Journal: Bioactive Materials

Article Title: Microfluidic chip-integrated vascularized endometrial complexes: Mitochondrial function and paracrine crosstalk enhance regenerative potential

doi: 10.1016/j.bioactmat.2025.08.035

Figure Lengend Snippet: Single-cell RNA sequencing (scRNA-seq) analysis of the dynamically cultured EO and HEO complexes. a. UMAP visualization of 12 samples across four culture conditions: HEOe (HEO + estrogen), HEOp (HEO + estrogen + progesterone + cAMP), EOe (EO + estrogen), EOp (EO + estrogen + progesterone + cAMP). Endometrial epithelial cell clusters were annotated as main Epi (mEpi), ribosome-related Epi (rEpi), proliferative Epi (pEpi), and ciliated Epi (cEpi). Endometrial stromal cell clusters were annotated as main Str (mStr), ribosome-related Str (rStr), proliferative Str (pStr), and a second proliferative Str cluster (pStr2); the endothelial cell clusters included main endothelial cells (mEndo) and ribosome-related endothelial cells (rEndo). b–c. Relative abundance of epithelial (b) and stromal (c) clusters within their respective compartments across groups.]d. Comparison of the percentage of proliferative epithelial cells (pEpi, left panel) and proliferative stromal cells (pStr, right panel) between the dynamically cultured EO and HEO complexes under estrogen culture.e. Validation of enhanced proliferation in HEO complexes by flow cytometry. Significantly higher proportions of EdU + cells in epithelial cells of HEO vs. EO complexes under estrogen culture.f. Ligand-receptor pairs exhibiting significant differences in communication probability between HEOe and EOe complexes. Pathways of biological interest include WNT7A, WNT5A , BMP6 , and LGALS9. Dot size represents the maximum communication probability across compared groups; dot color indicates statistical significance ( P < 0.05). Cluster abbreviations: Epi (endometrial epithelial cells), Str (endometrial stromal cells) and Endo (endothelial cells).g. Circle plot showing the BMP6, LGALS9, WNT7A, WNT5A signaling networks among different cell types in EOe and HEOe complexes.

Article Snippet: The spheroids were stimulated with 100 μL of ECM with different treatments, including WNT5A (200 ng/mL, CSB-EP026138HU, CUSABIO, Wuhan, China), WNT7A (100 ng/mL, P06680 , Solarbio), DKK1 (200 ng/mL, C12B, Novoprotein), BOX5 (HY-123071A, MCE), anti-WNT5A neutralizing antibody (2 μg/mL, MAB645, R&D), anti-WNT7A neutralizing antibody (10 μg/mL, sc-365665, Santa Cruz), or VEGFA (10 ng/mL, C744, Novoprotein) as a positive control, by adding dropwise onto the collagen matrix.

Techniques: RNA Sequencing, Cell Culture, Comparison, Biomarker Discovery, Flow Cytometry

Journal: Bioactive Materials

Article Title: Microfluidic chip-integrated vascularized endometrial complexes: Mitochondrial function and paracrine crosstalk enhance regenerative potential

doi: 10.1016/j.bioactmat.2025.08.035

Figure Lengend Snippet: Endometrial epithelial and stromal cells enhance the formation of vascular networks in HUVECs within the HEO complex through WNT7A and WNT5A signaling pathways. a. Representative images of sprouting assays using HUVEC spheroids treated as indicated. HUVEC spheroids cultured with basic ECM medium are shown as the blank group (Control). VEGFA added to the basic ECM medium served as the positive control. Scale bar: 400 μm. b. Quantification of sprouting number was determined by Image J software. ∗ P < 0.05 versus Control; # P < 0.05 versus WNT7A; $ P < 0.05 versus DKK1; ^ P < 0.05 versus WNT5A; & P < 0.05 versus BOX5; one-way ANOVA was conducted for statistical analysis. c. Representative images of the tube formation of HUVECs on Matrigel surface treated as indicated. HUVECs cultured with basic ECM medium are shown as the blank (Control). VEGFA added to the basic ECM medium served as the positive control. Scale bar: 400 μm. d. Quantification of branch number using Image J software. ∗ P < 0.05 versus Control; $ P < 0.05 versus DKK1; &P < 0.05 versus BOX5; one-way ANOVA was conducted for statistical analysis.e. Vascular networks of complex with HUVECs and ESCs (HE), complex with HUVECs and EEOs (HO), and HEO treated with WNT7A (or its inhibitor DKK1) and WNT5A (or its inhibitor BOX5) as indicated. Vascular networks are depicted in green. Scale bar: 100 μm.f. Quantification of the vessel-positive area within the formed vascular network across different groups. ∗P < 0.05, ∗∗P < 0.01, Student's t-test.g. The electrical resistance of HEO and HE with DKK1 and WNT7A treatments, respectively. ∗∗∗P < 0.001, two-way ANOVA.h. The electrical resistance of HEO and HO with BOX5 and WNT5A treatments, respectively. ∗∗∗P < 0.001, two-way ANOVA.

Article Snippet: The spheroids were stimulated with 100 μL of ECM with different treatments, including WNT5A (200 ng/mL, CSB-EP026138HU, CUSABIO, Wuhan, China), WNT7A (100 ng/mL, P06680 , Solarbio), DKK1 (200 ng/mL, C12B, Novoprotein), BOX5 (HY-123071A, MCE), anti-WNT5A neutralizing antibody (2 μg/mL, MAB645, R&D), anti-WNT7A neutralizing antibody (10 μg/mL, sc-365665, Santa Cruz), or VEGFA (10 ng/mL, C744, Novoprotein) as a positive control, by adding dropwise onto the collagen matrix.

Techniques: Protein-Protein interactions, Cell Culture, Control, Positive Control, Software