Multiple Differentiation Capacity Research Articles

The Role of Mesenchymal Stromal Cells-Derived Small Extracellular Vesicles in Diabetes and Its Chronic Complications

Mesenchymal stromal cells (MSCs) are applied in regenerative medicine of several tissues and organs nowadays by virtue of their self-renewal capabilities, multiple differentiation capacity, potent immunomodulatory properties, and their ability to be favourably cultured and manipulated. With the continuous development of “cell-free therapy” research, MSC-derived small extracellular vesicles (MSC-sEVs) have increasingly become a research hotspot in the treatment of various diseases. Small extracellular vesicles (SEVs) are membrane vesicles with diameters of 30 to 150 nm that mediate signal transduction between adjacent or distal cells or organs by delivering non-coding RNA, protein, and DNA. The contents and effects of sEVs vary depending on the properties of the originating cell. In recent years, MSC-sEVs have been found to play an important role in the occurrence and development of diabetes mellitus as a new way of communication between cells. Diabetes mellitus is a common metabolic disease in clinic. Its complications of the heart, brain, kidney, eyes, and peripheral nerves are a serious threat to human health and has been a hot issue for clinicians. MSC-sEVs could be applied to repair or prevent damage from the complications of diabetes mellitus through anti-inflammatory effects, reduction of endoplasmic reticulum-related protein stress, polarization of M2 macrophages, and increasing autophagy. Therefore, we highly recommend that MSC-sEVs-based therapies to treat diabetes mellitus and its chronic complication be further explored. The analysis of the role and molecular mechanisms of MSC-sEVs in diabetes and its related complications will provide new idea and insights for the prevention and treatment of diabetes.

Identification of alpha-enolase as a potential immunogenic molecule during allogeneic transplantation of human adipose-derived mesenchymal stromal cells

Background aimsGiven their low immunogenicity, immunoregulatory effects and multiple differentiation capacity, mesenchymal stromal cells (MSCs) have the potential to be used for “off-the-shelf” cell therapy to treat various diseases. However, the allorejection of MSCs indicates that they are not fully immune-privileged. In this study, the authors investigated the immunogenicity of human adipose-derived MSCs (Ad-MSCs) and identified potential immunogenic molecules. MethodsTo evaluate the immunogenicity of human Ad-MSCs in vivo, cells were transplanted into humanized mice (hu-mice), then T-cell infiltration and clearance of human Ad-MSCs were observed by immunofluorescence and bioluminescence imaging. One-way mixed lymphocyte reaction and flow cytometry were performed to evaluate the immunogenicity of human Ad-MSCs in vitro. High-throughput T-cell receptor (TCR) repertoire sequencing and mass spectrometry were applied to identified potential immunogenic molecules. ResultsThe authors observed that allogeneic Ad-MSCs recruited human T cells and caused faster clearance in hu-mice than non-humanized NOD.Cg-Prkdcscid IL2rgtm1Wjl/SzJ (NSG) mice. The proliferation and activation of T cells were significantly enhanced during in vitro co-culture with human Ad-MSCs. In addition, the level of HLA-II expression on human Ad-MSCs was dramatically increased after co-culture with human peripheral blood mononuclear cells (PBMCs). High-throughput sequencing was applied to analyze the TCR repertoire of the Ad-MSC-recruited T cells to identify dominant TCR CDR3 sequences. Using synthesized TCR CDR3 peptides, the authors identified several potential immunogenic candidates, including alpha-enolase (ENO1). The ENO1 expression level of Ad-MSCs significantly increased after co-culture with PBMCs, whereas ENO1 inhibitor (ENOblock) treatment decreased the expression level of ENO1 and Ad-MSC-induced proliferation of T cells. ConclusionsThe authors’ findings improve the understanding of the immunogenicity of human Ad-MSCs and provide a theoretical basis for the safe clinical application of allogeneic MSC therapy.

