Impact of N-methyl-N-nitrosourea administration in female Sprague-Dawley rats whose dams were submitted to protein restriction: effects on susceptibility to mammary carcinogenesis in adult life

Abstract

Previous studies have indicated that maternal low protein induces changes in mammary gland development and increases the susceptibility to chemically-induced mammary carcinogenesis after three doses of N-methyl-N-nitrosurea (MNU, 50 mg/kg each) in female offspring Wistar rats (a resistant strain). Using Sprague-Dawley (SD) rats (a susceptible strain), we will investigate the effect of maternal low protein and only a single dose of MNU on mammary gland development and the risk for the development of mammary tumors in females offspring. Pregnant female SD will be fed a normal protein diet (NPD group, 17% protein) or low protein diet (LPD group, 6% protein) from gestational day 0 (DG0) until postnatal day 21 (PND 21, weaning). After weaning, the female offspring will be fed a commercial diet and euthanized at specific points. At PND 28 or 35, females from NPD and LPD groups will receive a single dose of 50 mg/kg of MNU and some females will be euthanized 24 hours after MNU application. Females from NPD and LPD groups will also be followed up to the DPN 250 to evaluate the appearance of MNU-induced mammary tumors. At PND 29 or 36, the abdominal mammary glands will be removed for developmental analysis (longitudinal and lateral diameter measurements and terminal end buds count), cell proliferation (Ki -67), apoptosis (caspase-3) and estrogen receptor alpha (RE-±) expression by immunohistochemistry. The appearance of the first tumor (latency) will be recorded and females with tumor> 2 cm in diameter will be euthanized. Tumor latency and mean number of tumors will be compared between NPD and LPD groups. At the end of the study, it is expected to evaluate the deleterious effects of maternal low protein on the mammary gland development, as well as, to establish the more adequate window for MNU administration (PND 28 or 35) with potential to increase the risk for development of mammary tumors in female offspring. (AU)