Included in this dossier are three articles that explore three interesting topics: the governmental policies on tissue engineering in Latin America, the approaches and methods used to elaborate a bio-tooth, and the applications of platelet-rich fibrin in regenerative dentistry. ; Se incluyen en este dossier tres artículos que exploran tres temas interesantes: las políticas gubernamentales en materia de ingeniería tisular en Latinoamérica, los enfoques y métodos empleados para elaborar un biodiente, y las aplicaciones de la fibrina rica en plaquetas en odontología regenerativa. ; Incluídos neste dossiê estão três artigos que exploram três tópicos interessantes: as políticas governamentais sobre engenharia de tecidos na América Latina, as abordagens e métodos usados para elaborar um bio-dente e as aplicações da fibrina rica em plaquetas na odontologia regenerativa.
Patients with severe limbal damage and limbal stem cell deficiency are a therapeutic challenge. We evaluated four decellularization protocols applied to the full-thickness and half-thickness porcine limbus, and we used two cell types to recellularize the decellularized limbi. The results demonstrated that all protocols achieved efficient decellularization. However, the method that best preserved the transparency and composition of the limbus extracellular matrix was the use of 0.1% SDS applied to the half-thickness limbus. Recellularization with the limbal epithelial cell line SIRC and human adipose-derived mesenchymal stem cells (hADSCs) was able to generate a stratified epithelium able to express the limbal markers p63, pancytokeratin, and crystallin Z from day 7 in the case of SIRC and after 14–21 days of induction when hADSCs were used. Laminin and collagen IV expression was detected at the basal lamina of both cell types at days 14 and 21 of follow-up. Compared with control native limbi, tissues recellularized with SIRC showed adequate picrosirius red and alcian blue staining intensity, whereas limbi containing hADSCs showed normal collagen staining intensity. These preliminary results suggested that the limbal substitutes generated in this work share important similarities with the native limbus and could be potentially useful in the future. ; Spanish Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica (I+D+i) of the Spanish Ministry of Economy and Competitiveness (Instituto de Salud Carlos III), Grants FIS PI20/0317 and ICI21-00010, cofinanced by FEDER funds (European Union). This work was also supported by grant PI-0086-2020 from Consejería de Salud y Familias, Junta de Andalucía, Spain, and grant B-CTS-504-UGR20 (Proyectos de I+D+i en el marco del Programa Operativo FEDER Andalucía 2014–2020) from the University of Granada, Consejería de Transformación Económica, Industria, Conocimiento y Universidades, Junta de Andalucía, and European Union (cofinanced by FEDER funds).
Background: Treatment of patients affected by severe burns is challenging, especially due to the high risk of Pseudomonas infection. In the present work, we have generated a novel model of bioartificial human dermis substitute by tissue engineering to treat infected wounds using fibrin-agarose biomaterials functionalized with nanostructured lipid carriers (NLCs) loaded with two anti-Pseudomonas antibiotics: sodium colistimethate (SCM) and amikacin (AMK). Results: Results show that the novel tissue-like substitutes have strong antibacterial effect on Pseudomonas cultures, directly proportional to the NLC concentration. Free DNA quantification, WST-1 and Caspase 7 immunohistochemical assays in the functionalized dermis substitute demonstrated that neither cell viability nor cell proliferation were affected by functionalization in most study groups. Furthermore, immunohistochemistry for PCNA and KI67 and histochemistry for collagen and proteoglycans revealed that cells proliferated and were metabolically active in the functionalized tissue with no differences with controls. When functionalized tissues were biomechanically characterized, we found that NLCs were able to improve some of the major biomechanical properties of these artificial tissues, although this strongly depended on the type and concentration of NLCs. Conclusions: These results suggest that functionalization of fibrin-agarose human dermal substitutes with antibioticloaded NLCs is able to improve the antibacterial and biomechanical properties of these substitutes with no detectable side effects. This opens the door to future clinical use of functionalized tissues. ; NanoGSkin project of EuroNanoMed-III (ERA-NET Cofund scheme of the Horizon 2020 Research and Innovation Framework Programme), EU ; Instituto de Salud Carlos III AC17/00013 ; Centro para el Desarrollo Tecnológico Industrial -CDTI 00108589 ; Spanish Government ; Junta de Andalucía PE-0395-2019 ; Fundacion Benefica Anticancer San Francisco Javier y Santa Candida, Granada, Spain ; Department of Economic Development and Infrastructure of the Basque Government budget, through the HAZITEK business R + D support program ZE-2017/00014 ; European Union (EU) OTRI.35A-07
Temporo-mandibular joint disc disorders are highly prevalent in adult populations. Autologous chondrocyte implantation is a well-established method for the treatment of several chondral defects. However, very few studies have been carried out using human fibrous chondrocytes from the temporo-mandibular joint (TMJ). One of the main drawbacks associated to chondrocyte cell culture is the possibility that chondrocyte cells kept in culture tend to de-differentiate and to lose cell viability under in in-vitro conditions. In this work, we have isolated human temporo-mandibular joint fibrochondrocytes (TMJF) from human disc and we have used a highly-sensitive technique to determine cell viability, cell proliferation and gene expression of nine consecutive cell passages to determine the most appropriate cell passage for use in tissue engineering and future clinical use. Our results revealed that the most potentially viable and functional cell passages were P5–P6, in which an adequate equilibrium between cell viability and the capability to synthesize all major extracellular matrix components exists. The combined action of pro-apoptotic (TRAF5, PHLDA1) and anti-apoptotic genes (SON, HTT, FAIM2) may explain the differential cell viability levels that we found in this study. These results suggest that TMJF should be used at P5–P6 for cell therapy protocols. ; This work was supported by University of Granada-Campus de Excelencia Internacional, Subprograma de I+D+I y Transferencia and by the Spanish Ministry of Economy and Competitiveness, grant IPT-300000-2010-017 (INNPACTO program), co-financed by the European Regional Development Fund, European Union.
