Ligament injury in adult zebrafish triggers ECM remodeling and cell dedifferentiation for scar-free regeneration

Abstract

After traumatic injury, healing of mammalian ligaments is typically associated with fibrotic scarring as opposed to scar-free regeneration. In contrast, here we show that the ligament supporting the jaw joint of adult zebrafish is capable of rapid and complete scar-free healing. Following surgical transection of the jaw joint ligament, we observe breakdown of ligament tissue adjacent to the cut sites, expansion of mesenchymal tissue within the wound site, and then remodeling of extracellular matrix (ECM) to a normal ligament morphology. Lineage tracing of mature ligamentocytes following transection shows that they dedifferentiate, undergo cell cycle re-entry, and contribute to the regenerated ligament. Single-cell RNA sequencing of the regenerating ligament reveals dynamic expression of ECM genes in neural-crest-derived mesenchymal cells, as well as diverse immune cells expressing the endopeptidase-encoding gene legumain. Analysis of legumain mutant zebrafish shows a requirement for early ECM remodeling and efficient ligament regeneration. Our study establishes a new model of adult scar-free ligament regeneration and highlights roles of immune-mesenchyme cross-talk in ECM remodeling that initiates regeneration.

Single-cell analysis reveals distinct fibroblast plasticity during tenocyte regeneration in zebrafish

“…Consistently, uninjured tenocytes have been shown to become activated and give rise to new tenocytes after maxillary superficial tendon transection in adult zebrafish jaw ( 76 ). Similarly, lineage tracing shows that mature scxa + ligamentocytes contribute to the complete regeneration of jaw joint ligament following transection in adult zebrafish ( 78 ). In contrast to these studies, we find that unablated tenocytes show no active response to laser-mediated tendon injury.…”

“…Tendons are vital connective tissues connecting muscle to bone and tendon injury is often associated with devastating physical disability and reduced mobility (75). Unlike mammals, we and others find that zebrafish can completely regenerate their tendon structure after injury (76)(77)(78). Using this powerful zebrafish model, we idenLfy sclerotome-derived perivascular/intersLLal fibroblasts as the cellular source of newly regenerated tenocytes upon tendon injury.…”

Programmable promoter editing for precise control of transgene expression

“…DIAL can potentially integrate into the abundance of recombinase-based tools for lineage tracing to allow increase the setpoint of transgenes at specific developmental windows and within specific subpopulations. [67][68][69][70][71] While DIAL provides a number of features and insights into gene regulation, there remain limitations and opportunities to expand the capabilities. At high expression of the synthetic transcription factor, DIAL outputs do not change.…”

“…Given the ubiquity of recombinase tools in model organisms and cell lines, DIAL may integrate with diverse recombinase-based tools for lineage tracing. [72][73][74][75][76] We show that Cre activity can be instigated by orthogonal control systems, including external user-defined cues like modRNA and small molecules, opening opportunities for changing setpoints at a specific timepoint. 58,77 While the DIAL framework provides many insights on engineering synthetic promoter systems, there remain limitations and opportunities to expand the capabilities.…”