13 Real time quality monitoring of engineered tissue for regenerative medicine
Despite the growing demand and interest in biofabrication in the field of tissue engineering for customized biological constructs, there are currently no efficient methods for monitoring function and physiological behaviour over time. This gap is a big disadvantage, since direct monitoring of fundamental metabolic parameters can improve the standardization of cell culture experiments and contribute to spread the use of customized biological constructs in the clinical scenario. Here, we created a non-invasive strategy to study cells with miniaturized, electrochemical micro-biosensors that allow in situ real-time monitoring. We developed sensors for oxygen and glucose, respectively using a copper wire inserted into a silica capillary tube, partly filled with a graphite-loaded epoxy resin, and a platinum wire. The sensors were introduced into the tissue plates through modified tissue plate lids, and. the detection of oxygen was obtained by means of an electrochemical reduction, i.e. applying a cathodic potential to the carbon-disk surface. Signal acquisition and calibration were performed using a 4 channel potentiostat. The sensors were not damaged over a period of 21 days in cell culture medium and proved to be non-cytotoxic for up to 7 days. Conventional chondrogenic protocols contain high glucose concentrations (25 mM), whereby electrochemical biosensor detection is still often limited to lower glucose ranges. We thus investigated the effect of different glucose concentrations on the chondrogenic differentiation of bone marrow-derived mesenchymal stem cells as well as tested if the addition of lipids could support differentiation suffering from low energy resources. Low glucose concentrations (5 mM) turned out to decrease cell matrix deposition during differentiation, while a supplementation of lipids could not compensate for this effect. These findings suggest a pivotal role of glycolysis versus beta fatty acid oxidation during commitment, which we envision to further pursue by a current study with alternative intermediates of glycolysis.