Induction Of Mesenchymal Stem Cells Research Articles

Induction of mesenchymal stem cell (MSC) differentiation from iPS cells using MSC medium.

Mesenchymal stem cells (MSCs) and induced pluripotent stem (iPS) cells have great potential as cell sources for tissue engineering and regenerative medicine. This study aimed to investigate whether iPS cells can be differentiated into MSCs using MSCGM, a commercially available MSC culture system. The cells were characterized by flow cytometry, immunostaining, and gene expression analyses. We also examined their potential to differentiate into osteoblasts and chondrocytes. Our results showed that iPS cells cultured in MSCGM (iPS-MSCGM) exhibited a fibroblast-like morphology and expressed CD73 and CD90 genes, as well as positive markers for CD73, CD90, and CD105. Moreover, iPS-MSCGM cells demonstrated the ability to differentiate into osteoblasts and chondrocytes in vitro. This study demonstrates a new and simple method for inducing the differentiation of iPS cells to MSCs using MSCGM.

Differentiation Induction of Mesenchymal Stem Cells by a Au Delivery Platform.

Au decorated with type I collagen (Col) was used as a core material to cross-link with stromal cell-derived factor 1α (SDF1α) in order to investigate biological performance. The Au-based nanoparticles were subjected to physicochemical determination using scanning electron microscopy (SEM), dynamic light scattering (DLS) and ultraviolet-visible (UV-Vis) and Fourier-transform infrared spectroscopy (FTIR). Mesenchymal stem cells (MSCs) were used to evaluate the biocompatibility of this nanoparticle using the MTT assay and measuring reactive oxygen species (ROS) production. Also, the biological effects of the SDF-1α-conjugated nanoparticles (Au-Col-SDF1α) were assessed and the mechanisms were explored. Furthermore, we investigated the cell differentiation-inducing potential of these conjugated nanoparticles on MSCs toward endothelial cells, neurons, osteoblasts and adipocytes. We then ultimately explored the process of cell entry and transportation of the nanoparticles. Using a mouse animal model and retro-orbital sinus injection, we traced in vivo biodistribution to determine the biosafety of the Au-Col-SDF1α nanoparticles. In summary, our results indicate that Au-Col is a promising drug delivery system; it can be used to carry SDF1α to improve MSC therapeutic efficiency.

Role of Primary Cilia in Skeletal Disorders.

Primary cilia are highly conserved microtubule-based organelles that project from the cell surface into the extracellular environment and play important roles in mechanosensation, mechanotransduction, polarity maintenance, and cell behaviors during organ development and pathological changes. Intraflagellar transport (IFT) proteins are essential for cilium formation and function. The skeletal system consists of bones and connective tissue, including cartilage, tendons, and ligaments, providing support, stability, and movement to the body. Great progress has been achieved in primary cilia and skeletal disorders in recent decades. Increasing evidence suggests that cells with cilium defects in the skeletal system can cause numerous human diseases. Moreover, specific deletion of ciliary proteins in skeletal tissues with different Cre mice resulted in diverse malformations, suggesting that primary cilia are involved in the development of skeletal diseases. In addition, the intact of primary cilium is essential to osteogenic/chondrogenic induction of mesenchymal stem cells, regarded as a promising target for clinical intervention for skeletal disorders. In this review, we summarized the role of primary cilia and ciliary proteins in the pathogenesis of skeletal diseases, including osteoporosis, bone/cartilage tumor, osteoarthritis, intervertebral disc degeneration, spine scoliosis, and other cilium-related skeletal diseases, and highlighted their promising treatment methods, including using mesenchymal stem cells. Our review tries to present evidence for primary cilium as a promising target for clinical intervention for skeletal diseases.

