Adipose-derived stromal cells: Their identity and uses in clinical trials, an update

In adults, adipose tissue is abundant and can be easily sampled using liposuction. Largely involved in obesity and associated metabolic disorders, it is now described as a reservoir of immature stromal cells. These cells, called adipose-derived stromal cells (ADSCs) must be distinguished from the crude stromal vascular fraction (SVF) obtained after digestion of adipose tissue. ADSCs share many features with mesenchymal stem cells derived from bone marrow, including paracrine activity, but they also display some specific features, including a greater angiogenic potential. Their angiogenic properties as well as their paracrine activity suggest a putative tumor-promoting role for ADSCs although contradictory data have been published on this issue. Both SVF cells and ADSCs are currently being investigated in clinical trials in several fields (chronic inflammation, ischemic diseases, etc. ). Apart from a phase Ⅲ trial on the treatment of fistula,most of these are in phaseⅠand use autologous cells. In the near future, the end results of these trials should provide a great deal of data on the safety of ADSC use.     

Current evidence on potential of adipose derived stem cells to enhance bone regeneration and future projection

Injuries to the postnatal skeleton are naturally repaired through successive steps involving specific cell types in a process collectively termed “bone regeneration”. Although complex, bone regeneration occurs through a series of well-orchestrated stages wherein endogenous bone stem cells play a central role. In most situations, bone regeneration is successful; however, there are instances when it fails and creates non-healing injuries or fracture nonunion requiring surgical or therapeutic interventions. Transplantation of adult or mesenchymal stem cells (MSCs) defined by the International Society for Cell and Gene Therapy (ISCT) as CD105+CD90+CD73+CD45-CD34-CD14orCD11b-CD79αorCD19-HLA-DR- is being investigated as an attractive therapy for bone regeneration throughout the world. MSCs isolated from adipose tissue, adipose-derived stem cells (ADSCs), are gaining increasing attention since this is the most abundant source of adult stem cells and the isolation process for ADSCs is straightforward. Currently, there is not a single Food and Drug Administration (FDA) approved ADSCs product for bone regeneration. Although the safety of ADSCs is established from their usage in numerous clinical trials, the bone-forming potential of ADSCs and MSCs, in general, is highly controversial. Growing evidence suggests that the ISCT defined phenotype may not represent bona fide osteoprogenitors. Transplantation of both ADSCs and the CD105^- sub-population of ADSCs has been reported to induce bone regeneration. Most notably, cells expressing other markers such as CD146, AlphaV, CD200, PDPN, CD164, CXCR4, and PDGFRα have been shown to represent osteogenic sub-population within ADSCs. Amongst other strategies to improve the bone-forming ability of ADSCs, modulation of VEGF, TGF-β1 and BMP signaling pathways of ADSCs has shown promising results. The U.S. FDA reveals that 73% of Investigational New Drug applications for stem cell-based products rely on CD105 expression as the “positive” marker for adult stem cells. A concerted effort involving the scientific community, clinicians, industries, and regulatory bodies to redefine ADSCs using powerful selection markers and strategies to modulate signaling pathways of ADSCs will speed up the therapeutic use of ADSCs for bone regeneration.     

脂肪干细胞在泌尿系统疾病中的研究进展

脂肪干细胞(adipose-derived stem cells,ADSCs)是一类从脂肪分离出来的具有自我更新及多向分化潜能的成体干细胞,ADSCs具有高度的可塑性,可分化成多种类型的细胞。与其他干细胞相比,ADSCs具有来源充足,取材方便,供体易接受等独特优势,已成为基础医学及临床治疗的研究热点。ADSCs诱导分化和移植可有效治疗多种组织损伤性疾病,改善或修复器官功能,近年来ADSCs作为细胞疗法及组织工程的新型种子细胞在泌尿系统疾病治疗中取得了重大进展。本文重点讨论ADSCs的生物学特性及其在泌尿系统疾病中的应用前景。     

Adipose tissue-derived stem cells show considerable promise for regenerative medicine applications

