Responsive Giant Vesicles filled with Poly(N-isopropylacrylamide) Sols or Gels

4 pagesInternational audienceWe prepared giant unilamellar vesicles (GUVs) enclosing solutions or covalent gels of Poly(Nisopropylacrylamide) (PolyNipam). Concentrated suspensions of GUVs were prepared by applying an alternative field on a lipid film hydrated by a monomer solution containing N-isopropylacrylamide, crosslinker (N,N-methylene-bis-acrylamide), initiator and sucrose. Vesicle inner medium was polymerised and crosslinked by UV irradiation of the suspension, yielding viscous vesicles enclosing a solution of linear PolyNipam chains (when no bisacrylamide was used) or elastic vesicles filled with a covalent PolyNipam gel. We show that gel-filled vesicles are responsive systems triggered by the temperature: they shrink, reducing by a factor eight their volume below the critical temperature (32 ◦C in water, lower in glucose solution) and re-swell in a reversible and reproducible way upon decreasing temperature. In both cases, we show that the vesicle lipid membrane interacts with the internal polymer, resulting in an strong resistance of the vesicles to external mechanical stresses (enhanced tension of lysis)

Fluctuations of a membrane nanotube covered with an actin sleeve

Many biological functions rely on the reshaping of cell membranes, in particular into nanotubes, which are covered in vivo by dynamic actin networks. Nanotubes are subject to thermal fluctuations, but the effect of these on cell functions is unknown. Here, we form nanotubes from liposomes using an optically trapped bead adhering to the liposome membrane. From the power spectral density of this bead, we study the nanotube fluctuations in the range of membrane tensions measured in vivo. We show that an actin sleeve covering the nanotube damps its high frequency fluctuations because of the network viscoelasticity. Our work paves the way for further studies on the effect of nanotube fluctuations in cellular functions

F-actin mechanics control spindle centring in the mouse zygote

International audienceMitotic spindle position relies on interactions between astral microtubules nucleated by centrosomes and a rigid cortex. Some cells, such as mouse oocytes, do not possess centrosomes and astral microtubules. These cells rely only on actin and on a soft cortex to position their spindle off-centre and undergo asymmetric divisions. While the first mouse embryonic division also occurs in the absence of centrosomes, it is symmetric and not much is known on how the spindle is positioned at the exact cell centre. Using interdisciplinary approaches, we demonstrate that zygotic spindle positioning follows a three-step process: (1) coarse centring of pronuclei relying on the dynamics of an F-actin/Myosin-Vb meshwork; (2) fine centring of the metaphase plate depending on a high cortical tension; (3) passive maintenance at the cell centre. Altogether, we show that F-actin-dependent mechanics operate the switch between asymmetric to symmetric division required at the oocyte to embryo transition

Développement et étude d'objets biomimétiques stimulables : Vésicules géantes encapsulant des systèmes visco-élastiques de poly(NIPAM)

The goal of this PhD was to design responsive biomimetic objects from Giant Unilamellar Vesicles and PolyNipam solutions or gels. We highlighted the coupling between the lipid membrane and the internal polyNipam medium, which confers those composite vesicles thermoresponsive properties, and the behaviour of such objects during thermal transition has been investigated by quantitative fluorescence studies. We have also measured the influence of the coupling between polyNipam chains and lipid membrane on the bending modulus of the membranes. Then the mechanical properties of the composite vesicles have been measured using micropipette aspiration technique, showing the relevance of those objects as cellular mechanical models. Further studies concerning adhesion properties, membrane tether extrusion and lipid extraction from composite vesicles have been adressed

Développement et étude d'objets biomimétiques stimulables : Vésicules géantes encapsulant des systèmes visco-élastiques de poly(NIPAM)

The goal of this PhD was to design responsive biomimetic objects from Giant Unilamellar Vesicles and PolyNipam solutions or gels. We highlighted the coupling between the lipid membrane and the internal polyNipam medium, which confers those composite vesicles thermoresponsive properties, and the behaviour of such objects during thermal transition has been investigated by quantitative fluorescence studies. We have also measured the influence of the coupling between polyNipam chains and lipid membrane on the bending modulus of the membranes. Then the mechanical properties of the composite vesicles have been measured using micropipette aspiration technique, showing the relevance of those objects as cellular mechanical models. Further studies concerning adhesion properties, membrane tether extrusion and lipid extraction from composite vesicles have been adressed

Studying actin-induced cell shape changes using Giant Unilamellar Vesicles and reconstituted actin networks

International audienceCell shape changes that are fuelled by the dynamics of the actomyosin cytoskeleton control cellular processes such as motility and division. However, the mechanisms of interplay between cell membranes and actomyosin are complicated to decipher in the complex environment of the cytoplasm. Using biomimetic systems offers an alternative approach to studying cell shape changes in assays with controlled biochemical composition. Biomimetic systems allow quantitative experiments that can help to build physical models describing the processes of cell shape changes. This article reviews works in which actin networks are reconstructed inside or outside cell-sized Giant Unilamellar Vesicles (GUVs), which are models of cell membranes. We show how various actin networks affect the shape and mechanics of GUVs and how some cell shape changes can be reproduced in vitro using these minimal systems

Responsive viscoelastic giant lipid vesicles filled with a poly(N-isopropylacrylamide) artificial cytoskeleton

International audienceResponsive giant lipid vesicles filled with aqueous PolyNipam sol (SFV) or gel (GFV) were prepared by ultra-violet polymerisation performed in situ. Upon crossing the lower critical transition temperature of PolyNipam, SFVs and GFVs undergo a significant change of their structural and mechanical properties or a drastic volume transition, respectively. Rheometric and micropipette experiments show that both internal viscosity of SFVs and internal shear modulus of GFVs are tunable over several orders of magnitude and lie in the range observed for living cells. Moreover, the vesicle membrane is strongly bound to the internal polymer medium, making these systems interesting for mimicking the basic mechanical behaviour of passive living cells

Interplay between membrane tension and the actin cytoskeleton determines shape changes

International audienceThe ability of mammalian cells to deform their membrane relies on the action of the cytoskeleton. In particular, the dynamics of the actin cytoskeleton, assembling at the plasma membrane, plays a crucial role in controlling cell shape. Many proteins are involved to ensure proper growth of the actin network at the cell membrane. The detailed structure of this network regulates the force that is necessary for membrane deformation. We address here how the presence of capping proteins, which limit the length of actin filaments and thus affects network topology, influences membrane shape. We use a system of liposomes, activated to polymerize actin at their surface, and placed in a mixture of purified proteins that reconstitutes actin dynamics. Our system also allows the variation of membrane tension by deflating the liposomes. We show that membrane deformations are clearly favored in the presence of capping proteins in the actin network. Moreover, in the absence of capping proteins, membrane deformations appear only when the liposomes are deflated. Our results unveil that the interplay between membrane tension and actin network structure and dynamics governs cell shape

Remodelling of membrane tubules by the actin cytoskeleton

International audienc

Methods for Assessing Oocyte Quality: A Review of Literature

International audienceThe rate of infertility continues to rise in the world for several reasons, including the age of conception and current lifestyle. We list in this paper potential non-invasive and invasive techniques to assess oocyte quality. We searched the database PubMed using the terms “oocytes AND quality AND evaluation”. In the first part, we study the morphological criteria, compartment by compartment, to then focus in a second part on more objective techniques such as genetics, molecular, apoptosis, or human follicular fluid that contain biologically active molecules. The main criteria used to assess oocyte quality are morphological; however, several other techniques have been studied in women to improve oocyte quality assessment, but most of them are invasive and not usable in routine