Defining the role of syndecan-4 in mechanotransduction using surface-modification approaches

PubMed Central

Bellin, Robert M.; Kubicek, James D.; Frigault, Matthew J.; Kamien, Andrew J.; Steward, Robert L.; Barnes, Hillary M.; DiGiacomo, Michael B.; Duncan, Luke J.; Edgerly, Christina K.; Morse, Elizabeth M.; Park, Chan Young; Fredberg, Jeffrey J.; Cheng, Chao-Min; LeDuc, Philip R.

2009-01-01

The ability of cells to respond to external mechanical stimulation is a complex and robust process involving a diversity of molecular interactions. Although mechanotransduction has been heavily studied, many questions remain regarding the link between physical stimulation and biochemical response. Of significant interest has been the contribution of the transmembrane proteins involved, and integrins in particular, because of their connectivity to both the extracellular matrix and the cytoskeleton. Here, we demonstrate the existence of a mechanically based initiation molecule, syndecan-4. We first demonstrate the ability of syndecan-4 molecules to support cell attachment and spreading without the direct extracellular binding of integrins. We also examine the distribution of focal adhesion-associated proteins through controlling surface interactions of beads with molecular specificity in binding to living cells. Furthermore, after adhering cells to elastomeric membranes via syndecan-4-specific attachments we mechanically strained the cells via our mechanical stimulation and polymer surface chemical modification approach. We found ERK phosphorylation similar to that shown for mechanotransductive response for integrin-based cell attachments through our elastomeric membrane-based approach and optical magnetic twisting cytometry for syndecan-4. Finally, through the use of cytoskeletal disruption agents, this mechanical signaling was shown to be actin cytoskeleton dependent. We believe that these results will be of interest to a wide range of fields, including mechanotransduction, syndecan biology, and cell–material interactions. PMID:20080785

Potential mechanisms for the effects of tea extracts on the attachment, biofilm formation and cell size of Streptococcus mutans.

PubMed

Wang, Yi; Lee, Sui M; Dykes, Gary A

2013-01-01

Tea can inhibit the attachment of Streptococcus mutans to surfaces and subsequent biofilm formation. Five commercial tea extracts were screened for their ability to inhibit attachment and biofilm formation by two strains of S. mutans on glass and hydroxyapatite surfaces. The mechanisms of these effects were investigated using scanning electron microscopy (SEM) and phytochemical screening. The results indicated that extracts of oolong tea most effectively inhibited attachment and extracts of pu-erh tea most effectively inhibited biofilm formation. SEM images showed that the S. mutans cells treated with extracts of oolong tea, or grown in medium containing extracts of pu-erh tea, were coated with tea components and were larger with more rounded shapes. The coatings on the cells consisted of flavonoids, tannins and indolic compounds. The ratio of tannins to simple phenolics in each of the coating samples was ∼3:1. This study suggests potential mechanisms by which tea components may inhibit the attachment and subsequent biofilm formation of S. mutans on tooth surfaces, such as modification of cell surface properties and blocking of the activity of proteins and the structures used by the bacteria to interact with surfaces.

In vitro study of antibiotic effect on bacterial adherence to acrylic intraocular lenses.

PubMed

Gaál, Valéria; Kilár, Ferenc; Acs, Barnabás; Szijjártó, Zsuzsanna; Kocsis, Béla; Kustos, Ildikó

2005-11-10

Implantation of artificial intraocular lenses into the eye during ophthalmic surgical procedures ensures an unliving surface on which bacterial pathogens may attach and form biofilms. Despite antibiotic treatment bacteria growing in biofilms might cause inflammation and serious complications. In this study the adhesive ability of 7 Staphylococcus aureus and 11 coagulase-negative Staphylococcus (CNS) strains to the surface of acrylic intraocular lenses had been examined by the ultrasonic method. In untreated cases adhesion of the S. aureus and CNS strains did not differ significantly. We could not demonstrate significant differences between the adhesive ability of the standard strains and the clinical isolates. In this study a single--60 min long--antibiotic (ciprofloxacin and tobramycin) treatment had been applied, that correlate well with the single or intermittant antibiotic prophylaxis of patients. Ciprofloxacin administration was able to reduce significantly the number of attached cells on the surface of acrylic lenses both in the case of S. aureus and CNS strains. Dependence of the effect from concentration could also be demonstrated. Tobramycin treatment was able to inhibit significantly the attachment of S. aureus cells. Despite the debate on antibiotic prophylaxis we presented in our experiments that a single antibiotic administration can decrease the attachment of bacterial cells to the surface of acrylic intraocular lenses, and might be effective in the prevention of postoperative endophthalmitis, that is a rare but serious complication of ophthalmic surgery.

The effect of MTAD, an endodontic irrigant, on fibroblast attachment to periodontally affected root surfaces: A SEM analysis

PubMed Central

Ghandi, Mostafa; Houshmand, Behzad; Nekoofar, Mohammad H.; Tabor, Rachel K.; Yadeghari, Zahra; Dummer, Paul M. H.

2013-01-01

Background: Root surface debridement (RSD) is necessary to create an environment suitable for reattachment of the periodontium. Root surface conditioning may aid the formation of a biocompatible surface suitable for cell reattachment. BioPure™ MTAD (mixture of Doxycycline, citric acid and a detergent) is an endodontic irrigant with antibacterial properties and the ability to remove smear layer. It was hypothesized that MTAD may be useful for root surface conditioning. The efficacy of MTAD as a conditioner was measured by examining fibroblast attachment to root surfaces. Materials and Methods: Thirty-two specimens of human teeth with advanced periodontal disease were used. The surfaces were root planed until smooth. Half of the specimens were treated with 0.9% saline and the other samples with Biopure MTAD. As a negative control group, five further samples were left unscaled with surface calculus. Human gingival fibroblast cells HGF1-PI1 were cultured and poured over the tooth specimens and incubated. After fixation, the samples were sputter-coated with gold and examined with a SEM. The morphology and number of attached, fixed viable cells were examined. The data was analysed using the Mann-Whitney-U statistical test. Results: There was no significant difference between the numbers of attached cells in the experimental group treated with MTAD and the control group treated with saline. Little or no attached cells were seen in the negative control group. Conclusion: RSD created an environment suitable for cell growth and attachment in a laboratory setting. The use of MTAD did not promote the attachment and growth of cells on the surface of human roots following RSD. PMID:23869124

Affinity Electrophoresis Using Ligands Attached To Polymers

NASA Technical Reports Server (NTRS)

Van Alstine, James M.; Snyder, Robert S.; Harris, J. M.; Brooks, D. E.

1990-01-01

In new technique, reduction of electrophoretic mobilities by addition of polyethylene glycol to ligands increases electrophoretic separabilities. In immuno-affinity electrophoresis, modification of ligands extends specificity of electrophoretic separation to particles having surface electric-charge structures otherwise making them electrophoretically inseparable. Modification of antibodies by polyethylene glycol greatly reduces ability to aggregate while enhancing ability to affect electrophoretic mobilities of cells. In hydrophobic-affinity electrophoresis, addition of polyethylene glycol reduces tendency toward aggregation of cells or macromolecules.

Probiotics inhibit enteropathogenic E. coli adherence in vitro by inducing intestinal mucin gene expression.

PubMed

Mack, D R; Michail, S; Wei, S; McDougall, L; Hollingsworth, M A

1999-04-01

Probiotic agents, live microorganisms with beneficial effects for the host, may offer an alternative to conventional antimicrobials in the treatment and prevention of enteric infections. The probiotic agents Lactobacillus plantarum 299v and Lactobacillus rhamnosus GG quantitatively inhibited the adherence of an attaching and effacing pathogenic Escherichia coli to HT-29 intestinal epithelial cells but did not inhibit adherence to nonintestinal HEp-2 cells. HT-29 cells were grown under conditions that induced high levels of either MUC2 or MUC3 mRNA, but HEp-2 cells expressed only minimal levels of MUC2 and no MUC3 mRNA. Media enriched for MUC2 and MUC3 mucin were added exogenously to binding assays and were shown to be capable of inhibiting enteropathogen adherence to HEp-2 cells. Incubation of L. plantarum 299v with HT-29 cells increased MUC2 and MUC3 mRNA expression levels. From these in vitro studies, we propose the hypothesis that the ability of probiotic agents to inhibit adherence of attaching and effacing organisms to intestinal epithelial cells is mediated through their ability to increase expression of MUC2 and MUC3 intestinal mucins.

Identification of a Population of Epidermal Squamous Cell Carcinoma Cells with Enhanced Potential for Tumor Formation

PubMed Central

Adhikary, Gautam; Grun, Dan; Kerr, Candace; Balasubramanian, Sivaprakasam; Rorke, Ellen A.; Vemuri, Mohan; Boucher, Shayne; Bickenbach, Jackie R.; Hornyak, Thomas; Xu, Wen; Fisher, Matthew L.; Eckert, Richard L.

2013-01-01

Epidermal squamous cell carcinoma is among the most common cancers in humans. These tumors are comprised of phenotypically diverse populations of cells that display varying potential for proliferation and differentiation. An important goal is identifying cells from this population that drive tumor formation. To enrich for tumor-forming cells, cancer cells were grown as spheroids in non-attached conditions. We show that spheroid-selected cells form faster growing and larger tumors in immune-compromised mice as compared to non-selected cells. Moreover, spheroid-selected cells gave rise to tumors following injection of as few as one hundred cells, suggesting these cells have enhanced tumor-forming potential. Cells isolated from spheroid-selected tumors retain an enhanced ability to grow as spheroids when grown in non-attached culture conditions. Thus, these tumor-forming cells retain their phenotype following in vivo passage as tumors. Detailed analysis reveals that spheroid-selected cultures are highly enriched for expression of epidermal stem cell and embryonic stem cell markers, including aldehyde dehydrogenase 1, keratin 15, CD200, keratin 19, Oct4, Bmi-1, Ezh2 and trimethylated histone H3. These studies indicate that a subpopulation of cells that possess stem cell-like properties and express stem cell markers can be derived from human epidermal cancer cells and that these cells display enhanced ability to drive tumor formation. PMID:24376802

MicroRNA miR-30 family regulates non-attachment growth of breast cancer cells

PubMed Central

2013-01-01

Background A subset of breast cancer cells displays increased ability to self-renew and reproduce breast cancer heterogeneity. The characterization of these so-called putative breast tumor-initiating cells (BT-ICs) may open the road for novel therapeutic strategies. As microRNAs (miRNAs) control developmental programs in stem cells, BT-ICs may also rely on specific miRNA profiles for their sustained activity. To explore the notion that miRNAs may have a role in sustaining BT-ICs, we performed a comprehensive profiling of miRNA expression in a model of putative BT-ICs enriched by non-attachment growth conditions. Results We found breast cancer cells grown under non-attachment conditions display a unique pattern of miRNA expression, highlighted by a marked low expression of miR-30 family members relative to parental cells. We further show that miR-30a regulates non-attachment growth. A target screening revealed that miR-30 family redundantly modulates the expression of apoptosis and proliferation-related genes. At least one of these targets, the anti-apoptotic protein AVEN, was able to partially revert the effect of miR-30a overexpression. Finally, overexpression of miR-30a in vivo was associated with reduced breast tumor progression. Conclusions miR30-family regulates the growth of breast cancer cells in non-attachment conditions. This is the first analysis of target prediction in a whole family of microRNAs potentially involved in survival of putative BT-ICs. PMID:23445407

Attachment Capability of Antagonistic Yeast Rhodotorula glutinis to Botrytis cinerea Contributes to Biocontrol Efficacy.

PubMed

Li, Boqiang; Peng, Huaimin; Tian, Shiping

2016-01-01

Rhodotorula glutinis as an antagonism show good biocontrol performance against various post-harvest diseases in fruits. In the present study, strong attachment capability of R. glutinis to spores and hyphae of Botrytis cinerea was observed. Further analysis showed that certain protein components on the yeast cell surface played critical role during the interaction between R. glutinis and B. cinerea. The components mainly distributed at the poles of yeast cells and might contain glycosylation modification, as tunicamycin treated yeast cells lost attachment capability to B. cinerea. To investigate contributions of attachment capability of R. glutinis to its biocontrol efficacy, yeast cells were mutagenized with 3% methane-sulfonic acid ethyl ester (EMS), and a mutant CE4 with stable non-attaching phenotype was obtained. No significant difference was found on colony, cell morphology, reproductive ability, and capsule formation between the mutant and wild-type. However, there was a distinct difference in India ink positive staining patterns between the two strains. Moreover, wild-type strain of R. glutinis showed better performance on inhibiting spore germination and mycelial growth of B. cinerea than CE4 strain when yeast cells and B. cinerea were co-cultured in vitro. In biocontrol assay, both wild-type and CE4 strains showed significant biocontrol efficacy against gray mold caused by B. cinerea in apple fruit, whereas, control effect of CE4 strain was lower than that of wild-type. Our findings provided new evidences that attachment capability of R. glutinis to B. cinerea contributed to its biocontrol efficacy.

Attachment Capability of Antagonistic Yeast Rhodotorula glutinis to Botrytis cinerea Contributes to Biocontrol Efficacy

PubMed Central

Li, Boqiang; Peng, Huaimin; Tian, Shiping

2016-01-01

Rhodotorula glutinis as an antagonism show good biocontrol performance against various post-harvest diseases in fruits. In the present study, strong attachment capability of R. glutinis to spores and hyphae of Botrytis cinerea was observed. Further analysis showed that certain protein components on the yeast cell surface played critical role during the interaction between R. glutinis and B. cinerea. The components mainly distributed at the poles of yeast cells and might contain glycosylation modification, as tunicamycin treated yeast cells lost attachment capability to B. cinerea. To investigate contributions of attachment capability of R. glutinis to its biocontrol efficacy, yeast cells were mutagenized with 3% methane-sulfonic acid ethyl ester (EMS), and a mutant CE4 with stable non-attaching phenotype was obtained. No significant difference was found on colony, cell morphology, reproductive ability, and capsule formation between the mutant and wild-type. However, there was a distinct difference in India ink positive staining patterns between the two strains. Moreover, wild-type strain of R. glutinis showed better performance on inhibiting spore germination and mycelial growth of B. cinerea than CE4 strain when yeast cells and B. cinerea were co-cultured in vitro. In biocontrol assay, both wild-type and CE4 strains showed significant biocontrol efficacy against gray mold caused by B. cinerea in apple fruit, whereas, control effect of CE4 strain was lower than that of wild-type. Our findings provided new evidences that attachment capability of R. glutinis to B. cinerea contributed to its biocontrol efficacy. PMID:27199931

Ion-implanted polytetrafluoroethylene enhances Saccharomyces cerevisiae biofilm formation for improved immobilization

PubMed Central

Tran, Clara T. H.; Kondyurin, Alexey; Hirsh, Stacey L.; McKenzie, David R.; Bilek, Marcela M. M.

2012-01-01

The surface of polytetrafluoroethylene (PTFE) was modified using plasma immersion ion implantation (PIII) with the aim of improving its ability to immobilize yeast. The density of immobilized cells on PIII-treated and -untreated PTFE was compared as a function of incubation time over 24 h. Rehydrated yeast cells attached to the PIII-treated PTFE surface more rapidly, with higher density, and greater attachment strength than on the untreated surface. The immobilized yeast cells were removed mechanically or chemically with sodium hydroxide and the residues left on the surfaces were analysed with Fourier transform infrared spectroscopy-attenuated total reflection (FTIR-ATR) and X-ray photoelectron spectroscopy (XPS). The results revealed that the mechanism of cell attachment on both surfaces differs and a model is presented for each. Rapid attachment on the PIII-treated surface occurs through covalent bonds of cell wall proteins and the radicals on the treated surface. In contrast, on the untreated surface, only physisorbed molecules were found in the residue and lipids were more highly concentrated than proteins. The presence of lipids in the residue was found to be a consequence of damage to the plasma membrane during the rehydration process and the increased cell stress was also apparent by the amount of Hsp12 in the protein residue. The immobilized yeast cells on PIII-treated PTFE were found to be as active as yeast cells in suspension. PMID:22696486

Photodynamic toxicity of hematoporphyrin derivatives to human keratinocytes in culture.

PubMed

Kappus, H; Reinhold, C; Artuc, M

Human keratinocytes in culture were able to take up hematoporphyrin derivatives (HPDs) used during photodynamic chemotherapy of tumors. In the absence of light, HPDs showed no cytotoxic effects to keratinocytes. However, after irradiation with visible light, HPDs induced immediate cytotoxicity as measured by the neutral red uptake assay. On the other hand, cell attachment as measured by protein estimation was not affected. When the cells treated with HPDs and irradiated with light were cultured for a further 72 h, they partially lost their ability to attach to the collagen surface. Most of the cells remaining attached after 72 h were no longer viable following treatment with HPDs and light. All parameters measured depended on the intracellular concentration of HPDs used (7-50 ng/10(5) cells) and the time of irradiation (0-30 min). These results suggest that human keratinocytes are a good model to study cytotoxic effects of photodynamically active drugs. Further, keratinocytes were unable to recover after damage caused by HPDs and light.

Establishment of highly metastatic KRAS mutant lung cancer cell sublines in long-term three-dimensional low attachment cultures

PubMed Central

Nakano, Tomoyuki; Kanai, Yoshihiko; Amano, Yusuke; Yoshimoto, Taichiro; Matsubara, Daisuke; Shibano, Tomoki; Tamura, Tomoko; Oguni, Sachiko; Katashiba, Shizuka; Ito, Takeshi; Murakami, Yoshinori; Fukayama, Masashi; Murakami, Takashi; Endo, Shunsuke; Niki, Toshiro

2017-01-01

Decreased cell-substratum adhesion is crucially involved in metastasis. Previous studies demonstrated that lung cancer with floating cell clusters in histology is more likely to develop metastasis. In the present study, we investigated whether cancer cells in long-term, three-dimensional low attachment cultures acquire high metastatic potential; these cells were then used to examine the mechanisms underlying metastasis. Two KRAS-mutated adenocarcinoma cell lines (A549 and H441) were cultured and selected on ultra-low attachment culture dishes, and the resulting cells were defined as FL (for floating) sublines. Cancer cells were inoculated into NOD/SCID mice via an intracardiac injection, and metastasis was evaluated using luciferase-based imaging and histopathology. In vitro cell growth (in attachment or suspension cultures), migration, and invasion were assayed. A whole genomic analysis was performed to identify key molecular alterations in FL sublines. Upon detachment on low-binding dishes, parental cells initially formed rounded spheroids with limited growth activity. However, over time in cultures, cells gradually formed smaller spheroids that grew slowly, and, after 3–4 months, we obtained FL sublines that regained prominent growth potential in suspension cultures. On ordinary dishes, FL cells reattached and exhibited a more spindle-shaped morphology than parental cells. No marked differences were observed in cell growth with attachment, migration, or invasion between FL sublines and parental cell lines; however, FL cells exhibited markedly increased growth potential under suspended conditions in vitro and stronger metastatic abilities in vivo. A genomic analysis identified epithelial-mesenchymal transition (EMT) and c-Myc amplification in A549-FL and H441-FL cells, respectively, as candidate mechanisms for metastasis. The growth potential of FL cells was markedly inhibited by lentiviral ZEB1 knockdown in A549-FL cells and by the inhibition of c-Myc through lentiviral knockdown or the pharmacological inhibitor JQ1 in H441-FL cells. Long-term three-dimensional low attachment cultures may become a useful method for investigating the mechanisms underlying metastasis mediated by decreased cell-substratum adhesion. PMID:28786996

Iodine susceptibility of pseudomonads grown attached to stainless steel surfaces

NASA Technical Reports Server (NTRS)

Pyle, B. H.; McFeters, G. A.

1990-01-01

Pseudomonads were adapted to grow in phosphate-buffered water and on stainless steel surfaces to study the iodine sensitivity of attached and planktonic cells. Cultures adapted to low nutrient growth were incubated at room temperature in a circulating reactor system with stainless steel coupons to allow biofilm formation on the metal surfaces. In some experiments, the reactor was partially emptied and refilled with buffer at each sampling time to simulate a "fill-and-draw" water system. Biofilms of attached bacteria, resuspended biofilm bacteria, and reactor suspension, were exposed to 1 mg l-1 iodine for 2 min. Attached bacterial populations which established on coupons within 3 to 5 days displayed a significant increase in resistance to iodine. Increased resistance was also observed for resuspended cells from the biofilm and planktonic bacteria in the system suspension. Generally, intact biofilms and resuspended biofilm cells were most resistant, followed by planktonic bacteria and phosphate buffer cultures. Thus, biofilm formation on stainless steel surfaces within water systems can result in significantly increased disinfection resistance of commonly-occurring water-borne bacteria that may enhance their ability to colonise water treatment and distribution systems.

Differential Attachment of Salmonella enterica and Enterohemorrhagic Escherichia coli to Alfalfa, Fenugreek, Lettuce, and Tomato Seeds

PubMed Central

Cui, Yue; Walcott, Ronald

2017-01-01

ABSTRACT Vegetable seeds have the potential to disseminate and transmit foodborne bacterial pathogens. This study was undertaken to assess the abilities of selected Salmonella and enterohemorrhagic Escherichia coli (EHEC) strains to attach to fungicide-treated versus untreated, and intact versus mechanically damaged, seeds of alfalfa, fenugreek, lettuce, and tomato. Surface-sanitized seeds (2 g) were exposed to four individual strains of Salmonella or EHEC at 20°C for 5 h. Contaminated seeds were rinsed twice, each with 10 ml of sterilized water, before being soaked overnight in 5 ml of phosphate-buffered saline at 4°C. The seeds were then vortexed vigorously for 1 min, and pathogen populations in seed rinse water and soaking buffer were determined using a standard plate count assay. In general, the Salmonella cells had higher attachment ratios than the EHEC cells. Lettuce seeds by unit weight had the highest numbers of attached Salmonella or EHEC cells, followed by tomato, alfalfa, and fenugreek seeds. In contrast, individual fenugreek seeds had more attached pathogen cells, followed by lettuce, alfalfa, and tomato seeds. Significantly more Salmonella and EHEC cells attached to mechanically damaged seeds than to intact seeds (P < 0.05). Although, on average, significantly more Salmonella and EHEC cells were recovered from untreated than fungicide-treated seeds (P < 0.05), fungicide treatment did not significantly affect the attachment of individual bacterial strains to vegetable seeds (P > 0.05), with a few exceptions. This study fills gaps in the current body of literature and helps explain bacterial interactions with vegetable seeds with differing surface characteristics. IMPORTANCE Vegetable seeds, specifically sprout seeds, have the potential to disseminate and transmit foodborne bacterial pathogens. This study investigated the interaction between two important bacterial pathogens, i.e., Salmonella and EHEC, and vegetable seeds with differing surface characteristics. This research helps understand whether seed surface structure, integrity, and fungicide treatment affect the interaction between bacterial cells and vegetable seeds. PMID:28130295

Differential Attachment of Salmonella enterica and Enterohemorrhagic Escherichia coli to Alfalfa, Fenugreek, Lettuce, and Tomato Seeds.

PubMed

Cui, Yue; Walcott, Ronald; Chen, Jinru

2017-04-01

Vegetable seeds have the potential to disseminate and transmit foodborne bacterial pathogens. This study was undertaken to assess the abilities of selected Salmonella and enterohemorrhagic Escherichia coli (EHEC) strains to attach to fungicide-treated versus untreated, and intact versus mechanically damaged, seeds of alfalfa, fenugreek, lettuce, and tomato. Surface-sanitized seeds (2 g) were exposed to four individual strains of Salmonella or EHEC at 20°C for 5 h. Contaminated seeds were rinsed twice, each with 10 ml of sterilized water, before being soaked overnight in 5 ml of phosphate-buffered saline at 4°C. The seeds were then vortexed vigorously for 1 min, and pathogen populations in seed rinse water and soaking buffer were determined using a standard plate count assay. In general, the Salmonella cells had higher attachment ratios than the EHEC cells. Lettuce seeds by unit weight had the highest numbers of attached Salmonella or EHEC cells, followed by tomato, alfalfa, and fenugreek seeds. In contrast, individual fenugreek seeds had more attached pathogen cells, followed by lettuce, alfalfa, and tomato seeds. Significantly more Salmonella and EHEC cells attached to mechanically damaged seeds than to intact seeds ( P < 0.05). Although, on average, significantly more Salmonella and EHEC cells were recovered from untreated than fungicide-treated seeds ( P < 0.05), fungicide treatment did not significantly affect the attachment of individual bacterial strains to vegetable seeds ( P > 0.05), with a few exceptions. This study fills gaps in the current body of literature and helps explain bacterial interactions with vegetable seeds with differing surface characteristics. IMPORTANCE Vegetable seeds, specifically sprout seeds, have the potential to disseminate and transmit foodborne bacterial pathogens. This study investigated the interaction between two important bacterial pathogens, i.e., Salmonella and EHEC, and vegetable seeds with differing surface characteristics. This research helps understand whether seed surface structure, integrity, and fungicide treatment affect the interaction between bacterial cells and vegetable seeds. Copyright © 2017 American Society for Microbiology.

Fibroblastic interactions with high-porosity Ti-6Al-4V metal foam.

PubMed

Cheung, Serene; Gauthier, Maxime; Lefebvre, Louis-Philippe; Dunbar, Michael; Filiaggi, Mark

2007-08-01

A novel metallic Ti-6Al-4V foam in development at the National Research Council of Canada was investigated for its ability to foster cell attachment and growth using a fibroblast cell culture model. The foam was manufactured via a powder metallurgical process that could produce interconnected porosity greater than 70%. Cell attachment was assessed after 6 and 24 h, while proliferation was examined after 3 and 7 days. Ingrown fibroblasts displayed a number of different morphologies; some fibroblasts were spread thinly in close apposition with the irregular surface, or more often had several anchorage points and extended in three dimensions as they spanned pore space. It was also demonstrated that fibroblasts were actively migrating through the porous scaffold over a 14-day period. In a 60-day extended culture, fibroblasts were bridging and filling macropores and had extensively infiltrated the foams. Overall, it was established that this foam was supportive of cell attachment and proliferation, migration through the porous network, and that it was capable of sustaining a large cell population.

Monocyte-mediated delivery of polymeric backpacks to inflamed tissues: a generalized strategy to deliver drugs to treat inflammation.

PubMed

Anselmo, Aaron C; Gilbert, Jonathan B; Kumar, Sunny; Gupta, Vivek; Cohen, Robert E; Rubner, Michael F; Mitragotri, Samir

2015-02-10

Targeted delivery of drugs and imaging agents to inflamed tissues, as in the cases of cancer, Alzheimer's disease, Parkinson's disease, and arthritis, represents one of the major challenges in drug delivery. Monocytes possess a unique ability to target and penetrate into sites of inflammation. Here, we describe a broad approach to take advantage of the natural ability of monocytes to target and deliver flat polymeric particles ("Cellular Backpacks") to inflamed tissues. Cellular backpacks attach strongly to the surface of monocytes but do not undergo phagocytosis due to backpack's size, disk-like shape and flexibility. Following attachment of backpacks, monocytes retain important cellular functions including transmigration through an endothelial monolayer and differentiation into macrophages. In two separate in vivo inflammation models, backpack-laden monocytes exhibit increased targeting to inflamed tissues. Cellular backpacks, and their abilities to attach to monocytes without impairing monocyte functions and 'hitchhike' to a variety of inflamed tissues, offer a new platform for both cell-mediated therapies and broad targeting of inflamed tissues. Copyright © 2014 Elsevier B.V. All rights reserved.

Involvement of flocculin in negative potential-applied ITO electrode adhesion of yeast cells

PubMed Central

Koyama, Sumihiro; Tsubouchi, Taishi; Usui, Keiko; Uematsu, Katsuyuki; Tame, Akihiro; Nogi, Yuichi; Ohta, Yukari; Hatada, Yuji; Kato, Chiaki; Miwa, Tetsuya; Toyofuku, Takashi; Nagahama, Takehiko; Konishi, Masaaki; Nagano, Yuriko; Abe, Fumiyoshi

2015-01-01

The purpose of this study was to develop novel methods for attachment and cultivation of specifically positioned single yeast cells on a microelectrode surface with the application of a weak electrical potential. Saccharomyces cerevisiae diploid strains attached to an indium tin oxide/glass (ITO) electrode to which a negative potential between −0.2 and −0.4 V vs. Ag/AgCl was applied, while they did not adhere to a gallium-doped zinc oxide/glass electrode surface. The yeast cells attached to the negative potential-applied ITO electrodes showed normal cell proliferation. We found that the flocculin FLO10 gene-disrupted diploid BY4743 mutant strain (flo10Δ /flo10Δ) almost completely lost the ability to adhere to the negative potential-applied ITO electrode. Our results indicate that the mechanisms of diploid BY4743 S. cerevisiae adhesion involve interaction between the negative potential-applied ITO electrode and the Flo10 protein on the cell wall surface. A combination of micropatterning techniques of living single yeast cell on the ITO electrode and omics technologies holds potential of novel, highly parallelized, microchip-based single-cell analysis that will contribute to new screening concepts and applications. PMID:26187908

Multifunctional polymer-capped mesoporous silica nanoparticles for pH-responsive targeted drug delivery.

PubMed

Niedermayer, Stefan; Weiss, Veronika; Herrmann, Annika; Schmidt, Alexandra; Datz, Stefan; Müller, Katharina; Wagner, Ernst; Bein, Thomas; Bräuchle, Christoph

2015-05-07

A highly stable modular platform, based on the sequential covalent attachment of different functionalities to the surface of core-shell mesoporous silica nanoparticles (MSNs) for targeted drug delivery is presented. A reversible pH-responsive cap system based on covalently attached poly(2-vinylpyridine) (PVP) was developed as drug release mechanism. Our platform offers (i) tuneable interactions and release kinetics with the cargo drug in the mesopores based on chemically orthogonal core-shell design, (ii) an extremely robust and reversible closure and release mechanism based on endosomal acidification of the covalently attached PVP polymer block, (iii) high colloidal stability due to a covalently coupled PEG shell, and (iv) the ability to covalently attach a wide variety of dyes, targeting ligands and other functionalities at the outer periphery of the PEG shell. The functionality of the system was demonstrated in several cell studies, showing pH-triggered release in the endosome, light-triggered endosomal escape with an on-board photosensitizer, and efficient folic acid-based cell targeting.

Stochasticity of bacterial attachment and its predictability by the extended derjaguin-landau-verwey-overbeek theory.

PubMed

Chia, Teck Wah R; Nguyen, Vu Tuan; McMeekin, Thomas; Fegan, Narelle; Dykes, Gary A

2011-06-01

Bacterial attachment onto materials has been suggested to be stochastic by some authors but nonstochastic and based on surface properties by others. We investigated this by attaching pairwise combinations of two Salmonella enterica serovar Sofia (S. Sofia) strains (with different physicochemical and attachment properties) with one strain each of S. enterica serovar Typhimurium, S. enterica serovar Infantis, or S. enterica serovar Virchow (all with similar physicochemical and attachment abilities) in ratios of 0.428, 1, and 2.333 onto glass, stainless steel, Teflon, and polysulfone. Attached bacterial cells were recovered and counted. If the ratio of attached cells of each Salmonella serovar pair recovered was the same as the initial inoculum ratio, the attachment process was deemed stochastic. Experimental outcomes from the study were compared to those predicted by the extended Derjaguin-Landau-Verwey-Overbeek (XDLVO) theory. Significant differences (P < 0.05) between the initial and the attached ratios for serovar pairs containing S. Sofia S1296a for all different ratios were apparent for all materials. For S. Sofia S1635-containing pairs, 7 out of 12 combinations of serovar pairs and materials had attachment ratios not significantly different (P > 0.05) from the initial ratio of 0.428. Five out of 12 and 10 out of 12 samples had attachment ratios not significantly different (P > 0.05) from the initial ratios of 1 and 2.333, respectively. These results demonstrate that bacterial attachment to different materials is likely to be nonstochastic only when the key physicochemical properties of the bacteria were significantly different (P < 0.05) from each other. XDLVO theory could successfully predict the attachment of some individual isolates to particular materials but could not be used to predict the likelihood of stochasticity in pairwise attachment experiments.

Viral Capsid DNA Aptamer Conjugates as Multivalent Cell Targeting Vehicles

PubMed Central

Tong, Gary J.; Hsiao, Sonny C.; Carrico, Zachary M.; Francis, Matthew B.

2009-01-01

Nucleic acid aptamers offer significant potential as convenient and evolvable targeting groups for drug delivery. To attach them to the surface of a genome-free viral capsid carrier, an efficient oxidative coupling strategy has been developed. The method involves the periodate-mediated reaction of phenylene diamine substituted oligonucleotides with aniline groups installed on the outer surface of the capsid shells. Up to 60 DNA strands can be attached to each viral capsid with no apparent loss of base-pairing capabilities or protein stability. The ability of the capsids to bind specific cellular targets was demonstrated through the attachment of a 41-nucleotide sequence that targets a tyrosine kinase receptor on Jurkat T cells. After the installation of a fluorescent dye on the capsid interior, capsids bearing the cell-targeting sequence showed significant levels of binding to the cells relative to control samples. Colocalization experiments using confocal microscopy indicated that the capsids were endocytosed and trafficked to lysosomes for degradation. These observations suggest that aptamer-labeled capsids could be used for the targeted drug delivery of acid-labile prodrugs that would be preferentially released upon lysosomal acidification. PMID:19603808

Bacterial response to different surface chemistries fabricated by plasma polymerization on electrospun nanofibers.

PubMed

Abrigo, Martina; Kingshott, Peter; McArthur, Sally L

2015-12-06

Control over bacterial attachment and proliferation onto nanofibrous materials constitutes a major challenge for a variety of applications, including filtration membranes, protective clothing, wound dressings, and tissue engineering scaffolds. To develop effective devices, the interactions that occur between bacteria and nanofibers with different morphological and physicochemical properties need to be investigated. This paper explores the influence of fiber surface chemistry on bacterial behavior. Different chemical functionalities were generated on the surface of electrospun polystyrene nanofibers through plasma polymerization of four monomers (acrylic acid, allylamine, 1,7-octadiene, and 1,8-cineole). The interactions of Escherichia coli with the surface modified fibers were investigated through a combination of scanning electron microscopy and confocal laser scanning microscopy. Fiber wettability, surface charge, and chemistry were found to affect the ability of bacterial cells to attach and proliferate throughout the nanofiber meshes. The highest proportion of viable cells attachment occurred on the hydrophilic amine rich coating, followed by the hydrophobic octadiene. The acrylic acid coating rich in carboxyl groups showed a significantly lower attraction of bacterial cells. The 1,8-cineole retained the antibacterial activity of the monomer, resulting with a high proportion of dead isolated cells attached onto the fibers. Results showed that the surface chemistry properties of nanofibrous membranes can be strategically tuned to control bacterial behavior.

The use of plasma-activated covalent attachment of early domains of tropoelastin to enhance vascular compatibility of surfaces

PubMed Central

Hiob, Matti A.; Wise, Steven G.; Kondyurin, Alexey; Waterhouse, Anna; Bilek, Marcela M.; Ng, Martin K. C.; Weiss, Anthony S.

