An interdisciplinary approach to the control of signaling at cell-cell junctions

  • Viasnoff lab
  • Viasnoff lab

Controlled micro environments and microprobes. From micro-dishes to micro-niches.

We developed and patented a technology to create microwells with 3D differential protein coating that allows to coat the sides, top and bottom of the wells with different protein. We use these wells to present cell with geometric, rheological and chemical cues organized in 3D. We use the same technique to develop microfluidic devices to apply localized forces on cell-cell contact by control aspiration. We are now looking into coupling this approach to waveguide to perform super resolution microscopy in 3D. (In collaboration with Vincent Studer CNRS Bordeaux, France).

3D microwells: Development of micro-wells with controlled geometry and 3D chemical cues

We are exploring various possibilities to perform local chemical grafting on acrylate based microwells to recreate a microenvironment with controlled geometry, chemistry and rheology. This include protein adsorption, 3D protein patterning, in situ growth of hydrogel and inorganic surface treatments. Patent application number GB1213654.5.

 

3D super resolution imaging

Coll Vincent Studer (CNRS)
High risk Top secret project. Coming soon, hopefully.

 

Microfluidic tools to test cell adhesion

We developed a 3D microfluidic device that allows local stretching and shearing of a cell-cell contact by application of suction forces.

 
 

 

Mechanosensitive response and mesoscopic organization of adherens junction.

We use our microwell approach to control the formation of Adherens junction (junction mediated by Cadherin) and to study the influence of the cell cytoskeleton organization on the morphology, ultra-structure of the junction as well as the local mechanical stress map. Our goal is to construct a multiscale model to describe what sets the adhesive strength of an Adherens junction.

Cell-doublets in suspension

Coll J.P.Thiery (Astar)
We image cell doublet immobilized in non adhesive well and we study the formation of the contact for two cells stacked on each other.

 

Quantitative evaluation of the tension at the cell-cell contact.

Coll Y.Toyama (MBI)
We use laser ablation and pipette aspiration to establish the relation between the tension at a cell cell contact and the shape of the junction. We perform these experiments on cell doublet or controlled cell aggregate. We study the influence of the structuration of the cytoskeleton by ECM interaction on the tension at the cell-cell contact.

 

Reconstitution of artificial cell-cell contacts on supported bilayer

Coll J.Groves (Berkeley)
We reconstitute E-cad on restricted supported bilayers and study the role of the confinement and E-cad mobility on the structuration of this artificial cell-cell junction.

 

Polarization of the cell structure

We study the influence of the formation of the cell-cell junction on the structuration of the nucleus and other organelles inside the cytoskeleton for cell doublet and in controlled aggregates.

 
 

 

Mechanobiology of Hepatocytes. Morpho-functional control of bile canaliculi.

Hepatic apical lumen, or bile canaliculi (BC) is tubular network formed in between hepatocytes to maintain a clean microenvironment by draining metabolic waste out of liver. However, it is poorly known how BC develops into tubular shapes, especially how ECM regulates this process because of technical limitation on controlling 3D ECM presentation in microscale. Here, we propose a micro well system the side and bottom of which can be differentially coated with different chemicals, such as ECM protein fibronectin and passivation reagent pluronic acid.

Hepatocytes in μWells are healthy till at least 4 days after seeding and form functional Bile Canaliculi. Hepatocyte couplets orientate themselves into top-bottom manner in the μWells with bottoms coated by fibronectin, meanwhile adapt a side-side fashion in the μWells with bottoms passivated by pluronic acid and sides coated by fibronectin. In the μWells with bottom coated by fibronectin, interestingly, BC develops into more tubular shape when the sides are coated by fibronectin, comparing with the one when the sides are passivated by pluronic acid. This indicates that the ECM presentation under BC plays an important role to lead BC into tubular shape. Thus, we invented a new method to differentially control 3D ECM presentation in microscale. By varying ECM configurations, we could not only regulate BC orientation but also demonstrate the importance of ECM as a guidance for BC tubular shape formation.

Our patented hepatocytes culture system

We used our microwell technology to culture primary hepatocytes over days with no loss of function at the single doublet level or in microcolonies.

 

Morpho-metabolic relation in BC formation

Coll H.Yu (IBN/MBI)
By presenting the appropriate micro-environment to hepatocytes (micro-niches) we control the organization of the cytoskeleton and its influence on the morphology of bile canaliculi. We aim at elucidating the relationship between the shape and metabolism of Bile Canaliculi.

 

Controlling self organization and extension of Bile canaliculi

We further develop our technique to control the growth of bile canaliculi across many cell junctions. We study the physical and biochemical basis of the structure formation. Our goal is to recreate in-vivo like Bile Canaliculi.