Both the WNT/β-catenin and hedgehog signaling pathways are important in the regulation of limb development, chondrocyte differentiation, and degeneration of articular cartilage in osteoarthritis (OA). It is not clear how these signaling pathways interact in interzone cell differentiation and synovial joint morphogenesis. Here, we determined that constitutive activation of hedgehog signaling specifically within interzone cells induces joint morphological changes by selectively inhibiting β-catenin–induced Fgf18 expression. Stabilization of β-catenin or treatment with FGF18 rescued hedgehog-induced phenotypes. Hedgehog signaling induced expression of a dominant negative isoform of TCF7L2 (dnTCF7L2) in interzone progeny, which may account for the selective regulation of β-catenin target genes observed. Knockdown of TCF7L2 isoforms in mouse chondrocytes rescued hedgehog signaling–induced Fgf18 downregulation, while overexpression of the human dnTCF7L2 orthologue (dnTCF4) in human chondrocytes promoted the expression of catabolic enzymes associated with OA. Similarly, expression of dnTCF4 in human chondrocytes positively correlated with the aggrecanase ADAMTS4. Consistent with our developmental findings, activation of β-catenin also attenuated hedgehog-induced or surgically induced articular cartilage degeneration in mouse models of OA. Thus, our results demonstrate that hedgehog inhibits selective β-catenin target gene expression to direct interzone progeny fates and articular cartilage development and disease. Moreover, agents that increase β-catenin activity have the potential to therapeutically attenuate articular cartilage degeneration as part of OA.

To address large gaps in our understanding of the molecular regulation of articular and growth plate cartilage development in humans, we used our directed differentiation approach to generate these distinct cartilage tissues from human embryonic stem cells. The resulting transcriptomic profiles of hESC-derived articular and growth plate chondrocytes were similar to fetal epiphyseal and growth plate chondrocytes, with respect to genes both known and previously unknown to cartilage biology. With the goal to characterize the regulatory landscapes accompanying these respective transcriptomes, we mapped chromatin accessibility in hESC-derived chondrocyte lineages, and mouse embryonic chondrocytes, using ATAC-sequencing. Integration of the expression dataset with the differentially accessible genomic regions revealed lineage-specific gene regulatory networks. We validated functional interactions of two transcription factors (TFs) (RUNX2 in growth plate chondrocytes and RELA in articular chondrocytes) with their predicted genomic targets. The maps we provide thus represent a framework for probing regulatory interactions governing chondrocyte differentiation. This work constitutes a substantial step towards comprehensive and comparative molecular characterizations of distinct chondrogenic lineages and sheds new light on human cartilage development and biology.

Objectives Progressive pseudorheumatoid arthropathy of childhood (PPAC), caused by deficiency of WNT1 inducible signalling pathway protein 3 (WISP3), has been challenging to study because no animal model of the disease exists and cartilage recovered from affected patients is indistinguishable from common end-stage osteoarthritis. Therefore, to gain insights into why precocious articular cartilage failure occurs in this disease, we made in vitro derived articular cartilage using isogenic WISP3-deficient and WISP3-sufficient human pluripotent stem cells (hPSCs). Methods We generated articular cartilage-like tissues from induced-(i) PSCs from two patients with PPAC and one wild-type human embryonic stem cell line in which we knocked out WISP3. We compared these tissues to in vitro-derived articular cartilage tissues from two isogenic WISP3-sufficient control lines using histology, bulk RNA sequencing, single cell RNA sequencing and in situ hybridisation. Results WISP3-deficient and WISP3-sufficient hPSCs both differentiated into articular cartilage-like tissues that appeared histologically similar. However, the transcriptomes of WISP3-deficient tissues differed significantly from WISP3-sufficient tissues and pointed to increased TGFβ, TNFα/NFκB, and IL-2/STAT5 signalling and decreased oxidative phosphorylation. Single cell sequencing and in situ hybridisation revealed that WISP3-deficient cartilage contained a significantly higher fraction (~4 fold increase, p<0.001) of superficial zone chondrocytes compared with deeper zone chondrocytes than did WISP3-sufficient cartilage. Conclusions WISP3-deficient and WISP3-sufficient hPSCs can be differentiated into articular cartilage-like tissues, but these tissues differ in their transcriptomes and in the relative abundances of chondrocyte subtypes they contain. These findings provide important starting points for in vivo studies when an animal model of PPAC or presymptomatic patient-derived articular cartilage becomes available.

Over 100 scientists with common interests in human development, disease and regeneration gathered in late September 2016 for The Company of Biologists' second ‘From Stem Cells to Human Development’ meeting held in historic Southbridge. In this Meeting Review, we highlight some of the exciting new findings that were presented, and discuss emerging themes and convergences in human development and disease that arose during these discussions.