Neural Differentiation of Mesenchymal Stem Cells Influences Their Chemotactic Responses to Stromal Cell-Derived Factor-1α

Mesenchymal stem cells (MSCs) are proposed as a promising source for cell-based therapies in neural disease. Although increasing numbers of studies have been devoted to the delineation of factors involved in the migration of MSCs, the relationship between the chemotactic response and the differentiation status of these cells is still unclear. In the present study, we demonstrated that MSCs in varying neural differentiation states display various chemotactic responses to stromal cell-derived factor-1α (SDF-1α). The chemotactic responses of MSCs under different differentiation stages in response to SDF-1α were analyzed by Boyden chamber, and the results showed that cells of undifferentiation, 24-h preinduction, 5-h induction, and 18-h maintenance states displayed a stronger chemotactic response to SDF-1α, while 48-h maintenance did not. Further, we found that the phosphorylation levels of PI3K/Akt, ERK1/2, SAPK/JNK, and p38MAPK are closely related to the differentiation states of MSCs subjected to SDF-1α, and finally, inhibition of SAPK/JNK signaling significantly attenuates SDF-1α-stimulated transfilter migration of MSCs of undifferentiation, 24-h preinduction, 18-h maintenance, and 48-h maintenance, but not MSCs of 5-h induction. Meanwhile, interference with PI3K/Akt, p38MAPK, or ERK1/2 signaling prevents only cells at certain differentiation state from migrating in response to SDF-1α. Collectively, these results demonstrate that MSCs in varying neural differentiation states have different migratory capacities, thereby illuminating optimization of the therapeutic potential of MSCs to be used for neural regeneration after injury.     

Mesenchymal stem cell therapy and acute graft-versus-host disease: a review

Mesenchymal stem cells (MSCs) are being widely studied as potential cell therapy agents due to their immunomodulatory properties, which have been established by in vitro studies and in several clinical trials. Within this context, mesenchymal stem cell therapy appears to hold substantial promise, particularly in the treatment of conditions involving autoimmune and inflammatory components. Nevertheless, many research findings are still contradictory, mostly due to difficulties in characterization of the effects of MSCs in vivo. The purpose of this review is to report the mechanisms underlying mesenchymal stem cell therapy for acute graft-versus-host disease, particularly with respect to immunomodulation, migration, and homing, as well as report clinical applications described in the literature.     

Roles of db-cAMP,IBMX and RA in Aspects of Neural Differentiation of Cord Blood Derived Mesenchymal-Like Stem Cells

Mesenchymal stem cells (MSCs) have multilineage differentiation potential which includes cell lineages of the central nervous system; hence MSCs might be useful in the treatment of neurodegenerative diseases such as Parkinson''s disease. Although mesenchymal stem cells have been shown to differentiate into the neural lineage, there is still little knowledge about the underlying mechanisms of differentiation particularly towards specialized neurons such as dopaminergic neurons. Here, we show that MSCs derived from human umbilical cord blood (MSC^hUCBs) are capable of expressing tyrosine hydroxylase (TH) and Nurr1, markers typically associated with DA neurons. We also found differential phosphorylation of TH isoforms indicating the presence of post-translational mechanisms possibly activating and modifying TH in MSC^hUCB. Furthermore, functional dissection of components in the differentiation medium revealed that dibutyryl-cAMP (db-cAMP), 3-isobutyl-1-methylxanthine (IBMX) and retinoic acid (RA) are involved in the regulation of Nurr1 and Neurofilament-L expression as well as in the differential phosphorylation of TH. We also demonstrate a possible inhibitory role of the protein kinase A signaling pathway in the phosphorylation of specific TH isoforms.     

Mesenchymal stem cells in arthritic diseases

Mesenchymal stem cells (MSCs), the nonhematopoietic progenitor cells found in various adult tissues, are characterized by their ease of isolation and their rapid growth in vitro while maintaining their differentiation potential, allowing for extensive culture expansion to obtain large quantities suitable for therapeutic use. These properties make MSCs an ideal candidate cell type as building blocks for tissue engineering efforts to regenerate replacement tissues and repair damaged structures as encountered in various arthritic conditions. Osteoarthritis (OA) is the most common arthritic condition and, like rheumatoid arthritis (RA), presents an inflammatory environment with immunological involvement and this has been an enduring obstacle that can potentially limit the use of cartilage tissue engineering. Recent advances in our understanding of the functions of MSCs have shown that MSCs also possess potent immunosuppression and anti-inflammation effects. In addition, through secretion of various soluble factors, MSCs can influence the local tissue environment and exert protective effects with an end result of effectively stimulating regeneration in situ. This function of MSCs can be exploited for their therapeutic application in degenerative joint diseases such as RA and OA. This review surveys the advances made in the past decade which have led to our current understanding of stem cell biology as relevant to diseases of the joint. The potential involvement of MSCs in the pathophysiology of degenerative joint diseases will also be discussed. Specifically, we will explore the potential of MSC-based cell therapy of OA and RA by means of functional replacement of damaged cartilage via tissue engineering as well as their anti-inflammatory and immunosuppressive activities.     

