Dedifferentiated Fat Cell Research Articles

Muscle regeneration therapy using dedifferentiated fat cell (DFAT) for anal sphincter dysfunction.

We investigated the effects of mouse-derived DFAT on the myogenic differentiation of a mouse-derived myoblast cell line (C2C12) and examined the therapeutic effects of rat-derived DFAT on anal sphincter injury using a rat model. C2C12 cells were cultured using DMEM and DFAT-conditioned medium (DFAT-CM), evaluating MyoD and Myogenin gene expression via RT-PCR. DFAT was locally administered to model rats with anorectal sphincter dysfunction 3days post-CTX injection. Therapeutic effects were assessed through functional assessment, including anal pressure measurement using solid-state manometry pre/post-CTX, and on days 1, 3, 7, 10, 14, 17, and 21 post-DFAT administration. Histological evaluation involved anal canal excision on days 1, 3, 7, 14, and 21 after CTX administration, followed by hematoxylin-eosin staining. C2C12 cells cultured with DFAT-CM exhibited increased MyoD and Myogenin gene expression compared to control. Anal pressure measurements revealed early recovery of resting pressure in the DFAT-treated group. Histologically, DFAT-treated rats demonstrated an increase in mature muscle cells within newly formed muscle fibers on days 14 and 21 after CTX administration, indicating enhanced muscle tissue repair. DFAT demonstrated the potential to enhance histological and functional muscle tissue repair.These findings propose DFAT as a novel therapeutic approach for anorectal sphincter dysfunction treatment.

2α-Substituted Vitamin D Derivatives Effectively Enhance the Osteoblast Differentiation of Dedifferentiated Fat Cells.

The active form of vitamin D3, 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3], is a principal regulator of calcium homeostasis through activation of the vitamin D receptor (VDR). Previous studies have shown that 2α-(3-hydroxypropyl)-1,25D3 (O1C3) and 2α-(3-hydroxypropoxy)-1,25D3 (O2C3), vitamin D derivatives resistant to inactivation enzymes, can activate VDR, induce leukemic cell differentiation, and increase blood calcium levels in rats more effectively than 1,25(OH)2D3. In this study, to further investigate the usefulness of 2α-substituted vitamin D derivatives, we examined the effects of O2C3, O1C3, and their derivatives on VDR activity in cells and mouse tissues and on osteoblast differentiation of dedifferentiated fat (DFAT) cells, a cell type with potential therapeutic application in regenerative medicine. In cell culture experiments using kidney-derived HEK293 cells, intestinal mucosa-derived CaCO2 cells, and osteoblast-derived MG63 cells, and in mouse experiments, O2C2, O2C3, O1C3, and O1C4 had a weaker effect than or equivalent effect to 1,25(OH)2D3 in VDR transactivation and induction of the VDR target gene CYP24A1, but they enhanced osteoblast differentiation in DFAT cells equally to or more effectively than 1,25(OH)2D3. In long-term treatment with the compound without the medium change (7 days), the derivatives enhanced osteoblast differentiation more effectively than 1,25(OH)2D3. O2C3 and O1C3 were more stable than 1,25(OH)2D3 in DFAT cell culture. These results indicate that 2α-substituted vitamin D derivatives, such as inactivation-resistant O2C3 and O1C3, are more effective than 1,25(OH)2D3 in osteoblast differentiation of DFAT cells, suggesting potential roles in regenerative medicine with DFAT cells and other multipotent cells.

Comparative study of dedifferentiated fat cell and adipose-derived stromal cell sheets for periodontal tissue regeneration: In vivo and in vitro evidence.

