Genetic models for lineage tracing in musculoskeletal development, injury, and healing

Abstract

Musculoskeletal development and later post-natal homeostasis are highly dynamic processes, marked by rapid structural and functional changes across very short periods of time. Adult anatomy and physiology are derived from pre-existing cellular and biochemical states. Consequently, these early developmental states guide and predict the future of the system as a whole. Tools have been developed to mark, trace, and follow specific cells and their progeny either from one developmental state to the next or between circumstances of health and disease. There are now many such technologies alongside a library of molecular markers which may be utilized in conjunction to allow for precise development of unique cell 'lineages'. In this review, we first describe the development of the musculoskeletal system beginning as an embryonic germ layer and at each of the key developmental stages that follow. We then discuss these structures in the context of adult tissues during homeostasis, injury, and repair. Special focus is given in each of these sections to the key genes involved which may serve as markers of lineage or later in post-natal tissues. We then finish with a technical assessment of lineage tracing and the techniques and technologies currently used to mark cells, tissues, and structures within the musculoskeletal system.

Keywords: Bone; Developmental biology; Fate mapping; Lineage tracing; Model; Skeletomuscular system.

Figure 1:

(A) Stages of fracture repair. (B) inducible cre strategy using an osx-cre-ERT2 transgenic mouse crossed with a Rosa-tdTomato mouse. After tamoxifen injection, osterix-expressing cells will express tdTomato, as shown here in post-operative day 3 section of a tibial injury. Immunofluorescence against osterix revealed co-labeling of osx and tomato-positive cells. (C) Non-inducible labeling strategy utilizing a LepR-cre and tdTomato mouse in which all LepR-expressing cells and their progeny will express tdTomato.

Figure 2:

Schematic depicting the difference between a (A) global knockout and a (B) conditional knockout strategy. (C) Inducible conditional knockout using the cre-ERT2 system, in which the cre recombinase is translocated into the nucleus only after administration of tamoxifen. (D) Cell ablation using an inducible conditional strategy in which the toxin receptor is expressed in cells after cre recombination excised the stop codon located adjacent to the coding sequence of the receptor. Once the toxin is delivered systemically, only cells that express the toxin receptor will be ablated. (E, F) Tetracycline-responsive elements either activate or inactivate the cre recombinase, resulting in inhibition or activation of downstream gene expression.

Figure 3:

(A) Schematic of different recombinase strategies that result in either (B) deletion, (C) inversion or (D) translocation.