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Blebs are spherical cellular protrusions that occur in many physiological situations. Two distinct phases make up the life of a bleb, each of which have their own biology and physics: expansion, which lasts ∼30 s, and retraction, which lasts ∼2 min. We investigate these phases using optical microscopy and simple theoretical concepts, seeking information on blebbing itself, and on cytomechanics in general. We show that bleb nucleation depends on pressure, membrane-cortex adhesion energy, and membrane tension, and test this experimentally. Bleb growth occurs through a combination of bulk flow of lipids and delamination from the cell cortex via the formation and propagation of tears. In extreme cases, this can give rise to a traveling wave around the cell periphery, known as “circus movement.” When growth stalls, an actin cortex reforms under the bleb membrane, and retraction starts, driven by myosin-II. Using flicker spectroscopy, we find that retracting blebs are fivefold more rigid than expanding blebs, an increase entirely explained by the properties of the newly formed cortical actin mesh. Finally, using artificially nucleated blebs as pressure sensors, we show that cells rounded up in mitosis possess a substantial intracellular pressure. 相似文献
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Marion?Bichet Candie?Joly Ahmed?Hadj Henni Thomas?Guilbert Marie?Xémard Vincent?Tafani Vanessa?Lagal Guillaume?Charras Isabelle?TardieuxEmail author 《BMC biology》2014,12(1):773
Background
The public health threats imposed by toxoplasmosis worldwide and by malaria in sub-Saharan countries are directly associated with the capacity of their related causative agents Toxoplasma and Plasmodium, respectively, to colonize and expand inside host cells. Therefore, deciphering how these two Apicomplexan protozoan parasites access their host cells has been highlighted as a priority research with the perspective of designing anti-invasive molecules to prevent diseases. Central to the mechanism of invasion for both genera is mechanical force, which is thought to be applied by the parasite at the interface between the two cells following assembly of a unique cell-cell junction but this model lacks direct evidence and has been challenged by recent genetic studies. In this work, using parasites expressing the fluorescent core component of this junction, we analyze characteristic features of the kinematics of penetration of more than 1,000 invasion events.Results
The majority of invasion events occur with a typical forward rotational progression of the parasite through a static junction into an invaginating host cell plasma membrane. However, if parasites encounter resistance and if the junction is not strongly anchored to the host cell cortex, as when parasites do not secrete the toxofilin protein and, therefore, are unable to locally remodel the cortical actin cytoskeleton, the junction travels retrogradely with the host cell membrane along the parasite surface allowing the formation of a functional vacuole. Kinetic measurements of the invasive trajectories strongly support a similar parasite driven force in both static and capped junctions, both of which lead to successful invasion. However, about 20% of toxofilin mutants fail to enter and eventually disengage from the host cell membrane while the secreted RhOptry Neck (RON2) molecules are posteriorally capped before being cleaved and released in the medium. By contrast in cells characterized by low cortex tension and high cortical actin dynamics junction capping and entry failure are drastically reduced.Conclusions
This kinematic analysis newly highlights that to invade cells parasites need to engage their motor with the junction molecular complex where force is efficiently applied only upon proper anchorage to the host cell membrane and cortex.25.
The purpose of this study was to investigate the need for and choice of stratification factors, and the effects of blinding
and placebo in a clinical experiment. Eighty dogs with canine hip dysplasia (CHD) were included in a randomized, placebo-controlled
and double blind clinical trial with stratified parallel group design, in which body weight and degree of CHD were used as
stratification factors. Thirty-eight dogs were allocated to gold bead implantation and 42 to placebo. After six months, 33
of the 42 placebo-treated dogs received gold bead implantation in an open study lasting a further 18 months. The main outcome
variable in the study was change in pain signs of CHD as assessed by the owner. No significant difference in the main outcome
variable, regardless of the treatment given, could be detected in the two chosen stratification factors. The only factor to
influence the main outcome variable significantly was age. The blinding procedure used in the study, in which 60% of the owners
correctly guessed the treatment given, was found sufficient. Of those who guessed the treatment erroneously, 88% believed
the treatment given was gold bead implantation. The treatment efficacy after six months in the blinded treatment group was
found to be significantly larger compared to the efficacy obtained in the open study. A significant placebo effect was therefore
detected. Conclusion and Clinical Relevance: The age of the dogs influenced the outcome of the CHD treatment, and is recommended
as a stratification factor. A significant placebo effect has to be expected and an optimal blinding procedure is necessary
in similar clinical studies. 相似文献
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Naomi Eidelman Alan Boyde Andrew J Bushby Peter GT Howell Jirun Sun Dale E Newbury Frederick W Miller Pamela G Robey Lisa G Rider 《Arthritis research & therapy》2009,11(5):R159-21
Introduction
Calcified deposits (CDs) in skin and muscles are common in juvenile dermatomyositis (DM), and less frequent in adult DM. Limited information exists about the microstructure and composition of these deposits, and no information is available on their elemental composition and contents, mineral density (MD) and stiffness. We determined the microstructure, chemical composition, MD and stiffness of CDs obtained from DM patients. 相似文献27.
