Department of Pediatric Surgery, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Pediatr Surg Int. 2024 Aug 14;40(1):225. doi: 10.1007/s00383-024-05809-7.
Megacystis microcolon intestinal hypoperistalsis syndrome (MMIHS) is defined as a congenital visceral myopathy with genetic mutations. However, the etiology and pathophysiology are not fully understood. We aimed to generate a gene leiomodin-1a (lmod1a) modification technique to establish a zebrafish model of MMIHS.
We targeted lmod1a in zebrafish using CRISPR/Cas9. After confirming the genotype, we measured the expression levels of the target gene and protein associated with MMIHS. A gut transit assay and spatiotemporal mapping were conducted to analyze the intestinal function.
Genetic confirmation showed a 5-base-pair deletion in exon 1 of lmod1a, which caused a premature stop codon. We observed significant mRNA downregulation of lmod1a, myh11, myod1, and acta2 and the protein expression of Lmod1 and Acta2 in the mutant group. A functional analysis of the lmod1a mutant zebrafish showed that its intestinal peristalsis was fewer, slower, and shorter in comparison to the wild type.
This study showed that targeted deletion of lmod1a in zebrafish resulted in depletion of MMIHS-related genes and proteins, resulting in intestinal hypoperistalsis. This model may have the potential to be utilized in future therapeutic approaches, such as drug discovery screening and gene repair therapy for MMIHS.
巨膀胱-小结肠-肠蠕动不良综合征（MMIHS）被定义为一种具有基因突变的先天性内脏肌病。然而，其病因和病理生理学尚未完全阐明。我们旨在建立一个 lmod1a 基因修饰技术，以建立 MMIHS 的斑马鱼模型。
我们使用 CRISPR/Cas9 靶向斑马鱼中的 lmod1a。在确认基因型后，我们测量了与 MMIHS 相关的靶基因和蛋白的表达水平。进行了肠道转运试验和时空映射分析，以分析肠道功能。
遗传确证显示 lmod1a 的外显子 1 中存在 5 个碱基对缺失，导致过早的终止密码子。我们观察到 lmod1a、myh11、myod1 和 acta2 的 mRNA 表达显著下调，并且突变体组中 Lmod1 和 Acta2 的蛋白表达也下调。lmod1a 突变体斑马鱼的功能分析表明，其肠道蠕动次数更少、速度更慢、长度更短。
本研究表明，lmod1a 在斑马鱼中的靶向缺失导致与 MMIHS 相关的基因和蛋白耗竭，从而导致肠道蠕动不良。该模型可能有潜力用于未来的治疗方法，如药物发现筛选和 MMIHS 的基因修复治疗。
