GPR48 regulates epithelial cell proliferation and migration by activating EGFR during eyelid development.
OBJECTIVE
Eyelid development is a dynamic process involving cell proliferation, differentiation, and migration regulated by a number of growth factors and cytokines. Mice deficient in the orphan G protein-coupled receptor 48 (GPR48) showed an eye open at birth (EOB) phenotype. In this study, the authors attempted to clarify the role of GPR48 in eyelid development and the molecular mechanisms leading to the EOB phenotype.


METHODS
Phenotypic analysis of the eyelids of Gpr48(-/-) mice was carried out using histology and scanning electron microscopy. GPR48 expression pattern was determined using X-gal staining. In vitro scratch assay was used to determine cell motility defects in Gpr48(-)(/)(-) keratinocytes. The molecular mechanism underlying GPR48-mediated eyelid closure was explored using Western blot and immunostaining analyses. Expression levels of EGFR and its phosphorylated counterpart were examined in Gpr48(-/-) and wild-type keratinocytes and in eyelids.


RESULTS
GPR48 is highly expressed in the epithelium and apical mesenchymal cells of eyelids during embryonic development. Detailed analysis revealed that Gpr48(-/-) mice exhibited delayed leading-edge extension, reduced filopodia formation, and decreased rounded periderm cell formation around eyelid margins. Keratinocytes lacking GPR48 are defective in cell proliferation and migration with reduced F-actin staining. In addition, the phosphorylation of EGFR was dramatically decreased in cultured keratinocytes and developing eyelids in the absence of GPR48.


CONCLUSIONS
Inactivation of GPR48 induces the EOB phenotype by reducing epithelial cell proliferation and migration, indicating that GPR48 plays an essential role in eyelid development. Furthermore, GPR48 contributes to eyelid development through the regulation of the EGFR signaling pathway.
Jin
Chang
C
School of Ophthalmology and Optometry, Eye Hospital, Wenzhou Medical College, Wenzhou, Zhejiang, PR China.
Yin
Furong
F
Lin
Mimi
M
Li
Hongxia
H
Wang
Zhenlian
Z
Weng
Jinsheng
J
Liu
Mingyao
M
Da Dong
Xiang
X
Qu
Jia
J
Tu
LiLi
L
D000818:Animals; D015153:Blotting, Western; D001973:Bromodeoxyuridine / Q000378:metabolism; D002465:Cell Movement / Q000502:physiology*; D049109:Cell Proliferation; D002478:Cells, Cultured; D066246:ErbB Receptors / Q000378:metabolism*; D005143:Eyelids / Q000196:embryology* / Q000378:metabolism / Q000648:ultrastructure; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D020868:Gene Silencing / Q000502:physiology; D005838:Genotype; D007124:Immunoenzyme Techniques; D020287:In Situ Nick-End Labeling; D015603:Keratinocytes / Q000166:cytology* / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008855:Microscopy, Electron, Scanning; D010766:Phosphorylation; D043562:Receptors, G-Protein-Coupled / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology
C496200:LGR4 protein, mouse; D043562:Receptors, G-Protein-Coupled; D066246:ErbB Receptors; D001973:Bromodeoxyuridine