Xenogeneic Mesenchymal Stem Cells in the Formation of Hyaline Cartilage in Osteochondral Goat Failure

Background: Osteochondral knee failures are among the most common causes of disability among the elderly human population and animal athletes. The xenogeneic transplantation of mesenchymal stem cells is a questionable therapeutic alternative that, despite the low expression of Major Histocompatibility Complex type II by these cells, still has relevantuncertainties about the safety and clinical efficacy. The main objective of the present study was to investigate whether the xenogeneic transplantation of mesenchymal stem cells induces hyaline cartilage formation, without histopathological evidence of rejection, in osteochondralfailures of goats.Materials, Methods & Results: Five female goats were used, submitted to three surgical osteocondral failures in the right knee, treated with xenogenic mesenchymal stem cells of dental pulp, xenogenic platelet-rich plasma and hemostatic sponge of hydrolyzed collagen, respectively. The lesions were evaluated after 60 days of treatment, aiming to identify thepresence of hyaline cartilage or fibrocartilage and the subchondral bone pattern (regenerated or disorganized). Transplantation of xenogenic mesenchymal stem cells induced predominant formation of hyaline cartilage (P 0.05). Macroscopically, the lesions of the stem cell treated group showed formation of firm repair tissue, opaque staining, integrated with adjacent cartilage and with the failure filling almost completely. The groups treated with PRP and hemostatic sponge of hydrolyzed collagen presented, on average, partial filling of the lesion, with irregular shape and darkened coloration.Discussion. The absence of macroscopic and histopathological evidences of an inflammatory process on the surface and in the internal portion of the osteochondral lesions treated with xenogeneic stem cells, probably due to the low expression of Major Histocompatibility Complex type II by these cells, which would theoretically induce low rejection response. Such observations are of great importance, since graft-versus- host disease syndrome is a serious condition, responsible for the low therapeutic efficacy with transplantation of cells or grafts in humans. The formation of fibrocartilage, although without macro and microscopic evidence of degeneration or necrosis, in the osteochondral failures treated with PRP and hemostatic collagen sponge suggest that paracrine factors of the local microenvironment of the osteochondral failure are possibly responsible for the formation of fibrocartilaginous tissue or by inhibition of normal cartilage formation. The fibrocartilage formed in the Plasmaand Control groups, contributed to the commitment in the filling of the lesion, contrasting with the almost complete fill of the lesions treated with stem cells. The xenotransplantation of mesenchymal stem cells induced formation of hyaline cartilage and did not promote histopathological evidence of rejection in osteochondral lesions of goat knees. The treatments with PRP and hemostatic sponge of hydrolyzed collagen induced greater formation of fibrocartilaginous cartilaginous surface in the osteochondral failures

The influence of Aloe vera with mesenchymal stem cells from dental pulp on bone regeneration: characterization and treatment of non-critical defects of the tibia in rats

Abstract Objective This study aimed to evaluate the inflammatory effect and bone formation in sterile surgical failures after implantation of a collagen sponge with mesenchymal stem cells from human dental pulp (hDPSCs) and Aloe vera. Material and Methods Rattus norvegicus (n=75) were divided into five experimental groups according to treatment: G1) control (blood clot); G2) Hemospon®; G3) Hemospon® in a culture medium enriched with 8% Aloe vera; G4) Hemospon® in a culture medium containing hDPSCs and G5) Hemospon® in a culture medium enriched with 8% Aloe vera and hDPSCs. On days 7, 15 and 30, the animals were euthanized, and the tibia was dissected for histological, immunohistochemistry and immunofluorescence analyses. The results were analyzed using nonparametric Kruskal-Wallis test and Dunn’s post-test. Results On days 7 and 15, the groups with Aloe vera had less average acute inflammatory infiltrate compared to the control group and the group with Hemospon® (p<0.05). No statistically significant difference was found between the groups regarding bone formation at the three experimental points in time. Osteopontin expression corroborated the intensity of bone formation. Fluorescence microscopy revealed positive labeling with Q-Tracker® in hDPSCs before transplantation and tissue repair. Conclusion The results suggest that the combination of Hemospon®, Aloe vera and hDPSCs is a form of clinical treatment for the repair of non-critical bone defects that reduces the inflammatory cascade’s effects

