Journal: Bioactive Materials

Article Title: Inhalable PD-L1-engineered hybrid cellular vesicles suppress excessive neutrophil activation and restore mitochondrial homeostasis to alleviate ischemia–reperfusion lung injury and pneumonia

doi: 10.1016/j.bioactmat.2026.03.024

Figure Lengend Snippet: Characterization of Res-PD-L1@nmEVs . (A) Schematic illustration of the Res-PD-L1@nmEVs synthesis procedure. (B-D) Representative transmission electron microscopy (TEM) images, dynamic light scattering (DLS) size distributions, and zeta potential measurements of nEVs, PD-L1@mEVs, PD-L1@nmEVs, and Res-PD-L1@nmEVs. (E) PD-L1 expression in PD-L1-overexpressing MSCs (OE-PD-L1) and negative control (NC) MSCs, and CD11b expression in HL60 cells before and after DMSO stimulation, as determined by Western blot. (F) Expression levels of neutrophil membrane markers (CD11b, CXCR2, RAGE, TLR2) and the exosomal marker CD63 in the four EV types. (G) Fluorescence co-localization images of DiO-labeled nEVs (green) and DiL-labeled PD-L1@mEVs (red) after fusion, demonstrating hybrid vesicle formation. (H) Size stability of Res-PD-L1@nmEVs stored at 4 °C and 37 °C for 7 days. (I-K) Binding and neutralization capacity of Res-PD-L1@nmEVs against inflammatory cytokines (TNF-α, IL-6, IL-1β) in vitro. ∗ vs. 0ug/ml; # vs. 100 μg/ml, p < 0.05, n = 5.

Article Snippet: For detecting neutrophil characteristic proteins, the primary antibodies comprised CD11b (HA722075, HUABIO, China), CXCR2 (ER1906-87, HUABIO, China), RAGE (ET1702-27, HUABIO, China), TLR2 (ET1705-92, HUABIO, China), TNFR1(RT1624, HUABIO, China), IL6R ( AWA43078 , Abiowell, China) and the exosome marker CD63 (ET1607-2, HUABIO, China).

Techniques: Transmission Assay, Electron Microscopy, Zeta Potential Analyzer, Expressing, Negative Control, Western Blot, Membrane, Marker, Fluorescence, Labeling, Binding Assay, Neutralization, In Vitro

Journal: Bioactive Materials

Article Title: Inhalable PD-L1-engineered hybrid cellular vesicles suppress excessive neutrophil activation and restore mitochondrial homeostasis to alleviate ischemia–reperfusion lung injury and pneumonia

doi: 10.1016/j.bioactmat.2026.03.024

Figure Lengend Snippet: Res-PD-L1@nmEVs Suppresses Neutrophil Activation and Preserves Mitochondrial Integrity via PD-L1 Delivery (A-B) Rats subjected to lung IRI received nebulized administration of different formulations (Res, nEVs, PD-L1@mEVs, PD-L1@nmEVs, or Res-PD-L1@nmEVs) before ischemia and after reperfusion. Lung tissues were collected 2 h post-reperfusion. (A) Representative immunofluorescence images showing the expression and localization of CD11b (green), MPO (red), and PD-1 (yellow) in lung sections across treatment groups. (B) Enlarged view of the IRI group from (A). (C-D) mRNA levels of CD95 (C) and CD206 (D) in lung tissues. (E-F) Levels of myeloperoxidase (MPO) (E) and matrix metalloproteinase-9 (MMP-9) (F) in bronchoalveolar lavage fluid (BALF). (G-I) (G) Representative transmission electron microscopy (TEM) images of lung tissues (scale bar: 2 μm). (H) Proportion of damaged mitochondria. (I) Average number of mitophagic events per cell. (J) Immunofluorescence co-localization of mitochondrial marker TOMM20 (red) and EpCAM (green) in lung tissues (nuclei stained with DAPI, scale bar: 50 μm). (K-L) Protein expression levels of Beclin-1 (K) and LC3 (L) in lung tissues, with insets showing immunofluorescence co-localization of Beclin-1 (green) and LC3 (red) across treatment groups (nuclei stained with DAPI, scale bar: 50 μm). ∗ vs. Sham; # vs. IRI; & vs. IRI + PD-L1@nmEVs, p < 0.05.

Article Snippet: For detecting neutrophil characteristic proteins, the primary antibodies comprised CD11b (HA722075, HUABIO, China), CXCR2 (ER1906-87, HUABIO, China), RAGE (ET1702-27, HUABIO, China), TLR2 (ET1705-92, HUABIO, China), TNFR1(RT1624, HUABIO, China), IL6R ( AWA43078 , Abiowell, China) and the exosome marker CD63 (ET1607-2, HUABIO, China).

