Biology subjects

Chevallet, M.

Publications and source records attributed to Chevallet, M..

4 recordsLinked to original sources

bioRxiv2026

A general methodology for liver sinusoid fenestration analysis based on 3D electron microscopy data

The liver has a complex architecture composed of millions of lobules. Within these lobules, hepatocytes, the main hepatic cells, are organized in rows separated by blood capillaries known as sinusoids. These capillaries are lined by liver sinusoidal endothelial cells (LSEC) that form a very specific fenestrated endothelium essential for the exchange of metabolites and proteins between the blood and hepatocytes. Alterations in the size and number of LSEC fenestrations are associated with the onset and the progression of various liver diseases. The analysis of liver architecture is thus of utmost importance for advancing our knowledge of liver ultrastructure and its alterations. Liver architecture has been studied since decades, mainly using 2D electron microscopy, and more recently using advanced super-resolution fluorescence microscopy. In recent years, volume electron microscopy techniques, including focused ion beam-scanning electron microscopy (FIB-SEM) progressed and nowadays enable the 3D reconstruction of biological ultrastructures down to nanometer resolution. However, the analysis of large volumes (e.g., several tens of {micro}m3) remains challenging due to various constraints in the segmentation of large datasets. In the current study, we developed a workflow to semi-automatically segment hepatic sinusoids from FIB-SEM mice liver datasets using the CNN-based (convolutional neural network) tool known as "nnU-Net", after fine-tuning a ground truth model. We also implemented tools for semi-automatic quantification of LSEC fenestrae diameters and sinusoid porosity from segmented datasets. This workflow enabled us to compare the distribution of LSEC fenestrae diameters in wild-type versus Bmp9-deleted mice, a hepatic factor known to be involved in fenestration maintenance. Our results confirm the importance of BMP9 for LSEC differentiation. Therefore, the developed methodology represents a valuable tool for characterizing the fenestrated endothelium under various physiological and pathological conditions.

cell biology↗

bioRxiv2024

E cadherin appears to be an essential on/off switch for initiating bile canaliculi formation

The mechanisms underlying cell polarization are fundamental in biology, yet they are not fully understood. This is particularly true for hepatocytes, which exhibit a complex polarization, enabling the formation of the bile canaliculi (BCs) network that is essential for liver excretion functions. To identify key proteins involved in hepatocyte polarization and BC formation, we performed a proteomic approach to compare the human hepatocyte cell line HepG2 to its sub clone HepG2/C3A which shows much greater efficiency in forming mature BCs. We localized LimA1 and Espin to the BC for the first time, suggesting their important role there, and confirmed the presence of NHE-RF1. Using a protein repression strategy directed against selected proteins, we highlighted the essential role of E cadherin in the initiation of BC formation. Our data showed, for the first time, that in absence of E cadherin, hepatocytes lose their ability to form BCs.

cell biology↗

bioRxiv2020

Correlative transmission electron microscopy and high-resolution hard X-ray fluorescence microscopy of cell sections to measure trace elements concentrations at the organelle level

Metals are essential to all forms of life and their concentration and distribution in the organisms are tightly regulated. Indeed, in their free form, metal ions are toxic. Therefore, an excess of physiologic metal ions or the uptake of non-physiologic metal ions can be highly detrimental for the organisms. It is thus fundamental to understand metals distribution and dynamics in physiologic or disrupted conditions, for instance in metal-related pathologies or upon environmental exposure to metals. Elemental imaging techniques can serve this purpose, by allowing the visualization and the quantification of metal species in a tissue or down to the interior of a cell. Among these techniques, synchrotron radiation-based X-ray fluorescence (SR-XRF) microscopy is the most sensitive to date, and great progresses were made to reach spatial resolutions as low as 20x20 nm2. Until recently, 2D XRF mapping was used on whole cells, thus summing up the signal from the whole thickness of the cell. In the last two years, we have developed a methodology to work on thin cell sections, in order to analyze the metal content at the level of the organelle. Herein, we propose a correlative method to couple SR-XRF to electron microscopy, with the aim to quantify the elemental content in an organelle of interest. As a proof-of-concept, the technique was applied to the analysis of mitochondria from hepatocytes exposed to silver nanoparticles. It was thus possible to identify mitochondria with higher concentration of Ag(I) ions compared to the surrounding cytosol. The versatility of the method makes it suitable to answer a large panel of biological questions, for instance related to metal homeostasis in biological organisms.

cell biology↗

bioRxiv2019

Visualization of silver nanoparticle intracellular trafficking revealed nuclear translocation of silver ions leading to nuclear receptor impairment

The impact on human health of the increasing use of silver nanoparticles (AgNPs) in medical devices remains understudied, even though AgNP-containing dressings are known to release silver in the bloodstream leading to accumulation and slow clearance in the liver. Cellular studies have shown the intracellular dissolution of AgNPs within endo-lysosomes followed by Ag(I) binding to biomolecular thiolate-containing molecules. However, the precise subcellular distribution of Ag(I) and the nature of the disrupted physiological pathways remained unknown. Novel imaging approaches enabled us to visualize the trafficking of AgNP-containing lysosomes towards a perinuclear location and a direct nuclear transfer of Ag(I) species with accumulation in the nucleoli. These Ag(I) species impaired nuclear receptor activity, disrupting critical mechanisms of liver physiology in very low dose exposure scenarios, thus justifying further research into defining a framework for the safe use of AgNPs.

pharmacology and toxicology↗