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Sample records for inhibits cell motility

  1. Lichen Secondary Metabolite, Physciosporin, Inhibits Lung Cancer Cell Motility.

    PubMed

    Yang, Yi; Park, So-Yeon; Nguyen, Thanh Thi; Yu, Young Hyun; Nguyen, Tru Van; Sun, Eun Gene; Udeni, Jayalal; Jeong, Min-Hye; Pereira, Iris; Moon, Cheol; Ha, Hyung-Ho; Kim, Kyung Keun; Hur, Jae-Seoun; Kim, Hangun

    2015-01-01

    Lichens produce various unique chemicals that can be used for pharmaceutical purposes. To screen for novel lichen secondary metabolites showing inhibitory activity against lung cancer cell motility, we tested acetone extracts of 13 lichen samples collected in Chile. Physciosporin, isolated from Pseudocyphellaria coriacea (Hook f. & Taylor) D.J. Galloway & P. James, was identified as an effective compound and showed significant inhibitory activity in migration and invasion assays against human lung cancer cells. Physciosporin treatment reduced both protein and mRNA levels of N-cadherin with concomitant decreases in the levels of epithelial-mesenchymal transition markers such as snail and twist. Physciosporin also suppressed KITENIN (KAI1 C-terminal interacting tetraspanin)-mediated AP-1 activity in both the absence and presence of epidermal growth factor stimulation. Quantitative real-time PCR analysis showed that the expression of the metastasis suppressor gene, KAI1, was increased while that of the metastasis enhancer gene, KITENIN, was dramatically decreased by physciosporin. Particularly, the activity of 3'-untranslated region of KITENIN was decreased by physciosporin. Moreover, Cdc42 and Rac1 activities were decreased by physciosporin. These results demonstrated that the lichen secondary metabolite, physciosporin, inhibits lung cancer cell motility through novel mechanisms of action. PMID:26371759

  2. Lichen Secondary Metabolite, Physciosporin, Inhibits Lung Cancer Cell Motility

    PubMed Central

    Yang, Yi; Park, So-Yeon; Nguyen, Thanh Thi; Yu, Young Hyun; Nguyen, Tru Van; Sun, Eun Gene; Udeni, Jayalal; Jeong, Min-Hye; Pereira, Iris; Moon, Cheol; Ha, Hyung-Ho; Kim, Kyung Keun; Hur, Jae-Seoun; Kim, Hangun

    2015-01-01

    Lichens produce various unique chemicals that can be used for pharmaceutical purposes. To screen for novel lichen secondary metabolites showing inhibitory activity against lung cancer cell motility, we tested acetone extracts of 13 lichen samples collected in Chile. Physciosporin, isolated from Pseudocyphellaria coriacea (Hook f. & Taylor) D.J. Galloway & P. James, was identified as an effective compound and showed significant inhibitory activity in migration and invasion assays against human lung cancer cells. Physciosporin treatment reduced both protein and mRNA levels of N-cadherin with concomitant decreases in the levels of epithelial-mesenchymal transition markers such as snail and twist. Physciosporin also suppressed KITENIN (KAI1 C-terminal interacting tetraspanin)-mediated AP-1 activity in both the absence and presence of epidermal growth factor stimulation. Quantitative real-time PCR analysis showed that the expression of the metastasis suppressor gene, KAI1, was increased while that of the metastasis enhancer gene, KITENIN, was dramatically decreased by physciosporin. Particularly, the activity of 3’-untranslated region of KITENIN was decreased by physciosporin. Moreover, Cdc42 and Rac1 activities were decreased by physciosporin. These results demonstrated that the lichen secondary metabolite, physciosporin, inhibits lung cancer cell motility through novel mechanisms of action. PMID:26371759

  3. Inhibition of tumor cell proliferation and motility by fibroblasts is both contact and soluble factor dependent

    PubMed Central

    Alkasalias, Twana; Flaberg, Emilie; Kashuba, Vladimir; Alexeyenko, Andrey; Pavlova, Tatiana; Savchenko, Andrii; Szekely, Laszlo; Klein, George; Guven, Hayrettin

    2014-01-01

    Normal human and murine fibroblasts can inhibit proliferation of tumor cells when cocultured in vitro. The inhibitory capacity varies depending on the donor and the site of origin of the fibroblast. We showed previously that effective inhibition requires formation of a morphologically intact fibroblast monolayer before seeding of the tumor cells. Here we show that inhibition is extended to motility of tumor cells and we dissect the factors responsible for these inhibitory functions. We find that inhibition is due to two different sets of molecules: (i) the extracellular matrix (ECM) and other surface proteins of the fibroblasts, which are responsible for contact-dependent inhibition of tumor cell proliferation; and (ii) soluble factors secreted by fibroblasts when confronted with tumor cells (confronted conditioned media, CCM) contribute to inhibition of tumor cell proliferation and motility. However, conditioned media (CM) obtained from fibroblasts alone (nonconfronted conditioned media, NCM) did not inhibit tumor cell proliferation and motility. In addition, quantitative PCR (Q-PCR) data show up-regulation of proinflammatory genes. Moreover, comparison of CCM and NCM with an antibody array for 507 different soluble human proteins revealed differential expression of growth differentiation factor 15, dickkopf-related protein 1, endothelial-monocyte-activating polypeptide II, ectodysplasin A2, Galectin-3, chemokine (C-X-C motif) ligand 2, Nidogen1, urokinase, and matrix metalloproteinase 3. PMID:25404301

  4. Polarized cell motility induces hydrogen peroxide to inhibit cofilin via cysteine oxidation.

    PubMed

    Cameron, Jenifer M; Gabrielsen, Mads; Chim, Ya Hua; Munro, June; McGhee, Ewan J; Sumpton, David; Eaton, Philip; Anderson, Kurt I; Yin, Huabing; Olson, Michael F

    2015-06-01

    Mesenchymal cell motility is driven by polarized actin polymerization [1]. Signals at the leading edge recruit actin polymerization machinery to promote membrane protrusion, while matrix adhesion generates tractive force to propel forward movement. To work effectively, cell motility is regulated by a complex network of signaling events that affect protein activity and localization. H2O2 has an important role as a diffusible second messenger [2], and mediates its effects through oxidation of cysteine thiols. One cell activity influenced by H2O2 is motility [3]. However, a lack of sensitive and H2O2-specific probes for measurements in live cells has not allowed for direct observation of H2O2 accumulation in migrating cells or protrusions. In addition, the identities of proteins oxidized by H2O2 that contribute to actin dynamics and cell motility have not been characterized. We now show, as determined by fluorescence lifetime imaging microscopy, that motile cells generate H2O2 at membranes and cell protrusions and that H2O2 inhibits cofilin activity through oxidation of cysteines 139 (C139) and 147 (C147). Molecular modeling suggests that C139 oxidation would sterically hinder actin association, while the increased negative charge of oxidized C147 would lead to electrostatic repulsion of the opposite negatively charged surface. Expression of oxidation-resistant cofilin impairs cell spreading, adhesion, and directional migration. These findings indicate that H2O2 production contributes to polarized cell motility through localized cofilin inhibition and that there are additional proteins oxidized during cell migration that might have similar roles.

  5. Shielding of the Geomagnetic Field Alters Actin Assembly and Inhibits Cell Motility in Human Neuroblastoma Cells

    PubMed Central

    Mo, Wei-Chuan; Zhang, Zi-Jian; Wang, Dong-Liang; Liu, Ying; Bartlett, Perry F.; He, Rong-Qiao

    2016-01-01

    Accumulating evidence has shown that absence of the geomagnetic field (GMF), the so-called hypomagnetic field (HMF) environment, alters the biological functions in seemingly non-magnetosensitive cells and organisms, which indicates that the GMF could be sensed by non-iron-rich and non-photo-sensing cells. The underlying mechanisms of the HMF effects on those cells are closely related to their GMF sensation but remain poorly understood so far. Previously, we found that the HMF represses expressions of genes associated with cell migration and cytoskeleton assembly in human neuroblastoma cells (SH-SY5Y cell line). Here, we measured the HMF-induced changes on cell morphology, adhesion, motility and actin cytoskeleton in SH-SY5Y cells. The HMF inhibited cell adhesion and migration accompanied with a reduction in cellular F-actin amount. Moreover, following exposure to the HMF, the number of cell processes was reduced and cells were smaller in size and more round in shape. Furthermore, disordered kinetics of actin assembly in vitro were observed during exposure to the HMF, as evidenced by the presence of granule and meshed products. These results indicate that elimination of the GMF affects assembly of the motility-related actin cytoskeleton, and suggest that F-actin is a target of HMF exposure and probably a mediator of GMF sensation. PMID:27029216

  6. Knockdown of Golgi phosphoprotein 2 inhibits hepatocellular carcinoma cell proliferation and motility

    PubMed Central

    Liu, Yiming; Zhang, Xiaodi; Sun, Ting; Jiang, Junchang; Li, Ying; Chen, Mingliang; Wei, Zhen; Jiang, Weiqin; Zhou, Linfu

    2016-01-01

    Golgi phosphoprotein 2 (GP73) is highly expressed in hepatocellular carcinoma (HCC) cells, where it serves as a biomarker and indicator of disease progression. We used MTS assays, anchorage-independent cell colony formation assays and a xenograft tumor model to show that GP73-specific siRNAs inhibit HCC proliferation in HepG2, SMMC-7721, and Huh7 cell lines and in vivo. Following GP73 silencing, levels of p-Rb, a factor related to metastasis, were reduced, but cell cycle progression was unaffected. Our results suggest that GP73 silencing may not directly suppress proliferation, but may instead inhibit cell motility. Results from proliferation assays suggest GP73 reduces expression of epithelial mesenchymal transition (EMT)-related factors and promotes cell motility, while transwell migration and invasion assays indicated a possible role in metastasis. Immunofluorescence co-localization microscopy and immunoblotting showed that GP73 decreases expression of N-cadherin and E-cadherin, two key factors in EMT, which may in turn decrease intracellular adhesive forces and promote cell motility. This study confirmed that GP73 expression leads to increased expression of EMT-related proteins and that GP73 silencing reduces HCC cell migration in vitro. These findings suggest that GP73 silencing through siRNA delivery may provide a novel low-toxicity therapy for the inhibition of tumor proliferation and metastasis. PMID:26870893

  7. MYBPH inhibits NM IIA assembly via direct interaction with NMHC IIA and reduces cell motility

    SciTech Connect

    Hosono, Yasuyuki; Usukura, Jiro; Yamaguchi, Tomoya; Yanagisawa, Kiyoshi; Suzuki, Motoshi; Takahashi, Takashi

    2012-11-09

    Highlights: Black-Right-Pointing-Pointer MYBPH inhibits NMHC IIA assembly and cell motility. Black-Right-Pointing-Pointer MYBPH interacts to assembly-competent NM IIA. Black-Right-Pointing-Pointer MYBPH inhibits RLC and NMHC IIA, independent components of NM IIA. -- Abstract: Actomyosin filament assembly is a critical step in tumor cell migration. We previously found that myosin binding protein H (MYBPH) is directly transactivated by the TTF-1 lineage-survival oncogene in lung adenocarcinomas and inhibits phosphorylation of the myosin regulatory light chain (RLC) of non-muscle myosin IIA (NM IIA) via direct interaction with Rho kinase 1 (ROCK1). Here, we report that MYBPH also directly interacts with an additional molecule, non-muscle myosin heavy chain IIA (NMHC IIA), which was found to occur between MYBPH and the rod portion of NMHC IIA. MYBPH inhibited NMHC IIA assembly and reduced cell motility. Conversely, siMYBPH-induced increased motility was partially, yet significantly, suppressed by blebbistatin, a non-muscle myosin II inhibitor, while more profound effects were attained by combined treatment with siROCK1 and blebbistatin. Electron microscopy observations showed well-ordered paracrystals of NMHC IIA reflecting an assembled state, which were significantly less frequently observed in the presence of MYBPH. Furthermore, an in vitro sedimentation assay showed that a greater amount of NMHC IIA was in an unassembled state in the presence of MYBPH. Interestingly, treatment with a ROCK inhibitor that impairs transition of NM IIA from an assembly-incompetent to assembly-competent state reduced the interaction between MYBPH and NMHC IIA, suggesting that MYBPH has higher affinity to assembly-competent NM IIA. These results suggest that MYBPH inhibits RLC and NMHC IIA, independent components of NM IIA, and negatively regulates actomyosin organization at 2 distinct steps, resulting in firm inhibition of NM IIA assembly.

  8. A20 inhibits the motility of HCC cells induced by TNF-α.

    PubMed

    Wang, Xianteng; Ma, Chao; Zong, Zhaoyun; Xiao, Ying; Li, Na; Guo, Chun; Zhang, Lining; Shi, Yongyu

    2016-03-22

    Metastasis of hepatocellular carcinoma (HCC) can be facilitated by TNF-α, a prototypical inflammatory cytokine in the HCC microenvironment. A20 is a negative regulator of NF-κB signaling pathway. In the present study we ask whether A20 plays a role in HCC metastasis. We found that A20 expression was downregulated in the invasive cells of microvascular invasions (MVI) compared with the noninvasive cells in 89 tissue samples from patients with HCC by immunochemistry methods. Overexpression of A20 in HCC cell lines inhibited their motility induced by TNF-α. Furthermore, the overexpression of A20 inhibited epithelial-mesenchymal transition (EMT), FAK activation and RAC1 activity. By contrast, knockdown of A20 in one HCC cell line results in the converse. In addition, the overexpression of A20 restrained the formation of MVI in HCC xenograft in nude mice treated with TNF-α. All the results suggested that A20 functioned as a negative regulator in motility of HCC cells induced by TNF-α. PMID:26909601

  9. A20 inhibits the motility of HCC cells induced by TNF-α

    PubMed Central

    Xiao, Ying; Li, Na; Guo, Chun; Zhang, Lining; Shi, Yongyu

    2016-01-01

    Metastasis of hepatocellular carcinoma (HCC) can be facilitated by TNF-α, a prototypical inflammatory cytokine in the HCC microenvironment. A20 is a negative regulator of NF-κB signaling pathway. In the present study we ask whether A20 plays a role in HCC metastasis. We found that A20 expression was downregulated in the invasive cells of microvascular invasions (MVI) compared with the noninvasive cells in 89 tissue samples from patients with HCC by immunochemistry methods. Overexpression of A20 in HCC cell lines inhibited their motility induced by TNF-α. Furthermore, the overexpression of A20 inhibited epithelial-mesenchymal transition (EMT), FAK activation and RAC1 activity. By contrast, knockdown of A20 in one HCC cell line results in the converse. In addition, the overexpression of A20 restrained the formation of MVI in HCC xenograft in nude mice treated with TNF-α. All the results suggested that A20 functioned as a negative regulator in motility of HCC cells induced by TNF-α. PMID:26909601

  10. Prodigiosin inhibits motility and activates bacterial cell death revealing molecular biomarkers of programmed cell death.

    PubMed

    Darshan, N; Manonmani, H K

    2016-12-01

    The antimicrobial activity of prodigiosin from Serratia nematodiphila darsh1, a bacterial pigment was tested against few food borne bacterial pathogens Bacillus cereus, Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli. The mode of action of prodigiosin was studied. Prodigiosin induced bactericidal activity indicating a stereotypical set of biochemical and morphological feature of Programmed cell death (PCD). PCD involves DNA fragmentation, generation of ROS, and expression of a protein with caspase-like substrate specificity in bacterial cells. Prodigiosin was observed to be internalized into bacterial cells and was localized predominantly in the membrane and the nuclear fraction, thus, facilitating intracellular trafficking and then binding of prodigiosin to the bacterial DNA. Corresponding to an increasing concentration of prodigiosin, the level of certain proteases were observed to increase in bacteria studied, thus initiating the onset of PCD. Prodigiosin at a sub-inhibitory concentration inhibits motility of pathogens. Our observations indicated that prodigiosin could be a promising antibacterial agent and could be used in the prevention of bacterial infections. PMID:27460563

  11. Prodigiosin inhibits motility and activates bacterial cell death revealing molecular biomarkers of programmed cell death.

    PubMed

    Darshan, N; Manonmani, H K

    2016-12-01

    The antimicrobial activity of prodigiosin from Serratia nematodiphila darsh1, a bacterial pigment was tested against few food borne bacterial pathogens Bacillus cereus, Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli. The mode of action of prodigiosin was studied. Prodigiosin induced bactericidal activity indicating a stereotypical set of biochemical and morphological feature of Programmed cell death (PCD). PCD involves DNA fragmentation, generation of ROS, and expression of a protein with caspase-like substrate specificity in bacterial cells. Prodigiosin was observed to be internalized into bacterial cells and was localized predominantly in the membrane and the nuclear fraction, thus, facilitating intracellular trafficking and then binding of prodigiosin to the bacterial DNA. Corresponding to an increasing concentration of prodigiosin, the level of certain proteases were observed to increase in bacteria studied, thus initiating the onset of PCD. Prodigiosin at a sub-inhibitory concentration inhibits motility of pathogens. Our observations indicated that prodigiosin could be a promising antibacterial agent and could be used in the prevention of bacterial infections.

  12. Cucurbitacin I Inhibits Cell Motility by Indirectly Interfering with Actin Dynamics

    PubMed Central

    Knecht, David A.; LaFleur, Rebecca A.; Kahsai, Alem W.; Argueta, Christian E.; Beshir, Anwar B.; Fenteany, Gabriel

    2010-01-01

    Background Cucurbitacins are plant natural products that inhibit activation of the Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway by an unknown mechanism. They are also known to cause changes in the organization of the actin cytoskeleton. Methodology/Principal Findings We show that cucurbitacin I potently inhibits the migration of Madin-Darby canine kidney (MDCK) cell sheets during wound closure, as well as the random motility of B16-F1 mouse melanoma cells, but has no effect on movement of Dictyostelium discoideum amoebae. Upon treatment of MDCK or B16-F1 cells with cucurbitacin I, there is a very rapid cessation of motility and gradual accumulation of filamentous actin aggregates. The cellular effect of the compound is similar to that observed when cells are treated with the actin filament-stabilizing agent jasplakinolide. However, we found that, unlike jasplakinolide or phallacidin, cucurbitacin I does not directly stabilize actin filaments. In in vitro actin depolymerization experiments, cucurbitacin I had no effect on the rate of actin filament disassembly at the nanomolar concentrations that inhibit cell migration. At elevated concentrations, the depolymerization rate was also unaffected, although there was a delay in the initiation of depolymerization. Therefore, cucurbitacin I targets some factor involved in cellular actin dynamics other than actin itself. Two candidate proteins that play roles in actin depolymerization are the actin-severing proteins cofilin and gelsolin. Cucurbitacin I possesses electrophilic reactivity that may lead to chemical modification of its target protein, as suggested by structure-activity relationship data. However, mass spectrometry revealed no evidence for modification of purified cofilin or gelsolin by cucurbitacin I. Conclusions/Significance Cucurbitacin I results in accumulation of actin filaments in cells by a unique indirect mechanism. Furthermore, the proximal target of

  13. Silibinin inhibits triple negative breast cancer cell motility by suppressing TGF-β2 expression.

    PubMed

    Kim, Sangmin; Han, Jeonghun; Jeon, Myeongjin; You, Daeun; Lee, Jeongmin; Kim, Hee Jung; Bae, Sarang; Nam, Seok Jin; Lee, Jeong Eon

    2016-08-01

    Transforming growth factor-beta (TGF-β) is a multifunctional cytokine that regulates many biological events including cell motility and angiogenesis. Here, we investigated the role of elevated TGF-β2 level in triple negative breast cancer (TNBC) cells and the inhibitory effect of silibinin on TGF-β2 action in TNBC cells. Breast cancer patients with high TGF-β2 expression have a poor prognosis. The levels of TGF-β2 expression increased significantly in TNBC cells compared with those in non-TNBC cells. In addition, cell motility-related genes such as fibronectin (FN) and matrix metalloproteinase-2 (MMP-2) expression also increased in TNBC cells. Basal FN, MMP-2, and MMP-9 expression levels decreased in response to LY2109761, a dual TGF-β receptor I/II inhibitor, in TNBC cells. TNBC cell migration also decreased in response to LY2109761. Furthermore, we observed that TGF-β2 augmented the FN, MMP-2, and MMP-9 expression levels in a time- and dose-dependent manner. In contrast, TGF-β2-induced FN, MMP-2, and MMP-9 expression levels decreased significantly in response to LY2109761. Interestingly, we found that silibinin decreased TGF-β2 mRNA expression level but not that of TGF-β1 in TNBC cells. Cell migration as well as basal FN and MMP-2 expression levels decreased in response to silibinin. Furthermore, silibinin significantly decreased TGF-β2-induced FN, MMP-2, and MMP-9 expression levels and suppressed the lung metastasis of TNBC cells. Taken together, these results suggest that silibinin suppresses metastatic potential of TNBC cells by inhibiting TGF-β2 expression in TNBC cells. Thus, silibinin may be a promising therapeutic drug to treat TNBC.

  14. All-trans-retinoic Acid Modulates the Plasticity and Inhibits the Motility of Breast Cancer Cells

    PubMed Central

    Zanetti, Adriana; Affatato, Roberta; Centritto, Floriana; Fratelli, Maddalena; Kurosaki, Mami; Barzago, Maria Monica; Bolis, Marco; Terao, Mineko; Garattini, Enrico; Paroni, Gabriela

    2015-01-01

    All-trans-retinoic acid (ATRA) is a natural compound proposed for the treatment/chemoprevention of breast cancer. Increasing evidence indicates that aberrant regulation of epithelial-to-mesenchymal transition (EMT) is a determinant of the cancer cell invasive and metastatic behavior. The effects of ATRA on EMT are largely unknown. In HER2-positive SKBR3 and UACC812 cells, showing co-amplification of the ERBB2 and RARA genes, ATRA activates a RARα-dependent epithelial differentiation program. In SKBR3 cells, this causes the formation/reorganization of adherens and tight junctions. Epithelial differentiation and augmented cell-cell contacts underlie the anti-migratory action exerted by the retinoid in cells exposed to the EMT-inducing factors EGF and heregulin-β1. Down-regulation of NOTCH1, an emerging EMT modulator, is involved in the inhibition of motility by ATRA. Indeed, the retinoid blocks NOTCH1 up-regulation by EGF and/or heregulin-β1. Pharmacological inhibition of γ-secretase and NOTCH1 processing also abrogates SKBR3 cell migration. Stimulation of TGFβ contributes to the anti-migratory effect of ATRA. The retinoid switches TGFβ from an EMT-inducing and pro-migratory determinant to an anti-migratory mediator. Inhibition of the NOTCH1 pathway not only plays a role in the anti-migratory action of ATRA; it is relevant also for the anti-proliferative activity of the retinoid in HCC1599 breast cancer cells, which are addicted to NOTCH1 for growth/viability. This effect is enhanced by the combination of ATRA and the γ-secretase inhibitor N-(N-(3,5-difluorophenacetyl)-l-alanyl)-S-phenylglycine t-butyl ester, supporting the concept that the two compounds act at the transcriptional and post-translational levels along the NOTCH1 pathway. PMID:26018078

  15. Daucus carota Pentane/Diethyl Ether Fraction Inhibits Motility and Reduces Invasion of Cancer Cells.

    PubMed

    Zgheib, Perla; Daher, Costantine F; Mroueh, Mohamad; Nasrallah, Anita; Taleb, Robin I; El-Sibai, Mirvat

    2014-01-01

    Daucus carota (DC) is a herb used in folklore medicine in Lebanon to treat numerous diseases including cancer. Recent studies in our laboratory on DC oil and its fractions revealed potent anticancer activities in vitro and in vivo. The present study aims to investigate the effect of the most potent DC fraction, pentane/diethyl ether (50:50), on lung, skin, breast and glioblastoma cancer cell motility and invasion. Upon treatment, a pronounced decrease in cancer cell motility was observed in the 4 cell lines. The treatment also led to a decrease in cancer cell invasion and an increased cell adhesion. Additionally, the DC fraction caused a decrease in the activation of the ρ-GTPases Rac and CDC42, a finding that may partially explain the treatment-induced decrease in cell motility. The current study demonstrates a crucial effect of the DC pentane/diethyl ether fraction on cancer cell motility and metastasis, making it a potential candidate for cancer therapy specifically targeting cancer motility and metastasis.

  16. Daucus carota Pentane/Diethyl Ether Fraction Inhibits Motility and Reduces Invasion of Cancer Cells.

    PubMed

    Zgheib, Perla; Daher, Costantine F; Mroueh, Mohamad; Nasrallah, Anita; Taleb, Robin I; El-Sibai, Mirvat

    2014-01-01

    Daucus carota (DC) is a herb used in folklore medicine in Lebanon to treat numerous diseases including cancer. Recent studies in our laboratory on DC oil and its fractions revealed potent anticancer activities in vitro and in vivo. The present study aims to investigate the effect of the most potent DC fraction, pentane/diethyl ether (50:50), on lung, skin, breast and glioblastoma cancer cell motility and invasion. Upon treatment, a pronounced decrease in cancer cell motility was observed in the 4 cell lines. The treatment also led to a decrease in cancer cell invasion and an increased cell adhesion. Additionally, the DC fraction caused a decrease in the activation of the ρ-GTPases Rac and CDC42, a finding that may partially explain the treatment-induced decrease in cell motility. The current study demonstrates a crucial effect of the DC pentane/diethyl ether fraction on cancer cell motility and metastasis, making it a potential candidate for cancer therapy specifically targeting cancer motility and metastasis. PMID:26088465

  17. Claudin-18 inhibits cell proliferation and motility mediated by inhibition of phosphorylation of PDK1 and Akt in human lung adenocarcinoma A549 cells.

    PubMed

    Shimobaba, Shun; Taga, Saeko; Akizuki, Risa; Hichino, Asami; Endo, Satoshi; Matsunaga, Toshiyuki; Watanabe, Ryo; Yamaguchi, Masahiko; Yamazaki, Yasuhiro; Sugatani, Junko; Ikari, Akira

    2016-06-01

    Abnormal expression of claudin subtypes has been reported in various cancers. However, the pathological role of each claudin has not been clarified in detail. Claudin-18 was absent in human non-small cell and small cell lung cancers, although it is expressed in normal lung tissues. Here, we examined the effect of claudin-18 expression on the expression of junctional proteins, cell proliferation, and cell motility using human lung adenocarcinoma A549 cells. Real-time PCR and western blotting showed that exogenous expression of claudin-18 had no effect on the expression of junctional proteins including claudin-1, zonula occludens-1 (ZO-1), occludin, and E-cadherin. Claudin-18 was mainly distributed in cell-cell contact areas concomitant with ZO-1. Cell proliferation was significantly decreased at 48 and 72h after seeding of claudin 18-expressing cells. Claudin-18 suppressed cell motility, whereas it increased cell death in anoikis. Claudin-18 decreased phosphorylated (p)-3-phosphoinositide-dependent protein kinase-1 (PDK1) and p-Akt levels without affecting p-epidermal growth factor receptor and p-phosphatidylinositol-3 kinase (PI3K) levels. Furthermore, claudin-18 was bound with PDK1 and suppressed the nuclear localization of PDK1. We suggest that claudin-18 suppresses the abnormal proliferation and motility of lung epithelial cells mediated by inhibition of the PI3K/PDK1/Akt signaling pathway.

  18. Targeting the MAP kinase pathway in astrocytoma cells using a recombinant anthrax lethal toxin as a way to inhibit cell motility and invasion.

    PubMed

    Al-Dimassi, Saleh; Salloum, Gilbert; Saykali, Bechara; Khoury, Oula; Liu, Shihui; Leppla, Stephen H; Abi-Habib, Ralph; El-Sibai, Mirvat

    2016-05-01

    Malignant astrocytomas are highly invasive into adjacent and distant regions of the normal brain. Understanding and targeting cancer cell invasion is an important therapeutic approach. Cell invasion is a complex process that replies on many signaling pathways including the mitogen-activated protein (MAP) kinase (MAPK). In many cell lines, the use of MAPK-targeted drugs proved to be a potential method to inhibit cancer cell motility. In the present study, we use a recombinant anthrax lethal toxin (LeTx), which selectively inhibits the MAPK pathway, in order to target invasion. LeTx proved ineffective on cell survival in astrocytoma (as well as normal cells). However, astrocytoma cells that were treated with LeTx showed a significant decrease in cell motility as seen by wound healing as well as random 2D motility in serum. The cells also showed a decrease in invasion across a collagen matrix. The effect of LeTx on cell migration was mediated though the deregulation of Rho GTPases, which play a role in cell motility. Finally, the effect of LeTx on cell migration and Rho GTPases was mimicked by the inhibition of the MAPK pathway. In this study, we describe for the first time the effect of the LeTx on cancer cell motility and invasion not cell survival making it a potentially selective brain tumor invasion inhibitor. PMID:26984023

  19. EGFR inhibition in glioma cells modulates Rho signaling to inhibit cell motility and invasion and cooperates with temozolomide to reduce cell growth.

    PubMed

    Ramis, Guillem; Thomàs-Moyà, Elena; Fernández de Mattos, Silvia; Rodríguez, José; Villalonga, Priam

    2012-01-01

    Enforced EGFR activation upon gene amplification and/or mutation is a common hallmark of malignant glioma. Small molecule EGFR tyrosine kinase inhibitors, such as erlotinib (Tarceva), have shown some activity in a subset of glioma patients in recent trials, although the reported data on the cellular basis of glioma cell responsiveness to these compounds have been contradictory. Here we have used a panel of human glioma cell lines, including cells with amplified or mutant EGFR, to further characterize the cellular effects of EGFR inhibition with erlotinib. Dose-response and cellular growth assays indicate that erlotinib reduces cell proliferation in all tested cell lines without inducing cytotoxic effects. Flow cytometric analyses confirm that EGFR inhibition does not induce apoptosis in glioma cells, leading to cell cycle arrest in G(1). Interestingly, erlotinib also prevents spontaneous multicellular tumour spheroid growth in U87MG cells and cooperates with sub-optimal doses of temozolomide (TMZ) to reduce multicellular tumour spheroid growth. This cooperation appears to be schedule-dependent, since pre-treatment with erlotinib protects against TMZ-induced cytotoxicity whereas concomitant treatment results in a cooperative effect. Cell cycle arrest in erlotinib-treated cells is associated with an inhibition of ERK and Akt signaling, resulting in cyclin D1 downregulation, an increase in p27(kip1) levels and pRB hypophosphorylation. Interestingly, EGFR inhibition also perturbs Rho GTPase signaling and cellular morphology, leading to Rho/ROCK-dependent formation of actin stress fibres and the inhibition of glioma cell motility and invasion.

  20. α-TEA inhibits the growth and motility of human colon cancer cells via targeting RhoA/ROCK signaling.

    PubMed

    Yao, Jialin; Gao, Peng; Xu, Yang; Li, Zhaozhu

    2016-09-01

    Colon or colorectal cancer is a common type of human cancer, which originates in the intestine crassum or the rectum. In the United States, colorectal cancer has one of the highest rates of cancer‑related mortality. Investigating novel chemotherapeutic approaches is significant in the treatment of cancers, such as colorectal cancer. α-tocopherol ether-linked acetic acid (α-TEA) is a potent anticancer agent in multiple types of human cancer. However, its effect remains to be determined in colon cancer. In this study, HCT116 and SW480 human colon cancer cells were used to investigate the anticancer role of α-TEA. It was demonstrated that α-TEA inhibited cell proliferation, migration and invasion in colon cancer cells. Furthermore, it was shown that α-TEA downregulated the activity of RhoA and phosphorylated Rho-associated protein kinase (ROCK) substrate myosin light chain (MLC) using a pull-down assay and western blotting, respectively, implying that the RhoA/ROCK pathway is involved in α-TEA-mediated cell growth and motility inhibition. In order to confirm this hypothesis a RhoA inhibitor (clostridium botulinum C3 exoenzyme), a ROCK inhibitor (Y27632) and RhoA small interfering (si)RNA were applied to block RhoA/ROCK signaling. This resulted in the attenuation of MLC phosphorylation, and augmentation of α-TEA-mediated growth and motility inhibition in colon cancer cells. In conclusion, these results indicate that α-TEA inhibits growth and motility in colon cancer cells possibly by targeting RhoA/ROCK signaling. Moreover, combined with RhoA or ROCK inhibitors, α-TEA may exhibit a more effective inhibitory role in colon cancer.

  1. α-TEA inhibits the growth and motility of human colon cancer cells via targeting RhoA/ROCK signaling

    PubMed Central

    Yao, Jialin; Gao, Peng; Xu, Yang; Li, Zhaozhu

    2016-01-01

    Colon or colorectal cancer is a common type of human cancer, which originates in the intestine crassum or the rectum. In the United States, colorectal cancer has one of the highest rates of cancer-related mortality. Investigating novel chemotherapeutic approaches is significant in the treatment of cancers, such as colorectal cancer. α-tocopherol ether-linked acetic acid (α-TEA) is a potent anticancer agent in multiple types of human cancer. However, its effect remains to be determined in colon cancer. In this study, HCT116 and SW480 human colon cancer cells were used to investigate the anticancer role of α-TEA. It was demonstrated that α-TEA inhibited cell proliferation, migration and invasion in colon cancer cells. Furthermore, it was shown that α-TEA downregulated the activity of RhoA and phosphorylated Rho-associated protein kinase (ROCK) substrate myosin light chain (MLC) using a pull-down assay and western blotting, respectively, implying that the RhoA/ROCK pathway is involved in α-TEA-mediated cell growth and motility inhibition. In order to confirm this hypothesis a RhoA inhibitor (clostridium botulinum C3 exoenzyme), a ROCK inhibitor (Y27632) and RhoA small interfering (si)RNA were applied to block RhoA/ROCK signaling. This resulted in the attenuation of MLC phosphorylation, and augmentation of α-TEA-mediated growth and motility inhibition in colon cancer cells. In conclusion, these results indicate that α-TEA inhibits growth and motility in colon cancer cells possibly by targeting RhoA/ROCK signaling. Moreover, combined with RhoA or ROCK inhibitors, α-TEA may exhibit a more effective inhibitory role in colon cancer. PMID:27432222

  2. α-TEA inhibits the growth and motility of human colon cancer cells via targeting RhoA/ROCK signaling.

    PubMed

    Yao, Jialin; Gao, Peng; Xu, Yang; Li, Zhaozhu

    2016-09-01

    Colon or colorectal cancer is a common type of human cancer, which originates in the intestine crassum or the rectum. In the United States, colorectal cancer has one of the highest rates of cancer‑related mortality. Investigating novel chemotherapeutic approaches is significant in the treatment of cancers, such as colorectal cancer. α-tocopherol ether-linked acetic acid (α-TEA) is a potent anticancer agent in multiple types of human cancer. However, its effect remains to be determined in colon cancer. In this study, HCT116 and SW480 human colon cancer cells were used to investigate the anticancer role of α-TEA. It was demonstrated that α-TEA inhibited cell proliferation, migration and invasion in colon cancer cells. Furthermore, it was shown that α-TEA downregulated the activity of RhoA and phosphorylated Rho-associated protein kinase (ROCK) substrate myosin light chain (MLC) using a pull-down assay and western blotting, respectively, implying that the RhoA/ROCK pathway is involved in α-TEA-mediated cell growth and motility inhibition. In order to confirm this hypothesis a RhoA inhibitor (clostridium botulinum C3 exoenzyme), a ROCK inhibitor (Y27632) and RhoA small interfering (si)RNA were applied to block RhoA/ROCK signaling. This resulted in the attenuation of MLC phosphorylation, and augmentation of α-TEA-mediated growth and motility inhibition in colon cancer cells. In conclusion, these results indicate that α-TEA inhibits growth and motility in colon cancer cells possibly by targeting RhoA/ROCK signaling. Moreover, combined with RhoA or ROCK inhibitors, α-TEA may exhibit a more effective inhibitory role in colon cancer. PMID:27432222

  3. 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

  4. Cell motility on nanotopography

    NASA Astrophysics Data System (ADS)

    Kimura, Masahiro; Tsai, Irene; Green, Angelo; Jacobson, Bruce; Russell, Thomas

    2003-03-01

    Cell motility is strongly influenced by the structure of the substratum. Understanding cells motility on a surface has significant applications both in vivo and in vitro applications, such as biological sensors and hip replacement. A gradient surface is used to study the effect of the lateral nanotopography on cell motility. A gradient surface is generated by block copolymer and homopolymer blends, where the concentration of the components varies uniformly across the surface. The two homopolymers phase separate on the micron scale and this length scale gradually decrease to the nanoscopic, i.e. microphase separation of the diblock, as the copolymer concentration increases. Quantitative analysis of the speed of cell migration is correlated to the lateral length scale of the surface.

  5. Modeling collective cell motility

    NASA Astrophysics Data System (ADS)

    Rappel, Wouter-Jan

    Eukaryotic cells often move in groups, a critical aspect of many biological and medical processes including wound healing, morphogenesis and cancer metastasis. Modeling can provide useful insights into the fundamental mechanisms of collective cell motility. Constructing models that incorporate the physical properties of the cells, however, is challenging. Here, I discuss our efforts to build a comprehensive cell motility model that includes cell membrane properties, cell-substrate interactions, cell polarity, and cell-cell interaction. The model will be applied to a variety of systems, including motion on micropatterned substrates and the migration of border cells in Drosophila. This work was supported by NIH Grant No. P01 GM078586 and NSF Grant No. 1068869.

  6. Huaier Aqueous Extract Inhibits Ovarian Cancer Cell Motility via the AKT/GSK3β/β-Catenin Pathway

    PubMed Central

    Jia, Nan; Yu, Yinhua; Hua, Keqin; Feng, Weiwei

    2013-01-01

    Traditional Chinese medicine has gained popularity due to its ability to kill tumor cells. Recently, the apoptotic and anti-angiogenic effects of Trametes robiniophila murr (Huaier) have been investigated. The aim of this study was to investigate its effect on cell mobility and tumor growth in ovarian cancer. Cell viability and motility were measured using SRB, scratch and migration assays. Cell apoptosis was analysed by annexin V/PI staining. Using a reverse-phase protein array (RPPA) assay, we analyzed the levels of 153 proteins and/or phosphorylations in Huaier-treated and untreated cells. Huaier inhibited cell viability and induced both early and late apoptosis in SKOV3, SKOV3.ip1 and Hey cells in a time- and dose-dependent manner. Cell invasiveness and migration were also suppressed significantly. The RPPA results showed significant differences (of at least 30%; P <0.05) in the levels of 7 molecules in SKOV3 cells and 10 in SKOV3.ip1 cells between the untreated and treated cells. Most of the molecules identified play roles in cell proliferation, apoptosis or cell adhesion/invasion. Western blot analysis further validated that Huaier treatment resulted in decreased AKT phosphorylation, enhanced expression of total GSK3β, inhibition of the phosphorylation of GSK3β on S9, reduction of both cytoplasmic β-catenin expression and nuclear β-catenin translocation, and transcriptional repression of several Wnt/β-catenin target genes (DIXDC1, LRP6, WNT5A, and cyclin D1). After knocking down GSK3β, β-catenin expression could not be inhibited by Huaier. Finally, Huaier inhibited the growth of ovarian tumor xenografts in vivo. These studies indicate that Huaier inhibits tumor cell mobility in ovarian cancer via the AKT/GSK3β/β-catenin signaling pathway. PMID:23667667

  7. Overexpression of WISP-1 down-regulated motility and invasion of lung cancer cells through inhibition of Rac activation.

    PubMed

    Soon, Lilian L; Yie, Ting-An; Shvarts, Anita; Levine, Arnold J; Su, Fei; Tchou-Wong, Kam-Meng

    2003-03-28

    Wnt-induced-secreted-protein-1 (WISP-1) is a cysteine-rich, secreted factor belonging to the CCN family. These proteins have been implicated in the inhibition of metastasis; however, the mechanisms involved have not been described. We demonstrated that overexpression of WISP-1 in H460 lung cancer cells inhibited lung metastasis and in vitro cell invasion and motility. We investigated the possibility that WISP-1 may regulate activation of Rac, a small GTPase important for cytoskeletal reorganizations during motility. In an indirect assay, WISP-1-expressing cells exhibited marked reduction in Rac activation compared with control cells. Blocking antibodies to alpha(v)beta(5) and alpha(1) integrins restored Rac activation in WISP-1 cells, suggesting that the inhibitory effect of WISP-1 on Rac lies downstream of integrins. Constitutively activated Rac mutant (RacG12V) was transfected into WISP-1 cells to restore Rac activation and these WISP-1/RacG12V transfectants were used for further studies. We performed microarray and real-time PCR analyses to identify genes involved in invasion that may be differentially regulated by WISP-1. Here, we showed decreased expression of metalloproteinase-1 (MMP-1) in WISP-1 cells compared with controls but increased expression in WISP-1/RacG12V cells. In an invasion assay across collagen I, an MMP-1 target matrix, WISP-1 cells were significantly less invasive compared with controls, whereas WISP-1/RacG12V cells showed elevated invasion levels. This work illustrates a negatively regulated pathway by WISP-1 involving integrins and Rac in the down-regulation of invasion.

  8. Angiopoietin-like protein 1 suppresses SLUG to inhibit cancer cell motility.

    PubMed

    Kuo, Tsang-Chih; Tan, Ching-Ting; Chang, Yi-Wen; Hong, Chih-Chen; Lee, Wei-Jiunn; Chen, Min-Wei; Jeng, Yung-Ming; Chiou, Jean; Yu, Pei; Chen, Pai-Sheng; Wang, Ming-Yang; Hsiao, Michael; Su, Jen-Liang; Kuo, Min-Liang

    2013-03-01

    Angiopoietin-like protein 1 (ANGPTL1) is a potent regulator of angiogenesis. Growing evidence suggests that ANGPTL family proteins not only target endothelial cells but also affect tumor cell behavior. In a screen of 102 patients with lung cancer, we found that ANGPTL1 expression was inversely correlated with invasion, lymph node metastasis, and poor clinical outcomes. ANGPTL1 suppressed the migratory, invasive, and metastatic capabilities of lung and breast cancer cell lines in vitro and reduced metastasis in mice injected with cancer cell lines overexpressing ANGPTL1. Ectopic expression of ANGPTL1 suppressed the epithelial-to-mesenchymal transition (EMT) by reducing the expression of the zinc-finger protein SLUG. A microRNA screen revealed that ANGPTL1 suppressed SLUG by inducing expression of miR-630 in an integrin α(1)β(1)/FAK/ERK/SP1 pathway-dependent manner. These results demonstrate that ANGPTL1 represses lung cancer cell motility by abrogating the expression of the EMT mediator SLUG.

  9. Downregulation of VANGL1 Inhibits Cellular Invasion Rather than Cell Motility in Hepatocellular Carcinoma Cells Without Stimulation

    PubMed Central

    Toylu, Asli; Atabey, Nese; Sercan, Zeynep; Sakizli, Meral

    2015-01-01

    Aims: The Wnt planar cell polarity (PCP) pathway is one of the Wnt pathways which plays a critical role in cell proliferation and fate. The VANGL1 protein is one of Wnt-PCP pathway components. It is known that Wnt-PCP pathway has major roles in cell motility but its role in hepatocellular carcinoma (HCC) progression through invasion and metastasis needs to be clarified. Methods: We silenced VANGL1 gene expression in the HepG2 HCC cell line by stable transfection with a vector containing siRNA template for VANGL1 and investigated the change in cell invasion and motility. Results: Transfected cells with the siRNA template showed significantly suppressed invasive capacity when compared to controls although cellular motility was only slightly affected. Conclusion: Our study showed a basal role for VANGL1 with respect to the invasive capacity of HCC cells. This suggests that the Wnt-PCP pathway may play a role in progression of HCC through cellular invasion but further studies are needed to clarify its role in cell motility. PMID:25874746

  10. PKA and CDK5 can phosphorylate specific serines on the intracellular domain of podoplanin (PDPN) to inhibit cell motility.

    PubMed

    Krishnan, Harini; Retzbach, Edward P; Ramirez, Maria I; Liu, Tong; Li, Hong; Miller, W Todd; Goldberg, Gary S

    2015-07-01

    Podoplanin (PDPN) is a transmembrane glycoprotein that promotes tumor cell migration, invasion, and cancer metastasis. In fact, PDPN expression is induced in many types of cancer. Thus, PDPN has emerged as a functionally relevant cancer biomarker and chemotherapeutic target. PDPN contains 2 intracellular serine residues that are conserved between species ranging from mouse to humans. Recent studies indicate that protein kinase A (PKA) can phosphorylate PDPN in order to inhibit cell migration. However, the number and identification of specific residues phosphorylated by PKA have not been defined. In addition, roles of other kinases that may phosphorylate PDPN to control cell migration have not been investigated. We report here that cyclin dependent kinase 5 (CDK5) can phosphorylate PDPN in addition to PKA. Moreover, results from this study indicate that PKA and CDK5 cooperate to phosphorylate PDPN on both intracellular serine residues to decrease cell motility. These results provide new insight into PDPN phosphorylation dynamics and the role of PDPN in cell motility. Understanding novel mechanisms of PDPN intracellular signaling could assist with designing novel targeted chemotherapeutic agents and procedures. PMID:25959509

  11. N-(2-methyl-indol-1H-5-yl)-1-naphthalenesulfonamide: A novel reversible antimitotic agent inhibiting cancer cell motility.

    PubMed

    Aceves-Luquero, Clara; Galiana-Roselló, Cristina; Ramis, Guillem; Villalonga-Planells, Ruth; García-España, Enrique; Fernández de Mattos, Silvia; Peláez, Rafael; Llinares, José M; González-Rosende, M Eugenia; Villalonga, Priam

    2016-09-01

    A series of compounds containing the sulfonamide scaffold were synthesized and screened for their in vitro anticancer activity against a representative panel of human cancer cell lines, leading to the identification of N-(2-methyl-1H-indol-5-yl)-1-naphthalenesulfonamide (8e) as a compound showing a remarkable activity across the panel, with IC50 values in the nanomolar-to-low micromolar range. Cell cycle distribution analysis revealed that 8e promoted a severe G2/M arrest, which was followed by cellular senescence as indicated by the detection of senescence-associated β-galactosidase (SA-β-gal) in 8e-treated cells. Prolonged 8e treatment also led to the onset of apoptosis, in correlation with the detection of increased Caspase 3/7 activities. Despite increasing γ-H2A.X levels, a well-established readout for DNA double-strand breaks, in vitro DNA binding studies with 8e did not support interaction with DNA. In agreement with this, 8e failed to activate the cellular DNA damage checkpoint. Importantly, tubulin staining showed that 8e promoted a severe disorganization of microtubules and mitotic spindle formation was not detected in 8e-treated cells. Accordingly, 8e inhibited tubulin polymerization in vitro in a dose-dependent manner and was also able to robustly inhibit cancer cell motility. Docking analysis revealed a compatible interaction with the colchicine-binding site of tubulin. Remarkably, these cellular effects were reversible since disruption of treatment resulted in the reorganization of microtubules, cell cycle re-entry and loss of senescent markers. Collectively, our data suggest that this compound may be a promising new anticancer agent capable of both reducing cancer cell growth and motility. PMID:27349984

  12. Chlorotoxin Fused to IgG-Fc Inhibits Glioblastoma Cell Motility via Receptor-Mediated Endocytosis.

    PubMed

    Kasai, Tomonari; Nakamura, Keisuke; Vaidyanath, Arun; Chen, Ling; Sekhar, Sreeja; El-Ghlban, Samah; Okada, Masashi; Mizutani, Akifumi; Kudoh, Takayuki; Murakami, Hiroshi; Seno, Masaharu

    2012-01-01

    Chlorotoxin is a 36-amino acid peptide derived from Leiurus quinquestriatus (scorpion) venom, which has been shown to inhibit low-conductance chloride channels in colonic epithelial cells. Chlorotoxin also binds to matrix metalloproteinase-2 and other proteins on glioma cell surfaces. Glioma cells are considered to require the activation of matrix metalloproteinase-2 during invasion and migration. In this study, for targeting glioma, we designed two types of recombinant chlorotoxin fused to human IgG-Fcs with/without a hinge region. Chlorotoxin fused to IgG-Fcs was designed as a dimer of 60 kDa with a hinge region and a monomer of 30 kDa without a hinge region. The monomeric and dimeric forms of chlorotoxin inhibited cell proliferation at 300 nM and induced internalization in human glioma A172 cells. The monomer had a greater inhibitory effect than the dimer; therefore, monomeric chlorotoxin fused to IgG-Fc was multivalently displayed on the surface of bionanocapsules to develop a drug delivery system that targeted matrix metalloproteinase-2. The target-dependent internalization of bionanocapsules in A172 cells was observed when chlorotoxin was displayed on the bionanocapsules. This study indicates that chlorotoxin fused to IgG-Fcs could be useful for the active targeting of glioblastoma cells.

  13. A Cronobacter turicensis O1 Antigen-Specific Monoclonal Antibody Inhibits Bacterial Motility and Entry into Epithelial Cells

    PubMed Central

    Lehner, Angelika; Dietrich, Richard; Kleinsteuber, Ina; Canals, Rocío; Zurfluh, Katrin; Weiner, Kerstin; Märtlbauer, Erwin

    2014-01-01

    Cronobacter turicensis is an opportunistic foodborne pathogen that can cause a rare but sometimes lethal infection in neonates. Little is known about the virulence mechanisms and intracellular lifestyle of this pathogen. In this study, we developed an IgG monoclonal antibody (MAb; MAb 2G4) that specifically recognizes the O1 antigen of C. turicensis cells. The antilipopolysaccharide antibody bound predominantly monovalently to the O antigen and reduced bacterial growth without causing cell agglutination. Furthermore, binding of the antibody to the O1 antigen of C. turicensis cells caused a significant reduction of the membrane potential which is required to energize flagellar rotation, accompanied by a decreased flagellum-based motility. These results indicate that binding of IgG to the O antigen of C. turicensis causes a direct antimicrobial effect. In addition, this feature of the antibody enabled new insight into the pathogenicity of C. turicensis. In a tissue culture infection model, pretreatment of C. turicensis with MAb 2G4 showed no difference in adhesion to human epithelial cells, whereas invasion of bacteria into Caco-2 cells was significantly inhibited. PMID:25534937

  14. Cell Proliferation and Motility Are Inhibited by G1 Phase Arrest in 15-kDa Selenoprotein-Deficient Chang Liver Cells

    PubMed Central

    Bang, Jeyoung; Huh, Jang Hoe; Na, Ji-Woon; Lu, Qiao; Carlson, Bradley A.; Tobe, Ryuta; Tsuji, Petra A.; Gladyshev, Vadim N.; Hatfield, Dolph L.; Lee, Byeong Jae

    2015-01-01

    The 15-kDa selenoprotein (Sep15) is a selenoprotein residing in the lumen of the endoplasmic reticulum (ER) and implicated in quality control of protein folding. Herein, we established an inducible RNAi cell line that targets Sep15 mRNA in Chang liver cells. RNAi-induced Sep15 deficiency led to inhibition of cell proliferation, whereas cell growth was resumed after removal of the knockdown inducer. Sep15-deficient cells were arrested at the G1 phase by upregulating p21 and p27, and these cells were also characterized by ER stress. In addition, Sep15 deficiency led to the relocation of focal adhesions to the periphery of the cell basement and to the decrease of the migratory and invasive ability. All these changes were reversible depending on Sep15 status. Rescuing the knockdown state by expressing a silent mutant Sep15 mRNA that is resistant to siRNA also reversed the phenotypic changes. Our results suggest that SEP15 plays important roles in the regulation of the G1 phase during the cell cycle as well as in cell motility in Chang liver cells, and that this selenoprotein offers a novel functional link between the cell cycle and cell motility. PMID:25728752

  15. Shape determination in motile cells

    NASA Astrophysics Data System (ADS)

    Mogilner, Alex

    2010-03-01

    Flat, simple shaped, rapidly gliding fish keratocyte cell is the model system of choice to study cell motility. The cell motile appendage, lamellipod, has a characteristic bent-rectangular shape. Recent experiments showed that the lamellipodial geometry is tightly correlated with cell speed and with actin dynamics. These quantitative data combined with computational modeling suggest that a model for robust actin treadmill inside the 'unstretchable membrane bag'. According to this model, a force balance between membrane tension and growing and pushing actin network distributed unevenly along the cell periphery can explain the cell shape and motility. However, when adhesion of the cell to the surface weakens, the actin dynamics become less regular, and myosin-powered contraction starts playing crucial role in stabilizing the cell shape. I will illustrate how the combination of theoretical and experimental approaches helped to unravel the keratocyte motile behavior.

  16. Inhibition of breast cancer cell motility with a non-cyclooxygenase inhibitory derivative of sulindac by suppressing TGFβ/miR-21 signaling

    PubMed Central

    Ma, Ruixia; Feng, Xiangling; Li, Wei; Piazza, Gary A.; Xi, Yaguang

    2016-01-01

    Compelling efficacy on intervention of tumorigenesis by nonsteroidal anti-inflammatory drugs (NSAIDs) has been documented intensively. However, the toxicities related to cyclooxygenase (COX) inhibition resulting in suppression of physiologically important prostaglandins limit their clinical use for human cancer chemoprevention. A novel derivative of the NSAID sulindac sulfide (SS), referred as sulindac sulfide amide (SSA), was recently developed, which lacks COX inhibitory activity, yet shows greater suppressive effect than SS on growth of various cancer cells. In this study, we focus on the inhibitory activity of SSA on breast tumor cell motility, which has not been studied previously. Our results show that SSA treatment at non-cytotoxic concentrations can specifically reduce breast tumor cell motility without influencing tumor cell growth, and the mechanism of action involves the suppression of TGFβ signaling by directly blocking Smad2/3 phosphorylation. Moreover, miR-21, a well-documented oncogenic miRNA for promoting tumor cell metastasis, was also found to be involved in inhibitory activity of SSA in breast tumor cell motility through the modulation of TGFβ pathway. In conclusion, we demonstrate that a non-COX inhibitory derivative of sulindac can inhibit breast tumor metastasis by a mechanism involving the TGFβ/miR-21 signaling axis. PMID:26769851

  17. Inhibition of breast cancer cell motility with a non-cyclooxygenase inhibitory derivative of sulindac by suppressing TGFβ/miR-21 signaling.

    PubMed

    Yi, Bin; Chang, Hong; Ma, Ruixia; Feng, Xiangling; Li, Wei; Piazza, Gary A; Xi, Yaguang

    2016-02-16

    Compelling efficacy on intervention of tumorigenesis by nonsteroidal anti-inflammatory drugs (NSAIDs) has been documented intensively. However, the toxicities related to cyclooxygenase (COX) inhibition resulting in suppression of physiologically important prostaglandins limit their clinical use for human cancer chemoprevention. A novel derivative of the NSAID sulindac sulfide (SS), referred as sulindac sulfide amide (SSA), was recently developed, which lacks COX inhibitory activity, yet shows greater suppressive effect than SS on growth of various cancer cells. In this study, we focus on the inhibitory activity of SSA on breast tumor cell motility, which has not been studied previously. Our results show that SSA treatment at non-cytotoxic concentrations can specifically reduce breast tumor cell motility without influencing tumor cell growth, and the mechanism of action involves the suppression of TGFβ signaling by directly blocking Smad2/3 phosphorylation. Moreover, miR-21, a well-documented oncogenic miRNA for promoting tumor cell metastasis, was also found to be involved in inhibitory activity of SSA in breast tumor cell motility through the modulation of TGFβ pathway. In conclusion, we demonstrate that a non-COX inhibitory derivative of sulindac can inhibit breast tumor metastasis by a mechanism involving the TGFβ/miR-21 signaling axis.

  18. Toxigenic diversity of two different RAPD groups of Stachybotrys chartarum isolates analyzed by potential for trichothecene production and for boar sperm cell motility inhibition.

    PubMed

    Peltola, J; Niessen, L; Nielsen, K F; Jarvis, B B; Andersen, B; Salkinoja-Salonen, M; Möller, E M

    2002-11-01

    Thirty-one isolates of Stachybotrys chartarum from indoor and outdoor environments were analyzed for the presence of the trichodiene synthase (Tri5) gene, trichothecenes, boar sperm cell motility inhibition, and randomly amplified polymorphic DNA banding patterns (RAPDs). Twenty-two S. chartarum isolates tested positive for the Tri5 gene and nine were negative when tested using novel Tri5 gene-specific PCR primer pair. The Tri5 gene positive isolates contained satratoxins (five isolates) or the simple trichothecene, trichodermol (11 isolates). The Tri5 gene negative isolates did not produce satratoxins or trichodermol. Nineteen S. chartarum isolates, distributed among the Tri5 gene negative and positive groups, inhibited boar spermatozoan motility at concentrations of < or = 60 microg of crude cell extract/mL. The inhibition of motility was independent of satratoxins or atranones. Unweighted pair group method of arithmetic averages (UPGMA) cluster analysis of RAPD fragments clustered the 31 S. chartarum isolates in two distinct groups designated as RAPD groups 1 and 2. The grouping of S. chartarum isolates obtained by UPGMA cluster analysis of RAPD fragments was identical to the grouping obtained by Tri5 gene-specific PCR. This indicates that the S. chartarum isolates belonging to different groups were genetically distinct in a much wider area than just the Tri5 gene. PMID:12556129

  19. 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

  20. High mobility group Box-1 inhibits cancer cell motility and metastasis by suppressing activation of transcription factor CREB and nWASP expression.

    PubMed

    Zuo, Zhenghong; Che, Xun; Wang, Yulei; Li, Bowen; Li, Jingxia; Dai, Wei; Lin, Charles P; Huang, Chuanshu

    2014-09-15

    The ability to metastasize is a hallmark of malignant tumors, and metastasis is the principal cause of death of cancer patients. The High Mobility Group Box-1 (HMGB1) is a multifunction protein that serves as both a chromatin protein and an extracellular signaling molecule. Our current study demonstrated a novel mechanism of HMGB1 in the regulation of cancer cell actin polymerization, cell skeleton formation, cancer cell motility and metastasis. We found that knockdown of HMGB1 in human lung cancer A549 cells significantly increased cell β-actin polymerization, cell skeleton formation, cancer cell migration and invasion in vitro, as well as metastasis in vivo. And this increase could be inhibited by treatment of HMGB1 knockdown cells with recombinant human HMGB1. Further studies discovered that HMGB1 suppressed phosphorylation, nuclear translocation, and activation of CREB, by inhibiting nuclear translocation of PKA catalytic subunit. This reduces nWASP mRNA transcription and expression, further impairing cancer cell motility. Our findings on the novel mechanism underlying the HMGB1 anti-metastatic effect on cancer provides significant insight into the understanding of the nature of HMGB1 in cancer invasion and metastasis, further serving as key information for utilization of HMGB1 and its regulated downstream components as new targets for cancer therapy. PMID:25277185

  1. Activation of β2-adrenergic receptor by (R,R’)-4’-methoxy-1-naphthylfenoterol inhibits proliferation and motility of melanoma cells

    PubMed Central

    Wnorowski, Artur; Sadowska, Mariola; Paul, Rajib K.; Singh, Nagendra S.; Boguszewska-Czubara, Anna; Jimenez, Lucita; Abdelmohsen, Kotb; Toll, Lawrence; Jozwiak, Krzysztof; Bernier, Michel; Wainer, Irving W.

    2015-01-01

    (R,R’)-4’-methoxy-1-naphthylfenoterol [(R,R’)-MNF] is a highly-selective β2 adrenergic receptor (β2-AR) agonist. Incubation of a panel of human-derived melanoma cell lines with (R,R’)-MNF resulted in a dose- and time-dependent inhibition of motility as assessed by in vitro wound healing and xCELLigence migration and invasion assays. Activity of (R,R’)-MNF positively correlated with the β2-AR expression levels across tested cell lines. The anti-motility activity of (R,R’)-MNF was inhibited by the β2-AR antagonist ICI-118,551 and the protein kinase A inhibitor H-89. The adenylyl cyclase activator forskolin and the phosphodiesterase 4 inhibitor Ro 20–1724 mimicked the ability of (R,R’)-MNF to inhibit migration of melanoma cell lines in culture, highlighting the importance of cAMP for this phenomenon. (R,R’)-MNF caused significant inhibition of cell growth in β2-AR-expressing cells as monitored by radiolabeled thymidine incorporation and xCELLigence system. The MEK/ERK cascade functions in cellular proliferation, and constitutive phosphorylation of MEK and ERK at their active sites was significantly reduced upon β2-AR activation with (R,R’)-MNF. Protein synthesis was inhibited concomitantly both with increased eEF2 phosphorylation and lower expression of tumor cell regulators, EGF receptors, cyclin A and MMP-9. Taken together, these results identified β2-AR as a novel potential target for melanoma management, and (R,R’)-MNF as an efficient trigger of anti-tumorigenic cAMP/PKA-dependent signaling in β2-AR-expressing lesions. PMID:25703025

  2. Hyaluronan stimulates pancreatic cancer cell motility

    PubMed Central

    Cheng, Xiao-Bo; Kohi, Shiro; Koga, Atsuhiro; Hirata, Keiji; Sato, Norihiro

    2016-01-01

    Hyaluronan (HA) accumulates in pancreatic ductal adenocarcinoma (PDAC), but functional significance of HA in the aggressive phenotype remains unknown. We used different models to investigate the effect of HA on PDAC cell motility by wound healing and transwell migration assay. Changes in cell motility were examined in 8 PDAC cell lines in response to inhibition of HA production by treatment with 4-methylumbelliferone (4-MU) and to promotion by treatment with 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or by co-culture with tumor-derived stromal fibroblasts. We also investigated changes in cell motility by adding exogenous HA. Additionally, mRNA expressions of hyaluronan synthases and hyaluronidases were examined using real time RT-PCR. Inhibition of HA by 4-MU significantly decreased the migration, whereas promotion of HA by TPA or co-culture with tumor-derived fibroblasts significantly increased the migration of PDAC cells. The changes in HA production by these treatments tended to be associated with changes in HAS3 mRNA expression. Furthermore, addition of exogenous HA, especially low-molecular-weight HA, significantly increased the migration of PDAC cells. These findings suggest that HA stimulates PDAC cell migration and thus represents an ideal therapeutic target to prevent invasion and metastasis. PMID:26684359

  3. Elastic mismatch enhances cell motility

    NASA Astrophysics Data System (ADS)

    Bresler, Yony; Palmieri, Benoit; Grant, Martin

    In recent years, the study of physics phenomena in cancer has drawn considerable attention. In cancer metastasis, a soft cancer cell leaves the tumor, and must pass through the endothelium before reaching the bloodstream. Using a phase-field model we have shown that the elasticity mismatch between cells alone is sufficient to enhance the motility of thesofter cancer cell by means of bursty migration, in agreement with experiment. We will present further characterization of these behaviour, as well as new possible applications for this model.

  4. Rgnef (p190RhoGEF) Knockout Inhibits RhoA Activity, Focal Adhesion Establishment, and Cell Motility Downstream of Integrins

    PubMed Central

    Miller, Nichol L. G.; Lawson, Christine; Chen, Xiao Lei; Lim, Ssang-Taek; Schlaepfer, David D.

    2012-01-01

    Background Cell migration is a highly regulated process that involves the formation and turnover of cell-matrix contact sites termed focal adhesions. Rho-family GTPases are molecular switches that regulate actin and focal adhesion dynamics in cells. Guanine nucleotide exchange factors (GEFs) activate Rho-family GTPases. Rgnef (p190RhoGEF) is a ubiquitous 190 kDa GEF implicated in the control of colon carcinoma and fibroblast cell motility. Principal Findings Rgnef exon 24 floxed mice (Rgnefflox) were created and crossed with cytomegalovirus (CMV)-driven Cre recombinase transgenic mice to inactivate Rgnef expression in all tissues during early development. Heterozygous RgnefWT/flox (Cre+) crosses yielded normal Mendelian ratios at embryonic day 13.5, but Rgnefflox/flox (Cre+) mice numbers at 3 weeks of age were significantly less than expected. Rgnefflox/flox (Cre+) (Rgnef−/−) embryos and primary mouse embryo fibroblasts (MEFs) were isolated and verified to lack Rgnef protein expression. When compared to wildtype (WT) littermate MEFs, loss of Rgnef significantly inhibited haptotaxis migration, wound closure motility, focal adhesion number, and RhoA GTPase activation after fibronectin-integrin stimulation. In WT MEFs, Rgnef activation occurs within 60 minutes upon fibronectin plating of cells associated with RhoA activation. Rgnef−/− MEF phenotypes were rescued by epitope-tagged Rgnef re-expression. Conclusions Rgnef−/− MEF phenotypes were due to Rgnef loss and support an essential role for Rgnef in RhoA regulation downstream of integrins in control of cell migration. PMID:22649559

  5. All-trans-retinoic Acid Modulates the Plasticity and Inhibits the Motility of Breast Cancer Cells: ROLE OF NOTCH1 AND TRANSFORMING GROWTH FACTOR (TGFβ).

    PubMed

    Zanetti, Adriana; Affatato, Roberta; Centritto, Floriana; Fratelli, Maddalena; Kurosaki, Mami; Barzago, Maria Monica; Bolis, Marco; Terao, Mineko; Garattini, Enrico; Paroni, Gabriela

    2015-07-17

    All-trans-retinoic acid (ATRA) is a natural compound proposed for the treatment/chemoprevention of breast cancer. Increasing evidence indicates that aberrant regulation of epithelial-to-mesenchymal transition (EMT) is a determinant of the cancer cell invasive and metastatic behavior. The effects of ATRA on EMT are largely unknown. In HER2-positive SKBR3 and UACC812 cells, showing co-amplification of the ERBB2 and RARA genes, ATRA activates a RARα-dependent epithelial differentiation program. In SKBR3 cells, this causes the formation/reorganization of adherens and tight junctions. Epithelial differentiation and augmented cell-cell contacts underlie the anti-migratory action exerted by the retinoid in cells exposed to the EMT-inducing factors EGF and heregulin-β1. Down-regulation of NOTCH1, an emerging EMT modulator, is involved in the inhibition of motility by ATRA. Indeed, the retinoid blocks NOTCH1 up-regulation by EGF and/or heregulin-β1. Pharmacological inhibition of γ-secretase and NOTCH1 processing also abrogates SKBR3 cell migration. Stimulation of TGFβ contributes to the anti-migratory effect of ATRA. The retinoid switches TGFβ from an EMT-inducing and pro-migratory determinant to an anti-migratory mediator. Inhibition of the NOTCH1 pathway not only plays a role in the anti-migratory action of ATRA; it is relevant also for the anti-proliferative activity of the retinoid in HCC1599 breast cancer cells, which are addicted to NOTCH1 for growth/viability. This effect is enhanced by the combination of ATRA and the γ-secretase inhibitor N-(N-(3,5-difluorophenacetyl)-l-alanyl)-S-phenylglycine t-butyl ester, supporting the concept that the two compounds act at the transcriptional and post-translational levels along the NOTCH1 pathway. PMID:26018078

  6. All-trans-retinoic Acid Modulates the Plasticity and Inhibits the Motility of Breast Cancer Cells: ROLE OF NOTCH1 AND TRANSFORMING GROWTH FACTOR (TGFβ).

    PubMed

    Zanetti, Adriana; Affatato, Roberta; Centritto, Floriana; Fratelli, Maddalena; Kurosaki, Mami; Barzago, Maria Monica; Bolis, Marco; Terao, Mineko; Garattini, Enrico; Paroni, Gabriela

    2015-07-17

    All-trans-retinoic acid (ATRA) is a natural compound proposed for the treatment/chemoprevention of breast cancer. Increasing evidence indicates that aberrant regulation of epithelial-to-mesenchymal transition (EMT) is a determinant of the cancer cell invasive and metastatic behavior. The effects of ATRA on EMT are largely unknown. In HER2-positive SKBR3 and UACC812 cells, showing co-amplification of the ERBB2 and RARA genes, ATRA activates a RARα-dependent epithelial differentiation program. In SKBR3 cells, this causes the formation/reorganization of adherens and tight junctions. Epithelial differentiation and augmented cell-cell contacts underlie the anti-migratory action exerted by the retinoid in cells exposed to the EMT-inducing factors EGF and heregulin-β1. Down-regulation of NOTCH1, an emerging EMT modulator, is involved in the inhibition of motility by ATRA. Indeed, the retinoid blocks NOTCH1 up-regulation by EGF and/or heregulin-β1. Pharmacological inhibition of γ-secretase and NOTCH1 processing also abrogates SKBR3 cell migration. Stimulation of TGFβ contributes to the anti-migratory effect of ATRA. The retinoid switches TGFβ from an EMT-inducing and pro-migratory determinant to an anti-migratory mediator. Inhibition of the NOTCH1 pathway not only plays a role in the anti-migratory action of ATRA; it is relevant also for the anti-proliferative activity of the retinoid in HCC1599 breast cancer cells, which are addicted to NOTCH1 for growth/viability. This effect is enhanced by the combination of ATRA and the γ-secretase inhibitor N-(N-(3,5-difluorophenacetyl)-l-alanyl)-S-phenylglycine t-butyl ester, supporting the concept that the two compounds act at the transcriptional and post-translational levels along the NOTCH1 pathway.

  7. Integrin-free tetraspanin CD151 can inhibit tumor cell motility upon clustering and is a clinical indicator of prostate cancer progression

    PubMed Central

    Palmer, T. D.; Martínez, C. H.; Vasquez; Hebron, K.; Jones-Paris, C.; Arnold, S.A.; Chan, S.M.; Chalasani, V.; Gomez-Lemus, J.A.; Williams, A.K.; Chin, J.L.; Giannico, G.A.; Ketova, T.; Lewis, J.D.; Zijlstra, A.

