Biological effects of silk fibroin 3D scaffolds on stem cells from human exfoliated deciduous teeth (SHEDs).

The aim is to investigate in vitro biological effects of silk fibroin 3D scaffolds on stem cells from human exfoliated deciduous teeth (SHEDs) in terms of proliferation, morphological appearance, cell viability, and expression of mesenchymal stem cell markers. Silk fibroin 3D scaffolding materials may represent promising suitable scaffolds for their application in regenerative endodontic therapy approaches. SHEDs were cultured in silk fibroin 3D scaffolds. Then, cell numbers were counted and the Alamar blue colorimetric assay was used to analyse cell proliferation after 24, 48, 72, and 168 h of culture. The morphological features of SHEDs cultured on silk fibroin scaffolds were evaluated by scanning electron microscopy (SEM). Finally, cell viability and the expression of mesenchymal stem cell markers were analysed by flow cytometry. One-way analysis of variance (ANOVA) followed by a Bonferroni post-test was performed (P < 0.05). At 24 and 48 h of culture, SHED proliferation on scaffolds was modest compared to the control although still significant (p < 0.05). However, cell proliferation progressively increased from 72 to 168 h compared with the control (p < 0.001; p < 0.01). In addition, flow cytometry analysis showed that the culture of SHEDs on silk fibroin scaffolds did not significantly alter the level of expression of the mesenchymal markers CD73, CD90, or CD105 up to 168 h; in addition, cell viability in silk fibroin was similar to than obtained in plastic. Moreover, SEM studies revealed a suitable degree of proliferation, cell spreading, and attachment, especially after 168 h of culture. The findings from the current study suggest that silk fibroin 3D scaffolds had a favourable effect on the biological responses of SHEDs. Further in vivo investigations are required to confirm these results.

Effect of aqueous and particulate silk fibroin in a rat model of experimental colitis.

Silk fibroin (SF) has anti-inflammatory properties and promotes wound healing. Moreover, SF particles act as carriers of active drugs against intestinal inflammation due to their capacity to deliver the compound to the damaged colonic tissue. The present work assesses the effect of SF in the trinitrobenzenesulfonic acid model of rat colitis that resembles human intestinal inflammation. SF (8mg/kg) was administered in aqueous solution orally and in two particulate formats by intrarectal route, following two technologies: spray drying to make microparticles and desolvation in organic solvent to produce nanoparticles. SF treatments ameliorated the colonic damage, reduced neutrophil infiltration and improved the compromised oxidative status of the colon. They also reduced the gene expression of pro-inflammatory cytokines like IL-1ß and the anti-inflammatory cytokine IL-10. Moreover, they improved the intestinal wall integrity by increasing the gene expression of some of its markers (villin, trefoil factor-3 and mucins), thus accelerating the healing. The immunomodulatory properties of SF particles were also tested in vitro in macrophages: they activated the immune response in basal conditions without increasing it after a pro-inflammatory insult. In conclusion, SF particles could be useful as carriers to deliver active drugs to the damaged intestinal colon with additional anti-inflammatory and healing properties.

Effects of composite films of silk fibroin and graphene oxide on the proliferation, cell viability and mesenchymal phenotype of periodontal ligament stem cells.

