Molecular analysis of coronal perisutural tissues in a craniosynostotic rabbit model using polymerase chain reaction suppression subtractive hybridization.
Molecular analysis of coronal perisutural tissues in a craniosynostotic rabbit model using polymerase chain reaction suppression subtractive hybridization.
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DOI：
10.1097/prs.0b013e31821740e8
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发表时间：
2011-07
影响因子：
3.6
通讯作者：
Cooper GM
中科院分区：
文献类型：
--
作者：
Cray JJ Jr;Gallo PH;Durham EL;Losee JE;Mooney MP;Kathju S;Cooper GM
在美国，颅缝早闭（颅穹窿缝过早融合）的发病率为2，000 - 3，000例活产婴儿中有1例。这种情况会导致颅内压升高，头部形状严重改变，智力迟钝。我们以前曾描述过一群遗传性冠状缝骨性结合的兔子。该模型有助于描述与颅缝早闭相关的术后颅面生长。由于缺乏用于兔的分子工具，该模型的分子分析受到限制。为了了解颅缝早闭的发病机制，我们采用聚合酶链反应-抑制性扣除杂交（PCR SSH）比较了野生型（WT）和颅缝早闭（CS）兔缝周组织的基因表达。对来自10日龄WT（n=3）和CS（n=3）兔的合并的颅骨样品的RNA进行PCR SSH，以获得在WT组织中富集（在CS组织中低表达）或在CS组织中富集（与WT相比在CS中过表达）的cDNA克隆。鉴定了大约140个回收的在CS组织中上调的cDNA克隆和130个回收的WT组织克隆的差异表达。其中，通过定量逆转录酶（RT）-PCR确认了4个基因在CS缝合组织中过表达：β-珠蛋白、骨桥蛋白（SPP 1）、β-珠蛋白和组织蛋白酶K（CTSK）。两个基因被证实在CS样品中低表达：COL 3A 1和RNF 12。这些基因产物在我们的天然CS模型中的差异表达似乎是正常骨形成/吸收途径差异的结果。
In the United States the incidence of craniosynostosis (premature fusion of the sutures of the cranial vault) is 1 in 2,000-3,000 live births. The condition can cause increased intracranial pressure, severely altered head shape, and mental retardation. We have previously described a colony of rabbits with heritable coronal suture synostosis. This model has been instrumental in describing the post-surgical craniofacial growth associated with craniosynostosis. The molecular analysis of this model has been limited by the lack of molecular tools for use in rabbits. In order to understand the pathogenesis of craniosynostosis, we compared gene expression in perisutural tissues between wild-type (WT) and craniosynostotic (CS) rabbits using polymerase chain reaction-suppression subtractive hybridization (PCR SSH). PCR SSH was performed on RNA derived from pooled samples of calvariae from 10-day old WT (n=3) and CS (n=3) rabbits to obtain cDNA clones that are either enriched in WT tissues (underexpressed in CS tissue) or enriched in CS tissues (overexpressed in CS compared to WT). Differential expression was identified for approximately 140 recovered cDNA clones upregulated in CS tissues and 130 recovered clones for WT tissues. Of these, four genes were confirmed by quantitative reverse-transcriptase (RT)-PCR as being overexpressed in CS sutural tissue: β-globin, osteopontin (SPP1), SPARC, and cathepsin K (CTSK). Two genes were confirmed to be underexpressed in the CS samples: COL3A1 and RNF12. The differential expression of these gene products in our naturally occurring CS model appears to be the result of differences in the normal bone formation/resorption pathway.