Continuous purification and culture of rat type 1 and type 2 alveolar epithelial cells by magnetic cell sorting
磁细胞分选连续纯化培养大鼠1型和2型肺泡上皮细胞
磁気セルソーティングによるラット1型および2型肺胞上皮細胞の連続精製および培養
자기 세포 분류에 의한 랫트 1형 및 2형 폐포 상피 세포의 연속 정제 및 배양
Purificación continua y cultivo de células epiteliales alveolares tipo 1 y tipo 2 de rata mediante clasificación celular magnética
Purification et culture en continu de cellules épithéliales alvéolaires de rat de type 1 et de type 2 par tri cellulaire magnétique
Непрерывная очистка и культивирование альвеолярных эпителиальных клеток крыс 1-го и 2-го типов методом магнитной сортировки клеток
Di Liu 刘迪 ¹, Jian-Hui Sun 孙剑会 ², Hua-Cai Zhang 张华才 ¹, Jian-Xin Jiang 蒋建新 ¹, Ling Zeng 曾灵 ¹
¹ State Key Laboratory of Trauma, Burns and Combined Injuries, Daping Hospital, Army Medical University, Chongqing, 400042, China
中国 重庆 陆军军医大学大坪医院 创伤、烧伤与复合伤研究国家重点实验室
² Medical Center of Trauma and War injury, Daping Hospital, Army Medical University, Chongqing, 400042, China
中国 重庆 陆军军医大学大坪医院 战创伤医学中心
The incidence of acute lung injury (ALI) in severe trauma patients is 48% and the mortality rate following acute respiratory distress syndrome evolved from ALI is up to 68.5%. Alveolar epithelial type 1 cells (AEC1s) and type 2 cells (AEC2s) are the key cells in the repair of injured lungs as well as fetal lung development. Therefore, the purification and culture of AEC1s and AEC2s play an important role in the research of repair and regeneration of lung tissue.
Methods
Sprague-Dawley rats (3-4 weeks, 120-150 g) were purchase for experiment. Dispase and DNase I were jointly used to digest lung tissue to obtain a single-cell suspension, and then magnetic bead cell sorting was used to isolate AEC1s from the single-cell suspension by using AEC1s cell membrane marker protein T1a. Afterwards, alveolar epithelial cell membrane marker protein EpCAM was used to sort the remaining AEC2s. Cell purity was identified by immunofluorescence staining and flow cytometry.
Results
The purity of AEC1s and AEC2s was 88.3% ± 3.8% and 92.6% ± 2.7%. The cell growth was observed as follows: AEC1s stretched within the 12 h-16 h, but the cells proliferated slowly; while AEC2s began to stretch after 24 h and proliferated rapidly from the 2nd day and began to differentiate after 3 days.
Conclusion
AEC1s and AEC2s sorted by this method have high purity and good viability. Therefore, our method provides a new approach for the isolation and culture of AEC1s and AEC2s as well as a new strategy for the research of lung repair and regeneration.