Autologous platelet concentrates in regenerative endodontic treatment

Introduction: Autologous platelet concentrates, have recently emerged as a possible tool to improve regenerative procedures in the medical field. Objective: To analyze the literature on autologous platelet concentrates in regenerative endodontics, such as: Platelet Rich Plasma, Platelet Rich Fibrin, PRP/PRF Vs Blood Clot as a scaffold in regenerative endodontics. Methodology: Using the keywords Regenerative Endodontic Treatment, Autologous Platelet Concentrates, Immature Teeth, Platelet-rich Fibrin, Platelet-rich Plasma, Revitalization Endodontics, the MEDLINE/PubMed and ScienceDirect databases were searched, with emphasis on the last 5 years. It was evaluated with the PRISMA and AMSTAR-2 guidelines. Results: Platelet-rich plasma releases growth factors during the first hours after placement and induction of mesenchymal stem cells. Platelet Rich Fibrin is a second-generation platelet concentrate that releases growth factors more slowly, forming a network that traps leukocytes and platelets, liquid platelet rich fibrin has a higher concentration of leukocytes, which enhances immune defense in LPS-induced inflammation. Compared to blood clot, PRP and PRF show improved healing results, reducing infection and inflammation. Conclusion: The use of autologous platelet concentrates presents a more controlled scaffold in regenerative endodontics, with successful results, such as periapical healing, root thickening and apical closure.

Mg-BGNs/DCECM Composite Scaffold for Cartilage Regeneration: A Preliminary In Vitro Study.

Cartilage lesions can lead to progressive cartilage degeneration; moreover, they involve the subchondral bone, resulting in osteoarthritis (OA) onset and progression. Bioactive glasses, with the dual function of supporting both bone and cartilage regeneration, have become a promising biomaterial for cartilage/bone engineering applications. This is especially true for those containing therapeutic ions, which act as ion delivery systems and may further promote cartilage repair. In this study, we successfully fabricated Mg-containing bioactive glass nanospheres (Mg-BGNs) and constructed three different scaffolds, DCECM, Mg-BGNs-1/DCECM (1% Mg-BGNs), and Mg-BGNs-2/DCECM (10% Mg-BGNs) scaffold, by incorporating Mg-BGNs into decellularized cartilage extracellular matrix (DCECM). All three scaffolds showed favorable microarchitectural and ion controlled-release properties within the ideal range of pore size for tissue engineering applications. Furthermore, all scaffolds showed excellent biocompatibility and no signs of toxicity. Most importantly, the addition of Mg-BGNs to the DCECM scaffolds significantly promoted cell proliferation and enhanced chondrogenic differentiation induction of mesenchymal stem cells (MSCs) in pellet culture in a dose-dependent manner. Collectively, the multifunctional Mg-BGNs/DCECM composite scaffold not only demonstrated biocompatibility but also a significant chondrogenic response. Our study suggests that the Mg-BGNs/DCECM composite scaffold would be a promising tissue engineering tool for osteochondral lesions, with the ability to simultaneously stimulate articular cartilage and subchondral bone regeneration.

Generation of a novel model of bioengineered human oral mucosa with increased vascularization potential.

ObjectiveThe 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.BackgroundOral 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.MethodsThree 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.ConclusionOur 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.

Assessment of a novel activity of Acacia nilotica leaf extract for chondrogenesis induction of Mesenchymal stem cells for dental applications

BackgroundThe Dearth of the self-rejuvenation capability of cartilage tissue is one of the frequent problems encountered by the dentist and orthopedic surgeons. Acacia nilotica commonly known as (babul, thorny acacia, Egyptian acacia or thorn mimosa) has wide range of medicinal uses. The current study was carried out to investigate the herbal-based chondrogenesis inducing ability of leaves extract of A. nilotica in Mesenchymal stem cells (MSCs). PurposeTo investigate the ability of Acacia nilotica leaves extract in chondrogenesis induction from Mesenchymal stem cells Study designThe experimental were designed according to the completely randomized design (CRD). MethodsDifferent concentration of water and ethanol extract of plant leaves were tested for its inhibitory activity on HDP-SC. The HDP-SCs were stimulated to differentiate into chondrocytes in the presence of water extract of A. nilotica leaves. The study was accomplished by assessing the glycosaminoglycan (GAG), aggrecan, collagen 2α1 (Col2α1), and sox9 in the cellular matrix required for cartilage tissue formation. The MSCs revealed surface markers like CD45, CD90, CD105, and CD166 cultured on specialized chondrogenic differentiation medium in presence of leaves extract of A. nilotica. ResultsThe aqueous leaves extract showed a time-dependent increase in the concentration of glycosaminoglycan (GAG) deposition on 7th, 14th, and 21st day of incubation. The chondrogenic specific markers like aggrecan and col2α1were up-regulated from 7th to 21st days of incubation showing the highest protein expression on the 21st day. The sox9 protein was substantially enhanced on the 7th to 21thday of incubation of HDP-SC with leaves extract of A. nilotica compared to standard TGF-β inducer. ConclusionThe results demonstrate that aqueous leaves extract of A. nilotica promotes chondrogenesis of Mesenchymal stem cells. The study suggests that water leaves extract of “Babool"can be used as an alternative strategy for treatment of dental pulp through chondrogenic differentiation. A. nilotica as a source of many active secondary metabolites which may serve as potential candidates for development of novel lead compounds. This research confirmed the usefulness of this plant A. nilotica as a characteristic feature of medical plant and pharmaceutical. Further studies in the future are required to identify the role of the rest of the active compounds.

