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Hepatocarcinoma exhibits wide variation in incidence world-wide. While it is uncommon in the United States and most of Western Europe, it represents the 1st commonest cause of cancer death in Taiwan. The epidemiological study of the disease indicates a significant higher incidence in the male than in the female. Recent evidence indicates that nitric oxide synthase ( NOS ) is present in fresh human tumor tissues and the enzyme activity is correlated positively with tumor grade , suggesting the association of nitric oxide ( NO ) with tumor progression. In human hepatoma cell lines , the role of NO in the cell proliferation has not been fully investigated. NO , a highly diffusable , colorless , free radical gas , may act as an intracellular or intercellular messenger. NO is synthesized from L-arginine by NOS. Three types of NOS have been described - the constitutive neuronal ( nNOS ) and endothelial ( eNOS ) types and the inducible ( iNOS ) macrophage type. NO has a wide variety of physiological functions , including cardiovascular , immunologic as well as neuronal actions. Some of these effects appear to be mediated by the activation of guanylyl cyclase. At elevated concentrations , NO can either be cytostatic/cytotoxic or cytoprotective. It has been shown that plasma concentrations of sex hormone , NO2-/NO3- ( NO metabolites ) as well as transforming growth factor b were elevated in hepatocarcinoma patients. In this study , we employed human hepatocarcinoma cell line HepT2 to investigate if endogenous NOS activity can be modulated by hormones , especially sex hormones , and if cell growth correlates with NO production. Our results showed that following a 48h exposure to cells , 17b-estradiol ( E2 ) at 10-7- 10-5 M inhibited cell growth and increased NO production. Triiodothyronine ( T3 ; known to increase NO ) also increased NO production , but have no effect on cell growth. Gonadotropin-releasing hormone ( GnRH agonist ; known to increase NO ) suppressed cell growth without affecting NO production. Studies with combination of hormones revealed that both T3 and GnRH agonist enhanced the E2 action on cell growth or NO production. The NOS inhibitor , L-NAME , inhibited the E2-induced NO production without affecting the E2 inhibited cell growth. Neither progesterone nor testosterone influenced cell growth or NO production. Studies concerning the influence of E2 and T3 on iNOS and estrogen receptor ( ER ) mRNA revealed E2 at 10-6、10-5 M for 48h increased the iNOS mRNA expression , however , this effect was abolished by T3. ERa mRNA expression was enhanced by 10-7 M E2 , but the enhancement was replaced by attenuation at higher concentration of E2 ( 10-5 M ). T3 antagonized the biphasic regulation of ERa mRNA by E2. In summary , E2 , but not progesterone or testosterone , inhibited cell growth and elevated NO production. These two responses to E2 were augmented by T3 or GnRH agonist. However , cell growth inhibition was not necessarily accompanied by increased NO production. The effect of E2 on iNOS and ERa mRNA expression antagonized by T3 , did not correlate well with E2 action on cell growth or NO production. The mechanism by which E2 inhibits cell growth and induces NO production in HepT2 cells remains to be determined.
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