Machine learning reveals mesenchymal breast carcinoma cell adaptation in response to matrix stiffness.

Epithelial-mesenchymal transition (EMT) and its reverse process, mesenchymal-epithelial transition (MET), are believed to play key roles in facilitating the metastatic cascade. Metastatic lesions often exhibit a similar epithelial-like state to that of the primary tumour, in particular, by forming carcinoma cell clusters via E-cadherin-mediated junctional complexes. However, the factors enabling mesenchymal-like micrometastatic cells to resume growth and reacquire an epithelial phenotype in the target organ microenvironment remain elusive. In this study, we developed a workflow using image-based cell profiling and machine learning to examine morphological, contextual and molecular states of individual breast carcinoma cells (MDA-MB-231). MDA-MB-231 heterogeneous response to the host organ microenvironment was modelled by substrates with controllable stiffness varying from 0.2kPa (soft tissues) to 64kPa (bone tissues). We identified 3 distinct morphological cell types (morphs) varying from compact round-shaped to flattened irregular-shaped cells with lamellipodia, predominantly populating 2-kPa and >16kPa substrates, respectively. These observations were accompanied by significant changes in E-cadherin and vimentin expression. Furthermore, we demonstrate that the bone-mimicking substrate (64kPa) induced multicellular cluster formation accompanied by E-cadherin cell surface localisation. MDA-MB-231 cells responded to different substrate stiffness by morphological adaptation, changes in proliferation rate and cytoskeleton markers, and cluster formation on bone-mimicking substrate. Our results suggest that the stiffest microenvironment can induce MET.

Incoherent wavefront reconstruction by a retroemission device containing a thin fluorescent film: theory.

A retroemission device (REM) is an incoherent holographic device that represents a lenslet array situated on a substrate containing fluorescent material. Each lenslet focuses each wavelet of an optical wavefront incident on the REM device into a diffraction-limited volume (voxel) in the fluorescent material, so that the voxel coordinates encode the angle of incidence and curvature of the wavelet. The back-propagating fraction of the excited fluorescence is collected by the lenslet and quasi-collimated into a back-propagating wavelet. All wavelets are combined to reconstruct the incident wavefront propagating in the backward direction. We present a theoretical model of REM based on Fresnel-Kirchhoff approximation describing the reconstructed 3D image characteristics versus the thickness of the fluorescence film at the focal plane of the lenslets. Results of the computer simulations of the REM-based images of a point source, two axially separated point sources and an extended object (a circular rim) situated in the sagittal plane are presented. These results speak in favor of using a fluorescence film of minimum diffraction-limited thickness at the lenslet back focal plane. This REM structure minimizes the fluorescence background and improves the 3D imaging resolution in virtue of the exclusion of out-of-voxel fluorescence contributions to the reconstructed wavefront.

Tracing upconversion nanoparticle penetration in human skin.

Due to their unique optical properties upconversion nanoparticles (UCNPs) provide exceptionally high contrast for imaging of true nanoparticle distribution in excised human skin. It makes possible to show penetration of solid nanoparticles in skin treated with chemical enhancers. We demonstrated tracing upconversion nanoparticles in excised human skin by means of optical microscopy at the discrete particle level sensitivity to obtain their penetration profiles, which was validated by laser-ablation inductively-coupled-plasma mass-spectrometry. To demonstrate utilities of our method, UCNPs were coated with polymers, formulated in water and chemical enhancers, and applied on excised human skin mounted on Franz cells, followed by imaging using a custom-built laser-scanning microscope. To evaluate the toxicity impact on skin by polymer-coated UCNPs, we introduced a tissue engineering model of viable epidermis made of decellularized chick embryo skin seeded with keratinocytes. UCNPs formulated in water stopped in stratum corneum, whereas UCNPs formulated in ethanol-water solution crossed stratum corneum and reached viable epidermis - hence, the enhancement effect for solid nanoparticles was detected by optical microscopy. All polymer-coated UCNPs were found nontoxic within the accepted safety levels. The keratinocyte resilience to polyethyleneimine-coated UCNPs was surprising considering cytotoxicity of polyethyleneimine to two-dimensional cell cultures.