Promotion of wound healing using adipose-derived stem cells in radiation ulcer of a rat model

Background

Wound healing is a complex biologic process that involves the integration of inflammation, mitosis, angiogenesis, synthesis, and remodeling of the extracellular matrix. However, some wounds fail to heal properly and become chronic. Although some simulated chronic wound models have been established, an efficient approach to treat chronic wounds in animal models has not been determined. The aim of this study was to develop a modified rat model simulating the chronic wounds caused by clinical radiation ulcers and examine the treatment of chronic wounds with adipose-derived stem cells.

Results

Sprague–Dawley rats were irradiated with an electron beam, and wounds were created. The rats received treatment with adipose-derived stem cells (ASCs), and a wound-healing assay was performed. The wound sizes after ASC treatment for 3 weeks were significantly smaller compared with the control condition (p < 0.01). Histological observations of the wound edge and immunoblot analysis of the re-epithelialization region both indicated that the treatment with ASCs was associated with the development of new blood vessels. Cell-tracking experiments showed that ASCs were colocalized with endothelial cell markers in ulcerated tissues.

Conclusions

We established a modified rat model of radiation-induced wounds and demonstrated that ASCs accelerate wound-healing.     

Adipose-derived mesenchymal stromal/stem cells: An update on their phenotype in vivo and in vitro

Adipose tissue is a rich, ubiquitous and easily acces-sible source for multipotent stromal/stem cells and has, therefore, several advantages compared to other sourc-es of mesenchymal stromal/stem cells. Several studies have tried to identify the origin of the stromal/stem cell population within adipose tissue in situ. This is a complicated attempt because no marker has currently been described which unambiguously identifies native adipose-derived stromal/stem cells(ASCs). Isolated and cultured ASCs are a non-uniform preparation consisting of several subsets of stem and precursor cells. Cultured ASCs are characterized by their expression of a panel of markers(and the absence of others), whereas their in vitro phenotype is dynamic. Some markers were ex-pressed de novo during culture, the expression of some markers is lost. For a long time, CD34 expression was solely used to characterize haematopoietic stem and progenitor cells, but now it has become evident that it is also a potential marker to identify an ASC subpopula-tion in situ and after a short culture time. Nevertheless, long-term cultured ASCs do not express CD34, perhaps due to the artificial environment. This review gives an update of the recently published data on the origin and phenotype of ASCs both in vivo and in vitro. In addition, the composition of ASCs(or their subpopula-tions) seems to vary between different laboratories andpreparations. This heterogeneity of ASC preparationsmay result from different reasons. One of the main problems in comparing results from different laborato-ries is the lack of a standardized isolation and culture protocol for ASCs. Since many aspects of ASCs, suchas the differential potential or the current use in clinical trials, are fully described in other recent reviews, this review further updates the more basic research issues concerning ASCs' subpopulations, heterogeneity andculture standardization.     

脂肪干细胞在再生医学中的应用

再生医学是一门研究如何促进创伤与组织再生及功能重建的新兴学科,主要通过研究干细胞分化、机体等正常组织创伤修复与再生等机制来维持、修复、再生或改善损伤组织和器官功能。脂肪干细胞（adipose-derived stem cells,ASCs）是近年来从脂肪组织中分离得到的一种具有多向分化潜能的干细胞,是一种足量的、可用于实际的、有一定吸引力的自体细胞代替的供体资源,并能够广泛的用于组织修复、再生、发育的可塑性及细胞治疗等研究中。阐述了脂肪干细胞在旁分泌、软组织重建及损伤修复、骨骼肌重建、心血管重建、神经系统重建及癌症转移与入侵方面的作用模式,概括总结了目前利用脂肪干细胞参与的临床治疗方法,以期对脂肪干细胞在再生医学中应用研究提供参考。     

WJSC 6th Anniversary Special Issues（2）:Mesenchymal stem cellsAdipose mesenchymal stem cells in the field of bone tissue engineering

Bone tissue engineering represents one of the most challenging emergent fields for scientists and clinicians.Current failures of autografts and allografts in many pathological conditions have prompted researchers to find new biomaterials able to promote bone repair or regeneration with specific characteristics of biocompatibility,biodegradability and osteoinductivity.Recent advancements for tissue regeneration in bone defects have occurred by following the diamond concept and combining the use of growth factors and mesenchymal stem cells(MSCs).In particular,a more abundant and easily accessible source of MSCs was recently discovered in adipose tissue.These adipose stem cells(ASCs)can be obtained in large quantities with little donor site morbidity or patient discomfort,in contrast to the invasive and painful isolation of bone marrow MSCs.The osteogenic potential of ASCs on scaffolds has been examined in cell cultures and animal models,with only a few cases reporting the use of ASCs for successful reconstruction or accelerated healing of defects of the skull and jaw in patients.Although these reports extend our limited knowledge concerning the use of ASCs for osseous tissue repair and regeneration,the lack of standardization in applied techniques makes the comparison between studies difficult.Additional clinical trials are needed to assess ASC therapy and address potential ethical and safety concerns,which must be resolved to permit application in regenerative medicine.     