To compare the efficacy of adipocyte-derived dedifferentiated fat (DFAT) cell and adipose-derived stromal cell (ADSC) sheets for regenerative treatment of intra-bony periodontal defects. DFAT cells were obtained using the ceiling culture method and were compared with ADSCs using Cell Counting Kit-8, colony formation assay, surface antigen identification, and multilineage differentiation assays. DFAT and ADSC sheets were prepared in a cell sheet culture medium. The biological characteristics of DFAT cell and ADSC sheets were compared using haematoxylin and eosin staining, quantitative reverse transcription polymerase chain reaction, and immunofluorescence staining. Micro-computed tomography and histological staining were used to compare the effects of the two cell sheets on the repair of periodontal intra-bony defects in rats. DFAT cells and ADSCs demonstrated mesenchymal stem cell characteristics. Both cell types were CD29-, CD90-, and CD146-positive and CD31-, CD34-, and CD45-negative. DFAT cells and ADSCs exhibited similar osteogenic and adipogenic differentiation capabilities and colony formation ability. DFAT cells displayed stronger proliferation capabilities compared with ADSCs. Compared with the ADSC sheets, DFAT cell sheets exhibited a higher expression of periodontal-related genes and proteins and greater ability to regenerate periodontal tissue. Our findings suggest that DFAT cell sheets are an ideal seed cell source and form of cell delivery for periodontal intra-bony defects.

Implantation of dedifferentiated fat cells ameliorated antineutrophil cytoplasmic antibody glomerulonephritis by immunosuppression and increases in tumor necrosis factor-stimulated gene-6

IntroductionThe implantation of dedifferentiated fat (DFAT) cells has been shown to exert immunosuppressive effects. To develop DFAT cell therapy for antineutrophil cytoplasmic antibody (ANCA) glomerulonephritis, the effects of the implantation of DFAT cells on ANCA glomerulonephritis were investigated in mice.MethodsPKH26-labeled DFAT cells (105) were infused through the posterior orbital venous plexus to investigate delivery of DFAT cells in ICR mice. DFAT cells (105) were also implanted in SCG mice as a model for ANCA glomerulonephritis. Expression of tumor necrosis factor-stimulated gene-6 (TSG-6) mRNA and protein in kidney was evaluated, and the expression of microRNAs associated with TSG-6 in plasma, lung and kidney was analyzed. Expressions of CD44, prostaglandin (PG) E2, interleukin (IL)-10, IL-1β, tumor necrosis factor (TNF)-α mRNAs, C–C motif chemokine ligand 17 (CCL-17) and monocyte chemoattractant protein (MCP)-1 proteins were measured in kidney from SCG mice implanted with DFAT cells.ResultsAfter their intravenous infusion, almost all DFAT cells were trapped in the lung and not delivered into the kidney. Implantation of DFAT cells in SCG mice suppressed glomerular crescent formation, decreased urinary protein excretions and increased expression of TSG-6 mRNA, protein and immunostaining in kidney from these mice. Increased expression of microRNA 23b-3p in plasma, kidney and lung; decreased expression of CD44 mRNA; and increased expression of PGE2 and IL-10 mRNAs were also observed in kidney from these mice. Implantation of DFAT cells also decreased the expression of TNF-α and MCP-1 proteins and increased that of CCL-17 protein in kidney from the SCG mice. Survival rates were higher in SCG mice implanted with DFAT cells than in SCG mice without implantation.ConclusionMechanisms underlying the effects of improvement of ANCA glomerulonephritis are associated with immunosuppressive effects by TSG-6 and the transition of M1–M2 macrophages, suggesting that implantation of DFAT cells may become a cell therapy for ANCA glomerulonephritis.

OR26. Dedifferentiated Fat Cells versus Adipose Derived Mesenchymal Stem Cells Experimental Study about Adipose-derived Stem Cells for Cardiomyocyte Precursor using Traditional and Insert Culture

Abstract Background and Aims The discovery of dedifferentiated fat cell (DFAT) raises the question, are AMSC and DFAT the same type of mesenchymal stem cell (MSC)? We aim to analyze the immunocytochemistry (ICC) profile of both cells and the correlation between different methods with different passage to GATA-4 and cTnT expression in vitro. Methods and Results Human subcutaneous-fat tissue was isolated and divided into, first group, using single filtration by 100 µm cell strainer and second group using double filtration with addition of 70 µm strainer after the 100 µm. Cell obtained from both groups were labeled AMSC and DFAT respectively. Prior to passaging, CD34, CD45, CD90, and CD105 were analyze by ICC. After passaging, passage 1 and 4 was exposed to Gibco PSC Cardiomyocyte Differentiation Kit for 21 days. Colonies from days 7, -14, and -21 were analyze by ICC for GATA-4 and cTnT expression. Post hoc analysis shown that there is no significance difference in MSC markers expression between groups. There were significance differences in GATA-4 expression on all observation days, which decreased significantly from the 14th to the 21st day. Statistic significancy also found on cTnT expression, with a significant increase from day 14 to day 21. The filter variable consistently has a significant effect on the quantity of GATA-4 and cTnT expressions. Conclusion The study shows that different culture methods, presumably the use of filters, will have a significant effect on the differentiation of AMSC and DFAT into cardiomyocytes.

