Bolander Franklyn F
Department of Biological Sciences, University of South Carolina, Columbia, 29208, USA.
Mol Cell Endocrinol. 2005 Dec 21;245(1-2):105-10. doi: 10.1016/j.mce.2005.11.001. Epub 2005 Dec 5.
In mammary epithelial cells, prolactin (PRL) activates at least two signaling pathways: Jak/Stat and nitric oxide (NO). The former induces differentiation as measured by alpha-lactalbumin accumulation, while experiments with sodium nitroprusside (SNP) show that NO inhibits differentiation. In order to resolve this apparent contradiction, the kinetics, inducibility, and cellular localization of NO production and sensitivity in mammary cells were examined. First, mammary cells remained responsive to PRL throughout the incubation with respect to NO production. Second, although desensitization occurred with continuous PRL exposure, recovery began as quickly as 30 min after PRL withdrawal. Since PRL is secreted in pulses in vivo, complete desensitization was not a likely explanation for the cells' escape from NO inhibition. Finally, the cellular site of transduction was examined using the caveolar disrupting agent, methyl-beta-cyclodextrin (MBCD). MBCD inhibited the accumulation of PRL-induced NO but not alpha-lactalbumin. This finding was confirmed by membrane fractionation studies where the PRL-induced NO production occurred primarily in caveolae and PRL-stimulated tyrosine phosphorylation of Stat5, which transcribes the alpha-lactalbumin gene, occurred predominantly in noncaveolar membranes. Finally, endogenous elevations of NO by arginine did not inhibit differentiation. As such, the inhibition seen with SNP appeared to be an artifact of the ubiquitous generation of NO from SNP. Physiologically, PRL induces NO only in caveolae and this restricted distribution does not interfere with differentiation.
在乳腺上皮细胞中，催乳素（PRL）激活至少两条信号通路：Jak/Stat和一氧化氮（NO）。前者通过α-乳白蛋白积累来衡量诱导分化，而用硝普钠（SNP）进行的实验表明NO抑制分化。为了解决这一明显的矛盾，研究了乳腺细胞中NO产生的动力学、诱导性、细胞定位以及敏感性。首先，在整个孵育过程中，乳腺细胞对PRL刺激产生NO仍保持反应。其次，尽管持续暴露于PRL会导致脱敏，但在撤去PRL后30分钟内就开始恢复。由于PRL在体内是以脉冲形式分泌的，完全脱敏不太可能是细胞逃避NO抑制的原因。最后，使用小窝破坏剂甲基-β-环糊精（MBCD）来研究转导的细胞位点。MBCD抑制PRL诱导的NO积累，但不抑制α-乳白蛋白的积累。膜分离研究证实了这一发现，其中PRL诱导的NO产生主要发生在小窝中，而转录α-乳白蛋白基因的Stat5的PRL刺激的酪氨酸磷酸化主要发生在非小窝膜中。最后，精氨酸引起的内源性NO升高并不抑制分化。因此，SNP所见的抑制似乎是SNP普遍产生NO的假象。从生理学角度来看，PRL仅在小窝中诱导NO产生，而这种受限的分布并不干扰分化。
