Shaken, but not stirred: how vortical flow drives small-scale aggregations of gyrotactic phytoplankton

NASA Astrophysics Data System (ADS)

Barry, Michael; Durham, William; Climent, Eric; Stocker, Roman

2011-11-01

Coastal ocean observations reveal that motile phytoplankton form aggregations at the Kolmogorov scale (mm-cm), whereas non-motile cells do not. We propose a new mechanism for the formation of this small-scale patchiness based on the interplay of turbulence and gyrotactic motility. Counterintuitively, turbulence does not stir a plankton suspension to homogeneity but drives aggregations instead. Through controlled laboratory experiments we show that the alga Heterosigma akashiwo rapidly forms aggregations in a cavity-driven vortical flow that approximates Kolmogorov eddies. Gyrotactic motility is found to be the key ingredient for aggregation, as non-motile cells remain randomly distributed. Observations are in remarkable agreement with a 3D model, and the validity of this mechanism for generating patchiness has been extended to realistic turbulent flows using Direct Numerical Simulations. Because small-scale patchiness influences rates of predation, sexual reproduction, infection, and nutrient competition, this result indicates that gyrotactic motility can profoundly affect phytoplankton ecology.

Type IV pili interactions promote intercellular association and moderate swarming of Pseudomonas aeruginosa

PubMed Central

Anyan, Morgen E.; Amiri, Aboutaleb; Harvey, Cameron W.; Tierra, Giordano; Morales-Soto, Nydia; Driscoll, Callan M.; Alber, Mark S.; Shrout, Joshua D.

2014-01-01

Pseudomonas aeruginosa is a ubiquitous bacterium that survives in many environments, including as an acute and chronic pathogen in humans. Substantial evidence shows that P. aeruginosa behavior is affected by its motility, and appendages known as flagella and type IV pili (TFP) are known to confer such motility. The role these appendages play when not facilitating motility or attachment, however, is unclear. Here we discern a passive intercellular role of TFP during flagellar-mediated swarming of P. aeruginosa that does not require TFP extension or retraction. We studied swarming at the cellular level using a combination of laboratory experiments and computational simulations to explain the resultant patterns of cells imaged from in vitro swarms. Namely, we used a computational model to simulate swarming and to probe for individual cell behavior that cannot currently be otherwise measured. Our simulations showed that TFP of swarming P. aeruginosa should be distributed all over the cell and that TFP−TFP interactions between cells should be a dominant mechanism that promotes cell−cell interaction, limits lone cell movement, and slows swarm expansion. This predicted physical mechanism involving TFP was confirmed in vitro using pairwise mixtures of strains with and without TFP where cells without TFP separate from cells with TFP. While TFP slow swarm expansion, we show in vitro that TFP help alter collective motion to avoid toxic compounds such as the antibiotic carbenicillin. Thus, TFP physically affect P. aeruginosa swarming by actively promoting cell−cell association and directional collective motion within motile groups to aid their survival. PMID:25468980

Roles of ion transport in control of cell motility.

PubMed

Stock, Christian; Ludwig, Florian T; Hanley, Peter J; Schwab, Albrecht

2013-01-01

Cell motility is an essential feature of life. It is essential for reproduction, propagation, embryonic development, and healing processes such as wound closure and a successful immune defense. If out of control, cell motility can become life-threatening as, for example, in metastasis or autoimmune diseases. Regardless of whether ciliary/flagellar or amoeboid movement, controlled motility always requires a concerted action of ion channels and transporters, cytoskeletal elements, and signaling cascades. Ion transport across the plasma membrane contributes to cell motility by affecting the membrane potential and voltage-sensitive ion channels, by inducing local volume changes with the help of aquaporins and by modulating cytosolic Ca(2+) and H(+) concentrations. Voltage-sensitive ion channels serve as voltage detectors in electric fields thus enabling galvanotaxis; local swelling facilitates the outgrowth of protrusions at the leading edge while local shrinkage accompanies the retraction of the cell rear; the cytosolic Ca(2+) concentration exerts its main effect on cytoskeletal dynamics via motor proteins such as myosin or dynein; and both, the intracellular and the extracellular H(+) concentration modulate cell migration and adhesion by tuning the activity of enzymes and signaling molecules in the cytosol as well as the activation state of adhesion molecules at the cell surface. In addition to the actual process of ion transport, both, channels and transporters contribute to cell migration by being part of focal adhesion complexes and/or physically interacting with components of the cytoskeleton. The present article provides an overview of how the numerous ion-transport mechanisms contribute to the various modes of cell motility.

Effects of Motility and Adsorption Rate Coefficient on Transport of Bacteria through Saturated Porous Media

PubMed Central

Camper, Anne K.; Hayes, Jason T.; Sturman, Paul J.; Jones, Warren L.; Cunningham, Alfred B.

1993-01-01

Three strains of Pseudomonas fluorescens with different motility rates and adsorption rate coefficients were injected into porous-medium reactors packed with l-mm-diameter glass spheres. Cell breakthrough, time to peak concentration, tailing, and cell recovery were measured at three interstitial pore velocities (higher than, lower than, and much lower than the maximal bacterial motility rate). All experiments were done with distilled water to reduce the effects of growth and chemotaxis. Contrary to expectations, motility did not result in either early breakthrough or early time to peak concentration at flow velocities below the motility rate. Bacterial size exclusion effects were shown to affect breakthrough curve shape at the very low flow velocity, but no such effect was seen at the higher flow velocity. The tendency of bacteria to adsorb to porous-medium surfaces, as measured by adsorption rate coefficients, profoundly influenced transport characteristics. Cell recoveries were shown to be correlated with the ratio of advective to adsorptive transport in the reactors. Adsorption rate coefficients were found to be better predictors of microbial transport phenomena than individual characteristics, such as size, motility, or porous-medium hydrodynamics. PMID:16349075

Impairment of Swimming Motility by Antidiarrheic Lactobacillus acidophilus Strain LB Retards Internalization of Salmonella enterica Serovar Typhimurium within Human Enterocyte-Like Cells▿

PubMed Central

Liévin-Le Moal, Vanessa; Amsellem, Raymonde; Servin, Alain L.

2011-01-01

We report that both culture and the cell-free culture supernatant (CFCS) of Lactobacillus acidophilus strain LB (Lactéol Boucard) have the ability (i) to delay the appearance of Salmonella enterica serovar Typhimurium strain SL1344-induced mobilization of F-actin and, subsequently, (ii) to retard cell entry by S. Typhimurium SL1344. Time-lapse imaging and Western immunoblotting showed that S. Typhimurium SL1344 swimming motility, as represented by cell tracks of various types, was rapidly but temporarily blocked without affecting the expression of FliC flagellar propeller protein. We show that the product(s) secreted by L. acidophilus LB that supports the inhibitory activity is heat stable and of low molecular weight. The product(s) caused rapid depolarization of the S. Typhimurium SL1344 cytoplasmic membrane without affecting bacterial viability. We identified inhibition of swimming motility as a newly discovered mechanism by which the secreted product(s) of L. acidophilus strain LB retards the internalization of the diarrhea-associated pathogen S. enterica serovar Typhimurium within cultured human enterocyte-like cells. PMID:21825295

Next generation sequencing analysis reveals that the ribonucleases RNase II, RNase R and PNPase affect bacterial motility and biofilm formation in E. coli.

PubMed

Pobre, Vânia; Arraiano, Cecília M

2015-02-14

The RNA steady-state levels in the cell are a balance between synthesis and degradation rates. Although transcription is important, RNA processing and turnover are also key factors in the regulation of gene expression. In Escherichia coli there are three main exoribonucleases (RNase II, RNase R and PNPase) involved in RNA degradation. Although there are many studies about these exoribonucleases not much is known about their global effect in the transcriptome. In order to study the effects of the exoribonucleases on the transcriptome, we sequenced the total RNA (RNA-Seq) from wild-type cells and from mutants for each of the exoribonucleases (∆rnb, ∆rnr and ∆pnp). We compared each of the mutant transcriptome with the wild-type to determine the global effects of the deletion of each exoribonucleases in exponential phase. We determined that the deletion of RNase II significantly affected 187 transcripts, while deletion of RNase R affects 202 transcripts and deletion of PNPase affected 226 transcripts. Surprisingly, many of the transcripts are actually down-regulated in the exoribonuclease mutants when compared to the wild-type control. The results obtained from the transcriptomic analysis pointed to the fact that these enzymes were changing the expression of genes related with flagellum assembly, motility and biofilm formation. The three exoribonucleases affected some stable RNAs, but PNPase was the main exoribonuclease affecting this class of RNAs. We confirmed by qPCR some fold-change values obtained from the RNA-Seq data, we also observed that all the exoribonuclease mutants were significantly less motile than the wild-type cells. Additionally, RNase II and RNase R mutants were shown to produce more biofilm than the wild-type control while the PNPase mutant did not form biofilms. In this work we demonstrate how deep sequencing can be used to discover new and relevant functions of the exoribonucleases. We were able to obtain valuable information about the transcripts affected by each of the exoribonucleases and compare the roles of the three enzymes. Our results show that the three exoribonucleases affect cell motility and biofilm formation that are two very important factors for cell survival, especially for pathogenic cells.

Inoculation density and nutrient level determine the formation of mushroom-shaped structures in Pseudomonas aeruginosa biofilms

NASA Astrophysics Data System (ADS)

Ghanbari, Azadeh; Dehghany, Jaber; Schwebs, Timo; Müsken, Mathias; Häussler, Susanne; Meyer-Hermann, Michael

2016-09-01

Pseudomonas aeruginosa often colonises immunocompromised patients and the lungs of cystic fibrosis patients. It exhibits resistance to many antibiotics by forming biofilms, which makes it hard to eliminate. P. aeruginosa biofilms form mushroom-shaped structures under certain circumstances. Bacterial motility and the environment affect the eventual mushroom morphology. This study provides an agent-based model for the bacterial dynamics and interactions influencing bacterial biofilm shape. Cell motility in the model relies on recently published experimental data. Our simulations show colony formation by immotile cells. Motile cells escape from a single colony by nutrient chemotaxis and hence no mushroom shape develops. A high number density of non-motile colonies leads to migration of motile cells onto the top of the colonies and formation of mushroom-shaped structures. This model proposes that the formation of mushroom-shaped structures can be predicted by parameters at the time of bacteria inoculation. Depending on nutrient levels and the initial number density of stalks, mushroom-shaped structures only form in a restricted regime. This opens the possibility of early manipulation of spatial pattern formation in bacterial colonies, using environmental factors.

Inoculation density and nutrient level determine the formation of mushroom-shaped structures in Pseudomonas aeruginosa biofilms.

PubMed

Ghanbari, Azadeh; Dehghany, Jaber; Schwebs, Timo; Müsken, Mathias; Häussler, Susanne; Meyer-Hermann, Michael

2016-09-09

Pseudomonas aeruginosa often colonises immunocompromised patients and the lungs of cystic fibrosis patients. It exhibits resistance to many antibiotics by forming biofilms, which makes it hard to eliminate. P. aeruginosa biofilms form mushroom-shaped structures under certain circumstances. Bacterial motility and the environment affect the eventual mushroom morphology. This study provides an agent-based model for the bacterial dynamics and interactions influencing bacterial biofilm shape. Cell motility in the model relies on recently published experimental data. Our simulations show colony formation by immotile cells. Motile cells escape from a single colony by nutrient chemotaxis and hence no mushroom shape develops. A high number density of non-motile colonies leads to migration of motile cells onto the top of the colonies and formation of mushroom-shaped structures. This model proposes that the formation of mushroom-shaped structures can be predicted by parameters at the time of bacteria inoculation. Depending on nutrient levels and the initial number density of stalks, mushroom-shaped structures only form in a restricted regime. This opens the possibility of early manipulation of spatial pattern formation in bacterial colonies, using environmental factors.

Relationships between rabbit semen characteristics and fertilising ability after insemination.

PubMed

Theau-Clément, M; Ailloud, E; Sanchez, A; Saleil, G; Brun, J M

2016-03-01

This study aimed to analyse the relationship between rabbit semen characteristics and semen fertilising ability after insemination, which is generally found to be weak. Our hypothesis was that using high semen dilutions (1 : 19), non-oestrus-stimulated does, and homospermic inseminations would make it easier to predict semen fertilising ability. Semen characteristics were evaluated on 275 ejaculates of 128 INRA1001 bucks, distributed into five successive batches. A total of 1970 inseminations were performed. The continuous semen variables were subdivided into three classes of similar size to account for any non-linear relationship between semen characteristics and fertilising ability. Mass motility was divided into two classes according to the presence or absence of waves under microscope observation. Libido, the presence or absence of gel, volume, percentage of progressive sperms, curvilinear velocity, beat frequency of the flagellum, and straightness and linearity of sperm movement did not affect fertility, prolificacy or productivity. It was confirmed that mass motility, estimated by visual observation under the microscope, significantly influenced fertility as well as the percentage of motile and of rapid sperms, and the amplitude of lateral head displacement, estimated by a computer-assisted semen analysis system. To a lesser extent, the percentage of motile cells and of rapid cells significantly influenced prolificacy. Consequently, mass motility and the percentage of motile cells significantly influenced rabbit doe productivity (+1 live births/AI when the semen showed at least a beginning of wave movement, or when the percentage of motile cells was >84%). Interestingly, a gain of 1.5 rabbits was observed when the percentage of rapid cells changed from 64% to 79%, whereas productivity significantly dropped beyond 83% of rapid cells, reflecting a non-linear relationship.

Determinants of the epithelial-muscular axis on embryonic stem cell-derived gut-like structures.

PubMed

Luo, Yi; Takaki, Miyako; Misawa, Hiromi; Matsuyoshi, Hiroko; Sasahira, Tomonori; Chihara, Yoshitomo; Fujii, Kiyomu; Ohmori, Hitoshi; Kuniyasu, Hiroki

2010-01-01

Dome-like structures with epithelial-muscular layers resembling the gut have been derived from mouse embryonic stem (ES) cells. These domes have been reported to show spontaneous contractions and are called ES gut. In the present study, we examined the epithelial-muscular axis of these domes by detecting differentiation markers. A normal epithelial-muscular axis was exhibited in the domes with spontaneous motility, whereas the domes without spontaneous motility showed either an inverted or obscure axis. To investigate the factors affecting the epithelial-muscular axis, we examined the expression of hedgehog signaling factors in the domes. Expression of hedgehog family factors was detected in the epithelial components of the domes with motility, whereas this expression was inverted or obscure in the domes without motility. Out of the 25 domes, 10 of the 10 motility (+) domes showed a normal epithelial-muscular axis, whereas 14 of the 15 motility (-) domes lacked a normal epithelial-muscular axis. This implies that activin A upregulated the expression of sonic hedgehog and intestinal alkaline phosphatase in the embryoid bodies. These findings suggest that the motility of the ES gut depends on the domes' epithelial-muscular axis. Copyright © 2010 S. Karger AG, Basel.

Metabolic Adaptations of Azospirillum brasilense to Oxygen Stress by Cell-to-Cell Clumping and Flocculation

PubMed Central

Bible, Amber N.; Khalsa-Moyers, Gurusahai K.; Mukherjee, Tanmoy; Green, Calvin S.; Mishra, Priyanka; Purcell, Alicia; Aksenova, Anastasia; Hurst, Gregory B.

2015-01-01

The ability of bacteria to monitor their metabolism and adjust their behavior accordingly is critical to maintain competitiveness in the environment. The motile microaerophilic bacterium Azospirillum brasilense navigates oxygen gradients by aerotaxis in order to locate low oxygen concentrations that can support metabolism. When cells are exposed to elevated levels of oxygen in their surroundings, motile A. brasilense cells implement an alternative response to aerotaxis and form transient clumps by cell-to-cell interactions. Clumping was suggested to represent a behavior protecting motile cells from transiently elevated levels of aeration. Using the proteomics of wild-type and mutant strains affected in the extent of their clumping abilities, we show that cell-to-cell clumping represents a metabolic scavenging strategy that likely prepares the cells for further metabolic stresses. Analysis of mutants affected in carbon or nitrogen metabolism confirmed this assumption. The metabolic changes experienced as clumping progresses prime cells for flocculation, a morphological and metabolic shift of cells triggered under elevated-aeration conditions and nitrogen limitation. The analysis of various mutants during clumping and flocculation characterized an ordered set of changes in cell envelope properties accompanying the metabolic changes. These data also identify clumping and early flocculation to be behaviors compatible with the expression of nitrogen fixation genes, despite the elevated-aeration conditions. Cell-to-cell clumping may thus license diazotrophy to microaerophilic A. brasilense cells under elevated oxygen conditions and prime them for long-term survival via flocculation if metabolic stress persists. PMID:26407887

A Nutrient-Tunable Bistable Switch Controls Motility in Salmonella enterica Serovar Typhimurium

PubMed Central

Koirala, Santosh; Mears, Patrick; Sim, Martin; Golding, Ido; Chemla, Yann R.; Aldridge, Phillip D.

2014-01-01

ABSTRACT Many bacteria are motile only when nutrients are scarce. In contrast, Salmonella enterica serovar Typhimurium is motile only when nutrients are plentiful, suggesting that this bacterium uses motility for purposes other than foraging, most likely for host colonization. In this study, we investigated how nutrients affect motility in S. enterica and found that they tune the fraction of motile cells. In particular, we observed coexisting populations of motile and nonmotile cells, with the distribution being determined by the concentration of nutrients in the growth medium. Interestingly, S. enterica responds not to a single nutrient but apparently to a complex mixture of them. Using a combination of experimentation and mathematical modeling, we investigated the mechanism governing this behavior and found that it results from two antagonizing regulatory proteins, FliZ and YdiV. We also found that a positive feedback loop involving the alternate sigma factor FliA is required, although its role appears solely to amplify FliZ expression. We further demonstrate that the response is bistable: that is, genetically identical cells can exhibit different phenotypes under identical growth conditions. Together, these results uncover a new facet of the regulation of the flagellar genes in S. enterica and further demonstrate how bacteria employ phenotypic diversity as a general mechanism for adapting to change in their environment. PMID:25161191

Divalent Cation Control of Flagellar Motility in African Trypanosomes

NASA Astrophysics Data System (ADS)

Westergard, Anna M.; Hutchings, Nathan R.

2005-03-01

Changes in calcium concentration have been shown to dynamically affect flagellar motility in several eukaryotic systems. The African trypanosome is a monoflagellated protozoan parasite and the etiological agent of sleeping sickness. Although cell motility has been implicated in disease progression, very little is currently known about biochemical control of the trypanosome flagellum. In this study, we assess the effects of extracellular changes in calcium and nickel concentration on trypanosome flagellar movement. Using a flow through chamber, we determine the relative changes in motility in individual trypanosomes in response to various concentrations of calcium and nickel, respectively. Extracellular concentrations of calcium and nickel (as low as 100 micromolar) significantly inhibit trypanosome cell motility. The effects are reversible, as indicated by the recovery of motion after removal of the calcium or nickel from the chamber. We are currently investigating the specific changes in flagellar oscillation and coordination that result from calcium and nickel, respectively. These results verify the presence of a calcium-responsive signaling mechanism(s) that regulates flagellar beat in trypanosomes.

Metabolic Adaptations of Azospirillum brasilense to Oxygen Stress by Cell-to-Cell Clumping and Flocculation

DOE PAGES

Bible, Amber N.; Khalsa-Moyers, Gurusahai K.; Mukherjee, Tanmoy; ...

2015-09-25

The ability of bacteria to monitor their metabolism and adjust their behavior accordingly is critical to maintain competitiveness in the environment. The motile microaerophilic bacteriumAzospirillum brasilensenavigates oxygen gradients by aerotaxis in order to locate low oxygen concentrations that can support metabolism. When cells are exposed to elevated levels of oxygen in their surroundings, motileA. brasilensecells implement an alternative response to aerotaxis and form transient clumps by cell-to-cell interactions. Clumping was suggested to represent a behavior protecting motile cells from transiently elevated levels of aeration. Using the proteomics of wild-type and mutant strains affected in the extent of their clumping abilities,more » we show that cell-to-cell clumping represents a metabolic scavenging strategy that likely prepares the cells for further metabolic stresses. Analysis of mutants affected in carbon or nitrogen metabolism confirmed this assumption. The metabolic changes experienced as clumping progresses prime cells for flocculation, a morphological and metabolic shift of cells triggered under elevated-aeration conditions and nitrogen limitation. The analysis of various mutants during clumping and flocculation characterized an ordered set of changes in cell envelope properties accompanying the metabolic changes. These data also identify clumping and early flocculation to be behaviors compatible with the expression of nitrogen fixation genes, despite the elevated-aeration conditions. Finally, cell-to-cell clumping may thus license diazotrophy to microaerophilicA. brasilensecells under elevated oxygen conditions and prime them for long-term survival via flocculation if metabolic stress persists.« less

Metabolic Adaptations of Azospirillum brasilense to Oxygen Stress by Cell-to-Cell Clumping and Flocculation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bible, Amber N.; Khalsa-Moyers, Gurusahai K.; Mukherjee, Tanmoy

The ability of bacteria to monitor their metabolism and adjust their behavior accordingly is critical to maintain competitiveness in the environment. The motile microaerophilic bacteriumAzospirillum brasilensenavigates oxygen gradients by aerotaxis in order to locate low oxygen concentrations that can support metabolism. When cells are exposed to elevated levels of oxygen in their surroundings, motileA. brasilensecells implement an alternative response to aerotaxis and form transient clumps by cell-to-cell interactions. Clumping was suggested to represent a behavior protecting motile cells from transiently elevated levels of aeration. Using the proteomics of wild-type and mutant strains affected in the extent of their clumping abilities,more » we show that cell-to-cell clumping represents a metabolic scavenging strategy that likely prepares the cells for further metabolic stresses. Analysis of mutants affected in carbon or nitrogen metabolism confirmed this assumption. The metabolic changes experienced as clumping progresses prime cells for flocculation, a morphological and metabolic shift of cells triggered under elevated-aeration conditions and nitrogen limitation. The analysis of various mutants during clumping and flocculation characterized an ordered set of changes in cell envelope properties accompanying the metabolic changes. These data also identify clumping and early flocculation to be behaviors compatible with the expression of nitrogen fixation genes, despite the elevated-aeration conditions. Finally, cell-to-cell clumping may thus license diazotrophy to microaerophilicA. brasilensecells under elevated oxygen conditions and prime them for long-term survival via flocculation if metabolic stress persists.« less

Characteristics of sperm motility in boar semen diluted in different extenders and stored for seven days at 18 degrees C.

PubMed

Estienne, Mark J; Harper, Allen F; Day, Jennifer L

2007-11-01

Although numerous extenders exist for diluting boar semen, little research has been conducted comparing commercial extenders with regard to maintaining sperm motility during storage. The objective was to use a computer- assisted sperm analysis system to assess motility of boar spermatozoa diluted in Beltsville Thawing Solution, Merck-III, Androhep-lite, Sperm Aid, MR-A, Modena, X-Cell, VSP, and Vital. Ejaculates from boars (n=10) were collected and sub-samples were diluted (35x10(6) spermatozoa/ml) in the different extenders and stored for seven days at 18 degrees. Extender by day interactions were detected (p<0.01) and on each day post collection, there were numerically small, but statistically significant differences in characteristics of sperm motility among extenders. For example, on day 7, the percentages of motile and progressively motile spermatozoa were highest (p<0.05) in X-Cell (90.7%) and Modena (63.9%), respectively. The average velocity measured over the actual point-to-point track followed by the sperm cell (VCL; 198.2 microm/s) and path velocity of the smoothed cell path (VAP; 106.4 microm/s) were highest (p<0.05) in Vital and Modena, respectively. Average velocity measured in a straight line from the beginning to the end of the track (VSL; 78.3 microm/s), average value of the ratio VSL/VAP (straightness; 73.2) and average value of the ratio VSL/VCL (linearity; 44.1) on day 7 were highest in Androhep-lite. In summary, changes in sperm motility during storage were affected by the extender utilized, but with the exception of Sperm Aid, all extenders maintained a high degree of sperm motility through 7 days of storage.

Cancer Cell Glycocalyx Mediates Mechanostransduction and Flow-Regulated Invasion

PubMed Central

Qazi, Henry; Palomino, Rocio; Shi, Zhong-Dong; Munn, Lance L.; Tarbell, John M.

2014-01-01

Mammalian cells are covered by a surface proteoglycan (glycocalyx) layer, and it is known that blood vessel-lining endothelial cells use the glycocalyx to sense and transduce the shearing forces of blood flow into intracellular signals. Tumor cells in vivo are exposed to forces from interstitial fluid flow that may affect metastatic potential but are not reproduced by most in vitro cell motility assays. We hypothesized that glycocalyx-mediated mechanotransduction of interstitial flow shear stress is an un-recognized factor that can significantly enhance metastatic cell motility and play a role in augmentation of invasion. Involvement of MMP levels, cell adhesion molecules (CD44, α3 integrin), and glycocalyx components (heparan sulfate and hyaluronan) were investigated in a cell/collagen gel suspension model designed to mimic the interstitial flow microenvironment. Physiologic levels of flow upregulated MMP levels and enhanced the motility of metastatic cells. Blocking the flow-enhanced expression of MMP actvity or adhesion molecules (CD44 and integrins) resulted in blocking the flow-enhanced migratory activity. The presence of a glycocalyx-like layer was verified around tumor cells, and the degradation of this layer by hyaluronidase and heparinase blocked the flow-regulated invasion. This study shows for the first time that interstitial flow enhancement of metastatic cell motility can be mediated by the cell surface glycocalyx – a potential target for therapeutics. PMID:24077103

PTP-PEST targets a novel tyrosine site in p120 catenin to control epithelial cell motility and Rho GTPase activity.

PubMed

Espejo, Rosario; Jeng, Yowjiun; Paulucci-Holthauzen, Adriana; Rengifo-Cam, William; Honkus, Krysta; Anastasiadis, Panos Z; Sastry, Sarita K

2014-02-01

Tyrosine phosphorylation is implicated in regulating the adherens junction protein, p120 catenin (p120), however, the mechanisms are not well defined. Here, we show, using substrate trapping, that p120 is a direct target of the protein tyrosine phosphatase, PTP-PEST, in epithelial cells. Stable shRNA knockdown of PTP-PEST in colon carcinoma cells results in an increased cytosolic pool of p120 concomitant with its enhanced tyrosine phosphorylation and decreased association with E-cadherin. Consistent with this, PTP-PEST knockdown cells exhibit increased motility, enhanced Rac1 and decreased RhoA activity on a collagen substrate. Furthermore, p120 localization is enhanced at actin-rich protrusions and lamellipodia and has an increased association with the guanine nucleotide exchange factor, VAV2, and cortactin. Exchange factor activity of VAV2 is enhanced by PTP-PEST knockdown whereas overexpression of a VAV2 C-terminal domain or DH domain mutant blocks cell motility. Analysis of point mutations identified tyrosine 335 in the N-terminal domain of p120 as the site of PTP-PEST dephosphorylation. A Y335F mutant of p120 failed to induce the 'p120 phenotype', interact with VAV2, stimulate cell motility or activate Rac1. Together, these data suggest that PTP-PEST affects epithelial cell motility by controlling the distribution and phosphorylation of p120 and its availability to control Rho GTPase activity.

PTP-PEST targets a novel tyrosine site in p120 catenin to control epithelial cell motility and Rho GTPase activity

PubMed Central

Espejo, Rosario; Jeng, Yowjiun; Paulucci-Holthauzen, Adriana; Rengifo-Cam, William; Honkus, Krysta; Anastasiadis, Panos Z.; Sastry, Sarita K.

2014-01-01

ABSTRACT Tyrosine phosphorylation is implicated in regulating the adherens junction protein, p120 catenin (p120), however, the mechanisms are not well defined. Here, we show, using substrate trapping, that p120 is a direct target of the protein tyrosine phosphatase, PTP-PEST, in epithelial cells. Stable shRNA knockdown of PTP-PEST in colon carcinoma cells results in an increased cytosolic pool of p120 concomitant with its enhanced tyrosine phosphorylation and decreased association with E-cadherin. Consistent with this, PTP-PEST knockdown cells exhibit increased motility, enhanced Rac1 and decreased RhoA activity on a collagen substrate. Furthermore, p120 localization is enhanced at actin-rich protrusions and lamellipodia and has an increased association with the guanine nucleotide exchange factor, VAV2, and cortactin. Exchange factor activity of VAV2 is enhanced by PTP-PEST knockdown whereas overexpression of a VAV2 C-terminal domain or DH domain mutant blocks cell motility. Analysis of point mutations identified tyrosine 335 in the N-terminal domain of p120 as the site of PTP-PEST dephosphorylation. A Y335F mutant of p120 failed to induce the ‘p120 phenotype’, interact with VAV2, stimulate cell motility or activate Rac1. Together, these data suggest that PTP-PEST affects epithelial cell motility by controlling the distribution and phosphorylation of p120 and its availability to control Rho GTPase activity. PMID:24284071

Exosome secretion affects social motility in Trypanosoma brucei

PubMed Central

Shaked, Hadassa; Arvatz, Gil; Tkacz, Itai Dov; Binder, Lior; Waldman Ben-Asher, Hiba; Okalang, Uthman; Chikne, Vaibhav; Cohen-Chalamish, Smadar; Michaeli, Shulamit

2017-01-01

Extracellular vesicles (EV) secreted by pathogens function in a variety of biological processes. Here, we demonstrate that in the protozoan parasite Trypanosoma brucei, exosome secretion is induced by stress that affects trans-splicing. Following perturbations in biogenesis of spliced leader RNA, which donates its spliced leader (SL) exon to all mRNAs, or after heat-shock, the SL RNA is exported to the cytoplasm and forms distinct granules, which are then secreted by exosomes. The exosomes are formed in multivesicular bodies (MVB) utilizing the endosomal sorting complexes required for transport (ESCRT), through a mechanism similar to microRNA secretion in mammalian cells. Silencing of the ESCRT factor, Vps36, compromised exosome secretion but not the secretion of vesicles derived from nanotubes. The exosomes enter recipient trypanosome cells. Time-lapse microscopy demonstrated that cells secreting exosomes or purified intact exosomes affect social motility (SoMo). This study demonstrates that exosomes are delivered to trypanosome cells and can change their migration. Exosomes are used to transmit stress signals for communication between parasites. PMID:28257521

Cell cycles and proliferation patterns in Haematococcus pluvialis

NASA Astrophysics Data System (ADS)

Zhang, Chunhui; Liu, Jianguo; Zhang, Litao

2017-09-01

Most studies on Haematococcus pluvialis have been focused on cell growth and astaxanthin accumulation; far less attention has been paid to cell cycles and proliferation patterns. The purpose of this study was to clarify cell cycles and proliferation patterns in H. pluvialis microscopically using a camera and video recorder system. The complicated life history of H. pluvialis can be divided into two stages: the motile stage and the non-motile stage. All the cells can be classified into forms as follows: motile cell, nonmotile cell, zoospore and aplanospore. The main cell proliferation, both in the motile phase and non-motile phase in H. pluvialis, is by asexual reproduction. Under normal growth conditions, a motile cell usually produces two, sometimes four, and exceptionally eight zoospores. Under unfavorable conditions, the motile cell loses its flagella and transforms into a non-motile cell, and the non-motile cell usually produces 2, 4 or 8 aplanospores, and occasionally 20-32 aplanospores, which further develop into non-motile cells. Under suitable conditions, the non-motile cell is also able to release zoospores. The larger non-motile cells produce more than 16 zoospores, and the smaller ones produce 4 or 8 zoospores. Vegetative reproduction is by direct cell division in the motile phase and by occasional cell budding in the non-motile phase. There is, as yet, no convincing direct evidence for sexual reproduction.

Slits Affect the Timely Migration of Neural Crest Cells via Robo Receptor

PubMed Central

Giovannone, Dion; Reyes, Michelle; Reyes, Rachel; Correa, Lisa; Martinez, Darwin; Ra, Hannah; Gomez, Gustavo; Kaiser, Josh; Ma, Le; Stein, Mary-Pat; de Bellard, Maria Elena

2013-01-01

SUMMARY Background Neural crest cells emerge by delamination from the dorsal neural tube and give rise to various components of the peripheral nervous system in vertebrate embryos. These cells change from non-motile into highly motile cells migrating to distant areas before further differentiation. Mechanisms controlling delamination and subsequent migration of neural crest cells are not fully understood. Slit2, a chemorepellant for axonal guidance that repels and stimulates motility of trunk neural crest cells away from the gut has recently been suggested to be a tumor suppressor molecule. The goal of this study was to further investigate the role of Slit2 in trunk neural crest cell migration by constitutive expression in neural crest cells. Results We found that Slit gain-of-function significantly impaired neural crest cell migration while Slit loss-of-function favored migration. In addition, we observed that the distribution of key cytoskeletal markers was disrupted in both gain and loss of function instances. Conclusions These findings suggest that Slit molecules might be involved in the processes that allow neural crest cells to begin migration and transitioning to a mesenchymal type. PMID:22689303

Oxidative Damage to Rhesus Macaque Spermatozoa Results in Mitotic Arrest and Transcript Abundance Changes in Early Embryos1

PubMed Central

Burruel, Victoria; Klooster, Katie L.; Chitwood, James; Ross, Pablo J.; Meyers, Stuart A.

2013-01-01

ABSTRACT Our objective was to determine whether oxidative damage of rhesus macaque sperm induced by reactive oxygen species (ROS) in vitro would affect embryo development following intracytoplasmic sperm injection (ICSI) of metaphase II (MII) oocytes. Fresh rhesus macaque spermatozoa were treated with ROS as follows: 1 mM xanthine and 0.1 U/ml xanthine oxidase (XXO) at 37°C and 5% CO2 in air for 2.25 h. Sperm were then assessed for motility, viability, and lipid peroxidation. Motile ROS-treated and control sperm were used for ICSI of MII oocytes. Embryo culture was evaluated for 3 days for development to the eight-cell stage. Embryos were fixed and stained for signs of cytoplasmic and nuclear abnormalities. Gene expression was analyzed by RNA-Seq in two-cell embryos from control and treated groups. Exposure of sperm to XXO resulted in increased lipid peroxidation and decreased sperm motility. ICSI of MII oocytes with motile sperm induced similar rates of fertilization and cleavage between treatments. Development to four- and eight-cell stage was significantly lower for embryos generated with ROS-treated sperm than for controls. All embryos produced from ROS-treated sperm demonstrated permanent embryonic arrest and varying degrees of degeneration and nuclear fragmentation, changes that are suggestive of prolonged senescence or apoptotic cell death. RNA-Seq analysis of two-cell embryos showed changes in transcript abundance resulting from sperm treatment with ROS. Differentially expressed genes were enriched for processes associated with cytoskeletal organization, cell adhesion, and protein phosphorylation. ROS-induced damage to sperm adversely affects embryo development by contributing to mitotic arrest after ICSI of MII rhesus oocytes. Changes in transcript abundance in embryos destined for mitotic arrest is evident at the two-cell stage of development. PMID:23904511

Merkel Cell Polyomavirus Small T Antigen Drives Cell Motility via Rho-GTPase-Induced Filopodium Formation.

PubMed

Stakaitytė, Gabrielė; Nwogu, Nnenna; Dobson, Samuel J; Knight, Laura M; Wasson, Christopher W; Salguero, Francisco J; Blackbourn, David J; Blair, G Eric; Mankouri, Jamel; Macdonald, Andrew; Whitehouse, Adrian

2018-01-15

Cell motility and migration is a complex, multistep, and multicomponent process intrinsic to progression and metastasis. Motility is dependent on the activities of integrin receptors and Rho family GTPases, resulting in the remodeling of the actin cytoskeleton and formation of various motile actin-based protrusions. Merkel cell carcinoma (MCC) is an aggressive skin cancer with a high likelihood of recurrence and metastasis. Merkel cell polyomavirus (MCPyV) is associated with the majority of MCC cases, and MCPyV-induced tumorigenesis largely depends on the expression of the small tumor antigen (ST). Since the discovery of MCPyV, a number of mechanisms have been suggested to account for replication and tumorigenesis, but to date, little is known about potential links between MCPyV T antigen expression and the metastatic nature of MCC. Previously, we described the action of MCPyV ST on the microtubule network and how it impacts cell motility and migration. Here, we demonstrate that MCPyV ST affects the actin cytoskeleton to promote the formation of filopodia through a mechanism involving the catalytic subunit of protein phosphatase 4 (PP4C). We also show that MCPyV ST-induced cell motility is dependent upon the activities of the Rho family GTPases Cdc42 and RhoA. In addition, our results indicate that the MCPyV ST-PP4C interaction results in the dephosphorylation of β 1 integrin, likely driving the cell motility pathway. These findings describe a novel mechanism by which a tumor virus induces cell motility, which may ultimately lead to cancer metastasis, and provides opportunities and strategies for targeted interventions for disseminated MCC. IMPORTANCE Merkel cell polyomavirus (MCPyV) is the most recently discovered human tumor virus. It causes the majority of cases of Merkel cell carcinoma (MCC), an aggressive skin cancer. However, the molecular mechanisms implicating MCPyV-encoded proteins in cancer development are yet to be fully elucidated. This study builds upon our previous observations, which demonstrated that the MCPyV ST antigen enhances cell motility, providing a potential link between MCPyV protein expression and the highly metastatic nature of MCC. Here, we show that MCPyV ST remodels the actin cytoskeleton, promoting the formation of filopodia, which is essential for MCPyV ST-induced cell motility, and we also implicate the activity of specific Rho family GTPases, Cdc42 and RhoA, in these processes. Moreover, we describe a novel mechanism for the activation of Rho-GTPases and the cell motility pathway due to the interaction between MCPyV ST and the cellular phosphatase catalytic subunit PP4C, which leads to the specific dephosphorylation of β1 integrin. These findings may therefore provide novel strategies for therapeutic intervention for disseminated MCC. Copyright © 2018 Stakaitytė et al.

MRP-1/CD9 gene transduction regulates the actin cytoskeleton through the downregulation of WAVE2.

PubMed

Huang, C-L; Ueno, M; Liu, D; Masuya, D; Nakano, J; Yokomise, H; Nakagawa, T; Miyake, M

2006-10-19

Motility-related protein-1 (MRP-1/CD9) is involved in cell motility. We studied the change in the actin cytoskeleton, and the expression of actin-related protein (Arp) 2 and Arp3 and the Wiskott-Aldrich syndrome protein (WASP) family according to MRP-1/CD9 gene transduction into HT1080 cells. The frequency of cells with lamellipodia was significantly lower in MRP-1/CD9-transfected HT1080 cells than in control HT1080 cells (P<0.0001). MRP-1/CD9 gene transduction affected the subcellular localization of Arp2 and Arp3 proteins. Furthermore, MRP-1/CD9 gene transduction induced a downregulation of WAVE2 expression (P<0.0001). However, no difference was observed in the expression of Arp2, Arp3 or other WASPs. A neutralizing anti-MRP-1/CD9 monoclonal antibody inhibited downregulation of WAVE2 in MRP-1/CD9-transfected HT1080 cells (P<0.0001), and reversed the morphological effects of MRP-1/CD9 gene transduction. Furthermore, downregulation of WAVE2 by transfection of WAVE2-specific small interfering RNA (siRNA) mimicked the morphological effects of MRP-1/CD9 gene transduction and suppressed cell motility. However, transfection of each siRNA for Wnt1, Wnt2b1 or Wnt5a did not affect WAVE2 expression. Transfection of WAVE2-specific siRNA also did not affect expressions of these Wnts. These results indicate that MRP-1/CD9 regulates the actin cytoskeleton by downregulating of the WAVE2, through the Wnt-independent signal pathway.

Loss of ascl1a prevents secretory cell differentiation within the zebrafish intestinal epithelium resulting in a loss of distal intestinal motility

PubMed Central

Roach, Gillian; Wallace, Rachel Heath; Cameron, Amy; Ozel, Rifat Emrah; Hongay, Cintia F.; Baral, Reshica; Andreescu, Silvana; Wallace, Kenneth N.

2013-01-01

The vertebrate intestinal epithelium is renewed continuously from stem cells at the base of the crypt in mammals or base of the fold in fish over the life of the organism. As stem cells divide, newly formed epithelial cells make an initial choice between a secretory or enterocyte fate. This choice has previously been demonstrated to involve Notch signaling as well as Atonal and Her transcription factors in both embryogenesis and adults. Here, we demonstrate that in contrast to the atoh1 in mammals, ascl1a is responsible for formation of secretory cells in zebrafish. ascl1a−/− embryos lack all intestinal epithelial secretory cells and instead differentiate into enterocytes. ascl1a−/− embryos also fail to induce intestinal epithelial expression of deltaD suggesting that ascl1a plays a role in initiation of Notch signaling. Inhibition of Notch signaling increases the number of ascl1a and deltaD expressing intestinal epithelial cells as well as the number of developing secretory cells during two specific time periods: between 30 and 34 hpf and again between 64 and 74 hpf. Loss of enteroendocrine products results in loss of anterograde motility in ascl1a−/− embryos. 5HT produced by enterochromaffin cells is critical in motility and secretion within the intestine. We find that addition of exogenous 5HT to ascl1a−/− embryos at near physiological levels (measured by differential pulse voltammetry) induce anterograde motility at similar levels to wild type velocity, distance, and frequency. Removal or doubling the concentration of 5HT in WT embryos does not significantly affect anterograde motility, suggesting that the loss of additional enteroendocrine products in ascl1a−/− embryos also contributes to intestinal motility. Thus, zebrafish intestinal epithelial cells appear to have a common secretory progenitor from which all subtypes form. Loss of enteroendocrine cells reveals the critical need for enteroendocrine products in maintenance of normal intestinal motility. PMID:23353550

Myeloperoxidase Oxidized LDL Interferes with Endothelial Cell Motility through miR-22 and Heme Oxygenase 1 Induction: Possible Involvement in Reendothelialization of Vascular Injuries

PubMed Central

Daher, Jalil; Martin, Maud; Rousseau, Alexandre; Nuyens, Vincent; Fayyad-Kazan, Hussein; Van Antwerpen, Pierre; Courbebaisse, Guy; Martiat, Philippe; Badran, Bassam; Dequiedt, Frank

2014-01-01

Cardiovascular disease linked to atherosclerosis is the leading cause of death worldwide. Atherosclerosis is mainly linked to dysfunction in vascular endothelial cells and subendothelial accumulation of oxidized forms of LDL. In the present study, we investigated the role of myeloperoxidase oxidized LDL (Mox-LDL) in endothelial cell dysfunction. We studied the effect of proinflammatory Mox-LDL treatment on endothelial cell motility, a parameter essential for normal vascular processes such as angiogenesis and blood vessel repair. This is particularly important in the context of an atheroma plaque, where vascular wall integrity is affected and interference with its repair could contribute to progression of the disease. We investigated in vitro the effect of Mox-LDL on endothelial cells angiogenic properties and we also studied the signalling pathways that could be affected by analysing Mox-LDL effect on the expression of angiogenesis-related genes. We report that Mox-LDL inhibits endothelial cell motility and tubulogenesis through an increase in miR-22 and heme oxygenase 1 expression. Our in vitro data indicate that Mox-LDL interferes with parameters associated with angiogenesis. They suggest that high LDL levels in patients would impair their endothelial cell capacity to cope with a damaged endothelium contributing negatively to the progression of the atheroma plaque. PMID:25530680

Dose-response effects of estrogenic mycotoxins (zearalenone, alpha- and beta-zearalenol) on motility, hyperactivation and the acrosome reaction of stallion sperm

PubMed Central

2011-01-01

Background The aim of this study was to investigate the in vitro effects of the Fusarium fungus-derived mycotoxin, zearalenone and its derivatives alpha-zearalenol and beta-zearalenol on motility parameters and the acrosome reaction of stallion sperm. Since the toxic effects of zearalenone and its derivatives are thought to result from their structural similarity to 17beta-estradiol, 17beta-estradiol was used as a positive control for 'estrogen-like' effects. Methods Stallion spermatozoa were exposed in vitro to zearalenone, alpha-zearalenol, beta-zearalenol or 17beta-estradiol at concentrations ranging from 1 pM - 0.1 mM. After 2 hours exposure, motility parameters were evaluated by computer-assisted analysis, and acrosome integrity was examined by flow cytometry after staining with fluoroscein-conjugated peanut agglutinin. Results Mycotoxins affected sperm parameters only at the highest concentration tested (0.1 mM) after 2 hours exposure. In this respect, all of the compounds reduced the average path velocity, but only alpha-zearalenol reduced percentages of motile and progressively motile sperm. Induction of motility patterns consistent with hyperactivation was stimulated according to the following rank of potency: alpha-zearalenol >17beta-estradiol > zearalenone = beta-zearalenol. The hyperactivity-associated changes observed included reductions in straight-line velocity and linearity of movement, and an increase in the amplitude of lateral head displacement, while curvilinear velocity was unchanged. In addition, whereas alpha- and beta- zearalenol increased the percentages of live acrosome-reacted sperm, zearalenone and 17beta-estradiol had no apparent effect on acrosome status. In short, alpha-zearalenol inhibited normal sperm motility, but stimulated hyperactive motility in the remaining motile cells and simultaneously induced the acrosome reaction. Beta-zearalenol induced the acrosome reaction without altering motility. Conversely, zearalenone and 17beta-estradiol did not induce the acrosome reaction but induced hyperactive motility albeit to a different extent. Conclusions Apparently, the mycotoxin zearalenone has 17beta-estradiol-like estrogenic activity that enables it to induce hyperactivated motility of equine sperm cells, whereas the zearalenol derivatives induce premature completion of the acrosome reaction and thereby adversely affect stallion sperm physiology. The alpha form of zearalenol still possessed the estrogenic ability to induce hyperactivated motility, whereas its beta stereo-isomere had lost this property. PMID:21970729

Metabolic adaptations of Azospirillum brasilense to oxygen stress by cell-to-cell clumping and flocculation.

PubMed

Bible, Amber N; Khalsa-Moyers, Gurusahai K; Mukherjee, Tanmoy; Green, Calvin S; Mishra, Priyanka; Purcell, Alicia; Aksenova, Anastasia; Hurst, Gregory B; Alexandre, Gladys

2015-12-01

The ability of bacteria to monitor their metabolism and adjust their behavior accordingly is critical to maintain competitiveness in the environment. The motile microaerophilic bacterium Azospirillum brasilense navigates oxygen gradients by aerotaxis in order to locate low oxygen concentrations that can support metabolism. When cells are exposed to elevated levels of oxygen in their surroundings, motile A. brasilense cells implement an alternative response to aerotaxis and form transient clumps by cell-to-cell interactions. Clumping was suggested to represent a behavior protecting motile cells from transiently elevated levels of aeration. Using the proteomics of wild-type and mutant strains affected in the extent of their clumping abilities, we show that cell-to-cell clumping represents a metabolic scavenging strategy that likely prepares the cells for further metabolic stresses. Analysis of mutants affected in carbon or nitrogen metabolism confirmed this assumption. The metabolic changes experienced as clumping progresses prime cells for flocculation, a morphological and metabolic shift of cells triggered under elevated-aeration conditions and nitrogen limitation. The analysis of various mutants during clumping and flocculation characterized an ordered set of changes in cell envelope properties accompanying the metabolic changes. These data also identify clumping and early flocculation to be behaviors compatible with the expression of nitrogen fixation genes, despite the elevated-aeration conditions. Cell-to-cell clumping may thus license diazotrophy to microaerophilic A. brasilense cells under elevated oxygen conditions and prime them for long-term survival via flocculation if metabolic stress persists. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Ionic imbalance, in addition to molecular crowding, abates cytoskeletal dynamics and vesicle motility during hypertonic stress

PubMed Central

Nunes, Paula; Roth, Isabelle; Meda, Paolo; Féraille, Eric; Brown, Dennis; Hasler, Udo

2015-01-01

Cell volume homeostasis is vital for the maintenance of optimal protein density and cellular function. Numerous mammalian cell types are routinely exposed to acute hypertonic challenge and shrink. Molecular crowding modifies biochemical reaction rates and decreases macromolecule diffusion. Cell volume is restored rapidly by ion influx but at the expense of elevated intracellular sodium and chloride levels that persist long after challenge. Although recent studies have highlighted the role of molecular crowding on the effects of hypertonicity, the effects of ionic imbalance on cellular trafficking dynamics in living cells are largely unexplored. By tracking distinct fluorescently labeled endosome/vesicle populations by live-cell imaging, we show that vesicle motility is reduced dramatically in a variety of cell types at the onset of hypertonic challenge. Live-cell imaging of actin and tubulin revealed similar arrested microfilament motility upon challenge. Vesicle motility recovered long after cell volume, a process that required functional regulatory volume increase and was accelerated by a return of extracellular osmolality to isosmotic levels. This delay suggests that, although volume-induced molecular crowding contributes to trafficking defects, it alone cannot explain the observed effects. Using fluorescent indicators and FRET-based probes, we found that intracellular ATP abundance and mitochondrial potential were reduced by hypertonicity and recovered after longer periods of time. Similar to the effects of osmotic challenge, isovolumetric elevation of intracellular chloride concentration by ionophores transiently decreased ATP production by mitochondria and abated microfilament and vesicle motility. These data illustrate how perturbed ionic balance, in addition to molecular crowding, affects membrane trafficking. PMID:26045497

O-GlcNAcylation affects β-catenin and E-cadherin expression, cell motility and tumorigenicity of colorectal cancer.

PubMed

Harosh-Davidovich, Shani Ben; Khalaila, Isam

2018-03-01

O-GlcNAcylation, the addition of β-N-acetylglucosamine (O-GlcNAc) moiety to Ser/Thr residues, is a sensor of the cell metabolic state. Cancer diseases such as colon, lung and breast cancer, possess deregulated O-GlcNAcylation. Studies during the last decade revealed that O-GlcNAcylation is implicated in cancer tumorigenesis and proliferation. The Wnt/β-catenin signaling pathway and cadherin-mediated adhesion are also implicated in epithelial-mesenchymal transition (EMT), a key cellular process in invasion and cancer metastasis. Often, deregulation of the Wnt pathway is caused by altered phosphorylation of its components. Specifically, phosphorylation of Ser or Thr residues of β-catenin affects its location and interaction with E-cadherin, thus facilitating cell-cell adhesion. Consistent with previous studies, the current study indicates that β-catenin is O-GlcNAcylated. To test the effect of O-GlcNAcylation on cell motility and how O-GlcNAcylation might affect β-catenin and E-cadherin functions, the enzyme machinery of O-GlcNAcylation was modulated either with chemical inhibitors or by gene silencing. When O-GlcNAcase (OGA) was inhibited, a global elevation of protein O-GlcNAcylation and increase in the expression of E-cadherin and β-catenin were noted. Concomitantly with enhanced O-GlcNAcylation, β-catenin transcriptional activity were elevated. Additionally, fibroblast cell motility was enhanced. Stable silenced cell lines with adenoviral OGA or adenoviral O-GlcNAc transferase (OGT) were established. Consistent with the results obtained by OGA chemical inhibition by TMG, OGT-silencing led to a significant reduction in β-catenin level. In vivo, murine orthotropic colorectal cancer model indicates that elevated O-GlcNAcylation leads to increased mortality rate, tumor and metastasis development. However, reduction in O-GlcNAcylation promoted survival that could be attributed to attenuated tumor and metastasis development. The results described herein provide circumstantial clues that O-GlcNAcylation deregulates β-catenin and E-cadherin expression and activity in fibroblast cell lines and this might influence EMT and cell motility, which may further influence tumor development and metastasis. Copyright © 2018 Elsevier Inc. All rights reserved.

Down-regulation of UDP-glucose dehydrogenase affects glycosaminoglycans synthesis and motility in HCT-8 colorectal carcinoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Tsung-Pao; Pan, Yun-Ru; Fu, Chien-Yu

2010-10-15

UDP-glucose dehydrogenase (UGDH) catalyzes oxidation of UDP-glucose to yield UDP-glucuronic acid, a precursor of hyaluronic acid (HA) and other glycosaminoglycans (GAGs) in extracellular matrix. Although association of extracellular matrix with cell proliferation and migration has been well documented, the importance of UGDH in these behaviors is not clear. Using UGDH-specific small interference RNA to treat HCT-8 colorectal carcinoma cells, a decrease in both mRNA and protein levels of UGDH, as well as the cellular UDP-glucuronic acid and GAG production was observed. Treatment of HCT-8 cells with either UGDH-specific siRNA or HA synthesis inhibitor 4-methylumbelliferone effectively delayed cell aggregation into multicellularmore » spheroids and impaired cell motility in both three-dimensional collagen gel and transwell migration assays. The reduction in cell aggregation and migration rates could be restored by addition of exogenous HA. These results indicate that UGDH can regulate cell motility through the production of GAG. The enzyme may be a potential target for therapeutic intervention of colorectal cancers.« less

Double-Stranded RNA-Dependent Protein Kinase Regulates the Motility of Breast Cancer Cells

PubMed Central

Xu, Mei; Chen, Gang; Wang, Siying; Liao, Mingjun; Frank, Jacqueline A.; Bower, Kimberly A.; Zhang, Zhuo; Shi, Xianglin; Luo, Jia

2012-01-01

Double-stranded RNA (dsRNA)-dependent protein kinase (PKR) is an interferon-induced protein kinase that plays a central role in the anti-viral process. Due to its pro-apoptotic and anti-proliferative action, there is an increased interest in PKR modulation as an anti-tumor strategy. PKR is overexpressed in breast cancer cells; however, the role of PKR in breast cancer cells is unclear. The expression/activity of PKR appears inversely related to the aggressiveness of breast cancer cells. The current study investigated the role of PKR in the motility/migration of breast cancer cells. The activation of PKR by a synthesized dsRNA (PIC) significantly decreased the motility of several breast cancer cell lines (BT474, MDA-MB231 and SKBR3). PIC inhibited cell migration and blocked cell membrane ruffling without affecting cell viability. PIC also induced the reorganization of the actin cytoskeleton and impaired the formation of lamellipodia. These effects of PIC were reversed by the pretreatment of a selective PKR inhibitor. PIC also activated p38 mitogen-activated protein kinase (MAPK) and its downstream MAPK-activated protein kinase 2 (MK2). PIC-induced activation of p38 MAPK and MK2 was attenuated by the PKR inhibitor and the PKR siRNA, but a selective p38 MAPK inhibitor (SB203580) or other MAPK inhibitors did not affect PKR activity, indicating that PKR is upstream of p38 MAPK/MK2. Cofilin is an actin severing protein and regulates membrane ruffling, lamellipodia formation and cell migration. PIC inhibited cofilin activity by enhancing its phosphorylation at Ser3. PIC activated LIM kinase 1 (LIMK1), an upstream kinase of cofilin in a p38 MAPK-dependent manner. We concluded that the activation of PKR suppressed cell motility by regulating the p38 MAPK/MK2/LIMK/cofilin pathway. PMID:23112838

Phosphorylation of Tyrosine Residues 31 and 118 on Paxillin Regulates Cell Migration through an Association with Crk in Nbt-II Cells

PubMed Central

Petit, Valérie; Boyer, Brigitte; Lentz, Delphine; Turner, Christopher E.; Thiery, Jean Paul; Vallés, Ana M.

2000-01-01

Identification of signaling molecules that regulate cell migration is important for understanding fundamental processes in development and the origin of various pathological conditions. The migration of Nara Bladder Tumor II (NBT-II) cells was used to determine which signaling molecules are specifically involved in the collagen-mediated locomotion. We show here that paxillin is tyrosine phosphorylated after induction of motility on collagen. Overexpression of paxillin mutants in which tyrosine 31 and/or tyrosine 118 were replaced by phenylalanine effectively impaired cell motility. Moreover, stimulation of motility by collagen preferentially enhanced the association of paxillin with the SH2 domain of the adaptor protein CrkII. Mutations in both tyrosine 31 and 118 diminished the phosphotyrosine content of paxillin and prevented the formation of the paxillin–Crk complex, suggesting that this association is necessary for collagen-mediated NBT-II cell migration. Other responses to collagen, such as cell adhesion and spreading, were not affected by these mutations. Overexpression of wild-type paxillin or Crk could bypass the migration-deficient phenotype. Both the SH2 and the SH3 domains of CrkII are shown to play a critical role in this collagen-mediated migration. These results demonstrate the important role of the paxillin–Crk complex in the collagen-induced cell motility. PMID:10704446

Sperm Motility in Flow

NASA Astrophysics Data System (ADS)

Guasto, Jeffrey; Juarez, Gabriel; Stocker, Roman

2012-11-01

A wide variety of plants and animals reproduce sexually by releasing motile sperm that seek out a conspecific egg, for example in the reproductive tract for mammals or in the water column for externally fertilizing organisms. Sperm are aided in their quest by chemical cues, but must also contend with hydrodynamic forces, resulting from laminar flows in reproductive tracts or turbulence in aquatic habitats. To understand how velocity gradients affect motility, we subjected swimming sperm to a range of highly-controlled straining flows using a cross-flow microfluidic device. The motion of the cell body and flagellum were captured through high-speed video microscopy. The effects of flow on swimming are twofold. For moderate velocity gradients, flow simply advects and reorients cells, quenching their ability to cross streamlines. For high velocity gradients, fluid stresses hinder the internal bending of the flagellum, directly inhibiting motility. The transition between the two regimes is governed by the Sperm number, which compares the external viscous stresses with the internal elastic stresses. Ultimately, unraveling the role of flow in sperm motility will lead to a better understanding of population dynamics among aquatic organisms and infertility problems in humans.

Increased count, motility, and total motile sperm cells collected across three consecutive ejaculations within 24 h of oocyte retrieval: implications for management of men presenting with low numbers of motile sperm for assisted reproduction.

PubMed

Said, Al-Hasen; Reed, Michael L

2015-07-01

The purpose of this study was to quantitate changes in seminal volume, sperm count, motility, qualitative forward progression, and total motile sperm cells per ejaculate, across three consecutive ejaculates collected from individuals within 24 h preceding an IVF cycle. Men presenting with oligoasthenozoospermia or asthenozoospemia attempted three ejaculates within 24 h preceding IVF. Ejaculate 1 was produced the afternoon prior to oocyte retrieval, and ejaculates 2 and 3 were produced the morning of oocyte retrieval with 2-3 h between collections. Ejaculates 1 and 2 were extended 1:1 v/v with room temperature rTYBS. Test tubes were placed into a beaker of room temperature water, then placed at 4 °C for gradual cooling. Ejaculate 3 was not extended, but pooled with ejaculates 1 and 2 and processed for intracytoplasmic sperm injection (ICSI). Out of 109 oocyte retrievals, 28 men were asked to attempt multiple consecutive ejaculations. Among this population, 25/28 (89.3 %) were successful, and 3/28 men (10.7 %) could only produce two ejaculates. Mean volumes for ejaculates 1, 2, and 3 were significantly different from each other (p < 0.01); the volume decreased for each ejaculate. Mean sperm counts, motility, qualitative forward progression, and total motile cells per ejaculate for the ejaculates1, 2, and 3 demonstrated the following: ejaculates 2 and 3 were not significantly different, but counts, motility, and total motile sperm were improved over ejaculate 1 (p < 0.01). Pooling three consecutive ejaculates within 24 h increased the numbers of available motile sperm in this population by 8-fold compared to the first ejaculate alone, facilitating avoidance of sperm cryopreservation and additional centrifugation steps that could affect sperm viability and/or function.

TEMPORARY STORAGE OF BOVINE SEMEN CRYOPRESERVED IN LIQUID NITROGEN ON DRY ICE AND REFREEZING OF FROZEN-THAWED SEMEN.

PubMed

Abdussamad, A M; Gauly, M; Holtz, W

2015-01-01

Two experiments were conducted. The purpose of Experiment 1 was to investigate whether viability of bovine semen stored in liquid nitrogen (-196°C) will be adversely affected by temporary exposure to dry ice (-79°C). It was convincingly shown that post thaw-motility was not affected, regardless whether semen was thawed immediately or after being returned to liquid nitrogen. Shipping or temporary storage on dry ice, thus, is a viable option. In Experiment 2, refreezing of frozen-thawed semen was attempted. The proportion of motile spermatozoa was reduced by a factor of ten to between 6.0 % and 7.4 %, regardless whether thawing occurred directly after removal from liquid nitrogen or after an interim period on dry ice. When semen was refrozen on dry ice before being returned to liquid nitrogen, motility rates were significantly improved (13.0 % to 17.0 %, P<0.05). In both experiments sperm cells that remained motile displayed vigorous forward movement and normal morphological appearance.

In vitro and in vivo study of transcriptome alternation induced by butyrate in cattle using deep RNA-seq

USDA-ARS?s Scientific Manuscript database

Short-chain fatty acids (SCFAs,), especially butyrate, affect cell differentiation, proliferation, and motility. Furthermore, butyrate induces cell cycle arrest and apoptosis through its inhibition on histone deacetylases (HDACs). Butyrate is a potent inducer of histone hyper-acetylation in cells a...

Motile cilia of human airway epithelia contain hedgehog signaling components that mediate noncanonical hedgehog signaling.

PubMed

Mao, Suifang; Shah, Alok S; Moninger, Thomas O; Ostedgaard, Lynda S; Lu, Lin; Tang, Xiao Xiao; Thornell, Ian M; Reznikov, Leah R; Ernst, Sarah E; Karp, Philip H; Tan, Ping; Keshavjee, Shaf; Abou Alaiwa, Mahmoud H; Welsh, Michael J

2018-02-06

Differentiated airway epithelia produce sonic hedgehog (SHH), which is found in the thin layer of liquid covering the airway surface. Although previous studies showed that vertebrate HH signaling requires primary cilia, as airway epithelia mature, the cells lose primary cilia and produce hundreds of motile cilia. Thus, whether airway epithelia have apical receptors for SHH has remained unknown. We discovered that motile cilia on airway epithelial cells have HH signaling proteins, including patched and smoothened. These cilia also have proteins affecting cAMP-dependent signaling, including Gα i and adenylyl cyclase 5/6. Apical SHH decreases intracellular levels of cAMP, which reduces ciliary beat frequency and pH in airway surface liquid. These results suggest that apical SHH may mediate noncanonical HH signaling through motile cilia to dampen respiratory defenses at the contact point between the environment and the lung, perhaps counterbalancing processes that stimulate airway defenses. Copyright © 2018 the Author(s). Published by PNAS.

Cellular Scale Anisotropic Topography Guides Schwann Cell Motility

PubMed Central

Mitchel, Jennifer A.; Hoffman-Kim, Diane

2011-01-01

Directed migration of Schwann cells (SC) is critical for development and repair of the peripheral nervous system. Understanding aspects of motility specific to SC, along with SC response to engineered biomaterials, may inform strategies to enhance nerve regeneration. Rat SC were cultured on laminin-coated microgrooved poly(dimethyl siloxane) platforms that were flat or presented repeating cellular scale anisotropic topographical cues, 30 or 60 µm in width, and observed with timelapse microscopy. SC motion was directed parallel to the long axis of the topography on both the groove floor and the plateau, with accompanying differences in velocity and directional persistence in comparison to SC motion on flat substrates. In addition, feature dimension affected SC morphology, alignment, and directional persistence. Plateaus and groove floors presented distinct cues which promoted differential motility and variable interaction with the topographical features. SC on the plateau surfaces tended to have persistent interactions with the edge topography, while SC on the groove floors tended to have infrequent contact with the corners and walls. Our observations suggest the capacity of SC to be guided without continuous contact with a topographical cue. SC exhibited a range of distinct motile morphologies, characterized by their symmetry and number of extensions. Across all conditions, SC with a single extension traveled significantly faster than cells with more or no extensions. We conclude that SC motility is complex, where persistent motion requires cellular asymmetry, and that anisotropic topography with cellular scale features can direct SC motility. PMID:21949703

Hypoxic stellate cells of pancreatic cancer stroma regulate extracellular matrix fiber organization and cancer cell motility.

PubMed

Sada, Masafumi; Ohuchida, Kenoki; Horioka, Kohei; Okumura, Takashi; Moriyama, Taiki; Miyasaka, Yoshihiro; Ohtsuka, Takao; Mizumoto, Kazuhiro; Oda, Yoshinao; Nakamura, Masafumi

2016-03-28

Desmoplasia and hypoxia in pancreatic cancer mutually affect each other and create a tumor-supportive microenvironment. Here, we show that microenvironment remodeling by hypoxic pancreatic stellate cells (PSCs) promotes cancer cell motility through alteration of extracellular matrix (ECM) fiber architecture. Three-dimensional (3-D) matrices derived from PSCs under hypoxia exhibited highly organized parallel-patterned matrix fibers compared with 3-D matrices derived from PSCs under normoxia, and promoted cancer cell motility by inducing directional migration of cancer cells due to the parallel fiber architecture. Microarray analysis revealed that procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) in PSCs was the gene that potentially regulates ECM fiber architecture under hypoxia. Stromal PLOD2 expression in surgical specimens of pancreatic cancer was confirmed by immunohistochemistry. RNA interference-mediated knockdown of PLOD2 in PSCs blocked parallel fiber architecture of 3-D matrices, leading to decreased directional migration of cancer cells within the matrices. In conclusion, these findings indicate that hypoxia-induced PLOD2 expression in PSCs creates a permissive microenvironment for migration of cancer cells through architectural regulation of stromal ECM in pancreatic cancer. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

X-linked Inhibitor of Apoptosis Protein (XIAP) Mediates Cancer Cell Motility via Rho GDP Dissociation Inhibitor (RhoGDI)-dependent Regulation of the Cytoskeleton*

PubMed Central

Liu, Jinyi; Zhang, Dongyun; Luo, Wenjing; Yu, Yonghui; Yu, Jianxiu; Li, Jingxia; Zhang, Xinhai; Zhang, Baolin; Chen, Jingyuan; Wu, Xue-Ru; Rosas-Acosta, Germán; Huang, Chuanshu

2011-01-01

X-linked inhibitor of apoptosis protein (XIAP) overexpression has been found to be associated with malignant cancer progression and aggression in individuals with many types of cancers. However, the molecular basis of XIAP in the regulation of cancer cell biological behavior remains largely unknown. In this study, we found that a deficiency of XIAP expression in human cancer cells by either knock-out or knockdown leads to a marked reduction in β-actin polymerization and cytoskeleton formation. Consistently, cell migration and invasion were also decreased in XIAP-deficient cells compared with parental wild-type cells. Subsequent studies demonstrated that the regulation of cell motility by XIAP depends on its interaction with the Rho GDP dissociation inhibitor (RhoGDI) via the XIAP RING domain. Furthermore, XIAP was found to negatively regulate RhoGDI SUMOylation, which might affect its activity in controlling cell motility. Collectively, our studies provide novel insights into the molecular mechanisms by which XIAP regulates cancer invasion and offer a further theoretical basis for setting XIAP as a potential prognostic marker and specific target for treatment of cancers with metastatic properties. PMID:21402697

Disruption of TgPHIL1 Alters Specific Parameters of Toxoplasma gondii Motility Measured in a Quantitative, Three-Dimensional Live Motility Assay

PubMed Central

Leung, Jacqueline M.; Rould, Mark A.; Konradt, Christoph; Hunter, Christopher A.; Ward, Gary E.

2014-01-01

T. gondii uses substrate-dependent gliding motility to invade cells of its hosts, egress from these cells at the end of its lytic cycle and disseminate through the host organism during infection. The ability of the parasite to move is therefore critical for its virulence. T. gondii engages in three distinct types of gliding motility on coated two-dimensional surfaces: twirling, circular gliding and helical gliding. We show here that motility in a three-dimensional Matrigel-based environment is strikingly different, in that all parasites move in irregular corkscrew-like trajectories. Methods developed for quantitative analysis of motility parameters along the smoothed trajectories demonstrate a complex but periodic pattern of motility with mean and maximum velocities of 0.58±0.07 µm/s and 2.01±0.17 µm/s, respectively. To test how a change in the parasite's crescent shape might affect trajectory parameters, we compared the motility of Δphil1 parasites, which are shorter and wider than wild type, to the corresponding parental and complemented lines. Although comparable percentages of parasites were moving for all three lines, the Δphil1 mutant exhibited significantly decreased trajectory lengths and mean and maximum velocities compared to the parental parasite line. These effects were either partially or fully restored upon complementation of the Δphil1 mutant. These results show that alterations in morphology may have a significant impact on T. gondii motility in an extracellular matrix-like environment, provide a possible explanation for the decreased fitness of Δphil1 parasites in vivo, and demonstrate the utility of the quantitative three-dimensional assay for studying parasite motility. PMID:24489670

Inference of cell-cell interactions from population density characteristics and cell trajectories on static and growing domains.

PubMed

Ross, Robert J H; Yates, C A; Baker, R E

2015-06-01

A key feature of cell migration is how cell movement is affected by cell-cell interactions. Furthermore, many cell migratory processes such as neural crest stem cell migration [Thomas and Erickson, 2008; McLennan et al., 2012] occur on growing domains or in the presence of a chemoattractant. Therefore, it is important to study interactions between migrating cells in the context of domain growth and directed motility. Here we compare discrete and continuum models describing the spatial and temporal evolution of a cell population for different types of cell-cell interactions on static and growing domains. We suggest that cell-cell interactions can be inferred from population density characteristics in the presence of motility bias, and these population density characteristics for different cell-cell interactions are conserved on both static and growing domains. We also study the expected displacement of a tagged cell, and show that different types of cell-cell interactions can give rise to cell trajectories with different characteristics. These characteristics are conserved in the presence of domain growth, however, they are diminished in the presence of motility bias. Our results are relevant for researchers who study the existence and role of cell-cell interactions in biological systems, so far as we suggest that different types of cell-cell interactions could be identified from cell density and trajectory data. Copyright © 2015 Elsevier Inc. All rights reserved.

Mouse and human HSPC immobilization in liquid culture by CD43- or CD44-antibody coating.

PubMed

Loeffler, Dirk; Wang, Weijia; Hopf, Alois; Hilsenbeck, Oliver; Bourgine, Paul E; Rudolf, Fabian; Martin, Ivan; Schroeder, Timm

2018-03-29

Keeping track of individual cell identifications is imperative to the study of dynamic single-cell behavior over time. Highly motile hematopoietic stem and progenitor cells (HSPCs) migrate quickly and do not adhere, and thus must be imaged very frequently to keep cell identifications. Even worse, they are also flushed away during medium exchange. To overcome these limitations, we tested antibody coating for reducing HSPC motility in vitro. Anti-CD43- and anti-CD44-antibody coating reduced the cell motility of mouse and human HSPCs in a concentration-dependent manner. This enables 2-dimensional (2D) colony formation without cell mixing in liquid cultures, massively increases time-lapse imaging throughput, and also maintains cell positions during media exchange. Anti-CD43 but not anti-CD44 coating reduces mouse HSPC proliferation with increasing concentrations. No relevant effects on cell survival or myeloid and megakaryocyte differentiation of hematopoietic stem cells and multipotent progenitors 1-5 were detected. Human umbilical cord hematopoietic CD34 + cell survival, proliferation, and differentiation were not affected by either coating. This approach both massively simplifies and accelerates continuous analysis of suspension cells, and enables the study of their behavior in dynamic rather than static culture conditions over time. © 2018 by The American Society of Hematology.

Hydrogen peroxide stimulates cell motile activity through LPA receptor-3 in liver epithelial WB-F344 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shibata, Ayano; Tanabe, Eriko; Inoue, Serina

2013-04-12

Highlights: •Hydrogen peroxide stimulates cell motility of WB-F344 cells. •LPA{sub 3} is induced by hydrogen peroxide in WB-F344 cells. •Cell motility by hydrogen peroxide is inhibited in LPA{sub 3} knockdown cells. •LPA signaling is involved in cell migration by hydrogen peroxide. -- Abstract: Hydrogen peroxide which is one of reactive oxygen species (ROS) mediates a variety of biological responses, including cell proliferation and migration. In the present study, we investigated whether lysophosphatidic acid (LPA) signaling is involved in cell motile activity stimulated by hydrogen peroxide. The rat liver epithelial WB-F344 cells were treated with hydrogen peroxide at 0.1 or 1more » μM for 48 h. In cell motility assays, hydrogen peroxide treated cells showed significantly high cell motile activity, compared with untreated cells. To measure the expression levels of LPA receptor genes, quantitative real time RT-PCR analysis was performed. The expressions of LPA receptor-3 (Lpar3) in hydrogen peroxide treated cells were significantly higher than those in control cells, but not Lpar1 and Lpar2 genes. Next, to assess the effect of LPA{sub 3} on cell motile activity, the Lpar3 knockdown cells from WB-F344 cells were also treated with hydrogen peroxide. The cell motile activity of the knockdown cells was not stimulated by hydrogen peroxide. Moreover, in liver cancer cells, hydrogen peroxide significantly activated cell motility of Lpar3-expressing cells, but not Lpar3-unexpressing cells. These results suggest that LPA signaling via LPA{sub 3} may be mainly involved in cell motile activity of WB-F344 cells stimulated by hydrogen peroxide.« less

Myxococcus xanthus Gliding Motors Are Elastically Coupled to the Substrate as Predicted by the Focal Adhesion Model of Gliding Motility

PubMed Central

Balagam, Rajesh; Litwin, Douglas B.; Czerwinski, Fabian; Sun, Mingzhai; Kaplan, Heidi B.; Shaevitz, Joshua W.; Igoshin, Oleg A.

2014-01-01

Myxococcus xanthus is a model organism for studying bacterial social behaviors due to its ability to form complex multi-cellular structures. Knowledge of M. xanthus surface gliding motility and the mechanisms that coordinated it are critically important to our understanding of collective cell behaviors. Although the mechanism of gliding motility is still under investigation, recent experiments suggest that there are two possible mechanisms underlying force production for cell motility: the focal adhesion mechanism and the helical rotor mechanism, which differ in the biophysics of the cell–substrate interactions. Whereas the focal adhesion model predicts an elastic coupling, the helical rotor model predicts a viscous coupling. Using a combination of computational modeling, imaging, and force microscopy, we find evidence for elastic coupling in support of the focal adhesion model. Using a biophysical model of the M. xanthus cell, we investigated how the mechanical interactions between cells are affected by interactions with the substrate. Comparison of modeling results with experimental data for cell-cell collision events pointed to a strong, elastic attachment between the cell and substrate. These results are robust to variations in the mechanical and geometrical parameters of the model. We then directly measured the motor-substrate coupling by monitoring the motion of optically trapped beads and find that motor velocity decreases exponentially with opposing load. At high loads, motor velocity approaches zero velocity asymptotically and motors remain bound to beads indicating a strong, elastic attachment. PMID:24810164

The effect of surface characteristics on the transport of multiple Escherichia coli isolates in large scale columns of quartz sand.

PubMed

Lutterodt, G; Basnet, M; Foppen, J W A; Uhlenbrook, S

2009-02-01

Bacteria properties play an important role in the transport of bacteria in groundwater, but their role, especially for longer transport distances (>0.5 m) has not been studied. Thereto, we studied the effects of cell surface hydrophobicity, outer surface potential (OSP), cell sphericity, motility, and Ag43 protein expression on the outer cell surface for a number of E. coli strains, obtained from the environment on their transport behavior in columns of saturated quartz sand of 5 m height in two solutions: demineralized (DI) water and artificial groundwater (AGW). In DI water, sticking efficiencies ranged between 0.1 and 0.4 at the column inlet, and then decreased with transport distance to 0.02-0.2. In AGW, sticking efficiencies were on average 1log-unit higher than those in DI (water). Bacteria motility and Ag43 expression affected attachment with a (high) statistical significance. In contrast, hydrophobicity, OSP and cell sphericity did not significantly correlate with sticking efficiency. However, for transport distances more than 0.33 m, the correlation between sticking efficiency, Ag43 expression, and motility became insignificant. We concluded that Ag43 and motility played an important role in E. coli attachment to quartz grain surfaces, and that the transport distance dependent sticking efficiency reductions were caused by motility and Ag43 expression variations within a population. The implication of our findings is that less motile bacteria with little or no Ag43 expression may travel longer distances once they enter groundwater environments. In future studies, the possible effect of bacteria surface structures, like fimbriae, pili and surface proteins on bacteria attachment need to be considered more systematically in order to arrive at more meaningful inter-population comparisons of the transport behavior of E. coli strains in aquifers.

Acetylcholinesterase-R increases germ cell apoptosis but enhances sperm motility

PubMed Central

Mor, I; Sklan, EH; Podoly, E; Pick, M; Kirschner, M; Yogev, L; Bar-Sheshet Itach, S; Schreiber, L; Geyer, B; Mor, T; Grisaru, D; Soreq, H

2008-01-01

Abstract Changes in protein subdomains through alternative splicing often modify protein-protein interactions, altering biological processes. A relevant example is that of the stress-induced up-regulation of the acetylcholinesterase (AChE-R) splice variant, a common response in various tissues. In germ cells of male transgenic TgR mice, AChE-R excess associates with reduced sperm differentiation and sperm counts. To explore the mechanism(s) by which AChE-R up-regulation affects spermatogenesis, we identified AChE-R's protein partners through a yeast two-hybrid screen. In meiotic spermatocytes from TgR mice, we detected AChE-R interaction with the scaffold protein RACK1 and elevated apoptosis. This correlated with reduced scavenging by RACK1 of the pro-apoptotic TAp73, an outcome compatible with the increased apoptosis. In contrast, at later stages in sperm development, AChE-R's interaction with the glycolytic enzyme enolase-α elevates enolase activity. In transfected cells, enforced AChE-R excess increased glucose uptake and adenosine tri-phosphate (ATP) levels. Correspondingly, TgR sperm cells display elevated ATP levels, mitochondrial hyperactivity and increased motility. In human donors' sperm, we found direct association of sperm motility with AChE-R expression. Interchanging interactions with RACK1 and enolase-α may hence enable AChE-R to affect both sperm differentiation and function by participating in independent cellular pathways. PMID:18194455

Hydrogen peroxide stimulates cell motile activity through LPA receptor-3 in liver epithelial WB-F344 cells.

PubMed

Shibata, Ayano; Tanabe, Eriko; Inoue, Serina; Kitayoshi, Misaho; Okimoto, Souta; Hirane, Miku; Araki, Mutsumi; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi

2013-04-12

Hydrogen peroxide which is one of reactive oxygen species (ROS) mediates a variety of biological responses, including cell proliferation and migration. In the present study, we investigated whether lysophosphatidic acid (LPA) signaling is involved in cell motile activity stimulated by hydrogen peroxide. The rat liver epithelial WB-F344 cells were treated with hydrogen peroxide at 0.1 or 1 μM for 48 h. In cell motility assays, hydrogen peroxide treated cells showed significantly high cell motile activity, compared with untreated cells. To measure the expression levels of LPA receptor genes, quantitative real time RT-PCR analysis was performed. The expressions of LPA receptor-3 (Lpar3) in hydrogen peroxide treated cells were significantly higher than those in control cells, but not Lpar1 and Lpar2 genes. Next, to assess the effect of LPA3 on cell motile activity, the Lpar3 knockdown cells from WB-F344 cells were also treated with hydrogen peroxide. The cell motile activity of the knockdown cells was not stimulated by hydrogen peroxide. Moreover, in liver cancer cells, hydrogen peroxide significantly activated cell motility of Lpar3-expressing cells, but not Lpar3-unexpressing cells. These results suggest that LPA signaling via LPA3 may be mainly involved in cell motile activity of WB-F344 cells stimulated by hydrogen peroxide. Copyright © 2013 Elsevier Inc. All rights reserved.

Dose Dependent Side Effect of Superparamagnetic Iron Oxide Nanoparticle Labeling on Cell Motility in Two Fetal Stem Cell Populations

PubMed Central

Diana, Valentina; Bossolasco, Patrizia; Moscatelli, Davide; Silani, Vincenzo; Cova, Lidia

2013-01-01

Multipotent stem cells (SCs) could substitute damaged cells and also rescue degeneration through the secretion of trophic factors able to activate the endogenous SC compartment. Therefore, fetal SCs, characterized by high proliferation rate and devoid of ethical concern, appear promising candidate, particularly for the treatment of neurodegenerative diseases. Super Paramagnetic Iron Oxide nanoparticles (SPIOn), routinely used for pre-clinical cell imaging and already approved for clinical practice, allow tracking of transplanted SCs and characterization of their fate within the host tissue, when combined with Magnetic Resonance Imaging (MRI). In this work we investigated how SPIOn could influence cell migration after internalization in two fetal SC populations: human amniotic fluid and chorial villi SCs were labeled with SPIOn and their motility was evaluated. We found that SPIOn loading significantly reduced SC movements without increasing production of Reactive Oxygen Species (ROS). Moreover, motility impairment was directly proportional to the amount of loaded SPIOn while a chemoattractant-induced recovery was obtained by increasing serum levels. Interestingly, the migration rate of SPIOn labeled cells was also significantly influenced by a degenerative surrounding. In conclusion, this work highlights how SPIOn labeling affects SC motility in vitro in a dose-dependent manner, shedding the light on an important parameter for the creation of clinical protocols. Establishment of an optimal SPIOn dose that enables both a good visualization of grafted cells by MRI and the physiological migration rate is a main step in order to maximize the effects of SC therapy in both animal models of neurodegeneration and clinical studies. PMID:24244310

Keratinocyte Motility Is Affected by UVA Radiation-A Comparison between Normal and Dysplastic Cells.

PubMed

Niculiţe, Cristina M; Nechifor, Marina T; Urs, Andreea O; Olariu, Laura; Ceafalan, Laura C; Leabu, Mircea

2018-06-07

UVA radiation induces multiple and complex changes in the skin, affecting epidermal cell behavior. This study reports the effects of UVA exposure on normal (HaCaT) and dysplastic (DOK) keratinocytes. The adherence, spreading and proliferation were investigated by time-lapse measurement of cell layer impedance on different matrix proteins. Prior to UVA exposure, the time required for adherence and spreading did not differ significantly for HaCaT and DOK cells, while spreading areas were larger for HaCaT cells. Under UVA exposure, HaCaT and DOK cells behavior differed in terms of movement and proliferation. The cells' ability to cover the denuded surface and individual cell trajectories were recorded by time-lapse videomicroscopy, during wound healing experiments. Dysplastic keratinocytes showed more sensitivity to UVA, exhibiting transient deficiencies in directionality of movement and a delay in re-coating the denuded area. The actin cytoskeleton displayed a cortical organization immediately after irradiation, in both cell lines, similar to mock-irradiated cells. Post-irradiation, DOK cells displayed a better organization of stress fibers, persistent filopodia, and new, stronger focal contacts. In conclusion, after UVA exposure HaCaT and DOK cells showed a different behavior in terms of adherence, spreading, motility, proliferation, and actin cytoskeleton dynamics, with the dyplastic keratinocytes being more sensitive.

Inhibitory effects of LPA1 on cell motile activities stimulated by hydrogen peroxide and 2,3-dimethoxy-1,4-naphthoquinone in fibroblast 3T3 cells.

PubMed

Hirane, Miku; Araki, Mutsumi; Dong, Yan; Honoki, Kanya; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi

2013-11-08

Reactive oxygen species (ROS) are known to mediate a variety of biological responses, including cell motility. Recently, we indicated that lysophosphatidic acid (LPA) receptor-3 (LPA3) increased cell motile activity stimulated by hydrogen peroxide. In the present study, we assessed the role of LPA1 in the cell motile activity mediated by ROS in mouse fibroblast 3T3 cells. 3T3 cells were treated with hydrogen peroxide and 2,3-dimethoxy-1,4-naphthoquinone (DMNQ) at concentrations of 0.1 and 1 μM for 48 h. In cell motility assays with Cell Culture Inserts, the cell motile activities of 3T3 cells treated with hydrogen peroxide and DMNQ were significantly higher than those of untreated cells. 3T3 cells treated with hydrogen peroxide and DMNQ showed elevated expression levels of the Lpar3 gene, but not the Lpar1 and Lpar2 genes. To investigate the effects of LPA1 on the cell motile activity induced by hydrogen peroxide and DMNQ, Lpar1-overexpressing (3T3-a1) cells were generated from 3T3 cells and treated with hydrogen peroxide and DMNQ. The cell motile activities stimulated by hydrogen peroxide and DMNQ were markedly suppressed in 3T3-a1 cells. These results suggest that LPA signaling via LPA1 inhibits the cell motile activities stimulated by hydrogen peroxide and DMNQ in 3T3 cells. Copyright © 2013 Elsevier Inc. All rights reserved.

Transcriptomic analysis displays the effect of (-)-roemerine on the motility and nutrient uptake in Escherichia coli.

PubMed

Ayyildiz, Dilara; Arga, Kazim Yalcin; Avci, Fatma Gizem; Altinisik, Fatma Ece; Gurer, Caglayan; Gulsoy Toplan, Gizem; Kazan, Dilek; Wozny, Katharina; Brügger, Britta; Mertoglu, Bulent; Sariyar Akbulut, Berna

2017-08-01

Among the different families of plant alkaloids, (-)-roemerine, an aporphine type, was recently shown to possess significant antibacterial activity in Escherichia coli. Based on the increasing demand for antibacterials with novel mechanisms of action, the present work investigates the potential of the plant-derived alkaloid (-)-roemerine as an antibacterial in E. coli cells using microarray technology. Analysis of the genome-wide transcriptional reprogramming in cells after 60 min treatment with 100 μg/mL (-)-roemerine showed significant changes in the expression of 241 genes (p value <0.05 and fold change >2). Expression of selected genes was confirmed by qPCR. Differentially expressed genes were classified into functional categories to map biological processes and molecular pathways involved. Cellular activities with roles in carbohydrate transport and metabolism, energy production and conversion, lipid transport and metabolism, amino acid transport and metabolism, two-component signaling systems, and cell motility (in particular, the flagellar organization and motility) were among metabolic processes altered in the presence of (-)-roemerine. The down-regulation of the outer membrane proteins probably led to a decrease in carbohydrate uptake rate, which in turn results in nutrient limitation. Consequently, energy metabolism is slowed down. Interestingly, the majority of the expressional alterations were found in the flagellar system. This suggested reduction in motility and loss in the ability to form biofilms, thus affecting protection of E. coli against host cell defense mechanisms. In summary, our findings suggest that the antimicrobial action of (-)-roemerine in E. coli is linked to disturbances in motility and nutrient uptake.

Sperm quality analysis in XX, XY and YY males of the Nile tilapia (Oreochromis niloticus).

PubMed

Gennotte, V; François, E; Rougeot, C; Ponthier, J; Deleuze, S; Mélard, C

2012-07-01

In Nile tilapia (Oreochromis niloticus), individuals with atypical sexual genotype are commonly used in farming (use of YY males to produce all-male offspring), but they also constitute major tools to study sex determinism mechanisms. In other species, sexual genotype and sex reversal procedures affect different aspects of biology, such as growth, behavior and reproductive success. The aim of this study was to assess the influence of sexual genotype on sperm quality in Nile tilapia. Milt characteristics were compared in XX (sex-reversed), XY and YY males in terms of gonadosomatic index, sperm count, sperm motility and duration of sperm motility. Sperm motility was measured by computer-assisted sperm analysis (CASA) quantifying several parameters: total motility, progressive motility, curvilinear velocity, straight line velocity, average path velocity and linearity. None of the sperm traits measured significantly differed between the three genotypes. Mean values of gonadosomatic index, sperm concentration and sperm motility duration of XX, XY and YY males, respectively ranged from 0.92 to 1.33%, from 1.69 to 2.22 ×10(9) cells mL(-1) and from 18'04″ to 27'32″. Mean values of total motility and curvilinear velocity 1 min after sperm activation, respectively ranged from 53 to 58% and from 71 to 76 μm s(-1) for the three genotypes. After 3 min of activity, all the sperm motility and velocity parameters dropped by half and continued to slowly decrease thereafter. Seven min after activation, only 9 to 13% of spermatozoa were still progressive. Our results prove that neither sexual genotype nor hormonal sex reversal treatments affect sperm quality in male Nile tilapias with atypical sexual genotype. Copyright © 2012 Elsevier Inc. All rights reserved.

Different induction of LPA receptors by chemical liver carcinogens regulates cellular functions of liver epithelial WB-F344 cells.

PubMed

Hirane, Miku; Ishii, Shuhei; Tomimatsu, Ayaka; Fukushima, Kaori; Takahashi, Kaede; Fukushima, Nobuyuki; Honoki, Kanya; Tsujiuchi, Toshifumi

2016-11-01

Lysophosphatidic acid (LPA) signaling via LPA receptors (LPA 1 to LPA 6 ) mediates a variety of cellular functions, including cell motility. In the present study, we investigated the effects of LPA receptors on cell motile activity during multi-stage hepatocarcinogenesis in rat liver epithelial WB-F344 cells treated with chemical liver carcinogens. Cells were treated with a initiator (N-nitrosodiethylamine (DEN)) and three promoters (phenobarbital (PB), okadaic acid (OA) and clofibrate) every 24 h for 2 days. Cell motile activity was elevated by DEN, correlating with Lpar3 expression. PB, OA, and clofibrate elevated Lpar1 expression and inhibited cell motile activity. To evaluate the effects of long-term treatment on cell motility, cells were treated with DEN and/or PB for at least 6 months. Lpar3 expression and cell motile activity were significantly elevated by the long-term DEN treatment with or without further PB treatment. In contrast, long-term PB treatment with or without further DEN elevated Lpar1 expression and inhibited cell motility. When the synthesis of extracellular LPA was blocked by a potent ATX inhibitor S32826 before cell motility assay, the cell motility induced by DEN and PB was markedly suppressed. These results suggest that activation of the different LPA receptors may regulate the biological functions of cells treated with chemical carcinogens. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

Two distinct mTORC2-dependent pathways converge on Rac1 to drive breast cancer metastasis.