0Adipose-derived stem cells: Implications in tissue regeneration

Adipose-derived stem cells(ASCs) are mesenchymal stem cells(MSCs) that are obtained from abundant adipose tissue, adherent on plastic culture flasks, can be expanded in vitro, and have the capacity to differ-entiate into multiple cell lineages. Unlike bone marrow-derived MSCs, ASCs can be obtained from abundant adipose tissue by a minimally invasive procedure, which results in a high number of cells. Therefore, ASCs are promising for regenerating tissues and organs dam-aged by injury and diseases. This article reviews the implications of ASCs in tissue regeneration.     

Mesenchymal stem cell-derived exosomes: A novel and potential remedy for cutaneous wound healing and regeneration

Poor healing of cutaneous wounds is a common medical problem in the field of traumatology. Due to the intricate pathophysiological processes of wound healing, the use of conventional treatment methods, such as chemical molecule drugs and traditional dressings, have been unable to achieve satisfactory outcomes. Within recent years, explicit evidence suggests that mesenchymal stem cells (MSCs) have great therapeutic potentials on skin wound healing and regeneration. However, the direct application of MSCs still faces many challenges and difficulties. Intriguingly, exosomes as cell-secreted granular vesicles with a lipid bilayer membrane structure and containing specific components from the source cells may emerge to be excellent substitutes for MSCs. Exosomes derived from MSCs (MSC-exosomes) have been demonstrated to be beneficial for cutaneous wound healing and accelerate the process through a variety of mechanisms. These mechanisms include alleviating inflammation, promoting vascularization, and promoting proliferation and migration of epithelial cells and fibroblasts. Therefore, the application of MSC-exosomes may be a promising alternative to cell therapy in the treatment of cutaneous wounds and could promote wound healing through multiple mechanisms simultaneously. This review will provide an overview of the role and the mechanisms of MSC-derived exosomes in cutaneous wound healing, and elaborate the potentials and future perspectives of MSC-exosomes application in clinical practice.     

Adipose-derived stem cells: Implications in tissue regeneration

Adipose-derived stem cells (ASCs) are mesenchymal stem cells (MSCs) that are obtained from abundant adipose tissue, adherent on plastic culture flasks, can be expanded in vitro, and have the capacity to differentiate into multiple cell lineages. Unlike bone marrow-derived MSCs, ASCs can be obtained from abundant adipose tissue by a minimally invasive procedure, which results in a high number of cells. Therefore, ASCs are promising for regenerating tissues and organs damaged by injury and diseases. This article reviews the implications of ASCs in tissue regeneration.     

Osteogenic potential: comparison between bone marrow and adipose-derived mesenchymal stem cells

Bone tissue engineering(BTE) is now a promising re-search issue to improve the drawbacks from traditional bone grafting procedure such as limited donor sources and possible complications. Stem cells are one of the major factors in BTE due to the capability of self re-newal and multi-lineage differentiation. Unlike embry-onic stem cells, which are more controversial in ethical problem, adult mesenchymal stem cells are considered to be a more appropriate cell source for BTE. Bone marrow mesenchymal stem cells(BMSCs) are the ear-liest-discovered and well-known stem cell source using in BTE. However, the low stem cell yield requiring long expansion time in vitro, pain and possible morbidities during bone marrow aspiration and poor proliferation and osteogenic ability at old age impede its' clinical ap-plication. Afterwards, a new stem cell source coming from adipose tissue, so-called adipose-derived stemcells(ASCs), is found to be more suitable in clinical ap-plication because of high stem cells yield from lipoaspi-rates, faster cell proliferation and less discomfort and morbidities during harvesting procedure. However, the osteogenic capacity of ASCs is now still debated be-cause most papers described the inferior osteogenesis of ASCs than BMSCs. A better understanding of the osteogenic differences between ASCs and BMSCs is crucial for future selection of cells in clinical application for BTE. In this review, we describe the commonality and difference between BMSCs and ASCs by cell yield, cell surface markers and multiple-differentiation poten-tial. Then we compare the osteogenic capacity in vitro and bone regeneration ability in vivo between BMSCs and ASCs based on the literatures which utilized both BMSCs and ASCs simultaneously in their articles. The outcome indicated both BMSCs and ASCs exhibited the osteogenic ability to a certain extent both in-vitro and in-vivo. However, most in-vitro study papers verified the inferior osteogenesis of ASCs; conversely, in-vivo research reviews revealed more controversies in this issue. We expect the new researchers can have a quick understanding of the progress in this filed and design a more comprehensive research based on this review.     

