Migdal, C. | bioRxiv Science

bioRxiv2023

2D cell culture on soft hydrogel drives an endoplasmic reticulum stress-dependent S quiescence underlying molecular traits of pulmonary basal cells

Cell culture on soft matrix, either in 2D and 3D, preserves the characteristics of progenitors. However, the mechanism by which the mechanical microenvironment determines progenitor phenotype, and its relevance to human biology, remains poorly described. Here we designed multi-well hydrogel plates with a high degree of physico-chemical uniformity to reliably address the molecular mechanism underlying cell state modification driven by physiological stiffness. Cell cycle, differentiation and metabolic activity could be studied in parallel assays, showing that the soft environment promotes an atypical S-phase quiescence and prevents cell drift, while preserving the differentiation capacities of human bronchoepithelial cells. These softness-sensitive responses are driven by defects in proteostasis and enhanced basal endoplasmic reticulum stress. The analysis of available single cell data of the human lung also showed that this non-conventional state coming from the soft extracellular environment is indeed consistent with molecular feature of pulmonary basal cells. Overall, this study demonstrates that mechanical mimicry in 2D culture supports allows to maintain progenitor cells in a state of high physiological relevance for characterizing novel molecular events that govern progenitor biology in human tissues.

cell biology↗

bioRxiv2019

Glial cell mechanosensitivity is reversed by adhesion cues

Brain tissues demonstrate heterogeneous mechanical properties, which evolve with aging and pathologies. The observation in these tissues of smooth to sharp rigidity gradients raises the question of brain cells responses to both different values of rigidity and their spatial variations. Here, we use recent techniques of hydrogel photopolymerization to achieve stiffness structuration down to micrometer resolution. We investigate primary neuron adhesion and orientation as well as glial cell adhesive and proliferative properties on multi-rigidity polyacrylamide hydrogels presenting a uniform density of adhesive molecules. We first observed that neurons grow following rigidity gradients. Then, our main observation is that glial cell adhesion and proliferation can be enhanced on stiff or on soft regions depending on the adhesive coating of the hydrogel, i. e. fibronectin or poly-L-lysine/laminin. This behavior was unchanged in the presence or not of neuronal cells. In addition, and contrarily to other cell types, glial cells were not confined by sharp, micron-scaled gradients of rigidity. Our observations suggest that their mechanosensitivity could involve adheison-related mechanosensitive pathways that are specific to brain tissues. SIGNIFICANCEBy growing primary brain cells on 2D multi-rigidity polyacrylamide hydrogels, we show that favorable culture conditions for glial cells switch from stiff to soft substrates when changing the adhesive ligands from fibronectin to poly-L-lysine/laminin. Together with neurons, glial cells thus provide a unique example where soft is preferred to stiff, but unlike neurons, this preference can be reversed by changing the nature of the coating. We additionally show that contrarily to other cell types, glial cells are deformed by subcellular gradients of rigidity but cannot be confined by these rigidity gradients. These observations point that glial cell use a very specific, integrin-related machinery for rigidity sensing.

biophysics↗