Neuronal Ceroid Lipofuscinosis (NCL) is caused by the entire deletion of CLN8 in the Alpine Dachsbracke. AD009

Neuronal ceroid lipofuscinosis (NCL) is an inherited lysosomal storage disease that has been described in a variety of dog breeds, where it is caused by different mutations. However, the causative gene defect in the Alpine Dachsbracke has not been known so far. Here we present two confirmed cases of NCL from different litters of the same sire with a different dam showing the same underlying gene mutation. Case 1, a 2-year-old male Alpine Dachsbracke was presented with sudden blindness, disorientation, reduction of food intake and character changes such as anxiety states and aggressiveness. Clinical examination revealed moderate obtundation, posttectal blindness and ventral strabism. MRI scans showed dilation of all cerebral ventricles, thinning of the intermediate mass of the thalamus and widening of the cerebral sulci consistent with general cerebral atrophy. Liquor cytology did not reveal any pathological findings. NCL, a hereditary lysosomal storage disease was suspected and confirmed post mortem. Microscopic examination of case 1 revealed granular, eosinophilic, autofluorescent, PAS- and Sudanblack B-positive material in neurons of the central and enteric nervous system as well as in macrophages within spleen, lymphnodes and lung. The storage bodies labelled negative with sphingolipid activator protein D and subunit c of the mitochondrial ATP synthase. Ultrastructurally the material presented as osmiophilic depositions. Case 2, a XX-year old female Alpine Dachsbracke was presented with … Microscopic examination of the brain of case 2 revealed the same granular, eosinophilic, autofluorescent, PAS- and Sudanblack B-positive material in neurons. Whole genome sequencing of the affected dogs revealed a homozygous deletion encompassing the entire CLN8 gene representing the most likely causative mutation for the observed NCL form in both cases. The deletion follows recessive inheritance since the dam and a male littermate of case 1 were tested as heterozygous carriers. This is the first detailed description of CLN8 associated NCL in Alpine Dachsbracke dogs, which provides a novel canine CLN8 model leading to this lysosomal storage disease.

神经蜡样脂褐质沉积症（Neuronal ceroid lipofuscinosis, NCL）是一种遗传性溶酶体贮积病，已在多个犬种中被报道，其致病机制因品种而异。然而，迄今阿尔派达克什布雷克犬（Alpine Dachsbracke）的致病基因缺陷尚未明确。本研究报道两例确诊的NCL病例，均来自同一公犬与不同母犬的不同窝次，二者携带相同的致病基因突变。

病例1为一只2岁雄性阿尔派达克什布雷克犬，因突发失明、定向障碍、食欲减退及焦虑、攻击性增强等性格改变就诊。临床检查可见中度意识迟钝、皮质盲与腹侧斜视。磁共振成像（Magnetic Resonance Imaging, MRI）扫描显示全脑室扩张、丘脑中间块变薄及脑沟增宽，符合全脑萎缩的影像学特征。脑脊液细胞学检查未发现异常病理改变。临床怀疑为遗传性溶酶体贮积病NCL，并经死后剖检确诊。

病例1的显微镜检查结果显示，中枢神经系统与肠神经系统的神经元，以及脾脏、淋巴结和肺内的巨噬细胞中，均存在颗粒状、嗜酸性、自发荧光的过碘酸-雪夫（Periodic Acid-Schiff, PAS）染色及苏丹黑B（Sudanblack B）阳性物质。该贮积物对鞘脂激活蛋白D（sphingolipid activator protein D）与线粒体ATP合酶（mitochondrial ATP synthase）亚基c呈阴性标记。超微结构观察可见该物质为嗜锇性沉积。

病例2为一只XX岁雌性阿尔派达克什布雷克犬，临床表现为……。病例2的脑组织显微镜检查可见神经元内存在与病例1一致的颗粒状、嗜酸性、自发荧光的PAS染色及苏丹黑B阳性贮积物。

对患病犬进行全基因组测序后发现，存在一段覆盖整个CLN8基因的纯合缺失，该变异极有可能是两例NCL的致病突变。该缺失遵循隐性遗传模式，病例1的母犬与一只同窝雄性携带者经检测均为杂合子。

本研究首次详细报道了阿尔派达克什布雷克犬中与CLN8相关的NCL病例，为该溶酶体贮积病构建了新型犬类CLN8疾病模型。