Stem-cell-mediated bone repair has been used in clinical trials for the regeneration of large craniomaxillofacial defects, to slow the process of bone degeneration in patients with osteonecrosis of ...the femoral head and for prophylactic treatment of distal tibial fractures. Successful regenerative outcomes in these investigations have provided a solid foundation for wider use of stromal cells in skeletal repair therapy. However, employing stromal cells to facilitate or enhance bone repair is far from being adopted into clinical practice. Scientific, technical, practical and regulatory obstacles prevent the widespread therapeutic use of stromal cells. Ironically, one of the major challenges lies in the limited understanding of the mechanisms via which transplanted cells mediate regeneration. Animal models have been used to provide insight, but these models largely fail to reproduce the nuances of human diseases and bone defects. Consequently, the development of targeted approaches to optimize cell-mediated outcomes is difficult. In this Review, we highlight the successes and challenges reported in several clinical trials that involved the use of bone-marrow-derived mesenchymal or adipose-tissue-derived stromal cells. We identify several obstacles blocking the mainstream use of stromal cells to enhance skeletal repair and highlight technological innovations or areas in which novel techniques might be particularly fruitful in continuing to advance the field of skeletal regenerative medicine.
Fat grafting is used to restore breast defects after surgical resection of breast tumors. Supplementing fat grafts with adipose tissue-derived stromal/stem cells (ASCs) is proposed to improve the ...regenerative/restorative ability of the graft and retention. However, long term safety for ASC grafting in proximity of residual breast cancer cells is unknown. The objective of this study was to determine the impact of human ASCs derived from abdominal lipoaspirates of three donors, on a human breast cancer model that exhibits early metastasis.
Human MDA-MB-231 breast cancer cells represents "triple negative" breast cancer that exhibits early micrometastasis to multiple mouse organs 1. Human ASCs were derived from abdominal adipose tissue from three healthy female donors. Indirect co-culture of MDA-MB-231 cells with ASCs, as well as direct co-culture demonstrated that ASCs had no effect on MDA-MB-231 growth. Indirect co-culture, and ASC conditioned medium (CM) stimulated migration of MDA-MB-231 cells. ASC/RFP cells from two donors co-injected with MDA-MB-231/GFP cells exhibited a donor effect for stimulation of primary tumor xenografts. Both ASC donors stimulated metastasis. ASC/RFP cells were viable, and integrated with MDA-MB-231/GFP cells in the tumor. Tumors from the co-injection group of one ASC donor exhibited elevated vimentin, matrix metalloproteinase-9 (MMP-9), IL-8, VEGF and microvessel density. The co-injection group exhibited visible metastases to the lung/liver and enlarged spleen not evident in mice injected with MDA-MB-231/GFP alone. Quantitation of the total area of GFP fluorescence and human chromosome 17 DNA in mouse organs, H&E stained paraffin sections and fluorescent microscopy confirmed multi-focal metastases to lung/liver/spleen in the co-injection group without evidence of ASC/RFP cells.
Human ASCs derived from abdominal lipoaspirates of two donors stimulated metastasis of MDA-MB-231 breast tumor xenografts to multiple mouse organs. MDA-MB-231 tumors co-injected with ASCs from one donor exhibited partial EMT, expression of MMP-9, and increased angiogenesis.
The emerging field of regenerative medicine will require a reliable source of stem cells in addition to biomaterial scaffolds and cytokine growth factors. Adipose tissue represents an abundant and ...accessible source of adult stem cells with the ability to differentiate along multiple lineage pathways. The isolation, characterization, and preclinical and clinical application of adipose-derived stem cells (ASCs) are reviewed in this article.
Abstract Background aims Adipose tissue is a rich and very convenient source of cells for regenerative medicine therapeutic approaches. However, a characterization of the population of ...adipose-derived stromal and stem cells (ASCs) with the greatest therapeutic potential remains unclear. Under the authority of International Federation of Adipose Therapeutics and International Society for Cellular Therapy, this paper sets out to establish minimal definitions of stromal cells both as uncultured stromal vascular fraction (SVF) and as an adherent stromal/stem cells population. Methods Phenotypic and functional criteria for the identification of adipose-derived cells were drawn from the literature. Results In the SVF, cells are identified phenotypically by the following markers: CD45-CD235a-CD31-CD34+. Added value may be provided by both a viability marker and the following surface antigens: CD13, CD73, CD90 and CD105. The fibroblastoid colony-forming unit assay permits the evaluation of progenitor frequency in the SVF population. In culture, ASCs retain markers in common with other mesenchymal stromal/stem cells (MSCs), including CD90, CD73, CD105, and CD44 and remain negative for CD45 and CD31. They can be distinguished from bone-marrow-derived MSCs by their positivity for CD36 and negativity for CD106. The CFU-F assay is recommended to calculate population doublings capacity of ASCs. The adipocytic, chondroblastic and osteoblastic differentiation assays serve to complete the cell identification and potency assessment in conjunction with a quantitative evaluation of the differentiation either biochemically or by reverse transcription polymerase chain reaction. Conclusions The goal of this paper is to provide initial guidance for the scientific community working with adipose-derived cells and to facilitate development of international standards based on reproducible parameters.
