Hiura K, Lim S S, Little S P, Lin S, Sato M
Lilly Research Laboratories, Indianapolis, Indiana, USA.
Cell Motil Cytoskeleton. 1995;30(4):272-84. doi: 10.1002/cm.970300405.
The expression and localization of tensin and cortactin were examined in osteoclast precursors in comparison with isolated osteoclasts on various substrates. Initially, the ability of hen monocytes to differentiate into osteoclasts was evaluated on plastic or glass, and compared to differentiation on bone. Specifically, monocytes were isolated from the medullary bones of egg-laying hens maintained on a Ca-deficient diet. Differentiation was monitored morphologically and by quantitation of the ability to form Howship's lacunae in bone slices or resorb radiolabeled bone particles of 20-53 microns diameter. These cells differentiated into tartrate resistant acid phosphatase (TRAP)-positive, bone resorbing, multinucleated syncytia in the presence of cytosine-1-beta-D-arabinofuranoside in a time dependent manner (day 1-6). Differentiation into osteoclast-like cells was similar whether cultured on plastic, on glass, or on bone. When compared to GAP-DH control levels, tensin and cortactin mRNA levels increased by 7- and 10-fold, respectively, by day 6. Tensin and cortactin protein levels also increased by 6- and 15-fold, respectively, by day 6. Immunofluorescence of differentiating precursors showed that tensin localized between regions of cell to cell contact and colocalized with vinculin in podosomes of osteoclast-like cells and of real osteoclasts. Cortactin immunofluorescence was not detectable in monocytes but localized inside tensin/vinculin podosome structures after fusion into osteoclast-like cells and in freshly isolated osteoclasts. Both tensin and cortactin were associated with attachment complexes used by osteoclast-like cells and osteoclasts to resorb bone. Specifically, punctate cortactin staining was observed inside tensin staining which formed a double ring structure at the membrane/bone interface of resorbing osteoclasts. These data showed that tensin and cortactin can be used as osteoclast differentiation markers, that participate in attachment complexes used to resorb bone, and that tensin may participate in the fusion process of osteoclast precursors.
在破骨细胞前体中检测了张力蛋白（tensin）和皮层肌动蛋白（cortactin）的表达及定位，并与在各种基质上分离得到的破骨细胞进行比较。最初，评估了母鸡单核细胞在塑料或玻璃上分化为破骨细胞的能力，并与在骨上的分化情况进行比较。具体而言，从维持缺钙饮食的产蛋母鸡的骨髓骨中分离单核细胞。通过形态学观察以及对在骨切片中形成Howship腔隙的能力或吸收直径为20 - 53微米的放射性标记骨颗粒的能力进行定量分析来监测分化情况。在胞嘧啶 - 1 - β - D - 阿拉伯呋喃糖苷存在的情况下，这些细胞以时间依赖性方式（第1 - 6天）分化为抗酒石酸酸性磷酸酶（TRAP）阳性、具有骨吸收能力的多核合体。无论在塑料、玻璃还是骨上培养，向破骨细胞样细胞的分化情况相似。与GAP - DH对照水平相比，到第6天时，张力蛋白和皮层肌动蛋白的mRNA水平分别增加了7倍和10倍。到第6天时，张力蛋白和皮层肌动蛋白的蛋白质水平也分别增加了6倍和15倍。对分化前体的免疫荧光检测显示，张力蛋白定位于细胞间接触区域之间，并与破骨细胞样细胞和真正破骨细胞的足体中的纽蛋白共定位。在单核细胞中未检测到皮层肌动蛋白免疫荧光，但在融合为破骨细胞样细胞后以及新鲜分离的破骨细胞中，皮层肌动蛋白定位于张力蛋白/纽蛋白足体结构内部。张力蛋白和皮层肌动蛋白都与破骨细胞样细胞和破骨细胞用于吸收骨的附着复合物相关。具体而言，在张力蛋白染色内部观察到点状皮层肌动蛋白染色，其在吸收性破骨细胞的膜/骨界面处形成双环结构。这些数据表明，张力蛋白和皮层肌动蛋白可作为破骨细胞分化标志物，参与用于吸收骨的附着复合物，并且张力蛋白可能参与破骨细胞前体的融合过程。
