Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Isolation and Characterization of a Novel Population of Pluripotent Stem Cells Derived from Human Adipose Tissue

ABSTRACT: Transcriptional comparison of two subpopulations of stem cells isolated from lipoaspirated human adipose tissue, profiling Adipose Stem Cells (ASCs) and Multilineage Differentiating Stress-Enduring Adipose Tissue cells (MUSE-ATs). Experiments focused in the isolation and characterization of a new population of adipose tissue (AT) derived pluripotent stem cells, termed MUSE-ATs, which are isolated using severe cellular stress conditions, including long-term exposure to the proteolytic enzyme collagenase, serum deprivation, low temperatures and hypoxia. Under these conditions, a homogenous, highly purified population of Muse-AT cells is isolated without the utilization of cell sorting methods. The goal of this experiment was to characterize MUSE-ATs, as well as compare them to the endogenous subpopulation of ASCs for reference. Single condition experiment, MUSE-ATs vs. ASCs. Biological replicates: 3 MUSE-ATs replicates, 3 ASCs replicates for reference.

ORGANISM(S): Homo sapiens

SUBMITTER: Saleh Heneidi

PROVIDER: E-GEOD-46353 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Adipose tissue is a plentiful and easily accessible source of mesenchymal stem cells that have been shown to be multi potent and possibly have immuno suppressive capacity(Zuk, Zhu et al. 2002; McIntosh, Zvonic et al. 2006; Prichard, Reichert et al. 2008). Several studies have identified hypoxia and the molecular effector of reduced oxygen tension, HIF1Î± to be essential in the differentiation processes, stem cell proliferation and survival as well as in maintenance of stem cell characteristics (Lennon, Edmison et al. 2001; Goda, Ryan et al. 2003; Wang, Fermor et al. 2005; Lin, Lee et al. 2006; Malladi, Xu et al. 2007; Ohnishi, Yasuda et al. 2007). The fact that oxygen has a central role in cell development and metabolism makes the regulation of oxygen tension a valuable tool in the attempt of exploiting stem cells to their full potential. In the current study we investigate the transcriptional changes caused by two weeks of monolayer hypoxic expansion of human adipose tissue derived stem cells (ASCs). Application of the XVivo hypoxic workstation (BioSpherix, Redfield, NY) made it possible to conduct the expansion of six ASC lines at four different hypoxic conditions in parallel covering the range from mild (15% oxygen) to full hypoxic (1% oxygen) conditions. In this comprehensive investigation the Illumina bead microarray platform was employed to give a profound and new insight in the molecular changes effectuated by hypoxic expansion. The findings of this study demonstrate the significance of biologic variation in the transcriptional response to mild hypoxic expansion and display evidence that full hypoxic growth conditions at 1% oxygen tension selects for a more prolific cell population of reduced chondrogenic potential. Keywords: hypoxia response To investigate the transcriptional response to hypoxia, adipose tissue derived stem cells (ASCs) from six female individuals were cultured for 14 days at different oxygen tensions (ambient, 15%, 10%, 5% and 1%). Gene expression levels at each oxygen tension were compared to ambient conditions (Ambient oxygen tension) using Illumina Human-6 v2.0 microarrays.

Project description:The treatment of bone defects caused by infection, trauma or neoplasms remains a clinical challenge. Autologous bone transplantation is limited by availability, donor site morbidity and surgical risk factors. This has given rise to stromal/stem-cell based therapy. Bone marrow derived stromal cells (BMSCs) have been studied to a large extent and show high regenerative potential but their use is limited by availability, donor site morbidity and the relatively low cell yield as they represent only <0.1% of cell harvested from bone marrow aspirate. At the same time, they are the closest mesenchymal stromal cells for bone tissue engineering given their tissue origin and, unlike other mesenchymal stromal cells, can support the formation of hematopoietic marrow. Adipose tissue derived stromal cells (ASCs) as part of the stromal vascular fraction of adipose tissue can as well undergo osteogenic differentiation but can be additionally isolated in a sufficient quantity from lipoaspirate after liposuction of abundant subcutaneous fat tissue. Here, it has been shown that there are no major differences in regard to proliferation or differentiation capacity of ASCs derived from subcutaneous fat of different anatomical regions. It has been shown that BMSCs are more prone to senescence during expansion and passage than ASCs and that ageing impacts proliferative capabilities of BMSCs more than that of ASCs while it has also been reported that osteogenic differentiation capacity is least impacted by age. Multiple studies have compared the characteristics of these two mesenchymal stromal cells in regard to bone tissue engineering in vitro. Most studies point to inferior extracellular matrix mineralization and lower expression of key osteogenic transcription markers like Runx2 in osteogenic differentiated ASCs compared to BMSCs. On the other hand, a study by Rath et al. found contrary results using particular culturing conditions like 3D bioglass scaffolds. An intraindividual comparison of human MSCs of three donors cultured on decellularized porcine bone confirmed superior osteogenic capacity of BMSCs compared to ASCs. In contrast to BMSCs, ASCs were not able to induce heterogenic ossification in a mouse model. In a sheep tibia defect model application of BMSCs resulted in a significantly higher amount of newly formed bone tissue. Importantly, Osteogenic differentiated ASCs do not support the formation of a hematopoietic marrow. Proteomics enables large-scale analysis of proteins present in a cell type and can be used to identify differentially regulated key proteins in a comparative approach. A comparative proteomic analysis of BMSCs and ASCs by Roche et al. in 2009 identified 556 proteins with 78% of these not being differentially regulated between these two cell populations, regarded as high similarity. Another comparative proteomic study of 2016 by Jeon et al. found 90 differentially regulated proteins out of 3000 total identified proteins. Both studies do not specify a number of different tissue donors and in part using cell lines. Looking for differences upon osteogenic differentiation, transcriptomic comparison of osteogenic differentiated porcine ASCs and BMSCs has been performed, resulting in 21 differentially expressed genes after 21 days of osteogenic culture conditions. Still, it remains unanswered, which are the key distinctive features of osteogenic differentiated ASCs and BMSCs at protein level that might help address the abovementioned weaknesses of ASCs in bone tissue engineering/regeneration for translational research. To overcome this need, an intraindividual comparative DIA based proteomic analysis of osteogenic differentiated human BMSC and ASCs was performed in this study.

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Isolation and Characterization of a Novel Population of Pluripotent Stem Cells Derived from Human Adipose Tissue

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