Beaudry K L, Parsons C L M, Ellis S E, Akers R M
Department of Dairy Sciences, Virginia Polytechnic Institute and State University, Blacksburg 24061.
Department of Dairy Sciences, Virginia Polytechnic Institute and State University, Blacksburg 24061.
J Dairy Sci. 2016 Jan;99(1):796-804. doi: 10.3168/jds.2015-9972. Epub 2015 Nov 5.
Prepubertal mammary development involves elongation and branching of ducts and stromal tissue remodeling. This process is highly regulated and in mice is known to be affected by the presence of innate immune cells. Whether or not such immune cells are present or involved in bovine mammary development is unknown. For the first time, we determined the presence, location (relative to mammary ductal structures), and changes in numbers of eosinophils, mast cells, and macrophages in prepubertal bovine mammary tissue, and evaluated the effects of age, ovariectomy, and exogenous estrogen on numbers of each cell type. Chemical stains and immunofluorescence were used to identify the 3 cell types in formalin-fixed, paraffin-embedded mammary tissue from prepubertal female calves from 3 archived tissue sets. The ontogeny tissue set included samples of mammary tissue from female calves (n=4/wk) from birth to 6 wk of age. The ovary tissue set contained samples from ovary intact and ovariectomized heifers allowing us to investigate the influence of the ovaries on immune cells in the developing mammary gland in prepubertal heifers. Nineteen animals were intact or ovariectomized 30 d before sampling; they were 90, 120, or 150 d old at the time of sampling. A third tissue set, the estrogen set, allowed us to determine the effect of exogenous estrogen on innate immune cells in the gland. Eosinophils were identified via Luna staining, mast cells by May-Grunwald Giemsa staining, and macrophages with immunofluorescence. Key findings were that more eosinophils and mast cells were observed in near versus far stroma in the ontogeny and ovary tissue sets but not estrogen. More macrophages were observed in near versus far stroma in ontogeny animals. Eosinophils were more abundant in the younger animals, and fewer macrophages tended to be observed in ovariectomized heifers as compared with intact heifers and estrogen treatment resulted in a reduction in cell numbers. In summary, we show for the first time that innate immune cells are present in prepubertal bovine mammary tissue, localization varies by immune cell type, and abundance is related to proximity of epithelial structures and physiological state. We suggest a likely role for these cells in control of bovine mammary growth and ductal development.
青春期前乳腺发育涉及导管的伸长和分支以及基质组织重塑。这个过程受到高度调控，在小鼠中已知会受到先天免疫细胞的影响。此类免疫细胞是否存在于牛乳腺发育过程中或参与其中尚不清楚。我们首次确定了青春期前牛乳腺组织中嗜酸性粒细胞、肥大细胞和巨噬细胞的存在、位置（相对于乳腺导管结构）以及数量变化，并评估了年龄、卵巢切除和外源性雌激素对每种细胞类型数量的影响。使用化学染色和免疫荧光来鉴定来自3个存档组织集的青春期前雌性犊牛福尔马林固定、石蜡包埋乳腺组织中的这3种细胞类型。个体发育组织集包括从出生到6周龄的雌性犊牛（每周4头）的乳腺组织样本。卵巢组织集包含来自卵巢完整和卵巢切除的小母牛的样本，使我们能够研究卵巢对青春期前小母牛发育中乳腺内免疫细胞的影响。19只动物在采样前30天保持完整或进行了卵巢切除；采样时它们的年龄分别为90、120或150天。第三个组织集，即雌激素集，使我们能够确定外源性雌激素对乳腺内先天免疫细胞的影响。通过卢娜染色鉴定嗜酸性粒细胞，用美-格染色鉴定肥大细胞，用免疫荧光鉴定巨噬细胞。主要发现是，在个体发育和卵巢组织集中，与远基质相比，近基质中观察到更多的嗜酸性粒细胞和肥大细胞，但在雌激素组中并非如此。在个体发育动物中，近基质中观察到的巨噬细胞比远基质中更多。嗜酸性粒细胞在较年轻的动物中更为丰富，与完整小母牛相比，卵巢切除的小母牛中观察到的巨噬细胞往往较少，并且雌激素处理导致细胞数量减少。总之，我们首次表明先天免疫细胞存在于青春期前牛乳腺组织中，定位因免疫细胞类型而异，丰度与上皮结构的接近程度和生理状态有关。我们认为这些细胞在控制牛乳腺生长和导管发育中可能发挥作用。
