Effects of conditioned medium obtained from human adipose-derived stem cells on skin inflammation.

Introduction: Cell therapy using adipose-derived mesenchymal stem cells (ASCs) is a promising avenue of regenerative medicine for the treatment of various diseases. It has been considered that ASCs exert their therapeutic effects through the secretion of multiple factors that are critical for tissue remodeling or the suppression of inflammation. Recently, conditioned medium (CM) from ASCs that contains a complex of secreted factors has received attention as a cost-effective alternative to cell therapy. Methods: We investigated the effects of CM obtained from ASCs (ASCs-CM) using human dermal fibroblasts (hDFs) and human epidermal keratinocytes with or without interleukin (IL)-1ß and examined mRNA levels of marker genes. We also examined alterations in cell proliferation and morphology of hDFs following treatment with ASCs-CM. We further investigated the effects of ASCs-CM treatment on prevention of skin inflammation using a mouse model. Results: In hDFs and human epidermal keratinocytes, the ASCs-CM treatment suppressed pro-inflammatory factors and enhanced regenerative and remodeling factors with or without interleukin (IL)-1ß exposure. The ASCs-CM treatment also enhanced cell proliferation of hDFs and prevented morphological changes in response to IL-1ß exposure. Furthermore, in a mouse model of skin inflammation, treatment with ASCs-CM reduced the inflammatory reactions, including redness and thickness. Conclusions: CM from ASCs may represent a potential alternative to ASC therapy for the treatment of inflammatory skin conditions.

Development of hydroxyapatite-coated nonwovens for efficient isolation of somatic stem cells from adipose tissues.

Adipose-derived stem cells (ASCs) are an attractive cell source for cell therapy. Despite the increasing number of clinical applications, the methodology for ASC isolation is not optimized for every individual. In this study, we developed an effective material to stabilize explant cultures from small-fragment adipose tissues. Methods: Polypropylene/polyethylene nonwoven sheets were coated with hydroxyapatite (HA) particles. Adipose fragments were then placed on these sheets, and their ability to trap tissue was monitored during explant culture. The yield and properties of the cells were compared to those of cells isolated by conventional collagenase digestion. Results: Hydroxyapatite-coated nonwovens immediately trapped adipose fragments when placed on the sheets. The adhesion was stable even in culture media, leading to cell migration and proliferation from the tissue along with the nonwoven fibers. A higher fiber density further enhanced cell growth. Although cells on nonwoven explants could not be fully collected with cell dissociation enzymes, the cell yield was significantly higher than that of conventional monolayer culture without impacting stem cell properties. Conclusions: Hydroxyapatite-coated nonwovens are useful for the effective primary explant culture of connective tissues without enzymatic cell dissociation.

Divergence in chondrogenic potential between in vitro and in vivo of adipose- and synovial-stem cells from mouse and human.

BACKGROUND: Somatic stem cell transplantation has been performed for cartilage injury, but the reparative mechanisms are still conflicting. The chondrogenic potential of stem cells are thought as promising features for cartilage therapy; however, the correlation between their potential for chondrogenesis in vitro and in vivo remains undefined. The purpose of this study was to investigate the intrinsic chondrogenic condition depends on cell types and explore an indicator to select useful stem cells for cartilage regeneration. METHODS: The chondrogenic potential of two different stem cell types derived from adipose tissue (ASCs) and synovium (SSCs) of mice and humans was assessed using bone morphogenic protein-2 (BMP2) and transforming growth factor-ß1 (TGFß1). Their in vivo chondrogenic potential was validated through transplantation into a mouse osteochondral defect model. RESULTS: All cell types showed apparent chondrogenesis under the combination of BMP2 and TGFß1 in vitro, as assessed by the formation of proteoglycan- and type 2 collagen (COL2)-rich tissues. However, our results vastly differed with those observed following single stimulation among species and cell types; apparent chondrogenesis of mouse SSCs was observed with supplementation of BMP2 or TGFß1, whereas chondrogenesis of mouse ASCs and human SSCs was observed with supplementation of BMP2 not TGFß1. Human ASCs showed no obvious chondrogenesis following single stimulation. Mouse SSCs showed the formation of hyaline-like cartilage which had less fibrous components (COL1/3) with supplementation of TGFß1. However, human cells developed COL1/3+ tissues with all treatments. Transcriptomic analysis for TGFß receptors and ligands of cells prior to chondrogenic induction did not indicate their distinct reactivity to the TGFß1 or BMP2. In the transplanted site in vivo, mouse SSCs formed hyaline-like cartilage (proteoglycan+/COL2+/COL1-/COL3-) but other cell types mainly formed COL1/3-positive fibrous tissues in line with in vitro reactivity to TGFß1. CONCLUSION: Optimal chondrogenic factors driving chondrogenesis from somatic stem cells are intrinsically distinct among cell types and species. Among them, the response to TGFß1 may possibly represent the fate of stem cells when locally transplanted into cartilage defects.

