Journal: ACS Nano

Article Title: Carrier-Free Nanoassembly Suppresses Phase Separation via Ribosome-Inspired Crowding Control for Enhanced Chemo-Immunotherapy

doi: 10.1021/acsnano.5c17081

Figure Lengend Snippet: Mechanism of tumor growth inhibition by OSI-027. (A) Apoptosis of PDAC cells by the OSI-027-treated THP-1-cell system and (B) quantification of the positive rate of apoptosis as a percentage. (C) Phagocytosis of PDAC cells by the OSI-027-treated THP-1-cell system and (D) quantification of the positive rate of phagocytosis as a percentage. (E) A schematic of the HUVEC culture supernatant of the OSI-027-treated coculture system. (F) Microvascular density of the PDAC cell and THP-1-cell coculture system after OSI-027 treatment and (G) quantification of the area ratio of the microvascular density percentages. Data are shown as mean ± SD ( n = 3). (* p < 0.05, ** p < 0.01 and *** p < 0.001).

Article Snippet: The mTOR inhibitor (OSI-027) and Annexin V/propidium iodide (PI) apoptosis assay kits were purchased from MedChemExpress (NJ, USA).

Techniques: Inhibition

Journal: Advanced Science

Article Title: Targeting the ATX‐LPA Axis Overcomes TKI Resistance and Immunosuppression in Renal Cell Carcinoma via Dual Inhibition of AKT/mTOR and TBK1/IRF3 Pathways

doi: 10.1002/advs.76352

Figure Lengend Snippet: ENPP2 drives cabozantinib resistance through its enzymatic activity and LPA biosynthesis. (A) CCK‐8 viability assays of ACHN‐R and 786‐R cells treated with 10 µM GLPG1690, 10 µM HA130, or DMSO control, across a concentration gradient of cabozantinib for 48 h. (B, C) Representative images (B) and total area quantification (C) of colony formation assays for resistant cells treated with 5 µM cabozantinib in the presence of 10 µM GLPG1690, 10 µM HA130, or DMSO. (D, E) Representative Annexin V/DAPI flow cytometry plots (D) and apoptosis rate quantification (E) of 786‐R and ACHN‐R cells transduced with control shRNA (NC) or shENPP2, and treated with 5 µM cabozantinib plus 10 µM GLPG1690, 10 µM HA130, or DMSO for 48 h. (F) ELISA quantification of secreted LPA levels in cell culture supernatants from shNC and shENPP2 RCC cells cultured in low‐serum medium (0.5% FBS) for 6 h. (G) Colony formation assays of parental 786‐O and ACHN cells treated with DMSO control, 5 µM cabozantinib alone, or 5 µM cabozantinib combined with 10 µM exogenous LPA. (H, I) Flow cytometric apoptosis analysis (H) and quantification (I) of parental cells treated with 5 µM cabozantinib and co‐treated with DMSO or exogenous LPA (C1 = 10 µM; C2 = 20 µM) for 48 h. (J, K) CCK‐8 viability assays of 786‐R and ACHN‐R cells expressing shNC or shENPP2, treated with cabozantinib at the indicated concentrations with or without 20 µM LPA. (L, M) CCK‐8 assays demonstrating that overexpression of wild‐type ENPP2 (oeENPP2) enhanced TKI resistance in sensitive cell lines, while no such effect was observed in cells that overexpressed the enzymatically inactive T210A mutant. (N) Correlations between ENPP2 (top) or PLPP1 (bottom) expression and selected functional states of tumor cells obtained from the CancerSea database. All quantitative in vitro data are presented as mean ± SD from n = 3 independent experiments. Statistical significance was determined using an unpaired two‐tailed Student's t‐test (F), one‐way ANOVA (C, E, G, I), or two‐way ANOVA (A, J, K, L, M), followed by Tukey's post hoc test. ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: For apoptosis analysis, cells were collected and evaluated by the Annexin V‐APC/propidium iodide (PI) apoptosis kit (AP‐107, Multi Science) according to the manufacturer's instructions.

Techniques: Activity Assay, CCK-8 Assay, Control, Concentration Assay, Flow Cytometry, Transduction, shRNA, Enzyme-linked Immunosorbent Assay, Cell Culture, Expressing, Over Expression, Mutagenesis, Functional Assay, In Vitro, Two Tailed Test

Journal: Advanced Science

Article Title: Targeting the ATX‐LPA Axis Overcomes TKI Resistance and Immunosuppression in Renal Cell Carcinoma via Dual Inhibition of AKT/mTOR and TBK1/IRF3 Pathways

