Localization of endothelin-converting enzyme in bovine optic nerve and retina

Adnan Dibas, Ganesh Prasanna, Thomas Yorio

Research output: Contribution to journalArticleResearchpeer-review

8 Citations (Scopus)

Abstract

A significant loss and remodeling of the lamina cribrosa tissue leading to the excavation of the optic nerve is seen in glaucoma. Elevated endothelin-1 (ET-1) levels are detected in the aqueous humor of patients of open-angle glaucoma and in the plasma of patients with normal-tension glaucoma. Optic nerve damage, including axonal loss, can be mimicked by ET-1 injection near the optic nerve. ET-1 is produced from its precursor Big ET-1 (38 amino acids) by endothelin-converting enzyme (ECE). Although ET-1 and its receptors have been identified in the retina, little is known of the distribution of ECE at the optic nerve. Presently, ET-1 receptors and Big ET-1 converting activities were characterized in bovine optic nerve and the retina. The ETB receptor was detected in both the optic nerve and retina by immunoblotting and cross-linking, using 125I-ET-1. However, the ETA receptor was detected only in the retina. Big ET-1 conversion activities were detected in the plasma membrane (PM) of bovine retina, but not in the PM of the optic nerve. The retinal PM Big ET-1 converting activity was inhibited by phosphoramidon, thiorphan, and acidification. Furthermore, ECE cytosolic activities were detected in both the optic nerve and retina. Unlike the PM-ECE, cytosolic Big ET-1 converting activities were activated by acidification (pH 6.4), suggesting the involvement of ECE-2-like activity and/or cathepsin activity. Pepstatin, a potent inhibitor of cathepsins, inhibited the optic nerve (ON) cytosolic conversion of Big ET-1 peptide by 50%, and the combination of pepstatin and phosphoramidon, a potent inhibitor of ECE, inhibited the ON cytosolic activity by 86%. By contrast, the combination of both inhibitors weakly inhibited the cytosolic retinal Big ET-1 converting activity. Western blotting revealed the presence of ECE-1 at the PM of the retina not the ON. ECE-2 and cathpesins B, D, and L were detected only in the cytosol of both the retina and ON. In summary, it appears that ET-1 could be produced in the retina and optic nerve by at least two ECE subtypes and, perhaps, cathepsins. Big ET-1 converting activity may be an important target in preventing ET-1-induced optic nerve pathology.

Original languageEnglish
Pages (from-to)288-297
Number of pages10
JournalJournal of Ocular Pharmacology and Therapeutics
Volume21
Issue number4
DOIs
StatePublished - 1 Aug 2005

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Endothelin-1
Optic Nerve
Retina
Cathepsins
Cell Membrane
Endothelin A Receptors
Endothelin-Converting Enzymes
Thiorphan
Low Tension Glaucoma
Aqueous Humor
Open Angle Glaucoma
Immunoblotting
Glaucoma
Cytosol
Western Blotting

Cite this

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title = "Localization of endothelin-converting enzyme in bovine optic nerve and retina",
abstract = "A significant loss and remodeling of the lamina cribrosa tissue leading to the excavation of the optic nerve is seen in glaucoma. Elevated endothelin-1 (ET-1) levels are detected in the aqueous humor of patients of open-angle glaucoma and in the plasma of patients with normal-tension glaucoma. Optic nerve damage, including axonal loss, can be mimicked by ET-1 injection near the optic nerve. ET-1 is produced from its precursor Big ET-1 (38 amino acids) by endothelin-converting enzyme (ECE). Although ET-1 and its receptors have been identified in the retina, little is known of the distribution of ECE at the optic nerve. Presently, ET-1 receptors and Big ET-1 converting activities were characterized in bovine optic nerve and the retina. The ETB receptor was detected in both the optic nerve and retina by immunoblotting and cross-linking, using 125I-ET-1. However, the ETA receptor was detected only in the retina. Big ET-1 conversion activities were detected in the plasma membrane (PM) of bovine retina, but not in the PM of the optic nerve. The retinal PM Big ET-1 converting activity was inhibited by phosphoramidon, thiorphan, and acidification. Furthermore, ECE cytosolic activities were detected in both the optic nerve and retina. Unlike the PM-ECE, cytosolic Big ET-1 converting activities were activated by acidification (pH 6.4), suggesting the involvement of ECE-2-like activity and/or cathepsin activity. Pepstatin, a potent inhibitor of cathepsins, inhibited the optic nerve (ON) cytosolic conversion of Big ET-1 peptide by 50{\%}, and the combination of pepstatin and phosphoramidon, a potent inhibitor of ECE, inhibited the ON cytosolic activity by 86{\%}. By contrast, the combination of both inhibitors weakly inhibited the cytosolic retinal Big ET-1 converting activity. Western blotting revealed the presence of ECE-1 at the PM of the retina not the ON. ECE-2 and cathpesins B, D, and L were detected only in the cytosol of both the retina and ON. In summary, it appears that ET-1 could be produced in the retina and optic nerve by at least two ECE subtypes and, perhaps, cathepsins. Big ET-1 converting activity may be an important target in preventing ET-1-induced optic nerve pathology.",
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Localization of endothelin-converting enzyme in bovine optic nerve and retina. / Dibas, Adnan; Prasanna, Ganesh; Yorio, Thomas.

