Brown adipose tissue (BAT) plays a key role in the evolutionarily conserved mechanisms underlying energy homeostasis in mammals. It is characterized by fat vacuoles 5-10 mm in diameter and expression of uncoupling protein one, central to the regulation of thermogenesis. In the human newborn, BAT depots are typically grouped around the vasculature and solid organs. These depots maintain body temperature during cold exposure by warming the blood before its distribution to the periphery. They also ensure an optimal temperature for biochemical reactions within solid organs. BAT had been thought to involute throughout childhood and adolescence. Recent studies, however, have confirmed the presence of active BAT in adult humans with depots residing in cervical, supraclavicular, mediastinal, paravertebral, and suprarenal regions. While human pluripotent stem cells have been differentiated into functional brown adipocytes in vitro and brown adipocyte progenitor cells have been identified in murine skeletal muscle and white adipose tissue, multipotent metabolically active BAT-derived stem cells from a single depot have not been identified in adult humans to date. Here, we demonstrate a clonogenic population of metabolically active BAT stem cells residing in adult humans that can: (a) be expanded in vitro; (b) exhibit multilineage differentiation potential; and (c) functionally differentiate into metabolically active brown adipocytes. Our study defines a new target stem cell population that can be activated to restore energy homeostasis in vivo for the treatment of obesity and related metabolic disorders. STEM CELLS 2014;32:572-581

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There have been various forms of mesenchymal stem cell-like (MSC-like) cells isolated from umbilical cords (UCs). The isolation of umbilical cord lining stem cells (ULSCs) may be of great value for those interested in a possible treatment to several disease/disorders. Unlike umbilical cord blood cells, these cells are unique because they can be expanded to therapeutically relevant numbers and cryopreserved for several different uses. Here we efficiently isolate stem cells from a small segment of pre- and postnatal UCs, and obtain therapeutically relevant amounts of ULSCs within 3 weeks. We demonstrate their growth potential and characterize them using immunocytochemistry, flow cytometry, and RT-PCR. In addition, we differentiate ULSCs into multiple lineages. Pre- and postnatal ULSCs are morphologically similar to mesenchymal stem cells (MSCs) and easily expand to greater than 70 population doublings. They express pluripotent markers Oct4 and nanog at the protein and RNA level. Flow cytometry demonstrates that they express markers indicative of MSCs in addition to high SSEA-4 expression. ULSCs are easily differentiated into osteogenic, adipogenic, chondrogenic, cardiogenic, and neurogenic cells. Pre- and postnatal ULSCs are characteristically similar in respect to their growth, marker expression, and plasticity, demonstrating they are highly conserved throughout development. ULSCs have phenotypic and genotypic properties of MSCs. These studies demonstrate the therapeutic potential of an otherwise discarded tissue. They are a perfect HLA match for the donor and an excellent match for immediate family members; therefore, they may serve as a therapeutic cell source.

BackgroundMesenchymal stem cells (MSCs) represent an attractive avenue for cellular therapies targeting degenerative diseases. MSC in vitro expansion is required in order to obtain therapeutic numbers during the manufacturing process. It is known that culture conditions impact cellular properties and behavior after in vivo transplantation. In this study, we aimed at evaluating the benefit of hypoxic culturing of human bone marrow derived mesenchymal stem cells on cell fitness and whole genome expression and discussed its implication on cellular therapies targeting orthopedic diseases such as chronic lower back pain.MethodsHuman bone marrow mesenchymal stem cells (hBMMSCs) were isolated from fresh human anticoagulated whole bone marrow and were cultured side by side in atmospheric (20% O2) and hypoxic (5% O2) oxygen partial pressure for up to 3 passages. Stem cell fitness was assessed by clonogenic assay, cell surface marker expression and differentiation potential. Whole genome expression was performed by mRNA sequencing. Data from clonogenic assays, cell surface marker by flow cytometry and gene expression by quantitative PCR were analyzed by two-tailed paired Student’s t-test. Data from mRNA sequencing were aligned to hg19 using Tophat-2.0.13 and analyzed using Cufflinks-2.1.1.ResultsHypoxic culturing of hBMMSCs had positive effects on cell fitness, as evidenced by an increased clonogenicity and improved differentiation potential towards adipocyte and chondrocyte lineages. No difference in osteoblast differentiation or in cell surface markers were observed. Only a small subset of genes (34) were identified by mRNA sequencing to be significantly dysregulated by hypoxia. When clustered by biological function, these genes were associated with chondrogenesis and cartilage metabolism, inflammation and immunomodulation, cellular survival, migration and proliferation, vasculogenesis and angiogenesis.ConclusionsHypoxic culturing positively impacted hBMMSCs fitness and transcriptome, potentially improving inherent properties of these cells that are critical for the development of successful cellular therapies. Hypoxic culturing should be considered for the in vitro expansion of hBMMSCs during manufacturing of cellular therapies targeting orthopedic disorders such as lower back pain.

The therapeutic use of stem cells to treat diseases and injuries is a promising tool in regenerative medicine. The umbilical cord provides a rich source of stem cells; we have previously reported a population of stem cells isolated from Wharton's jelly. In this report, we aimed to isolate a novel cell population that was different than those found in Wharton's jelly. We isolated stem cells from the subepithelial layer of the umbilical cord; the cells could be expanded for greater than 90 population doubling and had mesenchymal stem cell characteristics, expressing CD9, SSEA4, CD44, CD90, CD166, CD73, and CD146 but were negative for STRO-1. The cells can be directionally differentiated and undergo osteo-, chondro-, adipo-, and cardiogenesis. In addition, we have identified for the first time that mesenchymal stem cells isolated from umbilical cord can produce microvesicles, termed exosomes. This is the first report describing a stem cell population isolated from the subepithelial layer of the umbilical cord. Given the growth capacity, multilineage potential, and most importantly the low levels of HLA-ABC, we propose that this novel cell isolated from the subepithelial layer of umbilical cord is an ideal candidate for allogeneic cell-based therapy.