Journal: Bioactive Materials

Article Title: Injection site dictates the immune response to a biodegradable polymer and corresponding collagen regeneration

doi: 10.1016/j.bioactmat.2026.04.004

Figure Lengend Snippet: LA released from T-gel degradation induces macrophage M2 polarization and promotes fibroblast activation and collagen production. (A) Flow cytometry analysis of the association between T-gel degradation products and macrophage M2 polarization ( n = 3, data represent mean ± s.d.). ∗∗∗∗P < 0.0001. (B) RNA-seq analysis of M2 macrophage marker expression in tissues one week after ID and SC injection of T-gel. (C–D) Immunofluorescence analysis of CD206 (C) and FAPα (D) expression in tissues ( n = 3, data represent mean ± s.d.). ∗∗P < 0.01. (E) Schematic diagram of the co-culture system showing RAW264.7 macrophages pretreated with T-gel extract medium that were subsequently co-cultured with L929 fibroblasts. (F–G) Collagen expression in L929 cells from the co-culture system assessed by Western blot (F) and ELISA (G). “C” represents RAW264.7 pretreated with blank medium; “T” represents RAW264.7 pretreated with T-gel extract medium. n = 3, data represent mean ± s.d. ∗P < 0.05 and ∗∗P < 0.01. (H) ELISA analysis of TGFβ expression in RAW264.7 cells treated with T-gel extract medium ( n = 3, data represent mean ± s.d.). ∗∗P < 0.01. (I) Schematic representation of LA, released during T-gel degradation, programming macrophages toward an M2 phenotype and subsequently promoting fibroblast activation and collagen secretion.

Article Snippet: Following permeabilization, cells were stained with Phycoerythrin (PE)-conjugated anti-mouse CD206 antibody (Elabscience, E-AB-F1135D) for 30 min at 4 °C in the dark.

Techniques: Activation Assay, Flow Cytometry, RNA Sequencing, Marker, Expressing, Injection, Immunofluorescence, Co-Culture Assay, Cell Culture, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Bioactive Materials

Article Title: Spatiotemporally engineered microneedle for microenvironment remodeling propels mucosal regeneration after tracheal mucosal injury

doi: 10.1016/j.bioactmat.2026.01.026

Figure Lengend Snippet: Evaluation of mucosal status after tracheal mucosal injury in rabbits . (a) Gross morphology and Masson staining of mucosal defects (n = 3). (b – f) Tracheal samples were collected on Day 10, and RNA sequencing was performed to assess biological differences between Native and Model groups (n = 3). (b) Principal components analysis of samples. (c) Volcano plot of DEGs. (d) KEGG enrichment analysis of DEGs. (e) Chord plot of enriched KEGG terms. (f) Heatmaps of DEGs associated with inflammation and oxidative stress. (g) IF staining of FISH (marker of bacteria, pink), iNOS (marker of M1 macrophages, orange), CD206 (marker of M2 macrophages, green), and immunohistochemical staining of CD31 (marker of endothelial cells) in various samples, blood vessels are denoted by triangles.

Article Snippet: IF staining was performed for iNOS (Abcam, ab283655), CD206 (Proteintech, 18704-1-AP), and CD31 (Abcam, ab28364) to assess inflammatory marker expression, and vascularization, respectively.

Techniques: Staining, RNA Sequencing, Marker, Bacteria, Immunohistochemical staining

Journal: Bioactive Materials

Article Title: Spatiotemporally engineered microneedle for microenvironment remodeling propels mucosal regeneration after tracheal mucosal injury

doi: 10.1016/j.bioactmat.2026.01.026

Figure Lengend Snippet: Gel-AgNA/MgGA MN remodel the microenvironment after TMI by regulating infection, inflammation, oxidative stress, and vascular disruption. (a) FISH (marker of bacteria, red) staining of harvested samples after treated with Gel, Gel-AgNA, Gel-MgGA, and Gel-AgNA/MgGA MN. (b, c) Quantitative analysis of bacterial coverage and fluorescence intensity of FISH (n = 9). (d) Heatmaps of DEGs related to inflammation and oxidative stress. (e) RT-PCR analysis showing the relative expression levels of IL-1, TNF-α, HIF-1α, and IL-6 (n = 3). (f) IF staining of iNOS (marker of M1 macrophages, red) and CD206 (marker of M2 macrophages, green). (g) Quantitative analysis of iNOS and CD206 fluorescence intensity. (h) RT-PCR analysis showing the relative expression levels of Arg1 and CD206. (i) IF staining of CD31 (marker of blood vessels, red). (j, k) RT-PCR analysis showing the relative expression levels of VEGF and HIF-1α (n = 3).

Article Snippet: IF staining was performed for iNOS (Abcam, ab283655), CD206 (Proteintech, 18704-1-AP), and CD31 (Abcam, ab28364) to assess inflammatory marker expression, and vascularization, respectively.