    2013-01-01

    Normal physiology relies on the organization of transmembrane proteins by molecular scaffolds, such as tetraspanins. Oncogenesis frequently involves changes in their organization or expression. The tetraspanin CD151 is thought to contribute to cancer progression through direct interaction with the laminin-binding integrins α3β1 and α6β1. However, this interaction cannot explain the ability of CD151 to control migration in the absence of these integrins or on non-laminin substrates. We demonstrate that CD151 can regulate tumor cell migration without direct integrin binding and that integrin-free CD151 (CD151free) correlates clinically with tumor progression and metastasis. Clustering CD151free through its integrin-binding domain promotes accumulation in areas of cell-cell contact leading to enhanced adhesion and inhibition of tumor cell motility in vitro and in vivo. CD151free clustering is a strong regulator of motility even in the absence of α3 expression but requires PKCα, suggesting that CD151 can control migration independent of its integrin associations. The histological detection of CD151free in prostate cancer correlates with poor patient outcome. When CD151free is present, patients are more likely to recur after radical prostatectomy and progression to metastatic disease is accelerated. Multivariable analysis identifies CD151free as an independent predictor of survival. Moreover, the detection of CD151free can stratify survival among patients with elevated PSA. Cumulatively these studies demonstrate that a subpopulation of CD151 exists on the surface of tumor cells that can regulate migration independent of its integrin partner. The clinical correlation of CD151free with prostate cancer progression suggests that it may contribute to the disease and predict cancer progression. PMID:24220242

  8. SIRT1 inhibits the mouse intestinal motility and epithelial proliferation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    SIRT1 inhibits the mouse intestinal motility and epithelial proliferation. Sirtuin 1 (SIRT1), a NAD+-dependent histone deacetylase, is involved in a wide array of cellular processes, including glucose homeostasis, energy metabolism, proliferation and apoptosis, and immune response. However, it is un...

  9. [Sodium houttuyfonate inhibits virulence related motility of Pseudomonas aeruginosa].

    PubMed

    Wu, Da-qiang; Huang, Wei-feng; Duan, Qiang-jun; Cheng, Hui-juan; Wang, Chang-zhong

    2015-04-01

    Sodium houttuyfonate (SH) is a derivative of effective component of a Chinese material medica, Houttuynia cordata, which is applied in anti-infection of microorganism. But, the antimicrobial mechanisms of SH still remain unclear. Here, we firstly discovered that SH effectively inhibits the three types of virulence related motility of.Pseudomonas aeruginosa, i.e., swimming, twitching and swarming. The plate assay results showed that the inhibitory action of SH against swimming and twitching in 24 h and swarming in 48 h is dose-dependent; and bacteria nearly lost all of the motile activities under the concentration of 1 x minimum inhibitory concentration (MIC) (512 mg x L(-1) same as azithromycin positive group (1 x MIC, 16 mg x L(-1)). Furthermore, we found that the expression of structural gene flgB and pilG is down-regulated by SH, which implies that inhibitory mechanism of SH against motility of P. aeruginosa may be due to the inhibition of flagella and pili bioformation of P. aeruginosa by SR Therefore, our presented results firstly demonstrate that SH effectively inhibits the motility activities of P. aeruginosa, and suggest that SH could be a promising antipseudomonas agents in clinic. PMID:26281603

  10. The motility of Chara corallina myosin was inhibited reversibly by 2,3-butanedione monoxime (BDM).

    PubMed

    Funaki, Keisuke; Nagata, Ayumi; Akimoto, Youka; Shimada, Kiyo; Ito, Kohji; Yamamoto, Keiichi

    2004-09-01

    We studied the effects of 2,3-butanedione monoxime (BDM) on the cytoplasmic streaming of Chara corallina and on the motility of myosin prepared from the same plant to examine whether this reagent really affects the plant class XI myosin. It was found that BDM inhibited both cytoplasmic streaming and the motility of myosin at a very similar concentration range (10-100 mM). BDM introduced directly into tonoplast-free cells also inhibited cytoplasmic streaming. These results suggested that effect of BDM on cytoplasmic streaming was exerted through myosin and not through ion channels at least in Chara corallina, though a very high concentration of BDM was required.

  11. Inhibition of Pseudomonas aeruginosa Swarming Motility by 1-Naphthol and Other Bicyclic Compounds Bearing Hydroxyl Groups

    PubMed Central

    Oura, Hiromu; Tashiro, Yosuke; Toyofuku, Masanori; Ueda, Kousetsu; Kiyokawa, Tatsunori; Ito, Satoshi; Takahashi, Yurika; Lee, Seunguk; Nojiri, Hideaki; Nakajima-Kambe, Toshiaki; Uchiyama, Hiroo; Futamata, Hiroyuki

    2015-01-01

    Many bacteria convert bicyclic compounds, such as indole and naphthalene, to oxidized compounds, including hydroxyindoles and naphthols. Pseudomonas aeruginosa, a ubiquitous bacterium that inhabits diverse environments, shows pathogenicity against animals, plants, and other microorganisms, and increasing evidence has shown that several bicyclic compounds alter the virulence-related phenotypes of P. aeruginosa. Here, we revealed that hydroxyindoles (4- and 5-hydroxyindoles) and naphthalene derivatives bearing hydroxyl groups specifically inhibit swarming motility but have minor effects on other motilities, including swimming and twitching, in P. aeruginosa. Further analyses using 1-naphthol showed that this effect is also associated with clinically isolated hyperswarming P. aeruginosa cells. Swarming motility is associated with the dispersion of cells from biofilms, and the addition of 1-naphthol maintained biofilm biomass without cell dispersion. We showed that this 1-naphthol-dependent swarming inhibition is independent of changes of rhamnolipid production and the intracellular level of signaling molecule cyclic-di-GMP (c-di-GMP). Transcriptome analyses revealed that 1-naphthol increases gene expression associated with multidrug efflux and represses gene expression associated with aerotaxis and with pyochelin, flagellar, and pilus synthesis. In the present study, we showed that several bicyclic compounds bearing hydroxyl groups inhibit the swarming motility of P. aeruginosa, and these results provide new insight into the chemical structures that inhibit the specific phenotypes of P. aeruginosa. PMID:25681177

  12. Inhibition of Pseudomonas aeruginosa swarming motility by 1-naphthol and other bicyclic compounds bearing hydroxyl groups.

    PubMed

    Oura, Hiromu; Tashiro, Yosuke; Toyofuku, Masanori; Ueda, Kousetsu; Kiyokawa, Tatsunori; Ito, Satoshi; Takahashi, Yurika; Lee, Seunguk; Nojiri, Hideaki; Nakajima-Kambe, Toshiaki; Uchiyama, Hiroo; Futamata, Hiroyuki; Nomura, Nobuhiko

    2015-04-01

    Many bacteria convert bicyclic compounds, such as indole and naphthalene, to oxidized compounds, including hydroxyindoles and naphthols. Pseudomonas aeruginosa, a ubiquitous bacterium that inhabits diverse environments, shows pathogenicity against animals, plants, and other microorganisms, and increasing evidence has shown that several bicyclic compounds alter the virulence-related phenotypes of P. aeruginosa. Here, we revealed that hydroxyindoles (4- and 5-hydroxyindoles) and naphthalene derivatives bearing hydroxyl groups specifically inhibit swarming motility but have minor effects on other motilities, including swimming and twitching, in P. aeruginosa. Further analyses using 1-naphthol showed that this effect is also associated with clinically isolated hyperswarming P. aeruginosa cells. Swarming motility is associated with the dispersion of cells from biofilms, and the addition of 1-naphthol maintained biofilm biomass without cell dispersion. We showed that this 1-naphthol-dependent swarming inhibition is independent of changes of rhamnolipid production and the intracellular level of signaling molecule cyclic-di-GMP (c-di-GMP). Transcriptome analyses revealed that 1-naphthol increases gene expression associated with multidrug efflux and represses gene expression associated with aerotaxis and with pyochelin, flagellar, and pilus synthesis. In the present study, we showed that several bicyclic compounds bearing hydroxyl groups inhibit the swarming motility of P. aeruginosa, and these results provide new insight into the chemical structures that inhibit the specific phenotypes of P. aeruginosa. PMID:25681177

  13. 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

  14. Octreotide acetate inhibits motility in the rabbit distal colon.

    PubMed

    John, K D; Ballantyne, G H; Modlin, I M

    1997-01-01

    Octreotide, the long-acting somatostatin analogue, has been reported to modulate gastrointestinal motility in both animals and humans. A role in colonic peristalsis and a possible clinical application in common disorders, such as chronic constipation and irritable bowel syndrome, have not been evaluated. It has been previously suggested that octreotide promotes the descending relaxation of the peristaltic reflex arc. We hypothesized that this effect may involve inhibition of the motility index (MI) of the distal colon. To test this proposal, we studied peristalsis in isolated rabbit colons and also in the intact distal colons of anesthetized rabbits undergoing octreotide administration. Left colons of New Zealand white rabbits were harvested, placed in an isolated organ chamber and perfused with Krebs-Ringer bicarbonate solution via the inferior mesenteric artery. In a separate preparation, the colons were left in situ. Motility was quantified with a 6-port continuous infusion manometry catheter. The MI (mm Hg/min) was calculated by integration of the area of the digitalized signal (8/s), which reflected high-pressure peaks of different magnitudes. High-pressure waves were defined as > 20 mm Hg. Octreotide was infused via the inferior mesenteric artery in the isolated specimen or the lateral ear vein in the anesthetized animals in concentrations of 10(-12) to 10(-6) M. Octreotide inhibited high-pressure waves in a dose-dependent manner. These effects resulted in a decreased MI, with the maximum inhibition of 24.6% at 10(-11) M (p < 0.05 by ANOVA). At that concentration, the number of peaks > 20 mm Hg were reduced by 62.2%. The data indicate that octreotide decreases the MI by inhibition of high-pressure waves in the distal rabbit colon. These findings are consistent with the proposal that somatostatin may augment descending relaxation of the peristaltic reflex arc. This effect is independent of neural modulation.

  15. Hydrodynamic Contributions to Amoeboid Cell Motility

    NASA Astrophysics Data System (ADS)

    Lewis, Owen; Guy, Robert

    2012-11-01

    Understanding the methods by which cells move is a fundamental problem in modern biology. Recent evidence has shown that the fluid dynamics of cytoplasm can play a vital role in cellular motility. The slime mold Physarum polycephalum provides an excellent model organism for the study of amoeboid motion. In this research, we use a simply analytic model in conjuction with computational experiments to investigate intracellular fluid flow in a simple model of Physarum. Of particlar interest are stresses generated by cytoplasmic flow which may be used to aid in cellular motility. In our numerical model, the Immersed Boundary Method is used to account for such stresses. We investigate the relationship between contraction waves, flow waves, adhesion, and locomotive forces in an attempt to characterize conditions necessary to generate directed motion.

  16. Hydrodynamic Contributions to Amoeboid Cell Motility

    NASA Astrophysics Data System (ADS)

    Lewis, Owen; Guy, Robert

    2011-11-01

    Understanding the methods by which cells move is a fundamental problem in modern biology. Recent evidence has shown that the fluid dynamics of cytoplasm can play a vital role in cellular motility. The slime mold Physarum polycephalum provides an excellent model organism for the study of amoeboid motion. In this research, we use both analytic and computational models to investigate intracellular fluid flow in a simple model of Physarum. In both models, of we are specifically interested in stresses generated by cytoplasmic flow which act in the direction of cellular motility. In our numerical model, the Immersed Boundary Method is used to account for such stresses. We investigate the relationship between contraction waves, low waves and locomotive forces, and attempt characterize conditions necessary to generate directed motion.

  17. Quantum Dot-Based Cell Motility Assay

    SciTech Connect

    Gu, Weiwei; Pellegrino, Teresa; Parak Wolfgang J; Boudreau,Rosanne; Le Gros, Mark A.; Gerion, Daniele; Alivisatos, A. Paul; Larabell, Carolyn A.

    2005-06-06

    Because of their favorable physical and photochemical properties, colloidal CdSe/ZnS-semiconductor nanocrystals (commonly known as quantum dots) have enormous potential for use in biological imaging. In this report, we present an assay that uses quantum dots as markers to quantify cell motility. Cells that are seeded onto a homogeneous layer of quantum dots engulf and absorb the nanocrystals and, as a consequence, leave behind a fluorescence-free trail. By subsequently determining the ratio of cell area to fluorescence-free track area, we show that it is possible to differentiate between invasive and noninvasive cancer cells. Because this assay uses simple fluorescence detection, requires no significant data processing, and can be used in live-cell studies, it has the potential to be a powerful new tool for discriminating between invasive and noninvasive cancer cell lines or for studying cell signaling events involved in migration.

  18. Reduced Protein Synthesis Fidelity Inhibits Flagellar Biosynthesis and Motility.

    PubMed

    Fan, Yongqiang; Evans, Christopher R; Ling, Jiqiang

    2016-01-01

    Accurate translation of the genetic information from DNA to protein is maintained by multiple quality control steps from bacteria to mammals. Genetic and environmental alterations have been shown to compromise translational quality control and reduce fidelity during protein synthesis. The physiological impact of increased translational errors is not fully understood. While generally considered harmful, translational errors have recently been shown to benefit cells under certain stress conditions. In this work, we describe a novel regulatory pathway in which reduced translational fidelity downregulates expression of flagellar genes and suppresses bacterial motility. Electron microscopy imaging shows that the error-prone Escherichia coli strain lacks mature flagella. Further genetic analyses reveal that translational errors upregulate expression of a small RNA DsrA through enhancing its transcription, and deleting DsrA from the error-prone strain restores motility. DsrA regulates expression of H-NS and RpoS, both of which regulate flagellar genes. We demonstrate that an increased level of DsrA in the error-prone strain suppresses motility through the H-NS pathway. Our work suggests that bacteria are capable of switching on and off the flagellar system by altering translational fidelity, which may serve as a previously unknown mechanism to improve fitness in response to environmental cues. PMID:27468805

  19. Reduced Protein Synthesis Fidelity Inhibits Flagellar Biosynthesis and Motility

    PubMed Central

    Fan, Yongqiang; Evans, Christopher R.; Ling, Jiqiang

    2016-01-01

    Accurate translation of the genetic information from DNA to protein is maintained by multiple quality control steps from bacteria to mammals. Genetic and environmental alterations have been shown to compromise translational quality control and reduce fidelity during protein synthesis. The physiological impact of increased translational errors is not fully understood. While generally considered harmful, translational errors have recently been shown to benefit cells under certain stress conditions. In this work, we describe a novel regulatory pathway in which reduced translational fidelity downregulates expression of flagellar genes and suppresses bacterial motility. Electron microscopy imaging shows that the error-prone Escherichia coli strain lacks mature flagella. Further genetic analyses reveal that translational errors upregulate expression of a small RNA DsrA through enhancing its transcription, and deleting DsrA from the error-prone strain restores motility. DsrA regulates expression of H-NS and RpoS, both of which regulate flagellar genes. We demonstrate that an increased level of DsrA in the error-prone strain suppresses motility through the H-NS pathway. Our work suggests that bacteria are capable of switching on and off the flagellar system by altering translational fidelity, which may serve as a previously unknown mechanism to improve fitness in response to environmental cues. PMID:27468805

  20. 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.

  1. Correlation of cell membrane dynamics and cell motility

    PubMed Central

    2011-01-01

    Background Essential events of cell development and homeostasis are revealed by the associated changes of cell morphology and therefore have been widely used as a key indicator of physiological states and molecular pathways affecting various cellular functions via cytoskeleton. Cell motility is a complex phenomenon primarily driven by the actin network, which plays an important role in shaping the morphology of the cells. Most of the morphology based features are approximated from cell periphery but its dynamics have received none to scant attention. We aim to bridge the gap between membrane dynamics and cell states from the perspective of whole cell movement by identifying cell edge patterns and its correlation with cell dynamics. Results We present a systematic study to extract, classify, and compare cell dynamics in terms of cell motility and edge activity. Cell motility features extracted by fitting a persistent random walk were used to identify the initial set of cell subpopulations. We propose algorithms to extract edge features along the entire cell periphery such as protrusion and retraction velocity. These constitute a unique set of multivariate time-lapse edge features that are then used to profile subclasses of cell dynamics by unsupervised clustering. Conclusions By comparing membrane dynamic patterns exhibited by each subclass of cells, correlated trends of edge and cell movements were identified. Our findings are consistent with published literature and we also identified that motility patterns are influenced by edge features from initial time points compared to later sampling intervals. PMID:22372978

  2. 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

  3. 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.

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

    SciTech Connect

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

    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 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 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.

  5. [The flagellum: from cell motility to morphogenesis].

    PubMed

    Kohl, Linda; Robinson, Derrick; Bastin, Philippe

    2003-01-01

    Flagella and cilia are elaborate cytoskeletal structures conserved from protists to mammals, where they fulfil functions related to motility or sensitivity. We demonstrate a novel role for the flagellum in the control of cell morphogenesis and division of Trypanosoma brucei. To investigate flagellum functions, its formation was perturbed by inducible RNA interference silencing of components required for intraflagellar transport (IFT), a dynamic process necessary for flagellum assembly. First, we show that down-regulation of IFT leads to assembly of a shorter flagellum. Strikingly, cells with a shorter flagellum are smaller, with a direct correlation between flagellum length and cell size. Detailed morphogenetic analysis reveals that the tip of the new flagellum defines the point where cytokinesis is initiated. Furthermore, when new flagellum formation is completely blocked, non-flagellated cells are very short, lose their normal shape and polarity and fail to undergo cytokinesis. We show that flagellum elongation controls formation of cytoskeletal structures present in the cell body that act as molecular organisers of the cell.

  6. Computational and Modeling Strategies for Cell Motility

    NASA Astrophysics Data System (ADS)

    Wang, Qi; Yang, Xiaofeng; Adalsteinsson, David; Elston, Timothy C.; Jacobson, Ken; Kapustina, Maryna; Forest, M. Gregory

    A predictive simulation of the dynamics of a living cell remains a fundamental modeling and computational challenge. The challenge does not even make sense unless one specifies the level of detail and the phenomena of interest, whether the focus is on near-equilibrium or strongly nonequilibrium behavior, and on localized, subcellular, or global cell behavior. Therefore, choices have to be made clear at the outset, ranging from distinguishing between prokaryotic and eukaryotic cells, specificity within each of these types, whether the cell is "normal," whether one wants to model mitosis, blebs, migration, division, deformation due to confined flow as with red blood cells, and the level of microscopic detail for any of these processes. The review article by Hoffman and Crocker [48] is both an excellent overview of cell mechanics and an inspiration for our approach. One might be interested, for example, in duplicating the intricate experimental details reported in [43]: "actin polymerization periodically builds a mechanical link, the lamellipodium, connecting myosin motors with the initiation of adhesion sites, suggesting that the major functions driving motility are coordinated by a biomechanical process," or to duplicate experimental evidence of traveling waves in cells recovering from actin depolymerization [42, 35]. Modeling studies of lamellipodial structure, protrusion, and retraction behavior range from early mechanistic models [84] to more recent deterministic [112, 97] and stochastic [51] approaches with significant biochemical and structural detail. Recent microscopic-macroscopic models and algorithms for cell blebbing have been developed by Young and Mitran [116], which update cytoskeletal microstructure via statistical sampling techniques together with fluid variables. Alternatively, whole cell compartment models (without spatial details) of oscillations in spreading cells have been proposed [35, 92, 109] which show positive and negative feedback

  7. Patterns of periodic holes created by increased cell motility

    PubMed Central

    Chen, Ting-Hsuan; Guo, Chunyan; Zhao, Xin; Yao, Yucheng; Boström, Kristina I.; Wong, Margaret N.; Tintut, Yin; Demer, Linda L.; Ho, Chih-Ming; Garfinkel, Alan

    2012-01-01

    The reaction and diffusion of morphogens is a mechanism widely used to explain many spatial patterns in physics, chemistry and developmental biology. However, because experimental control is limited in most biological systems, it is often unclear what mechanisms account for the biological patterns that arise. Here, we study a biological model of cultured vascular mesenchymal cells (VMCs), which normally self-organize into aggregates that form into labyrinthine configurations. We use an experimental control and a mathematical model that includes reacting and diffusing morphogens and a third variable reflecting local cell density. With direct measurements showing that cell motility was increased ninefold and threefold by inhibiting either Rho kinase or non-muscle myosin-II, respectively, our experimental results and mathematical modelling demonstrate that increased motility alters the multicellular pattern of the VMC cultures, from labyrinthine to a pattern of periodic holes. These results suggest implications for the tissue engineering of functional replacements for trabecular or spongy tissue such as endocardium and bone. PMID:22649581

  8. LRRK2 G2019S mutation attenuates microglial motility by inhibiting focal adhesion kinase

    PubMed Central

    Choi, Insup; Kim, Beomsue; Byun, Ji-Won; Baik, Sung Hoon; Huh, Yun Hyun; Kim, Jong-Hyeon; Mook-Jung, Inhee; Song, Woo Keun; Shin, Joo-Ho; Seo, Hyemyung; Suh, Young Ho; Jou, Ilo; Park, Sang Myun; Kang, Ho Chul; Joe, Eun-Hye

    2015-01-01

    In response to brain injury, microglia rapidly extend processes that isolate lesion sites and protect the brain from further injury. Here we report that microglia carrying a pathogenic mutation in the Parkinson's disease (PD)-associated gene, G2019S-LRRK2 (GS-Tg microglia), show retarded ADP-induced motility and delayed isolation of injury, compared with non-Tg microglia. Conversely, LRRK2 knockdown microglia are highly motile compared with control cells. In our functional assays, LRRK2 binds to focal adhesion kinase (FAK) and phosphorylates its Thr–X–Arg/Lys (TXR/K) motif(s), eventually attenuating FAK activity marked by decreased pY397 phosphorylation (pY397). GS-LRRK2 decreases the levels of pY397 in the brain, microglia and HEK cells. In addition, treatment with an inhibitor of LRRK2 kinase restores pY397 levels, decreased pTXR levels and rescued motility of GS-Tg microglia. These results collectively suggest that G2019S mutation of LRRK2 may contribute to the development of PD by inhibiting microglial response to brain injury. PMID:26365310

  9. 3D timelapse analysis of muscle satellite cell motility.

    PubMed

    Siegel, Ashley L; Atchison, Kevin; Fisher, Kevin E; Davis, George E; Cornelison, D D W

    2009-10-01

    Skeletal muscle repair and regeneration requires the activity of satellite cells, a population of myogenic stem cells scattered throughout the tissue and activated to proliferate and differentiate in response to myotrauma or disease. While it seems likely that satellite cells would need to navigate local muscle tissue to reach damaged areas, relatively little data on such motility exist, and most studies have been with immortalized cell lines. We find that primary satellite cells are significantly more motile than myoblast cell lines, and that adhesion to laminin promotes primary cell motility more than fourfold over other substrates. Using timelapse videomicroscopy to assess satellite cell motility on single living myofibers, we have identified a requirement for the laminin-binding integrin alpha 7 beta 1 in satellite cell motility, as well as a role for hepatocyte growth factor in promoting directional persistence. The extensive migratory behavior of satellite cells resident on muscle fibers suggests caution when determining, based on fixed specimens, whether adjacent cells are daughters from the same mother cell. We also observed more persistent long-term contact between individual satellite cells than has been previously supposed, potential cell-cell attractive and repulsive interactions, and migration between host myofibers. Based on such activity, we assayed for expression of "pathfinding" cues, and found that satellite cells express multiple guidance ligands and receptors. Together, these data suggest that satellite cell migration in vivo may be more extensive than currently thought, and could be regulated by combinations of signals, including adhesive haptotaxis, soluble factors, and guidance cues.

  10. Cell biology (Communication arising): Tubulin acetylation and cell motility

    NASA Astrophysics Data System (ADS)

    Palazzo, Alexander; Ackerman, Brian; Gundersen, Gregg G.

    2003-01-01

    Although the protein tubulin is known to undergo several post-translational modifications that accumulate in stable but not dynamic microtubules inside cells, the function of these modifications is unknown. Hubbert et al. have shown that the enzyme HDAC6 (for histone deacetylase 6) reverses the post-translational acetylation of tubulin, and provide evidence that reducing tubulin acetylation enhances cell motility. They also suggest that decreasing tubulin acetylation reduces microtubule stability. However, we find that microtubule stabilization is not promoted by tubulin acetylation. We conclude that the alteration in cell motility observed by Hubbert et al. in cells overexpressing HDAC6 results not from changes in the formation of stable microtubules, but from alterations in the degree of tubulin acetylation.

  11. Mannose-Binding Lectin Inhibits the Motility of Pathogenic Salmonella by Affecting the Driving Forces of Motility and the Chemotactic Response

    PubMed Central

    Nakamura, Shuichi; Islam, Md. Shafiqul; Guo, Yijie; Ihara, Kohei; Tomioka, Rintaro; Masuda, Mizuki; Yoneyama, Hiroshi; Isogai, Emiko

    2016-01-01

    Mannose-binding lectin (MBL) is a key pattern recognition molecule in the lectin pathway of the complement system, an important component of innate immunity. MBL functions as an opsonin which enhances the sequential immune process such as phagocytosis. We here report an inhibitory effect of MBL on the motility of pathogenic bacteria, which occurs by affecting the energy source required for motility and the signaling pathway of chemotaxis. When Salmonella cells were treated with a physiological concentration of MBL, their motile fraction and free-swimming speed decreased. Rotation assays of a single flagellum showed that the flagellar rotation rate was significantly reduced by the addition of MBL. Measurements of the intracellular pH and membrane potential revealed that MBL affected a driving force for the Salmonella flagellum, the electrochemical potential difference of protons. We also found that MBL treatment increased the reversal frequency of Salmonella flagellar rotation, which interfered with the relative positive chemotaxis toward an attractive substrate. We thus propose that the motility inhibition effect of MBL may be secondarily involved in the attack against pathogens, potentially facilitating the primary role of MBL in the complement system. PMID:27104738

  12. Time course of "escape" from calcitonin-induced inhibition of motility and resorption of disaggregated osteoclasts.

    PubMed

    Kanehisa, J

    1989-01-01

    The reversible calcitonin (CT)-induced inhibition of osteoclastic activity has been studied to clarify the mechanisms responsible for the so-called "escape phenomenon." Osteoclasts disaggregated from neonatal rabbits were cultured on glass coverslips or thin bovine bone slices. Resorption activity was evaluated by using time-lapse recording and scanning electron microscopy. Addition of CT to the cultures caused most osteoclasts on glass surfaces to be immotile and contracted. From 1.5 h onward, in cultures with CT, osteoclasts started to escape from CT-induced quiescence independently of other cells. CT also prevented osteoclasts on bone slices from excavating bone while concomitant cell immobility occurred. Inhibited osteoclasts were able to regain apparent bone-resorbing potency only after resumption of cytoplasmic immobility. The resumption of bone resorption could begin as early as 9.7 h after CT addition. The observations indicate that CT-induced inhibition of osteoclastic bone resorption is associated with inhibition of cytoplasmic motility and that the "escape" phenomenon reflects resumption of activity of osteoclasts that were previously inhibited by CT action rather than the resportive activity of newly formed osteoclasts. PMID:2765310

  13. SHP-2 phosphatase activity is required for PECAM-1-dependent cell motility.

    PubMed

    Zhu, Jing-Xu; Cao, Gaoyuan; Williams, James T; Delisser, Horace M

    2010-10-01

    Platelet endothelial cell adhesion molecule-1 (PECAM-1) has been implicated in endothelial cell motility during angiogenesis. Although there is evidence that SHP-2 plays a role in PECAM-1-dependent cell motility, the molecular basis of the activity of SHP-2 in this process has not been defined. To investigate the requirement of SHP-2 in PECAM-1-dependent cell motility, studies were done in which various constructs of SHP-2 were expressed in cell transfectants expressing PECAM-1. We observed that the levels of PECAM-1 tyrosine phosphorylation and SHP-2 association with PECAM-1 were significantly increased in cells expressing a phosphatase-inactive SHP-2 mutant, suggesting that the level of PECAM-1 tyrosine phosphorylation, and thus SHP-2 binding are regulated in part by bound, catalytically active SHP-2. We subsequently found that expression of PECAM-1 stimulated wound-induced migration and the formation of filopodia (a morphological feature of motile cells). These activities were associated with increased mitogen-activated protein kinase (MAPK) activation and the dephosphorylation of paxillin (an event implicated in the activation of MAPK). The phosphatase-inactive SHP-2 mutant, however, suppressed these PECAM-1-dependent phenomena, whereas the activity of PECAM-1 expressing cells was not altered by expression of wild-type SHP-2 or SHP-2 in which the scaffold/adaptor function had been disabled. Pharmacological inhibition of SHP-2 phosphatase activity also suppressed PECAM-1-dependent motility. Furthermore, PECAM-1 expression also stimulates tube formation, but none of the SHP-2 constructs affected this process. These findings therefore suggest a model for the involvement of SHP-2 in PECAM-1-dependent motility in which SHP-2, recruited by its interaction with PECAM-1, targets paxillin to ultimately activate the MAPK pathway and downstream events required for cell motility. PMID:20631249

  14. T Cell Motility as Modulator of Interactions with Dendritic Cells

    PubMed Central

    Stein, Jens V.

    2015-01-01

    It is well established that the balance of costimulatory and inhibitory signals during interactions with dendritic cells (DCs) determines T cell transition from a naïve to an activated or tolerant/anergic status. Although many of these molecular interactions are well reproduced in reductionist in vitro assays, the highly dynamic motility of naïve T cells in lymphoid tissue acts as an additional lever to fine-tune their activation threshold. T cell detachment from DCs providing suboptimal stimulation allows them to search for DCs with higher levels of stimulatory signals, while storing a transient memory of short encounters. In turn, adhesion of weakly reactive T cells to DCs presenting peptides presented on major histocompatibility complex with low affinity is prevented by lipid mediators. Finally, controlled recruitment of CD8+ T cells to cognate DC–CD4+ T cell clusters shapes memory T cell formation and the quality of the immune response. Dynamic physiological lymphocyte motility therefore constitutes a mechanism to mitigate low avidity T cell activation and to improve the search for “optimal” DCs, while contributing to peripheral tolerance induction in the absence of inflammation. PMID:26579132

  15. Effects of Hedera helix L. extracts on rat prostate cancer cell proliferation and motility

    PubMed Central

    Gumushan-Aktas, Hatice; Altun, Seyhan

    2016-01-01

    Hedera helix L., a member of Araliaceae family, has antiproliferative, cytotoxic, antimicrobial, antifungal, antiprotozoal and anti-inflammatory effects, and is used in cosmetics. The aim of the present study was to investigate the effect of treatment with extracts of leaves and unripened fruits of H. helix on rat prostate cancer cell lines with markedly different metastatic potentials: Mat-LyLu cells (strongly metastatic) and AT-2 cells (weakly metastatic). The effects of the extracts on cell kinetics and migration were determined. Tetrodotoxin was used to block the voltage-gated sodium channels (VGSCs) associated specifically with Mat-LyLu cells. Cell proliferation was detected spectrophotometrically using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay. The mitotic index was determined using the Feulgen staining method. Lateral motility was quantified by wound-healing assays. The results of the present study demonstrated that cell kinetics (proliferation and mitotic activity) and motility were inhibited by ethanolic leaf extract of H. helix. The ethanolic extract of H. helix fruit suppressed Mat-LyLu cell migration, with no effect on proliferation. The opposite effects were observed in AT-2 cells; migration was not affected but proliferation was inhibited. In conclusion, the ethanolic fruit extract of H. helix may inhibit the cell migration of Mat-LyLu cells by blocking VGSCs. However, the effect of ethanolic leaf extract of H. helix treatment on the lateral motility of the cancer cells is unclear. PMID:27698887

  16. Effects of Hedera helix L. extracts on rat prostate cancer cell proliferation and motility

    PubMed Central

    Gumushan-Aktas, Hatice; Altun, Seyhan

    2016-01-01

    Hedera helix L., a member of Araliaceae family, has antiproliferative, cytotoxic, antimicrobial, antifungal, antiprotozoal and anti-inflammatory effects, and is used in cosmetics. The aim of the present study was to investigate the effect of treatment with extracts of leaves and unripened fruits of H. helix on rat prostate cancer cell lines with markedly different metastatic potentials: Mat-LyLu cells (strongly metastatic) and AT-2 cells (weakly metastatic). The effects of the extracts on cell kinetics and migration were determined. Tetrodotoxin was used to block the voltage-gated sodium channels (VGSCs) associated specifically with Mat-LyLu cells. Cell proliferation was detected spectrophotometrically using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay. The mitotic index was determined using the Feulgen staining method. Lateral motility was quantified by wound-healing assays. The results of the present study demonstrated that cell kinetics (proliferation and mitotic activity) and motility were inhibited by ethanolic leaf extract of H. helix. The ethanolic extract of H. helix fruit suppressed Mat-LyLu cell migration, with no effect on proliferation. The opposite effects were observed in AT-2 cells; migration was not affected but proliferation was inhibited. In conclusion, the ethanolic fruit extract of H. helix may inhibit the cell migration of Mat-LyLu cells by blocking VGSCs. However, the effect of ethanolic leaf extract of H. helix treatment on the lateral motility of the cancer cells is unclear.

  17. Single cell motility and trail formation in populations of microglia

    NASA Astrophysics Data System (ADS)

    Lee, Kyoung Jin

    2009-03-01

    Microglia are a special type of glia cell in brain that has immune responses. They constitute about 20 % of the total glia population within the brain. Compared to other glia cells, microglia are very motile, constantly moving to destroy pathogens and to remove dead neurons. While doing so, they exhibit interesting body shapes, have cell-to-cell communications, and have chemotatic responses to each other. Interestingly, our recent in vitro studies show that their unusual motile behaviors can self-organize to form trails, similar to those in populations of ants. We have studied the changes in the physical properties of these trails by varying the cell population density and by changing the degree of spatial inhomogeneities (``pathogens''). Our experimental observations can be quite faithfully reproduced by a simple mathematical model involving many motile cells whose mechanical motion are driven by actin polymerization and depolymerization process within the individual cell body and by external chemical gradients.

  18. 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

    2013-11-01

    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.

  19. L1 cell adhesion molecule induces melanoma cell motility by activation of mitogen-activated protein kinase pathways.

    PubMed

    Yi, Young-Su; Baek, Kwang-Soo; Cho, Jae Youl

    2014-06-01

    L1 cell adhesion molecule (L1CAM) is highly expressed in various types of cancer cells and has been implicated in the control of cell proliferation and motility. Recently, L1CAM was reported to induce the motility of melanoma cells, but the mechanism of this induction remains poorly understood. In this study, we investigated the molecular mechanisms by which L1CAM induces the motility of melanoma cells. Unlike other types of cancer cells, B16F10 melanoma cells highly expressed L1CAM at both the RNA and protein levels, and the expression of L1CAM induced AP-1 activity. In accordance to AP-1 activation, MAPK signaling pathways were activated by L1CAM. Inhibition of L1CAM expression by L1CAM-specific siRNA suppressed the activation of MAPKs such as ERK and p38. However, no significant change was observed in JNK activation. As expected, upstream MAP2K, MKK3/6, MAP3K, and TAK1 were also deactivated by the inhibition of L1CAM expression. L1CAM induced the motility of B16F10 cells. Inhibition of L1CAM expression suppressed migration and invasion of B16F10 cells, but no suppressive effect was observed on their proliferation and anti-apoptotic resistance. Treatment of B16F10 cells with U0126, an ERK inhibitor, or SB203580, a p38 inhibitor, suppressed the migration and invasion abilities of B16F10 cells. Taken together, our results suggest that L1CAM induces the motility of B16F10 melanoma cells via the activation of MAPK pathways. This finding provides a more detailed molecular mechanism of L1CAM-mediated induction of melanoma cell motility. PMID:24974583

  20. 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

  1. Cell motility and antibiotic tolerance of bacterial swarms

    NASA Astrophysics Data System (ADS)

    Zuo, Wenlong

    Many bacteria species can move across moist surfaces in a coordinated manner known as swarming. It is reported that swarm cells show higher tolerance to a wide variety of antibiotics than planktonic cells. We used the model bacterium E. coli to study how motility affects the antibiotic tolerance of swarm cells. Our results provide new insights for the control of pathogenic invasion via regulating cell motility. Mailing address: Room 306 Science Centre North Block, The Chinese University of Hong Kong, Shatin, N.T. Hong Kong SAR. Phone: +852-3943-6354. Fax: +852-2603-5204. E-mail: zwlong@live.com.

  2. Form and Function in Cell Motility: From Fibroblasts to Keratocytes

    PubMed Central

    Herant, Marc; Dembo, Micah

    2010-01-01

    Abstract It is plain enough that a horse is made for running, but similar statements about motile cells are not so obvious. Here the basis for structure-function relations in cell motility is explored by application of a new computational technique that allows realistic three-dimensional simulations of cells migrating on flat substrata. With this approach, some cyber cells spontaneously display the classic irregular protrusion cycles and handmirror morphology of a crawling fibroblast, and others the steady gliding motility and crescent morphology of a fish keratocyte. The keratocyte motif is caused by optimal recycling of the cytoskeleton from the back to the front so that more of the periphery can be devoted to protrusion. These calculations are a step toward bridging the gap between the integrated mechanics and biophysics of whole cells and the microscopic molecular biology of cytoskeletal components. PMID:20409459

  3. Actin–myosin network reorganization breaks symmetry at the cell rear to spontaneously initiate polarized cell motility

    PubMed Central

    Yam, Patricia T.; Wilson, Cyrus A.; Ji, Lin; Hebert, Benedict; Barnhart, Erin L.; Dye, Natalie A.; Wiseman, Paul W.; Danuser, Gaudenz; Theriot, Julie A.

    2007-01-01

    We have analyzed the spontaneous symmetry breaking and initiation of actin-based motility in keratocytes (fish epithelial cells). In stationary keratocytes, the actin network flow was inwards and radially symmetric. Immediately before motility initiation, the actin network flow increased at the prospective cell rear and reoriented in the perinuclear region, aligning with the prospective axis of movement. Changes in actin network flow at the cell front were detectable only after cell polarization. Inhibition of myosin II or Rho kinase disrupted actin network organization and flow in the perinuclear region and decreased the motility initiation frequency, whereas increasing myosin II activity with calyculin A increased the motility initiation frequency. Local stimulation of myosin activity in stationary cells by the local application of calyculin A induced directed motility initiation away from the site of stimulation. Together, these results indicate that large-scale actin–myosin network reorganization and contractility at the cell rear initiate spontaneous symmetry breaking and polarized motility of keratocytes. PMID:17893245

  4. Lipopolysaccharide of Marinobacter litoralis inhibits swarming motility and biofilm formation in Pseudomonas aeruginosa PA01.

    PubMed

    Sardar, Raj Kumar; Kavita, Kumari; Jha, Bhavanath

    2015-06-01

    The lipopolysaccharide (LPS) was isolated from a marine bacterium identified as Marinobacter litoralis BK09 using 16S rRNA gene sequence similarity analysis. The GCMS analysis showed that the LPS contained 3-hydroxy-dodecanoic acid (C12:0 3OH) (49%), dodecanoic acid (C12:0) (24%) and decanoic acid (C10:0) (19%) as major fatty acids, and the polysaccharide constituents were fucose (53.79%), xylose (28.04%) and mannose (18.15%). The LPS almost completely inhibited swarming motility in Pseudomonas aeruginosa PA01. It also reduced biofilm formation by 50% with no adverse effect on cell growth. The production of virulence factor such as pyocyanin pigment was reduced (∼40%) by the LPS. The LPS did not show any limulus amoebocyte lysate (LAL) gelation activity. The repression of swarming motility, pyocyanin production and biofilm formation by the LPS suggests its potential application against P. aeruginosa infection. This is the first report on characterization and application of LPS from M. litoralis. PMID:25843881

  5. Helical motion of the cell body enhances Caulobacter crescentus motility.

    PubMed

    Liu, Bin; Gulino, Marco; Morse, Michael; Tang, Jay X; Powers, Thomas R; Breuer, Kenneth S

    2014-08-01

    We resolve the 3D trajectory and the orientation of individual cells for extended times, using a digital tracking technique combined with 3D reconstructions. We have used this technique to study the motility of the uniflagellated bacterium Caulobacter crescentus and have found that each cell displays two distinct modes of motility, depending on the sense of rotation of the flagellar motor. In the forward mode, when the flagellum pushes the cell, the cell body is tilted with respect to the direction of motion, and it precesses, tracing out a helical trajectory. In the reverse mode, when the flagellum pulls the cell, the precession is smaller and the cell has a lower translation distance per rotation period and thus a lower motility. Using resistive force theory, we show how the helical motion of the cell body generates thrust and can explain the direction-dependent changes in swimming motility. The source of the cell body precession is believed to be associated with the flexibility of the hook that connects the flagellum to the cell body.

  6. Cooperative cell motility during tandem locomotion of amoeboid cells

    PubMed Central

    Bastounis, Effie; Álvarez-González, Begoña; del Álamo, Juan C.; Lasheras, Juan C.; Firtel, Richard A.

    2016-01-01

    Streams of migratory cells are initiated by the formation of tandem pairs of cells connected head to tail to which other cells subsequently adhere. The mechanisms regulating the transition from single to streaming cell migration remain elusive, although several molecules have been suggested to be involved. In this work, we investigate the mechanics of the locomotion of Dictyostelium tandem pairs by analyzing the spatiotemporal evolution of their traction adhesions (TAs). We find that in migrating wild-type tandem pairs, each cell exerts traction forces on stationary sites (∼80% of the time), and the trailing cell reuses the location of the TAs of the leading cell. Both leading and trailing cells form contractile dipoles and synchronize the formation of new frontal TAs with ∼54-s time delay. Cells not expressing the lectin discoidin I or moving on discoidin I–coated substrata form fewer tandems, but the trailing cell still reuses the locations of the TAs of the leading cell, suggesting that discoidin I is not responsible for a possible chemically driven synchronization process. The migration dynamics of the tandems indicate that their TAs’ reuse results from the mechanical synchronization of the leading and trailing cells’ protrusions and retractions (motility cycles) aided by the cell–cell adhesions. PMID:26912787

  7. Inhibition of Mitochondrial Complex I Leads to Decreased Motility and Membrane Integrity Related to Increased Hydrogen Peroxide and Reduced ATP Production, while the Inhibition of Glycolysis Has Less Impact on Sperm Motility

    PubMed Central

    Plaza Davila, María; Martin Muñoz, Patricia; Tapia, Jose A.; Ortega Ferrusola, Cristina; Balao da Silva C, Carolina; Peña, Fernando J.

    2015-01-01

    Mitochondria have been proposed as the major source of reactive oxygen species in somatic cells and human spermatozoa. However, no data regarding the role of mitochondrial ROS production in stallion spermatozoa are available. To shed light on the role of the mitochondrial electron transport chain in the origin of oxidative stress in stallion spermatozoa, specific inhibitors of complex I (rotenone) and III (antimycin-A) were used. Ejaculates from seven Andalusian stallions were collected and incubated in BWW media at 37°C in the presence of rotenone, antimycin-A or control vehicle. Incubation in the presence of these inhibitors reduced sperm motility and velocity (CASA analysis) (p<0.01), but the effect was more evident in the presence of rotenone (a complex I inhibitor). These inhibitors also decreased ATP content. The inhibition of complexes I and III decreased the production of reactive oxygen species (p<0.01) as assessed by flow cytometry after staining with CellRox deep red. This observation suggests that the CellRox probe mainly identifies superoxide and that superoxide production may reflect intense mitochondrial activity rather than oxidative stress. The inhibition of complex I resulted in increased hydrogen peroxide production (p<0.01). The inhibition of glycolysis resulted in reduced sperm velocities (p<0.01) without an effect on the percentage of total motile sperm. Weak and moderate (but statistically significant) positive correlations were observed between sperm motility, velocity and membrane integrity and the production of reactive oxygen species. These results indicate that stallion sperm rely heavily on oxidative phosphorylation (OXPHOS) for the production of ATP for motility but also require glycolysis to maintain high velocities. These data also indicate that increased hydrogen peroxide originating in the mitochondria is a mechanism involved in stallion sperm senescence. PMID:26407142

  8. Inhibition of Mitochondrial Complex I Leads to Decreased Motility and Membrane Integrity Related to Increased Hydrogen Peroxide and Reduced ATP Production, while the Inhibition of Glycolysis Has Less Impact on Sperm Motility.

    PubMed

    Plaza Davila, María; Martin Muñoz, Patricia; Tapia, Jose A; Ortega Ferrusola, Cristina; Balao da Silva C, Carolina; Peña, Fernando J

    2015-01-01

    Mitochondria have been proposed as the major source of reactive oxygen species in somatic cells and human spermatozoa. However, no data regarding the role of mitochondrial ROS production in stallion spermatozoa are available. To shed light on the role of the mitochondrial electron transport chain in the origin of oxidative stress in stallion spermatozoa, specific inhibitors of complex I (rotenone) and III (antimycin-A) were used. Ejaculates from seven Andalusian stallions were collected and incubated in BWW media at 37 °C in the presence of rotenone, antimycin-A or control vehicle. Incubation in the presence of these inhibitors reduced sperm motility and velocity (CASA analysis) (p<0.01), but the effect was more evident in the presence of rotenone (a complex I inhibitor). These inhibitors also decreased ATP content. The inhibition of complexes I and III decreased the production of reactive oxygen species (p<0.01) as assessed by flow cytometry after staining with CellRox deep red. This observation suggests that the CellRox probe mainly identifies superoxide and that superoxide production may reflect intense mitochondrial activity rather than oxidative stress. The inhibition of complex I resulted in increased hydrogen peroxide production (p<0.01). The inhibition of glycolysis resulted in reduced sperm velocities (p<0.01) without an effect on the percentage of total motile sperm. Weak and moderate (but statistically significant) positive correlations were observed between sperm motility, velocity and membrane integrity and the production of reactive oxygen species. These results indicate that stallion sperm rely heavily on oxidative phosphorylation (OXPHOS) for the production of ATP for motility but also require glycolysis to maintain high velocities. These data also indicate that increased hydrogen peroxide originating in the mitochondria is a mechanism involved in stallion sperm senescence.

  9. Effect of gingerol on colonic motility via inhibition of calcium channel currents in rats

    PubMed Central

    Cai, Zheng-Xu; Tang, Xu-Dong; Wang, Feng-Yun; Duan, Zhi-Jun; Li, Yu-Chun; Qiu, Juan-Juan; Guo, Hui-Shu

    2015-01-01

    AIM: To investigate the effect of gingerol on colonic motility and the action of L-type calcium channel currents in this process. METHODS: The distal colon was cut along the mesenteric border and cleaned with Ca2+-free physiological saline solution. Muscle strips were removed and placed in Ca2+-free physiological saline solution, which was oxygenated continuously. Longitudinal smooth muscle samples were prepared by cutting along the muscle strips and were then placed in a chamber. Mechanical contractile activities of isolated colonic segments in rats were recorded by a 4-channel physiograph. Colon smooth muscle cells were dissociated by enzymatic digestion. L-type calcium currents were recorded using the conventional whole-cell patch-clamp technique. RESULTS: Gingerol inhibited the spontaneous contraction of colonic longitudinal smooth muscle in a dose-dependent manner with inhibition percentages of 13.3% ± 4.1%, 43.4% ± 3.9%, 78.2% ± 3.6% and 80.5% ± 4.5% at 25 μmol/L, 50 μmol/L, 75 μmol/L and 100 μmol/L, respectively (P < 0.01). Nifedipine, an L-type calcium channel blocker, diminished the inhibition of colonic motility by gingerol. Gingerol inhibited L-type calcium channel currents in colonic longitudinal myocytes of rats. At a 75 μmol/L concentration of gingerol, the percentage of gingerol-induced inhibition was diminished by nifedipine from 77.1% ± 4.2% to 42.6% ± 3.6% (P < 0.01). Gingerol suppressed IBa in a dose-dependent manner, and the inhibition rates were 22.7% ± 2.38%, 35.77% ± 3.14%, 49.78% ± 3.48% and 53.78% ± 4.16% of control at 0 mV, respectively, at concentrations of 25 μmol/L, 50 μmol/L, 75 μmol/L and 100 μmol/L (P < 0.01). The steady-state activation curve was shifted to the right by treatment with gingerol. The value of half activation was -14.23 ± 1.12 mV in the control group and -10.56 ± 1.04 mV in the 75 μmol/L group (P < 0.05) with slope factors, Ks, of 7.16 ± 0.84 and 7.02 ± 0.93 (P < 0.05) in the control and 75

  10. 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.

  11. Inhibitory Activity of (+)-Usnic Acid against Non-Small Cell Lung Cancer Cell Motility

    PubMed Central

    Yang, Yi; Nguyen, Thanh Thi; Jeong, Min-Hye; Crişan, Florin; Yu, Young Hyun; Ha, Hyung-Ho; Choi, Kyung Hee; Jeong, Hye Gwang; Jeong, Tae Cheon; Lee, Kwang Youl; Kim, Kyung Keun; Hur, Jae-Seoun; Kim, Hangun

    2016-01-01

    Lichens are symbiotic organisms that produce various unique chemicals that can be used for pharmaceutical purposes. With the aim of screening new anti-cancer agents that inhibit cancer cell motility, we tested the inhibitory activity of seven lichen species collected from the Romanian Carpathian Mountains against migration and invasion of human lung cancer cells and further investigated the molecular mechanisms underlying their anti-metastatic activity. Among them, Alectoria samentosa, Flavocetraria nivalis, Alectoria ochroleuca, and Usnea florida showed significant inhibitory activity against motility of human lung cancer cells. HPLC results showed that usnic acid is the main compound in these lichens, and (+)-usnic acid showed similar inhibitory activity that crude extract have. Mechanistically, β-catenin-mediated TOPFLASH activity and KITENIN-mediated AP-1 activity were decreased by (+)-usnic acid treatment in a dose-dependent manner. The quantitative real-time PCR data showed that (+)-usnic acid decreased the mRNA level of CD44, Cyclin D1 and c-myc, which are the downstream target genes of both β-catenin/LEF and c-jun/AP-1. Also, Rac1 and RhoA activities were decreased by treatment with (+)-usnic acid. Interestingly, higher inhibitory activity for cell invasion was observed when cells were treated with (+)-usnic acid and cetuximab. These results implied that (+)-usnic acid might have potential activity in inhibition of cancer cell metastasis, and (+)-usnic acid could be used for anti-cancer therapy with a distinct mechanisms of action. PMID:26751081

  12. Rapid actin-dependent viral motility in live cells.

    PubMed

    Vaughan, Joshua C; Brandenburg, Boerries; Hogle, James M; Zhuang, Xiaowei

    2009-09-16

    During the course of an infection, viruses take advantage of a variety of mechanisms to travel in cells, ranging from diffusion within the cytosol to active transport along cytoskeletal filaments. To study viral motility within the intrinsically heterogeneous environment of the cell, we have developed a motility assay that allows for the global and unbiased analysis of tens of thousands of virus trajectories in live cells. Using this assay, we discovered that poliovirus exhibits anomalously rapid intracellular movement that was independent of microtubules, a common track for fast and directed cargo transport. Such rapid motion, with speeds of up to 5 microm/s, allows the virus particles to quickly explore all regions of the cell with the exception of the nucleus. The rapid, microtubule-independent movement of poliovirus was observed in multiple human-derived cell lines, but appeared to be cargo-specific. Other cargo, including a closely related picornavirus, did not exhibit similar motility. Furthermore, the motility is energy-dependent and requires an intact actin cytoskeleton, suggesting an active transport mechanism. The speed of this microtubule-independent but actin-dependent movement is nearly an order of magnitude faster than the fastest speeds reported for actin-dependent transport in animal cells, either by actin polymerization or by myosin motor proteins.

  13. HES6 enhances the motility of alveolar rhabdomyosarcoma cells

    SciTech Connect

    Wickramasinghe, Caroline M; Domaschenz, Renae; Amagase, Yoko; Williamson, Daniel; Missiaglia, Edoardo; Shipley, Janet; Murai, Kasumi; Jones, Philip H

    2013-01-01

    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). Knockdown 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.

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

    SciTech Connect

    Gearhart, Debra A. . E-mail: dgearhar@mcg.edu; Sickles, Dale W.; Buccafusco, Jerry J.; Prendergast, Mark A.; Terry, Alvin V.

    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), and 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.

  15. Differential effects on cell motility, embryonic stem cell self-renewal and senescence by diverse Src kinase family inhibitors

    SciTech Connect

    Tamm, Christoffer Galito, Sara Pijuan Anneren, Cecilia

    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 in 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.

  16. Tyrosine kinase-mediated axial motility of basal cells revealed by intravital imaging

    PubMed Central

    Roy, Jeremy; Kim, Bongki; Hill, Eric; Visconti, Pablo; Krapf, Dario; Vinegoni, Claudio; Weissleder, Ralph; Brown, Dennis; Breton, Sylvie

    2016-01-01

    Epithelial cells are generally considered to be static relative to their neighbours. Basal cells in pseudostratified epithelia display a single long cytoplasmic process that can cross the tight junction barrier to reach the lumen. Using in vivo microscopy to visualize the epididymis, a model system for the study of pseudostratified epithelia, we report here the surprising discovery that these basal cell projections—which we call axiopodia—periodically extend and retract over time. We found that axiopodia extensions and retractions follow an oscillatory pattern. This movement, which we refer to as periodic axial motility (PAM), is controlled by c-Src and MEK1/2–ERK1/2. Therapeutic inhibition of tyrosine kinase activity induces a retraction of these projections. Such unexpected cell motility may reflect a novel mechanism by which specialized epithelial cells sample the luminal environment. PMID:26868824

  17. PKCα-Mediated Signals Regulate the Motile Responses of Cochlear Outer Hair Cells.

    PubMed

    Park, Channy; Kalinec, Federico

    2015-05-01

    There is strong evidence that changes in the actin/spectrin-based cortical cytoskeleton of outer hair cells (OHCs) regulate their motile responses as well as cochlear amplification, the process that optimizes the sensitivity and frequency selectivity of the mammalian inner ear. Since a RhoA/protein kinase C (PKC)-mediated pathway is known to inhibit the actin-spectrin interaction in other cell models, we decided to investigate whether this signaling cascade could also participate in the regulation of OHC motility. We used high-speed video microscopy and confocal microscopy to explore the effects of pharmacological activation of PKCα, PKCβI, PKCβII, PKCδ, PKCε, and PKCζ with lysophosphatidic acid (LPA) and their inhibition with bisindolylmaleimide I, as well as inhibition of RhoA and Rho-associated protein kinase (ROCK) with C3 and Y-27632, respectively. Motile responses were induced in isolated guinea pig OHCs by stimulation with an 8 V/cm external alternating electrical field as 50 Hz bursts of square wave pulses (100 ms on/off). We found that LPA increased expression of PKCα and PKCζ only, with PKCα, but not PKCζ, phosphorylating the cytoskeletal protein adducin of both Ser-726 and Thr-445. Interestingly, however, inhibition of PKCα reduced adducin phosphorylation only at Ser-726. We also determined that LPA activation of a PKCα-mediated signaling pathway simultaneously enhanced OHC electromotile amplitude and cell shortening, and facilitated RhoA/ROCK/LIMK1-mediated cofilin phosphorylation. Altogether, our results suggest that PKCα-mediated signals, probably via adducin-mediated inhibition of actin-spectrin binding and cofilin-mediated depolymerization of actin filaments, play an essential role in the homeostatic regulation of OHC motility and cochlear amplification. PMID:25954875

  18. 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.

  19. Zebrafish germ cells: motility and guided migration.

    PubMed

    Paksa, Azadeh; Raz, Erez

    2015-10-01

    In the course of embryonic development, the process of cell migration is critical for establishment of the embryonic body plan, for morphogenesis and for organ function. Investigating the molecular mechanisms underlying cell migration is thus crucial for understanding developmental processes and clinical conditions resulting from abnormal cell migration such as cancer metastasis. The long-range migration of primordial germ cells toward the region at which the gonad develops occurs in embryos of various species and thus constitutes a useful in vivo model for single-cell migration. Recent studies employing zebrafish embryos have greatly contributed to the understanding of the mechanisms facilitating the migration of these cells en route to their target.

  20. PAK4 kinase activity and somatic mutation promote carcinoma cell motility and influence inhibitor sensitivity

    PubMed Central

    Whale, Andrew D.; Dart, Anna; Holt, Mark; Jones, Gareth E.; Wells, Claire M.

    2012-01-01

    Hepatocyte growth factor (HGF) and its receptor (c-Met) are associated with cancer cell motility and invasiveness. p21-activated kinase 4 (PAK4), a potential therapeutic target, is recruited to and activated by c-Met. In response, PAK4 phosphorylates LIM kinase 1 (LIMK1) in an HGF-dependent manner in metastatic prostate carcinoma cells. PAK4 overexpression is known to induce increased cell migration speed but the requirement for kinase activity has not been established. We have used a panel of PAK4 truncations and mutations in a combination of over-expression and RNAi rescue experiments to determine the requirement for PAK4 kinase activity during carcinoma cell motility downstream of HGF. We find that neither the kinase domain alone nor a PAK4 mutant unable to bind Cdc42 is able to fully rescue cell motility in a PAK4-deficient background. Nevertheless, we find that PAK4 kinase activity and associated LIMK1 activity are essential for carcinoma cell motility, highlighting PAK4 as a potential anti-metastatic therapeutic target. We also show here that overexpression of PAK4 harboring a somatic mutation, E329K, increased the HGF-driven motility of metastatic prostate carcinoma cells. E329 lies within the G-loop region of the kinase. Our data suggest E329K mutation leads to a modest increase in kinase activity conferring resistance to competitive ATP inhibitors in addition to promoting cell migration. The existence of such a mutation may have implications for the development of PAK4-specific competitive ATP inhibitors should PAK4 be further explored for clinical inhibition. PMID:22689056

  1. Cell Motility Resulting form Spontaneous Polymerization Waves

    NASA Astrophysics Data System (ADS)

    Kruse, Karsten

    2014-03-01

    The crawling of living cells on solid substrates is often driven by the actin cytoskeleton, a network of structurally polar filamentous proteins that is intrinsically driven by the hydrolysis of ATP. How cells organize their actin network during crawling is still poorly understood. A possible general mechanism underlying actin organization has been offered by the observation of spontaneous actin polymerization waves in various different cell types. We use a theoretical approach to investigate the possible role of spontaneous actin waves on cell crawling. To this end, we develop a meanfield framework for studying spatiotemporal aspects of actin assembly dynamics, which helped to identify possible origins of self-organized actin waves. The impact of these waves on cell crawling is then investigated by using a phase-field approach to confine the actin network to a cellular domain. We find that spontaneous actin waves can lead to directional or amoeboidal crawling. In the latter case, the cell performs a random walk. Within our deterministic framework, this behavior is due to complex spiral waves inside the cell. Finally, we compare the seemingly random motion of our model cells to the dynamics of cells of the human immune system. These cells patrol the body in search for infected cells and we discuss possible implications of our theory for the search process' efficiency. Work was funded by the DFG through KR3430/1, GK1276, and SFB 1027.

  2. Tyrosine kinase inhibitor STI571 enhances thyroid cancer cell motile response to Hepatocyte Growth Factor.

    PubMed

    Frasca, F; Vigneri, P; Vella, V; Vigneri, R; Wang, J Y

    2001-06-28

    The Hepatocyte Growth Factor (HGF) and its receptor Met are physiological regulators of cell migration. HGF and Met have also been implicated in tumor progression and metastasis. We show here that the tyrosine kinase inhibitor STI571 has a stimulatory effect on HGF-induced migration and branching morphogenesis in thyroid cancer but not in primary or immortalized thyroid epithelial cells. These stimulatory effects of STI571 are observed at a concentration that is clinically relevant. The STI571-enhanced motile response can be correlated with an increase in the Met receptor tyrosine phosphorylation as well as ERK and Akt activation by HGF. Interestingly, one of the targets of STI571, namely the c-Abl tyrosine kinase, is activated by HGF and is recruited at the migrating edge of thyroid cancer cells. These data suggests that c-Abl and/or STI571-inhibited tyrosine kinases can negatively regulate the Met receptor to restrain the motile response in thyroid cancer cells.

  3. Wild-type p53 controls cell motility and invasion by dual regulation of MET expression

    PubMed Central

    Hwang, Chang-Il; Matoso, Andres; Corney, David C.; Flesken-Nikitin, Andrea; Körner, Stefanie; Wang, Wei; Boccaccio, Carla; Thorgeirsson, Snorri S.; Comoglio, Paolo M.; Hermeking, Heiko; Nikitin, Alexander Yu.

    2011-01-01

    Recent observations suggest that p53 mutations are responsible not only for growth of primary tumors but also for their dissemination. However, mechanisms involved in p53-mediated control of cell motility and invasion remain poorly understood. By using the primary ovarian surface epithelium cell culture, we show that conditional inactivation of p53 or expression of its mutant forms results in overexpression of MET receptor tyrosine kinase, a crucial regulator of invasive growth. At the same time, cells acquire increased MET-dependent motility and invasion. Wild-type p53 negatively regulates MET expression by two mechanisms: (i) transactivation of MET-targeting miR-34, and (ii) inhibition of SP1 binding to MET promoter. Both mechanisms are not functional in p53 absence, but mutant p53 proteins retain partial MET promoter suppression. Accordingly, MET overexpression, cell motility, and invasion are particularly high in p53-null cells. These results identify MET as a critical effector of p53 and suggest that inhibition of MET may be an effective antimetastatic approach to treat cancers with p53 mutations. These results also show that the extent of advanced cancer traits, such as invasion, may be determined by alterations in individual components of p53/MET regulatory network. PMID:21831840

  4. Extending the molecular clutch beyond actin-based cell motility

    NASA Astrophysics Data System (ADS)

    Havrylenko, Svitlana; Mezanges, Xavier; Batchelder, Ellen; Plastino, Julie

    2014-10-01

    Many cell movements occur via polymerization of the actin cytoskeleton beneath the plasma membrane at the front of the cell, forming a protrusion called a lamellipodium, while myosin contraction squeezes forward the back of the cell. In what is known as the ‘molecular clutch’ description of cell motility, forward movement results from the engagement of the acto-myosin motor with cell-matrix adhesions, thus transmitting force to the substrate and producing movement. However during cell translocation, clutch engagement is not perfect, and as a result, the cytoskeleton slips with respect to the substrate, undergoing backward (retrograde) flow in the direction of the cell body. Retrograde flow is therefore inversely proportional to cell speed and depends on adhesion and acto-myosin dynamics. Here we asked whether the molecular clutch was a general mechanism by measuring motility and retrograde flow for the Caenorhabditis elegans sperm cell in different adhesive conditions. These cells move by adhering to the substrate and emitting a dynamic lamellipodium, but the sperm cell does not contain an acto-myosin cytoskeleton. Instead the lamellipodium is formed by the assembly of major sperm protein, which has no biochemical or structural similarity to actin. We find that these cells display the same molecular clutch characteristics as acto-myosin containing cells. We further show that retrograde flow is produced both by cytoskeletal assembly and contractility in these cells. Overall this study shows that the molecular clutch hypothesis of how polymerization is transduced into motility via adhesions is a general description of cell movement regardless of the composition of the cytoskeleton.

  5. Extending the molecular clutch beyond actin-based cell motility

    PubMed Central

    Havrylenko, Svitlana; Mezanges, Xavier; Batchelder, Ellen; Plastino, Julie

    2014-01-01

    Many cell movements occur via polymerization of the actin cytoskeleton beneath the plasma membrane at the front of the cell, forming a protrusion called a lamellipodium, while myosin contraction squeezes forward the back of the cell. In what is known as the “molecular clutch” description of cell motility, forward movement results from the engagement of the acto-myosin motor with cell-matrix adhesions, thus transmitting force to the substrate and producing movement. However during cell translocation, clutch engagement is not perfect, and as a result, the cytoskeleton slips with respect to the substrate, undergoing backward (retrograde) flow in the direction of the cell body. Retrograde flow is therefore inversely proportional to cell speed and depends on adhesion and acto-myosin dynamics. Here we asked whether the molecular clutch was a general mechanism by measuring motility and retrograde flow for the Caenorhabditis elegans sperm cell in different adhesive conditions. These cells move by adhering to the substrate and emitting a dynamic lamellipodium, but the sperm cell does not contain an acto-myosin cytoskeleton. Instead the lamellipodium is formed by the assembly of Major Sperm Protein (MSP), which has no biochemical or structural similarity to actin. We find that these cells display the same molecular clutch characteristics as acto-myosin containing cells. We further show that retrograde flow is produced both by cytoskeletal assembly and contractility in these cells. Overall this study shows that the molecular clutch hypothesis of how polymerization is transduced into motility via adhesions is a general description of cell movement regardless of the composition of the cytoskeleton. PMID:25383039

  6. Activation of microbubbles by low-intensity pulsed ultrasound enhances the cytotoxicity of curcumin involving apoptosis induction and cell motility inhibition in human breast cancer MDA-MB-231 cells.

    PubMed

    Li, Yixiang; Wang, Pan; Chen, Xiyang; Hu, Jianmin; Liu, Yichen; Wang, Xiaobing; Liu, Quanhong

    2016-11-01

    Ultrasound and microbubbles-mediated drug delivery has become a promising strategy to promote drug delivery and its therapeutic efficacy. The aim of this research was to assess the effects of microbubbles (MBs)-combined low-intensity pulsed ultrasound (LPUS) on the delivery and cytotoxicity of curcumin (Cur) to human breast cancer MDA-MB-231 cells. Under the experimental condition, MBs raised the level of acoustic cavitation and enhanced plasma membrane permeability; and cellular uptake of Cur was notably improved by LPUS-MBs treatment, aggravating Cur-induced MDA-MB-231 cells death. The combined treatment markedly caused more obvious changes of cell morphology, F-actin cytoskeleton damage and cell migration inhibition. Our results demonstrated that combination of MBs and LPUS may be an efficient strategy for improving anti-tumor effect of Cur, suggesting a potential effective method for antineoplastic therapy. PMID:27245953

  7. Activation of microbubbles by low-intensity pulsed ultrasound enhances the cytotoxicity of curcumin involving apoptosis induction and cell motility inhibition in human breast cancer MDA-MB-231 cells.

    PubMed

    Li, Yixiang; Wang, Pan; Chen, Xiyang; Hu, Jianmin; Liu, Yichen; Wang, Xiaobing; Liu, Quanhong

    2016-11-01

    Ultrasound and microbubbles-mediated drug delivery has become a promising strategy to promote drug delivery and its therapeutic efficacy. The aim of this research was to assess the effects of microbubbles (MBs)-combined low-intensity pulsed ultrasound (LPUS) on the delivery and cytotoxicity of curcumin (Cur) to human breast cancer MDA-MB-231 cells. Under the experimental condition, MBs raised the level of acoustic cavitation and enhanced plasma membrane permeability; and cellular uptake of Cur was notably improved by LPUS-MBs treatment, aggravating Cur-induced MDA-MB-231 cells death. The combined treatment markedly caused more obvious changes of cell morphology, F-actin cytoskeleton damage and cell migration inhibition. Our results demonstrated that combination of MBs and LPUS may be an efficient strategy for improving anti-tumor effect of Cur, suggesting a potential effective method for antineoplastic therapy.

  8. Where to Go: Breaking the Symmetry in Cell Motility

    PubMed Central

    2016-01-01

    Cell migration in the “correct” direction is pivotal for many biological processes. Although most work is devoted to its molecular mechanisms, the cell’s preference for one direction over others, thus overcoming intrinsic random motility, epitomizes a profound principle that underlies all complex systems: the choice of one axis, in structure or motion, from a uniform or symmetric set of options. Explaining directional motility by an external chemo-attractant gradient does not solve but only shifts the problem of causation: whence the gradient? A new study in PLOS Biology shows cell migration in a self-generated gradient, offering an opportunity to take a broader look at the old dualism of extrinsic instruction versus intrinsic symmetry-breaking in cell biology. PMID:27196433

  9. Region-Specific Microtubule Transport in Motile Cells

    PubMed Central

    Yvon, Anne-Marie C.; Wadsworth, Patricia

    2000-01-01

    Photoactivation and photobleaching of fluorescence were used to determine the mechanism by which microtubules (MTs) are remodeled in PtK2 cells during fibroblast-like motility in response to hepatocyte growth factor (HGF). The data show that MTs are transported during cell motility in an actomyosin-dependent manner, and that the direction of transport depends on the dominant force in the region examined. MTs in the leading lamella move rearward relative to the substrate, as has been reported in newt cells (Waterman-Storer, C.M., and E.D. Salmon. 1997. J. Cell Biol. 139:417–434), whereas MTs in the cell body and in the retraction tail move forward, in the direction of cell locomotion. In the transition zone between the peripheral lamella and the cell body, a subset of MTs remains stationary with respect to the substrate, whereas neighboring MTs are transported either forward, with the cell body, or rearward, with actomyosin retrograde flow. In addition to transport, the photoactivated region frequently broadens, indicating that individual marked MTs are moved either at different rates or in different directions. Mark broadening is also observed in nonmotile cells, indicating that this aspect of transport is independent of cell locomotion. Quantitative measurements of the dissipation of photoactivated fluorescence show that, compared with MTs in control nonmotile cells, MT turnover is increased twofold in the lamella of HGF-treated cells but unchanged in the retraction tail, demonstrating that microtubule turnover is regionally regulated. PMID:11086002

  10. The Influence of Electric Field and Confinement on Cell Motility

    PubMed Central

    Huang, Yu-Ja; Samorajski, Justin; Kreimer, Rachel; Searson, Peter C.

