Human optic nerve head astrocytes as a target for endothelin-1.
OBJECTIVE
To determine whether human optic nerve head astrocytes (hONAs) are target cells for the actions of endothelin (ET)-1, a potent vasoactive peptide, by causing astrocyte proliferation, as occurs in glaucomatous optic nerve heads. ET-1 levels are elevated in glaucomatous eyes, and administration of ET-1 to the retina causes glial activation and optic nerve damage in animal models in a manner similar to that observed in glaucoma.


METHODS
Well-characterized hONAs were used in this study. Cell proliferation of hONAs was assessed, after ET-1 treatment under serum-free culture conditions, with both a formazan assay and [3H]thymidine uptake. ET receptor involvement for cell proliferation was determined with BQ788 (an ETB antagonist), BQ610 (an ETA antagonist), PD142893 (an ET(A/B) mixed antagonist), and sarafotoxin 6C (S6C; an ET(B) agonist). ET-1-induced intracellular calcium ([Ca2+]i) in hONAs was measured by fura-2 imaging. RT-PCR was used to determine whether hONAs express mRNA for preproET-1, ET(A), and ET(B) receptors.


RESULTS
ET-1 (10 and 100 nM) caused a time-dependent proliferation of hONAs, which was completely blocked by PD142893, as detected by two different cell proliferation assays. The effects of ET-1 were blocked by BQ788 and were also mimicked by S6C, indicative of the involvement of ET(B) receptor activation. ET-1-induced elevation in [Ca2+]i, and cell proliferation were both blocked completely by the ET(A) antagonist BQ610, suggesting ET(A) receptor involvement. The hONAs expressed mRNA for ET(A) and ET(B) receptors as well as preproET-1, suggesting that these cells may also be a source for ET-1 in the optic nerve head.


CONCLUSIONS
ET-1 induces astroglial proliferation in cultured human optic nerve head astrocytes through ET(A/B) receptor activation. This is similar to the proliferation of ET-1 in brain astrocytes. These findings suggest that ET-1, which is elevated in glaucoma, could cause proliferation of ONAs in the optic nerve head.
Prasanna
Ganesh
G
Department of Pharmacology and Neuroscience, Division of Cell Biology and Genetics, University of North Texas Health Science Center, Fort Worth, Texas 76107, USA.
Krishnamoorthy
Raghu
R
Clark
Abbot F
AF
Wordinger
Robert J
RJ
Yorio
Thomas
T
D000368:Aged; D000369:Aged, 80 and over; D001253:Astrocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1 / Q000494:pharmacology*; D006801:Humans; D009898:Optic Disk / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D016895:Culture Media, Serum-Free; D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1; D012333:RNA, Messenger; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin; D002118:Calcium