Neuropathy in the Altered Bone Marrow Microenvironment of Patients with Essential Thrombocythemia and the Effects of Interferonα-2b on Them

Background: ET is one of the classical Philadelphia chromosome-negative MPNs . BM-MSCs may involve in the pathogenesis of MPN. Bone marrow neural tissue, closely related to hematological diseases , is composed of sympathetic nervous system (SNS) fibres , ensheathing Schwann cells, supporting nestin+ MSCs . There has been a longstanding interest in using interferon (IFN) to control myeloproliferation in patients with ET. Until now, little is known about whether IFN had effects on the BM-MSCs from patients with ET.Objective: This study aims to systematically and thoroughly assess the biological and functional characteristics of BM-MSCs and SNS in patients with ET and explored the effects of IFNα-2b exerted on them.Methods: (1) The morphology of BM-MSCs was observed under an inverted microscope. (2) The immunophenotype markers of BM-MSCs were examined by flow cytometry. (3) BM-MSCs were induced to differentiate into adipocytes and osteoblasts, which were identified with corresponding staining. (4) The proliferation rate, senescence and apoptosis of BM-MSCs was detected.(5)Nestin+cell number ,sympathetic nerve fibres , schwann cells and nestin mRNA expression were detected by IHC and RT-PCR respectively.(6)CXCL12 and norepinephrine was detected by ELISA . (7) Umbilical cord derived CD34+ cells were purified and co-cultured with BM-MSCs to assess the function of BM-MSCs to support hematopoiesis . Hematopoietic growth factors such as SCF, IL-6 and VEGF were detected by ELISA . (8) Peripheral blood mononuclear cells (PBMCs) were isolated from normal controls and co-cultured with BM-MSCs to assess the suppression capacity of BM-MSCs in the proliferation of PBMCs. PBMCs were also co-cultured with BM-MSCs to assess the contribution of BM-MSCs in promoting the proliferation of Tregs. (9) Compared with ET derived BM-MSCs in the absence of IFNα-2b, the ET derived BM-MSCs were examined the changes in above biological and functional characteristics with the presence of IFNα-2b.Results: (1) No significant differences were found in morphology between BM-MSCs from patients with ET and normal controls. Both BM-MSCs exhibited bipolar spindle-like cells with a whirlpool-like array. (2) BM-MSCs from patients with ET and normal controls also had no discrepancies in immunophenotype characteristics. (3) BM-MSCs from ET showed increased proliferation, lower apoptosis capacity and decreased senescence. (4)In MPN patients, nestin+cell number and nestin messenger RNA expression were markedly increased. (5)Nevertheless , BM-MSCs from ET patients expressed lower levels CXCL12 than healthy controls. (6)Sympathetic nerve fibres and ensheathing Schwann cells were markedly reduced in BM of ET patients , with lower levels of norepinephrine secreted. (7) BM-MSCs from ET patients were more difficult to induce to osteoblasts than healthy controls. (8) Both BM-MSCs from ET patients and healthy controls had the potential to support hematopoiesis. However, BM-MSCs from ET patients had reduced capacity to amplify CD34+ cells and maintain long-term hematopoiesis. BM-MSCs from ET patients expressed lower levels SCF, IL-6 and VEGF than healthy controls. (9) BM-MSCs from ET were defective to suppress the proliferation of PBMCs and could promote PBMCs to differentiate into regulatory T cells. (10) IFNα-2b had no effect on the morphology, immunophenotype and JAK2 mutant allele of BM-MSCs derived from ET patients. IFNα-2b was capable of suppressing the proliferation of BM-MSCs, but increasing its apoptosis and its function of supporting hematopoiesis. IFNα-2b also ameliorated its defective in inhibiting the proliferation of PBMCs and promotes PBMCs to differentiate into regulatory T cells.Conclusion: (1) BM-MSCs from ET patients were multiply defective both in biological properties and functions. (2) BM-MSCs from ET were abnormal in proliferation,senescence, apoptosis and differentiation. (3) BM-MSCs from ET had reduced capacity to maintain hematopoiesis. (4) BM-MSCs from ET were defective in maintaining immune homeostasis. (5) Neuropathy is obvious in ET patients. (6) IFNα-2b affects BM-MSCs from ET patients in multiple differentiation capacities, proliferation, apoptosis and the capacity to support long-term hematopoiesis and immune regulation. DisclosuresNo relevant conflicts of interest to declare.

Mesenchymal stem cell-derived small extracellular vesicles and bone regeneration.