    2013-01-01

    The ability of cells to sense and respond to endogenous electric fields is important in processes such as wound healing, development, and nerve regeneration. In cell culture, many epithelial and endothelial cell types respond to an electric field of magnitude similar to endogenous electric fields by moving preferentially either parallel or antiparallel to the field vector, a process known as galvanotaxis. Here we report on the influence of dc electric field and confinement on the motility of fibroblast cells using a chip-based platform. From analysis of cell paths we show that the influence of electric field on motility is much more complex than simply imposing a directional bias towards the cathode or anode. The cell velocity, directedness, as well as the parallel and perpendicular components of the segments along the cell path are dependent on the magnitude of the electric field. Forces in the directions perpendicular and parallel to the electric field are in competition with one another in a voltage-dependent manner, which ultimately govern the trajectories of the cells in the presence of an electric field. To further investigate the effects of cell reorientation in the presence of a field, cells are confined within microchannels to physically prohibit the alignment seen in 2D environment. Interestingly, we found that confinement results in an increase in cell velocity both in the absence and presence of an electric field compared to migration in 2D. PMID:23555674

  11. The influence of electric field and confinement on cell motility.

    PubMed

    Huang, Yu-Ja; Samorajski, Justin; Kreimer, Rachel; Searson, Peter C

    2013-01-01

    The ability of cells to sense and respond to endogenous electric fields is important in processes such as wound healing, development, and nerve regeneration. In cell culture, many epithelial and endothelial cell types respond to an electric field of magnitude similar to endogenous electric fields by moving preferentially either parallel or antiparallel to the field vector, a process known as galvanotaxis. Here we report on the influence of dc electric field and confinement on the motility of fibroblast cells using a chip-based platform. From analysis of cell paths we show that the influence of electric field on motility is much more complex than simply imposing a directional bias towards the cathode or anode. The cell velocity, directedness, as well as the parallel and perpendicular components of the segments along the cell path are dependent on the magnitude of the electric field. Forces in the directions perpendicular and parallel to the electric field are in competition with one another in a voltage-dependent manner, which ultimately govern the trajectories of the cells in the presence of an electric field. To further investigate the effects of cell reorientation in the presence of a field, cells are confined within microchannels to physically prohibit the alignment seen in 2D environment. Interestingly, we found that confinement results in an increase in cell velocity both in the absence and presence of an electric field compared to migration in 2D.

  12. STIM1 overexpression promotes colorectal cancer progression, cell motility and COX-2 expression.

    PubMed

    Wang, J-Y; Sun, J; Huang, M-Y; Wang, Y-S; Hou, M-F; Sun, Y; He, H; Krishna, N; Chiu, S-J; Lin, S; Yang, S; Chang, W-C

    2015-08-13

    Tumor metastasis is the major cause of death among cancer patients, with >90% of cancer-related death attributable to the spreading of metastatic cells to secondary organs. Store-operated Ca(2+) entry (SOCE) is the predominant Ca(2+) entry mechanism in most cancer cells, and stromal interaction molecule 1 (STIM1) is the endoplasmic reticulum (ER) Ca(2+) sensor for store-operated channels. Here we reported that the STIM1 was overexpressed in colorectal cancer (CRC) patients. STIM1 overexpression in CRC was significantly associated with tumor size, depth of invasion, lymph node metastasis status and serum levels of carcinoembryonic antigen. Furthermore, ectopic expression of STIM1 promoted CRC cell motility, while depletion of STIM1 with short hairpin RNA inhibited CRC cell migration. Our data further suggested that STIM1 promoted CRC cell migration through increasing the expression of cyclooxygenase-2 (COX-2) and production of prostaglandin E2 (PGE2). Importantly, ectopically expressed COX-2 or exogenous PGE2 were able to rescue migration defect in STIM1 knockdown CRC cells, and inhibition of COX-2 with ibuprofen and indomethacin abrogated STIM1-mediated CRC cell motility. In short, our data provided clinicopathological significance for STIM1 and SOCE in CRC progression, and implicated a role for COX-2 in STIM1-mediated CRC metastasis. Our studies also suggested a new approach to inhibit STIM1-mediated metastasis with COX-2 inhibitors.

  13. Actin-dependent motility of melanosomes from fish retinal pigment epithelial (RPE) cells investigated using in vitro motility assays.

    PubMed

    McNeil, E L; Tacelosky, D; Basciano, P; Biallas, B; Williams, R; Damiani, P; Deacon, S; Fox, C; Stewart, B; Petruzzi, N; Osborn, C; Klinger, K; Sellers, J R; Smith, C King

    2004-06-01

    Melanosomes (pigment granules) within retinal pigment epithelial (RPE) cells of fish and amphibians undergo massive migrations in response to light conditions to control light flux to the retina. Previous research has shown that melanosome motility within apical projections of dissociated fish RPE cells requires an intact actin cytoskeleton, but the mechanisms and motors involved in melanosome transport in RPE have not been identified. Two in vitro motility assays, the Nitella assay and the sliding filament assay, were used to characterize actin-dependent motor activity of RPE melanosomes. Melanosomes applied to dissected filets of the Characean alga, Nitella, moved along actin cables at a mean rate of 2 microm/min, similar to the rate of melanosome motility in dissociated, cultured RPE cells. Path lengths of motile melanosomes ranged from 9 to 37 microm. Melanosome motility in the sliding filament assay was much more variable, ranging from 0.4-33 microm/min; 70% of velocities ranged from 1-15 microm/min. Latex beads coated with skeletal muscle myosin II and added to Nitella filets moved in the same direction as RPE melanosomes, indicating that the motility is barbed-end directed. Immunoblotting using antibodies against myosin VIIa and rab27a revealed that both proteins are enriched on melanosome membranes, suggesting that they could play a role in melanosome transport within apical projections of fish RPE.

  14. p27 as Jekyll and Hyde: regulation of cell cycle and cell motility.

    PubMed

    Larrea, Michelle D; Wander, Seth A; Slingerland, Joyce M

    2009-11-01

    p27 is a key regulator of cell proliferation. While it opposes cell cycle progression by binding to and inhibiting cyclin E-Cdk2, T157/T198 phosphorylation of p27 promotes its assembly of D-type cyclin-CDKs. In addition to its actions on the cell cycle, p27 regulates CDK-independent cytoplasmic functions. In human cancers, oncogenic activation of the PI3K signaling pathway often results in cytoplasmic mislocalization of p27. Cytoplasmic p27 plays an important role in cell motility and migration; it binds RhoA and modulates activation of the RhoA/ROCK cascade. p27:RhoA binding is facilitated by p27 phosphorylation at threonine 198. Accumulation of cytoplasmic p27 leads to increased cellular motility, a critical event in tumor metastasis. Further characterization of post-translational modifications governing p27 localization and its action on RhoA and the actin cytoskeleton may provide critical insights into human cancer metastasis.

  15. Dpp signaling directs cell motility and invasiveness during epithelial morphogenesis.

    PubMed

    Ninov, Nikolay; Menezes-Cabral, Sofia; Prat-Rojo, Carla; Manjón, Cristina; Weiss, Alexander; Pyrowolakis, George; Affolter, Markus; Martín-Blanco, Enrique

    2010-03-23

    Tissue remodeling in development and disease involves the coordinated invasion of neighboring territories and/or the replacement of entire cell populations. Cell guidance, cell matching, transitions from passive to migratory epithelia, cell growth and death, and extracellular matrix remodeling all impinge on epithelial spreading. Significantly, the extracellular signals that direct these activities and the specific cellular elements and mechanisms regulated by these signals remain in most cases to be identified. To address these issues, we performed an analysis of histoblasts (Drosophila abdominal epithelial founder cells) on their transition from a dormant state to active migration replacing obsolete larval epidermal cells (LECs). We found that during expansion, Decapentaplegic (Dpp) secreted from surrounding LECs leads to graded pathway activation in cells at the periphery of histoblast nests. Across nests, Dpp activity confers differential cellular behavior and motility by modulating cell-cell contacts, the organization and activity of the cytoskeleton, and histoblast attachment to the substrate. Furthermore, Dpp also prevents the premature death of LECs, allowing the coordination of histoblast expansion to LEC delamination. Dpp signaling activity directing histoblast spreading and invasiveness mimics transforming growth factor-beta and bone morphogenetic proteins' role in enhancing the motility and invasiveness of cancer cells, resulting in the promotion of metastasis. PMID:20226662

  16. Collective cell motility promotes chemotactic prowess and resistance to chemorepulsion.

    PubMed

    Malet-Engra, Gema; Yu, Weimiao; Oldani, Amanda; Rey-Barroso, Javier; Gov, Nir S; Scita, Giorgio; Dupré, Loïc

    2015-01-19

    Collective cell migration is a widespread biological phenomenon, whereby groups of highly coordinated, adherent cells move in a polarized fashion. This migration mode is a hallmark of tissue morphogenesis during development and repair and of solid tumor dissemination. In addition to circulating as solitary cells, lymphoid malignancies can assemble into tissues as multicellular aggregates. Whether malignant lymphocytes are capable of coordinating their motility in the context of chemokine gradients is, however, unknown. Here, we show that, upon exposure to CCL19 or CXCL12 gradients, malignant B and T lymphocytes assemble into clusters that migrate directionally and display a wider chemotactic sensitivity than individual cells. Physical modeling recapitulates cluster motility statistics and shows that intracluster cell cohesion results in noise reduction and enhanced directionality. Quantitative image analysis reveals that cluster migration runs are periodically interrupted by transitory rotation and random phases that favor leader cell turnover. Additionally, internalization of CCR7 in leader cells is accompanied by protrusion retraction, loss of polarity, and the ensuing replacement by new leader cells. These mechanisms ensure sustained forward migration and resistance to chemorepulsion, a behavior of individual cells exposed to steep CCL19 gradients that depends on CCR7 endocytosis. Thus, coordinated cluster dynamics confer distinct chemotactic properties, highlighting unexpected features of lymphoid cell migration. PMID:25578904

  17. HDAC6 Modulates Cell Motility by Altering the Acetylation Level of Cortactin

    PubMed Central

    Zhang, Xiaohong; Yuan, Zhigang; Zhang, Yingtao; Yong, Sarah; Salas-Burgos, Alexis; Koomen, John; Olashaw, Nancy; Parsons, J. Thomas; Yang, Xiang-Jiao; Dent, Sharon R.; Yao, Tso-Pang; Lane, William S.; Seto, Edward

    2009-01-01

    Summary Histone deacetylase 6 (HDAC6) is a tubulin-specific deacetylase that regulates microtubule-dependent cell movement. In this study, we identify the F-actin-binding protein, cortactin, as a HDAC6 substrate. We demonstrate that HDAC6 binds cortactin and that overexpression of HDAC6 leads to hypoacetylation of cortactin, while inhibition of HDAC6 activity leads to cortactin hyperacetylation. HDAC6 alters the ability of cortactin to bind F-actin by modulating a “charge patch” in its repeat region. Introduction of charge-preserving or charge-neutralizing mutations in this cortactin repeat region correlates with the gain or loss of F-actin binding ability, respectively. Cells expressing a charge-neutralizing cortactin mutant were less motile than control cells or cells expressing a charge-preserving mutant. These findings suggest that, in addition to its role in microtubule-dependent cell motility, HDAC6 influences actin-dependent cell motility by altering the acetylation status of cortactin, which, in turn, changes the F-actin binding activity of cortactin. PMID:17643370

  18. Membrane tension and cytoskeleton organization in cell motility

    NASA Astrophysics Data System (ADS)

    Sens, Pierre; Plastino, Julie

    2015-07-01

    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.

  19. 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

  20. Electroacupuncture at ST25 inhibits jejunal motility: Role of sympathetic pathways and TRPV1

    PubMed Central

    Yu, Zhi; Zhang, Na; Lu, Chun-Xia; Pang, Ting-Ting; Wang, Kai-Yue; Jiang, Jing-Feng; Zhu, Bing; Xu, Bin

    2016-01-01

    AIM: To investigate whether electroacupuncture (EA) at ST25 affects jejunal motility in vivo and if so, whether a sympathetic pathway is involved. METHODS: Jejunal motility was assessed using a manometric balloon placed in the jejunum approximately about 3-5 cm away from the suspensory ligament of the duodenum in anesthetized animals. The effects of EA at ST25 were measured in male Sprague-Dawley rats, some of which were treated with propranolol or clenbuterol (EA intensities: 1, 3, 5, 7, and 9 mA), and in male transient receptor potential vanilloid-1 (TRPV1) (capsaicin receptor) knockout mice (EA intensities: 1, 2, and 4 mA). RESULTS: Anesthetized rats exhibited three types of fasting jejunal motor patterns (types A, B, and C), and only type C rats responded to EA stimulation. In type C rats, EA at ST25 significantly suppressed the motor activity of the jejunum in an intensity-dependent manner. The inhibitory effect of EA was weakened by propranolol (β adrenoceptor antagonist) and disappeared with clenbuterol (β adrenoceptor agonist) induced inhibition of motility, suggesting that the effect of EA on motility is mediated via a sympathetic pathway. Compared with wild-type mice, EA at ST25 was less effective in TRPV1 knockout mice, suggesting that this multi-modal sensor channel participates in the mechanism. CONCLUSION: EA at ST25 was found to inhibit jejunal motility in an intensity-dependent manner, via a mechanism in which sympathetic nerves and TRPV1 receptors play an important role. PMID:26855542

  1. Membrane tension feedback on shape and motility of eukaryotic cells

    NASA Astrophysics Data System (ADS)

    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 in 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.

  2. 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

  3. Involvement of ephrin receptor A4 in pancreatic cancer cell motility and invasion

    PubMed Central

    LIU, CHENGLI; HUANG, HUI; WANG, CHENG; KONG, YALIN; ZHANG, HONGYI

    2014-01-01

    Ephrin (EPH) receptors can be classified into EPHA and EPHB receptors and are important in diverse cellular processes. EPHA4, a member of the EPHA receptors, has been demonstrated to be elevated in various human cancers and involved in the tumor progression. However, the role of EPHA4 in pancreatic cancer cells remains unclear. Therefore, the present study transfected Panc-1 and BxPC-3 cells with small interfering RNA (siRNA) to knockdown the expression of EPHA4. Wound healing and invasion assays were then performed to assess the effect of EPHA4 knockdown on the motility and invasion of pancreatic cancer cells. The results demonstrated that the knockdown of EPHA4 by siRNA inhibits the motility and invasion of pancreatic cancer cells. Furthermore, gelatin zymography assay showed that EPHA4 may regulate the activity of matrix metalloproteinase (MMP)-2. In addition, the knockdown of EPHA4 increased the expression of epithelial (E)-cadherin, as well as decreased the expression of Snail. Overall, these results suggested that EPHA4 may promote the motility and invasion of pancreatic cancer cells via the upregulation of MMP-2 and Snail, as well as the downregulation of E-cadherin. Thus, EPHA4 may act as a useful target for the treatment of pancreatic cancer. PMID:24932309

  4. Retraction in amoeboid cell motility powered by cytoskeletal dynamics.

    PubMed

    Miao, Long; Vanderlinde, Orion; Stewart, Murray; Roberts, Thomas M

    2003-11-21

    Cells crawl by coupling protrusion of their leading edge with retraction of their cell body. Protrusion is generated by the polymerization and bundling of filaments, but the mechanism of retraction is less clear. We have reconstituted retraction in vitro by adding Yersinia tyrosine phosphatase to the major sperm protein-based motility apparatus assembled from Ascaris sperm extracts. Retraction in vitro parallels that observed in vivo and is generated primarily by disassembly and rearrangement of the cytoskeleton. Therefore, cytoskeletal dynamics alone, unassisted by conventional motors, are able to generate both of these central components of amoeboid locomotion.

  5. A kinetic mechanism for cell sorting based on local variations in cell motility

    PubMed Central

    Strandkvist, Charlotte; Juul, Jeppe; Baum, Buzz; Kabla, Alexandre J.; Duke, Tom

    2014-01-01

    Our current understanding of cell sorting relies on physical difference, either in the interfacial properties or motile force, between cell types. But is such asymmetry a prerequisite for cell sorting? We test this using a minimal model in which the two cell populations are identical with respect to their physical properties and differences in motility arise solely from how cells interact with their surroundings. The model resembles the Schelling model used in social sciences to study segregation phenomena at the scale of societies. Our results demonstrate that segregation can emerge solely from cell motility being a dynamic property that changes in response to the local environment of the cell, but that additional mechanisms are necessary to reproduce the envelopment behaviour observed in vitro. The time course of segregation follows a power law, in agreement with the scaling reported from experiment and in other models of motility-driven segregation. PMID:25485079

  6. 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

  7. Lattice-free models of directed cell motility

    NASA Astrophysics Data System (ADS)

    Irons, Carolyn; Plank, Michael J.; Simpson, Matthew J.

    2016-01-01

    Directed cell migration often occurs when individual cells move in response to an external chemical stimulus. Cells can respond by moving in either the direction of increasing (chemoattraction) or decreasing (chemorepulsion) concentration. Many previous models of directed cell migration use a lattice-based framework where agents undergo a lattice-based random walk and the direction of nearest-neighbour motility events is biased in a preferred direction. Such lattice-based models can lead to unrealistic configurations of agents, since the agents always move on an artificial lattice structure which is never observed experimentally. We present a lattice-free model of directed cell migration that incorporates two key features. First, agents move on a continuous domain, with the possibility that there is some preferred direction of motion. Second, to be consistent with experimental observations, we enforce a crowding mechanism so that motility events that would lead to agent overlap are not permitted. We compare simulation data from the new lattice-free model with a more traditional lattice-based model. To provide additional insight into the lattice-free model, we construct an approximate conservation statement which corresponds to a nonlinear advection-diffusion equation in the continuum limit. The solution of this mean-field model compares well with averaged data from the individual-based model.

  8. Cadmium inhibits motility, activities of plasma membrane Ca(2+)-ATPase and axonemal dynein-ATPase of human spermatozoa.

    PubMed

    Da Costa, R; Botana, D; Piñero, S; Proverbio, F; Marín, R

    2016-05-01

    Cd(2+) has been associated with decreased sperm motility in individuals exposed to this element, such as smokers. Among other factors, this lowered motility could be the result of inhibition exerted by Cd(2+) on the activity of the sperm ATPases associated with sperm motility. In this study, we evaluated the plasma membrane Ca(2+)-ATPase and the axonemal dynein-ATPase activities as well as sperm motility, in the presence of different free Cd(2+) concentrations in the assay media. It was found that spermatozoa incubated for 5 h in a medium containing 25 nm free Cd(2+) showed a significant inhibition of progressive motility, reaching values even lower at higher Cd(2+) concentrations. In addition, it was found that the activity of the plasma membrane Ca(2+)-ATPase reached maximal inhibition at 50 nm free Cd(2+), with a K50% inhibition of 18.3 nm free Cd(2+). The dynein-ATPase activity was maximally inhibited by 25 nm free Cd(2+) in the assay medium, with a K50% inhibition of 11.3 nm Cd(2+). Our results indicate that the decreased activity of the sperm ATPases might have a critical importance in the biochemical mechanisms underlying the decreased sperm motility of individuals exposed to Cd(2+). PMID:26259968

  9. Extracellular Vesicles from Vascular Endothelial Cells Promote Survival, Proliferation and Motility of Oligodendrocyte Precursor Cells

    PubMed Central

    Kurachi, Masashi; Mikuni, Masahiko; Ishizaki, Yasuki

    2016-01-01

    We previously examined the effect of brain microvascular endothelial cell (MVEC) transplantation on rat white matter infarction, and found that MVEC transplantation promoted remyelination of demyelinated axons in the infarct region and reduced apoptotic death of oligodendrocyte precursor cells (OPCs). We also found that the conditioned medium (CM) from cultured MVECs inhibited apoptosis of cultured OPCs. In this study, we examined contribution of extracellular vesicles (EVs) contained in the CM to its inhibitory effect on OPC apoptosis. Removal of EVs from the CM by ultracentrifugation reduced its inhibitory effect on OPC apoptosis. To confirm whether EVs derived from MVECs are taken up by cultured OPCs, we labeled EVs with PKH67, a fluorescent dye, and added them to OPC cultures. Many vesicular structures labeled with PKH67 were found within OPCs immediately after their addition. Next we examined the effect of MVEC-derived EVs on OPC behaviors. After 2 days in culture with EVs, there was significantly less pyknotic and more BrdU-positive OPCs when compared to control. We also examined the effect of EVs on motility of OPCs. OPCs migrated longer in the presence of EVs when compared to control. To examine whether these effects on cultured OPCs are shared by EVs from endothelial cells, we prepared EVs from conditioned media of several types of endothelial cells, and tested their effects on cultured OPCs. EVs from all types of endothelial cells we examined reduced apoptosis of OPCs and promoted their motility. Identification of the molecules contained in EVs from endothelial cells may prove helpful for establishment of effective therapies for demyelinating diseases. PMID:27403742

  10. Motile apparatus in Vallisneria leaf cells. I. Organization of microfilaments.

    PubMed

    Yamaguchi, Y; Nagai, R

    1981-04-01

    The organization of the microfilaments in epidermal cells of Vallisneria leaves was investigated with respect to the induction of cytoplasmic streaming (secondary streaming). In many of the epidermal cells, cytoplasm exhibited rotational streaming along the anticlinal wall of the cell after exposure around the anticlinal wall. The bundles were arrayed in parallel to the streaming direction. They were recognized usually as 10-40 closely packed dense dots in cross-section. The spacing between bundles was not even. Bundles tended to form groups of 4 to 5 in which the spacing between bundles was usually 0.3 to 0.5 micrometer. The microfilaments were identified as F-actin. Together with the fact that rotational streaming in Vallisneria cells by cytochalasin B, the motile mechanism of secondary streaming was concluded to be similar in its essential features to the cytoplasmic streaming seen in Characean cell (primary streaming). In epidermal cells that had been kept under low-intensity light the cytoplasm and the cytoplasmic streaming occurred in these cells. The bundles of microfilaments remained in the very thin layer of cytoplasm lining the anticlinal wall, although they were fewer and somewhat loosely packed. EGTA at appropriate concentration could induce cytoplasmic streaming in these cells. The mechanism of the induction is discussed on the basis of the effectiveness of EGTA and the requirement of a low concentration of free Ca2+ for cytoplasmic streaming in Characean cells. PMID:6792210

  11. Na,K-ATPase β-Subunit Is Required for Epithelial Polarization, Suppression of Invasion, and Cell Motility

    PubMed Central

    Rajasekaran, Sigrid A.; Palmer, Lawrence G.; Quan, Karina; Harper, Jeffrey F.; Ball, William J.; Bander, Neil H.; Soler, Alejandro Peralta; Rajasekaran, Ayyappan K.

    2001-01-01

    The cell adhesion molecule E-cadherin has been implicated in maintaining the polarized phenotype of epithelial cells and suppression of invasiveness and motility of carcinoma cells. Na,K-ATPase, consisting of an α- and β-subunit, maintains the sodium gradient across the plasma membrane. A functional relationship between E-cadherin and Na,K-ATPase has not previously been described. We present evidence that the Na,K-ATPase plays a crucial role in E-cadherin–mediated development of epithelial polarity, and suppression of invasiveness and motility of carcinoma cells. Moloney sarcoma virus-transformed Madin-Darby canine kidney cells (MSV-MDCK) have highly reduced levels of E-cadherin and β1-subunit of Na,K-ATPase. Forced expression of E-cadherin in MSV-MDCK cells did not reestablish epithelial polarity or inhibit the invasiveness and motility of these cells. In contrast, expression of E-cadherin and Na,K-ATPase β1-subunit induced epithelial polarization, including the formation of tight junctions and desmosomes, abolished invasiveness, and reduced cell motility in MSV-MDCK cells. Our results suggest that E-cadherin–mediated cell-cell adhesion requires the Na,K-ATPase β-subunit's function to induce epithelial polarization and suppress invasiveness and motility of carcinoma cells. Involvement of the β1-subunit of Na,K-ATPase in the polarized phenotype of epithelial cells reveals a novel link between the structural organization and vectorial ion transport function of epithelial cells. PMID:11179415

  12. Trifluoperazine inhibits the MgATPase activity and in vitro motility of conventional and unconventional myosins.

    PubMed

    Sellers, James R; Wang, Fei; Chantler, Peter D

    2003-01-01

    Trifluoperazine, a calmodulin antagonist, has recently been shown to inhibit the MgATPase activity of scallop myosin in the absence of light chain dissociation (Patel et al. (2000) J Biol Chem 275: 4880-4888). To investigate the generality of this observation and the mechanism by which it occurs, we have examined the ability of trifluoperazine to inhibit the enzymatic properties of other conventional and unconventional myosins. We show that trifluoperazine can inhibit the actin-activated MgATPase activity of rabbit skeletal muscle myosin II heavy meromyosin (HMM), phosphorylated turkey gizzard smooth muscle myosin II HMM, phosphorylated human nonmuscle myosin IIA HMM and myosin V subfragment-1 (S1). In all cases half maximal inhibition occurred at 50-75 microM trifluoperazine while light chains (myosin II) or calmodulin (myosin V) remained associated with the heavy chains. In vitro motility of all myosins tested was completely inhibited by trifluoperazine. Chymotryptic digestion of baculovirus-expressed myosin V HMM possessing only two calmodulin binding sites yielded a minimal motor fragment with no bound calmodulin. The MgATPase of this fragment was inhibited by trifluoperazine over the same range of concentrations as the S1 fragment of myosin.

  13. Synchronization of Spontaneous Active Motility of Hair Cell Bundles

    PubMed Central

    Zhang, Tracy-Ying; Ji, Seung; Bozovic, Dolores

    2015-01-01

    Hair cells of the inner ear exhibit an active process, believed to be crucial for achieving the sensitivity of auditory and vestibular detection. One of the manifestations of the active process is the occurrence of spontaneous hair bundle oscillations in vitro. Hair bundles are coupled by overlying membranes in vivo; hence, explaining the potential role of innate bundle motility in the generation of otoacoustic emissions requires an understanding of the effects of coupling on the active bundle dynamics. We used microbeads to connect small groups of hair cell bundles, using in vitro preparations that maintain their innate oscillations. Our experiments demonstrate robust synchronization of spontaneous oscillations, with either 1:1 or multi-mode phase-locking. The frequency of synchronized oscillation was found to be near the mean of the innate frequencies of individual bundles. Coupling also led to an improved regularity of entrained oscillations, demonstrated by an increase in the quality factor. PMID:26540409

  14. Synchronization of Spontaneous Active Motility of Hair Cell Bundles.

    PubMed

    Zhang, Tracy-Ying; Ji, Seung; Bozovic, Dolores

    2015-01-01

    Hair cells of the inner ear exhibit an active process, believed to be crucial for achieving the sensitivity of auditory and vestibular detection. One of the manifestations of the active process is the occurrence of spontaneous hair bundle oscillations in vitro. Hair bundles are coupled by overlying membranes in vivo; hence, explaining the potential role of innate bundle motility in the generation of otoacoustic emissions requires an understanding of the effects of coupling on the active bundle dynamics. We used microbeads to connect small groups of hair cell bundles, using in vitro preparations that maintain their innate oscillations. Our experiments demonstrate robust synchronization of spontaneous oscillations, with either 1:1 or multi-mode phase-locking. The frequency of synchronized oscillation was found to be near the mean of the innate frequencies of individual bundles. Coupling also led to an improved regularity of entrained oscillations, demonstrated by an increase in the quality factor. PMID:26540409

  15. Influence of engineered surface on cell directionality and motility.

    PubMed

    Tang, Qing Yuan; Tong, Wing Yin; Shi, Jue; Shi, Peng; Lam, Yun Wah; Pang, Stella W

    2014-03-01

    Control of cell migration is important in numerous key biological processes, and is implicated in pathological conditions such as cancer metastasis and inflammatory diseases. Many previous studies indicated that cell migration could be guided by micropatterns fabricated on cell culture surfaces. In this study, we designed a polydimethylsiloxane cell culture substrate with gratings punctuated by corners and ends, and studied its effects on the behavior of MC3T3-E1 osteoblast cells. MC3T3-E1 cells elongated and aligned with the gratings, and the migration paths of the cells appeared to be guided by the grating pattern. Interestingly, more than 88% of the cells cultured on these patterns were observed to reverse their migration directions at least once during the 16 h examination period. Most of the reversal events occurred at the corners and the ends of the pattern, suggesting these localized topographical features induce an abrupt loss in directional persistence. Moreover, the cell speed was observed to increase temporarily right after each directional reversal. Focal adhesion complexes were more well-established in cells on the angular gratings than on flat surfaces, but the formation of filipodia appeared to be imbalanced at the corners and the ends, possibly leading to the loss of directional persistence. This study describes the first engineered cell culture surface that consistently induces changes in the directional persistence of adherent cells. This will provide an experimental model for the study of this phenomenon and a valuable platform to control the cell motility and directionality, which can be used for cell screening and selection. PMID:24589941

  16. Reversible stimulation of lymphocyte motility by cultured high endothelial cells: mediation by L-selectin.

    PubMed Central

    Harris, H; Miyasaka, M

    1995-01-01

    Lymphocyte emigration from blood into peripheral lymph nodes is mediated by specialized high endothelial cells (HEC) lining the postcapillary venules. A current model for this process postulates that it occurs in three steps: weak, selectin-mediated interactions tether lymphocytes to the blood vessel wall; the lymphocytes are activated to increase the affinity of integrin-dependent adhesion and enhance motility; and finally the lymphocytes migrate actively across the endothelial cell layer. Some features of this model are simulated in vitro by cultured HEC, which support the adhesion and transmigration of lymphocytes. In particular, cultured HEC stimulate lymphocytes to change shape from spherical to polar. This shape change provides a convenient assay of the motility activation of lymphocytes. In this paper it is shown that this occurs without the lymphocytes becoming tightly adherent, but depends on contact with the endothelial cell surface. The shape change is labile: non-adherent polar lymphocytes removed from HEC revert to round with a half-time of less than 8 min. Reagents which block the interaction of L-selectin with its ligands inhibit the HEC-induced shape change; these include mannose-6-phosphate, fucoidan, polyphosphomannan ester, treatment of HEC with sialidases and an anti-L-selectin monoclonal antibody known to block its lectin function. The change in shape is partially inhibited by antisera to the L-selectin ligand GlyCAM-1. Thus it is concluded that in this in vitro system, L-selectin-mediated binding of lymphocytes to HEC is essential for optimal induction of the shape change. Lymphocytes change shape in response to cultured HEC without loss of surface L-selectin, although activation stimuli are known to promote shedding of neutrophil L-selectin as well as motility and increased adhesiveness. However, the lymphocyte change in shape is a reversible process, and this may have implications for the nature and sequence of the signals transmitted from

  17. PTP1B inhibitor promotes endothelial cell motility by activating the DOCK180/Rac1 pathway

    PubMed Central

    Wang, Yuan; Yan, Feng; Ye, Qing; Wu, Xiao; Jiang, Fan

    2016-01-01

    Promoting endothelial cell (EC) migration is important not only for therapeutic angiogenesis, but also for accelerating re-endothelialization after vessel injury. Several recent studies have shown that inhibition of protein tyrosine phosphatase 1B (PTP1B) may promote EC migration and angiogenesis by enhancing the vascular endothelial growth factor receptor-2 (VEGFR2) signalling. In the present study, we demonstrated that PTP1B inhibitor could promote EC adhesion, spreading and migration, which were abolished by the inhibitor of Rac1 but not RhoA GTPase. PTP1B inhibitor significantly increased phosphorylation of p130Cas, and the interactions among p130Cas, Crk and DOCK180; whereas the phosphorylation levels of focal adhesion kinase, Src, paxillin, or Vav2 were unchanged. Gene silencing of DOCK180, but not Vav2, abrogated the effects of PTP1B inhibitor on EC motility. The effects of PTP1B inhibitor on EC motility and p130Cas/DOCK180 activation persisted in the presence of the VEGFR2 antagonist. In conclusion, we suggest that stimulation of the DOCK180 pathway represents an alternative mechanism of PTP1B inhibitor-stimulated EC motility, which does not require concomitant VEGFR2 activation as a prerequisite. Therefore, PTP1B inhibitor may be a useful therapeutic strategy for promoting EC migration in cardiovascular patients in which the VEGF/VEGFR functions are compromised. PMID:27052191

  18. Motility, viability, and calcium in the sperm cells.

    PubMed

    Parodi, Jorge

    2014-04-01

    Sperm cells are complicated in vitro models. Their viability is limited, and physiology is complex. The study of their properties is of great application in the animal production as viable and functional gametes are essential. It has been shown that the decrease of sperm cell viability parallels an increase of the reactive oxygen species (ROS). Reactive oxygen species is secondary to normal metabolic processes of the cell-like flagellar movement. There is evidence of strategies that reduce ROS levels by using exogenous or endogenous antioxidants with the intention that seminal plasma protects the sperm cells and increases viability. Perhaps viability can increase by reducing that flagellar movement which is regulated by calcium. The phenomenon has not been fully characterized, but it is established that in certain mammalian models, the entrance of calcium via specific channels such as CATsper or voltage-dependent channels, signals flagellar movement. Previous reports have indicated that a change in the concentration of calcium or if the temperature is altered, the function of mammal sperm cells is reduced or blocked and viability prolonged. Fish sperm can remain immobile for several weeks but when activated the number of mobile and viable sperm is reduced at a faster rate. However, if the cells are not mobilized the semen can be preserved for longer periods. As presented in this paper, this supports the notion that by modulating calcium channels to reduce motility the viability of these cells can increase.

  19. The interplay between cell motility and tissue architecture

    NASA Astrophysics Data System (ADS)

    Tanner, Kandice

    2013-03-01

    Glandular tissue form arboreal networks comprised of acini and tubes. Loss of structure is concomitant with the in vivo pathologic state. In vitro models have been shown to recapitulate the functional units of the mammary gland and other organs. Despite our much improved understanding gleaned from both in vitro and in vivo interrogation, the mechanisms by which cells are able to achieve the correct tissue organization remain elusive. How do single mammary epithelial cells form polarized acini when cultured in a surrogate basement membrane gel but not on 2D surfaces? Simply put, how does a cell know which way is up? Why do malignant breast cells show a differential response in that they form non-polarized aggregates? Recently, it was determined that non-malignant cells undergo multiple rotations to establish acini while tumor cells are randomly motile during tumor formation. Can it be that a tumor cell has simply lost its way. This research was supported by the Intramural Research Program of the NIH, National Cancer Institute.

  20. The role of palladin in actin organization and cell motility

    PubMed Central

    Goicoechea, Silvia M.; Arneman, Daniel; Otey, Carol A.

    2008-01-01

    Palladin is a widely expressed protein found in stress fibers, focal adhesions, growth cones, Z-discs, and other actin-based subcellular structures. It belongs to a small gene family that includes the Z-disc proteins myopalladin and myotilin, all of which share similar Ig-like domains. Recent advances have shown that palladin shares with myotilin the ability to bind directly to F-actin, and to crosslink actin filaments into bundles, in vitro. Studies in a variety of cultured cells suggest that the actin-organizing activity of palladin plays a central role in promoting cell motility. Correlative evidence also supports this hypothesis, as palladin levels are typically upregulated in cells that are actively migrating: in developing vertebrate embryos, in cells along a wound edge, and in metastatic cancer cells. Recently, a mutation in the human palladin gene was implicated in an unusually penetrant form of inherited pancreatic cancer, which has stimulated new ideas about the role of palladin in invasive cancer. PMID:18342394

  1. Vimentin-mediated regulation of cell motility through modulation of beta4 integrin protein levels in oral tumor derived cells.

    PubMed

    Dmello, Crismita; Sawant, Sharada; Alam, Hunain; Gangadaran, Prakash; Tiwari, Richa; Dongre, Harsh; Rana, Neha; Barve, Sai; Costea, Daniela Elena; Chaukar, Davendra; Kane, Shubhada; Pant, Harish; Vaidya, Milind

    2016-01-01

    Vimentin expression correlates well with migratory and invasive potential of the carcinoma cells. The molecular mechanism by which vimentin regulates cell motility is not yet clear. Here, we addressed this issue by depleting vimentin in oral squamous cell carcinoma derived cell line. Vimentin knockdown cells showed enhanced adhesion and spreading to laminin-5. However, we found that they were less invasive as compared to the vector control cells. In addition, signaling associated with adhesion behavior of the cell was increased in vimentin knockdown clones. These findings suggest that the normal function of β4 integrin as mechanical adhesive device is enhanced upon vimentin downregulation. As a proof of principle, the compromised invasive potential of vimentin depleted cells could be rescued upon blocking with β4 integrin adhesion-blocking (ASC-8) antibody or downregulation of β4 integrin in vimentin knockdown background. Interestingly, plectin which associates with α6β4 integrin in the hemidesmosomes, was also found to be upregulated in vimentin knockdown clones. Furthermore, experiments on lysosome and proteasome inhibition revealed that perhaps vimentin regulates the turnover of β4 integrin and plectin. Moreover, an inverse association was observed between vimentin expression and β4 integrin in oral squamous cell carcinoma (OSCC). Collectively, our results show a novel role of vimentin in modulating cell motility by destabilizing β4 integrin-mediated adhesive interactions. Further, vimentin-β4 integrin together may prove to be useful markers for prognostication of human oral cancer.

  2. Motorized RhoGAP myosin IXb (Myo9b) controls cell shape and motility

    PubMed Central

    Hanley, Peter J.; Xu, Yan; Kronlage, Moritz; Grobe, Kay; Schön, Peter; Song, Jian; Sorokin, Lydia; Schwab, Albrecht; Bähler, Martin

    2010-01-01

    Directional motility is a fundamental function of immune cells, which are recruited to sites of pathogen invasion or tissue damage by chemoattractant signals. To move, cells need to generate lamellipodial membrane protrusions at the front and retract the trailing end. These elementary events are initiated by Rho-family GTPases, which cycle between active GTP-bound and inactive GDP-bound states. How the activity of these “molecular switches” is spatially coordinated is only beginning to be understood. Here, we show that myosin IXb (Myo9b), a Rho GTPase-activating protein (RhoGAP) expressed in immune cells, is essential for coordinating the activity of Rho. We generated Myo9b-deficient mice and show that Myo9b−/− macrophages have strikingly defective spreading and polarization. Furthermore, Myo9b−/− macrophages fail to generate lamellipodia in response to a chemoattractant, and migration in a chemotactic gradient is severely impaired. Inhibition of Rho rescues the Myo9b−/− phenotype, but impairs tail retraction. We also found that Myo9b is important in vivo. Chemoattractant-induced monocyte recruitment to the peritoneal cavity is substantially reduced in Myo9b−/− mice. Thus, we identify the “motorized Rho inhibitor” Myo9b as a key molecular component required for spatially coordinated cell shape changes and motility. PMID:20566876

  3. Paxillin controls directional cell motility in response to physical cues

    PubMed Central

    Sero, Julia E.; German, Alexandra E.; Mammoto, Akiko; Ingber, Donald E.

    2012-01-01

    Physical cues from the extracellular environment that influence cell shape and directional migration are transduced into changes in cytoskeletal organization and biochemistry through integrin-based cell adhesions to extracellular matrix (ECM). Paxillin is a focal adhesion (FA) scaffold protein that mediates integrin anchorage to the cytoskeleton, and has been implicated in regulation of FA assembly and cell migration. To determine whether paxillin is involved in coupling mechanical distortion with directional movement, cell shape was physically constrained by culturing cells on square-shaped fibronectin-coated adhesive islands surrounded by non-adhesive barrier regions that were created with a microcontact printing technique. Square-shaped cells preferentially formed FAs and extended lamellipodia from their corner regions when stimulated with PDGF, and loss of paxillin resulted in loss of this polarized response. Selective expression of the N- and C-terminal domains of paxillin produced opposite, but complementary, effects on suppressing or promoting lamellipodia formation in different regions of square cells, which corresponded to directional motility defects in vitro. Paxillin loss or mutation was also shown to affect the formation of circular dorsal ruffles, and this corresponded to changes in cell invasive behavior in 3D. This commentary addresses the implications of these findings in terms of how a multifunctional FA scaffold protein can link physical cues to cell adhesion, protrusion and membrane trafficking so as to control directional migration in 2D and 3D. We also discuss how microengineered ECM islands and in vivo model systems can be used to further elucidate the functions of paxillin in directional migration. PMID:23076140

  4. GP73 N-glycosylation at Asn144 reduces hepatocellular carcinoma cell motility and invasiveness

    PubMed Central

    Jiang, Kai; Li, Wei; Zhang, Qinle; Yan, Guoquan; Guo, Kun; Zhang, Shu; Liu, Yinkun

    2016-01-01

    Golgi Protein 73 (GP73) is a potential liver disease glycobiomarker warranting comprehensive analyses of its glycan structure and glycosylation function. In this study, we used mass spectrometry to identify glycosylation sites and the glycan structure, high-throughput lectin microarray to provide rapid and sensitive profiling of glycoconjugates, and site-directed mutagenesis to clarify the impact of glycans on target glycoproteins in vivo. We identified three GP73 N-glycosylation sites: Asn109, Asn144 and Asn398. We found five glycoforms on Asn144, including biantennary, triantennary and fucosylated glycans. Removal of N-glycans at Asn144 enhanced the motility and invasiveness of hepatocellular carcinoma cells, possibly due to inhibition of cell adhesion related to the changes of cell membrane glycosylation. This study increases our understanding of the functional relevance of GP73 glycosylation and suggests that Asn144-deleted GP73 can influence the progression and metastasis of hepatocellular carcinoma. PMID:26993603

  5. Akt2 knock-down reveals its contribution to human lung cancer cell proliferation, growth, motility, invasion and endothelial cell tube formation.

    PubMed

    Attoub, Samir; Arafat, Kholoud; Hammadi, Nasseredine Kamel; Mester, Jan; Gaben, Anne-Marie

    2015-01-01

    The Akt/PKB serine/threonine protein kinase consists of three isoforms: Akt-1, -2 and -3. Their overexpression has been detected in human cancers, but their roles in cancer progression are unclear. We investigated the impact of specific silencing of Akt1 and Akt2 on human lung cancer cell proliferation, colony growth, motility, and invasion in vitro as well as tumor growth in vivo using human Non-Small Cell Lung Cancer cells LNM35, and on the vascular tube formation using HUVEC cells. Although silencing of Akt1 decreased cellular invasion at least in part via COX-2 inhibition, it had almost no effect on cell motility, proliferation, colony formation, and angiogenesis. Transient as well as stable silencing of Akt2 resulted in a strong inhibition of Rb phosphorylation associated with a decrease in cellular proliferation and colony formation, leading to the inhibition of tumor growth in the xenograft model. Silencing of Akt2 also reduced cellular motility and invasion in vitro, presumably via COX-2 inhibition. Moreover, silencing of Akt2 in the HUVEC cells resulted in the inhibition of their spontaneous angiogenic phenotype. Altogether, these results indicate that Akt2 plays an important role in lung cancer progression and can be a promising target for lung cancer therapy. PMID:26234648

  6. Akt2 knock-down reveals its contribution to human lung cancer cell proliferation, growth, motility, invasion and endothelial cell tube formation

    PubMed Central

    Attoub, Samir; Arafat, Kholoud; Kamel Hammadi, Nasseredine; Mester, Jan; Gaben, Anne-Marie

    2015-01-01

    The Akt/PKB serine/threonine protein kinase consists of three isoforms: Akt-1, −2 and −3. Their overexpression has been detected in human cancers, but their roles in cancer progression are unclear. We investigated the impact of specific silencing of Akt1 and Akt2 on human lung cancer cell proliferation, colony growth, motility, and invasion in vitro as well as tumor growth in vivo using human Non-Small Cell Lung Cancer cells LNM35, and on the vascular tube formation using HUVEC cells. Although silencing of Akt1 decreased cellular invasion at least in part via COX-2 inhibition, it had almost no effect on cell motility, proliferation, colony formation, and angiogenesis. Transient as well as stable silencing of Akt2 resulted in a strong inhibition of Rb phosphorylation associated with a decrease in cellular proliferation and colony formation, leading to the inhibition of tumor growth in the xenograft model. Silencing of Akt2 also reduced cellular motility and invasion in vitro, presumably via COX-2 inhibition. Moreover, silencing of Akt2 in the HUVEC cells resulted in the inhibition of their spontaneous angiogenic phenotype. Altogether, these results indicate that Akt2 plays an important role in lung cancer progression and can be a promising target for lung cancer therapy. PMID:26234648

  7. The ileal brake--inhibition of jejunal motility after ileal fat perfusion in man.

    PubMed Central

    Spiller, R C; Trotman, I F; Higgins, B E; Ghatei, M A; Grimble, G K; Lee, Y C; Bloom, S R; Misiewicz, J J; Silk, D B

    1984-01-01

    The possibility that malabsorbed fat passing through the human ileum exerts an inhibitory feedback control on jejunal motility has been investigated in 24 normal subjects by perfusing the ileum with a fat containing solution designed to produce ileal luminal fat concentrations similar to those in steatorrhoea (30-40 mg/ml). Mean transit times through a 30 cm saline perfused jejunal segment were measured by a dye dilution technique. Thirty minutes after ileal fat perfusion, mean transit times rose markedly to 18.9 +/- 2.5 minutes from a control value of 7.5 +/- 0.9 minutes (n = 5; p less than 0.05). This was associated with an increase in volume of the perfused segment which rose to 175.1 +/- 22.9 ml (control 97.6 +/- 10.3 ml, n = 5; p less than 0.05). Transit times and segmental volumes had returned towards basal values 90 minutes after completing the fat perfusion. Further studies showed that ileal fat perfusion produced a pronounced inhibition of jejunal pressure wave activity, percentage duration of activity falling from a control level of 40.3 +/- 5.0% to 14.9 +/- 2.8% in the hour after ileal perfusion (p less than 0.01). Ileal fat perfusion was associated with marked rises in plasma enteroglucagon and neurotensin, the peak values (218 +/- 37 and 68 +/- 13.1 pmol/l) being comparable with those observed postprandially in coeliac disease. These observations show the existence in man of an inhibitory intestinal control mechanism, whereby ileal fat perfusion inhibits jejunal motility and delays caudal transit of jejunal contents. PMID:6706215

  8. OBJECTIVE ANALYSIS OF SPERM MOTILITY IN THE LAKE STURGEON, ACIPENSER FULVESCENS: ACTIVATION AND INHIBITION CONDITIONS

    EPA Science Inventory

    An objective analysis of the duration of motility of sperm from the lake sturgeon, Acipenser fulvescens, has been performed using computer-assisted sperm motion analysis at 200 frames/s. Motility was measured in both 1993 and 1994. The percentage of activated motile sperm and the...

  9. Asynchrony in the growth and motility responses to environmental changes by individual bacterial cells

    SciTech Connect

    Umehara, Senkei; Hattori, Akihiro; Inoue, Ippei; Yasuda, Kenji . E-mail: yasuda.bmi@tmd.ac.jp

    2007-05-04

    Knowing how individual cells respond to environmental changes helps one understand phenotypic diversity in a bacterial cell population, so we simultaneously monitored the growth and motility of isolated motile Escherichia coli cells over several generations by using a method called on-chip single-cell cultivation. Starved cells quickly stopped growing but remained motile for several hours before gradually becoming immotile. When nutrients were restored the cells soon resumed their growth and proliferation but remained immotile for up to six generations. A flagella visualization assay suggested that deflagellation underlies the observed loss of motility. This set of results demonstrates that single-cell transgenerational study under well-characterized environmental conditions can provide information that will help us understand distinct functions within individual cells.

  10. Influence of Helical Cell Shape on Motility of Helicobacter Pylori

    NASA Astrophysics Data System (ADS)

    Hardcastle, Joseph; Martinez, Laura; Salama, Nina; Bansil, Rama; Boston University Collaboration; University of Washington Collaboration

    2014-03-01

    Bacteria's body shape plays an important role in motility by effecting chemotaxis, swimming mechanisms, and swimming speed. A prime example of this is the bacteria Helicobacter Pylori;whose helical shape has long been believed to provide an advantage in penetrating the viscous mucus layer protecting the stomach lining, its niche environment. To explore this we have performed bacteria tracking experiments of both wild-type bacteria along with mutants, which have a straight rod shape. A wide distribution of speeds was found. This distribution reflects both a result of temporal variation in speed and different shape morphologies in the bacterial population. Our results show that body shape plays less role in a simple fluid. However, in a more viscous solution the helical shape results in increased swimming speeds. In addition, we use experimentally obtained cell shape measurements to model the hydrodynamic influence of cell shape on swimming speed using resistive force theory. The results agree with the experiment, especially when we fold in the temporal distribution. Interestingly, our results suggest distinct wild-type subpopulations with varying number of half helices can lead to different swimming speeds. NSF PHY

  11. Tumor suppressor KAI1 affects integrin {alpha}v{beta}3-mediated ovarian cancer cell adhesion, motility, and proliferation

    SciTech Connect

    Ruseva, Zlatna; Geiger, Pamina Xenia Charlotte; Hutzler, Peter; Kotzsch, Matthias; Luber, Birgit; Schmitt, Manfred; Gross, Eva; Reuning, Ute

    2009-06-10

    The tetraspanin KAI1 had been described as a metastasis suppressor in many different cancer types, a function for which associations of KAI1 with adhesion and signaling receptors of the integrin superfamily likely play a role. In ovarian cancer, integrin {alpha}v{beta}3 correlates with tumor progression and its elevation in vitro provoked enhanced cell adhesion accompanied by significant increases in cell motility and proliferation in the presence of its major ligand vitronectin. In the present study, we characterized integrin {alpha}v{beta}3-mediated tumor biological effects as a function of cellular KAI1 restoration and proved for the first time that KAI1, besides its already known physical crosstalk with {beta}1-integrins, also colocalizes with integrin {alpha}v{beta}3. Functionally, elevated KAI1 levels drastically increased integrin {alpha}v{beta}3/vitronectin-dependent ovarian cancer cell adhesion. Since an intermediate level of cell adhesive strength is required for optimal cell migration, we next studied ovarian cancer cell motility as a function of KAI1 restoration. By time lapse video microscopy, we found impaired integrin {alpha}v{beta}3/vitronectin-mediated cell migration most probably due to strongly enhanced cellular immobilization onto the adhesion-supporting matrix. Moreover, KAI1 reexpression significantly diminished cell proliferation. These data strongly indicate that KAI1 may suppress ovarian cancer progression by inhibiting integrin {alpha}v{beta}3/vitronectin-provoked tumor cell motility and proliferation as important hallmarks of the oncogenic process.

  12. 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.

  13. Platelet endothelial cell adhesion molecule-1 modulates endothelial cell motility through the small G-protein Rho.

    PubMed

    Gratzinger, Dita; Canosa, Sandra; Engelhardt, Britta; Madri, Joseph A

    2003-08-01

    Platelet endothelial cell adhesion molecule-1 (PECAM-1), an immunoglobulin family vascular adhesion molecule, is involved in endothelial cell migration and angiogenesis (1, 2). We found that endothelial cells lacking PECAM-1 exhibit increased single cell motility and extension formation but poor wound healing migration, reminiscent of cells in which Rho activity has been suppressed by overexpressing a GTPase-activating protein (3). The ability of PECAM-1 to restore wound healing migration to PECAM-1-deficient cells was independent of its extracellular domain or signaling via its immunoreceptor tyrosine-based inhibitory motif. PECAM-1-deficient endothelial cells had a selective defect in RhoGTP loading, and inhibition of Rho activity mimicked the PECAM-1-deficient phenotype of increased chemokinetic single cell motility at the expense of coordinated wound healing migration. The wound healing advantage of PECAM-1-positive endothelial cells was not only Rho mediated but pertussis toxin inhibitable, characteristic of migration mediated by heterotrimeric G-protein-linked seven-transmembrane receptor signaling such as signaling in response to the serum sphingolipid sphingosine-1-phosphate (S1P) (4, 5). Indeed, we found that the wound healing defect of PECAM-1 null endothelial cells is minimized in sphingolipid-depleted media; moreover, PECAM-1 null endothelial cells fail to increase their migration in response to S1P. We have also found that PECAM-1 localizes to rafts and that in its absence heterotrimeric G-protein components are differentially recruited to rafts, providing a potential mechanism for PECAM-1-mediated coordination of S1P signaling. PECAM-1 may thus support the effective S1P/RhoGTP signaling required for wound healing endothelial migration by allowing for the spatially directed, coordinated activation of Galpha signaling pathways. PMID:12890700

  14. Galectin-3-induced cell spreading and motility relies on distinct signaling mechanisms compared to fibronectin.

    PubMed

    More, Shyam K; Chiplunkar, Shubhada V; Kalraiya, Rajiv D

    2016-05-01

    Secreted galectin-3 often gets incorporated into extracellular matrix and is utilized by cancer cells for spreading, movement, and metastatic dissemination. Here we investigate molecular mechanisms by which galectin-3 brings about these effects and compare it with fibronectin. Imaging of cells spread on fibronectin showed stress fibers throughout cell body, however, galectin-3-induced formation of parallel actin bundles in the lamellipodial region resulting in unique morphological features. FRAP analysis showed that the actin turnover in the lamellipodial region was much higher in cells spread on galectin-3 as compared to that on fibronectin. Rac1 activation is correlated with lamellipodial organization on both the substrates. Activation of Akt and Rac1, the regulators of actin dynamics, show inverse correlation with each other on both galectin-3 and fibronectin. Activation of Erk however, remained similar. Further, inhibition of activation of Akt and Erk inhibited spreading and motility of cells on galectin-3 but not on fibronectin. The results very comprehensively demonstrate distinct signaling pathways that regulate microfilament organization, lamellipodial structures, spreading, and movement of cells plated on galectin-3 as opposed to fibronectin.

  15. A role for cortactin in Listeria monocytogenes invasion of NIH 3T3 cells, but not in its intracellular motility.

    PubMed

    Barroso, Consuelo; Rodenbusch, Stacia E; Welch, Matthew D; Drubin, David G

    2006-04-01

    Cortactin is an F-actin binding protein that binds to the Arp2/3 complex, stimulates its actin nucleation activity, and inhibits actin filament debranching. Using RNA interference directed against cortactin, we explored the importance of cortactin for several processes involving dynamic actin assembly. Silencing cortactin expression was efficiently achieved in HeLa and NIH 3T3 cells, with less than 5% of cortactin expression in siRNA-treated cells. Surprisingly, endocytosis in HeLa and NIH 3T3 cells, and cell migration rates, were not altered by RNAi-mediated cortactin silencing. Listeria utilizes actin-based motility to move within and spread among mammalian host cells; its actin-clouds and tails recruit cortactin. We explored the role of cortactin during the Listeria life cycle in cortactin "knockdown" NIH 3T3 cells. Interestingly, cortactin siRNA-treated cells showed a significant reduction in the efficiency of the bacteria invasion in NIH 3T3 cells. However, cortactin depletion did not interfere with assembly of Listeria actin clouds or actin tails, or Listeria intracellular motility or speed. Therefore, our findings suggest that cortactin plays a role in Listeria internalization, but not in the formation of actin clouds and tails, or in bacteria intracellular motility.

  16. Dystroglycan is involved in laminin-1-stimulated motility of Müller glial cells: combined velocity and directionality analysis.

    PubMed

    Méhes, Elöd; Czirók, András; Hegedüs, Balázs; Szabó, Bálint; Vicsek, Tamás; Satz, Jakob; Campbell, Kevin; Jancsik, Veronika

    2005-03-01

    We investigate the role of dystroglycan, a major laminin-1 receptor and central member of the dystrophin-glycoprotein complex, in the laminin-1 induced motility of cultured Muller glial cells. Binding of laminin-1 to dystroglycan was prevented by IIH6, a function-blocking monoclonal antibody against alpha-dystroglycan. As an alternative means of inhibition, we used heparin to mask the dystroglycan binding site of the laminin-1, known to overlap with heparin binding sites. Cell motility was characterized in a two-dimensional motility assay based on computer-controlled videomicroscopy and statistical analysis of cellular trajectories. We obtained data on both the cell velocity and the diffusion index, a measure of direction-changing frequency. Both means of inhibition of dystroglycan function led to a significant decrease in the ability of laminin-1 to stimulate cell migration. At the same time, dystroglycan function does not appear to be involved in laminin-1-dependent increase in process dynamism and direction-changing activity.

  17. A model for cell motility on soft bio-adhesive substrates.

    PubMed

    Sarvestani, Alireza S

    2011-02-24

    Mechanical stiffness of bio-adhesive substrates has been recognized as a major regulator of cell motility. We present a simple physical model to study the crawling locomotion of a contractile cell on a soft elastic substrate. The mechanism of rigidity sensing is accounted for using Schwarz's two-spring model Schwarz et al. (2006). The predicted dependency between the speed of motility and substrate stiffness is qualitatively consistent with experimental observations. The model demonstrates that the rigidity dependent motility of cells is rooted in the regulation of actomyosin contractile forces by substrate deformation at each anchorage point. On stiffer substrates, the traction forces required for cell translocation acquire larger magnitude but show weaker asymmetry which leads to slower cell motility. On very soft substrates, the model predicts a biphasic relationship between the substrate rigidity and the speed of locomotion, over a narrow stiffness range, which has been observed experimentally for some cell types. PMID:21106198

  18. Fibroblastic reticular cell-derived lysophosphatidic acid regulates confined intranodal T-cell motility

    PubMed Central

    Takeda, Akira; Kobayashi, Daichi; Aoi, Keita; Sasaki, Naoko; Sugiura, Yuki; Igarashi, Hidemitsu; Tohya, Kazuo; Inoue, Asuka; Hata, Erina; Akahoshi, Noriyuki; Hayasaka, Haruko; Kikuta, Junichi; Scandella, Elke; Ludewig, Burkhard; Ishii, Satoshi; Aoki, Junken; Suematsu, Makoto; Ishii, Masaru; Takeda, Kiyoshi; Jalkanen, Sirpa; Miyasaka, Masayuki; Umemoto, Eiji

    2016-01-01

    Lymph nodes (LNs) are highly confined environments with a cell-dense three-dimensional meshwork, in which lymphocyte migration is regulated by intracellular contractile proteins. However, the molecular cues directing intranodal cell migration remain poorly characterized. Here we demonstrate that lysophosphatidic acid (LPA) produced by LN fibroblastic reticular cells (FRCs) acts locally to LPA2 to induce T-cell motility. In vivo, either specific ablation of LPA-producing ectoenzyme autotaxin in FRCs or LPA2 deficiency in T cells markedly decreased intranodal T cell motility, and FRC-derived LPA critically affected the LPA2-dependent T-cell motility. In vitro, LPA activated the small GTPase RhoA in T cells and limited T-cell adhesion to the underlying substrate via LPA2. The LPA-LPA2 axis also enhanced T-cell migration through narrow pores in a three-dimensional environment, in a ROCK-myosin II-dependent manner. These results strongly suggest that FRC-derived LPA serves as a cell-extrinsic factor that optimizes T-cell movement through the densely packed LN reticular network. DOI: http://dx.doi.org/10.7554/eLife.10561.001 PMID:26830463

  19. Theory of deformable substrates for cell motility studies.

    PubMed Central

    Peterson, M A

    1996-01-01

    Linear theory is used to relate the tractions F applied by a cell to the resulting deformation of fluid, viscoelastic, or solid substrates. The theory is used to fit data in which the motion of a fluid surface in the neighborhood of a motile keratocyte is visualized with the aid of embedded beads. The data are best fit by modeling the surface layer as a two-dimensional, nearly incompressible fluid. The data favor this model over another plausible model, the planar free boundary of a three-dimensional fluid. In the resulting diagrams for the distribution of F, it is found that both curl F and div F are concentrated in the lateral extrema of the lamellipodium. In a second investigation, a nonlinear theory of weak wrinkles in a solid substrate is proposed. The in-plane stress tensor plays the role of a metric. Compression wrinkles are found in regions where this metric is negative definite. Tension wrinkles arise, in linear approximation, at points on the boundary between positive definite and indefinite regions, and are conjectured to be stabilized by nonlinear effects. Data for the wrinkles that would be produced by keratocyte traction are computed, and these agree qualitatively with observed keratocyte wrinkles. Images FIGURE 7 PMID:8842205

  20. Tiam1/Rac1 signals contribute to the proliferation and chemoresistance, but not motility, of chronic lymphocytic leukemia cells.

    PubMed

    Hofbauer, Sebastian W; Krenn, Peter W; Ganghammer, Sylvia; Asslaber, Daniela; Pichler, Ulrike; Oberascher, Karin; Henschler, Reinhard; Wallner, Michael; Kerschbaum, Hubert; Greil, Richard; Hartmann, Tanja N

    2014-04-01

    Signals from the tumor microenvironment promote the migration, survival, and proliferation of chronic lymphocytic leukemia (CLL) cells. Rho GTPases control various signaling pathways downstream of microenvironmental cues. Here, we analyze the function of Rac1 in the motility and proliferation of CLL cells. We found decreased transcription of the Rac guanine nucleotide exchange factors Tiam1 and Vav1 in unstimulated peripheral blood CLL cells with almost complete loss of Tiam1 but increased transcription of the potential Rac antagonist RhoH. Consistently, stimulation of CLL cells with the chemokine CXCL12 induced RhoA but not Rac1 activation, whereas chemokine-induced CLL cell motility was Rac1-independent. Coculture of CLL cells with activated T cells induced their activation and subsequent proliferation. Here, Tiam1 expression was induced in the malignant cells in line with increased Ki-67 and c-Myc expression. Rac1 or Tiam1 knockdown using siRNA or treatment with the Tiam1/Rac inhibitor NSC-23766 attenuated c-Myc transcription. Furthermore, treatment of CLL cells with NSC-23766 reduced their proliferation. Rac inhibition also antagonized the chemoresistance of activated CLL cells toward fludarabine. Collectively, our data suggest a dynamic regulation of Rac1 function in the CLL microenvironment. Rac inhibition could be of clinical use by selectively interfering with CLL cell proliferation and chemoresistance.

  1. 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.

  2. Effects of cochlear loading on the motility of active outer hair cells

    PubMed Central

    Ó Maoiléidigh, Dáibhid; Hudspeth, A. J.

    2013-01-01

    Outer hair cells (OHCs) power the amplification of sound-induced vibrations in the mammalian inner ear through an active process that involves hair-bundle motility and somatic motility. It is unclear, though, how either mechanism can be effective at high frequencies, especially when OHCs are mechanically loaded by other structures in the cochlea. We address this issue by developing a model of an active OHC on the basis of observations from isolated cells, then we use the model to predict the response of an active OHC in the intact cochlea. We find that active hair-bundle motility amplifies the receptor potential that drives somatic motility. Inertial loading of a hair bundle by the tectorial membrane reduces the bundle’s reactive load, allowing the OHC’s active motility to influence the motion of the cochlear partition. The system exhibits enhanced sensitivity and tuning only when it operates near a dynamical instability, a Hopf bifurcation. This analysis clarifies the roles of cochlear structures and shows how the two mechanisms of motility function synergistically to create the cochlear amplifier. The results suggest that somatic motility evolved to enhance a preexisting amplifier based on active hair-bundle motility, thus allowing mammals to hear high-frequency sounds. PMID:23509256

  3. A nanoscale characterization of the interaction of a novel alginate oligomer with the cell surface and motility of Pseudomonas aeruginosa.

    PubMed

    Powell, Lydia C; Pritchard, Manon F; Emanuel, Charlotte; Onsøyen, Edvar; Rye, Philip D; Wright, Chris J; Hill, Katja E; Thomas, David W

    2014-03-01

    Pseudomonas aeruginosa (PA) biofilm-associated infections are a common cause of morbidity in chronic respiratory disease and represent a therapeutic challenge. Recently, the ability of a novel alginate oligomer (OligoG) to potentiate the effect of antibiotics against gram-negative, multi-drug-resistant bacteria and inhibit biofilm formation in vitro has been described. Interaction of OligoG with the cell surface of PA was characterized at the nanoscale using atomic force microscopy (AFM), zeta potential measurement (surface charge), and sizing measurements (dynamic light scattering). The ability of OligoG to modify motility was studied in motility assays. AFM demonstrated binding of OligoG to the bacterial cell surface, which was irreversible after exposure to hydrodynamic shear (5,500 × g). Zeta potential analysis (pH 5-9; 0.1-0.001 M NaCl) demonstrated that binding was associated with marked changes in the bacterial surface charge (-30.9 ± 0.8 to -47.0 ± 2.3 mV; 0.01 M NaCl [pH 5]; P < 0.001). Sizing analysis demonstrated that alteration of surface charge was associated with cell aggregation with a 2- to 3-fold increase in mean particle size at OligoG concentrations greater than 2% (914 ± 284 to 2599 ± 472 nm; 0.01 M NaCl [pH 5]; P < 0.001). These changes were associated with marked dose-dependent inhibition in bacterial swarming motility in PA and Burkholderia spp. The ability of OligoG to bind to a bacterial surface, modulate surface charge, induce microbial aggregation, and inhibit motility represents important direct mechanisms by which antibiotic potentiation and biofilm disruption is affected. These results highlight the value of combining multiple nanoscale technologies to further our understanding of the mechanisms of action of novel antibacterial therapies.

  4. Simulated Hypergravity Alters Vascular Smooth Muscle Cell Proliferation and Motility

    NASA Technical Reports Server (NTRS)

    Hunt, Shameka; Bettis, Barika; Harris-Hooker, Sandra; Sanford, Gary L.

    1997-01-01

    The cellular effects of gravity are poorly understood due to its constancy and nonavailability of altered gravitational models. Such an understanding is crucial for prolonged space flights. In these studies, we assessed the influence of centrifugation at 6G (HGrav) on vascular smooth muscle (SMC) mobility and proliferation. Cells were: (a) plated at low density and subjected to HGrav for 24-72 hr for proliferation studies, or (b) grown to confluency, subjected to HGrav, mechanically denuded and monitored for cell movement into the denuded area. Controls were maintained under normogravity. SMC showed a 50% inhibition of growth under HGrav and 10% serum; HGrav and low serum resulted in greater growth inhibition. The rate of movement of SMC into the denuded area was 2-3-fold higher under HGrav in low serum compared to controls, but similar in 10% serum. These studies show that HGrav has significant effects on SMC growth and mobility, which are dependent on serum levels.

  5. Daclatasvir inhibits hepatitis C virus NS5A motility and hyper-accumulation of phosphoinositides

    PubMed Central

    Chukkapalli, Vineela; Berger, Kristi L.; Kelly, Sean M.; Thomas, Meryl; Deiters, Alexander; Randall, Glenn

    2014-01-01

    Combinations of direct-acting antivirals (DAAs) against the hepatitis C virus (HCV) have the potential to revolutionize the HCV therapeutic regime. An integral component of DAA combination therapies are HCV NS5A inhibitors. It has previously been proposed that NS5A DAAs inhibit two functions of NS5A: RNA replication and virion assembly. In this study, we characterize the impact of a prototype NS5A DAA, daclatasvir (DCV), on HCV replication compartment formation. DCV impaired HCV replicase localization and NS5A motility. In order to characterize the mechanism behind altered HCV replicase localization, we examined the impact of DCV on the interaction of NS5A with its essential cellular cofactor, phosphatidylinositol-4-kinase III α (PI4KA). We observed that DCV does not inhibit PI4KA directly, nor does it impair early events of the NS5A-PI4KA interaction that can occur when NS5A is expressed alone. NS5A functions that are unaffected by DCV include PI4KA binding, as determined by co-immunoprecipitation, and a basal accumulation of the PI4KA product, PI4P. However, DCV impairs late steps in PI4KA activation that requires NS5A expressed in the context of the HCV polyprotein. These NS5A functions include hyper-stimulation of PI4P levels and appropriate replication compartment formation. The data are most consistent with a model wherein DCV inhibits conformational changes in the NS5A protein or protein complex formations that occur in the context of HCV polyprotein expression and stimulate PI4P hyper-accumulation and replication compartment formation. PMID:25546252

  6. URI promotes gastric cancer cell motility, survival, and resistance to adriamycin in vitro

    PubMed Central

    Hu, Xiaoxia; Zhang, Fei; Luo, Dongwei; Li, Na; Wang, Qian; Xu, Zhonghai; Bian, Huiqin; Liang, Yuting; Lu, Yaojuan; Zheng, Qiping; Gu, Junxia

    2016-01-01

    Unconventional prefoldin RPB5 interactor (URI), a RNA polymerase II Subunit 5-Interacting protein, is known to participate in the regulation of nutrient-sensitive mTOR-dependent transcription programs. Multiple studies have recently demonstrated that URI functions as an oncoprotein, possibly through the mTOR pathway, and regulates tumor cell motility, invasion, and metastasis. However, whether and how URI plays a role in gastric oncogenesis has not been elucidated. Due to drug resistance, recurrence and metastasis, the prognosis of gastric cancer remains poor. This study aims to explore the effects of URI on gastric cancer cells by focusing on their migratory ability and resistance to adriamycin. URI was over-expressed or knocked-down in MGC-803 and HGC-27 gastric cancer cells using URI plasmid or siRNA transfection approach. The cell viability, apoptosis, and migration ability were then examined by the CCK-8 assay, flow cytometer Annexin V/PI staining, and the Transwell cell migration assay respectively. The protein levels of apoptosis and EMT related genes were detected by western blot. The results showed that overexpression of URI promoted while knock-down of URI inhibited gastric cancer cell proliferation. URI overexpression resulted in increased Bcl-2 expression but decreased levels of Bax, cleaved PARP-1 and cleaved caspase-3. Conversely, cells treated with URI siRNA showed increased adriamycin induced apoptosis, along with reduced Bcl-2, but increased Bax, cleaved PARP-1 and cleaved caspase-3 expression. We have also shown that overexpression of URI enhanced cancer cell proliferation and migration with higher levels of Snail and Vimentin, whereas knockdown of URI in MGC-803 and HGC-27 cells inhibited proliferation and migration with decreased Snail and Vimentin expression. Together, our results support that URI promotes cell survival and mobility and acts as a chemotherapeutics resistant protein in MGC-803 and HGC-27 cells. URI might be a potential biomarker

  7. Cell division resets polarity and motility for the bacterium Myxococcus xanthus.

    PubMed

    Harvey, Cameron W; Madukoma, Chinedu S; Mahserejian, Shant; Alber, Mark S; Shrout, Joshua D

    2014-11-01

    Links between cell division and other cellular processes are poorly understood. It is difficult to simultaneously examine division and function in most cell types. Most of the research probing aspects of cell division has experimented with stationary or immobilized cells or distinctly asymmetrical cells. Here we took an alternative approach by examining cell division events within motile groups of cells growing on solid medium by time-lapse microscopy. A total of 558 cell divisions were identified among approximately 12,000 cells. We found an interconnection of division, motility, and polarity in the bacterium Myxococcus xanthus. For every division event, motile cells stop moving to divide. Progeny cells of binary fission subsequently move in opposing directions. This behavior involves M. xanthus Frz proteins that regulate M. xanthus motility reversals but is independent of type IV pilus "S motility." The inheritance of opposing polarity is correlated with the distribution of the G protein RomR within these dividing cells. The constriction at the point of division limits the intracellular distribution of RomR. Thus, the asymmetric distribution of RomR at the parent cell poles becomes mirrored at new poles initiated at the site of division. PMID:25157084

  8. Effects of zinc oxide nanoparticles on Kupffer cell phagosomal motility, bacterial clearance, and liver function

    PubMed Central

    Watson, Christa Y; Molina, Ramon M; Louzada, Andressa; Murdaugh, Kimberly M; Donaghey, Thomas C; Brain, Joseph D

    2015-01-01

    Background Zinc oxide engineered nanoparticles (ZnO ENPs) have potential as nanomedicines due to their inherent properties. Studies have described their pulmonary impact, but less is known about the consequences of ZnO ENP interactions with the liver. This study was designed to describe the effects of ZnO ENPs on the liver and Kupffer cells after intravenous (IV) administration. Materials and methods First, pharmacokinetic studies were conducted to determine the tissue distribution of neutron-activated 65ZnO ENPs post-IV injection in Wistar Han rats. Then, a noninvasive in vivo method to assess Kupffer cell phagosomal motility was employed using ferromagnetic iron particles and magnetometry. We also examined whether prior IV injection of ZnO ENPs altered Kupffer cell bactericidal activity on circulating Pseudomonas aeruginosa. Serum and liver tissues were collected to assess liver-injury biomarkers and histological changes, respectively. Results We found that the liver was the major site of initial uptake of 65ZnO ENPs. There was a time-dependent decrease in tissue levels of 65Zn in all organs examined, refecting particle dissolution. In vivo magnetometry showed a time-dependent and transient reduction in Kupffer cell phagosomal motility. Animals challenged with P. aeruginosa 24 hours post-ZnO ENP injection showed an initial (30 minutes) delay in vascular bacterial clearance. However, by 4 hours, IV-injected bacteria were cleared from the blood, liver, spleen, lungs, and kidneys. Seven days post-ZnO ENP injection, creatine phosphokinase and aspartate aminotransferase levels in serum were significantly increased. Histological evidence of hepatocyte damage and marginated neutrophils were observed in the liver. Conclusion Administration of ZnO ENPs transiently inhibited Kupffer cell phagosomal motility and later induced hepatocyte injury, but did not alter bacterial clearance from the blood or killing in the liver, spleen, lungs, or kidneys. Our data show that

  9. Altering the motility of Trypanosoma cruzi with rabbit polyclonal anti-peptide antibodies reduces infection to susceptible mammalian cells.