Enhancement of jaw bone regeneration via ERK1/2 activation using dedifferentiated fat cells

Background aimsMesenchymal stem/stromal cells (MSCs) are multipotent and self-renewing cells that are extensively used in tissue engineering. Adipose tissues are known to be the source of two types of MSCs; namely, adipose tissue–derived MSCs (ASCs) and dedifferentiated fat (DFAT) cells. Although ASCs are sometimes transplanted for clinical cytotherapy, the effects of DFAT cell transplantation on mandibular bone healing remain unclear. MethodsThe authors assessed whether DFAT cells have osteogenerative potential compared with ASCs in rats in vitro. In addition, to elucidate the ability of DFAT cells to regenerate the jaw bone, the authors examined the effects of DFAT cells on new bone formation in a mandibular defect model in (i) 30-week-old rats and (ii) ovariectomy-induced osteoporotic rats in vivo. ResultsOsteoblast differentiation with bone morphogenetic protein 2 (BMP-2) or osteogenesis-induced medium upregulated the osteogenesis-related molecules in DFAT cells compared with those in ASCs. BMP-2 activated the phosphorylation signaling pathways of ERK1/2 and Smad2 in DFAT cells, but minor Smad1/5/9 activation was noted in ASCs. The transplantation of DFAT cells into normal or ovariectomy-induced osteoporotic rats with mandibular defects promoted new bone formation compared with that seen with ASCs. ConclusionsDFAT cells promoted osteoblast differentiation and new bone formation through ERK1/2 and Smad2 signaling pathways in vitro. The transplantation of DFAT cells promoted new mandibular bone formation in vivo compared with that seen with ASCs. These results suggest that transplantation of ERK1/2-activated DFAT cells shorten the mandibular bone healing process in cytotherapy.

Promoting Effect of Dedifferentiated Fat Cell Transplantation on Bone Formation in a Mouse Femoral Fracture Model

Promoting Effect of Dedifferentiated Fat Cell Transplantation on Bone Formation in a Mouse Femoral Fracture Model

Therapeutic potential of mature adipocyte-derived dedifferentiated fat cells for inflammatory bowel disease

PurposeOur previous studies demonstrated that mature adipocyte-derived dedifferentiated fat (DFAT) cells possess similar multipotency as mesenchymal stem cells. Here, we examined the immunoregulatory potential of DFAT cells in vitro and the therapeutic effect of DFAT cell transplantation in a mouse inflammatory bowel disease (IBD) model.MethodsThe effect of DFAT cell co-culture on T cell proliferation and expression of immunosuppression-related genes in DFAT cells were evaluated. To create IBD, CD4+CD45RBhigh T cells were intraperitoneally injected into SCID mice. One week later, DFAT cells (1 × 105, DFAT group) or saline (Control group) were intraperitoneally injected. Subsequently bodyweight was measured every week and IBD clinical and histological scores were evaluated at 5 weeks after T cell administration.ResultsThe T cell proliferation was inhibited by co-cultured DFAT cells in a cell density-dependent manner. Gene expression of TRAIL, IDO1, and NOS2 in DFAT cells was upregulated by TNFα stimulation. DFAT group improved IBD-associated weight loss, IBD clinical and histological scores compared to Control group.ConclusionDFAT cells possess immunoregulatory potential and the cell transplantation promoted recovery from colon damage and improved clinical symptoms in the IBD model. DFAT cells could play an important role in the treatment of IBD.

