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.