In vivo reprogramming of wound-resident cells generates skin epithelial tissue.

Large cutaneous ulcers are, in severe cases, life threatening1,2. As the global population ages, non-healing ulcers are becoming increasingly common1,2. Treatment currently requires the transplantation of pre-existing epithelial components, such as skin grafts, or therapy using cultured cells2. Here we develop alternative supplies of epidermal coverage for the treatment of these kinds of wounds. We generated expandable epithelial tissues using in vivo reprogramming of wound-resident mesenchymal cells. Transduction of four transcription factors that specify the skin-cell lineage enabled efficient and rapid de novo epithelialization from the surface of cutaneous ulcers in mice. Our findings may provide a new therapeutic avenue for treating skin wounds and could be extended to other disease situations in which tissue homeostasis and repair are impaired.

Enrichment isolation of adipose-derived stem/stromal cells from the liquid portion of liposuction aspirates with the use of an adherent column.

BACKGROUND AIMS: Adipose-derived stem/progenitor cells (ASCs) are typically obtained from the lipoaspirates; however, a smaller number of ASCs can be isolated without enzymatic digestion from the infranatant liposuction aspirate fluid (LAF). We evaluated the effectiveness of an adherent column, currently used to isolate mesenchymal stromal cells from bone marrow, to isolate LAF cells. METHODS: We applied peripheral blood (PB), PB mixed with cultured ASCs (PB-ASC), and LAF solution to the column and divided it into two fractions, the adherent (positive) and the non-adherent (negative) fractions. We compared this method with hypotonic hemolysis (lysis) for the red blood cell count, nucleated cells count and cell compositions as well as functional properties of isolated mesenchymal cells. RESULTS: The column effectively removed red blood cells, though the removal efficiency was slightly inferior to hemolysis. After column processing of PB-ASC, 60.5% of ASCs (53.2% by lysis) were selectively collected in the positive fraction, and the negative fraction contained almost no ASCs. After processing of LAF solution, nucleated cell yields were comparable between the column and hemolysis; however, subsequent adherent culture indicated that a higher average ASC yield was obtained from the column-positive samples than from the lysis samples, suggesting that the column method may be superior to hemolysis for obtaining viable ASCs. Mesenchymal differentiation and network formation assays showed no statistical differences in ASC functions between the lysis and column-positive samples. CONCLUSIONS: Our results suggest that a column with non-woven rayon and polyethylene fabrics is useful for isolating stromal vascular fraction cells from LAF solutions for clinical applications.

Efficacy of serial excisions of melanocytic nevi on the face using a carbon dioxide laser: a cosmetic point of view.

UNLABELLED: In cosmetic clinical practice, many patients express the desire for removal of melanocytic nevi, especially those on the face. The carbon dioxide (CO2) laser currently is the preferred for treatment of such lesions because of less scar formation, less bleeding, and simplicity of the procedure. However, if the diameter of the lesion is greater than 5 mm, many clinicians prefer surgical resection to laser resection because laser resection of relatively large nevi often results in the formation of a conspicuous dimple. The authors developed a serial laser excision method for melanocytic nevi larger than 5 mm, with division of the lesion into multiple segments, which allowed the ablated area to gain optimal granulation and reepithelialization, leading to a satisfactory cosmetic appearance. This novel procedure was performed for 25 patients with melanocytic nevi ranging from 5 to 10 mm in diameter. The lesions were divided into two segments in 21 patients, three segments in 3 patients, and four segments in 1 patient. The divided parts of the lesions were ablated by CO2 laser serially at intervals of 2-4 weeks. All the patients obtained optimal granulation and epithelialization in the treated lesions, and the final appearance was satisfactory. Although the therapeutic period was longer than for a surgical excision or a single laser treatment, the new treatment approach of serial excision by CO2 laser achieved favorable outcomes for the treatment of relatively large nevi 5-10 mm in size. LEVEL OF EVIDENCE V: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Reinnervation of segmented latissimus dorsi muscle with the distal stump of the thoracodorsal nerve: A preliminary experimental study in rats.

