Suppression of pre-B cell colony formation by catecholamine oxidation

The sympathetic nervous system regulates many aspects of lymphocyte function by stimulating adrenergic receptors with catecholamines. To determine the effects of β-adrenergic receptor (β-AR) signaling on B cell development, we utilized the pre-B cell colony forming unit (CFU-Pre-B) assay and found that isoproterenol (ISO), a β-AR selective catecholamine, potently suppresses pre-B cell colony formation. Further investigation using selective agonists and knockout mice showed that the colony suppression was independent of adrenergic receptor signaling. This colony suppression was replicated by treatment with epinephrine and other catecholamines and was reversed with the addition of an antioxidant. Catecholamine oxidation was reasoned to cause the suppression, as this reaction generates reactive oxygen species (ROS) and toxic products. Further experiments with CFU-Pre-B assays revealed a sensitivity to ambient oxygen and oxidative stress generated by menadione treatment. Analysis of cells within the ISO treated colonies revealed alterations in the distribution of pre- and pro-B cells at different stages of development and indicated delays in maturation. ISO treatment was not found to be immediately toxic within the timeframe of expected ISO oxidation, but a loss of cells with colony forming potential was observable once colonies started to grow. B cell progenitors showed a greater sensitivity to ISO or menadione treatment than myeloid and erythroid progenitors. Our study demonstrates an unexpected hypersensitivity of B cell progenitors to oxidative stress generated by catecholamine oxidation. This study also highlights the importance of additional controls when assessing neuroimmune interactions assumed to be mediated by signaling through adrenergic or dopaminergic receptors.