In ovo gene manipulation of melanocytes and their adjacent keratinocytes during skin pigmentation of chicken embryos.
Dev Growth Differ
; 57(3): 232-41, 2015 Apr.
Article
in En
| MEDLINE
| ID: mdl-25739909
ABSTRACT
During skin pigmentation in avians and mammalians, melanin is synthesized in the melanocytes, and subsequently transferred to adjacently located keratinocytes, leading to a wide coverage of the body surface by melanin-containing cells. The behavior of melanocytes is influenced by keratinocytes shown mostly by in vitro studies. However, it has poorly been investigated how such intercellular cross-talk is regulated in vivo because of a lack of suitable experimental models. Using chicken embryos, we developed a method that enables in vivo gene manipulations of melanocytes and keratinocytes, where these cells are separately labeled by different genes. Two types of gene transfer techniques were combined one was a retrovirus-mediated gene infection into the skin/keratinocytes, and the other was the in ovo DNA electroporation into neural crest cells, the origin of melanocytes. Since the Replication-Competent Avian sarcoma-leukosis virus long terminal repeat with Splice acceptor (RCAS) infection was available only for the White leghorn strain showing little pigmentation, melanocytes prepared from the Hypeco nera (pigmented) were back-transplanted into embryos of White leghorn. Prior to the transplantation, enhanced green fluorescent protein (EGFP)(+) Neo(r+) -electroporated melanocytes from Hypeco nera were selectively grown in G418-supplemented medium. In the skin of recipient White leghorn embryos infected with RCAS-mOrange, mOrange(+) keratinocytes and transplanted EGFP(+) melanocytes were frequently juxtaposed each other. High-resolution confocal microscopy also revealed that transplanted melanocytes exhibited normal behaviors regarding distribution patterns of melanocytes, dendrite morphology, and melanosome transfer. The method described in this study will serve as a useful tool to understand the mechanisms underlying intercellular regulations during skin pigmentation in vivo.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Ovum
/
Skin Pigmentation
/
Keratinocytes
/
Melanocytes
Limits:
Animals
Language:
En
Journal:
Dev Growth Differ
Year:
2015
Document type:
Article
Affiliation country: