Journal: Journal of Cellular and Molecular Medicine

Article Title: Exosomal HMGB1 Orchestrates NSCLC Progression and Immunosuppressive Macrophage Polarisation Through the TLR4 / NF ‐ κB / IL ‐6/ STAT3 Signalling Cascade

doi: 10.1111/jcmm.71050

Figure Lengend Snippet: Targeting HMGB1 signalling improves therapeutic outcomes in NSCLC. (A) Correlation analysis between immune infiltration scores and HMGB1 expression in 491 LUAD and 500 LUSC patients from the TCGA database. (B) Correlation between HMGB1 expression and the distribution of various immune cell subsets in LUAD and LUSC patients. (C, D) THP‐1–derived M0 macrophages were treated with PBS, HMGB1 (10 or 100 ng) or exosomes derived from vector or HMGB1 OE cells (cell‐to‐exosome ratio = 1:10). M1 macrophage markers (CD86, CD80, iNOS) and M2 markers (CD206, IL‐10, Arg1) were quantified by PCR. (E) Lewis tumour‐bearing mice were treated with PBS, HMGB1 OE‐derived exosomes (1 × 10 10 exosomes per mouse, twice per week), anti‐PD‐1 antibody (RMP1‐14, 200 μg per mouse, twice per week) or combination therapy ( n = 5 per group). Tumour volumes and apoptosis levels in tumour tissues (day 25) were assessed. (F) PC9 cells were treated with PBS or exosomes from HMGB1 OE cells (cell‐to‐exosome ratio = 1:10), followed by Osimertinib (50 nM, 48 h), and apoptosis was measured. (G) A549 and PC9 cells were similarly treated with PBS or HMGB1 OE‐derived exosomes, followed by Cisplatin (5 μM, 48 h), and apoptosis was analysed. (H) A549 and PC9 cells were similarly treated with paclitaxel (10 μM, 48 h) under the same conditions, and cell apoptosis was determined. (I) A549‐bearing mice were treated with HMGB1 OE‐derived exosomes (1 × 10 10 exosomes per mouse), followed by PBS, paclitaxel (PTX, 10 mg/kg, twice per week), STAT3 inhibitor (5 mg/kg, twice per week) or combination therapy. (J) Schematic diagram illustrating the proposed mechanism: HMGB1 upregulates TLR4, thereby activating the NF‐κB–IL‐6 axis and stimulating JAK2/STAT3 signalling to promote tumour progression. Concurrently, HMGB1 facilitates M2 macrophage polarisation.

Article Snippet: Human NSCLC cell lines (A549, PC9), human embryonic kidney cells (HEK293T) and the human monocytic leukaemia cell line THP‐1 were obtained from the American Type Culture Collection (ATCC, USA).

Techniques: Expressing, Derivative Assay, Plasmid Preparation

Journal: bioRxiv

Article Title: Increased CD33 levels tune activation and function of induced human microglial cells through inhibition of the TREM2 pathway

doi: 10.64898/2026.01.28.701050

Figure Lengend Snippet: (A) Schematic of the TREM2-CD33 pathway. CD33 inhibition acts on SYK activation, upstream of microglial activation pathways AKT, ERK, p38, and JNK. (B) Protein lysates isolated from THP-1 monocytes 48 hours after transduction were used to quantify percent SYK phosphorylation by MSD. (MOI = 31250) (C) 48 hours post-transduction, THP-1 cells (MOI = 31250) were treated with increasing concentrations of TREM2-targeting antibody agonist (0-10 μg/mL). After 5 minutes, protein lysates were isolated and used to quantify percent SYK phosphorylation by MSD. (D) 48-hours post-transduction of increasing concentrations of AAV6-CD33 (MOI = 0-31250), THP-1 cells were treated with a TREM2-targeting antibody agonist (2.5 ug/mL). After 5 minutes, protein lysates were isolated and used to quantify percent SYK phosphorylation by MSD. (E) Percent AKT and GSK3β phosphorylation was quantified from HMC3 cell lysates 48 hours after transduction of AAV6-CD33, quantified by MSD and ELISA, respectively. (F) Percent ERK and p38-MAPK phosphorylation was quantified from HMC3 cell lysates 48 hours after transduction of AAV6-CD33, quantified by ELISA. (G) Percent JNK phosphorylation was quantified from HMC3 cell lysates 48 hours after transduction of AAV6-CD33, quantified by ELISA. (H) (left to right) Percent expression of secreted IL10, IL1β, TNFα, and IL6 in conditioned media from HMC3 cells 24 hours after LPS challenge and 48 hours after transduction of AAV6-CD33 was quantified by MSD. Data represent mean ± SEM using one-tailed unpaired t-test (A, E-G), and two-way ANOVA using uncorrected Fisher’s LSD with a single pooled variance (H). Data points (n-numbers) are plotted on each bar graph and scatter plot each representing an independent experimental replicate. Line plots are representative of three and two independent experimental replicates (C, D, respectively).

Article Snippet: THP-1 Cells: The human monocytic cell line THP-1 (TIB-202) was obtained from ATCC.

Techniques: Inhibition, Activation Assay, Isolation, Transduction, Phospho-proteomics, Enzyme-linked Immunosorbent Assay, Expressing, One-tailed Test