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Sample records for adherent cell types

  1. A Clark-type oxygen chip for in situ estimation of the respiratory activity of adhering cells.

    PubMed

    Wu, Ching-Chou; Luk, Hsiang-Ning; Lin, Yen-Ting Tsai; Yuan, Chia-Yin

    2010-04-15

    A Clark-type oxygen chip consisting of a polydimethylsiloxane (PDMS) reservoir containing an amino group-modified PDMS oxygen-permeable membrane (OPM) and a glass substrate containing a three-electrode detector has been constructed by using microfabrication techniques, and it is utilized for in situ measurement of the respiration activity of adhering cells. Use of the alginate sol electrolyte and the electroplating Ag/AgCl pseudo-reference electrode can effectively diminish the crosstalk between the electrochemical electrodes and supply a stable potential for the detection of dissolved oxygen, respectively. The Clark-type oxygen chips possess only 1.00% residual current, response time of 13.4s and good linearity with a correlation coefficient of 0.9933. The modification of amino groups for the OPM obviously facilitates the adhesion of HeLa cells onto the PDMS OPM surface and allows the cells to spread after 2h of incubation. The oxygen consumption of the cells in the cell-adhesion process increases with the adhesion time, and the increment of cellular oxygen consumption per minute reaches a maximum after 30 min of incubation. Moreover, the change in the respiration activity of adhering HeLa cells stimulated by the high concentration of glucose or propofol anaesthetic can be monitored in real time with the Clark-type oxygen chip. PMID:20188913

  2. Sialidases Affect the Host Cell Adherence and Epsilon Toxin-Induced Cytotoxicity of Clostridium perfringens Type D Strain CN3718

    PubMed Central

    Li, Jihong; Sayeed, Sameera; Robertson, Susan; Chen, Jianming; McClane, Bruce A.

    2011-01-01

    Clostridium perfringens type B or D isolates, which cause enterotoxemias or enteritis in livestock, produce epsilon toxin (ETX). ETX is exceptionally potent, earning it a listing as a CDC class B select toxin. Most C. perfringens strains also express up to three different sialidases, although the possible contributions of those enzymes to type B or D pathogenesis remain unclear. Type D isolate CN3718 was found to carry two genes (nanI and nanJ) encoding secreted sialidases and one gene (nanH) encoding a cytoplasmic sialidase. Construction in CN3718 of single nanI, nanJ and nanH null mutants, as well as a nanI/nanJ double null mutant and a triple sialidase null mutant, identified NanI as the major secreted sialidase of this strain. Pretreating MDCK cells with NanI sialidase, or with culture supernatants of BMC206 (an isogenic CN3718 etx null mutant that still produces sialidases) enhanced the subsequent binding and cytotoxic effects of purified ETX. Complementation of BMC207 (an etx/nanH/nanI/nanJ null mutant) showed this effect is mainly attributable to NanI production. Contact between BMC206 and certain mammalian cells (e.g., enterocyte-like Caco-2 cells) resulted in more rapid sialidase production and this effect involved increased transcription of BMC206 nanI gene. BMC206 was shown to adhere to some (e.g. Caco-2 cells), but not all mammalian cells, and this effect was dependent upon sialidase, particularly NanI, expression. Finally, the sialidase activity of NanI (but not NanJ or NanH) could be enhanced by trypsin. Collectively these in vitro findings suggest that, during type D disease originating in the intestines, trypsin may activate NanI, which (in turn) could contribute to intestinal colonization by C. perfringens type D isolates and also increase ETX action. PMID:22174687

  3. Haemophilus influenzae Type f Hijacks Vitronectin Using Protein H To Resist Host Innate Immunity and Adhere to Pulmonary Epithelial Cells.

    PubMed

    Al-Jubair, Tamim; Mukherjee, Oindrilla; Oosterhuis, Sharon; Singh, Birendra; Su, Yu-Ching; Fleury, Christophe; Blom, Anna M; Törnroth-Horsefield, Susanna; Riesbeck, Kristian

    2015-12-15

    The incidence of invasive Haemophilus influenzae type b (Hib) disease has significantly decreased since the introduction of an efficient vaccine against Hib. However, in contrast to Hib, infections caused by H. influenzae serotype f (Hif) are emerging. We recently did a whole genome sequencing of an invasive Hif isolate, and reported that Hif interacts with factor H by expressing protein H (PH). In this study, upon screening with various human complement regulators, we revealed that PH is also a receptor for vitronectin (Vn), an abundant plasma protein that regulates the terminal pathway of the human complement system in addition to being a component of the extracellular matrix. Bacterial Vn binding was significantly reduced when the lph gene encoding PH was deleted in an invasive Hif isolate. The dissociation constant (KD) of the interaction between recombinant PH and Vn was 2.2 μM, as revealed by Biolayer interferometry. We found that PH has different regions for simultaneous interaction with both Vn and factor H, and that it recognized the C-terminal part of Vn (aa 352-362). Importantly, PH-dependent Vn binding resulted in better survival of the wild-type Hif or PH-expressing Escherichia coli when exposed to human serum. Finally, we observed that PH mediated an increased bacterial adherence to alveolar epithelial cells in the presence of Vn. In conclusion, our study reveals that PH most likely plays an important role in Hif pathogenesis by increasing serum resistance and adhesion to the airways. PMID:26538390

  4. Physics of adherent cells

    NASA Astrophysics Data System (ADS)

    Schwarz, Ulrich S.; Safran, Samuel A.

    2013-07-01

    One of the most unique physical features of cell adhesion to external surfaces is the active generation of mechanical force at the cell-material interface. This includes pulling forces generated by contractile polymer bundles and networks, and pushing forces generated by the polymerization of polymer networks. These forces are transmitted to the substrate mainly by focal adhesions, which are large, yet highly dynamic adhesion clusters. Tissue cells use these forces to sense the physical properties of their environment and to communicate with each other. The effect of forces is intricately linked to the material properties of cells and their physical environment. Here a review is given of recent progress in our understanding of the role of forces in cell adhesion from the viewpoint of theoretical soft matter physics and in close relation to the relevant experiments.

  5. Suppression of BCG cell wall induced delayed-type hypersensitivity by BCG pre-treatment. I. Induction of adherent suppressor cells by live BCG injection and their characterization.

    PubMed Central

    Kato, K; Yamamoto, K I; Kakinuma, M; Ishihara, C; Azuma, I

    1981-01-01

    Previous injections of live Bacillus Calmette-Guérin (BCG) in mice produced a suppression of delayed-type hypersensitivity (DTH) induced by oil-treated BCG cell walls (CW). This phenomenon was analysed by the macrophage migration inhibition (MI) test in which peritoneal exudate cells (PEC) from live BCG-injected mice were mixed with PEC from BCG CW-immunized mice, with the result that the former cells suppressed the MI activity in the latter. We considered the Mi test to be a reliable method for demonstrating the existence of suppressor cells induced by the injection of live BCG. Moreover, we found that the adherent cells of PEC possessed a suppressive effect which was retained even after treatment with either anti-mouse Ig or anti-brain associated theta (BA theta) antigen; that the PEC from mice injected with live BCG on at least the 12th day before cell harvesting showed the suppression; and that the suppression operated across the H-2 barrier. PMID:6450731

  6. Contributions of NanI Sialidase to Caco-2 Cell Adherence by Clostridium perfringens Type A and C Strains Causing Human Intestinal Disease

    PubMed Central

    Li, Jihong

    2014-01-01

    Previous studies showed that Clostridium perfringens type D animal disease strain CN3718 uses NanI sialidase for adhering to enterocyte-like Caco-2 cells. The current study analyzed whether NanI is similarly important when type A and C human intestinal disease strains attach to Caco-2 cells. A PCR survey determined that the nanI gene was absent from typical type A food poisoning (FP) strains carrying a chromosomal enterotoxin (CPE) gene or the genetically related type C Darmbrand (Db) strains. However, the nanI gene was present in type A strains from healthy humans, type A strains causing CPE-associated antibiotic-associated diarrhea (AAD) or sporadic diarrhea (SD), and type C Pig-Bel strains. Consistent with NanI sialidase being the major C. perfringens sialidase when produced, FP and Db strains had little supernatant sialidase activity compared to other type A or C human intestinal strains. All type A and C human intestinal strains bound to Caco-2 cells, but NanI-producing strains had higher attachment levels. When produced, NanI can contribute to host cell attachment of human intestinal disease strains, since a nanI null mutant constructed in type A SD strain F4969 had lower Caco-2 cell adhesion than wild-type F4969 or a complemented strain. Further supporting a role for NanI in host cell attachment, sialidase inhibitors reduced F4969 adhesion to Caco-2 cells. Collectively, these results suggest that NanI may contribute to the intestinal attachment and colonization needed for the chronic diarrhea of CPE-associated AAD and SD, but this sialidase appears to be dispensable for the acute pathogenesis of type A FP or type C enteritis necroticans. PMID:25135687

  7. Adherence of Bilophila wadsworthia to cultured human embryonic intestinal cells.

    PubMed

    Gerardo, S H; Garcia, M M; Wexler, H M; Finegold, S M

    1998-02-01

    Adherence of Bilophila wadsworthia to the cultured human embryonic intestinal cell line, Intestine 407 (Int 407), varied among the strains tested from strongly adherent (76-100% cells positive for one or more adherent bacteria) to non- or weakly adherent (0-25% positive cells). Although negative staining revealed that infrequent cells of an adherent strain, WAL 9077, the adherent type-strain, WAL 7959, and a non-adherent strain, WAL 8448, expressed loosely associated fimbrial structures, a role for these structures in adhesion could not be confirmed with either scanning or thin-section electron micrography. Ruthenium red staining of thin-section preparations and subsequent electron microscopy failed to reveal an extensive extracellular polysaccharide layer. SDS-PAGE analysis of crude outer membrane fractions of WAL 9077 and WAL 8448 demonstrated clear differences in their major and minor outer membrane protein components. Thus, we postulate that the adherence of B. wadsworthia to Int 407 cells is mediated by an outer membrane or cell wall component. PMID:16887620

  8. Topography Influences Adherent Cell Regulation of Osteoclastogenesis.

    PubMed

    Nagasawa, M; Cooper, L F; Ogino, Y; Mendonca, D; Liang, R; Yang, S; Mendonca, G; Uoshima, K

    2016-03-01

    The importance of osteoclast-mediated bone resorption in the process of osseointegration has not been widely considered. In this study, cell culture was used to investigate the hypothesis that the function of implant-adherent bone marrow stromal cells (BMSCs) in osteoclastogenesis is influenced by surface topography. BMSCs isolated from femur and tibia of Sprague-Dawley rats were seeded onto 3 types of titanium surfaces (smooth, micro, and nano) and a control surface (tissue culture plastic) with or without osteogenic supplements. After 3 to 14 d, conditioned medium (CM) was collected. Subsequently, rat bone marrow-derived macrophages (BMMs) were cultured in media supplemented with soluble receptor activator of NF-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) as well as BMSC CM from each of the 4 surfaces. Gene expression levels of soluble RANKL, osteoprotegerin, tumor necrosis factor α, and M-CSF in cultured BMSCs at different time points were measured by real-time polymerase chain reaction. The number of differentiated osteoclastic cells was determined after tartrate-resistant acid phosphatase staining. Analysis of variance and t test were used for statistical analysis. The expression of prominent osteoclast-promoting factors tumor necrosis factor α and M-CSF was increased by BMSCs cultured on both micro- and nanoscale titanium topographies (P < 0.01). BMSC CM contained a heat-labile factor that increased BMMs osteoclastogenesis. CM from both micro- and nanoscale surface-adherent BMSCs increased the osteoclast number (P < 0.01). Difference in surface topography altered BMSC phenotype and influenced BMM osteoclastogenesis. Local signaling by implant-adherent cells at the implant-bone interface may indirectly control osteoclastogenesis and bone accrual around endosseous implants. PMID:26553885

  9. Antibodies Directed against Shiga-Toxin Producing Escherichia coli Serotype O103 Type III Secreted Proteins Block Adherence of Heterologous STEC Serotypes to HEp-2 Cells

    PubMed Central

    Desin, Taseen S.; Townsend, Hugh G.; Potter, Andrew A.

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) serotype O103 is a zoonotic pathogen that is capable of causing hemorrhagic colitis and hemolytic uremic syndrome (HUS) in humans. The main animal reservoir for STEC is ruminants and hence reducing the levels of this pathogen in cattle could ultimately lower the risk of STEC infection in humans. During the process of infection, STECO103 uses a Type III Secretion System (T3SS) to secrete effector proteins (T3SPs) that result in the formation of attaching and effacing (A/E) lesions. Vaccination of cattle with STEC serotype O157 T3SPs has previously been shown to be effective in reducing shedding of STECO157 in a serotype-specific manner. In this study, we tested the ability of rabbit polyclonal sera against individual STECO103 T3SPs to block adherence of the organism to HEp-2 cells. Our results demonstrate that pooled sera against EspA, EspB, EspF, NleA and Tir significantly lowered the adherence of STECO103 relative to pre-immune sera. Likewise, pooled anti-STECO103 sera were also able to block adherence by STECO157. Vaccination of mice with STECO103 recombinant proteins induced strong IgG antibody responses against EspA, EspB, NleA and Tir but not against EspF. However, the vaccine did not affect fecal shedding of STECO103 compared to the PBS vaccinated group over the duration of the experiment. Cross reactivity studies using sera against STECO103 recombinant proteins revealed a high degree of cross reactivity with STECO26 and STECO111 proteins implying that sera against STECO103 proteins could potentially provide neutralization of attachment to epithelial cells by heterologous STEC serotypes. PMID:26451946

  10. Antibodies Directed against Shiga-Toxin Producing Escherichia coli Serotype O103 Type III Secreted Proteins Block Adherence of Heterologous STEC Serotypes to HEp-2 Cells.

    PubMed

    Desin, Taseen S; Townsend, Hugh G; Potter, Andrew A

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) serotype O103 is a zoonotic pathogen that is capable of causing hemorrhagic colitis and hemolytic uremic syndrome (HUS) in humans. The main animal reservoir for STEC is ruminants and hence reducing the levels of this pathogen in cattle could ultimately lower the risk of STEC infection in humans. During the process of infection, STECO103 uses a Type III Secretion System (T3SS) to secrete effector proteins (T3SPs) that result in the formation of attaching and effacing (A/E) lesions. Vaccination of cattle with STEC serotype O157 T3SPs has previously been shown to be effective in reducing shedding of STECO157 in a serotype-specific manner. In this study, we tested the ability of rabbit polyclonal sera against individual STECO103 T3SPs to block adherence of the organism to HEp-2 cells. Our results demonstrate that pooled sera against EspA, EspB, EspF, NleA and Tir significantly lowered the adherence of STECO103 relative to pre-immune sera. Likewise, pooled anti-STECO103 sera were also able to block adherence by STECO157. Vaccination of mice with STECO103 recombinant proteins induced strong IgG antibody responses against EspA, EspB, NleA and Tir but not against EspF. However, the vaccine did not affect fecal shedding of STECO103 compared to the PBS vaccinated group over the duration of the experiment. Cross reactivity studies using sera against STECO103 recombinant proteins revealed a high degree of cross reactivity with STECO26 and STECO111 proteins implying that sera against STECO103 proteins could potentially provide neutralization of attachment to epithelial cells by heterologous STEC serotypes. PMID:26451946

  11. AHCC Activation and Selection of Human Lymphocytes via Genotypic and Phenotypic Changes to an Adherent Cell Type: A Possible Novel Mechanism of T Cell Activation

    PubMed Central

    Olamigoke, Loretta; Mansoor, Elvedina; Mann, Vivek; Ellis, Ivory; Okoro, Elvis; Wakame, Koji; Fuji, Hajime; Kulkarni, Anil; Francoise Doursout, Marie; Sundaresan, Alamelu

    2015-01-01

    Active Hexose Correlated Compound (AHCC) is a fermented mushroom extract and immune supplement that has been used to treat a wide range of health conditions. It helps in augmentation of the natural immune response and affects immune cell activation and outcomes. The goal of this project was to study and understand the role and mechanisms of AHCC supplementation in the prevention of immunosuppression through T cell activation. The method described here involves “in vitro” culturing of lymphocytes, exposing them to different concentrations of AHCC (0 μg/mL, 50 μg/mL, 100 μg/mL, 250 μg/mL, and 500 μg/mL) at 0 hours. Interestingly, clumping and aggregation of the cells were seen between 24 and 72 hours of incubation. The cells lay down extracellular matrix, which become adherent, and phenotypical changes from small rounded lymphocytes to large macrophage-like, spindle shaped, elongated, fibroblast-like cells even beyond 360 hours were observed. These are probably translated from genotypic changes in the cells since the cells propagate for at least 3 to 6 generations (present observations). RNA isolated was subjected to gene array analysis. We hypothesize that cell adhesion is an activation and survival pathway in lymphocytes and this could be the mechanism of AHCC activation in human lymphocytes. PMID:26788109

  12. AHCC Activation and Selection of Human Lymphocytes via Genotypic and Phenotypic Changes to an Adherent Cell Type: A Possible Novel Mechanism of T Cell Activation.

    PubMed

    Olamigoke, Loretta; Mansoor, Elvedina; Mann, Vivek; Ellis, Ivory; Okoro, Elvis; Wakame, Koji; Fuji, Hajime; Kulkarni, Anil; Francoise Doursout, Marie; Sundaresan, Alamelu

    2015-01-01

    Active Hexose Correlated Compound (AHCC) is a fermented mushroom extract and immune supplement that has been used to treat a wide range of health conditions. It helps in augmentation of the natural immune response and affects immune cell activation and outcomes. The goal of this project was to study and understand the role and mechanisms of AHCC supplementation in the prevention of immunosuppression through T cell activation. The method described here involves "in vitro" culturing of lymphocytes, exposing them to different concentrations of AHCC (0 μg/mL, 50 μg/mL, 100 μg/mL, 250 μg/mL, and 500 μg/mL) at 0 hours. Interestingly, clumping and aggregation of the cells were seen between 24 and 72 hours of incubation. The cells lay down extracellular matrix, which become adherent, and phenotypical changes from small rounded lymphocytes to large macrophage-like, spindle shaped, elongated, fibroblast-like cells even beyond 360 hours were observed. These are probably translated from genotypic changes in the cells since the cells propagate for at least 3 to 6 generations (present observations). RNA isolated was subjected to gene array analysis. We hypothesize that cell adhesion is an activation and survival pathway in lymphocytes and this could be the mechanism of AHCC activation in human lymphocytes. PMID:26788109

  13. Robotic adherent cell injection for characterizing cell-cell communication.

    PubMed

    Liu, Jun; Siragam, Vinayakumar; Gong, Zheng; Chen, Jun; Fridman, Michael D; Leung, Clement; Lu, Zhe; Ru, Changhai; Xie, Shaorong; Luo, Jun; Hamilton, Robert M; Sun, Yu

    2015-01-01

    Compared to robotic injection of suspended cells (e.g., embryos and oocytes), fewer attempts were made to automate the injection of adherent cells (e.g., cancer cells and cardiomyocytes) due to their smaller size, highly irregular morphology, small thickness (a few micrometers thick), and large variations in thickness across cells. This paper presents a robotic system for automated microinjection of adherent cells. The system is embedded with several new capabilities: automatically locating micropipette tips; robustly detecting the contact of micropipette tip with cell culturing surface and directly with cell membrane; and precisely compensating for accumulative positioning errors. These new capabilities make it practical to perform adherent cell microinjection truly via computer mouse clicking in front of a computer monitor, on hundreds and thousands of cells per experiment (versus a few to tens of cells as state of the art). System operation speed, success rate, and cell viability rate were quantitatively evaluated based on robotic microinjection of over 4000 cells. This paper also reports the use of the new robotic system to perform cell-cell communication studies using large sample sizes. The gap junction function in a cardiac muscle cell line (HL-1 cells), for the first time, was quantified with the system. PMID:25073160

  14. Type IV Pili in Francisella tularensis: Roles of pilF and pilT in Fiber Assembly, Host Cell Adherence, and Virulence ▿

    PubMed Central

    Chakraborty, Subhra; Monfett, Michael; Maier, Tamara M.; Benach, Jorge L.; Frank, Dara W.; Thanassi, David G.

    2008-01-01

    Francisella tularensis, a highly virulent facultative intracellular bacterium, is the causative agent of tularemia. Genome sequencing of all F. tularensis subspecies revealed the presence of genes that could encode type IV pili (Tfp). The live vaccine strain (LVS) expresses surface fibers resembling Tfp, but it was not established whether these fibers were indeed Tfp encoded by the pil genes. We show here that deletion of the pilF putative Tfp assembly ATPase in the LVS resulted in a complete loss of surface fibers. Disruption of the pilT putative disassembly ATPase also caused a complete loss of pili, indicating that pilT functions differently in F. tularensis than in model Tfp systems such as those found in Pseudomonas aeruginosa and Neisseria spp. The LVS pilF and pilT mutants were attenuated for virulence in a mouse model of tularemia by the intradermal route. Furthermore, although absence of pili had no effect on the ability of the LVS to replicate intracellularly, the pilF and pilT mutants were defective for adherence to macrophages, pneumocytes, and hepatocytes. This work confirms that the surface fibers expressed by the LVS are encoded by the pil genes and provides evidence that the Francisella pili contribute to host cell adhesion and virulence. PMID:18426883

  15. Vaccine production: upstream processing with adherent or suspension cell lines.

    PubMed

    Genzel, Yvonne; Rödig, Jana; Rapp, Erdmann; Reichl, Udo

    2014-01-01

    The production of viral vaccines in cell culture can be accomplished with primary, diploid, or continuous (transformed) cell lines. Each cell line, each virus type, and each vaccine preparation require the specific design of upstream and downstream processing. Media have to be selected as well as production vessels, cultivation conditions, and modes of operation. Many viruses only replicate to high titers in adherently growing cells, but similar to processes established for recombinant protein production, an increasing number of suspension cell lines is being evaluated for future use. Here, we describe key issues to be considered for the establishment of large-scale virus production in bioreactors. As an example upstream processing of cell culture-derived influenza virus production is described in more detail for adherently growing and for suspension cells. In particular, use of serum-containing, serum-free, and chemically defined media as well as choice of cultivation vessel are considered. PMID:24297427

  16. Micromolded Arrays for Separation of Adherent Cells

    PubMed Central

    Wang, Yuli; Phillips, Colleen; Xu, Wei; Pai, Jeng-Hao; Dhopeshwarkar, Rahul; Sims, Christopher E.; Allbritton, Nancy

    2010-01-01

    We present an efficient, yet inexpensive, approach for isolating viable single cells or colonies from a mixed population. This cell microarray platform possesses innovations in both the array manufacture and the manner of target cell release. Arrays of microwells with bases composed of detachable concave elements, termed microrafts, were fabricated by a dip-coating process using a polydimethylsiloxane mold as the template and the array substrate. This manufacturing approach enabled the use of materials other than photoresists to create the array elements. Thus microrafts possessing low autofluorescence could be fabricated for fluorescence-based identification of cells. Cells plated on the microarray settled and attached at the center of the wells due to the microrafts’ concavity. Individual microrafts were readily dislodged by the action of a needle inserted through the compliant polymer substrate. The hard polymer material (polystyrene or epoxy resin) of which the microrafts were composed protected the cells from damage by the needle. For cell analysis and isolation, cells of interest were identified using a standard inverted microscope and microrafts carrying target cells were dislodged with the needle. The released cells/microrafts could be efficiently collected, cultured and clonally expanded. During the separation and collection procedures, the cells remained adherent and provided a measure of protection during manipulation, thus providing an extremely high single-cell cloning rate (>95%). Generation of a transfected cell line based on expression of a fluorescent protein demonstrated an important application for performing on-chip cell separations. PMID:20838672

  17. A method for incorporating macromolecules into adherent cells

    PubMed Central

    1984-01-01

    We describe a simple method for loading exogenous macromolecules into the cytoplasm of mammalian cells adherent to tissue culture dishes. Culture medium was replaced with a thin layer of fluorescently labeled macromolecules, the cells were harvested from the substrate by scraping with a rubber policeman, transferred immediately to ice cold media, washed, and then replated for culture. We refer to the method as "scrape-loading." Viability of cells was 50-60% immediately after scrape-loading and was 90% for those cells remaining after 24 h of culture. About 40% of adherent, well-spread fibroblasts contained fluorescent molecules 18 h after scrape-loading of labeled dextrans, ovalbumin, or immunoglobulin-G. On average, 10(7) dextran molecules (70,000-mol wt) were incorporated into each fibroblast by scrape- loading in 10 mg/ml dextran. The extent of loading depended on the concentration and molecular weight of the dextrans used. A fluorescent analog of actin could also be loaded into fibroblasts where it labeled stress fibers. HeLa cells, a macrophage-like cell line, 1774A.1, and human neutrophils were all successfully loaded with dextran by scraping. The method of scrape-loading should be applicable to a broad range of adherent cell types, and useful for loading of diverse kinds of macromolecules. PMID:6201494

  18. Active mechanics and geometry of adherent cells and cell colonies

    NASA Astrophysics Data System (ADS)

    Banerjee, Shiladitya

    2014-03-01

    Measurements of traction stresses exerted by adherent cells or cell colonies on elastic substrates have yielded new insight on how the mechanical and geometrical properties of the substrate regulate cellular force distribution, mechanical energy, spreading, morphology or stress ber architecture. We have developed a generic mechanical model of adherent cells as an active contractile gel mechanically coupled to an elastic substrate and to neighboring cells in a tissue. The contractile gel model accurately predicts the distribution of cellular and traction stresses as observed in single cell experiments, and captures the dependence of cell shape, traction stresses and stress ber polarization on the substrate's mechanical and geometrical properties. The model further predicts that the total strain energy of an adherent cell is solely regulated by its spread area, in agreement with recent experiments on micropatterned substrates with controlled geometry. When used to describe the behavior of colonies of adherent epithelial cells, the model demonstrates the crucial role of the mechanical cross-talk between intercellular and extracellular adhesion in regulating traction force distribution. Strong intercellular mechanical coupling organizes traction forces to the colony periphery, whereas weaker intercellular coupling leads to the build up of traction stresses at intercellular junctions. Furthermore, in agreement with experiments on large cohesive keratinocyte colonies, the model predicts a linear scaling of traction forces with the colony size. This scaling suggests the emergence of an effective surface tension as a scale-free material property of the adherent tissue, originating from actomyosin contractility.

  19. Broadening cell selection criteria with micropallet arrays of adherent cells.

    PubMed

    Wang, Yuli; Young, Grace; Aoto, Phillip C; Pai, Jeng-Hao; Bachman, Mark; Li, G P; Sims, Christopher E; Allbritton, Nancy L

    2007-10-01

    A host of technologies exists for the separation of living, nonadherent cells, with separation decisions typically based on fluorescence or immunolabeling of cells. Methods to separate adherent cells as well as to broaden the range of possible sorting criteria would be of high value and complementary to existing strategies. Cells were cultured on arrays of releasable pallets. The arrays were screened and individual cell(s)/pallets were released and collected. Conventional fluorescence and immunolabeling of cells were compatible with the pallet arrays, as were separations based on gene expression. By varying the size of the pallet and the number of cells cultured on the array, single cells or clonal colonies of cells were isolated from a heterogeneous population. Since cells remained adherent throughout the isolation process, separations based on morphologic characteristics, for example cell shape, were feasible. Repeated measurements of each cell in an array were performed permitting the selection of cells based on their temporal behavior, e.g. growth rate. The pallet array system provides the flexibility to select and collect adherent cells based on phenotypic and temporal criteria and other characteristics not accessible by alternative methods. PMID:17559133

  20. Contractile network models for adherent cells.

    PubMed

    Guthardt Torres, P; Bischofs, I B; Schwarz, U S

    2012-01-01

    Cells sense the geometry and stiffness of their adhesive environment by active contractility. For strong adhesion to flat substrates, two-dimensional contractile network models can be used to understand how force is distributed throughout the cell. Here we compare the shape and force distribution for different variants of such network models. In contrast to Hookean networks, cable networks reflect the asymmetric response of biopolymers to tension versus compression. For passive networks, contractility is modeled by a reduced resting length of the mechanical links. In actively contracting networks, a constant force couple is introduced into each link in order to model contraction by molecular motors. If combined with fixed adhesion sites, all network models lead to invaginated cell shapes, but only actively contracting cable networks lead to the circular arc morphology typical for strongly adhering cells. In this case, shape and force distribution are determined by local rather than global determinants and thus are suited to endow the cell with a robust sense of its environment. We also discuss nonlinear and adaptive linker mechanics as well as the relation to tissue shape. PMID:22400597

  1. Contractile network models for adherent cells

    NASA Astrophysics Data System (ADS)

    Guthardt Torres, P.; Bischofs, I. B.; Schwarz, U. S.

    2012-01-01

    Cells sense the geometry and stiffness of their adhesive environment by active contractility. For strong adhesion to flat substrates, two-dimensional contractile network models can be used to understand how force is distributed throughout the cell. Here we compare the shape and force distribution for different variants of such network models. In contrast to Hookean networks, cable networks reflect the asymmetric response of biopolymers to tension versus compression. For passive networks, contractility is modeled by a reduced resting length of the mechanical links. In actively contracting networks, a constant force couple is introduced into each link in order to model contraction by molecular motors. If combined with fixed adhesion sites, all network models lead to invaginated cell shapes, but only actively contracting cable networks lead to the circular arc morphology typical for strongly adhering cells. In this case, shape and force distribution are determined by local rather than global determinants and thus are suited to endow the cell with a robust sense of its environment. We also discuss nonlinear and adaptive linker mechanics as well as the relation to tissue shape.

  2. Adherence of Helicobacter pylori to primary human gastrointestinal cells.

    PubMed Central

    Clyne, M; Drumm, B

    1993-01-01

    Helicobacter pylori adheres only to gastric cells in vivo. However, the organism adheres to a wide variety of nongastric cells in vitro. In this study, we have used flow cytometry to assess the adherence of H. pylori to primary epithelial cells isolated from gastric, duodenal, and colonic biopsy specimens by collagenase digestion. After incubation of bacteria and cells together and subsequent staining with a two-stage fluorescein isothiocyanate-labelled H. pylori antibody method, cells with adherent bacteria could be easily distinguished from cells without bacteria. Binding to Kato III cells (a gastric adenocarcinoma cell line) was saturable when bacteria and cells were mixed at a ratio of 250:1. Adherence to cells isolated from gastric biopsy specimens was significantly better than adherence to cells isolated from duodenal or colonic biopsy specimens. Almost 70% of gastric cells had bacteria bound, in contrast to 30% of duodenal cells and 32% of colonic cells (P < 0.0001). There was no correlation between expression of hemagglutinins by the bacteria and ability to bind to either Kato III cells or primary epithelial cells isolated from gastric biopsy specimens. In view of the strict tropism that the organism exhibits in vivo for gastric cells, the results of this study indicate that primary cells are ideal for assessing the factors that might play a role in the pathogenesis of disease caused by the organism. Images PMID:8406792

  3. Human placenta-derived adherent cells induce tolerogenic immune responses.

    PubMed

    Liu, Wei; Morschauser, Andrew; Zhang, Xin; Lu, Xiaohua; Gleason, Joseph; He, Shuyang; Chen, Hong-Jung; Jankovic, Vladimir; Ye, Qian; Labazzo, Kristen; Herzberg, Uri; Albert, Vivian R; Abbot, Stewart E; Liang, Bitao; Hariri, Robert

    2014-05-01

    Human placenta-derived adherent cells (PDAC cells) are a culture expanded, undifferentiated mesenchymal-like population derived from full-term placental tissue, with immunomodulatory and anti-inflammatory properties. PDA-001 (cenplacel-L), an intravenous formulation of PDAC cells, is in clinical development for the treatment of autoimmune and inflammatory diseases. To elucidate the mechanisms underlying the immunoregulatory properties of PDAC cells, we investigated their effects on immune cell populations, including T cells and dendritic cells (DC) in vitro and in vivo. PDAC cells suppressed T-cell proliferation in an OT-II T-cell adoptive transfer model, reduced the severity of myelin oligodendrocyte glycoprotein peptide-induced experimental autoimmune encephalomyelitis and ameliorated inflammation in a delayed type hypersensitivity response model. In vitro, PDAC cells suppressed T-cell proliferation and inhibited Th1 and Th17 differentiation. Analysis of tissues derived from PDAC cell-treated animals revealed diminished CD86 expression on splenic DC, suggesting that they can also modulate DC populations. Furthermore, PDAC cells modulate the differentiation and maturation of mouse bone marrow-derived DC. Similarly, human DC differentiated from CD14(+) monocytes in the presence of PDAC cells acquired a tolerogenic phenotype. These tolerogenic DC failed to induce allogeneic T-cell proliferation and differentiation toward Th1, but skewed T-cell differentiation toward Th2. Inhibition of cyclo-oxygenase-2 activity resulted in a significant, but not complete, abrogation of PDAC cells' effects on DC phenotype and function, implying a role for prostaglandin E2 in PDAC-mediated immunomodulation. This study identifies modulation of DC differentiation toward immune tolerance as a key mechanism underlying the immunomodulatory activities of PDAC cells. PMID:25505962

  4. Effect of hydrostatic pressure on the viability of non-adherent HL-60 cells

    NASA Astrophysics Data System (ADS)

    Yabuki, Takahiro; Yamanoha, Banri; Shimizu, Akio

    2013-06-01

    We investigated the effect of hydrostatic pressure on the viability of non-adherent HL-60 cell line derived from leukemic cells over a high pressure range. The HL-60 cells are resistant to pressures of up to 100 MPa under pressurization for 20 min at 25°C. However, cell viability decreased markedly between 100 and 200 MPa, and almost all cells died above 200 MPa. In the case of pressures up to 25 MPa at 25°C for four days, the viability of HL-60 cells was inhibited by increasing the pressure above 20 MPa. Although high viability was observed between 1.6 and 2.0 MPa for adherent astrocytes, viability did not change over pressures up to 2.0 MPa in the case of non-adherent HL-60 cells. It is thought that the response of cells to pressure varies among cell types.

  5. Enhanced adherence of methicillin-resistant Staphylococcus pseudintermedius sequence type 71 to canine and human corneocytes

    PubMed Central

    2014-01-01

    The recent worldwide spread of methicillin-resistant Staphylococcus pseudintermedius (MRSP) in dogs is a reason for concern due to the typical multidrug resistance patterns displayed by some MRSP lineages such as sequence type (ST) 71. The objective of this study was to compare the in vitro adherence properties between MRSP and methicillin-susceptible (MSSP) strains. Four MRSP, including a human and a canine strain belonging to ST71 and two canine non-ST71 strains, and three genetically unrelated MSSP were tested on corneocytes collected from five dogs and six humans. All strains were fully characterized with respect to genetic background and cell wall-anchored protein (CWAP) gene content. Seventy-seven strain-corneocyte combinations were tested using both exponential- and stationary-phase cultures. Negative binomial regression analysis of counts of bacterial cells adhering to corneocytes revealed that adherence was significantly influenced by host and strain genotype regardless of bacterial growth phase. The two MRSP ST71 strains showed greater adherence than MRSP non-ST71 (p < 0.0001) and MSSP (p < 0.0001). This phenotypic trait was not associated to any specific CWAP gene. In general, S. pseudintermedius adherence to canine corneocytes was significantly higher compared to human corneocytes (p < 0.0001), but the MRSP ST71 strain of human origin adhered equally well to canine and human corneocytes, suggesting that MRSP ST71 may be able to adapt to human skin. The genetic basis of the enhanced in vitro adherence of ST71 needs to be elucidated as this phenotypic trait may be associated to the epidemiological success and zoonotic potential of this epidemic MRSP clone. PMID:24957656

  6. Proliferative activity of vervet monkey bone marrow-derived adherent cells

    SciTech Connect

    Kramvis, A.; Garnett, H.M.

    1987-11-01

    Vervet monkey bone marrow-derived adherent cell population cultured in Fischer's medium supplemented with 12.5% fetal calf serum and 12.5% horse serum consists of two cell shapes: fusiform (type I) and polygonal (type II). Limiting-dilution cloning of the cells suggested that the two morphologically distinct cell types belong to the same cellular system even though they differ in their proliferative capabilities. The labeling index of type II cells, as measured by autoradiography, was found to be consistently lower than that of type I cells. It is probable that these two phenotypes represent different stages of differentiation, where progenitor type I gives rise to type II cells. The bone marrow-derived adherent cells were found to be cytokinetically at rest in vivo, using the thymidine suicide test, and relatively radioresistant with a D0 = 2.1 Gy and n = 2.36 at the time of explantation from the bone. Furthermore, in culture these cells are characterized by a relatively long cell cycle of 60 h, where the length of the S phase is 30 h, G2 is 12 h, M is 6 h, and G1 is 12 h. Thus, the vervet monkey bone marrow-derived adherent cells represent a cell population with a low turnover rate both in vivo and in vitro.

  7. Adherence of Tritrichomonas foetus to bovine vaginal epithelial cells.

    PubMed Central

    Corbeil, L B; Hodgson, J L; Jones, D W; Corbeil, R R; Widders, P R; Stephens, L R

    1989-01-01

    Adherence of Tritrichomonas foetus to bovine vaginal epithelial cells (VECs) in vitro was investigated with fresh washed bovine VECs and log-phase cultures of T. foetus. Observation under phase-contrast microscopy showed that T. foetus usually adhered first by the posterior flagellum and later by the body. Significantly more keratinized squamous epithelial cells were detected with attached parasites than nonkeratinized round epithelial cells. The optimal pH range for attachment was 6.0 to 7.5, with peak attachment at pH 6.5 for squamous VECs. Surface-reactive bovine antiserum to T. foetus prevented adherence to bovine squamous VECs. Inhibition of adherence occurred at nonagglutinating, nonimmobilizing serum dilutions. Antiserum fractions enriched for immunoglobulin G1 inhibited adherence, but fractions enriched for immunoglobulin G2 did not. The inhibitory antiserum was specific for several medium- to high-molecular-weight membrane antigens as detected in Western blots (immunoblots). The ability of surface-reactive antibodies to prevent adherence and to agglutinate and immobilize T. foetus indicates that they may be protective. Images PMID:2471692

  8. Micropallet arrays for the separation of single, adherent cells.

    PubMed

    Salazar, Georgina To'a; Wang, Yuli; Young, Grace; Bachman, Mark; Sims, Christopher E; Li, G P; Allbritton, Nancy L

    2007-01-15

    The selection and collection of single cells from within a heterogeneous population is required to produce genetically engineered cell lines, to develop new stem cell lines, and for single-cell studies. We describe a new platform for the positive selection of single live mammalian cells while the cells remain adherent to their growth surface. Cells were grown on arrays of microfabricated, releasable elements composed of SU-8 polymer termed "cell pallets". The presence of air between the elements restricted the cells to the top surfaces of the pallets. Single pallets situated within large arrays of pallets were released on demand using a single, focused, laser pulse. The laser pulses were low in energy (2-5 muJ) and did not detach nearby, nontargeted pallets. Since the SU-8 pallets and the underlying glass substrate were optically transparent, the cells on the pallets could be visualized by microscopy before and after release. Over 90% of cells remained attached to the pallet during laser-based release. The feasibility of growing the cells from the released pallets into clonal colonies was demonstrated. The pallet array system permits adherent cells to be inspected using conventional microscopy and selected cells released for further analysis. The ability to assess cells while they remain adherent to a surface will broaden the number of attributes that can be utilized for cell separation, for example, cell shape, cytoskeletal properties, and other attributes. PMID:17222037

  9. Effective interventions to improve medication adherence in Type 2 diabetes: a systematic review

    PubMed Central

    Williams, Joni L Strom; Walker, Rebekah J; Smalls, Brittany L; Campbell, Jennifer A; Egede, Leonard E

    2014-01-01

    SUMMARY Aim Medication adherence is associated with improved outcomes in diabetes. Interventions have been established to help improve medication adherence; however, the most effective interventions in patients with Type 2 diabetes remain unclear. The goal of this study was to distinguish whether interventions were effective and identify areas for future research. Methods Medline was searched for articles published between January 2000 and May 2013, and a reproducible strategy was used. Study eligibility criteria included interventions measuring medication adherence in adults with Type 2 diabetes. Results Twenty seven studies met the inclusion criteria and 13 showed a statistically significant change in medication adherence. Conclusion Heterogeneity of the study designs and measures of adherence made it difficult to identify effective interventions that improved medication adherence. Additionally, medication adherence may not be solely responsible for achieving glycemic control. Researchers must emphasize tailored interventions that optimize management and improve outcomes, and examine the need for clear indicators of medication adherence. PMID:25214893

  10. Elasticity of adherent active cells on a compliant substrate

    NASA Astrophysics Data System (ADS)

    Banerjee, Shiladitya; Mertz, Aaron F.; Dufresne, Eric R.; Marchetti, M. Cristina

    2012-02-01

    We present a continuum mechanical model of rigidity sensing by livings cells adhering to a compliant substrate. The cell or cell colony is modeled as an elastic active gel, adapting recently developed continuum theories of active viscoelastic fluids. The coupling to the substrate enters as a boundary condition that relates the cell's deformation field to local stress gradients. In the presence of activity, the substrate induces spatially inhomogeneous contractile stresses and deformations, with a power law dependence of the total traction forces on cell or colony size. This is in agreement with recent experiments on keratinocyte colonies adhered to fibronectin coated surfaces. In the presence of acto-myosin activity, the substrate also enhances the cell polarization, breaking the cell's front-rear symmetry. Maximal polarization is observed when the substrate stiffness matches that of the cell, in agreement with experiments on stem cells.

  11. Growth and adherence on stainless steel by Enterococcus faecium cells.

    PubMed

    Andrade, N J; Ajao, D B; Zottola, E A

    1998-11-01

    Enterococcus faecium isolated from Brazilian raw milk was used in this study. For growth studies, E. faecium was inoculated into 10% RSM (reconstituted skim milk) and MRS both, incubated at 6.5 and 9 degrees C for 10 days and at 30, 42, and 45 degrees C for 48 h. Cells were enumerated after spread-plating onto MRS agar and incubating at 30 degrees C for 48 h. The ability of E. faecium cells to adhere to stainless-steel chips (6 by 6 by 1 mm, AISI 304, finish #4) was investigated. MRS broth containing stainless steel chips was inoculated to an initial concentration of 10(3) or 10(6) CFU/ml of E. faecium. Adherent cells were stained with acridine orange and enumerated by epifluorescence microscopy. E. faecium grew between 6.5 and 42 degrees C in MRS and between 9 and 40 degrees C in RSM. In MRS broth with 10(6) or 10(3) CFU/ml, the g (generation time) values were 0.62 and 0.42 h and R (growth rate) values were 1.6 and 2.4 h-1. Values of R = 2.3 h-1 and g = 0.43 h were determined for E. faecium growing in RSM with 10(3) CFU/ml. In MRS broth, for samples with a starting concentration of 10(6) cells per ml, adherence to stainless-steel chips was first observed at 2 h. However, adherence was first observed at 4 h in samples with an initial concentration of 10(3) cells per ml. After 10 h of exposure the number of adherent cells was similar for all samples regardless of initial inoculum. These results indicate that E. faecium readily adheres to stainless steel. It also underscores the need to control E. faecium by using appropriate low storage temperatures and adequate sanitizing practices in the dairy industry. PMID:9829184

  12. Satisfaction with the Health Care Provider and Regimen Adherence in Minority Youth with Type 1 Diabetes.

    PubMed

    Taylor, Cortney J; La Greca, Annette; Valenzuela, Jessica M; Hsin, Olivia; Delamater, Alan M

    2016-09-01

    To assess whether satisfaction with the health-care provider is related to regimen adherence among primarily minority youth with type 1 diabetes. Youth with type 1 diabetes (n = 169; M age = 13.88; 52 % female; 70 % Hispanic) and their parents completed questionnaires that assessed their own satisfaction with the health-care provider and youths' adherence to diabetes self-care behaviors. Higher youth and parent patient-provider relationship satisfaction was associated with higher regimen adherence. Gender affected the relationship between satisfaction and regimen adherence, such that for girls, greater satisfaction was associated with better adherence; this was not the case for boys. Patient satisfaction with the health care provider is important for regimen adherence among primarily minority youth with type 1 diabetes, particularly for girls. Future research might focus on improving youths' relationships with their health care providers as a potential pathway to improve regimen adherence. PMID:27365095

  13. Microchamber Device for Detection of Transporter Activity of Adherent Cells

    PubMed Central

    Tsugane, Mamiko; Uejima, Etsuko; Suzuki, Hiroaki

    2015-01-01

    We present a method to detect the transporter activity of intact adherent cells using a microchamber device. When adherent cells are seeded onto the poly-di-methyl siloxane substrate having microchambers with openings smaller than the size of a cell, the cells form a confluent layer that covers the microchambers, creating minute, confined spaces. As substances exported across the cell membrane accumulate, transporter activity can be detected by observing the fluorescence intensity increase in the microchamber. We tested the microchamber device with HeLa cells over-expressing MDR1, an ATP-binding cassette transporter, and succeeded in detecting the transport of fluorescence-conjugated paclitaxel, the anti-cancer drug, at the single-cell level. PMID:25853126

  14. Surface modification of closed plastic bags for adherent cell cultivation

    NASA Astrophysics Data System (ADS)

    Lachmann, K.; Dohse, A.; Thomas, M.; Pohl, S.; Meyring, W.; Dittmar, K. E. J.; Lindenmeier, W.; Klages, C.-P.

    2011-07-01

    In modern medicine human mesenchymal stem cells are becoming increasingly important. However, a successful cultivation of this type of cells is only possible under very specific conditions. Of great importance, for instance, are the absence of contaminants such as foreign microbiological organisms, i.e., sterility, and the chemical functionalization of the ground on which the cells are grown. As cultivation of these cells makes high demands, a new procedure for cell cultivation has been developed in which closed plastic bags are used. For adherent cell growth chemical functional groups have to be introduced on the inner surface of the plastic bag. This can be achieved by a new, atmospheric-pressure plasma-based method presented in this paper. The method which was developed jointly by the Fraunhofer IST and the Helmholtz HZI can be implemented in automated equipment as is also shown in this contribution. Plasma process gases used include helium or helium-based gas mixtures (He + N2 + H2) and vapors of suitable film-forming agents or precursors such as APTMS, DACH, and TMOS in helium. The effect of plasma treatment is investigated by FTIR-ATR spectroscopy as well as surface tension determination based on contact angle measurements and XPS. Plasma treatment in nominally pure helium increases the surface tension of the polymer foil due to the presence of oxygen traces in the gas and oxygen diffusing through the gas-permeable foil, respectively, reacting with surface radical centers formed during contact with the discharge. Primary amino groups are obtained on the inner surface by treatment in mixtures with nitrogen and hydrogen albeit their amount is comparably small due to diffusion of oxygen through the gas-permeable bag, interfering with the plasma-amination process. Surface modifications introducing amino groups on the inner surface turned out to be most efficient in the promotion of cell growth.

  15. Adherence, accumulation, and cell division of a natural adherent bacterial population.

    PubMed Central

    Bloomquist, C G; Reilly, B E; Liljemark, W F

    1996-01-01

    Developing dental bacterial plaques formed in vivo on enamel surfaces were examined in specimens from 18 adult volunteers during the first day of plaque formation. An intraoral model placing enamel pieces onto teeth was used to study bacterial plaque populations developing naturally to various cell densities per square millimeter of surface area of the enamel (W. F. Liljemark, C. G. Bloomquist, C. L. Bandt, B. L. Philstrom, J. E. Hinrichs, and L. F. Wolff, Oral Microbiol. Immunol. 8:5-15, 1993). Radiolabeled nucleoside incorporation was used to measure DNA synthesis concurrent with the taking of standard viable cell counts of the plaque samples. Results showed that in vivo plaque formation began with the rapid adherence of bacteria until ca. 12 to 32% of the enamel's salivary pellicle was saturated (ca. 2.5 x 10(5) to 6.3 x 10(5) cells per mm2). The pioneer adherent species were predominantly those of the "sanguis streptococci." At the above-noted density, the bacteria present on the salivary pellicle incorporated low levels of radiolabeled nucleoside per viable cell. As bacterial numbers reached densities between 8.0 x 10(5) and 2.0 x 10(6) cells per mm2, there was a small increase in the incorporation of radiolabeled nucleosides per cell. At 2.5 x 10(6) to 4.0 x 10(6) cells per mm2 of enamel surface, there was a marked increase in the incorporation of radiolabeled nucleosides per cell which appeared to be cell-density dependent. The predominant species group in developing dental plaque films during density-dependent growth was the sanguis streptococci; however, most other species present showed similar patterns of increased DNA synthesis as the density noted above approached 2.5 x 10(6) to 4.0 x 10(6) cells per mm2. PMID:8576054

  16. Silk screen based dual spin-filter module for perfusion culture of adherent and non-adherent mammalian cells.

    PubMed

    Kamthan, Shweta; Gomes, James; Roychoudhury, Pradip K

    2014-08-01

    Spin-filters have been primarily used for producing therapeutic proteins from mammalian cells. However, disposability and/or high filter clogging of the existing spin-filter systems affect the process economy and productivity. Hence, to address these drawbacks a reusable dual spin-filter module for perfusion culture of adherent and non-adherent mammalian cells was designed. Two non-woven Bombyx mori silk layers were used as filter screen; the outer layer was conducive to cell attachment whilst the inner was non-conducive. Adherent cells can be cultured either in suspended mode using its inner single module or as monolayer of cells using its dual concentric module. We achieved 30 % higher urokinase productivity as compared to the stainless-steel spin-filter during perfusion experiments of adherent human kidney cells in suspended mode. This was due to the hydrophobic and negatively-charged silk screen that allows clog-free perfusion culture for prolonged periods. PMID:24737079

  17. Effect of Lewis blood group antigen expression on bacterial adherence to COS-1 cells.

    PubMed Central

    Gaffney, R A; Schaeffer, A J; Anderson, B E; Duncan, J L

    1994-01-01

    Epithelial cells from secretor individuals demonstrate decreased bacterial adherence compared with cells from nonsecretors. Lewis blood group antigen expression is one component of the secretor/nonsecretor phenotype and several epidemiologic studies have suggested a link between Lewis blood group antigen phenotype and susceptibility to urinary tract infections. In this study, we examined the possibility that the expression of the difucosylated Lewis blood group determinants, Leb and Ley (associated with the secretor phenotype), made cells less susceptible to Escherichia coli adherence by masking receptors for pili. COS-1 cells, which do not produce Lewis (Lea, Leb, Le(x), and Ley) blood group antigens, were used as target cells for bacterial adherence. The surface blood group antigen expression pattern of the cells was then modified by cotransfection with plasmids containing DNA inserts encoding alpha (1,2)-fucosyltransferase and alpha (1,3)- and alpha (1,4)-fucosyltransferases, resulting in the expression of Leb and Ley. E. coli HB101 expressing various adhesins (type 1, PapJ96, PapIA2, PapAD110, Prs, and S) from recombinant plasmids bound equally well to untransfected cells and transfected cells expressing Lea and Le(x) (nonsecretor phenotype) and Leb and Ley (secretor phenotype) antigens. We conclude that the presence of Leb and Ley antigens on cells from secretors does not alone mask receptors for E. coli pili or hinder bacterial adherence. PMID:8005692

  18. Initial adherence of EPEC, EHEC and VTEC to host cells

    PubMed Central

    Bardiau, Marjorie; Szalo, Mihai; Mainil, Jacques G.

    2010-01-01

    Initial adherence to host cells is the first step of the infection of enteropathogenic Escherichia coli (EPEC), enterohaemorrhagic Escherichia coli (EHEC) and verotoxigenic Escherichia coli (VTEC) strains. The importance of this step in the infection resides in the fact that (1) adherence is the first contact between bacteria and intestinal cells without which the other steps cannot occur and (2) adherence is the basis of host specificity for a lot of pathogens. This review describes the initial adhesins of the EPEC, EHEC and VTEC strains. During the last few years, several new adhesins and putative colonisation factors have been described, especially in EHEC strains. Only a few adhesins (BfpA, AF/R1, AF/R2, Ral, F18 adhesins) appear to be host and pathotype specific. The others are found in more than one species and/or pathotype (EPEC, EHEC, VTEC). Initial adherence of EPEC, EHEC and VTEC strains to host cells is probably mediated by multiple mechanisms. PMID:20423697

  19. Measuring insulin adherence among adults with type 2 diabetes.

    PubMed

    Osborn, Chandra Y; Gonzalez, Jeffery S

    2016-08-01

    Non-adherence to insulin is common and associated with suboptimal health. We adapted the Morisky Medication Adherence Scale to specify insulin adherence (MIAS) and compared it to the Adherence to Refills and Medication Scale for Diabetes (ARMS-D) and the Summary of Diabetes Self-Care Activities medications subscale (SDSCA-MS) and an insulin-specific (SDSCA-IS) version. A sample of 144 insulin-treated adults (58 % African American/Black, 34 % Caucasian/White, 8 % Other/Mixed race; 6.9 % Hispanic) completed these measures along with a HbA1C test. The internal consistency and factor structure of the MIAS were adequate; 59 % of participants forgot to take insulin and 46 % reported non-adherence. The MIAS was associated with the ARMS-D, SDSCA-MS, and SDSCA-IS (p < 0.001), and higher MIAS scores were marginally associated with better self-rated health (p = 0.057), but significantly associated with fewer emergency room visits (p = 0.001), and better HbA1C (p = 0.001). The MIAS is a valid and reliable insulin adherence assessment tool for practice and research applications. PMID:27062271

  20. A Selective and Purification-Free Strategy for Labeling Adherent Cells with Inorganic Nanoparticles.

    PubMed

    Gao, Yu; Lim, Jing; Yeo, David Chen Loong; Liao, Shanshan; Lans, Malin; Wang, Yaqi; Teoh, Swee-Hin; Goh, Bee Tin; Xu, Chenjie

    2016-03-01

    Cellular labeling with inorganic nanoparticles such as magnetic iron oxide nanoparticles, quantum dots, and fluorescent silica nanoparticles is an important method for the noninvasive visualization of cells using various imaging modalities. Currently, this is mainly achieved through the incubation of cultured cells with the nanoparticles that eventually reach the intracellular compartment through specific or nonspecific internalization. This classic method is advantageous in terms of simplicity and convenience, but it suffers from issues such as difficulties in fully removing free nanoparticles (suspended in solution) and the lack of selectivity on cell types. This article reports an innovative strategy for the specific labeling of adherent cells without the concern of freely suspended nanoparticles. This method relies on a nanocomposite film that is prepared by homogeneously dispersing nanoparticles within a biodegradable polymeric film. When adherent cells are seeded on the film, they adhere, spread, and filtrate into the film through the micropores formed during the film fabrication. The pre-embedded nanoparticles are thus internalized by the cells during this infiltration process. As an example, fluorescent silica nanoparticles were homogeneously distributed within a polycaprolactone film by utilizing cryomilling and heat pressing. Upon incubation within physiological buffer, no silica nanoparticles were released from the nanocomposite film even after 20 d of incubation. However, when adherent cells (e.g., human mesenchymal stem cells) were grown on the film, they became fluorescent after 3 d, which suggests internalization of silica nanoparticles by cells. In comparison, the suspension cells (e.g., monocytes) in the medium remained nonfluorescent no matter whether there was the presence of adherent cells or not. This strategy eventually allowed the selective and concomitant labeling of mesenchymal stem cells during their harvest from bone marrow aspiration

  1. Patterned Thermoresponsive Microgel Coatings for Noninvasive Processing of Adherent Cells.

    PubMed

    Uhlig, Katja; Wegener, Thomas; He, Jian; Zeiser, Michael; Bookhold, Johannes; Dewald, Inna; Godino, Neus; Jaeger, Magnus; Hellweg, Thomas; Fery, Andreas; Duschl, Claus

    2016-03-14

    Cultivation of adherently growing cells in artificial environments is of utmost importance in medicine and biotechnology to accomplish in vitro drug screening or to investigate disease mechanisms. Precise cell manipulation, like localized control over adhesion, is required to expand cells, to establish cell models for novel therapies and to perform noninvasive cell experiments. To this end, we developed a method of gentle, local lift-off of mammalian cells using polymer surfaces, which are reversibly and repeatedly switchable between a cell-attractive and a cell-repellent state. This property was introduced through micropatterned thermoresponsive polymer coatings formed from colloidal microgels. Patterning was obtained through automated nanodispensing or microcontact printing, making use of unspecific electrostatic interactions between microgels and substrates. This process is much more robust against ambient conditions than covalent coupling, thus lending itself to up-scaling. As an example, wound healing assays were accomplished at 37 °C with highly increased precision in microfluidic environments. PMID:26879608

  2. Adherence in adolescents with Type 1 diabetes: strategies and considerations for assessment in research and practice

    PubMed Central

    Gandhi, Kajal; Vu, Bach-Mai K; Eshtehardi, Sahar S; Wasserman, Rachel M; Hilliard, Marisa E

    2015-01-01

    Suboptimal adherence remains a significant concern for adolescents with Type 1 diabetes, the treatment regimen for which is complex and includes numerous behaviors. Accurate assessment of adherence is critical for effective healthcare and to measure trial outcomes. Without a valid biomarker of adherence, assessment strategies must rely on measuring management behaviors. This paper provides an overview of approaches to measure adherence, with an emphasis on contemporary, validated measures that are appropriate for current diabetes care. Objective measures include electronic data from diabetes management devices. Subjective measures include self/parent-report questionnaires, structured interviews and diaries/logbooks. Practical strategies for selecting measurement approaches for clinical and research purposes are reviewed, and implications of adherence assessment for clinical care delivery and adherence-promotion are discussed. PMID:27066110

  3. Non-adherence in type 2 diabetes: practical considerations for interpreting the literature

    PubMed Central

    Blackburn, David F; Swidrovich, Jaris; Lemstra, Mark

    2013-01-01

    The rising prevalence of type 2 diabetes poses a serious threat to human health and the viability of many health care systems around the world. Although several prescription medications can play a vital role in controlling symptoms and preventing complications, non-adherence to these therapies is highly prevalent and has been linked to increases in morbidity, mortality, and health care costs. Although a vast array of significant adherence predictors has been identified, the ability to explain or predict non-adherence with known risk-factors remains poor. Further, the definitions, outcomes, and various measures used in the non-adherence literature can be misleading for the unfamiliar reviewer. In this narrative review, a practical overview of important considerations for interpreting adherence endpoints and measures is discussed. Also, an organizational framework is proposed to consider published adherence interventions. This framework may allow for a unique appreciation into areas of limited knowledge and thus highlights targets for future research. PMID:23487395

  4. Association between patients’ beliefs and oral antidiabetic medication adherence in a Chinese type 2 diabetic population

    PubMed Central

    Wu, Ping; Liu, Naifeng

    2016-01-01

    Purpose The objective of this study was to identify, using the theory of planned behavior (TPB), patients’ beliefs about taking oral antidiabetic drugs (OADs) as prescribed, and to measure the correlations between beliefs and medication adherence. Patients and methods We performed a cross-sectional study of type 2 diabetic patients using structured questionnaires in a Chinese tertiary hospital. A total of 130 patients were enrolled to be interviewed about TPB variables (behavioral, normative, and control beliefs) relevant to medication adherence. Medication adherence was assessed using the eight-item Morisky Medication Adherence Scale (MMAS-8). Spearman’s rank correlation was used to assess the association between TPB and MMAS-8. Logistic regression analysis was performed to examine the relationship between different variables and MMAS-8, with statistical significance determined at P<0.05. Results From 130 eligible Chinese patients with an average age of 60.6 years and a male proportion of 50.8%, a nonsignificant relationship between behavioral, normative, and the most facilitating control beliefs and OAD adherence was found in our study. Having the OADs on hand (P=0.037) was the only facilitating control belief associated with adherence behavior. Being away from home or eating out (P=0.000), not accepting the disease (P=0.000), ignorance of life-long drug adherence (P=0.038), being busy (P=0.001), or poor memory (P=0.008) were control belief barriers found to be correlated with poor adherence. TPB is the only important determinant influencing OAD adherence among all the factors (P=0.011). Conclusion The results indicate that the TPB model could be used to examine adherence to OADs. One facilitating control belief, and most of the barrier control beliefs of TPB were related to medication adherence among Chinese type 2 diabetes inpatients. It will be helpful to understand patients’ self-medication and provide methods to develop instruments for identifying

  5. Automated and Online Characterization of Adherent Cell Culture Growth in a Microfabricated Bioreactor

    PubMed Central

    Jaccard, Nicolas; Macown, Rhys J.; Super, Alexandre; Griffin, Lewis D.; Veraitch, Farlan S.

    2014-01-01

    Adherent cell lines are widely used across all fields of biology, including drug discovery, toxicity studies, and regenerative medicine. However, adherent cell processes are often limited by a lack of advances in cell culture systems. While suspension culture processes benefit from decades of development of instrumented bioreactors, adherent cultures are typically performed in static, noninstrumented flasks and well-plates. We previously described a microfabricated bioreactor that enables a high degree of control on the microenvironment of the cells while remaining compatible with standard cell culture protocols. In this report, we describe its integration with automated image-processing capabilities, allowing the continuous monitoring of key cell culture characteristics. A machine learning–based algorithm enabled the specific detection of one cell type within a co-culture setting, such as human embryonic stem cells against the background of fibroblast cells. In addition, the algorithm did not confuse image artifacts resulting from microfabrication, such as scratches on surfaces, or dust particles, with cellular features. We demonstrate how the automation of flow control, environmental control, and image acquisition can be employed to image the whole culture area and obtain time-course data of mouse embryonic stem cell cultures, for example, for confluency. PMID:24692228

  6. Dynamic mechanical measurement of the viscoelasticity of single adherent cells

    NASA Astrophysics Data System (ADS)

    Corbin, Elise A.; Adeniba, Olaoluwa O.; Ewoldt, Randy H.; Bashir, Rashid

    2016-02-01

    Many recent studies on the viscoelasticity of individual cells link mechanics with cellular function and health. Here, we introduce a measurement of the viscoelastic properties of individual human colon cancer cells (HT-29) using silicon pedestal microelectromechanical systems (MEMS) resonant sensors. We demonstrate that the viscoelastic properties of single adherent cells can be extracted by measuring a difference in vibrational amplitude of our resonant sensor platform. The magnitude of vibration of the pedestal sensor is measured using a laser Doppler vibrometer (LDV). A change in amplitude of the sensor, compared with the driving amplitude (amplitude ratio), is influenced by the mechanical properties of the adhered cells. The amplitude ratio of the fixed cells was greater than the live cells, with a p-value <0.0001. By combining the amplitude shift with the resonant frequency shift measure, we determined the elastic modulus and viscosity values of 100 Pa and 0.0031 Pa s, respectively. Our method using the change in amplitude of resonant MEMS devices can enable the determination of a refined solution space and could improve measuring the stiffness of cells.

  7. A Localized Adherence-Like Pattern as a Second Pattern of Adherence of Classic Enteropathogenic Escherichia coli to HEp-2 Cells That Is Associated with Infantile Diarrhea

    PubMed Central

    Scaletsky, Isabel C. A.; Pedroso, Margareth Z.; Oliva, Carlos A. G.; Carvalho, Rozane L. B.; Morais, Mauro B.; Fagundes-Neto, Ulysses

    1999-01-01

    Escherichia coli strains that cause nonbloody diarrhea in infants are known to present three distinct patterns of adherence to epithelial cells, namely, localized (LA), diffuse (DA), and aggregative (AA) adherence. Strains with LA (typical Enteropathogenic Escherichia coli [EPEC]) are well recognized as a cause of secretory diarrhea, but the role of strains with DA (DAEC) is controversial, and strains with AA (EAEC) have been more frequently related to persistent diarrhea whereas its relationship with acute diarrhea is not well defined. To determine the relationship of the different types of E. coli adherence patterns with acute diarrhea (lasting less than 14 days) and persistent diarrhea (lasting more than 14 days) in São Paulo, Brazil, we studied stool specimens from 40 infants under 1 year of age with diarrhea and 40 age-matched control infants without any gastrointestinal symptoms. Twenty-eight (35.0%) of eighty cases yielded adherent E. coli (HEp-2 cells). Strains with localized and aggregative adherence were associated with acute and persistent diarrhea. A total of 11.2% of the adherent strains were typical EPEC serotypes and hybridized with the enteroadherence factor probe; 5.0% were EAEC and hybridized with the EAEC probe. DAEC strains were isolated from 10.0% of patients and 7.5% of controls and did not hybridize with the two probes used (daaC and AIDA-I). Strains with a localized adherence-like pattern (atypical EPEC) were found significantly more frequently (P = 0.028) in cultures from children with diarrhea (17.5%) than in controls (2.5%). PMID:10377120

  8. A Prestressed Cable Network Model of the Adherent Cell Cytoskeleton

    PubMed Central

    Coughlin, Mark F.; Stamenović, Dimitrije

    2003-01-01

    A prestressed cable network is used to model the deformability of the adherent cell actin cytoskeleton. The overall and microstructural model geometries and cable mechanical properties were assigned values based on observations from living cells and mechanical measurements on isolated actin filaments, respectively. The models were deformed to mimic cell poking (CP), magnetic twisting cytometry (MTC) and magnetic bead microrheometry (MBM) measurements on living adherent cells. The models qualitatively and quantitatively captured the fibroblast cell response to the deformation imposed by CP while exhibiting only some qualitative features of the cell response to MTC and MBM. The model for CP revealed that the tensed peripheral actin filaments provide the key resistance to indentation. The actin filament tension that provides mechanical integrity to the network was estimated at ∼158 pN, and the nonlinear mechanical response during CP originates from filament kinematics. The MTC and MBM simulations revealed that the model is incomplete, however, these simulations show cable tension as a key determinant of the model response. PMID:12547813

  9. Evidence that extracellular components function in adherence of Actinobacillus actinomycetemcomitans to epithelial cells.

    PubMed Central

    Meyer, D H; Fives-Taylor, P M

    1993-01-01

    Extracellular microvesicles and a highly proteinaceous polymer associated with a leukotoxin-producing strain, Actinobacillus actinomycetemcomitans SUNY 75, were shown to increase adherence of other weakly adherent A. actinomycetemcomitans strains to KB epithelial cells. Images PMID:8406899

  10. Endometrial Mesenchymal Stem Cells Isolated from Menstrual Blood by Adherence

    PubMed Central

    Du, Xue; Yuan, Qing; Qu, Ye; Zhou, Yuan; Bei, Jia

    2016-01-01

    Objective. To find a convenient and efficient way to isolate MSCs from human menstrual blood and to investigate their biological characteristics, proliferative capacity, and secretion levels. Methods. MSCs were isolated from menstrual blood of 3 healthy women using adherence. Cell immunological phenotype was examined by flow cytometry; the adipogenic, osteogenic, and chondrogenic differentiation of MSCs was examined by Oil-Red-O staining, ALP staining, and Alcian Blue staining, respectively; and the secretion of cytokines, including vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), and insulin-like growth factor-1 (IGF-1), was detected using enzyme-linked immunosorbent assay. Results. MB-MSCs were successfully isolated from human menstrual blood using adherence. They were positive for CD73, CD105, CD29, and CD44, but negative for CD31 and CD45. The differentiated MB-MSCs were positive for ALP staining, Oil-Red-O staining, and Alcian Blue staining. In addition, they could secrete antiapoptotic cytokines, such as VEGF, IGF-1, and HGF. Conclusion. It is feasible to isolate MSCs from human menstrual blood, thus avoiding invasive procedures and ethical controversies. Adherence could be a promising alternative to the density gradient centrifugation for the isolation of MSCs from menstrual blood. PMID:26681948

  11. A fully automated system for adherent cells microinjection.

    PubMed

    Becattini, Gabriele; Mattos, Leonardo S; Caldwell, Darwin G

    2014-01-01

    This paper proposes an automated robotic system to perform cell microinjections to relieve human operators from this highly difficult and tedious manual procedure. The system, which uses commercial equipment currently found on most biomanipulation laboratories, consists of a multitask software framework combining computer vision and robotic control elements. The vision part features an injection pipette tracker and an automatic cell targeting system that is responsible for defining injection points within the contours of adherent cells in culture. The main challenge is the use of bright-field microscopy only, without the need for chemical markers normally employed to highlight the cells. Here, cells are identified and segmented using a threshold-based image processing technique working on defocused images. Fast and precise microinjection pipette positioning over the automatically defined targets is performed by a two-stage robotic system which achieves an average injection rate of 7.6 cells/min with a pipette positioning precision of 0.23 μm. The consistency of these microinjections and the performance of the visual targeting framework were experimentally evaluated using two cell lines (CHO-K1 and HEK) and over 500 cells. In these trials, the cells were automatically targeted and injected with a fluorescent marker, resulting in a correct cell detection rate of 87% and a successful marker delivery rate of 67.5%. These results demonstrate that the new system is capable of better performances than expert operators, highlighting its benefits and potential for large-scale application. PMID:24403406

  12. Computational Tension Mapping of Adherent Cells Based on Actin Imaging

    PubMed Central

    Manifacier, Ian; Milan, Jean-Louis; Jeanneau, Charlotte; Chmilewsky, Fanny; Chabrand, Patrick; About, Imad

    2016-01-01

    Forces transiting through the cytoskeleton are known to play a role in adherent cell activity. Up to now few approaches haves been able to determine theses intracellular forces. We thus developed a computational mechanical model based on a reconstruction of the cytoskeleton of an adherent cell from fluorescence staining of the actin network and focal adhesions (FA). Our custom made algorithm converted the 2D image of an actin network into a map of contractile interactions inside a 2D node grid, each node representing a group of pixels. We assumed that actin filaments observed under fluorescence microscopy, appear brighter when thicker, we thus presumed that nodes corresponding to pixels with higher actin density were linked by stiffer interactions. This enabled us to create a system of heterogeneous interactions which represent the spatial organization of the contractile actin network. The contractility of this interaction system was then adapted to match the level of force the cell truly exerted on focal adhesions; forces on focal adhesions were estimated from their vinculin expressed size. This enabled the model to compute consistent mechanical forces transiting throughout the cell. After computation, we applied a graphical approach on the original actin image, which enabled us to calculate tension forces throughout the cell, or in a particular region or even in single stress fibers. It also enabled us to study different scenarios which may indicate the mechanical role of other cytoskeletal components such as microtubules. For instance, our results stated that the ratio between intra and extra cellular compression is inversely proportional to intracellular tension. PMID:26812601

  13. Computational Tension Mapping of Adherent Cells Based on Actin Imaging.

    PubMed

    Manifacier, Ian; Milan, Jean-Louis; Jeanneau, Charlotte; Chmilewsky, Fanny; Chabrand, Patrick; About, Imad

    2016-01-01

    Forces transiting through the cytoskeleton are known to play a role in adherent cell activity. Up to now few approaches haves been able to determine theses intracellular forces. We thus developed a computational mechanical model based on a reconstruction of the cytoskeleton of an adherent cell from fluorescence staining of the actin network and focal adhesions (FA). Our custom made algorithm converted the 2D image of an actin network into a map of contractile interactions inside a 2D node grid, each node representing a group of pixels. We assumed that actin filaments observed under fluorescence microscopy, appear brighter when thicker, we thus presumed that nodes corresponding to pixels with higher actin density were linked by stiffer interactions. This enabled us to create a system of heterogeneous interactions which represent the spatial organization of the contractile actin network. The contractility of this interaction system was then adapted to match the level of force the cell truly exerted on focal adhesions; forces on focal adhesions were estimated from their vinculin expressed size. This enabled the model to compute consistent mechanical forces transiting throughout the cell. After computation, we applied a graphical approach on the original actin image, which enabled us to calculate tension forces throughout the cell, or in a particular region or even in single stress fibers. It also enabled us to study different scenarios which may indicate the mechanical role of other cytoskeletal components such as microtubules. For instance, our results stated that the ratio between intra and extra cellular compression is inversely proportional to intracellular tension. PMID:26812601

  14. Characteristics and response of mouse bone marrow derived novel low adherent mesenchymal stem cells acquired by quantification of extracellular matrix

    PubMed Central

    Zheng, Ri-Cheng; Heo, Seong-Joo; Koak, Jai-Young; Lee, Joo-Hee; Park, Ji-Man

    2014-01-01

    PURPOSE The aim of present study was to identify characteristic and response of mouse bone marrow (BM) derived low-adherent bone marrow mesenchymal stem cells (BMMSCs) obtained by quantification of extracellular matrix (ECM). MATERIALS AND METHODS Non-adherent cells acquired by ECM coated dishes were termed low-adherent BMMSCs and these cells were analyzed by in vitro and in vivo methods, including colony forming unit fibroblast (CFU-f), bromodeoxyuridine (BrdU), multi-potential differentiation, flow cytometry and transplantation into nude mouse to measure the bone formation ability of these low-adherent BMMSCs. Titanium (Ti) discs with machined and anodized surfaces were prepared. Adherent and low-adherent BMMSCs were cultured on the Ti discs for testing their proliferation. RESULTS The amount of CFU-f cells was significantly higher when non-adherent cells were cultured on ECM coated dishes, which was made by 7 days culturing of adherent BMMSCs. Low-adherent BMMSCs had proliferation and differentiation potential as adherent BMMSCs in vitro. The mean amount bone formation of adherent and low-adherent BMMSCs was also investigated in vivo. There was higher cell proliferation appearance in adherent and low-adherent BMMSCs seeded on anodized Ti discs than machined Ti discs by time. CONCLUSION Low-adherent BMMSCs acquired by ECM from non-adherent cell populations maintained potential characteristic similar to those of the adherent BMMSCs and therefore could be used effectively as adherent BMMSCs in clinic. PMID:25352957

  15. Systematic review of adherence rates by medication class in type 2 diabetes: a study protocol

    PubMed Central

    McGovern, Andrew; Tippu, Zayd; Hinton, William; Munro, Neil; Whyte, Martin; de Lusignan, Simon

    2016-01-01

    Introduction Treatment options for type 2 diabetes are becoming increasingly complex with people often prescribed multiple medications, and may include both oral and injectable therapies. There is ongoing debate about which drug classes provide the optimum second-line and third-line treatment options. In the real world, patient adherence and persistence determines medication effectiveness. A better understanding of adherence may help inform the choice of second-line and third-line drug classes. Methods and analysis This systematic review will compare adherence and persistence rates across the different classes of medication available to people with type 2 diabetes. It will include all identified studies comparing medication adherence or persistence between two or more glucose-lowering medications in people with type 2 diabetes. Research databases (MEDLINE, EMBASE, The Cochrane Library, The Register of Controlled Trials, PsychINFO and CINAHL) will be searched for relevant articles, using a comprehensive search strategy. All identified medication trials and observational studies will be included which compare adherence or persistence across classes of diabetes medication. The characteristics and outcomes of all the included studies will be reported along with a study quality grade, assessed using the Cochrane Risk Assessment Tool. The quality of adjustment for confounders of adherence or persistence will be reported for each study. Where multiple (n ≥3) studies provide compare adherence or persistence across the same 2 medication classes, a meta-analysis will be performed. Ethics and dissemination No ethics approval is required. This review and meta-analysis (where possible) will provide important information on the relative patient adherence and persistence, with the different classes of diabetes therapies. Once complete, the results will be made available by peer-reviewed publication. Trial registration number CRD42015027865. PMID:26928029

  16. Regulatory focus and adherence to self-care behaviors among adults with type 2 diabetes.

    PubMed

    Avraham, Rinat; Van Dijk, Dina; Simon-Tuval, Tzahit

    2016-09-01

    The aims of this study were, first, to test the association between regulatory focus of adults with type 2 diabetes and their adherence to two types of self-care behaviors - lifestyle change (e.g. physical activity and diet) and medical care regimens (blood-glucose monitoring, foot care and medication usage). Second, to explore whether a fit between the message framing and patients' regulatory focus would improve their intentions to adhere specifically when the type of behavior fits the patients' regulatory focus as well. A cross-sectional study was conducted among 130 adults with type 2 diabetes who were hospitalized in an academic medical center. The patients completed a set of questionnaires that included their diabetes self-care activities, regulatory focus, self-esteem and demographic, socioeconomic and clinical data. In addition, participants were exposed to either a gain-framed or a loss-framed message, and were then asked to indicate their intention to improve adherence to self-care behaviors. A multivariable linear regression model revealed that promoters reported higher adherence to lifestyle change behaviors than preventers did (B = .60, p = .028). However, no effect of regulatory focus on adherence to medical care regimens was found (B = .46, p = .114). In addition, preventers reported higher intentions to adhere to medical care behaviors when the message framing was congruent with prevention focus (B = 1.16, p = .023). However, promoters did not report higher intentions to adhere to lifestyle behaviors when the message framing was congruent with promotion focus (B = -.16, p = .765). These findings justify the need to develop tailor-made interventions that are adjusted to both patients' regulatory focus and type of health behavior. PMID:26576471

  17. Motivational Interviewing to Promote Adherence Behaviors in Pediatric Type 1 Diabetes

    PubMed Central

    Hilliard, Marisa E.; Anderson, Barbara J.

