Department of Dentistry - Orthodontics and Craniofacial Biology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen, the Netherlands; Department of Animal Ecology and Physiology, Radboud University, Nijmegen, the Netherlands.
Department of Animal Ecology and Physiology, Radboud University, Nijmegen, the Netherlands.
Mech Dev. 2020 Sep;163:103632. doi: 10.1016/j.mod.2020.103632. Epub 2020 Jul 12.
Valproic acid (VPA) is an anti-epileptic drug known to cause congenital craniofacial abnormalities, including orofacial clefts (OFC). The exact mechanisms by which VPA leads to craniofacial skeletal malformations are poorly understood. In this study, we investigated the effects of VPA on cartilage and bone formation in the zebrafish larval head during 1-13 hpf (early) and 25-37 hpf (late) development in which cranial neural crest cells (CNCCs) arise and then proliferate and differentiate, respectively. Double-staining for cartilage and bone at 5 dpf revealed that VPA reduced cartilage and bone formation in a dose-dependent manner after both early or late exposure. Several different CNCC-derived cartilage and bone elements were affected in both groups. In the early group (100 μM VPA), the posterior head length and the ethmoid plate were reduced in length (both p < 0.01), while mineralization of 4 out of 9 bone elements was often lacking (all p < 0.01). In the late group (100 μM VPA), also the posterior head length was reduced as well as the length of the ceratohyals (both p < 0.01). Similar to early exposure, mineralization of 3 out of 9 bone elements was often lacking (all p < 0.01). These results indicate that both CNCC formation (early) and differentiation (late) are hampered by VPA treatment, of which the consequences for bone and cartilage formation are persistent at 5 dpf. Indeed, we also found that the expression of several genes related to cartilage and bone was upregulated at 5 dpf. These data indicate a compensatory reaction to the lack of cartilage and bone. Altogether, VPA seems to induce craniofacial malformations via disturbed CNCC function leading to defects in cartilage and bone formation.
丙戊酸（VPA）是一种抗癫痫药物，已知其会导致先天性颅面畸形，包括口腔面裂（OFC）。VPA 导致颅面骨骼畸形的确切机制尚未完全清楚。在这项研究中，我们研究了 VPA 在斑马鱼幼虫头部的软骨和骨骼形成中的作用，分别在颅神经嵴细胞（CNCCs）出现和增殖分化的 1-13 hpf（早期）和 25-37 hpf（晚期）发育过程中进行。在 5 dpf 时对软骨和骨骼进行双重染色显示，VPA 在早期或晚期暴露后均以剂量依赖性方式减少软骨和骨骼的形成。两组均影响了几种不同的 CNCC 衍生的软骨和骨骼元素。在早期组（100 μM VPA）中，后头部长度和筛板长度均缩短（均 p<0.01），而 9 个骨骼元素中的 4 个的矿化通常缺乏（均 p<0.01）。在晚期组（100 μM VPA）中，后头部长度和角质骨的长度也缩短（均 p<0.01）。与早期暴露相似，9 个骨骼元素中的 3 个的矿化通常缺乏（均 p<0.01）。这些结果表明，VPA 处理既阻碍了 CNCC 的形成（早期），也阻碍了分化（晚期），其对骨骼和软骨形成的后果在 5 dpf 时仍持续存在。实际上，我们还发现与软骨和骨骼相关的几个基因的表达在 5 dpf 时上调。这些数据表明了对软骨和骨骼缺乏的补偿反应。总之，VPA 似乎通过干扰 CNCC 功能诱导颅面畸形，从而导致软骨和骨骼形成缺陷。
