Gunpreet Oberoi1,2,3, Klara Janjić1,2, Anna Sonja Müller1,2, Barbara Schädl2,4,5, Andreas Moritz1,2, Hermann Agis6,7. 1. Department of Conservative Dentistry and Periodontology, University Clinic of Dentistry, Medical University of Vienna, Sensengasse 2a, 1090, Vienna, Austria. 2. Austrian Cluster for Tissue Regeneration, Vienna, Austria. 3. Center for Medical Physics and Biomedical Engineering, Medical University of Vienna, Vienna, Austria. 4. Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Vienna, Austria. 5. University Clinic of Dentistry, Medical University of Vienna, Vienna, Austria. 6. Department of Conservative Dentistry and Periodontology, University Clinic of Dentistry, Medical University of Vienna, Sensengasse 2a, 1090, Vienna, Austria. hermann.agis@meduniwien.ac.at. 7. Austrian Cluster for Tissue Regeneration, Vienna, Austria. hermann.agis@meduniwien.ac.at.
Abstract
OBJECTIVES: The factors that contribute to the morphological changes of dental pulp cell-derived microtissues are unknown. Here, we investigated the contraction dynamics of rod-shaped microtissues derived from dental pulp cells and examined the underlying cell signaling pathways. METHODS: Human dental pulp cells were seeded into agarose molds to assemble into rod-shaped microtissues. Resazurin- and tetrazolium-based cytotoxicity assays, Live/Dead staining, and hematoxylin and eosin staining for histological evaluation of rods were performed. Rod contraction was evaluated and measured for a period of 10 days. The role of TGF-β, phosphoinositide 3-kinase (PI3K)/AKT, and mitogen-activated protein kinase (MAPK) signaling pathway was analyzed. RESULTS: Dental pulp cells readily assembled into rods, maintaining the geometric shape for 48 h. Following this period, they condensed to form stable spheroidal structures that remained vital for 10 days from seeding. Inhibition of phosphoinositide 3-kinase signaling pathway by LY294002 significantly prolonged the diminution in the length of rods formed by dental pulp cells. TGF-β and pharmacological inhibition of TGF-β signaling did not show pronounced effects. CONCLUSION: Overall, dental pulp cells readily formed rod-shaped patterns of microtissues which, over a period of time, condensed into more stable spheroidal structures. Hence, technologies like bioprinting, using direct fabrication of microtissues need to consider the contraction dynamics. CLINICAL RELEVANCE: The field of regenerative endodontology will benefit from our findings as it can be applied as a novel platform to test the impact of pharmacological agents, biomaterials, and regenerative approaches including bioprinting.
OBJECTIVES: The factors that contribute to the morphological changes of dental pulp cell-derived microtissues are unknown. Here, we investigated the contraction dynamics of rod-shaped microtissues derived from dental pulp cells and examined the underlying cell signaling pathways. METHODS:Human dental pulp cells were seeded into agarose molds to assemble into rod-shaped microtissues. Resazurin- and tetrazolium-based cytotoxicity assays, Live/Dead staining, and hematoxylin and eosin staining for histological evaluation of rods were performed. Rod contraction was evaluated and measured for a period of 10 days. The role of TGF-β, phosphoinositide 3-kinase (PI3K)/AKT, and mitogen-activated protein kinase (MAPK) signaling pathway was analyzed. RESULTS: Dental pulp cells readily assembled into rods, maintaining the geometric shape for 48 h. Following this period, they condensed to form stable spheroidal structures that remained vital for 10 days from seeding. Inhibition of phosphoinositide 3-kinase signaling pathway by LY294002 significantly prolonged the diminution in the length of rods formed by dental pulp cells. TGF-β and pharmacological inhibition of TGF-β signaling did not show pronounced effects. CONCLUSION: Overall, dental pulp cells readily formed rod-shaped patterns of microtissues which, over a period of time, condensed into more stable spheroidal structures. Hence, technologies like bioprinting, using direct fabrication of microtissues need to consider the contraction dynamics. CLINICAL RELEVANCE: The field of regenerative endodontology will benefit from our findings as it can be applied as a novel platform to test the impact of pharmacological agents, biomaterials, and regenerative approaches including bioprinting.
Entities:
Keywords:
Contraction dynamics; PI3K; Pulp regeneration; Rod microtissues; TGF-β