PubMed

Morrison Joly, Meghan; Williams, Michelle M; Hicks, Donna J; Jones, Bayley; Sanchez, Violeta; Young, Christian D; Sarbassov, Dos D; Muller, William J; Brantley-Sieders, Dana; Cook, Rebecca S

2017-06-30

The importance of the mTOR complex 2 (mTORC2) signaling complex in tumor progression is becoming increasingly recognized. HER2-amplified breast cancers use Rictor/mTORC2 signaling to drive tumor formation, tumor cell survival and resistance to human epidermal growth factor receptor 2 (HER2)-targeted therapy. Cell motility, a key step in the metastatic process, can be activated by mTORC2 in luminal and triple negative breast cancer cell lines, but its role in promoting metastases from HER2-amplified breast cancers is not yet clear. Because Rictor is an obligate cofactor of mTORC2, we genetically engineered Rictor ablation or overexpression in mouse and human HER2-amplified breast cancer models for modulation of mTORC2 activity. Signaling through mTORC2-dependent pathways was also manipulated using pharmacological inhibitors of mTOR, Akt, and Rac. Signaling was assessed by western analysis and biochemical pull-down assays specific for Rac-GTP and for active Rac guanine nucleotide exchange factors (GEFs). Metastases were assessed from spontaneous tumors and from intravenously delivered tumor cells. Motility and invasion of cells was assessed using Matrigel-coated transwell assays. We found that Rictor ablation potently impaired, while Rictor overexpression increased, metastasis in spontaneous and intravenously seeded models of HER2-overexpressing breast cancers. Additionally, migration and invasion of HER2-amplified human breast cancer cells was diminished in the absence of Rictor, or upon pharmacological mTOR kinase inhibition. Active Rac1 was required for Rictor-dependent invasion and motility, which rescued invasion/motility in Rictor depleted cells. Rictor/mTORC2-dependent dampening of the endogenous Rac1 inhibitor RhoGDI2, a factor that correlated directly with increased overall survival in HER2-amplified breast cancer patients, promoted Rac1 activity and tumor cell invasion/migration. The mTORC2 substrate Akt did not affect RhoGDI2 dampening, but partially increased Rac1 activity through the Rac-GEF Tiam1, thus partially rescuing cell invasion/motility. The mTORC2 effector protein kinase C (PKC)α did rescue Rictor-mediated RhoGDI2 downregulation, partially rescuing Rac-guanosine triphosphate (GTP) and migration/motility. These findings suggest that mTORC2 uses two coordinated pathways to activate cell invasion/motility, both of which converge on Rac1. Akt signaling activates Rac1 through the Rac-GEF Tiam1, while PKC signaling dampens expression of the endogenous Rac1 inhibitor, RhoGDI2.

Second-harmonic generation scattering directionality predicts tumor cell motility in collagen gels

NASA Astrophysics Data System (ADS)

Burke, Kathleen A.; Dawes, Ryan P.; Cheema, Mehar K.; Van Hove, Amy; Benoit, Danielle S. W.; Perry, Seth W.; Brown, Edward

2015-05-01

Second-harmonic generation (SHG) allows for the analysis of tumor collagen structural changes throughout metastatic progression. SHG directionality, measured through the ratio of the forward-propagating to backward-propagating signal (F/B ratio), is affected by collagen fibril diameter, spacing, and disorder of fibril packing within a fiber. As tumors progress, these parameters evolve, producing concurrent changes in F/B. It has been recently shown that the F/B of highly metastatic invasive ductal carcinoma (IDC) breast tumors is significantly different from less metastatic tumors. This suggests a possible relationship between the microstructure of collagen, as measured by the F/B, and the ability of tumor cells to locomote through that collagen. Utilizing in vitro collagen gels of different F/B ratios, we explored the relationship between collagen microstructure and motility of tumor cells in a "clean" environment, free of the myriad cells, and signals found in in vivo. We found a significant relationship between F/B and the total distance traveled by the tumor cell, as well as both the average and maximum velocities of the cells. Consequently, one possible mechanism underlying the observed relationship between tumor F/B and metastatic output in IDC patient samples is a direct influence of collagen structure on tumor cell motility.

Inhibition of Swarming motility of Pseudomonas aeruginosa by Methanol extracts of Alpinia officinarum Hance. and Cinnamomum tamala T. Nees and Eberm.

PubMed

Lakshmanan, Divya; Nanda, Jishudas; Jeevaratnam, K

2018-06-01

Bacterial drug resistance is a challenge in clinical settings, especially in countries like India. Hence, discovery of novel alternative therapeutics has become a necessity in the fight against drug resistance. Compounds that inhibit bacterial virulence properties form new therapeutic alternatives. Pseudomonas aeruginosa is an opportunistic, nosocomial pathogen that infects immune-compromised patients. Swarming motility is an important virulence property of Pseudomonas which aids it in reaching host cells under nutrient limiting conditions. Here, we report the screening of five plant extracts against swarming motility of P. aeruginosa and show that methanol extracts of Alpinia officinarum and Cinnamomum tamala inhibit swarming motility at 5 μg mL -1 without inhibiting its growth. These extracts did not inhibit swimming and twitching motilities indicating a mode of action specific to swarming pathway. Preliminary experiments indicated that rhamnolipid production was not affected. This study reveals the potential of the two plants in anti-virulence drug discovery.

The GIT–PIX complexes regulate the chemotactic response of rat basophilic leukaemia cells

PubMed Central

Gavina, Manuela; Za, Lorena; Molteni, Raffaella; Pardi, Ruggero; Curtis, Ivan de

2009-01-01

Background information. Cell motility entails the reorganization of the cytoskeleton and membrane trafficking for effective protrusion. The GIT–PIX protein complexes are involved in the regulation of cell motility and adhesion and in the endocytic traffic of members of the family of G-protein-coupled receptors. We have investigated the function of the endogenous GIT complexes in the regulation of cell motility stimulated by fMLP (formyl-Met-Leu-Phe) peptide, in a rat basophilic leukaemia RBL-2H3 cell line stably expressing an HA (haemagglutinin)-tagged receptor for the fMLP peptide. Results. Our analysis shows that RBL cells stably transfected with the chemoattractant receptor expressed both GIT1–PIX and GIT2–PIX endogenous complexes. We have used silencing of the different members of the complex by small interfering RNAs to study the effects on a number of events linked to agonist-induced cell migration. We found that cell adhesion was not affected by depletion of any of the proteins of the GIT complex, whereas agonist-enhanced cell spreading was inhibited. Analysis of agonist-stimulated haptotactic cell migration indicated a specific positive effect of GIT1 depletion on trans-well migration. The internalization of the formyl-peptide receptor was also inhibited by depletion of GIT1 and GIT2. The effects of the GIT complexes on trafficking of the receptors was confirmed by an antibody-enhanced agonist-induced internalization assay, showing that depletion of PIX, GIT1 or GIT2 protein caused decreased perinuclear accumulation of internalized receptors. Conclusions. Our results show that endogenous GIT complexes are involved in the regulation of chemoattractant-induced cell motility and receptor trafficking, and support previous findings indicating an important function of the GIT complexes in the regulation of different G-protein-coupled receptors. Our results also indicate that endogenous GIT1 and GIT2 regulate distinct subsets of agonist-induced responses and suggest a possible functional link between the control of receptor trafficking and the regulation of cell motility by GIT proteins. PMID:19912111

Exosome secretion promotes chemotaxis of cancer cells.

PubMed

Sung, Bong Hwan; Weaver, Alissa M

2017-03-04

Migration of cells toward chemical cues, or chemotaxis, is important for many biologic processes such as immune defense, wound healing and cancer metastasis. Although chemotaxis is thought to occur in cancer cells, it is less well characterized than chemotaxis of professional immune cells such as neutrophils. Here, we show that cancer cell chemotaxis relies on secretion of exosome-type extracellular vesicles. Migration of fibrosarcoma cells toward a gradient of exosome-depleted serum was diminished by knockdown of the exosome secretion regulator Rab27a. Rescue experiments in which chemotaxis chambers were coated with purified extracellular vesicles demonstrate that exosomes but not microvesicles affect both speed and directionality of migrating cells. Chamber coating with purified fibronectin and fibronectin-depleted exosomes demonstrates that the exosome cargo fibronectin promotes cell speed but cannot account for the role of exosomes in promoting directionality of fibrosarcoma cell movement during chemotaxis. These experiments indicate that exosomes contain multiple motility-promoting cargoes that contribute to different aspects of cell motility.

Bioconvection in Cultures of the Calcifying Unicellular Alga Pleurochrysis Carterae

NASA Technical Reports Server (NTRS)

Montufar-Solis, Dina; Duke, P. Jackie; Marsh, Mary E.

2003-01-01

The unicellular, marine, calcifying alga P leurochiysis carterae--a model to study cell morphogenesis, cell polarity, calcification, gravitaxis, reproduction and development-- has extremely flexible culture requirements. Support studies for a flight experiment addressing cell motility suggested that cell density (cells/ml) affects cell movement in P. carterae cultures through the gradual establishment of bioconvection as the culture grows. To assess the effect of cell density on direction of the movement, without the effects of aging of the culture, swimming behavior was analyzed in aliquots from a series of dilutions obtained from a stock culture. Results showed that at low concentrations cells swim randomly. As the concentration increases, upswimming patterns overtake random swimming. Gradually, up and down movement patterns prevail, representative of bioconvection. This oriented swimming of P. carterae occurs in a wide range of concentrations, adding to the list of flexible requirements, in this case, cell concentration, to be used for spaceflight studies addressing cell motility and bioconvection in a unicellular model of biologically directed mineralization.

Bioinformatic dissecting of TP53 regulation pathway underlying butyrate-induced histone modification in epigenetic regulation

USDA-ARS?s Scientific Manuscript database

Butyrate affects cell proliferation, differentiation and motility. Butyrate inhibits histone deacetylase (HDAC) activities and induces cell cycle arrest and apoptosis. TP53 is one of the most active upstream regulators discovered by IPA in our RNA sequencing data set. The TP53 signaling pathway pl...

Vancomycin Reduces Cell Wall Stiffness and Slows Swim Speed of the Lyme Disease Bacterium.

PubMed

Harman, Michael W; Hamby, Alex E; Boltyanskiy, Ross; Belperron, Alexia A; Bockenstedt, Linda K; Kress, Holger; Dufresne, Eric R; Wolgemuth, Charles W

2017-02-28

Borrelia burgdorferi, the spirochete that causes Lyme disease, is a tick-transmitted pathogen that requires motility to invade and colonize mammalian and tick hosts. These bacteria use a unique undulating flat-wave shape to penetrate and propel themselves through host tissues. Previous mathematical modeling has suggested that the morphology and motility of these spirochetes depends crucially on the flagellar/cell wall stiffness ratio. Here, we test this prediction using the antibiotic vancomycin to weaken the cell wall. We found that low to moderate doses of vancomycin (≤2.0 μg/mL for 24 h) produced small alterations in cell shape and that as the dose was increased, cell speed decreased. Vancomycin concentrations >1.0 μg/mL also inhibited cell growth and led to bleb formation on a fraction of the cells. To quantitatively assess how vancomycin affects cell stiffness, we used optical traps to bend unflagellated mutants of B. burgdorferi. We found that in the presence of vancomycin, cell wall stiffness gradually decreased over time, with a 40% reduction in the bending stiffness after 36 h. Under the same conditions, the swimming speed of wild-type B. burgdorferi slowed by ∼15%, with only marginal changes to cell morphology. Interestingly, our biophysical model for the swimming dynamics of B. burgdorferi suggested that cell speed should increase with decreasing cell stiffness. We show that this discrepancy can be resolved if the periplasmic volume decreases as the cell wall becomes softer. These results provide a testable hypothesis for how alterations of cell wall stiffness affect periplasmic volume regulation. Furthermore, since motility is crucial to the virulence of B. burgdorferi, the results suggest that sublethal doses of antibiotics could negatively impact spirochete survival by impeding their swim speed, thereby enabling their capture and elimination by phagocytes. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Computer-based identification of a novel LIMK1/2 inhibitor that synergizes with salirasib to destabilize the actin cytoskeleton.

PubMed

Mashiach-Farkash, Efrat; Rak, Roni; Elad-Sfadia, Galit; Haklai, Roni; Carmeli, Shmuel; Kloog, Yoel; Wolfson, Haim J

2012-06-01

Neurofibromin regulates cell motility via three distinct GTPase pathways acting through two different domains, the Ras GTPase-activating protein-related domain (GRD) and the pre-GRD domain. First, the GRD domain inhibits Ras-dependent changes in cell motility through the mitogen activated protein cascade. Second, it also regulates Rho-dependent (Ras-independent) changes by activating LIM kinase 2 (LIMK2), an enzyme that phosphorylates and inactivates cofilin (an actin-depolymerizing factor). Third, the pre-GRD domain acts through the Rac1 GTPase, that activate the P21 activated kinase 1 (PAK1)-LIMK1-cofilin pathway. We employed molecular modeling to identify a novel inhibitor of LIMK1/2. The active sites of an ephrin-A receptor (EphA3) and LIMK2 showed marked similarity (60%). On testing a known inhibitor of EphA3, we found that it fits to the LIMK1/2-ATP binding site and to the latter's substrate-binding pockets. We identified a similar compound, T56-LIMKi, and found that it inhibits LIMK1/2 kinase activities. It blocked the phosphorylation of cofilin which led to actin severance and inhibition of tumor cell migration, tumor cell growth, and anchorage-independent colony formation in soft agar. Because modulation of LIMK by neurofibromin is not affected by the Ras inhibitor Salirasib, we examined the combined effect of Salirasib and T56-LIMKi each of which can affect cell motility by a distinct pathway. We found that their combined action on cell proliferation and stress-fiber formation in neurofibromin-deficient cells was synergistic. We suggest that this drug combination may be developed for treatment of neurofibromatosis and cancer.

Computer-Based Identification of a Novel LIMK1/2 Inhibitor that Synergizes with Salirasib to Destabilize the Actin Cytoskeleton

PubMed Central

Elad-Sfadia, Galit; Haklai, Roni; Carmeli, Shmuel; Kloog, Yoel; Wolfson, Haim J.

2012-01-01

Neurofibromin regulates cell motility via three distinct GTPase pathways acting through two different domains, the Ras GTPase-activating protein-related domain (GRD) and the pre-GRD domain. First, the GRD domain inhibits Ras-dependent changes in cell motility through the mitogen activated protein cascade. Second, it also regulates Rho-dependent (Ras-independent) changes by activating LIM kinase 2 (LIMK2), an enzyme that phosphorylates and inactivates cofilin (an actin-depolymerizing factor). Third, the pre-GRD domain acts through the Rac1 GTPase, that activate the P21 activated kinase 1 (PAK1)-LIMK1-cofilin pathway. We employed molecular modeling to identify a novel inhibitor of LIMK1/2. The active sites of an ephrin-A receptor (EphA3) and LIMK2 showed marked similarity (60%). On testing a known inhibitor of EphA3, we found that it fits to the LIMK1/2-ATP binding site and to the latter's substrate-binding pockets. We identified a similar compound, T56-LIMKi, and found that it inhibits LIMK1/2 kinase activities. It blocked the phosphorylation of cofilin which led to actin severance and inhibition of tumor cell migration, tumor cell growth, and anchorage-independent colony formation in soft agar. Because modulation of LIMK by neurofibromin is not affected by the Ras inhibitor Salirasib, we examined the combined effect of Salirasib and T56-LIMKi each of which can affect cell motility by a distinct pathway. We found that their combined action on cell proliferation and stress-fiber formation in neurofibromin-deficient cells was synergistic. We suggest that this drug combination may be developed for treatment of neurofibromatosis and cancer. PMID:22776759

betaPIX controls cell motility and neurite extension by regulating the distribution of GIT1.

PubMed

Za, Lorena; Albertinazzi, Chiara; Paris, Simona; Gagliani, Mariacristina; Tacchetti, Carlo; de Curtis, Ivan

2006-07-01

Cell motility entails the reorganization of the cytoskeleton and membrane trafficking for effective protrusion. GIT1/p95-APP1 is a member of a family of GTPase-activating proteins for ARF GTPases that affect endocytosis, adhesion and migration. GIT1 associates with paxillin and a complex including the Rac/Cdc42 exchanging factors PIX/Cool and the kinase PAK. In this study, we show that overexpression of betaPIX induces the accumulation of endogenous and overexpressed GIT1 at large structures similar to those induced by an ArfGAP-defective mutant of GIT1 (p95-C2). Immunohistochemical analysis and immunoelectron microscopy reveal that these structures include the endogenous transferrin receptor. Time-lapse analysis during motogenic stimuli shows that the formation and perinuclear accumulation of the p95-C2-positive structures is paralleled by inhibition of lamellipodium formation and cell retraction. Both dimerization and a functional SH3 domain of betaPIX are required for the accumulation of GIT1 in fibroblasts, which is prevented by the monomeric PIX-PG-DeltaLZ. This mutant also prevents the formation of endocytic aggregates and inhibition of neurite outgrowth in retinal neurons expressing p95-C2. Our results indicate that betaPIX is an important regulator of the subcellular distribution of GIT1, and suggest that alteration in the level of expression of the complex affects the endocytic compartment and cell motility.

Colony Expansion of Socially Motile Myxococcus xanthus Cells Is Driven by Growth, Motility, and Exopolysaccharide Production

PubMed Central

Patra, Pintu; Kissoon, Kimberley; Cornejo, Isabel; Kaplan, Heidi B.; Igoshin, Oleg A.

2016-01-01

Myxococcus xanthus, a model organism for studies of multicellular behavior in bacteria, moves exclusively on solid surfaces using two distinct but coordinated motility mechanisms. One of these, social (S) motility is powered by the extension and retraction of type IV pili and requires the presence of exopolysaccharides (EPS) produced by neighboring cells. As a result, S motility requires close cell-to-cell proximity and isolated cells do not translocate. Previous studies measuring S motility by observing the colony expansion of cells deposited on agar have shown that the expansion rate increases with initial cell density, but the biophysical mechanisms involved remain largely unknown. To understand the dynamics of S motility-driven colony expansion, we developed a reaction-diffusion model describing the effects of cell density, EPS deposition and nutrient exposure on the expansion rate. Our results show that at steady state the population expands as a traveling wave with a speed determined by the interplay of cell motility and growth, a well-known characteristic of Fisher’s equation. The model explains the density-dependence of the colony expansion by demonstrating the presence of a lag phase–a transient period of very slow expansion with a duration dependent on the initial cell density. We propose that at a low initial density, more time is required for the cells to accumulate enough EPS to activate S-motility resulting in a longer lag period. Furthermore, our model makes the novel prediction that following the lag phase the population expands at a constant rate independent of the cell density. These predictions were confirmed by S motility experiments capturing long-term expansion dynamics. PMID:27362260

Colony Expansion of Socially Motile Myxococcus xanthus Cells Is Driven by Growth, Motility, and Exopolysaccharide Production.

PubMed

Patra, Pintu; Kissoon, Kimberley; Cornejo, Isabel; Kaplan, Heidi B; Igoshin, Oleg A

2016-06-01

Myxococcus xanthus, a model organism for studies of multicellular behavior in bacteria, moves exclusively on solid surfaces using two distinct but coordinated motility mechanisms. One of these, social (S) motility is powered by the extension and retraction of type IV pili and requires the presence of exopolysaccharides (EPS) produced by neighboring cells. As a result, S motility requires close cell-to-cell proximity and isolated cells do not translocate. Previous studies measuring S motility by observing the colony expansion of cells deposited on agar have shown that the expansion rate increases with initial cell density, but the biophysical mechanisms involved remain largely unknown. To understand the dynamics of S motility-driven colony expansion, we developed a reaction-diffusion model describing the effects of cell density, EPS deposition and nutrient exposure on the expansion rate. Our results show that at steady state the population expands as a traveling wave with a speed determined by the interplay of cell motility and growth, a well-known characteristic of Fisher's equation. The model explains the density-dependence of the colony expansion by demonstrating the presence of a lag phase-a transient period of very slow expansion with a duration dependent on the initial cell density. We propose that at a low initial density, more time is required for the cells to accumulate enough EPS to activate S-motility resulting in a longer lag period. Furthermore, our model makes the novel prediction that following the lag phase the population expands at a constant rate independent of the cell density. These predictions were confirmed by S motility experiments capturing long-term expansion dynamics.

Functional proteomic analysis reveals the involvement of KIAA1199 in breast cancer growth, motility and invasiveness

PubMed Central

2014-01-01

Background KIAA1199 is a recently identified novel gene that is up-regulated in human cancer with poor survival. Our proteomic study on signaling polarity in chemotactic cells revealed KIAA1199 as a novel protein target that may be involved in cellular chemotaxis and motility. In the present study, we examined the functional significance of KIAA1199 expression in breast cancer growth, motility and invasiveness. Methods We validated the previous microarray observation by tissue microarray immunohistochemistry using a TMA slide containing 12 breast tumor tissue cores and 12 corresponding normal tissues. We performed the shRNA-mediated knockdown of KIAA1199 in MDA-MB-231 and HS578T cells to study the role of this protein in cell proliferation, migration and apoptosis in vitro. We studied the effects of KIAA1199 knockdown in vivo in two groups of mice (n = 5). We carried out the SILAC LC-MS/MS based proteomic studies on the involvement of KIAA1199 in breast cancer. Results KIAA1199 mRNA and protein was significantly overexpressed in breast tumor specimens and cell lines as compared with non-neoplastic breast tissues from large-scale microarray and studies of breast cancer cell lines and tumors. To gain deeper insights into the novel role of KIAA1199 in breast cancer, we modulated KIAA1199 expression using shRNA-mediated knockdown in two breast cancer cell lines (MDA-MB-231 and HS578T), expressing higher levels of KIAA1199. The KIAA1199 knockdown cells showed reduced motility and cell proliferation in vitro. Moreover, when the knockdown cells were injected into the mammary fat pads of female athymic nude mice, there was a significant decrease in tumor incidence and growth. In addition, quantitative proteomic analysis revealed that knockdown of KIAA1199 in breast cancer (MDA-MB-231) cells affected a broad range of cellular functions including apoptosis, metabolism and cell motility. Conclusions Our findings indicate that KIAA1199 may play an important role in breast tumor growth and invasiveness, and that it may represent a novel target for biomarker development and a novel therapeutic target for breast cancer. PMID:24628760

Computational approaches to substrate-based cell motility

DOE PAGES

Ziebert, Falko; Aranson, Igor S.

2016-07-15

Substrate-based crawling motility of eukaryotic cells is essential for many biological functions, both in developing and mature organisms. Motility dysfunctions are involved in several life-threatening pathologies such as cancer and metastasis. Motile cells are also a natural realization of active, self-propelled ‘particles’, a popular research topic in nonequilibrium physics. Finally, from the materials perspective, assemblies of motile cells and evolving tissues constitute a class of adaptive self-healing materials that respond to the topography, elasticity, and surface chemistry of the environment and react to external stimuli. Although a comprehensive understanding of substrate-based cell motility remains elusive, progress has been achieved recentlymore » in its modeling on the whole cell level. Furthermore we survey the most recent advances in computational approaches to cell movement and demonstrate how these models improve our understanding of complex self-organized systems such as living cells.« less

RSK is a principal effector of the RAS-ERK pathway for eliciting a coordinate, pro-motile/invasive gene program and phenotype in epithelial cells

PubMed Central

Doehn, Ulrik; Hauge, Camilla; Frank, Scott R.; Jensen, Claus J.; Duda, Katarzyna; Nielsen, Jakob V.; Cohen, Michael S.; Johansen, Jens V.; Winther, Benny R.; Lund, Leif R.; Winther, Ole; Taunton, Jack; Hansen, Steen H.; Frödin, Morten

2013-01-01

SUMMARY The RAS-stimulated RAF-MEK-ERK pathway confers epithelial cells with critical motile and invasive capacities during embryonic development, tissue regeneration and carcinoma progression. Yet many mechanisms by which ERK exerts this control remain elusive. Here, we demonstrate that the ERK-activated kinase RSK is necessary to induce motility and invasive capacities in non-transformed epithelial cells and carcinoma cells. RSK is moreover sufficient to induce certain motile responses. Expression profiling analysis revealed that a primary role of RSK is to induce transcription of potent pro-motile/invasive gene program by FRA1-dependent and independent mechanisms. Strikingly, the program enables RSK to coordinately modulate the extracellular environment, the intracellular motility apparatus, and receptors mediating communication between these compartments to stimulate motility and invasion. These findings uncover a general mechanism whereby the RAS-ERK pathway controls epithelial cell motility by identifying RSK as a key effector, from which emanates multiple highly coordinate transcription-dependent mechanisms for stimulation of motility and invasive properties. PMID:19716794

Involvement of gut microbiota in association between GLP-1/GLP-1 receptor expression and gastrointestinal motility.

PubMed

Yang, Mo; Fukui, Hirokazu; Eda, Hirotsugu; Xu, Xin; Kitayama, Yoshitaka; Hara, Ken; Kodani, Mio; Tomita, Toshihiko; Oshima, Tadayuki; Watari, Jiro; Miwa, Hiroto

2017-04-01

The microbiota in the gut is known to play a pivotal role in host physiology by interacting with the immune and neuroendocrine systems in gastrointestinal (GI) tissues. Glucagon-like peptide 1 (GLP-1), a gut hormone, is involved in metabolism as well as GI motility. We examined how gut microbiota affects the link between GLP-1/GLP-1 receptor (GLP-1R) expression and motility of the GI tract. Germ-free (GF) mice (6 wk old) were orally administered a fecal bacterial suspension prepared from specific pathogen-free (SPF) mice, and then after fecal transplantation (FT) GI tissues were obtained from the GF mice at various time points. The expression of GLP-1 and its receptor was examined by immunohistochemistry, and gastrointestinal transit time (GITT) was measured by administration of carmine red solution. GLP-1 was expressed in endocrine cells in the colonic mucosa, and GLP-1R was expressed in myenteric neural cells throughout the GI wall. GLP-1R-positive cells throughout the GI wall were significantly fewer in GF mice with FT than in GF mice without gut microbiota reconstitution. GITT was significantly shorter in GF mice with FT than in control GF mice without FT and correlated with the number of GLP-1R-positive cells throughout the GI wall. GITT was significantly longer in GF control mice than in SPF mice. When those mice were treated with GLP-1 agonist extendin4, GITT was significantly longer in the GF mice. The gut microbiota may accelerate or at least modify GI motility while suppressing GLP-1R expression in myenteric neural cells throughout the GI tract. NEW & NOTEWORTHY The gut microbiota has been intensively studied, because it plays a pivotal role in various aspects of host physiology. On the other hand, glucagon-like peptide 1 (GLP-1) plays important roles in metabolism as well as gastrointestinal motility. In the present study, we have suggested that the gut microbiota accelerates gastrointestinal motility while suppressing the expression of GLP-1 receptor in myenteric neural cells throughout the gastrointestinal tract. We believe that this article is very timely and suggestive work. Copyright © 2017 the American Physiological Society.

On-command on/off switching of progenitor cell and cancer cell polarized motility and aligned morphology via a cytocompatible shape memory polymer scaffold.

PubMed

Wang, Jing; Quach, Andy; Brasch, Megan E; Turner, Christopher E; Henderson, James H

2017-09-01

In vitro biomaterial models have enabled advances in understanding the role of extracellular matrix (ECM) architecture in the control of cell motility and polarity. Most models are, however, static and cannot mimic dynamic aspects of in vivo ECM remodeling and function. To address this limitation, we present an electrospun shape memory polymer scaffold that can change fiber alignment on command under cytocompatible conditions. Cellular response was studied using the human fibrosarcoma cell line HT-1080 and the murine mesenchymal stem cell line C3H/10T1/2. The results demonstrate successful on-command on/off switching of cell polarized motility and alignment. Decrease in fiber alignment causes a change from polarized motility along the direction of fiber alignment to non-polarized motility and from aligned to unaligned morphology, while increase in fiber alignment causes a change from non-polarized to polarized motility along the direction of fiber alignment and from unaligned to aligned morphology. In addition, the findings are consistent with the hypothesis that increased fiber alignment causes increased cell velocity, while decreased fiber alignment causes decreased cell velocity. On-command on/off switching of cell polarized motility and alignment is anticipated to enable new study of directed cell motility in tumor metastasis, in cell homing, and in tissue engineering. Copyright © 2017 Elsevier Ltd. All rights reserved.

Mitochondrial respiration is sensitive to cytoarchitectural breakdown.

PubMed

Kandel, Judith; Angelin, Alessia A; Wallace, Douglas C; Eckmann, David M

2016-11-07

An abundance of research suggests that cellular mitochondrial and cytoskeletal disruption are related, but few studies have directly investigated causative connections between the two. We previously demonstrated that inhibiting microtubule and microfilament polymerization affects mitochondrial motility on the whole-cell level in fibroblasts. Since mitochondrial motility can be indicative of mitochondrial function, we now further characterize the effects of these cytoskeletal inhibitors on mitochondrial potential, morphology and respiration. We found that although they did not reduce mitochondrial inner membrane potential, cytoskeletal toxins induced significant decreases in basal mitochondrial respiration. In some cases, basal respiration was only affected after cells were pretreated with the calcium ionophore A23187 in order to stress mitochondrial function. In most cases, mitochondrial morphology remained unaffected, but extreme microfilament depolymerization or combined intermediate doses of microtubule and microfilament toxins resulted in decreased mitochondrial lengths. Interestingly, these two particular exposures did not affect mitochondrial respiration in cells not sensitized with A23187, indicating an interplay between mitochondrial morphology and respiration. In all cases, inducing maximal respiration diminished differences between control and experimental groups, suggesting that reduced basal respiration originates as a largely elective rather than pathological symptom of cytoskeletal impairment. However, viability experiments suggest that even this type of respiration decrease may be associated with cell death.

Pattern-formation mechanisms in motility mutants of Myxococcus xanthus

PubMed Central

Starruß, Jörn; Peruani, Fernando; Jakovljevic, Vladimir; Søgaard-Andersen, Lotte; Deutsch, Andreas; Bär, Markus

2012-01-01

Formation of spatial patterns of cells is a recurring theme in biology and often depends on regulated cell motility. Motility of the rod-shaped cells of the bacterium Myxococcus xanthus depends on two motility machineries, type IV pili (giving rise to S-motility) and the gliding motility apparatus (giving rise to A-motility). Cell motility is regulated by occasional reversals. Moving M. xanthus cells can organize into spreading colonies or spore-filled fruiting bodies, depending on their nutritional status. To ultimately understand these two pattern-formation processes and the contributions by the two motility machineries, as well as the cell reversal machinery, we analyse spatial self-organization in three M. xanthus strains: (i) a mutant that moves unidirectionally without reversing by the A-motility system only, (ii) a unidirectional mutant that is also equipped with the S-motility system, and (iii) the wild-type that, in addition to the two motility systems, occasionally reverses its direction of movement. The mutant moving by means of the A-engine illustrates that collective motion in the form of large moving clusters can arise in gliding bacteria owing to steric interactions of the rod-shaped cells, without the need of invoking any biochemical signal regulation. The two-engine strain mutant reveals that the same phenomenon emerges when both motility systems are present, and as long as cells exhibit unidirectional motion only. From the study of these two strains, we conclude that unidirectional cell motion induces the formation of large moving clusters at low and intermediate densities, while it results in vortex formation at very high densities. These findings are consistent with what is known from self-propelled rod models, which strongly suggests that the combined effect of self-propulsion and volume exclusion interactions is the pattern-formation mechanism leading to the observed phenomena. On the other hand, we learn that when cells occasionally reverse their moving direction, as observed in the wild-type, cells form small but strongly elongated clusters and self-organize into a mesh-like structure at high enough densities. These results have been obtained from a careful analysis of the cluster statistics of ensembles of cells, and analysed in the light of a coagulation Smoluchowski equation with fragmentation. PMID:24312730

The cell fate determinant Scribble is required for maintenance of hematopoietic stem cell function.

PubMed

Mohr, Juliane; Dash, Banaja P; Schnoeder, Tina M; Wolleschak, Denise; Herzog, Carolin; Tubio Santamaria, Nuria; Weinert, Sönke; Godavarthy, Sonika; Zanetti, Costanza; Naumann, Michael; Hartleben, Björn; Huber, Tobias B; Krause, Daniela S; Kähne, Thilo; Bullinger, Lars; Heidel, Florian H

2018-05-01

Cell fate determinants influence self-renewal potential of hematopoietic stem cells. Scribble and Llgl1 belong to the Scribble polarity complex and reveal tumor-suppressor function in drosophila. In hematopoietic cells, genetic inactivation of Llgl1 leads to expansion of the stem cell pool and increases self-renewal capacity without conferring malignant transformation. Here we show that genetic inactivation of its putative complex partner Scribble results in functional impairment of hematopoietic stem cells (HSC) over serial transplantation and during stress. Although loss of Scribble deregulates transcriptional downstream effectors involved in stem cell proliferation, cell signaling, and cell motility, these effectors do not overlap with transcriptional targets of Llgl1. Binding partner analysis of Scribble in hematopoietic cells using affinity purification followed by mass spectometry confirms its role in cell signaling and motility but not for binding to polarity modules described in drosophila. Finally, requirement of Scribble for self-renewal capacity also affects leukemia stem cell function. Thus, Scribble is a regulator of adult HSCs, essential for maintenance of HSCs during phases of cell stress.

Membrane tension feedback on shape and motility of eukaryotic cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Winkler, Benjamin; Aranson, Igor S.; Ziebert, Falko

2016-04-01

In the framework of a phase field model of a single cell crawling on a substrate, we investigate how the properties of the cell membrane affect the shape and motility of the cell. Since the membrane influences the cell dynamics on multiple levels and provides a nontrivial feedback, we consider the following fundamental interactions: (i) the reduction of the actin polymerization rate by membrane tension; (ii) area conservation of the cell’s two-dimensional cross-section vs. conservation of the circumference (i.e. membrane inextensibility); and (iii) the contribution from the membrane’s bending energy to the shape and integrity of the cell. As inmore » experiments, we investigate two pertinent observables — the cell’s velocity and its aspect ratio. We find that the most important effect is the feedback of membrane tension on the actin polymerization. Bending rigidity has only minor effects, visible mostly in dynamic reshaping events, as exemplified by collisions of the cell with an obstacle.« less

Mathematical models of cell motility.

PubMed

Flaherty, Brendan; McGarry, J P; McHugh, P E

2007-01-01

Cell motility is an essential biological action in the creation, operation and maintenance of our bodies. Developing mathematical models elucidating cell motility will greatly advance our understanding of this fundamental biological process. With accurate models it is possible to explore many permutations of the same event and concisely investigate their outcome. While great advancements have been made in experimental studies of cell motility, it now has somewhat fallen on mathematical models to taking a leading role in future developments. The obvious reason for this is the complexity of cell motility. Employing the processing power of today's computers will give researches the ability to run complex biophysical and biochemical scenarios, without the inherent difficulty and time associated with in vitro investigations. Before any great advancement can be made, the basics of cell motility will have to be well-defined. Without this, complicated mathematical models will be hindered by their inherent conjecture. This review will look at current mathematical investigations of cell motility, explore the reasoning behind such work and conclude with how best to advance this interesting and challenging research area.

Shear-induced orientational dynamics and spatial heterogeneity in suspensions of motile phytoplankton

PubMed Central

Barry, Michael T.; Rusconi, Roberto; Guasto, Jeffrey S.; Stocker, Roman

2015-01-01

Fluid flow, ubiquitous in natural and man-made environments, has the potential to profoundly impact the transport of microorganisms, including phytoplankton in aquatic habitats and bioreactors. Yet, the effect of ambient flow on the swimming behaviour of phytoplankton has remained poorly understood, largely owing to the difficulty of observing cell–flow interactions at the microscale. Here, we present microfluidic experiments where we tracked individual cells for four species of motile phytoplankton exposed to a spatially non-uniform fluid shear rate, characteristic of many flows in natural and artificial environments. We observed that medium-to-high mean shear rates (1–25 s−1) produce heterogeneous cell concentrations in the form of regions of accumulation and regions of depletion. The location of these regions relative to the flow depends on the cells' propulsion mechanism, body shape and flagellar arrangement, as captured by an effective aspect ratio. Species having a large effective aspect ratio accumulated in the high-shear regions, owing to shear-induced alignment of the swimming orientation with the fluid streamlines. Species having an effective aspect ratio close to unity exhibited little preferential accumulation at low-to-moderate flow rates, but strongly accumulated in the low-shear regions under high flow conditions, potentially owing to an active, behavioural response of cells to shear. These observations demonstrate that ambient fluid flow can strongly affect the motility and spatial distribution of phytoplankton and highlight the rich dynamics emerging from the interaction between motility, morphology and flow. PMID:26538558

Cell motility in cancer invasion and metastasis: insights from simple model organisms.

PubMed

Stuelten, Christina H; Parent, Carole A; Montell, Denise J

2018-05-01

Metastasis remains the greatest challenge in the clinical management of cancer. Cell motility is a fundamental and ancient cellular behaviour that contributes to metastasis and is conserved in simple organisms. In this Review, we evaluate insights relevant to human cancer that are derived from the study of cell motility in non-mammalian model organisms. Dictyostelium discoideum, Caenorhabditis elegans, Drosophila melanogaster and Danio rerio permit direct observation of cells moving in complex native environments and lend themselves to large-scale genetic and pharmacological screening. We highlight insights derived from each of these organisms, including the detailed signalling network that governs chemotaxis towards chemokines; a novel mechanism of basement membrane invasion; the positive role of E-cadherin in collective direction-sensing; the identification and optimization of kinase inhibitors for metastatic thyroid cancer on the basis of work in flies; and the value of zebrafish for live imaging, especially of vascular remodelling and interactions between tumour cells and host tissues. While the motility of tumour cells and certain host cells promotes metastatic spread, the motility of tumour-reactive T cells likely increases their antitumour effects. Therefore, it is important to elucidate the mechanisms underlying all types of cell motility, with the ultimate goal of identifying combination therapies that will increase the motility of beneficial cells and block the spread of harmful cells.

Epidermal keratinocyte polarity and motility require Ca2+ influx through TRPV1

PubMed Central

Graham, David M.; Huang, Ling; Robinson, Kenneth R.; Messerli, Mark A.

2013-01-01

Summary Ca2+ has long been known to play an important role in cellular polarity and guidance. We studied the role of Ca2+ signaling during random and directed cell migration to better understand whether Ca2+ directs cell motility from the leading edge and which ion channels are involved in this function by using primary zebrafish keratinocytes. Rapid line-scan and time-lapse imaging of intracellular Ca2+ (Ca2+i) during migration and automated image alignment enabled us to characterize and map the spatiotemporal changes in Ca2+i. We show that asymmetric distributions of lamellipodial Ca2+ sparks are encoded in frequency, not amplitude, and that they correlate with cellular rotation during migration. Directed migration during galvanotaxis increases the frequency of Ca2+ sparks over the entire lamellipod; however, these events do not give rise to asymmetric Ca2+i signals that correlate with turning. We demonstrate that Ca2+-permeable channels within these cells are mechanically activated and include several transient receptor potential family members, including TRPV1. Last, we demonstrate that cell motility and Ca2+i activity are affected by pharmacological agents that target TRPV1, indicating a novel role for this channel during cell migration. PMID:23943873

Sperm motility in fishes. (II) Effects of ions and osmolality: a review.

PubMed

Alavi, Sayyed Mohammad Hadi; Cosson, Jacky

2006-01-01

The spermatozoa of most fish species are immotile in the testis and seminal plasma. Therefore, motility is induced after the spermatozoa are released into the aqueous environment during natural reproduction or into the diluent during artificial reproduction. There are clear relationships between seminal plasma composition and osmolality and the duration of fish sperm motility. Various parameters such as ion concentrations (K+, Na+, and Ca2+), osmotic pressure, pH, temperature and dilution rate affect motility. In the present paper, we review the roles of these ions on sperm motility in Salmonidae, Cyprinidae, Acipenseridae and marine fishes, and their relationship with seminal plasma composition. Results in the literature show that: 1. K+ is a key ion controlling sperm motility in Salmonidae and Acipenseridae in combination with osmotic pressure; this control is more simple in other fish species: sperm motility is prevented when the osmotic pressure is high (Cyprinidae) or low (marine fishes) compared to that of the seminal fluid. 2. Cations (mostly divalent, such as Ca2+) are antagonistic with the inhibitory effect of K+ on sperm motility. 3. In many species, Ca2+ influx and K+ or Na+ efflux through specific ionic channels change the membrane potential and eventually lead to an increase in cAMP concentration in the cell, which constitutes the initiation signal for sperm motility in Salmonidae. 4. Media that are hyper- and hypo-osmotic relative to seminal fluid trigger sperm motility in marine and freshwater fishes, respectively. 5. The motility of fish spermatozoa is controlled through their sensitivity to osmolality and ion concentrations. This phenomenon is related to ionic channel activities in the membrane and governs the motility mechanisms of axonemes.

Cytoskeletal self-organization in neuromorphogenesis.

PubMed

Dehmelt, Leif

2014-01-01

Self-organization of dynamic microtubules via interactions with associated motors plays a critical role in spindle formation. The microtubule-based mechanisms underlying other aspects of cellular morphogenesis, such as the formation and development of protrusions from neuronal cells is less well understood. In a recent study, we investigated the molecular mechanism that underlies the massive reorganization of microtubules induced in non-neuronal cells by expression of the neuronal microtubule stabilizer MAP2c. In that study we directly observed cortical dynein complexes and how they affect the dynamic behavior of motile microtubules in living cells. We found that stationary dynein complexes transiently associate with motile microtubules near the cell cortex and that their rapid turnover facilitates efficient microtubule transport. Here, we discuss our findings in the larger context of cellular morphogenesis with specific focus on self-organizing principles from which cellular shape patterns such as the thin protrusions of neurons can emerge.

Effect of Cell Aspect Ratio on Swarming Bacteria

NASA Astrophysics Data System (ADS)

Ilkanaiv, Bella; Kearns, Daniel B.; Ariel, Gil; Be'er, Avraham

2017-04-01

Swarming bacteria collectively migrate on surfaces using flagella, forming dynamic whirls and jets that consist of millions of individuals. Because some swarming bacteria elongate prior to actual motion, cell aspect ratio may play a significant role in the collective dynamics. Extensive research on self-propelled rodlike particles confirms that elongation promotes alignment, strongly affecting the dynamics. Here, we study experimentally the collective dynamics of variants of swarming Bacillus subtilis that differ in length. We show that the swarming statistics depends on the aspect ratio in a critical, fundamental fashion not predicted by theory. The fastest motion was obtained for the wild-type and variants that are similar in length. However, shorter and longer cells exhibit anomalous, non-Gaussian statistics and nonexponential decay of the autocorrelation function, indicating lower collective motility. These results suggest that the robust mechanisms to maintain aspect ratios may be important for efficient swarming motility. Wild-type cells are optimal in this sense.

Cytoskeletal self-organization in neuromorphogenesis

PubMed Central

Dehmelt, Leif

2014-01-01

Self-organization of dynamic microtubules via interactions with associated motors plays a critical role in spindle formation. The microtubule-based mechanisms underlying other aspects of cellular morphogenesis, such as the formation and development of protrusions from neuronal cells is less well understood. In a recent study, we investigated the molecular mechanism that underlies the massive reorganization of microtubules induced in non-neuronal cells by expression of the neuronal microtubule stabilizer MAP2c. In that study we directly observed cortical dynein complexes and how they affect the dynamic behavior of motile microtubules in living cells. We found that stationary dynein complexes transiently associate with motile microtubules near the cell cortex and that their rapid turnover facilitates efficient microtubule transport. Here, we discuss our findings in the larger context of cellular morphogenesis with specific focus on self-organizing principles from which cellular shape patterns such as the thin protrusions of neurons can emerge. PMID:24847718

Turbulent unmixing: how marine turbulence drives patchy distributions of motile phytoplankton

NASA Astrophysics Data System (ADS)

Durham, William; Climent, Eric; Barry, Michael; de Lillo, Filippo; Boffetta, Guido; Cencini, Massimo; Stocker, Roman

2013-11-01

Centimeter-scale patchiness in the distribution of phytoplankton increases the efficacy of many important ecological interactions in the marine food web. We show that turbulent fluid motion, usually synonymous with mixing, instead triggers intense small-scale patchiness in the distribution of motile phytoplankton. We use a suite of experiments, direct numerical simulations of turbulence, and analytical tools to show that turbulent shear and acceleration directs the motility of cells towards well-defined regions of flow, increasing local cell concentrations more than ten fold. This motility-driven `unmixing' offers an explanation for why motile cells are often more patchily distributed than non-motile cells and provides a mechanistic framework to understand how turbulence, whose strength varies profoundly in marine environments, impacts ocean productivity.

Exopolysaccharide-Independent Social Motility of Myxococcus xanthus

PubMed Central

Hu, Wei; Hossain, Muhaiminu; Lux, Renate; Wang, Jing; Yang, Zhe; Li, Yuezhong; Shi, Wenyuan

2011-01-01

Social motility (S motility), the coordinated movement of large cell groups on agar surfaces, of Myxococcus xanthus requires type IV pili (TFP) and exopolysaccharides (EPS). Previous models proposed that this behavior, which only occurred within cell groups, requires cycles of TFP extension and retraction triggered by the close interaction of TFP with EPS. However, the curious observation that M. xanthus can perform TFP-dependent motility at a single-cell level when placed onto polystyrene surfaces in a highly viscous medium containing 1% methylcellulose indicated that “S motility” is not limited to group movements. In an apparent further challenge of the previous findings for S motility, mutants defective in EPS production were found to perform TFP-dependent motility on polystyrene surface in methylcellulose-containing medium. By exploring the interactions between pilin and surface materials, we found that the binding of TFP onto polystyrene surfaces eliminated the requirement for EPS in EPS- cells and thus enabled TFP-dependent motility on a single cell level. However, the presence of a general anchoring surface in a viscous environment could not substitute for the role of cell surface EPS in group movement. Furthermore, EPS was found to serve as a self-produced anchoring substrate that can be shed onto surfaces to enable cells to conduct TFP-dependent motility regardless of surface properties. These results suggested that in certain environments, such as in methylcellulose solution, the cells could bypass the need for EPS to anchor their TPF and conduct single-cell S motility to promote exploratory movement of colonies over new specific surfaces. PMID:21245931

Enforcing host cell polarity: an apicomplexan parasite strategy towards dissemination.

PubMed

Baumgartner, Martin

2011-08-01

The propagation of apicomplexan parasites through transmitting vectors is dependent on effective dissemination of parasites inside the mammalian host. Intracellular Toxoplasma and Theileria parasites face the challenge that their spread inside the host depends in part on the motile capacities of their host cells. In response, these parasites influence the efficiency of dissemination by altering adhesive and/or motile properties of their host cells. Theileria parasites do so by targeting signalling pathways that control host cell actin dynamics. The resulting enforced polar host cell morphology facilitates motility and invasiveness, by establishing focal adhesion and invasion structures at the leading edge of the infected cell. This parasite strategy highlights mechanisms of motility regulation that are also likely relevant for immune or cancer cell motility. Copyright © 2011 Elsevier Ltd. All rights reserved.

Regulation of T-lymphocyte motility, adhesion and de-adhesion by a cell surface mechanism directed by low density lipoprotein receptor-related protein 1 and endogenous thrombospondin-1.

PubMed

Talme, Toomas; Bergdahl, Eva; Sundqvist, Karl-Gösta

2014-06-01

T lymphocytes are highly motile and constantly reposition themselves between a free-floating vascular state, transient adhesion and migration in tissues. The regulation behind this unique dynamic behaviour remains unclear. Here we show that T cells have a cell surface mechanism for integrated regulation of motility and adhesion and that integrin ligands and CXCL12/SDF-1 influence motility and adhesion through this mechanism. Targeting cell surface-expressed low-density lipoprotein receptor-related protein 1 (LRP1) with an antibody, or blocking transport of LRP1 to the cell surface, perturbed the cell surface distribution of endogenous thrombospondin-1 (TSP-1) while inhibiting motility and potentiating cytoplasmic spreading on intercellular adhesion molecule 1 (ICAM-1) and fibronectin. Integrin ligands and CXCL12 stimulated motility and enhanced cell surface expression of LRP1, intact TSP-1 and a 130,000 MW TSP-1 fragment while preventing formation of a de-adhesion-coupled 110 000 MW TSP-1 fragment. The appearance of the 130 000 MW TSP-1 fragment was inhibited by the antibody that targeted LRP1 expression, inhibited motility and enhanced spreading. The TSP-1 binding site in the LRP1-associated protein, calreticulin, stimulated adhesion to ICAM-1 through intact TSP-1 and CD47. Shear flow enhanced cell surface expression of intact TSP-1. Hence, chemokines and integrin ligands up-regulate a dominant motogenic pathway through LRP1 and TSP-1 cleavage and activate an associated adhesion pathway through the LRP1-calreticulin complex, intact TSP-1 and CD47. This regulation of T-cell motility and adhesion makes pro-adhesive stimuli favour motile responses, which may explain why T cells prioritize movement before permanent adhesion.

Electrical stimulation of gut motility guided by an in silico model

NASA Astrophysics Data System (ADS)

Barth, Bradley B.; Henriquez, Craig S.; Grill, Warren M.; Shen, Xiling

2017-12-01

Objective. Neuromodulation of the central and peripheral nervous systems is becoming increasingly important for treating a diverse set of diseases—ranging from Parkinson’s Disease and epilepsy to chronic pain. However, neuromodulation of the gastrointestinal (GI) tract has achieved relatively limited success in treating functional GI disorders, which affect a significant population, because the effects of stimulation on the enteric nervous system (ENS) and gut motility are not well understood. Here we develop an integrated neuromechanical model of the ENS and assess neurostimulation strategies for enhancing gut motility, validated by in vivo experiments. Approach. The computational model included a network of enteric neurons, smooth muscle fibers, and interstitial cells of Cajal, which regulated propulsion of a virtual pellet in a model of gut motility. Main results. Simulated extracellular stimulation of ENS-mediated motility revealed that sinusoidal current at 0.5 Hz was more effective at increasing intrinsic peristalsis and reducing colon transit time than conventional higher frequency rectangular current pulses, as commonly used for neuromodulation therapy. Further analysis of the model revealed that the 0.5 Hz sinusoidal currents were more effective at modulating the pacemaker frequency of interstitial cells of Cajal. To test the predictions of the model, we conducted in vivo electrical stimulation of the distal colon while measuring bead propulsion in awake rats. Experimental results confirmed that 0.5 Hz sinusoidal currents were more effective than higher frequency pulses at enhancing gut motility. Significance. This work demonstrates an in silico GI neuromuscular model to enable GI neuromodulation parameter optimization and suggests that low frequency sinusoidal currents may improve the efficacy of GI pacing.

Higher cell stiffness indicating lower metastatic potential in B16 melanoma cell variants and in (-)-epigallocatechin gallate-treated cells.

PubMed

Watanabe, Tatsuro; Kuramochi, Hiromi; Takahashi, Atsushi; Imai, Kazue; Katsuta, Naoko; Nakayama, Tomonobu; Fujiki, Hirota; Suganuma, Masami

2012-05-01

To understand how nanomechanical stiffness affects metastatic potential, we studied the relationship between cell migration, a characteristic of metastasis, and cell stiffness using atomic force microscopy (AFM), which can measure stiffness (elasticity) of individual living cells. Migration and cell stiffness of three metastatic B16 melanoma variants (B16-F10, B16-BL6, and B16-F1 cells), and also effects of (-)-epigallocatechin gallate (EGCG), were studied using Transwell assay and AFM. Migration of B16-F10 and B16-BL6 cells was 3 and 2 times higher than that of B16-F1 cells in Transwell assay, and cell stiffness determined by AFM was also different among the three variants, although they have similar morphologies and the same growth rates: Means of Young's modulus were 350.8 ± 4.8 Pa for B16-F10 cells, 661.9 ± 16.5 Pa for B16-BL6 cells, and 727.2 ± 13.0 Pa for B16-F1 cells. AFM measurements revealed that highly motile B16-F10 cells have low cell stiffness, and low motile and metastatic B16-F1 cells have high cell stiffness: Nanomechanical stiffness is inversely correlated with migration potential. Treatment of highly motile B16-F10 cells with EGCG increased cell stiffness 2-fold and inhibited migration of the cells. Our study with AFM clearly demonstrates that cell stiffness is a reliable quantitative indicator of migration potential, and very likely metastatic potential, even in morphologically similar cells. And increased cell stiffness may be a key nanomechanical feature in inhibition of metastasis.

Oncofetal Chondroitin Sulfate Glycosaminoglycans Are Key Players in Integrin Signaling and Tumor Cell Motility.

PubMed

Clausen, Thomas Mandel; Pereira, Marina Ayres; Al Nakouzi, Nader; Oo, Htoo Zarni; Agerbæk, Mette Ø; Lee, Sherry; Ørum-Madsen, Maj Sofie; Kristensen, Anders Riis; El-Naggar, Amal; Grandgenett, Paul M; Grem, Jean L; Hollingsworth, Michael A; Holst, Peter J; Theander, Thor; Sorensen, Poul H; Daugaard, Mads; Salanti, Ali

2016-12-01

Many tumors express proteoglycans modified with oncofetal chondroitin sulfate glycosaminoglycan chains (ofCS), which are normally restricted to the placenta. However, the role of ofCS in cancer is largely unknown. The function of ofCS in cancer was analyzed using the recombinant ofCS-binding VAR2CSA protein (rVAR2) derived from the malaria parasite, Plasmodium falciparum We demonstrate that ofCS plays a key role in tumor cell motility by affecting canonical integrin signaling pathways. Binding of rVAR2 to tumor cells inhibited the interaction of cells with extracellular matrix (ECM) components, which correlated with decreased phosphorylation of Src kinase. Moreover, rVAR2 binding decreased migration, invasion, and anchorage-independent growth of tumor cells in vitro Mass spectrometry of ofCS-modified proteoglycan complexes affinity purified from tumor cell lines on rVAR2 columns revealed an overrepresentation of proteins involved in cell motility and integrin signaling, such as integrin-β1 (ITGB1) and integrin-α4 (ITGA4). Saturating concentrations of rVAR2 inhibited downstream integrin signaling, which was mimicked by knockdown of the core chondroitin sulfate synthesis enzymes β-1,3-glucuronyltransferase 1 (B3GAT1) and chondroitin sulfate N-acetylgalactosaminyltransferase 1 (CSGALNACT1). The ofCS modification was highly expressed in both human and murine metastatic lesions in situ and preincubation or early intravenous treatment of tumor cells with rVAR2 inhibited seeding and spreading of tumor cells in mice. This was associated with a significant increase in survival of the animals. These data functionally link ofCS modifications with cancer cell motility and further highlights ofCS as a novel therapeutic cancer target. The cancer-specific expression of ofCS aids in metastatic phenotypes and is a candidate target for therapy. Mol Cancer Res; 14(12); 1288-99. ©2016 AACR. ©2016 American Association for Cancer Research.

C-NAP1 and rootletin restrain DNA damage-induced centriole splitting and facilitate ciliogenesis.

PubMed

Conroy, Pauline C; Saladino, Chiara; Dantas, Tiago J; Lalor, Pierce; Dockery, Peter; Morrison, Ciaran G

2012-10-15

Cilia are found on most human cells and exist as motile cilia or non-motile primary cilia. Primary cilia play sensory roles in transducing various extracellular signals, and defective ciliary functions are involved in a wide range of human diseases. Centrosomes are the principal microtubule-organizing centers of animal cells and contain two centrioles. We observed that DNA damage causes centriole splitting in non-transformed human cells, with isolated centrioles carrying the mother centriole markers CEP170 and ninein but not kizuna or cenexin. Loss of centriole cohesion through siRNA depletion of C-NAP1 or rootletin increased radiation-induced centriole splitting, with C-NAP1-depleted isolated centrioles losing mother markers. As the mother centriole forms the basal body in primary cilia, we tested whether centriole splitting affected ciliogenesis. While irradiated cells formed apparently normal primary cilia, most cilia arose from centriolar clusters, not from isolated centrioles. Furthermore, C-NAP1 or rootletin knockdown reduced primary cilium formation. Therefore, the centriole cohesion apparatus at the proximal end of centrioles may provide a target that can affect primary cilium formation as part of the DNA damage response.

Lysophosphatidic acid receptor-5 negatively regulates cellular responses in mouse fibroblast 3T3 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dong, Yan; Hirane, Miku; Araki, Mutsumi

2014-04-04

Highlights: • LPA{sub 5} inhibits the cell growth and motile activities of 3T3 cells. • LPA{sub 5} suppresses the cell motile activities stimulated by hydrogen peroxide in 3T3 cells. • Enhancement of LPA{sub 5} on the cell motile activities inhibited by LPA{sub 1} in 3T3 cells. • The expression and activation of Mmp-9 were inhibited by LPA{sub 5} in 3T3 cells. • LPA signaling via LPA{sub 5} acts as a negative regulator of cellular responses in 3T3 cells. - Abstract: Lysophosphatidic acid (LPA) signaling via G protein-coupled LPA receptors (LPA{sub 1}–LPA{sub 6}) mediates a variety of biological functions, including cellmore » migration. Recently, we have reported that LPA{sub 1} inhibited the cell motile activities of mouse fibroblast 3T3 cells. In the present study, to evaluate a role of LPA{sub 5} in cellular responses, Lpar5 knockdown (3T3-L5) cells were generated from 3T3 cells. In cell proliferation assays, LPA markedly stimulated the cell proliferation activities of 3T3-L5 cells, compared with control cells. In cell motility assays with Cell Culture Inserts, the cell motile activities of 3T3-L5 cells were significantly higher than those of control cells. The activity levels of matrix metalloproteinases (MMPs) were measured by gelatin zymography. 3T3-L5 cells stimulated the activation of Mmp-2, correlating with the expression levels of Mmp-2 gene. Moreover, to assess the co-effects of LPA{sub 1} and LPA{sub 5} on cell motile activities, Lpar5 knockdown (3T3a1-L5) cells were also established from Lpar1 over-expressing (3T3a1) cells. 3T3a1-L5 cells increased the cell motile activities of 3T3a1 cells, while the cell motile activities of 3T3a1 cells were significantly lower than those of control cells. These results suggest that LPA{sub 5} may act as a negative regulator of cellular responses in mouse fibroblast 3T3 cells, similar to the case for LPA{sub 1}.« less

Analysis of a minimal Rho-GTPase circuit regulating cell shape

NASA Astrophysics Data System (ADS)

Holmes, William R.; Edelstein-Keshet, Leah

2016-08-01

Networks of Rho-family GTPases regulate eukaryotic cell polarization and motility by controlling assembly and contraction of the cytoskeleton. The mutually inhibitory Rac-Rho circuit is emerging as a central, regulatory hub that can affect the shape and motility phenotype of eukaryotic cells. Recent experimental manipulation of the amounts of Rac and Rho or their regulators (guanine nucleotide-exchange factors, GTPase-activating proteins, guanine nucleotide dissociation inhibitors) have been shown to bias the prevalence of these different states and promote transitions between them. Here we show that part of this data can be understood in terms of inherent Rac-Rho mutually inhibitory dynamics. We analyze a spatio-temporal mathematical model of Rac-Rho dynamics to produce a detailed set of predictions of how parameters such as GTPase rates of activation and total amounts affect cell decisions (such as Rho-dominated contraction, Rac-dominated spreading, and spatially segregated Rac-Rho polarization). We find that in some parameter regimes, a cell can take on any of these three fates depending on its environment or stimuli. We also predict how experimental manipulations (corresponding to parameter variations) can affect cell shapes observed. Our methods are based on local perturbation analysis (a kind of nonlinear stability analysis), and an approximation of nonlinear feedback by sharp switches. We compare the Rac-Rho model to an even simpler single-GTPase (‘wave-pinning’) model and demonstrate that the overall behavior is inherent to GTPase properties, rather than stemming solely from network topology.

Lysophosphatidic acid receptor activation affects the C13NJ microglia cell line proteome leading to alterations in glycolysis, motility, and cytoskeletal architecture

PubMed Central

Bernhart, Eva; Kollroser, Manfred; Rechberger, Gerald; Reicher, Helga; Heinemann, Akos; Schratl, Petra; Hallström, Seth; Wintersperger, Andrea; Nusshold, Christoph; DeVaney, Trevor; Zorn-Pauly, Klaus; Malli, Roland; Graier, Wolfgang; Malle, Ernst; Sattler, Wolfgang

2014-01-01

Microglia, the immunocompetent cells of the CNS, are rapidly activated in response to injury and microglia migration towards and homing at damaged tissue plays a key role in CNS regeneration. Lysophosphatidic acid (LPA) is involved in signaling events evoking microglia responses through cognate G protein-coupled receptors. Here we show that human immortalized C13NJ microglia express LPA receptor subtypes LPA1, LPA2, and LPA3 on mRNA and protein level. LPA activation of C13NJ cells induced Rho and extracellular signal-regulated kinase activation and enhanced cellular ATP production. In addition, LPA induced process retraction, cell spreading, led to pronounced changes of the actin cytoskeleton and reduced cell motility, which could be reversed by inhibition of Rho activity. To get an indication about LPA-induced global alterations in protein expression patterns a 2-D DIGE/LC-ESI-MS proteomic approach was applied. On the proteome level the most prominent changes in response to LPA were observed for glycolytic enzymes and proteins regulating cell motility and/or cytoskeletal dynamics. The present findings suggest that naturally occurring LPA is a potent regulator of microglia biology. This might be of particular relevance in the pathophysiological context of neurodegenerative disorders where LPA concentrations can be significantly elevated in the CNS. PMID:19899077

Expression of transmembrane 4 superfamily (TM4SF) proteins and their role in hepatic stellate cell motility and wound healing migration.

PubMed

Mazzocca, Antonio; Carloni, Vinicio; Sciammetta, Silvia; Cordella, Claudia; Pantaleo, Pietro; Caldini, Anna; Gentilini, Paolo; Pinzani, Massimo

2002-09-01

Migration of activated hepatic stellate cells (HSC) is a key event in the progression of liver fibrosis. Little is known about transmembrane proteins involved in HSC motility. Tetraspanins (TM4SF) have been implicated in cell development, differentiation, motility and tumor cell invasion. We evaluated the expression and function of four TM4SF, namely CD9, CD81, CD63 and CD151, and their involvement in HSC migration, adhesion, and proliferation. All TM4SF investigated were highly expressed at the human HSC surface with different patterns of intracellular distribution. Monoclonal antibodies directed against the four TM4SF inhibited HSC migration induced by extracellular matrix proteins in both wound healing and haptotaxis assays. This inhibition was independent of the ECM substrates employed (collagen type I or IV, laminin), and was comparable to that obtained by incubating the cells with an anti-beta1 blocking mAb. Importantly, cell adhesion was unaffected by the incubation with the same antibodies. Co-immunoprecipitation studies revealed different patterns of association between the four TM4SF studied and beta1 integrin. Finally, anti-TM4SF antibodies did not affect HSC growth. These findings provide the first characterization of tetraspanins expression and of their role in HSC migration, a key event in liver tissue wound healing and fibrogenesis.

Characterizing motility dynamics in human RPE cells

NASA Astrophysics Data System (ADS)

Liu, Zhuolin; Kurokawa, Kazuhiro; Zhang, Furu; Miller, Donald T.

2017-02-01

Retinal pigment epithelium (RPE) cells are vital to health of the outer retina, however, are often compromised in ageing and ocular diseases that lead to blindness. Early manifestation of RPE disruption occurs at the cellular level, but while in vivo biomarkers at this scale hold considerable promise, RPE cells have proven extremely challenging to image in the living human eye. Recently we addressed this problem by using organelle motility as a novel contrast agent to enhance the RPE cell in conjunction with 3D resolution of adaptive optics-optical coherence tomography (AO-OCT) to section the RPE layer. In this study, we expand on the central novelty of our method - organelle motility - by characterizing the dynamics of the motility in individual RPE cells, important because of its direct link to RPE physiology. To do this, AO-OCT videos of the same retinal patch were acquired at approximately 1 min intervals or less, time stamped, and registered in 3D with sub-cellular accuracy. Motility was quantified by an exponential decay time constant, the time for motility to decorrelate the speckle field across an RPE cell. In two normal subjects, we found the decay time constant to be just 3 seconds, thus indicating rapid motility in normal RPE cells.

The Azospirillum brasilense Che1 chemotaxis pathway controls swimming velocity, which affects transient cell-to-cell clumping.

PubMed

Bible, Amber; Russell, Matthew H; Alexandre, Gladys

2012-07-01

The Che1 chemotaxis-like pathway of Azospirillum brasilense contributes to chemotaxis and aerotaxis, and it has also been found to contribute to regulating changes in cell surface adhesive properties that affect the propensity of cells to clump and to flocculate. The exact contribution of Che1 to the control of chemotaxis and flocculation in A. brasilense remains poorly understood. Here, we show that Che1 affects reversible cell-to-cell clumping, a cellular behavior in which motile cells transiently interact by adhering to one another at their nonflagellated poles before swimming apart. Clumping precedes and is required for flocculation, and both processes appear to be independently regulated. The phenotypes of a ΔaerC receptor mutant and of mutant strains lacking cheA1, cheY1, cheB1, or cheR1 (alone or in combination) or with che1 deleted show that Che1 directly mediates changes in the flagellar swimming velocity and that this behavior directly modulates the transient nature of clumping. Our results also suggest that an additional receptor(s) and signaling pathway(s) are implicated in mediating other Che1-independent changes in clumping identified in the present study. Transient clumping precedes the transition to stable clump formation, which involves the production of specific extracellular polysaccharides (EPS); however, production of these clumping-specific EPS is not directly controlled by Che1 activity. Che1-dependent clumping may antagonize motility and prevent chemotaxis, thereby maintaining cells in a metabolically favorable niche.

The Azospirillum brasilense Che1 Chemotaxis Pathway Controls Swimming Velocity, Which Affects Transient Cell-to-Cell Clumping

PubMed Central

Bible, Amber; Russell, Matthew H.

2012-01-01

The Che1 chemotaxis-like pathway of Azospirillum brasilense contributes to chemotaxis and aerotaxis, and it has also been found to contribute to regulating changes in cell surface adhesive properties that affect the propensity of cells to clump and to flocculate. The exact contribution of Che1 to the control of chemotaxis and flocculation in A. brasilense remains poorly understood. Here, we show that Che1 affects reversible cell-to-cell clumping, a cellular behavior in which motile cells transiently interact by adhering to one another at their nonflagellated poles before swimming apart. Clumping precedes and is required for flocculation, and both processes appear to be independently regulated. The phenotypes of a ΔaerC receptor mutant and of mutant strains lacking cheA1, cheY1, cheB1, or cheR1 (alone or in combination) or with che1 deleted show that Che1 directly mediates changes in the flagellar swimming velocity and that this behavior directly modulates the transient nature of clumping. Our results also suggest that an additional receptor(s) and signaling pathway(s) are implicated in mediating other Che1-independent changes in clumping identified in the present study. Transient clumping precedes the transition to stable clump formation, which involves the production of specific extracellular polysaccharides (EPS); however, production of these clumping-specific EPS is not directly controlled by Che1 activity. Che1-dependent clumping may antagonize motility and prevent chemotaxis, thereby maintaining cells in a metabolically favorable niche. PMID:22522896

Zebrafish mutations affecting cilia motility share similar cystic phenotypes and suggest a mechanism of cyst formation that differs from pkd2 morphants

PubMed Central

Sullivan-Brown, Jessica; Schottenfeld, Jodi; Okabe, Noriko; Hostetter, Christine L.; Serluca, Fabrizio C.; Thiberge, Stephan Y.; Burdine, Rebecca D.

2008-01-01

Zebrafish are an attractive model for studying the earliest cellular defects occurring during renal cyst formation because its kidney (the pronephros) is simple and genes that cause cystic kidney diseases (CKD) in humans, cause pronephric dilations in zebrafish. By comparing phenotypes in three different mutants, locke, swt and kurly, we find that dilations occur prior to 48 hpf in the medial tubules, a location similar to where cysts form in some mammalian diseases. We demonstrate that the first observeable phenotypes associated with dilation include cilia motility and luminal remodeling defects. Importantly, we show that some phenotypes common to human CKD, such as an increased number of cells, are secondary consequences of dilation. Despite having differences in cilia motility, locke, swt and kurly share similar cystic phenotypes, suggesting that they function in a common pathway. To begin to understand the molecular mechanisms involved in cyst formation, we have cloned the swt mutation and find that it encodes a novel leucine rich repeat containing protein (LRRC50), which is thought to function in correct dynein assembly in cilia. Finally, we show that knockdown of polycystic kidney disease 2 (pkd2) specifically causes glomerular cysts and does not affect cilia motility, suggesting multiple mechanisms exist for cyst formation. PMID:18178183

Toward the reconstitution of synthetic cell motility

PubMed Central

Siton-Mendelson, Orit; Bernheim-Groswasser, Anne

2016-01-01

ABSTRACT Cellular motility is a fundamental process essential for embryonic development, wound healing, immune responses, and tissues development. Cells are mostly moving by crawling on external, or inside, substrates which can differ in their surface composition, geometry, and dimensionality. Cells can adopt different migration phenotypes, e.g., bleb-based and protrusion-based, depending on myosin contractility, surface adhesion, and cell confinement. In the few past decades, research on cell motility has focused on uncovering the major molecular players and their order of events. Despite major progresses, our ability to infer on the collective behavior from the molecular properties remains a major challenge, especially because cell migration integrates numerous chemical and mechanical processes that are coupled via feedbacks that span over large range of time and length scales. For this reason, reconstituted model systems were developed. These systems allow for full control of the molecular constituents and various system parameters, thereby providing insight into their individual roles and functions. In this review we describe the various reconstituted model systems that were developed in the past decades. Because of the multiple steps involved in cell motility and the complexity of the overall process, most of the model systems focus on very specific aspects of the individual steps of cell motility. Here we describe the main advancement in cell motility reconstitution and discuss the main challenges toward the realization of a synthetic motile cell. PMID:27019160

Targeting tumor cell motility to prevent metastasis

PubMed Central

Palmer, Trenis D.; Ashby, William J.; Lewis, John D.; Zijlstra, Andries

2011-01-01

Mortality and morbidity in patients with solid tumors invariably results from the disruption of normal biological function caused by disseminating tumor cells. Tumor cell migration is under intense investigation as the underlying cause of cancer metastasis. The need for tumor cell motility in the progression of metastasis has been established experimentally and is supported empirically by basic and clinical research implicating a large collection of migration-related genes. However, there are few clinical interventions designed to specifically target the motility of tumor cells and adjuvant therapy to specifically prevent cancer cell dissemination is severely limited. In an attempt to define motility targets suitable for treating metastasis, we have parsed the molecular determinants of tumor cell motility into five underlying principles including cell autonomous ability, soluble communication, cell-cell adhesion, cell-matrix adhesion, and integrating these determinants of migration on molecular scaffolds. The current challenge is to implement meaningful and sustainable inhibition of metastasis by developing clinically viable disruption of molecular targets that control these fundamental capabilities. PMID:21664937

Multiciliated Cells in Animals.

PubMed

Meunier, Alice; Azimzadeh, Juliette

2016-12-01

Many animal cells assemble single cilia involved in motile and/or sensory functions. In contrast, multiciliated cells (MCCs) assemble up to 300 motile cilia that beat in a coordinate fashion to generate a directional fluid flow. In the human airways, the brain, and the oviduct, MCCs allow mucus clearance, cerebrospinal fluid circulation, and egg transportation, respectively. Impairment of MCC function leads to chronic respiratory infections and increased risks of hydrocephalus and female infertility. MCC differentiation during development or repair involves the activation of a regulatory cascade triggered by the inhibition of Notch activity in MCC progenitors. The downstream events include the simultaneous assembly of a large number of basal bodies (BBs)-from which cilia are nucleated-in the cytoplasm of the differentiating MCCs, their migration and docking at the plasma membrane associated to an important remodeling of the actin cytoskeleton, and the assembly and polarization of motile cilia. The direction of ciliary beating is coordinated both within cells and at the tissue level by a combination of planar polarity cues affecting BB position and hydrodynamic forces that are both generated and sensed by the cilia. Herein, we review the mechanisms controlling the specification and differentiation of MCCs and BB assembly and organization at the apical surface, as well as ciliary assembly and coordination in MCCs. Copyright © 2016 Cold Spring Harbor Laboratory Press; all rights reserved.

Bacterial spread from cell to cell: beyond actin-based motility.

PubMed

Kuehl, Carole J; Dragoi, Ana-Maria; Talman, Arthur; Agaisse, Hervé

2015-09-01

Several intracellular pathogens display the ability to propagate within host tissues by displaying actin-based motility in the cytosol of infected cells. As motile bacteria reach cell-cell contacts they form plasma membrane protrusions that project into adjacent cells and resolve into vacuoles from which the pathogen escapes, thereby achieving spread from cell to cell. Seminal studies have defined the bacterial and cellular factors that support actin-based motility. By contrast, the mechanisms supporting the formation of protrusions and their resolution into vacuoles have remained elusive. Here, we review recent advances in the field showing that Listeria monocytogenes and Shigella flexneri have evolved pathogen-specific mechanisms of bacterial spread from cell to cell. Copyright © 2015 Elsevier Ltd. All rights reserved.

Influence of Boar and Semen Parameters on Motility and Acrosome Integrity in Liquid Boar Semen Stored for Five Days

PubMed Central

2002-01-01

Ninety ejaculates from a total of 76 AI boars were extended in Beltsville Thawing Solution (BTS). Boar identity, breed, weight of the ejaculate and sperm concentration were registered. Motility and acrosome integrity were assessed after storage at 16–18°C for 6, 30, 54, 78, and 102 h. Storage time had a significant influence on both motility (p < 0.01) and acrosome integrity (p < 0.001). The Least Square Means for percentage of motility showed a small decline from 79.8% after 6 h of storage to 78.4% at 102 h. Motility at 78 and 102 h was significantly different from motility at 6 h (p < 0.05). The percentage of sperm cells with normal acrosomes declined throughout the experiment. The Least Square Means for 6, 30, 54, 78, and 102 h of storage were 93.9%, 90.6%, 88.0%, 84.8%, and 78.2%, respectively. The decrease in acrosome integrity from one storage time to the next was highly significant throughout the trial (p < 0.001). There was a significant influence of boar (p < 0.001) and sperm concentration (p < 0.01) on motility, while acrosome integrity was affected only by boar (p < 0.001). Breed of the boars and weight of the ejaculate did not influence the dependent variables. PMID:12071116

An Emerging Allee Effect Is Critical for Tumor Initiation and Persistence

PubMed Central

Böttger, Katrin; Hatzikirou, Haralambos; Voss-Böhme, Anja; Cavalcanti-Adam, Elisabetta Ada; Herrero, Miguel A.; Deutsch, Andreas

2015-01-01

Tumor cells develop different strategies to cope with changing microenvironmental conditions. A prominent example is the adaptive phenotypic switching between cell migration and proliferation. While it has been shown that the migration-proliferation plasticity influences tumor spread, it remains unclear how this particular phenotypic plasticity affects overall tumor growth, in particular initiation and persistence. To address this problem, we formulate and study a mathematical model of spatio-temporal tumor dynamics which incorporates the microenvironmental influence through a local cell density dependence. Our analysis reveals that two dynamic regimes can be distinguished. If cell motility is allowed to increase with local cell density, any tumor cell population will persist in time, irrespective of its initial size. On the contrary, if cell motility is assumed to decrease with respect to local cell density, any tumor population below a certain size threshold will eventually extinguish, a fact usually termed as Allee effect in ecology. These results suggest that strategies aimed at modulating migration are worth to be explored as alternatives to those mainly focused at keeping tumor proliferation under control. PMID:26335202

In Vitro Motility of Liver Connexin Vesicles along Microtubules Utilizes Kinesin Motors*

PubMed Central

Fort, Alfredo G.; Murray, John W.; Dandachi, Nadine; Davidson, Michael W.; Dermietzel, Rolf; Wolkoff, Allan W.; Spray, David C.

2011-01-01

Trafficking of the proteins that form gap junctions (connexins) from the site of synthesis to the junctional domain appears to require cytoskeletal delivery mechanisms. Although many cell types exhibit specific delivery of connexins to polarized cell sites, such as connexin32 (Cx32) gap junctions specifically localized to basolateral membrane domains of hepatocytes, the precise roles of actin- and tubulin-based systems remain unclear. We have observed fluorescently tagged Cx32 trafficking linearly at speeds averaging 0.25 μm/s in a polarized hepatocyte cell line (WIF-B9), which is abolished by 50 μm of the microtubule-disrupting agent nocodazole. To explore the involvement of cytoskeletal components in the delivery of connexins, we have used a preparation of isolated Cx32-containing vesicles from rat hepatocytes and assayed their ATP-driven motility along stabilized rhodamine-labeled microtubules in vitro. These assays revealed the presence of Cx32 and kinesin motor proteins in the same vesicles. The addition of 50 μm ATP stimulated vesicle motility along linear microtubule tracks with velocities of 0.4–0.5 μm/s, which was inhibited with 1 mm of the kinesin inhibitor AMP-PNP (adenylyl-imidodiphosphate) and by anti-kinesin antibody but only minimally affected by 5 μm vanadate, a dynein inhibitor, or by anti-dynein antibody. These studies provide evidence that Cx32 can be transported intracellularly along microtubules and presumably to junctional domains in cells and highlight an important role of kinesin motor proteins in microtubule-dependent motility of Cx32. PMID:21536677

Soy lecithin interferes with mitochondrial function in frozen-thawed ram spermatozoa.

PubMed

Del Valle, I; Gómez-Durán, A; Holt, W V; Muiño-Blanco, T; Cebrián-Pérez, J A

2012-01-01

Egg yolk and milk are the 2 major membrane cryoprotectants commonly used in freezing media for the long-term preservation of semen (alone or in combination with others). However, in recent years, there have been increasing arguments against the use of egg yolk or milk because of the risk of introducing diseases through the use of cryopreserved semen. In this study, we analyzed the protective effect of lecithin as an alternative to egg yolk for the cryopreservation of ram semen, using a range of functional markers for sperm viability, motility, apoptosis, and mitochondrial functionality analyses (mitochondrial inner membrane surface [MIMS], mitochondrial inner membrane potential [MIMP], and cell membrane potential) as methods of assessment in samples diluted in 3 different media: Tris-citrate-glucose as control and 2 media supplemented with soy lecithin or egg yolk. The results showed that lecithin was able to effectively protect certain sperm quality characteristics against freezing-induced damage. However, lecithin induced loss of mitochondrial membrane potential or mitochondrial loss that was not reflected by modifications in sperm motility in fresh semen. MIMS and MIMP values decreased in thawed lecithin-treated samples, concomitant with a lower (P < .05) percentage of total and progressively motile cells, compared with those in egg yolk-containing samples. Further incubation of thawed samples revealed changes in motility and mitochondrial functionality that otherwise would not have been detected. These results indicated that lecithin may have affected the inner mitochondrial membrane in frozenthawed spermatozoa and confirmed that sublethal damages that seriously affect sperm functionality, not detected by classic sperm quality analyses, can be evidenced by changes in the inner mitochondrial membrane surface. These findings strengthen the relationship between mitochondrial membrane potential and motility and show that the mitochondrial alterations induced by the cryopreservation process could be specific targets for the improvement of semen cryopreservation protocols.

A model of the effects of cancer cell motility and cellular adhesion properties on tumour-immune dynamics.

PubMed

Frascoli, Federico; Flood, Emelie; Kim, Peter S

2017-06-01

We present a three-dimensional model simulating the dynamics of an anti-cancer T-cell response against a small, avascular, early-stage tumour. Interactions at the tumour site are accounted for using an agent-based model (ABM), while immune cell dynamics in the lymph node are modelled as a system of delay differential equations (DDEs). We combine these separate approaches into a two-compartment hybrid ABM-DDE system to capture the T-cell response against the tumour. In the ABM at the tumour site, movement of tumour cells is modelled using effective physical forces with a specific focus on cell-to-cell adhesion properties and varying levels of tumour cell motility, thus taking into account the ability of cancer cells to spread and form clusters. We consider the effectiveness of the immune response over a range of parameters pertaining to tumour cell motility, cell-to-cell adhesion strength and growth rate. We also investigate the dependence of outcomes on the distribution of tumour cells. Low tumour cell motility is generally a good indicator for successful tumour eradication before relapse, while high motility leads, almost invariably, to relapse and tumour escape. In general, the effect of cell-to-cell adhesion on prognosis is dependent on the level of tumour cell motility, with an often unpredictable cross influence between adhesion and motility, which can lead to counterintuitive effects. In terms of overall tumour shape and structure, the spatial distribution of cancer cells in clusters of various sizes has shown to be strongly related to the likelihood of extinction. © The authors 2016. Published by Oxford University Press on behalf of the Institute of Mathematics and its Applications. All rights reserved.

Effects of reactive oxygen species on sperm function.

PubMed

Guthrie, H D; Welch, G R

2012-11-01

Reactive oxygen species (ROS) formation and membrane lipid peroxidation have been recognized as problems for sperm survival and fertility. The precise roles and detection of superoxide (SO), hydrogen peroxide (HP), and membrane lipid peroxidation have been problematic, because of the low specificity and sensitivity of the established chemiluminescence assay technologies. We developed flow cytometric assays to measure SO, HP, membrane lipid peroxidation, and inner mitochondrial transmembrane potential in boar sperm. These methods were sufficiently sensitive to permit detection of early changes in ROS formation in sperm cells that were still viable. Basal ROS formation and membrane lipid peroxidation in the absence of ROS generators were low in viable sperm of both fresh and frozen-thawed boar semen, affecting less than 4% of the sperm cells on average. However, this is not the case in other species, as human, bovine, and poultry sperm have large increases in sperm ROS formation, lipid peroxidation, loss of motility, and death in vitro. Closer study of the effects of ROS formation on the relationship between sperm motility and ATP content in boar sperm was conducted using menadione (mitochondrial SO generator) and HP treatment. Menadione or HP caused an immediate disruption of motility with delayed or no decrease in sperm ATP content, respectively. Overall, the inhibitory effects of ROS on motility point to a mitochondrial-independent mechanism. The reduction in motility may have been due to a ROS-induced lesion in ATP utilization or in the contractile apparatus of the flagellum. Published by Elsevier Inc.

RON kinase isoforms demonstrate variable cell motility in normal cells.

PubMed

Greenbaum, Alissa; Rajput, Ashwani; Wan, Guanghua

2016-09-01

Aberrant RON (Recepteur d'Origine Nantais) tyrosine kinase activation causes the epithelial cell to evade normal growth pathways, resulting in unregulated cell proliferation, increased cell motility and decreased apoptosis. Wildtype (wt) RON has been shown to play a role in metastasis of epithelial malignancies. It presents an important potential therapeutic target for colorectal, breast, gastric and pancreatic cancer. Little is known about functional differences amongst RON isoforms RON155, RON160 and RON165. The purpose of this study was to determine the effect of various RON kinase isoforms on cell motility. Cell lines with stable expression of wtRON were generated by inserting the coding region of RON in pTagRFP (tagged red fluorescence protein plasmid). The expression constructs of RON variants (RON155, RON160 and RON165) were generated by creating a mutagenesis-based wtRON-pTag RFP plasmid and stably transfected into HEK 293 cells. The wound closure scratch assay was used to investigate the effect on cell migratory capacity of wild type RON and its variants. RON transfected cells demonstrated increased cell motility compared to HEK293 control cells. RON165 cell motility was significantly increased compared to RON160 (mean percentage of wound covered 37.37% vs. 32.40%; p = 0.03). RON tyrosine kinase isoforms have variable cell motility. This may reflect a difference in the behavior of malignant epithelial cells and their capacity for metastasis.

Glycolysis is the primary bioenergetic pathway for cell motility and cytoskeletal remodeling in human prostate and breast cancer cells.

PubMed

Shiraishi, Takumi; Verdone, James E; Huang, Jessie; Kahlert, Ulf D; Hernandez, James R; Torga, Gonzalo; Zarif, Jelani C; Epstein, Tamir; Gatenby, Robert; McCartney, Annemarie; Elisseeff, Jennifer H; Mooney, Steven M; An, Steven S; Pienta, Kenneth J

2015-01-01

The ability of a cancer cell to detach from the primary tumor and move to distant sites is fundamental to a lethal cancer phenotype. Metabolic transformations are associated with highly motile aggressive cellular phenotypes in tumor progression. Here, we report that cancer cell motility requires increased utilization of the glycolytic pathway. Mesenchymal cancer cells exhibited higher aerobic glycolysis compared to epithelial cancer cells while no significant change was observed in mitochondrial ATP production rate. Higher glycolysis was associated with increased rates of cytoskeletal remodeling, greater cell traction forces and faster cell migration, all of which were blocked by inhibition of glycolysis, but not by inhibition of mitochondrial ATP synthesis. Thus, our results demonstrate that cancer cell motility and cytoskeleton rearrangement is energetically dependent on aerobic glycolysis and not oxidative phosphorylation. Mitochondrial derived ATP is insufficient to compensate for inhibition of the glycolytic pathway with regard to cellular motility and CSK rearrangement, implying that localization of ATP derived from glycolytic enzymes near sites of active CSK rearrangement is more important for cell motility than total cellular ATP production rate. These results extend our understanding of cancer cell metabolism, potentially providing a target metabolic pathway associated with aggressive disease.

Glycolysis is the primary bioenergetic pathway for cell motility and cytoskeletal remodeling in human prostate and breast cancer cells

PubMed Central

Shiraishi, Takumi; Verdone, James E.; Huang, Jessie; Kahlert, Ulf D.; Hernandez, James R.; Torga, Gonzalo; Zarif, Jelani C.; Epstein, Tamir; Gatenby, Robert; McCartney, Annemarie; Elisseeff, Jennifer H.; Mooney, Steven M.; An, Steven S.; Pienta, Kenneth J.

2015-01-01

The ability of a cancer cell to detach from the primary tumor and move to distant sites is fundamental to a lethal cancer phenotype. Metabolic transformations are associated with highly motile aggressive cellular phenotypes in tumor progression. Here, we report that cancer cell motility requires increased utilization of the glycolytic pathway. Mesenchymal cancer cells exhibited higher aerobic glycolysis compared to epithelial cancer cells while no significant change was observed in mitochondrial ATP production rate. Higher glycolysis was associated with increased rates of cytoskeletal remodeling, greater cell traction forces and faster cell migration, all of which were blocked by inhibition of glycolysis, but not by inhibition of mitochondrial ATP synthesis. Thus, our results demonstrate that cancer cell motility and cytoskeleton rearrangement is energetically dependent on aerobic glycolysis and not oxidative phosphorylation. Mitochondrial derived ATP is insufficient to compensate for inhibition of the glycolytic pathway with regard to cellular motility and CSK rearrangement, implying that localization of ATP derived from glycolytic enzymes near sites of active CSK rearrangement is more important for cell motility than total cellular ATP production rate. These results extend our understanding of cancer cell metabolism, potentially providing a target metabolic pathway associated with aggressive disease. PMID:25426557

Effects of minimal exposures to atmospheric pressure plasma on the activity of Salmonella Typhimurium: Deactivation of bacterial motility and suppression of host-cell invasion.

PubMed

Park, Jin-Sung; Kim, Kijung; Han, Je-Hyun; Gweon, Bomi; Ko, Ung Hyun; Yoo, Suk Jae; Choe, Wonho; Shin, Jennifer H

2016-09-01

Atmospheric pressure plasma (APP) has been shown effective in sterilization by reducing the number of viable microbes during surface cleaning, food processing, or human tissue treatment. For safe conduct, the majority of previous research focused on complete abolition of microbes, which may require severe treatments. Our aim is to investigate the minimal treatment conditions necessary for effective inactivation of bacteria in such a manner that the APP treated bacteria would not be able to harm the host cells. For this, we ought to identify the objective criteria to make the bacteria dysfunctional. We choose the motile properties and the host-cell invasion capability as two measures to quantify the pathogenic state of bacteria. In this paper, we investigated how the APP treatment in a minimal dosage affects the activity of Salmonella Typhimurium. At 100 W and 15 kHz for 20 s, the APP treatment effectively suppressed active "run and tumble" type motility and induced formation of abnormally long structures. With 20 s exposure, the bacterial cells failed to cause pyroptosis in the host cells with >90% survival after 12 h of co-incubation. Our results suggest novel measures to evaluate the functional pathogenic state for identifying safe APP treatment conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

Inhibition of cell proliferation and migration by oxidative stress from ascorbate-driven juglone redox cycling in human bladder-derived T24 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kviecinski, M.R., E-mail: mrkviecinski@hotmail.com; Pedrosa, R.C., E-mail: rozangelapedrosa@gmail.com; Felipe, K.B., E-mail: kakabettega@yahoo.com.br

2012-05-04

Highlights: Black-Right-Pointing-Pointer The cytotoxicity of juglone is markedly increased by ascorbate. Black-Right-Pointing-Pointer T24 cell death by oxidative stress is necrosis-like. Black-Right-Pointing-Pointer Redox cycling by juglone/ascorbate inhibits cell proliferation. Black-Right-Pointing-Pointer Cellular migration is impaired by juglone/ascorbate. -- Abstract: The effects of juglone on T24 cells were assessed in the presence and absence of ascorbate. The EC{sub 50} value for juglone at 24 h decreased from 28.5 {mu}M to 6.3 {mu}M in the presence of ascorbate. In juglone-treated cells, ascorbate increased ROS formation (4-fold) and depleted GSH (65%). N-acetylcysteine or catalase restricted the juglone/ascorbate-mediated effects, highlighting the role of oxidative stress inmore » juglone cytotoxicity. Juglone alone or associated with ascorbate did not cause caspase-3 activation or PARP cleavage, suggesting necrosis-like cell death. DNA damage and the mild ER stress caused by juglone were both enhanced by ascorbate. In cells treated with juglone (1-5 {mu}M), a concentration-dependent decrease in cell proliferation was observed. Ascorbate did not impair cell proliferation but its association with juglone led to a clonogenic death state. The motility of ascorbate-treated cells was not affected. Juglone slightly restricted motility, but cells lost their ability to migrate most noticeably when treated with juglone plus ascorbate. We postulate that juglone kills cells by a necrosis-like mechanism inhibiting cell proliferation and the motility of T24 cells. These effects are enhanced in the presence of ascorbate.« less

Angiopoietin-like 4 prevents metastasis through inhibition of vascular permeability and tumor cell motility and invasiveness.

