Murine marrow‐derived mesenchymal stem cell: isolation, <i>in vitro</i> expansion, and characterization

TLDR

There is a lack of information about murine MSCs despite extensive study of human and other species MSCs. The study aims to establish long‑term in‑vitro expansion conditions for murine bone‑marrow MSCs and to characterize their morphology, surface markers, and growth kinetics, thereby enabling future therapeutic applications. The authors cultured plastic‑adherent murine BM cells for over 50 passages, assessing morphology, surface marker expression, and growth kinetics. The expanded cells differentiated into osteogenic and adipogenic lineages, supported hematopoietic stem cell growth, expressed CD44, CD49e, CD29, and Sca‑1, and were estimated to occur at a frequency of 1 per 11,300–27,000 nucleated cells in adult BALB/c and C57Bl/6 mice. Summary.

Abstract

Summary. In spite of the attention given to the study of mesenchymal stem cells (MSCs) derived from the bone marrow (BM) of humans and other species, there is a lack of information about murine MSCs. We describe the establishment of conditions for the in vitro expansion of plastic‐adherent cells from murine BM for over 50 passages, and provide their characterization regarding morphology, surface marker profile and growth kinetics. These cells were shown to differentiate along osteogenic and adipogenic pathways, and to support the growth and differentiation of haematopoietic stem cells, and were thus operationally defined as murine mesenchymal stem cells (mMSCs). mMSCs were positive for the surface markers CD44, CD49e, CD29 and Sca‐1, and exhibited a homogeneous, distinctive morphology. Their frequency in the BM of adult BALB/c and C57Bl/6 mice, normal or knockout for the α ‐L‐iduronidase ( IDUA ) gene, was preliminarily estimated to be 1 per 11 300–27 000 nucleated cells. The emergence of a defined methodology for the culture of mMSCs, as well as a comprehensive understanding of their biology, will make the development of cellular and genetic therapy protocols in murine models possible, and provide new perspectives in the field of adult stem cells research.