Journal: bioRxiv

Article Title: Multi‑omic and spatial analysis reveals tumour‑derived paracrine signals drive suppressive macrophage polarisation via activation of the cAMP-CREB axis in glioblastoma

doi: 10.64898/2026.02.19.706754

Figure Lengend Snippet: (A) Western blot of THP-1 differentiated macrophages stimulated for 30 min, with conditioned media (CM) from a panel of GBM cell lines and probed using antibodies for, total CREB, phospho-CREB (pCREB) and b-actin. Data shown is representative of three independent experiments. (B) CREB transactivation activity in THP-1 macrophages transfected with a CREB luciferase reporter plasmid after stimulation with MU41 GBM cell conditioned medium (CM). CREB activity is presented as Relative Light Units (RLU) per mg of protein lysate. Statistical significance was determined using one-way ANOVA, followed by Tukey’s test for multiple comparisons (a=0.05). *(p<0.05). Error bars are median ± interquartile range. (C) Expression of CD2026 in THP-1 differentiated macrophages incubated with or without GBM cell (MU41) conditioned medium (CM) in the presence or absence of the CREB inhibitor (CREBi), 666-15 (1mM). (D) Multiplex IHC image showing phospho-AKT (pAKT), phospho-ERK1/2 (pERK) and pCREB expression in BMDM-TAMs in GBM tissue. Scale bar is 50mm. (E) Percentage of CD68 + pCREB + MDM co-expressing pAKT and pERK1/2 in GBM tissue (n=3). Error bars are S.E.M. (F) Cell lysates from THP-1 differentiated macrophages were probed for expression of pCREB, CREB, b-actin, pAKT, AKT, pERK and ERK following a 30-minute stimulation with GBM-CM, in combination with either a PI3K inhibitor (PI3Ki) BKM-120 (2 mM), a MAPK inhibitor (MAPKi) U0126 (10 mM), a cAMP inhibitor (cAMPi) H89 (10 mM) and a CREB inhibitor (CREBi) 666-15 (1 mM). MU41-CM with DMSO, and DMSO in fresh media (RPMI) ‘medium/med’, are negative controls, basal medium without GBM-CM.

Article Snippet: Human GBM cell lines (U-87MG, LN-229, LN-18, U-118MG, T98G, A172) obtained from the ATCC, and patient-derived GBM cells (MU41) were maintained in Dulbecco’s Modified Eagle’s Medium (DMEM) (Thermo Fisher Scientific) with 10% fetal calf serum (FCS) (Bovogen, Australia, #SFBS-F) and penicillin-streptomycin-amphotericin B (Gibco Anti-Anti, Thermo Fisher Scientific).

Techniques: Western Blot, Activity Assay, Transfection, Luciferase, Plasmid Preparation, Expressing, Incubation, Multiplex Assay

Journal: Carcinogenesis

Article Title: The novel agent phospho-glycerol-ibuprofen-amide (MDC-330) inhibits glioblastoma growth in mice: an effect mediated by cyclin D1

doi: 10.1093/carcin/bgw017

Figure Lengend Snippet: Cell kinetic effect of PGIA in GBM cells. (A) LN-229, U118-MG, U87-MG and LN-18 cells were treated with PGIA for 24h. Cell proliferation assay based on BrdU incorporation into DNA during the S-phase of the cell cycle. The percentage of BrdU-positive cells is shown in the right. (B) Cell death by apoptosis was determined by flow cytometry using the dual staining (annexin V and propidium iodide) in U87-MG cells treated with increasing concentrations of PGIA for 24h. Results are expressed as fold-increase compared with the percentage of annexin V(+) cells in the control group. (C) Differential cytotoxic effect of PGIA in GBM cells compared with NHA. Apoptosis was determined by flow cytometry in NHA, LN-18 and LN-229 cells incubated without or with 200 µM of PGIA for 24h. Results are expressed as fold-increase compared with the percentages of apoptotic cells in the control cells. (D) Cell cycle analysis of cells treated with and without PGIA for 24h.

Article Snippet: A panel of four human GBM (U87-MG, LN-18, LN-229 and U118-MG) cell lines were obtained from the American Type Culture Collection (Manassas, VA).

Techniques: Proliferation Assay, BrdU Incorporation Assay, Flow Cytometry, Staining, Control, Incubation, Cell Cycle Assay

Journal: Carcinogenesis

Article Title: The novel agent phospho-glycerol-ibuprofen-amide (MDC-330) inhibits glioblastoma growth in mice: an effect mediated by cyclin D1

doi: 10.1093/carcin/bgw017

Figure Lengend Snippet: PGIA reduces GBM xenograft growth. (A) PGIA inhibits the growth of human U118-MG xenografts. U118-MG tumor volume growth over time for vehicle control (■), ibuprofen (20mg/kg/day 5×/week (●) and PGIA 20mg/kg/day 5×/week (▲) treated mice. *Significantly different compared with control group (P < 0.01), #significantly different compared with ibuprofen group (P = 0.023), one-way ANOVA test. (B) PGIA inhibits the growth of human U87-MG xenografts. U87-MG tumor volume growth over time for vehicle control (♦), and PGIA 20mg/kg/day (■) treated mice. *Significantly different compared with control group (P < 0.01), one-way ANOVA test. (C) Ki-67 and p21 immunostaining were performed on tumor sections, and photographs were taken at ×20 magnification. Representative images are shown (left). Results were expressed as percentage of Ki-67+ or p21+ cells ± SEM per 20× field (right). *Significance compared with control group; P < 0.05. (D) Levels of p21 were measured by immunoblot in total fractions isolated from LN-229 and LN-18 cells treated with PGIA. β-Actin = loading control.

Article Snippet: A panel of four human GBM (U87-MG, LN-18, LN-229 and U118-MG) cell lines were obtained from the American Type Culture Collection (Manassas, VA).

Techniques: Control, Immunostaining, Western Blot, Isolation