ASCs were cultured under normoxic conditions in complete DMEM up to 80% confluency for all experiments. Two different methods were used to induce hypoxia at 2% and <0.1% O2 concentration. For the 2% O2 concentration, ASCs were cultured in a humidified incubator (Thermo) at 37°C, 2% O2. For a hypoxic environment with less than 0.1% O2, the hypoxia system Anaero Pouch–Anaero; (Mitsubishi Gas Chemical Company Inc., Japan) was used as previously described (Stubbs et al., 2012 (link)). The final indoor gas compositions were <0.1% O2 and 15% CO2. In order to mimic ischemia and anoxia in vitro, the OGD model was established using a previously published method (Chen et al., 2019 (link)). Similarly, in our study, the OGD exposure means that ASCs were cultured in serum- and glucose-free DMEM under hypoxic conditions (<0.1% O2) for 24 h.
To evaluate the effect of the degree and duration of hypoxia on ASCs exposed to OGD, the following experimental groups were established:
Free full text: Click here