Radio Nicholas M, Doctor John S, Witt-Enderby Paula A
Division of Pharmaceutical Sciences, Duquesne University, Pittsburgh, PA 15282, USA.
J Pineal Res. 2006 May;40(4):332-42. doi: 10.1111/j.1600-079X.2006.00318.x.
The goals of this study were to determine (a) if melatonin enhances human adult mesenchymal stem cell (hAMSC) differentiation into osteoblasts as assessed by measuring alkaline phosphatase (ALP) enzyme activity, and (b) identify potential signal transduction pathways that mediate this process. ALP activity significantly increased in hAMSCs following a 10-day incubation in osteogenic medium, relative to hAMSCs incubated in basal growth medium alone. Melatonin (50 nm), added in combination with the osteogenic medium, significantly increased ALP activity relative to osteogenic medium alone. Co-exposure of hAMSCs to osteogenic medium supplemented with melatonin and either pertussis toxin or the melatonin receptor antagonists, luzindole or 4P-PDOT (MT2 receptor selective), inhibited the melatonin-induced increase in ALP activity, indicating the involvement of melatonin receptors, in particular, MT2 receptors. Assessment of melatonin receptor function following exposure to osteogenic medium containing either vehicle or melatonin produced dichotomous results. That is, if the differentiation of hAMSCs into an osteoblast was induced by osteogenic medium alone, then 2-[125I]-iodomelatonin binding and melatonin receptor function increased. However, examination of melatonin receptor function following chronic melatonin exposure, an exposure that resulted in a 50% enhancement in ALP activity, revealed that these receptors were desensitized. This was reflected by a complete loss in specific 2-[125I]-iodomelatonin binding as well as melatonin efficacy to inhibit forskolin-induced cAMP accumulation. Further characterization of the mechanisms underlying melatonin's effects on these differentiation processes revealed that MEK (1/2) and ERK (1/2), epidermal growth factor receptors, metalloproteinase and clathrin-mediated endocytosis were essential while PKA was not. Our results are consistent with a role for melatonin in osteoblast differentiation. If so, then, the decrease in plasma melatonin levels observed in humans during late adulthood may further enhance susceptibility to osteoporosis.
(a) 褪黑素是否能增强人成年间充质干细胞（hAMSC）向成骨细胞的分化，这通过测量碱性磷酸酶（ALP）活性来评估；(b) 确定介导这一过程的潜在信号转导途径。与仅在基础生长培养基中培养的hAMSC相比，在成骨培养基中培养10天后，hAMSC中的ALP活性显著增加。与单独的成骨培养基相比，添加褪黑素（50 nM）与成骨培养基联合使用时，显著增加了ALP活性。hAMSC与添加了褪黑素的成骨培养基以及百日咳毒素或褪黑素受体拮抗剂鲁辛朵或4P-PDOT（MT2受体选择性拮抗剂）共同暴露，抑制了褪黑素诱导的ALP活性增加，表明褪黑素受体尤其是MT2受体参与其中。在暴露于含载体或褪黑素的成骨培养基后评估褪黑素受体功能产生了二分结果。也就是说，如果hAMSC向成骨细胞的分化仅由成骨培养基诱导，那么2-[125I]-碘褪黑素结合和褪黑素受体功能会增加。然而，在长期暴露于褪黑素后（这种暴露导致ALP活性提高50%）检查褪黑素受体功能，发现这些受体脱敏。这表现为特异性2-[125I]-碘褪黑素结合完全丧失以及褪黑素抑制福斯高林诱导的cAMP积累的功效丧失。对褪黑素影响这些分化过程的潜在机制的进一步表征表明，MEK（1/2）和ERK（1/2）、表皮生长因子受体、金属蛋白酶和网格蛋白介导的内吞作用是必不可少的，而PKA则不是。我们的结果与褪黑素在成骨细胞分化中的作用一致。如果是这样，那么在成年后期人类中观察到的血浆褪黑素水平下降可能会进一步增加患骨质疏松症的易感性。
