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Wensi Tao, Juan Ayala-Haedo, Matthew Field, Daniel Pelaez, Sara Wester; RNA-Sequencing Gene Expression Profiling of Orbital Adipose Derived Stem Cell Population of Thyroid Associated Orbitopathy (TAO). Invest. Ophthalmol. Vis. Sci. 2017;58(8):5154.
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© ARVO (1962-2015); The Authors (2016-present)
To characterize the cellular differences in gene expression profiling of orbital adipose-derived stem cells (OASC) from normal controls and patients with thyroid associated orbitopathy (TAO).
Samples were collected from medial orbital fat pad from controls and TAO patients undergoing orbital decompression. A total of 9 patients were recruited, 5 patients with active TAO and 4 controls without history of thyroid dysfunction. OASC were isolated by enzymatic digestion followed by centrifugation and clonal expansion.OASCs were differentiated into three lineages: chondrocytes, osteocytes, and adipocytes.RNA sequencing analysis was performed and results were compared to assess for differences in gene expression. FDR of 0.05 and 1.5 fold change in gene expression were used for cut off filtering. The top hits were subsequently confirmed by RT-PCR.
Isolated OASC express high levels of mesenchymal stem cell markers (CD 90, Nestin, CD146, and PDGFR), but low levels of the pluripotent stem cell markers (KLF4, OCT4, Nanog, Sox2). OASC isolated from TAO patients exhibited an increase in adipogenesic potential, and a decreased chondrogenesic and osteogenesic induction. RNA-seq disclosed 26 differentially expressed genes when comparing TAO and controls, including genes in the HOX family, Wnt signaling pathways and Zic family proteins. 3 up-regulated genes DSC3B, HOXB2 and KCNA4 and 2 down-regulated genes ZIC4 and WNT16 were confirmed by RT-PCR. We found an upregulation of HOX genes and down-regulation of WNT singling pathway in the OASC from TAO patients compared with control.
Our results demonstrate that there are intrinsic genetic and cellular differences in the OASC populations derived from TAO patients and controls. Since restricted expression patterning HOX genes imply their specific regional functions, the upregulation of HOX gens in the OACS from active TAO patients may be related to developmental abnormality in the orbital adipose tissue related to TAO. Meanwhile, Wnt signaling pathway is known to be a negative regulator for adipogenesis, down-regulation of WNT singling pathway can lead to an enhanced adipocyte differentiation in TAO patients. These changes in genetic expression profile of these two cell populations might be implicated in the pathogenesis of thyroid eye disease.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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