The use of adipose-derived stem cells as sheets for wound healing

Cellular therapies have shown immense promise in the treatment of nonhealing wounds. Cell sheets are an emerging strategy in tissue engineering, and these cell sheets are promising as a delivery method of mesenchymal stem cells to the wound bed. Cell sheet technology utilizes temperature dependent polymers to allow for lifting of cultured cells and extracellular matrix without the use of digestive enzymes. While mesenchymal stem cells (MSCs) have shown success in cell sheets for myocardial repair, examination of cell sheets in the field of wound healing has been limited. We previously developed a novel cell sheet composed of human adipose-derived stem cells (ASCs). Both single and triple layer cell sheets were examined in a full-thickness murine wound model. The treatment cell sheets were compared with untreated controls and analyzed at timepoints of 7, 14, 18 and 21 d. The ASC cell sheets showed increased healing at 7, 14 and 18 d, and this effect was increased in the triple layer cell sheet group. Future development of these cell sheets will focus on increasing angiogenesis in the wound bed, utilizing multiple cell types, and examining allogeneic cell sheets. Here we review our experiment, expand upon our future directions and discuss the potential of an off-the-shelf cell sheet. In the field of wound healing, such a cell sheet is both clinically and scientifically exciting.     

Hair follicle stem cell progeny heal blisters while pausing skin development

Injury in adult tissue generally reactivates developmental programs to foster regeneration, but it is not known whether this paradigm applies to growing tissue. Here, by employing blisters, we show that epidermal wounds heal at the expense of skin development. The regenerated epidermis suppresses the expression of tissue morphogenesis genes accompanied by delayed hair follicle (HF) growth. Lineage tracing experiments, cell proliferation dynamics, and mathematical modeling reveal that the progeny of HF junctional zone stem cells, which undergo a morphological transformation, repair the blisters while not promoting HF development. In contrast, the contribution of interfollicular stem cell progeny to blister healing is small. These findings demonstrate that HF development can be sacrificed for the sake of epidermal wound regeneration. Our study elucidates the key cellular mechanism of wound healing in skin blistering diseases.     

Effects of radiation and chemotherapy on adipose stem cells: Implications for use in fat grafting in cancer patients

Autologous fat transplantation is a versatile tool in reconstructive surgery. Adipose-derived stem cells (ASCs) increase survival of fat grafts and thus are increasingly used for breast reconstruction in breast cancer patients. However, radiation and/or chemotherapy have been proposed to inhibit soft tissue regeneration in wound healing thus suggesting alteration in stem cell pathways. Therefore, elucidating effects of radiation and chemotherapy on ASCs is critical if one desires to enhance the survival of fat grafts in patients. This review outlines our work evaluating the function and recoverability of ASCs from radiation or chemotherapy patients, focusing specifically on their availability as a source of autologous stem cells for fat grafting and breast reconstruction in cancer patients. Even though evidence suggests radiation and chemotherapy negatively influence ASCs at the cellular level, the efficiency of the isolation and differentiation capacity did not appear influenced in patients after receiving chemotherapy treatment, although fat from radiated patients exhibited significantly altered ASC differentiation into endothelial-like cells. Further, the in vitro growth rates of patient’s ASCs do not differ significantly before or after treatment. Taken together, these studies suggest ASCs as an important new tool for grafting and reconstruction even when radiation and chemotherapy treatment are involved.     

Adipose tissue,angiogenesis and angio-MIR under physiological and pathological conditions

Angiogenesis is a crucial process for the maintenance of normal tissue physiology and it is involved in tissue remodeling and regeneration. This process is essential for adipose tissue maintenance. The adipose tissue is composed by different cell types including stromal vascular cells as well as adipose stem cells (ASCs). In particular, ASCs are multipotent somatic stem cells that are able to differentiate and secrete several growth factors; they are recently emerging as a new cell reservoir for novel therapies and strategies in many diseases. Several studies suggest that ASCs have peculiar properties and participate in different disease-related processes such as angiogenesis. Furthermore, pathological expansion of adipose tissue brings to hypoxia, a major condition of unhealthy angiogenesis.Recent evidences have shown that microRNAs (miRNAs) play a crucial role also on ASCs as they take part in stemness maintenance, proliferation, and differentiation. It has been suggested that some miRNAs (MIR126, MIR31, MIR221 MIR222, MIR17-92 cluster, MIR30, MIR100 and MIR486) are directly involved in the angiogenic process by controlling multiple genes involved in this pathway. With the present review, we aim at providing an updated summary of the importance of adipose tissue under physiological and pathological conditions and of its relationship with neovascularization process. In particular, we report an overview of the most important miRNAs involved in angiogenesis focusing on ASCs. Hopefully the data presented will bring benefit in developing new therapeutic strategies.     