Stimulating factors for regulation of osteogenic and chondrogenic differentiation of mesenchymal stem cells

Mesenchymal stem cells (MSCs), distributed in many tissues in the human body, are multipotent cells capable of differentiating in specific directions. It is usually considered that the differentiation process of MSCs depends on specialized external stimulating factors, including cell signaling pathways, cytokines, and other physical stimuli. Recent findings have revealed other underrated roles in the differentiation process of MSCs, such as material morphology and exosomes. Although relevant achievements have substantially advanced the applicability of MSCs, some of these regulatory mechanisms still need to be better understood. Moreover, limitations such as long-term survival in vivo hinder the clinical application of MSCs therapy. This review article summarizes current knowledge regarding the differentiation patterns of MSCs under specific stimulating factors.     

低氧对 MSCs 增殖和分化的影响

摘要： 间充质干细胞(mesenchymal stem cells, MSCs)是具有自我更新、 多向分化和强可塑性的细胞, 具有分化为血液、 骨、 软骨、 脂 肪、 肌肉、 表皮、 上皮、 神经等组织的潜能, 受到再生医学研究的关注。目前已有研究表明将 MSCs 移植到多种损伤组织中都能改 善损伤组织的功能。文章在简要回顾了低氧环境对 MSCs 增殖和分化的研究内容和有关理论争论基础上重点介绍了缺氧诱导因 子 （ HIF ）通路对 MSCs 增殖和分化的影响。文章阐述了低氧环境对 MSCs 向成骨,成软骨,成脂及成神经元方向分化的影响。 由于 人体组织内生理条件下的氧张力远远小于大气中的氧张力 （21% ）, 采用低氧培养 MSCs 的研究方法得出的结论将更加贴近实际 MSCs 在人体内的增殖、分化情况。因此研究 MSCs 在低氧张力环境中增殖、分化的能力将为 MSCs 能成功移植到体内并发挥作 用提供保障。     

低氧对MSCs增殖和分化的影响

间充质干细胞（mesenchymal stemcells, MSCs）是具有自我更新、多向分化和强可塑性的细胞,具有分化为血液、骨、软骨、脂肪、肌肉、表皮、上皮、神经等组织的潜能,受到再生医学研究的关注。目前已有研究表明将MSCs 移植到多种损伤组织中都能改善损伤组织的功能。文章在简要回顾了低氧环境对MSCs增殖和分化的研究内容和有关理论争论基础上重点介绍了缺氧诱导因子（HIF）通路对MSCs 增殖和分化的影响。文章阐述了低氧环境对MSCs向成骨,成软骨,成脂及成神经元方向分化的影响。由于人体组织内生理条件下的氧张力远远小于大气中的氧张力（21%）,采用低氧培养MSCs 的研究方法得出的结论将更加贴近实际MSCs在人体内的增殖、分化情况。因此研究MSCs 在低氧张力环境中增殖、分化的能力将为MSCs 能成功移植到体内并发挥作用提供保障。     

Differentiation Potential of Mesenchymal Stem Cells and Stimulation of Nerve Regeneration

Mesenchymal stem cells (MSCs) are widely used in experimental research on cell therapy intended for the stimulation of repair processes in damaged tissues and organs. The present review summarizes the results of studies devoted to the possible directions of MSC differentiation after the transplantation of these cells into damaged nerves or special engineered structures of biological and artificial biodegradable materials that join the ends of a damaged nerve (nerve conduits). Data on exogenous MSC differentiation into Schwann cells, pericytes, smooth muscle cells, endotheliocytes, and other cell types are presented. Methods for preliminary MSC differentiation in vitro and examples of beneficial effects of these cells transplanted into damaged conductive nerves on nerve regeneration are given. The fate of exogenous MSCs placed into an unnatural biological niche remains poorly characterized and requires further studies, as emphasized in the review.     