Techniques: Infection, Disruption, Marker, Bacteria, Staining, Fluorescence, Reverse Transcription Polymerase Chain Reaction, Expressing

Journal: Regenerative Therapy

Article Title: Lung cancer cell-derived exosomal EHF drives M2 macrophage polarization via transcriptional activation of RNF41 to promote tumor progression

doi: 10.1016/j.reth.2026.101104

Figure Lengend Snippet: EHF promotes lung cancer cell malignancy and M2 macrophage polarization. A549 and H520 cells were transfected with control (Ctrl) or EHF-knockdown (KD-EHF) plasmids. (A-B) Western blot and quantification of EHF protein levels in A549 and H520 cells. (C-D) EdU staining and quantification of EdU + cells in A549 and H520 cells, assessing proliferation. (E-H) Glucose uptake and lactate production assays were conducted using commercial kits to evaluate glycolytic output. (I-J) Flow cytometric analysis of apoptosis. (K-L) Transwell assay was employed to examine cell migration. (M − P) THP-1 cells were differentiated into THP-1-M0 macrophages with 100 ng/mL PMA for 24 h. Culture supernatants from Ctrl- or KD-EHF–transfected A549 and H520 cells were collected and co-incubated with THP-1-M0 macrophages. (M − N) Flow cytometric analysis was performed to assess the proportion of CD206 + M2-polarized macrophages. (O–P) ELISA was conducted to quantify the secretion of M2-associated cytokines (TGF-β1, IL-10, and VEGFA) in the supernatants of THP-1-M0 macrophages. Data are presented as mean ± SD. P < 0.05 denotes statistical significance.

Article Snippet: Membranes were blocked with 5% skim milk in TBST, incubated with primary antibodies against EHF (1:1000, Catalog #: 27195-1-AP, Proteintech, Wuhan, China), TSG101 (1:1000, Catalog #: 14497-1-AP, Proteintech), Calnexin (1:1000, Catalog #: 10427-2-AP, Proteintech), CD9 (1:2000, Catalog #: 60232-1-Ig, Proteintech), RNF41 (1:500, Catalog #: 17233-1-AP, Proteintech), CD206 (1:1000, Catalog #: 18704-1-AP, Proteintech), TGF-β1 (1:1000, Catalog #: 26155-1-AP, Proteintech), VEGFA (1:1000, Catalog #: ab46154, Abcam, Cambridge, UK), and β-actin (1:1000, Catalog #: 66009-1-Ig, Proteintech) overnight at 4 °C, followed by HRP-conjugated secondary antibodies (1:5000, Catalog #: ab6721/ab205719, Abcam) for 1 h at room temperature.

Techniques: Transfection, Control, Knockdown, Western Blot, Staining, Transwell Assay, Migration, Incubation, Enzyme-linked Immunosorbent Assay

Journal: Regenerative Therapy

Article Title: Lung cancer cell-derived exosomal EHF drives M2 macrophage polarization via transcriptional activation of RNF41 to promote tumor progression

doi: 10.1016/j.reth.2026.101104

Figure Lengend Snippet: Exosomal EHF reprograms THP-1-M0 macrophages to promote lung cancer cell malignancy and angiogenesis via paracrine signaling. (A) Schematic of the paracrine co-culture system: Exosomes from control (Ctrl) or EHF-knockdown (KD-EHF) A549/H520 cells were co-incubated with THP-1-M0 macrophages. The medium from these macrophages was used to treat A549/H520 cells or HUVECs. (B-E) THP-1-M0 macrophages were treated with three conditions: blank control (no exosome), exosomes from control-transfected A549/H520 cells (Exo Ctrl ), or exosomes from EHF-knockdown A549/H520 cells (Exo KD−EHF ). (B–C) Flow cytometry analysis of CD206 + M2 macrophage proportions in THP-1-M0 cells. (D-E) ELISA measurement of M2-associated cytokines (TGF-β1, IL-10, and VEGFA). (F-M) The medium from exosome treated macrophages was then used to incubate A549/H520 cells or HUVECs. (F-G) EdU staining and quantification of EdU + proliferating A549 and H520 cells. (H–I) Flow cytometric analysis of apoptosis in A549 and H520 cells. (J-K) Transwell migration assays and quantification of migrated A549 and H520 cells. (L-M) Tube formation assays and quantification of tube numbers in HUVECs. Data are presented as mean ± SD. P < 0.05 denotes statistical significance.

Article Snippet: Membranes were blocked with 5% skim milk in TBST, incubated with primary antibodies against EHF (1:1000, Catalog #: 27195-1-AP, Proteintech, Wuhan, China), TSG101 (1:1000, Catalog #: 14497-1-AP, Proteintech), Calnexin (1:1000, Catalog #: 10427-2-AP, Proteintech), CD9 (1:2000, Catalog #: 60232-1-Ig, Proteintech), RNF41 (1:500, Catalog #: 17233-1-AP, Proteintech), CD206 (1:1000, Catalog #: 18704-1-AP, Proteintech), TGF-β1 (1:1000, Catalog #: 26155-1-AP, Proteintech), VEGFA (1:1000, Catalog #: ab46154, Abcam, Cambridge, UK), and β-actin (1:1000, Catalog #: 66009-1-Ig, Proteintech) overnight at 4 °C, followed by HRP-conjugated secondary antibodies (1:5000, Catalog #: ab6721/ab205719, Abcam) for 1 h at room temperature.

