Journal: Cells

Article Title: Serglycin Cooperates with the Unfolded Protein Response Pathway and Inflammation to Drive Glioblastoma Cell Survival

doi: 10.3390/cells15080660

Figure Lengend Snippet: Loss of SRGN diminishes the expression of UPR markers in GBM cell lines and results in ER stress susceptibility. ( A ) Assessment of the mRNA levels of UPR mediators in LN-18 shSCR and LN-18 shSRGN cells through real-time qPCR. ( B ) Relative mRNA levels of SRGN in LN-18 and U251-MG cell lines. ( C ) SRGN suppression levels in U251-MG siSRGN cells via real-time qPCR and ( D ) phase-contrast microscopy images upon SRGN silencing in U251-MG cells (48 h). ( E ) Relative proliferation rates of U251-MG siControl and U251-MG siSRGN cells after transient SRGN silencing (48 h). ( F ) Evaluation of the gene expression of UPR markers in U251-MG siControl and U251-MG siSRGN cells (48 h). ( G ) IC50 values for LN-18 cell lines upon TM treatment (48 h). ( H ) Wound closure assay in LN-18 cell lines after TM treatment for 24 h and 48 h. Statistically significant differences compared to control are shown by asterisk: * ( p ≤ 0.05).

Article Snippet: The LN-18 GBM cell line was obtained from the American Type Culture Collection (ATCC).

Techniques: Expressing, Microscopy, Gene Expression, Wound Closure Assay, Control

Journal: Cells

Article Title: Serglycin Cooperates with the Unfolded Protein Response Pathway and Inflammation to Drive Glioblastoma Cell Survival

doi: 10.3390/cells15080660

Figure Lengend Snippet: SRGN depletion attenuates the activation profile of UPR in GBM cell lines and renders cells non-responsive to ER stress. ( A ) Western blot analysis of UPR effectors’ protein and phosphorylated levels in TM-treated (48 h) LN-18 cells. ( B ) Immunofluorescence staining for ATF4 (green) and nuclei (blue) depicting the distribution and nuclear accumulation of ATF4 in TM-treated (48 h) LN-18 shSCR cells. ATF4 was not detected in LN-18 shSRGN cells under relative exposure. Scale bar 25 μm. ( C ) Western blot analysis of the basal UPR protein and phosphorylated levels in U251-MG siControl and U251-MG siSRGN cells, following transient SRGN silencing (72 h). All blots are representative of at least three independent experimental repetitions.

Article Snippet: The LN-18 GBM cell line was obtained from the American Type Culture Collection (ATCC).

Techniques: Activation Assay, Western Blot, Immunofluorescence, Staining

Journal: Cells

Article Title: Serglycin Cooperates with the Unfolded Protein Response Pathway and Inflammation to Drive Glioblastoma Cell Survival

doi: 10.3390/cells15080660

Figure Lengend Snippet: SRGN-expressing cells possess a pro-survival UPR mechanism and demand global UPR activation. Assessment of LN-18 shSCR and LN-18 shSRGN cells’ viability upon treatment with ( A ) MKC8866 (IRE1 RNase activity inhibitor), ( B ) Kira6 (IRE1 kinase activity inhibitor), ( C ) GSK2606414 (PERK kinase activity inhibitor) and ( D ) Ceapin-A7 (ATF6 TF activity inhibitor) for 24 h incubation at 0.1, 1 and 10 μΜ final concentrations. Statistically significant differences compared to control are shown by bars and asterisk: * ( p ≤ 0.05).

Article Snippet: The LN-18 GBM cell line was obtained from the American Type Culture Collection (ATCC).

Techniques: Expressing, Activation Assay, Activity Assay, Incubation, Control

Journal: Cells

Article Title: Serglycin Cooperates with the Unfolded Protein Response Pathway and Inflammation to Drive Glioblastoma Cell Survival

doi: 10.3390/cells15080660

Figure Lengend Snippet: SRGN suppression provokes an apoptotic phenotype in GBM cells. ( A ) Western blot analysis of the intact and cleaved forms of caspase-3 and PARP-1, as well as the active and total Akt forms in TM-treated (48 h) LN-18 cells. ( B ) caspase-3 and PARP-1 proteolysis assessed through Western blotting in U251-MG cells after transient SRGN knock-down (72 h). ( C ) Zymography assay for determining the active intracellular CTSB isoforms in acidic pH upon TM treatment (48 h) in LN-18 cell lines. Real-time qPCR for evaluating the relative mRNA levels of ( D ) CTSB , ( E ) BAX , ( F ) Bcl-2 and ( G ) Beclin-1 in TM-treated (48 h) LN-18 cell lines. ( H ) Basal mRNA levels of BAX , Bcl-2 and Beclin-1 in U251-MG cells upon SRGN depletion (48 h). All blots and zymography images are representative of at least three independent experimental repetitions. Statistically significant differences compared to control are shown by bars and asterisk: * ( p ≤ 0.05).

