Genetically Engineered Mouse Models Unveil Mechanisms and Therapeutic Strategies for GMPPB-Associated Dystroglycanopathy I

Recent studies have highlighted a significant association between mutations in GDP-mannose pyrophosphorylase B (GMPPB) gene with dystroglycanopathy (DGP), a rare disease characterized by neuromuscular phenotypes. To interrogate the molecular mechanisms driving the onset and progression and to identify potential therapeutic approaches for GMPPB-associated DGP, we here constructed genetically engineered mice models of Gmppb. We show that Gmppb knockout and P32L mutant mice are lethal in the homozygous form, whereas homologous R287Q mutants are viable. Heterozygous Gmppb-P32L mutant mice exhibit reduced muscle strength, decreased locomotor ability, elevated creatine kinase levels and increased centrally nucleated myofibers. Furthermore, loss of GMPPB results in defective differentiation of muscle stem cells, leading to failure to develop into mature myotubes and diminished muscle regeneration capability. Biochemical and transcriptomic analyses indicate that loss of GMPPB is associated with significant alterations in O-linked glycosylation level, intracellular Ca2+ storage and release and the Wnt/β-catenin signaling pathway. Pharmacological activation of the Wnt pathway alleviated disruption of muscle differentiation and regeneration post muscle injury in Gmppb-deficient models. Additionally, adeno-associated virus (AAV)-mediated gene replacement therapy successfully ameliorated the muscular phenotypes. Collectively, our findings provide direct evidence that impaired muscle stem cell differentiation contributes to GMPPB-associated dystroglycanopathy. Wnt pathway agonists and AAV gene therapy represent potential effective intervention strategies for treating DGP disease. To investigate the influence of GMPPB on muscle development, we created three genetically engineered mouse models: GMPPB knockout mice, GMPPB-P32L mice, and GMPPB-R287Q mice. Subsequently, we performed bioinformatics analysis on the transcriptome data obtained from muscle tissues of both GMPPB mutant mice and wild-type mice.

近年来的研究表明，GDP-甘露糖焦磷酸化酶B（GDP-mannose pyrophosphorylase B, GMPPB）基因突变与肌营养不良蛋白聚糖病（dystroglycanopathy, DGP）存在显著关联——后者是一类以神经肌肉表型为特征的罕见疾病。为探究驱动该疾病发生与进展的分子机制，并挖掘GMPPB相关性DGP的潜在治疗策略，本研究构建了Gmppb基因工程小鼠模型。
研究结果显示，纯合子Gmppb敲除小鼠及P32L突变小鼠均致死，而纯合子R287Q突变小鼠可正常存活。杂合子Gmppb-P32L突变小鼠表现出肌肉力量下降、运动能力降低、肌酸激酶水平升高以及中央核肌纤维增多的表型。
进一步研究发现，GMPPB缺失会导致肌肉干细胞分化缺陷，使其无法发育为成熟肌管，并削弱肌肉再生能力。生化与转录组分析表明，GMPPB缺失会显著改变O-连接糖基化水平、细胞内钙储存与释放功能，以及Wnt/β-连环蛋白信号通路（Wnt/β-catenin signaling pathway）。
对Wnt通路进行药理学激活，可缓解Gmppb缺陷模型中肌肉损伤后的肌肉分化与再生障碍。此外，腺相关病毒（adeno-associated virus, AAV）介导的基因替换疗法成功改善了该疾病的肌肉表型。
综上，本研究的发现直接证实，肌肉干细胞分化受损是GMPPB相关性肌营养不良蛋白聚糖病的核心致病机制之一。Wnt通路激动剂与AAV基因疗法有望成为治疗DGP的有效干预手段。
为探究GMPPB对肌肉发育的影响，本研究构建了三类基因工程小鼠模型：GMPPB敲除小鼠、GMPPB-P32L突变小鼠及GMPPB-R287Q突变小鼠。随后，我们对取自GMPPB突变小鼠与野生型小鼠肌肉组织的转录组数据进行了生物信息学分析。