Dependence of corneal stem/progenitor cells on ocular surface innervation.
OBJECTIVE
Neurotrophic keratopathy (NK) is a corneal degeneration associated with corneal nerve dysfunction. It can cause corneal epithelial defects, stromal thinning, and perforation. However, it is not clear if and to which extent epithelial stem cells are affected in NK. The purpose of this study was to identify the relationship between corneolimbal epithelial progenitor/stem cells and sensory nerves using a denervated mouse model of NK.


METHODS
NK was induced in mice by electrocoagulation of the ophthalmic branch of the trigeminal nerve. The absence of corneal nerves was confirmed with β-III tubulin immunostaining and blink reflex test after 7 days. ATP-binding cassette subfamily G member 2 (ABCG2), p63, and hairy enhancer of split 1 (Hes1) were chosen as corneolimbal stem/progenitor cell markers and assessed in denervated mice versus controls by immunofluorescent microscopy and real-time PCR. In addition, corneolimbal stem/progenitor cells were detected as side population cells using flow cytometry, and colony-forming efficiency assay was performed to assess their function.


RESULTS
ABCG2, p63, and Hes1 immunostaining were significantly decreased in denervated eyes after 7 days. Similarly, the expression levels of ABCG2, p63, K15, Hes1, and N-cadherin transcripts were also significantly decreased in denervated eyes. Stem/progenitor cells measured as side population from NK mice were decreased by approximately 75% compared with normals. In addition, the authors found a significant (P = 0.038) reduction in colony-forming efficiency of stem/progenitor cells harvested from denervated eyes.


CONCLUSIONS
Corneolimbal stem/progenitor cells are significantly reduced after depletion of sensory nerves. The data suggest a critical role of innervation in maintaining stem cells and/or the stem cell niche.
Schepens Eye Research Institute, Boston, Massachusetts 02114, USA.
Hiroki
H
Ueno
Giulio
G
Ferrari
Takaaki
T
Hattori
Daniel R
DR
Saban
Kishore R
KR
Katikireddy
Sunil K
SK
Chauhan
Reza
R
Dana
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000096:biosynthesis / Q000235:genetics*; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000096:biosynthesis / Q000235:genetics*; D015415:Biomarkers / Q000378:metabolism; D002470:Cell Survival; D003315:Cornea / Q000294:innervation* / Q000378:metabolism / Q000473:pathology; D003316:Corneal Diseases / Q000209:etiology / Q000235:genetics / Q000473:pathology*; D003714:Denervation / Q000379:methods; D004195:Disease Models, Animal; D004572:Electrolysis; D019573:Epithelium, Corneal / Q000378:metabolism / Q000473:pathology; D005434:Flow Cytometry; D005786:Gene Expression Regulation; D016147:Genes, Tumor Suppressor; D018257:Helix-Loop-Helix Motifs; D018398:Homeodomain Proteins / Q000096:biosynthesis / Q000235:genetics*; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence; D010750:Phosphoproteins / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D013234:Stem Cells / Q000378:metabolism / Q000473:pathology*; D015534:Trans-Activators / Q000096:biosynthesis / Q000235:genetics*; D000072056:Transcription Factor HES-1; D061221:Trigeminal Nerve Injuries / Q000150:complications / Q000378:metabolism / Q000473:pathology
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D051792:Basic Helix-Loop-Helix Proteins; D015415:Biomarkers; D018398:Homeodomain Proteins; D010750:Phosphoproteins; D012333:RNA, Messenger; D015534:Trans-Activators; D000072056:Transcription Factor HES-1; C501412:Abcg2 protein, mouse; C489734:Hes1 protein, mouse; C486419:Trp63 protein, mouse