Methods: Embryonic day 14-18 (E14-18) and neonatal ICR mice were used in this study. Expression of cartducin mRNA in the condylar cartilage was examined by in situ hybridization and RT-PCR. Condylar cartilage and Meckel's cartilage from E14.5 mice were cultured for 4 days with AS-ODN, and tissues were retrieved for immnohitochemistry and real-time PCR analysis of cartilage marker proteins.
Results: RT-PCR analysis showed that cartducin started to be expressed in the condylar anlagen and also in Meckel's cartilage at E 14. At E15, cartducin mRNA was detected in the proliferating zone of condylar cartilage by in situ hybridization. At E16, mRNA was detected from fibrous cell zone to upper hypertrophic cell zone. Cartducin abrogation by antisense oligonucleotides induced curvature-form dysmorphogenesis of Meckel's cartilage and sever perichondrium atrophy, compared to random sequence-ODN and ODN-free control groups. Significant inhibition of the cartilage matrix synthesis and reduction of the number of chondrocytes were also observed in the AS-ODN treated condylar cartilage. Immunostaining for aggrecan, type X collagen, Tenascin-C and Type I collagen were colocalized within the matrix. Reai-time PCR analyzes indicated that mRNA levels of aggrecan, collagen types II and X were dramatically reduced in the AS-ODN treated condylar cartilage.
Conclusion: These results suggest that cartducin may be involved in the maintaining perichondrium as well as the regulation of condylar cartilage formation.