Monitoring the biology stability of human umbilical cord-derived mesenchymal stem cells during long-term culture in serum-free medium

Mesenchymal stem cells (MSCs) are multipotent adult stem cells that have an immunosuppressive effect. The biological stability of MSCs in serum-free medium during long-term culture in vitro has not been elucidated clearly. The morphology, immunophenotype and multi-lineage potential were analyzed at passages 3, 5, 10, 15, 20, and 25 (P₃, P₅, P₁₀, P₁₅, P₂₀, and P₂₅, respectively). The cell cycle distribution, apoptosis, and karyotype of human umbilical cord-derived (hUC)-MSCs were analyzed at P₃, P₅, P₁₀, P₁₅, P₂₀, and P₂₅. From P₃ to P₂₅, the three defining biological properties of hUC-MSCs [adherence to plastic, specific surface antigen expression, multipotent differentiation potential] met the standards proposed by the International Society for Cellular Therapy for definition of MSCs. The cell cycle distribution analysis at the P₂₅ showed that the percentage of cells at G0/G1 was increased, compared with the cells at P₃ (P < 0.05). Cells at P₂₅ displayed an increase in the apoptosis rate (to 183 %), compared to those at P₃ (P < 0.01). Within subculture generations 3–20 (P₃–P₂₀), the differences between the cell apoptotic rates were not statistically significant (P > 0.05). There were no detectable chromosome eliminations, displacements, or chromosomal imbalances, as assessed by the karyotyping guidelines of the International System for Human Cytogenetic Nomenclature (ISCN, 2009). Long-term culture affects the biological stability of MSCs in serum-free MesenCult-XF medium. MSCs can be expanded up to the 25th passage without chromosomal changes by G-band. The best biological activity period and stability appeared between the third to 20th generations.     

Migration properties of adipose-tissue-derived mesenchymal stromal cells cocultured with activated monocytes in vitro

Mesenchymal stromal cells (MSCs) are a promising tool in regenerative medicine. MSC migration to damaged inflammatory sites (homing) is essential for tissue repair. We have studied the migration properties of adipose-tissue-derived MSCs (AT-MSC) after their cocultivation with activated monocytes from the THP-1 cell line. We observed the increased migration rate of AT-MSC in vitro with the lack of chemoattractant gradient and to the platelet-derived growth factor (PDGF BB), which is a well-known chemoattractant for cells of mesenchymal origin. Moreover, the rate of directional AT-MSC migration through fibronectin was also increased. We demonstrated that signaling via PDGFR-β activated through the binding of integrin receptors with an extracellular matrix is a possible mechanism for stimulation of cellular migration under simulated inflammatory conditions.     

Efficient Generation of Neural Stem Cell-Like Cells from Rat Adipose Derived Stem Cells After Lentiviral Transduction with Green Fluorescent Protein

Neural stem cells (NSCs) can be isolated from nervous tissues or derived from embryonic stem cells. However, their procurement for clinical applications is limited, and there is a need for alternative types of cell that have NSCs properties. In the present study, the differentiation potential of rat adipose-derived stem cells (ADSCs) was evaluated by infecting these cells with a lentiviral vector-encoding green fluorescent protein (GFP). ADSCs transduced with lentivirus were able to generate NSC-like cells, without any effects on their growth, phenotype, and normal differentiation potential. NSC-like cells derived from ADSCs formed neurospheres and expressed high levels of the neural progenitor marker nestin. In the absence of selected growth factors, these neurospheres differentiated into neurons expressing NeuN and MAP2 and GFAP-expressing glia, as determined by immunocytochemistry, Western blotting, and quantitative real-time polymerase chain reaction. These results demonstrate that ADSCs can be induced to generate neurospheres that have NSC-like properties and may thus constitute a potential source of cells in stem cell therapy for neurological disorders.     

Human mesenchymal stem cells - current trends and future prospective

Stem cells are cells specialized cell, capable of renewing themselves through cell division and can differentiate into multi-lineage cells. These cells are categorized as embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs) and adult stem cells. Mesenchymal stem cells (MSCs) are adult stem cells which can be isolated from human and animal sources. Human MSCs (hMSCs) are the non-haematopoietic, multipotent stem cells with the capacity to differentiate into mesodermal lineage such as osteocytes, adipocytes and chondrocytes as well ectodermal (neurocytes) and endodermal lineages (hepatocytes). MSCs express cell surface markers like cluster of differentiation (CD)29, CD44, CD73, CD90, CD105 and lack the expression of CD14, CD34, CD45 and HLA (human leucocyte antigen)-DR. hMSCs for the first time were reported in the bone marrow and till now they have been isolated from various tissues, including adipose tissue, amniotic fluid, endometrium, dental tissues, umbilical cord and Wharton''s jelly which harbours potential MSCs. hMSCs have been cultured long-term in specific media without any severe abnormalities. Furthermore, MSCs have immunomodulatory features, secrete cytokines and immune-receptors which regulate the microenvironment in the host tissue. Multilineage potential, immunomodulation and secretion of anti-inflammatory molecules makes MSCs an effective tool in the treatment of chronic diseases. In the present review, we have highlighted recent research findings in the area of hMSCs sources, expression of cell surface markers, long-term in vitro culturing, in vitro differentiation potential, immunomodulatory features, its homing capacity, banking and cryopreservation, its application in the treatment of chronic diseases and its use in clinical trials.     