Rapid Induction of Mesenchymal Stem Cells to Neural Cells

Background:-Human umbilical cord blood- mesenchymal stem cells(HUCB-MSCs) hold great promise invitro neural differentiation and therapy for neurodegenerative disorders. It has demonstrated that the numberand differentiating potential of bone marrow mesenchymal stem cells (MSCs) decrease with age. Therefore,the search for alternative sources of MSCs is of significant value.Objective:-To determine the possibility of obtaining clonally expanded UCB-MSCs and evaluate their rapidinduction and differentiation into neural cells in vitro.Methods:-MSCs were isolated from HUCB by combining gradient density centrifugation with plasticadherence. In order to identify factors that able to lead to neural differentiation, the cultured cells weretreated with retinoic acid (RA) and B-mercaptoethanol (BME). Differentiating characterization of UCBMSCswere detected by immunocytochemistry analysis.Results:-UCB-MSCs appeared like the fibroblast cell and these cells were extensively expanded in culture.The results showed that UCB-MSCs were positive for CD71and CD90 but were negative for CD34. Theimmunocytochemistry staining indicated that the differentiated cells give positive response for nestinemarker. The result confirmed for neural progenitor differentiation of MSCs.Conclusion: The results of this study confirmed that UCB provides a great source of stem cells for using intreatment of neurodegenerative disorders of the central nervous system.

Akt signaling is activated by TGF\u03b22 and impacts tenogenic induction of mesenchymal stem cells

BackgroundTissue engineered and regenerative approaches for treating tendon injuries are challenged by the limited information on the cellular signaling pathways driving tenogenic differentiation of stem cells. Members of the transforming growth factor (TGF) β family, particularly TGFβ2, play a role in tenogenesis, which may proceed via Smad-mediated signaling. However, recent evidence suggests some aspects of tenogenesis may be independent of Smad signaling, and other pathways potentially involved in tenogenesis are understudied. Here, we examined the role of Akt/mTORC1/P70S6K signaling in early TGFβ2-induced tenogenesis of mesenchymal stem cells (MSCs) and evaluated TGFβ2-induced tenogenic differentiation when Smad3 is inhibited.MethodsMouse MSCs were treated with TGFβ2 to induce tenogenesis, and Akt or Smad3 signaling was chemically inhibited using the Akt inhibitor, MK-2206, or the Smad3 inhibitor, SIS3. Effects of TGFβ2 alone and in combination with these inhibitors on the activation of Akt signaling and its downstream targets mTOR and P70S6K were quantified using western blot analysis, and cell morphology was assessed using confocal microscopy. Levels of the tendon marker protein, tenomodulin, were also assessed.ResultsTGFβ2 alone activated Akt signaling during early tenogenic induction. Chemically inhibiting Akt prevented increases in tenomodulin and attenuated tenogenic morphology of the MSCs in response to TGFβ2. Chemically inhibiting Smad3 did not prevent tenogenesis, but appeared to accelerate it. MSCs treated with both TGFβ2 and SIS3 produced significantly higher levels of tenomodulin at 7 days and morphology appeared tenogenic, with localized cell alignment and elongation. Finally, inhibiting Smad3 did not appear to impact Akt signaling, suggesting that Akt may allow TGFβ2-induced tenogenesis to proceed during disruption of Smad3 signaling.ConclusionsThese findings show that Akt signaling plays a role in TGFβ2-induced tenogenesis and that tenogenesis of MSCs can be initiated by TGFβ2 during disruption of Smad3 signaling. These findings provide new insights into the signaling pathways that regulate tenogenic induction in stem cells.

