Person:

Millet, Emil

In wound healing, tissue growth, and certain cancers, the epithelial or the endothelial monolayer sheet expands. Within the expanding monolayer sheet, migration of the individual cell is strongly guided by physical forces imposed by adjacent cells. This process is called plithotaxis and was discovered using Monolayer Stress Microscopy (MSM). MSM rests upon certain simplifying assumptions, however, concerning boundary conditions, cell material properties and system dimensionality. To assess the validity of these assumptions and to quantify associated errors, here we report new analytical, numerical, and experimental investigations. For several commonly used experimental monolayer systems, the simplifying assumptions used previously lead to errors that are shown to be quite small. Out-of-plane components of displacement and traction fields can be safely neglected, and characteristic features of intercellular stresses that underlie plithotaxis remain largely unaffected. Taken together, these findings validate Monolayer Stress Microscopy within broad but well-defined limits of applicability.

Background: The aminoglycoside antibiotic gentamicin is an ototoxic drug and has been used experimentally to investigate cochlear damage induced by noise. We have investigated the changes in the protein profile associated with caveolae in gentamicin treated and untreated spiral ligament (SL) pericytes, specialized cells in the blood labyrinth barrier of the inner ear microvasculature. Pericytes from various microvascular beds express caveolae, protein and cholesterol rich microdomains, which can undergo endocytosis and transcytosis to transport small molecules in and out the cells. A different protein profile in transport-specialized caveolae may induce pathological changes affecting the integrity of the blood labyrinth barrier and ultimately contributing to hearing loss. Method Caveolae isolation from treated and untreated cells is achieved through ultracentrifugation of the lysates in discontinuous gradients. Mass spectrometry (LC-MS/MS) analysis identifies the proteins in the two groups. Proteins segregating with caveolae isolated from untreated SL pericytes are then compared to caveolae isolated from SL pericytes treated with the gentamicin for 24 h. Data are analyzed using bioinformatic tools. Results: The caveolae proteome in gentamicin treated cells shows that 40% of total proteins are uniquely associated with caveolae during the treatment, and 15% of the proteins normally associated with caveolae in untreated cell are suppressed. Bioinformatic analysis of the data shows a decreased expression of proteins involved in genetic information processing, and an increase in proteins involved in metabolism, vesicular transport and signal transduction in gentamicin treated cells. Several Rab GTPases proteins, ubiquitous transporters, uniquely segregate with caveolae and are significantly enriched in gentamicin treated cells. Conclusion: We report that gentamicin exposure modifies protein profile of caveolae from SL pericytes. We identified a pool of proteins which are uniquely segregating with caveolae during the treatment, mainly participating in metabolic and biosynthetic pathways, in transport pathways and in genetic information processing. Finally, we show for the first time proteins associated with caveolae SL pericytes linked to nonsyndromic hearing loss. Electronic supplementary material The online version of this article (10.1186/s12953-018-0132-x) contains supplementary material, which is available to authorized users.