PubMed

Galaup, Ariane; Cazes, Aurelie; Le Jan, Sebastien; Philippe, Josette; Connault, Elisabeth; Le Coz, Emmanuelle; Mekid, Halima; Mir, Lluis M; Opolon, Paule; Corvol, Pierre; Monnot, Catherine; Germain, Stephane

2006-12-05

Angiopoietin-like 4 (ANGPTL4), a secreted protein of the angiopoietin-like family, is induced by hypoxia in both tumor and endothelial cells as well as in hypoxic perinecrotic areas of numerous cancers. Here, we investigated whether ANGPTL4 might affect tumor growth as well as metastasis. Metastatic 3LL cells were therefore xenografted into control mice and mice in which ANGPTL4 was expressed by using in vivo DNA electrotransfer. Whereas primary tumors grew at a similar rate in both groups, 3LL cells metastasized less efficiently to the lungs of mice that expressed ANGPTL4. Fewer 3LL emboli were observed in primary tumors, suggesting that intravasation of 3LL cells was inhibited by ANGPTL4. Furthermore, melanoma B16F0 cells injected into the retro-orbital sinus also metastasized less efficiently in mice expressing ANGPTL4. Although B16F0 cells were observed in lung vessels, they rarely invaded the parenchyma, suggesting that ANGPTL4 affects extravasation. In addition, recombinant B16F0 cells that overexpress ANGPTL4 were generated, showing a lower capacity for in vitro migration, invasion, and adhesion than control cells. Expression of ANGPTL4 induced reorganization of the actin cytoskeleton through inhibition of actin stress fiber formation and vinculin localization at focal contacts. Together, these results show that ANGPTL4, through its action on both vascular and tumor compartments, prevents the metastatic process by inhibiting vascular activity as well as tumor cell motility and invasiveness.

Angiopoietin-like 4 prevents metastasis through inhibition of vascular permeability and tumor cell motility and invasiveness

PubMed Central

Galaup, Ariane; Cazes, Aurelie; Le Jan, Sebastien; Philippe, Josette; Connault, Elisabeth; Le Coz, Emmanuelle; Mekid, Halima; Mir, Lluis M.; Opolon, Paule; Corvol, Pierre; Monnot, Catherine; Germain, Stephane

2006-01-01

Angiopoietin-like 4 (ANGPTL4), a secreted protein of the angiopoietin-like family, is induced by hypoxia in both tumor and endothelial cells as well as in hypoxic perinecrotic areas of numerous cancers. Here, we investigated whether ANGPTL4 might affect tumor growth as well as metastasis. Metastatic 3LL cells were therefore xenografted into control mice and mice in which ANGPTL4 was expressed by using in vivo DNA electrotransfer. Whereas primary tumors grew at a similar rate in both groups, 3LL cells metastasized less efficiently to the lungs of mice that expressed ANGPTL4. Fewer 3LL emboli were observed in primary tumors, suggesting that intravasation of 3LL cells was inhibited by ANGPTL4. Furthermore, melanoma B16F0 cells injected into the retro-orbital sinus also metastasized less efficiently in mice expressing ANGPTL4. Although B16F0 cells were observed in lung vessels, they rarely invaded the parenchyma, suggesting that ANGPTL4 affects extravasation. In addition, recombinant B16F0 cells that overexpress ANGPTL4 were generated, showing a lower capacity for in vitro migration, invasion, and adhesion than control cells. Expression of ANGPTL4 induced reorganization of the actin cytoskeleton through inhibition of actin stress fiber formation and vinculin localization at focal contacts. Together, these results show that ANGPTL4, through its action on both vascular and tumor compartments, prevents the metastatic process by inhibiting vascular activity as well as tumor cell motility and invasiveness. PMID:17130448

Differentiation Affects the Release of Exosomes from Colon Cancer Cells and Their Ability to Modulate the Behavior of Recipient Cells.

PubMed

Lucchetti, Donatella; Calapà, Federica; Palmieri, Valentina; Fanali, Caterina; Carbone, Federica; Papa, Alfredo; De Maria, Ruggero; De Spirito, Marco; Sgambato, Alessandro

2017-07-01

Exosomes are involved in intercellular communication. We previously reported that sodium butyrate-induced differentiation of HT29 colon cancer cells is associated with a reduced CD133 expression. Herein, we analyzed the role of exosomes in the differentiation of HT29 cells. Exosomes were prepared using ultracentrifugation. Gene expression levels were evaluated by real-time PCR. The cell proliferation rate was assessed by MTT assay and with the electric cell-substrate impedance sensing system, whereas cell motility was assessed using the scratch test and confocal microscopy. Sodium butyrate-induced differentiation of HT29 and Caco-2 cells increased the levels of released exosomes and their expression of CD133. Cell differentiation and the decrease of cellular CD133 expression levels were prevented by blocking multivesicular body maturation. Exosomes released by HT29 differentiating cells carried increased levels of miRNAs, induced an increased proliferation and motility of both colon cancer cells and normal fibroblasts, increased the colony-forming efficiency of cancer cells, and reduced the sodium butyrate-induced differentiation of HT29 cells. Such effects were associated with an increased phosphorylation level of both Src and extracellular signal regulated kinase proteins and with an increased expression of epithelial-to-mesenchymal transition-related genes. Release of exosomes is affected by differentiation of colon cancer cells; exosomes might be used by differentiating cells to get rid of components that are no longer necessary but might continue to exert their effects on recipient cells. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Motility modes of the parasite Trypanosoma brucei

NASA Astrophysics Data System (ADS)

Temel, Fatma Zeynep; Qu, Zijie; McAllaster, Michael; de Graffenried, Christopher; Breuer, Kenneth

2015-11-01

The parasitic single-celled protozoan Trypanosoma brucei causes African Sleeping Sickness, which is a fatal disease in humans and animals that threatens more than 60 million people in 36 African countries. Cell motility plays a critical role in the developmental phases and dissemination of the parasite. Unlike many other motile cells such as bacteria Escherichia coli or Caulobacter crescentus, the flagellum of T. brucei is attached along the length of its awl-like body, producing a unique mode of motility that is not fully understood or characterized. Here, we report on the motility of T. brucei, which swims using its single flagellum employing both rotating and undulating propulsion modes. We tracked cells in real-time in three dimensions using fluorescent microscopy. Data obtained from experiments using both short-term tracking within the field of view and long-term tracking using a tracking microscope were analyzed. Motility modes and swimming speed were analyzed as functions of cell size, rotation rate and undulation pattern. Research supported by NSF.

Mechanical stress as a regulator of cell motility

NASA Astrophysics Data System (ADS)

Putelat, T.; Recho, P.; Truskinovsky, L.

2018-01-01

The motility of a cell can be triggered or inhibited not only by an applied force but also by a mechanically neutral force couple. This type of loading, represented by an applied stress and commonly interpreted as either squeezing or stretching, can originate from extrinsic interaction of a cell with its neighbors. To quantify the effect of applied stresses on cell motility we use an analytically transparent one-dimensional model accounting for active myosin contraction and induced actin turnover. We show that stretching can polarize static cells and initiate cell motility while squeezing can symmetrize and arrest moving cells. We show further that sufficiently strong squeezing can lead to the loss of cell integrity. The overall behavior of the system depends on the two dimensionless parameters characterizing internal driving (chemical activity) and external loading (applied stress). We construct a phase diagram in this parameter space distinguishing between static, motile, and collapsed states. The obtained results are relevant for the mechanical understanding of contact inhibition and the epithelial-to-mesenchymal transition.

Image velocimetry and spectral analysis enable quantitative characterization of larval zebrafish gut motility.

PubMed

Ganz, J; Baker, R P; Hamilton, M K; Melancon, E; Diba, P; Eisen, J S; Parthasarathy, R

2018-05-02

Normal gut function requires rhythmic and coordinated movements that are affected by developmental processes, physical and chemical stimuli, and many debilitating diseases. The imaging and characterization of gut motility, especially regarding periodic, propagative contractions driving material transport, are therefore critical goals. Previous image analysis approaches have successfully extracted properties related to the temporal frequency of motility modes, but robust measures of contraction magnitude, especially from in vivo image data, remain challenging to obtain. We developed a new image analysis method based on image velocimetry and spectral analysis that reveals temporal characteristics such as frequency and wave propagation speed, while also providing quantitative measures of the amplitude of gut motion. We validate this approach using several challenges to larval zebrafish, imaged with differential interference contrast microscopy. Both acetylcholine exposure and feeding increase frequency and amplitude of motility. Larvae lacking enteric nervous system gut innervation show the same average motility frequency, but reduced and less variable amplitude compared to wild types. Our image analysis approach enables insights into gut dynamics in a wide variety of developmental and physiological contexts and can also be extended to analyze other types of cell movements. © 2018 John Wiley & Sons Ltd.

A Cell Number Counting Factor Regulates Akt/Protein Kinase B To Regulate Dictyostelium discoideum Group Size

PubMed Central

Gao, Tong; Knecht, David; Tang, Lei; Hatton, R. Diane; Gomer, Richard H.

2004-01-01

Little is known about how individual cells can organize themselves to form structures of a given size. During development, Dictyostelium discoideum aggregates in dendritic streams and forms groups of ∼20,000 cells. D. discoideum regulates group size by secreting and simultaneously sensing a multiprotein complex called counting factor (CF). If there are too many cells in a stream, the associated high concentration of CF will decrease cell-cell adhesion and increase cell motility, causing aggregation streams to break up. The pulses of cyclic AMP (cAMP) that mediate aggregation cause a transient translocation of Akt/protein kinase B (Akt/PKB) to the leading edge of the plasma membrane and a concomitant activation of the kinase activity, which in turn stimulates motility. We found that countin− cells (which lack bioactive CF) and wild-type cells starved in the presence of anticountin antibodies (which block CF activity) showed a decreased level of cAMP-stimulated Akt/PKB membrane translocation and kinase activity compared to parental wild-type cells. Recombinant countin has the bioactivity of CF, and a 1-min treatment of cells with recombinant countin potentiated Akt/PKB translocation to membranes and Akt/PKB activity. Western blotting of total cell lysates indicated that countin does not affect the total level of Akt/PKB. Fluorescence microscopy of cells expressing an Akt/PKB pleckstrin homology domain-green fluorescent protein (PH-GFP) fusion protein indicated that recombinant countin and anti-countin antibodies do not obviously alter the distribution of Akt/PKB PH-GFP when it translocates to the membrane. Our data indicate that CF increases motility by potentiating the cAMP-stimulated activation and translocation of Akt/PKB. PMID:15470246

Enterolactone alters FAK-Src signaling and suppresses migration and invasion of lung cancer cell lines.

PubMed

Chikara, Shireen; Lindsey, Kaitlin; Borowicz, Pawel; Christofidou-Solomidou, Melpo; Reindl, Katie M

2017-01-09

Systemic toxicity of chemotherapeutic agents and the challenges associated with targeting metastatic tumors are limiting factors for current lung cancer therapeutic approaches. To address these issues, plant-derived bioactive components have been investigated for their anti-cancer properties because many of these agents are non-toxic to healthy tissues. Enterolactone (EL) is a flaxseed-derived mammalian lignan that has demonstrated anti-migratory properties for various cancers, but EL has not been investigated in the context of lung cancer, and its anticancer mechanisms are ill-defined. We hypothesized that EL could inhibit lung cancer cell motility by affecting the FAK-Src signaling pathway. Non-toxic concentrations of EL were identified for A549 and H460 human lung cancer cells by conducting 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-Dephenyltetrazolium Bromide (MTT) assays. The anti-migratory and anti-invasive potential of EL for lung cancer cell lines was determined by scratch wound healing and Matrigel® invasion assays. Changes in filamentous actin (F-actin) fiber density and length in EL-treated cells were determined using phalloidin-conjugated rhodamine dye and fluorescent microscopy. Vinculin expression in focal adhesions upon EL treatment was determined by immunocytochemistry. Gene and protein expression levels of FAK-Src signaling molecules in EL-treated lung cancer cells were determined using PCR arrays, qRT-PCR, and western blotting. Non-toxic concentrations of EL inhibited lung cancer cell migration and invasion in a concentration- and time-dependent manner. EL treatment reduced the density and number of F-actin fibers in lung cancer cell lines, and reduced the number and size of focal adhesions. EL decreased phosphorylation of FAK and its downstream targets, Src, paxillin, and decreased mRNA expression of cell motility-related genes, RhoA, Rac1, and Cdc42 in lung cancer cells. Our data suggest that EL suppresses lung cancer cell motility and invasion by altering FAK activity and subsequent activation of downstream proteins needed for focal adhesion formation and cytoskeletal rearrangement. Therefore, administration of EL may serve as a safe and complementary approach for inhibiting lung tumor cell motility, invasion, and metastasis.

Retinoic acid-induced pancreatic stellate cell quiescence reduces paracrine Wnt-β-catenin signaling to slow tumor progression.

PubMed

Froeling, Fieke E M; Feig, Christine; Chelala, Claude; Dobson, Richard; Mein, Charles E; Tuveson, David A; Clevers, Hans; Hart, Ian R; Kocher, Hemant M

2011-10-01

Patients with pancreatic ductal adenocarcinoma are deficient in vitamin A, resulting in activation of pancreatic stellate cells (PSCs). We investigated whether restoration of retinol to PSCs restores their quiescence and affects adjacent cancer cells. PSCs and cancer cell lines (AsPc1 and Capan1) were exposed to doses and isoforms of retinoic acid (RA) in 2-dimensional and 3-dimensional culture conditions (physiomimetic organotypic culture). The effects of all-trans retinoic acid (ATRA) were studied in LSL-KrasG12D/+;LSL-Trp53R172H/+;Pdx-1-Cre mice, a model of human pancreatic ductal adenocarcinoma. After incubation with ATRA, PSCs were quiescent and had altered expression of genes that regulate proliferation, morphology, and motility; genes that encode cytoskeletal proteins and cytokines; and genes that control other functions, irrespective of culture conditions or dosage. In the organotypic model, and in mice, ATRA induced quiescence of PSCs and thereby reduced cancer cell proliferation and translocation of β-catenin to the nucleus, increased cancer cell apoptosis, and altered tumor morphology. ATRA reduced the motility of PSCs, so these cells created a "wall" at the junction between the tumor and the matrix that prevented cancer cell invasion. Restoring secreted frizzled-related protein 4 (sFRP4) secretion to quiescent PSCs reduced Wnt-β-catenin signaling in cancer cells and their invasive ability. Human primary and metastatic pancreatic tumor tissues stained strongly for cancer cell nuclear β-catenin but had low levels of sFRP4 (in cancer cells and PSCs). RA induces quiescence and reduces motility of PSCs, leading to reduced proliferation and increased apoptosis of surrounding pancreatic cancer cells. RA isoforms might be developed as therapeutic reagents for pancreatic cancer. Copyright © 2011 AGA Institute. Published by Elsevier Inc. All rights reserved.

Regulation of human genome expression and RNA splicing by human papillomavirus 16 E2 protein.

PubMed

Gauson, Elaine J; Windle, Brad; Donaldson, Mary M; Caffarel, Maria M; Dornan, Edward S; Coleman, Nicholas; Herzyk, Pawel; Henderson, Scott C; Wang, Xu; Morgan, Iain M

2014-11-01

Human papillomavirus 16 (HPV16) is causative in human cancer. The E2 protein regulates transcription from and replication of the viral genome; the role of E2 in regulating the host genome has been less well studied. We have expressed HPV16 E2 (E2) stably in U2OS cells; these cells tolerate E2 expression well and gene expression analysis identified 74 genes showing differential expression specific to E2. Analysis of published gene expression data sets during cervical cancer progression identified 20 of the genes as being altered in a similar direction as the E2 specific genes. In addition, E2 altered the splicing of many genes implicated in cancer and cell motility. The E2 expressing cells showed no alteration in cell growth but were altered in cell motility, consistent with the E2 induced altered splicing predicted to affect this cellular function. The results present a model system for investigating E2 regulation of the host genome. Copyright © 2014 Elsevier Inc. All rights reserved.

Bioengineered intestinal muscularis complexes with long-term spontaneous and periodic contractions

PubMed Central

Wang, Qianqian; Wang, Ke; Solorzano-Vargas, R. Sergio; Lin, Po-Yu; Walthers, Christopher M.; Thomas, Anne-Laure; Martín, Martín G.

2018-01-01

Although critical for studies of gut motility and intestinal regeneration, the in vitro culture of intestinal muscularis with peristaltic function remains a significant challenge. Periodic contractions of intestinal muscularis result from the coordinated activity of smooth muscle cells (SMC), the enteric nervous system (ENS), and interstitial cells of Cajal (ICC). Reproducing this activity requires the preservation of all these cells in one system. Here we report the first serum-free culture methodology that consistently maintains spontaneous and periodic contractions of murine and human intestinal muscularis cells for months. In this system, SMC expressed the mature marker myosin heavy chain, and multipolar/dipolar ICC, uniaxonal/multipolar neurons and glial cells were present. Furthermore, drugs affecting neural signals, ICC or SMC altered the contractions. Combining this method with scaffolds, contracting cell sheets were formed with organized architecture. With the addition of intestinal epithelial cells, this platform enabled up to 11 types of cells from mucosa, muscularis and serosa to coexist and epithelial cells were stretched by the contracting muscularis cells. The method constitutes a powerful tool for mechanistic studies of gut motility disorders and the functional regeneration of the engineered intestine. PMID:29718926

Defective phagosome motility and degradation in cell nonautonomous RPE pathogenesis of a dominant macular degeneration.

PubMed

Esteve-Rudd, Julian; Hazim, Roni A; Diemer, Tanja; Paniagua, Antonio E; Volland, Stefanie; Umapathy, Ankita; Williams, David S

2018-05-22

Stargardt macular dystrophy 3 (STGD3) is caused by dominant mutations in the ELOVL4 gene. Like other macular degenerations, pathogenesis within the retinal pigment epithelium (RPE) appears to contribute to the loss of photoreceptors from the central retina. However, the RPE does not express ELOVL4 , suggesting photoreceptor cell loss in STGD3 occurs through two cell nonautonomous events: mutant photoreceptors first affect RPE cell pathogenesis, and then, second, RPE dysfunction leads to photoreceptor cell death. Here, we have investigated how the RPE pathology occurs, using a STGD3 mouse model in which mutant human ELOVL4 is expressed in the photoreceptors. We found that the mutant protein was aberrantly localized to the photoreceptor outer segment (POS), and that resulting POS phagosomes were degraded more slowly in the RPE. In cell culture, the mutant POSs are ingested by primary RPE cells normally, but the phagosomes are processed inefficiently, even by wild-type RPE. The mutant phagosomes excessively sequester RAB7A and dynein, and have impaired motility. We propose that the abnormal presence of ELOVL4 protein in POSs results in phagosomes that are defective in recruiting appropriate motor protein linkers, thus contributing to slower degradation because their altered motility results in slower basal migration and fewer productive encounters with endolysosomes. In the transgenic mouse retinas, the RPE accumulated abnormal-looking phagosomes and oxidative stress adducts; these pathological changes were followed by pathology in the neural retina. Our results indicate inefficient phagosome degradation as a key component of the first cell nonautonomous event underlying retinal degeneration due to mutant ELOVL4.

Optimization of the sperm:oocyte ratio and sperm economy in the artificial reproduction of Rhamdia quelen using fructose as a sperm motility modulator.

PubMed

Adames, Maurício Spagnolo; de Toledo, Cesar Pereira Rebechi; Neumann, Giovano; Buzzi, Alexandre Henrique; Buratto, Cíntia Nara; Piana, Pitágoras Augusto; Bombardelli, Robie Allan

2015-10-01

This research was conducted to evaluate the effects of fructose as a modulator of sperm motility and its effects on the reduction in number of sperm cells in IVF using cryopreserved Rhamdia quelen semen. Sperm activation occurred in solutions containing fructose (0.0, 0.9, 1.8, 2.7, 3.6 and 4.5%). The sperm motility rate, velocity and duration of sperm motility were assessed by polynomial regression analysis and grouped by the principal component analysis (PCA). Then, the oocytes were mixed with semen at proportions of 1×10(4), 3×10(4), 5×10(4), 7×10(4) and 9×10(4) for the sperm:oocyte ratio and fertilization was induced by the activation of gametes with the fructose-containing solutions. The fertilization, hatching and larval normality rate were evaluated by response surface protocol and were further grouped by PCA. All sperm variables were affected by the activating solutions, and the most desirable theoretical results for the rate of sperm motility were obtained when using a solution containing 2.85% fructose. In the IVF and incubation assays, there was an interactive effect between the motile sperm:oocyte ratio and the fructose concentration on the rates of oocyte fertilization, hatching and on the clustered index for reproductive success. The results suggest the possibility of reducing the sperm cells on IVF by 17.77% when using a solution containing 2.28% fructose. In conclusion, the use of solutions containing fructose at concentrations that maximize sperm movement allow the reduction of the motile sperm:oocyte ratio, thus promoting sperm metabolic efficiencies and contributing to the feasibility of using cryopreserved semen at a large-scale in IVF. Copyright © 2015 Elsevier B.V. All rights reserved.

The CsgA and Lpp proteins affect HEp-2 cell invasion, motility, and biofilm formation in a strain of Escherichia coli O157:H7

USDA-ARS?s Scientific Manuscript database

In Escherichia coli O157:H7 strain ATCC 43895, a guanine to thymine transversion in the csgD promoter created strain 43895OR. Strain 43895OR produces an abundant extracellular matrix rich in curli fibers, forms biofilm on solid surfaces, invades cultured epithelial cells, and is more virulent in mic...

Microfabricated systems and assays for studying the cytoskeletal organization, micromechanics, and motility patterns of cancerous cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huda, Sabil; Pilans, Didzis; Makurath, Monika

Cell motions are driven by coordinated actions of the intracellular cytoskeleton – actin, microtubules (MTs) and substrate/focal adhesions (FAs). This coordination is altered in metastatic cancer cells resulting in deregulated and increased cellular motility. Microfabrication tools, including photolithography, micromolding, microcontact printing, wet stamping and microfluidic devices have emerged as a powerful set of experimental tools with which to probe and define the differences in cytoskeleton organization/dynamics and cell motility patterns in non-metastatic and metastatic cancer cells. In this paper, we discuss four categories of microfabricated systems: (i) micropatterned substrates for studying of cell motility sub-processes (for example, MT targeting ofmore » FAs or cell polarization); (ii) systems for studying cell mechanical properties, (iii) systems for probing overall cell motility patterns within challenging geometric confines relevant to metastasis (for example, linear and ratchet geometries), and (iv) microfluidic devices that incorporate co-cultures of multiple cell types and chemical gradients to mimic in vivo intravasation/extravasation steps of metastasis. Finally, together, these systems allow for creating controlled microenvironments that not only mimic complex soft tissues, but are also compatible with live cell high-resolution imaging and quantitative analysis of single cell behavior.« less

Microfabricated systems and assays for studying the cytoskeletal organization, micromechanics, and motility patterns of cancerous cells

DOE PAGES

Huda, Sabil; Pilans, Didzis; Makurath, Monika; ...

2014-08-28

Cell motions are driven by coordinated actions of the intracellular cytoskeleton – actin, microtubules (MTs) and substrate/focal adhesions (FAs). This coordination is altered in metastatic cancer cells resulting in deregulated and increased cellular motility. Microfabrication tools, including photolithography, micromolding, microcontact printing, wet stamping and microfluidic devices have emerged as a powerful set of experimental tools with which to probe and define the differences in cytoskeleton organization/dynamics and cell motility patterns in non-metastatic and metastatic cancer cells. In this paper, we discuss four categories of microfabricated systems: (i) micropatterned substrates for studying of cell motility sub-processes (for example, MT targeting ofmore » FAs or cell polarization); (ii) systems for studying cell mechanical properties, (iii) systems for probing overall cell motility patterns within challenging geometric confines relevant to metastasis (for example, linear and ratchet geometries), and (iv) microfluidic devices that incorporate co-cultures of multiple cell types and chemical gradients to mimic in vivo intravasation/extravasation steps of metastasis. Finally, together, these systems allow for creating controlled microenvironments that not only mimic complex soft tissues, but are also compatible with live cell high-resolution imaging and quantitative analysis of single cell behavior.« less

In vitro motility evaluation of aggregated cancer cells by means of automatic image processing.

PubMed

De Hauwer, C; Darro, F; Camby, I; Kiss, R; Van Ham, P; Decaesteker, C

1999-05-01

Set up of an automatic image processing based method that enables the motility of in vitro aggregated cells to be evaluated for a number of hours. Our biological model included the PC-3 human prostate cancer cell line growing as a monolayer on the bottom of Falcon plastic dishes containing conventional culture media. Our equipment consisted of an incubator, an inverted phase contrast microscope, a Charge Coupled Device (CCD) video camera, and a computer equipped with an image processing software developed in our laboratory. This computer-assisted microscope analysis of aggregated cells enables global cluster motility to be evaluated. This analysis also enables the trajectory of each cell to be isolated and parametrized within a given cluster or, indeed, the trajectories of individual cells outside a cluster. The results show that motility inside a PC-3 cluster is not restricted to slight motion due to cluster expansion, but rather consists of a marked cell movement within the cluster. The proposed equipment enables in vitro aggregated cell motility to be studied. This method can, therefore, be used in pharmacological studies in order to select anti-motility related compounds. The compounds selected by the equipment described could then be tested in vivo as potential anti-metastatic.

ChpA Controls Twitching Motility and Broadly Affects Gene Expression in the Biological Control Agent Lysobacter enzymogenes.

PubMed

Zhou, Mimi; Shen, Danyu; Xu, Gaoge; Liu, Fengquan; Qian, Guoliang

2017-05-01

Lysobacter enzymogenes (L. enzymogenes) is an agriculturally important Gram-negative bacterium that employs T4P (type IV pili)-driven twitching motility to exhibit its antifungal function. Yet, it is still unclear how this bacterium regulates its twitching motility. Here, by using strain OH11 as the working model organism, we showed that a hybrid two-component system ChpA acts as a positive regulator in controlling twitching motility in L. enzymogenes. ChpA is a hybrid TCS (two-component transduction system) contains 7 domains including those for auto-phosphorylation and phosphate group transfer, as well as a phosphate receiver (REC) domain. Mutation of chpA completely abolished the wild-type twitching motility, as evidenced by the absence of mobile cells at the margin of the mutant colonies. Further studies of domain-deletion and phenotypic characterization reveal that domains responsible for phosphorylation and phosphotransfer, but not the REC domain, were indispensable for ChpA in regulating twitching motility. Transcriptome analyses of the chpA knockout strain indicated that ChpA was extensively involved in controlling expression of a wide variety of genes (totaling 243). The products of these differentially expressed genes were involved in multiple physiological and biological functions in L. enzymogenes. Thus, we have not only identified a new regulator controlling twitching motility in L. enzymogenes, but also provided the first report demonstrating the broad impact of the conserved ChpA in gene regulation in Gram-negative bacteria.

Optical trapping reveals propulsion forces, power generation and motility efficiency of the unicellular parasites Trypanosoma brucei brucei

NASA Astrophysics Data System (ADS)

Stellamanns, Eric; Uppaluri, Sravanti; Hochstetter, Axel; Heddergott, Niko; Engstler, Markus; Pfohl, Thomas

2014-10-01

Unicellular parasites have developed sophisticated swimming mechanisms to survive in a wide range of environments. Cell motility of African trypanosomes, parasites responsible for fatal illness in humans and animals, is crucial both in the insect vector and the mammalian host. Using millisecond-scale imaging in a microfluidics platform along with a custom made optical trap, we are able to confine single cells to study trypanosome motility. From the trapping characteristics of the cells, we determine the propulsion force generated by cells with a single flagellum as well as of dividing trypanosomes with two fully developed flagella. Estimates of the dissipative energy and the power generation of single cells obtained from the motility patterns of the trypanosomes within the optical trap indicate that specific motility characteristics, in addition to locomotion, may be required for antibody clearance. Introducing a steerable second optical trap we could further measure the force, which is generated at the flagellar tip. Differences in the cellular structure of the trypanosomes are correlated with the trapping and motility characteristics and in consequence with their propulsion force, dissipative energy and power generation.

Optical trapping reveals propulsion forces, power generation and motility efficiency of the unicellular parasites Trypanosoma brucei brucei.

PubMed

Stellamanns, Eric; Uppaluri, Sravanti; Hochstetter, Axel; Heddergott, Niko; Engstler, Markus; Pfohl, Thomas

2014-10-01

Unicellular parasites have developed sophisticated swimming mechanisms to survive in a wide range of environments. Cell motility of African trypanosomes, parasites responsible for fatal illness in humans and animals, is crucial both in the insect vector and the mammalian host. Using millisecond-scale imaging in a microfluidics platform along with a custom made optical trap, we are able to confine single cells to study trypanosome motility. From the trapping characteristics of the cells, we determine the propulsion force generated by cells with a single flagellum as well as of dividing trypanosomes with two fully developed flagella. Estimates of the dissipative energy and the power generation of single cells obtained from the motility patterns of the trypanosomes within the optical trap indicate that specific motility characteristics, in addition to locomotion, may be required for antibody clearance. Introducing a steerable second optical trap we could further measure the force, which is generated at the flagellar tip. Differences in the cellular structure of the trypanosomes are correlated with the trapping and motility characteristics and in consequence with their propulsion force, dissipative energy and power generation.

A quantitative evaluation of cell migration by the phagokinetic track motility assay.

PubMed

Nogalski, Maciej T; Chan, Gary C T; Stevenson, Emily V; Collins-McMillen, Donna K; Yurochko, Andrew D

2012-12-04

Cellular motility is an important biological process for both unicellular and multicellular organisms. It is essential for movement of unicellular organisms towards a source of nutrients or away from unsuitable conditions, as well as in multicellular organisms for tissue development, immune surveillance and wound healing, just to mention a few roles(1,2,3). Deregulation of this process can lead to serious neurological, cardiovascular and immunological diseases, as well as exacerbated tumor formation and spread(4,5). Molecularly, actin polymerization and receptor recycling have been shown to play important roles in creating cellular extensions (lamellipodia), that drive the forward movement of the cell(6,7,8). However, many biological questions about cell migration remain unanswered. The central role for cellular motility in human health and disease underlines the importance of understanding the specific mechanisms involved in this process and makes accurate methods for evaluating cell motility particularly important. Microscopes are usually used to visualize the movement of cells. However, cells move rather slowly, making the quantitative measurement of cell migration a resource-consuming process requiring expensive cameras and software to create quantitative time-lapsed movies of motile cells. Therefore, the ability to perform a quantitative measurement of cell migration that is cost-effective, non-laborious, and that utilizes common laboratory equipment is a great need for many researchers. The phagokinetic track motility assay utilizes the ability of a moving cell to clear gold particles from its path to create a measurable track on a colloidal gold-coated glass coverslip(9,10). With the use of freely available software, multiple tracks can be evaluated for each treatment to accomplish statistical requirements. The assay can be utilized to assess motility of many cell types, such as cancer cells(11,12), fibroblasts(9), neutrophils(13), skeletal muscle cells(14), keratinocytes(15), trophoblasts(16), endothelial cells(17), and monocytes(10,18-22). The protocol involves the creation of slides coated with gold nanoparticles (Au°) that are generated by a reduction of chloroauric acid (Au(3+)) by sodium citrate. This method was developed by Turkevich et al. in 1951(23) and then improved in the 1970s by Frens et al.(24,25). As a result of this chemical reduction step, gold particles (10-20 nm in diameter) precipitate from the reaction mixture and can be applied to glass coverslips, which are then ready for use in cellular migration analyses(9,26,27). In general, the phagokinetic track motility assay is a quick, quantitative and easy measure of cellular motility. In addition, it can be utilized as a simple high-throughput assay, for use with cell types that are not amenable to time-lapsed imaging, as well as other uses depending on the needs of the researcher. Together, the ability to quantitatively measure cellular motility of multiple cell types without the need for expensive microscopes and software, along with the use of common laboratory equipment and chemicals, make the phagokinetic track motility assay a solid choice for scientists with an interest in understanding cellular motility.

Thiols of flagellar proteins are essential for progressive motility in human spermatozoa.

PubMed

Cabrillana, María Eugenia; Monclus, María de Los Ángeles; Lancellotti, Tania Estefania Sáez; Boarelli, Paola Vanina; Vincenti, Amanda Edith; Fornés, Miguel Matias; Sanabria, Eduardo Alfredo; Fornés, Miguel Walter

2017-07-01

Male infertility is a disorder of the reproductive system defined by the failure to achieve a clinical pregnancy after 12 months or more of regular unprotected sexual intercourse. The presence of low-motile or immotile spermatozoa is one of many causes of infertility; however, this observation provides little or no information regarding the pathogenesis of the malfunction. Good sperm motility depends on correct assembly of the sperm tail in the testis and efficient maturation during epididymal transit. Thiols of flagellar proteins, such as outer dense fibre protein 1 (ODF1), are oxidised to form disulfides during epididymal transit and the spermatozoa become motile. This study was designed to determine how oxidative changes in protein thiol status affect progressive motility in human spermatozoa. Monobromobimane (mBBr) was used as a specific thiol marker and disruptor of sperm progressive motility. When mBBr was blocked by dithiothreitol it did not promote motility changes. The analysis of mBBr-treated spermatozoa revealed a reduction of progressive motility and an increased number of spermatozoa with non-progressive motility without affecting ATP production. Laser confocal microscopy and western blot analysis showed that one of the mBBr-positive proteins reacted with an antibody to ODF1. Monobromobimane fluorescence intensity of the sperm tail was lower in normozoospermic than asthenozoospermic men, suggesting that thiol oxidation in spermatozoa of asthenozoospermic men is incomplete. Our findings indicate that mBBr affects the thiol status of ODF1 in human spermatozoa and interferes with progressive motility.

Periodic growth of bacterial colonies

NASA Astrophysics Data System (ADS)

Yamazaki, Yoshihiro; Ikeda, Takemasa; Shimada, Hirotoshi; Hiramatsu, Fumiko; Kobayashi, Naoki; Wakita, Jun-ichi; Itoh, Hiroto; Kurosu, Sayuri; Nakatsuchi, Michio; Matsuyama, Tohey; Matsushita, Mitsugu

2005-06-01

The formation of concentric ring colonies by bacterial species Bacillus subtilis and Proteus mirabilis has been investigated experimentally, focusing our attention on the dependence of local cell density upon the bacterial motility. It has been confirmed that these concentric ring colonies reflect the periodic change of the bacterial motility between motile cell state and immotile cell state. We conclude that this periodic change is macroscopically determined neither by biological factors (i.e., biological clock) nor by chemical factors (chemotaxis as inhibitor). And our experimental results strongly suggest that the essential factor for the change of the bacterial motility during concentric ring formation is the local cell density.

Diatom Cell Size, Coloniality and Motility: Trade-Offs between Temperature, Salinity and Nutrient Supply with Climate Change

PubMed Central

Svensson, Filip; Norberg, Jon; Snoeijs, Pauline

2014-01-01

Reduction in body size has been proposed as a universal response of organisms, both to warming and to decreased salinity. However, it is still controversial if size reduction is caused by temperature or salinity on their own, or if other factors interfere as well. We used natural benthic diatom communities to explore how “body size” (cells and colonies) and motility change along temperature (2–26°C) and salinity (0.5–7.8) gradients in the brackish Baltic Sea. Fourth-corner analysis confirmed that small cell and colony sizes were associated with high temperature in summer. Average community cell volume decreased linearly with 2.2% per °C. However, cells were larger with artificial warming when nutrient concentrations were high in the cold season. Average community cell volume increased by 5.2% per °C of artificial warming from 0 to 8.5°C and simultaneously there was a selection for motility, which probably helped to optimize growth rates by trade-offs between nutrient supply and irradiation. Along the Baltic Sea salinity gradient cell size decreased with decreasing salinity, apparently mediated by nutrient stoichiometry. Altogether, our results suggest that climate change in this century may polarize seasonality by creating two new niches, with elevated temperature at high nutrient concentrations in the cold season (increasing cell size) and elevated temperature at low nutrient concentrations in the warm season (decreasing cell size). Higher temperature in summer and lower salinity by increased land-runoff are expected to decrease the average cell size of primary producers, which is likely to affect the transfer of energy to higher trophic levels. PMID:25279720

Diatom cell size, coloniality and motility: trade-offs between temperature, salinity and nutrient supply with climate change.

PubMed

Svensson, Filip; Norberg, Jon; Snoeijs, Pauline

2014-01-01

Reduction in body size has been proposed as a universal response of organisms, both to warming and to decreased salinity. However, it is still controversial if size reduction is caused by temperature or salinity on their own, or if other factors interfere as well. We used natural benthic diatom communities to explore how "body size" (cells and colonies) and motility change along temperature (2-26°C) and salinity (0.5-7.8) gradients in the brackish Baltic Sea. Fourth-corner analysis confirmed that small cell and colony sizes were associated with high temperature in summer. Average community cell volume decreased linearly with 2.2% per °C. However, cells were larger with artificial warming when nutrient concentrations were high in the cold season. Average community cell volume increased by 5.2% per °C of artificial warming from 0 to 8.5°C and simultaneously there was a selection for motility, which probably helped to optimize growth rates by trade-offs between nutrient supply and irradiation. Along the Baltic Sea salinity gradient cell size decreased with decreasing salinity, apparently mediated by nutrient stoichiometry. Altogether, our results suggest that climate change in this century may polarize seasonality by creating two new niches, with elevated temperature at high nutrient concentrations in the cold season (increasing cell size) and elevated temperature at low nutrient concentrations in the warm season (decreasing cell size). Higher temperature in summer and lower salinity by increased land-runoff are expected to decrease the average cell size of primary producers, which is likely to affect the transfer of energy to higher trophic levels.

Comparative Plasmodium gene overexpression reveals distinct perturbation of sporozoite transmission by profilin.

PubMed

Sato, Yuko; Hliscs, Marion; Dunst, Josefine; Goosmann, Christian; Brinkmann, Volker; Montagna, Georgina N; Matuschewski, Kai

2016-07-15

Plasmodium relies on actin-based motility to migrate from the site of infection and invade target cells. Using a substrate-dependent gliding locomotion, sporozoites are able to move at fast speed (1-3 μm/s). This motility relies on a minimal set of actin regulatory proteins and occurs in the absence of detectable filamentous actin (F-actin). Here we report an overexpression strategy to investigate whether perturbations of F-actin steady-state levels affect gliding locomotion and host invasion. We selected two vital Plasmodium berghei G-actin-binding proteins, C-CAP and profilin, in combination with three stage-specific promoters and mapped the phenotypes afforded by overexpression in all three extracellular motile stages. We show that in merozoites and ookinetes, additional expression does not impair life cycle progression. In marked contrast, overexpression of C-CAP and profilin in sporozoites impairs circular gliding motility and salivary gland invasion. The propensity for productive motility correlates with actin accumulation at the parasite tip, as revealed by combinations of an actin-stabilizing drug and transgenic parasites. Strong expression of profilin, but not C-CAP, resulted in complete life cycle arrest. Comparative overexpression is an alternative experimental genetic strategy to study essential genes and reveals effects of regulatory imbalances that are not uncovered from deletion-mutant phenotyping. © 2016 Sato et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

The Toxoplasma Acto-MyoA Motor Complex Is Important but Not Essential for Gliding Motility and Host Cell Invasion

PubMed Central

Jackson, Allison J.; Whitelaw, Jamie A.; Pall, Gurman; Black, Jennifer Ann; Ferguson, David J. P.; Tardieux, Isabelle; Mogilner, Alex; Meissner, Markus

2014-01-01

Apicomplexan parasites are thought to actively invade the host cell by gliding motility. This movement is powered by the parasite's own actomyosin system, and depends on the regulated polymerisation and depolymerisation of actin to generate the force for gliding and host cell penetration. Recent studies demonstrated that Toxoplasma gondii can invade the host cell in the absence of several core components of the invasion machinery, such as the motor protein myosin A (MyoA), the microneme proteins MIC2 and AMA1 and actin, indicating the presence of alternative invasion mechanisms. Here the roles of MyoA, MLC1, GAP45 and Act1, core components of the gliding machinery, are re-dissected in detail. Although important roles of these components for gliding motility and host cell invasion are verified, mutant parasites remain invasive and do not show a block of gliding motility, suggesting that other mechanisms must be in place to enable the parasite to move and invade the host cell. A novel, hypothetical model for parasite gliding motility and invasion is presented based on osmotic forces generated in the cytosol of the parasite that are converted into motility. PMID:24632839

Epidermal growth factor promotes a mesenchymal over an amoeboid motility of MDA-MB-231 cells embedded within a 3D collagen matrix

NASA Astrophysics Data System (ADS)

Geum, Dongil T.; Kim, Beum Jun; Chang, Audrey E.; Hall, Matthew S.; Wu, Mingming

2016-01-01

The receptor of epidermal growth factor (EGFR) critically regulates tumor cell invasion and is a potent therapeutic target for treatment of many types of cancers, including carcinomas and glioblastomas. It is known that EGF regulates cell motility when tumor cells are embedded within a 3D biomatrix. However, roles of EGF in modulating tumor cell motility phenotype are largely unknown. In this article, we report that EGF promotes a mesenchymal over an amoeboid motility phenotype using a malignant breast tumor cell line, MDA-MB-231, embedded within a 3D collagen matrix. Amoeboid cells are rounded in shape, while mesenchymal cells are elongated, and their migrations are governed by a distinctly different set of biomolecules. Using single cell tracking analysis, we also show that EGF promotes cell dissemination through a significant increase in cell persistence along with a moderate increase of speed. The increase of persistence is correlated with the increase of the percentage of the mesenchymal cells within the population. Our work reveals a novel role of microenvironmental cue, EGF, in modulating heterogeneity and plasticity of tumor cell motility phenotype. In addition, it suggests a potential visual cue for diagnosing invasive states of breast cancer cells. This work can be easily extended beyond breast cancer cells.

Mechanism of Actin-Based Motility

NASA Astrophysics Data System (ADS)

Pantaloni, Dominique; Le Clainche, Christophe; Carlier, Marie-France

2001-05-01

Spatially controlled polymerization of actin is at the origin of cell motility and is responsible for the formation of cellular protrusions like lamellipodia. The pathogens Listeria monocytogenes and Shigella flexneri, which undergo actin-based propulsion, are acknowledged models of the leading edge of lamellipodia. Actin-based motility of the bacteria or of functionalized microspheres can be reconstituted in vitro from only five pure proteins. Movement results from the regulated site-directed treadmilling of actin filaments, consistent with observations of actin dynamics in living motile cells and with the biochemical properties of the components of the synthetic motility medium.

[Response of the algae Gymnodinium kovalevskii (Dinophyta) to exposure to synthetic detergents and distillation].

PubMed

Aĭzdaĭcher, N A

2000-01-01

The effects of synthetic detergents and combined effects of synthetic detergents and water freshening on growth characteristics of the alga Gymnodinium kovalevskii (Dinophyta) were studied. Low concentrations of synthetic detergents (0.1 and 1.0 mg/l) stimulated the algal growth. Elevated concentrations inhibited cell division, affected their motility and induced morphological changes. Contamination with synthetic detergents adversely affected the adaptation plasticity of algae with respect to salinity.

Motile hepatocellular carcinoma cells preferentially secret sugar metabolism regulatory proteins via exosomes.

PubMed

Zhang, Jing; Lu, Shaohua; Zhou, Ye; Meng, Kun; Chen, Zhipeng; Cui, Yizhi; Shi, Yunfeng; Wang, Tong; He, Qing-Yu

2017-07-01

Exosomes are deliverers of critically functional proteins, capable of transforming target cells in numerous cancers, including hepatocellular carcinoma (HCC). We hypothesize that the motility of HCC cells can be featured by comparative proteome of exosomes. Hence, we performed the super-SILAC-based MS analysis on the exosomes secreted by three human HCC cell lines, including the non-motile Hep3B cell, and the motile 97H and LM3 cells. More than 1400 exosomal proteins were confidently quantified in each MS analysis with highly biological reproducibility. We justified that 469 and 443 exosomal proteins represented differentially expressed proteins (DEPs) in the 97H/Hep3B and LM3/Hep3B comparisons, respectively. These DEPs focused on sugar metabolism-centric canonical pathways per ingenuity pathway analysis, which was consistent with the gene ontology analysis on biological process enrichment. These pathways included glycolysis I, gluconeogenesis I and pentose phosphate pathways; and the DEPs enriched in these pathways could form a tightly connected network. By analyzing the relative abundance of proteins and translating mRNAs, we found significantly positive correlation between exosomes and cells. The involved exosomal proteins were again focusing on sugar metabolism. In conclusion, motile HCC cells tend to preferentially export more sugar metabolism-associated proteins via exosomes that differentiate them from non-motile HCC cells. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Observing planar cell polarity in multiciliated mouse airway epithelial cells.

PubMed

Vladar, Eszter K; Lee, Yin Loon; Stearns, Tim; Axelrod, Jeffrey D

2015-01-01

The concerted movement of cilia propels inhaled contaminants out of the lungs, safeguarding the respiratory system from toxins, pathogens, pollutants, and allergens. Motile cilia on the multiciliated cells (MCCs) of the airway epithelium are physically oriented along the tissue axis for directional motility, which depends on the planar cell polarity (PCP) signaling pathway. The MCCs of the mouse respiratory epithelium have emerged as an important model for the study of motile ciliogenesis and the PCP signaling mechanism. Unlike other motile ciliated or planar polarized tissues, airway epithelial cells are relatively easily accessible and primary cultures faithfully model many of the essential features of the in vivo tissue. There is growing interest in understanding how cells acquire and polarize motile cilia due to the impact of mucociliary clearance on respiratory health. Here, we present methods for observing and quantifying the planar polarized orientation of motile cilia both in vivo and in primary culture airway epithelial cells. We describe how to acquire and evaluate electron and light microscopy images of ciliary ultrastructural features that reveal planar polarized orientation. Furthermore, we describe the immunofluorescence localization of PCP pathway components as a simple readout for airway epithelial planar polarization and ciliary orientation. These methods can be adapted to observe ciliary orientation in other multi- and monociliated cells and to detect PCP pathway activity in any tissue or cell type. Copyright © 2015 Elsevier Inc. All rights reserved.

Effect of HSP27 on Human Breast Tumor Cell Growth and Motility

DTIC Science & Technology

1999-08-01

the small stress protein, HSP27 , on growth and motility characteristics of normal and tumor-derived human mammary cell lines. We hypothesized that...cells overexpressing HSP27 would show increased motility, altered chemotactic properties, increased resistance to heat killing and to certain drugs...Donna has prepared and studied 19 clonal MDA23 1 breast tumor cell lines that overexpress human HSP27 , and determined that, while heat resistance is

Mena invasive (MenaINV) promotes multicellular streaming motility and transendothelial migration in a mouse model of breast cancer

PubMed Central

Roussos, Evanthia T.; Balsamo, Michele; Alford, Shannon K.; Wyckoff, Jeffrey B.; Gligorijevic, Bojana; Wang, Yarong; Pozzuto, Maria; Stobezki, Robert; Goswami, Sumanta; Segall, Jeffrey E.; Lauffenburger, Douglas A.; Bresnick, Anne R.; Gertler, Frank B.; Condeelis, John S.

2011-01-01

We have shown previously that distinct Mena isoforms are expressed in invasive and migratory tumor cells in vivo and that the invasion isoform (MenaINV) potentiates carcinoma cell metastasis in murine models of breast cancer. However, the specific step of metastatic progression affected by this isoform and the effects on metastasis of the Mena11a isoform, expressed in primary tumor cells, are largely unknown. Here, we provide evidence that elevated MenaINV increases coordinated streaming motility, and enhances transendothelial migration and intravasation of tumor cells. We demonstrate that promotion of these early stages of metastasis by MenaINV is dependent on a macrophage–tumor cell paracrine loop. Our studies also show that increased Mena11a expression correlates with decreased expression of colony-stimulating factor 1 and a dramatically decreased ability to participate in paracrine-mediated invasion and intravasation. Our results illustrate the importance of paracrine-mediated cell streaming and intravasation on tumor cell dissemination, and demonstrate that the relative abundance of MenaINV and Mena11a helps to regulate these key stages of metastatic progression in breast cancer cells. PMID:21670198

The alternatively-included 11a sequence modifies the effects of Mena on actin cytoskeletal organization and cell behavior

PubMed Central

Balsamo, Michele; Mondal, Chandrani; Carmona, Guillaume; McClain, Leslie M.; Riquelme, Daisy N.; Tadros, Jenny; Ma, Duan; Vasile, Eliza; Condeelis, John S.; Lauffenburger, Douglas A.; Gertler, Frank B.

2016-01-01

During tumor progression, alternative splicing gives rise to different Mena protein isoforms. We analyzed how Mena11a, an isoform enriched in epithelia and epithelial-like cells, affects Mena-dependent regulation of actin dynamics and cell behavior. While other Mena isoforms promote actin polymerization and drive membrane protrusion, we find that Mena11a decreases actin polymerization and growth factor-stimulated membrane protrusion at lamellipodia. Ectopic Mena11a expression slows mesenchymal-like cell motility, while isoform-specific depletion of endogenous Mena11a in epithelial-like tumor cells perturbs cell:cell junctions and increases membrane protrusion and overall cell motility. Mena11a can dampen membrane protrusion and reduce actin polymerization in the absence of other Mena isoforms, indicating that it is not simply an inactive Mena isoform. We identify a phosphorylation site within 11a that is required for some Mena11a-specific functions. RNA-seq data analysis from patient cohorts demonstrates that the difference between mRNAs encoding constitutive Mena sequences and those containing the 11a exon correlates with metastasis in colorectal cancer, suggesting that 11a exon exclusion contributes to invasive phenotypes and leads to poor clinical outcomes. PMID:27748415

Mena invasive (MenaINV) promotes multicellular streaming motility and transendothelial migration in a mouse model of breast cancer.

PubMed

Roussos, Evanthia T; Balsamo, Michele; Alford, Shannon K; Wyckoff, Jeffrey B; Gligorijevic, Bojana; Wang, Yarong; Pozzuto, Maria; Stobezki, Robert; Goswami, Sumanta; Segall, Jeffrey E; Lauffenburger, Douglas A; Bresnick, Anne R; Gertler, Frank B; Condeelis, John S

2011-07-01

We have shown previously that distinct Mena isoforms are expressed in invasive and migratory tumor cells in vivo and that the invasion isoform (Mena(INV)) potentiates carcinoma cell metastasis in murine models of breast cancer. However, the specific step of metastatic progression affected by this isoform and the effects on metastasis of the Mena11a isoform, expressed in primary tumor cells, are largely unknown. Here, we provide evidence that elevated Mena(INV) increases coordinated streaming motility, and enhances transendothelial migration and intravasation of tumor cells. We demonstrate that promotion of these early stages of metastasis by Mena(INV) is dependent on a macrophage-tumor cell paracrine loop. Our studies also show that increased Mena11a expression correlates with decreased expression of colony-stimulating factor 1 and a dramatically decreased ability to participate in paracrine-mediated invasion and intravasation. Our results illustrate the importance of paracrine-mediated cell streaming and intravasation on tumor cell dissemination, and demonstrate that the relative abundance of Mena(INV) and Mena11a helps to regulate these key stages of metastatic progression in breast cancer cells.

The alternatively-included 11a sequence modifies the effects of Mena on actin cytoskeletal organization and cell behavior.

PubMed

Balsamo, Michele; Mondal, Chandrani; Carmona, Guillaume; McClain, Leslie M; Riquelme, Daisy N; Tadros, Jenny; Ma, Duan; Vasile, Eliza; Condeelis, John S; Lauffenburger, Douglas A; Gertler, Frank B

2016-10-17

During tumor progression, alternative splicing gives rise to different Mena protein isoforms. We analyzed how Mena11a, an isoform enriched in epithelia and epithelial-like cells, affects Mena-dependent regulation of actin dynamics and cell behavior. While other Mena isoforms promote actin polymerization and drive membrane protrusion, we find that Mena11a decreases actin polymerization and growth factor-stimulated membrane protrusion at lamellipodia. Ectopic Mena11a expression slows mesenchymal-like cell motility, while isoform-specific depletion of endogenous Mena11a in epithelial-like tumor cells perturbs cell:cell junctions and increases membrane protrusion and overall cell motility. Mena11a can dampen membrane protrusion and reduce actin polymerization in the absence of other Mena isoforms, indicating that it is not simply an inactive Mena isoform. We identify a phosphorylation site within 11a that is required for some Mena11a-specific functions. RNA-seq data analysis from patient cohorts demonstrates that the difference between mRNAs encoding constitutive Mena sequences and those containing the 11a exon correlates with metastasis in colorectal cancer, suggesting that 11a exon exclusion contributes to invasive phenotypes and leads to poor clinical outcomes.

Myocardin-related transcription factors and SRF are required for cytoskeletal dynamics and experimental metastasis.

PubMed

Medjkane, Souhila; Perez-Sanchez, Cristina; Gaggioli, Cedric; Sahai, Erik; Treisman, Richard

2009-03-01

Rho GTPases control cytoskeletal dynamics through cytoplasmic effectors and regulate transcriptional activation through myocardin-related transcription factors (MRTFs), which are co-activators for serum response factor (SRF). We used RNA interference to investigate the contribution of the MRTF-SRF pathway to cytoskeletal dynamics in MDA-MB-231 breast carcinoma and B16F2 melanoma cells, in which basal MRTF-SRF activity is Rho-dependent. Depletion of MRTFs or SRF reduced cell adhesion, spreading, invasion and motility in culture, without affecting proliferation or inducing apoptosis. MRTF-depleted tumour cell xenografts showed reduced cell motility but proliferated normally. Tumour cells depleted of MRTF or SRF failed to colonize the lung from the bloodstream, being unable to persist after their arrival in the lung. Only a few genes show MRTF-dependent expression in both cell lines. Two of these, MYH9 (NMHCIIa) and MYL9 (MLC2), are also required for invasion and lung colonization. Conversely, expression of activated MAL/MRTF-A increases lung colonization by poorly metastatic B16F0 cells. Actin-based cell behaviour and experimental metastasis thus require Rho-dependent nuclear signalling through the MRTF-SRF network.

Microscopic Analysis of Bacterial Motility at High Pressure

PubMed Central

Nishiyama, Masayoshi; Sowa, Yoshiyuki

2012-01-01

The bacterial flagellar motor is a molecular machine that converts an ion flux to the rotation of a helical flagellar filament. Counterclockwise rotation of the filaments allows them to join in a bundle and propel the cell forward. Loss of motility can be caused by environmental factors such as temperature, pH, and solvation. Hydrostatic pressure is also a physical inhibitor of bacterial motility, but the detailed mechanism of this inhibition is still unknown. Here, we developed a high-pressure microscope that enables us to acquire high-resolution microscopic images, regardless of applied pressures. We also characterized the pressure dependence of the motility of swimming Escherichia coli cells and the rotation of single flagellar motors. The fraction and speed of swimming cells decreased with increased pressure. At 80 MPa, all cells stopped swimming and simply diffused in solution. After the release of pressure, most cells immediately recovered their initial motility. Direct observation of the motility of single flagellar motors revealed that at 80 MPa, the motors generate torque that should be sufficient to join rotating filaments in a bundle. The discrepancy in the behavior of free swimming cells and individual motors could be due to the applied pressure inhibiting the formation of rotating filament bundles that can propel the cell body in an aqueous environment. PMID:22768943

The N-terminal leucine-zipper motif in PTRF/cavin-1 is essential and sufficient for its caveolae-association

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wei, Zhuang; Laboratory of System Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031; Zou, Xinle

2015-01-16

Highlight: • The N-terminal leucine-zipper motif in PTRF/cavin-1 determines caveolar association. • Different cellular localization of PTRF/cavin-1 influences its serine 389 and 391 phosphorylation state. • PTRF/cavin-1 regulates cell motility via its caveolar association. - Abstract: PTRF/cavin-1 is a protein of two lives. Its reported functions in ribosomal RNA synthesis and in caveolae formation happen in two different cellular locations: nucleus vs. plasma membrane. Here, we identified that the N-terminal leucine-zipper motif in PTRF/cavin-1 was essential for the protein to be associated with caveolae in plasma membrane. It could counteract the effect of nuclear localization sequence in the molecule (AAmore » 235–251). Deletion of this leucine-zipper motif from PTRF/cavin-1 caused the mutant to be exclusively localized in nuclei. The fusion of this leucine-zipper motif with histone 2A, which is a nuclear protein, could induce the fusion protein to be exported from nucleus. Cell migration was greatly inhibited in PTRF/cavin-1{sup −/−} mouse embryonic fibroblasts (MEFs). The inhibited cell motility could only be rescued by exogenous cavin-1 but not the leucine-zipper motif deleted cavin-1 mutant. Plasma membrane dynamics is an important factor in cell motility control. Our results suggested that the membrane dynamics in cell migration is affected by caveolae associated PTRF/cavin-1.« less

In vitro motility of cells from human epidermoid carcinomas. A study by phase-contrast and reflection-contrast cinematography.

PubMed

Haemmerli, G; Sträuli, P

1981-05-15

The motile behavior of six cell lines derived from human squamous carcinomas (two from the larynx, four from the tongue) was studied by cinematography under phase- and reflection-contrast illumination. The recorded cell activities consist in spreading, stationary and translocation motility, and aggregate formation. Within this common pattern, quantitative modifications ("sub-pattern") are stable properties of the individual cells lines. Such modifications are particularly evident with regard to the dynamic texture of the aggregates which ranges from loose, netlike structures to compact islands with smooth borders. Accordingly, the intensity of cell traffic within and around the aggregates varies considerably. It is discussed to what extent the in vitro motility of the carcinoma cell populations reflects their behavior in the organism and thus the significance of cell movements for invasion.

Viscosity-dependent variations in the cell shape and swimming manner of Leptospira.

PubMed

Takabe, Kyosuke; Tahara, Hajime; Islam, Md Shafiqul; Affroze, Samia; Kudo, Seishi; Nakamura, Shuichi

2017-02-01

Spirochaetes are spiral or flat-wave-shaped Gram-negative bacteria that have periplasmic flagella between the peptidoglycan layer and outer membrane. Rotation of the periplasmic flagella transforms the cell body shape periodically, allowing the cell to swim in aqueous environments. Because the virulence of motility-deficient mutants of pathogenic species is drastically attenuated, motility is thought to be an essential virulence factor in spirochaetes. However, it remains unknown how motility practically contributes to the infection process. We show here that the cell body configuration and motility of the zoonotic spirochaete Leptospira changes depending on the viscosity of the medium. Leptospira swim and reverse the swimming direction by transforming the cell body. Motility analysis showed that the frequency of cell shape transformation was increased by increasing the viscosity of the medium. The increased cell body transformation induced highly frequent reversal of the swimming direction. A simple kinetic model based on the experimental results shows that the viscosity-induced increase in reversal limits cell migration, resulting in the accumulation of cells in high-viscosity regions. This behaviour could facilitate the colonization of the spirochaete on host tissues covered with mucosa.

Effect of HSP27 on Human Breast Tumor Cell Growth and Motility.

DTIC Science & Technology

1997-09-01

the small heat shock protein, Hsp27 , on growth and motility characteristics of human mammary tumor cell lines. Since Hsp27 regulates actin...microfilament dynamics, we hypothesize that cells expressing high levels of Hsp27 will show increased motility and altered chemotactic properties, in addition to...significantly elevated levels of Hsp27 has proven to be daunting. Down regulation of Hsp27 levels in MCF7 cells using antisense technology has also

Disruption of the A-Kinase Anchoring Domain in Flagellar Radial Spoke Protein 3 Results in Unregulated Axonemal cAMP-dependent Protein Kinase Activity and Abnormal Flagellar Motility

PubMed Central

Gaillard, Anne R.; Fox, Laura A.; Rhea, Jeanne M.; Craige, Branch

2006-01-01

Biochemical studies of Chlamydomonas flagellar axonemes revealed that radial spoke protein (RSP) 3 is an A-kinase anchoring protein (AKAP). To determine the physiological role of PKA anchoring in the axoneme, an RSP3 mutant, pf14, was transformed with an RSP3 gene containing a mutation in the PKA-binding domain. Analysis of several independent transformants revealed that the transformed cells exhibit an unusual phenotype: a fraction of the cells swim normally; the remainder of the cells twitch feebly or are paralyzed. The abnormal/paralyzed motility is not due to an obvious deficiency of radial spoke assembly, and the phenotype cosegregates with the mutant RSP3. We postulated that paralysis was due to failure in targeting and regulation of axonemal cAMP-dependent protein kinase (PKA). To test this, reactivation experiments of demembranated cells were performed in the absence or presence of PKA inhibitors. Importantly, motility in reactivated cell models mimicked the live cell phenotype with nearly equal fractions of motile and paralyzed cells. PKA inhibitors resulted in a twofold increase in the number of motile cells, rescuing paralysis. These results confirm that flagellar RSP3 is an AKAP and reveal that a mutation in the PKA binding domain results in unregulated axonemal PKA activity and inhibition of normal motility. PMID:16571668

Displacement correlations between a single mesenchymal-like cell and its nucleus effectively link subcellular activities and motility in cell migration analysis

NASA Astrophysics Data System (ADS)

Lan, Tian; Cheng, Kai; Ren, Tina; Arce, Stephen Hugo; Tseng, Yiider

2016-09-01

Cell migration is an essential process in organism development and physiological maintenance. Although current methods permit accurate comparisons of the effects of molecular manipulations and drug applications on cell motility, effects of alterations in subcellular activities on motility cannot be fully elucidated from those methods. Here, we develop a strategy termed cell-nuclear (CN) correlation to parameterize represented dynamic subcellular activities and to quantify their contributions in mesenchymal-like migration. Based on the biophysical meaning of the CN correlation, we propose a cell migration potential index (CMPI) to measure cell motility. When the effectiveness of CMPI was evaluated with respect to one of the most popular cell migration analysis methods, Persistent Random Walk, we found that the cell motility estimates among six cell lines used in this study were highly consistent between these two approaches. Further evaluations indicated that CMPI can be determined using a shorter time period and smaller cell sample size, and it possesses excellent reliability and applicability, even in the presence of a wide range of noise, as might be generated from individual imaging acquisition systems. The novel approach outlined here introduces a robust strategy through an analysis of subcellular locomotion activities for single cell migration assessment.

Ring-Shaped Microlanes and Chemical Barriers as a Platform for Probing Single-Cell Migration.

PubMed

Schreiber, Christoph; Segerer, Felix J; Wagner, Ernst; Roidl, Andreas; Rädler, Joachim O

2016-05-31

Quantification and discrimination of pharmaceutical and disease-related effects on cell migration requires detailed characterization of single-cell motility. In this context, micropatterned substrates that constrain cells within defined geometries facilitate quantitative readout of locomotion. Here, we study quasi-one-dimensional cell migration in ring-shaped microlanes. We observe bimodal behavior in form of alternating states of directional migration (run state) and reorientation (rest state). Both states show exponential lifetime distributions with characteristic persistence times, which, together with the cell velocity in the run state, provide a set of parameters that succinctly describe cell motion. By introducing PEGylated barriers of different widths into the lane, we extend this description by quantifying the effects of abrupt changes in substrate chemistry on migrating cells. The transit probability decreases exponentially as a function of barrier width, thus specifying a characteristic penetration depth of the leading lamellipodia. Applying this fingerprint-like characterization of cell motion, we compare different cell lines, and demonstrate that the cancer drug candidate salinomycin affects transit probability and resting time, but not run time or run velocity. Hence, the presented assay allows to assess multiple migration-related parameters, permits detailed characterization of cell motility, and has potential applications in cell biology and advanced drug screening.

aPKCζ affects directed cell migration through the regulation of myosin light chain phosphorylation

PubMed Central

Petrov, Daria; Dahan, Inbal; Cohen-Kfir, Einav; Ravid, Shoshana

2017-01-01

ABSTRACT Cell motility is an essential cellular process for a variety of biological events. It requires cross-talk between the signaling and the cytoskeletal systems. Despite the recognized importance of aPKCζ for cell motility, there is little understanding of the mechanism by which aPKCζ mediates extracellular signals to the cytoskeleton. In the present study, we report that aPKCζ is required for the cellular organization of acto-non-muscle myosin II (NMII) cytoskeleton, for proper cell adhesion and directed cell migration. We show that aPKCζ mediates EGF-dependent RhoA activation and recruitment to the cell membrane. We also show that aPKCζ mediates EGF-dependent myosin light chain (MRLC) phosphorylation that is carried out by Rho-associated protein kinase (ROCK), and that aPKCζ is required for EGF-dependent phosphorylation and inhibition of the myosin phosphatase targeting subunit (MYPT). Finally, we show that aPKCζ mediates the spatial organization of the acto-NMII cytoskeleton in response to EGF stimulation. Our data suggest that aPKCζ is an essential component regulator of acto-NMII cytoskeleton organization leading to directed cell migration, and is a mediator of the EGF signal to the cytoskeleton. PMID:27541056

Comparative Plasmodium gene overexpression reveals distinct perturbation of sporozoite transmission by profilin

PubMed Central

Sato, Yuko; Hliscs, Marion; Dunst, Josefine; Goosmann, Christian; Brinkmann, Volker; Montagna, Georgina N.; Matuschewski, Kai

2016-01-01

Plasmodium relies on actin-based motility to migrate from the site of infection and invade target cells. Using a substrate-dependent gliding locomotion, sporozoites are able to move at fast speed (1–3 μm/s). This motility relies on a minimal set of actin regulatory proteins and occurs in the absence of detectable filamentous actin (F-actin). Here we report an overexpression strategy to investigate whether perturbations of F-actin steady-state levels affect gliding locomotion and host invasion. We selected two vital Plasmodium berghei G-actin–binding proteins, C-CAP and profilin, in combination with three stage-specific promoters and mapped the phenotypes afforded by overexpression in all three extracellular motile stages. We show that in merozoites and ookinetes, additional expression does not impair life cycle progression. In marked contrast, overexpression of C-CAP and profilin in sporozoites impairs circular gliding motility and salivary gland invasion. The propensity for productive motility correlates with actin accumulation at the parasite tip, as revealed by combinations of an actin-stabilizing drug and transgenic parasites. Strong expression of profilin, but not C-CAP, resulted in complete life cycle arrest. Comparative overexpression is an alternative experimental genetic strategy to study essential genes and reveals effects of regulatory imbalances that are not uncovered from deletion-mutant phenotyping. PMID:27226484

Physical models of collective cell motility: from cell to tissue

NASA Astrophysics Data System (ADS)

Camley, B. A.; Rappel, W.-J.

2017-03-01

In this article, we review physics-based models of collective cell motility. We discuss a range of techniques at different scales, ranging from models that represent cells as simple self-propelled particles to phase field models that can represent a cell’s shape and dynamics in great detail. We also extensively review the ways in which cells within a tissue choose their direction, the statistics of cell motion, and some simple examples of how cell-cell signaling can interact with collective cell motility. This review also covers in more detail selected recent works on collective cell motion of small numbers of cells on micropatterns, in wound healing, and the chemotaxis of clusters of cells.

Adenovirus‑mediated overexpression of cystic fibrosis transmembrane conductance regulator enhances invasiveness and motility of serous ovarian cancer cells.

PubMed

Xu, Jiao; Lin, Liangbo; Yong, Min; Dong, Xiaojing; Yu, Tinghe; Hu, Lina

2016-01-01

The cystic fibrosis transmembrane conductance regulator (CFTR) belongs to the adenosine triphosphate‑binding cassette transporter family, members of which are involved in several types of cancer. Previous studies by our group reported that CFTR was highly expressed in serous ovarian cancer (SOC) tissues, and that knockdown of CFTR suppressed the proliferation of ovarian cancer in vitro and in vivo. Thus, the aim of the present study was to construct a recombinant adenoviral vector for the expression of the human CFTR gene in order to study the role of CFTR overexpression in the malignant invasion and migration of SOC cells in vitro. The present study then focused on the mechanisms of the role of CFTR in the migratory and invasive malignant properties of SOC cells. The CFTR gene was inserted into an adenoviral vector by using the AdEasy system in order to obtain the Ad‑CFTR overexpression vector, which was used to transfect the A2780 SOC cell line. Reverse-transcription polymerase chain reaction, western blot analysis and immunofluorescence were performed to detect the expression and localization of CFTR. Cell invasion and motility of the transfected cells compared with those of control cells were observed using Transwell and wound healing assays. A ~4,700 bp fragment of the CFTR gene was confirmed to be correctly cloned in the adenoviral vector and amplification of Ad‑CFTR was observed in HEK293 cells during package. After 48 h of transfection with Ad‑CFTR, ~90% of A2780 cells were red fluorescence protein‑positive. Immunofluorescence showed that following transfection, CFTR expression was increased and CFTR was located in the cell membrane and cytoplasm. CFTR overexpression was shown to enhance the invasion and motility of A2780 cells in vitro. Furthermore, the effects of CFTR overexpression on the activation c‑Src signaling were observed by western blot analysis. CFTR overexpressing cells showed the lowest activity of phospho‑Src (Tyr530), suggesting that CFTR may affect the activation of c‑Src signaling. The results of the present study demonstrated that adenovirus‑mediated CFTR overexpression enhanced cell invasion and motility of SOC cells in vitro. Furthermore, CFTR may be critical for the activation of c‑Src signaling.

DJ-1 KNOCK-DOWN IMPAIRS ASTROCYTE MITOCHONDRIAL FUNCTION

PubMed Central

LARSEN, N. J.; AMBROSI, G.; MULLETT, S. J.; BERMAN, S. B.; HINKLE, D. A.

2012-01-01

Mitochondrial dysfunction has long been implicated in the pathogenesis of Parkinson’s disease (PD). PD brain tissues show evidence for mitochondrial respiratory chain Complex I deficiency. Pharmacological inhibitors of Complex I, such as rotenone, cause experimental parkinsonism. The cytoprotective protein DJ-1, whose deletion is sufficient to cause genetic PD, is also known to have mitochondria-stabilizing properties. We have previously shown that DJ-1 is over-expressed in PD astrocytes, and that DJ-1 deficiency impairs the capacity of astrocytes to protect co-cultured neurons against rotenone. Since DJ-1 modulated, astrocyte-mediated neuroprotection against rotenone may depend upon proper astrocytic mitochondrial functioning, we hypothesized that DJ-1 deficiency would impair astrocyte mitochondrial motility, fission/fusion dynamics, membrane potential maintenance, and respiration, both at baseline and as an enhancement of rotenone-induced mitochondrial dysfunction. In astrocyte-enriched cultures, we observed that DJ-1 knock-down reduced mitochondrial motility primarily in the cellular processes of both untreated and rotenone treated cells. In these same cultures, DJ-1 knock-down did not appreciably affect mitochondrial fission, fusion, or respiration, but did enhance rotenone-induced reductions in the mitochondrial membrane potential. In neuron–astrocyte co-cultures, astrocytic DJ-1 knock-down reduced astrocyte process mitochondrial motility in untreated cells, but this effect was not maintained in the presence of rotenone. In the same co-cultures, astrocytic DJ-1 knock-down significantly reduced mitochondrial fusion in the astrocyte cell bodies, but not the processes, under the same conditions of rotenone treatment in which DJ-1 deficiency is known to impair astrocyte-mediated neuroprotection. Our studies therefore demonstrated the following new findings: (i) DJ-1 deficiency can impair astrocyte mitochondrial physiology at multiple levels, (ii) astrocyte mitochondrial dynamics vary with sub-cellular region, and (iii) the physical presence of neurons can affect astrocyte mitochondrial behavior. PMID:21907265

Optical trapping reveals propulsion forces, power generation and motility efficiency of the unicellular parasites Trypanosoma brucei brucei

PubMed Central

Stellamanns, Eric; Uppaluri, Sravanti; Hochstetter, Axel; Heddergott, Niko; Engstler, Markus; Pfohl, Thomas

2014-01-01

Unicellular parasites have developed sophisticated swimming mechanisms to survive in a wide range of environments. Cell motility of African trypanosomes, parasites responsible for fatal illness in humans and animals, is crucial both in the insect vector and the mammalian host. Using millisecond-scale imaging in a microfluidics platform along with a custom made optical trap, we are able to confine single cells to study trypanosome motility. From the trapping characteristics of the cells, we determine the propulsion force generated by cells with a single flagellum as well as of dividing trypanosomes with two fully developed flagella. Estimates of the dissipative energy and the power generation of single cells obtained from the motility patterns of the trypanosomes within the optical trap indicate that specific motility characteristics, in addition to locomotion, may be required for antibody clearance. Introducing a steerable second optical trap we could further measure the force, which is generated at the flagellar tip. Differences in the cellular structure of the trypanosomes are correlated with the trapping and motility characteristics and in consequence with their propulsion force, dissipative energy and power generation. PMID:25269514

CD28-CD80 interactions control regulatory T cell motility and immunological synapse formation1,2

PubMed Central

Thauland, Timothy J.; Koguchi, Yoshinobu; Dustin, Michael L.; Parker, David C.

2014-01-01

Regulatory T cells (Tregs) are essential for tolerance to self and environmental antigens, acting in part by downmodulating costimulatory molecules on the surface of dendritic cells (DCs) and altering naïve CD4 T cell-DC interactions. Here, we show that Tregs form stable conjugates with DCs before, but not after, they decrease surface expression of the costimulatory molecule CD80 on the DCs. We use supported planar bilayers to show that Tregs dramatically slow down, but maintain a highly polarized and motile phenotype after recognizing antigen in the absence of costimulation. These motile cells are characterized by distinct accumulations of LFA-1-ICAM-1 in the lamella and TCR-MHC in the uropod, consistent with a motile immunological synapse or ‘kinapse’. However, in the presence of high, but not low, concentrations of CD80, Tregs form stationary, symmetrical synapses. Using blocking antibodies, we show that, while CTLA-4 is required for CD80 downmodulation, CD28-CD80 interactions are critical for modulating Treg motility in the presence of antigen. Together, these results support the hypothesis that Tregs are tuned to alter their motility depending on costimulatory signals. PMID:25355918

Involvement of Mismatch Repair in the Reciprocal Control of Motility and Adherence of Uropathogenic Escherichia coli

PubMed Central

Cooper, Lauren A.; Simmons, Lyle A.

2012-01-01

Type 1 fimbriae and flagella, two surface organelles critical for colonization of the urinary tract by uropathogenic Escherichia coli (UPEC), mediate opposing virulence objectives. Type 1 fimbriae facilitate adhesion to mucosal cells and promote bacterial persistence in the urinary tract, while flagella propel bacteria through urine and along mucous layers during ascension to the upper urinary tract. Using a transposon screen of the E. coli CFT073 fim locked-ON (L-ON) mutant, a construct that constitutively expresses type 1 fimbriae and represses motility, we identified six mutants that exhibited a partial restoration of motility. Among these six mutated genes was mutS, which encodes a component of the methyl-directed mismatch repair (MMR) system. When complemented with mutS in trans, motility was again repressed. To determine whether the MMR system, in general, is involved in this reciprocal control, we characterized the effects of gene deletions of other MMR components on UPEC motility. Isogenic deletions of mutS, mutH, and mutL were constructed in both wild-type CFT073 and fim L-ON backgrounds. All MMR mutants showed an increase in motility in the wild-type background, and ΔmutH and ΔmutS mutations increased motility in the fim L-ON background. Cochallenge of the wild-type strain with an MMR-defective strain showed a subtle but significant competitive advantage in the bladder and spleen for the MMR mutant using the murine model of ascending urinary tract infection after 48 h. Our findings demonstrate that the MMR system generally affects the reciprocal regulation of motility and adherence and thus could contribute to UPEC pathogenesis during urinary tract infections. PMID:22473602

Oxidative stress negatively affects human sperm mitochondrial respiration.

PubMed

Ferramosca, Alessandra; Pinto Provenzano, Sara; Montagna, Daniela Domenica; Coppola, Lamberto; Zara, Vincenzo

2013-07-01

To correlate the level of oxidative stress in serum and seminal fluid and the level of sperm deoxyribonucleic acid (DNA) fragmentation with sperm mitochondrial respiratory efficiency. Sperm mitochondrial respiratory activity was evaluated with a polarographic assay of oxygen consumption carried out in hypotonically treated sperm cells. A possible relationship between sperm mitochondrial respiratory efficiency, the level of oxidative stress, and the level of sperm DNA fragmentation was investigated. Sperm motility was positively correlated with mitochondrial respiration but negatively correlated with oxidative stress and DNA fragmentation. Interestingly, sperm mitochondrial respiratory activity was negatively affected by oxidative stress and DNA fragmentation. Our data indicate that sperm mitochondrial respiration is decreased in patients with high levels of reactive oxygen species by an uncoupling between electron transport and adenosine triphosphate synthesis. This reduction in mitochondrial functionality might be 1 of the reasons responsible for the decrease in spermatozoa motility. Copyright © 2013 Elsevier Inc. All rights reserved.

Cortactin as a Target for FAK in the Regulation of Focal Adhesion Dynamics

PubMed Central

Ghassemian, Majid; Schlaepfer, David D.

2012-01-01

Background Efficient cell movement requires the dynamic regulation of focal adhesion (FA) formation and turnover. FAs are integrin-associated sites of cell attachment and establish linkages to the cellular actin cytoskeleton. Cells without focal adhesion kinase (FAK), an integrin-activated tyrosine kinase, exhibit defects in FA turnover and cell motility. Cortactin is an actin binding adaptor protein that can influence FA dynamics. FAK and cortactin interact, but the cellular role of this complex remains unclear. Principal Findings Using FAK-null fibroblasts stably reconstituted with green fluorescent protein (GFP) tagged FAK constructs, we find that FAK activity and FAK C-terminal proline-rich region 2 (PRR2) and PRR3 are required for FA turnover and cell motility. Cortactin binds directly to FAK PRR2 and PRR3 sites via its SH3 domain and cortactin expression is important in promoting FA turnover and GFP-FAK release from FAs. FAK-cortactin binding is negatively-regulated by FAK activity and associated with cortactin tyrosine phosphorylation. FAK directly phosphorylates cortactin at Y421 and Y466 and over-expression of cortactin Y421, Y466, and Y482 mutated to phenylalanine (3YF) prevented FAK-enhanced FA turnover and cell motility. However, phospho-mimetic cortactin mutated to glutamic acid (3YE) did not affect FA dynamics and did not rescue FA turnover defects in cells with inhibited FAK activity or with PRR2-mutated FAK that does not bind cortactin. Conclusions Our results support a model whereby FAK-mediated FA remodeling may occur through the formation of a FAK-cortactin signaling complex. This involves a cycle of cortactin binding to FAK, cortactin tyrosine phosphorylation, and subsequent cortactin-FAK dissociation accompanied by FA turnover and cell movement. PMID:22952866

Bacterial cell motility of Burkholderia gut symbiont is required to colonize the insect gut.

PubMed

Lee, Jun Beom; Byeon, Jin Hee; Jang, Ho Am; Kim, Jiyeun Kate; Yoo, Jin Wook; Kikuchi, Yoshitomo; Lee, Bok Luel

2015-09-14

We generated a Burkholderia mutant, which is deficient of an N-acetylmuramyl-l-alanine amidase, AmiC, involved in peptidoglycan degradation. When non-motile ΔamiC mutant Burkholderia cells harboring chain form were orally administered to Riptortus insects, ΔamiC mutant cells were unable to establish symbiotic association. But, ΔamiC mutant complemented with amiC gene restored in vivo symbiotic association. ΔamiC mutant cultured in minimal medium restored their motility with single-celled morphology. When ΔamiC mutant cells harboring single-celled morphology were administered to the host insect, this mutant established normal symbiotic association, suggesting that bacterial motility is essential for the successful symbiosis between host insect and Burkholderia symbiont. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Multiple scale model for cell migration in monolayers: Elastic mismatch between cells enhances motility

NASA Astrophysics Data System (ADS)

Palmieri, Benoit; Bresler, Yony; Wirtz, Denis; Grant, Martin

2015-07-01

We propose a multiscale model for monolayer of motile cells that comprise normal and cancer cells. In the model, the two types of cells have identical properties except for their elasticity; cancer cells are softer and normal cells are stiffer. The goal is to isolate the role of elasticity mismatch on the migration potential of cancer cells in the absence of other contributions that are present in real cells. The methodology is based on a phase-field description where each cell is modeled as a highly-deformable self-propelled droplet. We simulated two types of nearly confluent monolayers. One contains a single cancer cell in a layer of normal cells and the other contains normal cells only. The simulation results demonstrate that elasticity mismatch alone is sufficient to increase the motility of the cancer cell significantly. Further, the trajectory of the cancer cell is decorated by several speed “bursts” where the cancer cell quickly relaxes from a largely deformed shape and consequently increases its translational motion. The increased motility and the amplitude and frequency of the bursts are in qualitative agreement with recent experiments.

Functional and proteomic analyses reveal that wxcB is involved in virulence, motility, detergent tolerance, and biofilm formation in Xanthomonas campestris pv. vesicatoria.

PubMed

Park, Hye-Jee; Jung, Ho Won; Han, Sang-Wook

2014-09-26

The bacterial envelope possesses diverse functions, including protection against environmental stress and virulence factors for host infection. Here, we report the function of wxcB in Xanthomonas campestris pv. vesicatoria (Xcv), a causal agent of bacterial leaf spot disease in tomato and pepper. To characterize roles of wxcB, we generated a knockout mutant (XcvΔwxcB) and found that the virulence of the mutant was weaker than that of the wild type in tomato plants. To predict the mechanism affected by wxcB, we compared protein expressions between the wild type and the mutant. Expression of 152 proteins showed a greater than 2-fold difference. Proteins involved in motility and cell wall/membrane were the most abundant. Through phenotypic assays, we further demonstrated that the mutant displayed reduced motility and tolerance to treatment, but it showed increased biofilm formation. Interestingly, the LPS profile was unchanged. These results lead to new insights into the functions of wxcB that is associated with cell wall/membrane functions, which contributes to pathogen virulence. Copyright © 2014 Elsevier Inc. All rights reserved.

Effects of number and configuration of flagella on motility of Helicobacter species.

NASA Astrophysics Data System (ADS)

Constantino, Maira A.; Sharba, Sinan; Shen, Zeli; Fox, James G.; Haesebrouck, Freddy; Linden, Sara; Bansil, Rama

Helicobacters are ulcer-causing bacteria that colonize the viscoelastic gastric mucus layer of mammals. Previous studies have shown that motility and colonization are affected by helical body shape, number and configuration of flagella. In a recent study, using fast time-resolution and high-magnification 2-D phase-contrast microscopy to image individual helical and rod-shaped H. pylori we measured the rotation rate of the cell body and flagella and found that helical shape produces less than 15% changes in swimming speeds as compared to the rod-shaped cell. Motility of H. pylori was strongly influenced by its multiple unipolar flagella. Here we compare rotational and translational speeds of H. cetorum and H. suis which have bipolar flagella, with H. cetorum having single bipolar flagella and H. suis having multiple flagella. Preliminary results show that H. suis bacteria swim slower but rotate at the same rate as H. pylori and present two swimming modes. It can swim as a pusher, with one active rotating bundle and one inactive bundle, wrapped around the body or with both bundles active. Similar work on H. cetorum is ongoing and will also be presented. NSF PHY 1410798.

Eukaryotic Chemotaxis: A Network of Signaling Pathways Controls Motility, Directional Sensing, and Polarity

PubMed Central

Swaney, Kristen F.; Huang, Chuan-Hsiang; Devreotes, Peter N.

2015-01-01

Chemotaxis, the directed migration of cells in chemical gradients, is a vital process in normal physiology and in the pathogenesis of many diseases. Chemotactic cells display motility, directional sensing, and polarity. Motility refers to the random extension of pseudopodia, which may be driven by spontaneous actin waves that propagate through the cytoskeleton. Directional sensing is mediated by a system that detects temporal and spatial stimuli and biases motility toward the gradient. Polarity gives cells morphologically and functionally distinct leading and lagging edges by relocating proteins or their activities selectively to the poles. By exploiting the genetic advantages of Dictyostelium, investigators are working out the complex network of interactions between the proteins that have been implicated in the chemotactic processes of motility, directional sensing, and polarity. PMID:20192768

The Aeromonas caviae AHA0618 gene modulates cell length and influences swimming and swarming motility

PubMed Central

Lowry, Rebecca C; Parker, Jennifer L; Kumbhar, Ramhari; Mesnage, Stephane; Shaw, Jonathan G; Stafford, Graham P

2015-01-01

Aeromonas caviae is motile via a polar flagellum in liquid culture, with a lateral flagella system used for swarming on solid surfaces. The polar flagellum also has a role in cellular adherence and biofilm formation. The two subunits of the polar flagellum, FlaA and FlaB, are posttranslationally modified by O-linked glycosylation with pseudaminic acid on 6–8 serine and threonine residues within the central region of these proteins. This modification is essential for the formation of the flagellum. Aeromonas caviae possesses the simplest set of genes required for bacterial glycosylation currently known, with the putative glycosyltransferase, Maf1, being described recently. Here, we investigated the role of the AHA0618 gene, which shares homology (37% at the amino acid level) with the central region of a putative deglycosylation enzyme (HP0518) from the human pathogen Helicobacter pylori, which also glycosylates its flagellin and is proposed to be part of a flagellin deglycosylation pathway. Phenotypic analysis of an AHA0618 A. caviae mutant revealed increased swimming and swarming motility compared to the wild-type strain but without any detectable effects on the glycosylation status of the polar flagellins when analyzed by western blot analysis or mass spectroscopy. Bioinformatic analysis of the protein AHA0618, demonstrated homology to a family of l,d-transpeptidases involved in cell wall biology and peptidoglycan cross-linking (YkuD-like). Scanning electron microscopy (SEM) and fluorescence microscopy analysis of the wild-type and AHA0618-mutant A. caviae strains revealed the mutant to be subtly but significantly shorter than wild-type cells; a phenomenon that could be recovered when either AHA0618 or H. pylori HP0518 were introduced. We can therefore conclude that AHA0618 does not affect A. caviae behavior by altering polar flagellin glycosylation levels but is likely to have a role in peptidoglycan processing at the bacterial cell wall, consequently altering cell length and hence influencing motility. PMID:25515520

Delta opioid receptor on equine sperm cells: subcellular localization and involvement in sperm motility analyzed by computer assisted sperm analyzer (CASA)

PubMed Central

2010-01-01

Background Opioid receptors and endogenous opioid peptides act not only in the control of nociceptive pathways, indeed several reports demonstrate the effects of opiates on sperm cell motility and morphology suggesting the importance of these receptors in the modulation of reproduction in mammals. In this study we investigated the expression of delta opioid receptors on equine spermatozoa by western blot/indirect immunofluorescence and its relationship with sperm cell physiology. Methods We analyzed viability, motility, capacitation, acrosome reaction and mitochondrial activity in the presence of naltrindole and DPDPE by means of a computer assisted sperm analyzer and a fluorescent confocal microscope. The evaluation of viability, capacitation and acrosome reaction was carried out by the double CTC/Hoechst staining, whereas mitochondrial activity was assessed by means of MitoTracker Orange dye. Results We showed that in equine sperm cells, delta opioid receptor is expressed as a doublet of 65 and 50 kDa molecular mass and is localized in the mid piece of tail; we also demonstrated that naltrindole, a delta opioid receptor antagonist, could be utilized in modulating several physiological parameters of the equine spermatozoon in a dose-dependent way. We also found that low concentrations of the antagonist increase sperm motility whereas high concentrations show the opposite effect. Moreover low concentrations hamper capacitation, acrosome reaction and viability even if the percentage of cells with active mitochondria seems to be increased; the opposite effect is exerted at high concentrations. We have also observed that the delta opioid receptor agonist DPDPE is scarcely involved in affecting the same parameters at the employed concentrations. Conclusions The results described in this paper add new important details in the comprehension of the mammalian sperm physiology and suggest new insights for improving reproduction and for optimizing equine breeding. PMID:20579355

A light-regulated type I pilus contributes to Acinetobacter baumannii biofilm, motility and virulence functions.

PubMed

Wood, Cecily R; Ohneck, Emily J; Edelmann, Richard E; Actis, Luis A

2018-06-11

Transcriptional analyses of A. baumannii ATCC 17978 showed that the expression of A1S_2091 was enhanced in cells cultured in darkness at 24°C through a process that depended on the BlsA photoreceptor. Disruption of A1S_2091, a component of the A1S_2088-A1S_2091 polycistronic operon predicted to code for a type I chaperone/usher pilus assembly system, abolished surface motility and pellicle formation but significantly enhanced biofilm formation on plastic by bacteria cultured under darkness. Based on these observations, the A1S_2088-A1S_2091 operon was named the p hoto- r egulated p ilus ABCD ( prpABCD ) operon, with A1S_2091 coding for the PrpA pilin subunit. Unexpectedly, the comparative analyses of ATCC 17978 and prpA isogenic mutant cells cultured at 37°C showed the expression of light-regulated biofilm biogenesis and motility functions under a temperature condition that drastically affects BlsA production and its light sensing activity. These assays also suggest that 17978 cells produce alternative light-regulated adhesins and/or pili systems that enhance bacterial adhesion and biofilm formation both at 24°C and 37°C on plastic as well as on the surface of polarized A549 alveolar epithelial cells, where formation of bacterial filaments and cell chains was significantly enhanced. The inactivation of prpA also resulted in a significant reduction in virulence when tested using the Galleria mellonella virulence model. All these observations provide strong evidence showing the capacity of A. baumannii to sense light and interact with biotic and abiotic surfaces using undetermined alternative sensing and regulatory systems as well as alternative adherence and motility cellular functions that allow this pathogen to persist in different ecological niches. Copyright © 2018 American Society for Microbiology.

