INHIBITION OF ENDOTOXIN-INDUCED NITRIC-OXIDE SYNTHASE PRODUCTION IN MICROGLIAL CELLS BY THE PRESENCE OF ASTROGLIAL CELLS - A ROLE FOR TRANSFORMING GROWTH-FACTOR-BETA

In mixed glial cell cultures from cerebral cortices of newborn rats, e ndotoxin induces inducible nitric oxide (iNOS), nitric oxide (NO), and interleukin-1 beta (IL-1 beta) production in microglial cells. Earlie r we demonstrated that endotoxin induced iNOS but not IL-1 beta expres sion in microglial cells is inhibited by the presence of astroglial ce lls. In the present paper we describe studies on the mechanism by whic h astroglial cells exert selective suppressive action on iNOS expressi on by microglial cells. Expression of iNOS and IL-1 beta was studied b y single or double label immunocytochemical techniques and cell identi fication was performed with GSA-I-B-4-isolectin and an antibody agains t GFAP. Production of IL-1 beta and NO was determined by measurement o f IL-1 beta and nitrite concentrations in cell lysates and the culture medium, respectively. TGF beta, a cytokine known to inhibit NO produc tion by endotoxin challenged macrophages, was measured in culture medi um of mixed glial cell cultures using a bioassay. Microglial, astrogli al, and mixed glial cell cultures produced similar concentrations of T GF beta. The potential effect of TGF beta was studiedby using immunone utralizing antibodies against TGF beta 1 and TGF beta 2 on the inducti on of iNOS in microglial cells in the presence of astroglial cells. In cubation of the mixed glial cell culture with these TGF beta antibodie s (3 mu g/ml) markedly increased endotoxin-induced NO production and i NOS expression in microglial cells, whereas the production of IL-1 bet a was not affected. The antibodies against TGF beta 1 and TGF beta 2 m arginally increased NO production in pure microglial cell cultures, no netheless in cultures of purified microglial cells recombinant TGF bet a 1 and TGF beta 2 together with endotoxin inhibited NO production. We conclude that the presence of astroglial cells is essential for the i nhibitory effect of TGF beta on NO production by microglial cells (pos sibly) by activation of TGF beta or by increasing the sensitivity of m icroglial cells for TGF beta. (C) 1997 Wiley-Liss, Inc.