Hye-Ryung Choi

Isolation of human epidermal stem cells by adherence and the reconstruction of skin equivalents

Abstract.The isolation of human epidermal stem cells is critical for their clinical applications. In the present study, we isolated three populations of epidermal keratinocytes according to their ability to adhere to collagen type IV: i.e., rapidly adhering (RA), slowly adhering (SA), and non-adhering (NA) cells. The aim of this study was to characterize RA cells and to investigate the possibility of using these cells for epidermis reconstruction. To identify RA cells, flow cytometric analysis was performed using anti-α6 integrin and anti-CD71 antibodies. RA cells express high levels of α6 integrin and low levels of CD71, which are considered as markers of an epidermal stem cell nature. Furthermore, electron microscopy showed that RA cells are small and have a high nuclear to cytoplasmic ratio, whereas SA and NA cells have well-developed cellular organelles and abundant tonofilaments. Western blot analysis showed that RA cells are slow cycling and express p63, a putative epidermal stem cell marker, whereas SA and NA cells express c-Myc, which is known to regulate stem cell fate. To compare epidermal regenerative abilities, skin equivalents (SEs) were made using RA, SA, and NA cells. The epidermis constructed from RA cells was well formed compared to those formed from SA or NA cells. In addition, only SEs with RA cells expressed α6 integrin and β1 integrin at the basal layer. These results indicate that RA cells represent epidermal stem cells and are predominately comprised of stem cells. Therefore, the isolation of RA cells using a simple technique offers a potential route to their clinical application, because they are easily isolated and provide a high yield of epidermal stem cells.

Effect of tranexamic acid on melasma: a clinical trial with histological evaluation

Backgroundu2002 Melasma is associated with epidermal hyperpigmentation, weak basement membrane, vascular proliferation and increased numbers of mast cell. Tranexamic acid (TXA), a plasmin inhibitor, is reported to improve melasma when injected locally. However, the effects of oral and topical TXA on melasma have not been well studied and the underlying mechanism remains unclear.

Rapid healing and reduced erythema after ablative fractional carbon dioxide laser resurfacing combined with the application of autologous platelet-rich plasma.

BACKGROUND Fractional carbon dioxide laser resurfacing (FxCR) has shown considerable efficacy in reducing wrinkles, although complications such as scarring and prolonged erythema are more common and down‐time is longer than with nonablative laser treatment. Platelet‐rich plasma (PRP), a high concentration of platelets in a small volume of plasma, is known to enhance tissue healing. OBJECTIVE To evaluate the benefits of PRP in the wound healing process after FxCR. MATERIALS AND METHODS Twenty‐five subjects were treated with FxCR on the bilateral inner arms. PRP was prepared from 10 mL of whole blood and applied on a randomly allocated side, with normal saline being used as the contralateral control. Transepidermal water loss (TEWL) and skin color were measured on both sides. Skin biopsies were also taken from five subjects on day 28. RESULTS Significantly faster recovery of TEWL was seen on the PRP‐treated side. The erythema index and melanin index on the PRP‐treated side were lower than on the control side. Biopsy specimens from the PRP‐treated side showed thicker collagen bundles than those from the control side. CONCLUSION Application of autologous PRP is an effective method for enhancing wound healing and reducing transient adverse effects after FxCR treatment. The authors have indicated no significant interest with commercial supporters.

Synthesis and dual biological effects of hydroxycinnamoyl phenylalanyl/prolyl hydroxamic acid derivatives as tyrosinase inhibitor and antioxidant.

We previously reported that caffeoyl-amino acidyl-hydroxamic acid (CA-Xaa-NHOH) acted as both a good antioxidant and tyrosinase inhibitor, in particular when caffeic acid was conjugated with proline or amino acids having aromatic ring like phenylalanine. Here, various hydroxycinnamic acid (HCA) derivatives were further conjugated with phenylalanyl hydroxamic acid and prolyl hydroxamic acid (HCA-Phe-NHOH and HCA-Pro-NHOH) to study the structure and activity relationship as both antioxidants and tyrosinase inhibitors. When their biological activities were evaluated, all HCA-Phe-NHOH and HCA-Pro-NHOH exhibited enhanced antioxidant activity compared to HCA alone. Moreover, derivatives of caffeic acid, ferulic acid, and sinapic acid inhibited lipid peroxidation more efficiently than vitamin E analogue (Trolox). In addition, derivatives of caffeic acid and sinapic acid efficiently inhibited tyrosinase activity and reduced melanin content in melanocytes Mel-Ab cell.

