Abstract：
BACKGROUND: Previously, we discovered a strain of Kunming mice, referred to as the KMush/ush strain, that exhibited notably abnormal electroretinogram (ERG) readings and elevated thresholds for auditory brainstem responses (ABRs), which resembled the characteristics of Usher Syndrome (USH). We successfully identified the pathogenic genes, Pde6b and Adgrv1, after KMush/ush crossbred with CBA/CaJ mice, referred to as CBA-1ush/ush, CBA-2ush/ush or CBA-2ush/ush. In this investigation, we crossbred KMush/ush and CBA/J mice to establish novel recombinant inbred lines and analysed their phenotypic and genotypic characteristics.
METHODS: ERG readings, ABR testing, fundus morphology, histological examination of the retina and inner ear, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis, western blotting, DNA sequence analysis and behavioural experiments were performed to assess the phenotypes and genotypes of the progeny lines.
RESULTS: No obvious waveforms in the ERG were detected in F1 hybrid mice while normal ABR results were recorded. The F2 hybrids, which were called J1ush/ush or J2ush/ush, exhibited segregated hearing-loss phenotypes. J1ush/ush mice had a retinitis pigmentosa (RP) phenotype with elevated ABR thresholds, whereas J2ush/ush mice exhibited only the RP phenotype. Interestingly, J1ush/ush mice showed significantly higher ABR thresholds than wild-type mice at 28 days post born (P28), and RT-qPCR and DNA-sequencing analysis showed that Adgrv1 gene expression was significantly altered in J1ush/ush mice, but histological analysis showed no significant structural changes in the organ of Corti or spiral ganglia. Further elevation of ABR-related hearing thresholds by P56 manifested only as a reduced density of spiral ganglion cells, which differed significantly from the previous pattern of cochlear alterations in CBA-2ush/ush mice.
CONCLUSIONS: We successfully introduced the hearing-loss phenotype of inbred mice with USH into CBA/J mice, which provides a good animal model for future studies on the important physiological roles of the Adgrv1 gene in inner-ear structure and for therapeutic studies targeting Adgrv1-mutated USH.
METHODS: ERG readings, ABR testing, fundus morphology, histological examination of the retina and inner ear, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis, western blotting, DNA sequence analysis and behavioural experiments were performed to assess the phenotypes and genotypes of the progeny lines.
RESULTS: No obvious waveforms in the ERG were detected in F1 hybrid mice while normal ABR results were recorded. The F2 hybrids, which were called J1ush/ush or J2ush/ush, exhibited segregated hearing-loss phenotypes. J1ush/ush mice had a retinitis pigmentosa (RP) phenotype with elevated ABR thresholds, whereas J2ush/ush mice exhibited only the RP phenotype. Interestingly, J1ush/ush mice showed significantly higher ABR thresholds than wild-type mice at 28 days post born (P28), and RT-qPCR and DNA-sequencing analysis showed that Adgrv1 gene expression was significantly altered in J1ush/ush mice, but histological analysis showed no significant structural changes in the organ of Corti or spiral ganglia. Further elevation of ABR-related hearing thresholds by P56 manifested only as a reduced density of spiral ganglion cells, which differed significantly from the previous pattern of cochlear alterations in CBA-2ush/ush mice.
CONCLUSIONS: We successfully introduced the hearing-loss phenotype of inbred mice with USH into CBA/J mice, which provides a good animal model for future studies on the important physiological roles of the Adgrv1 gene in inner-ear structure and for therapeutic studies targeting Adgrv1-mutated USH.
摘要：
背景：以前，我们发现了一种昆明小鼠,称为KMush/Mush菌株，表现出明显异常的视网膜电图（ERG）读数和听觉脑干反应（ABR）阈值升高，类似于Usher综合征（USH）的特征。我们成功地鉴定了致病基因,Pde6b和Adgrv1，在KMush/ush与CBA/CaJ小鼠杂交后，称为CBA-1ush/ush，CBA-2ush/ush或CBA-2ush/ush。在这次调查中,我们杂交KMush/ush和CBA/J小鼠以建立新的重组自交系，并分析其表型和基因型特征。
方法：ERG读数，ABR测试,眼底形态学,视网膜和内耳的组织学检查，逆转录-定量聚合酶链反应（RT-qPCR）分析，西方印迹,进行DNA序列分析和行为实验以评估后代系的表型和基因型。
结果：在F1杂种小鼠的ERG中未检测到明显的波形，而记录到正常的ABR结果。F2杂种，它们被称为J1ush/ush或J2ush/ush，表现出隔离的听力损失表型。J1ush/ush小鼠具有视网膜色素变性（RP）表型，ABR阈值升高，而J2ush/ush小鼠仅表现出RP表型。有趣的是,J1ush/ush小鼠在出生后28天表现出明显高于野生型小鼠的ABR阈值（P28），RT-qPCR和DNA测序分析表明，J1ush/ush小鼠的Adgrv1基因表达显著改变，但是组织学分析显示Corti或螺旋神经节器官没有明显的结构变化。通过P56进一步提高ABR相关的听力阈值仅表现为螺旋神经节细胞的密度降低，与CBA-2ush/ush小鼠的耳蜗改变模式显着不同。
结论：我们成功地将USH近交系小鼠的听力损失表型引入CBA/J小鼠，这为未来研究Adgrv1基因在内耳结构中的重要生理作用以及针对Adgrv1突变USH的治疗研究提供了良好的动物模型。
方法：ERG读数，ABR测试,眼底形态学,视网膜和内耳的组织学检查，逆转录-定量聚合酶链反应（RT-qPCR）分析，西方印迹,进行DNA序列分析和行为实验以评估后代系的表型和基因型。
结果：在F1杂种小鼠的ERG中未检测到明显的波形，而记录到正常的ABR结果。F2杂种，它们被称为J1ush/ush或J2ush/ush，表现出隔离的听力损失表型。J1ush/ush小鼠具有视网膜色素变性（RP）表型，ABR阈值升高，而J2ush/ush小鼠仅表现出RP表型。有趣的是,J1ush/ush小鼠在出生后28天表现出明显高于野生型小鼠的ABR阈值（P28），RT-qPCR和DNA测序分析表明，J1ush/ush小鼠的Adgrv1基因表达显著改变，但是组织学分析显示Corti或螺旋神经节器官没有明显的结构变化。通过P56进一步提高ABR相关的听力阈值仅表现为螺旋神经节细胞的密度降低，与CBA-2ush/ush小鼠的耳蜗改变模式显着不同。
结论：我们成功地将USH近交系小鼠的听力损失表型引入CBA/J小鼠，这为未来研究Adgrv1基因在内耳结构中的重要生理作用以及针对Adgrv1突变USH的治疗研究提供了良好的动物模型。
