Mammary gland macro- and microstructural development during the first pregnancy following in utero heat stress exposure

Introduction In utero heat stress during late gestation impairs heifer growth and mammary development, reducing first-lactation productivity. However, whether these prenatal effects persist through the heifer’s first gestation remains unclear. This study evaluated the carryover effects of late-gestation heat stress on growth, endocrine profiles, and mammary gland development during first pregnancy. Methods Multiparous cows were exposed during the last 54 d of gestation to either heat stress (shade only; HS) or active cooling (shade, fans, and soakers; CL) under subtropical conditions, generating in utero heat-stressed (IUHS) and cooled (IUCL) heifers. After birth, all heifers were managed identically. Once pregnant, body weight, hip height, chest girth, and body length were recorded monthly. Teat traits and mammary gland (MG) dimensions were assessed at 2, 4, 6, and 9 months of gestation (MOG). Plasma progesterone was measured monthly, and estradiol at 8 and 9 MOG. At 6 and 9 MOG, high-resolution imaging and 3D MG reconstructions were performed, and biopsies were collected for histological analyses, including quantification of mammary epithelial cells (MEC), alveoli, lumen area, and MEC area. Fertility, calf birth weight, and milk production outcomes were also evaluated. Results IUHS heifers required more services per pregnancy and had shorter gestation lengths compared with IUCL, whereas overall growth traits and circulating hormones were similar between groups. At 6 MOG, IUHS heifers exhibited longer teats, greater inter-teat distances, and increased MG surface area. By 9 MOG, IUHS heifers displayed narrower, more elongated glands with greater teat spacing and eccentricity. Histologically, IUHS heifers had fewer MEC per alveolus and smaller alveolar areas. Milk production differences were minimal, with reduced yield in IUHS animals observed only at week 16 of lactation. Discussion Prenatal heat stress induces persistent alterations in mammary gland morphology and microstructure that extend into first gestation, despite limited effects on whole-body growth and endocrine profiles. These changes in alveolar development and MEC abundance may underlie subtle impairments in lactational performance, highlighting the long-term biological significance of late-gestation heat stress on mammary function.