Studio di biomateriali usati come scaffold per Tissue Engineering e loro caratterizzazione con tecniche spettroscopiche vibrazionali e di analisi termica

This research investigated someone of the main problems connected to the application of Tissue Engineering in the prosthetic field, in particular about the characterization of the scaffolding materials and biomimetic strategies adopted in order to promote the implant integration. The spectroscopic and thermal analysis techniques were usefully applied to characterize the chemico-physical properties of the materials such as – crystallinity; – relative composition in case of composite materials; – Structure and conformation of polymeric and peptidic chains; – mechanism and degradation rate; – Intramolecular and intermolecular interactions (hydrogen bonds, aliphatic interactions). This kind of information are of great importance in the comprehension of the interactions that scaffold undergoes when it is in contact with biological tissues; this information are fundamental to predict biodegradation mechanisms and to understand how chemico-physical properties change during the degradation process. In order to fully characterize biomaterials, this findings must be integrated by information relative to mechanical aspects and in vitro and in vivo behavior thanks to collaborations with biomedical engineers and biologists. This study was focussed on three different systems that correspond to three different strategies adopted in Tissue Engineering: biomimetic replica of fibrous 3-D structure of extracellular matrix (PCL-PLLA), incorporation of an apatitic phase similar to bone inorganic phase to promote biomineralization (PCL-HA), surface modification with synthetic oligopeptides that elicit the interaction with osteoblasts. The characterization of the PCL-PLLA composite underlined that the degradation started along PLLA fibres, which are more hydrophylic, and they serve as a guide for tissue regeneration. Moreover it was found that some cellular lines are more active in the colonization of the scaffold. In the PCL-HA composite, the weight ratio between the polymeric and the inorganic phase plays an essential role both in the degradation process and in the biomineralization of the material. The study of self-assembling peptides allowed to clarify the influence of primary structure on intermolecular and intermolecular interactions, that lead to the formation of the secondary structure and it was possible to find a new class of oligopeptides useful to functionalize materials surface. Among the analytical techniques used in this study, Raman vibrational spectroscopy played a major role, being non-destructive and non-invasive, two properties that make it suitable to degradation studies and to morphological characterization. Also micro-IR spectroscopy was useful in the comprehension of peptide structure on oxidized titanium: up to date this study was one of the first to employ this relatively new technique in the biomedical field.

Human bone: the tissue characteristics determining its mechanical behaviour

The present thesis illustrates the research carried out during the PhD studies in Bioengineering. The research was aimed to characterise the human bone tissue, with particular regard to the differences between cortical and trabecular bone. The bone tissue characteristics that affect its mechanical properties were verified or identified, using an experimental approach, to corroborate or refute hypotheses based on the state of the art in bone tissue biomechanics. The studies presented in the present PhD thesis were designed to investigate aspects of bone tissue biomechanics, which were in need of a more in-depth examination since the data found in the literature was contradictory or scarce. In particular, the work was focalised on the characterisation of the basic structure of the bone tissue (groups of lamellae), its composition, its spatial organisation (trabecular bone microarchitecture) and their influence on the mechanical properties. In conclusion, the present thesis integrates eight different studies on the characterisation of bone tissue. A more in-depth examination of some of the aspects of bone tissue biomechanics where the data found in the literature was contradictory or scarce was performed. Bone tissue was investigated at several scales, from its composition up to its spatial organization, to determine which parameters influence the mechanical behaviour of the tissue. It was found that although the composition and real density of bone tissue are similar, the differences in structure at different levels cause differences between the two types of bone tissue (cortical and trabecular) in mechanical properties. However, the apparent density can still be considered a good predictor of the mechanical properties of both cortical and trabecular bone. Finally, it was found that the bone tissue characteristics might change when a pathology is present, as demonstrated for OA.

Electrospun Polymeric Scaffolds with Enhanced Biomimetic Properties for Tissue Engineering Applications

This PhD Thesis is focused on the development of fibrous polymeric scaffolds for tissue engineering applications and on the improvement of scaffold biomimetic properties. Scaffolds were fabricated by electrospinning, which allows to obtain scaffolds made of polymeric micro or nanofibers. Biomimetism was enhanced by following two approaches: (1) the use of natural biopolymers, and (2) the modification of the fibers surface chemistry. Gelatin was chosen for its bioactive properties and cellular affinity, however it lacks in mechanical properties. This problem was overcome by adding poly(lactic acid) to the scaffold through co-electrospinning and mechanical properties of the composite constructs were assessed. Gelatin effectively improves cell growth and viability and worth noting, composite scaffolds of gelatin and poly(lactic acid) were more effective than a plain gelatin scaffold. Scaffolds made of pure collagen fibers were fabricated. Modification of collagen triple helix structure in electrospun collagen fibers was studied. Mechanical properties were evaluated before and after crosslinking. The crosslinking procedure was developed and optimized by using - for the first time on electrospun collagen fibers - the crosslinking reactant 1,4-butanediol diglycidyl ether, with good results in terms of fibers stabilization. Cell culture experiments showed good results in term of cell adhesion and morphology. The fiber surface chemistry of electrospun poly(lactic acid) scaffold was modified by plasma treatment. Plasma did not affect thermal and mechanical properties of the scaffold, while it greatly increased its hydrophilicity by the introduction of carboxyl groups at the fiber surface. This fiber functionalization enhanced the fibroblast cell viability and spreading. Surface modifications by chemical reactions were conducted on electrospun scaffolds made of a polysophorolipid. The aim was to introduce a biomolecule at the fiber surface. By developing a series of chemical reactions, one oligopeptide every three repeating units of polysophorolipid was grafted at the surface of electrospun fibers.