In vitro macrophage endotoxin tolerance: Defective in vitro macrophage MAPkinase signal transduction after LPS pretreatment is not present in macrophages from C3H/HeJ endotoxin resistant mice

Altered endotoxin (LPS) signal transduction in macrophages (M phi) may medi ate development of organ dysfunction in sepsis. C3H/HeJ M phi have a specif ic genetic defect that renders them "tolerant" to in vitro LPS activation. LPS tolerance can be induced in normal C3H/HeN M phi following in vitro LPS pretreatment. In these experiments, in vitro LPS-stimulated activation of M phi mitogen-activated protein (MAP) kinases were compared in C3H/HeJ and C3H/HeN mice. C3H/HeJ and C3H/HeN M phi were cultured +/- 10 ng/mL LPS pret reatment for 24 h, then stimulated with 0-1,000 ng/mL LPS for 6 h. Western blots were performed on lysates with monoclonal antibody to active ERK1,2 ( p42/44), stress-activated protein kinase (SAPK, p54/46), and p38 kinase. Su pernatant TNF or IL-l was determined by bioassay. High dose LPS stimulation activated ERK, SAPK, and p38 kinases in both C3H/HeN and C3H/HeJ M phi, ER K activation, p46 SAPK, and p38 activation were inhibited in C3H/HeN M phi after LPS pretreatment, whereas they were unchanged or increased in HeJ M p hi. TNF secretion was significantly decreased in C3H/HeN M phi following LP S pretreatment, but absent in C3H/HeJ M phi at all times. M phi from normal C3H/HeN mice rendered endotoxin tolerant by in vitro, low dose LPS pretrea tment have specific signal transduction defects that are not present in gen etically LPS hyporesponsive C3H/HeJ mice.