Chondroitin Sulfate Promotes the Proliferation of Keloid Fibroblasts Through Activation of the Integrin and Protein Kinase B Pathways.

Keloids are dermal fibroproliferative tumors that arise beyond the boundary of the original wound edges and invades adjacent tissue. Keloids are characterized by the extensive production of extracellular matrix (ECM) and abnormal fibroblast proliferation. Chondroitin sulfate (CS) is one of the major structural components of cartilage and ECM. Recently, we reported the over-accumulation of CS in keloid lesions. Keloid-derived fibroblasts (KFs) and normal dermal fibroblasts (NFs) were incubated with CS. The fibroblast proliferation rate was analyzed using a tetrazolium salt colorimetric assay. The activation of the intracellular signaling pathway was analyzed by Western blotting. Wortmannin, a PI3K inhibitor, and anti-integrin antibodies were tested to investigate the mechanism of the CS-induced cell proliferation. CS strongly stimulated the proliferation of KFs, but not NFs. The analysis of the intracellular signal transduction pathway revealed that the stimulation effect of CS on KF proliferation was due to the activation of the protein kinase B (AKT) pathway and that integrin α1 was responsible for this phenomenon. We revealed that CS probably activates the AKT pathway through integrin to induce KF proliferation. CS may be a novel clinical therapeutic target in keloids.

HtrA1 Is Specifically Up-Regulated in Active Keloid Lesions and Stimulates Keloid Development.

Keloids occur after failure of the wound healing process; inflammation persists, and various treatments are ineffective. Keloid pathogenesis is still unclear. We have previously analysed the gene expression profiles in keloid tissue and found that HtrA1 was markedly up-regulated in the keloid lesions. HtrA1 is a serine protease suggested to play a role in the pathogenesis of various diseases, including age-related macular degeneration and osteoarthritis, by modulating extracellular matrix or cell surface proteins. We analysed HtrA1 localization and its role in keloid pathogenesis. Thirty keloid patients and twelve unrelated patients were enrolled for in situ hybridization, immunohistochemical, western blot, and cell proliferation analyses. Fibroblast-like cells expressed more HtrA1 in active keloid lesions than in surrounding lesions. The proportion of HtrA1-positive cells in keloids was significantly higher than that in normal skin, and HtrA1 protein was up-regulated relative to normal skin. Silencing HtrA1 gene expression significantly suppressed cell proliferation. HtrA1 was highly expressed in keloid tissues, and the suppression of the HtrA1 gene inhibited the proliferation of keloid-derived fibroblasts. HtrA1 may promote keloid development by accelerating cell proliferation and remodelling keloid-specific extracellular matrix or cell surface molecules. HtrA1 is suggested to have an important role in keloid pathogenesis.

ß-Adrenergic Receptor Blockers Reduce the Occurrence of Keloids and Hypertrophic Scars after Cardiac Device Implantation: A Single-Institution Case-Control Study.

BACKGROUND: Keloids and hypertrophic scars are characterized by excessive proliferation of fibroblasts; abnormal accumulation of extracellular matrix; and clinical findings of raised, red, itchy, and painful lesions. There are few sufficient interventions for keloids, and the development of new therapeutic agents is urgently needed. Several studies suggest that a therapeutic possibility is ß-adrenergic receptor blocker treatment. METHODS: In this single-center case-control study, patients who had undergone cardiac device implantation 7 to 23 months earlier were identified. The implantation incision scars of the patients were deemed to be normal or abnormal depending on their redness. The cases (abnormal scars) and controls (normal scars) were compared in terms of their ß-blocker use rates. RESULTS: Of the 45 eligible patients, 12 and 33 patients were cases and controls, respectively. The cases tended to be less likely to have taken blockers than the controls (25 percent versus 45.5 percent). This difference became significant when the patients whose scars were diagnosed 7 or 8 months after implantation were excluded from the analysis: the age-adjusted odds ratios of the patients who were diagnosed 8 to 23 and 9 to 23 months after implantation were 0.10 (95 percent CI, 0.00 to 0.83; p = 0.0309) and 0.11 (95 percent CI, 0.00 to 0.98; p = 0.047), respectively. CONCLUSIONS: ß-Blockers may be an effective alternative modality for preventing and treating keloids and hypertrophic scars. Large-scale multicenter prospective studies that use histology to diagnose scars and diagnose the postoperative scars at the most suitable period are needed to confirm the effectiveness of blockers for abnormal scars. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, III.

