Genetic context modulates aging and degeneration in the murine retina

Background Age is the principal risk factor for neurodegeneration in both the retina and brain. The retina and brain share many biological properties; thus, insights into retinal aging and degeneration may shed light onto similar processes in the brain. Genetic makeup strongly influences susceptibility to age-related retinal disease. However, studies investigating retinal aging have not sufficiently accounted for genetic diversity. Therefore, examining molecular aging in the retina across different genetic backgrounds will enhance our understanding of human-relevant aging and degeneration in both the retina and brain—potentially improving therapeutic approaches to these debilitating conditions. Methods Transcriptomics and proteomics were employed to elucidate retinal aging signatures in nine genetically diverse mouse strains (C57BL/6J, 129S1/SvlmJ, NZO/HlLtJ, WSB/EiJ, CAST/EiJ, PWK/PhK, NOD/ShiLtJ, A/J, and BALB/cJ) across lifespan. These data predicted human disease-relevant changes in WSB and NZO strains. Accordingly, B6, WSB and NZO mice were subjected to human-relevant in vivo examinations at 4, 8, 12, and/or 18M, including: slit lamp, fundus imaging, optical coherence tomography, fluorescein angiography, and pattern/full-field electroretinography. Retinal morphology, vascular structure, and cell counts were assessed ex vivo. Results We identified common molecular aging signatures across the nine mouse strains, which included genes associated with photoreceptor function and immune activation. Genetic background strongly modulated these aging signatures. Analysis of cell type-specific marker genes predicted age-related loss of photoreceptors and retinal ganglion cells (RGCs) in WSB and NZO, respectively. Fundus exams revealed retinitis pigmentosa-relevant pigmentary abnormalities in WSB retinas and diabetic retinopathy (DR)-relevant cotton wool spots and exudates in NZO retinas. Profound photoreceptor dysfunction and loss were confirmed in WSB. Molecular analyses indicated changes in photoreceptor-specific proteins prior to loss, suggesting photoreceptor-intrinsic dysfunction in WSB. In addition, age-associated RGC dysfunction, loss, and concomitant microvascular dysfunction was observed in NZO mice. Proteomic analyses revealed an early reduction in protective antioxidant processes, which may underlie increased susceptibility to DR-relevant pathology in NZO. Conclusions Genetic context is a strong determinant of retinal aging, and our multi-omics resource can aid in understanding age-related diseases of the eye and brain. Our investigations identified and validated WSB and NZO mice as improved preclinical models relevant to common retinal neurodegenerative diseases. Overall design: 1/2 Retinas were processed for RNA-sequencing from 9 strains of mice, of both sexes and at 3 ages. We used the resulting data to identify strain-, age- and strain-by-age effects across the samples.

背景：年龄是视网膜与大脑神经退行性病变的首要风险因素。视网膜与大脑共享诸多生物学特性；因此，解析视网膜衰老与退行性病变的相关机制，或可为大脑中类似的病理过程提供研究思路。遗传构成对年龄相关性视网膜疾病的易感性具有显著影响。然而，现有关于视网膜衰老的研究尚未充分考虑遗传多样性。因此，探究不同遗传背景下视网膜的分子衰老特征，将有助于加深我们对视网膜与大脑中与人类相关的衰老及退行性病变的理解，或可改善这类致残性疾病的治疗策略。

方法：研究采用转录组学（transcriptomics）与蛋白质组学（proteomics）技术，解析了9种遗传多样性小鼠品系（C57BL/6J、129S1/SvlmJ、NZO/HlLtJ、WSB/EiJ、CAST/EiJ、PWK/PhK、NOD/ShiLtJ、A/J及BALB/cJ）在整个生命周期中的视网膜衰老特征。上述数据预测了WSB与NZO品系中与人类疾病相关的分子变化。据此，我们对B6、WSB及NZO小鼠在4、8、12及/或18月龄时开展了符合人类临床标准的体内检测，包括：裂隙灯检查、眼底成像、光学相干断层扫描（optical coherence tomography）、荧光素眼底血管造影（fluorescein angiography）以及图形/全视野视网膜电图（electroretinography）。此外，还对视网膜形态、血管结构以及细胞计数进行了离体评估。

结果：我们在9种小鼠品系中鉴定出了共通的分子衰老特征，其中包括与感光细胞功能及免疫激活相关的基因。遗传背景对这些衰老特征具有显著的调控作用。对细胞类型特异性标记基因的分析预测，WSB与NZO品系小鼠分别会出现感光细胞与视网膜神经节细胞（retinal ganglion cells, RGCs）的年龄相关性丢失。眼底检查显示，WSB小鼠视网膜出现了与色素性视网膜炎相关的色素异常，而NZO小鼠视网膜则出现了与糖尿病视网膜病变（diabetic retinopathy, DR）相关的棉絮斑及渗出物。我们在WSB小鼠中验证了严重的感光细胞功能异常与丢失。分子分析表明，在感光细胞丢失前，感光细胞特异性蛋白即已出现表达变化，提示WSB小鼠存在感光细胞内在的功能异常。此外，在NZO小鼠中还观察到了年龄相关性RGC功能异常、丢失以及伴随的微血管功能障碍。蛋白质组学分析显示，保护性抗氧化通路出现早期下调，这或为NZO小鼠易患DR相关病理改变的潜在机制。

结论：遗传背景是决定视网膜衰老进程的关键因素，本研究构建的多组学资源将有助于理解眼部与大脑的年龄相关性疾病。我们的研究鉴定并验证了WSB与NZO小鼠可作为与常见视网膜神经退行性疾病相关的更优临床前模型（preclinical models）。

整体实验设计：从9种小鼠品系（涵盖雌雄两性及3个年龄阶段）中采集的半数视网膜样本被用于RNA测序（RNA-sequencing），我们利用所得数据鉴定了样本中品系、年龄以及品系-年龄交互作用带来的差异效应。