    PubMed

    Finkelsztein, Eli J; Diaz-Soto, Juan C; Vargas-Zambrano, Juan C; Suesca, Elizabeth; Guzmán, Fanny; López, Manuel C; Thomas, M Carmen; Forero-Shelton, Manu; Cuellar, Adriana; Puerta, Concepción J; González, John M

    2015-03-01

    Trypanosoma cruzi's trypomastigotes are highly active and their incessant motility seems to be important for mammalian host cell infection. The kinetoplastid membrane protein-11 (KMP-11) is a protein expressed in all parasite stages, which induces a cellular and humoral immune response in the infected host, and is hypothesized to participate in the parasite's motility. An N-terminal peptide from KMP-11, termed K1 or TcTLE, induced polyclonal antibodies that inhibit parasitic invasion of Vero cells. The goal of this study was to evaluate the motility and infectivity of T. cruzi when exposed to polyclonal anti-TcTLE antibodies. Rabbits were immunized with TcTLE peptide along with FIS peptide as an immunomodulator. ELISA assay results showed that post-immunization sera contained high titers of polyclonal anti-TcTLE antibodies, which were also reactive against the native KMP-11 protein and live parasites as detected by immunofluorescence and flow cytometry assays. Trypomastigotes of T. cruzi were incubated with pre- or post-immunization sera, and infectivity to human astrocytes was assessed by Giemsa staining/light microscope and flow cytometry using carboxyfluorescein diacetate succinimidyl ester (CFSE) labeled parasites. T. cruzi infection in astrocytes decreased approximately by 30% upon incubation with post-immunization sera compared with pre-immunization sera. Furthermore, trypomastigotes were recorded by video microscopy and the parasite's flagellar speed was calculated by tracking the flagella. Trypomastigotes exposed to post-immunization sera had qualitative alterations in motility and significantly slower flagella (45.5 µm/s), compared with those exposed to pre-immunization sera (69.2 µm/s). In summary, polyclonal anti-TcTLE serum significantly reduced the parasite's flagellar speed and cell infectivity. These findings support that KMP-11 could be important for parasite motility, and that by targeting its N-terminal peptide infectivity can be reduced.

  10. Swimming motility plays a key role in the stochastic dynamics of cell clumping

    NASA Astrophysics Data System (ADS)

    Qi, Xianghong; Nellas, Ricky B.; Byrn, Matthew W.; Russell, Matthew H.; Bible, Amber N.; Alexandre, Gladys; Shen, Tongye

    2013-04-01

    Dynamic cell-to-cell interactions are a prerequisite to many biological processes, including development and biofilm formation. Flagellum induced motility has been shown to modulate the initial cell-cell or cell-surface interaction and to contribute to the emergence of macroscopic patterns. While the role of swimming motility in surface colonization has been analyzed in some detail, a quantitative physical analysis of transient interactions between motile cells is lacking. We examined the Brownian dynamics of swimming cells in a crowded environment using a model of motorized adhesive tandem particles. Focusing on the motility and geometry of an exemplary motile bacterium Azospirillum brasilense, which is capable of transient cell-cell association (clumping), we constructed a physical model with proper parameters for the computer simulation of the clumping dynamics. By modulating mechanical interaction (‘stickiness’) between cells and swimming speed, we investigated how equilibrium and active features affect the clumping dynamics. We found that the modulation of active motion is required for the initial aggregation of cells to occur at a realistic time scale. Slowing down the rotation of flagellar motors (and thus swimming speeds) is correlated to the degree of clumping, which is consistent with the experimental results obtained for A. brasilense.

  11. Alpha actinin-1 regulates cell-matrix adhesion organization in keratinocytes: consequences for skin cell motility.

    PubMed

    Hamill, Kevin J; Hiroyasu, Sho; Colburn, Zachary T; Ventrella, Rosa V; Hopkinson, Susan B; Skalli, Omar; Jones, Jonathan C R

    2015-04-01

    The migration of keratinocytes in wound healing requires coordinated activities of the motility machinery of a cell, the cytoskeleton, and matrix adhesions. In this study, we assessed the role of alpha actinin-1 (ACTN1), one of the two alpha actinin isoforms expressed in keratinocytes, in skin cell migration via a small hairpin RNA-mediated knockdown approach. Keratinocytes deficient in ACTN1 exhibit changes in their actin cytoskeleton organization, a loss in front-rear polarity, and impaired lamellipodial dynamics. They also display aberrant directed motility and move slower compared with their wild-type counterparts. Moreover, they have abnormally arranged matrix adhesion sites. Specifically, the focal adhesions in ACTN1 knockdown keratinocytes are not organized as distinct entities. Rather, focal adhesion proteins are arranged in a circle subjacent to cortical fibers of actin. In the same cells, hemidesmosome proteins arrange in cat paw patterns, more typical of confluent, stationary cells, and β4 integrin dynamics are reduced in knockdown cells compared with control keratinocytes. In summary, our data suggest a mechanism by which ACTN1 determines the motility of keratinocytes by regulating the organization of the actin cytoskeleton, focal adhesion, and hemidesmosome proteins complexes, thereby modulating cell speed, lamellipodial dynamics, and directed migration. PMID:25431851

  12. Increased aerobic glycolysis is important for the motility of activated VSMC and inhibited by indirubin-3′-monoxime

    PubMed Central

    Heiss, Elke H.; Schachner, Daniel; Donati, Maddalena; Grojer, Christoph S.; Dirsch, Verena M.

    2016-01-01

    Increased aerobic glycolysis is a recognized feature of multiple cellular phenotypes and offers a potential point for drug interference, as pursued by anti-tumor agents targeting the Warburg effect. This study aimed at examining the role of aerobic glycolysis for migration of vascular smooth muscle cells (VSMC) and its susceptibility to the small molecule indirubin-3′-monoxime (I3MO). Activation of VSMC with platelet-derived growth factor (PDGF) resulted in migration and increased glycolytic activity which was accompanied by an increased glucose uptake and hexokinase (HK) 2 expression. Inhibition of glycolysis or hexokinase by pharmacological agents or siRNA-mediated knockdown significantly reduced the migratory behavior in VSMC without affecting cell viability or early actin cytoskeleton rearrangement. I3MO, previously recognized as inhibitor of VSMC migration, was able to counteract the PDGF-activated increase in glycolysis and HK2 abundance. Activation of signal transducer and activator of transcription (STAT) 3 could be identified as crucial event in upregulation of HK2 and glycolytic activity in PDGF-stimulated VSMC and as point of interference for I3MO. I3MO did not inhibit hypoxia-inducible factor (HIF)1α-dependent transcription nor influence miRNA 143 levels, other potential regulators of HK2 levels. Overall, we demonstrate that increased aerobic glycolysis is an important factor for the motility of activated VSMC and that the anti-migratory property of I3MO may partly depend on impairment of glycolysis via a compromised STAT3/HK2 signaling axis. PMID:27185663

  13. Cell Shapes and Traction Forces Determine Stress in Motile Confluent Tissue

    NASA Astrophysics Data System (ADS)

    Yang, Xingbo; Bi, Dapeng; Czajkowski, Michael; Manning, Lisa; Marchetti, Cristina

    Collective cell migration is a highly regulated process involved in wound healing, cancer metastasis and morphogenesis. The understanding of the regulatory mechanism requires the study of mechanical interactions among cells that coordinate their active motion. To this end, we develop a method that determines cellular forces and tissue stresses from experimentally accessible cell shapes and traction forces. This approach allows us for the first time to calculate membrane tensions and hydrostatic pressures at a cellular level in collective migrating cell layers out of equilibrium. It helps us understand the mechanical origin of tissue stresses as previous inferred using Traction Force Microscopy (TFM). We test this approach on a new model of motile confluent tissue, which we term Self-propelled Voronoi Model (SPV) that incorporates cell elasticity, Contractility and motility. With the model, we explore the mechanical properties of confluent motile tissue as a function of cell activities and cell shapes in various geometries.

  14. Cyclin-dependent kinase 5 activity controls cell motility and metastatic potential of prostate cancer cells.

    PubMed

    Strock, Christopher J; Park, Jong-In; Nakakura, Eric K; Bova, G Steven; Isaacs, John T; Ball, Douglas W; Nelkin, Barry D

    2006-08-01

    We show here that cyclin-dependent kinase 5 (CDK5), a known regulator of migration in neuronal development, plays an important role in prostate cancer motility and metastasis. P35, an activator of CDK5 that is indicative of its activity, is expressed in a panel of human and rat prostate cancer cell lines, and is also expressed in 87.5% of the human metastatic prostate cancers we examined. Blocking of CDK5 activity with a dominant-negative CDK5 construct, small interfering RNA, or roscovitine resulted in changes in the microtubule cytoskeleton, loss of cellular polarity, and loss of motility. Expression of a dominant-negative CDK5 in the highly metastatic Dunning AT6.3 prostate cancer cell line also greatly impaired invasive capacity. CDK5 activity was important for spontaneous metastasis in vivo; xenografts of AT6.3 cells expressing dominant-negative CDK5 had less than one-fourth the number of lung metastases exhibited by AT6.3 cells expressing the empty vector. These results show that CDK5 activity controls cell motility and metastatic potential in prostate cancer.

  15. Nuclear actin modulates cell motility via transcriptional regulation of adhesive and cytoskeletal genes

    PubMed Central

    Sharili, Amir S.; Kenny, Fiona N.; Vartiainen, Maria K.; Connelly, John T.

    2016-01-01

    The actin cytoskeleton is a classic biomechanical mediator of cell migration. While it is known that actin also shuttles in and out of the nucleus, its functions within this compartment remain poorly understood. In this study, we investigated how nuclear actin regulates keratinocyte gene expression and cell behavior. Gene expression profiling of normal HaCaT keratinocytes compared to HaCaTs over-expressing wild-type β-actin or β-actin tagged with a nuclear localization sequence (NLS-actin), identified multiple adhesive and cytoskeletal genes, such as MYL9, ITGB1, and VCL, which were significantly down-regulated in keratinocytes with high levels of nuclear actin. In addition, genes associated with transcriptional regulation and apoptosis were up-regulated in cells over expressing NLS-actin. Functionally, accumulation of actin in the nucleus altered cytoskeletal and focal adhesion organization and inhibited cell motility. Exclusion of endogenous actin from the nucleus by knocking down Importin 9 reversed this phenotype and enhanced cell migration. Based on these findings, we conclude that the level of actin in the nucleus is a transcriptional regulator for tuning keratinocyte migration. PMID:27650314

  16. Nuclear actin modulates cell motility via transcriptional regulation of adhesive and cytoskeletal genes.

    PubMed

    Sharili, Amir S; Kenny, Fiona N; Vartiainen, Maria K; Connelly, John T

    2016-01-01

    The actin cytoskeleton is a classic biomechanical mediator of cell migration. While it is known that actin also shuttles in and out of the nucleus, its functions within this compartment remain poorly understood. In this study, we investigated how nuclear actin regulates keratinocyte gene expression and cell behavior. Gene expression profiling of normal HaCaT keratinocytes compared to HaCaTs over-expressing wild-type β-actin or β-actin tagged with a nuclear localization sequence (NLS-actin), identified multiple adhesive and cytoskeletal genes, such as MYL9, ITGB1, and VCL, which were significantly down-regulated in keratinocytes with high levels of nuclear actin. In addition, genes associated with transcriptional regulation and apoptosis were up-regulated in cells over expressing NLS-actin. Functionally, accumulation of actin in the nucleus altered cytoskeletal and focal adhesion organization and inhibited cell motility. Exclusion of endogenous actin from the nucleus by knocking down Importin 9 reversed this phenotype and enhanced cell migration. Based on these findings, we conclude that the level of actin in the nucleus is a transcriptional regulator for tuning keratinocyte migration. PMID:27650314

  17. Interaction of Motility, Directional Sensing, and Polarity Modules Recreates the Behaviors of Chemotaxing Cells

    PubMed Central

    Shi, Changji; Huang, Chuan-Hsiang; Devreotes, Peter N.; Iglesias, Pablo A.

    2013-01-01

    Chemotaxis involves the coordinated action of separable but interrelated processes: motility, gradient sensing, and polarization. We have hypothesized that these are mediated by separate modules that account for these processes individually and that, when combined, recreate most of the behaviors of chemotactic cells. Here, we describe a mathematical model where the modules are implemented in terms of reaction-diffusion equations. Migration and the accompanying changes in cellular morphology are demonstrated in simulations using a mechanical model of the cell cortex implemented in the level set framework. The central module is an excitable network that accounts for random migration. The response to combinations of uniform stimuli and gradients is mediated by a local excitation, global inhibition module that biases the direction in which excitability is directed. A polarization module linked to the excitable network through the cytoskeleton allows unstimulated cells to move persistently and, for cells in gradients, to gradually acquire distinct sensitivity between front and back. Finally, by varying the strengths of various feedback loops in the model we obtain cellular behaviors that mirror those of genetically altered cell lines. PMID:23861660

  18. Brain-derived neurotrophic factor regulates cell motility in human colon cancer.

    PubMed

    Huang, Ssu-Ming; Lin, Chingju; Lin, Hsiao-Yun; Chiu, Chien-Ming; Fang, Chia-Wei; Liao, Kuan-Fu; Chen, Dar-Ren; Yeh, Wei-Lan

    2015-06-01

    Brain-derived neurotrophic factor (BDNF) is a potent neurotrophic factor that has been shown to affect cancer cell metastasis and migration. In the present study, we investigated the mechanisms of BDNF-induced cell migration in colon cancer cells. The migratory activities of two colon cancer cell lines, HCT116 and SW480, were found to be increased in the presence of human BDNF. Heme oxygenase-1 (HO)-1 is known to be involved in the development and progression of tumors. However, the molecular mechanisms that underlie HO-1 in the regulation of colon cancer cell migration remain unclear. Expression of HO-1 protein and mRNA increased in response to BDNF stimulation. The BDNF-induced increase in cell migration was antagonized by a HO-1 inhibitor and HO-1 siRNA. Furthermore, the expression of vascular endothelial growth factor (VEGF) also increased in response to BDNF stimulation, as did VEGF mRNA expression and transcriptional activity. The increase in BDNF-induced cancer cell migration was antagonized by a VEGF-neutralizing antibody. Moreover, transfection with HO-1 siRNA effectively reduced the increased VEGF expression induced by BDNF. The BDNF-induced cell migration was regulated by the ERK, p38, and Akt signaling pathways. Furthermore, BDNF-increased HO-1 and VEGF promoter transcriptional activity were inhibited by ERK, p38, and AKT pharmacological inhibitors and dominant-negative mutants in colon cancer cells. These results indicate that BDNF increases the migration of colon cancer cells by regulating VEGF/HO-1 activation through the ERK, p38, and PI3K/Akt signaling pathways. The results of this study may provide a relevant contribution to our understanding of the molecular mechanisms by which BDNF promotes colon cancer cell motility.

  19. Computer-assisted quantification of motile and invasive capabilities of cancer cells.

    PubMed

    Kumar, Karthiga Santhana; Pillong, Max; Kunze, Jens; Burghardt, Isabel; Weller, Michael; Grotzer, Michael A; Schneider, Gisbert; Baumgartner, Martin

    2015-10-21

    High-throughput analysis of cancer cell dissemination and its control by extrinsic and intrinsic cellular factors is hampered by the lack of adequate and efficient analytical tools for quantifying cell motility. Oncology research would greatly benefit from such a methodology that allows to rapidly determine the motile behaviour of cancer cells under different environmental conditions, including inside three-dimensional matrices. We combined automated microscopy imaging of two- and three-dimensional cell cultures with computational image analysis into a single assay platform for studying cell dissemination in high-throughput. We have validated this new approach for medulloblastoma, a metastatic paediatric brain tumour, in combination with the activation of growth factor signalling pathways with established pro-migratory functions. The platform enabled the detection of primary tumour and patient-derived xenograft cell sensitivity to growth factor-dependent motility and dissemination and identified tumour subgroup-specific responses to selected growth factors of excellent diagnostic value.

  20. Computer-assisted quantification of motile and invasive capabilities of cancer cells

    PubMed Central

    Kumar, Karthiga Santhana; Pillong, Max; Kunze, Jens; Burghardt, Isabel; Weller, Michael; Grotzer, Michael A.; Schneider, Gisbert; Baumgartner, Martin

    2015-01-01

    High-throughput analysis of cancer cell dissemination and its control by extrinsic and intrinsic cellular factors is hampered by the lack of adequate and efficient analytical tools for quantifying cell motility. Oncology research would greatly benefit from such a methodology that allows to rapidly determine the motile behaviour of cancer cells under different environmental conditions, including inside three-dimensional matrices. We combined automated microscopy imaging of two- and three-dimensional cell cultures with computational image analysis into a single assay platform for studying cell dissemination in high-throughput. We have validated this new approach for medulloblastoma, a metastatic paediatric brain tumour, in combination with the activation of growth factor signalling pathways with established pro-migratory functions. The platform enabled the detection of primary tumour and patient-derived xenograft cell sensitivity to growth factor-dependent motility and dissemination and identified tumour subgroup-specific responses to selected growth factors of excellent diagnostic value. PMID:26486848

  1. Thrombin induces expression of twist and cell motility via the hypoxia-inducible factor-1α translational pathway in colorectal cancer cells.

    PubMed

    Chang, Li-Hsun; Chen, Chun-Han; Huang, Der-Yi; Pai, Hui-Chen; Pan, Shiow-Lin; Teng, Che-Ming

    2011-04-01

    Deep vein thrombosis associated with advanced cancer is known as Trousseau's syndrome. We hypothesized that thrombin, an activator of protease-activated receptor (PAR)-1 and PAR-4 contributes to tumor metastasis. In this study, we demonstrated that thrombin and the PAR-1 activating peptide (AP) SFLLRN, but not the PAR-4 AP GYPGKF, induced HIF-1α activities, protein expression, and cell motility in colorectal cancer cells, and these actions were significantly inhibited by the PAR-1 antagonist SCH79797. Moreover, thrombin-induced HIF-1α activity and cell motility were blocked by inhibiting important mediators of signaling transduction, including the ERK, PI3K, and mTOR pathways. These results showed that thrombin induced HIF-1α protein expression through PAR-1 and HIF-1α translational de novo protein synthesis. Twist can regulate epithelial-mesenchymal transition (EMT) and increase tumor metastasis. However, we observed that thrombin-induced HIF-1α increased Twist mRNA and its protein level was mediated by the modulation of PAR-1 activation and the HIF-1α translational pathway. In addition, Twist could increase N-cadherin but not E-cadherin to promote tumor metastasis. Overexpression of dominant-negative HIF-1α reversed thrombin-mediated Twist and Twist-induced N-cadherin expression. Moreover, siTwist inhibited Twist-induced N-cadherin and Thrombin-induced cell motility. In conclusion, our study showed that thrombin-induced HIF-1α upregulated Twist at the transcriptional level to enhance cell motility. These findings show that thrombin upregulates Twist via HIF-1α to make tumor cells malignant and also establish a link between the coagulation disorder and cancer metastasis. PMID:20857420

  2. Putrescine importer PlaP contributes to swarming motility and urothelial cell invasion in Proteus mirabilis.

    PubMed

    Kurihara, Shin; Sakai, Yumi; Suzuki, Hideyuki; Muth, Aaron; Phanstiel, Otto; Rather, Philip N

    2013-05-31

    Previously, we reported that the speA gene, encoding arginine decarboxylase, is required for swarming in the urinary tract pathogen Proteus mirabilis. In addition, this previous study suggested that putrescine may act as a cell-to-cell signaling molecule (Sturgill, G., and Rather, P. N. (2004) Mol. Microbiol. 51, 437-446). In this new study, PlaP, a putative putrescine importer, was characterized in P. mirabilis. In a wild-type background, a plaP null mutation resulted in a modest swarming defect and slightly decreased levels of intracellular putrescine. In a P. mirabilis speA mutant with greatly reduced levels of intracellular putrescine, plaP was required for the putrescine-dependent rescue of swarming motility. When a speA/plaP double mutant was grown in the presence of extracellular putrescine, the intracellular levels of putrescine were greatly reduced compared with the speA mutant alone, indicating that PlaP functioned as the primary putrescine importer. In urothelial cell invasion assays, a speA mutant exhibited a 50% reduction in invasion when compared with wild type, and this defect could be restored by putrescine in a PlaP-dependent manner. The putrescine analog Triamide-44 partially inhibited the uptake of putrescine by PlaP and decreased both putrescine stimulated swarming and urothelial cell invasion in a speA mutant.

  3. 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.

  4. Human NK cell development requires CD56-mediated motility and formation of the developmental synapse

    PubMed Central

    Mace, Emily M.; Gunesch, Justin T.; Dixon, Amera; Orange, Jordan S.

    2016-01-01

    While distinct stages of natural killer (NK) cell development have been defined, the molecular interactions that shape human NK cell maturation are poorly understood. Here we define intercellular interactions between developing NK cells and stromal cells which, through contact-dependent mechanisms, promote the generation of mature, functional human NK cells from CD34+ precursors. We show that developing NK cells undergo unique, developmental stage-specific sustained and transient interactions with developmentally supportive stromal cells, and that the relative motility of NK cells increases as they move through development in vitro and ex vivo. These interactions include the formation of a synapse between developing NK cells and stromal cells, which we term the developmental synapse. Finally, we identify a role for CD56 in developmental synapse structure, NK cell motility and NK cell development. Thus, we define the developmental synapse leading to human NK cell functional maturation. PMID:27435370

  5. Is there a role for human pluripotent stem cells in modelling interstitial cells of Cajal and gut motility disorders?

    PubMed

    Meng, Wenbo; Zhou, Jerry; Elliott, Ross; Murphy, Patricia; Ho, Vincent; O'Connor, Michael

    2015-01-01

    Gastrointestinal motility disorders affect millions of people worldwide, resulting in significant morbidity and mortality. Current treatments for these disorders are inadequate and often provide little to no relief for patients. As a result, gastrointestinal motility disorders produce substantial long-term social and economic burdens in both developed and developing countries. These limited treatment options arise largely from our relatively poor understanding of the molecular etiology for the majority of gastrointestinal motility disorders. In turn, this is due to our limited access to normal or diseased human gut tissue for use in research. In particular while the interstitial cells of Cajal (ICC) are known to be important for gastrointestinal motility, little is known of how these cells function or how they are involved in disease initiation and progression. The advent of human pluripotent stem cell technology offers an opportunity to generate large amounts of human tissue for both research and clinical applications. The application of this technology to gastrointestinal motility disorders is currently only in its infancy and as yet no studies have described ICC production from human pluripotent cells. By considering the present understanding of the anatomical, cellular and molecular basis of gut motility with particular emphasis on ICC, this review provides a clear framework for the application of human pluripotent stem cell technology to answer fundamental questions of ICC involvement in gut motility. PMID:25391378

  6. Microfabricated Systems and Assays for Studying the Cytoskeletal Organization, Micromechanics, and Motility Patterns of Cancerous Cells

    PubMed Central

    Huda, Sabil; Pilans, Didzis; Makurath, Monika; Hermans, Thomas

    2015-01-01

    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 review, we discuss four categories of microfabricated systems: (i) micropatterned substrates for studying of cell motility sub-processes (for example, MT targeting of 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 cells types and chemical gradients to mimic in vivo intravasation/extravasation steps of metastasis. 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. PMID:26900544

  7. 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

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

    PubMed

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

    2013-04-15

    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 34hpf and again between 64 and 74hpf. 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

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

    PubMed

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

    2013-04-15

    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 34hpf and again between 64 and 74hpf. 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.

  10. Rapid actions of plasma membrane estrogen receptors regulate motility of mouse embryonic stem cells through a profilin-1/cofilin-1-directed kinase signaling pathway.

    PubMed

    Yun, Seung Pil; Ryu, Jung Min; Kim, Mi Ok; Park, Jae Hong; Han, Ho Jae

    2012-08-01

    Long-term estrogen actions are vital for driving cell growth, but more recent evidence suggests that estrogen mediates more rapid cellular effects. However, the function of estradiol-17β (E(2))-BSA in mouse embryonic stem cells has not been reported. Therefore, we examined the role of E(2)-BSA in mouse embryonic stem cell motility and its related signal pathways. E(2)-BSA (10(-8) m) significantly increased motility after 24 h incubation and increased filamentous (F)-actin expression; these effects were inhibited by the estrogen receptor antagonist ICI 182,780, indicating that E(2)-BSA bound membrane estrogen receptors and initiated a signal. E(2)-BSA increased c-Src and focal adhesion kinase (FAK) phosphorylation, which was attenuated by ICI 182,780. The E(2)-BSA-induced increase in epidermal growth factor receptor (EGFR) phosphorylation was inhibited by Src inhibitor PP2. As a downstream signal molecule, E(2)-BSA activated cdc42 and increased formation of a complex with the neural Wiskott-Aldrich syndrome protein (N-WASP)/cdc42/transducer of cdc42-dependent actin assembly-1 (TOCA-1), which was inhibited by FAK small interfering RNA (siRNA) and EGFR inhibitor AG 1478. In addition, E(2)-BSA increased profilin-1 expression and cofilin-1 phosphorylation, which was blocked by cdc42 siRNA. Subsequently, E(2)-BSA induced an increase in F-actin expression, and cell motility was inhibited by each signal pathway-related siRNA molecule or inhibitors but not by cofilin-1 siRNA. A combined treatment of cofilin-1 siRNA and E(2)-BSA increased F-actin expression and cell motility more than that of E(2)-BSA alone. These data demonstrate that E(2)-BSA stimulated motility by interacting with profilin-1/cofilin-1 and F-actin through FAK- and c-Src/EGFR transactivation-dependent N-WASP/cdc42/TOCA-1 complex.

  11. Eukaryotic Translation Initiation Factor 3a (eIF3a) Promotes Cell Proliferation and Motility in Pancreatic Cancer

    PubMed Central

    2016-01-01

    Identifying a target molecule that is crucially involved in pancreatic tumor growth and metastasis is necessary in developing an effective treatment. The study aimed to investigate the role of the eukaryotic translation initiation factor 3a (eIF3a) in the cell proliferation and motility in pancreatic cancer. Our data showed that the expression of eIF3a was upregulated in pancreatic ductal adenocarcinoma as compared with its expression in normal pancreatic tissues. Knockdown of eIF3a by a specific shRNA caused significant decreases in cell proliferation and clonogenic abilities in pancreatic cancer SW1990 and Capan-1 cells. Consistently, the pancreatic cancer cell growth rates were also impaired in xenotransplanted mice. Moreover, wound-healing assay showed that depletion of eIF3a significantly slowed down the wound recovery processes in SW1990 and Capan-1 cells. Transwell migration and invasion assays further showed that cell migration and invasion abilities were significantly inhibited by knockdown of eIF3a in SW1990 and Capan-1 cells. Statistical analysis of eIF3a expression in 140 cases of pancreatic ductal adenocarcinoma samples revealed that eIF3a expression was significantly associated with tumor metastasis and TNM staging. These analyses suggest that eIF3a contributes to cell proliferation and motility in pancreatic ductal adenocarcinoma. PMID:27550487

  12. MT1-MMP dependent repression of the tumor suppressor SPRY4 contributes to MT1-MMP driven melanoma cell motility

    PubMed Central

    Shaverdashvili, Khvaramze; Zhang, Keman; Osman, Iman; Honda, Kord; Jobava, Rauli; Bedogni, Barbara

    2015-01-01

    Metastatic melanoma is the deadliest of all skin cancers. Despite progress in diagnostics and treatment of melanoma, the prognosis for metastatic patients remains poor. We previously showed that Membrane-type 1 Matrix Metalloproteinase (MT1-MMP) is one of the drivers of melanoma metastasis. Classically, MT1-MMP regulates a verity of cellular functions including cell-to-cell interaction and cell-to-matrix communication. Recently, MT1-MMP has been found to also modulate gene expression. To specifically assess MT1-MMP dependent gene regulation in melanoma, microarray gene expression analysis was performed in a melanoma cell line whose metastatic properties depend on the activity of MT1-MMP. We identified the tumor suppressor gene SPRY4 as a new transcriptional target of MT1-MMP that is negatively regulated by the protease. Knockdown of MT1-MMP enhances SPRY4 expression at the mRNA and protein level. SPRY4 expression inversely correlates with that of MT1-MMP in melanoma samples and importantly, correlates with melanoma patient survival. SPRY4 modulates MT1-MMP dependent cell migration such that inhibition of SPRY4 rescues cell migration that has been impaired by MT1-MMP knock down. MT1-MMP decreases SPRY4 in part through an MMP2/RAC1 axis we previously show promotes cell motility downstream of MT1-MMP. These results identify the tumor suppressor SPRY4 as a novel molecular effector of MT1-MMP affecting melanoma cell motility. PMID:26392417

  13. Eukaryotic Translation Initiation Factor 3a (eIF3a) Promotes Cell Proliferation and Motility in Pancreatic Cancer.

    PubMed

    Wang, Shu Qian; Liu, Yu; Yao, Min Ya; Jin, Jing

    2016-10-01

    Identifying a target molecule that is crucially involved in pancreatic tumor growth and metastasis is necessary in developing an effective treatment. The study aimed to investigate the role of the eukaryotic translation initiation factor 3a (eIF3a) in the cell proliferation and motility in pancreatic cancer. Our data showed that the expression of eIF3a was upregulated in pancreatic ductal adenocarcinoma as compared with its expression in normal pancreatic tissues. Knockdown of eIF3a by a specific shRNA caused significant decreases in cell proliferation and clonogenic abilities in pancreatic cancer SW1990 and Capan-1 cells. Consistently, the pancreatic cancer cell growth rates were also impaired in xenotransplanted mice. Moreover, wound-healing assay showed that depletion of eIF3a significantly slowed down the wound recovery processes in SW1990 and Capan-1 cells. Transwell migration and invasion assays further showed that cell migration and invasion abilities were significantly inhibited by knockdown of eIF3a in SW1990 and Capan-1 cells. Statistical analysis of eIF3a expression in 140 cases of pancreatic ductal adenocarcinoma samples revealed that eIF3a expression was significantly associated with tumor metastasis and TNM staging. These analyses suggest that eIF3a contributes to cell proliferation and motility in pancreatic ductal adenocarcinoma. PMID:27550487

  14. Regulation of Motility, Invasion and Metastatic Potential of Squamous Cell Carcinoma by 1,25D3

    PubMed Central

    Ma, Yingyu; Yu, Wei-Dong; Su, Bing; Seshadri, Mukund; Luo, Wei; Trump, Donald L.; Johnson, Candace S.

    2012-01-01

    BACKGROUND 1,25D3, the active metabolite of vitamin D, has been shown to exhibit broad spectrum anti-tumor activity in xenograft animal models. However, its activity against metastatic disease has not been extensively investigated. METHODS Squamous cell carcinoma (SCC) or 1,25D3-resistant variant SCC-DR cells were treated with 1,25D3. Actin organization was examined by immunofluorescence assay. Cell migration was assessed by “wound” healing and chemotactic migration assay. Cell invasion was assessed by Matrigel-based invasion assay and in situ zymography. MMP-2 and MMP-9 expression and secretion was examined by immunoblot analysis and ELISA, respectively. E-cadherin expression was assessed by flow cytometry, immunoblot analysis and immunohistochemistry. Knockdown of E-cadherin was achieved by siRNA. Experimental metastasis mouse model was done by intravenous injection of tumor cells. Lung tumor development was assessed by magnetic resonance imaging, gross observation and histology. RESULTS SCC cellular morphology and actin organization were altered by 10 nM of 1,25D3. 1,25D3 inhibited SCC cell motility and invasion, which was associated with reduced expression and secretion of MMP-2 and MMP-9. 1,25D3 promoted the expression of E-cadherin. These findings were not observed in SCC-DR cells. Knock down of E-cadherin rescued 1,25D3-inhibited cell migration. Intravenous injection of SCC or SCC-DR cells resulted in the establishment of extensive pulmonary lesions in saline-treated C3H mice. Treatment with 1,25D3 resulted in a marked reduction in the formation of lung tumor colonies in animals injected with SCC but not SCC-DR cells. CONCLUSIONS 1,25D3 suppresses SCC cell motility, invasion and metastasis, partially through the promotion of E-cadherin-mediated cell-cell adhesion. PMID:22833444

  15. The Influence of Non Polar and Polar Molecules in Mouse Motile Cells Membranes and Pure Lipid Bilayers

    PubMed Central

    Sierra-Valdez, Francisco J.; Forero-Quintero, Linda S.; Zapata-Morin, Patricio A.; Costas, Miguel; Chavez-Reyes, Arturo; Ruiz-Suárez, Jesús C.

    2013-01-01

    We report an experimental study of mouse sperm motility that shows chief aspects characteristic of neurons: the anesthetic (produced by tetracaine) and excitatory (produced by either caffeine or calcium) effects and their antagonic action. While tetracaine inhibits sperm motility and caffeine has an excitatory action, the combination of these two substances balance the effects, producing a motility quite similar to that of control cells. We also study the effects of these agents (anesthetic and excitatory) on the melting points of pure lipid liposomes constituted by 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and dipalmitoyl phosphatidic acid (DPPA). Tetracaine induces a large fluidization of the membrane, shifting the liposomes melting transition temperature to much lower values. The effect of caffeine is null, but its addition to tetracaine-doped liposomes greatly screen the fluidization effect. A high calcium concentration stiffens pure lipid membranes and strongly reduces the effect of tetracaine. Molecular Dynamics Simulations are performed to further understand our experimental findings at the molecular level. We find a strong correlation between the effect of antagonic molecules that could explain how the mechanical properties suitable for normal cell functioning are affected and recovered. PMID:23565149

  16. Different Motile Behaviors of Human Hematopoietic Stem versus Progenitor Cells at the Osteoblastic Niche

    PubMed Central

    Foster, Katie; Lassailly, François; Anjos-Afonso, Fernando; Currie, Erin; Rouault-Pierre, Kevin; Bonnet, Dominique

    2015-01-01

    Summary Despite advances in our understanding of interactions between mouse hematopoietic stem cells (HSCs) and their niche, little is known about communication between human HSCs and the microenvironment. Using a xenotransplantation model and intravital imaging, we demonstrate that human HSCs display distinct motile behaviors to their hematopoietic progenitor cell (HPC) counterparts, and the same pattern can be found between mouse HSCs and HPCs. HSCs become significantly less motile after transplantation, while progenitor cells remain motile. We show that human HSCs take longer to find their niche than previously expected and suggest that the niche be defined as the position where HSCs stop moving. Intravital imaging is the only technique to determine where in the bone marrow stem cells stop moving, and future analyses should focus on the environment surrounding the HSC at this point. PMID:26455414

  17. Autocrine motility factor promotes HER2 cleavage and signaling in breast cancer cells

    PubMed Central

    Kho, Dhong Hyo; Nangia-Makker, Pratima; Balan, Vitaly; Hogan, Victor; Tait, Larry; Wang, Yi; Raz, Avraham

    2013-01-01

    Trastuzumab (Herceptin®) is an effective targeted therapy in HER2 overexpressing human breast carcinoma. However, many HER2-positive patients initially or eventually become resistant to this treatment, so elucidating mechanisms of trastuzumab resistance that emerge in breast carcinoma cells is clinically important. Here we show that autocrine motility factor (AMF) binds to HER2 and induces cleavage to the ectodomain-deleted and constitutively active form p95HER2. Mechanistic investigations indicated that interaction of AMF with HER2 triggers HER2 phosphorylation and metalloprotease-mediated ectodomain shedding, activating PI3K and MAPK signaling and ablating the ability of trastuzumab to inhibit breast carcinoma cell growth. Further, we found that HER2 expression and AMF secretion were inversely related in breast carcinoma cells. Based on this evidence that AMF may contribute to HER2-mediated breast cancer progression, our findings suggest that AMF-HER2 interaction might be a novel target for therapeutic management of breast cancer patients whose disease is resistant to trastuzumab. PMID:23248119

  18. Downregulation of TFPI in breast cancer cells induces tyrosine phosphorylation signaling and increases metastatic growth by stimulating cell motility

    PubMed Central

    2011-01-01

    Background Increased hemostatic activity is common in many cancer types and often causes additional complications and even death. Circumstantial evidence suggests that tissue factor pathway inhibitor-1 (TFPI) plays a role in cancer development. We recently reported that downregulation of TFPI inhibited apoptosis in a breast cancer cell line. In this study, we investigated the effects of TFPI on self-sustained growth and motility of these cells, and of another invasive breast cancer cell type (MDA-MB-231). Methods Stable cell lines with TFPI (both α and β) and only TFPIβ downregulated were created using RNA interference technology. We investigated the ability of the transduced cells to grow, when seeded at low densities, and to form colonies, along with metastatic characteristics such as adhesion, migration and invasion. Results Downregulation of TFPI was associated with increased self-sustained cell growth. An increase in cell attachment and spreading was observed to collagen type I, together with elevated levels of integrin α2. Downregulation of TFPI also stimulated migration and invasion of cells, and elevated MMP activity was involved in the increased invasion observed. Surprisingly, equivalent results were observed when TFPIβ was downregulated, revealing a novel function of this isoform in cancer metastasis. Conclusions Our results suggest an anti-metastatic effect of TFPI and may provide a novel therapeutic approach in cancer. PMID:21849050

  19. 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

  20. On-Chip Clonal Analysis of Glioma-Stem-Cell Motility and Therapy Resistance.

    PubMed

    Gallego-Perez, Daniel; Chang, Lingqian; Shi, Junfeng; Ma, Junyu; Kim, Sung-Hak; Zhao, Xi; Malkoc, Veysi; Wang, Xinmei; Minata, Mutsuko; Kwak, Kwang J; Wu, Yun; Lafyatis, Gregory P; Lu, Wu; Hansford, Derek J; Nakano, Ichiro; Lee, L James

    2016-09-14

    Enhanced glioma-stem-cell (GSC) motility and therapy resistance are considered to play key roles in tumor cell dissemination and recurrence. As such, a better understanding of the mechanisms by which these cells disseminate and withstand therapy could lead to more efficacious treatments. Here, we introduce a novel micro-/nanotechnology-enabled chip platform for performing live-cell interrogation of patient-derived GSCs with single-clone resolution. On-chip analysis revealed marked intertumoral differences (>10-fold) in single-clone motility profiles between two populations of GSCs, which correlated well with results from tumor-xenograft experiments and gene-expression analyses. Further chip-based examination of the more-aggressive GSC population revealed pronounced interclonal variations in motility capabilities (up to ∼4-fold) as well as gene-expression profiles at the single-cell level. Chip-supported therapy resistance studies with a chemotherapeutic agent (i.e., temozolomide) and an oligo RNA (anti-miR363) revealed a subpopulation of CD44-high GSCs with strong antiapoptotic behavior as well as enhanced motility capabilities. The living-cell-interrogation chip platform described herein enables thorough and large-scale live monitoring of heterogeneous cancer-cell populations with single-cell resolution, which is not achievable by any other existing technology and thus has the potential to provide new insights into the cellular and molecular mechanisms modulating glioma-stem-cell dissemination and therapy resistance.

  1. Pre- and postsynaptic mechanisms in the clonidine- and oxymetazoline-induced inhibition of gastric motility in the rat.

    PubMed

    Zádori, Zoltán S; Shujaa, Nashwan; Fülöp, Katalin; Dunkel, Petra; Gyires, Klára

    2007-10-01

    The inhibitory effect of clonidine (non-selective alpha2-adrenoceptor agonist) and oxymetazoline (alpha2A-adrenoceptor selective agonist) was compared on basal and stimulated gastric motor activity (gastric tone and contractions) using the balloon method in the rat. It was shown that oxymetazoline (0.2-1.7 micromol/kg, i.v.) decreased the basal motility, while clonidine (1.9-3.8 micromol/kg, i.v.) failed to affect it. When motility was stimulated centrally by insulin (5 IU/rat, i.v.), both clonidine (1.9-3.8 micromol/kg, i.v.) and oxymetazoline (0.1-3.4 micromol/kg, i.v.) inhibited the gastric motor activity. However, while the effect of clonidine was antagonized by the non-selective alpha2-adrenoceptor antagonist yohimbine (5 micromol/kg, i.v.) and the alpha2A-adrenoceptor selective antagonist BRL 44408 (3 micromol/kg, i.v.), the effect of oxymetazoline was only partially affected. Prazosin (alpha1- and alpha2B-adrenoceptor antagonist, 0.07-0.28 micromol/kg, i.v.) also failed to reverse the effect of oxymetazoline. Furthermore, when gastric motility was stimulated peripherally by activation of postsynaptic cholinergic muscarinic receptors by the combination of carbachol (0.14 micromol/kg, i.v.) and hexamethonium (37 micromol/kg, i.v.), clonidine (3.8 micromol/kg, i.v.) failed to affect the increased motor activity, however, oxymetazoline (0.8-3.4 micromol/kg, i.v.) exerted a pronounced inhibition. These results suggest that different mechanisms may be involved in the inhibitory effect of clonidine and oxymetazoline; while clonidine reduces the gastric motility by activation of presynaptic alpha2-adrenoceptors, postsynaptic component in the effect of oxymetazoline has also been raised.

  2. Evolution of cell motility in an individual-based model of tumour growth

    PubMed Central

    Gerlee, P.; Anderson, A.R.A.

    2009-01-01

    Tumour invasion is driven by proliferation and importantly migration into the surrounding tissue. Cancer cell motility is also critical in the formation of metastases and is therefore a fundamental issue in cancer research. In this paper we investigate the emergence of cancer cell motility in an evolving tumour population using an individual-based modelling approach. In this model of tumour growth each cell is equipped with a microenvironment response network that determines the behaviour or phenotype of the cell based on the local environment. The response network is modelled using a feed-forward neural network, which is subject to mutations when the cells divide. With this model we have investigated the impact of the micro-environment on the emergence of a motile invasive phenotype. The results show that when a motile phenotype emerges the dynamics of the model are radically changed and we observe faster growing tumours exhibiting diffuse morphologies. Further we observe that the emergence of a motile subclone can occur in a wide range of micro-environmental growth conditions. Iterated simulations showed that in identical growth conditions the evolutionary dynamics either converge to a proliferating or migratory phenotype, which suggests that the introduction of cell motility into the model changes the shape of fitness landscape on which the cancer cell population evolves and that it now contains several local maxima. This could have important implications for cancer treatments which focus on the gene level, as our results show that several distinct genotypes and critically distinct phenotypes can emerge and become dominant in the same micro-environment. PMID:19285513

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

    PubMed Central

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

    2008-01-01

    Background 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. Methods 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. Results 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 ≥ 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. Conclusion 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. PMID:19025616

  4. Metabolic activity of sperm cells: correlation with sperm cell concentration, viability and motility in the rabbit.

    PubMed

    Sabés-Alsina, Maria; Planell, Núria; Gil, Sílvia; Tallo-Parra, Oriol; Maya-Soriano, Maria José; Taberner, Ester; Piles, Miriam; Sabés, Manel; Lopez-Bejar, Manel

    2016-10-01

    The resazurin reduction test (RRT) is a useful technique to assess the metabolic rate of sperm cells. RRT depends on the ability of metabolically active cells to reduce the non-fluorescent dye resazurin to the fluorescent resorufin. The aim of this study was to develop a vital fluorometric method to evaluate metabolic activity of rabbit sperm cells. Twenty-five rabbit males were included in the study. Viability and morphology, motility and metabolic activity were evaluated using an eosin-nigrosin staining, a computer-assisted semen analysis (CASA) and the RRT, respectively. Spearman rank correlation analysis was used to determine the correlation between RRT and semen parameters. After evaluation, a concentration of 10 × 106 sperm cells/ml was selected for further experiments with RRT. No significant correlation was found between the RRT results and the motility parameters. However, after RRT a significant positive correlation between relative fluorescence units and the percentage of alive spermatozoa (r = 0.62; P = 0.001) and a negative one with the percentage of sperm cells with acrosomic abnormalities (r = -0.45; P < 0.05) were detected. The vital assessment of metabolic rate of sperm cells by RRT could provide more information about semen quality than other routine semen analysis, correlating with sperm viability and acrosome status information.

  5. 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.

  6. [Inhibition of menstrual uterine motility with four beta-adrenergic drugs (author's transl)].

    PubMed

    Cifuentes, R; Cobo, E

    1981-01-01

    Effects of the sublingual administration of four beta-adrenoceptor drugs on the uterine motility in 40 normal menstruating women were studied. The drugs and total doses tested were: orciprenaline (40 mg), Partusisten (10 mg), salbutamol (8 mg) and isoxsuprine (40 mg). The uterine and antidiuretic activities were studied before and after administration of each one. All those drugs employed reduced greatly the uterine contractions in all the patients. The cardiovascular side-effects were minimal and well tolerated. It suggested that the adrenergic system has an important role in the control of uterine motility during human menstruation.

  7. Human epididymis protein 4 (HE4) plays a key role in ovarian cancer cell adhesion and motility

    SciTech Connect

    Lu, Renquan; Sun, Xinghui; Xiao, Ran; Zhou, Lei; Gao, Xiang; Guo, Lin

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer We generated stable transduced HE4 overexpression and knockdown cells. Black-Right-Pointing-Pointer HE4 was associated with EOC cell adhesion and motility. Black-Right-Pointing-Pointer HE4 might have some effects on activation of EGFR-MAPK signaling pathway. Black-Right-Pointing-Pointer HE4 play an important role in EOC tumorigenicity. -- Abstract: Human epididymis protein 4 (HE4) is a novel and specific biomarker for epithelial ovarian cancer (EOC). We previously demonstrated that serum HE4 levels were significantly elevated in the majority of EOC patients but not in subjects with benign disease or healthy controls. However, the precise mechanism of HE4 protein function is unknown. In this study, we generated HE4-overexpressing SKOV3 cells and found that stably transduced cells promoted cell adhesion and migration. Knockdown of HE4 expression was achieved by stable transfection of SKOV3 cells with a construct encoding a short hairpin DNA directed against the HE4 gene. Correspondingly, the proliferation and spreading ability of HE4-expressed cells were inhibited by HE4 suppression. Mechanistically, impaired EGFR and Erk1/2 phosphorylation were observed in cells with HE4 knockdown. The phosphorylation was restored when the knockdown cells were cultured in conditioned medium containing HE4. Moreover, in vivo tumorigenicity showed that HE4 suppression markedly inhibited the growth of tumors. This suggests that expression of HE4 is associated with cancer cell adhesion, migration and tumor growth, which can be related to its effects on the EGFR-MAPK signaling pathway. Our results provide evidence of the cellular and molecular mechanisms that may underlie the motility-promoting role of HE4 in EOC progression. The role of HE4 as a target for gene-based therapy might be considered in future studies.

  8. Phosphorylation and interaction of Myopodin by Integrin-Link Kinase Lead to Suppression of Cell Growth and Motility in Prostate Cancer Cells

    PubMed Central

    Yu, Yan-Ping; Luo, Jian-Hua

    2011-01-01

    Myopodin is a tumor suppressor gene that suppresses growth of prostate and urothelial carcinomas. However, the mechanism of myopodin tumor suppressor activity or signaling that leads to activation of myopodin remains unclear. In this report, we showed that the N-terminus of myopodin binds integrin-linked kinase (ILK) both in vivo and in vitro. An ILK interaction motif of 78 amino acids (amino acids 82–157) was identified in the N-terminus region of myopodin. Induction of ILK dependent kinase activity by integrin α7 led to phosphorylation of myopodin both in vivo and in vitro. Knocking down ILK dramatically reduced the inhibition of cell growth and motility mediated by myopodin. A mutant of myopodin lacking the ILK interaction motif is inactive in suppressing the growth and motility of PC3 cells. As a result, this study showed a novel and critical signaling pathway that leads to activation of myopodin. PMID:21643011

  9. Signaling Pathways Used by Ergot Alkaloids to Inhibit Bovine Sperm Motility

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ergot alkaloids exert their toxic or pharmaceutical effects through membrane receptor-mediated activities. This study investigated the signaling pathways involved in the in vitro inhibitory effects of both ergotamine (ET) and dihydroergotamine (DEHT) on bovine sperm motility using specific inhibitor...

  10. Cortical Contractility Triggers a Stochastic Switch to Fast Amoeboid Cell Motility

    PubMed Central

    Ruprecht, Verena; Wieser, Stefan; Callan-Jones, Andrew; Smutny, Michael; Morita, Hitoshi; Sako, Keisuke; Barone, Vanessa; Ritsch-Marte, Monika; Sixt, Michael; Voituriez, Raphaël; Heisenberg, Carl-Philipp

    2015-01-01

    Summary 3D amoeboid cell migration is central to many developmental and disease-related processes such as cancer metastasis. Here, we identify a unique prototypic amoeboid cell migration mode in early zebrafish embryos, termed stable-bleb migration. Stable-bleb cells display an invariant polarized balloon-like shape with exceptional migration speed and persistence. Progenitor cells can be reversibly transformed into stable-bleb cells irrespective of their primary fate and motile characteristics by increasing myosin II activity through biochemical or mechanical stimuli. Using a combination of theory and experiments, we show that, in stable-bleb cells, cortical contractility fluctuations trigger a stochastic switch into amoeboid motility, and a positive feedback between cortical flows and gradients in contractility maintains stable-bleb cell polarization. We further show that rearward cortical flows drive stable-bleb cell migration in various adhesive and non-adhesive environments, unraveling a highly versatile amoeboid migration phenotype. PMID:25679761

  11. Structural insight of a concentration-dependent mechanism by which YdiV inhibits Escherichia coli flagellum biogenesis and motility

    PubMed Central

    Li, Bingqing; Li, Ning; Wang, Feng; Guo, Liming; Huang, Yan; Liu, Xiuhua; Wei, Tiandi; Zhu, Deyu; Liu, Cuilan; Pan, Hongfang; Xu, Sujuan; Wang, Hong-Wei; Gu, Lichuan

    2012-01-01

    YdiV is a negative regulator of cell motility. It interacts with FlhD4C2 complex, a product of flagellar master operon, which works as the transcription activator of all other flagellar operons. Here, we report the crystal structures of YdiV and YdiV2–FlhD2 complex at 1.9 Å and 2.9 Å resolutions, respectively. Interestingly, YdiV formed multiple types of complexes with FlhD4C2. YdiV1–FlhD4C2 and YdiV2–FlhD4C2 still bound to DNA, while YdiV3–FlhD4C2 and YdiV4–FlhD4C2 did not. DNA bound FlhD4C2 through wrapping around the FlhC subunit rather than the FlhD subunit. Structural analysis showed that only two peripheral FlhD subunits were accessible for YdiV binding, forming the YdiV2–FlhD4C2 complex without affecting the integrity of ring-like structure. YdiV2–FlhD2 structure and the negative staining electron microscopy reconstruction of YdiV4–FlhD4C2 suggested that the third and fourth YdiV molecule bound to the FlhD4C2 complex through squeezing into the ring-like structure of FlhD4C2 between the two internal D subunits. Consequently, the ring-like structure opened up, and the complex lost DNA-binding ability. Thus, YdiV inhibits FlhD4C2 only at relatively high concentrations. PMID:23002140

  12. Microtubule-dependent motility and orientation of the cortical endoplasmic reticulum in elongating characean internodal cells.

    PubMed

    Foissner, Ilse; Menzel, Diedrik; Wasteneys, Geoffrey O

    2009-03-01

    Motility of the endoplasmic reticulum (ER) is predominantly microtubule- dependent in animal cells but thought to be entirely actomyosin-dependent in plant cells. Using live cell imaging and transmission electron microscopy to examine ER motility and structural organization in giant internodal cells of characean algae, we discovered that at the onset of cell elongation, the cortical ER situated near the plasma membrane formed a tight meshwork of predominantly transverse ER tubules that frequently coaligned with microtubules. Microtubule depolymerization increased mesh size and decreased the dynamics of the cortical ER. In contrast, perturbing the cortical actin array with cytochalasins did not affect the transverse orientation but decreased mesh size and increased ER dynamics. Our data suggest that myosin-dependent ER motility is confined to the ER strands in the streaming endoplasm, while the more sedate cortical ER uses microtubule-based mechanisms for organization and motility during early stages of cell elongation. We show further that the ER has an inherent, NEM-sensitive dynamics which can be altered via interaction with the cytoskeleton and that tubule formation and fusion events are cytoskeleton-independent.

  13. Silencing of phosphoglucose isomerase/autocrine motility factor decreases U87 human glioblastoma cell migration.

    PubMed

    Li, Yang; Wei, Zhenqing; Dong, Bin; Lian, Zhigang; Xu, Yinghui

    2016-04-01

    Phosphoglucose isomerase/autocrine motility factor (PGI/AMF) is secreted by tumors and influences tumor growth and metastasis. In order to investigate the effects of silencing PGI/AMF on the migration and the sphere forming abilities of human glioblastoma U87 cells, as well as on the side population cells (SPCs), PGI/AMF was silenced using siRNA. Western blot analysis and RT-qPCR were used to assess the expression of PGI/AMF, Akt and SRY (sex determining region Y)-box 2 (SOX2). Wound healing, migration and tumorsphere formation assays were performed to assess invasion and metastatic potential. The proportion of SPCs was determined using Hoechst 33342 dye and flow cytometric analysis. PGI/AMF silencing inhibited the wound healing capacity and migration ability of U87 cells by 52.6 and 80.4%, respectively, compared with the scrambled siRNA (both P<0.001). Silencing of PGI/AMF decreased the proportion of SPCs in the U87 cells by 80.9% (P<0.01). The silencing of PGI/AMF decreased the number and size of tumorspheres by 53.1 and 39.9%, respectively, compared with the scrambled siRNA (both P<0.01). The silencing of PGI/AMF decreased the levels of phosphorylated Akt (-71.9%, P<0.001) compared with the scrambled siRNA, as well as the levels of the stemness marker, SOX2 (-61.7%, P<0.01). Taken together, these findings suggest that PGI/AMF silencing decreases migration, tumorsphere formation as well as the proportion of SPCs in glioblastoma U87 cells. We suggest that the Akt pathway is involved, and our results provide a potential new target for the treatment of glioblastoma.

  14. Potential anthelmintic: D-psicose inhibits motility, growth and reproductive maturity of L1 larvae of Caenorhabditis elegans.

    PubMed

    Sato, Masashi; Kurose, Hiroyuki; Yamasaki, Toru; Izumori, Ken

    2008-04-01

    No anthelmintic sugars have yet been identified. Eight ketohexose stereoisomers (D- and L-forms of psicose, fructose, tagatose and sorbose), along with D-galactose and D-glucose, were examined for potency against L1 stage Caenorhabditis elegans fed Escherichia coli. Of the sugars, D-psicose specifically inhibited the motility, growth and reproductive maturity of the L1 stage. D-Psicose probably interferes with the nematode nutrition. The present results suggest that D-psicose, one of the rare sugars, is a potential anthelmintic.

  15. The dynein cortical anchor Num1 activates dynein motility by relieving Pac1/LIS1-mediated inhibition

    PubMed Central

    Lammers, Lindsay G.

    2015-01-01

    Cortically anchored dynein orients the spindle through interactions with astral microtubules. In budding yeast, dynein is offloaded to Num1 receptors from microtubule plus ends. Rather than walking toward minus ends, dynein remains associated with plus ends due in part to its association with Pac1/LIS1, an inhibitor of dynein motility. The mechanism by which dynein is switched from “off” at the plus ends to “on” at the cell cortex remains unknown. Here, we show that overexpression of the coiled-coil domain of Num1 specifically depletes dynein–dynactin–Pac1/LIS1 complexes from microtubule plus ends and reduces dynein-Pac1/LIS1 colocalization. Depletion of dynein from plus ends requires its microtubule-binding domain, suggesting that motility is required. An enhanced Pac1/LIS1 affinity mutant of dynein or overexpression of Pac1/LIS1 rescues dynein plus end depletion. Live-cell imaging reveals minus end–directed dynein–dynactin motility along microtubules upon overexpression of the coiled-coil domain of Num1, an event that is not observed in wild-type cells. Our findings indicate that dynein activity is directly switched “on” by Num1, which induces Pac1/LIS1 removal. PMID:26483554

  16. The dynein cortical anchor Num1 activates dynein motility by relieving Pac1/LIS1-mediated inhibition.

    PubMed

    Lammers, Lindsay G; Markus, Steven M

    2015-10-26

    Cortically anchored dynein orients the spindle through interactions with astral microtubules. In budding yeast, dynein is offloaded to Num1 receptors from microtubule plus ends. Rather than walking toward minus ends, dynein remains associated with plus ends due in part to its association with Pac1/LIS1, an inhibitor of dynein motility. The mechanism by which dynein is switched from "off" at the plus ends to "on" at the cell cortex remains unknown. Here, we show that overexpression of the coiled-coil domain of Num1 specifically depletes dynein-dynactin-Pac1/LIS1 complexes from microtubule plus ends and reduces dynein-Pac1/LIS1 colocalization. Depletion of dynein from plus ends requires its microtubule-binding domain, suggesting that motility is required. An enhanced Pac1/LIS1 affinity mutant of dynein or overexpression of Pac1/LIS1 rescues dynein plus end depletion. Live-cell imaging reveals minus end-directed dynein-dynactin motility along microtubules upon overexpression of the coiled-coil domain of Num1, an event that is not observed in wild-type cells. Our findings indicate that dynein activity is directly switched "on" by Num1, which induces Pac1/LIS1 removal. PMID:26483554

  17. The dynein cortical anchor Num1 activates dynein motility by relieving Pac1/LIS1-mediated inhibition.

    PubMed

    Lammers, Lindsay G; Markus, Steven M

    2015-10-26

    Cortically anchored dynein orients the spindle through interactions with astral microtubules. In budding yeast, dynein is offloaded to Num1 receptors from microtubule plus ends. Rather than walking toward minus ends, dynein remains associated with plus ends due in part to its association with Pac1/LIS1, an inhibitor of dynein motility. The mechanism by which dynein is switched from "off" at the plus ends to "on" at the cell cortex remains unknown. Here, we show that overexpression of the coiled-coil domain of Num1 specifically depletes dynein-dynactin-Pac1/LIS1 complexes from microtubule plus ends and reduces dynein-Pac1/LIS1 colocalization. Depletion of dynein from plus ends requires its microtubule-binding domain, suggesting that motility is required. An enhanced Pac1/LIS1 affinity mutant of dynein or overexpression of Pac1/LIS1 rescues dynein plus end depletion. Live-cell imaging reveals minus end-directed dynein-dynactin motility along microtubules upon overexpression of the coiled-coil domain of Num1, an event that is not observed in wild-type cells. Our findings indicate that dynein activity is directly switched "on" by Num1, which induces Pac1/LIS1 removal.

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

    SciTech Connect

    Chen, Na; Sato, Daisuke; Saiki, Yuriko; Sunamura, Makoto; Fukushige, Shinichi; Horii, Akira

    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 this 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.

  19. Simulating T cell motility in the lymph node paracortex with a packed lattice geometry

    PubMed Central

    Bogle, Gib; Dunbar, P. Rod

    2009-01-01

    Agent-based simulation modeling of T cell trafficking, activation and proliferation in the lymph node paracortex requires a model for cell motility. Such a model must be able to reproduce the observed random-walk behaviour of T cells, while accommodating large numbers of tightly packed cells, and must be computationally efficient. We report the development of a motility model, based on a 3-D lattice geometry, that meets these objectives. Cells make discrete jumps between neighbouring lattice sites in directions that are randomly determined from specified discrete probability distributions, which are defined by a small number of parameters. It is shown that the main characteristics of the random motion of T cells as typically observed in vivo can be reproduced by suitable specification of model parameters. The model is computationally highly efficient, and provides a suitable engine for a model capable of simulating the full T cell population of the paracortex. PMID:18711399

  20. Intermediate filament Nestin and the cell motility in cancer - a review.

    PubMed

    Shi, A-M; Tao, Z-Q; Wang, X; Wang, Y-Q

    2016-06-01

    The intermediate filaments (Ifs) constitute the cytoskeleton which is a key feature of both prokaryotic and eukaryotic cells. The IFs are expressed throughout life and are involved in the regulation of cell differentiation, homeostasis, ageing and pathogenesis. The IFs not only provide structural integrity to the cell, but they are involved also in a range of cellular functions from organelle trafficking and cell migration to signaling transduction. The IFs are highly dynamic proteins, able to respond and adapt their network rapidly in response to intra- and extra-cellular cues. In cancer, these IFs play a crucial role with regard to cell invasion vial cell motility. The present review article will enlighten information about important IF nestin with regard to its role in cancer cell motility and invasion. PMID:27383310

  1. Wide-ranging effects of eight cytochalasins and latrunculin A and B on intracellular motility and actin filament reorganization in characean internodal cells.

    PubMed

    Foissner, Ilse; Wasteneys, Geoffrey O

    2007-04-01

    Numerous forms of cytochalasins have been identified and, although they share common biological activity, they may differ considerably in potency. We investigated the effects of cytochalasins A, B, C, D, E, H and J and dihydrocytochalasin B in an ideal experimental system for cell motility, the giant internodal cells of the characean alga Nitella pseudoflabellata. Cytochalasins D (60 microM) and H (30 microM) were found to be most suited for fast and reversible inhibition of actin-based motility, while cytochalasins A and E arrested streaming at lower concentrations but irreversibly. We observed no clear correlation between the ability of cytochalasins to inhibit motility and the actual disruption of the subcortical actin bundle tracks on which myosin-dependent motility occurs. Indeed, the actin bundles remained intact at the time of streaming cessation and disassembled only after one to several days' treatment. Even when applied at concentrations lower than that required to inhibit cytoplasmic streaming, all of the cytochalasins induced reorganization of the more labile cortical actin filaments into actin patches, swirling clusters or short rods. Latrunculins A and B arrested streaming only after disrupting the subcortical actin bundles, a process requiring relatively high concentrations (200 microM) and very long treatment periods of >1 d. Latrunculins, however, worked synergistically with cytochalasins. A 1 h treatment with 15 nM latrunculin A and 4 microM cytochalasin D induced reversible fragmentation of subcortical actin bundles and arrested cytoplasmic streaming. Our findings provide insights into the mechanisms by which cytochalasins and latrunculins interfere with characean actin to inhibit motility.

  2. 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.

  3. Socs36E attenuates STAT signaling to optimize motile cell specification in the Drosophila ovary.

    PubMed

    Monahan, Amanda J; Starz-Gaiano, Michelle

    2013-07-15

    The Janus kinase/Signal transducers and activators of transcription (JAK/STAT) pathway determines cell fates by regulating gene expression. One example is the specification of the motile cells called border cells during Drosophila oogenesis. It has been established that too much or too little STAT activity disrupts follicle cell identity and cell motility, which suggests the signaling must be precisely regulated. Here, we find that Suppressor of cytokine signaling at 36E (Socs36E) is a necessary negative regulator of JAK/STAT signaling during border cell specification. We find when STAT signaling is too low to induce migration in the presumptive border cell population, nearby follicle cells uncharacteristically become invasive to enable efficient migration of the cluster. We generated a genetic null allele that reveals Socs36E is required in the anterior follicle cells to limit invasive behavior to an optimal number of cells. We further show Socs36E genetically interacts with the required STAT feedback inhibitor apontic (apt) and APT's downstream target, mir-279, and provide evidence that suggests APT directly regulates Socs36E transcriptionally. Our work shows Socs36E plays a critical role in a genetic circuit that establishes a boundary between the motile border cell cluster and its non-invasive epithelial neighbors through STAT attenuation. PMID:23583584

  4. Induction of focal adhesions and motility in Drosophila S2 cells.

    PubMed

    Ribeiro, Susana A; D'Ambrosio, Michael V; Vale, Ronald D

    2014-12-01

    Focal adhesions are dynamic structures that interact with the extracellular matrix on the cell exterior and actin filaments on the cell interior, enabling cells to adhere and crawl along surfaces. We describe a system for inducing the formation of focal adhesions in normally non-ECM-adherent, nonmotile Drosophila S2 cells. These focal adhesions contain the expected molecular markers such as talin, vinculin, and p130Cas, and they require talin for their formation. The S2 cells with induced focal adhesions also display a nonpolarized form of motility on vitronectin-coated substrates. Consistent with findings in mammalian cells, the degree of motility can be tuned by changing the stiffness of the substrate and was increased after the depletion of PAK3, a p21-activated kinase. A subset of nonmotile, nonpolarized cells also exhibited focal adhesions that rapidly assembled and disassembled around the cell perimeter. Such cooperative and dynamic fluctuations of focal adhesions were decreased by RNA interference (RNAi) depletion of myosin II and focal adhesion kinase, suggesting that this behavior requires force and focal adhesion maturation. These results demonstrate that S2 cells, a cell line that is well studied for cytoskeletal dynamics and readily amenable to protein manipulation by RNAi, can be used to study the assembly and dynamics of focal adhesions and mechanosensitive cell motility.

  5. Nanoparticle-functionalized polymer platform for controlling metastatic cancer cell adhesion, shape, and motility.

    PubMed

    Lee, Hyojin; Jang, Yeongseon; Seo, Jinhwa; Nam, Jwa-Min; Char, Kookheon

    2011-07-26

    Controlling and understanding the changes in metastatic cancer cell adhesion, shape, and motility are of paramount importance in cancer research, diagnosis, and treatment. Here, we used gold nanoparticles (AuNPs) as nanotopological structures and protein nanocluster forming substrates. Cell adhesion controlling proteins [in this case, fibronection (Fn) and ephrinB3] were modified to AuNPs, and these particles were then modified to the layer-by-layer (LbL) polymer surface that offers a handle for tuning surface charge and mechanical property of a cell-interfacing substrate. We found that metastatic cancer cell adhesion is affected by nanoparticle density on a surface, and ∼140 particles per 400 μm(2) (∼1.7 μm spacing between AuNPs) is optimal for effective metastatic cell adhesion. It was also shown that the AuNP surface density and protein nanoclustering on a spherical AuNP are controlling factors for the efficient interfacing and signaling of metastatic cancer cells. Importantly, the existence of nanotopological features (AuNPs in this case) is much more critical in inducing more dramatic changes in metastatic cell adhesion, protrusion, polarity, and motility than the presence of a cell adhesion protein, Fn, on the surface. Moreover, cell focal adhesion and motility-related paxillin clusters were heavily formed in cell lamellipodia and filopodia and high expression of phospho-paxillins were observed when the cells were cultured on either an AuNP or Fn-modified AuNP polymer surface. The ephrin signaling that results in the decreased expression of paxillin was found to be more effective when ephrins were modified to the AuNP surface than when ephrinB3 was directly attached to the polymer film. The overall trend for cell motility change is such that a nanoparticle-modified LbL surface induces higher cell motility and the AuNP modification to the LbL surface results in more pronounced change in cell motility than Fn or ephrin modification to the LbL surface.

