Expression of heat shock protein 47(Hsp47) mRNA levels in rabbit connective tissues during the response to injury and in pregnancy.
Hsp47 (also termed "colligin") is a 47 kDa protein that is localized in the ER and cis-Golgi vesicles of fibrocytes, chondrocytes, and other collagen-secreting cells. Under stress conditions, Hsp47 expression is upregulated as part of the heat shock/stress response that mitigates cell damage from noxious stimuli such as elevated temperature, heavy metals, and oxidative stress. Under non-stress conditions, Hsp47 functions as a collagen-specific molecular chaperone that facilitates intracellular procollagen polypeptide synthesis, and triple helix assembly in connective tissues. Previously it has been shown that levels of collagen-specific gene expression are significantly altered in ligaments, menisci, and other connective tissues of the rabbit following surgically induced injuries (increased), and during pregnancy (decreased). The present study was undertaken to determine whether expression of mRNA for the Hsp47 collagen-binding stress protein was also influenced in these experimental models. Since no sequence information was available on the rabbit Hsp47 gene, a partial cDNA for rabbit Hsp47 was first isolated and cloned using reverse transcriptase PCR (RT-PCR) with degenerate oligonucleotide primers. Rabbit Hsp47 sequence-specific primers then designed enabled analysis of Hsp47 mRNA expression in rabbit connective tissues using semiquantitative RT-PCR. It was found that Hsp47 expression is affected in a complex, tissue-specific manner by injury and pregnancy. Hsp47 transcript levels were elevated in the medial collateral ligament (MCL) of the rabbit knee following surgical induction of a gap injury. Transection of the anterior cruciate ligament (ACL), which leads to chronic progressive damage to menisci of the rabbit knee joint, was accompanied by an upregulation of Hsp47 expression in the medial and lateral menisci. Hsp47 mRNA levels were depressed during pregnancy in the kidney and ACL of primigravid adolescent rabbits, but were not altered in corneal tissue during pregnancy or in the ACL of skeletally mature multiparous females. The changes in Hsp47 transcript levels within these connective tissues following injury/pregnancy often, but not always, paralleled changes in collagen-specific gene expression.
Hart
D A
DA
McCaig Centre for Joint Injury and Arthritis Research, Faculty of Medicine, University of Calgary, AB, Canada.
Reno
C
C
Hellio Le Graverand
M P
MP
Hoffman
L
L
Kulyk
W
W
D000818:Animals; D016118:Anterior Cruciate Ligament / Q000378:metabolism* / Q000601:surgery; D000070598:Anterior Cruciate Ligament Injuries; D002921:Cicatrix / Q000378:metabolism*; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D003315:Cornea / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D008297:Male; D017888:Medial Collateral Ligament, Knee / Q000293:injuries / Q000378:metabolism / Q000601:surgery; D008592:Menisci, Tibial / Q000378:metabolism*; D011247:Pregnancy; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011817:Rabbits
D006360:Heat-Shock Proteins; D012333:RNA, Messenger; D003094:Collagen