Optimizing a Human Osteoarthritic Articular ChondrocyteCulture System as a Test Platform for Therapeutics: A Pilot Study Comparing Extracellular Matrix Protection Factor 1 to Its Hyaluronic Acid Conjugated Counter Part

Date of Award

6-2018

Degree Type

Thesis

Degree Name

Master of Science in Biomedical Sciences

First Advisor

Marina D'Angelo, PhD

Second Advisor

Michael McGuinness, PhD

Third Advisor

Kathyrn Behling, MD, PhD

Abstract

Osteoarthritis (OA) is a disease that leads to the loss of cartilage. Articular cartilage is composed of chondrocytes surrounded by an extracellular matrix (ECM). During OA the ECM becomes degraded due to the imbalance of anabolic and catabolic processes. Many factors contribute to the disease process, including matrix metalloproteinase 13 (MMP-13), which targets collagen type II for degradation, a major structural protein of the ECM. Our group previously developed a three-dimensional, serum-free culture system of primary, human osteoarthritic articular chondrocytes (HOACs) to test the efficacy of a novel MMP-13 inhibitor, Extracellular Matrix Protection Factor 1 (ECPF-1), on collagen degradation. In this study, we optimized our culture system and compared the efficacy ofECPF-1 when conjugated to hyaluronic acid (HA), HA-ECPF-1. HOA Cs were isolated from patients who underwent a total knee arthroplasty. To optimize the system, we combined both the greatest and least pathology sides together and plated the number of viable cells at a density of 1.8 x 106 cells per 0.5 milliliter of alginate. Chondrocytes were treated with 250nM ofECPF-1, 250nM HA-ECPF-1, HA alone or serum-free media as a control. Conditioned media (CM) and alginate-associated matrix (AAM) were isolated on days 2, 5, 8 or 11 and analyzed for collagen metabolism. We successfully scaled-down our culture system so that we plated at least 4 fold more wells per patient. We pooled material in the smaller cultures to measure clinically relevant collagen degradation endpoints. Treating the patients with HA-ECPF-1 and ECPF-1 under mixed cultures of greatest and least pathology cells showed no significant effect and variation occurred across patients. When we cultured the HOA Cs to day 11, less total collagen I was produced and more collagen II was maintained in the AAM suggesting a lesser pathologic phenotype. Thus, the HOAC culture system would be best used as 24-72 hour cultures created from the least pathology cells only to test efficacy of potential OA therapeutics. In this investigation we demonstrated the efficacy of the new formulation, HA-ECPF-1, underscoring the use of HOACs to test several treatment conditions from one patient when screening potential OA therapeutics.

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