Journal: Molecular medicine reports

Article Title: BCG stimulation promotes dendritic cell proliferation and expression of VDR and CYP27B1 in vitamin D‑deficient mice.

doi: 10.3892/mmr.2019.10780

Figure Lengend Snippet: Figure 2. Effect of BCG on BMDC differentiation and maturation. (A) Cell morphology during BMDC induction from days 1 to 6, as observed by light micros- copy. (B) BCG stimulated BMDC maturation. (C) Phenotypic changes of the BMDCs induced by BCG stimulation and (D) the levels of the surface molecules of DCs. All images were captured under identical magnification and microscopy conditions. **P<0.01 vs. 0 mg/ml in the normal control or vitamin D‑deficiency groups. ##P<0.01, vitamin D‑deficiency group vs. normal control group. Scale bar, 20 µm. BCG, Bacillus Calmette‑Guérin; BMDC, bone marrow‑derived dendritic cell; CD11c, integrin alpha‑X; CD80, T‑lymphocyte activation antigen CD80; MHC‑I, major histocompatibility complex class I; MHC‑II, major histocompatibility complex class II; CD86, T‑lymphocyte activation antigen CD86.

Article Snippet: The cell concentration was adjusted to 1x106 cells/ml and the cells were fixed with 0.1 ml polyformaldehyde for 30 min at 25 ̊C Subsequently, fluorescein isothiocyanate‐labeled integrin alpha‐X (CD11c; 1:50; 130‐110‐700; Miltenyi Biotech), T‐lymphocyte activation antigen CD86 (CD86; 1:50; 130‐102‐506; Miltenyi Biotech), T-lymphocyte activation antigen cd80 (cd80; 1:50; 130‐116‐459; Miltenyi Biotech), major histocompatibility complex class i (MHc-i; 1:50; ab95572; abcam) and MHC‐II (1:50; 130‐112‐386; Miltenyi Biotech) were added, respectively.

Techniques: Microscopy, Control, Activation Assay, Immunopeptidomics

Journal: iScience

Article Title: Microglial transcription profiles in mouse and human are driven by APOE4 and sex.

doi: 10.1016/j.isci.2021.103238

Figure Lengend Snippet: Figure 2. Sex and APOE genotype drive amyloid pathology and microglial gene expression profiles in APOE-FAD mice (A) Representative images of immunohistochemical staining for amyloid-b (red) and the microglial marker IBA1 (green) in male and female APOE3- and APOE4-FAD mice. Scale bar, 50 mm. (B) Samples separate by APOE and sex along PC1, with APOE3-FAD males clustering separately from mice of both sexes and APOE3-FAD females. A large proportion of the genes driving this separation (76.4%) are DAM-APOE genes. APOE3-FAD males are shown in yellow, APOE3-FAD females in gray, APOE4-FAD males in magenta, and APOE4-FAD females in teal. (C) Heatmap of the DAM-APOE genes associated with sample clustering along PC1. Gene expression levels were normalized, scaled, and centered and are displayed as Z scores. (D and E) Levels of the DAM-APOE marker, CD11c, are increased in APOE3-FAD females and in APOE4-FAD mice of both sexes, both at the (D) transcript and (E) protein levels. (D) Gene expression of Itgax in normalized counts; data are presented as mean (GSEM) values; n = 5–7/group. *p < 0.05, **p < 0.01; one-way ANOVA with Tukey’s multiple comparisons test correction. (E) Representative immunofluorescence images of microglia (IBA1, red) and CD11c (green) in the hippocampal subiculum of male and female APOE3- and APOE4-FAD mice. Scale bar, 25 mm.

Article Snippet: For IBA1/CD11c double labeling, antigen retrieval was performed using boiling EDTA and sections were incubated for 2 days at 4 C in primary antibodies directed against IBA1 (1:500) and CD11c (#NB110-97871, 1:500, Novus Biologicals).

