Lo Surdo, Jessica
(2013)
Toward Improved Characterization of Human Mesenchymal Stem Cells for Use in Cellular Therapies: A Regulatory Science Perspective.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
Cellular therapies hold great potential to treat a variety of medical conditions. Product characterization of cellular therapies is particularly difficult, as they pose regulatory challenges due to donor heterogeneity and a lack of standard lot release tests that can reliably predict in vivo outcomes. In particular, multipotent stromal cells, also called mesenchymal stem cells (MSCs), are potentially valuable as a cellular therapy due to their regenerative capacity and immunosuppressive function. Due to the required expansion and inherent heterogeneity of MSCs, quantitative approaches capable of measuring differentiation capacity and immunosuppressive function between donors and passages on a per cell basis are needed. To address this unmet need, a sample set of human MSCs comprised of eight donors was created, cultured to early and late passages, and novel quantitative bioassays were established capable of measuring adipogenic and osteogenic differentiation on a per cell basis, as well as an in vitro assay to measure immunosuppressive function. Based on existing bioassays, MSCs demonstrate a decrease in overall proliferative potential and colony forming unit capacity, while expression of hallmark MSC surface markers remain unchanged. Utilizing automated microscopy techniques, adipogenic and osteogenic differentiation potential was quantified on a per cell basis, allowing us to directly assess the role of donor variability and in vitro culture on MSC function. Overall, donor variability and a decrease in differentiation potential with passage was demonstrated based on these quantitative assays. Since culture-expanded MSCs increase in cell size, this parameter was utilized to enrich for small cells, which demonstrated that the small cell population is more stem-like based on these applied quantitative bioassays. Lastly, immunosuppressive function of human MSCs on murine-derived clonal T cells was assessed utilizing a novel in vitro xenogeneic model system. Human MSCs can inhibit murine T cell activation, rendering this an ideal system to assess immunosuppressive function of MSCs in vitro. In conclusion, novel methods were established to quantify MSC function, and these findings were correlated with other previously-established quantitative bioassays to better understand the role of donor variability and passaging on MSC potency. Taken together, these quantitative approaches provide valuable tools to measure MSC quality, and supports continued efforts to improve characterization strategies for cellular therapies.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
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| ETD Committee: |
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| Date: |
25 September 2013 |
| Date Type: |
Publication |
| Defense Date: |
26 April 2013 |
| Approval Date: |
25 September 2013 |
| Submission Date: |
10 July 2013 |
| Access Restriction: |
5 year -- Restrict access to University of Pittsburgh for a period of 5 years. |
| Number of Pages: |
172 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Swanson School of Engineering > Bioengineering |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
mesenchymal stem cells, differentiation, characterization |
| Date Deposited: |
25 Sep 2013 12:54 |
| Last Modified: |
25 Sep 2018 05:15 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/19298 |
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