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Sviluppo di scaffolds biologici da omento e muscolo scheletrico mediante processi di decellularizzazione
Full text linkABSTRACT
BACKGROUND
The availability of organs and tissues, obtained in laboratory from biological material, is one of the main demands in the event of loss of substance, due to congenital or post-traumatic defects. For this purpose, bioengineered tissues have been developed through decellularization and recellularization processes.
AIM OF THE STUDY
The objectives of this study were:
• Development of a new type of biological scaffold derived from omentum that preserves the structural integrity of the extracellular matrix and the vascular network of the native tissue even after a phase of decellularization.
• Development of a further biological scaffold from skeletal muscle, derived from the decellularization of the rectus muscle of the rabbit, for the reconstruction of a defect in the abdominal wall.
MATERIAL AND METHODS
As regards the first part of the experiment, samples of omentum, from rat and human, were decellularized through physical reactions (freezing/thawing and mechanical agitation), chemical (EDTA and isopropanol) and enzymatic (trypsin, lipases and endonucleases) methods involving the removal of cells and their lipid content.
Samples were analyzed at various stages by histological staining (hematoxylin and eosin, azan Mallory, van Gieson, PAS, Sudan, Oil Red), immunohistochemical reactions (anti- CD31, -CD34, α-smooth muscle actin) and stained with DAPI. Furthermore, evaluations about the quantitative concentration of genomic DNA at the end of decellularization were carried out with a spectrophotometer.
Attempts have been made to recellularize scaffold by cells of the stromal vascular fraction (SVF), isolated from samples of human lipoaspirate.
It has, also, developed an additional scaffolds by decellularization of samples of the rectus muscle of the rabbit. Samples were subjected to physical (freezing/thawing), chemical (EDTA and Triton X -100) and enzymatic (DNase) methods and results were analyzed by histological staining (hematoxylin and eosin, azan Mallory, van Gieson).
Thereafter, the scaffold was implanted in a rabbit receiver in correspondence with a surgical defect in the abdominal wall with loss of substance. After three weeks, the implant was studied by ultrasound examination in vivo and, after sacrifice of the animal, it was analyzed with the histological stainings mentioned above.
RESULTS
Histological stainings confirmed the effectiveness of the decellularization process with the total loss of its lipid component of the omentum. This led to the creation of an acellular scaffold in which the three-dimensional organization of the collagen, elastic, and reticular fibers was preserved. Also, the preservation of the vascular network was highlighted.
Preliminary studies have shown, moreover, the possibility of recellularization of the scaffold through the introduction of SVF cells.
Regarding the second part of the project, it was shown that the muscle-derived scaffold has maintained the integrity of the extracellular matrix and the structure of the vascular channels, in the removal of cellular elements.
Following implantation in the receiving rabbit, a solution of continuity with the adjacent tissues has been highlighted, by ultrasound in vivo, with the absence of herniation .
After the sacrifice of the animal, histological analysis of the scaffold showed the presence of reparative tissue and new vascular channels in the seat of the abdominal defect.
Future perspectives will be to recellularize the scaffold with progenitor cells to promote muscle regeneration.
CONCLUSIONS
We obtained a new type of acellular biological structure resulting from the omentum, through the application of the process of decellularization, which allows to minimize possible alterations of the extracellular matrix during the removal of the cells.
The scaffold was preliminarily recellularized with cells of the stromal vascular fraction, but further experiments will focus on the further evolution of in vitro recellularization and in vivo implantation.
The decellularization of the muscle tissue and its implantation in vivo has allowed to identify a biological structure capable of offering a valid alternative to the materials currently used for the defects repair in the abdominal wall.
Also in this direction we will evaluate in the future the possibility of recellularization of the scaffold with progenitor cells
Development of biological scaffolds through decellularization of human arteries and veins
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No full textFunctional maturation of the carotid body in the postnatal period relies partly on structural and neurochemical changes, which are reviewed here. Structural changes include changes in cytological composition, and increases in glomic tissue volume, dense-cored granules of type I cells, synapses of type I cells with type II cells and afferent nerve fibres. Vascular volume also increases, but in the same proportion as extravascular volume. During maturation, the carotid body also shows higher density and hypoxic sensitivity of K(+)-channels and an increased hypoxic [Ca(2+)](i) response. Modulation of content and release of catecholamine occurs, together with decreased expression of tyrosine hydroxylase and dopamine β-hydroxylase and increased expression of choline acetyltransferase. Expression of dopamine 2 receptor and nicotinic α3 and α7 receptor subunits increases, and muscarinic M1 receptor protein, nicotinic α4 and β2 receptor subunits and adenosine receptor 1 decrease. Maturation of the carotid body may also be explained with reference to the developmentally regulated expression of trophic factors and their receptors
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Modelli sperimentali di vasi di piccolo calibro bioingegnerizzati per applicazioni microchirurgiche
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