CIHR Group in Skeletal Development and Remodeling, Department of Physiology and Pharmacology, Schulich School of Medicine and Dentistry, The University of Western Ontario, London, Ontario, Canada.
J Cell Mol Med. 2009 Sep;13(9B):3497-516. doi: 10.1111/j.1582-4934.2009.00684.x.
Elucidating the signalling pathways that regulate chondrocyte differentiation, such as the actin cytoskeleton and Rho GTPases, during development is essential for understanding of pathological conditions of cartilage, such as chondrodysplasias and osteoarthritis. Manipulation of actin dynamics in tibia organ cultures isolated from E15.5 mice results in pronounced enhancement of endochondral bone growth and specific changes in growth plate architecture. Global changes in gene expression were examined of primary chondrocytes isolated from embryonic tibia, treated with the compounds cytochalasin D, jasplakinolide (actin modifiers) and the ROCK inhibitor Y27632. Cytochalasin D elicited the most pronounced response and induced many features of hypertrophic chondrocyte differentiation. Bioinformatics analyses of microarray data and expression validation by real-time PCR and immunohistochemistry resulted in the identification of the nuclear receptor retinoid related orphan receptor-alpha (Ror-alpha) as a novel putative regulator of chondrocyte hypertrophy. Expression of Ror-alpha target genes, (Lpl, fatty acid binding protein 4 [Fabp4], Cd36 and kruppel-like factor 5 [Klf15]) were induced during chondrocyte hypertrophy and by cytochalasin D and are cholesterol dependent. Stimulation of Ror-alpha by cholesterol results in increased bone growth and enlarged, rounded cells, a phenotype similar to chondrocyte hypertrophy and to the changes induced by cytochalasin D, while inhibition of cholesterol synthesis by lovastatin inhibits cytochalasin D induced bone growth. Additionally, we show that in a mouse model of cartilage specific (Col2-Cre) Rac1, inactivation results in increased Hif-1alpha (a regulator of Rora gene expression) and Ror-alpha(+) cells within hypertrophic growth plates. We provide evidence that cholesterol signalling through increased Ror-alpha expression stimulates chondrocyte hypertrophy and partially mediates responses of cartilage to actin dynamics.
阐明调控软骨细胞分化的信号通路，如肌动蛋白细胞骨架和 Rho GTPases，对于理解软骨的病理状况，如软骨发育不良和骨关节炎，是至关重要的。在从 E15.5 小鼠分离的胫骨器官培养物中操纵肌动蛋白动力学会导致软骨内骨生长显著增强，并导致生长板结构发生特定变化。研究了从小鼠胚胎胫骨中分离的原代软骨细胞中基因表达的全局变化，这些细胞用细胞松弛素 D、jasplakinolide（肌动蛋白修饰剂）和 ROCK 抑制剂 Y27632 处理。细胞松弛素 D 引起的反应最为明显，并诱导了许多肥大软骨细胞分化的特征。微阵列数据分析的生物信息学分析和实时 PCR 和免疫组织化学的表达验证导致鉴定核受体视黄酸相关孤儿受体-α（Ror-α）为软骨细胞肥大的新型潜在调节剂。Ror-α 靶基因（Lpl、脂肪酸结合蛋白 4 [Fabp4]、Cd36 和 Krüppel 样因子 5 [Klf15]）的表达在软骨细胞肥大过程中以及细胞松弛素 D 和胆固醇依赖性诱导。胆固醇刺激 Ror-α 导致骨生长增加和细胞增大、变圆，表现型类似于软骨细胞肥大和细胞松弛素 D 诱导的变化，而洛伐他汀抑制胆固醇合成抑制细胞松弛素 D 诱导的骨生长。此外，我们还表明，在软骨特异性（Col2-Cre）Rac1 的小鼠模型中，失活导致肥大生长板中 Hif-1alpha（Rora 基因表达的调节剂）和 Ror-α（+）细胞增加。我们提供的证据表明，胆固醇信号通过增加 Ror-α 表达刺激软骨细胞肥大，并部分介导软骨对肌动蛋白动力学的反应。