The stromal-vascular cell fraction (SVF) of adipose tissue can be an abundant source of both multipotent and pluripotent stem cells, known as adipose-derived stem cells or adipose tissue-derived stromal cells (ADSCs). The SVF also contains vascular cells, targeted progenitor cells, and preadipocytes. Stromal cells isolated from adipose tissue express common surface antigens, show the ability to adhere to plastic, and produce forms that resemble fibroblasts. They are characterized by a high proliferation potential and the ability to differentiate into cells of meso-, ecto- and endodermal origin. Although stem cells obtained from an adult organism have smaller capabilities for differentiation in comparison to embryonic and induced pluripotent stem cells (iPSs), the cost of obtaining them is significantly lower. The 40 years of research that mainly focused on the potential of bone marrow stem cells (BMSCs) revealed a number of negative factors: the painful sampling procedure, frequent complications, and small cell yield. The number of stem cells in adipose tissue is relatively large, and obtaining them is less invasive. Sampling through simple procedures such as liposuction performed under local anesthesia is less painful, ensuring patient comfort. The isolated cells are easily grown in culture, and they retain their properties over many passages. That is why adipose tissue has recently been treated as an attractive alternative source of stem cells. Essential aspects of ADSC biology and their use in regenerative medicine will be analyzed in this article.     

Transplantation of adipose-derived stem cells overexpressing hHGF into cardiac tissue

The delivery of adipose-derived stem cells (ADSCs) for promoting tissue repair has become a potential new therapy, while hepatocyte growth factor (HGF) is an important growth factor with angiogenic, antifibrotic, and anti-inflammatory benefits. Therefore, transplantation of ADSCs into acute myocardial infarction (AMI) may improve cardiac function through angiogenesis and anti-fibrosis, and that hHGF may enhance these effects. ADSCs were isolated from human subcutaneous adipose tissue. Lentivirus vector encoding human HGF (lenti-hHGF) was constructed and infected into ADSCs. Results indicated that transplantation of ADSCs led to improvement of left ventricular function, explained partly through their ability to differentiate into endothelial cells, resulting in increased blood flow and decreased fibrosis. Furthermore, hHGF enhanced these effects. This suggests that ADSCs combined with HGF gene transfer may be a useful strategy for the treatment of patients with ischemic heart disease.     

Therapeutic Potential of Human Adipose-Derived Stem Cells (ADSCs) from Cancer Patients: A Pilot Study

Mesenchymal stem cells from adipose tissue (ADSCs) are an important source of cells for regenerative medicine. The therapeutic effect of culture-expanded adipose derived stem cells has been shown; however, optimal xeno-free culture conditions remain to be determined. Cancer patients, specifically those undergoing invasive surgery, constitute a subgroup of patients who could benefit from autologous stem cell transplantation. Although regenerative potential of their ADSCs could be affected by the disease and/or treatment, we are not aware of any study that has evaluated the therapeutic potential of ADSCs isolated from cancer patients in reference to that of ADSCs derived from healthy subjects. Here we report that ADSCs isolated from subabdominal adipose tissue of patients with urological neoplasms yielded similar growth kinetics, presented equivalent mesenchymal surface markers and showed similar differentiation potential into distinct mesodermal cell lineages: adipocytes, chondroblasts and osteoblasts than ADSCs isolated from adipose tissue of age-matched non-oncogenic participants, all under xeno-free growth culture conditions. Molecular karyotyping of patient expanded ADSCs genomes showed no disease-related alterations indicating their safety. In addition, vesicles <100 nm identified as exosomes (EXOs) which may be at least partly responsible for the attributed therapeutic paracrine effects of the ADSCs were effectively isolated from ADSCs and showed equivalent miRNA content regardless they were derived from cancer patients or non-oncogenic participants indicating that the repair capabilities of xeno-free expanded ADSCs are not compromised by patient condition and therefore their xeno-free culture expanded ADSCs should be suitable for autologous stem cell transplantation in a clinical setting.     

Transplantation of human adipose tissue derived-SVF enhance liver function through high anti-inflammatory property

Although human adipose tissue-derived stromal vascular fraction (SVF) has been considered a promising source of stem cells, its characteristics relevant to treatment of a damaged liver have not been fully elucidated. In the present study, we sought to characterize the property of human SVF and determine the therapeutic utility of SVF in the liver cirrhosis model. We performed microarray, quantitative (q)-PCR experiments, and in vivo therapeutic assays using a liver cirrhotic mouse model. q-PCR results revealed that hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF)-A, Interleukin (IL)-10 and microRNA (miR)-146 were more highly upregulated in SVF than in human adipose-derived mesenchymal stem cells (ASCs). The SVF culture medium (CM) inhibited the activation of hepatic stellate cells in vitro. Injection of SVF significantly suppressed TAA-induced liver fibrosis and repaired liver function by inhibition of infiltrating inflammatory cells and induction of capillary/hepatocyte regeneration in vivo. Injection of IL-10 siRNA treated SVF cells decreased anti-inflammation and anti-fibrotic effects in TAA-induced mice liver. Our data indicate that SVF show a high anti-inflammatory property for treating fibrotic liver diseases through IL-10 secretion. Therefore, SVF might be a novel therapeutic alternative for the treatment of liver cirrhosis in clinical settings.     