Paramecium swimming and ciliary beating patterns: a study on four RNA interference mutations.

PubMed

Funfak, Anette; Fisch, Cathy; Abdel Motaal, Hatem T; Diener, Julien; Combettes, Laurent; Baroud, Charles N; Dupuis-Williams, Pascale

2015-01-01

Paramecium cells swim and feed by beating their thousands of cilia in coordinated patterns. The organization of these patterns and its relationship with cell motility has been the subject of a large body of work, particularly as a model for ciliary beating in human organs where similar organization is seen. However the rapid motion of the cells makes quantitative measurements very challenging. Here we provide detailed measurements of the swimming of Paramecium cells from high-speed video at high magnification, as they move in microfluidic channels. An image analysis protocol allows us to decouple the cell movement from the motion of the cilia, thus allowing us to measure the ciliary beat frequency (CBF) and the spatio-temporal organization into metachronal waves along the cell periphery. Two distinct values of the CBF appear at different regions of the cell: most of the cilia beat in the range of 15 to 45 Hz, while the cilia in the peristomal region beat at almost double the frequency. The body and peristomal CBF display a nearly linear relation with the swimming velocity. Moreover the measurements do not display a measurable correlation between the swimming velocity and the metachronal wave velocity on the cell periphery. These measurements are repeated for four RNAi silenced mutants, where proteins specific to the cilia or to their connection to the cell base are depleted. We find that the mutants whose ciliary structure is affected display similar swimming to the control cells albeit with a reduced efficiency, while the mutations that affect the cilia's anchoring to the cell lead to strongly reduced ability to swim. This reduction in motility can be related to a loss of coordination between the ciliary beating in different parts of the cell.

ALDH isozymes downregulation affects cell growth, cell motility and gene expression in lung cancer cells.

PubMed

Moreb, Jan S; Baker, Henry V; Chang, Lung-Ji; Amaya, Maria; Lopez, M Cecilia; Ostmark, Blanca; Chou, Wayne

2008-11-24

Aldehyde dehydrogenase isozymes ALDH1A1 and ALDH3A1 are highly expressed in non small cell lung cancer. Neither the mechanisms nor the biologic significance for such over expression have been studied. We have employed oligonucleotide microarrays to analyze changes in gene profiles in A549 lung cancer cell line in which ALDH activity was reduced by up to 95% using lentiviral mediated expression of siRNA against both isozymes (Lenti 1+3). Stringent analysis methods were used to identify gene expression patterns that are specific to the knock down of ALDH activity and significantly different in comparison to wild type A549 cells (WT) or cells similarly transduced with green fluorescent protein (GFP) siRNA. We confirmed significant and specific down regulation of ALDH1A1 and ALDH3A1 in Lenti 1+3 cells and in comparison to 12 other ALDH genes detected. The results of the microarray analysis were validated by real time RT-PCR on RNA obtained from Lenti 1+3 or WT cells treated with ALDH activity inhibitors. Detailed functional analysis was performed on 101 genes that were significantly different (P < 0.001) and their expression changed by > or = 2 folds in the Lenti 1+3 group versus the control groups. There were 75 down regulated and 26 up regulated genes. Protein binding, organ development, signal transduction, transcription, lipid metabolism, and cell migration and adhesion were among the most affected pathways. These molecular effects of the ALDH knock-down are associated with in vitro functional changes in the proliferation and motility of these cells and demonstrate the significance of ALDH enzymes in cell homeostasis with a potentially significant impact on the treatment of lung cancer.

Uncovering the Mystery of Gliding Motility in the Myxobacteria

PubMed Central

Nan, Beiyan; Zusman, David R.

2012-01-01

Bacterial gliding motility is the smooth movement of cells on solid surfaces unaided by flagella or pili. Many diverse groups of bacteria exhibit gliding, but the mechanism of gliding motility has remained a mystery since it was first observed more than a century ago. Recent studies on the motility of Myxococcus xanthus, a soil myxobacterium, suggest a likely mechanism for gliding in this organism. About forty M. xanthus genes were shown to be involved in gliding motility, and some of their protein products were labeled and localized within cells. These studies suggest that gliding motility in M. xanthus involves large multiprotein structural complexes, regulatory proteins, and cytoskeletal filaments. In this review, we summarize recent experiments that provide the basis for this emerging view of M. xanthus motility. We also discuss alternative models for gliding. PMID:21910630

In Azospirillum brasilense, mutations in flmA or flmB genes affect polar flagellum assembly, surface polysaccharides, and attachment to maize roots.

PubMed

Rossi, Fernando Ariel; Medeot, Daniela Beatriz; Liaudat, Juan Pablo; Pistorio, Mariano; Jofré, Edgardo

2016-09-01

Azospirillum brasilense is a soil bacterium capable of promoting plant growth. Several surface components were previously reported to be involved in the attachment of A. brasilense to root plants. Among these components are the exopolysaccharide (EPS), lipopolysaccharide (LPS) and the polar flagellum. Flagellin from polar flagellum is glycosylated and it was suggested that genes involved in such a posttranslational modification are the same ones involved in the biosynthesis of sugars present in the O-antigen of the LPS. In this work, we report on the characterization of two homologs present in A. brasilense Cd, to the well characterized flagellin modification genes, flmA and flmB, from Aeromonas caviae. We show that mutations in either flmA or flmB genes of A. brasilense resulted in non-motile cells due to alterations in the polar flagellum assembly. Moreover, these mutations also affected the capability of A. brasilense cells to adsorb to maize roots and to produce LPS and EPS. By generating a mutant containing the polar flagellum affected in their rotation, we show the importance of the bacterial motility for the early colonization of maize roots. Copyright © 2016 Elsevier GmbH. All rights reserved.

Hydrodynamic effects on microcapillary motility and chemotaxis assays of Methylosinus trichosporium OB3b.

PubMed Central

Shonnard, D R; Taylor, R T; Tompson, A; Knapp, R B

1992-01-01

A study of the random motility and chemotaxis of Methylosinus trichosporium OB3b was conducted by using Palleroni-chamber microcapillary assay procedures. Under the growth conditions employed, this methanotroph was observed qualitatively with a microscope to be either slightly motile or essentially nonmotile. However, the cells did not not respond in the microcapillary assays in the manner expected for nonmotile Brownian particles. As a consequence, several hydrodynamic effects on these Palleroni microcapillary assays were uncovered. In the random-motility microcapillary assay, nondiffusive cell accumulations occurred that were strongly dependent upon cell concentration. An apparent minimal random-motility coefficient (mu) for this bacterial cell of 1.0 x 10(-7) cm2/s was estimated from microcapillary assays. A simple alternative spectrophotometric assay, based upon gravitational settling, was developed and shown to be an improvement over the Palleroni microcapillary motility assay for M. trichosporium OB3b in that it yielded a more-accurate threefold-lower random-motility coefficient. In addition, it provided a calculation of the gravitational-settling velocity. In the chemotaxis microcapillary assay, the apparent chemotactic responses were strongest for the highest test-chemical concentrations in the microcapillaries, were correlated with microcapillary fluid density, and were strongly dependent upon the microcapillary volume. A simple method to establish the maximal concentration of a chemical that can be tested and to quantify any contributions of abiotic convection is described. Investigators should be aware of the potential problems due to density-driven convection when using these commonly employed microcapillary assays for studying cells which have low motilities. PMID:1444383

Quantifying the roles of random motility and directed motility using advection-diffusion theory for a 3T3 fibroblast cell migration assay stimulated with an electric field.

PubMed

Simpson, Matthew J; Lo, Kai-Yin; Sun, Yung-Shin

2017-03-17

Directed cell migration can be driven by a range of external stimuli, such as spatial gradients of: chemical signals (chemotaxis); adhesion sites (haptotaxis); or temperature (thermotaxis). Continuum models of cell migration typically include a diffusion term to capture the undirected component of cell motility and an advection term to capture the directed component of cell motility. However, there is no consensus in the literature about the form that the advection term takes. Some theoretical studies suggest that the advection term ought to include receptor saturation effects. However, others adopt a much simpler constant coefficient. One of the limitations of including receptor saturation effects is that it introduces several additional unknown parameters into the model. Therefore, a relevant research question is to investigate whether directed cell migration is best described by a simple constant tactic coefficient or a more complicated model incorporating saturation effects. We study directed cell migration using an experimental device in which the directed component of the cell motility is driven by a spatial gradient of electric potential, which is known as electrotaxis. The electric field (EF) is proportional to the spatial gradient of the electric potential. The spatial variation of electric potential across the experimental device varies in such a way that there are several subregions on the device in which the EF takes on different values that are approximately constant within those subregions. We use cell trajectory data to quantify the motion of 3T3 fibroblast cells at different locations on the device to examine how different values of the EF influences cell motility. The undirected (random) motility of the cells is quantified in terms of the cell diffusivity, D, and the directed motility is quantified in terms of a cell drift velocity, v. Estimates D and v are obtained under a range of four different EF conditions, which correspond to normal physiological conditions. Our results suggest that there is no anisotropy in D, and that D appears to be approximately independent of the EF and the electric potential. The drift velocity increases approximately linearly with the EF, suggesting that the simplest linear advection term, with no additional saturation parameters, provides a good explanation of these physiologically relevant data. We find that the simplest linear advection term in a continuum model of directed cell motility is sufficient to describe a range of different electrotaxis experiments for 3T3 fibroblast cells subject to normal physiological values of the electric field. This is useful information because alternative models that include saturation effects involve additional parameters that need to be estimated before a partial differential equation model can be applied to interpret or predict a cell migration experiment.

Intermittent Ca2+ signals mediated by Orai1 regulate basal T cell motility

PubMed Central

Greenberg, Milton L; Jairaman, Amit; Akunwafo, Chijioke; Leverrier, Sabrina; Yu, Ying; Parker, Ian; Dynes, Joseph L

2017-01-01

Ca2+ influx through Orai1 channels is crucial for several T cell functions, but a role in regulating basal cellular motility has not been described. Here, we show that inhibition of Orai1 channel activity increases average cell velocities by reducing the frequency of pauses in human T cells migrating through confined spaces, even in the absence of extrinsic cell contacts or antigen recognition. Utilizing a novel ratiometric genetically encoded cytosolic Ca2+ indicator, Salsa6f, which permits real-time monitoring of cytosolic Ca2+ along with cell motility, we show that spontaneous pauses during T cell motility in vitro and in vivo coincide with episodes of cytosolic Ca2+ signaling. Furthermore, lymph node T cells exhibited two types of spontaneous Ca2+ transients: short-duration ‘sparkles’ and longer duration global signals. Our results demonstrate that spontaneous and self-peptide MHC-dependent activation of Orai1 ensures random walk behavior in T cells to optimize immune surveillance. PMID:29239723

Surface association and the MreB cytoskeleton regulate pilus production, localization and function in Pseudomonas aeruginosa.

PubMed

Cowles, Kimberly N; Gitai, Zemer

2010-06-01

Spatial organization of bacterial proteins influences many cellular processes, including division, chromosome segregation and motility. Virulence-associated proteins also localize to specific destinations within bacterial cells. However, the functions and mechanisms of virulence factor localization remain largely unknown. In this work, we demonstrate that polar assembly of the Pseudomonas aeruginosa PAO1 type IV pilus is regulated by surface association in a manner that affects gene transcription, protein levels and protein localization. We also uncover one mechanism for this regulation that acts through the actin homologue MreB. Inactivation of MreB leads to mislocalization of the pilus retraction ATPase PilT, mislocalization of the pili themselves and a reduction in motility. Furthermore, the role of MreB in polar localization of PilT is modulated by surface association, corroborating our results that environmental factors influence the regulation of pilus production. Specifically, MreB mediates both the initiation and maintenance of PilT localization when cells are grown in suspension but only affects the initiation of localization when cells are grown on a surface. Together, these results suggest that the bacterial cytoskeleton provides a mechanism for the polar localization of P. aeruginosa pili and demonstrate that protein localization may represent an important aspect of virulence factor regulation in bacterial pathogens.

The beneficial effect of genetically engineered Schwann cells with enhanced motility in peripheral nerve regeneration: review.

PubMed

Gravvanis, A I; Lavdas, A A; Papalois, A; Tsoutsos, D A; Matsas, R

2007-01-01

The importance of Schwann cells in promoting nerve regeneration across a conduit has been extensively reported in the literature, and Schwann cell motility has been acknowledged as a prerequisite for myelination of the peripheral nervous system during regeneration after injury. Review of recent literature and retrospective analysis of our studies with genetically modified Schwann Cells with increased motility in order to identify the underlying mechanism of action and outline the future trends in peripheral nerve repair. Schwann cell transduction with the pREV-retrovirus, for expression of Sialyl-Transferase-X, resulting in conferring Polysialyl-residues (PSA) on NCAM, increases their motility in-vitro and ensures nerve regeneration through silicone tubes after end-to-side neurorraphy in the rat sciatic nerve model, thus significantly promoting fiber maturation and functional outcome. An artificial nerve graft consisting of a type I collagen tube lined with the genetically modified Schwann cells with increased motility, used to bridge a defect in end-to-end fashion in the rat sciatic nerve model, was shown to promote nerve regeneration to a level equal to that of a nerve autograft. The use of genetically engineered Schwann cells with enhanced motility for grafting endoneural tubes promotes axonal regeneration, by virtue of the interaction of the transplanted cells with regenerating axonal growth cones as well as via the recruitment of endogenous Schwann cells. It is envisaged that mixed populations of Schwann cells, expressing PSA and one or more trophic factors, might further enhance the regenerating and remyelinating potential of the lesioned nerves.

Interstitial flows promote an amoeboid cell phenotype and motility of breast cancer cells

NASA Astrophysics Data System (ADS)

Tung, Chih-Kuan; Huang, Yu Ling; Zheng, Angela; Wu, Mingming

2015-03-01

Lymph nodes, the drainage systems for interstitial flows, are clinically known to be the first metastatic sites of many cancer types including breast and prostate cancers. Here, we demonstrate that breast cancer cell morphology and motility is modulated by interstitial flows in a cell-ECM adhesion dependent manner. The average aspect ratios of the cells are significantly lower (or are more amoeboid like) in the presence of the flow in comparison to the case when the flow is absent. The addition of exogenous adhesion molecules within the extracellular matrix (type I collagen) enhances the overall aspect ratio (or are more mesenchymal like) of the cell population. Using measured cell trajectories, we find that the persistence of the amoeboid cells (aspect ratio less than 2.0) is shorter than that of mesenchymal cells. However, the maximum speed of the amoeboid cells is larger than that of mesenchymal cells. Together these findings provide the novel insight that interstitial flows promote amoeboid cell morphology and motility and highlight the plasticity of tumor cell motility in response to its biophysical environment. Supported by NIH Grant R21CA138366.

Flagellum Density Regulates Proteus mirabilis Swarmer Cell Motility in Viscous Environments

PubMed Central

Tuson, Hannah H.; Copeland, Matthew F.; Carey, Sonia; Sacotte, Ryan

2013-01-01

Proteus mirabilis is an opportunistic pathogen that is frequently associated with urinary tract infections. In the lab, P. mirabilis cells become long and multinucleate and increase their number of flagella as they colonize agar surfaces during swarming. Swarming has been implicated in pathogenesis; however, it is unclear how energetically costly changes in P. mirabilis cell morphology translate into an advantage for adapting to environmental changes. We investigated two morphological changes that occur during swarming—increases in cell length and flagellum density—and discovered that an increase in the surface density of flagella enabled cells to translate rapidly through fluids of increasing viscosity; in contrast, cell length had a small effect on motility. We found that swarm cells had a surface density of flagella that was ∼5 times larger than that of vegetative cells and were motile in fluids with a viscosity that inhibits vegetative cell motility. To test the relationship between flagellum density and velocity, we overexpressed FlhD4C2, the master regulator of the flagellar operon, in vegetative cells of P. mirabilis and found that increased flagellum density produced an increase in cell velocity. Our results establish a relationship between P. mirabilis flagellum density and cell motility in viscous environments that may be relevant to its adaptation during the infection of mammalian urinary tracts and movement in contact with indwelling catheters. PMID:23144253

An increase or a decrease in myosin II phosphorylation inhibits macrophage motility

PubMed Central

1991-01-01

Myosin II purified from mammalian non-muscle cells is phosphorylated on the 20-kD light chain subunit (MLC20) by the Ca2+/calmodulin-dependent enzyme myosin light chain kinase (MLCK). The importance of MLC20 phosphorylation in regulating cell motility was investigated by introducing either antibodies to MLCK (MK-Ab) or a Ca2+/calmodulin- independent, constitutively active form of MLCK (MK-) into macrophages. The effects of these proteins on cell motility were then determined using a quantitative chemotaxis assay. Chemotaxis is significantly diminished in macrophages containing MK-Ab compared to macrophages containing control antibodies. Moreover, there is an inverse relationship between the number of cells that migrate and the amount of MK-Ab introduced into cells. Interestingly, there is also an inverse relationship between the number of cells that migrate and the amount of MK- introduced into cells. Other experiments demonstrated that MK-Ab decreased intracellular MLC20 phosphorylation while MK- increased MLC20 phosphorylation. MK- also increased the amount of myosin associated with the cytoskeleton. These data demonstrate that the regulation of MLCK is an important aspect of cell motility and suggest that MLC20 phosphorylation must be maintained within narrow limits during translational motility by mammalian cells. PMID:2071674

The Interplay between Signaling and Metabolism in Breast Cancer Cell Motility and Metastasis

NASA Astrophysics Data System (ADS)

Tsarfaty, Ilan

2013-03-01

The initiation and growth of tumor metastases require tumor cells go through a transition between collective-to-individual cell migration. Understanding the molecular, cellular and physical mechanisms of these different migration modes is limited. We focus on the tumor cell migration induced by Hepatocyte Growth Factor / Scatter Factor (HGF/SF) - Met-signaling, a master regulator of cell motility in normal and malignant processes. Met has been implicated in tumorigenesis and metastasis and several Met targeting agents have been introduced into the clinic, and are currently in all phases of clinical trials Our analysis demonstrates that Met signaling dramatically alter the morpho-kinetic dynamics of collective migration of tumor cells. It induce a ``wave'' of increasing velocities that propagates back from the leading edge, increases cells' orientation and cooperation capabilities. In parallel Met signaling induces amoeboid cell motility that increased cell individuality. The decision making regarding the motility mode is dependent on the extent of activation of unique signal and metabolic cues. We present a combination of molecular imaging, conceptual and modeling framework for the analysis and assessment of the collective mesenchymal to epithelial versus amoeboid motility. Combined together our analysis can contribute to the understanding of metastasis and personalizing anti Met targeted therapy.

Microbial dispersal in unsaturated porous media: Characteristics of motile bacterial cell motions in unsaturated angular pore networks

NASA Astrophysics Data System (ADS)

Ebrahimi, Ali N.; Or, Dani

2014-09-01

The dispersal rates of self-propelled microorganisms affect their spatial interactions and the ecological functioning of microbial communities. Microbial dispersal rates affect risk of contamination of water resources by soil-borne pathogens, the inoculation of plant roots, or the rates of spoilage of food products. In contrast with the wealth of information on microbial dispersal in water replete systems, very little is known about their dispersal rates in unsaturated porous media. The fragmented aqueous phase occupying complex soil pore spaces suppress motility and limits dispersal ranges in unsaturated soil. The primary objective of this study was to systematically evaluate key factors that shape microbial dispersal in model unsaturated porous media to quantify effects of saturation, pore space geometry, and chemotaxis on characteristics of principles that govern motile microbial dispersion in unsaturated soil. We constructed a novel 3-D angular pore network model (PNM) to mimic aqueous pathways in soil for different hydration conditions; within the PNM, we employed an individual-based model that considers physiological and biophysical properties of motile and chemotactic bacteria. The effects of hydration conditions on first passage times in different pore networks were studied showing that fragmentation of aquatic habitats under dry conditions sharply suppresses nutrient transport and microbial dispersal rates in good agreement with limited experimental data. Chemotactically biased mean travel speed of microbial cells across 9 mm saturated PNM was ˜3 mm/h decreasing exponentially to 0.45 mm/h for the PNM at matric potential of -15 kPa (for -35 kPa, dispersal practically ceases and the mean travel time to traverse the 9 mm PNM exceeds 1 year). Results indicate that chemotaxis enhances dispersal rates by orders of magnitude relative to random (diffusive) motions. Model predictions considering microbial cell sizes relative to available liquid pathways sizes were in good agreement with experimental results for unsaturated soils. The new modeling platform enables quantitative consideration of key biophysical factors (e.g., pore space heterogeneities and hydration conditions) governing microbial interactions in 3-D soil pore spaces.

Heat shock protein 27 regulates human prostate cancer cell motility and metastatic progression

PubMed Central

Voll, Eric A; Ogden, Irene M; Pavese, Janet M; Huang, XiaoKe; Xu, Li; Jovanovic, Borko D; Bergan, Raymond C

2014-01-01

Prostate cancer (PCa) is the most common form of cancer in American men. Mortality from PCa is caused by the movement of cancer cells from the primary organ to form metastatic tumors at distant sites. Heat shock protein 27 (HSP27) is known to increase human PCa cell invasion and its overexpression is associated with metastatic disease. The role of HSP27 in driving PCa cell movement from the prostate to distant metastatic sites is unknown. Increased HSP27 expression increased metastasis as well as primary tumor mass. In vitro studies further examined the mechanism of HSP27-induced metastatic behavior. HSP27 did not affect cell detachment, adhesion, or migration, but did increase cell invasion. Cell invasion was dependent upon matrix metalloproteinase 2 (MMP-2), whose expression was increased by HSP27. In vivo, HSP27 induced commensurate changes in MMP-2 expression in tumors. These findings demonstrate that HSP27 drives metastatic spread of cancer cells from the prostate to distant sites, does so across a continuum of expression levels, and identifies HSP27-driven increases in MMP-2 expression as functionally relevant. These findings add to prior studies demonstrating that HSP27 increases PCa cell motility, growth and survival. Together, they demonstrate that HSP27 plays an important role in PCa progression. PMID:24798191

Fungal networks shape dynamics of bacterial dispersal and community assembly in cheese rind microbiomes.

PubMed

Zhang, Yuanchen; Kastman, Erik K; Guasto, Jeffrey S; Wolfe, Benjamin E

2018-01-23

Most studies of bacterial motility have examined small-scale (micrometer-centimeter) cell dispersal in monocultures. However, bacteria live in multispecies communities, where interactions with other microbes may inhibit or facilitate dispersal. Here, we demonstrate that motile bacteria in cheese rind microbiomes use physical networks created by filamentous fungi for dispersal, and that these interactions can shape microbial community structure. Serratia proteamaculans and other motile cheese rind bacteria disperse on fungal networks by swimming in the liquid layers formed on fungal hyphae. RNA-sequencing, transposon mutagenesis, and comparative genomics identify potential genetic mechanisms, including flagella-mediated motility, that control bacterial dispersal on hyphae. By manipulating fungal networks in experimental communities, we demonstrate that fungal-mediated bacterial dispersal can shift cheese rind microbiome composition by promoting the growth of motile over non-motile community members. Our single-cell to whole-community systems approach highlights the interactive dynamics of bacterial motility in multispecies microbiomes.

From immobilized cells to motile cells on a bed-of-nails: effects of vertical nanowire array density on cell behaviour

PubMed Central

Persson, Henrik; Li, Zhen; Tegenfeldt, Jonas O.; Oredsson, Stina; Prinz, Christelle N.

2015-01-01

The field of vertical nanowire array-based applications in cell biology is growing rapidly and an increasing number of applications are being explored. These applications almost invariably rely on the physical properties of the nanowire arrays, creating a need for a better understanding of how their physical properties affect cell behaviour. Here, we investigate the effects of nanowire density on cell migration, division and morphology for murine fibroblasts. Our results show that few nanowires are sufficient to immobilize cells, while a high nanowire spatial density enables a ”bed-of-nails” regime, where cells reside on top of the nanowires and are fully motile. The presence of nanowires decreases the cell proliferation rate, even in the “bed-of-nails” regime. We show that the cell morphology strongly depends on the nanowire density. Cells cultured on low (0.1 μm−2) and medium (1 μm−2) density substrates exhibit an increased number of multi-nucleated cells and micronuclei. These were not observed in cells cultured on high nanowire density substrates (4 μm−2). The results offer important guidelines to minimize cell-function perturbations on nanowire arrays. Moreover, these findings offer the possibility to tune cell proliferation and migration independently by adjusting the nanowire density, which may have applications in drug testing. PMID:26691936

Suppressive effects of 3-bromopyruvate on the proliferation and the motility of hepatocellular carcinoma cells.

PubMed

Tomizawa, Minoru; Shinozaki, Fuminobu; Motoyoshi, Yasufumi; Sugiyama, Takao; Yamamoto, Shigenori; Ishige, Naoki

2016-01-01

The compound 3-bromopyruvate (3BP) is an analogue of pyruvate, which is the final product of glycolysis that enters the citric acid cycle. The present study aimed to investigate the suppressive effects of 3BP on the proliferation and motility of hepatocellular carcinoma (HCC) cells. HLF and PLC/PRF/5 cells were cultured with 3BP and subjected to an MTS assay. Apoptosis was analyzed by hematoxylin and eosin staining. Cell motility was analyzed using a scratch assay. Real-time quantitative polymerase chain reaction (PCR) was performed to determine the expression levels of cyclin D1 and matrix metalloproteinase (MMP)9. Proliferation of both cell lines was significantly suppressed by 3BP at 100 µM (P<0.05). The expression level of cyclin D1 was decreased after 3BP treatment at 100 µM in both cell lines (P<0.05). Pyknotic nuclei were observed in the cells cultured with 3BP at 100 µM. These results revealed that 3BP suppressed cell proliferation, decreased the expression of cyclin D1, and induced apoptosis in HCC cells. 3BP significantly suppressed motility in both cell lines (P<0.05). The expression level of MMP9 was significantly decreased (P<0.05). 3BP suppressed the proliferation and motility of HCC cells by decreasing the expression of cyclin D1 and MMP9.

Targeting ion channels for the treatment of gastrointestinal motility disorders

PubMed Central

Beyder, Arthur

2012-01-01

Gastrointestinal (GI) functional and motility disorders are highly prevalent and responsible for long-term morbidity and sometimes mortality in the affected patients. It is estimated that one in three persons has a GI functional or motility disorder. However, diagnosis and treatment of these widespread conditions remains challenging. This partly stems from the multisystem pathophysiology, including processing abnormalities in the central and peripheral (enteric) nervous systems and motor dysfunction in the GI wall. Interstitial cells of Cajal (ICCs) are central to the generation and propagation of the cyclical electrical activity and smooth muscle cells (SMCs) are responsible for electromechanical coupling. In these and other excitable cells voltage-sensitive ion channels (VSICs) are the main molecular units that generate and regulate electrical activity. Thus, VSICs are potential targets for intervention in GI motility disorders. Research in this area has flourished with advances in the experimental methods in molecular and structural biology and electrophysiology. However, our understanding of the molecular mechanisms responsible for the complex and variable electrical behavior of ICCs and SMCs remains incomplete. In this review, we focus on the slow waves and action potentials in ICCs and SMCs. We describe the constituent VSICs, which include voltage-gated sodium (NaV), calcium (CaV), potassium (KV, KCa), chloride (Cl–) and nonselective ion channels (transient receptor potentials [TRPs]). VSICs have significant structural homology and common functional mechanisms. We outline the approaches and limitations and provide examples of targeting VSICs at the pores, voltage sensors and alternatively spliced sites. Rational drug design can come from an integrated view of the structure and mechanisms of gating and activation by voltage or mechanical stress. PMID:22282704

The Effect of Curcumin on Intracellular pH (pHi), Membrane Hyperpolarization and Sperm Motility.

PubMed

Naz, Rajesh K

2014-04-01

Curcumin has shown to affect sperm motility and function in vitro and fertility in vivo. The molecular mechanism(s) by which curcumin affects sperm motility has not been delineated. Since modulation of intracellular pH (pHi) and plasma membrane polarization is involved in sperm motility, the present study was conducted to investigate the effect of curcumin on these sperm (human and murine) parameters. The effect of curcumin on sperm forward motility was examined by counting percentages of forward moving sperm. The effect of curcumin on intracellular pH (pHi) was measured by the fluorescent pH indicator 2,7-bicarboxyethyl-5,6-carboxyfluorescein-acetoxymethyl ester (BCECF-AM). The effect of curcumin on plasma membrane polarization was examined using the fluorescence sensitive dye bis (1,3-dibarbituric acid)-trimethine oxanol [DiBAC4(3)]. Curcumin caused a concentration-dependent (p<0.05) decrease in forward motility of both human and mouse sperm. It also caused a concentration-dependent decrease in intracellular pH (pHi) in both human and mouse sperm. Curcumin induced significant (p<0.05) hyperpolarization of the plasma membrane in both human and mouse sperm. These findings indicate that curcumin inhibits sperm forward motility by intracellular acidification and hyperpolarization of sperm plasma membrane. This is the first study to our knowledge which examined the effect of curcumin on sperm pHi and membrane polarization that affect sperm forward motility. These exciting findings will have application in deciphering the signal transduction pathway involved in sperm motility and function and in development of a novel non-steroidal contraceptive for infertility.

Automated measurement of cell motility and proliferation

PubMed Central

Bahnson, Alfred; Athanassiou, Charalambos; Koebler, Douglas; Qian, Lei; Shun, Tongying; Shields, Donna; Yu, Hui; Wang, Hong; Goff, Julie; Cheng, Tao; Houck, Raymond; Cowsert, Lex

2005-01-01

Background Time-lapse microscopic imaging provides a powerful approach for following changes in cell phenotype over time. Visible responses of whole cells can yield insight into functional changes that underlie physiological processes in health and disease. For example, features of cell motility accompany molecular changes that are central to the immune response, to carcinogenesis and metastasis, to wound healing and tissue regeneration, and to the myriad developmental processes that generate an organism. Previously reported image processing methods for motility analysis required custom viewing devices and manual interactions that may introduce bias, that slow throughput, and that constrain the scope of experiments in terms of the number of treatment variables, time period of observation, replication and statistical options. Here we describe a fully automated system in which images are acquired 24/7 from 384 well plates and are automatically processed to yield high-content motility and morphological data. Results We have applied this technology to study the effects of different extracellular matrix compounds on human osteoblast-like cell lines to explore functional changes that may underlie processes involved in bone formation and maintenance. We show dose-response and kinetic data for induction of increased motility by laminin and collagen type I without significant effects on growth rate. Differential motility response was evident within 4 hours of plating cells; long-term responses differed depending upon cell type and surface coating. Average velocities were increased approximately 0.1 um/min by ten-fold increases in laminin coating concentration in some cases. Comparison with manual tracking demonstrated the accuracy of the automated method and highlighted the comparative imprecision of human tracking for analysis of cell motility data. Quality statistics are reported that associate with stage noise, interference by non-cell objects, and uncertainty in the outlining and positioning of cells by automated image analysis. Exponential growth, as monitored by total cell area, did not linearly correlate with absolute cell number, but proved valuable for selection of reliable tracking data and for disclosing between-experiment variations in cell growth. Conclusion These results demonstrate the applicability of a system that uses fully automated image acquisition and analysis to study cell motility and growth. Cellular motility response is determined in an unbiased and comparatively high throughput manner. Abundant ancillary data provide opportunities for uniform filtering according to criteria that select for biological relevance and for providing insight into features of system performance. Data quality measures have been developed that can serve as a basis for the design and quality control of experiments that are facilitated by automation and the 384 well plate format. This system is applicable to large-scale studies such as drug screening and research into effects of complex combinations of factors and matrices on cell phenotype. PMID:15831094

Treatment with insulin uncovers the motogenic capacity of nitric oxide in aortic smooth muscle cells: dependence on Gab1 and Gab1-SHP2 association.

PubMed

Dixit, Madhulika; Zhuang, Daming; Ceacareanu, Bogdan; Hassid, Aviv

2003-11-14

Contrary to the antimotogenic effect of NO in dedifferentiated vascular smooth muscle cells (VSMCs), we have reported that NO stimulates the motility of differentiated cultured VSMC isolated from adult rats. This process involves upregulation of protein tyrosine phosphatase SHP2, followed by downregulation of RhoA activity. In the present study, we tested the hypothesis that insulin alters the motogenic phenotype of cultured rat aortic smooth muscle cells exposed to NO from inhibition to stimulation of cell motility. We demonstrate for the first time that NO stimulates the motility of VSMCs cultured for several days in the presence but not the absence of insulin. Moreover, we show that NO blocks PDGF-induced cell motility in insulin-naive but not in insulin-treated cells. We also demonstrate that the scaffold adapter protein Gab1, considered a physiological activator of protein tyrosine phosphatase SHP2, increases cell motility in the presence but not the absence of insulin. In cells cultured in the presence of insulin, overexpression of Gab1 mimics, whereas a dominant-negative allele of Gab1 (Gab1YF) blocks, the motility-stimulatory effect of NO. Cotransfection experiments with dominant-negative Gab1 and wild-type SHP2 or wild-type Gab1 and dominant-negative SHP2 indicate that the two proteins work together as a functional unit to induce motility. Because chronic insulin can increase the levels of phosphatidylinositol 3 (PI3) kinase in several models of hyperinsulinemia, we also tested the potential involvement of this enzyme in mechanisms leading to increased cell motility. We found that the motogenic effect of NO, Gab1, and SHP2 was blocked by the selective PI3 kinase inhibitor LY294002, suggesting a requirement of PI3 kinase in mediating motogenesis. These observations may be relevant to molecular mechanisms related to the pathogenesis of vascular disease in hyperinsulinemic diabetes. The full text of this article is available online at http://www.circresaha.org.

Mucosal fluid glycoprotein DMBT1 suppresses twitching motility and virulence of the opportunistic pathogen Pseudomonas aeruginosa

PubMed Central

Evans, David J.; Fleiszig, Suzanne M. J.

2017-01-01

It is generally thought that mucosal fluids protect underlying epithelial surfaces against opportunistic infection via their antimicrobial activity. However, our published data show that human tear fluid can protect against the major opportunistic pathogen Pseudomonas aeruginosa independently of bacteriostatic activity. Here, we explored the mechanisms for tear protection, focusing on impacts of tear fluid on bacterial virulence factor expression. Results showed that tear fluid suppressed twitching motility, a type of surface-associated movement conferred by pili. Previously, we showed that twitching is critical for P. aeruginosa traversal of corneal epithelia, exit from epithelial cells after internalization, and corneal virulence. Inhibition of twitching by tear fluid was dose-dependent with dilutions to 6.25% retaining activity. Purified lactoferrin, lysozyme, and contrived tears containing these, and many other, tear components lacked the activity. Systematic protein fractionation, mass spectrometry, and immunoprecipitation identified the glycoprotein DMBT1 (Deleted in Malignant Brain Tumors 1) in tear fluid as required. DMBT1 purified from human saliva also inhibited twitching, as well as P. aeruginosa traversal of human corneal epithelial cells in vitro, and reduced disease pathology in a murine model of corneal infection. DMBT1 did not affect PilA expression, nor bacterial intracellular cyclicAMP levels, and suppressed twitching motility of P. aeruginosa chemotaxis mutants (chpB, pilK), and an adenylate cyclase mutant (cyaB). However, dot-immunoblot assays showed purified DMBT1 binding of pili extracted from PAO1 suggesting that twitching inhibition may involve a direct interaction with pili. The latter could affect extension or retraction of pili, their interactions with biotic or abiotic surfaces, or cause their aggregation. Together, the data suggest that DMBT1 inhibition of twitching motility contributes to the mechanisms by which mucosal fluids protect against P. aeruginosa infection. This study also advances our understanding of how mucosal fluids protect against infection, and suggests directions for novel biocompatible strategies to protect our surface epithelia against a major opportunistic pathogen. PMID:28489917

HIV-1 Nef interferes with host cell motility by deregulation of Cofilin.

PubMed

Stolp, Bettina; Reichman-Fried, Michal; Abraham, Libin; Pan, Xiaoyu; Giese, Simone I; Hannemann, Sebastian; Goulimari, Polyxeni; Raz, Erez; Grosse, Robert; Fackler, Oliver T

2009-08-20

HIV-1 Nef is a key factor in AIDS pathogenesis. Here, we report that Nef potently inhibits motility of fibroblasts and chemotaxis of HIV-1-infected primary human T lymphocytes toward the chemokines SDF-1alpha, CCL-19, and CCL-21 ex vivo. Furthermore, Nef inhibits guided motility of zebrafish primordial germ cells toward endogenous SDF-1a in vivo. These migration defects result from Nef-mediated inhibition of the actin remodeling normally triggered by migratory stimuli. Nef strongly induces phosphorylation of cofilin, inactivating this evolutionarily conserved actin-depolymerizing factor that promotes cell motility when unphosphorylated. Nef-dependent cofilin deregulation requires association of Nef with the cellular kinase Pak2. Disruption of Nef-Pak2 association restores the cofilin phosphorylation levels and actin remodeling that facilitate cell motility. We conclude that HIV-1 Nef alters Pak2 function, which directly or indirectly inactivates cofilin, thereby restricting migration of infected T lymphocytes as part of a strategy to optimize immune evasion and HIV-1 replication.

Stimulation of glioma cell motility by expression, proteolysis, and release of the L1 neural cell recognition molecule.

PubMed

Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S

2009-10-29

Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding alphavbeta5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain.

Stimulation of glioma cell motility by expression, proteolysis, and release of the L1 neural cell recognition molecule

PubMed Central

Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S

2009-01-01

Background Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. Results L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding αvβ5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Conclusion Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain. PMID:19874583

An Ribonuclease T2 Family Protein Modulates Acinetobacter baumannii Abiotic Surface Colonization

PubMed Central

Jacobs, Anna C.; Blanchard, Catlyn E.; Catherman, Seana C.; Dunman, Paul M.; Murata, Yoshihiko

2014-01-01

Acinetobacter baumannii is an emerging bacterial pathogen of considerable medical concern. The organism's transmission and ability to cause disease has been associated with its propensity to colonize and form biofilms on abiotic surfaces in health care settings. To better understand the genetic determinants that affect biomaterial attachment, we performed a transposon mutagenesis analysis of abiotic surface-colonization using A. baumannii strain 98-37-09. Disruption of an RNase T2 family gene was found to limit the organism's ability to colonize polystyrene, polypropylene, glass, and stainless steel surfaces. DNA microarray analyses revealed that in comparison to wild type and complemented cells, the RNase T2 family mutant exhibited reduced expression of 29 genes, 15 of which are predicted to be associated with bacterial attachment and surface-associated motility. Motility assays confirmed that RNase T2 mutant displays a severe motility defect. Taken together, our results indicate that the RNase T2 family protein identified in this study is a positive regulator of A. baumannii's ability to colonize inanimate surfaces and motility. Moreover, the enzyme may be an effective target for the intervention of biomaterial colonization, and consequently limit the organism's transmission within the hospital setting. PMID:24489668

Apoptosis Signal-Regulating Kinase 1 Is Involved in Brain-Derived Neurotrophic Factor (BDNF)-Enhanced Cell Motility and Matrix Metalloproteinase 1 Expression in Human Chondrosarcoma Cells

PubMed Central

Lin, Chih-Yang; Chang, Sunny Li-Yun; Fong, Yi-Chin; Hsu, Chin-Jung; Tang, Chih-Hsin

2013-01-01

Chondrosarcoma is the primary malignancy of bone that is characterized by a potent capacity to invade locally and cause distant metastasis, and is therefore associated with poor prognoses. Chondrosarcoma further shows a predilection for metastasis to the lungs. The brain-derived neurotrophic factor (BDNF) is a small molecule in the neurotrophin family of growth factors that is associated with the disease status and outcome of cancers. However, the effect of BDNF on cell motility in human chondrosarcoma cells is mostly unknown. Here, we found that human chondrosarcoma cell lines had significantly higher cell motility and BDNF expression compared to normal chondrocytes. We also found that BDNF increased cell motility and expression of matrix metalloproteinase-1 (MMP-1) in human chondrosarcoma cells. BDNF-mediated cell motility and MMP-1 up-regulation were attenuated by Trk inhibitor (K252a), ASK1 inhibitor (thioredoxin), JNK inhibitor (SP600125), and p38 inhibitor (SB203580). Furthermore, BDNF also promoted Sp1 activation. Our results indicate that BDNF enhances the migration and invasion activity of chondrosarcoma cells by increasing MMP-1 expression through a signal transduction pathway that involves the TrkB receptor, ASK1, JNK/p38, and Sp1. BDNF thus represents a promising new target for treating chondrosarcoma metastasis. PMID:23892595

Modulation of Chlamydomonas reinhardtii flagellar motility by redox poise

PubMed Central

Wakabayashi, Ken-ichi; King, Stephen M.

2006-01-01

Redox-based regulatory systems are essential for many cellular activities. Chlamydomonas reinhardtii exhibits alterations in motile behavior in response to different light conditions (photokinesis). We hypothesized that photokinesis is signaled by variations in cytoplasmic redox poise resulting from changes in chloroplast activity. We found that this effect requires photosystem I, which generates reduced NADPH. We also observed that photokinetic changes in beat frequency and duration of the photophobic response could be obtained by altering oxidative/reductive stress. Analysis of reactivated cell models revealed that this redox poise effect is mediated through the outer dynein arms (ODAs). Although the global redox state of the thioredoxin-related ODA light chains LC3 and LC5 and the redox-sensitive Ca2+-binding subunit of the docking complex DC3 did not change upon light/dark transitions, we did observe significant alterations in their interactions with other flagellar components via mixed disulfides. These data indicate that redox poise directly affects ODAs and suggest that it may act in the control of flagellar motility. PMID:16754958

Roles of quorum sensing molecules from Rhizobium etli RT1 in bacterial motility and biofilm formation.

PubMed

Dixit, Swarnita; Dubey, Ramesh Chand; Maheshwari, Dinesh Kumar; Seth, Prahlad Kishore; Bajpai, Vivek K

Strain RT1 was isolated from root nodules of Lens culinaris (a lentil) and characterized as Rhizobium etli (a Gram-negative soil-borne bacterium) by 16S rDNA sequencing and phylogenetic analysis. The signaling molecules produced by R. etli (RT1) were detected and identified by high-performance liquid chromatography coupled with mass spectrometry. The most abundant and biologically active N-acyl homoserine lactone molecules (3-oxo-C 8 -HSL and 3-OH-C 14 -HSL) were detected in the ethyl acetate extract of RT1. The biological role of 3-oxo-C 8 -HSL was evaluated in RT1. Bacterial motility and biofilm formation were affected or modified on increasing concentrations of 3-oxo-C 8 -HSL. Results confirmed the existence of cell communication in RT1 mediated by 3-oxo-C 8 -HSL, and positive correlations were found among quorum sensing, motility and biofilm formation in RT1. Copyright © 2017 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Reduction in ins-7 gene expression in non-neuronal cells of high glucose exposed Caenorhabditis elegans protects from reactive metabolites, preserves neuronal structure and head motility, and prolongs lifespan.

PubMed

Mendler, Michael; Riedinger, Christin; Schlotterer, Andrea; Volk, Nadine; Fleming, Thomas; Herzig, Stephan; Nawroth, Peter P; Morcos, Michael

2017-02-01

Glucose derived metabolism generates reactive metabolites affecting the neuronal system and lifespan in C. elegans. Here, the role of the insulin homologue ins-7 and its downstream effectors in the generation of high glucose induced neuronal damage and shortening of lifespan was studied. In C. elegans high glucose conditions induced the expression of the insulin homologue ins-7. Abrogating ins-7 under high glucose conditions in non-neuronal cells decreased reactive oxygen species (ROS)-formation and accumulation of methylglyoxal derived advanced glycation endproducts (AGEs), prevented structural neuronal damage and normalised head motility and lifespan. The restoration of lifespan by decreased ins-7 expression was dependent on the concerted action of sod-3 and glod-4 coding for the homologues of iron-manganese superoxide dismutase and glyoxalase 1, respectively. Under high glucose conditions mitochondria-mediated oxidative stress and glycation are downstream targets of ins-7. This impairs the neuronal system and longevity via a non-neuronal/neuronal crosstalk by affecting sod-3 and glod-4, thus giving further insight into the pathophysiology of diabetic complications. Copyright © 2017 Elsevier Inc. All rights reserved.

Chlorpyrifos, chlorpyrifos-oxon, and diisopropylfluorophosphate inhibit kinesin-dependent microtubule motility

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gearhart, Debra A.; Sickles, Dale W.; Buccafusco, Jerry J.

2007-01-01

Diisopropylfluorophosphate, originally developed as a chemical warfare agent, is structurally similar to nerve agents, and chlorpyrifos has extensive worldwide use as an agricultural pesticide. While inhibition of cholinesterases underlies the acute toxicity of these organophosphates, we previously reported impaired axonal transport in the sciatic nerves from rats treated chronically with subthreshold doses of chlorpyrifos. Those data indicate that chlorpyrifos (and/or its active metabolite, chlorpyrifos-oxon) might directly affect the function of kinesin and/or microtubules-the principal proteins that mediate anterograde axonal transport. The current report describes in vitro assays to assess the concentration-dependent effects of chlorpyrifos (0-10 {mu}M), chlorpyrifos-oxon (0-10 {mu}M), andmore » diisopropylfluorophosphate (0-0.59 nM) on kinesin-dependent microtubule motility. Preincubating bovine brain microtubules with the organophosphates did not alter kinesin-mediated microtubule motility. In contrast, preincubation of bovine brain kinesin with diisopropylfluorophosphate, chlorpyrifos, or chlorpyrifos-oxon produced a concentration-dependent increase in the number of locomoting microtubules that detached from the kinesin-coated glass cover slip. Our data suggest that the organophosphates-chlorpyrifos-oxon, chlorpyrifos, and diisopropylfluorophosphate-directly affect kinesin, thereby disrupting kinesin-dependent transport on microtubules. Kinesin-dependent movement of vesicles, organelles, and other cellular components along microtubules is fundamental to the organization of all eukaryotic cells, especially in neurons where organelles and proteins synthesized in the cell body must move down long axons to pre-synaptic sites in nerve terminals. We postulate that disruption of kinesin-dependent intracellular transport could account for some of the long-term effects of organophosphates on the peripheral and central nervous system.« less

Energy output of a single outer hair cell: Effect of resonance

NASA Astrophysics Data System (ADS)

Iwasa, Kuni H.

2018-05-01

The ability of the mammalian ear in processing high frequency sounds, up to ˜100 kHz, is based on the capability of outer hair cells (OHCs) in responding to stimulation at high frequencies. These cells show a unique motility in their cell body coupled with charge movement. With this motile element, voltage changes generated by stimuli at their hair bundles drive the cell body and that, in turn, amplifies the signal. In vitro experiments show that the movement of these charges significantly increases the membrane capacitance, limiting the motile activity by an additional attenuation of voltage changes. It was found, however, that such an effect is due to the absence of mechanical load. In the presence of mechanical load, particularly inertial load, such as under in vivo conditions, the movement of motile charges should reduce the membrane capacitance, enhancing the mechanical power output.

A minimal physical model for crawling cells

NASA Astrophysics Data System (ADS)

Tiribocchi, Adriano; Tjhung, Elsen; Marenduzzo, Davide; Cates, Michael E.

Cell motility in higher organisms (eukaryotes) is fundamental to biological functions such as wound healing or immune response, and is also implicated in diseases such as cancer. For cells crawling on solid surfaces, considerable insights into motility have been gained from experiments replicating such motion in vitro. Such experiments show that crawling uses a combination of actin treadmilling (polymerization), which pushes the front of a cell forward, and myosin-induced stress (contractility), which retracts the rear. We present a simplified physical model of a crawling cell, consisting of a droplet of active polar fluid with contractility throughout, but treadmilling connected to a thin layer near the supporting wall. The model shows a variety of shapes and/or motility regimes, some closely resembling cases seen experimentally. Our work supports the view that cellular motility exploits autonomous physical mechanisms whose operation does not need continuous regulatory effort.

A minimal physical model captures the shapes of crawling cells

NASA Astrophysics Data System (ADS)

Tjhung, E.; Tiribocchi, A.; Marenduzzo, D.; Cates, M. E.

2015-01-01

Cell motility in higher organisms (eukaryotes) is crucial to biological functions ranging from wound healing to immune response, and also implicated in diseases such as cancer. For cells crawling on hard surfaces, significant insights into motility have been gained from experiments replicating such motion in vitro. Such experiments show that crawling uses a combination of actin treadmilling (polymerization), which pushes the front of a cell forward, and myosin-induced stress (contractility), which retracts the rear. Here we present a simplified physical model of a crawling cell, consisting of a droplet of active polar fluid with contractility throughout, but treadmilling connected to a thin layer near the supporting wall. The model shows a variety of shapes and/or motility regimes, some closely resembling cases seen experimentally. Our work strongly supports the view that cellular motility exploits autonomous physical mechanisms whose operation does not need continuous regulatory effort.

Self-organized cell motility

NASA Astrophysics Data System (ADS)

Du, Xinxin; Doubrovinski, Konstantin

2011-03-01

Cell migration plays a key role in a wide range of biological phenomena, such as morphogenesis, chemotaxis, and wound healing. Cell locomotion relies on the cytoskeleton, a meshwork of filamentous proteins, intrinsically out of thermodynamic equilibrium and cross-linked by molecular motors, proteins that turn chemical energy into mechanical work. In the course of locomotion, cells remain polarized, i.e. they retain a single direction of motion in the absence of external cues. Traditionally, polarization has been attributed to intracellular signaling. However, recent experiments show that polarization may be a consequence of self-organized cytoskeletal dynamics. Our aim is to elucidate the mechanisms by which persistent unidirectional locomotion may arise through simple mechanical interactions of the cytoskeletal proteins. To this end, we develop a simple physical description of cytoskeletal dynamics. We find that the proposed description accounts for a range of phenomena associated with cell motility, including spontaneous polarization, persistent unidirectional motion, and the co-existence of motile and non-motile states.

Motor-driven intracellular transport powers bacterial gliding motility.

PubMed

Sun, Mingzhai; Wartel, Morgane; Cascales, Eric; Shaevitz, Joshua W; Mignot, Tâm

2011-05-03

Protein-directed intracellular transport has not been observed in bacteria despite the existence of dynamic protein localization and a complex cytoskeleton. However, protein trafficking has clear potential uses for important cellular processes such as growth, development, chromosome segregation, and motility. Conflicting models have been proposed to explain Myxococcus xanthus motility on solid surfaces, some favoring secretion engines at the rear of cells and others evoking an unknown class of molecular motors distributed along the cell body. Through a combination of fluorescence imaging, force microscopy, and genetic manipulation, we show that membrane-bound cytoplasmic complexes consisting of motor and regulatory proteins are directionally transported down the axis of a cell at constant velocity. This intracellular motion is transmitted to the exterior of the cell and converted to traction forces on the substrate. Thus, this study demonstrates the existence of a conserved class of processive intracellular motors in bacteria and shows how these motors have been adapted to produce cell motility.

L1 stimulation of human glioma cell motility correlates with FAK activation

PubMed Central

Yang, Muhua; Li, Yupei; Chilukuri, Kalyani; Brady, Owen A.; Boulos, Magdy I.; Kappes, John C.

2011-01-01

The neural adhesion/recognition protein L1 (L1CAM; CD171) has been shown or implicated to function in stimulation of cell motility in several cancer types, including high-grade gliomas. Our previous work demonstrated the expression and function of L1 protein in stimulation of cell motility in rat glioma cells. However, the mechanism of this stimulation is still unclear. This study further investigated the function of L1 and L1 proteolysis in human glioblastoma multiforme (GBM) cell migration and invasion, as well as the mechanism of this stimulation. L1 mRNA was found to be present in human T98G GBM cell line but not in U-118 MG grade III human glioma cell line. L1 protein expression, proteolysis, and release were found in T98G cells and human surgical GBM cells by Western blotting. Exosome-like vesicles released by T98G cells were purified and contained full-length L1. In a scratch assay, T98G cells that migrated into the denuded scratch area exhibited upregulation of ADAM10 protease expression coincident with loss of surface L1. GBM surgical specimen cells exhibited a similar loss of cell surface L1 when xenografted into the chick embryo brain. When lentivirally introduced shRNA was used to attenuate L1 expression, such T98G/shL1 cells exhibited significantly decreased cell motility by time lapse microscopy in our quantitative Super Scratch assay. These cells also showed a decrease in FAK activity and exhibited increased focal complexes. L1 binding integrins which activate FAK were found in T98G and U-118 MG cells. Addition of L1 ectodomain-containing media (1) rescued the decreased cell motility of T98G/shL1 cells and (2) increased cell motility of U-118 MG cells but (3) did not further increase T98G cell motility. Injection of L1-attenuated T98G/shL1 cells into embryonic chick brains resulted in the absence of detectable invasion compared to control cells which invaded brain tissue. These studies support a mechanism where glioma cells at the edge of a cell mass upregulate ADAM10 to proteolyze surface L1 and the resultant ectodomain increases human glioma cell migration and invasion by binding to integrin receptors, activating FAK, and increasing turnover of focal complexes. PMID:21373966

L1 stimulation of human glioma cell motility correlates with FAK activation.

PubMed

Yang, Muhua; Li, Yupei; Chilukuri, Kalyani; Brady, Owen A; Boulos, Magdy I; Kappes, John C; Galileo, Deni S

2011-10-01

The neural adhesion/recognition protein L1 (L1CAM; CD171) has been shown or implicated to function in stimulation of cell motility in several cancer types, including high-grade gliomas. Our previous work demonstrated the expression and function of L1 protein in stimulation of cell motility in rat glioma cells. However, the mechanism of this stimulation is still unclear. This study further investigated the function of L1 and L1 proteolysis in human glioblastoma multiforme (GBM) cell migration and invasion, as well as the mechanism of this stimulation. L1 mRNA was found to be present in human T98G GBM cell line but not in U-118 MG grade III human glioma cell line. L1 protein expression, proteolysis, and release were found in T98G cells and human surgical GBM cells by Western blotting. Exosome-like vesicles released by T98G cells were purified and contained full-length L1. In a scratch assay, T98G cells that migrated into the denuded scratch area exhibited upregulation of ADAM10 protease expression coincident with loss of surface L1. GBM surgical specimen cells exhibited a similar loss of cell surface L1 when xenografted into the chick embryo brain. When lentivirally introduced shRNA was used to attenuate L1 expression, such T98G/shL1 cells exhibited significantly decreased cell motility by time lapse microscopy in our quantitative Super Scratch assay. These cells also showed a decrease in FAK activity and exhibited increased focal complexes. L1 binding integrins which activate FAK were found in T98G and U-118 MG cells. Addition of L1 ectodomain-containing media (1) rescued the decreased cell motility of T98G/shL1 cells and (2) increased cell motility of U-118 MG cells but (3) did not further increase T98G cell motility. Injection of L1-attenuated T98G/shL1 cells into embryonic chick brains resulted in the absence of detectable invasion compared to control cells which invaded brain tissue. These studies support a mechanism where glioma cells at the edge of a cell mass upregulate ADAM10 to proteolyze surface L1 and the resultant ectodomain increases human glioma cell migration and invasion by binding to integrin receptors, activating FAK, and increasing turnover of focal complexes.

Effects of transforming growth factor-beta1 on cell motility, collagen gel contraction, myofibroblastic differentiation, and extracellular matrix expression of human adipose-derived stem cell.

PubMed

Kakudo, Natsuko; Kushida, Satoshi; Suzuki, Kenji; Ogura, Tsunetaka; Notodihardjo, Priscilla Valentin; Hara, Tomoya; Kusumoto, Kenji

2012-12-01

Human adipose-derived stem cells (ASCs) are adult pluripotent stem cells, and their usefulness in plastic surgery has garnered attention in recent years. Although, there have been expectations that ASCs might function in wound repair and regeneration, no studies to date have examined the role of ASCs in the mechanism that promotes wound-healing. Transforming growth factor-beta1 (TGF-β1) is a strong candidate cytokine for the triggering of mesenchymal stem cell migration, construction of extracellular matrices, and differentiation of ASCs into myofibroblasts. Cell proliferation, motility, and differentiation, as well as extracellular matrix production, play an important role in wound-healing. We have evaluated the capacity of ASCs to proliferate and their potential to differentiate into phenotypic myofibroblasts, as well as their cell motility and collagen gel contraction ability, when cultured with TGF-β1. Cell motility was analyzed using a wound-healing assay. ASCs that differentiated into myofibroblasts expressed the gene for alpha-smooth muscle actin, and its protein expression was detected immunohistochemically. The extracellular matrix expression in ASCs was evaluated using real-time RT-PCR. Based on the results, we conclude that human ASCs have the potential for cell motility, extracellular matrix gene expression, gel contraction, and differentiation into myofibroblasts and, therefore, may play an important role in the wound-healing process.

SLP-76-ADAP adaptor module regulates LFA-1 mediated costimulation and T cell motility.

PubMed

Wang, Hongyan; Wei, Bin; Bismuth, Georges; Rudd, Christopher E

2009-07-28

Although adaptor ADAP (FYB) and its binding to SLP-76 has been implicated in TcR-induced "inside-out" signaling for LFA-1 activation in T cells, little is known regarding its role in LFA-1-mediated "outside-in" signaling. In this study, we demonstrate that ADAP and SLP-76-ADAP binding are coupled to LFA-1 costimulation of IL-2 production, F-actin clustering, cell polarization, and T cell motility. LFA-1 enhancement of anti-CD3-induced IL-2 production was completely dependent on SLP-76-ADAP binding. Further, anti-CD3 was found to require CD11a ligation by antibody or ICAM1 to cause T cell polarization. ADAP augmented this polarization induced by anti-CD3/CD11a, but not by anti-CD3 alone. ADAP expression with LFA-1 ligation alone was sufficient to polarize T cells directly and to increase T cell motility whereas the loss of ADAP in ADAP-/- primary T cells reduced motility. A mutant lacking SLP-76-binding sites (M12) blocked LFA-1 costimulation of IL-2 production, polarization, and motility. LFA-1-ADAP polarization was also dependent on src kinases, Rho GTPases, phospholipase C, and phosphoinositol 3-kinase. Our findings provide evidence of an obligatory role for the SLP-76-ADAP module in LFA-1-mediated costimulation in T cells.

A simple and accurate rule-based modeling framework for simulation of autocrine/paracrine stimulation of glioblastoma cell motility and proliferation by L1CAM in 2-D culture.

PubMed

Caccavale, Justin; Fiumara, David; Stapf, Michael; Sweitzer, Liedeke; Anderson, Hannah J; Gorky, Jonathan; Dhurjati, Prasad; Galileo, Deni S

2017-12-11

Glioblastoma multiforme (GBM) is a devastating brain cancer for which there is no known cure. Its malignancy is due to rapid cell division along with high motility and invasiveness of cells into the brain tissue. Simple 2-dimensional laboratory assays (e.g., a scratch assay) commonly are used to measure the effects of various experimental perturbations, such as treatment with chemical inhibitors. Several mathematical models have been developed to aid the understanding of the motile behavior and proliferation of GBM cells. However, many are mathematically complicated, look at multiple interdependent phenomena, and/or use modeling software not freely available to the research community. These attributes make the adoption of models and simulations of even simple 2-dimensional cell behavior an uncommon practice by cancer cell biologists. Herein, we developed an accurate, yet simple, rule-based modeling framework to describe the in vitro behavior of GBM cells that are stimulated by the L1CAM protein using freely available NetLogo software. In our model L1CAM is released by cells to act through two cell surface receptors and a point of signaling convergence to increase cell motility and proliferation. A simple graphical interface is provided so that changes can be made easily to several parameters controlling cell behavior, and behavior of the cells is viewed both pictorially and with dedicated graphs. We fully describe the hierarchical rule-based modeling framework, show simulation results under several settings, describe the accuracy compared to experimental data, and discuss the potential usefulness for predicting future experimental outcomes and for use as a teaching tool for cell biology students. It is concluded that this simple modeling framework and its simulations accurately reflect much of the GBM cell motility behavior observed experimentally in vitro in the laboratory. Our framework can be modified easily to suit the needs of investigators interested in other similar intrinsic or extrinsic stimuli that influence cancer or other cell behavior. This modeling framework of a commonly used experimental motility assay (scratch assay) should be useful to both researchers of cell motility and students in a cell biology teaching laboratory.

A free-boundary model of a motile cell explains turning behavior.

PubMed

Nickaeen, Masoud; Novak, Igor L; Pulford, Stephanie; Rumack, Aaron; Brandon, Jamie; Slepchenko, Boris M; Mogilner, Alex

2017-11-01

To understand shapes and movements of cells undergoing lamellipodial motility, we systematically explore minimal free-boundary models of actin-myosin contractility consisting of the force-balance and myosin transport equations. The models account for isotropic contraction proportional to myosin density, viscous stresses in the actin network, and constant-strength viscous-like adhesion. The contraction generates a spatially graded centripetal actin flow, which in turn reinforces the contraction via myosin redistribution and causes retraction of the lamellipodial boundary. Actin protrusion at the boundary counters the retraction, and the balance of the protrusion and retraction shapes the lamellipodium. The model analysis shows that initiation of motility critically depends on three dimensionless parameter combinations, which represent myosin-dependent contractility, a characteristic viscosity-adhesion length, and a rate of actin protrusion. When the contractility is sufficiently strong, cells break symmetry and move steadily along either straight or circular trajectories, and the motile behavior is sensitive to conditions at the cell boundary. Scanning of a model parameter space shows that the contractile mechanism of motility supports robust cell turning in conditions where short viscosity-adhesion lengths and fast protrusion cause an accumulation of myosin in a small region at the cell rear, destabilizing the axial symmetry of a moving cell.

Characterization of Amoeba proteus myosin VI immunoanalog.

PubMed

Dominik, Magdalena; Kłopocka, Wanda; Pomorski, Paweł; Kocik, Elzbieta; Redowicz, Maria Jolanta

2005-07-01

Amoeba proteus, the highly motile free-living unicellular organism, has been widely used as a model to study cell motility. However, molecular mechanisms underlying its unique locomotion and intracellular actin-based-only trafficking remain poorly understood. A search for myosin motors responsible for vesicular transport in these giant cells resulted in detection of 130-kDa protein interacting with several polyclonal antibodies against different tail regions of human and chicken myosin VI. This protein was binding to actin in the ATP-dependent manner, and immunoprecipitated with anti-myosin VI antibodies. In order to characterize its possible functions in vivo, its cellular distribution and colocalization with actin filaments and dynamin II during migration and pinocytosis were examined. In migrating amoebae, myosin VI immunoanalog localized to vesicular structures, particularly within the perinuclear and sub-plasma membrane areas, and colocalized with dynamin II immunoanalog and actin filaments. The colocalization was even more evident in pinocytotic cells as proteins concentrated within pinocytotic pseudopodia. Moreover, dynamin II and myosin VI immunoanalogs cosedimented with actin filaments, and were found on the same isolated vesicles. Blocking endogenous myosin VI immunoanalog with anti-myosin VI antibodies inhibited the rate of pseudopodia protrusion (about 19% decrease) and uroidal retraction (about 28% decrease) but did not affect cell morphology and the manner of cell migration. Treatment with anti-human dynamin II antibodies led to changes in directionality of amebae migration and affected the rate of only uroidal translocation (about 30% inhibition). These results indicate that myosin VI immunoanalog is expressed in protist Amoeba proteus and may be involved in vesicle translocation and cell locomotion.

Environmental Stress-mediated EPS Production Shape Microbial Activity on Various Hydrated Rough Surfaces

NASA Astrophysics Data System (ADS)

Wang, G.; Liu, L.; Chen, G.

2016-12-01

The complex environmental physical and chemical processes and interplay with the associating biological responses are keys to understanding the environmental microbiology ensconced in environmental remediation, water quality control, food safety, nutrient cycling, and etc., yet remain poorly understood. Using experimental micromodels, we study how environmental conditions (e.g., hydration fluctuation, nutrient limitation, pH variation, etc.) affect microbial extracellular polymeric substances (EPS) production and their configuration within various hydrated surfaces, and impacts on microbial motility, surface attachment, aggregation, and other bioremediation activities. To elucidate the potential mechanisms underlying the complex bio-physicochemical processes, we developed an individual-based and spatio-temporally resolved modeling platform that explicitly considers microscale aqueous-phase configuration and nutrient transport/diffusion and associated biophysical processes affecting individual microbial cell life history. We quantitatively explore the effects of the above microscale environmental processes on bio-physicochemical interactions affecting microbial growth, motility, surface attachment and aggregation, and shaping population interactions and functions. Simulation scenarios of microbial induced pollutant (e.g., roxarsone) biotransformation on various hydrated rough surfaces will also be present.

Aldolase sequesters WASP and affects WASP/Arp2/3-stimulated actin dynamics.

PubMed

Ritterson Lew, Carolyn; Tolan, Dean R

2013-08-01

In addition to its roles in sugar metabolism, fructose-1,6-bisphosphate aldolase (aldolase) has been implicated in cellular functions independent from these roles, termed "moonlighting functions." These moonlighting functions likely involve the known aldolase-actin interaction, as many proteins with which aldolase interacts are involved in actin-dependent processes. Specifically, aldolase interacts both in vitro and in cells with Wiskott-Aldrich Syndrome Protein (WASP), a protein involved in controlling actin dynamics, yet the function of this interaction remains unknown. Here, the effect of aldolase on WASP-dependent processes in vitro and in cells is investigated. Aldolase inhibits WASP/Arp2/3-dependent actin polymerization in vitro. In cells, knockdown of aldolase results in a decreased rate of cell motility and cell spreading, two WASP-dependent processes. Expression of exogenous aldolase rescues these defects. Whether these effects of aldolase on WASP-dependent processes were due to aldolase catalysis or moonlighting functions is tested using aldolase variants defective in either catalytic or actin-binding activity. While the actin-binding deficient aldolase variant is unable to inhibit actin polymerization in vitro and is unable to rescue cell motility defects in cells, the catalytically inactive aldolase is able to perform these functions, providing evidence that aldolase moonlighting plays a role in WASP-mediated processes. Copyright © 2013 Wiley Periodicals, Inc.

Soy lecithin replaces egg yolk for cryopreservation of human sperm without adversely affecting postthaw motility, morphology, sperm DNA integrity, or sperm binding to hyaluronate.

PubMed

Reed, Michael L; Ezeh, Peace C; Hamic, Amanda; Thompson, Douglas J; Caperton, Charles L

2009-11-01

Semen specimens (one ejaculate from each of 20 consenting study participants) were subjected to routine semen analysis, an in vitro sperm binding assay (HBA), and a sperm chromatin dispersion assay (HaloSperm), both before and after cryopreservation using cryoprotectant media supplemented with either egg yolk or soy lecithin. Comparing the equivalency of the two phospholipid cryopreservation supplements with regard to postthaw functional parameters demonstrated that there were no statistically significant differences between the two supplements for [1] recovery of motile sperm, [2] maintenance of sperm cell morphology, [3] maintenance of the ability of sperm to bind to hyaluronate in vitro, or [4] maintenance of sperm DNA integrity.

Influences of thermal acclimation and acute temperature change on the motility of epithelial wound-healing cells (keratocytes) of tropical, temperate and Antarctic fish.

PubMed

Ream, Rachael A; Theriot, Julie A; Somero, George N

2003-12-01

The ability to heal superficial wounds is an important element in an organism's repertoire of adaptive responses to environmental stress. In fish, motile cells termed keratocytes are thought to play important roles in the wound-healing process. Keratocyte motility, like other physiological rate processes, is likely to be dependent on temperature and to show adaptive variation among differently thermally adapted species. We have quantified the effects of acute temperature change and thermal acclimation on actin-based keratocyte movement in primary cultures of keratocytes from four species of teleost fish adapted to widely different thermal conditions: two eurythermal species, the longjaw mudsucker Gillichthys mirabilis (environmental temperature range of approximately 10-37 degrees C) and a desert pupfish, Cyprinodon salinus (10-40 degrees C), and two species from stable thermal environments, an Antarctic notothenioid, Trematomus bernacchii (-1.86 degrees C), and a tropical clownfish, Amphiprion percula (26-30 degrees C). For all species, keratocyte speed increased with increasing temperature. G. mirabilis and C. salinus keratocytes reached maximal speeds at 25 degrees C and 35 degrees C, respectively, temperatures within the species' normal thermal ranges. Keratocytes of the stenothermal species continued to increase in speed as temperature increased above the species' normal temperature ranges. The thermal limits of keratocyte motility appear to exceed those of whole-organism thermal tolerance, notably in the case of T. bernacchii. Keratocytes of T. bernacchii survived supercooling to -6 degrees C and retained motility at temperatures as high as 20 degrees C. Mean keratocyte speed was conserved at physiological temperatures for the three temperate and tropical species, which suggests that a certain rate of motility is advantageous for wound healing. However, there was no temperature compensation in speed of movement for keratocytes of the Antarctic fish, which have extremely slow rates of movement at physiological temperatures. Keratocytes from all species moved in a persistent, unidirectional manner at low temperatures but at higher temperatures began to take more circular or less-persistent paths. Thermal acclimation affected the persistence and turning magnitude of keratocytes, with warmer acclimations generally yielding more persistent cells that followed straighter paths. However, acclimation did not alter the effect of experimental temperature on cellular speed. These findings suggest that more than one temperature-sensitive mechanism may govern cell motility: the rate-limiting process(es) responsible for speed is distinct from the mechanism(s) underlying directionality and persistence. Keratocytes represent a useful study system for evaluating the effects of temperature at the cellular level and for studying adaptive variation in actin-based cellular movement and capacity for wound healing.

Measuring Borrelia burgdorferi Motility and Chemotaxis.

PubMed

Zhang, Kai; Li, Chunhao

2018-01-01

Swimming plate, cell motion tracking, and capillary tube assays are very useful tools to quantitatively measure bacterial motility and chemotaxis. These methods were modified and applied to study Borrelia burgdorferi motility and chemotaxis. By using these methods, numerous motility and chemotaxis mutants have been characterized and several chemoattractants were identified. With the assistance of these tools, the role of motility and chemotaxis in the pathogenicity of B. burgdorferi has been established. In addition, these tools also facilitate the study of motility and chemotaxis in other spirochetes.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takahashi, Atsushi; Graduate School of Science and Engineering, Saitama University, Saitama 338-8570; Green Tea Laboratory, Saitama Prefectural Agriculture and Forestry Research Center, Saitama 358-0042

Highlights: •EGCG reduced cell motility of highly metastatic human lung cancer cells. •EGCG increased cell stiffness of the cells, indicating the inhibition of phenotypes of EMT. •EGCG inhibited expression of vimentin and Slug in the cells at the leading edge of scratch. •Treatment of MβCD increased cell stiffness, and inhibited cell motility and vimentin expression. •Inhibition of EMT phenotypes with EGCG is a mechanism-based inhibition of cancer metastasis. -- Abstract: Cell motility and cell stiffness are closely related to metastatic activity of cancer cells. (−)-Epigallocatechin gallate (EGCG) has been shown to inhibit spontaneous metastasis of melanoma cell line into themore » lungs of mice, so we studied the effects of EGCG on cell motility, cell stiffness, and expression of vimentin and Slug, which are molecular phenotypes of epithelial–mesenchymal transition (EMT). Treatments of human non-small cell lung cancer cell lines H1299 and Lu99 with 50 and 100 μM EGCG reduced cell motility to 67.5% and 43.7% in H1299, and 71.7% and 31.5% in Lu99, respectively in in vitro wound healing assay. Studies on cell stiffness using atomic force microscope (AFM) revealed that treatment with 50 μM EGCG increased Young’s modulus of H1299 from 1.24 to 2.25 kPa and that of Lu99 from 1.29 to 2.28 kPa, showing a 2-fold increase in cell stiffness, i.e. rigid elasticity of cell membrane. Furthermore, treatment with 50 μM EGCG inhibited high expression of vimentin and Slug in the cells at a leading edge of scratch. Methyl-β-cyclodextrin, a reagent to deplete cholesterol in plasma membrane, showed inhibition of EMT phenotypes similar that by EGCG, suggesting that EGCG induces inhibition of EMT phenotypes by alteration of membrane organization.« less

Bacteria rolling: motilities of rosette colonies in Caulobacter crescentus

NASA Astrophysics Data System (ADS)

Zeng, Yu; Liu, Bin

2016-11-01

The aquatic bacterium Caulobacter crescentus has two life cycle stages with distinct motilities: freely swimming swarmer cells and immotile stalked cells. Here, we show a new type of movement performed by freely suspended rosettes, spontaneous aggregates of stalked cells aligned radially relative to each other. Reproductive rosette members generate predivisional daughter cells with flagella, inducing rotations of the rosette as a whole. Such rotations exhibit dynamic angular velocities and lead to intermittent linear movements along liquid-solid interfaces, resembling rolling movements. We reconstructed the translational and rotational dynamics of the rosette movements from high-speed filming and long-term tracking. A mechanical model was developed to explain the hydrodynamic mechanism underlying such motilities. Our study illustrated a nontrivial mechanism for clustered bacteria to achieve motilities and sheds light on the adaptive significance of the collective behaviors of microorganisms in complex fluid environments.

Emergence of HGF/SF-Induced Coordinated Cellular Motility

PubMed Central

Zaritsky, Assaf; Natan, Sari; Ben-Jacob, Eshel; Tsarfaty, Ilan

2012-01-01

Collective cell migration plays a major role in embryonic morphogenesis, tissue remodeling, wound repair and cancer invasion. Despite many decades of extensive investigations, only few analytical tools have been developed to enhance the biological understanding of this important phenomenon. Here we present a novel quantitative approach to analyze long term kinetics of bright field time-lapse wound healing. Fully-automated spatiotemporal measures and visualization of cells' motility and implicit morphology were proven to be sound, repetitive and highly informative compared to single-cell tracking analysis. We study cellular collective migration induced by tyrosine kinase-growth factor signaling (Met-Hepatocyte Growth Factor/Scatter Factor (HGF/SF)). Our quantitative approach is applied to demonstrate that collective migration of the adenocarcinoma cell lines is characterized by simple morpho-kinetics. HGF/SF induces complex morpho-kinetic coordinated collective migration: cells at the front move faster and are more spread than those further away from the wound edge. As the wound heals, distant cells gradually accelerate and enhance spread and elongation –resembling the epithelial to mesenchymal transition (EMT), and then the cells become more spread and maintain higher velocity than cells located closer to the wound. Finally, upon wound closure, front cells halt, shrink and round up (resembling mesenchymal to epithelial transition (MET) phenotype) while distant cells undergo the same process gradually. Met inhibition experiments further validate that Met signaling dramatically alters the morpho-kinetic dynamics of the healing wound. Machine-learning classification was applied to demonstrate the generalization of our findings, revealing even subtle changes in motility patterns induced by Met-inhibition. It is concluded that activation of Met-signaling induces an elaborated model in which cells lead a coordinated increased motility along with gradual differentiation-based collective cell motility dynamics. Our quantitative phenotypes may guide future investigation on the molecular and cellular mechanisms of tyrosine kinase-induced coordinate cell motility and morphogenesis in metastasis. PMID:22970283

Modular control of endothelial sheet migration

PubMed Central

Vitorino, Philip; Meyer, Tobias

2008-01-01

Growth factor-induced migration of endothelial cell monolayers enables embryonic development, wound healing, and angiogenesis. Although collective migration is widespread and therapeutically relevant, the underlying mechanism by which cell monolayers respond to growth factor, sense directional signals, induce motility, and coordinate individual cell movements is only partially understood. Here we used RNAi to identify 100 regulatory proteins that enhance or suppress endothelial sheet migration into cell-free space. We measured multiple live-cell migration parameters for all siRNA perturbations and found that each targeted protein primarily regulates one of four functional outputs: cell motility, directed migration, cell–cell coordination, or cell density. We demonstrate that cell motility regulators drive random, growth factor-independent motility in the presence or absence of open space. In contrast, directed migration regulators selectively transduce growth factor signals to direct cells along the monolayer boundary toward open space. Lastly, we found that regulators of cell–cell coordination are growth factor-independent and reorient randomly migrating cells inside the sheet when boundary cells begin to migrate. Thus, cells transition from random to collective migration through a modular control system, whereby growth factor signals convert boundary cells into pioneers, while cells inside the monolayer reorient and follow pioneers through growth factor-independent migration and cell–cell coordination. PMID:19056882

Rab coupling protein mediated endosomal recycling of N-cadherin influences cell motility.

PubMed

Lindsay, Andrew J; McCaffrey, Mary W

2017-12-01

Rab coupling protein (RCP) is a Rab GTPase effector that functions in endosomal recycling. The RCP gene is frequently amplified in breast cancer, leading to increased cancer aggressiveness. Furthermore, RCP enhances the motility of ovarian cancer cells by coordinating the recycling of α5β1 integrin and EGF receptor to the leading edge of migrating cells. Here we report that RCP also influences the motility of lung adenocarcinoma cells. Knockdown of RCP inhibits the motility of A549 cells in 2D and 3D migration assays, while its overexpression enhances migration in these assays. Depletion of RCP leads to a reduction in N-cadherin protein levels, which could be restored with lysosomal inhibitors. Trafficking assays revealed that RCP knockdown inhibits the return of endocytosed N-cadherin to the cell surface. We propose that RCP regulates the endosomal recycling of N-cadherin, and in its absence N-cadherin is diverted to the degradative pathway. The increased aggressiveness of tumour cells that overexpress RCP may be due to biased recycling of N-cadherin in metastatic cancer cells.

Androgen receptor and chemokine receptors 4 and 7 form a signaling axis to regulate CXCL12-dependent cellular motility.

PubMed

Hsiao, Jordy J; Ng, Brandon H; Smits, Melinda M; Wang, Jiahui; Jasavala, Rohini J; Martinez, Harryl D; Lee, Jinhee; Alston, Jhullian J; Misonou, Hiroaki; Trimmer, James S; Wright, Michael E

2015-03-31

Identifying cellular signaling pathways that become corrupted in the presence of androgens that increase the metastatic potential of organ-confined tumor cells is critical to devising strategies capable of attenuating the metastatic progression of hormone-naïve, organ-confined tumors. In localized prostate cancers, gene fusions that place ETS-family transcription factors under the control of androgens drive gene expression programs that increase the invasiveness of organ-confined tumor cells. C-X-C chemokine receptor type 4 (CXCR4) is a downstream target of ERG, whose upregulation in prostate-tumor cells contributes to their migration from the prostate gland. Recent evidence suggests that CXCR4-mediated proliferation and metastasis of tumor cells is regulated by CXCR7 through its scavenging of chemokine CXCL12. However, the role of androgens in regulating CXCR4-mediated motility with respect to CXCR7 function in prostate-cancer cells remains unclear. Immunocytochemistry, western blot, and affinity-purification analyses were used to study how androgens influenced the expression, subcellular localization, and function of CXCR7, CXCR4, and androgen receptor (AR) in LNCaP prostate-tumor cells. Moreover, luciferase assays and quantitative polymerase chain reaction (qPCR) were used to study how chemokines CXCL11 and CXCL12 regulate androgen-regulated genes (ARGs) in LNCaP prostate-tumor cells. Lastly, cell motility assays were carried out to determine how androgens influenced CXCR4-dependent motility through CXCL12. Here we show that, in the LNCaP prostate-tumor cell line, androgens coordinate the expression of CXCR4 and CXCR7, thereby promoting CXCL12/CXCR4-mediated cell motility. RNA interference experiments revealed functional interactions between AR and CXCR7 in these cells. Co-localization and affinity-purification experiments support a physical interaction between AR and CXCR7 in LNCaP cells. Unexpectedly, CXCR7 resided in the nuclear compartment and modulated AR-mediated transcription. Moreover, androgen-mediated cell motility correlated positively with the co-localization of CXCR4 and CXCR7 receptors, suggesting that cell migration may be linked to functional CXCR4/CXCR7 heterodimers. Lastly, CXCL12-mediated cell motility was CXCR7-dependent, with CXCR7 expression required for optimal expression of CXCR4 protein. Overall, our results suggest that inhibition of CXCR7 function might decrease the metastatic potential of organ-confined prostate cancers.

Characterisation of chicken TES and its role in cell spreading and motility.

PubMed

Griffith, Elen; Coutts, Amanda S; Black, Donald M

2004-03-01

Previously we identified TES as a candidate tumour suppressor gene that is located at human chromosome 7q31.1. More recently, we and others have shown TES to encode a novel LIM domain protein that localises to focal adhesions. Here, we present the cloning and functional analysis of the chicken orthologue of TES, cTES. The TES proteins are highly conserved between chicken and human, showing 89% identity at the amino acid level. We show that the cTES protein localised at focal adhesions, actin stress fibres, and sites of cell-cell contact, and GST-cTES can pull-down zyxin and actin. To investigate a functional role for cTES, we looked at the effect of its overexpression on cell spreading and cell motility. Cells overexpressing cTES showed increased cell spreading on fibronectin, and decreased cell motility, compared to RCAS vector transfected control cells. The data from our studies with cTES support our previous findings with human TES and further implicate TES as a member of a complex of proteins that function together to regulate cell adhesion and additionally demonstrate a role for TES in cell motility. Copyright 2004 Wiley-Liss, Inc.

Characterization of Pro-Inflammatory Flagellin Proteins Produced by Lactobacillus ruminis and Related Motile Lactobacilli

PubMed Central

Neville, B. Anne; Forde, Brian M.; Claesson, Marcus J.; Darby, Trevor; Coghlan, Avril; Nally, Kenneth; Ross, R. Paul; O’Toole, Paul W.

2012-01-01

Lactobacillus ruminis is one of at least twelve motile but poorly characterized species found in the genus Lactobacillus. Of these, only L. ruminis has been isolated from mammals, and this species may be considered as an autochthonous member of the gastrointestinal microbiota of humans, pigs and cows. Nine L. ruminis strains were investigated here to elucidate the biochemistry and genetics of Lactobacillus motility. Six strains isolated from humans were non-motile while three bovine isolates were motile. A complete set of flagellum biogenesis genes was annotated in the sequenced genomes of two strains, ATCC25644 (human isolate) and ATCC27782 (bovine isolate), but only the latter strain produced flagella. Comparison of the L. ruminis and L. mali DSM20444T motility loci showed that their genetic content and gene-order were broadly similar, although the L. mali motility locus was interrupted by an 11.8 Kb region encoding rhamnose utilization genes that is absent from the L. ruminis motility locus. Phylogenetic analysis of 39 motile bacteria indicated that Lactobacillus motility genes were most closely related to those of motile carnobacteria and enterococci. Transcriptome analysis revealed that motility genes were transcribed at a significantly higher level in motile L. ruminis ATCC27782 than in non-motile ATCC25644. Flagellin proteins were isolated from L. ruminis ATCC27782 and from three other Lactobacillus species, while recombinant flagellin of aflagellate L. ruminis ATCC25644 was expressed and purified from E. coli. These native and recombinant Lactobacillus flagellins, and also flagellate L. ruminis cells, triggered interleukin-8 production in cultured human intestinal epithelial cells in a manner suppressed by short interfering RNA directed against Toll-Like Receptor 5. This study provides genetic, transcriptomic, phylogenetic and immunological insights into the trait of flagellum-mediated motility in the lactobacilli. PMID:22808200

Clinically relevant enhancement of human sperm motility using compounds with reported phosphodiesterase inhibitor activity

PubMed Central

Tardif, Steve; Madamidola, Oladipo A.; Brown, Sean G.; Frame, Lorna; Lefièvre, Linda; Wyatt, Paul G.; Barratt, Christopher L.R.; Martins Da Silva, Sarah J.

2014-01-01

STUDY QUESTION Can we identify compound(s) with reported phosphodiesterase inhibitor (PDEI) activity that could be added to human spermatozoa in vitro to enhance their motility without compromising other sperm functions? SUMMARY ANSWER We have identified several compounds that produce robust and effective stimulation of sperm motility and, importantly, have a positive response on patient samples. WHAT IS KNOWN ALREADY For >20 years, the use of non-selective PDEIs, such as pentoxifylline, has been known to influence the motility of human spermatozoa; however, conflicting results have been obtained. It is now clear that human sperm express several different phosphodiesterases and these are compartmentalized at different regions of the cells. By using type-specific PDEIs, differential modulation of sperm motility may be achieved without adversely affecting other functions such as the acrosome reaction (AR). STUDY DESIGN, SIZE, DURATION This was a basic medical research study examining sperm samples from normozoospermic donors and subfertile patients attending the Assisted Conception Unit (ACU), Ninewells Hospital Dundee for diagnostic semen analysis, IVF and ICSI. Phase 1 screened 43 commercially available compounds with reported PDEI activity to identify lead compounds that stimulate sperm motility. Samples were exposed (20 min) to three concentrations (1, 10 and 100 µM) of compound, and selected candidates (n = 6) progressed to Phase 2, which provided a more comprehensive assessment using a battery of in vitro sperm function tests. PARTICIPANTS/MATERIALS, SETTING, METHODS All healthy donors and subfertile patients were recruited at the Medical Research Institute, University of Dundee and ACU, Ninewells Hospital Dundee (ethical approval 08/S1402/6). In Phase 1, poor motility cells recovered from the 40% interface of the discontinuous density gradient were used as surrogates for patient samples. Pooled samples from three to four different donors were utilized in order to reduce variability and increase the number of cells available for simultaneous examination of multiple compounds. During Phase 2 testing, semen samples from 23 patients attending for either routine diagnostic andrology assessment or IVF/ICSI were prepared and exposed to selected compounds. Additionally, 48 aliquots of prepared samples, surplus to clinical use, were examined from IVF (n = 32) and ICSI (n = 16) patients to further determine the effects of selected compounds under clinical conditions of treatment. Effects of compounds on sperm motility were assessed by computer-assisted sperm analysis. A modified Kremer test using methyl cellulose was used to assess sperm functional ability to penetrate into viscous media. Sperm acrosome integrity and induction of apoptosis were assessed using the acrosomal content marker PSA-FITC and annexin V kit, respectively. MAIN RESULTS AND THE ROLE OF CHANCE In Phase 1, six compounds were found to have a strong effect on poor motility samples with a magnitude of response of ≥60% increase in percentage total motility. Under capacitating and non-capacitating conditions, these compounds significantly (P ≤ 0.05) increased the percentage of total and progressive motility. Furthermore, these compounds enhanced penetration into a cervical mucus substitute (P ≤ 0.05). Finally, the AR was not significantly induced and these compounds did not significantly increase the externalization of phosphatidylserine (P = 0.6, respectively). In general, the six compounds maintained the stimulation of motility over long periods of time (180 min) and their effects were still observed after their removal. In examinations of clinical samples, there was a general observation of a more significant stimulation of sperm motility in samples with lower baseline motility. In ICSI samples, compounds #26, #37 and #38 were the most effective at significantly increasing total motility (88, 81 and 79% of samples, respectively) and progressive motility (94, 93 and 81% of samples, respectively). In conclusion, using a two-phased drug discovery screening approach including the examination of clinical samples, 3/43 compounds were identified as promising candidates for further study. LIMITATIONS, REASONS FOR CAUTION This is an in vitro study and caution must be taken when extrapolating the results. Data for patients were from one assessment and thus the robustness of responses needs to be established. The n values for ICSI samples were relatively small. WIDER IMPLICATIONS OF THE FINDINGS We have systematically screened and identified several compounds that have robust and effective stimulation (i.e. functional significance with longevity and no toxicity) of total and progressive motility under clinical conditions of treatment. These compounds could be clinical candidates with possibilities in terms of assisted reproductive technology options for current or future patients affected by asthenozoospermia or oligoasthenozoospermia. STUDY FUNDING/COMPETING INTEREST(S) This study was funded primarily by the MRC (DPFS) but with additional funding from the Wellcome Trust, Tenovus (Scotland), University of Dundee, NHS Tayside and Scottish Enterprise. The authors have no competing interests. A patent (#WO2013054111A1) has been published containing some of the information presented in this manuscript. PMID:25124668

Gene position in a long operon governs motility development in Bacillus subtilis

PubMed Central

Cozy, Loralyn M.; Kearns, Daniel B.

2010-01-01

Growing cultures of Bacillus subtilis bifurcate into subpopulations of motile individuals and non-motile chains of cells that are differentiated at the level of gene expression. The motile cells are ON and the chaining cells are OFF for transcription that depends on RNA polymerase and the alternative sigma factor σD. Here we show that chaining cells were OFF for σD-dependent gene expression because σD levels fell below a threshold, and σD activity was inhibited by the anti-sigma factor FlgM. The probability that σD exceeded the threshold was governed by the position of the sigD genes. The proportion of ON cells increased when sigD was artificially moved forward in the 27kb fla/che operon. In addition, we identified a new σD-dependent promoter that increases sigD expression and may provide positive feedback to stabilize the ON state. Finally, we demonstrate that ON/OFF motility states in B. subtilis are a form of development because mosaics of stable and differentiated epigenotypes were evident when the normally dispersed bacteria were forced to grow in one dimension. PMID:20233303

miR-145-dependent targeting of junctional adhesion molecule A and modulation of fascin expression are associated with reduced breast cancer cell motility and invasiveness.