Behavior and biocompatibility of rabbit bone marrow mesenchymal stem cells with bacterial cellulose membrane

Background Tissue engineering has been shown to exhibit great potential for the creation of biomaterials capable of developing into functional tissues. Cellular expansion and integration depends on the quality and surface-determinant factors of the scaffold, which are required for successful biological implants. The objective of this research was to characterize and evaluate the in vitro characteristics of rabbit bone marrow mesenchymal stem cells (BM-MSCs) associated with a bacterial cellulose membrane (BCM). We assessed the adhesion, expansion, and integration of the biomaterial as well as its ability to induce macrophage activation. Finally, we evaluated the cytotoxicity and toxicity of the BCM. Methods Samples of rabbit bone marrow were collected. Mesenchymal stem cells were isolated from medullary aspirates to establish fibroblast colony-forming unit assay. Osteogenic, chondrogenic, and adipogenic differentiation was performed. Integration with the BCM was assessed by scanning electron microscopy at 1, 7, and 14 days. Cytotoxicity was assessed via the production of nitric oxide, and BCM toxicity was assessed with the MTT assay; phagocytic activity was also determined. Results The fibroblastoid colony-forming unit (CFU-F) assay showed cells with a fibroblastoid morphology organized into colonies, and distributed across the culture area surface. In the growth curve, two distinct phases, lag and log phase, were observed at 15 days. Multipotentiality of the cells was evident after induction of osteogenic, chondrogenic, and adipogenic lineages. Regarding the BM-MSCs’ bioelectrical integration with the BCM, BM-MSCs were anchored in the BCM in the first 24 h. On day 7 of culture, the cytoplasm was scattered, and on day 14, the cells were fully integrated with the biomaterial. We also observed significant macrophage activation; analysis of the MTT assay and the concentration of nitric oxide revealed no cytotoxicity of the biomaterial. Conclusion The BCM allowed the expansion and biointegration of bone marrow progenitor cells with a stable cytotoxic profile, thus presenting itself as a biomaterial with potential for tissue engineering

Plasticidade estrutural e isolamento de células progenitoras do cordão umbilical de cutias (Dasyprocta prymnolopha) criadas em cativeiro

The agouti has been used as an experimental model in several studies focused on reproductive biology. The umbilical cord, an embryonic attachment that connects the foetus to the placenta, has been reported as an important anatomical site for obtaining stem cells. The objective of this study was to describe macro- and microscopically the umbilical cord of agoutis at different stages of gestation, to expand and cultivate in vitro the progenitor cells and to report their morphological characteristics. Seven cutias were submitted to caesarean section to collect the umbilical cords: five were destined for studies of cord structure in different stages of gestation (30, 35, 50, 75 and 100 days postcoital), and two were collected in the third stage of gestation for isolation and cell culture. The umbilical cord of cutias assumes a spiral arrangement, with veins and arteries on it starting 50 days after coitus. The arteries present an outer layer of smooth muscle fibres in a longitudinal and circular arrangement and a medium layer of smooth muscle fibres with only longitudinal and intimate orientation and coated by the endothelium. The veins consist of longitudinal smooth muscle fibres with an extract of smooth muscle cells, and the endothelium, in all analysed gestational phases, is a structure bounded by simple pavement epithelial tissue originating from the amnion, adhered to Wharton's Jelly and forming the umbilical vessels and allantoid duct. The proposed protocol allowed the collection of a high cellular concentration of umbilical cord progenitor cells from viable cutias.A cutia vem sendo utilizada como modelo experimental em diversos estudos voltados à biologia reprodutiva. O cordão umbilical, anexo embrionário que une o feto à placenta, tem sido relatado como um importante sítio anatômico para obtenção de células-tronco. O objetivo deste estudo foi descrever macro e microscopicamente o cordão umbilical de cutias, em fases diferentes da gestação, expandir e cultivar in vitro as células progenitoras e relatar suas características morfológicas. Foram utilizadas sete cutias submetidas à cesariana para a coleta dos cordões umbilicais, cinco foram destinadas aos estudos da estrutura do cordão, em diferentes estágios de gestação (30, 35, 50, 75 e 100 dias pós-coito), e duas, no terço final da gestação, para isolamento e cultivo celular. O cordão umbilical de cutia assume disposição espiralada, com veias e artérias sobre ele a partir dos 50 dias após o coito. As artérias apresentam camada externa de fibras musculares lisas, disposição longitudinal e circular, camada média de fibras musculares lisas, apenas com disposição longitudinal e íntima revestida pelo endotélio. As veias constituídas por fibras musculares lisas longitudinais com um extrato de células musculares lisas e pelo endotélio. Em todas as fases gestacionais analisadas é uma estrutura delimitada por tecido epitelial simples pavimentoso, proveniente do âmnio, aderido a Geleia de Wharton e com formação de vasos umbilicais e ducto alantóide. O protocolo proposto permitiu a coleta de células progenitoras do cordão umbilical de cutias, viáveis com elevada concentração celular