2013-01-01

All current metallic vascular prostheses, including stents, exhibit suboptimal biocompatibility. Improving the re-endothelialization and reducing the thrombogenicity of these devices would substantially improve their clinical efficacy. Tropoelastin (TE), the soluble precursor of elastin, mediates favorable endothelial cell interactions while having low thrombogenicity. Here we show that constructs of TE corresponding to the first 10 (“N10”) and first 18 (“N18”) N-terminal domains of the molecule facilitate endothelial cell attachment and proliferation equivalent to the performance of full-length TE. This N-terminal ability contrasts with the known role of the C-terminus of TE in facilitating cell attachment, particularly of fibroblasts. When immobilized on a plasma-activated coating (“PAC”), N10 and N18 retained their bioactivity and endothelial cell interactive properties, demonstrating attachment and proliferation equivalent to full-length TE. In whole blood assays, both N10 and N18 maintained the low thrombogenicity of PAC. Furthermore, these N-terminal constructs displayed far greater resistance to protease degradation by blood serine proteases kallikrein and thrombin than did full-length TE. When immobilized onto a PAC surface, these shorter constructs form a modified metal interface to establish a platform technology for biologically compatible, implantable cardiovascular devices. PMID:23863453

Control of Surface Chemistry, Substrate Stiffness, and Cell Function in a Novel Terpolymer Methacrylate Library

PubMed Central

Joy, Abraham; Cohen, Daniel M.; Luk, Arnold; Anim-Danso, Emmanuel; Chen, Christopher; Kohn, Joachim

2011-01-01

A focused library of methacrylate terpolymers was synthesized to explore the effects of varying surface chemistry and adhesive peptide ligands on cell function. The chemical diversity of methacrylate monomers enabled construction of a library of polymers in which one can systematically vary the chemical composition to achieve a wide range of contact angle, Young's modulus, and Tg values. Furthermore, the materials were designed to allow surface immobilization of bioactive peptides. We then examined the effects of these material compositions on protein adsorption and cell attachment, proliferation, and differentiation. We observed that chemical composition of the polymers was an important determinant for NIH 3T3 cell attachment and proliferation, as well as human mesenchymal stem cell differentiation, and correlated directly with the ability of the polymers to adsorb proteins that mediate cell adhesion. Importantly, functionalization of the methacrylate terpolymer library with an adhesive GRGDS peptide normalized cellular responses. RGD-functionalized polymers uniformly exhibited robust attachment, proliferation, and differentiation irrespective of the underlying substrate chemistry. These studies provide a library-based approach to rapidly explore the biological functionality of biomaterials with a wide range of compositions, and highlights the importance of cell and protein cell adhesion in predicting their performance. PMID:21226505

Measles virus attachment proteins with impaired ability to bind CD46 interact more efficiently with the homologous fusion protein

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

Corey, Elizabeth A.; Iorio, Ronald M.; Program in Immunology and Virology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655

2009-01-05

Fusion promotion by measles virus (MV) depends on an interaction between the hemagglutinin (H) and fusion (F) glycoproteins. Amino acid substitutions in MV H that drastically reduce hemagglutinating activity result in an increase in the amount of H (primarily the 74 kDa isoform) detectable in a complex with F at the cell surface. This is in direct contrast to the loss of the ability to detect a complex between the fusion protein of Newcastle disease virus and most attachment proteins that lack receptor binding activity. These opposing results provide support for the existence of different mechanisms for the regulation ofmore » fusion by these two paramyxoviruses.« less

Histatin 1 Enhances Cell Adhesion to Titanium in an Implant Integration Model.

PubMed

van Dijk, I A; Beker, A F; Jellema, W; Nazmi, K; Wu, G; Wismeijer, D; Krawczyk, P M; Bolscher, J G M; Veerman, E C I; Stap, J

2017-04-01

Cellular adhesion is essential for successful integration of dental implants. Rapid soft tissue integration is important to create a seal around the implant and prevent infections, which commonly cause implant failure and can result in bone loss. In addition, soft tissue management is important to obtain good dental aesthetics. We previously demonstrated that the salivary peptide histatin 1 (Hst1) causes a more than 2-fold increase in the ability of human adherent cells to attach and spread on a glass surface. Cells treated with Hst1 attached more rapidly and firmly to the substrate and to each other. In the current study, we examine the potential application of Hst1 for promotion of dental implant integration. Our results show that Hst1 enhances the attachment and spreading of soft tissue cell types (oral epithelial cells and fibroblasts) to titanium (Ti) and hydroxyapatite (HAP), biomaterials that have found wide applications as implant material in dentistry and orthopedics. For improved visualization of cell adhesion to Ti, we developed a novel technique that uses sputtering to deposit a thin, transparent layer of Ti onto glass slides. This approach allows detailed, high-resolution analysis of cell adherence to Ti in real time. Furthermore, our results suggest that Hst1 has no negative effects on cell survival. Given its natural occurrence in the oral cavity, Hst1 could be an attractive agent for clinical application. Importantly, even though Hst1 is specific for saliva of humans and higher primates, it stimulated the attachment and spreading of canine cells, paving the way for preclinical studies in canine models.

Small molecules targeting LapB protein prevent Listeria attachment to catfish muscle

PubMed Central

Das, Bhaskar; Lawrence, Mark

2017-01-01

Listeria monocytogenes is a Gram-positive foodborne pathogen and the causative agent of listeriosis. L. monocytogenes lapB gene encodes a cell wall surface anchor protein, and mutation of this gene causes Listeria attenuation in mice. In this work, the potential role of Listeria LapB protein in catfish fillet attachment was investigated. To achieve this, boron-based small molecules designed to interfere with the active site of the L. monocytogenes LapB protein were developed, and their ability to prevent L. monocytogenes attachment to fish fillet was tested. Results indicated that seven out of nine different small molecules were effective in reducing the Listeria attachment to catfish fillets. Of these, three small molecules (SM3, SM5, and SM7) were highly effective in blocking Listeria attachment to catfish fillets. This study suggests an alternative strategy for reduction of L. monocytogenes contamination in fresh and frozen fish products. PMID:29253892

The basolateral vesicle sorting machinery and basolateral proteins are recruited to the site of enteropathogenic E. coli microcolony growth at the apical membrane.

PubMed

Pedersen, Gitte A; Jensen, Helene H; Schelde, Anne-Sofie B; Toft, Charlotte; Pedersen, Hans N; Ulrichsen, Maj; Login, Frédéric H; Amieva, Manuel R; Nejsum, Lene N

2017-01-01

Foodborne Enteropathogenic Escherichia coli (EPEC) infections of the small intestine cause diarrhea especially in children and are a major cause of childhood death in developing countries. EPEC infects the apical membrane of the epithelium of the small intestine by attaching, effacing the microvilli under the bacteria and then forming microcolonies on the cell surface. We first asked the question where on epithelial cells EPEC attaches and grows. Using models of polarized epithelial monolayers, we evaluated the sites of initial EPEC attachment to the apical membrane and found that EPEC preferentially attached over the cell-cell junctions and formed microcolonies preferentially where three cells come together at tricellular tight junctions. The ability of EPEC to adhere increased when host cell polarity was compromised yielding EPEC access to basolateral proteins. EPEC pedestals contain basolateral cytoskeletal proteins. Thus, we asked if attached EPEC causes reorganization the protein composition of the host cell plasma membrane at sites of microcolony formation. We found that EPEC microcolony growth at the apical membrane resulted in a local accumulation of basolateral plasma membrane proteins surrounding the microcolony. Basolateral marker protein aquaporin-3 localized to forming EPEC microcolonies. Components of the basolateral vesicle targeting machinery were re-routed. The Exocyst (Exo70) was recruited to individual EPEC as was the basolateral vesicle SNARE VAMP-3. Moreover, several Rab variants were also recruited to the infection site, and their dominant-negative equivalents were not. To quantitatively study the recruitment of basolateral proteins, we created a pulse of the temperature sensitive basolateral VSVG, VSVG3-SP-GFP, from the trans-Golgi Network. We found that after release from the TGN, significantly more VSVG3-SP-GFP accumulated at the site of microcolony growth than on equivalent membrane regions of uninfected cells. This suggests that trafficking of vesicles destined for the basolateral membrane are redirected to the apical site of microcolony growth. Thus, in addition to disrupting host cell fence function, local host cell plasma membrane protein composition is changed by altered protein trafficking and recruitment of basolateral proteins to the apical microcolony. This may aid EPEC attachment and subsequent microcolony growth.

The basolateral vesicle sorting machinery and basolateral proteins are recruited to the site of enteropathogenic E. coli microcolony growth at the apical membrane

PubMed Central

Pedersen, Gitte A.; Jensen, Helene H.; Schelde, Anne-Sofie B.; Toft, Charlotte; Pedersen, Hans N.; Ulrichsen, Maj; Login, Frédéric H.; Amieva, Manuel R.

2017-01-01

Foodborne Enteropathogenic Escherichia coli (EPEC) infections of the small intestine cause diarrhea especially in children and are a major cause of childhood death in developing countries. EPEC infects the apical membrane of the epithelium of the small intestine by attaching, effacing the microvilli under the bacteria and then forming microcolonies on the cell surface. We first asked the question where on epithelial cells EPEC attaches and grows. Using models of polarized epithelial monolayers, we evaluated the sites of initial EPEC attachment to the apical membrane and found that EPEC preferentially attached over the cell-cell junctions and formed microcolonies preferentially where three cells come together at tricellular tight junctions. The ability of EPEC to adhere increased when host cell polarity was compromised yielding EPEC access to basolateral proteins. EPEC pedestals contain basolateral cytoskeletal proteins. Thus, we asked if attached EPEC causes reorganization the protein composition of the host cell plasma membrane at sites of microcolony formation. We found that EPEC microcolony growth at the apical membrane resulted in a local accumulation of basolateral plasma membrane proteins surrounding the microcolony. Basolateral marker protein aquaporin-3 localized to forming EPEC microcolonies. Components of the basolateral vesicle targeting machinery were re-routed. The Exocyst (Exo70) was recruited to individual EPEC as was the basolateral vesicle SNARE VAMP-3. Moreover, several Rab variants were also recruited to the infection site, and their dominant-negative equivalents were not. To quantitatively study the recruitment of basolateral proteins, we created a pulse of the temperature sensitive basolateral VSVG, VSVG3-SP-GFP, from the trans-Golgi Network. We found that after release from the TGN, significantly more VSVG3-SP-GFP accumulated at the site of microcolony growth than on equivalent membrane regions of uninfected cells. This suggests that trafficking of vesicles destined for the basolateral membrane are redirected to the apical site of microcolony growth. Thus, in addition to disrupting host cell fence function, local host cell plasma membrane protein composition is changed by altered protein trafficking and recruitment of basolateral proteins to the apical microcolony. This may aid EPEC attachment and subsequent microcolony growth. PMID:28636623

Covalent binding of nanoliposomes to the surface of magnetotactic bacteria for the synthesis of self-propelled therapeutic agents.

PubMed

Taherkhani, Samira; Mohammadi, Mahmood; Daoud, Jamal; Martel, Sylvain; Tabrizian, Maryam

2014-05-27

The targeted and effective delivery of therapeutic agents remains an unmet goal in the field of controlled release systems. Magnetococcus marinus MC-1 magnetotactic bacteria (MTB) are investigated as potential therapeutic carriers. By combining directional magnetotaxis-microaerophilic control of these self-propelled agents, a larger amount of therapeutics can be delivered surpassing the diffusion limits of large drug molecules toward hard-to-treat hypoxic regions in solid tumors. The potential benefits of these carriers emphasize the need to develop an adequate method to attach therapeutic cargos, such as drug-loaded nanoliposomes, without substantially affecting the cell's ability to act as delivery agents. In this study, we report on a strategy for the attachment of liposomes to MTB (MTB-LP) through carbodiimide chemistry. The attachment efficacy, motility, and magnetic response of the MTB-LP were investigated. Results confirm that a substantial number of nanoliposomes (∼70) are efficiently linked with MTB without compromising functionality and motility. Cytotoxicity assays using three different cell types (J774, NIH/3T3, and Colo205) reveal that liposomal attachments to MTB formulation improve the biocompatibility of MTB, whereas attachment does not interfere with liposomal uptake.

Shear Stress Regulates Adhesion and Rolling of CD44+ Leukemic and Hematopoietic Progenitor Cells on Hyaluronan

PubMed Central

Christophis, Christof; Taubert, Isabel; Meseck, Georg R.; Schubert, Mario; Grunze, Michael; Ho, Anthony D.; Rosenhahn, Axel

2011-01-01

Leukemic cells and human hematopoietic progenitor cells expressing CD44 receptors have the ability to attach and roll on hyaluronan. We investigated quantitatively the adhesion behavior of leukemic cell lines and hematopoietic progenitor cells on thin films of the polysaccharides hyaluronan and alginate in a microfluidic system. An applied flow enhances the interaction between CD44-positive cells and hyaluronan if a threshold shear stress of 0.2 dyn/cm2 is exceeded. At shear stress ∼1 dyn/cm2, the cell rolling speed reaches a maximum of 15 μm/s. Leukemic Jurkat and Kasumi-1 cells lacking CD44-expression showed no adhesion or rolling on the polysaccharides whereas the CD44-expressing leukemic cells KG-1a, HL-60, K-562, and hematopoietic progenitor cells attached and rolled on hyaluronan. Interestingly, the observations of flow-induced cell rolling are related to those found in the recruitment of leukocytes to inflammatory sites and the mechanisms of stem-cell homing into the bone marrow. PMID:21806926

Antibacterial Au nanostructured surfaces

NASA Astrophysics Data System (ADS)

Wu, Songmei; Zuber, Flavia; Brugger, Juergen; Maniura-Weber, Katharina; Ren, Qun

2016-01-01

We present here a technological platform for engineering Au nanotopographies by templated electrodeposition on antibacterial surfaces. Three different types of nanostructures were fabricated: nanopillars, nanorings and nanonuggets. The nanopillars are the basic structures and are 50 nm in diameter and 100 nm in height. Particular arrangement of the nanopillars in various geometries formed nanorings and nanonuggets. Flat surfaces, rough substrate surfaces, and various nanostructured surfaces were compared for their abilities to attach and kill bacterial cells. Methicillin-resistant Staphylococcus aureus, a Gram-positive bacterial strain responsible for many infections in health care system, was used as the model bacterial strain. It was found that all the Au nanostructures, regardless their shapes, exhibited similar excellent antibacterial properties. A comparison of live cells attached to nanotopographic surfaces showed that the number of live S. aureus cells was <1% of that from flat and rough reference surfaces. Our micro/nanofabrication process is a scalable approach based on cost-efficient self-organization and provides potential for further developing functional surfaces to study the behavior of microbes on nanoscale topographies.We present here a technological platform for engineering Au nanotopographies by templated electrodeposition on antibacterial surfaces. Three different types of nanostructures were fabricated: nanopillars, nanorings and nanonuggets. The nanopillars are the basic structures and are 50 nm in diameter and 100 nm in height. Particular arrangement of the nanopillars in various geometries formed nanorings and nanonuggets. Flat surfaces, rough substrate surfaces, and various nanostructured surfaces were compared for their abilities to attach and kill bacterial cells. Methicillin-resistant Staphylococcus aureus, a Gram-positive bacterial strain responsible for many infections in health care system, was used as the model bacterial strain. It was found that all the Au nanostructures, regardless their shapes, exhibited similar excellent antibacterial properties. A comparison of live cells attached to nanotopographic surfaces showed that the number of live S. aureus cells was <1% of that from flat and rough reference surfaces. Our micro/nanofabrication process is a scalable approach based on cost-efficient self-organization and provides potential for further developing functional surfaces to study the behavior of microbes on nanoscale topographies. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr06157a

A naturally occurring nanomaterial from the Sundew (Drosera) for tissue engineering.

PubMed

Lenaghan, S C; Serpersu, K; Xia, L; He, W; Zhang, M

2011-12-01

In recent years advances have been made in the design of novel materials for tissue engineering through the use of polysaccharides. This study evaluated the ability of a naturally secreted polysaccharide adhesive from the Sundew (Drosera capensis) as a support for cell growth. The Sundew adhesive has several advantages including its high elasticity and antibiotic nature. By coating glass cover slips with the Sundew adhesive, a network of nanofibers was generated that was capable of promoting attachment and differentiation of a model neuronal cell line, PC-12. We also demonstrated the potential of this material for repairing bone and soft tissue injuries, by testing attachment of osteoblasts and endothelial cells. Finally, it was determined that the Sundew biomaterial was stable through testing by atomic force microscopy and prolonged cell growth. This work has proven the capabilities of using a nanomaterial derived from the Sundew adhesive for the purpose of tissue engineering.

Basic mechanism for biorientation of mitotic chromosomes is provided by the kinetochore geometry and indiscriminate turnover of kinetochore microtubules

PubMed Central

Zaytsev, Anatoly V.; Grishchuk, Ekaterina L.

2015-01-01

Accuracy of chromosome segregation relies on the ill-understood ability of mitotic kinetochores to biorient, whereupon each sister kinetochore forms microtubule (MT) attachments to only one spindle pole. Because initial MT attachments result from chance encounters with the kinetochores, biorientation must rely on specific mechanisms to avoid and resolve improper attachments. Here we use mathematical modeling to critically analyze the error-correction potential of a simplified biorientation mechanism, which involves the back-to-back arrangement of sister kinetochores and the marked instability of kinetochore–MT attachments. We show that a typical mammalian kinetochore operates in a near-optimal regime, in which the back-to-back kinetochore geometry and the indiscriminate kinetochore–MT turnover provide strong error-correction activity. In human cells, this mechanism alone can potentially enable normal segregation of 45 out of 46 chromosomes during one mitotic division, corresponding to a mis-segregation rate in the range of 10−1–10−2 per chromosome. This theoretical upper limit for chromosome segregation accuracy predicted with the basic mechanism is close to the mis-segregation rate in some cancer cells; however, it cannot explain the relatively low chromosome loss in diploid human cells, consistent with their reliance on additional mechanisms. PMID:26424798

Identification of Pro-Differentiation p53 Target Genes and Evaluation of Expression in Normal and Malignant Mammary Gland

DTIC Science & Technology

2009-04-01

progenitor cells ’ ability to survive under non-attachment culture conditions, which is composed of primary mammoshpere and secondary mammosphere. Both... cell lineages in early mouse development depend on SOX2 function. Genes Dev, 17: 126-140, 2003 8. Chambers, I., Colby, D., Robertson, M., Nichols, J...Rivett, D., Jones, K., and Dalton, S. LIF/STAT3 controls ES cell self-renewal and pluripotency by a Myc-dependent mechanism. Development , 132: 885-896

Virion Glycoprotein-Mediated Immune Evasion by Human Cytomegalovirus: a Sticky Virus Makes a Slick Getaway

PubMed Central

Gardner, Thomas J.

2016-01-01

SUMMARY The prototypic herpesvirus human cytomegalovirus (CMV) exhibits the extraordinary ability to establish latency and maintain a chronic infection throughout the life of its human host. This is even more remarkable considering the robust adaptive immune response elicited by infection and reactivation from latency. In addition to the ability of CMV to exist in a quiescent latent state, its persistence is enabled by a large repertoire of viral proteins that subvert immune defense mechanisms, such as NK cell activation and major histocompatibility complex antigen presentation, within the cell. However, dissemination outside the cell presents a unique existential challenge to the CMV virion, which is studded with antigenic glycoprotein complexes targeted by a potent neutralizing antibody response. The CMV virion envelope proteins, which are critical mediators of cell attachment and entry, possess various characteristics that can mitigate the humoral immune response and prevent viral clearance. Here we review the CMV glycoprotein complexes crucial for cell attachment and entry and propose inherent properties of these proteins involved in evading the CMV humoral immune response. These include viral glycoprotein polymorphism, epitope competition, Fc receptor-mediated endocytosis, glycan shielding, and cell-to-cell spread. The consequences of CMV virion glycoprotein-mediated immune evasion have a major impact on persistence of the virus in the population, and a comprehensive understanding of these evasion strategies will assist in designing effective CMV biologics and vaccines to limit CMV-associated disease. PMID:27307580

Mother-Child Attachment From Infancy to the Preschool Years: Predicting Security and Stability.

PubMed

Meins, Elizabeth; Bureau, Jean-François; Fernyhough, Charles

2018-05-01

Relations between maternal mind-mindedness (appropriate and nonattuned mind-related comments), children's age-2 perspective-taking abilities, and attachment security at 44 (n = 165) and 51 (n = 128) months were investigated. Nonattuned comments predicted insecure preschool attachment, via insecure 15-month attachment security (44-month attachment) and poorer age-2 perspective-taking abilities (51-month attachment). With regard to attachment stability, higher perspective-taking abilities distinguished the stable secure groups from (a) the stable insecure groups and (b) children who changed from secure to insecure (at trend level). These effects were independent of child gender, stressful life events, and socioeconomic status (SES). The contribution of these findings to our understanding of stability and change in attachment security from infancy to the preschool years is discussed. © 2017 The Authors. Child Development © 2017 Society for Research in Child Development, Inc.

Teratogen metabolism: activation of thalidomide and thalidomide analogues to products that inhibit the attachment of cells to concanavalin A coated plastic surfaces.

PubMed

Braun, A G; Weinreb, S L

1984-05-01

Thalidomide metabolites inhibited the attachment of tumor cells to concanavalin A coated polyethylene surfaces. Thalidomide, itself, was non-inhibitory. Thalidomide activation to inhibitory products required hepatic microsomes, an NADPH-generating system, and molecular oxygen. Production of inhibitory metabolites was unaffected by either epoxide hydrolase or 1,2-epoxy-3,3,3-trichloropropane (TCPO), an inhibitor of epoxide hydrolase endogenous to hepatic S9 fraction. Therefore, the attachment inhibitor was probably not an arene oxide. Inhibition was not accompanied by cytotoxicity, as judged by trypan blue exclusion. Although uninduced hepatic microsomes from mice, rats and dogs had similar abilities to activate thalidomide, microsomes from Aroclor 1254 induced rats were relatively inactive in the system. Inhibitory metabolites were generated from the thalidomide analogues EM8 , EM12 , EM16 , EM87 , EM136 , EM255 , E350 , phthalimide, phthalimido-phthalimide, indan, 1- indanone and 1,3- indandione . Glutarimide , glutamic acid and phthalic acid did not activate to inhibitory products.

Characterization and Imaging of Antibody-Coated Gold Nanoparticles for Targeted Treatment of Microbial Keratitis

NASA Astrophysics Data System (ADS)

Mahan, Matthew

Microbial keratitis (MK) is an infection of the cornea by pathogenic organisms that causes inflammation and irritation. It can lead to full or partial blindness if left untreated. Current clinical treatment methods rely on high frequency application of topical drugs which are subject to the issues of patient compliance and microbial resistance. In this work, gold nanoparticles (AuNP) were proposed as an alternative treatment method in light-based therapies. Particle formulation methods were investigated and assessed using transmission electron microscopy (TEM) and ultraviolet/visible spectroscopy (UV-Vis). AuNP of 20 nm diameter were used as platforms to attach monoclonal antibodies anti-FLAG or anti-F1 to enhance their cell-targeting ability as well as polyethylene glycol to reduce non-specific binding and protein adsorption. These functionalized particles were qualitatively assessed using UV-Vis. The antibody-functionalized AuNP were then assessed for their ability to attach directly to Pseudomonas aeruginosa, expressing FLAG peptide, or Aspergillus fumigatus, expressing the F1 receptor. Attachment was imaged using dark field microscopy, transmission electron microscopy, and fluorescence microscopy.

Computational modeling of in vitro biological responses on polymethacrylate surfaces

PubMed Central

Ghosh, Jayeeta; Lewitus, Dan Y; Chandra, Prafulla; Joy, Abraham; Bushman, Jared; Knight, Doyle; Kohn, Joachim

2011-01-01

The objective of this research was to examine the capabilities of QSPR (Quantitative Structure Property Relationship) modeling to predict specific biological responses (fibrinogen adsorption, cell attachment and cell proliferation index) on thin films of different polymethacrylates. Using 33 commercially available monomers it is theoretically possible to construct a library of over 40,000 distinct polymer compositions. A subset of these polymers were synthesized and solvent cast surfaces were prepared in 96 well plates for the measurement of fibrinogen adsorption. NIH 3T3 cell attachment and proliferation index were measured on spin coated thin films of these polymers. Based on the experimental results of these polymers, separate models were built for homo-, co-, and terpolymers in the library with good correlation between experiment and predicted values. The ability to predict biological responses by simple QSPR models for large numbers of polymers has important implications in designing biomaterials for specific biological or medical applications. PMID:21779132

Derivation of Cinnamon Blocks Leukocyte Attachment by Interacting with Sialosides

PubMed Central

Lin, Wei-Ling; Guu, Shih-Yun; Tsai, Chan-Chuan; Prakash, Ekambaranellore; Viswaraman, Mohan; Chen, Hsing-Bao; Chang, Chuan-Fa

2015-01-01

Molecules derived from cinnamon have demonstrated diverse pharmacological activities against infectious pathogens, diabetes and inflammatory diseases. This study aims to evaluate the effect of the cinnamon-derived molecule IND02 on the adhesion of leukocytes to host cells. The anti-inflammatory ability of IND02, a pentameric procyanidin type A polyphenol polymer isolated from cinnamon alcohol extract, was examined. Pretreatment with IND02 significantly reduced the attachment of THP-1 cells or neutrophils to TNF-α-activated HUVECs or E-selectin/ICAM-1, respectively. IND02 also reduced the binding of E-, L- and P-selectins with sialosides. Furthermore, IND02 could agglutinate human red blood cells (RBC), and the agglutination could be disrupted by sialylated glycoprotein. Our findings demonstrate that IND02, a cinnamon-derived compound, can interact with sialosides and block the binding of selectins and leukocytes with sialic acids. PMID:26076445

Thrombospondin-induced adhesion of human keratinocytes.

PubMed Central

Varani, J; Nickoloff, B J; Riser, B L; Mitra, R S; O'Rourke, K; Dixit, V M

1988-01-01

Human epidermal keratinocytes obtained from normal skin attached and spread on thrombospondin (TSP)-coated plastic dishes but failed to attach and spread on untreated plastic culture dishes or dishes coated with fibronectin or laminin. These cells produced minimal amounts of immunoreactive TSP. Keratinocytes established in culture on MCDB 153 medium and maintained for one to three passages in an undifferentiated state by continued cultivation in this low Ca2+-containing medium attached and spread on plastic dishes as well as on TSP-coated dishes. These cells also secreted significant amounts of TSP into the culture medium. When the keratinocytes were incubated for one day in MCDB 153 medium supplemented with high Ca2+ or in MEM (which also contains high Ca2+), there was decreased secretion of TSP into the culture medium concomitant with a reduction in attachment and spreading on plastic culture dishes. Proteolytic fragments of TSP were examined for stimulation of keratinocyte attachment and spreading. A 140-kd fragment produced by removal of the 25-kd heparin-binding domain had similar activity to the intact molecule while the 25-kd fragment was without effect. Further proteolytic treatment of the 140-kd fragment gave rise to a fragment consisting of 120 kd and 18-D moieties held together in disulphide linkage. This fragment did not support attachment or spreading. This study reveals that normal epidermal keratinocytes grown under conditions that maintain the undifferentiated state are able to produce TSP and utilize it as an attachment factor. When keratinocytes are grown under conditions that promote differentiation, ability to produce and utilize TSP is diminished. Since TSP is present at the dermal-epidermal junction and because TSP promotes keratinocyte attachment and spreading, this molecule may play an important role in maintaining normal growth of the basal cell layer and may also participate in reepithelialization during wound repair. Images PMID:2452837

Sexual dimorphism in the attachment ability of the ladybird beetle Coccinella septempunctata on soft substrates

NASA Astrophysics Data System (ADS)

Heepe, Lars; Petersen, Dennis S.; Tölle, Lisa; Wolff, Jonas O.; Gorb, Stanislav N.

2017-01-01

Many insects possess adhesive foot pads, which enable reliable attachment to diverse and unpredictable substrates. The function of these adhesive organs was shown to be affected by environmental conditions such as substrate roughness, chemistry, and ambient humidity. So far, the attachment ability of insects and also that of spiders and geckos has been tested on rigid substrates only. However, the natural habitats of climbing animals may provide a variety of substrate stiffness ranging from rigid rock surfaces to soft, biofilm covered substrates. In order to test the effect of different substrate stiffness on the attachment ability of insects, we have performed friction experiments with female and male ladybird beetles Coccinella septempunctata on smooth silicone elastomer substrates of different stiffness, using a centrifugal force tester. Whereas in females, the attachment ability was not affected by the substrate stiffness within the range of tested stiffness, males showed decreasing attachment ability with decreasing substrate stiffness. This sexual dimorphism in attachment ability is explained by the presence of a specialized, discoidal seta type in males, which is not present in females. It is argued that discoidal setae, when softer if compared to the substrate, may show an advantageous peak-free interfacial stress distribution when being pulled off the substrate. For such setae being stiffer if compared the substrate, they potentially show increased edge stress concentration. In this case, lower pull-off forces are expected, in agreement with the experimentally obtained results. With the present study, we demonstrate for the first time that the substrate stiffness may have an effect on the attachment ability of climbing animals, which may also be of relevance for technical and medical applications involving adhesion to soft substrates.

Cytotoxic Tumor-Targeting Peptides From In Vivo Phage Display.

PubMed

Northup, Jessica R Newton; Deutscher, Susan L

2016-01-01

We previously utilized an in vivo peptide phage display selection technique, which included the use of detergent elution of phage from excised tumor, to obtain tumor-targeting phage with the ability to extravasate the vasculature and bind directly to prostate tumor tissue. It is hypothesized that this same in vivo phage selection technique can be used to functionally select for molecules that not only bind to cancer cells but also kill them. Here we analyzed two different in vivo phage display selected phage clones, G1 and H5, retrieved from PC-3 human prostate carcinoma xenografted tumors. First, cell de-attachment as an endpoint criterion for apoptosis and cell cycle was examined. After 2.5 hours incubation with G1 phage, PC-3 cell attachment was reduced by 23.8% and the percent of cell population in M phase reduced by 32.1%. In comparison, PC-3 cells incubated with H5 phage had a reduction of 25.0% cell attachment and 33.6% of cell population in M phase. These changes in combination with elevated caspase activation within cells in M phase, and no significant changes to G1/G0 or S phase cell populations suggest that the cytotoxic phages are targeting actively dividing PC-3 cells. Microscopic studies were also performed to further analyze the nature of cytotoxicity of these two phage clones. It was found that G1 phage induced and co- localized with tubulin based projections within apoptotic cells, while H5 phage did not. These phage may form the foundation for a new class of targeted prostate cancer therapeutic agents.

Locomotion and attachment of leaf beetle larvae Gastrophysa viridula (Coleoptera, Chrysomelidae).

PubMed

Zurek, Daniel B; Gorb, Stanislav N; Voigt, Dagmar

2015-02-06

While adult green dock leaf beetles Gastrophysa viridula use tarsal adhesive setae to attach to and walk on smooth vertical surfaces and ceilings, larvae apply different devices for similar purposes: pretarsal adhesive pads on thoracic legs and a retractable pygopod at the 10th abdominal segment. Both are soft smooth structures and capable of wet adhesion. We studied attachment ability of different larval instars, considering the relationship between body weight and real contact area between attachment devices and the substrate. Larval gait patterns were analysed using high-speed video recordings. Instead of the tripod gait of adults, larvae walked by swinging contralateral legs simultaneously while adhering by the pygopod. Attachment ability of larval instars was measured by centrifugation on a spinning drum, revealing that attachment force decreases relative to weight. Contributions of different attachment devices to total attachment ability were investigated by selective disabling of organs by covering them with melted wax. Despite their smaller overall contact area, tarsal pads contributed to a larger extent to total attachment ability, probably because of their distributed spacing. Furthermore, we observed different behaviour in adults and larvae when centrifuged: while adults gradually slipped outward on the centrifuge drum surface, larvae stayed at the initial position until sudden detachment.

Locomotion and attachment of leaf beetle larvae Gastrophysa viridula (Coleoptera, Chrysomelidae)

PubMed Central

Zurek, Daniel B.; Gorb, Stanislav N.; Voigt, Dagmar

2015-01-01

While adult green dock leaf beetles Gastrophysa viridula use tarsal adhesive setae to attach to and walk on smooth vertical surfaces and ceilings, larvae apply different devices for similar purposes: pretarsal adhesive pads on thoracic legs and a retractable pygopod at the 10th abdominal segment. Both are soft smooth structures and capable of wet adhesion. We studied attachment ability of different larval instars, considering the relationship between body weight and real contact area between attachment devices and the substrate. Larval gait patterns were analysed using high-speed video recordings. Instead of the tripod gait of adults, larvae walked by swinging contralateral legs simultaneously while adhering by the pygopod. Attachment ability of larval instars was measured by centrifugation on a spinning drum, revealing that attachment force decreases relative to weight. Contributions of different attachment devices to total attachment ability were investigated by selective disabling of organs by covering them with melted wax. Despite their smaller overall contact area, tarsal pads contributed to a larger extent to total attachment ability, probably because of their distributed spacing. Furthermore, we observed different behaviour in adults and larvae when centrifuged: while adults gradually slipped outward on the centrifuge drum surface, larvae stayed at the initial position until sudden detachment. PMID:25657837

Effect of bioactive extruded PLA/HA composite films on focal adhesion formation of preosteoblastic cells.

PubMed

Persson, Maria; Lorite, Gabriela S; Kokkonen, Hanna E; Cho, Sung-Woo; Lehenkari, Petri P; Skrifvars, Mikael; Tuukkanen, Juha

2014-09-01

The quality of the initial cell attachment to a biomaterial will influence any further cell function, including spreading, proliferation, differentiation and viability. Cell attachment is influenced by the material's ability to adsorb proteins, which is related to the surface chemistry and topography of the material. In this study, we incorporated hydroxyapatite (HA) particles into a poly(lactic acid) (PLA) composite and evaluated the surface structure and the effects of HA density on the initial cell attachment in vitro of murine calvarial preosteoblasts (MC3T3-EI). Scanning electron microscopy (SEM), atomic force microscopy (AFM) and infrared spectroscopy (FTIR) showed that the HA particles were successfully incorporated into the PLA matrix and located at the surface which is of importance in order to maintain the bioactive effect of the HA particles. SEM and AFM investigation revealed that the HA density (particles/area) as well as surface roughness increased with HA loading concentration (i.e. 5, 10, 15 and 20wt%), which promoted protein adsorption. Furthermore, the presence of HA on the surface enhanced cell spreading, increased the formation of actin stress fibers and significantly improved the expression of vinculin in MC3T3-E1 cells which is a key player in the regulation of cell adhesion. These results suggest the potential utility of PLA/HA composites as biomaterials for use as a bone substitute material and in tissue engineering applications. Copyright © 2014 Elsevier B.V. All rights reserved.

The effect of Me2SO overexposure during cryopreservation on HOS TE85 and hMSC viability, growth and quality.