PubMed

Götte, M; Mohr, C; Koo, C-Y; Stock, C; Vaske, A-K; Viola, M; Ibrahim, S A; Peddibhotla, S; Teng, Y H-F; Low, J-Y; Ebnet, K; Kiesel, L; Yip, G W

2010-12-16

Micro RNAs are small non-coding RNAs, which regulate fundamental cellular and developmental processes at the transcriptional and translational level. In breast cancer, miR-145 expression is downregulated compared with healthy control tissue. As several predicted targets of miR-145 potentially regulate cell motility, we aimed at investigating a potential role for miR-145 in breast cancer cell motility and invasiveness. Assisted by Affymetrix array technology, we demonstrate that overexpression of miR-145 in MDA-MB-231, MCF-7, MDA-MB-468 and SK-BR-3 breast cancer cells and in Ishikawa endometrial carcinoma cells leads to a downregulation of the cell-cell adhesion protein JAM-A and of the actin bundling protein fascin. Moreover, podocalyxin and Serpin E1 mRNA levels were downregulated, and gamma-actin, transgelin and MYL9 were upregulated upon miR-145 overexpression. These miR-145-dependent expression changes drastically decreased cancer cell motility, as revealed by time-lapse video microscopy, scratch wound closure assays and matrigel invasion assays. Immunofluorescence microscopy demonstrated restructuring of the actin cytoskeleton and a change in cell morphology by miR-145 overexpression, resulting in a more cortical actin distribution, and reduced actin stress fiber and filopodia formation. Nuclear rotation was observed in 10% of the pre-miR-145 transfected MDA-MB-231 cells, accompanied by a reduction of perinuclear actin. Luciferase activation assays confirmed direct miR-145-dependent regulation of the 3'UTR of JAM-A, whereas siRNA-mediated knockdown of JAM-A expression resulted in decreased motility and invasiveness of MDA-MB-231 and MCF-7 breast cancer cells. Our data identify JAM-A and fascin as novel targets of miR-145, firmly establishing a role for miR-145 in modulating breast cancer cell motility. Our data provide a rationale for future miR-145-targeted approaches of antimetastatic cancer therapy.

Analysis of a Spontaneous Non-Motile and Avirulent Mutant Shows That FliM Is Required for Full Endoflagella Assembly in Leptospira interrogans.

PubMed

Fontana, Célia; Lambert, Ambroise; Benaroudj, Nadia; Gasparini, David; Gorgette, Olivier; Cachet, Nathalie; Bomchil, Natalia; Picardeau, Mathieu

2016-01-01

Pathogenic Leptospira strains are responsible for leptospirosis, a worldwide emerging zoonotic disease. These spirochetes are unique amongst bacteria because of their corkscrew-like cell morphology and their periplasmic flagella. Motility is reported as an important virulence determinant, probably favoring entry and dissemination of pathogenic Leptospira in the host. However, proteins constituting the periplasmic flagella and their role in cell shape, motility and virulence remain poorly described. In this study, we characterized a spontaneous L. interrogans mutant strain lacking motility, correlated with the loss of the characteristic hook-shaped ends, and virulence in the animal model. Whole genome sequencing allowed the identification of one nucleotide deletion in the fliM gene resulting in a premature stop codon, thereby preventing the production of flagellar motor switch protein FliM. Genetic complementation restored cell morphology, motility and virulence comparable to those of wild type cells. Analyses of purified periplasmic flagella revealed a defect in flagella assembly, resulting in shortened flagella compared to the wild type strain. This also correlated with a lower amount of major filament proteins FlaA and FlaB. Altogether, these findings demonstrate that FliM is required for full and correct assembly of the flagella which is essential for motility and virulence.

Alignment of cellular motility forces with tissue flow as a mechanism for efficient wound healing

PubMed Central

Basan, Markus; Elgeti, Jens; Hannezo, Edouard; Rappel, Wouter-Jan; Levine, Herbert

2013-01-01

Recent experiments have shown that spreading epithelial sheets exhibit a long-range coordination of motility forces that leads to a buildup of tension in the tissue, which may enhance cell division and the speed of wound healing. Furthermore, the edges of these epithelial sheets commonly show finger-like protrusions whereas the bulk often displays spontaneous swirls of motile cells. To explain these experimental observations, we propose a simple flocking-type mechanism, in which cells tend to align their motility forces with their velocity. Implementing this idea in a mechanical tissue simulation, the proposed model gives rise to efficient spreading and can explain the experimentally observed long-range alignment of motility forces in highly disordered patterns, as well as the buildup of tensile stress throughout the tissue. Our model also qualitatively reproduces the dependence of swirl size and swirl velocity on cell density reported in experiments and exhibits an undulation instability at the edge of the spreading tissue commonly observed in vivo. Finally, we study the dependence of colony spreading speed on important physical and biological parameters and derive simple scaling relations that show that coordination of motility forces leads to an improvement of the wound healing process for realistic tissue parameters. PMID:23345440

HES6 enhances the motility of alveolar rhabdomyosarcoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wickramasinghe, Caroline M; MRC Laboratory of Molecular Biology, Addenbrooke's Hospital Cambridge, CB2 0QH; Domaschenz, Renae

Absract: HES6, a member of the hairy-enhancer-of-split family of transcription factors, plays multiple roles in myogenesis. It is a direct target of the myogenic transcription factor MyoD and has been shown to regulate the formation of the myotome in development, myoblast cell cycle exit and the organization of the actin cytoskeleton during terminal differentiation. Here we investigate the expression and function of HES6 in rhabdomyosarcoma, a soft tissue tumor which expresses myogenic genes but fails to differentiate into muscle. We show that HES6 is expressed at high levels in the subset of alveolar rhabdomyosarcomas expressing PAX/FOXO1 fusion genes (ARMSp). Knockdownmore » of HES6 mRNA in the ARMSp cell line RH30 reduces proliferation and cell motility. This phenotype is rescued by expression of mouse Hes6 which is insensitive to HES6 siRNA. Furthermore, expression microarray analysis indicates that the HES6 knockdown is associated with a decrease in the levels of Transgelin, (TAGLN), a regulator of the actin cytoskeleton. Knockdown of TAGLN decreases cell motility, whilst TAGLN overexpression rescues the motility defect resulting from HES6 knockdown. These findings indicate HES6 contributes to the pathogenesis of ARMSp by enhancing both proliferation and cell motility.« less

Fennel induces cytotoxic effects against testicular germ cells in mice; evidences for suppressed pre-implantation embryo development.

PubMed

Minas, Aram; Najafi, Gholamreza; Jalali, Ali Shalizar; Razi, Mazdak

2018-05-15

Foeniculum vulgare (FVE; fennel) is an aromatic plant belonging to Umbelliferae family, which is widely used in traditional societies because of its different pharmaceutical properties. To uncover the fennel-derived essential oil (FVEO)-induced effects on male reproductive potential, 24 mature male albino mice were divided into, control, 0.37, 0.75, and 1.5 mg kg -1 FVEO-received groups. Following 35 days, the animals were euthanized and the testicular tissue and sperm samples were collected. The histological alterations, tubular differentiation (TDI), spermiogenesis (SPI) indices, apoptosis ratio, and RNA damage of germinal cells were analyzed. Moreover, the sperm count, motility, viability, chromatin condensation, and DNA fragmentation were assessed. Finally, the pre-implantation embryo development including; the percentage of zygote, 2-cell embryos and blastocysts were assessed. Observations showed that the FVEO, dose dependently, increased histological damages, resulted in germ cells dissociation, depletion, nuclear shrinkage and significantly (P < .05) decreased tubular differentiation and spermiogenesis ratios. Moreover, the FVEO-received animals (more significantly in 1.5 mg kg -1 -received group) exhibited decreased sperm count, viability, and motility and represented enhanced percentage of sperms with decondensed chromatin and DNA fragmentation. Finally, the animals in FVEO-received group showed diminished zygote formation and represented decreased pre-implantation embryo development compared to control animals. In conclusion, our data showed that, FVEO albeit at higher doses, is able to adversely affect cellular DNA and RNA contents, which in turn is able to negatively affect the sperm count and morphology. All these impairments are able to negatively affect the fertilization potential as well as pre-implantation embryo development. © 2018 Wiley Periodicals, Inc.

S100A4 is frequently overexpressed in lung cancer cells and promotes cell growth and cell motility

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Na; Sato, Daisuke; Saiki, Yuriko

2014-05-09

Highlights: • We observed frequent overexpression of S100A4 in lung cancer cell lines. • Knockdown of S100A4 suppressed proliferation in lung cancer cells. • Forced expression of S100A4 accelerated cell motility in lung cancer cells. • PRDM2 was found to be one of the downstream suppressed genes of S100A4. - Abstract: S100A4, a small calcium-binding protein belonging to the S100 protein family, is commonly overexpressed in a variety of tumor types and is widely accepted to associate with metastasis by regulating the motility and invasiveness of cancer cells. However, its biological role in lung carcinogenesis is largely unknown. In thismore » study, we found that S100A4 was frequently overexpressed in lung cancer cells, irrespective of histological subtype. Then we performed knockdown and forced expression of S100A4 in lung cancer cell lines and found that specific knockdown of S100A4 effectively suppressed cell proliferation only in lung cancer cells with S100A4-overexpression; forced expression of S100A4 accelerated cell motility only in S100A4 low-expressing lung cancer cells. PRDM2 and VASH1, identified as novel upregulated genes by microarray after specific knockdown of S100A4 in pancreatic cancer, were also analyzed, and we found that PRDM2 was significantly upregulated after S100A4-knockdown in one of two analyzed S100A4-overexpressing lung cancer cells. Our present results suggest that S100A4 plays an important role in lung carcinogenesis by means of cell proliferation and motility by a pathway similar to that in pancreatic cancer.« less

Changes of Photosynthetic Behaviors and Photoprotection during Cell Transformation and Astaxanthin Accumulation in Haematococcus pluvialis Grown Outdoors in Tubular Photobioreactors.

PubMed

Zhang, Litao; Su, Fang; Zhang, Chunhui; Gong, Fengying; Liu, Jianguo

2016-12-26

The cell transformation from green motile cells to non-motile cells and astaxanthin accumulation can be induced in the green alga Haematococcus pluvialis cultured outdoors. In the initial 3 d of incubation (cell transformation phase), light absorption and photosynthetic electron transport became more efficient. After five days of incubation (astaxanthin accumulation phase), the light absorption per active reaction center (ABS/RC) increased, but the efficiency of electron transport ( ψ o ) and the quantum yield of electron transport ( φ Eo ) decreased with increased time, indicating that the capacity of photosynthetic energy utilization decreased significantly during astaxanthin accumulation, leading to an imbalance between photosynthetic light absorption and energy utilization. It would inevitably aggravate photoinhibition under high light, e.g., at midday. However, the level of photoinhibition in H. pluvialis decreased as the incubation time increased, which is reflected by the fact that F v / F m determined at midday decreased significantly in the initial 3 d of incubation, but was affected very little after seven days of incubation, compared with that determined at predawn. This might be because the non-photochemical quenching, plastid terminal oxidase, photosystem I cyclic electron transport, defensive enzymes and the accumulated astaxanthin can protect cells against photoinhibition.

Changes of Photosynthetic Behaviors and Photoprotection during Cell Transformation and Astaxanthin Accumulation in Haematococcus pluvialis Grown Outdoors in Tubular Photobioreactors

PubMed Central

Zhang, Litao; Su, Fang; Zhang, Chunhui; Gong, Fengying; Liu, Jianguo

2016-01-01

The cell transformation from green motile cells to non-motile cells and astaxanthin accumulation can be induced in the green alga Haematococcus pluvialis cultured outdoors. In the initial 3 d of incubation (cell transformation phase), light absorption and photosynthetic electron transport became more efficient. After five days of incubation (astaxanthin accumulation phase), the light absorption per active reaction center (ABS/RC) increased, but the efficiency of electron transport (ψo) and the quantum yield of electron transport (φEo) decreased with increased time, indicating that the capacity of photosynthetic energy utilization decreased significantly during astaxanthin accumulation, leading to an imbalance between photosynthetic light absorption and energy utilization. It would inevitably aggravate photoinhibition under high light, e.g., at midday. However, the level of photoinhibition in H. pluvialis decreased as the incubation time increased, which is reflected by the fact that Fv/Fm determined at midday decreased significantly in the initial 3 d of incubation, but was affected very little after seven days of incubation, compared with that determined at predawn. This might be because the non-photochemical quenching, plastid terminal oxidase, photosystem I cyclic electron transport, defensive enzymes and the accumulated astaxanthin can protect cells against photoinhibition. PMID:28035956

Thermal stratification hinders gyrotactic micro-organism rising in free-surface turbulence

NASA Astrophysics Data System (ADS)

Lovecchio, Salvatore; Zonta, Francesco; Marchioli, Cristian; Soldati, Alfredo

2017-05-01

Thermal stratification in water bodies influences the exchange of heat, momentum, and chemical species across the air-water interface by modifying the sub-surface turbulence characteristics. Turbulence modifications may in turn prevent small motile algae (phytoplankton, in particular) from reaching the heated surface. We examine how different regimes of stable thermal stratification affect the motion of these microscopic organisms (modelled as gyrotactic self-propelling cells) in a free-surface turbulent channel flow. This archetypal setup mimics an environmentally plausible situation that can be found in lakes and oceans. Results from direct numerical simulations of turbulence coupled with Lagrangian tracking reveal that rising of bottom-heavy self-propelling cells depends strongly on the strength of stratification, especially near the thermocline where high temperature and velocity gradients occur: Here hydrodynamic shear may disrupt directional cell motility and hamper near-surface accumulation. For all gyrotactic re-orientation times considered in this study (spanning two orders of magnitude), we observe a reduction of the cell rising speed and temporary confinement under the thermocline: If re-orientation is fast, cells eventually trespass the thermocline within the simulated time span; if re-orientation is slow, confinement lasts much longer because cells align in the streamwise direction and their vertical swimming is practically annihilated.

Rab proteins: The key regulators of intracellular vesicle transport

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bhuin, Tanmay; Roy, Jagat Kumar, E-mail: jkroy@bhu.ac.in

2014-10-15

Vesicular/membrane trafficking essentially regulates the compartmentalization and abundance of proteins within the cells and contributes in many signalling pathways. This membrane transport in eukaryotic cells is a complex process regulated by a large and diverse array of proteins. A large group of monomeric small GTPases; the Rabs are essential components of this membrane trafficking route. Most of the Rabs are ubiquitously expressed proteins and have been implicated in vesicle formation, vesicle motility/delivery along cytoskeleton elements and docking/fusion at target membranes through the recruitment of effectors. Functional impairments of Rabs affecting transport pathways manifest different diseases. Rab functions are accompanied bymore » cyclical activation and inactivation of GTP-bound and GDP-bound forms between the cytosol and membranes which is regulated by upstream regulators. Rab proteins are characterized by their distinct sub-cellular localization and regulate a wide variety of endocytic, transcytic and exocytic transport pathways. Mutations of Rabs affect cell growth, motility and other biological processes. - Highlights: • Rab proteins regulate different signalling pathways. • Deregulation of Rabs is the fundamental causes of a variety of human diseases. • This paper gives potential directions in developing therapeutic targets. • This paper also gives ample directions for modulating pathways central to normal physiology. • These are the huge challenges for drug discovery and delivery in near future.« less

Cellular Motility--Experiments on Contractile and Motile Mechanisms in the Slime Mould, Physarum Polycephalum

ERIC Educational Resources Information Center

Holmes, R. P.; Stewart, P. R.

1977-01-01

Actin and myosin have now been demonstrated to be important constituents of many eukaryotic cells. Their role is primarily that of a contractile system underlying all aspects of cellular motility. Described here is a simple experimental system to demonstrate quantitatively aspects of motility and its regulation in a slime mold. (Author/MA)

Interplay between type IV pili activity and exopolysaccharides secretion controls motility patterns in single cells of Myxococcus xanthus

PubMed Central

Hu, Wei; Gibiansky, Maxsim L.; Wang, Jing; Wang, Chuandong; Lux, Renate; Li, Yuezhong; Wong, Gerard C. L.; Shi, Wenyuan

2016-01-01

Myxococcus xanthus performs coordinated social motility of cell groups through the extension and retraction of type IV pili (TFP) on solid surfaces, which requires both TFP and exopolysaccharides (EPS). By submerging cells in a liquid medium containing 1% methylcellulose, M. xanthus TFP-driven motility was induced in isolated cells and independently of EPS. We measured and analyzed the movements of cells using community tracking algorithms, which combine single-cell resolution with statistics from large sample populations. Cells without significant multi-cellular social interactions have surprisingly complex behaviors: EPS− cells exhibited a pronounced increase in the tendency to stand vertically and moved with qualitatively different characteristics than other cells. A decrease in the EPS secretion of cells correlates with a higher instantaneous velocity, but with lower directional persistence in trajectories. Moreover, EPS− cells do not adhere to the surface as strongly as wild-type and EPS overproducing cells, and display a greater tendency to have large deviations between the direction of movement and the cell axis, with cell velocity showing only minimal dependence on the direction of movement. The emerging picture is that EPS does not simply provide rheological resistance to a single mechanism but rather that the availability of EPS impacts motility pattern. PMID:26821939

The Wnt5A/Protein Kinase C Pathway Mediates Motility in Melanoma Cells via the Inhibition of Metastasis Suppressors and Initiation of an Epithelial to Mesenchymal Transition*S

PubMed Central

Dissanayake, Samudra K.; Wade, Michael; Johnson, Carrie E.; O’Connell, Michael P.; Leotlela, Poloko D.; French, Amanda D.; Shah, Kavita V.; Hewitt, Kyle J.; Rosenthal, Devin T.; Indig, Fred E.; Jiang, Yuan; Nickoloff, Brian J.; Taub, Dennis D.; Trent, Jeffrey M.; Moon, Randall T.; Bittner, Michael; Weeraratna, Ashani T.

2008-01-01

We have shown that Wnt5A increases the motility of melanoma cells. To explore cellular pathways involving Wnt5A, we compared gain-of-function (WNT5A stable transfectants) versus loss-of-function (siRNA knockdown) of WNT5A by microarray analysis. Increasing WNT5A suppressed the expression of several genes, which were re-expressed after small interference RNA-mediated knockdown of WNT5A. Genes affected by WNT5A include KISS-1, a metastasis suppressor, and CD44, involved in tumor cell homing during metastasis. This could be validated at the protein level using both small interference RNA and recombinant Wnt5A (rWnt5A). Among the genes up-regulated by WNT5A was the gene vimentin, associated with an epithelial to mesenchymal transition (EMT), which involves decreases in E-cadherin, due to up-regulation of the transcriptional repressor, Snail. rWnt5A treatment increases Snail and vimentin expression, and decreases E-cadherin, even in the presence of dominant-negativeTCF4, suggesting that this activation is independent of Wnt/β-catenin signaling. Because Wnt5A can signal via protein kinase C (PKC), the role of PKC in Wnt5A-mediated motility and EMT was also assessed using PKC inhibition and activation studies. Treating cells expressing low levels of Wnt5A with phorbol ester increased Snail expression inhibiting PKC in cells expressing high levels of Wnt5A decreased Snail. Furthermore, inhibition of PKC before Wnt5A treatment blocked Snail expression, implying that Wnt5A can potentiate melanoma metastasis via the induction of EMT in a PKC-dependent manner. PMID:17426020

Azospirillum brasilense Chemotaxis Depends on Two Signaling Pathways Regulating Distinct Motility Parameters

PubMed Central

Mukherjee, Tanmoy; Kumar, Dhivya; Burriss, Nathan; Xie, Zhihong

2016-01-01

ABSTRACT The genomes of most motile bacteria encode two or more chemotaxis (Che) systems, but their functions have been characterized in only a few model systems. Azospirillum brasilense is a motile soil alphaproteobacterium able to colonize the rhizosphere of cereals. In response to an attractant, motile A. brasilense cells transiently increase swimming speed and suppress reversals. The Che1 chemotaxis pathway was previously shown to regulate changes in the swimming speed, but it has a minor role in chemotaxis and root surface colonization. Here, we show that a second chemotaxis system, named Che4, regulates the probability of swimming reversals and is the major signaling pathway for chemotaxis and wheat root surface colonization. Experimental evidence indicates that Che1 and Che4 are functionally linked to coordinate changes in the swimming motility pattern in response to attractants. The effect of Che1 on swimming speed is shown to enhance the aerotactic response of A. brasilense in gradients, likely providing the cells with a competitive advantage in the rhizosphere. Together, the results illustrate a novel mechanism by which motile bacteria utilize two chemotaxis pathways regulating distinct motility parameters to alter movement in gradients and enhance the chemotactic advantage. IMPORTANCE Chemotaxis provides motile bacteria with a competitive advantage in the colonization of diverse niches and is a function enriched in rhizosphere bacterial communities, with most species possessing at least two chemotaxis systems. Here, we identify the mechanism by which cells may derive a significant chemotactic advantage using two chemotaxis pathways that ultimately regulate distinct motility parameters. PMID:27068592

Azospirillum brasilense Chemotaxis Depends on Two Signaling Pathways Regulating Distinct Motility Parameters.

PubMed

Mukherjee, Tanmoy; Kumar, Dhivya; Burriss, Nathan; Xie, Zhihong; Alexandre, Gladys

2016-06-15

The genomes of most motile bacteria encode two or more chemotaxis (Che) systems, but their functions have been characterized in only a few model systems. Azospirillum brasilense is a motile soil alphaproteobacterium able to colonize the rhizosphere of cereals. In response to an attractant, motile A. brasilense cells transiently increase swimming speed and suppress reversals. The Che1 chemotaxis pathway was previously shown to regulate changes in the swimming speed, but it has a minor role in chemotaxis and root surface colonization. Here, we show that a second chemotaxis system, named Che4, regulates the probability of swimming reversals and is the major signaling pathway for chemotaxis and wheat root surface colonization. Experimental evidence indicates that Che1 and Che4 are functionally linked to coordinate changes in the swimming motility pattern in response to attractants. The effect of Che1 on swimming speed is shown to enhance the aerotactic response of A. brasilense in gradients, likely providing the cells with a competitive advantage in the rhizosphere. Together, the results illustrate a novel mechanism by which motile bacteria utilize two chemotaxis pathways regulating distinct motility parameters to alter movement in gradients and enhance the chemotactic advantage. Chemotaxis provides motile bacteria with a competitive advantage in the colonization of diverse niches and is a function enriched in rhizosphere bacterial communities, with most species possessing at least two chemotaxis systems. Here, we identify the mechanism by which cells may derive a significant chemotactic advantage using two chemotaxis pathways that ultimately regulate distinct motility parameters. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

VEGF-A/VEGFR-2 signaling plays an important role for the motility of pancreas cancer cells.

PubMed

Doi, Yosuke; Yashiro, Masakazu; Yamada, Nobuya; Amano, Ryosuke; Noda, Satoru; Hirakawa, Kosei

2012-08-01

Pancreatic cancer is one of the most lethal solid tumors. Vascular endothelial growth factor receptors (VEGFRs) are expressed not only by endothelial cells but also by pancreatic cancer cells. VEGFRs might play an important role for the development of pancreatic cancer cells. The purpose of this study was to evaluate the efficacy of VEGF/VEGFR-2-targeted therapy in pancreatic carcinoma. Five pancreatic carcinoma cell lines were used. The expression level of VEGFR-2 of cancer cells was examined by RT-PCR and Western blot. The effects of VEGFs, bevacizumab as an anti-VEGF antibody, sunitinib as a tyrosine kinase inhibitor against VEGFRs, and VEGF-R2 siRNA on the motility activity of pancreatic cancer cells were examined by invasion assay and wound healing assay. The effect of VEGF, bevacizumab, and sunitinib on the phosphorylation of VEGFR-2 and downstream effecter molecules, MAPK and PI3K, was examined by western blot. Pancreatic cancer cell lines expressed VEGFR-2. VEGF-A significantly increased the motility of pancreas cancer cells, which was inhibited by VEGFR-2 siRNA. Conditioned medium from pancreas cancer cells significantly stimulated the motility of pancreas cancer cells. VEGF/VEGFR inhibitors, bevacizumab and sunitinib, significantly decreased the motility of pancreas cancer cells. VEGFR-2 phosphorylation level of pancreas cancer cells was increased by VEGF-A. Bevacizumab and sunitinib decreased the level of VEGFR-2 phosphorylation, p-ERK, and p-Akt expression. VEGF-A decreased zonula occludens (ZO-1) or ZO-2 expression in pancreas cancer cells. VEGF-A/VEGFR-2 signaling plays an important role in inducing invasion and migration of pancreatic cancer cells.

L1CAM stimulates glioma cell motility and proliferation through the fibroblast growth factor receptor.

PubMed

Mohanan, Vishnu; Temburni, Murali K; Kappes, John C; Galileo, Deni S

2013-04-01

The L1CAM cell adhesion/recognition molecule (L1, CD171) and fibroblast growth factor receptor (FGFR) both are expressed by human high-grade glioma cells, but their potential actions in controlling cell behavior have not been linked. L1 actions in cancer cells have been attributed mainly to integrin receptors, and we demonstrated previously that L1-stimulated glioma cell migration correlates with integrin expression, increased focal adhesion kinase activation and focal complex turnover. Our analyses of datasets revealed FGFR is overexpressed in glioma regardless of grade, while ADAM10 metalloprotease expression increases with glioma grade. Here, we used dominant-negative and short hairpin RNA approaches to inhibit the activation of FGFR1 and expression of L1, respectively. An L1 peptide that inhibits L1-FGFR interaction and PD173074, a chemical inhibitor of FGFR1 activity, also were used to elucidate the involvement of L1-FGFR interactions on glioma cell behavior. Time-lapse cell motility studies and flow cytometry cell cycle analyses showed that L1 operates to increase glioma cell motility and proliferation through FGFR activation. Shutdown of both L1 expression and FGFR activity in glioma cells resulted in a complete termination of cell migration in vitro. These studies show for the first time that soluble L1 ectodomain (L1LE) acts on glioma cells through FGFRs, and that FGFRs are used by glioma cells for increasing motility as well as proliferation in response to activation by L1LE ligand. Thus, effective treatment of high-grade glioma may require simultaneous targeting of L1, FGFRs, and integrin receptors, which would reduce glioma cell motility as well as proliferation.

Endothelial cell motility, coordination and pattern formation during vasculogenesis.

PubMed

Czirok, Andras

2013-01-01

How vascular networks assemble is a fundamental problem of developmental biology that also has medical importance. To explain the organizational principles behind vascular patterning, we must understand how can tissue level structures be controlled through cell behavior patterns like motility and adhesion that, in turn, are determined by biochemical signal transduction processes? We discuss the various ideas that have been proposed as mechanisms for vascular network assembly: cell motility guided by extracellular matrix alignment (contact guidance), chemotaxis guided by paracrine and autocrine morphogens, and multicellular sprouting guided by cell-cell contacts. All of these processes yield emergent patterns, thus endothelial cells can form an interconnected structure autonomously, without guidance from an external pre-pattern. © 2013 Wiley Periodicals, Inc.

Simultaneous measurement of sensor-protein dynamics and motility of a single cell by on-chip microcultivation system

PubMed Central

Inoue, Ippei; Shiomi, Daisuke; Kawagishi, Ikuro; Yasuda, Kenji

2004-01-01

Measurement of the correlation between sensor-protein expression, motility and environmental change is important for understanding the adaptation process of cells during their change of generation. We have developed a novel assay exploiting the on-chip cultivation system, which enabled us to observe the change of the localization of expressed sensor-protein and the motility for generations. Localization of the aspartate sensitive sensor protein at two poles in Escherichia coli decreased quickly after the aspartate was added into the cultivation medium. However, it took more than three generations for recovering the localization after the removal of aspartate from the medium. Moreover, the tumbling frequency was strongly related to the localization of the sensor protein in a cell. The results indicate that the change of the spatial localization of sensor protein, which was inherited for more than three generations, may contribute to cells, motility as the inheritable information. PMID:15119953

Quantitative in vivo whole genome motility screen reveals novel therapeutic targets to block cancer metastasis.

PubMed

Stoletov, Konstantin; Willetts, Lian; Paproski, Robert J; Bond, David J; Raha, Srijan; Jovel, Juan; Adam, Benjamin; Robertson, Amy E; Wong, Francis; Woolner, Emma; Sosnowski, Deborah L; Bismar, Tarek A; Wong, Gane Ka-Shu; Zijlstra, Andries; Lewis, John D

2018-06-14

Metastasis is the most lethal aspect of cancer, yet current therapeutic strategies do not target its key rate-limiting steps. We have previously shown that the entry of cancer cells into the blood stream, or intravasation, is highly dependent upon in vivo cancer cell motility, making it an attractive therapeutic target. To systemically identify genes required for tumor cell motility in an in vivo tumor microenvironment, we established a novel quantitative in vivo screening platform based on intravital imaging of human cancer metastasis in ex ovo avian embryos. Utilizing this platform to screen a genome-wide shRNA library, we identified a panel of novel genes whose function is required for productive cancer cell motility in vivo, and whose expression is closely associated with metastatic risk in human cancers. The RNAi-mediated inhibition of these gene targets resulted in a nearly total (>99.5%) block of spontaneous cancer metastasis in vivo.

Persistent enhancement of bacterial motility increases tumor penetration.

PubMed

Thornlow, Dana N; Brackett, Emily L; Gigas, Jonathan M; Van Dessel, Nele; Forbes, Neil S

2015-11-01

Motile bacteria can overcome the transport limitations that hinder many cancer therapies. Active bacteria can penetrate through tissue to deliver treatment to resistant tumor regions. Bacterial therapy has had limited success, however, because this motility is heterogeneous, and within a population many individuals are non-motile. In human trials, heterogeneity led to poor dispersion and incomplete tumor colonization. To address these problems, a swarm-plate selection method was developed to increase swimming velocity. Video microscopy was used to measure the velocity distribution of selected bacteria and a microfluidic tumor-on-a-chip device was used to measure penetration through tumor cell masses. Selection on swarm plates increased average velocity fourfold, from 4.9 to 18.7 μm/s (P < 0.05) and decreased the number of non-motile individuals from 51% to 3% (P < 0.05). The selected phenotype was both robust and stable. Repeating the selection process consistently increased velocity and eliminated non-motile individuals. When selected strains were cryopreserved and subcultured for 30.1 doublings, the high-motility phenotype was preserved. In the microfluidic device, selected Salmonella penetrated deeper into cell masses than unselected controls. By 10 h after inoculation, control bacteria accumulated in the front 30% of cell masses, closest to the flow channel. In contrast, selected Salmonella accumulated in the back 30% of cell masses, farthest from the channel. Selection increased the average penetration distance from 150 to 400 μm (P < 0.05). This technique provides a simple and rapid method to generate high-motility Salmonella that has increased penetration and potential for greater tumor dispersion and clinical efficacy. © 2015 Wiley Periodicals, Inc.

The Pseudomonas aeruginosa ribbon-helix-helix DNA-binding protein AlgZ (AmrZ) controls twitching motility and biogenesis of type IV pili.

PubMed

Baynham, Patricia J; Ramsey, Deborah M; Gvozdyev, Borys V; Cordonnier, Ellen M; Wozniak, Daniel J

2006-01-01

Pseudomonas aeruginosa is an opportunistic pathogen that is commonly found in water and soil. In order to colonize surfaces with low water content, P. aeruginosa utilizes a flagellum-independent form of locomotion called twitching motility, which is dependent upon the extension and retraction of type IV pili. This study demonstrates that AlgZ, previously identified as a DNA-binding protein absolutely required for transcription of the alginate biosynthetic operon, is required for twitching motility. AlgZ may be required for the biogenesis or function of type IV pili in twitching motility. Transmission electron microscopy analysis of an algZ deletion in nonmucoid PAO1 failed to detect surface pili. To examine expression and localization of PilA (the major pilin subunit), whole-cell extracts and cell surface pilin preparations were analyzed by Western blotting. While the PilA levels present in whole-cell extracts were similar for wild-type P. aeruginosa and P. aeruginosa with the algZ deletion, the amount of PilA on the surface of the cells was drastically reduced in the algZ mutant. Analysis of algZ and algD mutants indicates that the DNA-binding activity of AlgZ is essential for the regulation of twitching motility and that this is independent of the role of AlgZ in alginate expression. These data show that AlgZ DNA-binding activity is required for twitching motility independently of its role in alginate production and that this involves the surface localization of type IV pili. Given this new role in twitching motility, we propose that algZ (PA3385) be designated amrZ (alginate and motility regulator Z).

Balance between cell-substrate adhesion and myosin contraction determines the frequency of motility initiation in fish keratocytes.

PubMed

Barnhart, Erin; Lee, Kun-Chun; Allen, Greg M; Theriot, Julie A; Mogilner, Alex

2015-04-21

Cells are dynamic systems capable of spontaneously switching among stable states. One striking example of this is spontaneous symmetry breaking and motility initiation in fish epithelial keratocytes. Although the biochemical and mechanical mechanisms that control steady-state migration in these cells have been well characterized, the mechanisms underlying symmetry breaking are less well understood. In this work, we have combined experimental manipulations of cell-substrate adhesion strength and myosin activity, traction force measurements, and mathematical modeling to develop a comprehensive mechanical model for symmetry breaking and motility initiation in fish epithelial keratocytes. Our results suggest that stochastic fluctuations in adhesion strength and myosin localization drive actin network flow rates in the prospective cell rear above a critical threshold. Above this threshold, high actin flow rates induce a nonlinear switch in adhesion strength, locally switching adhesions from gripping to slipping and further accelerating actin flow in the prospective cell rear, resulting in rear retraction and motility initiation. We further show, both experimentally and with model simulations, that the global levels of adhesion strength and myosin activity control the stability of the stationary state: The frequency of symmetry breaking decreases with increasing adhesion strength and increases with increasing myosin contraction. Thus, the relative strengths of two opposing mechanical forces--contractility and cell-substrate adhesion--determine the likelihood of spontaneous symmetry breaking and motility initiation.

Autocrine motility factor (neuroleukin, phosphohexose isomerase) induces cell movement through 12-lipoxygenase-dependent tyrosine phosphorylation and serine dephosphorylation events.

PubMed

Timár, J; Tóth, S; Tóvári, J; Paku, S; Raz, A

1999-01-01

Autocrine motility factor (AMF) is one of the motility cytokines regulating tumor cell migration, therefore identification of the signaling pathway coupled with it has critical importance. Previous studies revealed several elements of this pathway predominated by lipoxygenase-PKC activations but the role for tyrosine kinases remained questionable. Motility cytokines frequently have mitogenic effect as well, producing activation of overlapping signaling pathways therefore we have used B16a melanoma cells as models where AMF has exclusive motility effect. Our studies revealed that in B16a cells AMF initiated rapid (1-5 min) activation of the protein tyrosine kinase (PTK) cascade inducing phosphorylation of 179, 125, 95 and 40/37 kD proteins which was mediated by upstream cyclo- and lipoxygenases. The phosphorylated proteins were localized to the cortical actin-stress fiber attachment zones in situ by confocal microscopy. On the other hand, AMF receptor activation induced significant decrease in overall serine-phosphorylation level of cellular proteins accompanied by serine phosphorylation of 200, 90, 78 and 65 kd proteins. The decrease in serine phosphorylation was independent of PTKs, PKC as well as cyclo- and lipoxygenases. However, AMF induced robust translocation of PKCalpha to the stress fibers and cortical actin suggesting a critical role for this kinase in the generation of the motility signal. Based on the significant decrease in serine phosphorylation after AMF stimulus in B16a cells we postulated the involvement of putative serine/threonine phosphatase(s) upstream lipoxygenase and activation of the protein tyrosine kinase cascade downstream cyclo- and lipoxygenase(s) in the previously identified autocrine motility signal.

Motor-driven intracellular transport powers bacterial gliding motility

PubMed Central

Sun, Mingzhai; Wartel, Morgane; Cascales, Eric; Shaevitz, Joshua W.; Mignot, Tâm

2011-01-01

Protein-directed intracellular transport has not been observed in bacteria despite the existence of dynamic protein localization and a complex cytoskeleton. However, protein trafficking has clear potential uses for important cellular processes such as growth, development, chromosome segregation, and motility. Conflicting models have been proposed to explain Myxococcus xanthus motility on solid surfaces, some favoring secretion engines at the rear of cells and others evoking an unknown class of molecular motors distributed along the cell body. Through a combination of fluorescence imaging, force microscopy, and genetic manipulation, we show that membrane-bound cytoplasmic complexes consisting of motor and regulatory proteins are directionally transported down the axis of a cell at constant velocity. This intracellular motion is transmitted to the exterior of the cell and converted to traction forces on the substrate. Thus, this study demonstrates the existence of a conserved class of processive intracellular motors in bacteria and shows how these motors have been adapted to produce cell motility. PMID:21482768

COX inhibition excites enteric nerves that affect motility, alkaline secretion, and permeability in rat duodenum.

PubMed

Nylander, O; Hällgren, A; Sababi, M

2001-11-01

In anesthetized rats, the cyclooxygenase (COX) inhibitor indomethacin induces duodenal motility, increases duodenal mucosal alkaline secretion (DMAS), and evokes a transient increase in duodenal paracellular permeability (DPP). To examine whether enteric nerves influence these responses, the duodenum was perfused with lidocaine. Motility was assessed by measuring intraluminal pressure, and DPP was determined as blood-to-lumen clearance of (51)Cr-EDTA. DMAS was assessed by titration. In control animals, few contractions occurred during saline perfusion and lidocaine did not alter this condition. Perfusion with 0.03-0.1% lidocaine did not affect DMAS or DPP whereas 0.3-1% lidocaine reduced DMAS and increased DPP. Indomethacin induced motility and doubled DMAS. Application of 0.03% lidocaine on the duodenal serosa reduced motility and DMAS whereas 0.03% lidocaine applied luminally inhibited DMAS only. Higher concentrations of lidocaine abolished the increase in DMAS and changed the motility pattern to numerous low-amplitude contractions, the latter effect being blocked by iloprost. The lidocaine-induced increases in DPP were markedly higher than in controls. We conclude that indomethacin activates enteric nerves that induce motility, increase DMAS, and decrease DPP.

Cranial neural crest recycle surface integrins in a substratum-dependent manner to promote rapid motility.

PubMed

Strachan, Lauren R; Condic, Maureen L

2004-11-08

Cell migration is essential for proper development of numerous structures derived from embryonic neural crest cells (NCCs). Although the migratory pathways of NCCs have been determined, the molecular mechanisms regulating NCC motility remain unclear. NCC migration is integrin dependent, and recent work has shown that surface expression levels of particular integrin alpha subunits are important determinants of NCC motility in vitro. Here, we provide evidence that rapid cranial NCC motility on laminin requires integrin recycling. NCCs showed both ligand- and receptor-specific integrin regulation in vitro. On laminin, NCCs accumulated internalized laminin but not fibronectin receptors over 20 min, whereas on fibronectin neither type of receptor accumulated internally beyond 2 min. Internalized laminin receptors colocalized with receptor recycling vesicles and were subsequently recycled back to the cell surface. Blocking receptor recycling with bafilomycin A inhibited NCC motility on laminin, indicating that substratum-dependent integrin recycling is essential for rapid cranial neural crest migration.

Consequences of Location-Dependent Organ of Corti Micro-Mechanics

PubMed Central

Liu, Yanju; Gracewski, Sheryl M.; Nam, Jong-Hoon

2015-01-01

The cochlea performs frequency analysis and amplification of sounds. The graded stiffness of the basilar membrane along the cochlear length underlies the frequency-location relationship of the mammalian cochlea. The somatic motility of outer hair cell is central for cochlear amplification. Despite two to three orders of magnitude change in the basilar membrane stiffness, the force capacity of the outer hair cell’s somatic motility, is nearly invariant over the cochlear length. It is puzzling how actuators with a constant force capacity can operate under such a wide stiffness range. We hypothesize that the organ of Corti sets the mechanical conditions so that the outer hair cell’s somatic motility effectively interacts with the media of traveling waves—the basilar membrane and the tectorial membrane. To test this hypothesis, a computational model of the gerbil cochlea was developed that incorporates organ of Corti structural mechanics, cochlear fluid dynamics, and hair cell electro-physiology. The model simulations showed that the micro-mechanical responses of the organ of Corti are different along the cochlear length. For example, the top surface of the organ of Corti vibrated more than the bottom surface at the basal (high frequency) location, but the amplitude ratio was reversed at the apical (low frequency) location. Unlike the basilar membrane stiffness varying by a factor of 1700 along the cochlear length, the stiffness of the organ of Corti complex felt by the outer hair cell remained between 1.5 and 0.4 times the outer hair cell stiffness. The Y-shaped structure in the organ of Corti formed by outer hair cell, Deiters cell and its phalange was the primary determinant of the elastic reactance imposed on the outer hair cells. The stiffness and geometry of the Deiters cell and its phalange affected cochlear amplification differently depending on the location. PMID:26317521

Pumilacidin-Like Lipopeptides Derived from Marine Bacterium Bacillus sp. Strain 176 Suppress the Motility of Vibrio alginolyticus

PubMed Central

Xiu, Pengyuan; Liu, Rui

2017-01-01

ABSTRACT Bacterial motility is a crucial factor during the invasion and colonization processes of pathogens, which makes it an attractive therapeutic drug target. Here, we isolated a marine bacterium (Vibrio alginolyticus strain 178) from a seamount in the tropical West Pacific that exhibits vigorous motility on agar plates and severe pathogenicity to zebrafish. We found that V. alginolyticus 178 motility was significantly suppressed by another marine bacterium, Bacillus sp. strain 176, isolated from the same niche. We isolated, purified, and characterized two different cyclic lipopeptides (CLPs) from Bacillus sp. 176 using high-performance liquid chromatography, mass spectrometry, and nuclear magnetic resonance spectroscopy. The two related CLPs have a pumilacidin-like structure and were both effective inhibitors of V. alginolyticus 178 motility. The CLPs differ by only one methylene group in their fatty acid chains. In addition to motility suppression, the CLPs also induced cell aggregation in the medium and reduced adherence of V. alginolyticus 178 to glass substrates. Notably, upon CLP treatment, the expression levels of two V. alginolyticus flagellar assembly genes (flgA and flgP) dropped dramatically. Moreover, the CLPs inhibited biofilm formation in several other strains of pathogenic bacteria without inducing cell death. This study indicates that CLPs from Bacillus sp. 176 show promise as antimicrobial lead compounds targeting bacterial motility and biofilm formation with a low potential for eliciting antibiotic resistance. IMPORTANCE Pathogenic bacteria often require motility to establish infections and subsequently spread within host organisms. Thus, motility is an attractive therapeutic target for the development of novel antibiotics. We found that cyclic lipopeptides (CLPs) produced by marine bacterium Bacillus sp. strain 176 dramatically suppress the motility of the pathogenic bacterium Vibrio alginolyticus strain 178, reduce biofilm formation, and promote cellular aggregation without inducing cell death. These findings suggest that CLPs hold great promise as potential drug candidates targeting bacterial motility and biofilm formation with a low overall potential for triggering antibiotic resistance. PMID:28389538

Pumilacidin-Like Lipopeptides Derived from Marine Bacterium Bacillus sp. Strain 176 Suppress the Motility of Vibrio alginolyticus.

PubMed

Xiu, Pengyuan; Liu, Rui; Zhang, Dechao; Sun, Chaomin

2017-06-15

Bacterial motility is a crucial factor during the invasion and colonization processes of pathogens, which makes it an attractive therapeutic drug target. Here, we isolated a marine bacterium ( Vibrio alginolyticus strain 178) from a seamount in the tropical West Pacific that exhibits vigorous motility on agar plates and severe pathogenicity to zebrafish. We found that V. alginolyticus 178 motility was significantly suppressed by another marine bacterium, Bacillus sp. strain 176, isolated from the same niche. We isolated, purified, and characterized two different cyclic lipopeptides (CLPs) from Bacillus sp. 176 using high-performance liquid chromatography, mass spectrometry, and nuclear magnetic resonance spectroscopy. The two related CLPs have a pumilacidin-like structure and were both effective inhibitors of V. alginolyticus 178 motility. The CLPs differ by only one methylene group in their fatty acid chains. In addition to motility suppression, the CLPs also induced cell aggregation in the medium and reduced adherence of V. alginolyticus 178 to glass substrates. Notably, upon CLP treatment, the expression levels of two V. alginolyticus flagellar assembly genes ( flgA and flgP ) dropped dramatically. Moreover, the CLPs inhibited biofilm formation in several other strains of pathogenic bacteria without inducing cell death. This study indicates that CLPs from Bacillus sp. 176 show promise as antimicrobial lead compounds targeting bacterial motility and biofilm formation with a low potential for eliciting antibiotic resistance. IMPORTANCE Pathogenic bacteria often require motility to establish infections and subsequently spread within host organisms. Thus, motility is an attractive therapeutic target for the development of novel antibiotics. We found that cyclic lipopeptides (CLPs) produced by marine bacterium Bacillus sp. strain 176 dramatically suppress the motility of the pathogenic bacterium Vibrio alginolyticus strain 178, reduce biofilm formation, and promote cellular aggregation without inducing cell death. These findings suggest that CLPs hold great promise as potential drug candidates targeting bacterial motility and biofilm formation with a low overall potential for triggering antibiotic resistance. Copyright © 2017 American Society for Microbiology.

Purinergic A2b Receptor Activation by Extracellular Cues Affects Positioning of the Centrosome and Nucleus and Causes Reduced Cell Migration.

PubMed

Ou, Young; Chan, Gordon; Zuo, Jeremy; Rattner, Jerome B; van der Hoorn, Frans A

2016-07-15

The tight, relative positioning of the nucleus and centrosome in mammalian cells is important for the regulation of cell migration. Under pathophysiological conditions, the purinergic A2b receptor can regulate cell motility, but the underlying mechanism remains unknown. Expression of A2b, normally low, is increased in tissues experiencing adverse physiological conditions, including hypoxia and inflammation. ATP is released from such cells. We investigated whether extracellular cues can regulate centrosome-nucleus positioning and cell migration. We discovered that hypoxia as well as extracellular ATP cause a reversible increase in the distance between the centrosome and nucleus and reduced cell motility. We uncovered the underlying pathway: both treatments act through the A2b receptor and specifically activate the Epac1/RapGef3 pathway. We show that cells lacking A2b do not respond in this manner to hypoxia or ATP but transfection of A2b restores this response, that Epac1 is critically involved, and that Rap1B is important for the relative positioning of the centrosome and nucleus. Our results represent, to our knowledge, the first report demonstrating that pathophysiological conditions can impact the distance between the centrosome and nucleus. Furthermore, we identify the A2b receptor as a central player in this process. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Purinergic A2b Receptor Activation by Extracellular Cues Affects Positioning of the Centrosome and Nucleus and Causes Reduced Cell Migration*

PubMed Central

Ou, Young; Chan, Gordon; Zuo, Jeremy; Rattner, Jerome B.; van der Hoorn, Frans A.

2016-01-01

The tight, relative positioning of the nucleus and centrosome in mammalian cells is important for the regulation of cell migration. Under pathophysiological conditions, the purinergic A2b receptor can regulate cell motility, but the underlying mechanism remains unknown. Expression of A2b, normally low, is increased in tissues experiencing adverse physiological conditions, including hypoxia and inflammation. ATP is released from such cells. We investigated whether extracellular cues can regulate centrosome-nucleus positioning and cell migration. We discovered that hypoxia as well as extracellular ATP cause a reversible increase in the distance between the centrosome and nucleus and reduced cell motility. We uncovered the underlying pathway: both treatments act through the A2b receptor and specifically activate the Epac1/RapGef3 pathway. We show that cells lacking A2b do not respond in this manner to hypoxia or ATP but transfection of A2b restores this response, that Epac1 is critically involved, and that Rap1B is important for the relative positioning of the centrosome and nucleus. Our results represent, to our knowledge, the first report demonstrating that pathophysiological conditions can impact the distance between the centrosome and nucleus. Furthermore, we identify the A2b receptor as a central player in this process. PMID:27226580

RickA Expression Is Not Sufficient to Promote Actin-Based Motility of Rickettsia raoultii

PubMed Central

Balraj, Premanand; Karkouri, Khalid El; Vestris, Guy; Espinosa, Leon; Raoult, Didier; Renesto, Patricia

2008-01-01

Background Rickettsia raoultii is a novel Rickettsia species recently isolated from Dermacentor ticks and classified within the spotted fever group (SFG). The inability of R. raoultii to spread within L929 cells suggests that this bacterium is unable to polymerize host cell actin, a property exhibited by all SFG rickettsiae except R. peacocki. This result led us to investigate if RickA, the protein thought to generate actin nucleation, was expressed within this rickettsia species. Methodology/Principal Findings Amplification and sequencing of R. raoultii rickA showed that this gene encoded a putative 565 amino acid protein highly homologous to those found in other rickettsiae. Using immunofluorescence assays, we determined that the motility pattern (i.e. microcolonies or cell-to-cell spreading) of R. raoultii was different depending on the host cell line in which the bacteria replicated. In contrast, under the same experimental conditions, R. conorii shares the same phenotype both in L929 and in Vero cells. Transmission electron microscopy analysis of infected cells showed that non-motile bacteria were free in the cytosol instead of enclosed in a vacuole. Moreover, western-blot analysis demonstrated that the defect of R. raoultii actin-based motility within L929 cells was not related to lower expression of RickA. Conclusion/Significance These results, together with previously published data about R. typhi, strongly suggest that another factor, apart from RickA, may be involved with be responsible for actin-based motility in bacteria from the Rickettsia genus. PMID:18612416

Influence of seminal plasma on the kinematics of boar spermatozoa during freezing.

PubMed

Rodríguez-Martínez, H; Saravia, F; Wallgren, M; Roca, J; Peña, F J

2008-11-01

Sperm motility is, for its relation to cell viability and fertility, a central component of the spermiogram, where consideration of motion patterns allows discrimination of sub-populations among boar spermatozoa. Extension and cryo-preservation imposes changes in these patterns in connection to handling, additives, temperature changes and the removal of boar seminal plasma (BSP) which apparently makes spermatozoa susceptible to oxidative stress, thus affecting survival and motility post-thaw. Detailed kinematic analyses during sperm cooling are sparse, particularly when considering the instrumentation and settings used for analyses, the effect of extenders, and of the BSP the processed spermatozoa are exposed to. Spermatozoa present in the first collectable 10mL of the sperm-rich fraction of the ejaculate (portion 1, P1-BSP), have shown an increased ability to sustain motility during and after cryo-preservation than spermatozoa immersed in the rest of the ejaculate (portion 2, P2). When P2-spermatozoa were cleansed from their BSP and exposed for 60min to pooled P1-BSP, their motility post-thaw increased to similar levels as P1-spermatozoa. This BSP-influence is sire-dependent, presumably related to the protein concentration in the different ejaculate portions, and apparently unrelated to changes in membrane integrity or membrane stability through conventional, controlled cooling.

In Silico Reconstitution of Actin-Based Symmetry Breaking and Motility

PubMed Central

Dayel, Mark J.; Akin, Orkun; Landeryou, Mark; Risca, Viviana; Mogilner, Alex; Mullins, R. Dyche

2009-01-01

Eukaryotic cells assemble viscoelastic networks of crosslinked actin filaments to control their shape, mechanical properties, and motility. One important class of actin network is nucleated by the Arp2/3 complex and drives both membrane protrusion at the leading edge of motile cells and intracellular motility of pathogens such as Listeria monocytogenes. These networks can be reconstituted in vitro from purified components to drive the motility of spherical micron-sized beads. An Elastic Gel model has been successful in explaining how these networks break symmetry, but how they produce directed motile force has been less clear. We have combined numerical simulations with in vitro experiments to reconstitute the behavior of these motile actin networks in silico using an Accumulative Particle-Spring (APS) model that builds on the Elastic Gel model, and demonstrates simple intuitive mechanisms for both symmetry breaking and sustained motility. The APS model explains observed transitions between smooth and pulsatile motion as well as subtle variations in network architecture caused by differences in geometry and conditions. Our findings also explain sideways symmetry breaking and motility of elongated beads, and show that elastic recoil, though important for symmetry breaking and pulsatile motion, is not necessary for smooth directional motility. The APS model demonstrates how a small number of viscoelastic network parameters and construction rules suffice to recapture the complex behavior of motile actin networks. The fact that the model not only mirrors our in vitro observations, but also makes novel predictions that we confirm by experiment, suggests that the model captures much of the essence of actin-based motility in this system. PMID:19771152

Matriptase is required for the active form of hepatocyte growth factor induced Met, focal adhesion kinase and protein kinase B activation on neural stem/progenitor cell motility.

PubMed

Fang, Jung-Da; Lee, Sheau-Ling

2014-07-01

Hepatocyte growth factor (HGF) is a chemoattractant and inducer for neural stem/progenitor (NS/P) cell migration. Although the type II transmembrane serine protease, matriptase (MTP) is an activator of the latent HGF, MTP is indispensable on NS/P cell motility induced by the active form of HGF. This suggests that MTP's action on NS/P cell motility involves mechanisms other than proteolytic activation of HGF. In the present study, we investigate the role of MTP in HGF-stimulated signaling events. Using specific inhibitors of phosphatidylinositol-3-kinase (PI3K), protein kinase B (Akt) or focal adhesion kinase (FAK), we demonstrated that in NS/P cells HGF-activated c-Met induces PI3k-Akt signaling which then leads to FAK activation. This signaling pathway ultimately induces MMP2 expression and NS/P cell motility. Knocking down of MTP in NS/P cells with specific siRNA impaired HGF-stimulation of c-Met, Akt and FAK activation, blocked HGF-induced production of MMP2 and inhibited HGF-stimulated NS/P cell motility. MTP-knockdown NS/P cells cultured in the presence of recombinant protein of MTP protease domain or transfected with the full-length wild-type but not the protease-defected MTP restored HGF-responsive events in NS/P cells. In addition to functioning as HGF activator, our data revealed novel function of MTP on HGF-stimulated c-Met signaling activation. Copyright © 2014. Published by Elsevier B.V.

Effect of semen preparation on casa motility results in cryopreserved bull spermatozoa.

PubMed

Contri, Alberto; Valorz, Claudio; Faustini, Massimo; Wegher, Laura; Carluccio, Augusto

2010-08-01

Computer-assisted sperm analyzers (CASA) have become the standard tool for evaluating sperm motility and kinetic patterns because they provide objective data for thousands of sperm tracks. However, these devices are not ready-to-use and standardization of analytical practices is a fundamental requirement. In this study, we evaluated the effects of some settings, such as frame rate and frames per field, chamber and time of analysis, and samples preparations, including thawing temperature, sperm sample concentration, and media used for dilution, on the kinetic results of bovine frozen-thawed semen using a CASA. In Experiment 1, the frame rate (30-60 frame/s) significantly affected motility parameters, whereas the number of frames per field (30 or 45) did not seem to affect sperm kinetics. In Experiment 2, the thawing protocol affects sperm motility and kinetic parameters. Sperm sample concentration significantly limited the opportunity to perform the analysis and the kinetic results. A concentration of 100 and 50 x 10(6) sperm/mL limited the device's ability to perform the analysis or gave wrong results, whereas 5, 10, 20, and 30 x 10(6) sperm/mL concentrations allowed the analysis to be performed, but with different results (Experiment 3). The medium used for the dilution of the sample, which is fundamental for a correct sperm head detection, affects sperm motility results (Experiment 4). In this study, Makler and Leja chambers were used to perform the semen analysis with CASA devices. The chamber used significantly affected motility results (Experiment 5). The time between chamber loading and analysis affected sperm velocities, regardless of chamber used. Based on results recorded in this study, we propose that the CASA evaluation of motility of bovine frozen-thawed semen using Hamilton-Thorne IVOS 12.3 should be performed using a frame rate of 60 frame/s and 30 frames per field. Semen should be diluted at least at 20 x 10(6) sperm/mL using PBS. Furthermore, it is necessary to consider the type of chamber used and perform the analysis within 1 or 2 min, regardless of the chamber used. Copyright 2010 Elsevier Inc. All rights reserved.

The prevalence of Human Papilloma Virus (HPV) infection in the oligospermic and azoospermic men.

PubMed

Nasseri, Sherko; Monavari, Seyed Hamidreza; Keyvani, Hossein; Nikkhoo, Bahram; Vahabpour Roudsari, Rouhollah; Khazeni, Mohammad

2015-01-01

Human papilloma virus (HPV) infection is one of the most common sexually transmitted diseases that affects men like women and infected cutaneous and mucosal squamous epithelium. The aim of the present study was to determine the prevalence of HPV in the semen of oligospermic, azoospermic and normal patients. From June 2012 to June 2013, a total of 90 individuals were enrolled in this cross sectional comparative study. The participants were classified into three groups (oligospermia, azoosprmia and normal). This classification was based on a new WHO reference values for human semen characteristics published on 2010. After extraction of DNA from specimens L1 gene of HPV was amplified by nested polymerase chain reaction (Nested-PCR) and the PCR products of positive specimens were genotyped using INNO-LiPA HPV Genotyping Extra assay. Among 50 confirmed oligospermic male, 15 were HPV DNA positive (30%). In azoospemic group we had 8 HPV DNA positive (40%) and in normal group just 3 of 20(15%) samples were positive. Statistical assessment was done with SPSS v.15. Chi-square test showed no significant relationship between 3 groups results. Based on independent samples t-test, we found statistical significant relationship for sperm count (p<0.05) and sperm motility (slow) (p<0.05) in oligospermic group positive samples compared with negative. In the present study, 13 HPV genotypes were detected among positive samples. HPV genotypes 16, 45 in the high risk group and 6,11,42 in the low risk group were more frequent than the others. The current study shows that HPV infection can affect on sperm count and motility and decrease count of sperm cell and decrease motility capability of these cells.

Effect of semen extender supplementation with cysteine on postthaw sperm quality, DNA damage, and fertilizing ability in the common carp (Cyprinus carpio).

PubMed

Öğretmen, Fatih; İnanan, Burak Evren; Kutluyer, Filiz; Kayim, Murathan

2015-06-01

Amino acids have an important biological role for prevention of cell damage during cryopreservation. The objective of this study is to determine the effects of cysteine on postthaw sperm motility, duration of sperm motility, DNA damage, and fertility in the common carp (Cyprinus carpio). Sperm collected from 10 individuals was cryopreserved in extenders containing different cysteine concentrations (2.5, 5, 10, and 20 mM). Semen samples diluted at the ratio of 1:9 by the extenders were subjected to cryopreservation. After dilution, the semen was aspirated into 0.25-mL straws; the straws were placed on the tray, frozen in nitrogen vapor, and plunged into liquid nitrogen. DNA damage was evaluated by comet assay after cryopreservation. Our results indicated that an increase in the concentration of cysteine caused a significant increase in the motility rate and duration of sperm in the common carp (C carpio; P < 0.05). Comparing all concentrations of cysteine, the best concentration of cysteine was 20 mM. Higher postthaw motility (76.00 ± 1.00%) and fertilization (97.00 ± 1.73%) rates were obtained with the extender at the concentration of 20 mM. Supplementation of the extender with cysteine was increased the fertilization and hatching rate and decreased DNA damage. Consequently, cysteine affected the motility, fertilization, and DNA damage positively, and extenders could be supplemented with cysteine. Copyright © 2015 Elsevier Inc. All rights reserved.

Changes in p53 expression in mouse fibroblasts can modify motility and extracellular matrix organization.

PubMed

Alexandrova, A; Ivanov, A; Chumakov, P; Kopnin, B; Vasiliev, J

2000-11-23

Effects of p53 expression on cell morphology and motility were studied using the derivatives of p53-null 10(1) mouse fibroblasts with tetracycline-regulated expression of exogenous human p53. Induction of p53 expression was accompanied by significant decrease in extracellular matrix (fibronectin) and reduction of matrix fibrils, diminution of the number and size of focal contacts, decrease of cell areas, establishment of more elongated cell shape and alterations of actin cytoskeleton (actin bundles became thinner, their number and size decreased). Expression of His175 and Gln22/ Ser23 p53 mutants caused no such effects. To study the influence of p53 expression on cell motility we used wound technique and videomicroscopy observation of single living cells. It was found that induction of p53 expression led to increase of lamellar activity of cell edge. However, in spite of enhanced lamellar activity p53-expressing cells migrated to shorter distance and filled the narrow wound in longer time as compared with their p53-null counterparts. Possible mechanisms of the influence of p53 expression on cell morphology and motility are discussed.

Regulation of adipose-tissue-derived stromal cell orientation and motility in 2D- and 3D-cultures by direct-current electrical field.

PubMed

Yang, Gang; Long, Haiyan; Ren, Xiaomei; Ma, Kunlong; Xiao, Zhenghua; Wang, Ying; Guo, Yingqiang

2017-02-01

Cell alignment and motility play a critical role in a variety of cell behaviors, including cytoskeleton reorganization, membrane-protein relocation, nuclear gene expression, and extracellular matrix remodeling. Direct current electric field (EF) in vitro can direct many types of cells to align vertically to EF vector. In this work, we investigated the effects of EF stimulation on rat adipose-tissue-derived stromal cells (ADSCs) in 2D-culture on plastic culture dishes and in 3D-culture on various scaffold materials, including collagen hydrogels, chitosan hydrogels and poly(L-lactic acid)/gelatin electrospinning fibers. Rat ADSCs were exposed to various physiological-strength EFs in a homemade EF-bioreactor. Changes of morphology and movements of cells affected by applied EFs were evaluated by time-lapse microphotography, and cell survival rates and intracellular calcium oscillations were also detected. Results showed that EF facilitated ADSC morphological changes, under 6 V/cm EF strength, and that ADSCs in 2D-culture aligned vertically to EF vector and kept a good cell survival rate. In 3D-culture, cell galvanotaxis responses were subject to the synergistic effect of applied EF and scaffold materials. Fast cell movement and intracellular calcium activities were observed in the cells of 3D-culture. We believe our research will provide some experimental references for the future study in cell galvanotaxis behaviors. © 2017 Japanese Society of Developmental Biologists.

Lysophosphatidic acid signaling via LPA{sub 1} and LPA{sub 3} regulates cellular functions during tumor progression in pancreatic cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fukushima, Kaori; Takahashi, Kaede; Yamasaki, Eri

Lysophosphatidic acid (LPA) signaling via G protein-coupled LPA receptors exhibits a variety of biological effects, such as cell proliferation, motility and differentiation. The aim of this study was to evaluate the roles of LPA{sub 1} and LPA{sub 3} in cellular functions during tumor progression in pancreatic cancer cells. LPA{sub 1} and LPA{sub 3} knockdown cells were generated from PANC-1 cells. The cell motile and invasive activities of PANC-1 cells were inhibited by LPA{sub 1} and LPA{sub 3} knockdown. In gelatin zymography, LPA{sub 1} and LPA{sub 3} knockdown cells indicated the low activation of matrix metalloproteinase-2 (MMP-2) in the presence ofmore » LPA. Next, to assess whether LPA{sub 1} and LPA{sub 3} regulate cellular functions induced by anticancer drug, PANC-1 cells were treated with cisplatin (CDDP) for approximately 6 months. The cell motile and invasive activities of long-term CDDP treated cells were markedly higher than those of PANC-1 cells, correlating with the expression levels of LPAR1 and LPAR3 genes. In soft agar assay, the long-term CDDP treated cells formed markedly large sized colonies. In addition, the cell motile and invasive activities enhanced by CDDP were significantly suppressed by LPA{sub 1} and LPA{sub 3} knockdown as well as colony formation. These results suggest that LPA signaling via LPA{sub 1} and LPA{sub 3} play an important role in the regulation of cellular functions during tumor progression in PANC-1 cells. - Highlights: • The cell motile and invasive activities of PANC-1 cells were stimulated by LPA{sub 1} and LPA{sub 3}. • LPA{sub 1} and LPA{sub 3} enhanced MMP-2 activation in PANC-1 cells. • The expressions of LPAR1 and LPAR3 genes were elevated in PANC-1 cells treated with cisplatin. • The cell motile and invasive activities of PANC-1 cells treated with cisplatin were suppressed by LPA{sub 1} and LPA{sub 3} knockdown. • LPA{sub 1} and LPA{sub 3} are involved in the regulation of cellular functions during tumor progression in PANC-1 cells.« less

Peroxisomes, lipid droplets, and endoplasmic reticulum “hitchhike” on motile early endosomes

PubMed Central

Guimaraes, Sofia C.; Schuster, Martin; Bielska, Ewa; Dagdas, Gulay; Kilaru, Sreedhar; Meadows, Ben R.A.; Schrader, Michael

2015-01-01

Intracellular transport is mediated by molecular motors that bind cargo to be transported along the cytoskeleton. Here, we report, for the first time, that peroxisomes (POs), lipid droplets (LDs), and the endoplasmic reticulum (ER) rely on early endosomes (EEs) for intracellular movement in a fungal model system. We show that POs undergo kinesin-3– and dynein-dependent transport along microtubules. Surprisingly, kinesin-3 does not colocalize with POs. Instead, the motor moves EEs that drag the POs through the cell. PO motility is abolished when EE motility is blocked in various mutants. Most LD and ER motility also depends on EE motility, whereas mitochondria move independently of EEs. Covisualization studies show that EE-mediated ER motility is not required for PO or LD movement, suggesting that the organelles interact with EEs independently. In the absence of EE motility, POs and LDs cluster at the growing tip, whereas ER is partially retracted to subapical regions. Collectively, our results show that moving EEs interact transiently with other organelles, thereby mediating their directed transport and distribution in the cell. PMID:26620910

A Cyanobacterium Capable of Swimming Motility

NASA Astrophysics Data System (ADS)

Waterbury, John B.; Willey, Joanne M.; Franks, Diana G.; Valois, Frederica W.; Watson, Stanley W.

1985-10-01

A novel cyanobacterium capable of swimming motility wass isolated in pure culture from several locations in the Atlantic Ocean. It is a small unicellular form, assignable to the genus Synechococcus, that is capable of swimming through liquids at speeds of 25 micrometers per second. Light microscopy revealed that the motile cells display many features characteristic of bacterial flagellar motility. However, electron microscopy failed to reveal flagella and shearing did not arrest motility, indicating that the cyanobacterium may be propelled by a novel mechanism.

Differential Expression of Virulence Genes and Motility in Ralstonia (Pseudomonas) solanacearum during Exponential Growth.

PubMed

Clough, S J; Flavier, A B; Schell, M A; Denny, T P

1997-03-01

A complex network regulates virulence in Ralstonia solanacearum (formerly Pseudomonas solanacearum); central to this system is PhcA, a LysR-type transcriptional regulator. We report here that two PhcA-regulated virulence factors, endoglucanase (Egl) and acidic exopolysaccharide I (EPS I), and motility are expressed differentially during exponential growth in batch cultures. Tests with strains carrying lacZ fusions in a wild-type genetic background revealed that expression (on a per-cell basis) of phcA was constant but expression of egl and epsB increased 20- to 50-fold during multiplication from 1 x 10(sup7) to 5 x 10(sup8) CFU/ml. Expression of xpsR, an intermediate regulator downstream of PhcA in the regulatory cascade for eps expression, was similar to that of epsB and egl. Motility track photography revealed that all strains were essentially nonmotile at 10(sup6) CFU/ml. As cell density increased, 30 to 50% of wild-type cells were motile between 10(sup7) and 10(sup8) CFU/ml, but this population was again nonmotile at 10(sup9) CFU/ml. In contrast, about 60% of the cells of phcB and phcA mutants remained motile at 10(sup9) CFU/ml. Expression of phcB, which is not positively regulated by PhcA, was the inverse of epsB, egl, and xpsR (i.e., it decreased 20-fold at high cell density). PhcB is essential for production of an extracellular factor, tentatively identified as 3-hydroxypalmitic acid methyl ester (3-OH PAME), that might act as an exponential-phase signal to activate motility or expression of virulence genes. However, growth of the lacZ fusion strains in medium containing excess 3-OH PAME did not result in motility or expression of virulence genes at dramatically lower cell densities, suggesting that 3-OH PAME is not the only factor controlling these traits.

The long non-coding RNA PCSEAT exhibits an oncogenic property in prostate cancer and functions as a competing endogenous RNA that associates with EZH2.

PubMed

Yang, Xiaohui; Wang, Liang; Li, Rong; Zhao, Yuhui; Gu, Yinmin; Liu, Siying; Cheng, Tianyou; Huang, Kuohsiang; Yuan, Yi; Song, Dalong; Gao, Shan

2018-07-12

Prostate cancer (PCa) is the most common malignancy and the leading cause of cancer deaths in males. Recent studies demonstrate that long non-coding RNAs (lncRNAs) are involved in many aspects of PCa. However, their biological roles in PCa remain imperfectly understood. Here,wecharacterized anlncRNA, PCaspecific expression and EZH2-associatedtranscript (PCSEAT, annotated as PRCAT38), which is specifically overexpressedin PCa. We further demonstrated that knockdown of PCSEAT results in the reduction of PCa cell growth and motility, and overexpression of PCSEAT reverses these phenotypes. Furthermore, bioactive PCSEAT is incorporated into exosomes and transmitted to adjacent cells, thus promoting cell proliferation and motility. Mechanistically, we found that PCSEAT promotes cell proliferation, at least in part by affecting miR-143-3p- and miR-24-2-5p-mediated regulation of EZH2, suggesting that PCSEAT and EZH2 competitively 'sponge' miR-143-3p and miR-24-2-5p.Overall, ourresultsrevealthat PCSEAT is specifically overexpressed in PCa patients and a potential oncogene in PCa cells via mediating EZH2 activity, indicating that PCSEAT may be a potential therapeutic target in PCa. Copyright © 2018 Elsevier Inc. All rights reserved.

Effect of Porous Media Particle Size on Bacterial Motility and Chemotaxis

NASA Astrophysics Data System (ADS)

Olson, M. S.; Smith, J. A.; Ford, R. M.; Fernandez, E. J.

2003-12-01

Many soil-inhabiting bacteria that degrade chemical contaminants are both motile and chemotactic. Chemotaxis refers to the ability of bacteria to sense pollutant concentration gradients in water and preferentially swim toward regions of high pollutant concentration, and is thought to be important in guiding subsurface microbial populations toward chemical contaminants. Bacterial motion consists of a series of smooth-swimming runs interrupted by changes in direction. In the presence of a chemical gradient, bacteria bias their frequency of changing direction and demonstrate longer run lengths in the direction of increasing attractant concentration. One concern when studying bacterial chemotaxis in porous media is that in small pores, the porous media may interrupt the extended run lengths of bacteria swimming in the direction of a positive chemical gradient. The purpose of this study is to examine how a decrease in particle size affects the motility and chemotactic response of bacteria traveling through porous media. We employ an innovative technique for noninvasive visualization of changes in bacterial density distributions in a packed column as a function of time. Paramagnetic magnetite particles are attached to the surface of Pseudomonas putida F1 cells using an antibody. Bacterial distributions within a column of glass-coated polystyrene beads are imaged using magnetic resonance imaging (MRI), with a spatial resolution of 300 μ m. Experiments are conducted with both 250-300 μ m beads and 90-150 μ m beads. Bacteria labeled with magnetite are introduced into a specially designed chromatography column packed with glass-coated polystyrene beads. Bacterial migration is monitored over time using MRI, with and without the presence of a chemical gradient of trichloroethylene (TCE). Comparisons of the motility and chemotactic transport coefficients for Pseudomonas putida F1 cells traveling through different-sized samples of porous media in the presence of TCE will be presented and discussed. Results suggest a decrease in both motility and chemotaxis in the smaller-sized porous media.

Loss of Dishevelleds disrupts planar polarity in ependymal motile cilia and results in hydrocephalus

PubMed Central

Ohata, Shinya; Nakatani, Jin; Herranz-Pérez, Vicente; Cheng, JrGang; Belinson, Haim; Inubushi, Toshiro; Snider, William D.; García-Verdugo, Jose Manuel; Wynshaw-Boris, Anthony; Álvarez-Buylla, Arturo

2014-01-01

SUMMARY Defects in ependymal (E) cells, which line the ventricle and generate cerebrospinal fluid flow through ciliary beating, can cause hydrocephalus. Dishevelled genes (Dvls) are essential for Wnt signaling and Dvl2 has been shown to localize to the rootlet of motile cilia. Using the hGFAP-Cre;Dvl1−/−;2flox/flox;3+/− mouse, we show that compound genetic ablation of Dvls causes hydrocephalus. In hGFAP-Cre;Dvl1−/−;2flox/flox;3+/− mutants, E cells differentiated normally, but the intracellular and intercellular rotational alignments of ependymal motile cilia were disrupted. As a consequence, the fluid flow generated by the hGFAP-Cre;Dvl1−/−;2flox/flox;3+/− E cells was significantly slower than that observed in control mice. Dvls were also required for the proper positioning of motile cilia on the apical surface. Tamoxifen-induced conditional removal of Dvls in adult mice also resulted in defects in intracellular rotational alignment and positioning of ependymal motile cilia. These results suggest that Dvls are continuously required for E cell planar polarity and may prevent hydrocephalus. PMID:25043421

Genetic analysis of vertebrate sensory hair cell mechanosensation: the zebrafish circler mutants.

PubMed

Nicolson, T; Rüsch, A; Friedrich, R W; Granato, M; Ruppersberg, J P; Nüsslein-Volhard, C

1998-02-01

The molecular basis of sensory hair cell mechanotransduction is largely unknown. In order to identify genes that are essential for mechanosensory hair cell function, we characterized a group of recently isolated zebrafish motility mutants. These mutants are defective in balance and swim in circles but have no obvious morphological defects. We examined the mutants using calcium imaging of acoustic-vibrational and tactile escape responses, high resolution microscopy of sensory neuroepithelia in live larvae, and recordings of extracellular hair cell potentials (microphonics). Based on the analyses, we have identified several classes of genes. Mutations in sputnik and mariner affect hair bundle integrity. Mutant astronaut and cosmonaut hair cells have relatively normal microphonics and thus appear to affect events downstream of mechanotransduction. Mutant orbiter, mercury, and gemini larvae have normal hair cell morphology and yet do not respond to acoustic-vibrational stimuli. The microphonics of lateral line hair cells of orbiter, mercury, and gemini larvae are absent or strongly reduced. Therefore, these genes may encode components of the transduction apparatus.

Motility of vestibular hair cells in the chick.

PubMed

Ogata, Y; Sekitani, T

1993-01-01

Recent studies of the outer hair cells in cochlea have demonstrated active motilities. However, very little study has been done on the vestibular hair cells (VHCs). The present study shows the motile response of the VHCs induced by application of Ca2+/ATP promoting contraction. Reversible cell shape changes could be shown in 10 of 16 isolated type I hair cells and 9 of 15 isolated type II hair cells by applying the contraction solution. Furthermore, the sensory hair bundles in the utricular epithelium pivoted around the base and stood perpendicularly to the apical borderline of the epithelium in response to the application of the same solution. It is suggested that the contraction of the isolated VHCs may be transferred to tension which causes the sensory hair bundles to restrict their motion in normal tissue, instead of changing the cell shape.

Invadopodia-associated proteins blockade as a novel mechanism for 6-shogaol and pterostilbene to reduce breast cancer cell motility and invasion.

PubMed

Hong, Bo-Han; Wu, Chi-Hao; Yeh, Chi-Tai; Yen, Gow-Chin

2013-05-01

Invadopodia are actin-rich membrane protrusions of tumor cells that are thought to initiate the local migration and invasion during cancer metastasis. The blockade of invadopodia-associated proteins has been reported as a promising approach for prevention of tumor metastasis. The aim of this study was to investigate the modulatory effects of 6-shogaol and pterostilbene on invadopodia in aggressive breast cancer cells. By wound-healing, transwell, and gelatin zymography assays, we found that 6-shogaol and pterostilbene effectively attenuated the motility and invasion of MDA-MB-231 cells, and suppressed the activities of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9). Further investigation into the underlying molecular mechanisms revealed that the levels of key modulators of invadopodium maturation, including c-Src kinase, cortactin, and membrane type 1-matrix metalloproteinase (MT1-MMP) decreased when cells were treated with 6-shogaol or pterostilbene. These data suggest that the repression of these factors might affect the maturation of invadopodia, inhibiting the metastasis of MDA-MB-231 cells. In conclusion, the present study demonstrates for the first time that 6-shogaol and pterostilbene can inhibit invadopodium formation and MMP activity in highly invasive breast cancer cells. We suggest that these compounds may be clinically useful in chemopreventive treatments for metastatic breast cancer. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cryopreservation and ensuing in vitro fertilization ability of boar spermatozoa from epididymides stored at 4 degrees C.

PubMed

Kikuchi, K; Nagai, T; Kashiwazaki, N; Ikeda, H; Noguchi, J; Shimada, A; Soloy, E; Kaneko, H

1998-09-01

The influence of prolonged storage of boar epididymides on post-thaw sperm motility, and in vitro fertilization was evaluated. Twenty pairs of epididymides were obtained from Large White boars, and spermatozoa from one of each of the pairs were immediately collected and frozen (control group). The remaining epididymides were cooled to 4 degrees C and stored for 1, 2 or 3 d, after which spermatozoa were collected and frozen (experimental groups Day 1, 2 and 3, respectively). Sperm motility was maintained throughout the dilution procedure and then dropped (P < 0.01) after freezing and thawing. During storage the motility of nonfrozen spermatozoa decreased significantly (P < 0.01), reaching a value equal to that of frozen-thawed spermatozoa on Day 3. In vitro fertilization experiments revealed significantly (P < 0.05) lower penetration rates using Day 1, 2 and 3 stored spermatozoa (12, 13 and 2%, respectively) than that of the control group (40%). Oocyte penetration ability seemed to be reflected by acrosome integrity. However, the motility of spermatozoa with the ability to penetrate oocytes in Day 1 and Day 2 groups did not differ from that of the controls. The motility of spermatozoa lacking penetration ability, on the other hand, gradually decreased as the storage period was prolonged. This suggests that the sperm motility and penetration ability are affected by different mechanisms during the cold storage of epididymides. Finally, control and experimental groups exhibited high incidences of monospermic penetration (64 to 90%) and of male pronuclear formation (67 to 71%). These data suggest that cryopreservation of spermatozoa from boar epididymides stored at 4 degrees C for 1 to 2 d can be used for conserving male germ cells when epididymal spermatozoa can not be collected immediately and cryopreserved.

Coordinated Cyclic-Di-GMP Repression of Salmonella Motility through YcgR and Cellulose

PubMed Central

Zorraquino, Violeta; García, Begoña; Latasa, Cristina; Echeverz, Maite; Toledo-Arana, Alejandro; Valle, Jaione

2013-01-01

Cyclic di-GMP (c-di-GMP) is a secondary messenger that controls a variety of cellular processes, including the switch between a biofilm and a planktonic bacterial lifestyle. This nucleotide binds to cellular effectors in order to exert its regulatory functions. In Salmonella, two proteins, BcsA and YcgR, both of them containing a c-di-GMP binding PilZ domain, are the only known c-di-GMP receptors. BcsA, upon c-di-GMP binding, synthesizes cellulose, the main exopolysaccharide of the biofilm matrix. YcgR is dedicated to c-di-GMP-dependent inhibition of motility through its interaction with flagellar motor proteins. However, previous evidences indicate that in the absence of YcgR, there is still an additional element that mediates motility impairment under high c-di-GMP levels. Here we have uncovered that cellulose per se is the factor that further promotes inhibition of bacterial motility once high c-di-GMP contents drive the activation of a sessile lifestyle. Inactivation of different genes of the bcsABZC operon, mutation of the conserved residues in the RxxxR motif of the BcsA PilZ domain, or degradation of the cellulose produced by BcsA rescued the motility defect of ΔycgR strains in which high c-di-GMP levels were reached through the overexpression of diguanylate cyclases. High c-di-GMP levels provoked cellulose accumulation around cells that impeded flagellar rotation, probably by means of steric hindrance, without affecting flagellum gene expression, exportation, or assembly. Our results highlight the relevance of cellulose in Salmonella lifestyle switching as an architectural element that is both essential for biofilm development and required, in collaboration with YcgR, for complete motility inhibition. PMID:23161026

Differential KrasV12 protein levels control a switch regulating lung cancer cell morphology and motility

PubMed Central

Schäfer, C.; Mohan, A.; Burford, W.; Driscoll, M. K.; Ludlow, A. T.; Wright, W. E.; Shay, J. W.; Danuser, G.

2016-01-01

Introduction Oncogenic Kras mutations are important drivers of lung cancer development and metastasis. They are known to activate numerous cellular signaling pathways implicated in enhanced proliferation, survival, tumorigenicity and motility during malignant progression. Objectives Most previous studies of Kras in cancer have focused on the comparison of cell states in the absence or presence of oncogenic Kras mutations. Here we show that differential expression of the constitutively active mutation KrasV12 has profound effects on cell morphology and motility that drive metastatic processes. Methods The study relies on lung cancer cell transformation models, patient-derived lung cancer cell lines, and human lung tumor sections combined with molecular biology techniques, live-cell imaging and staining methods. Results Our analysis shows two cell functional states driven by KrasV12 protein levels: a non-motile state associated with high KrasV12 levels and tumorigenicity, and a motile state associated with low KrasV12 levels and cell dissemination. Conversion between the states is conferred by differential activation of a mechano-sensitive double-negative feedback between KrasV12/ERK/Myosin II and matrix-adhesion signaling. KrasV12 expression levels change upon cues such as hypoxia and integrin-mediated cell-matrix adhesion, rendering KrasV12 levels an integrator of micro-environmental signals that translate into cellular function. By live cell imaging of tumor models we observe shedding of mixed high and low KrasV12 expressers forming multi-functional collectives with potentially optimal metastatic properties composed of a highly mobile and a highly tumorigenic unit. Discussion Together these data highlight previously unappreciated roles for the quantitative effects of expression level variation of oncogenic signaling molecules in conferring fundamental alterations in cell function regulation required for cancer progression. PMID:29057096

Cell motility assays.

PubMed

Hague, Angela; Jones, Gareth E

2008-10-01

This report summarises practical aspects to measuring cell motility in culture. The methods described here were discussed at a 1-day European Tissue Culture Society (ETCS-UK) workshop organised by John Masters and Gareth E Jones that was held at University College London on 19th April 2007.

StarD13 is a tumor suppressor in breast cancer that regulates cell motility and invasion

PubMed Central

HANNA, SAMER; KHALIL, BASSEM; NASRALLAH, ANITA; SAYKALI, BECHARA A.; SOBH, RANIA; NASSER, SELIM; EL-SIBAI, MIRVAT

2014-01-01

Breast cancer is one of the most commonly diagnosed cancers in women around the world. In general, the more aggressive the tumor, the more rapidly it grows and the more likely it metastasizes. Members of the Rho subfamily of small GTP-binding proteins (GTPases) play a central role in breast cancer cell motility and metastasis. The switch between active GTP-bound and inactive GDP-bound state is regulated by guanine nucleotide exchange factors (GEFs), GTPase-activating proteins (GAPs) and guanine-nucleotide dissociation inhibitors (GDIs). We studied the role of StarD13, a recently identified Rho-GAP that specifically inhibits the function of RhoA and Cdc42. We aimed to investigate its role in breast cancer proliferation and metastasis. The levels of expression of this Rho-GAP in tumor tissues of different grades were assayed using immunohistochemistry. We observed that, while the level of StarD13 expression decreases in cancer tissues compared to normal tissues, it increases as the grade of the tumor increased. This was consistent with the fact that although StarD13 was indeed a tumor suppressor in our breast cancer cells, as seen by its effect on cell proliferation, it was needed for cancer cell motility. In fact, StarD13 knockdown resulted in an inhibition of cell motility and cells were not able to detach their tail and move forward. Our study describes, for the first time, a tumor suppressor that plays a positive role in cancer motility. PMID:24627003

Screen for Slit/Robo signaling in trunk neural cells reveals new players.

PubMed

Martinez, Darwin; Zuhdi, Nora; Reyes, Michelle; Ortega, Blanca; Giovannone, Dion; Lee, Vivian M; de Bellard, Maria Elena

2018-06-01

Slits ligands and their Robo receptors are involved in quite disparate cell signaling pathways that include axon guidance, cell proliferation, cell motility and angiogenesis. Neural crest cells emerge by delamination from neural cells in the dorsal neural tube, and give rise to various components of the peripheral nervous system in vertebrates. It is well established that these cells change from a non-migratory to a highly migratory state allowing them to reach distant regions before they differentiate. However, but the mechanism controlling this delamination and subsequent migration are still not fully understood. The repulsive Slit ligand family members, have been classified also as true tumor suppressor molecules. The present study explored in further detail what possible Slit/Robo signals are at play in the trunk neural cells and neural crest cells by carrying out a microarray after Slit2 gain of function in trunk neural tubes. We found that in addition to molecules known to be downstream of Slit/Robo signaling, there were a large set of molecules known to be important in maintaining cells in non-motile, epithelia phenotype. Furthermore, we found new molecules previously not associated with Slit/Robo signaling: cell proliferation markers, Ankyrins and RAB intracellular transporters. Our findings suggest that neural crest cells use and array of different Slit/Robo pathways during their transformation from non-motile to highly motile cells. Copyright © 2018. Published by Elsevier B.V.

Effect of tributyltin on adenylate content and enzyme activities of teleost sperm: a biochemical approach to study the mechanisms of toxicant reduced spermatozoa motility.

PubMed

Rurangwa, E; Biegniewska, A; Slominska, E; Skorkowski, E F; Ollevier, F

2002-03-01

The effects of tributyltin (TBT) on the energy metabolism and motility of fish spermatozoa were investigated in vitro in African catfish and common carp. A significant (P<0.05) decrease of the duration and the intensity of motility was observed in catfish spermatozoa exposed to 0.27 microg/l TBT for 24 h. Exposure of catfish spermatozoa to 2.7-27 microg/l TBT caused an instant decrease in ATP content. In the presence of 27 microg/l TBT approximately 55% of the initial ATP concentration in catfish semen was lost after 60 min incubation while AMP concentrations increased and the total adenine nucleotide (TAN) pool remained unchanged. The reduction in sperm ATP levels could not be attributed to cell death since viability decreased only slightly over the period of exposure. In carp by contrast, none of the adenylates concentrations studied (ATP, ADP and AMP) were affected by TBT exposure at any experimental condition. However, carp sperm motility was significantly reduced by exposure to 2.7 microg/l TBT. Among the enzymes investigated only lactate dehydrogenase (LDH) in catfish sperm was significantly (P<0.01) affected by 27 microg/l TBT treatment with a reduction in activity of approximately 75%. Compared with carp sperm before TBT exposure, that of catfish had lower adenylate contents and overall lower enzymatic activities; this explains its slower sperm velocity and shorter duration of movement as measured by computer assisted sperm analysis (CASA). The present in vitro study shows that catfish spermatozoa are more sensitive to TBT exposure (and probably to other toxicants) than those of carp.

Influence of salinomycin treatment on division and movement of individual cancer cells cultured in normoxia or hypoxia evaluated with time-lapse digital holographic microscopy

PubMed Central

Kamlund, Sofia; Strand, Daniel; Janicke, Birgit; Alm, Kersti; Oredsson, Stina

2017-01-01

ABSTRACT Most studies on new cancer drugs are based on population-derived data, where the absence of response of a small population may pass unnoticed. Thus, individual longitudinal tracking of cells is important for the future development of efficient cancer treatments. We have used digital holographic microscopy to track individual JIMT-1 human breast cancer cells and L929 mouse fibroblast cultivated in normoxia or hypoxia. In addition, JIMT-1 cells were treated with salinomycin, a cancer stem cell targeting compound. Three-day time-lapse movies were captured and individual cells were analysed with respect to cell division (cell cycle length) and cell movement. Comparing population-doubling time derived from population-based growth curves and individual cell cycle time data from time-lapse movies show that the former hide a sub-population of dividing cells. Salinomycin treatment increased the motility of cells, however, this motility did not result in an increased distant migration i.e. the cells increased their local movement. MCF-7 breast cancer cells showed similar motility behaviour as salinomycin-treated JIMT-1 cells. We suggest that combining features, such as motility and migration, can be used to distinguish cancer cells with mesenchymal (JIMT-1) and epithelial (MCF-7) features. The data clearly emphasize the importance of longitudinal cell tracking to understand the biology of individual cells under different conditions. PMID:28933990

Sperm Mitochondrial Integrity Is Not Required for Hyperactivated Motility, Zona Binding, or Acrosome Reaction in the Rhesus Macaque1

PubMed Central

Hung, Pei-hsuan; Miller, Marion G.; Meyers, Stuart A.; VandeVoort, Catherine A.

2008-01-01

Whether the main energy source for sperm motility is from oxidative phosphorylation or glycolysis has been long-debated in the field of reproductive biology. Using the rhesus monkey as a model, we examined the role of glycolysis and oxidative phosphorylation in sperm function by using alpha-chlorohydrin (ACH), a glycolysis inhibitor, and pentachlorophenol (PCP), an oxidative phosphorylation uncoupler. Sperm treated with ACH showed no change in percentage of motile sperm, although sperm motion was impaired. The ACH-treated sperm did not display either hyperactivity- or hyperactivation-associated changes in protein tyrosine phosphorylation. When treated with PCP, sperm motion parameters were affected by the highest level of PCP (200 μM); however, PCP did not cause motility impairments even after chemical activation. Sperm treated with PCP were able to display hyperactivity and tyrosine phosphorylation after chemical activation. In contrast with motility measurements, treatment with either the glycolytic inhibitor or the oxidative phosphorylation inhibitor did not affect sperm-zona binding and zona-induced acrosome reaction. The results suggest glycolysis is essential to support sperm motility, hyperactivity, and protein tyrosine phosphorylation, while energy from oxidative phosphorylation is not necessary for hyperactivated sperm motility, tyrosine phosphorylation, sperm-zona binding, and acrosome reaction in the rhesus macaque. PMID:18480469

Choline dehydrogenase polymorphism rs12676 is a functional variation and is associated with changes in human sperm cell function.

