Modelling collagen fibril self-assembly from extracellular medium in embryonic tendon

Abstract

Collagen is a key structural component of multicellular organisms and is arranged in a highly organised manner. In structural tissues such as tendons, collagen forms bundles of parallels fibres between cells, which appear within a 24 hour window between E13.5 and E14.5 during mouse embryonic development. Current models assume that the organised structure of collagen requires cellular control, whereby cells actively lay down collagen fibrils from cell surfaces. However, such models appear incompatible with the time- and length-scales of fibril formation. We propose a phase-transition model to account for the rapid development of ordered fibrils in tendon tissues, reducing reliance on active cellular processes. We develop phase-field crystal simulations of collagen fibrillogenesis in solution domains derived from electron micrograph images of inter-cellular spaces in embryonic tendon and compare results qualitatively and quantitatively to observed patterns of fibril formation. To test the prediction of this phase-transition model that free protomeric collagen should exist in the intercellular spaces prior to the formation of observable fibrils, we use laser-capture microdissection, coupled with mass spectrometry, which demonstrates steadily increasing free collagen in intercellular spaces up to E13.5, followed by a rapid reduction of free collagen that coincides with the appearance of less soluble collagen fibrils. The model and measurements together provide evidence for extracellular self-assembly of collagen fibrils in embryonic mouse tendon, supporting a new model for collagen fibrillogenesis during embryonic development.

Molecular analysis of the extracellular microenvironment: from form to function

“…Modeling data that included electron microscopy and laser capture microdissection coupled with proteomics suggested that not all assembly is dependent on cellular processes and may be driven by mechanical loading [32]. In this model, free triple helical segments were observed secreted early in development and self-assembled through processes of mechanical loading, and aggregation in the absence of cells [32]. As part of extracellular assembly, crosslinks are formed between collagen types as well as other ECM proteins.…”

“…Cellular processing involves, in some ways, the presence of fibropositors or membrane channels that form template spaces for organizing individual or groups of fibrils [31]. Modeling data that included electron microscopy and laser capture microdissection coupled with proteomics suggested that not all assembly is dependent on cellular processes and may be driven by mechanical loading [32]. In this model, free triple helical segments were observed secreted early in development and self-assembled through processes of mechanical loading, and aggregation in the absence of cells [32].…”

“…Current studies leverage using multiplexed markers of disease to target specific regions for microproteomics [72,150]. This is a powerful approach for comprehensive proteomic assessment of the ECM, especially when paired with other modalities such as SEM or nonlinear birefringent techniques [32]. An advantage is the proteomic depth achieved by microproteomics when compared to other tissue imaging proteomics.…”