Expression of a Constitutively Active Form of Hck in Chondrocytes Activates Wnt and Hedgehog Signaling Pathways, and Induces Chondrocyte Proliferation in Mice
Expression of a Constitutively Active Form of Hck in Chondrocytes Activates Wnt and Hedgehog Signaling Pathways, and Induces Chondrocyte Proliferation in Mice
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DOI：
10.3390/ijms21082682
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发表时间：
2020-04-01
影响因子：
5.6
通讯作者：
Komori, Toshihisa
中科院分区：
文献类型：
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作者：
Matsuura, Viviane K. S. Kawata;Yoshida, Carolina Andrea;Komori, Toshihisa
Runx2 是软骨细胞增殖和成熟所必需的。在软骨细胞中Runx2靶基因的搜索中，我们发现Runx2上调软骨细胞中造血细胞激酶（Hck）的表达，Hck是Src酪氨酸激酶家族的成员，并且Hck在野生型小鼠的软骨肢体骨骼中表达较高，而在Runx2（-/-）小鼠的软骨肢体骨骼中表达较低，并且 Runx2 结合 Hck 的启动子区域。为了研究 Hck 在软骨细胞中的功能，使用 Col2a1 启动子/增强子产生了表达 Hck 组成型活性形式 (Hck(CA)) 的转基因小鼠。后肢骨骼融合，胫骨变成一个大的圆形肿块，生长板明显紊乱。软骨细胞成熟延迟到 E16.5，但此后加速。 BrdU 标记但非末端脱氧核苷酸转移酶介导的 dUTP 缺口末端标记 (TUNEL) 阳性的软骨细胞增加。此外，Hck 敲除减少了原代软骨细胞的增殖。在使用来自 Hck(CA) 转基因小鼠后肢 RNA 的微阵列和实时 RT-PCR 分析中，Wnt（Wnt10b、Tcf7、Lef1、Dkk1）和刺猬（Ihh、Ptch1 和 Gli1）信号通路基因的表达上调。这些发现表明，Hck的表达受Runx2调控，在软骨细胞中高表达，Hck(CA)激活Wnt和hedgehog信号通路，促进软骨细胞增殖而不增加细胞凋亡。
Runx2 is required for chondrocyte proliferation and maturation. In the search of Runx2 target genes in chondrocytes, we found that Runx2 up-regulated the expression of hematopoietic cell kinase (Hck), which is a member of the Src tyrosine kinase family, in chondrocytes, that Hck expression was high in cartilaginous limb skeletons of wild-type mice but low in those of Runx2(-/-) mice, and that Runx2 bound the promoter region of Hck. To investigate the functions of Hck in chondrocytes, transgenic mice expressing a constitutively active form of Hck (Hck(CA)) were generated using the Col2a1 promoter/enhancer. The hind limb skeletons were fused, the tibia became a large, round mass, and the growth plate was markedly disorganized. Chondrocyte maturation was delayed until E16.5 but accelerated thereafter. BrdU-labeled, but not terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive, chondrocytes were increased. Furthermore, Hck knock-down reduced the proliferation of primary chondrocytes. In microarray and real-time RT-PCR analyses using hind limb RNA from Hck(CA) transgenic mice, the expression of Wnt (Wnt10b, Tcf7, Lef1, Dkk1) and hedgehog (Ihh, Ptch1, and Gli1) signaling pathway genes was upregulated. These findings indicated that Hck, whose expression is regulated by Runx2, is highly expressed in chondrocytes, and that Hck(CA) activates Wnt and hedgehog signaling pathways, and promotes chondrocyte proliferation without increasing apoptosis.