Healthy human primary bm mscs

Three month old Lewis rats were euthanized in accordance with approvals from the Institutional Animal Care and Use Committee (IACUC) of Rhode Island Hospital. Menisci were isolated using sterile tools, rinsed with sterile PBS, and cultured in DMEM++ in a 37°C cell incubator for 24–48 hours prior to experiments. For ex-plant repair experiments, at least three menisci were used per experimental group. A radial tear was created in the inner anterior horn and each menisci was placed into a 96-well culture plate. Torn menisci were treated by co-culturing with 1.0×10⁵ healthy human primary BM-MSCs (ATCC, Manassas, VA), CPCL3, primary C-PCs, or no cell (control group). All cells were fluorescently labeled with Vibrant Dil Cell Labeling Solution (ThermoFisher Scientific, Waltham, MA, USA) according to the manufacturer. In experiments involving SDF-1, menisci were pre-treated with SDF-1 recombinant protein (200ng/mL) (PeproTech Inc., Rocky Hill, NJ) for 15 minutes followed by treatment with 1.0×10⁵ fluorescently labeled CPCL3 cells. Images were taken 3, 5, 10, 17 and 20 days after treatment using a Nikon Eclipse 90i microscope. Gene expression analysis was conducted using mRNA collected from 20 day samples.