410 PLATELET-ACTIVATING FACTOR INDUCES DE NOVO SYNTHESIS OF MTOR IN TERMINALLY DIFFERENTIATED POLYMORPHONUCLEAR LEUKOCYTES

BackgroundThe mammalian target of rapamycin (mTOR) is known to play a pivotal role in the differentiation of hematopoietic stem cells (HSCs) via control of cell cycling. In addition to this function, mTOR is known to mediate translation of constitutively expressed mRNAs. Specifically, we have demonstrated the role of mTOR in translational regulation of key inflammatory mediators. In this investigation we hypothesized that an inflammatory agonist would increase synthesis of mTOR in a pool of terminally differentiated polymorphonuclear leukocytes (PMN).MethodsHuman HSCs were isolated from umbilical cord blood obtained from full-term neonates. This population of HSCs underwent proliferation and differentiation into mature PMNs. These cells were purified by magnetically selecting cells expressing the CD16+ cell surface antigen. The CD16+ cells were exposed to platelet-activating factor (PAF) 10 nM or HBSS for 0, 60, or 120 minutes. The cells were subsequently fixed and examined by immunocytochemistry for their expression of mTOR.ResultsThe population of terminally differentiated PMNs demonstrated increased expression of mTOR following stimulation with PAF compared with control. Further, this population of cells demonstrated a time-dependent increase in production of mTOR consistent with the expected rapid response of primary host defense cells.ConclusionsHSC-derived, terminally differentiated PMNs demonstrate functional integrity of inflammatory mediator-induced mTOR expression. This capacity likely enables the cell to control the rapid synthesis of other mediators of inflammation. This finding will be key for further investigation using this model to perform loss of mTOR function experiments on downstream inflammatory mediators and the concomitant alteration in inflammatory processes.