Session Type: ACR Poster Session A
Session Time: 9:00AM-11:00AM
Background/Purpose: Mesenchymal stem cells are multipotent stromal cells that can differentiate into tissue-specific lineages, such as osteocytes and chondrocytes. Their multipotency has made them an intriguing candidate for potential cellular therapy for rheumatic and musculoskeletal diseases; however, a substantial improvement in the understanding of the intracellular signaling, function, and regulation of mesenchymal stem cells is warranted before therapeutic use. Our data suggest G protein-coupled receptor kinase 3 (GRK3) functions as a negative regulator of G protein-coupled receptor (GPCR) signaling of sphingosine-1-phosphate receptor (S1PR) on bone marrow-derived mesenchymal stem cells (BmMSCs) and lack of such regulation alters BmMSC functionality.
Methods: BmMSCs were isolated from GRK3-deficient (Grk3-/-) and wildtype mice for evaluations of differentiation (chondrogenic, adipogenic, and osteogenic), CXCL12 secretion (ELISA), and proliferation (CCK8 assay). To better understand the functional differences noted with Grk3-/- BmMSCs, we investigated GPCR signaling through S1PR stimulation (ERK1/2 western blot). GRK3 β-arrestin recruitment to S1PR1 was demonstrated by TANGO, and S1PR1 internalization was determined by flow cytometry quantification of surface receptor expression.
Results: Grk3-/- BmMSCs have enhanced proliferation and osteogenic differentiation ex vivo compared to wildtype BmMSCs in identical culture conditions and passages. Grk3-/- BmMSCs also secrete more CXCL12, an essential chemokine for hematopoietic stem and progenitor development, and Grk3-/- mice have increased hematopoietic cell numbers isolated from the bone marrow. However, Grk3-/- mice do not show enhanced trabecular or cortical bone density, which suggest GRK3 deficiency in BmMSCs may have more pronounced effects on hematopoiesis as opposed to mature bone development in vivo. Both Grk3-/- BmMSC proliferation and osteogenic differentiation were reduced to wildtype level upon sphingosine kinase inhibitor treatment, and Grk3-/- BmMSCs have sustained ERK1/2 signaling upon stimulation of S1PR with S1P ligand. In addition, we report GRK3 recruits β-arrestin to the C-terminus of S1PR1, and we demonstrate BmMSCs lacking GRK3 regulation have impaired S1PR1 internalization.
Conclusion: Our work suggests GRK3 regulates S1PR on BmMSCs and lack of such regulation induces increased cellular proliferation and osteogenic differentiation, indicating GRK3 may serve as a potential therapeutic target to alter BmMSC functionality.
To cite this abstract in AMA style:Brozowski JM, Timoshchenko RG, Serafin DS, Koontz J, Allyn B, Eudy AM, Harris T, Abraham D, Rubin CT, Rubin J, Allbritton N, Billard MJ, Tarrant TK. G Protein-Coupled Receptor Kinase 3 Modulates Mesenchymal Stem Cell Proliferation and Differentiation through Sphingosine-1-Phosphate Receptor Regulation [abstract]. Arthritis Rheumatol. 2017; 69 (suppl 10). https://acrabstracts.org/abstract/g-protein-coupled-receptor-kinase-3-modulates-mesenchymal-stem-cell-proliferation-and-differentiation-through-sphingosine-1-phosphate-receptor-regulation/. Accessed February 22, 2020.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/g-protein-coupled-receptor-kinase-3-modulates-mesenchymal-stem-cell-proliferation-and-differentiation-through-sphingosine-1-phosphate-receptor-regulation/