PubMed

Johnson, Amy R; Lao, Sai; Wang, Tongwen; Galanko, Joseph A; Zeisel, Steven H

2012-01-01

Approximately 15% of couples are affected by infertility and up to half of these cases arise from male factor infertility. Unidentified genetic aberrations such as chromosomal deletions, translocations and single nucleotide polymorphisms (SNPs) may be the underlying cause of many cases of idiopathic male infertility. Deletion of the choline dehydrogenase (Chdh) gene in mice results in decreased male fertility due to diminished sperm motility; sperm from Chdh(-/-) males have decreased ATP concentrations likely stemming from abnormal sperm mitochondrial morphology and function in these cells. Several SNPs have been identified in the human CHDH gene that may result in altered CHDH enzymatic activity. rs12676 (G233T), a non-synonymous SNP located in the CHDH coding region, is associated with increased susceptibility to dietary choline deficiency and risk of breast cancer. We now report evidence that this SNP is also associated with altered sperm motility patterns and dysmorphic mitochondrial structure in sperm. Sperm produced by men who are GT or TT for rs12676 have 40% and 73% lower ATP concentrations, respectively, in their sperm. rs12676 is associated with decreased CHDH protein in sperm and hepatocytes. A second SNP located in the coding region of IL17BR, rs1025689, is linked to altered sperm motility characteristics and changes in choline metabolite concentrations in sperm.

Choline Dehydrogenase Polymorphism rs12676 Is a Functional Variation and Is Associated with Changes in Human Sperm Cell Function

PubMed Central

Johnson, Amy R.; Lao, Sai; Wang, Tongwen; Galanko, Joseph A.; Zeisel, Steven H.

2012-01-01

Approximately 15% of couples are affected by infertility and up to half of these cases arise from male factor infertility. Unidentified genetic aberrations such as chromosomal deletions, translocations and single nucleotide polymorphisms (SNPs) may be the underlying cause of many cases of idiopathic male infertility. Deletion of the choline dehydrogenase (Chdh) gene in mice results in decreased male fertility due to diminished sperm motility; sperm from Chdh−/− males have decreased ATP concentrations likely stemming from abnormal sperm mitochondrial morphology and function in these cells. Several SNPs have been identified in the human CHDH gene that may result in altered CHDH enzymatic activity. rs12676 (G233T), a non-synonymous SNP located in the CHDH coding region, is associated with increased susceptibility to dietary choline deficiency and risk of breast cancer. We now report evidence that this SNP is also associated with altered sperm motility patterns and dysmorphic mitochondrial structure in sperm. Sperm produced by men who are GT or TT for rs12676 have 40% and 73% lower ATP concentrations, respectively, in their sperm. rs12676 is associated with decreased CHDH protein in sperm and hepatocytes. A second SNP located in the coding region of IL17BR, rs1025689, is linked to altered sperm motility characteristics and changes in choline metabolite concentrations in sperm. PMID:22558321

Motility problems in the intellectually challenged child, adolescent, and young adult.

PubMed

Martinelli, Massimo; Staiano, Annamaria

2011-12-01

Gastrointestinal (GI) motility problems represent an important cause of morbidity and sometimes mortality in patients affected by developmental disorders. This article describes motility disorders in Down syndrome, cerebral palsy, familial dysautonomia, and Williams syndrome. These problems do not often receive appropriate attention, either because priority is given to other medical aspects of the disorder, or because of the inability of affected children to communicate their symptoms. A better approach to the diagnosis and treatment of GI disorders is required to improve quality of life and minimize morbidity and mortality among patients with developmental disorders.

Transposon Insertions of magellan-4 That Impair Social Gliding Motility in Myxococcus xanthus

PubMed Central

Youderian, Philip; Hartzell, Patricia L.

2006-01-01

Myxococcus xanthus has two different mechanisms of motility, adventurous (A) motility, which permits individual cells to glide over solid surfaces, and social (S) motility, which permits groups of cells to glide. To identify the genes involved in S-gliding motility, we mutagenized a ΔaglU (A−) strain with the defective transposon, magellan-4, and screened for S− mutants that form nonmotile colonies. Sequence analysis of the sites of the magellan-4 insertions in these mutants and the alignment of these sites with the M. xanthus genome sequence show that two-thirds of these insertions lie within 27 of the 37 nonessential genes known to be required for social motility, including those necessary for the biogenesis of type IV pili, exopolysaccharide, and lipopolysaccharide. The remaining insertions also identify 31 new, nonessential genes predicted to encode both structural and regulatory determinants of S motility. These include three tetratricopeptide repeat proteins, several regulators of transcription that may control the expression of genes involved in pilus extension and retraction, and additional enzymes involved in polysaccharide metabolism. Three insertions that abolish S motility lie within genes predicted to encode glycolytic enzymes, suggesting that the signal for pilus retraction may be a simple product of exopolysaccharide catabolism. PMID:16299386

Different computer-assisted sperm analysis (CASA) systems highly influence sperm motility parameters.

PubMed

Boryshpolets, S; Kowalski, R K; Dietrich, G J; Dzyuba, B; Ciereszko, A

2013-10-15

In this study, we examined different computer-assisted sperm analysis (CASA) systems (CRISMAS, Hobson Sperm Tracker, and Image J CASA) on the exact same video recordings to evaluate the differences in sperm motility parameters related to the specific CASA used. To cover a wide range of sperm motility parameters, we chose 12-second video recordings at 25 and 50 Hz frame rates after sperm motility activation using three taxonomically distinct fish species (sterlet: Acipenser ruthenus L.; common carp: Cyprinus carpio L.; and rainbow trout: Oncorhynchus mykiss Walbaum) that are characterized by essential differences in sperm behavior during motility. Systematically higher values of velocity and beat cross frequency (BCF) were observed in video recordings obtained at 50 Hz frame frequency compared with 25 Hz for all three systems. Motility parameters were affected by the CASA and species used for analyses. Image J and CRISMAS calculated higher curvilinear velocity (VCL) values for rainbow trout and common carp at 25 Hz frequency compared with the Hobson Sperm Tracker, whereas at 50 Hz, a significant difference was observed only for rainbow trout sperm recordings. No significant difference was observed between the CASA systems for sterlet sperm motility at 25 and 50 Hz. Additional analysis of 1-second segments taken at three time points (1, 6, and 12 seconds of the recording) revealed a dramatic decrease in common carp and rainbow trout sperm speed. The motility parameters of sterlet spermatozoa did not change significantly during the 12-second motility period and should be considered as a suitable model for longer motility analyses. Our results indicated that the CASA used can affect motility results even when the same motility recordings are used. These results could be critically altered by the recording quality, time of analysis, and frame rate of camera, and could result in erroneous conclusions. Copyright © 2013 Elsevier Inc. All rights reserved.

Delamination of neural crest cells requires transient and reversible Wnt inhibition mediated by Dact1/2.

PubMed

Rabadán, M Angeles; Herrera, Antonio; Fanlo, Lucia; Usieto, Susana; Carmona-Fontaine, Carlos; Barriga, Elias H; Mayor, Roberto; Pons, Sebastián; Martí, Elisa

2016-06-15

Delamination of neural crest (NC) cells is a bona fide physiological model of epithelial-to-mesenchymal transition (EMT), a process that is influenced by Wnt/β-catenin signalling. Using two in vivo models, we show that Wnt/β-catenin signalling is transiently inhibited at the time of NC delamination. In attempting to define the mechanism underlying this inhibition, we found that the scaffold proteins Dact1 and Dact2, which are expressed in pre-migratory NC cells, are required for NC delamination in Xenopus and chick embryos, whereas they do not affect the motile properties of migratory NC cells. Dact1/2 inhibit Wnt/β-catenin signalling upstream of the transcriptional activity of T cell factor (TCF), which is required for EMT to proceed. Dact1/2 regulate the subcellular distribution of β-catenin, preventing β-catenin from acting as a transcriptional co-activator to TCF, yet without affecting its stability. Together, these data identify a novel yet important regulatory element that inhibits β-catenin signalling, which then affects NC delamination. © 2016. Published by The Company of Biologists Ltd.

[Thyroid-intestinal motility interactions summary].

PubMed

Pustorino, S; Foti, M; Calipari, G; Pustorino, E; Ferraro, R; Guerrisi, O; Germanotta, G

2004-12-01

Thyroid diseases may be related to gastrointestinal motility symptoms. Such symptoms can vary in degree and, sometimes, are the only clue of a thyroid disease or, at least, the first. The mechanism by which the thyroid hormones can influence gastrointestinal motility, even if not still completely elucidated, can be found in a synergism between a direct effect of the thyronins and an indirect effect mediated by cathecolamines on the muscle cell receptors. Neck discomfort and dysphagia are common findings in patients with thyroid diseases. Hyper- and hypothyroidism can impair esophageal motility, modifying pharyngo-esophageal structure and/or muscular function and interacting with the neuro-humoral regulation of the esophageal peristalsis. Oesophageal motility alterations, observed in patients affected by small non-toxic goiter, are less understandable. At the gastro-duodenal level, basic and postprandial electric rhythm alterations have been observed in hyperthyroid patients, often associated with delayed gastric emptying, too. In such patients, the autonomous nervous system dysfunction may even modify the neuro-hormonal mutual regulation (vagal influence decrease) of the gastro-duodenal myoelectric activity. Hypothyroidism may cause a delay of the gastric emptying too, but such pattern may also be related to an associated autoimmune disease or to an independent chronic modification of the gastric mucosa. Diarrhoea and malabsorption are common findings together with hyperthyroidism, whereas constipation is frequently observed in hypothyroidism. The clinically most demanding situation is certainly the secondary chronic intestinal pseudo-obstruction syndrome, which involves the bowel in most cases, but may also show up by means of a mega-small bowel or a mega-duodenum, or even all of the above. In conclusion it may be stated that: 1) thyroid diseases may be related to symptoms due to digestive motility dysfunction. 2) Any segment of the gastrointestinal trait may be involved. 3) The typical clinical manifestations of the thyroid illnesses may be borderline, missing or concealed by other intercurrent illnesses, especially in the elderly patients. 4) Motility-related digestive symptoms may conceal an underlying, easily misdetected, thyroid disease and must be therefore carefully analyzed.

Determining the relative contribution and hierarchy of hha and qseBC in the regulation of flagellar motility of Escherichia coli O157:H7.

PubMed

Sharma, Vijay K; Casey, Thomas A

2014-01-01

In recent studies, we demonstrated that a deletion of hha caused increased secretion of locus of enterocyte encoded adherence proteins and reduced motility of enterohemorrhagic Escherichia coli (EHEC) O157:H7. In addition to the importance of hha in positive regulation of motility, a two-component quorum sensing pathway encoded by the qseBC genes has been shown to activate bacterial motility in response to mammalian stress hormones epinephrine and norepinephrine as well as bacterially produced autoinducer-3. In this study, we compared regulatory contribution and hierarchy of hha, a member of the Hha/YmoA family of nucleoid-associated proteins, to that of qseBC in the expression of EHEC O157:H7 motility. Since norepinephrine affects motility of EHEC O157:H7 through a qseBC-encoded two-component quorum sensing signaling, we also determined whether the hha-mediated regulation of motility is affected by norepinephrine and whether this effect is qseBC dependent. We used single (Δhha or ΔqseC) and double (Δhha ΔqseC) deletion mutants to show that hha exerts a greater positive regulatory effect in comparison to qseBC on the expression of motility by EHEC O157:H7. We also show that Hha is hierarchically superior in transcriptional regulation of motility than QseBC because transcription of qseC was significantly reduced in the hha deletion mutant compared to that in the parental and the hha-complemented mutant strains. These results suggest that hha regulates motility of EHEC O157:H7 directly as well as indirectly by controlling the transcription of qseBC.

Determining the Relative Contribution and Hierarchy of hha and qseBC in the Regulation of Flagellar Motility of Escherichia coli O157:H7

PubMed Central

Sharma, Vijay K.; Casey, Thomas A.

2014-01-01

In recent studies, we demonstrated that a deletion of hha caused increased secretion of locus of enterocyte encoded adherence proteins and reduced motility of enterohemorrhagic Escherichia coli (EHEC) O157:H7. In addition to the importance of hha in positive regulation of motility, a two-component quorum sensing pathway encoded by the qseBC genes has been shown to activate bacterial motility in response to mammalian stress hormones epinephrine and norepinephrine as well as bacterially produced autoinducer-3. In this study, we compared regulatory contribution and hierarchy of hha, a member of the Hha/YmoA family of nucleoid-associated proteins, to that of qseBC in the expression of EHEC O157:H7 motility. Since norepinephrine affects motility of EHEC O157:H7 through a qseBC-encoded two-component quorum sensing signaling, we also determined whether the hha-mediated regulation of motility is affected by norepinephrine and whether this effect is qseBC dependent. We used single (Δhha or ΔqseC) and double (Δhha ΔqseC) deletion mutants to show that hha exerts a greater positive regulatory effect in comparison to qseBC on the expression of motility by EHEC O157:H7. We also show that Hha is hierarchically superior in transcriptional regulation of motility than QseBC because transcription of qseC was significantly reduced in the hha deletion mutant compared to that in the parental and the hha-complemented mutant strains. These results suggest that hha regulates motility of EHEC O157:H7 directly as well as indirectly by controlling the transcription of qseBC. PMID:24465756

Paxillin Mediates Sensing of Physical Cues and Regulates Directional Cell Motility by Controlling Lamellipodia Positioning

PubMed Central

Sero, Julia E.; Thodeti, Charles K.; Mammoto, Akiko; Bakal, Chris; Thomas, Sheila; Ingber, Donald E.

2011-01-01

Physical interactions between cells and the extracellular matrix (ECM) guide directional migration by spatially controlling where cells form focal adhesions (FAs), which in turn regulate the extension of motile processes. Here we show that physical control of directional migration requires the FA scaffold protein paxillin. Using single-cell sized ECM islands to constrain cell shape, we found that fibroblasts cultured on square islands preferentially activated Rac and extended lamellipodia from corner, rather than side regions after 30 min stimulation with PDGF, but that cells lacking paxillin failed to restrict Rac activity to corners and formed small lamellipodia along their entire peripheries. This spatial preference was preceded by non-spatially constrained formation of both dorsal and lateral membrane ruffles from 5–10 min. Expression of paxillin N-terminal (paxN) or C-terminal (paxC) truncation mutants produced opposite, but complementary, effects on lamellipodia formation. Surprisingly, pax−/− and paxN cells also formed more circular dorsal ruffles (CDRs) than pax+ cells, while paxC cells formed fewer CDRs and extended larger lamellipodia even in the absence of PDGF. In a two-dimensional (2D) wound assay, pax−/− cells migrated at similar speeds to controls but lost directional persistence. Directional motility was rescued by expressing full-length paxillin or the N-terminus alone, but paxN cells migrated more slowly. In contrast, pax−/− and paxN cells exhibited increased migration in a three-dimensional (3D) invasion assay, with paxN cells invading Matrigel even in the absence of PDGF. These studies indicate that paxillin integrates physical and chemical motility signals by spatially constraining where cells will form motile processes, and thereby regulates directional migration both in 2D and 3D. These findings also suggest that CDRs may correspond to invasive protrusions that drive cell migration through 3D extracellular matrices. PMID:22194823

Immature germ cells in semen - correlation with total sperm count and sperm motility.

PubMed

Patil, Priya S; Humbarwadi, Rajendra S; Patil, Ashalata D; Gune, Anita R

2013-07-01

Current data regarding infertility suggests that male factor contributes up to 30% of the total cases of infertility. Semen analysis reveals the presence of spermatozoa as well as a number of non-sperm cells, presently being mentioned in routine semen report as "round cells" without further differentiating them into leucocytes or immature germ cells. The aim of this work was to study a simple, cost-effective, and convenient method for differentiating the round cells in semen into immature germ cells and leucocytes and correlating them with total sperm counts and motility. Semen samples from 120 males, who had come for investigation for infertility, were collected, semen parameters recorded, and stained smears studied for different round cells. Statistical analysis of the data was done to correlate total sperm counts and sperm motility with the occurrence of immature germ cells and leucocytes. The average shedding of immature germ cells in different groups with normal and low sperm counts was compared. The clinical significance of "round cells" in semen and their differentiation into leucocytes and immature germ cells are discussed. Round cells in semen can be differentiated into immature germ cells and leucocytes using simple staining methods. The differential counts mentioned in a semen report give valuable and clinically relevant information. In this study, we observed a negative correlation between total count and immature germ cells, as well as sperm motility and shedding of immature germ cells. The latter was statistically significant with a P value 0.000.

Distinct patterns of primary and motile cilia in Rathke's cleft cysts and craniopharyngioma subtypes.

PubMed

Coy, Shannon; Du, Ziming; Sheu, Shu-Hsien; Woo, Terri; Rodriguez, Fausto J; Kieran, Mark W; Santagata, Sandro

2016-12-01

Cilia are highly conserved organelles, which serve critical roles in development and physiology. Motile cilia are expressed in a limited range of tissues, where they principally regulate local extracellular fluid dynamics. In contrast, primary cilia are expressed by many vertebrate cell types during interphase, and are intimately involved in the cell cycle and signal transduction. Notably, primary cilia are essential for vertebrate hedgehog pathway activity. Improved detection of motile cilia may assist in the diagnosis of some pathologic entities such as Rathke's cleft cysts, whereas characterizing primary cilia in neoplastic tissues may implicate cilia-dependent signaling pathways as critical for tumorigenesis. We show that immunohistochemistry for the nuclear transcription factor FOXJ1, a master regulator of motile ciliogenesis, robustly labels the motile ciliated epithelium of Rathke's cleft cysts. FOXJ1 expression discriminates Rathke's cleft cysts from entities in the sellar/suprasellar region with overlapping histologic features such as craniopharyngiomas. Co-immunohistochemistry for FOXJ1 and markers that highlight motile cilia such as acetylated tubulin (TUBA4A) and the small GTPase ARL13B further enhance the ability to identify diagnostic epithelial cells. In addition to highlighting motile cilia, ARL13B immunohistochemistry also robustly highlights primary cilia in formalin-fixed paraffin-embedded sections. Primary cilia are present throughout the neoplastic epithelium of adamantinomatous craniopharyngioma, but are limited to basally oriented cells near the fibrovascular stroma in papillary craniopharyngioma. Consistent with this differing pattern of primary ciliation, adamantinomatous craniopharyngiomas express significantly higher levels of SHH, and downstream targets such as PTCH1 and GLI2, compared with papillary craniopharyngiomas. In conclusion, motile ciliated epithelium can be readily identified using immunohistochemistry for FOXJ1, TUBA4A, and ARL13B, facilitating the diagnosis of Rathke's cleft cysts. Primary cilia can be identified by ARL13B immunohistochemistry in routine pathology specimens. The widespread presence of primary cilia in adamantinomatous craniopharyngioma implicates cilia-dependent hedgehog signaling in the pathogenesis of adamantinomatous craniopharyngioma.

Distinct Patterns of Primary and Motile Cilia in Rathke’s Cleft Cysts and Craniopharyngioma Subtypes

PubMed Central

Coy, Shannon; Du, Ziming; Sheu, Shu-Hsien; Woo, Terri; Rodriguez, Fausto J.; Kieran, Mark W.; Santagata, Sandro

2017-01-01

Cilia are highly conserved organelles which serve critical roles in development and physiology. Motile cilia are expressed in a limited range of tissues, where they principally regulate local extracellular fluid dynamics. In contrast, primary cilia are expressed by many vertebrate cell types during interphase, and are intimately involved in the cell cycle and signal transduction. Notably, primary cilia are essential for vertebrate hedgehog pathway activity. Improved detection of motile cilia may assist in the diagnosis of some pathologic entities such as Rathke’s cleft cysts while characterizing primary cilia in neoplastic tissues may implicate cilia-dependent signaling pathways as critical for tumorigenesis. We show that immunohistochemistry for the nuclear transcription factor FOXJ1, a master regulator of motile ciliogenesis, robustly labels the motile ciliated epithelium of Rathke’s cleft cysts. FOXJ1 expression discriminates Rathke’s cleft cysts from entities in the sellar/suprasellar region with overlapping histologic features such as craniopharyngiomas. Co-immunohistochemistry for FOXJ1 and markers that highlight motile cilia such as acetylated tubulin (TUBA4A) and the small GTPase ARL13B further enhance the ability to identify diagnostic epithelial cells. In addition to highlighting motile cilia, ARL13B immunohistochemistry also robustly highlights primary cilia in formalin-fixed paraffin-embedded sections. Primary cilia are present throughout the neoplastic epithelium of adamantinomatous craniopharyngioma, but are limited to basally oriented cells near the fibrovascular stroma in papillary craniopharyngioma. Consistent with this differing pattern of primary ciliation, adamantinomatous craniopharyngiomas express significantly higher levels of SHH, and downstream targets such as PTCH1 and GLI2, compared to papillary craniopharyngiomas. In conclusion, motile ciliated epithelium can be readily identified using immunohistochemistry for FOXJ1, TUBA4A and ARL13B, facilitating the diagnosis of Rathke’s cleft cyst. Primary cilia can be identified by ARL13B immunohistochemistry in routine pathology specimens. The widespread presence of primary cilia in adamantinomatous craniopharyngioma implicates cilia-dependent hedgehog signaling in the pathogenesis of adamantinomatous craniopharyngioma. PMID:27562488

Dynamic localization of HmpF regulates type IV pilus activity and directional motility in the filamentous cyanobacterium Nostoc punctiforme.

PubMed

Cho, Ye Won; Gonzales, Alfonso; Harwood, Thomas V; Huynh, Jessica; Hwang, Yeji; Park, Jun Sang; Trieu, Anthony Q; Italia, Parth; Pallipuram, Vivek K; Risser, Douglas D

2017-10-01

Many cyanobacteria exhibit surface motility powered by type 4 pili (T4P). In the model filamentous cyanobacterium Nostoc punctiforme, the T4P systems are arrayed in static, bipolar rings in each cell. The chemotaxis-like Hmp system is essential for motility and the coordinated polar accumulation of PilA on cells in motile filaments, while the Ptx system controls positive phototaxis. Using transposon mutagenesis, a gene, designated hmpF, was identified as involved in motility. Synteny among filamentous cyanobacteria and the similar expression patterns for hmpF and hmpD imply that HmpF is part of the Hmp system. Deletion of hmpF produced a phenotype distinct from other hmp genes, but indistinguishable from pilB or pilQ. Both an HmpF-GFPuv fusion protein, and PilA, as assessed by in situ immunofluorescence, displayed coordinated, unipolar localization at the leading pole of each cell. Reversals were modulated by changes in light intensity and preceded by the migration of HmpF-GFPuv to the lagging cell poles. These results are consistent with a model where direct interaction between HmpF and the T4P system activates pilus extension, the Hmp system facilitates coordinated polarity of HmpF to establish motility, and the Ptx system modulates HmpF localization to initiate reversals in response to changes in light intensity. © 2017 John Wiley & Sons Ltd.

Quantification of Dynamic Morphological Drug Responses in 3D Organotypic Cell Cultures by Automated Image Analysis

PubMed Central

Härmä, Ville; Schukov, Hannu-Pekka; Happonen, Antti; Ahonen, Ilmari; Virtanen, Johannes; Siitari, Harri; Åkerfelt, Malin; Lötjönen, Jyrki; Nees, Matthias

2014-01-01

Glandular epithelial cells differentiate into complex multicellular or acinar structures, when embedded in three-dimensional (3D) extracellular matrix. The spectrum of different multicellular morphologies formed in 3D is a sensitive indicator for the differentiation potential of normal, non-transformed cells compared to different stages of malignant progression. In addition, single cells or cell aggregates may actively invade the matrix, utilizing epithelial, mesenchymal or mixed modes of motility. Dynamic phenotypic changes involved in 3D tumor cell invasion are sensitive to specific small-molecule inhibitors that target the actin cytoskeleton. We have used a panel of inhibitors to demonstrate the power of automated image analysis as a phenotypic or morphometric readout in cell-based assays. We introduce a streamlined stand-alone software solution that supports large-scale high-content screens, based on complex and organotypic cultures. AMIDA (Automated Morphometric Image Data Analysis) allows quantitative measurements of large numbers of images and structures, with a multitude of different spheroid shapes, sizes, and textures. AMIDA supports an automated workflow, and can be combined with quality control and statistical tools for data interpretation and visualization. We have used a representative panel of 12 prostate and breast cancer lines that display a broad spectrum of different spheroid morphologies and modes of invasion, challenged by a library of 19 direct or indirect modulators of the actin cytoskeleton which induce systematic changes in spheroid morphology and differentiation versus invasion. These results were independently validated by 2D proliferation, apoptosis and cell motility assays. We identified three drugs that primarily attenuated the invasion and formation of invasive processes in 3D, without affecting proliferation or apoptosis. Two of these compounds block Rac signalling, one affects cellular cAMP/cGMP accumulation. Our approach supports the growing needs for user-friendly, straightforward solutions that facilitate large-scale, cell-based 3D assays in basic research, drug discovery, and target validation. PMID:24810913

PED/PEA-15 Controls Fibroblast Motility and Wound Closure by ERK1/2-Dependent Mechanisms

PubMed Central

Buonomo, Roberta; Giacco, Ferdinando; Vasaturo, Angela; Caserta, Sergio; Guido, Stefano; Pagliara, Valentina; Garbi, Corrado; Mansueto, Gelsomina; Cassese, Angela; Perruolo, Giuseppe; Oriente, Francesco; Miele, Claudia; Beguinot, Francesco; Formisano, Pietro

2012-01-01

Cell migration is dependent on the control of signaling events that play significant roles in creating contractile force and in contributing to wound closure. We evaluated wound closure in fibroblasts from mice overexpressing (TgPED) or lacking ped/pea-15 (KO), a gene overexpressed in patients with type 2 diabetes. Cultured skin fibroblasts isolated from TgPED mice showed a significant reduction in the ability to recolonize wounded area during scratch assay, compared to control fibroblasts. This difference was observed both in the absence and in the presence of mytomicin C, an inhibitor of mitosis. In time-lapse experiments, TgPED fibroblasts displayed about twofold lower velocity and diffusion coefficient, as compared to controls. These changes were accompanied by reduced spreading and decreased formation of stress fibers and focal adhesion plaques. At the molecular level, TgPED fibroblasts displayed decreased RhoA activation and increased abundance of phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2). Inhibition of ERK1/2 activity by PD98059 restored RhoA activation, cytoskeleton organization and cell motility, and almost completely rescued wound closure of TgPED fibroblasts. Interestingly, skin fibroblasts isolated from KO mice displayed an increased wound closure ability. In vivo, healing of dorsal wounds was delayed in TgPED and accelerated in KO mice. Thus, PED/PEA-15 may affect fibroblast motility by a mechanism, at least in part, mediated by ERK1/2. J. Cell. Physiol. 227: 2106–2116, 2012. © 2011 Wiley Periodicals, Inc. PMID:21780113

Termination factor Rho: From the control of pervasive transcription to cell fate determination in Bacillus subtilis

PubMed Central

Nicolas, Pierre; Repoila, Francis; Bardowski, Jacek; Aymerich, Stéphane

2017-01-01

In eukaryotes, RNA species originating from pervasive transcription are regulators of various cellular processes, from the expression of individual genes to the control of cellular development and oncogenesis. In prokaryotes, the function of pervasive transcription and its output on cell physiology is still unknown. Most bacteria possess termination factor Rho, which represses pervasive, mostly antisense, transcription. Here, we investigate the biological significance of Rho-controlled transcription in the Gram-positive model bacterium Bacillus subtilis. Rho inactivation strongly affected gene expression in B. subtilis, as assessed by transcriptome and proteome analysis of a rho–null mutant during exponential growth in rich medium. Subsequent physiological analyses demonstrated that a considerable part of Rho-controlled transcription is connected to balanced regulation of three mutually exclusive differentiation programs: cell motility, biofilm formation, and sporulation. In the absence of Rho, several up-regulated sense and antisense transcripts affect key structural and regulatory elements of these differentiation programs, thereby suppressing motility and biofilm formation and stimulating sporulation. We dissected how Rho is involved in the activity of the cell fate decision-making network, centered on the master regulator Spo0A. We also revealed a novel regulatory mechanism of Spo0A activation through Rho-dependent intragenic transcription termination of the protein kinase kinB gene. Altogether, our findings indicate that distinct Rho-controlled transcripts are functional and constitute a previously unknown built-in module for the control of cell differentiation in B. subtilis. In a broader context, our results highlight the recruitment of the termination factor Rho, for which the conserved biological role is probably to repress pervasive transcription, in highly integrated, bacterium-specific, regulatory networks. PMID:28723971

Genetic Analysis of Collective Motility of Paenibacillus sp. NAIST15-1

PubMed Central

Kobayashi, Kazuo; Kanesaki, Yu

2016-01-01

Bacteria have developed various motility mechanisms to adapt to a variety of solid surfaces. A rhizosphere isolate, Paenibacillus sp. NAIST15-1, exhibited unusual motility behavior. When spotted onto 1.5% agar media, Paenibacillus sp. formed many colonies, each of which moved around actively at a speed of 3.6 μm/sec. As their density increased, each moving colony began to spiral, finally forming a static round colony. Despite its unusual motility behavior, draft genome sequencing revealed that both the composition and organization of flagellar genes in Paenibacillus sp. were very similar to those in Bacillus subtilis. Disruption of flagellar genes and flagellar stator operons resulted in loss of motility. Paenibacillus sp. showed increased transcription of flagellar genes and hyperflagellation on hard agar media. Thus, increased flagella and their rotation drive Paenibacillus sp. motility. We also identified a large extracellular protein, CmoA, which is conserved only in several Paenibacillus and related species. A cmoA mutant could neither form moving colonies nor move on hard agar media; however, motility was restored by exogenous CmoA. CmoA was located around cells and enveloped cell clusters. Comparison of cellular behavior between the wild type and cmoA mutant indicated that extracellular CmoA is involved in drawing water out of agar media and/or smoothing the cell surface interface. This function of CmoA probably enables Paenibacillus sp. to move on hard agar media. PMID:27764113

Fungal lectin of Peltigera canina induces chemotropism of compatible Nostoc cells by constriction-relaxation pulses of cyanobiont cytoskeleton.

PubMed

Díaz, Eva Maria; Vicente-Manzanares, Miguel; Sacristan, Mara; Vicente, Carlos; Legaz, Maria-Estrella

2011-10-01

A glycosylated arginase acting as a fungal lectin from Peltigera canina is able to produce recruitment of cyanobiont Nostoc cells and their adhesion to the hyphal surface. This implies that the cyanobiont would develop organelles to motility towards the chemoattractant. However when visualized by transmission electron microscopy, Nostoc cells recently isolated from P. canina thallus do not reveal any motile, superficial organelles, although their surface was covered by small spindles and serrated layer related to gliding. The use of S-(3,4-dichlorobenzyl)isothiourea, blebbistatin, phalloidin and latrunculin A provide circumstantial evidence that actin microfilaments rather than MreB, the actin-like protein from prokaryota, and, probably, an ATPase which develops contractile function similar to that of myosin II, are involved in cell motility. These experimental facts, the absence of superficial elements (fimbriae, pili or flagellum) related to cell movement, and the appearance of sunken cells during of after movement verified by scanning electron microscopy, support the hypothesis that the motility of lichen cyanobionts could be achieved by contraction-relaxation episodes of the cytoskeleton induced by fungal lectin act as a chemoattractant.

Fungal lectin of Peltigera canina induces chemotropism of compatible Nostoc cells by constriction-relaxation pulses of cyanobiont cytoskeleton

PubMed Central

Díaz, Eva Maria; Vicente-Manzanares, Miguel; Sacristan, Mara; Legaz, Maria-Estrella

2011-01-01

A glycosylated arginase acting as a fungal lectin from Peltigera canina is able to produce recruitment of cyanobiont Nostoc cells and their adhesion to the hyphal surface. This implies that the cyanobiont would develop organelles to motility toward the chemoattractant. However when visualized by transmission electron microscopy, Nostoc cells recently isolated from P. canina thallus do not reveal any motile, superficial organelles, although their surface was covered by small spindles and serrated layer related to gliding. The use of S-(3,4-dichlorobenzyl)isothiourea, blebbistatin, phalloidin and latrunculin A provide circumstantial evidence that actin microfilaments rather than MreB, the actin-like protein from prokaryota, and probably, an ATPase which develops contractile function similar to that of myosin II, are involved in cell motility. These experimental facts, the absence of superficial elements (fimbriae, pili or flagellum) related to cell movement, and the appearance of sunken cells during of after movement verified by scanning electron microscopy, support the hypothesis that the motility of lichen cyanobionts could be achieved by contraction-relaxation episodes of the cytoskeleton induced by fungal lectin act as a chemoattractant. PMID:21897128

Essential role of protein kinase C zeta in transducing a motility signal induced by superoxide and a chemotactic peptide, fMLP.

PubMed

Kuribayashi, Kageaki; Nakamura, Kiminori; Tanaka, Maki; Sato, Tsutomu; Kato, Junji; Sasaki, Katsunori; Takimoto, Rishu; Kogawa, Katsuhisa; Terui, Takeshi; Takayama, Tetsuji; Onuma, Takayuki; Matsunaga, Takuya; Niitsu, Yoshiro

2007-03-26

Under various pathological conditions, including infection, malignancy, and autoimmune diseases, tissues are incessantly exposed to reactive oxygen species produced by infiltrating inflammatory cells. We show augmentation of motility associated with morphological changes of human squamous carcinoma SASH1 cells, human peripheral monocytes (hPMs), and murine macrophage-like cell line J774.1 by superoxide stimulation. We also disclose that motility of hPMs and J774.1 induced by a chemotactic peptide (N-formyl-methionyl-leucyl-phenylalanine [fMLP]) was inhibited by superoxide dismutase or N-acetylcystein, indicating stimulation of motility by superoxide generated by fMLP stimulation. In these cells, protein kinase C (PKC) zeta was activated to phosphorylate RhoGDI-1, which liberated RhoGTPases, leading to their activation. These events were inhibited by dominant-negative PKCzeta in SASH1 cells, myristoylated PKCzeta peptides in hPMs and J774.1, or a specific inhibitor of RhoGTPase in SASH1, hPMs, and J774.1. These results suggest a new approach for manipulation of inflammation as well as tumor cell invasion by targeting this novel signaling pathway.

Essential role of protein kinase C ζ in transducing a motility signal induced by superoxide and a chemotactic peptide, fMLP

PubMed Central

Kuribayashi, Kageaki; Nakamura, Kiminori; Tanaka, Maki; Sato, Tsutomu; Kato, Junji; Sasaki, Katsunori; Takimoto, Rishu; Kogawa, Katsuhisa; Terui, Takeshi; Takayama, Tetsuji; Onuma, Takayuki; Matsunaga, Takuya; Niitsu, Yoshiro

2007-01-01

Under various pathological conditions, including infection, malignancy, and autoimmune diseases, tissues are incessantly exposed to reactive oxygen species produced by infiltrating inflammatory cells. We show augmentation of motility associated with morphological changes of human squamous carcinoma SASH1 cells, human peripheral monocytes (hPMs), and murine macrophage-like cell line J774.1 by superoxide stimulation. We also disclose that motility of hPMs and J774.1 induced by a chemotactic peptide (N-formyl-methionyl-leucyl-phenylalanine [fMLP]) was inhibited by superoxide dismutase or N-acetylcystein, indicating stimulation of motility by superoxide generated by fMLP stimulation. In these cells, protein kinase C (PKC) ζ was activated to phosphorylate RhoGDI-1, which liberated RhoGTPases, leading to their activation. These events were inhibited by dominant-negative PKCζ in SASH1 cells, myristoylated PKCζ peptides in hPMs and J774.1, or a specific inhibitor of RhoGTPase in SASH1, hPMs, and J774.1. These results suggest a new approach for manipulation of inflammation as well as tumor cell invasion by targeting this novel signaling pathway. PMID:17389234

Reduced migration of MLH1 deficient colon cancer cells depends on SPTAN1.

PubMed

Hinrichsen, Inga; Ernst, Benjamin Philipp; Nuber, Franziska; Passmann, Sandra; Schäfer, Dieter; Steinke, Verena; Friedrichs, Nicolaus; Plotz, Guido; Zeuzem, Stefan; Brieger, Angela

2014-01-24

Defects in the DNA mismatch repair (MMR) protein MLH1 are frequently observed in sporadic and hereditary colorectal cancers (CRC). Affected tumors generate much less metastatic potential than the MLH1 proficient forms. Although MLH1 has been shown to be not only involved in postreplicative MMR but also in several MMR independent processes like cytoskeletal organization, the connection between MLH1 and metastasis remains unclear. We recently identified non-erythroid spectrin αII (SPTAN1), a scaffolding protein involved in cell adhesion and motility, to interact with MLH1. In the current study, the interaction of MLH1 and SPTAN1 and its potential consequences for CRC metastasis was evaluated. Nine cancer cell lines as well as fresh and paraffin embedded colon cancer tissue from 12 patients were used in gene expression studies of SPTAN1 and MLH1. Co-expression of SPTAN1 and MLH1 was analyzed by siRNA knock down of MLH1 in HeLa, HEK293, MLH1 positive HCT116, SW480 and LoVo cells. Effects on cellular motility were determined in MLH1 deficient HCT116 and MLH1 deficient HEK293T compared to their MLH1 proficient sister cells, respectively. MLH1 deficiency is clearly associated with SPTAN1 reduction. Moreover, siRNA knock down of MLH1 decreased the mRNA level of SPTAN1 in HeLa, HEK293 as well as in MLH1 positive HCT116 cells, which indicates a co-expression of SPTAN1 by MLH1. In addition, cellular motility of MLH1 deficient HCT116 and MLH1 deficient HEK293T cells was impaired compared to the MLH1 proficient sister clones. Consequently, overexpression of SPTAN1 increased migration of MLH1 deficient cells while knock down of SPTAN1 decreased cellular mobility of MLH1 proficient cells, indicating SPTAN1-dependent migration ability. These data suggest that SPTAN1 levels decreased in concordance with MLH1 reduction and impaired cellular mobility in MLH1 deficient colon cancer cells. Therefore, aggressiveness of MLH1-positive CRC might be related to SPTAN1.

Reduced migration of MLH1 deficient colon cancer cells depends on SPTAN1

PubMed Central

2014-01-01

Introduction Defects in the DNA mismatch repair (MMR) protein MLH1 are frequently observed in sporadic and hereditary colorectal cancers (CRC). Affected tumors generate much less metastatic potential than the MLH1 proficient forms. Although MLH1 has been shown to be not only involved in postreplicative MMR but also in several MMR independent processes like cytoskeletal organization, the connection between MLH1 and metastasis remains unclear. We recently identified non-erythroid spectrin αII (SPTAN1), a scaffolding protein involved in cell adhesion and motility, to interact with MLH1. In the current study, the interaction of MLH1 and SPTAN1 and its potential consequences for CRC metastasis was evaluated. Methods Nine cancer cell lines as well as fresh and paraffin embedded colon cancer tissue from 12 patients were used in gene expression studies of SPTAN1 and MLH1. Co-expression of SPTAN1 and MLH1 was analyzed by siRNA knock down of MLH1 in HeLa, HEK293, MLH1 positive HCT116, SW480 and LoVo cells. Effects on cellular motility were determined in MLH1 deficient HCT116 and MLH1 deficient HEK293T compared to their MLH1 proficient sister cells, respectively. Results MLH1 deficiency is clearly associated with SPTAN1 reduction. Moreover, siRNA knock down of MLH1 decreased the mRNA level of SPTAN1 in HeLa, HEK293 as well as in MLH1 positive HCT116 cells, which indicates a co-expression of SPTAN1 by MLH1. In addition, cellular motility of MLH1 deficient HCT116 and MLH1 deficient HEK293T cells was impaired compared to the MLH1 proficient sister clones. Consequently, overexpression of SPTAN1 increased migration of MLH1 deficient cells while knock down of SPTAN1 decreased cellular mobility of MLH1 proficient cells, indicating SPTAN1-dependent migration ability. Conclusions These data suggest that SPTAN1 levels decreased in concordance with MLH1 reduction and impaired cellular mobility in MLH1 deficient colon cancer cells. Therefore, aggressiveness of MLH1-positive CRC might be related to SPTAN1. PMID:24456667

The Major Outer Membrane Protein MopB Is Required for Twitching Movement and Affects Biofilm Formation and Virulence in Two Xylella fastidiosa strains.

PubMed

Chen, Hongyu; Kandel, Prem P; Cruz, Luisa F; Cobine, Paul A; De La Fuente, Leonardo

2017-11-01

MopB is a major outer membrane protein (OMP) in Xylella fastidiosa, a bacterial plant pathogen that causes losses on many economically important crops. Based on in silico analysis, the uncharacterized MopB protein of X. fastidiosa contains a β-barrel structure with an OmpA-like domain and a predicted calcium-binding motif. Here, MopB function was studied by mutational analysis taking advantage of the natural competence of X. fastidiosa. Mutants of mopB were constructed in two different X. fastidiosa strains, the type strain Temecula and the more virulent WM1-1. Deletion of the mopB gene impaired cell-to-cell aggregation, surface attachment, and biofilm formation in both strains. Interestingly, mopB deletion completely abolished twitching motility. Electron microscopy of the bacterial cell surface revealed that mopB deletion eliminated type IV and type I pili formation, potentially caused by destabilization of the outer membrane. Both mopB mutants showed reduced virulence using tobacco (Nicotiana tabacum) as a host under greenhouse conditions. These results suggest that MopB has pleiotropic functions in biofilm formation and twitching motility and is important for virulence of X. fastidiosa.

The nexin link and B-tubule glutamylation maintain the alignment of outer doublets in the ciliary axoneme.

PubMed

Alford, Lea M; Stoddard, Daniel; Li, Jennifer H; Hunter, Emily L; Tritschler, Douglas; Bower, Raqual; Nicastro, Daniela; Porter, Mary E; Sale, Winfield S

2016-06-01

We developed quantitative assays to test the hypothesis that the N-DRC is required for integrity of the ciliary axoneme. We examined reactivated motility of demembranated drc cells, commonly termed "reactivated cell models." ATP-induced reactivation of wild-type cells resulted in the forward swimming of ∼90% of cell models. ATP-induced reactivation failed in a subset of drc cell models, despite forward motility in live drc cells. Dark-field light microscopic observations of drc cell models revealed various degrees of axonemal splaying. In contrast, >98% of axonemes from wild-type reactivated cell models remained intact. The sup-pf4 and drc3 mutants, unlike other drc mutants, retain most of the N-DRC linker that interconnects outer doublet microtubules. Reactivated sup-pf4 and drc3 cell models displayed nearly wild-type levels of forward motility. Thus, the N-DRC linker is required for axonemal integrity. We also examined reactivated motility and axoneme integrity in mutants defective in tubulin polyglutamylation. ATP-induced reactivation resulted in forward swimming of >75% of tpg cell models. Analysis of double mutants defective in tubulin polyglutamylation and different regions of the N-DRC indicate B-tubule polyglutamylation and the distal lobe of the linker region are both important for axonemal integrity and normal N-DRC function. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Transcriptomic Profiling Suggests That Promysalin Alters the Metabolic Flux, Motility, and Iron Regulation in Pseudomonas putida KT2440.

PubMed

Giglio, Krista M; Keohane, Colleen E; Stodghill, Paul V; Steele, Andrew D; Fetzer, Christian; Sieber, Stephan A; Filiatrault, Melanie J; Wuest, William M

2018-06-05

Promysalin, a secondary metabolite produced by P. putida RW10S1, is a narrow-spectrum antibiotic that targets P. aeruginosa over other Pseudomonas spp. P. putida KT2440, a nonproducing strain, displays increased swarming motility and decreased pyoverdine production in the presence of exogenous promysalin. Herein, proteomic and transcriptomic experiments were used to provide insight about how promysalin elicits responses in PPKT2440 and rationalize its species selectivity. RNA-sequencing results suggest that promysalin affects PPKT2440 by (1) increasing swarming in a flagella-independent manner; (2) causing cells to behave as if they were experiencing an iron-deficient environment, and (3) shifting metabolism away from glucose conversion to pyruvate via the Entner-Doudoroff pathway. These findings highlight nature's ability to develop small molecules with specific targets, resulting in exquisite selectivity.

Three-dimensional matrix fiber alignment modulates cell migration and MT1-MMP utility by spatially and temporally directing protrusions

NASA Astrophysics Data System (ADS)

Fraley, Stephanie I.; Wu, Pei-Hsun; He, Lijuan; Feng, Yunfeng; Krisnamurthy, Ranjini; Longmore, Gregory D.; Wirtz, Denis

2015-10-01

Multiple attributes of the three-dimensional (3D) extracellular matrix (ECM) have been independently implicated as regulators of cell motility, including pore size, crosslink density, structural organization, and stiffness. However, these parameters cannot be independently varied within a complex 3D ECM protein network. We present an integrated, quantitative study of these parameters across a broad range of complex matrix configurations using self-assembling 3D collagen and show how each parameter relates to the others and to cell motility. Increasing collagen density resulted in a decrease and then an increase in both pore size and fiber alignment, which both correlated significantly with cell motility but not bulk matrix stiffness within the range tested. However, using the crosslinking enzyme Transglutaminase II to alter microstructure independently of density revealed that motility is most significantly predicted by fiber alignment. Cellular protrusion rate, protrusion orientation, speed of migration, and invasion distance showed coupled biphasic responses to increasing collagen density not predicted by 2D models or by stiffness, but instead by fiber alignment. The requirement of matrix metalloproteinase (MMP) activity was also observed to depend on microstructure, and a threshold of MMP utility was identified. Our results suggest that fiber topography guides protrusions and thereby MMP activity and motility.

Step-wise loss of bacterial flagellar torsion confers progressive phagocytic evasion.

PubMed

Lovewell, Rustin R; Collins, Ryan M; Acker, Julie L; O'Toole, George A; Wargo, Matthew J; Berwin, Brent

2011-09-01

Phagocytosis of bacteria by innate immune cells is a primary method of bacterial clearance during infection. However, the mechanisms by which the host cell recognizes bacteria and consequentially initiates phagocytosis are largely unclear. Previous studies of the bacterium Pseudomonas aeruginosa have indicated that bacterial flagella and flagellar motility play an important role in colonization of the host and, importantly, that loss of flagellar motility enables phagocytic evasion. Here we use molecular, cellular, and genetic methods to provide the first formal evidence that phagocytic cells recognize bacterial motility rather than flagella and initiate phagocytosis in response to this motility. We demonstrate that deletion of genes coding for the flagellar stator complex, which results in non-swimming bacteria that retain an initial flagellar structure, confers resistance to phagocytic binding and ingestion in several species of the gamma proteobacterial group of Gram-negative bacteria, indicative of a shared strategy for phagocytic evasion. Furthermore, we show for the first time that susceptibility to phagocytosis in swimming bacteria is proportional to mot gene function and, consequently, flagellar rotation since complementary genetically- and biochemically-modulated incremental decreases in flagellar motility result in corresponding and proportional phagocytic evasion. These findings identify that phagocytic cells respond to flagellar movement, which represents a novel mechanism for non-opsonized phagocytic recognition of pathogenic bacteria.

Step-Wise Loss of Bacterial Flagellar Torsion Confers Progressive Phagocytic Evasion

PubMed Central

Lovewell, Rustin R.; Collins, Ryan M.; Acker, Julie L.; O'Toole, George A.; Wargo, Matthew J.; Berwin, Brent

2011-01-01

Phagocytosis of bacteria by innate immune cells is a primary method of bacterial clearance during infection. However, the mechanisms by which the host cell recognizes bacteria and consequentially initiates phagocytosis are largely unclear. Previous studies of the bacterium Pseudomonas aeruginosa have indicated that bacterial flagella and flagellar motility play an important role in colonization of the host and, importantly, that loss of flagellar motility enables phagocytic evasion. Here we use molecular, cellular, and genetic methods to provide the first formal evidence that phagocytic cells recognize bacterial motility rather than flagella and initiate phagocytosis in response to this motility. We demonstrate that deletion of genes coding for the flagellar stator complex, which results in non-swimming bacteria that retain an initial flagellar structure, confers resistance to phagocytic binding and ingestion in several species of the gamma proteobacterial group of Gram-negative bacteria, indicative of a shared strategy for phagocytic evasion. Furthermore, we show for the first time that susceptibility to phagocytosis in swimming bacteria is proportional to mot gene function and, consequently, flagellar rotation since complementary genetically- and biochemically-modulated incremental decreases in flagellar motility result in corresponding and proportional phagocytic evasion. These findings identify that phagocytic cells respond to flagellar movement, which represents a novel mechanism for non-opsonized phagocytic recognition of pathogenic bacteria. PMID:21949654

Actin Filament Polymerization Regulates Gliding Motility by Apicomplexan ParasitesV⃞

PubMed Central

Wetzel, D.M.; Håkansson, S.; Hu, K.; Roos, D.; Sibley, L.D.

2003-01-01

Host cell entry by Toxoplasma gondii depends critically on actin filaments in the parasite, yet paradoxically, its actin is almost exclusively monomeric. In contrast to the absence of stable filaments in conventional samples, rapid-freeze electron microscopy revealed that actin filaments were formed beneath the plasma membrane of gliding parasites. To investigate the role of actin filaments in motility, we treated parasites with the filament-stabilizing drug jasplakinolide (JAS) and monitored the distribution of actin in live and fixed cells using yellow fluorescent protein (YFP)-actin. JAS treatment caused YFP-actin to redistribute to the apical and posterior ends, where filaments formed a spiral pattern subtending the plasma membrane. Although previous studies have suggested that JAS induces rigor, videomicroscopy demonstrated that JAS treatment increased the rate of parasite gliding by approximately threefold, indicating that filaments are rate limiting for motility. However, JAS also frequently reversed the normal direction of motility, disrupting forward migration and cell entry. Consistent with this alteration, subcortical filaments in JAS-treated parasites occurred in tangled plaques as opposed to the straight, roughly parallel orientation observed in control cells. These studies reveal that precisely controlled polymerization of actin filaments imparts the correct timing, duration, and directionality of gliding motility in the Apicomplexa. PMID:12589042

Hyaluronan (HA) interacting proteins RHAMM and hyaluronidase impact prostate cancer cell behavior and invadopodia formation in 3D HA-based hydrogels.

PubMed

Gurski, Lisa A; Xu, Xian; Labrada, Lyana N; Nguyen, Ngoc T; Xiao, Longxi; van Golen, Kenneth L; Jia, Xinqiao; Farach-Carson, Mary C

2012-01-01

To study the individual functions of hyaluronan interacting proteins in prostate cancer (PCa) motility through connective tissues, we developed a novel three-dimensional (3D) hyaluronic acid (HA) hydrogel assay that provides a flexible, quantifiable, and physiologically relevant alternative to current methods. Invasion in this system reflects the prevalence of HA in connective tissues and its role in the promotion of cancer cell motility and tissue invasion, making the system ideal to study invasion through bone marrow or other HA-rich connective tissues. The bio-compatible cross-linking process we used allows for direct encapsulation of cancer cells within the gel where they adopt a distinct, cluster-like morphology. Metastatic PCa cells in these hydrogels develop fingerlike structures, "invadopodia", consistent with their invasive properties. The number of invadopodia, as well as cluster size, shape, and convergence, can provide a quantifiable measure of invasive potential. Among candidate hyaluronan interacting proteins that could be responsible for the behavior we observed, we found that culture in the HA hydrogel triggers invasive PCa cells to differentially express and localize receptor for hyaluronan mediated motility (RHAMM)/CD168 which, in the absence of CD44, appears to contribute to PCa motility and invasion by interacting with the HA hydrogel components. PCa cell invasion through the HA hydrogel also was found to depend on the activity of hyaluronidases. Studies shown here reveal that while hyaluronidase activity is necessary for invadopodia and inter-connecting cluster formation, activity alone is not sufficient for acquisition of invasiveness to occur. We therefore suggest that development of invasive behavior in 3D HA-based systems requires development of additional cellular features, such as activation of motility associated pathways that regulate formation of invadopodia. Thus, we report development of a 3D system amenable to dissection of biological processes associated with cancer cell motility through HA-rich connective tissues.

Effect of various commercial buffers on sperm viability and capacitation.

PubMed

Andrisani, Alessandra; Donà, Gabriella; Ambrosini, Guido; Bonanni, Guglielmo; Bragadin, Marcantonio; Cosmi, Erich; Clari, Giulio; Armanini, Decio; Bordin, Luciana

2014-08-01

A wide variety of sperm preparation protocols are currently available for assisted conception. They include density gradient separation and washing methods. Both aim at isolating and capacitating as much motile sperm as possible for subsequent oocyte fertilization. The aim of this study was to examine the effects of four commercial sperm washing buffers on sperm viability and capacitation. Semen samples from 48 healthy donors (normal values of sperm count, motility, morphology, and volume) were analyzed. After separation (density gradient 40/80%), sperm were incubated in various buffers then analysed for reactive oxygen species (ROS) production, viability, tyrosine phosphorylation (Tyr-P), cholera toxin B subunit (CTB) labeling, and the acrosome reaction (AR). The buffers affected ROS generation in various ways resulting either in rapid cell degeneration (when the amount of ROS was too high for cell survival) or the inability of the cells to maintain correct functioning (when ROS were too few). Only when the correct ROS generation curve was maintained, suitable membrane reorganization, evidenced by CTB labeling was achieved, leading to the highest percentages of both Tyr-P- and acrosome-reacted-cells. Distinguishing each particular pathological state of the sperm sample would be helpful to select the preferred buffer treatment since both ROS production and membrane reorganization can be significantly altered by commercial buffers.

Spermatozoa from the maned wolf (Chrysocyon brachyurus) display typical canid hyper-sensitivity to osmotic and freezing-induced injury, but respond favorably to dimethyl sulfoxide.

PubMed

Johnson, Amy E M; Freeman, Elizabeth W; Wildt, David E; Songsasen, Nucharin

2014-06-01

We assessed the influences of medium osmolality, cryoprotectant and cooling and warming rate on maned wolf (Chrysocyon brachyurus) spermatozoa. Ejaculates were exposed to Ham's F10 medium (isotonic control) or to this medium plus NaCl (350-1000mOsm), sucrose (369 and 479mOsm), 1M glycerol (1086mOsm) or dimethyl sulfoxide (Me2SO, 1151mOsm) for 10 min. Each sample then was diluted back into Ham's medium and assessed for sperm motility and plasma membrane integrity. Although glycerol and Me2SO had no influence (P>0.05), NaCl and sucrose solutions affected sperm motility (P<0.05), but not membrane integrity. Motility of sperm exposed to <600mOsm NaCl or sucrose was less (P<0.05) than fresh ejaculate, but comparable (P>0.05) to the control. As osmolality of the NaCl solution increased, motility decreased to <5%. In a separate study, ejaculates were diluted in Test Yolk Buffer containing 1M glycerol or Me2SO and cooled from 5°C to -120°C at -57.8°C, -124.2°C or -67.0°C/min, frozen in LN2, thawed in a water bath for 30s at 37°C or 10s at 50°C, and then assessed for motility, plasma- and acrosomal membrane integrity. Cryopreservation markedly (P<0.05) reduced sperm motility by 70% compared to fresh samples. Higher (P<0.05) post-thaw motility (20.0±1.9% versus 13.5±2.1%) and membrane integrity (51.2±1.7% versus 41.5±2.2%) were observed in samples cryopreserved in Me2SO than in glycerol. Cooling rates influenced survival of sperm cryopreserved in glycerol with -57.8°C/min being advantageous (P<0.05). The findings demonstrate that although maned wolf spermatozoa are similar to domestic dog sperm in their sensitivity to osmotic-induced motility damage, the plasma membranes tolerate dehydration, and the cells respond favorably to Me2SO as a cryoprotectant. Published by Elsevier Inc.

A Mena invasion isoform potentiates EGF-induced carcinoma cell invasion and metastasis.

PubMed

Philippar, Ulrike; Roussos, Evanthia T; Oser, Matthew; Yamaguchi, Hideki; Kim, Hyung-Do; Giampieri, Silvia; Wang, Yarong; Goswami, Sumanta; Wyckoff, Jeffrey B; Lauffenburger, Douglas A; Sahai, Erik; Condeelis, John S; Gertler, Frank B

2008-12-01

The spread of cancer during metastatic disease requires that tumor cells subvert normal regulatory networks governing cell motility to invade surrounding tissues and migrate toward blood and lymphatic vessels. Enabled (Ena)/vasodilator-stimulated phosphoprotein (VASP) proteins regulate cell motility by controlling the geometry of assembling actin networks. Mena, an Ena/VASP protein, is upregulated in the invasive subpopulation of breast cancer cells. In addition, Mena is alternately spliced to produce an invasion isoform, Mena(INV). Here we show that Mena and Mena(INV) promote carcinoma cell motility and invasiveness in vivo and in vitro, and increase lung metastasis. Mena and Mena(INV) potentiate epidermal growth factor (EGF)-induced membrane protrusion and increase the matrix degradation activity of tumor cells. Interestingly, Mena(INV) is significantly more effective than Mena in driving metastases and sensitizing cells to EGF-dependent invasion and protrusion. Upregulation of Mena(INV) could therefore enable tumor cells to invade in response to otherwise benign EGF stimulus levels.

Genistein inhibits cell invasion and motility by inducing cell differentiation in murine osteosarcoma cell line LM8.

PubMed

Nakamura, Atsushi; Aizawa, Junichi; Sakayama, Kenshi; Kidani, Teruki; Takata, Tomoyo; Norimatsu, Yoshiaki; Miura, Hiromasa; Masuno, Hiroshi

2012-09-26

One of the problems associated with osteosarcoma is the frequent formation of micrometastases in the lung prior to diagnosis because the development of metastatic lesions often causes a fatal outcome. Therefore, the prevention of pulmonary metastases during the early stage of tumor development is critical for the improvement of the prognosis of osteosarcoma patients. In Japan, soy is consumed in a wide variety of forms, such as miso soup and soy sauce. The purpose of this study is to investigate the effect of genistein, an isoflavone found in soy, on the invasive and motile potential of osteosarcoma cells. LM8 cells were treated for 3 days with various concentrations of genistein. The effect of genistein on cell proliferation was determined by DNA measurement in the cultures and 5-bromo-2'-deoxyuridine (BrdU) incorporation study. The assays of cell invasion and motility were performed using the cell culture inserts with either matrigel-coated membranes or uncoated membranes in the invasion chambers. The expression and secretion of MMP-2 were determined by immunohistochemistry and gelatin zymography. The subcellular localization and cellular level of β-catenin were determined by immunofluorescence and Western blot. For examining cell morphology, the ethanol-fixed cells were stained with hematoxylin-eosin (H&E). The expression of osteocalcin mRNA was determined by reverse transcription-polymerase chain reaction (RT-PCR). Genistein dose-dependently inhibits cell proliferation. Genistein-treated cells were less invasive and less motile than untreated cells. The expression and secretion of MMP-2 were lower in the genistein-treated cultures than in the untreated cultures. β-Catenin in untreated cells was located in the cytoplasm and/or nucleus, while in genistein-treated cells it was translocated near to the plasma membrane. The level of β-catenin was higher in genistein-treated cells than in untreated cells. Treatment of LM8 cells with genistein induced morphological changes, markedly decreased the formation of multilayer masses of cells, and markedly increased the expression of osteocalcin mRNA. Genistein decreased invasive and motile potential by inducing cell differentiation in LM8 cells. Genistein may be useful as an anti-metastatic drug for osteosarcoma through its differentiation-inducing effects.

Labelling of living mammalian spermatozoa with the fluorescent thiol alkylating agent, monobromobimane (MB): immobilization upon exposure to ultraviolet light and analysis of acrosomal status

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cummins, J.M.; Fleming, A.D.; Crozet, N.

1986-03-01

Living spermatozoa of seven mammalian species were treated with the thiol-alkylating fluorescent labelling compound, monobromobimane (MBBR). MB-labelling alone had no effect on sperm motility, nor on the time course or ability of golden hamster spermatozoa to undergo the acrosome reaction when capacitated in vitro. Exposure of MB-labelled spermatozoa to ultraviolet (UV) light and excitation of the MB fluorochrome resulted in virtually immediate immobilization of the spermatozoa without affecting acrosomal status. UV exposure of unlabelled spermatozoa for up to 30 sec had no effect upon motility. Immobilization of MB-labelled spermatozoa depended on the midpiece being irradiated, as irradiation of the headmore » alone, or of the more distal parts of the principal piece, had little or no effect upon motility. Labelling with MB followed by immobilization of individually selected spermatozoa was most useful for detailing the course and site of occurrence of the acrosome reaction during penetration of the cumulus oophorus by golden hamster spermatozoa in vitro. In these often hyperactivated spermatozoa, precise determination of the acrosomal status could not often otherwise be made due to the difficulty in visualizing the acrosomal region of a vigorously thrashing, hyperactivated spermatozoon. This technique should prove valuable in a variety of studies on sperm motility, capacitation and fertilization, and could also be extended to other cell systems.« less

Monocarboxylate transporter 1 contributes to growth factor-induced tumor cell migration independent of transporter activity

PubMed Central

Gray, Alana L.; Coleman, David T.; Shi, Runhua; Cardelli, James A.

2016-01-01

Tumor progression to metastatic disease contributes to the vast majority of incurable cancer. Understanding the processes leading to advanced stage cancer is important for the development of future therapeutic strategies. Here, we establish a connection between tumor cell migration, a prerequisite to metastasis, and monocarboxylate transporter 1 (MCT1). MCT1 transporter activity is known to regulate aspects of tumor progression and, as such, is a clinically relevant target for treating cancer. Knockdown of MCT1 expression caused decreased hepatocyte growth factor (HGF)-induced as well as epidermal growth factor (EGF)-induced tumor cell scattering and wound healing. Western blot analysis suggested that MCT1 knockdown (KD) hinders signaling through the HGF receptor (c-Met) but not the EGF receptor. Exogenous, membrane-permeable MCT1 substrates were not able to rescue motility in MCT1 KD cells, nor was pharmacologic inhibition of MCT1 able to recapitulate decreased cell motility as seen with MCT1 KD cells, indicating transporter activity of MCT1 was dispensable for EGF- and HGF-induced motility. These results indicate MCT1 expression, independent of transporter activity, is required for growth factor-induced tumor cell motility. The findings presented herein suggest a novel function for MCT1 in tumor progression independent of its role as a monocarboxylate transporter. PMID:27127175

Quantitative Analysis of Intracellular Motility Based on Optical Flow Model

PubMed Central

Li, Heng

2017-01-01

Analysis of cell mobility is a key issue for abnormality identification and classification in cell biology research. However, since cell deformation induced by various biological processes is random and cell protrusion is irregular, it is difficult to measure cell morphology and motility in microscopic images. To address this dilemma, we propose an improved variation optical flow model for quantitative analysis of intracellular motility, which not only extracts intracellular motion fields effectively but also deals with optical flow computation problem at the border by taking advantages of the formulation based on L1 and L2 norm, respectively. In the energy functional of our proposed optical flow model, the data term is in the form of L2 norm; the smoothness of the data changes with regional features through an adaptive parameter, using L1 norm near the edge of the cell and L2 norm away from the edge. We further extract histograms of oriented optical flow (HOOF) after optical flow field of intracellular motion is computed. Then distances of different HOOFs are calculated as the intracellular motion features to grade the intracellular motion. Experimental results show that the features extracted from HOOFs provide new insights into the relationship between the cell motility and the special pathological conditions. PMID:29065574

How Helicobacter pylori urease may affect external pH and influence growth and motility in the mucus environment: evidence from in-vitro studies.

PubMed

Sidebotham, Ramon L; Worku, Mulugeta L; Karim, Q Najma; Dhir, Nirmal K; Baron, J Hugh

2003-04-01

Survival of Helicobacter pylori is dependent upon urease in the cytoplasm and at the bacterial surface. We have sought to clarify how alkaline ammonium salts, released from urea by this enzyme, might alter mucus pH and so affect growth and motility of the bacterium in the gastric mucus environment. Experiments were conducted in vitro to determine how the growth and motility of H. pylori are affected by changes in external pH, and how the bacterium, by hydrolysing urea, alters the pH of the bicarbonate buffer that occurs at the gastric mucosal surface. These data were fitted into experimental models that describe how pH varies within the mucus layer in the acid-secreting stomach. H. pylori was motile between pH 5 and 8, with optimal motility at pH 5. It grew between pH 6 and 8, with optimal growth at pH 6. The bacterium had urease activity between pH 2.7 and 7.4, as evidenced by pH rises in bicarbonate-buffered solutions of urea. Changes in buffer pH were dependent upon initial pH and urea concentration, with the greatest rate of pH change occurring at pH 3. Modelling experiments utilizing these data indicated that (1) in the absence of urease, H. pylori growth and motility in the mucus layer would be restricted severely by low mucus pH in the acid-secreting stomach, and (2) urease will sometimes inhibit H. pylori growth and motility in the mucus layer by elevating the pH of the mucus environment above pH 8. Urease is essential to the growth and motility of H. pylori in the mucus layer in the acid-secreting stomach, but, paradoxically, sometimes it might suppress colonization by raising the mucus pH above 8. This latter effect may protect the bacteria from the adverse consequences of overpopulation.

Sperm cryopreservation affects postthaw motility, but not embryogenesis or larval growth in the Brazilian fish Brycon insignis (Characiformes).

PubMed

Viveiros, A T M; Isaú, Z A; Caneppele, D; Leal, M C

2012-09-01

Sperm cryopreservation is an important method for preserving genetic information and facilitating artificial reproduction. The objective was to investigate whether the cryopreservation process affects postthaw sperm motility, embryogenesis, and larval growth in the fish Brycon insignis. Sperm was diluted in methyl glycol and Beltsville Thawing solution, frozen in a nitrogen vapor vessel (dry shipper) and stored in liquid nitrogen. Half of the samples were evaluated both subjectively (% of motile sperm and motility quality score-arbitrary grading system from 0 [no movement] to 5 [rapidly swimming sperm]) and in a computer-assisted sperm analyzer (CASA; percentage of motile sperm and velocity). The other half was used for fertilization and the evaluation of embryogenesis (cleavage and gastrula stages), hatching rate, percentage of larvae with normal development and larval growth up to 112 days posthatching (dph). Fresh sperm was analyzed subjectively (percentage of motile sperm and motility quality score) and used as the control. In the subjective analysis, sperm motility significantly decreased from 100% motile sperm and quality score of 5 in fresh sperm to 54% motile sperm and quality score of 3 after thawing. Under computer-assisted sperm analyzer evaluation, postthaw sperm had 67% motile sperm, 122 μm/sec of curvilinear velocity, 87 μm/sec of straight-line velocity and 103 μm/sec of average path velocity. There were no significant differences between progenies (pooled data) for the percentage of viable embryos in cleavage (62%) or gastrula stages (24%) or in the hatching rate (24%), percentage of normal hatched larvae (93%), larval body weight (39.8 g), or standard length (12.7 cm) at 112 days posthatching. Based on these findings, cryopreserved sperm can be used as a tool to restore the population of endangered species, such as B. insignis, as well as for aquaculture purposes, without any concern regarding quality of the offspring. Copyright © 2012 Elsevier Inc. All rights reserved.

Wnt5A Activates the Calpain-Mediated Cleavage of Filamin A

PubMed Central

O’Connell, Michael P.; Fiori, Jennifer L.; Baugher, Katherine M.; Indig, Fred E.; French, Amanda D.; Camilli, Tura C.; Frank, Brittany P.; Earley, Rachel; Hoek, Keith S.; Hasskamp, Joanne H.; Elias, E. George; Taub, Dennis D.; Bernier, Michel; Weeraratna, Ashani T.

2009-01-01

We have previously shown that Wnt5A and ROR2, an orphan tyrosine kinase receptor, interact to mediate melanoma cell motility. In other cell types, this can occur through the interaction of ROR2 with the cytoskeletal protein filamin A. Here, we found that filamin A protein levels correlated with Wnt5A levels in melanoma cells. Small interfering RNA (siRNA) knockdown of WNT5A decreased filamin A expression. Knockdown of filamin A also corresponded to a decrease in melanoma cell motility. In metastatic cells, filamin A expression was predominant in the cytoplasm, which western analysis indicated was due to the cleavage of filamin A in these cells. Treatment of nonmetastatic melanoma cells with recombinant Wnt5A increased filamin A cleavage, and this could be prevented by the knockdown of ROR2 expression. Further, BAPTA-AM chelation of intracellular calcium also inhibited filamin A cleavage, leading to the hypothesis that Wnt5A/ROR2 signaling could cleave filamin A through activation of calcium-activated proteases, such as calpains. Indeed, WNT5A knockdown decreased calpain 1 expression, and by inhibiting calpain 1 either pharmacologically or using siRNA, it decreased cell motility. Our results indicate that Wnt5A activates calpain-1, leading to the cleavage of filamin A, which results in a remodeling of the cytoskeleton and an increase in melanoma cell motility. PMID:19177143

Nr-CAM is a target gene of the beta-catenin/LEF-1 pathway in melanoma and colon cancer and its expression enhances motility and confers tumorigenesis.

PubMed

Conacci-Sorrell, Maralice E; Ben-Yedidia, Tamar; Shtutman, Michael; Feinstein, Elena; Einat, Paz; Ben-Ze'ev, Avri

2002-08-15

beta-catenin and plakoglobin (gamma-catenin) are homologous molecules involved in cell adhesion, linking cadherin receptors to the cytoskeleton. beta-catenin is also a key component of the Wnt pathway by being a coactivator of LEF/TCF transcription factors. To identify novel target genes induced by beta-catenin and/or plakoglobin, DNA microarray analysis was carried out with RNA from cells overexpressing either protein. This analysis revealed that Nr-CAM is the gene most extensively induced by both catenins. Overexpression of either beta-catenin or plakoglobin induced Nr-CAM in a variety of cell types and the LEF/TCF binding sites in the Nr-CAM promoter were required for its activation by catenins. Retroviral transduction of Nr-CAM into NIH3T3 cells stimulated cell growth, enhanced motility, induced transformation, and produced rapidly growing tumors in nude mice. Nr-CAM and LEF-1 expression was elevated in human colon cancer tissue and cell lines and in human malignant melanoma cell lines but not in melanocytes or normal colon tissue. Dominant negative LEF-1 decreased Nr-CAM expression and antibodies to Nr-CAM inhibited the motility of B16 melanoma cells. The results indicate that induction of Nr-CAM transcription by beta-catenin or plakoglobin plays a role in melanoma and colon cancer tumorigenesis, probably by promoting cell growth and motility.

Cell motility and ECM proteolysis regulate tumor growth and tumor relapse by altering the fraction of cancer stem cells and their spatial scattering

NASA Astrophysics Data System (ADS)

Kumar, Sandeep; Kulkarni, Rahul; Sen, Shamik

2016-06-01

Tumors consist of multiple cell sub-populations including cancer stem cells (CSCs), transiently amplifying cells and terminally differentiated cells (TDCs), with the CSC fraction dictating the aggressiveness of the tumor and drug sensitivity. In epithelial cancers, tumor growth is influenced greatly by properties of the extracellular matrix (ECM), with cancer progression associated with an increase in ECM density. However, the extent to which increased ECM confinement induced by an increase in ECM density influences tumor growth and post treatment relapse dynamics remains incompletely understood. In this study, we use a cellular automata-based discrete modeling approach to study the collective influence of ECM density, cell motility and ECM proteolysis on tumor growth, tumor heterogeneity, and tumor relapse after drug treatment. We show that while increased confinement suppresses tumor growth and the spatial scattering of CSCs, this effect can be reversed when cells become more motile and proteolytically active. Our results further suggest that, in addition to the absolute number of CSCs, their spatial positioning also plays an important role in driving tumor growth. In a nutshell, our study suggests that, in confined environments, cell motility and ECM proteolysis are two key factors that regulate tumor growth and tumor relapse dynamics by altering the number and spatial distribution of CSCs.

Quantitative analysis of Plasmodium ookinete motion in three dimensions suggests a critical role for cell shape in the biomechanics of malaria parasite gliding motility.

PubMed

Kan, Andrey; Tan, Yan-Hong; Angrisano, Fiona; Hanssen, Eric; Rogers, Kelly L; Whitehead, Lachlan; Mollard, Vanessa P; Cozijnsen, Anton; Delves, Michael J; Crawford, Simon; Sinden, Robert E; McFadden, Geoffrey I; Leckie, Christopher; Bailey, James; Baum, Jake

2014-05-01

Motility is a fundamental part of cellular life and survival, including for Plasmodium parasites--single-celled protozoan pathogens responsible for human malaria. The motile life cycle forms achieve motility, called gliding, via the activity of an internal actomyosin motor. Although gliding is based on the well-studied system of actin and myosin, its core biomechanics are not completely understood. Currently accepted models suggest it results from a specifically organized cellular motor that produces a rearward directional force. When linked to surface-bound adhesins, this force is passaged to the cell posterior, propelling the parasite forwards. Gliding motility is observed in all three life cycle stages of Plasmodium: sporozoites, merozoites and ookinetes. However, it is only the ookinetes--formed inside the midgut of infected mosquitoes--that display continuous gliding without the necessity of host cell entry. This makes them ideal candidates for invasion-free biomechanical analysis. Here we apply a plate-based imaging approach to study ookinete motion in three-dimensional (3D) space to understand Plasmodium cell motility and how movement facilitates midgut colonization. Using single-cell tracking and numerical analysis of parasite motion in 3D, our analysis demonstrates that ookinetes move with a conserved left-handed helical trajectory. Investigation of cell morphology suggests this trajectory may be based on the ookinete subpellicular cytoskeleton, with complementary whole and subcellular electron microscopy showing that, like their motion paths, ookinetes share a conserved left-handed corkscrew shape and underlying twisted microtubular architecture. Through comparisons of 3D movement between wild-type ookinetes and a cytoskeleton-knockout mutant we demonstrate that perturbation of cell shape changes motion from helical to broadly linear. Therefore, while the precise linkages between cellular architecture and actomyosin motor organization remain unknown, our analysis suggests that the molecular basis of cell shape may, in addition to motor force, be a key adaptive strategy for malaria parasite dissemination and, as such, transmission. © 2014 The Authors. Cellular Microbiology published by John Wiley & Sons Ltd.

Curvature-Guided Motility of Microalgae in Geometric Confinement

NASA Astrophysics Data System (ADS)

Ostapenko, Tanya; Schwarzendahl, Fabian Jan; Böddeker, Thomas J.; Kreis, Christian Titus; Cammann, Jan; Mazza, Marco G.; Bäumchen, Oliver

2018-02-01

Microorganisms, such as bacteria and microalgae, often live in habitats consisting of a liquid phase and a plethora of interfaces. The precise ways in which these motile microbes behave in their confined environment remain unclear. Using experiments and Brownian dynamics simulations, we study the motility of a single Chlamydomonas microalga in an isolated microhabitat with controlled geometric properties. We demonstrate how the geometry of the habitat controls the cell's navigation in confinement. The probability of finding the cell swimming near the boundary increases with the wall curvature, as seen for both circular and elliptical chambers. The theory, utilizing an asymmetric dumbbell model of the cell and steric wall interactions, captures this curvature-guided navigation quantitatively with no free parameters.

Diversification of caldesmon-linked actin cytoskeleton in cell motility

PubMed Central

Mayanagi, Taira

2011-01-01

The actin cytoskeleton plays a key role in regulating cell motility. Caldesmon (CaD) is an actin-linked regulatory protein found in smooth muscle and non-muscle cells that is conserved among a variety of vertebrates. It binds and stabilizes actin filaments, as well as regulating actin-myosin interaction in a calcium (Ca2+)/calmodulin (CaM)- and/or phosphorylation-dependent manner. CaD function is regulated qualitatively by Ca2+/CaM and by its phosphorylation state and quantitatively at the mRNA level, by three different transcriptional regulation of the CALD1 gene. CaD has numerous functions in cell motility, such as migration, invasion and proliferation, exerted via the reorganization of the actin cytoskeleton. Here we will outline recent findings regarding CaD's structural features and functions. PMID:21350330

A Submersible, Off-Axis Holographic Microscope for Detection of Microbial Motility and Morphology in Aqueous and Icy Environments

PubMed Central

Lindensmith, Christian A.; Rider, Stephanie; Bedrossian, Manuel; Wallace, J. Kent; Serabyn, Eugene; Showalter, G. Max; Deming, Jody W.; Nadeau, Jay L.

2016-01-01

Sea ice is an analog environment for several of astrobiology’s near-term targets: Mars, Europa, Enceladus, and perhaps other Jovian or Saturnian moons. Microorganisms, both eukaryotic and prokaryotic, remain active within brine channels inside the ice, making it unnecessary to penetrate through to liquid water below in order to detect life. We have developed a submersible digital holographic microscope (DHM) that is capable of resolving individual bacterial cells, and demonstrated its utility for immediately imaging samples taken directly from sea ice at several locations near Nuuk, Greenland. In all samples, the appearance and motility of eukaryotes were conclusive signs of life. The appearance of prokaryotic cells alone was not sufficient to confirm life, but when prokaryotic motility occurred, it was rapid and conclusive. Warming the samples to above-freezing temperatures or supplementing with serine increased the number of motile cells and the speed of motility; supplementing with serine also stimulated chemotaxis. These results show that DHM is a useful technique for detection of active organisms in extreme environments, and that motility may be used as a biosignature in the liquid brines that persist in ice. These findings have important implications for the design of missions to icy environments and suggest ways in which DHM imaging may be integrated with chemical life-detection suites in order to create more conclusive life detection packages. PMID:26812683

A Submersible, Off-Axis Holographic Microscope for Detection of Microbial Motility and Morphology in Aqueous and Icy Environments.

PubMed

Lindensmith, Christian A; Rider, Stephanie; Bedrossian, Manuel; Wallace, J Kent; Serabyn, Eugene; Showalter, G Max; Deming, Jody W; Nadeau, Jay L

2016-01-01

Sea ice is an analog environment for several of astrobiology's near-term targets: Mars, Europa, Enceladus, and perhaps other Jovian or Saturnian moons. Microorganisms, both eukaryotic and prokaryotic, remain active within brine channels inside the ice, making it unnecessary to penetrate through to liquid water below in order to detect life. We have developed a submersible digital holographic microscope (DHM) that is capable of resolving individual bacterial cells, and demonstrated its utility for immediately imaging samples taken directly from sea ice at several locations near Nuuk, Greenland. In all samples, the appearance and motility of eukaryotes were conclusive signs of life. The appearance of prokaryotic cells alone was not sufficient to confirm life, but when prokaryotic motility occurred, it was rapid and conclusive. Warming the samples to above-freezing temperatures or supplementing with serine increased the number of motile cells and the speed of motility; supplementing with serine also stimulated chemotaxis. These results show that DHM is a useful technique for detection of active organisms in extreme environments, and that motility may be used as a biosignature in the liquid brines that persist in ice. These findings have important implications for the design of missions to icy environments and suggest ways in which DHM imaging may be integrated with chemical life-detection suites in order to create more conclusive life detection packages.

Changes of sperm quality and hormone receptors in the rat testis after exposure to methamphetamine.

PubMed

Nudmamud-Thanoi, Sutisa; Sueudom, Wanvipa; Tangsrisakda, Nareelak; Thanoi, Samur

2016-10-01

Methamphetamine (METH) is known to damage neurons and induce psychosis. It can also induce apoptosis in seminiferous tubules and affect sperm quality. The present study was carried out to investigate the effect of a rat model of METH addiction on sperm quality and expression of progesterone receptors (PR) and estrogen receptors (ER) in the testis. Sperm quality parameters including sperm motility, sperm morphology and sperm concentration were examined. Protein and gene expressions PR, ERα and ERβ were studied using immunohistochemistry and reverse transcriptase-polymerase chain reaction, respectively. The percentages of normal sperm motility and normal sperm morphology were significantly decreased in animals receiving METH, especially in escalating dose (ED METH) and escalating dose-binge (ED-binge METH) groups when compared with control. In addition, sperm concentrations in ED METH and ED-binge METH groups were numerically decreased. PR, ERα and ERβ immunoreactive cells were significantly decreased in spermatogonia, spermatogenic cells and especially in Sertoli cells in all METH-treated groups. Furthermore, messenger RNA expression of PR, ERα and ERβ were also significantly decreased in all METH-treated animals. These results indicate that METH can induce abnormal sperm quality. These changes of sperm quality may relate to the reduction of PR, ERα and ERβ expressions in male germ cells and Sertoli cells which are essential for spermatogenesis and development of sperm.

Resolvin E1 inhibits dendritic cell migration in the skin and attenuates contact hypersensitivity responses

PubMed Central

Sawada, Yu; Hanakawa, Sho; Nakamizo, Satoshi; Murata, Teruasa; Ueharaguchi-Tanada, Yuri; Ono, Sachiko; Amano, Wataru; Nakajima, Saeko; Egawa, Gyohei; Tanizaki, Hideaki; Otsuka, Atsushi; Kitoh, Akihiko; Dainichi, Teruki; Ogawa, Narihito; Kobayashi, Yuichi; Yokomizo, Takehiko; Arita, Makoto; Nakamura, Motonobu; Miyachi, Yoshiki

2015-01-01

Resolvin E1 (RvE1) is a lipid mediator derived from ω3 polyunsaturated fatty acids that exerts potent antiinflammatory roles in several murine models. The antiinflammatory mechanism of RvE1 in acquired immune responses has been attributed to attenuation of cytokine production by dendritic cells (DCs). In this study, we newly investigated the effect of RvE1 on DC motility using two-photon microscopy in a contact hypersensitivity (CHS) model and found that RvE1 impaired DC motility in the skin. In addition, RvE1 attenuated T cell priming in the draining lymph nodes and effector T cell activation in the skin, which led to the reduced skin inflammation in CHS. In contrast, leukotriene B4 (LTB4) induced actin filament reorganization in DCs and increased DC motility by activating Cdc42 and Rac1 via BLT1, which was abrogated by RvE1. Collectively, our results suggest that RvE1 attenuates cutaneous acquired immune responses by inhibiting cutaneous DC motility, possibly through LTB4-BLT1 signaling blockade. PMID:26438363

Involvement of FFA1 and FFA4 in the regulation of cellular functions during tumor progression in colon cancer cells.

PubMed

Takahashi, Kaede; Fukushima, Kaori; Onishi, Yuka; Minami, Kanako; Otagaki, Shiho; Ishimoto, Kaichi; Fukushima, Nobuyuki; Honoki, Kanya; Tsujiuchi, Toshifumi

2018-08-01

Free fatty acid receptor 1 (FFA1) and FFA4 mediate a variety of biological responses through binding of medium- and long-chain free fatty acids. The aim of this study was to investigate an involvement of FFA1 and FFA4 in the regulation of cellular functions during tumor progression in colon cancer cells. The long-term fluorouracil (5-FU) and cisplatin (CDDP) treated cells were generated from DLD1 cells (DLD-5FU and DLD-CDDP cells, respectively). FFAR1 expressions were lower in DLD-5FU and DLD-CDDP cells than in DLD1 cells. In contrast, DLD-5FU and DLD-CDDP cells showed the high FFAR4 expressions, compared with DLD1 cells. The cell motile activities of DLD-5FU and DLD-CDDP cells were reduced by GW9508 which is an agonist of FFA1 and FFA4. Moreover, GW1100, an antagonist of FFA1, inhibited the cell motile activities of DLD-5FU and DLD-CDDP cells. To evaluate whether FFA1 and FFA4 regulate the enhancement of cell motility, invasion and colony formation, highly migratory (hmDLD1) cells were established from DLD1 cells. FFAR1 expression was significantly higher in hmDLD1 cells than in DLD1 cells, but no change of FFAR4 expression was observed. The elevated cell motile and invasive activities and colony formation of hmDLD1 cells were suppressed by FFA1 inhibition. These results suggest that FFA1 and FFA4 are involved in the regulation of cellular functions during tumor progression in colon cancer DLD1 cells. Copyright © 2018 Elsevier Inc. All rights reserved.

Migration of fresh and cryopreserved human spermatozoa in polyacrylamide gel.

PubMed

Goldstein, M C; Wix, L S; Foote, R H; Feldschuh, R; Feldschuh, J

1982-05-01

The ability of freshly collected and frozen human spermatozoa to migrate in round capillary tubes containing specially formulated polyacrylamide gel was investigated, using 33 ejaculates from 27 donors. Each semen sample was divided; one portion was left undiluted, and the other portion was diluted to 50 x 10(6) sperm/ml. Glycerol was used as the cryoprotectant. The percentage of motile sperm cells was determined before and after freezing. Fresh semen contained a higher percentage of motile cells, which migrated farther than those of cryopreserved-thawed semen. Various correlations between the percentage of motile sperm and migration distance ranged from 0.57 to 0.62. There was a low positive correlation of migration distance with sperm cell concentration per milliliter, r = 0.25 to 0.34; and thus adjusting semen samples to a standard sperm concentration improved the accuracy of the test only slightly. The regression coefficient of migration distance on the percentage of motile sperm in fresh semen was 0.65, indicating that for each 10% increase in sperm motility, migration distance is predicted to increase 6.5 mm. Five batches of polyacrylamide gel gave uniform results, and the application of this stable gel to fertility investigations is discussed.

Emergence of complex behavior in pili-based motility in early stages of P. aeruginosa surface adaptation

NASA Astrophysics Data System (ADS)

Brill-Karniely, Yifat; Jin, Fan; Wong, Gerard C. L.; Frenkel, Daan; Dobnikar, Jure

2017-04-01

Pseudomonas aeruginosa move across surfaces by using multiple Type IV Pili (TFP), motorized appendages capable of force generation via linear extension/retraction cycles, to generate surface motions collectively known as twitching motility. Pseudomonas cells arrive at a surface with low levels of piliation and TFP activity, which both progressively increase as the cells sense the presence of a surface. At present, it is not clear how twitching motility emerges from these initial minimal conditions. Here, we build a simple model for TFP-driven surface motility without complications from viscous and solid friction on surfaces. We discover the unanticipated structural requirement that TFP motors need to have a minimal amount of effective angular rigidity in order for cells to perform the various classes of experimentally-observed motions. Moreover, a surprisingly small number of TFP are needed to recapitulate movement signatures associated with twitching: Two TFP can already produce movements reminiscent of recently observed slingshot type motion. Interestingly, jerky slingshot motions characteristic of twitching motility comprise the transition region between different types of observed crawling behavior in the dynamical phase diagram, such as self-trapped localized motion, 2-D diffusive exploration, and super-diffusive persistent motion.

T Lymphocyte Migration: An Action Movie Starring the Actin and Associated Actors.

PubMed

Dupré, Loïc; Houmadi, Raïssa; Tang, Catherine; Rey-Barroso, Javier

2015-01-01

The actin cytoskeleton is composed of a dynamic filament meshwork that builds the architecture of the cell to sustain its fundamental properties. This physical structure is characterized by a continuous remodeling, which allows cells to accomplish complex motility steps such as directed migration, crossing of biological barriers, and interaction with other cells. T lymphocytes excel in these motility steps to ensure their immune surveillance duties. In particular, actin cytoskeleton remodeling is a key to facilitate the journey of T lymphocytes through distinct tissue environments and to tune their stop and go behavior during the scanning of antigen-presenting cells. The molecular mechanisms controlling actin cytoskeleton remodeling during T lymphocyte motility have been only partially unraveled, since the function of many actin regulators has not yet been assessed in these cells. Our review aims to integrate the current knowledge into a comprehensive picture of how the actin cytoskeleton drives T lymphocyte migration. We will present the molecular actors that control actin cytoskeleton remodeling, as well as their role in the different T lymphocyte motile steps. We will also highlight which challenges remain to be addressed experimentally and which approaches appear promising to tackle them.

Coupling of protein localization and cell movements by a dynamically localized response regulator in Myxococcus xanthus

PubMed Central

Leonardy, Simone; Freymark, Gerald; Hebener, Sabrina; Ellehauge, Eva; Søgaard-Andersen, Lotte

2007-01-01

Myxococcus xanthus cells harbor two motility machineries, type IV pili (Tfp) and the A-engine. During reversals, the two machineries switch polarity synchronously. We present a mechanism that synchronizes this polarity switching. We identify the required for motility response regulator (RomR) as essential for A-motility. RomR localizes in a bipolar, asymmetric pattern with a large cluster at the lagging cell pole. The large RomR cluster relocates to the new lagging pole in parallel with cell reversals. Dynamic RomR localization is essential for cell reversals, suggesting that RomR relocalization induces the polarity switching of the A-engine. The analysis of RomR mutants shows that the output domain targets RomR to the poles and the receiver domain is essential for dynamic localization. The small GTPase MglA establishes correct RomR polarity, and the Frz two-component system regulates dynamic RomR localization. FrzS localizes with Tfp at the leading pole and relocates in an Frz-dependent manner to the opposite pole during reversals; FrzS and RomR localize and oscillate independently. The Frz system synchronizes these oscillations and thus the synchronous polarity switching of the motility machineries. PMID:17932488

Visualizing Breast Cancer Cell Interaction with Tumor-Infiltrating Lymphocytes During Immunotherapy

DTIC Science & Technology

2013-04-01

in order to assess the motility cells in this tissue. To do that, CXCR6 GFP/+ mice were injected with 4T1 CFP cells, on day 30, metastasis were...Figure1: Imaging of metastasis A: Image of a metastase at day 30 after tumor implantation. 4T1-CFP tumor cells (blue), CXCR6 GFP/+ infiltrating...recognition or not. Figure 2 : Motility of CXCR6 -GFP+ cells in the periphery and core of the tumor. A-Image of 4T1-CFP tumor cells (blue), CXCR6 GFP

Evaluation of role of Notch3 signaling pathway in human lung cancer cells.

