医学
肺移植
移植
荧光素酶
肺
生物发光成像
免疫组织化学
转基因
基因传递
病理
气道
遗传增强
腺相关病毒
男科
生物
重组DNA
内科学
转染
外科
基因
载体(分子生物学)
生物化学
作者
Samuel J. Kesseli,Madison Krischak,Qimeng Gao,Trevor J. Gonzalez,Min Zhang,Samantha E. Halpern,Riley Kahan,Mingqing Song,Niki Huffman,Huimin Xu,Nader Abraham,Aravind Asokan,Andrew S. Barbas,Matthew G. Hartwig
标识
DOI:10.1016/j.jtcvs.2022.08.050
摘要
Objective Adeno-associated virus is a clinically used gene therapy vector but has not been studied in lung transplantation. We sought to determine the efficacy of adeno-associated virus delivery during static cold storage via the airway versus the pulmonary artery before lung transplantation in a rodent model. Methods Lewis rat lung grafts were treated with a dose of 8e8 or 4e9 viral genome/μL recombinant adeno-associated virus subtype-9 vectors containing firefly luciferase genomes administered via the pulmonary artery or airway during cold storage. A control group did not receive adeno-associated virus. Recipient syngeneic rats then underwent single left lung transplantation. Animals underwent bioluminescence imaging on postoperative days 7, 14, 28, and 56. Explanted tissues were prepared as lysates to quantify luciferase activity. Immunohistochemistry was performed to evaluate cellular transgene expression patterns. Results Control animals with no luminescent signal produced a background radiance of 6.1e4 p/s/cm2/sr. In the airway delivery group, mean radiance was greater than the control at 4e9 viral genome/μL postoperative day 7 radiance 6.9e4 p/s/cm2/sr (P = .04). In the pulmonary artery delivery group, we observed greater in vivo luminescence in animals receiving 4e9 viral genome/μL compared with all other groups. However, analysis of tissue lysate revealed greater luminescence in the airway delivery group and suggested off-target expression in heart and liver tissue in the pulmonary artery delivery group. Immunohistochemistry demonstrated transgene staining in distal airway epithelium and alveoli but sparing of the vasculature in the airway delivery group. Conclusions Adeno-associated virus mediates gene transduction during static cold storage in rat lung isografts when administered via the airway and pulmonary artery. Airway administration leads to robust transgene expression in respiratory epithelial cells, whereas pulmonary artery administration targets alternative cell types and increases extrapulmonary transgene expression.
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