Morikuni Tobita

Concise Review: Adipose‐Derived Stem Cells as a Novel Tool for Future Regenerative Medicine

The potential use of stem cell‐based therapies for the repair and regeneration of various tissues and organs offers a paradigm shift that may provide alternative therapeutic solutions for a number of diseases. The use of either embryonic stem cells (ESCs) or induced pluripotent stem cells in clinical situations is limited due to cell regulations and to technical and ethical considerations involved in the genetic manipulation of human ESCs, even though these cells are, theoretically, highly beneficial. Mesenchymal stem cells seem to be an ideal population of stem cells for practical regenerative medicine, because they are not subjected to the same restrictions. In particular, large number of adipose‐derived stem cells (ASCs) can be easily harvested from adipose tissue. Furthermore, recent basic research and preclinical studies have revealed that the use of ASCs in regenerative medicine is not limited to mesodermal tissue but extends to both ectodermal and endodermal tissues and organs, although ASCs originate from mesodermal lineages. Based on this background knowledge, the primary purpose of this concise review is to summarize and describe the underlying biology of ASCs and their proliferation and differentiation capacities, together with current preclinical and clinical data from a variety of medical fields regarding the use of ASCs in regenerative medicine. In addition, future directions for ASCs in terms of cell‐based therapies and regenerative medicine are discussed. STEM CELLS 2012;30:804–810

New Japanese Initiatives on Stem Cell Therapies

Two laws aiming to provide a new legal framework to promote regenerative medicine, while ensuring the efficacy and safety of the treatments, came into effect in Japan on November 25, 2014. The scope of these laws is briefly described here.

The Effect of Adipose-Derived Stem Cells on Ischemia-Reperfusion Injury: Immunohistochemical and Ultrastructural Evaluation

Background: Advances in the treatment of reperfusion injury have created an opportunity for plastic surgeons to apply these treatments to flaps and implanted tissues. The authors examined the direct and indirect effects of adipose-derived stem cells on ischemia-reperfusion injury on a skin flap model to determine the in vivo differentiation of adipose-derived stem cells to endothelial cells; the levels of vascular endothelial growth factor (VEGF), transforming growth factor-&bgr;, and fibroblast growth factor; and the ultrastructural changes apparent with scanning electron microscopy to clarify the initial events and the following cascades. Methods: Two identical cranial based random flaps with a dimension of 1 × 5 cm were elevated on the dorsums of 20 ICR mice. The left flap was designated as the control and the right flap was injected with adipose-derived stem cells. The flaps were then subjected to 6 hours of ischemia by clamping the pedicle, and then reperfusion. Results: The mean viable flap length in the control and experimental groups was 15.2 ± 3.4 mm and 24.4 ± 2.9 mm, respectively. The mean viable flap area in the control and experimental groups was 12.9 ± 4.1 mm2 and 21.8 ± 3.7 mm2, respectively. The in vivo differentiation of adipose-derived stem cells to endothelial cells was observed. The immunohistochemical stainings, VEGF, transforming growth factor-&bgr;, and fibroblast growth factor revealed increased levels in the experimental groups. Scanning electron microscopy indicated mild injury in the experimental group. Conclusions: The adipose-derived stem cells could prevent ischemia-reperfusion injury, mainly by regulating the growth factors. Although VEGF was the foremost inhibitor of injury, the overall cascade was enhanced by adipose-derived stem cells, with the help of the other growth factors.

Adipose tissue-derived mesenchymal stem cells and platelet-rich plasma: stem cell transplantation methods that enhance stemness.

Because of their ease of isolation and relative abundance, adipose-derived mesenchymal stem cells (ASCs) are a particularly attractive autologous cell source for various therapeutic purposes. ASCs retain a high proliferation capacity in vitro and have the ability to undergo extensive differentiation into multiple cell lineages. Moreover, ASCs secrete a wide range of growth factors that can stimulate tissue regeneration. Therefore, the clinical use of ASCs is feasible. However, the potential of ASCs differs depending on the donor’s medical condition, including diseases such as diabetes. Recent studies demonstrated that ASCs from diabetic donors exhibit reduced proliferative potential and a smaller proportion of stem cell marker-positive cells. Therefore, to ensure the success of regenerative medicine, tissue engineering methods must be improved by the incorporation of factors that increase the proliferation and differentiation of stem/progenitor cells when autologous cells are used. Platelet-rich plasma (PRP), which contains high levels of diverse growth factors that can stimulate stem cell proliferation and cell differentiation in the context of tissue regeneration, has recently been identified as a biological material that could be applied to tissue regeneration. Thus, co-transplantation of ASCs and PRP represents a promising novel approach for cell therapy in regenerative medicine. In this review, we describe the potential benefits of adding PRP to ASCs and preclinical and clinical studies of this approach in various medical fields. We also discuss the mechanisms of PRP action and future cell-based therapies using co-transplantation of ASCs and PRP.

