Pilot study design.

Fabry disease (FD) is an X-linked disorder of glycosphingolipid metabolism caused by mutations in the GLA gene encoding alpha-galactosidase A (α-Gal). Loss of α-Gal activity leads to progressive lysosomal accumulation of α-Gal substrate, predominately globotriaosylceramide (Gb3) and its deacylated derivative globotriaosylsphingosine (lyso‐Gb3). FD manifestations include early onset neuropathic pain, gastrointestinal symptoms, and later onset life-threatening renal, cardiovascular and cerebrovascular disorders. Current treatments can preserve kidney function but are not very effective in preventing progression of cardiovascular pathology which remains the most common cause of premature death in FD patients. There is a significant need for a translational model that could be used for testing cardiac efficacy of new drugs. Two mouse models of FD have been developed. The α-Gal A-knockout (GlaKO) model is characterized by progressive tissue accumulation of Gb3 and lyso-Gb3 but does not develop any Fabry pathology besides mild peripheral neuropathy. Reports of minor cardiac function abnormalities in GlaKO model are inconsistent between different studies. Recently, G3Stg/GlaKO was generated by crossbreeding GlaKO with transgenic mice expressing human Gb3 synthase. G3Stg/GlaKO demonstrate higher tissue substrate accumulation and develop cellular and tissue pathologies. Functional renal pathology analogous to that found in early stages of FD has also been described in this model. The objective of this study is to characterize cardiac phenotype in GlaKO and G3Stg/GlaKO mice using echocardiography. Longitudinal assessments of cardiac wall thickness, mass and function were performed in GlaKO and wild-type (WT) littermate controls from 5–13 months of age. G3Stg/GlaKO and WT mice were assessed between 27–28 weeks of age due to their shortened lifespan. Several cardiomyopathy characteristics of early Fabry pathology were found in GlaKO mice, including mild cardiomegaly [up-to-25% increase in left ventricular (LV mass)] with no significant LV wall thickening. The LV internal diameter was significantly wider (up-to-24% increase at 9-months), when compared to the age-matched WT. In addition, there were significant increases in the end-systolic, end-diastolic volumes and stroke volume, suggesting volume overload. Significant reduction in Global longitudinal strain (GLS) measuring local myofiber contractility of the LV was also detected at 13-months. Similar GLS reduction was also reported in FD patients. Parameters such as ejection fraction, fractional shortening and cardiac output were either only slightly affected or were not different from controls. On the other hand, some of the cardiac findings in G3Stg/GlaKO mice were inconsistent with Fabry cardiomyopathy seen in FD patients. This could be potentially an artifact of the Gb3 synthase overexpression under a strong ubiquitous promoter. In conclusion, GlaKO mouse model presents mild cardiomegaly, mild cardiac dysfunction, but significant cardiac volume overload and functional changes in GLS that can be used as translational biomarkers to determine cardiac efficacy of novel treatment modalities. The level of tissue Gb3 accumulation in G3Stg/GlaKO mouse more closely recapitulates the level of substrate accumulation in FD patients and may provide better translatability of the efficacy of new therapeutics in clearing pathological substrates from cardiac tissues. But interpretation of the effect of treatment on cardiac structure and function in this model should be approached with caution.

法布里病（Fabry disease, FD）是一种由编码α-半乳糖苷酶A（alpha-galactosidase A, α-Gal）的GLA基因发生突变所引发的X连锁糖鞘脂代谢紊乱性疾病。α-Gal活性缺失会导致α-Gal底物进行性溶酶体蓄积，主要底物为神经三己糖酰基鞘氨醇（globotriaosylceramide, Gb3）及其脱酰基衍生物脱酰基神经三己糖鞘氨醇（globotriaosylsphingosine, lyso‐Gb3）。FD的临床表现包括早发性神经性疼痛、胃肠道症状，以及晚发性的危及生命的肾、心血管及脑血管病变。现有治疗手段可保护肾功能，但在阻止心血管病变进展方面效果有限，而心血管病变仍是FD患者过早死亡的最常见原因。目前亟需可用于评估新药心脏疗效的转化模型。目前已开发出两种FD小鼠模型。α-半乳糖苷酶A基因敲除（α-Gal A-knockout, GlaKO）模型以Gb3与lyso‐Gb3进行性组织蓄积为特征，但除轻度周围神经病外，不会出现任何法布里病相关病理表现，不同研究关于GlaKO模型存在轻微心脏功能异常的报道并不一致。近期，研究人员通过将GlaKO小鼠与表达人Gb3合酶的转基因小鼠杂交繁育，构建了G3Stg/GlaKO模型，该模型的组织底物蓄积水平更高，且会出现细胞及组织病理改变，此外该模型中还出现了与FD早期阶段类似的功能性肾脏病理表现。本研究的目的为利用超声心动图对GlaKO与G3Stg/GlaKO小鼠的心脏表型进行表征。研究人员对5~13月龄的GlaKO小鼠及野生型（wild-type, WT）同窝对照小鼠进行了心脏壁厚度、质量及功能的纵向评估。由于G3Stg/GlaKO小鼠寿命较短，对其的评估在27~28周龄时完成。研究在GlaKO小鼠中发现了早期FD病理的多项心肌病特征，包括轻度心脏肥大[左心室（left ventricular, LV）质量最多增加25%]，但无明显左心室壁增厚。与同年龄匹配的WT小鼠相比，GlaKO小鼠的左心室内径显著增宽，9月龄时最多增加24%。此外，其收缩末期、舒张末期容积及每搏输出量均显著升高，提示存在容量负荷过重。13月龄时还检测到左心室肌纤维收缩力指标整体纵向应变（Global longitudinal strain, GLS）显著降低，该表型在FD患者中也已有报道。射血分数、短轴缩短率及心输出量等参数仅受轻微影响，或与对照组无显著差异。另一方面，G3Stg/GlaKO小鼠的部分心脏表现与FD患者的法布里心肌病并不相符，这可能是在强泛启动子驱动下Gb3合酶过表达所导致的潜在人为假象。综上，GlaKO小鼠模型存在轻度心脏肥大、轻度心脏功能异常，但同时存在显著的心脏容量负荷过重及GLS功能改变，这些可作为评估新型治疗手段心脏疗效的转化生物标志物。G3Stg/GlaKO小鼠的组织Gb3蓄积水平更贴近FD患者的底物蓄积水平，可更好地反映新型治疗手段清除心脏组织病理底物的疗效，但在解释该模型中心脏结构与功能的治疗效应时需谨慎。