Mesenchymal stem cells (MSCs) and MSC‐derived small extracellular vesicles (sEVs) are promising candidates for cell‐based and cell‐free regenerative medicine, respectively. By virtue of their multiple lineage differentiation capacity, MSCs have been implicated as an ideal tool for bone and cartilage regeneration. However, later observations attributed such regenerative effects to MSC‐secreted paracrine factors. Exosomes, endosomal originated sEVs carrying lipid, protein and nucleic acid cargoes, were identified as components of the MSC secretome and propagated the key regenerative and immunoregulatory characteristics of parental MSCs. Here, exosome biogenesis, the molecular composition of exosomes, sEV‐cell interactions and the effects on key bone homeostasis cells are reviewed. MSC‐derived sEVs show to promote neovascularization and bone and cartilage regeneration in preclinical disease models. The mechanisms include the transfer of molecules, including microRNAs, mRNAs and proteins, to other key cells. MSC‐derived sEVs are interesting candidates as biopharmaceuticals for drug delivery and for the engineering of biologically functionalized materials. Although major exploratory efforts have been made for therapeutic development, the secretion, distribution and biological effects of MSC‐derived sEVs in bone and cartilage regeneration are not fully understood. Moreover, techniques for high‐yield production, purity and storage need to be optimized before effective and safe MSC‐derived sEVs therapies are realized.

Investigation of platelet-rich plasma in increasing proliferation and migration of endometrial mesenchymal stem cells and improving pregnancy outcome of patients with thin endometrium.

Platelet-rich plasma (PRP) contains abundant growth factors and is gradually used in the field of reproduction. A thin endometrium is recognized as a critical factor in embryo implantation failure. Endometrial mesenchymal stem cells (EnMSCs), which were isolated from human menstrual blood, are highly proliferative and show multiple differentiation capacity. The current study was to investigate the effect of PRP on the proliferation and migration of EnMSCs, and the effectiveness of PRP in the treatment of patients with thin endometrium. EnMSCs were treated with PRP in vitro, followed by measuring cell proliferation, migration, and adhesion by using CCK8, scratch, and adhesion test, respectively. Twenty patients undergoing in vitro fertilization (IVF) with refractory thin endometrium history were given PRP by infusion into the uterine cavity after the treatment of hormone replacement therapy (HRT). All components of PRP significantly stimulated the growth, migration, and adhesion of EnMSCs when compared with the negative control. Cell proliferation and migration were induced by PRP in a dose-dependent manner with maximum proliferation at a 2% PRP dose. The clinical data showed that successful endometrial expansion and pregnancy were discovered in 12 patients after PRP infusion, and the pregnancy rate increased to 60%. Intrauterine PRP infusion represents a new way for female patients with thin endometrium with poor response. This study lays the foundations for the potential treatment of thin endometrium with PRP in vivo.

Effects of platelet-rich plasma on the activity of human menstrual blood-derived stromal cells in vitro

BackgroundHuman menstrual blood-derived stromal cells (MenSCs) are highly proliferative and show multiple differentiation capacity. The convenience and non-invasiveness make MenSC a novel cell source for regenerative medicine applications. Platelet-rich plasma (PRP) contains abundant growth factors which are beneficial to wound healing. However, the influence of PRP on MenSCs remains elusive. Here, we evaluated the role of PRP in MenSCs proliferation and assessed the effects of PRP on endometrial receptivity regulation in vitro.MethodsMenSCs cultured with 10% activated PRP were compared with those cultured with 10% fetal bovine serum (FBS). Differences in cell proliferation, differentiation, and endometrial receptivity-related gene expression were evaluated.ResultsNotably, 10% activated PRP significantly promoted MenSCs proliferation and adipogenic/osteogenic differentiation while suppressing apoptosis. Expression of the mesenchymal stem cells (MSCs) marker CD105 and the perivascular markers SUSD2 and CD146 were elevated after PRP treatment. Moreover, short-term PRP stimulation activated the phosphorylation of Akt and signal transducer and activator of transcription 3 (STAT3) pathways, upregulated expression of FoxO1, LIF, and IL1-β, and downregulated IL-6.ConclusionsIn summary, PRP could promote MenSC proliferation, markedly accelerate cell stemness, and evaluate MenSC functions by enhancing the expression of angiogenesis and endometrial receptivity markers, suggesting its potential use as a promising supplement for MenSCs in endometrial regenerative medicine. Our results provide a theoretical basis for the clinical application of co-transplantation of PRP combined with MenSCs.

