Mesenchymal stem cells and skin wound repair and regeneration: possibilities and questions

Adult mesenchymal stem cells (MSCs) have the capacity for self-renewal and for differentiating into a variety of cells and tissues. They may leave their niche to migrate to remote tissues and play a critical role in wound repair and tissue regeneration. Because of their multipotency, easy isolation and culture, highly expansive potential, and immunosuppression properties, these cells may be an attractive therapeutic tool for regenerative medicine and tissue engineering. Several studies have indicated a contribution of MSCs to reconstituting skin in cutaneous wounds, but problems still need resolution before MSCs can be widely used clinically. This review focuses mainly on the benefits of MSCs in skin wound healing and tissue regeneration and on the questions that remain to be answered before MSCs can be used in clinical practice. This study was supported in part by the National Natural Science Foundation of China (30730090, 30672176, 30500194) and by State Key Development Program of Basic Research of China (973 Program, 2005CB522603).     

Mesenchymal stem cells-derived exosomal microRNAs contribute to wound inflammation

Clinical and experimental studies have highlighted the significance of inflammation in coordinating wound repair and regeneration.However,it remains challenging to control the inflammatory response and tolerance at systemic levels without causing toxicity to injured tissues.Mesenchymal stem cells(MSCs) possess potent immunomodulatory properties and facilitate tissue repair by releasing exosomes,which generate a suitable microenvironment for inflammatory resolution.Exosomes contain several effective bioactive molecules and act as a cell-cell communication vehicle to influence cellular activities in recipient cells.During this process,the horizontal transfer of exosomal microRNAs(miRNAs) to acceptor cells,where they regulate target gene expression,is of particular interest for understanding the basic biology of inflammation ablation,tissue homeostasis,and development of therapeutic approaches.In this review,we describe a signature of three specific miRNAs(miR-21,miR-146 a,and miR-181) present in human umbilical cord MSC-derived exosomes(MSC-EXO) identified microarray chip analysis and focus on the inflammatory regulatory functions of these immune-related miRNAs.We also discuss the potential mechanisms contributing to the resolution of wound inflammation and tissue healing.     

Adult bone-marrow-derived mesenchymal stem cells contribute to wound healing of skin appendages

Adult bone-marrow-derived mesenchymal stem cells (MSCs) are well-established as having the capacity to differentiate into cells with mesodermal, ectodermal, and endodermal characteristics and can leave their niche to home toward and engraft within foreign tissues. To investigate whether adult MSCs contribute to the repair of skin appendages after injury, BrdU-labeled MSCs were co-cultured with heat-shocked confluent sweat gland cells (SGCs) in vitro and later intravenously injected into full-thickness skin wounds in rats. When adult MSCs were co-cultured with heat-shocked SGCs, a subset of adult MSCs differentiated into SGCs, the percentage of differentiation being enhanced by epidermal growth factor and the injured microenviroment, but weakened by PD98059. The ERK (extracellular signal-regulated kinase) pathway, especially pERK, was involved in the phenotype conversion of human MSCs into human SGC. Labeled MSCs were noted in hair follicles, sebaceous glands, blood vessels, and dermis in full-thickness wounds, and the incorporated cells in hair follicles and sebaceous glands were also positive for pan-cytokeratin. After wound healing, some labeled MSCs returned to the bone marrow, whereas other were retained in the dermis. We conclude that adult MSCs have the capacity to dock at specific sites, to contribute to wound healing of skin appendages, and to home toward marrow, and that engraftment of bone-marrow-derived cells is a functional event.This work was supported in part by the National Basic Science and Development Program (973 Program and 2005CB522603) and the National Natural Science Foundation of China (30230370 and 30500194).     

