Cd73 561014

Immunophenotyping of eMSCs [cluster of differentiation (CD) marker expression] was performed using an Epics XL flow cytometer (Beckman Coulter, Inc., Brea, CA, USA). A single cell suspension was obtained using 0.05% trypsin/EDTA. Cells (1×10⁶) were suspended in 1 ml PBS containing 5% FCS. Subsequently, according to the manufacturer's instructions, the cells were incubated at room temperature for 30 min with the following mouse anti-human fluorescein isothiocyanate-conjugated monoclonal antibodies: anti-CD34 (555821; 1:250), CD45 (555482; 1:250) and CD90 (555595; 1:250), and phycoerythrin-conjugated monoclonal antibodies: Anti-CD19 (557835; 1:250), CD73 (561014; 1:200), CD105 (560839; 1:250), CD146 (561013; 1:200) and human leukocyte antigen (HLA)-DR (555812; 1:200; all BD Pharmingen, San Diego, CA, USA). All antibodies were pre-diluted for use according to the recommended volume per test. For each test, 1×10⁶ cells in 100 µl PBS were used. For CD73, CD146 and (HLA)-DR the volume per test was 20 µl. Cells were analyzed by flow cytometry.

Recombinant human BMP2 was provided by Daewoong Pharmaceutical Co., Ltd. (Seoul, Korea). Primary Abs against β-actin (#3700), p-STAT1 (Ser701) (#7649s), p-STAT1 (Tyr727) (#9177s), total STAT1 (#9172s) were purchased from Cell Signaling Technology Inc. (Beverly, MA, USA), and inducible nitric oxide synthase (iNOS, sc-650) and IDO (sc-53978) were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). Secondary Abs were purchased from Invitrogen (Carlsbad, CA, USA; mouse: 31430; rabbit: 32460). FACS Abs against HLA-DR (307603). CD4 (100411), CD25 (101907), and FOXP3 (320007) were purchased from BioLegend, Inc. (San Diego, CA, USA), and CD105 (560839), CD45 (560973), CD34 (560940), and CD73 (561014) were purchased from BD Biosciences (San Jose, CA, USA).