Single-cell RNA sequencing in cornea research: Insights into limbal stem cells and their niche regulation

The corneal epithelium is composed of stratified squamous epithelial cells on the outer surface of the eye, which acts as a protective barrier and is critical for clear and stable vision. Its continuous renewal or wound healing depends on the proliferation and differentiation of limbal stem cells (LSCs), a cell population that resides at the limbus in a highly regulated niche. Dysfunction of LSCs or their niche can cause limbal stem cell deficiency, a disease that is manifested by failed epithelial wound healing or even blindness. Nevertheless, compared to stem cells in other tissues, little is known about the LSCs and their niche. With the advent of single-cell RNA sequencing, our understanding of LSC characteristics and their microenvironment has grown considerably. In this review, we summarized the current findings from single-cell studies in the field of cornea research and focused on important advancements driven by this technology, including the heterogeneity of the LSC population, novel LSC markers and regulation of the LSC niche, which will provide a reference for clinical issues such as corneal epithelial wound healing, ocular surface reconstruction and interventions for related diseases.     

A novel composition for the culture of human adipose stem cells which includes complement C3

Adipose tissue is an easily accessible and abundant source of stem cells. Adipose stem cells (ASCs) are currently being researched as treatment options for repair and regeneration of damaged tissues. The standard culture conditions used for expansion of ASCs contain fetal bovine serum (FBS) which is undefined, could transmit known and unknown adventitious agents, and may cause adverse immune reactions. We have described a novel culture condition which excludes the use of FBS and characterised the resulting culture. Human ASCs were cultured in the novel culture medium, which included complement protein C3. These cultures, called C-ASCs, were compared with ASCs cultured in medium supplemented with FBS. Analysis of ASCs for surface marker profile, proliferation characteristics and differentiation potential indicated that the C-ASCs were similar to ASCs cultured in medium containing FBS. Using a specific inhibitor, we show that C3 is required for the survival of C-ASCs. This novel composition lends itself to being developed into a defined condition for the routine culture of ASCs for basic and clinical applications.     

组蛋白乙酰化对成体干细胞生物学性状的影响

成体干细胞（adult stem cells,ASCs）是指存在于一种已经分化组织中的未分化细胞,它们可以再生修复损伤的组织和器官,是组织工程和细胞治疗的理想细胞。但是ASCs在体外扩增过程中容易发生自主分化和衰老,影响其在临床的广泛应用。组蛋白乙酰化作为表观遗传调节的重要机制,参与细胞分化、衰老及凋亡等众多细胞活动的调控。该文就组蛋白乙酰化对成体干细胞生物学性状的影响进行综述。     

Exosomes from adipose‐derived stem cells and application to skin wound healing

Skin wound healing is an intractable problem that represents an urgent clinical need. To solve this problem, a large number of studies have focused on the use of exosomes (EXOs) derived from adipose‐derived stem cells (ADSCs). This review describes the mechanisms whereby ADSCs‐EXOs regulate wound healing and their clinical application. In the wound, ADSCs‐EXOs modulate immune responses and inflammation. They also promote angiogenesis, accelerate proliferation and re‐epithelization of skin cells, and regulate collagen remodelling which inhibits scar hyperplasia. Compared with ADSCs therapeutics, ADSCs‐EXOs have highly stability and are easily stored. Additionally, they are not rejected by the immune system and have a homing effect and their dosage can be easily controlled. ADSCs‐EXOs can improve fat grafting and promote wound healing in patients with diabetes mellitus. They can also act as a carrier and combined scaffold for treatment, leading to scarless cutaneous repair. Overall, ADSCs‐EXOs have the potential to be used in the clinic to promote wound healing.     