doi: 10.1002/advs.76352

Figure Lengend Snippet: LPAR1 mediates ENPP2‐driven TKI resistance by activating a TBK1‐AKT/mTOR cascade. (A) Baseline mRNA expression profiles of LPAR1–6 in 786‐O and ACHN cell lines, determined by qRT‐PCR. (B) Immunoblot analysis of downstream cascade activation (TBK1, IRF3, AKT, and mTOR) in ENPP2‐overexpressing RCC cells treated with DMSO or the specific LPAR1 antagonist AM966. (C) Immunoblot analysis confirming that genetic depletion of LPAR2 (siLPAR2 #1 and #2) fails to abrogate ENPP2‐induced downstream signaling. (D–G) In vitro viability evaluated by CCK‐8 assays across cabozantinib concentration gradients (D, E) and flow cytometric apoptosis analysis (F, G) of RCC cells following LPAR1 silencing (D, F, G) or LPAR2 silencing (E), demonstrating that cabozantinib sensitization is LPAR1‐dependent. (H, I) Representative in vivo bioluminescence images (H) and quantification of tumor growth kinetics (I) for 786‐O xenografts harboring stable LPAR1 knockdown under cabozantinib treatment (n = 5 mice per group). (J, K) Immunoblot analysis of AKT and mTOR phosphorylation following TBK1 knockdown (J) or ectopic TBK1 overexpression (K). (L) Immunoblot analysis of TBK1 and AKT phosphorylation in RCC cells overexpressing a constitutively active AKT mutant (ca‐AKT). (M) Immunoblot analysis of TBK1 and AKT phosphorylation in ENPP2‐overexpressing cells following TBK1 or AKT silencing. (N) EdU incorporation assays in RCC cells following TBK1 depletion. (O, P) CCK‐8 cell viability assays of TBK1‐depleted RCC cells across a cabozantinib concentration gradient. (Q) Colony formation assays of TBK1‐depleted RCC cells under cabozantinib treatment. (R, S) Representative flow cytometry plots (R) and apoptosis rate quantification (S) demonstrating that ectopic expression of TBK1 (oeTBK1) effectively rescues cell survival in ENPP2‐depleted cells under cabozantinib treatment. Data are presented as mean ± SD from n = 3 independent in vitro experiments or n = 5 mice per group. Statistical significance was determined using a one‐way ANOVA (G, N, S), or two‐way ANOVA (D, E, I, O, P), followed by Tukey's post hoc test. ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: For apoptosis analysis, cells were collected and evaluated by the Annexin V‐APC/propidium iodide (PI) apoptosis kit (AP‐107, Multi Science) according to the manufacturer's instructions.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Activation Assay, In Vitro, CCK-8 Assay, Concentration Assay, In Vivo, Knockdown, Phospho-proteomics, Over Expression, Mutagenesis, Flow Cytometry

Journal: Nature Communications

Article Title: Fusobacterium periodonticum promotes colorectal tumorigenesis via decanoic acid-driven neutrophil chemotaxis

doi: 10.1038/s41467-026-74591-y

Figure Lengend Snippet: A Flow cytometry analysis of neutrophil apoptosis after coculture with sodium decanoate (DeaNa), sodium palmitate (PaNa), and PBS. Quantitative graphs showing the proportion of late apoptotic/necrotic (Annexin V⁺/PI⁺) neutrophils ( B ) and early apoptotic (Annexin V⁺/PI⁻) neutrophils ( C ) under different treatments ( n = 3). D Schematic representation of neutrophil migration in vitro for 3 h at 37 °C. Created in BioRender. Yang, Q. (2026) https://BioRender.com/fm3t1j5 . Representative images of migrated neutrophils on the lower membrane stained with DAPI and photographed using the multifunctional enzyme reader. E Numbers of migrating neutrophils ( n = 3). F Volcano plot of differential gene expression in sodium decanoate-treated and PBS-treated neutrophils. G Heatmap showing the DEGs identified via RNA-seq in the presence of decanoic acid or PBS. H Quantitative reverse transcription PCR (qRT‒PCR) validation of gene expression changes in neutrophils treated with decanoic acid, palmitic acid and PBS ( n = 4). I Cell supernatant CXCL8 expression in neutrophils treated with decanoic acid, palmitic acid and PBS, as determined by ELISA ( n = 4). J Simplified schematic illustrating the coculture assay of neutrophils with Caco-2 cells, detailing the experimental setup. Created in BioRender. Yang, Q. (2026) https://BioRender.com/74ys0y4 . K qRT‒PCR validation of gene expression changes in Caco-2 cells after coculture with different fatty-acid-treated neutrophils ( n = 3). Data are shown as mean ± SEM, with representative results from at least three independent experiments. P values were determined by one-way ANOVA with Holm–Šidák test ( B , C ), one-way ANOVA with Dunnett’s test ( E , K ), one-way ANOVA with Tukey test ( H – I ). DeaNa, sodium decanoate; PaNa, sodium palmitate. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001 and ns P > 0.05. Exact P values: B DeaNa vs PBS, P = 0.0465, PaNa vs PBS, P = 0.0002; C DeaNa vs PBS, P < 0.0001, PaNa vs PBS, P < 0.0001; E DeaNa vs PBS, P = 0.0342, PaNa vs PBS, P = 0.4726; H Cxcl2 , DeaNa vs PBS, P = 0.0015, PaNa vs PBS, P = 0.0001; Cxcl8 , DeaNa vs PBS, P = 0.0120, PaNa vs PBS, P = 0.0725; Cpt1a , DeaNa vs PBS, P = 0.0441, PaNa vs PBS, P < 0.0001; Osm , DeaNa vs PBS, P = 0.0078, PaNa vs PBS, P = 0.0019; Vegfa , DeaNa vs PBS, P = 0.0319, PaNa vs PBS, P = 0.0009. I DeaNa vs PBS, P = 0.0016, PaNa vs PBS, P = 0.0101. K CXCL8, DeaNa vs PBS, P < 0.0001, PaNa vs PBS, P = 0.9138; OSM, DeaNa vs PBS, P = 0.0865, PaNa vs PBS, P = 0.0031; VEGFA, DeaNa vs PBS, P = 0.0053, PaNa vs PBS, P < 0.0001; AREG, DeaNa vs PBS, P = 0.0451, PaNa vs PBS, P = 0.0391; PTGS2, DeaNa vs PBS, P = 0.0015, PaNa vs PBS, P = 0.0224.

Article Snippet: After treatment, cells were collected and stained using an Annexin V-FITC/Propidium Iodide (PI)-PE Apoptosis Detection Kit (MedChemExpress, USA).

Techniques: Flow Cytometry, Migration, In Vitro, Membrane, Staining, Gene Expression, RNA Sequencing, Reverse Transcription, Biomarker Discovery, Expressing, Enzyme-linked Immunosorbent Assay, Co-culture Assay