Expression of Evc2 in craniofacial tissues and craniofacial bone defects in Evc2 knockout mouse.
Expression of Evc2 in craniofacial tissues and craniofacial bone defects in Evc2 knockout mouse.
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DOI：
10.1016/j.archoralbio.2016.05.002
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发表时间：
2016-08
影响因子：
3
通讯作者：
Mochida Y
中科院分区：
文献类型：
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作者：
Badri MK;Zhang H;Ohyama Y;Venkitapathi S;Alamoudi A;Kamiya N;Takeda H;Ray M;Scott G;Tsuji T;Kunieda T;Mishina Y;Mochida Y
我们的目标是确定 EVC2 在颅面组织中的表达，并使用 Evc2 敲除 (KO) 小鼠模型研究 Evc2 缺陷对颅面骨的影响。 Evc2 KO 小鼠是通过引入提前终止密码子，然后引入与 β-半乳糖苷酶 (LacZ) 融合的内部核糖体进入位点而产生的。制备来自野生型 (WT)、杂合子 (Het) 和纯合 Evc2 KO 小鼠的样品。使用抗 β-半乳糖苷酶、抗 EVC2 和抗 SOX9 抗体进行 LacZ 染色和免疫组织化学 (IHC)。颅面骨被阿尔新蓝和茜素红染色。 KO中LacZ活性主要观察在内脏颅骨的前部。在 KO 和 Het 胚胎中，使用抗 β-半乳糖苷酶抗体进行 IHC，在许多软骨区域中鉴定出表达 Evc2 的细胞。在 WT 胚胎的这些区域观察到内源 EVC2 蛋白。抗SOX9抗体双重标记表明这些细胞主要是软骨细胞。在成年阶段，EVC2在鼻骨和蝶枕骨联合软骨细胞、前上颌骨和下颌骨的骨细胞和内皮样细胞中表达。骨骼双重染色表明，与WT相比，KO中观察到EVC2表达的颅面骨存在形态缺陷。据我们所知，我们的研究首次鉴定了颅面组织中 Evc2 表达细胞的类型。与表达模式一致，在 Evc2 KO 小鼠中发现异常的颅面骨形态，表明 EVC2 在颅面生长和发育过程中可能很重要。
Our objectives were to determine the expression of EVC2 in craniofacial tissues and investigate the effect of Evc2 deficiency on craniofacial bones using Evc2 knockout (KO) mouse model. Evc2 KO mice were generated by introducing a premature stop codon followed by the Internal Ribosomal Entry Site fused to β-galactosidase (LacZ). Samples from wild-type (WT), heterozygous (Het) and homozygous Evc2 KO mice were prepared. LacZ staining and immunohistochemistry (IHC) with anti-β-galactosidase, anti-EVC2 and anti-SOX9 antibodies were performed. The craniofacial bones were stained with alcian blue and alizarin red. The LacZ activity in KO was mainly observed in the anterior parts of viscerocranium. The Evc2-expressing cells were identified in many cartilageous regions by IHC with anti-β-galactosidase antibody in KO and Het embryos. The endogenous EVC2 protein was observed in these areas in WT embryos. Double labeling with anti-SOX9 antibody showed that these cells were mainly chondrocytes. At adult stages, the expression of EVC2 was found in chondrocytes of nasal bones and spheno-occipital synchondrosis, and osteocytes and endothelial-like cells of the premaxilla and mandible. The skeletal double staining demonstrated that craniofacial bones, where the expression of EVC2 was observed, in KO had the morphological defects as compared to WT. To our knowledge, our study was the first to identify the types of Evc2-expressing cells in craniofacial tissues. Consistent with the expression pattern, abnormal craniofacial bone morphology was found in the Evc2 KO mice, suggesting that EVC2 may be important during craniofacial growth and development.