Protocol for isolation of human bone marrow stromal cells and characterization of cellular metabolism

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Bone marrow stromal cells (BMSCs) serve as a valuable reservoir of multipotent stem cells important in the regulation of bone homeostasis and energy metabolism. Here, we present a protocol for isolating human BMSCs (hBMSCs) and characterizing their cellular metabolism related to hBMSC functional properties. We describe steps for bioenergetics, cell senescence, and production of reactive oxygen species (ROS), together with description of the data analysis. These assays provide information on hBMSC metabolic status valuable to regenerative medicine and therapeutic applications. For complete details on the use and execution of this protocol, please refer to Tencerova et al. 1 • Isolation and cultivation of hBMSCs from bone marrow aspirates • Steps to measure hBMSC bioenergetics, ROS production, and senescent phenotype • Data analysis of metabolic assays and troubleshooting Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Bone marrow stromal cells (BMSCs) serve as a valuable reservoir of multipotent stem cells important in the regulation of bone homeostasis and energy metabolism. Here, we present a protocol for isolating human BMSCs (hBMSCs) and characterizing their cellular metabolism related to hBMSC functional properties. We describe steps for bioenergetics, cell senescence, and production of reactive oxygen species (ROS), together with description of the data analysis. These assays provide information on hBMSC metabolic status valuable to regenerative medicine and therapeutic applications.

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