Cytokines in adipose-derived mesenchymal stem cells promote the healing of liver disease

Adipose-derived mesenchymal stem cells(ADSCs) are a treatment cell source for patients with chronic liver injury. ADSCs are characterized by being harvested from the patient's own subcutaneous adipose tissue, a high cell yield(i.e., reduced immune rejection response), accumulation at a disease nidus, suppression of excessive immune response, production of various growth factors and cytokines, angiogenic effects, antiapoptotic effects, and control of immune cells via cellcell interaction. We previously showed that conditioned medium of ADSCs promoted hepatocyte proliferation and improved the liver function in a mouse model of acute liver failure. Furthermore, as found by many other groups, the administration of ADSCs improved liver tissue fibrosis in a mouse model of liver cirrhosis. A comprehensive protein expression analysis by liquid chromatography with tandem mass spectrometry showed that the various cytokines and chemokines produced by ADSCs promote the healing of liver disease. In this review, we examine the ability of expressed protein components of ADSCs to promote healing in cell therapy for liver disease. Previous studies demonstrated that ADSCs are a treatment cell source for patients with chronic liver injury. This review describes the various cytokines and chemokines produced by ADSCs that promote the healing of liver disease.     

Therapeutic applications of adipose-derived stromal vascular fractions in osteoarthritis

Osteoarthritis (OA) is considered to be a highly heterogeneous disease with progressive cartilage loss, subchondral bone remodeling, and low-grade inflammation. It is one of the world's leading causes of disability. Most conventional clinical treatments for OA are palliative drugs, which cannot fundamentally cure this disease. The stromal vascular fraction (SVF) from adipose tissues is a heterogeneous cell population. According to previous studies, it contains a large number of mesenchymal stem cells, which have been used to treat OA with good therapeutic results. This safe, simple, and effective therapy is expected to be applied and promoted in the future. In this paper, the detailed pathogenesis, diagnosis, and current clinical treatments for OA are introduced. Then, clinical studies and the therapeutic mechanism of SVF for the treatment of OA are summarized.     

Current applications of adipose-derived stem cells and their future perspectives

Adult stem cells have a great potential to treat various diseases. For these cell-based therapies, adipose-derived stem cells(ADSCs) are one of the most promising stem cell types, including embryonic stem cells(ESCs) and induced pluripotent stem cells(iPSCs). ESCs and iPSCs have taken center stage due to their pluripotency. However, ESCs and iPSCs have limitations in ethical issues and in identification of characteristics, respectively. Unlike ESCs and iPSCs, ADSCs do not have such limitations and are not only easily obtained but also uniquely expandable. ADSCs can differentiate into adipocytes, osteoblasts, chondrocytes, myocytes and neurons under specific differentiation conditions, and these kinds of differentiation potential of ADSCs could be applied in regenerative medicine e.g., skin reconstruction, bone and cartilage formation, etc. In this review, the current status of ADSC isolation, differentiation and their therapeutic applications are discussed.     

Adipose-Derived Stromal Vascular Fraction Cell Effects on a Rodent Model of Thin Endometrium

Endometrial dysfunction affects approximately 1% of infertile women, and there is currently no standard therapy for improving fertility treatment outcomes in these patients. In our study, we utilized a rodent model of thin endometrium to test whether intrauterine application of adipose-derived stromal vascular fraction cells (SVF) could improve morphological and physiological markers of endometrial receptivity. Using anhydrous ethanol, endometrial area and gland density were significantly reduced in our model of thin endometrium. Application of SVF was associated with a 29% reduction in endometrial vascular endothelial growth factor (VEGF) expression and significant increases in uterine artery systolic/diastolic velocity ratios and resistance index values, suggesting reduced diastolic microvascular tone. However, no significant improvements in endometrial area or gland density were observed following SVF treatment. 3D confocal imaging demonstrated poor engraftment of SVF cells into recipient tissue, which likely contributed to the negative results of this study. We suspect modified treatment protocols utilizing adjuvant estrogen and/or tail vein cell delivery may improve SVF retention and therapeutic response in subsequent studies. SVF is an easily-obtainable cell product with regenerative capability that may have a future role in the treatment of infertile women with endometrial dysfunction.     