PubMed

Morris, Timothy J; Picken, Andrew; Sharp, Duncan M C; Slater, Nigel K H; Hewitt, Christopher J; Coopman, Karen

2016-12-01

With the cell therapy industry continuing to grow, the ability to preserve clinical grade cells, including mesenchymal stem cells (MSCs), whilst retaining cell viability and function remains critical for the generation of off-the-shelf therapies. Cryopreservation of MSCs, using slow freezing, is an established process at lab scale. However, the cytotoxicity of cryoprotectants, like Me 2 SO, raises questions about the impact of prolonged cell exposure to cryoprotectant at temperatures >0 °C during processing of large cell batches for allogenic therapies prior to rapid cooling in a controlled rate freezer or in the clinic prior to administration. Here we show that exposure of human bone marrow derived MSCs to Me 2 SO for ≥1 h before freezing, or after thawing, degrades membrane integrity, short-term cell attachment efficiency and alters cell immunophenotype. After 2 h's exposure to Me 2 SO at 37 °C post-thaw, membrane integrity dropped to ∼70% and only ∼50% of cells retained the ability to adhere to tissue culture plastic. Furthermore, only 70% of the recovered MSCs retained an immunophenotype consistent with the ISCT minimal criteria after exposure. We also saw a similar loss of membrane integrity and attachment efficiency after exposing osteoblast (HOS TE85) cells to Me 2 SO before, and after, cryopreservation. Overall, these results show that freezing medium exposure is a critical determinant of product quality as process scale increases. Defining and reporting cell sensitivity to freezing medium exposure, both before and after cryopreservation, enables a fair judgement of how scalable a particular cryopreservation process can be, and consequently whether the therapy has commercial feasibility. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Laser microsurgery of higher plant cell walls permits patch-clamp access

NASA Technical Reports Server (NTRS)

Henriksen, G. H.; Taylor, A. R.; Brownlee, C.; Assmann, S. M.; Evans, M. L. (Principal Investigator)

1996-01-01

Plasma membranes of guard cells in epidermal peels of Vicia faba and Commelina communis can be made accessible to a patch-clamp pipet by removing a small portion (1-3 micrometers in diameter) of the guard cell wall using a microbeam of ultraviolet light generated by a nitrogen laser. Using this laser microsurgical technique, we have measured channel activity across plasma membranes of V. faba guard cells in both cell-attached and isolated patch configurations. Measurements made in the inside-out patch configuration revealed two distinct K(+)-selective channels. Major advantages of the laser microsurgical technique include the avoidance of enzymatic protoplast isolation, the ability to study cell types that have been difficult to isolate as protoplasts or for which enzymatic isolation protocols result in protoplasts not amenable to patch-clamp studies, the maintenance of positional information in single-channel measurements, reduced disruption of cell-wall-mediated signaling pathways, and the ability to investigate intercellular signaling through studies of cells remaining situated within tissue.

Engineering of Surface Functionality onto Polystyrene Microcarriers for the Attachment and Growth of Human Endothelial Cells

NASA Astrophysics Data System (ADS)

Xiong, Gordon M.; Foord, John S.; Griffiths, Jon-Paul; Parker, Emily M.; Moloney, Mark G.; Choong, Cleo

2014-08-01

This work reports the effects of introducing diverse chemical functionalities onto the surface of polystyrene microcarrier beads on their ability to function as injectable cell carriers. Cellular adhesion and proliferation, as well as cellular outgrowths from microcarrier surfaces, using human umbilical vein endothelial cells (HUVECs), were examined in detail. It was observed that initial cell adhesion appeared to be most significantly decreased by hydrophobicity, whilst cell proliferation appeared to be improved in most chemical functional groups over unmodified polystyrene. Overall, our study highlights the importance of surface chemistry in directing the growth and function of human endothelial cells.

Interaction of herpes simplex virus glycoprotein gC with mammalian cell surface molecules.

PubMed Central

Tal-Singer, R; Peng, C; Ponce De Leon, M; Abrams, W R; Banfield, B W; Tufaro, F; Cohen, G H; Eisenberg, R J

1995-01-01

The entry of herpes simplex virus (HSV) into mammalian cells is a multistep process beginning with an attachment step involving glycoproteins gC and gB. A second step requires the interaction of glycoprotein gD with a cell surface molecule. We explored the interaction between gC and the cell surface by using purified proteins in the absence of detergent. Truncated forms of gC and gD, gC1(457t), gC2(426t), and gD1(306t), lacking the transmembrane and carboxyl regions were expressed in the baculovirus system. We studied the ability of these proteins to bind to mammalian cells, to bind to immobilized heparin, to block HSV type 1 (HSV-1) attachment to cells, and to inhibit plaque formation by HSV-1. Each of these gC proteins bound to conformation-dependent monoclonal antibodies and to human complement component C3b, indicating that they maintained the same conformation of gC proteins expressed in mammalian cells. Biotinylated gC1(457t) and gC2(426t) each bind to several cell lines. Binding was inhibited by an excess of unlabeled gC but not by gD, indicating specificity. The attachment of gC to cells involves primarily heparan sulfate proteoglycans, since heparitinase treatment of cells reduced gC binding by 50% but had no effect on gD binding. Moreover, binding of gC to two heparan sulfate-deficient L-cell lines, gro2C and sog9, both of which are mostly resistant to HSV infection, was markedly reduced. Purified gD1 (306t), however, bound equally well to the two mutant cell lines. In contrast, saturating amounts of gC1(457t) interfered with HSV-1 attachment to cells but failed to block plaque formation, suggesting a role for gC in attachment but not penetration. A mutant form of gC lacking residues 33 to 123, gC1(delta 33-123t), expressed in the baculovirus system, bound significantly less well to cells than did gC1(457t) and competed poorly with biotinylated gC1(457t) for binding. These results suggest that residues 33 to 123 are important for gC attachment to cells. In contrast, both the mutant and wild-type forms of gC bound to immobilized heparin, indicating that binding of these proteins to the cell surface involves more than a simple interaction with heparin. To determine that the contribution of the N-terminal region of gC is important for HSV attachment, we compared several properties of a mutant HSV-1 which contains gC lacking amino acids 33 to 123 to those of its parental virus, which contains full-length gC. The mutant bound less well to cells than the parental virus but exhibited normal growth properties.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:7769707

[Preparation of elastic porous cell scaffold fabricated with combined polydimethylsiloxane (PDMS) and hydroxyapatite (HA)].

PubMed

Yang, Yang; Lan, Ding; Huang, Yan; Li, Yanming; Wang, Yuren; Sun, Lianwen; Fan, Yubo

2014-06-01

Polydimethylsiloxane (PDMS) and hydroxyapatite (HA) were combined in our laboratory to fabricate an elastic porous cell scaffold with pore-forming agent, and then the scaffold was used as culture media for rat bone marrow derived mesenchymal stem cells (rBMSCs). Different porous materials (square and circular in shape) were prepared by different pore-forming agents (NaCl or paraffin spheres) with adjustable porosity (62%-76%). The HA crystals grew on the wall of hole when the material was exposed to SBF solutions, showing its biocompatibility and ability to support the cells to attach on the materials.

Substratum interfacial energetic effects on the attachment of marine bacteria

NASA Astrophysics Data System (ADS)

Ista, Linnea Kathryn

Biofilms represent an ancient, ubiquitous and influential form of life on earth. Biofilm formation is initiated by attachment of bacterial cells from an aqueous suspension onto a suitable attachment substratum. While in certain, well studied cases initial attachment and subsequent biofilm formation is mediated by specific ligand-receptor pairs on the bacteria and attachment substratum, in the open environment, including the ocean, it is assumed to be non-specific and mediated by processes similar to those that drive adsorption of colloids at the water-solid interface. Colloidal principles are studied to determine the molecular and physicochemical interactions involved in the attachment of the model marine bacterium, Cobetia marina to model self-assembled monolayer surfaces. In the simplest application of colloidal principles the wettability of attachment substrata, as measured by the advancing contact angle of water (theta AW) on the surface, is frequently used as an approximation for the surface tension. We demonstrate the applicability of this approach for attachment of C. marina and algal zoospores and extend it to the development of a means to control attachment and release of microorganisms by altering and tuning surface thetaAW. In many cases, however, thetaAW does not capture all the information necessary to model attachment of bacteria to attachment substrata; SAMs with similar thetaAW attach different number of bacteria. More advanced colloidal models of initial bacterial attachment have evolved over the last several decades, with the emergence of the model proposed by van Oss, Chaudhury and Good (VCG) as preeminent. The VCG model enables calculation of interfacial tensions by dividing these into two major interactions thought to be important at biointerfaces: apolar, Lifshitz-van der Waals and polar, Lewis acid-base (including hydrogen bonding) interactions. These interfacial tensions are combined to yield DeltaGadh, the free energy associated with attachment of bacteria to a substratum. We use VCG to model DeltaGadh and interfacial tensions as they relate to model bacterial attachment on SAMs that accumulate cells to different degrees. Even with the more complex interactions measured by VCG, surface energy of the attachment substratum alone was insufficient to predict attachment. VCG was then employed to model attachment of C. marina to a series of SAMs varying systematically in the number of ethylene glycol residues present in the molecule; an identical series has been previously shown to vary dramatically in the number of cells attached as a function of ethylene glycols present. Our results indicate that while VCG adequately models the interfacial tension between water and ethylene glycol SAMs in a manner that predicts bacterial attachment, DeltaGadh as calculated by VCG neither qualitatively nor quantitatively reflects the attachment data. The VCG model, thus, fails to capture specific information regarding the interactions between the attaching bacteria, water, and the SAM. We show that while hydrogen-bond accepting interactions are very well captured by this model, the ability for SAMs and bacteria to donate hydrogen bonds is not adequately described as the VCG model is currently applied. We also describe ways in which VCG fails to capture two specific biological aspects that may be important in bacterial attachment to surfaces:1.) specific interactions between molecules on the surface and bacteria and 2.) bacterial cell surface heterogeneities that may be important in differential attachment to different substrata.

Spatial Periodicity of Escherichia coli K-12 Biofilm Microstructure Initiates during a Reversible, Polar Attachment Phase of Development and Requires the Polysaccharide Adhesin PGA

PubMed Central

Agladze, Konstantin; Wang, Xin; Romeo, Tony

2005-01-01

Using fast Fourier transform (FFT) analysis, we previously observed that cells within Escherichia coli biofilm are organized in nonrandom or periodic spatial patterns (K. Agladze et al., J. Bacteriol. 185:5632-5638, 2003). Here, we developed a gravity displacement assay for examining cell adherence and used it to quantitatively monitor the formation of two distinct forms of cell attachment, temporary and permanent, during early biofilm development. Temporarily attached cells were mainly surface associated by a cell pole; permanent attachments were via the lateral cell surface. While temporary attachment precedes permanent attachment, both forms can coexist in a population. Exposure of attached cells to gravity liberated an unattached population capable of rapidly reassembling a new monolayer, composed of temporarily attached cells, and possessing periodicity. A csrA mutant, which forms biofilm more vigorously than its wild-type parent, exhibited an increased proportion of permanently attached cells and a form of attachment that was not apparent in the parent strain, permanent polar attachment. Nevertheless, it formed periodic attachment patterns. In contrast, biofilm mutants with altered lipopolysaccharide synthesis (waaG) exhibited increased cell-cell interactions, bypassed the polar attachment step, and produced FFT spectra characteristic of aperiodic cell distribution. Mutants lacking the polysaccharide adhesin β-1,6-N-acetyl-d-glucosamine (ΔpgaC) also exhibited aperiodic cell distribution, but without apparent cell-cell interactions, and were defective in forming permanent attachments. Thus, spatial periodicity of biofilm microstructure is genetically determined and evident during the formation of temporary cell surface attachments. PMID:16321928

Comparison of osteoblast-like cell responses to calcium silicate and tricalcium phosphate ceramics in vitro.

PubMed

Ni, Siyu; Chang, Jiang; Chou, Lee; Zhai, Wanyin

2007-01-01

Calcium silicate ceramics have been proposed as new bone repair biomaterials, since they have proved to be bioactive, degradable, and biocompatible. Beta-tricalcium phosphate ceramic is a well-known degradable material for bone repair. This study compared the effects of CaSiO3 (alpha-, and beta-CaSiO3) and beta-Ca3(PO4)2 (beta-TCP) ceramics on the early stages of rat osteoblast-like cell attachment, proliferation, and differentiation. Osteoblast-like cells were cultured directly on CaSiO3 (alpha-, and beta-CaSiO3) and beta-TCP ceramics. Attachment of a greater number of cells was observed on CaSiO3 (alpha-, and beta-CaSiO3) ceramics compared with beta-TCP ceramics after incubation for 6 h. SEM observations showed an intimate contact between cells and the substrates, significant cells adhesion, and that the cells spread and grew on the surfaces of all the materials. In addition, the proliferation rate and alkaline phosphatase (ALP) activity of the cells on the CaSiO3 (alpha-, and beta-CaSiO3) ceramics were improved when compared with the beta-TCP ceramics. In the presence of CaSiO3, elevated levels of calcium and silicon in the culture medium were observed throughout the 7-day culture period. In conclusion, the results of the present study revealed that CaSiO3 ceramics showed greater ability to support cell attachment, proliferation, and differentiation than beta-TCP ceramic. 2006 Wiley Periodicals, Inc.

Microbiological study of biofilm formation in isolates of Salmonella enterica Typhimurium DT104 and DT104b cultured from the modern pork chain.

PubMed

O'Leary, Denis; Cabe, Evonne M Mc; McCusker, Matthew P; Martins, Marta; Fanning, Séamus; Duffy, Geraldine

2013-01-15

The purpose of this study was to characterise 172 Salmonella Typhimurium isolates taken from the pork chain for their biofilm forming abilities and to analyse their potential to survive on food processing surfaces. Many Salmonella have the ability to form biofilms. These natural structures, elaborated by bacteria are important in food production because their formation contributes to bacterial survival. Adherent bacterial cells are more resilient to displacement strategies including physical and chemical procedures as a consequence of their altered more resistant phenotype. By improving our understanding of the nature of biofilms, this data could positively contribute to the development and implementation of eradication strategies. In this study, Salmonella Typhimurium DT104 and DT104b were investigated for their ability to form biofilms on a range of different surfaces under defined environmental growth conditions. Phenotypic characterisation involved examining colony morphology on indicator agars, assessing their ability to survive chlorine-based challenges and investigating their ability to attach to stainless steel and to plastic surfaces. All bacterial isolates were investigated for the presence of Salmonella genomic island I (SGI1) which is thought to enhance efficient biofilm formation. It was found that the majority of strains possess biofilm forming capabilities but successful attachment is highly dependent on the surface on which the biofilm is forming. The strains readily attached to stainless steel and plastic surfaces and survived high chlorine concentrations. Molecular and phenotypic comparisons of strong and weak biofilm forming strains indicate that biofilm development is not solely dependent on the acquirement of SGI1. Copyright © 2012 Elsevier B.V. All rights reserved.

Compositions and methods for adoptive and active immunotherapy

DOEpatents

Fahmy, Tarek; Steenblock, Erin

2014-01-14

Modular aAPCs and methods of their manufacture and use are provided. The modular aAPCs are constructed from polymeric microparticles. The aAPCs include encapsulated cytokines and coupling agents which modularly couple functional elements including T cell receptor activators, co-stimulatory molecules and adhesion molecules to the particle. The ability of these aAPCs to release cytokines in a controlled manner, coupled with their modular nature and ease of ligand attachment, results in an ideal, tunable APC capable of stimulating and expanding primary T cells.

Fabrication of three-dimensional scaffolds using precision extrusion deposition with an assisted cooling device.

PubMed

Hamid, Q; Snyder, J; Wang, C; Timmer, M; Hammer, J; Guceri, S; Sun, W

2011-09-01

In the field of biofabrication, tissue engineering and regenerative medicine, there are many methodologies to fabricate a building block (scaffold) which is unique to the target tissue or organ that facilitates cell growth, attachment, proliferation and/or differentiation. Currently, there are many techniques that fabricate three-dimensional scaffolds; however, there are advantages, limitations and specific tissue focuses of each fabrication technique. The focus of this initiative is to utilize an existing technique and expand the library of biomaterials which can be utilized to fabricate three-dimensional scaffolds rather than focusing on a new fabrication technique. An expanded library of biomaterials will enable the precision extrusion deposition (PED) device to construct three-dimensional scaffolds with enhanced biological, chemical and mechanical cues that will benefit tissue generation. Computer-aided motion and extrusion drive the PED to precisely fabricate micro-scaled scaffolds with biologically inspired, porosity, interconnectivity and internal and external architectures. The high printing resolution, precision and controllability of the PED allow for closer mimicry of tissues and organs. The PED expands its library of biopolymers by introducing an assisting cooling (AC) device which increases the working extrusion temperature from 120 to 250 °C. This paper investigates the PED with the integrated AC's capabilities to fabricate three-dimensional scaffolds that support cell growth, attachment and proliferation. Studies carried out in this paper utilized a biopolymer whose melting point is established to be 200 °C. This polymer was selected to illustrate the newly developed device's ability to fabricate three-dimensional scaffolds from a new library of biopolymers. Three-dimensional scaffolds fabricated with the integrated AC device should illustrate structural integrity and ability to support cell attachment and proliferation.

Smart Biointerface with Photoswitched Functions between Bactericidal Activity and Bacteria-Releasing Ability.

PubMed

Wei, Ting; Zhan, Wenjun; Yu, Qian; Chen, Hong

2017-08-09

Smart biointerfaces with capability to regulate cell-surface interactions in response to external stimuli are of great interest for both fundamental research and practical applications. Smart surfaces with "ON/OFF" switchability for a single function such as cell attachment/detachment are well-known and useful, but the ability to switch between two different functions may be seen as the next level of "smart". In this work reported, a smart supramolecular surface capable of switching functions reversibly between bactericidal activity and bacteria-releasing ability in response to UV-visible light is developed. This platform is composed of surface-containing azobenzene (Azo) groups and a biocidal β-cyclodextrin derivative conjugated with seven quaternary ammonium salt groups (CD-QAS). The surface-immobilized Azo groups in trans form can specially incorporate CD-QAS to achieve a strongly bactericidal surface that kill more than 90% attached bacteria. On irradiation with UV light, the Azo groups switch to cis form, resulting in the dissociation of the Azo/CD-QAS inclusion complex and release of dead bacteria from the surface. After the kill-and-release cycle, the surface can be easily regenerated for reuse by irradiation with visible light and reincorporation of fresh CD-QAS. The use of supramolecular chemistry represents a promising approach to the realization of smart, multifunctional surfaces, and has the potential to be applied to diverse materials and devices in the biomedical field.

Myrip couples the capture of secretory granules by the actin-rich cell cortex and their attachment to the plasma membrane.

PubMed

Huet, Sébastien; Fanget, Isabelle; Jouannot, Ouardane; Meireles, Patricia; Zeiske, Tim; Larochette, Nathanaël; Darchen, François; Desnos, Claire

2012-02-15

Exocytosis of secretory granules (SGs) requires their delivery to the actin-rich cell cortex followed by their attachment to the plasma membrane (PM). How these reactions are executed and coordinated is still unclear. Myrip, which is also known as Slac-2c, binds to the SG-associated GTPase Rab27 and is thought to promote the delivery of SGs to the PM by recruiting the molecular motor myosin Va. Myrip also interacts with actin and the exocyst complex, suggesting that it may exert multiple roles in the secretory process. By combining total internal reflection fluorescence microscopy, single-particle tracking, a photoconversion-based assay, and mathematical modeling, we show that, in human enterochromaffin cells, Myrip (1) inhibits a class of SG motion characterized by fast and directed movement, suggesting that it facilitates the dissociation of SGs from microtubules; (2) enhances their motion toward the PM and the probability of SG attachment to the PM; and (3) increases the characteristic time of immobilization at the PM, indicating that it is a component of the molecular machinery that tether SGs to the PM. Remarkably, while the first two effects of Myrip depend on its ability to recruit myosin Va on SGs, the third is myosin Va independent but relies on the C-terminal domain of Myrip. We conclude that Myrip couples the retention of SGs in the cell cortex, their transport to the PM, and their attachment to the PM, and thus promotes secretion. These three steps of the secretory process are thus intimately coordinated.

Nanoparticle bioconjugate for controlled cellular delivery of doxorubicin

NASA Astrophysics Data System (ADS)

Sangtani, Ajmeeta; Petryayeva, Eleonora; Wu, Miao; Susumu, Kimihiro; Oh, Eunkeu; Huston, Alan L.; Lasarte-Aragones, Guillermo; Medintz, Igor L.; Algar, W. Russ; Delehanty, James B.

2018-02-01

Nanoparticle (NP)-mediated drug delivery offers the potential to overcome limitations of systemic delivery, including the ability to specifically target cargo and control release of NP-associated drug cargo. Doxorubicin (DOX) is a widely used FDA-approved cancer therapeutic; however, multiple side effects limit its utility. Thus, there is wide interest in modulating toxicity after cell delivery. Our goal here was to realize a NP-based DOX-delivery system that can modulate drug toxicity by controlling the release kinetics of DOX from the surface of a hard NP carrier. To achieve this, we employed a quantum dot (QD) as a central scaffold which DOX was appended via three different peptidyl linkages (ester, disulfide, hydrazone) that are cleavable in response to various intracellular conditions. Attachment of a cell penetrating peptide (CPP) containing a positively charged polyarginine sequence facilitates endocytosis of the ensemble. Polyhistidine-driven metal affinity coordination was used to self-assemble both peptides to the QD surface, allowing for fine control over both the ratio of peptides attached to the QD as well as DOX dose delivered to cells. Microplate-based Förster resonance energy transfer assays confirmed the successful ratiometric assembly of the conjugates and functionality of the linkages. Cell delivery experiments and cytotoxicity assays were performed to compare the various cleavable linkages to a control peptide where DOX is attached through an amide bond. The role played by various attachment chemistries used in QD-peptide-drug assemblies and their implications for the rationale in design of NPbased constructs for drug delivery is described here.

Role of curli and cellulose expression by Escherichia coli O157:H7 on the cell’s ability to attach to spinach

USDA-ARS?s Scientific Manuscript database

Shiga-toxigenic Escherichia coli O157:H7 (STEC) outbreaks have been linked to consumption of fresh produce. Cellular appendages, such as curli fibers have been suggested to be involved in STEC persistence in fresh produce as these curli are critical in biofilm formation and adherence to animal cell...

The multifunctional wound dressing with core-shell structured fibers prepared by coaxial electrospinning

NASA Astrophysics Data System (ADS)

Wei, Qilin; Xu, Feiyang; Xu, Xingjian; Geng, Xue; Ye, Lin; Zhang, Aiying; Feng, Zengguo

2016-06-01

The non-woven wound dressing with core-shell structured fibers was prepared by coaxial electrospinning. The polycaprolactone (PCL) was electrospun as the fiber's core to provide mechanical strength whereas collagen was fabricated into the shell in order to utilize its good biocompatibility. Simultaneously, the silver nanoparticles (Ag-NPs) as anti-bacterial agent were loaded in the shell whereas the vitamin A palmitate (VA) as healing-promoting drug was encapsulated in the core. Resulting from the fiber's core-shell structure, the VA released from the core and Ag-NPs present in the shell can endow the dressing both heal-promoting and anti-bacteria ability simultaneously, which can greatly enhance the dressing's clinical therapeutic effect. The dressing can maintain high swelling ratio of 190% for 3 d indicating its potential application as wet dressing. Furthermore, the dressing's anti-bacteria ability against Staphylococcus aureus was proved by in vitro anti-bacteria test. The in vitro drug release test showed the sustainable release of VA within 72 h, while the cell attachment showed L929 cells can well attach on the dressing indicating its good biocompatibility. In conclusion, the fabricated nanofibrous dressing possesses multiple functions to benefit wound healing and shows promising potential for clinical application.

Strengths and Difficulties as Correlates of Attachment Style in Institutionalized and Non-Institutionalized Children with Below-Average Intellectual Abilities

ERIC Educational Resources Information Center

Muris, Peter; Maas, Anneke

2004-01-01

The current study examined attachment style, strengths, and difficulties in institutionalized and non-institutionalized children with below-average intellectual abilities. Parents/caregivers and teachers of the children completed a brief measure of attachment style and the Strengths and Difficulties Questionnaire, which assesses the most important…

Competition for attachment of aquaculture candidate probiotic and pathogenic bacteria on fish intestinal mucus.

PubMed

Vine, N G; Leukes, W D; Kaiser, H; Daya, S; Baxter, J; Hecht, T

2004-06-01

Probiotics for aquaculture are generally only selected by their ability to produce antimicrobial metabolites; however, attachment to intestinal mucus is important in order to remain within the gut of its host. Five candidate probiotics (AP1-AP5), isolated from the clownfish, Amphiprion percula (Lacepéde), were examined for their ability to attach to fish intestinal mucus and compete with two pathogens, Aeromonas hydrophila and Vibrio alginolyticus. Two different radioactive isotopes were used to quantify competition between pathogens and probionts. Attachment of the pathogens was enhanced by the presence of the candidate probiotics. However, the addition of the candidate probiotics after the pathogens resulted in reduced pathogen attachment. Only AP5 caused lower attachment success of V. alginolyticus when added before the pathogen. When AP5 was added first, the average attachment change was 41% compared with 72% when added after V. alginolyticus, suggesting that the probiotic is displaced but that enhanced attachment of the pathogen does not occur. Conversely, when V. alginolyticus was added first, followed by AP5, attachment change was 37% while AP5 had 92% attachment change when added second. This implies that the pathogen was displaced by the candidate probiotic and therefore it appeared that, based on the ability of probiont AP5 to attach to mucus, the growth of the pathogen in the digestive tract might be suppressed by the candidate probiont's presence.

Antibacterial Au nanostructured surfaces.

PubMed

Wu, Songmei; Zuber, Flavia; Brugger, Juergen; Maniura-Weber, Katharina; Ren, Qun

2016-02-07

We present here a technological platform for engineering Au nanotopographies by templated electrodeposition on antibacterial surfaces. Three different types of nanostructures were fabricated: nanopillars, nanorings and nanonuggets. The nanopillars are the basic structures and are 50 nm in diameter and 100 nm in height. Particular arrangement of the nanopillars in various geometries formed nanorings and nanonuggets. Flat surfaces, rough substrate surfaces, and various nanostructured surfaces were compared for their abilities to attach and kill bacterial cells. Methicillin-resistant Staphylococcus aureus, a Gram-positive bacterial strain responsible for many infections in health care system, was used as the model bacterial strain. It was found that all the Au nanostructures, regardless their shapes, exhibited similar excellent antibacterial properties. A comparison of live cells attached to nanotopographic surfaces showed that the number of live S. aureus cells was <1% of that from flat and rough reference surfaces. Our micro/nanofabrication process is a scalable approach based on cost-efficient self-organization and provides potential for further developing functional surfaces to study the behavior of microbes on nanoscale topographies.

Salmonella enterica isolates from layer farm environments are able to form biofilm on eggshell surfaces.

PubMed

Pande, Vivek V; McWhorter, Andrea R; Chousalkar, Kapil K

2016-08-01

This study examined the eggshell biofilm forming ability of Salmonella enterica isolates recovered from egg farms. Multicellular behaviour and biofilm production were examined at 22 and 37°C by Congo red morphology and the crystal violet staining assay. The results indicated that the biofilm forming behaviour of Salmonella isolates was dependent on temperature and associated with serovars. Significantly greater biofilm production was observed at 22°C compared with 37°C. The number of viable biofilm cells attached to eggshells after incubation for 48 h at 22°C was significantly influenced by serovar. Scanning electron microscopic examination revealed firm attachment of bacterial cells to the eggshell surface. The relative expression of csgD and adrA gene was significantly higher in eggshell biofilm cells of S. Mbandaka and S. Oranienburg. These findings demonstrate that Salmonella isolates are capable of forming biofilm on the eggshell surface and that this behaviour is influenced by temperature and serovar.

Development of magnesium calcium phosphate biocement for bone regeneration.

PubMed

Jia, Junfeng; Zhou, Huanjun; Wei, Jie; Jiang, Xin; Hua, Hong; Chen, Fangping; Wei, Shicheng; Shin, Jung-Woog; Liu, Changsheng

2010-08-06

Magnesium calcium phosphate biocement (MCPB) with rapid-setting characteristics was fabricated by using the mixed powders of magnesium oxide (MgO) and calcium dihydrogen phosphate (Ca(H(2)PO(4))(2).H(2)O). The results revealed that the MCPB hardened after mixing the powders with water for about 7 min, and the compressive strength reached 43 MPa after setting for 1 h, indicating that the MCPB had a short setting time and high initial mechanical strength. After the acid-base reaction of MCPB containing MgO and Ca(H(2)PO(4))(2).H(2)O in a molar ratio of 2 : 1, the final hydrated products were Mg(3)(PO(4))(2) and Ca(3)(PO(4))(2). The MCPB was degradable in Tris-HCl solution and the degradation ratio was obviously higher than calcium phosphate biocement (CPB) because of its fast dissolution. The attachment and proliferation of the MG(63) cells on the MCPB were significantly enhanced in comparison with CPB, and the alkaline phosphatase activity of MG(63) cells on the MCPB was significantly higher than on the CPB at 7 and 14 days. The MG(63) cells with normal phenotype spread well on the MCPB surfaces, and were attached in close proximity to the substrate, as seen by scanning electron microscopy (SEM). The results demonstrated that the MCPB had a good ability to support cell attachment, proliferation and differentiation, and exhibited good cytocompatibility.

Significance of a Posttranslational Modification of the PilA Protein of Geobacter sulfurreducens for Surface Attachment, Biofilm Formation, and Growth on Insoluble Extracellular Electron Acceptors.

PubMed

Richter, Lubna V; Franks, Ashley E; Weis, Robert M; Sandler, Steven J

2017-04-15

Geobacter sulfurreducens , an anaerobic metal-reducing bacterium, possesses type IV pili. These pili are intrinsic structural elements in biofilm formation and, together with a number of c -type cytochromes, are thought to serve as conductive nanowires enabling long-range electron transfer (ET) to metal oxides and graphite anodes. Here, we report that a posttranslational modification of a nonconserved amino acid residue within the PilA protein, the structural subunit of the type IV pili, is crucial for growth on insoluble extracellular electron acceptors. Matrix-assisted laser desorption ionization (MALDI) mass spectrometry of the secreted PilA protein revealed a posttranslational modification of tyrosine-32 with a moiety of a mass consistent with a glycerophosphate group. Mutating this tyrosine into a phenylalanine inhibited cell growth with Fe(III) oxides as the sole electron acceptor. In addition, this amino acid substitution severely diminished biofilm formation on graphite surfaces and impaired current output in microbial fuel cells. These results demonstrate that the capability to attach to insoluble electron acceptors plays a crucial role for the cells' ability to utilize them. The work suggests that glycerophosphate modification of Y32 is a key factor contributing to the surface charge of type IV pili, influencing the adhesion of Geobacter to specific surfaces. IMPORTANCE Type IV pili are bacterial appendages that function in cell adhesion, virulence, twitching motility, and long-range electron transfer (ET) from bacterial cells to insoluble extracellular electron acceptors. The mechanism and role of type IV pili for ET in Geobacter sulfurreducens is still a subject of research. In this study, we identified a posttranslational modification of the major G. sulfurreducens type IV pilin, suggested to be a glycerophosphate moiety. We show that a mutant in which the glycerophosphate-modified tyrosine-32 is replaced with a phenylalanine has reduced abilities for ET and biofilm formation compared with those of the wild type. The results show the importance of the glycerophosphate-modified tyrosine for surface attachment and electron transfer in electrode- or Fe(III)-respiring G. sulfurreducens cells. Copyright © 2017 American Society for Microbiology.

Significance of a Posttranslational Modification of the PilA Protein of Geobacter sulfurreducens for Surface Attachment, Biofilm Formation, and Growth on Insoluble Extracellular Electron Acceptors

PubMed Central

Franks, Ashley E.; Weis, Robert M.; Sandler, Steven J.

2017-01-01

ABSTRACT Geobacter sulfurreducens, an anaerobic metal-reducing bacterium, possesses type IV pili. These pili are intrinsic structural elements in biofilm formation and, together with a number of c-type cytochromes, are thought to serve as conductive nanowires enabling long-range electron transfer (ET) to metal oxides and graphite anodes. Here, we report that a posttranslational modification of a nonconserved amino acid residue within the PilA protein, the structural subunit of the type IV pili, is crucial for growth on insoluble extracellular electron acceptors. Matrix-assisted laser desorption ionization (MALDI) mass spectrometry of the secreted PilA protein revealed a posttranslational modification of tyrosine-32 with a moiety of a mass consistent with a glycerophosphate group. Mutating this tyrosine into a phenylalanine inhibited cell growth with Fe(III) oxides as the sole electron acceptor. In addition, this amino acid substitution severely diminished biofilm formation on graphite surfaces and impaired current output in microbial fuel cells. These results demonstrate that the capability to attach to insoluble electron acceptors plays a crucial role for the cells' ability to utilize them. The work suggests that glycerophosphate modification of Y32 is a key factor contributing to the surface charge of type IV pili, influencing the adhesion of Geobacter to specific surfaces. IMPORTANCE Type IV pili are bacterial appendages that function in cell adhesion, virulence, twitching motility, and long-range electron transfer (ET) from bacterial cells to insoluble extracellular electron acceptors. The mechanism and role of type IV pili for ET in Geobacter sulfurreducens is still a subject of research. In this study, we identified a posttranslational modification of the major G. sulfurreducens type IV pilin, suggested to be a glycerophosphate moiety. We show that a mutant in which the glycerophosphate-modified tyrosine-32 is replaced with a phenylalanine has reduced abilities for ET and biofilm formation compared with those of the wild type. The results show the importance of the glycerophosphate-modified tyrosine for surface attachment and electron transfer in electrode- or Fe(III)-respiring G. sulfurreducens cells. PMID:28138101

Effect of micro-nano-hybrid structured hydroxyapatite bioceramics on osteogenic and cementogenic differentiation of human periodontal ligament stem cell via Wnt signaling pathway.

PubMed

Mao, Lixia; Liu, Jiaqiang; Zhao, Jinglei; Chang, Jiang; Xia, Lunguo; Jiang, Lingyong; Wang, Xiuhui; Lin, Kaili; Fang, Bing

2015-01-01

The surface structure of bioceramic scaffolds is crucial for its bioactivity and osteoinductive ability, and in recent years, human periodontal ligament stem cells have been certified to possess high osteogenic and cementogenic differential ability. In the present study, hydroxyapatite (HA) bioceramics with micro-nano-hybrid surface (mnHA [the hybrid of nanorods and microrods]) were fabricated via hydrothermal reaction of the α-tricalcium phosphate granules as precursors in aqueous solution, and the effects of mnHA on the attachment, proliferation, osteogenic and cementogenic differentiations of human periodontal ligament stem cells as well as the related mechanisms were systematically investigated. The results showed that mnHA bioceramics could promote cell adhesion, proliferation, alkaline phosphatase (ALP) activity, and expression of osteogenic/cementogenic-related markers including runt-related transcription factor 2 (Runx2), ALP, osteocalcin (OCN), cementum attachment protein (CAP), and cementum protein (CEMP) as compared to the HA bioceramics with flat and dense surface. Moreover, mnHA bioceramics stimulated gene expression of low-density lipoprotein receptor-related protein 5 (LRP5) and β-catenin, which are the key genes of canonical Wnt signaling. Moreover, the stimulatory effect on ALP activity and osteogenic and cementogenic gene expression, including that of ALP, OCN, CAP, CEMP, and Runx2 of mnHA bioceramics could be repressed by canonical Wnt signaling inhibitor dickkopf1 (Dkk1). The results suggested that the HA bioceramics with mnHA could act as promising grafts for periodontal tissue regeneration.

The relationship between adult attachment orientation and child self-regulation in eating: The mediating role of persuasive-controlling feeding practices.

PubMed

Powell, Elisabeth M; Frankel, Leslie A; Umemura, Tomo; Hazen, Nancy

2017-08-01

The present study examines the hypothesis that adult attachment orientation, specifically anxious attachment, is related to children's diminished ability to self-regulate their food intake, and that this relationship is mediated by parents' persuasive-controlling feeding practices. Two hundred and sixty five mothers and fathers of preschool children completed online questionnaires that included measures of Adult Attachment Orientation, Parental Persuasive-Controlling Feeding Practices, and Child Self-Regulation of Eating. Structural equation modeling revealed a significant relationship between parental anxious attachment and child self-regulatory abilities, which was fully mediated by parental persuasive-controlling feeding. Also as predicted, parents' avoidant attachment was found to be unrelated to persuasive-controlling feeding and child self-regulated eating. Findings suggest that parents with an anxious attachment orientation may be more likely than other parents to try to use persuasive techniques to control their children's food intake, which may impair children's ability to regulate their food intake, increasing their obesity risk. Implications for intervention are discussed. Copyright © 2017 Elsevier Ltd. All rights reserved.