In: Journal of Ocular Pharmacology and Therapeutics, Vol. 21, No. 4, 01.08.2005, p. 288-297.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

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AU - Dibas, Adnan

AU - Prasanna, Ganesh

AU - Yorio, Thomas

PY - 2005/8/1

Y1 - 2005/8/1

N2 - A significant loss and remodeling of the lamina cribrosa tissue leading to the excavation of the optic nerve is seen in glaucoma. Elevated endothelin-1 (ET-1) levels are detected in the aqueous humor of patients of open-angle glaucoma and in the plasma of patients with normal-tension glaucoma. Optic nerve damage, including axonal loss, can be mimicked by ET-1 injection near the optic nerve. ET-1 is produced from its precursor Big ET-1 (38 amino acids) by endothelin-converting enzyme (ECE). Although ET-1 and its receptors have been identified in the retina, little is known of the distribution of ECE at the optic nerve. Presently, ET-1 receptors and Big ET-1 converting activities were characterized in bovine optic nerve and the retina. The ETB receptor was detected in both the optic nerve and retina by immunoblotting and cross-linking, using 125I-ET-1. However, the ETA receptor was detected only in the retina. Big ET-1 conversion activities were detected in the plasma membrane (PM) of bovine retina, but not in the PM of the optic nerve. The retinal PM Big ET-1 converting activity was inhibited by phosphoramidon, thiorphan, and acidification. Furthermore, ECE cytosolic activities were detected in both the optic nerve and retina. Unlike the PM-ECE, cytosolic Big ET-1 converting activities were activated by acidification (pH 6.4), suggesting the involvement of ECE-2-like activity and/or cathepsin activity. Pepstatin, a potent inhibitor of cathepsins, inhibited the optic nerve (ON) cytosolic conversion of Big ET-1 peptide by 50%, and the combination of pepstatin and phosphoramidon, a potent inhibitor of ECE, inhibited the ON cytosolic activity by 86%. By contrast, the combination of both inhibitors weakly inhibited the cytosolic retinal Big ET-1 converting activity. Western blotting revealed the presence of ECE-1 at the PM of the retina not the ON. ECE-2 and cathpesins B, D, and L were detected only in the cytosol of both the retina and ON. In summary, it appears that ET-1 could be produced in the retina and optic nerve by at least two ECE subtypes and, perhaps, cathepsins. Big ET-1 converting activity may be an important target in preventing ET-1-induced optic nerve pathology.

AB - A significant loss and remodeling of the lamina cribrosa tissue leading to the excavation of the optic nerve is seen in glaucoma. Elevated endothelin-1 (ET-1) levels are detected in the aqueous humor of patients of open-angle glaucoma and in the plasma of patients with normal-tension glaucoma. Optic nerve damage, including axonal loss, can be mimicked by ET-1 injection near the optic nerve. ET-1 is produced from its precursor Big ET-1 (38 amino acids) by endothelin-converting enzyme (ECE). Although ET-1 and its receptors have been identified in the retina, little is known of the distribution of ECE at the optic nerve. Presently, ET-1 receptors and Big ET-1 converting activities were characterized in bovine optic nerve and the retina. The ETB receptor was detected in both the optic nerve and retina by immunoblotting and cross-linking, using 125I-ET-1. However, the ETA receptor was detected only in the retina. Big ET-1 conversion activities were detected in the plasma membrane (PM) of bovine retina, but not in the PM of the optic nerve. The retinal PM Big ET-1 converting activity was inhibited by phosphoramidon, thiorphan, and acidification. Furthermore, ECE cytosolic activities were detected in both the optic nerve and retina. Unlike the PM-ECE, cytosolic Big ET-1 converting activities were activated by acidification (pH 6.4), suggesting the involvement of ECE-2-like activity and/or cathepsin activity. Pepstatin, a potent inhibitor of cathepsins, inhibited the optic nerve (ON) cytosolic conversion of Big ET-1 peptide by 50%, and the combination of pepstatin and phosphoramidon, a potent inhibitor of ECE, inhibited the ON cytosolic activity by 86%. By contrast, the combination of both inhibitors weakly inhibited the cytosolic retinal Big ET-1 converting activity. Western blotting revealed the presence of ECE-1 at the PM of the retina not the ON. ECE-2 and cathpesins B, D, and L were detected only in the cytosol of both the retina and ON. In summary, it appears that ET-1 could be produced in the retina and optic nerve by at least two ECE subtypes and, perhaps, cathepsins. Big ET-1 converting activity may be an important target in preventing ET-1-induced optic nerve pathology.

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