Techniques: Activation Assay, Immunofluorescence, Expressing, Transmission Assay, Electron Microscopy, Marker, Staining

Journal: STAR Protocols

Article Title: Protocol for isolating and culturing microglia from the adult mouse brain using a magnetic-activated cell sorting system

doi: 10.1016/j.xpro.2026.104471

Figure Lengend Snippet: Representative workflow of isolation and culture of adult mouse microglia (A and B) Dissect the brain into small pieces on ice in Petri dish. (C) Collect cell pellets in C-tubes following mechanical/enzymatic dissociation using a gentleMACS dissociator. (D) Preparation of cell straining and debris removal processes. (E) Perform debris removal by carefully overlaying ice-cold PBS onto the cell suspension. (F) After centrifugation, identify three layers; the middle, yellowish layer corresponds to myelin dna debris. (G) Enrich CD11b+ cells by magnetic separation using appropriate columns. (H) Seed isolated microglia onto 6-wll culture plates for downstream assays.

Article Snippet: CD11b Microbead (Clone: M1/70.15.11.5) (1:10 ratio) , Miltenyi Biotech , Cat#130-049-601; RRID: AB_2927911.

Techniques: Isolation, Suspension, Centrifugation

Journal: STAR Protocols

Article Title: Protocol for isolating and culturing microglia from the adult mouse brain using a magnetic-activated cell sorting system

doi: 10.1016/j.xpro.2026.104471

Figure Lengend Snippet: Flow cytometry for MACS-isolated microglia purity (A and B) Total CD11b-positive cells from MACS columns; (C) Cell viability assessed by Zombie Red staining; (D) Infiltrating leukocytes, including neutrophils (Ly6G+) and T lymphocytes (CD3+); (E) Proportions of microglia (CD11b+CD45low) versus monocytes/border-associated macrophages (CD11b+CD45high); (F) Proportion of resting (homeostatic) microglia identified as CD11b+TMEM119+ cells.

Article Snippet: CD11b Microbead (Clone: M1/70.15.11.5) (1:10 ratio) , Miltenyi Biotech , Cat#130-049-601; RRID: AB_2927911.

Techniques: Flow Cytometry, Isolation, Staining

Journal: PLOS One

Article Title: Inhibition of neutrophil infiltration and NETs formation ameliorates neuropsychiatric and renal dysfunction in MRL/lpr mice with lupus

doi: 10.1371/journal.pone.0348011

Figure Lengend Snippet: A) Upper half: Multiplex immunofluorescence staining of CD11b, CD16, and MAP2 in brain sections from various groups of mice, with localized positive areas enlarged; white arrowheads indicate CD11b/CD16 double-positive cells. Lower half: Multiplex immunofluorescence staining of NE and MPO, with localized positive areas enlarged. The brain region schematic used as a reference is derived from the Allen Brain Atlas. B) and C) Statistical count of CD11b positive and CD11b/CD16 double-positive cells across different groups. D) and E) Semi-quantitative analysis of fluorescence intensity for NE and MPO in different groups. Statistical significance is indicated as follows: * p < 0.05, ** p < 0.01 compared to the MRL/MpJ group; # p < 0.05, ## p < 0.01 compared to the MRL/lpr+Avacopan group.

Article Snippet: Sections were incubated with primary antibodies overnight at 4°C, including: CD11b (1:2000, GB15058, Servicebio), CD16 (1:200, GB14028, Servicebio), MAP2 (1:2000, GB11128-2, Servicebio), NE (1:8000, ET1702-78, HUABIO), MPO (1:500, GB11224, Servicebio).

Techniques: Multiplex Assay, Immunofluorescence, Staining, Derivative Assay, Fluorescence

Journal: PLOS One

Article Title: Inhibition of neutrophil infiltration and NETs formation ameliorates neuropsychiatric and renal dysfunction in MRL/lpr mice with lupus

doi: 10.1371/journal.pone.0348011

Figure Lengend Snippet: A) Kidney function assessment of various mouse groups at week 20, evaluated by measuring the urine albumin-to-creatinine ratio. B) Upper half: PAS staining of kidney tissue was used to visualize renal tissue architecture and structural injury. Immune cell infiltration was assessed by multiplex immunofluorescence staining for CD11b and CD16; lower half: Multiplex immunofluorescence staining for CD11b, CD16, NE, MPO with localized positive areas enlarged. C) and D) Statistical count of CD11b positive and CD11b/CD16 double-positive cells across different groups. E) and F) Semi-quantitative analysis of fluorescence intensity for NE and MPO in different groups; * indicates comparisons of NE fluorescence intensity, # indicates comparisons of MPO fluorescence intensity. * p < 0.05, ** p < 0.01 compared to the MRL/MpJ group; # p < 0.05, ## p < 0.01 compared to the MRL/lpr+Avacopan group. G) Western blot bands for IL-6, IL-17, TNF-α, and β-actin in different groups. H and I) Semi-quantitative analysis of Western blot results for brain and kidney tissues respectively; * p < 0.05, ** p < 0.01.

