mRNA in situ hybridization of TIGR/MYOC in human trabecular meshwork.
OBJECTIVE
To determine the distribution of mRNA expression of the trabecular meshwork-induced glucocorticoid response protein/myocilin (TIGR/MYOC) in human trabecular meshwork.


METHODS
In situ hybridization using a 1.25-kb probe obtained from reverse transcription-polymerase chain reaction of TIGR/MYOC cDNA was performed to determine the location of cell labeling within the different regions of the meshwork. The effect of dexamethasone on the pattern of labeling was studied in organ cultured meshwork. Trabecular meshwork from three sources was studied: enucleated eyes obtained at autopsy, trabeculectomy specimens obtained during filtration surgery, and meshworks from anterior segments in perfusion organ culture. Hybridization was performed on frozen sections, paraffin sections, and sections from JB-4 plastic-embedded tissue.


RESULTS
Labeling for TIGR/MYOC mRNA was present in most trabecular cells of the uveal, corneoscleral, and juxtacanalicular regions but only variably present in the endothelial cells of Schlemm's canal. A similar pattern was found in the trabeculectomy specimens from eyes with primary open-angle or pseudoexfoliative glaucoma. Dexamethasone treatment increased the labeling intensity and number of labeled cells in meshwork, and also the number of labeled endothelial cells of Schlemm's canal. Fresh tissue processed within 12 hours postmortem gave more consistent labeling than older tissue, although some label was found up to 48 hours postmortem. Labeling was found in tissue from all three sources, and with all three embedding techniques; JB-4 sections provided the best morphologic resolution.


CONCLUSIONS
In situ hybridization reveals that mRNA expression for TIGR/MYOC is present in most cells in all regions of the meshwork but only variably present in the endothelial cells of Schlemm's canal. Dexamethasone treatment increased the number and intensity of labeled cells, and also increased the number of labeled cells in the endothelial lining of Schlemm's canal.
Wang
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X
Department of Ophthalmology, Mayo Clinic and Mayo Foundation, Rochester, Minnesota 55905, USA.
Johnson
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DH
D000368:Aged; D000369:Aged, 80 and over; D003598:Cytoskeletal Proteins; D017931:DNA Primers / Q000737:chemistry; D003907:Dexamethasone / Q000494:pharmacology; D017889:Exfoliation Syndrome / Q000378:metabolism* / Q000473:pathology / Q000601:surgery; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism; D005902:Glaucoma, Open-Angle / Q000378:metabolism* / Q000473:pathology / Q000601:surgery; D005938:Glucocorticoids / Q000494:pharmacology; D006023:Glycoproteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D017403:In Situ Hybridization; D008875:Middle Aged; D009924:Organ Culture Techniques; D015347:RNA Probes / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism*; D014129:Trabecular Meshwork / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D014130:Trabeculectomy; D000099179:Myocilin
D003598:Cytoskeletal Proteins; D017931:DNA Primers; D003907:Dexamethasone; D005136:Eye Proteins; D005938:Glucocorticoids; D006023:Glycoproteins; D015347:RNA Probes; D012333:RNA, Messenger; D000099179:Myocilin