IADR Abstract Archives
Photobiomodulation Activated Odontoblast Dynamics: a Function of Matrix Mechanical Properties
Objectives: The predominant focus of regenerative dentistry has been genetic manipulations, yet the epigenetic role of extracellular matrix (ECM) mechanics is poorly investigated. This project aims to dissect the cellular responses of odontoblasts to matrix mechanics by replicating a wound-like soft environment and assessing the role of Photobiomodulation (PBM) treatments for dentin regeneration.
Methods: Polydimethylsiloxane (PDMS, Sylgard 184) matrices, composed of a 10:1 ratio (base: curing agent) were poured into 12-well polystyrene plates and cured for 24 hours in a 95°C oven. Mechanical stiffness was evaluated using a Shore A Durometer (Insize) and surface topology was analyzed with an Atomic Force Microscope (Bruker). Following sterilization and serum coating, an odontoblast cell line was seeded (100,000 cells/well) in hypoxic (1 µg/mL Cobalt chloride) and low serum (0.2% FBS) conditions to simulate wound-like environments. Cell adhesion assay was performed at 5 hours to examine cell-substrate interaction and cell viability was assessed at 24 hours with AlamarBlue assay. PBM was performed (810 nm laser, CW, 10 mW/cm2, 5 min; 4.5 p.J/cm2, 1 Einstein). To examine the mechanistic pathways involved, Seahorse analysis and bulk RNA sequencing were performed (Illumina NextSeq 550 system).
Results: The PDMS substrate exhibited a lower shore stiffness (1.1 ± 1.6 MPa) than polystyrene (~1 GPa). Reduced cell viability and proliferation were observed on the softer matrix (n = 3, p < 0.05), which were rescued by PBM treatment, while cell adhesion remained unaffected. Notably, TGF-β1 and FAK inhibition enhanced cell viability, and non-uniform morphology was observed, whereas SB-431542 had no such effect. RNA sequencing demonstrated activation of cell survival and mitochondrial signaling pathways via PBM treatment. Future studies utilizing Seahorse analysis and qPCR will further elucidate the underlying molecular mechanisms.
Conclusions: The results from this study suggest that precision engineering of biomaterial mechanical properties can promote wound healing.
Methods: Polydimethylsiloxane (PDMS, Sylgard 184) matrices, composed of a 10:1 ratio (base: curing agent) were poured into 12-well polystyrene plates and cured for 24 hours in a 95°C oven. Mechanical stiffness was evaluated using a Shore A Durometer (Insize) and surface topology was analyzed with an Atomic Force Microscope (Bruker). Following sterilization and serum coating, an odontoblast cell line was seeded (100,000 cells/well) in hypoxic (1 µg/mL Cobalt chloride) and low serum (0.2% FBS) conditions to simulate wound-like environments. Cell adhesion assay was performed at 5 hours to examine cell-substrate interaction and cell viability was assessed at 24 hours with AlamarBlue assay. PBM was performed (810 nm laser, CW, 10 mW/cm2, 5 min; 4.5 p.J/cm2, 1 Einstein). To examine the mechanistic pathways involved, Seahorse analysis and bulk RNA sequencing were performed (Illumina NextSeq 550 system).
Results: The PDMS substrate exhibited a lower shore stiffness (1.1 ± 1.6 MPa) than polystyrene (~1 GPa). Reduced cell viability and proliferation were observed on the softer matrix (n = 3, p < 0.05), which were rescued by PBM treatment, while cell adhesion remained unaffected. Notably, TGF-β1 and FAK inhibition enhanced cell viability, and non-uniform morphology was observed, whereas SB-431542 had no such effect. RNA sequencing demonstrated activation of cell survival and mitochondrial signaling pathways via PBM treatment. Future studies utilizing Seahorse analysis and qPCR will further elucidate the underlying molecular mechanisms.
Conclusions: The results from this study suggest that precision engineering of biomaterial mechanical properties can promote wound healing.
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