The
tumor-promoting effect of
nodularin during
carcinogenesis was investigated. Male Fischer 344 rats were injected with
nodularin for 10 weeks from week 3 after
N-nitrosodiethylamine initiation without partial
hepatectomy. Rats were further maintained for 10 weeks after the cessation of
nodularin and were periodically killed. In contrast to the minimal foci in the DEN and
nodularin alone groups, treatment with DEN and
nodularin produced four kinds of nodules with eosinophilic, clear, mixed and basophilic cells. After the cessation of
nodularin, the maximally increased number, but not the area, of
glutathione S-transferase placental form-positive [GST-P(+)] nodules at week 12 decreased significantly and the appearance of two types of hyperplastic nodules was noted by GST-P immunostaining; homogeneously stained dense nodules (DN) and heterogeneously stained pale nodules (PN), which appeared only after the cessation of
nodularin. DN were well circumscribed by
enzyme-altered cells, as opposed to poorly in PN. Moreover, normal-appearing hepatocytes replaced the
enzyme-altered cells in PN. In contrast to the higher
PCNA index in GST-P(+) DN, the background level returned to that of the control at week 15.
PCNA indices in DN were significantly higher than in PN, which were still higher than the control, indicating that
nodularin affected the
PCNA index differentially in the altered and unaltered hepatocytes. However,
nodularin without DEN initiation significantly increased the
PCNA index through initial cell death and subsequent hepatocyte proliferation. These results suggest that: (i)
nodularin has a promoting effect by inducing hepatocyte proliferation in both
enzyme-altered hyperplastic nodules and the surrounding parenchyma; (ii) proliferation is transient in background cells but not in
enzyme-altered hepatocytes; (iii) GST-P(+) DN can be regarded as progressive and GST-P(+) PN as regressive, revealed by both immunohistochemistry and
PCNA index.