The Effect of Autologous Dedifferentiated Fat Cell Transplantation in a Pig Model of Hindlimb Ischemia

The Effect of Autologous Dedifferentiated Fat Cell Transplantation in a Pig Model of Hindlimb Ischemia

Transplantation of dedifferentiated fat cell-derived micromass pellets contributed to cartilage repair in the rat osteochondral defect model

Mature adipocyte-derived dedifferentiated fat (DFAT) cells possesses the ability to proliferate effectively and the potential to differentiate into multiple linages of mesenchymal tissue; similar to adipose-derived stem cells (ASCs). The purpose of this study is to examine the effects of DFAT cell transplantation on cartilage repair in a rat model of osteochondral defects. Full-thickness osteochondral defects were created in the knees of Sprague-Dawley rats bilaterally. Cartilage-like micromass pellets were prepared from green fluorescent protein (GFP)-labeled rat DFAT cells and subsequently transplanted into the affected right knee of these rats. Defects in the left knee were used as a control. Macroscopic and microscopic changes of treated and control defects were evaluated up to 12 weeks post-treatment with DFAT cells. To observe the transplanted cells, sectioned femurs were immunostained for GFP and type II collagen. DFAT cells formed micromass pellets expressing characteristics of immature cartilage invitro. In the DFAT cell-transplanted limbs, the defects were completely filled with white micromass pellets as early as 2 weeks post-treatment. These limbs became smooth at 4 weeks. Conversely, the defects in the control limbs were still not repaired by 4 weeks. Macroscopic ICRS scores at 2 and 4 weeks were significantly higher in the DFAT cells-transplanted limbs compared to those of the control limbs. The modified O'Driscol histological scores for the DFAT cell-transplanted limbs were significantly higher than those of the control limbs at corresponding time points. GFP-positive DAFT cells were detected in the transplanted area at 2 weeks but hardly visible at 12 weeks post-operation. Transplantation of DFAT cell-derived micromass pellets contribute to cartilage repair in a rat osteochondral defect model. DFAT cell transplantation may be a viable therapeutic strategy for the repair of osteochondral injuries.

Transplantation of dedifferentiation fat cells promotes intervertebral disc regeneration in a rat intervertebral disc degeneration model

Our group has reported that mature adipocyte-derived dedifferentiated fat (DFAT) cells show multilineage differentiation potential similar to that observed in mesenchymal stem cells. In the present study, we examined whether DFAT cell transplantation could contribute to intervertebral disc regeneration using a rat intervertebral disc degeneration (IDD) model. The IDD was created in Sprague-Dawley rats by puncturing at level of caudal intervertebral disc under fluoroscopy. One week after injury, rat DFAT cells (5 × 104, DFAT group, n = 13) or phosphate-buffered saline (PBS, control group, n = 13) were injected into the intervertebral disc. Percent disc height index (%DHI) was measured every week and histology of injured disc was evaluated at 8 weeks after transplantation. Radiographic analysis revealed that the %DHI in the DFAT group significantly higher than that in the control group at 2–3 weeks after transplantation. Histological analysis revealed that ectopic formation of nucleus pulposus (NP)-like tissue at the outer layer of annulus fibrosus was frequently observed in the DFAT group but not in the control group. Transplantation experiments using green fluorescent protein (GFP)-labeled DFAT cells revealed that the ectopic NP-like tissue was positive for GFP, suggesting direct differentiation of DFAT cells into NP-like cells. In conclusion, DFAT cell transplantation promoted the regeneration of intervertebral disc and improved intervertebral disc height in the rat IDD model. Because adipose tissue is abundant and easily accessible, DFAT cell transplantation may be an attractive therapeutic strategy against IDD.

Dedifferentiated Fat Cells as a Novel Source for Cell Therapy to Target Neonatal Hypoxic-Ischemic Encephalopathy

