Fig. 5: In vitro cleavage of cellular Endoglin by thrombin.

a Immunofluorescence staining of Endoglin was conducted using the Endoglin/CD105 P4A4-Alexa 488 antibody (green) and vectashield mounting medium with DAPI (blue) after treating ECFC with increasing concentrations of Thr-H (0.01, 0.1 and 1 µM). Observations were carried out using confocal microscopy at 20x magnification. Phase photos at 10x magnification were taken for each condition to confirm the presence of the cell monolayer. b Endoglin fluorescence intensity following ECFC treatment with increasing concentrations of Thr-H (0.01, 0.1 and 1 µM) was quantified using ImageJ software (n = 3). c Plots showing the quantification of Eng fluorescence intensity in ECFC following treatment with 1 µM of Thr-H, alongside the number of nuclei relative to the surface. d Immunofluorescence staining of Eng using the Endoglin/CD105 P4A4-Alexa 488 antibody in different cell types, including HUVECs and MSCs, mounted with Vectashield containing DAPI. HUVECs were additionally stained with VE-Cadherin (VE-CAD) using a Texas Red-conjugated antibody, and MSCs were stained with Phalloidin-Alexa 555 to assess cell integrity after treatment with Thr. e Plots showing the quantification of Eng fluorescence intensity in HUVEC and MSC following treatment with 1 µM of Thr-H, alongside the number of nuclei relative to the surface. f ELISA kit assay results for sEng in supernatants of ECFC, HUVEC and MSC treated or not with 1 µM of Thr-H. The data is presented as mean ± S.D. for a minimum of n = 4. Statistical significance was considered at *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.