Journal: Frontiers in Immunology

Article Title: Uterine macrophages and NK cells exhibit population and gene-level changes after implantation but maintain pro-invasive properties

doi: 10.3389/fimmu.2024.1364036

Figure Lengend Snippet: Pre- and post-implantation bulk CD14+CD64+ macrophages/monocytes promote invasion of primary EVTs as strongly as pre- and post-implantation bulk NK cells ex vivo on an implantation-on-a-chip device. (A) Graphical representation of the experimental approach taken to prepare and load purified endometrial and decidual bulk NK cells (CD56+CD3- cells, top flow cytometry plots) and bulk macrophages (CD14+CD64+, bottom flow cytometry plots) onto the implantation-on-a-chip (IOC). Shown are raw flow cytometry data of pre- and post-sort purity of NK cells and macrophages. Events shown have undergone forward and side scatter gating, singlet discrimination, live/dead discrimination, and CD45+ gating. “Fetal” denotes the channel of the IOC containing fluorescently labeled fetal EVTs, “ECM” denotes the channel of the IOC containing mock extracellular matrix, and “Maternal vessel” denotes the channel of the IOC containing endothelial cells, as described in the Methods section. Note the horizontal orientation of the actual devices. For purposes of data acquisition and interpretation, the IOC is imaged and shown in (B, C) in the vertical orientation, with the fetal channel at the top, ECM in the middle, and the maternal vessel channel at the bottom. (B) Brightfield imaging showing successful embedding of purified endometrial macrophages (Endo Mac), first trimester decidual macrophages (FT Mac), endometrial NK cells (Endo NK) and first trimester decidual NK cells (FT NK) onto the IOC. (C) Representative fluorescent imaging of labeled EVTs invading into the ECM and into the maternal vessel channel on the indicated post-seeding day. Days 1 and 3 post-seeding are shown. Channels on each device are separated by white dotted lines. After 1 day, note that most of the EVTs in any condition have not yet entered the ECM. After 3 days of culture, most of the EVTs are seen in the ECM channel in the macrophage and NK cell conditions. Scale bars represent 200μM. (D) Quantification of invasion area of EVTs beyond the fetal channel at day 3 of culture. One-way ANOVA with Tukey’s multiple comparison test was performed (n = 3 independent biological samples and 3 independent devices per biological sample). Data are presented as mean ± SD. Area of invasion at day 1 was not significantly different among groups. Invasion area of EVTs in the macrophage and NK cell conditions were all significantly different than the control (no immune cell) condition at day 3 of culture, ****, p<0.0001.

Article Snippet: A commercially available human primary endometrial microvascular endothelial cell line (HEMEC, ScienCell, #7010) cultured in recommended endothelial cell medium (ECM, ScienCell, #1001) was used between P1-5.

Techniques: Ex Vivo, Purification, Flow Cytometry, Labeling, Imaging, Comparison, Control

Journal: bioRxiv

Article Title: Cell Chirality of Micropatterned Endometrial Microvascular Endothelial Cells

doi: 10.1101/2023.10.20.563368

Figure Lengend Snippet: (A) Representative image of actin immunofluorescent staining of human endometrial microvascular endothelial cells (HEMECs) on the microarray extracellular matrix (ECM) island. (B) Heat map of average percent clockwise fibers/island on every ECM combination. (C) Quantification of actin fiber orientation for specific ECM combinations. (D) Representative examples of percent clockwise cells. (E) Heat map of average percent clockwise cells/island on every ECM combination. (F) Quantification of cell-boundary based orientation on specific ECM combinations. (G) Representative Images for calculating nuclear-centrosome axis-based left-right cell orientation. (H) Heat map of average percent left-oriented cells/island on specific ECM combinations. (I) Quantification of left-right orientation on specific ECM combinations. Representative images of stained HEMECs cultured on (J) collagen 3 + laminin and (K) fibronectin + decorin. Scale bars: 100 μm.

Article Snippet: Human endometrial microvascular endothelial cells (HEMEC; ScienCell Catalog #7010) were maintained as per the manufacturer’s instructions.

Techniques: Staining, Microarray, Cell Culture