A novel transgenic mouse model expressing mutant TTR gene causing oculoleptomeningeal type of ATTRm amyloidosis
A novel transgenic mouse model expressing mutant TTR gene causing oculoleptomeningeal type of ATTRm amyloidosis
复制标题
一种表达突变TTR基因的新型转基因小鼠模型，引起眼脑膜型ATTRm淀粉样变性
DOI：
10.1080/13506129.2019.1583190
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发表时间：
2019
期刊：
影响因子：
5.5
通讯作者：
Ando Y.
中科院分区：
文献类型：
--
作者：
Isoguchi A.;Ueda M.;Kanenawa K.;Masuda T.;Misumi Y.;Yamashita T.;Obayashi K.;Ando Y.
背景遗传性转甲状腺素蛋白（ATTRm）淀粉样变性是一种由 TTR 突变引起的常染色体显性遗传性疾病，表现为全身器官衰竭[引文 1]。迄今为止，在 ATTRm 淀粉样变性患者中发现了超过 140 种 TTR 突变 [引文 2]，其中一些突变据报道导致眼脑膜型 ATTRm 淀粉样变性 [引文 1，引文 2]。眼部淀粉样蛋白沉积物主要来源于视网膜色素上皮（RPE）中产生的变体TTR。肝移植 (LT) 已作为 ATTRm 淀粉样变性的治疗方法进行。然而，即使在 LT 后，眼部症状也经常进展 [引文 3]，因为 LT 后 RPE 中继续产生变异 TTR。在本研究中，我们构建了一种表达人类 ATTR Tyr114Cys 基因的新型转基因小鼠模型，该基因会导致人类软脑膜淀粉样变性[引文 4，引文 5]，并研究了该模型小鼠的组织病理学和生化特征。材料和方法将含有具有 Tyr114Cys 突变的 TTR 基因的人类基因组 DNA 显微注射到 C57BL/6 胚胎中。尾部 DNA 用于确认转基因。使用从小鼠每个组织部位分离的 RNA，通过实时 PCR 分析人 ATTR Tyr114Cys 基因表达。使用抗人 TTR 抗体通过免疫组织化学检查每个组织部位的人 ATTR Tyr114Cys 表达和沉积。还通过免疫印迹分析了血浆、脑脊液 (CSF) 和房水中的人 ATTR。刚果红染色检测小鼠组织样本中的淀粉样蛋白沉积。结果人ATTR Tyr114Cys基因在肝脏、大脑脉络丛、视网膜和睫状体中表达。免疫组织化学染色还显示，人 ATTR Tyr114Cys 在肝脏、脉络丛和朗格汉斯胰岛中表达，并且人 ATTR Tyr114Cys 在多个器官中积累。那些人类 ATTR 沉积物不会与刚果红发生反应。免疫印迹分析表明血浆、脑脊液和房水中含有人ATTR Tyr114Cys。讨论和结论我们发现我们的新型TG小鼠除了肝脏和大脑外，还在眼睛中表达人ATTR Tyr114Cys。为了调查人类 ATTR Tyr114Cys 是否形成淀粉样蛋白沉积并导致眼部功能障碍，还需要进一步的研究。总之，我们表达 ATTR Tyr114Cys 的新型转基因小鼠可能是眼脑膜 ATTRm 淀粉样变性的有用模型。
BackgroundHereditary transthyretin (ATTRm) amyloidosis is an autosomal dominant hereditary disease caused by TTR mutations and shows systemic organ failures [Citation 1]. To date, more than 140 kinds of TTR mutations were identified in ATTRm amyloidosis patients [Citation 2] and some of those mutations reportedly caused oculoleptomeningeal type of ATTRm amyloidosis [Citation 1, Citation 2]. Ocular amyloid deposits are mainly derived from variant TTR produced in the retinal pigment epithelium (RPE). Liver transplantation (LT) has been performed as a treatment for ATTRm amyloidosis. However, ocular symptoms often progress even after LT [Citation 3] because variant TTR continues to be produced in RPE after LT. In this study, we generated a novel transgenic mouse model expressing human ATTR Tyr114Cys gene which causes leptomeningeal amyloidosis in human [Citation 4, Citation 5] and investigated histopathological and biochemical features of this model mouse.Materials and methodsGenomic human DNA containing the TTR gene with a Tyr114Cys mutation was microinjected into C57BL/6 embryos. Tail DNA was used to confirm the transgene. Human ATTR Tyr114Cys gene expression was analysed by real-time PCR using RNA isolated from each tissue site of the mice. Human ATTR Tyr114Cys expression and deposition were examined in each tissue site by immunohistochemistry with an anti-human TTR antibody. Human ATTR was also analysed in plasma, cerebrospinal fluid (CSF) and aqueous humour by immunoblotting. Congo red staining was performed to detect amyloid deposits in tissue samples of the mice.ResultsGene expression of the human ATTR Tyr114Cys was demonstrated in liver, choroid plexus of the brain, retina and the ciliary body. Immunohistochemical staining also revealed that human ATTR Tyr114Cys was expressed in liver, choroid plexus and pancreatic islets of Langerhans and human ATTR Tyr114Cys accumulated in several organs. Those human ATTR deposits did not react with Congo red. Immunoblotting analyses indicated that human ATTR Tyr114Cys was contained in plasma, CSF and aqueous humour.Discussion and conclusionsWe found that our novel TG mice expressed human ATTR Tyr114Cys in the eyes in addition to liver and brain. To investigate whether human ATTR Tyr114Cys forms amyloid deposits and causes ocular dysfunctions, further studies are needed. In conclusion, our novel type of transgenic mouse expressing ATTR Tyr114Cys may be a useful model of oculoleptomeningeal ATTRm amyloidosis.