Abstract
Understanding how the local cellular environment influences cell metabolism, phenotype and matrix synthesis is crucial to engineering functional tissue grafts of a clinically relevant scale. The objective of this study was to investigate how the local oxygen environment within engineered cartilaginous tissues is influenced by factors such as cell source, environmental oxygen tension and the cell seeding density. Furthermore, the subsequent impact of such factors on both the cellular oxygen consumption rate and cartilage matrix synthesis were also examined. Bone marrow derived stem cells (BMSCs), infrapatellar fat pad derived stem cells (FPSCs) and chondrocytes (CCs) were seeded into agarose hydrogels and stimulated with transforming growth factor-β3 (TGF- β3). The local oxygen concentration was measured within the center of the constructs, and numerical modeling was employed to predict oxygen gradients and the average oxygen consumption rate within the engineered tissues. The cellular oxygen consumption rate of hydrogel encapsulated CCs remained relatively unchanged with time in culture. In contrast, stem cells were found to possess a relatively high initial oxygen consumption rate, but adopted a less oxidative, more chondrocyte-like oxygen consumption profile following chondrogenic differentiation, resulting in net increases in engineered tissue oxygenation. Furthermore, a greater reduction in oxygen uptake was observed when the oxygen concentration of the external cell culture environment was reduced. In general, cartilage matrix deposition was found to be maximal in regions of low oxygen, but collagen synthesis was inhibited in very low (less than 2%) oxygen regions. These findings suggest that promoting an oxygen consumption profile similar to that of chondrocytes might be considered a key determinant to the success of stem cell-based cartilage tissue engineering strategies.
Highlights
IntroductionThe efficacy of stem cell based cartilage tissue engineering strategies is typically evaluated based on changes in the expression of chondrogenic matrix related genes, as well as the resulting biochemical composition and biomechanical function of the graft (Kafienah and Sims, 2004; Cernanec et al, 2007; Fehrer et al, 2007; Buckley et al, 2010; Meyer et al, 2010; Farrell et al, 2012, 2014; Thorpe et al, 2012; Carroll et al, 2014)
Agarose hydrogels seeded with either Bone marrow derived stem cells (BMSCs), fat pad derived stem cells (FPSCs) or CCs were cultured for 24 days in chondrogenic medium
On day 0, the lowest O2c levels were measured in BMSC seeded constructs (0.0% ± 0.0), whereas the highest levels were measured in the CC seeded constructs (4.6% ± 0.3)
Summary
The efficacy of stem cell based cartilage tissue engineering strategies is typically evaluated based on changes in the expression of chondrogenic matrix related genes, as well as the resulting biochemical composition and biomechanical function of the graft (Kafienah and Sims, 2004; Cernanec et al, 2007; Fehrer et al, 2007; Buckley et al, 2010; Meyer et al, 2010; Farrell et al, 2012, 2014; Thorpe et al, 2012; Carroll et al, 2014). MSCs, typically reside in tissues such as bone marrow, which has been shown to possess oxygen levels in the range of 4–7% in vivo (Grant and Smith, 1963; Kofoed et al, 1985; Harrison et al, 2002), and due to their perivascular niche (as reviewed by Caplan, 2007; da Silva Meirelles et al, 2008), are likely to experience oxygen levels at least at the higher end of this range This would suggest that undifferentiated MSCs possess a different oxygen consumption rate to that of differentiated chondrocytes. Introducing gradients of regulatory factors such as oxygen into cell laden constructs, for example through the use of novel culture systems (Thorpe et al, 2013), may be critical to engineering tissues with spatial complexity mimicking that observed in articular cartilage
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