Hypoxic-Mediated Oxidative Stress Condition and Hydroxyapatite-Inducing Osteogenic Differentiation of Human Mesenchymal Stem Cells: A Mathematical Modeling Study.

Avascular necrosis (AVN) of the bones remains a major clinical challenge. Fractures in the talus, the scaphoid, and the neck of the femur are especially challenging to heal due to the low blood vessel network and the lack of collateral blood supply. These fractures are associated with high rates of nonunion and increased infections that require repeated operations. Conventional treatments by autografting or allografting bone replacement and synthetic bone implants have limitations, including the invasiveness of operative procedures, tissue supply insufficiency, and the risk of host rejection. The advancement in tissue engineering has revealed the potential of stem cells as restorative agents for bone injuries. The administration of mesenchymal stem cells (MSCs) into the talus, the scaphoid, and the neck of the femur could produce enhanced osteogenesis via the manipulation of MSC culture conditions. In this study, we used hydroxyapatite as the nanomaterial, and hypoxic milieu to enhance MSC differentiation capacity into the osteogenic lineage, allowing for more rapid and efficient bone cell replacement treatment. Our results demonstrate 1% oxygen and 12.5 μg/mL of hydroxyapatite (HAP) as the optimal conditions to incorporate the osteogenic medium for the osteogenic induction of MSCs. We also established a proof of concept that the addition of HAP and hypoxic conditions could augment the osteoinductive capacity of MSCs. We also developed an accurate mathematical model to support future bone cell replacement therapy.

Differentiation of Human Mesenchymal Stem Cells from Wharton's Jelly Towards Neural Stem Cells Using A Feasible and Repeatable Protocol.

The transplantation of neural stem cells (NSCs) capable of regenerating to the cells of the central nervous system (CNS) is a promising strategy in the treatment of CNS diseases and injury. As previous studies have highlighted mesenchymal stem cells (MSCs) as a source of NSCs, this study aimed to develop a feasible, efficient, and reproducible method for the neural induction of MSCs isolated from Wharton’s jelly (hWJ-MSCs). We induced neural differentiation in a monolayer culture using epidermal growth factor, basic fibroblast growth factor, N2, and B27 supplements. This resulted in a homogenous population of proliferating cells that expressed certain neural markers at both the protein and mRNA levels. Flow cytometry and immunocytochemistry confirmed the expression of neural markers: nestin, sex-determining region Y (SRY) box 1 and 2 (SOX1 and SOX2), microtubule-associated protein 2 (MAP2), and glial fibrillary acidic protein (GFAP). The qRT-PCR analysis revealed significantly enhanced expression of nestin and MAP2 in differentiated cells. This study confirms that it is possible to generate NSCs-like cells from hWJ-MSCs in a 2D culture using a practical method. However, the therapeutic effectiveness of such differentiated cells should be extended to confirm the terminal differentiation ability and electrophysiological properties of neurons derived from them.

The effect of growth factors on induction of mesenchymal stem cell to the skeletal muscle cells.

The objective of this study was to evaluate the effect of IGF1 and FGF2 growth factors on the differentiation of human adipose tissue‑derived mesenchymal stem cells (MSCs) into skeletal muscle cells. MSCs were divided into four groups. Group I (control group) was incubated only in myogenic medium, group II was incubated by adding 100 ng/ml FGF2 to the myogenic medium, group III was incubated by adding 20 ng/ml IGF1 to the myogenic medium, group IV was incubated by adding 100 ng/ml FGF2 and 20 ng/ml IGF1 to the myogenic medium. Cells dyed with MyoD1 antibodies were analyzed by flow cytometry so as to determine the myogenic differentiation on day 28. It was confirmed that 11.5 % of the control group, 39.2 % of the FGF2 group, 23.1 % of the IGF1 group, and 39.3 % of the IGF1-FGF2 group showed differentiation. Our results show that IGF1 and FGF2 have a positive contribution to myogenic differentiation of MSCs. They contribute to the studies related to muscle diseases and their treatment by the fact that growth factors support the feature of regeneration capacity and differentiation (Tab. 2, Fig. 7, Ref. 23).