Techniques: Gene Expression, Immunohistochemical staining, Staining, Marker

Journal: Journal of nanobiotechnology

Article Title: Lactiplantibacillus plantarum -derived extracellular vesicles alleviate acute lung injury by inhibiting ferroptosis of macrophages.

doi: 10.1186/s12951-025-03405-y

Figure Lengend Snippet: Fig. 2 LpEVs alleviated lung inflammation and injury in mice. A. Schematic depiction of the experiment. B. Representative images showing histopatho logical alterations in lungs of different treatment groups. ICR mice (n = 5) were sitimulated with LPS (2.5 mg/kg, intratracheally) for 3 h, then administered PBS or LpEVs (100 µg) via the tail vein injection method. Mice were euthanized 24 h post-treatment. Scale bar = 200 μm. C. Representative F4/80 and MPO IHC staining images in different lungs. Scale bars represent 50 μm. D. Protein concentration in BALF. E. Cell counts in BALF. F. The concentration of cytokines/chemokines in the BALF was assessed using ELISA. G. Flow cytometry analysis of neutrophils (CD11b+ Ly6G+) in the BALF. H. Flow cytometry analysis of macrophages (CD11b+ F4/80+) in the BALF. I. Flow cytometry analysis of alveolar macrophages (CD11c+ Siglec F+) in the BALF

Article Snippet: The population of alveolar macrophages (anti-CD11c, anti-Siglec F) (E-AB-F0991E, Elabscience, Wuhan, China) (12-1702- 82, Thermo Fisher, USA), macrophages (anti-F4/80, anti-CD11b) (E-AB-F0995D, E-AB-F1081E, Elabscience, Wuhan, China), and neutrophils (anti-CD11b, anti-Ly6G) (E-AB-F1081E, E-AB-F1108C, Wuhan, China) were assessed using flow cytometry with specific antibodies.

Techniques: Injection, Immunohistochemistry, Protein Concentration, Concentration Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry

Journal: Journal of nanobiotechnology

Article Title: Lactiplantibacillus plantarum -derived extracellular vesicles alleviate acute lung injury by inhibiting ferroptosis of macrophages.

doi: 10.1186/s12951-025-03405-y

Figure Lengend Snippet: Fig. 4 LpEVs treatment inhibited LPS-induced ferroptosis in vivo. A. The levels of reactive oxygen species (ROS) in lung BALF cells were evaluated using DCFH-DA. The left panel displays a flow cytometry diagram, while the right panel shows the mean fluorescence intensity of ROS. B. Relative MDA content in lungs. C. Relative Fe2+ content in lungs. D. Relative GSH content in lungs. E. Relative mRNA expression of ferroptotic-related genes were analyzed by qRT-PCR. F. The expression levels of ferroptotic-related proteins in lungs were analyzed by western blotting. ACTIN is adopted as the loading control. G. Representative IHC staining images for GPX4, SLC7A11 and ACSL4 were obtained from different lung samples. Scale bars represent 50 μm. H. The statisti cal graph of the expression of GPX4, SLC7A11 and ACSL4 in IHC staining. I. Representative IHC staining images for macrophages M1 marker CD86 and M1 marker CD206 in lungs. Scale bars represent 50 μm. J. The statistical graph of the expression of CD86 and CD206 in IHC staining. Quantification of proteins expresion levels in IHC staining were determined by image J software. The values for the LPS-PBS and LPS-EV were expressed as percentages of the values for the Sham group. *P < 0.05, **P < 0.01, ***P < 0.001, ***P < 0.0001 and ns, no significant difference

Article Snippet: The population of alveolar macrophages (anti-CD11c, anti-Siglec F) (E-AB-F0991E, Elabscience, Wuhan, China) (12-1702- 82, Thermo Fisher, USA), macrophages (anti-F4/80, anti-CD11b) (E-AB-F0995D, E-AB-F1081E, Elabscience, Wuhan, China), and neutrophils (anti-CD11b, anti-Ly6G) (E-AB-F1081E, E-AB-F1108C, Wuhan, China) were assessed using flow cytometry with specific antibodies.

Techniques: In Vivo, Flow Cytometry, Fluorescence, Expressing, Quantitative RT-PCR, Western Blot, Control, Immunohistochemistry, Marker, Software

Journal: Journal of nanobiotechnology

Article Title: Lactiplantibacillus plantarum -derived extracellular vesicles alleviate acute lung injury by inhibiting ferroptosis of macrophages.

doi: 10.1186/s12951-025-03405-y

Figure Lengend Snippet: Fig. 5 LpEVs facilitated the transition of macrophages towards an anti-inflammatory phenotype. A. Confocal laser scanning microscopy images dis played the presence of Dio-labeled LpEVs in MH-S cells. Green: Dio labeled LpEVs; Blue: Hoechst. Scale bar = 20 μm. B. The Dio-positive MH-S cells were determined by flow cytometry. C. Relative mRNA expression levels of inflammatory factors were assessed by qRT-PCR. M1 proinflammatory genes: Il1β, Il6, Tnfα, and iNos, M2 anti-inflammatory genes: Arg-1, and Cd206. The M1 macrophages were induced by LPS treatment, and M2 macrophages were induced by IL-4 treatment. D. The levels of pro-inflammatory and anti-inflammatory cytokines in the medium supernatants were quantified using ELISA. E. The expression of CD86 (an M1 marker) on MH-S cells 12 h after exposure to LPS was assessed using flow cytometry, with representative histograms and average relative mean fluorescence intensity (MFI) provided. F. Flow cytometry was used to detect the expression of CD206, an M2 marker, on MH-S cells 12 h after exposure to IL4

