Yu Ze-Yan, McKay Karen, van Asperen Peter, Zheng Maolin, Fleming Jane, Ginn Samantha L, Kizana Eddy, Latham Margot, Feneley Michael P, Kirkland Peter D, Rowe Peter B, Lumbers Eugenie R, Alexander Ian E
Department of Respiratory Medicine, The Children's Hospital at Westmead, Westmead, Sydney, NSW, Australia.
J Gene Med. 2007 Jun;9(6):429-39. doi: 10.1002/jgm.1039.
Development of effective and durable gene therapy for treatment of the respiratory manifestations of cystic fibrosis remains a formidable challenge. Obstacles include difficulty in achieving efficient gene transfer to mature airway epithelium and the need to stably transduce self-renewing epithelial progenitor cells in order to avoid loss of transgene expression through epithelial turnover. Targeting the developing airway epithelium during fetal life offers the prospect of circumventing these challenges.
In the current study we investigated vesicular stomatitis virus glycoprotein (VSVg)-pseudotyped HIV-1-derived lentivirus vector-mediated gene transfer to the airway epithelium of mid-gestation fetal lambs, both in vitro and in vivo. In the in vitro studies epithelial sheet explants and lung organ culture were used to examine transduction of the proximal and more distal airway epithelium, respectively. For the in vivo studies, vector was delivered directly into the proximal airway.
We found that even during the early pseudoglandular and canalicular phases of lung development, occurring through mid-gestation, the proximal bronchial airway epithelium was relatively mature and highly resistant to lentivirus-mediated transduction. In contrast, the more distal bronchiolar airway epithelium was relatively permissive for transduction although the absolute levels achieved remained low.
This result is promising as the bronchiolar airway epithelium is a major site of pathology in the cystic fibrosis airway, and much higher levels of transduction are likely to be achieved by developing strategies that increase the amount of vector reaching the more distal airway after intratracheal delivery.
开发有效且持久的基因疗法用于治疗囊性纤维化的呼吸道表现仍然是一项艰巨的挑战。障碍包括难以实现向成熟气道上皮的高效基因转移，以及需要稳定转导自我更新的上皮祖细胞以避免因上皮更新而导致转基因表达丧失。在胎儿期靶向发育中的气道上皮提供了规避这些挑战的前景。
在本研究中，我们在体外和体内研究了水泡性口炎病毒糖蛋白（VSVg）假型化的源自HIV-1的慢病毒载体介导的基因转移至妊娠中期胎羊的气道上皮。在体外研究中，分别使用上皮片外植体和肺器官培养来检查近端和更远端气道上皮的转导情况。对于体内研究，将载体直接递送至近端气道。
我们发现，即使在肺发育的早期假腺泡期和小管期（直至妊娠中期），近端支气管气道上皮也相对成熟，并且对慢病毒介导的转导具有高度抗性。相比之下，更远端的细支气管气道上皮对转导相对敏感，尽管所达到的绝对水平仍然较低。
这一结果很有前景，因为细支气管气道上皮是囊性纤维化气道病变的主要部位，通过开发在气管内递送后增加到达更远端气道的载体量的策略，可能会实现更高水平的转导。