To date, nerve stumps have been dissected at the proximal side of the donor muscle for reinnervation of the muscle in free neurovascular muscle transfer. Herein, we examined the use of the distal thoracodorsal nerve, dissected from the muscle belly at the distal side of the latissimus dorsi muscle, for the reinnervation of muscle. The rat right latissimus dorsi muscle was employed as the model for our study. Twenty Wistar rats were used in this study. A rectangular muscle segment was dissected with the distal stump of dominant thoracodorsal nerve. After rotation of muscle, the distal nerve stump was sutured to a severed proximal recipient thoracodorsal nerve (n = 5). The degree of reinnervation through the distal nerve stump was compared with control groups that received proximal-to-proximal nerve sutures (n = 5), nerves that were not severed (n = 5), and severed nerves that were not sutured (n = 5) using electrophysiological, histological, and muscular volume assessments. Reinnervation of the distal nerve stump was confirmed by the contraction of the muscle following electrical stimulation and electromyography. Crossing of axons into motor endplates was confirmed by histology. Results of these assays were similar to that of the proximal nerve suture group. The volume of muscle in the distal nerve suture group was not significant different from that of the proximal nerve suture group (P = 0.63). It was demonstrated that the distal stump of the thoracodorsal nerve can be used to innervate segmented latissimus dorsi muscle. This novel procedure for the reinnervation of transplanted muscle deserves further investigations.

Therapeutic potential of fibroblast growth factor-2 for hypertrophic scars: upregulation of MMP-1 and HGF expression.

Although hypertrophic scars (HTSs) and keloids are challenging problems, their pathogenesis is not well understood, making therapy difficult. We showed that matrix metalloproteinase (MMP)-1 expression was downregulated in HTS compared with normal skin from the same patients, whereas type 1 and 3 collagen and transforming growth factor-ß (TGF-ß) were upregulated. These differences, however, were not seen in cultured fibroblasts, suggesting the involvement of microenvironmental factors in the pathogenesis of HTS. Fibroblast growth factor-2 (FGF-2) highly upregulated the expression of MMP-1 and hepatocyte growth factor (HGF) in both HTS-derived and control fibroblasts; the upregulation was reversed by extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) inhibitors. An animal study using human HTS tissue implanted into nude mice indicated that controlled-release FGF-2 resulted in significantly less weight and decreased hydroxyproline content in HTS. Degradation of collagen fibers in FGF-2-treated HTS was also confirmed histologically. Western blotting showed that FGF-2-treated HTS expressed significantly higher MMP-1 protein than control. Decreased MMP-1 expression may be an important transcriptional change in HTS, and its reversal as well as upregulation of HGF by FGF-2 could be a new therapeutic approach for HTS.

Adipose injury-associated factors mitigate hypoxia in ischemic tissues through activation of adipose-derived stem/progenitor/stromal cells and induction of angiogenesis.

Based on the analysis of exudates from injured adipose tissue, we prepared a mixture containing the injury-associated growth factors at the same proportion as the exudates, named adipose injury cocktail (AIC). We hypothesized that AIC induces a series of regenerating and angiogenic processes without actual wounding. The purpose of this study is to elucidate the therapeutic potentials of AIC. AIC preferentially activated adipose-derived stem/progenitor/stromal cells (ASCs) to proliferate, migrate, and form networks compared with vascular endothelial cells, whereas vascular endothelial growth factor did not induce mitogenesis or chemotaxis in human ASCs. Each component growth factor of AIC was differently responsible for the ASC activation. AIC-treated ASCs tended to differentiate into adipocytes or vessel-constituting cells rather than into other cell types. In ischemic adipose tissues of mice, induced by either a surgical intervention or diabetes, AIC administration enhanced proliferation, especially of CD31(-)/CD34(+) ASCs, and mitigated tissue hypoxia by increasing capillary density and reducing fibrogenesis. These results suggest that AIC may have therapeutic potentials for various ischemic/hypoxic conditions by inducing adipose remodeling and neovascularization through activation of ASCs and other cells. Treatment with AIC has many advantages over cell-based therapies regarding morbidity, cost, and physical risks and may be used as an alternative therapy for improving tissue oxygen.

Purification and characterization of human adipose-resident microvascular endothelial progenitor cells.

