Institute of Pharmacology and Toxicology, Medical Faculty, RWTH Aachen University, Wendlingweg 2, 52074, Aachen, Germany.
Institute of Occupational, Social and Environmental Medicine, Medical Faculty, RWTH Aachen University, Pauwelsstraße 30, 52074, Aachen, Germany.
Respir Res. 2022 Sep 20;23(1):261. doi: 10.1186/s12931-022-02169-5.
Respiratory diseases represent a global health burden. Because research on therapeutic strategies of airway diseases is essential, the technique of precision-cut lung slices (PCLS) has been developed and widely studied. PCLS are an alternative ex vivo model and have the potential to replace and reduce in vivo animal models. So far, the majority of studies was conducted with short-term cultivated PCLS (≤ 72 h). As there is large interest in research of chronic diseases and chronic toxicity, feasibility of cultivating human PCLS long-term over 2 weeks and recently over 4 weeks was investigated by another research group with successful results. Our aim was to establish a model of long-term cultivated rat PCLS over a period of 29 days.
Rat PCLS were cultured for 29 days and analysed regarding viability, histopathology, reactivity and gene expression at different time points during cultivation.
Cultivation of rat PCLS over a 29-day time period was successful with sustained viability. Furthermore, the ability of bronchoconstriction was maintained between 13 and 25 days, depending on the mediator. However, reduced relaxation, altered sensitivity and increased respiratory tone were observed. Regarding transcription, alteration in gene expression pattern of the investigated target genes was ascertained during long-term cultivation with mixed results. Furthermore, the preparation of PCLS seems to influence messenger ribonucleic acid (mRNA) expression of most target genes. Moreover, the addition of fetal bovine serum (FBS) to the culture medium did not improve viability of PCLS. In contrast to medium without FBS, FBS seems to affect measurements and resulted in marked cellular changes of metaplastic and/or regenerative origin.
Overall, a model of long-term cultivated rat PCLS which stays viable for 29 days and reactive for at least 13 days could be established. Before long-term cultivated PCLS can be used for in-depth study of chronic diseases and chronic toxicity, further investigations have to be made.
呼吸道疾病是全球健康负担。由于气道疾病治疗策略的研究至关重要，因此开发了并广泛研究了精密切割肺切片（PCLS）技术。PCLS 是一种替代的离体模型，具有替代和减少体内动物模型的潜力。到目前为止，大多数研究都是使用短期培养的 PCLS（≤72 小时）进行的。由于人们对慢性疾病和慢性毒性的研究兴趣很大，另一个研究小组成功地研究了长期培养人类 PCLS 超过 2 周和最近超过 4 周的可行性。我们的目标是建立一个培养 29 天的大鼠 PCLS 长期培养模型。
将大鼠 PCLS 培养 29 天，并在培养过程中的不同时间点分析其活力、组织病理学、反应性和基因表达。
大鼠 PCLS 培养 29 天成功，活力持续。此外，取决于介质，在 13 至 25 天之间保持支气管收缩能力。然而，观察到松弛减少、敏感性改变和呼吸张力增加。关于转录，在长期培养过程中，确定了所研究靶基因的基因表达模式的改变，结果喜忧参半。此外，PCLS 的制备似乎会影响大多数靶基因的信使核糖核酸（mRNA）表达。此外，向培养基中添加胎牛血清（FBS）并没有提高 PCLS 的活力。与不含 FBS 的培养基相比，FBS 似乎会影响测量结果，并导致具有化生和/或再生起源的明显细胞变化。
总之，建立了一种可长期培养大鼠 PCLS 的模型，该模型可在 29 天内保持活力，并至少在 13 天内保持反应性。在长期培养的 PCLS 可用于深入研究慢性疾病和慢性毒性之前，还需要进行进一步的研究。