Mesenchymal stem cells: Paracrine signaling and differentiation during cutaneous wound repair

Cutaneous wounds persist as a health care crisis in spite of increased understanding of the cellular and molecular responses to injury. Contributing significantly to this crisis is the lack of reliable therapies for treatment of wounds that are slow to heal including chronic wounds and deep dermal wounds that develop hypertrophic scars. This article will review the growing evidence demonstrating the promise of multipotent mesenchymal stem/stromal (MSCs) for the treatment of impaired wound healing. MSCs are often referred to as mesenchymal stem cells despite concerns that these cells are not truly stem cells given the lack of evidence demonstrating self-renewal in vivo. Regardless, abundant evidence demonstrates the therapeutic potential of MSCs for repair and regeneration of damaged tissue due to injury or disease. To date, MSC treatment of acute and chronic wounds results in accelerated wound closure with increased epithelialization, granulation tissue formation and angiogenesis. Although there is evidence for MSC differentiation in the wound, most of the therapeutic effects are likely due to MSCs releasing soluble factors that regulate local cellular responses to cutaneous injury. Important challenges need to be overcome before MSCs can be used effectively to treat wounds that are slow to heal.     

Mesenchymal stem cells: Potential role in corneal wound repair and transplantation

Corneal diseases are a major cause of blindness in the world. Although great progress has been achieved in the treatment of corneal diseases, wound healing after severe corneal damage and immunosuppressive therapy after corneal transplantation remain prob-lematic. Mesenchymal stem cells(MSCs) derived from bone marrow or other adult tissues can differentiate into various types of mesenchymal lineages, such as osteocytes, adipocytes, and chondrocytes, both in vivo and in vitro. These cells can further differentiate into specific cell types under specific conditions. MSCs migrate to injury sites and promote wound healing by secreting anti-inflammatory and growth factors. In ad-dition, MSCs interact with innate and acquired immune cells and modulate the immune response through their powerful paracrine function. Over the last decade, MSCs have drawn considerable attention because of their beneficial properties and promising therapeutic prospective. Furthermore, MSCs have been applied to various studies related to wound healing, autoim-mune diseases, and organ transplantation. This review discusses the potential functions of MSCs in protecting corneal tissue and their possible mechanisms in corneal wound healing and corneal transplantation.     

Mesenchymal stem cells promote alveolar epithelial cell wound repair in vitro through distinct migratory and paracrine mechanisms

Background

Mesenchymal stem cells (MSC) are in clinical trials for widespread indications including musculoskeletal, neurological, cardiac and haematological disorders. Furthermore, MSC can ameliorate pulmonary fibrosis in animal models although mechanisms of action remain unclear. One emerging concept is that MSCs may have paracrine, rather than a functional, roles in lung injury repair and regeneration.

Methods

To investigate the paracrine role of human MSC (hMSC) on pulmonary epithelial repair, hMSC-conditioned media (CM) and a selected cohort of hMSC-secretory proteins (identified by LC-MS/MS mass spectrometry) were tested on human type II alveolar epithelial cell line A549 cells (AEC) and primary human small airway epithelial cells (SAEC) using an in vitro scratch wound repair model. A 3D direct-contact wound repair model was further developed to assess the migratory properties of hMSC.

Results

We demonstrate that MSC-CM facilitates AEC and SAEC wound repair in serum-dependent and –independent manners respectively via stimulation of cell migration. We also show that the hMSC secretome contains an array of proteins including Fibronectin, Lumican, Periostin, and IGFBP-7; each capable of influencing AEC and SAEC migration and wound repair stimulation. In addition, hMSC also show a strong migratory response to AEC injury as, supported by the observation of rapid and effective AEC wound gap closure by hMSC in the 3D model.

Conclusion

These findings support the notion for clinical application of hMSCs and/or their secretory factors as a pharmacoregenerative modality for the treatment of idiopathic pulmonary fibrosis (IPF) and other fibrotic lung disorders.     

Mesenchymal stem cells contribute to endogenous FVIII:c production

Besides the liver, it has been difficult to identify which organ(s) and/or cellular component(s) contribute significantly to the production of human FVIII:c (FVIII). Thus far, only endothelial cells have been shown to constitute a robust extrahepatic source of FVIII, possibly explaining both the diverse presence of FVIII mRNA in the body, and the observed increase in FVIII levels during liver failure. Here, we investigate whether human mesenchymal stem cells (MSC), ubiquitously present in different organs, could also contribute to FVIII production. MSC isolated from human lung, liver, brain, and bone marrow expressed FVIII message as determined by quantitative‐RT‐PCR. Using an antibody specific for FVIII, confocal microscopy, and umbilical cord‐derived endothelial cells (HUVEC) as a negative control, we demonstrated that, in MSC, FVIII protein was not stored in granules; rather, it localized to the perinuclear region. Furthermore, functional FVIII was detected in MSC supernatants and cell lysates by aPTT and chromogenic assays. These results demonstrate that MSC can contribute at low levels to the functional FVIII pool, and advance the understanding of the physiology of FVIII production and secretion. J. Cell. Physiol. © 2012 Wiley Periodicals, Inc.     