Human adipose tissue is a rich source of adipose-derived stem cells (ASCs) and vascular endothelial progenitor cells (EPCs). However, no standardized method has been established for the isolation and purification of adipose-derived EPCs (AEPCs). The aim of this study was to establish a method for the isolation and purification of AEPCs. The stromal vascular fraction (SVF) was extracted from human lipoaspirates, and the CD45-CD31+ fraction of the SVF was collected by magnetic-activated cell sorting (MACS). The CD45-CD31+ fraction was cultured for 4.5 days, followed by a second MACS separation to collect the CD31+ fraction. Purified AEPCs were expanded without being overwhelmed by proliferating ASCs, indicating that a high level (> 95%) of AEPC purification is a key factor for their successful isolation and expansion. AEPCs exhibited typical endothelial markers, including CD31, von Willebrand factor, and the isolectin-B4 binding capacity. AEPCs formed colonies, comparable to cultured human umbilical vein endothelial cells (HUVECs). Both AEPCs and HUVECs formed capillary-like networks in the tube formation assay, with no significant difference in network lengths. We are the first to establish a purification and expansion method to isolate these cells. Because adipose tissue is a clinically accessible and abundant tissue, AEPCs may have potential advantages as a therapeutic tool for regenerative medicine.

A prospective randomized controlled study of oral tranexamic acid for preventing postinflammatory hyperpigmentation after Q-switched ruby laser.

BACKGROUND: Postinflammatory hyperpigmentation (PIH) is the most common skin complication in Asians after invasive cosmetic treatments. OBJECTIVE: To determine whether oral tranexamic acid (TA) reduces the incidence of PIH after Q-switched ruby laser (QSRL) treatment. METHODS AND MATERIALS: Thirty-two Japanese women underwent QSRL treatment for senile lentigines on the face. They were randomly divided into two groups that did (n=15) and did not (n=17) receive oral TA treatment (750 mg/d) for the first 4 weeks after QSRL treatment. Nineteen participants had melasma-like maculae at baseline. Clinical and colorimetric assessments were performed at baseline and 2 and 4 weeks later. RESULTS: Pigmentation was effectively treated using QSRL at 2 weeks, but PIH was frequently seen at 4 weeks. There was no significant difference in the incidence of PIH between participants who received oral TA and those who did not. The presence of melasma did not influence the effectiveness of the treatment. CONCLUSION: Although oral TA has been reported to have depigmentation effects, it may not be effective for preventing PIH after QSRL. Considering the dosage and duration of treatment, an optimal protocol may be needed to induce the efficacy of this treatment to achieve the PIH-preventing effect of oral TA.

IFATS collection: Fibroblast growth factor-2-induced hepatocyte growth factor secretion by adipose-derived stromal cells inhibits postinjury fibrogenesis through a c-Jun N-terminal kinase-dependent mechanism.

Adipose-derived stem/stromal cells (ASCs) not only function as tissue-specific progenitor cells but also are multipotent and secrete angiogenic growth factors, such as hepatocyte growth factor (HGF), under certain circumstances. However, the biological role and regulatory mechanism of this secretion have not been well studied. We focused on the role of ASCs in the process of adipose tissue injury and repair and found that among injury-associated growth factors, fibroblast growth factor-2 (FGF-2) strongly promoted ASC proliferation and HGF secretion through a c-Jun N-terminal kinase (JNK) signaling pathway. In a mouse model of ischemia-reperfusion injury of adipose tissue, regenerative changes following necrotic and apoptotic changes were seen for 2 weeks. Acute release of FGF-2 by injured adipose tissue was followed by upregulation of HGF. During the adipose tissue remodeling process, adipose-derived 5-bromo-2-deoxyuridine-positive cells were shown to be ASCs (CD31-CD34+). Inhibition of JNK signaling inhibited the activation of ASCs and delayed the remodeling process. In addition, inhibition of FGF-2 or JNK signaling prevented postinjury upregulation of HGF and led to increased fibrogenesis in the injured adipose tissue. Increased fibrogenesis also followed the administration of a neutralizing antibody against HGF. FGF-2 released from injured tissue acts through a JNK signaling pathway to stimulate ASCs to proliferate and secrete HGF, contributing to the regeneration of adipose tissue and suppression of fibrogenesis after injury. This study revealed a functional role for ASCs in the response to injury and provides new insight into the therapeutic potential of ASCs.

Progenitor-enriched adipose tissue transplantation as rescue for breast implant complications.

Breast enhancement with artificial implants is one of the most frequently performed cosmetic surgeries but is associated with various complications, such as capsular contracture, that lead to implant removal or replacement at a relatively high rate. For replacement, we used transplantation of progenitor-supplemented adipose tissue (cell-assisted lipotransfer; CAL) in 15 patients. The stromal vascular fraction containing adipose tissue progenitor cells obtained from liposuction aspirates was used to enrich for progenitor cells in the graft. Overall, clinical results were very satisfactory, and no major abnormalities were seen on magnetic resonance imaging or mammogram after 12 months. Postoperative atrophy of injected fat was minimal and did not change substantially after 2 months. Surviving fat volume at 12 months was 155 +/- 50 mL (Right; mean +/- SD) and 143 +/- 80 mL (Left) following lipoinjection from an initial mean of 264 mL. These preliminary results suggest that CAL is a suitable methodology for the replacement of breast implants.