Neonatal hypoxic-ischemic (HI) encephalopathy (HIE) remains a major cause of mortality and persistent neurological disabilities in affected individuals. At present, hypothermia is considered to be the only applicable treatment option, although growing evidence suggests that cell-based therapy might achieve better outcomes. Dedifferentiated fat (DFAT) cells are derived from mature adipocytes via a dedifferentiation strategy called ceiling culture. Their abundance and ready availability might make them an ideal therapeutic tool for the treatment of HIE. In the present study, we aimed to determine whether the outcome of HIE can be improved by DFAT cell treatment. HI injury was achieved by ligating the left common carotid artery in 7-day-old rat pups, followed by 1-h exposure to 8% O₂. Subsequently, the severity of damage was assessed by diffusion-weighted magnetic resonance imaging to assign animals to equivalent groups. 24 h after hypoxia, DFAT cells were injected at 10⁵ cells/pup into the right external jugular vein. To evaluate brain damage in the acute phase, a group of animals was sacrificed 48 h after the insult, and paraffin sections of the brain were stained to assess several acute injury markers. In the chronic phase, the behavioral outcome was measured by performing a series of behavioral tests. From the 24th day of age, the sensorimotor function was examined by evaluating the initial forepaw placement on a cylinder wall and the latency to falling from a rotarod treadmill. The cognitive function was tested with the novel object recognition (NOR) test. In vitro conditioned medium (CM) prepared from cultured DFAT cells was added at various concentrations to neuronal cell cultures, which were then exposed to oxygen-glucose deprivation (OGD). The number of cells that stained positive for the apoptosis marker active caspase-3 decreased by 73 and 52% in the hippocampus and temporal cortex areas of the brain, respectively, in the DFAT-treated pups. Similarly, the numbers of ED-1-positive cells (activated microglia) decreased by 66 and 44%, respectively, in the same areas in the DFAT-treated group. The number of cells positive for the oxidative stress marker 4-hydroxyl-2-nonenal decreased by 68 and 50% in the hippocampus and the parietal cortex areas, respectively, in the DFAT-treated group. The HI insult led to a motor deficit according to the rotarod treadmill and cylinder test, where it significantly affected the vehicle group, whereas no difference was confirmed between the DFAT and sham groups. However, the NOR test indicated no significant differences between any of the groups. DFAT treatment did not reduce the infarct volume, which was confirmed immunohistochemically. According to in vitro experiments, the cell death rates in the DFAT-CM-treated cells were significantly lower than those in the controls when DFAT-CM was added 48 h prior to OGD. The treatment effect of adding DFAT-CM 24 h prior to OGD was also significant. Our results indicate that intravenous injection with DFAT cells is effective for ameliorating HI brain injury, possibly via paracrine effects.

マウス急性移植片対宿主病モデルに対する脱分化脂肪細胞移植の治療効果

移植片対宿主病 (graft versus host disease: GVHD) は造血幹細胞移植 (hematopoietic cell transplantation: HCT) の致死的合併症の一つとして知られている．近年ステロイド抵抗性急性GVHD に対し間葉系幹細胞 (mesenchymal stem cell: MSC) を用いた細胞治療の臨床応用が始まっている． 脱分化脂肪細胞 (dedifferentiated fat cell: DFAT) にはMSC と同様の免疫調節作用があることが確認されており，DFAT をGVHD の治療に応用できる可能性がある．本検討ではマウス急性GVHD モデルを作製し，尾静脈からDFAT を投与し治療効果を検討した．DFAT を投与すると生存率が改善傾向を示し，体重減少が抑制され，臨床スコアが改善した．本検討により，DFAT が急性GVHD の細胞治療のソースとなりうることを示した．

Transplantation of mature adipocyte-derived dedifferentiated fat cells for the treatment of vesicoureteral reflux in a rat model.

Autologous cells potentially provide an ideal injectable substance for management in vesicoureteral reflux (VUR). The aim of this study is to examine the effects of mature adipocyte-derived dedifferentiated fat (DFAT) cell transplantation on VUR in a rat bladder pressurization-induced VUR model. To create VUR, Sprague-Dawley rats underwent urethral clamping and placement of cystostomy followed by intravesical pressurization. Rat DFAT cells (1×106 cells, DFAT group, n=5) or saline (control group, n=5) was then injected into the bilateral vesicoureteral junctions. Two weeks later, VUR grade was evaluated on cystography. The number of apoptotic cells in the renal pelvic urothelium, the ureteral inner/outer diameter ratio and the area of connective tissue in the posterior bladder wall were measured. The reflux grade in the DFAT group was significantly lower than that in the control group. The number of apoptotic cells in the renal pelvic urothelium, ureteral inner/outer diameter ratio and connective tissue area in DFAT group were significantly lower in comparison with the control group. DFAT cell transplantation improved VUR and exerted nephroprotective effects in a rat VUR model.

Smooth muscle-like tissue constructs with circumferentially oriented cells formed by the cell fiber technology.