    2015-01-01

    Suboptimal regimen adherence among youth with type 1 diabetes (T1D) is a common challenge for patients, families, and providers. Motivational interviewing (MI) is a brief communication style designed to elicit intrinsic motivation and strengthen commitment to behavior change goals. As pediatric MI research expands, a critical review of its evidence base and applicability to promote adherence behaviors for youth with T1D is needed. This review introduces the core tenets of MI and clinical applications in T1D, synthesizes the existing MI research in T1D, and discusses the next steps in MI research. Overall, mixed results for MI interventions in T1D reflect variations in research study design and clinical implementation. Targeting adherence rather than glycemic outcomes typically demonstrates greater results, highlighting the promise of MI to facilitate meaningful and enduring improvements in youths’ T1D adherence behaviors. PMID:25142716

  18. Evaluation of a Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) assay (Keystone Sym)

    EPA Science Inventory

    Our goal is to establish an in vitro model system to evaluate chemical effects using a single stem cell culture technique that would improve throughput and provide quantitative markers of differentiation and cell number. To this end, we have used an adherent cell differentiation ...

  19. Factors involved in adherence of lactobacilli to human Caco-2 cells.

    PubMed Central

    Greene, J D; Klaenhammer, T R

    1994-01-01

    A quantitative assay performed with bacterial cells labelled with [3H]thymidine was used to investigate factors involved in the adherence of human isolates Lactobacillus acidophilus BG2FO4 and NCFM/N2 and Lactobacillus gasseri ADH to human Caco-2 intestinal cells. For all three strains, adherence was concentration dependent, greater at acidic pH values, and significantly greater than adherence of a control dairy isolate, Lactobacillus delbrueckii subsp. bulgaricus 1489. Adherence of L. acidophilus BG2FO4 and NCFM/N2 was decreased by protease treatment of the bacterial cells, whereas adherence of L. gasseri ADH either was not affected or was enhanced by protease treatment. Putative surface layer proteins were identified on L. acidophilus BG2FO4 and NCFM/N2 cells but were not involved in adherence. Periodate oxidation of bacterial cell surface carbohydrates significantly reduced adherence of L. gasseri ADH, moderately reduced adherence of L. acidophilus BG2FO4, and had no effect on adherence of L. acidophilus NCFM/N2. These results indicate that Lactobacillus species adhere to human intestinal cells via mechanisms which involve different combinations of carbohydrate and protein factors on the bacterial cell surface. The involvement of a secreted bridging protein, which has been proposed as the primary mediator of adherence of L. acidophilus BG2FO4 in spent culture supernatant (M.-H. Coconnier, T. R. Klaenhammer, S. Kernéis, M.-F. Bernet, and A. L. Servin, Appl. Environ. Microbiol. 58:2034-2039, 1992), was not confirmed in this study. Rather, a pH effect on Caco-2 cells contributed significantly to the adherence of this strain in spent culture supernatant.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:7811085

  20. Adherence to statin therapy in patients with type 2 diabetes: An important dilemma

    PubMed Central

    Farsaei, Shadi; Sabzghabaee, Ali Mohammad; Amini, Masoud; Zargarzadeh, Amir Hooshang

    2015-01-01

    Background: Despite the importance of patients’ adherence to their drug treatments for achieving desired therapeutic goals and the proven role 3-hydroxy-3-methylglutaryl coenzyme A inhibitors (statins) for the health status of patients with cardiovascular diseases, there is not enough information regarding diabetic patients’ adherence to statin therapy in developing countries. In this clinical study we aimed to assess the adherence of diabetes type 2 patients to statin therapy in a research based community clinic in Iran. Materials and Methods: In this prospective clinical study which was done at Isfahan Endocrinology and Metabolism Research Center, 204 diabetic type 2 patients under treatment with statin were interviewed twice and their demographic data (age, gender, body mass index, education), statin information (type, dose) and their serum lipid profile were recorded. Three months after the initial visits, patients were assessed using pill counting method and according to patients’ self-reporting and also assessed low-density lipoprotein (LDL) cholesterol goal attainment <100 mg/dl. Results: Adherence rate was 79.7% and 69% according to pill counting and self-reporting among study population. Moreover, 68.4% of patients achieved their LDL cholesterol goal of <100 mg/dl and adherent patients reached therapeutic goal significantly more than those who were considered non-adherence to statin therapy (P < 0.01). Conclusion: Adherence to statin therapy, as reflected by pill count method, is significantly related to LDL cholesterol goal achievement in patients with diabetes and dyslipidemia. Pill count method can be used to identify patients who are nonadherent to statin therapy and at high risk for failure to attain LDL cholesterol goals. PMID:25983760

  1. Patterns and obstacles to oral antidiabetic medications adherence among type 2 diabetics in Ismailia, Egypt: a cross section study

    PubMed Central

    Heissam, Khaled; Abuamer, Zeinab; El-Dahshan, Nahed

    2015-01-01

    Introduction Diabetes is a costly and increasingly common chronic disease. Effective management of diabetes to achieve glycemic control improves patient quality of life. Adherence rates to drug regimens in patients with type 2 diabetes are relatively low and vary widely between populations. There are many factors that could affect patient adherence to drug therapy. The aim of the present study was assessing patterns and obstacles to adherence of type 2 diabetic patients to their oral hypoglycemic drugs. Methods The present work is a descriptive cross section study, carried on type 2 diabetic patients who were on oral hypoglycemic drugs. Data concerning adherence to drugs was assessed using measure treatment adherence scale (MTA). Results A total of 372 (55.59% males and 44.41% females) patients with type-2 diabetes fulfilled the inclusion criteria and included in the study. Among the participants, 26.1% were found to have good adherence, 47.9% had a fair adherence, and 26% had poor adherence. Conclusion The overall rate of medication adherence among the diabetic patients population was suboptimal and non-acceptable. Evaluation of adherence is vital for patients with diabetes in order to determine factors and barriers affecting the adherence and to manage them. PMID:26113919

  2. Type A Behaviors, Hostility, Anger and Exercise Adherence.

    ERIC Educational Resources Information Center

    Goffaux, Jacqueline; And Others

    A study was conducted to examine the relationship between the components of the Type A behavior pattern and the maintenance of exercise participation in a 5-month physical fitness program. Metropolitan Government employees (N=200) volunteered to participate in a pilot health promotion program. Physical fitness activities (supervised walking,…

  3. Adherence of Candida to cultured vascular endothelial cells: mechanisms of attachment and endothelial cell penetration.

    PubMed

    Rotrosen, D; Edwards, J E; Gibson, T R; Moore, J C; Cohen, A H; Green, I

    1985-12-01

    To elucidate the pathogenesis of hematogenous Candida infections, we developed an in vitro model of Candida adherence to and penetration of human endothelial cells. We enhanced or inhibited adherence in order to probe mechanisms of attachment. Adherence of Candida albicans showed a linear relation to Candida inoculum (range, 10(2)-10(5) cfu, r = .99, P less than .01) and exceeded that of less virulent Candida species and that of Saccharomyces cerevisiae (P less than .01). Candida immune serum blocked attachment (greater than 95% inhibition; P less than .001), however, this activity was abolished by immunoprecipitation of immune serum with C. albicans mannan (P less than .001) and was unaffected by immunoprecipitation with S. cerevisiae mannan or by adsorption with particulate chitin. Adherence was diminished by exposing C. albicans to heat (greater than 99% inhibition; P less than .01), UV light (98% inhibition; P less than .01), or sodium periodate (greater than 72% inhibition; P less than .01). An extract from heat-exposed C. albicans blocked adherence (greater than 51% inhibition; P less than .001). Transmission electron microscopy demonstrated that viable or killed Candida organisms were attached to endothelial cells, were enveloped by membrane processes from the endothelial cell surface, and were incorporated into the endothelial cells within phagosomes. Cytochalasin B blocked incorporation without blocking surface attachment. PMID:3905987

  4. Muscle-derived stem cells isolated as non-adherent population give rise to cardiac, skeletal muscle and neural lineages

    SciTech Connect

    Arsic, Nikola; Mamaeva, Daria; Lamb, Ned J.; Fernandez, Anne

    2008-04-01

    Stem cells with the ability to differentiate in specialized cell types can be extracted from a wide array of adult tissues including skeletal muscle. Here we have analyzed a population of cells isolated from skeletal muscle on the basis of their poor adherence on uncoated or collagen-coated dishes that show multi-lineage differentiation in vitro. When analysed under proliferative conditions, these cells express stem cell surface markers Sca-1 (65%) and Bcrp-1 (80%) but also MyoD (15%), Neuronal {beta} III-tubulin (25%), GFAP (30%) or Nkx2.5 (1%). Although capable of growing as non-attached spheres for months, when given an appropriate matrix, these cells adhere giving rise to skeletal muscle, neuronal and cardiac muscle cell lineages. A similar cell population could not be isolated from either bone marrow or cardiac tissue suggesting their specificity to skeletal muscle. When injected into damaged muscle, these non-adherent muscle-derived cells are retrieved expressing Pax7, in a sublaminar position characterizing satellite cells and participate in forming new myofibers. These data show that a non-adherent stem cell population can be specifically isolated and expanded from skeletal muscle and upon attachment to a matrix spontaneously differentiate into muscle, cardiac and neuronal lineages in vitro. Although competing with resident satellite cells, these cells are shown to significantly contribute to repair of injured muscle in vivo supporting that a similar muscle-derived non-adherent cell population from human muscle may be useful in treatment of neuromuscular disorders.

  5. A validated measure of adherence to antibiotic prophylaxis in children with sickle cell disease

    PubMed Central

    Duncan, Natalie A; Kronenberger, William G; Hampton, Kisha C; Bloom, Ellen M; Rampersad, Angeli G; Roberson, Christopher P; Shapiro, Amy D

    2016-01-01

    Background Antibiotic prophylaxis is a mainstay in sickle cell disease management. However, adherence is estimated at only 66%. This study aimed to develop and validate a Sickle Cell Antibiotic Adherence Level Evaluation (SCAALE) to promote systematic and detailed adherence evaluation. Methods A 28-item questionnaire was created, covering seven adherence areas. General Adherence Ratings from the parent and one health care provider and medication possession ratios were obtained as validation measures. Results Internal consistency was very good to excellent for the total SCAALE (α=0.89) and four of the seven subscales. Correlations between SCAALE scores and validation measures were strong for the total SCAALE and five of the seven subscales. Conclusion The SCAALE provides a detailed, quantitative, multidimensional, and global measurement of adherence and can promote clinical care and research. PMID:27354768

  6. Toxicity Minimized Cryoprotectant Addition and Removal Procedures for Adherent Endothelial Cells

    PubMed Central

    Davidson, Allyson Fry; Glasscock, Cameron; McClanahan, Danielle R.; Benson, James D.; Higgins, Adam Z.

    2015-01-01

    Ice-free cryopreservation, known as vitrification, is an appealing approach for banking of adherent cells and tissues because it prevents dissociation and morphological damage that may result from ice crystal formation. However, current vitrification methods are often limited by the cytotoxicity of the concentrated cryoprotective agent (CPA) solutions that are required to suppress ice formation. Recently, we described a mathematical strategy for identifying minimally toxic CPA equilibration procedures based on the minimization of a toxicity cost function. Here we provide direct experimental support for the feasibility of these methods when applied to adherent endothelial cells. We first developed a concentration- and temperature-dependent toxicity cost function by exposing the cells to a range of glycerol concentrations at 21°C and 37°C, and fitting the resulting viability data to a first order cell death model. This cost function was then numerically minimized in our state constrained optimization routine to determine addition and removal procedures for 17 molal (mol/kg water) glycerol solutions. Using these predicted optimal procedures, we obtained 81% recovery after exposure to vitrification solutions, as well as successful vitrification with the relatively slow cooling and warming rates of 50°C/min and 130°C/min. In comparison, conventional multistep CPA equilibration procedures resulted in much lower cell yields of about 10%. Our results demonstrate the potential for rational design of minimally toxic vitrification procedures and pave the way for extension of our optimization approach to other adherent cell types as well as more complex systems such as tissues and organs. PMID:26605546

  7. Raman micro-spectroscopy study of living SH-SY5Y cells adhering on different substrates.

    PubMed

    Caponi, S; Mattana, S; Ricci, M; Sagini, K; Urbanelli, L; Sassi, P; Morresi, A; Emiliani, C; Dalla Serra, M; Iannotta, S; Musio, C; Fioretto, D

    2016-01-01

    In this paper we test the ability of Raman micro-spectroscopy and Raman mapping to investigate the status of cells grown in adhesion on different substrates. The spectra of immortalized SH-SY5Y cells, grown on silicon and on metallic substrates are compared with those obtained for the same type of cells adhering on organic polyaniline (PANI), a memristive substrate chosen to achieve a living bio-hybrid system. Raman spectra give information on the status of the single cell, its local biochemical composition, and on the modifications induced by the substrate interaction. The good agreement between Raman spectra collected from cells adhering on different substrates confirms that the PANI, besides allowing the cell growth, doesn't strongly affect the general biochemical properties of the cell. The investigation of the cellular state in a label free condition is challenging and the obtained results confirm the Raman ability to achieve this information. PMID:26256426

  8. A rapid and sensitive fluorometric microassay for determining cell mediated cytotoxicity to adherent growing cell lines.

    PubMed

    Krüger-Krasagakes, S; Garbe, C; Kossman, P; Orfanos, C E

    1992-11-25

    In order to measure cell mediated cytotoxicity to adherent growing cell lines in vitro more rapidly and conveniently, a fluorometric microassay was developed and results were compared with those obtained by the 51Cr release assay. The fluorometric method is based on the hydrolysis of the fluorochrome 4-methylumbelliferyl heptanoate (MUH) by intracellular esterases of viable cells. Melanoma cell monolayers were incubated with lymphokine activated killer (LAK) cells for 4 h at various effector: target (E:T) cell ratios (E:T = 16, 8, 4, 2:1). Thereafter surviving adherent melanoma cells were stained with MUH for 30 min and fluorescence was measured directly in a 96 well plate reader. For the calculation of LAK cell cytotoxicity fluorescence values were corrected for the number of nonspecifically detached tumor cells during the washes and the number of nonspecifically adherent LAK cells. Using identical target and effector cell preparations both assays showed a nearly proportional increase of percentage cytotoxicity with rising numbers of lymphocytes. Compared with the 51Cr release assay, however, higher cytotoxicity values were obtained with the fluorometric MUH microassay: 57% with MUH versus 26% with 51Cr and 39% versus 14% for cell lines StML-11 and SKMel-28, respectively (E:T ratio = 16:1). The higher cytotoxicity rates obtained with the fluorometric MUH microassay were not due to the additional 30 min staining with MUH or due to nonspecific hydrolysis of MUH by extracellular esterases released from damaged cells, as could be shown by a series of experiments. In conclusion, a simple and rapid fluorometric microassay has been developed showing reliable reproducibility and a higher sensitivity compared with the 51Cr release assay for the determination of cellular cytotoxicity to adherent growing cell lines, avoiding hazardous radioactive labels. PMID:1431156

  9. The role of alginate in Pseudomonas aeruginosa EPS adherence, viscoelastic properties and cell attachment.

    PubMed

    Orgad, Oded; Oren, Yoram; Walker, Sharon L; Herzberg, Moshe

    2011-08-01

    Among various functions, extracellular polymeric substances (EPS) provide microbial biofilms with mechanical stability and affect initial cell attachment, the first stage in the biofilm formation process. The role of alginate, an abundant polysaccharide in Pseudomonas aeruginosa biofilms, in the viscoelastic properties and adhesion kinetics of EPS was analyzed using a quartz crystal microbalance with dissipation (QCM-D) monitoring technology. EPS was extracted from two P. aeruginosa biofilms, a wild type strain, PAO1, and a mucoid strain, PAOmucA22 that over-expresses alginate production. The higher alginate content in the EPS originating from the mucoid biofilms was clearly shown to increase both the rate and the extent of attachment of the EPS, as well as the layer's thickness. Also, the presence of calcium and elevated ionic strength increased the thickness of the EPS layer. Dynamic light scattering (DLS) showed that the presence of calcium and elevated ionic strength induced intermolecular attractive interactions in the mucoid EPS molecules. For the wild type EPS, in the presence of calcium, an elevated shift in the distribution of the diffusion coefficients was observed with DLS due to a more compacted conformation of the EPS molecules. Moreover, the alginate over-expression effect on EPS adherence was compared to the effect of alginate over-expression on P. aeruginosa cell attachment. In a parallel plate flow cell, under similar hydraulic and aquatic conditions as those applied for the EPS adsorption tests in the QCM-D flow cell, reduced adherence of the mucoid strain was clearly observed compared to the wild type isogenic bacteria. The results suggest that alginate contributes to steric hindrance and shielding of cell surface features and adhesins that are known to promote cell attachment. PMID:21797737

  10. Isolation and manipulation of living adherent cells by micromolded magnetic rafts

    PubMed Central

    Gach, Philip C.; Wang, Yuli; Phillips, Colleen; Sims, Christopher E.; Allbritton, Nancy L.

    2011-01-01

    A new strategy for magnetically manipulating and isolating adherent cells with extremely high post-collection purity and viability is reported. Micromolded magnetic elements (termed microrafts) were fabricated in an array format and used as culture surfaces and carriers for living, adherent cells. A poly(styrene-co-acrylic acid) polymer containing well dispersed magnetic nanoparticles was developed for creating the microstructures by molding. Nanoparticles of γFe2O3 at concentrations up to 1% wt.∕wt. could be used to fabricate microrafts that were optically transparent, highly magnetic, biocompatible, and minimally fluorescent. To prevent cellular uptake of nanoparticles from the magnetic polymer, a poly(styrene-co-acrylic acid) layer lacking γFe2O3 nanoparticles was placed over the initial magnetic microraft layer to prevent cellular uptake of the γFe2O3 during culture. The microraft surface geometry and physical properties were altered by varying the polymer concentration or layering different polymers during fabrication. Cells plated on the magnetic microrafts were visualized using standard imaging techniques including brightfield, epifluorescence, and confocal microscopy. Magnetic microrafts possessing cells of interest were dislodged from the array and efficiently collected with an external magnet. To demonstrate the feasibility of cell isolation using the magnetic microrafts, a mixed population of wild-type cells and cells stably transfected with a fluorescent protein was plated onto an array. Microrafts possessing single, fluorescent cells were released from the array and magnetically collected. A post-sorting single-cell cloning rate of 92% and a purity of 100% were attained. PMID:22007266

  11. Hepatitis B virus efficiently infects non-adherent hepatoma cells via human sodium taurocholate cotransporting polypeptide

    PubMed Central

    Okuyama-Dobashi, Kaori; Kasai, Hirotake; Tanaka, Tomohisa; Yamashita, Atsuya; Yasumoto, Jun; Chen, Wenjia; Okamoto, Toru; Maekawa, Shinya; Watashi, Koichi; Wakita, Takaji; Ryo, Akihide; Suzuki, Tetsuro; Matsuura, Yoshiharu; Enomoto, Nobuyuki; Moriishi, Kohji

    2015-01-01

    Sodium taurocholate cotransporting polypeptide (NTCP) has been reported as a functional receptor for hepatitis B virus (HBV) infection. However, HBV could not efficiently infect HepG2 cells expressing NTCP (NTCP-HepG2 cells) under adherent monolayer-cell conditions. In this study, NTCP was mainly detected in the basolateral membrane region, but not the apical site, of monolayer NTCP-HepG2 cells. We hypothesized that non-adherent cell conditions of infection would enhance HBV infectivity. Non-adherent NTCP-HepG2 cells were prepared by treatment with trypsin and EDTA, which did not degrade NTCP in the membrane fraction. HBV successfully infected NTCP-HepG2 cells at a viral dose 10 times lower in non-adherent phase than in adherent phase. Efficient infection of non-adherent NTCP-HepG2 cells with blood-borne or cell-culture-derived HBV was observed and was remarkably impaired in the presence of the myristoylated preS1 peptide. HBV could also efficiently infect HepaRG cells under non-adherent cell conditions. We screened several compounds using our culture system and identified proscillaridin A as a potent anti-HBV agent with an IC50 value of 7.2 nM. In conclusion, non-adherent host cell conditions of infection augmented HBV infectivity in an NTCP-dependent manner, thus providing a novel strategy to identify anti-HBV drugs and investigate the mechanism of HBV infection. PMID:26592202

  12. Comparison of adherence and persistence among adults with type 2 diabetes mellitus initiating saxagliptin or linagliptin

    PubMed Central

    Farr, Amanda M; Sheehan, John J; Davis, Brian M; Smith, David M

    2016-01-01

    Background Adherence and persistence to antidiabetes medications are important to control blood glucose levels among individuals with type 2 diabetes mellitus (T2D). Objectives The objective of this study was to compare adherence and persistence over a 12-month period between patients initiating saxagliptin and patients initiating linagliptin, two dipeptidyl peptidase-4 inhibitors. Methods This retrospective cohort study was conducted in MarketScan® Commercial and Medicare Supplemental claims databases. Patients with T2D initiating saxagliptin or linagliptin between January 1, 2009, and June 30, 2013, were selected. Patients were required to be at least 18 years old and have 12 months of continuous enrollment prior to and following initiation. Adherence and persistence to initiated medication were measured over the 12 months after initiation using outpatient pharmacy claims. Patients were considered adherent if the proportion of days covered was ≥0.80. Patients were considered nonpersistent (or to have discontinued) if there was a gap of >60 days without initiated medication on hand. Multivariable logistic regression and multivariable Cox proportional hazard models were fit to compare adherence and persistence, respectively, between the two cohorts. Results There were 21,599 saxagliptin initiators (mean age 55 years; 53% male) and 5,786 linagliptin initiators (mean age 57 years; 54% male) included in the study sample. Over the 12-month follow-up, 46% of saxagliptin initiators and 42% of linagliptin initiators were considered adherent and 47% of saxagliptin initiators and 51% of linagliptin initiators discontinued their initiated medication. After controlling for patient characteristics, saxagliptin initiation was associated with significantly greater odds of being adherent (adjusted odds ratio =1.212, 95% CI 1.140–1.289) and significantly lower hazards of discontinuation (adjusted hazard ratio =0.887, 95% CI 0.850–0.926) compared with linagliptin initiation

  13. Fluorescence assay for the detection of adherent Candida yeasts to target cells in microtest plates.

    PubMed

    Borg-von Zepelin, M; Wagner, T

    1995-01-01

    We describe an assay based on photometric analysis for the measurement of adherence of Candida species to epithelial target cells (Vero cell line). Adherent Candida cells were detected by staining the cells with the fluorescent dye Calcofluor white (CFW), which binds to chitin and glucan in the yeasts. The tests were performed on microtest plates, which were analysed automatically by fluorescence plate readers. The assay is based on the following steps: (i) coating of the microtest plates with target cells (e.g. Vero cells); (ii) infection with Candida: (iii) staining of Candida with CFW; (iv) rinsing to remove non-adherent Candida cells and unbound dye; (v) detection of adherent fluorescent Candida cells. The test was able to detect 4 x 10(4) cells ml-1. The standard deviation was +/- 8%. Day-to-day variation was +/- 10% at most. The adherence of strains of different Candida species was assayed by a standard procedure. The results confirmed the order of adherence, with C. albicans ranking first, followed by C. tropicalis, C. parapsilosis and C. glabrata. PMID:8569807

  14. Trichomonas vaginalis lipophosphoglycan mutants have reduced adherence and cytotoxicity to human ectocervical cells.

    PubMed

    Bastida-Corcuera, Felix D; Okumura, Cheryl Y; Colocoussi, Angie; Johnson, Patricia J

    2005-11-01

    The extracellular human pathogen Trichomonas vaginalis is covered by a dense glycocalyx thought to play a role in host-parasite interactions. The main component of the glycocalyx is lipophosphoglycan (LPG), a polysaccharide anchored in the plasma membrane by inositol phosphoceramide. To study the role of LPG in trichomonads, we produced T. vaginalis LPG mutants by chemical mutagenesis and lectin selection and characterized them using morphological, biochemical, and functional assays. Two independently selected LPG mutants, with growth rates comparable to that of the wild-type (parent) strain, lost the ability to bind the lectins Ricinnus comunis agglutinin I (RCA120) and wheat germ agglutinin, indicating alterations in surface galactose and glucosamine residues. LPG isolated from mutants migrated faster than parent strain LPG on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, suggesting the mutants had shorter LPG molecules. Dionex high-performance anion exchange chromatography with pulsed amperometric detection analyses revealed galactosamine, glucosamine, galactose, glucose, mannose/xylose, and rhamnose as the main monosaccharides of T. vaginalis parent strain LPG. LPG from both mutants showed a reduction of galactose and glucosamine, corresponding with the reduced size of their LPG molecules and inability to bind the lectins RCA120 and wheat germ agglutinin. Mutant parasites were defective in attachment to plastic, a characteristic associated with avirulent strains of T. vaginalis. Moreover, the mutants were less adherent and less cytotoxic to human vaginal ectocervical cells in vitro than the parental strain. Finally, while parent strain LPG could inhibit the attachment of parent strain parasites to vaginal cells, LPG from either mutant could not inhibit attachment. These combined results demonstrate that T. vaginalis adherence to host cells is LPG mediated and that an altered LPG leads to reduced adherence and cytotoxicity of this parasite. PMID

  15. Trichomonas vaginalis Lipophosphoglycan Mutants Have Reduced Adherence and Cytotoxicity to Human Ectocervical Cells

    PubMed Central

    Bastida-Corcuera, Felix D.; Okumura, Cheryl Y.; Colocoussi, Angie; Johnson, Patricia J.

    2005-01-01

    The extracellular human pathogen Trichomonas vaginalis is covered by a dense glycocalyx thought to play a role in host-parasite interactions. The main component of the glycocalyx is lipophosphoglycan (LPG), a polysaccharide anchored in the plasma membrane by inositol phosphoceramide. To study the role of LPG in trichomonads, we produced T. vaginalis LPG mutants by chemical mutagenesis and lectin selection and characterized them using morphological, biochemical, and functional assays. Two independently selected LPG mutants, with growth rates comparable to that of the wild-type (parent) strain, lost the ability to bind the lectins Ricinnus comunis agglutinin I (RCA120) and wheat germ agglutinin, indicating alterations in surface galactose and glucosamine residues. LPG isolated from mutants migrated faster than parent strain LPG on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, suggesting the mutants had shorter LPG molecules. Dionex high-performance anion exchange chromatography with pulsed amperometric detection analyses revealed galactosamine, glucosamine, galactose, glucose, mannose/xylose, and rhamnose as the main monosaccharides of T. vaginalis parent strain LPG. LPG from both mutants showed a reduction of galactose and glucosamine, corresponding with the reduced size of their LPG molecules and inability to bind the lectins RCA120 and wheat germ agglutinin. Mutant parasites were defective in attachment to plastic, a characteristic associated with avirulent strains of T. vaginalis. Moreover, the mutants were less adherent and less cytotoxic to human vaginal ectocervical cells in vitro than the parental strain. Finally, while parent strain LPG could inhibit the attachment of parent strain parasites to vaginal cells, LPG from either mutant could not inhibit attachment. These combined results demonstrate that T. vaginalis adherence to host cells is LPG mediated and that an altered LPG leads to reduced adherence and cytotoxicity of this parasite. PMID

  16. Concordance between two methods in measuring treatment adherence in patients with type 2 diabetes

    PubMed Central

    López-Simarro, Flora; Brotons, Carlos; Moral, Irene; Aguado-Jodar, Alba; Cols-Sagarra, Cèlia; Miravet-Jiménez, Sònia

    2016-01-01

    Objective We analyzed the concordance between two methods for measuring treatment adherence (TA) and studied the determinants of TA in patients with type 2 diabetes mellitus. Methods We conducted a cross-sectional descriptive study in a primary care center, involving 320 diabetic patients. TA was measured using the Haynes–Sackett (H–S) adherence test during the patient interview and based on pharmacy refill data. TA was calculated globally and by drug groups (antihypertensive, lipid-lowering, and antidiabetic drugs). Results Poor TA as measured by the H–S test was observed in 11.2% of the patients. Based on pharmacy refill data, there was a poor global TA rate of 30.3%, which was 33.3%, 26.6%, and 34.2% for oral antidiabetic, antihypertensive, and lipid-lowering drugs, respectively. Concordance between the two methods was poor. There was no relationship between the degree of disease control and TA as measured by the H–S test. Good TA measured based on pharmacy refill data for antidiabetic and antihypertensive drugs was associated with lower glycosylated hemoglobin and diastolic blood pressure values, respectively. Patients with good global TA showed lower glycosylated hemoglobin, diastolic blood pressure, and low-density lipoprotein cholesterol values. The multivariate analysis found good oral antidiabetic adherence to be associated to free pharmacy service; good antihypertensive drug adherence to the existence of comorbidities; and good lipid-lowering drug adherence to a history of ischemic heart disease, and a more experienced physician and/or female physician. Conclusion Concordance between the two methods in assessing TA was low. Approximately one-third of the patients with type 2 diabetes mellitus presented poor TA in relation to antihypertensive, lipid-lowering, and antidiabetic medication. An improved TA was associated with a better control of the studied parameters. Comorbidities, such as ischemic heart disease and access to free pharmacy service

  17. Mitogenic activation of B cells in vitro: the properties of adherent accessory cells as revealed by partition analysis.

    PubMed

    Kettman, J R; Soederberg, A; Lefkovits, I

    1986-08-15

    The requirement of B cells activated by mitogen (dextran sulfate plus lipopolysaccharide) for accessory cells was studied by partition analysis. Small numbers of splenic B cells were activated to clonal growth, as determined by visual inspection, and to immunoglobulin (Ig) synthesis, as determined by release of Ig into the culture fluid. By placing irradiated adherent cells in the periphery of the microculture wells and forcing responding cells to different areas of the well (slant experiments), it was observed that no cell contact was necessary for B cell activation, and that "promoted" contact ("Rock and Roll" experiments) does not increase the efficiency of activation. Sequential microcultures suggest that only some irradiated adherent cells act as accessory cells, but they can perform this function to more than one B cell. Attempts to perform limiting dilution analysis by varying irradiated adherent cell input showed non-single-hit behavior. When the data were rearranged, taking into account the distribution of irradiated adherent cells, then single-hit behavior with about 1 to 5% of irradiated adherent cells acting as an accessory cells for B cell clonal activation was observed. The evidence suggests that an uncommon irradiated adherent cell releases a soluble factor necessary for B cell activation and/or clonal proliferation. PMID:3488344

  18. Late Adherent Human Bone Marrow Stromal Cells Form Bone and Restore the Hematopoietic Microenvironment In Vivo

    PubMed Central

    Vianna, Verônica Fernandes; Bonfim, Danielle Cabral; Cavalcanti, Amanda dos Santos; Fernandes, Marco Cury; Kahn, Suzana Assad; Casado, Priscila Ladeira; Lima, Inayá Correa; Murray, Samuel S.; Murray, Elsa J. Brochmann; Duarte, Maria Eugenia Leite

    2013-01-01

    Bone marrow stromal cells (BMSCs) are a valuable resource for skeletal regenerative medicine because of their osteogenic potential. In spite of the very general term “stem cell,” this population of cells is far from homogeneous, and different BMSCs clones have greatly different phenotypic properties and, therefore, potentially different therapeutic potential. Adherence to a culture flask surface is a primary defining characteristic of BMSCs. We hypothesized that based on the adherence time we could obtain an enriched population of cells with a greater therapeutic potential. We characterized two populations of bone marrow-derived cells, those that adhered by three days (R-cells) and those that did not adhere by three days but did by six days (L-cells). Clones derived from L-cells could be induced into adipogenic, chondrogenic, and osteogenic differentiation in vitro. L-cells appeared to have greater proliferative capacity, as manifested by larger colony diameter and clones with higher CD146 expression. Only clones from L-cells developed bone marrow stroma in vivo. We conclude that the use of late adherence of BMSCs is one parameter that can be used to enrich for cells that will constitute a superior final product for cell therapy in orthopedics. PMID:23710460

  19. Filamentous hemagglutinin has a major role in mediating adherence of Bordetella pertussis to human WiDr cells.

    PubMed Central

    Urisu, A; Cowell, J L; Manclark, C R

    1986-01-01

    [35S]methionine-labeled Bordetella pertussis adhered to monolayers of WiDr cells, an epitheliumlike cell line from a human intestinal carcinoma. Adherence was proportional to the density of the WiDr cells and to the concentration of B. pertussis in the assay. Adherence of virulent phase I strains Tohama phase I, 114, and BP338 was much greater than adherence of avirulent strains Tohama phase III and 423 phase IV. Mutants deficient in the production of the filamentous hemagglutinin (FHA) were hemagglutination negative and adhered to WiDr cells much less efficiently than the parent strains. Preincubation of B. pertussis cells with FHA increased their hemagglutination activity and adherence to WiDr cells. Goat antibody to FHA inhibited, in a dose-dependent manner, the adherence of strain Tohama I but not the adherence of FHA-deficient mutant Tohama 325. At similar protein concentrations, normal goat antibody, goat antibody to pertussis toxin, or the Fab fragments of goat antibody to serotype 2 fimbriae had no effect on adherence. Also, an FHA-positive strain without fimbriae showed high adherence, while a fimbriated FHA-deficient mutant adhered poorly. Our data indicate that FHA plays a major role in adherence of B. pertussis to human WiDr cells. Fimbriae do not appear to mediate attachment of B. pertussis to WiDr cells. PMID:2872165

  20. Trends in adherence to dietary recommendations among Korean type 2 diabetes mellitus patients

    PubMed Central

    2015-01-01

    BACKGROUND/OBJECTIVES The current study examined trends in adherence to dietary recommendations and compared the levels of adherence between diagnosed and undiagnosed subjects with type 2 diabetes mellitus (T2DM) in Korea over the past 14 years. SUBJECTS/METHODS Data were collected from the 1998-2012 Korea National Health and Nutrition Examination Surveys (KNHANES). Diagnosed diabetes was defined as giving a positive response to questions about awareness of the disease, a physician's diagnosis of diabetes, or medical treatment for diabetes, whereas undiagnosed diabetes was defined as having a fasting glucose level ≥ 126 mg/dl. Assessment of adherence level was based on 6 components of dietary guidelines, considering meal patterns and intake levels of calories, carbohydrates, vegetable/seaweed, sodium, and alcohol. The participants received 1 point if they met the criteria for each of the 6 components, and the total possible score ranged from 0 to 6 points. Multivariate generalized linear regression was performed, taking into account the complex survey design. RESULTS Among all diabetic patients aged 30 years or older, the proportion of diagnosed diabetes increased dramatically, from 40.9% in 1998 to 75.9% in 2012 (P for trend < 0.001). The overall adherence levels to dietary recommendations were low and did not significantly differ between diagnosed and undiagnosed subjects with T2DM for all survey years. Several improvements were observed, including increased adherence to maintaining sufficient vegetable/seaweed consumption (increased from 0.12 to 0.16 points) and limiting sodium intake (increased from 0.12-0.13 points to 0.19-0.24 points; P for trend < 0.001), while adherence to maintaining moderate alcohol consumption decreased. CONCLUSIONS Analysis of data collected by the KNHANES indicates that Korean T2DM patients have poor adherence to dietary recommendations and maintenance of a healthy lifestyle, regardless of disease awareness. This finding suggests

  1. Dietary Adherence and Mealtime Behaviors in Young Children with Type 1 Diabetes on Intensive Insulin Therapy

    PubMed Central

    Patton, Susana R.; Dolan, Lawrence M.; Chen, Ming; Powers, Scott W.

    2013-01-01

    Diet is an important component of diabetes treatment and integral to successful management. While intensive insulin therapy can allow patients to eat more freely, it is not known how the rapid uptake of intensive therapy in young children with type 1 diabetes has impacted their diet and if diet and healthful eating in young children correlates with mealtime behaviors and glycemic control. This study examined diet, mealtime behaviors, and glucose control in a sample of 39 young children on intensive therapy. This was a one-sample, cross-sectional study. Children had a mean age of 5.1±1.1 years. Children’s 3-day diet diaries were assessed using a deviation scale (measure of adherence) and a healthy eating index. Mealtime behaviors were assessed using the Behavioral Pediatric Feeding Assessment Scale. Children’s glucose control was measured using continuous glucose monitoring. Children’s mean carbohydrate intake was 72%±24% of the recommended levels based on their age, sex, size, and activity level, and children exceeded national guidelines for percentage of calories from fat and saturated fat. A more healthful diet correlated with fewer child mealtime behavior problems, but better dietary adherence correlated with more parent mealtime behavior problems. Even in the context of intensive management, diet can be problematic for young children with type 1 diabetes. Parent-reported problems with mealtime behaviors seem to correlate with healthy eating and dietary adherence. PMID:23351629

  2. The Reliability and Validity of the Perceived Dietary Adherence Questionnaire for People with Type 2 Diabetes

    PubMed Central

    Asaad, Ghada; Sadegian, Maryam; Lau, Rita; Xu, Yunke; Soria-Contreras, Diana C.; Bell, Rhonda C.; Chan, Catherine B.

    2015-01-01

    Nutrition therapy is essential for diabetes treatment, and assessment of dietary intake can be time consuming. The purpose of this study was to develop a reliable and valid instrument to measure diabetic patients’ adherence to Canadian diabetes nutrition recommendations. Specific information derived from three, repeated 24-h dietary recalls of 64 type 2 diabetic patients, aged 59.2 ± 9.7 years, was correlated with a total score and individual items of the Perceived Dietary Adherence Questionnaire (PDAQ). Test-retest reliability was completed by 27 type 2 diabetic patients, aged 62.8 ± 8.4 years. The correlation coefficients for PDAQ items versus 24-h recalls ranged from 0.46 to 0.11. The intra-class correlation (0.78) was acceptable, indicating good reliability. The results suggest that PDAQ is a valid and reliable measure of diabetes nutrition recommendations. Because it is quick to administer and score, it may be useful as a screening tool in research and as a clinical tool to monitor dietary adherence. PMID:26198247

  3. Type D Personality Predicts Poor Medication Adherence in Chinese Patients with Type 2 Diabetes Mellitus: A Six-Month Follow-Up Study

    PubMed Central

    Li, Xuemei; Zhang, Shengfa; Xu, Huiwen; Tang, Xinfeng; Zhou, Huixuan; Yuan, Jiaqi; Wang, Xiaohua; Qu, Zhiyong; Wang, Fugang; Zhu, He; Guo, Shuai

    2016-01-01

    Background Type D personality and medication nonadherence have been shown to be associated with poor health outcomes. Type D personality is associated with poor medication adherence in patients with coronary artery disease, myocardial infarction, and heart failure. However, the relationship between type D personality and medication adherence in patients with Type 2 Diabetes Mellitus (T2DM) remains unknown. This study aims to examine whether type D personality was associated with medication adherence in patients with T2DM. Design and Settings A follow-up study was conducted in general hospital of the People's Liberation Army in Beijing. Methods 412 T2DM patients (205 females), who were recruited by circular systematic random sampling, provided demographic and baseline data about medical information and completed measures of Type D personality. Then, 330 patients went on to complete a self-report measure of medication adherence at the sixth month after baseline data collection. Chi-square test, t tests, and hierarchical multiple regression analyses were conducted, as needed. Results Patients with type D personality were significantly more likely to have poor medication adherence (p<0.001). Type D personality predicts poor medication adherence before and after controlling for covariates when it was analyzed as a categorical variable. However, the dimensional construct of type D personality was not associated with medication adherence when analyzed as a continuous variable. Conclusion Although, as a dimensional construct, type D personality may not reflect the components of the personality associated with poor medication adherence in patients with T2DM, screening for type D personality may help to identify those who are at higher risk of poor medication adherence. Interventions, aiming to improve medication adherence, should be launched for these high-risk patients. PMID:26894925

  4. Type of hemodialysis and preference for behavioral involvement: interactive effects on adherence in end-stage renal disease.

    PubMed

    Christensen, A J; Smith, T W; Turner, C W; Holman, J M; Gregory, M C

    1990-01-01

    Examined the effects of hemodialysis type (i.e., staff controlled, in center vs. patient controlled, home) and patient preference for behavioral involvement on adherence and emotional adjustment in a sample of 53 patients with end-stage renal disease. Consistent with person x treatment interaction models, higher levels of preference for behavioral involvement were associated with better dietary adherence (i.e., lower serum potassium) for patients receiving dialysis at home but worse dietary adherence for patients receiving treatment in a dialysis center. A similar though weaker patient x treatment type matching pattern was observed for fluid-intake adherence (i.e., interdialytic weight gain). No effects were observed for patients' self-reported depression levels. Possible mechanisms for the interactional effect on adherence are discussed. PMID:2331980

  5. Examining the interaction of parental involvement and parenting style in predicting adherence in youth with type 1 diabetes

    PubMed Central

    Landers, Sara E.; Friedrich, Elizabeth A.; Jawad, Abbas F.; Miller, Victoria A.

    2016-01-01

    Introduction This study examined whether aspects of parenting style (specifically, warmth, autonomy support, and coercion) moderated the association between parental involvement and adherence in youth with type 1 diabetes. Methods Children ages 8–16 years with type 1 diabetes and a parent completed assessments of parental involvement, parenting style, and adherence. Results Parent autonomy support and coercion were associated with adherence but warmth was not. Child report of more parental involvement was associated with better adherence. Warmth, autonomy support, and coercion were not moderators. Discussion The findings underscore the importance of parental involvement, operationalized as responsibility for diabetes tasks, and parenting style, specifically coercion and autonomy support, for adherence in pediatric chronic illness management. Longitudinal research is needed to better understand how and why dimensions of involvement (e.g., responsibility, monitoring, support) vary over time and whether they impact outcomes differentially. PMID:26866945

  6. Assessment of self-reported adherence among patients with type 2 diabetes in Matlala District Hospital, Limpopo Province

    PubMed Central

    Adegbola, Sadeen A.; Govender, Indiran; Ogunbanjo, Gboyega A.O.

    2016-01-01

    Introduction Complications associated with Diabetes Mellitus are a burden to health services, especially in resource poor settings. These complications are associated with substandard care and poor adherence to treatment plans. The aim of the study was to assess the self-reported adherence to treatment amongst patients with type 2 diabetes in Matlala District Hospital, Limpopo Province. Methods This cross-sectional study used convenience sampling with a standardised, validated questionnaire. Data were collected over 4 months, and Microsoft Excel was used for data capturing. Results We found that 137 (70%) of the participants considered themselves adherent to their diabetes medication. Younger age (p = 0.028), current employment (p = 0.018) and keeping appointment were factors significantly associated with adherence. Reasons given for poor adherence were that the clinic did not have their pills (29%), they had forgotten to take their medication (16%) and gone travelling without taking enough pills (14%). Reasons given for poor adherences to a healthy lifestyle were being too old (29%), 22% had no specific reason, 13% struggled to motivate themselves and 10% simply forgot what to do. Sixty-eight percent of the adhered participants recommended the use of medication at meal times, 14% set a reminder, and 8% used the assistance of a treatment supporter Conclusions and recommendations The study revealed a higher than expected reported level of adherence to diabetes treatment. Further research is needed to assess whether self-reported adherence corresponds to the metabolic control of the patients and to improve services. PMID:27543285

  7. Mechanical Restrictions on Biological Responses by Adherent Cells within Collagen Gels

    PubMed Central

    Simon, D.D.; Horgan, C.O.; Humphrey, J.D.

    2012-01-01

    Cell-seeded collagen and fibrin gels represent excellent assays for studying interactions between adherent interstitial cells and the three-dimensional extracellular matrix in which they reside. Over one hundred papers have employed the free-floating collagen gel assay alone since its introduction in 1979 and much has been learned about mechanobiological responses of diverse types of cells. Yet, given that mechanobiology is the study of biological responses by cells to mechanical stimuli that must respect the basic laws of mechanics, we must quantify better the mechanical conditions that are imposed on or arise in cell-seeded gels. In this paper, we suggest that cell responses and associated changes in matrix organization within the classical free-floating gel assay are highly restricted by the mechanics. In particular, many salient but heretofore unexplained or misinterpreted observations in free-floating gels can be understood in terms of apparent cell-mediated residual stress fields that satisfy quasi-static equilibria and continuity of tractions. There is a continuing need, therefore, to bring together the allied fields of mechanobiology and biomechanics as we continue to elucidate cellular function within both native connective tissues and tissue equivalents that are used in basic scientific investigations or regenerative medicine. PMID:23022259

  8. The effect of adherent and phagocytic cells on human lymphocyte PHA responsiveness.

    PubMed

    Potter, M R; Moore, M

    1977-01-01

    The effect of small numbers of adherent and phagocytic cells on the human peripheral blood lymphocyte response to PHA was examined by depleting these cells from lymphocyte preparations. Lymphocyte preparations obtained by centrifugation on Ficoll--Triosil, which contained on average 85% lymphocytes, responded well to PHA. Depletion of cells adhering to nylon fibre, giving a population containing on average 95% lymphocytes, resulted in a considerably reduced response. Depletion of cells that adhered to plastic or ingested iron powder to give populations containing on average 90% lymphocytes, also reduced the PHA response, but to a lesser extent. Reduction in PHA responsiveness correlated with increasing lymphocyte purity. The responsiveness of nylon-column-filtered cells could be restored by adding a small number of cells from a monocyte-rich population. PMID:300303

  9. Depression in Chinese patients with type 2 diabetes: associations with hyperglycemia, hypoglycemia, and poor treatment adherence

    PubMed Central

    Zhang, Yuying; Ting, Rose ZW; Yang, Wenying; Jia, Weiping; Li, Wenhui; Ji, Linong; Guo, Xiaohui; Kong, Alice PS; Wing, Yun‐Kwok; Luk, Andrea OY; Sartorius, Norman; Morisky, Donald E; Oldenburg, Brian; Weng, Jianping

    2015-01-01

    Abstract Background We hypothesize that depression in type 2 diabetes might be associated with poor glycemic control, in part due to suboptimal self‐care. We tested this hypothesis by examining the associations of depression with clinical and laboratory findings in a multicenter survey of Chinese type 2 diabetic patients. Method 2538 patients aged 18–75 years attending hospital‐based clinics in four cities in China underwent detailed clinical‐psychological‐behavioral assessment during a 12‐month period between 2011 and 2012. Depression was diagnosed if Patient Health Questionnaire‐9 (PHQ‐9) score ≥10. Diabetes self‐care and medication adherence were assessed using the Summary of Diabetes Self‐care Activities and the 4‐item Morisky medication adherence scale respectively. Results In this cross‐sectional study (mean age: 56.4 ± 10.5[SD] years, 53% men), 6.1% (n = 155) had depression. After controlling for study sites, patients with depression had higher HbA 1c (7.9 ± 2.0 vs. 7.7 ± 2.0%, P = 0.008) and were less likely to achieve HbA 1c goal of <7.0% (36.2% vs 45.6%, P = 0.004) than those without depression. They were more likely to report hypoglycemia and to have fewer days of being adherent to their recommended diet, exercise, foot care and medication. In logistic regression, apart from young age, poor education, long disease duration, tobacco use, high body mass index, use of insulin, depression was independently associated with failure to attain HbA 1c target (Odds Ratio [OR] = 1.56, 95%CI:1.05–2.32, P = 0.028). The association between depression and glycemic control became non‐significant after inclusion of adherence to diet, exercise and medication (OR = 1.48, 95% CI 0.99–2.21, P = 0.058). Conclusion Depression in type 2 diabetes was closely associated with hyperglycemia and hypoglycemia, which might be partly mediated through poor treatment adherence. PMID:25349949

  10. Cloning of a genetic determinant from Clostridium difficile involved in adherence to tissue culture cells and mucus.

    PubMed Central

    Karjalainen, T; Barc, M C; Collignon, A; Trollé, S; Boureau, H; Cotte-Laffitte, J; Bourlioux, P

    1994-01-01

    Our laboratory has previously shown that Clostridium difficile adherence to Caco-2 cells is greatly enhanced after heat shock at 60 degrees C and that it is mediated by a proteinaceous surface component. The experiments described here show that C. difficile could adhere to several types of tissue culture cells (Vero, HeLa, and KB) after heat shock. The type of culture medium (liquid or solid, with or without blood) had little effect on adhesion. To clone the adhesin gene, polyclonal antibodies against C. difficile heated at 60 degrees C were used to screen a genomic library of C. difficile constructed in lambda ZapII. Ten positive clones were identified in the library, one of which (pCL6) agglutinated several types of erythrocytes in the presence of mannose. In Western blots (immunoblots), this clone expressed in Escherichia coli a 40- and a 27-kDa protein; a 27-kDa protein has been previously identified in the surface extracts of heat-shocked C. difficile as a possible adhesin. The clone adhered to Vero, Caco-2, KB, and HeLa cells; the adherence was blocked by anti-C. difficile antibodies, by a surface extract of C. difficile, and by mucus isolated from axenic mice. Furthermore, the clone could attach ex vivo to intestinal mucus isolated from axenic mice. Preliminary studies on the receptor moieties implicated in C. difficile adhesion revealed that glucose and galactose could partially block adhesion to tissue culture cells, as did di- or trisaccharides containing these sugars, suggesting that the adhesin is a lectin. In addition, N-acetylgalactosamine, a component of mucus, and gelatin partially impeded cell attachment. Images PMID:7927694

  11. Quality of life associated with treatment adherence in patients with type 2 diabetes: a cross-sectional study

    PubMed Central

    Martínez, Yolanda V; Prado-Aguilar, Carlos A; Rascón-Pacheco, Ramón A; Valdivia-Martínez, José J

    2008-01-01

    Background Despite certain contradictions, an association has been identified between adherence to drug treatment and the quality of life in patients with type 2 diabetes. The contradictions observed emphasize the importance of using different methods to measure treatment adherence, or the association of psychological precursors of adherence with quality of life. For this reason, we have used an indirect method to measure adherence (pill count), as well as two adherence behaviour precursors (attitude and knowledge), to assess the association between adherence and the quality of life in type 2 diabetes patients. Methods A cross-sectional comparative study on a random sample of 238 type 2 diabetic patients was carried out over one year in four family medicine units of the Mexican Institute of Social Security (IMSS) in Aguascalientes, Mexico. Treatment adherence was measured using the indirect method of pill count to assess adherence behaviour, obtaining information at two home visits. In the first we recorded the medicine prescribed and in the second, we counted the medicine remaining to determine the proportion of the medicine taken. We also assessed two adherence behaviour precursors: the patients' knowledge regarding their medical prescription measured through a structured questionnaire; and attitudes to treatment adherence using a Likert scale. Quality of life was measured through the WHOQOL-100 (the WHO Quality of Life questionnaire). Information concerning both knowledge and attitude was obtained through interviews with the patients. A multiple linear regression model was constructed to establish the relationship between each quality of life domain and the variables related to adherence, controlling for covariates. Results There was no association between quality of life and treatment adherence behaviour. However, the combination of strong knowledge and a positive attitude was associated with five of the six quality of life domains. Conclusion The results

  12. Adherence to human lung microvascular endothelial cells (HMVEC-L) of Plasmodium vivax isolates from Colombia

    PubMed Central

    2013-01-01

    Background For years Plasmodium vivax has been considered the cause of benign malaria. Nevertheless, it has been observed that this parasite can produce a severe disease comparable to Plasmodium falciparum. It has been suggested that some physiopathogenic processes might be shared by these two species, such as cytoadherence. Recently, it has been demonstrated that P. vivax-infected erythrocytes (Pv-iEs) have the capacity to adhere to endothelial cells, in which intercellular adhesion molecule-1 (ICAM-1) seems to be involved in this process. Methods Adherence capacity of 21 Colombian isolates, from patients with P. vivax mono-infection to a microvascular line of human lung endothelium (HMVEC-L) was assessed in static conditions and binding was evaluated at basal levels or in tumor necrosis factor (TNF) stimulated cells. The adherence specificity for the ICAM-1 receptor was determined through inhibition with an anti-CD54 monoclonal antibody. Results The majority of P. vivax isolates, 13 out of 21 (61.9%), adhered to the HMVEC-L cells, but P. vivax adherence was at least seven times lower when compared to the four P. falciparum isolates. Moreover, HMVEC-L stimulation with TNF led to an increase of 1.6-fold in P. vivax cytoadhesion, similar to P. falciparum isolates (1.8-fold) at comparable conditions. Also, blockage of ICAM-1 receptor with specific antibodies showed a significant 50% adherence reduction. Conclusions Plasmodium vivax isolates found in Colombia are also capable of adhering specifically in vitro to lung endothelial cells, via ICAM-1 cell receptor, both at basal state and after cell stimulation with TNF. Collectively, these findings reinforce the concept of cytoadherence for P. vivax, but here, to a different endothelial cell line and using geographical distinct isolates, thus contributing to understanding P. vivax biology. PMID:24080027

  13. Fibronectin-binding protein of Streptococcus pyogenes: sequence of the binding domain involved in adherence of streptococci to epithelial cells.

    PubMed Central

    Talay, S R; Valentin-Weigand, P; Jerlström, P G; Timmis, K N; Chhatwal, G S

    1992-01-01

    The sequence of the fibronectin-binding domain of the fibronectin-binding protein of Streptococcus pyogenes (Sfb protein) was determined, and its role in streptococcal adherence was investigated by use of an Sfb fusion protein in adherence studies. A 1-kb DNA fragment coding for the binding domain of Sfb protein was cloned into the expression vector pEX31 to produce an Sfb fusion protein consisting of the N-terminal part of MS2 polymerase and a C-terminal fragment of the streptococcal protein. Induction of the vector promoter resulted in hyperexpression of fibronectin-binding fusion protein in the cytoplasm of the recombinant Escherichia coli cells. Sequence determination of the cloned 1-kb fragment revealed an in-frame reading frame for a 268-amino-acid peptide composed of a 37-amino-acid sequence which is completely repeated three times and incompletely repeated a fourth time. Cloning of one repeat into pEX31 resulted in expression of small fusion peptides that show fibronectin-binding activity, indicating that one repeat contains at least one binding domain. Each repeat exhibits two charged domains and shows high homology with the 38-amino-acid D3 repeat of the fibronectin-binding protein of Staphylococcus aureus. Sequence comparison with other streptococcal ligand-binding surface proteins, including M protein, failed to reveal significant homology, which suggests that Sfb protein represents a novel type of functional protein in S. pyogenes. The Sfb fusion protein isolated from the cytoplasm of recombinant cells was purified by fast protein liquid chromatography. It showed a strong competitive inhibition of fibronectin binding to S. pyogenes and of the adherence of bacteria to cultured epithelial cells. In contrast, purified streptococcal lipoteichoic acid showed only a weak inhibition of fibronectin binding and streptococcal adherence. These results demonstrate that Sfb protein is directly involved in the fibronectin-mediated adherence of S. pyogenes to

  14. A Comparative Evaluation of Adherence of Microorganism to Different Types of Brackets: A Scanning Electron Microscopic Study

    PubMed Central

    Shashidhar, E P; Sahitya, M; Sunil, T; Murthy, Anup R; Rani, M S

    2015-01-01

    Background: The purpose of this study was to evaluate and compare the adherence of microorganism to different types of brackets using the scanning electron microscope (SEM). A double-blinded study was undertaken to evaluate and adherence of microorganisms to different types of brackets using SEM. Materials and Methods: At random, 12 patients reporting for treatment to the department of Orthodontics VS Dental College and Hospital were selected. Four types of brackets were included in the present study stainless steel, titanium, composite, and ceramic. Brackets were bonded to teeth of the patient on all the four quadrants. The teeth included for bonding were lateral incisor, canine, first premolar, and second premolar. The brackets were left for 72 h. After 72 h brackets were debonded, and they were evaluated by SEM for adherence of microorganism in the slot and tie wings surface. The SEM images were graded, and the adherence of microorganism to the brackets in the surfaces and the four different quadrants were recorded. Results: There is a significant difference in adherence of microorganisms to the various types of brackets (P < 0.001) and the surfaces (P < 0.05) included in the study. However, there is no significance in the mean adherence of microorganisms in the different quadrants (P > 0.05) included in the study. The interaction of bracket/surface, bracket/quadrant, surface/quadrants was analyzed, there was no significance of comparison of bracket/surfaces/quadrant but the interaction of bracket/quadrant was found to be significant (<0.011). The interaction of bracket/surfaces/quadrant was also found to be significant (<0.003). Conclusion: The maximum adherence of microorganisms was observed with the composite bracket material and the least adherence of microorganisms was observed with the titanium bracket material. The adherence of microorganisms is relatively more in the slot area, when compare to the tie wings surface maximum adherence of microorganism is

  15. Inhibitory effects of α-cyperone on adherence and invasion of avian pathogenic Escherichia coli O78 to chicken type II pneumocytes.

    PubMed

    Zhang, Li-Yan; Lv, Shuang; Wu, Shuai-Cheng; Guo, Xun; Xia, Fang; Hu, Xi-Rou; Song, Zhou; Zhang, Cui; Qin, Qian-Qian; Fu, Ben-Dong; Yi, Peng-Fei; Shen, Hai-Qing; Wei, Xu-Bin

    2014-05-15

    Avian pathogenic Escherichia coli (APEC) are extra-intestinal pathogenic E. coli, and usually cause avian septicemia through breaching the blood-gas barrier. Type II pneumocytes play an important role of maintaining the function of the blood-gas barrier. However, the mechanism of APEC injuring type II pneumocytes remains unclear. α-cyperone can inhibit lung cell injury induced by Staphylococcus aureus. In order to explore whether α-cyperone regulates the adherence and invasion of APEC-O78 to chicken type II pneumocytes, we successfully cultured chicken type II pneumocytes. The results showed that α-cyperone significantly decreased the adherence of APEC-O78 to chicken type II pneumocytes. In addition, α-cyperone inhibited actin cytoskeleton polymerization induced by APEC-O78 through down regulating the expression of Nck-2, Cdc42 and Rac1. These results provide new evidence for the prevention of colibacillosis in chicken. PMID:24629766

  16. Relationship of cell surface morphology and composition of Streptococcus salivarius K+ to adherence and hydrophobicity.

    PubMed Central

    Weerkamp, A H; van der Mei, H C; Slot, J W

    1987-01-01

    The cell surfaces of a range of variants of Streptococcus salivarius HB, altered in cell wall antigen composition, were compared with those of the parent with respect to adherence, ability to adsorb to hexadecane, morphology, and exposure of lipoteichoic acid (LTA). Adherence to host surfaces was measured by using both saliva-coated hydroxyapatite beads and tissue-cultured HeLa cells, and interbacterial adherence was measured by using Veillonella alcalescens V1 cells. Progressive loss of the protease-sensitive fibril classes was generally associated with decreasing ability to adsorb to hexadecane. However, increased exposure of protein antigen C (AgC) increased the apparent hydrophobicity of the cell. This correlated with the finding that AgC was the most hydrophobic of the solubilized fibrillar cell wall antigens. Collectively, this demonstrates that adsorption to hydrophobic ligands is directly related to the density of the fibrillar layer on the cells and the properties and surface exposure of specific fibril classes. The involvement of hydrophobic interactions in AgC-associated attachment was suggested by its sensitivity to low levels of the hydrophobic bond-breaking agent tetramethyl urea, although the reduction was not to the level of adherence observed with strains lacking AgC. However, hydrophobicity was less essential to other adherence reactions. Circumstantial evidence, including immunoelectron microscopy, showing that LTA was virtually absent from the fibrillar layer, whole-cell enzyme-linked immunosorbent assay, suggesting that surface exposure of LTA related inversely to the density of the fibrillar layer, and agarose gel electrophoresis, showing that LTA was not specifically associated with protein fibrillar antigens, strongly suggested that LTA does not confer hydrophobic properties to these cells and is not involved in adherence reactions associated with the cell wall protein antigens. Images PMID:3804445

  17. A Review of Adolescent Adherence in Type 1 Diabetes and the Untapped Potential of Diabetes Providers to Improve Outcomes

    PubMed Central

    Datye, Karishma A.; Moore, Daniel J.; Russell, William E.

    2015-01-01

    Only 21 % of adolescents with type 1 diabetes (T1D) meet glycemic goals set forth by the American Diabetes Association. Adherence to therapy is a particular concern in this population, and the association between poor adherence and worsening glycemic control indicates that there is a critical need to improve adherence to therapy in adolescents with T1D. In this article, we review barriers to adherence in adolescents with T1D and discuss interventions aimed at improving adherence to therapy and glycemic control. Interventions include technology-based applications, family-based therapies, motivational interviewing, and others. Notably, less than 10 % of the interventions reviewed are provider-led, clinic-based interventions, and few have focused on regimen-related aspects of adherence. This article also outlines the importance of provider communication and the role of providers in facilitating adherence behaviors in adolescents with T1D. Finally, we suggest future directions of research to improve adherence to therapy in adolescents with T1D. PMID:26084580

  18. Further observations on the specific red cell adherence test: effects of radiation therapy

    SciTech Connect

    Richie, J.P.; Yap, W.T.

    1981-04-01

    To assess the effects of radiation therapy on the specific red cell adherence test we have evaluated 33 patients who underwent cystectomy for bladder cancer and in whom radiotherapy had been used. With this test negative tumors were found in 32 of the 33 cases. In a second series of 10 patients histologic examinations were done by biopsy before radiotherapy and by subsequent microscopic examination of the cystectomy specimen. The specific red cell adherence test results remained constant in all of these cases. These findings strongly suggest that 1) the specific red cell adherence test does remain negative after radiotherapy and 2) this test is a valuable prognosticator of the future likelihood of invasion in all patients with transitional cell carcinoma of the bladder.

  19. Akkermansia muciniphila Adheres to Enterocytes and Strengthens the Integrity of the Epithelial Cell Layer.

    PubMed

    Reunanen, Justus; Kainulainen, Veera; Huuskonen, Laura; Ottman, Noora; Belzer, Clara; Huhtinen, Heikki; de Vos, Willem M; Satokari, Reetta

    2015-06-01

    Akkermansia muciniphila is a Gram-negative mucin-degrading bacterium that resides in the gastrointestinal tracts of humans and animals. A. muciniphila has been linked with intestinal health and improved metabolic status in obese and type 2 diabetic subjects. Specifically, A. muciniphila has been shown to reduce high-fat-diet-induced endotoxemia, which develops as a result of an impaired gut barrier. Despite the accumulating evidence of the health-promoting effects of A. muciniphila, the mechanisms of interaction of the bacterium with the host have received little attention. In this study, we used several in vitro models to investigate the adhesion of A. muciniphila to the intestinal epithelium and its interaction with the host mucosa. We found that A. muciniphila adheres strongly to the Caco-2 and HT-29 human colonic cell lines but not to human colonic mucus. In addition, A. muciniphila showed binding to the extracellular matrix protein laminin but not to collagen I or IV, fibronectin, or fetuin. Importantly, A. muciniphila improved enterocyte monolayer integrity, as shown by a significant increase in the transepithelial electrical resistance (TER) of cocultures of Caco-2 cells with the bacterium. Further, A. muciniphila induced interleukin 8 (IL-8) production by enterocytes at cell concentrations 100-fold higher than those for Escherichia coli, suggesting a very low level of proinflammatory activity in the epithelium. In conclusion, our results demonstrate that A. muciniphila adheres to the intestinal epithelium and strengthens enterocyte monolayer integrity in vitro, suggesting an ability to fortify an impaired gut barrier. These results support earlier associative in vivo studies and provide insights into the interaction of A. muciniphila with the host. PMID:25795669

  20. Akkermansia muciniphila Adheres to Enterocytes and Strengthens the Integrity of the Epithelial Cell Layer

    PubMed Central

    Reunanen, Justus; Kainulainen, Veera; Huuskonen, Laura; Ottman, Noora; Belzer, Clara; Huhtinen, Heikki; de Vos, Willem M.

    2015-01-01

    Akkermansia muciniphila is a Gram-negative mucin-degrading bacterium that resides in the gastrointestinal tracts of humans and animals. A. muciniphila has been linked with intestinal health and improved metabolic status in obese and type 2 diabetic subjects. Specifically, A. muciniphila has been shown to reduce high-fat-diet-induced endotoxemia, which develops as a result of an impaired gut barrier. Despite the accumulating evidence of the health-promoting effects of A. muciniphila, the mechanisms of interaction of the bacterium with the host have received little attention. In this study, we used several in vitro models to investigate the adhesion of A. muciniphila to the intestinal epithelium and its interaction with the host mucosa. We found that A. muciniphila adheres strongly to the Caco-2 and HT-29 human colonic cell lines but not to human colonic mucus. In addition, A. muciniphila showed binding to the extracellular matrix protein laminin but not to collagen I or IV, fibronectin, or fetuin. Importantly, A. muciniphila improved enterocyte monolayer integrity, as shown by a significant increase in the transepithelial electrical resistance (TER) of cocultures of Caco-2 cells with the bacterium. Further, A. muciniphila induced interleukin 8 (IL-8) production by enterocytes at cell concentrations 100-fold higher than those for Escherichia coli, suggesting a very low level of proinflammatory activity in the epithelium. In conclusion, our results demonstrate that A. muciniphila adheres to the intestinal epithelium and strengthens enterocyte monolayer integrity in vitro, suggesting an ability to fortify an impaired gut barrier. These results support earlier associative in vivo studies and provide insights into the interaction of A. muciniphila with the host. PMID:25795669

  1. The cell-cell junctions of mammalian testes: I. The adhering junctions of the seminiferous epithelium represent special differentiation structures.

    PubMed

    Domke, Lisa M; Rickelt, Steffen; Dörflinger, Yvette; Kuhn, Caecilia; Winter-Simanowski, Stefanie; Zimbelmann, Ralf; Rosin-Arbesfeld, Rina; Heid, Hans; Franke, Werner W

    2014-09-01

    The seminiferous tubules and the excurrent ducts of the mammalian testis are physiologically separated from the mesenchymal tissues and the blood and lymph system by a special structural barrier to paracellular translocations of molecules and particles: the "blood-testis barrier", formed by junctions connecting Sertoli cells with each other and with spermatogonial cells. In combined biochemical as well as light and electron microscopical studies we systematically determine the molecules located in the adhering junctions of adult mammalian (human, bovine, porcine, murine, i.e., rat and mouse) testis. We show that the seminiferous epithelium does not contain desmosomes, or "desmosome-like" junctions, nor any of the desmosome-specific marker molecules and that the adhering junctions of tubules and ductules are fundamentally different. While the ductules contain classical epithelial cell layers with E-cadherin-based adherens junctions (AJs) and typical desmosomes, the Sertoli cells of the tubules lack desmosomes and "desmosome-like" junctions but are connected by morphologically different forms of AJs. These junctions are based on N-cadherin anchored in cytoplasmic plaques, which in some subforms appear thick and dense but in other subforms contain only scarce and loosely arranged plaque structures formed by α- and β-catenin, proteins p120, p0071 and plakoglobin, together with a member of the striatin family and also, in rodents, the proteins ZO-1 and myozap. These N-cadherin-based AJs also include two novel types of junctions: the "areae adhaerentes", i.e., variously-sized, often very large cell-cell contacts and small sieve-plate-like AJs perforated by cytoplasm-to-cytoplasm channels of 5-7 nm internal diameter ("cribelliform junctions"). We emphasize the unique character of this epithelium that totally lacks major epithelial marker molecules and structures such as keratin filaments and desmosomal elements as well as EpCAM- and PERP-containing junctions. We also

  2. Impact Mediated Loading Cytoplasmic Loading of Macromolecules into Adherent Cells

    NASA Technical Reports Server (NTRS)

    Clarke, Mark S. F.; Feeback, Daniel L.; Vanderburg, Charles R.

    2003-01-01

    The advent of modern molecular biology, including the development of gene array technologies, has resulted in an explosion of information concerning the specific genes activated during normal cellular development, as well as those associated with a variety of pathological conditions. These techniques have served as a highly efficient, broacI.-based screening approach for those specific genes involved. in regulating normal cellular physiology and identifying candidate genes directly associated with the etiology of specific disease states. However, this approach provides information at the transcriptional' level only and does not necessarily indicate . that the gene in question is in fact translated ito a protein, or whether or not post-translational modification of the protein occurs. The critical importance of post-translational modification (i.e. phosphorylation, glycosylation, sialyation, etc.) to protein function has been recognized with regard to a number of proteins involved in a variety of important disease states. For example, altered glycosylation of beta-amyloid precursor protein results in an increase in the amount of beta-amyloid peptide generated and hence secreted as insoluble extracellular amyloid deposits (Georgopoulou, McLaughlin et al. 2001; Walter, Fluhrer et al. 2001), a pathological hal1nark of Alzheimer's disease. Abnormal phosphorylaion of synapsin I has been linked to alterations in synaptic vesicle trafficking leading to defective neurotransmission in Huntington's disease (Lievens, Woodman et al. 2002). Altered phosphorylation of the TAU protein involved in microtubule function has been linked to a number of neurodegenative diseases such as Alzheimer's disease (Billingsley and Kincaid 1997; Sanchez, Alvarez-Tllada et a1. 2001). Aberrant siaIyation of cell/I surface antigens has been detected in a number of different tumor cell types and has been linked to the acquisition of a neoplastic phenotype (Sell 1990), while improper' sia1yation of

  3. Human Cardiac-Derived Adherent Proliferating Cells Reduce Murine Acute Coxsackievirus B3-Induced Myocarditis

    PubMed Central

    Miteva, Kapka; Haag, Marion; Peng, Jun; Savvatis, Kostas; Becher, Peter Moritz; Seifert, Martina; Warstat, Katrin; Westermann, Dirk; Ringe, Jochen; Sittinger, Michael; Schultheiss, Heinz-Peter

    2011-01-01

    Background Under conventional heart failure therapy, inflammatory cardiomyopathy typically has a progressive course, indicating a need for alternative therapeutic strategies to improve long-term outcomes. We recently isolated and identified novel cardiac-derived cells from human cardiac biopsies: cardiac-derived adherent proliferating cells (CAPs). They have similarities with mesenchymal stromal cells, which are known for their anti-apoptotic and immunomodulatory properties. We explored whether CAPs application could be a novel strategy to improve acute Coxsackievirus B3 (CVB3)-induced myocarditis. Methodology/Principal Findings To evaluate the safety of our approach, we first analyzed the expression of the coxsackie- and adenovirus receptor (CAR) and the co-receptor CD55 on CAPs, which are both required for effective CVB3 infectivity. We could demonstrate that CAPs only minimally express both receptors, which translates to minimal CVB3 copy numbers, and without viral particle release after CVB3 infection. Co-culture of CAPs with CVB3-infected HL-1 cardiomyocytes resulted in a reduction of CVB3-induced HL-1 apoptosis and viral progeny release. In addition, CAPs reduced CD4 and CD8 T cell proliferation. All CAPs-mediated protective effects were nitric oxide- and interleukin-10-dependent and required interferon-γ. In an acute murine model of CVB3-induced myocarditis, application of CAPs led to a decrease of cardiac apoptosis, cardiac CVB3 viral load and improved left ventricular contractility parameters. This was associated with a decline in cardiac mononuclear cell activity, an increase in T regulatory cells and T cell apoptosis, and an increase in left ventricular interleukin-10 and interferon-γ mRNA expression. Conclusions We conclude that CAPs are a unique type of cardiac-derived cells and promising tools to improve acute CVB3-induced myocarditis. PMID:22174827

  4. Awareness, agreement, adoption and adherence to type 2 diabetes mellitus guidelines: a survey of Indonesian primary care physicians

    PubMed Central

    2014-01-01

    Background To assess the degree of awareness, agreement, adoption and adherence of physicians in Indonesia to type 2 diabetes mellitus guidelines, and their association with characteristics of the responders. Methods Questionnaire survey among General Practitioners (GPs) attending the Indonesian Association of Family Practitioners annual conference in November 2012. The proportion of GPs who were aware of, agreed with, adopted and adhered to the seven recommendations in the guidelines (screening for diabetes, diagnosis, lifestyle modification, use of sulfonylurea, target blood glucose, target blood pressure and use of statin) were calculated in the total number of responders. Results Of the 399 GPs participating, 383 (89%) were aware of the existence of Indonesian type 2 diabetes guidelines. Awareness for each recommendation varied from 66 to 91%. The recommendation to use a random blood glucose test for diagnosing patients with classic diabetes symptoms had the least awareness (265/399, 66%) and least agreement (163/399, 41%). The recommendation on statin use was the least adopted (192/399, 48%), while the least adherence (7/399, 2%) was found for the recommendation on screening for diabetes for patients with risk factors. Years of practice experience and proportion of diabetes patients seen in their practice were independently related with adherence to statin prescription. Conclusions High awareness of the Indonesian type 2 diabetes guideline does not necessary lead to adoption or adherence to recommendations important for outcomes and quality of care. The awareness-to-adherence model helps in identifying barriers for the use of guidelines. PMID:24755412

  5. Obedience and motivation as mechanisms for adherence to medication: a study in obese type 2 diabetic patients

    PubMed Central

    Reach, Gérard

    2011-01-01

    Objective To clarify the mechanisms of adherence. Methods A cross-sectional, multicenter French study using a self-questionnaire administered by 116 general practitioners to 782 obese type 2 diabetic patients. Results The analysis of 670 completed questionnaires revealed a strong association between the adherence to medication and the behavior of fastening the seatbelt when seated in the rear of a car. Multivariate analysis indicated that this behavior was an independent determinant of adherence to medication (odds ratio [OR] 2.3, 95% confidence interval [CI] 1.4–3.6, P < 0.001) with the same OR as the motivation to adhere to medical prescriptions (OR 2.2, 95% CI 1.3–3.6, P = 0.003) in a model with good accuracy (area under the receiver operating characteristic curve 0.774). A multiple correspondence analysis suggested that adherence to medication and seatbelt behavior are “homologous” behaviors, with homology between phenomena defined by the fact that they share a common etiology. Conclusion Adherence may have two dimensions: passive (obedience, the main determinant of seatbelt behavior) and active (motivation). This conclusion has theoretical and practical implications. Firstly, empowerment through patient education can be defined as a process that replaces the passive mechanism of adherence in patients’ minds with an active, conscious choice. Secondly, recognizing these two dimensions may help to establish a tailored patient-physician relationship to prevent nonadherence. PMID:22114466

  6. Adhesion and function of rat liver cells adherent to silk fibroin/collagen blend films.

    PubMed

    Cirillo, B; Morra, M; Catapano, G

    2004-01-01

    Collagen is often used in bioartificial livers as a biomimetic coating to promote liver cell adhesion and differentiation. Animal proteins are expensive and expose the host to risks of cross-species infection due to contamination with prions. Silk fibroin (SF) is a biocompatible protein produced by Bombyx mori silk worms and possibly an alternative to collagen. We prepared SF-collagen blend films with different SF content adherent to the bottom of standard tissue culture dishes, and characterized their surface morphology by SEM, their wettability and examined them for their capacity to support rat liver cell adhesion and metabolism. Cell metabolism was characterized by estimating the rate at which cells eliminated ammonia and synthesized urea for up to 48h of culture. SF-containing films were smooth, clear and more wettable than collagen. Cells readily adhered, formed junctions and small size aggregates on all films. As many cells adhered on SF as on collagen films. Cell adhesion to high collagen content blend films could not be reliably estimated because cells dwelt in the large cavities in the film. The effect of SF on cell metabolism differed with the investigated metabolic pathway. However, cells on SF-containing films eliminated ammonia and synthesized urea at rates generally comparable to, for urea synthesis at times higher than, that of cells on collagen. These results suggest that silk fibroin is a suitable substratum for liver cell attachment and culture, and a potential alternative to collagen as a biomimetic coating. PMID:14984185

  7. Localized, macromolecular transport for thin, adherent, single cells via an automated, single cell electroporation biomanipulator.

    PubMed

    Sakaki, Kelly; Esmaeilsabzali, Hadi; Massah, Shabnam; Prefontaine, Gratien G; Dechev, Nikolai; Burke, Robert D; Park, Edward J

    2013-11-01

    Single cell electroporation (SCE), via microcapillary, is an effective method for molecular, transmembrane transport used to gain insight on cell processes with minimal preparation. Although possessing great potential, SCE is difficult to execute and the technology spans broad fields within cell biology and engineering. The technical complexities, the focus and expertise demanded during manual operation, and the lack of an automated SCE platform limit the widespread use of this technique, thus the potential of SCE has not been realized. In this study, an automated biomanipulator for SCE is presented. Our system is capable of delivering molecules into the cytoplasm of extremely thin cellular features of adherent cells. The intent of the system is to abstract the technical challenges and exploit the accuracy and repeatability of automated instrumentation, leaving only the focus of the experimental design to the operator. Each sequence of SCE including cell and SCE site localization, tip-membrane contact detection, and SCE has been automated. Positions of low-contrast cells are localized and "SCE sites" for microcapillary tip placement are determined using machine vision. In addition, new milestones within automated cell manipulation have been achieved. The system described herein has the capability of automated SCE of "thin" cell features less than 10 μm in thickness. Finally, SCE events are anticipated using visual feedback, while monitoring fluorescing dye entering the cytoplasm of a cell. The execution is demonstrated by inserting a combination of a fluorescing dye and a reporter gene into NIH/3T3 fibroblast cells. PMID:23771309

  8. Proteins other than the Locus of Enterocyte Effacement-encoded proteins may contribute to Escherichia coli O157:H7 adherence to bovine rectoanal junction stratified squamous epithelial cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this study, the Type III Secretion System (TTSS) proteins considered critical for Escherichia coli O157 (O157) adherence to the follicle-associated epithelial (FAE) cells at the bovine recto-anal junction (RAJ), did not appear to contribute to O157 adherence to the RAJ squamous epithelial (RSE) ...

