PJSC Human Stem Cells Institute, Moscow, Russia.
LLC Genotarget, Skolkovo Innovation Center, Moscow, Russia.
Bull Exp Biol Med. 2023 Apr;174(6):768-773. doi: 10.1007/s10517-023-05789-z. Epub 2023 May 9.
We studied the effects of a dual-vector DYSF gene delivery system based on adeno-associated virus serotype 9 capsids on pathological manifestations of dysferlinopathy in skeletal muscles of Bla/J mice lacking DYSF expression. The mice received intravenous injection of 3×10 genomic copies of the virus containing the dual-vector system. M. gastrocnemius, m. psoas major, m. vastus lateralis, and m. gluteus superficialis were isolated for histological examination in 3, 6, and 12 weeks after treatment. Healthy wild-type (C57BL/6) mice served as positive control and were sacrificed 3 weeks after injection of 150 μl of 0.9% NaCl into the caudal vein. To detect dysferlin in muscle cryosections, immunohistochemical analysis with diagnostic antibodies was performed; paraffin sections were stained with hematoxylin and eosin for morphometric analysis. After administration of gene-therapeutic constructs, muscle fibers with membrane or cytoplasmic dysferlin location were detected in all examined muscles. The proportion of necrotic muscle fibers decreased, the number of muscle fibers with central location of the nucleus increased, and the mean cross-section area of the muscle fibers decreased.
我们研究了基于腺相关病毒血清型 9 衣壳的双载体 DYSF 基因传递系统对缺乏 DYSF 表达的 Bla/J 小鼠骨骼肌中肌营养不良蛋白病病理表现的影响。这些小鼠接受了含有双载体系统的 3×10 基因组拷贝病毒的静脉注射。在治疗后 3、6 和 12 周，分离 M. gastrocnemius、m. psoas major、m. vastus lateralis 和 m. gluteus superficialis 进行组织学检查。健康的野生型（C57BL/6）小鼠作为阳性对照，在尾静脉注射 150μl 0.9%NaCl 后 3 周处死。为了在肌肉冷冻切片中检测肌营养不良蛋白，使用诊断抗体进行免疫组织化学分析；对石蜡切片进行苏木精和伊红染色进行形态计量学分析。在给予基因治疗构建体后，在所有检查的肌肉中均检测到具有膜或细胞质肌营养不良蛋白定位的肌纤维。坏死肌纤维的比例减少，核中央定位的肌纤维数量增加，肌纤维的平均横截面积减小。