The proper functioning of many organs and tissues containing smooth muscles greatly depends on the intricate organization of the smooth muscle cells oriented in appropriate directions. Consequently controlling the cellular orientation in three-dimensional (3D) cellular constructs is an important issue in engineering tissues of smooth muscles. However, the ability to precisely control the cellular orientation at the microscale cannot be achieved by various commonly used 3D tissue engineering building blocks such as spheroids. This paper presents the formation of coiled spring-shaped 3D cellular constructs containing circumferentially oriented smooth muscle-like cells differentiated from dedifferentiated fat (DFAT) cells. By using the cell fiber technology, DFAT cells suspended in a mixture of extracellular proteins possessing an optimized stiffness were encapsulated in the core region of alginate shell microfibers and uniformly aligned to the longitudinal direction. Upon differentiation induction to the smooth muscle lineage, DFAT cell fibers self-assembled to coiled spring structures where the cells became circumferentially oriented. By changing the initial core-shell microfiber diameter, we demonstrated that the spring pitch and diameter could be controlled. 21 days after differentiation induction, the cell fibers contained high percentages of ASMA-positive and calponin-positive cells. Our technology to create these smooth muscle-like spring constructs enabled precise control of cellular alignment and orientation in 3D. These constructs can further serve as tissue engineering building blocks for larger organs and cellular implants used in clinical treatments.

Dedifferentiated fat cells: A cell source for regenerative medicine.

The identification of an ideal cell source for tissue regeneration remains a challenge in the stem cell field. The ability of progeny cells to differentiate into other cell types is important for the processes of tissue reconstruction and tissue engineering and has clinical, biochemical or molecular implications. The adaptation of stem cells from adipose tissue for use in regenerative medicine has created a new role for adipocytes. Mature adipocytes can easily be isolated from adipose cell suspensions and allowed to dedifferentiate into lipid-free multipotent cells, referred to as dedifferentiated fat (DFAT) cells. Compared to other adult stem cells, the DFAT cells have unique advantages in their abundance, ease of isolation and homogeneity. Under proper condition in vitro and in vivo, the DFAT cells have exhibited adipogenic, osteogenic, chondrogenic, cardiomyogenc, angiogenic, myogenic, and neurogenic potentials. In this review, we first discuss the phenomena of dedifferentiation and transdifferentiation of cells, and then dedifferentiation of adipocytes in particular. Understanding the dedifferentiation process itself may contribute to our knowledge of normal growth processes, as well as mechanisms of disease. Second, we highlight new developments in DFAT cell culture and summarize the current understanding of DFAT cell properties. The unique features of DFAT cells are promising for clinical applications such as tissue regeneration.

脱分化脂肪細胞(DFAT) における血管新生効果の検討

治療的血管新生において，脱分化脂肪細胞(dedifferentiated fat cell: DFAT) が血管内皮細胞にどのような作用を与えるかについては十分に解明されていない． 本研究では，green fluorescent protein (GFP) 標識DFAT またはGFP 標識脂肪組織由来幹細胞(adiposederived stem cell: ASC) を血管内皮細胞と共培養することにより，細胞間相互作用による血管内皮細胞の増殖能，管腔形成能の変化を比較検討した．さらに蛍光免疫染色やリアルタイムRT-PCR を用いて，DFAT が血管構成細胞であるペリサイトへの分化能について，ASC と比較して検討した．その結果，DFAT とASC はともに，血管内皮細胞の細胞増殖能，管腔形成能を促進することが明らかになった．また血管内皮細胞との共培養により，DFAT はペリサイト特異的マーカーであるNG2, PDGFR-β の発現が増強し，ASC と比較してペリサイトへの分化能が高い可能性が示唆された．これらの結果よりDFAT は治療的血管新生における有用な治療用細胞ソースとなりうることが示された．

The osteoblastic differentiation ability of human dedifferentiated fat cells is higher than that of adipose stem cells from the buccal fat pad