In regenerative dentistry, stem cell-based therapy often requires a scaffold to deliver cells and/or growth factors to the injured site. Graphene oxide (GO) and silk fibroin (SF) are promising biomaterials for tissue engineering as they are both non toxic and promote cell proliferation. On the other hand, periodontal ligament stem cells (PDLSCs) are mesenchymal stem cells readily accessible with a promising use in cell therapy. The purpose of this study was to investigate the effects of composite films of GO, SF and GO combined with fibroin in the mesenchymal phenotype, viability, adhesion and proliferation rate of PDLSCs. PDLSCs obtained from healthy extracted teeth were cultured on GO, SF or combination of GO and SF films up to 10 days. Adhesion level of PDSCs on the different biomaterials were evaluated after 12 h of culture, whereas proliferation rate of cells was assessed using the MTT assay. Level of apoptosis was determined using Annexin-V and 7-AAD and mesenchymal markers expression of PDLSCs were analyzed by flow cytometry. At day 7 of culture, MTT experiments showed a high rate of proliferation of PDLSCs growing on GO films compared to the other tested biomaterials, although it was slightly lower than in plastic (control). However PDLSCs growing in fibroin or GO plus fibroin films showed a discrete proliferation. Importantly, at day 10 of culture it was observed a significant increase in PDLSCs proliferation rate in GO films compared to plastic (P < 0.05), as well as in GO plus fibroin compared to fibroin alone (P < 0.001). Flow cytometry analysis showed that culture of PDLSCs in fibroin, GO or GO plus fibroin films did not significantly alter the level of expression of the mesenchymal markers CD73, CD90 or CD105 up to 168 h, being the cell viability in GO even better than obtained in plastic. Our findings suggest that the combination of human dental stem cells/fibroin/GO based-bioengineered constructs have strong potential for their therapeutic use in regenerative dentistry.

Ingeniería tisular del tejido óseo. Diseño y desarrollo de materiales híbridos biológicamente activos basados en vitrocerámicas para sustitución ósea / Bone tissue engineering. Design and development of biologically active vitroceramic-based hybrid materials to be used as bone substitutes

Objetivo: Describir el desarrollo y caracterización de un material vitrocerámico y la respuesta inicial de células madre mesenquimales adultas (MSC-A) aisladas de la medula ósea. Material y metodología: El material se obtuvo por calentamiento de un vidrio 55SiO2-41CaO-4P2O5 (mol/%) por el método sol-gel. Las células se aislaron por aspirados directos de cresta ilíaca de pacientes adultos jóvenes. Se estudió el grado de adherencia, proliferación y diferenciación a osteoblastos de las MSC-A sembradas sobre el material. La diferenciación celular se evaluó mediante la producción de osteocalcina y la pérdida del marcador mesenquimal CD90. La proliferación celular sobre el sustrato se realizó mediante el ensayo de reducción de sales de tetrazolio. El material sembrado se implantó en un defecto crítico realizado en fémur de conejo para valorar su capacidad osteorregeneradora, y se observó mediante TAC. Resultados: Las MSC-A se adhirieron, expandieron, proliferaron y produjeron matriz extracelular mineralizada sobre el material durante el tiempo en cultivo, al mismo tiempo que mostraron fenotipo osteoblástico, e incrementaron la producción de osteocalcina y la pérdida de expresión de CD90. El material se reabsorbió parcialmente al final del estudio. Conclusión: El material es citocompatible, osteoconductor, bioactivo, con capacidad de promover la diferenciación de MSC-A a osteoblastos y la neoformación ósea después de su implantación en asociación con MSC-A; es una matriz adecuada para la regeneración del tejido óseo (AU)

Purpose: To describe the development and characterization of a vitroceramic material as well as the initial response of adult mesenchymal stem (MSCs-A) isolated from bone marrow. Material and methodology: The material was obtained by heating glass with composition in mol% 55SiO2-41CaO-4P2O5 by a sol gel method. Cells were isolated from direct iliac crest aspirates from young adult patients. An analysis was performed of the degree of adhesion, proliferation and osteoblastic differentiation of MSCs-A seeded onto the material. Cell differentiation was evaluated through the production of osteocalcin and the loss of the CD90 mesenchymal marker. Cell proliferation on the substrate was performed using the tetrazolium salt reduction method. The seeded material was implanted in a critical defect caused in a rabbit femur in order to determine its osteogenerating capacity; CT observations were carried out. Results: MSCs-A se bound to the material, expanded, proliferated and produced mineralized extracellular matrix on the material during the culture period. At the same time, they showed an osteoblastic phenotype, increasing osteocalcin production and losing CD90 expression. The material was partially resorbed at the end of the study. Conclusion: The material is cytocompatible, osteoconductive, bioactive and has a capacity to promote osteoblastic differentiation of MSCs-A as well as new bone formation following its implantation in association with MSCs-A; an appropriate matrix for bone tissue regeneration (AU)