(355d) Assessing Human Articular Cartilage Transcriptome Layers with RNA Sequencing

Osteoarthritis, a chronic disease, remains an issue for adults that causes cartilage degradation within a joint . According to the Centers for Disease Control and Prevention (2023), over 32.5 million adults in the US are affected by osteoarthritis (OA) [1]. In this study we seek to understand the connection between tissue engineering and genetics to regenerate human articular cartilage (hAC). We purpose to validate a protocol for RNA isolation and characterize the transcriptome of hAC in a tri-layer fashion via bulk RNA sequencing (bulk-RNA-seq). Additionally, we aim to analyze the transcriptome of normal articular cartilage in comparison to the hAC chemical composition and physical properties. We are relating these properties to the tri-layers of hAC through histological staining with Safranin O—Fast green and imaging with differential interference contrast (DIC) microscopy. We are relating these properties to superficial, middle, and deep zone with a cryotome procedure, RNA extracted, and qualifiedd by Bioanalyzer. Next, we generate bulk RNA sequencing of hAC layer-by-layer and compare results to early passaging of Mesenchymal Stromal Cells (MSC) and tissues intended for Matrix-Induced Autologous Chondrocyte Implantation (MACI). We will use differential gene expression (DE) analysis by DESeq2 R package software for bulk-RNA-seq. The result will be interpreted in terms of differentiation from MSCs to gene expression patterns of tri-layer hAC. We will report on development and validation of protocols for isolating cells and their subsequent characterization with application in regenerating the tri-layered hAC transcriptome stimulatory bioreactors used in our laboratory and corresponding properties of the extracellular matrix (ECM).

[1] “Osteoarthritis | CDC.” Accessed: Apr. 05, 2024. [Online]. Available: https://www.cdc.gov/arthritis/types/osteoarthritis.htm