In Vitro Evaluation of Endothelial Progenitor Cells from Adipose Tissue as Potential Angiogenic Cell Sources for Bladder Angiogenesis

Autologous endothelial progenitor cells (EPCs) might be alternative angiogenic cell sources for vascularization of tissue-engineered bladder, while isolation and culture of EPCs from peripheral blood in adult are usually time-consuming and highly inefficient. Recent evidence has shown that EPCs also exist in the adipose tissue. As adipose tissue is plentiful in the human body and can be easily harvested through a minimally invasive method, the aim of this study was to culture and characterize EPCs from adipose tissue (ADEPCs) and investigate their potential for the neovascularization of tissue-engineered bladder. Adipose stromal vascular fraction (SVF) was isolated and used for the culture of ADEPCs and adipose derived stem cells (ADSCs). After SVF was cultured for one week, ADEPCs with typical cobblestone morphology emerged and could be isolated from ADSCs according to their different responses to trypsinization. Rat bladder smooth muscle cells (RBSMCs) were isolated and cultured from rat bladder. RBSMCs exhibited typical spindle-shaped morphology. ADEPCs had higher proliferative potential than ADSCs and RBSMCs. ADEPCs stained positive for CD34, Stro-1, VEGFR-2, eNOS and CD31 but negative for α-SMA, CD14 and CD45. ADSCs stained positive for CD34, Stro-1 and α-SMA but negative for VEGFR-2, eNOS, CD31, CD14 and CD45. RBSMCs stained only positive for α-SMA. ADEPCs could be expanded from a single cell at an early passage to a cell cluster containing more than 10,000 cells. ADEPCs were able to uptake DiI-Ac-LDL, bind UEA-1 and form capillary-like structures in three-dimensional scaffolds (Matrigel and bladder acellular matrix). ADEPCs were also able to enhance the human umbilical vein endothelial cells’ capability of capillary-like tube formation on Matrigel. Additionally, significantly higher levels of mRNA and protein of vascular endothelial growth factor were found in ADEPCs than in RBSMCs. These results suggest the potential use of ADEPCs as angiogenic cell sources for engineering bladder tissue.     

Current applications of adipose-derived stem cells and their future perspectives简

Adult stem cells have a great potential to treat various diseases. For these cell-based therapies, adipose-derived stem cells (ADSCs) are one of the most promising stem cell types, including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). ESCs and iPSCs have taken center stage due to their pluripotency. However, ESCs and iPSCs have limitations in ethical issues and in identification of characteristics, respectively. Unlike ESCs and iPSCs, ADSCs do not have such limitations and are not only easily obtained but also uniquely expandable. ADSCs can differentiate into adipocytes, osteoblasts, chondrocytes, myocytes and neurons under specific differentiation conditions, and these kinds of differentiation potential of ADSCs could be applied in regenerative medicine e.g., skin reconstruction, bone and cartilage formation, etc. In this review, the current status of ADSC isolation, differentiation and their therapeutic applications are discussed.     

In Vitro Evaluation of Different Methods of Handling Human Liposuction Aspirate and Their Effect on Adipocytes and Adipose Derived Stem Cells

Nowadays, fat tissue transplantation is widely used in regenerative and reconstructive surgery. However, a shared method of lipoaspirate handling for ensuring a good quality fat transplant has not yet been established. The study was to identify a method to recover from the lipoaspirate samples the highest number of human viable adipose tissue‐derived stem cells (hADSCs) included in stromal vascular fraction (SVF) cells and of adipocytes suitable for transplantation, avoiding an extreme handling. We compared the lipoaspirate spontaneous stratification (10‐20‐30 min) with the centrifugation technique at different speeds (90‐400‐1500 × g). After each procedure, lipoaspirate was separated into top oily lipid layer, liquid fraction, “middle layer”, and bottom layer. We assessed the number of both adipocytes in the middle layer and SVF cells in all layers. The histology of middle layer and the surface phenotype of SVF cells by stemness markers (CD105+, CD90+, CD45?) was analyzed as well. The results showed a normal architecture in all conditions except for samples centrifuged at 1500 × g. In both methods, the flow cytometry analysis showed that greater number of ADSCs was in middle layer; in the fluid portion and in bottom layer was not revealed significant expression levels of stemness markers. Our findings indicate that spontaneous stratification at 20 min and centrifugation at 400 × g are efficient approaches to obtain highly viable ADSCs cells and adipocytes, ensuring a good thickness of lipoaspirate for autologous fat transfer. Since an important aspect of surgery practice consists of gain time, the 400 × g centrifugation could be the recommended method when the necessary instrumentation is available. J. Cell. Physiol. 230: 1974–1981, 2015. © 2015 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.