Mesenchymal stem cells stimulate intestinal stem cells to repair radiation-induced intestinal injury

The loss of stem cells residing in the base of the intestinal crypt has a key role in radiation-induced intestinal injury. In particular, Lgr5⁺ intestinal stem cells (ISCs) are indispensable for intestinal regeneration following exposure to radiation. Mesenchymal stem cells (MSCs) have previously been shown to improve intestinal epithelial repair in a mouse model of radiation injury, and, therefore, it was hypothesized that this protective effect is related to Lgr5⁺ ISCs. In this study, it was found that, following exposure to radiation, transplantation of MSCs improved the survival of the mice, ameliorated intestinal injury and increased the number of regenerating crypts. Furthermore, there was a significant increase in Lgr5⁺ ISCs and their daughter cells, including Ki67⁺ transient amplifying cells, Vil1⁺ enterocytes and lysozyme⁺ Paneth cells, in response to treatment with MSCs. Crypts isolated from mice treated with MSCs formed a higher number of and larger enteroids than those from the PBS group. MSC transplantation also reduced the number of apoptotic cells within the small intestine at 6 h post-radiation. Interestingly, Wnt3a and active β-catenin protein levels were increased in the small intestines of MSC-treated mice. In addition, intravenous delivery of recombinant mouse Wnt3a after radiation reduced damage in the small intestine and was radioprotective, although not to the same degree as MSC treatment. Our results show that MSCs support the growth of endogenous Lgr5⁺ ISCs, thus promoting repair of the small intestine following exposure to radiation. The molecular mechanism of action mediating this was found to be related to increased activation of the Wnt/β-catenin signaling pathway.The epithelium of the small intestine contains crypts and villi. Intestinal stem cells (ISCs) reside in the base of the crypts and are responsible for maintaining intestinal epithelial homeostasis and regeneration following injury.^{1, 2} Recent studies have identified two populations of stem cells in the small intestine of mice called Lgr5⁺ and Bmi1⁺ ISCs.^{3, 4, 5, 6, 7, 8, 9, 10, 11} Lgr5⁺ ISCs, also known as crypt base columnar cells (CBCs), are interspersed among the Paneth cells and are active rapidly cycling stem cells.¹² A single Lgr5⁺ ISC can grow to form ‘enteroids'' in vitro that develop into all the differentiated cell types found in the intestinal crypt.¹³ Conversely, Bmi1⁺ cells are a population of ISCs located at position +4 relative to the base of the crypt, and are quiescent, slowly cycling stem cells.¹⁴ The loss of ISCs has a critical role in radiation-induced intestinal injury (RIII).^{15, 16, 17, 18} Apoptosis of stem cells because of exposure to radiation prevents normal re-epithelialization of the intestines. Therefore, enhancing the survival of ISCs following radiation is a potential effective treatment for RIII.Mesenchymal stem cells (MSCs) possess significant potential as a therapeutic for tissue damage because of their ability to regulate inflammation, inhibit apoptosis, promote angiogenesis, and support the growth and differentiation of local stem and progenitor cells.^{19, 20} However, the mechanisms by which MSCs mediate these beneficial effects remain unclear, although it has been suggested that MSCs may actively secrete a broad range of bioactive molecules with immunomodulatory (PGE2, IDO, NO, HLA-G5, TSG-6, IL-6, IL-10 and IL-1RA), mitogenic (TGFα/β, HGF, IGF-1, bFGF and EGF), angiogenic (VEGF and TGF-β1) and/or anti-apoptotic (STC-1 and SFRP2) properties that function to modulate the regenerative environment at the site of injury.²¹ Upon re-establishment of the microenvironment following damage, the surviving endogenous stem and progenitor cells can then regenerate the injured tissue completely.Our previous study, as well as other published studies, has found that systemic administration of MSCs improves intestinal epithelial repair in an animal model of radiation injury.^{22, 23, 24, 25} Following MSC treatment, radiation-induced lesions in mice were significantly smaller than those in the control group. However, the mechanism behind this protective effect is not fully understood. Lgr5⁺ ISCs have been previously shown to be indispensable for radiation-induced intestinal regeneration.²⁶ Therefore, in this study, we tested whether the therapeutic effects of MSCs in response to RIII are related to the Lgr5⁺ population of resident ISCs.     