Q&amp;A: Mesenchymal stem cells — where do they come from and is it important?

Mesenchymal stem — or stromal — cells (MSCs) have been administered in hundreds of clinical trials for multiple indications, making them some of the most commonly used selected regenerative cells. Paradoxically, MSCs have also long remained the least characterized stem cells regarding native identity and natural function, being isolated retrospectively in long-term culture. Recent years have seen progress in our understanding of the natural history of these cells, and candidate native MSCs have been identified within fetal and adult organs. Beyond basic knowledge, deciphering the biology of innate MSCs may have important positive consequences for the therapeutic use of these cells.     

Deferoxamine promotes mesenchymal stem cell homing in noise‐induced injured cochlea through PI3K/AKT pathway

Objective

Over 5% of the world's population suffers from disabling hearing loss. Stem cell homing in target tissue is an important aspect of cell‐based therapy, which its augmentation increases cell therapy efficiency. Deferoxamine (DFO) can induce the Akt activation, and phosphorylation status of AKT (p‐AKT) upregulates CXC chemokine receptor‐4 (CXCR4) expression. We examined whether DFO can enhance mesenchymal stem cells (MSCs) homing in noise‐induced damaged cochlea by PI3K/AKT dependent mechanism.

Materials and Methods

Mesenchymal stem cells were treated with DFO. AKT, p‐AKT protein and hypoxia inducible factor 1‐ α (HIF‐1α) and CXCR4 gene and protein expression was evaluated by RT‐ PCR and Western blot analysis. For in vivo assay, rats were assigned to control, sham, noise exposure groups without any treatment or receiving normal, DFO‐treated and DFO +LY294002 (The PI3K inhibitor)‐treated MSCs. Following chronic exposure to 115 dB white noise, MSCs were injected into the rat cochlea through the round window. Number of Hoechst‐ labelled cells was determined in the endolymph after 24 hours.

Results

Deferoxamine increased P‐AKT, HIF‐1α and CXCR4 expression in MSCs compared to non‐treated cells. DFO pre‐conditioning significantly increased the homing ability of MSCs into injured ear compared to normal MSCs. These effects of DFO were blocked by LY294002.

Conclusions

Pre‐conditioning of MSCs by DFO before transplantation can improve stem cell homing in the damaged cochlea through PI3K/AKT pathway activation.

     

Mesenchymal stem cells neuronal differentiation ability: a real perspective for nervous system repair?

Mesenchymal Stem Cells (MSCs) are a bone marrow-derived population present in adult tissues that possess the important property of dividing when called upon and of differentiating into specialized cells. The evidence that MSCs were able to transdifferentiate into specialized cells of tissues different from bone marrow, in particular into nervous cells, opened up the possibility of using MSCs to substitute damaged neurons, that are normally not replaced but lost, in order to repair the Nervous System. The first neuronal differentiation protocols were based on the use of a mixture of toxic drugs which induced MSCs to rapidly acquire a neuronal-like morphology with the expression of specific neuronal markers. However, many subsequent studies demonstrated that the morphological and molecular modifications of MSCs were probably due to a stress response, rather than to a real differentiation into neuronal cells, thus throwing into question the possible use of MSCs to repair the nervous system. Currently, some papers are suggesting again that it may be possible to induce neuronal differentiation of MSCs by using several differentiation protocols, and by accompanying the morphological evidence of differentiation with functional evidence, thus demonstrating that MSC-derived cells not only seem to be neurons, but that they also function like neurons. In this review, we have attempted to shed light on the capacity of MSCs to genuinely differentiate into nervous cells, and to identify the most reliable protocols for obtaining neurons from MSCs for nervous system repair.     