Regeneration of elastic fibers by three-dimensional culture on a collagen scaffold and the addition of latent TGF-ß binding protein 4 to improve elastic matrix deposition.

The objective of this study was to investigate the effects of latent TGF-ß binding protein 4 (LTBP-4) on elastic fiber regeneration in three-dimensional cultures of human dermal fibroblasts (HDFs). Appropriate collagen scaffold for elastic fiber regeneration was also examined. Collagen sponges cross-linked at 120 °C and composed of small pores (25 µm on average) was favorable for elastic fiber regeneration by HDFs. Addition of LTBP-4, followed by culture for 21 days, accelerated elastic fiber accumulation within the scaffolds. Conditioned scaffolds containing either HDFs or LTBP-4-built mature elastic fibers were implanted between the dermis and the cutaneous muscle of mice. The combined use of HDFs and LTBP-4 resulted in thicker tissues containing elastic fibers. These results indicate that weakly cross-linked collagen sponges can be used as scaffolds for regenerating elastic fibers both in vitro and in vivo, and that the addition of LTBP-4 accelerates the deposition of both elastin and fibrillin-1, and increases cell proliferation. These techniques may be useful for generating cutaneous or cardiovascular tissue equivalents; furthermore, they may serve as a useful method for the three-dimensional analyses of drugs used to treat skin diseases or to examine the microstructure of elastin networks.

The Shear Wave Velocity on Elastography Correlates with the Clinical Symptoms and Histopathological Features of Keloids.

BACKGROUND: Keloids present as red, painful lesions causing serious functional and cosmetic problems; however, there is no consensus regarding tools for objectively evaluating keloids. To demonstrate the utility of shear wave elastography in keloids, we investigated the correlations between clinical symptoms, ultrasound shear wave velocity, and histopathological findings. METHODS: Three patients with keloids containing both red hypertrophic and mature areas were evaluated using the shear wave velocity and histopathological findings. RESULTS: The results indicate that the shear wave velocity is high in active hypertrophic areas and low in mature areas. The areas with high elastography values exhibited numerous fibrillar collagenous matrices forming a whorled pattern with hyalinized tissue on hematoxylin-eosin staining corresponding with metachromasia on toluidine blue staining. In the mature area, the collagen fibers were oriented parallel to each other without metachromasia. CONCLUSIONS: Shear wave elastography provides quantitative estimates of tissue stiffness that correlate with the clinical symptoms and histopathological findings of the keloid lesions and can be used to assess the activity of keloids.

A toe keloid after syndactyly release treated with surgical excision and intralesional steroid injection.

SUMMARY: A keloid is a benign fibroproliferative disease of unknown etiology. Although it is common among Asians, the development of keloid on the foot is rare. We experienced a case of a keloid which arose on the foot of a 4-year-old boy after the surgical release of syndactyly. He had congenital cutaneous syndactyly of the third and fourth toes. After the reconstructive operation was performed when the patient was 2 years old, the wound became hypertrophic and grew to 37 × 37 × 8 mm. After the diagnosis of keloid based on a pathological examination, the keloid was resected completely. The web was reconstructed with a planter rectangular flap, and the skin defects were covered with a full-thickness skin graft. After the operation, we administered 5 intralesional steroid injections. Finally, the keloid was diminished 2 years after the operation.

A retrospective analysis on the proper size of tissue expanders to treat scalp lesions.

BACKGROUND: Tissue expanders have become established instruments for scalp reconstruction. However, selection of the size of the expander has not been investigated systematically, and it generally depends on the experience of the surgeon. METHODS: We retrospectively analyzed 21 patients who had undergone treatment for scalp lesions using tissue expanders without any complications and measured 2 variables: the volume of the expanders per area of the excised lesions and the hypothetical stretched functional skin width relative to the width of the excised lesions. We also sought to evaluate the relationship between these 2 variables and the need for revision surgery during the postoperative course. RESULTS: The need for revision surgery was statistically higher in patients with a volume of 7 ml/cm(2) lesion or less and width of functional skin of less than 2.5 cm/cm lesion (P < 0.05). For scar repairs, the required size and volume of the expanders tended to be larger than those required for any other lesions. CONCLUSIONS: Expanders that generate functional skin at least more than 2.5 times the width of the lesion and have a volume more than 7 ml/cm(2) lesion are necessary to cover scalp lesions without complications.

