Mitogenic and metabolic actions of epidermal growth factor on rat articular chondrocytes: modulation by fetal calf serum, transforming growth factor-beta, and tyrphostin.
The effects of human recombinant epidermal growth factor (EGF) on rat articular chondrocytes from humeral and femoral head cartilage of 21-day-old Wistar rats were analyzed. The cells were cultured under standard conditions as monolayers. Cell proliferation was studied by [3H]thymidine incorporation and determination of DNA content, proteoglycan synthesis by [35S]sulfate incorporation, and collagen synthesis by [3H]proline incorporation. The presence of specific receptors was confirmed by [125I]-EGF binding and that of EGF and EGF-receptor (EGF-R) mRNA by reverse transcription and the polymerase chain reaction. EGF (0.5-2.5 ng/ml) stimulated [3H]thymidine incorporation and increased DNA content of cultures. The effect was strongest when serum concentration was low (< or =1%) and was lost at high (> or =7.5%) serum concentrations. The EGF-induced effect on deoxynucleic acid synthesis was inhibited by transforming growth factor-beta and tyrphostin, a tyrosine kinase inhibitor that blocks the phosphorylation of tyrosine residues on EGF-R. Cultured rat articular chondrocytes possess a single class of high-affinity binding sites (Kd 0.18 nM). There were about 4.5 x 10(9) binding sites per microgram of DNA or about 37,800 binding sites per cell with 8.3 pg DNA per cell. Cultured cells contained EGF mRNA and EGF-R mRNA. Incubation of cells with EGF for 24 h decreased the EGF mRNA transcripts and increased the EGF-R mRNA levels. These findings suggest that EGF probably takes part in the regulation of chondrocyte activity under normal and presumably pathological conditions.
Ribault
D
D
INSERM U.349, Paris, France.
Khatib
A M
AM
Panasyuk
A
A
Barbara
A
A
Bouizar
Z
Z
Mitrovic
R D
RD
D000818:Animals; D001769:Blood; D002358:Cartilage, Articular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D002417:Cattle; D002453:Cell Cycle / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis; D004247:DNA / Q000096:biosynthesis; D015536:Down-Regulation; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D066246:ErbB Receptors / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D008297:Male; D009570:Nitriles / Q000494:pharmacology*; D011509:Proteoglycans / Q000096:biosynthesis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D009570:Nitriles; D011509:Proteoglycans; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D004815:Epidermal Growth Factor; D003094:Collagen; D004247:DNA; D066246:ErbB Receptors