TGF-beta is specifically expressed in human dermal papilla cells and modulates hair folliculogenesis.

Dermal papilla cells (DPCs) in the mammalian hair follicle have been shown to develop hair follicles through epithelial-mesenchymal interactions. A cell therapy to regenerate human hair is theoretically possible by expanding autologous human DPCs (hDPCs) and transplanting them into bald skin, though much remains to be overcome before clinical success. In this study, we compared gene signatures of hDPCs at different passages and human dermal fibroblasts, and found transforming growth factor (TGF)-beta(2) to be highly expressed in cultured hDPCs. Keratinocyte conditioned medium, which is known to help preserve the hair-inducing capacity of hDPCs, up-regulated TGF-beta(2) expression of hDPCs and also enhanced their alkaline phosphatase (ALP) activity, a known index for hair-inductive capacity. Through screening of components secreted from keratinocytes, the vitamin D(3) analogue was found to promote TGF-beta(2) expression and ALP activity of hDPCs. In animal hair folliculogenesis models using rat epidermis and expanded hDPCs, inhibition of TGF-beta(2) signalling at the ligand or receptor level significantly impaired hair folliculogenesis and maturation. These results suggest an important role for TGF-beta(2) in hair follicle morphogenesis and provide insights into the establishment of future cell therapies for hair regrowth by transplanting expanded DPCs.

Differential expression of stem-cell-associated markers in human hair follicle epithelial cells.

Several putative biomarkers have been suggested for identifying murine follicular stem cells; however, human hair follicles have a different pattern of biomarker expression, and follicular stem cell isolation methods have not been established. To isolate a stem cell population applicable to clinical settings, we conducted a comprehensive survey of the expression of stem-cell-associated (K15, CD200, CD34, and CD271) and other biomarkers (K1, K14, CD29, and CD49f) in immunohistological sections of the human epidermis and follicular outer root sheath (ORS). We also examined freshly isolated and cultured epidermal or follicular cells with single- and multicolor flow cytometry or immunocytochemistry. After sorting cells by CD200, CD34, and forward scatter (FSC) values (cell size), colony-forming assays were performed. We found that biomarkers were differentially expressed in the epidermis and ORS. Basal bulge cells were mainly K15+CD200+CD34(-)CD271(-), and suprabasal cells were K15(-)CD200+CD34(-)CD271(-). We categorized follicular cells into nine subpopulations according to biomarker expression profiles. The CD200+CD34(-) bulge cells had much higher colony-forming abilities than the CD34+ population, and were divided into two subpopulations: a CD200+CD34(-)FSC(high) (K15-rich, basal) and a CD200+CD34(-)FSC(low) (K15-poor, suprabasal) population. The former formed fewer but larger-sized colonies than the latter. Follicular epithelial cell cultivation resulted in loss of K15, CD200, CD34, and CD271 expression, but maintenance of K14, CD29, and CD49f expression. We found that the bulge contained two populations with different localizations, cell sizes, and colony-forming abilities. We showed that K15, CD200, CD34, and CD271 were useful biomarkers for characterizing freshly isolated human follicular epithelial cells in diverse stages of differentiation.

Evaluation of animal models for the hair-inducing capacity of cultured human dermal papilla cells.

BACKGROUND: The dermal papilla (DP) interacts with epithelial cells for folliculogenesis. For translational research on cell therapies for hair regrowth with cultured human DP cells (hDPCs), a model to evaluate the capacity of hDPCs to induce hair formation is inevitable. METHODS: Chamber models were constructed by transplanting 4 different combinations of mouse or human epithelial and mesenchymal cells into a silicone chamber implanted onto the back of nude mice. In parallel, 3 types of sandwich constructs were created by inserting hDPCs or human DP tissue between the epidermis and dermis of isolated rat footpad skin or human facial skin, and subcutaneously transplanting the constructs into the back of nude mice. Four to six weeks later, skin sections of each model were histologically examined. RESULTS: Folliculoneogenesis was detected in both chamber and sandwich models, although the induction rate and maturity of the hair follicles varied among cell combination subgroups in each model. The difference in hair induction rate was not statistically significant between 2 representative chamber and sandwich subgroups using cultured hDPCs. The sandwich model, however, required fewer hDPCs, did not require human keratinocytes, and exhibited a higher rate of successful sample collection. CONCLUSIONS: Although there is no significant difference in hair induction rate, the sandwich model using cultured hDPCs and the rat sole skin is more feasible than the chamber model using human cultured keratinocytes and hDPCs as a tool to evaluate the hair-inducing capacity of cultured hDPCs.