ObjectivesThe purpose of this study was to evaluate and compare the osteoblastic differentiation ability of dedifferentiated fat (DFAT) cells and adipose stem cells (ASCs) from the buccal fat pad (BFP).Materials and methodsWe isolated human DFAT cells and ASCs from the BFP of a patient who underwent oral and maxillofacial surgery and then analyzed their cell surface antigens by flow cytometry. Then, the cells were cultured in osteogenic medium for 14 days. Measurement of bone-specific alkaline phosphatase (BAP), osteocalcin (OCN), and calcium deposition and alizarin red staining were performed to evaluate the osteoblastic differentiation ability of both cell types.ResultsASCs and DFAT cells were positive for CD90 and CD105 and negative for CD11b, CD34, and CD45. BAP (days 3 and 7), OCN (day 14), and calcium deposition (days 7 and 14) within DFAT cell cultures were significantly higher than those in ASC cultures. The alizarin red-stained area in DFAT cell cultures, which indicates mineralized matrix deposition, was stained more strongly than that in ASC cultures.ConclusionsThe cell surface antigens of ASCs and DFAT cells tend to be similar. Furthermore, the osteoblastic differentiation ability of human DFAT cells is higher than that of ASCs from the BFP.Clinical relevanceIsolation of DFAT cells from the BFP has an esthetic advantage because the BFP can be obtained via the oral cavity without injury to the external body surface. Therefore, we consider that DFAT cells from the BFP are an ideal cell source for bone tissue engineering.

Current status of drug therapies for osteoporosis and the search for stem cells adapted for bone regenerative medicine

A number of factors can lead to bone disorders such as osteoporosis, in which the balance of bone resorption vs. bone formation is upset (i.e., more bone is resorbed than is formed). The result is a loss of bone mass, with a concomitant decrease in bone density. Drugs for osteoporosis can be broadly classified as "bone resorption inhibitors", which impede bone resorption by osteoclasts, and "bone formation accelerators", which augment bone formation by osteoblasts. Here, we describe representative drugs in each class, i.e., the bisphosphonates and the parathyroid hormone. In addition, we introduce two novel bone formation accelerators, SST-VEDI and SSH-BMI, which are currently under investigation by our research group. On the other hand, regenerative therapy, characterized by (ideally) the use of a patient's own cells to regenerate lost tissue, is now a matter of global interest. At present, candidate cell sources for regenerative therapy include embryonic stem cells (created from embryos based on the fertilization of oocytes), induced pluripotent stem cells (created artificially by using somatic cells as the starting material), and somatic stem cells (found in the tissues of the adult body). This review summarizes the identifying features and the therapeutic potential of each of these stem cell types for bone regenerative medicine. Although a number of different kinds of somatic stem cells have been reported, we turn our attention toward two that are of particular interest for prospective applications in bone repair: the dedifferentiated fat cell, and the deciduous dental pulp-derived stem cell.

Osteogenic Effects of Dedifferentiated Fat Cell Transplantation in Rabbit Models of Bone Defect and Ovariectomy-Induced Osteoporosis

We have previously reported that mature adipocyte-derived dedifferentiated fat (DFAT) cells have a high proliferative activity and the potential to differentiate into lineages of mesenchymal tissue similar to bone marrow mesenchymal stem cells (MSCs). In the present study, we examined the effects of autologous DFAT cell transplantation on bone regeneration in a rabbit bone defect model and an ovariectomy (OVX)-induced osteoporosis model. The formation of tissue-engineered bone (TEB) was observed when rabbit DFAT cells were loaded onto a β-tricalcium phosphate (TCP)/collagen sponge and cultured in an osteogenic differentiation medium for 3 weeks. Autologous implantation of DFAT cell-mediated TEB constructs promoted bone regeneration in a rabbit tibial defect model. Regenerated bone tissue induced by transplantation of DFAT cell-mediated TEB constructs was histologically well differentiated and exhibited higher bone strength in a three-point bending test compared to that induced by the β-TCP/collagen sponge alone. In OVX-induced osteoporosis model rabbits, DFAT cells were obtained with the osteogenic activity similar to cells from healthy rabbits. Intrabone marrow injection of autologous DFAT cells significantly increased the bone mineral density (BMD) at the injected site in the OVX rabbits. Transplanted DFAT cells remained mainly on the injection side of the bone marrow by at least 28 days after intrabone marrow injection and a part of them expressed osteocalcin. In conclusion, these results demonstrate that autologous implantation of DFAT cells contributed to bone regeneration in a rabbit bone defect model and an OVX-induced osteoporosis model. DFAT cells may be an attractive cell source for cell-based bone tissue engineering to treat nonunion fractures in all patients, including those with osteoporosis.