Document Type

Thesis

Degree Name

Master of Science (MSc)

Department

Health Science

Faculty/School

Faculty of Science

First Advisor

Stephanie DeWitte-Orr

Advisor Role

Supervisor

Second Advisor

Diane Gregory

Advisor Role

Co-Supervisor

Abstract

Low back pain (LBP) is a leading cause of disability worldwide and is commonly associated with intervertebral disc (IVD) degeneration. During IVD degeneration (IVDD), extracellular matrix fragments, including decorin, are released and can function as damage-associated molecular patterns (DAMPs), activating innate immune responses through pattern recognition receptors (PRRs). Pathogen-associated molecular patterns (PAMPs), such as long double-stranded RNA (LdsRNA), also activate innate immune signaling and may additionally induce the RNA interference (RNAi) pathway under specific conditions. Activation of PRRs by DAMPs or PAMPs can stimulate resident and immune cells to secrete pro-inflammatory cytokines including monocyte chemoattractant protein (MCP-1). The RNAi pathway is activated by dsRNA which is processed into small interfering (si)RNAs that guide sequence-specific targeting of complementary messenger RNAs (mRNAs), resulting in reduced protein expression. Although both DAMPs and PAMPs stimulate inflammatory signaling, differences in how resident disc cells and immune cells respond to these stimuli remain unclear. This study evaluated the responses of NR8383 macrophages and annulus fibrosus (AF) cells following treatment with the DAMP, decorin, and the PAMP, LdsRNA. Specifically, this study aimed to evaluate MCP-1 production following decorin treatment, assess the effects of decorin on AF wound healing and evaluate MCP-1 production following LdsRNA treatment with a focus on potential RNAi-mediated suppression. To investigate sequence-specific effects, cells were treated with both sequence-specific MCP-1 LdsRNA and non-sequence-specific mCherry LdsRNA synthesized through in vitro transcription. Both cell types were treated with varying concentrations of decorin and LdsRNA, and secreted MCP-1 protein levels were quantified using enzyme-linked immunosorbent assays (ELISAs). AF cells demonstrated significant differences in MCP-1 production across decorin concentrations and treatment durations, with higher decorin doses inducing earlier MCP-1 production, while lower doses resulted in delayed responses. In contrast, unstimulated NR8383 macrophages exhibited high levels of MCP-1 production across all time points. Following LdsRNA treatment, NR8383 macrophages demonstrated significant differences in MCP-1 production between sequence-specific and non-sequence-specific LdsRNA when collapsed across doses, whereas AF cells did not demonstrate significant responses. Collectively, these findings suggest distinct responses of  NR8383 macrophages and AF cells to DAMP and PAMP stimulation under the experimental conditions used, highlighting the importance of considering cell type when investigating inflammatory mechanisms associated with IVD degeneration and LBP.

Convocation Year

2026

Convocation Season

Fall

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Available for download on Tuesday, August 17, 2027

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