  9. Microfluidic Probe for Single-Cell Lysis and Analysis in Adherent Tissue Culture

    PubMed Central

    Lauffenburger, Douglas A.; Han, Jongyoon

    2014-01-01

    Single-cell analysis provides information critical to understanding key disease processes that are characterized by significant cellular heterogeneity. Few current methods allow single-cell measurement without removing cells from the context of interest, which not only destroys contextual information but also may perturb the process under study. Here we present a microfluidic probe that lyses single adherent cells from standard tissue culture and captures the contents to perform single-cell biochemical assays. We use this probe to measure kinase and housekeeping protein activities, separately or simultaneously, from single human hepatocellular carcinoma cells in adherent culture. This tool has the valuable ability to perform measurements that clarify connections between extracellular context, signals and responses, especially in cases where only a few cells exhibit a characteristic of interest. PMID:24594667

  10. The Malaysian Medication Adherence Scale (MALMAS): Concurrent Validity Using a Clinical Measure among People with Type 2 Diabetes in Malaysia

    PubMed Central

    Lai, Pauline Siew Mei; Morisky, Donald E.

    2015-01-01

    Medication non-adherence is a prevalent problem worldwide but up to today, no gold standard is available to assess such behavior. This study was to evaluate the psychometric properties, particularly the concurrent validity of the English version of the Malaysian Medication Adherence Scale (MALMAS) among people with type 2 diabetes in Malaysia. Individuals with type 2 diabetes, aged 21 years and above, using at least one anti-diabetes agent and could communicate in English were recruited. The MALMAS was compared with the 8-item Morisky Medication Adherence Scale (MMAS-8) to assess its convergent validity while concurrent validity was evaluated based on the levels of glycated hemoglobin (HbA1C). Participants answered the MALMAS twice: at baseline and 4 weeks later. The study involved 136 participants. The MALMAS achieved acceptable internal consistency (Cronbach’s alpha=0.565) and stable reliability as the test-retest scores showed fair correlation (Spearman’s rho=0.412). The MALMAS has good correlation with the MMAS-8 (Spearman’s rho=0.715). Participants who were adherent to their anti-diabetes medications had significantly lower median HbA1C values than those who were non-adherence (7.90 versus 8.55%, p=0.032). The odds of participants who were adherent to their medications achieving good glycemic control was 3.36 times (95% confidence interval: 1.09-10.37) of those who were non-adherence. This confirms the concurrent validity of the MALMAS. The sensitivity of the MALMAS was 88.9% while its specificity was 29.6%. The findings of this study further substantiates the reliability and validity of the MALMAS, in particular its concurrent validity and sensitivity for assessing medication adherence of people with type 2 diabetes in Malaysia. PMID:25909363

  11. Cell-mediated cytotoxicity of bovine mononuclear cells to IBRV-infected cells: dependence on Sephadex G-10 adherent cells.

    PubMed

    Campos, M; Rossi, C R

    1985-04-01

    Following intranasal inoculation of cattle with infectious bovine rhinotracheitis virus (IBRV) mononuclear cells that produced a genetically unrestricted cytotoxic response against IBRV-infected, but not against uninfected cells, were present in peripheral blood. Cytotoxicity was detected between 6 and 14 days after primary infection in a 20 h, but not in a 5 h, 51Cr-release assay. Cytotoxic activity was present in peripheral blood mononuclear cells from infected and subsequently hyperimmunized cattle for a considerably longer time. Neither natural cytotoxicity, antibody-dependent cell cytotoxicity, nor antibody produced during the assay was responsible for the cytotoxicity. However, cytotoxicity was dependent upon an adherent mononuclear cell that was partially removed by passage over nylon wool and completely removed by passage over Sephadex G-10. PMID:2408374

  12. Traction Stresses Exerted by Adherent Cells: From Angiogenesis to Metastasis

    NASA Astrophysics Data System (ADS)

    Reinhart-King, Cynthia

    2010-03-01

    Cells exert traction stresses against their substrate that mediate their ability to sense the mechanical properties of their microenvironment. These same forces mediate cell adhesion, migration and the formation of stable cell-cell contacts during tissue formation. In this talk, I will present our data on the traction stresses generated by endothelial cells and metastatic breast cancer cells focused on understanding the processes of angiogenesis and metastasis, respectively. In the context of capillary formation, our data indicate that the mechanics of the substrate play a critical role in establishing endothelial cell-cell contacts. On more compliant substrates, endothelial cell shape and traction stresses polarize and promote the formation of stable cell-cell contacts. On stiffer substrates, traction stresses are less polarized and cell connectivity is disrupted. These data indicate that the mechanical properties of the microenvironment may drive cell connectivity and the formation of stable cell-cell contacts through the reorientation of traction stresses. In our studies of metastatic cell migration, we have found that traction stresses increase with increasing metastatic potential. We investigated three lines of varying metastatic potential (MCF10A, MCF7 and MDAMB231). MDAMB231, which are the most invasive, exert the most significant forces as measured by Traction Force Microscopy. These data present the possibility that cellular traction stress generation aids in the ability of metastatic cells to migrate through the matrix-dense tumor microenvironment. Such measurements are integral to link the mechanical and chemical microenvironment with the resulting response of the cell in health and disease.

  13. [Ability of Staphylococcus cohnii strains to adhere to epithelial cells and solid surfaces in the hospital environment].

    PubMed

    Waldon, Edyta; Szewczyk, Eligia M

    2002-01-01

    Presented study describes abilities of staphylococci to adhere to exfoliated cheek and uroepithelial epithelium cells and to various surfaces such as plastics, glass and steel. The subject of the study were strains of Staphylococcus cohnii ssp. cohnii and Staphylococcus cohnii ssp. urealyticus isolated from Intensive Care Unit of Pediatric Hospital. Staphylococcus cohnii ssp.cohnii adhered in great number to epithelial cells. However, the adhesion differed by individual strains. We did not find relationship between slime production and adherence to epithelial cell. Most of investigated strains adhered closely to surfaces--especially of plastics and glass. This phenomenon was stronger in the presence of culture medium and phosphate buffer. PMID:12185691

  14. In vitro inhibition of Helicobacter pylori growth and adherence to gastric mucosal cells by Pycnogenol.

    PubMed

    Rohdewald, Peter; Beil, Winfried

    2008-05-01

    The emergence of antibiotic resistant H. pylori strains has necessitated the identification of alternative additive therapies for the treatment of this infection. The study tested whether a specific pine bark extract (Pycnogenol is effective in inhibiting the growth and adherence of H. pylori in vitro. Inhibition of H. pylori growth by Pycnogenol was tested in liquid medium as well as in an in vitro model by using sessile bacteria attached to AGS cells. Adherence was determined by co-incubation of gastric cells with Pycnogenol and H. pylori in vitro. Pycnogenol inhibited H. pylori growth in suspension with an MIC(50) of 12.5 microg/mL. Growth of H. pylori in infected cells was reduced to 10% of the control value by 125 microg/mL Pycnogenol. Adherence of H. pylori to gastric cells was reduced by 70% after 3 h incubation with 125 microg/mL Pycnogenol. The results show a significant, yet limited inhibition of growth and adherence of H. pylori to gastric cells by Pycnogenol. In vivo studies have to demonstrate the clinical relevance of these findings. PMID:18350522

  15. The many ways adherent cells respond to applied stretch.

    PubMed

    Sears, Candice; Kaunas, Roland

    2016-05-24

    Cells in various tissues are subjected to mechanical stress and strain that have profound effects on cell architecture and function. The specific response of the cell to applied strain depends on multiple factors, including cell contractility, spatial and temporal strain pattern, and substrate dimensionality and rigidity. Recent work has demonstrated that the cell response to applied strain depends on a complex combination of these factors, but the way these factors interact to elicit a specific response is not intuitive. We submit that an understanding of the integrated response of a cell to these factors will provide new insight into mechanobiology and contribute to the effective design of deformable engineered scaffolds meant to provide appropriate mechanical cues to the resident cells. PMID:26515245

  16. RGD-functionalized spherulites as targeted vectors captured by adherent cultured cells.

    PubMed

    Chenevier, P; Delord, B; Amédée, J; Bareille, R; Ichas, F; Roux, D

    2002-12-16

    Spherulites are multilamellar vesicles consisting of concentric shells that can encapsulate small organic molecules or macromolecules. We investigate the possibility of targeting neutral spherulites to adherent culture cells by functionalizing their surface with RGD-containing ligands. The strength and specificity of association of RGD spherulites with several cell lines (EAhy 926 endothelial cell line, human umbilical vein endothelial cell (HUVEC) and human osteoprogenitor (HOP) primary cells) was studied, and the molecular interaction of RGD spherulites with the EAhy 926 cell surface was investigated. We show that, after binding to cells, spherulites are internalized. PMID:12431780

  17. Screening ToxCast™ Phase I Chemicals in a Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) Assay

    EPA Science Inventory

    An Adherent Cell Differentiation and Cytotoxicity (ACDC) in vitro assay with mouse embryonic stem cells was used to screen the ToxCast Phase I chemical library for effects on cellular differentiation and cell number. The U.S. Environmental Protection Agency (EPA) established the ...

  18. Cell prestress. I. Stiffness and prestress are closely associated in adherent contractile cells

    NASA Technical Reports Server (NTRS)

    Wang, Ning; Tolic-Norrelykke, Iva Marija; Chen, Jianxin; Mijailovich, Srboljub M.; Butler, James P.; Fredberg, Jeffrey J.; Stamenovic, Dimitrije; Ingber, D. E. (Principal Investigator)

    2002-01-01

    The tensegrity hypothesis holds that the cytoskeleton is a structure whose shape is stabilized predominantly by the tensile stresses borne by filamentous structures. Accordingly, cell stiffness must increase in proportion with the level of the tensile stress, which is called the prestress. Here we have tested that prediction in adherent human airway smooth muscle (HASM) cells. Traction microscopy was used to measure the distribution of contractile stresses arising at the interface between each cell and its substrate; this distribution is called the traction field. Because the traction field must be balanced by tensile stresses within the cell body, the prestress could be computed. Cell stiffness (G) was measured by oscillatory magnetic twisting cytometry. As the contractile state of the cell was modulated with graded concentrations of relaxing or contracting agonists (isoproterenol or histamine, respectively), the mean prestress ((t)) ranged from 350 to 1,900 Pa. Over that range, cell stiffness increased linearly with the prestress: G (Pa) = 0.18(t) + 92. While this association does not necessarily preclude other interpretations, it is the hallmark of systems that secure shape stability mainly through the prestress. Regardless of mechanism, these data establish a strong association between stiffness of HASM cells and the level of tensile stress within the cytoskeleton.

  19. The agreement of patient-reported versus observed medication adherence in type 2 diabetes mellitus (T2DM)

    PubMed Central

    Kelly, Katherine; Grau-Sepulveda, Maria V; Goldstein, Benjamin A; Spratt, Susan E; Wolfley, Anne; Hatfield, Vicki; Murphy, Monica; Jones, Ellen; Granger, Bradi B

    2016-01-01

    Objective Medication adherence in type 2 diabetes mellitus (T2DM) improves glycemic control and is associated with reduced adverse clinical events, and accurately assessing adherence assessment is important. We aimed to determine agreement between two commonly used adherence measures—the self-reported Morisky Medication Adherence Scale (MMAS) and direct observation of medication use by nurse practitioners (NPs) during home visits—and determine the relationship between each measure and glycated hemoglobin (HbA1c). Research design and methods We evaluated agreement between adherence measures in the Southeastern Diabetes Initiative (SEDI) prospective clinical intervention home visit cohort, which included high-risk patients (n=430) in 4 SEDI-participating counties. The mean age was 58.7 (SD 11.6) years. The majority were white (n=210, 48.8%), female (n=236, 54.9%), living with a partner (n=316, 74.5%), and insured by Medicare/Medicaid (n=361, 84.0%). Medication adherence was dichotomized to ‘adherent’ or ‘not adherent’ using established cut-points. Inter-rater agreement was evaluated using Cohen's κ coefficient. Relationships among adherence measures and HbA1c were evaluated using the Wilcoxon rank-sum test and c-statistics. Results Fewer patients (n=261, 61%) were considered adherent by self-reported MMAS score versus the NP-observed score (n=338; 79%). Inter-rater agreement between the two adherence measures was fair (κ=0.24; 95% CI 0.15 to 0.33; p<0.0001). Higher adherence was significantly associated with lower HbA1c levels for both measures, yet discrimination was weak (c-statistic=0.6). Conclusions Agreement between self-reported versus directly observed medication adherence was lower than expected. Though scores for both adherence measures were significantly associated with HbA1c, neither discriminated well for discrete levels of HbA1c. PMID:27403322

  20. Escherichia coli isolated from a Crohn's disease patient adheres, invades, and induces inflammatory responses in polarized intestinal epithelial cells.

    PubMed

    Eaves-Pyles, Tonyia; Allen, Christopher A; Taormina, Joanna; Swidsinski, Alexander; Tutt, Christopher B; Jezek, G Eric; Islas-Islas, Martha; Torres, Alfredo G

    2008-07-01

    Inflammatory diseases of the intestinal tract are a major health concern both in the United States and around the world. Evidence now suggests that a new category of Escherichia coli, designated Adherent Invasive E. coli (AIEC) is highly prevalent in Crohn's Disease (CD) patients. AIEC strains have been shown to colonize and adhere to intestinal epithelial cells (IEC). However, the role AIEC strains play in the induction of an inflammatory response is not known. Therefore, we examined several E. coli strains (designated LF82, O83:H1, 6604 and 6655) that were isolated from CD patients for their ability to induce inflammation in two IEC, Caco-2BBe and T-84 cells. Results showed that each strain had varying abilities to adhere to and invade IEC as well as induced cytokine secretion from polarized IEC. However, E. coli O83:H1 displayed the best characteristics of AIEC strains as compared to the prototype AIEC strain LF82, inducing cytokine secretion from IEC and promoting immune cell migration through IEC. Upon further analysis, E. coli O83:H1 did not harbor virulence genes present in known pathogenic intestinal organisms. Further characterization of E. coli O83:H1 virulence determinants showed that a non-flagellated O83:H1 strain significantly decreased the organism's ability to adhere to and invade both IEC and elicit IEC cytokine secretion compared to the wild type and complemented strains. These findings demonstrate that E. coli O83:H1 possesses the characteristics of the AIEC LF82 strain that may contribute to the low-grade, chronic inflammation observed in Crohn's disease. PMID:17900983

  1. Triggering Death of Adherent Cells with Ultraviolet Radiation.

    PubMed

    Crowley, Lisa C; Waterhouse, Nigel J

    2016-01-01

    Ultraviolet (UV) radiation is a convenient stimulus for triggering cell death that is available in most laboratories. We use a Stratalinker UV cross-linker because it is a safe, cheap, reliable, consistent, and easily controlled source of UV irradiation. This protocol describes using a Stratalinker to trigger UV-induced death of HeLa cells. PMID:27371593

  2. Adherence of Candida albicans to a cell surface polysaccharide receptor on Streptococcus gordonii.

    PubMed Central

    Holmes, A R; Gopal, P K; Jenkinson, H F

    1995-01-01

    Candida albicans ATCC 10261 and CA2 bound to cells of the oral bacteria Streptococcus gordonii, Streptococcus oralis, and Streptococcus sanguis when these bacteria were immobilized onto microtiter plate wells, but they did not bind to cells of Streptococcus mutans or Streptococcus salivarius. Cell wall polysaccharide was extracted with alkali from S. gordonii NCTC 7869, the streptococcal species to which C. albicans bound with highest affinity, and was effective in blocking the coaggregation of C. albicans and S. gordonii cells in the fluid phase. When fixed to microtiter plate wells, the S. gordonii polysaccharide was bound by all strains of C. albicans tested. The polysaccharide contained Rha, Glc, GalNAc, GlcNAc, and Gal and was related compositionally to previously characterized cell wall polysaccharides from strains of S. oralis and S. sanguis. The adherence of yeast cells to the immobilized polysaccharide was not inhibitable by a number of saccharides. Antiserum raised to the S. gordonii NCTC 7869 polysaccharide blocked adherence of C. albicans ATCC 10261 to the polysaccharide. The results identify a complex cell wall polysaccharide of S. gordonii as the coaggregation receptor for C. albicans. Adherent interactions of yeast cells with streptococci and other bacteria may be important for colonization of both hard and soft oral surfaces by C. albicans. PMID:7729891

  3. Partners' representations of diabetes as mediators between patients' representations and adherence to self-care behaviors, in type 2 diabetes.

    PubMed

    Pereira, M Graça; Pedras, Susana; Machado, José C; Ferreira, Gabriela

    2016-09-01

    The purpose of this study was to analyze partners' representations of diabetes as mediators between patients' illness representations and adherence to all self-care behaviors, in recently diagnosed type 2 diabetes (T2DM) patients. The sample included 340 patients and their respective partners. The instruments used were: Revised Summary of Diabetes Self-Care Activities (RSDSCA); Medication Adherence Report Scale (MARS); and the Brief Illness Perception Questionnaire  (Brief-IPQ). A mediational effect of partners' representation of diabetes consequences was found between the same patients' representations and  exercise, foot care, and self-monitoring of blood glucose. Partners' representations of personal and treatment control, were mediators between the same partners' representations and self-monitoring of blood glucose. No partners' representations mediated patients' representation and adherence to medication or diet . This study emphasized partners' representations on patient's adherence to exercise, foot care and monitoring of blood glucose, in recent diagnosed T2DM patients. Interventions to promote adherence in T2DM should promote convergence between patients and partners' diabetes representations. This study provides some evidence for the need to treat T2DM within the dyad to improve adherence, starting after the diagnosis. PMID:26718034

  4. The functions of the variable lipoprotein family of Mycoplasma hyorhinis in adherence to host cells.

    PubMed

    Xiong, Qiyan; Wang, Jia; Ji, Yan; Ni, Bo; Zhang, Bixiong; Ma, Qinghong; Wei, Yanna; Xiao, Shaobo; Feng, Zhixin; Liu, Maojun; Shao, Guoqing

    2016-04-15

    Mycoplasma hyorhinis (M. hyorhinis) is a swine pathogen that is associated with various human cancers and contamination in cell cultures. However, no studies on the adhesion molecules of this pathogen have yet been reported. The variable lipoprotein (Vlp) family is an important surface component of M. hyorhinis. Herein, we performed several experiments to identify the function of the Vlp family in adherence to host cells. Seven recombinant Vlp (rVlp) proteins were expressed in Escherichia coli and purified by affinity chromatography. The potential role of rVlp adherence to pig kidney (PK-15) and swine tracheal epithelial (STEC) cells was then studied by indirect immunofluorescence assay and microtiter plate adherence assay. Adhesion of M. hyorhinis to PK-15 and STEC cells was specifically inhibited by the addition of a cocktail of rVlp proteins. The rVlp protein mixture was shown to bind to both PK-15 and STEC cells. The binding increased in a dose-dependent manner and could be blocked by antisera against the rVlp proteins. Most of the rVlp proteins could bind individually to both PK-15 and STEC cells except for rVlpD and rVlpF, which bound only to STEC cells. Because Vlp members vary in size among different strains and generations, they may vary in their cytoadhesion capabilities in various strains. In summary, the present results indicate that the Vlp family functions as adhesins of M. hyorhinis. PMID:27016761

  5. Optical painting and fluorescence activated sorting of single adherent cells labelled with photoswitchable Pdots.

    PubMed

    Kuo, Chun-Ting; Thompson, Alison M; Gallina, Maria Elena; Ye, Fangmao; Johnson, Eleanor S; Sun, Wei; Zhao, Mengxia; Yu, Jiangbo; Wu, I-Che; Fujimoto, Bryant; DuFort, Christopher C; Carlson, Markus A; Hingorani, Sunil R; Paguirigan, Amy L; Radich, Jerald P; Chiu, Daniel T

    2016-01-01

    The efficient selection and isolation of individual cells of interest from a mixed population is desired in many biomedical and clinical applications. Here we show the concept of using photoswitchable semiconducting polymer dots (Pdots) as an optical 'painting' tool, which enables the selection of certain adherent cells based on their fluorescence, and their spatial and morphological features, under a microscope. We first develop a Pdot that can switch between the bright (ON) and dark (OFF) states reversibly with a 150-fold contrast ratio on irradiation with ultraviolet or red light. With a focused 633-nm laser beam that acts as a 'paintbrush' and the photoswitchable Pdots as the 'paint', we select and 'paint' individual Pdot-labelled adherent cells by turning on their fluorescence, then proceed to sort and recover the optically marked cells (with 90% recovery and near 100% purity), followed by genetic analysis. PMID:27118210

  6. Optical painting and fluorescence activated sorting of single adherent cells labelled with photoswitchable Pdots

    PubMed Central

    Kuo, Chun-Ting; Thompson, Alison M.; Gallina, Maria Elena; Ye, Fangmao; Johnson, Eleanor S.; Sun, Wei; Zhao, Mengxia; Yu, Jiangbo; Wu, I-Che; Fujimoto, Bryant; DuFort, Christopher C.; Carlson, Markus A.; Hingorani, Sunil R.; Paguirigan, Amy L.; Radich, Jerald P.; Chiu, Daniel T.

    2016-01-01

    The efficient selection and isolation of individual cells of interest from a mixed population is desired in many biomedical and clinical applications. Here we show the concept of using photoswitchable semiconducting polymer dots (Pdots) as an optical ‘painting' tool, which enables the selection of certain adherent cells based on their fluorescence, and their spatial and morphological features, under a microscope. We first develop a Pdot that can switch between the bright (ON) and dark (OFF) states reversibly with a 150-fold contrast ratio on irradiation with ultraviolet or red light. With a focused 633-nm laser beam that acts as a ‘paintbrush' and the photoswitchable Pdots as the ‘paint', we select and ‘paint' individual Pdot-labelled adherent cells by turning on their fluorescence, then proceed to sort and recover the optically marked cells (with 90% recovery and near 100% purity), followed by genetic analysis. PMID:27118210

  7. Comparative study of the radiobiological effects induced on adherent vs suspended cells by BNCT, neutrons and gamma rays treatments.

    PubMed

    Cansolino, L; Clerici, A M; Zonta, C; Dionigi, P; Mazzini, G; Di Liberto, R; Altieri, S; Ballarini, F; Bortolussi, S; Carante, M P; Ferrari, M; González, S J; Postuma, I; Protti, N; Santa Cruz, G A; Ferrari, C

    2015-12-01

    The present work is part of a preclinical in vitro study to assess the efficacy of BNCT applied to liver or lung coloncarcinoma metastases and to limb osteosarcoma. Adherent growing cell lines can be irradiated as adherent to the culture flasks or as cell suspensions, differences in radio-sensitivity of the two modalities of radiation exposure have been investigated. Dose related cell survival and cell cycle perturbation results evidenced that the radiosensitivity of adherent cells is higher than that of the suspended ones. PMID:26256647

  8. Both enzymatic and non-enzymatic properties of heat-labile enterotoxin are responsible for LT-enhanced adherence of enterotoxigenic Escherichia coli to porcine IPEC-J2 cells.

    PubMed

    Fekete, Peter Z; Mateo, Kristina S; Zhang, Weiping; Moxley, Rodney A; Kaushik, Radhey S; Francis, David H

    2013-06-28

    Previous studies in piglets indicate that heat labile enterotoxin (LT) expression enhances intestinal colonization by K88 adhesin-producing enterotoxigenic Escherichia coli (ETEC) as wild-type ETEC adhered to intestinal epithelium in substantially greater numbers than did non-toxigenic constructs. Enzymatic activity of the toxin was also shown to contribute to the adhesion of ETEC and non-ETEC bacteria to epithelial cells in culture. To further characterize the contribution of LT to host cell adhesion, a nontoxigenic, K88-producing E. coli was transformed with either the gene encoding for LT holotoxin, a catalytically-attenuated form of the toxin [LT(R192G)], or LTB subunits, and resultant changes in bacterial adherence to IPEC-J2 porcine intestinal epithelial cells were measured. Strains expressing LT holotoxin or mutants were able to adhere in significantly higher numbers to IPEC-J2 cells than was an isogenic, toxin-negative construct. LT+ strains were also able to significantly block binding of a wild-type LT+ ETEC strain to IPEC-J2 cells. Adherence of isogenic strains to IPEC-J2 cells was unaltered by cycloheximide treatment, suggesting that LT enhances ETEC adherence to IPEC-J2 cells independent of host cell protein synthesis. However, pretreating IPEC-J2 cells with LT promoted adherence of negatively charged latex beads (a surrogate for bacteria which carry a negative change), which adherence was inhibited by cycloheximide, suggesting LT may induce a change in epithelial cell membrane potential. Overall, these data suggest that LT may enhance ETEC adherence by promoting an association between LTB and epithelial cells, and by altering the surface charge of the host plasma membrane to promote non-specific adherence. PMID:23517763

  9. Protective effects of osmolytes in cryopreserving adherent neuroblastoma (Neuro-2a) cells.

    PubMed

    Bailey, Trisha L; Wang, Mian; Solocinski, Jason; Nathan, Britto P; Chakraborty, Nilay; Menze, Michael A

    2015-12-01

    A simple method to cryopreserve adherent monolayers of neuronal cells is currently not available, but the development of this technique could facilitate numerous applications in the field of biomedical engineering, cell line development, and drug screening. However, complex tissues of some exceptional animals survive freezing in nature. These animals are known to accumulate several small molecular weight solutes prior to freezing. Following a similar strategy, we investigated the effects of osmolytes such as trehalose, proline, and sucrose as additives to the traditional cryoprotectant dimethyl sulfoxide (Me2SO) in modulating the cryopreservation outcome of mouse neuroblastoma (Neuro-2a) cells. Neuro-2a cells adhered to cell culture plates were incubated for 24 h at varying concentrations of trehalose, proline, sucrose and combinations of these compounds. Cells were cryopreserved for 24 h and cell viability post-freezing and thawing was quantified by trypan blue exclusion assay. On average, only 13.5% of adherent cells survived freezing in the presence of 10% Me2SO alone (control). Pre-incubation of cells with medium containing both trehalose and proline severely decreased cell proliferation, but increased cell recovery to about 53% of control. Furthermore, characterization using Raman microspectroscopy revealed that the addition of both trehalose and proline to 10% Me2SO substantially increased the size, and altered the nature, of ice crystals formed during freezing. Our results suggest that pre-incubation of Neuro-2a cells with trehalose and proline in combination provides cell protection along with alterations of ice structure in order to increase cell survival post-freezing. PMID:26408850

  10. Fibrinogen-Induced Streptococcus mutans Biofilm Formation and Adherence to Endothelial Cells

    PubMed Central

    Lombardo Bedran, Telma Blanca; Azelmat, Jabrane; Palomari Spolidorio, Denise

    2013-01-01

    Streptococcus mutans, the predominant bacterial species associated with dental caries, can enter the bloodstream and cause infective endocarditis. The aim of this study was to investigate S. mutans biofilm formation and adherence to endothelial cells induced by human fibrinogen. The putative mechanism by which biofilm formation is induced as well as the impact of fibrinogen on S. mutans resistance to penicillin was also evaluated. Bovine plasma dose dependently induced biofilm formation by S. mutans. Of the various plasma proteins tested, only fibrinogen promoted the formation of biofilm in a dose-dependent manner. Scanning electron microscopy observations revealed the presence of complex aggregates of bacterial cells firmly attached to the polystyrene support. S. mutans in biofilms induced by the presence of fibrinogen was markedly resistant to the bactericidal effect of penicillin. Fibrinogen also significantly increased the adherence of S. mutans to endothelial cells. Neither S. mutans cells nor culture supernatants converted fibrinogen into fibrin. However, fibrinogen is specifically bound to the cell surface of S. mutans and may act as a bridging molecule to mediate biofilm formation. In conclusion, our study identified a new mechanism promoting S. mutans biofilm formation and adherence to endothelial cells which may contribute to infective endocarditis. PMID:24222906

  11. Sortase inhibitor phenyl vinyl sulfone inhibits Renibacterium salmoninarum adherence and invasion of host cells.

    PubMed

    Sudheesh, Ponnerassery S; Crane, Samuel; Cain, Kenneth D; Strom, Mark S

    2007-12-13

    Renibacterium salmoninarum, the causative agent of bacterial kidney disease in salmonid fishes, is a Gram-positive diplococcobacillus belonging to the family Micrococcaceae. Analysis of the genome sequence of the bacterium demonstrated the presence of a sortase homolog (srtD), a gene specifying an enzyme found in Gram-positive bacteria and required for covalent anchoring of cell surface proteins. Interference of sortase activity is being examined as a target for therapeutic prevention of infection by several pathogenic Gram-positive bacterial species. In silico analysis identified 8 open reading frames containing sortase recognition motifs, suggesting these proteins are translocated to the bacterial cell wall. The sortase and potential sortase substrate genes are transcribed in R. salmoninarum, suggesting they encode functional proteins. Treatment of R. salmoninarum with phenyl vinyl sulfone (PVS) significantly reduced bacterial adherence to Chinook salmon fibronectin. In addition, the ability of the PVS-treated bacteria to adhere to Chinook salmon embryo cells (CHSE-214) in vitro was dramatically reduced compared to that of untreated bacteria. More importantly, PVS-treated bacteria were unable to invade and replicate within CHSE-214 cells (demonstrated by an intracellular growth assay and by light microscopy). When treated with PVS, R. salmoninarum was not cytopathic to CHSE-214 cells, whereas untreated bacteria produced cytopathology within a few days. These findings clearly show that PVS, a small molecule drug and a known sortase inhibitor, can interfere with the ability of R. salmoninarum to adhere and colonize fish cells, with a corresponding decrease in virulence. PMID:18286808

  12. Pseudomonas cepacia adherence to respiratory epithelial cells is enhanced by Pseudomonas aeruginosa

    SciTech Connect

    Saiman, L.; Cacalano, G.; Prince, A. )

    1990-08-01

    Pseudomonas aeruginosa and Pseudomonas cepacia are both opportunistic pathogens of patients with cystic fibrosis. The binding characteristics of these two species were compared to determine if they use similar mechanisms to adhere to respiratory epithelial cells. P. cepacia 249 was shown to be piliated, but there was no detectable homology between P. aeruginosa pilin gene probes and P. cepacia genomic DNA. P. cepacia and P. aeruginosa did not appear to compete for epithelial receptors. In the presence of purified P. aeruginosa pili, the adherence of 35S-labeled strain 249 to respiratory epithelial monolayers was unaffected, while that of P. aeruginosa PAO1 was decreased by 55%. The binding of P. cepacia 249 and 715j was increased by 2.4-fold and 1.5-fold, respectively, in the presence of an equal inoculum of PAO1. Interbacterial agglutination contributed to the increased adherence of P. cepacia, as the binding of 249 was increased twofold in the presence of irradiated PAO1. PAO1 exoproducts had a marked effect in enhancing the ability of the P. cepacia strains to adhere to the epithelial monolayers. A PAO1 supernatant increased the binding of 249 by eightfold and that of 715j by fourfold. Thus, there appears to be a synergistic relationship between P. aeruginosa and P. cepacia in which PAO1 exoproducts modify the epithelial cell surface, exposing receptors and facilitating increased P. cepacia attachment.

  13. Associations between adherence and outcomes among older, type 2 diabetes patients: evidence from a Medicare Supplemental database

    PubMed Central

    Boye, Kristina Secnik; Curtis, Sarah E; Lage, Maureen J; Garcia-Perez, Luis-Emilio

    2016-01-01

    Objective To examine the association between adherence to glucose-lowering agents and patient outcomes, including costs, acute-care resource utilization, and complications, in an older, type 2 diabetic population. Data and methods The study used Truven’s Medicare Supplemental database from July 1, 2009 to June 30, 2014. Patients aged 65 years or older were included if they had at least two type 2 diabetes diagnoses and received a glucose-lowering agent from July 1, 2010 through June 30, 2011. Multivariable analyses examined the relationships among 3-year patient outcomes and levels of adherence, proxied by the proportion of days covered. Outcomes included all-cause medical costs, diabetes-related medical costs, acute-care resource utilization, and acute complications. Results In this study (N=123,235), higher adherence was linked to reduced costs and improved health outcomes. For example, comparing an individual with adherence of proportion of days covered <20% to one with proportion of days covered ≥80% illustrates an average saving of $28,824 in total 3-year costs. Furthermore, a 1% increase in adherence among 1,000 patients was associated with all-cause savings of $65,464 over 3 years. The probability of a hospitalization, an emergency room (ER) visit, or an acute complication decreased monotonically as adherence levels got higher, as did the number of hospitalizations, ER visits, and days hospitalized (P<0.005). Conclusion Higher adherence was associated with substantially less need for acute care, as indicated by a lowered probability of hospitalization or ER use, a reduced risk of an acute complication, and a decreased number of hospitalizations, ER visits, and days hospitalized. Higher adherence was also generally associated with lower all-cause and diabetes-related total costs, despite higher drug costs. These lower total costs were driven by the diminished acute care and outpatient costs. Results suggest that higher glucose-lowering agent adherence is

  14. Manipulation of a quasi-natural cell block for high-efficiency transplantation of adherent somatic cells.

    PubMed

    Chung, H J; Hassan, M M; Park, J O; Kim, H J; Hong, S T

    2015-05-01

    Recent advances have raised hope that transplantation of adherent somatic cells could provide dramatic new therapies for various diseases. However, current methods for transplanting adherent somatic cells are not efficient enough for therapeutic applications. Here, we report the development of a novel method to generate quasi-natural cell blocks for high-efficiency transplantation of adherent somatic cells. The blocks were created by providing a unique environment in which cultured cells generated their own extracellular matrix. Initially, stromal cells isolated from mice were expanded in vitro in liquid cell culture medium followed by transferring the cells into a hydrogel shell. After incubation for 1 day with mechanical agitation, the encapsulated cell mass was perforated with a thin needle and then incubated for an additional 6 days to form a quasi-natural cell block. Allograft transplantation of the cell block into C57BL/6 mice resulted in perfect adaptation of the allograft and complete integration into the tissue of the recipient. This method could be widely applied for repairing damaged cells or tissues, stem cell transplantation, ex vivo gene therapy, or plastic surgery. PMID:25742639

  15. Manipulation of a quasi-natural cell block for high-efficiency transplantation of adherent somatic cells

    PubMed Central

    Chung, H.J.; Hassan, M.M.; Park, J.O.; Kim, H.J.; Hong, S.T.

    2015-01-01

    Recent advances have raised hope that transplantation of adherent somatic cells could provide dramatic new therapies for various diseases. However, current methods for transplanting adherent somatic cells are not efficient enough for therapeutic applications. Here, we report the development of a novel method to generate quasi-natural cell blocks for high-efficiency transplantation of adherent somatic cells. The blocks were created by providing a unique environment in which cultured cells generated their own extracellular matrix. Initially, stromal cells isolated from mice were expanded in vitro in liquid cell culture medium followed by transferring the cells into a hydrogel shell. After incubation for 1 day with mechanical agitation, the encapsulated cell mass was perforated with a thin needle and then incubated for an additional 6 days to form a quasi-natural cell block. Allograft transplantation of the cell block into C57BL/6 mice resulted in perfect adaptation of the allograft and complete integration into the tissue of the recipient. This method could be widely applied for repairing damaged cells or tissues, stem cell transplantation, ex vivo gene therapy, or plastic surgery. PMID:25742639

  16. An in vitro clonal assay of adherent stem cells (ASC) in mouse marrow.

    PubMed

    Reincke, U; Rosenblatt, M; Hellman, S

    1984-11-01

    Hematopoietic stem cells with high proliferative capacity can be assayed when stromal bone marrow cultures are overlaid with limiting dilutions of marrow samples. This leads to hematopoietic growth after 4 weeks in a fraction of cultures, consistent with expectations based on Poisson statistics. It will be shown that monoclonal cultures are obtained that last from 2 to 15 weeks and that can generate up to several million mature granulocytes. The originating clone-forming cell is named adherent stem cell (ASC) because of its adherence to plastic or stromal surfaces. The ASC is comparable to the CFU-S in frequency, proliferative capacity and in its ability to give rise to CFU-S. As an unexpected additional finding we report that a mode of "clonal succession" was apparent in cultures which expressed more than one clone. PMID:6490726

  17. Mycoplasma pulmonis Vsa proteins and polysaccharide modulate adherence to pulmonary epithelial cells.

    PubMed

    Bolland, Jeffrey R; Dybvig, Kevin

    2012-06-01

    The Mycoplasma pulmonis Vsa proteins are a family of size- and phase-variable lipoproteins that shield the mycoplasmas from complement and modulate attachment to abiotic surfaces. Mycoplasmas producing a long Vsa protein hemadsorb poorly and yet are proficient at colonizing rats and mice. The effect of the length of the Vsa protein on the attachment of mycoplasmas to epithelial cells has not been previously explored. We find that independent of Vsa isotype, mycoplasmas producing a long Vsa protein with many tandem repeats adhere poorly to murine MLE-12 cells compared with mycoplasmas producing a short Vsa. We also find that mutants lacking the EPS-I polysaccharide of M. pulmonis exhibited decreased adherence to MLE-12 cells, even though it has been shown previously that such mutants have an enhanced ability to form a biofilm. PMID:22428866

  18. A mannose-specific adherence mechanism in Lactobacillus plantarum conferring binding to the human colonic cell line HT-29.

    PubMed Central

    Adlerberth, I; Ahrne, S; Johansson, M L; Molin, G; Hanson, L A; Wold, A E

    1996-01-01

    Two Lactobacillus plantarum strains of human intestinal origin, strains 299 (= DSM 6595) and 299v (= DSM 9843), have proved to be efficient colonizers of the human intestine under experimental conditions. These strains and 17 other L. plantarum strains were tested for the ability to adhere to cells of the human colonic cell line HT-29.L.plantarum 299 and 299v and nine other L. plantarum strains, including all six strains that belong to the same genetic subgroup as L. plantarum 299 and 299v, adhered to HT-29 cells in a manner that could be inhibited by methyl-alpha-D-mannoside. The ability to adhere to HT-29 cells correlated with an ability to agglutinate cells of Saccharomyces cerevisiae and erythrocytes in a mannose-sensitive manner and with adherence to D-mannose-coated agarose beads. L. plantarum 299 and 299v adhered to freshly isolated human colonic and ileal enterocytes, but the binding was not significantly inhibited by methyl-alpha-D-mannoside. Periodate treatment of HT-29 cells abolished mannose-sensitive adherence, confirming that the cell-bound receptor was of carbohydrate nature. Proteinase K treatment of the bacteria also abolished adherence, indicating that the binding involved protein structures on the bacterial cell surface. Thus, a mannose-specific adhesin has been identified in L. plantarum; this adhesin could be involved in the ability to colonize the intestine. PMID:8779562

  19. Catechin-based procyanidins from Peumus boldus Mol. aqueous extract inhibit Helicobacter pylori urease and adherence to adenocarcinoma gastric cells.

    PubMed

    Pastene, Edgar; Parada, Víctor; Avello, Marcia; Ruiz, Antonieta; García, Apolinaria

    2014-11-01

    In this work, the anti-Helicobacter pylori effect of an aqueous extract from dried leaves of Peumus boldus Mol. (Monimiaceae) was evaluated. This extract displayed high inhibitory activity against H. pylori urease. Therefore, in order to clarify the type of substances responsible for such effect, a bioassay-guided fractionation strategy was carried out. The active compounds in the fractions were characterized through different chromatographic methods (RP-HPLC; HILIC-HPLC). The fraction named F5 (mDP = 7.8) from aqueous extract was the most active against H. pylori urease with an IC50  = 15.9 µg gallic acid equivalents (GAE)/mL. HPLC analysis evidenced that F5 was composed mainly by catechin-derived proanthocyanidins (LC-MS and phloroglucinolysis). The anti-adherent effect of boldo was assessed by co-culture of H. pylori and AGS cells. Both the aqueous extract and F5 showed an anti-adherent effect in a concentration-dependent manner. An 89.3% of inhibition was reached at 2.0 mg GAE/mL of boldo extract. In conjunction, our results suggest that boldo extract has a potent anti-urease activity and anti-adherent effect against H. pylori, properties directly linked with the presence of catechin-derived proanthocyanidins. PMID:24853276

  20. Establishment and characterization of a novel, spontaneously immortalized retinoblastoma cell line with adherent growth.

    PubMed

    Kim, Jeong Hun; Kim, Jin Hyoung; Yu, Young Suk; Kim, Dong Hun; Kim, Chong Jai; Kim, Kyu-Won

    2007-09-01

    Retinoblastoma is the most common intraocular cancer of childhood, however, only a few cultured retinoblastoma cell lines are available to date. In the present study, we established a new human retinoblastoma cell line with adherent growth, named SNUOT-Rb1. The SNUOT-Rb1 cell line was established from an eye with retinoblastoma, which was enucleated from a 3-year-old Korean child. SNUOT-Rb1 has morphological and biochemical characteristics common to previous human retinoblastoma cell line, Y79: morphological features of fibroblast- or ganglion-like cells, and biochemical features of expression of glial fibrillary acidic protein and neuron-specific enolase. However, compared to Y79, SNUOT-Rb1 has a unique characteristic of growing in adherence, and the doubling time of SNUOT-Rb1 is shorter than Y79 in adherent or floating growth. In analysis of the tumorigenic potential of SNUOT-Rb1 in nude mice, orthotopic implantation of SNUOT-Rb1 mimics the pattern of local growth of retinoblastoma. In comparative genomic hybridization analysis, we found that SNUOT-Rb1 has significant chromosomal imbalances on chromosome 3, 9, 10, 11, 14, 16, 17, and 22. Therefore, SNUOT-Rb1 could be useful in studying the biological and genetic characteristics of retinoblastoma for insights into the heredity and genetics of childhood cancer. PMID:17671685

  1. Internet-Based Contingency Management to Improve Adherence with Blood Glucose Testing Recommendations for Teens with Type 1 Diabetes

    ERIC Educational Resources Information Center

    Raiff, Bethany R.; Dallery, Jesse

    2010-01-01

    The current study used Internet-based contingency management (CM) to increase adherence with blood glucose testing to at least 4 times daily. Four teens diagnosed with Type 1 diabetes earned vouchers for submitting blood glucose testing videos over a Web site. Participants submitted a mean of 1.7 and 3.1 blood glucose tests per day during the 2…

  2. Canine PHA-stimulated adherent cell enhance interferon-gamma production and proliferation of autologous peripheral blood mononuclear cells.

    PubMed

    Ide, Kaori; Momoi, Yasuyuki; Iwasaki, Toshiroh

    2005-03-01

    Dendritic cells are specialized antigen-presenting cells with immuno-modulating functions that are attractive for clinical applications for cancer immunotherapy. This study examined immunostimulatory functions of phytohemagglutinin (PHA)-stimulated adherent cells (PHA-Ad cells) from peripheral blood mononuclear cells (PBMCs) in dogs. PHA-Ad cells enhanced interferon-gamma from autologous PBMC in vitro. PHA-Ad cells also stimulated antigen-independent proliferation of peripheral blood lymphocytes. These results suggest that PHA-Ad cells from PBMC possess a stimulatory function to evoke anti-tumour immunity and that they demonstrate potential for therapeutic applications in dogs. PMID:19379211

  3. Relationship between cell surface composition of Candida albicans and adherence to acrylic after growth on different carbon sources.

    PubMed Central

    McCourtie, J; Douglas, L J

    1981-01-01

    The adherence of Candida albicans to acrylic was measured in vitro after growth of the yeast to stationary phase in defined medium containing glucose, sucrose, galactose, fructose, or maltose as the carbon source. In each case, yeast adherence was proportional to the concentration of sugar in the growth medium, but equimolar concentrations of different sugars promoted adherence to different extents. In vitro adherence was further increased by the addition of divalent cations to assay mixtures but was inhibited when saliva-treated acrylic strips were used or when yeasts were suspended in mixed saliva during the assay. The rate of spheroplast formation of yeasts grown in media containing a 500 mM concentration of the different sugars correlated well with the relative adherence of the cells to acrylic. Galactose-grown yeasts were most resistant to spheroplast formation with Zymolyase-5000 and most adherent to acrylic, whereas fructose-grown organisms were least resistant to spheroplast formation and least adherent to acrylic. These results indicate that when grown to stationary phase in media containing high concentrations of certain sugars, C. albicans undergoes a change in cell surface composition which facilitates its adherence to acrylic surfaces. Electron microscopy of yeasts harvested from such media revealed the presence of an additional surface layer which may be responsible for this enhanced adherence. Images PMID:7019091

  4. Increased neutrophil adherence and adhesion molecule mRNA expression in endothelial cells during selenium deficiency.

    PubMed

    Maddox, J F; Aherne, K M; Reddy, C C; Sordillo, L M

    1999-05-01

    Leukocyte aggregation and activation on endothelial cells (EC) are important preliminary events in leukocyte migration into tissue and subsequent inflammation. Thus, an increase in leukocyte adherence has the potential to affect inflammatory disease outcome. Selenium (Se) is an integral part of the antioxidant enzyme glutathione peroxidase (GSH-Px) and plays an important role in the maintenance of the redox state of a cell. Se supplementation in the bovine has been shown to improve the outcome of acute mastitis caused by coliform bacteria, in part by enhancing the speed of neutrophil migration into the affected mammary gland. However, the mechanisms by which Se modulates neutrophil migration have not been elucidated. Therefore, an in vitro model of Se deficiency in primary bovine mammary artery EC was used to examine the impact of Se status on the adhesive properties of EC. The effect of Se on functional activities was examined by measuring neutrophil adherence to Se-deficient and Se-supplemented EC. Se-deficient EC showed significantly enhanced neutrophil adherence when stimulated with tumor necrosis factor alpha (TNF-alpha) for 4 or 24 h, interleukin-1 for 12 h, or H2O2 for 20 min (P < 0.05). To determine the mechanisms underlying these changes in neutrophil adherence, the expression of EC adhesion molecules, ICAM-1, E-selectin, and P-selectin were examined at the molecular level by a competitive reverse transcription-polymerase chain reaction. Results revealed higher mRNA expression for E-selectin and ICAM-1 in Se-deficient EC stimulated with TNF-alpha for 3 and 6 h, and greater expression of P-selectin mRNA in Se-supplemented EC with 3-h TNF-alpha stimulation. These studies provide new information to establish the role of Se nutrition in the initiation of leukocyte adherence to endothelium. PMID:10331495

  5. Neurosphere and adherent culture conditions are equivalent for malignant glioma stem cell lines.

    PubMed

    Rahman, Maryam; Reyner, Karina; Deleyrolle, Loic; Millette, Sebastien; Azari, Hassan; Day, Bryan W; Stringer, Brett W; Boyd, Andrew W; Johns, Terrance G; Blot, Vincent; Duggal, Rohit; Reynolds, Brent A

    2015-03-01

    Certain limitations of the neurosphere assay (NSA) have resulted in a search for alternative culture techniques for brain tumor-initiating cells (TICs). Recently, reports have described growing glioblastoma (GBM) TICs as a monolayer using laminin. We performed a side-by-side analysis of the NSA and laminin (adherent) culture conditions to compare the growth and expansion of GBM TICs. GBM cells were grown using the NSA and adherent culture conditions. Comparisons were made using growth in culture, apoptosis assays, protein expression, limiting dilution clonal frequency assay, genetic affymetrix analysis, and tumorigenicity in vivo. In vitro expansion curves for the NSA and adherent culture conditions were virtually identical (P=0.24) and the clonogenic frequencies (5.2% for NSA vs. 5.0% for laminin, P=0.9) were similar as well. Likewise, markers of differentiation (glial fibrillary acidic protein and beta tubulin III) and proliferation (Ki67 and MCM2) revealed no statistical difference between the sphere and attachment methods. Several different methods were used to determine the numbers of dead or dying cells (trypan blue, DiIC, caspase-3, and annexin V) with none of the assays noting a meaningful variance between the two methods. In addition, genetic expression analysis with microarrays revealed no significant differences between the two groups. Finally, glioma cells derived from both methods of expansion formed large invasive tumors exhibiting GBM features when implanted in immune-compromised animals. A detailed functional, protein and genetic characterization of human GBM cells cultured in serum-free defined conditions demonstrated no statistically meaningful differences when grown using sphere (NSA) or adherent conditions. Hence, both methods are functionally equivalent and remain suitable options for expanding primary high-grade gliomas in tissue culture. PMID:25806119

  6. Poor medication adherence in type 2 diabetes: recognizing the scope of the problem and its key contributors

    PubMed Central

    Polonsky, William H; Henry, Robert R

    2016-01-01

    At least 45% of patients with type 2 diabetes (T2D) fail to achieve adequate glycemic control (HbA1c <7%). One of the major contributing factors is poor medication adherence. Poor medication adherence in T2D is well documented to be very common and is associated with inadequate glycemic control; increased morbidity and mortality; and increased costs of outpatient care, emergency room visits, hospitalization, and managing complications of diabetes. Poor medication adherence is linked to key nonpatient factors (eg, lack of integrated care in many health care systems and clinical inertia among health care professionals), patient demographic factors (eg, young age, low education level, and low income level), critical patient beliefs about their medications (eg, perceived treatment inefficacy), and perceived patient burden regarding obtaining and taking their medications (eg, treatment complexity, out-of-pocket costs, and hypoglycemia). Specific barriers to medication adherence in T2D, especially those that are potentially modifiable, need to be more clearly identified; strategies that target poor adherence should focus on reducing medication burden and addressing negative medication beliefs of patients. Solutions to these problems would require behavioral innovations as well as new methods and modes of drug delivery. PMID:27524885

  7. Temperature-induced labelling of Fluo-3 AM selectively yields brighter nucleus in adherent cells

    SciTech Connect

    Meng, Guixian; Pan, Leiting; Li, Cunbo; Hu, Fen; Shi, Xuechen; Lee, Imshik; Drevenšek-Olenik, Irena; Zhang, Xinzheng; Xu, Jingjun

    2014-01-17

    Highlights: •We detailedly examine temperature effects of Fluo-3 AM labelling in adherent cells. •4 °C Loading and 20 °C de-esterification of Fluo-3 AM yields brighter nuclei. •Brighter nuclei labelling by Fluo-3 AM also depends on cell adhesion quality. •A qualitative model of the brighter nucleus is proposed. -- Abstract: Fluo-3 is widely used to study cell calcium. Two traditional approaches: (1) direct injection and (2) Fluo-3 acetoxymethyl ester (AM) loading, often bring conflicting results in cytoplasmic calcium ([Ca{sup 2+}]{sub c}) and nuclear calcium ([Ca{sup 2+}]{sub n}) imaging. AM loading usually yields a darker nucleus than in cytoplasm, while direct injection always induces a brighter nucleus which is more responsive to [Ca{sup 2+}]{sub n} detection. In this work, we detailedly investigated the effects of loading and de-esterification temperatures on the fluorescence intensity of Fluo-3 in response to [Ca{sup 2+}]{sub n} and [Ca{sup 2+}]{sub c} in adherent cells, including osteoblast, HeLa and BV2 cells. Interestingly, it showed that fluorescence intensity of nucleus in osteoblast cells was about two times larger than that of cytoplasm when cells were loaded with Fluo-3 AM at 4 °C and allowed a subsequent step for de-esterification at 20 °C. Brighter nuclei were also acquired in HeLa and BV2 cells using the same experimental condition. Furthermore, loading time and adhesion quality of cells had effect on fluorescence intensity. Taken together, cold loading and room temperature de-esterification treatment of Fluo-3 AM selectively yielded brighter nucleus in adherent cells.

  8. Upon impact: the fate of adhering Pseudomonas fluorescens cells during nanofiltration.

    PubMed

    Habimana, Olivier; Semião, Andrea J C; Casey, Eoin

    2014-08-19

    Nanofiltration (NF) is a high-pressure membrane filtration process increasingly applied in drinking water treatment and water reuse processes. NF typically rejects divalent salts, organic matter, and micropollutants. However, the efficiency of NF is adversely affected by membrane biofouling, during which microorganisms adhere to the membrane and proliferate to create a biofilm. Here we show that adhered Pseudomonas fluorescens cells under high permeate flux conditions are met with high fluid shear and convective fluxes at the membrane-liquid interface, resulting in their structural damage and collapse. These results were confirmed by fluorescent staining, flow cytometry, and scanning electron microscopy. This present study offers a "first-glimpse" of cell damage and death during the initial phases of bacterial adhesion to NF membranes and raises a key question about the role of this observed phenomena during early-stage biofilm formation under permeate flux and cross-flow conditions. PMID:25072514

  9. Adherence to a home-based exercise program and incidence of cardiovascular disease in type 2 diabetes patients.

    PubMed

    Shinji, S; Shigeru, M; Ryusei, U; Mitsuru, M; Shigehiro, K

    2007-10-01

    The aim of this study was to examine the association between adherence to a home-based exercise program and the incidence of cardiovascular disease (CVD) in patients with type 2 diabetes. We investigated 102 patients with type 2 diabetes aged 35 to 75 years, and followed them prospectively for 17.2 months. Before enrollment, all patients received a traditional exercise prescription. The exercise program consisted of a daily walking exercise at home for 20 - 30 minutes. Self-reported adherence to the exercise program and the incidence of CVD were confirmed by information obtained from telephone interviews. There were 38 dropouts among the patients in the exercise program. Dropouts were significantly younger than completers. The rate of obesity was significantly higher among the dropouts than among the completers. No differences were observed between the two groups for gender, history of CVD and other clinical characteristics. During the follow-up, we documented 8 new cases of CVD. The incidence of CVD during the follow-up was 1.56 percent among the program completers and 18.4 percent among the dropouts. Adherence to the home-based exercise was inversely related to the incidence of CVD (p < 0.01). These associations persisted after adjustment for age and other covariates. In conclusion, adherence to an exercise program is associated with a reduced incidence of CVD among patients with type 2 diabetes. PMID:17436204

  10. Non-invasive and non-destructive measurements of confluence in cultured adherent cell lines.

    PubMed

    Busschots, Steven; O'Toole, Sharon; O'Leary, John J; Stordal, Britta

    2015-01-01

    Many protocols used for measuring the growth of adherent monolayer cells in vitro are invasive, destructive and do not allow for the continued, undisturbed growth of cells within flasks. Protocols often use indirect methods for measuring proliferation. Microscopy techniques can analyse cell proliferation in a non-invasive or non-destructive manner but often use expensive equipment and software algorithms. In this method images of cells within flasks are captured by photographing under a standard inverted phase contract light microscope using a digital camera with a camera lens adaptor. Images are analysed for confluence using ImageJ freeware resulting in a measure of confluence known as an Area Fraction (AF) output. An example of the AF method in use on OVCAR8 and UPN251 cell lines is included. •Measurements of confluence from growing adherent cell lines in cell culture flasks is obtained in a non-invasive, non-destructive, label-free manner.•The technique is quick, affordable and eliminates sample manipulation.•The technique provides an objective, consistent measure of when cells reach confluence and is highly correlated to manual counting with a haemocytometer. The average correlation co-efficient from a Spearman correlation (n = 3) was 0.99 ± 0.008 for OVCAR8 (p = 0.01) and 0.99 ± 0.01 for UPN251 (p = 0.01) cell lines. PMID:26150966

  11. Non-invasive and non-destructive measurements of confluence in cultured adherent cell lines

    PubMed Central

    Busschots, Steven; O’Toole, Sharon; O’Leary, John J.; Stordal, Britta

    2014-01-01

    Many protocols used for measuring the growth of adherent monolayer cells in vitro are invasive, destructive and do not allow for the continued, undisturbed growth of cells within flasks. Protocols often use indirect methods for measuring proliferation. Microscopy techniques can analyse cell proliferation in a non-invasive or non-destructive manner but often use expensive equipment and software algorithms. In this method images of cells within flasks are captured by photographing under a standard inverted phase contract light microscope using a digital camera with a camera lens adaptor. Images are analysed for confluence using ImageJ freeware resulting in a measure of confluence known as an Area Fraction (AF) output. An example of the AF method in use on OVCAR8 and UPN251 cell lines is included. • Measurements of confluence from growing adherent cell lines in cell culture flasks is obtained in a non-invasive, non-destructive, label-free manner. • The technique is quick, affordable and eliminates sample manipulation. • The technique provides an objective, consistent measure of when cells reach confluence and is highly correlated to manual counting with a haemocytometer. The average correlation co-efficient from a Spearman correlation (n = 3) was 0.99 ± 0.008 for OVCAR8 (p = 0.01) and 0.99 ± 0.01 for UPN251 (p = 0.01) cell lines. PMID:26150966

  12. Effects of mononuclear phagocyte system modulating agents on Fc and C3 receptors of adherent cells.

    PubMed Central

    Hitomi, M.; Shimizu, F.

    1985-01-01

    Agents which modulate the mononuclear phagocyte system (MPS) were examined for their effects on Fc and C3 receptors of adherent cells (A-cells) as judged by rosette formation. Dextran sulphate, carrageenan, and immune complexes, known as MPS suppressants, reduced the percentage of receptor-positive A-cells, while levamisole, known as a MPS-activator, increased the percentage in vitro. The changes in the percentage of Fc receptor were parallel to those of the C3 receptor in vitro. The effects of these agents were also examined in vivo. PMID:2408651

  13. The expression of nonagglutinating fimbriae and its role in Proteus mirabilis adherence to epithelial cells.

    PubMed

    Tolson, D L; Harrison, B A; Latta, R K; Lee, K K; Altman, E

    1997-08-01

    Proteus mirabilis is a common causative agent of human urinary tract infections, especially in catheterized patients and in those patients with structural abnormalities of the urinary tract. In addition to the production of hemolysin and urease, fimbriae-mediated adherence to uroepithelial cells and kidney epithelium may be essential for virulence of P. mirabilis. A single P. mirabilis strain is capable of expressing several morphologically distinct fimbrial species, which can each be favoured by specific in vitro growth conditions. The fimbrial species reported to date include mannose-resistant/Proteus-like fimbriae, ambient temperature fimbriae, P. mirabilis fimbriae, and nonagglutinating fimbriae (NAF). Here, using intact bacteria or purified NAF as immunogens, we have generated the first reported NAF-specific monoclonal antibodies (mAbs). Bacteria expressing NAF as their only fimbrial species adhered strongly to a number of cell lines in vitro, including uroepithelial cell lines. Binding of P. mirabilis was markedly reduced following preincubation with NAF-specific mAbs and Fab fragments. The presence of NAF with highly conserved N-terminal sequences on all P. mirabilis strains so far examined, combined with the ability of both anti-NAF mAbs and purified NAF molecules to inhibit P. mirabilis adherence in vitro, suggests that NAF may contribute to the pathogenesis of P. mirabilis. PMID:9304781

  14. Imaging deformation of adherent cells due to shear stress using quantitative phase imaging.

    PubMed

    Eldridge, Will J; Sheinfeld, Adi; Rinehart, Matthew T; Wax, Adam

    2016-01-15

    We present a platform for detecting cellular deformations from mechanical stimuli, such as fluid shear stress, using rapid quantitative phase imaging. Rapid quantitative phase imaging was used to analyze changes in the optical path length of adherent skin cancer cells during mechanical displacement. Both the whole-cell phase displacement and the resultant shift of the cellular center of mass were calculated over the duration of the stimulus. Whole-cell phase displacement images were found to match expectation. Furthermore, center-of-mass shifts of adherent cells were found to resemble that of a one-dimensional Kelvin-Voigt (KV) viscoelastic solid. Cellular steady-state displacements from step fluid shear stimuli were found to be linearly related to the shear stress. Shear stiffness constants for cells exposed to a cytoskeletal disrupting toxin were found to be significantly lower than unexposed cells. This novel technique allows for elastographic analysis of whole-cell effective shear stiffness without the use of an exogenous force applicator, a specialized culture substrate, or tracking net perimeter movement of the cell. PMID:26766712

  15. Array of Biodegradable Microraftsfor Isolation and Implantation of Living, Adherent Cells

    PubMed Central

    Wang, Yuli; Phillips, Colleen N.; Herrera, Gabriela S.; Sims, Christopher E.; Yeh, Jen Jen; Allbritton, Nancy L.

    2013-01-01

    A new strategy for efficient sorting and implantation of viable adherent cells into animals is described. An array of biodegradable micro-structures (microrafts) was fabricated using a polydimethylsiloxane substrate for micromolding poly(lactic-co-glycolic acid) (PLGA). Screening various forms of PLGA determined that the suitability of PLGA for microraft manufacture, biocompatibility and in vitro degradation was dependent on molecular weight and lactic/glycolic ratio. Cells plated on the array selectively attached to the microrafts and could be identified by their fluorescence, morphology or other criteria. The cells were efficiently dislodged and collected from the array using a microneedle device. The platform was used to isolate specific cells from a mixed population establishing the ability to sort target cells for direct implantation. As a proof of concept, fluorescently conjugated microrafts carrying tumor cells stably expressing luciferase were isolated from an array and implanted subcutaneously into mice. In vivo bio-luminescence imaging confirmed the growth of a tumor in the recipient animals. Imaging of tissue sections from the tumors demonstrated in vivo degradation of the implanted microrafts. The process is a new strategy for isolating and delivering a small number of adherent cells for animal implantation with potential applications in tissue repair, tumor induction, in vivo differentiation of stem cells and other biomedical research. PMID:23930219

  16. A visual targeting system for the microinjection of unstained adherent cells.

    PubMed

    Becattini, Gabriele; Mattos, Leonardo S; Caldwell, Darwin G

    2013-02-01

    Automatic localization and targeting are critical steps in automating the process of microinjecting adherent cells. This process is currently performed manually by highly trained operators and is characterized as a laborious task with low success rate. Therefore, automation is desired to increase the efficiency and consistency of the operations. This research offers a contribution to this procedure through the development of a vision system for a robotic microinjection setup. Its goals are to automatically locate adherent cells in a culture dish and target them for a microinjection. Here the major concern was the achievement of an error-free targeting system to guarantee high consistency in microinjection experiments. To accomplish this, a novel visual targeting algorithm integrating different image processing techniques was proposed. This framework employed defocusing microscopy to highlight cell features and improve cell segmentation and targeting reliability. Three main image processing techniques, operating at three different focus levels in a bright field (BF) microscope, were used: an anisotropic contour completion (ACC) method, a local intensity variation background-foreground classifier, and a grayscale threshold-based segmentation. The proposed framework combined information gathered by each of these methods using a validation map and this was shown to provide reliable cell targeting results. Experiments conducted with sets of real images from two different cell lines (CHO-K1 and HEK), which contained a total of more than 650 cells, yielded flawless targeting results along with a cell detection ratio greater than 50%. PMID:23287416

  17. Immune adherence in renal glomeruli. Complement receptor sites on glomerular capillary epithelial cells.

    PubMed Central

    Burkholder, P. M.; Oberley, T. D.; Barber, T. A.; Beacom, A.; Koehler, C.

    1977-01-01

    Several very recent reports have indicated the presence of receptor sites for the third component of complement in human but not other vertebrate renal glomeruli. The present study constitutes a demonstration that the glomerular capillary epithelial cell bears this receptor, detectable with either EAC complexes (EAC1423b) or fluores ceinated zymosan-C3 (ZC3b) complexes, Fresh, unfixed frozen sections of normal or diseased human kidneys, mechanically isolated human glomeruli, dissociated glomerular cells, and glomeruli and golmerular cells maintained in tissue culture were examined with various EAC complexes or ZC3b and examined by phase light microscopy, fluorescence microscopy, or transmission and scanning electron microscopy. Clearly, by scanning electron microscopy it was determined that glomerular capillary epithelial cells bind the immune-adherence EAC indicator cells. Because glomeruli or glomerular epithelial cells did not bind E, EA, EACI, EAC14, or EAC142 but did bind EAC1423b or ZC3b, it is concluded that C3b (activated bound fragment of the third component of complement) is responsible for the immune-adherence reaction in glomeruli. Preliminary examination of diseased renal biopsies indicates that sclerotic glomeruli, focal segmental sclerotic or proliferative glomerular capillary lesions, and proliferative epithelial crescents are immune-adherence negative. Furthermore, a clear or consistent inverse relationship between glomerular capillary deposits of C3 which presumably might block epithelial C3 receptor sites, and immune-adherence reactivity with EAC in vitro was not as evident in this study as reported previously by other investigators. Nevertheless, it is still attractive to conceive that glomerular C3 receptor sites might be responsible for binding of antigen-antibody-complement complexes and formation of immune-complex deposits, at least on the epimembranous (subepithelial) surface of glomerular capillary walls. Inability to demonstrate this

  18. Predicting Noninsulin Antidiabetic Drug Adherence Using a Theoretical Framework Based on the Theory of Planned Behavior in Adults With Type 2 Diabetes: A Prospective Study.

    PubMed

    Zomahoun, Hervé Tchala Vignon; Moisan, Jocelyne; Lauzier, Sophie; Guillaumie, Laurence; Grégoire, Jean-Pierre; Guénette, Line

    2016-04-01

    Understanding the process behind noninsulin antidiabetic drug (NIAD) nonadherence is necessary for designing effective interventions to resolve this problem. This study aimed to explore the ability of the theory of planned behavior (TPB), which is known as a good predictor of behaviors, to predict the future NIAD adherence in adults with type 2 diabetes. We conducted a prospective study of adults with type 2 diabetes. They completed a questionnaire on TPB variables and external variables. Linear regression was used to explore the TPB's ability to predict future NIAD adherence, which was prospectively measured as the proportion of days covered by at least 1 NIAD using pharmacy claims data. The interaction between past NIAD adherence and intention was tested. The sample included 340 people. There was an interaction between past NIAD adherence and intention to adhere to the NIAD (P = 0.032). Intention did not predict future NIAD adherence in the past adherers and nonadherers groups, but its association measure was high among past nonadherers (β = 5.686, 95% confidence interval [CI] -10.174, 21.546). In contrast, intention was mainly predicted by perceived behavioral control both in the past adherers (β = 0.900, 95% CI 0.796, 1.004) and nonadherers groups (β = 0.760, 95% CI 0.555, 0.966). The present study suggests that TPB is a good tool to predict intention to adhere and future NIAD adherence. However, there was a gap between intention to adhere and actual adherence to the NIAD, which is partly explained by the past adherence level in adults with type 2 diabetes. PMID:27082543

  19. Predicting Noninsulin Antidiabetic Drug Adherence Using a Theoretical Framework Based on the Theory of Planned Behavior in Adults With Type 2 Diabetes

    PubMed Central

    Zomahoun, Hervé Tchala Vignon; Moisan, Jocelyne; Lauzier, Sophie; Guillaumie, Laurence; Grégoire, Jean-Pierre; Guénette, Line

    2016-01-01

    Abstract Understanding the process behind noninsulin antidiabetic drug (NIAD) nonadherence is necessary for designing effective interventions to resolve this problem. This study aimed to explore the ability of the theory of planned behavior (TPB), which is known as a good predictor of behaviors, to predict the future NIAD adherence in adults with type 2 diabetes. We conducted a prospective study of adults with type 2 diabetes. They completed a questionnaire on TPB variables and external variables. Linear regression was used to explore the TPB's ability to predict future NIAD adherence, which was prospectively measured as the proportion of days covered by at least 1 NIAD using pharmacy claims data. The interaction between past NIAD adherence and intention was tested. The sample included 340 people. There was an interaction between past NIAD adherence and intention to adhere to the NIAD (P = 0.032). Intention did not predict future NIAD adherence in the past adherers and nonadherers groups, but its association measure was high among past nonadherers (β = 5.686, 95% confidence interval [CI] −10.174, 21.546). In contrast, intention was mainly predicted by perceived behavioral control both in the past adherers (β = 0.900, 95% CI 0.796, 1.004) and nonadherers groups (β = 0.760, 95% CI 0.555, 0.966). The present study suggests that TPB is a good tool to predict intention to adhere and future NIAD adherence. However, there was a gap between intention to adhere and actual adherence to the NIAD, which is partly explained by the past adherence level in adults with type 2 diabetes. PMID:27082543

  20. Towards high-throughput automated targeted femtosecond laser-based transfection of adherent cells

    NASA Astrophysics Data System (ADS)

    Antkowiak, Maciej; Torres-Mapa, Maria Leilani; Gunn-Moore, Frank; Dholakia, Kishan

    2011-03-01

    Femtosecond laser induced cell membrane poration has proven to be an attractive alternative to the classical methods of drug and gene delivery. It is a selective, sterile, non-contact technique that offers a highly localized operation, low toxicity and consistent performance. However, its broader application still requires the development of robust, high-throughput and user-friendly systems. We present a system capable of unassisted enhanced targeted optoinjection and phototransfection of adherent mammalian cells with a femtosecond laser. We demonstrate the advantages of a dynamic diffractive optical element, namely a spatial light modulator (SLM) for precise three dimensional positioning of the beam. It enables the implementation of a "point-and-shoot" system in which using the software interface a user simply points at the cell and a predefined sequence of precisely positioned doses can be applied. We show that irradiation in three axial positions alleviates the problem of exact beam positioning on the cell membrane and doubles the number of viably optoinjected cells when compared with a single dose. The presented system enables untargeted raster scan irradiation which provides transfection of adherent cells at the throughput of 1 cell per second.

  1. Evaluation of Gelatin Microparticles as Adherent-Substrates for Mesenchymal Stem Cells in a Hydrogel Composite.

    PubMed

    Lu, Steven; Lee, Esther J; Lam, Johnny; Tabata, Yasuhiko; Mikos, Antonios G

    2016-06-01

    Due to the lack of cell-adhesive moieties in traditional synthetic hydrogels, the present work investigated the use of degradable gelatin microparticles (GMPs) as temporary adherent substrates for anchorage-dependent mesenchymal stem cells (MSCs). MSCs were seeded onto GMPs of varying crosslinking densities and sizes to investigate their role on influencing MSC differentiation and aggregation. The MSC-seeded GMPs were then encapsulated in poly(ethylene glycol)-based hydrogels and cultured in serum-free, growth factor-free osteochondral medium. Non-seeded MSCs co-encapsulated with GMPs in the hydrogels were used as a control for comparison. Over the course of 35 days, MSCs seeded on GMPs exhibited more cell-cell contacts, greater chondrogenic potential, and a down-regulation of osteogenic markers compared to the controls. Although the factors of GMP crosslinking and size had nominal influence on MSC differentiation and aggregation, GMPs demonstrate potential as an adherent-substrate for improving cell delivery from hydrogel scaffolds by facilitating cell-cell contacts and improving MSC differentiation. PMID:26935924

  2. Slow-Adhering Stem Cells Derived from Injured Skeletal Muscle Have Improved Regenerative Capacity

    PubMed Central

    Mu, Xiaodong; Xiang, Guosheng; Rathbone, Christopher R.; Pan, Haiying; Bellayr, Ian H.; Walters, Thomas J.; Li, Yong

    2011-01-01

    A wide variety of myogenic cell sources have been used for repair of injured and diseased muscle including muscle stem cells, which can be isolated from skeletal muscle as a group of slow-adhering cells on a collagen-coated surface. The therapeutic use of muscle stem cells for improving muscle regeneration is promising; however, the effect of injury on their characteristics and engraftment potential has yet to be described. In the present study, slow-adhering stem cells (SASCs) from both laceration-injured and control noninjured skeletal muscles in mice were isolated and studied. Migration and proliferation rates, multidifferentiation potentials, and differences in gene expression in both groups of cells were compared in vitro. Results demonstrated that a larger population of SASCs could be isolated from injured muscle than from control noninjured muscle. In addition, SASCs derived from injured muscle demonstrated improved migration, a higher rate of proliferation and multidifferentiation, and increased expression of Notch1, STAT3, Msx1, and MMP2. Moreover, when transplanted into dystrophic muscle in MDX/SCID mice, SASCs from injured muscle generated greater engraftments with a higher capillary density than did SASCs from control noninjured muscle. These data suggest that traumatic injury may modify stem cell characteristics through trophic factors and improve the transplantation potential of SASCs in alleviating skeletal muscle injuries and diseases. PMID:21684246

  3. Types of Stem Cells

    MedlinePlus

    ... PDF) Download an introduction to stem cells and stem cell research. Stem Cell Glossary Stem cell terms to know. ... stem cells blog from the International Society for Stem Cell Research. Learn About Stem Cells From Lab to You ...

  4. Applications of electroporation of adherent cells in situ, on a partly conductive slide.

    PubMed

    Raptis, L H; Brownell, H L; Liu, S K; Firth, K L; MacKenzie, L W; Stiles, C D; Alberta, J A

    1995-10-01

    Nontraumatic, simple, and reproducible procedures for the introduction of nonpermeant molecules into adherent mammalian cells by in situ electroporation are described. Cells are grown on a glass slide, half of which is coated with electrically conductive, optically transparent, indium-tin oxide. An electric pulse is applied in the presence of the molecules to be introduced, and their effect on the cellular phenotype can be observed. The cells growing on the nonconductive side of the slide do not receive any pulse and serve as controls. Careful adjustment of electric field strength can achieve the introduction of the molecules into essentially 100% of the cells, and this treatment causes no detectable disruption to cellular metabolism. This is applied in the presence of the fluorescent dye, Lucifer yellow, causing its penetration into the cells growing on the conductive half of the slide. The migration of the dye to the nonelectroporated cells growing on the nonconductive area is microscopically observed under fluorescence illumination. PMID:8556428

  5. Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) assay-Book Chapter*

    EPA Science Inventory

    There are thousands of environmental chemicals for which there is limited toxicological information, motivating the development and application of in vitro systems to profile the biological effects of xenobiotic exposure and predict their potential developmental hazard. An adhere...

  6. Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) Assay: Book Chapter

    EPA Science Inventory

    There are thousands of environmental chemicals for which there is limited toxicological information, motivating the development and application of in vitro systems to profile the biological effects of xenobiotic exposure and predict their potential developmental hazard. An adher...

  7. Cell Phone-Based and Adherence Device Technologies for HIV Care and Treatment in Resource-Limited Settings: Recent Advances.

    PubMed

    Campbell, Jeffrey I; Haberer, Jessica E

    2015-12-01

    Numerous cell phone-based and adherence monitoring technologies have been developed to address barriers to effective HIV prevention, testing, and treatment. Because most people living with HIV and AIDS reside in resource-limited settings (RLS), it is important to understand the development and use of these technologies in RLS. Recent research on cell phone-based technologies has focused on HIV education, linkage to and retention in care, disease tracking, and antiretroviral therapy adherence reminders. Advances in adherence devices have focused on real-time adherence monitors, which have been used for both antiretroviral therapy and pre-exposure prophylaxis. Real-time monitoring has recently been combined with cell phone-based technologies to create real-time adherence interventions using short message service (SMS). New developments in adherence technologies are exploring ingestion monitoring and metabolite detection to confirm adherence. This article provides an overview of recent advances in these two families of technologies and includes research on their acceptability and cost-effectiveness when available. It additionally outlines key challenges and needed research as use of these technologies continues to expand and evolve. PMID:26439917

  8. Let Visuals Tell the Story: Medication Adherence in Patients with Type II Diabetes Captured by a Novel Ingestion Sensor Platform

    PubMed Central

    Behzadi, Yashar; Littlewort, Gwen

    2015-01-01

    Background Chronic diseases such as diabetes require high levels of medication adherence and patient self-management for optimal health outcomes. A novel sensing platform, Digital Health Feedback System (Proteus Digital Health, Redwood City, CA), can for the first time detect medication ingestion events and physiological measures simultaneously, using an edible sensor, personal monitor patch, and paired mobile device. The Digital Health Feedback System (DHFS) generates a large amount of data. Visual analytics of this rich dataset may provide insights into longitudinal patterns of medication adherence in the natural setting and potential relationships between medication adherence and physiological measures that were previously unknown. Objective Our aim was to use modern methods of visual analytics to represent continuous and discrete data from the DHFS, plotting multiple different data types simultaneously to evaluate the potential of the DHFS to capture longitudinal patterns of medication-taking behavior and self-management in individual patients with type II diabetes. Methods Visualizations were generated using time domain methods of oral metformin medication adherence and physiological data obtained by the DHFS use in 5 patients with type II diabetes over 37-42 days. The DHFS captured at-home metformin adherence, heart rate, activity, and sleep/rest. A mobile glucose monitor captured glucose testing and level (mg/dl). Algorithms were developed to analyze data over varying time periods: across the entire study, daily, and weekly. Following visualization analysis, correlations between sleep/rest and medication ingestion were calculated across all subjects. Results A total of 197 subject days, encompassing 141,840 data events were analyzed. Individual continuous patch use varied between 87-98%. On average, the cohort took 78% (SD 12) of prescribed medication and took 77% (SD 26) within the prescribed ±2-hour time window. Average activity levels per subjects ranged

  9. Group A Streptococcus Adheres to Pharyngeal Epithelial Cells with Salivary Proline-rich Proteins via GrpE Chaperone Protein*

    PubMed Central

    Murakami, Jumpei; Terao, Yutaka; Morisaki, Ichijiro; Hamada, Shigeyuki; Kawabata, Shigetada

    2012-01-01

    Group A Streptococcus pyogenes (GAS) is an important human pathogen that frequently causes pharyngitis. GAS organisms can adhere to and invade pharyngeal epithelial cells, which are overlaid by salivary components. However, the role of salivary components in GAS adhesion to pharyngeal cells has not been reported precisely. We collected human saliva and purified various salivary components, including proline-rich protein (PRP), statherin, and amylase, and performed invasion assays. The GAS-HEp-2 association ratio (invasion/adhesion ratio) and invasion ratio of GAS were increased significantly with whole human saliva and PRP, while the anti-PRP antibody inhibited the latter. GAS strain NY-5, which lacks M and F proteins on the cell surface, was promoted to cohere with HEp-2 cells by whole human saliva and PRP. The 28-kDa protein of GAS bound to PRP and was identified as GrpE, a chaperone protein, whereas the N-terminal of GrpE was found to bind to PRP. A GrpE-deficient mutant of GAS strain B514Sm, TR-45, exhibited a reduced ability to adhere to and invade HEp-2 cells. Microscopic observations showed the GrpE was mainly expressed on the surface of the cell division site of GAS. Furthermore, GrpE-deficient mutants of GAS and Streptococcus pneumoniae showed an elongated morphology as compared with the wild type. Taken together, this is the first study to show an interaction between salivary PRP and GAS GrpE, which plays an important role in GAS infection on the pharynx, whereas the expression of GrpE on the surface of GAS helps to maintain morphology. PMID:22566698

  10. Measles virus hemagglutinin mediates monocyte aggregation and increased adherence to measles-infected endothelial cells.