Article Snippet: The population of alveolar macrophages (anti-CD11c, anti-Siglec F) (E-AB-F0991E, Elabscience, Wuhan, China) (12-1702- 82, Thermo Fisher, USA), macrophages (anti-F4/80, anti-CD11b) (E-AB-F0995D, E-AB-F1081E, Elabscience, Wuhan, China), and neutrophils (anti-CD11b, anti-Ly6G) (E-AB-F1081E, E-AB-F1108C, Wuhan, China) were assessed using flow cytometry with specific antibodies.

Techniques: Confocal Laser Scanning Microscopy, Labeling, Flow Cytometry, Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Marker, Fluorescence

Journal: Journal of nanobiotechnology

Article Title: Lactiplantibacillus plantarum -derived extracellular vesicles alleviate acute lung injury by inhibiting ferroptosis of macrophages.

doi: 10.1186/s12951-025-03405-y

Figure Lengend Snippet: Fig. 6 LpEVs attenuated ferroptosis in macrophages. A. The ROS level in MH-S was assessed by flow cytometry. B. Relative MDA content in MH-S. C. Rela tive GSH content in MH-S. D. The intercellular Fe2+ was detected by FerroOrange (a specific probe of Fe2+). Scale bar = 100 μm. E. The expression levels of ferroptotic-related proteins and NRF2-related proteins in MH-S were detected by western blot. ACTIN is adopted as the loading control. F. SLC7A11, GPX4 and ACSL4 immunofluorescent staining results of differently treated MH-S. Scale bar = 20 μm. G. NRF2 and HO-1 immunofluorescent staining results of differently treated MH-S. Scale bar = 20 μm

Article Snippet: The population of alveolar macrophages (anti-CD11c, anti-Siglec F) (E-AB-F0991E, Elabscience, Wuhan, China) (12-1702- 82, Thermo Fisher, USA), macrophages (anti-F4/80, anti-CD11b) (E-AB-F0995D, E-AB-F1081E, Elabscience, Wuhan, China), and neutrophils (anti-CD11b, anti-Ly6G) (E-AB-F1081E, E-AB-F1108C, Wuhan, China) were assessed using flow cytometry with specific antibodies.

Techniques: Flow Cytometry, Expressing, Western Blot, Control, Staining

Journal: Journal of nanobiotechnology

Article Title: Lactiplantibacillus plantarum -derived extracellular vesicles alleviate acute lung injury by inhibiting ferroptosis of macrophages.

doi: 10.1186/s12951-025-03405-y

Figure Lengend Snippet: Fig. 7 LpEVs regulated ferroptosis in macrophages via delivery of cbn-let-7. A. miRNA microarray analyze the miRNA expression profile of LpEVs. B. The 4 miRNAs (cel-mir-4937, cbn-let-7, ngi-miR-92a and gsa-mir-9394b) expression level in LpEVs. C. cbn-let-7 expression in PBS and LpEVs treated MH-S cells. D, E. Structure and luciferase result of dual luciferase reporter gene. F. The relative expression of cbn-let-7 in NC, cbn-let-7 mimic and inhibitor treated MH-S cells. G. The protein expression of of ACSL4 and GPX4 in NC, cbn-let-7 mimic and inhibitor treated MH-S cells. H. The expression of cbn-let-7 in different treated cells. I. The protein expression of of ACSL4 and GPX4 in different treated cells. ACTIN is adopted as the loading control. J. The intercellular Fe2+ in different treated cells was detected by FerroOrange (a specific probe of Fe2+)

Article Snippet: The population of alveolar macrophages (anti-CD11c, anti-Siglec F) (E-AB-F0991E, Elabscience, Wuhan, China) (12-1702- 82, Thermo Fisher, USA), macrophages (anti-F4/80, anti-CD11b) (E-AB-F0995D, E-AB-F1081E, Elabscience, Wuhan, China), and neutrophils (anti-CD11b, anti-Ly6G) (E-AB-F1081E, E-AB-F1108C, Wuhan, China) were assessed using flow cytometry with specific antibodies.

Techniques: Microarray, Expressing, Luciferase, Control