Gallery

Acini were grown to maturity in blank Matrigel before being transferred to a soft or stiff gel for an additional 7 days. The nuclei are shown in blue, and cells that have divided in the past 24 hours are shown in red. The laminin meshwork (shown in yellow) encapsulates cells in soft matrices. However, this protective barrier is disrupted after culture in high stiffness gels. DAPI (blue), laminin-332 (yellow), EdU (red); scale bar 20 μm.

Acini were grown to maturity in blank Matrigel for 14 days ("Day 0") before being transferred to a soft or stiff gel for an additional 7 days. Nuclei are shown in blue, cell membranes in green, and basement membrane components in yellow (laminin) or red (collagen IV). Mature acini transferred to soft gels maintain a thin, continuous sheet of basement membrane across their outer surface. Conversely, acini transferred to a stiff microenvironment often show loss of polarisation. DAPI (blue), CAAX (green), laminin-332 (yellow), collagen IV (red); scale bar 20 μm.

Mature acini transferred to stiff gels but treated with the BH3 mimetic ABT-737 had phenotypes more representative of acini in soft microenvironments. DAPI (blue), CAAX (green), laminin-332 (yellow), C3* (red); scale bar 20 μm.

Cells were either (a) untreated, (b) engineered to overexpress the anti-apoptotic Bcl2-family protein Bcl-xL, or (c) drug-treated with ROCK inhibitor (Y-27632), an MMP inhibitor (batimastat), blebbistatin (myosin II inhibitor), or ABT-737 (BH3 mimetic). DAPI (blue), CAAX (green), laminin-332 (yellow); scale bar 20 μm.

Fluorescent stacks can be segmented, labelled, and linked to track individual cell motion.

Acini were grown to maturity for 14 days in Matrigel, transferred to a stiff gel for 7 days, and then transferred to a soft gel for an additional 7 days. This double transfer largely reestablished basement membrane organisation but did not result in a decrease in acinar volume or cell number. DAPI (blue), CAAX (green), laminin-332 (yellow); scale bar 20 μm.

Various maximum intensity projections of acini embedded in their native environment. DAPI (blue), CAAX (green), various (red); scale bar 20 μm.

Mature acini transferred to stiff gels exhibit a loss of basement membrane organisation, accompanied by loss of organised architecture, and ultimate cellular disassociation. DAPI (blue), CAAX (green), laminin-332 (yellow); scale bar 20 μm.

Various individual slices of acini embedded in their native environment. DAPI (blue), CAAX (green), various (red); scale bar 20 μm.