P007 - In Vitro Modelling of Human Cartilage: Comparing Traditional Culture Methods with Pregenerate Organ-on-a-Chip System
- T. Hopkins (Oswestry, GB)
- T. Hopkins (Oswestry, GB)
- P. Jasobhar (Cardiff, GB)
- H. McCarthy (Oswestry, GB)
- J. Rosser (Vienna, AT)
- S. Ali (Vienna, AT)
- C. Hulme (Oswestry, GB)
- K. Wright (Oswestry, GB)
Abstract
Purpose
In vitro modelling of human cartilage is commonly used to understand injury and disease in the tissue and to test treatment options. Organ-on-a-chip (OOAC) is an emerging modelling technology, incorporating cell culture chambers and microfluidics, that seeks to improve current modelling capabilities by increasing physiological relevance. Here, we assess the Pregenerate OOAC system to investigate human chondrocyte responses to pro-inflammatory stimuli cf. traditional chondrocyte culture techniques.
Methods and Materials
The Pregenerate OOAC consists of a central cell chamber (volume: 7.5µl) connected to a medium channel with an inlet at either end; six of such culture systems are contained on one chip (Figure 1A). Chondrocytes were isolated from macroscopically intact cartilage obtained during knee arthroplasty (n=5) and culture-expanded in monolayer to passage 2. Chondrocytes were encapsulated in Fibrin hydrogel and 2,200 chondrocytes loaded into each cell chamber of the microfluidic chips. Parallel cultures were set in high-density pellets (200k cells/pellet) or in monolayer (5000 cells/cm2). After 21 days, cultures were stimulated with a pro-inflammatory cocktail (50ng/ml TNF-α, 10ng/ml IL-1β and 25ng/ml IFN-γ) for 24 hours prior to assessing gene expression levels of COL1A1, COL2A1, COL10, ACAN, SOX9, MMP1, MMP3, MMP13 and ADAMTS-5.
Results
Fluorescence microscopy demonstrated evenly distributed chondrocytes within the culture chamber (Figure 1B). Pro-inflammatory stimulation down-regulated COL1A1, COL2A1, ACAN and SOX-9 in monolayer and pellet cultures only; COL10 was down-regulated in monolayer and pellet cultures but up-regulated in the Pregenerate chip. MMP3 and MMP13 were up-regulated in all three culture systems. ADAMTS-5 was down-regulated in the chips only. There was no change in expression of MMP1 in any of the culture systems.
Conclusion
These results demonstrate distinct differences in human chondrocyte responses to pro-inflammatory stimuli in the Pregenerate microfluidic chip cf. traditional culture conditions. Further work is needed to investigate these differences and which system is more synonymous with in vivo behaviour.
