Double-ratiometric fluorescence imaging of H(2)Se and O(2)˙(−) under hypoxia for exploring Na(2)SeO(3)-induced HepG2 cells' apoptosis

Sodium selenite (Na(2)SeO(3)), as an anti-tumor drug for inducing tumor cells' apoptosis, has been widely studied under normoxic conditions and has been shown to exhibit oxidative stress process-induced apoptosis. However, since the real tumor environment is hypoxic, the actual mechanism is still unclear. Hence, considering the main metabolite of Na(2)SeO(3) in the metabolic process to be hydrogen selenide (H(2)Se) and also that it can be converted to superoxide anion (O(2)˙(−)) instantaneously in the presence of oxygen, a dual-ratiometric fluorescence imaging system for simultaneous monitoring of the changes of H(2)Se and O(2)˙(−) induced by Na(2)SeO(3)-guided tumor cell apoptosis under hypoxic conditions was constructed. Two molecular probes NIR-H(2)Se and dihydroethidium were used to detect H(2)Se and O(2)˙(−), respectively, whereas Rhodamine 110 was used as the reference fluorophore. Notably, H(2)Se levels significantly increased under hypoxic conditions, but there was no change in the level of O(2)˙(−), which is inconsistent with the results of the previous researches. Therefore, we hypothesize that the mechanism of Na(2)SeO(3)-induced apoptosis for tumor cells is caused by reductive stress; also, this method can be applied for the future study of other anti-cancer selenium compounds.