ABSTRACT

Microcystins and nodularin, isolated from toxic blue-green algae, are hepatotoxic monocyclic polypeptides. Both microcystins and nodularin inhibited in vitro protein phosphatase activity present in a cytosolic fraction of mouse liver, bound to the okadaic acid receptors, protein phosphatases 1 and 2A, and thus resulted in the increase of phosphoproteins; this was referred to as the apparent "activation" of protein kinases. Their concentrations causing 50% of the maximal effects are comparable to that of okadaic acid, a potent protein phosphatase inhibitor and a potent tumor promoter, in the nanomolar range of concentration. The increase of phosphoproteins was observed in rat primary cultured hepatocytes and was subsequently associated with morphological changes, which appeared to be a step in the process of hepatotoxicity. The well-known hepatotoxic compounds, alpha-amanitin and phalloidin, did not show any effects similar to those of microcystins, nodularin and okadaic acid. It is suggested that the hepatotoxicity of microcystins and nodularin may result from inhibition of protein phosphatases and the increase of phosphoproteins.

INTRODUCTIONVarious kinds of compounds have been found to inhibit tumor promotion in mouse skin. These include protease inhibitors, vitamin A derivatives, anti-inflammatory steroids, phosphodiesterase inhibitors, lipoxygenase inhibitors, phospholipase A, inhibitors, inhibitors of polyamine synthesis and of histidine decarboxylase, and calmodulin antagonists (Troll et al., 1970;Hozumi et a/., 1972; Verma et a/., 1979;Schwarz et al., 1977;Fischer et al., 1982;Nakadate et al., 1982; Takigawa et al., 1982; Umezawa et al., 1983; Belman and Troll, 1974;Nishino et al., 1984a;Slaga et al., 1982). Tumor promotion can be inhibited by use of various known inhibitors of specific biochemical reactions induced by tumor promoters, but we considered that the inhibitory effects of natural compounds present in daily foods should also be investigated.Recently, quercetin was found to inhibit the tumor promoting activity of 12-U-tetradecanoylphorbol-13-acetate (TPA), or teleocidin in two-stage carcinogenesis experiments in mouse skin initiated with 7,12-dimethylbenz(a)anthracene (DMBA) (Kato et al., 1983;Nishino et al., 1984a;Fujiki et al., 1986). In addition, we demonstrated that other flavonoids inhibit the in uitro and in vivo effects induced by tumor promoters (Horiuchi et al., 1986). Since quercetin and other flavonoids are present in foods, such as vegetables and fruits, we realized that antitumor promoters are present in daily foods.Every day Japanese people drink green tea, which contains (-)-epigallocatechin gallate, EGCG (Fig. I), as its main polyphenolic constituent. Therefore, in this work, we studied the influence of EGCG on tumor promotion such as its effects on the specific binding of [3H]TPA to a mouse particulate fraction in uitro, its effect on phorbol ester receptors in uiuo and its effect on activation of protein kinase C by teleocidin in uitro. EGCG inhibited tumor promotion by teleocidin in a two-stage chemical carcinogenesis experiment on mouse skin. MATERIALS AND METHODSMaterials. The preparation of EGCG used contained EGCG (85%), ( -)-epicatechin (10%) and ( -)-epicatechin gallate ( 5 % ) , as determined by high performance liquid chromatography (HPLC) on a YMC-A312 (ODS) column (6 x 150 mm), with water-acetonitrile cetic acid (85 : 10 : 5) as a solvent, monitoring the UV absorption at 254nm. This EGCG was isolated from Japanese green tea leaves as follows: the tea leaves were extracted with a boiling mixture of methanol-water ( I : l), the solvent was evaporated, and the residue was dissolved in water and applied to a column of DIAION HP-20 [ Mitsubishi, Kasei, Japan]. Material was eluted with stepwise increasing concentrations of methanol, and fractions containing EGCG were detected by HPLC. These fractions were pooled, concentrated, and extracted with ethyl acetate. The ethyl acetate extract was chromatographed on a column of Toyo-pearl HW-40 (coarse) [Toyo Soda, Japan], by elution with a water- Inhibition of specific I'HJTPA binding. A particulate fraction containing phorbol ester receptors was prepared...

Rectal carcinoid tumors that satisfy the following three conditions are indicated for local resection, including endoscopic polypectomy: a maximum diameter of < or = 10 mm, no invasion of the muscularis propria, and no depression or ulceration in the lesion. Endoscopic ultrasonography also is useful for estimating the depth of invasion of rectal carcinoid tumors and for determining whether endoscopic polypectomy is indicated.

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