    PubMed

    Soilu-Hänninen, M; Hänninen, A; Ilonen, J; Salmi, A; Salonen, R

    1996-09-01

    The effect of measles virus (MV) infection on monocyte adhesion was studied using human peripheral blood monocytes and monocytic and endothelial cell lines. The infection of monocytic U-937 cells led to the formation of large cellular aggregates. Aggregation was independent of intercellular adhesion molecule-1 (ICAM-1)/lymphocyte function-associated antigen-1 (LFA-1), but could be inhibited by monoclonal antibodies (mAb) against the MV hemagglutinin glycoprotein (MV-H). mAb against the MV receptor, CD46, also blocked aggregation. No significant changes in the cell surface expression of adhesion molecules CD11a, CD11b, CD11c, CD18, CD54, CD44, CD49d (alpha 4-integrin) and CD62L (L-selectin) were observed on MV-infected monocytes. Infection of a human endothelial cell line, EAhy 926 (HEC), with MV led to a two-fold increase in 1CAM-1 expression and a two-fold increase in monocyte adherence to the HEC (from 22 +/- 1.6% to 42 +/- 4.8%). However, ICAM-1 mAb reduced monocyte adhesion to the control and MV-infected HEC to a similar degree, whereas anti-MV-H antibodies abolished the difference between binding to infected and control HEC. We conclude that MV hemagglutinin mediated both the homo typic aggregation in infected monocyte cultures and increased monocyte adherence to the infected endothelial cells. PMID:8884738

  11. Enterotoxigenic Escherichia coli TibA Glycoprotein Adheres to Human Intestine Epithelial Cells

    PubMed Central

    Lindenthal, Christoph; Elsinghorst, Eric A.

    2001-01-01

    Enterotoxigenic Escherichia coli (ETEC) is capable of invading epithelial cell lines derived from the human ileum and colon. Two separate invasion loci (tia and tib) that direct noninvasive E. coli strains to adhere to and invade cultured human intestine epithelial cells have previously been isolated from the classical ETEC strain H10407. The tib locus directs the synthesis of TibA, a 104-kDa outer membrane glycoprotein. Synthesis of TibA is directly correlated with the adherence and invasion phenotypes of the tib locus, suggesting that this protein is an adhesin and invasin. Here we report the purification of TibA and characterization of its biological activity. TibA was purified by continuous-elution preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Purified TibA was biotin labeled and then shown to bind to HCT8 human ileocecal epithelial cells in a specific and saturable manner. Unlabeled TibA competed with biotin-labeled TibA, suggesting the presence of a specific TibA receptor in HCT8 cells. These results show that TibA acts as an adhesin. Polyclonal anti-TibA antiserum inhibited invasion of ETEC strain H10407 and of recombinant E. coli bearing tib locus clones, suggesting that TibA also acts as an invasin. The ability of TibA to direct epithelial cell adhesion suggests a role for this protein in ETEC pathogenesis. PMID:11119488

  12. Enterotoxigenic Escherichia coli TibA glycoprotein adheres to human intestine epithelial cells.

    PubMed

    Lindenthal, C; Elsinghorst, E A

    2001-01-01

    Enterotoxigenic Escherichia coli (ETEC) is capable of invading epithelial cell lines derived from the human ileum and colon. Two separate invasion loci (tia and tib) that direct noninvasive E. coli strains to adhere to and invade cultured human intestine epithelial cells have previously been isolated from the classical ETEC strain H10407. The tib locus directs the synthesis of TibA, a 104-kDa outer membrane glycoprotein. Synthesis of TibA is directly correlated with the adherence and invasion phenotypes of the tib locus, suggesting that this protein is an adhesin and invasin. Here we report the purification of TibA and characterization of its biological activity. TibA was purified by continuous-elution preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Purified TibA was biotin labeled and then shown to bind to HCT8 human ileocecal epithelial cells in a specific and saturable manner. Unlabeled TibA competed with biotin-labeled TibA, suggesting the presence of a specific TibA receptor in HCT8 cells. These results show that TibA acts as an adhesin. Polyclonal anti-TibA antiserum inhibited invasion of ETEC strain H10407 and of recombinant E. coli bearing tib locus clones, suggesting that TibA also acts as an invasin. The ability of TibA to direct epithelial cell adhesion suggests a role for this protein in ETEC pathogenesis. PMID:11119488

  13. Adherence to hydroxyurea medication by children with sickle cell disease (SCD) using an electronic device: a feasibility study.

    PubMed

    Inoue, Susumu; Kodjebacheva, Gergana; Scherrer, Tammy; Rice, Gary; Grigorian, Matthew; Blankenship, Jeremy; Onwuzurike, Nkechi

    2016-08-01

    Adherence to hydroxyurea (HU) is a significant modifying factor in sickle cell vaso-occlusive pain. We conducted a study using an electronic medication container-monitor-reminder device (GlowCap™) to track adherence and determine whether use of this device affected rates of HU adherence. Subjects were regular attendees to our clinic. They were given a 37-item questionnaire and were asked to use a GlowCap containing HU. When the device cap is opened, it makes a remote "medication taken" record. The device also provides usage reminder in the form of lights and alarm sounds if the cap opening is delayed. Nineteen subjects participated in the survey, and 17 in the intervention phase. Of the 17, 12 had reliable adherence data. Seventeen caregivers of patients and two patients completed the survey. Two most common barriers to adherence identified were lack of reminders and absence of medicine home delivery. The intervention component of this study, which used both the electronic (GlowCap) method and medication possession ratio showed that the median adherence rate for the 12 patients evaluated was 85 %. The GlowCap device accurately kept a record of adherence rates. This device may be an effective tool for increasing HU medication adherence. PMID:27225236

  14. Experimental evidence for the role of lipids in adherence of Candida spp. to human buccal epithelial cells.

    PubMed Central

    Ghannoum, M A; Burns, G R; Elteen, K A; Radwan, S S

    1986-01-01

    Lipids extracted from Candida albicans and C. tropicalis, but not from the weakly adherent C. pseudotropicalis, significantly blocked in vitro adherence of the respective yeast cells to buccal epithelial cells. The percentage of reduction from control values ranged between 16.4 and 42.1%, depending on the species, the strain, and the solvent used for lipid extraction. The constituent lipid classes of both the acetone and chloroform-methanol extracts of C. albicans ATCC 10231 were qualitatively and quantitatively analyzed. The individual classes were isolated by preparative thin-layer chromatography and then tested for their effects on the adherence of this strain to buccal epithelial cells. Individual phospholipids, sterols, and steryl esters blocked adherence significantly (between 15.5 and 55.7% reduction). Triacylglycerols and free fatty acids showed no effect whatsoever. The same results were obtained when standard lipid samples were investigated. Images PMID:3759234

  15. Cell shape-dependent shear stress on adherent cells in a micro-physiologic system as revealed by FEM.

    PubMed

    Pfister, C; Bozsak, C; Wolf, P; Demmel, F; Brischwein, M

    2015-05-01

    Flow-induced shear stress on adherent cells leads to biochemical signaling and mechanical responses of the cells. To determine the flow-induced shear stress on adherent cells cultured in a micro-scaled reaction chamber, we developed a suitable finite element method model. The influence of the most important parameters-cell shape, cell density, shear modulus and fluid velocity-was investigated. Notably, the cell shape strongly influences the resulting shear stress. Long and smooth cells undergo lower shear stress than more rounded cells. Changes in the curvature of the cells lead to stress peaks and single cells experience higher shear stress values than cells of a confluent monolayer. The computational results of the fluid flow simulation were validated experimentally. We also analyzed the influence of flow-induced shear stress on the metabolic activity and shape of L929, a mouse fibroblast cell line, experimentally. The results indicate that threshold stress values for continuous flow conditions cannot be transferred to quasi static flow conditions interrupted by short fluid exchange events. PMID:25856467

  16. Interleukin-3 greatly expands non-adherent endothelial forming cells with pro-angiogenic properties.

    PubMed

    Moldenhauer, Lachlan M; Cockshell, Michaelia P; Frost, Lachlan; Parham, Kate A; Tvorogov, Denis; Tan, Lih Y; Ebert, Lisa M; Tooley, Katie; Worthley, Stephen; Lopez, Angel F; Bonder, Claudine S

    2015-05-01

    Circulating endothelial progenitor cells (EPCs) provide revascularisation for cardiovascular disease and the expansion of these cells opens up the possibility of their use as a cell therapy. Herein we show that interleukin-3 (IL3) strongly expands a population of human non-adherent endothelial forming cells (EXnaEFCs) with low immunogenicity as well as pro-angiogenic capabilities in vivo, making their therapeutic utilisation a realistic option. Non-adherent CD133(+) EFCs isolated from human umbilical cord blood and cultured under different conditions were maximally expanded by day 12 in the presence of IL3 at which time a 350-fold increase in cell number was obtained. Cell surface marker phenotyping confirmed expression of the hematopoietic progenitor cell markers CD133, CD117 and CD34, vascular cell markers VEGFR2 and CD31, dim expression of CD45 and absence of myeloid markers CD14 and CD11b. Functional experiments revealed that EXnaEFCs exhibited classical properties of endothelial cells (ECs), namely binding of Ulex europaeus lectin, up-take of acetylated-low density lipoprotein and contribution to EC tube formation in vitro. These EXnaEFCs demonstrated a pro-angiogenic phenotype within two independent in vivo rodent models. Firstly, a Matrigel plug assay showed increased vascularisation in mice. Secondly, a rat model of acute myocardial infarction demonstrated reduced heart damage as determined by lower levels of serum creatinine and a modest increase in heart functionality. Taken together, these studies show IL3 as a potent growth factor for human CD133(+) cell expansion with clear pro-angiogenic properties (in vitro and in vivo) and thus may provide clinical utility for humans in the future. PMID:25900163

  17. Aspergillus fumigatus MedA governs adherence, host cell interactions and virulence

    PubMed Central

    Gravelat, Fabrice N.; Ejzykowicz, Daniele E.; Chiang, Lisa Y.; Chabot, Josée C.; Urb, Mirjam; Macdonald, K. Denyese; al-Bader, Nadia; Filler, Scott G.; Sheppard, Donald C.

    2010-01-01

    In medically important fungi, regulatory elements that control development and asexual reproduction often govern the expression of virulence traits. We therefore cloned the Aspergillus fumigatus developmental modifier MedA and characterized its role in conidiation, host cell interactions and virulence. As in the model organism Aspergillus nidulans, disruption of medA in A. fumigatus dramatically reduced conidiation. However, the conidiophore morphology was markedly different between the two species. Further, gene expression analysis suggested that MedA governs conidiation through different pathways in A. fumigatus compared to A. nidulans. The A. fumigatus ΔmedA strain was impaired in biofilm production and adherence to plastic, as well as adherence to pulmonary epithelial cells, endothelial cells and fibronectin in vitro. The ΔmedA strain also had reduced capacity to damage pulmonary epithelial cells, and stimulate pro-inflammatory cytokine mRNA and protein expression. Consistent with these results, the A. fumigatus ΔmedA strain also exhibited reduced virulence in both an invertebrate and a mammalian model of invasive aspergillosis. Collectively these results suggest that the downstream targets of A. fumigatus MedA mediate virulence, and may provide novel therapeutic targets for invasive aspergillosis. PMID:19889083

  18. The Rickettsia conorii Autotransporter Protein Sca1 Promotes Adherence to Nonphagocytic Mammalian Cells ▿ †

    PubMed Central

    Riley, Sean P.; Goh, Kenneth C.; Hermanas, Timothy M.; Cardwell, Marissa M.; Chan, Yvonne G. Y.; Martinez, Juan J.

    2010-01-01

    The pathogenesis of spotted fever group (SFG) Rickettsia species, including R. conorii and R. rickettsii, is acutely dependent on adherence to and invasion of host cells, including cells of the mammalian endothelial system. Bioinformatic analyses of several rickettsia genomes revealed the presence of a cohort of genes designated sca genes that are predicted to encode proteins with homology to autotransporter proteins of Gram-negative bacteria. Previous work demonstrated that three members of this family, rOmpA (Sca0), Sca2, and rOmpB (Sca5) are involved in the interaction with mammalian cells; however, very little was known about the function of other conserved rickettsial Sca proteins. Here we demonstrate that sca1, a gene present in nearly all SFG rickettsia genomes, is actively transcribed and expressed in R. conorii cells. Alignment of Sca1 sequences from geographically diverse SFG Rickettsia species showed that there are high degrees of sequence identity and conservation of these sequences, suggesting that Sca1 may have a conserved function. Using a heterologous expression system, we demonstrated that production of R. conorii Sca1 in the Escherichia coli outer membrane is sufficient to mediate attachment to but not invasion of a panel of cultured mammalian epithelial and endothelial cells. Furthermore, preincubation of a recombinant Sca1 peptide with host cells blocked R. conorii cell association. Together, these results demonstrate that attachment to mammalian cells can be uncoupled from the entry process and that Sca1 is involved in the adherence of R. conorii to host cells. PMID:20176791

  19. Fucoidans Disrupt Adherence of Helicobacter pylori to AGS Cells In Vitro

    PubMed Central

    Chua, Eng-Guan; Verbrugghe, Phebe; Perkins, Timothy T.; Tay, Chin-Yen

    2015-01-01

    Fucoidans are complex sulphated polysaccharides derived from abundant and edible marine algae. Helicobacter pylori is a stomach pathogen that persists in the hostile milieu of the human stomach unless treated with antibiotics. This study aims to provide preliminary data to determine, in vitro, if fucoidans can inhibit the growth of H. pylori and its ability to adhere to gastric epithelial cells (AGS). We analysed the activity of three different fucoidan preparations (Fucus A, Fucus B, and Undaria extracts). Bacterial growth was not arrested or inhibited by the fucoidan preparations supplemented into culture media. All fucoidans, when supplemented into tissue culture media at 1000 µg mL−1, were toxic to AGS cells and reduced the viable cell count significantly. Fucoidan preparations at 100 µg mL−1 were shown to significantly reduce the number of adherent H. pylori. These in vitro findings provide the basis for further studies on the clinical use of sulphated polysaccharides as complementary therapeutic agents. PMID:26604968

  20. Fucoidans Disrupt Adherence of Helicobacter pylori to AGS Cells In Vitro.

    PubMed

    Chua, Eng-Guan; Verbrugghe, Phebe; Perkins, Timothy T; Tay, Chin-Yen

    2015-01-01

    Fucoidans are complex sulphated polysaccharides derived from abundant and edible marine algae. Helicobacter pylori is a stomach pathogen that persists in the hostile milieu of the human stomach unless treated with antibiotics. This study aims to provide preliminary data to determine, in vitro, if fucoidans can inhibit the growth of H. pylori and its ability to adhere to gastric epithelial cells (AGS). We analysed the activity of three different fucoidan preparations (Fucus A, Fucus B, and Undaria extracts). Bacterial growth was not arrested or inhibited by the fucoidan preparations supplemented into culture media. All fucoidans, when supplemented into tissue culture media at 1000 µg mL(-1), were toxic to AGS cells and reduced the viable cell count significantly. Fucoidan preparations at 100 µg mL(-1) were shown to significantly reduce the number of adherent H. pylori. These in vitro findings provide the basis for further studies on the clinical use of sulphated polysaccharides as complementary therapeutic agents. PMID:26604968

  1. Extending metabolome coverage for untargeted metabolite profiling of adherent cultured hepatic cells.

    PubMed

    García-Cañaveras, Juan Carlos; López, Silvia; Castell, José Vicente; Donato, M Teresa; Lahoz, Agustín

    2016-02-01

    MS-based metabolite profiling of adherent mammalian cells comprises several challenging steps such as metabolism quenching, cell detachment, cell disruption, metabolome extraction, and metabolite measurement. In LC-MS, the final metabolome coverage is strongly determined by the separation technique and the MS conditions used. Human liver-derived cell line HepG2 was chosen as adherent mammalian cell model to evaluate the performance of several commonly used procedures in both sample processing and LC-MS analysis. In a first phase, metabolite extraction and sample analysis were optimized in a combined manner. To this end, the extraction abilities of five different solvents (or combinations) were assessed by comparing the number and the levels of the metabolites comprised in each extract. Three different chromatographic methods were selected for metabolites separation. A HILIC-based method which was set to specifically separate polar metabolites and two RP-based methods focused on lipidome and wide-ranging metabolite detection, respectively. With regard to metabolite measurement, a Q-ToF instrument operating in both ESI (+) and ESI (-) was used for unbiased extract analysis. Once metabolite extraction and analysis conditions were set up, the influence of cell harvesting on metabolome coverage was also evaluated. Therefore, different protocols for cell detachment (trypsinization or scraping) and metabolism quenching were compared. This study confirmed the inconvenience of trypsinization as a harvesting technique, and the importance of using complementary extraction solvents to extend metabolome coverage, minimizing interferences and maximizing detection, thanks to the use of dedicated analytical conditions through the combination of HILIC and RP separations. The proposed workflow allowed the detection of over 300 identified metabolites from highly polar compounds to a wide range of lipids. PMID:26769129

  2. In vitro adherence patterns of Shigella serogroups to bovine recto-anal junction squamous epithelial (RSE) cells are similar to those of Escherichia coli O157

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The aim of this study was to determine whether Shigella species, which are human gastrointestinal pathogens, can adhere to cattle recto-anal junction squamous epithelial (RSE) cells using a recently standardized adherence assay, and to compare their adherence patterns to that of Escherichia coli O15...

  3. Relationships among stressful life events and physiological markers, treatment adherence, and psychosocial functioning among youth with type 2 diabetes

    PubMed Central

    Walders-Abramson, Natalie; Venditti, Elizabeth M.; Ievers-Landis, Carolyn E.; Anderson, Barbara; ghormli, Laure El; Geffner, Mitchell; Kaplan, Joan; Koontz, Michaela B.; Saletsky, Ron; Payan, Marisa; Yasuda, Patrice

    2014-01-01

    Objective To examine the relationship between stressful life events and physiological measures, medication adherence, depressive symptoms, and impaired quality of life in adolescents with recent onset type 2 diabetes (T2D). Study design Data were collected from 497 ethnically diverse participants (66% female) in the final year of the Treatment Options for type 2 Diabetes in Adolescents and Youth (TODAY) multi-center clinical trial. Exposure to 32 possible events over the prior year and rating of subsequent distress or upset were collected by self-report, and summarized in a major stressors score. The score was analyzed for relationship to glycemic control (HbA1c and treatment failure), BMI, diagnosis of hypertension or triglycerides dyslipidemia, oral medication adherence, presence of depressive symptoms, and impaired quality of life. Results The total number of major stressful life events was calculated, with 33% of the sample reporting none, 67% at least one, 47% at least two, 33% at least three, and 20% reporting four or more. There were no associations between major stressors score and physiological measures or diagnosis of comorbidities. The odds of medication non-adherence increased significantly from those reporting at least one major stressor (odds ratio=1.58, p=0.0265) to those reporting at least 4 (odds ratio=2.70, p=0.0009). Significant odds of elevated depressive symptoms and impaired quality of life were also found with increased reporting of major stressors. Conclusions Exposure to major stressful life events is association with lower adherence and impaired psychosocial functioning among adolescents with T2D. PMID:24948348

  4. Modification of Solid Phase Red Cell Adherence Assay for the Detection of Platelet Antibodies in Patients With Thrombocytopenia

    PubMed Central

    Vongchan, Preeyanat; Nawarawong, Weerasak; Linhardt, Robert J.

    2009-01-01

    Platelet refractoriness is caused by HLA antibodies and platelet-specific antibodies. Current methods used to detect antiplatelet antibodies have limitations. Solid phase red cell adherence (SPRCA) lacks sensitivity and requires a second assay using chloroquine-treated intact platelets to specify the response due to anti-HLA. We modified SPRCA by using 2 types of antihuman platelet antibodies with different specificities toward platelet lysate and tested samples from 361 patients (69 with unexplained thrombocytopenia and 292 with poor response to platelet transfusions not explicable by alloimmunization or the clinical situation) and 50 from healthy volunteers. Our method compared favorably with platelet suspension direct immunofluorescence. All samples from healthy volunteers were negative; of the samples from the patient population, 240 were positive (147 samples had only antiplatelet and 3 samples had only anti-HLA antibodies). This modified technique had a sensitivity of 98% and a specificity of 91%. PMID:18701420

  5. Lactobacilli Interfere with Streptococcus pyogenes Hemolytic Activity and Adherence to Host Epithelial Cells.

    PubMed

    Saroj, Sunil D; Maudsdotter, Lisa; Tavares, Raquel; Jonsson, Ann-Beth

    2016-01-01

    Streptococcus pyogenes [Group A streptococcus (GAS)], a frequent colonizer of the respiratory tract mucosal surface, causes a variety of human diseases, ranging from pharyngitis to the life-threatening streptococcal toxic shock-like syndrome. Lactobacilli have been demonstrated to colonize the respiratory tract. In this study, we investigated the interference of lactobacilli with the virulence phenotypes of GAS. The Lactobacillus strains L. rhamnosus Kx151A1 and L. reuteri PTA-5289, but not L. salivarius LMG9477, inhibited the hemolytic activity of S. pyogenes S165. The inhibition of hemolytic activity was attributed to a decrease in the production of streptolysin S (SLS). Conditioned medium (CM) from the growth of L. rhamnosus Kx151A1 and L. reuteri PTA-5289 was sufficient to down-regulate the expression of the sag operon, encoding SLS. The Lactobacillus strains L. rhamnosus Kx151A1, L. reuteri PTA-5289, and L. salivarius LMG9477 inhibited the initial adherence of GAS to host epithelial cells. Intriguingly, competition with a combination of Lactobacillus species reduced GAS adherence to host cells most efficiently. The data suggest that an effector molecule released from certain Lactobacillus strains attenuates the production of SLS at the transcriptional level and that combinations of Lactobacillus strains may protect the pharyngeal mucosa more efficiently from the initial colonization of GAS. The effector molecules released from Lactobacillus strains affecting the virulence phenotypes of pathogens hold potential in the development of a new generation of therapeutics. PMID:27524981

  6. Lactobacilli Interfere with Streptococcus pyogenes Hemolytic Activity and Adherence to Host Epithelial Cells

    PubMed Central

    Saroj, Sunil D.; Maudsdotter, Lisa; Tavares, Raquel; Jonsson, Ann-Beth

    2016-01-01

    Streptococcus pyogenes [Group A streptococcus (GAS)], a frequent colonizer of the respiratory tract mucosal surface, causes a variety of human diseases, ranging from pharyngitis to the life-threatening streptococcal toxic shock-like syndrome. Lactobacilli have been demonstrated to colonize the respiratory tract. In this study, we investigated the interference of lactobacilli with the virulence phenotypes of GAS. The Lactobacillus strains L. rhamnosus Kx151A1 and L. reuteri PTA-5289, but not L. salivarius LMG9477, inhibited the hemolytic activity of S. pyogenes S165. The inhibition of hemolytic activity was attributed to a decrease in the production of streptolysin S (SLS). Conditioned medium (CM) from the growth of L. rhamnosus Kx151A1 and L. reuteri PTA-5289 was sufficient to down-regulate the expression of the sag operon, encoding SLS. The Lactobacillus strains L. rhamnosus Kx151A1, L. reuteri PTA-5289, and L. salivarius LMG9477 inhibited the initial adherence of GAS to host epithelial cells. Intriguingly, competition with a combination of Lactobacillus species reduced GAS adherence to host cells most efficiently. The data suggest that an effector molecule released from certain Lactobacillus strains attenuates the production of SLS at the transcriptional level and that combinations of Lactobacillus strains may protect the pharyngeal mucosa more efficiently from the initial colonization of GAS. The effector molecules released from Lactobacillus strains affecting the virulence phenotypes of pathogens hold potential in the development of a new generation of therapeutics. PMID:27524981

  7. Role of specific determinants in mannan of Candida albicans serotype A in adherence to human buccal epithelial cells.

    PubMed Central

    Miyakawa, Y; Kuribayashi, T; Kagaya, K; Suzuki, M; Nakase, T; Fukazawa, Y

    1992-01-01

    Candida albicans serotype A (C. albicans A) possesses a specific antigen, designated antigen 6, which resides in mannans on the cell surface. To determine the role of the mannan moiety of the C. albicans cell wall in adherence to buccal epithelial cells, we used antigen 6-deficient mutants which had been isolated by screening with an agglutinating monoclonal antibody against antigen 6 (MAb-6). 1H nuclear magnetic resonance spectral analysis of the purified mannans from the mutants showed a loss of the signals related to that beta-linkage of the side chains. Moreover, acetolyzed fragments of the mutant mannans showed a decreased amount of mannohexaose and mannopentaose. The mutant yeast cells exhibited significantly reduced ability to adhere both to exfoliated buccal epithelial cells and to a human buccal cell line. A number of strains of C. albicans A, C. tropicalis, and C. glabrata, all of which bear antigen 6, showed significantly higher adherence to the cell line than did those of C. albicans serotype B, which lack antigen 6. The whole mannan from the C. albicans A parent inhibited the adherence of C. albicans A to epithelial cells dose dependently, whereas mannan from a mutant strains did not. Moreover, C. albicans A treated with MAb-6 or polyclonal factor 6 serum showed reduced adherence. A close correlation was found between adhesive ability and agglutinability with MAb-6 in the C. albicans A parent, the antigenic mutants, and their spontaneous revertants. These results suggest that so far as mannan adhesion is concerned, serotype A-specific determinants are largely involved in the mechanisms of adherence of C. albicans A to human buccal epithelial cells. PMID:1375200

  8. Lithium attenuates lead induced toxicity on mouse non-adherent bone marrow cells.

    PubMed

    Banijamali, Mahsan; Rabbani-Chadegani, Azra; Shahhoseini, Maryam

    2016-07-01

    Lead is a poisonous heavy metal that occurs in all parts of environment and causes serious health problems in humans. The aim of the present study was to investigate the possible protective effect of lithium against lead nitrate induced toxicity in non-adherent bone marrow stem cells. Trypan blue and MTT assays represented that exposure of the cells to different concentrations of lead nitrate decreased viability in a dose dependent manner, whereas, pretreatment of the cells with lithium protected the cells against lead toxicity. Lead reduced the number and differentiation status of bone marrow-derived precursors when cultured in the presence of colony stimulating factor (CSF), while the effect was attenuated by lithium. The cells treated with lead nitrate exhibited cell shrinkage, DNA fragmentation, anion superoxide production, but lithium prevented lead action. Moreover, apoptotic indexes such as PARP cleavage and release of HMGB1 induced by lead, were protected by lithium, suggesting anti-apoptotic effect of lithium. Immunoblot analysis of histone H3K9 acetylation indicated that lithium overcame lead effect on acetylation. In conclusion, lithium efficiently reduces lead toxicity suggesting new insight into lithium action which may contribute to increased cell survival. It also provides a potentially new therapeutic strategy for lithium and a cost-effective approach to minimize destructive effects of lead on bone marrow stem cells. PMID:27259346

  9. Viability of adhered bacterial cells: tracking MinD protein oscillations

    NASA Astrophysics Data System (ADS)

    Barrett, Matt; Colville, Keegan; Schultz-Nielsen, Chris; Jericho, Manfred; Dutcher, John

    2010-03-01

    To study bacterial cells using atomic force microscopy, it is necessary to immobilize the cells on a substrate. Because bacterial cells and common substrates such as glass and mica have a net negative charge, positively charged polymers such as poly-L-lysine (PLL) and polyethyleneimine (PEI) are commonly used as adhesion layers. However, the use of adhesion polymers could stress the cell and even render it inviable. Viable E. coli cells use oscillations of Min proteins along the axis of the rod-shaped cells to ensure accurate cell division. By tagging MinD proteins with GFP, oscillations can be observed using fluorescence microscopy. For a healthy cell in an ideal environment, the oscillation period is measured to be ˜40 s. Prior experiments have shown that PLL increases the oscillation period significantly (up to 80%). In the present study, we have used epifluorescence and total internal reflection fluorescence (TIRF) to track MinD protein oscillations in E. coli bacteria adhered to a variety of positively charged polymers on mica as a function of polymer surface coverage.

  10. In vitro inhibition of calcium oxalate crystallization and crystal adherence to renal tubular epithelial cells by Terminalia arjuna.

    PubMed

    Mittal, A; Tandon, S; Singla, S K; Tandon, C

    2016-04-01

    Urolithiasis is a multifactorial disease and remains a public health problem around the world. Of all types of renal stones, calcium oxalate (CaOx) is the most common composition formed in the urinary system of the patients with urolithiasis. The present study is aimed at evaluating the antiurolithiatic properties of the Tris-Cl extract (TE) of Terminalia arjuna (T. arjuna). The antilithiatic activity of TE of T. arjuna was investigated on nucleation, aggregation, and growth of the CaOx crystals, as well as its protective potency was tested on oxalate-induced cell injury of NRK-52E renal epithelial cells. Also, in vitro antioxidant activity of TE T. arjuna bark was also determined. The TE of T. arjuna exhibited a concentration-dependent inhibition of nucleation and growth of CaOx crystals. Inhibition of aggregation of CaOx crystals remains constant. When NRK-52E cells were injured by exposure to oxalate for 48 h, the TE prevented the cells from injury and CaOx crystal adherence resulting in increased cell viability in a dose-dependent manner. The TE also scavenged the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radicals with an IC50 at 51.72 µg/mL. The results indicated that T. arjuna is a potential candidate for phytotherapy against urolithiasis as it attains the ability to inhibit CaOx crystallization and scavenge DPPH free radicals in vitro along with a cytoprotective role. PMID:26424092

  11. The HAART cell phone adherence trial (WelTel Kenya1): a randomized controlled trial protocol

    PubMed Central

    Lester, Richard T; Mills, Edward J; Kariri, Antony; Ritvo, Paul; Chung, Michael; Jack, William; Habyarimana, James; Karanja, Sarah; Barasa, Samson; Nguti, Rosemary; Estambale, Benson; Ngugi, Elizabeth; Ball, T Blake; Thabane, Lehana; Kimani, Joshua; Gelmon, Lawrence; Ackers, Marta; Plummer, Francis A

    2009-01-01

    Background The objectives are to compare the effectiveness of cell phone-supported SMS messaging to standard care on adherence, quality of life, retention, and mortality in a population receiving antiretroviral therapy (ART) in Nairobi, Kenya. Methods and Design A multi-site randomized controlled open-label trial. A central randomization centre provided opaque envelopes to allocate treatments. Patients initiating ART at three comprehensive care clinics in Kenya will be randomized to receive either a structured weekly SMS ('short message system' or text message) slogan (the intervention) or current standard of care support mechanisms alone (the control). Our hypothesis is that using a structured mobile phone protocol to keep in touch with patients will improve adherence to ART and other patient outcomes. Participants are evaluated at baseline, and then at six and twelve months after initiating ART. The care providers keep a weekly study log of all phone based communications with study participants. Primary outcomes are self-reported adherence to ART and suppression of HIV viral load at twelve months scheduled follow-up. Secondary outcomes are improvements in health, quality of life, social and economic factors, and retention on ART. Primary analysis is by 'intention-to-treat'. Sensitivity analysis will be used to assess per-protocol effects. Analysis of covariates will be undertaken to determine factors that contribute or deter from expected and determined outcomes. Discussion This study protocol tests whether a novel structured mobile phone intervention can positively contribute to ART management in a resource-limited setting. Trial Registration Trial Registration Number: NCT00830622 PMID:19772596

  12. NLRP3 protects alveolar barrier integrity by an inflammasome-independent increase of epithelial cell adherence.

    PubMed

    Kostadinova, Elena; Chaput, Catherine; Gutbier, Birgitt; Lippmann, Juliane; Sander, Leif E; Mitchell, Timothy J; Suttorp, Norbert; Witzenrath, Martin; Opitz, Bastian

    2016-01-01

    Bacterial pneumonia is a major cause of acute lung injury and acute respiratory distress syndrome, characterized by alveolar barrier disruption. NLRP3 is best known for its ability to form inflammasomes and to regulate IL-1β and IL-18 production in myeloid cells. Here we show that NLRP3 protects the integrity of the alveolar barrier in a mouse model of Streptococcus pneumoniae-induced pneumonia, and ex vivo upon treatment of isolated perfused and ventilated lungs with the purified bacterial toxin, pneumolysin. We reveal that the preserving effect of NLRP3 on the lung barrier is independent of inflammasomes, IL-1β and IL-18. NLRP3 improves the integrity of alveolar epithelial cell monolayers by enhancing cellular adherence. Collectively, our study uncovers a novel function of NLRP3 by demonstrating that it protects epithelial barrier function independently of inflammasomes. PMID:27476670

  13. NLRP3 protects alveolar barrier integrity by an inflammasome-independent increase of epithelial cell adherence

    PubMed Central

    Kostadinova, Elena; Chaput, Catherine; Gutbier, Birgitt; Lippmann, Juliane; Sander, Leif E.; Mitchell, Timothy J.; Suttorp, Norbert; Witzenrath, Martin; Opitz, Bastian

    2016-01-01

    Bacterial pneumonia is a major cause of acute lung injury and acute respiratory distress syndrome, characterized by alveolar barrier disruption. NLRP3 is best known for its ability to form inflammasomes and to regulate IL-1β and IL-18 production in myeloid cells. Here we show that NLRP3 protects the integrity of the alveolar barrier in a mouse model of Streptococcus pneumoniae-induced pneumonia, and ex vivo upon treatment of isolated perfused and ventilated lungs with the purified bacterial toxin, pneumolysin. We reveal that the preserving effect of NLRP3 on the lung barrier is independent of inflammasomes, IL-1β and IL-18. NLRP3 improves the integrity of alveolar epithelial cell monolayers by enhancing cellular adherence. Collectively, our study uncovers a novel function of NLRP3 by demonstrating that it protects epithelial barrier function independently of inflammasomes. PMID:27476670

  14. Investigation on cytoskeleton dynamics for non-adherent cells under point-like stimuli

    NASA Astrophysics Data System (ADS)

    Miccio, Lisa; Memmolo, Pasquale; Merola, Francesco; Mugnano, Martina; Fusco, Sabato; Paciello, Antonio; Ferraro, Pietro; Netti, Paolo A.

    2015-05-01

    In the present paper, Holographic Optical Tweezers (HOT) is employed to trap and manage functionalized micrometric latex beads with the aim at probing cellular forces in no-adherent state. For the first time at best of our knowledge, a suspended cell, subjected to mechanical stress, structures its cytoskeleton when anchored to point-like bonds. We exploit the HOT arrangement to induce mechanical deformation in suspended NIH 3T3 fibroblast. Our investigation is devoted to understand the inner cell mechanism when it is mechanically stressed by point-like stimulus without the substrate influence. In our experiment, cell adhesion is prevented and the stimulus is applied through latex beads trapped by HOT and positioned externally to the cell membrane. Our aims are devoted to analyze cell response during the transition from an homogeneous and isotropic structure (as it's in suspension) to a mechanically stressed state. To analyze the cell material interaction we combine the HOT arrangement with two imaging systems: a Digital Holography (DH) setup in microscope configuration that is an investigation method useful for quantitative, label-free and full-field analysis of low contrast object and a fluorescence modulus. HOT are exploited to induce cellular response to specific stimuli while DH allows to measure such responses in no-invasive way. Finally, fluorescence imaging is added to discriminate the inner cell structures.

  15. Adherence to a Mediterranean-type dietary pattern and cognitive decline in a community population123

    PubMed Central

    Tangney, Christine C; Kwasny, Mary J; Li, Hong; Wilson, Robert S; Evans, Denis A; Morris, Martha Clare

    2011-01-01

    Background: Many of the foods abundant in the traditional Mediterranean diet, such as vegetables and fish, have been associated with slower cognitive decline. Objective: We investigated whether adherence to a Mediterranean dietary pattern or to the Healthy Eating Index–2005 (HEI-2005) is associated with cognitive change in older adults. Design: This article is based on analyses of data from an ongoing longitudinal study in adults aged ≥65 y known as the Chicago Health and Aging Project (CHAP). CHAP participants (2280 blacks and 1510 whites) with ≥2 cognitive assessments were evaluated for adherence to 1) the Mediterranean dietary pattern (MedDiet; maximum score: 55) and 2) the HEI-2005 (maximum score: 100). For both scoring systems, higher scores connote greater adherence. Cognitive function was assessed at 3-y intervals on the basis of a composite measure of global cognition. Linear mixed models were used to examine the association of dietary scores to change in cognitive function. Mean follow-up time was 7.6 y. Results: Mean (±SD) scores for participants were 28.2 ± 0.1 for the MedDiet and 61.2 ± 9.6 for the HEI-2005. White participants had higher energy-adjusted MedDiet scores but lower HEI-2005 scores than did black participants. Higher MedDiet scores were associated with slower rates of cognitive decline (β = +0.0014 per 1-point increase, SEE = 0.0004, P = 0.0004) after adjustment for age, sex, race, education, participation in cognitive activities, and energy. No such associations were observed for HEI-2005 scores. Conclusion: The Mediterranean dietary pattern as captured by the MedDiet scoring system may reduce the rate of cognitive decline with older age. PMID:21177796

  16. Curli modulates adherence of Escherichia coli O157 to bovine recto-anal junction squamous epithelial cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Our recent studies have shown that Intimin and the Locus of Enterocyte Effacement-encoded proteins do not play a role in Escherichia coli O157 (O157) adherence to the bovine recto-anal junction squamous epithelial cells (RSE) cells. Hence, to define factors that play a contributory role, we investi...

  17. The effects of disodium cromoglycate on enhanced adherence of Haemophilus influenzae to A549 cells infected with respiratory syncytial virus.

    PubMed

    Fukasawa, Chie; Ishiwada, Naruhiko; Ogita, Junko; Hishiki, Haruka; Kohno, Yoichi

    2009-08-01

    Nontypeable Haemophilus influenzae (NTHi) secondary infection often complicates respiratory syncytial virus (RSV) infections. Previous studies have revealed that RSV infections enhance NTHi adherence to airway epithelial cells. In this study, we investigated the effects of disodium cromoglycate (DSCG) and corticosteroids, which are frequently used for the treatment of wheezing often related to RSV infections, on the adherence of NTHi to RSV-infected A549 cells. DSCG inhibited enhanced adherence of NTHi to RSV-infected A549 cells, whereas dexamethasone (Dex) and fluticasone propionate (Fp) did not. DSCG suppressed the expression of ICAM-1, which is one of the NTHi receptors. Furthermore, DSCG exhibited an inhibitory effect on RSV infections. It is suggested that DSCG exerts an anti-RSV effect, and consequently attenuates the expression of NTHi receptors. PMID:19390482

  18. Laser-generated Micro-bubbles for Molecular Delivery to Adherent Cells

    NASA Astrophysics Data System (ADS)

    Genc, Suzanne Lee

    We examine the use of optical breakdown in aqueous media as a means to deliver molecules into live adherent cell cultures. This process, called optoinjection (OI), is affected both by the media composition and the cellular exposure to hydrodynamic stresses associated with the cavitation bubble formed by the optical breakdown process. Here we explore the possibility of performing OI using laser microbeams focused at low numerical aperture to provide conditions where OI can be performed at high-throughput. We first investigate the effect of media composition on plasma and cavitation bubble formation. We make the discovery that irradiation of minimal essential media, supports the formation of low-density plasmas (LDP) resulting in the generation of small (2--20 mum radius) cavitation bubbles. This provides gentle specific hydrodynamic perturbations to single or small groups of cells. The addition of supplemental fetal bovine serum to the medium prevents the formation LDPs and the resulting avalanche ionization generates larger (> 100 mum radius) bubbles and more violent hydrodynamic effects. Second, using high-speed photography we provide the first visualization of LDP-generated cavitation bubbles at precise offset locations relative to a boundary on which a cell monolayer can be cultured. These images depict the cellular exposure to different hydrodynamic conditions depending on the normalized offset distance (gamma = s/Rmax) and show how it affects the cellular exposure to shear stresses upon bubble expansion and different distributions of bubble energy upon collapse. Lastly, we examine the effects of pulse energy, parameters, and single vs. multiple laser exposures on the ability to deliver 3-5 kDa dextrans into adherent cells using both small (< 20 mum) and large (100mu m) radius bubbles. For single exposures, we identify several conditions under which OI can be optimized: (a) conditions where cell viability is maximized (˜90%) but optoinjection of viable cells

  19. Role of the Amino-Terminal Region of Porphyromonas gingivalis Fimbriae in Adherence to Epithelial Cells

    PubMed Central

    Sojar, Hakimuddin T.; Han, Yiping; Hamada, Nobushiro; Sharma, Ashu; Genco, Robert J.

    1999-01-01

    Porphyromonas gingivalis fimbriae elicit many responses in eukaryotic cells, including mitogenicity, cytokine production, epithelial cell invasion, and cellular immune response. Specific domains of the major fimbrial protein (FimA) have been shown to be important in triggering some of these functions. The goal of the present study was to identify the domain(s) of P. gingivalis FimA responsible for specific interaction with human mucosal epithelial cells. Fimbriated P. gingivalis strains have been shown to bind to buccal epithelial cells, whereas nonfimbriated strains bind at low levels or not at all. This and other studies provide evidence that FimA mediates the adherence of P. gingivalis to oral epithelial cells. To determine the specific region(s) of P. gingivalis FimA involved in epithelial cell binding, specific antipeptide antibodies were used to inhibit the binding of iodinated purified fimbriae as well as the binding of P. gingivalis cells to epithelial cells. Antibodies directed against peptides 49 to 68 (VVMANTAGAMELVGKTLAEVK) and 69 to 90 (ALTTELTAENQEAAGLIMTAEP) were found to highly inhibit both the binding of fimbriae and the binding of P. gingivalis cells to epithelial cells. The antibody against FimA peptides 69 to 90 also reacted with P. gingivalis fimbriae in immunogold labeling and immunoblot analysis, thereby indicating that this peptide domain is exposed on the surface of fimbriae. Our results suggest that the amino-terminal domain corresponding to amino acid residues 49 to 90 of the fimbrillin protein is a major epithelial cell binding domain of P. gingivalis fimbriae. PMID:10531284

  20. Enhanced adherence of mouse fibroblast and vascular cells to plasma modified polyethylene.

    PubMed

    Reznickova, Alena; Novotna, Zdenka; Kolska, Zdenka; Kasalkova, Nikola Slepickova; Rimpelova, Silvie; Svorcik, Vaclav

    2015-01-01

    Since the last decade, tissue engineering has shown a sensational promise in providing more viable alternatives to surgical procedures for harvested tissues, implants and prostheses. Biomedical polymers, such as low-density polyethylene (LDPE), high-density polyethylene (HDPE) and ultra-high molecular weight polyethylene (UHMWPE), were activated by Ar plasma discharge. Degradation of polymer chains was examined by determination of the thickness of ablated layer. The amount of an ablated polymer layer was measured by gravimetry. Contact angle, measured by goniometry, was studied as a function of plasma exposure and post-exposure aging times. Chemical structure of modified polymers was characterized by angle resolved X-ray photoelectron spectroscopy. Surface chemistry and polarity of the samples were investigated by electrokinetic analysis. Changes in surface morphology were followed using atomic force microscopy. Cytocompatibility of plasma activated polyethylene foils was studied using two distinct model cell lines; VSMCs (vascular smooth muscle cells) as a model for vascular graft testing and connective tissue cells L929 (mouse fibroblasts) approved for standardized material cytotoxicity testing. Specifically, the cell number, morphology, and metabolic activity of the adhered and proliferated cells on the polyethylene matrices were studied in vitro. It was found that the plasma treatment caused ablation of the polymers, resulting in dramatic changes in their surface morphology and roughness. ARXPS and electrokinetic measurements revealed oxidation of the polymer surface. It was found that plasma activation has a positive effect on the adhesion and proliferation of VSMCs and L929 cells. PMID:25953566

  1. Effects of pressure and temperature on the survival rate of adherent A-172 cells

    NASA Astrophysics Data System (ADS)

    Yasuhara, Ryo; Kushida, Ryo; Ishii, Shiwori; Yamanoha, Banri; Shimizu, Akio

    2013-06-01

    Preservation of cells under high pressure is an important alternative to cryopreservation. We studied the effect of temperature (4, 25, 37°C) and pressure (0.1-350 MPa) on the survival rate of A-172 glioblastoma cells. The survival rate was not changed by brief (10 min) pressurization of up to 150 MPa, but the survival rate began to decrease from 150 MPa, and most of the A-172 cells died when treated with over 200 MPa. Lengthy pressurization (4 days) at lower pressure (upto 20.1 MPa) without medium exchange showed complex results. The survival rate of cells preserved at 25°C showed two maxima at 1.6 and 20.1 MPa. After preservation, cells adhered and proliferated in the same way as normal cells when cultured at 37°C in a CO2 incubator. The other two temperatures, 4° and 37°C, showed no maximum survival rate. Therefore, a high survival rate can be maintained with high pressure treatment.

  2. The impact of environmental changes upon the microrheological response of adherent cells.

    PubMed

    Picard, C; Donald, A

    2009-10-01

    The mechanical behaviour of adherent cells cultured in vitro is known to be dependent on the mechanical properties of the substrate. We show that this mechanical behaviour is also strongly affected by the cells' environment. We focus here on the impact of temperature and pH. Experiments carried out on individual cells in a tuneable environment reveal that the intra-cellular mechanical behaviour exhibits large and fast changes when the external cell environment is changed. Fast passive microrheometry measurements allow for the precise characterisation of the transient regime observed during a temperature drop. When maintained at a non-physiological temperature, the cells reach a stabilised state distinct from the state observed in physiological conditions. The perturbation can be reversed but exhibits hysteretic behaviour when physiological conditions are restored. The transient regime observed during the recovery process is found to be different from the transient regime observed when leaving physiological conditions. A modified generalized Stokes-Einstein equation taking into account the cell activity through an effective temperature is proposed here to fit the experimental results. Excellent agreement between the model and the measurements is obtained for time lags from 10⁻³ to 1 s considered in this study. PMID:19551417

  3. Soluble fibrin augments platelet/tumor cell adherence in vitro and in vivo, and enhances experimental metastasis.

    PubMed

    Biggerstaff, J P; Seth, N; Amirkhosravi, A; Amaya, M; Fogarty, S; Meyer, T V; Siddiqui, F; Francis, J L

    1999-01-01

    There is considerable evidence for a relationship between hemostasis and malignancy. Since platelet adhesion to tumor cells has been implicated in the metastatic process and plasma levels of fibrinogen (Fg) and soluble fibrin (sFn) monomer are increased in cancer, we hypothesized that these molecules might enhance tumor-platelet interaction. We therefore studied binding of sFn monomer to tumor cells in a static microplate adhesion assay and determined the effect of pre-treating tumor cells with sFn on tumor cell-induced thrombocytopenia and experimental metastasis. Soluble fibrin (produced by adding thrombin to FXIII- and plasminogen-free Fg in the presence of Gly-Pro-Arg-Pro-amide (GPRP-NH2) significantly increased platelet adherence to tumor cells. This effect was primarily mediated by the integrins alphaIIb beta3 on the platelet and CD 54 (ICAM-1) on the tumor cells. Platelets adhered to untreated A375 cells (28 +/- 8 platelets/tumor cell) and this was not significantly affected by pre-treatment of the tumor cells with fibrinogen or GPRP-NH2. Although thrombin treatment increased adherence, pre-incubation of the tumor cells with sFn resulted in a further increase in platelet binding to tumor cells. In contrast to untreated tumor cells, intravenous injection of sFn-treated A 375 cells reduced the platelet count in anticoagulated mice, supporting the in vitro finding that sFn enhanced tumor cell-platelet adherence. In a more aggressive model of experimental metastasis, treating tumor cells with sFn enhanced lung seeding by 65% compared to untreated cells. Extrapolation of our data to the clinical situation suggests that coagulation activation, and subsequent increase in circulating Fn monomer, may enhance platelet adhesion to circulating tumor cells and thereby facilitate metastatic spread. PMID:10919717

  4. Adherent-phagocytic cells influence suppressed concanavalin-A induced proliferation of spleen lymphoid cells in copper deficient rats

    SciTech Connect

    Kramer, T.R.; Briske-Anderson, M.; Johnson, W.T.

    1986-03-01

    Weanling male Lewis rats (N = 10/group) were fed ad-libitum for 42 days diets based on AIN standards containing 21% casein, 5% safflower oil, and deficient (0.6 ..mu..g/g) or adequate (5.6 ..mu..g/g) levels of cu. Cu-deficient rats showed typical biochemical and hematological changes. Immunological changes exhibited by Cu-deficient rats were influenced by the presence of splenic adherent-phagocytic cells (macrophage-like), but not by cytochrome-c oxidase activity of spleen lymphoid cells (SLC). Decreased proliferation was exhibited by concanavalin-A (Con-A) stimulated SLC of Cu-deficient rats. Following removal of plastic-adherent phagocytic cells from the SLC suspensions, equivalent proliferation was exhibited by Con-A stimulated nonadherent-SLC of Cu-deficient and Cu-adequate rats. Decreased cytochrome-c oxidase activity was exhibited by both unstimulated SLC and nonadherent-SLC of Cu-deficient rats, but decreased proliferation was exhibited only in Con-A stimulated SLC of Cu-deficient rats. These findings indicate that nonadherent splenic T-lymphocytes of Cu-deficient rats are not impaired in their ability to proliferate, and that cytochrome-c oxidase activity in unstimulated lymphoid cells of Cu-deficient rats is apparently not related to levels of proliferation by the Con-A stimulated cells.

  5. Reduction of Escherichia coli adherence to uroepithelial bladder cells after consumption of cranberry juice: a double-blind randomized placebo-controlled cross-over trial.

    PubMed

    Di Martino, P; Agniel, R; David, K; Templer, C; Gaillard, J L; Denys, P; Botto, H

    2006-02-01

    To determine the efficacy of the consumption of cranberry juice versus placebo with regard to the presence of in vitro bacterial anti-adherence activity in the urine of healthy volunteers. Twenty healthy volunteers, 10 men and 10 women, were included. The study was a double-blind, randomized, placebo-controlled, and cross-over study. In addition to normal diet, each volunteer received at dinner a single dose of 750 ml of a total drink composed of: (1) 250 ml of the placebo and 500 ml of mineral water, or (2) 750 ml of the placebo, or (3) 250 ml of the cranberry juice and 500 ml of mineral water, or (4) 750 ml of the cranberry juice. Each volunteer took the four regimens successively in a randomly order, with a washout period of at least 6 days between every change in regimen. The first urine of the morning following cranberry or placebo consumption was collected and used to support bacterial growth. Six uropathogenic Escherichia coli strains (all expressing type 1 pili; three positive for the gene marker for P-fimbriae papC and three negative for papC), previously isolated from patients with symptomatic urinary tract infections, were grown in urine samples and tested for their ability to adhere to the T24 bladder cell line in vitro. There were no significant differences in the pH or specific gravity between the urine samples collected after cranberry or placebo consumption. We observed a dose dependent significant decrease in bacterial adherence associated with cranberry consumption. Adherence inhibition was observed independently from the presence of genes encoding type P pili and antibiotic resistance phenotypes. Cranberry juice consumption provides significant anti-adherence activity against different E. coli uropathogenic strains in the urine compared with placebo. PMID:16397814

  6. Xanthomonas citri subsp. citri type IV Pilus is required for twitching motility, biofilm development, and adherence.

    PubMed

    Dunger, German; Guzzo, Cristiane R; Andrade, Maxuel O; Jones, Jeffrey B; Farah, Chuck S

    2014-10-01

    Bacterial type IV pili (T4P) are long, flexible surface filaments that consist of helical polymers of mostly pilin subunits. Cycles of polymerization, attachment, and depolymerization mediate several pilus-dependent bacterial behaviors, including twitching motility, surface adhesion, pathogenicity, natural transformation, escape from immune system defense mechanisms, and biofilm formation. The Xanthomonas citri subsp. citri strain 306 genome codes for a large set of genes involved in T4P biogenesis and regulation and includes several pilin homologs. We show that X. citri subsp. citri can exhibit twitching motility in a manner similar to that observed in other bacteria such as Pseudomonas aeruginosa and Xylella fastidiosa and that this motility is abolished in Xanthomonas citri subsp. citri knockout strains in the genes coding for the major pilin subunit PilAXAC3241, the ATPases PilBXAC3239 and PilTXAC2924, and the T4P biogenesis regulators PilZXAC1133 and FimXXAC2398. Microscopy analyses were performed to compare patterns of bacterial migration in the wild-type and knockout strains and we observed that the formation of mushroom-like structures in X. citri subsp. citri biofilm requires a functional T4P. Finally, infection of X. citri subsp. citri cells by the bacteriophage (ΦXacm4-11 is T4P dependent. The results of this study improve our understanding of how T4P influence Xanthomonas motility, biofilm formation, and susceptibility to phage infection. PMID:25180689

  7. Lactoferrin affects the adherence and invasion of Streptococcus dysgalactiae ssp. dysgalactiae in mammary epithelial cells.

    PubMed

    O'Halloran, Fiona; Beecher, Christine; Chaurin, Valerie; Sweeney, Torres; Giblin, Linda

    2016-06-01

    Streptococcus dysgalactiae ssp. dysgalactiae is an important causative agent of bovine mastitis worldwide. Lactoferrin is an innate immune protein that is associated with many functions including immunomodulatory, antiproliferative, and antimicrobial properties. This study aimed to investigate the interactions between lactoferrin and a clinical bovine mastitis isolate, Strep. dysgalactiae ssp. dysgalactiae DPC5345. Initially a deliberate in vivo bovine intramammary challenge was performed with Strep. dysgalactiae DPC5345. Results demonstrated a significant difference in lactoferrin mRNA levels in milk cells between the control and infused quarters 7h postinfusion. Milk lactoferrin levels in the Strep. dysgalactiae DPC5345 infused quarters were significantly increased compared with control quarters at 48h postinfusion. In vitro studies demonstrated that lactoferrin had a bacteriostatic effect on the growth of Strep. dysgalactiae DPC5345 and significantly decreased the ability of the bacteria to internalize into HC-11 mammary epithelial cells. Confocal microscopy images of HC-11 cells exposed to Strep. dysgalactiae and lactoferrin further supported this effect by demonstrating reduced invasion of bacteria to HC-11 cells. The combined data suggest that a bovine immune response to Strep. dysgalactiae infection includes a significant increase in lactoferrin expression in vivo, and based on in vitro data, lactoferrin limits mammary cell invasion of this pathogen by binding to the bacteria and preventing its adherence. PMID:27016824

  8. Surface glycosaminoglycans mediate adherence between HeLa cells and Lactobacillus salivarius Lv72

    PubMed Central

    2013-01-01

    Background The adhesion of lactobacilli to the vaginal surface is of paramount importance to develop their probiotic functions. For this reason, the role of HeLa cell surface proteoglycans in the attachment of Lactobacillus salivarius Lv72, a mutualistic strain of vaginal origin, was investigated. Results Incubation of cultures with a variety of glycosaminoglycans (chondroitin sulfate A and C, heparin and heparan sulfate) resulted in marked binding interference. However, no single glycosaminoglycan was able to completely abolish cell binding, the sum of all having an additive effect that suggests cooperation between them and recognition of specific adhesins on the bacterial surface. In contrast, chondroitin sulfate B enhanced cell to cell attachment, showing the relevance of the stereochemistry of the uronic acid and the sulfation pattern on binding. Elimination of the HeLa surface glycosaminoglycans with lyases also resulted in severe adherence impairment. Advantage was taken of the Lactobacillus-glycosaminoglycans interaction to identify an adhesin from the bacterial surface. This protein, identify as a soluble binding protein of an ABC transporter system (OppA) by MALDI-TOF/(MS), was overproduced in Escherichia coli, purified and shown to interfere with L. salivarius Lv72 adhesion to HeLa cells. Conclusions These data suggest that glycosaminoglycans play a fundamental role in attachment of mutualistic bacteria to the epithelium that lines the cavities where the normal microbiota thrives, OppA being a bacterial adhesin involved in the process. PMID:24044741

  9. Unidirectional Movement of Cellulose Synthase Complexes in Arabidopsis Seed Coat Epidermal Cells Deposit Cellulose Involved in Mucilage Extrusion, Adherence, and Ray Formation1[OPEN

    PubMed Central

    Lam, Patricia; Young, Robin; DeBolt, Seth

    2015-01-01

    CELLULOSE SYNTHASE5 (CESA5) synthesizes cellulose necessary for seed mucilage adherence to seed coat epidermal cells of Arabidopsis (Arabidopsis thaliana). The involvement of additional CESA proteins in this process and details concerning the manner in which cellulose is deposited in the mucilage pocket are unknown. Here, we show that both CESA3 and CESA10 are highly expressed in this cell type at the time of mucilage synthesis and localize to the plasma membrane adjacent to the mucilage pocket. The isoxaben resistant1-1 and isoxaben resistant1-2 mutants affecting CESA3 show defects consistent with altered mucilage cellulose biosynthesis. CESA3 can interact with CESA5 in vitro, and green fluorescent protein-tagged CESA5, CESA3, and CESA10 proteins move in a linear, unidirectional fashion around the cytoplasmic column of the cell, parallel with the surface of the seed, in a pattern similar to that of cortical microtubules. Consistent with this movement, cytological evidence suggests that the mucilage is coiled around the columella and unwinds during mucilage extrusion to form a linear ray. Mutations in CESA5 and CESA3 affect the speed of mucilage extrusion and mucilage adherence. These findings imply that cellulose fibrils are synthesized in an ordered helical array around the columella, providing a distinct structure to the mucilage that is important for both mucilage extrusion and adherence. PMID:25926481

  10. Stress response of adherent cells on a polymer blend surface composed of a segmented polyurethane and MPC copolymers.

    PubMed

    Sawada, Shin-Ichi; Iwasaki, Yasuhiko; Nakabayashi, Nobuo; Ishihara, Kazuhiko

    2006-12-01

    To better understand the effect of 2-methacryloyloxyethyl phosphorylcholine (MPC) copolymer in improving the biocompatibility of segmented polyurethane (SPU), the expression of heat shock protein (HSP) mRNA in HeLa S3 cells adhered on SPU blended with MPC copolymers was measured. Conventionally, MPC copolymers (PMEH) were synthesized by changing the feed ratios of MPC and 2-ethylhexyl methacrylate. X-ray photoelectron spectroscopic analysis of the SPU/PMEH film indicated that the surface concentration of MPC units on the SPU/PMEH film increased with an increase in PMEH composition. HeLa S3 cells were cultured on SPU/PMEH films. The number of adherent cells on the SPU/PMEH films decreased with an increase in the concentration of PMEH. When the PMEH composition was greater than 0.5 wt %, cell adhesion and proliferation decreased markedly. Expressions of HSP27 and HSP47 mRNA were detected using the reverse transcription-polymerase chain reaction (RT-PCR). After incubation for 24 h, both the HSP mRNA expressions in the HeLa S3 cells showed no significant differences among all samples. In HeLa S3 cells that adhered to the SPU film for 48 h, the expressions of HSP27 and HSP47 mRNA increased significantly when compared with those incubated for 24 h. In contrast, the two kinds of mRNA expressions decreased in the HeLa S3 cells that adhered to the SPU/PMEH films for 48 h. From these results, we concluded that PMEH was quite important in suppressing the stress response of adherent HeLa S3 cells. Therefore, SPU/PMEH blend polymers are useful as implantable biomedical materials. PMID:16758458

  11. Human placenta-derived adherent cells improve cardiac performance in mice with chronic heart failure.

    PubMed

    Chen, Hong-Jung; Chen, Chien-Hsi; Chang, Ming-Yao; Tsai, Da-Ching; Baum, Ellen Z; Hariri, Robert; Herzberg, Uri; Hsieh, Patrick C H

    2015-03-01

    Human placenta-derived adherent cells (PDACs) are a culture-expanded, undifferentiated mesenchymal-like population derived from full-term placental tissue, with immunomodulatory, anti-inflammatory, angiogenic, and neuroprotective properties. PDA-001 (cenplacel-L), an intravenous formulation of PDAC cells, is in clinical development for the treatment of autoimmune and inflammatory diseases. We tested the therapeutic effects of PDA-001 in mice with chronic heart failure (CHF). Three weeks after transaortic constriction surgery to induce CHF, the mice underwent direct intramyocardial (IM) or i.v. injection of PDA-001 at a high (0.5 × 10(6) cells per mouse), medium (0.5 × 10(5) cells per mouse), or low (0.5 × 10(4) cells per mouse) dose. The mice were sacrificed 4 weeks after treatment. Echocardiography and ventricular catheterization showed that IM injection of PDA-001 significantly improved left ventricular systolic and diastolic function compared with injection of vehicle or i.v. injection of PDA-001. IM injection of PDA-001 also decreased cardiac fibrosis, shown by trichrome staining in the vicinity of the injection sites. Low-dose treatment showed the best improvement in cardiac performance compared with the medium- and high-dose groups. In another independent study to determine the mechanism of action with bromodeoxyuridine labeling, the proliferation rates of endothelial cells and cardiomyocytes were significantly increased by low or medium IM dose PDA-001. However, no surviving PDA-001 cells were detected in the heart 1 month after injection. In vivo real-time imaging consistently revealed that the PDA-001 cells were detectable only within 2 days after IM injection of luciferase-expressing PDA-001. Together, these results have demonstrated the cardiac therapeutic potential of PDA-001, likely through a paracrine effect. PMID:25673767

  12. Interactions between Periodontal Bacteria and Human Oral Epithelial Cells: Fusobacterium nucleatum Adheres to and Invades Epithelial Cells

    PubMed Central

    Han, Yiping W.; Shi, Wenyuan; Huang, George T.-J.; Kinder Haake, Susan; Park, No-Hee; Kuramitsu, Howard; Genco, Robert J.

    2000-01-01

    Bacteria are causative agents of periodontal diseases. Interactions between oral bacteria and gingival epithelial cells are essential aspects of periodontal infections. Using an in vitro tissue culture model, a selected group of gram-negative anaerobic bacteria frequently associated with periodontal diseases, including Bacteroides forsythus, Campylobacter curvus, Eikenella corrodens, Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia, were examined for their ability to adhere to and invade primary cultures of human gingival epithelial cells (HGEC). The effects of these bacteria on the production of interleukin-8 (IL-8), a proinflammatory chemokine, were also measured. These studies provided an initial demonstration that F. nucleatum adhered to and invaded HGEC and that this was accompanied by high levels of IL-8 secretion from the epithelial cells. The attachment and invasion characteristics of F. nucleatum were also tested using KB cells, an oral epithelial cell line. The invasion was verified by transmission electron microscopy and with metabolic inhibitors. Invasion appeared to occur via a “zipping” mechanism and required the involvement of actins, microtubules, signal transduction, protein synthesis, and energy metabolism of the epithelial cell, as well as protein synthesis by F. nucleatum. A spontaneous mutant, lam, of F. nucleatum, isolated as defective in autoagglutination, was unable to attach to or invade HGEC or KB cells, further indicating the requirement of bacterial components in these processes. Sugar inhibition assays indicated that lectin-like interactions were involved in the attachment of F. nucleatum to KB cells. Investigation of these new virulence phenotypes should improve our understanding of the role of F. nucleatum in periodontal infections. PMID:10816455

  13. A computational model of the response of adherent cells to stretch and changes in substrate stiffness

    PubMed Central

    Lutchen, Kenneth R.; Suki, Béla

    2014-01-01

    Cells in the body exist in a dynamic mechanical environment where they are subject to mechanical stretch as well as changes in composition and stiffness of the underlying extracellular matrix (ECM). However, the underlying mechanisms by which cells sense and adapt to their dynamic mechanical environment, in particular to stretch, are not well understood. In this study, we hypothesized that emergent phenomena at the level of the actin network arising from active structural rearrangements driven by nonmuscle myosin II molecular motors play a major role in the cellular response to both stretch and changes in ECM stiffness. To test this hypothesis, we introduce a simple network model of actin-myosin interactions that links active self-organization of the actin network to the stiffness of the network and the traction forces generated by the network. We demonstrate that such a network replicates not only the effect of changes in substrate stiffness on cellular traction and stiffness and the dependence of rate of force development by a cell on the stiffness of its substrate, but also explains the physical response of adherent cells to transient and cyclic stretch. Our results provide strong indication that network phenomena governed by the active reorganization of the actin-myosin structure plays an important role in cellular mechanosensing and response to both changes in ECM stiffness and externally applied mechanical stretch. PMID:24408996

  14. CsrRS and environmental pH regulate group B streptococcus adherence to human epithelial cells and extracellular matrix.

    PubMed

    Park, Su Eun; Jiang, Shengmei; Wessels, Michael R

    2012-11-01

    Streptococcus agalactiae (group B Streptococcus or GBS) is a common colonizer of the gastrointestinal and genital tracts and an important cause of invasive infections in newborn infants and in adults with predisposing chronic conditions or advanced age. Attachment to epithelial surfaces at mucosal sites is a critical step in the successful colonization of a human host, and regulation of this process is likely to play an important role in both commensalism and dissemination to cause invasive disease. We found that inactivation of the CsrRS (or CovRS) two-component system increased GBS adherence to epithelial cells derived from human vaginal, cervical, and respiratory epithelium, as well as increasing adherence to extracellular matrix proteins and increasing biofilm formation on polystyrene. Neutral (as opposed to acidic) pH enhanced GBS binding to vaginal epithelial cells and to fibrinogen and fibronectin, effects that were partially dependent on CsrRS. The regulatory effects of CsrRS and environmental pH on bacterial adherence correlated with their effects on the expression of multiple surface adhesins, as assessed by quantitative reverse transcription-PCR. We conclude that GBS adherence to epithelial and abiotic surfaces is regulated by the CsrRS two-component system and by environmental pH through their regulatory effects on the expression of bacterial surface adhesins. Dynamic regulation of GBS adherence enhances the organism's adaptability to survival in multiple niches in the human host. PMID:22949550

  15. CsrRS and Environmental pH Regulate Group B Streptococcus Adherence to Human Epithelial Cells and Extracellular Matrix

    PubMed Central

    Park, Su Eun; Jiang, Shengmei

    2012-01-01

    Streptococcus agalactiae (group B Streptococcus or GBS) is a common colonizer of the gastrointestinal and genital tracts and an important cause of invasive infections in newborn infants and in adults with predisposing chronic conditions or advanced age. Attachment to epithelial surfaces at mucosal sites is a critical step in the successful colonization of a human host, and regulation of this process is likely to play an important role in both commensalism and dissemination to cause invasive disease. We found that inactivation of the CsrRS (or CovRS) two-component system increased GBS adherence to epithelial cells derived from human vaginal, cervical, and respiratory epithelium, as well as increasing adherence to extracellular matrix proteins and increasing biofilm formation on polystyrene. Neutral (as opposed to acidic) pH enhanced GBS binding to vaginal epithelial cells and to fibrinogen and fibronectin, effects that were partially dependent on CsrRS. The regulatory effects of CsrRS and environmental pH on bacterial adherence correlated with their effects on the expression of multiple surface adhesins, as assessed by quantitative reverse transcription-PCR. We conclude that GBS adherence to epithelial and abiotic surfaces is regulated by the CsrRS two-component system and by environmental pH through their regulatory effects on the expression of bacterial surface adhesins. Dynamic regulation of GBS adherence enhances the organism's adaptability to survival in multiple niches in the human host. PMID:22949550

  16. Family variables as moderators between beliefs towards medicines and adherence to self-care behaviors and medication in type 2 diabetes.

    PubMed

    Pereira, M Graça; Pedras, Susana; Machado, José Cunha

    2014-06-01

    This study analyzed whether family variables such as marital adjustment, partner support, family coping, and family stress moderated the relationship between negative beliefs about medicines and adherence to self-care behaviors (diet, glucose monitoring, exercise, foot care, and medication), in Type 2 diabetes patients. The sample was composed of 387 individuals with Type 2 diabetes, diagnosed in the past 12 months. Patients were assessed on self-care behaviors in diabetes, medication adherence, beliefs about medicines, family coping, family stress, marital adjustment, and partner support. The results showed marital adjustment, family coping, partner support, and family stress as moderators in the relationship between negative beliefs and adherence. Patients with negative beliefs regarding medicines, but who reported good marital adjustment and family coping were more likely to test their blood glucose; and if they reported low support from their partners were less likely to adhere to their prescribed diet. Finally, patients with negative beliefs about medicines, but who reported high family stress, were less likely to take their medication. The results emphasize the importance of family variables on adherence to self-care behaviors and medication. This study revealed the importance of including partners on interventions regarding Type 2 diabetes because they seem to play an important role in patient's adherence. PMID:24707825

  17. Prevalence of Escherichia coli strains with localized, diffuse, and aggregative adherence to HeLa cells in infants with diarrhea and matched controls.

    PubMed Central

    Gomes, T A; Blake, P A; Trabulsi, L R

    1989-01-01

    To determine the possible role of Escherichia coli strains with three different patterns of adherence to HeLa cells in causing diarrhea in infants in São Paulo, Brazil, we studied stool specimens from 100 infants up to 1 year of age with acute diarrheal illnesses and 100 age-matched control infants without recent diarrhea. E. coli with localized adherence to HeLa cells was much more common in patients (23%) than in controls (2%) (P less than 0.0001) and was detected more frequently than rotavirus (19%) was in patients, even though the study was conducted during the coldest months of the year. Most (80%) of the E. coli colonies with localized adherence were of traditional enteropathogenic E. coli serotypes. Little difference was found between patients and controls in the rate of isolation of E. coli with diffuse adherence (31 and 32%, respectively) or aggregative adherence (10 and 8%, respectively). A genetic probe used to detect a plasmid-mediated adhesin which confers expression of localized adherence proved to be 100% sensitive and 99.9% specific in detecting E. coli with localized adherence to HeLa cells. Although E. coli strains with localized adherence have now been shown to be enteric pathogens in several parts of the world, the role of strains showing diffuse adherence and aggregative adherence is still uncertain. PMID:2563383

  18. Characterization of Binding of Candida albicans to Small Intestinal Mucin and Its Role in Adherence to Mucosal Epithelial Cells

    PubMed Central

    de Repentigny, Louis; Aumont, Francine; Bernard, Karine; Belhumeur, Pierre

    2000-01-01

    In order to approximate and adhere to mucosal epithelial cells, Candida must traverse the overlying mucus layer. Interactions of Candida species with mucin and human buccal epithelial cells (BECs) were thus investigated in vitro. Binding of the Candida species to purified small intestinal mucin showed a close correlation with their hierarchy of virulence. Significant differences (P < 0.05) were found among three categories of Candida species adhering highly (C. dubliniensis, C. tropicalis, and C. albicans), moderately (C. parapsilosis and C. lusitaniae) or weakly (C. krusei and C. glabrata) to mucin. Adherence of C. albicans to BECs was quantitatively inhibited by graded concentrations of mucin. However, inhibition of adherence was reversed by pretreatment of mucin with pronase or C. albicans secretory aspartyl proteinase Sap2p but not with sodium periodate. Saturable concentration- and time-dependent binding of mucin to C. albicans was abrogated by pronase or Sap2p treatment of mucin but was unaffected by β-mercaptoethanol, sodium periodate, neuraminidase, lectins, or potentially inhibitory sugars. Probing of membrane blots of the mucin with C. albicans revealed binding of the yeast to the 66-kDa cleavage product of the 118-kDa C-terminal glycopeptide of mucin. Although no evidence was found for the participation of C. albicans cell surface mannoproteins in specific receptor-ligand binding to mucin, inhibition of binding by p-nitrophenol (1 mM) and tetramethylurea (0.36 M) revealed that hydrophobic interactions are involved in adherence of C. albicans to mucin. These results suggest that C. albicans may both adhere to and enzymatically degrade mucins by the action of Saps, and that both properties may act to modulate Candida populations in the oral cavity and gastrointestinal tract. PMID:10816460

  19. The role of Listeria monocytogenes cell wall surface anchor protein LapB in virulence, adherence, and intracellular replication

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lmof2365_2117 is a Listeria monocytogenes putative cell wall surface anchor protein with a conserved domain found in collagen binding proteins. We constructed a deletion mutation in lmof2365_2117 in serotype 4b strain F2365, evaluated its virulence, and determined its ability to adhere and invade co...

  20. Adherence of Candida albicans to human buccal epithelial cells: host-induced protein synthesis and signaling events.

    PubMed Central

    Bailey, A; Wadsworth, E; Calderone, R

    1995-01-01

    The synthesis of proteins by Candida albicans was studied following adherence of blastoconidia to human buccal epithelial cells (HBEC). Initially, labeling of HBEC, C. albicans, and HBEC-C. albicans with [35S]methionine was performed. After a 3-h incubation and prior to labeling with [35S]methionine, the cultures were treated with cycloheximide to prevent HBEC protein synthesis. The HBEC-C. albicans mixture as well as C. albicans and HBEC incubated separately were extracted with beta-mercaptoethanol (beta-ME). These extracts as well as the cell residue (solubilized by boiling with sodium dodecyl sulfate [SDS]) were examined by SDS-polyacrylamide gel electrophoresis and autoradiography. In comparison to cultures of C. albicans incubated without HBEC, proteins with molecular masses of approximately 52 to 56 kDa from beta-ME extracts and from SDS-solubilized cells were observed only from adhering cultures. In addition, unlabeled beta-ME extracts were electrotransferred to nitrocellulose and immunoblotted with antiphosphotyrosine antibodies to determine whether cell signaling events were occurring during adherence. Proteins with molecular masses of 54 and 60 kDa were recognized only in mixed cultures of C. albicans and HBEC. These data indicate that following adherence of C. albicans to HBEC, new Candida proteins are expressed. Further, these events are accompanied by the expression of signal proteins, presumably of Candida origin. PMID:7822023

  1. Automatic segmentation and quantification of fluorescing microspheres adhering to capillary endothelial cells in the rat lung

    NASA Astrophysics Data System (ADS)

    Albert, Thomas A.; Fingar, Victor H.; Taber, Scott W.; Wieman, Thomas J.

    1995-05-01

    Adhesion molecules present in the cellular membrane of the endothelium provide sites of leukocyte adherence as a first step in the process of leukocyte migration into the interstitium. New evidence suggests the same adhesion proteins may be responsible for the spread of metastatic tumors by providing a location for tumor cell attachment. A method was sought to quantitate the degree of adhesion molecule expression in the pulmonary capillary endothelium using a recently developed animal model which allows for viewing the lung surface in vivo. Videoimages of the pulmonary vascular system were gathered using this new lung chamber technique. A fully automated digital image processing and analysis (DIPA) system was also developed to estimate the level of intercellular adhesion molecule-1 (ICAM-1) expression on the capillary endothelial cells in these videoimages. Fluorescent microspheres were immunologically bound to the ICAM-1 molecules present on the endothelial cell surface. The DIPA system then located and quantified the fluorescent spots present in the videoimages. The ability of this system to locate and measure the fluorescence was compared with human measurements of the same images.

  2. Synergistic role of curli and cellulose in cell adherence and biofilm formation of attaching and effacing Escherichia coli and identification of Fis as a negative regulator of curli

    PubMed Central

    Saldaña, Zeus; Xicohtencatl-Cortes, Juan; Avelino, Fabiola; Phillips, Alan D.; Kaper, James B.; Puente, José L.; Girón, Jorge A.