Stem cells have been long looked at as possible therapeutic vehicles for different health related problems. Among the different existing stem cell populations, Adipose- derived Stem Cells (ASCs) have ...been gathering attention in the last 10 years. When compared to other stem cells populations and sources, ASCs can be easily isolated while providing simultaneously higher yields upon the processing of adipose tissue. Similar to other stem cell populations, it was initially thought that the main potential of ASCs for regenerative medicine approaches was intimately related to their differentiation capability. Although this is true, there has been an increasing body of literature describing the trophic effects of ASCs on the protection, survival and differentiation of variety of endogenous cells/tissues. Moreover, they have also shown to possess an immunomodulatory character. This effect is closely related to the ASCs' secretome and the soluble factors found within it. Molecules such as hepatocyte growth factor (HGF), granulocyte and macrophage colony stimulating factors, interleukins (ILs) 6, 7, 8 and 11, tumor necrosis factor-alpha (TNF-alpha), vascular endothelial growth factor (VEGF), brain derived neurotrophic factor (BDNF), nerve growth factor (NGF), adipokines and others have been identified within the ASCs' secretome. Due to its importance regarding future applications for the field of regenerative medicine, we aim, in the present review, to make a comprehensive analysis of the literature relating to the ASCs' secretome and its relevance to the immune and central nervous system, vascularization and cardiac regeneration. The concluding section will highlight some of the major challenges that remain before ASCs can be used for future clinical applications.
Facial deformities require precise reconstruction of the appearance and function of the original tissue. The current standard of care-the use of bone harvested from another region in the body-has ...major limitations, including pain and comorbidities associated with surgery. We have engineered one of the most geometrically complex facial bones by using autologous stromal/stem cells, native bovine bone matrix, and a perfusion bioreactor for the growth and transport of living grafts, without bone morphogenetic proteins. The ramus-condyle unit, the most eminent load-bearing bone in the skull, was reconstructed using an image-guided personalized approach in skeletally mature Yucatán minipigs (human-scale preclinical model). We used clinically approved decellularized bovine trabecular bone as a scaffolding material and crafted it into an anatomically correct shape using image-guided micromilling to fit the defect. Autologous adipose-derived stromal/stem cells were seeded into the scaffold and cultured in perfusion for 3 weeks in a specialized bioreactor to form immature bone tissue. Six months after implantation, the engineered grafts maintained their anatomical structure, integrated with native tissues, and generated greater volume of new bone and greater vascular infiltration than either nonseeded anatomical scaffolds or untreated defects. This translational study demonstrates feasibility of facial bone reconstruction using autologous, anatomically shaped, living grafts formed in vitro, and presents a platform for personalized bone tissue engineering.
A diverse array of environmental factors contributes to the overall control of stem cell activity. In particular, new data continue to mount on the influence of the extracellular matrix (ECM) on stem ...cell fate through physical interactions with cells, such as the control of cell geometry, ECM geometry/topography at the nanoscale, ECM mechanical properties, and the transmission of mechanical or other biophysical factors to the cell. Here, we review some of the physical processes by which cues from the ECM can influence stem cell fate, with particular relevance to the use of stem cells in tissue engineering and regenerative medicine.
The ability to isolate, expand and differentiate adult stem cells into a chondrogenic lineage is an important step in the development of tissue engineering approaches for cartilage repair or ...regeneration for the treatment of joint injury or osteoarthritis, as well as for their application in plastic or reconstructive surgery. Adipose-derived stem cells (ASCs) provide an abundant and easily accessible source of adult stem cells for use in such regenerative approaches. This protocol first describes the isolation of ASCs from liposuction aspirate. The cell culture conditions provided for ASC expansion provide a large number of multipotent stem cells. Instructions for growth factor-based induction of ASCs into chondrocyte-like cells using either cell pellet or alginate bead systems are detailed. These methods are similar to those published for chondrogenesis of bone marrow-derived mesenchymal stem cells but distinct because of the unique nature of ASCs. Investigators can expect consistent differentiation of ASCs, allowing for slight variation as a result of donor and serum lot effects. Approximately 10-12 weeks are needed for the entire process of ASC isolation, including the characterization of chondrocyte-like cells, which is also described.
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