  6. CRL2(LRR-1) targets a CDK inhibitor for cell cycle control in C. elegans and actin-based motility regulation in human cells.

    PubMed

    Starostina, Natalia G; Simpliciano, Jennifer M; McGuirk, Michael A; Kipreos, Edward T

    2010-11-16

    The Cip/Kip CDK inhibitor (CKI) p21(Cip1/WAF1) has a critical role in the nucleus to limit cell proliferation by inhibiting CDK-cyclin complexes. In contrast, cytoplasmic p21 regulates cell survival and the actin cytoskeleton. These divergent functions for p21 in different cellular compartments suggest the necessity for complex regulation. In this study, we identify the CRL2(LRR-1) ubiquitin ligase as a conserved regulator of Cip/Kip CKIs that promotes the degradation of C. elegans CKI-1 and human p21. The nematode CRL2(LRR-1) complex negatively regulates nuclear CKI-1 levels to ensure G1-phase cell cycle progression in germ cells. In contrast, human CRL2(LRR1) targets cytoplasmic p21, acting as a critical regulator of cell motility that promotes a nonmotile stationary cell state by preventing p21 from inhibiting the Rho/ROCK/LIMK pathway. Inactivation of human CRL2(LRR1) leads to the activation of the actin-depolymerizing protein cofilin, dramatic reorganization of the actin cytoskeleton, and increased cell motility.

  7. Extracellular ATP induces hyperpolarization and motility stimulation of ciliary cells.

    PubMed Central

    Tarasiuk, A; Bar-Shimon, M; Gheber, L; Korngreen, A; Grossman, Y; Priel, Z

    1995-01-01

    Cellular membrane potential and ciliary motility were examined in tissues cultures prepared from frog palate and esophagus epithelia. Addition of micromolar concentrations of extracellular ATP caused membrane hyperpolarization and enhanced the beat frequency. These two effects of ATP were 1) dose dependent, reaching a maximum at 10 microM ATP; 2) dependent on the presence of extracellular Ca2+ or Mg2+; 3) insensitive to inhibitors of voltage-gated calcium channels; 4) abolished after depleting the intracellular Ca2+ stores with thapsigargin; 5) attenuated by quinidine (1 mM), Cs+ (5-20 mM), and replacement of extracellular Na+ by K+; 6) insensitive to charybdotoxin (5-20 nM), TEA (1-20 microM), and apamin (0.1-1 microM); 7) independent of initial membrane potential; and 8) unaffected by amiloride. In addition, extracellular ATP induced an appreciable rise in intracellular Ca2+. Addition of thapsigargin caused an initial enhancement of the ciliary beat frequency and membrane hyperpolarization. These results strongly suggest the involvement of calcium-dependent potassium channels in the response to ATP. The results show that moderate hyperpolarization is closely associated with a sustained enhancement of ciliary beating by extracellular ATP. Images FIGURE 6 PMID:7756536

  8. The Regulation of RhoA at Focal Adhesions by StarD13 is Important for Astrocytoma Cell Motility

    PubMed Central

    Khalil, Bassem D.; Hanna, Samer; Saykali, Bechara A.; El-Sitt, Sally; Nasrallah, Anita; Marston, Daniel; El-Sabban, Marwan; Hahn, Klaus M.; Symons, Marc; El-Sibai, Mirvat

    2015-01-01

    Malignant astrocytomas are highly invasive into adjacent and distant regions of the normal brain. Rho GTPases are small monomeric G proteins that play important roles in cytoskeleton rearrangement, cell motility, and tumor invasion. In the present study, we show that the knock down of StarD13, a GTPase activating protein (GAP) for RhoA and Cdc42, inhibits astrocytoma cell migration through modulating focal adhesion dynamics and cell adhesion. This effect is mediated by the resulting constitutive activation of RhoA and the subsequent indirect inhibition of Rac. Using Total Internal Reflection Fluorescence (TIRF)-based Förster Resonance Energy Transfer (FRET), we show that RhoA activity localizes with focal adhesions at the basal surface of astrocytoma cells. Moreover, the knock down of StarD13 inhibits the cycling of RhoA activation at the rear edge of cells, which makes them defective in retracting their tail. This study highlights the importance of the regulation of RhoA activity in focal adhesions of astrocytoma cells and establishes StarD13 as a GAP playing a major role in this process. PMID:24333506

  9. Regulation of Chlamydomonas flagella and ependymal cell motile cilia by ceramide-mediated translocation of GSK3.

    PubMed

    Kong, Ji Na; Hardin, Kara; Dinkins, Michael; Wang, Guanghu; He, Qian; Mujadzic, Tarik; Zhu, Gu; Bielawski, Jacek; Spassieva, Stefka; Bieberich, Erhard

    2015-12-01

    Cilia are important organelles formed by cell membrane protrusions; however, little is known about their regulation by membrane lipids. We characterize a novel activation mechanism for glycogen synthase kinase-3 (GSK3) by the sphingolipids phytoceramide and ceramide that is critical for ciliogenesis in Chlamydomonas and murine ependymal cells, respectively. We show for the first time that Chlamydomonas expresses serine palmitoyl transferase (SPT), the first enzyme in (phyto)ceramide biosynthesis. Inhibition of SPT in Chlamydomonas by myriocin led to loss of flagella and reduced tubulin acetylation, which was prevented by supplementation with the precursor dihydrosphingosine. Immunocytochemistry showed that (phyto)ceramide was colocalized with phospho-Tyr-216-GSK3 (pYGSK3) at the base and tip of Chlamydomonas flagella and motile cilia in ependymal cells. The (phyto)ceramide distribution was consistent with that of a bifunctional ceramide analogue UV cross-linked and visualized by click-chemistry-mediated fluorescent labeling. Ceramide depletion, by myriocin or neutral sphingomyelinase deficiency (fro/fro mouse), led to GSK3 dephosphorylation and defective flagella and cilia. Motile cilia were rescued and pYGSK3 localization restored by incubation of fro/fro ependymal cells with exogenous C24:1 ceramide, which directly bound to pYGSK3. Our findings suggest that (phyto)ceramide-mediated translocation of pYGSK into flagella and cilia is an evolutionarily conserved mechanism fundamental to the regulation of ciliogenesis.

  10. Regulation of Chlamydomonas flagella and ependymal cell motile cilia by ceramide-mediated translocation of GSK3

    PubMed Central

    Kong, Ji Na; Hardin, Kara; Dinkins, Michael; Wang, Guanghu; He, Qian; Mujadzic, Tarik; Zhu, Gu; Bielawski, Jacek; Spassieva, Stefka; Bieberich, Erhard

    2015-01-01

    Cilia are important organelles formed by cell membrane protrusions; however, little is known about their regulation by membrane lipids. We characterize a novel activation mechanism for glycogen synthase kinase-3 (GSK3) by the sphingolipids phytoceramide and ceramide that is critical for ciliogenesis in Chlamydomonas and murine ependymal cells, respectively. We show for the first time that Chlamydomonas expresses serine palmitoyl transferase (SPT), the first enzyme in (phyto)ceramide biosynthesis. Inhibition of SPT in Chlamydomonas by myriocin led to loss of flagella and reduced tubulin acetylation, which was prevented by supplementation with the precursor dihydrosphingosine. Immunocytochemistry showed that (phyto)ceramide was colocalized with phospho–Tyr-216-GSK3 (pYGSK3) at the base and tip of Chlamydomonas flagella and motile cilia in ependymal cells. The (phyto)ceramide distribution was consistent with that of a bifunctional ceramide analogue UV cross-linked and visualized by click-chemistry–mediated fluorescent labeling. Ceramide depletion, by myriocin or neutral sphingomyelinase deficiency (fro/fro mouse), led to GSK3 dephosphorylation and defective flagella and cilia. Motile cilia were rescued and pYGSK3 localization restored by incubation of fro/fro ependymal cells with exogenous C24:1 ceramide, which directly bound to pYGSK3. Our findings suggest that (phyto)ceramide-mediated translocation of pYGSK into flagella and cilia is an evolutionarily conserved mechanism fundamental to the regulation of ciliogenesis. PMID:26446842

  11. Insulin-like growth factor binding protein-3 interacts with autocrine motility factor/phosphoglucose isomerase (AMF/PGI) and inhibits the AMF/PGI function.

    PubMed

    Mishra, Suresh; Raz, Avrahram; Murphy, Liam J

    2004-04-01

    Autocrine motility factor/phosphoglucose isomerase (AMF/PGI) was identified as a binding partner for insulin-like growth factor binding protein-3 (IGFBP-3) in solubilized T47D and MCF-7 human breast cancer cell membranes. The interaction between AMF/PGI and IGFBP-3 was verified by cross-linking biotinylated IGFBP-3 to intact cells. After solubilization of the membranes, the biotinylated complexes were precipitated with streptavidin-agarose conjugate and analyzed by SDS-PAGE. A M(r) approximately 80,000 complex was identified when the nitrocellulose membranes were probed either with streptavidin-horseradish peroxidase conjugate or AMF/PGI antiserum confirming the cross-linking of IGFBP-3 to AMF/PGI. The interaction between IGFBP-3 and AMF/PGI was also further confirmed by ligand blotting of purified AMF/PGI using biotinylated IGFBP-3. Both glycosylated and nonglycosylated IGFBP-3 inhibited the catalytic activity of AMF/PGI in a dose-dependent fashion. In addition, IGFBP-3 inhibited the binding of AMF/PGI to breast cancer cells and AMF/PGI-induced migration of both T47D and MCF-7 human breast cancer cells. IGFBP-3 also decreased the phosphorylation of AMF/PGI and reduced the translocation of AMF/PGI to the cell membrane and AMF/PGI. AMF/PGI resulted in a dose-dependent inhibition of IGFBP-3 induced apoptosis in T47D and MCF-7 cells. In summary, we have identified AMF/PGI as a membrane-associated binding partner for IGFBP-3 in breast cancer cells. The ability of IGFBP-3 to bind and inhibit the actions of AMF/PGI may have some role in the antiproliferative proapoptotic effects of IGFBP-3.

  12. Merkel Cell Polyomavirus Small T Antigen Mediates Microtubule Destabilization To Promote Cell Motility and Migration

    PubMed Central

    Knight, Laura M.; Stakaityte, Gabriele; Wood, Jennifer, J.; Abdul-Sada, Hussein; Griffiths, David A.; Howell, Gareth J.; Wheat, Rachel; Blair, G. Eric; Steven, Neil M.; Macdonald, Andrew; Blackbourn, David J.

    2014-01-01

    ABSTRACT Merkel cell carcinoma (MCC) is an aggressive skin cancer of neuroendocrine origin with a high propensity for recurrence and metastasis. Merkel cell polyomavirus (MCPyV) causes the majority of MCC cases due to the expression of the MCPyV small and large tumor antigens (ST and LT, respectively). Although a number of molecular mechanisms have been attributed to MCPyV tumor antigen-mediated cellular transformation or replication, to date, no studies have investigated any potential link between MCPyV T antigen expression and the highly metastatic nature of MCC. Here we use a quantitative proteomic approach to show that MCPyV ST promotes differential expression of cellular proteins implicated in microtubule-associated cytoskeletal organization and dynamics. Intriguingly, we demonstrate that MCPyV ST expression promotes microtubule destabilization, leading to a motile and migratory phenotype. We further highlight the essential role of the microtubule-associated protein stathmin in MCPyV ST-mediated microtubule destabilization and cell motility and implicate the cellular phosphatase catalytic subunit protein phosphatase 4C (PP4C) in the regulation of this process. These findings suggest a possible molecular mechanism for the highly metastatic phenotype associated with MCC. IMPORTANCE Merkel cell polyomavirus (MCPyV) causes the majority of cases of Merkel cell carcinoma (MCC), an aggressive skin cancer with a high metastatic potential. However, the molecular mechanisms leading to virally induced cancer development have yet to be fully elucidated. In particular, no studies have investigated any potential link between the virus and the highly metastatic nature of MCC. We demonstrate that the MCPyV small tumor antigen (ST) promotes the destabilization of the host cell microtubule network, which leads to a more motile and migratory cell phenotype. We further show that MCPyV ST induces this process by regulating the phosphorylation status of the cellular microtubule

  13. GAR22β regulates cell migration, sperm motility, and axoneme structure.

    PubMed

    Gamper, Ivonne; Fleck, David; Barlin, Meltem; Spehr, Marc; El Sayad, Sara; Kleine, Henning; Maxeiner, Sebastian; Schalla, Carmen; Aydin, Gülcan; Hoss, Mareike; Litchfield, David W; Lüscher, Bernhard; Zenke, Martin; Sechi, Antonio

    2016-01-15

    Spatiotemporal cytoskeleton remodeling is pivotal for cell adhesion and migration. Here we investigated the function of Gas2-related protein on chromosome 22 (GAR22β), a poorly characterized protein that interacts with actin and microtubules. Primary and immortalized GAR22β(-/-) Sertoli cells moved faster than wild-type cells. In addition, GAR22β(-/-) cells showed a more prominent focal adhesion turnover. GAR22β overexpression or its reexpression in GAR22β(-/-) cells reduced cell motility and focal adhesion turnover. GAR22β-actin interaction was stronger than GAR22β-microtubule interaction, resulting in GAR22β localization and dynamics that mirrored those of the actin cytoskeleton. Mechanistically, GAR22β interacted with the regulator of microtubule dynamics end-binding protein 1 (EB1) via a novel noncanonical amino acid sequence, and this GAR22β-EB1 interaction was required for the ability of GAR22β to modulate cell motility. We found that GAR22β is highly expressed in mouse testes, and its absence resulted in reduced spermatozoa generation, lower actin levels in testes, and impaired motility and ultrastructural disorganization of spermatozoa. Collectively our findings identify GAR22β as a novel regulator of cell adhesion and migration and provide a foundation for understanding the molecular basis of diverse cytoskeleton-dependent processes. PMID:26564797

  14. GAR22β regulates cell migration, sperm motility, and axoneme structure

    PubMed Central

    Gamper, Ivonne; Fleck, David; Barlin, Meltem; Spehr, Marc; Sayad, Sara El; Kleine, Henning; Maxeiner, Sebastian; Schalla, Carmen; Aydin, Gülcan; Hoss, Mareike; Litchfield, David W.; Lüscher, Bernhard; Zenke, Martin; Sechi, Antonio

    2016-01-01

    Spatiotemporal cytoskeleton remodeling is pivotal for cell adhesion and migration. Here we investigated the function of Gas2-related protein on chromosome 22 (GAR22β), a poorly characterized protein that interacts with actin and microtubules. Primary and immortalized GAR22β−/− Sertoli cells moved faster than wild-type cells. In addition, GAR22β−/− cells showed a more prominent focal adhesion turnover. GAR22β overexpression or its reexpression in GAR22β−/− cells reduced cell motility and focal adhesion turnover. GAR22β–actin interaction was stronger than GAR22β–microtubule interaction, resulting in GAR22β localization and dynamics that mirrored those of the actin cytoskeleton. Mechanistically, GAR22β interacted with the regulator of microtubule dynamics end-binding protein 1 (EB1) via a novel noncanonical amino acid sequence, and this GAR22β–EB1 interaction was required for the ability of GAR22β to modulate cell motility. We found that GAR22β is highly expressed in mouse testes, and its absence resulted in reduced spermatozoa generation, lower actin levels in testes, and impaired motility and ultrastructural disorganization of spermatozoa. Collectively our findings identify GAR22β as a novel regulator of cell adhesion and migration and provide a foundation for understanding the molecular basis of diverse cytoskeleton-dependent processes. PMID:26564797

  15. Spironolactone inhibits podocyte motility via decreasing integrin β1 and increasing integrin β3 in podocytes under high-glucose conditions.

    PubMed

    Li, Zhuo; Zhang, Li; Shi, Wei; Chen, Yuanhan; Zhang, Hong; Liu, Shuangxin; Liang, Xinling; Ling, Ting; Yu, Chunping; Huang, Zhongshun; Tan, Xiaofan; Zhao, Xinchen; Ye, Zhiming; Zhang, Bin; Wang, Wenjian; Li, Ruizhao; Ma, Jianchao

    2015-11-01

    Integrin β1 and β3 expression by podocytes is required to maintain glomerular structural integrity. Previous studies have shown that aldosterone (ALD) is involved in glomerular podocyte injury, and mineralocorticoid receptor (MR) blocker spironolactone effectively reduces proteinuria in patients with diabetic nephropathy. The present study was designed to observe the effects of spironolactone on β1 and β3 integrin expression and podocyte motility under in vitro diabetic conditions. Immortalized mouse podocytes were cultured in media containing normal glucose (NG) levels, high glucose (HG) or HG plus spironolacton. The expression of β1 and β3 integrin in podocytes was detected by reverse transcription quantitative polymerase chain reaction, immunofluorescence and western blot analyses. The effects of spironolacton on podocyte motility was further evaluated using a wound healing assay. HG stimulation markedly decreased mRNA and protein expression of integrin β1, and significantly increased mRNA and protein expression of integrin β3 in cultured podocytes. However, simultaneous treatment with spironolacton (10‑7 mol/l) significantly attenuated HG-mediated increases in integrin β3 and decreases in integrin β1 expression. Furthermore, the migration of podocytes induced by HG was abrogated by concomitant treatment with spironolacton. In conclusion, the present study suggested that HG decreased the expression of integrin β1 in cultured podocytes, accompanied with an increase of integrin β3. Spironolactone inhibited cell motility and stabilized podoctyes treated with HG, probably through partly normalizing the expression of integrin β1 and decreasing the expression of integrin β3. PMID:26352002

  16. Automated single-cell motility analysis on a chip using lensfree microscopy

    PubMed Central

    Pushkarsky, Ivan; Lyb, Yunbo; Weaver, Westbrook; Su, Ting-Wei; Mudanyali, Onur; Ozcan, Aydogan; Di Carlo, Dino

    2014-01-01

    Quantitative cell motility studies are necessary for understanding biophysical processes, developing models for cell locomotion and for drug discovery. Such studies are typically performed by controlling environmental conditions around a lens-based microscope, requiring costly instruments while still remaining limited in field-of-view. Here we present a compact cell monitoring platform utilizing a wide-field (24 mm2) lensless holographic microscope that enables automated single-cell tracking of large populations that is compatible with a standard laboratory incubator. We used this platform to track NIH 3T3 cells on polyacrylamide gels over 20 hrs. We report that, over an order of magnitude of stiffness values, collagen IV surfaces lead to enhanced motility compared to fibronectin, in agreement with biological uses of these structural proteins. The increased throughput associated with lensfree on-chip imaging enables higher statistical significance in observed cell behavior and may facilitate rapid screening of drugs and genes that affect cell motility. PMID:24739819

  17. Uncovering the Mechanism of Trapping and Cell Orientation during Neisseria gonorrhoeae Twitching Motility

    PubMed Central

    Zaburdaev, Vasily; Biais, Nicolas; Schmiedeberg, Michael; Eriksson, Jens; Jonsson, Ann-Beth; Sheetz, Michael P.; Weitz, David A.

    2014-01-01

    Neisseria gonorrheae bacteria are the causative agent of the second most common sexually transmitted infection in the world. The bacteria move on a surface by means of twitching motility. Their movement is mediated by multiple long and flexible filaments, called type IV pili, that extend from the cell body, attach to the surface, and retract, thus generating a pulling force. Moving cells also use pili to aggregate and form microcolonies. However, the mechanism by which the pili surrounding the cell body work together to propel bacteria remains unclear. Understanding this process will help describe the motility of N. gonorrheae bacteria, and thus the dissemination of the disease which they cause. In this article we track individual twitching cells and observe that their trajectories consist of alternating moving and pausing intervals, while the cell body is preferably oriented with its wide side toward the direction of motion. Based on these data, we propose a model for the collective pili operation of N. gonorrheae bacteria that explains the experimentally observed behavior. Individual pili function independently but can lead to coordinated motion or pausing via the force balance. The geometry of the cell defines its orientation during motion. We show that by changing pili substrate interactions, the motility pattern can be altered in a predictable way. Although the model proposed is tangibly simple, it still has sufficient robustness to incorporate further advanced pili features and various cell geometries to describe other bacteria that employ pili to move on surfaces. PMID:25296304

  18. Uncovering the mechanism of trapping and cell orientation during Neisseria gonorrhoeae twitching motility.

    PubMed

    Zaburdaev, Vasily; Biais, Nicolas; Schmiedeberg, Michael; Eriksson, Jens; Jonsson, Ann-Beth; Sheetz, Michael P; Weitz, David A

    2014-10-01

    Neisseria gonorrheae bacteria are the causative agent of the second most common sexually transmitted infection in the world. The bacteria move on a surface by means of twitching motility. Their movement is mediated by multiple long and flexible filaments, called type IV pili, that extend from the cell body, attach to the surface, and retract, thus generating a pulling force. Moving cells also use pili to aggregate and form microcolonies. However, the mechanism by which the pili surrounding the cell body work together to propel bacteria remains unclear. Understanding this process will help describe the motility of N. gonorrheae bacteria, and thus the dissemination of the disease which they cause. In this article we track individual twitching cells and observe that their trajectories consist of alternating moving and pausing intervals, while the cell body is preferably oriented with its wide side toward the direction of motion. Based on these data, we propose a model for the collective pili operation of N. gonorrheae bacteria that explains the experimentally observed behavior. Individual pili function independently but can lead to coordinated motion or pausing via the force balance. The geometry of the cell defines its orientation during motion. We show that by changing pili substrate interactions, the motility pattern can be altered in a predictable way. Although the model proposed is tangibly simple, it still has sufficient robustness to incorporate further advanced pili features and various cell geometries to describe other bacteria that employ pili to move on surfaces. PMID:25296304

  19. 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

  20. PP2A in the regulation of cell motility and invasion.

    PubMed

    Basu, Sunanda

    2011-02-01

    Cell motility is a very critical phenomenon that plays an important role in the development of eukaryotic organisms. One of the well studied cell motility phenomena is chemotaxis, which is described as a directional movement of cell in response to changes in external chemotactic gradient. Numerous studies conducted both in unicellular organism and in mammalian cells have demonstrated the importance of phosphatidylionositol-3 kinase (PI3K) in this process. In addition, it is now well established that although PI3K plays an activation role in chemotaxis, the role of phosphatases is also critical to maintain this dynamic cyclical process. Protein phosphatase 2A (PP2A) is a major serine/threonine phosphatase that is a key player in regulating PI3K signaling. PP2A is abundantly and ubiquitously expressed and has been highly conserved during the evolution of eukaryotes. PP2A is composed of three protein subunits, A, B, and C. Subunit 'A' is a 60-65 kDa structural component, 'C' is a 36-38 kDa catalytic subunit, and 'B' is a 54-130 kDa regulatory subunit. The core complex of PP2A is comprised of the A and C subunits, which are tightly associated and this dimeric core complexes with the regulatory B subunit. The B subunit determines the substrate specificity as well as the spatial and temporal functions of PP2A. PP2A plays an important role in regulating multiple signal transduction pathways, including cell-cycle regulation, cell-growth and development, cytoskeleton dynamics, and cell motility. This review focuses on the role of PP2A in regulating motility of normal and transformed cells.

  1. Regulation of actin-dependent cytoplasmic motility by type II phytochrome occurs within seconds in Vallisneria gigantea epidermal cells.

    PubMed

    Takagi, Shingo; Kong, Sam-Geun; Mineyuki, Yoshinobu; Furuya, Masaki

    2003-02-01

    The effects of light on actin-dependent cytoplasmic motility in epidermal cells of green leaves of the aquatic angiosperm Vallisneria gigantea were investigated quantitatively using a custom-made dynamic image analyzer. Cytoplasmic motility was measured by monitoring changes in the brightness of individual pixels on digitized images taken sequentially under infrared light. Acceleration and deceleration of cytoplasmic motility were regulated photoreversibly by type II phytochrome(s). This phytochrome-dependent induction of cytoplasmic motility did not occur uniformly in cytoplasm but took place as scattered patches in which no particular organelles, including nucleus, existed. The induction became detectable at 2.5 s after the start of irradiation with pulsed red light. In cells exposed to microbeam irradiation, cytoplasmic motility was induced only in sites in the cytoplasm that were irradiated directly, whereas nonirradiated neighboring areas were unaffected. The effect was short-lived, disappearing within a few minutes, and no signal was transmitted from an irradiated cell to its neighbors. Anti-phytochrome antibody-responsive protein(s) was detectable in the leaf extract by immunoblot and zinc blot analyses and in cryosections of the epidermis by immunocytochemistry. Although the phytochrome-dependent cytoplasmic motility was blocked by exogenously applied latrunculin B or cytochalasins, treatment of the dark-adapted cells with Ca(2+)-chelating reagents induced the cytoplasmic motility. We have proposed a model for the phytochrome regulation of cytoplasmic motility as one of the earliest responses to a light stimulus. PMID:12566576

  2. Regulation of Actin-Dependent Cytoplasmic Motility by Type II Phytochrome Occurs within Seconds in Vallisneria gigantea Epidermal Cells

    PubMed Central

    Takagi, Shingo; Kong, Sam-Geun; Mineyuki, Yoshinobu; Furuya, Masaki

    2003-01-01

    The effects of light on actin-dependent cytoplasmic motility in epidermal cells of green leaves of the aquatic angiosperm Vallisneria gigantea were investigated quantitatively using a custom-made dynamic image analyzer. Cytoplasmic motility was measured by monitoring changes in the brightness of individual pixels on digitized images taken sequentially under infrared light. Acceleration and deceleration of cytoplasmic motility were regulated photoreversibly by type II phytochrome(s). This phytochrome-dependent induction of cytoplasmic motility did not occur uniformly in cytoplasm but took place as scattered patches in which no particular organelles, including nucleus, existed. The induction became detectable at 2.5 s after the start of irradiation with pulsed red light. In cells exposed to microbeam irradiation, cytoplasmic motility was induced only in sites in the cytoplasm that were irradiated directly, whereas nonirradiated neighboring areas were unaffected. The effect was short-lived, disappearing within a few minutes, and no signal was transmitted from an irradiated cell to its neighbors. Anti-phytochrome antibody–responsive protein(s) was detectable in the leaf extract by immunoblot and zinc blot analyses and in cryosections of the epidermis by immunocytochemistry. Although the phytochrome-dependent cytoplasmic motility was blocked by exogenously applied latrunculin B or cytochalasins, treatment of the dark-adapted cells with Ca2+-chelating reagents induced the cytoplasmic motility. We have proposed a model for the phytochrome regulation of cytoplasmic motility as one of the earliest responses to a light stimulus. PMID:12566576

  3. Automated characterization of cell shape changes during amoeboid motility by skeletonization

    PubMed Central

    2010-01-01

    Background The ability of a cell to change shape is crucial for the proper function of many cellular processes, including cell migration. One type of cell migration, referred to as amoeboid motility, involves alternating cycles of morphological expansion and retraction. Traditionally, this process has been characterized by a number of parameters providing global information about shape changes, which are insufficient to distinguish phenotypes based on local pseudopodial activities that typify amoeboid motility. Results We developed a method that automatically detects and characterizes pseudopodial behavior of cells. The method uses skeletonization, a technique from morphological image processing to reduce a shape into a series of connected lines. It involves a series of automatic algorithms including image segmentation, boundary smoothing, skeletonization and branch pruning, and takes into account the cell shape changes between successive frames to detect protrusion and retraction activities. In addition, the activities are clustered into different groups, each representing the protruding and retracting history of an individual pseudopod. Conclusions We illustrate the algorithms on movies of chemotaxing Dictyostelium cells and show that our method makes it possible to capture the spatial and temporal dynamics as well as the stochastic features of the pseudopodial behavior. Thus, the method provides a powerful tool for investigating amoeboid motility. PMID:20334652

  4. A specific PTPRC/CD45 phosphorylation event governed by stem cell chemokine CXCL12 regulates primitive hematopoietic cell motility.

    PubMed

    Williamson, Andrew J K; Pierce, Andrew; Jaworska, Ewa; Zhou, Cong; Aspinall-O'Dea, Mark; Lancashire, Lee; Unwin, Richard D; Abraham, Sheela A; Walker, Michael J; Cadecco, Sara; Spooncer, Elaine; Holyoake, Tessa L; Whetton, Anthony D

    2013-11-01

    CXCL12 governs cellular motility, a process deregulated by hematopoietic stem cell oncogenes such as p210-BCR-ABL. A phosphoproteomics approach to the analysis of a hematopoietic progenitor cell line treated with CXCL12 and the Rac 1 and 2 inhibitor NSC23766 has been employed to objectively discover novel mechanisms for regulation of stem cells in normal and malignant hematopoiesis. The proteomic data sets identified new aspects of CXCL12-mediated signaling and novel features of stem cell regulation. We also identified a novel phosphorylation event in hematopoietic progenitor cells that correlated with motile response and governed by the chemotactic factor CXCL12. The novel phosphorylation site on PTPRC/CD45; a protein tyrosine phosphatase, was validated by raising an antibody to the site and also using a mass spectrometry absolute quantification strategy. Site directed mutagenesis and inhibitor studies demonstrated that this single phosphorylation site governs hematopoietic progenitor cell and lymphoid cell motility, lies downstream from Rac proteins and potentiates Src signaling. We have also demonstrated that PTPRC/CD45 is down-regulated in leukemogenic tyrosine kinase expressing cells. The use of discovery proteomics has enabled further understanding of the regulation of PTPRC/CD45 and its important role in cellular motility in progenitor cells.

  5. Electric Cell-substrate Impedance Sensing for the Quantification of Endothelial Proliferation, Barrier Function, and Motility

    PubMed Central

    Szulcek, Robert; Bogaard, Harm Jan; van Nieuw Amerongen, Geerten P.

    2014-01-01

    Electric Cell-substrate Impedance Sensing (ECIS) is an in vitro impedance measuring system to quantify the behavior of cells within adherent cell layers. To this end, cells are grown in special culture chambers on top of opposing, circular gold electrodes. A constant small alternating current is applied between the electrodes and the potential across is measured. The insulating properties of the cell membrane create a resistance towards the electrical current flow resulting in an increased electrical potential between the electrodes. Measuring cellular impedance in this manner allows the automated study of cell attachment, growth, morphology, function, and motility. Although the ECIS measurement itself is straightforward and easy to learn, the underlying theory is complex and selection of the right settings and correct analysis and interpretation of the data is not self-evident. Yet, a clear protocol describing the individual steps from the experimental design to preparation, realization, and analysis of the experiment is not available. In this article the basic measurement principle as well as possible applications, experimental considerations, advantages and limitations of the ECIS system are discussed. A guide is provided for the study of cell attachment, spreading and proliferation; quantification of cell behavior in a confluent layer, with regard to barrier function, cell motility, quality of cell-cell and cell-substrate adhesions; and quantification of wound healing and cellular responses to vasoactive stimuli. Representative results are discussed based on human microvascular (MVEC) and human umbilical vein endothelial cells (HUVEC), but are applicable to all adherent growing cells. PMID:24747269

  6. How Cells Integrate Complex Stimuli: The Effect of Feedback from Phosphoinositides and Cell Shape on Cell Polarization and Motility

    PubMed Central

    Edelstein-Keshet, Leah

    2012-01-01

    To regulate shape changes, motility and chemotaxis in eukaryotic cells, signal transduction pathways channel extracellular stimuli to the reorganization of the actin cytoskeleton. The complexity of such networks makes it difficult to understand the roles of individual components, let alone their interactions and multiple feedbacks within a given layer and between layers of signalling. Even more challenging is the question of if and how the shape of the cell affects and is affected by this internal spatiotemporal reorganization. Here we build on our previous 2D cell motility model where signalling from the Rho family GTPases (Cdc42, Rac, and Rho) was shown to organize the cell polarization, actin reorganization, shape change, and motility in simple gradients. We extend this work in two ways: First, we investigate the effects of the feedback between the phosphoinositides (PIs) , and Rho family GTPases. We show how that feedback increases heights and breadths of zones of Cdc42 activity, facilitating global communication between competing cell “fronts”. This hastens the commitment to a single lamellipodium initiated in response to multiple, complex, or rapidly changing stimuli. Second, we show how cell shape feeds back on internal distribution of GTPases. Constraints on chemical isocline curvature imposed by boundary conditions results in the fact that dynamic cell shape leads to faster biochemical redistribution when the cell is repolarized. Cells with frozen cytoskeleton, and static shapes, consequently respond more slowly to reorienting stimuli than cells with dynamic shape changes, the degree of the shape-induced effects being proportional to the extent of cell deformation. We explain these concepts in the context of several in silico experiments using our 2D computational cell model. PMID:22396633

  7. Rab11a differentially modulates epidermal growth factor-induced proliferation and motility in immortal breast cells.

    PubMed

    Palmieri, Diane; Bouadis, Amina; Ronchetti, Ruban; Merino, Maria J; Steeg, Patricia S

    2006-11-01

    The development of cancer prevention strategies depends on the elucidation of molecular pathways underlying oncogenesis. In a previous proteomic study of matched normal breast ducts and Ductal Carcinoma in Situ (DCIS), we identified overexpression of Rab11a in DCIS. Rab11a is not well studied in cancer, but is known to regulate the recycling of internalized cell surface proteins and receptors from the early endosome through the trans-Golgi network. Using immunohistochemistry, we confirmed our observation, noting increased Rab11a expression in 19 of 22 (86%) DCIS cases compared to matched normal breast epithelium. To study the function of Rab11a, immortal, nontumorigenic MCF10A breast cells were stimulated with ligands to the EGF receptor (EGFR) after transfection with empty vector (control), Rab11a, or a S25N dominant-negative (DN) Rab11a. Using an iodinated ligand:receptor recycling assay, transfection of Rab11a accelerated, while DN-Rab11a postponed EGFR recycling in vitro. The signaling and in vitro phenotypic consequences of Rab11a expression and function were studied. Transfection of DN-Rab11a increased Erk1/2 activation downstream of EGF, but exerted no effect on the Akt pathway. Expression of DN-Rab11a inhibited MCF10A proliferation by 50-60%, and also inhibited anchorage-dependent colonization. Notably, DN-Rab11a transfection increased motility toward EGFR ligands. The data provide a first demonstration that Rab11a modulates EGFR recycling, and promotes the proliferation but inhibits the motility of an immortal breast line, consistent with the DCIS phenotype.

  8. Archaeal Signal Transduction: Impact of Protein Phosphatase Deletions on Cell Size, Motility, and Energy Metabolism in Sulfolobus acidocaldarius*

    PubMed Central

    Reimann, Julia; Esser, Dominik; Orell, Alvaro; Amman, Fabian; Pham, Trong Khoa; Noirel, Josselin; Lindås, Ann-Christin; Bernander, Rolf; Wright, Phillip C.; Siebers, Bettina; Albers, Sonja-Verena

    2013-01-01

    In this study, the in vitro and in vivo functions of the only two identified protein phosphatases, Saci-PTP and Saci-PP2A, in the crenarchaeal model organism Sulfolobus acidocaldarius were investigated. Biochemical characterization revealed that Saci-PTP is a dual-specific phosphatase (against pSer/pThr and pTyr), whereas Saci-PP2A exhibited specific pSer/pThr activity and inhibition by okadaic acid. Deletion of saci_pp2a resulted in pronounced alterations in growth, cell shape and cell size, which could be partially complemented. Transcriptome analysis of the three strains (Δsaci_ptp, Δsaci_pp2a and the MW001 parental strain) revealed 155 genes that were differentially expressed in the deletion mutants, and showed significant changes in expression of genes encoding the archaella (archaeal motility structure), components of the respiratory chain and transcriptional regulators. Phosphoproteome studies revealed 801 unique phosphoproteins in total, with an increase in identified phosphopeptides in the deletion mutants. Proteins from most functional categories were affected by phosphorylation, including components of the motility system, the respiratory chain, and regulatory proteins. In the saci_pp2a deletion mutant the up-regulation at the transcript level, as well as the observed phosphorylation pattern, resembled starvation stress responses. Hypermotility was also observed in the saci_pp2a deletion mutant. The results highlight the importance of protein phosphorylation in regulating essential cellular processes in the crenarchaeon S. acidocaldarius. PMID:24078887

  9. Ophiobolin A from Bipolaris oryzae perturbs motility and membrane integrities of porcine sperm and induces cell death on mammalian somatic cell lines.

    PubMed

    Bencsik, Ottó; Papp, Tamás; Berta, Máté; Zana, Annamária; Forgó, Péter; Dombi, György; Andersson, Maria A; Salkinoja-Salonen, Mirja; Vágvölgyi, Csaba; Szekeres, András

    2014-09-01

    Bipolaris oryzae is a phytopathogenic fungus causing a brown spot disease in rice, and produces substance that strongly perturbs motility and membrane integrities of boar spermatozoa. The substance was isolated from the liquid culture of the fungal strain using extraction and a multi-step semi-preparative HPLC procedures. Based on the results of mass spectrometric and 2D NMR techniques, the bioactive molecule was identified as ophiobolin A, a previously described sesterterpene-type compound. The purified ophiobolin A exhibited strong motility inhibition and viability reduction on boar spermatozoa. Furthermore, it damaged the sperm mitochondria significantly at sublethal concentration by the dissipation of transmembrane potential in the mitochondrial inner membrane, while the plasma membrane permeability barrier remained intact. The study demonstrated that the cytotoxicity of ophiobolin A toward somatic cell lines is higher by 1-2 orders of magnitude compared to other mitochondriotoxic mycotoxins, and towards sperm cells unique by replacing the progressive motility by shivering tail beating at low exposure concentration. PMID:25251540

  10. Cell Motility Is Decreased in Macrophages Activated by Cancer Cell-Conditioned Medium

    PubMed Central

    Go, Ahreum; Ryu, Yun-Kyoung; Lee, Jae-Wook; Moon, Eun-Yi

    2013-01-01

    Macrophages play a role in innate immune responses to various foreign antigens. Many products from primary tumors influence the activation and transmigration of macrophages. Here, we investigated a migration of macrophages stimulated with cancer cell culture-conditioned medium (CM). Macrophage activation by treatment with CM of B16F10 cells were judged by the increase in protein levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX2). The location where macrophages were at 4 h-incubation with control medium or CM was different from where they were at 5 h-incubation in culture dish. Percentage of superimposed macrophages at every 1 h interval was gradually increased by CM treatment as compared to control. Total coverage of migrated track expressed in coordinates was smaller and total distance of migration was shorter in CM-treated macrophages than that in control. Rac1 activity in CM-treated macrophages was also decreased as compared to that in control. When macrophages were treated with CM in the presence of dexamethasone (Dex), an increase in COX2 protein levels, and a decrease in Rac1 activity and total coverage of migration were reversed. In the meanwhile, biphasic changes were detected by Dex treatment in section distance of migration at each time interval, which was more decreased at early time and then increased at later time. Taken together, data demonstrate that macrophage motility could be reduced in accordance with activation in response to cancer cell products. It suggests that macrophage motility could be a novel marker to monitor cancer-associated inflammatory diseases and the efficacy of anti-inflammatory agents. PMID:24404340

  11. Manipulating directional cell motility using intracellular superparamagnetic nanoparticles

    NASA Astrophysics Data System (ADS)

    Bradshaw, Michael; Clemons, Tristan D.; Ho, Diwei; Gutiérrez, Lucía; Lázaro, Francisco J.; House, Michael J.; St. Pierre, Timothy G.; Fear, Mark W.; Wood, Fiona M.; Iyer, K. Swaminathan

    2015-03-01

    This study investigated the ability for magnetic nanoparticles to influence cellular migration in the presence of an external magnetic field. We found that the direction of migrating keratinocytes can be controlled and the migration speed of fibroblasts can be increased with the internalisation of these nanoparticles in the presence of a magnetic field. The possibility of shepherding cells towards a region of interest through the use of internalized nanoparticles is an attractive prospect for cell tracking, cell therapies, and tissue engineering applications.This study investigated the ability for magnetic nanoparticles to influence cellular migration in the presence of an external magnetic field. We found that the direction of migrating keratinocytes can be controlled and the migration speed of fibroblasts can be increased with the internalisation of these nanoparticles in the presence of a magnetic field. The possibility of shepherding cells towards a region of interest through the use of internalized nanoparticles is an attractive prospect for cell tracking, cell therapies, and tissue engineering applications. Electronic supplementary information (ESI) available: Nanoparticle characterisation, supporting experimental data, video time course study of cellular uptake of the nanoparticles and complete experimental details are all provided in the ESI. See DOI: 10.1039/c4nr06594h

  12. Direct Correlation between Motile Behavior and Protein Abundance in Single Cells.

    PubMed

    Dufour, Yann S; Gillet, Sébastien; Frankel, Nicholas W; Weibel, Douglas B; Emonet, Thierry

    2016-09-01

    Understanding how stochastic molecular fluctuations affect cell behavior requires the quantification of both behavior and protein numbers in the same cells. Here, we combine automated microscopy with in situ hydrogel polymerization to measure single-cell protein expression after tracking swimming behavior. We characterized the distribution of non-genetic phenotypic diversity in Escherichia coli motility, which affects single-cell exploration. By expressing fluorescently tagged chemotaxis proteins (CheR and CheB) at different levels, we quantitatively mapped motile phenotype (tumble bias) to protein numbers using thousands of single-cell measurements. Our results disagreed with established models until we incorporated the role of CheB in receptor deamidation and the slow fluctuations in receptor methylation. Beyond refining models, our central finding is that changes in numbers of CheR and CheB affect the population mean tumble bias and its variance independently. Therefore, it is possible to adjust the degree of phenotypic diversity of a population by adjusting the global level of expression of CheR and CheB while keeping their ratio constant, which, as shown in previous studies, confers functional robustness to the system. Since genetic control of protein expression is heritable, our results suggest that non-genetic diversity in motile behavior is selectable, supporting earlier hypotheses that such diversity confers a selective advantage. PMID:27599206

  13. Tyrosyl Phosphorylated PAK1 Regulates Breast Cancer Cell Motility in Response to Prolactin through Filamin A

    PubMed Central

    Hammer, Alan; Rider, Leah; Oladimeji, Peter; Cook, Leslie; Li, Quanwen; Mattingly, Raymond R.

    2013-01-01

    The p21-activated serine-threonine kinase (PAK1) is activated by small GTPase-dependent and -independent mechanisms and regulates cell motility. Both PAK1 and the hormone prolactin (PRL) have been implicated in breast cancer by numerous studies. We have previously shown that the PRL-activated tyrosine kinase JAK2 (Janus tyrosine kinase 2) phosphorylates PAK1 in vivo and identified tyrosines (Tyr) 153, 201, and 285 in the PAK1 molecule as sites of JAK2 tyrosyl phosphorylation. Here, we have used human breast cancer T47D cells stably overexpressing PAK1 wild type or PAK1 Y3F mutant in which Tyr(s) 153, 201, and 285 were mutated to phenylalanines to demonstrate that phosphorylation of these three tyrosines are required for maximal PRL-dependent ruffling. In addition, phosphorylation of these three tyrosines is required for increased migration of T47D cells in response to PRL as assessed by two independent motility assays. Finally, we show that PAK1 phosphorylates serine (Ser) 2152 of the actin-binding protein filamin A to a greater extent when PAK1 is tyrosyl phosphorylated by JAK2. Down-regulation of PAK1 or filamin A abolishes the effect of PRL on cell migration. Thus, our data presented here bring some insight into the mechanism of PRL-stimulated motility of breast cancer cells. PMID:23340249

  14. Direct Correlation between Motile Behavior and Protein Abundance in Single Cells

    PubMed Central

    Gillet, Sébastien; Frankel, Nicholas W.; Weibel, Douglas B.

    2016-01-01

    Understanding how stochastic molecular fluctuations affect cell behavior requires the quantification of both behavior and protein numbers in the same cells. Here, we combine automated microscopy with in situ hydrogel polymerization to measure single-cell protein expression after tracking swimming behavior. We characterized the distribution of non-genetic phenotypic diversity in Escherichia coli motility, which affects single-cell exploration. By expressing fluorescently tagged chemotaxis proteins (CheR and CheB) at different levels, we quantitatively mapped motile phenotype (tumble bias) to protein numbers using thousands of single-cell measurements. Our results disagreed with established models until we incorporated the role of CheB in receptor deamidation and the slow fluctuations in receptor methylation. Beyond refining models, our central finding is that changes in numbers of CheR and CheB affect the population mean tumble bias and its variance independently. Therefore, it is possible to adjust the degree of phenotypic diversity of a population by adjusting the global level of expression of CheR and CheB while keeping their ratio constant, which, as shown in previous studies, confers functional robustness to the system. Since genetic control of protein expression is heritable, our results suggest that non-genetic diversity in motile behavior is selectable, supporting earlier hypotheses that such diversity confers a selective advantage. PMID:27599206

  15. 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

  16. The actin gene ACT1 is required for phagocytosis, motility, and cell separation of Tetrahymena thermophila.

    PubMed

    Williams, Norman E; Tsao, Che-Chia; Bowen, Josephine; Hehman, Gery L; Williams, Ruth J; Frankel, Joseph

    2006-03-01

    A previously identified Tetrahymena thermophila actin gene (C. G. Cupples and R. E. Pearlman, Proc. Natl. Acad. Sci. USA 83:5160-5164, 1986), here called ACT1, was disrupted by insertion of a neo3 cassette. Cells in which all expressed copies of this gene were disrupted exhibited intermittent and extremely slow motility and severely curtailed phagocytic uptake. Transformation of these cells with inducible genetic constructs that contained a normal ACT1 gene restored motility. Use of an epitope-tagged construct permitted visualization of Act1p in the isolated axonemes of these rescued cells. In ACT1Delta mutant cells, ultrastructural abnormalities of outer doublet microtubules were present in some of the axonemes. Nonetheless, these cells were still able to assemble cilia after deciliation. The nearly paralyzed ACT1Delta cells completed cleavage furrowing normally, but the presumptive daughter cells often failed to separate from one another and later became reintegrated. Clonal analysis revealed that the cell cycle length of the ACT1Delta cells was approximately double that of wild-type controls. Clones could nonetheless be maintained for up to 15 successive fissions, suggesting that the ACT1 gene is not essential for cell viability or growth. Examination of the cell cortex with monoclonal antibodies revealed that whereas elongation of ciliary rows and formation of oral structures were normal, the ciliary rows of reintegrated daughter cells became laterally displaced and sometimes rejoined indiscriminately across the former division furrow. We conclude that Act1p is required in Tetrahymena thermophila primarily for normal ciliary motility and for phagocytosis and secondarily for the final separation of daughter cells.

  17. Pancreatic spasmolytic polypeptide protects the gastric mucosa but does not inhibit acid secretion or motility.

    PubMed

    McKenzie, C; Marchbank, T; Playford, R J; Otto, W; Thim, L; Parsons, M E

    1997-07-01

    The objectives of these studies were to examine whether the trefoil peptide porcine pancreatic spasmolytic polypeptide (PSP) had gastric mucosal protectant properties similar to its human equivalent human spasmolytic polypeptide (hSP) and to confirm the antisecretory and antimotility action of the peptide. PSP and recombinant hSP reduced gastric mucosal damage caused by a combination of subcutaneous indomethacin and restraint stress in the conscious rat. At a dose of 500 micrograms/kg bolus plus 500 micrograms.kg-1.h-1 sc, PSP significantly reduced the total area of damage by 58%. PSP at a dose of 150 micrograms/kg iv had no inhibitory effect on pentagastrin-stimulated gastric acid secretion in the perfused stomachs of anesthetized rats. This lack of antisecretory activity was confirmed in vitro using an isolated stomach preparation from the immature rat. PSP and hSP at concentrations up to 800 nM did not inhibit electrically or chemically evoked contractions of the guinea pig ileum and duodenum in vitro. Thus antisecretory and antimotility actions do not underlie the mucosal protectant properties of PSP. PSP did, however, stimulate cell migration, and this may, at least in part, account for its protectant properties.

  18. Comparison of actin and cell surface dynamics in motile fibroblasts

    PubMed Central

    1992-01-01

    We have investigated the dynamic behavior of actin in fibroblast lamellipodia using photoactivation of fluorescence. Activated regions of caged resorufin (CR)-labeled actin in lamellipodia of IMR 90 and MC7 3T3 fibroblasts were observed to move centripetally over time. Thus in these cells, actin filaments move centripetally relative to the substrate. Rates were characteristic for each cell type; 0.66 +/- 0.27 microns/min in IMR 90 and 0.36 +/- 0.16 microns/min in MC7 3T3 cells. In neither case was there any correlation between the rate of actin movement and the rate of lamellipodial protrusion. The half-life of the activated CR-actin filaments was approximately 1 min in IMR 90 lamellipodia, and approximately 3 min in MC7 3T3 lamellipodia. Thus continuous filament turnover accompanies centripetal movement. In both cell types, the length of time required for a section of the actin meshwork to traverse the lamellipodium was several times longer than the filament half-life. The dynamic behavior of the dorsal surface of the cell was also observed by tracking lectin-coated beads on the surface and phase-dense features within lamellipodia of MC7 3T3 cells. The movement of these dorsal features occurred at rates approximately three times faster than the rate of movement of the underlying bulk actin cytoskeleton, even when measured in the same individual cells. Thus the transport of these dorsal features must occur by some mechanism other than simple attachment to the moving bulk actin cytoskeleton. PMID:1400580

  19. Chemokine-guided cell migration and motility in zebrafish development.

    PubMed

    Bussmann, Jeroen; Raz, Erez

    2015-05-12

    Chemokines are vertebrate-specific, structurally related proteins that function primarily in controlling cell movements by activating specific 7-transmembrane receptors. Chemokines play critical roles in a large number of biological processes and are also involved in a range of pathological conditions. For these reasons, chemokines are at the focus of studies in developmental biology and of clinically oriented research aimed at controlling cancer, inflammation, and immunological diseases. The small size of the zebrafish embryos, their rapid external development, and optical properties as well as the large number of eggs and the fast expansion in genetic tools available make this model an extremely useful one for studying the function of chemokines and chemokine receptors in an in vivo setting. Here, we review the findings relevant to the role that chemokines play in the context of directed single-cell migration, primarily in neutrophils and germ cells, and compare it to the collective cell migration of the zebrafish lateral line. We present the current knowledge concerning the formation of the chemokine gradient, its interpretation within the cell, and the molecular mechanisms underlying the cellular response to chemokine signals during directed migration.

  20. Constitutively Active 5-HT Receptors: An Explanation of How 5-HT Antagonists Inhibit Gut Motility in Species Where 5-HT is Not an Enteric Neurotransmitter?

    PubMed Central

    Spencer, Nick J.

    2015-01-01

    Antagonists of 5-Hydroxytryptamine (5-HT) receptors are well known to inhibit gastrointestinal (GI)-motility and transit in a variety of mammals, including humans. Originally, these observations had been interpreted by many investigators (including us) as evidence that endogenous 5-HT plays a major role in GI motility. This seemed a logical assumption. However, the story changed dramatically after recent studies revealed that 5-HT antagonists still blocked major GI motility patterns (peristalsis and colonic migrating motor complexes) in segments of intestine depleted of all 5-HT. Then, these results were further supported by Dr. Gershons' laboratory, which showed that genetic deletion of all genes that synthesizes 5-HT had minor, or no inhibitory effects on GI transit in vivo. If 5-HT was essential for GI motility patterns and transit, then one would expect major disruptions in motility and transit when 5-HT synthesis was genetically ablated. This does not occur. The inhibitory effects of 5-HT antagonists on GI motility clearly occur independently of any 5-HT in the gut. Evidence now suggests that 5-HT antagonists act on 5-HT receptors in the gut which are constitutively active, and don't require 5-HT for their activation. This would explain a long-standing mystery of how 5-HT antagonists inhibit gut motility in species like mice, rats, and humans where 5-HT is not an enteric neurotransmitter. Studies are now increasingly demonstrating that the presence of a neurochemical in enteric neurons does not mean they function as neurotransmitters. Caution should be exercised when interpreting any inhibitory effects of 5-HT antagonists on GI motility. PMID:26732863

  1. Motility of Mycoplasma pneumoniae.

    PubMed Central

    Radestock, U; Bredt, W

    1977-01-01

    Cell of Mycoplasma pneumoniae FH gliding on a glass surface in liquid medium were examined by microscopic observation and quantitatively by microcinematography (30 frames per min). Comparisons were made only within the individual experiments. The cells moved in an irregular pattern with numerous narrow bends and circles. They never changed their leading end. The average speed (without pauses) was relatively constant between o.2 and 0.5 mum/s. The maximum speed was about 1.5 to 2.0 mum/s. The movements were interrupted by resting periods of different lengths and frequency. Temperature, viscosity, pH, and the presence of yeast extract in the medium influenced the motility significantly; changes in glucose, calcium ions, and serum content were less effective. The movements were affected by iodoacetate, p-mercuribenzoate, and mitomycin C at inhibitory or subinhibitory concentrations. Sodium fluoride, sodium cyanide, dinitrophenol, chloramphenicol, puromycin, cholchicin, and cytochalasin B at minimal inhibitory concentrations did not affect motility. The movements were effectively inhibited by anti-M. pneumoniae antiserum. Studies with absorbed antiserum suggested that the surface components involved in motility are heat labile. The gliding of M. pneumoniae cells required an intact energy metabolism and the proteins involved seemed to have a low turnover. Images PMID:14925

  2. Concerted Action of Two Formins in Gliding Motility and Host Cell Invasion by Toxoplasma gondii

    PubMed Central

    Daher, Wassim; Plattner, Fabienne; Carlier, Marie-France; Soldati-Favre, Dominique

    2010-01-01

    The invasive forms of apicomplexan parasites share a conserved form of gliding motility that powers parasite migration across biological barriers, host cell invasion and egress from infected cells. Previous studies have established that the duration and direction of gliding motility are determined by actin polymerization; however, regulators of actin dynamics in apicomplexans remain poorly characterized. In the absence of a complete ARP2/3 complex, the formin homology 2 domain containing proteins and the accessory protein profilin are presumed to orchestrate actin polymerization during host cell invasion. Here, we have undertaken the biochemical and functional characterization of two Toxoplasma gondii formins and established that they act in concert as actin nucleators during invasion. The importance of TgFRM1 for parasite motility has been assessed by conditional gene disruption. The contribution of each formin individually and jointly was revealed by an approach based upon the expression of dominant mutants with modified FH2 domains impaired in actin binding but still able to dimerize with their respective endogenous formin. These mutated FH2 domains were fused to the ligand-controlled destabilization domain (DD-FKBP) to achieve conditional expression. This strategy proved unique in identifying the non-redundant and critical roles of both formins in invasion. These findings provide new insights into how controlled actin polymerization drives the directional movement required for productive penetration of parasites into host cells. PMID:20949068

  3. Antisense inhibition of hyaluronan synthase-2 in human osteosarcoma cells inhibits hyaluronan retention and tumorigenicity

    SciTech Connect

    Nishida, Yoshihiro . E-mail: ynishida@med.nagoya-u.ac.jp; Knudson, Warren; Knudson, Cheryl B.; Ishiguro, Naoki

    2005-07-01

    Osteosarcoma is a common malignant bone tumor associated with childhood and adolescence. The results of numerous studies have suggested that hyaluronan plays an important role in regulating the aggressive behavior of various types of cancer cells. However, no studies have addressed hyaluronan with respect to osteosarcomas. In this investigation, the mRNA expression copy number of three mammalian hyaluronan synthases (HAS) was determined using competitive RT-PCR in the osteoblastic osteosarcoma cell line, MG-63. MG-63 are highly malignant osteosarcoma cells with an abundant hyaluronan-rich matrix. The results demonstrated that HAS-2 is the predominant HAS in MG-63. Accumulation of intracellular hyaluronan increased in association with the proliferative phase of these cells. The selective inhibition of HAS-2 mRNA in MG-63 cells by antisense phosphorothioate oligonucleotides resulted in reduced hyaluronan accumulation by these cells. As expected, the reduction in hyaluronan disrupted the assembly of cell-associated matrices. However, of most interest, coincident with the reduction in hyaluronan, there was a substantial decrease in cell proliferation, a decrease in cell motility and a decrease in cell invasiveness. These data suggest that hyaluronan synthesized by HAS-2 in MG-63 plays a crucial role in osteosarcoma cell proliferation, motility, and invasion.

  4. Programmable manipulation of motile cells in optoelectronic tweezers using a grayscale image

    NASA Astrophysics Data System (ADS)

    Choi, Wonjae; Nam, Seong-Won; Hwang, Hyundoo; Park, Sungsu; Park, Je-Kyun

    2008-10-01

    This paper describes a grayscale optoelectronic tweezers (OET) which allows adjustment of the electric field strength at each position of OET. A grayscale light image was used to pattern vertical electric field strength on an OET. As an electric field depends on the brightness at each point, the brighter light patterns generate the stronger electric field in the OET. Its feasibility for application to cell manipulation was demonstrated by aligning highly motile protozoan cells in vertical direction. Depending on the brightness of each pixel, the behaviors of aligned cells varied due to the different electric field strength to each cell.

  5. The neuronal differentiation factor NeuroD1 downregulates the neuronal repellent factor Slit2 expression and promotes cell motility and tumor formation of neuroblastoma.

    PubMed

    Huang, Peng; Kishida, Satoshi; Cao, Dongliang; Murakami-Tonami, Yuko; Mu, Ping; Nakaguro, Masato; Koide, Naoshi; Takeuchi, Ichiro; Onishi, Akira; Kadomatsu, Kenji

    2011-04-15

    The basic helix-loop-helix transcription factor NeuroD1 has been implicated in the neurogenesis and early differentiation of pancreatic endocrine cells. However, its function in relation to cancer has been poorly examined. In this study, we found that NeuroD1 is involved in the tumorigenesis of neuroblastoma. NeuroD1 was strongly expressed in a hyperplastic region comprising neuroblasts in the celiac sympathetic ganglion of 2-week-old MYCN transgenic (Tg) mice and was consistently expressed in the subsequently generated neuroblastoma tissue. NeuroD1 knockdown by short hairpin RNA (shRNA) resulted in motility inhibition of the human neuroblastoma cell lines, and this effect was reversed by shRNA-resistant NeuroD1. The motility inhibition by NeuroD1 knockdown was associated with induction of Slit2 expression, and knockdown of Slit2 could restore cell motility. Consistent with this finding, shRNA-resistant NeuroD1 suppressed Slit2 expression. NeuroD1 directly bound to the first and second E-box of the Slit2 promoter region. Moreover, we found that the growth of tumor spheres, established from neuroblastoma cell lines in MYCN Tg mice, was suppressed by NeuroD1 suppression. The functions identified for NeuroD1 in cell motility and tumor sphere growth may suggest a link between NeuroD1 and the tumorigenesis of neuroblastoma. Indeed, tumor formation of tumor sphere-derived cells was significantly suppressed by NeuroD1 knockdown. These data are relevant to the clinical features of human neuroblastoma: high NeuroD1 expression was closely associated with poor prognosis. Our findings establish the critical role of the neuronal differentiation factor NeuroD1 in neuroblastoma as well as its functional relationship with the neuronal repellent factor Slit2.

  6. Effects of PNPG on cell growth cycle, motility machinery and quorum sensing in Serratia marcescens.

    PubMed

    Wei, Jun-Rong; Horng, Yu-Tze; Lai, Hsin-Chih; Luh, Kwen-Tay; Ho, Shen-Wu

    2004-02-01

    p-Nitrophenylglycerol (PNPG) effectively inhibits swarming of the enterobacterium Proteus mirabilis. The underlying mechanism of inhibition is unclear. We have now found that both PNPG also inhibits motility and swarming in another enterobacterium, Serratia marcescens. While the peak promoter activities of the flagellar master operon (flhDCSm), the flagellin structural gene (hagSm) and the nuclease gene (nucASm) in S. marcescens increased with increasing PNPG concentration, the expression of these genes was delayed in accordance with the reduced growth rate. As the quorum-sensing system is involved in the regulation of swarming in S. marcescens, we also examined the effect of PNPG on the production of quorum-sensing signal molecules and found that their expression was delayed with a reduced level. PNPG, therefore, had a pleiotropic effect on all aspects of S. marcescens physiology relating to swarming. The underlying molecular mechanism remains to be elucidated.

  7. Automated tracking and laser micromanipulation of motile cells

    NASA Astrophysics Data System (ADS)

    Stuhrmann, B.; Gögler, M.; Betz, T.; Ehrlicher, A.; Koch, D.; Käs, J.

    2005-03-01

    Control over neuronal growth is a prerequisite for the creation of defined in vitro neuronal networks as assays for the elucidation of interneuronal communication. Neuronal growth has been directed by focusing a near-infrared laser beam at a nerve cell's leading edge [A. Ehrlicher, T. Betz, B. Stuhrmann, D. Koch, V. Milner, M. G. Raizen, and J. Käs, Proc. Natl. Acad. Sci. U.S.A. 99, 16024 (2002)]. The setup reported by Ehrlicher et al. was limited to local laser irradiation and relied on a great deal of subjective interaction since the laser beam could only be steered manually. To overcome the drawbacks of the reported setup, we developed and here present a fully automated low-contrast edge detection software package, which responds to detected cell morphological changes by rapidly actuating laser steering devices, such as acousto-optical deflectors or moving mirrors, thus enabling experiments with minimum human interference. The resulting radiation patterns can be arbitrary functions of space, time, and cell morphology, and are calculated by experiment specific feedback routines. Data processing is repeated on the order of 1s allowing rapid reactions to morphological changes. The strengths of our program are the combination of real-time low contrast shape detection with complex feedback mechanisms, as well as easy adaptability due to a modular programming concept. In this article we demonstrate automated optical guidance; however, the software is easily adaptable to other problems requiring automated rapid responses of equipment to changes in the morphology of low contrast objects.

  8. Effect of cell physicochemical characteristics and motility on bacterial transport in groundwater.

    PubMed

    Becker, Matthew W; Collins, Samantha A; Metge, David W; Harvey, Ronald W; Shapiro, Allen M

    2004-04-01

    The influence of physicochemical characteristics and motility on bacterial transport in groundwater were examined in flow-through columns. Four strains of bacteria isolated from a crystalline rock groundwater system were investigated, with carboxylate-modified and amidine-modified latex microspheres and bromide as reference tracers. The bacterial isolates included a gram-positive rod (ML1), a gram-negative motile rod (ML2), a nonmotile mutant of ML2 (ML2m), and a gram-positive coccoid (ML3). Experiments were repeated at two flow velocities, in a glass column packed with glass beads, and in another packed with iron-oxyhydroxide coated glass beads. Bacteria breakthrough curves were interpreted using a transport equation that incorporates a sorption model from microscopic observation of bacterial deposition in flow-cell experiments. The model predicts that bacterial desorption rate will decrease exponentially with the amount of time the cell is attached to the solid surface. Desorption kinetics appeared to influence transport at the lower flow rate, but were not discernable at the higher flow rate. Iron-oxyhydroxide coatings had a lower-than-expected effect on bacterial breakthrough and no effect on the microsphere recovery in the column experiments. Cell wall type and shape also had minor effects on breakthrough. Motility tended to increase the adsorption rate, and decrease the desorption rate. The transport model predicts that at field scale, desorption rate kinetics may be important to the prediction of bacteria transport rates.

  9. BMP promotes motility and represses growth of smooth muscle cells by activation of tandem Wnt pathways

    PubMed Central

    de Jesus Perez, Vinicio A.; Ali, Ziad; Alastalo, Tero-Pekka; Ikeno, Fumiaki; Sawada, Hirofumi; Lai, Ying-Ju; Kleisli, Thomas; Spiekerkoetter, Edda; Qu, Xiumei; Rubinos, Laura H.; Ashley, Euan; Amieva, Manuel; Dedhar, Shoukat

    2011-01-01

    We present a novel cell-signaling paradigm in which bone morphogenetic protein 2 (BMP-2) consecutively and interdependently activates the wingless (Wnt)–β-catenin (βC) and Wnt–planar cell polarity (PCP) signaling pathways to facilitate vascular smooth muscle motility while simultaneously suppressing growth. We show that BMP-2, in a phospho-Akt–dependent manner, induces βC transcriptional activity to produce fibronectin, which then activates integrin-linked kinase 1 (ILK-1) via α4-integrins. ILK-1 then induces the Wnt–PCP pathway by binding a proline-rich motif in disheveled (Dvl) and consequently activating RhoA-Rac1–mediated motility. Transfection of a Dvl mutant that binds βC without activating RhoA-Rac1 not only prevents BMP-2–mediated vascular smooth muscle cell motility but promotes proliferation in association with persistent βC activity. Interfering with the Dvl-dependent Wnt–PCP activation in a murine stented aortic graft injury model promotes extensive neointima formation, as shown by optical coherence tomography and histopathology. We speculate that, in response to injury, factors that subvert BMP-2–mediated tandem activation of Wnt–βC and Wnt–PCP pathways contribute to obliterative vascular disease in both the systemic and pulmonary circulations. PMID:21220513

  10. Effect of cell physicochemical characteristics and motility on bacterial transport in groundwater

    USGS Publications Warehouse

    Becker, M.W.; Collins, S.A.; Metge, D.W.; Harvey, R.W.; Shapiro, A.M.

    2004-01-01

    The influence of physicochemical characteristics and motility on bacterial transport in groundwater were examined in flow-through columns. Four strains of bacteria isolated from a crystalline rock groundwater system were investigated, with carboxylate-modified and amidine-modified latex microspheres and bromide as reference tracers. The bacterial isolates included a gram-positive rod (ML1), a gram-negative motile rod (ML2), a nonmotile mutant of ML2 (ML2m), and a gram-positive coccoid (ML3). Experiments were repeated at two flow velocities, in a glass column packed with glass beads, and in another packed with iron-oxyhydroxide coated glass beads. Bacteria breakthrough curves were interpreted using a transport equation that incorporates a sorption model from microscopic observation of bacterial deposition in flow-cell experiments. The model predicts that bacterial desorption rate will decrease exponentially with the amount of time the cell is attached to the solid surface. Desorption kinetics appeared to influence transport at the lower flow rate, but were not discernable at the higher flow rate. Iron-oxyhydroxide coatings had a lower-than-expected effect on bacterial breakthrough and no effect on the microsphere recovery in the column experiments. Cell wall type and shape also had minor effects on breakthrough. Motility tended to increase the adsorption rate, and decrease the desorption rate. The transport model predicts that at field scale, desorption rate kinetics may be important to the prediction of bacteria transport rates. ?? 2003 Elsevier B.V. All rights reserved.

  11. 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.

  12. Antihuman epidermal growth factor receptor 2 antibody herceptin inhibits autocrine motility factor (AMF) expression and potentiates antitumor effects of AMF inhibitors.

    PubMed

    Talukder, Amjad H; Bagheri-Yarmand, Rozita; Williams, Ruth R E; Ragoussis, Jiannis; Kumar, Rakesh; Raz, Avraham

    2002-10-01

    Overexpression of the human epidermal growth factor receptor (HER) 2 has been linked to the development and maintenance of malignant phenotypes in breast tumors. In addition, the growth and dissemination of human cancers are regulated in part by the autocrine motility factor (AMF)/phosphoglucose isomerase shown to be up-regulated by heregulin (HRG) in breast cancer cells. This study was undertaken to explore the effect of anti-HER2 monoclonal antibody 4D5 [Herceptin (HCT)] on AMF expression and the potential of its augmentation by specific simple sugar AMF inhibitors. Here we show that HCT treatment of high HER2-expressing breast cancer SK-BR3, BT-474, and ZR-75R cells resulted in down-regulation of AMF mRNA and protein. HCT inhibited the ability of HRG to induce AMF expression in cells with a normal HER2 level, and HCT-mediated down-regulation could be reversed by HRG treatment in breast cancer cells with a high HER2 level. HCT also inhibited transcription from a chimeric pGL3-Luc vector-based reporter system containing the 1.8-kb promoter region of human AMF. Treatment of breast cancer cells with the combination of HCT and specific AMF inhibitors, erythrose 4-phosphate or D-mannose 6-phosphate, resulted in an additive inhibitory effect on both the growth rate and invasiveness of cells as compared with treatment with each agent alone. Results presented here suggest that HCT can effectively block both ligand-induced and constitutive expression of AMF associated with high HER2 overexpression, implying a role of the AMF pathway in the action of HCT. Accordingly, the combination of AMF inhibitor with HCT can potentiate the growth-inhibitory and anti-invasive action of HCT in breast cancer cells.

  13. 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.

  14. On an evolution equation in a cell motility model

    NASA Astrophysics Data System (ADS)

    Mizuhara, Matthew S.; Berlyand, Leonid; Rybalko, Volodymyr; Zhang, Lei

    2016-04-01

    This paper deals with the evolution equation of a curve obtained as the sharp interface limit of a non-linear system of two reaction-diffusion PDEs. This system was introduced as a phase-field model of (crawling) motion of eukaryotic cells on a substrate. The key issue is the evolution of the cell membrane (interface curve) which involves shape change and net motion. This issue can be addressed both qualitatively and quantitatively by studying the evolution equation of the sharp interface limit for this system. However, this equation is non-linear and non-local and existence of solutions presents a significant analytical challenge. We establish existence of solutions for a wide class of initial data in the so-called subcritical regime. Existence is proved in a two step procedure. First, for smooth (H2) initial data we use a regularization technique. Second, we consider non-smooth initial data that are more relevant from the application point of view. Here, uniform estimates on the time when solutions exist rely on a maximum principle type argument. We also explore the long time behavior of the model using both analytical and numerical tools. We prove the nonexistence of traveling wave solutions with nonzero velocity. Numerical experiments show that presence of non-linearity and asymmetry of the initial curve results in a net motion which distinguishes it from classical volume preserving curvature motion. This is done by developing an algorithm for efficient numerical resolution of the non-local term in the evolution equation.