Advances of mesenchymal stem cells in treatment of graft-versus-host disease

The development of graft-versus-host disease (GVHD) significantly affects the efficacy of allogeneic hematopoietic stem cell transplantation (allo-HSCT). Glucocorticoid combined with immunosuppressive agents is the first-line treatment of GVHD, but there are still some patients with poor response. Therefore, the treatment of GVHD requires more effective and safer methods. Mesenchymal stem cell (MSC) is a class of non-hematopoietic stem cells with self-renewal and multiple differentiation capacity. They can regulate immune cells and play immunosuppressive effects under certain conditions. MSC will be a new therapy method for GVHD in the future. In order to provide database of MSC application, this review literatures on advances of the effectiveness, dose, route of administration, efficacy factors, complications and other aspects of MSC in treatment of GVHD. Key words: Mesenchymal stem cells; Graft vs host disease; Treatment outcome

MALP-2, an agonist of TLR6, promotes the immune status without affecting the differentiation capacity of umbilical cord mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are increasingly used in cell-based therapy due to their multiple differentiation capacity, low expression of co-stimulatory factors and immunosuppressive effect. However, accumulating studies reported the recognition and rejection of engrafted MSCs, which eventually led to the fail of clinical trials. Toll-like receptors (TLRs) are important in mediating the immune response. In the present study, macrophage-activated lipopeptide-2 (MALP-2) was introduced to activate the TLR6 pathway in umbilical cord MSCs (UCMSCs). PBLs isolated from healthy volunteers were co-cultured with UCMSCs to measure whether activation of TLR6 of UCMSCs could stimulate immune responses. Reverse transcription-quantitative polymerase chain reaction and immunohistochemistry were performed to detect pro-inflammatory molecules and differentiation status of UCMSCs, respectively. The results indicated that activation of TLR6 in UCMSCs increased the proliferation of peripheral blood leukocytes (PBLs) and enhanced the release of lactate dehydrogenase in damaged UCMSCs, which confirmed the role of TLR6 in promoting the immunogenicity of UCMSCs. Furthermore, quantitative polymerase chain reaction demonstrated that the expression of proinflammatory molecules (including IL-1β, IL-6, IL-8, IL-10, CCL1 and CCL4) was induced, whereas the expression of stem cell markers (Klf4 and Nanog) was inhibited. The differentiation results indicated that activation of TLR6 had no effect on the differentiation capacity of UCMSCs. All these findings suggest that stimulation of TLR6 pathway may increase the immunogenicity of UCMSCs in in vitro detections. In conclusion, the results of the current study indicated a new role of TLR6 in regulating the biological function of UCMSCs.

Porphyromonas gingivalis can invade periodontal ligament stem cells

BackgroundPorphyromonas gingivalis is strongly associated with the development, progression, severity and recurrence of periodontitis. Periodontal ligament stem cells (PDLSCs) play an important role in the maintenance of periodontal tissue self-renewal and repair. The purpose of this study was to investigate the ability of P. gingivalis to infect PDLSCs using an in vitro monolayer model.MethodsWe separated and cultured primary PDLSCs using the tissue block with limiting dilution method. The efficiency of P. gingivalis (ATCC 33277) infection of PDLSCs was measured using agar plate culture and quantitative polymerase chain reaction (q-PCR) methods. PDLSCs infected with P. gingivalis were also observed by transmission electron microscopy.ResultsWe assessed stem cell properties including cell morphology, clone formation, growth activity, cell surface antigens and multiple differentiation capacity. The infection rates of P. gingivalis in PDLSC at MOIs of 50, 100, 200, and 500 were 5.83%, 8.12%, 7.77% and 7.53% according to the agar plate culture method. By q-PCR, the efficiencies of P. gingivalis infection of PDLSCs at MOIs of 50, 100, 200, and 500 were 6.74%, 10.56%, 10.36% and 9.78%, respectively. Overall, the infection efficiency based on q-PCR was higher than that according to agar plate culture. Using transmission electron microscopy, we verified that P. gingivalis (ATCC 33277) could infect and invade PDLSCs after 2 h of incubation, and endocytic vacuoles were not found surrounding the internalized bacteria.ConclusionsIn conclusion, our data demonstrate that P. gingivalis can invade PDLSCs.