Diffusible signal factor-repressed extracellular traits enable attachment of Xylella fastidiosa to insect vectors and transmission.

PubMed

Baccari, Clelia; Killiny, Nabil; Ionescu, Michael; Almeida, Rodrigo P P; Lindow, Steven E

2014-01-01

The hypothesis that a wild-type strain of Xylella fastidiosa would restore the ability of rpfF mutants blocked in diffusible signal factor production to be transmitted to new grape plants by the sharpshooter vector Graphocephala atropunctata was tested. While the rpfF mutant was very poorly transmitted by vectors irrespective of whether they had also fed on plants infected with the wild-type strain, wild-type strains were not efficiently transmitted if vectors had fed on plants infected with the rpfF mutant. About 100-fewer cells of a wild-type strain attached to wings of a vector when suspended in xylem sap from plants infected with an rpfF mutant than in sap from uninfected grapes. The frequency of transmission of cells suspended in sap from plants that were infected by the rpfF mutant was also reduced over threefold. Wild-type cells suspended in a culture supernatant of an rpfF mutant also exhibited 10-fold less adherence to wings than when suspended in uninoculated culture media. A factor released into the xylem by rpfF mutants, and to a lesser extent by the wild-type strain, thus inhibits their attachment to, and thus transmission by, sharpshooter vectors and may also enable them to move more readily through host plants.

Interaction between endothelial cells and albumin encapsulated droplets in Poiseuille flow

NASA Astrophysics Data System (ADS)

Seda, Robinson; Fowlkes, J. Brian; Bull, Joseph

2012-11-01

Acoustic droplet vaporization (ADV) of DDFP encapsulated microdroplets has the ability to transform these emulsions into larger gas emboli capable of occluding blood vessels for therapy. An albumin shell is able to stabilize the droplet's superheated core, but can also interact with endothelial cells (EC) at the vessel wall if in close proximity. Radial migration of these microdroplets could bring them close enough to make this interaction possible leading to bioeffects that include cell detachment and death if an ADV event occurs. The purpose of this study is to investigate the hydrodynamic conditions (i.e. shear stresses) that make possible this EC-droplet interaction. A flow chamber coated with a monolayer of EC and connected to a syringe pump is used to flow a DDFP droplet solution at physiological shear stresses (1-50 dyne/cm2) and inspected for droplet attachment. Droplets have been observed to interact and reversibly attach to EC in a static environment, thus it is expected that at low shear stress values interaction and further attachment will be possible. Knowing the flow conditions at which this interaction is likely to occur will aid in preventative measures to avoid significant bioeffects associated with ADV near the vessel wall. This work is supported by NIH grant R01EB006476.

“Ins” and “Outs” of mesenchymal stem cell osteogenesis in regenerative medicine

PubMed Central

Yamaguchi, Dean T

2014-01-01

Repair and regeneration of bone requires mesenchymal stem cells that by self-renewal, are able to generate a critical mass of cells with the ability to differentiate into osteoblasts that can produce bone protein matrix (osteoid) and enable its mineralization. The number of human mesenchymal stem cells (hMSCs) diminishes with age and ex vivo replication of hMSCs has limited potential. While propagating hMSCs under hypoxic conditions may maintain their ability to self-renew, the strategy of using human telomerase reverse transcriptase (hTERT) to allow for hMSCs to prolong their replicative lifespan is an attractive means of ensuring a critical mass of cells with the potential to differentiate into various mesodermal structural tissues including bone. However, this strategy must be tempered by the oncogenic potential of TERT-transformed cells, or their ability to enhance already established cancers, the unknown differentiating potential of high population doubling hMSCs and the source of hMSCs (e.g., bone marrow, adipose-derived, muscle-derived, umbilical cord blood, etc.) that may provide peculiarities to self-renewal, differentiation, and physiologic function that may differ from non-transformed native cells. Tissue engineering approaches to use hMSCs to repair bone defects utilize the growth of hMSCs on three-dimensional scaffolds that can either be a base on which hMSCs can attach and grow or as a means of sequestering growth factors to assist in the chemoattraction and differentiation of native hMSCs. The use of whole native extracellular matrix (ECM) produced by hMSCs, rather than individual ECM components, appear to be advantageous in not only being utilized as a three-dimensional attachment base but also in appropriate orientation of cells and their differentiation through the growth factors that native ECM harbor or in simulating growth factor motifs. The origin of native ECM, whether from hMSCs from young or old individuals is a critical factor in “rejuvenating” hMSCs from older individuals grown on ECM from younger individuals. PMID:24772237

Fast Response, Open-Celled Porous, Shape Memory Effect Actuators with Integrated Attachments

NASA Technical Reports Server (NTRS)

Jardine, Andrew Peter (Inventor)

2015-01-01

This invention relates to the exploitation of porous foam articles exhibiting the Shape Memory Effect as actuators. Each foam article is composed of a plurality of geometric shapes, such that some geometric shapes can fit snugly into or around rigid mating connectors that attach the Shape Memory foam article intimately into the load path between a static structure and a moveable structure. The foam is open-celled, composed of a plurality of interconnected struts whose mean diameter can vary from approximately 50 to 500 microns. Gases and fluids flowing through the foam transfer heat rapidly with the struts, providing rapid Shape Memory Effect transformations. Embodiments of porous foam articles as torsional actuators and approximately planar structures are disposed. Simple, integral connection systems exploiting the ability to supply large loads to a structure, and that can also supply hot and cold gases and fluids to effect rapid actuation are also disposed.

Effect of the surfactant Tween 80 on the detachment and dispersal of Salmonella enterica Thompson single cells and aggregates from cilantro leaves as revealed by image analysis

USDA-ARS?s Scientific Manuscript database

Biofilms formed by human enteric pathogens on plants are a great concern to the produce industry. Salmonella enterica has the ability to form biofilms and large aggregates on leaf surfaces, including on cilantro leaves. Aggregates that remained attached after rigorous washing of cilantro leaves and ...

Patient satisfaction and masticatory efficiency of single implant-retained mandibular overdentures using the stud and magnetic attachments.

PubMed

Cheng, Tao; Sun, Guifeng; Huo, Jingyu; He, Xiaoji; Wang, Yining; Ren, Yan-Fang

2012-11-01

To study patient satisfaction and masticatory efficiency of single implant-retained mandibular overdentures using the stud and magnetic attachments in a randomized clinical trial with a crossover design. Patients received a single implant placed in the midline of the mandible and either a stud (Locator) or a magnetic (Magfit) attachment, assigned at random. Patient satisfaction, including patient comfort, speech, chewing ability and retention, and masticatory efficiency measured by chewing peanuts, were assessed before and 3 months after attachment insertion. Patient satisfaction and masticatory efficiency were evaluated again 3 months after insertion of the alternate attachment bodies. The outcomes were compared before and after insertion of the attachments and between the two types of attachments using Wilcoxon signed rank tests. Patient overall satisfaction, comfort, speech, chewing ability, and retention improved significantly after insertion of both types of attachment bodies (p<0.05). Masticatory efficiencies also increased in both the Locator and the Magfit groups (p<0.05). There were no statistically significant differences in patient overall satisfaction, comfort, speech, and retention between the two types of attachments (p>0.05). The Locator attachments performed better in perceived chewing ability than the Magfit (p<0.05), but there was no statistically significant difference in masticatory efficiency between the two attachment types (p>0.05). Clinical outcomes were significantly improved in single implant-retained mandibular overdentures using either the Locator or the Magfit magnetic attachments. There was no difference in masticatory efficiency between the two attachment types. Copyright © 2012 Elsevier Ltd. All rights reserved.

Application of nanostructured biochips for efficient cell transfection microarrays

NASA Astrophysics Data System (ADS)

Akkamsetty, Yamini; Hook, Andrew L.; Thissen, Helmut; Hayes, Jason P.; Voelcker, Nicolas H.

2007-01-01

Microarrays, high-throughput devices for genomic analysis, can be further improved by developing materials that are able to manipulate the interfacial behaviour of biomolecules. This is achieved both spatially and temporally by smart materials possessing both switchable and patterned surface properties. A system had been developed to spatially manipulate both DNA and cell growth based upon the surface modification of highly doped silicon by plasma polymerisation and polyethylene grafting followed by masked laser ablation for formation of a pattered surface with both bioactive and non-fouling regions. This platform has been successfully applied to transfected cell microarray applications with the parallel expression of genes by utilising its ability to direct and limit both DNA and cell attachment to specific sites. One of the greatest advantages of this system is its application to reverse transfection, whereupon by utilising the switchable adsorption and desorption of DNA using a voltage bias, the efficiency of cell transfection can be enhanced. However, it was shown that application of a voltage also reduces the viability of neuroblastoma cells grown on a plasma polymer surface, but not human embryonic kidney cells. This suggests that the application of a voltage may not only result in the desorption of bound DNA but may also affect attached cells. The characterisation of a DNA microarray by contact printing has also been investigated.

Plasmid-dependent attachment of Agrobacterium tumefaciens to plant tissue culture cells.

PubMed

Matthysse, A G; Wyman, P M; Holmes, K V

1978-11-01

Kinetic, microscopic, and biochemical studies show that virulent Ti (tumor inducing)-plasmid-containing strains of Agrobacterium attach to normal tobacco and carrot tissue culture cells. Kinetic studies showed that virulent strains of A. tumefaciens attach to the plant tissue culture cells in increasing numbers during the first 1 to 2 h of incubation of the bacteria with the plant cells. Five Ti-plasmid-containing virulent Agrobacterium strains showed greater attachment to tobacco cells than did five avirulent strains. Light and scanning electron microscopic observations confirmed that virulent strains showed little attachment. Bacterial attachment was blocked by prior incubation of the plant cells with lipopolysaccharide extracted from A. tumefaciens, but not from A. radiobacter, suggesting that bacterial lipopolysaccharide is one of the components involved in the attachment process. At least one other bacterial product may be required for attachment in tissue culture because the virulent A. tumefaciens NT1, which lacks the Ti plasmid, does not itself attach to tobacco cells, but its lipopolysaccharide does inhibit the attachment of virulent strains.

Contribution of Spores to the Ability of Clostridium difficile To Adhere to Surfaces

PubMed Central

Joshi, Lovleen Tina; Phillips, Daniel S.; Williams, Catrin F.; Alyousef, Abdullah

2012-01-01

Clostridium difficile is the commonest cause of hospital-acquired infection in the United Kingdom. We characterized the abilities of 21 clinical isolates to form spores; to adhere to inorganic and organic surfaces, including stainless steel and human adenocarcinoma cells; and to germinate. The composition of culture media had a significant effect on spore formation, as significantly more spores were produced in brain heart infusion broth (Student's t test; P = 0.018). The spore surface relative hydrophobicity (RH) varied markedly (14 to 77%) and was correlated with the ability to adhere to stainless steel. We observed no correlation between the ribotype and the ability to adhere to steel. When the binding of hydrophobic (DS1813; ribotype 027; RH, 77%) and hydrophilic (DS1748; ribotype 002; RH, 14%) spores to human gut epithelial cells at different stages of cell development was examined, DS1813 spores adhered more strongly, suggesting the presence of surface properties that aid attachment to human cells. Electron microscopy studies revealed the presence of an exosporium surrounding DS1813 spores that was absent from spores of DS1748. Finally, the ability of spores to germinate was found to be strain and medium dependent. While the significance of these findings to the disease process has yet to be determined, this study has highlighted the importance of analyzing multiple isolates when attempting to characterize the behavior of a bacterial species. PMID:22923404

A novel member of the SAF (scaffold attachment factor)-box protein family inhibits gene expression and induces apoptosis

PubMed Central

Chan, Ching Wan; Lee, Youn-Bok; Uney, James; Flynn, Andrea; Tobias, Jonathan H.; Norman, Michael

2007-01-01

The SLTM [SAF (scaffold attachment factor)-like transcription modulator] protein contains a SAF-box DNA-binding motif and an RNA-binding domain, and shares an overall identity of 34% with SAFB1 {scaffold attachment factor-B1; also known as SAF-B (scaffold attachment factor B), HET [heat-shock protein 27 ERE (oestrogen response element) and TATA-box-binding protein] or HAP (heterogeneous nuclear ribonucleoprotein A1-interacting protein)}. Here, we show that SLTM is localized to the cell nucleus, but excluded from nucleoli, and to a large extent it co-localizes with SAFB1. In the nucleus, SLTM has a punctate distribution and it does not co-localize with SR (serine/arginine) proteins. Overexpression of SAFB1 has been shown to exert a number of inhibitory effects, including suppression of oestrogen signalling. Although SLTM also suppressed the ability of oestrogen to activate a reporter gene in MCF-7 breast-cancer cells, inhibition of a constitutively active β-galactosidase gene suggested that this was primarily the consequence of a generalized inhibitory effect on transcription. Measurement of RNA synthesis, which showed a particularly marked inhibition of [3H]uridine incorporation into mRNA, supported this conclusion. In addition, analysis of cell-cycle parameters, chromatin condensation and cytochrome c release showed that SLTM induced apoptosis in a range of cultured cell lines. Thus the inhibitory effects of SLTM on gene expression appear to result from generalized down-regulation of mRNA synthesis and initiation of apoptosis consequent upon overexpressing the protein. While indicating a crucial role for SLTM in cellular function, these results also emphasize the need for caution when interpreting phenotypic changes associated with manipulation of protein expression levels. PMID:17630952

Adhesion of Biodegradative Anaerobic Bacteria to Solid Surfaces

PubMed Central

van Schie, Paula M.; Fletcher, Madilyn

1999-01-01

In order to exploit the ability of anaerobic bacteria to degrade certain contaminants for bioremediation of polluted subsurface environments, we need to understand the mechanisms by which such bacteria partition between aqueous and solid phases, as well as the environmental conditions that influence partitioning. We studied four strictly anaerobic bacteria, Desulfomonile tiedjei, Syntrophomonas wolfei, Syntrophobacter wolinii, and Desulfovibrio sp. strain G11, which theoretically together can constitute a tetrachloroethylene- and trichloroethylene-dechlorinating consortium. Adhesion of these organisms was evaluated by microscopic determination of the numbers of cells that attached to glass coverslips exposed to cell suspensions under anaerobic conditions. We studied the effects of the growth phase of the organisms on adhesion, as well as the influence of electrostatic and hydrophobic properties of the substratum. Results indicate that S. wolfei adheres in considerably higher numbers to glass surfaces than the other three organisms. Starvation greatly decreases adhesion of S. wolfei and Desulfovibrio sp. strain G11 but seems to have less of an effect on the adhesion of the other bacteria. The presence of Fe3+ on the substratum, which would be electropositive, significantly increased the adhesion of S. wolfei, whereas the presence of silicon hydrophobic groups decreased the numbers of attached cells of all species. Measurements of transport of cells through hydrophobic-interaction and electrostatic-interaction columns indicated that all four species had negatively charged cell surfaces and that D. tiedjei and Desulfovibrio sp. strain G11 possessed some hydrophobic cell surface properties. These findings are an early step toward understanding the dynamic attachment of anaerobic bacteria in anoxic environments. PMID:10543826

Physical Properties and Cellular Responses to Crosslinkable Poly(Propylene Fumarate)/Hydroxyapatite Nanocomposites

PubMed Central

Lee, Kee-Won; Wang, Shanfeng; Yaszemski, Michael J.; Lu, Lichun

2008-01-01

A series of crosslinkable nanocomposites has been developed using hydroxyapatite (HA) nanoparticles and poly(propylene fumarate) (PPF). PPF/HA nanocomposites with four different weight fractions of HA nanoparticles have been characterized in terms of thermal and mechanical properties. To assess surface chemistry of crosslinked PPF/HA nanocomposites, their hydrophilicity and capability of adsorbing proteins have been determined using static contact angle measurement and MicroBCA protein assay kit after incubation with 10% fetal bovine serum (FBS), respectively. In vitro cell studies have been performed using MC3T3-E1 mouse pre-osteoblast cells to investigate the ability of PPF/HA nanocomposites to support cell attachment, spreading, and proliferation after 1, 4, and 7 days. By adding HA nanoparticles to PPF, the mechanical properties of crosslinked PPF/HA nanocomposites have not been increased due to the initially high modulus of crosslinked PPF. However, hydrophilicity and serum protein adsorption on the surface of nanocomposites have been significantly increased, resulting in enhanced cell attachment, spreading, and proliferation after 4 days of cell seeding. These results indicate that crosslinkable PPF/HA nanocomposites are useful for hard tissue replacement because of excellent mechanical strength and osteoconductivity. PMID:18403013

Attachment, invasion, chemotaxis, and proteinase expression of B16-BL6 melanoma cells exhibiting a low metastatic phenotype after exposure to dietary restriction of tyrosine and phenylalanine.

PubMed

Uhlenkott, C E; Huijzer, J C; Cardeiro, D J; Elstad, C A; Meadows, G G

1996-03-01

We previously reported that low levels of tyrosine (Tyr) and phenylalanine (Phe) alter the metastatic phenotype of B16-BL6 (BL6) murine melanoma and select for tumor cell populations with decreased lung colonizing ability. To more specifically characterize the effects of Tyr and Phe restriction on the malignant phenotype of BL6, we investigated in vitro attachment, invasion, proteinase expression, and chemotaxis of high and low metastatic BL6 variants. High metastatic variant cells were isolated from subcutaneous tumors of mice fed a nutritionally complete diet (ND cells) and low metastatic variant cells were isolated from mice fed a diet restricted in Tyr and Phe (LTP cells). Results indicate that attachment to reconstituted basement membrane (Matrigel) was significantly reduced in LTP cells as compared to ND cells. Attachment to collagen IV, laminin, and fibronectin were similar between the two variants. Invasion through Matrigel and growth factor-reduced Matrigel were significantly decreased in LTP cells as compared to ND cells. Zymography revealed the presence of M(r) 92,000 and M(r) 72,000 progelatinases, tissue plasminogen activator, and urokinase plasminogen activator in the conditioned medium of both variants; however, there were no differences in activity of these secreted proteinases between the two variants. Growth of the variants on growth factor-reduced Matrigel similarly induced expression of the M(r) 92,000 progelatinase. The variants exhibited similar chemotactic responses toward laminin. However, the chemotactic response toward fibronectin by LTP cells was significantly increased. MFR5, a monoclonal antibody which selectively blocks function of the alpha 5 chain of the alpha 5 beta 1 integrin, VLA-5, decreased the chemotactic response toward fibronectin of ND cells by 37%; the chemotactic response by LTP cells was reduced by 49%. This effect was specific for fibronectin-mediated chemotaxis since the chemotaxis toward laminin and invasion through Matrigel were not altered by the presence of MFR5. The surface expression of VLA-5 was significantly increased in LTP cells as compared to ND cells by flow cytometric analysis. These observations suggest that limitation of Tyr and Phe either directly modifies BL6 or selects for subpopulations with altered in vitro invasion, chemotaxis, and integrin expression.

Epigallocatechin Gallate-Modified Gelatin Sponges Treated by Vacuum Heating as a Novel Scaffold for Bone Tissue Engineering.

PubMed

Honda, Yoshitomo; Takeda, Yoshihiro; Li, Peiqi; Huang, Anqi; Sasayama, Satoshi; Hara, Eiki; Uemura, Naoya; Ueda, Mamoru; Hashimoto, Masanori; Arita, Kenji; Matsumoto, Naoyuki; Hashimoto, Yoshiya; Baba, Shunsuke; Tanaka, Tomonari

2018-04-11

Chemical modification of gelatin using epigallocatechin gallate (EGCG) promotes bone formation in vivo. However, further improvements are required to increase the mechanical strength and bone-forming ability of fabricated EGCG-modified gelatin sponges (EGCG-GS) for practical applications in regenerative therapy. In the present study, we investigated whether vacuum heating-induced dehydrothermal cross-linking of EGCG-GS enhances bone formation in critical-sized rat calvarial defects. The bone-forming ability of vacuum-heated EGCG-GS (vhEGCG-GS) and other sponges was evaluated by micro-computed tomography and histological staining. The degradation of sponges was assessed using protein assays, and cell morphology and proliferation were verified by scanning electron microscopy and immunostaining using osteoblastic UMR106 cells in vitro. Four weeks after the implantation of sponges, greater bone formation was detected for vhEGCG-GS than for EGCG-GS or vacuum-heated gelatin sponges (dehydrothermal cross-linked sponges without EGCG). In vitro experiments revealed that the relatively low degradability of vhEGCG-GS supports cell attachment, proliferation, and cell-cell communication on the matrix. These findings suggest that vacuum heating enhanced the bone forming ability of EGCG-GS, possibly via the dehydrothermal cross-linking of EGCG-GS, which provides a scaffold for cells, and by maintaining the pharmacological effect of EGCG.

An Ribonuclease T2 Family Protein Modulates Acinetobacter baumannii Abiotic Surface Colonization

PubMed Central

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

2014-01-01

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

A Chemically Defined Carrier for the Delivery of Human Mesenchymal Stem/Stromal Cells to Skin Wounds

PubMed Central

Walker, Nathan G.; Mistry, Anita R.; Smith, Louise E.; Eves, Paula C.; Tsaknakis, Grigorios; Forster, Simon; Watt, Suzanne M.

2012-01-01

Skin has a remarkable capacity for regeneration, but age- and diabetes-related vascular problems lead to chronic non-healing wounds for many thousands of U.K. patients. There is a need for new therapeutic approaches to treat these resistant wounds. Donor mesenchymal stem/stromal cells (MSCs) have been shown to assist cutaneous wound healing by accelerating re-epithelialization. The aim of this work was to devise a low risk and convenient delivery method for transferring these cells to wound beds. Plasma polymerization was used to functionalize the surface of medical-grade silicone with acrylic acid. Cells attached well to these carriers, and culture for up to 3 days on the carriers did not significantly affect their phenotype or ability to support vascular tubule formation. These carriers were then used to transfer MSCs onto human dermis. Cell transfer was confirmed using an MTT assay to assess viable cell numbers and enhanced green fluorescent protein–labeled MSCs to demonstrate that the cells post-transfer attached to the dermis. We conclude that this synthetic carrier membrane is a promising approach for delivery of therapeutic MSCs and opens the way for future studies to evaluate its impact on repairing difficult skin wounds. PMID:21943098

Plasmid-dependent attachment of Agrobacterium tumefaciens to plant tissue culture cells.

PubMed Central

Matthysse, A G; Wyman, P M; Holmes, K V

1978-01-01

Kinetic, microscopic, and biochemical studies show that virulent Ti (tumor inducing)-plasmid-containing strains of Agrobacterium attach to normal tobacco and carrot tissue culture cells. Kinetic studies showed that virulent strains of A. tumefaciens attach to the plant tissue culture cells in increasing numbers during the first 1 to 2 h of incubation of the bacteria with the plant cells. Five Ti-plasmid-containing virulent Agrobacterium strains showed greater attachment to tobacco cells than did five avirulent strains. Light and scanning electron microscopic observations confirmed that virulent strains showed little attachment. Bacterial attachment was blocked by prior incubation of the plant cells with lipopolysaccharide extracted from A. tumefaciens, but not from A. radiobacter, suggesting that bacterial lipopolysaccharide is one of the components involved in the attachment process. At least one other bacterial product may be required for attachment in tissue culture because the virulent A. tumefaciens NT1, which lacks the Ti plasmid, does not itself attach to tobacco cells, but its lipopolysaccharide does inhibit the attachment of virulent strains. Images PMID:730370

Cell adhesion pattern created by OSTE polymers.

PubMed

Liu, Wenjia; Li, Yiyang; Ding, Xianting

2017-04-24

Engineering surfaces with functional polymers is a crucial issue in the field of micro/nanofabrication and cell-material interface studies. For many applications of surface patterning, it does not need cells to attach on the whole surface. Herein, we introduce a novel polymer fabrication protocol of off-stoichiometry thiol-ene (OSTE) polymers to create heterogeneity on the surface by utilizing 3D printing and soft-lithography. By choosing two OSTE polymers with different functional groups, we create a pattern where only parts of the surface can facilitate cell adhesion. We also study the hydrophilic property of OSTE polymers by mixing poly(ethylene glycol) (PEG) directly with pre-polymers and plasma treatments afterwards. Moreover, we investigate the effect of functional groups' excess ratio and hydrophilic property on the cell adhesion ability of OSTE polymers. The results show that the cell adhesion ability of OSTE materials can be tuned within a wide range by the coupling effect of functional groups' excess ratio and hydrophilic property. Meanwhile, by mixing PEG with pre-polymers and undergoing oxygen plasma treatment afterward can significantly improve the hydrophilic property of OSTE polymers.

Attachment of Salmonella strains to a plant cell wall model is modulated by surface characteristics and not by specific carbohydrate interactions.

PubMed

Tan, Michelle Sze-Fan; Moore, Sean C; Tabor, Rico F; Fegan, Narelle; Rahman, Sadequr; Dykes, Gary A

2016-09-15

Processing of fresh produce exposes cut surfaces of plant cell walls that then become vulnerable to human foodborne pathogen attachment and contamination, particularly by Salmonella enterica. Plant cell walls are mainly composed of the polysaccharides cellulose, pectin and hemicelluloses (predominantly xyloglucan). Our previous work used bacterial cellulose-based plant cell wall models to study the interaction between Salmonella and the various plant cell wall components. We demonstrated that Salmonella attachment was favoured in the presence of pectin while xyloglucan had no effect on its attachment. Xyloglucan significantly increased the attachment of Salmonella cells to the plant cell wall model only when it was in association with pectin. In this study, we investigate whether the plant cell wall polysaccharides mediate Salmonella attachment to the bacterial cellulose-based plant cell wall models through specific carbohydrate interactions or through the effects of carbohydrates on the physical characteristics of the attachment surface. We found that none of the monosaccharides that make up the plant cell wall polysaccharides specifically inhibit Salmonella attachment to the bacterial cellulose-based plant cell wall models. Confocal laser scanning microscopy showed that Salmonella cells can penetrate and attach within the tightly arranged bacterial cellulose network. Analysis of images obtained from atomic force microscopy revealed that the bacterial cellulose-pectin-xyloglucan composite with 0.3 % (w/v) xyloglucan, previously shown to have the highest number of Salmonella cells attached to it, had significantly thicker cellulose fibrils compared to other composites. Scanning electron microscopy images also showed that the bacterial cellulose and bacterial cellulose-xyloglucan composites were more porous when compared to the other composites containing pectin. Our study found that the attachment of Salmonella cells to cut plant cell walls was not mediated by specific carbohydrate interactions. This suggests that the attachment of Salmonella strains to the plant cell wall models were more dependent on the structural characteristics of the attachment surface. Pectin reduces the porosity and space between cellulose fibrils, which then forms a matrix that is able to retain Salmonella cells within the bacterial cellulose network. When present with pectin, xyloglucan provides a greater surface for Salmonella cells to attach through the thickening of cellulose fibrils.

Alginate-Encapsulation for the Improved Hypothermic Preservation of Human Adipose-Derived Stem Cells.

PubMed

Swioklo, Stephen; Constantinescu, Andrei; Connon, Che J

2016-03-01

Despite considerable progress within the cell therapy industry, unmet bioprocessing and logistical challenges associated with the storage and distribution of cells between sites of manufacture and the clinic exist. We examined whether hypothermic (4°C-23°C) preservation of human adipose-derived stem cells could be improved through their encapsulation in 1.2% calcium alginate. Alginate encapsulation improved the recovery of viable cells after 72 hours of storage. Viable cell recovery was highly temperature-dependent, with an optimum temperature of 15°C. At this temperature, alginate encapsulation preserved the ability for recovered cells to attach to tissue culture plastic on rewarming, further increasing its effect on total cell recovery. On attachment, the cells were phenotypically normal, displayed normal growth kinetics, and maintained their capacity for trilineage differentiation. The number of cells encapsulated (up to 2 × 10(6) cells per milliliter) did not affect viable cell recovery nor did storage of encapsulated cells in a xeno-free, serum-free,current Good Manufacturing Practice-grade medium. We present a simple, low-cost system capable of enhancing the preservation of human adipose-derived stem cells stored at hypothermic temperatures, while maintaining their normal function. The storage of cells in this manner has great potential for extending the time windows for quality assurance and efficacy testing, distribution between the sites of manufacture and the clinic, and reducing the wastage associated with the limited shelf life of cells stored in their liquid state. ©AlphaMed Press.

Protein attachment to silane-functionalized porous silicon: A comparison of electrostatic and covalent attachment.

PubMed

Baranowska, Malgorzata; Slota, Agata J; Eravuchira, Pinkie J; Alba, Maria; Formentin, Pilar; Pallarès, Josep; Ferré-Borrull, Josep; Marsal, Lluís F

2015-08-15

Porous silicon (pSi) is a prosperous biomaterial, biocompatible, and biodegradable. Obtaining regularly functionalized pSi surfaces is required in many biotechnology applications. Silane-PEG-NHS (triethoxysilane-polyethylene-glycol-N-hydroxysuccinimide) is useful for single-molecule studies due to its ability to attach to only one biomolecule. We investigate the functionalization of pSi with silane-PEG-NHS and compare it with two common grafting agents: APTMS (3-aminopropylotrimethoxysilane) as electrostatic linker, and APTMS modified with glutaraldehyde as covalent spacer. We show the arrangement of two proteins (collagen and bovine serum albumin) as a function of the functionalization and of the pore size. FTIR is used to demonstrate correct functionalization while fluorescence confocal microscopy reveals that silane-PEG-NHS results in a more uniform protein distribution. Reflection interference spectroscopy (RIfS) is used to estimate the attachment of linker and proteins. The results open a way to obtain homogenous chemical modified silicon supports with a great value in biosensing, drug delivery and cell biology. Copyright © 2015 Elsevier Inc. All rights reserved.

Differentiation and characteristics of the enhanced green fluorescent protein gene transgenic goat neural stem cells cultured in attached and non-attached plates.

PubMed

Zheng, Yue-Mao; Dang, Yong-Hui; Qiu, Shuang; Qi, Ying-Pei; Xu, Yong-Ping; Sai, Wu-Jia-Fu

2011-08-01

The aims of this study were (i) to determine whether NSCs (neural stem cells) could be isolated from the brain of embryonic day 98 fetal goat, (ii) to determine if these stem cells have the capability of multipotent differentiation following transfection with a reporter gene, EGFP (enhanced green fluorescent protein) and (iii) to study the characteristics of the stem cells cultured in attached and non-attached plates. NSCs were isolated from embryonic day 98 fetal goat brain, transfected with EGFP gene using lipofection, and subcultured in attached and non-attached plates respectively. The transgenic stem cells were induced to differentiate into osteogenic and endothelial cells in vitro respectively. Markers associated with undifferentiated NSCs and their differentiated cells were tested by RT-PCR (reverse transcription-PCR). The results demonstrated that stem cells could be isolated from embryonic day 98 fetal goat brain, and EGFP gene could be transfected into the cells. The transgenic NSCs were capable of self-renewal, a defining property of stem cells, and were grown as free-floating neurospheres in non-attached plates. When the neurospheres were transferred and cultured in attached plates, cells migrate from the neurospheres and are grown as spindle cells. The stem cells were grown as quasi-circular cells when the single stem cells were cultured in attached plates. Both the NSCs cultured in non-attached and attached plates could express Hes1 (hairy and enhancer of split 1), Oct4 (octamer-binding protein 4), Nanog, Sox2 [SRY (sex-determining region Y)-box 2] and Nestin, while following differentiation cells expressed markers for osteogenic cells (Osteocalcin+ and Osteonectin+) and endothelium (CD34+ and eNOS+). The results demonstrated that the goat EGFP gene transgenic NSCs have the capability of multipotent differentiation, which means that the transgenic NSCs may be useful in cell transplantation studies in future.

Effect of micro-nano-hybrid structured hydroxyapatite bioceramics on osteogenic and cementogenic differentiation of human periodontal ligament stem cell via Wnt signaling pathway

PubMed Central

Mao, Lixia; Liu, Jiaqiang; Zhao, Jinglei; Chang, Jiang; Xia, Lunguo; Jiang, Lingyong; Wang, Xiuhui; Lin, Kaili; Fang, Bing

2015-01-01

The surface structure of bioceramic scaffolds is crucial for its bioactivity and osteoinductive ability, and in recent years, human periodontal ligament stem cells have been certified to possess high osteogenic and cementogenic differential ability. In the present study, hydroxyapatite (HA) bioceramics with micro-nano-hybrid surface (mnHA [the hybrid of nanorods and microrods]) were fabricated via hydrothermal reaction of the α-tricalcium phosphate granules as precursors in aqueous solution, and the effects of mnHA on the attachment, proliferation, osteogenic and cementogenic differentiations of human periodontal ligament stem cells as well as the related mechanisms were systematically investigated. The results showed that mnHA bioceramics could promote cell adhesion, proliferation, alkaline phosphatase (ALP) activity, and expression of osteogenic/cementogenic-related markers including runt-related transcription factor 2 (Runx2), ALP, osteocalcin (OCN), cementum attachment protein (CAP), and cementum protein (CEMP) as compared to the HA bioceramics with flat and dense surface. Moreover, mnHA bioceramics stimulated gene expression of low-density lipoprotein receptor-related protein 5 (LRP5) and β-catenin, which are the key genes of canonical Wnt signaling. Moreover, the stimulatory effect on ALP activity and osteogenic and cementogenic gene expression, including that of ALP, OCN, CAP, CEMP, and Runx2 of mnHA bioceramics could be repressed by canonical Wnt signaling inhibitor dickkopf1 (Dkk1). The results suggested that the HA bioceramics with mnHA could act as promising grafts for periodontal tissue regeneration. PMID:26648716

Swimming Motility Reduces Deposition to Silica Surfaces

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

Lu, Nanxi; Massoudieh, Arash; Liang, Xiaomeng

The role of swimming motility on bacterial transport and fate in porous media was evaluated. We present microscopic evidence showing that strong swimming motility reduces attachment of Azotobacter vinelandii cells to silica surfaces. Applying global and cluster statistical analyses to microscopic videos taken under non-flow conditions, wild type, flagellated A. vinelandii strain DJ showed strong swimming ability with an average speed of 13.1 μm/s, DJ77 showed impaired swimming averaged at 8.7 μm/s, and both the non-flagellated JZ52 and chemically treated DJ cells were non-motile. Quantitative analyses of trajectories observed at different distances above the collector of a radial stagnation pointmore » flow cell (RSPF) revealed that both swimming and non-swimming cells moved with the flow when at a distance of at least 20 μm from the collector surface. Near the surface, DJ cells showed both horizontal and vertical movement diverging them from reaching surfaces, while chemically treated DJ cells moved with the flow to reach surfaces, suggesting that strong swimming reduced attachment. In agreement with the RSPF results, the deposition rates obtained for two-dimensional multiple-collector micromodels were also lowest for DJ, while DJ77 and JZ52 showed similar values. Strong swimming specifically reduced deposition on the upstream surfaces of the micromodel collectors.« less

Advanced tissue engineering scaffold design for regeneration of the complex hierarchical periodontal structure.