  15. Hepatocyte growth factor is a potent angiogenic factor which stimulates endothelial cell motility and growth

    PubMed Central

    1992-01-01

    Hepatocyte Growth Factor (HGF, also known as Scatter Factor) is a powerful mitogen or motility factor in different cells, acting through the tyrosine kinase receptor encoded by the MET protooncogene. Endothelial cells express the MET gene and expose at the cell surface the mature protein (p190MET) made of a 50 kD (alpha) subunit disulfide linked to a 145-kD (beta) subunit. HGF binding to endothelial cells identifies two sites with different affinities. The higher affinity binding site (Kd = 0.35 nM) corresponds to the p190MET receptor. Sub- nanomolar concentrations of HGF, but not of a recombinant inactive precursor, stimulate the receptor kinase activity, cell proliferation and motility. HGF induces repairs of a wound in endothelial cell monolayer. HGF stimulates the scatter of endothelial cells grown on three-dimensional collagen gels, inducing an elongated phenotype. In the rabbit cornea, highly purified HGF promotes neovascularization at sub-nanomolar concentrations. HGF lacks activities related to hemostasis-thrombosis, inflammation and endothelial cells accessory functions. These data show that HGF is an in vivo potent angiogenic factor and in vitro induces endothelial cells to proliferate and migrate. PMID:1383237

  16. Modeling invasion of brain tissue by glioblastoma cells: ECM alignment and motility

    NASA Astrophysics Data System (ADS)

    Sander, L. M.

    2013-03-01

    A key stage in the development of highly malignant brain tumors (Glioblastoma Multiforme) is invasion of normal brain tissue by motile cells moving through a crowded, complex environment. Evidence from in vitro experiments suggests the cell motion is accompanied by considerable deformation and alignment of the extra-cellular matrix (ECM) of the brain. In the case of breast cancer, alignment effects of this sort have been seen in vivo. We have modeled features of this system including stress confinement in the non-linear elasticity of the ECM and contact guidance of the cell motion.

  17. Reassessing the mechanics of parasite motility and host-cell invasion.

    PubMed

    Tardieux, Isabelle; Baum, Jake

    2016-08-29

    The capacity to migrate is fundamental to multicellular and single-celled life. Apicomplexan parasites, an ancient protozoan clade that includes malaria parasites (Plasmodium) and Toxoplasma, achieve remarkable speeds of directional cell movement. This rapidity is achieved via a divergent actomyosin motor system, housed within a narrow compartment that lies underneath the length of the parasite plasma membrane. How this motor functions at a mechanistic level during motility and host cell invasion is a matter of debate. Here, we integrate old and new insights toward refining the current model for the function of this motor with the aim of revitalizing interest in the mechanics of how these deadly pathogens move. PMID:27573462

  18. Actomyosin-based motility of endoplasmic reticulum and chloroplasts in Vallisneria mesophyll cells.

    PubMed

    Liebe, S; Menzel, D

    1995-01-01

    Intracellular localization and motile behaviour of the endoplasmic reticulum (ER), plastids and mitochondria were studied in living mesophyll cells of Vallisneria using the vital fluorochrome 3,3'-dihexyloxacarbocyanine iodide (DIOC6(3)). In quiescent cells, the ER was composed of a three-dimensional network of tubular and lamellar elements. Chloroplasts were distributed evenly throughout the cell periphery and appeared embedded within the ER network. The ER network was relatively stationary, with the exception of rare motile episodes occurring as movement of tubular ER strands and adjacent areas of the polygonal network in localized areas of the cell. During experimental induction of streaming, most of the lamellar ER elements transformed into tubules and together with the chloroplasts they began to translocate to the anticlinal walls to establish the circular streaming around the circumference of the cell. Microwave-accelerated fixation followed by immunofluorescence revealed an hitherto unknown phase of actin reorganization occurring within the cells and most interestingly at the surface of the chloroplasts during streaming induction. Myosin was localized in an ER-like pattern in quiescent as well as in streaming cells, with bright fluorescent label localized on mitochondria and proplastids. In addition, myosin label appeared on the surface of the chloroplasts, preferentially in streaming mesophyll cells. Motile activities were impeded by the actin-depolymerizing drug cytochalasin D (CD), the thioreagent N-ethylmaleimide (NEM), and thapsigargin, an inhibitor of the ER-Ca(2+)-ATPase. These inhibitors also interfered with the integrity of actin filaments, the intracellular distribution of myosin and calcium-homeostasis, respectively. These effects suggested an obligate association of at least one type of myosin with the membranes of ER and smaller organelles and are consistent with the appearance of another type of myosin on the chloroplast surface upon streaming

  19. The role of filament-packing dynamics in powering amoeboid cell motility

    PubMed Central

    Miao, Long; Vanderlinde, Orion; Liu, Jun; Grant, Richard P.; Wouterse, Alan; Shimabukuro, Katsuya; Philipse, Albert; Stewart, Murray; Roberts, Thomas M.

    2008-01-01

    Although several models have been proposed to account for how cytoskeleton polymerization drives protrusion in cell motility, the precise mechanism remains controversial. Here, we show that, in addition to force exerted directly against the membrane by growing filaments, the way elongating filaments pack also contributes to protrusion by generating an expansion of the cytoskeleton gel. Tomography shows that filament packing in the major sperm protein (MSP) -based nematode sperm-motility machinery resembles that observed with rigid rods. Maximum rod-packing density decreases dramatically as the rods lengthen. Therefore, as filaments elongate, the cytoskeleton gel expands to accommodate their packing less densely. This volume expansion combines with polymerization to drive protrusion. Consistent with this hypothesis, an engineered MSP mutant that generates shorter filaments shows higher filament-packing density and slower movement. PMID:18385381

  20. Cytoskeletal architecture and cell motility remain unperturbed in mouse embryonic fibroblasts from Plk3 knockout mice

    PubMed Central

    Michel, Daniel R; Mun, Kyu-Shik; Ho, Chia-Chi

    2016-01-01

    Polo-like kinase 3 (Plk3) is best known for its involvement in cell cycle checkpoint regulation following exposure to cytotoxicants or induction of DNA damage. Yet, Plk3 has also been implicated in roles beyond those of cellular responses to DNA damage. Here, we have investigated the proposition, suggested by the Plk literature, that Plk3 regulates cytoskeletal architecture and cell functions mediated by the cytoskeleton. To this end, we have assayed mouse embryonic fibroblasts (MEFs) generated from both Plk3 knockout and wild-type mice. In particular, we asked whether Plk3 is involved in actin fiber and microtubule integrity, cell migration, cell attachment, and/or cell invasion. Our results demonstrate that functional Plk3 is not critical for the regulation of cytoskeletal integrity, cell morphology, cell adhesion, or motility in MEFs. PMID:26843517

  1. Choreography of Cell Motility and Interaction Dynamics Imaged by Two-Photon Microscopy in Lymphoid Organs

    PubMed Central

    Cahalan, Michael D.; Parker, Ian

    2009-01-01

    The immune system is the most diffuse cellular system in the body. Accordingly, long-range migration of cells and short-range communication by local chemical signaling and by cell-cell contacts are vital to the control of an immune response. Cellular homing and migration within lymphoid organs, antigen recognition, and cell signaling and activation are clearly vital during an immune response, but these events had not been directly observed in vivo until recently. Introduced to the field of immunology in 2002, two-photon microscopy is the method of choice for visualizing living cells deep within native tissue environments, and it is now revealing an elegant cellular choreography that underlies the adaptive immune response to antigen challenge. We review cellular dynamics and molecular factors that contribute to basal motility of lymphocytes in the lymph node and cellular interactions leading to antigen capture and recognition, T cell activation, B cell activation, cytolytic effector function, and antibody production. PMID:18173372

  2. A novel serotonin-secreting cell type regulates ciliary motility in the mucociliary epidermis of Xenopus tadpoles.

    PubMed

    Walentek, Peter; Bogusch, Susanne; Thumberger, Thomas; Vick, Philipp; Dubaissi, Eamon; Beyer, Tina; Blum, Martin; Schweickert, Axel

    2014-04-01

    The embryonic skin of Xenopus tadpoles serves as an experimental model system for mucociliary epithelia (MCE) such as the human airway epithelium. MCEs are characterized by the presence of mucus-secreting goblet and multiciliated cells (MCCs). A third cell type, ion-secreting cells (ISCs), is present in the larval skin as well. Synchronized beating of MCC cilia is required for directional transport of mucus. Here we describe a novel cell type in the Xenopus laevis larval epidermis, characterized by serotonin synthesis and secretion. It is termed small secretory cell (SSC). SSCs are detectable at early tadpole stages, unlike MCCs and ISCs, which are specified at early neurulation. Subcellularly, serotonin was found in large, apically localized vesicle-like structures, which were entirely shed into the surrounding medium. Pharmacological inhibition of serotonin synthesis decreased the velocity of cilia-driven fluid flow across the skin epithelium. This effect was mediated by serotonin type 3 receptor (Htr3), which was expressed in ciliated cells. Knockdown of Htr3 compromised flow velocity by reducing the ciliary motility of MCCs. SSCs thus represent a distinct and novel entity of the frog tadpole MCE, required for ciliary beating and mucus transport across the larval skin. The identification and characterization of SSCs consolidates the value of the Xenopus embryonic skin as a model system for human MCEs, which have been known for serotonin-dependent regulation of ciliary beat frequency. PMID:24598162

  3. The Membrane-associated Protein, Supervillin, Accelerates F-actin-dependent Rapid Integrin Recycling and Cell Motility

    PubMed Central

    Fang, Zhiyou; Takizawa, Norio; Wilson, Korey A.; Smith, Tara C.; Delprato, Anna; Davidson, Michael W.; Lambright, David G.; Luna, Elizabeth J.

    2010-01-01

    In migrating cells, the cytoskeleton coordinates signal transduction and re-distributions of transmembrane proteins, including integrins and growth factor receptors. Supervillin is an F-actin- and myosin II-binding protein that tightly associates with signaling proteins in cholesterol-rich, “lipid raft” membrane microdomains. We show here that supervillin also can localize with markers for early and sorting endosomes (EE/SE) and with overexpressed components of the Arf6 recycling pathway in the cell periphery. Supervillin tagged with the photoswitchable fluorescent protein, tdEos, moves both into and away from dynamic structures resembling podosomes at the basal cell surface. Rapid integrin recycling from EE/SE is inhibited in supervillin-knockdown cells, but the rates of integrin endocytosis and recycling from the perinuclear recycling center (PNRC) are unchanged. A lack of synergy between supervillin knockdown and the actin filament barbed-end inhibitor, cytochalasin D, suggests that both treatments affect actin-dependent rapid recycling. Supervillin also enhances signaling from the epidermal growth factor receptor (EGFR) to extracellular signal-regulated kinases 1 and 2 (ERK) and increases the velocity of cell translocation. These results suggest that supervillin, F-actin, and associated proteins may coordinate a rapid, basolateral membrane recycling pathway that contributes to ERK signaling and actin-based cell motility. PMID:20331534

  4. Vimentin and post-translational modifications in cell motility during cancer - a review.

    PubMed

    Shi, A-M; Tao, Z-Q; Li, R; Wang, Y-Q; Wang, X; Zhao, J

    2016-07-01

    The post-translational modifications (PTMs) are defined as the covalent modification or enzymatic modification of proteins during or after protein biosynthesis. Proteins are synthesized by ribosomes translating mRNA into polypeptide chains, which may then undergo PTM to form the mature protein product. PTMs are important components in cell signaling. Moreover, it is a known fact that PTM regulation offers an immense array and depth of regulatory possibilities. The present review article will focus on their possible role in cancer cell motility with special reference to vimentin, an intermediate filament (IF), as the later is an important process responsible for life-threatening state viz. cancer metastasis. PMID:27383311

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

    PubMed

    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. PMID:25625899

  6. Live cell imaging of neuronal growth cone motility and guidance in vitro

    PubMed Central

    Suter, Daniel M.

    2013-01-01

    Summary The neuronal growth cone, a highly motile structure at the tip of neuronal processes, is an excellent model system for studying directional cell movements. While biochemical and genetic approaches unveiled molecular interactions between ligand, receptor, signaling and cytoskeleton-associated proteins controlling axonal growth and guidance, in vitro live cell imaging has emerged as a crucial approach for dissecting cellular mechanisms of growth cone motility and guidance. Important insights into these mechanisms have been gained from studies using the large growth cones elaborated by Aplysia californica neurons, an outstanding model system for live cell imaging for a number of reasons. Identified neurons can be isolated and imaged at room temperature. Aplysia growth cones are 5–10 times larger than growth cones from other species, making them suitable for quantitative high-resolution imaging of cytoskeletal protein dynamics and biophysical approaches. Lastly, protein, RNA, fluorescent probes and small molecules can be microinjected into the neuronal cell body for localization and functional studies. The following chapter describes culturing of Aplysia bag cell neurons, live cell imaging of neuronal growth cones using differential interference contrast and fluorescent speckle microscopy as well as the restrained bead interaction assay to induce adhesion-mediated growth cone guidance in vitro. PMID:21748670

  7. 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.

  8. Oligomycin A-induced inhibition of mitochondrial ATP-synthase activity suppresses boar sperm motility and in vitro capacitation achievement without modifying overall sperm energy levels.

    PubMed

    Ramió-Lluch, Laura; Yeste, Marc; Fernández-Novell, Josep M; Estrada, Efrén; Rocha, Luiz; Cebrián-Pérez, José A; Muiño-Blanco, Teresa; Concha, Ilona I; Ramírez, Alfredo; Rodríguez-Gil, Joan E

    2014-01-01

    Incubation of boar spermatozoa in a capacitation medium with oligomycin A, a specific inhibitor of the F0 component of the mitochondrial ATP synthase, induced an immediate and almost complete immobilisation of cells. Oligomycin A also inhibited the ability of spermatozoa to achieve feasible in vitro capacitation (IVC), as measured through IVC-compatible changes in motility patterns, tyrosine phosphorylation levels of the acrosomal p32 protein, membrane fluidity and the ability of spermatozoa to achieve subsequent, progesterone-induced in vitro acrosome exocytosis (IVAE). Both inhibitory effects were caused without changes in the rhythm of O2 consumption, intracellular ATP levels or mitochondrial membrane potential (MMP). IVAE was accompanied by a fast and intense peak in O2 consumption and ATP levels in control spermatozoa. Oligomycin A also inhibited progesterone-induced IVAE as well as the concomitant peaks of O2 consumption and ATP levels. The effect of oligomycin on IVAE was also accompanied by concomitant alterations in the IVAE-induced changes on intracellular Ca(2+) levels and MMP. Our results suggest that the oligomycin A-sensitive mitochondrial ATP-synthase activity is instrumental in the achievement of an adequate boar sperm motion pattern, IVC and IVAE. However, this effect seems not to be linked to changes in the overall maintenance of adequate energy levels in stages other than IVAE.

  9. A microfluidic perfusion platform for cultivation and screening study of motile microalgal cells

    PubMed Central

    Eu, Young-Jae; Park, Hye-Sun; Kim, Dong-Pyo; Wook Hong, Jong

    2014-01-01

    Systematic screening of algal cells is getting huge interest due to their capability of producing lipid-based biodiesel. Here, we introduce a new microfluidic platform composed of an array of perfusion chambers designed for long-term cultivation and preliminary screening of motile microalgal cells through loading and releasing of cells to and from the chambers. The chemical environment in each perfusion chamber was independently controlled for 5 days. The effect of nitrogen-depletion on the lipid production, phototaxis behavior in the absence of Ca2+, and cytotoxic effect of herbicide on microalgal cells was successfully monitored and compared with simultaneous control experiments on the platform. The present methodology could be extended to effective screening of algal cells and various cell lines for the production of biodiesel and other useful chemicals. PMID:24803962

  10. Three dimensional template matching segmentation method for motile cells in 3D+t video sequences.

    PubMed

    Pimentel, J A; Corkidi, G

    2010-01-01

    In this work, we describe a segmentation cell method oriented to deal with experimental data obtained from 3D+t microscopical volumes. The proposed segmentation technique takes advantage of the pattern of appearances exhibited by the objects (cells) from different focal planes, as a result of the object translucent properties and its interaction with light. This information allows us to discriminate between cells and artifacts (dust an other) with equivalent size and shape that are present in the biological preparation. Using a simple correlation criteria, the method matches a 3D video template (extracted from a sample of cells) with the motile cells contained into the biological sample, obtaining a high rate of true positives while discarding artifacts. In this work, our analysis is focused on sea urchin spermatozoa cells but is applicable to many other microscopical structures having the same optical properties. PMID:21096252

  11. STAT3 interacts with Skp2/p27/p21 pathway to regulate the motility and invasion of gastric cancer cells.

    PubMed

    Wei, Zheng; Jiang, Xian; Qiao, Haiquan; Zhai, Bo; Zhang, Lianfeng; Zhang, Qiang; Wu, Yuanhong; Jiang, Hongchi; Sun, Xueying

    2013-04-01

    The interleukin-6 (IL-6)/Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway mediates cell proliferation and migration. S-phase kinase-associated protein-2 (Skp2) catalyzes the ubiquitylation of p27 and p21. Here we investigated that the cross-talk of the two pathways regulates motility and invasion of gastric cancer SGC7901 and MGC803 cells. Both cell lines endogenously secret IL-6, and blockage of IL-6 or JAK2 inhibited the activation of JAK2 and STAT3. Depletion of STAT3 downregulated Skp2 expression, and thereby increased the expression of p27 and p21. The depletion of STAT3 inhibited the ability of cells to migrate and invade, and impaired the cellular cytoskeleton mainly microtubules; while the depletion of p27 partially restored the impaired ability to migrate, and reversed the impaired microfilaments, further inhibited the ability to invade, but had little effect on microtubules and cellular adhering ability of STAT3-depleted cells. STAT3 depletion inhibited the activity of RhoA and the interaction with stathmin, downregulated the expression of pFAK (phosphorylated focal adhesion kinase), acetylated-tubulin, RECK (reversion-inducing-cysteine-rich protein with kazal motifs) and Sp1, upregulated E-cadherin, and reduced the activities of MMP (matrix metalloproteinase)-2 and -9. The depletion of p27 increased RhoA (Ras homolog family member A) activity, upregulated RECK, and downregulated E-cadherin and Sp1 in STAT3-depleted cells. The results indicate that the interaction between STAT3 and Skp2/p27/p21 pathway plays an important role in mediating the motility, migration and invasion of gastric cancer cells, and inhibition of STAT3 may be a useful therapeutic approach for metastasis of gastric cancer, but caution needs to be taken for its effects on Skp2/p27/p21 pathway. PMID:23333463

  12. Motility voltage sensor of the outer hair cell resides within the lateral plasma membrane.

    PubMed Central

    Huang, G; Santos-Sacchi, J

    1994-01-01

    The outer hair cell (OHC) from the organ of Corti is believed to be responsible for the mammal's exquisite sense of hearing. A membrane-based motile response of this cell underlies the initial processing of acoustic energy. The voltage-dependent capacitance of the OHC, possibly reflecting charge movement of the motility voltage sensor, was measured in cells during intracellular dialysis of trypsin under whole cell voltage clamp. Within 10 min after dialysis, light and electron microscopic examination revealed that the subplasmalemmal structures, including the cytoskeletal framework and subsurface cisternae, were disrupted and/or detached from adjacent plasma membrane. Dialysis of heat-inactivated trypsin produced no changes in cell structure. Simultaneous measures of linear and nonlinear membrane capacitance revealed minimal changes, indicating that contributions by subsurface structures to the generation of the nonlinear capacitance are unlikely. This study strongly suggests that voltage-dependent charge movement in the OHC reflects properties of the force generator's voltage sensor and that the sensor/motor resides solely within the lateral plasma membrane. Images PMID:7991617

  13. Superdiffusive cell motility on 2D substrates modeled as a persistent Lévy walk

    NASA Astrophysics Data System (ADS)

    Passucci, Giuseppe; Brasch, Megan E.; Deakin, Nicholas O.; Turner, Christopher E.; Henderson, James H.; Manning, M. Lisa

    Cell motility is an essential part of many biological processes such as morphogenesis, wound healing and tumorigenesis. We quantified cell motility by tracking mouse fibroblast and human breast carcinoma nuclei to construct cell trajectories. The mean-squared displacement of these trajectories reveals that cell motion is super diffusive, where displacements scale faster than t 1 / 2 in all directions. Existing self-propelled particle (SPP) models that do not explicitly incorporate ensemble heterogeneity are unable to predict this super-diffusive behavior. Therefore we developed a run-and-tumble SPP model with Levy distributed run times that captures observed super-diffusive behavior in the mean-squared displacement as well as scaling collapse exponents of displacement probability distributions which match those of mouse fibroblast and human breast carcinoma cell trajectories. We additionally introduced small fluctuations in particle orientation during runs, which generates a crossover from super-diffusive to diffusive dynamics at a very long times. This timescale can be extracted in experiments from the velocity auto-correlation function, allowing us to explicitly test this model prediction.

  14. Ras activation mediates WISP-1-induced increases in cell motility and matrix metalloproteinase expression in human osteosarcoma.

    PubMed

    Wu, Chien-Lin; Tsai, Hsiao-Chi; Chen, Zhen-Wei; Wu, Chi-Ming; Li, Te-Mao; Fong, Yi-Chin; Tang, Chih-Hsin

    2013-12-01

    WISP-1 is a cysteine-rich protein that belongs to the CCN (Cyr61, CTGF, Nov) family of matrix cellular proteins. Osteosarcoma is a type of highly malignant tumor with a potent capacity to invade locally and cause distant metastasis. However, the effect of WISP-1 on migration activity in human osteosarcoma cells is mostly unknown. In this study, we first found that the expression of WISP-1 in osteosarcoma patients was significantly higher than that in normal bone and corrected with tumor stage. Exogenous treatment of osteosarcoma cells with WISP-1 promoted cell motility and matrix metalloproteinase (MMP)-2 and MMP-9 expression. In addition, the Ras and Raf-1 inhibitor or siRNA abolished WISP-1-induced cell migration and MMP expression. On the other hand, activation of the Ras, Raf-1, MEK, ERK, and NF-κB signaling pathway after WISP-1 treatment was demonstrated, and WISP-1-induced expression of MMPs and migration activity were inhibited by the specific inhibitor, and mutant of MEK, ERK, and NF-κB cascades. Taken together, our results indicated that WISP-1 enhances the migration of osteosarcoma cells by increasing MMP-2 and MMP-9 expression through the integrin receptor, Ras, Raf-1, MEK, ERK, and NF-κB signal transduction pathway.

  15. A genome-wide RNAi screen for microtubule bundle formation and lysosome motility regulation in Drosophila S2 cells

    PubMed Central

    Jolly, Amber L.; Luan, Chi-Hao; Dusel, Brendon E.; Dunne, Sara Fernandez; Winding, Michael; Dixit, Vishrut J.; Robins, Chloe; Saluk, Jennifer L.; Logan, David J.; Carpenter, Anne E.; Sharma, Manu; Dean, Deborah; Cohen, Andrew R.; Gelfand, Vladimir I.

    2016-01-01

    Summary Long-distance intracellular transport of organelles, mRNA, and proteins (“cargo”) occurs along the microtubule cytoskeleton by the action of kinesin and dynein motor proteins; the vast network of factors involved in regulating intracellular cargo transport are still unknown. We capitalize on the Drosophila melanogaster S2 model cell system to monitor lysosome transport along microtubule bundles, which require enzymatically active kinesin-1 motor protein for their formation. We use an automated tracking program and a naïve Bayesian classifier for the multivariate motility data to analyze 15,683 gene phenotypes, and find 98 proteins involved in regulating lysosome motility along microtubules and 48 involved in the formation of microtubule filled processes in S2 cells. We identify innate immunity genes, ion channels and signaling proteins having a role in lysosome motility regulation, and find an unexpected relationship between the dynein motor, Rab7a and lysosome motility regulation. PMID:26774481

  16. 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

  17. Progesterone inhibits mast cell secretion.

    PubMed

    Vasiadi, M; Kempuraj, D; Boucher, W; Kalogeromitros, D; Theoharides, T C

    2006-01-01

    Mast cells are involved in allergic reactions, where they secrete numerous vasoactive, inflammatory and nociceptive mediators in response to immunoglobulin E (IgE) and antigen. However, they have also been implicated in inflammatory conditions, such as painful bladder syndrome/interstitial cystitis (PBS/IC), irritable bowel syndrome (IBS) and migraines, all of which occur more often in women and are exacerbated during ovulation, but are suppressed during pregnancy. Mast cells express high affinity estrogen receptors and estradiol augments their secretion, while tamoxifen inhibits it. Here we report that progesterone (100 nM), but not the structurally related cholesterol, inhibits histamine secretion from purified rat peritoneal mast cells stimulated immunologically or by substance P (SP), an effect also documented by electron microscopy. These results suggest that mast cell secretion may be regulated by progesterone and may explain the reduced symptoms of certain inflammatory conditions during pregnancy.

  18. CXCR4/CXCL12 signaling impacts enamel progenitor cell proliferation and motility in the dental stem cell niche.

    PubMed

    Yokohama-Tamaki, Tamaki; Otsu, Keishi; Harada, Hidemitsu; Shibata, Shunichi; Obara, Nobuko; Irie, Kazuharu; Taniguchi, Akiyoshi; Nagasawa, Takashi; Aoki, Kazunari; Caliari, Steven R; Weisgerber, Daniel W; Harley, Brendan A C

    2015-12-01

    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.

  19. Live Imaging of Influenza Infection of the Trachea Reveals Dynamic Regulation of CD8+ T Cell Motility by Antigen

    PubMed Central

    Lambert Emo, Kris; Hyun, Young-min; Barilla, Christopher; Gerber, Scott; Fowell, Deborah; Kim, Minsoo

    2016-01-01

    During a primary influenza infection, cytotoxic CD8+ T cells need to infiltrate the infected airways and engage virus-infected epithelial cells. The factors that regulate T cell motility in the infected airway tissue are not well known. To more precisely study T cell infiltration of the airways, we developed an experimental model system using the trachea as a site where live imaging can be performed. CD8+ T cell motility was dynamic with marked changes in motility on different days of the infection. In particular, significant changes in average cell velocity and confinement were evident on days 8–10 during which the T cells abruptly but transiently increase velocity on day 9. Experiments to distinguish whether infection itself or antigen affect motility revealed that it is antigen, not active infection per se that likely affects these changes as blockade of peptide/MHC resulted in increased velocity. These observations demonstrate that influenza tracheitis provides a robust experimental foundation to study molecular regulation of T cell motility during acute virus infection. PMID:27644089

  20. Live Imaging of Influenza Infection of the Trachea Reveals Dynamic Regulation of CD8+ T Cell Motility by Antigen.

    PubMed

    Lambert Emo, Kris; Hyun, Young-Min; Reilly, Emma; Barilla, Christopher; Gerber, Scott; Fowell, Deborah; Kim, Minsoo; Topham, David J

    2016-09-01

    During a primary influenza infection, cytotoxic CD8+ T cells need to infiltrate the infected airways and engage virus-infected epithelial cells. The factors that regulate T cell motility in the infected airway tissue are not well known. To more precisely study T cell infiltration of the airways, we developed an experimental model system using the trachea as a site where live imaging can be performed. CD8+ T cell motility was dynamic with marked changes in motility on different days of the infection. In particular, significant changes in average cell velocity and confinement were evident on days 8-10 during which the T cells abruptly but transiently increase velocity on day 9. Experiments to distinguish whether infection itself or antigen affect motility revealed that it is antigen, not active infection per se that likely affects these changes as blockade of peptide/MHC resulted in increased velocity. These observations demonstrate that influenza tracheitis provides a robust experimental foundation to study molecular regulation of T cell motility during acute virus infection. PMID:27644089

  1. Fibroblasts Cultured on Nanowires Exhibit Low Motility, Impaired Cell Division, and DNA Damage

    PubMed Central

    Persson, Henrik; Købler, Carsten; Mølhave, Kristian; Samuelson, Lars; Tegenfeldt, Jonas O; Oredsson, Stina; Prinz, Christelle N

    2013-01-01

    Nanowires are commonly used as tools for interfacing living cells, acting as biomolecule-delivery vectors or electrodes. It is generally assumed that the small size of the nanowires ensures a minimal cellular perturbation, yet the effects of nanowires on cell migration and proliferation remain largely unknown. Fibroblast behaviour on vertical nanowire arrays is investigated, and it is shown that cell motility and proliferation rate are reduced on nanowires. Fibroblasts cultured on long nanowires exhibit failed cell division, DNA damage, increased ROS content and respiration. Using focused ion beam milling and scanning electron microscopy, highly curved but intact nuclear membranes are observed, showing no direct contact between the nanowires and the DNA. The nanowires possibly induce cellular stress and high respiration rates, which trigger the formation of ROS, which in turn results in DNA damage. These results are important guidelines to the design and interpretation of experiments involving nanowire-based transfection and electrical characterization of living cells. PMID:23813871

  2. Fibroblasts cultured on nanowires exhibit low motility, impaired cell division, and DNA damage.

    PubMed

    Persson, Henrik; Købler, Carsten; Mølhave, Kristian; Samuelson, Lars; Tegenfeldt, Jonas O; Oredsson, Stina; Prinz, Christelle N

    2013-12-01

    Nanowires are commonly used as tools for interfacing living cells, acting as biomolecule-delivery vectors or electrodes. It is generally assumed that the small size of the nanowires ensures a minimal cellular perturbation, yet the effects of nanowires on cell migration and proliferation remain largely unknown. Fibroblast behaviour on vertical nanowire arrays is investigated, and it is shown that cell motility and proliferation rate are reduced on nanowires. Fibroblasts cultured on long nanowires exhibit failed cell division, DNA damage, increased ROS content and respiration. Using focused ion beam milling and scanning electron microscopy, highly curved but intact nuclear membranes are observed, showing no direct contact between the nanowires and the DNA. The nanowires possibly induce cellular stress and high respiration rates, which trigger the formation of ROS, which in turn results in DNA damage. These results are important guidelines to the design and interpretation of experiments involving nanowire-based transfection and electrical characterization of living cells.

  3. An extracellular matrix, calmodulin-binding protein from Dictyostelium with EGF-like repeats that enhance cell motility.

    PubMed

    Suarez, Andres; Huber, Robert J; Myre, Michael A; O'Day, Danton H

    2011-07-01

    CyrA is a novel cysteine-rich protein with four EGFL repeats that was isolated using the calmodulin (CaM) binding overlay technique (CaMBOT), suggesting it is a CaM-binding protein (CaMBP). The full-length 63kDa cyrA is cleaved into two major C-terminal fragments, cyrA-C45 and cyrA-C40. A putative CaM-binding domain was detected and both CaM-agarose binding and CaM immunoprecipitation verified that cyrA-C45 and cyrA-C40 each bind to CaM in both a Ca(2+)-dependent and -independent manner. cyrA-C45 was present continuously throughout growth and development but was secreted at high levels during the multicellular slug stage of Dictyostelium development. At this time, cyrA localizes to the extracellular matrix (ECM). ECM purification verified the presence of cyrA-C45. An 18 amino acid peptide (DdEGFL1) from the first EGFL repeat sequence of cyrA (EGFL1) that is present in both cyrA-C45 and -C40 enhances both random cell motility and cAMP-mediated chemotaxis. Here we reveal that the dose-dependent enhancement of motility by DdEGFL1 is related to the time of cell starvation. Addition of DdEGFL1 also inhibits cyrA proteolysis. The status of cyrA as an extracellular CaMBP was further clarified by the demonstration that CaM is secreted during development. Antagonism of CaM with W7 resulted in enhanced cyrA proteolysis suggesting a functional role for extracellular CaM in protecting CaMBPs from proteolysis. cyrA is the first extracellular CaMBP identified in Dictyostelium and since it is an ECM protein with EGF-like repeats that enhance cell motility and it likely also represents the first matricellular protein identified in a lower eukaryote. PMID:21402150

  4. Natural plant products inhibits growth and alters the swarming motility, biofilm formation, and expression of virulence genes in enteroaggregative and enterohemorrhagic Escherichia coli.

    PubMed

    García-Heredia, Alam; García, Santos; Merino-Mascorro, José Ángel; Feng, Peter; Heredia, Norma

    2016-10-01

    The purpose of this study was to determine the effects of plant products on the growth, swarming motility, biofilm formation and virulence gene expression in enterohemorrhagic Escherichia coli O157:H7 and enteroaggregative E. coli strain 042 and a strain of O104:H4 serotype. Extracts of Lippia graveolens and Haematoxylon brassiletto, and carvacrol, brazilin were tested by an antimicrobial microdilution method using citral and rifaximin as controls. All products showed bactericidal activity with minimal bactericidal concentrations ranging from 0.08 to 8.1 mg/ml. Swarming motility was determined in soft LB agar. Most compounds reduced swarming motility by 7%-100%; except carvacrol which promoted motility in two strains. Biofilm formation studies were done in microtiter plates. Rifaximin inhibited growth and reduced biofilm formation, but various concentrations of other compounds actually induced biofilm formation. Real time PCR showed that most compounds decreased stx2 expression. The expression of pic and rpoS in E. coli 042 were suppressed but in E. coli O104:H4 they varied depending on compounds. In conclusion, these extracts affect E. coli growth, swarming motility and virulence gene expression. Although these compounds were bactericidal for pathogenic E. coli, sublethal concentrations had varied effects on phenotypic and genotypic traits, and some increased virulence gene expression.

  5. Calcium Signalling Triggered by NAADP in T Cells Determines Cell Shape and Motility During Immune Synapse Formation

    PubMed Central

    Nebel, Merle; Zhang, Bo; Odoardi, Francesca; Flügel, Alexander; Potter, Barry V. L.; Guse, Andreas H.

    2016-01-01

    Nicotinic acid adenine dinucleotide phosphate (NAADP) has been implicated as an initial Ca2+ trigger in T cell Ca2+ signalling, but its role in formation of the immune synapse in CD4+ effector T cells has not been analysed. CD4+ T cells are activated by the interaction with peptide-MHCII complexes on the surface of antigen-presenting cells. Establishing a two-cell system including primary rat CD4+ T cells specific for myelin basic protein and rat astrocytes enabled us to mirror this activation process in vitro and to analyse Ca2+ signalling, cell shape changes and motility in T cells during formation and maintenance of the immune synapse. After immune synapse formation, T cells showed strong, antigen-dependent increases in free cytosolic calcium concentration ([Ca2+]i). Analysis of cell shape and motility revealed rounding and immobilization of T cells depending on the amplitude of the Ca2+ signal. NAADP-antagonist BZ194 effectively blocked Ca2+ signals in T cells evoked by the interaction with antigen-presenting astrocytes. BZ194 reduced the percentage of T cells showing high Ca2+ signals thereby supporting the proposed trigger function of NAADP for global Ca2+ signalling. Taken together, the NAADP signalling pathway is further confirmed as a promising target for specific pharmacological intervention to modulate T cell activation. PMID:27747143

  6. Effects of Adhesion Dynamics and Substrate Compliance on the Shape and Motility of Crawling Cells

    PubMed Central

    Ziebert, Falko; Aranson, Igor S.

    2013-01-01

    Computational modeling of eukaryotic cells moving on substrates is an extraordinarily complex task: many physical processes, such as actin polymerization, action of motors, formation of adhesive contacts concomitant with both substrate deformation and recruitment of actin etc., as well as regulatory pathways are intertwined. Moreover, highly nontrivial cell responses emerge when the substrate becomes deformable and/or heterogeneous. Here we extended a computational model for motile cell fragments, based on an earlier developed phase field approach, to account for explicit dynamics of adhesion site formation, as well as for substrate compliance via an effective elastic spring. Our model displays steady motion vs. stick-slip transitions with concomitant shape oscillations as a function of the actin protrusion rate, the substrate stiffness, and the rates of adhesion. Implementing a step in the substrate’s elastic modulus, as well as periodic patterned surfaces exemplified by alternating stripes of high and low adhesiveness, we were able to reproduce the correct motility modes and shape phenomenology found experimentally. We also predict the following nontrivial behavior: the direction of motion of cells can switch from parallel to perpendicular to the stripes as a function of both the adhesion strength and the width ratio of adhesive to non-adhesive stripes. PMID:23741334

  7. From individual cell motility to collective behaviors: insights from a prokaryote, Myxococcus xanthus.

    PubMed

    Zhang, Yong; Ducret, Adrien; Shaevitz, Joshua; Mignot, Tâm

    2012-01-01

    In bird flocks, fish schools, and many other living organisms, regrouping among individuals of the same kin is frequently an advantageous strategy to survive, forage, and face predators. However, these behaviors are costly because the community must develop regulatory mechanisms to coordinate and adapt its response to rapid environmental changes. In principle, these regulatory mechanisms, involving communication between individuals, may also apply to cellular systems which must respond collectively during multicellular development. Dissecting the mechanisms at work requires amenable experimental systems, for example, developing bacteria. Myxococcus xanthus, a Gram-negative delatproteobacterium, is able to coordinate its motility in space and time to swarm, predate, and grow millimeter-size spore-filled fruiting bodies. A thorough understanding of the regulatory mechanisms first requires studying how individual cells move across solid surfaces and control their direction of movement, which was recently boosted by new cell biology techniques. In this review, we describe current molecular knowledge of the motility mechanism and its regulation as a lead-in to discuss how multicellular cooperation may have emerged from several layers of regulation: chemotaxis, cell-cell signaling, and the extracellular matrix. We suggest that Myxococcus is a powerful system to investigate collective principles that may also be relevant to other cellular systems.

  8. OXPHOS dysfunction regulates integrin-β1 modifications and enhances cell motility and migration.

    PubMed

    Nunes, Joana B; Peixoto, Joana; Soares, Paula; Maximo, Valdemar; Carvalho, Sandra; Pinho, Salome S; Vieira, Andre F; Paredes, Joana; Rego, Ana C; Ferreira, Ildete L; Gomez-Lazaro, Maria; Sobrinho-Simoes, Manuel; Singh, Keshav K; Lima, Jorge

    2015-04-01

    Mitochondria are central organelles for cellular metabolism. In cancer cells, mitochondrial oxidative phosphorylation (OXPHOS) dysfunction has been shown to promote migration, invasion, metastization and apoptosis resistance. With the purpose of analysing the effects of OXPHOS dysfunction in cancer cells and the molecular players involved, we generated cybrid cell lines harbouring either wild-type (WT) or mutant mitochondrial DNA (mtDNA) [tRNAmut cybrids, which harbour the pathogenic A3243T mutation in the leucine transfer RNA gene (tRNAleu)]. tRNAmut cybrids exhibited lower oxygen consumption and higher glucose consumption and lactate production than WT cybrids. tRNAmut cybrids displayed increased motility and migration capacities, which were associated with altered integrin-β1 N-glycosylation, in particular with higher levels of β-1,6-N-acetylglucosamine (GlcNAc) branched N-glycans. This integrin-β1 N-glycosylation pattern was correlated with higher levels of membrane-bound integrin-β1 and also with increased binding to fibronectin. When cultured in vitro, tRNAmut cybrids presented lower growth rate than WT cybrids, however, when injected in nude mice, tRNAmut cybrids produced larger tumours and showed higher metastatic potential than WT cybrids. We conclude that mtDNA-driven OXPHOS dysfunction correlates with increased motility and migration capacities, through a mechanism that may involve the cross talk between cancer cell mitochondria and the extracellular matrix.

  9. Nodal signaling regulates endodermal cell motility and actin dynamics via Rac1 and Prex1

    PubMed Central

    Housley, Michael P.; Weiner, Orion D.

    2012-01-01

    Embryo morphogenesis is driven by dynamic cell behaviors, including migration, that are coordinated with fate specification and differentiation, but how such coordination is achieved remains poorly understood. During zebrafish gastrulation, endodermal cells sequentially exhibit first random, nonpersistent migration followed by oriented, persistent migration and finally collective migration. Using a novel transgenic line that labels the endodermal actin cytoskeleton, we found that these stage-dependent changes in migratory behavior correlated with changes in actin dynamics. The dynamic actin and random motility exhibited during early gastrulation were dependent on both Nodal and Rac1 signaling. We further identified the Rac-specific guanine nucleotide exchange factor Prex1 as a Nodal target and showed that it mediated Nodal-dependent random motility. Reducing Rac1 activity in endodermal cells caused them to bypass the random migration phase and aberrantly contribute to mesodermal tissues. Together, our results reveal a novel role for Nodal signaling in regulating actin dynamics and migration behavior, which are crucial for endodermal morphogenesis and cell fate decisions. PMID:22945937

  10. A novel GRK2/HDAC6 interaction modulates cell spreading and motility

    PubMed Central

    Lafarga, Vanesa; Aymerich, Ivette; Tapia, Olga; Mayor, Federico; Penela, Petronila

    2012-01-01

    Cell motility and adhesion involves dynamic microtubule (MT) acetylation/deacetylation, a process regulated by enzymes as HDAC6, a major cytoplasmic α-tubulin deacetylase. We identify G protein-coupled receptor kinase 2 (GRK2) as a key novel stimulator of HDAC6. GRK2, which levels inversely correlate with the extent of α-tubulin acetylation in epithelial cells and fibroblasts, directly associates with and phosphorylates HDAC6 to stimulate α-tubulin deacetylase activity. Remarkably, phosphorylation of GRK2 itself at S670 specifically potentiates its ability to regulate HDAC6. GRK2 and HDAC6 colocalize in the lamellipodia of migrating cells, leading to local tubulin deacetylation and enhanced motility. Consistently, cells expressing GRK2-K220R or GRK2-S670A mutants, unable to phosphorylate HDAC6, exhibit highly acetylated cortical MTs and display impaired migration and protrusive activity. Finally, we find that a balanced, GRK2/HDAC6-mediated regulation of tubulin acetylation differentially modulates the early and late stages of cellular spreading. This novel GRK2/HDAC6 functional interaction may have important implications in pathological contexts. PMID:22193721

  11. Human Schlafen 5 (SLFN5) Is a Regulator of Motility and Invasiveness of Renal Cell Carcinoma Cells

    PubMed Central

    Sassano, Antonella; Mavrommatis, Evangelos; Arslan, Ahmet Dirim; Kroczynska, Barbara; Beauchamp, Elspeth M.; Khuon, Satya; Chew, Ten-Leong; Green, Kathleen J.; Munshi, Hidayatullah G.; Verma, Amit K.

    2015-01-01

    We provide evidence that human SLFN5, an interferon (IFN)-inducible member of the Schlafen (SLFN) family of proteins, exhibits key roles in controlling motility and invasiveness of renal cell carcinoma (RCC) cells. Our studies define the mechanism by which this occurs, demonstrating that SLFN5 negatively controls expression of the matrix metalloproteinase 1 gene (MMP-1), MMP-13, and several other genes involved in the control of malignant cell motility. Importantly, our data establish that SLFN5 expression correlates with a better overall survival in a large cohort of patients with RCC. The inverse relationship between SLFN5 expression and RCC aggressiveness raises the possibility of developing unique therapeutic approaches in the treatment of RCC, by modulating SLFN5 expression. PMID:26012550

  12. Cyclin-Dependent Kinase 5 (CDK5) Controls Melanoma Cell Motility, Invasiveness, and Metastatic Spread—Identification of a Promising Novel therapeutic target1

    PubMed Central

    Bisht, Savita; Nolting, Jens; Schütte, Ute; Haarmann, Jens; Jain, Prashi; Shah, Dhruv; Brossart, Peter; Flaherty, Patrick; Feldmann, Georg

    2015-01-01

    Despite considerable progress in recent years, the overall prognosis of metastatic malignant melanoma remains poor, and curative therapeutic options are lacking. Therefore, better understanding of molecular mechanisms underlying melanoma progression and metastasis, as well as identification of novel therapeutic targets that allow inhibition of metastatic spread, are urgently required. The current study provides evidence for aberrant cyclin-dependent kinase 5 (CDK5) activation in primary and metastatic melanoma lesions by overexpression of its activator protein CDK5R1/p35. Moreover, using melanoma in vitro model systems, shRNA-mediated inducible knockdown of CDK5 was found to cause marked inhibition of cell motility, invasiveness, and anchorage-independent growth, while at the same time net cell growth was not affected. In vivo, CDK5 knockdown inhibited growth of orthotopic xenografts as well as formation of lung and liver colonies in xenogenic injection models mimicking systemic metastases. Inhibition of lung metastasis was further validated in a syngenic murine melanoma model. CDK5 knockdown was accompanied by dephosphorylation and overexpression of caldesmon, and concomitant caldesmon knockdown rescued cell motility and proinvasive phenotype. Finally, it was found that pharmacological inhibition of CDK5 activity by means of roscovitine as well as by a novel small molecule CDK5-inhibitor, N-(5-isopropylthiazol-2-yl)-3-phenylpropanamide, similarly caused marked inhibition of invasion/migration, colony formation, and anchorage-independent growth of melanoma cells. Thus, experimental data presented here provide strong evidence for a crucial role of aberrantly activated CDK5 in melanoma progression and metastasis and establish CDK5 as promising target for therapeutic intervention. PMID:26310376

  13. Reduced Parasite Motility and Micronemal Protein Secretion by a p38 MAPK Inhibitor Leads to a Severe Impairment of Cell Invasion by the Apicomplexan Parasite Eimeria tenella

    PubMed Central

    Bussière, Françoise I.; Le Vern, Yves; Niepceron, Alisson; Silvestre, Anne; de Sablet, Thibaut; Lacroix-Lamandé, Sonia; Laurent, Fabrice

    2015-01-01

    E. tenella infection is associated with a severe intestinal disease leading to high economic losses in poultry industry. Mitogen activated protein kinases (MAPKs) are implicated in early response to infection and are divided in three pathways: p38, extracellular signal-regulated protein kinase (ERK) and c-Jun N-terminal kinase (JNK). Our objective was to determine the importance of these kinases on cell invasion by E. tenella. We evaluated the effect of specific inhibitors (ERK: PD98059, JNKII: SP600125, p38 MAPK: SB203580) on the invasion of epithelial cells. Incubation of SP600125 and SB203580 with epithelial cells and parasites significantly inhibited cell invasion with the highest degree of inhibition (90%) for SB203580. Silencing of the host p38α MAPK expression by siRNA led to only 20% decrease in cell invasion. In addition, when mammalian epithelial cells were pre-treated with SB203580, and washed prior infection, a 30% decrease in cell invasion was observed. This decrease was overcome when a p38 MAPK activator, anisomycin was added during infection. This suggests an active but limited role of the host p38 MAPK in this process. We next determined whether SB203580 has a direct effect on the parasite. Indeed, parasite motility and secretion of micronemal proteins (EtMIC1, 2, 3 and 5) that are involved in cell invasion were both decreased in the presence of the inhibitor. After chasing the inhibitor, parasite motility and secretion of micronemal proteins were restored and subsequently cell invasion. SB203580 inhibits cell invasion by acting partly on the host cell and mainly on the parasite. PMID:25689363

  14. Slow motility in hair cells of the frog amphibian papilla: Myosin light chain-mediated shape change

    PubMed Central

    Farahbakhsh, Nasser A.; Narins, Peter M.

    2008-01-01

    Using video, fluorescence and confocal microscopy, quantitative analysis and modeling, we investigated intracellular processes mediating the calcium/calmodulin (Ca2+/CaM)-dependent slow motility in hair cells dissociated from the rostral region of amphibian papilla, one of the two auditory organs in frogs. The time course of shape changes in these hair cells during the period of pretreatment with several specific inhibitors, as well as their response to the calcium ionophore, ionomycin, were recorded and compared. These cells respond to ionomycin with a tri-phasic shape change: an initial phase of iso-volumetric length decrease; a period of concurrent shortening and swelling; and the final phase of increase in both length and volume. We found that both the myosin light chain kinase inhibitor, ML-7, and antagonists of the multifunctional Ca2+/CaM-dependent kinases, KN-62 and KN-93, inhibit the iso-volumetric shortening phase of the response to ionomycin. The type 1 protein phosphatase inhibitors, calyculin A and okadaic acid induce minor shortening on their own, but do not significantly alter the phase 1 response. However, they appear to counter effects of the inhibitors of Ca2+/CaM-dependent kinases. We hypothesize that an active actomyosin-based process mediates the iso-volumetric shortening in the frog rostral amphibian papillar hair cells. PMID:18534795

  15. Slow motility in hair cells of the frog amphibian papilla: myosin light chain-mediated shape change.

    PubMed

    Farahbakhsh, Nasser A; Narins, Peter M

    2008-07-01

    Using video, fluorescence and confocal microscopy, quantitative analysis and modeling, we investigated intracellular processes mediating the calcium/calmodulin (Ca(2+)/CaM)-dependent slow motility in hair cells dissociated from the rostral region of amphibian papilla, one of the two auditory organs in frogs. The time course of shape changes in these hair cells during the period of pretreatment with several specific inhibitors, as well as their response to the calcium ionophore, ionomycin, were recorded and compared. These cells respond to ionomycin with a tri-phasic shape change: an initial phase of iso-volumetric length decrease; a period of concurrent shortening and swelling; and the final phase of increase in both length and volume. We found that both the myosin light chain kinase inhibitor, ML-7, and antagonists of the multifunctional Ca(2+)/CaM-dependent kinases, KN-62 and KN-93, inhibit the iso-volumetric shortening phase of the response to ionomycin. The type 1 protein phosphatase inhibitors, calyculin A and okadaic acid induce minor shortening on their own, but do not significantly alter phase 1 response. However, they appear to counter effects of the inhibitors of Ca(2+)/CaM-dependent kinases. We hypothesize that an active actomyosin-based process mediates the iso-volumetric shortening in the frog rostral amphibian papillar hair cells.

  16. Atrazine represses S100A4 gene expression and TPA-induced motility in HepG2 cells.

    PubMed

    Peyre, Ludovic; Zucchini-Pascal, Nathalie; Rahmani, Roger

    2014-03-01

    Atrazine (ATZ) is probably the most widely used herbicide in the world. However there are still many controversies regarding its impacts on human health. Our investigations on the role of pesticides in liver dysfunctions have led us to detect an inhibition of FSP1 expression of 70% at 50μm and around 95% at 500μM of ATZ (p<0.01). This gene encodes the protein S100a4 and is a clinical biomarker of epithelial-mesenchymal transition (EMT), a key step in the metastatic process. Here we investigated the possible effect of ATZ on cell migration and noticed that it prevents the EMT and motility of the HepG2 cells induced by the phorbol ester TPA. ATZ decreases Fak pathway activation but has no effect on the Erk1/2 pathway known to be involved in metastasis in this cell line. These results suggest that ATZ could be involved in cell homeostasis perturbation, potentially through a S100a4-dependant mechanism. PMID:24211529

  17. ADAM17 Promotes Motility, Invasion, and Sprouting of Lymphatic Endothelial Cells.

    PubMed

    Mężyk-Kopeć, Renata; Wyroba, Barbara; Stalińska, Krystyna; Próchnicki, Tomasz; Wiatrowska, Karolina; Kilarski, Witold W; Swartz, Melody A; Bereta, Joanna

    2015-01-01

    Tumor-associated lymphatic vessels actively participate in tumor progression and dissemination. ADAM17, a sheddase for numerous growth factors, cytokines, receptors, and cell adhesion molecules, is believed to promote tumor development, facilitating both tumor cell proliferation and migration, as well as tumor angiogenesis. In this work we addressed the issue of whether ADAM17 may also promote tumor lymphangiogenesis. First, we found that ADAM17 is important for the migratory potential of immortalized human dermal lymphatic endothelial cells (LEC). When ADAM17 was stably silenced in LEC, their proliferation was not affected, but: (i) single-cell motility, (ii) cell migration through a 3D Matrigel/collagen type I matrix, and (iii) their ability to form sprouts in a 3D matrix were significantly diminished. The differences in the cell motility between ADAM17-proficient and ADAM17-silenced cells were eliminated by inhibitors of EGFR and HER2, indicating that ADAM17-mediated shedding of growth factors accounts for LEC migratory potential. Interestingly, ADAM17 depletion affected the integrin surface expression/functionality in LEC. ADAM17-silenced cells adhered to plastic, type I collagen, and fibronectin faster than their ADAM17-proficient counterparts. The difference in adhesion to fibronectin was abolished by a cyclic RGD peptide, emphasizing the involvement of integrins in the process. Using a soluble receptor array, we identified BIG-H3 among several candidate proteins involved in the phenotypic and behavioral changes of LEC upon ADAM17 silencing. In additional assays, we confirmed the increased expression of BIG-H3, as well as TGFβ2 in ADAM17-silenced LEC. The antilymphangiogenic effects of ADAM17 silencing in lymphatic endothelial cells suggest further relevance of ADAM17 as a potential target in cancer therapy. PMID:26176220

  18. ADAM17 Promotes Motility, Invasion, and Sprouting of Lymphatic Endothelial Cells

    PubMed Central

    Stalińska, Krystyna; Próchnicki, Tomasz; Wiatrowska, Karolina; Kilarski, Witold W.; Swartz, Melody A.; Bereta, Joanna

    2015-01-01

    Tumor-associated lymphatic vessels actively participate in tumor progression and dissemination. ADAM17, a sheddase for numerous growth factors, cytokines, receptors, and cell adhesion molecules, is believed to promote tumor development, facilitating both tumor cell proliferation and migration, as well as tumor angiogenesis. In this work we addressed the issue of whether ADAM17 may also promote tumor lymphangiogenesis. First, we found that ADAM17 is important for the migratory potential of immortalized human dermal lymphatic endothelial cells (LEC). When ADAM17 was stably silenced in LEC, their proliferation was not affected, but: (i) single-cell motility, (ii) cell migration through a 3D Matrigel/collagen type I matrix, and (iii) their ability to form sprouts in a 3D matrix were significantly diminished. The differences in the cell motility between ADAM17-proficient and ADAM17-silenced cells were eliminated by inhibitors of EGFR and HER2, indicating that ADAM17-mediated shedding of growth factors accounts for LEC migratory potential. Interestingly, ADAM17 depletion affected the integrin surface expression/functionality in LEC. ADAM17-silenced cells adhered to plastic, type I collagen, and fibronectin faster than their ADAM17-proficient counterparts. The difference in adhesion to fibronectin was abolished by a cyclic RGD peptide, emphasizing the involvement of integrins in the process. Using a soluble receptor array, we identified BIG-H3 among several candidate proteins involved in the phenotypic and behavioral changes of LEC upon ADAM17 silencing. In additional assays, we confirmed the increased expression of BIG-H3, as well as TGFβ2 in ADAM17-silenced LEC. The antilymphangiogenic effects of ADAM17 silencing in lymphatic endothelial cells suggest further relevance of ADAM17 as a potential target in cancer therapy. PMID:26176220

  19. ADAM17 Promotes Motility, Invasion, and Sprouting of Lymphatic Endothelial Cells.

    PubMed

    Mężyk-Kopeć, Renata; Wyroba, Barbara; Stalińska, Krystyna; Próchnicki, Tomasz; Wiatrowska, Karolina; Kilarski, Witold W; Swartz, Melody A; Bereta, Joanna

    2015-01-01

    Tumor-associated lymphatic vessels actively participate in tumor progression and dissemination. ADAM17, a sheddase for numerous growth factors, cytokines, receptors, and cell adhesion molecules, is believed to promote tumor development, facilitating both tumor cell proliferation and migration, as well as tumor angiogenesis. In this work we addressed the issue of whether ADAM17 may also promote tumor lymphangiogenesis. First, we found that ADAM17 is important for the migratory potential of immortalized human dermal lymphatic endothelial cells (LEC). When ADAM17 was stably silenced in LEC, their proliferation was not affected, but: (i) single-cell motility, (ii) cell migration through a 3D Matrigel/collagen type I matrix, and (iii) their ability to form sprouts in a 3D matrix were significantly diminished. The differences in the cell motility between ADAM17-proficient and ADAM17-silenced cells were eliminated by inhibitors of EGFR and HER2, indicating that ADAM17-mediated shedding of growth factors accounts for LEC migratory potential. Interestingly, ADAM17 depletion affected the integrin surface expression/functionality in LEC. ADAM17-silenced cells adhered to plastic, type I collagen, and fibronectin faster than their ADAM17-proficient counterparts. The difference in adhesion to fibronectin was abolished by a cyclic RGD peptide, emphasizing the involvement of integrins in the process. Using a soluble receptor array, we identified BIG-H3 among several candidate proteins involved in the phenotypic and behavioral changes of LEC upon ADAM17 silencing. In additional assays, we confirmed the increased expression of BIG-H3, as well as TGFβ2 in ADAM17-silenced LEC. The antilymphangiogenic effects of ADAM17 silencing in lymphatic endothelial cells suggest further relevance of ADAM17 as a potential target in cancer therapy.

  20. Heparin regulates B6FS cell motility through a FAK/actin cytoskeleton axis

    PubMed Central

    Voudouri, Kallirroi; Nikitovic, Dragana; Berdiaki, Aikaterini; Papachristou, Dionysios J.; Tsiaoussis, John; Spandidos, Demetrios A.; Tsatsakis, Aristides M.; Tzanakakis, George N.

    2016-01-01

    Soft tissue sarcomas are rare, heterogeneous tumors of mesenchymal origin with an aggressive behavior. Heparin is a mixture of heavily sulfated, linear glycosaminoglycan (GAG) chains, which participate in the regulation of various cell biological functions. Heparin is considered to have significant anticancer capabilities, although the mechanisms involved have not been fully defined. In the present study, the effects of unfractionated heparin (UFH) and low-molecular-weight heparin (LMWH) on B6FS fibrosarcoma cell motility were examined. Both preparations of heparin were shown to both enhance B6FS cell adhesion (p<0.01 and p<0.05), and migration (p<0.05), the maximal effect being evident at the concentration of 10 µg/ml. The utilization of FAK-deficient cells demonstrated that the participation of FAK was obligatory for heparin-dependent fibrosarcoma cell adhesion (p<0.05). The results of confocal microscopy indicated that heparin was taken up by the B6FS cells, and that UFH and LMWH induced F-actin polymerization. Heparitinase digestion demonstrated that the endogenous heparan sulfate (HS) chains did not affect the motility of the B6FS cells (p>0.05, not significant). In conclusion, both UFH and LMWH, through a FAK/actin cytoskeleton axis, promoted the adhesion and migration of B6FS fibrosarcoma cells. Thus, our findings indicate that the responsiveness of fibrosarcoma cells to the exogenous heparin/HS content of the cancer microenvironment may play a role in their ability to become mobile and metastasize. PMID:27572115

  1. Picropodophyllin and sorafenib synergistically suppress the proliferation and motility of hepatocellular carcinoma cells

    PubMed Central

    TOMIZAWA, MINORU; SHINOZAKI, FUMINOBU; MOTOYOSHI, YASUFUMI; SUGIYAMA, TAKAO; YAMAMOTO, SHIGENORI; SUEISHI, MAKOTO

    2014-01-01

    Resistance is one limitation of sorafenib in the treatment of hepatocellular carcinoma (HCC). Insulin-like growth factor-1 receptor (IGF-1R) is involved in cancer cell proliferation. To assess the potential synergistic antitumor effects of picropodophyllin (PPP), an IGF-1R inhibitor, HLF and PLC/PRL/5, HCC cells were treated with PPP alone or PPP in combination with sorafenib, a multikinase inhibitor. Normal human umbilical vein endothelial cells (HUVECs) were also used to analyze the antiangiogenic effects of the drugs. HCC cells and HUVECs were cultured on 96-well plates, and then treated with PPP, with and without the addition of sorafenib. A 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium inner salt assay and hematoxylin and eosin staining were then performed 48 h later. The HCC cells were also analyzed using scratch assays and hematoxylin and eosin staining after 48 h. The proliferation of HLF, PLC/PRF/5 and HUVEC cells was suppressed by the combination of 0.2 μM PPP and 3 μM sorafenib more effectively than by 10 μM sorafenib alone. The motility of HLF and PLC/PRF/5 cells was also suppressed to a greater extent with the combination of PPP at 0.2 μM and sorafenib at 3 μM than with sorafenib at 10 μM alone. The cells that had been treated with 0.2 μM PPP and 3 μM sorafenib also exhibited pyknotic nuclei, which is characteristic of apoptosis. In conclusion, PPP enhanced sorafenib-mediated suppression of proliferation and motility in HCC cells. Therefore, the combination of PPP and sorafenib may exert antitumor and antiangiogenic effects. PMID:25289088

  2. Smad4-dependent TGF-β Signaling Suppresses RON Receptor Tyrosine Kinase-dependent Motility and Invasion of Pancreatic Cancer Cells*

    PubMed Central

    Zhao, Shujie; Ammanamanchi, Sudhakar; Brattain, Michael; Cao, Lin; Thangasamy, Amalraj; Wang, Jing; Freeman, James W.

    2008-01-01

    Transforming growth factorβ (TGF-β) signals through Smad-dependent and Smad-independent pathways. However, Smad signaling is altered by allelic deletion or intragenic mutation of the Smad4 gene in more than half of pancreatic ductal adenocarcinomas. We show here that loss of Smad4-dependent signaling leads to aberrant expression of RON, a phosphotyrosine kinase receptor, and that signaling by RON cooperates with Smad4-independent TGF-β signaling to promote cell motility and invasion. Restoring Smad4 expression in a pancreatic ductal adenocarcinoma cell line that is deficient in Smad4 repressed RON expression. Conversely, small interference RNA knock down of Smad4 or blocking TGF-β signaling with a TGF-β type I receptor kinase inhibitor in Smad4-intact cell lines induced RON expression. TGF-β-induced motility and invasion were inhibited in cells that express Smad4 and that have low levels of RON compared with isogenically matched cells that were deficient in Smad4. Furthermore, knocking down RON expression in Smad4-deficient cells suppressed TGF-β-mediated motility and invasion. We further determined that Smad4-dependent signaling regulated RON expression at the transcriptional level by real-time reverse transcription PCR and RON promoter luciferase reporter assays. Functional inactivation by site-directed mutations of two Smad binding sites on the RON promoter inhibited TGF-β-mediated repression of RON promoter activity. These studies indicate that loss of Smad4 contributes to aberrant RON expression and that cross-talk of Smad4-independent TGF-β signaling and the RON pathway promotes an invasive phenotype. PMID:18310076

  3. A structure activity-relationship study of the bacterial signal molecule HHQ reveals swarming motility inhibition in Bacillus atrophaeus.

    PubMed

    Reen, F Jerry; Shanahan, Rachel; Cano, Rafael; O'Gara, Fergal; McGlacken, Gerard P

    2015-05-21

    The sharp rise in antimicrobial resistance has been matched by a decline in the identification and clinical introduction of new classes of drugs to target microbial infections. Thus new approaches are being sought to counter the pending threat of a post-antibiotic era. In that context, the use of non-growth limiting small molecules, that target virulence behaviour in pathogens, has emerged as a solution with real clinical potential. We have previously shown that two signal molecules (HHQ and PQS) from the nosocomial pathogen Pseudomonas aeruginosa have modulatory activity towards other microorganisms. This current study involves the synthesis and evaluation of analogues of HHQ towards swarming and biofilm virulence behaviour in Bacillus atrophaeus, a soil bacterium and co-inhibitor with P. aeruginosa. Compounds with altered C6-C8 positions on the anthranilate-derived ring of HHQ, display a surprising degree of biological specificity, with certain candidates displaying complete motility inhibition. In contrast, anti-biofilm activity of the parent molecule was completely lost upon alteration at any position indicating a remarkable degree of specificity and delineation of phenotype. PMID:25880413

  4. Helicobacter pylori strains vary cell shape and flagellum number to maintain robust motility in viscous environments.

    PubMed

    Martínez, Laura E; Hardcastle, Joseph M; Wang, Jeffrey; Pincus, Zachary; Tsang, Jennifer; Hoover, Timothy R; Bansil, Rama; Salama, Nina R

    2016-01-01

    The helical shape of the human stomach pathogen Helicobacter pylori has been suggested to provide mechanical advantage for penetrating the viscous stomach mucus layer. Using single-cell tracking and quantitative morphology analysis, we document marked variation in cell body helical parameters and flagellum number among H. pylori strains leading to distinct and broad speed distributions in broth and viscous gastric mucin media. These distributions reflect both temporal variation in swimming speed and morphologic variation within the population. Isogenic mutants with straight-rod morphology showed 7-21% reduction in speed and a lower fraction of motile bacteria. Mutational perturbation of flagellum number revealed a 19% increase in speed with 4 versus 3 median flagellum number. Resistive force theory modeling incorporating variation of both cell shape and flagellum number predicts qualitative speed differences of 10-30% among strains. However, quantitative comparisons suggest resistive force theory underestimates the influence of cell body shape on speed for helical shaped bacteria.

  5. Bacterial Quorum Sensing Molecule N-3-Oxo-Dodecanoyl-L-Homoserine Lactone Causes Direct Cytotoxicity and Reduced Cell Motility in Human Pancreatic Carcinoma Cells

    PubMed Central

    Kumar, Ashwath S.; Bryan, Jeffrey N.; Kumar, Senthil R.

    2014-01-01

    In spite of chemotherapeutic and surgical advances, pancreatic cancer continues to have a dismal prognosis. Metastasis due to tumor cell migration remains the most critical challenge in treating pancreatic cancer, and conventional chemotherapy is rarely curative. In the quest for more novel molecules to fight this disease, we tested the hypothesis that the Pseudomonas aeruginosa quorum sensing signal molecule N-3-oxo-dodecanoyl-L-homoserine lactone (O-DDHSL) would be cytotoxic to and reduce mobility of pancreatic carcinoma cells (Panc-1 and Aspc-1). Results showed a decrease in cell viability from apoptosis, diminished colony formation, and inhibition of migration of the evaluated pancreatic carcinoma cell lines. Also, cell viability decreased in the presence of O-DDHSL when cells were grown in matrigel basement membrane matrix. While messenger RNA for IQGAP-1 decreased in Panc-1 and HPDE cells upon exposure to O-DDHSL, no change was observed in Aspc-1 cells. Cofilin mRNA expression was found to be increased in both HPDE and Panc-1 cells with marginal decrease in Aspc-1 cells. RhoC, a Rho-family GTPase involved in cell motility, increased in the presence of O-DDHSL, suggesting a possible compensatory response to alteration in other migration associated genes. Our results indicate that O-DDHSL could be an effective biomolecule in eukaryotic systems with multimodal function for essential molecular targeting in pancreatic cancer. PMID:25188245

  6. Modulation of intracellular calcium levels by calcium lactate affects colon cancer cell motility through calcium-dependent calpain.

    PubMed

    Sundaramoorthy, Pasupathi; Sim, Jae Jun; Jang, Yeong-Su; Mishra, Siddhartha Kumar; Jeong, Keun-Yeong; Mander, Poonam; Chul, Oh Byung; Shim, Won-Sik; Oh, Seung Hyun; Nam, Ky-Youb; Kim, Hwan Mook

    2015-01-01

    Cancer cell motility is a key phenomenon regulating invasion and metastasis. Focal adhesion kinase (FAK) plays a major role in cellular adhesion and metastasis of various cancers. The relationship between dietary supplementation of calcium and colon cancer has been extensively investigated. However, the effect of calcium (Ca2+) supplementation on calpain-FAK-motility is not clearly understood. We sought to identify the mechanism of FAK cleavage through Ca2+ bound lactate (CaLa), its downstream signaling and role in the motility of human colon cancer cells. We found that treating HCT116 and HT-29 cells with CaLa immediately increased the intracellular Ca2+ (iCa2+) levels for a prolonged period of time. Ca2+ influx induced cleavage of FAK into an N-terminal FAK (FERM domain) in a dose-dependent manner. Phosphorylated FAK (p-FAK) was also cleaved in to its p-N-terminal FAK. CaLa increased colon cancer cells motility. Calpeptin, a calpain inhibitor, reversed the effects of CaLa on FAK and pFAK cleavage in both cancer cell lines. The cleaved FAK translocates into the nucleus and modulates p53 stability through MDM2-associated ubiquitination. CaLa-induced Ca2+ influx increased the motility of colon cancer cells was mediated by calpain activity through FAK and pFAK protein destabilization. In conclusion, these results suggest that careful consideration may be given in deciding dietary Ca2+ supplementation to patient undergoing treatment for metastatic cancer.

  7. A small-molecule inhibitor of T. gondii motility induces the posttranslational modification of myosin light chain-1 and inhibits myosin motor activity.

    PubMed

    Heaslip, Aoife T; Leung, Jacqueline M; Carey, Kimberly L; Catti, Federica; Warshaw, David M; Westwood, Nicholas J; Ballif, Bryan A; Ward, Gary E

    2010-01-15

    Toxoplasma gondii is an obligate intracellular parasite that enters cells by a process of active penetration. Host cell penetration and parasite motility are driven by a myosin motor complex consisting of four known proteins: TgMyoA, an unconventional Class XIV myosin; TgMLC1, a myosin light chain; and two membrane-associated proteins, TgGAP45 and TgGAP50. Little is known about how the activity of the myosin motor complex is regulated. Here, we show that treatment of parasites with a recently identified small-molecule inhibitor of invasion and motility results in a rapid and irreversible change in the electrophoretic mobility of TgMLC1. While the precise nature of the TgMLC1 modification has not yet been established, it was mapped to the peptide Val46-Arg59. To determine if the TgMLC1 modification is responsible for the motility defect observed in parasites after compound treatment, the activity of myosin motor complexes from control and compound-treated parasites was compared in an in vitro motility assay. TgMyoA motor complexes containing the modified TgMLC1 showed significantly decreased motor activity compared to control complexes. This change in motor activity likely accounts for the motility defects seen in the parasites after compound treatment and provides the first evidence, in any species, that the mechanical activity of Class XIV myosins can be modulated by posttranslational modifications to their associated light chains.

