Expression of collagens I, III, IV and V mRNA in excimer wounded rat cornea: analysis by semi-quantitative PCR.
A semi-quantitative polymerase chain reaction (PCR) methodology was used to evaluate the kinetic changes occurring in collagens I, III, IV and V mRNA in rat cornea following excimer laser keratectomy. cDNA was synthesized from RNA extracted from rat cornea at various times following excimer laser photoablative keratectomy. Collagen cDNA sequences were subsequently amplified using specific sets of oligonucleotide primers. Competitive PCR amplification was carried out using an internal standard so that a semi-quantitative analysis of message for synthesis of collagen types I, III, IV and V could be performed and time course dynamics of message for these collagens studied. There was a biphasic increase in the levels of collagens III, IV and V mRNA following excimer laser keratectomy. Collagen I mRNA levels demonstrated a more sustained increase and were still elevated at 6 weeks following wounding. Collagens IV and V mRNA showed the largest increase with an approximate three fold increase over controls between 4 days and 1 week. Our results demonstrate that upregulation of stromal collagens I, III, and V mRNA and basement membrane collagen IV mRNA occurs in rat cornea following excimer laser keratectomy.
Rhoads Molecular Immunology, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, USA.
W J
WJ
Power
A H
AH
Kaufman
J
J
Merayo-Lloves
V
V
Arrunategui-Correa
C S
CS
Foster
D000818:Animals; D001483:Base Sequence; D003094:Collagen / Q000096:biosynthesis* / Q000235:genetics; D003315:Cornea / Q000378:metabolism* / Q000601:surgery; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000138:chemical synthesis; D004587:Electrophoresis, Agar Gel; D005260:Female; D054018:Lasers, Excimer; D008969:Molecular Sequence Data; D018903:Photorefractive Keratectomy; D016133:Polymerase Chain Reaction / Q000379:methods*; D012313:RNA / Q000302:isolation & purification; D012333:RNA, Messenger / Q000096:biosynthesis*; D051381:Rats; D015854:Up-Regulation; D014945:Wound Healing / Q000502:physiology*
D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D012313:RNA; D003094:Collagen