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Coupling of the radiosensitivity of melanocyte stem cells to their dormancy during the hair cycle
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Abstracts / Journal of Dermatological Science 84 (2016) e1–e88
the altered genes, suggesting that alteration of gene expression by UVB irradiation was different among cell-types. http://dx.doi.org/10.1016/j.jdermsci.2016.08.245 Category 13(P13): Pigmentation and Melanoma P13-01[II-4] Coupling of the radiosensitivity of melanocyte stem cells to their dormancy during the hair cycle Makiko Ueno 1,∗ , Takahiro Aoto 2 , Yasuaki Mohri 2 , Hiroo Yokozeki 1 , Emi K. Nishimura 2 1 Department of Dermatology, Tokyo Medical and Dental University Graduate school of Medicine, Tokyo, Japan 2 Department of Stem Cell Biology, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan
Current studies have revealed that stem cells are more radiosensitive than mature cells. As somatic stem cells are generally slow-cycling cells and are mostly kept in a quiescent state, this conflicts with Bergonié and Tribondeau’s law that actively mitotic cells are the most radiosensitive. In this study, we focused on hair graying to understand the stress-resistance of melanocyte stem cells (McSCs). We irradiated C57BL/6 mice at different hair cycle stages and observed the graying of hair follicles. Then we used DctH2B-GFP transgenic mice which enables the stable visualization of McSCs and an anti-Kit monoclonal antibody which selectively eradicates amplifying McSCs to investigate the coupling of the radiosensitivity of melanocyte stem cells to their cell cycle status. We succeeded in chasing the fate of McSCs throughout the hair cycle and found that McSCs show different radiosensitivities depending on their state during each hair cycle. Hair was significantly grayed when catagen-telogen hair follicles were irradiated, while hair did not show any significant hair graying when anagen hair follicles were irradiated. Histological analyses demonstrated that quiescent McSCs are rather radiosensitive but the coexistence of quiescent and non-quiescent McSCs provides the stem cell pool with radioresistance. The irradiated quiescent McSCs prematurely differentiate in the niche upon their activation without sufficiently renewing themselves or providing mature melanocytes to the hair bulb for hair pigmentation. These data indicate that tissue radiosensitivity is largely dependent on the state of somatic stem cells under their local microenvironment. http://dx.doi.org/10.1016/j.jdermsci.2016.08.246
P13-02[II-5] Induction of melanocytes and fibroblasts from multilineage-differentiating stress-enduring (Muse) cells derived from human adipose tissue Takeshi Yamauchi 1,∗ , Kenshi Yamasaki 1 , Kenichiro Tsuchiyama 1 , Saaya Koike 1 , Mai Inoue 1 , Fumitaka Ogura 2 , Shohei Wakao 2 , Mari Dezawa 2 , Setsuya Aiba 1 1
Department of Dermatology, Tohoku University Graduate School of Medicine, Miyagi, Japan 2 Department of Stem Cell Biology and Histology, Tohoku University Graduate School of Medicine, Japan Multilineage-differentiating stress-enduring (Muse) cell is a multipotent somatic stem cell existing mesenchymal tissues such as dermis. Muse cells are distinct from iPS cells and embryonic stem cells, which require gene transduction and complicated manipulations. To come insight of the clinical usages of Muse cells for skin diseases, we aimed to establish Muse cells from adipose tissues and induce cutaneous cells. We collected human adipose-derived stem cells (hASC) from human adipose tissues obtained from surgical specimens. We identified 2.9 ± 0.35% of SSEA3-positive cells in hASC (hASC-Muse). hASC-Muse expressed alkaline phosphatase, smooth muscle actin, neurofilament and ␣-fetoprotein, indicating hASC-Muse possessed pluripotency. Culturing with 10 factors including Wnt3a, hASC-Muse induced genes of MITF, KIT, TRP1 and gp100 at 3 weeks, DCT at 5 weeks, and tyrosinase at 6 weeks (hASC-Muse-MC). Along with induction of melanogenetic genes, hASC-Muse-MC showed tyrosinase activity after 6 weeks culture. After 6 weeks of hASC-Muse-MC induction, we observed greater L-DOPA activity and melanin contents when hASC-MuseMC was stimulated with bovine pituitary extract or ␣-MSH. Using skin reconstituted culture model, we confirmed hASC-Muse-MC migrate into the basal layer of reconstituted epidermis. We further examined if hASC-Muse differentiates into fibroblasts. After 2week culture with TGF-2 and ascorbic acid, hASC-Muse expressed fibroblast markers such as collagen I and IV (hASC-Muse-Fb). hASCMuse-MC and hASC-Muse-Fb have lower telomerase activity than human primary melanocytes and fibroblasts. These showed that we could generate functional melanocytes and fibroblasts from hASC-Muse without any external gene transduction. http://dx.doi.org/10.1016/j.jdermsci.2016.08.247 P13-03[C09-01] Expression of activation induced deaminase (AID) and BRAFV600E mutation in malignant melanoma Daisuke Omoto ∗ , Emi Mashima, Yumiko Sakuragi, Natsuko Saito, Takashi Yamaguchi, Reiko Watabe, Haruna Yoshioka, Kana Hiromasa, Sanehito Haruyama, Rieko Kubo, Manabu Yoshioka, Daisuke Nishio, Motonobu Nakamura Department of Dermatology, University of Occupational and Environmental Health, Japan Activation-induced cytidine deaminase (AID), a member of a cytidine deaminase family, is essential for somatic hypermutation and class-switch recombination, in immunoglobulin genes. Recently, several studies have shown that abnormally expressed AID acts as a genome mutator that contributes to malignancy such as adult T cell leukemia/lymphoma (ATLL) and gastric cancer.