Neurogenin-1 Overexpression Increases the Therapeutic Effects of Mesenchymal Stem Cells through Enhanced Engraftment in an Ischemic Rat Brain.

Background and ObjectivesStem cell therapy is a promising strategy for treating neurological diseases but its effectiveness is influenced by the route of administration and the characteristics of the stem cells. We determined whether neural induction of mesenchymal stem cells (MSCs) was beneficial when the cells were delivered intra-arterially through the carotid artery.Methods and ResultsMSCs were neurally induced using a retroviral vector expressing the neurogenic transcription factor neurogenin-1 (Ngn1). The LacZ gene encoding bacterial β-galactosidase was used as a control. Ischemic stroke was induced by transluminal occlusion of the middle cerebral artery and 3 days later the MSCs were delivered intra-arterially through the internal carotid artery. Magnetic resonance imaging analysis indicated that compared to MSCs expressing LacZ (MSCs/LacZ), MSCs expressing Ngn1 (MSCs/Ngn1) exhibited increased recruitment to the ischemic region and populated this area for a longer duration. Immunohistochemical analysis indicated that compared to MSCs/LacZ, MSCs/Ngn1 more effectively alleviated neurological dysfunction by blocking secondary damage associated with neuronal cell death and brain inflammation. Microarray and real-time PCR analysis indicated that MSCs/Ngn1 exhibited increased expression of chemotactic cytokine receptors, adherence to endothelial cells, and migration ability.ConclusionsNeural induction with Ngn1 increases the homing ability of MSCs, enhancing their engraftment efficiency in the ischemic rat brain. Intra-arterial delivery of neurally induced MSCs/Ngn1 3 days after ischemic injury blocks neuronal cell death and inflammation, and improves functional recovery. Thus, intra-arterial administration of stem cells with neural properties may be a novel therapy for the treatment of ischemic stroke.

Wharton's Jelly Mesenchymal Stem Cell: Various Protocols for Isolation and Differentiation of Hepatocyte-Like Cells; Narrative Review.

There are several differentiation methods for mesenchymal stem cells (MSCs) into hepatocyte-like cell. Investigators reported various hepatic differentiation protocols such as modifying culturing conditions or using various growth factors/cytokines. In this literature review, we compared different MSCs extraction and isolation protocols from Wharton’s jelly (WJ) and explored various MSCs differentiation methods. Various protocols have been recommended for MSCs isolated from WJ, such as enzymatic, enzymatic-explant, and explant methods. In the explant method, valuable time is wasted, but the cost and biological contaminations are reduced and the number of isolated cells is high. However, other features, such as immune phenotype and multiline-age differentiation capacity, do not differ from other methods. There are also several differentiation methods for hepatocyte-like cell including the induction of MSC by cytokines and growth factors, and the differentiation of MSC in 2- and 3-dimensional matrix (2D and 3D). Among several cytokines, hepatocyte growth factor (HGF) and fibroblast growth factor (FGF) are essential. In the early stage of the differentiation, 2D culture is useful, and in the development stage, 3D culture system with HGF and FGF cytokines are more effective in the process of differentiation. Some studies have used 3D culture system in biocompatible scaffolds, such as alginate, collagen, gelatin, and peptide-Gly-Leu-amide (PGLA). In conclusion, Wharton’s jelly-Mesenchymal stem cells (WJ-MSCs) can be considered as an appropriate source for hepatocyte differentiation. Moreover, we introduced the explant method as the most effective protocol. This review attempted to highlight factors in hepatocyte differentiation, but the most effective protocol is not still unknown.

