Gas exchange is essential for bioreactor cultivation of tissue engineered cartilage

Abstract

Tissue engineered cartilage can be grown in vitro if the necessary physical and biochemical factors are present in the tissue culture environment. Cell metabolism and tissue composition were studied for engineered cartilage cultured for 5 weeks using bovine articular chondrocytes, polymer scaffolds (5 mm diameter × 2 mm thick fibrous discs), and rotating bioreactors. Medium pH and concentrations of oxygen, carbon dioxide, glucose, lactate, ammonia, and glycosoaminoglycan (GAG) were varied by altering the exchange rates of gas and medium in the bioreactors. Cell–polymer constructs were assessed with respect to histomorphology, biochemical composition and metabolic activity. Low oxygen tension (∼40 mmHg) and low pH (∼6.7) were associated with anaerobic cell metabolism (yield of lactate on glucose, YL/G, of 2.2 mol/mol) while higher oxygen tension (∼80 mmHg) and higher pH (∼7.0) were associated with more aerobic cell metabolism (YL/G of 1.65–1.79 mol/mol). Under conditions of infrequent medium replacement (50% once per week), cells utilized more economical pathways such that glucose consumption and lactate production both decreased, cell metabolism remained relatively aerobic (YL/G of 1.67 mol/mol) and the resulting constructs were cartilaginous. More aerobic conditions generally resulted in larger constructs containing higher amounts of cartilaginous tissue components, while anaerobic conditions suppressed chondrogenesis in 3D tissue constructs. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 63: 197–205, 1999.

Computational Study of Culture Conditions and Nutrient Supply in Cartilage Tissue Engineering

“…With respect to matrix synthesis, it has to be emphasized that not only is the presence or absence of nutrients relevant, but also the concentration level (4,21,(26)(27)(28). In general, for specific culture conditions, nutrient levels in the construct are determined by the ratio between the characteristic times for diffusion and uptake.…”

“…From studies using a tracer dye it was concluded that spinner flasks and RWV bioreactors closely approximated perfectly mixed conditions (55). However, the measured oxygen level in the medium ranged from 65% to 87% of the equilibrium with the incubator atmosphere in RWV and spinner flask culture (11,27). In addition, depending on the local flow environment concentrations around the construct may be lower due to cellular uptake (12).…”

“…In eq 1 the source terms q ox for oxygen and q glc for glucose are given by where V [mol cell -1 s -1 ] is the maximum uptake rate, K m [mol m -3 ] is the concentration at which the half- From the literature as summarized in Table 1, it is evident that there is a wide range of oxygen and glucose uptake rates for chondrocytes. For the reference situation, we use the values V ox ) 6.70 × 10 -15 mol cell -1 h -1 and V glc ) 6.12 × 10 -14 mol cell -1 h -1 , which were established for the same culture system for long-term culture under tissue engineering conditions (21,27). In addition, the maximum values for the uptake parameters (Table 1) were used, namely, V ox ) 1 × 10 -13 mol cell -1 h -1 and V glc ) 4.0 × 10 -13 mol cell -1 h -1 .…”

“…The initial value for glucose was set to 25 mM, based on medium containing 4.5 g L -1 glucose and no serum (25,27). The initial value for lactate is zero.…”

“…Most modeling studies consider only one metabolite, most often oxygen. It has been shown, however, that GAG synthesis is affected by nutrient availability (25), pH (26), and oxygen (27)(28)(29)(30)(31)(32)(33)(34). Complicated interactions exist between chondrocyte energy balance and matrix metabolism, and the precise regulation mechanisms remain largely unknown (28,(35)(36)(37)(38)(39)(40).…”