Immediate evaluation of neovascularization in a grafted bilayered artificial dermis using laser Doppler imaging.

BACKGROUND: A bilayered artificial dermis is widely applied for skin defects. Its collagen sponge is biodegraded and replaced with dermis-like tissue after application. There is no reliable method for quantitatively evaluating the blood flow of artificial dermis. In this study, we used laser Doppler imaging to evaluate the perfusion of artificial dermis. MATERIALS AND METHODS: Twelve patients treated with artificial dermis and secondary skin grafting were included. We measured the perfusion unit just after application of artificial dermis, 1 week after, and before skin grafting. RESULTS: Secondary skin grafts of 6 patients took completely, and the others showed partial necrosis. Laser Doppler imaging could detect blood flow in the artificial dermis, and a significant difference was observed in perfusion units between the "complete take" group and "partial necrosis" group before grafting (P < 0.05). CONCLUSIONS: Laser Doppler imaging could be a useful and noninvasive technique for the evaluation of blood flow to the artificial dermis before grafting.

Severe Acute Radiodermatitis in a Keloid Patient with Takayasu's Arteritis.

SUMMARY: Although combination therapy for keloid including postoperative radiation therapy (RT) is common, the radiation toxicity of RT in a patient with a history of collagen vascular disease has not been fully recognized. We experienced a case of an acute radiodermatitis in a patient with keloid. This patient had a chest keloid because of the bypass surgery for Takayasu's arteritis. After we performed an excision and postoperative RT, severe radiodermatitis occurred. We speculate that the higher single dose and the use of electron beams may be related to the onset of severe acute radiodermatitis in this case. It should be kept in mind that there is a risk of exacerbation of radiation toxicity in patients with collagen vascular disease.

Novel collagen/gelatin scaffold with sustained release of basic fibroblast growth factor: clinical trial for chronic skin ulcers.

Chronic skin ulcers such as diabetic ulcers and venous leg ulcers are increasing and are a costly problem in healthcare. We have developed a novel artificial dermis, collagen/gelatin sponge (CGS), which is capable of sustained release of basic fibroblast growth factor (bFGF) for more than 10 days. The objective of this study was to investigate the safety and efficacy of CGS impregnated with bFGF in the treatment of chronic skin ulcers. Patients with chronic skin ulcers that had not healed in at least 4 weeks were treated with CGS impregnated with bFGF at 7 or 14 µg/cm(2) after debridement, and the wound bed improvement was assessed 14 days after application. Wound bed improvement was defined as a granulated and epithelialized area on day 14 with a proportion to the baseline wound area after debridement of 50% or higher. The wound area, the wound area on day 14, and the granulation area on day 14 were independently measured by blinded reviewers in a central review using digital images of wounds taken with a calibrator. Patients were followed up until 28 days after application to observe the adverse reactions related to the application of CGS. From May 2010 to June 2011, 17 patients were enrolled and, in 16 patients, the wound bed improved. Among the randomized patients in step 2, no significant difference was seen between the low-dose group and the high-dose group. No serious adverse reactions were observed. Adverse reactions with a clear causal relationship to the study treatment were mild and patients quickly recovered from them. This study is the first-in-man clinical trial of CGS and showed the safety and efficacy of CGS impregnated with bFGF in the treatment of chronic skin ulcers. This combination therapy could be a promising therapy for chronic skin ulcers.

Multipotent stem cells are effectively collected from adult human cheek skin.

Skin-derived precursor (SKP) cells are a valuable resource for tissue engineering and regenerative medicine, because they represent multipotent stem cells that differentiate into neural and mesodermal progenies. Previous studies suggest that the stem cell pool decreases with age. Here, we show that human multipotent SKP cells can be efficiently collected from adult cheek/chin skin, even in aged individuals of 70-78years. SKP cells were isolated from 38 skin samples by serum-free sphere culture and examined for the ability to differentiate into neural and mesodermal lineages. The number of spheres obtained from adult facial skin was significantly higher than that of trunk or extremity skin. SKP cells derived from cheek/chin skin exhibited a high ability to differentiate into neural and mesodermal cells relative to those derived from eyelid, trunk, or extremity skin. Furthermore, cheek/chin skin SKP cells were shown to express markers for undifferentiated stem cells, including a high expression level of the Sox9 gene. These results indicate that cheek/chin skin is useful for the recovery of multipotent stem cells for tissue engineering and regenerative therapy.