UP-REGULATION OF LPS-INDUCED INOS ACTIVITY IN DIBUTYRYL-CYCLIC-AMP DIFFERENTIATED RAT ASTROCYTES

AIM: To study the effect of dBcAMP on bacterial endotoxin ISS-induced NOS activity. METHODS: Microscopic changes were observed. Nitrite leve ls were measured by fluorometric assay. NOS activity was measured by c itrulline assay. RESULTS: Within 3-4 h after the addition of dBcAMP 1 mmol . L-1 to culture medium, a morphological transformation reminisce nt of in vivo, differentiation occurred. Coincubation with LPS and dBc AMP 1 mmol . L-1 resulted in a marked increase in the nitrite producti on as compared with LPS alone. This increase was concentration- and ti me-dependent with a maximal effect after 24 h treatment. Nitrite produ ction stimulated by LPS is parallel to the degree of cell differentiat ion. After a 24-h costimulation with LPS and dBcAMP, L-citrulline form ation assay revealed a 3-fold increase in NOS activity over LPS treatm ent alone, Simultaneous incubation with L-NAME, completely inhibited t he stimulation effect of LSS/dBcAMP on nitrite production, Cycloheximi de and dactinomycin also suppressed enhancement of NOS activity stimul ated by LPS/ dBcAMP, both in nitrite production and citrulline assay, indicating that the enhancement of NOS activity was due to the express ion of inducible NOS (iNOS) gene and protein. CONCLUSION: Inflammatory signals can trigger astrocytes to express substantially different lev els of iNOS depending on their degree of differentiation.