PubMed

Hassan, Wael Abdo; Yoshida, Ryoji; Kudoh, Shinji; Motooka, Yamato; Ito, Takaaki

2016-05-01

There is still a debate on the extent to which Notch3 signaling is involved in lung carcinogenesis and whether such function is dependent on cancer type or not. To evaluate Notch3 expression in different types of human lung cancer cells. Notch3 was detected in human lung cancer cell lines and in tissues. Then, small interfering RNA (siRNA) was used to down-regulate the expression of Notch3 in H69AR small cell lung carcinoma (SCLC) cells; two non-small cell lung carcinoma (NSCLC) cells; A549 adenocarcinoma (ADC); and H2170 squamous cell carcinoma (SCC). In addition, Notch3 intracellular domain (N3ICD) plasmid was transfected into H1688 human SCLC cells. We observed the effect of deregulating Notch3 signaling on the following cell properties: Notch-related proteins, cell morphology, adhesion, epithelial-mesenchymal transition (EMT), motility, proliferation and neuroendocrine (NE) features of SCLC. Notch3 is mainly expressed in NSCLC, and the expression of Notch1, Hes1 and Jagged1 is affected by Notch3. Notch3 has opposite functions in SCLC and NSCLC, being a tumor suppressor in the former and tumor promoting in the latter, in the context of cell adhesion, EMT and motility. Regarding cell proliferation, we found that inhibiting Notch3 in NSCLC decreases cell proliferation and induces apoptosis in NSCLC. Notch3 has no effect on cell proliferation or NE features of SCLC. Notch3 signaling in lung carcinoma is dependent on cell type. In SCLC, Notch3 behaves as a tumor suppressor pathway, while in NSCLC it acts as a tumor-promoting pathway.

Swimming Motility Reduces Deposition to Silica Surfaces

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Nanxi; Massoudieh, Arash; Liang, Xiaomeng

The role of swimming motility on bacterial transport and fate in porous media was evaluated. We present microscopic evidence showing that strong swimming motility reduces attachment of Azotobacter vinelandii cells to silica surfaces. Applying global and cluster statistical analyses to microscopic videos taken under non-flow conditions, wild type, flagellated A. vinelandii strain DJ showed strong swimming ability with an average speed of 13.1 μm/s, DJ77 showed impaired swimming averaged at 8.7 μm/s, and both the non-flagellated JZ52 and chemically treated DJ cells were non-motile. Quantitative analyses of trajectories observed at different distances above the collector of a radial stagnation pointmore » flow cell (RSPF) revealed that both swimming and non-swimming cells moved with the flow when at a distance of at least 20 μm from the collector surface. Near the surface, DJ cells showed both horizontal and vertical movement diverging them from reaching surfaces, while chemically treated DJ cells moved with the flow to reach surfaces, suggesting that strong swimming reduced attachment. In agreement with the RSPF results, the deposition rates obtained for two-dimensional multiple-collector micromodels were also lowest for DJ, while DJ77 and JZ52 showed similar values. Strong swimming specifically reduced deposition on the upstream surfaces of the micromodel collectors.« less

Calcium/calmodulin and cAMP/protein kinase-A pathways regulate sperm motility in the stallion.

PubMed

Lasko, Jodi; Schlingmann, Karen; Klocke, Ann; Mengel, Grace Ann; Turner, Regina

2012-06-01

In spite of the importance of sperm motility to fertility in the stallion, little is known about the signaling pathways that regulate motility in this species. In other mammals, calcium/calmodulin signaling and the cyclic AMP/protein kinase-A pathway are involved in sperm motility regulation. We hypothesized that these pathways also were involved in the regulation of sperm motility in the stallion. Using immunoblotting, calmodulin and the calmodulin-dependent protein kinase II β were shown to be present in stallion sperm and with indirect immunofluorescence calmodulin was localized to the acrosome and flagellar principal piece. Additionally, inhibition of either calmodulin or protein kinase-A significantly reduced sperm motility without affecting viability. Following inhibition of calmodulin, motility was not restored with agonists of the cyclic AMP/protein kinase-A pathway. These data suggest that calcium/calmodulin and cyclic AMP/protein kinase-A pathways are involved in the regulation of stallion sperm motility. The failure of cyclic AMP/protein kinase-A agonists to restore motility of calmodulin inhibited sperm suggests that both pathways may be required to support normal motility. Copyright © 2012 Elsevier B.V. All rights reserved.

A new holistic 3D non-invasive analysis of cellular distribution and motility on fibroin-alginate microcarriers using light sheet fluorescent microscopy

PubMed Central

Pierini, Michela; Bevilacqua, Alessandro; Torre, Maria Luisa; Lucarelli, Enrico

2017-01-01

Cell interaction with biomaterials is one of the keystones to developing medical devices for tissue engineering applications. Biomaterials are the scaffolds that give three-dimensional support to the cells, and are vectors that deliver the cells to the injured tissue requiring repair. Features of biomaterials can influence the behaviour of the cells and consequently the efficacy of the tissue-engineered product. The adhesion, distribution and motility of the seeded cells onto the scaffold represent key aspects, and must be evaluated in vitro during the product development, especially when the efficacy of a specific tissue-engineered product depends on viable and functional cell loading. In this work, we propose a non-invasive and non-destructive imaging analysis for investigating motility, viability and distribution of Mesenchymal Stem Cells (MSCs) on silk fibroin-based alginate microcarriers, to test the adhesion capacity of the fibroin coating onto alginate which is known to be unsuitable for cell adhesion. However, in depth characterization of the biomaterial is beyond the scope of this paper. Scaffold-loaded MSCs were stained with Calcein-AM and Ethidium homodimer-1 to detect live and dead cells, respectively, and counterstained with Hoechst to label cell nuclei. Time-lapse Light Sheet Fluorescent Microscopy (LSFM) was then used to produce three-dimensional images of the entire cells-loaded fibroin/alginate microcarriers. In order to quantitatively track the cell motility over time, we also developed an open source user friendly software tool called Fluorescent Cell Tracker in Three-Dimensions (F-Tracker3D). Combining LSFM with F-Tracker3D we were able for the first time to assess the distribution and motility of stem cells in a non-invasive, non-destructive, quantitative, and three-dimensional analysis of the entire surface of the cell-loaded scaffold. We therefore propose this imaging technique as an innovative holistic tool for monitoring cell-biomaterial interactions, and as a tool for the design, fabrication and functionalization of a scaffold as a medical device. PMID:28817694

Silibinin inhibits fibronectin induced motility, invasiveness and survival in human prostate carcinoma PC3 cells via targeting integrin signaling

PubMed Central

Deep, Gagan; Kumar, Rahul; Jain, Anil K; Agarwal, Chapla; Agarwal, Rajesh

2014-01-01

Prostate cancer (PCA) is the 2nd leading cause of cancer-related deaths among men in the United States. Preventing or inhibiting metastasis-related events through non-toxic agents could be a useful approach for lowering high mortality among PCA patients. We have earlier reported that natural flavonoid silibinin possesses strong anti-metastatic efficacy against PCA however, mechanism/s of its action still remains largely unknown. One of the major events during metastasis is the replacement of cell-cell interaction with integrins-based cell-matrix interaction that controls motility, invasiveness and survival of cancer cells. Accordingly, here we examined silibinin effect on advanced human PCA PC3 cells' interaction with extracellular matrix component fibronectin. Silibinin (50-200 μM) treatment significantly decreased the fibronectin (5 μg/ml)-induced motile morphology via targeting actin cytoskeleton organization in PC3 cells. Silibinin also decreased the fibronectin-induced cell proliferation and motility but significantly increased cell death in PC3 cells. Silibinin also inhibited the PC3 cells invasiveness in Transwell invasion assays with fibronectin or cancer associated fibroblasts (CAFs) serving as chemoattractant. Importantly, PC3-luc cells cultured on fibronectin showed rapid dissemination and localized in lungs following tail vein injection in athymic male nude mice; however, in silibinin-treated PC3-luc cells, dissemination and lung localization was largely compromised. Molecular analyses revealed that silibinin treatment modulated the fibronectin-induced expression of integrins (α5, αV, β1 and β3), actin-remodeling (FAK, Src, GTPases, ARP2 and cortactin), apoptosis (cPARP and cleaved caspase 3), EMT (E-cadherin and β-catenin), and cell survival (survivin and Akt) related signaling molecules in PC3 cells. Furthermore, PC3-xenograft tissue analyses confirmed the inhibitory effect of silibinin on fibronectin and integrins expression. Together, these results showed that silibinin targets PCA cells' interaction with fibronectin and inhibits their motility, invasiveness and survival; thus further supporting silibinin use in PCA intervention including its metastatic progression. PMID:25285031

Silibinin inhibits fibronectin induced motility, invasiveness and survival in human prostate carcinoma PC3 cells via targeting integrin signaling.

PubMed

Deep, Gagan; Kumar, Rahul; Jain, Anil K; Agarwal, Chapla; Agarwal, Rajesh

2014-10-01

Prostate cancer (PCA) is the 2nd leading cause of cancer-related deaths among men in the United States. Preventing or inhibiting metastasis-related events through non-toxic agents could be a useful approach for lowering high mortality among PCA patients. We have earlier reported that natural flavonoid silibinin possesses strong anti-metastatic efficacy against PCA however, mechanism/s of its action still remains largely unknown. One of the major events during metastasis is the replacement of cell-cell interaction with integrins-based cell-matrix interaction that controls motility, invasiveness and survival of cancer cells. Accordingly, here we examined silibinin effect on advanced human PCA PC3 cells' interaction with extracellular matrix component fibronectin. Silibinin (50-200 μM) treatment significantly decreased the fibronectin (5 μg/ml)-induced motile morphology via targeting actin cytoskeleton organization in PC3 cells. Silibinin also decreased the fibronectin-induced cell proliferation and motility but significantly increased cell death in PC3 cells. Silibinin also inhibited the PC3 cells invasiveness in Transwell invasion assays with fibronectin or cancer associated fibroblasts (CAFs) serving as chemoattractant. Importantly, PC3-luc cells cultured on fibronectin showed rapid dissemination and localized in lungs following tail vein injection in athymic male nude mice; however, in silibinin-treated PC3-luc cells, dissemination and lung localization was largely compromised. Molecular analyses revealed that silibinin treatment modulated the fibronectin-induced expression of integrins (α5, αV, β1 and β3), actin-remodeling (FAK, Src, GTPases, ARP2 and cortactin), apoptosis (cPARP and cleaved caspase 3), EMT (E-cadherin and β-catenin), and cell survival (survivin and Akt) related signaling molecules in PC3 cells. Furthermore, PC3-xenograft tissue analyses confirmed the inhibitory effect of silibinin on fibronectin and integrins expression. Together, these results showed that silibinin targets PCA cells' interaction with fibronectin and inhibits their motility, invasiveness and survival; thus further supporting silibinin use in PCA intervention including its metastatic progression.

Endocytic reawakening of motility in jammed epithelia

NASA Astrophysics Data System (ADS)

Malinverno, Chiara; Corallino, Salvatore; Giavazzi, Fabio; Bergert, Martin; Li, Qingsen; Leoni, Marco; Disanza, Andrea; Frittoli, Emanuela; Oldani, Amanda; Martini, Emanuele; Lendenmann, Tobias; Deflorian, Gianluca; Beznoussenko, Galina V.; Poulikakos, Dimos; Ong, Kok Haur; Uroz, Marina; Trepat, Xavier; Parazzoli, Dario; Maiuri, Paolo; Yu, Weimiao; Ferrari, Aldo; Cerbino, Roberto; Scita, Giorgio

2017-05-01

Dynamics of epithelial monolayers has recently been interpreted in terms of a jamming or rigidity transition. How cells control such phase transitions is, however, unknown. Here we show that RAB5A, a key endocytic protein, is sufficient to induce large-scale, coordinated motility over tens of cells, and ballistic motion in otherwise kinetically arrested monolayers. This is linked to increased traction forces and to the extension of cell protrusions, which align with local velocity. Molecularly, impairing endocytosis, macropinocytosis or increasing fluid efflux abrogates RAB5A-induced collective motility. A simple model based on mechanical junctional tension and an active cell reorientation mechanism for the velocity of self-propelled cells identifies regimes of monolayer dynamics that explain endocytic reawakening of locomotion in terms of a combination of large-scale directed migration and local unjamming. These changes in multicellular dynamics enable collectives to migrate under physical constraints and may be exploited by tumours for interstitial dissemination.

Imaging host cell-Leishmania interaction dynamics implicates parasite motility, lysosome recruitment, and host cell wounding in the infection process.

PubMed

Forestier, Claire-Lise; Machu, Christophe; Loussert, Celine; Pescher, Pascale; Späth, Gerald F

2011-04-21

Leishmania donovani causes human visceral leishmaniasis. The parasite infectious cycle comprises extracellular flagellated promastigotes that proliferate inside the insect vector, and intracellular nonmotile amastigotes that multiply within infected host cells. Using primary macrophages infected with virulent metacyclic promastigotes and high spatiotemporal resolution microscopy, we dissect the dynamics of the early infection process. We find that motile promastigotes enter macrophages in a polarized manner through their flagellar tip and are engulfed into host lysosomal compartments. Persistent intracellular flagellar activity leads to reorientation of the parasite flagellum toward the host cell periphery and results in oscillatory parasite movement. The latter is associated with local lysosomal exocytosis and host cell plasma membrane wounding. These findings implicate lysosome recruitment followed by lysosome exocytosis, consistent with parasite-driven host cell injury, as key cellular events in Leishmania host cell infection. This work highlights the role of promastigote polarity and motility during parasite entry. Copyright © 2011 Elsevier Inc. All rights reserved.

Notch/Her12 signalling modulates, motile/immotile cilia ratio downstream of Foxj1a in zebrafish left-right organizer

PubMed Central

Sampaio, Pedro; Pestana, Sara; Pinto, Andreia; Vaz, Andreia; Roxo-Rosa, Mónica; Gardner, Rui; Lopes, Telma; Schilling, Britta; Henry, Ian; Saúde, Leonor

2017-01-01

Foxj1a is necessary and sufficient to specify motile cilia. Using transcriptional studies and slow-scan two-photon live imaging capable of identifying the number of motile and immotile cilia, we now established that the final number of motile cilia depends on Notch signalling (NS). We found that despite all left-right organizer (LRO) cells express foxj1a and the ciliary axonemes of these cells have dynein arms, some cilia remain immotile. We identified that this decision is taken early in development in the Kupffer’s Vesicle (KV) precursors the readout being her12 transcription. We demonstrate that overexpression of either her12 or Notch intracellular domain (NICD) increases the number of immotile cilia at the expense of motile cilia, and leads to an accumulation of immotile cilia at the anterior half of the KV. This disrupts the normal fluid flow intensity and pattern, with consequent impact on dand5 expression pattern and left-right (L-R) axis establishment. PMID:28875937

Malaria parasite LIMP protein regulates sporozoite gliding motility and infectivity in mosquito and mammalian hosts

PubMed Central

Santos, Jorge M; Egarter, Saskia; Zuzarte-Luís, Vanessa; Kumar, Hirdesh; Moreau, Catherine A; Kehrer, Jessica; Pinto, Andreia; da Costa, Mário; Franke-Fayard, Blandine; Janse, Chris J; Frischknecht, Friedrich; Mair, Gunnar R

2017-01-01

Gliding motility allows malaria parasites to migrate and invade tissues and cells in different hosts. It requires parasite surface proteins to provide attachment to host cells and extracellular matrices. Here, we identify the Plasmodium protein LIMP (the name refers to a gliding phenotype in the sporozoite arising from epitope tagging of the endogenous protein) as a key regulator for adhesion during gliding motility in the rodent malaria model P. berghei. Transcribed in gametocytes, LIMP is translated in the ookinete from maternal mRNA, and later in the sporozoite. The absence of LIMP reduces initial mosquito infection by 50%, impedes salivary gland invasion 10-fold, and causes a complete absence of liver invasion as mutants fail to attach to host cells. GFP tagging of LIMP caused a limping defect during movement with reduced speed and transient curvature changes of the parasite. LIMP is an essential motility and invasion factor necessary for malaria transmission. DOI: http://dx.doi.org/10.7554/eLife.24109.001 PMID:28525314

The CRISPR/Cas system inhibited the pro-oncogenic effects of alternatively spliced fibronectin extra domain A via editing the genome in salivary adenoid cystic carcinoma cells.

PubMed

Wang, H-C; Yang, Y; Xu, S-Y; Peng, J; Jiang, J-H; Li, C-Y

2015-07-01

To identify the association of fibronectin (FN) extra domain A (EDA) with the progression of salivary adenoid cystic carcinoma (SACC). Accordingly, the exclusion of EDA exon through the CRISPR/Cas9 system was investigated as the rescue for such pro-oncogenic splicing. SACC-83 cells were transiently transfected with plasmids containing recombinant EDA, and the cellular growth and motility were then accessed in vitro. Epithelial-mesenchymal transition (EMT) was investigated with immunohistochemistry, Western blot, and real-time PCR analysis. SACC tissues from 81 patients were used to access the associations between EDA+FN and clinical-pathological parameters. CRISPR/Cas9 plasmids containing sgRNA were designed and co-transfected into SACC-83 cells; the effects of EDA knockout on cellular growth and motility were then accessed. The recombinant EDA exhibited little effect on the proliferation of SACC cells, but significantly promoted the migration and invasion of the cells (P < 0.05), accompanied with upregulated EMT (P < 0.05); consistently, the expression of EDA+FN was positively associated with the metastasis, nerve invasion and recurrence of SACC (P < 0.05). Furthermore, the EDA knockout from the FN gene in most SACC cells resulted in a decrease in cell motility and invasion, as well as prolonged population doubling time, compared with untreated SACC-83 cells (P < 0.05). The EDA domain significantly promoted the motility of SACC cells, and positively associated with the tumor progression in patients with SACC. Thus, it is a potential risk factor and also a therapeutic target for SACC. The CRISPR/Cas9 system may control a pro-oncogenic splicing process through the exclusion of EDA exon from the FN gene, leading to inhibition of motility, invasion and proliferation of cancer cells. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Potent blockade of hepatocyte growth factor-stimulated cell motility, matrix invasion and branching morphogenesis by antagonists of Grb2 Src homology 2 domain interactions.

PubMed

Atabey, N; Gao, Y; Yao, Z J; Breckenridge, D; Soon, L; Soriano, J V; Burke, T R; Bottaro, D P

2001-04-27

Hepatocyte growth factor (HGF) stimulates mitogenesis, motogenesis, and morphogenesis in a wide range of cellular targets during development, homeostasis and tissue regeneration. Inappropriate HGF signaling occurs in several human cancers, and the ability of HGF to initiate a program of protease production, cell dissociation, and motility has been shown to promote cellular invasion and is strongly linked to tumor metastasis. Upon HGF binding, several tyrosines within the intracellular domain of its receptor, c-Met, become phosphorylated and mediate the binding of effector proteins, such as Grb2. Grb2 binding through its SH2 domain is thought to link c-Met with downstream mediators of cell proliferation, shape change, and motility. We analyzed the effects of Grb2 SH2 domain antagonists on HGF signaling and observed potent blockade of cell motility, matrix invasion, and branching morphogenesis, with ED(50) values of 30 nm or less, but only modest inhibition of mitogenesis. These compounds are 1000-10,000-fold more potent anti-motility agents than any previously characterized Grb2 SH2 domain antagonists. Our results suggest that SH2 domain-mediated c-Met-Grb2 interaction contributes primarily to the motogenic and morphogenic responses to HGF, and that these compounds may have therapeutic application as anti-metastatic agents for tumors where the HGF signaling pathway is active.

Gliding Motility of Babesia bovis Merozoites Visualized by Time-Lapse Video Microscopy

PubMed Central

Asada, Masahito; Goto, Yasuyuki; Yahata, Kazuhide; Yokoyama, Naoaki; Kawai, Satoru; Inoue, Noboru; Kaneko, Osamu; Kawazu, Shin-ichiro

2012-01-01

Background Babesia bovis is an apicomplexan intraerythrocytic protozoan parasite that induces babesiosis in cattle after transmission by ticks. During specific stages of the apicomplexan parasite lifecycle, such as the sporozoites of Plasmodium falciparum and tachyzoites of Toxoplasma gondii, host cells are targeted for invasion using a unique, active process termed “gliding motility”. However, it is not thoroughly understood how the merozoites of B. bovis target and invade host red blood cells (RBCs), and gliding motility has so far not been observed in the parasite. Methodology/Principal Findings Gliding motility of B. bovis merozoites was revealed by time-lapse video microscopy. The recorded images revealed that the process included egress of the merozoites from the infected RBC, gliding motility, and subsequent invasion into new RBCs. The gliding motility of B. bovis merozoites was similar to the helical gliding of Toxoplasma tachyzoites. The trails left by the merozoites were detected by indirect immunofluorescence assay using antiserum against B. bovis merozoite surface antigen 1. Inhibition of gliding motility by actin filament polymerization or depolymerization indicated that the gliding motility was driven by actomyosin dependent process. In addition, we revealed the timing of breakdown of the parasitophorous vacuole. Time-lapse image analysis of membrane-stained bovine RBCs showed formation and breakdown of the parasitophorous vacuole within ten minutes of invasion. Conclusions/Significance This is the first report of the gliding motility of B. bovis. Since merozoites of Plasmodium parasites do not glide on a substrate, the gliding motility of B. bovis merozoites is a notable finding. PMID:22506073

Emerging ciliopathies: are respiratory cilia compromised in Usher syndrome?

PubMed

Piatti, G; De Santi, M M; Brogi, M; Castorina, P; Ambrosetti, U

2014-01-01

Usher syndrome is a ciliopathy involving photoreceptors and cochlear hair cells (sensory cilia): since sensory and motor ciliopathies can overlap, we analysed the respiratory cilia (motile) in 17 patients affected by Usher syndrome and 18 healthy control subject. We studied the mucociliary transport time with the saccharine test, ciliary motility and ultrastructure of respiratory cilia obtained by nasal brushing; we also recorded the classical respiratory function values by spirometry. All enrolled subjects showed normal respiratory function values. The mean mucociliary transport time with saccharine was 22.33 ± 17.96 min, which is in the range of normal values. The mean ciliary beat frequency of all subjects was 8.81 ± 2.18 Hz, which is a value approaching the lower physiological limit. None of the classical ciliary alterations characterizing the "ciliary primary dyskinesia" was detected, although two patients showed alterations in number and arrangement of peripheral microtubules and one patient had abnormal ciliary roots. Respiratory cilia in Usher patients don't seem to have evident ultrastructural alterations, as expected, but the fact that the ciliary motility appeared slightly reduced could emphasize that a rigid distinction between sensory and motor ciliopathies may not reflect what really occurs. Copyright © 2014 Elsevier Inc. All rights reserved.

Computerized in vitro test for chemical toxicity based on tetrahymena swimming patterns

NASA Technical Reports Server (NTRS)

Noever, David A.; Matsos, Helen C.; Cronise, Raymond J.; Looger, Loren L.; Relwani, Rachna A.; Johnson, Jacqueline U.

1994-01-01

An apparatus and method for rapidly determining chemical toxicity was evaluated. The toxicity monitor includes an automated scoring of how motile biological cells (Tetrahymena pyriformis) slow down or otherwise change their swimming patterns in a hostile chemical environment. The device, called the Motility Assay Apparatus (MAA) is tested for 30 second determination of chemical toxicity in 20 aqueous samples containing trace organics and salts. With equal or better detection limits, results compare favorably to in vivo animal tests of eye irritancy, in addition to agreeing for all chemicals with previous manual evaluations of single cell motility.

Guava Leaf Extract Inhibits Quorum-Sensing and Chromobacterium violaceum Induced Lysis of Human Hepatoma Cells: Whole Transcriptome Analysis Reveals Differential Gene Expression

PubMed Central

Tiwary, Bipransh Kumar; Kumar, Anoop

2014-01-01

Quorum sensing (QS) is a process mediated via small molecules termed autoinducers (AI) that allow bacteria to respond and adjust according to the cell population density by altering the expression of multitudinous genes. Since QS governs numerous bioprocesses in bacteria, including virulence, its inhibition promises to be an ideal target for the development of novel therapeutics. We found that the aqueous leaf extract of Psidium guajava (GLE) exhibited anti-QS properties as evidenced by inhibition of violacein production in Chromobacterium violaceum and swarming motility of Pseudomonas aeruginosa. The gram-negative bacterium, C. violaceum is a rare pathogen with high mortality rate. In this study, perhaps for the first time, we identified the target genes of GLE in C. violaceum MTCC 2656 by whole transcriptome analysis on Ion Torrent. Our data revealed that GLE significantly down-regulated 816 genes at least three fold, with p value≤0.01, which comprises 19% of the C. violaceum MTCC 2656 genome. These genes were distributed throughout the genome and were associated with virulence, motility and other cellular processes, many of which have been described as quorum regulated in C. violaceum and other gram negative bacteria. Interestingly, GLE did not affect the growth of the bacteria. However, consistent with the gene expression pattern, GLE treated C. violaceum cells were restrained from causing lysis of human hepatoma cell line, HepG2, indicating a positive relationship between the QS-regulated genes and pathogenicity. Overall, our study proposes GLE as a QS inhibitor (QSI) with the ability to attenuate virulence without affecting growth. To the best of our knowledge, this is the first report which provides with a plausible set of candidate genes regulated by the QS system in the neglected pathogen C. violaceum. PMID:25229331

Guava leaf extract inhibits quorum-sensing and Chromobacterium violaceum induced lysis of human hepatoma cells: whole transcriptome analysis reveals differential gene expression.

PubMed

Ghosh, Runu; Tiwary, Bipransh Kumar; Kumar, Anoop; Chakraborty, Ranadhir

2014-01-01

Quorum sensing (QS) is a process mediated via small molecules termed autoinducers (AI) that allow bacteria to respond and adjust according to the cell population density by altering the expression of multitudinous genes. Since QS governs numerous bioprocesses in bacteria, including virulence, its inhibition promises to be an ideal target for the development of novel therapeutics. We found that the aqueous leaf extract of Psidium guajava (GLE) exhibited anti-QS properties as evidenced by inhibition of violacein production in Chromobacterium violaceum and swarming motility of Pseudomonas aeruginosa. The gram-negative bacterium, C. violaceum is a rare pathogen with high mortality rate. In this study, perhaps for the first time, we identified the target genes of GLE in C. violaceum MTCC 2656 by whole transcriptome analysis on Ion Torrent. Our data revealed that GLE significantly down-regulated 816 genes at least three fold, with p value ≤ 0.01, which comprises 19% of the C. violaceum MTCC 2656 genome. These genes were distributed throughout the genome and were associated with virulence, motility and other cellular processes, many of which have been described as quorum regulated in C. violaceum and other gram negative bacteria. Interestingly, GLE did not affect the growth of the bacteria. However, consistent with the gene expression pattern, GLE treated C. violaceum cells were restrained from causing lysis of human hepatoma cell line, HepG2, indicating a positive relationship between the QS-regulated genes and pathogenicity. Overall, our study proposes GLE as a QS inhibitor (QSI) with the ability to attenuate virulence without affecting growth. To the best of our knowledge, this is the first report which provides with a plausible set of candidate genes regulated by the QS system in the neglected pathogen C. violaceum.

The identification of specialized pacemaking cells in the anal sphincters.

PubMed

Shafik, Ahmed; El Sibai, Olfat; Ahmed, Ismail

2006-07-01

Interstitial cells of Cajal (ICC) are claimed to generate the electrical activity in the colon and stomach. As the external (EAS) and internal (IAS) anal sphincters exhibit resting electrical activity, we hypothesized the presence of ICC in these sphincters. This hypothesis was investigated in the current study. Specimens from the EAS and IAS were taken from normal areas of the anorectum which had been surgically excised by abdominoperineal operation for rectal cancer of 28 patients (16 men, 12 women, mean age 42.2+/-4.8 years). The specimens were subjected to c-kit immunohistochemistry. Controls for the specificity of the antisera consisted of tissue incubation with normal rabbit serum substituted for the primary antiserum. Fusiform, c-kit positive, ICC-like cells were detected in the anal sphincters; they had dendritic processes. They were clearly distinguishable from the non-branching, c-kit negative smooth and striated muscle cells of the anal sphincters. The specimens contained also c-kit positive mast cells, but they had a rounded body with no dendritic processes. Immunoreactivity was absent in negative controls in which the primary antibody was omitted. We have identified, for the first time, cells in EAS and IAS with morphological and immunological phenotypes similar to ICCs of the gut. These cells appear to be responsible for initiating the slow waves recorded from the anal sphincters and for controlling their activity. A deficiency or absence of these cells may affect the anal motile activity. Studies are needed to explore the role of these cells in anal motility disorders.

Targeting the Human Papillomavirus E6 and E7 Oncogenes through Expression of the Bovine Papillomavirus Type 1 E2 Protein Stimulates Cellular Motility▿†

PubMed Central

Morrison, Monique A.; Morreale, Richard J.; Akunuru, Shailaja; Kofron, Matthew; Zheng, Yi; Wells, Susanne I.

2011-01-01

Expression of the high-risk human papillomavirus (HPV) E6 and E7 oncogenes is essential for the initiation and maintenance of cervical cancer. The repression of both was previously shown to result in activation of their respective tumor suppressor targets, p53 and pRb, and subsequent senescence induction in cervical cancer cells. Consequently, viral oncogene suppression is a promising approach for the treatment of HPV-positive tumors. One well-established method of E6/E7 repression involves the reexpression of the viral E2 protein which is usually deleted in HPV-positive cancer cells. Here, we show that, surprisingly, bovine papillomavirus type 1 (BPV1) E2 but not RNA interference-mediated E6/E7 repression in HPV-positive cervical cancer cells stimulates cellular motility and invasion. Migration correlated with the dynamic formation of cellular protrusions and was dependent upon cell-to-cell contact. While E2-expressing migratory cells were senescent, migration was not a general feature of cellular senescence or cell cycle arrest and was specifically observed in HPV-positive cervical cancer cells. Interestingly, E2-expressing cells not only were themselves motile but also conferred increased motility to admixed HeLa cervical cancer cells. Together, our data suggest that repression of the viral oncogenes by E2 stimulates the motility of E6/E7-targeted cells as well as adjacent nontargeted cancer cells, thus raising the possibility that E2 expression may unfavorably increase the local invasiveness of HPV-positive tumors. PMID:21835799

Generation of Compartmentalized Pressure by a Nuclear Piston Governs Cell Motility in 3D Matrix*

PubMed Central

Petrie, Ryan J.; Koo, Hyun; Yamada, Kenneth M.

2017-01-01

Cells use actomyosin contractility to move through three-dimensional (3D) extracellular matrix. Contractility affects the type of protrusions cells use to migrate in 3D, but the mechanisms are unclear. Here we found that contractility generated high-pressure lobopodial protrusions in cells migrating in a 3D matrix. In these cells, the nucleus physically divided the cytoplasm into forward and rear compartments. Actomyosin contractility with the nucleoskeleton-intermediate filament linker protein nesprin 3 pulled the nucleus forward and pressurized the front of the cell. Reducing expression of nesprin 3 reduced and equalized the intracellular pressure. Thus, the nucleus can act as a piston that physically compartmentalizes the cytoplasm and increases the hydrostatic pressure between the nucleus and the leading edge to drive lamellipodia-independent 3D cell migration. PMID:25170155

The dynamic instability in the hook/flagellum system that triggers bacterial flicks

NASA Astrophysics Data System (ADS)

Jabbarzadeh, Mehdi; Fu, Henry

2017-11-01

Dynamical bending, buckling, and polymorphic transformations of the flagellum are known to affect bacterial motility, but run-reverse-flick motility of monotrichous bacteria also involves the even more flexible hook, which connects the flagellum to the cell body. Here, we identify the dynamic buckling mechanism that produces flicks in Vibrio alginolyticus. Estimates of forces and torques on the hook from experimental observations suggest that flicks are triggered at stresses below the hook's static Euler buckling criterion. Using an accurate linearization of the Kirchoff rod model for the hook in a model of a swimming bacterium with rigid flagellum, we show that as hook stiffness decreases there is a transition from on-axis flagellar rotation with small hook deflections to flagellar precession with large deflections. When flagellum flexibility is incorporated, the precession is disrupted by significant flagellar bending - i.e., incipient flicks. The predicted onset of dynamic instabilities corresponds well with experimentally observed flick events.

Human basal body basics.

PubMed

Vertii, Anastassiia; Hung, Hui-Fang; Hehnly, Heidi; Doxsey, Stephen

2016-01-01

In human cells, the basal body (BB) core comprises a ninefold microtubule-triplet cylindrical structure. Distal and subdistal appendages are located at the distal end of BB, where they play indispensable roles in cilium formation and function. Most cells that arrest in the G0 stage of the cell cycle initiate BB docking at the plasma membrane followed by BB-mediated growth of a solitary primary cilium, a structure required for sensing the extracellular environment and cell signaling. In addition to the primary cilium, motile cilia are present in specialized cells, such as sperm and airway epithelium. Mutations that affect BB function result in cilia dysfunction. This can generate syndromic disorders, collectively called ciliopathies, for which there are no effective treatments. In this review, we focus on the features and functions of BBs and centrosomes in Homo sapiens.

Unique Directional Motility of Influenza C Virus Controlled by Its Filamentous Morphology and Short-Range Motions.

PubMed

Sakai, Tatsuya; Takagi, Hiroaki; Muraki, Yasushi; Saito, Mineki

2018-01-15

Influenza virus motility is based on cooperation between two viral spike proteins, hemagglutinin (HA) and neuraminidase (NA), and is a major determinant of virus infectivity. To translocate a virus particle on the cell surface, HA molecules exchange viral receptors and NA molecules accelerate the receptor exchange of HA. This type of virus motility was recently identified in influenza A virus (IAV). To determine if other influenza virus types have a similar receptor exchange mechanism-driven motility, we investigated influenza C virus (ICV) motility on a receptor-fixed glass surface. This system excludes receptor mobility, which makes it more desirable than a cell surface for demonstrating virus motility by receptor exchange. Like IAV, ICV was observed to move across the receptor-fixed surface. However, in contrast to the random movement of IAV, a filamentous ICV strain, Ann Arbor/1/50 (AA), moved in a straight line, in a directed manner, and at a constant rate, whereas a spherical ICV strain, Taylor/1233/47 (Taylor), moved randomly, similar to IAV. The AA and Taylor viruses each moved with a combination of gradual (crawling) and rapid (gliding) motions, but the distances of crawling and gliding for the AA virus were shorter than those of the Taylor virus. Our findings indicate that like IAV, ICV also has a motility that is driven by the receptor exchange mechanism. However, compared with IAV movement, filamentous ICV movement is highly regulated in both direction and speed. Control of ICV movement is based on its specific motility employing short crawling and gliding motions as well as its own filamentous morphology. IMPORTANCE Influenza virus enters into a host cell for infection via cellular endocytosis. Human influenza virus infects epithelial cells of the respiratory tract, the surfaces of which are hidden by abundant cilia that are inactive in endocytosis. An open question is the manner by which the virus migrates to endocytosis-active domains. In analyzing individual virus behaviors through single-virus tracking, we identified a novel function of the hemagglutinin and esterase of influenza C virus (ICV) as the motility machinery. Hemagglutinin iteratively exchanges a viral receptor, causing virus movement. Esterase degrades the receptors along the trajectory traveled by the virus and prevents the virus from moving backward, causing directional movement. We propose that ICV has a unique motile machinery directionally controlled via hemagglutinin sensing the receptor density manipulated by esterase. Copyright © 2018 Sakai et al.

Inactivation of ferric uptake regulator (Fur) attenuates Helicobacter pylori J99 motility by disturbing the flagellar motor switch and autoinducer-2 production.

PubMed

Lee, Ai-Yun; Kao, Cheng-Yen; Wang, Yao-Kuan; Lin, Ssu-Yuan; Lai, Tze-Ying; Sheu, Bor-Shyang; Lo, Chien-Jung; Wu, Jiunn-Jong

2017-08-01

Flagellar motility of Helicobacter pylori has been shown to be important for the bacteria to establish initial colonization. The ferric uptake regulator (Fur) is a global regulator that has been identified in H. pylori which is involved in the processes of iron uptake and establishing colonization. However, the role of Fur in H. pylori motility is still unclear. Motility of the wild-type, fur mutant, and fur revertant J99 were determined by a soft-agar motility assay and direct video observation. The bacterial shape and flagellar structure were evaluated by transmission electron microscopy. Single bacterial motility and flagellar switching were observed by phase-contrast microscopy. Autoinducer-2 (AI-2) production in bacterial culture supernatant was analyzed by a bioluminescence assay. The fur mutant showed impaired motility in the soft-agar assay compared with the wild-type J99 and fur revertant. The numbers and lengths of flagellar filaments on the fur mutant cells were similar to those of the wild-type and revertant cells. Phenotypic characterization showed similar swimming speed but reduction in switching rate in the fur mutant. The AI-2 production of the fur mutant was dramatically reduced compared with wild-type J99 in log-phase culture medium. These results indicate that Fur positively modulates H. pylori J99 motility through interfering with bacterial flagellar switching. © 2017 John Wiley & Sons Ltd.

Rifaximin decreases virulence of Crohn's disease-associated Escherichia coli and epithelial inflammatory responses.

PubMed

Dogan, Belgin; Fu, Jing; Zhang, Shiying; Scherl, Ellen J; Simpson, Kenneth W

2018-05-01

Escherichia coli with an adherent and invasive pathotype (AIEC) is implicated in the pathogenesis of Crohn's disease (CD). Rifaximin improves symptoms in mild-to-moderate CD. It is unclear if this outcome is due to its effects on bacteria or intestinal epithelial inflammatory responses. We examined the effects of rifaximin on the growth and virulence of CD-associated E. coli and intestinal epithelial inflammatory responses. Seven well-characterized CD-associated E. coli strains (six AIEC, one non-AIEC; four rifaximin-resistant, three sensitive) were evaluated. We assessed the effects of rifaximin on CD-associated E. coli growth, adhesion to, and invasion of epithelial cells, virulence gene expression, motility, and survival in macrophages. Additionally, we determined the effects of rifaximin on intestinal epithelial inflammatory responses. In vitro rifaximin exerted a dose-dependent effect on the growth of sensitive strains but did not affect the growth of resistant strains. Rifaximin reduced adhesion, invasion, virulence gene expression and motility of CD-associated E. coli in a manner that was independent of its antimicrobial effect. Furthermore, rifaximin reduced IL-8 secretion from pregnane X receptor-expressing T84 colonic epithelial cells. The effect of rifaximin on adhesion was largely attributable to its action on bacteria, whereas decreases in invasion and cytokine secretion were due to its effect on the epithelium. In conclusion, our results show that rifaximin interferes with multiple steps implicated in host-AIEC interactions related to CD, including adhesion to, and invasion of epithelial cells, virulence gene expression, motility, and pro-inflammatory cytokine secretion. Further study is required to determine the relationship of these effects to clinical responses in CD patients.

Spontaneous symmetry breaking in active droplets provides a generic route to motility

PubMed Central

Tjhung, Elsen; Marenduzzo, Davide; Cates, Michael E.

2012-01-01

We explore a generic mechanism whereby a droplet of active matter acquires motility by the spontaneous breakdown of a discrete symmetry. The model we study offers a simple representation of a “cell extract” comprising, e.g., a droplet of actomyosin solution. (Such extracts are used experimentally to model the cytoskeleton). Actomyosin is an active gel whose polarity describes the mean sense of alignment of actin fibres. In the absence of polymerization and depolymerization processes (‘treadmilling’), the gel’s dynamics arises solely from the contractile motion of myosin motors; this should be unchanged when polarity is inverted. Our results suggest that motility can arise in the absence of treadmilling, by spontaneous symmetry breaking (SSB) of polarity inversion symmetry. Adapting our model to wall-bound cells in two dimensions, we find that as wall friction is reduced, treadmilling-induced motility falls but SSB-mediated motility rises. The latter might therefore be crucial in three dimensions where frictional forces are likely to be modest. At a supracellular level, the same generic mechanism can impart motility to aggregates of nonmotile but active bacteria; we show that SSB in this (extensile) case leads generically to rotational as well as translational motion. PMID:22797894

Motility and more: the flagellum of Trypanosoma brucei

PubMed Central

Langousis, Gerasimos; Hill, Kent L.

2014-01-01

A central feature of trypanosome cell biology and life cycle is the parasite’s single flagellum, which is an essential and multifunctional organelle involved in cell propulsion, morphogenesis and cytokinesis. The flagellar membrane is also a specialized subdomain of the cell surface that harbors multiple parasite virulence factors with roles in signaling and host-parasite interactions. In this review, we discuss the structure, assembly and function of the trypanosome flagellum, including canonical roles in cell motility as well as novel and emerging roles in cell morphogenesis and host-parasite interaction. PMID:24931043

Peroxisome Proliferator-Activated Receptor-γ Ligands Alter Breast Cancer Cell Motility through Modulation of the Plasminogen Activator System

PubMed Central

Carter, Jennifer C.; Church, Frank C.

2011-01-01

We investigated peroxisome proliferator-activated receptor-γ (PPAR-γ) ligands effect on cell motility and the plasminogen activator system using normal MCF-10A and malignant MCF-10CA1 cell lines. Ciglitazone reduced both wound-induced migration and chemotaxis. However, the effect was not reversed with pretreatment of cells with the PPAR-γ-specific antagonist GW9662. Immunoblot analysis of conditioned media showed ciglitazone decreased plasminogen activator inhibitor-1 (PAI-1) in both cell lines; this effect was also unaltered by PPAR-γ antagonism. Alternatively, treatment with the ω-6 fatty acid arachidonic acid (ArA), but not the ω-3 fatty acid docosahexanoic acid, increased both MCF-10A cell migration and cell surface uPA activity. Pretreatment with a PPAR-γ antagonist reversed these effects, suggesting that ArA mediates its effect on cell motility and uPA activity through PPAR-γ activation. Collectively, the data suggest PPAR-γ ligands have a differential effect on normal and malignant cell migration and the plasminogen activation system, resulting from PPAR-γ-dependent and PPAR-γ-independent effects. PMID:22131991

A computational model of amoeboid cell swimming in unbounded medium and through obstacles

NASA Astrophysics Data System (ADS)

Campbell, Eric; Bagchi, Prosenjit

2017-11-01

Pseudopod-driven motility is commonly observed in eukaryotic cells. Pseudopodia are actin-rich protrusions of the cellular membrane which extend, bifurcate, and retract in cycles resulting in amoeboid locomotion. While actin-myosin interactions are responsible for pseudopod generation, cell deformability is crucial concerning pseudopod dynamics. Because pseudopodia are highly dynamic, cells are capable of deforming into complex shapes over time. Pseudopod-driven motility represents a multiscale and complex process, coupling cell deformation, protein biochemistry, and cytoplasmic and extracellular fluid motion. In this work, we present a 3D computational model of amoeboid cell swimming in an extracellular medium (ECM). The ECM is represented as a fluid medium with or without obstacles. The model integrates full cell deformation, a coarse-grain reaction-diffusion system for protein dynamics, and fluid interaction. Our model generates pseudopodia which bifurcate and retract, showing remarkable similarity to experimental observations. Influence of cell deformation, protein diffusivity and cytoplasmic viscosity on the swimming speed is analyzed in terms of altered pseudopod dynamics. Insights into the role of matrix porosity and obstacle size on cell motility are also provided. Funded by NSF CBET 1438255.

Physical Model of the Dynamic Instability in an Expanding Cell Culture

PubMed Central

Mark, Shirley; Shlomovitz, Roie; Gov, Nir S.; Poujade, Mathieu; Grasland-Mongrain, Erwan; Silberzan, Pascal

2010-01-01

Abstract Collective cell migration is of great significance in many biological processes. The goal of this work is to give a physical model for the dynamics of cell migration during the wound healing response. Experiments demonstrate that an initially uniform cell-culture monolayer expands in a nonuniform manner, developing fingerlike shapes. These fingerlike shapes of the cell culture front are composed of columns of cells that move collectively. We propose a physical model to explain this phenomenon, based on the notion of dynamic instability. In this model, we treat the first layers of cells at the front of the moving cell culture as a continuous one-dimensional membrane (contour), with the usual elasticity of a membrane: curvature and surface-tension. This membrane is active, due to the forces of cellular motility of the cells, and we propose that this motility is related to the local curvature of the culture interface; larger convex curvature correlates with a stronger cellular motility force. This shape-force relation gives rise to a dynamic instability, which we then compare to the patterns observed in the wound healing experiments. PMID:20141748

Motile activities of Dictyostelium discoideum differ from those in Protista or vertebrate animal cells.

PubMed

Waligórska, Agnieszka; Wianecka-Skoczeń, Magdalena; Korohoda, Włodzimierz

2007-01-01

Cell movement in the amoebae Dictyostelium discoideum has been examined in media differing in monovalent cation concentration (i.e. Na+ and K+). Under isotonic or even slightly hypertonic conditions, the cells move equally well in solutions in which either potassium or sodium ions dominate. However, in strongly hypertonic solutions the amoebae showed motility in a 2% potassium chloride solution, but remained motionless in a hypertonic 2% sodium chloride solution. This inhibition of D. discoideum amoebae movement in a hypertonic sodium chloride solution was fully reversible. Such behaviour corresponds to that of plant, fungi, and some invertebrate animal cells rather than protozoan or vertebrate cells. These observations suggest that studies using D. discoideum as a model for cell motility in vertebrate animal tissue cells should be considered with caution, and would seem to confirm the classification of cellular slime moulds as related rather to Fungi than to Protista. This also shows that the cell membrane models should consider the asymmetry in sodium/potassium ion concentrations found in vertebrate animal cells as one of various possibilities.

miR-22 suppresses the proliferation and invasion of gastric cancer cells by inhibiting CD151

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Xun; Yu, Honggang, E-mail: honggang_yuwh@163.com; Lu, Xinyao

2014-02-28

Highlights: • miR-22 was decreased in GC tissue samples and cell lines. • miR-22 suppressed GC cell growth and motility in vitro. • CD151 was a direct target of miR-22. • miR-22 suppressed GC cell growth and motility by inhibiting CD151. - Abstract: Gastric cancer (GC) is the second common cause of cancer-related death worldwide. microRNAs (miRNAs) play important roles in the carcinogenesis of GC. Here, we found that miR-22 was significantly decreased in GC tissue samples and cell lines. Ectopic overexpression of miR-22 remarkably suppressed cell proliferation and colony formation of GC cells. Moreover, overexpression of miR-22 significantly suppressedmore » migration and invasion of GC cells. CD151 was found to be a target of miR-22. Furthermore, overexpression of CD151 significantly attenuated the tumor suppressive effect of miR-22. Taken together, miR-22 might suppress GC cells growth and motility partially by inhibiting CD151.« less

Mycoplasma pneumoniae Protein P30 Is Required for Cytadherence and Associated with Proper Cell Development

PubMed Central

Romero-Arroyo, Cynthia E.; Jordan, Jarrat; Peacock, Susan J.; Willby, Melisa J.; Farmer, Mark A.; Krause, Duncan C.

1999-01-01

The attachment organelle of Mycoplasma pneumoniae is a polar, tapered cell extension containing an intracytoplasmic, electron-dense core. This terminal structure is the leading end in gliding motility, and its duplication is thought to precede cell division, raising the possibility that mutations affecting cytadherence also confer a defect in motility or cell development. Mycoplasma surface protein P30 is associated with the attachment organelle, and P30 mutants II-3 and II-7 do not cytadhere. In this study, the recombinant wild-type but not the mutant II-3 p30 allele restored cytadherence when transformed into P30 mutants by recombinant transposon delivery. The mutations associated with loss of P30 in mutant II-3 and reacquisition of P30 in cytadhering revertants thereof were identified by nucleotide sequencing of the p30 gene. Morphological abnormalities that included ovoid or multilobed cells having a poorly defined tip structure were associated with loss of P30. Digital image analysis confirmed quantitatively the morphological differences noted visually. Transformation of the P30 mutants with the wild-type p30 allele restored a normal morphology, as determined both visually and by digital image analysis, suggesting that P30 plays a role in mycoplasma cell development. Finally, the P30 mutants localized the adhesin protein P1 to the terminal organelle, indicating that P30 is not involved in P1 trafficking but may be required for its receptor-binding function. PMID:9973332

Transposon tagging of genes for cell-cell interactions in Myxococcus xanthus.

PubMed Central

Kalos, M; Zissler, J

1990-01-01

The prokaryote Myxococcus xanthus is a model for cell interactions important in multicellular behavior. We used the transposon TnphoA to specifically identify genes for cell-surface factors involved in cell interactions. From a library of 10,700 insertions of TnphoA, we isolated 36 that produced alkaline phosphatase activity. Three TnphoA insertions tagged cell motility genes, called cgl, which control the adventurous movement of cells. The products of the tagged cgl genes could function in trans upon other cells and were localized primarily in the cell envelope and extracellular space, consistent with TnphoA tagging genes for extracellular factors controlling motility. Images PMID:2172982

The role of drebrin in glioma migration and invasion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Terakawa, Yuzo; Department of Neurosurgery, Osaka City University Graduate School of Medicine, Osaka; Agnihotri, Sameer

Glioblastoma (GBM) is the most common primary brain tumor in adults. Despite current advances in therapy consisting of surgery followed by chemotherapy and radiation, the overall survival rate still remains poor. Therapeutic failures are partly attributable to the highly infiltrative nature of tumor adjacent to normal brain parenchyma. Recently, evidence is mounting to suggest that actin cytoskeleton dynamics are critical components of the cell invasion process. Drebrin is an actin-binding protein involved in the regulation of actin filament organization, and plays a significant role in cell motility; however, the role of drebrin in glioma cell invasiveness has not yet beenmore » fully elucidated. Therefore, this study was aimed to clarify the role of drebrin in glioma cell morphology and cell motility. Here we show that drebrin is expressed in glioma cell lines and in operative specimens of GBM. We demonstrate that stable overexpression of drebrin in U87 cells leads to alterations in cell morphology, and induces increased invasiveness in vitro while knockdown of drebrin in U87 cells by small interfering RNA (siRNA) decreases invasion and migration. In addition, we show that depletion of drebrin by siRNA alters glioma cell morphology in A172 GBM cell line. Our results suggest that drebrin contributes to the maintenance of cell shape, and may play an important role in glioma cell motility. - Highlights: ► Drebrin is an actin-binding protein aberrantly expressed in several cancers. ► Role of drebrin in glioma cell morphology and motility is previously unknown. ► We demonstrate that drebrin is expressed in 40% of glioblastoma specimens. ► Drebrin plays a significant role in modulating glioma cell migration and invasion.« less

T Cell Interstitial Migration: Motility Cues from the Inflamed Tissue for Micro- and Macro-Positioning.

PubMed

Gaylo, Alison; Schrock, Dillon C; Fernandes, Ninoshka R J; Fowell, Deborah J

2016-01-01

Effector T cells exit the inflamed vasculature into an environment shaped by tissue-specific structural configurations and inflammation-imposed extrinsic modifications. Once within interstitial spaces of non-lymphoid tissues, T cells migrate in an apparent random, non-directional, fashion. Efficient T cell scanning of the tissue environment is essential for successful location of infected target cells or encounter with antigen-presenting cells that activate the T cell's antimicrobial effector functions. The mechanisms of interstitial T cell motility and the environmental cues that may promote or hinder efficient tissue scanning are poorly understood. The extracellular matrix (ECM) appears to play an important scaffolding role in guidance of T cell migration and likely provides a platform for the display of chemotactic factors that may help to direct the positioning of T cells. Here, we discuss how intravital imaging has provided insight into the motility patterns and cellular machinery that facilitates T cell interstitial migration and the critical environmental factors that may optimize the efficiency of effector T cell scanning of the inflamed tissue. Specifically, we highlight the local micro-positioning cues T cells encounter as they migrate within inflamed tissues, from surrounding ECM and signaling molecules, as well as a requirement for appropriate long-range macro-positioning within distinct tissue compartments or at discrete foci of infection or tissue damage. The central nervous system (CNS) responds to injury and infection by extensively remodeling the ECM and with the de novo generation of a fibroblastic reticular network that likely influences T cell motility. We examine how inflammation-induced changes to the CNS landscape may regulate T cell tissue exploration and modulate function.

Activation of Transforming Growth Factor Beta 1 Signaling in Gastric Cancer-associated Fibroblasts Increases Their Motility, via Expression of Rhomboid 5 Homolog 2, and Ability to Induce Invasiveness of Gastric Cancer Cells.

PubMed

Ishimoto, Takatsugu; Miyake, Keisuke; Nandi, Tannistha; Yashiro, Masakazu; Onishi, Nobuyuki; Huang, Kie Kyon; Lin, Suling Joyce; Kalpana, Ramnarayanan; Tay, Su Ting; Suzuki, Yuka; Cho, Byoung Chul; Kuroda, Daisuke; Arima, Kota; Izumi, Daisuke; Iwatsuki, Masaaki; Baba, Yoshifumi; Oki, Eiji; Watanabe, Masayuki; Saya, Hideyuki; Hirakawa, Kosei; Baba, Hideo; Tan, Patrick

2017-07-01

Fibroblasts that interact with cancer cells are called cancer-associated fibroblasts (CAFs), which promote progression of different tumor types. We investigated the characteristics and functions of CAFs in diffuse-type gastric cancers (DGCs) by analyzing features of their genome and gene expression patterns. We isolated CAFs and adjacent non-cancer fibroblasts (NFs) from 110 gastric cancer (GC) tissues from patients who underwent gastrectomy in Japan from 2008 through 2016. Cells were identified using specific markers of various cell types by immunoblot and flow cytometry. We selected pairs of CAFs and NFs for whole-exome and RNA sequencing analyses, and compared expression of specific genes using quantitative reverse transcription PCR. Protein levels and phosphorylation were compared by immunoblot and immunofluorescence analyses. Rhomboid 5 homolog 2 (RHBDF2) was overexpressed from a transgene in fibroblasts or knocked down using small interfering RNAs. Motility and invasiveness of isolated fibroblasts and GC cell lines (AGS, KATOIII, MKN45, NUGC3, NUGC4, OCUM-2MD3 and OCUM-12 cell lines) were quantified by real-time imaging analyses. We analyzed 7 independent sets of DNA microarray data from patients with GC and associated expression levels of specific genes with patient survival times. Nude mice were given injections of OCUM-2MD3 in the stomach wall; tumors and metastases were collected and analyzed by immunohistochemistry. Many of the genes with increased expression in CAFs compared with NFs were associated with transforming growth factor beta 1 (TGFB1) activity. When CAFs were cultured in extracellular matrix, they became more motile than NFs; DGC cells incubated with CAFs were also more motile and invasive in vitro than DGC cells not incubated with CAFs. When injected into nude mice, CAF-incubated DGC cells invaded a greater number of lymphatic vessels than NF-incubated DGC cells. We identified RHBDF2 as a gene overexpressed in CAFs compared with NFs. Knockdown of RHBDF2 in CAFs reduced their elongation and motility in response to TGFB1, whereas overexpression of RHBDF2 in NFs increased their motility in extracellular matrix. RHBDF2 appeared to regulate oncogenic and non-canonical TGFB1 signaling. Knockdown of RHBDF2 in CAFs reduced cleavage of the TGFB receptor 1 (TGFBR1) by ADAM metallopeptidase domain 17 (ADAM17 or TACE) and reduced expression of genes that regulate motility. Incubation of NFs with in interleukin 1 alpha (IL1A), IL1B or tumor necrosis factor, secreted by DGCs, increased fibroblast expression of RHBDF2. Simultaneous high expression of these cytokines in GC samples was associated with shorter survival times of patients. In CAFs isolated from human DGCs, we observed increased expression of RHBDF2, which regulates TGFB1 signaling. Expression of RHBDF2 in fibroblasts is induced by inflammatory cytokines (such as IL1A, IL1B, and tumor necrosis factor) secreted by DGCs. RHBDF2 promotes cleavage of TGFBR1 by activating TACE and motility of CAFs in response to TGFB1. These highly motile CAFs induce DGCs to invade extracellular matrix and lymphatic vessels in nude mice. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Interplay of differential cell mechanical properties, motility, and proliferation in emergent collective behavior of cell co-cultures

NASA Astrophysics Data System (ADS)

Sutter, Leo; Kolbman, Dan; Wu, Mingming; Ma, Minglin; Das, Moumita

The biophysics of cell co-cultures, i.e. binary systems of cell populations, is of great interest in many biological processes including formation of embryos, and tumor progression. During these processes, different types of cells with different physical properties are mixed with each other, with important consequences for cell-cell interaction, aggregation, and migration. The role of the differences in their physical properties in their collective behavior remains poorly understood. Furthermore, until recently most theoretical studies of collective cell migration have focused on two dimensional systems. Under physiological conditions, however, cells often have to navigate three dimensional and confined micro-environments. We study a confined, three-dimensional binary system of interacting, active, and deformable particles with different physical properties such as deformability, motility, adhesion, and division rates using Langevin Dynamics simulations. Our findings may provide insights into how the differences in and interplay between cell mechanical properties, division, and motility influence emergent collective behavior such as cell aggregation and segregation experimentally observed in co-cultures of breast cancer cells and healthy breast epithelial cells. This work was partially supported by a Cottrell College Science Award.

Polarization of cells and soft objects driven by mechanical interactions: consequences for migration and chemotaxis.

PubMed

Leoni, M; Sens, P

2015-02-01

We study a generic model for the polarization and motility of self-propelled soft objects, biological cells, or biomimetic systems, interacting with a viscous substrate. The active forces generated by the cell on the substrate are modeled by means of oscillating force multipoles at the cell-substrate interface. Symmetry breaking and cell polarization for a range of cell sizes naturally "emerge" from long range mechanical interactions between oscillating units, mediated both by the intracellular medium and the substrate. However, the harnessing of cell polarization for motility requires substrate-mediated interactions. Motility can be optimized by adapting the oscillation frequency to the relaxation time of the system or when the substrate and cell viscosities match. Cellular noise can destroy mechanical coordination between force-generating elements within the cell, resulting in sudden changes of polarization. The persistence of the cell's motion is found to depend on the cell size and the substrate viscosity. Within such a model, chemotactic guidance of cell motion is obtained by directionally modulating the persistence of motion, rather than by modulating the instantaneous cell velocity, in a way that resembles the run and tumble chemotaxis of bacteria.

Differential effects of immunosuppressive drugs on T-cell motility.

PubMed

Datta, A; David, R; Glennie, S; Scott, D; Cernuda-Morollon, E; Lechler, R I; Ridley, A J; Marelli-Berg, F M

2006-12-01

The best-characterized mechanism of the action of immunosuppressive drugs is to prevent T-cell clonal expansion, thus containing the magnitude of the ensuing immune response. As T-cell recruitment to the inflammatory site is another key step in the development of T-cell-mediated inflammation, we analyzed and compared the effects of two commonly used immunosuppressants, cyclosporin A (CsA) and the rapamycin-related compound SDZ-RAD, on the motility of human CD4+ T cells. We show that CsA, but not SDZ-RAD, inhibits T-cell transendothelial migration in vitro. CsA selectively impaired chemokine-induced T-cell chemotaxis while integrin-mediated migration was unaffected. The inhibition of T-cell chemotaxis correlated with reduced AKT/PKB but not ERK activation following exposure to the chemokine CXCL-12/SDF-1. In addition, CsA, but not SDZ-RAD, prevents some T-cell receptor-mediated effects on T-cell motility. Finally, we show that CsA, but not SDZ-RAD inhibits tissue infiltration by T cells in vivo. Our data suggest a prominent antiinflammatory role for CsA in T-cell-mediated tissue damage, by inhibiting T-cell trafficking into tissues in addition to containing clonal expansion.

Effect of post-thaw addition of seminal plasma on motility, viability and chromatin integrity of cryopreserved donkey jack (Equus asinus) spermatozoa.

PubMed

Sabatini, C; Mari, G; Mislei, B; Love, Cc; Panzani, D; Camillo, F; Rota, A

2014-12-01

Pregnancy rates in donkeys after artificial insemination with cryopreserved semen are still low, compared to the horse species. Addition of autologous seminal plasma to frozen-thawed semen appeared to improve pregnancy rates. The aims of this study were to evaluate (1) sperm motility and plasma membrane integrity after thawing (T0) and after one and 2 h (T1 and T2) of post-thaw incubation in either 0% (SP0) or 70% (SP70) autologous seminal plasma and (2) sperm motility, plasma membrane integrity and DNA quality (%COMP-αt) after thawing (T0) and after 2 and 4 h (T2 and T4) of post-thaw incubation in either 0% (SP0), 5% (SP5) or 20% (SP20) homologous seminal plasma. In experiment 1, seminal plasma decreased total and progressive sperm motility and plasma membrane intact spermatozoa immediately after dilution and at all following time points (p < 0.05). In experiment 2, total and progressive motility did not differ between treatments immediately after dilution and between SP0 and SP5 at T2, while they were lower in both SP5 and SP20 than in SP0 at T4. Plasma membrane intact sperm cells did not differ between SP0 and SP5 and were lower in SP20 at all time points. DNA quality was not affected by treatment immediately after dilution and was significantly worse for SP20 after 4 h of incubation (p < 0.05). The post-thaw addition of seminal plasma at the tested concentrations did not improve donkey frozen semen characteristics in vitro over time. © 2014 Blackwell Verlag GmbH.

Computer-assisted sperm analysis of fresh epididymal cat spermatozoa and the impact of cool storage (4 degrees C) on sperm quality.

PubMed

Filliers, M; Rijsselaere, T; Bossaert, P; De Causmaecker, V; Dewulf, J; Pope, C E; Van Soom, A

2008-12-01

Epididymal cat sperm is commonly used for in vitro fertilization. Because of the high variability in preparation protocols and methods of evaluation, sperm quality may vary considerably between experiments and laboratories. The aims of the present study were (1) to describe an epididymal sperm preparation protocol to produce clean, highly motile samples using density gradient centrifugation, (2) to provide reference values of computer-assisted semen analysis (CASA) parameters of fresh epididymal cat sperm after density gradient centrifugation and (3) to investigate the effect of cool storage on various spermatozoa characteristics. After slicing the epididymides, viable and motile sperm cells were isolated using Percoll centrifugation. Sperm motility parameters were subsequently assessed using CASA in experiment 1. In experiment 2, fresh (day 0) sperm samples were evaluated for motility parameters (HTR) and stained for assessment of acrosomal status (FITC-PSA), morphology (eosin/nigrosin (E/N)), membrane integrity (E/N and SYBR((R))14-PI) and DNA fragmentation (TUNEL). After addition of a Tris-glucose-citrate diluent containing 20% egg yolk, samples were cooled to 4 degrees C and reassessed on d1, d3, d5, d7 and d10. Cool storage impaired most motility and velocity parameters: MOT, PMOT, VAP, VSL, VCL, BCF, RAPID and the percentage of normal spermatozoa showed a decrease over time (P<0.05) as compared to fresh samples. In contrast, STR, ALH, membrane integrity, DNA fragmentation and the percentage of acrosome intact spermatozoa were not affected by cool storage. However, the influence of cool storage of cat spermatozoa on subsequent in vitro embryo development and quality after IVF requires further investigation.

Phenazines affect biofilm formation by Pseudomonas aeruginosa in similar ways at various scales

PubMed Central

Ramos, Itzel; Dietrich, Lars E. P.; Price-Whelan, Alexa; Newman, Dianne K.

2010-01-01

Pseudomonads produce phenazines, a group of small, redox-active compounds with diverse physiological functions. In this study, we compared the phenotypes of Pseudomonas aeruginosa strain PA14 and a mutant unable to synthesize phenazines in flow cell and colony biofilms quantitatively. Although phenazine production does not impact the ability of PA14 to attach to surfaces, as has been shown for Pseudomonas chlororaphis (Maddula, 2006; Maddula, 2008), it influences swarming motility and the surface-to-volume ratio of mature biofilms. These results indicate that phenazines affect biofilm development across a large range of scales, but in unique ways for different Pseudomonas species. PMID:20123017

Incubation of human sperm with micelles made from glycerophospholipid mixtures increases sperm motility and resistance to oxidative stress

PubMed Central

Costa, Carlos; Bassaizteguy, Verónica; Cardozo, Romina; Montes, José; Settineri, Robert; Nicolson, Garth L.

2018-01-01

Membrane integrity is essential in maintaining sperm viability, signaling, and motility, which are essential for fertilization. Sperm are highly susceptible to oxidative stress, as they are rich in sensitive polyunsaturated fatty acids (PUFA), and are unable to synthesize and repair many essential membrane constituents. Because of this, sperm cellular membranes are important targets of this process. Membrane Lipid Replacement (MLR) with glycerophospholipid mixtures (GPL) has been shown to ameliorate oxidative stress in cells, restore their cellular membranes, and prevent loss of function. Therefore, we tested the effects of MLR on sperm by tracking and monitoring GPL incorporation into their membrane systems and studying their effects on sperm motility and viability under different experimental conditions. Incubation of sperm with mixtures of exogenous, unoxidized GPL results in their incorporation into sperm membranes, as shown by the use of fluorescent dyes attached to GPL. The percent overall (total) sperm motility was increased from 52±2.5% to 68±1.34% after adding GPL to the incubation media, and overall sperm motility was recovered from 7±2% after H2O2 treatment to 58±2.5%)(n = 8, p<0.01) by the incorporation of GPL into sperm membranes. When sperm were exposed to H2O2, the mitochondrial inner membrane potential (MIMP), monitored using the MIMP tracker dye JC-1 in flow cytometry, diminished, whereas the addition of GPL prevented the decrease in MIMP. Confocal microscopy with Rhodamine-123 and JC-1 confirmed the mitochondrial localization of the dyes. We conclude that incubation of human sperm with glycerolphospholipids into the membranes of sperm improves sperm viability, motility, and resistance to oxidizing agents like H2O2. This suggests that human sperm might be useful to test innovative new treatments like MLR, since such treatments could improve fertility when it is adversely affected by increased oxidative stress. PMID:29856778

TMEM16A inhibition impedes capacitation and acquisition of hyperactivated motility in guinea pig sperm.

PubMed

Cordero-Martínez, Joaquín; Reyes-Miguel, Tania; Rodríguez-Páez, Lorena; Garduño-Siciliano, Leticia; Maldonado-García, Deneb; Roa-Espitia, Ana L; Hernández-González, Enrique O

2018-07-01

Ca 2+ -activated Cl - channels (CaCCs) are anionic channels that regulate many important physiological functions associated with chloride and calcium flux in some somatic cells. The molecular identity of CaCCs was revealed to be TMEM16A and TMEM16B (also known as Anoctamin or ANO1 and ANO2, respectively) in all eukaryotes. A recent study suggests the presence of TMEM16A in human sperm and a relationship with the rhZP-induced acrosome reaction. However, to the best of our knowledge, little is known about the role of TMEM16A in other spermatic processes such as capacitation or motility. In this study, we evaluated the effects of two TMEM16A antagonists on capacitation, acrosome reaction, and motility in guinea pig sperm; these antagonists were T16Ainh-A01, belonging to a second generation of potent antagonists of TMEM16A, and niflumic acid (NFA), a well-known antagonist of TMEM16A (CaCCs). First of all, we confirmed that the absence of Cl - in the capacitation medium changes motility parameters, capacitation, and the progesterone-induced acrosome reaction. Using a specific antibody, TMEM16A was found as a protein band of ∼120 kDa, which localization was in the apical crest of the acrosome and the middle piece of the flagellum. Inhibition of TMEM16A by T16Ainh-A01 affected sperm physiology by reducing capacitation, blocking the progesterone-induced acrosome reaction under optimal capacitation conditions, inhibiting progressive motility, and the acquisition of hyperactivated motility, diminishing [Ca 2+ ]i, and increasing [Cl - ]i. These changes in sperm kinematic parameters provide new evidence of the important role played by TMEM16A in the production of sperm capable of fertilizing oocytes. © 2018 Wiley Periodicals, Inc.

Engineering bacterial motility towards hydrogen-peroxide.

PubMed

Virgile, Chelsea; Hauk, Pricila; Wu, Hsuan-Chen; Shang, Wu; Tsao, Chen-Yu; Payne, Gregory F; Bentley, William E

2018-01-01

Synthetic biologists construct innovative genetic/biological systems to treat environmental, energy, and health problems. Many systems employ rewired cells for non-native product synthesis, while a few have employed the rewired cells as 'smart' devices with programmable function. Building on the latter, we developed a genetic construct to control and direct bacterial motility towards hydrogen peroxide, one of the body's immune response signaling molecules. A motivation for this work is the creation of cells that can target and autonomously treat disease, the latter signaled by hydrogen peroxide release. Bacteria naturally move towards a variety of molecular cues (e.g., nutrients) in the process of chemotaxis. In this work, we engineered bacteria to recognize and move towards hydrogen peroxide, a non-native chemoattractant and potential toxin. Our system exploits oxyRS, the native oxidative stress regulon of E. coli. We first demonstrated H2O2-mediated upregulation motility regulator, CheZ. Using transwell assays, we showed a two-fold increase in net motility towards H2O2. Then, using a 2D cell tracking system, we quantified bacterial motility descriptors including velocity, % running (of tumble/run motions), and a dynamic net directionality towards the molecular cue. In CheZ mutants, we found that increased H2O2 concentration (0-200 μM) and induction time resulted in increased running speeds, ultimately reaching the native E. coli wild-type speed of ~22 μm/s with a ~45-65% ratio of running to tumbling. Finally, using a microfluidic device with stable H2O2 gradients, we characterized responses and the potential for "programmed" directionality towards H2O2 in quiescent fluids. Overall, the synthetic biology framework and tracking analysis in this work will provide a framework for investigating controlled motility of E. coli and other 'smart' probiotics for signal-directed treatment.

Transplantation of enteric nervous system stem cells rescues nitric oxide synthase deficient mouse colon

PubMed Central

McCann, Conor J.; Cooper, Julie E.; Natarajan, Dipa; Jevans, Benjamin; Burnett, Laura E.; Burns, Alan J.; Thapar, Nikhil

2017-01-01

Enteric nervous system neuropathy causes a wide range of severe gut motility disorders. Cell replacement of lost neurons using enteric neural stem cells (ENSC) is a possible therapy for these life-limiting disorders. Here we show rescue of gut motility after ENSC transplantation in a mouse model of human enteric neuropathy, the neuronal nitric oxide synthase (nNOS−/−) deficient mouse model, which displays slow transit in the colon. We further show that transplantation of ENSC into the colon rescues impaired colonic motility with formation of extensive networks of transplanted cells, including the development of nNOS+ neurons and subsequent restoration of nitrergic responses. Moreover, post-transplantation non-cell-autonomous mechanisms restore the numbers of interstitial cells of Cajal that are reduced in the nNOS−/− colon. These results provide the first direct evidence that ENSC transplantation can modulate the enteric neuromuscular syncytium to restore function, at the organ level, in a dysmotile gastrointestinal disease model. PMID:28671186

Genistein inhibits cell invasion and motility by inducing cell differentiation in murine osteosarcoma cell line LM8

PubMed Central

2012-01-01

Background One of the problems associated with osteosarcoma is the frequent formation of micrometastases in the lung prior to diagnosis because the development of metastatic lesions often causes a fatal outcome. Therefore, the prevention of pulmonary metastases during the early stage of tumor development is critical for the improvement of the prognosis of osteosarcoma patients. In Japan, soy is consumed in a wide variety of forms, such as miso soup and soy sauce. The purpose of this study is to investigate the effect of genistein, an isoflavone found in soy, on the invasive and motile potential of osteosarcoma cells. Methods LM8 cells were treated for 3 days with various concentrations of genistein. The effect of genistein on cell proliferation was determined by DNA measurement in the cultures and 5-bromo-2’-deoxyuridine (BrdU) incorporation study. The assays of cell invasion and motility were performed using the cell culture inserts with either matrigel-coated membranes or uncoated membranes in the invasion chambers. The expression and secretion of MMP-2 were determined by immunohistochemistry and gelatin zymography. The subcellular localization and cellular level of β-catenin were determined by immunofluorescence and Western blot. For examining cell morphology, the ethanol-fixed cells were stained with hematoxylin-eosin (H&E). The expression of osteocalcin mRNA was determined by reverse transcription-polymerase chain reaction (RT-PCR). Results Genistein dose-dependently inhibits cell proliferation. Genistein-treated cells were less invasive and less motile than untreated cells. The expression and secretion of MMP-2 were lower in the genistein-treated cultures than in the untreated cultures. β-Catenin in untreated cells was located in the cytoplasm and/or nucleus, while in genistein-treated cells it was translocated near to the plasma membrane. The level of β-catenin was higher in genistein-treated cells than in untreated cells. Treatment of LM8 cells with genistein induced morphological changes, markedly decreased the formation of multilayer masses of cells, and markedly increased the expression of osteocalcin mRNA. Conclusions Genistein decreased invasive and motile potential by inducing cell differentiation in LM8 cells. Genistein may be useful as an anti-metastatic drug for osteosarcoma through its differentiation-inducing effects. PMID:23013480

Prestin-based outer hair cell electromotility in knockin mice does not appear to adjust the operating point of a cilia-based amplifier

PubMed Central

Gao, Jiangang; Wang, Xiang; Wu, Xudong; Aguinaga, Sal; Huynh, Kristin; Jia, Shuping; Matsuda, Keiji; Patel, Manish; Zheng, Jing; Cheatham, MaryAnn; He, David Z.; Dallos, Peter; Zuo, Jian

2007-01-01

The remarkable sensitivity and frequency selectivity of the mammalian cochlea is attributed to a unique amplification process that resides in outer hair cells (OHCs). Although the mammalian-specific somatic motility is considered a substrate of cochlear amplification, it has also been proposed that somatic motility in mammals simply acts as an operating-point adjustment for the ubiquitous stereocilia-based amplifier. To address this issue, we created a mouse model in which a mutation (C1) was introduced into the OHC motor protein prestin, based on previous results in transfected cells. In C1/C1 knockin mice, localization of C1-prestin, as well as the length and number of OHCs, were all normal. In OHCs isolated from C1/C1 mice, nonlinear capacitance and somatic motility were both shifted toward hyperpolarization, so that, compared with WT controls, the amplitude of cycle-by-cycle (alternating, or AC) somatic motility remained the same, but the unidirectional (DC) component reversed polarity near the OHC's presumed in vivo resting membrane potential. No physiological defects in cochlear sensitivity or frequency selectivity were detected in C1/C1 or C1/+ mice. Hence, our results do not support the idea that OHC somatic motility adjusts the operating point of a stereocilia-based amplifier. However, they are consistent with the notion that the AC component of OHC somatic motility plays a dominant role in mammalian cochlear amplification. PMID:17640919

Gut microbial products regulate murine gastrointestinal motility via Toll-like Receptor 4 signaling

PubMed Central

Anitha, Mallappa; Vijay-Kumar, Matam; Sitaraman, Shanthi V.; Gewirtz, Andrew T.; Srinivasan, Shanthi

2012-01-01

Background & Aims Altered gastrointestinal motility is associated with significant morbidity and health care costs. Toll-like receptors regulate intestinal homeostasis. We examined the roles of Toll-like receptor (TLR)4 signaling in survival of enteric neurons and gastrointestinal motility. Methods We assessed changes in intestinal motility by assessing stool frequency, bead expulsion, and isometric muscle recordings of colonic longitudinal muscle strips from mice that do not express TLR4 (Tlr4Lps-d or TLR4−/−) or Myd88 (Myd88−/−), in wild-type germ-free mice or wild-type mice depleted of the microbiota, and in mice with neural crest-specific deletion of Myd88 (Wnt1Cre+/−/Myd88fl/fl). We studied the effects of the TLR4 agonist lipopolysaccharide (LPS) on survival of cultured, immortalized fetal enteric neurons (IM-FEN) and enteric neuronal cells isolated from wild-type and Tlr4Lps-d mice at embryonic day 13.5. Results There was a significant delay in gastrointestinal motility and reduced numbers of nitrergic neurons in TLR4Lps-d, TLR4−/−, and Myd88−/− mice, compared with wild-type mice. A similar phenotype was observed in germ-free mice, mice depleted of intestinal microbiota, and Wnt1Cre+/−/Myd88fl/fl mice. Incubation of enteric neuronal cells with LPS led to activation of the transcription factor NF-κB and increased cell survival. Conclusions Interactions between enteric neurons and microbes increases neuron survival and gastrointestinal motility in mice. LPS activation of TLR4 and NF-κB appears to promote survival of enteric neurons. Factors that regulate TLR4 signaling by neurons might be developed to alter gastrointestinal motility. PMID:22732731

Rosiglitazone Improves Stallion Sperm Motility, ATP Content, and Mitochondrial Function.

PubMed

Swegen, Aleona; Lambourne, Sarah Renay; Aitken, R John; Gibb, Zamira

2016-11-01

Media used for equine sperm storage often contain relatively high concentrations of glucose, even though stallion spermatozoa preferentially utilize oxidative phosphorylation (OXPHOS) over glycolysis to generate ATP and support motility. Rosiglitazone is an antidiabetic compound that enhances metabolic flexibility and glucose utilization in various cell types, but its effects on sperm metabolism are unknown. This study investigated the effects of rosiglitazone on stallion sperm function in vitro, along with the possible role of AMP-activated protein kinase (AMPK) in mediating these effects. Spermatozoa were incubated with or without rosiglitazone, GW9662 (an antagonist of peroxisome proliferator-activating receptor-gamma), and compound C (CC; an AMPK inhibitor). Sperm motility, viability, reactive oxygen species production, mitochondrial membrane potential (mMP), ATP content, and glucose uptake capacity were measured. Samples incubated with rosiglitazone displayed significantly higher motility, percentage of cells with normal mMP, ATP content, and glucose uptake capacity, while sperm viability was unaffected. The percentage of spermatozoa positive for mitochondrial ROS was also significantly lower in rosiglitazone-treated samples. AMPK localized to the sperm midpiece, and its phosphorylation, was increased in rosiglitazone-treated spermatozoa. CC decreased sperm AMPK phosphorylation and reduced sperm motility, and successfully inhibited the effects of rosiglitazone. Inclusion of rosiglitazone in a room temperature sperm storage medium maintained sperm motility above 60% for 6 days, attaining significantly higher motility than sperm stored in control media. The ability of rosiglitazone to substantially alleviate the time-dependent deterioration of stallion spermatozoa by diverting metabolism away from OXPHOS and toward glycolysis has novel implications for the long-term, functional preservation of these cells. © 2016 by the Society for the Study of Reproduction, Inc.

T Cell Interstitial Migration: Motility Cues from the Inflamed Tissue for Micro- and Macro-Positioning

PubMed Central

Gaylo, Alison; Schrock, Dillon C.; Fernandes, Ninoshka R. J.; Fowell, Deborah J.

2016-01-01

Effector T cells exit the inflamed vasculature into an environment shaped by tissue-specific structural configurations and inflammation-imposed extrinsic modifications. Once within interstitial spaces of non-lymphoid tissues, T cells migrate in an apparent random, non-directional, fashion. Efficient T cell scanning of the tissue environment is essential for successful location of infected target cells or encounter with antigen-presenting cells that activate the T cell’s antimicrobial effector functions. The mechanisms of interstitial T cell motility and the environmental cues that may promote or hinder efficient tissue scanning are poorly understood. The extracellular matrix (ECM) appears to play an important scaffolding role in guidance of T cell migration and likely provides a platform for the display of chemotactic factors that may help to direct the positioning of T cells. Here, we discuss how intravital imaging has provided insight into the motility patterns and cellular machinery that facilitates T cell interstitial migration and the critical environmental factors that may optimize the efficiency of effector T cell scanning of the inflamed tissue. Specifically, we highlight the local micro-positioning cues T cells encounter as they migrate within inflamed tissues, from surrounding ECM and signaling molecules, as well as a requirement for appropriate long-range macro-positioning within distinct tissue compartments or at discrete foci of infection or tissue damage. The central nervous system (CNS) responds to injury and infection by extensively remodeling the ECM and with the de novo generation of a fibroblastic reticular network that likely influences T cell motility. We examine how inflammation-induced changes to the CNS landscape may regulate T cell tissue exploration and modulate function. PMID:27790220

Hepatitis B core protein promotes liver cancer metastasis through miR-382-5p/DLC-1 axis.

PubMed

Du, Juan; Bai, Fuxiang; Zhao, Peiqing; Li, Xiaoyan; Li, Xueen; Gao, Lifen; Ma, Chunhong; Liang, Xiaohong

2018-01-01

The hepatitis B virus core protein (HBc), also named core antigen, is well-known for its key role in viral capsid formation and virus replication. Recently, studies showed that HBc has the potential to control cell biology activity by regulating host gene expression. Here, we utilized miRNA microarray to identify 24 upregulated miRNAs and 21 downregulated miRNAs in HBc-expressed HCC cells, which were involved in multiple biological processes, including cell motility. Consistently, the in vitro transwell assay and the in vivo tail-vein injection model showed HBc promotion on HCC metastasis. Further, the miRNA-target gene network analysis displayed that the deleted in liver cancer (DLC-1) gene, an important negative regulator for cell motility, was potentially targeted by several differentially expressed miRNAs in HBc-introduced cells. Introduction of miRNAs mimics or inhibitors and 3'UTR luciferase activity assay proved that miR-382-5p efficiently suppressed DLC-1 expression and its 3'-UTR luciferase reporter activity. Importantly, cotransfection of miR-382-5p mimics/inhibitors and the DLC-1 expression vector almost abrogated HBc promotion on cell motility, indicating that the miR-382-5p/DLC-1 axis is important for mediating HBc-enhanced HCC motility. Clinical HCC samples also showed a negative correlation between miR-382-5p and DLC-1 expression level. Furthermore, HBc-positive HCC tissues showed high miR-382-5p level and reduced DLC-1 expression. In conclusion, our findings revealed that HBc promoted HCC motility by regulating the miR-382-5p/DLC-1 axis, which might provide a novel target for clinical diagnosis and treatment. Copyright © 2017 Elsevier B.V. All rights reserved.

Single cell swimming dynamics of Listeria monocytogenes using a nanoporous microfluidic platform

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wright, Evan; Neethirajan, Suresh; Warriner, Keith

2014-01-01

Listeria monocytogenes remains a significant foodborne pathogen due to its virulence and ability to become established in food processing facilities. The pathogen is characterized by its ability to grow over a wide temperature range and withstand a broad range of stresses. The following reports on the chemotaxis and motility of the L. monocytogenes when exposed to relatively small concentrations of acetic acid. Using the developed nanoporous microfluidic device to precisely modulate the cellular environment, we exposed the individual Listeria cells to acetic acid and, in real time and with high resolution, observed how the cells reacted to the change inmore » their surroundings. Our results showed that concentrations of acetic acid below 10 mM had very little, if any, effect on the motility. However, when exposed to 100 mM acetic acid, the cells exhibited a sharp drop in velocity and displayed a more random pattern of motion. These results indicate that at appropriate concentrations, acetic acid has the ability to disable the flagellum of the cells, thus impairing their motility. This drop in motility has numerous effects on the cell; its main effects being the obstruction of the cell's ability to properly form biofilms and a reduction in the overall infectivity of the cells. Since these characteristics are especially useful in controlling the proliferation of L. monocytogenes, acetic acid shows potential for application in the food industry as an active compound in designing a food packaging environment and as an antimicrobial agent.« less

The relationship between gastrointestinal motility and gastric dilatation-volvulus in dogs.

PubMed

Gazzola, Krista M; Nelson, Laura L

2014-09-01

Gastric dilatation-volvulus (GDV) is a devastating disease that most commonly affects large and giant-breed dogs. Though a number of risk factors have been associated with the development of GDV, the etiology of GDV remains unclear. Abnormal gastric motility patterns and delayed gastric emptying have been previously described in dogs following GDV. Work evaluating the effects of gastropexy procedures and changes to gastric motility after experimental GDV has not found the same changes as those found in dogs with naturally occurring GDV. Although the role of abnormal gastric motility in dogs with GDV will need to be clarified with additional research, such study is likely to be facilitated by improved access to and development of noninvasive measurement techniques for the evaluation of gastric emptying and other motility parameters. In particular, the availability of Food and Drug Administration-approved wireless motility devices for the evaluation of gastrointestinal motility is particularly promising in the study of GDV and other functional gastrointestinal diseases of large and giant-breed dogs. Published by Elsevier Inc.

Competitive Stem Cell Recruitment by Multiple Cytotactic Cues

PubMed Central

Mendelson, Avital; Cheung, Yukkee; Paluch, Kamila; Chen, Mo; Kong, Kimi; Tan, Jiali; Dong, Ziming; Sia, Samuel K.; Mao, Jeremy J.

2014-01-01

A multitude of cytotactic cues direct cell migration in development, cancer metastasis and wound healing. However, our understanding of cell motility remains fragmented partially because current migration devices only allow the study of independent factors. We developed a cell motility assay that allows competitive recruitment of a given cell population simultaneously by gradients of multiple cytotactic cues, observable under real-time imaging. Well-defined uniform gradients of cytotactic cues can be independently generated and sustained in each channel. As a case study, bone marrow mesenchymal stem/stromal cells (MSCs) were exposed to 15 cytokines that are commonly present in arthritis. Cytokines that induced robust recruitment of MSCs in multiple groups were selected to ‘compete’ in a final round to yield the most chemotactic factor(s) based on cell migration numbers, distances, migration indices and motility over time. The potency of a given cytokine in competition frequently differed from its individual action, substantiating the need to test multiple cytokines concurrently due to synergistic or antagonistic effects. This new device has the rare capacity to screen molecules that induce cell migration in cancer therapy, drug development and tissue regeneration. PMID:23364311

The physics of the unconventional motility strategy of euglenids

NASA Astrophysics Data System (ADS)

Arroyo, Marino; Noselli, Giovanni; Desimone, Antonio

Euglenids are a family of unicellular protists, which use flagella to move in a fluid. However, they are also capable of performing elegantly concerted large amplitude deformations of the cell shape, in what is known as metaboly. To perform metaboly, euglenids use an elaborate cortical complex capable of actively imposing spatially modulated shear deformations on the cell surface. This mode of cell deformation has been linked to motility, but biophysical studies have demonstrated that it leads to very small swimming velocities as compared to flagellar locomotion. Furthermore, why would these cells possess two elaborate apparatus for the same function remains unclear. In this work, we combine experimental observations of euglena gracilis cells with theoretical models to shed light into the function of metaboly. The theoretical models account for the force generation and shape evolution at the cell envelop, together with the mechanical interaction of the cell with its environment. We characterize the efficiency of the two modes of locomotion of this cells in terms of the physical nature of their environment. ERC AdG 340685 MicroMotility.

Antibiotic Algae by Chemical Surface Engineering.

PubMed

Kerschgens, Isabel P; Gademann, Karl

2018-03-02

Chemical cell-surface engineering is a tool for modifying and altering cellular functions. Herein, we report the introduction of an antibiotic phenotype to the green alga Chlamydomonas reinhardtii by chemically modifying its cell surface. Flow cytometry and confocal microscopy studies demonstrated that a hybrid of the antibiotic vancomycin and a 4-hydroxyproline oligomer binds reversibly to the cell wall without affecting the viability or motility of the cells. The modified cells were used to inhibit bacterial growth of Gram-positive Bacillus subtilis cultures. Delivery of the antibiotic from the microalgae to the bacterial cells was verified by microscopy. Our studies provide compelling evidence that 1) chemical surface engineering constitutes a useful tool for the introduction of new, previously unknown functionality, and 2) living microalgae can serve as new platforms for drug delivery. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Actin-based motility of Listeria: Right-handed helical trajectories

NASA Astrophysics Data System (ADS)

Rangarajan, Murali

2012-06-01

Bacteria such as Listeria monocytogenes recruit cellular machinery to move in and between cells. Understanding the mechanism of motility, including force and torque generation and the resultant displacements, holds keys to numerous applications in medicine and biosensing. In this work, a simple back-of-the-envelope calculation is presented to illustrate that a biomechanical model of actin-based motility of a rigid surface through persistently attached filaments propelled by affinity-modulated molecular motors can produce a right-handed helical trajectory consistent with experimental observations. The implications of the mechanism to bacterial motility are discussed.

WASP and SCAR are evolutionarily conserved in actin-filled pseudopod-based motility

PubMed Central

2017-01-01

Diverse eukaryotic cells crawl through complex environments using distinct modes of migration. To understand the underlying mechanisms and their evolutionary relationships, we must define each mode and identify its phenotypic and molecular markers. In this study, we focus on a widely dispersed migration mode characterized by dynamic actin-filled pseudopods that we call “α-motility.” Mining genomic data reveals a clear trend: only organisms with both WASP and SCAR/WAVE—activators of branched actin assembly—make actin-filled pseudopods. Although SCAR has been shown to drive pseudopod formation, WASP’s role in this process is controversial. We hypothesize that these genes collectively represent a genetic signature of α-motility because both are used for pseudopod formation. WASP depletion from human neutrophils confirms that both proteins are involved in explosive actin polymerization, pseudopod formation, and cell migration. WASP and WAVE also colocalize to dynamic signaling structures. Moreover, retention of WASP together with SCAR correctly predicts α-motility in disease-causing chytrid fungi, which we show crawl at >30 µm/min with actin-filled pseudopods. By focusing on one migration mode in many eukaryotes, we identify a genetic marker of pseudopod formation, the morphological feature of α-motility, providing evidence for a widely distributed mode of cell crawling with a single evolutionary origin. PMID:28473602

Association of Lis1 with outer arm dynein is modulated in response to alterations in flagellar motility

PubMed Central

Rompolas, Panteleimon; Patel-King, Ramila S.; King, Stephen M.