Pharmaceutical induction of ApoE secretion by multipotent mesenchymal stromal cells (MSCs)

Background

Apolipoprotein E (ApoE) is a molecular scavenger in the blood and brain. Aberrant function of the molecule causes formation of protein and lipid deposits or "plaques" that characterize Alzheimer's disease (AD) and atherosclerosis. There are three human isoforms of ApoE designated ε2, ε3, and ε4. Each isoform differentially affects the structure and function of the protein and thus the development of disease. Homozygosity for ApoE ε4 is associated with atherosclerosis and Alzheimer's disease whereas ApoE ε2 and ε3 tend to be protective. Furthermore, the ε2 form may cause forms of hyperlipoproteinemia. Therefore, introduction of ApoE ε3 may be beneficial to patients that are susceptible to or suffering from these diseases. Mesenchymal stem cells or multipotent mesenchymal stromal cells (MSCs) are adult progenitor cells found in numerous tissues. They are easily expanded in culture and engraft into host tissues when administered appropriately. Furthermore, MSCs are immunosuppressive and have been reported to engraft as allogeneic transplants. In our previous study, mouse MSCs (mMSCs) were implanted into the brains of ApoE null mice, resulting in production of small amounts of ApoE in the brain and attenuation of cognitive deficits. Therefore human MSCs (hMSCs) are a promising vector for the administration of ApoE ε3 in humans.

Results

Unlike mMSCs, hMSCs were found not to express ApoE in culture; therefore a molecular screen was performed for compounds that induce expression. PPARγ agonists, neural stem cell conditioned medium, osteo-inductive media, dexamethasone, and adipo-inductive media (AIM) were tested. Of the conditions tested, only AIM or dexamethasone induced sustained secretion of ApoE in MSCs and the duration of secretion was only limited by the length of time MSCs could be sustained in culture. Upon withdrawal of the inductive stimuli, the ApoE secretion persisted for a further 14 days.

Conclusion

The data demonstrated that pre-treatment and perhaps co-administration of MSCs homozygous for ApoE ε3 and dexamethasone may represent a novel therapy for severe instances of AD, atherosclerosis and other ApoE-related diseases.     

Detection of cytokines in supernatant from hematopoietic stem/progenitor cells co-cultured with mesenchymal stem cells and endothelial progenitor cells

This study aimed to investigate the significance of cytokine expression in supernatant from hematopoietic stem/progenitor cells (HSCs/HPCs) co-cultured with mesenchymal stem cells (MSCs) or endothelial progenitor cells (EPCs). Mononuclear cells (MNCs) were isolated from normal human umbilical cord blood and then cultured solely or co-cultured with MSCs or EPCs. Changes in the number of MNCs and HSCs/HPCs were observed, and MNC proliferation was tested by carboxyfluorescein diacetate succinimidyl ester. The cultured supernatants of the treated MSCs and EPCs were collected at 24 h after co-culture and used to determine the concentrations of IL-3, IL-6, stem cell factor (SCF), TPO, Flt3l, and VEGF. The total number and proliferation of MNCs increased significantly when co-cultured with MSCs or EPCs than when cultured alone, particularly when MNCs were co-cultured with EPCs. The differences in IL-3 and Flt3l concentrations between groups were not significant. However, IL-6 in the MSC group was significantly higher than that in the two other groups. The SCF and TPO concentrations were highly expressed in the EPC group. The VEGF concentrations in the MSC group and the EPC group were higher than those in the control group. These results indicated that MSCs and EPCs possibly favor the proliferation of MNCs and HSCs/HPCs. IL-6 and VEGF may be related to hematopoietic reconstitution and homing ability of HSCs/HPCs. TPO may have a specific relationship with the promotion of HSCs/HPCs differentiation.     

细胞外基质硬度调控间充质干细胞分化

新近研究表叽细胞外基质（extracellularmatrix,ECM）的物理性质,特别是硬度或弹性,能对细胞的黏附、铺展、迁移、增殖、分化和凋亡等多种功能和行为产生重要影响。间充质干细胞（mesenchymalstemcells,MSCs）是组织工程和细胞治疗的理想种子细胞。ECM硬度可诱导MSCs向脂肪、软骨、神经、肌肉和骨等方向分化。该文综合论述了ECM硬度对干细胞分化的影响,涵盖了构建ECM硬度的测量、调控与表征等,不同培养条件下干细胞对硬度的响应和分化以及硬度和其他因素的联合作用;在此基础上,进一步论述了干细胞分化过程中细胞感应ECM硬度并转化为生物学信号的机制和信号通路。该文还总结了在ECM硬度调控干细胞分化行为领域最新的研究进展情况,较为系统地分析了材料学、细胞生物学、分子生物学水平的主要影响因素,并对本领域未来需要重点研究的问题进行了展望。