LIPOPOLYSACCHARIDE IMPAIRS MACROPHAGE CYTOPLASMIC PH REGULATION UNDER CONDITIONS SIMULATING THE INFLAMMATORY MICROENVIRONMENT

Within the acidic inflammatory milieu, macrophages (mphis) must maintain their cytoplasmic pH (pH(i)) within a range conducive to optimal function. It was previously shown that metabolism of L-arginine at concentrations present in vitro in RPMI medium (1.14 mM) impairs the ability of mphis to regulate pH(i). However, concentrations of L-arginine in vivo reportedly range from approximately 100 muM in serum to less-than-or-equal-to 50 muM in wounds. To investigate the potential in vivo relevance of this inhibition, mphi pH(i) regulation was examined following incubation with low concentrations of L-arginine that mimic the inflammatory microenvironment, in the presence or absence of lipopolysaccharide (LPS). pH(i) regulation was evaluated as the ability of thioglycolate-elicited murine peritoneal mphis to recover from an imposed cytoplasmic acid load. The mphi pH(i) was measured using a pH-sensitive fluorescent probe. Following incubation for 2 h in the absence of LPS, the pH(i) recovery rate was equivalent in cells incubated with and without L-arginine. Coincubation with LPS, however, resulted in marked inhibition of pH(i) recovery at L-arginine concentrations as low as 12.5 muM. The inhibition was not due to LPS alone, since LPS without L-arginine was not inhibitory. Inhibition of pH(i) recovery was observed at LPS concentrations ranging from 10 ng/ml to 10 mug/ml. The L-arginine-dependent inhibition was apparent within 60 min of exposure to LPS, in both freshly harvested cells and cells preincubated for 2 h in the absence of L-arginine and then exposed to both L-arginine and LPS. Under conditions mimicking the in vivo setting, LPS-stimulated L-arginine metabolism impairs mphi pH(i) regulation. Modulation of pH(i) by this mechanism may compromise mphi function within the acidic microenvironment of inflammation.