Eyelid Thread-lifting Treatment for Periocular Infantile Hemangioma in an Extremely Premature Low-birth-weight Infant.

Periocular infantile hemangioma (IH) sometimes hinders proper eye opening and limits the visual field. Even short-term deprivation of hemilateral vision may cause amblyopia in infants during the visually sensitive period and, thus, necessitates prompt treatment. Propranolol is a commonly known remedy for IH, though it may not act sufficiently to rapidly resolve a vision-threatening critical condition in an infant. Low birth weight is a significant risk factor for IH incidence. Despite the high incidence, evidence-based treatment for IH in preterm infants has not been established. This report describes periocular IH of an extremely premature low-birth-weight infant (body weight: 526 g) whose lesion showed no reduction after oral administration of propranolol for 2 weeks but was successfully treated by topical steroid injection and eyelid thread-lifting management, with no eventual visual impairment.

Cell-assisted lipotransfer for facial lipoatrophy: efficacy of clinical use of adipose-derived stem cells.

BACKGROUND: Lipoinjection is a promising treatment, but its efficacy in recontouring facial lipoatrophy remains to be established. OBJECTIVE: The objective was to evaluate the efficacy and adverse effects of lipoinjection and supplementation of adipose-derived stem/stromal cells (ASCs) to adipose grafts. METHODS: To overcome drawbacks of autologous lipoinjection, we have developed a novel strategy called cell-assisted lipotransfer (CAL). In CAL, stromal vascular fraction containing ASCs was freshly isolated from half of an aspirated fat sample and attached to the other half of aspirated fat sample with the fat acting as a scaffold. This process converts relatively ASC-poor aspirated fat into ASC-rich fat. We performed conventional lipoinjection (non-CAL; n=3) or CAL (n=3) on six patients with facial lipoatrophy due to lupus profundus or Parry-Romberg syndrome. RESULTS: All patients obtained improvement in facial contour, but the CAL group had a better clinical improvement score than did the non-CAL patients, although the difference did not reach statistical significance (p=.11). Adipose necrosis was found in one non-CAL case who took perioperative oral corticosteroids. CONCLUSION: Our results suggest that CAL is both effective and safe and potentially superior to conventional lipoinjection for facial recontouring. The authors have indicated no significant interest with commercial supporters.

Efficient isolation and culture of endothelial cells from venous malformation using the Rho-associated protein kinase inhibitor Y27632.

BACKGROUND: The investigation of primary cells from a pathological lesion can elucidate the pathogenesis of diseases, but, for vascular malformations in humans, such basic research is still stagnant, because the isolation and culture of vascular endothelial cells (ECs) is very difficult. To obtain a sufficient amount of ECs from venous malformation (VM) this study took advantage of a Rho-associated protein kinase inhibitor, Y27632, which had been used for the efficient procurement of primary keratinocytes. METHODS: ECs were isolated and cultured from VM lesions, combining enzymatic digestion, cell sorting, and Y27632. The proliferative effect of Y27632 on ECs was examined by proliferation assay. The characteristics of the ECs cultured with Y27632 by EC marker expression and tube formation assay were also examined. RESULTS: Y27632 enhanced the proliferation of ECs and elongated the senescence of the cells. The expression of specific markers of ECs such as von Willebrand factor, endothelin-1, and VE-cadherin, was confirmed in the cells cultured with Y27632. In a tube formation assay, the cells cultured with Y27632 showed higher tube formation ability compared to the cells cultured without Y27632, indicating that Y27632 promoted the angiogenic capability of ECs. CONCLUSIONS: The protocol using Y27632 offers a new EC culture methodology and provides a new option for the biological investigation of vascular malformations. This new method will contribute to other types of vascular biology research as well.

Differential contributions of graft-derived and host-derived cells in tissue regeneration/remodeling after fat grafting.

