u118 mg cell line  (ATCC)

Journal: iScience

Article Title: Phosphatidylserine receptors TIM-1 and AXL mediate tick-borne Powassan virus entry

doi: 10.1016/j.isci.2025.113930

Figure Lengend Snippet: Genomic sequence analysis and infectious U118 neuronal cell culture system (A) Phylogenetic analysis of five Powassan viral genome sequences from known lineage I and II strains. GenBank accession numbers are provided along with strain information including location and year of isolation. Phylogenetic tree was constructed using the neighbor-joining method with pairwise distance estimation following ClustalW alignment program. (B) Matrix panel shows the percent identity in the upper right and nucleotide divergence in the lower left of aligned Powassan viral genomic sequences (ClustalW program, DNASTAR Lasergene MegAlign 15). (C) Graph shows the RT-qPCR analysis of POWV (LB), POWV ( M11665 ), and DTV (SPO) genome replication in U118 cells 24 and 48 hpi. The y axis shows genome copies per 50 ng of total RNA in log scale. Error bars represent the SD of three biological replicates. Student’s t test. ∗ p < 0.01, ∗∗ p < 0.001, ∗∗∗ p < 0.0001, ∗∗∗∗ p < 0.00001. (D) Representative immunofluorescence images of mock and LB-, DTV- ( M11665 ), and DTV- (SPO) infected U118 neuronal cells detected by viral NS1 at 24 and 48 hpi (scale bars, 25 μm). Red, NS1 viral protein; blue, DAPI. (E) Western blot analysis of infected cell lysates probed with the anti-M protein antibody targeting LB, DTV ( M11665 ), and DTV (SPO) at 24 and 48 hpi. Cells were infected with each virus at an MOI of 1. Two independent experiments were conducted. Representative data are presented.

Article Snippet: U118 brain cells (HTB-15, ATCC) were cultured as monolayers in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 1X non-essential amino acids (NEAAs), penicillin (100 units/ml), and streptomycin (100 units/ml).

Techniques: Sequencing, Cell Culture, Isolation, Construct, Genomic Sequencing, Quantitative RT-PCR, Immunofluorescence, Infection, Western Blot, Virus

Journal: iScience

Article Title: Phosphatidylserine receptors TIM-1 and AXL mediate tick-borne Powassan virus entry

doi: 10.1016/j.isci.2025.113930

Figure Lengend Snippet: Expression analysis of TIM-1, TIM-4, AXL, TYRO3, and MERTK on Vero E6 and U118 cells and viral entry-neutralization assay (A) FSC/SSC plot with gating and histograms from flow analysis are shown. Black line shows cell staining by control goat antibody, while the red line represents staining by antibodies against specific PtdSer receptors. The value indicated in the histogram corresponds to the expression level of each receptor protein calculated by subtracting the mean fluorescent intensity of the control antibody-stained cells from that of the cells stained with an antibody specific to a PtdSer receptor. ND, not detected. Note: TIM-1 and AXL expression on the Vero E6 cell surface was found. TIM-4, TYRO3, and MERTK were not expressed. On the U118 cell surface, only AXL expression was found. (B) Schematic representation of neutralization assay using 100 plaque-forming units of virus incubated with soluble forms of TIM-1 receptor Fc-sTIM1 (sTIM1dMLDR801) or control protein Fc-NS (NCFcCQ R801). (C) Graphs show fold inhibition of indicated viral replication following neutralization with sTIM1dMLDR801. Error bars represent the SD using three biological replicates. Student’s t test. ∗ p < 0.01, ∗∗ p < 0.001, ∗∗∗ p < 0.0001. Representative data from two independent experiments were provided.

Article Snippet: U118 brain cells (HTB-15, ATCC) were cultured as monolayers in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 1X non-essential amino acids (NEAAs), penicillin (100 units/ml), and streptomycin (100 units/ml).

Techniques: Expressing, Neutralization, Staining, Control, Virus, Incubation, Inhibition

Journal: Journal of Neuroimmune Pharmacology

Article Title: Inhibition of TRAF3IP2 Modulates NAMPT and NAD Metabolism in Glioblastoma

doi: 10.1007/s11481-025-10252-z

Figure Lengend Snippet: Inhibition of TRAF3IP2 reduces NAMPT expression in glioblastoma. Analysis of glioblastoma samples from TCGA (red bars) compared to non-malignant brain samples (blue bars) from GTEx shows an increase in the expression of TRAF3IP2 and NAMPT mRNA, accessed through GEPIA 2 ( http://gepia2.cancer-pku.cn ) ( A ). Kaplan-Meier curve analysis of overall survival from TCGA sample analysis using the Xena platform ( https://xena.ucsc.edu/ ) reveals an inverse association between TRAF3IP2 gene expression and length of survival in glioblastoma patients ( B ). Levels of mRNA expression, measured through qRT PCR, show reduced TRAF3IP2, NAMPT and SIRT1 expression in U87 TRAF3IP2KD and U118 TRAF3IP2KD cells, compared to U87 SCR and U118 TRAF3IP2KD , respectively ( p < 0.05) ( C ). Silencing TRAF3IP2 reduces relative NAD levels in glioblastoma cell lines, compared to scrambled controls (SCR). The level of NAD was measured in U87 TRAF3IP2KD and U87 SCR in presence or absence of NAMPT inhibitor “FK866” (DMSO was used as solvent for FK866) ( D ). Silencing TRAF3IP2 decreases ATP production mechanism, measured through reduced oxygen consumption rate (OCR) in Agilent Seahorse XF Cell Mito Stress Test kit, in glioblastoma cell line KNS (Compared to SCR controls) ( E ). Triplicate experiments, * P < 0.05, ** P < 0.01, *** P < 0.005, **** P < 0.001

Article Snippet: Glioblastoma U-87 MG (U87) and U-118 MG (U118) cell lines were purchased from ATCC (Rockville, MD) and Pediatric glioblastoma KNS42 cells (KNS) were obtained from the JCRB (Japan Cancer Research Resources) cell bank.