Defects in skin gamma delta T cell function contribute to delayed wound repair in rapamycin-treated mice

Disruptions in the normal program of tissue repair can result in poor wound healing, which perturbs the integrity of barrier tissues such as the skin. Such defects in wound repair occur in transplant recipients treated with the immunosuppressant drug rapamycin (sirolimus). Intraepithelial lymphocytes, such as gammadelta T cells in the skin, mediate tissue repair through the production of cytokines and growth factors. The capacity of skin-resident T cells to function during rapamycin treatment was analyzed in a mouse model of wound repair. Rapamycin treatment renders skin gammadelta T cells unable to proliferate, migrate, and produce normal levels of growth factors. The observed impairment of skin gammadelta T cell function is directly related to the inhibitory action of rapamycin on mammalian target of rapamycin. Skin gammadelta T cells treated with rapamycin are refractory to IL-2 stimulation and attempt to survive in the absence of cytokine and growth factor signaling by undergoing autophagy. Normal wound closure can be restored in rapamycin-treated mice by addition of the skin gammadelta T cell-produced factor, insulin-like growth factor-1. These studies not only reveal that mammalian target of rapamycin is a master regulator of gammadelta T cell function but also provide a novel mechanism for the increased susceptibility to nonhealing wounds that occurs during rapamycin administration.     

Adipocyte lineage cells contribute to the skin stem cell niche to drive hair cycling

In mammalian skin, multiple types of resident cells are required to create a functional tissue and support tissue homeostasis and regeneration. The cells that compose the epithelial stem cell niche for skin homeostasis and regeneration are not well defined. Here, we identify adipose precursor cells within the skin and demonstrate that their dynamic regeneration parallels the activation of skin stem cells. Functional analysis of adipocyte lineage cells in mice with defects in adipogenesis and in transplantation experiments revealed that intradermal adipocyte lineage cells are necessary and sufficient to drive follicular stem cell activation. Furthermore, we implicate PDGF expression by immature adipocyte cells in the regulation of follicular stem cell activity. These data highlight adipogenic cells as skin niche cells that positively regulate skin stem cell activity, and suggest that adipocyte lineage cells may alter epithelial stem cell function clinically.     

Mesenchymal stem cells to treat type 1 diabetes

What is clear is we are in the era of the stem cell and its potential in ameliorating human disease. Our perspective is generated from an in vivo model in a large animal that offers significant advantages (complete transplantation tolerance, large size and long life span). This review is an effort to meld our preclinical observations with others for the reader and to outline potential avenues to improve the present outlook for patients with diabetes. This effort exams the history or background of stem cell research in the laboratory and the clinic, types of stem cells, pluripotency or lack thereof based on a variety of pre-clinical investigations attempting endocrine pancreas recovery using stem cell transplantation. The focus is on the use of hematopoietic and mesenchymal stem cells. This review will also examine recent clinical experience following stem cell transplantation in patients with type 1 diabetes.     

Mesenchymal stem cells are highly resistant to sulfur mustard

The effect of sulfur mustard (SM) to the direct injured tissues of the skin, eyes and airways is well investigated. Little is known about the effect of SM to mesenchymal stem cells (MSC). However, this is an interesting aspect. Comparing the clinical picture of SM it is known today that MSC play an important role e.g. in chronic impaired wound healing. Therefore we wanted to get an understanding about how SM affects MSC and if these findings might become useful to get a better understanding of the effect of sulfur mustard gas with respect to skin wounds.     

