Cd11b c

Expression of CD90, CD105, CD45, and CD11b/c (eBioscience, USA) on MSCs surface was assessed by flow cytometry (Becton Dickinson, USA). The expression of CD11b/c, CD80, CD83, CD86, and MHC II (eBioscience, USA) on DCs surface was identified using flow cytometry. Each group of DCs were collected on the sixth day. After washing with PBS, the cells were immunolabeled with monoclonal anti-CD80, anti-CD83, anti-CD86, and anti-MHC II (eBioscience, USA) antibodies, as well as their isotype control antibodies. After that they were incubated in darkness at 4°C for 30 min and were detected using FACSCalibur flow cytometer.

All of the experiments and surgical procedures were performed in accordance with the ethical guidelines issued by the ISO 10993–1:2009 and approved by the local Bioethics Committees of Faculty of Biology, M.V. Lomonosov Moscow State University (#16.1 dated 28 May 2021). Mesenchymal stem cells (MSCs) were isolated from femurs of young (3–5 days old) Wistar rats and cultured for 2 weeks in DMEM (Dubecco’s Modified Eagle Medium, PanEco, Russia), which was supplemented with 10% fetal calf serum (FCS, Biological Industries, Israel) and 100 U/mL penicillin. MSCs were cultivated for three passages. The cells were removed via incubation in a trypsin–versene solution for 5 min and then counted using hemocytometer. Later on, a 10⁵ cell suspension in 100 μL of PE buffer (2 mL EDTA-0.5% ETS in PBS) was prepared and incubated with antibodies to positive surface markers of MSC phenotype: CD90 and CD29; negative surface markers: CD45 and CD11b/c (eBioscience, San Diego, CA, USA); the dye cell viability 7-Aminoactinomycin D (7AAD) in the dark for 40 min at a temperature of 5 °C. Cells were then washed in PBS once and extracted via centrifugation, which was followed by a flow cytometer examination (FACS ARIA II, Franklin Lakes, NJ, USA). Analysis of the results and graphs were performed using the Flowing Software 2.5.1 (Supplemental Materials, Figure S2).