COOPERATIVE TURNING-ON OF THIN FILAMENT
COOPERATIVE TURNING-ON OF THIN FILAMENT
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DOI：
10.1016/8756-3282(93)90232-y
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发表时间：
1993-01-01
影响因子：
1.2
通讯作者：
ISHII, Y
中科院分区：
文献类型：
--
作者：
ISHII, Y
脊柱后纵韧带骨化（OPLL）是东方人椎管狭窄和脊髓病的常见原因。OPLL的特征是韧带组织中异位新骨形成。为探讨OPLL的发病机制，本研究对人后纵韧带细胞进行了体外培养，并对其形态学和生化特性进行了研究。来自正常脊柱韧带的对照组的细胞培养物没有显示出任何成骨特性。相反，从OPLL患者获得的细胞系（OG 1-OG 5）显示成骨细胞的几种不同表型特征。OG 1细胞表现出典型的成骨细胞样表型特征（即，体外钙化、高碱性磷酸酶[ALP]活性和甲状旁腺激素[PTH]引起的cAMP水平升高）。所有细胞系（OG 1-OG 5）通过显著增加cAMP水平对PTH和PGE 2作出反应。ALP活性在不同细胞系之间存在差异。OG 1和OG 2细胞显示出高水平的ALP活性。与非骨化组细胞相比，OG 3和OG 4细胞的活性较高，而OG 5细胞的活性不明显。仅在OG 3细胞中，CT引起cAMP水平和ALP活性的增加，并且其刺激作用表明CT对OPLL患者的韧带细胞具有直接的体外作用以刺激成骨细胞分化。很明显，来自OPLL韧带的一些细胞具有成骨细胞的几种表型特征，但来自无骨化韧带的细胞没有显示出任何成骨细胞特性。原肌球蛋白和肌钙蛋白的荧光探针监测了肌球蛋白亚片段1（S1）结合引起的细丝从“关闭”到“打开”的状态变化。在不存在SI的情况下，即使在Ca 2+的存在下，细丝也处于“关闭”状态。肌球蛋白头以及Ca 2+是打开细丝所必需的。Ca 2+与肌钙蛋白的结合使两种细丝状态之间的平衡向“转向状态”状态移动，从而促进细丝的开启。结合和交换实验表明，从肌钙蛋白中去除Ca 2+仅在“关闭”状态下稳定细丝复合物。因此，在肌球蛋白头的情况下由Ca 2+诱导的结构变化似乎在确定两个细丝状态之间的平衡中是重要的，即使Ca 2+本身不打开细丝。
Ossification of the posterior longitudinal ligament of the spine (OPLL) is a common cause of spinal canal stenosis and myelopathy in Orientals. OPLL is characterized by heterotopic new bone formation in ligamentous tissue. To investigate the pathogenesis of OPLL, human posterior longitudinal ligament cells were cultured and their in vitro morphological and biochemical characteristics were studied. Cell cultures from control subjects with normal spinal ligaments did not show any osteoblastic properties. In contrast, cell lines (OG1-OG5) obtained from an OPLL patient showed several different phenotypic characteristics for osteoblasts. OG1 cells showed typical osteoblast-like phenotypic characteristics (i.e., in vitro calcification, high alkaline phosphatase [ALP] activity, and elevation of cAMP levels by parathyroid hormone [PTH]). All cell lines (OG1-OG5) responded to PTH and PGE2 by markedly increasing cAMP levels. ALP activities varied among the cell lines. The OG1 and OG2 cells exhibited a high level of ALP activity. Compared with cell lines from the non-ossification group, the activities were higher in the OG3 and OG4 cells, but not significantly in the OG5 cells. Only in the OG3 cells, CT caused an increase in cAMP level and ALP activity, and its stimulatory effects demonstrated that CT had a direct, in vitro action on ligament cells of OPLL patients to stimulate osteoblastic differentiation. It is clear that some cells from ligaments with OPLL had several phenotypes characteristic of osteoblasts, but cells from ligaments without ossification did not show any osteoblastic properties. This observation is considered to be an important clue to understanding the pathophysiology of OPLL.Fluorescence probes on tropomyosin and troponin monitored the state change of the thin filament from the 'turned-off' to the 'turned-on' induced by the binding of myosin subfragment 1 (S1). In the absence of SI, the thin filament is in the 'turned-off' state even in the presence of Ca2+. The myosin head as well as Ca2+ is required for the turning-on of the thin filament. The binding of Ca2+ to troponin shifts the equilibrium between the two thin filament states towards the 'turned-state' state, facilitating the turning-on of the thin filament. The binding and the exchange experiments showed that removal of Ca2+ from troponin stabilized the thin filament complex only in the 'turned-off' state. Thus, the structural changes induced by Ca2+ in the absence of myosin head appears to be important in determining the equilibrium between the two thin filament state, even though Ca2+ itself does not turn on the thin filament.