Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences (SGGW), Nowoursynowska 159, 02-776 Warsaw, Poland.
Eur J Cell Biol. 2011 Oct;90(10):854-64. doi: 10.1016/j.ejcb.2011.06.007. Epub 2011 Aug 24.
Autophagy is a catabolic process providing an alternative energy source for cells under stressful conditions such as starvation, growth factor deprivation or hypoxia. During involution of the bovine mammary gland autophagy is induced in mammary epithelial cells (MECs) as a survival mechanism, and is tightly regulated by hormones and growth factors necessary for gland development. In the present study we adapted the three-dimensional culture model to investigate the role of autophagy during formation of alveoli-like structures by bovine BME-UV1 MECs grown on extracellular matrix (ECM) components. Using confocal microscopy and Western-blot analyses of autophagic and apoptotic markers: LC3, and cleaved caspase-3, we showed that autophagy was induced in centrally localized cells within the developing acini. These cells lacked a direct contact with ECM, and formed a distinct population from the outer layer of cells. Induction of autophagy preceded apoptosis, but did not inhibit the formation of a hollow lumen. In the presence of steroid hormones: 17β-estradiol and progesterone, although autophagy was augmented, acini formation proceeded normally. In contrast, the major lactogenic hormone: prolactin, which supports functional differentiation of alveoli, did not alter induction of autophagy within the spheroids. BME-UV1 cells cultured on Matrigel in the presence of growth factors IGF-I and EGF formed larger, underdeveloped acini without lumens due to caspase-3 inhibition, and sustained autophagy in the centre of the spheroids, while TGF-β1 accelerated apoptosis, and increased autophagy significantly. Our observations suggest that sex steroids 17β-estradiol and progesterone, as well as growth factor TGF-β1 may regulate the development of the bovine mammary gland by inducing autophagy in addition to regulating proliferation and apoptosis of MECs. These data indicate that autophagy may play an important role during alveolargenesis.
自噬是一种分解代谢过程，可为细胞提供应激条件下（如饥饿、生长因子缺乏或缺氧）的替代能源。在牛乳腺的退化过程中，自噬被诱导发生在乳腺上皮细胞（MEC）中，作为一种生存机制，并且受到激素和生长因子的严格调节，这些激素和生长因子对于乳腺发育是必需的。在本研究中，我们采用了三维培养模型来研究自噬在牛 BME-UV1 MEC 形成肺泡样结构过程中的作用，这些细胞在细胞外基质（ECM）成分上生长。通过共聚焦显微镜和自噬和凋亡标志物（LC3 和 cleaved caspase-3）的 Western blot 分析，我们发现自噬被诱导发生在发育中的小泡中央定位的细胞中。这些细胞与 ECM 没有直接接触，并且形成了与外层细胞不同的群体。自噬的诱导先于凋亡，但并不抑制中空腔的形成。在类固醇激素 17β-雌二醇和孕酮存在的情况下，尽管自噬增强了，但小泡的形成仍正常进行。相比之下，主要的泌乳激素：催乳素，它支持肺泡的功能分化，并没有改变小泡内自噬的诱导。在生长因子 IGF-I 和 EGF 的存在下，BME-UV1 细胞在 Matrigel 上培养形成更大的、无腔的未成熟小泡，这是由于 caspase-3 抑制所致，并且中心的小泡维持自噬，而 TGF-β1 加速了凋亡，并显著增加了自噬。我们的观察结果表明，性类固醇 17β-雌二醇和孕酮以及生长因子 TGF-β1 可能通过诱导 MEC 的自噬来调节牛乳腺的发育，除了调节增殖和凋亡之外。这些数据表明，自噬可能在肺泡发生过程中发挥重要作用。
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