Elsevier

Food and Chemical Toxicology

Volume 37, Issue 12, December 1999, Pages 1159-1166
Food and Chemical Toxicology

Research Section
Effect of urinary pH on the progression of urinary bladder tumours

https://doi.org/10.1016/S0278-6915(99)00111-8Get rights and content

Abstract

Systemic alkalosis has been postulated to enhance tumorigenesis, whereas systemic acidosis has been implicated to exert a favourable influence on tumour control and regression. In the present study the urinary pH was influenced by feeding acid-forming or base-forming diets, and the effect of alkaline or acid urine on the early and late progression phase of urinary bladder carcinogenicity was investigated in male Wistar rats. Bladder lesions were initiated by N-butyl-N-(4-hydroxybutyl) nitrosamine (0.05% BBN in the drinking water during 4 weeks) and promoted by sodium bicarbonate (3.4% NaHCO3 in the diet during 15 or 25 weeks). After short- (15 week) and more long-term (25 week) promotion with NaHCO3, groups of 20 rats were fed a diet containing the acidifying salt ammoniumchloride (2.1% NH4Cl) or the control diet. All surviving rats were killed after a total study duration of 52 weeks. Additional control groups were, after initiation, fed diets containing NaHCO3 and killed after 15 wk or 25 wk of promotion, or at the end of the study. In rats fed diets with added salts, water intake and the amount of urine produced were increased and the urinary density was decreased compared to rats fed control diet. During NaHCO3 feeding, urinary pH and sodium concentration were increased. During NH4Cl feeding, urinary pH was decreased and urinary chloride and calcium concentrations were increased. Initiation by BBN followed by treatment with NaHCO3 caused a high incidence of papillary/nodular hyperplasia, papillomas and carcinomas of the bladder epithelium. These lesions progressed with time or longer duration of NaHCO3 promotion. A tumour protective effect of urinary acidification by NH4Cl was not found. In fact, both acidification and prolonged alkalinization tended to aggravate the malignancy of bladder carcinomas.

Introduction

Since the beginning of this century, systemic alkalosis has been associated with a variety of malignant neoplasms and has been implicated as an enhancing factor in tumorigenesis. In contrast, a favourable influence of systemic acidosis on tumour control and regression has long been implicated (reviewed by Harguindey, 1982). Several reports have indicated that metabolic acidosis inhibits tumour growth in experimental tumours. Verne and Roth (1963)reported inhibition of the carcinogenicity of subcutaneous implantations of oestradiol benzoate in rabbits by acidification of drinking water. In another study it was found that lowering the systemic pH by ammonium chloride inhibited the growth of solid tumours and leukaemias in mice (Anghileri, 1975). Hydrochloric acid added to laboratory food has resulted in an increased rate of regression of subcutaneously implanted sarcoma 180 (Harguindey et al., 1979). Also in tissue culture, low environmental pH has been shown to inhibit cell proliferation, survival and activity of tumour cells (Tannock and Rotin, 1989; Wike-Hooley et al., 1984).

Spontaneous cell death has been observed to occur within regions of solid tumours. Although the causes of spontaneous cell death within tumours are not known, the poorly developed vasculature may contribute to this process, by failing to provide adequate nutrients or to remove catabolytes (Vaupel et al., 1989). Owing to the limited range of diffusion of oxygen within tissues, regions within solid tumours tend to become hypoxic. It is well known that the extracellular fluid in malignant tumours is acidic relative to that in normal tissue (Jähde et al., 1990; Wike-Hooley et al., 1984). Hypoxic regions of tumours are likely to be acidic because of dependence on glycolysis as a major source of metabolic energy, leading to accumulation of lactic acid and hydrolysis of ATP, with consequent reduction of extracellular pH (Hochachka and Mommsen, 1983; Maidorn et al., 1993). It has been hypothesized that the combination of hypoxia and lower extracellular pH may be responsible for cell death (Boyer et al., 1993; Rotin et al., 1986).

In the present study it was investigated whether tumour progression can be inhibited by a chronically decreased environmental pH. The urinary bladder was selected as target organ because, in contrast to the pH of blood and interstitial fluid which is controlled within narrow limits, the urinary pH can easily be manipulated.

Two-stage urinary bladder carcinogenesis, promoted by increases in both urinary sodium ion concentration and pH, could be inhibited by lowering the urinary pH through simultaneous oral administration of ammonium chloride during promotion (Fukushima et al., 1986; Hagiwara et al., 1994). The objective of the present study was to investigate the effect of low environmental pH on the progression of preneoplastic urinary bladder lesions initiated by BBN and promoted by short- (15 wk) or more long-term (25 wk) promotion with NaHCO3.

Section snippets

Chemicals

N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) was purchased from Organische Chemicaliën Service (Haarlem, The Netherlands). Sodium hydrogen carbonate (NaHCO3, purity>99%) was obtained from Boom b.v. (Meppel, The Netherlands). Ammonium chloride (NH4Cl) was obtained from Merck (Germany) (Art. 1145; Zur Analyse).

Animals

Weanling, SPF-bred male Wistar rats (Crl:WI(WU)BR) were purchased from Charles River Wiga GmbH (Sulzfeld, Germany) and acclimatized for 7 days. At the start of the study the rats were about

Results

Five rats were killed in extremis or found dead during the course of the study. All deaths occurred in groups with prolonged promotion with NaHCO3; namely, one rat of group E (in wk 38), two rats of group F (in wk 42 and 45) and two rats of group G (in wk 42 and 45). Growth curves are presented in Fig. 2. In the final stage of the study, mean body weights in all groups were lower than in group B (which received the control diet during the major part of the study). These differences were

Discussion

Treatment with BBN (initiation), followed by treatment with NaHCO3 (promotion) appeared to be an effective way to induce urinary bladder carcinogenesis in rats, causing high incidences of simple and P/N hyperplasia, papillomas and carcinomas of the urothelium. These results confirm the well established correlation between increased urinary pH and sodium concentration, and the occurrence of cell proliferation, hyperplasia and neoplasia of bladder urothelium, both in vivo (Fukushima et al., 1986;

Acknowledgements

We would like to thank Dr A.J.M. Hagenaars for statistical evaluation and Professor Dr V.J. Feron for his advice and stimulation.

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