Endotoxin activation of mitogen-activated protein kinase in THP-1 cells; diminished activation following endotoxin desensitization

The signal transduction events occurring in monocytes in response to endotoxin (LPS) stimulation are incompletely delineated, although pertussis toxin (PT)-sensitive G proteins and the mitogen-activated protein kinase (MAPK) cascade have been implicated. Cellular desensitization in response to 18-h pre-exposure to 1 mu g/mL LPS alters signal transduction pathways of cellular activation and decreases production of certain inflammatory mediators such as thromboxane (Tx)B-2, the stable metabolite of TxA(2). We hypothesized that LPS stimulation of the human monocyte cell hue THP-I occurs ria MAPK activation, and that LPS desensitization, induced by pre-exposure to LPS, is associated with altered signaling through the MAPK cascade, involvement of a specific MAPK, ERK, in LPS-stimulated TxB(2) production was further tested using a specific MAPK cascade inhibitor, PD98059 (PD), PD inhibited LPS and phorbol myristate acetate (PMA)-stimulated ERK activation as demonstrated by immunoblots using anti-activated ERK antibodies, PD significantly inhibited LPS and PMA-stimulated TxB(2) synthesis to non-detectable levels, suggesting an involvement of MAPK in LPS-stimulated activation. Because PT-sensitive G proteins mediate LPS-stimulated signal transduction, their role in MAPK activation was tested. Pretreatment with PT inhibited basal and LPS-stimulated, but not PMA-stimulated ERK activation. Activation of ERK after LPS desensitization was also assessed. LPS pre-exposure resulted in a profound decrease in LPS-stimulated activation of ERK, but did not affect PMA activation of ERK. These data implicate the involvement of the MAPK cascade in LPS-stimulated activation of THP-1 cells and suggest coupling of Gi proteins and MAPKs in LPS-stimulated events, LPS desensitization is associated with decreased MAPK activation, but does not impair MAPK activation by PMA, Thus, LPS desensitization appears to selectively alter signal transduction upstream of ERK.