Article Snippet: The LN-18 GBM cell line was obtained from the American Type Culture Collection (ATCC).

Techniques: Western Blot, Knockdown, Zymography, Control

Journal: Cells

Article Title: Serglycin Cooperates with the Unfolded Protein Response Pathway and Inflammation to Drive Glioblastoma Cell Survival

doi: 10.3390/cells15080660

Figure Lengend Snippet: SRGN orchestrates the inflammatory response in GBM cells. ( A ) Relative mRNA levels of TLRs and TNFRs in LN-18 shSCR and LN-18 shSRGN cell lines assessed through real-time PCR. ( B ) Immunofluorescence staining for P-p65 (green) and nuclei (blue) depicting the distribution and nuclear accumulation of phosphorylated p65 subunit in LN-18 cells at basal conditions. P-p65 was not detected in LN-18 shSRGN cells under relative exposure. Scale bar 25 μm. Real-time qPCR for evaluating the relative mRNA levels of ( C ) TLR2 , ( D ) TLR4 , ( E ) TNFRI , ( F ) TNFRII , ( G ) IL-1β , ( H ) IL-8 and ( I ) CXCL-1 in TM-treated (48 h) LN-18 cell lines. ( J ) Basal mRNA levels of TLR4 and IL-1β in U251-MG cells upon transient SRGN silencing (48 h) assessed through real-time qPCR. ( K ) p65/NF-kB phosphorylated and total forms in TM-treated (48 h) LN-18 cell lines evaluated by Western blotting. All blots are representative of at least three independent experimental repetitions. Statistically significant differences compared to control are shown by bars and asterisk: * ( p ≤ 0.05).

Article Snippet: The LN-18 GBM cell line was obtained from the American Type Culture Collection (ATCC).

Techniques: Real-time Polymerase Chain Reaction, Immunofluorescence, Staining, Western Blot, Control

Journal: Molecular Therapy Oncology

Article Title: Potency-enhancing mutations in E3-19K and i-leader increase the cytolytic activity of the PH20/ SPAM1 -armed oncolytic adenovirus Ad5Δ24RGD

doi: 10.1016/j.omton.2026.201137

Figure Lengend Snippet: The 19K SS and iL Q125Ter modifications mutually increase the potency of OAds regardless of the type of fiber modification (A) Sizes of MTT-stained plaques in cancer cell monolayers under an agarose overlay. Two to three independent experiments were conducted. The total number ( n ) of analyzed plaques is indicated for each group. Data are shown as means (SD) and analyzed using Brown-Forsythe and Welch ANOVA tests with Games-Howell post hoc multiple comparisons. (B) IC 50 values derived from viability curves after infection of cells (2.5 × 10 4 per well) in suspension with serial 3-fold dilutions of the indicated OAds. Data were collected on day 6 post-infection for all cell cultures except LN18 and LN229 cells infected with Ad5Δ24RGD-derived OAds (on day 7). Two to three independent experiments with technical triplicates were performed. Data are shown as means (SD). Only ≥3-fold differences from the parental virus Ad5Δ24RGD are indicated. (C) Sizes of fluorescent plaques and MTT-stained plaques in cancer cell monolayers under an agarose overlay. Three to four independent experiments were performed. The total number ( n ) of analyzed plaques is indicated for each group. Data are shown as means (SD) and analyzed using Brown-Forsythe and Welch ANOVA tests and Games-Howell post hoc multiple comparisons. ∗∗∗∗ p ≤ 0.0001; ∗∗∗ p ≤ 0.001; ∗∗ p ≤ 0.01; ∗ p ≤ 0.05; NS, not significant. See also .

Article Snippet: Human embryonic kidney HEK293, lung adenocarcinoma A549 and H441, prostate adenocarcinoma LNCap and PC3, hepatocellular carcinoma HepG2 and Hep3B, glioblastoma LN18, LN229, DBTRG, T98G, and U87 (American Type Culture Collection, ATCC), murine glioma CT-2A (#SCC194, Sigma) and GL261 cells (#ACC802, DSMZ Cell Culture Collection), and human short-term glioma (III–IV grade) primary cell cultures (GliSav, GliShat, GliBah, GliDu, AG-AASH, GBM100622, and GBM160323) were grown in Dulbecco’s modified Eagle medium (DMEM) with L-alanyl-glutamine, 4.5 g/L glucose, and Na pyruvate (#C415, Paneco), supplemented with 10% fetal bovine serum (FBS; HyClone/Cytiva, neoFroxx, or Capricorn Scientific) and penicillin-streptomycin solution at a final concentration of 50 U/mL and 50 μg/mL (#А063, Paneco).

Techniques: Modification, Staining, Derivative Assay, Infection, Suspension, Virus