脐血干细胞与创面修复

脐血干细胞是一类具有多向分化潜能的原始祖细胞,具备自我更新和增殖的能力,在特定条件诱导下可以分化为不同细胞,逐渐作为临床组织工程的来源细胞。近年来随着对脐血干细胞的不断研究,发现其在创面修复中具有明显的优势,成为创面临床治疗的一条新途径。本文从脐血干细胞的生物学特性、采集与冻存、体外扩增等方面对创面修复的研究进行综述。     

脐血干细胞与创面修复

脐血干细胞是一类具有多向分化潜能的原始祖细胞,具备自我更新和增殖的能力,在特定条件诱导下可以分化为不同细胞,逐渐作为临床组织工程的来源细胞。近年来随着对脐血干细胞的不断研究,发现其在创面修复中具有明显的优势,成为创面临床治疗的一条新途径。本文从脐血干细胞的生物学特性、采集与冻存、体外扩增等方面对创面修复的研究进行综述。     

Autoreactive T cells induce in vitro BM mesenchymal stem cell transdifferentiation to neural stem cells

BACKGROUND: The degree of post-injury inflammation of the damaged area of a spinal cord is the main difference between the natural successful repair in inferior vertebrates and failure in superior vertebrates. The treatment of rats with anti-myelin lymphocytes after experimental spinal cord injury induces their functional recovery. On the other hand, mesenchymal stem cells (MSC) from adult BM implanted in injured areas recover the morphology and function of spinal cord in mammals. The purpose of this study was to determine whether there is a direct relationship between anti-nervous tissue T cells and MSC reparatory properties. METHODS: Circulating autoreactive lymphocytes of patients with spinal cord injuries and amyotrophic lateral sclerosis were isolated and activated in vitro. These cells were cocultured with autologous MSC for 2-15 days. Cocultures of non-selected lymphocytes were used as controls. RESULTS: After 48 h of coculture, MSC adopted a spindle shape with polarization of the cytoplasm that resembled bipolar neurons. Their nuclei diminished the nucleolus number and the chromatin lost its granular appearance. After 15 days of culture the cells developed the typical structure of a neural network. No morphologic changes were observed in control cultures. The differentiated cells reacted positively to tubuline III, GFAP and nestin. No differences were observed between the different patient cell sources. DISCUSSION: We observed that autoreactive cells may induce the transdifferentiation of MSC to neural stem cells. This T-cell-MSC interaction may be a common phenomenon during physiologic nerve tissue repair.     

Epithelial stem cells and implications for wound repair

Activation of epithelial stem cells and efficient recruitment of their proliferating progeny plays a critical role in cutaneous wound healing. The reepithelialized wound epidermis has a mosaic composition consisting of progeny that can be traced back both to epidermal and several types of hair follicle stem cells. The contribution of hair follicle stem cells to wound epidermis is particularly intriguing as it involves lineage identity change from follicular to epidermal. Studies from our laboratory show that hair follicle-fated bulge stem cells commit only transient amplifying epidermal progeny that participate in the initial wound re-epithelialization, but eventually are outcompeted by other epidermal clones and largely disappear after a few months. Conversely, recently described stem cell populations residing in the isthmus portion of hair follicle contribute long-lasting progeny toward wound epidermis and, arguably, give rise to new interfollicular epidermal stem cells. The role of epithelial stem cells during wound healing is not limited to regenerating stratified epidermis. By studying regenerative response in large cutaneous wounds, our laboratory uncovered that epithelial cells in the center of the wound can acquire greater morphogenetic plasticity and, together with the underlying wound dermis, can engage in an embryonic-like process of hair follicle neogenesis. Future studies should uncover the cellular and signaling basis of this remarkable adult wound regeneration phenomenon.     

