A novel gene therapy for ARPKD based on CFTR.

IF 3.3 3区 医学 Q1 GASTROENTEROLOGY & HEPATOLOGY
Cristian Ciobanu, Patricia Outeda, William B Guggino, Liudmila Cebotaru
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引用次数: 0

Abstract

Autosomal recessive polycystic kidney disease (ARPKD) is associated with cysts derived from abnormal bile ducts. We focused on targeting the cysts and show that a gene therapy for ARPKD that targets the abnormal bile ducts is feasible. We injected 1-mo-old, Pkhd1del3-4/del3-4 mice intraperitoneally with 2 × 1012 particles/kg of adeno-associated virus (AAV1) containing either a GFP vector or a truncated cystic fibrosis transmembrane conductance regulator (CFTR) vector, Δ27-264-CFTR, or left them untreated. Two months after treatment, the cyst area and size in the liver were lower in the CFTR vector-treated mice than in mice receiving the GFP vector. We detected vector genomes and mRNA expression only in mice receiving the corresponding CFTR or GFP vector. We observed abundant GFP immunofluorescence in the cholangiocytes of the cysts and also saw expression of GFP and CFTR proteins above background levels in the corresponding treated mice. CFTR immunofluorescence was predominantly apically located in the ARPKD cholangiocytes, but after CFTR vector installation, it increased in the basolateral membrane. We stained mouse livers with Maackia amurensis lectin (MAL) or Sambucus nigra lectin (SNA), specific for α2,3- and α2,6-N-linked sialic acid, respectively, to detect the presence of sialic acid moieties contributing to AAV1 binding. Although immunofluorescent SNA was detected in the wild-type bile ducts, MAL 1 was not. MAL immunofluorescence was present in remarkably high levels on the apical surfaces of the cysts in cholangiocytes, offering a good target for AAV gene therapy. A gene therapy using an AAV1-based vector containing a truncated CFTR could be therapeutic in ARPKD.NEW & NOTEWORTHY Autosomal recessive polycystic kidney disease (ARPKD) causes severe disease in babies in the womb. Those who survive the neonatal period face chronic kidney and liver disease throughout their life. The overall goal of our study here is to develop a gene therapy to treat ARPKD.

一种基于CFTR的ARPKD基因治疗新方法。
常染色体隐性多囊肾病与异常胆管囊肿有关。我们的重点是针对囊肿,并表明针对异常胆管的ARPKD基因治疗是可行的。我们给1个月大的Pkhd1del3-4/del3-4小鼠腹腔注射2 × 1012颗粒/kg的AAV1, AAV1含有GFP载体或截断的CFTR载体Δ27-264-CFTR,或者不进行治疗。治疗2个月后,CFTR载体处理的小鼠肝脏囊肿面积和大小低于GFP载体处理的小鼠。我们仅在接受相应CFTR或GFP载体的小鼠中检测载体基因组和mRNA表达。我们在囊肿胆管细胞中观察到丰富的GFP免疫荧光,并且在相应处理的小鼠中GFP和CFTR蛋白的表达高于背景水平。CFTR免疫荧光主要位于ARPKD胆管细胞的顶端,但在CFTR载体安装后,其在基底外侧膜上的荧光增加。我们分别用α2,3-和α2,6- n -链唾液酸特异性的黑黑树胶凝集素(SNA)和黑树胶凝集素(MAL)对小鼠肝脏进行染色,检测与AAV1结合相关的唾液酸片段的存在。尽管在野生型胆管中检测到免疫荧光SNA,但未检测到MAL 1。MAL免疫荧光在胆管细胞囊肿的顶端表面表现出显著的高水平,为AAV基因治疗提供了一个很好的靶点。结论:利用含有截断CFTR的aav1载体对ARPKD进行基因治疗是可行的。
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来源期刊
CiteScore
9.40
自引率
2.20%
发文量
104
审稿时长
1 months
期刊介绍: The American Journal of Physiology-Gastrointestinal and Liver Physiology publishes original articles pertaining to all aspects of research involving normal or abnormal function of the gastrointestinal tract, hepatobiliary system, and pancreas. Authors are encouraged to submit manuscripts dealing with growth and development, digestion, secretion, absorption, metabolism, and motility relative to these organs, as well as research reports dealing with immune and inflammatory processes and with neural, endocrine, and circulatory control mechanisms that affect these organs.
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