Expression of tenascin-C splice variants in normal and bullous keratopathy human corneas.
OBJECTIVE
To characterize the expression patterns of tenascin-C (TN-C) splice variants in normal corneas and in those affected by pseudophakic-aphakic bullous keratopathy (PBK-ABK).


METHODS
Alternatively spliced variants of TN-C mRNA from normal and age-matched human corneas with PBK-ABK were analyzed by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot hybridization, using beta2-microglobulin as a housekeeping gene to normalize the samples. Normal and PBK-ABK corneas were studied by immunofluorescence and western blot analysis with antibodies to specific fibronectin type III-like (FN-III) repeats of TN-C.


RESULTS
Tenascin-C mRNA expression was detected in epithelial, stromal, and endothelial cells of normal and PBK-ABK central corneas, although the protein was seen only in diseased corneas. Assessed by RT-PCR, PBK-ABK corneas expressed approximately three times more total TN-C mRNA than did normal corneas. Four major TN-C mRNA variants (with no FN-III insertional repeats or with retained insertional repeats D, A1, or A1+D) and three minor variants (with retained repeats A1+A2, A1+A2+D, or A1+A2+B+D) were much more abundant in PBK-ABK than in normal corneas. Repeat A1 was more abundant in PBK-ABK TN-C protein than repeats A2, A3, B, or D. Major TN-C variants in PBK-ABK corneas were in the range of 190 kDa to 240 kDa.


CONCLUSIONS
Expression of TN-C mRNA and protein is higher in PBK-ABK corneas than in normal corneas. This increase mainly concerns relatively small TN-C splice variants that may affect corneal cell adhesion and migration and contribute to the exacerbation of PBK-ABK.
Ljubimov
A V
AV
Ophthalmology Research Laboratories, Burns and Allen Research Institute, Cedars-Sinai Medical Center, University of California Los Angeles Medical School Affiliate, 90048, USA.
Saghizadeh
M
M
Spirin
K S
KS
Khin
H L
HL
Lewin
S L
SL
Zardi
L
L
Bourdon
M A
MA
Kenney
M C
MC
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D017398:Alternative Splicing; D015153:Blotting, Western; D003315:Cornea / Q000378:metabolism*; D003316:Corneal Diseases / Q000378:metabolism* / Q000473:pathology; D017931:DNA Primers / Q000737:chemistry; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008875:Middle Aged; D015345:Oligonucleotide Probes / Q000737:chemistry; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism*; D019063:Tenascin / Q000235:genetics / Q000378:metabolism*
D017931:DNA Primers; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D019063:Tenascin