TUMOR-NECROSIS-FACTOR STIMULATES SYSTEM X(AG)(-) TRANSPORT ACTIVITY IN HUMAN ENDOTHELIUM

System X(AG)(-) is responsible for the carrier-mediated Na+-independen t transport of anionic amino acids such as glutamate and aspartate acr oss the plasma membrane of cells. In order to examine a possible role for cytokines in regulating System X(AG)(-) activity, the effect of TN F on [H-3]glutamate transport in cultured human umbilical vein endothe lial cells (HUVECs) was studied. Carrier-mediated glutamate uptake was accomplished by two high-affinity carriers, predominantly by a Na+-in dependent carrier (System x(AG)(-), 75% of total glutamate uptake) and , to a lesser extent by a Na+-dependent carrier (System X(AG)(-), 24% of total uptake). TNF treatment (10 ng/ml far 10 hr) resulted in an 80 % increase in Na+-independent glutamate transport activity with no cha nge in System X(AG)(-) activity. The TNF stimulatory effect was blocke d by actinomycin D and cycloheximide. TNF treatment increased System X (AG)(-) glutamate transporter V-max by 51% (control V-max = 2359 +/- 3 45 pmole/mg protein/min vs TNF V-max = 3569 +/- 436 pmole/mg protein/m in, P < 0.01) without altering transporter affinity (control K-m, 229 +/- 40 mu M glutamate vs TNF K-m = 224 +/- 60 mu M glutamate, P = NS). The protein kinase C (PKC) inhibitor chelerythrine chloride had no ef fect on the TNF-stimulated glutamate transport, indicating that the au gmented glutamate transport was not mediated by PKC activation. These data indicate that the TNF-stimulated glutamate transport in HUVECs re quires do novo protein synthesis, possibly of the System x(AG)(-) tran sporter protein itself. Accelerated glutamate transport provides a pre cursor for the biosynthesis of macromolecules and glutamine. (C) 1995 Academic Press, Inc.