Journal: Bioactive Materials

Article Title: Bioengineered extracellular vesicles escape lysosomal degradation and deliver Tet-PKM2 for macrophage immunometabolic reprogramming and periodontitis treatment

doi: 10.1016/j.bioactmat.2026.01.002

Figure Lengend Snippet: Generation of bioengineered LEVs (LEVs@TA) through in situ TA modifications. ( A ) Schematic illustration showing the interaction of TA with the phospholipid bilayer of LEVs via hydrogen bonding and the uptake of LEVs@TA by macrophages. ( B ) The percentages of CY5-TA-modified cells following incubation with gradient concentrations of CY5-TA (0, 0.1, 1, 5, and 10 μM) for 24 h (flow cytometry assay). ( C ) Colocalization of CY5-TA on HEK293T cells following incubation in 10 μM CY5-TA for 24 h (fluorescence microscopy). The nuclei were stained with DAPI (blue). ( D ) The percentages of CY5-TA-modified LEVs following incubation with gradient concentrations of CY5-TA (0, 10, 20, 50, and 100 μM) for 24 h (flow cytometry assay). ( E ) Colocalization of CY5-TA and PKH67-labeled LEVs (green) following incubation with 100 μM CY5-TA for 24 h (fluorescence microscopy). ( F ) Snapshots of CGMD simulations depicting the uptake of LEVs and LEVs@TA by macrophages at 0, 5, 10, 15, and 20 ns. ( G ) Representative in vivo fluorescence images showing good stability of DIO-labeled-LEVs@CY5-TA in vivo .

Article Snippet: PKH67-labeled SEVs and LEVs resuspended in complete medium were used to treat RAW 264.7 cells for 24 h. The cells were then fixed with 4 % PFA (Coolaber), permeabilized, and stained for cytoskeletal visualization using fluorescein phalloidin (1:1000 dilution; MCE) for 30 min.

Techniques: In Situ, Modification, Incubation, Flow Cytometry, Fluorescence, Microscopy, Staining, Labeling, In Vivo

Journal: Bioactive Materials

Article Title: Bioengineered extracellular vesicles escape lysosomal degradation and deliver Tet-PKM2 for macrophage immunometabolic reprogramming and periodontitis treatment

doi: 10.1016/j.bioactmat.2026.01.002

Figure Lengend Snippet: Endo/lysosomal escape capacity of bioengineered LEVs@TA following uptake by macrophages. ( A ) Schematic illustration showing the endo/lysosomal escape process of LEVs@TA within the cytoplasm of macrophages. After uptake by macrophages, LEVs@TA were entrapped within endo/lysosomes, and then TA underwent protonation and disassembled from LEVs in an acidic environment, leading to rupture of the endo/lysosomal structure. ( B ) Snapshots of CGMD simulations showing the disassembly of TA and LEVs in the lysosomal environment. ( C ) Colocalization of LysoTracker-labeled endo/lysosomes (violet) and PKH67-labeled LEVs or LEVs@TA (green) (fluorescence microscopy). The nuclei were stained with Hoechst (blue). ( D ) Quantification of the colocalization of endo/lysosomes and LEVs or LEVs@TA using the Pearson correlation coefficient ( n = 12). ( E ) Schematic illustration showing the leakage of calcein into the cytosol when TA diffused from LEVs@TA and destabilized the endo/lysosomal membranes. ( F ) The distribution of calcein (green) in macrophages treated with PBS, LEVs, and LEVs@TA (fluorescence microscopy). (G) Representative TEM images of macrophages showing the structure of lysosomes in macrophages treated with LEVs and LEVs@TA. The data are expressed as the mean ± SEM. Statistical analysis was performed with Student's t -test ( D ). ∗∗∗ p < 0.001 indicates significant differences between the indicated columns.

Article Snippet: PKH67-labeled SEVs and LEVs resuspended in complete medium were used to treat RAW 264.7 cells for 24 h. The cells were then fixed with 4 % PFA (Coolaber), permeabilized, and stained for cytoskeletal visualization using fluorescein phalloidin (1:1000 dilution; MCE) for 30 min.