多能干细胞诱导分化及体细胞转分化为心肌细胞的研究进展

心血管疾病是威胁人类健康的重大疾病,而心肌细胞数量逐渐减少,甚至衰竭是其核心病变。心肌细胞补偿性替代治疗是未来用于治疗这类疾病的重要手段,因此,心肌细胞的来源和有效治疗将成为关键。目前,心肌细胞构建的主要方法有多能干细胞诱导分化成心肌祖细胞或心肌细胞、心源性心肌祖细胞,以及体细胞重编程等。其中,多能干细胞向心肌细胞分化是最常用的方法;而体细胞转分化技术相较于传统的诱导多潜能干细胞衍生心肌细胞缩短了时间窗,为潜在的心血管疾病治疗提供了另一种思路。随着获取心肌细胞效率及其质量的提升,未来心血管疾病的治疗将有望获得重大突破。     

Bone marrow-derived stem cells contribute skin regeneration in skin and soft tissue expansion

Skin and soft tissue expansion stimulates the proliferation of skin epidermal basal cells and increase the dermal collagen deposition and angiogenesis. To explore the contribution of bone marrow‐derived stem cells (BMSCs) to the generation of “new” skin during the expansion, we used a chimeric mouse model in which the donor C57BL mice were engrafted with the bone marrow of enhanced green fluorescent protein (EGFP) transgenic mice. BMSCs were collected from the tibia and femur of EGFP⁺ transgenic mice, and then injected into normal C57BL mice via the tail vein (chimeric mice). Skin was obtained at different times (days 0, 7, 14, 21, 28, and 35). Skin stromal‐derived factor‐1 (SDF‐1) expression was evaluated. The number, distribution, and phenotype changes of EGFP⁺ cells in the skin were also evaluated by means of fluorescent microscopy. EGFP⁺ cells were present stably in the normal skin. The number of EGFP⁺ cells of the Group A mice changed with the tension, and reached the peak on day 21(17.1 ± 6.7%), as compared with either Group B (5.5 ± 1.0%) or Group C (5.1 ± 0.9%). The SDF‐1 expression in the expanded skin was significant increased (≈11‐fold, P < 0.01) compared to non‐expanded skin on day 21. Immunofluorescence showed EGFP⁺ cells were converted into vascular endothelial cells, epidermal cells, and spindle‐shaped dermal fibroblasts. Strain can promote the expression of SDF‐1 and facilitate the differentiation and proliferation of BMSCs in the expanded skin. J. Cell. Physiol. 226: 2834–2840, 2011. © 2011 Wiley‐Liss, Inc.     

Mesenchymal stem cells and collagen patches for anterior cruciate ligament repair

AIM: To investigate collagen patches seeded with mesenchymal stem cells(MSCs) and/or tenocytes(TCs) with regards to their suitability for anterior cruciate ligament(ACL) repair. METHODS: Dynamic intraligamentary stabilization utilizes a dynamic screw system to keep ACL remnants in place and promote biological healing, supplemented by collagen patches. How these scaffolds interact with cells and what type of benefit they provide has not yet been investigated in detail. Primary ACL-derived TCs and human bone marrow derived MSCs were seeded onto two different types of 3D collagen scaffolds, Chondro-Gide?(CG) and Novocart?(NC). Cells were seeded onto the scaffolds and cultured for 7 d either as a pure populations or as "premix" containing a 1:1 ratio of TCs to MSCs. Additionally, as controls, cells were seeded in monolayers and in co-cultures on both sides of porous high-density membrane inserts(0.4 μm). We analyzed the patches by real time polymerase chain reaction, glycosaminoglycan(GAG), DNA and hydroxyproline(HYP) content. To determine cell spreading and adherence in the scaffolds microscopic imaging techniques, i.e., confocal laser scanning microscopy(c LSM) and scanning electron microscopy(SEM), were applied.RESULTS: CLSM and SEM imaging analysis confirmed cell adherence onto scaffolds. The metabolic cell activity revealed that patches promote adherence and proliferation of cells. The most dramatic increase in absolute metabolic cell activity was measured for CG samples seeded with tenocytes or a 1:1 cell premix. Analysis of DNA content and c LSM imaging also indicated MSCs were not proliferating as nicely as tenocytes on CG. The HYP to GAG ratio significantly changed for the premix group, resulting from a slightly lower GAG content, demonstrating that the cells are modifying the underlying matrix. Real-time quantitativepolymerase chain reaction data indicated that MSCs showed a trend of differentiation towards a more tenogenic-like phenotype after 7 d.CONCLUSION: CG and NC are both cyto-compatible with primary MSCs and TCs; TCs seemed to perform better on these collagen patches than MSCs.