Cell Factory, Italy, and cultured using Roswell Park Memorial Institute medium 1640 (RPMI) supplemented with 10% FBS (Sigma, St. Louis, MO, USA), 2 mM L-glutamine (Sigma), 100 mM sodium pyruvate (Sigma), 100 U/mL penicillin, and 100 mg/mL streptomycin (Sigma) in a 5% CO 2 incubator at 37 °C.

Techniques: Selection, Biomarker Discovery, In Vitro