Post-injury hydraulic fracturing drives fissure formation in the zebrafish basal epidermal cell layer

Abstract

SummaryThe skin epithelium acts as the barrier between an organism’s internal and external environments. In zebrafish and other freshwater organisms, this barrier function requires withstanding a large osmotic pressure differential. Wounds breach this epithelium, causing a large disruption to the tissue microenvironment due to the mixing of isotonic interstitial fluid with the external hypotonic fresh water. Here we show that, following acute injury, the larval zebrafish epidermis undergoes a dramatic fissuring process that resembles hydraulic fracturing, driven by the influx of external fluid. The fissuring starts in the basal epidermal layer nearest to the wound, and then propagates at a constant rate through the tissue spanning over one hundred micrometers; during this process the outermost superficial epidermal layer remains intact. Fissuring is completely inhibited when larvae are wounded in an isotonic external media, suggesting that osmotic pressure gradients drive fissure. Additionally, fissuring partially depends on myosin II activity, as its inhibition reduces fissure propagation away from the wound. During and after fissuring, the basal layer forms large macropinosomes (with cross-sectional areas ranging from 1-10 µm2), presumably to clear the excess fluid. We conclude that excess external fluid entry through the wound and subsequent closure of the wound through actomyosin purse string contraction in the superficial cell layer causes fluid pressure buildup in the extracellular space of the zebrafish epidermis. This excess fluid pressure causes tissue to fissure, and eventually the fluid is cleared through macropinocytosis.

Confined keratocytes mimic in vivo migration and reveal volume‐speed relationship

“…Indeed, individual keratocytes swell to 50% of their pre‐wounding volume following normal wounding in freshwater (Kennard & Theriot, 2020), which could significantly boost speed if the volume‐speed relationship holds in the swelling regime. Hypotonic shock additionally leads to a massive fluid influx that increases the space between basal cells and the overlying periderm (Kennard, Sathe, Labuz, Prinz, & Theriot, 2022). This expansion of the epidermis may release some of the 2D constraints on keratocytes and provide additional speed benefits.…”

“…The following previously generated transgenic zebrafish lines were used: TgBAC(ΔNp63:Gal4) la213 (Rasmussen et al, 2015), Tg(UAS:Life-Act-EGFP) mu271 (Helker et al, 2013), Tg(UAS:mCherry) (Kennard et al, 2022).…”

Confined keratocytes mimic in vivo migration and reveal volume-speed relationship

“…Indeed, individual keratocytes swell to 50% of their pre-wounding volume following normal wounding in freshwater (Kennard & Theriot, 2020), which could significantly boost speed if the volume-speed relationship holds in the swelling regime. Hypotonic shock additionally leads to a massive fluid influx that increases the space between basal cells and the overlying periderm (Kennard et al, 2022). This expansion of the epidermis may release some of the 2D constraints on keratocytes and provide additional speed benefits.…”

“…The following previously generated transgenic zebrafish lines were used: TgBAC(ΔNp63:Gal4) la213 (Rasmussen et al, 2015), Tg(UAS:LifeAct-EGFP) mu271 (Helker et al, 2013), Tg(UAS:mCherry) (Kennard et al, 2022).…”

Inverse blebs operate as hydraulic pumps during mouse blastocyst formation

“…During embryonic development or the formation of organoids, relocation of IF is suspected to influence tissue jamming [9][10][11] or topological transitions when fluid lumens form 12 . In some of the most striking relocations of IF, pressurized fluid can even fracture tissues 3,13,14 . In large fluid compartments, such as lumens, the movement of IF can be controlled by specialized cell protrusions called cilia 15 , but in packed tissues we are only beginning to understand how cells control the fluid surrounding them 16 .…”