Mitochondrial Superoxide Signaling Contributes to Norepinephrine-Mediated T-Lymphocyte Cytokine Profiles

Norepinephrine (NE) produces multifaceted regulatory patterns in T-lymphocytes. Recently, we have shown that NE utilizes redox signaling as evidenced by increased superoxide (O(2)(●-)) causally linked to the observed changes in these cells; however, the source of this reactive oxygen species (ROS) remains elusive. Herein, we hypothesized that the source of increased O(2)(●-) in NE-stimulated T-lymphocytes is due to disruption of mitochondrial bioenergetics. To address this hypothesis, we utilized purified mouse splenic CD4+ and CD8+ T-lymphocytes stimulated with NE and assessed O(2)(●-) levels, mitochondrial metabolism, cellular proliferation, and cytokine profiles. We demonstrate that the increase in O(2)(●-) levels in response to NE is time-dependent and occurs at later points of T-lymphocyte activation. Moreover, the source of O(2)(●-) was indeed the mitochondria as evidenced by enhanced MitoSOX Red oxidation as well as abrogation of this signal by the addition of the mitochondrial-targeted O(2)(●-)-scavenging antioxidant MitoTempol. NE-stimulated T-lymphocytes also demonstrated decreased mitochondrial respiratory capacity, which suggests disruption of mitochondrial metabolism and the potential source of increased mitochondrial O(2)(●-). The effects of NE in regards to redox signaling appear to be adrenergic receptor-dependent as specific receptor antagonists could reverse the increase in O(2)(●-); however, differential receptors regulating these processes were observed in CD4+ versus CD8+ T-lymphocytes. Finally, mitochondrial O(2)(●-) was shown to be mechanistic to the NE-mediated T-lymphocyte phenotype as supplementation of MitoTempol could reverse specific changes in cytokine expression observed with NE treatment. Overall, these studies indicate that mitochondrial metabolism and O(2)(●-)-mediated redox signaling play a regulatory role in the T-lymphocyte response to NE.