Parathyroid hormone-Smad3 axis exerts anti-apoptotic action and augments anabolic action of transforming growth factor β in osteoblasts
Parathyroid hormone-Smad3 axis exerts anti-apoptotic action and augments anabolic action of transforming growth factor β in osteoblasts
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DOI：
10.1074/jbc.m302566200
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发表时间：
2003-12-26
影响因子：
4.8
通讯作者：
Chihara, K
中科院分区：
文献类型：
--
作者：
Sowa, H;Kaji, H;Chihara, K
虽然一些研究表明甲状旁腺激素（PTH）对骨具有合成代谢作用，但其确切机制尚不清楚。另一方面，转化生长因子β (tgf - β)大量储存在骨基质中，通过局部注射刺激啮齿动物的骨形成。虽然我们之前的研究表明Smad3是刺激骨形成的重要分子，但尚未有关于甲状旁腺激素对Smad3影响的报道。在本研究中，我们研究了PTH对小鼠成骨细胞Smad3的影响及其生理意义。PTH在MC3T3-E1和大鼠成骨细胞UMR-106中促进Smad3 mRNA在10 min内的表达和蛋白水平呈剂量依赖性。蛋白激酶A （PKA）激活剂和蛋白激酶C （PKC）激活剂增加Smad3蛋白水平，PKA和PKC抑制剂均能拮抗PTH诱导的Smad3，表明PTH通过PKA和PKC途径促进Smad3的产生。接下来，我们利用台盼蓝、转移酶介导的缺口末端标记和Hoechst染色检测PTH和Smad3在这些细胞中的抗凋亡作用。PTH预处理或Smad3过表达均可减少地塞米松和乙泊苷诱导的凋亡细胞数量。此外，显性阴性突变体Smad3DeltaC取消了pth诱导的抗凋亡作用。另一方面，PTH增强了tgf - β诱导的转录活性。此外，PTH增强了tgf - β诱导的I型胶原的产生，而不影响tgf - β减少的MC3T3-E1细胞的增殖。这些观察结果表明，PTH通过加速Smad3的转录活性来放大tgf - β的合成代谢作用。总之，我们首先证明了PTH-Smad3轴在成骨细胞中具有抗凋亡作用，并增强了tgf - β在成骨细胞中的合成代谢作用。因此，PTH- smad3轴可能参与了PTH的骨合成代谢作用。
Although several studies indicated that parathyroid hormone (PTH) exerted anabolic action on bone, its precise mechanisms have been unknown. On the other hand, transforming growth factor beta (TGF-beta), abundantly stored in bone matrix, stimulates bone formation with a local injection in rodents. Although our previous study suggested that Smad3 is an important molecule for the stimulation of bone formation, no reports have been available about the effects of PTH on Smad3. In this present study, we examined the effects of PTH on Smad3 and the physiological significance in mouse osteoblastic cells. PTH promoted the expression of Smad3 mRNA within 10 min and the protein level in a dose-dependent manner in MC3T3-E1 and rat osteoblastic UMR-106 cells. Protein kinase A (PKA) activator as well as protein kinase C (PKC) activators increased Smad3 protein level, and both PKA and PKC inhibitors antagonized PTH-induced Smad3, indicating that PTH promotes the production of Smad3 through both PKA and PKC pathways. Next, we examined anti-apoptotic effects of PTH and Smad3 in these cells, employing trypan blue, transferase-mediated nick end labeling, and Hoechst staining. Pretreatment with PTH or overexpression of Smad3 decreased the number of apoptotic cells induced by dexamethasone and etoposide. Moreover, a dominant negative mutant, Smad3DeltaC, abrogated PTH-induced anti-apoptotic effects. On the other hand, PTH augmented TGF-beta-induced transcriptional activity. Furthermore, PTH enhanced TGF-beta-induced production of type I collagen, whereas it did not affect TGF-beta-reduced proliferation in MC3T3-E1 cells. These observations indicated that PTH amplified the anabolic effects of TGF-beta by accelerating the transcriptional activity of Smad3. In conclusion, we first demonstrated that PTH-Smad3 axis exerts anti-apoptotic effects in osteoblasts and reinforces the anabolic action by TGF-beta in osteoblasts. Hence, PTH-Smad3 axis might be involved in the bone anabolic action of PTH.