Article Snippet: Sections were incubated with primary antibodies overnight at 4°C, including: CD11b (1:2000, GB15058, Servicebio), CD16 (1:200, GB14028, Servicebio), MAP2 (1:2000, GB11128-2, Servicebio), NE (1:8000, ET1702-78, HUABIO), MPO (1:500, GB11224, Servicebio).

Techniques: Staining, Multiplex Assay, Immunofluorescence, Fluorescence, Western Blot

Journal: Cell Reports Medicine

Article Title: Macrophage-mimetic photothermal nanotherapeutics regulate mitochondrial homeostasis and inflammatory cascades in lung ischemia-reperfusion injury

doi: 10.1016/j.xcrm.2026.102768

Figure Lengend Snippet: Preparation and characterization of Rg3@PACVs (A–C) Characterization of mPDA, Rg3@mPDA, and Rg3@PACVs using transmission electron microscopy (TEM) and dynamic light scattering (DLS) for hydrodynamic size distribution and zeta potential analysis (scale bars, 100 nm). (D) Elemental mapping of carbon (C), nitrogen (N), oxygen (O), phosphorus (P), and sulfur (S) in Rg3@PACVs (scale bars, 100 nm). (E) Fourier transform infrared spectroscopy (FTIR) spectra of mPDA, Rg3, Mac-CVs, and Rg3@PACVs. (F) The expression of macrophage-membrane markers including Toll-like receptor 2 (TLR2), integrin subunit alpha M (CD11b), C-X-C motif chemokine receptor 4 (CXCR4), receptor for advanced glycation end-products (RAGE), interleukin-6 receptor (IL-6R), and tumor necrosis factor receptor 1 (TNFR1) detected by digital western blot analysis. (G) Fluorescence of Dio-labeled Mac-CVs and Dil-labeled Rg3@mPDA after ultrasonic extrusion to form Dio/Dil-labeled Rg3@PACVs (scale bars, 1 μm). (H) Near-infrared (NIR, 808nm) images of Rg3, mPDA, Mac-CVs, and Rg3@PACVs in microcentrifuge tubes. (I and J) Temperature changes of Rg3@PACVs under different laser power intensities (I) and the photothermal effect curves of Rg3@PACVs at various concentrations (J). (K) Photothermal stability of Rg3@PACVs. (L) Size stability of Rg3@PACVs after 1-week storage at 4°C and 37°C.

Article Snippet: CD11b Recombinant Rabbit Monoclonal Antibody , HUABIO , Cat# HA722075; RRID: AB_3096756.

Techniques: Transmission Assay, Electron Microscopy, Zeta Potential Analyzer, Fourier Transform Infrared Spectroscopy, Spectroscopy, Expressing, Membrane, Western Blot, Fluorescence, Labeling

Journal: Cell Reports Medicine

Article Title: Macrophage-mimetic photothermal nanotherapeutics regulate mitochondrial homeostasis and inflammatory cascades in lung ischemia-reperfusion injury

doi: 10.1016/j.xcrm.2026.102768

Figure Lengend Snippet: Rg3@PACVs ameliorate lung injury by suppressing inflammatory responses in rat lung ischemia-reperfusion injury (IRI) (A) Lung injury assessment after tail vein injection of Rg3, PACVs, or Rg3@PACVs with near-infrared (NIR) irradiation during 2-h reperfusion following 1-h pulmonary ischemia. (B) IVIS images of major organs harvested 30 min and 1, 2, 4, 8, 12, and 24 h after intravenous injection. (C–E) Plasma levels of inflammatory cytokines IL-6 (C), IL-1β (D), and TNF-α (E). (F) Total cell counts in bronchoalveolar lavage fluid (BALF). (G and H) Hematoxylin and eosin (H&E) staining of lung tissue (scale bars, 50 μm) and lung injury scores. (I) Wet/dry weight ratio assessing pulmonary edema. (J and K) Immunofluorescence analysis of (J) neutrophil marker CD11b (red); (K) myeloperoxidase (MPO) intensity reflecting neutrophil degranulation (green); scale bars, 50 μm. (L) Zonula occludens-1 (ZO-1) expression (red) indicating alveolar-capillary barrier integrity; scale bars, 50 μm. (M) Apoptosis detection by TUNEL assay. Nuclei counterstained with DAPI (blue). Scale bars: 50 μm. Data are presented as the mean ± SD and analyzed using one-way ANOVA with Tukey’s post hoc test ( ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001 vs. Sham; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. IRI; & p < 0.05, && p < 0.01, &&& p < 0.001 vs. IRI+Rg3@PACVs; n = 5 biological replicates).

