Uro‐genital Development in Juvenile Sprague‐Dawley Rats: a Histomorphologic and Immunohistochemical Study

Abstract

Preclinical juvenile toxicity studies are crucial in the development of new drugs intended to be administered to children. While the normal histomorphological features of the uro‐genital systems in mature rats have been described, the physiological developmental changes occurring in neonatal and juvenile rats remain incompletely characterized. As the rat is the most commonly used species in toxicology studies, it is pivotal to further understand the normal tissue remodeling events, including apoptosis and cell proliferation occurring during the juvenile phase, in order to prevent mistaking these changes for pathological lesions. The objective of this study was to describe the key histomorphologic postnatal events occurring in the uro‐genital system of Sprague‐Dawley rats from birth to postnatal day (PnD) 30, with a special focus on cell proliferation and apoptosis. Formalin‐fixed and paraffin‐embedded kidneys, urinary bladder, ovaries, testes, epididymis, uterus and vagina were obtained from 51 Sprague Dawley rats divided in 13 timepoints according to age (PnD 1, 2, 4, 6, 8, 10, 14, 17, 21, 24, 26, 28, and 30). Whenever possible, equal numbers of females and males were used in each group. The ratios of the testes and kidney weights relative to the body weight was calculated for each timepoint, reaching a maximum value between PnD10 and PnD15 and on PnD30, respectively. Immunohistochemistry staining was performed to highlight apoptosis (caspase‐3) and cell proliferation (Ki‐67) when relevant. Hematoxylin and eosin slides revealed an increased cellularity in the kidney at PnD1 with the presence of a distinct nephrogenic zone in the outer one‐fourth of the cortex and contained different stages of developing nephrons. Nephrogenesis was characterized by abundant cell death and mitoses progressing from the outer cortex to the medulla, and ceased mostly by PnD14, as only few mitoses were observed at the cortico‐medullary junction at PnD21. At PnD1, the cuboidal epithelium lining the luminal surface of the urinary bladder consisted of two distinct cell layers and became 3 to 4 layers thick by PnD21. On PnD1, the ovarian parenchyma was hypercellular and diffusely composed of pockets of oocytes enveloped in stromal cells and, by PnD 26, a sharply demarcated cortico‐medullary separation was noticeable. The first uterine glands became visible around PnD14, whereas scattered apoptotic epithelial cells were still present up to PnD26. The vaginal epithelium showed multifocal mucification mixed with few apoptotic cells starting on PnD24. At PnD1, the seminiferous tubular epithelium was 1 to 2 layers thick, composed of spermatogonia and sertoli cells, and a seminiferous lumen was rarely noticeable. Pachytene spermatocytes were observed around the second week of life, while round spermatids were first visible on PnD26. This study showed that the urogenital system of Sprague‐Dawley rats is immature at birth and that different organs develop at various rates during the first month of age. Programmed cell death and cell proliferation coexist and should not be misinterpreted as drug‐related effects in toxicologic studies.Support or Funding InformationMitacs