M Tangney GT Winters WJ Mitchell 《Journal of industrial microbiology & biotechnology》2001,27(5):298-306
The utilization of maltose by Clostridium acetobutylicum ATCC 824 was investigated. Glucose was used preferentially to maltose, when both substrates were present in the medium. Maltose
phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) activity was detected in extracts prepared from cultures
grown on maltose, but not glucose or sucrose, as the sole carbon source. Extract fractionation and PTS reconstitution experiments
revealed that the specificity for maltose is contained entirely within the membrane in this organism. A putative gene system
for the maltose PTS was identified (from the C. acetobutylicum ATCC 824 genome sequence), encoding an enzyme IIMal and a maltose 6-phosphate hydrolase.
Journal of Industrial Microbiology & Biotechnology (2001) 27, 298–306.
Received 12 September 2000/ Accepted in revised form 30 November 2000 相似文献
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Proteins of the ERM family (ezrin, moesin, radixin) play a fundamental role in tethering the membrane to the cellular actin cortex as well as regulating cortical organization and mechanics. Overexpression of dominant inactive forms of ezrin leads to fragilization of the membrane-cortex link and depletion of moesin results in softer cortices that disrupt spindle orientation during cytokinesis. Therefore, the kinetics of association of ERM proteins with the cortex likely influence the timescale of cortical signaling events and the dynamics of membrane interfacing to the cortex. However, little is known about ERM protein turnover at the membrane-cortex interface. Here, we examined cortical ezrin dynamics using fluorescence recovery after photobleaching experiments and single-molecule imaging. Using multiexponential fitting of fluorescence recovery curves, we showed that ezrin turnover resulted from three molecular mechanisms acting on very different timescales. The fastest turnover process was due to association/dissociation from the F-actin cortex, suggesting that ezrin acts as a link that leads to low friction between the membrane and the cortex. The second turnover process resulted from association/dissociation of ezrin from the membrane and the slowest turnover process resulted from the slow diffusion of ezrin in the plane of the membrane. In summary, ezrin-mediated membrane-cortex tethering resulted from long-lived interactions with the membrane via the FERM domain coupled with shorter-lived interactions with the cortex. The slow diffusion of membranous ezrin and its interaction partners relative to the cortex signified that signals emanating from membrane-associated ezrin may locally act to modulate cortical organization and contractility. 相似文献
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Proteins of the ERM family (ezrin, moesin, radixin) play a fundamental role in tethering the membrane to the cellular actin cortex as well as regulating cortical organization and mechanics. Overexpression of dominant inactive forms of ezrin leads to fragilization of the membrane-cortex link and depletion of moesin results in softer cortices that disrupt spindle orientation during cytokinesis. Therefore, the kinetics of association of ERM proteins with the cortex likely influence the timescale of cortical signaling events and the dynamics of membrane interfacing to the cortex. However, little is known about ERM protein turnover at the membrane-cortex interface. Here, we examined cortical ezrin dynamics using fluorescence recovery after photobleaching experiments and single-molecule imaging. Using multiexponential fitting of fluorescence recovery curves, we showed that ezrin turnover resulted from three molecular mechanisms acting on very different timescales. The fastest turnover process was due to association/dissociation from the F-actin cortex, suggesting that ezrin acts as a link that leads to low friction between the membrane and the cortex. The second turnover process resulted from association/dissociation of ezrin from the membrane and the slowest turnover process resulted from the slow diffusion of ezrin in the plane of the membrane. In summary, ezrin-mediated membrane-cortex tethering resulted from long-lived interactions with the membrane via the FERM domain coupled with shorter-lived interactions with the cortex. The slow diffusion of membranous ezrin and its interaction partners relative to the cortex signified that signals emanating from membrane-associated ezrin may locally act to modulate cortical organization and contractility. 相似文献
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Marco Fritzsche Alexandre Lewalle Tom Duke Karsten Kruse Guillaume Charras 《Molecular biology of the cell》2013,24(6):757-767
The cell cortex is a thin network of actin, myosin motors, and associated proteins that underlies the plasma membrane in most eukaryotic cells. It enables cells to resist extracellular stresses, perform mechanical work, and change shape. Cortical structural and mechanical properties depend strongly on the relative turnover rates of its constituents, but quantitative data on these rates remain elusive. Using photobleaching experiments, we analyzed the dynamics of three classes of proteins within the cortex of living cells: a scaffold protein (actin), a cross-linker (α-actinin), and a motor (myosin). We found that two filament subpopulations with very different turnover rates composed the actin cortex: one with fast turnover dynamics and polymerization resulting from addition of monomers to free barbed ends, and one with slow turnover dynamics with polymerization resulting from formin-mediated filament growth. Our data suggest that filaments in the second subpopulation are on average longer than those in the first and that cofilin-mediated severing of formin-capped filaments contributes to replenishing the filament subpopulation with free barbed ends. Furthermore, α-actinin and myosin minifilaments turned over significantly faster than F-actin. Surprisingly, only one-fourth of α-actinin dimers were bound to two actin filaments. Taken together, our results provide a quantitative characterization of essential mechanisms underlying actin cortex homeostasis. 相似文献