Resveratrol augments therapeutic efficiency of mouse bone marrow mesenchymal stem cell-based therapy in experimental autoimmune encephalomyelitis

Experimental autoimmune encephalitis (EAE) is an inflammatory demyelinating disease, which served as a useful model providing considerable insights into the pathogenesis of multiple sclerosis (MS). Mouse bone marrow mesenchymal stem cells (mBM-MSC) were shown to have neuroprotection capabilities in EAE. Resveratrol is a small polyphenolic compound and possess therapeutic activity in various immune-mediated diseases. The sensitivity of mBM-MSCs to resveratrol was determined by an established cell-viability assay. Resveratrol-treated mBM-MSCs were also characterized with flow cytometry using MSC-specific surface markers and analyzed for their multiple differentiation capacities. EAE was induced in C57BL/6 mice by immunization with MOG35-55. Interferon gamma (IFN-γ)/tumor necrosis factor alpha (TNF-α) and interleukin-4 (IL-4)/interleukin-10 (IL-10), the hallmark cytokines that direct T helper type 1 (Th1) and Th2 development, were detected with enzyme-linked immunosorbent assay (ELISA). In vivo efficacy experiments showed that mBM-MSCs or resveratrol alone led to a significant reduction in clinical scores, and combined treatment resulted in even more prominent reduction. The combined treatment with mBM-MSCs and resveratrol enhanced the immunomodulatory effects, showing suppressed proinflammatory cytokines (IFN-γ, TNF-α) and increased anti-inflammatory cytokines (IL-4, IL-10). The combination of mBM-MSCs and resveratrol provides a novel potential experimental protocol for alleviating EAE symptoms.

Gold Nanoparticles Promote Proliferation of Human Periodontal Ligament Stem Cells and Have Limited Effects on Cells Differentiation

Gold nanoparticles (AuNPs) had been widely applied in the practice and advancement of chemistry, biology, and medicine due to facility of synthesis and versatility in surface functionalization. Recent studies had shown that AuNPs can be applied to cells, affecting cellular physiological processes such as proliferation and differentiation. In this study, four diameters of AuNPs (20, 40, 60, and 80 nm) were cocultured with human periodontal ligament cells (hPDLCs) at six different concentrations. The optimal size and concentration of AuNPs were selected to treat human periodontal ligament stem cells (hPDLSCs) to evaluate proliferation. Moreover, the influence of AuNPs on multiple differentiation capacity of hPDLSCs was clarified. The results revealed that AuNPs (60 nm, 56 μM) can effectively promote the proliferation of hPDLCs/hPDLSCsin vitro, slightly enhance osteoblastic differentiation, and have no effect on adipogenic differentiation. In addition, the expression of COL-1, Runx2, BSP, and OCN was upregulated in the presence of AuNPs (60 nm, 56 μM). These results indicated that AuNPs (60 nm, 56 μM) can effectively promote the proliferation of hPDLCs/hPDLSCs and have no significant effect on the differentiation of hPDLSCs. These results provide an insight on the advantage of implementing of AuNPs on hPDLSCs culture and expose the influence of these materials on periodontal tissue engineering.

Cancer stem cells: The potential of carbon ion beam radiation and new radiosensitizers (Review)

Cancer stem cells (CSCs) are a small population of cells in cancer with stem-like properties such as cell proliferation, multiple differentiation and tumor initiation capacities. CSCs are therapy-resistant and cause cancer metastasis and recurrence. One key issue in cancer therapy is how to target and eliminate CSCs, in order to cure cancer completely without relapse and metastasis. To target CSCs, many cell surface markers, DNAs and microRNAs are considered as CSC markers. To date, the majority of the reported markers are not very specific to CSCs and are also present in non-CSCs. However, the combination of several markers is quite valuable for identifying and targeting CSCs, although more specific identification methods are needed. While CSCs are considered as critical therapeutic targets, useful treatment methods remain to be established. Epigenetic gene regulators, microRNAs, are associated with tumor initiation and progression. MicroRNAs have been recently considered as promising therapeutic targets, which can alter the therapeutic resistance of CSCs through epigenetic modification. Moreover, carbon ion beam radiotherapy is a promising treatment for CSCs. Evidence indicates that the carbon ion beam is more effective against CSCs than the conventional X-ray beam. Combination therapies of radiosensitizing microRNAs and carbon ion beam radiotherapy may be a promising cancer strategy. This review focuses on the identification and treatment resistance of CSCs and the potential of microRNAs as new radiosensitizers and carbon ion beam radiotherapy as a promising therapeutic strategy against CSCs.

