Ha Tin-Yan, Chan See-Wing, Wang Zhangting, Law Patrick Wai Nok, Miu Kai-Kei, Lu Gang, Chan Wai-Yee
School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, Hong Kong SAR, China.
CUHK-SDU Joint Laboratory on Reproductive Genetics, School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong SAR, China.
Cell Mol Life Sci. 2025 Mar 2;82(1):99. doi: 10.1007/s00018-025-05622-y.
Campomelic Dysplasia (CD) is a rare congenital disease caused by haploinsufficiency (HI) in SOX9. Patients with CD typically present with skeletal abnormalities and 75% of them have sex reversal. In this study, we use CRISPR/Cas9 to generate a human induced pluripotent stem cell (hiPSC) model from a heathy male donor, based on a previously reported SOX9 splice site mutation in a CD patients. This hiPSCs-derived chondrocytes from heterozygotes (HT) and homozygotes (HM) SOX9 mutation carriers showed significant defects in chondrogenesis. Bulk RNA profiling revealed that the BMP-SMAD signaling pathway, ribosome-related, and chromosome segregation-related gene sets were altered in the HT chondrocytes. The profile also showed significant noggin upregulation in CD chondrocytes, with ChIP-qPCR confirming that SOX9 binds to the distal regulatory element of noggin. This suggests SOX9 plays a feedback role in the BMP signaling pathway by modulating noggin expression rather than acting solely as a downstream regulator. This provides further insights into its dosage sensitivity in chondrogenesis. Overexpression of SOX9 showed promising results with improved sulfated glycosaminoglycans (GAGs) aggregation and COL2A1 expression following differentiation. We hope this finding could provide a better understanding of the dosage-dependent role of SOX9 in chondrogenesis and contribute to the development of improved therapeutic targets for CD patients.
弯肢发育不良（CD）是一种由SOX9单倍剂量不足（HI）引起的罕见先天性疾病。CD患者通常表现出骨骼异常，其中75%的患者存在性反转。在本研究中，我们基于先前报道的一名CD患者的SOX9剪接位点突变，利用CRISPR/Cas9从一名健康男性供体生成了人诱导多能干细胞（hiPSC）模型。来自杂合子（HT）和纯合子（HM）SOX9突变携带者的这种hiPSC衍生软骨细胞在软骨生成方面表现出明显缺陷。大量RNA分析显示，HT软骨细胞中骨形态发生蛋白（BMP）-SMAD信号通路、核糖体相关和染色体分离相关基因集发生了改变。分析还显示CD软骨细胞中头蛋白明显上调，染色质免疫沉淀定量聚合酶链反应（ChIP-qPCR）证实SOX9与头蛋白的远端调控元件结合。这表明SOX9通过调节头蛋白表达在BMP信号通路中发挥反馈作用，而不是仅仅作为下游调节因子。这为其在软骨生成中的剂量敏感性提供了进一步的见解。SOX9的过表达显示出有希望的结果，分化后硫酸化糖胺聚糖（GAG）聚集和II型胶原蛋白（COL2A1）表达得到改善。我们希望这一发现能够更好地理解SOX9在软骨生成中的剂量依赖性作用，并有助于为CD患者开发更好的治疗靶点。
