Fig. 1: An immune-competent, microvascularized, human LOC device in a 96-well format.
From: An immune-competent lung-on-a-chip for modelling the human severe influenza infection response
a, Schematic of the 96-well plate format of the LOC device, showing the 2 × 4 grid formatted to fit on a bottomless 96-well plate, including a zoomed-in view of a single device and the cellular composition of each channel and the device seeding timeline, created using BioRender (https://BioRender.com/7wk1dzr). b,c, Representative images of the airway epithelium in the LOC device, showing DAPI in yellow and ciliated cell marker TUBB4A in magenta (scale bars, 100 µm), with inset showing TUBB4A signal situated at the top surface of the cell (scale bar, 10 µm) (b), as well as goblet cell marker Mucin5AC (magenta) and actin (yellow) (c). Scale bars, 11 µm. d, ECM staining of collagen IV (green) surrounding the formed vessels (CD31, yellow). Scale bars, 160 µm. e, Additional ECM staining of fibronectin (green) in the interstitial space along with CD31 (yellow) staining of the vessels. Scale bars, 160 µm. f, Incorporation of labelled interstitial fibroblasts (green) with vasculature (CD31, yellow). Scale bars, 160 µm. g, Incorporation of IMs (green) with vasculature (CD31, yellow). Scale bars, 80 µm. h, Representative images of the epithelium (ZO-1, yellow) with AMs (CD68, magenta). Scale bars, 100 µm. i, The microvascular network perfused with 70 kDa FITC-dextran. Scale bar, 500 µm. j, Timeframe images (Δt = 100 ms) of labelled PBMCs (red, arrows) entering the microvascular network through open lumens between ports of the device. k, Labelled PBMCs (cyan) perfused through the microvascular network (yellow), showing PBMCs entering through an open lumen. Scale bar, x-direction 160 µm, z-direction 50 µm. l, Bright-field image of whole blood perfused through the microvasculature. Scale bar, 160 µm. m, 10 kDa FITC-dextran permeability of the epithelium in LOC, Mϕ-LOC and IC-LOC devices, quantified by the airway concentration of FITC-dextran at the assay endpoint (2 h incubation). The dotted line represents the starting concentration of 250 μg ml−1. N = 3 independent devices, with statistical significance determined via ordinary one-way ANOVA followed by Tukey’s multiple comparisons test. NS, not signficant. Data are presented as mean values ± s.d. n, A graph depicting the major cell types observed in the IC-LOC and the human lung, as determined via scRNA-seq.
