Rapamycin Augments the NMDA-Mediated TNF Suppression of MRSA-Stimulated RAW264.7 Murine Macrophages

Abstract

Background. Methicillin-resistant Staphylococcus aureus (MRSA) can stimulate massive cytokine release. Ketamine suppresses tumor necrosis factor (TNF) secretion by MRSA-stimulated RAW264.7 macrophages, and the mechanism likely involves N-methyl-D-aspartic acid (NMDA) receptor antagonism. The downstream effects of NMDA-mediated TNF suppression, specifically the PI3K/Akt and mTOR modulation, have not been described. Methods. RAW264.7 cells were stimulated for 18 hrs with 105 to 107 CFU/mL inocula of either of two prototypical community-acquired- (CA-) MRSA isolates, USA300 strain LAC and USA400 strain MW2. Then we added the NMDA inhibitors ketamine or 2R-amino-5-phosphonopentanoate (AP5), NMDA substrate, LY294002, and rapamycin in various combinations. Results. NMDA inhibition suppressed TNF secretion by almost a third compared to the no-ketamine control. When NMDA substrate was added, the TNF secretion increased by 10%. Addition of LY294002 suppressed TNF production by macrophages by 20%. Rapamycin exhibited a concentration-dependent TNF induction-suppression response: induction at doses of 0.1 and 1 ng/mL and suppression at 10 and 100 ng/mL. Induction of TNF was abolished when LY294002 was added and the suppression became uniform. Ketamine-induced suppression of TNF secretion was intensified 10–15% when rapamycin was added, but not when LY294002 was added. Conclusion. These findings suggest that NMDA-induced TNF suppression can be augmented by concurrent mTOR inhibition.

Rapamycin unbalances the polarization of human macrophages to M1

“…In particular RAPA was shown to be a good candidate for pharmacological modulation of dendritic cells and CD4 + CD25 + regulatory T cells . Moreover, a growing body of evidence indicates that in myeloid phagocytes (monocytes, macrophages, granulocytes and myeloid dendritic cells), the mTOR pathway is crucial for survival and activation …”

“…[22][23][24][25][26][27] Moreover, a growing body of evidence indicates that in myeloid phagocytes (monocytes, macrophages, granulocytes and myeloid dendritic cells), the mTOR pathway is crucial for survival and activation. 19,[28][29][30][31] Plasticity is a hallmark of myeloid mononuclear phagocytes and in response to environmental signals these cells undergo different forms of polarized activation, the extremes of which are called classic or M1 and alternative or M2. 32,33 Although the central and pervasive action of RAPA in innate immune responses is becoming apparent, 30,34 its effect on macrophage viability or polarization is still discordant 19,31,35 or not yet studied.…”

“…19,[28][29][30][31] Plasticity is a hallmark of myeloid mononuclear phagocytes and in response to environmental signals these cells undergo different forms of polarized activation, the extremes of which are called classic or M1 and alternative or M2. 32,33 Although the central and pervasive action of RAPA in innate immune responses is becoming apparent, 30,34 its effect on macrophage viability or polarization is still discordant 19,31,35 or not yet studied.…”

Differential Role of Rapamycin and Torin/KU63794 in Inflammatory Response of 264.7 RAW Macrophages Stimulated by CA-MRSA

“…We have shown that the agonism of the NMDA receptor increases and antagonism of the same receptor diminishes the TNF response [ 6 ]. Downstream from the NMDA, the phosphoinositide-3 kinase/akt (PI3/AKT) also influences such response [ 6 , 7 ]. The next downstream signaling molecule is the mammalian target of Rapamycin (mTOR complex).…”

“…The role of mTOR as a central controller of cell growth, proliferation, and metabolism has been described but many aspects are unclear [ 8 , 9 ]. Previous studies by our group have shown that inhibition of mTOR by Rapamycin has a dose-dependent biphasic response on TNF secretion [ 7 ]. At very low doses, Rapamycin increases TNF production, but, at tenfold higher doses, TNF secretion is diminished.…”