Spanish Plan Nacional de Investigacion Cientifica, Desarrollo e Innovacion Tecnologica (I+D+I) of the Spanish Ministry of Science and Innovation (Instituto de Salud Carlos III), Grant/Award Number: FIS PI18/331, FIS PI21/00980, FIS PI18/332 and ICI19/00024; Consejeria de Salud y Familias, Junta de Andalucia, Spain, Grant/Award Number: PI-0442--2019; FEDER funds, European Union ; Objective: The aim of this study was to generate novel models of bioartificial human oral mucosa with increased vascularization potential for future use as an advanced therapies medicinal product, by using different vascular and mesenchymal stem cell sources. Background: Oral mucosa substitutes could contribute to the clinical treatment of complex diseases affecting the oral cavity. Although several models of artificial oral mucosa have been described, biointegration is a major issue that could be favored by the generation of novel substitutes with increased vascularization potential once grafted in vivo. Methods: Three types of mesenchymal stem cells (MSCs) were obtained from adipose tissue, bone marrow, and dental pulp, and their in vitro potential was evaluated by inducing differentiation to the endothelial lineage using conditioning media. Then, 3D models of human artificial oral mucosa were generated using biocompatible fibrin-agarose biomaterials combined with human oral mucosa fibroblasts and each type of MSC before and after induction to the endothelial lineage, using human umbilical vein endothelial cells (HUVEC) as controls. The vascularization potential of each oral mucosa substitute was assessed in vitro and in vivo in nude mice. Results: In vitro induction of MSCs kept in culture was able to increase the expression of VEGF, CD31, and vWF endothelial markers, especially in bone marrow and dental pulp-MSCs, and numerous proteins with a role in vasculogenesis become overexpressed. Then, in vivo grafting resulted in a significant increase in blood vessels formation at the interface area between the graft and the host tissues, with significantly positive expression of VEGF, CD31, vWF, and CD34 as compared to negative controls, especially when pre-differentiated MSCs derived from bone marrow and dental pulp were used. In addition, a significantly higher number of cells committed to the endothelial lineage expressing the same endothelial markers were found within the bioartificial tissue. Conclusion: Our results suggest that the use of pre-differentiated MSCs could contribute to a rapid generation of a vascular network that may favor in vivo biointegration of bioengineered human oral mucosa substitutes. ; Spanish Plan Nacional de Investigacion Cientifica, Desarrollo e Innovacion Tecnologica (I+D+I) of the Spanish Ministry of Science and Innovation (Instituto de Salud Carlos III) FIS PI18/331 FIS PI21/00980 FIS PI18/332 ICI19/00024 ; Junta de Andalucia PI-0442-2019 ; European Commission
Purpose: Human cornea substitutes generated by tissue engineering currently require limbal stem cells for the generation of orthotypical epithelial cell cultures. We recently reported that bioengineered corneas can be fabricated in vitro from a heterotypical source obtained from Wharton's jelly in the human umbilical cord (HWJSC). Methods: Here, we generated a partial thickness cornea model based on plastic compression nanostructured fibrin-agarose biomaterials with cornea epithelial cells on top, as an orthotypical model (HOC), or with HWJSC, as a heterotypical model (HHC), and determined their potential in vivo usefulness by implantation in an animal model. Results: No major side effects were seen 3 and 12 months after implantation of either bioengineered partial cornea model in rabbit corneas. Clinical results determined by slit lamp and optical coherence tomography were positive after 12 months. Histological and immunohistochemical findings demonstrated that in vitro HOC and HHC had moderate levels of stromal and epithelial cell marker expression, whereas in vivo grafted corneas were more similar to control corneas. Conclusion: These results suggest that both models are potentially useful to treat diseases requiring anterior cornea replacement, and that HHC may be an efficient alternative to the use of HOC which circumvents the need to generate cornea epithelial cell cultures. ; Spanish Plan Nacional de Investigacion Cientifica, Desarrollo e Innovacion Tecnologica (I CD Ci) from the Spanish Ministry of Science and Innovation: Instituto de Salud Carlos III FIS PI17/0391 FIS PI14/0955 ; European Union (EU) ; Spanish Plan Nacional de Investigacion Cientifica, Desarrollo e Innovacion Tecnologica (I CD Ci) from the Spanish Ministry of Science and Innovation: Ministry of Science, Innovation and Universities of Spain PGC2018-101904-A-I00 RTC-2017-6696-1
