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Chua A.W.C.,Skin Bank Unit | Chua A.W.C.,National University of Singapore | Ma D.,Reconstructive and Aesthetic Surgery | Gan S.U.,National University of Singapore | And 5 more authors.
Journal of Investigative Dermatology | Year: 2011

Keloids are found only in humans and the underlying biochemical mechanisms of their pathogenesis remain unknown. R-spondins (Rspos) are a relatively new group of secreted proteins known to be Wnt/Β-catenin signaling agonists, but their role in keloids has yet to be elucidated. We investigated the expression levels of R-spondin2 (Rspo2) in cell lysates and conditioned media of monocultures and co-cultures of fibroblasts and keratinocytes derived from keloids and normal skin. In this study we found increased protein expression and secretion of Rspo2 in respective monocultures of keloid fibroblasts and keratinocytes when compared with their normal counterparts. Double-chamber co-culture experiments implicated the role of keloid keratinocytes (KKs) in the induction of Rspo2 secretion from fibroblasts because of epithelial-mesenchymal interactions. Addition of recombinant human Rspo2 in culture increased the proliferation of keratinocytes and it acted synergistically with Wnt3a through the canonical Wnt/Β-catenin pathway. Overexpression of Rspo2 in normal fibroblasts brought about thicker epidermis when compared with control fibroblasts in a skin organotypic culture model. This observation coincides with the hyperproliferative phenotype of thickened epidermis seen in keloids. Taken together, the results suggest the possible double paracrine action of KKs in inducing higher expression of Rspo2 in fibroblasts that promotes keratinocyte proliferation and epidermal thickening. © 2011 The Society for Investigative Dermatology. Source

Wong M.,Reconstructive and Aesthetic Surgery | Chua A.,Skin Bank Unit | Tan B.-K.,Reconstructive and Aesthetic Surgery
European Journal of Plastic Surgery | Year: 2013

Systemically unwell patients with extensive wounds present a reconstructive challenge. Conventional skin grafting results in surgical morbidity and donor wounds that may not heal. We propose that cultured skin, in the form of cultured epithelial autografts, be used for wound coverage in these patients. We present two patients with chronic large wounds over the abdomen and the leg. Keratinocytes obtained from a skin biopsy were cultured on a fibrin mat and the cultured epithelial autograft (CEA)-fibrin construct was delivered onto the wound bed 3 weeks later. Initial graft take was 85 and 75 % respectively, and epithelization was complete by 3 weeks. Stable, good quality coverage was maintained at 1.5 years follow-up. We conclude that in a selected group of patients, CEA is a useful modality for the reconstruction of extensive defects with minimum donor site morbidity. The fibrin mat is an optimal system that supports the growth of keratinocytes and allows easy delivery to the wound bed. Level of Evidence: Level IV, therapeutic study. © 2012 Springer-Verlag Berlin Heidelberg. Source

Chua A.W.C.,Skin Bank Unit | Chua A.W.C.,National University of Singapore | Gan S.U.,National University of Singapore | Ting Y.,Skin Bank Unit | And 5 more authors.
Journal of Dermatological Science | Year: 2011

Background: Current evidence suggests the potential role of Wnt signalling in keloids pathogenesis but such literature remains scanty. We hypothesize that Wnt signalling is upregulated in keloid fibroblasts (KFs) and this promotes cellular growth, migration and extracellular matrix (ECM) production in such fibroblasts. Objectives: To verify the downregulation of secreted frizzled-related protein 1 (SFRP1), a Wnt inhibitor and test KFs sensitivity to Wnt3a treatment compared to NFs in terms of activation of Wnt/β-catenin, cellular growth, migration and ECM expressions. Next, to investigate if ectopic expression of SFRP1 and treatment of quercetin in KFs can reverse their phenotypes. Methods: Quantitative Real-time PCR and western blotting were used to verify SFRP1 expression in NFs and KFs. The fibroblasts were tested with Wnt3a conditioned media and its effects were tested for (1) the cells' sensitivity to direct Wnt signalling via the activation of TCF reporter assay and protein expression of β-catenin, (2) cellular growth, (3) cell migration and (4) expressions of ECM components. Finally KFs were stably transduced with SFRP1 and treated with 2 doses of quercetin. Results: Lower levels of SFRP1 were confirmed at mRNA and protein levels in KFs which partly explained their sensitivity to Wnt3a treatment in terms of higher Wnt activation, cellular growth and fibronectin expression. Interestingly, Wnt3a did not promote higher cell migration rate and increase in collagen I expression. Ectopic expression of SFRP1 and quercetin treatment was able to mitigate Wnt3a-mediated phenotype of KFs. Conclusions: Using SFRP1 or inhibitors of Wnt signalling might be one of the therapeutic solutions to treat keloid scarring. © 2011 Japanese Society for Investigative Dermatology. Source

