3β-Hydroxysterol-Delta24 reductase plays an important role in long bone growth by protecting chondrocytes from reactive oxygen species

Abstract

Desmosterolosis is an autosomal recessive disease caused by mutations in the 3β-hydroxysterol-Delta24 reductase (DHCR24) gene, with severe developmental anomalies including short limbs. We utilized DHCR24 knockout (KO) mice to study the underlying bone pathology. Because the KO mice died within a few hours after birth, we cultured metatarsal bones from newborn mice. The growth of bones from KO mice was significantly retarded after 1week of culture. Absence of proliferating chondrocytes in the growth plate and abnormal hypertrophy of prehypertrophic chondrocytes were observed in the bones from KO mice. Hypertrophic differentiation was evidenced by higher expression of Indian hedgehog, alkaline phosphatase, and matrix metalloproteinase 13. Since elevated levels of reactive oxygen species (ROS) during chondrogenesis are known to inhibit proliferation and to initiate chondrocyte hypertrophy in the growth plate, and since DHCR24 acts as a potent ROS scavenger, we hypothesized that the abnormal chondrocyte proliferation and differentiation in KO mice were due to decreased ROS scavenging activity. Treatment with an antioxidant, N-acetyl cysteine, could correct the abnormalities observed in the bones from KO mice. Treatment of bones from wild-type mice with U18666A, a chemical inhibitor of DHCR24, resulted in short broad bones with a disrupted proliferating zone. Treatment of ATDC cells with hydrogen peroxide (H(2)O(2)) induced hypertrophic changes as evidenced by the expression of the marker genes specific for hypertrophic chondrocyte differentiation. H(2)O(2)-induced hypertrophic change was prevented by adenoviral delivery of DHCR24. Induction of chondrocyte differentiation in ATDC cells by insulin was associated with increased ROS production that was markedly enhanced by treatment of ATDC5 cells with DHCR24 siRNA. This is the first demonstration that DHCR24 plays an important role in long bone growth by protecting chondrocytes from ROS.