Preferred Name
Nikki Cubbage
Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-No Derivative Works 4.0 International License.
Date of Graduation
5-9-2024
Semester of Graduation
Spring
Degree Name
Master of Science (MS)
Department
Department of Biology
Second Advisor
Timothy Bloss
Third Advisor
Kristopher Kubow
Abstract
Following injury, tissue repair allows for the restoration of the architecture and function of the injured tissue, while tissue replacement is characterized by severely damaged or non-regenerable tissue that is repaired by the laying down of extensive connective tissue, also known as fibrosis. The fibrotic process should be arrested at its early stage to have the greatest impact on preventing or reversing associate sequelae and promoting the regeneration of normal tissue. Sphingosine 1-phosphate (S1P), a metabolic product of cell membrane sphingolipids, has emerged as a pathway that may be involved in fibrosis. Most of S1P’s actions are mediated through the five G protein-coupled S1P receptors (S1PR 1–5) that respond to extracellular S1P to regulate multiple cellular processes. Here we used 2-dimensional (2D) and 3-dimensional (3D) in vitro cell culture models to characterize S1P in injury and post-traumatic fibrosis. We hypothesized that the S1P-S1PR axis plays a role in the formation of fibrosis after injury. To test our hypothesis the in vitro models used human bone-marrow mesenchymal stem cells (BM-MSCs) treated with transforming growth factor beta-1 (TGFβ1) to mimic injury (2D and 3D culture systems) or to induce the formation of fibrotic nodules (3D culture system). Quantitative PCR (qPCR) was used to investigate the expression levels of S1PRs, S1P pathway genes, and fibrotic genes, using GAPDH as a housekeeping gene. Overall, treatment of TGF��1 for 24 hours was significantly associated with all the genes investigated in most of the in vitro models. Upregulation of S1PR4 and S1PR5 was observed in the 2D injury model as well as upregulation of the collagen type 1 alpha 1 chain (COL1A1) gene in the 3D fibrotic nodule assay, which may suggest that the fibrotic nodule assay is a model of a later – more established – stage of the fibrotic process in comparison with the 2D injury model. Altogether, our results provide novel insights in the modulation of the expression of S1P during human post-traumatic fibrotic formation and support the notion that the S1P pathway may be a potential target in the prevention and/or treatment of fibrotic formation after injury.
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Biology Commons, Cell and Developmental Biology Commons, Medical Cell Biology Commons, Molecular Genetics Commons, Musculoskeletal Diseases Commons