  8. 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.

  9. Polo-like kinase 1 induces epithelial-to-mesenchymal transition and promotes epithelial cell motility by activating CRAF/ERK signaling

    PubMed Central

    Wu, Jianguo; Ivanov, Andrei I; Fisher, Paul B; Fu, Zheng

    2016-01-01

    Polo-like kinase 1 (PLK1) is a key cell cycle regulator implicated in the development of various cancers, including prostate cancer. However, the functions of PLK1 beyond cell cycle regulation remain poorly characterized. Here, we report that PLK1 overexpression in prostate epithelial cells triggers oncogenic transformation. It also results in dramatic transcriptional reprogramming of the cells, leading to epithelial-to-mesenchymal transition (EMT) and stimulation of cell migration and invasion. Consistently, PLK1 downregulation in metastatic prostate cancer cells enhances epithelial characteristics and inhibits cell motility. The signaling mechanisms underlying the observed cellular effects of PLK1 involve direct PLK1-dependent phosphorylation of CRAF with subsequent stimulation of the MEK1/2-ERK1/2-Fra1-ZEB1/2 signaling pathway. Our findings highlight novel non-canonical functions of PLK1 as a key regulator of EMT and cell motility in normal prostate epithelium and prostate cancer. This study also uncovers a previously unanticipated role of PLK1 as a potent activator of MAPK signaling. DOI: http://dx.doi.org/10.7554/eLife.10734.001 PMID:27003818

  10. BRE facilitates skeletal muscle regeneration by promoting satellite cell motility and differentiation.

    PubMed

    Xiao, Lihai; Lee, Kenneth Ka Ho

    2016-01-06

    The function of the Bre gene in satellite cells was investigated during skeletal muscle regeneration. The tibialis anterior leg muscle was experimentally injured in Bre knockout mutant (BRE-KO) mice. It was established that the accompanying muscle regeneration was impaired as compared with their normal wild-type counterparts (BRE-WT). There were significantly fewer pax7(+) satellite cells and smaller newly formed myofibers present in the injury sites of BRE-KO mice. Bre was required for satellite cell fusion and myofiber formation. The cell fusion index and average length of newly-formed BRE-KO myofibers were found to be significantly reduced as compared with BRE-WT myofibers. It is well established that satellite cells are highly invasive which confers on them the homing ability to reach the muscle injury sites. Hence, we tracked the migratory behavior of these cells using time-lapse microscopy. Image analysis revealed no difference in directionality of movement between BRE-KO and BRE-WT satellite cells but there was a significant decrease in the velocity of BRE-KO cell movement. Moreover, chemotactic migration assays indicated that BRE-KO satellite cells were significantly less responsive to chemoattractant SDF-1α than BRE-WT satellite cells. We also established that BRE normally protects CXCR4 from SDF-1α-induced degradation. In sum, BRE facilitates skeletal muscle regeneration by enhancing satellite cell motility, homing and fusion.

  11. BRE facilitates skeletal muscle regeneration by promoting satellite cell motility and differentiation

    PubMed Central

    Xiao, Lihai; Lee, Kenneth Ka Ho

    2016-01-01

    ABSTRACT The function of the Bre gene in satellite cells was investigated during skeletal muscle regeneration. The tibialis anterior leg muscle was experimentally injured in Bre knockout mutant (BRE-KO) mice. It was established that the accompanying muscle regeneration was impaired as compared with their normal wild-type counterparts (BRE-WT). There were significantly fewer pax7+ satellite cells and smaller newly formed myofibers present in the injury sites of BRE-KO mice. Bre was required for satellite cell fusion and myofiber formation. The cell fusion index and average length of newly-formed BRE-KO myofibers were found to be significantly reduced as compared with BRE-WT myofibers. It is well established that satellite cells are highly invasive which confers on them the homing ability to reach the muscle injury sites. Hence, we tracked the migratory behavior of these cells using time-lapse microscopy. Image analysis revealed no difference in directionality of movement between BRE-KO and BRE-WT satellite cells but there was a significant decrease in the velocity of BRE-KO cell movement. Moreover, chemotactic migration assays indicated that BRE-KO satellite cells were significantly less responsive to chemoattractant SDF-1α than BRE-WT satellite cells. We also established that BRE normally protects CXCR4 from SDF-1α-induced degradation. In sum, BRE facilitates skeletal muscle regeneration by enhancing satellite cell motility, homing and fusion. PMID:26740569

  12. Wnt Signaling in Cell Motility and Invasion: Drawing Parallels between Development and Cancer

    PubMed Central

    Sedgwick, Alanna E.; D’Souza-Schorey, Crislyn

    2016-01-01

    The importance of canonical and non-canonical Wnt signal transduction cascades in embryonic development and tissue homeostasis is well recognized. The aberrant activation of these pathways in the adult leads to abnormal cellular behaviors, and tumor progression is frequently a consequence. Here we discuss recent findings and analogies between Wnt signaling in developmental processes and tumor progression, with a particular focus on cell motility and matrix invasion and highlight the roles of the ARF (ADP-Ribosylation Factor) and Rho-family small GTP-binding proteins. Wnt-regulated signal transduction from cell surface receptors, signaling endosomes and/or extracellular vesicles has the potential to profoundly influence cell movement, matrix degradation and paracrine signaling in both development and disease. PMID:27589803

  13. Wnt Signaling in Cell Motility and Invasion: Drawing Parallels between Development and Cancer.

    PubMed

    Sedgwick, Alanna E; D'Souza-Schorey, Crislyn

    2016-01-01

    The importance of canonical and non-canonical Wnt signal transduction cascades in embryonic development and tissue homeostasis is well recognized. The aberrant activation of these pathways in the adult leads to abnormal cellular behaviors, and tumor progression is frequently a consequence. Here we discuss recent findings and analogies between Wnt signaling in developmental processes and tumor progression, with a particular focus on cell motility and matrix invasion and highlight the roles of the ARF (ADP-Ribosylation Factor) and Rho-family small GTP-binding proteins. Wnt-regulated signal transduction from cell surface receptors, signaling endosomes and/or extracellular vesicles has the potential to profoundly influence cell movement, matrix degradation and paracrine signaling in both development and disease. PMID:27589803

  14. No Correlates for Somatic Motility in Freeze-Fractured Hair-Cell Membranes of Lizards and Birds

    NASA Astrophysics Data System (ADS)

    Köppl, C.; Forge, A.; Manley, G. A.

    2003-02-01

    It is not known whether active processes in mammals and non-mammals are due to the same underlying mechanism. To address this, we studied the size and density of particles in hair-cell membranes in mammals, in a lizard, the Tokay gecko, and in a bird, the barn owl. We surmised that if the prominent particles described in mammalian outer-hair-cell membranes are responsible for cochlear motility, a similar occurrence in non-mammalian hair cells would argue for similar mechanisms. Particle densities differed, however, substantially from those of mammals, suggesting that non-mammals have no membrane-based motility.

  15. Single-gene tuning of Caulobacter cell cycle period and noise, swarming motility, and surface adhesion

    PubMed Central

    Lin, Yihan; Crosson, Sean; Scherer, Norbert F

    2010-01-01

    Sensor histidine kinases underlie the regulation of a range of physiological processes in bacterial cells, from chemotaxis to cell division. In the gram-negative bacterium Caulobacter crescentus, the membrane-bound histidine kinase, DivJ, is a polar-localized regulator of cell cycle progression and development. We show that DivJ localizes to the cell pole through a dynamic diffusion and capture mechanism rather than by active localization. Analysis of single C. crescentus cells in microfluidic culture demonstrates that controlled expression of divJ permits facile tuning of both the mean and noise of the cell division period. Simulations of the cell cycle that use a simplified protein interaction network capture previously measured oscillatory protein profiles, and recapitulate the experimental observation that deletion of divJ increases the cell cycle period and noise. We further demonstrate that surface adhesion and swarming motility of C. crescentus in semi-solid media can also be tuned by divJ expression. We propose a model in which pleiotropic control of polar cell development by the DivJ–DivK–PleC signaling pathway underlies divJ-dependent tuning of cell swarming and adhesion behaviors. PMID:21179017

  16. Approaches to myosin modelling in a two-phase flow model for cell motility

    NASA Astrophysics Data System (ADS)

    Kimpton, L. S.; Whiteley, J. P.; Waters, S. L.; Oliver, J. M.

    2016-04-01

    A wide range of biological processes rely on the ability of cells to move through their environment. Mathematical models have been developed to improve our understanding of how cells achieve motion. Here we develop models that explicitly track the cell's distribution of myosin within a two-phase flow framework. Myosin is a small motor protein which is important for contracting the cell's actin cytoskeleton and enabling cell motion. The two phases represent the actin network and the cytosol in the cell. We start from a fairly general description of myosin kinetics, advection and diffusion in the two-phase flow framework, then identify a number of sub-limits of the model that may be relevant in practice, two of which we investigate further via linear stability analyses and numerical simulations. We demonstrate that myosin-driven contraction of the actin network destabilizes a stationary steady state leading to cell motion, but that rapid diffusion of myosin and rapid unbinding of myosin from the actin network are stabilizing. We use numerical simulation to investigate travelling-wave solutions relevant to a steadily gliding cell and we consider a reduction of the model in which the cell adheres strongly to the substrate on which it is crawling. This work demonstrates that a number of existing models for the effect of myosin on cell motility can be understood as different sub-limits of our two-phase flow model.

  17. D1-type dopamine receptors inhibit growth cone motility in cultured retina neurons: evidence that neurotransmitters act as morphogenic growth regulators in the developing central nervous system.

    PubMed Central

    Lankford, K L; DeMello, F G; Klein, W L

    1988-01-01

    Precedent exists for the early development and subsequent down-regulation of neurotransmitter receptor systems in the vertebrate central nervous system, but the function of such embryonic receptors has not been established. Here we show that stimulation of early-developing dopamine receptors in avian retina cells greatly inhibits the motility of neuronal growth cones. Neurons from embryonic chicken retinas were cultured in low-density monolayers, and their growth cones were observed with phase-contrast or video-enhanced-contrast-differential-interference-contrast (VEC-DIC) microscopy. Approximately 25% of the neurons responded to micromolar dopamine with a rapid reduction in filopodial activity followed by a flattening of growth cones and retraction of neurites. The response occurred at all ages examined (embryonic day-8 retinal neurons cultured on polylysine-coated coverslips for 1-7 days), although neurite retraction was greatest in younger cultures. Effects of dopamine on growth cone function could be reversed by haloperidol or (+)-SCH 23390, whereas forskolin elicited a response similar to dopamine; these data show the response was receptor-mediated, acting through a D1-type system, and are consistent with the use of cAMP as a second messenger. The experiments provide strong support for the hypothesis that neurotransmitters, besides mediating transynaptic signaling in the adult, may have a role in neuronal differentiation as growth regulators. Images PMID:3380807

  18. D1-type dopamine receptors inhibit growth cone motility in cultured retina neurons: evidence that neurotransmitters act as morphogenic growth regulators in the developing central nervous system.

    PubMed Central

    Lankford, K L; DeMello, F G; Klein, W L

    1988-01-01

    Precedent exists for the early development and subsequent down-regulation of neurotransmitter receptor systems in the vertebrate central nervous system, but the function of such embryonic receptors has not been established. Here we show that stimulation of early-developing dopamine receptors in avian retina cells greatly inhibits the motility of neuronal growth cones. Neurons from embryonic chicken retinas were cultured in low-density monolayers, and their growth cones were observed with phase-contrast or video-enhanced-contrast-differential-interference-contrast (VEC-DIC) microscopy. Approximately 25% of the neurons responded to micromolar dopamine with a rapid reduction in filopodial activity followed by a flattening of growth cones and retraction of neurites. The response occurred at all ages examined (embryonic day-8 retinal neurons cultured on polylysine-coated coverslips for 1-7 days), although neurite retraction was greatest in younger cultures. Effects of dopamine on growth cone function could be reversed by haloperidol or (+)-SCH 23390, whereas forskolin elicited a response similar to dopamine; these data show the response was receptor-mediated, acting through a D1-type system, and are consistent with the use of cAMP as a second messenger. The experiments provide strong support for the hypothesis that neurotransmitters, besides mediating transynaptic signaling in the adult, may have a role in neuronal differentiation as growth regulators. Images PMID:3357895

  19. The activation of directional stem cell motility by green light-emitting diode irradiation.

    PubMed

    Ong, Wei-Kee; Chen, How-Foo; Tsai, Cheng-Ting; Fu, Yun-Ju; Wong, Yi-Shan; Yen, Da-Jen; Chang, Tzu-Hao; Huang, Hsien-Da; Lee, Oscar Kuang-Sheng; Chien, Shu; Ho, Jennifer Hui-Chun

    2013-03-01

    Light-emitting diode (LED) irradiation is potentially a photostimulator to manipulate cell behavior by opsin-triggered phototransduction and thermal energy supply in living cells. Directional stem cell motility is critical for the efficiency and specificity of stem cells in tissue repair. We explored that green LED (530 nm) irradiation directed the human orbital fat stem cells (OFSCs) to migrate away from the LED light source through activation of extracellular signal-regulated kinases (ERK)/MAP kinase/p38 signaling pathway. ERK inhibitor selectively abrogated light-driven OFSC migration. Phosphorylation of these kinases as well as green LED irradiation-induced cell migration was facilitated by increasing adenosine triphosphate (ATP) production in OFSCs after green LED exposure, and which was thermal stress-independent mechanism. OFSCs, which are multi-potent mesenchymal stem cells isolated from human orbital fat tissue, constitutionally express three opsins, i.e. retinal pigment epithelium-derived rhodopsin homolog (RRH), encephalopsin (OPN3) and short-wave-sensitive opsin 1 (OPN1SW). However, only two non-visual opsins, i.e. RRH and OPN3, served as photoreceptors response to green LED irradiation-induced OFSC migration. In conclusion, stem cells are sensitive to green LED irradiation-induced directional cell migration through activation of ERK signaling pathway via a wavelength-dependent phototransduction.

  20. WISP-1 increases MMP-2 expression and cell motility in human chondrosarcoma cells.

    PubMed

    Hou, Chun-Han; Chiang, Yi-Chun; Fong, Yi-Chin; Tang, Chih-Hsin

    2011-06-01

    Chondrosarcoma is a type of highly malignant tumor with a potent capacity to invade locally and cause distant metastasis. Chondrosarcoma shows a predilection for metastasis to the lungs. WISP-1 is a cysteine-rich protein that belongs to the CCN (Cyr61, CTGF, Nov) family of matricellular proteins. However, the effect of WISP-1 on migration activity in human chondrosarcoma cells is mostly unknown. Here we found that WISP-1 increased the migration and expression of matrix metalloproteinase (MMP)-2 in human chondrosarcoma cells (JJ012 cells). We also found that human chondrosarcoma tissues had significant expression of the WISP-1 which was higher than that in normal cartilage. α5β1 monoclonal antibody and MAPK kinase (MEK) inhibitors (PD98059 and U0126) inhibited the WISP-1-induced increase of the migration and MMP-2 up-regulation of chondrosarcoma cells. WISP-1 stimulation increased the phosphorylation of focal adhesion kinase (FAK), MEK and extracellular signal-regulated kinase (ERK). In addition, NF-κB inhibitors also suppressed the cell migration and MMP-2 expression enhanced by WISP-1. Moreover, WISP-1 increased NF-κB luciferase activity and binding of p65 to the NF-κB element on the MMP-2 promoter. Taken together, our results indicated that WISP-1 enhances the migration of chondrosarcoma cells by increasing MMP-2 expression through the α5β1 integrin receptor, FAK, MEK, ERK, p65 and NF-κB signal transduction pathway.

  1. Interplay between motility and cell-substratum adhesion in amoeboid cells

    PubMed Central

    Zhu, Xiaoying; Bouffanais, Roland; Yue, Dick K. P.

    2015-01-01

    The effective migration of amoeboid cells requires a fine regulation of cell-substratum adhesion. These entwined processes have been shown to be regulated by a host of biophysical and biochemical cues. Here, we reveal the pivotal role played by calcium-based mechanosensation in the active regulation of adhesion resulting in a high migratory adaptability. Using mechanotactically driven Dictyostelium discoideum amoebae, we uncover the existence of optimal mechanosensitive conditions—corresponding to specific levels of extracellular calcium—for persistent directional migration over physicochemically different substrates. When these optimal mechanosensitive conditions are met, noticeable enhancement in cell migration directionality and speed is achieved, yet with significant differences among the different substrates. In the same narrow range of calcium concentrations that yields optimal cellular mechanosensory activity, we uncovered an absolute minimum in cell-substratum adhesion activity, for all considered substrates, with differences in adhesion strength among them amplified. The blocking of the mechanosensitive ion channels with gadolinium—i.e., the inhibition of the primary mechanosensory apparatus—hampers the active reduction in substrate adhesion, thereby leading to the same undifferentiated and drastically reduced directed migratory response. The adaptive behavioral responses of Dictyostelium cells sensitive to substrates with varying physicochemical properties suggest the possibility of novel surface analyses based on the mechanobiological ability of mechanosensitive and guidable cells to probe substrates at the nanometer-to-micrometer level. PMID:26487898

  2. Computational estimates of membrane flow and tension gradient in motile cells.

    PubMed

    Fogelson, Ben; Mogilner, Alex

    2014-01-01

    All parts of motile cells, including the plasma membrane, have to translocate in the direction of locomotion. Both directed intracellular membrane transport coupled with polarized endo- and exocytosis and fluid flow in the plane of the plasma membrane can contribute to this overall plasma membrane translocation. It remains unclear how strong a force is required to generate this flow. We numerically solve Stokes equations for the viscous membrane flow across a flat plasma membrane surface in the presence of transmembrane proteins attached to the cytoskeleton and find the membrane tension gradient associated with this flow. This gradient is sensitive to the size and density of the transmembrane proteins attached to the cytoskeleton and can become significant enough to slow down cell movement. We estimate the influence of intracellular membrane transport and actin growth and contraction on the tension gradient, and discuss possible 'tank tread' flow at ventral and dorsal surfaces.

  3. Involvement of S100-related calcium-binding protein pEL98 (or mts1) in cell motility and tumor cell invasion.

    PubMed

    Takenaga, K; Nakamura, Y; Endo, H; Sakiyama, S

    1994-08-01

    We examined the relationship between cell motility and the expressions of pEL98 (mts1) mRNA and protein in various murine normal and transformed cells. The expression of pEL98 (mts1) in v-Ha-ras-transformed NIH3T3 cells and in normal rat kidney cells transformed by either v-Ha-ras or v-src was increased over that in the corresponding parental cells at both mRNA and protein levels. The expression in normal rat fibroblasts (3Y1) transformed by v-Ha-ras was also increased compared with that in 3Y1 cells. However, the expression of pEL98 (mts1) in 3Y1 cells transformed by v-src was increased in one clone (src 3Y1-K), but decreased in another clone (src 3Y1-H). The expression level of pEL98 (mts1) correlated well with cell motility, which was examined by measuring cell tracks by phagokinesis. In order to test direct involvement of the pEL98 (mts1) protein in cell motility, src 3Y1-H cells that showed low cell motility were transfected with pEL98 cDNA. The transfectants expressing large amounts of the pEL98 protein showed significantly higher cell motility than src 3Y1-H cells. The expression of pEL98 (mts1) was also found to be correlated with motile and invasive abilities in various clones derived from Lewis lung carcinoma. These results suggest that the pEL98 (mts1) protein plays a role in regulating cell motility and tumor cell invasiveness. PMID:7928629

  4. 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

  5. Actin on disease--studying the pathobiology of cell motility using Dictyostelium discoideum.

    PubMed

    Carnell, Michael J; Insall, Robert H

    2011-02-01

    The actin cytoskeleton in eukaryotic cells provides cell structure and organisation, and allows cells to generate forces against membranes. As such it is a central component of a variety of cellular structures involved in cell motility, cytokinesis and vesicle trafficking. In multicellular organisms these processes contribute towards embryonic development and effective functioning of cells of all types, most obviously rapidly moving cells like lymphocytes. Actin also defines and maintains the architecture of complex structures such as neuronal synapses and stereocillia, and is required for basic housekeeping tasks within the cell. It is therefore not surprising that misregulation of the actin cytoskeleton can cause a variety of disease pathologies, including compromised immunity, neurodegeneration, and cancer spread. Dictyostelium discoideum has long been used as a tool for dissecting the mechanisms by which eukaryotic cells migrate and chemotax, and recently it has gained precedence as a model organism for studying the roles of conserved pathways in disease processes. Dictyostelium's unusual lifestyle, positioned between unicellular and multicellular organisms, combined with ease of handling and strong conservation of actin regulatory machinery with higher animals, make it ideally suited for studying actin-related diseases. Here we address how research in Dictyostelium has contributed to our understanding of immune deficiencies and neurological defects in humans, and briefly discuss its future prospects for furthering our understanding of neurodegenerative disorders.

  6. Intracellular photoactivation of caged cGMP induces myosin II and actin responses in motile cells.

    PubMed

    Pfannes, Eva K B; Anielski, Alexander; Gerhardt, Matthias; Beta, Carsten

    2013-12-01

    Cyclic GMP (cGMP) is a ubiquitous second messenger in eukaryotic cells. It is assumed to regulate the association of myosin II with the cytoskeleton of motile cells. When cells of the social amoeba Dictyostelium discoideum are exposed to chemoattractants or to increased osmotic stress, intracellular cGMP levels rise, preceding the accumulation of myosin II in the cell cortex. To directly investigate the impact of intracellular cGMP on cytoskeletal dynamics in a living cell, we released cGMP inside the cell by laser-induced photo-cleavage of a caged precursor. With this approach, we could directly show in a live cell experiment that an increase in intracellular cGMP indeed induces myosin II to accumulate in the cortex. Unexpectedly, we observed for the first time that also the amount of filamentous actin in the cell cortex increases upon a rise in the cGMP concentration, independently of cAMP receptor activation and signaling. We discuss our results in the light of recent work on the cGMP signaling pathway and suggest possible links between cGMP signaling and the actin system. PMID:24136144

  7. Heparanase promotes myeloma progression by inducing mesenchymal features and motility of myeloma cells

    PubMed Central

    Trotter, Timothy N.; Peker, Deniz; Regal, Kellie M.; Javed, Amjad; Suva, Larry J.; Yang, Yang

    2016-01-01

    Bone dissemination and bone disease occur in approximately 80% of patients with multiple myeloma (MM) and are a major cause of patient mortality. We previously demonstrated that MM cell-derived heparanase (HPSE) is a major driver of MM dissemination to and progression in new bone sites. However the mechanism(s) by which HPSE promotes MM progression remains unclear. In the present study, we investigated the involvement of mesenchymal features in HPSE-promoted MM progression in bone. Using a combination of molecular, biochemical, cellular, and in vivo approaches, we demonstrated that (1) HPSE enhanced the expression of mesenchymal markers in both MM and vascular endothelial cells; (2) HPSE expression in patient myeloma cells positively correlated with the expression of the mesenchymal markers vimentin and fibronectin. Additional mechanistic studies revealed that the enhanced mesenchymal-like phenotype induced by HPSE in MM cells is due, at least in part, to the stimulation of the ERK signaling pathway. Finally, knockdown of vimentin in HPSE expressing MM cells resulted in significantly attenuated MM cell dissemination and tumor growth in vivo. Collectively, these data demonstrate that the mesenchymal features induced by HPSE in MM cells contribute to enhanced tumor cell motility and bone-dissemination. PMID:26849235

  8. Lrs14 transcriptional regulators influence biofilm formation and cell motility of Crenarchaea

    PubMed Central

    Orell, Alvaro; Peeters, Eveline; Vassen, Victoria; Jachlewski, Silke; Schalles, Sven; Siebers, Bettina; Albers, Sonja-Verena

    2013-01-01

    Like bacteria, archaea predominately exist as biofilms in nature. However, the environmental cues and the molecular mechanisms driving archaeal biofilm development are not characterized. Here we provide data suggesting that the transcriptional regulators belonging to the Lrs14-like protein family constitute a key regulatory factor during Sulfolobus biofilm development. Among the six lrs14-like genes encoded by Sulfolobus acidocaldarius, the deletion of three led to markedly altered biofilm phenotypes. Although Δsaci1223 and Δsaci1242 deletion mutants were impaired in biofilm formation, the Δsaci0446 deletion strain exhibited a highly increased extracellular polymeric substance (EPS) production, leading to a robust biofilm structure. Moreover, although the expression of the adhesive pili (aap) genes was upregulated, the genes of the motility structure, the archaellum (fla), were downregulated rendering the Δsaci0446 strain non-motile. Gel shift assays confirmed that Saci0446 bound to the promoter regions of fla and aap thus controlling the expression of both cell surface structures. In addition, genetic epistasis analysis using Δsaci0446 as background strain identified a gene cluster involved in the EPS biosynthetic pathway of S. acidocaldarius. These results provide insights into both the molecular mechanisms that govern biofilm formation in Crenarchaea and the functionality of the Lrs14-like proteins, an archaea-specific class of transcriptional regulators. PMID:23657363

  9. Melatonin enhances the human mesenchymal stem cells motility via melatonin receptor 2 coupling with Gαq in skin wound healing.

    PubMed

    Lee, Sei-Jung; Jung, Young Hyun; Oh, Sang Yub; Yun, Seung Pil; Han, Ho Jae

    2014-11-01

    Melatonin, a circadian rhythm-promoting molecule, has a variety of biological functions, but the functional role of melatonin in the motility of mesenchymal stem cells (MSCs) has yet to be studied. In a mouse skin excisional wound model, we found that transplantation of umbilical cord blood (UCB)-MSCs pretreated with melatonin enhanced wound closure, granulation, and re-epithelialization at mouse skin wound sites, where relatively more UCB-MSCs which were engrafted onto the wound site were detected. Thus, we identified the signaling pathway of melatonin, which affects the motility of UCB-MSCs. Melatonin (1 μm) significantly increased the motility of UCB-MSCs, which had been inhibited by the knockdown of melatonin receptor 2 (MT2). We found that Gαq coupled with MT2 and that the binding of Gαq to MT2 uniquely stimulated an atypical PKC isoform, PKCζ. Melatonin induced the phosphorylation of FAK and paxillin, which were concurrently downregulated by blocking of the PKC activity. Melatonin increased the levels of active Cdc42 and Arp2/3, and it has the ability to stimulate cytoskeletal reorganization-related proteins such as profilin-1, cofilin-1, and F-actin in UCB-MSCs. Finally, a lack of MT2 expression in UCB-MSCs during a mouse skin transplantation experiment resulted in impaired wound healing and less engraftment of stem cells at the wound site. These results demonstrate that melatonin signaling via MT2 triggers FAK/paxillin phosphorylation to stimulate reorganization of the actin cytoskeleton, which is responsible for Cdc42/Arp2/3 activation to promote UCB-MSCs motility.

  10. Peroxisome proliferator-activated receptor γ agonist efatutazone impairs transforming growth factor β2-induced motility of epidermal growth factor receptor tyrosine kinase inhibitor-resistant lung cancer cells.

    PubMed

    Serizawa, Masakuni; Murakami, Haruyasu; Watanabe, Masaru; Takahashi, Toshiaki; Yamamoto, Nobuyuki; Koh, Yasuhiro

    2014-06-01

    Epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKI) are effective for non-small cell lung cancers (NSCLC) with EGFR-activating mutations. However, most responders develop resistance. Efatutazone, a novel peroxisome proliferator-activated receptor gamma (PPARγ) agonist, is currently under clinical evaluation; it has antiproliferative effects and induces cellular morphological changes and differentiation. The present study investigated the effects of efatutazone in EGFR-TKI-resistant NSCLC cells, while focusing on cell motility. The PC-9-derived NSCLC cell lines PC-9ER and PC-9ZD, resistant to EGFR-TKI due to v-crk avian sarcoma virus CT10 oncogene homolog-like (CRKL) amplification-induced phosphatidylinositol 3-kinase (PI3K)/v-akt murine thymoma viral oncogene homolog (AKT) activation and an EGFR T790M mutation, respectively, were used. These cells exhibit enhanced cell motility due to transforming growth factor β (TGF-β)/Smad2 family member 2 (Smad2) pathway activation. Efatutazone had no growth-inhibitory effect on the tested cells but inhibited the motility of EGFR-TKI-resistant cells in wound closure and transwell assays. Efatutazone plus erlotinib treatment provided greater inhibition of PC-9ER cell migration than efatutazone or erlotinib alone. Efatutazone suppressed increased TGF-β2 secretion from both cell lines (shown by ELISA) and downregulation of TGF-β2 transcription (observed by quantitative RT-PCR). Immunoblot analysis and luciferase assays revealed that efatutazone suppressed Smad2 phosphorylation and its transcriptional activity. These results suggest that efatutazone inhibits cell motility by antagonizing the TGF-β/Smad2 pathway and effectively prevents metastasis in NSCLC patients with acquired resistance to EGFR-TKI regardless of the resistance mechanism.

  11. The HP0256 gene product is involved in motility and cell envelope architecture of Helicobacter pylori

    PubMed Central

    2010-01-01

    Background Helicobacter pylori is the causative agent for gastritis, and peptic and duodenal ulcers. The bacterium displays 5-6 polar sheathed flagella that are essential for colonisation and persistence in the gastric mucosa. The biochemistry and genetics of flagellar biogenesis in H. pylori has not been fully elucidated. Bioinformatics analysis suggested that the gene HP0256, annotated as hypothetical, was a FliJ homologue. In Salmonella, FliJ is a chaperone escort protein for FlgN and FliT, two proteins that themselves display chaperone activity for components of the hook, the rod and the filament. Results Ablation of the HP0256 gene in H. pylori significantly reduced motility. However, flagellin and hook protein synthesis was not affected in the HP0256 mutant. Transmission electron transmission microscopy revealed that the HP0256 mutant cells displayed a normal flagellum configuration, suggesting that HP0256 was not essential for assembly and polar localisation of the flagella in the cell. Interestingly, whole genome microarrays of an HP0256 mutant revealed transcriptional changes in a number of genes associated with the flagellar regulon and the cell envelope, such as outer membrane proteins and adhesins. Consistent with the array data, lack of the HP0256 gene significantly reduced adhesion and the inflammatory response in host cells. Conclusions We conclude that HP0256 is not a functional counterpart of FliJ in H. pylori. However, it is required for full motility and it is involved, possibly indirectly, in expression of outer membrane proteins and adhesins involved in pathogenesis and adhesion. PMID:20377912

  12. 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

  13. Fascin, an actin-bundling protein associated with cell motility, is upregulated in hormone receptor negative breastancer

    PubMed Central

    Grothey, A; Hashizume, R; Sahin, A A; McCrea, P D

    2000-01-01

    Loss of hormone receptor (HR) status in breast carcinomas is associated with increased tumour cell motility and invasiveness. In an immunohistological study of 58 primary breast cancers, oestrogen (ER) and progesterone (PR) receptor levels were inversely correlated with the expression of fascin, an actin-bundling protein associated with cell motility (P< 0.0001 and P = 0.0019, respectively). In addition, fascin was preferentially expressed in non-diploid tumours (P = 0.03). In summary, the upregulation of fascin in HR-negative breast cancers may contribute to their more aggressive behaviour. © 2000 Cancer Research Campaign PMID:10970687

  14. Signaling networks and cell motility: a computational approach using a phase field description.

    PubMed

    Marth, Wieland; Voigt, Axel

    2014-07-01

    The processes of protrusion and retraction during cell movement are driven by the turnover and reorganization of the actin cytoskeleton. Within a reaction-diffusion model which combines processes along the cell membrane with processes within the cytoplasm a Turing type instability is used to form the necessary polarity to distinguish between cell front and rear and to initiate the formation of different organizational arrays within the cytoplasm leading to protrusion and retraction. A simplified biochemical network model for the activation of GTPase which accounts for the different dimensionality of the cell membrane and the cytoplasm is used for this purpose and combined with a classical Helfrich type model to account for bending and stiffness effects of the cell membrane. In addition streaming within the cytoplasm and the extracellular matrix is taken into account. Combining these phenomena allows to simulate the dynamics of cells and to reproduce the primary phenomenology of cell motility. The coupled model is formulated within a phase field approach and solved using adaptive finite elements. PMID:23835784

  15. Regulation of T Cell Motility In Vitro and In Vivo by LPA and LPA2

    PubMed Central

    Knowlden, Sara A.; Capece, Tara; Popovic, Milan; Chapman, Timothy J.; Rezaee, Fariba; Kim, Minsoo; Georas, Steve N.

    2014-01-01

    Lysophosphatidic acid (LPA) and the LPA-generating enzyme autotaxin (ATX) have been implicated in lymphocyte trafficking and the regulation of lymphocyte entry into lymph nodes. High local concentrations of LPA are thought to be present in lymph node high endothelial venules, suggesting a direct influence of LPA on cell migration. However, little is known about the mechanism of action of LPA, and more work is needed to define the expression and function of the six known G protein-coupled receptors (LPA 1–6) in T cells. We studied the effects of 18∶1 and 16∶0 LPA on naïve CD4+ T cell migration and show that LPA induces CD4+ T cell chemorepulsion in a Transwell system, and also improves the quality of non-directed migration on ICAM-1 and CCL21 coated plates. Using intravital two-photon microscopy, lpa2−/− CD4+ T cells display a striking defect in early migratory behavior at HEVs and in lymph nodes. However, later homeostatic recirculation and LPA-directed migration in vitro were unaffected by loss of lpa2. Taken together, these data highlight a previously unsuspected and non-redundant role for LPA2 in intranodal T cell motility, and suggest that specific functions of LPA may be manipulated by targeting T cell LPA receptors. PMID:25003200

  16. Regulation of T cell motility in vitro and in vivo by LPA and LPA2.

    PubMed

    Knowlden, Sara A; Capece, Tara; Popovic, Milan; Chapman, Timothy J; Rezaee, Fariba; Kim, Minsoo; Georas, Steve N

    2014-01-01

    Lysophosphatidic acid (LPA) and the LPA-generating enzyme autotaxin (ATX) have been implicated in lymphocyte trafficking and the regulation of lymphocyte entry into lymph nodes. High local concentrations of LPA are thought to be present in lymph node high endothelial venules, suggesting a direct influence of LPA on cell migration. However, little is known about the mechanism of action of LPA, and more work is needed to define the expression and function of the six known G protein-coupled receptors (LPA 1-6) in T cells. We studied the effects of 18∶1 and 16∶0 LPA on naïve CD4+ T cell migration and show that LPA induces CD4+ T cell chemorepulsion in a Transwell system, and also improves the quality of non-directed migration on ICAM-1 and CCL21 coated plates. Using intravital two-photon microscopy, lpa2-/- CD4+ T cells display a striking defect in early migratory behavior at HEVs and in lymph nodes. However, later homeostatic recirculation and LPA-directed migration in vitro were unaffected by loss of lpa2. Taken together, these data highlight a previously unsuspected and non-redundant role for LPA2 in intranodal T cell motility, and suggest that specific functions of LPA may be manipulated by targeting T cell LPA receptors. PMID:25003200

  17. Interstitial flows promote an amoeboid over mesenchymal motility of breast cancer cells revealed by a three dimensional microfluidic model

    PubMed Central

    Huang, Yu Ling; Tung, Chih-kuan; Zheng, Anqi; Kim, Beum Jun; Wu, Mingming

    2015-01-01

    Malignant tumors are often associated with an elevated fluid pressure due to the abnormal growth of vascular vessels, and thus an increased interstitial flow out of the tumor. Recent in vitro work revealed that interstitial flows critically regulated tumor cell migration within a three dimensional biomatrix, and breast cancer cell migration behavior depended sensitively on the cell seeding density, chemokine availability and flow rates. In this paper, we focus on roles of interstitial flows in modulating heterogeneity of cancer cell motility phenotype within a three dimensional biomatrix. Using a newly developed microfluidic model, we show that breast cancer cells (MDA-MB-231) embedded in a 3D type I collagen matrix exhibit both an amoeboid and a mesenchymal motility, and interstitial flows promote the cell population towards the amoeboid motility phenotype. Furthermore, the addition of exogenous adhesion molecules (fibronectin) within the extracellular matrix (type I collagen) partially rescues the mesenchymal phenotype in the presence of the flow. Quantitative analysis of cell tracks and cell shape shows distinct differential migration characteristics of amoeboid and mesenchymal cells. Notably, the fastest moving cells belong to the subpopulation of amoeboid cells. Together, these findings highlight the important roles of biophysical forces in modulating tumor cell migration heterogeneity and plasticity, as well as the suitability of microfluidic models in interrogating tumor cell dynamics at single-cell and subpopulation level. PMID:26235230

  18. Activation of sperm motility in striped bass via a cAMP-independent pathway.

    PubMed

    He, Shuyang; Jenkins-Keeran, Karen; Woods, L Curry

    2004-05-01

    The objective of the present study was to identify the effect of osmolality, ions (K+, H+, Ca2+, Mg2+) and cAMP on the initiation of sperm motility in striped bass (Morone saxatilis). Striped bass spermatozoa remained motile in solutions isotonic to seminal plasma (350 mOsm/kg) until osmolality reached 600 mOsm/kg. K+ (0-100 mM) had no effect ( p>0.05 ) on sperm motility, and sperm displayed a high percentage of motility over a wide range of pH (6.0-8.5). Sperm motility could be initiated in Ca2+-free solutions. In contrast, sperm motility was inhibited (P<0.01) by solutions containing > or =10 mM Ca2+, and sperm could not be reactivated by a Ca2+-free solution. This Ca2+ inhibition was not affected by verapamil, a Ca2+ channel blocker. However, if sperm motility was first initiated in a Ca2+-free solution, the addition of Ca2+ solutions, up to 80 mM, failed to inhibit sperm motility, suggesting that Ca2+ inhibited the initiation of motility, but had no control of motile spermatozoa. Mg2+ solutions had similar inhibitory effects on sperm motility as Ca2+ solutions. Therefore, initiation of motility in striped bass sperm may be related to voltage-gated channels across the cell's plasma membrane. Membrane permeable cAMP did not initiate motility of quiescent, intact striped bass spermatozoa, and motility of demembranated sperm could be activated in the absence of cAMP.

  19. 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

  20. Slow motility in hair cells of the frog amphibian papilla: Ca2+-dependent shape changes.

    PubMed

    Farahbakhsh, Nasser A; Narins, Peter M

    2006-02-01

    We investigated the process of slow motility in non-mammalian auditory hair cells by recording the time course of shape change in hair cells of the frog amphibian papilla. The tall hair cells in the rostral segment of this organ, reported to be the sole recipients of efferent innervation, were found to shorten in response to an increase in the concentration of the intracellular free calcium. These shortenings are composed of two partially-overlapping phases: an initial rapid iso-volumetric contraction, followed by a slower length decrease accompanied with swelling. It is possible to unmask the iso-volumetric contraction by delaying the cell swelling with the help of K+ or Cl- channel inhibitors, quinine or furosemide. Furthermore, it appears that the longitudinal contraction in these cells is Ca2+-calmodulin-dependent: in the presence of W-7, a calmodulin inhibitor, only a slow, swelling phase could be observed. These findings suggest that amphibian rostral AP hair cells resemble their mammalian counterparts in expressing both a Ca2+-calmodulin-dependent contractile structure and an "osmotic" mechanism capable of mediating length change in response to extracellular stimuli. Such a mechanism might be utilized by the efferent neurotransmitters for adaptive modulation of mechano-electrical transduction, sensitivity enhancement, frequency selectivity, and protection against over-stimulation.

  1. [Cell growth and motility in culture (in vitro) under microgravity conditions. The Fibroblast Experiment].

    PubMed

    Tairbekov, M G; Margolis, L B; Baĭbakov, B A; Gabova, A V; Dergacheva, G B

    1994-01-01

    The experiment "Fibroblast" was performed in 1992 on biosatellite "Cosmos-2229" in onboard device "Biobox" designed by the order of European Space Agency. The main objective was elucidation of the mechanisms responsible for the effect of space flight factors, mostly microgravity, on cell culture. We studied time-related changes in growth, motility and some morphological characteristics of the cells in monolayer cultures on a solid substrate and in three-dimensional cultures supported by sponge gels. Studies were carried out on connective tissue cells isolated from the mouse embryos. Comparative after-flight analysis of the cell cultures exposed to space flight and of those under the normal gravity conditions (1 g) on the Earth has revealed some differences. The space flight conditions, mainly microgravity, induced marked changes in morphological characteristics and functional activity of the cultured fibroblasts: changes in the nucleus size and shape, retardation of cell growth and division rate. We believe that these changes may be due to weakening of intercellular contacts and cell adhesion to the substrate. These findings are important both for general biology and space medicine, specifically for the problems of tissue regeneration and wound healing under the conditions of long-term space flight.

  2. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture.

    PubMed

    Lestard, Nathalia R; Capella, Marcia A M

    2016-01-01

    Although music is part of virtually all cultures in the world, little is known about how it affects us. Since the beginning of this century several studies suggested that the response to music, and to sound in general, is complex and might not be exclusively due to emotion, given that cell types other than auditory hair cells can also directly react to audible sound. The present study was designed to better understand the direct effects of acoustic vibrations, in the form of music, in human cells in culture. Our results suggest that the mechanisms of cell growth arrest and/or cell death induced by acoustic vibrations are similar for auditory and nonauditory cells. PMID:27478480

  3. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture

    PubMed Central

    Lestard, Nathalia R.

    2016-01-01

    Although music is part of virtually all cultures in the world, little is known about how it affects us. Since the beginning of this century several studies suggested that the response to music, and to sound in general, is complex and might not be exclusively due to emotion, given that cell types other than auditory hair cells can also directly react to audible sound. The present study was designed to better understand the direct effects of acoustic vibrations, in the form of music, in human cells in culture. Our results suggest that the mechanisms of cell growth arrest and/or cell death induced by acoustic vibrations are similar for auditory and nonauditory cells. PMID:27478480

  4. Contact inhibition of locomotion determines cell-cell and cell-substrate forces in tissues.

    PubMed

    Zimmermann, Juliane; Camley, Brian A; Rappel, Wouter-Jan; Levine, Herbert

    2016-03-01

    Cells organized in tissues exert forces on their neighbors and their environment. Those cellular forces determine tissue homeostasis as well as reorganization during embryonic development and wound healing. To understand how cellular forces are generated and how they can influence the tissue state, we develop a particle-based simulation model for adhesive cell clusters and monolayers. Cells are contractile, exert forces on their substrate and on each other, and interact through contact inhibition of locomotion (CIL), meaning that cell-cell contacts suppress force transduction to the substrate and propulsion forces align away from neighbors. Our model captures the traction force patterns of small clusters of nonmotile cells and larger sheets of motile Madin-Darby canine kidney (MDCK) cells. In agreement with observations in a spreading MDCK colony, the cell density in the center increases as cells divide and the tissue grows. A feedback between cell density, CIL, and cell-cell adhesion gives rise to a linear relationship between cell density and intercellular tensile stress and forces the tissue into a nonmotile state characterized by a broad distribution of traction forces. Our model also captures the experimentally observed tissue flow around circular obstacles, and CIL accounts for traction forces at the edge. PMID:26903658

  5. Human Umbilical Cord Perivascular Cells Exhibited Enhanced Migration Capacity towards Hepatocellular Carcinoma in Comparison with Bone Marrow Mesenchymal Stromal Cells: A Role for Autocrine Motility Factor Receptor

    PubMed Central

    Aquino, Jorge B.; Malvicini, Mariana; Bolontrade, Marcela; Podhajcer, Osvaldo; Garcia, Mariana G.; Mazzolini, Guillermo

    2014-01-01

    Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. Unfortunately, the incidence and mortality associated with HCC are increasing. Therefore, new therapeutic strategies are urgently needed and the use of mesenchymal stromal cells (MSCs) as carrier of therapeutic genes is emerging as a promising option. Different sources of MSCs are being studied for cell therapy and bone marrow-derived cells are the most extensively explored; however, birth associated-tissues represent a very promising source. The aim of this work was to compare the in vitro and in vivo migration capacity between bone marrow MSCs (BM-MSCs) and human umbilical cord perivascular cells (HUCPVCs) towards HCC. We observed that HUCPVCs presented higher in vitro and in vivo migration towards factors released by HCC. The expression of autocrine motility factor (AMF) receptor, genes related with the availability of the receptor on the cell surface (caveolin-1 and -2) and metalloproteinase 3, induced by the receptor activation and important for cell migration, was increased in HUCPVCs. The chemotactic response towards recombinant AMF was increased in HUCPVCs compared to BM-MSCs, and its inhibition in the conditioned medium from HCC induced higher decrease in HUCPVC migration than in BM-MSC. Our results indicate that HUCPVCs could be a useful cellular source to deliver therapeutic genes to HCC. PMID:25147818

  6. Human umbilical cord perivascular cells exhibited enhanced migration capacity towards hepatocellular carcinoma in comparison with bone marrow mesenchymal stromal cells: a role for autocrine motility factor receptor.

    PubMed

    Bayo, Juan; Fiore, Esteban; Aquino, Jorge B; Malvicini, Mariana; Rizzo, Manglio; Peixoto, Estanislao; Alaniz, Laura; Piccioni, Flavia; Bolontrade, Marcela; Podhajcer, Osvaldo; Garcia, Mariana G; Mazzolini, Guillermo

    2014-01-01

    Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. Unfortunately, the incidence and mortality associated with HCC are increasing. Therefore, new therapeutic strategies are urgently needed and the use of mesenchymal stromal cells (MSCs) as carrier of therapeutic genes is emerging as a promising option. Different sources of MSCs are being studied for cell therapy and bone marrow-derived cells are the most extensively explored; however, birth associated-tissues represent a very promising source. The aim of this work was to compare the in vitro and in vivo migration capacity between bone marrow MSCs (BM-MSCs) and human umbilical cord perivascular cells (HUCPVCs) towards HCC. We observed that HUCPVCs presented higher in vitro and in vivo migration towards factors released by HCC. The expression of autocrine motility factor (AMF) receptor, genes related with the availability of the receptor on the cell surface (caveolin-1 and -2) and metalloproteinase 3, induced by the receptor activation and important for cell migration, was increased in HUCPVCs. The chemotactic response towards recombinant AMF was increased in HUCPVCs compared to BM-MSCs, and its inhibition in the conditioned medium from HCC induced higher decrease in HUCPVC migration than in BM-MSC. Our results indicate that HUCPVCs could be a useful cellular source to deliver therapeutic genes to HCC.

  7. Optical trapping and manipulation of single cells and motile microorganisms by laser diode radiation

    NASA Astrophysics Data System (ADS)

    Frediani, Carlo; Ascoli, Cesare; Lucia, S.; Verkerk, P.; Guidoni, L.; Fioretti, A.; Arimondo, Ennio

    1994-12-01

    Electromagnetic fields, emitted by laser sources, have been utilized in recent years for controlling the position and velocity of atoms, ions and microscopic neutral particles. In 1987 Ashkin has shown for the first time that cells too can be trapped by using a focused beam of laser radiation. The trapping is due to the interation between the electric dipole induced by the laser electric field in the cell and the electric field itself. In order to maximize the trapping effect and to avoid damage to the cells caused by excessive heating, the laser wavelength must be far from the absorption bands for both the cells and the solution where cells are kept, usually water. Our preliminary experiments, utilizing a 100 mW laser diode at 850 nm with suitable focusing, show that also free swimming (up to 100 micrometers /s) protozoa (Dunaliella salina) can be easily trapped, without apparent damage. The experimental set-up and the experiments on motile micro-organisms are presented. Possible biomedical applications are discussed.

  8. Cancer progression and tumor cell motility are associated with the FGFR4 Arg(388) allele.

    PubMed

    Bange, Johannes; Prechtl, Dieter; Cheburkin, Yuri; Specht, Katja; Harbeck, Nadia; Schmitt, Manfred; Knyazeva, Tatjana; Müller, Susanne; Gärtner, Silvia; Sures, Irmi; Wang, Hongyang; Imyanitov, Evgeny; Häring, Hans-Ulrich; Knayzev, Pjotr; Iacobelli, Stefano; Höfler, Heinz; Ullrich, Axel

    2002-02-01

    Expression analysis of genes encoding components of the phosphotyrosine signaling system by cDNA array hybridization revealed elevated levels of FGFR4 transcripts in several mammary carcinoma cell lines. In the FGFR4 gene transcript from MDA-MB-453 mammary carcinoma cells, a G to A conversion was discovered that results in the substitution of glycine by arginine at position 388 in the transmembrane domain of the receptor. The Arg(388) allele was also found in cell lines derived from a variety of other tumor types as well as in the germ-line of cancer patients and healthy individuals. Analysis of three geographically separated groups indicated that it occurs in approximately 50% of the human population. Investigation of the clinical data of 84 breast cancer patients revealed that homo- or heterozygous carriers of the Arg(388) allele had a significantly reduced disease-free survival time (P = 0.01) within a median follow-up of 62 months. Moreover, the FGFR4 Arg(388) allele was associated with early lymph node metastasis and advanced tumor-node-metastasis (TNM) stage in 82 colon cancer patients. Consistent with this finding, MDA-MB-231 mammary tumor cells expressing FGFR4 Arg(388) exhibited increased motility relative to cells expressing the FGFR4 Gly(388) isotype. Our results support the conclusion that the FGFR4 Arg(388) allele represents a determinant that is innocuous in healthy individuals but predisposes cancer patients for significantly accelerated disease progression.

  9. Androgens Regulate T47D Cells Motility and Invasion through Actin Cytoskeleton Remodeling

    PubMed Central

    Montt-Guevara, Maria Magdalena; Shortrede, Jorge Eduardo; Giretti, Maria Silvia; Giannini, Andrea; Mannella, Paolo; Russo, Eleonora; Genazzani, Alessandro David; Simoncini, Tommaso

    2016-01-01

    The relationship between androgens and breast cancer is controversial. Androgens have complex effects on breast cancer progression and metastasis. Moreover, androgen receptor (AR) is expressed in approximately 70 to 90% of invasive breast carcinomas, which has prognostic relevance in basal-like cancers and in triple-negative breast cancers. Recent studies have associated the actin-binding proteins of the ezrin–radixin–moesin (ERM) family with metastasis in endocrine-sensitive cancers. We studied on T47D breast cancer cells whether androgens with different characteristics, such as testosterone (T), dihydrotestosterone (DHT), and dehydroepiandrosterone (DHEA) may regulate breast cancer cell motility and invasion through the control of actin remodeling. We demonstrate that androgens promote migration and invasion in T47D via Moesin activation. We show that T and DHEA exert their actions via the AR and estrogen receptor (ER), while the non-aromatizable androgen – DHT – only recruits AR. We further report that androgen induced significant changes in actin organization with pseudopodia along with membrane ruffles formation, and this process is mediated by Moesin. Our work identifies novel mechanisms of action of androgens on breast cancer cells. Through the modulation of Moesin, androgens alter the architecture of cytoskeleton in T47D breast cancer cell and promote cell migration and invasion. These results could help to understand the biological actions of androgens on breast cancer and, eventually, to develop new strategies for breast cancer treatment. PMID:27746764

  10. 4D Tumorigenesis Model for Quantitating Coalescence, Directed Cell Motility and Chemotaxis, Identifying Unique Cell Behaviors, and Testing Anticancer Drugs.

    PubMed

    Kuhl, Spencer; Voss, Edward; Scherer, Amanda; Lusche, Daniel F; Wessels, Deborah; Soll, David R

    2016-01-01

    A 4D high-resolution computer-assisted reconstruction and motion analysis system has been developed and applied to the long-term (14-30 days) analysis of cancer cells migrating and aggregating within a 3D matrix. 4D tumorigenesis models more closely approximate the tumor microenvironment than 2D substrates and, therefore, are improved tools for elucidating the interactions within the tumor microenvironment that promote growth and metastasis. The model we describe here can be used to analyze the growth of tumor cells, aggregate coalescence, directed cell motility and chemotaxis, matrix degradation, the effects of anticancer drugs, and the behavior of immune and endothelial cells mixed with cancer cells. The information given in this chapter is also intended to acquaint the reader with computer-assisted methods and algorithms that can be used for high-resolution 3D reconstruction and quantitative motion analysis. PMID:27271907

  11. Quantitative analysis of signal transduction in motile and phototactic cells by computerized light stimulation and model based tracking

    NASA Astrophysics Data System (ADS)

    Streif, Stefan; Staudinger, Wilfried Franz; Oesterhelt, Dieter; Marwan, Wolfgang

    2009-02-01

    To investigate the responses of Halobacterium salinarum to stimulation with light (phototaxis and photokinesis), we designed an experimental setup consisting of optical devices for automatic video image acquisition and computer-controlled light stimulation, and developed algorithms to analyze physiological responses of the cells. Cells are categorized as motile and nonmotile by a classification scheme based on the square displacement of cell positions. Computerized tracking based on a dynamic model of the stochastic cell movement and a Kalman filter-based algorithm allows smoothed estimates of the cell tracks and the detection of physiological responses to complex stimulus patterns. The setup and algorithms were calibrated which allows quantitative measurements and systematic analysis of cellular sensing and response. Overall, the setup is flexible, extensible, and consists mainly of commercially available products. This facilitates modifications of the setup and algorithms for physiological studies of the motility of cells or microorganisms.

  12. Autocrine motility factor receptor promotes the proliferation of human acute monocytic leukemia THP-1 cells

    PubMed Central

    WANG, YINGCHAO; MA, LINA; WANG, CHUNMEI; SHENG, GUANGYAO; FENG, LEI; YIN, CHUYUN

    2015-01-01

    The aberrant activation of autocrine motility factor receptor (AMFR) has been implicated in several types of human cancer. The present study aimed to elucidate the effect of AMFR on the regulation of proliferation in an acute monocytic leukemia cell line, THP-1. THP-1 cells were transfected with AMFR-targeted small interfering (si)RNA and a plasmid encoding a truncated AMFR, AMFR-C, (pcDNA3.1-AMFR-C). The mRNA and protein levels of AMFR and the downstream targets, rho-associated, coiled-coil containing protein kinase 2 (ROCK2), cyclin D1, and B-cell lymphoma (Bcl)-2, were measured using reverse transcription-quantitatibe polymerase chain reaction and immunoblot analyses. The effects on cell cycle and apoptosis were investigated using flow cytometry. The present study successfully established the knockdown of AMFR and expression of AMFR-C in the THP-1 cells. Downregulation of AMFR induced cell cycle arrest at the G0/G1 phase, and increased apoptosis of the THP-1 cells (all P<0.05). The AMFR siRNA increased the percentage of early apoptotic cells between 3.88±1.43 and 19.58±4.29% (P<0.05). The expression levels of ROCK2, cyclin D1 and Bcl-2 were reduced by the downregulation of AMFR and enhanced by overexpression of AMFR-C. In conclusion, AMFR appears to be crucial for the proliferation of the THP-1 acute monocytic leukemia cell line. Therefore, AMFR may represent a potential target for the treatment of acute monocytic leukemia. PMID:26136223

  13. Cell motility and biofilm formation in Bacillus subtilis are affected by the ribosomal proteins, S11 and S21.

    PubMed

    Takada, Hiraku; Morita, Masato; Shiwa, Yuh; Sugimoto, Ryoma; Suzuki, Shota; Kawamura, Fujio; Yoshikawa, Hirofumi

    2014-01-01

    Bacillus subtilis differentiates into various cellular states in response to environmental changes. It exists in two states during the exponential growth phase: motile cells and connected chains of sessile cells. Here, we identified new regulators of cell motility and chaining, the ribosomal proteins S21 (rpsU) and S11 (rpsK). Their mutants showed impaired cell motility (observed in a laboratory strain) and robust biofilm formation (observed in an undomesticated strain). The two major operons for biofilm formation, tapA-sipW-tasA and epsA-O, were strongly expressed in the rpsU mutant, whereas the flagellin-encoding hag gene and other SigD-dependent motility regulons were not. Genetic analysis revealed that the mutation of remA, the transcriptional activator of the eps operon, is epistatic to that of rpsU, whereas the mutation of antagonistic regulators of SinR is not. Our studies demonstrate that S11 and S21 participate in the regulation of bistability via the RemA/RemB pathway.

  14. Modulation of Intracellular Calcium Levels by Calcium Lactate Affects Colon Cancer Cell Motility through Calcium-Dependent Calpain

    PubMed Central

    Sundaramoorthy, Pasupathi; Sim, Jae Jun; Jang, Yeong-Su; Mishra, Siddhartha Kumar; Jeong, Keun-Yeong; Mander, Poonam; Chul, Oh Byung; Shim, Won-Sik; Oh, Seung Hyun; Nam, Ky-Youb; Kim, Hwan Mook

    2015-01-01

    Cancer cell motility is a key phenomenon regulating invasion and metastasis. Focal adhesion kinase (FAK) plays a major role in cellular adhesion and metastasis of various cancers. The relationship between dietary supplementation of calcium and colon cancer has been extensively investigated. However, the effect of calcium (Ca2+) supplementation on calpain-FAK-motility is not clearly understood. We sought to identify the mechanism of FAK cleavage through Ca2+ bound lactate (CaLa), its downstream signaling and role in the motility of human colon cancer cells. We found that treating HCT116 and HT-29 cells with CaLa immediately increased the intracellular Ca2+ (iCa2+) levels for a prolonged period of time. Ca2+ influx induced cleavage of FAK into an N-terminal FAK (FERM domain) in a dose-dependent manner. Phosphorylated FAK (p-FAK) was also cleaved in to its p-N-terminal FAK. CaLa increased colon cancer cells motility. Calpeptin, a calpain inhibitor, reversed the effects of CaLa on FAK and pFAK cleavage in both cancer cell lines. The cleaved FAK translocates into the nucleus and modulates p53 stability through MDM2-associated ubiquitination. CaLa-induced Ca2+ influx increased the motility of colon cancer cells was mediated by calpain activity through FAK and pFAK protein destabilization. In conclusion, these results suggest that careful consideration may be given in deciding dietary Ca2+ supplementation to patient undergoing treatment for metastatic cancer. PMID:25629974

  15. Motility-dependence of the heterogenous staining of culture cells by a monoclonal anti-tropomyosin antibody

    PubMed Central

    1988-01-01

    A monoclonal antibody (CG1) which recognizes tropomyosin isoforms 1 and 3 of chicken embryo fibroblasts was used to detect what is a motility- dependent change in the availability of the antigenic determinant in tropomyosin molecules along microfilaments. Immunofluorescence microscopy with this antibody revealed a heterogenous staining pattern among chicken embryo fibroblasts cells such that a population (17%) of cells showed only background staining. Stress fibers in about half the population of the cells stained weakly with this antibody, while the stress fibers in another population of cells (35%) showed very strong staining. After glycerination or cytochalasin B treatment, all of the cells became positive in reaction to CG1 antibody, suggesting that the antigenic determinant was present in every cell. On the other hand, all of the cells after brief nonionic detergent treatment became negative to CG1 antibody. The CG1 staining pattern was not significantly changed in cells at different stages after release from colcemid blockage, nor was a brief treatment of cells with buffer containing 2 M urea, mild trypsin, chymotrypsin, or V.8 protease effective in changing the reactivity. However, most of the cells with a morphology typical of movement, and all of the contracted, glycerinated cells were strongly positive to CG1 antibody. These results suggest that the unmasking of the CG1 determinant may be motility-dependent. Immunoblot analysis showed that forced modification on the cysteine residue of tropomyosin molecules, caused either by performic acid oxidation or by disulfide cross-linking with the chemical 5,5'-dithiobis (2-nitrobenzoate), results in drastic changes in the reactivity of the different isoforms to CG1 antibody. These results indicate that the cysteine residue is involved in the CG1 determinant. The motility-dependent unmasking of this determinant may suggest an important role for nonmuscle tropomyosin in regulating cell motility. PMID:2448313

  16. Simulating the Complex Cell Design of Trypanosoma brucei and Its Motility

    PubMed Central

    Alizadehrad, Davod; Krüger, Timothy; Engstler, Markus; Stark, Holger

    2015-01-01

    The flagellate Trypanosoma brucei, which causes the sleeping sickness when infecting a mammalian host, goes through an intricate life cycle. It has a rather complex propulsion mechanism and swims in diverse microenvironments. These continuously exert selective pressure, to which the trypanosome adjusts with its architecture and behavior. As a result, the trypanosome assumes a diversity of complex morphotypes during its life cycle. However, although cell biology has detailed form and function of most of them, experimental data on the dynamic behavior and development of most morphotypes is lacking. Here we show that simulation science can predict intermediate cell designs by conducting specific and controlled modifications of an accurate, nature-inspired cell model, which we developed using information from live cell analyses. The cell models account for several important characteristics of the real trypanosomal morphotypes, such as the geometry and elastic properties of the cell body, and their swimming mechanism using an eukaryotic flagellum. We introduce an elastic network model for the cell body, including bending rigidity and simulate swimming in a fluid environment, using the mesoscale simulation technique called multi-particle collision dynamics. The in silico trypanosome of the bloodstream form displays the characteristic in vivo rotational and translational motility pattern that is crucial for survival and virulence in the vertebrate host. Moreover, our model accurately simulates the trypanosome's tumbling and backward motion. We show that the distinctive course of the attached flagellum around the cell body is one important aspect to produce the observed swimming behavior in a viscous fluid, and also required to reach the maximal swimming velocity. Changing details of the flagellar attachment generates less efficient swimmers. We also simulate different morphotypes that occur during the parasite's development in the tsetse fly, and predict a flagellar

  17. 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

  18. Special type of morphological reorganization induced by phorbol ester: reversible partition of cell into motile and stable domains

    SciTech Connect

    Dugina, V.B.; Svitkina, T.M.; Vasiliev, J.M.; Gelfand, I.M.

    1987-06-01

    The phorbol ester phorbol 12-myristate 13-acetate (PMA) induced reversible alteration of the shape of fibroblastic cells of certain transformed lines-namely, partition of the cells into two types of domains: motile body actively extending large lamellas and stable narrow cytoplasmic processes. Dynamic observations have shown that stable processes are formed from partially retracted lamellas and from contracted tail parts of cell bodies. Immunofluorescence microscopy and electron microscopy of platinum replicas of cytoskeleton have shown that PMA-induced narrow processes are rich in microtubules and intermediate filaments but relatively poor in actin microfilaments; in contrast, lamellas and cell bodies contained numerous microfilaments. Colcemid-induced depolymerization of microtubules led to contraction of PMA-induced processes; cytochalasin B prevented this contraction. It is suggested that PMA-induced separation of cell into motile and stable parts is due to directional movement of actin structures along the microtubular framework. Similar movements may play an important role in various normal morphogenetic processes.

  19. Live-Cell Imaging of Phagosome Motility in Primary Mouse RPE Cells.

    PubMed

    Hazim, Roni; Jiang, Mei; Esteve-Rudd, Julian; Diemer, Tanja; Lopes, Vanda S; Williams, David S

    2016-01-01

    The retinal pigment epithelium (RPE) is a post-mitotic epithelial monolayer situated between the light-sensitive photoreceptors and the choriocapillaris. Given its vital functions for healthy vision, the RPE is a primary target for insults that result in blinding diseases, including age-related macular degeneration (AMD). One such function is the phagocytosis and digestion of shed photoreceptor outer segments. In the present study, we examined the process of trafficking of outer segment disk membranes in live cultures of primary mouse RPE, using high speed spinning disk confocal microscopy. This approach has enabled us to track phagosomes, and determine parameters of their motility, which are important for their efficient degradation.

  20. Individual motile CD4+ T cells can participate in efficient multi-killing through conjugation to multiple tumor cells

    PubMed Central

    Liadi, Ivan; Singh, Harjeet; Romain, Gabrielle; Rey-Villamizar, Nicolas; Merouane, Amine; Adolacion, Jay R T.; Kebriaei, Partow; Huls, Helen; Qiu, Peng; Roysam, Badrinath; Cooper, Laurence J.N.; Varadarajan, Navin

    2015-01-01

    T cells genetically modified to express a CD19-specific chimeric antigen receptor (CAR) for the investigational treatment of B-cell malignancies comprise a heterogeneous population, and their ability to persist and participate in serial killing of tumor cells is a predictor of therapeutic success. We implemented Timelapse Imaging Microscopy In Nanowell Grids (TIMING) to provide direct evidence that CD4+CAR+ T cells (CAR4 cells) can engage in multi-killing via simultaneous conjugation to multiple tumor cells. Comparisons of the CAR4 cells and CD8+CAR+ T cells (CAR8 cells) demonstrate that while CAR4 cells can participate in killing and multi-killing, they do so at slower rates, likely due to the lower Granzyme B content. Significantly, in both sets of T cells, a minor sub-population of individual T cells identified by their high motility, demonstrated efficient killing of single tumor cells. By comparing both the multi-killer and single killer CAR+ T cells it appears that the propensity and kinetics of T-cell apoptosis was modulated by the number of functional conjugations. T cells underwent rapid apoptosis, and at higher frequencies, when conjugated to single tumor cells in isolation and this effect was more pronounced on CAR8 cells. Our results suggest that the ability of CAR+ T cells to participate in multi-killing should be evaluated in the context of their ability to resist activation induced cell death (AICD). We anticipate that TIMING may be utilized to rapidly determine the potency of T-cell populations and may facilitate the design and manufacture of next-generation CAR+ T cells with improved efficacy. PMID:25711538

  1. HMGB1 induces the overexpression of miR-222 and miR-221 and increases growth and motility in papillary thyroid cancer cells.

    PubMed

    Mardente, Stefania; Mari, Emanuela; Consorti, Fabrizio; Di Gioia, Cira; Negri, Rodolfo; Etna, Marilena; Zicari, Alessandra; Antonaci, Alfredo

    2012-12-01

    Experimental and epidemiological studies have revealed that chronic inflammation contributes to cancer progression and even predisposes to cellular transformation. Inflammatory infiltrates in papillary thyroid cancer include lymphocytes, macrophages and cytokines. High-mobility group box 1 protein (HMGB1) is a late inflammatory cytokine that signals danger to the immune system through the receptor for advanced glycation end-products (RAGE) and Toll-like receptor. The activation of the above receptors results in the secretion of growth, chemotactic and angiogenic factors that contribute to chronic inflammation. In this study, we suggest that apart from the activation of signal transduction pathways by the activation of RAGE, the indirect inhibition of cell cycle regulators [such as phosphatase and tensin homolog (PTEN)] may also cause an increase in cell growth and motility. MicroRNAs (miRNAs) have increasingly been implicated in regulating the malignant progression of cancer. MiR-221 and miR-222 have been found to be deregulated in human papillary thyroid carcinomas. They are involved in cell proliferation through the inhibition of the cell cycle regulator, p27kip1, in human papillary carcinomas. In this study, we show that HMGB1 increases the expression of miR-221 and miR-222 in primary cultures of excised papillary lesions and in an established papillary cancer cell line (BC PAP). The overexpression of oncogenic miR-221 and miR-222 caused by HMGB1 is associated with an increase in malignancy scores, namely cell growth and motility.

  2. Seizure-Induced Motility of Differentiated Dentate Granule Cells Is Prevented by the Central Reelin Fragment.

    PubMed

    Orcinha, Catarina; Münzner, Gert; Gerlach, Johannes; Kilias, Antje; Follo, Marie; Egert, Ulrich; Haas, Carola A

    2016-01-01

    Granule cell dispersion (GCD) represents a pathological widening of the granule cell layer in the dentate gyrus and it is frequently observed in patients with mesial temporal lobe epilepsy (MTLE). Recent studies in human MTLE specimens and in animal epilepsy models have shown that a decreased expression and functional inactivation of the extracellular matrix protein Reelin correlates with GCD formation, but causal evidence is still lacking. Here, we used unilateral kainate (KA) injection into the mouse hippocampus, an established MTLE animal model, to precisely map the loss of reelin mRNA-synthesizing neurons in relation to GCD along the septotemporal axis of the epileptic hippocampus. We show that reelin mRNA-producing neurons are mainly lost in the hilus and that this loss precisely correlates with the occurrence of GCD. To monitor GCD formation in real time, we used organotypic hippocampal slice cultures (OHSCs) prepared from mice which express enhanced green fluorescent protein (eGFP) primarily in differentiated dentate granule cells. Using life cell microscopy we observed that increasing doses of KA resulted in an enhanced motility of eGFP-positive granule cells. Moreover, KA treatment of OHSC resulted in a rapid loss of Reelin-producing interneurons mainly in the hilus, as observed in vivo. A detailed analysis of the migration behavior of individual eGFP-positive granule cells revealed that the majority of these neurons actively migrate toward the hilar region, where Reelin-producing neurons are lost. Treatment with KA and subsequent addition of the recombinant R3-6 Reelin fragment significantly prevented the movement of eGFP-positive granule cells. Together, these findings suggest that GCD formation is indeed triggered by a loss of Reelin in hilar interneurons. PMID:27516734

  3. Seizure-Induced Motility of Differentiated Dentate Granule Cells Is Prevented by the Central Reelin Fragment

    PubMed Central

    Orcinha, Catarina; Münzner, Gert; Gerlach, Johannes; Kilias, Antje; Follo, Marie; Egert, Ulrich; Haas, Carola A.