    2009-01-01

    Summary Curli are adhesive fimbriae of Escherichia coli and Salmonella enterica. Expression of curli (csgA) and cellulose (bcsA) is co-activated by the transcriptional activator CsgD. In this study, we investigated the contribution of curli and cellulose to the adhesive properties of enterohemorragic (EHEC) O157:H7 and enteropathogenic E. coli (EPEC) O127:H6. While single mutations in csgA, csgD, or bcsA in EPEC and EHEC had no dramatic effect on cell adherence, double csgAbcsA mutants were significantly less adherent than the single mutants or wild-type strains to human colonic HT-29 epithelial cells or to cow colon tissue in vitro. Over-expression of csgD (carried on plasmid pCP994) in a csgD mutant, but not in the single csgA or bscA mutants, led to significant increase in adherence and biofilm formation in EPEC and EHEC, suggesting that synchronized over-production of curli and cellulose enhances bacterial adherence. In line with this finding, csgD transcription was activated significantly in the presence of cultured epithelial cells as compared to growth in tissue culture medium. Analysis of the influence of virulence and global regulators in the production of curli in EPEC identified Fis (factor for inversion stimulation) as a, heretofore unrecognized, negative transcriptional regulator of csgA expression. An EPEC E2348/69Δfis produced abundant amounts of curli whereas a double fiscsgD mutant yielded no detectable curli production. Our data suggest that curli and cellulose act in concert to favor host colonization, biofilm formation, and survival in different environments. PMID:19187284

  3. Reduction of non-adherent behaviour in a Mexican-American adolescent with type 2 diabetes.

    PubMed

    Piven, Emily; Duran, Rene

    2014-03-01

    This single-subject research aimed to evaluate the effect of occupation-based activities to improve diabetes self-management skills in a non-adherent 19-year-old Mexican-American adolescent transitioning to young adulthood. Using a pre-test/post-test design, the subject's performance was re-evaluated with five standardized measures following an 8-week intervention. The subject made major improvements on the Diabetes Self-Efficacy Scale, Exercise Behaviour and in goal attainment of targeted behaviours on the basis of the Canadian Occupational Performance Measure. The Adapted Intrusiveness Rating Scale and the Social/Role Activities Limitations Scale revealed increased intrusiveness of diabetes in his life, once he finally embraced his need to prioritize diabetes self-care. The study illuminated how a culturally sensitive, occupation-based early intervention might potentially prevent or reduce debilitating complications in adulthood. The value of this study is its contribution to body of diabetes literature on the role of occupational therapist in secondary prevention with Mexican-Americans. Research suggestions included expansion of single-subject design with larger samples and higher-level research studies with adolescents from various cultural backgrounds. PMID:24532099

  4. Adherence of human mesenchymal stem cells on Ti and TiO2 nano-columnar surfaces fabricated by glancing angle sputter deposition

    NASA Astrophysics Data System (ADS)

    Motemani, Yahya; Greulich, Christina; Khare, Chinmay; Lopian, Michael; Buenconsejo, Pio John S.; Schildhauer, Thomas A.; Ludwig, Alfred; Köller, Manfred

    2014-02-01

    The interaction of human mesenchymal stem cells (hMSCs) with Ti and TiO2 nano-columnar surfaces fabricated using glancing angle sputter deposition was investigated. The adherence and proliferation of hMSCs on different nano-columnar surfaces, including vertical columns, slanted columns and chevrons, were examined with calcein-acetoxymethyl ester fluorescence staining and scanning electron microscopy. For comparison, adherence of hMSCs on compact, dense films was also studied. After 24 h and 7 days, adherent and viable cells were observed on both, Ti nano-columns as well as dense Ti films, which confirms the biocompatibility of these nanostructures. Very small pseudopodia with width of approximately 20-35 nm and length varying from 20 to 200 nm were observed between the nano-columns, independent of the type of the nano-columnar morphology. Large inter-column spacing and effectively increased surface area make these nanostructures promising candidates for bio-functionalization or drug loading on the surface of Ti-based implants.

  5. Cell surface adhesiveness of mouse sarcoma lines evaluated by latex particle adherence assay: correlation with growth behavior and electrophoretic mobility.

    PubMed

    Bubeník, J; Jandlová, T; Suhajová, E; Malkovský, M

    1979-01-01

    Using the latex particle adherence assay and five mouse sarcoma cell lines of the identical origin, etiology and genotype but differing in malignancy we attempted to correlate the degree of cell surface adhesiveness with growth behavior and electrophoretic mobility of cells. Higher tumorigenicity of four of the cell lines (Mc11--Mc14) was associated with lower cell surface adhesiveness and, conversely, lower malignancy of the fifth line (Mc15) with higher cell surface adhesiveness. No simple correlation or causal relationship was found among the electrophoretic mobility of the lines and other cellular characteristics. PMID:522921

  6. Platelet activating factor amplifies human neutrophil adherence to bovine endothelial cells: evidence for a lipoxygenase dependent mechanism.

    PubMed

    Damtew, B; Spagnuolo, P J

    1992-10-01

    Platelet activating factor (PAF) is a potent lipid mediator that induces the release of leukotrienes and prostaglandins from various cells and tissues. We examined the capacity of PAF alone and in combination with soluble stimuli to enhance eicosanoid synthesis and adherence of human neutrophils. Neutrophils were preincubated with PAF and washed before exposure to the soluble stimuli F-Met-Leu-Phe (FMLP), calcium ionophore A23187, and phorbol myristate acetate. Preincubation of neutrophils with 1 microM PAF enhanced the release of both LTB4 and LTC4 in response to each of the three agonists, in contrast with the unprimed neutrophils. Priming was specific for PAF since lyso-PAF was inactive. Priming concentrations of PAF also augmented the adherence of neutrophils to endothelium in the presence of the soluble agonists A23187, phorbol myristate acetate, and FMLP. The priming effect of PAF on eicosanoid release and neutrophil adherence was shown to have similar time- and dose-dependent effects. Further, the priming effects of PAF on adherence could be reversed by preincubation of neutrophils with the lipoxygenase inhibitors nordihydroguiaretic acid and 5,8,11,14-ETYA but not by preincubation with the cyclooxygenase inhibitor indomethacin. These data demonstrate that PAF amplifies neutrophil adherence to endothelium through a lipoxygenase dependent mechanism. PMID:1330924

  7. Role of M3 protein in the adherence and internalization of an invasive Streptococcus pyogenes strain by epithelial cells.

    PubMed

    Eyal, Osnat; Jadoun, Jeries; Bitler, Arcady; Skutelski, Ehud; Sela, Shlomo

    2003-10-15

    Streptococcus pyogenes utilizes multiple mechanisms for adherence to and internalization by epithelial cells. One of the molecules suggested of being involved in adherence and internalization is the M protein. Although strains of the M3 serotype form the second largest group isolated from patients with severe invasive diseases and fatal infections, not much information is known regarding the interactions of M3 protein with mammalian cells. In this study we have constructed an emm3 mutant of an invasive M3 serotype (SP268), and demonstrated that the M3 protein is involved in both adherence to and internalization by HEp-2 cells. Fibronectin promoted both adherence and internalization of SP268 in an M3-independent pathway. Utilizing speB and speB/emm3 double mutants, it was found that M3 protein is not essential for the maturation of SpeB, as was reported for the M1 protein. Increased internalization efficiency observed in both the speB and emm3/speB mutants suggested that inhibition of S. pyogenes internalization by SpeB is not related to the presence of an intact M3 protein. Thus, other proteins in SP268, which serve as targets for SpeB activity, have a prominent role in the internalization process. PMID:14522456

  8. Fimbria-mediated adherence of Candida albicans to glycosphingolipid receptors on human buccal epithelial cells.

    PubMed Central

    Yu, L; Lee, K K; Sheth, H B; Lane-Bell, P; Srivastava, G; Hindsgaul, O; Paranchych, W; Hodges, R S; Irvin, R T

    1994-01-01

    Candida albicans is an opportunist fungal pathogen that has the ability to adhere to host cell surface receptors via a number of adhesins. Yu et al. (L. Yu, K. K. Lee, K. Ens, P. C. Doig, M. R. Carpenter, W. Staddon, R. S. Hodges, W. Paranchych, and R. T. Irvin, Infect. Immun. 62:2834-2842, 1994) described the purification and initial characterization of a fimbrial adhesin from C. albicans. In this paper, we show that C. albicans fimbriae also bind to asialo-GM1 [gangliotetraosylceramide: beta Gal(1-3)beta GalNAc(1-4) beta Gal(1-4)beta Glc(1-1)Cer] immobilized on microtiter plates in a saturable and concentration-dependent manner. C. albicans fimbrial binding to exfoliated human buccal epithelial cells (BECs) was inhibited by asialo-GM1 in in vitro binding assays. The fimbriae interact with the glycosphingolipid receptors via the carbohydrate portion of the receptors, since fimbriae were observed to bind to synthetic beta GalNAc(1-4)beta Gal-protein conjugates and the disaccharide was able to inhibit binding of fimbriae to BECs in in vitro binding assays. We conclude from these results that the C. albicans yeast form expresses a fimbrial adhesin that binds to glycosphingolipids displayed on the surface of human BECs. Images PMID:8005674

  9. Protein phosphatase 2A activity is required for functional adherent junctions in endothelial cells.

    PubMed

    Kása, Anita; Czikora, István; Verin, Alexander D; Gergely, Pál; Csortos, Csilla

    2013-09-01

    Reversible Ser/Thr phosphorylation of cytoskeletal and adherent junction (AJ) proteins has a critical role in the regulation of endothelial cell (EC) barrier function. We have demonstrated earlier that protein phosphatase 2A (PP2A) activity is important in EC barrier integrity. In the present work, macro- and microvascular EC were examined and we provided further evidence on the significance of PP2A in the maintenance of EC cytoskeleton and barrier function with special focus on the Bα (regulatory) subunit of PP2A. Immunofluorescent staining revealed that the inhibition of PP2A results in changes in the organization of EC cytoskeleton as microtubule dissolution and actin re-arrangement were detected. Depletion of Bα regulatory subunit of PP2A had similar effect on the cytoskeleton structure of the cells. Furthermore, transendothelial electric resistance measurements demonstrated significantly slower barrier recovery of Bα depleted EC after thrombin treatment. AJ proteins, VE-cadherin and β-catenin, were detected along with Bα in pull-down assay. Also, the inhibition of PP2A (by okadaic acid or fostriecin) or depletion of Bα caused β-catenin translocation from the membrane to the cytoplasm in parallel with its phosphorylation on Ser552. In conclusion, our data suggest that the A/Bα/C holoenzyme form of PP2A is essential in EC barrier integrity both in micro- and macrovascular EC. PMID:23721711

  10. Influence of patients’ disease knowledge and beliefs about medicines on medication adherence: findings from a cross-sectional survey among patients with type 2 diabetes mellitus in Palestine

    PubMed Central

    2014-01-01

    Background Diabetes mellitus (DM) is a common serious health problem. Medication adherence is a key determinant of therapeutic success in patients with diabetes mellitus. The purpose of this study was to assess medication adherence and its potential association with beliefs and diabetes – related knowledge in patients with type II DM. Methods This study was carried out at Al-Makhfia governmental diabetes primary healthcare clinic in Nablus, Palestine. Main outcome of interest in the study was medication adherence. The Beliefs about Medicines Questionnaire (BMQ) was used to assess beliefs. Morisky Medication Adherence Scale (MMSA-8©) was used to assess medication adherence. The Michigan diabetes knowledge test (MDKT) was used to assess diabetes – related knowledge. Univariate and multivariate analysis were carried out using Statistical Package for Social Sciences (SPSS 20). Results Four hundred and five patients were interviewed. The mean ± SD age of the participants was 58.3 ± 10.4 (range = 28 – 90) years. More than half (53.3%) of the participants were females. Approximately 42.7% of the study sample were considered non-adherent (MMAS-8© score of < 6). Multivariate analysis showed that the following variables were significantly associated with non-adherence: disease-related knowledge, beliefs about necessity of anti-diabetic medications, concerns about adverse consequences of anti-diabetic medications and beliefs that medicines in general are essentially harmful. Diabetic patients with high knowledge score and those with strong beliefs in the necessity of their anti-diabetic medications were less likely to be non-adherent ([O.R = 0.87, 95% CI of 0.78 – 0.97] and [O.R = 0.93, 95% of 0.88 – 0.99] respectively). However, diabetic patients with high concerns about adverse consequences of anti-diabetic medications and those with high belief that all medicines are harmful were more likely to be non-adherent ([O.R = 1.09; 95% C

  11. Evaluation of a Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) assay (SOT)

    EPA Science Inventory

    The Embryonic Stem Cell Test (EST) has been used to evaluate the effects of xenobiotics using three endpoints, stem cell differentiation, stem cell viability and 3T3-cell viability. Our research goal is to establish amodel system that would evaluate chemical effects using a singl...

  12. Urokinase receptor-dependent and -independent p56/59(hck) activation state is a molecular switch between myelomonocytic cell motility and adherence.

    PubMed Central

    Chiaradonna, F; Fontana, L; Iavarone, C; Carriero, M V; Scholz, G; Barone, M V; Stoppelli, M P

    1999-01-01

    Anchorage-independent myelomonocytic cells acquire adherence within minutes of differentiation stimuli, such as the proteolytically inactive N-terminal fragment of urokinase binding to its cognate glycosylphosphatidylinositol (GPI)-anchored receptor. Here, we report that urokinase-treated differentiating U937 monocyte-like cells exhibit a rapid and transient inhibition of p56/59(hck) and p55(fgr) whereas no changes in the activity of other Src family kinases, such as p53/56(lyn) and p59(fyn) were observed. U937 transfectants expressing a kinase-defective (Lys267 to Met) p56/59(hck) variant exhibit enhanced adhesiveness and a marked F-actin redistribution in thin protruding structures. Conversely, urokinase as well as expression of wild-type or constitutively active (Tyr499 to Phe) p56/59(hck) stimulates the directional migration of uninduced U937 cells. Accordingly, expression of constitutively active or kinase inactive p56/59(hck) selectively prevents urokinase receptor-dependent induction of either adhesion or motility, indicating that a specific activation state of p56/59(hck) is required for each cell response. In conclusion, modulation of the intracellular p56/59(hck) tyrosine kinase activity switches cell motility towards adherence, providing a mutually exclusive mechanism to regulate these properties during monocyte/macrophage differentiation in vivo. PMID:10357814

  13. Butyrate modulates bacterial adherence on LS174T human colorectal cells by stimulating mucin secretion and MAPK signaling pathway

    PubMed Central

    Jung, Tae-Hwan; Park, Jeong Hyeon; Han, Kyoung-Sik

    2015-01-01

    BACKGROUND/OBJECTIVES Fermentation of dietary fiber results in production of various short chain fatty acids in the colon. In particular, butyrate is reported to regulate the physical and functional integrity of the normal colonic mucosa by altering mucin gene expression or the number of goblet cells. The objective of this study was to investigate whether butyrate modulates mucin secretion in LS174T human colorectal cells, thereby influencing the adhesion of probiotics such as Lactobacillus and Bifidobacterium strains and subsequently inhibiting pathogenic bacteria such as E. coli. In addition, possible signaling pathways involved in mucin gene regulation induced by butyrate treatment were also investigated. MATERIALS/METHODS Mucin protein content assay and periodic acid-Schiff (PAS) staining were performed in LS174T cells treated with butyrate at various concentrations. Effects of butyrate on the ability of probiotics to adhere to LS174T cells and their competition with E. coli strains were examined. Real time polymerase chain reaction for mucin gene expression and Taqman array 96-well fast plate-based pathway analysis were performed on butyrate-treated LS174T cells. RESULTS Treatment with butyrate resulted in a dose-dependent increase in mucin protein contents in LS174T cells with peak effects at 6 or 9 mM, which was further confirmed by PAS staining. Increase in mucin protein contents resulted in elevated adherence of probiotics, which subsequently reduced the adherent ability of E. coli. Treatment with butyrate also increased transcriptional levels of MUC3, MUC4, and MUC12, which was accompanied by higher gene expressions of signaling kinases and transcription factors involved in mitogen-activated protein kinase (MAPK) signaling pathways. CONCLUSIONS Based on our results, butyrate is an effective regulator of modulation of mucin protein production at the transcriptional and translational levels, resulting in changes in the adherence of gut microflora. Butyrate

  14. Glucosyltransferases of Viridans Group Streptococci Modulate Interleukin-6 and Adhesion Molecule Expression in Endothelial Cells and Augment Monocytic Cell Adherence

    PubMed Central

    Yeh, Chiou-Yueh; Chen, Jen-Yang; Chia, Jean-San

    2006-01-01

    Recruitment of monocytes plays important roles during vegetation formation and endocardial inflammation in the pathogenesis of infective endocarditis (IE). Bacterial antigens or modulins can activate endothelial cells through the expression of cytokines or adhesion molecules and modulate the recruitment of leukocytes. We hypothesized that glucosyltransferases (GTFs), modulins of viridans group streptococci, may act directly to up-regulate the expression of adhesion molecules and also interleukin-6 (IL-6) to augment monocyte attachment to endothelial cells. Using primary cultured human umbilical vein endothelial cells (HUVECs) as an in vitro model, we demonstrated that GTFs (in the cell-bound or free form) could specifically modulate the expression of IL-6, and also adhesion molecules, in a dose- and time-dependent manner. Results of inhibition assays suggested that enhanced expression of adhesion molecules was dependent on the activation of nuclear factor κB (NF-κB) and extracellular signal-regulated kinase and that p38 mitogen-activated protein kinase pathways also contributed to the release of IL-6. Streptococcus-infected HUVECs or treatment with purified IL-6 plus soluble IL-6 receptor α enhanced the expression of ICAM-1 and the adherence of the monocytic cell line U937. These results suggest that streptococcal GTFs might play an important role in recruiting monocytic cells during inflammation in IE through induction of adhesion molecules and IL-6, a cytokine involved in transition from neutrophil to monocyte recruitment. PMID:16428777

  15. Impact of the type of mask on the effectiveness of and adherence to continuous positive airway pressure treatment for obstructive sleep apnea*

    PubMed Central

    de Andrade, Rafaela Garcia Santos; Piccin, Vivien Schmeling; Nascimento, Juliana Araújo; Viana, Fernanda Madeiro Leite; Genta, Pedro Rodrigues; Lorenzi-Filho, Geraldo

    2014-01-01

    Continuous positive airway pressure (CPAP) is the gold standard for the treatment of obstructive sleep apnea (OSA). Although CPAP was originally applied with a nasal mask, various interfaces are currently available. This study reviews theoretical concepts and questions the premise that all types of interfaces produce similar results. We revised the evidence in the literature about the impact that the type of CPAP interface has on the effectiveness of and adherence to OSA treatment. We searched the PubMed database using the search terms "CPAP", "mask", and "obstructive sleep apnea". Although we identified 91 studies, only 12 described the impact of the type of CPAP interface on treatment effectiveness (n = 6) or adherence (n = 6). Despite conflicting results, we found no consistent evidence that nasal pillows and oral masks alter OSA treatment effectiveness or adherence. In contrast, most studies showed that oronasal masks are less effective and are more often associated with lower adherence and higher CPAP abandonment than are nasal masks. We concluded that oronasal masks can compromise CPAP OSA treatment adherence and effectiveness. Further studies are needed in order to understand the exact mechanisms involved in this effect. PMID:25610507

  16. Correlative VIS-fluorescence and soft X-ray cryo-microscopy/tomography of adherent cells.

    PubMed

    Hagen, Christoph; Guttmann, Peter; Klupp, Barbara; Werner, Stephan; Rehbein, Stefan; Mettenleiter, Thomas C; Schneider, Gerd; Grünewald, Kay

    2012-02-01

    Soft X-ray cryo-microscopy/tomography of vitreous samples is becoming a valuable tool in structural cell biology. Within the 'water-window' wavelength region (2.34-4.37nm), it provides absorption contrast images with high signal to noise ratio and resolution of a few tens of nanometer. Soft X-rays with wavelengths close to the K-absorption edge of oxygen penetrate biological samples with thicknesses in the micrometer range. Here, we report on the application of a recently established extension of the transmission soft X-ray cryo-microscope (HZB TXM) at the beamline U41-XM of the BESSY II electron storage ring by an in-column epi-fluorescence and reflected light cryo-microscope. We demonstrate the new capability for correlative fluorescence and soft X-ray cryo-microscopy/tomography of this instrument along a typical life science experimental approach - the correlation of a fluorophore-tagged protein (pUL34-GFP of pseudorabies virus, PrV, the nuclear membrane-anchored component of the nuclear egress complex of the Herpesviridae which interacts with viral pUL31) in PrV pUL34-GFP/pUL31 coexpressing mammalian cells, with virus-induced vesicular structures in the nucleus, expanding the nucleoplasmic reticulum. Taken together, our results demonstrate new possibilities to study the role of specific proteins in substructures of adherent cells, especially of the nucleus in toto, accessible to electron microscopy in thinned samples only. PMID:22210307

  17. Correlative VIS-fluorescence and soft X-ray cryo-microscopy/tomography of adherent cells

    PubMed Central

    Hagen, Christoph; Guttmann, Peter; Klupp, Barbara; Werner, Stephan; Rehbein, Stefan; Mettenleiter, Thomas C.; Schneider, Gerd; Grünewald, Kay

    2012-01-01

    Soft X-ray cryo-microscopy/tomography of vitreous samples is becoming a valuable tool in structural cell biology. Within the ‘water-window’ wavelength region (2.34–4.37 nm), it provides absorption contrast images with high signal to noise ratio and resolution of a few tens of nanometer. Soft X-rays with wavelengths close to the K-absorption edge of oxygen penetrate biological samples with thicknesses in the micrometer range. Here, we report on the application of a recently established extension of the transmission soft X-ray cryo-microscope (HZB TXM) at the beamline U41-XM of the BESSY II electron storage ring by an in-column epi-fluorescence and reflected light cryo-microscope. We demonstrate the new capability for correlative fluorescence and soft X-ray cryo-microscopy/tomography of this instrument along a typical life science experimental approach – the correlation of a fluorophore-tagged protein (pUL34-GFP of pseudorabies virus, PrV, the nuclear membrane-anchored component of the nuclear egress complex of the Herpesviridae which interacts with viral pUL31) in PrV pUL34-GFP/pUL31 coexpressing mammalian cells, with virus-induced vesicular structures in the nucleus, expanding the nucleoplasmic reticulum. Taken together, our results demonstrate new possibilities to study the role of specific proteins in substructures of adherent cells, especially of the nucleus in toto, accessible to electron microscopy in thinned samples only. PMID:22210307

  18. Plasmids in Yersinia enterocolitica serotypes O:3 and O:9: correlation with epithelial cell adherence in vitro.

    PubMed Central

    Vesikari, T; Nurmi, T; Mäki, M; Skurnik, M; Sundqvist, C; Granfors, K; Grönroos, P

    1981-01-01

    Human isolates of Yersinia enterocolitica serotypes O:3 (biotype 4) and O:9 (biotype 3) harbored plasmids sized approximately 47 and 44 megadaltons, respectively. No such plasmids were found in "apathogenic" strains of Y. enterocolitica belonging to biotype 1. There was a positive correlation among the presence of plasmid, autoagglutination, and adherence to and toxicity for HEp-2 cell cultures; all of these properties were lost by culturing at 37 degrees C in the absence of calcium. Strains of Y. enterocolitica O:3 and O:9 cured of the plasmids showed increased invasiveness in the HEp-2 cell culture model, but no invasiveness in guinea pig eye. It is suggested that the plasmids of Y. enterocolitica primarily determine epithelial cell adherence, but may also be associated with other pathogenic properties. Images PMID:7287174

  19. Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) assay

    EPA Science Inventory

    The Embryonic Stem Cell Test (EST) is an assay which evaluates xenobiotic-induced effects using three endpoints: mouse embryonic stem cell (mESC) differentiation, mESC viability, and 3T3-cell viability. Our research goal was to develop an improved high-throughput assay by establi...

  20. A Systematic Review of Interventions Addressing Adherence to Anti-Diabetic Medications in Patients with Type 2 Diabetes—Components of Interventions

    PubMed Central

    Sapkota, Sujata; Brien, Jo-anne E.; Greenfield, Jerry R.; Aslani, Parisa

    2015-01-01

    Background Poor adherence to anti-diabetic medications contributes to suboptimal glycaemic control in patients with type 2 diabetes (T2D). A range of interventions have been developed to promote anti-diabetic medication adherence. However, there has been very little focus on the characteristics of these interventions and how effectively they address factors that predict non-adherence. In this systematic review we assessed the characteristics of interventions that aimed to promote adherence to anti-diabetic medications. Method Using appropriate search terms in Medline, Embase, CINAHL, International Pharmaceutical Abstracts (IPA), PUBmed, and PsychINFO (years 2000–2013), we identified 52 studies which met the inclusion criteria. Results Forty-nine studies consisted of patient-level interventions, two provider-level interventions, and one consisted of both. Interventions were classified as educational (n = 7), behavioural (n = 3), affective, economic (n = 3) or multifaceted (a combination of the above; n = 40). One study consisted of two interventions. The review found that multifaceted interventions, addressing several non-adherence factors, were comparatively more effective in improving medication adherence and glycaemic target in patients with T2D than single strategies. However, interventions with similar components and those addressing similar non-adherence factors demonstrated mixed results, making it difficult to conclude on effective intervention strategies to promote adherence. Educational strategies have remained the most popular intervention strategy, followed by behavioural, with affective components becoming more common in recent years. Most of the interventions addressed patient-related (n = 35), condition-related (n = 31), and therapy-related (n = 20) factors as defined by the World Health Organization, while fewer addressed health care system (n = 5) and socio-economic-related factors (n = 13). Conclusion There is a noticeable shift in the literature

  1. Adherence of Clostridium thermocellum to cellulose.

    PubMed Central

    Bayer, E A; Kenig, R; Lamed, R

    1983-01-01

    The adherence of Clostridium thermocellum, a cellulolytic, thermophilic anaerobe, to its insoluble substrate (cellulose) was studied. The adherence phenomenon was determined to be selective for cellulose. The observed adherence was not significantly affected by various parameters, including salts, pH, temperature, detergents, or soluble sugars. A spontaneous adherence-defective mutant strain (AD2) was isolated from the wild-type strain YS. Antibodies were prepared against the bacterial cell surface and rendered specific to the cellulose-binding factor (CBF) by adsorption to mutant AD2 cells. By using these CBF-specific antibodies, crossed immunoelectrophoresis of cell extracts revealed a single discrete precipitation peak in the parent strain which was absent in the mutant. This difference was accompanied by an alteration in the polypeptide profile whereby sonicates of strain YS contained a 210,000-molecular-weight band which was missing in strain AD2. The CBF antigen could be removed from cell extracts by adsorption to cellulose. A combined gel-overlay--immunoelectrophoretic technique demonstrated that the cellulose-binding properties of the CBF were accompanied by carboxymethylcellulase activity. During the exponential phase of growth, a large part of the CBF antigen and related carboxymethylcellulase activity was associated with the cells of wild-type strain YS. However, the amounts decreased in stationary-phase cells. Cellobiose-grown mutant AD2 cells lacked the cell-associated CBF, but the latter was detected in the extracellular fluid. Increased levels of CBF were observed when cells were grown on cellulose. In addition, mutant AD2 regained cell-associated CBF together with the property of cellulose adherence. The presence of the CBF antigen and related adherence characteristics appeared to be a phenomenon common to other naturally occurring strains of this species. Images PMID:6630152

  2. An adherent cell perifusion technique to study the overall and sequential response of rat alveolar macrophages to toxic substances.

    PubMed Central

    Forget, G; Lacroix, M J; Cadieux, A; Calvert, R; Grose, J H; Sirois, P

    1983-01-01

    Essentially pure (97%) alveolar macrophages were isolated by bronchoalveolar lavage of rats with warm (37 degrees C) PBS solution. These cells were allowed to adhere to the inside walls of open-ended glass cylinders which were closed off at each end by three-way stopcocks. The adhering cells were perifused with RPMI-1640 medium supplemented with 5% fetal bovine serum for 18 hr at the rate of 1 mL/hr, and the effluent medium was collected automatically in 2-mL aliquots. Cell recoveries and viabilities did not differ from those found for Petri cultures treated similarly, indicating that the perifusion method under study offered an adequate milieu for short-term primary cultures. The alveolar macrophages in culture were subjected to the presence of particulate (chrysotile asbestos) and soluble (phorbol myristate) toxicants, and their response was monitored in the effluent medium by measuring the release of prostaglandins (PGE) by radioimmunoassay. A significant increase in the sequential release of PGE was observed in the presence of asbestos (100 micrograms/mL) or phorbol myristate (200 ng/mL). Treatment of the cells with indomethacin (20 microM) completely abolished the release of PGE stimulated with phorbol myristate. A cumulative response to the toxicants was also observed when cells were harvested manually from the chambers: asbestos caused a 2-fold increase in cell mortality relative to control, while phorbol myristate brought about a 3-fold increase in the number of dead cells. This effect was not prevented by the presence of indomethacin. Cell aggregation was also observed when cells were perifused in the presence of phorbol myristate, whether indomethacin was present or absent. Our results indicate that the cell perifusion system combines the advantages of conventional adherent cell cultures (viability, aggregation) with those of perifusion techniques (sequential metabolism studies). Images FIGURE 1. FIGURE 2. FIGURE 3. FIGURE 6. PMID:6641651

  3. Distinguishing cell type using epigenotype

    NASA Astrophysics Data System (ADS)

    Wytock, Thomas; Motter, Adilson E.

    Recently, researchers have proposed that unique cell types are attractors of their epigenetic dynamics including gene expression and chromatin conformation patterns. Traditionally, cell types have been classified by their function, morphology, cytochemistry, and other macroscopically observable properties. Because these properties are the result of many proteins working together, it should be possible to predict cell types from gene expression or chromatin conformation profiles. In this talk, I present a maximum entropy approach to identify and distinguish cell type attractors on the basis of correlations within these profiles. I will demonstrate the flexibility of this method through its separate application to gene expression and chromatin conformation datasets. I show that our method out-performs other machine-learning techniques and uncorrelated benchmarks. We adapt our method to predict growth rate from gene expression in E. coli and S. cerevisiae and compare our predictions with those from metabolic models. In addition, our method identifies a nearly convex region of state-space associated with each cell type attractor basin. Estimates of the growth rate and attractor basin make it possible to rationally control gene regulatory networks independent of a model. This research was supported by NSF-GRFP, NSF-GK12, GAANN, and Northwestern's NIH-NIGMS Molecular Biophysics Training Grant.

  4. The adherence of endothelial cells to Dacron induces the expression of the intercellular adhesion molecule (ICAM-1).

    PubMed Central

    Margiotta, M S; Robertson, F S; Greco, R S

    1992-01-01

    The intercellular adhesion molecule (ICAM-1) is a glycoprotein expressed by endothelial cells activated by cytokines. The lymphocyte-function-associated antigen (LFA-1) is an integrin expressed by activated white blood cells. Together, this receptor-ligand pair is responsible, in part, for the localization of neutrophils at sites of inflammation. Using an in vitro model, the authors studied the binding of antibodies against ICAM-1 by human saphenous vein endothelial cells (HSVEC) adherent to Dacron and control cultureware. After adherence to Dacron pretreated with fibronectin, 24% more HSVEC-bound antibody against ICAM-1 compared with HSVEC on controls. In contrast, 90% more HSVEC adherent to Dacron incubated with whole blood bound anti-ICAM-1 antibodies. These cells bound 17.7-fold greater amounts of antibody compared with HSVEC on controls. Pretreating Dacron with plasma resulted in no increase in antibody binding compared with control. Our studies suggest that the cellular components of blood in contact with Dacron create a microenvironment that activates HSVEC and enhances ICAM-1 expression. Induction of this adhesion molecule may play a pivotal role in the migration and localization of leukocytes at the site of the vascular prosthesis. PMID:1359845

  5. Management of type 2 diabetes mellitus: Adherence challenges in environments of low socio-economic status

    PubMed Central

    Phillips, Julie

    2014-01-01

    Abstract Background The efficacy of treatment for clients with diabetes is highly dependent on the individual's ability to manage the disease. Several constraints, such as poverty, illiteracy and insufficient resources (finances and specialised healthcare professionals), especially communities of low socio-economic status, could influence clients’ ability to manage their disease. Aim The main aim of this study was to outline the obstacles encountered by individuals with type 2 diabetes mellitus from an urban community with regard to management of their disease. Setting The study was conducted at a primary health care facility in the Western Cape, South Africa. Methods Ethical clearance was obtained from all relevant authorities. Eight (8) conveniently selected clients with type 2 diabetes mellitus per participating community healthcare centre (six approved centres in total) were invited to take part in focus group discussions. Twenty six clients, 15 females and 11 males, with a mean age of 58.92 years (SD = 7.33), agreed to participate. Audiotaped data were transcribed verbatim followed by content analysis and identification of themes. Results Themes that emerged were challenges with: a healthy eating plan, physical activity, financial constraints, other people's understanding of the disease, and service received at the community healthcare centre. Verbatim quotes were used to exemplify the themes. Conclusion Clients with type 2 diabetes mellitus experienced several challenges in the management of their disease. These challenges should be addressed to assist with better glycaemic control and to curb the emergence of diabetic complications and their attendant cost implications. PMID:26245413

  6. Adherence to GLP-1 receptor agonist therapy administered by once-daily or once-weekly injection in patients with type 2 diabetes in Germany

    PubMed Central

    Qiao, Qing; Ouwens, Mario JNM; Grandy, Susan; Johnsson, Kristina; Kostev, Karel

    2016-01-01

    Aim This study aimed to compare 6-month adherence to therapy with exenatide once weekly (Bydureon®) vs liraglutide once daily (Victoza®) in patients with type 2 diabetes under primary care in Germany. Methods A nationwide longitudinal prescription database (LRx), (between January 2011 and September 2014) was used to analyze adherence to therapy. The proportion of days covered (PDC) by prescription was used as a measure of adherence in the 6-month postindex period. Logistic regression analyses were performed to investigate the associations between glucagon-like peptide-1 receptor agonist therapy adjusting for age, sex, and cotherapy. Results Therapy was initiated in 5,449 patients with exenatide once weekly (age: 59.7±11.8 years; 51.4% were male) and in 24,648 patients with liraglutide once daily (age: 59.4±11.4 years; 49.7% were male). The median PDC was 0.88 for exenatide once weekly and 0.77 for liraglutide once daily (P<0.05). Once-weekly exenatide was associated with significantly higher adherence. Odds ratio (95% confidence interval) for having a PDC of ≥0.80 was 1.78 (1.62–1.96) for exenatide once weekly compared with liraglutide once daily after adjusting for age, sex, and cotherapy. Conclusion Adherence to treatment with exenatide once weekly was significantly increased compared to that with liraglutide once daily over 6 months in patients with type 2 diabetes. PMID:27418849

  7. A family of cell-adhering peptides homologous to fibrinogen C-termini

    SciTech Connect

    Levy-Beladev, Liron; Levdansky, Lilia; Gaberman, Elena; Friedler, Assaf; Gorodetsky, Raphael

    2010-10-08

    Research highlights: {yields} Cell-adhesive sequences homologous to fibrinogen C-termini exist in other proteins. {yields} The extended homologous cell-adhesive C-termini peptides family is termed Haptides. {yields} In membrane-like environment random coiled Haptides adopt a helical conformation. {yields} Replacing positively charged residues with alanine reduces Haptides activity. -- Abstract: A family of cell-adhesive peptides homologous to sequences on different chains of fibrinogen was investigated. These homologous peptides, termed Haptides, include the peptides C{beta}, preC{gamma}, and C{alpha}E, corresponding to sequences on the C-termini of fibrinogen chains {beta}, {gamma}, and {alpha}E, respectively. Haptides do not affect cell survival and rate of proliferation of the normal cell types tested. The use of new sensitive assays of cell adhesion clearly demonstrated the ability of Haptides, bound to inert matrices, to mediate attachment of different matrix-dependent cell types including normal fibroblasts, endothelial, and smooth muscle cells. Here we present new active Haptides bearing homologous sequences derived from the C-termini of other proteins, such as angiopoietin 1 and 2, tenascins C and X, and microfibril-associated glycoprotein-4. The cell adhesion properties of all the Haptides were found to be associated mainly with their 11 N-terminal residues. Mutated preC{gamma} peptides revealed that positively charged residues account for their attachment effect. These results suggest a mechanism of direct electrostatic interaction of Haptides with the cell membrane. The extended Haptides family may be applied in modulating adhesion of cells to scaffolds for tissue regeneration and for enhancement of nanoparticulate transfection into cells.

  8. Characterization of Three-Dimensional Retinal Tissue Derived from Human Embryonic Stem Cells in Adherent Monolayer Cultures.

    PubMed

    Singh, Ratnesh K; Mallela, Ramya K; Cornuet, Pamela K; Reifler, Aaron N; Chervenak, Andrew P; West, Michael D; Wong, Kwoon Y; Nasonkin, Igor O

    2015-12-01

    Stem cell-based therapy of retinal degenerative conditions is a promising modality to treat blindness, but requires new strategies to improve the number of functionally integrating cells. Grafting semidifferentiated retinal tissue rather than progenitors allows preservation of tissue structure and connectivity in retinal grafts, mandatory for vision restoration. Using human embryonic stem cells (hESCs), we derived retinal tissue growing in adherent conditions consisting of conjoined neural retina and retinal pigment epithelial (RPE) cells and evaluated cell fate determination and maturation in this tissue. We found that deriving such tissue in adherent conditions robustly induces all eye field genes (RX, PAX6, LHX2, SIX3, SIX6) and produces four layers of pure populations of retinal cells: RPE (expressing NHERF1, EZRIN, RPE65, DCT, TYR, TYRP, MITF, PMEL), early photoreceptors (PRs) (coexpressing CRX and RCVRN), inner nuclear layer neurons (expressing CALB2), and retinal ganglion cells [RGCs, expressing BRN3B and Neurofilament (NF) 200]. Furthermore, we found that retinal progenitors divide at the apical side of the hESC-derived retinal tissue (next to the RPE layer) and then migrate toward the basal side, similar to that found during embryonic retinogenesis. We detected synaptogenesis in hESC-derived retinal tissue, and found neurons containing many synaptophysin-positive boutons within the RGC and PR layers. We also observed long NF200-positive axons projected by RGCs toward the apical side. Whole-cell recordings demonstrated that putative amacrine and/or ganglion cells exhibited electrophysiological responses reminiscent of those in normal retinal neurons. These responses included voltage-gated Na(+) and K(+) currents, depolarization-induced spiking, and responses to neurotransmitter receptor agonists. Differentiation in adherent conditions allows generation of long and flexible pieces of 3D retinal tissue suitable for isolating transplantable slices of tissue

  9. Acute Shear Stress Direction Dictates Adherent Cell Remodeling and Verifies Shear Profile of Spinning Disc Assays

    PubMed Central

    Fuhrmann, Alexander; Engler, Adam J.

    2015-01-01

    Several methods have been developed to quantify population level changes in cell attachment strength given its large heterogeneity. One such method is the rotating disc chamber or “spinning disc” in which a range of shear forces are applied to attached cells to quantify detachment force, i.e. attachment strength, which can be heterogeneous within cell populations. However, computing the exact force vectors that act upon cells is complicated by complex flow fields and variable cell morphologies. Recent observations suggest that cells may remodel their morphology and align during acute shear exposure, but contrary to intuition, shear is not orthogonal to the radial direction. Here we theoretically derive the magnitude and direction of applied shear and demonstrate that cells, under certain physiological conditions, align in this direction within minutes. Shear force magnitude is also experimentally verified which validates that for spread cells shear forces and not torque or drag dominate in this assay, and demonstrates that the applied force per cell area is largely independent of initial morphology. These findings suggest that direct quantified comparison of the effects of shear on a wide array of cell types and conditions can be made with confidence using this assay without the need for computational or numerical modeling. PMID:25619322

  10. Compensatory Beliefs about Glucose Testing are Associated with Low Adherence to Treatment and Poor Metabolic Control in Adolescents with Type 1 Diabetes

    ERIC Educational Resources Information Center

    Rabiau, Marjorie A.; Knauper, Barbel; Nguyen, Thien-Kim; Sufrategui, Maria; Polychronakos, Constantin

    2009-01-01

    The goal of this research was to investigate whether compensatory beliefs (CBs) regarding glucose testing predict blood glucose levels and adherence to treatment in adolescents with type 1 diabetes. CBs are convictions that the negative effects of one behavior (e.g. not testing one's glucose level) can be compensated for by engaging in another…

  11. Isolation of a protein-containing cell surface component from Streptococcus sanguis which affects its adherence to saliva-coated hydroxyapatite.

    PubMed Central

    Liljemark, W F; Bloomquist, C G

    1981-01-01

    The isolation and partial characterization of a protein-containing cell surface component from Streptococcus sanguis which blocks the adherence of this microbe to saliva-coated hydroxyapatite are described. Several methods of extraction were attempted. Sonication of whole cells and cell walls proved to be the most successful and yielded biologically active adherence-blocking components. The adherence-blocking ability of these components was effective in intraspecies blocking experiments. The extract obtained from cell walls of S. sanguis was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and shown to contain one major and two to three minor bands when stained with Coomassie blue. The molecular weight of the major band was estimated to be 70,000 to 90,000. Gel filtration of the sonified cell wall extract on 10% agarose yielded two active adherence-blocking peaks, the void volume and a second peak. Images PMID:6273317

  12. Rethinking adherence.

    PubMed

    Steiner, John F

    2012-10-16

    In 2012, the Centers for Medicare & Medicaid Services (CMS) will introduce measures of adherence to oral hypoglycemic, antihypertensive, and cholesterol-lowering drugs into its Medicare Advantage quality program. To meet these quality goals, delivery systems will need to develop and disseminate strategies to improve adherence. The design of adherence interventions has too often been guided by the mistaken assumptions that adherence is a single behavior that can be predicted from readily available patient characteristics and that individual clinicians alone can improve adherence at the population level.Effective interventions require recognition that adherence is a set of interacting behaviors influenced by individual, social, and environmental forces; adherence interventions must be broadly based, rather than targeted to specific population subgroups; and counseling with a trusted clinician needs to be complemented by outreach interventions and removal of structural and organizational barriers. To achieve the adherence goals set by CMS, front-line clinicians, interdisciplinary teams, organizational leaders, and policymakers will need to coordinate efforts in ways that exemplify the underlying principles of health care reform. PMID:23070491

  13. Hydrodynamic Determinants of Cell Necrosis and Molecular Delivery Produced by Pulsed Laser Microbeam Irradiation of Adherent Cells

    PubMed Central

    Compton, Jonathan L.; Hellman, Amy N.; Venugopalan, Vasan

    2013-01-01

    Time-resolved imaging, fluorescence microscopy, and hydrodynamic modeling were used to examine cell lysis and molecular delivery produced by picosecond and nanosecond pulsed laser microbeam irradiation in adherent cell cultures. Pulsed laser microbeam radiation at λ = 532 nm was delivered to confluent monolayers of PtK2 cells via a 40×, 0.8 NA microscope objective. Using laser microbeam pulse durations of 180–1100 ps and pulse energies of 0.5–10.5 μJ, we examined the resulting plasma formation and cavitation bubble dynamics that lead to laser-induced cell lysis, necrosis, and molecular delivery. The cavitation bubble dynamics are imaged at times of 0.5 ns to 50 μs after the pulsed laser microbeam irradiation, and fluorescence assays assess the resulting cell viability and molecular delivery of 3 kDa dextran molecules. Reductions in both the threshold laser microbeam pulse energy for plasma formation and the cavitation bubble energy are observed with decreasing pulse duration. These energy reductions provide for increased precision of laser-based cellular manipulation including cell lysis, cell necrosis, and molecular delivery. Hydrodynamic analysis reveals critical values for the shear-stress impulse generated by the cavitation bubble dynamics governs the location and spatial extent of cell necrosis and molecular delivery independent of pulse duration and pulse energy. Specifically, cellular exposure to a shear-stress impulse J≳0.1 Pa s ensures cell lysis or necrosis, whereas exposures in the range of 0.035≲J≲0.1 Pa s preserve cell viability while also enabling molecular delivery of 3 kDa dextran. Exposure to shear-stress impulses of J≲0.035 Pa s leaves the cells unaffected. Hydrodynamic analysis of these data, combined with data from studies of 6 ns microbeam irradiation, demonstrates the primacy of shear-stress impulse in determining cellular outcome resulting from pulsed laser microbeam irradiation spanning a nearly two

  14. Hydrodynamic determinants of cell necrosis and molecular delivery produced by pulsed laser microbeam irradiation of adherent cells.

    PubMed

    Compton, Jonathan L; Hellman, Amy N; Venugopalan, Vasan

    2013-11-01

    Time-resolved imaging, fluorescence microscopy, and hydrodynamic modeling were used to examine cell lysis and molecular delivery produced by picosecond and nanosecond pulsed laser microbeam irradiation in adherent cell cultures. Pulsed laser microbeam radiation at λ = 532 nm was delivered to confluent monolayers of PtK2 cells via a 40×, 0.8 NA microscope objective. Using laser microbeam pulse durations of 180-1100 ps and pulse energies of 0.5-10.5 μJ, we examined the resulting plasma formation and cavitation bubble dynamics that lead to laser-induced cell lysis, necrosis, and molecular delivery. The cavitation bubble dynamics are imaged at times of 0.5 ns to 50 μs after the pulsed laser microbeam irradiation, and fluorescence assays assess the resulting cell viability and molecular delivery of 3 kDa dextran molecules. Reductions in both the threshold laser microbeam pulse energy for plasma formation and the cavitation bubble energy are observed with decreasing pulse duration. These energy reductions provide for increased precision of laser-based cellular manipulation including cell lysis, cell necrosis, and molecular delivery. Hydrodynamic analysis reveals critical values for the shear-stress impulse generated by the cavitation bubble dynamics governs the location and spatial extent of cell necrosis and molecular delivery independent of pulse duration and pulse energy. Specifically, cellular exposure to a shear-stress impulse J≳0.1 Pa s ensures cell lysis or necrosis, whereas exposures in the range of 0.035≲J≲0.1 Pa s preserve cell viability while also enabling molecular delivery of 3 kDa dextran. Exposure to shear-stress impulses of J≲0.035 Pa s leaves the cells unaffected. Hydrodynamic analysis of these data, combined with data from studies of 6 ns microbeam irradiation, demonstrates the primacy of shear-stress impulse in determining cellular outcome resulting from pulsed laser microbeam irradiation spanning a nearly two-orders-of-magnitude range of

  15. Stationary nanoliter droplet array with a substrate of choice for single adherent/nonadherent cell incubation and analysis

    PubMed Central

    Shemesh, Jonathan; Ben Arye, Tom; Avesar, Jonathan; Kang, Joo H.; Fine, Amir; Super, Michael; Meller, Amit; Ingber, Donald E.; Levenberg, Shulamit

    2014-01-01

    Microfluidic water-in-oil droplets that serve as separate, chemically isolated compartments can be applied for single-cell analysis; however, to investigate encapsulated cells effectively over prolonged time periods, an array of droplets must remain stationary on a versatile substrate for optimal cell compatibility. We present here a platform of unique geometry and substrate versatility that generates a stationary nanodroplet array by using wells branching off a main microfluidic channel. These droplets are confined by multiple sides of a nanowell and are in direct contact with a biocompatible substrate of choice. The device is operated by a unique and reversed loading procedure that eliminates the need for fine pressure control or external tubing. Fluorocarbon oil isolates the droplets and provides soluble oxygen for the cells. By using this approach, the metabolic activity of single adherent cells was monitored continuously over time, and the concentration of viable pathogens in blood-derived samples was determined directly by measuring the number of colony-formed droplets. The method is simple to operate, requires a few microliters of reagent volume, is portable, is reusable, and allows for cell retrieval. This technology may be particularly useful for multiplexed assays for which prolonged and simultaneous visual inspection of many isolated single adherent or nonadherent cells is required. PMID:25053808

  16. Adherence to the American Diabetes Association standards of care among patients with type 2 diabetes in primary care in Saudi Arabia

    PubMed Central

    Harbi, Turki J. Al; Tourkmani, Ayla M.; Al-Khashan, Hesham I.; Mishriky, Adel M.; Qahtani, Hala Al; Bakhiet, Ahmed

    2015-01-01

    Objectives: To assess adherence to 11 American Diabetes Association (ADA) standards of diabetic care. Methods: We conducted this one-year historical prospective study between October 2010 and September 2011 on 450 adult type 2 diabetes patients in a primary care center in Saudi Arabia. We used the definitions of the 2010 ADA standards of diabetic care processes and targets. Results: Four-hundred and fifty medical files were valid. The adherence to ADA process standards of measurement of glycated hemoglobin (HbA1c) was 68.7%, 92.9% for blood pressure, and 80.2% for serum lipids. Screening was lowest for nephropathy (35.6%), and highest for diabetic foot (72%). Adherence to medications ranged between 82.2% for antiplatelets, and 92.4% for dyslipidemia. For outcome standards, 24.2% of the patients had an HbA1c <7%, and 32.2% had controlled blood pressure (<130/80 mm Hg); and 58.5% achieved targeted low-density lipoproteins (LDL). Only 7.2% had glycemic control in addition to controlled blood pressure and targeted LDL level. An increasing trend of patients achieving glycemic control (<7%) was shown throughout follow-up (p=0.003). Conclusions: We found suboptimal adherence with many ADA standards of diabetic care among patients with type 2 diabetes treated at a primary care center in Saudi Arabia. The achievement of outcome standards, either singly or combined, is lower than the adherence rates. However, the figures show improvement in adherence during the follow-up period. PMID:25719589

  17. A Randomized Controlled Trial of Cognitive Behavioral Therapy for Adherence and Depression (CBT-AD) in Patients With Uncontrolled Type 2 Diabetes

    PubMed Central

    Safren, Steven A.; Gonzalez, Jeffrey S.; Wexler, Deborah J.; Psaros, Christina; Delahanty, Linda M.; Blashill, Aaron J.; Margolina, Aleksandra I.; Cagliero, Enrico

    2014-01-01

    OBJECTIVE To test cognitive behavioral therapy for adherence and depression (CBT-AD) in type 2 diabetes. We hypothesized that CBT-AD would improve adherence; depression; and, secondarily, hemoglobin A1c (A1C). RESEARCH DESIGN AND METHODS Eighty-seven adults with unipolar depression and uncontrolled type 2 diabetes received enhanced treatment as usual (ETAU), including medication adherence, self-monitoring of blood glucose (SMBG), and lifestyle counseling; a provider letter documented psychiatric diagnoses. Those randomized to the intervention arm also received 9–11 sessions of CBT-AD. RESULTS Immediately after acute treatment (4 months), adjusting for baseline, CBT-AD had 20.7 percentage points greater oral medication adherence on electronic pill cap (95% CI −31.14 to −10.22, P = 0.000); 30.2 percentage points greater SMBG adherence through glucometer downloads (95% CI −42.95 to −17.37, P = 0.000); 6.44 points lower depression scores on the Montgomery-Asberg Depression Rating Scale (95% CI 2.33–10.56, P = 0.002); 0.74 points lower on the Clinical Global Impression (95% CI 0.16–1.32, P = 0.01); and 0.72 units lower A1C (95% CI 0.29–1.15, P = 0.001) relative to ETAU. Analyses of 4-, 8-, and 12-month follow-up time points indicated that CBT-AD maintained 24.3 percentage points higher medication adherence (95% CI −38.2 to −10.3, P = 0.001); 16.9 percentage points greater SMBG adherence (95% CI −33.3 to −0.5, P = 0.043); and 0.63 units lower A1C (95% CI 0.06–1.2, P = 0.03) after acute treatment ended. For depression, there was some evidence of continued improvement posttreatment, but no between-group differences. CONCLUSIONS CBT-AD is an effective intervention for adherence, depression, and glycemic control, with enduring and clinically meaningful benefits for diabetes self-management and glycemic control in adults with type 2 diabetes and depression. PMID:24170758

  18. Method of making a membrane having hydrophilic and hydrophobic surfaces for adhering cells or antibodies by using atomic oxygen or hydroxyl radicals

    NASA Technical Reports Server (NTRS)

    Koontz, Steven L. (Inventor); Spaulding, Glenn F. (Inventor)

    1994-01-01

    A portion of an organic polymer article such as a membrane is made hydrophilic by exposing a hydrophobic surface of the article to a depth of about 50 to about 5000 angstroms to atomic oxygen or hydroxyl radicals at a temperature below 100C., preferably below 40 C, to form a hydrophilic uniform surface layer of hydrophilic hydroxyl groups. The atomic oxygen and hydroxyl radicals are generated by a flowing afterglow microwave discharge, and the surface is outside of a plasma produced by the discharge. A membrane having both hydrophilic and hydrophobic surfaces can be used in an immunoassay by adhering antibodies to the hydrophobic surface. In another embodiment, the membrane is used in cell culturing where cells adhere to the hydrophilic surface. Prior to adhering cells, the hydrophilic surface may be grafted with a compatibilizing compound. A plurality of hydrophilic regions bounded by adjacent hydrophobic regions can be produced such that a maximum of one cell per each hydrophilic region adheres.

  19. Genome Wide assessment of Early Osseointegration in Implant-Adherent Cells

    NASA Astrophysics Data System (ADS)

    Thalji, Ghadeer N.

    Objectives: To determine the molecular processes involved in osseointegration. Materials and methods: A structured literature review concerning in vitro and in vivo molecular assessment of osseointegration was performed. A rat and a human model were then used to identify the early molecular processes involved in osseointegration associated with a micro roughened and nanosurface superimposed featured implants. In the rat model, 32 titanium implants with surface topographies exhibiting a micro roughened (AT-II) and nanosurface superimposed featured implants (AT-I) were placed in the tibiae of 8 rats and subsequently harvested at 2 and 4 days after placement. Whereas in the human model, four titanium mini-implants with either a moderately roughened surface (TiOblast) or super-imposed nanoscale topography (Osseospeed) were placed in edentulous sites of eleven systemically healthy subjects and subsequently removed after 3 and 7 days. Total RNA was isolated from cells adherent to retrieved implants. A whole genome microarray using the Affymetrix 1.1 ST Array platform was used to describe the gene expression profiles that were differentially regulated by the implant surfaces. Results: The literature review provided evidence that particular topographic cues can be specifically integrated among the many extracellular signals received by the cell in its signal transduction network. In the rat model, functionally relevant categories related to ossification, skeletal system development, osteoblast differentiation, bone development and biomineral tissue development were upregulated and more prominent at AT-I compared to AT-II. In the human model, there were no significant differences when comparing the two-implant surfaces at each time point. However, the microarray identified several genes that were differentially regulated at day 7 vs. day 3 for both implant surfaces. Functionally relevant categories related to the extracellular matrix, collagen fibril organization and

  20. Mechanism of Adherence of Streptococcus mutans to Smooth Surfaces I. Roles of Insoluble Dextran-Levan Synthetase Enzymes and Cell Wall Polysaccharide Antigen in Plaque Formation

    PubMed Central

    Mukasa, Hidehiko; Slade, Hutton D.

    1973-01-01

    The mechanism of adherence of Streptococcus mutans to smooth glass surfaces has been studied. The results with both viable and heat-killed cells showed that the process required (i) the synthesis of a water-insoluble dextran-levan polymer by cell-bound enzymes and (ii) the participation of a binding site on the surface of the S. mutans cell. Synthesis of the polymer from sucrose in the presence of the cells was required for adherence, and indicates that an “active” form of the polymer was required. Polymer synthesized by cell-free S. mutans enzymes when added to S. mutans cells did not produce adherence. Purified antibody globulin, specific for the a-d site in the polysaccharide S. mutans group a antigen, completely inhibited adherence. Antibody to the second antigen present in the polysaccharide molecule, the a antigen, did not inhibit adherence. The evidence indicates that adherence did not require an antigenic binding site which might be common to all S. mutans strains. The orientation of the synthetase enzyme(s), antigenic binding site, and dextran-levan polymer on the cell surface is under study. Images PMID:4582634

  1. Syncytial-Type Cell Plates

    PubMed Central

    Otegui, Marisa; Staehelin, L. Andrew

    2000-01-01

    Cell wall formation in the syncytial endosperm of Arabidopsis was studied by using high-pressure-frozen/freeze-substituted developing seeds and immunocytochemical techniques. The endosperm cellularization process begins at the late globular embryo stage with the synchronous organization of small clusters of oppositely oriented microtubules (∼10 microtubules in each set) into phragmoplast-like structures termed mini-phragmoplasts between both sister and nonsister nuclei. These mini-phragmoplasts produce a novel kind of cell plate, the syncytial-type cell plate, from Golgi-derived vesicles ∼63 nm in diameter, which fuse by way of hourglass-shaped intermediates into wide (∼45 nm in diameter) tubules. These wide tubules quickly become coated and surrounded by a ribosome-excluding matrix; as they grow, they branch and fuse with each other to form wide tubular networks. The mini-phragmoplasts formed between a given pair of nuclei produce aligned tubular networks that grow centrifugally until they merge into a coherent wide tubular network with the mini-phragmoplasts positioned along the network margins. The individual wide tubular networks expand laterally until they meet and eventually fuse with each other at the sites of the future cell corners. Transformation of the wide tubular networks into noncoated, thin (∼27 nm in diameter) tubular networks begins at multiple sites and coincides with the appearance of clathrin-coated budding structures. After fusion with the syncytial cell wall, the thin tubular networks are converted into fenestrated sheets and cell walls. Immunolabeling experiments show that the cell plates and cell walls of the endosperm differ from those of the embryo and maternal tissue in two features: their xyloglucans lack terminal fucose residues on the side chain, and callose persists in the cell walls after the cell plates fuse with the parental plasma membrane. The lack of terminal fucose residues on xyloglucans suggests that these cell wall

  2. Role of flagella in adherence, internalization, and translocation of Campylobacter jejuni in nonpolarized and polarized epithelial cell cultures.

    PubMed Central

    Grant, C C; Konkel, M E; Cieplak, W; Tompkins, L S

    1993-01-01

    Previous studies of Campylobacter jejuni have suggested that flagellin is an adhesin for epithelial cells and that motility is a virulence factor of this bacterium. The role of flagella in the interactions of C. jejuni with nonpolarized and polarized epithelial cells was examined with flagellar mutants. Flagellated, nonmotile (flaA flaB+ Mot-) and nonflagellated, nonmotile (flaA flaB Mot-) mutants of C. jejuni were constructed by in vivo homologous recombination and gene replacement techniques. Both classes of mutants were found to adhere to cells of human epithelial origin (INT 407) equally well; however, on the basis of the percentage of the inoculum internalized, internalization of the flaA flaB Mot- mutants was decreased by factors ranging from approximately 30 to 40 compared with the parent. The flaA flaB+ Mot- mutant was internalized by the INT 407 cells at levels six- to sevenfold higher than the flaA flaB Mot- mutants. Both classes of mutants, unlike the parent, were unable to translocate across polarized Caco-2 monolayers. These results indicate that flagella are not involved in C. jejuni adherence to epithelial cells but that they do play a role in internalization. Furthermore, the results suggest that either the motility of C. jejuni or the product of flaA is essential for the bacterium to cross polarized epithelial cell monolayers. Images PMID:8478066

  3. Display of Cell Surface Sites for Fibronectin Assembly Is Modulated by Cell Adherence to 1F3 and C-Terminal Modules of Fibronectin

    PubMed Central

    Zhang, Qinghong; Annis, Douglas S.; Erickson, Harold P.; Mosher, Deane F.

    2009-01-01

    Background Fibronectin-null cells assemble soluble fibronectin shortly after adherence to a substrate coated with intact fibronectin but not when adherent to the cell-binding domain of fibronectin (modules 7F3-10F3). Interactions of adherent cells with regions of adsorbed fibronectin other than modules 7F3-10F3, therefore, are required for early display of the cell surface sites that initiate and direct fibronectin assembly. Methodology/Principal Findings To identify these regions, coatings of proteolytically derived or recombinant pieces of fibronectin containing modules in addition to 7F3-10F3 were tested for effects on fibronectin assembly by adherent fibronectin-null fibroblasts. Pieces as large as one comprising modules 2F3-14F3, which include the heparin-binding and cell adhesion domains, were not effective in supporting fibronectin assembly. Addition of module 1F3 or the C-terminal modules to modules 2F3-14F3 resulted in some activity, and addition of both 1F3 and the C-terminal modules resulted in a construct, 1F3-C, that best mimicked the activity of a coating of intact fibronectin. Constructs 1F3-C V0, 1F3-C V64, and 1F3-C Δ(V15F310F1) were all able to support fibronectin assembly, suggesting that 1F3 through 11F1 and/or 12F1 were important for activity. Coatings in which the active parts of 1F3-C were present in different proteins were much less active than intact 1F3-C. Conclusions These results suggest that 1F3 acts together with C-terminal modules to induce display of fibronectin assembly sites on adherent cells. PMID:19119318

  4. Vascular cell adhesion molecule-1 and the integrin VLA-4 mediate adhesion of human B cell precursors to cultured bone marrow adherent cells.

    PubMed Central

    Ryan, D H; Nuccie, B L; Abboud, C N; Winslow, J M

    1991-01-01

    Adhesion of B cell precursors to accessory cells in the bone marrow microenvironment may be required for normal early B cell development. Human bone marrow B cell precursors adhere more avidly than mature B cells to bone marrow-derived fibroblasts. To determine the mechanism of this adhesion, expression of adhesion proteins on human B precursor cells and cell lines was measured by flow cytometry. The very late antigen (VLA) integrins VLA-4 and VLA-5 were the only adhesion proteins expressed at higher levels in B cell precursors than mature B cells. Antibodies to the alpha and beta chains of VLA-4, but not VLA-5, significantly blocked binding to bone marrow-derived fibroblasts of immature B cells and cell lines. Although fibronectin is a ligand for VLA-4, anti-fibronectin antibody and a soluble fibronectin fragment containing the VLA-4 binding domain did not block adhesion, suggesting that VLA-4 is involved in adhesion of B cell precursors, but not as a fibronectin receptor. Vascular cell adhesion molecule-1 (VCAM-1), the other known counterreceptor for VLA-4, was identified on bone marrow-derived fibroblasts, and anti-VCAM-1 significantly blocked adhesion of normal B cell precursors to bone marrow-derived fibroblasts, indicating that VLA-4/VCAM-1 interactions are important in adhesion of B cell precursors to the bone marrow microenvironment. Images PMID:1715889

  5. Invitro study of adherent mandibular osteoblast-like cells on carrier materials.

    PubMed

    Turhani, D; Weissenböck, M; Watzinger, E; Yerit, K; Cvikl, B; Ewers, R; Thurnher, D

    2005-07-01

    Augmentation of the craniofacial region is necessary for many aesthetic and reconstructive procedures. Tissue engineering offers a new option to supplement existing treatment regimens. In this procedure, materials composed of hydroxyapatite (HA), of synthetic or natural origin, are used as scaffolds. The aim of this study was to evaluate the effects of three HA materials on cultured human osteoblasts in vitro. Explant cultures of cells from human alveolar bone were established. Human osteoblasts were cultured on the surface of HA calcified from red algae (C GRAFT/Algipore), deproteinized bovine HA (Bio-Oss) and bovine HA carrying the cell binding peptide P-15 (Pep Gen P-15). Cultured cells were evaluated with respect to cell attachment, proliferation and differentiation. Cells were cultured for 6 and 21 days under osteogenic differentiation conditions, and tissue-culture polystyrene dishes were used as control. The ability of cells to proliferate and form extracellular matrix on these scaffolds was assessed by a DNA quantification assay, protein synthesis analysis and by scanning electron microscopical examination. Osteogenic differentiation was screened by the expression of alkaline phosphatase. The osteoblastic phenotype of the cells was monitored using mRNA levels of the bone-related proteins including osteocalcin, osteopontin and collagen Type I. We found that cells cultured on C GRAFT/Algipore) and Pep Gen P-15 showed a continuous increase in DNA content and protein synthesis. Cells cultured on Bio-Oss showed a decrease in DNA content from Day 6 (P < 0.05) to Day 21 (P < 0.0001) and protein synthesis on Day 21 (P < 0.005). Alkaline phosphatase activity increased in cells grown on C GRAFT/Algipore and Pep Gen P-15 in contrast to cells grown on Bio-Oss, in which the lowest levels of activity could be observed on Day 21 (P < 0.05). Reverse transcriptase polymerase chain reaction analysis confirmed the osteoblastic phenotype of the cells grown on all three

  6. Effect of shear stress and of transmural pressure on cAMP-dependent responses of cells adhering to a biomaterial

    NASA Astrophysics Data System (ADS)

    Chotard-Ghodsnia, R.; Drochon, A.; Faucheux, N.; Nagel, M.-D.; Grebe, R.

    2002-02-01

    Biomaterials used in some bioreactors are porous and exposed to normal and tangential flow of physiological fluid. Flow-induced forces may influence the morphological and biochemical responses of cells adhering to these materials. The objective of this work is to examine the capacity of mechanical stress to cause changes in cell morphology via the cAMP pathway (cyclic adenosine monophosphate). This second messenger is known to modulate cell morphology in static conditions. In classical flow devices, cells are submitted to only tangential stresses. We designed a new flow system, a Hele-Shaw cell with a porous bottom wall, in order to take into account the influence of a transmural pressure. This flow chamber allows to follow up continuously the shape changes of cells that are adherent to a porous biomaterial (polyacrylonitrile) and are exposed to controlled levels of shear stress or transmural pressure. Mouse Swiss 3T3 fibroblasts exposed to a 1.1-Pa shear stress, as well as those exposed to a 84-mm Hg transmural pressure, round up (up to 50%) in a few minutes. If the cAMP pathway is inhibited when a mechanical stress is applied, cell rounding is significantly prevented. These observations suggest that flow-induced cell shape changes are cAMP-dependent. This conclusion is supported by an increased cAMP accumulation measured in cells under mechanical stress when compared to static experiments. Our in vitro flow system is thus useful to study the influence of transmural pressure or shear stress on the early morphological and biochemical responses of cells in contact with a biomaterial.

  7. Adherent invasive Escherichia coli strains from patients with Crohn's disease survive and replicate within macrophages without inducing host cell death.

    PubMed

    Glasser, A L; Boudeau, J; Barnich, N; Perruchot, M H; Colombel, J F; Darfeuille-Michaud, A

    2001-09-01

    Escherichia coli strains recovered from Crohn's disease (CD) lesions are able to adhere to and invade cultured intestinal epithelial cells. We analyzed the behavior within macrophages of adherent invasive E. coli (AIEC) strains isolated from patients with CD. All the 15 AIEC strains tested were able to replicate extensively within J774-A1 cells: the numbers of intracellular bacteria increased 2.2- to 74.2-fold at 48 h over that at 1 h postinfection. By use of murine peritoneal macrophages and human monocyte-derived-macrophages, the reference AIEC strain LF82 was confirmed to be able to survive intracellularly. Transmission electron micrographs of AIEC LF82-infected macrophages showed that at 24 h postinfection, infected cells harbored large vacuoles containing numerous bacteria, as a result of the fusion of several vacuoles occurring after 8 h postinfection. No lactate dehydrogenase (LDH) release, no sign of DNA fragmentation or degradation, and no binding to fluorescein isothlocyanate-labeled annexin V were observed with LF82-infected J774-A1 cells, even after 24 h postinfection. LF82-infected J774-A1 cells secreted 2.7-fold more tumor necrosis factor alpha (TNF-alpha) than cells stimulated with 1 microg of lipopolysaccharide (LPS)/ml. No release of interleukin-1beta was observed with LPS-prestimulated J774-A1 cells infected with AIEC LF82. These findings showed that (i) AIEC strains are able to survive and to replicate within macrophages, (ii) AIEC LF82 replication does not induce any cell death of the infected cells, and (iii) LF82-infected J774-A1 cells release high levels of TNF-alpha. These properties could be related to some features of CD and particularly to granuloma formation, one of the hallmarks of CD lesions. PMID:11500426

  8. Large-Scale Production of High-Quality Helper-Dependent Adenoviral Vectors Using Adherent Cells in Cell Factories

    PubMed Central

    Suzuki, Masataka; Cela, Racel; Clarke, Christian; Bertin, Terry K.; Mouriño, Susana

    2010-01-01

    Abstract The most efficient and widely used system for generating helper-dependent adenoviral vectors (HDAds) is the Cre/loxP system developed by Graham and co-workers (Parks, R.J., Chen, L., Anton, M., Sankar, U., Rudnicki, M.A., and Graham, F.L. [1996]. Proc. Natl. Acad. Sci. U. S. A. 93, 13565–13570). Alternative systems have been developed for HDAd production, but all are limited by the technical complexity of a three-component vector production system for reproducibly generating large quantities of adenovirus with high infectivity and low helper virus (HV) contamination. Recently, these problems were addressed by Ng and co-workers (Palmer, D., and Ng, P. [2003]. Mol Ther. 8, 846–852), who developed an improved system that combines the use of a suspension-adapted producer cell line expressing high levels of Cre recombinase, a HV resistant to mutation, and a refined purification protocol. With this system, >1 × 1013 highly infectious vector particles are easily produced without vector genome rearrangements and having very low HV contamination levels. However, the Ng system incorporates a spinner flask culture system that involves considerable time, effort, and tissue culture medium to produce HDAds. We have an alternative system to obtain comparable quantities with equivalent quality to the spinner flask approach but requiring reduced labor and lower volumes of medium. This method utilizes a 10-chamber cell factory with adherent cells to produce high infectivity of HDAds with minimal HV contamination while improving yield and reducing technical complexity, effort, and medium requirements. This system is easily translatable to the production of clinical-grade HDAds for human trials. PMID:19719388

  9. Effects of cryopreservation and phenylacetate on biological characters of adherent LAK cells from patients with hepatocellular carcinoma

    PubMed Central

    Zheng, Ning; Ye, Sheng-Long; Sun, Rui-Xia; Zhao, Yan; Tang, Zhao-You

    2002-01-01

    AIM: To improve the preparation of adherent lymphokine-activated killer (A-LAK) cells and to study the effects of cryopreservation and phenylacetate (PA) on biological characters of A-LAK cells. METHODS: A-LAK cells were obtained from peripheral blood mononuclear cells (PBMCs) of the patients with hepatocellular carcinoma (HCC) by using L-phenylalanine methyl ester (PME) to deplete immunosuppressive monocytes. Proliferative activity of SMMC7721 cell line after treatment with phenylacetate (PA) was observed. A-LAK cells were treated with the supernatant of SMMC7721 cells that had been pretreated with PA. The changes of proliferation, cytotoxicity and phenotype of A-LAK cells were investigated after cryopreservation. RESULTS: The expansion of A-LAK cells (96.79 ± 69.10 folds on Day 14) was significantly higher than that of non-adherent LAK (NA-LAK) cells (22.77 ± 13.20) as well as conventional LAK cells (4.64 ± 0.91). PA significantly suppressed the growth of SMMC7721 cells, and the inhibitor ratio was 46%. The supernatant of cultured tumor cells intensively suppressed the proliferation and cytotoxicity of A-LAK cells, but the suppressive effect of the supernatant was previously decreased after treatment with PA. Impairments in proliferation and cytotoxicity of A-LAK cells immediately after thawing of cryopreservation and recovery after reincubation with IL-2 were observed. The cytotoxicity of thawed A-LAK cells on Day 5 was significantly higher than that of fresh A-LAK before freezing (54.8% ± 10.2% vs 40.5% ± 6.4%). No significant change in the percentage of lymphocyte subsets was identified in frozen A-LAK cells as compared with that in the fresh control cells. CONCLUSION: A-LAK cells can be simply prepared by using PME, and showed a synergistic anti-tumor effect with the combination of PA. Cryopreservation can increase the immunoactivities of A-LAK cells from the patients with hepatocellular carcinoma. PMID:11925598

  10. Capsule of Streptococcus equi subsp. zooepidemicus hampers the adherence and invasion of epithelial and endothelial cells and is attenuated during internalization.

    PubMed

    Xu, Bin; Pei, Xiaomeng; Su, Yiqi; Ma, Zhe; Fan, Hongjie

    2016-08-01

    Direct interaction between pathogens and host cells often is a prerequisite for colonization, infection and dissemination. Regulated production of capsular polysaccharide (CPS), which is made of hyaluronic acid, is essential for the pathogenicity of Streptococcus equi subsp. Zooepidemicus (SEZ). Here, we constructed a CPS-deleted mutant and analyzed it along with the parental wild-type strain in attachment and invasion of mammalian epithelial and endothelial cell lines. The CPS-deleted mutant exhibited significant increase in adherence and invasion by several orders of magnitude compared with the wild-type strain through quantitative analysis and electron microscopy observation. After the wild-type strain was recovered from invaded cells, its morphology was analyzed by visual methods and scanning electron microscopy, which revealed that its capsule was almost completely absent. Capsule measurements showed a similar result in which CPS production was nearly attenuated to the same extent as in the CPS-deleted mutant. qPCR assays revealed a marked reduction in the transcriptional levels of the CPS biosynthesis genes, has operon. Moreover, the repression in capsular production was stable inheritance. Our findings indicate that SEZ is a facultative intracellular bacterium, capsule attenuation in SEZ contributes to attachment and invasion in interactions with host cells, and the active regulation of capsule breakdown is controlled by SEZ during internalization. PMID:27388015

  11. Empowerment, motivation, and medical adherence (EMMA): the feasibility of a program for patient-centered consultations to support medication adherence and blood glucose control in adults with type 2 diabetes

    PubMed Central

    Varming, Annemarie Reinhardt; Hansen, Ulla Møller; Andrésdóttir, Gudbjörg; Husted, Gitte Reventlov; Willaing, Ingrid

    2015-01-01

    Purpose To explore the feasibility of a research-based program for patient-centered consultations to improve medical adherence and blood glucose control in patients with type 2 diabetes. Patients and methods The patient-centered empowerment, motivation, and medical adherence (EMMA) consultation program consisted of three individual consultations and one phone call with a single health care professional (HCP). Nineteen patients with type 2 diabetes completed the feasibility study. Feasibility was assessed by a questionnaire-based interview with patients 2 months after the final consultation and interviews with HCPs. Patient participation was measured by 10-second event coding based on digital recordings and observations of the consultations. Results HCPs reported that EMMA supported patient-centered consultations by facilitating dialogue, reflection, and patient activity. Patients reported that they experienced valuable learning during the consultations, felt understood, and listened to and felt a trusting relationship with HCPs. Consultations became more person-specific, which helped patients and HCPs to discover inadequate diabetes self-management through shared decision-making. Compared with routine consultations, HCPs talked less and patients talked more. Seven of ten dialogue tools were used by all patients. It was difficult to complete the EMMA consultations within the scheduled time. Conclusion The EMMA program was feasible, usable, and acceptable to patients and HCPs. The use of tools elicited patients’ perspectives and facilitated patient participation and shared decision-making. PMID:26366060

  12. A Crosstalk Between K ras (Kirsten Rat Sarcoma Viral Oncogene Homologue) and Adherence Molecular Complex Leads to Disassociation of Cells-A Possible Contribution Towards Metastasis in Colorectal Cancer.