Techniques: Labeling, Fluorescence, Microscopy, Staining

Journal: STAR Protocols

Article Title: Protocol for spatial quantitative analysis of cell division events across the epidermis of developing Hibiscus trionum petals

doi: 10.1016/j.xpro.2026.104354

Figure Lengend Snippet: Staining and mounting of the Hibiscus trionum petals at early stages of development (A) Early-stage floral bud at the final dissection step, after propidium iodide staining (red color). The double arrow indicates the dissection site. Scale bar, 100 μm. (B) Petals being mounted in a drop of Hoyer’s medium. (C) Coverslip placed over the petals and gently to spread the medium evenly. (D) Example slide showing EdU-labeled petals. (E) Zoom view of the coverslip region of panel D. (F) Higher magnification of selected petals prepared for imaging. Scale bar, 1 mm.

Article Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Techniques: Staining, Dissection, Labeling, Imaging

Journal: STAR Protocols

Article Title: Protocol for spatial quantitative analysis of cell division events across the epidermis of developing Hibiscus trionum petals

doi: 10.1016/j.xpro.2026.104354

Figure Lengend Snippet: Imaging EdU-labeled Hibiscus trionum petals at early stages of development to visualize the repartition of cell division events across the adaxial epidermis (A) Slide positioned under an upright confocal microscope using a Plan-Apochromat 10×/0.45 objective lens. (B) Example confocal image showing EdU-labeled nuclei (green) and propidium iodide-stained membranes (red) in the adaxial epidermis of early-stage petals. Newly synthesized DNA is visualized via fluorescent nucleotide analog 5-ethynyl-2′-deoxyuridine (EdU). Scale bar: 100 μm.

Article Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Techniques: Imaging, Labeling, Microscopy, Staining, Synthesized

Journal: STAR Protocols

Article Title: Protocol for spatial quantitative analysis of cell division events across the epidermis of developing Hibiscus trionum petals

doi: 10.1016/j.xpro.2026.104354

Figure Lengend Snippet: Identifying EdU-labeled nuclei using spot detection (A) Step 1/3 -Initial parameter setup for spot detection. (B) Step 2/3. Channel selection and parameter adjustment for EdU-labeled nuclei. For Step 3/3, see .

Article Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Techniques: Labeling, Selection

Journal: STAR Protocols

Article Title: Protocol for spatial quantitative analysis of cell division events across the epidermis of developing Hibiscus trionum petals

doi: 10.1016/j.xpro.2026.104354

Figure Lengend Snippet: Select EdU-labeled nuclei of the central proximo-distal stripe for filtered spot analysis (Step 3/3) (A) Application of a quality filter (“Quality above automatic threshold”) to detect EdU-labeled nuclei. (B) Stripe selection by X-position. The detected nuclei are further filtered using the histogram slider to isolate a central stripe. This step enables quantitative analysis of nuclei relative position along the petal proximo–distal axis.

Article Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Techniques: Labeling, Selection

Journal: STAR Protocols

Article Title: Protocol for spatial quantitative analysis of cell division events across the epidermis of developing Hibiscus trionum petals

doi: 10.1016/j.xpro.2026.104354

Figure Lengend Snippet: Create your Scatter plot of the Edu-labeled nuclei position across the proximo-distal axis of the petal (A) Select region of interest. (B) Plot settings. (C) Exported final scatter plot.

Article Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Techniques: Labeling

Journal: STAR Protocols

Article Title: Protocol for spatial quantitative analysis of cell division events across the epidermis of developing Hibiscus trionum petals

doi: 10.1016/j.xpro.2026.104354

Figure Lengend Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Article Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Techniques: Labeling

Journal: STAR Protocols

Article Title: Protocol for spatial quantitative analysis of cell division events across the epidermis of developing Hibiscus trionum petals

doi: 10.1016/j.xpro.2026.104354

Figure Lengend Snippet: Analysis of the spatial distribution of EdU-labeled nuclei along the proximodistal axis of the petal (A) Example of an Excel file exported from Imaris, containing XYZ positions of EdU-labeled nuclei. (B) Structure of the formatted R input file (‘R_Results’ sheet), including six key columns required for analysis: Genotype, Stage, Petal, Y (nuclei position), L (petal length), and YNorm (normalized position). (C) Calculation of the relative position of each EdU-labeled nucleus by normalizing its Y coordinate to the total petal length (YNorm = Y/L). (D) Output from script section #1: Density plots of EdU-labeled nuclei per individual petals. (E) Output from script section #2: Pooled density plots of EdU-labeled nuclei per genotype.

Article Snippet: Example of output table showing EdU-labeled nuclei coordinates extracted from Imaris

Techniques: Labeling