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Sviluppo di un modello biomimetico in vitro di barriera alveolo-capillare che simula il movimento respiratorio in condizioni di interfaccia aria-liquido = Development of a biomimetic in vitro model of the alveolar-capillary barrier that mimics respiratory movement under air-liquid interface conditions

Massimiliano Collu

Sviluppo di un modello biomimetico in vitro di barriera alveolo-capillare che simula il movimento respiratorio in condizioni di interfaccia aria-liquido = Development of a biomimetic in vitro model of the alveolar-capillary barrier that mimics respiratory movement under air-liquid interface conditions.

Rel. Chiara Tonda Turo, Gianluca Ciardelli. Politecnico di Torino, Corso di laurea magistrale in Ingegneria Biomedica, 2023

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Abstract:

In vitro models that mimic the alveolar-capillary barrier are becoming essential research tools to test the efficacy of drugs and understand cellular responses to different stimuli, thanks to their ability to reproduce the physiological architecture and microenvironment of the human alveoli. It is well known that different mechanical forces, under physiological or pathological conditions, significantly influence cell behaviour, therefore playing an important role in maintaining the overall structure and function of the lung. Currently available systems do not reproduce all aspects of the alveolar microenvironment and the stimuli acting on it, and they are able to recreate only small components of the alveolar region. The aim of this work was to develop a biomimetic in vitro model to mimic the alveolar-capillary barrier using the commercial bioreactor LiveBox2 (IVTech S.R.L., Massarosa (LU), Italy). In particular, in this work, the commercial polyethylene terephthalate (PET) was replaced with an electrospun polycaprolactone/ gelatin (PCL/Gel) membrane to mimic the alveolar basement membrane. The biomimetic membrane was accommodated between the two bioreactor chambers to recreate the architecture of the alveolar-capillary barrier. Moreover, the ALI interaction module was used to reproduce the air-liquid interface (ALI) condition, experienced by alveolar epithelial cells within the alveolar region. A mechanical actuator was used to partially occlude the basal chamber outlet tube after flow activation, thus reproducing the stimulation that occurs on alveolar tissue during breathing. First, a pressure sensor was used to monitor the overpressure inside the basal chamber following the mechanical actuation. The measurements were repeated for two different membranes placed between bioreactor chambers: the commercial PET membrane, and the biomimetic PCL/Gel membrane. For both, a higher overpressure was observed as the occlusion level of the outlet tube increased, but since PCL/Gel membrane was more elastic, it required greater occlusion level. Then, cell culture tests were performed seeding epithelial (A549) and endothelial (HuLEC-5A) cells on the apical and basal surface of the PCL/Gel membrane, respectively. Cells were co-cultured in the opposite side of PCL/Gel modified Transwell® inserts and inside the bioreactor chambers to compare different experimental conditions (static, dynamic with only flow and dynamic with flow and mechanical actuation). The ALI condition was established after 3 days of co-culture by removing the A549 medium from the apical chamber. After 7 days, the staining of nuclei and cytoskeleton was performed to assess the cell adhesion and viability. Finally, the effect of different stimulation on cell behaviour was qualitatively evaluated by immunofluorescence staining.

Relators: Chiara Tonda Turo, Gianluca Ciardelli
Academic year: 2022/23
Publication type: Electronic
Number of Pages: 93
Subjects:
Corso di laurea: Corso di laurea magistrale in Ingegneria Biomedica
Classe di laurea: New organization > Master science > LM-21 - BIOMEDICAL ENGINEERING
Aziende collaboratrici: UNSPECIFIED
URI: http://webthesis.biblio.polito.it/id/eprint/27881
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