Microfluidic and Lab-on-a-Chip Systems for Cutaneous Wound Healing Studies

Abstract

Cutaneous wound healing is a complex, multi-stage process involving direct and indirect cell communication events with the aim of efficiently restoring the barrier function of the skin. One key aspect in cutaneous wound healing is associated with cell movement and migration into the physically, chemically, and biologically injured area, resulting in wound closure. Understanding the conditions under which cell migration is impaired and elucidating the cellular and molecular mechanisms that improve healing dynamics are therefore crucial in devising novel therapeutic strategies to elevate patient suffering, reduce scaring, and eliminate chronic wounds. Following the global trend towards the automation, miniaturization, and integration of cell-based assays into microphysiological systems, conventional wound healing assays such as the scratch assay and cell exclusion assay have recently been translated and improved using microfluidics and lab-on-a-chip technologies. These miniaturized cell analysis systems allow for precise spatial and temporal control over a range of dynamic microenvironmental factors including shear stress, biochemical and oxygen gradients to create more reliable in vitro models that resemble the in vivo microenvironment of a wound more closely on a molecular, cellular, and tissue level. The current review provides (a) an overview on the main molecular and cellular processes that take place during wound healing, (b) a brief introduction into conventional in vitro wound healing assays, and (c) a perspective on future cutaneous and vascular wound healing research using microfluidic technology.

Keywords: cell migration; cutaneous wound healing; lab-on-a-chip; microfluidics; microvasculature; skin; wound healing assay.

Figure 1

(A) Overview of the cellular processes during the three wound healing stages. (B) Schematic representation of pathways involved in wound healing, including receptors for fibroblast growth factors (FGFs), epidermal growth factors (EGFs), and transforming growth factor β (TGF-β).

Figure 2

In vitro wound healing assays. (A) Scratch assays most frequently use pipette tips to manually scratch and remove cells from a cell monolayer. (B) Cell exclusion assays block a wound area before cell adhesion with a physical barrier insert removed after the establishment of the cell layer integrity to create a well-defined wound.

Figure 3

Overview of state-of-the-art microfluidic wound healing assays, including cell-depletion (physical or enzymatic) and physical cell exclusion approaches in microfluidic channels. (A) the mechanical cell depletion approach, (B) the enzymatic cell depletion approach, and (C) the cell exclusion approach. In each approach cross-section, and top views of the microchannel during and after wounding are illustrated.

Figure 4

Microfluidic wound healing assays based on physical cell exclusion. (A–C) Pillar-based microfluidic wound healing to analyze the influence of (B) EGF concentration on wound closure and (C) the number of proliferative cells. (* p < 0.05 vs. control). Adapted with permission from ref. [28]. 2021, Elsevier.

Figure 5

Enzymatic microfluidic wound healing assays based on the laminar flow patterning of fluids. (A–C) Influence of the on-chip nanopattern direction on wound healing speed using enzymatic depletion of a central cell-free area using trypsin. Adapted with permission from [48]. (D,E) Influence of flow direction, shear, and VEGF on (E) endothelial migration rate and (D) wound healing directionality. (* Significant increase compared with control values (Student’s t-test, p < 0.05). # Significant increase compared with 100 ng/mL VEGF165 treatment (Student’s t-test, p < 0.05). Adapted with permission from ref. [49]. 2021, Elsevier.

Figure 6

(A) Wound-healing lab-on-a-chip system with four individual pneumatic and fluidic cell chambers. (B) Pneumatic actuation of a flexible membrane within the microfluidic device. (C) Direct comparison of a conventional scratch assay’s reproducibility and precision compared to pneumatically-actuated, automated physical cell depletion method for endothelial cells (HUVECs). Adapted from [53] with permission of The Royal Society of Chemistry. (D) Effect of growth factor bFGF and inhibitory agents mitomycin C (MMC) and MEK-inhibitor U0126 on dermal fibroblast migration and proliferation dynamics. (ns, non-significant; ** p  <  0.01; *** p  <  0.001; **** p  <  0.0001). Adapted with permission from ref. [54]. 2021, Elsevier.