Article Snippet: CD11b Recombinant Rabbit Monoclonal Antibody , HUABIO , Cat# HA722075; RRID: AB_3096756.

Techniques: Injection, Irradiation, Clinical Proteomics, Staining, Immunofluorescence, Marker, Expressing, TUNEL Assay

Journal: Cell Reports Medicine

Article Title: Macrophage-mimetic photothermal nanotherapeutics regulate mitochondrial homeostasis and inflammatory cascades in lung ischemia-reperfusion injury

doi: 10.1016/j.xcrm.2026.102768

Figure Lengend Snippet: Preparation and characterization of Rg3@PACVs (A–C) Characterization of mPDA, Rg3@mPDA, and Rg3@PACVs using transmission electron microscopy (TEM) and dynamic light scattering (DLS) for hydrodynamic size distribution and zeta potential analysis (scale bars, 100 nm). (D) Elemental mapping of carbon (C), nitrogen (N), oxygen (O), phosphorus (P), and sulfur (S) in Rg3@PACVs (scale bars, 100 nm). (E) Fourier transform infrared spectroscopy (FTIR) spectra of mPDA, Rg3, Mac-CVs, and Rg3@PACVs. (F) The expression of macrophage-membrane markers including Toll-like receptor 2 (TLR2), integrin subunit alpha M (CD11b), C-X-C motif chemokine receptor 4 (CXCR4), receptor for advanced glycation end-products (RAGE), interleukin-6 receptor (IL-6R), and tumor necrosis factor receptor 1 (TNFR1) detected by digital western blot analysis. (G) Fluorescence of Dio-labeled Mac-CVs and Dil-labeled Rg3@mPDA after ultrasonic extrusion to form Dio/Dil-labeled Rg3@PACVs (scale bars, 1 μm). (H) Near-infrared (NIR, 808nm) images of Rg3, mPDA, Mac-CVs, and Rg3@PACVs in microcentrifuge tubes. (I and J) Temperature changes of Rg3@PACVs under different laser power intensities (I) and the photothermal effect curves of Rg3@PACVs at various concentrations (J). (K) Photothermal stability of Rg3@PACVs. (L) Size stability of Rg3@PACVs after 1-week storage at 4°C and 37°C.

Article Snippet: Recombinant Anti-CD11b antibody , Servicebio , Cat# GB15058.

Techniques: Transmission Assay, Electron Microscopy, Zeta Potential Analyzer, Fourier Transform Infrared Spectroscopy, Spectroscopy, Expressing, Membrane, Western Blot, Fluorescence, Labeling

Journal: Cell Reports Medicine

Article Title: Macrophage-mimetic photothermal nanotherapeutics regulate mitochondrial homeostasis and inflammatory cascades in lung ischemia-reperfusion injury

doi: 10.1016/j.xcrm.2026.102768

Figure Lengend Snippet: Rg3@PACVs ameliorate lung injury by suppressing inflammatory responses in rat lung ischemia-reperfusion injury (IRI) (A) Lung injury assessment after tail vein injection of Rg3, PACVs, or Rg3@PACVs with near-infrared (NIR) irradiation during 2-h reperfusion following 1-h pulmonary ischemia. (B) IVIS images of major organs harvested 30 min and 1, 2, 4, 8, 12, and 24 h after intravenous injection. (C–E) Plasma levels of inflammatory cytokines IL-6 (C), IL-1β (D), and TNF-α (E). (F) Total cell counts in bronchoalveolar lavage fluid (BALF). (G and H) Hematoxylin and eosin (H&E) staining of lung tissue (scale bars, 50 μm) and lung injury scores. (I) Wet/dry weight ratio assessing pulmonary edema. (J and K) Immunofluorescence analysis of (J) neutrophil marker CD11b (red); (K) myeloperoxidase (MPO) intensity reflecting neutrophil degranulation (green); scale bars, 50 μm. (L) Zonula occludens-1 (ZO-1) expression (red) indicating alveolar-capillary barrier integrity; scale bars, 50 μm. (M) Apoptosis detection by TUNEL assay. Nuclei counterstained with DAPI (blue). Scale bars: 50 μm. Data are presented as the mean ± SD and analyzed using one-way ANOVA with Tukey’s post hoc test ( ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001 vs. Sham; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. IRI; & p < 0.05, && p < 0.01, &&& p < 0.001 vs. IRI+Rg3@PACVs; n = 5 biological replicates).

Article Snippet: Recombinant Anti-CD11b antibody , Servicebio , Cat# GB15058.

Techniques: Injection, Irradiation, Clinical Proteomics, Staining, Immunofluorescence, Marker, Expressing, TUNEL Assay