Use of CRISPR/dCas9 system for induction of mesenchymal stem cells

Use of CRISPR/dCas9 system for induction of mesenchymal stem cells

Strontium released bi-lineage scaffolds with immunomodulatory properties induce a pro-regenerative environment for osteochondral regeneration

The different lineage-specific biological properties of articular cartilage and subchondral bone present a great challenge in the construction of bi-lineage scaffolds for simultaneous osteochondral regeneration. To overcome this challenge, strontium incorporated calcium silicate (Sr-CS) ceramic was prepared for bi-lineage formation of scaffolds in this study. The positive result of Sr-CS in the regeneration of osteochondral defects was first proven by its improved effect on the osteogenesis and chondrogenesis induction of mesenchymal stem cells (MSCs). After that, scaffold-mediated macrophage polarization between classically activated inflammatory macrophages (termed M1Ф) and alternatively activated inflammatory macrophages (termed M2Ф) was assayed to investigate whether the incorporation of Sr into calcium silicate could alter host-to-scaffold immune response. Furthermore, the interactions between Sr-CS pretreated macrophages and MSCs differentiation were performed to prove the enhancement effect of suppressed inflammatory response on osteogenesis and chondrogenesis. In vivo transplantation showed that the Sr-CS scaffolds distinctly improved the regeneration of cartilage and subchondral bone, as compared to the calcium silicate scaffolds. On the one hand, the mechanism attributes to enhancement of strontium on the osteogenic and chondrogenic differentiation of MSCs. On the other hand, the reason can partially be attributed to suppressed synovial inflammatory response, which has improved effects on enhancement of osteogenesis and chondrogenesis. These findings suggest that monophasic Sr-CS scaffolds with a bi-lineage conducive property and an inflammatory response regulatory property represents a viable strategy for simultaneous regeneration of osteochondral defects.

Differentiation of Porcine Mesenchymal Stem Cells to Lung Epithelial Cells Using Media Conditioned by Human Primary Small Airway Epithelial Cells

Mesenchymal stem cells (MSCs) under defined conditions, can differentiate into many different cell types, and may be the key to treating a multitude of ailments including chronic lung injury. Although MSCs can reduce inflammation and alter immune response when injected into sites of injury, differentiating MSCs in vitro prior to injection may enhance their therapeutic actions. The aim of this study was to develop a simple and reliable method to differentiate porcine adipose‐derived (pAMSCs) and bone marrow‐derived mesenchymal stem cells (pBMSCs) to small airway epithelium (SAE) for future testing in a pig model of lung injury. Traditional induction of MSCs to SAE involves treatment with an artificial combination of factors that mimic the local milieu in vivo. A simpler and more physiologically‐relevant approach is to use naturally conditioned media that may yield more biologically active cells. We hypothesized that media conditioned with human primary small airway epithelial cells (hSAECs) can induce differentiation of pAMSCs and pBMSCs to SAE. hSAECs (ATCC) were seeded in serum‐free M199 media to create conditioned media which was collected 4 days after seeding for immediate use. pAMSCs and pBMSCs were isolated in‐house from juvenile Yorkshire pigs, and expanded to passage 3 prior to start of experiments. Fibronectin‐coated, 8‐well slides (for immunohistochemistry) or T75 flasks (to examine morphology) were seeded with either pAMSCs or pBMSCs (10,000 cells/cm2). Every 4 days for 16 days, a subset of cells were labeled with stem cell markers (anti‐FOXA1, anti‐CD44, anti‐CD105), small airway cell markers (anti‐SCGB3A2, anti‐RAGE), and neurofilament markers (anti‐neurofilament heavy polypeptide, anti‐68kDa neurofilament)), and media was exchanged with either conditioned media or unconditioned media. After 5 days, pBMSCs exposed to conditioned media transitioned to cuboidal and epithelial‐like cells. pBMSCs exposed to unconditioned media remained elongated and stem cell‐like. pAMSCs proliferated faster than pBMSCs, but yielded spindly overgrown cells in high density that did not differentiate in either conditioned or unconditioned media. Expression of proteins did not correlate with the morphological changes in pBMSCs. Although RAGE expression increased initially in response to conditioned media, levels were reduced after day 12. SCGB3A2 expression also increased over time, but was not different between the conditioned and unconditioned groups. The distinct change in morphology observed in pBMSCs exposed to conditioned media indicates a promising phenotypic change. Verification of the epithelial cell phenotype induced by the conditioned media with semi‐quantitative gene and protein expression of cell markers for small airway cells and their functional ability to produce surfactant remains to be determined. Our study indicates that conditioned media is able to induce differentiation of MSCs into epithelial cells. Further, our results revealed that the source of MSCs (adipose versus bone marrow), and the density of MSCs may be important factors in successful differentiation of MSCs.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

In Vitro Induction of Tendon-Specific Markers in Tendon Cells, Adipose- and Bone Marrow-Derived Stem Cells is Dependent on TGFβ3, BMP-12 and Ascorbic Acid Stimulation.