PubMed

Costa, Pedro F; Vaquette, Cédryck; Zhang, Qiyi; Reis, Rui L; Ivanovski, Saso; Hutmacher, Dietmar W

2014-03-01

This study investigated the ability of an osteoconductive biphasic scaffold to simultaneously regenerate alveolar bone, periodontal ligament and cementum. A biphasic scaffold was built by attaching a fused deposition modelled bone compartment to a melt electrospun periodontal compartment. The bone compartment was coated with a calcium phosphate (CaP) layer for increasing osteoconductivity, seeded with osteoblasts and cultured in vitro for 6 weeks. The resulting constructs were then complemented with the placement of PDL cell sheets on the periodontal compartment, attached to a dentin block and subcutaneously implanted into athymic rats for 8 weeks. Scanning electron microscopy, X-ray diffraction, alkaline phosphatase and DNA content quantification, confocal laser microscopy, micro computerized tomography and histological analysis were employed to evaluate the scaffold's performance. The in vitro study showed that alkaline phosphatase activity was significantly increased in the CaP-coated samples and they also displayed enhanced mineralization. In the in vivo study, significantly more bone formation was observed in the coated scaffolds. Histological analysis revealed that the large pore size of the periodontal compartment permitted vascularization of the cell sheets, and periodontal attachment was achieved at the dentin interface. This work demonstrates that the combination of cell sheet technology together with an osteoconductive biphasic scaffold could be utilized to address the limitations of current periodontal regeneration techniques. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Interaction of Chlamydia trachomatis organisms and HeLa 229 cells.

PubMed Central

Kuo, C C; Grayston, T

1976-01-01

The infection of HeLa 229 cells in monolayer culture with trachoma (B/TW-5/OT) and lymphogranuloma venereum (LGV) (L2/434/Bu) organism was studied in terms of two parameters: radioactivity counts of cell-associated tritium labeled organisms at the initial stage of inoculation for measurement of attachment, and inclusion counts of infection cells after incubation for measurement of growth. Factors affecting attachment and inclusion formation and correlation of the two are presented. It was shown that attachment is an important initial step in infection by Chlamydia trachomatis. The rate of attachment was temperature dependent. The attachment of LGV organisms was affected more profoundly by temperature than was that of trachoma organisms. Attachment and inclusion formation of trachoma and LGV organisms were inhibited by heparin. Diethylaminoethyl-dextran was again shown to enhance attachment and inclusion formation of trachoma but not LGV organisms. NaF had no effect on attachment, but inhibited inclusion formation of both trachoma and LGV organisms. Both attachment and inclusion formation of trachoma organisms were strongly enhanced by centrifugation of the inoculum onto the cell monolayer. Although inclusion formation of trachoma organism was much greater in susceptible cells (HeLa 229) than relatively insusceptible cells (fetal tonsil), attachment was only slightly greater. The results based on the test of two cell lines suggested that attachment prpbably is not a critical factor in determing a cell line's susceptibility to infection with trachoma organisms. PMID:179950

Chlorine-rich plasma polymer coating for the prevention of attachment of pathogenic fungal cells onto materials surfaces

NASA Astrophysics Data System (ADS)

Lamont-Friedrich, Stephanie J.; Michl, Thomas D.; Giles, Carla; Griesser, Hans J.; Coad, Bryan R.

2016-07-01

The attachment of pathogenic fungal cells onto materials surfaces, which is often followed by biofilm formation, causes adverse consequences in a wide range of areas. Here we have investigated the ability of thin film coatings from chlorinated molecules to deter fungal colonization of solid materials by contact killing of fungal cells reaching the surface of the coating. Coatings were deposited onto various substrate materials via plasma polymerization, which is a substrate-independent process widely used for industrial coating applications, using 1,1,2-trichloroethane as the process vapour. XPS surface analysis showed that the coatings were characterized by a highly chlorinated hydrocarbon polymer nature, with only a very small amount of oxygen incorporated. The activity of these coatings against human fungal pathogens was quantified using a recently developed, modified yeast assay and excellent antifungal activity was observed against Candida albicans and Candida glabrata. Plasma polymer surface coatings derived from chlorinated hydrocarbon molecules may therefore offer a promising solution to preventing yeast and mould biofilm formation on materials surfaces, for applications such as air conditioners, biomedical devices, food processing equipment, and others.

Novel Architectures for Achieving Direct Electron Transfer in Enzymatic Biofuel Cells

NASA Astrophysics Data System (ADS)

Blaik, Rita A.

Enzymatic biofuel cells are a promising source of alternative energy for small device applications, but still face the challenge of achieving direct electron transfer with high enzyme concentrations in a simple system. In this dissertation, methods of constructing electrodes consisting of enzymes attached to nanoparticle-enhanced substrates that serve as high surface area templates are evaluated. In the first method described, glucose oxidase is covalently attached to gold nanoparticles that are assembled onto genetically engineered M13 bacteriophage. The resulting anodes achieve a high peak current per area and a significant improvement in enzyme surface coverage. In the second system, fructose dehydrogenase, a membrane-bound enzyme that has the natural ability to achieve direct electron transfer, is immobilized into a matrix consisting of binders and carbon nanotubes to extend the lifetime of the anode. For the cathode, bilirubin oxidase is immobilized in a carbon nanotube and sol-gel matrix to achieve direct electron transfer. Finally, a full fuel cell consisting of both an anode and cathode is constructed and evaluated with each system described.

A DNA Aptamer Against Influenza A Virus: An Effective Inhibitor to the Hemagglutinin-Glycan Interactions.

PubMed

Li, Wenkai; Feng, Xinru; Yan, Xing; Liu, Keyi; Deng, Le

2016-06-01

Most therapeutical nucleic acid aptamers tend to inhibit protein-protein interactions and thereby function as antagonists. Attachment of the influenza virus surface glycoprotein hemagglutinin (HA) to sialic acid-containing host cell receptors (glycan) facilitates the initial stage of viral infection. Inhibition of the attachment may result in an antiviral effect on the proliferation of the influenza virus. To develop therapeutically interesting agents, we selected two single-stranded DNA (ssDNA) aptamers specific to the HA protein of H1N1 influenza virus (A/Puerto Rico/8/1934) through a procedure of systematic evolution of ligands by exponential enrichment. As it showed a higher binding affinity for HA protein (Kd = 78 ± 1 nM), aptamer 1 was tested for its ability to interfere with HA-glycan interactions using chicken red blood cell hemagglutination and microneutralization assays, which demonstrated that it significantly suppressed the viral infection in host cells. These results indicate that the isolated ssDNA aptamer may be developed as an antiviral agent against influenza through appropriate therapeutic formulation.

Sharing information about cancer with one's family is associated with improved quality of life.

PubMed

Lai, Carlo; Borrelli, Beatrice; Ciurluini, Paola; Aceto, Paola

2017-10-01

The aim of this study was to investigate the association between cancer patients' ability to share information about their illness with their social network and attachment style dimensions, alexithymia, and quality of life. We hypothesised that ability to share information about one's cancer with family, friends, and medical teams would be positively associated with quality of life and secure attachment and negatively associated with alexithymia. Forty-five cancer patients were recruited from the Psycho-oncology Unit of the San Camillo-Forlanini Hospital in Rome. We collected anamnestic data and self-report data on social sharing ability, quality of life, alexithymia, and attachment. Sharing with family (B = 4.66; SE = 1.82; β = .52; SE = 0.20; t(41) = 2.6; P = .0143) was the only predictor of global health status, and attachment security was the only predictor of mean social sharing (B = 0.25; SE = 0.06; β = .63; SE = 0.14; t(41) = 4.4; P < .0001). Encouraging patients to share information about their experience of cancer may help to improve their quality of life. Attachment security seems to promote social sharing. Psychological assessments of cancer patients should cover both ability to share information about one's cancer with family and attachment security. Copyright © 2016 John Wiley & Sons, Ltd.

FAP-α (Fibroblast activation protein-α) is involved in the control of human breast cancer cell line growth and motility via the FAK pathway

PubMed Central

2014-01-01

Background Fibroblast Activation Protein alpha (FAP-α) or seprase is an integral membrane serine peptidase. Previous work has not satisfactorily explained both the suppression and promotion effects that have been observed in cancer. The purpose of this work was to investigate the role of FAP-α in human breast cancer. Expression of FAP-α was characterized in primary tumour samples and in cell lines, along with the effects of FAP-α expression on in vitro growth, invasion, attachment and migration. Furthermore the potential interaction of FAP-α with other signalling pathways was investigated. Results FAP-α was significantly increased in patients with poor outcome and survival. In vitro results showed that breast cancer cells over expressing FAP-α had increased growth ability and impaired migratory ability. The growth of MDA-MB-231 cells and the adhesion and invasion ability of both MCF-7 cells and MDA-MB-231 cells were not dramatically influenced by FAP-α expression. Over-expression of FAP-α resulted in a reduction of phosphorylated focal adhesion kinase (FAK) level in both cells cultured in normal media and serum-free media. An inhibitor to FAK restored the reduced motility ability of both MCF-7exp cells and MDA-MB-231exp cells and prevented the change in phosphorylated FAK levels. However, inhibitors to PI3K, ERK, PLCϒ, NWASP, ARP2/3, and ROCK had no influence this. Conclusions FAP-α in significantly associated with poor outcome in patients with breast cancer. In vitro, FAP-α promotes proliferation and inhibits migration of breast cancer cells, potentially by regulating the FAK pathway. These results suggest FAP-α could be a target for future therapies. PMID:24885257

FAP-α (Fibroblast activation protein-α) is involved in the control of human breast cancer cell line growth and motility via the FAK pathway.

PubMed

Jia, Jun; Martin, Tracey Amanda; Ye, Lin; Jiang, Wen Guo

2014-05-21

Fibroblast Activation Protein alpha (FAP-α) or seprase is an integral membrane serine peptidase. Previous work has not satisfactorily explained both the suppression and promotion effects that have been observed in cancer. The purpose of this work was to investigate the role of FAP-α in human breast cancer. Expression of FAP-α was characterized in primary tumour samples and in cell lines, along with the effects of FAP-α expression on in vitro growth, invasion, attachment and migration. Furthermore the potential interaction of FAP-α with other signalling pathways was investigated. FAP-α was significantly increased in patients with poor outcome and survival. In vitro results showed that breast cancer cells over expressing FAP-α had increased growth ability and impaired migratory ability. The growth of MDA-MB-231 cells and the adhesion and invasion ability of both MCF-7 cells and MDA-MB-231 cells were not dramatically influenced by FAP-α expression. Over-expression of FAP-α resulted in a reduction of phosphorylated focal adhesion kinase (FAK) level in both cells cultured in normal media and serum-free media. An inhibitor to FAK restored the reduced motility ability of both MCF-7exp cells and MDA-MB-231exp cells and prevented the change in phosphorylated FAK levels. However, inhibitors to PI3K, ERK, PLCΥ, NWASP, ARP2/3, and ROCK had no influence this. FAP-α in significantly associated with poor outcome in patients with breast cancer. In vitro, FAP-α promotes proliferation and inhibits migration of breast cancer cells, potentially by regulating the FAK pathway. These results suggest FAP-α could be a target for future therapies.

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

Martínez Díez, Ana Luisa, E-mail: a.martinez@itma.es; Fraunhofer Institute for Solar Energy Systems ISE, Heidenhofstr. 2, 79110 Freiburg; Gutmann, Johannes

In this paper, we present a concentrator system based on a stack of fluorescent concentrators (FCs) and a bifacial solar cell. Coupling bifacial solar cells to a stack of FCs increases the performance of the system and preserves its efficiency when scaled. We used an approach to optimize a fluorescent solar concentrator system design based on a stack of multiple fluorescent concentrators (FC). Seven individual fluorescent collectors (20 mm×20 mm×2 mm) were realized by in-situ polymerization and optically characterized in regard to their ability to guide light to the edges. Then, an optimization procedure based on the experimental data ofmore » the individual FCs was carried out to determine the stack configuration that maximizes the total number of photons leaving edges. Finally, two fluorescent concentrator systems were realized by attaching bifacial silicon solar cells to the optimized FC stacks: a conventional system, where FC were attached to one side of the solar cell as a reference, and the proposed bifacial configuration. It was found that for the same overall FC area, the bifacial configuration increases the short-circuit current by a factor of 2.2, which is also in agreement with theoretical considerations.« less

Adhesive interactions between milk fat globule membrane and Lactobacillus rhamnosus GG inhibit bacterial attachment to Caco-2 TC7 intestinal cell.

PubMed

Guerin, Justine; Soligot, Claire; Burgain, Jennifer; Huguet, Marion; Francius, Gregory; El-Kirat-Chatel, Sofiane; Gomand, Faustine; Lebeer, Sarah; Le Roux, Yves; Borges, Frederic; Scher, Joël; Gaiani, Claire

2018-07-01

Milk is the most popular matrix for the delivery of lactic acid bacteria, but little is known about how milk impacts bacterial functionality. Here, the adhesion mechanisms of Lactobacillus rhamnosus GG (LGG) surface mutants to a milk component, the milk fat globule membrane (MFGM), were compared using atomic force microscopy (AFM). AFM results revealed the key adhesive role of the LGG SpaCBA pilus in relation to MFGM. A LGG mutant without exopolysaccharides but with highly exposed pili improved the number of adhesive events between LGG and MFGM compared to LGG wild type (WT). In contrast, the number of adhesive events decreased significantly for a LGG mutant without SpaCBA pili. Moreover, the presence of MFGM in the dairy matrix was found to decrease significantly the bacterial attachment ability to Caco-2 TC7 cells. This work thus demonstrated a possible competition between LGG adhesion to MFGM and to epithelial intestinal cells. This competition could negatively impact the adhesion capacity of LGG to intestinal cells in vivo, but requires further substantiation. Copyright © 2018 Elsevier B.V. All rights reserved.

Modulation of cell surface hydrophobicity and attachment of bacteria to abiotic surfaces and shrimp by Malaysian herb extracts.

PubMed

Hui, Yew Woh; Dykes, Gary A

2012-08-01

The use of simple crude water extracts of common herbs to reduce bacterial attachment may be a cost-effective way to control bacterial foodborne pathogens, particularly in developing countries. The ability of water extracts of three common Malaysian herbs (Andrographis paniculata, Eurycoma longifolia, and Garcinia atroviridis) to modulate hydrophobicity and attachment to surfaces of five food-related bacterial strains (Bacillus cereus ATCC 14576, Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 10145, Salmonella Enteritidis ATCC 13076, Staphylococcus aureus ATCC 25923) were determined. The bacterial attachment to hydrocarbon assay was used to determine bacterial hydrophobicity. Staining and direct microscopic counts were used to determine attachment of bacteria to glass and stainless steel. Plating on selective media was used to determine attachment of bacteria to shrimp. All extracts were capable of either significantly ( P < 0.05) increasing or decreasing bacterial surface hydrophobicity, depending on the herb extract and bacteria combination. Bacterial attachment to all surfaces was either significantly (P < 0.05) increased or decreased, depending on the herb extract and bacteria combination. Overall, hydrophobicity did not show a significant correlation (P > 0.05) to bacterial attachment. For specific combinations of bacteria, surface material, and plant extract, significant correlations (R > 0.80) between hydrophobicity and attachment were observed. The highest of these was observed for S. aureus attachment to stainless steel and glass after treatment with the E. longifolia extract (R = 0.99, P < 0.01). The crude water herb extracts in this study were shown to have the potential to modulate specific bacterial and surface interactions and may, with further work, be useful for the simple and practical control of foodborne pathogens.

Microplate Fluorescence Assay for Measurement of the Ability of Strains of Listeria monocytogenes from Meat and Meat-Processing Plants To Adhere to Abiotic Surfaces▿

PubMed Central

Gamble, Rachel; Muriana, Peter M.

2007-01-01

Listeria monocytogenes is a significant food-borne pathogen that is capable of adhering to and producing biofilms on processing equipment, making it difficult to eliminate from meat-processing environments and allowing potential contamination of ready-to-eat (RTE) products. We devised a fluorescence-based microplate method for screening isolates of L. monocytogenes for the ability to adhere to abiotic surfaces. Strains of L. monocytogenes were incubated for 2 days at 30°C in 96-well microplates, and the plates were washed in a plate washer. The retained cells were incubated for 15 min at 25°C with 5,6-carboxyfluorescein diacetate and washed again, and then the fluorescence was read with a plate reader. Several enzymatic treatments (protease, lipase, and cellulase) were effective in releasing adherent cells from the microplates, and this process was used for quantitation on microbiological media. Strongly adherent strains of L. monocytogenes were identified that had 15,000-fold-higher levels of fluorescence and 100,000-fold-higher plate counts in attachment assays than weakly adherent strains. Strongly adherent strains of L. monocytogenes adhered equally well to four different substrates (glass, plastic, rubber, and stainless steel); showed high-level attachment on microplates at 10, 20, 30, and 40°C; and showed significant differences from weakly adherent strains when examined by scanning electron microscopy. A greater incidence of strong adherence was observed for strains isolated from RTE meats than for those isolated from environmental surfaces. Analysis of surface adherence among Listeria isolates from processing environments may provide a better understanding of the molecular mechanisms involved in attachment and suggest solutions to eliminate them from food-processing environments. PMID:17586676

Effect of microbubble ligation to cells on ultrasound signal enhancement: implications for targeted imaging.

PubMed

Lankford, Miles; Behm, Carolyn Z; Yeh, James; Klibanov, Alexander L; Robinson, Peter; Lindner, Jonathan R

2006-10-01

Molecular imaging with contrast-enhanced ultrasound (CEU) relies on the detection of microbubbles retained in regions of disease. The aim of this study was to determine whether microbubble attachment to cells influences their acoustic signal generation and stability. Biotinylated microbubbles were attached to streptavidin-coated plates to derive density versus intensity relations during low- and high-power imaging. To assess damping from microbubble attachment to solid or cell surfaces, in vitro imaging was performed for microbubbles charge-coupled to methacrylate spheres and for vascular cell adhesion molecule-1-targeted microbubbles attached to endothelial cells. Signal enhancement on plates increased according to acoustic power and microbubble site density up to 300 mm. Microbubble signal was reduced by attachment to solid spheres during high- and low-power imaging but was minimally reduced by attachment to endothelial cells and only at low power. Attachment of targeted microbubbles to rigid surfaces results in damping and a reduction of their acoustic signal, which is not seen when microbubbles are attached to cells. A reliable concentration versus intensity relationship can be expected from microbubble attachment to 2-dimensional surfaces until a very high site density is reached.

Influence of Natural Organic Matter on Attachment Kinetics of Salmonella Typhimurium

NASA Astrophysics Data System (ADS)

Chowdhury, I.; Zorlu, O.; Hill, J. E.; Walker, S. L.

2011-12-01

Salmonella enterica serovar Typhimurium is one of the most common and virulent bacterial pathogens, usually found in food and water. This waterborne pathogen has been attributed to causing gastroenteritis and typhoid fever, leading to 16 million cases and over half a million deaths worldwide each year. Natural organic matter (NOM) is ubiquitous in environment and previous work has shown NOM to enhance the stability and transport of bacteria cells; hence NOM will certainly interact with Salmonella and affect its transport in environment. The objective of this study was to investigate the influence of NOM (Suwannee River humic acid standard II, SRHA) on the attachment kinetics of a model Salmonella (Salmonella enterica serovar Typhimurium SA5983) to glass. The transport study was conducted in a parallel plate flow chamber using fluorescent microscope to visualize the bacterial cells, which were tagged with green fluorescent protein (GFP). The solution pH was unadjusted, and the flow rate through parallel plate channel was 0.1 mL/min to simulate groundwater conditions. Parameters varied in this study were NOM presence, ion valence (K+, Ca2+) as well as cell growth phase (mid-exponential and late-exponential growth phases). These parameters were chosen because ion valence may alter the NOM conformation and capacity for bridging, as well growth phase impacts the cellular surface chemistry. Extensive characterization of the bacterial cells was conducted including measurements of electrophoretic mobility, hydrophobicity, acidity, surface charge density and extracellular polymeric substance content. Additionally, electrokintic characterization was conducted for the glass. Preliminary results demonstrated the sensitivity of cell attachment to ionic valence and cell growth phase. Also the addition of NOM reduced the attachment of the Salmonella cells significantly under all of these conditions. Without NOM, attachment efficiencies (α) in KCl were similar at both growth phases; however, in the presence of the divalent ion, α decreased as the cells aged. In presence of NOM and KCl, α was significantly lower at late exponential phase than mid exponential phase; whereas, the opposite was observed with divalent ions. These trends indicate the complex role of NOM, which is coupled with ion valence and growth phase, in the transport of Salmonella. Detailed results will be presented along with proposed mechanisms involved in the interactions between Salmonella and NOM. These mechanisms highlight the role this important naturally occurring macromolecule plays in the fate of Salmonella. This understanding will improve our ability to predict the behavior of this pathogen in environmentally relevant conditions.

Bacteria Interactions with Dying Diatoms

NASA Astrophysics Data System (ADS)

Smriga, S.; Juarez, G.; Fernandez, V.; Stocker, R.

2016-02-01

Dying phytoplankton are surrounded by microscale gradients of dissolved organic matter (DOM) that can attract bacteria. These 'phycospheres' may impact the trophic transfer of DOM in the marine microbial food web and enable the growth of bacterial populations, yet these effects remain poorly quantified particularly in relation to the physiological state of the phytoplankton. We dissected phycosphere interactions at unprecedented spatial and temporal resolution using the model diatom Thalassiosira weissflogii and the bacterium Marinobacter adhaerans. Diatom stress was stimulated by addition of polyunsaturated aldehyde (PUA) and both diatom and bacterial responses were captured via time-lapse fluorescence microscopy. We found that stressed diatoms underwent lysis 10-15 h after PUA treatment. Coordinated with the timing of this transition into phytodetritus, wild-type Marinobacter accumulated via chemotaxis near the diatoms immediately following lysis. In contrast, at lysis there was no accumulation of either a non-chemotactic or a non-motile mutant of Marinobacter, pointing to behavioral rather than demographic responses as drivers for the accumulation. Despite the lack of response, non-chemotactic as well as non-swimming bacterial cells that by chance attached to or were located near (<30 µm) stressed diatoms experienced more growth than cells further afield. Growth within the phycosphere was even greater after diatom lysis. Through quantification at the microscale, these results reveal that chemotaxis may precede rapid bacterial attachment to stressed and dying diatoms and may be integral to the microbial colonization of new phytodetritus during phytoplankton blooms and bloom collapses in coastal ecosystems. Even while chemotactic cells retain a growth advantage given their ability to sense and respond to lysis events, phycosphere DOM provides growth benefits to both motile and non-motile taxa that become attached to or happen to be co-located with new phytodetrital particles, thus likely influencing the composition of particle-attached microbial communities.

Biofilm formation by pathogenic Prototheca algae.

PubMed

Kwiecinski, J

2015-12-01

Prototheca microalgae are the only plants known to cause infections in humans and animals. The mechanisms of Prototheca infections are poorly understood, and no good treatments are available. Biofilms-surface-attached, three-dimensional microbial communities contributing to chronic infections-are formed by many pathogenic bacteria and fungi, but it is not known if Prototheca algae also have this ability. This study shows that various Prototheca species form biofilms composed of surface-attached cells in all growth phases, linked together by matrix containing DNA and polysaccharides. Biofilm formation was modulated by the presence of host plasma or milk. Compared to planktonic cells, Prototheca biofilms caused decreased release of IL-6 by mononuclear immune cells and responded differently to treatment with antimicrobials. Prototheca biofilms possibly contribute to chronic and hard-to-treat character of those algal infections. Prototheca algae are the only existing pathogenic plants. Almost nothing is known about mechanisms of Prototheca infections. This study identifies that, similar to pathogenic bacteria and fungi, Prototheca algae can form biofilms. These biofilms induce reduced immune cell activation relative to planktonic cells, and are also less susceptible to antimicrobials. Biofilm formation by Prototheca could be the first in vitro correlate of pathogenicity, opening a new research field for this pathogen. © 2015 The Society for Applied Microbiology.

Multifunctional Tannic Acid/Silver Nanoparticle-Based Mucoadhesive Hydrogel for Improved Local Treatment of HSV Infection: In Vitro and In Vivo Studies

PubMed Central

Szymańska, Emilia; Orłowski, Piotr; Tomaszewska, Emilia; Bąska, Piotr; Grobelny, Jarosław; Basa, Anna; Krzyżowska, Małgorzata

2018-01-01

Mucoadhesive gelling systems with tannic acid modified silver nanoparticles were developed for effective treatment of herpes virus infections. To increase nanoparticle residence time after local application, semi solid formulations designed from generally regarded as safe (GRAS) excipients were investigated for their rheological and mechanical properties followed with ex vivo mucoadhesive behavior to the porcine vaginal mucosa. Particular effort was made to evaluate the activity of nanoparticle-based hydrogels toward herpes simplex virus (HSV) type 1 and 2 infection in vitro in immortal human keratinocyte cell line and in vivo using murine model of HSV-2 genital infection. The effect of infectivity was determined by real time quantitative polymerase chain reaction, plaque assay, inactivation, attachment, penetration and cell-to-cell assessments. All analyzed nanoparticle-based hydrogels exhibited pseudoplastic and thixotropic properties. Viscosity and mechanical measurements of hydrogels were found to correlate with the mucoadhesive properties. The results confirmed the ability of nanoparticle-based hydrogels to affect viral attachment, impede penetration and cell-to-cell transmission, although profound differences in the activity evoked by tested preparations toward HSV-1 and HSV-2 were noted. In addition, these findings demonstrated the in vivo potential of tannic acid modified silver nanoparticle-based hydrogels for vaginal treatment of HSV-2 genital infection. PMID:29382085

Biophysical basis for convergent evolution of two veil-forming microbes.

PubMed

Petroff, Alexander P; Pasulka, Alexis L; Soplop, Nadine; Wu, Xiao-Lun; Libchaber, Albert

2015-11-01

Microbes living in stagnant water typically rely on chemical diffusion to draw nutrients from their environment. The sulfur-oxidizing bacterium Thiovulum majus and the ciliate Uronemella have independently evolved the ability to form a 'veil', a centimetre-scale mucous sheet on which cells organize to produce a macroscopic flow. This flow pulls nutrients through the community an order of magnitude faster than diffusion. To understand how natural selection led these microbes to evolve this collective behaviour, we connect the physical limitations acting on individual cells to the cell traits. We show how diffusion limitation and viscous dissipation have led individual T. majus and Uronemella cells to display two similar characteristics. Both of these cells exert a force of approximately 40 pN on the water and attach to boundaries by means of a mucous stalk. We show how the diffusion coefficient of oxygen in water and the viscosity of water define the force the cells must exert. We then show how the hydrodynamics of filter-feeding orient a microbe normal to the surface to which it attaches. Finally, we combine these results with new observations of veil formation and a review of veil dynamics to compare the collective dynamics of these microbes. We conclude that this convergent evolution is a reflection of similar physical limitations imposed by diffusion and viscosity acting on individual cells.

Biophysical basis for convergent evolution of two veil-forming microbes

PubMed Central

Petroff, Alexander P.; Pasulka, Alexis L.; Soplop, Nadine; Wu, Xiao-Lun; Libchaber, Albert

2015-01-01

Microbes living in stagnant water typically rely on chemical diffusion to draw nutrients from their environment. The sulfur-oxidizing bacterium Thiovulum majus and the ciliate Uronemella have independently evolved the ability to form a ‘veil’, a centimetre-scale mucous sheet on which cells organize to produce a macroscopic flow. This flow pulls nutrients through the community an order of magnitude faster than diffusion. To understand how natural selection led these microbes to evolve this collective behaviour, we connect the physical limitations acting on individual cells to the cell traits. We show how diffusion limitation and viscous dissipation have led individual T. majus and Uronemella cells to display two similar characteristics. Both of these cells exert a force of approximately 40 pN on the water and attach to boundaries by means of a mucous stalk. We show how the diffusion coefficient of oxygen in water and the viscosity of water define the force the cells must exert. We then show how the hydrodynamics of filter-feeding orient a microbe normal to the surface to which it attaches. Finally, we combine these results with new observations of veil formation and a review of veil dynamics to compare the collective dynamics of these microbes. We conclude that this convergent evolution is a reflection of similar physical limitations imposed by diffusion and viscosity acting on individual cells. PMID:26716000

Listeria monocytogenes biofilm-associated protein (BapL) may contribute to surface attachment of L. monocytogenes but is absent from many field isolates.

PubMed

Jordan, Suzanne J; Perni, Stefano; Glenn, Sarah; Fernandes, Isabel; Barbosa, Manuela; Sol, Manuela; Tenreiro, Rogerio P; Chambel, Lelia; Barata, Belarmino; Zilhao, Isabel; Aldsworth, Timothy G; Adriao, Andreia; Faleiro, M Leonor; Shama, Gilbert; Andrew, Peter W

2008-09-01

Listeria monocytogenes is a food-borne pathogen capable of adhering to a range of surfaces utilized within the food industry, including stainless steel. The factors required for the attachment of this ubiquitous organism to abiotic surfaces are still relatively unknown. In silico analysis of the L. monocytogenes EGD genome identified a putative cell wall-anchored protein (Lmo0435 [BapL]), which had similarity to proteins involved in biofilm formation by staphylococci. An insertion mutation was constructed in L. monocytogenes to determine the influence of this protein on attachment to abiotic surfaces. The results show that the protein may contribute to the surface adherence of strains that possess BapL, but it is not an essential requirement for all L. monocytogenes strains. Several BapL-negative field isolates demonstrated an ability to adhere to abiotic surfaces equivalent to that of BapL-positive strains. BapL is not required for the virulence of L. monocytogenes in mice.

Electrochemical Detection of Human Mesenchymal Stem Cell Differentiation on Fabricated Gold Nano-Dot Cell Chips.

PubMed

An, Jeung Hee; Kim, Seung U; Park, Mi-Kyung; Choi, Jeong Woo

2015-10-01

Human mesenchymal stem cells (MSCs) have the capacity for self-renewal and maintain pluripotency, which is defined by their ability to differentiate into cells such as osteoblasts, neurons, and glial cells. In this study, we report a method for defining the status of human MSCs based on electrochemical detection systems. Gold nano-dot structures were fabricated using a nanoporous alumina mask, and the structural formations were confirmed by scanning electron microscopy (SEM). Human MSCs were allowed to attach to RGD (Arg-Gly-Asp) peptide nanopatterned surfaces, and electrochemical tools were applied to the MSCs attached on the chip surface. The cultured MSCs were shown to differentiate into neural cell types, as indicated by immunocytochemical staining for tyrosine hydroxylase and beta tubulin III. Following treatment with basic fibroblast growth factor (bFGF) for 14 days, most of the B10 cells exhibited bipolar or multipolar morphology with branched processes, and the proportion of B10 cells expressing neuronal cell markers considerably increased. Electrophysiological recordings from MSCs treated with bFGF for 5-14 days were examined with cyclic voltammetry, and the electrochemical signals were shown to increase during differentiation from MSCs to neuronal cells. This human MSC cell line is a useful tool for studying organogenesis, specifically neurogenesis, and in addition, the cell line provides a valuable source of cells for cell therapy. The electrochemical measurement system proposed here could be utilized in electrical cell chips for numerous applications, including cell differentiation, disease diagnosis, drug detection, and on-site monitoring.

Visualisation of morphological interaction of diamond and silver nanoparticles with Salmonella Enteritidis and Listeria monocytogenes.

PubMed

Sawosz, Ewa; Chwalibog, André; Mitura, Katarzyna; Mitura, Stanisław; Szeliga, Jacek; Niemiec, Tomasz; Rupiewicz, Marlena; Grodzik, Marta; Sokołowska, Aleksandra

2011-09-01

Currently, medicine intensively searches for methods to transport drugs to a target (sick) point within the body. The objective of the present investigation was to evaluate morphological characteristics of the assembles of silver or diamond nanoparticles with Salmonella Enteritidis (G-) or Listeria monocytogenes (G+), to reveal possibilities of constructing nanoparticle-bacteria vehicles. Diamond nanoparticles (nano-D) were produced by the detonation method. Hydrocolloids of silver nanoparticles (nano-Ag) were produced by electric non-explosive patented method. Hydrocolloids of nanoparticles (200 microl) were added to bacteria suspension (200 microl) in the following order: nano-D + Salmonella E.; nano-D + Listeria monocytogenes; nano-Ag + Salmonella E; nano-Ag + Listeria monocytogenes. Samples were inspected by transmission electron microscopy. Visualisation of nanoparticles and bacteria interaction showed harmful effects of both nanoparticles on bacteria morphology. The most spectacular effect of nano-D were strong links between nano-D packages and the flagella of Salmonella E. Nano-Ag were closely attached to Listeria monocytogenes but not to Salmonella E. There was no evidence of entering nano-Ag inside Listeria monocytogenes but smaller particles were placed inside Salmonella E. The ability of nano-D to attach to the flagella and the ability of nano-Ag to penetrate inside bacteria cells can be utilized to design nano-bacteria vehicles, being carriers for active substances attached to nanoparticles.

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

PubMed

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.

Effect of bovine manure on fecal coliform attachment to soil and soil particles of different sizes.

PubMed

Guber, Andrey K; Pachepsky, Yakov A; Shelton, Daniel R; Yu, Olivia

2007-05-01

Manure-borne bacteria can be transported in runoff as free cells, cells attached to soil particles, and cells attached to manure particles. The objectives of this work were to compare the attachment of fecal coliforms (FC) to different soils and soil fractions and to assess the effect of bovine manure on FC attachment to soil and soil fractions. Three sand fractions of different sizes, the silt fraction, and the clay fraction of loam and sandy clay loam soils were separated and used along with soil samples in batch attachment experiments with water-FC suspensions and water-manure-FC suspensions. In the absence of manure colloids, bacterial attachment to soil, silt, and clay particles was much higher than the attachment to sand particles having no organic coating. The attachment to the coated sand particles was similar to the attachment to silt and clay. Manure colloids in suspensions decreased bacterial attachment to soils, clay and silt fractions, and coated sand fractions, but did not decrease the attachment to sand fractions without the coating. The low attachment of bacteria to silt and clay particles in the presence of manure colloids may cause predominantly free-cell transport of manure-borne FC in runoff.

The structural proteins of epidemic and historical strains of Zika virus differ in their ability to initiate viral infection in human host cells.

PubMed

Bos, Sandra; Viranaicken, Wildriss; Turpin, Jonathan; El-Kalamouni, Chaker; Roche, Marjolaine; Krejbich-Trotot, Pascale; Desprès, Philippe; Gadea, Gilles

2018-03-01

Mosquito-borne Zika virus (ZIKV) recently emerged in South Pacific islands and Americas where large epidemics were documented. In the present study, we investigated the contribution of the structural proteins C, prM and E in the permissiveness of human host cells to epidemic strains of ZIKV. To this end, we evaluated the capacity of the epidemic strain BeH819015 to infect epithelial A549 and neuronal SH-SY5Y cells in comparison to the African historical MR766 strain. For that purpose, we generated a molecular clone of BeH819015 and a chimeric clone of MR766 which contains the BeH819015 structural protein region. We showed that ZIKV containing BeH819015 structural proteins was much less efficient in cell-attachment leading to a reduced susceptibility of A549 and SH-SY5Y cells to viral infection. Our data illustrate a previously underrated role for C, prM, and E in ZIKV epidemic strain ability to initiate viral infection in human host cells. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Evidence of femtosecond-laser pulse induced cell membrane nanosurgery

NASA Astrophysics Data System (ADS)

Katchinskiy, Nir; Godbout, Roseline; Elezzabi, Abdulhakem Y.

2017-02-01

The mechanism of femtosecond laser nanosurgical attachment is investigated in the following article. Using sub-10 femtosecond laser pulses with 800 nm central wavelength were used to attach retinoblastoma cells. During the attachment process the cell membrane phospholipid bilayers hemifuse into one shared phospholipid bilayer, at the location of attachment. Transmission electron microscopy was used in order to verify the above hypothesis. Based on the imaging results, it was concluded that the two cell membrane coalesce to form one single shared membrane. The technique of cell-cell attachment via femtosecond laser pulses could potentially serve as a platform for precise cell membrane manipulation. Manipulation of the cellular membrane is valuable for studying diseases such as cancer; where the expression level of plasma proteins on the cell membrane is altered.