Associations of clinical outcomes and MRI findings in intra-articular administration of autologous adipose-derived stem cells for knee osteoarthritis.

INTRODUCTION: Clinical studies of intra-articular injection of mesenchymal stem cells for osteoarthritis (OA) indicate its efficacy. Here, we retrospectively investigated the associations of pretherapeutic magnetic resonance imaging (MRI) findings with the clinical outcomes up to 6 months, after intra-articular administration of adipose-derived stem cells (ASCs) to knee OA patients. METHODS: We first analyzed alterations of the visual analog scale (VAS) and knee injury and osteoarthritis outcome score (KOOS) in 57 knees of 34 patients from whom clinical scores were obtained before ASC therapy, and at 1, 3, and 6 months. Among the patients, we further examined MRI findings of 34 knees of 19 patients whose pretherapeutic MRI data were available. RESULTS: The mean improvement of VAS and KOOS-total during 6 months was 2.6 ± 4.0 (from 6.1 ± 2.5 to 3.5 ± 2.9, P < 0.001) and 10.2 ± 12.4 (from 54.4 ± 12.7 to 64.6 ± 13.8, P < 0.01), respectively. Scales related to pain and symptoms improved earlier than those related to activities of daily living (ADL) and sports/recreation. Improvement of VAS and KOOS-sports/recreation was significantly higher in patients with more severe cartilage lesions. Similarly, osteophyte lesions were associated significantly with improvement of VAS and KOOS-ADL, and BML was associated with KOOS-ADL and KOOS-sports/recreation. CONCLUSIONS: In intra-articular administration of autologous ASCs for knee OA, improvement of VAS and KOOS-sports/recreation was significantly higher in patients with more severe cartilage lesions. Similarly, osteophyte lesions were associated significantly with improvement of VAS and KOOS-ADL, and BML was associated with KOOS-ADL and KOOS-sports/recreation. Clinical studies with larger numbers of patients and various kinds of data are necessary to predict therapeutic effects.

Triple coverage of ischial ulcers with adipofascial turnover and fasciocutaneous flaps.

Despite a wide variety of flap options, ischial ulcers remain the most difficult pressure ulcers to treat. This article describes the authors' successful surgical procedure for coverage of ischial ulcers using adipofascial turnover flaps combined with a local fasciocutaneous flap. After debridement, the adipofascial flaps are harvested both cephalad and caudal to the defect. The flaps are then turned over to cover the exposed bone in a manner so as to overlap the two flaps. A local fasciocutaneous flap (Limberg flap) is applied to the raw surface of the turnover flaps. Twenty-two patients with ischial ulcers were treated using this surgical procedure. Overall, 86.4 percent of the flaps (19 of 22) healed primarily. Triple coverage with the combination of double adipofascial turnover flaps and a local fasciocutaneous flap allows for an easily performed and minimally invasive procedure, preservation of future flap options, and a soft-tissue supply sufficient for covering the prominence and bony prominence and filling dead space. This technique provides successful soft-tissue reconstruction for minor to moderate-size ischial pressure ulcers.

