Dept. of Orthopaedic Surgery, Washington University in Saint Louis, St. Louis, MO, 63110, USA.
Shriners Hospitals for Children-St. Louis, St. Louis, MO, 63110, USA.
Nat Commun. 2021 Jan 13;12(1):362. doi: 10.1038/s41467-020-20598-y.
The therapeutic application of human induced pluripotent stem cells (hiPSCs) for cartilage regeneration is largely hindered by the low yield of chondrocytes accompanied by unpredictable and heterogeneous off-target differentiation of cells during chondrogenesis. Here, we combine bulk RNA sequencing, single cell RNA sequencing, and bioinformatic analyses, including weighted gene co-expression analysis (WGCNA), to investigate the gene regulatory networks regulating hiPSC differentiation under chondrogenic conditions. We identify specific WNTs and MITF as hub genes governing the generation of off-target differentiation into neural cells and melanocytes during hiPSC chondrogenesis. With heterocellular signaling models, we further show that WNT signaling produced by off-target cells is responsible for inducing chondrocyte hypertrophy. By targeting WNTs and MITF, we eliminate these cell lineages, significantly enhancing the yield and homogeneity of hiPSC-derived chondrocytes. Collectively, our findings identify the trajectories and molecular mechanisms governing cell fate decision in hiPSC chondrogenesis, as well as dynamic transcriptome profiles orchestrating chondrocyte proliferation and differentiation.
人诱导多能干细胞（hiPSCs）在软骨再生中的治疗应用在很大程度上受到以下因素的阻碍：软骨细胞产量低，以及在软骨发生过程中细胞向异质的非靶细胞分化具有不可预测性。在这里，我们结合了批量 RNA 测序、单细胞 RNA 测序和包括加权基因共表达网络分析（WGCNA）在内的生物信息学分析，以研究调控 hiPSC 在软骨形成条件下分化的基因调控网络。我们确定了特定的 WNTs 和 MITF 作为调控 hiPSC 软骨发生过程中向神经细胞和黑素细胞产生非靶细胞分化的枢纽基因。通过异细胞信号模型，我们进一步表明，由非靶细胞产生的 WNT 信号负责诱导软骨细胞肥大。通过靶向 WNTs 和 MITF，我们消除了这些细胞谱系，显著提高了 hiPSC 来源的软骨细胞的产量和均一性。总的来说，我们的研究结果确定了 hiPSC 软骨生成中细胞命运决定的轨迹和分子机制，以及协调软骨细胞增殖和分化的动态转录组图谱。
