Musculoskeletal Disease Center, Veterans Affairs Loma Linda Healthcare System, 11201 Benton Street, Loma Linda, CA 92357, USA.
Department of Medicine, Loma Linda University, Loma Linda, CA 92354, USA.
Sci Rep. 2017 Mar 28;7:45408. doi: 10.1038/srep45408.
Based on our findings that PHD2 is a negative regulator of chondrocyte differentiation and that hypoxia signaling is implicated in the pathogenesis of osteoarthritis, we investigated the consequence of disruption of the Phd2 gene in chondrocytes on the articular cartilage phenotype in mice. Immunohistochemistry detected high expression of PHD2 in the superficial zone (SZ), while PHD3 and HIF-1α (target of PHD2) are mainly expressed in the middle-deep zone (MDZ). Conditional deletion of the Phd2 gene (cKO) in chondrocytes accelerated the transition of progenitors to hypertrophic (differentiating) chondrocytes as revealed by reduced SZ thickness, and increased MDZ thickness, as well as increased chondrocyte hypertrophy. Immunohistochemistry further revealed decreased levels of progenitor markers but increased levels of hypertrophy markers in the articular cartilage of the cKO mice. Treatment of primary articular chondrocytes, in vitro, with IOX2, a specific inhibitor of PHD2, promoted articular chondrocyte differentiation. Knockdown of Hif-1α expression in primary articular chondrocytes using lentiviral vectors containing Hif-1α shRNA resulted in reduced expression levels of Vegf, Glut1, Pgk1, and Col10 compared to control shRNA. We conclude that Phd2 is a key regulator of articular cartilage development that acts by inhibiting the differentiation of articular cartilage progenitors via modulating HIF-1α signaling.
基于我们的发现，PHD2 是软骨细胞分化的负调节剂，并且缺氧信号通路与骨关节炎的发病机制有关，我们研究了软骨细胞中 Phd2 基因的破坏对小鼠关节软骨表型的影响。免疫组化检测到 PHD2 在浅层区（SZ）高表达，而 PHD3 和 HIF-1α（PHD2 的靶标）主要在中深层区（MDZ）表达。软骨细胞中 Phd2 基因的条件缺失（cKO）导致祖细胞向肥大（分化）软骨细胞的转化加速，表现为 SZ 厚度减少，MDZ 厚度增加，以及软骨细胞肥大增加。免疫组化进一步显示，cKO 小鼠关节软骨中的祖细胞标志物水平降低，但肥大标志物水平升高。体外培养的原代关节软骨细胞用 IOX2（PHD2 的特异性抑制剂）处理可促进关节软骨细胞分化。使用含有 Hif-1α shRNA 的慢病毒载体敲低原代关节软骨细胞中的 Hif-1α 表达，与对照 shRNA 相比，Vegf、Glut1、Pgk1 和 Col10 的表达水平降低。我们得出结论，Phd2 是关节软骨发育的关键调节剂，通过调节 HIF-1α 信号通路抑制关节软骨祖细胞的分化来发挥作用。