Periodontal tissue regeneration by combined implantation of adipose tissue-derived stem cells and platelet-rich plasma in a canine model

BACKGROUND AIMS One goal of periodontal therapy is to regenerate periodontal tissues. Stem cells, growth factors and scaffolds and biomaterials are vital for the restoration of the architecture and function of complex tissues. Adipose tissue-derived stem cells (ASCs) are an ideal population of stem cells for practical regenerative medicine. In addition, platelet-rich plasma (PRP) can be useful for its ability to stimulate tissue regeneration. PRP contains various growth factors and may be useful as a cell carrier in stem cell therapies. The purpose of this study was to determine whether a mixture of ASCs and PRP promoted periodontal tissue regeneration in a canine model. METHODS Autologous ASCs and PRP were implanted into areas with periodontal tissue defects. Periodontal tissue defects that received PRP alone or non-implantation were also examined. Histologic, immunohistologic and x-ray studies were performed 1 or 2 months after implantation. The amount of newly formed bone and the scale of newly formed cementum in the region of the periodontal tissue defect were analyzed on tissue sections. RESULTS The areas of newly formed bone and cementum were greater 2 months after implantation of ASCs and PRP than at 1 month after implantation, and the radiopacity in the region of the periodontal tissue defect increased markedly by 2 months after implantation. The ASCs and PRP group exhibited periodontal tissue with the correct architecture, including alveolar bone, cementum-like structures and periodontal ligament-like structures, by 2 months after implantation. CONCLUSIONS These findings suggest that a combination of autologous ASCs and PRP promotes periodontal tissue regeneration that develops the appropriate architecture for this complex tissue.

Periodontal disease and periodontal tissue regeneration.

Periodontal disease leads to destruction of the periodontium such as alveolar bone, cementum, the periodontal ligament, and gingiva. Effective treatment for periodontal tissue regeneration is important, because periodontal disease is related to several systemic diseases. However, various conventional therapies for periodontal tissue regeneration have shown limited and variable clinical outcomes. Thus, there are ongoing efforts to identify an alternative cell source, such as stem cells, for the development of new tissue engineering therapies. In this review, periodontal disease and the application of tissue engineering for periodontal tissue regeneration are discussed. In particular, adipose-derived stem cells are presented as an agent for restoring periodontal tissue defects.

Direct and indirect effects of a combination of adipose-derived stem cells and platelet-rich plasma on bone regeneration.

A key goal for successful bone regeneration is to bridge a bone defect using healing procedures that are stable and durable. Adipose-derived stem cells (ASCs) have the potential to differentiate into bone. Meanwhile, platelet-rich plasma (PRP) is an interesting biological means to repair tissue by inducing chemotactic, proliferative, and anabolic cellular responses. This study evaluated bone regeneration using a combination of ASCs and PRP in a rat calvarial defect model. ASCs were isolated from inguinal fat pads of F344 inbred rats, while PRP was prepared from these rats. ASCs were cultured in control medium supplemented with 10% fetal bovine serum or 5% PRP in vitro. After 1 week, levels of growth factors including insulin-like growth factor-1, transforming growth factor-β1, hepatocyte growth factor, and vascular endothelial growth factor in the culture supernatant were measured by enzyme-linked immunosorbent assays. Moreover, the ASC/PRP admixture was transplanted into the rat calvarial defect. Microcomputed tomography, histological, and immunohistochemical (osteopontin and osteocalcin) analyses were performed at 4 and 8 weeks after transplantation. The in vitro study showed that the levels of growth factors secreted by ASCs were significantly increased by the addition of PRP. Transplantation of the ASC/PRP admixture had dramatic effects on bone regeneration overtime in comparison with rats that received other transplants. Furthermore, some ASCs directly differentiated into osteogenic cells in vivo. These findings suggest that the combination of ASCs and PRP has augmentative effects on bone regeneration. The ASC/PRP admixture may be a promising source for the clinical treatment of cranial defects.

Adipose-derived stem cells and periodontal tissue engineering.