Differentiation of Glioma Stem Cells and Progenitor Cells into Local Host Cell-Like Cells: A Study Based on Choroidcarcinoma Differentiation of Choroid Plexus of GFP Transgenic Nude Mouse.

The idea of multiple differentiation capacity of glioma stem cells and progentior cells (GSCPs) has been accepted by most of the researchers, but the effect of local environment on the differentiation of GSCPs is unclear. GSCPs SU2 and CM-Dil-stained C6 cells (C6-Dil) were injected into the brain of GFP transgenic nude mice. The xenografts were sectioned. Morphological changes of tumor cells that resided in the choroid plexus, molecular markers expression, and the relationship between the original tumor cells and host cells were studied carefully. The tumorigenicity rate was 40/40 (100%) in all of the inoculated nude mice. Cell morphology and molecular expression of neoplasm settled in the choroid plexus showed that choroidcarcinoma derived from GSCPs was developed. These results showed that GSCPs may have the multiple differentiation capacity, which can be induced by the local environment of host brain as NSCs, and cell fusion may play an important role in the transformation.

Mechanical stress stimulates the osteo/odontoblastic differentiation of human stem cells from apical papilla via erk 1/2 and JNK MAPK pathways.

Background Information. Stem cells from apical papilla (SCAPs) are a potent candidate for the apexogenesis/apexification due to their multiple differentiation capacity. During the orthodontic treatment of developing teeth, SCAPs in vivo are usually subjected to the cyclic stress induced by compression forces. However, it remains unclear whether mechanical stress can affect the proliferation and differentiation of human SCAPs. Results. Human SCAPs were isolated and stimulated by 200 g mechanical stimuli for 30 min and their proliferation and differentiation capacity were evaluated in vitro at different time points. MTT and FCM results demonstrated that cell proliferation was enhanced, while TEM findings showed the morphological and ultrastructural changes in stress-treated SCAPs. ALP activity and mineralization capacity of stress-treated SCAPs were upregulated . In the meantime, higher odontogenic and osteogenic differentiation were found in stress-treated SCAPs by real-time RT-PCR and Western blot, as indicated by the expression of related markers at both mRNA and protein levels. Moreover, the protein expressions of pJNK and pERK MAPK pathways were upregulated. Conclusion. Together, these findings suggest that mechanical stress is an important factor affecting the proliferation and differentiation of SCAPs via the activation of ERK and JNK signaling pathway.

A New Source of Mesenchymal Stem Cells for Articular Cartilage Repair

Background: Bone marrow (BM) has been considered as a major source of mesenchymal stem cells (MSCs), but it has many disadvantages in clinical application. However, MSCs from peripheral blood (PB) could be obtained by a less invasive method and be more beneficial for autologous transplantation than BM MSCs, which makes PB a promising source for articular cartilage repair in clinical use. Purpose: To assess whether MSCs from mobilized PB of New Zealand White rabbits have similar biological characteristics in vitro and chondrogenesis in vivo as BM MSCs. Study Design: Controlled laboratory study. Methods: A combined method of drug administration containing granulocyte colony stimulating factor (G-CSF) plus CXCR4 antagonist AMD3100 was adopted to mobilize the PB stem cells of adult New Zealand White rabbits in vitro. The isolated cells were identified as MSCs by morphological characteristics, surface markers, and differentiation potentials. A comparison between PB MSCs and BM MSCs was made in terms of biological characteristics in vitro and chondrogenesis in vivo. This issue was investigated from the aspects of morphology, immune phenotype, multiple differentiation capacity, expansion potential, antiapoptotic capacity, and ability to repair cartilage defects in vivo of PB MSCs compared with BM MSCs. Results: Peripheral blood MSCs were successfully mobilized by the method of combined drug administration, then isolated, expanded, and identified in vitro. No significant difference was found concerning the morphology, immune phenotype, and antiapoptotic capacity between PB MSCs and BM MSCs. Significantly, MSCs from both sources compounded with decalcified bone matrix showed the same ability to repair cartilage defects in vivo. For multipluripotency, BM MSCs exhibited a more osteogenic potential and higher proliferation capacity than PB MSCs, whereas PB MSCs possessed a stronger adipogenic and chondrogenic differentiation potential than BM MSCs in vitro. Conclusion: Although there are some differences in the proliferation and differentiation aspects between the 2 sources, PB MSCs share certain similar biological characteristics in vitro and chondrogenesis in vivo as BM MSCs. Clinical Relevance: These results suggest that PB MSCs are a new source of seed cells used in articular cartilage repair.

