Department of Medical Genetics and Developmental Biology, School of Basic Medical Sciences, Capital Medical University, Beijing 100069, China.
Center for Gene Diagnosis, Zhongnan Hospital, Wuhan University, Wuhan, Hubei 430071, China.
Int J Cardiol. 2018 Aug 15;265:188-194. doi: 10.1016/j.ijcard.2018.04.038. Epub 2018 Apr 11.
Atrial septal defect (ASD) is one of the most common cardiac malformations worldwide. Several genes have been identified so far, which can merely explain small proportion of all the cases, therefore, it is anticipated that there are additional genes causing ASD. The aims of this study were to identify the causal gene of ostium secundum atrial septal defect (ASDII) in a Chinese family.
Whole exome sequencing was performed in three affected members and one control in the ASDII family. We screened mutations of LBX2 in 300 unrelated ASD patients and validated in 400 normal controls by Sanger sequencing. LBX2 knockout zebrafish was generated by CRISPR/Cas9 to detect whether lbx2 deficiency influenced cardiac development.
A rare missense mutation in LBX2 (c.A403G: p.K135E) was identified as the pathogenic cause of ASD. Subsequent mutation screening revealed two missense variants in 3 of 300 sporadic patients. We observed expanded size of atrium and ventricle in LBX2 knockout zebrafish through hematoxylin-eosin staining, more incompact distribution of cardiac myocytes was also discovered in homozygote compared with in wildtype. Furthermore, we performed in situ hybridization of crip2 gene to trace the cardiac neural crest cells in the embryo stage and found that the migration of neural crest cells was obviously delayed in the homozygotes.
We identified LBX2 for the first time as a pathogenic gene of ASDII. LBX2 deficiency may cause abnormal development of heart through influencing the migration of neural crest cells and affect the process of cardiac septation.
房间隔缺损（ASD）是全球最常见的心脏畸形之一。迄今为止，已经确定了几个基因，但它们只能解释所有病例的一小部分，因此，预计还有其他导致 ASD 的基因。本研究的目的是确定一个中国家庭中继发孔型房间隔缺损（ASDII）的致病基因。
对 ASDII 家系的 3 名受影响成员和 1 名对照进行全外显子组测序。我们在 300 名无关 ASD 患者中筛选了 LBX2 的突变，并通过 Sanger 测序在 400 名正常对照中进行了验证。通过 CRISPR/Cas9 生成 LBX2 敲除斑马鱼，以检测 lbx2 缺失是否影响心脏发育。
在 LBX2 中发现了一个罕见的错义突变（c.A403G: p.K135E），这是 ASD 的致病原因。随后的突变筛查在 300 名散发性患者中的 3 名中发现了 2 个错义变体。通过苏木精-伊红染色，我们观察到 LBX2 敲除斑马鱼的心房和心室增大，与野生型相比，杂合子中心脏心肌细胞的分布也更加不紧凑。此外，我们还对 crip2 基因进行了原位杂交，以追踪胚胎期的心脏神经嵴细胞，发现神经嵴细胞的迁移在杂合子中明显延迟。
我们首次将 LBX2 鉴定为 ASDII 的致病基因。LBX2 缺失可能通过影响神经嵴细胞的迁移而导致心脏发育异常，并影响心脏间隔形成的过程。
