Session Type: ACR Poster Session C
Session Time: 9:00AM-11:00AM
Background/Purpose: Ankylosing spondylitis (AS) is an immune-mediated inflammatory disease associated with trabecular bone loss and concomitant aberrant bone formation in the spine. Over 40 genes or genetic regions have been associated with the risk of developing AS. The impact of these genes on disease pathogenesis and the cell types that mediate these effects remain largely unknown. To begin to address these questions we generated induced pluripotent stem cells (iPSCs) from fibroblasts of axial spondyloarthritis (AxSpA) patients and healthy controls (HC), differentiated them into disease-relevant cell lineages like mesenchymal stem cells (MSCs) and osteoblasts (OBs), and analyzed comprehensive gene expression patterns and expression levels of AS risk genes in iPSCs and MSCs using whole blood as a comparison.
Methods: Dermal fibroblasts from two AxSpA patients and one HC were reprogrammed using a Sendai virus vector encoding OCT4, SOX2, KLF4 and MYC. Virus-free iPSCs were differentiated into MSCs using a TGF-β inhibitor. MSCs were differentiated into osteoblasts, by culturing them in cell type specific differentiation media. Immunofluorescence microscopy and flow cytometry were conducted to evaluate the expression of lineage specific markers. Genome-wide gene expression patterns in iPSCs and MSCs were compared with whole blood using RNA-Seq followed by Principle Components Analysis (PCA) and Ingenuity Pathway Analysis (IPA).
Results: Reprogrammed lines expressed iPSC-specific genes and proteins, and iPSCs were successfully differentiated into functional MSCs. Gene expression analysis revealed distinct cell type-specific clusters and differentially expressed pathways identified by IPA. The expression of known AS risk genes in iPSCs and MSCs was compared to peripheral blood, revealing several genes that were highly enriched in MSCs (e.g. EDIL3, ANO6, HAPLN1) or expressed at high levels, but comparable to blood (TNFRSF1, TYK2, SH2B3, and STAT3). MSCs were further differentiated into osteoblasts. Differences in osteoblast mineralization potential between subject-derived lines were consistent in duplicate iPSC derivations, with the AxSpA patient exhibiting 3-fold higher mineralization than the HC along with increased expression of alkaline phosphatase (ALP) mRNA in patient lines, whereas other genes associated with osteoblast differentiation (etc. RUNX2, Osterix) were expressed equally in all lines.
Conclusion: AxSpA patient-derived iPSCs demonstrated pluripotency, lineage-specific gene expression patterns, and could be differentiated into disease-relevant cell types. A subset of risk genes was expressed predominantly in MSCs, underscoring the potential utility of these cells in studying pathogenic mechanisms. Moreover, enhanced OB mineralization potential was reproducible in separate iPSC derivations from the same individuals. We are currently generating more lines to determine whether this is a disease characteristic. In summary, skin fibroblast-derived iPSCs provide a powerful system to explore the functional genomics of AS risk genes that may impact bone formation.
To cite this abstract in AMA style:Layh-Schmitt G, Lu S, Lazowick E, Brooks S, Gadina M, Colbert RA. Characterization of Mesenchymal Stem Cells Generated from Axial Spondyloarthritis Patient-Derived Induced Pluripotent Stem Cells [abstract]. Arthritis Rheumatol. 2015; 67 (suppl 10). https://acrabstracts.org/abstract/characterization-of-mesenchymal-stem-cells-generated-from-axial-spondyloarthritis-patient-derived-induced-pluripotent-stem-cells/. Accessed October 16, 2021.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/characterization-of-mesenchymal-stem-cells-generated-from-axial-spondyloarthritis-patient-derived-induced-pluripotent-stem-cells/