ROLE OF NITRIC-OXIDE IN PANCREATIC TUMOR-GROWTH - IN-VIVO AND IN-VITRO STUDIES

Nitric oxide (NO), an endogenous free radical, has been implicated in a wide range of biological functions. NO is generated enzymatically fr om the terminal guanidinonitrogen of L-arginine by nitric oxide syntha se (NOS). Despite intensive investigations, the role of NO - either as the primary product of the L-arginine/NOS pathway or provided from th e NO donor sodium nitroprusside (SNP) - in carcinogenesis and tumour c ell growth remains unclear and controversial. The objective of this st udy was to examine the growth effects of NO on a ductal pancreatic ade nocarcinoma in the rat and on a human pancreatic tumour cell line (HA- hpc(2)). In vivo, both SNP and endogenous induction of NO by endotoxin s [lipopolysaccharide (LPS)] plus L-arginine significantly reduced the tumour growth. To investigate the mechanisms of NO anti-tumour growth action, the effects of either the SNP or L-arginine/NOS pathway were analysed on the HA-hpc, cell line. Nitrite/nitrate production, NOS act ivity and iNOS expression [assessed by reverse transcription-polymeras e chain reaction (RT-PCR)] were tested and related to growth (assessed by [H-3]thymidine incorporation assay) and apoptosis (assessed by int ernucleosomal DNA cleavage). SNP exerted a dual effect on tumour cells : stimulation of the proliferation up to 1 mM and inhibition at higher concentrations. These effects were related to NO production. Both pro liferative and cytostatic responses were inhibited by NO scavenger nyl -4,4,5,5-tetramethyl-hemidazoline-1-oxyl3-oxide (carboxy-PTIO). The ma rked apoptotic DNA fragmentation induced by SNP was also abolished by PTIO association. Unlike macrophages, the human pancreatic tumour cell s did not seem to express intrinsically the L-arginine/NOS pathway. Ma crophages were activated by HA-hpc, cells as well as by LPS plus cytok ines [interleukin (IL)-1 beta plus tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma]. In HA-hpc(2)/macrophage co-cultures, NOS activity and inducible NOS (iNOS) transcription were stimulated, where as an antiproliferative response was observed. These effects were rela ted to both macrophage amount and NO production. Addition of LPS plus cytokines to cc-cultures doubled iNOS activity, nitrite/nitrate produc tion and tumoricidal effect. These data suggest the involvement of NO in pancreatic tumour growth and support the fact that generation of hi gh levels of NO with potential production of endogenous reactive nitro gen intermediates may contribute to induction of apoptosis and tumour growth inhibition.