     

脂肪干细胞的诱导分化及其来源的研究

目的:通过组织块培养法得到脂肪干细胞(adipose-derived stem cells,ADSCs),探讨其诱导分化潜能,并初步研究ADSCs的来源。方法:用脂肪组织块培养法培养原代人ADSCs。第三代ADSCs进行成脂和成骨诱导分化,分别用油红O和茜素红S染色进行鉴定。脂肪组织块培养七天后取脂肪组织进行Hematoxylin-eosin Staining(HE)染色观察ADSCs组织分布。结果:用脂肪组织块培养法成功培养出原代人ADSCs。ADSCs传代到第8代,依然保持着良好的增殖能力和细胞形态。ADSCs能成功诱导成脂肪细胞和骨细胞。通过对培养七天后的脂肪组织块进行HE染色,发现ADSCs主要分布在脂肪组织的间质血管和结缔组织周围。结论:用脂肪组织块培养出来的ADSCs具有成脂和成骨分化的潜能。ADSCs主要定位于间质血管和结缔组织周围。     

基质血管组分(SVF)临床分离技术研究

脂肪组织易获取、组织相容性好且对供体影响小,可作为获得成体干细胞的重要来源。基质血管组分(SVF)是从脂肪中分离出来的包括脂源性干细胞(ADSC)和基质细胞的异质性细胞群。SVF促进组织的修复和再生已被大量的临床实验所证实,尤其是在美容整形和组织修复中的应用。早期,SVF通过酶消化法获得,随着近年来在临床中扩大应用,为确保患者安全和质量可控,开发出新型自动分离设备。同时,为符合一些国家监管要求,避免酶的使用,采用非酶消化法获取SVF。因此,该文主要针对基于酶消化法和非酶消化法已经发表临床分离方法和上市的相关设备作详细论述。     

Fat tissue: an underappreciated source of stem cells for biotechnology

Adipose tissue can be harvested in large amounts with minimal morbidity. It contains numerous cells types, including adipocytes, preadipocytes, vascular endothelial cells and vascular smooth muscle cells; it also contains cells that have the ability to differentiate into several lineages, such as fat, bone, cartilage, skeletal, smooth, and cardiac muscle, endothelium, hematopoietic cells, hepatocytes and neuronal cells. Cloning studies have shown that some adipose-derived stem cells (ADSCs) have multilineage differentiation potential. ADSCs are also capable of expressing multiple growth factors, including vascular endothelial growth factor and hepatocyte growth factor. Early, uncontrolled, non-randomized clinical research, applying fresh adipose-derived cells into a cranial defect or undifferentiated ADSCs into fistulas in Crohn's disease, has shown healing and an absence of side effects. The combination of these properties, and the large quantity of cells that can be obtained from fat, suggests that this tissue will be a useful tool in biotechnology.     

Autologous cell therapy for cisplatin-induced acute kidney injury by using non-expanded adipose tissue-derived cells

Abstract Background aims. Recent studies have demonstrated that cultured mesenchymal stromal cells derived from adipose tissue are useful for regenerative cell therapy. The stromal vascular fraction (SVF) can be obtained readily without culturing and may be clinically applicable. We investigated the therapeutic effects of SVF and used it in the treatment of acute kidney injury (AKI). Methods. Liposuction aspirates were obtained from healthy donors who had provided written informed consent. We harvested the SVF and determined the growth factor secretion and anti-apoptotic ability with conditioned medium. To investigate the effect of SVF on AKI, cisplatin was injected into rats and SVF was administrated into the subcupsula of the kidney. Results. Both human and rat SVF cells secreted vascular endothelial growth factor-A (VEGF) and hepatocyte growth factor (HGF). Human SVF-conditioned media had an anti-apoptotic effect, which was inhibited by anti-HGF antibody (Ab) but not by anti-VEGF Ab. In vivo, SVF significantly ameliorated renal function, attenuated tubular damage and increased the cortical blood flow speed. In the SVF-treated group, VEGF levels in the cortex and HGF levels in both the cortex and medulla, especially tubules in the medulla, were significantly higher. Immunostaining revealed that SVF cells expressing VEGF and HGF and remained in the subcapsule on day 14. Conclusions. The present study demonstrates that a subcapsular injection of non-expanded SVF cells ameliorates rat AKI, and that the mechanism probably involves secretion of renoprotective molecules. Administration of human SVF may be clinically applicable and useful as a novel autologous cell therapy against kidney diseases.