Techniques: Inhibition, Expressing, Gene Expression, Quantitative RT-PCR, Solvent

Journal: Journal of Neuroimmune Pharmacology

Article Title: Inhibition of TRAF3IP2 Modulates NAMPT and NAD Metabolism in Glioblastoma

doi: 10.1007/s11481-025-10252-z

Figure Lengend Snippet: Targeting TRAF3IP2 alters protein levels of NAD salvage pathway markers in glioblastoma. Protein levels of TRAF3IP2 and NAMPT in U87, U118, and KNS cell lines ( A ). Protein levels of SIRT1, total p53, Acetyl-p53[K382] (a-p53), and Phosphorylated-p53[S392] (p-p53) in U87, U118, and KNS cell lines ( B ). Imagej analysis of band intensity in western blots, relative to β-actin (TRAF3IP2KD vs. SCR) ( C ). Immunohistochemical analysis of the tumor tissues demonstrating that U87 TRAF3IP2KD -derived tumors had reduced TRAF3IP2 and NAMPT expression, compared to SCR control tumor tissues (Size bar = 50 µM) ( D ). Relative fluorometric intensity of ROS in U87 TRAF3IP2KD , compared to U87 SCR showed that silencing TRA3IP2 increases ROS levels ( E ). Protein expression of LKB1, p-LKB1, AMPK, p-AMPK in transduced U87, U118, and KNS glioblastoma cell lines (F). Triplicate experiments, (* P < 0.05)

Article Snippet: Glioblastoma U-87 MG (U87) and U-118 MG (U118) cell lines were purchased from ATCC (Rockville, MD) and Pediatric glioblastoma KNS42 cells (KNS) were obtained from the JCRB (Japan Cancer Research Resources) cell bank.

Techniques: Western Blot, Immunohistochemical staining, Derivative Assay, Expressing, Control

Journal: Journal of Neuroimmune Pharmacology

Article Title: Inhibition of TRAF3IP2 Modulates NAMPT and NAD Metabolism in Glioblastoma

doi: 10.1007/s11481-025-10252-z

Figure Lengend Snippet: TRAF3IP2 inhibition is associated with increased ROS presence and apoptosis. Gene expression analysis showed elevated levels of survivin in TRAF3IP2KD cells, compared to SCR controls (Triplicate experiments/cell lines, p < 0.05) ( A ). Western blot analysis of showed levels of survivin, total caspase-3 and cleaved caspase-3 protein in U87 and U118 cells ( B ). Scatterplot of glioblastoma cell groups and treatment types (U87 SCR+DMSO (i); U87 SCR+FK866 (ii); U87 TRAF3IP2KD+DMSO (iii); U87 TRAF3IP2KD+FK866 (iv)), with bar graph representation of Annexin V and PI staining of U87 (v) ( C ). * P < 0.05

Article Snippet: Glioblastoma U-87 MG (U87) and U-118 MG (U118) cell lines were purchased from ATCC (Rockville, MD) and Pediatric glioblastoma KNS42 cells (KNS) were obtained from the JCRB (Japan Cancer Research Resources) cell bank.

Techniques: Inhibition, Gene Expression, Western Blot, Staining

Journal: Journal of Neuroimmune Pharmacology

Article Title: Inhibition of TRAF3IP2 Modulates NAMPT and NAD Metabolism in Glioblastoma

doi: 10.1007/s11481-025-10252-z

Figure Lengend Snippet: Targeting TRAF3IP2 reduces mTOR associated cell growth markers and glycolysis and glioblastoma cells. Fold change difference in mTOR and mTORC associated markers Raptor and Rictor in TRAF3IP2KD cells, compared to SCR controls, in U87, U118 and KNS cell lines (Triplicate experiments/cell lines, p < 0.05) ( A ). Western blot of Raptor, p-Raptor, Rictor, and p-Rictor protein expression in U87 SCR and U87 TRAF3IP2KD cells ( B ). Agilent Seahorse XF Glycolytic stress test measuring glycolysis, glycolytic capacity, glycolytic reserve, and non-glycolytic acidification through assessment of real-time ECAR levels, normalized to total µg protein in cell culture wells, of glioblastoma cell lines U87 ( C ) and KNS ( D )

Article Snippet: Glioblastoma U-87 MG (U87) and U-118 MG (U118) cell lines were purchased from ATCC (Rockville, MD) and Pediatric glioblastoma KNS42 cells (KNS) were obtained from the JCRB (Japan Cancer Research Resources) cell bank.

Techniques: Western Blot, Expressing, Cell Culture