High Ringxiety: Attachment Anxiety Predicts Experiences of Phantom Cell Phone Ringing.

PubMed

Kruger, Daniel J; Djerf, Jaikob M

2016-01-01

Mobile cell phone users have reported experiencing ringing and/or vibrations associated with incoming calls and messages, only to find that no call or message had actually registered. We believe this phenomenon can be understood as a human signal detection issue, with potentially important influences from psychological attributes. We hypothesized that individuals higher in attachment anxiety would report more frequent phantom cell phone experiences, whereas individuals higher in attachment avoidance would report less frequent experiences. If these experiences are primarily psychologically related to attributes of interpersonal relationships, associations with attachment style should be stronger than for general sensation seeking. We also predicted that certain contexts would interact with attachment style to increase or decrease the likelihood of experiencing phantom cell phone calls and messages. Attachment anxiety directly predicted the frequency of phantom ringing and notification experiences, whereas attachment avoidance and sensation seeking did not directly predict frequency. Attachment anxiety and attachment avoidance interacted with contextual factors (expectations for a call or message and concerned about an issue that one may be contacted about) in the expected directions for predicting phantom cell phone experiences.

Gonococcal attachment to eukaryotic cells

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

James, J.F.; Lammel, C.J.; Draper, D.L.

The attachment of Neisseria gonorrhoeae to eukaryotic cells grown in tissue culture was analyzed by use of light and electron microscopy and by labeling of the bacteria with (/sup 3/H)- and (/sup 14/C)adenine. Isogenic piliated and nonpiliated N. gonorrhoeae from opaque and transparent colonies were studied. The results of light microscopy studies showed that the gonococci attached to cells of human origin, including Flow 2000, HeLa 229, and HEp 2. Studies using radiolabeled gonococci gave comparable results. Piliated N. gonorrhoeae usually attached in larger numbers than nonpiliated organisms, and those from opaque colonies attached more often than isogenic variants frommore » transparent colonies. Day-to-day variation in rate of attachment was observed. Scanning electron microscopy studies showed the gonococcal attachment to be specific for microvilli of the host cells. It is concluded that more N. gonorrhoeae from opaque colonies, as compared with isogenic variants from transparent colonies, attach to eukaryotic cells grown in tissue culture.« less

DNA binding, antioxidant, cytotoxicity (MTT, lactate dehydrogenase, NO), and cellular uptake studies of structurally different nickel(II) thiosemicarbazone complexes: synthesis, spectroscopy, electrochemistry, and X-ray crystallography.

PubMed

Prabhakaran, R; Kalaivani, P; Huang, R; Poornima, P; Vijaya Padma, V; Dallemer, F; Natarajan, K

2013-02-01

Three new nickel(II) thiosemicarbazone complexes have been synthesized and characterized by analytical, spectral, and single-crystal X-ray diffraction studies. In complex 1, the ligand 2-hydroxy-1-naphthaldehydethiosemicarbazone coordinated as a monobasic tridentate donor, whereas in complexes 2 and 3, the ligands salicylaldehyde-4(N)-ethylthiosemicarbazone and 2-hydroxy-1-naphthaldehyde-4(N)-ethylthiosemicarbazone coordinated as a dibasic tridentate donor. The DNA binding ability of the complexes in calf thymus DNA was explored by absorption and emission titration experiments. The antioxidant property of the new complexes was evaluated to test their free-radical scavenging ability. In vitro cytotoxicity assays were performed for the new complexes in A549 and HepG2 cell lines. The new compounds overcome cisplatin resistance in the A549 cell line and they were also active in the HepG2 cell line. The cellular uptake study showed the accumulation of the complexes in tumor cells depended on the nature of the ligand attached to the nickel ion.

Biofilm formation and interspecies interactions in mixed cultures of thermo-acidophilic archaea Acidianus spp. and Sulfolobus metallicus.

PubMed

Castro, Camila; Zhang, Ruiyong; Liu, Jing; Bellenberg, Sören; Neu, Thomas R; Donati, Edgardo; Sand, Wolfgang; Vera, Mario

2016-09-01

The understanding of biofilm formation by bioleaching microorganisms is of great importance for influencing mineral dissolution rates and to prevent acid mine drainage (AMD). Thermo-acidophilic archaea such as Acidianus, Sulfolobus and Metallosphaera are of special interest due to their ability to perform leaching at high temperatures, thereby enhancing leaching rates. In this work, leaching experiments and visualization by microscopy of cell attachment and biofilm formation patterns of the crenarchaeotes Sulfolobus metallicus DSM 6482(T) and the Acidianus isolates DSM 29038 and DSM 29099 in pure and mixed cultures on sulfur or pyrite were studied. Confocal laser scanning microscopy (CLSM) combined with fluorescent dyes as well as fluorescently labeled lectins were used to visualize different components (e.g. DNA, proteins or glycoconjugates) of the aforementioned species. The data indicate that cell attachment and the subsequently formed biofilms were species- and substrate-dependent. Pyrite leaching experiments coupled with pre-colonization and further inoculation with a second species suggest that both species may negatively influence each other during pyrite leaching with respect to initial attachment and pyrite dissolution rates. In addition, the investigation of binary biofilms on pyrite showed that both species were heterogeneously distributed on pyrite surfaces in the form of individual cells or microcolonies. Physical contact between the two species seems to occur, as revealed by specific lectins able to specifically bind single species within mixed cultures. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Substrate effects on endothelial cell adherence rates.

PubMed

Scott, W J; Mann, P

1990-01-01

Endothelial cell attachment to a synthetic substrate was studied using an in vitro model system. Attachment rate was defined as the number of tritium-labeled endothelial cells attached to a synthetic substrate after 30 minutes. The surface of the synthetic substrate was chemically modified with either laminin or fibronectin. Labeled endothelial cells attached more rapidly to synthetic substrate, chemically modified with biomolecules, as compared with the untreated substrate controls. Unlabeled endothelial cells were grown to confluency on a second set of modified and untreated substrates. The cells were removed with 1% Triton, and the rate of re-endothelialization with tritium-labeled endothelial cells was determined. The rate was 11-13 times that of the same cells on untreated substrate. These data confirm that biomolecules increase the attachment rate of endothelial cells to synthetic substrate, and also suggest that endothelial cells may secrete a Triton-insoluble product (Sigma, St. Louis, MO) into subendothelial matrix that increases re-endothelialization.

Study of the Ability of Bifidobacteria of Human Origin to Prevent and Treat Rotavirus Infection Using Colonic Cell and Mouse Models

PubMed Central

Darveau, André; Fliss, Ismaïl

2016-01-01

Rotavirus is the leading cause of severe acute gastroenteritis among children worldwide. Despite effective vaccines, inexpensive alternatives such as probiotics are needed. The aim of this study was to assess the ability of probiotic candidate Bifidobacterium thermophilum RBL67 to inhibit rotavirus infection. Bacterial adhesion to intestinal cells and interference with viral attachment were evaluated in vitro. B. thermophilum RBL67 displayed adhesion indexes of 625 ± 84 and 1958 ± 318 on Caco-2 and HT-29 cells respectively and was comparable or superior to four other bifidobacteria, including B. longum ATCC 15707 and B. pseudolongum ATCC 25526 strains. Incubation of B. thermophilum RBL67 for 30 min before (exclusion) and simultaneously (competition) with human rotavirus strain Wa decreased virus attachment by 2.0 ± 0.1 and 1.5 ± 0.1 log10 (by 99.0% and 96.8% respectively). Displacement of virus already present was negligible. In CD-1 suckling mice fed B. thermophilum RBL67 challenged with simian rotavirus SA-11, pre-infection feeding with RBL 67 was more effective than post-infection feeding, reducing the duration of diarrhea, limiting epithelial lesions, reducing viral replication in the intestine, accelerating recovery, and stimulating the humoral specific IgG and IgM response, without inducing any adverse effect. B. thermophilum RBL67 had little effect on intestinal IgA titer. These results suggest that humoral immunoglobulin might provide protection against the virus and that B. thermophilum RBL67 has potential as a probiotic able to inhibit rotavirus infection and ultimately reduce its spread. PMID:27727323

Enteropathogenic Escherichia coli O125:H6 Triggers Attaching and Effacing Lesions on Human Intestinal Biopsy Specimens Independently of Nck and TccP/TccP2▿

PubMed Central

Bai, Li; Schüller, Stephanie; Whale, Andrew; Mousnier, Aurelie; Marches, Olivier; Wang, Lei; Ooka, Tadasuke; Heuschkel, Robert; Torrente, Franco; Kaper, James B.; Gomes, Tânia A. T.; Xu, Jianguo; Phillips, Alan D.; Frankel, Gad

2008-01-01

Typical enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) employ either Nck, TccP/TccP2, or Nck and TccP/TccP2 pathways to activate the neuronal Wiskott-Aldrich syndrome protein (N-WASP) and to trigger actin polymerization in cultured cells. This phenotype is used as a marker for the pathogenic potential of EPEC and EHEC strains. In this paper we report that EPEC O125:H6, which represents a large category of strains, lacks the ability to utilize either Nck or TccP/TccP2 and hence triggers actin polymerization in vitro only inefficiently. However, we show that infection of human intestinal biopsies with EPEC O125:H6 results in formation of typical attaching and effacing lesions. Expression of TccP in EPEC O125:H6, which harbors an EHEC O157-like Tir, resulted in efficient actin polymerization in vitro and enhanced colonization of human intestinal in vitro organ cultures with detectable N-WASP and electron-dense material at the site of bacterial adhesion. These results show the existence of a natural category of EPEC that colonizes the gut mucosa using Nck- and TccP-independent mechanisms. Importantly, the results highlight yet again the fact that conclusions made on the basis of in vitro cell culture models cannot be extrapolated wholesale to infection of mucosal surfaces and that the ability to induce actin polymerization on cultured cells should not be used as a definitive marker for EPEC and EHEC virulence. PMID:17984209

Enteropathogenic Escherichia coli O125:H6 triggers attaching and effacing lesions on human intestinal biopsy specimens independently of Nck and TccP/TccP2.

PubMed

Bai, Li; Schüller, Stephanie; Whale, Andrew; Mousnier, Aurelie; Marches, Olivier; Wang, Lei; Ooka, Tadasuke; Heuschkel, Robert; Torrente, Franco; Kaper, James B; Gomes, Tânia A T; Xu, Jianguo; Phillips, Alan D; Frankel, Gad

2008-01-01

Typical enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) employ either Nck, TccP/TccP2, or Nck and TccP/TccP2 pathways to activate the neuronal Wiskott-Aldrich syndrome protein (N-WASP) and to trigger actin polymerization in cultured cells. This phenotype is used as a marker for the pathogenic potential of EPEC and EHEC strains. In this paper we report that EPEC O125:H6, which represents a large category of strains, lacks the ability to utilize either Nck or TccP/TccP2 and hence triggers actin polymerization in vitro only inefficiently. However, we show that infection of human intestinal biopsies with EPEC O125:H6 results in formation of typical attaching and effacing lesions. Expression of TccP in EPEC O125:H6, which harbors an EHEC O157-like Tir, resulted in efficient actin polymerization in vitro and enhanced colonization of human intestinal in vitro organ cultures with detectable N-WASP and electron-dense material at the site of bacterial adhesion. These results show the existence of a natural category of EPEC that colonizes the gut mucosa using Nck- and TccP-independent mechanisms. Importantly, the results highlight yet again the fact that conclusions made on the basis of in vitro cell culture models cannot be extrapolated wholesale to infection of mucosal surfaces and that the ability to induce actin polymerization on cultured cells should not be used as a definitive marker for EPEC and EHEC virulence.

On the use of magnets to disrupt the physiological compass of birds.

PubMed

Wang, K; Mattern, E; Ritz, T

2006-10-04

Behavioral researchers have attached magnets to birds during orientation experiments, assuming that such magnets will disrupt their ability to obtain magnetic information. Here, we investigate the effect of an attached magnet on the ability to derive directional information from a radical-pair based compass mechanism. We outline in some detail the geometrical symmetries that would allow a bird to identify magnetic directions in a radical-pair based compass. We show that the artificial field through an attached magnet will quickly disrupt the birds' ability to distinguish pole-ward from equator-ward headings, but that much stronger fields are necessary to disrupt their ability to detect the magnetic axis. Together with estimates of the functional limits of a radical-pair based compass, our calculations suggest that artificial fields of comparable size to the geomagnetic field are not generally sufficient to render a radical-pair based compass non-functional.

Assembly of Hepatitis Delta Virus: Particle Characterization, Including the Ability To Infect Primary Human Hepatocytes▿

PubMed Central

Gudima, Severin; He, Yiping; Meier, Anja; Chang, Jinhong; Chen, Rongji; Jarnik, Michal; Nicolas, Emmanuelle; Bruss, Volker; Taylor, John

2007-01-01

Efficient assembly of hepatitis delta virus (HDV) was achieved by cotransfection of Huh7 cells with two plasmids: one to provide expression of the large, middle, and small envelope proteins of hepatitis B virus (HBV), the natural helper of HDV, and another to initiate replication of the HDV RNA genome. HDV released into the media was assayed for HDV RNA and HBV envelope proteins and characterized by rate-zonal sedimentation, immunoaffinity purification, electron microscopy, and the ability to infect primary human hepatocytes. Among the novel findings were that (i) immunostaining for delta antigen 6 days after infection with 300 genome equivalents (GE) per cell showed only 1% of cells as infected, but this was increased to 16% when 5% polyethylene glycol was present during infection; (ii) uninfected cells did not differ from infected cells in terms of albumin accumulation or the presence of E-cadherin at cell junctions; and (iii) sensitive quantitative real-time PCR assays detected HDV replication even when the multiplicity of infection was 0.2 GE/cell. In the future, this HDV assembly and infection system can be further developed to better understand the mechanisms shared by HBV and HDV for attachment and entry into host cells. PMID:17229685

Physico-Mechanical Properties of HA/TCP Pellets and Their Three-Dimensional Biological Evaluation In Vitro.

PubMed

Kamalaldin, Nurulain 'Atikah; Jaafar, Mariatti; Zubairi, Saiful Irwan; Yahaya, Badrul Hisham

2018-01-04

The use of bioceramics, especially the combination of hydroxyapatite (HA) and β-tricalcium phosphate (β-TCP), as a three-dimensional scaffold in bone engineering is essential because together these elements constitute 60% of the bone content. Different ratios of HA and β-TCP were previously tested for their ability to produce suitable bioceramic scaffolds, which must be able to withstand high mechanical load. In this study, two ratios of HA/TCP (20:80 and 70:30) were used to create pellets, which then were evaluated in vitro to identify any adverse effects of using the material in bone grafting. Diametral tensile strength (DTS) and density testing was conducted to assess the mechanical strength and porosity of the pellets. The pellets then were tested for their toxicity to normal human fibroblast cells. In the toxicity assay, cells were incubated with the pellets for 3 days. At the end of the experiment, cell morphological changes were assessed, and the absorbance was read using PrestoBlue Cell Viability Reagent™. An inversely proportional relationship between DTS and porosity percentage was detected. Fibroblasts showed normal cell morphology in both treatments, which suggests that the HA/TCP pellets were not toxic. In the osteoblast cell attachment assay, cells were able to attach to the surface of both ratios, but cells were also able to penetrate inside the scaffold of the 70:30 pellets. This finding suggests that the 70:30 ratio had better osteoconduction properties than the 20:80 ratio.

Effects of nerve cells and adhesion molecules on nerve conduit for peripheral nerve regeneration

PubMed Central

Fiorellini, Joseph P.

2017-01-01

Background For peripheral nerve regeneration, recent attentions have been paid to the nerve conduits made by tissue-engineering technique. Three major elements of tissue-engineering are cells, molecules, and scaffolds. Methods In this study, the attachments of nerve cells, including Schwann cells, on the nerve conduit and the effects of both growth factor and adhesion molecule on these attachments were investigated. Results The attachment of rapidly-proliferating cells, C6 cells and HS683 cells, on nerve conduit was better than that of slowly-proliferating cells, PC12 cells and Schwann cells, however, the treatment of nerve growth factor improved the attachment of slowly-proliferating cells. In addition, the attachment of Schwann cells on nerve conduit coated with fibronectin was as good as that of Schwann cells treated with glial cell line-derived neurotrophic factor (GDNF). Conclusions Growth factor changes nerve cell morphology and affects cell cycle time. And nerve growth factor or fibronectin treatment is indispensable for Schwann cell to be used for implantation in artificial nerve conduits. PMID:29090249

Synthesis and biological evaluation of novel heterocyclic derivatives of combretastatin A-4.

PubMed

Nguyen, Thi Thanh Binh; Lomberget, Thierry; Tran, Ngoc Chau; Colomb, Evelyne; Nachtergaele, Lore; Thoret, Sylviane; Dubois, Joëlle; Guillaume, Joren; Abdayem, Rawad; Haftek, Marek; Barret, Roland

2012-12-01

A novel series of combretastatin A-4 heterocyclic analogues was prepared by replacement of the B ring with indole, benzofurane or benzothiophene, attached at the C2 position. These compounds were evaluated for their abilities to inhibit tubulin assembly: derivative cis3b, having a benzothiophene, showed an activity similar to those of colchicine or deoxypodophyllotoxine. The antiproliferative and antimitotic properties of cis3b against keratinocyte cancer cell lines were also evaluated and the intracellular organization of microtubules in the cells after treatment with both stereoisomers of 3b was also determined, using confocal microscopy. Copyright © 2012 Elsevier Ltd. All rights reserved.

Synergistic Effect of Carbon Nanotubes and Graphene on Diopside Scaffolds.

PubMed

Liu, Tingting; Wu, Ping; Gao, Chengde; Feng, Pei; Xiao, Tao; Deng, Youwen; Shuai, Cijun; Peng, Shuping

2016-01-01

A synergetic effect between carbon nanotubes (CNTs) and graphene on diopside (Di) scaffolds was demonstrated. 3D network architecture in the matrix was formed through the 1D CNTs inlaid among the 2D graphene platelets (GNPs). The mechanical properties of the CNTs/GNPs/Di scaffolds were significantly improved compared with the CNTs/Di scaffolds and GNPs/Di scaffolds. In addition, the scaffolds exhibited excellent apatite-forming ability, a modest degradation rate, and stable mechanical properties in simulated body fluid (SBF). Moreover, cell culturing tests indicated that the scaffolds supported the cells attachment and proliferation. Taken together, the CNTs/GNPs/Di scaffolds offered great potential for bone tissue engineering.

Aggregation of the rhizospheric bacterium Azospirillum brasilense in response to oxygen

NASA Astrophysics Data System (ADS)

Abdoun, Hamid; McMillan, Mary; Pereg, Lily

2016-04-01

Azospirillum brasilense spp. have ecological, scientific and agricultural importance. As model plant growth promoting rhizobacteria they interact with a large variety of plants, including important food and cash crops. Azospirillum strains are known for their production of plant growth hormones that enhance root systems and for their ability to fix nitrogen. Azospirillum cells transform in response to environmental cues. The production of exopolysaccharides and cell aggregation during cellular transformation are important steps in the attachment of Azospirillum to roots. We investigate signals that induce cellular transformation and aggregation in the Azospirillum and report on the importance of oxygen to the process of aggregation in this rhizospheric bacterium.

Experiential avoidance mediates the link between maternal attachment style and theory of mind.

PubMed

Vanwoerden, Salome; Kalpakci, Allison H; Sharp, Carla

2015-02-01

Theoretical and empirical models suggest a relation between attachment style and theory of mind (ToM) in childhood and adulthood; however, this link has not been evaluated to the same extent in adolescence. Additionally, these models typically fail to consider mechanisms by which attachment style affects ToM abilities. The present study sought to test a mediational model in which experiential avoidance mediates the relation between maternal attachment style and ToM. A sample of 282 adolescents (Mage=15.42years, SD=1.44, 62.8% female) was recruited from an inpatient psychiatric hospital. Findings revealed that maternal attachment style in females was related to ToM, through experiential avoidance. Specifically, those with a disorganized maternal attachment were most likely to engage in experiential avoidant cognitive and emotional strategies, which in turn related to lower levels of ToM ability. Implications and areas for future research are discussed. Copyright © 2014 Elsevier Inc. All rights reserved.

Physiologically Low Oxygen Enhances Biomolecule Production and Stemness of Mesenchymal Stem Cell Spheroids

PubMed Central

Shearier, Emily; Xing, Qi; Qian, Zichen

2016-01-01

Multicellular human mesenchymal stem cell (hMSC) spheroids have been demonstrated to be valuable in a variety of applications, including cartilage regeneration, wound healing, and neoangiogenesis. Physiological relevant low oxygen culture can significantly improve in vitro hMSC expansion by preventing cell differentiation. We hypothesize that hypoxia-cultured hMSC spheroids can better maintain the regenerative properties of hMSCs. In this study, hMSC spheroids were fabricated using hanging drop method and cultured under 2% O2 and 20% O2 for up to 96 h. Spheroid diameter and viability were examined, as well as extracellular matrix (ECM) components and growth factor levels between the two oxygen tensions at different time points. Stemness was measured among the spheroid culture conditions and compared to two-dimensional cell cultures. Spheroid viability and structural integrity were studied using different needle gauges to ensure no damage would occur when implemented in vivo. Spheroid attachment and integration within a tissue substitute were also demonstrated. The results showed that a three-dimensional hMSC spheroid cultured at low oxygen conditions can enhance the production of ECM proteins and growth factors, while maintaining the spheroids' stemness and ability to be injected, attached, and potentially be integrated within a tissue. PMID:26830500

Community dynamics of attached and free cells and the effects of attached cells on chalcopyrite bioleaching by Acidithiobacillus sp.

PubMed

Yang, Hailin; Feng, Shoushuai; Xin, Yu; Wang, Wu

2014-02-01

The community dynamics of attached and free cells of Acidithiobacillus sp. were investigated and compared during chalcopyrite bioleaching process. In the mixed strains system, Acidithiobacillus ferrooxidans was the dominant species at the early stage while Acidithiobacillus thiooxidans owned competitive advantage from the middle stage to the end of bioprocess. Meanwhile, compared to A. ferrooxidans, more significant effects of attached cells on free biomass with A. thiooxidans were shown in either the pure or mixed strains systems. Moreover, the effects of attached cells on key chemical parameters were also studied in different adsorption-deficient systems. Consistently, the greatest reduction of key chemical ion was shown with A. thiooxidans and the loss of bioleaching efficiency was high to 50.5%. These results all demonstrated the bioleaching function of attached cells was more efficient than the free cells, especially with A. thiooxidans. These notable results would help us to further understand the chalcopyrite bioleaching. Copyright © 2013 Elsevier Ltd. All rights reserved.

Pseudomonas aeruginosa cells attached to a surface display a typical proteome early as 20 minutes of incubation

PubMed Central

Crouzet, Marc; Claverol, Stéphane; Lomenech, Anne-Marie; Le Sénéchal, Caroline; Costaglioli, Patricia; Barthe, Christophe; Garbay, Bertrand; Bonneu, Marc

2017-01-01

Biofilms are present in all environments and often result in negative effects due to properties of the biofilm lifestyle and especially antibiotics resistance. Biofilms are associated with chronic infections. Controlling bacterial attachment, the first step of biofilm formation, is crucial for fighting against biofilm and subsequently preventing the persistence of infection. Thus deciphering the underlying molecular mechanisms involved in attachment could allow discovering molecular targets from it would be possible to develop inhibitors against bacterial colonization and potentiate antibiotherapy. To identify the key components and pathways that aid the opportunistic pathogen Pseudomonas aeruginosa in attachment we performed for the first time a proteomic analysis as early as after 20 minutes of incubation using glass wool fibers as a surface. We compared the protein contents of the attached and unattached bacteria. Using mass spectrometry, 3043 proteins were identified. Our results showed that, as of 20 minutes of incubation, using stringent quantification criteria 616 proteins presented a modification of their abundance in the attached cells compared to their unattached counterparts. The attached cells presented an overall reduced gene expression and characteristics of slow-growing cells. The over-accumulation of outer membrane proteins, periplasmic folding proteins and O-antigen chain length regulators was also observed, indicating a profound modification of the cell envelope. Consistently the sigma factor AlgU required for cell envelope homeostasis was highly over-accumulated in attached cells. In addition our data suggested a role of alarmone (p)ppGpp and polyphosphate during the early attachment phase. Furthermore, almost 150 proteins of unknown function were differentially accumulated in the attached cells. Our proteomic analysis revealed the existence of distinctive biological features in attached cells as early as 20 minutes of incubation. Analysis of some mutants demonstrated the interest of this proteomic approach in identifying genes involved in the early phase of adhesion to a surface. PMID:28678862

Suppression of adhesion-induced protein tyrosine phosphorylation decreases invasive and metastatic potentials of B16-BL6 melanoma cells by protein tyrosine kinase inhibitor genistein.

PubMed

Yan, C; Han, R

1997-01-01

Protein tyrosine kinase (PTK) appears to be involved in the activation of signaling during cell attachment to and spreading on extracellular matrix (ECM) in the metastatic cascade. To verify the assumption that PTK inhibitors might impair ECM signaling and prevent cancer metastasis, the highly metastatic B16-BL6 mouse melanoma cells were exposed to the PTK inhibitor genistein for 3 days. The ability of the cells to invade through reconstituted basement membrane (Matrigel) and to establish experimental pulmonary metastatic foci in C57BL/6 mice decreased after genistein exposure. The genistein-treated cells were also prevented from attaching to Matrigel and spread extremely poorly on the ECM substratum. Immunoblot analysis showed that tyrosine phosphorylation of a 125-kD protein in response to cell spreading on Matrigel was suppressed in the genistein-treated cells. Adhesion-induced protein tyrosine phosphorylation represents the earlier and specific event in the activation of ECM signaling, so this result implied ECM signaling was impaired in the treated cells. With immunofluorescence microscopy, the adhesion-induced tyrosine phosphorylated proteins were located at the pericytoplasms of well-spread cells, but not at the periphery of poorly spread genistein-treated cells. Therefore, this paper suggests that genistein might impair ECM signaling and subsequently prevent cancer cells from spreading well and invading or establishing metastasis through the suppression of adhesion-induced protein tyrosine phosphorylation. PTKs and adhesion-induced protein tyrosine phosphorylation might play a role in the control of invasion and metastasis.

Characterization of femtosecond-laser pulse induced cell membrane nanosurgical attachment.

PubMed

Katchinskiy, Nir; Godbout, Roseline; Elezzabi, Abdulhakem Y

2016-07-01

This article provides insight into the mechanism of femtosecond laser nanosurgical attachment of cells. We have demonstrated that during the attachment of two retinoblastoma cells using sub-10 femtosecond laser pulses, with 800 nm central wavelength, the phospholipid molecules of both cells hemifuse and form one shared phospholipid bilayer, at the attachment location. In order to verify the hypothesis that hemifusion takes place, transmission electron microscope images of the cell membranes of retinoblastoma cells were taken. It is shown that at the attachment interface, the two cell membranes coalesce and form one single membrane shared by both cells. Thus, further evidence is provided to support the hypothesis that laser-induced ionization process led to an ultrafast reversible destabilization of the phospholipid layer of the cellular membrane, which resulted in cross-linking of the phospholipid molecules in each membrane. This process of hemifusion occurs throughout the entire penetration depth of the femtosecond laser pulse train. Thus, the attachment between the cells takes place across a large surface area, which affirms our findings of strong physical attachment between the cells. The femtosecond laser pulse hemifusion technique can potentially provide a platform for precise molecular manipulation of cellular membranes. Manipulation of the cellular membrane is an important procedure that could aid in studying diseases such as cancer; where the expression level of plasma proteins on the cell membrane is altered.

Silencing by nuclear matrix attachment distinguishes cell-type specificity: association with increased proliferation capacity.

PubMed

Linnemann, Amelia K; Krawetz, Stephen A

2009-05-01

DNA loop organization by nuclear scaffold/matrix attachment is a key regulator of gene expression that may provide a means to modulate phenotype. We have previously shown that attachment of genes to the NaCl-isolated nuclear matrix correlates with their silencing in HeLa cells. In contrast, expressed genes were associated with the lithium 3,5-diiodosalicylate (LIS)-isolated nuclear scaffold. To define their role in determining phenotype matrix attached regions (MARs) on human chromosomes 14-18 were identified as a function of expression in a primary cell line. The locations of MARs in aortic adventitial fibroblast (AoAF) cells were very stable (r = 0.909) and 96% of genes attached at MARs are silent (P < 0.001). Approximately one-third of the genes uniquely expressed in AoAF cells were associated with the HeLa cell nuclear matrix and silenced. Comparatively, 81% were associated with the AoAF cell nuclear scaffold (P < 0.001) and expressed. This suggests that nuclear scaffold/matrix association mediates a portion of cell type-specific gene expression thereby modulating phenotype. Interestingly, nuclear matrix attachment and thus silencing of specific genes that regulate proliferation and maintain the integrity of the HeLa cell genome suggests that transformation may at least in part be achieved through aberrant nuclear matrix attachment.

Chlorine stress mediates microbial surface attachment in drinking water systems.

PubMed

Liu, Li; Le, Yang; Jin, Juliang; Zhou, Yuliang; Chen, Guowei

2015-03-01

Microbial attachment to drinking water pipe surfaces facilitates pathogen survival and deteriorates disinfection performance, directly threatening the safety of drinking water. Notwithstanding that the formation of biofilm has been studied for decades, the underlying mechanisms for the origins of microbial surface attachment in biofilm development in drinking water pipelines remain largely elusive. We combined experimental and mathematical methods to investigate the role of environmental stress-mediated cell motility on microbial surface attachment in chlorination-stressed drinking water distribution systems. Results show that at low levels of disinfectant (0.0-1.0 mg/L), the presence of chlorine promotes initiation of microbial surface attachment, while higher amounts of disinfectant (>1.0 mg/L) inhibit microbial attachment. The proposed mathematical model further demonstrates that chlorination stress (0.0-5.0 mg/L)-mediated microbial cell motility regulates the frequency of cell-wall collision and thereby controls initial microbial surface attachment. The results reveal that transport processes and decay patterns of chlorine in drinking water pipelines regulate microbial cell motility and, thus, control initial surface cell attachment. It provides a mechanistic understanding of microbial attachment shaped by environmental disinfection stress and leads to new insights into microbial safety protocols in water distribution systems.

Transforming growth factor-beta1 promotes the migration and invasion of sphere-forming stem-like cell subpopulations in esophageal cancer

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

Yue, Dongli; Zhang, Zhen; Li, Jieyao

Esophageal cancer is one of the most lethal solid malignancies. Mounting evidence demonstrates that cancer stem cells (CSCs) are able to cause tumor initiation, metastasis and responsible for chemotherapy and radiotherapy failures. As CSCs are thought to be the main reason of therapeutic failure, these cells must be effectively targeted to elicit long-lasting therapeutic responses. We aimed to enrich and identify the esophageal cancer cell subpopulation with stem-like properties and help to develop new target therapy strategies for CSCs. Here, we found esophageal cancer cells KYSE70 and TE1 could form spheres in ultra low attachment surface culture and be seriallymore » passaged. Sphere-forming cells could redifferentiate and acquire morphology comparable to parental cells, when return to adherent culture. The sphere-forming cells possessed the key criteria that define CSCs: persistent self-renewal, overexpression of stemness genes (SOX2, ALDH1A1 and KLF4), reduced expression of differentiation marker CK4, chemoresistance, strong invasion and enhanced tumorigenic potential. SB525334, transforming growth factor-beta 1(TGF-β1) inhibitor, significantly inhibited migration and invasion of sphere-forming stem-like cells and had no effect on sphere-forming ability. In conclusion, esophageal cancer sphere-forming cells from KYSE70 and TE1 cultured in ultra low attachment surface possess cancer stem cell properties, providing a model for CSCs targeted therapy. TGF-β1 promotes the migration and invasion of sphere-forming stem-like cells, which may guide future studies on therapeutic strategies targeting these cells. - Highlights: • Esophageal cancer sphere-forming cells possess cancer stem cell properties. • Sphere-forming cells enhance TGF-β1 pathway activity. • TGF-β 1 inhibitor suppresses the migration and invasion of sphere-forming cells.« less

Insights to the effects of free cells on community structure of attached cells and chalcopyrite bioleaching during different stages.

PubMed

Feng, Shoushuai; Yang, Hailin; Wang, Wu

2016-01-01

The effects of free cells on community structure of attached cells and chalcopyrite bioleaching by Acidithiobacillus sp. during different stages were investigated. The attached cells of Acidithiobacillus thiooxidans owned the community advantage from 14thd to the end of bioprocess in the normal system. The community structure of attached cells was greatly influenced in the free cells-deficient systems. Compared to A. thiooxidans, the attached cells community of Acidithiobacillus ferrooxidans had a higher dependence on its free cells. Meanwhile, the analysis of key biochemical parameters revealed that the effects of free cells on chalcopyrite bioleaching in different stages were diverse, ranging from 32.8% to 64.3%. The bioleaching contribution of free cells of A. ferrooxidans in the stationary stage (8-14thd) was higher than those of A. thiooxidans, while the situation was gradually reversed in the jarosite passivation inhibited stage (26-40thd). These results may be useful in guiding chalcopyrite bioleaching. Copyright © 2015 Elsevier Ltd. All rights reserved.

Influence of substrate diffusion on degradation of dibenzofuran and 3-chlorodibenzofuran by attached and suspended bacteria.

PubMed Central

Harms, H; Zehnder, A J

1994-01-01

Dibenzofuran uptake-associated kinetic parameters of suspended and attached Sphingomonas sp. strain HH19k cells were compared. The suspended cells were studied in a batch system, whereas glass beads in percolated columns were used as the solid support for attached cells. The maximum specific activities of cells in the two systems were the same. The apparent half-maximum uptake rate-associated concentrations (Kt') of attached cells, however, were considerably greater than those of suspended cells and depended on cell density and on percolation velocity. A mathematical model was developed to explain the observed differences in terms of substrate transport to the cells. This model was based on the assumptions that the intrinsic half-maximum uptake rate-associated concentration (Kt) was unchanged and that deviations of Kt' from Kt resulted from the stereometry and the hydrodynamics around the cells. Our calculations showed that (i) diffusion to suspended cells and to single attached cells is efficient and therefore only slightly affects Kt'; (ii) diffusion to cells located on crowded surfaces is considerably lower than that to single attached cells and greatly increases Kt', which depends on the cell density; (iii) the convective-diffusive transport to attached cells that occurs in a percolated column is influenced by the liquid flow and results in dependency of Kt' on the flow rate; and (iv) higher specific affinity of cells correlates with higher susceptibility to diffusion limitation. Properties of the experimental system which limited quantitative proof of exclusively transport-controlled variations of Kt' are discussed. PMID:8085817

The RhoA-ROCK-PTEN pathway as a molecular switch for anchorage dependent cell behavior.

PubMed

Yang, Seungwon; Kim, Hyun-Man

2012-04-01

The proliferation of anchorage-dependent cells of mesenchymal origin requires the attachment of the cells to substrates. Thus, cells that are poorly attached to substrates exhibit retarded cell cycle progression or apoptotic death. A major disadvantage of most polymers used in tissue engineering is their hydrophobicity; hydrophobic surfaces do not allow cells to attach firmly and, therefore, do not allow normal proliferation rates. In this study, we investigated the molecular mechanism underlying the reduced proliferation rate of cells that are poorly attached to substrates. There was an inverse relationship between the activity of the small GTPase RhoA (RhoA) and the cell proliferation rate. RhoA activity correlated inversely with the strength of cell adhesion to the substrates. The high RhoA activity in the cells poorly attached to substrates caused an increase in the activity of Rho-associated kinase (ROCK), a well-known effector of RhoA that upregulated the activity of phosphatase and tensin homolog (PTEN). The resulting activated PTEN downregulated Akt activity, which is essential for cell proliferation. Thus, the cells that were poorly attached to substrates showed low levels of cell proliferation because the RhoA-ROCK-PTEN pathway was hyperactive. In addition, RhoA activity seemed to be related to focal adhesion kinase (FAK) activity. Weak FAK activity in these poorly attached cells failed to downregulate the high RhoA activity that restrained cell proliferation. Interestingly, reducing the expression of any component of the RhoA-ROCK-PTEN pathway rescued the proliferation rate without physico-chemical surface modifications. Based on these results, we suggest that the RhoA-ROCK-PTEN pathway acts as a molecular switch to control cell proliferation and determine anchorage dependence. In cells that are poorly attached to substrates, its inhibition is sufficient to restore cell proliferation without the need for physico-chemical modification of the material surface. Copyright © 2012 Elsevier Ltd. All rights reserved.

