Introduction
Articular cartilage has a limited repair capacity (Hunziker, 1999)
leaving it susceptible to damage from trauma or diseases such as
osteoarthritis (OA). Although total joint arthroplasty is currently the
standard-of-care with generally good clinical outcomes, this technique
has limited function and longevity in the younger population. Therefore,
recent efforts have focused on developing biological-based approaches
for cartilage repair. One such promising approach is defect resurfacing
with engineered cartilage (Waldman et al. , 2002). While much
progress has been made, it still has proven difficult to produce tissue
suitable for the repair of clinically-sized defects (2 –
6 cm2) (Brittberg et al. , 1994). One of the
major shortcomings of most cartilage tissue engineering approaches is
the generally low accumulation of tissue constituents (Buschmann et al.,
1992; Vunjak-Novakovic et al., 1999; Waldman et al. , 2002). To
address this challenge, several strategies have been investigated, such
as: growth factor stimulation (Chen et al. , 2014; Lam et
al. , 2015), gene therapy (Chen et al. , 2014), mechanical
stimulation (Waldman et al. , 2003), and bioreactor cultivation
(Khan et al. , 2009). While each strategy can accelerate tissue
formation, these approaches either rely on use of overly sophisticated
methods and/or suffer from off-target effects, thereby requiring
substantial efforts to implement or optimize.
An alternative and straightforward approach may be to control nutrient
metabolism. Several studies have demonstrated that increased nutrient
availability (primarily by increasing media volume) can improve
cartilaginous tissue growth (Mauck et al. , 2003; Heywood et
al. , 2006; Khan et al. , 2009; Oze et al. , 2012). While
these studies did not identify a particular mechanism, this effect may
be due to resultant changes in glucose metabolism (Oze et al. ,
2012). Chondrocytes are known for their anaerobic metabolism of glucose
(Khan et al. , 2009), even in the presence of oxygen (Warburg-like
effect) (Lane et al. , 1977; Suits et al. , 2008). After
uptake, glucose is primarily metabolized through the glycolytic and
fermentation pathways resulting in the release of lactate. The aerobic
pathway is typically not well utilized and little of the pyruvate
generated from glycolysis is metabolized through the tricarboxylic acid
(TCA) cycle (Lane et al. , 1977). Interestingly however,
chondrocytes tend to synthesize more extracellular matrix (ECM) when
cultured under conditions that elicit a switch in metabolism from
anaerobic to mixed aerobic-anaerobic metabolism (Lane et al. ,
1977; Obradovic et al. , 1999; Khan et al. , 2009). We
postulate that this metabolic switch may be directed by alterations in
hypoxia inducible factor 1α (HIF-1α) signaling. The transition to
different metabolic states can result in the pooling of intracellular
metabolites, several of which have been shown to stabilize HIF-1α by
interfering with proline-hydroxylase-2 (PHD2) (Lu et al. , 2002;
Kim et al. , 2010; Ren et al. , 2011; De Saedeleer et
al. , 2012; Bailey & Nathan, 2018); the enzyme responsible for
initiating HIF-1α degradation in the presence of oxygen. For
chondrocytes, this pseudo-hypoxic state (hypoxic gene expression in the
absence of hypoxia) is especially important as the target genes of HIF-1
affect both chondrogenesis and cartilage homeostasis (Goldring &
Marcu, 2009; Dengler et al. , 2014). Thus, the
purpose of this study is to investigate the effect of increasing
nutrient availability on glucose metabolism, HIF-1 signaling, and
resultant tissue formation by primary articular chondrocytes.