INDUCTION OF NITRIC-OXIDE SYNTHASE IN SUBSETS OF MURINE PULMONARY FIBROBLASTS - EFFECT ON FIBROBLAST INTERLEUKIN-6 PRODUCTION

The purpose of this investigation was to determine whether mouse lung fibroblast subsets have the ability to produce nitric oxide (NO), and if so, to characterize the induction and effects of its synthesis. Pre viously, we isolated Thy1(+) and Thy1(-) subpopulations of mouse lung fibroblasts, which differ in terms of cytokine production, morphology, response to cytokines and radiation, and ability to present antigen t o T lymphocytes. When treated with the proinflammatory cytokines IFN-g amma, TNF-alpha, and IL-1 alpha, these fibroblast lines produce microm olar quantities of NO2- and NO3-, two stable end products of the NO pa thway. A combination of all three cytokines provided the greatest indu ction, and there was no measurable production of NO in unstimulated ce lls. Thy1(+) fibroblasts have fewer requirements for induction of NO p roduction than the Thy1(-) line, in that NO production could be induce d by only two of the above cytokines, where the Thy1(-) fibroblasts re quired all three. Inducible NO synthase (iNOS) mRNA was shown to be pr esent by the reverse transcriptase-polymerase chain reaction as early as 2 hr after cytokine treatment in both cell lines. Addition of the N O synthase inhibitors N-G-monomethyl-L-arginine and amino-guanidine in hibited production of NO2- and NO3-, but not iNOS mRNA. This inhibitio n was partially reversed by the addition of an excess of L-arginine. I nterestingly, inhibition of NO synthesis was shown to decrease IL-6 pr oduction by more than 50% in cytokine-treated Thy1(+) fibroblasts. The se results indicate for the first time that Thy1(+) and Thy1(-) mouse lung fibroblast subsets have the capability to produce NO to differing extents in response to cytokines and may therefore play an important role in the inflammatory response in the lung as well as in the progre ssion of lung disease. (C) 1994 Academic Press, Inc.