Santino P, Martignani E, Miretti S, Baratta M, Accornero P
Dipartimento di Scienze Veterinarie, Largo Braccini 2, 10095 Grugliasco (TO), Italy.
Gen Comp Endocrinol. 2017 Jun 1;247:87-96. doi: 10.1016/j.ygcen.2017.01.020. Epub 2017 Jan 22.
In the adult female, within the estrous cycle, the mammary gland undergoes multiple rounds of growth, with increased cellular proliferation, and involution, with increased apoptosis. The increase in proliferation is elicited by endocrine (Estrogen, Progesterone), as well as locally produced (epidermal growth factor, insulin-like growth factor, etc) growth factors. Among the genes that are modulated during cellular proliferation, immediate early genes play a fundamental role, being rapidly upregulated and then downregulated within the G0/G1 phase of the cell cycle, allowing the progression to the subsequent phases. Egrs (1-4) are immediate early genes that encode for transcription factors that promote, within different cell types and depending on the strength and duration of the stimuli, several different responses like mitogenesis, differentiation, apoptosis or even anti-apoptosis. In this work we have studied the mechanisms of modulation of the Egr family, in mammary epithelial cells of different origin (bovine, canine, feline, murine). Following stimulation with growth medium, Egr mRNA expression showed a strong upregulation reaching a peak at 45-60min, that rapidly declined. Among several cytokines, particularly important for mammary morphogenesis, that we have tested (EGF, IGF-I, insulin, estrogen, progesterone), only EGF upregulated Egrs to levels close to those elicited by growth medium. In order to understand how the Egr transcription factors were regulated, we have inhibited Erk 1/2 and PI3K, molecules that drive two major intracellular signaling pathways. Inhibition of the Erk 1/2 pathway totally abolished Egr upregulation mediated by growth medium or EGF. On the other hand, the PI3K-Akt pathway played a minor role on Egr levels, with a strong inhibitory effect on cat GH2 cells only, that could be ascribed to reduced Erk phosphorylation following PI3K inhibition. Finally we showed that addition of growth medium also upregulated that the mammary luminal marker cytokeratin 18, but only in the murine NMuMG cell line. This is the first manuscript describing how the Egr transcription factors are expressed in mammary epithelial cells of domestic animals and which growth factors and signaling pathways modulate their expression.
在成年雌性动物的发情周期中，乳腺会经历多轮生长，细胞增殖增加，随后是退化，细胞凋亡增加。增殖的增加是由内分泌（雌激素、孕酮）以及局部产生的（表皮生长因子、胰岛素样生长因子等）生长因子引发的。在细胞增殖过程中被调节的基因中，即刻早期基因起着基本作用，它们在细胞周期的G0/G1期迅速上调然后下调，从而使细胞进入后续阶段。Egr（1-4）是即刻早期基因，编码转录因子，根据刺激的强度和持续时间，在不同细胞类型中促进多种不同的反应，如促有丝分裂、分化、凋亡甚至抗凋亡。在这项工作中，我们研究了不同来源（牛、犬、猫、鼠）的乳腺上皮细胞中Egr家族的调节机制。用生长培养基刺激后，Egr mRNA表达强烈上调，在45-60分钟达到峰值，随后迅速下降。在我们测试的几种对乳腺形态发生特别重要的细胞因子（表皮生长因子、胰岛素样生长因子-I、胰岛素、雌激素、孕酮）中，只有表皮生长因子能将Egr上调至接近生长培养基引发的水平。为了了解Egr转录因子是如何被调节的，我们抑制了Erk 1/2和PI3K，这两种分子驱动两条主要的细胞内信号通路。抑制Erk 1/2通路完全消除了生长培养基或表皮生长因子介导的Egr上调。另一方面，PI3K-Akt通路对Egr水平的作用较小，仅对猫GH2细胞有强烈的抑制作用，这可能归因于PI3K抑制后Erk磷酸化减少。最后我们表明，添加生长培养基也上调了乳腺腔标记细胞角蛋白18，但仅在鼠NMuMG细胞系中。这是第一篇描述Egr转录因子在家畜乳腺上皮细胞中如何表达以及哪些生长因子和信号通路调节其表达的手稿。
