Effects of Extracellular Vesicles from Blood-Derived Products on Osteoarthritic Chondrocytes within an Inflammation Model.

Alexander Otahal,Markus Neubauer,Andrea De Luna,Olga Kuten-Pella,Karina Kramer,Zsombor Lacza,Lukas B Moser,Stefan Nehrer

doi:10.3390/ijms22137224

Abstract

Osteoarthritis (OA) is hallmarked by a progressive degradation of articular cartilage. One major driver of OA is inflammation, in which cytokines such as IL-6, TNF-α and IL-1β are secreted by activated chondrocytes, as well as synovial cells—including macrophages. Intra-articular injection of blood products—such as citrate-anticoagulated plasma (CPRP), hyperacute serum (hypACT), and extracellular vesicles (EVs) isolated from blood products—is gaining increasing importance in regenerative medicine for the treatment of OA. A co-culture system of primary OA chondrocytes and activated M1 macrophages was developed to model an OA joint in order to observe the effects of EVs in modulating the inflammatory environment. Primary OA chondrocytes were obtained from patients undergoing total knee replacement. Primary monocytes obtained from voluntary healthy donors and the monocytic cell line THP-1 were differentiated and activated into proinflammatory M1 macrophages. EVs were isolated by ultracentrifugation and characterized by nanoparticle tracking analysis and Western blot. Gene expression analysis of chondrocytes by RT-qPCR revealed increased type II collagen expression, while cytokine profiling via ELISA showed lower TNF-α and IL-1β levels associated with EV treatment. In conclusion, the inflammation model provides an accessible tool to investigate the effects of blood products and EVs in the inflammatory context of OA.

Highlights

interleukin 1β (IL-1β) and tumor necrosis factor α (TNF-α) can induce the expression of other proinflammatory cytokines—such as IL-6, IL-15, IL-17, and IL-18 [10,11,12,13]—and on the other hand, they promote the production of the matrix-degrading enzymes matrix metalloproteinases (MMPs)—MMP-1, MMP-3, MMP-13—as well as a disintegrin and metalloproteinase (ADAM) with thrombospondin-1 domains (ADAMTS)-4 and -5
Prior to setting up the co-culture system schematically presented in Figure 1A, extracellular vesicles (EVs) were enriched via ultracentrifugation (UC) from the blood products Citrate-anticoagulated PRP (CPRP) and hyperacute serum (hypACT)
With the exception of elevated ACAN or COL2 expression in response to hypACT blood products or hypACT EVs, respectively, a low quantity and quality of gene expression changes was detected in chondrocytes via room temperature (RT)-qPCR

Summary

Introduction

Risk factors favoring the onset of OA include obesity, genetic predisposition, trauma, muscle weakness, physical activity levels, bone density, and nutritional status [2,3]. It is hallmarked by cartilage damage, intraarticular synovitis, subchondral remodeling, and joint pain [4]. IL-1β and TNF-α can induce the expression of other proinflammatory cytokines—such as IL-6, IL-15, IL-17, and IL-18 [10,11,12,13]—and on the other hand, they promote the production of the matrix-degrading enzymes matrix metalloproteinases (MMPs)—MMP-1, MMP-3, MMP-13—as well as a disintegrin and metalloproteinase (ADAM) with thrombospondin-1 domains (ADAMTS)-4 and -5 These induce the destruction of the matrix proteins type II collagen, glycosaminoglycans, and proteoglycans [14,15,16]

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