Overexpression of dominant negative PARP interferes with tumor formation of HeLa cells in nude mice: evidence for increased tumor cell apoptosis in vivo.

PubMed

Hans, M A; Müller, M; Meyer-Ficca, M; Bürkle, A; Küpper, J H

1999-11-25

Poly(ADP-ribose) polymerase (PARP4) catalyzes the formation of ADP-ribose polymers covalently attached to proteins by using NAD+ as substrate. PARP is strongly activated by DNA single- or double-strand breaks and is thought to be involved in cellular responses to DNA damage. We characterized a dominant negative PARP mutant, i.e. the DNA-binding domain of this enzyme, whose overexpression in cells leads to increased genetic instability following DNA damage. In order to study whether PARP activity is also implicated in the process of tumorigenesis, we generated stably transfected HeLa cell clones with constitutive overexpression of dominant negative PARP and investigated tumor formation of these clones in nude mice. We found that inhibition of PARP activity dramatically reduces tumor forming ability of HeLa cells. Moreover, we provide strong evidence that the observed reduction in tumor forming ability is due to increased tumor cell apoptosis in vivo. Viewed together, our data and those from other groups show that inhibition of PARP enzyme activity interferes with DNA base excision repair and leads to increased genetic instability and recombination but, on the other hand, can sensitize cells to apoptotic stimuli and by this mechanism may prevent tumor formation.

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

PubMed Central

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

2014-01-01

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

Engineered microtopographies and surface chemistries direct cell attachment and function

NASA Astrophysics Data System (ADS)

Magin, Chelsea Marie

Harrison, in 1914, first recognized that cells respond to physicochemical cues such as substratum topography when he observed that fibroblasts elongated while cultured on spider silk. Recently, techniques developed in the micro-electronics industry have been used to create molds for producing microscaled topographies with various shapes and spatial arrangements. Although these patterning techniques are well-established, very little is known about the mechanisms underlying cell sensing and response to microtopographies. In this work cellular micro-environments with varying surface topographies and chemistries were evaluated with marine organisms and mammalian cells to investigate cellular sensing and response. Biofouling---the accumulation of micro-organisms, plants, and animals on submerged surfaces---is an environmental and economic concern. Engineered topographies, replicated in polydimethylsiloxane elastomer (PDMSe) and functionalized poly(ethylene glycol)-dimethacrylate (PEGDMA) hydrogels, were evaluated for inhibition of marine fouling organism attachment. Microtopographies replicated in PDMSe inhibited attachment of the marine bacterium, Cobetia marina up to 99% versus smooth. The average normalized attachment densities of cells of C. marina and zoospores of the green algae Ulva on PDMSe topographies scaled inversely with the Engineered Roughness Index (ERIII), a representation of surface energy. Attachment densities of Ulva from four assays and C. marina from two growth phases to PDMSe surfaces scaled inversely with one equation: ERI II multiplied by the Reynolds number of the organism (Re) (R 2 = 0.77). The same microtopographies created in PDMSe reduced the initial attachment density and attachment strength of cells of the diatoms Navicula incerta and Seminavis robusta compared to smooth PDMSe. The average normalized attachment density of Navicula after exposure to shear stress (48 Pa) was correlated with the contact area between the diatom and a topographically modified surface (R2=0.82). Functionalized PEGDMA hydrogels significantly reduced attachment and attachment strength of Navicula and C. marina. These hydrogels also reduced attachment of zoospores of Ulva compared to PDMSe. Attachment of Ulva to microtopographies in PDMSe and PEGDMA-co-HEMA negatively correlated with ERIII*Re (R2 = 0.94 and R2 = 0.99, respectively). Incorporating a surface energy term into this equation created a correlation between the attachment densities of cells from two evolutionarily diverse groups on substrates of two surface chemistries with an equation that describes the various microtopographies and surface chemistries in terms of surface energy (R2 = 0.80). The current Attachment Model can now be used to design engineered antifouling surface microtopographies and chemistries that inhibit the attachment of organisms from three evoluntionarily diverse groups. Hydrogels based on PEGDMA were also chosen as a substratum material for mammalian cell culture. Capturing endothelial progenitor cells (EPCs) and inducing differentiation into the endothelial cell (EC) phenotype is the ideal way to re-endothelialize a small-diameter vascular graft. Substratum elasticity has been reported to direct stem cell differentiation into specific lineages. Functionalized PEGDMA hydrogels provided good compliance, high fidelity of topographic features and sites for surface modification with biomolecules. Fibronectin grafting and topography both increased EC attachment. This combination of adjustable elasticity, surface chemistry and topography has the potential to promote the capture and differentiation of EPCs into a confluent EC monolayer. Engineered microtopographies replicated in PDMSe directed elongation and alignment of human coronary artery endothelial cells (HCAECs) and human coronary artery smooth muscle cells (HCASMCs) compared to smooth surfaces. Engineered cellular micro-environments were created with specific surface energies defined by chemistry and topography to successfully direct cell attachment and function.

Ruxolitinib and Polycation Combination Treatment Overcomes Multiple Mechanisms of Resistance of Pancreatic Cancer Cells to Oncolytic Vesicular Stomatitis Virus

PubMed Central

Felt, Sébastien A.; Droby, Gaith N.

2017-01-01

ABSTRACT Vesicular stomatitis virus (VSV) is a promising oncolytic virus (OV). Although VSV is effective against a majority of pancreatic ductal adenocarcinoma cell (PDAC) cell lines, some PDAC cell lines are highly resistant to VSV, and the mechanisms of resistance are still unclear. JAK1/2 inhibitors (such as ruxolitinib and JAK inhibitor I) strongly stimulate VSV replication and oncolysis in all resistant cell lines but only partially improve the susceptibility of resistant PDACs to VSV. VSV tumor tropism is generally dependent on the permissiveness of malignant cells to viral replication rather than on receptor specificity, with several ubiquitously expressed cell surface molecules playing a role in VSV attachment to host cells. However, as VSV attachment to PDAC cells has never been tested before, here we examined if it was possibly inhibited in resistant PDAC cells. Our data show a dramatically weaker attachment of VSV to HPAF-II cells, the most resistant human PDAC cell line. Although sequence analysis of low-density lipoprotein (LDL) receptor (LDLR) mRNA did not reveal any amino acid substitutions in this cell line, HPAF-II cells displayed the lowest level of LDLR expression and dramatically lower LDL uptake. Treatment of cells with various statins strongly increased LDLR expression levels but did not improve VSV attachment or LDL uptake in HPAF-II cells. However, LDLR-independent attachment of VSV to HPAF-II cells was dramatically improved by treating cells with Polybrene or DEAE-dextran. Moreover, combining VSV with ruxolitinib and Polybrene or DEAE-dextran successfully broke the resistance of HPAF-II cells to VSV by simultaneously improving VSV attachment and replication. IMPORTANCE Oncolytic virus (OV) therapy is an anticancer approach that uses viruses that selectively infect and kill cancer cells. This study focuses on oncolytic vesicular stomatitis virus (VSV) against pancreatic ductal adenocarcinoma (PDAC) cells. Although VSV is effective against most PDAC cells, some are highly resistant to VSV, and the mechanisms are still unclear. Here we examined if VSV attachment to cells was inhibited in resistant PDAC cells. Our data show very inefficient attachment of VSV to the most resistant human PDAC cell line, HPAF-II. However, VSV attachment to HPAF-II cells was dramatically improved by treating cells with polycations. Moreover, combining VSV with polycations and ruxolitinib (which inhibits antiviral signaling) successfully broke the resistance of HPAF-II cells to VSV by simultaneously improving VSV attachment and replication. We envision that this novel triple-combination approach could be used in the future to treat PDAC tumors that are highly resistant to OV therapy. PMID:28566376

The Role of Depression and Attachment Styles in Predicting Students' Addiction to Cell Phones.

PubMed

Ghasempour, Abdollah; Mahmoodi-Aghdam, Mansour

2015-01-01

The present study aimed at investigating the role of depression and attachment styles in predicting cell phone addiction. In this descriptive correlational study, a sample including 100 students of Payame Noor University (PNU), Reyneh Center, Iran, in the academic year of 2013-2014 was selected using volunteer sampling. Participants were asked to complete the adult attachment inventory (AAI), Beck depression inventory-13 (BDI-13) and the cell phone overuse scale (COS). Results of the stepwise multiple regression analysis showed that depression and avoidant attachment style were the best predictors of students' cell phone addiction (R(2) = 0.23). The results of this study highlighted the predictive value of depression and avoidant attachment style concerning students' cell phone addiction.

Electromicroinjection of particles into living cells

DOEpatents

Ray, F. Andrew; Cram, L. Scott; Galey, William R.

1988-01-01

Method and apparatus for introducing particles into living cells. Fluorescently-stained human chromosomes are introduced into cultured, mitotic Chinese hamster cells using electromicroinjection. The recipient cells frequently survived the physiological perturbation imposed by a successful chromosome injection. Successfully injected recipient cells maintained viability as evidenced by their ability to be expanded. The technique relies on the surface charge of fluorescently stained chromosomes and their ability to be attracted and repelled to and from the tip of a micropipette. The apparatus includes a micropipette having a tip suitable for piercing the membrane of a target cell and an electrode inserted into the lumen thereof. The target cells and suspended particles are located in an electrically conducted solution, and the lumen of the micropipette is filled with an electrically conducting solution which contacts the electrode located therein. A second electrode is also located in the conducting solution containing the target cells and particles. Voltages applied to the electrode within the micropipette attract the particles to the region of the tip thereof. The particles adhere to the surface of the micropipette with sufficient force that insertion of the micropipette tip and attached particle through the membrane of a target cell will not dislodge the particle. By applying a voltage having the opposite polarity of the attraction voltage, the particles are expelled from the micropipette to which is then withdrawn from the cell body.

Adult Attachment Affects Neural Response to Preference-Inferring in Ambiguous Scenarios: Evidence From an fMRI Study

PubMed Central

Zhang, Xing; Ran, Guangming; Xu, Wenjian; Ma, Yuanxiao; Chen, Xu

2018-01-01

Humans are highly social animals, and the ability to cater to the preferences of other individuals is encouraged by society. Preference-inferring is an important aspect of the theory of mind (TOM). Many previous studies have shown that attachment style is closely related to TOM ability. However, little is known about the effects of adult attachment style on preferences inferring under different levels of certainty. Here, we investigated how adult attachment style affects neural activity underlying preferences inferred under different levels of certainty by using functional magnetic resonance imaging (fMRI). The fMRI results demonstrated that adult attachment influenced the activation of anterior insula (AI) and inferior parietal lobule (IPL) in response to ambiguous preference-inferring. More specifically, in the ambiguous preference condition, the avoidant attached groups exhibited a significantly enhanced activation than secure and anxious attached groups in left IPL; the anxious attached groups exhibited a significantly reduced activation secure attached group in left IPL. In addition, the anxious attached groups exhibited a significantly reduced activation than secure and avoidant attached groups in left AI. These results were also further confirmed by the subsequent PPI analysis. The results from current study suggest that, under ambiguous situations, the avoidant attached individuals show lower sensitivity to the preference of other individuals and need to invest more cognitive resources for preference-reasoning; while compared with avoidant attached group, the anxious attached individuals express high tolerance for uncertainty and a higher ToM proficiency. Results from the current study imply that differences in preference-inferring under ambiguous conditions associated with different levels of individual attachment may explain the differences in interpersonal interaction. PMID:29559932

Characterization of somatic embryo attached structures in Feijoa sellowiana Berg. (Myrtaceae).

PubMed

Correia, Sandra M; Canhoto, Jorge M

2010-06-01

The presence of an attached organ to somatic embryos of angiosperms connecting the embryo to the supporting tissue has been a subject of controversy. This study shows that 67% of the morphologically normal somatic embryos of Feijoa sellowiana possess this type of organ and that its formation was not affected by culture media composition. Histological and ultrastructural analysis indicated that the attached structures of somatic embryos displayed a great morphological diversity ranging from a few cells to massive and columnar structures. This contrast with the simple suspensors observed in zygotic embryos which were only formed by five cells. As well as the suspensor of zygotic embryos, somatic embryo attached structures undergo a process of degeneration in later stages of embryo development. Other characteristic shared by zygotic suspensors and somatic embryo attached structures was the presence of thick cell walls surrounding the cells. Elongated thin filaments were often associated with the structures attached to somatic embryos, whereas in other cases, tubular cells containing starch grains connected the embryo to the supporting tissue. These characteristics associated with the presence of plasmodesmata in the cells of the attached structures seem to indicate a role on embryo nutrition. However, cell proliferation in the attached structures resulting into new somatic embryos may also suggest a more complex relationship between the embryo and the structures connecting it to the supporting tissue.

Cell attachment properties of Portland cement-based endodontic materials: biological and methodological considerations.

PubMed

Ahmed, Hany Mohamed Aly; Luddin, Norhayati; Kannan, Thirumulu Ponnuraj; Mokhtar, Khairani Idah; Ahmad, Azlina

2014-10-01

The attachment and spreading of mammalian cells on endodontic biomaterials are an area of active research. The purpose of this review is to discuss the cell attachment properties of Portland cement (PC)-based materials by using scanning electron microscope (SEM). In addition, methodological aspects and technical challenges are discussed. A PubMed electronic search was conducted by using appropriate key words to identify the available investigations on the cell attachment properties of PC-based endodontic materials. After retrieving the full text of related articles, the cross citations were also identified. A total of 23 articles published between January 1993 and October 2013 were identified. This review summarizes the cell attachment properties of commercial and experimental PC-based materials on different cell cultures by using SEM. Methodological procedures, technical challenges, and relevance of SEM in determining the biological profile of PC-based materials are discussed. SEM observations demonstrate that commercial MTA formulations show favorable cell attachment properties, which is consistent with their successful clinical outcomes. The favorable cell attachment properties of PC and its modified formulations support its potential use as a substitute for mineral trioxide aggregate. However, researchers should carefully select cell types for their SEM investigations that would be in contact with the proposed PC-based combinations in the clinical situation. Despite being a technical challenge, SEM provides useful information on the cell attachment properties of PC-based materials; however, other assays for cell proliferation and viability are essential to come up with an accurate in vitro biological profile of any given PC-based formulation. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

The Influence of Prior Modes of Growth, Temperature, Medium, and Substrate Surface on Biofilm Formation by Antibiotic-Resistant Campylobacter jejuni.

PubMed

Teh, Amy Huei Teen; Lee, Sui Mae; Dykes, Gary A

2016-12-01

Campylobacter jejuni is one of the most common causes of bacterial gastrointestinal food-borne infection worldwide. It has been suggested that biofilm formation may play a role in survival of these bacteria in the environment. In this study, the influence of prior modes of growth (planktonic or sessile), temperatures (37 and 42 °C), and nutrient conditions (nutrient broth and Mueller-Hinton broth) on biofilm formation by eight C. jejuni strains with different antibiotic resistance profiles was examined. The ability of these strains to form biofilm on different abiotic surfaces (stainless steel, glass, and polystyrene) as well as factors potentially associated with biofilm formation (bacterial surface hydrophobicity, auto-aggregation, and initial attachment) was also determined. The results showed that cells grown as sessile culture generally have a greater ability to form biofilm (P < 0.05) compared to their planktonic counterparts. Biofilm was also greater (P < 0.05) in lower nutrient media, while growth at different temperatures affects biofilm formation in a strain-dependent manner. The strains were able to attach and form biofilms on different abiotic surfaces, but none of them demonstrated strong, complex, or structured biofilm formation. There were no clear trends between the bacterial surface hydrophobicity, auto-aggregation, attachment, and biofilm formation by the strains. This finding suggests that environmental factors did affect biofilm formation by C. jejuni, and they are more likely to persist in the environment in the form of mixed-species rather than monospecies biofilms.

Silencing by nuclear matrix attachment distinguishes cell-type specificity: association with increased proliferation capacity

PubMed Central

Linnemann, Amelia K.; Krawetz, Stephen A.

2009-01-01

DNA loop organization by nuclear scaffold/matrix attachment is a key regulator of gene expression that may provide a means to modulate phenotype. We have previously shown that attachment of genes to the NaCl-isolated nuclear matrix correlates with their silencing in HeLa cells. In contrast, expressed genes were associated with the lithium 3,5-diiodosalicylate (LIS)-isolated nuclear scaffold. To define their role in determining phenotype matrix attached regions (MARs) on human chromosomes 14–18 were identified as a function of expression in a primary cell line. The locations of MARs in aortic adventitial fibroblast (AoAF) cells were very stable (r = 0.909) and 96% of genes attached at MARs are silent (P < 0.001). Approximately one-third of the genes uniquely expressed in AoAF cells were associated with the HeLa cell nuclear matrix and silenced. Comparatively, 81% were associated with the AoAF cell nuclear scaffold (P < 0.001) and expressed. This suggests that nuclear scaffold/matrix association mediates a portion of cell type-specific gene expression thereby modulating phenotype. Interestingly, nuclear matrix attachment and thus silencing of specific genes that regulate proliferation and maintain the integrity of the HeLa cell genome suggests that transformation may at least in part be achieved through aberrant nuclear matrix attachment. PMID:19276204

In vitro evaluation of demineralized freeze-dried bone allograft in combination with enamel matrix derivative.

PubMed

Miron, Richard J; Bosshardt, Dieter D; Laugisch, Oliver; Dard, Michel; Gemperli, Anja C; Buser, Daniel; Gruber, Reinhard; Sculean, Anton

2013-11-01

Preclinical and clinical studies suggest that a combination of enamel matrix derivative (EMD) with demineralized freeze-dried bone allograft (DFDBA) may improve periodontal wound healing and regeneration. To date, no single study has characterized the effects of this combination on in vitro cell behavior. The aim of this study is to test the ability of EMD to adsorb to the surface of DFDBA particles and determine the effect of EMD coating on downstream cellular pathways such as adhesion, proliferation, and differentiation of primary human osteoblasts and periodontal ligament (PDL) cells. DFDBA particles were precoated with EMD or human blood and analyzed for protein adsorption patterns via scanning electron microscopy. Cell attachment and proliferation were quantified using a commercial assay. Cell differentiation was analyzed using real-time polymerase chain reaction for genes encoding Runx2, alkaline phosphatase, osteocalcin, and collagen 1α1, and mineralization was assessed using alizarinred staining. Analysis of cell attachment revealed no significant differences among control, blood-coated, and EMD-coated DFDBA particles. EMD significantly increased cell proliferation at 3 and 5 days after seeding for both osteoblasts and PDL cells compared to control and blood-coated samples. Moreover, there were significantly higher messenger ribonucleic acid levels of osteogenic differentiation markers, including collagen 1α1, alkaline phosphatase, and osteocalcin, in osteoblasts and PDL cells cultured on EMD-coated DFDBA particles at 3, 7, and 14 days. The results suggest that the addition of EMD to DFDBA particles may influence periodontal regeneration by stimulating PDL cell and osteoblast proliferation and differentiation.

Physiological Changes of Surface Membrane in Lactobacillus with Prebiotics.

PubMed

Pan, Mingfang; Kumaree, Kishore K; Shah, Nagendra P

2017-03-01

Synbiotics are always considered to be beneficial in healthy manipulation of gut environment; however, the purpose of this research was to investigate the dominance of synbiotic over the individual potential of probiotics and prebiotics. Four different types of prebiotics, fructo-oligosaccharides, raffinose, inulin, and cellobiose, were evaluated based on their varying degree of polymerization, combined each with 2 different Lactobacilli strains, including Lactobacillus paracasei 276 and Lactobacillus plantarum WCFS1. The effects of synbiotics combination on the surface structure were evaluated by analyzing auto-aggregation, membrane hydrophobicity, and adhesion to Caco-2 cells. Our results showed that both Lactobacilli exhibited significantly greater degree of attachment to Caco-2 cells (23.31% and 16.85%, respectively) when using cellobiose as a substrate than with other prebiotics (P < 0.05). Intestinal adhesion ability was in correlation with the percent of auto-aggregation, both Lactobacillus exhibited higher percent of auto-aggregation in cellobiose compared to other prebiotics. These behavioral changes in terms of attachment and auto-aggregation were further supported with the changes noticed from infrared spectra (FT-IR). © 2017 Institute of Food Technologists®.

Freezing Responses in DMSO-Based Cryopreservation of Human iPS Cells: Aggregates Versus Single Cells.

PubMed

Li, Rui; Yu, Guanglin; Azarin, Samira M; Hubel, Allison

2018-05-01

Inadequate preservation methods of human induced pluripotent stem cells (hiPSCs) have impeded efficient reestablishment of cell culture after the freeze-thaw process. In this study, we examined roles of the cooling rate, seeding temperature, and difference between cell aggregates (3-50 cells) and single cells in controlled rate freezing of hiPSCs. Intracellular ice formation (IIF), post-thaw membrane integrity, cell attachment, apoptosis, and cytoskeleton organization were evaluated to understand the different freezing responses between hiPSC single cells and aggregates, among cooling rates of 1, 3, and 10°C/min, and between seeding temperatures of -4°C and -8°C. Raman spectroscopy images of ice showed that a lower seeding temperature (-8°C) did not affect IIF in single cells, but significantly increased IIF in aggregates, suggesting higher sensitivity of aggregates to supercooling. In the absence of IIF, Raman images showed greater variation of dimethyl sulfoxide concentration across aggregates than single cells, suggesting cryoprotectant transport limitations in aggregates. The ability of cryopreserved aggregates to attach to culture substrates did not correlate with membrane integrity for the wide range of freezing parameters, indicating inadequacy of using only membrane integrity-based optimization metrics. Lower cooling rates (1 and 3°C/min) combined with higher seeding temperature (-4°C) were better at preventing IIF and preserving cell function than a higher cooling rate (10°C/min) or lower seeding temperature (-8°C), proving the seeding temperature range of -7°C to -12°C from literature to be suboptimal. Unique f-actin cytoskeletal organization into a honeycomb-like pattern was observed in postpassage and post-thaw colonies and correlated with successful reestablishment of cell culture.

Microbiological test results using three urine pretreatment regimes with 316L stainless steel

NASA Technical Reports Server (NTRS)

Huff, Timothy L.

1993-01-01

Three urine pretreatments, (1) Oxone (Dupont) and sulfuric acid, (2) sodium hypochlorite and sulfuric acid, (3) and ozone, were studied for their ability to reduce microbial levels in urine and minimize surface attachment to 316L stainless steel coupons. Urine samples inoculated with Bacillus insolitus and a filamentous mold, organisms previously recovered from the vapor compression distillation subsystem of NASA Space Station Freedom water recovery test were tested in glass corrosion cells containing base or weld metal coupons. Microbial levels, changes in pH, color, turbidity, and odor of the fluid were monitored over the course of the 21-day test. Specimen surfaces were examined by scanning electron microscopy at completion of the test for microbial attachment. Ozonated urine samples were less turbid and had lower microbial levels than controls or samples receiving other pretreatments. Base metal coupons receiving pretreatment were relatively free of attached bacteria. However, well-developed biofilms were found in the heat-affected regions of welded coupons receiving Oxone and hypochlorite pretreatments. Few bacteria were observed in the same regions of the ozone pretreatment sample.

Modulation of osteoblast attachment and growth in vitro by inertial forces

NASA Astrophysics Data System (ADS)

Kacena, Melissa Ann

1999-11-01

Spaceflight exploration and associated experiments show that human bones lose in density during inertial unloading, due principally to their demineralization. This research project examines the effect of gravity on osteoblast attachment and function in various inertial environments. Chicken calvarial osteoblasts were cultured under the following inertial conditions: spaceflight, simulated shuttle launch accelerations and vibrations, centrifugation, clino-rotation, and inversion. Cultures exposed to these conditions were compared with cultures grown in the laboratory as static 1G controls. Electron and light microscopy revealed the number of total osteoblasts attached to their substrate. Biochemical assays discerned changes in viable cell number, alkaline phosphatase levels, and mineralization. Immunohistochemical assays were used to investigate differences in cytoskeletal and extracellular matrix protein concentrations in the cultures, the percentage of proliferative cells, and cell viability. Compared to controls, spaceflight results indicated that the number of attached osteoblast cells was reduced. Launch simulation data indicated that the associated accelerations and vibrations may contribute to the reduction of attached osteoblasts in spaceflight cultures. Following centrifugation, the number of attached cells was unaltered; however, immunostaining of actin, fibronectin, and vinculin did show alterations in cultures exposed to hypergravity. Confluent cultures that were right side up, inverted, and clino-rotated contained a comparable number of attached cells and functioned similarly on the basis of measured alkaline phosphatase and bound calcium content. Sparse clino-rotated or inverted cultures showed an immediate response of diminished viable osteoblast numbers, but this effect disappeared with time and all remaining attached cells functioned similarly (APase and bound calcium). On the basis of these data osteoblast attachment and function in confluent cultures is minimally, if at all, affected by alterations in inertial environments. However, in sparse cultures about half as many cells are found attached initially. The remaining attached cells appear to multiply and function normally. These results suggest that the effects of spaceflight on bone are thus not likely to be caused by direct intrinsic effects of gravity on single osteoblasts that can be simulated in laboratory experiments in vitro experiments.

Immobilization of Delftia tsuruhatensis in macro-porous cellulose and biodegradation of phenolic compounds in repeated batch process.

PubMed

Juarez Jimenez, B; Reboleiro Rivas, P; Gonzalez Lopez, J; Pesciaroli, C; Barghini, P; Fenice, M

2012-01-01

Delftia tsuruhatensis BM90, previously isolated from Tyrrhenian Sea and selected for its ability to degrade a wide array of phenolic compounds, was immobilized in chemically modified macro porous cellulose. The development of bacterial adhesion on the selected carrier was monitored by scanning electron microscopy. Evident colonization started already after 8h of incubation. After 72h, almost all the carrier surface was covered by the bacterial cells. Extracellular bacterial structures, such as pili or fimbriae, contributed to carrier colonization and cell attachment. Immobilized cells of D. tsuruhatensis were tested for their ability to biodegrade a pool of 20 phenols in repeated batch process. During the first activation batch (72h), 90% of phenols degradation was obtained already in 48h. In the subsequent batches (up to 360h), same degradation was obtained after 24h only. By contrast, free cells were slower: to obtain almost same degradation, 48h were needed. Thus, process productivity, achieved by the immobilized cells, was double than that of free cells. Specific activity was also higher suggesting that the use of immobilized D. tsuruhatensis BM90 could be considered very promising in order to obtain an efficient reusable biocatalyst for long-term treatment of phenols containing effluents. Copyright © 2011 Elsevier B.V. All rights reserved.

Mechanisms of Ricin Toxin Neutralization Revealed through Engineered Homodimeric and Heterodimeric Camelid Antibodies*

PubMed Central

Herrera, Cristina; Tremblay, Jacqueline M.; Shoemaker, Charles B.; Mantis, Nicholas J.

2015-01-01

Novel antibody constructs consisting of two or more different camelid heavy-chain only antibodies (VHHs) joined via peptide linkers have proven to have potent toxin-neutralizing activity in vivo against Shiga, botulinum, Clostridium difficile, anthrax, and ricin toxins. However, the mechanisms by which these so-called bispecific VHH heterodimers promote toxin neutralization remain poorly understood. In the current study we produced a new collection of ricin-specific VHH heterodimers, as well as VHH homodimers, and characterized them for their ability neutralize ricin in vitro and in vivo. We demonstrate that the VHH heterodimers, but not homodimers were able to completely protect mice against ricin challenge, even though the two classes of antibodies (heterodimers and homodimers) had virtually identical affinities for ricin holotoxin and similar IC50 values in a Vero cell cytotoxicity assay. The VHH heterodimers did differ from the homodimers in their ability to promote toxin aggregation in solution, as revealed through analytical ultracentrifugation. Moreover, the VHH heterodimers that were most effective at promoting ricin aggregation in solution were also the most effective at blocking ricin attachment to cell surfaces. Collectively, these data suggest that heterodimeric VHH-based neutralizing agents may function through the formation of antibody-toxin complexes that are impaired in their ability to access host cell receptors. PMID:26396190

Cell behavior on gallium nitride surfaces: peptide affinity attachment versus covalent functionalization.

PubMed

Foster, Corey M; Collazo, Ramon; Sitar, Zlatko; Ivanisevic, Albena

2013-07-02

Gallium nitride is a wide band gap semiconductor that demonstrates a unique set of optical and electrical properties as well as aqueous stability and biocompatibility. This combination of properties makes gallium nitride a strong candidate for use in chemical and biological applications such as sensors and neural interfaces. Molecular modification can be used to enhance the functionality and properties of the gallium nitride surface. Here, gallium nitride surfaces were functionalized with a PC12 cell adhesion promoting peptide using covalent and affinity driven attachment methods. The covalent scheme proceeded by Grignard reaction and olefin metathesis while the affinity driven scheme utilized the recognition peptide isolated through phage display. This study shows that the method of attaching the adhesion peptide influences PC12 cell adhesion and differentiation as measured by cell density and morphological analysis. Covalent attachment promoted monolayer and dispersed cell adhesion while affinity driven attachment promoted multilayer cell agglomeration. Higher cell density was observed on surfaces modified using the recognition peptide. The results suggest that the covalent and affinity driven attachment methods are both suitable for promoting PC12 cell adhesion to the gallium nitride surface, though each method may be preferentially suited for distinct applications.

Escherichia coli attachment to model particulates: The effects of bacterial cell characteristics and particulate properties.

PubMed

Liang, Xiao; Liao, Chunyu; Soupir, Michelle L; Jarboe, Laura R; Thompson, Michael L; Dixon, Philip M

2017-01-01

E. coli bacteria move in streams freely in a planktonic state or attached to suspended particulates. Attachment is a dynamic process, and the fraction of attached microorganisms is thought to be affected by both bacterial characteristics and particulate properties. In this study, we investigated how the properties of cell surfaces and stream particulates influence attachment. Attachment assays were conducted for 77 E. coli strains and three model particulates (ferrihydrite, Ca-montmorillonite, or corn stover) under environmentally relevant conditions. Surface area, particle size distribution, and total carbon content were determined for each type of particulate. Among the three particulates, attachment fractions to corn stover were significantly larger than the attachments to 2-line ferrihydrite (p-value = 0.0036) and Ca-montmorillonite (p-value = 0.022). Furthermore, attachment to Ca-montmorillonite and corn stover was successfully modeled by a Generalized Additive Model (GAM) using cell characteristics as predictor variables. The natural logarithm of the net charge on the bacterial surface had a significant, positive, and linear impact on the attachment of E. coli bacteria to Ca-montmorillonite (p-value = 0.013), but it did not significantly impact the attachment to corn stover (p-value = 0.36). The large diversities in cell characteristics among 77 E. coli strains, particulate properties, and attachment fractions clearly demonstrated the inadequacy of using a static parameter or linear coefficient to predict the attachment behavior of E. coli in stream water quality models.

3D culturing and differentiation of SH-SY5Y neuroblastoma cells on bacterial nanocellulose scaffolds.

PubMed

Innala, Marcus; Riebe, Ilse; Kuzmenko, Volodymyr; Sundberg, Johan; Gatenholm, Paul; Hanse, Eric; Johannesson, Sara

2014-10-01

A new in vitro model, mimicking the complexity of nerve tissue, was developed based on a bacterial nanocellulose (BNC) scaffold that supports 3D culturing of neuronal cells. BNC is extracellularly excreted by Gluconacetobacter xylinus (G. xylinus) in the shape of long non-aggregated nanofibrils. The cellulose network created by G. xylinus has good mechanical properties, 99% water content, and the ability to be shaped into 3D structures by culturing in different molds. Surface modification with trimethyl ammonium beta-hydroxypropyl (TMAHP) to induce a positive surface charge, followed by collagen I coating, has been used to improve cell adhesion, growth, and differentiation on the scaffold. In the present study, we used SH-SY5Y neuroblastoma cells as a neuronal model. These cells attached and proliferated well on the BNC scaffold, as demonstrated by scanning electron microscopy (SEM) and the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) assay. Following neuronal differentiation, we demonstrated functional action potentials (APs) by electrophysiological recordings, indicating the presence of mature neurons on the scaffolds. In conclusion, we have demonstrated for the first time that neurons can attach, proliferate, and differentiate on BNC. This 3D model based on BNC scaffolds could possibly be used for developing in vitro disease models, when combined with human induced pluripotent stem (iPS) cells (derived from diseased patients) for detailed investigations of neurodegenerative disease mechanisms and in the search for new therapeutics.

Mouse embryo attachment to substratum and interaction of trophoblast with cultured cells

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

Glass, R.H.; Spindle, A.I.; Pedersen, R.A.

1979-06-01

Hatching, attachment, and trophoblast outgrowth of mouse embryos in vitro were examined as a model for implantation. Mouse embryos attached and grew out on glass cover slips that were partially covered with cultured mouse cells (L cells, liver cells, transformed JLS-V11 cells, and teratocarcinoma cells). Scanning electron microscopy showed that processes of these cells made contact with trophoblast, but there was no evidence of cell lysis or of phagocytosis of the cells by trophoblast. Time-lapse cinematography showed that after contact the cultured mouse cells retracted from the trophoblast, which then spread into the areas vacated by those cells. This suggestsmore » a means by which the trophoblast gains entry into the endometrium without destruction of maternal cells. Neuraminidase (100 or 250 units/ml) had no effect on attachment of mouse embryos to glass. However, attachment was inhibited by trypsin at concentrations of 0.25%, 0.025%, and 0.0025%. Treatment of early blastocysts with diazooxo-norleucine, an inhibitor of glycoprotein synthesis, decreased the number of embryos hatching from the zona pellucida; treatment at the late blastocyst stage decreased hatching to a lesser extent. Among the late blastocysts that did hatch, the number forming trophoblast outgrowths was lower than in controls. These results suggest that glycoproteins may be of importance for embryo hatching, attachment, and outgrowth.« less

Effect of C-implantation on Nerve-Cell Attachment to Polystyrene Films

NASA Astrophysics Data System (ADS)

Sommani, Piyanuch; Tsuji, Hiroshi; Kitamura, Tsuyoshi; Hattori, Mitsutaka; Yamada, Tetsuya; Sato, Hiroko; Gotoh, Yasuhito; Ishikawa, Junzo

The surfaces of the polystyrene films spin-coated on glass were modified by carbon negative-ion implantation with various ion doses from 1×1014 to 3×1016 ions/cm2 at 5 and 10 keV. The implantation conditions with and without a pattering mask were for investigation of the cell-attachment properties and for evaluation of surface physical properties of contact angle, respectively. The contact angles of modified surface were investigated by pure water drop and air bubble method. The lowest angle value of the implanted films at 5 and 10 keV were approximately 72° at 3×1015 ions/cm2 after dipping in the de-ionized water for 2 hours. The lowering of contact angles on C-implanted surfaces when increase the ion dose is due to formation of the OH and C-O bonds. Nerve-cell-attachment properties of modified surface were investigated by the nerve-like cell of rat adrenal pheochromocytoma (PC12h) in vitro. After 2 days culture of the PC12h cells, no cells attached on the polystyrene films implanted with low ion dose from 1×1014 to 3×1014 ions/cm2. On the polystyrene films implanted with the dose order of 1015 ions/cm2, the cells selectively attached only on the implanted region. Whereas on the surfaces implanted with high dose such as 1×1016 and 3×1016 ions/cm2 mostly cells attached on the implanted region, and some attached on the unimplanted region, as well as cells were abnormal in shape and large size. Therefore, the suitable dose implantation for the selective-attachment of nerve-cells on the polystyrene films implanted at 5 and 10 keV were obtained around the dose order of 1015 ions/cm2, and the best condition for the selective attachment properties was at 3×1015 ions/cm2 corresponding to the lowest contact angle.

