Fig. 4: Microdroplet mechanical anisotropy enables cell invasion.

a F-actin (grey) and DAPI (blue) in OVCAR8 cells embedded in collagen or collagen-based organo-hydrogels (OHG) for 24 h (representative images from n = 3 gels). Microdroplet locations are indicated with an asterisk. b BODIPY (green), F-actin (grey) and nuclei (blue) staining of OVCAR8 cells embedded in collagen-based OHG (left; n = 3 gels) or BODIPY (green), mRFP (red) staining of mRFP-OVCAR8 cells invading into peritoneal adipose tissue (right; n = 3 tissues from 1 donor). Arrowheads indicate oil microdroplet (left) and adipocyte (right) deformations at the contact points with cells. c Quantification of OVCAR8 organotypic invasion depth into peritoneal adipose tissues after 7 d (n = 3 tissues from 1 donor). Rhombuses indicate the average. d Schematic representation of the Sylgard 184 PDMS-based OGH. e F-actin (grey) and nuclei (red) staining of OVCAR8 spheroids embedded for 7 d in collagen-based OHGs prepared with PDMS microdroplets of varying stiffness (representative images from n = 10 spheroids). f OVCAR8 spheroid area after 7 d in collagen-based OHGs with PDMS microdroplets prepared with varying crosslinker percentage, relative to average area in 2 wt% microbead OHGs (n = 10 spheroids). g Schematic representation of the norbornene-functionalised hyaluronic acid (NB-HA) OGH with norbornene-functionalised bovine serum albumin (NB-BSA) used for RGD presentation at the microdroplet surface. h F-actin (grey) and nuclei (red) of OVCAR8 spheroids after 7 d in NB-HA OHG with localised presentation of RGD (left). Relative OVCAR8 spheroid area in NB-HA OHG with localised RGD presentation after 7 d in culture (n = 11 spheroids; top right). BODIPY (green), nuclei (red), and F-actin (grey) staining of OVCAR8 cells in NB-HA OHG presenting RGD on the microdroplet surface (bottom right). Arrowhead indicates a cell spreading at the microdroplet-HA interface. i Schematic representation of the system to modulate microdroplet interfacial mechanics. j F-actin (grey) and nuclei (red) of OVCAR8 spheroids after 7 d in NB-HA OHGs (microdroplet-restricted RGD presentation) with varying microdroplet interfacial modulus (representative images from n = 9 spheroids). k OVCAR8 spheroid area in NB-HA OHGs with microdroplet-presenting RGD and varying protein nanosheet interfacial mechanics after 7 d in culture, relative to BSA-only control (n = 9 spheroids). For the data in (c and h), a two-sided unpaired t test was performed. One-way analysis of variance (ANOVA) with Tukey’s correction for multiple comparisons was performed for the data in (f and k). Scale bars, 25 μm (a, b), 50 μm (h, bottom right), 200 μm (e, h left, j). Source data are provided as a Source Data file.