Copper–GHK increases integrin expression and p63 positivity by keratinocytes

Glycyl-l-histidyl-l-lysyl (GHK) possesses a high affinity for copper(II) ions, with which it spontaneously forms a complex (copper–GHK). It is well known that copper–GHK plays a physiological role in the process of wound healing and tissue repair by stimulating collagen synthesis in fibroblasts. This study was conducted to investigate the effects of copper–GHK on keratinocytes. Proliferative effects were analyzed and hematoxylin and eosin staining and immunohistochemistry were conducted to evaluate the effects of copper–GHK in skin equivalent (SE) models. In addition, western blotting was performed. In monolayer cultured keratinocytes, copper–GHK increased the proliferation of keratinocytes. When the SE models were evaluated, basal cells became cuboidal when copper–GHK was added. Immunohistochemical analysis revealed that copper–GHK increased proliferating cell nuclear antigen (PCNA) and p63 positivity. Furthermore, the expression of integrin α6 and β1 increased in SE models, and these results were confirmed by Western blotting. The results of this study indicate that treatment with copper–GHK may increase the proliferative potential of basal keratinocytes by modulating the expression of integrins, p63 and PCNA. In addition, increased levels of p63, a putative stem cell marker of the skin, suggests that copper–GHK promotes the survival of basal stem cells in the skin.

Niche interactions in epidermal stem cells

Within the epidermis and dermis of the skin, cells secrete and are surrounded by the extracellular matrix (ECM), which provides structural and biochemical support. The ECM of the epidermis is the basement membrane, and collagen and other dermal components constitute the ECM of the dermis. There is significant variation in the composition of the ECM of the epidermis and dermis, which can affect cell to cell and cell to ECM interactions. These interactions, in turn, can influence biological responses, aging, and wound healing; abnormal ECM signaling likely contributes to skin diseases. Thus, strategies for manipulating cell-ECM interactions are critical for treating wounds and a variety of skin diseases. Many of these strategies focus on epidermal stem cells, which reside in a unique niche in which the ECM is the most important component; interactions between the ECM and epidermal stem cells play a major role in regulating stem cell fate. As they constitute a major portion of the ECM, it is likely that integrins and type IV collagens are important in stem cell regulation and maintenance. In this review, we highlight recent research-including our previous work-exploring the role that the ECM and its associated components play in shaping the epidermal stem cell niche.

Oligosaccharides of hyaluronic acid increased epidermal cell stemness by modulation of integrin expression

Hyaluronic acid (HA) is an abundant matrix component and is degraded into polymers of various sizes. These generated fragments appear to have properties that affect wound healing of the skin. In industry, small‐sized HA is used as a moisturizing agent but can have biologic effects when it is absorbed through the skin with barrier disruption.

Dual effects of caffeoyl-amino acidyl-hydroxamic acid as an antioxidant and depigmenting agent.

Anti-aging and depigmentation have both been an important subject of study for skin disease and the cosmetic industry. Caffeic acid (CA) has shown synergistically enhanced antioxidant activity when conjugated with amino acids. Hydroxamic acid (NHOH) is a well-known metal chelator, potentially having both tyrosinase inhibitory activity and free radical scavenging activity. We prepared caffeoyl-amino acidyl-hydroxamic acid (CA-Xaa-NHOH) and found that caffeoyl-prolyl-hydroxamic acid (CA-Pro-NHOH) contained excellent antioxidant activity and tyrosinase inhibitory activity by various bioassay systems. Also, CA-Pro-NHOH showed mild melanogenesis inhibitory activity in Mel-Ab cells.

Enhanced cell permeability of kojic acid-phenylalanine amide with metal complex.

Kojic acid-phenylalanine amide (KA-F-NH(2)), which showed an excellent tyrosinase inhibitory activity, did not inhibit melanogenesis in melanocyte due to its low cell permeability. To enhance its cell permeability by increasing lipophilicity, we prepared metal coordination compounds of KA-F-NH(2) and characterized them by FT-IR and ICP analysis. The metal complex of KA-F-NH(2) inhibited mushroom tyrosinase activity as much as KA-F-NH(2) and reduced melanin contents in melanocyte efficiently.

Kojic acid–amino acid amide metal complexes and their melanogenesis inhibitory activities

Tyrosinase plays a critical role in the early stages of the melanin synthetic pathway by catalyzing the oxidation of the substrate. Therefore, tyrosinase inhibitors have been intensively studied in both cosmetic and food industries to develop hypopigmentary agents and prevent enzymatic browning in food. Previously, we reported that kojic acid–amino acid amide (KA‐AA‐NH2) showed enhanced tyrosinase inhibitory activity compared with kojic acid alone, but this was not observed in a cell test because of poor cell permeability. To enhance cell permeability, we prepared copper and zinc complexes of KA‐AA‐NH2 and characterized them using FT‐IR spectroscopy, ESI‐MS spectrometry, and inductively coupled plasma analysis. We then showed that KA‐AA‐NH2 copper complexes exhibited melanogenesis inhibitory activity in Mel‐Ab cells. Copyright