Zhang Yan, Wei Wen-Bin, Zhao Junyang, Xu Xiaolin, Wang Fufeng
Department of Ophthalmology, Beijing Children's Hospital, Capital Medical University, National Center for Children's Health, Beijing, China.
Beijing Tongren Eye Center, Beijing Key Laboratory of Intraocular Tumor Diagnosis and Treatment, Beijing Tongren Hospital, Capital Medical University, Beijing, China.
Ophthalmic Genet. 2022 Dec;43(6):795-805. doi: 10.1080/13816810.2022.2098985. Epub 2022 Aug 8.
To characterize the spectrum of mosaic RB1 pathogenic alleles and map the distribution of mutant cells in available tissues from mosaic patients. Next-generation sequencing was performed on blood samples from 263 retinoblastoma families to identify mosaic RB1 variant alleles. A variety of available tissues were sampled to determine tissue distribution and fraction of mutant cells in five mosaic patients who consented to participate in mosaic pathogenic allele research. Twelve identified mosaic RB1 variants were all "null" pathogenic alleles and displayed reduced expressivity. The use of next-generation deep sequencing increased the sensitivity of mosaicism detection to 0.03% in the case of tissue DNA. In the five mosaic participants, we observed coherent but uneven, bilateral asymmetrical distribution of mutant cells across various tissues. They all carried early-embryonic mosaic pathogenic alleles and had significantly higher variant fractions in blood than in other tissues. Variant fractions of ipsilateral tissue samples were not concordant higher or lower compared with the contralateral side. Only ipsilateral conjunctival and oral epithelial cells showed concordance in mosaicism levels. No associations were observed between the laterality of affected eyes and variant fractions of any tissue type. NGS allows the detection of low-level mosaicism. Mosaic RB1 pathogenic alleles are prone to occur at very early stages of human embryonic development. With respect to genetic counseling, risk prediction should take into account unrecognized mosaicism. The underlying tissue distribution patterns of mosaic RB1 variant alleles remain to be determined.
为了表征镶嵌型RB1致病等位基因的谱系，并绘制镶嵌型患者可用组织中突变细胞的分布图。对263个视网膜母细胞瘤家族的血样进行了下一代测序，以鉴定镶嵌型RB1变异等位基因。对五名同意参与镶嵌型致病等位基因研究的患者的各种可用组织进行采样，以确定组织分布和突变细胞比例。鉴定出的12个镶嵌型RB1变异均为“无效”致病等位基因，且表现出表达降低。对于组织DNA，使用下一代深度测序将镶嵌现象检测的灵敏度提高到了0.03%。在这五名镶嵌型参与者中，我们观察到突变细胞在各种组织中呈现连贯但不均匀、双侧不对称的分布。他们都携带早期胚胎镶嵌型致病等位基因，血液中的变异比例明显高于其他组织。同侧组织样本的变异比例与对侧相比并无一致的高低之分。只有同侧结膜和口腔上皮细胞在镶嵌程度上表现出一致性。未观察到患眼的侧别与任何组织类型的变异比例之间存在关联。二代测序可检测低水平的镶嵌现象。镶嵌型RB1致病等位基因易于在人类胚胎发育的非常早期阶段出现。在遗传咨询方面，风险预测应考虑未被识别的镶嵌现象。镶嵌型RB1变异等位基因的潜在组织分布模式仍有待确定。
