The electrospinning is a highly promising technique for the fabrication of scaffolds, widely used in the field of tissue engineering. This technique allows to obtain fibrillar structures similar to those of the ECM, because they offer high surface areas and ease of manipulation of the properties to be suitable for different purposes. The solvent used for this work is that formed by the mixture of acids, called 'Green Solvent' because it represents an innovative solvent that has gained much attention in recent years. Its main characteristic is that it is less toxic and is especially great for getting the fine fibers, although it may seem that more harmful because it contains acid. While the polymers used are: Polyammide or Nylon 6 (PA6), Polycaprolactone (PCL), Poly L-lactic acid (PLLA) and the copolymer Poly L-lactide-co-ɛ-caprolactone (PLCL) blend of PCL and PLLA in 30:70 ratio in the mix of acids The fibers obtained with these polymer solutions have been characterized through a morphological analysis by SEM, and also with a mechanical characterization, which was carried through tensile tests and contact angle measurements to know the wettability of the samples. This choice is due to the desire to see if actually this solvent is really innovative and non-toxic. Biological studies were conducted at the Laboratory of Molecular Medicine of Pavia, where were carried out biocompatibility tests with assessment of the effect of the morphology and size of the fibers on cell viability. The cell line used in this case has been to endothelial, in particular, have been used ECV304, human endothelial cells. Further, biological analysis were made using the CLSM (Confocal Laser Scanning Microscope) to see the qualitatively expression of endothelial specific proteins, incurred through the quantitative analysis of the Western blotting technique and RT-PCR. Eventually, studies of hydrolytic degradability were performed in phosphate buffer to study the timing of degradation of the different scaffolds based on the chemical structure of the polymers which constitute them.
L’elettrofilatura (o electrospinning) è una tecnica estremamente promettente per la realizzazione di scaffold da impiegarsi nel campo dell’ingegneria tissutale. Tale tecnica permette di ottenere tessuti simili alle strutture fibrillari della ECM, perchè offrono elevate aree superficiali e facilità di manipolazione delle proprietà per adattarle a scopi differenti. Per la costruzione di questi scaffold nanofibrosi è stato utilizzato come solvente un mix di acidi: Acido Acetico: Acido Formico : Cloroformio. La scelta è ricaduta su questo solvente perché è ritenuto innovativo, risultando meno tossico rispetto a quelli usati comunemente. In questo solvento sono stati sciolti dei polimeri quali: Polyammide or Nylon 6 (PA6), Polycaprolactone (PCL), Poly L-lactic acid (PLLA) e il copolimero Poly L-lactide-co-ɛ-caprolactone (PLCL) che è il blend di PCL e PLLA in un rapport 30:70. Le fibre sono state caratterizzate attraverso un analisi morfologica mediante il SEM, sono state valutate le loro proprietà meccaniche e il loro grado di bagnabilità attraverso il contact angle. Nel laboratorio di Medicina Molecolare a Pavia, si è voluto testare la biocompatibilità di questi scaffold usando come linea cellulare le cellule endoteliali umane. Ulterori analisi biologiche sono state fatte utilizzando il CLSM (Confocal Laser Scanning Microscope) per verificare qualitativamente l’espressione di proteine specifiche dell’endotelio, affiancate ad analisi quantitative attraverso la tecnica del Western Blot e RT-PCR. Infine, sono stati eseguiti studi di biodegradabilità idrolitica in tampone fosfato, per studiare i tempi di degradazione dei diversi scaffold sulla base della struttura chimica dei polimeri che li costituiscono.
Nanofibre elettrospinnate per la rigenerazione tissutale: uno studio di biocompatibilità con cellule endoteliali umane
CARROZZINI, TATIANA
2015/2016
Abstract
The electrospinning is a highly promising technique for the fabrication of scaffolds, widely used in the field of tissue engineering. This technique allows to obtain fibrillar structures similar to those of the ECM, because they offer high surface areas and ease of manipulation of the properties to be suitable for different purposes. The solvent used for this work is that formed by the mixture of acids, called 'Green Solvent' because it represents an innovative solvent that has gained much attention in recent years. Its main characteristic is that it is less toxic and is especially great for getting the fine fibers, although it may seem that more harmful because it contains acid. While the polymers used are: Polyammide or Nylon 6 (PA6), Polycaprolactone (PCL), Poly L-lactic acid (PLLA) and the copolymer Poly L-lactide-co-ɛ-caprolactone (PLCL) blend of PCL and PLLA in 30:70 ratio in the mix of acids The fibers obtained with these polymer solutions have been characterized through a morphological analysis by SEM, and also with a mechanical characterization, which was carried through tensile tests and contact angle measurements to know the wettability of the samples. This choice is due to the desire to see if actually this solvent is really innovative and non-toxic. Biological studies were conducted at the Laboratory of Molecular Medicine of Pavia, where were carried out biocompatibility tests with assessment of the effect of the morphology and size of the fibers on cell viability. The cell line used in this case has been to endothelial, in particular, have been used ECV304, human endothelial cells. Further, biological analysis were made using the CLSM (Confocal Laser Scanning Microscope) to see the qualitatively expression of endothelial specific proteins, incurred through the quantitative analysis of the Western blotting technique and RT-PCR. Eventually, studies of hydrolytic degradability were performed in phosphate buffer to study the timing of degradation of the different scaffolds based on the chemical structure of the polymers which constitute them.È consentito all'utente scaricare e condividere i documenti disponibili a testo pieno in UNITESI UNIPV nel rispetto della licenza Creative Commons del tipo CC BY NC ND.
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https://hdl.handle.net/20.500.14239/23060