Effective combination of human bone marrow mesenchymal stem cells and minocycline in experimental autoimmune encephalomyelitis mice

IntroductionMultiple sclerosis (MS) is the most common inflammatory demyelinating disorder of the central nervous system (CNS). Minocycline ameliorates the clinical severity of MS and exhibits antiinflammatory, neuroprotective activities, and good tolerance for long-term use, whereas it is toxic to the CNS. Recently, the immunomodulation and neuroprotection capabilities of human bone marrow mesenchymal stem cells (hBM-MSCs) were shown in experimental autoimmune encephalomyelitis (EAE). In this study, we evaluated whether the combination of hBM-MSCs and a low-dose minocycline could produce beneficial effects in EAE mice.MethodsThe sensitivity of hBM-MSCs to minocycline was determined by an established cell-viability assay. Minocycline-treated hBM-MSCs were also characterized with flow cytometry by using MSC surface markers and analyzed for their multiple differentiation capacities. EAE was induced in C57BL/6 mice by using immunization with MOG35-55. Immunopathology assays were used to detect the inflammatory cells, demyelination, and neuroprotection. Interferon gamma (IFN-γ)/tumor necrosis factor alpha (TNF-α) and interleukin-4 (IL-4)/interleukin-10 (IL-10), the hallmark cytokines that direct Th1 and Th2 development, were detected with enzyme-linked immunosorbent assay (ELISA). terminal dUTP nick-end labeling (TUNEL) staining was performed to elucidate the cell apoptosis in the spinal cords of EAE mice.ResultsMinocycline did not affect the viability, surface phenotypes, or differentiation capacity of hBM-MSCs, while minocycline affected the viability of astrocytes at a high dose. In vivo efficacy experiments showed that combined treatment, compared to the use of minocycline or hBM-MSCs alone, resulted in a significant reduction in clinical scores, along with attenuation of inflammation, demyelination, and neurodegeneration. Moreover, the combined treatment with hBM-MSCs and minocycline enhanced the immunomodulatory effects, which suppressed proinflammatory cytokines (IFN-γ, TNF-α) and conversely increased anti-inflammatory cytokines (IL-4, IL-10). In addition, TUNEL staining also demonstrated a significant decrease of the number of apoptotic cells in the combined treatment compared with either treatment alone.ConclusionsThe combination of hBM-MSCs and minocycline provides a novel experimental protocol to enhance the therapeutic effects in MS.

The Comparison of Multilineage Differentiation of Bone Marrow and Adipose-Derived Mesenchymal Stem Cells

To compare the roles of adipose and bone marrow derived mesenchymal stem cells (AMSCs and BMSCs) in multiple differentiation capacity to provide a theoretical basis for stem cell transplantation. We isolated bone marrow and adipose derived mesenchymal stem cells and compared their phenotype, cell doubling time, the secretion of factors, and the ability of multi-differentiation. BMSCs and AMSCs were similar in cell phenotype and the differences existed only between the expression of CD106. The proliferation rate of AMSCs was faster than of BMSCs (doubling time 28h vs. 39h) and the capacity to suppress T cell proliferation and activation was weakened in AMSCs. In addition, both sources of cells were able to differentiate into bone, fat, and cartilage which proved their stem cell properties. Cell origin and abundance were decisive factors in stem cell applications and with the same premise as for AMSCs and BMSCs, adipose tissue is a more promising source of stem cells.

