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dc.contributor Crawford, Robert en_US
dc.contributor Fierro, Fernando en_US
dc.contributor.advisor Mulligan, Kimberly en_US
dc.contributor.author McEnerney, Amber
dc.date.accessioned 2018-05-16T18:20:08Z
dc.date.issued 2018-05-16
dc.date.submitted 2018-05
dc.identifier.uri http://hdl.handle.net/10211.3/202968
dc.description Project (M.A., Biological Sciences (Stem Cell))--California State University, Sacramento, 2018. en_US
dc.description.abstract Healthy bone relies on an equilibrium between bone-resorbing osteoclasts and bone-forming osteoblasts. Bone remodeling is therefore dependent on differentiation and activation of the associated osteoprogenitors. Discovering a way to supplement the bone marrow with osteoblast progenitor cells would be extremely beneficial for bone restoration in patients with brittle or degenerative bone disease. Recent fate-mapping studies have elucidated a niche residing in the metaphyseal plate of long bones, regulating the differentiation of osteoprogenitors. This niche cell-type is referred to as skeletal stem cells (SSCs). SSCs have the ability to differentiate into bone, cartilage and hematopoietic supportive stroma. However, human SSCs are understudied, likely due to the lack of an established, optimized sorting protocol enriching for this rare niche population. Based on recent work in mice, we have found that the combination of markers CD45- CD51+ CD146+ CD200lo enriches for a fraction of human cells capable of initiating colony forming units-fibroblast (CFU-F), a hallmark property of SSCs. Here we describe a novel isolation protocol for the putative human skeletal stem cell population. Before sorting, we obtained the mononuclear cell fraction by ficoll gradient centrifugation of the whole bone marrow biopsy. Our sorting strategy included magnetic columns to select the CD45ˉ population, followed by a CD146+ enrichment. Lastly, FACS sorting was used to obtain the CD45ˉCD51+CD200loCD146+ (SSC) population. The data includes isolated SSCs from seven different donors, ranging from 19 to 52 years old. We determined the frequency of SSCs, their ability to form colonies in a CFU-F assay, as well as their morphology and size. Since SSCs are present in very limited numbers in bone marrow, we also expanded the cells ex vivo. We found that freshly isolated SSCs displayed a greater expression of Mx1, Grem1 and Hmga2, when compared to the expanded SSC and Whole Bone Marrow Multipotent Stromal Cell (WBM-MSC) populations. After 14 days in culture, SSCs displayed many properties of WBM-MSCs, including fibroblastic morphology, immune phenotype, and gene expression. In addition, both cell populations successfully displayed osteogenic and adipogenic differentiation potential. Lastly, we show that freshly isolated human SSCs can efficiently home to bone marrow, opening the possibility of using these cells to treat systemic bone defects. This is a unique characteristic, as WBM-MSCs lack homing capabilities. With further investigation, human SSCs are likely to play a role in future regenerative therapeutics in orthopedic medicine. en_US
dc.description.sponsorship Biological Sciences (Stem Cell) en_US
dc.language.iso en_US en_US
dc.subject Osteoprogenitor en_US
dc.subject Bone marrow multipotent stromal cells en_US
dc.subject Osteoporosis en_US
dc.title Isolation and expansion of human skeletal stem cells en_US
dc.type Project en_US
dc.description.embargoterms 5 years en_US
dc.date.embargountil 2023-05-15T18:20:08Z

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