Innovative developments in the multidisciplinary field of tissue engineering have yielded various implementation strategies and the possibility of functional tissue regeneration. Technologic advances in the combination of stem cells, biomaterials, and growth factors have created unique opportunities to fabricate tissues in vivo and in vitro. The therapeutic potential of human multipotent mesenchymal stem cells (MSCs), which are harvested from bone marrow and adipose tissue, has generated increasing interest in a wide variety of biomedical disciplines. These cells can differentiate into a variety of tissue types, including bone, cartilage, fat, and nerve tissue. Adipose-derived stem cells have some advantages compared with other sources of stem cells, most notably that a large number of cells can be easily and quickly isolated from adipose tissue. In current clinical therapy for periodontal tissue regeneration, several methods have been developed and applied either alone or in combination, such as enamel matrix proteins, guided tissue regeneration, autologous/allogeneic/xenogeneic bone grafts, and growth factors. However, there are various limitations and shortcomings for periodontal tissue regeneration using current methods. Recently, periodontal tissue regeneration using MSCs has been examined in some animal models. This method has potential in the regeneration of functional periodontal tissues because the various secreted growth factors from MSCs might not only promote the regeneration of periodontal tissue but also encourage neovascularization of the damaged tissues. Adipose-derived stem cells are especially effective for neovascularization compared with other MSC sources. In this review, the possibility and potential of adipose-derived stem cells for regenerative medicine are introduced. Of particular interest, periodontal tissue regeneration with adipose-derived stem cells is discussed.

Prefabrication of tissue engineered bone grafts: an experimental study.

The purpose of this study was to determine whether angiogenesis could successfully be induced into bone tissue that was engineered by cultured adipose-derived stem cells with porous beta-tricalcium phosphate and whether its biologic properties could be maintained by flap prefabrication technique.Adipose-derived stem cells with porous beta-tricalcium phosphate were implanted into the superficial inferior epigastric artery flap of the Fisher rats. After prefabrication for 8 weeks, the prefabricated flaps were elevated and the pedicles were clamped for 4 hours. The samples were harvested after 2 weeks for analyses.Angiogenesis was significantly increased in the prefabricated groups (P < 0.05). There was no significant difference between the prefabricated and nonprefabricated groups in terms of the osteogenic capacity (P > 0.5).The promising results obtained with prefabrication in tissue engineered bone grafts encourage the clinical application of this technology. Thus, prefabrication may be a useful technique in any engineered bone tissue transfer.

Adipose-Derived Stem Cells Improve Collagenase-Induced Tendinopathy in a Rat Model

Background: Tendinopathy is a common and highly prevalent musculoskeletal disorder characterized by repetitive activity-related pain and focal tendon tenderness. Histopathologically, tendinopathic tissue mainly shows degenerative changes. Therefore, tendinopathy is not affected by anti-inflammatory therapies. A novel approach, including a stem cell–based therapy, may be beneficial for its treatment. Purpose/Hypothesis: The purpose of this study was to evaluate the effects of adipose-derived stem cells (ASCs) on tendon healing in a rat tendinopathy model. The hypothesis was that ASC transplantation would improve degeneration in collagenase-induced tendinopathy. Study Design: Controlled laboratory study. Methods: Sixteen F344/NSlc rats underwent collagenase injection into the Achilles tendon to induce tendinopathy. At 1 week after collagenase injection, 8 rats received ASCs (ASC group) and 8 received phosphate-buffered saline alone (PBS group). Animals were sacrificed at 4 or 12 weeks after ASC administration, and the degree of degeneration in each tendon was histologically evaluated according to the Bonar scale. The microstructure of healing tendons was observed by scanning electron microscopy. Reverse-transcription polymerase chain reaction (RT-PCR) was performed to measure the ratio of type III collagen messenger RNA (mRNA) to type I collagen mRNA in tendons. Results: The median Bonar scale score in the ASC and PBS groups was 2.5 and 5.33 at 4 weeks after treatment and 1.0 and 4.0 at 12 weeks after treatment, respectively. Histologically, the ASC group showed a significantly lower degree of tendon degeneration than the PBS group at both time points. In the RT-PCR analysis, the ratio of type III collagen to type I collagen was significantly lower in the ASC group than in the PBS group at 12 weeks after treatment. Moreover, this ratio decreased over time in the ASC group, whereas it increased over time in the PBS group. Conclusion: The study findings demonstrate that the application of ASCs results in significant improvement in the pathological findings associated with tendinopathy and the normalization of collagen ratios within the affected tendon. Clinical Relevance: Subcutaneous adipose tissue can be harvested easily, and ASC administration might have the potential to rapidly treat tendinopathy.