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Transcriptomic trajectory of the tendon and fibrocartilage fate switch [bulk RNA seq]

ABSTRACT: Transcriptomic trajectory of the tendon and fibrocartilage fate switch [bulk RNA seq]

PROVIDER: PRJNA646184 | ENA |

REPOSITORIES: ENA

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Transcriptomic trajectory of the tendon and fibrocartilage fate switch

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Transcriptomic trajectory of the tendon and fibrocartilage fate switch [bulk RNA seq]

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Transcriptional profiling of mESC-derived tendon and fibrocartilage cell fate switch [scRNA-seq]

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Transcriptomic trajectory of the tendon and fibrocartilage fate switch

Project description:This SuperSeries is composed of the SubSeries listed below.

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RNA-seq analysis of Scx-lineage cell depletion to investigate tendon cell functions during flexor tendon healing

Project description:Little is understood about the roles of tendon cells during flexor tendon healing. To better understand tendon cell functions, the Scx-Cre mouse was crossed to the DTR mouse model to facilitate scleraxis lineage cell depletion prior to acute flexor tendon injury and repair. WT (cre-) and experimental (cre+) mice underwent complete transection and repair of the flexor digitorum longus tendon. Repaired tendons were harvested at 14 and 28 days post-repair for bulk RNA-Seq analysis to examine possible mechanisms driving differential healing due to Scx lineage cell depletion.

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Project description:The musculoskeletal system is integrated by tendons that are characterized by expression of a functionally important transcription factor Scleraxis (Scx). To date, there has been no solid culture system for tenogenic differentiation. Here we developed the tenocyte induction method using induced pluripotent stem cells established from ScxGFP transgenic mice by monitoring fluorescence that reflects a series of dynamic differentiation process. Among several developmentally relevant factors, TGF-2 was the most potent inducer for differentiation of tenocytes expressing Tenomodulin, a mature differentiation marker. Single-cell RNA sequencing (scRNA-seq) revealed 11 distinct clusters including mature tenocyte population and tenogenic differentiation trajectory, which well recapitulates in vivo developmental process. Dataset of scRNA-seq and transcriptome of Scx deficient tendon enabled identification of Scx-related genes in which tendon-related genes are enriched. Collectively, this study leads to a new opportunity for tendon research and provides further insight into mechanistic understanding of the differentiation pathway to a tenogenic fate.

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Divergent cell differentiation trajectory in normal and heterotopic ossified soft connective tissues

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Electrospun scaffold micro-architecture induces an activated transcriptional phenotype within tendon fibroblasts. Bulk RNA-seq data.

Project description:Biomaterial augmentation of surgically repaired rotator cuff tendon tears aims to improve the high failure rates (~40%) of traditional repairs. Biomaterials that can alter cellular phenotypes through the provision of microscale topographical cues are now under development. We aimed to systematically evaluate the effect of topographic architecture on the cellular phenotype of fibroblasts from healthy and diseased tendons. Electrospun polydioxanone scaffolds with fiber diameters ranging from 300 to 4000 nm, in either a highly aligned or random configuration, were produced. Healthy tendon fibroblasts cultured for 7 days on scaffolds with highly aligned fibers demonstrated a distinctive elongated morphology, whilst those cultured on randomly configured fibers demonstrated a flattened and spread morphology. The effect of scaffold micro-architecture on the transcriptome of both healthy and diseased tendon fibroblasts was assessed with bulk RNA-seq. Both healthy (n=3) and diseased tendon cells (n=3) demonstrated a similar transcriptional response to architectural variants. Gene set enrichment analysis revealed that large diameter (≥ 2000 nm) aligned scaffolds induced an upregulation of genes involved in cellular replication and a downregulation of genes defining inflammatory responses and cell adhesion. Similarly, PDPN and CD248, markers of inflammatory or ‘activated’ fibroblasts, were downregulated during culture of both healthy and diseased fibroblasts on aligned scaffolds with large (≥ 2000 nm) fiber diameters. In conclusion scaffold architectures resembling that of disordered type III collagen, typically present during the earlier phases of wound healing, resulted in tendon fibroblast activation. Conversely, scaffolds mimicking aligned diameter collagen I fibrils, present during tissue remodelling, did not activate tendon derived fibroblasts. This has implications for the design of scaffolds used during rotator cuff repair augmentation.

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Depletion of Scleraxis-lineage cells during tendon healing transiently impairs multi-scale restoration of tendon structure during early healing

Project description:Tendons are composed of a heterogeneous cell environment, with Scleraxis-lineage (ScxLin) cells being the predominant population. Although ScxLin cells are required for maintenance of tendon homeostasis, their functions during tendon healing are unknown. To this end, we first characterized the spatiotemporal dynamics of ScxLin cells during tendon healing, and identified that the overall ScxLin pool continuously expands up to early remodeling healing phase. To better define the function of ScxLin cells during the late proliferative phase of healing, we inducibly depleted ScxLin cells from day 14-18 post-surgery using the Scx-Cre; Rosa-DTR mouse model, with local administration of diphtheria toxin inducing apoptosis of ScxLin cells in the healing tendon. At D28 post-surgery, ScxLin cell depleted tendons (DTRScxLin) had substantial impairments in structure and function, relative to WT, demonstrating the importance of ScxLin cells during tendon healing. Next, bulk RNAseq was utilized to identify the underlying mechanisms that were impaired with depletion and revealed that ScxLin depletion induced molecular and morphological stagnation of the healing process at D28. However, this stagnation was transient, such that by D56 tendon mechanics in DTRScxLin were not significantly different than wildtype repairs. Collectively, these data offer fundamental knowledge on the dynamics and roles of ScxLin cells during tendon healing.

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