Novel Scaffold Based on Chitosan Hydrogels/Phthalated Cashew Gum for Supporting Human Dental Pulp Stem Cells

Hydrogels are structures that have value for application in the area of tissue engineering because they mimic the extracellular matrix. Naturally obtained polysaccharides, such as chitosan (CH) and cashew gum, are materials with the ability to form polymeric networks due to their physicochemical properties. This research aimed to develop a scaffold based on chitosan and phthalated cashew tree gum and test it as a support for the growth of human mesenchymal stem cells. In this study, phthalation in cashew gum (PCG) was performed by using a solvent-free route. PCG-CH scaffold was developed by polyelectrolyte complexation, and its ability to support adherent stem cell growth was evaluated. The scaffold showed a high swelling rate. The pore sizes of the scaffold were analyzed by scanning electron microscopy. Human dental pulp stem cells (hDPSCs) were isolated, expanded, and characterized for their potential to differentiate into mesenchymal lineages and for their immunophenotypic profile. Isolated mesenchymal stem cells presented fibroblastoid morphology, plastic adhesion capacity, and differentiation in osteogenic, adipogenic, and chondrogenic lineages. Mesenchymal stem cells were cultured in scaffolds to assess cell adhesion and growth. The cells seeded on the scaffold showed typical morphology, attachment, and adequate distribution inside the matrix pores. Thus, cells seeded in the scaffold may improve the osteoinductive and osteoconductive properties of these biomaterials

Renal Progenitor Cells Have Higher Genetic Stability and Lower Oxidative Stress than Mesenchymal Stem Cells during In Vitro Expansion

In vitro senescence of multipotent cells has been commonly associated with DNA damage induced by oxidative stress. These changes may vary according to the sources of production and the studied lineages, which raises questions about the effect of growing time on genetic stability. This study is aimed at evaluating the evolution of genetic stability, viability, and oxidative stress of bone marrow mesenchymal stem cells (MSCBMsu) and renal progenitor cells of the renal cortex (RPCsu) of swine (Sus scrofa domesticus) in culture passages. P2, P5, and P9 were used for MSCBMsu and P1, P2, and P3 for RPCsu obtained by thawing. The experimental groups were submitted to MTT, apoptosis and necrosis assays, comet test, and reactive substance measurements of thiobarbituric acid (TBARS), nitrite, reduced glutathione (GSH), and catalase. The MTT test curve showed a mean viability of 1.14±0.62 and 1.12±0.54, respectively, for MSCBMsu and RPCsu. The percentages of MSCBMsu and RPCsu were presented, respectively, for apoptosis, an irregular and descending behavior, and necrosis, ascending and irregular. The DNA damage index showed higher intensity among the MSCBMsu in the P5 and P9 passages (p<0.05). In the TBARS evaluation, there was variation among the lines of RPCsu and MSCBMsu, presenting the last most significant variations (p<0.05). In the nitrite values, we identified only among the lines, in the passages P1 and P2, with the highest averages displayed by the MSCBMsu lineage (p<0.05). The measurement of antioxidant system activity showed high standards, identifying differences only for GSH values, in the RPCsu lineage, in P3 (p<0.05). This study suggests that the maintenance of cell culture in the long term induces lower regulation of oxidative stress, and RPCsu presents higher genetic stability and lower oxidative stress than MSCBMsu during in vitro expansion