Distal perforator-based fasciocutaneous V-Y flap for treatment of sacral pressure ulcers.

Although the gluteal V-Y advancement flap has been recognized as the most reliable method for management of sacral pressure ulcers, its limited mobility has been a challenging problem. The authors present a new modification of the V-Y advancement flap to overcome the problem. After débridement, a large triangle is designed to create a V-Yadvancement flap on the unilateral buttock and the medial half is elevated as a fasciocutaneous flap, preserving the distal perforators in the muscular attachment. Then an arc-shaped incision is made in the gluteus maximus muscle along with the lateral edge of the triangular flap. The split muscle is elevated at a depth above the deeper fascia until sufficient advancement of the flap is obtained. This full-thickness elevation of the gluteus maximus muscle from the distal (lateral) side avoids the impairment of perforators or their mother vessels and achieves great advancement. Thirty-one patients with sacral pressure defects larger than 8 cm in diameter were treated using this surgical procedure. Overall, 93.5 percent of the flaps (29 of 31) healed primarily. The largest defect that was closed with a unilateral flap was 16 cm in diameter. The present technique accomplishes remarkable excursion of the unilateral V-Y fasciocutaneous flap, with high flap reliability and preservation of the contralateral buttock as well as gluteus maximus muscle function.

Surgical correction for curly toe using open tenotomy of flexor digitorum brevis tendon.

Curly toe is a common congenital deformity characterized by flexion and varus deformity of the interphalangeal joints. Because this minor deformity is seldom accompanied with any symptoms, treatment strategy has rarely been discussed in detail in the literature. Eight toes in seven patients with curly toe were treated by open tenotomy of the medial slip of the flexor digitorum brevis tendon. If sufficient correction was not obtained, the collateral ligament and the volar plate of the proximal interphalangeal joint were dissected. The skin defect at the plantar base of the toe was covered using a local flap or a full-thickness skin graft. The median age at operation was 2 years 6 months (ranged from 8 months to 5 years 4 months). In all cases, contracture of the plantar skin at the base of the toe and tight FDB tendon were recognized to a variable degree. Postoperatively, overlapping of the affected toe was corrected in every case at a median follow-up of 2 years 9 months. However, flexion and/or varus deformity tended to remain to some degree in those patients with severe curly toe. Toes with moderate to severe deformity with overlapping beneath the adjacent toe are candidates for surgical correction, because spontaneous correction is unlikely and troublesome symptoms may occur as the child grows older. Surgical correction should be performed until 2-3 years of age. The postoperative result might be poor, if treated in the older age, because skeletal deformity is likely to occur. Open tenotomy of the FDB tendon is easy to perform, and toe function was seldom impaired.

Analysis of ischemia-reperfusion injury in a microcirculatory model of pressure ulcers.

The aim of this study was to establish a pressure ulcer model that visualizes the microcirculation, and to examine the participation of ischemia-reperfusion injury in the pathophysiology of pressure ulcers. An original system composed of a new skin fold chamber and compression device allowed loading quantitative vertical stress to the skin. An intravital microscopic technique enabled direct visualization of the microcirculation in the physiological condition and in response to pressure application. To estimate the effect of ischemia-reperfusion injury, animals were divided into two groups: the compression-release group (n = 8), in which the animals received four cycles of compression-release which consisted of 2 hours of compression followed by 1 hour of pressure release; and the compression alone group (n = 8) in which the animals underwent continuous compression for 8 hours. Functional capillary density was quantified before the compression procedure and on day 1 (35 hours) after the first evaluation. The cyclic compression-release procedure significantly decreased functional capillary density as compared to continuous compression, indicating that in our experimental setting repetition of ischemia-reperfusion cycle more severely damaged the microcirculation than single prolonged ischemic insult. This finding supports the significant contribution of ischemia-reperfusion injury to the pathophysiology of pressure ulcers at the level of dynamic in vivo microcirculation.