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Design of flexible substrates to investigate the effect of mechanical stimuli on cell cultures for tissue engineering research

Marta Tosini

Design of flexible substrates to investigate the effect of mechanical stimuli on cell cultures for tissue engineering research.

Rel. Diana Nada Caterina Massai, Giovanni Putame. Politecnico di Torino, Corso di laurea magistrale in Ingegneria Biomedica, 2021


Tissue engineering (TE) aims at generating functional tissue constructs to be used for repairing or replacing damaged human tissues or as in vitro tissue models for investigating the physiological/pathological tissue mechanisms. Biological constructs consist of two main elements: substrate and cells. Commonly, monolayer static cell cultures are performed by using flat rigid substrates. However, for promoting the formation of functional constructs, dynamic culture conditions on flexible substrates would be more appropriate in some cases. In the last decades, bioreactors have become crucial tools in TE research. Indeed, they allow for mimicking the native dynamic environment by providing specific physico-chemical stimuli in vitro. In particular, the in vitro mechanical stimulation of cells has been proven to affect their maturation and differentiation. Moreover, the mechanical and chemical features of the substrate are crucial factors acting on the cell fate. Notwithstanding different types of artificial or natural-derived substrates have been developed, there is still room for understanding of how such properties influence the cell behaviour, especially when coupled with dynamic culture conditions. In this scenario, the aim of this work was to design different flexible substrates with defined stiffness to be used within a tunable stretch bioreactor for dynamic cell culture. The development of the substrates followed three phases: design, manufacturing, and testing. Firstly, several substrates characterized by different geometries were designed using a CAD software, and two designs were eventually considered: 1) a substrate with a single square well, and 2) a substrate with two parallel rectangular wells. Dimensions and shapes of both designs were optimized by performing Finite Element Analyses. The Von Mises stresses, different components of the strain tensor, and the deflection of the well bottom were analyzed. The design was optimized to obtain uniaxial planar deformation at the bottom of the wells. Then, the corresponding molds were designed and 3D printed, and the substrates were manufactured by casting a polydimethylsiloxane solution within the molds. The substrates were used for preliminary cell experiments to investigate the effect of stretch on periodontal ligament stem cells. Finally, the Digital Image Correlation (DIC) method was adopted for measuring the actual deformation of the substrates and for validating the simulation outcomes. Simulations showed that a uniform deformation was obtained for a substrate with a single square well and for a substrate with two rectangular wells, both symmetrical on the vertical section to minimize the out-of-plane deflection of the bottom. Biological tests pointed out the need to improve the substrate transparency for microscope inspection, and the importance of substrate functionalization for cell adhesion. DIC tests are ongoing. In conclusion, this work led to the development of flexible substrates for providing controlled mechanical stimuli to the cultured cells. The proposed solutions allow improving reproducibility of culture conditions and reducing the amount of culture medium thanks to the well shape of the substrates, with a consequent cost reduction. Further tests will be performed and new substrate designs will be developed to increase the number of parallel wells available for each substrate, in order to enable a number of statistically significant biological experiments at once.

Relators: Diana Nada Caterina Massai, Giovanni Putame
Academic year: 2021/22
Publication type: Electronic
Number of Pages: 77
Additional Information: Tesi secretata. Fulltext non presente
Corso di laurea: Corso di laurea magistrale in Ingegneria Biomedica
Classe di laurea: New organization > Master science > LM-21 - BIOMEDICAL ENGINEERING
Aziende collaboratrici: Politecnico di Torino
URI: http://webthesis.biblio.polito.it/id/eprint/20173
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