    PubMed

    Murtaza, Bibi Nazia; Doak, Shareen; Morgan, Claire; Nadeem, Muhammad Shahid; Al-Ghanim, Khalid A; Shakoori, Abdul Rauf

    2016-10-01

    Constitutive activation of mutant K ras (Kirsten rat sarcoma viral oncogene homologue) and disassembly of E-cadherin-catenin complex (E-cadherin, α-catenin, β-catenin, and γ-catenin) play an important role in apoptosis, differentiation, and cell proliferation. In this study, the expression pattern of K ras and E-cadherin-catenin complex has been evaluated in normal and mutant colorectal cancer cell lines with an object to determine its impact on disassociation of cells from one another. We addressed the expression analysis of K ras with reference to its association with adherence molecules in two colorectal cancer cell lines, that is, Caco-2 (wild type K ras served as a control) and DLD1 (heterozygous mutation at codon 13) at message level by qRT-PCR and translational level by western blotting. Compared to the control Caco-2 cell lines, the K ras in DLD1 cell lines showed slightly higher values while α-catenin showed a slight lower (1.3-folds), β-catenin and E-cadherin showed significantly lower expression (4.2-fold decrease). It can be inferred that a possible cross talk exists between K ras and adherent junction mediated signalling. Mutation at codon 13 (G to D) leads to the overexpression of K ras and reduced expression of adherent junction complex resulting in metastasis. J. Cell. Biochem. 117: 2340-2345, 2016. © 2016 Wiley Periodicals, Inc. PMID:26945839

  13. Krüppel-Like Factor 4 Overexpression Initiates a Mesenchymal-to-Epithelial Transition and Redifferentiation of Human Pancreatic Cells following Expansion in Long Term Adherent Culture

    PubMed Central

    Docherty, Hilary M.; McGowan, Neil W. A.; Forbes, Shareen; Heremans, Yves; Forbes, Stuart J.; Heimberg, Harry; Casey, John; Docherty, Kevin

    2015-01-01

    A replenishable source of insulin-producing cells has the potential to cure type 1 diabetes. Attempts to culture and expand pancreatic β-cells in vitro have resulted in their transition from insulin-producing epithelial cells to mesenchymal stromal cells (MSCs) with high proliferative capacity but devoid of any hormone production. The aim of this study was to determine whether the transcription factor Krüppel-like factor 4 (KLF4), could induce a mesenchymal-to-epithelial transition (MET) of the cultured cells. Islet-enriched pancreatic cells, allowed to dedifferentiate and expand in adherent cell culture, were transduced with an adenovirus containing KLF4 (Ad-Klf4). Cells were subsequently analysed for changes in cell morphology by light microscopy, and for the presence of epithelial and pancreatic markers by immunocytochemistry and quantitative RT/PCR. Infection with Ad-Klf4 resulted in morphological changes, down-regulation of mesenchymal markers, and re-expression of both epithelial and pancreatic cell markers including insulin and transcription factors specific to β-cells. This effect was further enhanced by culturing cells in suspension. However, the effects of Ad-KLf4 were transient and this was shown to be due to increased apoptosis in Klf4-expressing cells. Klf4 has been recently identified as a pioneer factor with the ability to modulate the structure of chromatin and enhance reprogramming/transdifferentiation. Our results show that Klf4 may have a role in the redifferentiation of expanded pancreatic cells in culture, but before this can be achieved the off-target effects that result in increased apoptosis would need to be overcome. PMID:26457418

  14. Impact of release dynamics of laser-irradiated polymer micropallets on the viability of selected adherent cells

    PubMed Central

    Ma, Huan; Mismar, Wael; Wang, Yuli; Small, Donald W.; Ras, Mat; Allbritton, Nancy L.; Sims, Christopher E.; Venugopalan, Vasan

    2012-01-01

    We use time-resolved interferometry, fluorescence assays and computational fluid dynamics (CFD) simulations to examine the viability of confluent adherent cell monolayers to selection via laser microbeam release of photoresist polymer micropallets. We demonstrate the importance of laser microbeam pulse energy and focal volume position relative to the glass–pallet interface in governing the threshold energies for pallet release as well as the pallet release dynamics. Measurements using time-resolved interferometry show that increases in laser pulse energy result in increasing pallet release velocities that can approach 10 m s−1 through aqueous media. CFD simulations reveal that the pallet motion results in cellular exposure to transient hydrodynamic shear stress amplitudes that can exceed 100 kPa on microsecond timescales, and which produces reduced cell viability. Moreover, CFD simulation results show that the maximum shear stress on the pallet surface varies spatially, with the largest shear stresses occurring on the pallet periphery. Cell viability of confluent cell monolayers on the pallet surface confirms that the use of larger pulse energies results in increased rates of necrosis for those cells situated away from the pallet centre, while cells situated at the pallet centre remain viable. Nevertheless, experiments that examine the viability of these cell monolayers following pallet release show that proper choices for laser microbeam pulse energy and focal volume position lead to the routine achievement of cell viability in excess of 90 per cent. These laser microbeam parameters result in maximum pallet release velocities below 6 m s−1 and cellular exposure of transient hydrodynamic shear stresses below 20 kPa. Collectively, these results provide a mechanistic understanding that relates pallet release dynamics and associated transient shear stresses with subsequent cellular viability. This provides a quantitative, mechanistic basis for determining

  15. Lateral flagella are required for increased cell adherence, invasion and biofilm formation by Aeromonas spp.

    PubMed

    Gavín, Rosalina; Merino, Susana; Altarriba, Maria; Canals, Rocío; Shaw, Jonathan G; Tomás, Juan M

    2003-07-15

    Two types of flagella are responsible for motility in mesophilic Aeromonas strains. A polar unsheathed flagellum is expressed constitutively that allows the bacterium to swim in liquid environments and, in media where the polar flagellum is unable to propel the cell, Aeromonas express peritrichous lateral flagella. Recently, Southern blot analysis using a DNA probe based on the Aeromonas caviae Sch3N lateral flagellin gene sequence showed a good correlation between strains positive for the DNA probe, swarming motility and the presence of lateral flagella by microscopy. Here, we conclude that the easiest method for the detection of the lateral flagellin gene(s) is by PCR (polymerase chain reaction); this showed good correlation with swarming motility and the presence of lateral flagella. This was despite the high degree of DNA heterogeneity found in Aeromonas gene sequences. Furthermore, by reintroducing the laf (lateral flagella) genes into several mesophilic lateral-flagella-negative Aeromonas wild-type strains, we demonstrate that this surface structure enhances the adhesion to and invasion of HEp-2 cells and the capacity for biofilm formation in vitro. These results, together with previous data obtained using Laf- mutants, demonstrate that lateral flagella production is a pathogenic feature due to its enhancement of the interaction with eukaryotic cell surfaces. PMID:12855171

  16. Chemical stimulation of adherent cells by localized application of acetylcholine from a microfluidic system.

    PubMed

    Zibek, Susanne; Hagmeyer, Britta; Stett, Alfred; Stelzle, Martin

    2010-01-01

    Chemical stimulation of cells is inherently cell type selective in contrast to electro-stimulation. The availability of a system for localized application of minute amounts of chemical stimulants could be useful for dose related response studies to test new compounds. It could also bring forward the development of a novel type of neuroprostheses. In an experimental setup microdroplets of an acetylcholine solution were ejected from a fluidic microsystem and applied to the bottom of a nanoporous membrane. The solution traveled through the pores to the top of the membrane on which TE671 cells were cultivated. Calcium imaging was used to visualize cellular response with temporal and spatial resolution. Experimental demonstration of chemical stimulation for both threshold gated stimulation as well as accumulated dose-response was achieved by either employing acetylcholine as chemical stimulant or applying calcein uptake, respectively. Numerical modeling and simulation of transport mechanisms involved were employed to gain a theoretical understanding of the influence of pore size, concentration of stimulant and droplet volume on the spatial-temporal distribution of stimulant and on the cellular response. Diffusion, pressure driven flow and evaporation effects were taken into account. Fast stimulation kinetic is achieved with pores of 0.82 μm diameter, whereas sustained substance delivery is obtained with nanoporous membranes. In all cases threshold concentrations ranging from 0.01 to 0.015 μM acetylcholine independent of pore size were determined. PMID:21151808

  17. Chemical Stimulation of Adherent Cells by Localized Application of Acetylcholine from a Microfluidic System

    PubMed Central

    Zibek, Susanne; Hagmeyer, Britta; Stett, Alfred; Stelzle, Martin

    2010-01-01

    Chemical stimulation of cells is inherently cell type selective in contrast to electro-stimulation. The availability of a system for localized application of minute amounts of chemical stimulants could be useful for dose related response studies to test new compounds. It could also bring forward the development of a novel type of neuroprostheses. In an experimental setup microdroplets of an acetylcholine solution were ejected from a fluidic microsystem and applied to the bottom of a nanoporous membrane. The solution traveled through the pores to the top of the membrane on which TE671 cells were cultivated. Calcium imaging was used to visualize cellular response with temporal and spatial resolution. Experimental demonstration of chemical stimulation for both threshold gated stimulation as well as accumulated dose–response was achieved by either employing acetylcholine as chemical stimulant or applying calcein uptake, respectively. Numerical modeling and simulation of transport mechanisms involved were employed to gain a theoretical understanding of the influence of pore size, concentration of stimulant and droplet volume on the spatial-temporal distribution of stimulant and on the cellular response. Diffusion, pressure driven flow and evaporation effects were taken into account. Fast stimulation kinetic is achieved with pores of 0.82 μm diameter, whereas sustained substance delivery is obtained with nanoporous membranes. In all cases threshold concentrations ranging from 0.01 to 0.015 μM acetylcholine independent of pore size were determined. PMID:21151808

  18. EnP1, a Microsporidian Spore Wall Protein That Enables Spores To Adhere to and Infect Host Cells In Vitro▿

    PubMed Central

    Southern, Timothy R.; Jolly, Carrie E.; Lester, Melissa E.; Hayman, J. Russell

    2007-01-01

    Microsporidia are spore-forming fungal pathogens that require the intracellular environment of host cells for propagation. We have shown that spores of the genus Encephalitozoon adhere to host cell surface glycosaminoglycans (GAGs) in vitro and that this adherence serves to modulate the infection process. In this study, a spore wall protein (EnP1; Encephalitozoon cuniculi ECU01_0820) from E. cuniculi and Encephalitozoon intestinalis is found to interact with the host cell surface. Analysis of the amino acid sequence reveals multiple heparin-binding motifs, which are known to interact with extracellular matrices. Both recombinant EnP1 protein and purified EnP1 antibody inhibit spore adherence, resulting in decreased host cell infection. Furthermore, when the N-terminal heparin-binding motif is deleted by site-directed mutagenesis, inhibition of adherence is ablated. Our transmission immunoelectron microscopy reveals that EnP1 is embedded in the microsporidial endospore and exospore and is found in high abundance in the polar sac/anchoring disk region, an area from which the everting polar tube is released. Finally, by using a host cell binding assay, EnP1 is shown to bind host cell surfaces but not to those that lack surface GAGs. Collectively, these data show that given its expression in both the endospore and the exospore, EnP1 is a microsporidian cell wall protein that may function both in a structural capacity and in modulating in vitro host cell adherence and infection. PMID:17557882

  19. B-lymphocyte responses to trinitrophenyl-conjugated Ficoll: requirement for T lymphocytes and Ia-bearing adherent cells.

    PubMed Central

    Letvin, N L; Benacerraf, B; Germain, R N

    1981-01-01

    These studies were done to characterize the cellular requirements for B-lymphocyte responses to the haptenated polysaccharide trinitrophenyl-conjugated Ficoll. By using an in vitro microculture system, it was demonstrated that hapten-specific anti-trinitrophenyl-conjugated Ficoll plaque-forming cell responses by B lymphocytes require Ia-bearing adherent accessory cells and Thy 1+ Lyt 1+2- nylon wool-nonadherent (T) lymphocytes. Such T cells could be primed in vivo with nonderivatized Ficoll to show carrier-specific helper cell function for derivatized Ficoll responses in vitro. The implications of these findings for our understanding of b-lymphocyte triggering by so-called T-independent antigens are discussed. PMID:6975477

  20. Decreased Adherence of Enterohemorrhagic Escherichia coli to HEp-2 Cells in the Presence of Antibodies That Recognize the C-Terminal Region of Intimin

    PubMed Central

    Gansheroff, Lisa J.; Wachtel, Marian R.; O'Brien, Alison D.

    1999-01-01

    Antiserum raised against intimin from enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain 86-24 has been shown previously by our laboratory to inhibit adherence of this strain to HEp-2 cells. In the present study, we sought to identify the region(s) of intimin important for the effect of anti-intimin antisera on EHEC adherence and to determine whether antisera raised against intimin from an O157:H7 strain could reduce adherence of other strains. Compared to preimmune serum controls, polyclonal sera raised against the histidine-tagged intimin protein RIHisEae (intiminO157) or against His-tagged C-terminal fragments of intimin from strain 86-24 reduced adherence of this strain. Furthermore, an antibody fraction purified from the anti-RIHisEae serum that contained antibodies to the C-terminal third of intimin, the putative receptor-binding domain, also reduced adherence of strain 86-24, but a purified fraction containing antibodies to the N-terminal two-thirds of intimin did not inhibit adherence. The polyclonal anti-intiminO157 serum raised against RIHisEae inhibited, to different degrees, the adherence of another O157:H7 strain, an EHEC O55:H7 strain, one of two independent EHEC O111:NM isolates tested, and one of two EHEC O26:H11 strains tested. Adherence of the other O26:H11 and O111:NM strains and an EPEC O127:H6 strain was not reduced. Finally, immunoblot analysis indicated a correlation between the antigenic divergence in the C-terminal third of intimins from different strains and the capacity of anti-intiminO157 antiserum to reduce adherence of heterologous strains. Taken together, these data suggest that intiminO157 could be used as an immunogen to elicit adherence-blocking antibodies against O157:H7 strains and closely-related EHEC. PMID:10569757

  1. Acute myelogenous leukemia cells with the MLL-ELL translocation convert morphologically and functionally into adherent myofibroblasts

    SciTech Connect

    Tashiro, Haruko; Mizutani-Noguchi, Mitsuho; Shirasaki, Ryosuke

    2010-01-01

    Bone marrow-myofibroblasts, a major component of bone marrow-stroma, are reported to originate from hematopoietic stem cells. We show in this paper that non-adherent leukemia blasts can change into myofibroblasts. When myeloblasts from two cases of acute myelogenous leukemia with a fusion product comprising mixed lineage leukemia and RNA polymerase II elongation factor, were cultured long term, their morphology changed to that of myofibroblasts with similar molecular characteristics to the parental myeloblasts. The original leukemia blasts, when cultured on the leukemia blast-derived myofibroblasts, grew extensively. Leukemia blasts can create their own microenvironment for proliferation.

  2. Specific adherence of Borrelia burgdorferi extracellular vesicles to human endothelial cells in culture.

    PubMed Central

    Shoberg, R J; Thomas, D D

    1993-01-01

    Borrelia burgdorferi produces extracellular vesicles which contain some of the outer surface proteins of the bacterium (e.g., OspA and OspB). Borrelial vesicles, isolated by differential centrifugation and filtration, were tested for the ability to bind to cultured human umbilical vein endothelial (HUVE) cells in culture. The recently described lipoprotein OspD was expressed on vesicles. Vesicles exhibited differential expression of OspB and OspD in a relationship with passage number and medium serum supplement type, respectively. Qualitative immunoblotting analyses demonstrated dose-dependent, passage number-dependent adsorption of vesicles by HUVE cells. This adsorption was demonstrated to be dependent upon a borrelial component of the vesicle and not due to the presence of minor contamination with intact spirochetes. Quantitative experiments examining inhibition of B. burgdorferi-HUVE association as a function of prior vesicle-HUVE association demonstrated dependence upon (i) a borrelial component(s) in the vesicle, (ii) low passage number, and (iii) vesicle protein concentration. However, vesicle pretreatment of the HUVE cell monolayer was not requisite for this inhibition. Vesicles from highly passaged borrelias were noninhibitory for B. burgdorferi-HUVE cell association, regardless of the serum used to supplement the medium. The use of vesicles as a tool for studying B. burgdorferi pathogenesis and/or physiology is proposed. Images PMID:8359911

  3. Cellular Adherence, Glucosyltransferase Adsorption, and Glucan Synthesis of Streptococcus mutans AHT Mutants

    PubMed Central

    Koga, Toshihiko; Inoue, Masakazu

    1978-01-01

    Streptococcus mutans AHT mutants M1, M2, and M13 failed to adhere to a glass surface, whereas mutants M9 and M35 exhibited decreased and increased adherence, respectively, as compared with the parent strain, when grown in sucrose broth. Extracellular glucosyltransferase prepared from glucose-grown cultures of the adherent strains (wild type, M9, and M35) induced adherence of heat-killed cells of the homologous and heterologous streptococcal strains as well as of Escherichia coli K-12 and uncoated resin particles. The glucosyltransferase was adsorbed on all the streptococcal cells and glucan-coated resins, but not on E. coli cells and the uncoated resins. Glucosyltransferase from the nonadhering mutants (M1, M2, M13) neither was significantly adsorbed on nor induced adherence of any of the cells and resins. Cell-free enzymes from the glucose-grown adherent strains produced water-soluble and water-insoluble glucans, whereas those from the nonadhering mutants produced only water-soluble glucans. Small amounts of alkali-soluble, cell-associated glucan were recovered from the sucrose-grown nonadhering mutants. Thus, the relative proportions of glucosyltransferase isozymes elaborated by the S. mutans mutants, insofar as they affect the physico-chemical properties of the glucans produced, seem to determine the adherence abilities of the cells. The adsorption of glucosyltransferase on glucan molecules on the cell surface is not required for the adherence of S. mutans, but de novo glucan synthesis is important in the adherence process. PMID:631879

  4. Inhibition of Streptococcus pneumoniae adherence to human epithelial cells in vitro by the probiotic Lactobacillus rhamnosus GG

    PubMed Central

    2013-01-01

    Background Colonization of the nasopharynx by Streptococcus pneumoniae is considered a prerequisite for pneumococcal infections such as pneumonia and otitis media. Probiotic bacteria can influence disease outcomes through various mechanisms, including inhibition of pathogen colonization. Here, we examine the effect of the probiotic Lactobacillus rhamnosus GG (LGG) on S. pneumoniae colonization of human epithelial cells using an in vitro model. We investigated the effects of LGG administered before, at the same time as, or after the addition of S. pneumoniae on the adherence of four pneumococcal isolates. Results LGG significantly inhibited the adherence of all the pneumococcal isolates tested. The magnitude of inhibition varied with LGG dose, time of administration, and the pneumococcal isolate used. Inhibition was most effective when a higher dose of LGG was administered prior to establishment of pneumococcal colonization. Mechanistic studies showed that LGG binds to epithelial cells but does not affect pneumococcal growth or viability. Administration of LGG did not lead to any significant changes in host cytokine responses. Conclusions These findings demonstrate that LGG can inhibit pneumococcal colonization of human epithelial cells in vitro and suggest that probiotics could be used clinically to prevent the establishment of pneumococcal carriage. PMID:23561014

  5. Adherence to a Healthy Nordic Food Index Is Associated with a Lower Risk of Type-2 Diabetes—The Danish Diet, Cancer and Health Cohort Study

    PubMed Central

    Lacoppidan, Sandra Amalie; Kyrø, Cecilie; Loft, Steffen; Helnæs, Anne; Christensen, Jane; Hansen, Camilla Plambeck; Dahm, Christina Catherine; Overvad, Kim; Tjønneland, Anne; Olsen, Anja

    2015-01-01

    Background: Type-2 diabetes (T2D) prevalence is rapidly increasing worldwide. Lifestyle factors, in particular obesity, diet, and physical activity play a significant role in the etiology of the disease. Of dietary patterns, particularly the Mediterranean diet has been studied, and generally a protective association has been identified. However, other regional diets are less explored. Objective: The aim of the present study was to investigate the association between adherence to a healthy Nordic food index and the risk of T2D. The index consists of six food items: fish, cabbage, rye bread, oatmeal, apples and pears, and root vegetables. Methods: Data was obtained from a prospective cohort study of 57,053 Danish men and women aged 50–64 years, at baseline, of whom 7366 developed T2D (median follow-up: 15.3 years). The Cox proportional hazards model was used to assess the association between the healthy Nordic food index and risk of T2D, adjusted for potential confounders. Results: Greater adherence to the healthy Nordic food index was significantly associated with lower risk of T2D after adjusting for potential confounders. An index score of 5−6 points (high adherence) was associated with a statistically significantly 25% lower T2D risk in women (HR: 0.75, 95%CI: 0.61–0.92) and 38% in men (HR: 0.62; 95%CI: 0.53–0.71) compared to those with an index score of 0 points (poor adherence). Conclusion: Adherence to a healthy Nordic food index was found to be inversely associated with risk of T2D, suggesting that regional diets other than the Mediterranean may also be recommended for prevention of T2D. PMID:26506373

  6. Tracking in real time the crawling dynamics of adherent living cells with a high resolution surface plasmon microscope

    NASA Astrophysics Data System (ADS)

    Streppa, L.; Berguiga, L.; Boyer Provera, E.; Ratti, F.; Goillot, E.; Martinez Torres, C.; Schaeffer, L.; Elezgaray, Juan; Arneodo, A.; Argoul, F.

    2016-03-01

    We introduce a high resolution scanning surface plasmon microscope for long term imaging of living adherent mouse myoblast cells. The coupling of a high numerical aperture objective lens with a fibered heterodyne interferometer provides both enhanced sensitivity and long term stability. This microscope takes advantage of the plasmon resonance excitation and the amplification of the electromagnetic field in near-field distance to the gold coated coverslip. This plasmon enhanced evanescent wave microscopy is particularly attractive for the study of cell adhesion and motility since it can be operated without staining of the biological sample. We show that this microscope allows very long-term imaging of living samples, and that it can capture and follow the temporal deformation of C2C12 myoblast cell protusions (lamellipodia), during their migration on a at surface.

  7. Relationship between adherence to diet, glycemic control and cardiovascular risk factors in patients with type 1 diabetes: a nationwide survey in Brazil

    PubMed Central

    2014-01-01

    Background To determine the relationship between adherence to the diet reported by patients with type 1 diabetes under routine clinical care in Brazil, and demographic, socioeconomic status, glycemic control and cardiovascular risk factors. Methods This was a cross-sectional, multicenter study conducted between December 2008 and December 2010 in 28 public clinics in 20 Brazilian cities. The data was obtained from 3,180 patients, aged 22 ± 11.8 years (56.3% females, 57.4% Caucasians and 43.6% non-Caucasians). The mean time since diabetes diagnosis was 11.7 ± 8.1 years. Results Overall, 1,722 (54.2%) of the patients reported to be adherent to the diet without difference in gender, duration of diabetes and socioeconomic status. Patients who reported adherence to the diet had lower BMI, HbA1c, triglycerides, LDL-cholesterol, non HDL-cholesterol and diastolic blood pressure and had more HbA1c at goal, performed more frequently self-monitoring of blood glucose (p < 0.001), and reported less difficulties to follow specific schedules of diet plans (p < 0.001). Less patients who reported to be adherent were obese or overweight (p = 0.005). The quantity of food and time schedule of the meals were the most frequent complaints. Logistic regression analysis showed that ethnicity, (Caucasians, (OR 1.26 [1.09-1.47]), number of medical clinical visits in the last year (OR 1.10 [1.06-1.15]), carbohydrate counting, (OR 2.22 [1.49-3.30]) and diets recommended by diabetes societies’, (OR 1.57 [1.02-2.41]) were related to greater patients’ adherence (p < 0.05) and age, [adolescents (OR 0.60 [0.50-0.72]), high BMI (OR 0.58 [0.94-0.98]) and smoking (OR 0.58 [0.41-0.84]) with poor patients’ adherence (p < 0.01). Conclusions Our results suggest that it is necessary to rethink medical nutrition therapy in order to help patients to overcome barriers that impair an optimized adherence to the diet. PMID:24607084

  8. The anti-inflammatory drug nimesulide inhibits neutrophil adherence to and migration across monolayers of cytokine-activated endothelial cells.

    PubMed

    Dapino, P; Ottonello, L; Dallegri, F

    1994-01-01

    Neutrophil migration through the microvascular endothelium represents a fundamental event for the cell accumulation at sites of tissue injury. Owing to their capacity to modify the structural and functional characteristics of endothelial cells, inflammatory cytokines such as interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF alpha) play a pivotal role in directing circulating neutrophils away from the bloodstream to the interstitial tissue. In order to study neutrophil transendothelial migration, human umbilical vein endothelial cells were grown to confluence on the polycarbonate filter of two-compartment migration chambers. Pretreatment of the endothelial cell monolayers with TNF alpha for 4 h resulted in rapid migration of approximately 50% of subsequently added neutrophils across the layers. In contrast, < 10% of added neutrophils penetrated untreated endothelial monolayers. Using TNF alpha-treated endothelium, neutrophil transmigration was inhibited by the methane sulfonanilide anti-inflammatory drug nimesulide. Moreover, neutrophil adherence to TNF alpha-treated endothelial monolayers, cultured in microtiter wells, was markedly reduced by nimesulide. A linear correlation between the drug-dependent inhibition of neutrophil transmigration and neutrophil adherence was found. Finally, nimesulide did not interfere with the TNF alpha ability to convert resting endothelium into a pro-adhesive and pro-locomotory cell layer. The data suggest that nimesulide reduces neutrophil transendothelial migration primarily by limiting the cell anchorage to the TNF alpha-activated endothelium. Therefore, the drug has the potential to down-regulate neutrophil extravasation and, in turn, the burden of neutrophil oxidants and proteases leading to tissue injury at sites of inflammation. PMID:7824814

  9. Automated Method for the Rapid and Precise Estimation of Adherent Cell Culture Characteristics from Phase Contrast Microscopy Images

    PubMed Central

    Jaccard, Nicolas; Griffin, Lewis D; Keser, Ana; Macown, Rhys J; Super, Alexandre; Veraitch, Farlan S; Szita, Nicolas

    2014-01-01

    The quantitative determination of key adherent cell culture characteristics such as confluency, morphology, and cell density is necessary for the evaluation of experimental outcomes and to provide a suitable basis for the establishment of robust cell culture protocols. Automated processing of images acquired using phase contrast microscopy (PCM), an imaging modality widely used for the visual inspection of adherent cell cultures, could enable the non-invasive determination of these characteristics. We present an image-processing approach that accurately detects cellular objects in PCM images through a combination of local contrast thresholding and post hoc correction of halo artifacts. The method was thoroughly validated using a variety of cell lines, microscope models and imaging conditions, demonstrating consistently high segmentation performance in all cases and very short processing times (<1 s per 1,208 × 960 pixels image). Based on the high segmentation performance, it was possible to precisely determine culture confluency, cell density, and the morphology of cellular objects, demonstrating the wide applicability of our algorithm for typical microscopy image processing pipelines. Furthermore, PCM image segmentation was used to facilitate the interpretation and analysis of fluorescence microscopy data, enabling the determination of temporal and spatial expression patterns of a fluorescent reporter. We created a software toolbox (PHANTAST) that bundles all the algorithms and provides an easy to use graphical user interface. Source-code for MATLAB and ImageJ is freely available under a permissive open-source license. Biotechnol. Bioeng. 2014;111: 504–517. © 2013 Wiley Periodicals, Inc. PMID:24037521

  10. Receptor-like glycocompounds in human milk that inhibit classical and El Tor Vibrio cholerae cell adherence (hemagglutination).

    PubMed Central

    Holmgren, J; Svennerholm, A M; Lindblad, M

    1983-01-01

    The two biotypes of Vibrio cholerae were found to have cell-associated hemagglutinins which differ with regard to binding to different species of erythrocytes and inhibition by monosaccharides. A total of 12 classical V. cholerae strains (Inaba or Ogawa) strongly agglutinated human erythrocytes in a reaction specifically inhibited by L-fucose, whereas 12 El Tor strains preferably agglutinated chicken erythrocytes, a reaction reversed by D-mannose or by higher concentrations of D-fructose, D-glucose, alpha-methyl-D-mannoside, or sucrose. Milk from Swedish women inhibited both of these adherence reactions, and the predominating inhibitory activity for each reaction resisted boiling, was destroyed by periodate treatment, and bound a concanavalin A-Sepharose column, suggesting a carbohydrate structure. Further characterization indicated that the inhibitory activity for classical V. cholerae hemagglutination was distributed about equally on glycoprotein and free oligosaccharide, but was not present on glycolipid. The El Tor inhibiting activity, on the other hand, was almost exclusively of a high-molecular-weight glycoprotein nature. These results support our previous suggestion (Holmgren et al., Infect. Immun. 33:136-141, 1981) that human milk may contain receptor-like glycocompounds which can prevent bacterial adherence by competition with receptors on target cells. PMID:6295953

  11. Dual Pili Post-translational Modifications Synergize to Mediate Meningococcal Adherence to Platelet Activating Factor Receptor on Human Airway Cells

    PubMed Central

    Schulz, Benjamin L.; Power, Peter M.; Swords, W. Edward; Weiser, Jeffery N.; Apicella, Michael A.; Edwards, Jennifer L.; Jennings, Michael P.

    2013-01-01

    Pili of pathogenic Neisseria are major virulence factors associated with adhesion, twitching motility, auto-aggregation, and DNA transformation. Pili of N. meningitidis are subject to several different post-translational modifications. Among these pilin modifications, the presence of phosphorylcholine (ChoP) and a glycan on the pilin protein are phase-variable (subject to high frequency, reversible on/off switching of expression). In this study we report the location of two ChoP modifications on the C-terminus of N. meningitidis pilin. We show that the surface accessibility of ChoP on pili is affected by phase variable changes to the structure of the pilin-linked glycan. We identify for the first time that the platelet activating factor receptor (PAFr) is a key, early event receptor for meningococcal adherence to human bronchial epithelial cells and tissue, and that synergy between the pilin-linked glycan and ChoP post-translational modifications is required for pili to optimally engage PAFr to mediate adherence to human airway cells. PMID:23696740

  12. Spatially and temporally controlled gene transfer by electroporation into adherent cells on plasmid DNA-loaded electrodes.

    PubMed

    Yamauchi, Fumio; Kato, Koichi; Iwata, Hiroo

    2004-01-01

    Functional characterization of human genes is one of the most challenging tasks in current genomics. Owing to a large number of newly discovered genes, high-throughput methodologies are greatly needed to express in parallel each gene in living cells. To develop a method that allows efficient transfection of plasmids into adherent cells in spatial- and temporal-specific manners, we studied electric pulse-triggered gene transfer using a plasmid-loaded electrode. A plasmid was loaded on a gold electrode surface having an adsorbed layer of poly(ethyleneimine), and cells were then plated directly onto this modified surface. The plasmid was detached from the electrode by applying a short electric pulse and introduced into the cells cultured on the electrode, resulting in efficient gene expression, even in primary cultured cells. The location of transfected cells could be restricted within a small area on a micropatterned electrode, showing the versatility of the method for spatially controlled transfection. Plasmid transfection could also be performed in a temporally controlled manner without a marked loss of the efficiency when an electric pulse was applied within 3 days after cell plating. The method described here will provide an efficient means to transfer multiple genes, in parallel, into cultured mammalian cells for high-throughput reverse genetics research. PMID:15613595

  13. Distinct sites on tenascin-C mediate repellent or adhesive interactions with different neuronal cell types.

    PubMed

    Husmann, K; Carbonetto, S; Schachner, M

    1995-11-01

    In this study we have determined the binding specificities of four different neuronal cell types to tenascin-C (TN-C) and laminin using a cell adhesion assay. TN-C was repulsive for small cerebellar neurons and PC12 phaeochromocytoma cells, since after short-term adhesion to the substrate-bound molecule with a maximum of cell binding at 45 min, the cells detached from the substrate and after 22 h only about 25% of the originally adherent cells were still bound. For N2A neuroblastoma cells and retinal cells TN-C was an adhesive substrate, since the number of adherent cells did not decrease after the initial attachment period. All four cell types adhered well to laminin at all time points studied. For short-term adhesion of small cerebellar neurons and PC12 cells two binding sites were identified on TN-C, one being localized within the epidermal growth factor-like repeats three to five and the second within fibronectin type III-like repeats three and four. One binding site for N2A and retinal cells was localized within fibronectin type III-like repeat seven. Binding of small cerebellar neurons to TN-C was dependent on Ca2+, but not on Mg2+ and was inhibitable by polyclonal antibodies to beta 1 integrin. Short-term adhesion of small cerebellar neurons was also inhibitable with a mixture of recombinant fragments of TN-C encompassing the whole molecule, although the specific inhibitory activity of this mixture was ten-fold lower on a molar basis when compared to the native molecule. Our observations indicate that different neuronal cell types use distinct binding sites on TN-C for repellent or adhesive interactions and that beta 1 integrin is involved in the recognition event leading to repulsion of small cerebellar neurons. PMID:8821032

  14. High-molecular-mass lipopolysaccharides are involved in Actinobacillus pleuropneumoniae adherence to porcine respiratory tract cells.

    PubMed Central

    Paradis, S E; Dubreuil, D; Rioux, S; Gottschalk, M; Jacques, M

    1994-01-01

    Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia. The major adhesin of A. pleuropneumoniae has been identified as the lipopolysaccharides (LPSs) (M. Bélanger, D. Dubreuil, J. Harel, C. Girard, and M. Jacques, Infect. Immun. 58:3523-3530, 1990). Using immunoelectron microscopy and flow cytometry, we showed in the present study that LPSs were well exposed at the surface of this encapsulated microorganism. Immunolocalization with porcine lung and tracheal frozen sections showed that extracted LPS bound to the lung mesenchyme and vascular endothelium and to the tracheal epithelium, respectively. Inhibition of adherence of A. pleuropneumoniae with extracted LPS was also performed with lung and tracheal frozen sections. Acid hydrolysis of LPS revealed that the active component of LPS was not lipid A but the polysaccharides. LPSs from A. pleuropneumoniae serotypes 1 and 2 were separated by chromatography on Sephacryl S-300 SF, in the presence of sodium deoxycholate, according to their molecular masses. The adherence-inhibitory activity was found in the high-molecular-mass fractions. These high-molecular-mass fractions contained 2-keto-3-deoxyoctulosonic acid and neutral sugars, and they were recognized by a monoclonal antibody directed against A. pleuropneumoniae O antigen but not recognized by a monoclonal antibody against capsular antigen. Images PMID:8039902

  15. Pulmonary Alveolar Type II Cells Isolated from Rats

    PubMed Central

    Dobbs, Leland G.; Mason, Robert J.

    1979-01-01

    It is unclear what factors control the secretion of pulmonary surface active material from alveolar type II cells in vivo. Other workers have suggested that cholinergic stimuli, adrenergic stimuli, and prostaglandins may all stimulate secretion. We isolated type II cells from the lungs of rats by treatment with elastase, discontinuous density centrifugation, and adherence in primary culture. β-Adrenergic agonists, but not cholinergic agonists, caused an increase in the release of [14C]disaturated phosphatidylcholine, the major component of surface-active material, from type II cells in culture. The β-adrenergic effect was stereo-selective, (−)-isoproterenol being 50 times more potent than (+)-isoproterenol. Terbutaline, 10 μM, a noncatecholamine β-2 adrenergic agonist, caused a release of 2.0±0.5 (mean±SD) times the basal release of [14C]disaturated phosphatidylcholine in 3 h; the concentration of terbutaline causing half maximal stimulation was 800 nM. The terbutaline effect was blocked by propranolol, a β-adrenergic antagonist (calculated Kd = 6 nM), but not by phentolamine, an α-adrenergic antagonist. Isobutylmethylxanthine, a phosphodiesterase inhibitor, and 8-Br cyclic AMP, but not 8-Br cyclic guanosine monophosphate, also stimulated release. We conclude that type II cells secrete disaturated phosphatidylcholine in response to treatment with adrenergic stimulation. PMID:34631

  16. Heat-labile enterotoxin-induced activation of NF-κB and MAPK pathways in intestinal epithelial cells impacts enterotoxigenic Escherichia coli (ETEC) adherence.

    PubMed

    Wang, Xiaogang; Gao, Xiaofei; Hardwidge, Philip R

    2012-08-01

    Enterotoxigenic Escherichia coli (ETEC) causes human morbidity and mortality in developing nations and is an emerging threat to food safety in developed nations. The ETEC heat-labile enterotoxin (LT) not only causes diarrheal disease by deregulating host adenylate cyclase, but also enhances ETEC adherence to intestinal epithelial cells. The mechanism governing this LT pro-adherence phenotype is unclear. Here we investigated intestinal epithelial cell signal transduction pathways activated by ETEC and quantified the relative importance of these host pathways to LT-induced ETEC adherence. We show that ETEC activates both NF-κB and mitogen-activated protein kinase signalling pathways through mechanisms that are primarily dependent upon LT. LT-induced NF-κB activation depends upon the cAMP-dependent activation of the Ras-like GTPase Rap1 but is independent of protein kinase A (PKA). By using inhibitors of these pathways, we demonstrate that inhibiting the p38 mitogen-activated protein kinase prevents LT from increasing ETEC adherence. By contrast, the LT pro-adherence phenotype appears unrelated to both LT-induced Rap1 activity and to subsequent NF-κB activation. We speculate that LT may alter host signal transduction to induce the presentation of ligands for ETEC adhesins in such a way that promotes ETEC adherence. Our findings provide insight into previously unexplored functions of LT and their relative importance to ETEC virulence. PMID:22452361

  17. Pilus phase variation switches gonococcal adherence to invasion by caveolin-1-dependent host cell signaling.

    PubMed

    Faulstich, Michaela; Böttcher, Jan-Peter; Meyer, Thomas F; Fraunholz, Martin; Rudel, Thomas

    2013-01-01

    Many pathogenic bacteria cause local infections but occasionally invade into the blood stream, often with fatal outcome. Very little is known about the mechanism underlying the switch from local to invasive infection. In the case of Neisseria gonorrhoeae, phase variable type 4 pili (T4P) stabilize local infection by mediating microcolony formation and inducing anti-invasive signals. Outer membrane porin PorB(IA), in contrast, is associated with disseminated infection and facilitates the efficient invasion of gonococci into host cells. Here we demonstrate that loss of pili by natural pilus phase variation is a prerequisite for the transition from local to invasive infection. Unexpectedly, both T4P-mediated inhibition of invasion and PorB(IA)-triggered invasion utilize membrane rafts and signaling pathways that depend on caveolin-1-Y14 phosphorylation (Cav1-pY14). We identified p85 regulatory subunit of PI3 kinase (PI3K) and phospholipase Cγ1 as new, exclusive and essential interaction partners for Cav1-pY14 in the course of PorBIA-induced invasion. Active PI3K induces the uptake of gonococci via a new invasion pathway involving protein kinase D1. Our data describe a novel route of bacterial entry into epithelial cells and offer the first mechanistic insight into the switch from local to invasive gonococcal infection. PMID:23717204

  18. Basic Leucine Zipper (bZIP) Domain Transcription Factor MBZ1 Regulates Cell Wall Integrity, Spore Adherence, and Virulence in Metarhizium robertsii *

    PubMed Central

    Huang, Wei; Shang, Yanfang; Chen, Peilin; Cen, Kai; Wang, Chengshu

    2015-01-01

    Transcription factors (TFs) containing the basic leucine zipper (bZIP) domain are widely distributed in eukaryotes and display an array of distinct functions. In this study, a bZIP-type TF gene (MBZ1) was deleted and functionally characterized in the insect pathogenic fungus Metarhizium robertsii. The deletion mutant (ΔMBZ1) showed defects in cell wall integrity, adhesion to hydrophobic surfaces, and topical infection of insects. Relative to the WT, ΔMBZ1 was also impaired in growth and conidiogenesis. Examination of putative target gene expression indicated that the genes involved in chitin biosynthesis were differentially transcribed in ΔMBZ1 compared with the WT, which led to the accumulation of a higher level of chitin in mutant cell walls. MBZ1 exhibited negative regulation of subtilisin proteases, but positive control of an adhesin gene, which is consistent with the observation of effects on cell autolysis and a reduction in spore adherence to hydrophobic surfaces in ΔMBZ1. Promoter binding assays indicated that MBZ1 can bind to different target genes and suggested the possibility of heterodimer formation to increase the diversity of the MBZ1 regulatory network. The results of this study advance our understanding of the divergence of bZIP-type TFs at both intra- and interspecific levels. PMID:25673695

  19. Characterization of an ExoS Type III Translocation-Resistant Cell Line

    PubMed Central

    Rucks, Elizabeth A.; Olson, Joan C.

    2005-01-01

    Pseudomonas aeruginosa ExoS is a type III-secreted type III-secreted, bifunctional protein that causes diverse effects on eukaryotic cell function. The coculture of P. aeruginosa strains expressing ExoS with HL-60 myeloid cells revealed the cell line to be resistant to the toxic effects of ExoS. Differentiation of HL-60 cells with phorbol 12-myristate 13-acetate (TPA) rendered the cell line sensitive to ExoS. To understand the cellular basis for the alteration in sensitivity, undifferentiated and TPA-differentiated HL-60 cells were compared for differences in bacterial adherence, type III secretion induction, and ExoS translocation. These comparisons found that ExoS was translocated more efficiently in TPA-differentiated HL-60 cells than in undifferentiated cells. The studies support the ability of eukaryotic cells to influence P. aeruginosa TTS at the level of membrane translocation. PMID:15618208

  20. Impact on disinfection efficiency of cell load and of planktonic/adherent/detached state: case of Hafnia alvei inactivation by plasma activated water.

    PubMed

    Kamgang-Youbi, Georges; Herry, Jean-Marie; Brisset, Jean-Louis; Bellon-Fontaine, Marie-Noëlle; Doubla, Avaly; Naïtali, Murielle

    2008-12-01

    This paper describes the effects of initial microbial concentration and planktonic/adherent/detached states on the efficiency of plasma-activated water. This disinfecting solution was obtained by treating distilled water with an atmospheric pressure plasma produced by gliding electric discharges in humid air. The inactivation kinetics of planktonic cells of Hafnia alvei (selected as a bacterial model) were found to be of the first order. They were influenced by the initial microbial concentration. Efficiency decreased when the initial viable population N(0) increased, and the inactivation rate k(max) was linearly modified as a function of Log(10) (N(0)). This relation was used to compare planktonic, adherent, and detached cells independently from the level of population. Bacteria adhering to stainless steel and high-density polyethylene were also sensitive to treatment, but at a lower rate than their free-living counterparts. Moreover, cells detached from these solid substrates exhibited an inactivation rate lower than that of planktonic cells but similar to adherent bacteria. This strongly suggests the induction of a physiological modification to bacteria during the adhesion step, rendering adherent--and further detached--bacteria less susceptible to the treatment, when compared to planktonic bacteria. PMID:18769918

  1. Single cell dual adherent-suspension co-culture micro-environment for studying tumor-stromal interactions with functionally selected cancer stem-like cells.

    PubMed

    Chen, Yu-Chih; Zhang, Zhixiong; Fouladdel, Shamileh; Deol, Yadwinder; Ingram, Patrick N; McDermott, Sean P; Azizi, Ebrahim; Wicha, Max S; Yoon, Euisik

    2016-08-01

    Considerable evidence suggests that cancer stem-like cells (CSCs) are critical in tumor pathogenesis, but their rarity and transience has led to much controversy about their exact nature. Although CSCs can be functionally identified using dish-based tumorsphere assays, it is difficult to handle and monitor single cells in dish-based approaches; single cell-based microfluidic approaches offer better control and reliable single cell derived sphere formation. However, like normal stem cells, CSCs are heavily regulated by their microenvironment, requiring tumor-stromal interactions for tumorigenic and proliferative behaviors. To enable single cell derived tumorsphere formation within a stromal microenvironment, we present a dual adherent/suspension co-culture device, which combines a suspension environment for single-cell tumorsphere assays and an adherent environment for co-culturing stromal cells in close proximity by selectively patterning polyHEMA in indented microwells. By minimizing dead volume and improving cell capture efficiency, the presented platform allows for the use of small numbers of cells (<100 cells). As a proof of concept, we co-cultured single T47D (breast cancer) cells and primary cancer associated fibroblasts (CAF) on-chip for 14 days to monitor sphere formation and growth. Compared to mono-culture, co-cultured T47D have higher tumorigenic potential (sphere formation rate) and proliferation rates (larger sphere size). Furthermore, 96-multiplexed single-cell transcriptome analyses were performed to compare the gene expression of co-cultured and mono-cultured T47D cells. Phenotypic changes observed in co-culture correlated with expression changes in genes associated with proliferation, apoptotic suppression, tumorigenicity and even epithelial-to-mesechymal transition. Combining the presented platform with single cell transcriptome analysis, we successfully identified functional CSCs and investigated the phenotypic and transcriptome effects induced

  2. Plasticity of Hopx+ Type I alveolar cells to regenerate Type II cells in the lung

    PubMed Central

    Jain, Rajan; Barkauskas, Christina E.; Takeda, Norifumi; Bowie, Emily J.; Aghajanian, Haig; Wang, Qiaohong; Padmanabhan, Arun; Manderfield, Lauren J.; Gupta, Mudit; Li, Deqiang; Li, Li; Trivedi, Chinmay M.; Hogan, Brigid L. M.; Epstein, Jonathan A.

    2015-01-01

    The plasticity of differentiated cells in adult tissues undergoing repair is an area of intense research. Pulmonary alveolar Type II cells produce surfactant and function as progenitors in the adult, demonstrating both self-renewal and differentiation into gas exchanging Type I cells. In vivo, Type I cells are thought to be terminally differentiated and their ability to give rise to alternate lineages has not been reported. Here, we show that Hopx becomes restricted to Type I cells during development. However, unexpectedly, lineage-labeled Hopx+ cells both proliferate and generate Type II cells during adult alveolar regrowth following partial pneumonectomy. In clonal 3D culture, single Hopx+ Type I cells generate organoids composed of Type I and Type II cells, a process modulated by TGFβ signaling. These findings demonstrate unanticipated plasticity of Type I cells and a bi-directional lineage relationship between distinct differentiated alveolar epithelial cell types in vivo and in single cell culture. PMID:25865356

  3. Plasticity of Hopx(+) type I alveolar cells to regenerate type II cells in the lung.

    PubMed

    Jain, Rajan; Barkauskas, Christina E; Takeda, Norifumi; Bowie, Emily J; Aghajanian, Haig; Wang, Qiaohong; Padmanabhan, Arun; Manderfield, Lauren J; Gupta, Mudit; Li, Deqiang; Li, Li; Trivedi, Chinmay M; Hogan, Brigid L M; Epstein, Jonathan A

    2015-01-01

    The plasticity of differentiated cells in adult tissues undergoing repair is an area of intense research. Pulmonary alveolar type II cells produce surfactant and function as progenitors in the adult, demonstrating both self-renewal and differentiation into gas exchanging type I cells. In vivo, type I cells are thought to be terminally differentiated and their ability to give rise to alternate lineages has not been reported. Here we show that Hopx becomes restricted to type I cells during development. However, unexpectedly, lineage-labelled Hopx(+) cells both proliferate and generate type II cells during adult alveolar regrowth following partial pneumonectomy. In clonal 3D culture, single Hopx(+) type I cells generate organoids composed of type I and type II cells, a process modulated by TGFβ signalling. These findings demonstrate unanticipated plasticity of type I cells and a bidirectional lineage relationship between distinct differentiated alveolar epithelial cell types in vivo and in single-cell culture. PMID:25865356

  4. Protocol for SAMS (Support and Advice for Medication Study): A randomised controlled trial of an intervention to support patients with type 2 diabetes with adherence to medication

    PubMed Central

    Farmer, Andrew J; Prevost, A Toby; Hardeman, Wendy; Craven, Anthea; Sutton, Stephen; Griffin, Simon J; Kinmonth, Ann-Louise

    2008-01-01

    Background Although some interventions have been shown to improve adherence to medication for diabetes, results are not consistent. We have developed a theory-based intervention which we will evaluate in a well characterised population to test efficacy and guide future intervention development and trial design. Methods and Design The SAMS (Supported Adherence to Medication Study) trial is a primary care based multi-centre randomised controlled trial among 200 patients with type 2 diabetes and an HbA1c of 7.5% or above. It is designed to evaluate the efficacy of a two-component motivational intervention based on the Theory of Planned Behaviour and volitional action planning to support medication adherence compared with standard care. The intervention is delivered by practice nurses. Nurses were trained using a workshop approach with role play and supervised using assessment of tape-recorded consultations. The trial has a two parallel groups design with an unbalanced three-to-two individual randomisation eight weeks after recruitment with twelve week follow-up. The primary outcome is medication adherence measured using an electronic medication monitor over 12 weeks and expressed as the difference between intervention and control in mean percentage of days on which the correct number of medication doses is taken. Subgroup analyses will explore impact of number of medications taken, age, HbA1c, and self-reported adherence at baseline on outcomes. The study also measures the effect of dispensing medication to trial participants packaged in the electronic medication-monitoring device compared with conventional medication packaging. This will be achieved through one-to-one randomisation at recruitment to these conditions with assessment of the difference between groups in self-report of medication adherence and change in mean HbA1c from baseline to eight weeks. Anonymised demographic data are collected on non-respondents. Central randomisation is carried out independently

  5. Adhering heat-killed human Lactobacillus acidophilus, strain LB, inhibits the process of pathogenicity of diarrhoeagenic bacteria in cultured human intestinal cells.

    PubMed

    Coconnier, M H; Bernet, M F; Chauvière, G; Servin, A L

    1993-12-01

    Heat-killed L. acidophilus, strain LB, was tested for its ability to adhere in vitro onto human enterocyte-like Caco-2 and muco-secreting HT29-MTX cells in culture. The heat-killed LB bacteria exhibited a high adhesive property. A diffuse pattern of adhesion was observed to the undifferentiated cells, the apical brush border of the enterocytic cells, and to the mucus layer that covered the surface of the mucus-secreting cells. The inhibitory effect of heat-killed LB organisms against the human intestinal Caco-2 cell-adhesion and cell-invasion by a large variety of diarrhoeagenic bacteria was investigated. The following dose-dependent inhibitions were obtained: (i) against the cell-association of enterotoxigenic, diffusely-adhering and enteropathogenic Escherichia coli, Listeria monocytogenes, Yersinia pseudotuberculosis, and Salmonella typhimurium; (ii) against the cell-invasion by enteropathogenic Escherichia coli, Yersinia pseudotuberculosis, Listeria monocytogenes and Salmonella typhimurium. PMID:8188996

  6. Determinants of adherence to self-care behavior among women with type 2 diabetes: an explanation based on health belief model

    PubMed Central

    Karimy, Mahmood; Araban, Marzieh; Zareban, Iraj; Taher, Mohammad; Abedi, Ahmadreza

    2016-01-01

    Background: Self-care is an essential element in treating a person with diabetes; and managing diabetes is of prime importance. The aim of this study was to investigate the predictors of adherence to self-care behavior among women with Type 2 diabetes. Methods: This cross-sectional study was conducted on 210 female patients aged 30 to 60. Data collection tool was an anonymous valid and reliable questionnaire designed based on the Health Belief Model (HBM), which acquired information about the followings: Perceived susceptibility, severity, benefits, barriers, self-efficacy and diabetes self-care behavior. Data were analyzed by t-test, chisquare and regression analysis. Results: The multiple regression models revealed 59.9% of the variance of self-care behavior with self-efficacy, perceived barrier, benefit and susceptibility. Additionally, the highest weight for β (β=0.87) was found for self-efficacy. Self-care behavior was positively correlated with all HBM variables except for perceived barriers showing a negative correlation. Conclusion: The Health Belief Model may be used as a framework to design intervention programs in an attempt to improve adherence to self-care behaviors of women with diabetes. In addition, the results indicated that self-efficacy might play a more crucial role in developing self-care behaviors than t other HBM components. Therefore, if the focus is placed on self-efficacy when developing educational programs, it may increase the likelihood of adherence to self-care behavior. PMID:27493912

  7. Inhibition of mitogenesis induced by phytohemagglutinin and Lens culinaris lectin in adherent-cell supernatants treated with protein extract of Mycobacterium tuberculosis.

    PubMed Central

    Parra, C; Montaño, L F; Huesca, M; Rayón, I; Willms, K; Goodsaid, F

    1986-01-01

    Specific stimulation of T cells by phytohemagglutinin and Lens culinaris lectin was inhibited by a soluble factor(s) secreted by normal adherent cells stimulated with culture filtrate protein extract (CFPE) derived from bacterial cultures of Mycobacterium tuberculosis H37Ra (avirulent) and H37Rv (virulent). The induction of the inhibitory factor was blocked by the presence of hyperimmune antisera to H37Rv or H37Ra CFPE. The inhibitory factor did not seem to be a CFPE reprocessed by the adherent cells. Inhibitory activity was maximal in supernatants of adherent-cell cultures incubated for 48 h; the inhibitory factor was heat labile, and its production was dependent on the concentration of M. tuberculosis CFPE. A mouse monocyte-macrophage cell line, ATCC J774A.1, produced an identical inhibitory factor, thus excluding a non-macrophage-contaminating adherent cell as the source of the factor. This inhibitory factor also interfered with the recognition of phytohemagglutinin and Lens culinaris lectin by T cells. PMID:3082760

  8. A Serine-Threonine Kinase (StkP) Regulates Expression of the Pneumococcal Pilus and Modulates Bacterial Adherence to Human Epithelial and Endothelial Cells In Vitro

    PubMed Central

    Herbert, Jenny A.; Mitchell, Andrea M.; Mitchell, Timothy J.

    2015-01-01

    The pneumococcal serine threonine protein kinase (StkP) acts as a global regulator in the pneumococcus. Bacterial mutants deficient in StkP are less virulent in animal models of infection. The gene for this regulator is located adjacent to the gene for its cognate phosphatase in the pneumococcal genome. The phosphatase dephosphorylates proteins phosphorylated by StkP and has been shown to regulate a number of key pneumococcal virulence factors and to modulate adherence to eukaryotic cells. The role of StkP in adherence of pneumococci to human cells has not previously been reported. In this study we show StkP represses the pneumococcal pilus, a virulence factor known to be important for bacterial adhesion. In a serotype 4 strain regulation of the pilus by StkP modulates adherence to human brain microvascular endothelial cells (HBMEC) and human lung epithelial cells. This suggests that the pneumococcal pilus may play a role in adherence during infections such as meningitis and pneumonia. We show that regulation of the pilus occurs at the population level as StkP alters the number of pili-positive cells within a single culture. As far as we are aware this is the first gene identified outside of the pilus islet that regulates the biphasic expression of the pilus. These findings suggest StkPs role in cell division may be linked to regulation of expression of a cell surface adhesin. PMID:26090876

  9. Mitigation of Lethal Radiation Syndrome in Mice by Intramuscular Injection of 3D Cultured Adherent Human Placental Stromal Cells.

    PubMed

    Gaberman, Elena; Pinzur, Lena; Levdansky, Lilia; Tsirlin, Maria; Netzer, Nir; Aberman, Zami; Gorodetsky, Raphael

    2013-01-01

    Exposure to high lethal dose of ionizing radiation results in acute radiation syndrome with deleterious systemic effects to different organs. A primary target is the highly sensitive bone marrow and the hematopoietic system. In the current study C3H/HeN mice were total body irradiated by 7.7 Gy. Twenty four hrs and 5 days after irradiation 2×10(6) cells from different preparations of human derived 3D expanded adherent placental stromal cells (PLX) were injected intramuscularly. Treatment with batches consisting of pure maternal cell preparations (PLX-Mat) increased the survival of the irradiated mice from ∼27% to 68% (P<0.001), while cell preparations with a mixture of maternal and fetal derived cells (PLX-RAD) increased the survival to ∼98% (P<0.0001). The dose modifying factor of this treatment for both 50% and 37% survival (DMF50 and DMF37) was∼1.23. Initiation of the more effective treatment with PLX-RAD injection could be delayed for up to 48 hrs after irradiation with similar effect. A delayed treatment by 72 hrs had lower, but still significantly effect (p<0.05). A faster recovery of the BM and improved reconstitution of all blood cell lineages in the PLX-RAD treated mice during the follow-up explains the increased survival of the cells treated irradiated mice. The number of CD45+/SCA1+ hematopoietic progenitor cells within the fast recovering population of nucleated BM cells in the irradiated mice was also elevated in the PLX-RAD treated mice. Our study suggests that IM treatment with PLX-RAD cells may serve as a highly effective "off the shelf" therapy to treat BM failure following total body exposure to high doses of radiation. The results suggest that similar treatments may be beneficial also for clinical conditions associated with severe BM aplasia and pancytopenia. PMID:23823334

  10. Mitigation of Lethal Radiation Syndrome in Mice by Intramuscular Injection of 3D Cultured Adherent Human Placental Stromal Cells

    PubMed Central

    Gaberman, Elena; Pinzur, Lena; Levdansky, Lilia; Tsirlin, Maria; Netzer, Nir; Aberman, Zami; Gorodetsky, Raphael

    2013-01-01

    Exposure to high lethal dose of ionizing radiation results in acute radiation syndrome with deleterious systemic effects to different organs. A primary target is the highly sensitive bone marrow and the hematopoietic system. In the current study C3H/HeN mice were total body irradiated by 7.7 Gy. Twenty four hrs and 5 days after irradiation 2×106 cells from different preparations of human derived 3D expanded adherent placental stromal cells (PLX) were injected intramuscularly. Treatment with batches consisting of pure maternal cell preparations (PLX-Mat) increased the survival of the irradiated mice from ∼27% to 68% (P<0.001), while cell preparations with a mixture of maternal and fetal derived cells (PLX-RAD) increased the survival to ∼98% (P<0.0001). The dose modifying factor of this treatment for both 50% and 37% survival (DMF50 and DMF37) was∼1.23. Initiation of the more effective treatment with PLX-RAD injection could be delayed for up to 48 hrs after irradiation with similar effect. A delayed treatment by 72 hrs had lower, but still significantly effect (p<0.05). A faster recovery of the BM and improved reconstitution of all blood cell lineages in the PLX-RAD treated mice during the follow-up explains the increased survival of the cells treated irradiated mice. The number of CD45+/SCA1+ hematopoietic progenitor cells within the fast recovering population of nucleated BM cells in the irradiated mice was also elevated in the PLX-RAD treated mice. Our study suggests that IM treatment with PLX-RAD cells may serve as a highly effective “off the shelf” therapy to treat BM failure following total body exposure to high doses of radiation. The results suggest that similar treatments may be beneficial also for clinical conditions associated with severe BM aplasia and pancytopenia. PMID:23823334

  11. Short message service (SMS) reminders and real-time adherence monitoring improve antiretroviral therapy adherence in rural Uganda

    PubMed Central

    Haberer, Jessica E.; Musiimenta, Angella; Atukunda, Esther C.; Musinguzi, Nicholas; Wyatt, Monique A.; Ware, Norma C.; Bangsberg, David R.

    2016-01-01

    Objective: To explore the effects of four types of short message service (SMS) plus real-time adherence monitoring on antiretroviral therapy (ART) adherence: daily reminders, weekly reminders, reminders triggered after a late or missed dose (delivered to patients), and notifications triggered by sustained adherence lapses (delivered to patient-nominated social supporters). Design: Pilot randomized controlled trial. Methods: Sixty-three individuals initiating ART received a real-time adherence monitor and were randomized (1 : 1 : 1): (1) Scheduled SMS reminders (daily for 1 month, weekly for 2 months), then SMS reminders triggered by a late or missed dose (no monitoring signal within 2 h of expected dosing); SMS notifications to social supporters for sustained adherence lapses (no monitoring signal for >48 h) added after 3 months. (2) Triggered SMS reminders starting at enrolment; SMS notifications to social supporters added after 3 months. (3) Control: No SMS. HIV RNA was determined at 9 months. Percentage adherence and adherence lapses were compared by linear generalized estimating equations and Poisson regression, respectively. Results: Median age was 31 years, 65% were women, and median enrolment CD4+ cell count was 322 cells/μl 97% took once daily tenofovir/emtricitabine/efavirenz. Compared to control, adherence was 11.1% higher (P = 0.04) and more than 48-h lapses were less frequent (IRR 0.6, P = 0.02) in the scheduled SMS arm. Adherence and more than 48-h lapses were similar in the triggered SMS arm and control. No differences in HIV RNA were seen. Conclusion: Scheduled SMS reminders improved ART in the context of real-time monitoring. Larger studies are needed to determine the impact of triggered reminders and role of social supporters in improving adherence. PMID:26760452

  12. HIV Medication Adherence

    MedlinePlus

    HIV Treatment HIV Medication Adherence (Last updated 3/1/2016; last reviewed 3/1/2016) Key Points Medication adherence means sticking ... exactly as prescribed. Why is adherence to an HIV regimen important? Adherence to an HIV regimen gives ...

  13. A large mobility of hydrophilic molecules at the outmost layer controls the protein adsorption and adhering behavior with the actin fiber orientation of human umbilical vein endothelial cells (HUVEC).

    PubMed

    Kakinoki, Sachiro; Seo, Ji-Hun; Inoue, Yuuki; Ishihara, Kazuhiko; Yui, Nobuhiko; Yamaoka, Tetsuji

    2013-01-01

    Adhesion behaviors of human umbilical vein endothelial cells (HUVECs) are interestingly affected by the mobility of hydrophilic chains on the material surfaces. Surfaces with different molecular mobilities were prepared using ABA-type block copolymers consisting polyrotaxane (PRX) or poly(ethylene glycol) (PEG) central block (A block), and amphiphilic anchoring B blocks of poly(2-methacryloyloxyethyl phosphorylcholine-co-n-butyl methacrylate) (PMB). Two different molecular mobilities of the PRX chains were designed by using normal α-cyclodextrin (α-CD) or α-CD whose hydroxyl groups were converted to methoxy groups in a given ratio to improve its molecular mobility (PRX-PMB and OMe-PRX-PMB). The surface mobility of these materials was assessed as the mobility factor (Mf), which is measured by quartz crystal microbalance with dissipation monitoring system. HUVECs adhered on OMe-PRX-PMB surface much more than PRX-PMB and PMB-block-PEG-block-PMB (PEG-PMB) surfaces. These different HUVEC adhesions were correlated with the density of cell-binding site of adsorbed fibronectin. In addition, the alignment of the actin cytoskeleton of adhered HUVECs was strongly suppressed on the PEG-PMB, PRX-PMB, and OMe-PRX-PMB in response to the increased Mf value. Remarkably, the HUVECs adhered on the OMe-PRX-PMB surface with much less actin organization. We concluded that not only the cell adhesion but also the cellular function are regulated by the molecular mobility of the outmost material surfaces. PMID:23796033

  14. Parenting style, parent-youth conflict, and medication adherence in youth with type 2 diabetes participating in an intensive lifestyle change intervention.

    PubMed

    Saletsky, Ronald D; Trief, Paula M; Anderson, Barbara J; Rosenbaum, Paula; Weinstock, Ruth S

    2014-06-01

    Parenting behaviors and family conflict relate to type 1 diabetes outcomes in youth. Our purpose was to understand these relationships in parents and youth with type 2 diabetes (T2DM). The TODAY (Treatment Options for Type 2 Diabetes in Adolescents and Youth) trial enrolled youth (10-17 years) with T2DM and parent/guardian. For this ancillary study, we enrolled a sample of youth-parent pairs (N = 137) in 1 study arm (metformin plus lifestyle intervention). They completed questionnaires measuring parenting style related to normative (e.g., completing homework) and diabetes self-care (e.g., testing blood glucose) tasks, and parent-youth verbal conflict (baseline, 6, and 12 months). Parenting style was consistent across normative and diabetes tasks, with gradual increases in autonomy perceived by youth. Conversations were generally calm, with greater conflict regarding normative than diabetes tasks at baseline (youth: p < .001, parent: p = .01), 6 months (youth: p = .02, parent: p > .05), and 12 months (youth: p > .05., parent: p = .05). A permissive parenting style toward normative tasks and a less authoritarian style toward diabetes tasks, at baseline, predicted better medication adherence (8-12 months) (normative: adjusted R2 = 0.48, p < .001; diabetes: adjusted R2 = 0.47, p < .001). Parent-youth conflict did not predict medication adherence. Youth with T2DM who perceive more autonomy (less parental control) in day-to-day and diabetes tasks are more likely to adhere to medication regimens. It may be valuable to assess youth perceptions of parenting style and help parents understand youths' needs for autonomy. PMID:24548045

  15. Parenting Style, Parent-Youth Conflict, and Medication Adherence in Youth with Type 2 Diabetes Participating in an Intensive Lifestyle Change Intervention

    PubMed Central

    Saletsky, Ronald D.; Trief, Paula M.; Anderson, Barbara J.; Rosenbaum, Paula; Weinstock, Ruth S.

    2014-01-01

    INTRODUCTION Parenting behaviors and family conflict relate to type 1 diabetes outcomes in youth. The purpose of this study was to understand these relationships in parents and youth with type 2 diabetes (T2DM). METHODS The TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) trial enrolled youth (10-17 years) with recent-onset T2DM and parent/guardian. For this ancillary study, we enrolled a sample of youth-parent pairs (N =137) in one TODAY study arm (metformin plus lifestyle intervention). Parents and youths completed questionnaires to assess parenting style related to normative (e.g., completing homework) and diabetes self-care (e.g., testing blood glucose) tasks, and parent-youth verbal conflict (baseline, 6 and 12 months). RESULTS Parenting style was consistent across normative and diabetes tasks, with gradual increases in autonomy perceived by youth. Conversations were generally calm, with greater conflict regarding normative tasks than diabetes tasks at baseline (youth: p<0.001, parent: p=0.01), 6 months (youth: p=0.02, parent: p >0.05) and 12 months (youth: p> 0.05., parent: p=0.05). A permissive parenting style towards normative tasks and a less authoritarian style towards diabetes tasks, at baseline, predicted better medication adherence (8-12 months) (normative: adjusted R2=0.48, p<0.001; diabetes: adjusted R2 = 0.47, p<0.001). Parent-youth conflict did not predict medication adherence. DISCUSSION Youth with T2DM who perceive more autonomy (less parental control) in day-to-day and diabetes tasks are more likely to adhere to medication regimens. It may be valuable to assess youth perceptions of parenting style at onset of medication treatment and help parents understand youths’ needs for autonomy. PMID:24548045

  16. Cystone – An ayurvedic polyherbal formulation inhibits adherence of uropathogenic E. coli and modulates H2O2-induced toxicity in NRK-52E cells

    PubMed Central

    Vidyashankar, Satyakumar; Maheshkumar, Puttanarasaiah; Patki, Pralhad S

    2010-01-01

    Gentamicin is a widely used antibiotic for the treatment of adverse urinary tract infections (UTI), which in turn causes nephrotoxicity to uroepithelial cells and hence an alternative safe herbal remedy is much desired to compensate these toxic effects. The bacterial adhesion to the uroepithelial cells is the primary step in UTI and it induces various immunogenic reactions leading to the generation of reactive oxygen species (ROS), which are detrimental to the cells survival. Inhibition of bacterial adherence to urinary tract epithelial cells has been assumed to account for the beneficial action ascribed to cystone (an ayurvedic polyherbal formulation) in the prevention of UTI. In this study, we have examined the effect of cystone on the adherence of pathogenic [2-14C]-acetate labeled Escherichia coli (MTCC-729) to rat proximal renal tubular cells (NRK-52E cells). Further, the antioxidant property of cystone was studied using hydrogen peroxide (400 μM) as a pro-oxidant in NRK-52E cells. The results showed that cystone inhibited the adherence of E. coli to NRK-52E cells significantly. Additionally cystone effectively combats the toxicity induced by H2O2 in NRK-52E cells. The cytoprotective effect of cystone is brought about by inhibiting lipid peroxidation by 36% in cells treated with cystone compared to H2O2-treated cells without cystone. The antioxidant enzymes catalase, glutathione were increased by 53% and 68% respectively and superoxide dismutase activity was increased 3-fold. The glutathione content was significantly increased by 2.4-fold in NRK-52E cells treated with cystone compared to H2O2 control group. These results suggest that cystone effectively inhibits bacterial adherence to NRK-52E cells and attenuates H2O2-induced toxicity in NRK-52E cells by inhibiting lipid peroxidation and increasing the antioxidant defense mechanism. PMID:27186087

  17. Adherence Reduction of Campylobacter jejuni and Campylobacter coli Strains to HEp-2 Cells by Mannan Oligosaccharides and a High-Molecular-Weight Component of Cranberry Extract.

    PubMed

    Ramirez-Hernandez, Alejandra; Rupnow, John; Hutkins, Robert W

    2015-08-01

    Campylobacter infections are a leading cause of human bacterial gastroenteritis in the United States and are a major cause of diarrheal disease throughout the world. Colonization and subsequent infection and invasion of Campylobacter require that the bacteria adhere to the surface of host cells. Agents that inhibit adherence could be used prophylactically to reduce Campylobacter carriage and infection. Mannan oligosaccharides (MOS) have been used as a feed supplement in livestock animals to improve performance and to replace growth-promoting antibiotics. However, MOS and other nondigestible oligosaccharides may also prevent pathogen colonization by inhibiting adherence in the gastrointestinal tract. In addition, plant extracts, including those derived from cranberries, have been shown to have antiadherence activity against pathogens. The goal of this study was to assess the ability of MOS and cranberry fractions to serve as antiadherence agents against strains of Campylobacter jejuni and Campylobacter coli. Adherence experiments were performed using HEp-2 cells. Significant reductions in adherence of C. jejuni 29438, C. jejuni 700819, C. jejuni 3329, and C. coli 43485 were observed in the presence of MOS (up to 40 mg/ml) and with a high-molecular-weight fraction of cranberry extract (up to 3 mg/ml). However, none of the tested materials reduced adherence of C. coli BAA-1061. No additive effect in adherence inhibition was observed for an MOS-cranberry blend. These results suggest that both components, MOS and cranberry, could be used to reduce Campylobacter colonization and carriage in livestock animals and potentially limit human exposure to this pathogen. PMID:26219363

  18. Response of adherent cells to mechanical perturbations of the surrounding matrix.

    PubMed

    Ben-Yaakov, Dan; Golkov, Roman; Shokef, Yair; Safran, Samuel A

    2015-02-01

    We present a generic and unified theory to explain how cells respond to perturbations of their mechanical environment such as the presence of neighboring cells, slowly applied stretch, or gradients of matrix rigidity. Motivated by experiments, we calculate the local balance of forces that give rise to a tendency for the cell to locally move or reorient, with a focus on the contribution of feedback and homeostasis to cell contractility (manifested by a fixed displacement, strain or stress) that acts on the adhesions at the cell boundary. These forces can be either reinforced or diminished by elastic stresses due to mechanical perturbations of the matrix. Our model predicts these changes and how their balance with local protrusive forces that act on the cell's leading edge either increase or decrease the tendency of the cell to locally move (toward neighboring cells or rigidity gradients) or reorient (in the direction of slowly applied stretch or rigidity gradients). PMID:25604950

  19. Comparative genomic hybridization analysis of newly established retinoblastoma cell lines of adherent growth compared with Y79 of nonadherent growth.

    PubMed

    Kim, Jeong Hun; Kim, Jin Hyoung; Yu, Young Suk; Kim, Dong Hun; Kim, Yong Kyu; Kim, Kyu-Won

    2008-08-01

    Retinoblastoma (RB) shows cytogenetic aberrations involving genes other than RB gene located on 13q14. We analyzed genomic aberration in newly established RB cell lines SNUOT-RB1 and SNUOT-RB4 of adherent growth and Y79 cell line of nonadherent growth by microarray comparative genomic hybridization. SNUOT-RB1 showed 44 significant copy number changes (gain in 11 and loss in 33, P<0.0005). SNUOT-RB4 showed 42 significant copy number changes (gain in 8 and loss in 34, P<0.0005). Y79 cell line had the greatest gain of 19.65-fold in the locus of MYCN gene 2p24.1, whereas SNUOT-RB1 and SNUOT-RB4 showed no significant gain. SNUOT-RB1 and SNUOT-RB4 gained chromosomal copy numbers commonly in chromosome 11, especially in locus 11q13, which is responsible for cancer-related genes such as CCND1, MEN1, and FGF3. Losses of copy numbers occurred in chromosomes 3, 9, 10, 11, 16, and 17. In summary, SNUOT-RB1 and SNUOT-RB4 represented similar pattern in gain and loss of chromosomal copy number changes, while different from Y79. The loss of CYLD gene of tumor suppressor gene, 16q12-q13, was only on locus of common involvement in 3 cell lines. PMID:18799932

  20. Measurement of annexin V uptake and lactadherin labeling for the quantification of apoptosis in adherent Tca8113 and ACC-2 cells.

    PubMed

    Hu, T; Shi, J; Jiao, X; Zhou, J; Yin, X

    2008-09-01

    Phosphatidylserine (PS) exposure occurs during the cell death program and fluorescein-labeled lactadherin permits the detection of PS exposure earlier than annexin V in suspended cell lines. Adherent cell lines were studied for this apoptosis-associated phenomenon to determine if PS probing methods are reliable because specific membrane damage may occur during harvesting. Apoptosis was induced in the human tongue squamous carcinoma cell line (Tca8113) and the adenoid cystic carcinoma cell line (ACC-2) by arsenic trioxide. Cells were harvested with a modified procedure and labeled with lactadherin and/or annexin V. PS exposure was localized by confocal microscopy and apoptosis was quantified by flow cytometry. The detachment procedure without trypsinization did not induce cell damage. In competition binding experiments, phospholipid vesicles competed for more than 95 and 90% of lactadherin but only about 75 and 70% of annexin V binding to Tca8113 and ACC-2 cells. These data indicate that PS exposure occurs in three stages during the cell death program and that fluorescein-labeled lactadherin permitted the detection of early PS exposure. A similar pattern of PS exposure has been observed in two malignant cell lines with different adherence, suggesting that this pattern of PS exposure is common in adherent cells. Both lactadherin and annexin V could be used in adherent Tca8113 and ACC-2 cell lines when an appropriate harvesting procedure was used. Lactadherin is more sensitive than annexin V for the detection of PS exposure as the physical structure of PS in these blebs and condensed apoptotic cell surface may be more conducive to binding lactadherin than annexin V. PMID:18820763

  1. Improving physician's adherence to completing vaccination schedules for patients with type 2 diabetes attending non-communicable diseases clinics in West Bay Health Center, Qatar

    PubMed Central

    Tawfik, Hassan; Bashwar, Zelaikha; Al-Ali, Amal; Salem, Mohamed; Abdelbagi, Isameldin

    2015-01-01

    Incomplete vaccination for patients with type 2 diabetes attending non-communicable diseases (NCD) clinics is an issue that could affect patient's health and wellness negatively and puts patients at high risk of serious diseases. We aimed to improve physicians adherence to complete vaccination schedule for patients with type 2 diabetes attending NCD clinics in west bay health center according to American Diabetes Association (ADA) recommendation by 25% by January 2015. In the pre-intervention phase: the quality improvement team designed a checklist to collect the percentage of physician's adherence of prescription of the recommended vaccination for patients with type 2 diabetes. The percentage of complete vaccination in patients with diabetes attending NCD clinic in West Bay Health Center was 20% . In the intervention phase the intervention was in the form of: the creation a vaccination form and attached to the (NCD) progress note; to distribute and remind the physicians about the ADA guidelines vaccination recommendations; a summary of the vaccination schedule developed and attached to (NCD) form; development of vaccination reminder posters and posters in the waiting area, nurse station, and physician clinics and education and orientation sessions for NCD clinic staff. In the post-intervention phase the average percentage of complete vaccination in patients with diabetes attending NCD clinic in West Bay Health Center increased to 69%. PMID:26732463

  2. Rapidly Self-Renewing Human Multipotent Marrow Stromal Cells (hMSC) Express Sialyl Lewis X and Actively Adhere to Arterial Endothelium in a Chick Embryo Model System

    PubMed Central

    McFerrin, Harris E.; Olson, Scott D.; Gutschow, Miriam V.; Semon, Julie A.; Sullivan, Deborah E.; Prockop, Darwin J.

    2014-01-01

    Background There have been conflicting observations regarding the receptors utilized by human multipotent mesenchymal bone marrow stromal cells (hMSC) to adhere to endothelial cells (EC). To address the discrepancies, we performed experiments with cells prepared with a standardized, low-density protocol preserving a sub-population of small cells that are rapidly self-renewing. Methods Sialyl Lewis X (SLeX) and α4 integrin expression were determined by flow cytometry. Fucosyltransferase expression was determined by quantitative realtime RT-PCR. Cell adhesion assays were carried out with a panel of endothelial cells from arteries, veins and the microvasculature in vitro. In vivo experiments were performed to determine single cell interactions in the chick embryo chorioallantoic membrane (CAM). The CAM is a well-characterized respiratory organ allowing for time-lapse image acquisition of large numbers of cells treated with blocking antibodies against adhesion molecules expressed on hMSC. Results hMSC expressed α4 integrin, SLeX and fucosyltransferase 4 and adhered to human EC from arteries, veins and the microvasculature under static conditions in vitro. In vivo, hMSC rolled on and adhered to arterioles in the chick embryo CAM, whereas control melanoma cells embolized. Inhibition of α4 integrin and/or SLeX with blocking antibodies reduced rolling and adhesion in arterioles and increased embolism of hMSC. Conclusions The results demonstrated that rapidly self-renewing hMSC were retained in the CAM because they rolled on and adhered to respiratory arteriolar EC in an α4 integrin- and SLeX-dependent manner. It is therefore important to select cells based on their cell adhesion receptor profile as well as size depending on the intended target of the cell and the injection route. PMID:25144321

  3. Human Placenta-Derived Adherent Cells Prevent Bone loss, Stimulate Bone formation, and Suppress Growth of Multiple Myeloma in Bone

    PubMed Central

    Li, Xin; Ling, Wen; Pennisi, Angela; Wang, Yuping; Khan, Sharmin; Heidaran, Mohammad; Pal, Ajai; Zhang, Xiaokui; He, Shuyang; Zeitlin, Andy; Abbot, Stewart; Faleck, Herbert; Hariri, Robert; Shaughnessy, John D.; van Rhee, Frits; Nair, Bijay; Barlogie, Bart; Epstein, Joshua; Yaccoby, Shmuel

    2011-01-01

    Human placenta has emerged as a valuable source of transplantable cells of mesenchymal and hematopoietic origin for multiple cytotherapeutic purposes, including enhanced engraftment of hematopoietic stem cells, modulation of inflammation, bone repair, and cancer. Placenta-derived adherent cells (PDACs) are mesenchymal-like stem cells isolated from postpartum human placenta. Multiple myeloma is closely associated with induction of bone disease and large lytic lesions, which are often not repaired and are usually the sites of relapses. We evaluated the antimyeloma therapeutic potential, in vivo survival, and trafficking of PDACs in the severe combined immunodeficiency (SCID)–rab model of medullary myeloma-associated bone loss. Intrabone injection of PDACs into non-myelomatous and myelomatous implanted bone in SCID-rab mice promoted bone formation by stimulating endogenous osteoblastogenesis, and most PDACs disappeared from bone within 4 weeks. PDACs inhibitory effects on myeloma bone disease and tumor growth were dose-dependent and comparable with those of fetal human mesenchymal stem cells (MSCs). Intrabone, but not subcutaneous, engraftment of PDACs inhibited bone disease and tumor growth in SCID-rab mice. Intratumor injection of PDACs had no effect on subcutaneous growth of myeloma cells. A small number of intravenously injected PDACs trafficked into myelomatous bone. Myeloma cell growth rate in vitro was lower in coculture with PDACs than with MSCs from human fetal bone or myeloma patients. PDACs also promoted apoptosis in osteoclast precursors and inhibited their differentiation. This study suggests that altering the bone marrow microenvironment with PDAC cytotherapy attenuates growth of myeloma and that PDAC cytotherapy is a promising therapeutic approach for myeloma osteolysis. PMID:21732484

  4. Adherence to Phosphodiesterase Type 5 Inhibitors in the Treatment of Erectile Dysfunction in Long-Term Users: How Do Men Use the Inhibitors?