Techniques: Co-Culture Assay, Control, Knockdown, Incubation, Transfection, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Staining, Migration

Journal: Regenerative Therapy

Article Title: Lung cancer cell-derived exosomal EHF drives M2 macrophage polarization via transcriptional activation of RNF41 to promote tumor progression

doi: 10.1016/j.reth.2026.101104

Figure Lengend Snippet: RNF41 mediates exosomal EHF-driven pro-tumorigenic effects in lung cancer. (A) Schematic of the rescue co-culture system: THP-1-M0 macrophages (control or RNF41-overexpressing) were incubated with exosomes from control (Exo Ctrl ) or EHF-knockdown (Exo KD−EHF ) A549/H520 cells. The medium from these co-cultures was used to treat A549/H520 cells or HUVECs. (B–C) Flow cytometry was used to analyze the proportion of CD206 + M2-polarized macrophages in THP-1-M0 cells. (D-E) ELISA was employed to quantify the levels of M2-associated cytokines (TGF-β1, IL-10, and VEGFA) in the CM from macrophages. (F-G) EdU assays were carried out to measure the proliferation of A549 and H520 cells. (H–I) Annexin V/PI flow cytometry was used to assess the apoptosis of A549 and H520 cells treated with CM. (J-K) Transwell assays were performed to evaluate the migration of A549 and H520 cells. (L-M) Tube formation assays were conducted on HUVECs treated with CM. Data are presented as mean ± SD. P < 0.05 denotes statistical significance.

Article Snippet: Membranes were blocked with 5% skim milk in TBST, incubated with primary antibodies against EHF (1:1000, Catalog #: 27195-1-AP, Proteintech, Wuhan, China), TSG101 (1:1000, Catalog #: 14497-1-AP, Proteintech), Calnexin (1:1000, Catalog #: 10427-2-AP, Proteintech), CD9 (1:2000, Catalog #: 60232-1-Ig, Proteintech), RNF41 (1:500, Catalog #: 17233-1-AP, Proteintech), CD206 (1:1000, Catalog #: 18704-1-AP, Proteintech), TGF-β1 (1:1000, Catalog #: 26155-1-AP, Proteintech), VEGFA (1:1000, Catalog #: ab46154, Abcam, Cambridge, UK), and β-actin (1:1000, Catalog #: 66009-1-Ig, Proteintech) overnight at 4 °C, followed by HRP-conjugated secondary antibodies (1:5000, Catalog #: ab6721/ab205719, Abcam) for 1 h at room temperature.

Techniques: Co-Culture Assay, Control, Incubation, Knockdown, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Migration

Journal: Regenerative Therapy

Article Title: Lung cancer cell-derived exosomal EHF drives M2 macrophage polarization via transcriptional activation of RNF41 to promote tumor progression

doi: 10.1016/j.reth.2026.101104

Figure Lengend Snippet: EHF knockdown inhibits tumor growth and downregulates M2-associated molecules in v ivo. Nude mice were subcutaneously implanted with Ctrl or KD-EHF A549 cells. (A) Tumor volume was measured every 5 days over 30 days. (B) Tumor weight. (C) Representative images of excised tumors. (D) Western blot analysis of RNF41, CD206, TGF-β1, and VEGFA expression levels in tumor tissues. Data are presented as mean ± SD. P < 0.05 denotes statistical significance.

Article Snippet: Membranes were blocked with 5% skim milk in TBST, incubated with primary antibodies against EHF (1:1000, Catalog #: 27195-1-AP, Proteintech, Wuhan, China), TSG101 (1:1000, Catalog #: 14497-1-AP, Proteintech), Calnexin (1:1000, Catalog #: 10427-2-AP, Proteintech), CD9 (1:2000, Catalog #: 60232-1-Ig, Proteintech), RNF41 (1:500, Catalog #: 17233-1-AP, Proteintech), CD206 (1:1000, Catalog #: 18704-1-AP, Proteintech), TGF-β1 (1:1000, Catalog #: 26155-1-AP, Proteintech), VEGFA (1:1000, Catalog #: ab46154, Abcam, Cambridge, UK), and β-actin (1:1000, Catalog #: 66009-1-Ig, Proteintech) overnight at 4 °C, followed by HRP-conjugated secondary antibodies (1:5000, Catalog #: ab6721/ab205719, Abcam) for 1 h at room temperature.

Techniques: Knockdown, Western Blot, Expressing