Abnormal synthesis of cartilage-characteristic proteoglycan in azaserine-induced micromelial limbs.
Administration of azaserine (250 micrograms) to day-4 chick embryos in ovo was shown to induce micromelial limbs. In the present study, biosynthesis of cartilage-characteristic proteoglycan H (PG-H) as an index of limb chondrogenesis was examined in normal and micromelial hind limbs from day-7 chick embryos by biochemical and immunological methods. (1) Metabolic labelling of the micromelial limbs with [6-3H]-glucose and [35S]sulphate, followed by analysis of labelled proteoglycans by glycerol-density-gradient centrifugation under dissociative conditions, showed a marked reduction in PG-H synthesis. (2) PG-H synthesized by micromelial limbs differed from that synthesized by normal limbs in possessing a slower sedimenting velocity and much lower amounts of chondroitin sulphates. (3) The amount of PG-H core protein in micromelial limbs was significantly decreased to about 19% on a per limb basis and about 42% on a per DNA basis of that in normal limbs, as determined by e.l.i.s.a. (4) The transition from PG-M to PG-H during limb formation was retarded in micromelial limbs as judged by an indirect immunofluorescence technique using antibodies against PG-M and PG-H. (5) The deficiency of incorporation of labelled glucose into chondroitin sulphate chains of PG-H in micromelial limbs was partially restored by using [6-3H]-glucosamine as a precursor, suggesting that the synthesis of UDP-N-acetylhexosamine, required for chondroitin sulphate chain synthesis of PG-H in micromelial limbs, was decreased. These results suggest that the reduction in the synthesis of PG-H as well as the production of an abnormal form of PG-H during a critical period of limb morphogenesis may be important factors in explaining the micromelia induced by azaserine.
Honda
A
A
Department of Biochemistry, Tokyo College of Pharmacy, Japan.
Tsuboi
I
I
Kimata
K
K
Hirabayashi
Y
Y
Yamada
K
K
Mori
Y
Y
D000014:Abnormalities, Drug-Induced / Q000378:metabolism*; D000818:Animals; D001377:Azaserine / Q000633:toxicity*; D002356:Cartilage / Q000378:metabolism*; D002642:Chick Embryo; D006614:Hindlimb / Q000002:abnormalities* / Q000187:drug effects / Q000378:metabolism; D011509:Proteoglycans / Q000096:biosynthesis*
D011509:Proteoglycans; D001377:Azaserine