    PubMed Central

    Carvalheira, Ana; Forjaz, Vera; Pereira, Nuno Monteiro

    2014-01-01

    Introduction The high effectiveness of phosphodiesterase type 5 inhibitors (PDE5-i) in the treatment of erectile dysfunction (ED) has been demonstrated. However, previous research shows that PDE5-i treatments have high discontinuation rates. Aim The main goals of this study were to (i) characterize the way men use PDE5-i and (ii) analyze the adherence to treatment, identifying the factors that influence PDE5-i use. Methods A total of 148 men with clinical diagnosis for ED who maintained the treatment with PDE5-i for over 3 years were interviewed. Interviews concerning their ongoing treatment were carried out using a standardized questionnaire with quantitative and qualitative items. Main Outcome Measures Physiological measures included the intracavernous alprostadil injection test, associated with penile rigidometry and penile Doppler ultrasound. The qualitative measure included two questions: “Do you use the drug in every sexual intercourse?” and “How do you use the inhibitor?” Results ED causes were classified as venogenic (31%), arteriogenic (23%), psychogenic (18%), iatrogenic (13%), neurogenic (8%), and diabetic (7%). Participation rate was 71.8%. Of the 148 patients studied, 75% claimed not to use PDE5-i in every intercourse. Most used tadalafil (66%), followed by sildenafil (20%), vardenafil (10%), and 4% alternated the type of medicine. Four main categories emerged concerning the factors that determine the intake of PDE5-i in some intercourse situations and not in others: (i) psychological factors; (ii) medication-related factors; (iii) circumstantial factors; and (iv) relational factors. Conclusion The analysis of men's narratives revealed a combination of factors that influence the adherence to PDE5-i. The psychological and medication-related factors were the most prevalent. This study highlighted the importance of taking these factors into account, both at the time of prescription and during the follow-up in order to improve adherence

  5. Vertical nanopillars for in situ probing of nuclear mechanics in adherent cells.

    PubMed

    Hanson, Lindsey; Zhao, Wenting; Lou, Hsin-Ya; Lin, Ziliang Carter; Lee, Seok Woo; Chowdary, Praveen; Cui, Yi; Cui, Bianxiao

    2015-06-01

    The mechanical stability and deformability of the cell nucleus are crucial to many biological processes, including migration, proliferation and polarization. In vivo, the cell nucleus is frequently subjected to deformation on a variety of length and time scales, but current techniques for studying nuclear mechanics do not provide access to subnuclear deformation in live functioning cells. Here we introduce arrays of vertical nanopillars as a new method for the in situ study of nuclear deformability and the mechanical coupling between the cell membrane and the nucleus in live cells. Our measurements show that nanopillar-induced nuclear deformation is determined by nuclear stiffness, as well as opposing effects from actin and intermediate filaments. Furthermore, the depth, width and curvature of nuclear deformation can be controlled by varying the geometry of the nanopillar array. Overall, vertical nanopillar arrays constitute a novel approach for non-invasive, subcellular perturbation of nuclear mechanics and mechanotransduction in live cells. PMID:25984833

  6. Vertical nanopillars for in situ probing of nuclear mechanics in adherent cells

    NASA Astrophysics Data System (ADS)

    Hanson, Lindsey; Zhao, Wenting; Lou, Hsin-Ya; Lin, Ziliang Carter; Lee, Seok Woo; Chowdary, Praveen; Cui, Yi; Cui, Bianxiao

    2015-06-01

    The mechanical stability and deformability of the cell nucleus are crucial to many biological processes, including migration, proliferation and polarization. In vivo, the cell nucleus is frequently subjected to deformation on a variety of length and time scales, but current techniques for studying nuclear mechanics do not provide access to subnuclear deformation in live functioning cells. Here we introduce arrays of vertical nanopillars as a new method for the in situ study of nuclear deformability and the mechanical coupling between the cell membrane and the nucleus in live cells. Our measurements show that nanopillar-induced nuclear deformation is determined by nuclear stiffness, as well as opposing effects from actin and intermediate filaments. Furthermore, the depth, width and curvature of nuclear deformation can be controlled by varying the geometry of the nanopillar array. Overall, vertical nanopillar arrays constitute a novel approach for non-invasive, subcellular perturbation of nuclear mechanics and mechanotransduction in live cells.

  7. AFBI assay – Aptamer Fluorescence Binding and Internalization assay for cultured adherent cells

    PubMed Central

    Thiel, William H.; Giangrande, Paloma H.

    2016-01-01

    The SELEX (Systematic Evolution of Ligands by Exponential Enrichment) process allows for the enrichment of DNA or RNA aptamers from a complex nucleic acid library that are specific for a target molecule. The SELEX process has been adapted from identifying aptamers in vitro using recombinant target protein to cell-based methodologies (Cell-SELEX), where the targets are expressed on the surface of cells. One major advantage of Cell-SELEX is that the target molecules are maintained in a native confirmation. Additionally, Cell-SELEX may be used to discover novel therapeutic biomarkers by performing selections on diseased versus healthy cells. However, a caveat to Cell-SELEX is that testing of single aptamers identified in the selection is laborious, time-consuming, and expensive. The most frequently used methods to screen for aptamer binding and internalization on cells are flow cytometry and quantitative PCR (qPCR). While flow cytometry can directly assess binding of a fluorescently-labeled aptamer to a target, it requires significant starting material and is not easily scalable. qPCR-based approaches are highly sensitive but have non-negligible experiment-to-experiment variability due to the number of sample processing steps. Herein we describe a cell-based aptamer fluorescence binding and internalization (AFBI) assay. This assay requires minimal reagents and has few experimental steps/manipulations, thereby allowing for rapid screening of many aptamers and conditions simultaneously and direct quantitation of aptamer binding and internalization. PMID:26972784

  8. A functional assay for gap junctional examination; electroporation of adherent cells on indium-tin oxide.

    PubMed

    Geletu, Mulu; Guy, Stephanie; Firth, Kevin; Raptis, Leda

    2014-01-01

    In this technique, cells are cultured on a glass slide that is partly coated with indium-tin oxide (ITO), a transparent, electrically conductive material. A variety of molecules, such as peptides or oligonucleotides can be introduced into essentially 100% of the cells in a non-traumatic manner. Here, we describe how it can be used to study intercellular, gap junctional communication. Lucifer yellow penetrates into the cells when an electric pulse, applied to the conductive surface on which they are growing, causes pores to form through the cell membrane. This is electroporation. Cells growing on the nonconductive glass surface immediately adjacent to the electroporated region do not take up Lucifer yellow by electroporation but do acquire the fluorescent dye as it is passed to them via gap junctions that link them to the electroporated cells. The results of the transfer of dye from cell to cell can be observed microscopically under fluorescence illumination. This technique allows for precise quantitation of gap junctional communication. In addition, it can be used for the introduction of peptides or other non-permeant molecules, and the transfer of small electroporated peptides via gap junctions to inhibit the signal in the adjacent, non-electroporated cells is a powerful demonstration of signal inhibition. PMID:25350637

  9. AFBI assay - Aptamer Fluorescence Binding and Internalization assay for cultured adherent cells.

    PubMed

    Thiel, William H; Giangrande, Paloma H

    2016-07-01

    The SELEX (Systematic Evolution of Ligands by Exponential Enrichment) process allows for the enrichment of DNA or RNA aptamers from a complex nucleic acid library that are specific for a target molecule. The SELEX process has been adapted from identifying aptamers in vitro using recombinant target protein to cell-based methodologies (Cell-SELEX), where the targets are expressed on the surface of cells. One major advantage of Cell-SELEX is that the target molecules are maintained in a native confirmation. Additionally, Cell-SELEX may be used to discover novel therapeutic biomarkers by performing selections on diseased versus healthy cells. However, a caveat to Cell-SELEX is that testing of single aptamers identified in the selection is laborious, time-consuming, and expensive. The most frequently used methods to screen for aptamer binding and internalization on cells are flow cytometry and quantitative PCR (qPCR). While flow cytometry can directly assess binding of a fluorescently-labeled aptamer to a target, it requires significant starting material and is not easily scalable. qPCR-based approaches are highly sensitive but have non-negligible experiment-to-experiment variability due to the number of sample processing steps. Herein we describe a cell-based aptamer fluorescence binding and internalization (AFBI) assay. This assay requires minimal reagents and has few experimental steps/manipulations, thereby allowing for rapid screening of many aptamers and conditions simultaneously and direct quantitation of aptamer binding and internalization. PMID:26972784

  10. Antibody against the Carboxyl Terminus of Intimin α Reduces Enteropathogenic Escherichia coli Adherence to Tissue Culture Cells and Subsequent Induction of Actin Polymerization

    PubMed Central

    Carvalho, Humberto M.; Teel, Louise D.; Kokai-Kun, John F.; O'Brien, Alison D.

    2005-01-01

    The C-terminal third of intimin binds to its translocated receptor (Tir) to promote attaching and effacing lesion formation during infection with enteropathogenic Escherichia coli (EPEC). We observed that the adherence of EPEC strains to HEp-2 cells was reduced and that actin polymerization was blocked by antibody raised against the C-terminal third of intimin α. PMID:15784601

  11. Adherence to Mediterranean-style dietary pattern and risk of esophageal squamous cell carcinoma: a case-control study in Iran

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The benefit of adherence to a Mediterranean-style dietary pattern in relation to the risk of esophageal squamous cell carcinoma (ESCC) has not been investigated among non-Mediterranean high-risk populations. The objective of the present study was to examine the association of compliance with the Med...

  12. Exopolysaccharides of Lactobacillus reuteri: Their influence on adherence of E. coli to epithelial cells and inflammatory response.

    PubMed

    Kšonžeková, Petra; Bystrický, Peter; Vlčková, Silvia; Pätoprstý, Vladimír; Pulzová, Lucia; Mudroňová, Dagmar; Kubašková, Terézia; Csank, Tomáš; Tkáčiková, Ľudmila

    2016-05-01

    The aim of the study was to characterize exopolysaccharides (EPS) originated from Lactobacillus reuteri strain DSM 17938 (EPS-DSM17938) and L. reuteri strain L26 Biocenol™ (EPS-L26) and evaluate their influence on adherence of enterotoxigenic Escherichia coli (ETEC) to IPEC-1 cells and proinflammatory gene expression. Both EPS were d-glucan polysaccharides with higher molecular weight (Mw), but differing in spatial conformation and elicited variable cytokine profile. EPS-DSM17938, relatively linear polysaccharide with (1→4) and (1→6) glycosidic linkages, increased IL-1β gene expression (0.1mg/mL; P<0.05), while EPS-L26, more branched polysaccharide with (1→3) and (1→6) glycosidic linkages, exerted slight but statistically significant up-regulation of NF-κB, TNF-α and IL-6 mRNA (P<0.05). The most significant finding is that preincubation of IPEC-1 cells with both EPS followed by ETEC infection inhibit ETEC adhesion on IPEC-1 cells (P<0.01) and ETEC-induced gene expression of proinflammatory cytokine IL-1β and IL-6 (P<0.01). PMID:26876991

  13. Further evidence for the existence of 'homing' receptors on murine leukemia cells which mediate adherence to normal bone marrow stromal cells.

    PubMed

    Kamenov, B; Longenecker, B M

    1985-01-01

    A significant proportion of 131IUDR-labelled cells from murine leukemia cell lines L1210 and P388, but not the L5178Y lymphoma cell line, are retained in the bone marrow (B.M.) following i.v. injection into syngeneic mice. Following this, L1210 and P388 cells grow and rapidly replace the normal hematopoietic cells of the B.M. L1210 and P388 cells, but not several lymphoma cell lines, also bind avidly to monolayers of B.M. stromal cells (Dexter cultures) and soon overgrow the cultures following rapid cell proliferation. P388 cells bound equally well to confluent monolayers of B.M., whole mouse embryo and newborn mouse kidney while L1210 cells bound well to B.M. and whole mouse embryo but showed little binding to newborn kidney monolayers. The accumulation of the two leukemia cell lines in the B.M. was constant and indistinguishable over a 48-h period. In contrast, in both spleen and liver the number of L1210 cells decreased during the same period while P388 cells were retained at a constant level. Generally there was a lack of correlation of B.M. metastasis of a cell line and its metastasis to other organs although P388 cells, but not L1210 cells, demonstrated a tremendous capacity for metastatic growth in both spleen and liver. Normal B.M. cells were fused with the syngeneic SP2/0 murine myeloma fusor line and 10 hybridomas plus the SP2/0 parent were tested for in-vitro adherence to B.M. monolayers and in-vivo metastatic behavior. The same 3 (out of 10) hybridomas showed a high level of adherence to B.M. monolayers, high levels of retention of cells in the B.M. following i.v. injection, and rapid growth and takeover of the normal B.M. In marked contrast, neither the SP2/0 parent nor the remaining 7 hybridomas show significant adherence, B.M. retention or growth in the B.M. A distinct lack of correlation of B.M. vs liver or spleen metastasis was once again noted for the hybridomas although all of the hybridomas showed much less metastatic growth in the liver than

  14. Radial mobility and cytotoxic function of retroviral replicating vector transduced, non-adherent alloresponsive T lymphocytes.

    PubMed

    Erickson, Kate L; Hickey, Michelle J; Kato, Yuki; Malone, Colin C; Owens, Geoffrey C; Prins, Robert M; Liau, Linda M; Kasahara, Noriyuki; Kruse, Carol A

    2015-01-01

    We report a novel adaptation of the Radial Monolayer Cell Migration assay, first reported to measure the radial migration of adherent tumor cells on extracellular matrix proteins, for measuring the motility of fluorescently-labeled, non-adherent human or murine effector immune cells. This technique employs a stainless steel manifold and 10-well Teflon slide to focally deposit non-adherent T cells into wells prepared with either confluent tumor cell monolayers or extracellular matrix proteins. Light and/or multi-channel fluorescence microscopy is used to track the movement and behavior of the effector cells over time. Fluorescent dyes and/or viral vectors that code for fluorescent transgenes are used to differentially label the cell types for imaging. This method is distinct from similar-type in vitro assays that track horizontal or vertical migration/invasion utilizing slide chambers, agar or transwell plates. The assay allows detailed imaging data to be collected with different cell types distinguished by specific fluorescent markers; even specific subpopulations of cells (i.e., transduced/nontransduced) can be monitored. Surface intensity fluorescence plots are generated using specific fluorescence channels that correspond to the migrating cell type. This allows for better visualization of the non-adherent immune cell mobility at specific times. It is possible to gather evidence of other effector cell functions, such as cytotoxicity or transfer of viral vectors from effector to target cells, as well. Thus, the method allows researchers to microscopically document cell-to-cell interactions of differentially-labeled, non-adherent with adherent cells of various types. Such information may be especially relevant in the assessment of biologically-manipulated or activated immune cell types, where visual proof of functionality is desired with tumor target cells before their use for cancer therapy. PMID:25741775

  15. Production of the Escherichia coli Common Pilus by Uropathogenic E. coli Is Associated with Adherence to HeLa and HTB-4 Cells and Invasion of Mouse Bladder Urothelium

    PubMed Central

    Carrillo-Casas, Erika Margarita; Durán, Laura; Zhang, Yushan; Hernández-Castro, Rigoberto; Puente, José L.; Daaka, Yehia; Girón, Jorge A.

    2014-01-01

    Uropathogenic Escherichia coli (UPEC) strains cause urinary tract infections and employ type 1 and P pili in colonization of the bladder and kidney, respectively. Most intestinal and extra-intestinal E. coli strains produce a pilus called E. coli common pilus (ECP) involved in cell adherence and biofilm formation. However, the contribution of ECP to the interaction of UPEC with uroepithelial cells remains to be elucidated. Here, we report that prototypic UPEC strains CFT073 and F11 mutated in the major pilin structural gene ecpA are significantly deficient in adherence to cultured HeLa (cervix) and HTB-4 (bladder) epithelial cells in vitro as compared to their parental strains. Complementation of the ecpA mutant restored adherence to wild-type levels. UPEC strains produce ECP upon growth in Luria-Bertani broth or DMEM tissue culture medium preferentially at 26°C, during incubation with cultured epithelial cells in vitro at 37°C, and upon colonization of mouse bladder urothelium ex vivo. ECP was demonstrated on and inside exfoliated bladder epithelial cells present in the urine of urinary tract infection patients. The ability of the CFT073 ecpA mutant to invade the mouse tissue was significantly reduced. The presence of ECP correlated with the architecture of the biofilms produced by UPEC strains on inert surfaces. These data suggest that ECP can potentially be produced in the bladder environment and contribute to the adhesive and invasive capabilities of UPEC during its interaction with the host bladder. We propose that along with other known adhesins, ECP plays a synergistic role in the multi-step infection of the urinary tract. PMID:25036370

  16. Escherichia coli O157:H7 Strains That Persist in Feedlot Cattle Are Genetically Related and Demonstrate an Enhanced Ability To Adhere to Intestinal Epithelial Cells

    PubMed Central

    Carlson, Brandon A.; Nightingale, Kendra K.; Mason, Gary L.; Ruby, John R.; Choat, W. Travis; Loneragan, Guy H.; Smith, Gary C.; Sofos, John N.; Belk, Keith E.

    2009-01-01

    A longitudinal study was conducted to investigate the nature of Escherichia coli O157:H7 colonization of feedlot cattle over the final 100 to 110 days of finishing. Rectal fecal grab samples were collected from an initial sample population of 788 steers every 20 to 22 days and microbiologically analyzed to detect E. coli O157:H7. The identities of presumptive colonies were confirmed using a multiplex PCR assay that screened for gene fragments unique to E. coli O157:H7 (rfbE and fliCh7) and other key virulence genes (eae, stx1, and stx2). Animals were classified as having persistent shedding (PS), transient shedding (TS), or nonshedding (NS) status if they consecutively shed the same E. coli O157:H7 genotype (based on the multiplex PCR profile), exhibited variable E. coli O157 shedding, or never shed morphologically typical E. coli O157, respectively. Overall, 1.0% and 1.4% of steers were classified as PS and NS animals, respectively. Characterization of 132 E. coli O157:H7 isolates from PS and TS animals by pulsed-field gel electrophoresis (PFGE) typing yielded 32 unique PFGE types. One predominant PFGE type accounted for 53% of all isolates characterized and persisted in cattle throughout the study. Isolates belonging to this predominant and persistent PFGE type demonstrated an enhanced (P < 0.0001) ability to adhere to Caco-2 human intestinal epithelial cells compared to isolates belonging to less common PFGE types but exhibited equal virulence expression. Interestingly, the attachment efficacy decreased as the genetic divergence from the predominant and persistent subtype increased. Our data support the hypothesis that certain E. coli O157:H7 strains persist in feedlot cattle, which may be partially explained by an enhanced ability to colonize the intestinal epithelium. PMID:19617387

  17. Extracellular mass transport considerations for space flight research concerning suspended and adherent in vitro cell cultures

    NASA Technical Reports Server (NTRS)

    Klaus, David M.; Benoit, Michael R.; Nelson, Emily S.; Hammond, Timmothy G.

    2004-01-01

    Conducting biological research in space requires consideration be given to isolating appropriate control parameters. For in vitro cell cultures, numerous environmental factors can adversely affect data interpretation. A biological response attributed to microgravity can, in theory, be explicitly correlated to a specific lack of weight or gravity-driven motion occurring to, within or around a cell. Weight can be broken down to include the formation of hydrostatic gradients, structural load (stress) or physical deformation (strain). Gravitationally induced motion within or near individual cells in a fluid includes sedimentation (or buoyancy) of the cell and associated shear forces, displacement of cytoskeleton or organelles, and factors associated with intra- or extracellular mass transport. Finally, and of particular importance for cell culture experiments, the collective effects of gravity must be considered for the overall system consisting of the cells, their environment and the device in which they are contained. This does not, however, rule out other confounding variables such as launch acceleration, on orbit vibration, transient acceleration impulses or radiation, which can be isolated using onboard centrifuges or vibration isolation techniques. A framework is offered for characterizing specific cause-and-effect relationships for gravity-dependent responses as a function of the above parameters.

  18. A Method to Evaluate the Efficiency of Transfection Reagents in an Adherent Zebrafish Cell Line

    PubMed Central

    Aschberger, Teresa; Pelster, Bernd

    2013-01-01

    Abstract We present a simple and robust method to evaluate the transfection efficiency of commercially available transfection reagents intended to be established for use in nonmammalian cell lines. To illustrate the method, we compare the ability of four different reagents to transfect the embryonic zebrafish cell line Z3. Z3 cells were seeded in a 96-well plate and simultaneously transfected in several variations by using minimum volumes of transfection reagent and a vector DNA encoding an amplified version of green fluorescent protein (GFP). After 24 and 48 h, transfection efficiency was determined by a dual fluorescence plate reader measurement of GFP and Hoechst 33342 fluorescence, an indicator of cell density. Of the four different reagents tested, certain variations of JetPrime™ reagent and X-tremeGene™ HP reagent produced the highest fluorescence signal per cell after 24- and 48-h incubation, respectively. The simultaneous multivariate setup enables comparing different reagent/DNA combinations at different time points well, independent of cell growth variability or seeding density. PMID:23515475

  19. Milk digesta and milk protein fractions influence the adherence of Lactobacillus gasseri R and Lactobacillus casei FMP to human cultured cells.

    PubMed

    Volstatova, Tereza; Havlik, Jaroslav; Potuckova, Miroslava; Geigerova, Martina

    2016-08-10

    Adhesion to the intestinal epithelium is considered an important feature of probiotic bacteria, which may increase their persistence in the intestine, allowing them to exert their beneficial health effect or promote the colonisation process. However, this feature might be largely dependent on the host specificity or diet. In the present study, we investigated the effect of selected milks and milk protein fractions on the ability of selected lactobacilli to adhere to the cells of an intestinal model based on co-culture Caco-2/HT29-MTX cell lines. Most milk digesta did not significantly affect bacterial adhesion except for UHT-treated milk and sheep milk. The presence of UHT-treated milk digesta reduced the adhesion of Lactobacillus gasseri R by 61% but not that of Lactobacillus casei FMP. However, sheep milk significantly increased the adherence of L. casei FMP (P < 0.05) but not of L. gasseri R. Among the protein fractions, rennet casein (RCN) and bovine serum albumin (BSA) showed reproducible patterns and strain-specific effects on bacterial adherence. While RCN reduced the adherence of L. gasseri R to <50% compared to the control, it did not have a significant effect on L. casei FMP. In contrast, BSA reduced L. casei FMP adherence to a higher extent than that of L. gasseri R. Whey protein (WH) tended to increase the adherence of both strains by 130%-180%. Recently, interactions between the host diet and its microbiota have attracted considerable interest. Our results may explain one of the aspects of the role of milk in the development of microbiota or support of probiotic supplements. Based on our data, we conclude that the persistence of probiotic strains supplemented as part of dairy food or constitutional microbiota in the gut might be affected negatively or positively by the food matrix through complex strain or concentration dependent effects. PMID:27435508

  20. Study protocol for a randomized controlled trial to assess the feasibility of an open label intervention to improve hydroxyurea adherence in youth with sickle cell disease

    PubMed Central

    Smaldone, Arlene; Findley, Sally; Bakken, Suzanne; Matiz, L. Adriana; Rosenthal, Susan L.; Jia, Haomiao; Matos, Sergio; Manwani, Deepa; Green, Nancy S.

    2016-01-01

    Background Community health workers (CHW) are increasingly recognized as a strategy to improve health outcomes for the underserved with chronic diseases but has not been formally explored in adolescents with sickle cell disease (SCD). SCD primarily affects African American, Hispanic and other traditionally underserved populations. Hydroxyurea (HU), an oral, once-daily medication, is the only approved therapeutic drug for sickle cell disease and markedly reduces symptoms, morbidity and mortality and improves quality of life largely by increasing hemoglobin F blood levels. This paper presents the rationale, study design and protocol for an open label randomized controlled trial to improve parent-youth partnerships in self-management and medication adherence to HU in adolescents with SCD. Methods/Design A CHW intervention augmented by text messaging was designed for adolescents with SCD ages 10–18 years and their parents to improve daily HU adherence. Thirty adolescent parent dyads will be randomized with 2:1 intervention group allocation. Intervention dyads will establish a relationship with a culturally aligned CHW to identify barriers to HU use, identify cues to build a habit, and develop a dyad partnership to improve daily HU adherence and achieve their individualized “personal best” hemoglobin F target. Intervention feasibility, acceptability and efficacy will be assessed via a 2-site trial. Outcomes of interest are HU adherence, dyad self-management communication, quality of life, and resource use. Discussion Despite known benefits, poor HU adherence is common. If feasible and acceptable, the proposed intervention may improve health of underserved adolescents with SCD by enhancing long-term HU adherence. PMID:27327779

  1. Determinants of medication adherence among adults with type 2 diabetes mellitus in three Malaysian public health clinics: a cross-sectional study

    PubMed Central

    Chew, Boon-How; Hassan, Noor-Hasliza; Sherina, Mohd-Sidik

    2015-01-01

    Medication adherence (MA) in adults with type 2 diabetes mellitus (T2D) is associated with improved disease control (glycated hemoglobin, blood pressure, and lipid profile), lower rates of death and diabetes-related complications, increased quality of life, and decreased health care resource utilization. However, there is a paucity of data on the effect of diabetes-related distress, depression, and health-related quality of life on MA. This study examined factors associated with MA in adults with T2D at the primary care level. This was a cross-sectional study conducted in three Malaysian public health clinics, where adults with T2D were recruited consecutively in 2013. We used the 8-item Morisky Medication Adherence Scale (MMAS-8) to assess MA as the main dependent variable. In addition to sociodemographic data, we included diabetes-related distress, depressive symptoms, and health-related quality of life as independent variables. Independent association between the MMAS-8 score and its determinants was done using generalized linear models with a gamma distribution and log link function. The participant response rate was 93.1% (700/752). The majority were female (52.8%), Malay (52.9%), and married (79.1%). About 43% of patients were classified as showing low MA (MMAS-8 score <6). Higher income (adjusted odds ratio 0.90) and depressive symptoms (adjusted odds ratio 0.99) were significant independent determinants of medication non-adherence in young adults with T2D. Low MA in adults with T2D is a prevalent problem. Thus, primary health care providers in public health clinics should focus on MA counselling for adult T2D patients who are younger, have a higher income, and symptoms of depression. PMID:25999699

  2. Cell-Type-Specific Optogenetics in Monkeys.

    PubMed

    Namboodiri, Vijay Mohan K; Stuber, Garret D

    2016-09-01

    The recent advent of technologies enabling cell-type-specific recording and manipulation of neuronal activity spurred tremendous progress in neuroscience. However, they have been largely limited to mice, which lack the richness in behavior of primates. Stauffer et al. now present a generalizable method for achieving cell-type specificity in monkeys. PMID:27610562

  3. Immunosuppression associated with the development of chronic infections with Rickettsia tsutsugamushi: adherent suppressor cell activity and macrophage activation.

    PubMed Central

    Jerrells, T R

    1985-01-01

    Measures of general immunocompetency such as lymphocyte responses to mitogens and alloantigens and the ability to produce antibody to T-dependent and T-independent antigens were evaluated during the development of chronic infections with Rickettsia tsutsugamushi resulting from subcutaneous infection of BALB/c mice. It was found that a transient immunosuppression was demonstrable regardless of the infecting strain of rickettsiae; however, the immunosuppression produced by the Karp and Kato strains was more pronounced and longer lived. As a marked splenomegaly resulting from inflammatory macrophage influx accompanied this immunosuppression, mitogen- and antigen-induced lymphocyte proliferation was also evaluated after adherent cell depletion or in the presence of indomethacin, and both treatments significantly improved the responses. Isolated splenic macrophages were shown to suppress the responses of lymphocytes from naive mice as well as to exhibit parameters of activation including tumor cell cytolysis and cytostasis and the ability to inhibit the replication of R. tsutsugamushi in vitro. These data suggest an association between macrophage activation involved in rickettsial clearance and a transient immunosuppression. PMID:2931378

  4. Type II pneumocytes in mixed cell culture of human lung: a light and electron microscopic study.

    PubMed Central

    Bingle, L; Bull, T B; Fox, B; Guz, A; Richards, R J; Tetley, T D

    1990-01-01

    Alveolar Type II epithelial cells dedifferentiate rapidly in vitro. Studies with animal tissue suggest that cell-cell and extracellular matrix-cell interactions are important in the retention of Type II cell morphology in vitro. Thus, in this study with human tissue, alveolar Type II cells, alveolar macrophages, and spindle cells were prepared from the same sample of lung (obtained following lobectomy for cancer, n = 3), cocultured on glass cover slips or tissue culture plastic, and studied by light microscopy with scanning (SEM) and transmission (TEM) electron microscopy for 8 days. The primary cell isolates contained approximately 45% Type II cells; the remainder were macrophages or unidentifiable cells. Clusters, made up of a single layer of cuboidal Type II cells around a central core of connective tissue (largely collagen and some elastic tissue), formed above a monolayer of spindle cells. The Type II cells were morphologically similar to those seen in vivo. The cells were still cuboidal at 8 days but had lost their lamellar bodies, which were released into the medium via the apical surface. The clusters increased in size with time (area, microns 2: day 1, 29(5-143) x 10(2); day 8, 63(10-311) x 10(2); mean(range); p less than 0.02) without changing in number per culture, suggesting Type II cell proliferation. This may have been due to factors produced by the other cells and adherence to the extracellular matrix (ECM); (free collagen fibers, present in the original preparation, spindle cells, and/or Type II cells could be responsible for presence of ECM). We propose this as a useful model for the study of human Type II epithelial cells in vitro. Images FIGURE 1. a FIGURE 1. b FIGURE 1. c FIGURE 1. d FIGURE 1. e FIGURE 1. f FIGURE 2. a FIGURE 2. b FIGURE 2. c FIGURE 2. d FIGURE 2. e FIGURE 2. f FIGURE 2. g FIGURE 3. PMID:2384069

  5. Listeria monocytogenes listeriolysin O and phosphatidylinositol-specific phospholipase C affect adherence to epithelial cells.

    PubMed

    Krawczyk-Balska, Agata; Bielecki, Jacek

    2005-09-01

    Listeria monocytogenes, a foodborn intracellular animal and human pathogen, produces several exotoxins contributing to virulence. Among these are listeriolysin O (LLO), a pore-forming cholesterol-dependent hemolysin, and a phosphatidylinositol-specific phospholipase C (PI-PLC). LLO is known to play an important role in the escape of bacteria from the primary phagocytic vacuole of macrophages, and PI-PLC supports this process. Evidence is accumulating that LLO and PI-PLC are multifunctional virulence factors with many important roles in the host-parasite interaction other than phagosomal membrane disruption. LLO and PI-PLC may induce a number of host cell responses by modulating signal transduction of infected cells via intracellular Ca2+ levels and the metabolism of phospholipids. This would result in the activation of host phospholipase C and protein kinase C. In the present study, using Bacillus sub tilis strains expressing LLO, PI-PLC, and simultaneously LLO and PI-PLC, we show that LLO and PI-PLC enhance bacterial binding to epithelial cells Int407, with LLO being necessary and PI-PLC playing an accessory role. The results of this work suggest that these two listerial proteins act on epithelial cells prior to internalization. PMID:16391652

  6. Roles for Cell Wall Glycopeptidolipid in Surface Adherence and Planktonic Dispersal of Mycobacterium avium

    EPA Science Inventory

    The opportunistic pathogen Mycobacterium avium is a significant inhabitant of biofilms in drinking water distribution systems. M. avium expresses on its cell surface serovar-specific glycopeptidolipids (ssGPLs). Studies have implicated the core GPL in biofilm formation by M. aviu...

  7. Anti-adherence potential of Enterococcus durans cells and its cell-free supernatant on plastic and stainless steel against foodborne pathogens.

    PubMed

    Amel, Ait Meddour; Farida, Bendali; Djamila, Sadoun

    2015-07-01

    It is demonstrated that numerous bacteria are able to attach to surfaces of equipment used for food handling or processing. In this study, a strain of Enterococcus durans, originally isolated from a milking machine surface, was firstly studied for its biofilm formation potential on plastic and stainless steel supports. The strain was found to be a biofilm producer either at 25, 30 or 37 °C on polystyrene microtitre plates, with a best adherence level observed at 25 °C. En. durans showed a strong adhesion to stainless steel AISI-304. Antibacterial and anti-adherence activities of En. durans were tested against four foodborne pathogens (Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 27853 and Listeria innocua CLIP 74915) which were shown as biofilm producers on both plastic and stainless steel. En. durans cells and cell-free culture supernatant showed a significant (P < 0.05) inhibition potential of the pathogens either on solid media or in broth co-cultures. Characterization of the antibacterial substances indicated their proteinaceous nature which assigned them most probably to bacteriocins group. PMID:25466409

  8. Factors influencing human leukocyte adherence in vitro.

    PubMed

    Stepniewicz, W; Tchórzewski, H; Luciak, M

    1983-01-01

    Studies were performed on factors influencing leucocyte adherence in vitro. Blood condensation was found to increase leukocyte adherence. Addition of heparin, dextran or ethanol caused a significant reduction of white blood cell count in blood samples in comparison with blood mixed with sodium EDTA or ACD solution. This suggests the existence of two granulocyte subpopulations; viz, rapidly adhering and slowly adhering. Heparin enhanced granulocyte adherence, while dextran and ethanol decreased it. Five-day storage of ACD blood led to a decrease in granulocyte adherence, while addition of heparin or histamine to ACD blood prevented this change to occur. The glucose concentration of 1,000 mg/dl augmented granulocyte adherence, while higher glucose concentrations induced its progressive fall below the control values. There was no significant change of lymphocyte adherence during the experiments. PMID:6194070

  9. Monolithic cascade-type solar cells

    NASA Technical Reports Server (NTRS)

    Yamamoto, S.; Shibukawa, A.; Yamaguchi, M.

    1985-01-01

    Solar cells consist of a semiconductor base, a bottom cell with a band-gap energy of E1, and a top cell with a band-gap energy of E2, and 0.96 E1 1.36 eV and (0.80 E + 0.77) eV E2 (0.80 E1 + 0.92) eV. A monolithic cascade-type solar cell was prepared with an n(+)-type GaAs base, a GaInAs bottom solar cell, and a GaAiInAs top solar cell. The surface of the cell is coated with a SiO antireflection film. The efficiency of the cell is 32%.

  10. Reduction of Adherence of E. coli O157:H7 to HEp-2 Cells and to Bovine Large Intestinal Mucosal Explants by Colicinogenic E. coli

    PubMed Central

    Etcheverría, A. I.; Arroyo, G. H.; Alzola, R.; Parma, A. E.

    2011-01-01

    Enterohemorrhagic E. coli strains (EHEC) had emerged as foodborne pathogens and cause in human diarrhea and hemolytic-uremic syndrome. Because of the widespread distribution of EHEC serotypes and O157 and non-O157 in cattle population, its control will require interventions at the farm level such as the administration of probiotics that produce inhibitory metabolites. E. coli O157:H7 shows tissue tropisms for the gastrointestinal tract (GIT) of cattle. The aim of this study was to test the ability of a colicinogenic E. coli (isolated from bovine) to reduce the adherence of E. coli O157:H7 to HEp-2 cells and to GIT of cattle. We inoculated HEp-2 cells and bovine colon explants with both kinds of strains. Colicinogenic E. coli was able to reduce the adherence of E. coli O157:H7 to HEp-2 cells and to bovine tissues. PMID:23724308

  11. Modeling and visualizing cell type switching.

    PubMed

    Ghaffarizadeh, Ahmadreza; Podgorski, Gregory J; Flann, Nicholas S

    2014-01-01

    Understanding cellular differentiation is critical in explaining development and for taming diseases such as cancer. Differentiation is conventionally represented using bifurcating lineage trees. However, these lineage trees cannot readily capture or quantify all the types of transitions now known to occur between cell types, including transdifferentiation or differentiation off standard paths. This work introduces a new analysis and visualization technique that is capable of representing all possible transitions between cell states compactly, quantitatively, and intuitively. This method considers the regulatory network of transcription factors that control cell type determination and then performs an analysis of network dynamics to identify stable expression profiles and the potential cell types that they represent. A visualization tool called CellDiff3D creates an intuitive three-dimensional graph that shows the overall direction and probability of transitions between all pairs of cell types within a lineage. In this study, the influence of gene expression noise and mutational changes during myeloid cell differentiation are presented as a demonstration of the CellDiff3D technique, a new approach to quantify and envision all possible cell state transitions in any lineage network. PMID:24834107

  12. Surface-Mediated Stimuli Responsive Delivery of Organic Molecules from Porous Carriers to Adhered Cells.

    PubMed

    Ergün, Bahar; De Cola, Luisa; Galla, Hans-Joachim; Kehr, Nermin Seda

    2016-07-01

    The alternating layer-by-layer deposition of oppositely charged polyelectrolytes on fluorescence-dye-(Hst)-loaded zeolites L ((Hst) Zeo-PSS/PLL) is described. The arrays and nanocomposite (NC) hydrogels of (Hst) Zeo-PSS/PLL are prepared. The subsequent cell experiments show the potential application of arrays and NC hydrogels of (Hst) Zeo-PSS/PLL as alternative 2D- and 3D-surfaces, respectively, for 2D- and 3D-surface-mediated controlled organic molecules delivery applications. PMID:27114067

  13. Functional and phenotypic characterization of a protein from Lactobacillus acidophilus involved in cell morphology, stress tolerance and adherence to intestinal cells.

    PubMed

    O'Flaherty, Sarah J; Klaenhammer, Todd R

    2010-11-01

    Structural components of the cell surface have an impact on some of the beneficial attributes of probiotic bacteria. In silico analysis of the L. acidophilus NCFM genome sequence revealed the presence of a putative cell surface protein that was predicted to be a myosin cross-reactive antigen (MCRA). As MCRAs are conserved among many probiotic bacteria, we used the upp-based counterselective gene replacement system, designed recently for use in L. acidophilus, to determine the functional role of this gene (LBA649) in L. acidophilus NCFM. Phenotypic assays were undertaken with the parent strain (NCK1909) and deletion mutant (NCK2015) to assign a function for this gene. The growth of NCK2015 (ΔLBA649) was reduced in the presence of lactate, acetate, porcine bile and salt. Adhesion of NCK2015 to Caco-2 cells was substantially reduced for both stationary-phase (∼45 % reduction) and exponential-phase cells (∼50 % reduction). Analysis of NCK2015 by scanning electron microscopy revealed a longer cell morphology after growth in MRS broth compared to NCK1909. These results indicate a role for LBA649 in stress tolerance, cell wall division and adherence to Caco-2 cells. PMID:20829293

  14. The comparison of the effect of endodontic irrigation on cell adherence to root canal dentin.

    PubMed

    Ring, Karla C; Murray, Peter E; Namerow, Kenneth N; Kuttler, Sergio; Garcia-Godoy, Franklin

    2008-12-01

    The purpose of this study was to compare the effect of 10 different endodontic irrigation and chelating treatments on dental pulp stem cell (DPSC) attachment to root canal surfaces. Thirty-eight extracted human nondiseased single-canal teeth were cleaned and shaped using ProTaper and ProFile rotary instrumentation (Tulsa Dentsply, Tulsa, OK). The irrigation treatments investigated were 6% sodium hypochlorite, 2% chlorhexidine gluconate, Aquatine Endodontic Cleanser, and Morinda citrifolia juice. The irrigation treatments were used in conjunction with EDTA or MTAD. The instrumented teeth were immediately placed in cell culture with confluent DPSCs for 1 week. The number of attached DPSCs appeared to be correlated with the cytotoxicity of the root canal irrigating solution (analysis of variance, p < 0.0001). The presence or absence of the smear layer had little influence on DPSC activity (chi-square, p > 0.05). The results suggest that biocompatible irrigants are needed to promote DPSC attachment to root canal dentin, which is essential to accomplish some regenerative endodontic therapies. PMID:19026877

  15. Cell Wall-Associated Protein Antigens of Streptococcus salivarius: Purification, Properties, and Function in Adherence

    PubMed Central

    Weerkamp, Anton H.; Jacobs, Ton

    1982-01-01

    Three cell wall-associated protein antigens (antigens b, c, and d) were isolated from mutanolysin-solubilized cell walls of Streptococcus salivarius HB and purified to apparent homogeneity by a combination of ion-exchange chromatography, gel filtration, and immunoadsorption chromatography. Antigens b and c were also isolated from culture supernatants. Antigen b consisted of more than 80% protein and had an apparent molecular weight as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of 320,000. Antigen c consisted of 57% protein, about 30% neutral sugar, and about 13% amino sugar, and its glycoprotein nature was confirmed by specific staining techniques. During sodium dodecyl sulfate-polyacrylamide gel electrophoresis antigen c resolved into two or more bands, depending on the source or the isolation procedure, in the molecular weight range from 220,000 to 280,000. Antigen d consisted of 95% protein and was observed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis as two bands with molecular weights of 129,000 and 121,000. Under nondenaturing conditions all three antigens had molecular weights in the range from 1 × 106 to 3 × 106 as determined by gel filtration. The amino acid compositions of antigens b, c, and d were characterized by low amounts of basic amino acids and relatively high levels of nonpolar amino acids. Among oral streptococcal species antigens b and c were virtually restricted to strains of S. salivarius and most often to serotype I strains. Antigen b was recognized as the factor that mediates coaggregation of S. salivarius with Veillonella strains. The purified protein retained its biological activity. Antigen c could be linked to functions relating to adhesion of the streptococci to host tissues on the basis of its absence in mutant strains and blocking by specific antisera. The purified molecule had no detectable biological activity. Antigen d could not be linked to an established adhesion function. Images

  16. Adherence of non-O157 Shiga-toxin Escherichia coli to bovine recto-anal junction squamous epithelial cells appears to be mediated by mechanisms distinct from those used by O157

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study presents evidence that the pattern of adherence of clinically relevant non-O157 Shiga-toxin producing Escherichia coli (STEC) to bovine recto-anal junction squamous epithelial cells (RSE) is similar to that of O157, although the mechanisms of adherence appear to be distinct. Our results f...

  17. Surfactant protein D inhibits adherence of uropathogenic Escherichia coli to the bladder epithelial cells and the bacterium-induced cytotoxicity: a possible function in urinary tract.

    PubMed

    Kurimura, Yuichiro; Nishitani, Chiaki; Ariki, Shigeru; Saito, Atsushi; Hasegawa, Yoshihiro; Takahashi, Motoko; Hashimoto, Jiro; Takahashi, Satoshi; Tsukamoto, Taiji; Kuroki, Yoshio

    2012-11-16

    The adherence of uropathogenic Escherichia coli (UPEC) to the host urothelial surface is the first step for establishing UPEC infection. Uroplakin Ia (UPIa), a glycoprotein expressed on bladder urothelium, serves as a receptor for FimH, a lectin located at bacterial pili, and their interaction initiates UPEC infection. Surfactant protein D (SP-D) is known to be expressed on mucosal surfaces in various tissues besides the lung. However, the functions of SP-D in the non-pulmonary tissues are poorly understood. The purposes of this study were to investigate the possible function of SP-D expressed in the bladder urothelium and the mechanisms by which SP-D functions. SP-D was expressed in human bladder mucosa, and its mRNA was increased in the bladder of the UPEC infection model in mice. SP-D directly bound to UPEC and strongly agglutinated them in a Ca(2+)-dependent manner. Co-incubation of SP-D with UPEC decreased the bacterial adherence to 5637 cells, the human bladder cell line, and the UPEC-induced cytotoxicity. In addition, preincubation of SP-D with 5637 cells resulted in the decreased adherence of UPEC to the cells and in a reduced number of cells injured by UPEC. SP-D directly bound to UPIa and competed with FimH for UPIa binding. Consistent with the in vitro data, the exogenous administration of SP-D inhibited UPEC adherence to the bladder and dampened UPEC-induced inflammation in mice. These results support the conclusion that SP-D can protect the bladder urothelium against UPEC infection and suggest a possible function of SP-D in urinary tract. PMID:23012359

  18. Anti-Retroviral Lectins Have Modest Effects on Adherence of Trichomonas vaginalis to Epithelial Cells In Vitro and on Recovery of Tritrichomonas foetus in a Mouse Vaginal Model

    PubMed Central

    Chatterjee, Aparajita; Ratner, Daniel M.; Ryan, Christopher M.; Johnson, Patricia J.; O’Keefe, Barry R.; Secor, W. Evan; Anderson, Deborah J.; Robbins, Phillips W.; Samuelson, John

    2015-01-01

    Trichomonas vaginalis causes vaginitis and increases the risk of HIV transmission by heterosexual sex, while Tritrichomonas foetus causes premature abortion in cattle. Our goals were to determine the effects, if any, of anti-retroviral lectins, which are designed to prevent heterosexual transmission of HIV, on adherence of Trichomonas to ectocervical cells and on Tritrichomonas infections in a mouse model. We show that Trichomonas Asn-linked glycans (N-glycans), like those of HIV, bind the mannose-binding lectin (MBL) that is part of the innate immune system. N-glycans of Trichomonas and Tritrichomonas bind anti-retroviral lectins (cyanovirin-N and griffithsin) and the 2G12 monoclonal antibody, each of which binds HIV N-glycans. Binding of cyanovirin-N appears to be independent of susceptibility to metronidazole, the major drug used to treat Trichomonas. Anti-retroviral lectins, MBL, and galectin-1 cause Trichomonas to self-aggregate and precipitate. The anti-retroviral lectins also increase adherence of ricin-resistant mutants, which are less adherent than parent cells, to ectocervical cell monolayers and to organotypic EpiVaginal tissue cells. Topical application of either anti-retroviral lectins or yeast N-glycans decreases by 40 to 70% the recovery of Tritrichomonas from the mouse vagina. These results, which are explained by a few simple models, suggest that the anti-retroviral lectins have a modest potential for preventing or treating human infections with Trichomonas. PMID:26252012

  19. SALT-OVERLY SENSITIVE5 Mediates Arabidopsis Seed Coat Mucilage Adherence and Organization through Pectins.

    PubMed

    Griffiths, Jonathan S; Tsai, Allen Yi-Lun; Xue, Hui; Voiniciuc, Cătălin; Sola, Krešimir; Seifert, Georg J; Mansfield, Shawn D; Haughn, George W

    2014-05-01

    Interactions between cell wall polymers are critical for establishing cell wall integrity and cell-cell adhesion. Here, we exploit the Arabidopsis (Arabidopsis thaliana) seed coat mucilage system to examine cell wall polymer interactions. On hydration, seeds release an adherent mucilage layer strongly attached to the seed in addition to a nonadherent layer that can be removed by gentle agitation. Rhamnogalacturonan I (RG I) is the primary component of adherent mucilage, with homogalacturonan, cellulose, and xyloglucan constituting minor components. Adherent mucilage contains rays composed of cellulose and pectin that extend above the center of each epidermal cell. CELLULOSE SYNTHASE5 (CESA5) and the arabinogalactan protein SALT-OVERLY SENSITIVE5 (SOS5) are required for mucilage adherence through unknown mechanisms. SOS5 has been suggested to mediate adherence by influencing cellulose biosynthesis. We, therefore, investigated the relationship between SOS5 and CESA5. cesa5-1 seeds show reduced cellulose, RG I, and ray size in adherent mucilage. In contrast, sos5-2 seeds have wild-type levels of cellulose but completely lack adherent RG I and rays. Thus, relative to each other, cesa5-1 has a greater effect on cellulose, whereas sos5-2 mainly affects pectin. The double mutant cesa5-1 sos5-2 has a much more severe loss of mucilage adherence, suggesting that SOS5 and CESA5 function independently. Double-mutant analyses with mutations in MUCILAGE MODIFIED2 and FLYING SAUCER1 that reduce mucilage release through pectin modification suggest that only SOS5 influences pectin-mediated adherence. Together, these findings suggest that SOS5 mediates adherence through pectins and does so independently of but in concert with cellulose synthesized by CESA5. PMID:24808103

  20. The Use of Cell Phone Support for Non-adherent HIV-Infected Youth and Young Adults: An Initial Randomized and Controlled Intervention Trial

    PubMed Central

    Belzer, Marvin E.; Naar-King, Sylvie; Olson, Johanna; Sarr, Moussa; Thornton, Sarah; Kahana, Shoshana Y.; Gaur, Aditya H.; Clark, Leslie F.

    2014-01-01

    This randomized behavioral trial examined whether youth living with HIV (YLH) receiving cell-phone support with study funded phone plans, demonstrated improved adherence and viral control during the 24 week intervention and 24 weeks post-intervention compared to controls. Monday through Friday phone calls confirmed medications were taken, provided problem-solving support, and referred to services to address adherence barriers. Of 37 participants (ages 15–24), 62 % were male and 70 % were African American. Self-reported adherence was significantly higher in the intervention group compared to the control at 24 and 48 weeks for the past month (P = 0.007) and log 10 HIV VL was significantly lower at both 24 weeks (2.82 versus 4.52 P = 0.002) and 48 weeks (3.23 versus 4.23 P = 0.043). Adherence and viral load showed medium to large effect sizes across the 48 week study. This is the first study to demonstrate sustained clinically significant reductions in HIV VL using youth friendly technology. PMID:24271347

  1. Adherence of streptococcus pyogenes, Escherichia coli, and Pseudomonas aeruginosa to fibronectin-coated and uncoated epithelial cells.

    PubMed Central

    Abraham, S N; Beachey, E H; Simpson, W A

    1983-01-01

    The relationship between the variability in the fibronectin (Fn) content on human buccal epithelial cells and the capacity of the cells to bind gram-positive (Streptococcus pyogenes) or gram-negative (Escherichia coli or Pseudomonas aeruginosa) bacteria was investigated. Adhesion experiments performed with mixtures of epithelial cells and mixed suspensions of either S. pyogenes and E. coli or S. pyogenes and P. aeruginosa exhibited three major populations of buccal cells: one of these was able to bind S. pyogenes (gram positive) but neither of the gram-negative bacteria; a second population was able to bind the gram-negative but not the gram-positive bacteria; and a third was able to bind various numbers of both types of organisms. Further adhesion experiments performed with a mixture of epithelial cells and a mixed suspension of S. pyrogens, E. coli, and fluoresceinconjugated methacrylate beads coated with immune immunoglobulin G directed against Fn revealed that the epithelial cells recognizing the gram-positive bacteria were rich in Fn, whereas those recognizing the gram-negative organisms were poor in Fn. Immunoelectron microscopy confirmed that cells of S. pyogenes bound to epithelial cells coated with Fn, whereas cells of E. coli bound to epithelial cells lacking Fn. These results suggest that Fn on the surfaces of epithelial cells may modulate the ecology of the human oropharyngeal cavity, especially with respect to the colonization of these surfaces by pathogenic gram-negative or gram-positive bacteria. Images PMID:6411621

  2. Phenotypic analysis of nylon-wool-adherent suppressor cells that inhibit the effector process of tumour cell lysis by lymphokine-activated killer cells in patients with advanced gastric carcinoma.

    PubMed

    Koyama, S; Fukao, K

    1994-01-01

    The causes of down-regulation of cytotoxic immune responses in cancer patients have not been fully evaluated. We previously demonstrated that T-cell-growth-factor-activated peripheral blood lymphocytes (PBL) with the surface phenotype CD8+ CD11b-, from patients with widespread metastasis of gastric carcinoma, inhibited the effector process of lymphokine-activated-killer(LAK)-cell-mediated cytolysis. In this study, we examined suppressor cell activity in freshly prepared PBL from 18 patients with advanced gastric carcinoma, and 10 normal healthy individuals. The suppressor cell activity was assayed by recording whether or not PBL inhibited directly the effector process of LAK cell cytotoxicity. Most of the PBL suspensions from cancer patients showed that they contained a population of cells that can directly inhibit the effector phase of tumor cell lysis of the cytotoxic cells. To analyze further the PBL responsible for the suppression, the cells were passed over a nylon-wool column. Nylon-wool-adherent cells significantly augmented the suppression, while the cells passing through abrogated the suppressive effect. Most nylon-wool-adherent cells from 10 normal healthy controls did not inhibit the cytotoxic reaction. To determine further the suppressor-effector population in nylon-wool-adherent cells, negative-selection studies using CD8-, CD4- or CD11b-coated magnetic beads, and positive-selection studies using CD8- or CD4-coated magnetic beads were performed. Finally the results suggest that the suppressor-effector cells comprise at least two different surface phenotypes: CD8+ T and CD8-CD11b+ cells. The possible role of CD4+ T cells and HLA-DR+ LeuM3+ macrophages as suppressor cells was ruled out in nylon-wool-adherent cells. CD8+ T and possibly CD8-CD11b+ cells apparently suppressed the efferent limb of the antitumor immunity. The selective immune suppression mediated by these cells may partly be concerned with escape mechanisms of gastric carcinoma from the host

  3. Turning One Cell Type into Another.

    PubMed

    Slack, Jonathan M W

    2016-01-01

    The nature of cells in early embryos may be respecified simply by exposure to inducing factors. In later stage embryos, determined cell populations do not respond to inducing factors but may be respecified by other stimuli, especially the introduction of specific transcription factors. Fully differentiated cell types are hard to respecify by any method, but some degree of success can be achieved using selected combinations of transcription factors, and this may have clinical significance in the future. PMID:26969988

  4. Birth weight and later life adherence to unhealthy lifestyles in predicting type 2 diabetes: prospective cohort study

    PubMed Central

    Ley, Sylvia H; Tobias, Deirdre K; Chiuve, Stephanie E; VanderWeele, Tyler J; Rich-Edwards, Janet W; Curhan, Gary C; Willett, Walter C; Manson, JoAnn E; Hu, Frank B

    2015-01-01

    Objectives To prospectively assess the joint association of birth weight and established lifestyle risk factors in adulthood with incident type 2 diabetes and to quantitatively decompose the attributing effects to birth weight only, to adulthood lifestyle only, and to their interaction. Design Prospective cohort study. Setting Health Professionals Follow-up Study (1986-2010), Nurses’ Health Study (1980-2010), and Nurses’ Health Study II (1991-2011). Participants 149 794 men and women without diabetes, cardiovascular disease, or cancer at baseline. Main outcome measure Incident cases of type 2 diabetes, identified through self report and validated by a supplementary questionnaire. Unhealthy lifestyle was defined on the basis of body mass index, smoking, physical activity, alcohol consumption, and the alternate healthy eating index. Results During 20-30 years of follow-up, 11 709 new cases of type 2 diabetes were documented. The multivariate adjusted relative risk of type 2 diabetes was 1.45 (95% confidence interval 1.32 to 1.59) per kg lower birth weight and 2.10 (1.71 to 2.58) per unhealthy lifestyle factor. The relative risk of type 2 diabetes associated with a combination of per kg lower birth weight and per unhealthy lifestyle factor was 2.86 (2.26 to 3.63), which was more than the addition of the risk associated with each individual factor, indicating a significant interaction on an additive scale (P for interaction<0.001). The attributable proportions of joint effect were 22% (95% confidence interval 18.3% to 26.4%) to lower birth weight alone, 59% (57.1% to 61.5%) to unhealthy lifestyle alone, and 18% (13.9% to 21.3%) to their interaction. Conclusion Most cases of type 2 diabetes could be prevented by the adoption of a healthier lifestyle, but simultaneous improvement of both prenatal and postnatal factors could further prevent additional cases. PMID:26199273

  5. Biology of alveolar type II cells.

    PubMed

    Mason, Robert J

    2006-01-01

    The purpose of this review is to highlight the many metabolic properties of alveolar type II cells, their production of surfactant, their role in innate immunity, and their importance in the repair process after lung injury. The review is based on the medical literature and results from our laboratory. Type II cells produce and secrete pulmonary surfactant and for that purpose they need to synthesize the lipids of surfactant. One of the regulators of lipogenesis is the transcription factor sterol regulatory element binding protein-1c (SREBP-1c). This is a key transcription factor regulating fatty acid synthesis. Type II cells also proliferate to restore the epithelium after lung injury, clear alveolar fluid by transporting sodium from the apical to the basolateral surface, and participate in the innate immune response to inhaled materials and organisms. The type II cell is, in many ways, the defender of the alveolus. However, the type II cells work in concert with the other cells in the gas exchange regions of the lung to keep the alveoli open and reduce inflammation due to irritants in the air we breathe. PMID:16423262

  6. The cells of the dorsal iris involved in lens regeneration are myoepithelial cells whose cytoskeleton changes during cell type conversion.

    PubMed

    Yang, Y; Zalik, S E

    1994-06-01

    During newt lens regeneration, the pigmented epithelial cells (PECs) of the dorsal iris dedifferentiate and give rise to a new lens. We have studied the cytoskeleton of the PECs using iris flat mounts and sections. In flat-mount iris preparations stained by labelled phalloidin three main regions can be recognized: the pupillary (P) ring, the middle (M) ring, and the more external junctional (J) ring. The cells of the P ring that give rise to the lens have an elongated spindle shape and exhibit an elaborate cytoskeleton of actin filament bundles oriented along the long axis of the cells, reminiscent of myoepithelial or smooth muscle cells. These cells express smooth muscle-specific alpha actin, muscle gamma actin and cytokeratin II, and adhere to each other through the cell adhesion molecule A-CAM. During dedifferentiation, actin staining increases considerably as the actin filament bundles thicken and shorten and then accumulate preferentially in the apical and basel regions of the elongating lens fibres. Cytokeratin II, which is also organized as fibrils along the long axis of the normal iris PECs, increases progressively during dedifferentiation, when it is organized as a thick band surrounding the nucleus. The expression of this protein is repressed during lens fibre differentiation, but is retained in mitotic cells. The data suggest that during cell type conversion some cytoskeletal proteins increase and reorganize, while others disappear during lens fibre differentiation. PMID:7526744

  7. Manipulation of intestinal epithelial cell function by the cell contact-dependent type III secretion systems of Vibrio parahaemolyticus.

    PubMed

    O'Boyle, Nicky; Boyd, Aoife

    2014-01-01

    Vibrio parahaemolyticus elicits gastroenteritis by deploying Type III Secretion Systems (TTSS) to deliver effector proteins into epithelial cells of the human intestinal tract. The bacteria must adhere to the human cells to allow colonization and operation of the TTSS translocation apparatus bridging the bacterium and the host cell. This article first reviews recent advances in identifying the molecules responsible for intercellular adherence. V. parahaemolyticus possesses two TTSS, each of which delivers an exclusive set of effectors and mediates unique effects on the host cell. TTSS effectors primarily target and alter the activation status of host cell signaling proteins, thereby bringing about changes in the regulation of cellular behavior. TTSS1 is responsible for the cytotoxicity of V. parahaemolyticus, while TTSS2 is necessary for the enterotoxicity of the pathogen. Recent publications have elucidated the function of several TTSS effectors and their importance in the virulence of the bacterium. This review will explore the ability of the TTSS to manipulate activities of human intestinal cells and how this modification of cell function favors bacterial colonization and persistence of V. parahaemolyticus in the host. PMID:24455490

  8. Attachment role of gonococcal pili. Optimum conditions and quantitation of adherence of isolated pili to human cells in vitro.

    PubMed Central

    Pearce, W A; Buchanan, T M

    1978-01-01

    Gonoccocal pili facilitate attachment of virulent Neisseria gonorrhoeae to human cells. To characterize this attachment function, purified gonococcal pili isolated from four strains possessing antigenically distinct pili were radiolabeled with 125I and used to measure the attachment of pili to various human cells in vitro. Human buccal and cervical-vaginal mucosal epithealial cells, fallopian tube mucosa, and sperm bound pili in greater numbers per micrometer2 of surface area (1--10) than fetal tonsil fibroblasts, HeLa M cells, erythrocytes, or polymorphonuclear leukocytes. This cell specificity of attachment suggests a greater density of membrane pili binding sites on cells similar or identical to cells from natural sites of infection. The pili binding sites were quantitated as 1 X 10(4) per cervical-vaginal squamous cell. Pili of all antigenic types attached equally to a given cell type, implying that the attachment moiety of each pilus was similar. Attachement of gonoccocal pili to human cells occurred quickly with saturation of presumed receptor sites within 20--60 min. Attachment was temperature dependent (37 degrees greater than 20 degrees greater than 4 degrees C), and pH dependent (3.5 less than 4.5 less than 5.5 less than 7.5). Attachment was inhibited by antibody to pili (homologous pili Ab greater than heterologous Ab). The extent of possible protection against gonococcal infection due to inhibition of pili-mediated attachment might prove limited as a result of the considerable antigenic heterogeneity among pili and the observation that blockage of pili attachment is maximal only with antibody to pili of the infecting strain. Images PMID:96134

  9. Evaluation of 309 environmental chemicals using a mouse embryonic stem cell adherent cell differentiation and cytotoxicity assay.

    PubMed

    Chandler, Kelly J; Barrier, Marianne; Jeffay, Susan; Nichols, Harriette P; Kleinstreuer, Nicole C; Singh, Amar V; Reif, David M; Sipes, Nisha S; Judson, Richard S; Dix, David J; Kavlock, Robert; Hunter, Edward S; Knudsen, Thomas B

    2011-01-01

    The vast landscape of environmental chemicals has motivated the need for alternative methods to traditional whole-animal bioassays in toxicity testing. Embryonic stem (ES) cells provide an in vitro model of embryonic development and an alternative method for assessing developmental toxicity. Here, we evaluated 309 environmental chemicals, mostly food-use pesticides, from the ToxCast™ chemical library using a mouse ES cell platform. ES cells were cultured in the absence of pluripotency factors to promote spontaneous differentiation and in the presence of DMSO-solubilized chemicals at different concentrations to test the effects of exposure on differentiation and cytotoxicity. Cardiomyocyte differentiation (α,β myosin heavy chain; MYH6/MYH7) and cytotoxicity (DRAQ5™/Sapphire700™) were measured by In-Cell Western™ analysis. Half-maximal activity concentration (AC₅₀) values for differentiation and cytotoxicity endpoints were determined, with 18% of the chemical library showing significant activity on either endpoint. Mining these effects against the ToxCast Phase I assays (∼500) revealed significant associations for a subset of chemicals (26) that perturbed transcription-based activities and impaired ES cell differentiation. Increased transcriptional activity of several critical developmental genes including BMPR2, PAX6 and OCT1 were strongly associated with decreased ES cell differentiation. Multiple genes involved in reactive oxygen species signaling pathways (NRF2, ABCG2, GSTA2, HIF1A) were strongly associated with decreased ES cell differentiation as well. A multivariate model built from these data revealed alterations in ABCG2 transporter was a strong predictor of impaired ES cell differentiation. Taken together, these results provide an initial characterization of metabolic and regulatory pathways by which some environmental chemicals may act to disrupt ES cell growth and differentiation. PMID:21666745

  10. Evaluation of 309 Environmental Chemicals Using a Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity Assay

    PubMed Central

    Chandler, Kelly J.; Barrier, Marianne; Jeffay, Susan; Nichols, Harriette P.; Kleinstreuer, Nicole C.; Singh, Amar V.; Reif, David M.; Sipes, Nisha S.; Judson, Richard S.; Dix, David J.; Kavlock, Robert; Hunter, Edward S.; Knudsen, Thomas B.

    2011-01-01

    The vast landscape of environmental chemicals has motivated the need for alternative methods to traditional whole-animal bioassays in toxicity testing. Embryonic stem (ES) cells provide an in vitro model of embryonic development and an alternative method for assessing developmental toxicity. Here, we evaluated 309 environmental chemicals, mostly food-use pesticides, from the ToxCast™ chemical library using a mouse ES cell platform. ES cells were cultured in the absence of pluripotency factors to promote spontaneous differentiation and in the presence of DMSO-solubilized chemicals at different concentrations to test the effects of exposure on differentiation and cytotoxicity. Cardiomyocyte differentiation (α,β myosin heavy chain; MYH6/MYH7) and cytotoxicity (DRAQ5™/Sapphire700™) were measured by In-Cell Western™ analysis. Half-maximal activity concentration (AC50) values for differentiation and cytotoxicity endpoints were determined, with 18% of the chemical library showing significant activity on either endpoint. Mining these effects against the ToxCast Phase I assays (∼500) revealed significant associations for a subset of chemicals (26) that perturbed transcription-based activities and impaired ES cell differentiation. Increased transcriptional activity of several critical developmental genes including BMPR2, PAX6 and OCT1 were strongly associated with decreased ES cell differentiation. Multiple genes involved in reactive oxygen species signaling pathways (NRF2, ABCG2, GSTA2, HIF1A) were strongly associated with decreased ES cell differentiation as well. A multivariate model built from these data revealed alterations in ABCG2 transporter was a strong predictor of impaired ES cell differentiation. Taken together, these results provide an initial characterization of metabolic and regulatory pathways by which some environmental chemicals may act to disrupt ES cell growth and differentiation. PMID:21666745

  11. Increased protein secretion and adherence to HeLa cells by Shigella spp. following growth in the presence of bile salts.

    PubMed Central

    Pope, L M; Reed, K E; Payne, S M

    1995-01-01

    Growth of Shigella spp. in the presence of the bile salt deoxycholate or chenodeoxycholate enhanced the bacterial invasion of HeLa cells. Growth in the presence of other structurally similar bile salts or detergents had little or no effect. Deoxycholate-enhanced invasion was not observed when bacteria were exposed to deoxycholate at low temperatures or when chloramphenicol was added to the growth medium, indicating that bacterial growth and protein synthesis are required. Increased invasion is associated with the presence of an intact Shigella virulence plasmid and is correlated with increased secretion of a set of proteins, including the Ipa proteins, to the outer membrane and into the growth medium. The increased invasion induced by the bile salts appears to be due to increased adherence. The enhanced adherence was specific to Shigella spp., since the enteroinvasive Escherichia coli strains tested did not exhibit the effect in response to growth in bile salts. PMID:7642302

  12. Proper Regulation of Cdc42 Activity is Required for Tight Actin Concentration at the Equator during Cytokinesis in Adherent Mammalian Cells

    PubMed Central

    Zhu, Xiaodong; Wang, Junxia; Moriguchi, Kazuki; Liow, Lu Ting; Ahmed, Sohail; Kaverina, Irina; Murata-Hori, Maki

    2012-01-01

    Cytokinesis in mammalian cells requires actin assembly at the equatorial region. Although functions of RhoA in this process have been well established, additional mechanisms are likely involved. We have examined if Cdc42 is involved in actin assembly during cytokinesis. Depletion of Cdc42 had no apparent effects on the duration of cytokinesis, while overexpression of constitutively active Cdc42 (CACdc42) caused cytokinesis failure in normal rat kidney epithelial cells. Cells depleted of Cdc42 displayed abnormal cell morphology and caused a failure of tight accumulation of actin and RhoA at the equator. In contrast, in cells overexpressing CACdc42, actin formed abnormal bundles and RhoA was largely eliminated from the equator. Our results suggest that accurate regulation of Cdc42 activity is crucial for proper equatorial actin assembly and RhoA localization during cytokinesis. Notably, our observations also suggest that tight actin concentration is not essential for cytokinesis in adherent mammalian cells. PMID:21763307

  13. Replication of human immunodeficiency virus type 1 in primary dendritic cell cultures.

    PubMed Central

    Langhoff, E; Terwilliger, E F; Bos, H J; Kalland, K H; Poznansky, M C; Bacon, O M; Haseltine, W A

    1991-01-01

    The ability of the human immunodeficiency virus type 1 (HIV-1) to replicate in primary blood dendritic cells was investigated. Dendritic cells compose less than 1% of the circulating leukocytes and are nondividing cells. Highly purified preparations of dendritic cells were obtained using recent advances in cell fractionation. The results of these experiments show that dendritic cells, in contrast to monocytes and T cells, support the active replication of all strains of HIV-1 tested, including T-cell tropic and monocyte/macrophage tropic isolates. The dendritic cell cultures supported much more virus production than did cultures of primary unseparated T cells, CD4+ T cells, and adherent as well as nonadherent monocytes. Replication of HIV-1 in dendritic cells produces no noticeable cytopathic effect nor does it decrease total cell number. The ability of the nonreplicating dendritic cells to support high levels of replication of HIV-1 suggests that this antigen-presenting cell population, which is also capable of supporting clonal T-cell growth, may play a central role in HIV pathogenesis, serving as a source of continued infection of CD4+ T cells and as a reservoir of virus infection. Images PMID:1910172

  14. Cell elasticity with altered cytoskeletal architectures across multiple cell types.

    PubMed

    Grady, Martha E; Composto, Russell J; Eckmann, David M

    2016-08-01

    The cytoskeleton is primarily responsible for providing structural support, localization and transport of organelles, and intracellular trafficking. The structural support is supplied by actin filaments, microtubules, and intermediate filaments, which contribute to overall cell elasticity to varying degrees. We evaluate cell elasticity in five different cell types with drug-induced cytoskeletal derangements to probe how actin filaments and microtubules contribute to cell elasticity and whether it is conserved across cell type. Specifically, we measure elastic stiffness in primary chondrocytes, fibroblasts, endothelial cells (HUVEC), hepatocellular carcinoma cells (HUH-7), and fibrosarcoma cells (HT 1080) subjected to two cytoskeletal destabilizers: cytochalasin D and nocodazole, which disrupt actin and microtubule polymerization, respectively. Elastic stiffness is measured by atomic force microscopy (AFM) and the disruption of the cytoskeleton is confirmed using fluorescence microscopy. The two cancer cell lines showed significantly reduced elastic moduli values (~0.5kPa) when compared to the three healthy cell lines (~2kPa). Non-cancer cells whose actin filaments were disrupted using cytochalasin D showed a decrease of 60-80% in moduli values compared to untreated cells of the same origin, whereas the nocodazole-treated cells showed no change in elasticity. Overall, we demonstrate actin filaments contribute more to elastic stiffness than microtubules but this result is cell type dependent. Cancer cells behaved differently, exhibiting increased stiffness as well as stiffness variability when subjected to nocodazole. We show that disruption of microtubule dynamics affects cancer cell elasticity, suggesting therapeutic drugs targeting microtubules be monitored for significant elastic changes. PMID:26874250

  15. Cell culture models using rat primary alveolar type I cells

    PubMed Central

    Downs, Charles A.; Montgomery, David W.; Merkle, Carrie J.

    2011-01-01

    There is a lack of cell culture models using primary alveolar type I (AT I) cells. The purpose of this study was to develop cell culture models using rat AT I cells and microvascular endothelial cells from the lung (MVECL). Two types of model systems were developed: single and co-culture systems; additionally a 3-dimensional model system was developed. Pure AT I cell (96.3 ±2.7%) and MVECL (97.9 ±1.1 %) preparations were used. AT I cell morphology, mitochondrial number and distribution, actin filament arrangement and number of apoptotic cells at confluence, and telomere attrition were characterized. AT I cells maintained their morphometric characteristics through at least population doubling (PD) 35, while demonstrating telomere attrition through at least PD 100. Furthermore, AT I cells maintained the expression of their specific markers, T1α and AQ-5, through PD 42. For the co-cultures, AT I cells were grown on the top and MVECL were grown on the bottom of fibronectin coated 24 well Transwell Fluroblok™ filter inserts. Neither cell type transmigrated the 1 micron pores. Additionally AT I cells were grown in a thick layer of Matrigel® to create a 3-dimensional model in which primary AT I cells form ring-like structures that resemble an alveolus. The development of these model systems offers the opportunities to investigate AT I cell cells and their interactions with MVECL in response to pharmacological interventions and in the processes of disease, repair and regeneration. PMID:21624488

  16. Evaluation of 309 environmental chemicals using a mouse embryonic stem cell adherent cell differentiation and cytotoxicity assay

    EPA Science Inventory

    The vast landscape of environmental chemicals has motivated the need for alternative methods to traditional whole-animal bioassays in toxicity testing. Embryonic stem (ES) cells provide an in vitro model of embryonic development and an alternative method for assessing development...

  17. Cell culture models using rat primary alveolar type I cells.

    PubMed

    Downs, Charles A; Montgomery, David W; Merkle, Carrie J

    2011-10-01

    There is a lack of cell culture models using primary alveolar type I (AT I) cells. The purpose of this study was to develop cell culture models using rat AT I cells and microvascular endothelial cells from the lung (MVECL). Two types of model systems were developed: single and co-culture systems; additionally a 3-dimensional model system was developed. Pure AT I cell (96.3 ± 2.7%) and MVECL (97.9 ± 1.1%) preparations were used. AT I cell morphology, mitochondrial number and distribution, actin filament arrangement and number of apoptotic cells at confluence, and telomere attrition were characterized. AT I cells maintained their morphometric characteristics through at least population doubling (PD) 35, while demonstrating telomere attrition through at least PD 100. Furthermore, AT I cells maintained the expression of their specific markers, T1α and AQ-5, through PD 42. For the co-cultures, AT I cells were grown on the top and MVECL were grown on the bottom of fibronectin-coated 24-well Transwell Fluroblok™ filter inserts. Neither cell type transmigrated the 1 μm pores. Additionally, AT I cells were grown in a thick layer of Matrigel(®) to create a 3-dimensional model in which primary AT I cells form ring-like structures that resemble an alveolus. The development of these model systems offers the opportunities to investigate AT I cells and their interactions with MVECL in response to pharmacological interventions and in the processes of disease, repair and regeneration. PMID:21624488

  18. Multiple roles of Activin/Nodal, bone morphogenetic protein, fibroblast growth factor and Wnt/β-catenin signalling in the anterior neural patterning of adherent human embryonic stem cell cultures

    PubMed Central

    Lupo, Giuseppe; Novorol, Claire; Smith, Joseph R.; Vallier, Ludovic; Miranda, Elena; Alexander, Morgan; Biagioni, Stefano; Pedersen, Roger A.; Harris, William A.

    2013-01-01

    Several studies have successfully produced a variety of neural cell types from human embryonic stem cells (hESCs), but there has been limited systematic analysis of how different regional identities are established using well-defined differentiation conditions. We have used adherent, chemically defined cultures to analyse the roles of Activin/Nodal, bone morphogenetic protein (BMP), fibroblast growth factor (FGF) and Wnt/β-catenin signalling in neural induction, anteroposterior patterning and eye field specification in hESCs. We show that either BMP inhibition or activation of FGF signalling is required for effective neural induction, but these two pathways have distinct outcomes on rostrocaudal patterning. While BMP inhibition leads to specification of forebrain/midbrain positional identities, FGF-dependent neural induction is associated with strong posteriorization towards hindbrain/spinal cord fates. We also demonstrate that Wnt/β-catenin signalling is activated during neural induction and promotes acquisition of neural fates posterior to forebrain. Therefore, inhibition of this pathway is needed for efficient forebrain specification. Finally, we provide evidence that the levels of Activin/Nodal and BMP signalling have a marked influence on further forebrain patterning and that constitutive inhibition of these pathways represses expression of eye field genes. These results show that the key mechanisms controlling neural patterning in model vertebrate species are preserved in adherent, chemically defined hESC cultures and reveal new insights into the signals regulating eye field specification. PMID:23576785

  19. Multiple roles of Activin/Nodal, bone morphogenetic protein, fibroblast growth factor and Wnt/β-catenin signalling in the anterior neural patterning of adherent human embryonic stem cell cultures.

    PubMed

    Lupo, Giuseppe; Novorol, Claire; Smith, Joseph R; Vallier, Ludovic; Miranda, Elena; Alexander, Morgan; Biagioni, Stefano; Pedersen, Roger A; Harris, William A

    2013-04-01

    Several studies have successfully produced a variety of neural cell types from human embryonic stem cells (hESCs), but there has been limited systematic analysis of how different regional identities are established using well-defined differentiation conditions. We have used adherent, chemically defined cultures to analyse the roles of Activin/Nodal, bone morphogenetic protein (BMP), fibroblast growth factor (FGF) and Wnt/β-catenin signalling in neural induction, anteroposterior patterning and eye field specification in hESCs. We show that either BMP inhibition or activation of FGF signalling is required for effective neural induction, but these two pathways have distinct outcomes on rostrocaudal patterning. While BMP inhibition leads to specification of forebrain/midbrain positional identities, FGF-dependent neural induction is associated with strong posteriorization towards hindbrain/spinal cord fates. We also demonstrate that Wnt/β-catenin signalling is activated during neural induction and promotes acquisition of neural fates posterior to forebrain. Therefore, inhibition of this pathway is needed for efficient forebrain specification. Finally, we provide evidence that the levels of Activin/Nodal and BMP signalling have a marked influence on further forebrain patterning and that constitutive inhibition of these pathways represses expression of eye field genes. These results show that the key mechanisms controlling neural patterning in model vertebrate species are preserved in adherent, chemically defined hESC cultures and reveal new insights into the signals regulating eye field specification. PMID:23576785

  20. Combined strategies for optimal detection of the contact point in AFM force-indentation curves obtained on thin samples and adherent cells.

