Musculoskeletal Disease Center, Jerry L. Pettis VA Medical Center, Loma Linda, CA 92357, USA.
Departments of Medicine, Loma Linda University, Loma Linda, CA 92354, USA.
Cells. 2021 Aug 26;10(9):2200. doi: 10.3390/cells10092200.
We previously showed that conditional disruption of the gene in chondrocytes led to a massive increase in long bone trabecular bone mass. Loss of gene expression or inhibition of PHD2 activity by a specific inhibitor resulted in a several-fold compensatory increase in expression in chondrocytes. To determine if expression of PHD3 plays a role in endochondral bone formation, we conditionally disrupted the gene in chondrocytes by crossing floxed () mice with mice. Loss of expression in the chondrocytes of ; conditional knockout (cKO) mice was confirmed by real time PCR. At 16 weeks of age, neither body weight nor body length was significantly different in the cKO mice compared to ; wild-type (WT) mice. Areal BMD measurements of total body as well as femur, tibia, and lumbar skeletal sites were not significantly different between the cKO and WT mice at 16 weeks of age. Micro-CT measurements revealed significant gender differences in the trabecular bone volume adjusted for tissue volume at the secondary spongiosa of the femur and the tibia for both genotypes, but no genotype difference was found for any of the trabecular bone measurements of either the femur or the tibia. Trabecular bone volume of distal femur epiphysis was not different between cKO and WT mice. Histology analyses revealed cKO mice exhibited a comparable chondrocyte differentiation and proliferation, as evidenced by no changes in cartilage thickness and area in the cKO mice as compared to WT littermates. Consistent with the in vivo data, lentiviral shRNA-mediated knockdown of expression in chondrocytes did not affect the expression of markers of chondrocyte differentiation (, , , ). Our study found that but not expressed in chondrocytes regulates endochondral bone formation, and the compensatory increase in expression in the chondrocytes of cKO mice is not the cause for increased trabecular bone mass in cKO mice.
我们之前的研究表明，条件性破坏软骨细胞中的基因会导致长骨小梁骨量大量增加。基因表达的丧失或特定抑制剂对 PHD2 活性的抑制导致软骨细胞中表达的几倍代偿性增加。为了确定 PHD3 的表达是否在软骨内骨形成中起作用，我们通过将 floxed () 小鼠与 小鼠杂交，在软骨细胞中条件性破坏 基因。 ; 条件性敲除 (cKO) 小鼠中 基因表达的缺失通过实时 PCR 得到证实。在 16 周龄时，与 ; 野生型 (WT) 小鼠相比， cKO 小鼠的体重和体长均无显著差异。16 周龄时，cKO 和 WT 小鼠的全身、股骨、胫骨和腰椎骨骼部位的面积 BMD 测量值无显著差异。两种基因型的股骨和胫骨次级松质骨组织体积校正的小梁骨体积的微 CT 测量值存在显著的性别差异，但在股骨或胫骨的任何小梁骨测量值中均未发现基因型差异。cKO 和 WT 小鼠的远端股骨骨骺小梁骨体积无差异。组织学分析表明，cKO 小鼠表现出可比的软骨细胞分化和增殖，因为与 WT 同窝仔相比，cKO 小鼠的软骨厚度和面积没有变化。与体内数据一致，软骨细胞中 表达的慢病毒 shRNA 介导的敲低不会影响软骨细胞分化标志物的表达 (、、、)。我们的研究发现，表达于软骨细胞中的 而非 调节软骨内骨形成，并且 cKO 小鼠中软骨细胞中 表达的代偿性增加不是 cKO 小鼠中小梁骨量增加的原因。
