Expression and Localization of Ornithine Decarboxylase in Reversible Papillomatosis Induced by Uracil in Rat Bladder

Direct mechanical irritation by uracil calculi formed following feeding of 3% uracil in the diet to male rats produces severe papillary hyperplasia (papillomatosis, which is reversible) of bladder epithelium. To evaluate the mechanism of the appearance of uracil-induced papillomatosis, we examined the changes of the enzyme activity and the localization of ornithine decarboxylase (ODC), as well as polyamine biosynthesis, and epithelial proliferation, that accompany the sequential bladder epithelial changes following administration and withdrawal of uracil. Moreover, expression of ODC mRNA was investigated using northern blotting and localization of ODC mRNA was demonstrated using in situ hybridization. ODC activity during uracil administration was maintained at a high level compared to that in normal epithelium, but sharply decreased after cessation of uracil treatment. The accumulation of ODC protein was observed in the proliferating bladder epithelium by immunohistochemical examination and western blotting analysis, and even after cessation of treatment, the protein binding to anti-ODC antibody remained mildly elevated. Sequential changes of proliferating cell nuclear antigen (PCNA)-positive cells in the epithelium during the development and disappearance of papillomatosis correlated with ODC activity. ODC mRNA was expressed strongly in the proliferating epithelium in rats treated with uracil and weakly in normal epithelium, in accordance with the location of ODC protein. Consequently, our data demonstrate that cell proliferation in the development of papillomatosis is closely associated with polyamine metabolism, and moreover suggest that ODC activity is up-regulated at a post-translational step.     

Induction of the PAOh1/SMO polyamine oxidase by polyamine analogues in human lung carcinoma cells

Purpose The induction of polyamine catabolism has been directly associated with the cytotoxic response of various tumor types to the antitumor polyamine analogues. Initially, human polyamine catabolism was assumed to be under the control of a rate-limiting spermidine/spermine N¹-acetyltransferase (SSAT) that provides substrate for an acetylpolyamine oxidase (PAO). We have recently cloned a new polyamine analogue-inducible human polyamine oxidase (PAOh1/SMO) that efficiently uses spermine as a substrate. The induction of PAOh1/SMO in response to multiple polyamine analogues was examined in representative lung tumor cell lines.Methods Representatives of three different classes of antitumor polyamine analogues were examined for their ability to induce PAOh1/SMO.Results The human adenocarcinoma line, NCI A549 was found to be the most responsive line with respect to induction of PAOh1/SMO in response to analogue exposure. Similar to previous observations with SSAT expression, PAOh1/SMO induction was found to occur primarily in non-small-cell lung cancers cell lines. Using a series of polyamine analogues, it was found that the most potent inducers of PAOh1/SMO possessed multiple three-carbon linkers between nitrogens, as typified by N¹,N¹¹-bis(ethyl)norspermine.Conclusions Since PAOh1/SMO is an analogue-inducible enzyme that produces H₂O₂ as a metabolic product, it may play a significant role in determining the sensitivity of various human tumors to specific polyamine analogues.Abbreviations BENSpm N¹,N¹¹-bis(ethyl)norspermine - CHENSpm N¹-ethyl-N¹¹-(cycloheptyl)methyl-4,8,diazaundecane - CPENSpm N¹-ethyl-N¹¹-(cyclopropyl)methyl-4,8,diazaundecane - DAO Diamine oxidase - IPENSpm (S)-N¹-(2-methyl-1-butyl)-N¹¹-ethyl-4,8,diazaundecane - MAO Monoamine oxidase - MDL 72,527 (N¹,N⁴-bis(2,3-butadienyl)-1,4-butanediamine) - PAO Acetylpolyamine oxidase - PAOh1/SMO Human polyamine oxidase h1/spermine oxidase - SSAT Spermidine/spermine N¹-acetyltransferaseThis work was supported by NIH grants CA51085, CA58184, CA85509, and CA88843.