    PubMed

    Gavara, Núria

    2016-01-01

    Atomic Force Microscopy (AFM) is a widely used tool to study cell mechanics. Current AFM setups perform high-throughput probing of living cells, generating large amounts of force-indentations curves that are subsequently analysed using a contact-mechanics model. Here we present several algorithms to detect the contact point in force-indentation curves, a crucial step to achieve fully-automated analysis of AFM-generated data. We quantify and rank the performance of our algorithms by analysing a thousand force-indentation curves obtained on thin soft homogeneous hydrogels, which mimic the stiffness and topographical profile of adherent cells. We take advantage of the fact that all the proposed algorithms are based on sequential search strategies, and show that a combination of them yields the most accurate and unbiased results. Finally, we also observe improved performance when force-indentation curves obtained on adherent cells are analysed using our combined strategy, as compared to the classical algorithm used in the majority of previous cell mechanics studies. PMID:26891762

  1. Combined strategies for optimal detection of the contact point in AFM force-indentation curves obtained on thin samples and adherent cells

    PubMed Central

    Gavara, Núria

    2016-01-01

    Atomic Force Microscopy (AFM) is a widely used tool to study cell mechanics. Current AFM setups perform high-throughput probing of living cells, generating large amounts of force-indentations curves that are subsequently analysed using a contact-mechanics model. Here we present several algorithms to detect the contact point in force-indentation curves, a crucial step to achieve fully-automated analysis of AFM-generated data. We quantify and rank the performance of our algorithms by analysing a thousand force-indentation curves obtained on thin soft homogeneous hydrogels, which mimic the stiffness and topographical profile of adherent cells. We take advantage of the fact that all the proposed algorithms are based on sequential search strategies, and show that a combination of them yields the most accurate and unbiased results. Finally, we also observe improved performance when force-indentation curves obtained on adherent cells are analysed using our combined strategy, as compared to the classical algorithm used in the majority of previous cell mechanics studies. PMID:26891762

  2. Gain-of-Function Mutations in PDR1, a Regulator of Antifungal Drug Resistance in Candida glabrata, Control Adherence to Host Cells

    PubMed Central

    Vale-Silva, Luís; Ischer, Françoise; Leibundgut-Landmann, Salomé

    2013-01-01

    Candida glabrata is an emerging opportunistic pathogen that is known to develop resistance to azole drugs due to increased drug efflux. The mechanism consists of CgPDR1-mediated upregulation of ATP-binding cassette transporters. A range of gain-of-function (GOF) mutations in CgPDR1 have been found to lead not only to azole resistance but also to enhanced virulence. This implicates CgPDR1 in the regulation of the interaction of C. glabrata with the host. To identify specific CgPDR1-regulated steps of the host-pathogen interaction, we investigated in this work the interaction of selected CgPDR1 GOF mutants with murine bone marrow-derived macrophages and human acute monocytic leukemia cell line (THP-1)-derived macrophages, as well as different epithelial cell lines. GOF mutations in CgPDR1 did not influence survival and replication within macrophages following phagocytosis but led to decreased adherence to and uptake by macrophages. This may allow evasion from the host's innate cellular immune response. The interaction with epithelial cells revealed an opposite trend, suggesting that GOF mutations in CgPDR1 may favor epithelial colonization of the host by C. glabrata through increased adherence to epithelial cell layers. These data reveal that GOF mutations in CgPDR1 modulate the interaction with host cells in ways that may contribute to increased virulence. PMID:23460523

  3. Combined strategies for optimal detection of the contact point in AFM force-indentation curves obtained on thin samples and adherent cells

    NASA Astrophysics Data System (ADS)

    Gavara, Núria

    2016-02-01

    Atomic Force Microscopy (AFM) is a widely used tool to study cell mechanics. Current AFM setups perform high-throughput probing of living cells, generating large amounts of force-indentations curves that are subsequently analysed using a contact-mechanics model. Here we present several algorithms to detect the contact point in force-indentation curves, a crucial step to achieve fully-automated analysis of AFM-generated data. We quantify and rank the performance of our algorithms by analysing a thousand force-indentation curves obtained on thin soft homogeneous hydrogels, which mimic the stiffness and topographical profile of adherent cells. We take advantage of the fact that all the proposed algorithms are based on sequential search strategies, and show that a combination of them yields the most accurate and unbiased results. Finally, we also observe improved performance when force-indentation curves obtained on adherent cells are analysed using our combined strategy, as compared to the classical algorithm used in the majority of previous cell mechanics studies.

  4. Fuel cells: principles, types, fuels, and applications.

    PubMed

    Carrette, L; Friedrich, K A; Stimming, U

    2000-12-15

    During the last decade, fuel cells have received enormous attention from research institutions and companies as novel electrical energy conversion systems. In the near future, they will see application in automotive propulsion, distributed power generation, and in low power portable devices (battery replacement). This review gives an introduction into the fundamentals and applications of fuel cells: Firstly, the environmental and social factors promoting fuel cell development are discussed, with an emphasis on the advantages of fuel cells compared to the conventional techniques. Then, the main reactions, which are responsible for the conversion of chemical into electrical energy in fuel cells, are given and the thermodynamic and kinetic fundamentals are stated. The theoretical and real efficiencies of fuel cells are also compared to that of internal combustion engines. Next, the different types of fuel cells and their main components are explained and the related material issues are presented. A section is devoted to fuel generation and storage, which is of paramount importance for the practical aspects of fuel cell use. Finally, attention is given to the integration of the fuel cells into complete systems. PMID:23696319

  5. Carbon source-induced reprogramming of the cell wall proteome and secretome modulates the adherence and drug resistance of the fungal pathogen Candida albicans

    PubMed Central

    Ene, Iuliana V; Heilmann, Clemens J; Sorgo, Alice G; Walker, Louise A; de Koster, Chris G; Munro, Carol A; Klis, Frans M; Brown, Alistair J P

    2012-01-01

    The major fungal pathogen Candida albicans can occupy diverse microenvironments in its human host. During colonization of the gastrointestinal or urogenital tracts, mucosal surfaces, bloodstream, and internal organs, C. albicans thrives in niches that differ with respect to available nutrients and local environmental stresses. Although most studies are performed on glucose-grown cells, changes in carbon source dramatically affect cell wall architecture, stress responses, and drug resistance. We show that growth on the physiologically relevant carboxylic acid, lactate, has a significant impact on the C. albicans cell wall proteome and secretome. The regulation of cell wall structural proteins (e.g. Cht1, Phr1, Phr2, Pir1) correlated with extensive cell wall remodeling in lactate-grown cells and with their increased resistance to stresses and antifungal drugs, compared with glucose-grown cells. Moreover, changes in other proteins (e.g. Als2, Gca1, Phr1, Sap9) correlated with the increased adherence and biofilm formation of lactate-grown cells. We identified mating and pheromone-regulated proteins that were exclusive to lactate-grown cells (e.g. Op4, Pga31, Pry1, Scw4, Yps7) as well as mucosa-specific and other niche-specific factors such as Lip4, Pga4, Plb5, and Sap7. The analysis of the corresponding null mutants confirmed that many of these proteins contribute to C. albicans adherence, stress, and antifungal drug resistance. Therefore, the cell wall proteome and secretome display considerable plasticity in response to carbon source. This plasticity influences important fitness and virulence attributes known to modulate the behavior of C. albicans in different host microenvironments during infection. PMID:22997008

  6. Large-scale enrichment of mobilized CD34+ peripheral blood hematopoietic progenitors by removal of nylon wool-adherent mature cells.

    PubMed

    Di Nicola, M; Siena, S; Bregni, M; Ravagnani, F; Vitello, F; Belli, N; Dodero, A; Magni, M; Bonadonna, G; Gianni, A M

    1994-12-01

    With the aim of facilitating the ex vivo manipulation of peripheral blood hematopoietic progenitors (CPCs = circulating progenitor cells) collected by leukapheresis, we removed polymorphonuclear cells and monocytes that naturally adhere to nylon wool fibers. Leukapheresed cells harvested at the time of hematopoietic recovery after cancer therapy with high-dose cyclophosphamide plus hematopoietic growth factors were incubated with nylon wool fibers for 1 h at 37 degrees C. Evaluation of the cells non-adherent to the nylon wool in all experiments (n = 14) showed that the median recovery of nucleated cells and CPCs detected as CD34+ cells, CFU-GM and BFU-E was 16.4% (range 4.8%-34.0%), 60.0% (range 30.8-80.8%), 60.9% (range 33.4-74.5%) and 65.5% (range 30.8-69.2%), respectively. Therefore exposure to the nylon wool determined a selective removal of mature cells and a complementary enrichment of CPCs. The wide range of results depended on the significantly different cell compositions of the unmanipulated leukaphereses. The latter from patients receiving rhG-CSF (n = 10) comprised a median of 88.5% (range 77.8-93.8%) and 11.5% (range 6.2-22.2%) polymorphonuclear and mononuclear cells, respectively. In contrast, leukaphereses from patients receiving rhGM-CSF or PIXY321 (n = 4) comprised a median of 71.1% (range 55.4-85.0%) and 28.9% (range 15.0-44.6%) polymorphonuclear and mononuclear cells, respectively.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7536068

  7. Production of high-titer human influenza A virus with adherent and suspension MDCK cells cultured in a single-use hollow fiber bioreactor.

    PubMed

    Tapia, Felipe; Vogel, Thomas; Genzel, Yvonne; Behrendt, Ilona; Hirschel, Mark; Gangemi, J David; Reichl, Udo

    2014-02-12

    Hollow fiber bioreactors (HFBRs) have been widely described as capable of supporting the production of highly concentrated monoclonal antibodies and recombinant proteins. Only recently HFBRs have been proposed as new single-use platforms for production of high-titer influenza A virus. These bioreactors contain multiple hollow fiber capillary tubes that separate the bioreactor in an intra- and an extra-capillary space. Cells are usually cultured in the extra-capillary space and can grow to a very high cell concentration. This work describes the evaluation of the single-use hollow fiber bioreactor PRIMER HF (Biovest International Inc., USA) for production of influenza A virus. The process was setup, characterized and optimized by running a total of 15 cultivations. The HFBRs were seeded with either adherent or suspension MDCK cells, and infected with influenza virus A/PR/8/34 (H1N1), and the pandemic strain A/Mexico/4108/2009 (H1N1). High HA titers and TCID₅₀ of up to 3.87 log₁₀(HA units/100 μL) and 1.8 × 10(10)virions/mL, respectively, were obtained for A/PR/8/34 influenza strain. Influenza virus was collected by performing multiple harvests of the extra-capillary space during a virus production time of up to 12 days. Cell-specific virus yields between 2,000 and 8,000 virions/cell were estimated for adherent MDCK cells, and between 11,000 and 19,000 virions/cell for suspension MDCK.SUS2 cells. These results do not only coincide with the cell-specific virus yields obtained with cultivations in stirred tank bioreactors and other high cell density systems, but also demonstrate that HFBRs are promising and competitive single-use platforms that can be considered for commercial production of influenza virus. PMID:24269322

  8. Distinct Cell Clusters Touching Islet Cells Induce Islet Cell Replication in Association with Over-Expression of Regenerating Gene (REG) Protein in Fulminant Type 1 Diabetes

    PubMed Central

    Aida, Kaoru; Saitoh, Sei; Nishida, Yoriko; Yokota, Sadanori; Ohno, Shinichi; Mao, Xiayang; Akiyama, Daiichiro; Tanaka, Shoichiro; Awata, Takuya; Shimada, Akira; Oikawa, Youichi; Shimura, Hiroki; Furuya, Fumihiko; Takizawa, Soichi; Ichijo, Masashi; Ichijo, Sayaka; Itakura, Jun; Fujii, Hideki; Hashiguchi, Akinori; Takasawa, Shin; Endo, Toyoshi; Kobayashi, Tetsuro

    2014-01-01

    Background Pancreatic islet endocrine cell-supporting architectures, including islet encapsulating basement membranes (BMs), extracellular matrix (ECM), and possible cell clusters, are unclear. Procedures The architectures around islet cell clusters, including BMs, ECM, and pancreatic acinar-like cell clusters, were studied in the non-diabetic state and in the inflamed milieu of fulminant type 1 diabetes in humans. Result Immunohistochemical and electron microscopy analyses demonstrated that human islet cell clusters and acinar-like cell clusters adhere directly to each other with desmosomal structures and coated-pit-like structures between the two cell clusters. The two cell-clusters are encapsulated by a continuous capsule composed of common BMs/ECM. The acinar-like cell clusters have vesicles containing regenerating (REG) Iα protein. The vesicles containing REG Iα protein are directly secreted to islet cells. In the inflamed milieu of fulminant type 1 diabetes, the acinar-like cell clusters over-expressed REG Iα protein. Islet endocrine cells, including beta-cells and non-beta cells, which were packed with the acinar-like cell clusters, show self-replication with a markedly increased number of Ki67-positive cells. Conclusion The acinar-like cell clusters touching islet endocrine cells are distinct, because the cell clusters are packed with pancreatic islet clusters and surrounded by common BMs/ECM. Furthermore, the acinar-like cell clusters express REG Iα protein and secrete directly to neighboring islet endocrine cells in the non-diabetic state, and the cell clusters over-express REG Iα in the inflamed milieu of fulminant type 1 diabetes with marked self-replication of islet cells. PMID:24759849

  9. Hydroxy decenoic acid down regulates gtfB and gtfC expression and prevents Streptococcus mutans adherence to the cell surfaces

    PubMed Central

    2012-01-01

    Background 10-Hydroxy-2-decenoic acid, an unsaturated fatty acid is the most active and unique component to the royal jelly that has antimicrobial properties. Streptococcus mutans is associated with pathogenesis of oral cavity, gingivoperiodontal diseases and bacteremia following dental manipulations. In the oral cavity, S. mutans colonize the soft tissues including tongue, palate, and buccal mucosa. When considering the role of supragingival dental plaque in caries, the proportion of acid producing bacteria (particularly S. mutans), has direct relevance to the pathogenicity of the plaque. The genes that encode glucosyltransferases (gtfs) especially gtfB and gtfC are important in S. mutans colonization and pathogenesis. This study investigated the hydroxy-decenoic acid (HDA) effects on gtfB and gtfC expression and S. mutans adherence to cells surfaces. Methods Streptococcus mutans was treated by different concentrations of HPLC purified HDA supplied by Iran Beekeeping and Veterinary Association. Real time RT-PCR and western blot assays were conducted to evaluate gtfB and gtfC genes transcription and translation before and after HDA treatment. The bacterial attachment to the cell surfaces was evaluated microscopically. Results 500 μg ml-1 of HDA inhibited gtfB and gtfC mRNA transcription and its expression. The same concentration of HDA decreased 60% the adherence of S. mutans to the surface of P19 cells. Conclusion Hydroxy-decenoic acid prevents gtfB and gtfC expression efficiently in the bactericide sub-concentrations and it could effectively reduce S. mutans adherence to the cell surfaces. In the future, therapeutic approaches to affecting S. mutans could be selective and it’s not necessary to put down the oral flora completely. PMID:22839724

  10. Uptake and cellular distribution of nucleolar targeting peptides (NrTPs) in different cell types.

    PubMed

    Rodrigues, Margarida; Andreu, David; Santos, Nuno C

    2015-03-01

    Nucleolar targeting peptides (NrTPs) are a family of cell penetrating peptides (CPPs) derived from crotamine, a rattlesnake venom toxin. They were named NrTPs for their remarkable nucleolus-homing properties and have been studied for their potential as drug delivery vehicles. Live cell microscopy experiments were conducted to monitor NrTP uptake and distribution in different cell types, including primary cells (PBMCs and erythrocytes) and different immortalized cell lines (HeLa, BHK21, BV-173, and MOLT-4). Uptake dependence on cell type (primary vs. immortalized, suspension vs. adherent, cancer vs. healthy cells), peptide concentration and cell viability were evaluated. To gain further insight on the internalization mechanism, uptake kinetics was also monitored. Results showed the uptake and distribution pattern as strongly dependent on peptide sequence, peptide concentration and membrane constituents. Under similar conditions, NrTP6 is more internalized than NrTP1, NrTP2 and NrTP5. Additionally, while internalization of NrTP7 and NrTP8 may cause cytotoxicity, NrTP6 is noncytotoxic. Higher peptide concentrations can be correlated to nucleolar targeting, although even at low concentrations a residual number of cells reveal positive nucleolar labeling. NrTPs were successfully internalized into all cell types tested except erythrocytes. PMID:25620660

  11. Efficient entry of cell-penetrating peptide nona-arginine into adherent cells involves a transient increase in intracellular calcium

    PubMed Central

    Melikov, Kamran; Hara, Ann; Yamoah, Kwabena; Zaitseva, Elena; Zaitsev, Eugene; Chernomordik, Leonid V.

    2015-01-01

    Understanding the mechanism of entry of cationic peptides such as nona-arginine (R9) into cells remains an important challenge to their use as efficient drug-delivery vehicles. At nanomolar to low micromolar R9 concentrations and at physiological temperature, peptide entry involves endocytosis. In contrast, at a concentration ≥10 μM, R9 induces a very effective non-endocytic entry pathway specific for cationic peptides. We found that a similar entry pathway is induced at 1–2 μM concentrations of R9 if peptide application is accompanied by a rapid temperature drop to 15°C. Both at physiological and at sub-physiological temperatures, this entry mechanism was inhibited by depletion of the intracellular ATP pool. Intriguingly, we found that R9 at 10–20 μM and 37°C induces repetitive spikes in intracellular Ca2+ concentration. This Ca2+ signalling correlated with the efficiency of the peptide entry. Pre-loading cells with the Ca2+ chelator BAPTA (1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid) inhibited both Ca2+ spikes and peptide entry, suggesting that an increase in intracellular Ca2+ precedes and is required for peptide entry. One of the hallmarks of Ca2+ signalling is a transient cell-surface exposure of phosphatidylserine (PS), a lipid normally residing only in the inner leaflet of the plasma membrane. Blocking the accessible PS with the PS-binding domain of lactadherin strongly inhibited non-endocytic R9 entry, suggesting the importance of PS externalization in this process. To conclude, we uncovered a novel mechanistic link between calcium signalling and entry of cationic peptides. This finding will enhance our understanding of the properties of plasma membrane and guide development of future drug-delivery vehicles. PMID:26272944

  12. DNA typing of epithelial cells after strangulation.

    PubMed

    Wiegand, P; Kleiber, M

    1997-01-01

    DNA typing was carried out on epithelial cells which were transferred from the hands of the suspect onto the neck of the victim. In an experimental study 16 suspect-victim combinations were investigated for estimating the typing success. Alternatively to an attack against the neck, the upper arm was used for "strangulation". PCR typing was carried out using the short tandem repeat systems (STRs) HumCD4, HumVWF31A (VWA) and Hum-FIBRA (FGA) and the success rate was > 70% for all 3 systems. In most of the cases mixed patterns containing the phenotype of the suspect and the victim were obtained. In a case where strangulation was the cause of death, epithelial cells could be removed from the neck of the victim. The DNA pattern of the suspect could be successfully amplified using four STRs, demonstrating the applicability of this approach for practical casework. PMID:9274940

  13. Improving medication adherence in diabetes type 2 patients through Real Time Medication Monitoring: a Randomised Controlled Trial to evaluate the effect of monitoring patients' medication use combined with short message service (SMS) reminders

    PubMed Central

    2011-01-01

    Background Innovative approaches are needed to support patients' adherence to drug therapy. The Real Time Medication Monitoring (RTMM) system offers real time monitoring of patients' medication use combined with short message service (SMS) reminders if patients forget to take their medication. This combination of monitoring and tailored reminders provides opportunities to improve adherence. This article describes the design of an intervention study aimed at evaluating the effect of RTMM on adherence to oral antidiabetics. Methods/Design Randomised Controlled Trial (RCT) with two intervention arms and one control arm involving diabetes type 2 patients with suboptimal levels of adherence to oral antidiabetics (less than 80% based on pharmacy refill data). Patients in the first intervention arm use RTMM including SMS reminders and a personal webpage where they can monitor their medication use. Patients in the second intervention arm use RTMM without SMS reminders or webpage access. Patients in the control arm are not exposed to any intervention. Patients are randomly assigned to one of the three arms. The intervention lasts for six months. Pharmacy refill data of all patients are available from 11 months before, until 11 months after the start of the intervention. Primary outcome measure is adherence to oral antidiabetics calculated from: 1) data collected with RTMM, as a percentage of medication taken as prescribed, and as percentage of medication taken within the correct time interval, 2) refill data, taking the number of days for which oral antidiabetics are dispensed during the study period divided by the total number of days of the study period. Differences in adherence between the intervention groups and control group are studied using refill data. Differences in adherence between the two intervention groups are studied using RTMM data. Discussion The intervention described in this article consists of providing RTMM to patients with suboptimal adherence levels

  14. Cell-surface serglycin promotes adhesion of myeloma cells to collagen type I and affects the expression of matrix metalloproteinases.

    PubMed

    Skliris, Antonis; Labropoulou, Vassiliki T; Papachristou, Dionysios J; Aletras, Alexios; Karamanos, Nikos K; Theocharis, Achilleas D

    2013-05-01

    Serglycin (SG) is mainly expressed by hematopoetic cells as an intracellular proteoglycan. Multiple myeloma cells constitutively secrete SG, which is also localized on the cell surface in some cell lines. In this study, SG isolated from myeloma cells was found to interact with collagen type I (Col I), which is a major bone matrix component. Notably, myeloma cells positive for cell-surface SG (csSG) adhered significantly to Col I, compared to cells lacking csSG. Removal of csSG by treatment of the cells with chondroitinase ABC or blocking of csSG by an SG-specific polyclonal antibody significantly reduced the adhesion of myeloma cells to Col I. Significant up-regulation of expression of the matrix metalloproteinases MMP-2 and MMP-9 at both the mRNA and protein levels was observed when culturing csSG-positive myeloma cells on Col I-coated dishes or in the presence of soluble Col I. MMP-9 and MMP-2 were also expressed in increased amounts by myeloma cells in the bone marrow of patients with multiple myeloma. Our data indicate that csSG of myeloma cells affects key functional properties, such as adhesion to Col I and the expression of MMPs, and imply that csSG may serve as a potential prognostic factor and/or target for pharmacological interventions in multiple myeloma. PMID:23387827

  15. Hematopoietic Cancer Cell Lines Can Support Replication of Sabin Poliovirus Type 1

    PubMed Central

    van Eikenhorst, Gerco; de Gruijl, Tanja D.; van der Pol, Leo A.; Bakker, Wilfried A. M.

    2015-01-01

    Viral vaccines can be produced in adherent or in suspension cells. The objective of this work was to screen human suspension cell lines for the capacity to support viral replication. As the first step, it was investigated whether poliovirus can replicate in such cell lines. Sabin poliovirus type 1 was serially passaged on five human cell lines, HL60, K562, KG1, THP-1, and U937. Sabin type 1 was capable of efficiently replicating in three cell lines (K562, KG1, and U937), yielding high viral titers after replication. Expression of CD155, the poliovirus receptor, did not explain susceptibility to replication, since all cell lines expressed CD155. Furthermore, we showed that passaged virus replicated more efficiently than parental virus in KG1 cells, yielding higher virus titers in the supernatant early after infection. Infection of cell lines at an MOI of 0.01 resulted in high viral titers in the supernatant at day 4. Infection of K562 with passaged Sabin type 1 in a bioreactor system yielded high viral titers in the supernatant. Altogether, these data suggest that K562, KG1, and U937 cell lines are useful for propagation of poliovirus. PMID:25815312

  16. Investigation on cytoskeleton dynamics for no-adherent cells subjected to point-like stimuli by digital holographic microscopy and holographic optical trapping

    NASA Astrophysics Data System (ADS)

    Miccio, Lisa; Merola, Francesco; Memmolo, Pasquale; Mugnano, Martina; Fusco, Sabato; Netti, Paolo A.; Ferraro, Pietro

    2014-05-01

    Guiding, controlling and studying cellular functions are challenging themes in the biomedical field, as they are fundamental prerequisites for new therapeutic strategies from tissue regeneration to controlled drug delivery. In recent years, multidisciplinary studies in nanotechnology offer new tools to investigate important biophysical phenomena in response to the local physical characteristics of the extracellular environment, some examples are the mechanisms of cell adhesion, migration, communication and differentiation. Indeed for reproducing the features of the extracellular matrix in vitro, it is essential to develop active devices that evoke as much as possible the natural cellular environment. Our investigation is in the framework of studying and clarifying the biophysical mechanisms of the interaction between cells and the microenvironment in which they exist. We implement an optical tweezers setup to investigate cell material interaction and we use Digital Holography as non-invasive imaging technique in microscopy. We exploit Holographic Optical Tweezers arrangement in order to trap and manage functionalized micrometric latex beads to induce mechanical deformation in suspended cells. A lot of papers in literature examine the dynamics of the cytoskeleton when cells adhere on substrates and nowadays well established cell models are based on such research activities. Actually, the natural cell environment is made of a complex extracellular matrix and the single cell behavior is due to intricate interactions with the environment and are strongly correlated to the cell-cell interactions. Our investigation is devoted to understand the inner cell mechanism when it is mechanically stressed by point-like stimulus without the substrate influence.

  17. Identification of Vulnerable Cell Types in Major Brain Disorders Using Single Cell Transcriptomes and Expression Weighted Cell Type Enrichment.

    PubMed

    Skene, Nathan G; Grant, Seth G N

    2016-01-01

    The cell types that trigger the primary pathology in many brain diseases remain largely unknown. One route to understanding the primary pathological cell type for a particular disease is to identify the cells expressing susceptibility genes. Although this is straightforward for monogenic conditions where the causative mutation may alter expression of a cell type specific marker, methods are required for the common polygenic disorders. We developed the Expression Weighted Cell Type Enrichment (EWCE) method that uses single cell transcriptomes to generate the probability distribution associated with a gene list having an average level of expression within a cell type. Following validation, we applied EWCE to human genetic data from cases of epilepsy, Schizophrenia, Autism, Intellectual Disability, Alzheimer's disease, Multiple Sclerosis and anxiety disorders. Genetic susceptibility primarily affected microglia in Alzheimer's and Multiple Sclerosis; was shared between interneurons and pyramidal neurons in Autism and Schizophrenia; while intellectual disabilities and epilepsy were attributable to a range of cell-types, with the strongest enrichment in interneurons. We hypothesized that the primary cell type pathology could trigger secondary changes in other cell types and these could be detected by applying EWCE to transcriptome data from diseased tissue. In Autism, Schizophrenia and Alzheimer's disease we find evidence of pathological changes in all of the major brain cell types. These findings give novel insight into the cellular origins and progression in common brain disorders. The methods can be applied to any tissue and disorder and have applications in validating mouse models. PMID:26858593

  18. Identification of Vulnerable Cell Types in Major Brain Disorders Using Single Cell Transcriptomes and Expression Weighted Cell Type Enrichment

    PubMed Central

    Skene, Nathan G.; Grant, Seth G. N.

    2016-01-01

    The cell types that trigger the primary pathology in many brain diseases remain largely unknown. One route to understanding the primary pathological cell type for a particular disease is to identify the cells expressing susceptibility genes. Although this is straightforward for monogenic conditions where the causative mutation may alter expression of a cell type specific marker, methods are required for the common polygenic disorders. We developed the Expression Weighted Cell Type Enrichment (EWCE) method that uses single cell transcriptomes to generate the probability distribution associated with a gene list having an average level of expression within a cell type. Following validation, we applied EWCE to human genetic data from cases of epilepsy, Schizophrenia, Autism, Intellectual Disability, Alzheimer's disease, Multiple Sclerosis and anxiety disorders. Genetic susceptibility primarily affected microglia in Alzheimer's and Multiple Sclerosis; was shared between interneurons and pyramidal neurons in Autism and Schizophrenia; while intellectual disabilities and epilepsy were attributable to a range of cell-types, with the strongest enrichment in interneurons. We hypothesized that the primary cell type pathology could trigger secondary changes in other cell types and these could be detected by applying EWCE to transcriptome data from diseased tissue. In Autism, Schizophrenia and Alzheimer's disease we find evidence of pathological changes in all of the major brain cell types. These findings give novel insight into the cellular origins and progression in common brain disorders. The methods can be applied to any tissue and disorder and have applications in validating mouse models. PMID:26858593

  19. A Brief Adherence Intervention that Improved Glycemic Control: Mediation by Patterns of Adherence

    PubMed Central

    de Vries McClintock, Heather F.; Morales, Knashawn H.; Small, Dylan S.; Bogner, Hillary R.

    2014-01-01

    This study examined whether longitudinal adherence profiles mediated the relationship between a brief adherence intervention and glycemic control among patients with Type 2 diabetes. Adherence was assessed using the Medication Event Monitoring System (MEMS). Longitudinal analysis via growth curve mixture modeling was carried out to classify patients according to patterns of adherence to oral hypoglycemic agents. Hemoglobin A1c (HbA1c) assays were used to measure glycemic control as the clinical outcome. Across the whole sample, longitudinal adherence profiles mediated 35.2% (13.2%, 81.0%) of the effect of a brief adherence intervention on glycemic control (from odds ratio (OR) = 8.48, 95% CI (3.24, 22.2) to 4.00, 95% CI (1.34, 11.93)). Our results suggest that patients in the intervention had better glycemic control largely due to their greater likelihood of adherence to oral hypoglycemic agents. PMID:24913600

  20. The pathogenic potential of Helicobacter cinaedi isolated from non-human sources: adherence, invasion and translocation ability in polarized intestinal epithelial Caco-2 cells in vitro.

    PubMed

    Taniguchi, Takako; Yamazaki, Wataru; Saeki, Yuji; Takajo, Ichiro; Okayama, Akihiko; Hayashi, Tetsuya; Misawa, Naoaki

    2016-05-01

    Helicobacter cinaedi infection has been recognized as an increasingly important emerging disease in humans. Infection with H. cinaedi causes bacteremia, cellulitis and enteritis. H. cinaedi has been isolated from non-human sources, including dogs, cats and rodents; however, it remains unclear whether animal strains are pathogenic in humans and as zoonotic pathogens. In this study, H. cinaedi isolates were recovered from a dog and a hamster, and the ability of these isolates to adhere to, invade and translocate across polarized human intestinal epithelial Caco-2 cells was examined in vitro. To better understand the pathogenic potential of animal H. cinaedi isolates, these results were compared with those for a human strain that was isolated from a patient with bacteremia. The animal and human strains adhered to and invaded Caco-2 cells, but to a lesser degree than the C. jejuni 81-176 strain, which was used as a control. The integrity of tight junctions was monitored by measuring transepithelial electrical resistance (TER) with a membrane insert system. The TER values for all H. cinaedi strains did not change during the experimental periods compared with those of the controls; however, translocation of H. cinaedi from the apical side to the basolateral side was confirmed by cultivation and H. cinaedi-specific PCR, suggesting that the H. cinaedi strains translocated by transcellular route. This study demonstrated that H. cinaedi strains of animal origin might have a pathogenic potential in human epithelial cells as observed in a translocation assay in vitro with a human isolate. PMID:26685883

  1. The pathogenic potential of Helicobacter cinaedi isolated from non-human sources: adherence, invasion and translocation ability in polarized intestinal epithelial Caco-2 cells in vitro

    PubMed Central

    TANIGUCHI, Takako; YAMAZAKI, Wataru; SAEKI, Yuji; TAKAJO, Ichiro; OKAYAMA, Akihiko; HAYASHI, Tetsuya; MISAWA, Naoaki

    2015-01-01

    Helicobacter cinaedi infection has been recognized as an increasingly important emerging disease in humans. Infection with H. cinaedi causes bacteremia, cellulitis and enteritis. H. cinaedi has been isolated from non-human sources, including dogs, cats and rodents; however, it remains unclear whether animal strains are pathogenic in humans and as zoonotic pathogens. In this study, H. cinaedi isolates were recovered from a dog and a hamster, and the ability of these isolates to adhere to, invade and translocate across polarized human intestinal epithelial Caco-2 cells was examined in vitro. To better understand the pathogenic potential of animal H. cinaedi isolates, these results were compared with those for a human strain that was isolated from a patient with bacteremia. The animal and human strains adhered to and invaded Caco-2 cells, but to a lesser degree than the C. jejuni 81–176 strain, which was used as a control. The integrity of tight junctions was monitored by measuring transepithelial electrical resistance (TER) with a membrane insert system. The TER values for all H. cinaedi strains did not change during the experimental periods compared with those of the controls; however, translocation of H. cinaedi from the apical side to the basolateral side was confirmed by cultivation and H. cinaedi-specific PCR, suggesting that the H. cinaedi strains translocated by transcellular route. This study demonstrated that H. cinaedi strains of animal origin might have a pathogenic potential in human epithelial cells as observed in a translocation assay in vitro with a human isolate. PMID:26685883

  2. Improved assay for quantitating adherence of ruminal bacteria to cellulose.

    PubMed Central

    Rasmussen, M A; White, B A; Hespell, R B

    1989-01-01

    A quantitative technique suitable for the determination of adherence of ruminal bacteria to cellulose was developed. This technique employs adherence of cells to cellulose disks and alleviates the problem of nonspecific cell entrapment within cellulose particles. By using this technique, it was demonstrated that the adherence of Ruminococcus flavefaciens FD1 to cellulose was inhibited by formaldehyde, methylcellulose, and carboxymethyl cellulose. Adherence was unaffected by acid hydrolysates of methylcellulose, glucose, and cellobiose. PMID:2782879

  3. relA enhances the adherence of enteropathogenic Escherichia coli.

    PubMed

    Spira, Beny; Ferreira, Gerson Moura; de Almeida, Luiz Gustavo

    2014-01-01

    Enteropathogenic Escherichia coli (EPEC) is a known causative agent of diarrhea in children. In the process of colonization of the small intestine, EPEC synthesizes two types of adhesins, the bundle-forming pilus (BFP) and intimin. The BFP pilus is an adhesin associated with the initial stages of adherence of EPEC to epithelial cells, while the outer membrane protein intimin carries out the intimate adherence that takes place at the third stage of infection. BFP is encoded by the bfp operon located in plasmid EAF, present only in typical EPEC isolates, while eae, the gene that encodes intimin is situated in the LEE, a chromosomal pathogenicity island. Transcription of bfp and eae is regulated by the products of the perABC operon, also present in plasmid EAF. Here we show that deletion of relA, that encodes a guanosine penta and tetraphosphate synthetase impairs EPEC adherence to epithelial cells in vitro. In the absence of relA, the transcription of the regulatory operon perABC is reduced, resulting in lower levels of BFP and intimin. Bacterial adherence, BFP and intimin synthesis and perABC expression are restored upon complementation with the wild-type relA allele. PMID:24643076

  4. Role of Adherence in the Pathogenesis of Pseudomonas aeruginosa Lung Infection in Cystic Fibrosis Patients

    PubMed Central

    Woods, Donald E.; Bass, Joe A.; Johanson, W. G.; Straus, David C.

    1980-01-01

    A correlation has been demonstrated between the in vitro adherence of Pseudomonas aeruginosa to upper respiratory tract epithelium and colonization of the respiratory tract by this organism. Twenty patients with cystic fibrosis (CF) and 20 age-matched controls were examined in this study. All of the CF patients but none of the controls were colonized with P. aeruginosa at the time of study. P. aeruginosa adherence to isolated epithelial cells, as determined by an in vitro assay, was 19.1 ± 1.1 bacteria per buccal epithelial cell in the CF patients and 2.3 ± 0.3 bacteria per cell in the controls (P < 0.01). P. aeruginosa strains of the mucoid colony type adhered in significantly lower numbers to buccal epithelial cells than did strains of the rough colony type (1.8 + 0.1 versus 24.8 ± 0.9, P < 0.001). This difference might explain the common observation that the initial pseudomonas colonization of the respiratory tract of CF patients is due to organisms of the rough colony type. We have further demonstrated that increased P. aeruginosa adherence in vitro varies directly with the loss of a protease-sensitive glycoprotein, fibronectin, from the cell surface, as well as increased levels of salivary proteases in CF patients. When examined by a direct radioimmune binding assay, buccal cells from CF patients possessed only 17% of the total cell surface fibronectin present on similar cells obtained from controls. Salivary protease levels, as measured by 125I release from an 125I-labeled insoluble fibrin matrix, were increased about threefold in CF patients versus controls. Thus, colonization of the respiratory tract by P. aeruginosa in CF patients correlates well with buccal cell adherence of this organism; increased adherence is associated with decreased amounts of fibronectin on respiratory epithelial cell surfaces and increased levels of salivary proteases. PMID:7014444

  5. Regenerative Effects of Mesenchymal Stem Cells: Contribution of Muse Cells, a Novel Pluripotent Stem Cell Type that Resides in Mesenchymal Cells.

    PubMed

    Wakao, Shohei; Kuroda, Yasumasa; Ogura, Fumitaka; Shigemoto, Taeko; Dezawa, Mari

    2012-01-01

    Mesenchymal stem cells (MSCs) are easily accessible and safe for regenerative medicine. MSCs exert trophic, immunomodulatory, anti-apoptotic, and tissue regeneration effects in a variety of tissues and organs, but their entity remains an enigma. Because MSCs are generally harvested from mesenchymal tissues, such as bone marrow, adipose tissue, or umbilical cord as adherent cells, MSCs comprise crude cell populations and are heterogeneous. The specific cells responsible for each effect have not been clarified. The most interesting property of MSCs is that, despite being adult stem cells that belong to the mesenchymal tissue lineage, they are able to differentiate into a broad spectrum of cells beyond the boundary of mesodermal lineage cells into ectodermal or endodermal lineages, and repair tissues. The broad spectrum of differentiation ability and tissue-repairing effects of MSCs might be mediated in part by the presence of a novel pluripotent stem cell type recently found in adult human mesenchymal tissues, termed multilineage-differentiating stress enduring (Muse) cells. Here we review recently updated studies of the regenerative effects of MSCs and discuss their potential in regenerative medicine. PMID:24710542

  6. Regenerative Effects of Mesenchymal Stem Cells: Contribution of Muse Cells, a Novel Pluripotent Stem Cell Type that Resides in Mesenchymal Cells

    PubMed Central

    Wakao, Shohei; Kuroda, Yasumasa; Ogura, Fumitaka; Shigemoto, Taeko; Dezawa, Mari

    2012-01-01

    Mesenchymal stem cells (MSCs) are easily accessible and safe for regenerative medicine. MSCs exert trophic, immunomodulatory, anti-apoptotic, and tissue regeneration effects in a variety of tissues and organs, but their entity remains an enigma. Because MSCs are generally harvested from mesenchymal tissues, such as bone marrow, adipose tissue, or umbilical cord as adherent cells, MSCs comprise crude cell populations and are heterogeneous. The specific cells responsible for each effect have not been clarified. The most interesting property of MSCs is that, despite being adult stem cells that belong to the mesenchymal tissue lineage, they are able to differentiate into a broad spectrum of cells beyond the boundary of mesodermal lineage cells into ectodermal or endodermal lineages, and repair tissues. The broad spectrum of differentiation ability and tissue-repairing effects of MSCs might be mediated in part by the presence of a novel pluripotent stem cell type recently found in adult human mesenchymal tissues, termed multilineage-differentiating stress enduring (Muse) cells. Here we review recently updated studies of the regenerative effects of MSCs and discuss their potential in regenerative medicine. PMID:24710542

  7. SALT-OVERLY SENSITIVE5 Mediates Arabidopsis Seed Coat Mucilage Adherence and Organization through Pectins1[W][OPEN

    PubMed Central

    Griffiths, Jonathan S.; Tsai, Allen Yi-Lun; Xue, Hui; Voiniciuc, Cătălin; Šola, Krešimir; Seifert, Georg J.; Mansfield, Shawn D.; Haughn, George W.

    2014-01-01

    Interactions between cell wall polymers are critical for establishing cell wall integrity and cell-cell adhesion. Here, we exploit the Arabidopsis (Arabidopsis thaliana) seed coat mucilage system to examine cell wall polymer interactions. On hydration, seeds release an adherent mucilage layer strongly attached to the seed in addition to a nonadherent layer that can be removed by gentle agitation. Rhamnogalacturonan I (RG I) is the primary component of adherent mucilage, with homogalacturonan, cellulose, and xyloglucan constituting minor components. Adherent mucilage contains rays composed of cellulose and pectin that extend above the center of each epidermal cell. CELLULOSE SYNTHASE5 (CESA5) and the arabinogalactan protein SALT-OVERLY SENSITIVE5 (SOS5) are required for mucilage adherence through unknown mechanisms. SOS5 has been suggested to mediate adherence by influencing cellulose biosynthesis. We, therefore, investigated the relationship between SOS5 and CESA5. cesa5-1 seeds show reduced cellulose, RG I, and ray size in adherent mucilage. In contrast, sos5-2 seeds have wild-type levels of cellulose but completely lack adherent RG I and rays. Thus, relative to each other, cesa5-1 has a greater effect on cellulose, whereas sos5-2 mainly affects pectin. The double mutant cesa5-1 sos5-2 has a much more severe loss of mucilage adherence, suggesting that SOS5 and CESA5 function independently. Double-mutant analyses with mutations in MUCILAGE MODIFIED2 and FLYING SAUCER1 that reduce mucilage release through pectin modification suggest that only SOS5 influences pectin-mediated adherence. Together, these findings suggest that SOS5 mediates adherence through pectins and does so independently of but in concert with cellulose synthesized by CESA5. PMID:24808103

  8. Cell type-specific bipolar cell input to ganglion cells in the mouse retina.

    PubMed

    Neumann, S; Hüser, L; Ondreka, K; Auler, N; Haverkamp, S

    2016-03-01

    Many distinct ganglion cell types, which are the output elements of the retina, were found to encode for specific features of a visual scene such as contrast, color information or movement. The detailed composition of retinal circuits leading to this tuning of retinal ganglion cells, however, is apart from some prominent examples, largely unknown. Here we aimed to investigate if ganglion cell types in the mouse retina receive selective input from specific bipolar cell types or if they sample their synaptic input non-selectively from all bipolar cell types stratifying within their dendritic tree. To address this question we took an anatomical approach and immunolabeled retinae of two transgenic mouse lines (GFP-O and JAM-B) with markers for ribbon synapses and type 2 bipolar cells. We morphologically identified all green fluorescent protein (GFP)-expressing ganglion cell types, which co-stratified with type 2 bipolar cells and assessed the total number of bipolar input synapses and the proportion of synapses deriving from type 2 bipolar cells. Only JAM-B ganglion cells received synaptic input preferentially from bipolar cell types other than type 2 bipolar cells whereas the other analyzed ganglion cell types sampled their bipolar input most likely from all bipolar cell terminals within their dendritic arbor. PMID:26751712

  9. Parent report of mealtime behaviors in young children with type 1 diabetes mellitus: implications for better assessment of dietary adherence problems in the clinic.

    PubMed

    Patton, Susana R; Dolan, Lawrence M; Powers, Scott W

    2006-06-01

    Parents of children with a chronic illness that has a nutrition treatment component often report mealtime behavior problems. Although research suggests that parents of young children with type 1 diabetes mellitus (T1DM) perceive more mealtime behavior problems than parents of controls, no study has examined the pattern of mealtime behaviors reported by parents of children with T1DM. We examined parents' perceptions of mealtime behaviors of children with T1DM using the Behavioral Pediatric Feeding Assessment Scale (BPFAS). We hypothesized that parents of young children with T1DM would describe similar mealtime problems as has been found in families of children with clinical feeding problems. Data from 85 families of children with T1DM (mean = 5 +/- 1.5 years) were used. Factor analysis for children with T1DM identified a 6-factor solution. Four factors (child refusal, picky eater, stalling, food texture) were similar to factors identified in children with clinical feeding problems. Two unique factors, reflecting strict dietary requirements and intense disruptive behavior, were identified for children with T1DM. Consistent with our hypothesis, we conclude that patterns of mealtime behaviors appear similar for children with T1DM and children with clinical feeding problems. However, for young children with T1DM, unique problems exist, related to a strict feeding schedule consistent with the diabetes diet. Within routine diabetes care, the BPFAS is a valid and clinically useful tool to assess dietary adherence and mealtime behaviors in children. Monitoring via the BPFAS can identify families in need of behavioral interventions to improve mealtime functioning. PMID:16775516

  10. A standard ballroom and Latin dance program to improve fitness and adherence to physical activity in individuals with type 2 diabetes and in obesity

    PubMed Central

    2014-01-01

    Objective To test the effectiveness of a dance program to improve fitness and adherence to physical activity in subjects with type 2 diabetes and obesity. Research Design and Methods Following a motivational interviewing session, 100 subjects with diabetes and/or obesity were enrolled either in a dance program (DP, n = 42) or in a self-selected physical activity program (SSP, n = 58), according to their preferences. Outcome measures were reduced BMI/waist circumference, improved metabolic control in type 2 diabetes (−0.3% reduction of HbA1c) and improved fitness (activity expenditure >10 MET-hour/week; 10% increase in 6-min walk test (6MWT)). Target achievement was tested at 3 and 6 months, after adjustment for baseline data (propensity score). Results Attrition was lower in DP. Both programs significantly decreased body weight (on average, −2.6 kg; P < 0.001) and waist circumference (DP, −3.2 cm; SSP, −2.2; P < 0.01) at 3 months, and the results were maintained at 6 months. In DP, the activity-related energy expenditure averaged 13.5 ± 1.8 MET-hour/week in the first three months and 14.1 ± 3.0 in the second three-month period. In SSP, activity energy expenditure was higher but highly variable in the first three-month period (16.5 ± 13.9 MET-hour/week), and decreased in the following three months (14.2 ± 12.3; P vs. first period < 0.001). At three months, no differences in target achievement were observed between groups. After six months the odds to attain the MET, 6MWT and A1c targets were all significantly associated with DP. Conclusion Dance may be an effective strategy to implement physical activity in motivated subjects with type 2 diabetes or obesity (Clinical trial reg. no.NCT02021890, clinicaltrials.gov). PMID:25045404

  11. Impact of telephonic interviews on persistence and daily adherence to insulin treatment in insulin-naïve type 2 diabetes patients: dropout study

    PubMed Central

    Yavuz, Dilek Gogas; Bilen, Habip; Sancak, Seda; Garip, Tayfun; Hekimsoy, Zeliha; Sahin, Ibrahim; Yilmaz, Murat; Aydin, Hasan; Atmaca, Aysegul; Sert, Murat; Karakaya, Pinar; Arpaci, Dilek; Oguz, Aytekin; Guvener, Nilgun

    2016-01-01

    Objective The objective of this study is to evaluate the impact of sequential telephonic interviews on treatment persistence and daily adherence to insulin injections among insulin-naïve type 2 diabetes patients initiated on different insulin regimens in a 3-month period. Methods A total of 1,456 insulin-naïve patients with type 2 diabetes (mean [standard deviation, SD] age: 56.0 [12.0] years, 49.1% were females) initiated on insulin therapy and consecutively randomized to sequential (n=733) and single (n=723) telephonic interview groups were included. Data on insulin treatment and self-reported blood glucose values were obtained via telephone interview. Logistic regression analysis was performed for factors predicting increased likelihood of persistence and skipping an injection. Results Overall, 76.8% patients (83.2% in sequential vs 70.3% in single interview group, (P<0.001) remained on insulin treatment at the third month. Significantly higher rate for skipping doses was noted in basal bolus than in other regimens (27.0% vs 15.0% for premixed and 15.8% basal insulin, respectively, P<0.0001). Logistic regression analysis revealed sequential telephonic interview (odds ratio [OR], 1.531; 95% confidence interval [CI], 1.093–2.143; P=0.013), higher hemoglobin A1c levels (OR, 1.090; 95% CI, 0.999–1.189; P=0.049), and less negative appraisal of insulin therapy as significant predictors of higher persistence. Basal bolus regimen (OR, 1.583; 95% CI, 1.011–2.479; P=0.045) and higher hemoglobin A1c levels (OR, 1.114; 95% CI, 1.028–1.207; P=0.008) were the significant predictors of increased likelihood of skipping an injection. Conclusion Our findings revealed positive influence of sequential telephonic interview, although including no intervention in treatment, on achieving better treatment persistence in type 2 diabetes patients initiating insulin. PMID:27274207

  12. Hyaluronic Acid-Based Hydrogels as 3D Matrices for in Vitro Evaluation of Chemotherapeutic Drugs Using Poorly Adherent Prostate Cancer Cells

    PubMed Central

    Gurski, Lisa A.; Jha, Amit K.; Zhang, Chu; Jia, Xinqiao; Farach-Carson, Mary C.

    2009-01-01

    The current investigation aimed to develop a biomimetic, three-dimensional (3D) culture system for poorly adherent bone metastatic prostate cancer cells (C4-2B) for use as an in vitro platform for anti-cancer drug screening. To this end, hyaluronic acid (HA) derivatives carrying complementary aldehyde (HAALD) and hydrazide (HAADH) groups were synthesized and characterized. In situ encapsulation of C4-2B cells was achieved by simple mixing of HAALD and HAADH in the presence of the cells. Unlike two-dimensional (2D) monolayer culture in which cells adopt an atypical spread morphology, cells residing in the HA matrix formed distinct clustered structures which grew and merged, reminiscent of real tumors. Anti-cancer drugs added to the media surrounding the cell/gel construct diffused into the gel and killed the embedded cells. The HA hydrogel system was used successfully to test the efficacy of anti-cancer drugs including camptothecin, docetaxel, and rapamycin, alone and in combination, including specificity, dose and time responses. Responses of cells to anti-neoplastics differed between the 3D HA hydrogel and 2D monolayer systems. We suggest that the data obtained from 3D HA systems is superior to that from conventional 2D monolayers as the 3D system better reflects the bone metastatic microenvironment of the cancer cells. PMID:19695694

  13. Genetic relatedness and virulence properties of enteropathogenic Escherichia coli strains of serotype O119:H6 expressing localized adherence or localized and aggregative adherence-like patterns on HeLa cells.

    PubMed

    Garcia, Bruna G; Ooka, Tadasuke; Gotoh, Yasuhiro; Vieira, Mônica A M; Yamamoto, Denise; Ogura, Yoshitoshi; Girão, Dennys M; Sampaio, Suely C F; Melo, Alexis Bonfim; Irino, Kinue; Hayashi, Tetsuya; Gomes, Tânia A T

    2016-05-01

    Enteropathogenic Escherichia coli (EPEC) induce attaching and effacing (A/E) lesions in enterocytes and produce the bundle-forming pilus (BFP) contributing to the localized adherence (LA) pattern formation on HeLa cells. Enteroaggregative E. coli (EAEC) produce aggregative adherence (AA) on HeLa cells and form prominent biofilms. The ability to produce LA or AA is an important hallmark to classify fecal E. coli isolates as EPEC or EAEC, respectively. E. coli strains of serotype O119:H6 exhibit an LA+ phenotype and have been considered as comprising a clonal group of EPEC strains. However, we have recently identified O119:H6 EPEC strains that produce LA and an AA-like pattern concurrently (LA/AA-like+). In this study, we evaluated the relatedness of three LA/AA-like+ and three LA+ O119:H6 strains by comparing their virulence and genotypic properties. We first found that the LA/AA-like+ strains induced actin accumulation in HeLa cells (indicative of A/E lesions formation) and formed biofilms on abiotic surfaces more efficiently than the LA+ strains. MLST analysis showed that the six strains all belong to the ST28 complex. All strains carried multiple plasmids, but as plasmid profiles were highly variable, this cannot be used to differentiate LA/AA-like+ and LA+ strains. We further obtained their draft genome sequences and the complete sequences of four plasmids harbored by one LA/AA-like+ strain. Analysis of these sequences and comparison with 37 fully sequenced E. coli genomes revealed that both O119:H6 groups belong to the E. coli phylogroup B2 and are very closely related with only 58-67 SNPs found between LA/AA-like+ and LA+ strains. Search of the draft sequences of the six strains for adhesion-related genes known in EAEC and other E. coli pathotypes detected no genes specifically present in LA/AA-like+ strains. Unexpectedly however, we found that a large plasmid distinct from pEAF is responsible for the AA-like phenotype of the LA/AA-like+ strains. Although we

  14. Serum Proteins Enhance Dispersion Stability and Influence the Cytotoxicity and Dosimetry of ZnO Nanoparticles in Suspension and Adherent Cancer Cell Models.

    PubMed

    Anders, Catherine B; Chess, Jordan J; Wingett, Denise G; Punnoose, Alex

    2015-12-01

    Agglomeration and sedimentation of nanoparticles (NPs) within biological solutions is a major limitation in their use in many downstream applications. It has been proposed that serum proteins associate with the NP surface to form a protein corona that limits agglomeration and sedimentation. Here, we investigate the effect of fetal bovine serum (FBS) proteins on the dispersion stability, dosimetry, and NP-induced cytotoxicity of cationic zinc oxide nanoparticles (nZnO) synthesized via forced hydrolysis with a core size of 10 nm. Two different in vitro cell culture models, suspension and adherent, were evaluated by comparing a phosphate buffered saline (PBS) nZnO dispersion (nZnO/PBS) and an FBS-stabilized PBS nZnO dispersion (nZnO - FBS/PBS). Surface interactions of FBS on nZnO were analyzed via spectroscopic and optical techniques. Fourier transformed infrared spectroscopy (FTIR) confirmed the adsorption of negatively charged protein components on the cationic nZnO surface through the disappearance of surfaced-adsorbed carboxyl functional groups and the subsequent detection of vibrational modes associated with the protein backbone of FBS-associated proteins. Further confirmation of these interactions was noted in the isoelectric point shift of the nZnO from the characteristic pH of 9.5 to a pH of 6.1. In nZnO - FBS/PBS dispersions, the FBS reduced agglomeration and sedimentation behaviors to impart long-term improvements (>24 h) to the nZnO dispersion stability. Furthermore, mathematical dosimetry models indicate that nZnO - FBS/PBS dispersions had consistent NP deposition patterns over time unlike unstable nZnO/PBS dispersions. In suspension cell models, the stable nZnO - FBS/PBS dispersion resulted in a ~33 % increase in the NP-induced cytotoxicity for both Jurkat leukemic and Hut-78 lymphoma cancer cells. In contrast, the nZnO - FBS/PBS dispersion resulted in 49 and 71 % reductions in the cytotoxicity observed towards the adherent breast (T-47D) and prostate

  15. Serum Proteins Enhance Dispersion Stability and Influence the Cytotoxicity and Dosimetry of ZnO Nanoparticles in Suspension and Adherent Cancer Cell Models

    NASA Astrophysics Data System (ADS)

    Anders, Catherine B.; Chess, Jordan J.; Wingett, Denise G.; Punnoose, Alex

    2015-11-01

    Agglomeration and sedimentation of nanoparticles (NPs) within biological solutions is a major limitation in their use in many downstream applications. It has been proposed that serum proteins associate with the NP surface to form a protein corona that limits agglomeration and sedimentation. Here, we investigate the effect of fetal bovine serum (FBS) proteins on the dispersion stability, dosimetry, and NP-induced cytotoxicity of cationic zinc oxide nanoparticles (nZnO) synthesized via forced hydrolysis with a core size of 10 nm. Two different in vitro cell culture models, suspension and adherent, were evaluated by comparing a phosphate buffered saline (PBS) nZnO dispersion (nZnO/PBS) and an FBS-stabilized PBS nZnO dispersion (nZnO - FBS/PBS). Surface interactions of FBS on nZnO were analyzed via spectroscopic and optical techniques. Fourier transformed infrared spectroscopy (FTIR) confirmed the adsorption of negatively charged protein components on the cationic nZnO surface through the disappearance of surfaced-adsorbed carboxyl functional groups and the subsequent detection of vibrational modes associated with the protein backbone of FBS-associated proteins. Further confirmation of these interactions was noted in the isoelectric point shift of the nZnO from the characteristic pH of 9.5 to a pH of 6.1. In nZnO - FBS/PBS dispersions, the FBS reduced agglomeration and sedimentation behaviors to impart long-term improvements (>24 h) to the nZnO dispersion stability. Furthermore, mathematical dosimetry models indicate that nZnO - FBS/PBS dispersions had consistent NP deposition patterns over time unlike unstable nZnO/PBS dispersions. In suspension cell models, the stable nZnO - FBS/PBS dispersion resulted in a ~33 % increase in the NP-induced cytotoxicity for both Jurkat leukemic and Hut-78 lymphoma cancer cells. In contrast, the nZnO - FBS/PBS dispersion resulted in 49 and 71 % reductions in the cytotoxicity observed towards the adherent breast (T-47D) and prostate

  16. Defining cell types and states with single-cell genomics

    PubMed Central

    Trapnell, Cole

    2015-01-01

    A revolution in cellular measurement technology is under way: For the first time, we have the ability to monitor global gene regulation in thousands of individual cells in a single experiment. Such experiments will allow us to discover new cell types and states and trace their developmental origins. They overcome fundamental limitations inherent in measurements of bulk cell population that have frustrated efforts to resolve cellular states. Single-cell genomics and proteomics enable not only precise characterization of cell state, but also provide a stunningly high-resolution view of transitions between states. These measurements may finally make explicit the metaphor that C.H. Waddington posed nearly 60 years ago to explain cellular plasticity: Cells are residents of a vast “landscape” of possible states, over which they travel during development and in disease. Single-cell technology helps not only locate cells on this landscape, but illuminates the molecular mechanisms that shape the landscape itself. However, single-cell genomics is a field in its infancy, with many experimental and computational advances needed to fully realize its full potential. PMID:26430159

  17. Portrait of Candida albicans Adherence Regulators

    PubMed Central

    Finkel, Jonathan S.; Xu, Wenjie; Huang, David; Hill, Elizabeth M.; Desai, Jigar V.; Woolford, Carol A.; Nett, Jeniel E.; Taff, Heather; Norice, Carmelle T.; Andes, David R.; Lanni, Frederick; Mitchell, Aaron P.

    2012-01-01

    Cell-substrate adherence is a fundamental property of microorganisms that enables them to exist in biofilms. Our study focuses on adherence of the fungal pathogen Candida albicans to one substrate, silicone, that is relevant to device-associated infection. We conducted a mutant screen with a quantitative flow-cell assay to identify thirty transcription factors that are required for adherence. We then combined nanoString gene expression profiling with functional analysis to elucidate relationships among these transcription factors, with two major goals: to extend our understanding of transcription factors previously known to govern adherence or biofilm formation, and to gain insight into the many transcription factors we identified that were relatively uncharacterized, particularly in the context of adherence or cell surface biogenesis. With regard to the first goal, we have discovered a role for biofilm regulator Bcr1 in adherence, and found that biofilm regulator Ace2 is a major functional target of chromatin remodeling factor Snf5. In addition, Bcr1 and Ace2 share several target genes, pointing to a new connection between them. With regard to the second goal, our findings reveal existence of a large regulatory network that connects eleven adherence regulators, the zinc-response regulator Zap1, and approximately one quarter of the predicted cell surface protein genes in this organism. This limited yet sensitive glimpse of mutant gene expression changes had thus defined one of the broadest cell surface regulatory networks in C. albicans. PMID:22359502

  18. Epithelial cell invasion and adherence directed by the enterotoxigenic Escherichia coli tib locus is associated with a 104-kilodalton outer membrane protein.

    PubMed Central

    Elsinghorst, E A; Weitz, J A

    1994-01-01

    Enterotoxigenic Escherichia coli (ETEC) is capable of invading epithelial cell lines derived from the human colon and ileocecum. Two separate loci (tia and tib) that direct noninvasive E. coli HB101 to adhere to and invade intestinal epithelial cells have previously been cosmid cloned from ETEC H10407. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of cellular fractions from tib-positive HB101 shows that the tib locus directs the synthesis of a 104-kDa outer membrane protein (the TibA protein). The tib locus was subcloned to a maximum of 6.7 kb and mutagenized with transposon Tn5. Production of TibA was directly correlated with the capacity of the subclones and Tn5 mutants to invade and adhere to epithelial cells, suggesting that TibA was required for these phenotypes. The position and direction of transcription of the tibA gene were identified by complementation and in vivo T7 RNA polymerase-promoter induction experiments. The role of the tib locus in epithelial cell invasion was confirmed by the construction of chromosomal deletion derivatives in H10407. These deletion mutants invaded epithelial cells at about 15% of the parental level and were fully complemented by plasmids bearing the tib locus. The size and function of the TibA protein are similar to those of invasin from Yersinia pseudotuberculosis (103 kDa). However, a tib probe did not hybridize with the gene encoding invasin. Hybridization analyses of genomic DNA from a wide variety of pathogenic and nonpathogenic bacteria, including Salmonella, Shigella, Yersinia, and Escherichia species, indicate that the tib locus is unique to specific ETEC strains. Images PMID:8039917

  19. Epithelial cell invasion and adherence directed by the enterotoxigenic Escherichia coli tib locus is associated with a 104-kilodalton outer membrane protein.

    PubMed

    Elsinghorst, E A; Weitz, J A

    1994-08-01

    Enterotoxigenic Escherichia coli (ETEC) is capable of invading epithelial cell lines derived from the human colon and ileocecum. Two separate loci (tia and tib) that direct noninvasive E. coli HB101 to adhere to and invade intestinal epithelial cells have previously been cosmid cloned from ETEC H10407. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of cellular fractions from tib-positive HB101 shows that the tib locus directs the synthesis of a 104-kDa outer membrane protein (the TibA protein). The tib locus was subcloned to a maximum of 6.7 kb and mutagenized with transposon Tn5. Production of TibA was directly correlated with the capacity of the subclones and Tn5 mutants to invade and adhere to epithelial cells, suggesting that TibA was required for these phenotypes. The position and direction of transcription of the tibA gene were identified by complementation and in vivo T7 RNA polymerase-promoter induction experiments. The role of the tib locus in epithelial cell invasion was confirmed by the construction of chromosomal deletion derivatives in H10407. These deletion mutants invaded epithelial cells at about 15% of the parental level and were fully complemented by plasmids bearing the tib locus. The size and function of the TibA protein are similar to those of invasin from Yersinia pseudotuberculosis (103 kDa). However, a tib probe did not hybridize with the gene encoding invasin. Hybridization analyses of genomic DNA from a wide variety of pathogenic and nonpathogenic bacteria, including Salmonella, Shigella, Yersinia, and Escherichia species, indicate that the tib locus is unique to specific ETEC strains. PMID:8039917

  20. Capacity of different cell types to stimulate cytotoxic T lymphocyte precursor cells in the presence of interleukin 2.

    PubMed

    Dröge, W; Moyers, C; Wehrmaker, A; Schmidt, H; Panknin, S; Männel, D; Falk, W

    1984-06-01

    Plastic-adherent cells enriched for dendritic cells (AC) were found to be among the most potent stimulator cells for the activation of cytotoxic T lymphocytes (CTL) in vitro in the presence of interleukin 2 (IL 2) and a constant second set of allogeneic stimulator cells. Concanavalin A-activated nylon wool-nonadherent spleen cells ( CNWT ), concanavalin A-activated unfractionated spleen cells ( Cspl ), and some variants of the ESb T lymphoma line were equally effective as stimulator cells, however, and provoked a substantial cytotoxic response at concentrations of 10(4) cells per culture or less. In contrast, nonactivated nylon wool-nonadherent spleen cells ( NWT ) or unfractionated spleen cells (Spl) and cells of the P815 mastocytoma, the Meth A fibrosarcoma, and the T cell lymphomas Ly 5178 Eb and ESb did not stimulate cytotoxic responses at these cell concentrations. The strong stimulatory potential of the Cspl preparation was reduced by treatment with anti-Thy-1 antibody plus complement, whereas the stimulatory activity of the AC preparation was resistant to this treatment. All cell types tested expressed class I major histocompatibility antigens. Nonactivated NWT cells, in contrast to the CNWT preparation, showed no detectable staining with anti-I-E or anti-I-A antibodies and also a slightly weaker staining with class I antisera. Experiments with the tumor cell lines revealed, however, that there was no strict correlation between stimulatory potential and density of class I alloantigens or the expression of I-E determinants. Experiments on primary cytotoxic responses in vivo gave similar results. Experiments in cultures with a single set of stimulator cells and I region-compatible responder cells indicated that AC and Cspl or CNWT also have a markedly stronger capacity than NWT to induce IL 2-dependent DNA synthesis. PMID:6233360

  1. Soft-focused extracorporeal shock waves increase the expression of tendon-specific markers and the release of anti-inflammatory cytokines in an adherent culture model of primary human tendon cells.

    PubMed

    de Girolamo, Laura; Stanco, Deborah; Galliera, Emanuela; Viganò, Marco; Lovati, Arianna Barbara; Marazzi, Monica Gioia; Romeo, Pietro; Sansone, Valerio

    2014-06-01

    Focused extracorporeal shock waves have been found to upregulate the expression of collagen and to initiate cell proliferation in healthy tenocytes and to positively affect the metabolism of tendons, promoting the healing process. Recently, soft-focused extracorporeal shock waves have also been found to have a significant effect on tissue regeneration. However, very few in vitro reports have dealt with the application of this type of shock wave to cells, and in particular, no previous studies have investigated the response of tendon cells to this impulse. We devised an original model to investigate the in vitro effects of soft-focused shock waves on a heterogeneous population of human resident tendon cells in adherent monolayer culture. Our results indicate that soft-focused extracorporeal shock wave treatment (0.17 mJ/mm(2)) is able to induce positive modulation of cell viability, proliferation and tendon-specific marker expression, as well as release of anti-inflammatory cytokines. This could prefigure a new rationale for routine employment of soft-focused shock waves to treat the failed healing status that distinguishes tendinopathies. PMID:24631378

  2. Single cell transcriptional analysis reveals novel innate immune cell types.

    PubMed

    Kippner, Linda E; Kim, Jinhee; Gibson, Greg; Kemp, Melissa L

    2014-01-01

    Single-cell analysis has the potential to provide us with a host of new knowledge about biological systems, but it comes with the challenge of correctly interpreting the biological information. While emerging techniques have made it possible to measure inter-cellular variability at the transcriptome level, no consensus yet exists on the most appropriate method of data analysis of such single cell data. Methods for analysis of transcriptional data at the population level are well established but are not well suited to single cell analysis due to their dependence on population averages. In order to address this question, we have systematically tested combinations of methods for primary data analysis on single cell transcription data generated from two types of primary immune cells, neutrophils and T lymphocytes. Cells were obtained from healthy individuals, and single cell transcript expression data was obtained by a combination of single cell sorting and nanoscale quantitative real time PCR (qRT-PCR) for markers of cell type, intracellular signaling, and immune functionality. Gene expression analysis was focused on hierarchical clustering to determine the existence of cellular subgroups within the populations. Nine combinations of criteria for data exclusion and normalization were tested and evaluated. Bimodality in gene expression indicated the presence of cellular subgroups which were also revealed by data clustering. We observed evidence for two clearly defined cellular subtypes in the neutrophil populations and at least two in the T lymphocyte populations. When normalizing the data by different methods, we observed varying outcomes with corresponding interpretations of the biological characteristics of the cell populations. Normalization of the data by linear standardization taking into account technical effects such as plate effects, resulted in interpretations that most closely matched biological expectations. Single cell transcription profiling provides

  3. Association of Weight Loss and Medication Adherence Among Adults With Type 2 Diabetes Mellitus: SHIELD (Study to Help Improve Early evaluation and management of risk factors Leading to Diabetes)☆

    PubMed Central

    Grandy, Susan; Fox, Kathleen M.; Hardy, Elise

    2013-01-01

    Background Adherence to prescribed diabetes medications is suboptimal, which can lead to poor glycemic control and diabetic complications. Treatment-related weight gain is a side effect of some oral antidiabetic agents and insulin, which may negatively affect adherence to therapy. Objective This study investigated whether adults with type 2 diabetes mellitus (T2DM) who lost weight had better medication adherence than those who gained weight. Methods Weight change over 1 year (2007 to 2008) was assessed among respondents in the US Study to Help Improve Early evaluation and management of risk factors Leading to Diabetes (SHIELD). Weight loss of >1.0%, ≥3%, and ≥5% of weight was compared with weight gain of ≥1.0%. Medication adherence was assessed using the Morisky 4-item questionnaire for medication-taking behavior, with lower scores representing better adherence. Results There were 746 T2DM respondents who lost >1.0%, 483 who lost ≥3%, 310 who lost ≥5%, and 670 who gained ≥1.0% of weight. Each weight-loss group had significantly lower Morisky scores than the weight-gain group; mean scores of 0.389 versus 0.473 (P = 0.050) for the >1.0% weight-loss group, 0.365 versus 0.473 (P = 0.026) for the ≥3% weight-loss group, and 0.334 versus 0.473 (P = 0.014) for the ≥5% weight-loss group. Significantly fewer respondents who lost weight had received insulin, sulfonylurea, or thiazolidinedione therapy (57%) compared with respondents who gained weight (64%) (P = 0.002). Demographics, exercise habits, and dieting were similar between weight-loss and weight-gain groups. Conclusions T2DM respondents with weight loss had significantly better medication adherence and were less likely to be on treatment regimens that increase weight than T2DM respondents with weight gain. These findings suggest that strategies that lead to weight loss, including use of diabetes medications associated with weight loss, may improve medication adherence. PMID:24465048

  4. Participation of carnitine palmitoyltransferase in the synthesis of dipalmitoylphosphatidylcholine in rat alveolar type II cells.

    PubMed

    Arduini, A; Zibellini, G; Ferrari, L; Magnanimi, L; Dottori, S; Lohninger, A; Carminati, P

    2001-02-01

    We have investigated the role of carnitine palmitoyltransferase (EC 2.3.1.21) in pulmonar type II pneumocyte, a lung cell responsible for the synthesis of surface active lipids. Adult type II pneumocytes were isolated from rat lung and purified by differential adherence. When these lung cells were incubated with radioactive palmitate, the percentage of radioactivity recovered into dipalmitoylphosphatidylcholine (DPPC), a major surface active lipid, was almost 60% with respect to total phosphatidylcholine (PC) molecular species. Cellular lysates from type II pneumocytes contained detectable amount of carnitine palmitoyltransferase (CPT) activity (1 nmol/min/mg). Most of the CPT activity found in these cells could be inhibited by incubating them for 60 min with 5 microM tetradecylglycidic acid (TDGA), a specific and irreversible CPT inhibitor of the malonyl-CoA sensitive CPT isoform (CPT I). TDGA treatment of adult type II pneumocytes caused a significant reduction in the incorporation of radioactive palmitate into PC, though this effect did not seem to be specific for DPPC. TDGA affected the incorporation of radioactive palmitate at the sn2 rather than the sn1 position of the glycerol backbone of PC. The incorporation of radioactive palmitate into DPPC was also observed when these lung cells were incubated with palmitate-labeled palmitoyl-L-carnitine. Our data suggest that type II pneumocyte CPT may play an important role in remodelling PC fatty acid composition and hence DPPC synthesis. PMID:11330841

  5. Integration of Provider, Pharmacy, and Patient-Reported Data to Improve Medication Adherence for Type 2 Diabetes: A Controlled Before-After Pilot Study

    PubMed Central

    Alzeer, Abdullah H; Phillips, Erin O'Kelly; Marrero, David G

    2016-01-01

    Background Patients with diabetes often have poor adherence to using medications as prescribed. The reasons why, however, are not well understood. Furthermore, most health care delivery processes do not routinely assess medication adherence or the factors that contribute to poor adherence. Objective The objective of the study was to assess the feasibility of an integrated informatics approach to aggregating and displaying clinically relevant data with the potential to identify issues that may interfere with appropriate medication utilization and facilitate patient-provider communication during clinical encounters about strategies to improve medication use. Methods We developed a clinical dashboard within an electronic health record (EHR) system that uses data from three sources: the medical record, pharmacy claims, and a patient portal. Next, we implemented the dashboard into three community health centers. Health care providers (n=15) and patients with diabetes (n=96) were enrolled in a before-after pilot to test the system’s impact on medication adherence and clinical outcomes. To measure adherence, we calculated the proportion of days covered using pharmacy claims. Demographic, laboratory, and visit data from the EHR were analyzed using pairwise t tests. Perceived barriers to adherence were self-reported by patients. Providers were surveyed about their use and perceptions of the clinical dashboard. Results Adherence significantly and meaningfully improved (improvements ranged from 6%-20%) consistently across diabetes as well as cardiovascular drug classes. Clinical outcomes, including HbA1c, blood pressure, lipid control, and emergency department utilization remained unchanged. Only a quarter of patients (n=24) logged into the patient portal and completed psychosocial questionnaires about their barriers to taking medications. Conclusions Integrated approaches using advanced EHR, clinical decision support, and patient-controlled technologies show promise for

  6. NbHSWP11, a microsporidia Nosema bombycis protein, localizing in the spore wall and membranes, reduces spore adherence to host cell BME.

    PubMed

    Yang, Donglin; Dang, Xiaoqun; Peng, Pai; Long, Mengxian; Ma, Cheng; Qin, Junjie Jia Guowei; Wu, Haijing; Liu, Tie; Zhou, Xiaowei; Pan, Guoqing; Zhou, Zeyang

    2014-10-01

    Microsporidia are obligate intracellular parasites, and a derivative of fungi, which harbor a rigid spore wall to resist adverse environmental pressures. The spore wall protein, which is thought to be the first and direct protein interacting with the host cell, may play a key role in the process of microsporidia infection. In this study, we report a protein, NbHSWP11, with a dnaJ domain. The protein also has 6 heparin-binding motifs which are known to interact with extracellular glycosaminoglycans. Syntenic analysis indicated that gene loci of Nbhswp11 are conserved and syntenic between Nosema bombycis and Nosema ceranae. Phylogenetic tree analysis showed that Nbhswp11 clusters with fungal dnaJ proteins and has 98% identity with an N. bombycis dnaJ protein. Nbhswp11 was transcribed throughout the entire life stages, and gradually increased during 1-7 days, in a silkworm that was infected by N. bombycis, as determined by reverse-transcription PCR (RT-PCR). The recombinant protein NbHSWP11 (rSWP11-HIS) was obtained and purified using gene cloning and prokaryotic expression. Western blotting analysis displayed NbHSWP11 expressed in the total mature spore proteins and spore coat proteins. Indirect immunofluorescence assay revealed NbHSWP11 located at the spore wall of mature spores and the spore coats. Furthermore, immune electron microscopy showed that NbHSWP11 localized in the cytoplasm of the sporont. Within the developmental process of N. bombycis, a portion of NbHSWP11 is targeted to the spore wall of sporoblasts and mature spores. However, most of NbHSWP11 distributes on the membraneous structures of the sporoblast and mature spore. In addition, using a host cell binding assay, native protein NbHSWP11 in the supernatant of total soluble mature spore proteins is shown to bind to the host cell BmE surface. Finally, an antibody blocking assay showed that purified rabbit antibody of NbHSWP11 inhibits spore adherence and decreases the adherence rate of spores by 20

  7. Escherichia coli O157 adherence to the bovine recto-anal junction (RAJ) squamous epithelial cells is mediated by adhesins other than those encoded by genes on the Locus of Enterocyte Attachment (LEE) pathogenicity island

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Escherichia coli O157 (O157) persist in the gastrointestinal tracts (GIT) of bovine reservoirs primarily by adhering to the mucocutaneous, recto-anal junction (RAJ), comprising of both follicle-associated-epithelial (FAE) cells proximally and stratified squamous epithelial (RSE) cells distally. Whe...

  8. IgG1 antimycobacterial antibodies can reverse the inhibitory effect of pentoxifylline on tumour necrosis factor alpha (TNF-α) secreted by mycobacterial antigen-stimulated adherent cells

    PubMed Central

    THAKURDAS, S M; HASAN, Z; HUSSAIN, R

    2004-01-01

    Chronic inflammation associated with cachexia, weight loss, fever and arthralgia is the hallmark of advanced mycobacterial diseases. These symptoms are attributed to the chronic stimulation of tumour necrosis factor (TNF)-α. Mycobacterial components directly stimulate adherent cells to secrete TNF-α. We have shown recently that IgG1 antimycobacterial antibodies play a role in augmenting TNF-α in purified protein derivative (PPD)-stimulated adherent cells from non-BCG-vaccinated donors. We now show that IgG1 antibodies can also augment TNF-α expression in stimulated adherent cells obtained from BCG-vaccinated donors and this augmentation is not linked to interleukin (IL)-10 secretion. In addition IgG1 antimycobacterial antibodies can reverse the effect of TNF-α blockers such as pentoxifylline and thalidomide. These studies therefore have clinical implications for anti-inflammatory drug treatments which are used increasingly to alleviate symptoms associated with chronic inflammation. PMID:15086397

  9. A YadA-like autotransporter, Hag 1, in Veillonella atypica is a Multivalent Hemagglutinin Involved in Adherence to Oral Streptococci, Porphyromonas gingivalis, and Human Oral Buccal Cells

    PubMed Central

    Merritt, Justin; Qi, Fengxia

    2015-01-01

    Dental biofilm development is a sequential process, and adherence between microbes and the salivary pellicle (adhesion) as well as among different microbes (co-adhesion or coaggregation) plays a critical role in building a biofilm community. The Veillonella species are among the most predominant species in the oral cavity and coaggregate with many initial, early, middle and late colonizers. Similar to oral fusobacteria, they are also considered bridging species in biofilm development. However, the mechanism of this ability has yet to be reported, due to the previous lack of a genetic transformation system in the entire genus. In this study, we used our recently discovered transformable Veillonella strain, V. atypica OK5, to probe the mechanism of coaggregation between Veillonella species and other oral bacteria. By insertional inactivation of all 8 putative hemagglutinin genes, we identified one gene, hag1, which is involved in V. atypica coaggregation with the initial colonizers Streptococcus gordonii, Streptococcus oralis and Streptococcus cristatus, and the periodontal pathogen Porphyromonas gingivalis. The hag1 mutant also abolished adherence to human buccal cells. Inhibition assays using various chemical or physiological treatments suggest different mechanisms being involved in coaggregation with different partners. The entire hag1 gene was sequenced and shown to be the largest known bacterial hemagglutinin gene. PMID:25440509

  10. Inhibition of Shigella sonnei adherence to HT-29 cells by lactobacilli from Chinese fermented food and preliminary characterization of S-layer protein involvement.

    PubMed

    Zhang, Ying-Chun; Zhang, Lan-Wei; Tuo, Yan-Feng; Guo, Chun-Feng; Yi, Hua-Xi; Li, Jing-Yan; Han, Xue; Du, Ming

    2010-10-01

    In this study, seven lactobacilli with a high degree of antagonistic a