Abstracts / Journal of Dermatological Science 84 (2016) e1–e88
the altered genes, suggesting that alteration of gene expression by UVB irradiation was different among cell-types. http://dx.doi.org/10.1016/j.jdermsci.2016.08.245 Category 13(P13): Pigmentation and Melanoma P13-01[II-4] Coupling of the radiosensitivity of melanocyte stem cells to their dormancy during the hair cycle Makiko Ueno 1,∗ , Takahiro Aoto 2 , Yasuaki Mohri 2 , Hiroo Yokozeki 1 , Emi K. Nishimura 2 1 Department of Dermatology, Tokyo Medical and Dental University Graduate school of Medicine, Tokyo, Japan 2 Department of Stem Cell Biology, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan
Current studies have revealed that stem cells are more radiosensitive than mature cells. As somatic stem cells are generally slow-cycling cells and are mostly kept in a quiescent state, this conflicts with Bergonié and Tribondeau’s law that actively mitotic cells are the most radiosensitive. In this study, we focused on hair graying to understand the stress-resistance of melanocyte stem cells (McSCs). We irradiated C57BL/6 mice at different hair cycle stages and observed the graying of hair follicles. Then we used DctH2B-GFP transgenic mice which enables the stable visualization of McSCs and an anti-Kit monoclonal antibody which selectively eradicates amplifying McSCs to investigate the coupling of the radiosensitivity of melanocyte stem cells to their cell cycle status. We succeeded in chasing the fate of McSCs throughout the hair cycle and found that McSCs show different radiosensitivities depending on their state during each hair cycle. Hair was significantly grayed when catagen-telogen hair follicles were irradiated, while hair did not show any significant hair graying when anagen hair follicles were irradiated. Histological analyses demonstrated that quiescent McSCs are rather radiosensitive but the coexistence of quiescent and non-quiescent McSCs provides the stem cell pool with radioresistance. The irradiated quiescent McSCs prematurely differentiate in the niche upon their activation without sufficiently renewing themselves or providing mature melanocytes to the hair bulb for hair pigmentation. These data indicate that tissue radiosensitivity is largely dependent on the state of somatic stem cells under their local microenvironment. http://dx.doi.org/10.1016/j.jdermsci.2016.08.246
P13-02[II-5] Induction of melanocytes and fibroblasts from multilineage-differentiating stress-enduring (Muse) cells derived from human adipose tissue Takeshi Yamauchi 1,∗ , Kenshi Yamasaki 1 , Kenichiro Tsuchiyama 1 , Saaya Koike 1 , Mai Inoue 1 , Fumitaka Ogura 2 , Shohei Wakao 2 , Mari Dezawa 2 , Setsuya Aiba 1 1
Department of Dermatology, Tohoku University Graduate School of Medicine, Miyagi, Japan 2 Department of Stem Cell Biology and Histology, Tohoku University Graduate School of Medicine, Japan Multilineage-differentiating stress-enduring (Muse) cell is a multipotent somatic stem cell existing mesenchymal tissues such as dermis. Muse cells are distinct from iPS cells and embryonic stem cells, which require gene transduction and complicated manipulations. To come insight of the clinical usages of Muse cells for skin diseases, we aimed to establish Muse cells from adipose tissues and induce cutaneous cells. We collected human adipose-derived stem cells (hASC) from human adipose tissues obtained from surgical specimens. We identified 2.9 ± 0.35% of SSEA3-positive cells in hASC (hASC-Muse). hASC-Muse expressed alkaline phosphatase, smooth muscle actin, neurofilament and ␣-fetoprotein, indicating hASC-Muse possessed pluripotency. Culturing with 10 factors including Wnt3a, hASC-Muse induced genes of MITF, KIT, TRP1 and gp100 at 3 weeks, DCT at 5 weeks, and tyrosinase at 6 weeks (hASC-Muse-MC). Along with induction of melanogenetic genes, hASC-Muse-MC showed tyrosinase activity after 6 weeks culture. After 6 weeks of hASC-Muse-MC induction, we observed greater L-DOPA activity and melanin contents when hASC-MuseMC was stimulated with bovine pituitary extract or ␣-MSH. Using skin reconstituted culture model, we confirmed hASC-Muse-MC migrate into the basal layer of reconstituted epidermis. We further examined if hASC-Muse differentiates into fibroblasts. After 2week culture with TGF-2 and ascorbic acid, hASC-Muse expressed fibroblast markers such as collagen I and IV (hASC-Muse-Fb). hASCMuse-MC and hASC-Muse-Fb have lower telomerase activity than human primary melanocytes and fibroblasts. These showed that we could generate functional melanocytes and fibroblasts from hASC-Muse without any external gene transduction. http://dx.doi.org/10.1016/j.jdermsci.2016.08.247 P13-03[C09-01] Expression of activation induced deaminase (AID) and BRAFV600E mutation in malignant melanoma Daisuke Omoto ∗ , Emi Mashima, Yumiko Sakuragi, Natsuko Saito, Takashi Yamaguchi, Reiko Watabe, Haruna Yoshioka, Kana Hiromasa, Sanehito Haruyama, Rieko Kubo, Manabu Yoshioka, Daisuke Nishio, Motonobu Nakamura Department of Dermatology, University of Occupational and Environmental Health, Japan Activation-induced cytidine deaminase (AID), a member of a cytidine deaminase family, is essential for somatic hypermutation and class-switch recombination, in immunoglobulin genes. Recently, several studies have shown that abnormally expressed AID acts as a genome mutator that contributes to malignancy such as adult T cell leukemia/lymphoma (ATLL) and gastric cancer.