    2016-01-01

    Granule cell dispersion (GCD) represents a pathological widening of the granule cell layer in the dentate gyrus and it is frequently observed in patients with mesial temporal lobe epilepsy (MTLE). Recent studies in human MTLE specimens and in animal epilepsy models have shown that a decreased expression and functional inactivation of the extracellular matrix protein Reelin correlates with GCD formation, but causal evidence is still lacking. Here, we used unilateral kainate (KA) injection into the mouse hippocampus, an established MTLE animal model, to precisely map the loss of reelin mRNA-synthesizing neurons in relation to GCD along the septotemporal axis of the epileptic hippocampus. We show that reelin mRNA-producing neurons are mainly lost in the hilus and that this loss precisely correlates with the occurrence of GCD. To monitor GCD formation in real time, we used organotypic hippocampal slice cultures (OHSCs) prepared from mice which express enhanced green fluorescent protein (eGFP) primarily in differentiated dentate granule cells. Using life cell microscopy we observed that increasing doses of KA resulted in an enhanced motility of eGFP-positive granule cells. Moreover, KA treatment of OHSC resulted in a rapid loss of Reelin-producing interneurons mainly in the hilus, as observed in vivo. A detailed analysis of the migration behavior of individual eGFP-positive granule cells revealed that the majority of these neurons actively migrate toward the hilar region, where Reelin-producing neurons are lost. Treatment with KA and subsequent addition of the recombinant R3–6 Reelin fragment significantly prevented the movement of eGFP-positive granule cells. Together, these findings suggest that GCD formation is indeed triggered by a loss of Reelin in hilar interneurons. PMID:27516734

  4. Seizure-Induced Motility of Differentiated Dentate Granule Cells Is Prevented by the Central Reelin Fragment.

    PubMed

    Orcinha, Catarina; Münzner, Gert; Gerlach, Johannes; Kilias, Antje; Follo, Marie; Egert, Ulrich; Haas, Carola A

    2016-01-01

    Granule cell dispersion (GCD) represents a pathological widening of the granule cell layer in the dentate gyrus and it is frequently observed in patients with mesial temporal lobe epilepsy (MTLE). Recent studies in human MTLE specimens and in animal epilepsy models have shown that a decreased expression and functional inactivation of the extracellular matrix protein Reelin correlates with GCD formation, but causal evidence is still lacking. Here, we used unilateral kainate (KA) injection into the mouse hippocampus, an established MTLE animal model, to precisely map the loss of reelin mRNA-synthesizing neurons in relation to GCD along the septotemporal axis of the epileptic hippocampus. We show that reelin mRNA-producing neurons are mainly lost in the hilus and that this loss precisely correlates with the occurrence of GCD. To monitor GCD formation in real time, we used organotypic hippocampal slice cultures (OHSCs) prepared from mice which express enhanced green fluorescent protein (eGFP) primarily in differentiated dentate granule cells. Using life cell microscopy we observed that increasing doses of KA resulted in an enhanced motility of eGFP-positive granule cells. Moreover, KA treatment of OHSC resulted in a rapid loss of Reelin-producing interneurons mainly in the hilus, as observed in vivo. A detailed analysis of the migration behavior of individual eGFP-positive granule cells revealed that the majority of these neurons actively migrate toward the hilar region, where Reelin-producing neurons are lost. Treatment with KA and subsequent addition of the recombinant R3-6 Reelin fragment significantly prevented the movement of eGFP-positive granule cells. Together, these findings suggest that GCD formation is indeed triggered by a loss of Reelin in hilar interneurons.

  5. Heparin-binding proteins HB-GAM (pleiotrophin) and amphoterin in the regulation of cell motility.

    PubMed

    Rauvala, H; Huttunen, H J; Fages, C; Kaksonen, M; Kinnunen, T; Imai, S; Raulo, E; Kilpeläinen, I

    2000-09-01

    Fractionation of proteins from perinatal rat brain was monitored using a neurite outgrowth assay. Two neurite-promoting proteins, HB-GAM (heparin-binding growth-associated molecule; also known as pleiotrophin) and amphoterin, were isolated, cloned and produced by baculovirus expression for structural and functional studies. HB-GAM is highly expressed in embryonic and early post-natal fiber pathways of the nervous system, and it enhances axonal growth/guidance by binding to N-syndecan (syndecan-3) at the neuron surface. N-syndecan in turn communicates with the cytoskeleton through the cortactin/src-kinase pathway to enhance neurite extension. In addition to N-syndecan, the chondroitin sulfate proteoglycan RPTP beta/zeta (receptor-type tyrosine phosphatase beta/zeta) is implicated in the receptor mechanism of HB-GAM. HB-GAM is also prominently expressed in developing and regenerating bone as a matrix-bound cue for migration of osteoblasts/osteoblast precursors to the site of bone deposition. HB-GAM is suggested to regulate motility in osteoblasts through a similar mechanism as in neurons. Structural studies using heteronuclear NMR reveal two similar protein domains in HB-GAM, both consisting of three anti-parallel beta-strands. Search of sequence databases shows that the beta structures of HB-GAM and of the similar domains of MK (midkine) correspond to the thrombospondin type I (TSR) sequence motif. We suggest that the TSR sequence motif, found in several neurite outgrowth-promoting and other cell surface and matrix-binding proteins, defines a beta structure similar to those found in HB-GAM and MK. In general, amphoterin is highly expressed in immature and transformed cells. We suggest a model, according to which amphoterin is an autocrine/paracrine regulator of invasive migration. Amphoterin binds to RAGE (receptor of advanced glycation end products), an immunoglubulin superfamily member related to N-CAM (neural cell adhesion molecule), that communicates with the

  6. Reduced Contractility and Motility of Prostatic Cancer-Associated Fibroblasts after Inhibition of Heat Shock Protein 90

    PubMed Central

    Henke, Alex; Franco, Omar E.; Stewart, Grant D.; Riddick, Antony C.P.; Katz, Elad; Hayward, Simon W.; Thomson, Axel A.

    2016-01-01

    Background: Prostate cancer-associated fibroblasts (CAF) can stimulate malignant progression and invasion of prostatic tumour cells via several mechanisms including those active in extracellular matrix; Methods: We isolated CAF from prostate cancer patients of Gleason Score 6–10 and confirmed their cancer-promoting activity using an in vivo tumour reconstitution assay comprised of CAF and BPH1 cells. We tested the effects of heat shock protein 90 (HSP90) inhibitors upon reconstituted tumour growth in vivo. Additionally, CAF contractility was measured in a 3D collagen contraction assay and migration was measured by scratch assay; Results: HSP90 inhibitors dipalmitoyl-radicicol and 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) reduced tumour size and proliferation in CAF/BPH1 reconstituted tumours in vivo. We observed that the most contractile CAF were derived from patients with lower Gleason Score and of younger age compared with the least contractile CAF. HSP90 inhibitors radicicol and 17-DMAG inhibited contractility and reduced the migration of CAF in scratch assays. Intracellular levels of HSP70 and HSP90 were upregulated upon treatment with HSP90 inhibitors. Inhibition of HSP90 also led to a specific increase in transforming growth factor beta 2 (TGFβ2) levels in CAF; Conclusions: We suggest that HSP90 inhibitors act not only upon tumour cells, but also on CAF in the tumour microenvironment. PMID:27563925

  7. JNK Inhibition Inhibits Lateral Line Neuromast Hair Cell Development

    PubMed Central

    Cai, Chengfu; Lin, Jinchao; Sun, Shaoyang; He, Yingzi

    2016-01-01

    JNK signaling is known to play a role in regulating cell behaviors such as cell cycle progression, cell proliferation, and apoptosis, and recent studies have suggested important roles for JNK signaling in embryonic development. However, the precise function of JNK signaling in hair cell development remains poorly studied. In this study, we used the small molecule JNK inhibitor SP600125 to examine the effect of JNK signaling abrogation on the development of hair cells in the zebrafish lateral line neuromast. Our results showed that SP600125 reduced the numbers of both hair cells and supporting cells in neuromasts during larval development in a dose-dependent manner. Additionally, JNK inhibition strongly inhibited the proliferation of neuromast cells, which likely explains the decrease in the number of differentiated hair cells in inhibitor-treated larvae. Furthermore, western blot and in situ analysis showed that JNK inhibition induced cell cycle arrest through induction of p21 expression. We also showed that SP600125 induced cell death in developing neuromasts as measured by cleaved caspase-3 immunohistochemistry, and this was accompanied with an induction of p53 gene expression. Together these results indicate that JNK might be an important regulator in the development of hair cells in the lateral line in zebrafish by controlling both cell cycle progression and apoptosis. PMID:26903805

  8. Biochemical and Functional Characterization of the Interaction between Liprin-α1 and GIT1: Implications for the Regulation of Cell Motility

    PubMed Central

    Pettinato, Emanuela; Paris, Simona; Bachi, Angela; de Curtis, Ivan

    2011-01-01

    We have previously identified the scaffold protein liprin-α1 as an important regulator of integrin-mediated cell motility and tumor cell invasion. Liprin-α1 may interact with different proteins, and the functional significance of these interactions in the regulation of cell motility is poorly known. Here we have addressed the involvement of the liprin-α1 partner GIT1 in liprin-α1-mediated effects on cell spreading and migration. GIT1 depletion inhibited spreading by affecting the lamellipodia, and prevented liprin-α1-enhanced spreading. Conversely inhibition of the formation of the liprin-α1-GIT complex by expression of liprin-ΔCC3 could still enhance spreading, although to a lesser extent compared to full length liprin-α1. No cumulative effects were observed after depletion of both liprin-α1 and GIT1, suggesting that the two proteins belong to the same signaling network in the regulation of cell spreading. Our data suggest that liprin-α1 may compete with paxillin for binding to GIT1, while binding of βPIX to GIT1 was unaffected by the presence of liprin-α1. Interestingly, GIT and liprin-α1 reciprocally regulated their subcellular localization, since liprin-α1 overexpression, but not the GIT binding-defective liprin-ΔCC3 mutant, affected the localization of endogenous GIT at peripheral and mature central focal adhesions, while the expression of a truncated, active form of GIT1 enhanced the localization of endogenous liprin-α1 at the edge of spreading cells. Moreover, GIT1 was required for liprin-α1-enhanced haptotatic migration, although the direct interaction between liprin-α1 and GIT1 was not needed. Our findings show that the functional interaction between liprin-α1 and GIT1 cooperate in the regulation of integrin-dependent cell spreading and motility on extracellular matrix. These findings and the possible competition of liprin-α1 with paxillin for binding to GIT1 suggest that alternative binding of GIT1 to either liprin-α1 or paxillin

  9. Microfilaments and microtubules control the shape, motility, and subcellular distribution of cortical mitochondria in characean internodal cells.

    PubMed

    Foissner, I

    2004-12-01

    The shape, motility, and subcellular distribution of mitochondria in characean internodal cells were studied by visualizing fluorescent dyes with confocal laser scanning microscopy and conducting drug-inhibitor experiments. Shape, size, number, and distribution of mitochondria varied according to the growth status and the metabolic activity within the cell. Vermiform (sausage-shaped), disc-, or amoeba-like mitochondria were present in elongating internodes, whereas very young cells and older cells that had completed growth contained short, rodlike organelles only. Mitochondria were evenly distributed and passively transported in the streaming endoplasm. In the cortex, mitochondria were sandwiched between the plasma membrane and the stationary chloroplast files and distributed in relation to the pattern of pH banding. Highest mitochondrial densities were found at the acid, photosynthetically more active regions, whereas the alkaline sites contained fewer and smaller mitochondria. In the cortex of elongating cells, small mitochondria moved slowly along microtubules or actin filaments. The shape and motility of giant mitochondria depended on the simultaneous interaction with both cytoskeletal systems. There was no microtubule-dependent motility in the cortex of nonelongating mature cells and mitochondria only occasionally travelled along actin filaments. These observations suggest that mitochondria of characean internodes possess motor proteins for microtubules and actin filaments, both of which can be used either as tracks for migration or for immobilization. The cortical cytoskeleton probably controls the spatiotemporal distribution of mitochondria within the cell and promotes their association with chloroplasts, which is necessary for exchange of metabolites during photosynthesis and detoxification.

  10. Fluorescence imaging and spectroscopy of motile sperm cells and CHO cells in an optical trap (laser tweezers)

    NASA Astrophysics Data System (ADS)

    Koenig, Karsten; Liu, Yagang; Krasieva, Tatiana B.; Patrizio, Pasquale; Tadir, Yona; Sonek, Gregory J.; Berns, Michael W.; Tromberg, Bruce J.

    1995-05-01

    We describe fluorescence spectroscopy and imaging studies of optically trapped single Chinese hamster ovary (CHO) and motile human sperm cells. The NIR trapping beam was provided by a tunable, multimode continuous wave Ti:Sapphire laser. The beam was introduced into an inverted confocal laser scanning microscope. Fluorescence of cells in the single- beam gradient force optical trap was excited with a 488 nm microbeam (laser scanning microscopy) or with 365 nm radiation from a high- pressure mercury lamp. Modifications to NADH-attributed autofluorescence and Rhodamine- and Propidium Iodide-attributed xenofluorescence indicate a significant cell-damaging effect of 760 nm trapping beams. 760 nm effects produce a biological response comparable to UVA-induced oxidative stress and appear to be a consequence to two-photon absorption.

  11. Polyglycylation of Tubulin Is Essential and Affects Cell Motility and Division in Tetrahymena thermophila

    PubMed Central

    Xia, Lu; Hai, Bing; Gao, Yan; Burnette, Dylan; Thazhath, Rupal; Duan, Jianming; Bré, Marie-Helene; Levilliers, Nicolette; Gorovsky, Martin A.; Gaertig, Jacek

    2000-01-01

    We analyzed the role of tubulin polyglycylation in Tetrahymena thermophila using in vivo mutagenesis and immunochemical analysis with modification-specific antibodies. Three and five polyglycylation sites were identified at glutamic acids near the COOH termini of α- and β-tubulin, respectively. Mutants lacking all polyglycylation sites on α-tubulin have normal phenotype, whereas similar sites on β-tubulin are essential. A viable mutant with three mutated sites in β-tubulin showed reduced tubulin glycylation, slow growth and motility, and defects in cytokinesis. Cells in which all five polyglycylation sites on β-tubulin were mutated were viable if they were cotransformed with an α-tubulin gene whose COOH terminus was replaced by the wild-type COOH terminus of β-tubulin. In this double mutant, β-tubulin lacked detectable polyglycylation, while the α-β tubulin chimera was hyperglycylated compared with α-tubulin in wild-type cells. Thus, the essential function of polyglycylation of the COOH terminus of β-tubulin can be transferred to α-tubulin, indicating it is the total amount of polyglycylation on both α- and β-tubulin that is essential for survival. PMID:10831613

  12. Extracellular matrix rigidity governs smooth muscle cell motility in a biphasic fashion.

    PubMed

    Peyton, Shelly R; Putnam, Andrew J

    2005-07-01

    Increasing evidence suggests that mechanical cues inherent to the extracellular matrix (ECM) may be equally as critical as its chemical identity in regulating cell behavior. We hypothesized that the mechanical properties of the ECM directly regulate the motility of vascular smooth muscle cells (SMCs) and tested this hypothesis using polyacrylamide substrates with tunable mechanical properties. Quantification of the migration speed on uniformly compliant hydrogels spanning a range of stiffnesses (Young's moduli values from 1.0 to 308 kPa for acrylamide/bisacrylamide ratios between 5/0.1% and 15/1.2%, respectively) revealed a biphasic dependence on substrate compliance, suggesting the existence of an optimal substrate stiffness capable of supporting maximal migration. The value of this optimal stiffness shifted depending on the concentration of ECM protein covalently attached to the substrate. Specifically, on substrates presenting a theoretical density of 0.8 microg/cm(2) fibronectin, the maximum speed of 0.74 +/- 0.09 microm/min was achieved on a 51.9 kPa gel; on substrates presenting a theoretical density of 8.0 microg/cm(2) fibronectin, the maximum speed of 0.72 +/- 0.06 microm/min occurred on a softer 21.6 kPa gel. Pre-treatment of cells with Y27632, an inhibitor of the Rho/Rho-kinase (ROCK) pathway, reduced these observed maxima to values comparable to those on non-optimal stiffnesses. In parallel, quantification of TritonX-insoluble vinculin via Western blotting, coupled with qualitative fluorescent microscopy, revealed that the formation of focal adhesions and actin stress fibers also depends on ECM stiffness. Combined, these data suggest that the mechanical properties of the underlying ECM regulate Rho-mediated contractility in SMCs by disrupting a presumptive cell-ECM force balance, which in turn regulates cytoskeletal assembly and ultimately, cell migration.

  13. Daydreamer, a Ras effector and GSK-3 substrate, is important for directional sensing and cell motility

    PubMed Central

    Kölsch, Verena; Shen, Zhouxin; Lee, Susan; Plak, Katarzyna; Lotfi, Pouya; Chang, Jessica; Charest, Pascale G.; Romero, Jesus Lacal; Jeon, Taeck J.; Kortholt, Arjan; Briggs, Steven P.; Firtel, Richard A.

    2013-01-01

    How independent signaling pathways are integrated to holistically control a biological process is not well understood. We have identified Daydreamer (DydA), a new member of the Mig10/RIAM/lamellipodin (MRL) family of adaptor proteins that localizes to the leading edge of the cell. DydA is a putative Ras effector that is required for cell polarization and directional movement during chemotaxis. dydA− cells exhibit elevated F-actin and assembled myosin II (MyoII), increased and extended phosphoinositide-3-kinase (PI3K) activity, and extended phosphorylation of the activation loop of PKB and PKBR1, suggesting that DydA is involved in the negative regulation of these pathways. DydA is phosphorylated by glycogen synthase kinase-3 (GSK-3), which is required for some, but not all, of DydA's functions, including the proper regulation of PKB and PKBR1 and MyoII assembly. gskA− cells exhibit very strong chemotactic phenotypes, as previously described, but exhibit an increased rate of random motility. gskA− cells have a reduced MyoII response and a reduced level of phosphatidylinositol (3,4,5)-triphosphate production, but a highly extended recruitment of PI3K to the plasma membrane and highly extended kinetics of PKB and PKBR1 activation. Our results demonstrate that GSK-3 function is essential for chemotaxis, regulating multiple substrates, and that one of these effectors, DydA, plays a key function in the dynamic regulation of chemotaxis. PMID:23135995

  14. Cyclin-dependent kinase 5 regulates the proliferation, motility and invasiveness of lung cancer cells through its effects on cytoskeletal remodeling.

    PubMed

    Liu, Jun-Li; Gu, Run-Xia; Zhou, Xiao-Shu; Zhou, Fang-Zheng; Wu, Gang

    2015-09-01

    Determining the molecular phenotype is a key to understanding and predicting the metastatic potential and the prognosis for patients with lung cancer. Our previous study demonstrated that increased expression of cyclin‑dependent kinase 5 (CDK5) in patients with non‑small cell lung cancer (NSCLC) is associated with a poorer prognosis. The present study aimed to further investigate the underlying mechanism of CDK5 in vitro and in vivo using the A549 human NSCLC cell line. A 3‑(4,5‑dimethylthiazol‑2‑yl)‑2,5‑diphenyltetrazolium bromide assay was used to quantify the proliferation of the A549 cells; migration assay and invasiveness assays were performed using Transwell chambers and wound healing assays were used to assess cell motility, which was assessed by measuring the movement of cells. Inhibition of CDK5 by roscovitine and small interfering (si)RNA was used to investigate the mechanism of CDK5 in the process of A549 lung cancer cell proliferation, migration and invasion. The results demonstrated that functional inhibition of CDK5 using roscovitine and siRNA markedly suppressed the proliferation of A549 cells and resulted in a reduced tumor mass in vivo. In addition, the hinhibition of CDK5 reduced the migration and invasiveness of the A549 cells in vitro and in vivo. Notably, CDK5 inhibition also impaired tumor cell cytoskeletal remodeling and led to loss of cell polarity, which may partially explain the reduction of A549 cell mobility and invasiveness. The results of the present study revealed that CDK5 may be important in the regulation of migration and invasiveness in NSCLC through its effects on cytoskeletal remodeling. PMID:26018459

  15. Plant Lectin Can Target Receptors Containing Sialic Acid, Exemplified by Podoplanin, to Inhibit Transformed Cell Growth and Migration

    PubMed Central

    Shen, Yongquan; Acharya, Nimish K.; Han, Min; McNulty, Dean E.; Hasegawa, Hitoki; Hyodo, Toshinori; Senga, Takeshi; Geng, Jian-Guo; Kosciuk, Mary; Shin, Seung S.; Goydos, James S.; Temiakov, Dmitry; Nagele, Robert G.; Goldberg, Gary S.

    2012-01-01

    Cancer is a leading cause of death of men and women worldwide. Tumor cell motility contributes to metastatic invasion that causes the vast majority of cancer deaths. Extracellular receptors modified by α2,3-sialic acids that promote this motility can serve as ideal chemotherapeutic targets. For example, the extracellular domain of the mucin receptor podoplanin (PDPN) is highly O-glycosylated with α2,3-sialic acid linked to galactose. PDPN is activated by endogenous ligands to induce tumor cell motility and metastasis. Dietary lectins that target proteins containing α2,3-sialic acid inhibit tumor cell growth. However, anti-cancer lectins that have been examined thus far target receptors that have not been identified. We report here that a lectin from the seeds of Maackia amurensis (MASL) with affinity for O-linked carbohydrate chains containing sialic acid targets PDPN to inhibit transformed cell growth and motility at nanomolar concentrations. Interestingly, the biological activity of this lectin survives gastrointestinal proteolysis and enters the cardiovascular system to inhibit melanoma cell growth, migration, and tumorigenesis. These studies demonstrate how lectins may be used to help develop dietary agents that target specific receptors to combat malignant cell growth. PMID:22844530

  16. Nm23-h1 binds to gelsolin and inactivates its actin-severing capacity to promote tumor cell motility and metastasis.

    PubMed

    Marino, Natascia; Marshall, Jean-Claude; Collins, Joshua W; Zhou, Ming; Qian, Yongzhen; Veenstra, Timothy; Steeg, Patricia S

    2013-10-01

    Nm23-H1 has been identified as a metastasis suppressor gene, but its protein interactions have yet to be understood with any mechanistic clarity. In this study, we evaluated the proteomic spectrum of interactions made by Nm23-H1 in 4T1 murine breast cancer cells derived from tissue culture, primary mammary tumors, and pulmonary metastases. By this approach, we identified the actin-severing protein Gelsolin as binding partner for Nm23-H1, verifying their interaction by coimmunoprecipitation in 4T1 cells as well as in human MCF7, MDA-MB-231T, and MDA-MB-435 breast cancer cells. In Gelsolin-transfected cells, coexpression of Nm23-H1 abrogated the actin-severing activity of Gelsolin. Conversely, actin severing by Gelsolin was abrogated by RNA interference-mediated silencing of endogenous Nm23-H1. Tumor cell motility was negatively affected in parallel with Gelsolin activity, suggesting that Nm23-H1 binding inactivated the actin-depolymerizing function of Gelsolin to inhibit cell motility. Using indirect immunoflourescence to monitor complexes formed by Gelsolin and Nm23-H1 in living cells, we observed their colocalization in a perinuclear cytoplasmic compartment that was associated with the presence of disrupted actin stress fibers. In vivo analyses revealed that Gelsolin overexpression increased the metastasis of orthotopically implanted 4T1 or tail vein-injected MDA-MB-231T cells (P = 0.001 and 0.04, respectively), along with the proportion of mice with diffuse liver metastases, an effect ablated by coexpression of Nm23-H1. We observed no variation in proliferation among lung metastases. Our findings suggest a new actin-based mechanism that can suppress tumor metastasis.

  17. Trimebutine as a modulator of gastrointestinal motility.

    PubMed

    Lee, Hyun-Tai; Kim, Byung Joo

    2011-06-01

    Trimebutine has been used for treatment of both hypermotility and hypomotility disorders of the gastrointestinal (GI) tract, such as irritable bowel syndrome. In this issue, Tan et al. (2011) examined the concentration-dependent dual effects of trimebutine on colonic motility in guinea pig. The authors suggested that trimebutine attenuated colonic motility mainly through the inhibition of L-type Ca(2+) channels at higher concentrations, whereas, at lower concentrations, it depolarized membrane potentials by reducing BK(ca) currents, resulting in the enhancement of the muscle contractions. Trimebutine might be a plausible modulator of GI motility, which gives an insight in developing new prokinetic agents. Further studies to elucidate the effects of trimebutine on the interstitial cells of Cajal, the pacemaker in GI muscles would promote the therapeutic benefits as a GI modulator. PMID:21725804

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

    PubMed Central

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

    2016-01-01

    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. PMID:27670131

  19. 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.

  20. CD271 regulates the proliferation and motility of hypopharyngeal cancer cells

    PubMed Central

    Mochizuki, Mai; Tamai, Keiichi; Imai, Takayuki; Sugawara, Sayuri; Ogama, Naoko; Nakamura, Mao; Matsuura, Kazuto; Yamaguchi, Kazunori; Satoh, Kennichi; Sato, Ikuro; Motohashi, Hozumi; Sugamura, Kazuo; Tanaka, Nobuyuki

    2016-01-01

    CD271 (p75 neurotrophin receptor) plays both positive and negative roles in cancer development, depending on the cell type. We previously reported that CD271 is a marker for tumor initiation and is correlated with a poor prognosis in human hypopharyngeal cancer (HPC). To clarify the role of CD271 in HPC, we established HPC cell lines and knocked down the CD271 expression using siRNA. We found that CD271-knockdown completely suppressed the cells’ tumor-forming capability both in vivo and in vitro. CD271-knockdown also induced cell-cycle arrest in G0 and suppressed ERK phosphorylation. While treatment with an ERK inhibitor only partially inhibited cell growth, CDKN1C, which is required for maintenance of quiescence, was strongly upregulated in CD271-depleted HPC cells, and the double knockdown of CD271 and CDKN1C partially rescued the cells from G0 arrest. In addition, either CD271 depletion or the inhibition of CD271-RhoA signaling by TAT-Pep5 diminished the in vitro migration capability of the HPC cells. Collectively, CD271 initiates tumor formation by increasing the cell proliferation capacity through CDKN1C suppression and ERK-signaling activation, and by accelerating the migration signaling pathway in HPC. PMID:27469492

  1. Inhibition of β-N-acetylglucosaminidase by acetamide affects sperm motility and fertilization success of rainbow trout (Oncorhynchus mykiss) and Siberian sturgeon (Acipenser baerii).

    PubMed

    Sarosiek, B; Glogowski, J; Cejko, B I; Kujawa, R; Szczepkowski, M; Kuźmiński, H; Dobosz, S; Kowalski, R K

    2014-03-15

    β-N-Acetylglucosaminidase (β-NAGase) is an enzyme found in the sperm acrosome of numerous animal species including fish. Fish spermatozoa differ in their morphology including acrosome or acrosomeless aquasperm in chondrostean (e.g., sturgeon) and teleostean (e.g., rainbow trout). It has been shown that β-NAGase exists with high activity in both eggs and sperm of these species. The present study shows the potency of β-NAGase in fertilization. In rainbow trout, increase in sperm motility parameters (VAP and MOT) were observed in the presence of acetamide, an inhibitor for β-NAGase. In contrast, sperm motility parameters (VCL, VSL, VAP, MOT, and PRG) were reduced on the Siberian sturgeon in the presence of acetamide. The inhibition of the activity of β-NAGase in rainbow trout spermatozoa was led to a reduction in the number of fertilized eggs from 79% to 40%, whereas in sturgeon no change was observed in fertilization. Moreover, inhibition of β-NAGase in both spermatozoa and eggs of trout and sturgeon resulted in significant decrease in fertilization rate from 79% to 1% in rainbow trout and from 84% to 12% in Siberian sturgeon. Our research proves that β-NAGase can play a significant role in the fertilization process in teleosteans.

  2. THE DIFFERENTIAL REGULATION OF CELL MOTILE ACTIVITY THROUGH MATRIX STIFFNESS AND POROSITY IN THREE DIMENSIONAL COLLAGEN MATRICES

    PubMed Central

    Miron-Mendoza, Miguel; Seemann, Joachim; Grinnell, Frederick

    2010-01-01

    In three dimensional collagen matrices, cell motile activity results in collagen translocation, cell spreading and cell migration. Cells can penetrate into the matrix as well as spread and migrate along its surface. In the current studies, we quantitatively characterize collagen translocation, cell spreading and cell migration in relationship to collagen matrix stiffness and porosity. Collagen matrices prepared with 1 to 4 mg/ml collagen exhibited matrix stiffness (storage modulus measured by oscillating rheometry) increasing from 4 to 60 Pa and matrix porosity (measured by scanning electron microscopy) decreasing from 4 to 1 μm2. Over this collagen concentration range, the consequences of cell motile activity changed markedly. As collagen concentration increased, cells no longer were able to cause translocation of collagen fibrils. Cell migration increased and cell spreading changed from dendritic to more flattened and polarized morphology depending on location of cells within or on the surface of the matrix. Collagen translocation appeared to depend primarily on matrix stiffness, whereas cell spreading and migration were less dependent on matrix stiffness and more dependent on collagen matrix porosity. PMID:20537378

  3. Microscopic analysis of bacterial motility at high pressure.

    PubMed

    Nishiyama, Masayoshi; Sowa, Yoshiyuki

    2012-04-18

    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.

  4. Pancreatic Fibroblasts Stimulate the Motility of Pancreatic Cancer Cells through IGF1/IGF1R Signaling under Hypoxia

    PubMed Central

    Hirakawa, Toshiki; Yashiro, Masakazu; Doi, Yosuke; Kinoshita, Haruhito; Morisaki, Tamami; Fukuoka, Tatsunari; Hasegawa, Tsuyoshi; Kimura, Kenjiro; Amano, Ryosuke; Hirakawa, Kosei

    2016-01-01

    Pancreatic ductal adenocarcinoma (PDAC) is characterized by its hypovascularity, with an extremely poor prognosis because of its highly invasive nature. PDAC proliferates with abundant stromal cells, suggesting that its invasive activity might be controlled by intercellular interactions between cancer cells and fibroblasts. Using four PDAC cell lines and two pancreas cancer-associated fibroblasts (CAFs), the expression of insulin-like growth factor-1 (IGF1) and IGF1 receptor (IGF1R) was evaluated by RT-PCR, FACScan, western blot, or ELISA. Correlation between IGF1R and the hypoxia marker carbonic anhydrase 9 (CA9) was examined by immunohistochemical staining of 120 pancreatic specimens. The effects of CAFs, IGF1, and IGF1R inhibitors on the motility of cancer cells were examined by wound-healing assay or invasion assay under normoxia (20% O2) and hypoxia (1% O2). IGF1R expression was significantly higher in RWP-1, MiaPaCa-2, and OCUP-AT cells than in Panc-1 cells. Hypoxia increased the expression level of IGF1R in RWP-1, MiaPaCa-2, and OCUP-AT cells. CA9 expression was correlated with IGF1R expression in pancreatic specimens. CAFs produced IGF1 under hypoxia, but PDAC cells did not. A conditioned medium from CAFs, which expressed αSMA, stimulated the migration and invasion ability of MiaPaCa-2, RWP-1, and OCUP-AT cells. The motility of all PDAC cells was greater under hypoxia than under normoxia. The motility-stimulating ability of CAFs was decreased by IGF1R inhibitors. These findings might suggest that pancreas CAFs stimulate the invasion activity of PDAC cells through paracrine IGF1/IGF1R signaling, especially under hypoxia. Therefore the targeting of IGF1R signaling might represent a promising therapeutic approach in IGF1R-dependent PDAC. PMID:27487118

  5. 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

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

    2013-07-25

    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.

  6. Gonadotropin-releasing hormone type II (GnRH-II) agonist regulates the motility of human decidual endometrial stromal cells: possible effect on embryo implantation and pregnancy.

    PubMed

    Wu, Hsien-Ming; Huang, Hong-Yuan; Lee, Chyi-Long; Soong, Yung-Kuei; Leung, Peter C K; Wang, Hsin-Shih

    2015-04-01

    Invasion of the maternal decidua by extravillous trophoblast is an important process for embryo implantation and placentation in humans. Motile behavior of decidual endometrial stromal cells has been considered of critical importance for embryo implantation and programming of human pregnancy. The gonadotropin-releasing hormone (GnRH) effects in endometrium have raised concerns in reproduction. In the present study, we examined the action of GnRH-II agonist-promoted motility of human decidual endometrial stromal cells and the mechanisms of the action, indicating the role of GnRH-II agonist in embryo implantation and early pregnancy. Human decidual endometrial stromal cells were isolated from the decidual tissue from healthy women undergoing elective pregnancy termination of a normal pregnancy at 6- to 12-wk gestation, after informed consent. Cell motility was estimated by invasion and migration assay. Zymography and immunoblot analysis were performed to investigate the mechanisms of the GnRH-II action. The GnRH-I receptor (GnRH-IR) was expressed in human decidual tissue and endometrial stromal cells. The GnRH-II agonist promoted cell motility. Mitogen-activated protein kinase inhibitors abolished GnRH-II agonist-induced cell motility and activation of MMP-2 and MMP-9. GnRH-II agonist-mediated cell motility was suppressed by knockdown of endogenous GnRH-IR, MMP (matrix metalloproteinase)-2, and MMP-9 with small interfering RNA and MMP inhibitors. Our study demonstrates that the GnRH-II agonist promoted the cell motility of human decidual endometrial stromal cells through the GnRH-IR and the phosphorylation of extracellular signal-regulated protein kinase 1/2 and JNK-dependent activation of MMP-2 and MMP-9. Our findings represent a new concept regarding the mechanisms of GnRH-II-promoted cell motility, suggesting that GnRH-II agonist has strong effects on embryo implantation and decidual programming of human pregnancy. PMID:25761596

  7. CyrA, a matricellular protein that modulates cell motility in Dictyostelium discoideum.

    PubMed

    Huber, Robert J; Suarez, Andres; O'Day, Danton H

    2012-05-01

    CyrA, an extracellular matrix (slime sheath), calmodulin (CaM)-binding protein in Dictyostelium discoideum, possesses four tandem EGF-like repeats in its C-terminus and is proteolytically cleaved during asexual development. A previous study reported the expression and localization of CyrA cleavage products CyrA-C45 and CyrA-C40. In this study, an N-terminal antibody was produced that detected the full-length 63kDa protein (CyrA-C63). Western blot analyses showed that the intracellular expression of CyrA-C63 peaked between 12 and 16h of development, consistent with the time that cells are developing into a motile, multicellular slug. CyrA immunolocalization and CyrA-GFP showed that the protein localized to the endoplasmic reticulum, particularly its perinuclear component. CyrA-C63 secretion began shortly after the onset of starvation peaking between 8 and 16h of development. A pharmacological analysis showed that CyrA-C63 secretion was dependent on intracellular Ca(2+) release and active CaM, PI3K, and PLA2. CyrA-C63 bound to CaM both intra- and extracellularly and both proteins were detected in the slime sheath deposited by migrating slugs. In keeping with its purported function, CyrA-GFP over-expression enhanced cAMP-mediated chemotaxis and CyrA-C45 was detected in vinculin B (VinB)-GFP immunoprecipitates, thus providing a link between the increase in chemotaxis and a specific cytoskeletal component. Finally, DdEGFL1-FITC was detected on the membranes of cells capped with concanavalin A suggesting that a receptor exists for this peptide sequence. Together with previous studies, the data presented here suggests that CyrA is a bona fide matricellular protein in D. discoideum.

  8. Senescent stromal cells induce cancer cell migration via inhibition of RhoA/ROCK/myosin-based cell contractility.

    PubMed

    Aifuwa, Ivie; Giri, Anjil; Longe, Nick; Lee, Sang Hyuk; An, Steven S; Wirtz, Denis

    2015-10-13

    Cells induced into senescence exhibit a marked increase in the secretion of pro-inflammatory cytokines termed senescence-associated secretory phenotype (SASP). Here we report that SASP from senescent stromal fibroblasts promote spontaneous morphological changes accompanied by an aggressive migratory behavior in originally non-motile human breast cancer cells. This phenotypic switch is coordinated, in space and time, by a dramatic reorganization of the actin and microtubule filament networks, a discrete polarization of EB1 comets, and an unconventional front-to-back inversion of nucleus-MTOC polarity. SASP-induced morphological/migratory changes are critically dependent on microtubule integrity and dynamics, and are coordinated by the inhibition of RhoA and cell contractility. RhoA/ROCK inhibition reduces focal adhesions and traction forces, while promoting a novel gliding mode of migration. PMID:26483365

  9. Secreted or nonsecreted forms of acidic fibroblast growth factor produced by transfected epithelial cells influence cell morphology, motility, and invasive potential.

    PubMed Central

    Jouanneau, J; Gavrilovic, J; Caruelle, D; Jaye, M; Moens, G; Caruelle, J P; Thiery, J P

    1991-01-01

    Addition of exogenous acidic fibroblast growth factor (aFGF) to NBT-II epithelial carcinoma cells results in fibroblastic transformation and cell motility. We have generated aFGF-producing NBT-II cells by transfection with recombinant expression vectors containing human aFGF cDNA, or the human aFGF cDNA coupled to a signal peptide (SP) sequence. The effects of the nonsecreted and the secreted 16-kDa growth factor on the morphology, motility, and cell invasive potential (gelatinase activity) were compared. aFGF coupled to a SP was actively secreted out of the producing cells. The secretion of aFGF was not necessary for induction of gelatinase activity, as this was observed in NBT-II cells producing aFGF with or without SP. Production of aFGF, whether secreted or not secreted, resulted in increased in vitro motility of most isolated clones; however, there was no correlation between aFGF level and motility rate. The data suggest that expression of aFGF in NBT-II cells induces metastatic potential through an autocrine or intracrine mechanism. Images PMID:1707175

  10. Pediatric intestinal motility disorders

    PubMed Central

    Gfroerer, Stefan; Rolle, Udo

    2015-01-01

    Pediatric intestinal motility disorders affect many children and thus not only impose a significant impact on pediatric health care in general but also on the quality of life of the affected patient. Furthermore, some of these conditions might also have implications for adulthood. Pediatric intestinal motility disorders frequently present as chronic constipation in toddler age children. Most of these conditions are functional, meaning that constipation does not have an organic etiology, but in 5% of the cases, an underlying, clearly organic disorder can be identified. Patients with organic causes for intestinal motility disorders usually present in early infancy or even right after birth. The most striking clinical feature of children with severe intestinal motility disorders is the delayed passage of meconium in the newborn period. This sign is highly indicative of the presence of Hirschsprung disease (HD), which is the most frequent congenital disorder of intestinal motility. HD is a rare but important congenital disease and the most significant entity of pediatric intestinal motility disorders. The etiology and pathogenesis of HD have been extensively studied over the last several decades. A defect in neural crest derived cell migration has been proven as an underlying cause of HD, leading to an aganglionic distal end of the gut. Numerous basic science and clinical research related studies have been conducted to better diagnose and treat HD. Resection of the aganglionic bowel remains the gold standard for treatment of HD. Most recent studies show, at least experimentally, the possibility of a stem cell based therapy for HD. This editorial also includes rare causes of pediatric intestinal motility disorders such as hypoganglionosis, dysganglionosis, chronic intestinal pseudo-obstruction and ganglioneuromatosis in multiple endocrine metaplasia. Underlying organic pathologies are rare in pediatric intestinal motility disorders but must be recognized as early as

  11. Rab11-FIP3 is a Rab11-binding protein that regulates breast cancer cell motility by modulating the actin cytoskeleton

    PubMed Central

    Jing, Jian; Tarbutton, Elizabeth; Wilson, Gayle; Prekeris, Rytis

    2009-01-01

    Cell adhesion and motility are very dynamic processes that require the temporal and spatial coordination of many cellular structures. ADP-ribosylation factor 6 (Arf6) has emerged as master regulator of endocytic membrane traffic and cytoskeletal dynamics during cell movement. Recently, a novel Arf6-binding protein known as FIP3/arfophilin/eferin has been identified. In addition to Arf6, FIP3 also interacts with Rab11, a small monomeric GTPase that regulates endocytic membrane transport. Both Arf6 and Rab11 GTPases have been implicated in regulation of cell motility. Here we test the role of FIP3 in breast carcinoma cell motility. First, we demonstrate that FIP3 is associated with recycling endosomes that are present at the leading edge of motile cells. Second, we show that FIP3 is required for the motility of MDA-MB-231 breast carcinoma cells. Third, we demonstrate that FIP3 regulates Rac1-dependent actin cytoskeleton dynamics and modulates the formation and ruffling of lamellipodia. Finally, we demonstrate that FIP3 regulates the localization of Arf6 at the plasma membrane of MDA-MB-231 cells. Based on our data we propose that FIP3 affects cell motility by regulating Arf6 localization to the plasma membrane of the leading edge, thus regulating polarized Rac1 activation and actin dynamics. PMID:19327867

  12. Marrow-Derived Stem Cell Motility in 3D Synthetic Scaffold Is Governed by Geometry Along With Adhesivity and Stiffness

    PubMed Central

    Peyton, Shelly R.; Kalcioglu, Z. Ilke; Cohen, Joshua C.; Runkle, Anne P.; Van Vliet, Krystyn J.; Lauffenburger, Douglas A.

    2012-01-01

    Design of 3D scaffolds that can facilitate proper survival, proliferation, and differentiation of progenitor cells is a challenge for clinical applications involving large connective tissue defects. Cell migration within such scaffolds is a critical process governing tissue integration. Here, we examine effects of scaffold pore diameter, in concert with matrix stiffness and adhesivity, as independently tunable parameters that govern marrow-derived stem cell motility. We adopted an “inverse opal” processing technique to create synthetic scaffolds by crosslinking poly(ethylene glycol) at different densities (controlling matrix elastic moduli or stiffness) and small doses of a heterobifunctional monomer (controlling matrix adhesivity) around templating beads of different radii. As pore diameter was varied from 7 to 17 µm (i.e., from significantly smaller than the spherical cell diameter to approximately cell diameter), it displayed a profound effect on migration of these stem cells—including the degree to which motility was sensitive to changes in matrix stiffness and adhesivity. Surprisingly, the highest probability for substantive cell movement through pores was observed for an intermediate pore diameter, rather than the largest pore diameter, which exceeded cell diameter. The relationships between migration speed, displacement, and total path length were found to depend strongly on pore diameter. We attribute this dependence to convolution of pore diameter and void chamber diameter, yielding different geometric environments experienced by the cells within. PMID:21449030

  13. Depletion of enteroendocrine and mucus-secreting cells is associated with colorectal carcinogenesis severity and impaired intestinal motility in rats.

    PubMed

    Novaes, Rômulo Dias; Sequetto, Priscila Lima; Vilela Gonçalves, Reggiani; Cupertino, Marli Carmo; Santos, Eliziária Cardoso; Mello, Vanessa Joia; Araújo, Marta Rocha; Silva, Edson; Oliveira, Tânia Toledo

    2016-01-01

    This study investigated the relationship between enteroendocrine and mucus-secreting cells distribution, the severity of colonic mucosal injury and intestinal motility in experimental colorectal carcinogenesis. Using a standardized murine model of colorectal carcinogenesis, eight-weeks-old female Wistar rats weighting 147.30 ± 29.15g were randomized into two groups receiving a subcutaneous injection of 0.9% saline (control) or the chemical carcinogen 1,2-dimethylhydrazine (DMH) at 20 mg/kg per week during 10 weeks. Aberrant crypt foci (ACF) were more frequent in DMH group compared to control group (P < 0.001). The number of enteroendocrine and mucus-secreting cells, and intestinal motility was reduced in DMH animals (P < 0.05). The distribution of enteric neurons was similar in both groups. In DMH animals there was a direct correlation between colonic motility and distribution of enteroendocrine (R(2)  = 0.68, P < 0.05) and mucus-secreting cells (R(2)  = 0.77, P < 0.05). Inverse correlation between the number of ACF, mucus-secreting cells (R(2)  = -0.57, P < 0.05), and enteroendocrine cells (R(2)  = -0.74, P < 0.05) was also observed. There was inverse correlation between the severity of the mucosal lesion, the number of mucus-secreting cells (R(2)  = -0.83, P < 0.05) and enteroendocrine cells (R(2)  = -0.96, P < 0.05). There was a direct correlation between nucleolar organizer regions (AgNOR) and ACF number (R(2)  = 0.62; P < 0.01). Inverse correlation was also found between AgNOR, the number of mucus-secreting cells (R(2)  = -0.76; P < 0.001), and enteroendocrine cells (R(2)  = -0.86; P < 0.001). Taken together, the results indicated that colonic malignant transformation is related to depletion of mucus-secreting and enteroendocrine cells, which was a useful indicator of the evolutionary status of intestinal neoplasm, partially explaining the intestinal motility disorders in the

  14. MicroRNA networks regulated by all-trans retinoic acid and Lapatinib control the growth, survival and motility of breast cancer cells

    PubMed Central

    Kurosaki, Mami; Paroni, Gabriela; Zanetti, Adriana; Gianni, Maurizio; Bolis, Marco; Lupi, Monica; Tsykin, Anna; Goodall, Gregory J.; Garattini, Enrico

    2015-01-01

    SKBR3-cells, characterized by ERBB2/RARA co-amplification, represent a subgroup of HER2+ breast-cancers sensitive to all-trans retinoic acid (ATRA) and Lapatinib. In this model, the two agents alone or in combination modulate the expression of 174 microRNAs (miRs). These miRs and predicted target-transcripts are organized in four interconnected modules (Module-1 to -4). Module-1 and Module-3 consist of ATRA/Lapatinib up-regulated and potentially anti-oncogenic miRs, while Module-2 contains ATRA/Lapatinib down-regulated and potentially pro-oncogenic miRs. Consistent with this, the expression levels of Module-1/-3 and Module-2 miRs are higher and lower, respectively, in normal mammary tissues relative to ductal-carcinoma-in-situ, invasive-ductal-carcinoma and metastases. This indicates associations between tumor-progression and the expression profiles of Module-1 to -3 miRs. Similar associations are observed with tumor proliferation-scores, staging, size and overall-survival using TCGA (The Cancer Genome Atlas) data. Forced expression of Module-1 miRs, (miR-29a-3p; miR-874-3p) inhibit SKBR3-cell growth and Module-3 miRs (miR-575; miR-1225-5p) reduce growth and motility. Module-2 miRs (miR-125a; miR-193; miR-210) increase SKBR3 cell growth, survival and motility. Some of these effects are of general significance, being replicated in other breast cancer cell lines representing the heterogeneity of this disease. Finally, our study demonstrates that HIPK2-kinase and the PLCXD1-phospholipase-C are novel targets of miR-193a-5p/miR-210-3p and miR-575/miR-1225-5p, respectively. PMID:25961594

  15. Axonemal motility in Chlamydomonas.

    PubMed

    Wakabayashi, Ken-ichi; Kamiya, Ritsu

    2015-01-01

    Motile cilia and flagella rapidly propagate bending waves and produce water flow over the cell surface. Their function is important for the physiology and development of various organisms including humans. The movement is based on the sliding between outer doublet microtubules driven by axonemal dyneins, and is regulated by various axonemal components and environmental factors. For studies aiming to elucidate the mechanism of cilia/flagella movement and regulation, Chlamydomonas is an invaluable model organism that offers a variety of mutants. This chapter introduces standard methods for studying Chlamydomonas flagellar motility including analysis of swimming paths, measurements of swimming speed and beat frequency, motility reactivation in demembranated cells (cell models), and observation of microtubule sliding in disintegrating axonemes. Most methods may be easily applied to other organisms with slight modifications of the medium conditions.

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

    SciTech Connect

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

    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 multicellular 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.

  17. Additive effect by combination of Akt inhibitor, MK-2206, and PDGFR inhibitor, tyrphostin AG 1296, in suppressing anaplastic thyroid carcinoma cell viability and motility

    PubMed Central

    Che, Huan-yong; Guo, Hang-yuan; Si, Xu-wei; You, Qiao-ying; Lou, Wei-ying

    2014-01-01

    The phosphatidylinositol-3-kinase/Akt pathway and receptor tyrosine kinases regulate many tumorigenesis related cellular processes including cell metabolism, cell survival, cell motility, and angiogenesis. Anaplastic thyroid carcinoma (ATC) is a rare type of thyroid cancer with no effective systemic therapy. It has been shown that Akt activation is associated with tumor progression in ATC. Here we observed the additive effect between an Akt inhibitor (MK-2206) and a novel platelet-derived growth factor receptor inhibitor (tyrphostin AG 1296) in ATC therapy. We found an additive effect between MK-2206 and tyrphostin AG 1296 in suppressing ATC cell viability. The combination of MK-2206 and tyrphostin AG 1296 induces additive apoptosis, additive suppression of the Akt signaling pathway, as well as additive inhibition of cell migration and invasion of ATC cells. Furthermore, the combination of MK-2206 and tyrphostin AG 1296 induced additive suppression of ATC tumor growth in vivo. In summary, our studies suggest that the combination of Akt and receptor tyrosine kinase inhibitors may be an efficient therapeutic strategy for ATC treatment, which might shed new light on ATC therapy. PMID:24665203

  18. Ceramide limits phosphatidylinositol-3-kinase C2β-controlled cell motility in ovarian cancer: potential of ceramide as a metastasis-suppressor lipid.

    PubMed

    Kitatani, K; Usui, T; Sriraman, S K; Toyoshima, M; Ishibashi, M; Shigeta, S; Nagase, S; Sakamoto, M; Ogiso, H; Okazaki, T; Hannun, Y A; Torchilin, V P; Yaegashi, N

    2016-05-01

    Targeting cell motility, which is required for dissemination and metastasis, has therapeutic potential for ovarian cancer metastasis, and regulatory mechanisms of cell motility need to be uncovered for developing novel therapeutics. Invasive ovarian cancer cells spontaneously formed protrusions, such as lamellipodia, which are required for generating locomotive force in cell motility. Short interfering RNA screening identified class II phosphatidylinositol 3-kinase C2β (PI3KC2β) as the predominant isoform of PI3K involved in lamellipodia formation of ovarian cancer cells. The bioactive sphingolipid ceramide has emerged as an antitumorigenic lipid, and treatment with short-chain C6-ceramide decreased the number of ovarian cancer cells with PI3KC2β-driven lamellipodia. Pharmacological analysis demonstrated that long-chain ceramide regenerated from C6-ceramide through the salvage/recycling pathway, at least in part, mediated the action of C6-ceramide. Mechanistically, ceramide was revealed to interact with the PIK-catalytic domain of PI3KC2β and affect its compartmentalization, thereby suppressing PI3KC2β activation and its driven cell motility. Ceramide treatment also suppressed cell motility promoted by epithelial growth factor, which is a prometastatic factor. To examine the role of ceramide in ovarian cancer metastasis, ceramide liposomes were employed and confirmed to suppress cell motility in vitro. Ceramide liposomes had an inhibitory effect on peritoneal metastasis in a murine xenograft model of human ovarian cancer. Metastasis of PI3KC2β knocked-down cells was insensitive to treatment with ceramide liposomes, suggesting specific involvement of ceramide interaction with PI3KC2β in metastasis suppression. Our study identified ceramide as a bioactive lipid that limits PI3KC2β-governed cell motility, and ceramide is proposed to serve as a metastasis-suppressor lipid in ovarian cancer. These findings could be translated into developing ceramide

  19. Ceramide limits phosphatidylinositol-3-kinase C2β-controlled cell motility in ovarian cancer: potential of ceramide as a metastasis-suppressor lipid.

    PubMed

    Kitatani, K; Usui, T; Sriraman, S K; Toyoshima, M; Ishibashi, M; Shigeta, S; Nagase, S; Sakamoto, M; Ogiso, H; Okazaki, T; Hannun, Y A; Torchilin, V P; Yaegashi, N

    2016-05-01

    Targeting cell motility, which is required for dissemination and metastasis, has therapeutic potential for ovarian cancer metastasis, and regulatory mechanisms of cell motility need to be uncovered for developing novel therapeutics. Invasive ovarian cancer cells spontaneously formed protrusions, such as lamellipodia, which are required for generating locomotive force in cell motility. Short interfering RNA screening identified class II phosphatidylinositol 3-kinase C2β (PI3KC2β) as the predominant isoform of PI3K involved in lamellipodia formation of ovarian cancer cells. The bioactive sphingolipid ceramide has emerged as an antitumorigenic lipid, and treatment with short-chain C6-ceramide decreased the number of ovarian cancer cells with PI3KC2β-driven lamellipodia. Pharmacological analysis demonstrated that long-chain ceramide regenerated from C6-ceramide through the salvage/recycling pathway, at least in part, mediated the action of C6-ceramide. Mechanistically, ceramide was revealed to interact with the PIK-catalytic domain of PI3KC2β and affect its compartmentalization, thereby suppressing PI3KC2β activation and its driven cell motility. Ceramide treatment also suppressed cell motility promoted by epithelial growth factor, which is a prometastatic factor. To examine the role of ceramide in ovarian cancer metastasis, ceramide liposomes were employed and confirmed to suppress cell motility in vitro. Ceramide liposomes had an inhibitory effect on peritoneal metastasis in a murine xenograft model of human ovarian cancer. Metastasis of PI3KC2β knocked-down cells was insensitive to treatment with ceramide liposomes, suggesting specific involvement of ceramide interaction with PI3KC2β in metastasis suppression. Our study identified ceramide as a bioactive lipid that limits PI3KC2β-governed cell motility, and ceramide is proposed to serve as a metastasis-suppressor lipid in ovarian cancer. These findings could be translated into developing ceramide

  20. Ceramide limits phosphatidylinositol-3-kinase C2β-controlled cell motility in ovarian cancer: potential of ceramide as a metastasis-suppressor lipid

    PubMed Central

    Kitatani, Kazuyuki; Toshinori, Usui; Sriraman, Shravan Kumar; Toyoshima, Masafumi; Ishibashi, Masumi; Shigeta, Shogo; Nagase, Satoru; Sakamoto, Masahiro; Ogiso, Hideo; Okazaki, Toshiro; Hannun, Yusuf A.; Torchilin, Vladimir P.; Yaegashi, Nobuo

    2015-01-01

    Targeting cell motility, which is required for dissemination and metastasis, has therapeutic potential for ovarian cancer metastasis, and regulatory mechanisms of cell motility need to be uncovered for developing novel therapeutics. Invasive ovarian cancer cells spontaneously formed protrusions, such as lamellipodia, which are required for generating locomotive force in cell motility. siRNA screening identified class II phosphatidylinositol 3-kinase C2β (PI3KC2β) as the predominant isoform of PI3K involved in lamellipodia formation of ovarian cancer cells. The bioactive sphingolipid ceramide has emerged as an antitumorigenic lipid, and treatment with short-chain C6-ceramide decreased the number of ovarian cancer cells with PI3KC2β-driven lamellipodia. Pharmacological analysis demonstrated that long-chain ceramide regenerated from C6-ceramide through the salvage/recycling pathway, at least in part, mediated the action of C6-ceramide. Mechanistically, ceramide was revealed to interact with the PIK-catalytic domain of PI3KC2β and affect its compartmentalization, thereby suppressing PI3KC2β activation and its driven cell motility. Ceramide treatment also suppressed cell motility promoted by epithelial growth factor, which is a prometastatic factor. To examine the role of ceramide in ovarian cancer metastasis, ceramide liposomes were employed and confirmed to suppress cell motility in vitro. Ceramide liposomes had an inhibitory effect on peritoneal metastasis in a murine xenograft model of human ovarian cancer. Metastasis of PI3KC2β knocked-down cells was insensitive to treatment with ceramide liposomes, suggesting specific involvement of ceramide interaction with PI3KC2β in metastasis suppression. Our study identified ceramide as a bioactive lipid that limits PI3KC2β-governed cell motility, and ceramide is proposed to serve as a metastasis-suppressor lipid in ovarian cancer. These findings could be translated into developing ceramide-based therapy for

  1. Green tea flavan-3-ols and oligomeric proanthocyanidins inhibit the motility of infective larvae of Teladorsagia circumcincta and Trichostrongylus colubriformis in vitro.

    PubMed

    Molan, A L; Sivakumaran, S; Spencer, P A; Meagher, L P

    2004-12-01

    The effects of a hot water infusion and an aqueous acetone extract of green tea (Camellia sinensis) on the motility of infective larvae of the sheep nematodes Teladorsagia circumcincta and Trichostrongylus colubriformis were investigated under in vitro conditions. The infusion and extract dose-dependently inactivated the infective larvae as assessed by the larval migration inhibition (LMI) assay. To determine the components responsible for the inhibitory activity, the hot water infusion and aqueous acetone extract of green tea were fractionated on Sephadex LH-20 and the green tea extract fractions (GTE-I-VIII) characterised by mass spectrometry. The larvae were exposed to increasing concentrations of these GTE fractions. Fractions containing epigallocatechin gallate (EGCG) and proanthocyanidin oligomers were most effective. GTE fractions were more effective against T. circumcincta than T. colubriformis larvae as assessed by the LMI assay. PMID:15276775

  2. Flagella and pili-mediated near-surface single-cell motility mechanisms in P. aeruginosa.

    PubMed

    Conrad, Jacinta C; Gibiansky, Maxsim L; Jin, Fan; Gordon, Vernita D; Motto, Dominick A; Mathewson, Margie A; Stopka, Wiktor G; Zelasko, Daria C; Shrout, Joshua D; Wong, Gerard C L

    2011-04-01

    Bacterial biofilms are structured multicellular communities that are responsible for a broad range of infections. Knowing how free-swimming bacteria adapt their motility mechanisms near a surface is crucial for understanding the transition from the planktonic to the biofilm phenotype. By translating microscopy movies into searchable databases of bacterial behavior and developing image-based search engines, we were able to identify fundamental appendage-specific mechanisms for the surface motility of Pseudomonas aeruginosa. Type IV pili mediate two surface motility mechanisms: horizontally oriented crawling, by which the bacterium moves lengthwise with high directional persistence, and vertically oriented walking, by which the bacterium moves with low directional persistence and high instantaneous velocity, allowing it to rapidly explore microenvironments. The flagellum mediates two additional motility mechanisms: near-surface swimming and surface-anchored spinning, which often precedes detachment from a surface. Flagella and pili interact cooperatively in a launch sequence whereby bacteria change orientation from horizontal to vertical and then detach. Vertical orientation facilitates detachment from surfaces and thereby influences biofilm morphology. PMID:21463573

  3. Neutrophils lacking platelet-endothelial cell adhesion molecule-1 exhibit loss of directionality and motility in CXCR2-mediated chemotaxis.

    PubMed

    Wu, Yue; Stabach, Paul; Michaud, Michael; Madri, Joseph A

    2005-09-15

    Time-lapsed videomicroscopy was used to study the migration of platelet-endothelial cell adhesion molecule-1-deficient (PECAM-1(-/-)) murine neutrophils undergoing chemotaxis in Zigmond chambers containing IL-8, KC, or fMLP gradients. PECAM-1(-/-) neutrophils failed to translocate up the IL-8, KC, and fMLP gradients. Significant reductions in cell motility and cell spreading were also observed in IL-8 or KC gradients. In wild-type neutrophils, PECAM-1 and F-actin were colocalized at the leading fronts of polarized cells toward the gradient. In contrast, in PECAM-1(-/-) neutrophils, although F-actin also localized to the leading front of migrating cells, F-actin polymerization was unstable, and cycling was remarkably increased compared with that of wild-type neutrophils. This may be due to the decreased cytokine-induced mobilization of the actin-binding protein, moesin, into the cytoskeleton of PECAM-1(-/-) neutrophils. PECAM-1(-/-) neutrophils also exhibited intracellularly dislocalized Src homology 2 domain containing phosphatase 1 (SHP-1) and had less IL-8-induced SHP-1 phosphatase activity. These results suggest that PECAM-1 regulates neutrophil chemotaxis by modulating cell motility and directionality, in part through its effects on SHP-1 localization and activation. PMID:16148090

  4. One isoform of Arg/Abl2 tyrosine kinase is nuclear and the other seven cytosolic isoforms differently modulate cell morphology, motility and the cytoskeleton

    SciTech Connect

    Bianchi, Cristina; Torsello, Barbara; Di Stefano, Vitalba; Zipeto, Maria A.; Facchetti, Rita; Bombelli, Silvia; Perego, Roberto A.

    2013-08-01

    The non-receptor tyrosine kinase Abelson related gene (Arg/Abl2) regulates cell migration and morphogenesis by modulating the cytoskeleton. Arg promotes actin-based cell protrusions and spreading, and inhibits cell migration by attenuating stress fiber formation and contractility via activation of the RhoA inhibitor, p190RhoGAP, and by regulating focal adhesion dynamics also via CrkII phosphorylation. Eight full-length Arg isoforms with different N- and C-termini are endogenously expressed in human cells. In this paper, the eight Arg isoforms, subcloned in the pFLAG-CMV2 vector, were transfected in COS-7 cells in order to study their subcellular distribution and role in cell morphology, migration and cytoskeletal modulation. The transfected 1BSCTS Arg isoform has a nuclear distribution and phosphorylates CrkII in the nucleus, whilst the other isoforms are detected in the cytoplasm. The 1BLCTL, 1BSCTL, 1ASCTS isoforms were able to significantly decrease stress fibers, induce cell shrinkage and filopodia-like protrusions with a significant increase in p190RhoGAP phosphorylation. In contrast, 1ALCTL, 1ALCTS, 1ASCTL and 1BLCTS isoforms do not significantly decrease stress fibers and induce the formation of retraction tail-like protrusions. The 1BLCTL and 1ALCTL isoforms have different effects on cell migration and focal adhesions. All these data may open new perspectives to study the mechanisms of cell invasiveness. -Highlights: • Each of the eight Arg isoforms was transfected in COS-7 cells. • Only the 1BSCTS Arg isoform has a nuclear distribution in transfected cells. • The cytoplasmic isoforms and F-actin colocalize cortically and in cell protrusions. • Arg isoforms differently phosphorylate p190RhoGAP and CrkII. • Arg isoforms differently modulate stress fibers, cell protrusions and motility.

  5. The human PMR1 endonuclease stimulates cell motility by down regulating miR-200 family microRNAs.

    PubMed

    Gu, Shan-Qing; Gallego-Perez, Daniel; McClory, Sean P; Shi, Junfeng; Han, Joonhee; Lee, L James; Schoenberg, Daniel R

    2016-07-01

    The motility of MCF-7 cells increases following expression of a human PMR1 transgene and the current study sought to identify the molecular basis for this phenotypic change. Ensemble and single cell analyses show increased motility is dependent on the endonuclease activity of hPMR1, and cells expressing active but not inactive hPMR1 invade extracellular matrix. Nanostring profiling identified 14 microRNAs that are downregulated by hPMR1, including all five members of the miR-200 family and others that also regulate invasive growth. miR-200 levels increase following hPMR1 knockdown, and changes in miR-200 family microRNAs were matched by corresponding changes in miR-200 targets and reporter expression. PMR1 preferentially cleaves between UG dinucleotides within a consensus YUGR element when present in the unpaired loop of a stem-loop structure. This motif is present in the apical loop of precursors to most of the downregulated microRNAs, and hPMR1 targeting of pre-miRs was confirmed by their loss following induced expression and increase following hPMR1 knockdown. Introduction of miR-200c into hPMR1-expressing cells reduced motility and miR-200 target gene expression, confirming hPMR1 acts upstream of Dicer processing. These findings identify a new role for hPMR1 in the post-transcriptional regulation of microRNAs in breast cancer cells. PMID:27257068

  6. RanBPM Protein Acts as a Negative Regulator of BLT2 Receptor to Attenuate BLT2-mediated Cell Motility*

    PubMed Central

    Wei, Jun-Dong; Kim, Joo-Young; Kim, Ae-Kyoung; Jang, Sung Key; Kim, Jae-Hong

    2013-01-01

    BLT2, a low affinity receptor for leukotriene B4 (LTB4), is a member of the G protein-coupled receptor family and is involved in many signal transduction pathways associated with various cellular phenotypes, including chemotactic motility. However, the regulatory mechanism for BLT2 has not yet been demonstrated. To understand the regulatory mechanism of BLT2, we screened and identified the proteins that bind to BLT2. Using a yeast two-hybrid assay with the BLT2 C-terminal domain as bait, we found that RanBPM, a previously proposed scaffold protein, interacts with BLT2. We demonstrated the specific interaction between BLT2 and RanBPM by GST pulldown assay and co-immunoprecipitation assay. To elucidate the biological function of the RanBPM-BLT2 interaction, we evaluated the effects of RanBPM overexpression or knockdown. We found that BLT2-mediated motility was severely attenuated by RanBPM overexpression and that knockdown of endogenous RanBPM by shRNA strongly promoted BLT2-mediated motility, suggesting a negative regulatory function of RanBPM toward BLT2. Furthermore, we observed that the addition of BLT2 ligands caused the dissociation of BLT2 and RanBPM, thus releasing the negative regulatory effect of RanBPM. Finally, we propose that Akt-induced BLT2 phosphorylation at residue Thr355, which occurs after the addition of BLT2 ligands, is a potential mechanism by which BLT2 dissociates from RanBPM, resulting in stimulation of BLT2 signaling. Taken together, our results suggest that RanBPM acts as a negative regulator of BLT2 signaling to attenuate BLT2-mediated cell motility. PMID:23928309

  7. Missing-in-Metastasis regulates cell motility and invasion via PTPδ-mediated changes in SRC activity

    PubMed Central

    Chaudhary, Fauzia; Lucito, Robert; Tonks, Nicholas K.

    2014-01-01

    Missing in Metastasis (MIM), also known as MTSS1, is a scaffold protein that is down-regulated in multiple metastatic cancer cell lines compared to non-metastatic counterparts. MIM regulates cytoskeletal dynamics and actin polymerization, and has been implicated in the control of cell motility and invasion. MIM has also been shown to bind to a receptor PTP, PTPδ, an interaction that may provide a link between tyrosine phosphorylation-dependent signaling and metastasis. We used shRNA-mediated gene silencing to investigate the consequences of loss of MIM on the migration and invasion of the MCF10A mammary epithelial cell model of breast cancer. We observed that suppression of MIM by RNAi enhanced migration and invasion of MCF10A cells, effects that were associated with increased levels of PTPδ. Furthermore, analysis of human clinical data indicated that PTPδ was elevated in breast cancer samples when compared to normal tissue. We demonstrated that the SRC protein tyrosine kinase is a direct substrate of PTPδ and, upon suppression of MIM, we observed changes in the phosphorylation status of SRC, in particular the inhibitory site (Tyr 527) was hypophosphorylated, whereas the activating autophosphorylation site (Tyr 416) was hyperphosphorylated. Thus, the absence of MIM led to PTPδ-mediated activation of SRC. Finally, the SRC inhibitor SU6656 counteracted the effects of MIM suppression on cell motility and invasion. This study illustrates that both SRC and PTPδ have the potential to be therapeutic targets for metastatic tumors associated with loss of MIM. PMID:25287652

  8. 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.

  9. Pea Broth Enhances the Biocontrol Efficacy of Lysobacter capsici AZ78 by Triggering Cell Motility Associated with Biogenesis of Type IV Pilus.

    PubMed

    Tomada, Selena; Puopolo, Gerardo; Perazzolli, Michele; Musetti, Rita; Loi, Nazia; Pertot, Ilaria

    2016-01-01

    Bacterial cells can display different types of motility, due to the presence of external appendages such as flagella and type IV pili. To date, little information on the mechanisms involved in the motility of the Lysobacter species has been available. Recently, L. capsici AZ78, a biocontrol agent of phytopathogenic oomycetes, showed the ability to move on jellified pea broth. Pea broth medium improved also the biocontrol activity of L. capsici AZ78 against Plasmopara viticola under greenhouse conditions. Noteworthy, the quantity of pea residues remaining on grapevine leaves fostered cell motility in L. capsici AZ78. Based on these results, this unusual motility related to the composition of the growth medium was investigated in bacterial strains belonging to several Lysobacter species. The six L. capsici strains tested developed dendrite-like colonies when grown on jellified pea broth, while the development of dendrite-like colonies was not recorded in the media commonly used in motility assays. To determine the presence of genes responsible for biogenesis of the flagellum and type IV pili, the genome of L. capsici AZ78 was mined. Genes encoding structural components and regulatory factors of type IV pili were upregulated in L. capsici AZ78 cells grown on the above-mentioned medium, as compared with the other tested media. These results provide new insight into the motility mechanism of L. capsici members and the role of type IV pili and pea compounds on the epiphytic fitness and biocontrol features of L. capsici AZ78. PMID:27507963

  10. Pea Broth Enhances the Biocontrol Efficacy of Lysobacter capsici AZ78 by Triggering Cell Motility Associated with Biogenesis of Type IV Pilus

    PubMed Central

    Tomada, Selena; Puopolo, Gerardo; Perazzolli, Michele; Musetti, Rita; Loi, Nazia; Pertot, Ilaria

    2016-01-01

    Bacterial cells can display different types of motility, due to the presence of external appendages such as flagella and type IV pili. To date, little information on the mechanisms involved in the motility of the Lysobacter species has been available. Recently, L. capsici AZ78, a biocontrol agent of phytopathogenic oomycetes, showed the ability to move on jellified pea broth. Pea broth medium improved also the biocontrol activity of L. capsici AZ78 against Plasmopara viticola under greenhouse conditions. Noteworthy, the quantity of pea residues remaining on grapevine leaves fostered cell motility in L. capsici AZ78. Based on these results, this unusual motility related to the composition of the growth medium was investigated in bacterial strains belonging to several Lysobacter species. The six L. capsici strains tested developed dendrite-like colonies when grown on jellified pea broth, while the development of dendrite-like colonies was not recorded in the media commonly used in motility assays. To determine the presence of genes responsible for biogenesis of the flagellum and type IV pili, the genome of L. capsici AZ78 was mined. Genes encoding structural components and regulatory factors of type IV pili were upregulated in L. capsici AZ78 cells grown on the above-mentioned medium, as compared with the other tested media. These results provide new insight into the motility mechanism of L. capsici members and the role of type IV pili and pea compounds on the epiphytic fitness and biocontrol features of L. capsici AZ78. PMID:27507963

  11. 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