2012-01-01

The cytoplasmic dynein regulatory factor Lis1, which induces a persistent tight binding to microtubules and allows for transport of cargoes under high-load conditions, is also present in motile cilia/flagella. We observed that Lis1 levels in flagella of Chlamydomonas strains that exhibit defective motility due to mutation of various axonemal substructures were greatly enhanced compared with wild type; this increase was absolutely dependent on the presence within the flagellum of the outer arm dynein α heavy chain/light chain 5 thioredoxin unit. To assess whether cells might interpret defective motility as a “high-load environment,” we reduced the flagellar beat frequency of wild-type cells through enhanced viscous load and by reductive stress; both treatments resulted in increased levels of flagellar Lis1, which altered the intrinsic beat frequency of the trans flagellum. Differential extraction of Lis1 from wild-type and mutant axonemes suggests that the affinity of outer arm dynein for Lis1 is directly modulated. In cytoplasm, Lis1 localized to two punctate structures, one of which was located near the base of the flagella. These data reveal that the cell actively monitors motility and dynamically modulates flagellar levels of the dynein regulatory factor Lis1 in response to imposed alterations in beat parameters. PMID:22855525

Up-regulation of neogenin-1 increases cell proliferation and motility in gastric cancer

PubMed Central

Kim, Seok-Jun; Wang, Yuan-Guo; Lee, Hyun-Woo; Gu Kang, Hyeok; La, Sun-Hyuk; Ju Choi, Il; Irimura, Tatsuro; Ro, Jae Y.; Bresalier, Robert S.; Chun, Kyung-Hee

2014-01-01

Although elevated expression of neogenin-1 has been detected in human gastric cancer tissue, its role in gastric tumorigenesis remains unclear due to the lack of neogenin-1 studies in cancer. Therefore, we demonstrated here the function and regulatory mechanism of neogenin-1 in gastric cancer. Neogenin-1 ablation decreased proliferation and migration of gastric cancer cells, whereas its over-expression reversed these effects. Xenografted analyses using gastric cancer cells displayed statistically significant inhibition of tumor growth by neogenin-1 depletion. Interestingly, galectin-3 interacted with HSF-1 directly, which facilitated nuclear-localization and binding on neogenin-1 promoter to drive its transcription and gastric cancer cell motility. The galectin-3-increased gastric cancer cell motility was down-regulated by HSF-1 depletion. Moreover, the parallel expression patterns of galectin-3 and neogenin-1, as well as those of HSF-1 and neogenin-1, were detected in the malignant tissues of gastric cancer patients. Taken together, high-expression of neogenin-1 promotes gastric cancer proliferation and motility and its expression is regulated by HSF-1 and galectin-3 interaction. In addition, we propose further studies for neogenin-1 and its associated pathways to provide them as a proper target for gastric cancer therapy. PMID:24930499

Light-induced motility of thermophilic Synechococcus isolates from Octopus Spring, Yellowstone National Park.

PubMed

Ramsing, N B; Ferris, M J; Ward, D M

1997-06-01

This study demonstrates light-induced motility of two thermophilic Synechococcus isolates that are morphologically similar but that belong to different cyanobacterial lineages. Both isolates migrated away from densely inoculated streaks to form fingerlike projections extending toward or away from the light source, depending on the light intensity. However, the two isolates seemed to prefer widely different light conditions. The behavior of each isolate was controlled by several factors, including temperature, preacclimation of inocula, acclimation during the experiment, and strain-specific genetic preferences for different light conditions (adaptation). Time-lapse microscopy confirmed that these projections were formed by actively gliding cells and were not simply the outcome of directional cell division. The observed motility rates of individual cells of 0.1 to 0.3 micrometers s-1 agreed well with the distance traversed by the projections, 0.3 to 0.5 mm h-1, suggesting that most cells in each projection are travelling in the same direction. The finding of motility among two phylogenetically unaffiliated unicellular cyanobacteria suggests that this trait may be widespread among this group. If so, this would have important implications for experiments on colonization, succession, diel positioning, and photosynthetic activity in hot spring mats dominated by Synechococcus-like cyanobacteria.

Modeling and predictions of biphasic mechanosensitive cell migration altered by cell-intrinsic properties and matrix confinement.

PubMed

Pathak, Amit

2018-04-12

Motile cells sense the stiffness of their extracellular matrix (ECM) through adhesions and respond by modulating the generated forces, which in turn lead to varying mechanosensitive migration phenotypes. Through modeling and experiments, cell migration speed is known to vary with matrix stiffness in a biphasic manner, with optimal motility at an intermediate stiffness. Here, we present a two-dimensional cell model defined by nodes and elements, integrated with subcellular modeling components corresponding to mechanotransductive adhesion formation, force generation, protrusions and node displacement. On 2D matrices, our calculations reproduce the classic biphasic dependence of migration speed on matrix stiffness and predict that cell types with higher force-generating ability do not slow down on very stiff matrices, thus disabling the biphasic response. We also predict that cell types defined by lower number of total receptors require stiffer matrices for optimal motility, which also limits the biphasic response. For a cell type with robust biphasic migration on 2D surface, simulations in channel-like confined environments of varying width and height predict faster migration in more confined matrices. Simulations performed in shallower channels predict that the biphasic mechanosensitive cell migration response is more robust on 2D micro-patterns as compared to the channel-like 3D confinement. Thus, variations in the dimensionality of matrix confinement alters the way migratory cells sense and respond to the matrix stiffness. Our calculations reveal new phenotypes of stiffness- and topography-sensitive cell migration that critically depend on both cell-intrinsic and matrix properties. These predictions may inform our understanding of various mechanosensitive modes of cell motility that could enable tumor invasion through topographically heterogeneous microenvironments. © 2018 IOP Publishing Ltd.

Two-Photon Microscopy Imaging of thy1GFP-M Transgenic Mice: A Novel Animal Model to Investigate Brain Dendritic Cell Subsets In Vivo

PubMed Central

Laperchia, Claudia; Allegra Mascaro, Anna L.; Sacconi, Leonardo; Andrioli, Anna; Mattè, Alessandro; De Franceschi, Lucia; Grassi-Zucconi, Gigliola; Bentivoglio, Marina; Buffelli, Mario; Pavone, Francesco S.

2013-01-01

Transgenic mice expressing fluorescent proteins in specific cell populations are widely used for in vivo brain studies with two-photon fluorescence (TPF) microscopy. Mice of the thy1GFP-M line have been engineered for selective expression of green fluorescent protein (GFP) in neuronal populations. Here, we report that TPF microscopy reveals, at the brain surface of these mice, also motile non-neuronal GFP+ cells. We have analyzed the behavior of these cells in vivo and characterized in brain sections their immunophenotype. With TPF imaging, motile GFP+ cells were found in the meninges, subarachnoid space and upper cortical layers. The striking feature of these cells was their ability to move across the brain parenchyma, exhibiting evident shape changes during their scanning-like motion. In brain sections, GFP+ cells were immunonegative to antigens recognizing motile cells such as migratory neuroblasts, neuronal and glial precursors, mast cells, and fibroblasts. GFP+ non-neuronal cells exhibited instead the characteristic features and immunophenotype (CD11c and major histocompatibility complex molecule class II immunopositivity) of dendritic cells (DCs), and were immunonegative to the microglial marker Iba-1. GFP+ cells were also identified in lymph nodes and blood of thy1GFP-M mice, supporting their identity as DCs. Thus, TPF microscopy has here allowed the visualization for the first time of the motile behavior of brain DCs in situ. The results indicate that the thy1GFP-M mouse line provides a novel animal model for the study of subsets of these professional antigen-presenting cells in the brain. Information on brain DCs is still very limited and imaging in thy1GFP-M mice has a great potential for analyses of DC-neuron interaction in normal and pathological conditions. PMID:23409142

Modification of Salmonella Typhimurium Motility by the Probiotic Yeast Strain Saccharomyces boulardii

PubMed Central

Pontier-Bres, Rodolphe; Prodon, François; Munro, Patrick; Rampal, Patrick; Lemichez, Emmanuel; Peyron, Jean François; Czerucka, Dorota

2012-01-01

Background Motility is an important component of Salmonella enterica serovar Typhimurium (ST) pathogenesis allowing the bacteria to move into appropriate niches, across the mucus layer and invade the intestinal epithelium. In vitro, flagellum-associated motility is closely related to the invasive properties of ST. The probiotic yeast Saccharomyces boulardii BIOCODEX (S.b-B) is widely prescribed for the prophylaxis and treatment of diarrheal diseases caused by bacteria or antibiotics. In case of Salmonella infection, S.b-B has been shown to decrease ST invasion of T84 colon cell line. The present study was designed to investigate the impact of S.b-B on ST motility. Methodology/Principal Findings Experiments were performed on human colonic T84 cells infected by the Salmonella strain 1344 alone or in the presence of S.b-B. The motility of Salmonella was recorded by time-lapse video microscopy. Next, a manual tracking was performed to analyze bacteria dynamics (MTrackJ plugin, NIH image J software). This revealed that the speed of bacterial movement was modified in the presence of S.b-B. The median curvilinear velocity (CLV) of Salmonella incubated alone with T84 decreased from 43.3 µm/sec to 31.2 µm/sec in the presence of S.b-B. Measurement of track linearity (TL) showed similar trends: S.b-B decreased by 15% the number of bacteria with linear tract (LT) and increased by 22% the number of bacteria with rotator tract (RT). Correlation between ST motility and invasion was further established by studying a non-motile flagella-deficient ST strain. Indeed this strain that moved with a CLV of 0.5 µm/sec, presented a majority of RT and a significant decrease in invasion properties. Importantly, we show that S.b-B modified the motility of the pathogenic strain SL1344 and significantly decreased invasion of T84 cells by this strain. Conclusions This study reveals that S.b-B modifies Salmonella's motility and trajectory which may account for the modification of Salmonella's invasion. PMID:22442723

A biopolymer transistor: electrical amplification by microtubules.

PubMed

Priel, Avner; Ramos, Arnolt J; Tuszynski, Jack A; Cantiello, Horacio F

2006-06-15

Microtubules (MTs) are important cytoskeletal structures engaged in a number of specific cellular activities, including vesicular traffic, cell cyto-architecture and motility, cell division, and information processing within neuronal processes. MTs have also been implicated in higher neuronal functions, including memory and the emergence of "consciousness". How MTs handle and process electrical information, however, is heretofore unknown. Here we show new electrodynamic properties of MTs. Isolated, taxol-stabilized MTs behave as biomolecular transistors capable of amplifying electrical information. Electrical amplification by MTs can lead to the enhancement of dynamic information, and processivity in neurons can be conceptualized as an "ionic-based" transistor, which may affect, among other known functions, neuronal computational capabilities.

CXCR4/CXCL12 signaling impacts enamel progenitor cell proliferation and motility in the dental stem cell niche

PubMed Central

Otsu, Keishi; Harada, Hidemitsu; Shibata, Shunichi; Obara, Nobuko; Irie, Kazuharu; Taniguchi, Akiyoshi; Nagasawa, Takashi; Aoki, Kazunari; Caliari, Steven R.; Weisgerber, Daniel W.

2015-01-01

Dental stem cells are located at the proximal ends of rodent incisors. These stem cells reside in the dental epithelial stem cell niche, termed the apical bud. We focused on identifying critical features of a chemotactic signal in the niche. Here, we report that CXCR4/CXCL12 signaling impacts enamel progenitor cell proliferation and motility in dental stem cell niche cells. We report cells in the apical bud express CXCR4 mRNA at high levels while expression is restricted in the basal epithelium (BE) and transit-amplifying (TA) cell regions. Furthermore, the CXCL12 ligand is present in mesenchymal cells adjacent to the apical bud. We then performed gain- and loss-of-function analyses to better elucidate the role of CXCR4 and CXCL12. CXCR4-deficient mice contain epithelial cell aggregates, while cell proliferation in mutant incisors was also significantly reduced. We demonstrate in vitro that dental epithelial cells migrate toward sources of CXCL12, whereas knocking down CXCR4 impaired motility and resulted in formation of dense cell colonies. These results suggest that CXCR4 expression may be critical for activation of enamel progenitor cell division and that CXCR4/CXCL12 signaling may control movement of epithelial progenitors from the dental stem cell niche. PMID:26246398

Two problems in multiphase biological flows: Blood flow and particulate transport in microvascular network, and pseudopod-driven motility of amoeboid cells

NASA Astrophysics Data System (ADS)

Bagchi, Prosenjit

2016-11-01

In this talk, two problems in multiphase biological flows will be discussed. The first is the direct numerical simulation of whole blood and drug particulates in microvascular networks. Blood in microcirculation behaves as a dense suspension of heterogeneous cells. The erythrocytes are extremely deformable, while inactivated platelets and leukocytes are nearly rigid. A significant progress has been made in recent years in modeling blood as a dense cellular suspension. However, many of these studies considered the blood flow in simple geometry, e.g., straight tubes of uniform cross-section. In contrast, the architecture of a microvascular network is very complex with bifurcating, merging and winding vessels, posing a further challenge to numerical modeling. We have developed an immersed-boundary-based method that can consider blood cell flow in physiologically realistic and complex microvascular network. In addition to addressing many physiological issues related to network hemodynamics, this tool can be used to optimize the transport properties of drug particulates for effective organ-specific delivery. Our second problem is pseudopod-driven motility as often observed in metastatic cancer cells and other amoeboid cells. We have developed a multiscale hydrodynamic model to simulate such motility. We study the effect of cell stiffness on motility as the former has been considered as a biomarker for metastatic potential. Funded by the National Science Foundation.

Evaluation of epididymis storage temperature and cryopreservation conditions for improved mitochondrial membrane potential, membrane integrity, sperm motility and in vitro fertilization in bovine epididymal sperm.

PubMed

Nichi, M; Rijsselaere, T; Losano, Jda; Angrimani, Dsr; Kawai, Gkv; Goovaerts, Igf; Van Soom, A; Barnabe, V H; De Clercq, Jbp; Bols, Pej

2017-04-01

The maintaining of the epididymis at lower temperatures during storage and transport improves sperm quality. Our study aimed to test whether epididymis storage temperature (post-mortem) and sperm cryopreservation affect sperm kinetics, membrane integrity, mitochondrial potential and fertility capacity. Thirty-six epididymides were collected from 18 bulls after slaughter and divided into two groups: at 4 or 34°C for 2-3 hr. The sperm was collected from the epididymis cauda. The evaluation consisted of computer-assisted sperm analysis (CASA), SYBR14/PI/JC1 to evaluate membrane integrity, mitochondrial membrane potential (MMP) and measurement of lipid peroxidation (TBARS). The sperm was then frozen using an automatic device. After thawing, sperm samples were evaluated by the same variables and further in vitro fertilization rates. Cryopreservation negatively affected sperm motility in samples stored at 4 and 34°C. Nevertheless, the 4°C samples yielded higher rates of blastocyst formation. Pre-freeze sperm motility, progressive motility and velocity were higher in sperm from epididymis stored at 4°C while post-thaw sperm motility, progressive motility and velocity remained the same among samples from epididymis stored at 4 or 34°C. However, with regard to the kinetic patterns, samples collected from epididymis stored at 34°C had lower values when compared to those stored at 4°C prior the cryopreservation process. Our results indicate that epididymis handling conditions after cryopreservation may affect sperm quality after thawing, especially due to compromised MMP in sperm collected from epididymis stored at higher temperatures. © 2016 Blackwell Verlag GmbH.

Flow cytometry application in the assessment of sperm DNA integrity of men with asthenozoospermia.

PubMed

Piasecka, M; Gaczarzewicz, D; Laszczyńska, M; Starczewski, A; Brodowska, A

2007-01-01

Sperm genomic integrity and ultrastructural features of ejaculated spermatozoa contributing to the assessment of gamete fertility potential in patients with asthenozoospermia are discussed. The proportion of TUNEL-positive cells was significantly higher in the semen of patients with low sperm motility (n=40; p<0.01) as compared to men with normal sperm motility (n=54). Sperm DNA fragmentation negatively correlated (n=94) with sperm motility, sperm concentration, and integrity of the sperm cellular membrane (HOS-test). Two categories of patients were distinguished: (1) patients (23 out of 94 subjects) with < or = 4% of TUNEL-positive cells and (2) patients (71 subjects) with 4% of TUNEL-positive cells. A significant difference was noted in the sperm motility and HOS-test results between patients from both groups. Large numbers of immature spermatozoa with extensive cytoplasmic retention, ultrastructural chromatin and midpiece abnormalities, and conglomerates containing sperm fragments were present more frequently in the semen of asthenozoospermic subjects with >4% of TUNEL-positive sperm cells. Low sperm motility seems to be accompanied by serious defects of gamete chromatin expressed as diminished sperm genomic integrity and abnormal DNA condensation and by defects of sperm midpiece. These abnormalities may reflect developmental failure during the spermatogenic remodeling process. The DNA fragmentation test may be considered as an additional assay for the evaluation of spermatozoa beside standard analysis and taken together with electron microscopy may help to determine the actual number of "healthy" spermatozoa thereby playing an important role during diagnosis and treatment of male infertility.

Spirochete motility and morpholgy

NASA Astrophysics Data System (ADS)

Charon, Nyles

2004-03-01

Spirochetes have a unique structure, and as a result their motility is different from that of other bacteria. These organisms can swim in a highly viscous, gel-like medium, such as that found in connective tissue, that inhibits the motility of most other bacteria. In spirochetes, the organelles for motility, the periplasmic flagella, reside inside the cell within the periplasmic space. A given periplasmic flagellum is attached only at one end of the cell, and depending on the species, may or may not overlap in the center of the cell. The number of periplasmic flagella varies from species to species. These structures have been shown to be directly involved in motility and function by rotating within the periplasmic space (1). The present talk focuses on the spirochete that causes Lyme disease, Borrelia burgdorferi. In many bacterial species, cell shape is usually dictated by the peptidoyglycan layer of the cell wall. In the first part of the talk, results will be presented that the morphology of B. burgdorferi is the result of a complex interaction between the cell cylinder and the internal periplasmic flagella resulting in a cell with a flat-wave morphology. Backward moving, propagating waves enable these bacteria to swim and translate in a given direction. Using targeted mutagenesis, we inactivated the gene encoding the major periplasmic flagellar filament protein FlaB. The resulting flaB mutants not only were non-motile, but were rod-shaped (2). Western blot analysis indicated that flaB was no longer synthesized, and electron microscopy revealed that the mutants were completely deficient in periplasmic flagella. Our results indicate that the periplasmic flagella of B. burgdorferi have a skeletal function. These organelles dynamically interact with the rod-shaped cell cylinder to enable the cell to swim, and to confer in part its flat-wave morphology The latter part of the talk concerns the basis for asymmetrical rotation of the periplasmic flagella of B. burgdorferi during chemotaxis. In translational motility, the bundles of periplasmic flagella rotate in opposite directions. When not translating, they rotate in the same direction, and the cells flex. We present evidence that asymmetrical rotation of the bundles during translation does not depend upon the chemotaxis signal transduction system. The histidine kinase CheA is known to be an essential component in the signaling pathway for bacterial chemotaxis. Mutants of cheA in flagellated bacteria continually rotate their flagella in one direction. B. burgdorferi has two copies of cheA. We reasoned that if chemotaxis were essential for asymmetrical rotation of the flagellar bundles, and if the flagellar motors at both cell ends were identical, inactivation of the two cheA genes should result in cells that constant flex. To test this hypothesis, the signaling pathway was completely blocked by construction of a double cheA mutant. This mutant was completely deficient in chemotaxis. Rather than flexing, it failed to reverse, and it continually translated only in one direction. The results indicate that asymmetrical rotation does not depend upon the chemotaxis system but rather upon differences between the two flagellar bundles. We propose that certain factors within the spirochete localize at flagellar motors at one end of the cell to effect this asymmetry (3). References: 1. Charon, N.W. and S.F. Goldstein. 2002. The genetics of motility and chemotaxis of a fascinating group of bacteria: the spirochetes. Ann. Rev. Genetics. 36: 47-73. 2. Motaleb M.A., L. Corum, J.L Bono, A.F. Elias, P. Rosa, D.S. Samuels, N.W. Charon. 2000. Borrelia burgdorferi periplasmic flagella have both skeletal and motility functions. Proc Natl Acad Sci. 2000 97:10899-10904. 3. Li, C. R. Bakker, M. Motaleb, F. Cabello, M.L. Sartakova, and N.W. Charon. 2002. Asymmetrical flagellar rotation in Borrelia burgdorferi non-chemotaxis mutants. Proc. Natl. Acad. Sci. 99:6169-6174.

Membrane tension and cytoskeleton organization in cell motility.

PubMed

Sens, Pierre; Plastino, Julie

2015-07-15

Cell membrane shape changes are important for many aspects of normal biological function, such as tissue development, wound healing and cell division and motility. Various disease states are associated with deregulation of how cells move and change shape, including notably tumor initiation and cancer cell metastasis. Cell motility is powered, in large part, by the controlled assembly and disassembly of the actin cytoskeleton. Much of this dynamic happens in close proximity to the plasma membrane due to the fact that actin assembly factors are membrane-bound, and thus actin filaments are generally oriented such that their growth occurs against or near the membrane. For a long time, the membrane was viewed as a relatively passive scaffold for signaling. However, results from the last five years show that this is not the whole picture, and that the dynamics of the actin cytoskeleton are intimately linked to the mechanics of the cell membrane. In this review, we summarize recent findings concerning the role of plasma membrane mechanics in cell cytoskeleton dynamics and architecture, showing that the cell membrane is not just an envelope or a barrier for actin assembly, but is a master regulator controlling cytoskeleton dynamics and cell polarity.

Modelling cell motility and chemotaxis with evolving surface finite elements

PubMed Central

Elliott, Charles M.; Stinner, Björn; Venkataraman, Chandrasekhar

2012-01-01

We present a mathematical and a computational framework for the modelling of cell motility. The cell membrane is represented by an evolving surface, with the movement of the cell determined by the interaction of various forces that act normal to the surface. We consider external forces such as those that may arise owing to inhomogeneities in the medium and a pressure that constrains the enclosed volume, as well as internal forces that arise from the reaction of the cells' surface to stretching and bending. We also consider a protrusive force associated with a reaction–diffusion system (RDS) posed on the cell membrane, with cell polarization modelled by this surface RDS. The computational method is based on an evolving surface finite-element method. The general method can account for the large deformations that arise in cell motility and allows the simulation of cell migration in three dimensions. We illustrate applications of the proposed modelling framework and numerical method by reporting on numerical simulations of a model for eukaryotic chemotaxis and a model for the persistent movement of keratocytes in two and three space dimensions. Movies of the simulated cells can be obtained from http://homepages.warwick.ac.uk/∼maskae/CV_Warwick/Chemotaxis.html. PMID:22675164

Transient gene silencing of galectin-3 suppresses pancreatic cancer cell migration and invasion through degradation of β-catenin

PubMed Central

Kobayashi, Tsutomu; Shimura, Tatsuo; Yajima, Toshiki; Kubo, Norio; Araki, Kenichiro; Tsutsumi, Soichi; Suzuki, Hideki; Kuwano, Hiroyuki; Raz, Avraham

2013-01-01

Pancreatic cancer is a leading cause of cancer-related mortality and often has a poor prognosis because of its late diagnosis, aggressive local invasion, early metastasis, and poor response to chemotherapy. The chemotherapeutic agent gemcitabine is effective for treating advanced pancreatic cancer, but its efficacy remains less than satisfactory. It is expected that further investigation of pancreatic cancer cell invasion and development of strategies to block this process should improve the disease prognosis. In this study, we tested our hypothesis that galectin-3 (gal-3), a multifunctional member of the β-galactoside-binding protein family, may regulate pancreatic cancer cell motility, and silencing of it inhibit cell motility. Previous studies demonstrated that this protein is associated with tumor cell adhesion, proliferation, differentiation, angiogenesis, apoptosis, and metastasis. Here, we used gal-3 small interfering RNA (siRNA) to silence its expression in various pancreatic cancer cell lines to determine whether gal-3 regulates cell proliferation, migration and invasion in vitro. We found that silencing gal-3 reduced cellular migration and invasion, but failed to affect proliferation. In gal-3 siRNA-transfected cells, we detected a decrease in β-catenin expression, an important signal for cancer cell invasion, which was caused by down-regulation of phosphorylated Akt and GSK-3β. We also found that matrix metalloproteinase (MMP)-2 expression was reduced by gal-3 silencing. These results indicate that gal-3-mediated invasion via MMP-2 regulated by β-catenin degradation is initiated by Akt phosphorylation in pancreatic cancer cells. Our results suggest that gal-3 can be a novel therapeutic target in pancreatic cancer. PMID:21448903

Protein Tyrosine Phosphatase-PEST and β8 Integrin Regulate Spatiotemporal Patterns of RhoGDI1 Activation in Migrating Cells

PubMed Central

Lee, Hye Shin; Cheerathodi, Mujeeburahiman; Chaki, Sankar P.; Reyes, Steve B.; Zheng, Yanhua; Lu, Zhimin; Paidassi, Helena; DerMardirossian, Celine; Lacy-Hulbert, Adam; Rivera, Gonzalo M.

2015-01-01

Directional cell motility is essential for normal development and physiology, although how motile cells spatiotemporally activate signaling events remains largely unknown. Here, we have characterized an adhesion and signaling unit comprised of protein tyrosine phosphatase (PTP)-PEST and the extracellular matrix (ECM) adhesion receptor β8 integrin that plays essential roles in directional cell motility. β8 integrin and PTP-PEST form protein complexes at the leading edge of migrating cells and balance patterns of Rac1 and Cdc42 signaling by controlling the subcellular localization and phosphorylation status of Rho GDP dissociation inhibitor 1 (RhoGDI1). Translocation of Src-phosphorylated RhoGDI1 to the cell's leading edge promotes local activation of Rac1 and Cdc42, whereas dephosphorylation of RhoGDI1 by integrin-bound PTP-PEST promotes RhoGDI1 release from the membrane and sequestration of inactive Rac1/Cdc42 in the cytoplasm. Collectively, these data reveal a finely tuned regulatory mechanism for controlling signaling events at the leading edge of directionally migrating cells. PMID:25666508

Dia-Interacting Protein (DIP) Imposes Migratory Plasticity in mDia2-Dependent Tumor Cells in Three-Dimensional Matrices

PubMed Central

Wyse, Meghan M.; Lei, Jun; Nestor-Kalinoski, Andrea L.; Eisenmann, Kathryn M.

2012-01-01

Tumor cells rely upon membrane pliancy to escape primary lesions and invade secondary metastatic sites. This process relies upon localized assembly and disassembly cycles of F-actin that support and underlie the plasma membrane. Dynamic actin generates both spear-like and bleb structures respectively characterizing mesenchymal and amoeboid motility programs utilized by metastatic cells in three-dimensional matrices. The molecular mechanism and physiological trigger(s) driving membrane plasticity are poorly understood. mDia formins are F-actin assembly factors directing membrane pliancy in motile cells. mDia2 is functionally coupled with its binding partner DIP, regulating cortical actin and inducing membrane blebbing in amoeboid cells. Here we show that mDia2 and DIP co-tether to nascent blebs and this linkage is required for bleb formation. DIP controls mesenchymal/amoeboid cell interconvertability, while CXCL12 induces assembly of mDia2:DIP complexes to bleb cortices in 3D matrices. These results demonstrate how DIP-directed mDia2-dependent F-actin dynamics regulate morphological plasticity in motile cancer cells. PMID:23024796

Active unjamming of confluent cell layers

NASA Astrophysics Data System (ADS)

Marchetti, M. Cristina

Cell motion inside dense tissues governs many biological processes, including embryonic development and cancer metastasis, and recent experiments suggest that these tissues exhibit collective glassy behavior. Motivated by these observations, we have studied a model of dense tissues that combines self-propelled particle models and vertex models of confluent cell layers. In this model, referred to as self-propelled Voronoi (SPV), cells are described as polygons in a Voronoi tessellation with directed noisy cell motility and interactions governed by a shape energy that incorporates the effects of cell volume incompressibility, contractility and cell-cell adhesion. Using this model, we have demonstrated a new density-independent solid-liquid transition in confluent tissues controlled by cell motility and a cell-shape parameter measuring the interplay of cortical tension and cell-cell adhesion. An important insight of this work is that the rigidity and dynamics of cell layers depends sensitively on cell shape. We have also used the SPV model to test a new method developed by our group to determine cellular forces and tissue stresses from experimentally accessible cell shapes and traction forces, hence providing the spatio-temporal distribution of stresses in motile dense tissues. This work was done with Dapeng Bi, Lisa Manning and Xingbo Yang. MCM was supported by NSF-DMR-1305184 and by the Simons Foundation.

Oxidative stress reduces trophoblast FOXO1 and integrin β3 expression that inhibits cell motility.

PubMed

Chen, Chie-Pein; Chen, Cheng-Yi; Wu, Yi-Hsin; Chen, Chia-Yu

2018-06-08

Preeclampsia is a serious pregnancy complication associated with placental oxidative stress and impaired trophoblast migration. The mechanism of defective trophoblast migration remains unknown. Forkhead box O1 (FOXO1) is a transcription factor. Integrin β3 is involved in cell motility. We hypothesized that FOXO1 mediates expression of trophoblast integrin β3, which could be impaired by oxidative stress and have implications in preeclampsia. The expressions of FOXO1 and integrin β3 were significantly reduced in preeclamptic placentas (n = 15) compared to that of controls (n = 15; p < 0.01). HTR-8/SVneo and JEG-3 trophoblasts were transfected to express wild-type FOXO1-WT or constitutively-expressed nuclear mutant form, FOXO1-AAA. The FOXO1 in HTR-8/SVneo and 3A-Sub-E trophoblasts was silenced by small interfering RNA. AKT-mediated phosphorylation inactivated FOXO1, but FOXO1-AAA was not phosphorylated. The expression of trophoblast integrin β3 was significantly elevated by FOXO1 overexpression and inhibited by FOXO1 knockdown. FOXO1 regulates integrin β3 at the transcriptional level via binding to the putative FOXO1 response element site between position -1154 to -1139 (TGAGATGTTTTGAAAG) in HTR-8/SVneo trophoblasts. The level of phosphorylated FOXO1 was decreased, and the FOXO1 level was increased in trophoblasts treated with AKT inhibitor MK2206, leading to upregulation of integrin β3. The capabilities of trophoblast adhesion and migration were enhanced by FOXO1-overexpression or MK2206, and inhibited by silencing FOXO1 or oxidative stress with H 2 O 2 . These results suggest that FOXO1 enhances trophoblast integrin β3 expression, and mediates cell adhesion and migration. By affecting the expression of FOXO1 and cell motility in trophoblasts, oxidative stress plays a role in the development of preeclampsia. Copyright © 2018 Elsevier Inc. All rights reserved.

Evolution and Design Governing Signal Precision and Amplification in a Bacterial Chemosensory Pathway

PubMed Central

Espinosa, Leon; Baronian, Grégory; Molle, Virginie; Mauriello, Emilia M. F.; Brochier-Armanet, Céline; Mignot, Tâm

2015-01-01

Understanding the principles underlying the plasticity of signal transduction networks is fundamental to decipher the functioning of living cells. In Myxococcus xanthus, a particular chemosensory system (Frz) coordinates the activity of two separate motility systems (the A- and S-motility systems), promoting multicellular development. This unusual structure asks how signal is transduced in a branched signal transduction pathway. Using combined evolution-guided and single cell approaches, we successfully uncoupled the regulations and showed that the A-motility regulation system branched-off an existing signaling system that initially only controlled S-motility. Pathway branching emerged in part following a gene duplication event and changes in the circuit structure increasing the signaling efficiency. In the evolved pathway, the Frz histidine kinase generates a steep biphasic response to increasing external stimulations, which is essential for signal partitioning to the motility systems. We further show that this behavior results from the action of two accessory response regulator proteins that act independently to filter and amplify signals from the upstream kinase. Thus, signal amplification loops may underlie the emergence of new connectivity in signal transduction pathways. PMID:26291327

Cancer cell motility: lessons from migration in confined spaces

PubMed Central

Paul, Colin D.; Mistriotis, Panagiotis; Konstantopoulos, Konstantinos

2017-01-01

Time-lapse, deep-tissue imaging made possible by advances in intravital microscopy has demonstrated the importance of tumour cell migration through confining tracks in vivo. These tracks may either be endogenous features of tissues or be created by tumour or tumour-associated cells. Importantly, migration mechanisms through confining microenvironments are not predicted by 2D migration assays. Engineered in vitro models have been used to delineate the mechanisms of cell motility through confining spaces encountered in vivo. Understanding cancer cell locomotion through physiologically relevant confining tracks could be useful in developing therapeutic strategies to combat metastasis. PMID:27909339

Biometric assessment of prostate cancer's metastatic potential.

PubMed

Cooper, C R; Emmett, N; Harris-Hooker, S; Patterson, R; Cooke, D B

1994-01-01

Currently, no protocol exists that can assess the metastatic potential of prostate adenocarcinoma. The reason for this is partly due to the lack of information on cellular changes that result in a tumor cell's becoming metastatic. In this investigation, attempts were made to devise a method that correlated with the metastatic potential of AT-1, Mat-Lu, and Mat-LyLu cell lines of the Dunning R-3327 rat prostatic adenocarcinoma system. To accomplish this, we applied BioQuant biometric parameters, i.e., area, shape factor, and cell motility. AT-1 had a lower shape factor and a greater area as compared with the more highly metastatic Mat-Lu subline. No significant difference in area or shape factor was detected between the AT-1 cell line and the highly metastatic Mat-LyLu line. However, the lowly metastatic AT-1 line had less motility as compared with the Mat-Lu and Mat-LyLu lines. This study revealed that metastatic potential could be partially predicted via area and shape factor and accurately predicted via cell motility.

Colloid centrifugation of fresh stallion semen before cryopreservation decreased microorganism load of frozen-thawed semen without affecting seminal kinetics.

PubMed

Guimarães, T; Lopes, G; Pinto, M; Silva, E; Miranda, C; Correia, M J; Damásio, L; Thompson, G; Rocha, A

2015-01-15

Freezability of equine semen may be influenced by microorganism population of semen. The objective of this study was to verify the effect of single-layer density gradient centrifugation (SLC) of fresh semen before cryopreservation on semen's microbial load (ML) and sperm cells kinetics after freezing-thawing. For that, one ejaculate was collected from 20 healthy stallions and split into control (C) samples (cryopreserved without previous SLC) and SLC samples (subjected to SLC). Semen cryopreservation was performed according to the same protocol in both groups. Microbial load of each microorganism species and total microbial load (TML) expressed in colony-forming units (CFU/mL) as well as frozen-thawed sperm kinetics were assessed in both groups. Additional analysis of the TML was performed, subdividing the frozen-thawed samples in "suitable" (total motility ≥ 30%) and "unsuitable" (total motility < 30%) semen for freezing programs, and comparing the C and SLC groups within these subpopulations. After thawing, SLC samples had less (P < 0.05) TML (88.65 × 10(2) ± 83.8 × 10(2) CFU/mL) than C samples (155.69 × 10(2) ± 48.85 × 10(2) CFU/mL), mainly due to a reduction of Enterococcus spp. and Bacillus spp. A relationship between post-thaw motility and SLC effect on ML was noted, as only in samples with more than 30% total motility was ML reduced (P < 0.05) by SLC (from 51.33 × 10(2) ± 33.26 × 10(2) CFU/mL to 26.68 × 10(2) ± 12.39 × 10(2) CFU/mL in "suitable" frozen-thawed semen vs. 240.90 × 10(2) ± 498.20 × 10(2) to 139.30 × 10(2) ± 290.30 × 10(2) CFU/mL in "unsuitable" frozen-thawed semen). The effect of SLC on kinetics of frozen-thawed sperm cells was negligible. Copyright © 2015 Elsevier Inc. All rights reserved.

Osmotic characteristics and fertility of murine spermatozoa collected in different solutions.

PubMed

Si, Wei; Men, Hongsheng; Benson, James D; Critser, John K

2009-02-01

Osmotic stress is an important factor that can result in cell damage during cryopreservation. Before ejaculation or collection for cryopreservation, murine spermatozoa are stored in epididymal fluid, a physiologically hyperosmotic environment (approximately 415 mmol/kg). The objectives of this study were to determine the osmotic tolerance limits of sperm motion parameters of ICR and C57BL/6 mouse spermatozoa collected in isosmotic (290 mmol/kg) and hyperosmotic (415 mmol/kg) media, and the effect of the osmolality of sperm collection media on sperm fertility after cryopreservation. Our results indicate that murine spermatozoa collected in media with different osmolalities (290 and 415 mmol/kg Dulbecco's phosphate buffered saline (DPBS)) appeared to have different osmotic tolerances for the maintenance of sperm motility and other motion parameters in both mouse strains. The hypo- and hyperosmotic treatments decreased motility and affected other motion parameters of spermatozoa collected in 290 mmol/kg DPBS. The extent of the change of motion parameters after treatments corresponded with the levels of osmotic stress. However, for spermatozoa collected in 415 mmol/kg DPBS, exposure to 290 mmol/kg DPBS tended to increase sperm motility and the quality of their motion parameters. The osmolality of sperm collection medium can affect murine sperm fertility. Spermatozoa collected in 415 mmol/kg medium showed higher fertility compared with spermatozoa collected in 290 mmol/kg as assessed by IVF. Results characterizing murine sperm osmotic tolerance collected in media with different osmolalities from different strains and the effect of collection media osmolality on sperm fertility after cryopreservation will be useful in designing cryopreservation protocols.

Kinesin-1 and mitochondrial motility control by discrimination of structurally equivalent but distinct subdomains in Ran-GTP-binding domains of Ran-binding protein 2.

PubMed

Patil, Hemangi; Cho, Kyoung-in; Lee, James; Yang, Yi; Orry, Andrew; Ferreira, Paulo A

2013-03-27

The pleckstrin homology (PH) domain is a versatile fold that mediates a variety of protein-protein and protein-phosphatidylinositol lipid interactions. The Ran-binding protein 2 (RanBP2) contains four interspersed Ran GTPase-binding domains (RBD(n = 1-4)) with close structural homology to the PH domain of Bruton's tyrosine kinase. The RBD2, kinesin-binding domain (KBD) and RBD3 comprise a tripartite domain (R2KR3) of RanBP2 that causes the unfolding, microtubule binding and biphasic activation of kinesin-1, a crucial anterograde motor of mitochondrial motility. However, the interplay between Ran GTPase and R2KR3 of RanBP2 in kinesin-1 activation and mitochondrial motility is elusive. We use structure-function, biochemical, kinetic and cell-based assays with time-lapse live-cell microscopy of over 260,000 mitochondrial-motility-related events to find mutually exclusive subdomains in RBD2 and RBD3 towards Ran GTPase binding, kinesin-1 activation and mitochondrial motility regulation. The RBD2 and RBD3 exhibit Ran-GTP-independent, subdomain and stereochemical-dependent discrimination on the biphasic kinetics of kinesin-1 activation or regulation of mitochondrial motility. Further, KBD alone and R2KR3 stimulate and suppress, respectively, multiple biophysical parameters of mitochondrial motility. The regulation of the bidirectional transport of mitochondria by either KBD or R2KR3 is highly coordinated, because their kinetic effects are accompanied always by changes in mitochondrial motile events of either transport polarity. These studies uncover novel roles in Ran GTPase-independent subdomains of RBD2 and RBD3, and KBD of RanBP2, that confer antagonizing and multi-modal mechanisms of kinesin-1 activation and regulation of mitochondrial motility. These findings open new venues towards the pharmacological harnessing of cooperative and competitive mechanisms regulating kinesins, RanBP2 or mitochondrial motility in disparate human disorders.

Mitochondrial respiratory efficiency is positively correlated with human sperm motility.

PubMed

Ferramosca, Alessandra; Provenzano, Sara Pinto; Coppola, Lamberto; Zara, Vincenzo

2012-04-01

To correlate sperm mitochondrial respiratory efficiency with variations in sperm motility and with sperm morphologic anomalies. Sperm mitochondrial respiratory activity was evaluated with a polarographic assay of oxygen consumption carried out in hypotonically-treated sperm cells. A possible relationship among sperm mitochondrial respiratory efficiency, sperm motility, and morphologic anomalies was investigated. Mitochondrial respiratory efficiency was positively correlated with sperm motility and negatively correlated with the percentage of immotile spermatozoa. Moreover, midpiece defects impaired mitochondrial functionality. Our data indicate that an increase in sperm motility requires a parallel increase in mitochondrial respiratory capacity, thereby supporting the fundamental role played by mitochondrial oxidative phosphorylation in sperm motility of normozoospermic subjects. These results are of physiopathological relevance because they suggest that disturbances of sperm mitochondrial function and of energy production could be responsible for asthenozoospermia. Copyright © 2012 Elsevier Inc. All rights reserved.

Mitochondrial motility and vascular smooth muscle proliferation.

PubMed

Chalmers, Susan; Saunter, Christopher; Wilson, Calum; Coats, Paul; Girkin, John M; McCarron, John G

2012-12-01

Mitochondria are widely described as being highly dynamic and adaptable organelles, and their movement is thought to be vital for cell function. Yet, in various native cells, including those of heart and smooth muscle, mitochondria are stationary and rigidly structured. The significance of the differences in mitochondrial behavior to the physiological function of cells is unclear and was studied in single myocytes and intact resistance-sized cerebral arteries. We hypothesized that mitochondrial dynamics is controlled by the proliferative status of the cells. High-speed fluorescence imaging of mitochondria in live vascular smooth muscle cells shows that the organelle undergoes significant reorganization as cells become proliferative. In nonproliferative cells, mitochondria are individual (≈ 2 μm by 0.5 μm), stationary, randomly dispersed, fixed structures. However, on entering the proliferative state, mitochondria take on a more diverse architecture and become small spheres, short rod-shaped structures, long filamentous entities, and networks. When cells proliferate, mitochondria also continuously move and change shape. In the intact pressurized resistance artery, mitochondria are largely immobile structures, except in a small number of cells in which motility occurred. When proliferation of smooth muscle was encouraged in the intact resistance artery, in organ culture, the majority of mitochondria became motile and the majority of smooth muscle cells contained moving mitochondria. Significantly, restriction of mitochondrial motility using the fission blocker mitochondrial division inhibitor prevented vascular smooth muscle proliferation in both single cells and the intact resistance artery. These results show that mitochondria are adaptable and exist in intact tissue as both stationary and highly dynamic entities. This mitochondrial plasticity is an essential mechanism for the development of smooth muscle proliferation and therefore presents a novel therapeutic target against vascular disease.

Cell-to-cell stimulation of movement in nonmotile mutants of Myxococcus

PubMed Central

Hodgkin, Jonathan; Kaiser, Dale

1977-01-01

A large number of nonmotile mutants of the gliding bacterium Myxococcus xanthus have been isolated and partly characterized. About [unk] of these mutants are conditional mutants of a novel kind: mutant cells become transiently motile after contact with nonmutant cells or with cells of a different mutant type. These “stimulatable” mutants fall into five phenotypic classes (types B, C, D, E, and F). Most mutants are nonstimulatable (type A) and never become motile, but type A cells (and wild-type cells) can stimulate cells of any of the other five types. Stimulatable mutants of different types are capable of stimulating each other. For example, in a mixture of B and C cells, both become motile. Linkage analysis using a generalized transducing phage has shown that each of types B, C, D, E, and F corresponds to a single distinct genetic locus. Type A mutants, by contrast, belong to at least 17 different loci. Stimulation depends on close apposition of interacting cells, because stimulation does not occur when contact between cells is prevented. It is possible that the stimulatable mutants are defective in components of the gliding mechanism that can be exchanged between cells. Alternatively, they may be defective in a system of cell communication controlling the coordinated cell movements observed in Myxococcus. Images PMID:16592422

Flagellar motility is a key determinant of the magnitude of the inflammasome response to Pseudomonas aeruginosa.

PubMed

Patankar, Yash R; Lovewell, Rustin R; Poynter, Matthew E; Jyot, Jeevan; Kazmierczak, Barbara I; Berwin, Brent

2013-06-01

We previously demonstrated that bacterial flagellar motility is a fundamental mechanism by which host phagocytes bind and ingest bacteria. Correspondingly, loss of bacterial motility, consistently observed in clinical isolates from chronic Pseudomonas aeruginosa infections, enables bacteria to evade association and ingestion of P. aeruginosa by phagocytes both in vitro and in vivo. Since bacterial interactions with the phagocyte cell surface are required for type three secretion system-dependent NLRC4 inflammasome activation by P. aeruginosa, we hypothesized that reduced bacterial association with phagocytes due to loss of bacterial motility, independent of flagellar expression, will lead to reduced inflammasome activation. Here we report that inflammasome activation is reduced in response to nonmotile P. aeruginosa. Nonmotile P. aeruginosa elicits reduced IL-1β production as well as caspase-1 activation by peritoneal macrophages and bone marrow-derived dendritic cells in vitro. Importantly, nonmotile P. aeruginosa also elicits reduced IL-1β levels in vivo in comparison to those elicited by wild-type P. aeruginosa. This is the first demonstration that loss of bacterial motility results in reduced inflammasome activation and antibacterial IL-1β host response. These results provide a critical insight into how the innate immune system responds to bacterial motility and, correspondingly, how pathogens have evolved mechanisms to evade the innate immune system.

Flagellar Motility Is a Key Determinant of the Magnitude of the Inflammasome Response to Pseudomonas aeruginosa

PubMed Central

Patankar, Yash R.; Lovewell, Rustin R.; Poynter, Matthew E.; Jyot, Jeevan; Kazmierczak, Barbara I.

2013-01-01

We previously demonstrated that bacterial flagellar motility is a fundamental mechanism by which host phagocytes bind and ingest bacteria. Correspondingly, loss of bacterial motility, consistently observed in clinical isolates from chronic Pseudomonas aeruginosa infections, enables bacteria to evade association and ingestion of P. aeruginosa by phagocytes both in vitro and in vivo. Since bacterial interactions with the phagocyte cell surface are required for type three secretion system-dependent NLRC4 inflammasome activation by P. aeruginosa, we hypothesized that reduced bacterial association with phagocytes due to loss of bacterial motility, independent of flagellar expression, will lead to reduced inflammasome activation. Here we report that inflammasome activation is reduced in response to nonmotile P. aeruginosa. Nonmotile P. aeruginosa elicits reduced IL-1β production as well as caspase-1 activation by peritoneal macrophages and bone marrow-derived dendritic cells in vitro. Importantly, nonmotile P. aeruginosa also elicits reduced IL-1β levels in vivo in comparison to those elicited by wild-type P. aeruginosa. This is the first demonstration that loss of bacterial motility results in reduced inflammasome activation and antibacterial IL-1β host response. These results provide a critical insight into how the innate immune system responds to bacterial motility and, correspondingly, how pathogens have evolved mechanisms to evade the innate immune system. PMID:23529619

Isolation, characterization, and complementation of a motility mutant of Spiroplasma citri.

PubMed Central

Jacob, C; Nouzières, F; Duret, S; Bové, J M; Renaudin, J

1997-01-01

The helical mollicute Spiroplasma citri, when growing on low-agar medium, forms fuzzy colonies with occasional surrounding satellite colonies due to the ability of the spiroplasmal cells to move through the agar matrix. In liquid medium, these helical organisms flex, twist, and rotate rapidly. By using Tn4001 insertion mutagenesis, a motility mutant was isolated on the basis of its nondiffuse, sharp-edged colonies. Dark-field microscopy observations revealed that the organism flexed at a low frequency and had lost the ability to rotate about the helix axis. In this mutant, the transposon was shown to be inserted into an open reading frame encoding a putative polypeptide of 409 amino acids for which no significant homology with known proteins was found. The corresponding gene, named scm1, was recovered from the wild-type strain and introduced into the motility mutant by using the S. citri oriC plasmid pBOT1 as the vector. The appearance of fuzzy colonies and the observation that spiroplasma cells displayed rotatory and flexional movements showed the motile phenotype to be restored in the spiroplasmal transformants. The functional complementation of the motility mutant proves the scm1 gene product to be involved in the motility mechanism of S. citri. PMID:9244268

Dishevelled links basal body docking and orientation in ciliated epithelial cells

PubMed Central

Vladar, Eszter K.; Axelrod, Jeffrey D.

2014-01-01

Some epithelia contain cells with multiple, motile cilia that beat in a concerted fashion. New tools and experimental systems have facilitated molecular studies of cilium biogenesis and of the coordinated planar polarization of cilia that leads to their concerted motility. Recent, elegant work by Park and colleagues, using embryonic frog epidermis, demonstrates that Dishevelled (Dvl), a key regulator of both the Wnt/β-catenin and Planar Cell Polarity (PCP) pathways, controls both the docking and planar polarization of ciliary basal bodies. PMID:18819800

THE ROLE OF THREE CYTOPLASMIC FIBERS IN BHK-21 CELL MOTILITY

PubMed Central

Goldman, Robert D.

1971-01-01

Microtubule breakdown in the presence of 5 or 40 µg/ml of colchicine is observed in BHK-21/C13 fibroblast-like cells. Several morphological and physiological effects are noted in the absence of microtubules: (a) the cells transform from fibroblast-like to epithelial-like cells; (b) the normal pattern of intracellular birefringence changes and a juxtanuclear cap of birefringent filaments is formed; (c) time-lapse cinematography demonstrates that cell locomotion is inhibited in colchicine-treated cells, even though membrane ruffling persists. The results are discussed in terms of the specific roles of microtubules in cultured cell motility and possible functional relationships of the three types of cytoplasmic fibers seen in BHK-21 cells. PMID:4942774

Tyrosine kinases, drugs, and Shigella flexneri dissemination.

PubMed

Dragoi, Ana-Maria; Agaisse, Hervé

2014-01-01

Shigella flexneri is an enteropathogenic bacterium responsible for approximately 100 million cases of severe dysentery each year. S. flexneri colonization of the human colonic epithelium is supported by direct spread from cell to cell, which relies on actin-based motility. We have recently uncovered that, in intestinal epithelial cells, S. flexneri actin-based motility is regulated by the Bruton's tyrosine kinase (Btk). Consequently, treatment with Ibrutinib, a specific Btk inhibitor currently used in the treatment of B-cell malignancies, effectively impaired S. flexneri spread from cell to cell. Thus, therapeutic intervention capitalizing on drugs interfering with host factors supporting the infection process may represent an effective alternative to treatments with antimicrobial compounds.

Isolation, Culture, and Motility Measurements of Epidermal Melanocytes from GFP-Expressing Reporter Mice.

PubMed

Dagnino, Lina; Crawford, Melissa

2018-03-27

In this article, we provide a method to isolate primary epidermal melanocytes from reporter mice, which also allow targeted gene inactivation. The mice from which these cells are isolated are bred into a Rosa26 mT/mG reporter background, which results in GFP expression in the targeted melanocytic cell population. These cells are isolated and cultured to >95% purity. The cells can be used for gene expression studies, clonogenic experiments, and biological assays, such as capacity for migration. Melanocytes are slow moving cells, and we also provide a method to measure motility using individual cell tracking and data analysis.

Cellular mechanics and motility

NASA Astrophysics Data System (ADS)

Hénon, Sylvie; Sykes, Cécile

2015-10-01

The term motility defines the movement of a living organism. One widely known example is the motility of sperm cells, or the one of flagellar bacteria. The propulsive element of such organisms is a cilium(or flagellum) that beats. Although cells in our tissues do not have a flagellum in general, they are still able to move, as we will discover in this chapter. In fact, in both cases of movement, with or without a flagellum, cell motility is due to a dynamic re-arrangement of polymers inside the cell. Let us first have a closer look at the propulsion mechanism in the case of a flagellum or a cilium, which is the best known, but also the simplest, and which will help us to define the hydrodynamic general conditions of cell movement. A flagellum is sustained by cellular polymers arranged in semi-flexible bundles and flagellar beating generates cell displacement. These polymers or filaments are part of the cellular skeleton, or "cytoskeleton", which is, in this case, external to the cellular main body of the organism. In fact, bacteria move in a hydrodynamic regime in which viscosity dominates over inertia. The system is thus in a hydrodynamic regime of low Reynolds number (Box 5.1), which is nearly exclusively the case in all cell movements. Bacteria and their propulsion mode by flagella beating are our unicellular ancestors 3.5 billion years ago. Since then, we have evolved to form pluricellular organisms. However, to keep the ability of displacement, to heal our wounds for example, our cells lost their flagellum, since it was not optimal in a dense cell environment: cells are too close to each other to leave enough space for the flagella to accomplish propulsion. The cytoskeleton thus developed inside the cell body to ensure cell shape changes and movement, and also mechanical strength within a tissue. The cytoskeleton of our cells, like the polymers or filaments that sustain the flagellum, is also composed of semi-flexible filaments arranged in bundles, and also in cross-linked or branched networks. It is a highly dynamical system in which filaments are able to elongate or slide one on the other with the contribution of very active cellular proteins like molecular motors. The versatile properties of this cytoskeleton ensure the diversity of mechanical behaviors to explain cell rigidity as well as cell motility.

Autoinducer 2 controls biofilm formation in Escherichia coli through a novel motility quorum-sensing regulator (MqsR, B3022).

PubMed

González Barrios, Andrés F; Zuo, Rongjun; Hashimoto, Yoshifumi; Yang, Li; Bentley, William E; Wood, Thomas K

2006-01-01

The cross-species bacterial communication signal autoinducer 2 (AI-2), produced by the purified enzymes Pfs (nucleosidase) and LuxS (terminal synthase) from S-adenosylhomocysteine, directly increased Escherichia coli biofilm mass 30-fold. Continuous-flow cells coupled with confocal microscopy corroborated these results by showing the addition of AI-2 significantly increased both biofilm mass and thickness and reduced the interstitial space between microcolonies. As expected, the addition of AI-2 to cells which lack the ability to transport AI-2 (lsr null mutant) failed to stimulate biofilm formation. Since the addition of AI-2 increased cell motility through enhanced transcription of five motility genes, we propose that AI-2 stimulates biofilm formation and alters its architecture by stimulating flagellar motion and motility. It was also found that the uncharacterized protein B3022 regulates this AI-2-mediated motility and biofilm phenotype through the two-component motility regulatory system QseBC. Deletion of b3022 abolished motility, which was restored by expressing b3022 in trans. Deletion of b3022 also decreased biofilm formation significantly, relative to the wild-type strain in three media (46 to 74%) in 96-well plates, as well as decreased biomass (8-fold) and substratum coverage (19-fold) in continuous-flow cells with minimal medium (growth rate not altered and biofilm restored by expressing b3022 in trans). Deleting b3022 changed the wild-type biofilm architecture from a thick (54-mum) complex structure to one that contained only a few microcolonies. B3022 positively regulates expression of qseBC, flhD, fliA, and motA, since deleting b3022 decreased their transcription by 61-, 25-, 2.4-, and 18-fold, respectively. Transcriptome analysis also revealed that B3022 induces crl (26-fold) and flhCD (8- to 27-fold). Adding AI-2 (6.4 muM) increased biofilm formation of wild-type K-12 MG1655 but not that of the isogenic b3022, qseBC, fliA, and motA mutants. Adding AI-2 also increased motA transcription for the wild-type strain but did not stimulate motA transcription for the b3022 and qseB mutants. Together, these results indicate AI-2 induces biofilm formation in E. coli through B3022, which then regulates QseBC and motility; hence, b3022 has been renamed the motility quorum-sensing regulator gene (the mqsR gene).

Curcumin Attenuates β-catenin Signaling in Prostate Cancer Cells through Activation of Protein Kinase D1

PubMed Central

Sundram, Vasudha; Chauhan, Subhash C.; Ebeling, Mara; Jaggi, Meena

2012-01-01

Prostate cancer is the most commonly diagnosed cancer affecting 1 in 6 males in the US. Understanding the molecular basis of prostate cancer progression can serve as a tool for early diagnosis and development of novel treatment strategies for this disease. Protein Kinase D1 (PKD1) is a multifunctional kinase that is highly expressed in normal prostate. The decreased expression of PKD1 has been associated with the progression of prostate cancer. Therefore, synthetic or natural products that regulate this signaling pathway can serve as novel therapeutic modalities for prostate cancer prevention and treatment. Curcumin, the active ingredient of turmeric, has shown anti-cancer properties via modulation of a number of different molecular pathways. Herein, we have demonstrated that curcumin activates PKD1, resulting in changes in β-catenin signaling by inhibiting nuclear β-catenin transcription activity and enhancing the levels of membrane β-catenin in prostate cancer cells. Modulation of these cellular events by curcumin correlated with decreased cell proliferation, colony formation and cell motility and enhanced cell-cell aggregation in prostate cancer cells. In addition, we have also revealed that inhibition of cell motility by curcumin is mediated by decreasing the levels of active cofilin, a downstream target of PKD1. The potent anti-cancer effects of curcumin in vitro were also reflected in a prostate cancer xenograft mouse model. The in vivo inhibition of tumor growth also correlated with enhanced membrane localization of β-catenin. Overall, our findings herein have revealed a novel molecular mechanism of curcumin action via the activation of PKD1 in prostate cancer cells. PMID:22523587

Analysis of the Borrelia burgdorferi Cyclic-di-GMP-Binding Protein PlzA Reveals a Role in Motility and Virulence ▿

PubMed Central

Pitzer, Joshua E.; Sultan, Syed Z.; Hayakawa, Yoshihiro; Hobbs, Gerry; Miller, Michael R.; Motaleb, Md A.

2011-01-01

The cyclic-dimeric-GMP (c-di-GMP)-binding protein PilZ has been implicated in bacterial motility and pathogenesis. Although BB0733 (PlzA), the only PilZ domain-containing protein in Borrelia burgdorferi, was reported to bind c-di-GMP, neither its role in motility or virulence nor it's affinity for c-di-GMP has been reported. We determined that PlzA specifically binds c-di-GMP with high affinity (dissociation constant [Kd], 1.25 μM), consistent with Kd values reported for c-di-GMP-binding proteins from other bacteria. Inactivation of the monocistronically transcribed plzA resulted in an opaque/solid colony morphology, whereas the wild-type colonies were translucent. While the swimming pattern of mutant cells appeared normal, on swarm plates, mutant cells exhibited a significantly reduced swarm diameter, demonstrating a role of plzA in motility. Furthermore, the plzA mutant cells were significantly less infectious in experimental mice (as determined by 50% infectious dose [ID50]) relative to wild-type spirochetes. The mutant also had survival rates in fed ticks lower than those of the wild type. Consequently, plzA mutant cells failed to complete the mouse-tick-mouse infection cycle, indicating plzA is essential for the enzootic life cycle of B. burgdorferi. All of these defects were corrected when the mutant was complemented in cis. We propose that failure of plzA mutant cells to infect mice was due to altered motility; however, the possibility that an unidentified factor(s) contributed to interruption of the B. burgdorferi enzootic life cycle cannot yet be excluded. PMID:21357718

Hypercholesterolemia Impaired Sperm Functionality in Rabbits

PubMed Central

Monclus, Maria A.; Cabrillana, Maria E.; Clementi, Marisa A.; Espínola, Leandro S.; Cid Barría, Jose L.; Vincenti, Amanda E.; Santi, Analia G.; Fornés, Miguel W.

2010-01-01

Hypercholesterolemia represents a high risk factor for frequent diseases and it has also been associated with poor semen quality that may lead to male infertility. The aim of this study was to analyze semen and sperm function in diet-induced hypercholesterolemic rabbits. Twelve adult White New Zealand male rabbits were fed ad libitum a control diet or a diet supplemented with 0.05% cholesterol. Rabbits under cholesterol-enriched diet significantly increased total cholesterol level in the serum. Semen examination revealed a significant reduction in semen volume and sperm motility in hypercholesterolemic rabbits (HCR). Sperm cell morphology was seriously affected, displaying primarily a “folded head”-head fold along the major axe-, and the presence of cytoplasmic droplet on sperm flagellum. Cholesterol was particularly increased in acrosomal region when detected by filipin probe. The rise in cholesterol concentration in sperm cells was determined quantitatively by Gas chromatographic-mass spectrometric analyses. We also found a reduction of protein tyrosine phosphorylation in sperm incubated under capacitating conditions from HCR. Interestingly, the addition of Protein Kinase A pathway activators -dibutyryl-cyclic AMP and iso-butylmethylxanthine- to the medium restored sperm capacitation. Finally, it was also reported a significant decrease in the percentage of reacted sperm in the presence of progesterone. In conclusion, our data showed that diet-induced hypercholesterolemia adversely affects semen quality and sperm motility, capacitation and acrosomal reaction in rabbits; probably due to an increase in cellular cholesterol content that alters membrane related events. PMID:20976152

Covalent binding of nanoliposomes to the surface of magnetotactic bacteria for the synthesis of self-propelled therapeutic agents.

PubMed

Taherkhani, Samira; Mohammadi, Mahmood; Daoud, Jamal; Martel, Sylvain; Tabrizian, Maryam

2014-05-27

The targeted and effective delivery of therapeutic agents remains an unmet goal in the field of controlled release systems. Magnetococcus marinus MC-1 magnetotactic bacteria (MTB) are investigated as potential therapeutic carriers. By combining directional magnetotaxis-microaerophilic control of these self-propelled agents, a larger amount of therapeutics can be delivered surpassing the diffusion limits of large drug molecules toward hard-to-treat hypoxic regions in solid tumors. The potential benefits of these carriers emphasize the need to develop an adequate method to attach therapeutic cargos, such as drug-loaded nanoliposomes, without substantially affecting the cell's ability to act as delivery agents. In this study, we report on a strategy for the attachment of liposomes to MTB (MTB-LP) through carbodiimide chemistry. The attachment efficacy, motility, and magnetic response of the MTB-LP were investigated. Results confirm that a substantial number of nanoliposomes (∼70) are efficiently linked with MTB without compromising functionality and motility. Cytotoxicity assays using three different cell types (J774, NIH/3T3, and Colo205) reveal that liposomal attachments to MTB formulation improve the biocompatibility of MTB, whereas attachment does not interfere with liposomal uptake.

Low-cost motility tracking system (LOCOMOTIS) for time-lapse microscopy applications and cell visualisation.

PubMed

Lynch, Adam E; Triajianto, Junian; Routledge, Edwin

2014-01-01

Direct visualisation of cells for the purpose of studying their motility has typically required expensive microscopy equipment. However, recent advances in digital sensors mean that it is now possible to image cells for a fraction of the price of a standard microscope. Along with low-cost imaging there has also been a large increase in the availability of high quality, open-source analysis programs. In this study we describe the development and performance of an expandable cell motility system employing inexpensive, commercially available digital USB microscopes to image various cell types using time-lapse and perform tracking assays in proof-of-concept experiments. With this system we were able to measure and record three separate assays simultaneously on one personal computer using identical microscopes, and obtained tracking results comparable in quality to those from other studies that used standard, more expensive, equipment. The microscopes used in our system were capable of a maximum magnification of 413.6×. Although resolution was lower than that of a standard inverted microscope we found this difference to be indistinguishable at the magnification chosen for cell tracking experiments (206.8×). In preliminary cell culture experiments using our system, velocities (mean µm/min ± SE) of 0.81 ± 0.01 (Biomphalaria glabrata hemocytes on uncoated plates), 1.17 ± 0.004 (MDA-MB-231 breast cancer cells), 1.24 ± 0.006 (SC5 mouse Sertoli cells) and 2.21 ± 0.01 (B. glabrata hemocytes on Poly-L-Lysine coated plates), were measured and are consistent with previous reports. We believe that this system, coupled with open-source analysis software, demonstrates that higher throughput time-lapse imaging of cells for the purpose of studying motility can be an affordable option for all researchers.

Low-Cost Motility Tracking System (LOCOMOTIS) for Time-Lapse Microscopy Applications and Cell Visualisation

PubMed Central

Lynch, Adam E.; Triajianto, Junian; Routledge, Edwin

2014-01-01

Direct visualisation of cells for the purpose of studying their motility has typically required expensive microscopy equipment. However, recent advances in digital sensors mean that it is now possible to image cells for a fraction of the price of a standard microscope. Along with low-cost imaging there has also been a large increase in the availability of high quality, open-source analysis programs. In this study we describe the development and performance of an expandable cell motility system employing inexpensive, commercially available digital USB microscopes to image various cell types using time-lapse and perform tracking assays in proof-of-concept experiments. With this system we were able to measure and record three separate assays simultaneously on one personal computer using identical microscopes, and obtained tracking results comparable in quality to those from other studies that used standard, more expensive, equipment. The microscopes used in our system were capable of a maximum magnification of 413.6×. Although resolution was lower than that of a standard inverted microscope we found this difference to be indistinguishable at the magnification chosen for cell tracking experiments (206.8×). In preliminary cell culture experiments using our system, velocities (mean µm/min ± SE) of 0.81±0.01 (Biomphalaria glabrata hemocytes on uncoated plates), 1.17±0.004 (MDA-MB-231 breast cancer cells), 1.24±0.006 (SC5 mouse Sertoli cells) and 2.21±0.01 (B. glabrata hemocytes on Poly-L-Lysine coated plates), were measured and are consistent with previous reports. We believe that this system, coupled with open-source analysis software, demonstrates that higher throughput time-lapse imaging of cells for the purpose of studying motility can be an affordable option for all researchers. PMID:25121722

Directional control of lamellipodia extension by constraining cell shape and orienting cell tractional forces

NASA Technical Reports Server (NTRS)

Parker, Kevin Kit; Brock, Amy Lepre; Brangwynne, Cliff; Mannix, Robert J.; Wang, Ning; Ostuni, Emanuele; Geisse, Nicholas A.; Adams, Josephine C.; Whitesides, George M.; Ingber, Donald E.

2002-01-01

Directed cell migration is critical for tissue morphogenesis and wound healing, but the mechanism of directional control is poorly understood. Here we show that the direction in which cells extend their leading edge can be controlled by constraining cell shape using micrometer-sized extracellular matrix (ECM) islands. When cultured on square ECM islands in the presence of motility factors, cells preferentially extended lamellipodia, filopodia, and microspikes from their corners. Square cells reoriented their stress fibers and focal adhesions so that tractional forces were concentrated in these corner regions. When cell tension was dissipated, lamellipodia extension ceased. Mechanical interactions between cells and ECM that modulate cytoskeletal tension may therefore play a key role in the control of directional cell motility.

Differential effects on cell motility, embryonic stem cell self-renewal and senescence by diverse Src kinase family inhibitors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tamm, Christoffer, E-mail: christoffer.tamm@imbim.uu.se; Galito, Sara Pijuan, E-mail: sara.pijuan@imbim.uu.se; Anneren, Cecilia, E-mail: cecilia.anneren@imbim.uu.se

2012-02-15

The Src family of non-receptor tyrosine kinases (SFKs) has been shown to play an intricate role in embryonic stem (ES) cell maintenance. In the present study we have focused on the underlying molecular mechanisms responsible for the vastly different effects induced by various commonly used SFK inhibitors. We show that several diverse cell types, including fibroblasts completely lacking SFKs, cannot undergo mitosis in response to SU6656 and that this is caused by an unselective inhibition of Aurora kinases. In contrast, PP2 and PD173952 block motility immediately upon exposure and forces cells to grow in dense colonies. The subsequent halt inmore » proliferation of fibroblast and epithelial cells in the center of the colonies approximately 24 h post-treatment appears to be caused by cell-to-cell contact inhibition rather than a direct effect of SFK kinase inhibition. Interestingly, in addition to generating more homogenous and dense ES cell cultures, without any diverse effect on proliferation, PP2 and PD173652 also promote ES cell self-renewal by reducing the small amount of spontaneous differentiation typically observed under standard ES cell culture conditions. These effects could not be mirrored by the use of Gleevec, a potent inhibitor of c-Abl and PDGFR kinases that are also inhibited by PP2. -- Highlights: Black-Right-Pointing-Pointer SFK inhibitor SU6656 induces senescence in mouse ES cells. Black-Right-Pointing-Pointer SU6656 inhibits mitosis in a SFK-independent manner via cross-selectivity for Aurora kinases. Black-Right-Pointing-Pointer SFK inhibitor PP2 impairs cell motility in various cell lines, including mouse ES cells. Black-Right-Pointing-Pointer Ensuing impeded motility, PP2 inhibits proliferation of various cells lines except for mouse ES cells. Black-Right-Pointing-Pointer SFK inhibitors PP2 and PD173952 impede spontaneous differentiation in standard mouse ES culture maintenance.« less

hemingway is required for sperm flagella assembly and ciliary motility in Drosophila.

PubMed

Soulavie, Fabien; Piepenbrock, David; Thomas, Joëlle; Vieillard, Jennifer; Duteyrat, Jean-Luc; Cortier, Elisabeth; Laurençon, Anne; Göpfert, Martin C; Durand, Bénédicte

2014-04-01

Cilia play major functions in physiology and development, and ciliary dysfunctions are responsible for several diseases in humans called ciliopathies. Cilia motility is required for cell and fluid propulsion in organisms. In humans, cilia motility deficiencies lead to primary ciliary dyskinesia, with upper-airways recurrent infections, left-right asymmetry perturbations, and fertility defects. In Drosophila, we identified hemingway (hmw) as a novel component required for motile cilia function. hmw encodes a 604-amino acid protein characterized by a highly conserved coiled-coil domain also found in the human orthologue, KIAA1430. We show that HMW is conserved in species with motile cilia and that, in Drosophila, hmw is expressed in ciliated sensory neurons and spermatozoa. We created hmw-knockout flies and found that they are hearing impaired and male sterile. hmw is implicated in the motility of ciliated auditory sensory neurons and, in the testis, is required for elongation and maintenance of sperm flagella. Because HMW is absent from mature flagella, we propose that HMW is not a structural component of the motile axoneme but is required for proper acquisition of motile properties. This identifies HMW as a novel, evolutionarily conserved component necessary for motile cilium function and flagella assembly.

Stem cell motility enables a density-dependent rate of fate commitment during scaled resizing of adult organs

NASA Astrophysics Data System (ADS)

Du, Xinxin; O'Brien, Lucy; Riedel-Kruse, Ingmar

Many adult organs grow or shrink to accommodate fluctuating levels of physiological demand. Specifically, the intestine of the fruit fly (the midgut) expands four-fold in the number of mature cells and, proportionally, the number of stem cells when the fly eats. However, the cellular behaviors that give rise to this stem scaling are not well-understood. Here we present a biophysical model of the adult fly midgut. A set of differential equations can recapitulate the physiological kinetics of cells during midgut growth and shrinkage as long as the rate of stem cell fate commitment depends on the stem cell number density in the tissue. To elucidate the source of this dependence, we model the tissue in a 2D simulation with soft spheres, where stem cells choose fate commitment through Delta-Notch pathway interactions with other stem cells, a known process in fly midguts. We find that as long as stem cells exhibit a large enough amplitude of random motion through the tissue (`stem cell motility'), and explore a large enough `territory' in their lifetime, stem cell scaling can occur. These model observations are confirmed through in vivo live-imaging, where we indeed see that stem cells are motile in the fly midgut.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Geng, Tao; Smallwood, Chuck R.; Bredeweg, Erin L.

Modern live-cell imaging approaches permit real-time visualization of biological processes, yet limitations exist for unicellular organism isolation, culturing and long-term imaging that preclude fully understanding how cells sense and respond to environmental perturbations and the link between single-cell variability and whole-population dynamics. Here we present a microfluidic platform that provides fine control over the local environment with the capacity to replace media components at any experimental time point, and provides both perfused and compartmentalized cultivation conditions depending on the valve configuration. The functionality and flexibility of the platform were validated using both bacteria and yeast having different sizes, motility andmore » growth media. The demonstrated ability to track the growth and dynamics of both motile and non-motile prokaryotic and eukaryotic organisms emphasizes the versatility of the devices, which with further scale-up should enable studies in bioenergy and environmental research.« less

Effects of environment factors on initiation of sperm motility in sea cucumber Apostichopus japonicus (Selenka)

NASA Astrophysics Data System (ADS)

Yu, Li; Shao, Mingyu; Bao, Zhenmin; Hu, Jingjie; Zhang, Zhifeng

2011-06-01

Sperm of sea cucumber Apostichopus japonicus (Selenka) were quiescent in electrolyte NaCl solution and artificial seawater (ASW) and nonelectrolyte glucose and mannitol solutions when the osmolality was less than 200 mOsm kg-1. The sperm started to be motile as a result of increased osmolality, indicating an osmolality-dependent initiation of sperm motility in sea cucumber. After a brief incubation in hypotonic NaCl and glucose solutions with osmolalities of 200 and 400 mOsm kg-1, sperm lost partial motile ability. Sperm became immobilized when pH was 6.0 in NaCl, glucose and mannitol solutions, suggesting that an H+ release is involved in sperm activation. The decreased pH had no effect on the percentage of motile sperm in ASW, whereas it delayed the time period to reach the maximum motility (motilitymax). Extracellular Ca2+ in electrolyte solutions was not essential for motility stimulation but shortened the time of reaching motilitymax. When Ca2+ was mixed in nonelectrolyte solutions the sperm motility was completely suppressed. The K+ channel blocker, quinine, suppressed the sperm motility in electrolyte solution, showing a possible involvement of K+ transport in the process. High K+ concentration did not affect the sperm motility in NaCl solution, but decreased it in ASW and almost entirely suppressed it in nonelectrolyte solutions. The different effects of pH and K+ in ASW and NaCl solution indicate that external ions may also regulate sperm motility.

By activating matrix metalloproteinase-7, shear stress promotes chondrosarcoma cell motility, invasion and lung colonization.

PubMed

Guan, Pei-Pei; Yu, Xin; Guo, Jian-Jun; Wang, Yue; Wang, Tao; Li, Jia-Yi; Konstantopoulos, Konstantinos; Wang, Zhan-You; Wang, Pu

2015-04-20

Interstitial fluid flow and associated shear stress are relevant mechanical signals in cartilage and bone (patho)physiology. However, their effects on chondrosarcoma cell motility, invasion and metastasis have yet to be delineated. Using human SW1353, HS.819.T and CH2879 chondrosarcoma cell lines as model systems, we found that fluid shear stress induces the accumulation of cyclic AMP (cAMP) and interleukin-1β (IL-1β), which in turn markedly enhance chondrosarcoma cell motility and invasion via the induction of matrix metalloproteinase-7 (MMP-7). Specifically, shear-induced cAMP and IL-1β activate PI3-K, ERK1/2 and p38 signaling pathways, which lead to the synthesis of MMP-7 via transactivating NF-κB and c-Jun in human chondrosarcoma cells. Importantly, MMP-7 upregulation in response to shear stress exposure has the ability to promote lung colonization of chondrosarcomas in vivo. These findings offer a better understanding of the mechanisms underlying MMP-7 activation in shear-stimulated chondrosarcoma cells, and provide insights on designing new therapeutic strategies to interfere with chondrosarcoma invasion and metastasis.

Ecotoxicity evaluation of a liquid detergent using the automatic biotest ECOTOX.

PubMed

Azizullah, Azizullah; Richter, Peter; Ullah, Waheed; Ali, Imran; Häder, Donat-Peter

2013-08-01

Synthetic detergents are common pollutants reaching aquatic environments in different ways after usage at homes, institutions and industries. In this study a liquid detergent, used for dish washing, was evaluated for its toxicity during long- and short-term tests using the automatic biotest ECOTOX. Different parameters of Euglena gracilis like motility, swimming velocity, gravitactic orientation, cell compactness and cell growth were used as end points. In short-term experiments, the maximum adverse effects on motility, velocity, cell shape and gravitaxis were observed after 1 h of exposure. With further increase in exposure time to the detergent a slight recovery of these parameters was observed. In long-term experiments, the detergent caused severe disturbances to E. gracilis. Motility, cell growth and cell compactness (shape) with EC50 values of 0.064, 0.18 and 2.05 %, respectively, were found as the most sensitive parameters to detergent stress. There was a slight positive effect on gravitactic orientation at the lowest two concentrations; at higher concentrations of the detergent cells orientation was highly impaired giving EC50 values of 1.75 and 2.52 % for upward swimming and r-value, respectively.

Assaying Wnt5A-mediated Invasion in Melanoma Cells

PubMed Central

O'Connell, Michael P.; French, Amanda D.; Leotlela, Poloko D.; Weeraratna, Ashani T.

2009-01-01

Wnt5A has been implicated in melanoma metastasis, and the progression of other cancers including pancreatic, gastric, prostate and lung cancers. Assays to test motility and invasion include both in vivo assays, and in vitro assays. The former assays include the use of tail vein or footpad injections of metastatic cells, and are often laborious and expensive. In vitro invasion assays provide quick readouts that can help to establish conditions that either activate or inhibit melanoma cell motility, and to assess whether the conditions in question are worth translating into an in vivo model. Here we describe two standard methods for assaying motility and invasion in vitro including wound healing assays and Matrigel invasion assays (Boyden chamber assays). In addition, we and several other laboratories have previously shown that melanoma cells require MMP-2 for their invasion, and have recently shown that Wnt5A treatment can increase the levels of this enzyme in melanoma cells, as demonstrated by gelatin zymography. The use of these techniques can help to assess the migratory capacity of melanoma cells in response to Wnt treatment. PMID:19099260

Mechanics and polarity in cell motility

NASA Astrophysics Data System (ADS)

Ambrosi, D.; Zanzottera, A.

2016-09-01

The motility of a fish keratocyte on a flat substrate exhibits two distinct regimes: the non-migrating and the migrating one. In both configurations the shape is fixed in time and, when the cell is moving, the velocity is constant in magnitude and direction. Transition from a stable configuration to the other one can be produced by a mechanical or chemotactic perturbation. In order to point out the mechanical nature of such a bistable behaviour, we focus on the actin dynamics inside the cell using a minimal mathematical model. While the protein diffusion, recruitment and segregation govern the polarization process, we show that the free actin mass balance, driven by diffusion, and the polymerized actin retrograde flow, regulated by the active stress, are sufficient ingredients to account for the motile bistability. The length and velocity of the cell are predicted on the basis of the parameters of the substrate and of the cell itself. The key physical ingredient of the theory is the exchange among actin phases at the edges of the cell, that plays a central role both in kinematics and in dynamics.

Active motility in bimodular bacterial aggregates

NASA Astrophysics Data System (ADS)

Zeng, Yu; Liu, Bin

2017-11-01

Dispersal capability is essential for microorganisms to achieve long-distance translocation, thus crucial for their abundance in various environments. In general, active dispersals are attributed to the movements of self-powered planktonic cells, while sessile cells that live a colonial life often disperse passively through flow entrainments. Here, we report another means of active dispersal employed by aggregates of sessile cells. The spherical rosette colonies of the bacterium Caulobacter crescentus are aggregates of sessile stalked cells, of which a small proportion undergo cell division, grow active flagella and effect whole-rosette motility. We show that these rosettes actively disperse both in bulk water and near the solid-liquid interface. In particular, the proximity of a self-powered rosette to the solid surface promotes a rolling movement, leading to its persistent transportation along the solid boundary. The active dispersal of these rosettes demonstrated a novel mode of colonial transportation that is based on the division of labor between sessile and motile cells. The authors thank the support of National Science Foundation CREST: Center for Cellular and Biomolecular Machines at UC Merced (NSF-HRD-1547848).

Metformin Treatment Inhibits Motility and Invasion of Glioblastoma Cancer Cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Al Hassan, Marwa; Fakhoury, Isabelle; El Masri, Zeinab

Glioblastoma multiforme (GBM) is one of the most common and deadliest cancers of the central nervous system (CNS). GBMs high ability to infiltrate healthy brain tissues makes it difficult to remove surgically and account for its fatal outcomes. To improve the chances of survival, it is critical to screen for GBM-targeted anticancer agents with anti-invasive and antimigratory potential. Metformin, a commonly used drug for the treatment of diabetes, has recently emerged as a promising anticancer molecule. This prompted us, to investigate the anticancer potential of metformin against GBMs, specifically its effects on cell motility and invasion. The results show amore » significant decrease in the survival of SF268 cancer cells in response to treatment with metformin. Furthermore, metformin’s efficiency in inhibiting 2D cell motility and cell invasion in addition to increasing cellular adhesion was also demonstrated in SF268 and U87 cells. Finally, AKT inactivation by downregulation of the phosphorylation level upon metformin treatment was also evidenced. In conclusion, this study provides insights into the anti-invasive antimetastatic potential of metformin as well as its underlying mechanism of action.« less

Metformin Treatment Inhibits Motility and Invasion of Glioblastoma Cancer Cells

DOE PAGES

Al Hassan, Marwa; Fakhoury, Isabelle; El Masri, Zeinab; ...

2018-06-26

Glioblastoma multiforme (GBM) is one of the most common and deadliest cancers of the central nervous system (CNS). GBMs high ability to infiltrate healthy brain tissues makes it difficult to remove surgically and account for its fatal outcomes. To improve the chances of survival, it is critical to screen for GBM-targeted anticancer agents with anti-invasive and antimigratory potential. Metformin, a commonly used drug for the treatment of diabetes, has recently emerged as a promising anticancer molecule. This prompted us, to investigate the anticancer potential of metformin against GBMs, specifically its effects on cell motility and invasion. The results show amore » significant decrease in the survival of SF268 cancer cells in response to treatment with metformin. Furthermore, metformin’s efficiency in inhibiting 2D cell motility and cell invasion in addition to increasing cellular adhesion was also demonstrated in SF268 and U87 cells. Finally, AKT inactivation by downregulation of the phosphorylation level upon metformin treatment was also evidenced. In conclusion, this study provides insights into the anti-invasive antimetastatic potential of metformin as well as its underlying mechanism of action.« less

A common surfactant used in food packaging found to be toxic for reproduction in mammals.

PubMed

Nerin, Cristina; Canellas, Elena; Vera, Paula; Garcia-Calvo, Estefanía; Luque-Garcia, José Luis; Cámara, Carmen; Ausejo, Raquel; Miguel, Joaquín; Mendoza, Noelia

2018-03-01

Migration from a multilayer plastic material intended for food contact showed that 2,4,7,9-tetramethyl-5-decyne-4,7-diol mixture (surfynol), used as a surfactant in the adhesive employed to build the multilayer, was transferred to water and other food simulants in contact with the plastic. When these multilayer plastics were used for containing seminal doses for artificial insemination, it was found that fertility was seriously damaged in terms of motility, acrosome integrity, mitochondrial activity and penetration capacity in the cells, thus affecting male fertility. Quantitative proteomic analysis of exposed germinal cells demonstrated the inhibition of key proteins involved in the fertilization capacity by affecting the cytoskeleton, sperm motility, the energy machinery and sperm defense mechanisms against oxidation, therefore confirming the surfactant-induced male infertility. These results open up new and interesting perspectives for the study of reprotoxicity caused by different chemicals common in our daily lives. This paper demonstrates the toxicity for reproduction of a common surfactant used in food packaging and the scientific reasons why the sperm loses reproductive capacity in presence of this chemical. So, the surfactant affects the male fertility. The surfactant is present in many adhesives used either for building multilayer materials or to glue paper and plastic in food packaging. This is the first time that reprotoxicity is demonstrated for this compound. According to the theoretical approach Threshold of Toxicological Concern (TTC) the compound is highly toxic but experimental data did not exist so far. The study described in this paper and the results obtained open a door to further research in which male infertility caused by chemicals could be demonstrated. Copyright © 2018. Published by Elsevier Ltd.

Polymorphonuclear cell motility, ankylosing spondylitis, and HLA B27.

PubMed Central

Pease, C T; Fordham, J N; Currey, H L

1984-01-01

Polymorphonuclear leucocyte (PMN) function was studied in 29 subjects with ankylosing spondylitis (AS). Of these, 20 were HLA B27+ve and 9 B27-ve. There were 30 controls and, of these, 15 were B27+ve. Random and directed cell migration was measured by 2 techniques: migration through a micropore filter and migration under an agar film. The chemo-attractant was either case in-activated serum or zymosan-activated serum. By both techniques directed motility was increased in subjects with B27 or with AS when compared to the B27-ve controls. This suggests that the disease AS and the possession of B27 are both associated with increased PMN motility. PMID:6608924

Listeria monocytogenes DNA glycosylase AdiP affects flagellar motility, biofilm formation, virulence, and stress responses

USDA-ARS?s Scientific Manuscript database

The temperature-dependent alteration of flagellar motility gene expression is critical for the foodborne pathogen Listeria monocytogenes to respond to a changing environment. In this study, a genetic determinant, L. monocytogenes f2365_0220 (lmof2365_0220), encoding a putative protein that is struct...

Rac regulates vascular endothelial growth factor stimulated motility.

PubMed

Soga, N; Connolly, J O; Chellaiah, M; Kawamura, J; Hruska, K A

2001-01-01

During angiogenesis endothelial cells migrate towards a chemotactic stimulus. Understanding the mechanism of endothelial cell migration is critical to the therapeutic manipulation of angiogenesis and ultimately cancer prevention. Vascular endothelial growth factor (VEGF) is a potent chemotactic stimulus of endothelial cells during angiogenesis. The endothelial cell signal transduction pathway of VEGF represents a potential target for cancer therapy, but the mechanisms of post-receptor signal transduction including the roles of rho family GTPases in regulating the cytoskeletal effects of VEGF in endothelial cells are not understood. Here we analyze the mechanisms of cell migration in the mouse brain endothelial cell line (bEND3). Stable transfectants containing a tetracycline repressible expression vector were used to induce expression of Rac mutants. Endothelial cell haptotaxis was stimulated by constitutively active V12Rac on collagen and vitronectin coated supports, and chemotaxis was further stimulated by VEGF. Osteopontin coated supports were the most stimulatory to bEND3 haptotaxis, but VEGF was not effective in further increasing migration on osteopontin coated supports. Haptotaxis on support coated with collagen, vitronectin, and to a lesser degree osteopontin was inhibited by N17 Rac. N17 Rac expression blocked stimulation of endothelial cell chemotaxis by VEGF. As part of the chemotactic stimulation, VEGF caused a loss of actin organization at areas of cell-cell contact and increased stress fiber expression in endothelial cells which were directed towards pores in the transwell membrane. N17 Rac prevented the stimulation of cell-cell contact disruption and the stress fiber stimulation by VEGF. These data demonstrate two pathways of regulating endothelial cell motility, one in which Rac is activated by matrix/integrin stimulation and is a crucial modulator of endothelial cell haptotaxis. The other pathway, in the presence of osteopontin, is Rac independent. VEGF stimulated chemotaxis, is critically dependent on Rac activation. Osteopontin was a potent matrix activator of motility, and perhaps one explanation for the absence of a VEGF plus osteopontin effect is that osteopontin stimulated motility was inhibitory to the Rac pathway.

Treponema pallidum Invades Intercellular Junctions of Endothelial Cell Monolayers

NASA Astrophysics Data System (ADS)

Thomas, D. Denee; Navab, Mahamad; Haake, David A.; Fogelman, Alan M.; Miller, James N.; Lovett, Michael A.

1988-05-01

The pathogenesis of syphilis reflects invasive properties of Treponema pallidum, but the actual mode of tissue invasion is unknown. We have found two in vitro parallels of treponemal invasiveness. We tested whether motile T. pallidum could invade host cells by determining the fate of radiolabeled motile organisms added to a HeLa cell monolayer; 26% of treponemes associated with the monolayer in a trypsin-resistant niche, presumably between the monolayer and the surface to which it adhered, but did not attain intracellularity. Attachment of T. pallidum to cultured human and rabbit aortic and human umbilical vein endothelial cells was 2-fold greater than to HeLa cells. We added T. pallidum to aortic endothelial cells grown on membrane filters under conditions in which tight intercellular junctions had formed. T. pallidum was able to pass through the endothelial cell monolayers without altering tight junctions, as measured by electrical resistance. In contrast, heat-killed T. pallidum and the nonpathogen Treponema phagedenis biotype Reiter failed to penetrate the monolayer. Transmission electron micrographs of sections of the monolayer showed T. pallidum in intercellular junctions. Our in vitro observations suggest that these highly motile spirochetes may leave the circulation by invading the junctions between endothelial cells.

Computer-aided sperm analysis (CASA) in the medical laboratory: CASA in diagnostic andrology and assisted conception.

PubMed

Tomlinson, Mathew J; Naeem, Asad

2018-03-21

CASA has been used in reproductive medicine and pathology laboratories for over 25 years, yet the 'fertility industry' generally remains sceptical and has avoided automation, despite clear weaknesses in manual semen analysis. Early implementers had difficulty in validating CASA-Mot instruments against recommended manual methods (haemocytometer) due to the interference of seminal debris and non-sperm cells, which also affects the accuracy of grading motility. Both the inability to provide accurate sperm counts and a lack of consensus as to the value of sperm kinematic parameters appear to have continued to have a negative effect on CASA-Mot's reputation. One positive interpretation from earlier work is that at least one or more measures of sperm velocity adds clinical value to the semen analysis, and these are clearly more objective than any manual motility analysis. Moreover, recent CASA-Mot systems offer simple solutions to earlier problems in eliminating artefacts and have been successfully validated for sperm concentration; as a result, they should be viewed with more confidence in relation to motility grading. Sperm morphology and DNA testing both require an evidence-based consensus and a well-validated (reliable, reproducible) assay to be developed before automation of either can be of real clinical benefit.

Effect of storage in short--and long-term commercial semen extenders on the motility, plasma membrane and chromatin integrity of boar spermatozoa.

PubMed

De Ambrogi, Marco; Ballester, Juan; Saravia, Fernando; Caballero, Ignacio; Johannisson, Anders; Wallgren, Margareta; Andersson, Magnus; Rodriguez-Martinez, Heriberto

2006-10-01

For artificial insemination (AI) in pigs, preservation of liquid boar semen at 16-20 degrees C is still common practice as sperm cryopreservation remains suboptimal in this species. To meet the different needs of the swine industry, several extenders have been developed to preserve semen in liquid form for short--and long-term storage. In the present study, three different commercial extenders devised for short-term (BTS+) or long-term preservation (MR-A and X-Cell), were used to test whether storage of semen from four mature, fertile boars at 17 degrees C for 96 h would affect sperm characteristics relevant for fertility, such as motility, membrane integrity and chromatin stability. Computer-assisted sperm analysis, and stainings with the acylated membrane dye SYBR-14/propidium iodide, and acridine orange in connection with flow cytometry were used to evaluate these variables. Percentages of total motile spermatozoa decreased slightly, but significantly, after 72-96 h. While membrane integrity values varied during the period of study, no significant changes in either membrane integrity or chromatin stability were, however, registered. This suggests a customary 96-day storage at 17 degrees C in these extenders was too short an interval to cause losses of integrity in nuclear DNA in the boar population studied.