d-Amino acids differentially trigger an inflammatory environment in vitro

Abstract Studies in vivo have demonstrated that the accumulation of d -amino acids ( d -AAs) is associated with age-related diseases and increased immune activation. However, the underlying mechanism(s) of these observations are not well defined. The metabolism of d -AAs by d -amino oxidase (DAO) produces hydrogen peroxide (H 2 O 2 ), a reactive oxygen species involved in several physiological processes including immune response, cell differentiation, and proliferation. Excessive levels of H 2 O 2 contribute to oxidative stress and eventual cell death, a characteristic of age-related pathology. Here, we explored the molecular mechanisms of d -serine ( d -Ser) and d -alanine ( d -Ala) in human liver cancer cells, HepG2, with a focus on the production of H 2 O 2 the downstream secretion of pro-inflammatory cytokine and chemokine, and subsequent cell death. In HepG2 cells, we demonstrated that d -Ser decreased H 2 O 2 production and induced concentration-dependent depolarization of mitochondrial membrane potential (MMP). This was associated with the upregulation of activated NF-кB, pro-inflammatory cytokine, TNF-α, and chemokine, IL-8 secretion, and subsequent apoptosis. Conversely, d -Ala-treated cells induced H 2 O 2 production, and were also accompanied by the upregulation of activated NF-кB, TNF-α, and IL-8, but did not cause significant apoptosis. The present study confirms the role of both d -Ser and d -Ala in inducing inflammatory responses, but each via unique activation pathways. This response was associated with apoptotic cell death only with d -Ser. Further research is required to gain a better understanding of the mechanisms underlying d -AA-induced inflammation and its downstream consequences, especially in the context of aging given the wide detection of these entities in systemic circulation.