Preliminary Study on Dental Pulp Stem Cell–mediated Pulp Regeneration in Canine Immature Permanent Teeth

IntroductionThe health of human teeth depends on the integrity of the hard tissue and the activity of the pulp and periodontal tissues, which are responsible for nutritional supply. Without the nourishing of the pulp tissue, the possibility of tooth fracture can increase. In immature permanent teeth, root development may be influenced as well. This study explored the potential of using autologous dental pulp stem cells (DPSCs) to achieve pulp regeneration in a canine pulpless model. MethodsThe establishment of the pulpless animal model involved pulp extirpation and root canal preparation of young permanent incisor teeth in beagles. Autologous DPSCs were obtained from extracted first molars and expanded ex vivo to obtain a larger number of cells. The biological characteristics of canine DPSCs (cDPSCs) were analyzed both in vitro and in vivo by using the same method as used in human DPSCs. cDPSCs were transplanted into the pulpless root canal with Gelfoam as the scaffold, and root development was evaluated by radiographic and histologic analyses. ResultscDPSCs with rapid proliferation, multiple differentiation capacity, and development potential were successfully isolated and identified both in vitro and in vivo. After they were transplanted into the pulpless root canal with Gelfoam as the scaffold, DPSCs were capable of generating pulp-like tissues containing blood vessels and dentin-like tissue. Thickening of the root canal wall was also observed. ConclusionsThis study demonstrates the feasibility of using stem cell–mediated tissue engineering to realize pulp regeneration in immature teeth.

The comparition of biological characteristics and multilineage differentiation of bone marrow and adipose derived Mesenchymal stem cells

We compared the two sources of adipose and bone marrow-derived mesenchymal stem cells (BMSCs and AMSCs ) in multiple differentiation capacity and biological characteristics to provide a theoretical basis for stem cells transplantation. We isolated bone marrow- and adipose-derived mesenchymal stem cells and compared their phenotype,cell doubling time, the secretion of factors and their ability of multi-differentiation. We also compared their differences in T lymphocyte activation, proliferation and suppression. BMSCs and AMSCs were similar in cell phenotype and the differences existed only in the expression of CD106. On the proliferation rate, AMSCs were faster than BMSCs (doubling time 28 vs. 39h). In addition, both of these two sources of cells were able to differentiate into bone, fat and cartilage that proved their stem cells properties and the number of stem cell progenitors (CFU-F) from adipose tissue were 10 times larger than those from bone marrow. But AMSCs showed a diminished capacity for suppressing T lymphocyte proliferation and activation compared to BMSCs. Cell origin and abundance were decisive factors in stem cells applications and, in the same volume, with the same premise of AMSCs and BMSCs, adipose tissue is a more promising source of stem cells.

Increased Differentiation Capacity of Bone Marrow-Derived Mesenchymal Stem Cells in Aquaporin-5 Deficiency

Mesenchymal stem cells (MSCs) are adult stem cells with a self-renewal and multipotent capability and express extensively in multitudinous tissues. We found that water channel aquaporin-5 (AQP5) is expressed in bone marrow-derived MSCs (BMMSCs) in the plasma membrane pattern. BMMSCs from AQP5(-/-) mice showed significantly lower plasma membrane water permeability than those from AQP5(+/+) mice. In characterizing the cultured BMMSCs from AQP5(-/-) and AQP5(+/+) mice, we found no obvious differences in morphology and proliferation between the 2 genotypes. However, the multiple differentiation capacity was significantly higher in AQP5(-/-) than AQP5(+/+) BMMSCs as revealed by representative staining by Oil Red O (adipogenesis); Alizarin Red S and alkaline phosphatase (ALP; osteogenesis); and type II collagen and Safranin O (chondrogenesis) after directional induction. Relative mRNA expression levels of 3 lineage differentiation markers, including PPARγ2, C/EBPα, adipsin, collagen 1a, osteopontin, ALP, collagen 11a, collagen 2a, and aggrecan, were significantly higher in AQP5(-/-) -differentiating BMMSCs, supporting an increased differentiation capacity of AQP5(-/-) BMMSCs. Furthermore, a bone-healing process was accelerated in AQP5(-/-) mice in a drill-hole injury model. Mechanistic studies indicated a significantly lower apoptosis rate in AQP5(-/-) than AQP5(+/+) BMMSCs. Apoptosis inhibitor Z-VAD-FMK increased the differentiation capacity to a greater extent in AQP5(+/+) than AQP5(-/-) BMMSCs. We conclude that AQP5-mediated high plasma membrane water permeability enhances the apoptosis rate of differentiating BMMSCs, thus decreasing their differentiation capacity. These data implicate AQP5 as a novel determinant of differentiation of BMMSCs and therefore a new molecular target for regulating differentiation of BMMSCs during tissue repair and regeneration.