Mesenchymal Stem Cells (MSCs) and tissue-specific progenitors have been proposed as useful tools for regenerative medicine approaches in bone, cartilage and tendon-related pathologies. The differentiation of cells towards the desired, target tissue-specific lineage has demonstrated advantages in the application of cell therapies and tissue engineering. Unlike osteogenic and chondrogenic differentiation, there is no consensus on the best tenogenic induction protocol. Many growth factors have been proposed for this purpose, including BMP-12, b-FGF, TGF-β3, CTGF, IGF-1 and ascorbic acid (AA). In this study, different combinations of these growth factors have been tested in the context of a two-step differentiation protocol, in order to define their contribution to the induction and maintenance of tendon marker expression in adipose tissue and bone marrow derived MSCs and tendon cells (TCs), respectively. Our results demonstrate that TGF-β3 is the main inducer of scleraxis, an early expressed tendon marker, while at the same time inhibiting tendon markers normally expressed later, such as decorin. In contrast, we find that decorin is induced by BMP-12, b-FGF and AA. Our results provide new insights into the effect of different factors on the tenogenic induction of MSCs and TCs, highlighting the importance of differential timing in TGF-β3 stimulation.

Behavior of Mesenchymal Stem Cells Obtained From Dental Tissues: A Review of the Literature

The success of tissue engineering in combination with tissue regeneration depends on the behavior and cellular activity in the biological processes developed within a structure that functions as a support, better known as scaffolds, or directly at the site of the injury. The cell-cell and cell-biomaterial interaction are key factors for the induction of a specific cell behavior, together with the bioactive factors that allow the formation of the desired tissue. Mesenchymal Stem Cells (MSC) can be isolated from the umbilical cord and bone marrow; however, the behavior of Dental Pulp Stem Cells (DPSC) has been shown to have a high potential for the formation of bone tissue, and these cells have even been able to induce the process of angiogenesis. Advances in periodontal regeneration, dentin-pulp complex, and craniofacial bone defects through the induction of MSC obtained from tooth structures in in vitro-in vivo studies have permitted the obtaining of clinical evidence of the achievements obtained to date.

TGFβ2-induced tenogenesis impacts cadherin and connexin cell-cell junction proteins in mesenchymal stem cells

Tenogenic differentiation of stem cells is needed for tendon tissue engineering approaches. A current challenge is the limited information on the cellular-level changes during tenogenic induction. Tendon cells in embryonic and adult tendons possess an array of cell-cell junction proteins that include cadherins and connexins, but how these proteins are impacted by tenogenic differentiation is unknown. Our objective was to explore how tenogenic induction of mesenchymal stem cells (MSCs) using the transforming growth factor (TGF)β2 impacted protein markers of tendon differentiation and protein levels of N-cadherin, cadherin-11 and connexin-43. MSCs were treated with TGFβ2 for 21 days. At 3 days, TGFβ2-treated MSCs developed a fibroblastic morphology and significantly decreased levels of N-cadherin protein, which were maintained through 21 days. Similar decreases in protein levels were found for cadherin-11. Connexin-43 protein levels significantly increased at 3 days, but then decreased below control levels, though not significantly. Protein levels of scleraxis and tenomodulin were significantly increased at day 14 and 21, respectively. Taken together, our results indicate that TGFβ2 is an inducer of tendon marker proteins (scleraxis and tenomodulin) in MSCs and that tenogenesis alters the protein levels of N-cadherin, cadherin-11 and connexin-43. These findings suggest a role for connexin-43 early in tenogenesis, and show that early-onset and sustained decreases in N-cadherin and cadherin-11 may be novel markers of tenogenesis in MSCs.