BACKGROUND: Recent research indicates that the adipose tissue of nonvascularized grafts is completely remodeled within 3 months, although origins of next-generation cells are unclear. METHODS: Inguinal fat pads of green fluorescent protein mice and wild-type mice were cross-transplanted beneath the scalp. At 1, 2, 4, and 12 weeks after transplantation, grafted fat was harvested, weighed, and analyzed through immunohistochemistry, whole-mount staining, and flow cytometry of cell isolates. Bone marrow of green fluorescent protein mice was transplanted to wild-type mice (after irradiation). Eight weeks later, these mice also received fat grafts, which were analyzed as well. RESULTS: The majority of host-derived cells detected during remodeling of grafted fat were macrophages (>90 percent at the early stage; 60 percent at 12 weeks). Cell origins were analyzed at 12 weeks (i.e., when completely regenerated). At this point, mature adipocytes were largely derived from adipose-derived stem/stromal cells of grafts. Although vascular wall constituents were chiefly graft derived, vascular endothelial cells originated equally from graft and host bone marrow. Adipose-derived stem/stromal cells of regenerated fat were an admixture of grafted, host nonbone marrow, and host bone marrow cells. CONCLUSIONS: The above findings underscore the importance of adipose stem/stromal cells in the grafted fat for adipocyte regeneration. Host bone marrow and local tissues contributed substantially to capillary networks and provided new adipose-derived stem/stromal cells. An appreciation of mechanisms that are operant in this setting stands to improve clinical outcomes of fat grafting and cell-based therapies.

Degeneration, regeneration, and cicatrization after fat grafting: dynamic total tissue remodeling during the first 3 months.

BACKGROUND: Fat grafting is promising, but clinical outcomes are not always predictable. The mechanisms of tissue revascularization/regeneration, and tissue necrosis and subsequent absorption/fibrosis of the graft, are poorly understood. METHODS: An autologous inguinal fat pad was transplanted under the scalp of mice, and detailed cellular events during the first 3 months were investigated with immunohistochemistry. RESULTS: Except for the most superficial surviving zone, death of all adipocytes was confirmed at 1 week. Perilipin-positive small new adipocytes appeared at 1 week and peaked in number at 4 weeks in the regenerating zone (the second zone). In the most central necrotizing zone, adipogenesis did not occur and many inflammatory cells were observed after 2 weeks. CD34+/Ki67+ proliferating adipose stem/progenitor cells were seen at 1 to 4 weeks, but the majority of proliferating cells were MAC2+ monocytes/macrophages. Although CD206+ M1 macrophages surrounded oil droplets for phagocytosis, CD206+ M2 macrophages appeared in areas where adipocyte replacement failed and formed multiple layers for cicatrization of oil drop spaces. Adipogenesis was complete by 12 weeks, but stabilization of nonregenerated areas was still ongoing at that time. Lipid droplets derived from dead adipocytes were absorbed slowly and thus aided adipose remodeling by maintaining the space until adipocyte regeneration. CONCLUSIONS: Dynamic remodeling after fat grafting was confirmed. Adipocyte fate differed, depending on the microenvironment: intact survival, replacement with a new adipocyte, or replacement with cicatrization/oil cyst. This detailed understanding will help refine surgical grafting procedures and postoperative evaluation.

Stromal vascular fraction isolated from lipo-aspirates using an automated processing system: bench and bed analysis.

The heterogeneous stromal vascular fraction (SVF), containing adipose-derived stem/progenitor cells (ASCs), can be easily isolated through enzymatic digestion of aspirated adipose tissue. In clinical settings, however, strict control of technical procedures according to standard operating procedures and validation of cell-processing conditions are required. Therefore, we evaluated the efficiency and reliability of an automated system for SVF isolation from adipose tissue. SVF cells, freshly isolated using the automated procedure, showed comparable number and viability to those from manual isolation. Flow cytometric analysis confirmed an SVF cell composition profile similar to that after manual isolation. In addition, the ASC yield after 1 week in culture was also not significantly different between the two groups. Our clinical study, in which SVF cells isolated with the automated system were transplanted with aspirated fat tissue for soft tissue augmentation/reconstruction in 42 patients, showed satisfactory outcomes with no serious side-effects. Taken together, our results suggested that the automated isolation system is as reliable a method as manual isolation and may also be useful in clinical settings. Automated isolation is expected to enable cell-based clinical trials in small facilities with an aseptic room, without the necessity of a good manufacturing practice-level cell processing area.