Inflammatory bowel disease: Therapeutic limitations and prospective of the stem cell therapy

Inflammatory bowel disease(IBD), consisting primarily of ulcerative colitis and Crohn's disease, is a group of debilitating auto-immune disorders, which also increases the risk of colitis-associated cancer. However, due to the chronic nature of the disease and inconsistent treatment outcomes of current anti-IBD drugs(e.g., approximately 30% non-responders to anti-TNFα agents), and related serious side effects, about half of all IBD patients(in millions) turn to alternative treatment options. In this regard, mucosal healing is gaining acceptance as a measure of disease activity in IBD patients as recent studies have correlated the success of mucosal healing with improved prognosis. However, despite the increasing clinical realization of the significance of the concept of mucosal healing, its regulation and means of therapeutic targeting remain largely unclear. Here, stemcell therapy, which uses hematopoietic stem cells or mesenchymal stem cells, remains a promising option. Stem cells are the pluripotent cells with ability to differentiate into the epithelial and/or immune-modulatory cells. The overreaching concept is that the stem cells can migrate to the damaged areas of the intestine to provide curative help in the mucosal healing process. Moreover, by differentiating into the mature intestinal epithelial cells, the stem cells also help in restoring the barrier integrity of the intestinal lining and hence prevent the immunomodulatory induction, the root cause of the IBD. In this article, we elaborate upon the current status of the clinical management of IBD and potential role of the stem cell therapy in improving IBD therapy and patient's quality of life.     

The potential of adipose‐derived stem cells in craniofacial repair and regeneration

The recent identification of a mesenchymal stem cell population in adipose tissue has led to an abundance of research focused on the regenerative properties of these cells. As such, adipose‐derived stem cells (ASCs) and potential therapies in craniofacial regeneration have been widely studied. This review will discuss the identification and potential of ASCs, and specifically, preclinical and clinical studies using ASCs in craniofacial repair. Studies involving ASCs in the repair of defects caused by craniosynostosis and Treacher Collins syndrome will be discussed. A comprehensive review of the literature will be presented, focusing on fat grafting and biomaterials‐based approaches that include ASCs for craniofacial regeneration. (Part C) 96:95–97, 2012. © 2012 Wiley Periodicals, Inc.     

Antler stem cells and their potential in wound healing and bone regeneration

Compared to other vertebrates, the regenerative capacity of appendages in mammals is very limited. Deer antlers are an exception and can fully regenerate annually in postnatal mammals. This process is initiated by the antler stem cells (AnSCs). AnSCs can be divided into three types: (1) Antlerogenic periosteum cells (for initial pedicle and first antler formation); (2) Pedicle periosteum cells (for annual antler regeneration); and (3) Reserve mesenchyme cells (RMCs) (for rapid antler growth). Previous studies have demonstrated that AnSCs express both classic mesenchymal stem cells (MSCs) and embryonic stem cells (ESCs), and are able to differentiate into multiple cell types in vitro. Thus, AnSCs were defined as MSCs, but with partial ESC attributes. Near-perfect generative wound healing can naturally occur in deer, and wound healing can be achieved by the direct injection of AnSCs or topical application of conditioned medium of AnSCs in rats. In addition, in rabbits, the use of both implants with AnSCs and cell-free preparations derived from AnSCs can stimulate osteogenesis and repair defects of bone. A more comprehensive understanding of AnSCs will lay the foundation for developing an effective clinical therapy for wound healing and bone repair.     

Monolayer cell expansion conditions affect the chondrogenic potential of adipose-derived stem cells

Adipose-derived stem cells (ASCs) are an abundant, readily available population of multipotent progenitor cells that reside in adipose tissue. Isolated ASCs are typically expanded in monolayer on standard tissue culture plastic with a basal medium containing 10% fetal bovine serum. However, recent data suggest that altering the monolayer expansion conditions by using suspension culture plastic, adding growth factors to the medium, or adjusting the seeding density may affect the self-renewal rate, multipotency, and lineage-specific differentiation potential of the ASCs. We hypothesized that variation in any of these expansion conditions would influence the chondrogenic potential of ASCs. ASCs were isolated from human liposuction waste tissue and expanded through two passages with different tissue culture plastic, feed medium, and cell seeding densities. Once expanded, the cells were cast in an agarose gel and subjected to identical chondrogenic culture conditions for 7 days, at which point cell viability, radiolabel incorporation, and gene expression were measured. High rates of matrix synthesis upon chondrogenic induction were mostly associated with smaller cells, as indicated by cell width and area on tissue culture plastic, and it appears that expansion in a growth factor supplemented medium is important in maintaining this morphology. All end-point measures were highly dependent on the specific monolayer culture conditions. These results support the hypothesis that monolayer culture conditions may "prime" the cells or predispose them towards a specific phenotype and thus underscore the importance of early culture conditions in determining the growth and differentiation potential of ASCs.