Fibroblast elongation and dendritic extensions in constrained versus unconstrained microtissues

Area of Science:

• Cell Biology
• Biophysics
• Developmental Biology

Background:

• Cytoskeletal tension is crucial for biological processes like embryogenesis and in vitro cell sorting.
• In vitro, cell-derived tension can lead to microtissue failure.
• Directed self-assembly in micro-molds can create self-constraining microtissues.

Purpose of the Study:

• To investigate microtissue failure mechanisms driven by cell-generated tension.
• To analyze cell movement and morphology during constrained self-assembly.
• To explore scaffold-free models for studying fibroblast behavior.

Main Methods:

• Normal human fibroblasts (NHFs) were subjected to directed self-assembly in micro-molds.
• Green fluorescent protein-positive (GFP+) cells were used to track cell behavior.
• Pharmacologic interventions were employed to identify molecular pathways involved in failure.

Main Results:

• Constrained microtissues narrowed, thinned, and failed at their midpoints due to cellular contraction.
• Cells formed dendritic extensions that retracted as they elongated significantly before failure.
• Coordinated cell motility was observed over large distances within the microtissues.
• Myosin II and Rho kinase pathways were identified as critical for microtissue failure.

Conclusions:

• Microtissue failure under cell-derived tension is a coordinated process dependent on myosin II and Rho kinase.
• Inhibition of failure leads to altered cell morphology, characterized by shorter cells and increased extensions.
• Directed self-assembly with GFP+ cells provides a valuable scaffold-free model for studying fibroblast-populated collagen gels (FPCGs).