Researcher(s)
- Valerie West, Biomedical Engineering, University of Delaware
Faculty Mentor(s)
- Justin Parreno, Biological Sciences and Biomedical Engineering, University of Delaware
Abstract
Tendinosis as a result of tissue overload counterintuitively causes under stimulation of tendon cells, tenocytes. Previously in Parreno Lab, we demonstrated that cellular under stimulation alters cellular phenotype through a decrease in tenogenic expression levels and an increase in both chondrogenic and protease expression levels. Therefore, we hypothesize that this regulation of genes in response to cellular stress-deprivation is the result of actin depolymerization. Moreover, we speculate that actin depolymerization regulates genes through monomeric (globular; G-) actin-binding myocardin-related transcription factor (MRTF).
Leading up to this summer’s project, we began testing our hypothesis by exposing isolated primary tenocytes to Latrunculin A, sequestering G-actin and preventing actin polymerization, resulting in an increase in the proportion of G-actin to F-actin within the cells. Additionally, Latrunculin caused a nuclear export of MRTF from the nucleus of cells. Coinciding with nuclear export of MRTF are decreases to tenogenic genes (collagen-1, scleraxis, and a-smooth muscle actin) and increases to the expression of chondrogenic (Sox9) and proteases (Mmp-3 and Mmp-13). To determine if genes were regulated directly by MRTF, we exposed tenocytes to MRTF inhibitor, CCG1423. Exposure of cells to CCG1423 results in significant decreases to tenogenic genes, with minimal effects on chondrogenic or protease genes. Furthermore, we induce that actin depolymerization is a regulator of gene expression in tendon cells, partially through regulation of MRTF. To verify this, my summer research was centered around the measurement of protein levels in CCG1423 treated cells, through the WES protocol. col1a1 and alpha-smooth muscle actin were found to show decreased expression, but scleraxis proved unsuccessful using the WES. Ultimately, further understanding the regulation of gene expression during tendinosis by actin may lead to new therapeutic opportunities against disease progression.