Ma D.,Reconstructive and Aesthetic Surgery | Kua J.E.H.,Reconstructive and Aesthetic Surgery | Lim W.K.,Reconstructive and Aesthetic Surgery | Lim W.K.,Skin Bank Unit | And 3 more authors.
Cytotherapy | Year: 2015

Background aims: Little is published on the characterization and therapeutic potential of human mesenchymal cells derived from hair follicle (HF) dermal sheath (DS). In this study, we isolated and characterized HF DS-mesenchymal stromal cells (DS-MSCs) with respect to the bone marrow mesenchymal stromal cells (BM-MSCs). We further tested if DS-MSC-conditioned medium (CM), like what was previously reported for BM-MSC CM, has superior wound-healing properties, in both in vitro and in vivo wound models compared with skin fibroblast CM. Methods: DS-MSCs were isolated from HF and cultured in vitro to assess long-term growth potential, colony-forming efficiency (CFE), expression of CD surface markers and differentiation potential. The cytokine expression of DS-MSC CM was determined through an antibody-based protein array analysis. The wound-healing effects of the CM were tested in vitro with the use of human cell cultures and in vivo with the use of a diabetic mouse wound model. Results: In vitro results revealed that DS-MSCs have high growth capacity and CFE while displaying some phenotypes similar to BM-MSCs. DS-MSCs strongly expressed many surface markers expressed in BM-MSCs and could also differentiate into osteoblasts, chondrocytes and adipocytes. DS-MSCs secreted significantly higher proportions of paracrine factors such as interleukin-6 (IL-6), IL-8 and growth-related oncogene. DS-MSC-CM demonstrated enhanced wound-healing effects on human skin keratinocytes, fibroblasts and endothelial cells in vitro, and the wound-healing time in diabetic mice was found to be shorter, compared with vehicle controls. Conclusions: Human HF DS stromal cells demonstrated MSC-like properties and might be an alternative source for therapeutic use in wound healing. © 2015 International Society for Cellular Therapy. Source

Heng W.L.,National Heart Center Singapore | Seck T.,National Heart Center Singapore | Tay C.P.,National Heart Center Singapore | Chua A.,Skin Bank Unit | And 3 more authors.
Cell and Tissue Banking | Year: 2013

Established in 2008, the National Cardiovascular Homograft Bank (NCHB) has been instrumental in creating an available supply of cardiovascular tissues for implantation in Singapore. This article introduces its collaboration with Singapore General Hospital Skin Bank Unit. The procedure of homograft recovery, processing, cryopreservation and quality assurance are presented. Since its establishment, the NCHB has followed the guidelines set by the Ministry of Health Singapore and the American Association of Tissue Banks. A total of 57 homografts had been recovered and 40 homografts were determined to be suitable for clinical use. The most significant reasons for non-clinical use are positive microbiological culture or unsuitable graft condition. Crucial findings prompted reviews and implementation of new procedures to improve the safety of homograft recipients. These include (1) a change in antibiotic decontamination regime from penicillin and streptomycin to amikacin and vancomycin after a review and (2) mandating histopathogical examination since the discovery of cardiac sarcoidosis in a previously undiagnosed donor. Further, the NCHB also routinely performs dengue virus screening, for donors suspected of dengue infection. Cultural factors which affect the donation rate are also briefly explored. By 2010, 31 homografts had been implanted into recipients with congenital or acquired heart valve conditions. More than half of these recipients were children. Post-operative outcomes had been encouraging, with no report of adverse events attributed to implanted homografts. © 2012 The Author(s). Source

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