A preliminary study of growth characteristics of melanocytes co‐cultured with keratinocytes in vitro

To clarify the characteristic growth of melanocytes (MCs) and Keratinocytes (KCs) in vitro and discuss the mechanism of culturing autologous melanocytes in the treatment of vitiligo. Epidermis cells derived from normal skin tissues were isolated and cultured in vitro. Melanocytes in DOPA staining were observed. The expression level of markers in MCs was detected by qRT‐PCR and the percentage of MCs and KCs were detected by flow cytometry. Cells derived from normal skin tissues mainly included KCs, MCs, and fibroblasts. There were significant differences between the percentage of KC, MC, fibroblasts (P < 0.05), and the expression of Microphthalmia‐associated transcription factor (P < 0.05) and Tyrosinase‐related protein‐2 (P < 0.05) in the second, 10th, 20th, and 30th day. Significant differences were also found between the average numbers of MC stained by DOPA (P < 0.05) and the average percentage of MCs in the 10th, 20th, and 30th Day (P < 0.05). But there were no significant differences between the average percentage of KCs in the 10th, 20th, and 30th Day (P > 0.05) detected by flow cytometry. The number of MCs co‐cultured with KCs in vitro reached the maximum in the 20th Day and this co‐cultured model may contribute to the growth of MCs which could be used in the treatment of vitiligo.

[1]  D. Lipsker,et al.  MITF-High and MITF-Low Cells and a Novel Subpopulation Expressing Genes of Both Cell States Contribute to Intra- and Intertumoral Heterogeneity of Primary Melanoma , 2017, Clinical Cancer Research.

[2]  D. Spandidos,et al.  Soyasaponin Ag inhibits α-MSH-induced melanogenesis in B16F10 melanoma cells via the downregulation of TRP-2 , 2017, International journal of molecular medicine.

[3]  M. Kozłowska-Wojciechowska,et al.  SOX10-MITF pathway activity in melanoma cells , 2016, Archives of medical science : AMS.

[4]  T. Hirobe,et al.  Human fibroblasts treated with hydrogen peroxide stimulate human melanoblast proliferation and melanocyte differentiation, but inhibit melanocyte proliferation in serum-free co-culture system. , 2016, Journal of dermatological science.

[5]  Y. Seo,et al.  The activation of melanogenesis by p-CREB and MITF signaling with extremely low-frequency electromagnetic fields on B16F10 melanoma. , 2016, Life sciences.

[6]  Prakhar Bansal,et al.  Restoration of Pigmentation by Follicular Unit Extraction Transplant in Three Cases of Focal Vitiligo Recalcitrant to Therapy including with Previous Nonculture Melanocyte-keratinocyte Transplant , 2016, International journal of trichology.

[7]  G. Kratz,et al.  Human melanocytes mitigate keratinocyte-dependent contraction in an in vitro collagen contraction assay. , 2015, Burns : journal of the International Society for Burn Injuries.

[8]  Yue Sun,et al.  In vitro‐induced differentiation of bone marrow mesenchymal stem cells into melanocytes , 2015, Cell biology international.

[9]  M. Balañá,et al.  Epidermal stem cells and skin tissue engineering in hair follicle regeneration. , 2015, World journal of stem cells.

[10]  K. Sharquie,et al.  Melanocytes transplantation in patients with vitiligo using needling micrografting technique. , 2013, Journal of drugs in dermatology : JDD.

[11]  Ai-Young Lee.  Role of Keratinocytes in the Development of Vitiligo , 2012, Annals of dermatology.

[12]  I. Chu,et al.  Effects of hydroxybenzyl alcohols on melanogenesis in melanocyte-keratinocyte co-culture and monolayer culture of melanocytes , 2008 .

[13]  I. Chu,et al.  Effects of hydroxybenzyl alcohols on melanogenesis in melanocyte-keratinocyte co-culture and monolayer culture of melanocytes. , 2008, Journal of enzyme inhibition and medicinal chemistry.

[14]  W. Placek,et al.  Autologous Cultured Melanocytes in Vitiligo Treatment , 2007, Dermatologic surgery : official publication for American Society for Dermatologic Surgery [et al.].

[15]  J. Nordlund.  The melanocyte and the epidermal melanin unit: an expanded concept. , 2007, Dermatologic clinics.

[16]  F Beermann,et al.  Genetics of pigment cells: lessons from the tyrosinase gene family. , 2006, Histology and histopathology.

[17]  P. Grimes.  New insights and new therapies in vitiligo. , 2005, JAMA.

[18]  B. Gilchrest,et al.  Fate of melanocytes during development of the hair follicle pigmentary unit. , 2003, The journal of investigative dermatology. Symposium proceedings.

[19]  V. Hearing,et al.  A melanocyte-keratinocyte coculture model to assess regulators of pigmentation in vitro. , 2002, Analytical biochemistry.

[20]  S. Sohn,et al.  Melasma: histopathological characteristics in 56 Korean patients , 2002, The British journal of dermatology.

[21]  Jeffrey S. Chang,et al.  Transplant of Cultured Autologous Pure Melanocytes after Laser‐Abrasion for the Treatment of Segmental Vitiligo , 2000, The Journal of dermatology.

[22]  E. Matoušková,et al.  Tissue-engineered skin in the treatment of vitiligo lesions. , 2000, Folia biologica.

[23]  M. Philippe,et al.  Keratinocyte-melanocyte co-cultures and pigmented reconstructed human epidermis: models to study modulation of melanogenesis. , 1999, Cellular and molecular biology.

[24]  N. Kama,et al.  Autologous skin graft, human dura mater and polypropylene mesh for the repair of ventral abdominal hernias: an experimental study. , 1999, The European journal of surgery = Acta chirurgica.

[25]  M. Picardo,et al.  Chimeric human epidermal reconstructs to study the role of melanocytes and keratinocytes in pigmentation and photoprotection. , 1998, The Journal of investigative dermatology.

[26]  J. Mulliken,et al.  GRAFTING OF BURNS WITH CULTURED EPITHELIUM PREPARED FROM AUTOLOGOUS EPIDERMAL CELLS , 1981, The Lancet.

[27]  H. Green,et al.  Formation of a keratinizing epithelium in culture by a cloned cell line derived from a teratoma , 1975, Cell.

[28]  A. Ylppo.  Children's Clinic, Helsinki. , 1948, Lancet.