Nature of a Red Cell Sensitizing Substance from Streptococci1

PubMed Central

Jackson, Robert W.; Moskowitz, Merwin

1966-01-01

Jackson, Robert W. (Purdue University, Lafayette, Ind.), and Merwin Moskowitz. Nature of a red cell sensitizing substance from streptococci. J. Bacteriol. 91:2205–2209. 1966.—A method for purifying a streptococcal antigen which sensitizes red cells to agglutination by antiserum is described. The antigen, when purified by this method, is almost exclusively composed of glycerophosphate and d-alanine. The ratio of alanine to glycerophosphate varies from 1:5 to 1:3. The glycerophosphate is polymerized and is thus a teichoic acid. The polyglycerophosphate appears to be the antigenic determinant for agglutination. d-Alanine is readily removed by mild base and appears to be necessary for the attachment of the teichoic acid to red cells. Quantitative removal of alanine does not affect the ability of the polymer to absorb antibody from serum. PMID:5329284

Contributions of Hierarchical Attachments to Student Adaptation to College

ERIC Educational Resources Information Center

Michael, Cobi Sue

2017-01-01

The purpose of this study was to determine whether attachment, as measured dimensionally in a number of significant relationships (mother, father, romantic partner, best friend, social group, and institution), was associated with undergraduate students' ability to adapt to college. A second goal of this study was to determine if attachment exists…

Targeted drug delivery using genetically engineered diatom biosilica.

PubMed

Delalat, Bahman; Sheppard, Vonda C; Rasi Ghaemi, Soraya; Rao, Shasha; Prestidge, Clive A; McPhee, Gordon; Rogers, Mary-Louise; Donoghue, Jacqueline F; Pillay, Vinochani; Johns, Terrance G; Kröger, Nils; Voelcker, Nicolas H

2015-11-10

The ability to selectively kill cancerous cell populations while leaving healthy cells unaffected is a key goal in anticancer therapeutics. The use of nanoporous silica-based materials as drug-delivery vehicles has recently proven successful, yet production of these materials requires costly and toxic chemicals. Here we use diatom microalgae-derived nanoporous biosilica to deliver chemotherapeutic drugs to cancer cells. The diatom Thalassiosira pseudonana is genetically engineered to display an IgG-binding domain of protein G on the biosilica surface, enabling attachment of cell-targeting antibodies. Neuroblastoma and B-lymphoma cells are selectively targeted and killed by biosilica displaying specific antibodies sorbed with drug-loaded nanoparticles. Treatment with the same biosilica leads to tumour growth regression in a subcutaneous mouse xenograft model of neuroblastoma. These data indicate that genetically engineered biosilica frustules may be used as versatile 'backpacks' for the targeted delivery of poorly water-soluble anticancer drugs to tumour sites.

MC3T3-E1 cell response of amorphous phase/TiO2 nanocrystal composite coating prepared by microarc oxidation on titanium.

PubMed

Zhou, Rui; Wei, Daqing; Yang, Haoyue; Feng, Wei; Cheng, Su; Li, Baoqiang; Wang, Yaming; Jia, Dechang; Zhou, Yu

2014-06-01

Bioactive amorphous phase/TiO2 nanocrystal (APTN) composite coatings were fabricated by microarc oxidation (MAO) on Ti. The APTN coatings are composed of much amorphous phase with Si, Na, Ca, Ti and O elements and a few TiO2 nanocrystals. With increasing applied voltage, the micropore density of the APTN coating decreases and the micropore size of the APTN coating increases. The results indicate that less MC3T3-E1 cells attach on the APTN coatings as compared to Ti. However, the APTN coatings greatly enhance the cell proliferation ability and the activity of alkaline phosphatase. The amorphous phase and the concentrations of the released Ca and Si from the APTN coatings during cell culture have significant effects on the cell response. Copyright © 2014 Elsevier B.V. All rights reserved.

Adhesion of Asaia bogorensis to Glass and Polystyrene in the Presence of Cranberry Juice.

PubMed

Antolak, Hubert; Kregiel, Dorota; Czyzowska, Agata

2015-06-01

The aim of the study was to evaluate the adhesion abilities of the acetic acid bacterium Asaia bogorensis to glass and polystyrene in the presence of American cranberry (Vaccinium macrocarpon) juice. The strain of A. bogorensis used was isolated from spoiled commercial fruit-flavored drinking water. The cranberry juice was analyzed for polyphenols, organic acids, and carbohydrates using high-performance liquid chromatography and liquid chromatography-mass spectrometry techniques. The adhesive abilities of bacterial cells in culture medium supplemented with cranberry juice were determined using luminometry and microscopy. The viability of adhered and planktonic bacterial cells was determined by the plate count method, and the relative adhesion coefficient was calculated. This strain of A. bogorensis was characterized by strong adhesion properties that were dependent upon the type of surface. The highest level of cell adhesion was found on the polystyrene. However, in the presence of 10% cranberry juice, attachment of bacterial cells was three times lower. Chemical analysis of juice revealed the presence of sugars, organic acids, and anthocyanins, which were identified as galactosides, glucosides, and arabinosides of cyanidin and peonidin. A-type proanthocyanidins responsible for the antiadhesion properties of V. macrocarpon also were detected.

Synergistic Effect of Carbon Nanotubes and Graphene on Diopside Scaffolds

PubMed Central

Liu, Tingting; Wu, Ping; Gao, Chengde; Feng, Pei; Xiao, Tao; Deng, Youwen; Shuai, Cijun; Peng, Shuping

2016-01-01

A synergetic effect between carbon nanotubes (CNTs) and graphene on diopside (Di) scaffolds was demonstrated. 3D network architecture in the matrix was formed through the 1D CNTs inlaid among the 2D graphene platelets (GNPs). The mechanical properties of the CNTs/GNPs/Di scaffolds were significantly improved compared with the CNTs/Di scaffolds and GNPs/Di scaffolds. In addition, the scaffolds exhibited excellent apatite-forming ability, a modest degradation rate, and stable mechanical properties in simulated body fluid (SBF). Moreover, cell culturing tests indicated that the scaffolds supported the cells attachment and proliferation. Taken together, the CNTs/GNPs/Di scaffolds offered great potential for bone tissue engineering. PMID:27144173

Surface modification via strain-promoted click reaction facilitates targeted lentiviral transduction.

PubMed

Chu, Yanjie; Oum, Yoon Hyeun; Carrico, Isaac S

2016-01-01

As a result of their ability to integrate into the genome of both dividing and non-dividing cells, lentiviruses have emerged as a promising vector for gene delivery. Targeted gene transduction of specific cells and tissues by lentiviral vectors has been a major goal, which has proven difficult to achieve. We report a novel targeting protocol that relies on the chemoselective attachment of cancer specific ligands to unnatural glycans on lentiviral surfaces. This strategy exhibits minimal perturbation on virus physiology and demonstrates remarkable flexibility. It allows for targeting but can be more broadly useful with applications such as vector purification and immunomodulation. Copyright © 2015 Elsevier Inc. All rights reserved.

Attachment Anxiety is Linked to Alterations in Cortisol Production and Cellular Immunity

PubMed Central

Jaremka, Lisa M.; Glaser, Ronald; Loving, Timothy J.; Malarkey, William B.; Stowell, Jeffrey R.; Kiecolt-Glaser, Janice K.

2013-01-01

Although evidence suggests that attachment anxiety may increase risk for health problems, the mechanisms are not well understood. Married couples (N = 85, Mage = 38.67) provided saliva samples over three days and blood samples on two occasions. Participants with higher attachment anxiety produced more cortisol and had fewer numbers of CD3+ T-cells, CD45+ T-cells, CD3+CD4+ helper T-cells, and CD3+CD8+ cytotoxic T-cells than those with lower attachment anxiety. Higher cortisol was also related to fewer numbers of CD3+, CD45+, CD3+CD4+, and CD3+CD8+, which is mechanistically consistent with research showing that cortisol alters the cellular immune response. These data suggest that attachment anxiety may have physiological costs and provide a glimpse into the pathways through which social relationships impact health. The current study also extends attachment theory in an important new direction by utilizing a psychoneuroimmunological approach to the study of attachment anxiety, stress, and health. PMID:23307944

Sonication reduces the attachment of Salmonella Typhimurium ATCC 14028 cells to bacterial cellulose-based plant cell wall models and cut plant material.

PubMed

Tan, Michelle S F; Rahman, Sadequr; Dykes, Gary A

2017-04-01

This study investigated the removal of bacterial surface structures, particularly flagella, using sonication, and examined its effect on the attachment of Salmonella Typhimurium ATCC 14028 cells to plant cell walls. S. Typhimurium ATCC 14028 cells were subjected to sonication at 20 kHz to remove surface structures without affecting cell viability. Effective removal of flagella was determined by staining flagella of sonicated cells with Ryu's stain and enumerating the flagella remaining by direct microscopic counting. The attachment of sonicated S. Typhimurium cells to bacterial cellulose-based plant cell wall models and cut plant material (potato, apple, lettuce) was then evaluated. Varying concentrations of pectin and/or xyloglucan were used to produce a range of bacterial cellulose-based plant cell wall models. As compared to the non-sonicated controls, sonicated S. Typhimurium cells attached in significantly lower numbers (between 0.5 and 1.0 log CFU/cm 2 ) to all surfaces except to the bacterial cellulose-only composite without pectin and xyloglucan. Since attachment of S. Typhimurium to the bacterial cellulose-only composite was not affected by sonication, this suggests that bacterial surface structures, particularly flagella, could have specific interactions with pectin and xyloglucan. This study indicates that sonication may have potential applications for reducing Salmonella attachment during the processing of fresh produce. Copyright © 2016 Elsevier Ltd. All rights reserved.

XanoMatrix surfaces as scaffolds for mesenchymal stem cell culture and growth

PubMed Central

Bhardwaj, Garima; Webster, Thomas J

2016-01-01

Stem cells are being widely investigated for a wide variety of applications in tissue engineering due to their ability to differentiate into a number of cells such as neurons, osteoblasts, and fibroblasts. This ability of stem cells to differentiate into different types of cells is greatly based on mechanical and chemical cues received from their three-dimensional environments. All organs are formed by a number of cells linked together via an extracellular matrix (ECM). The ECM is a complex network of proteins and carbohydrates, which occupies intercellular spaces and regulates cellular activity by controlling cell adhesion, migration, proliferation, and differentiation. The ECM is composed of two main types of macromolecules, namely, polysaccharide glycosaminoglycans, which are covalently attached to proteins in the form of proteoglycans and fibrous proteins belonging to two functional groups, structural (collagen and elastin) and adhesive (fibronectin, laminin, vitronectin, etc). Tissue engineering is a multidisciplinary field that aims to develop biomimetic scaffolds that emulate properties of the ECM to help repair or regenerate diseased or damaged tissue. This study introduces one of these matrices, XanoMatrix, as an optimal scaffold for tissue engineering applications, in particular, for stem cell research, based on its composition, nanofibrous structure, and porosity. Results of this study suggest that XanoMatrix scaffolds are promising for stem cell tissue engineering applications and as improved cell culture inserts for studying stem cell functions (compared to traditional Corning and Falcon cell culture plates) and, thus, should be further studied. PMID:27354795

Boron nitride nanotubes for gene silencing.

PubMed

Şen, Özlem; Çobandede, Zehra; Emanet, Melis; Bayrak, Ömer Faruk; Çulha, Mustafa

2017-09-01

Non-viral gene delivery is increasingly investigated as an alternative to viral vectors due to low toxicity and immunogenicity, easy preparation, tissue specificity, and ability to transfer larger sizes of genes. In this study, boron nitride nanotubes (BNNTs) are functionalized with oligonucleotides (oligo-BNNTs). The morpholinos complementary to the oligonucleotides attached to the BNNTs (morpholino/oligo-BNNTs) are hybridized to silence the luciferase gene. The morpholino/oligo-BNNTs conjugates are administered to luciferase-expressing cells (MDA-MB-231-luc2) and the luciferase activity is monitored. The luciferase activity is decreased when MDA-MB-231-luc2 cells were treated with morpholino/oligo-BNNTs. The study suggests that BNNTs can be used as a potential vector to transfect cells. BNNTs are potential new nanocarriers for gene delivery applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Bioreactor and methods for producing synchronous cells

NASA Technical Reports Server (NTRS)

Helmstetter, Charles E. (Inventor); Thornton, Maureen (Inventor); Gonda, Steve (Inventor)

2005-01-01

Apparatus and methods are directed to a perfusion culture system in which a rotating bioreactor is used to grow cells in a liquid culture medium, while these cells are attached to an adhesive-treated porous surface. As a result of this arrangement and its rotation, the attached cells divide, with one cell remaining attached to the substrate, while the other cell, a newborn cell is released. These newborn cells are of approximately the same age, that are collected upon leaving the bioreactor. The populations of newborn cells collected are of synchronous and are minimally, if at all, disturbed metabolically.

Surface engineering approaches to micropattern surfaces for cell-based assays.

PubMed

Falconnet, Didier; Csucs, Gabor; Grandin, H Michelle; Textor, Marcus

2006-06-01

The ability to produce patterns of single or multiple cells through precise surface engineering of cell culture substrates has promoted the development of cellular bioassays that provide entirely new insights into the factors that control cell adhesion to material surfaces, cell proliferation, differentiation and molecular signaling pathways. The ability to control shape and spreading of attached cells and cell-cell contacts through the form and dimension of the cell-adhesive patches with high precision is important. Commitment of stem cells to different specific lineages depends strongly on cell shape, implying that controlled microenvironments through engineered surfaces may not only be a valuable approach towards fundamental cell-biological studies, but also of great importance for the design of cell culture substrates for tissue engineering. Furthermore, cell patterning is an important tool for organizing cells on transducers for cell-based sensing and cell-based drug discovery concepts. From a material engineering standpoint, patterning approaches have greatly profited by combining microfabrication technologies, such as photolithography, with biochemical functionalization to present to the cells biological cues in spatially controlled regions where the background is rendered non-adhesive ("non-fouling") by suitable chemical modification. The focus of this review is on the surface engineering aspects of biologically motivated micropatterning of two-dimensional (flat) surfaces with the aim to provide an introductory overview and critical assessment of the many techniques described in the literature. In particular, the importance of non-fouling surface chemistries, the combination of hard and soft lithography with molecular assembly techniques as well as a number of less well known, but useful patterning approaches, including direct cell writing, are discussed.

A novel pressed porous silicon-polycaprolactone composite as a dual-purpose implant for the delivery of cells and drugs to the eye.

PubMed

Irani, Yazad D; Tian, Yuan; Wang, Mengjia; Klebe, Sonja; McInnes, Steven J; Voelcker, Nicolas H; Coffer, Jeffery L; Williams, Keryn A

2015-10-01

Dysfunction of corneal epithelial stem cells can result in painful and blinding disease of the ocular surface. In such cases, treatment may involve transfer of growth factor and normal adult stem cells to the ocular surface. Our purpose was to develop an implantable scaffold for the delivery of drugs and cells to the ocular surface. We examined the potential of novel composite biomaterials fabricated from electrospun polycaprolactone (PCL) fibres into which nanostructured porous silicon (pSi) microparticles of varying sizes (150-250 μm or <40 μm) had been pressed. The PCL fabric provided a flexible support for mammalian cells, whereas the embedded pSi provided a substantial surface area for efficient delivery of adsorbed drugs and growth factors. Measurements of tensile strength of these composites revealed that the pSi did not strongly influence the mechanical properties of the polymer microfiber component for the Si loadings evaluated. Human lens epithelial cells (SRA01/04) attached to the composite materials, and exhibited enhanced attachment and growth when the materials were coated with foetal bovine serum. To examine the ability of the materials to deliver a small-drug payload, pSi microparticles were loaded with fluorescein diacetate prior to cell attachment. After 6 hours (h), cells exhibited intracellular fluorescence, indicative of transfer of the fluorescein diacetate into viable cells and its subsequent enzymatic conversion to fluorescein. To investigate loading of large-molecule biologics, murine BALB/c 3T3 cells, responsive to epidermal growth factor, insulin and transferrin, were seeded on composite materials. The cells showed significantly more proliferation at 48 h when seeded on composites loaded with these biologics, than on unloaded composites. No cell proliferation was observed on PCL alone, indicating the biologics had loaded into the pSi microparticles. Drug release, measured by ELISA for insulin, indicated a burst followed by a slower, continuous release over six days. When implanted under the rat conjunctiva, the most promising composite material did not cause significant neovascularization but did elicit a macrophage and mild foreign body response. These novel pressed pSi-PCL materials have potential for delivery of both small and large drugs that can be released in active form, and can support the growth of mammalian cells. Copyright © 2015 Elsevier Ltd. All rights reserved.

Effect of tissue scaffold topography on protein structure monitored by fluorescence spectroscopy.

PubMed

Portugal, Carla A M; Truckenmüller, Roman; Stamatialis, Dimitrios; Crespo, João G

2014-11-10

The impact of surface topography on the structure of proteins upon adhesion was assessed through non-invasive fluorescence monitoring. This study aimed at obtaining a better understanding about the role of protein structural status on cell-scaffold interactions. The changes induced upon adsorption of two model proteins with different geometries, trypsin (globular conformation) and fibrinogen (rod-shaped conformation) on poly-l-lactic acid (PLLA) scaffolds with different surface topographies, flat, fibrous and surfaces with aligned nanogrooves, were assessed by fluorescence spectroscopy monitoring, using tryptophan as structural probe. Hence, the maximum emission blue shift and the increase of fluorescence anisotropy observed after adsorption of globular and rod-like shaped proteins on surfaces with parallel nanogrooves were ascribed to more intense protein-surface interactions. Furthermore, the decrease of fluorescence anisotropy observed upon adsorption of proteins to scaffolds with fibrous morphology was more significant for rod-shaped proteins. This effect was associated to the ability of these proteins to adjust to curved surfaces. The additional unfolding of proteins induced upon adsorption on scaffolds with a fibrous morphology may be the reason for better cell attachment there, promoting an easier access of cell receptors to initially hidden protein regions (e.g. RGDS sequence), which are known to have a determinant role in cell attaching processes. Copyright © 2014 Elsevier B.V. All rights reserved.

Fibronectin Attachment Protein (FAP) From Bacillus Calmette-Guerin As Targeting Agent For Bladder Tumor Cells

PubMed Central

Coon, Brian G.; Crist, Scott; González-Bonet, Andrés M.; Kim, Hee-Kwon; Sowa, Jennifer; Thompson, David H.; Ratliff, Timothy L.; Aguilar, R. Claudio

2011-01-01

The adjuvant therapy of choice for superficial bladder cancer is the intravesical instillation of live Mycobacterium bovis Bacillus Calmette-Guerin (BCG). In spite of the fact that this therapy is the most effective treatment for superficial bladder cancer, intravesical administration of BCG is associated with high local morbidity and the potential for systemic infection. Therefore, there is a need for the development of safer, less toxic approaches to fight this disease. Since fibronectin attachment protein (FAP) is a key element in BCG retention and targeting to cells, we hypothesize that this protein can be used as targeting agent to deliver cytotoxic cargo for the treatment of bladder tumors. Here we evaluated the ability of bladder tumor cells to bind and endocytose FAP via fibronectin-integrin complexes. We found that microaggregation induced by an anti-FAP polyclonal antibody accelerated FAP uptake by T24 bladder tumor cells. FAP was determined to be internalized via a clathrin-independent, caveolae-dependent mechanism. Further, once within the endosomal compartment, FAP was targeted to the lysosomal compartment with negligible recycling to the plasma membrane. Importantly, we demonstrated that FAP microaggregation and internalization could also be triggered by multivalent Ni2+NTA-bearing liposomes. Overall, our studies validate the use of FAP as a targeting vector and provide the foundation for the design of more effective, less toxic bladder cancer therapeutics. PMID:21901746

Maintenance of a functional higher order chromatin structure: The role of the nuclear matrix in normal and disease states

PubMed Central

Linnemann, Amelia K.; Krawetz, Stephen A.

2010-01-01

Summary The ordered packaging of DNA within the nucleus of somatic cells reflects a dynamic supportive structure that facilitates stable transcription interrupted by intermittent cycles of extreme condensation. This dynamic mode of packing and unpacking chromatin is intimately linked to the ability of the genome to specifically complex with both histones and non-histone proteins. Understanding the underlying mechanism that governs the formation of higher order chromatin structures is a key to understanding how local architecture modulates transcription. In part, the formation of these structures appears to be regulated through genomic looping that is dynamically mediated by attachment to the nuclear scaffold/matrix at S/MARs, i.e., Scaffold/Matrix Attachment Regions. Although the mechanism guiding the formation and use of these higher-ordered structures remains unknown, S/MARs continue to reveal a multitude of roles in development and the pathogenesis of disease. PMID:20948980

Maintenance of a functional higher order chromatin structure: The role of the nuclear matrix in normal and disease states.

PubMed

Linnemann, Amelia K; Krawetz, Stephen A

2009-01-01

The ordered packaging of DNA within the nucleus of somatic cells reflects a dynamic supportive structure that facilitates stable transcription interrupted by intermittent cycles of extreme condensation. This dynamic mode of packing and unpacking chromatin is intimately linked to the ability of the genome to specifically complex with both histones and non-histone proteins. Understanding the underlying mechanism that governs the formation of higher order chromatin structures is a key to understanding how local architecture modulates transcription. In part, the formation of these structures appears to be regulated through genomic looping that is dynamically mediated by attachment to the nuclear scaffold/matrix at S/MARs, i.e., Scaffold/Matrix Attachment Regions. Although the mechanism guiding the formation and use of these higher-ordered structures remains unknown, S/MARs continue to reveal a multitude of roles in development and the pathogenesis of disease.

Attachment Security and Language Development in an Italian Sample: The Role of Premature Birth and Maternal Language

ERIC Educational Resources Information Center

Costantini, Alessandro; Cassibba, Rosalinda; Coppola, Gabrielle; Castoro, Germana

2012-01-01

We investigated the influence of biological immaturity and attachment security on linguistic development and tested whether maternal language mediated the impact of security on the child's linguistic abilities. Forty mother-child dyads were followed longitudinally, with the child's attachment security assessed at 24 months of age through trained…

Laminin and Fibronectin in Cell Adhesion: Enhanced Adhesion of Cells from Regenerating Liver to Laminin

NASA Astrophysics Data System (ADS)

Carlsson, Roland; Engvall, Eva; Freeman, Aaron; Ruoslahti, Erkki

1981-04-01

Laminin, a basement membrane glycoprotein isolated from cultures of mouse endodermal cells and rat yolk sac carcinoma cells, promoted the attachment of liver cells obtained from regenerating mouse liver. Cells from normal mouse liver attached readily to dishes coated with fibronectin but attached poorly to surfaces coated with laminin. Both proteins efficiently promoted the attachment of cells from livers undergoing regeneration. After regeneration, the attachment to laminin returned to the low levels found in animals not subjected to partial hepatectomy but attachment to fibronectin remained high. Immunofluorescent staining of sections of normal liver with antilaminin revealed the presence of laminin in or adjacent to the walls of the bile ducts and blood vessels. After induction of regeneration by partial hepatectomy, increased amounts of laminin appeared in the sinusoidal areas. After carbon tetrachloride poisoning, staining for laminin was especially pronounced in the necrotic and postnecrotic areas around the central veins. This additional expression of laminin was transient. It reached a maximum around 5-6 days after the injury and then gradually disappeared. These findings show that laminin is an adhesive protein. The increase of laminin in regenerating liver and the adhesiveness of cells from such livers to laminin suggest a role for laminin in the maintenance of a proper tissue organization during liver regeneration.

Reinjury risk of nano-calcium oxalate monohydrate and calcium oxalate dihydrate crystals on injured renal epithelial cells: aggravation of crystal adhesion and aggregation

PubMed Central

Gan, Qiong-Zhi; Sun, Xin-Yuan; Bhadja, Poonam; Yao, Xiu-Qiong; Ouyang, Jian-Ming

2016-01-01

Background Renal epithelial cell injury facilitates crystal adhesion to cell surface and serves as a key step in renal stone formation. However, the effects of cell injury on the adhesion of nano-calcium oxalate crystals and the nano-crystal-induced reinjury risk of injured cells remain unclear. Methods African green monkey renal epithelial (Vero) cells were injured with H2O2 to establish a cell injury model. Cell viability, superoxide dismutase (SOD) activity, malonaldehyde (MDA) content, propidium iodide staining, hematoxylin–eosin staining, reactive oxygen species production, and mitochondrial membrane potential (Δψm) were determined to examine cell injury during adhesion. Changes in the surface structure of H2O2-injured cells were assessed through atomic force microscopy. The altered expression of hyaluronan during adhesion was examined through laser scanning confocal microscopy. The adhesion of nano-calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) crystals to Vero cells was observed through scanning electron microscopy. Nano-COM and COD binding was quantitatively determined through inductively coupled plasma emission spectrometry. Results The expression of hyaluronan on the cell surface was increased during wound healing because of Vero cell injury. The structure and function of the cell membrane were also altered by cell injury; thus, nano-crystal adhesion occurred. The ability of nano-COM to adhere to the injured Vero cells was higher than that of nano-COD crystals. The cell viability, SOD activity, and Δψm decreased when nano-crystals attached to the cell surface. By contrast, the MDA content, reactive oxygen species production, and cell death rate increased. Conclusion Cell injury contributes to crystal adhesion to Vero cell surface. The attached nano-COM and COD crystals can aggravate Vero cell injury. As a consequence, crystal adhesion and aggregation are enhanced. These findings provide further insights into kidney stone formation. PMID:27382277

Strategies for cell manipulation and skeletal tissue engineering using high-throughput polymer blend formulation and microarray techniques.

PubMed

Khan, Ferdous; Tare, Rahul S; Kanczler, Janos M; Oreffo, Richard O C; Bradley, Mark

2010-03-01

A combination of high-throughput material formulation and microarray techniques were synergistically applied for the efficient analysis of the biological functionality of 135 binary polymer blends. This allowed the identification of cell-compatible biopolymers permissive for human skeletal stem cell growth in both in vitro and in vivo applications. The blended polymeric materials were developed from commercially available, inexpensive and well characterised biodegradable polymers, which on their own lacked both the structural requirements of a scaffold material and, critically, the ability to facilitate cell growth. Blends identified here proved excellent templates for cell attachment, and in addition, a number of blends displayed remarkable bone-like architecture and facilitated bone regeneration by providing 3D biomimetic scaffolds for skeletal cell growth and osteogenic differentiation. This study demonstrates a unique strategy to generate and identify innovative materials with widespread application in cell biology as well as offering a new reparative platform strategy applicable to skeletal tissues. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

Utilization of human DC-SIGN and L-SIGN for entry and infection of host cells by the New World arenavirus, Junín virus

PubMed Central

Belouzard, Sandrine; Cordo, Sandra M.; Candurra, Nélida A.; Whittaker, Gary R.

2014-01-01

The target cell tropism of enveloped viruses is regulated by interactions between viral proteins and cellular receptors determining susceptibility at a host cell, tissue or species level. However, a number of additional cell-surface moieties can also bind viral envelope glycoproteins and could act as capture receptors, serving as attachment factors to concentrate virus particles on the cell surface, or to disseminate the virus infection to target organs or susceptible cells within the host. Here, we used Junín virus (JUNV) or JUNV glycoprotein complex (GPC)-pseudotyped particles to study their ability to be internalized by the human C-type lectins hDC- or hL-SIGN. Our results provide evidence that hDC- and hL-SIGN can mediate the entry of Junín virus into cells, and may play an important role in virus infection and dissemination in the host. PMID:24183720

Implementation and assessment of an early home-based intervention on infant attachment organisation: the CAPEDP attachment study in France.

PubMed

Tereno, Susana; Guedeney, Nicole; Dugravier, Romain; Greacen, Tim; Saïas, Thomas; Tubach, Florence; Guédeney, Antoine

2013-06-01

Attachment is a long-term emotional link between infants and their mothers. Attachment quality influences subsequent psychosocial relationships, the ability to manage stress and, consequently, later mental health. Home intervention programmes targeting infant attachment have been implemented in several contexts with varying degrees of efficacy. Within the CAPEDP study (Parental Skills and Attachment in Early Childhood: reduction of risks linked to mental health problems and promotion of resilience), a subsample of 120 families were recruited with the objective of assessing the impact of this home-visiting programme on infant attachment organisation using the Strange Situation Procedure. The present paper describes the methodology used in this ancillary study.

Identification of Cell Adhesive Sequences in the N-terminal Region of the Laminin α2 Chain*

PubMed Central

Hozumi, Kentaro; Ishikawa, Masaya; Hayashi, Takemitsu; Yamada, Yuji; Katagiri, Fumihiko; Kikkawa, Yamato; Nomizu, Motoyoshi

2012-01-01

The laminin α2 chain is specifically expressed in the basement membrane surrounding muscle and nerve. We screened biologically active sequences in the mouse laminin N-terminal region of α2 chain using 216 soluble peptides and three recombinant proteins (rec-a2LN, rec-a2LN+, and rec-a2N) by both the peptide- or protein-coated plate and the peptide-conjugated Sepharose bead assays. Ten peptides showed cell attachment activity in the plate assay, and 8 peptides were active in the bead assay. Seven peptides were active in the both assays. Five peptides promoted neurite outgrowth with PC12 cells. To clarify the cellular receptors, we examined the effects of heparin and EDTA on cell attachment to 11 active peptides. Heparin inhibited cell attachment to 10 peptides, and EDTA significantly affected only A2-8 peptide (YHYVTITLDLQQ, mouse laminin α2 chain, 117–128)-mediated cell attachment. Cell attachment to A2-8 was also specifically inhibited by anti-integrin β1 and anti-integrin α2β1 antibodies. These results suggest that A2-8 promotes an integrin α2β1-mediated cell attachment. The rec-a2LN protein, containing the A2-8 sequence, bound to integrin α2β1 and cell attachment to rec-a2LN was inhibited by A2-8 peptide. Further, alanine substitution analysis of both the A2-8 peptide and the rec-a2LN+ protein revealed that the amino acids Ile-122, Leu-124, and Asp-125 were involved in integrin α2β1-mediated cell attachment, suggesting that the A2-8 site plays a functional role as an integrin α2β1 binding site in the LN module. These active peptides may provide new insights on the molecular mechanism of laminin-receptor interactions. PMID:22654118

ROCK Inhibition Promotes Attachment, Proliferation, and Wound Closure in Human Embryonic Stem Cell–Derived Retinal Pigmented Epithelium

PubMed Central

Croze, Roxanne H.; Thi, William J.; Clegg, Dennis O.

2016-01-01

Purpose Nonexudative (dry) age-related macular degeneration (AMD), a leading cause of blindness in the elderly, is associated with the loss of retinal pigmented epithelium (RPE) cells and the development of geographic atrophy, which are areas devoid of RPE cells and photoreceptors. One possible treatment option would be to stimulate RPE attachment and proliferation to replace dying/dysfunctional RPE and bring about wound repair. Clinical trials are underway testing injections of RPE cells derived from pluripotent stem cells to determine their safety and efficacy in treating AMD. However, the factors regulating RPE responses to AMD-associated lesions are not well understood. Here, we use cell culture to investigate the role of RhoA coiled coil kinases (ROCKs) in human embryonic stem cell–derived RPE (hESC-RPE) attachment, proliferation, and wound closure. Methods H9 hESC were spontaneously differentiated into RPE cells. hESC-RPE cells were treated with a pan ROCK1/2 or a ROCK2 only inhibitor; attachment, and proliferation and cell size within an in vitro scratch assay were examined. Results Pharmacological inhibition of ROCKs promoted hESC-RPE attachment and proliferation, and increased the rate of closure of in vitro wounds. ROCK inhibition decreased phosphorylation of cofilin and myosin light chain, suggesting that regulation of the cytoskeleton underlies the mechanism of action of ROCK inhibition. Conclusions ROCK inhibition promotes attachment, proliferation, and wound closure in H9 hESC-RPE cells. ROCK isoforms may have different roles in wound healing. Translational Relevance Modulation of the ROCK-cytoskeletal axis has potential in stimulating wound repair in transplanted RPE cells and attachment in cellular therapies. PMID:27917311

The chemotaxis-like Che1 pathway has an indirect role in adhesive cell properties of Azospirillum brasilense.

PubMed

Siuti, Piro; Green, Calvin; Edwards, Amanda Nicole; Doktycz, Mitchel J; Alexandre, Gladys

2011-10-01

The Azospirillum brasilense chemotaxis-like Che1 signal transduction pathway was recently shown to modulate changes in adhesive cell surface properties that, in turn, affect cell-to-cell aggregation and flocculation behaviors rather than flagellar-mediated chemotaxis. Attachment to surfaces and root colonization may be functions related to flocculation. Here, the conditions under which A. brasilense wild-type Sp7 and che1 mutant strains attach to abiotic and biotic surfaces were examined using in vitro attachment and biofilm assays combined with atomic force microscopy and confocal microscopy. The nitrogen source available for growth is found to be a major modulator of surface attachment by A. brasilense and could be promoted in vitro by lectins, suggesting that it depends on interaction with surface-exposed residues within the extracellular matrix of cells. However, Che1-dependent signaling is shown to contribute indirectly to surface attachment, indicating that distinct mechanisms are likely underlying flocculation and attachment to surfaces in A. brasilense. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Structural correlation between lipophilicity and lipopolysaccharide-sequestering activity in spermine-sulfonamide analogs.

PubMed

Burns, Mark R; Jenkins, Scott A; Vermeulen, Nicolas M; Balakrishna, Rajalakshmi; Nguyen, Thuan B; Kimbrell, Matthew R; David, Sunil A

2006-12-15

Lipopolysaccharides (LPS), otherwise termed 'endotoxins', are outer-membrane constituents of Gram-negative bacteria, and play a key role in the pathogenesis of 'Septic Shock', a major cause of mortality in the critically ill patient. We had previously defined the pharmacophore necessary for small molecules to specifically bind and neutralize this complex carbohydrate. A series of aryl and aliphatic spermine-sulfonamide analogs were synthesized and tested in a series of binding and cell-based assays in order to probe the effect of lipophilicity on sequestration ability. A strong correlation was indeed found, supporting the hypothesis that endotoxin-neutralizing ability involves a lipophilic or membrane attachment event. The research discussed herein may be useful for the design of additional carbohydrate recognizing molecules and endotoxin-neutralizing drugs.

The role of nanostructured mesoporous silicon in discriminating in vitro calcification for electrospun composite tissue engineering scaffolds

NASA Astrophysics Data System (ADS)

Fan, Dongmei; Akkaraju, Giridhar R.; Couch, Ernest F.; CanhamCurrent Address, Intrinsiq Materials Ltd, Geraldine Road, Malvern Wr14 3SZ Uk, Leigh T.; Coffer, Jeffery L.

2011-02-01

The impact of mesoporous silicon (PSi) particles-embedded either on the surface, or totally encapsulated within electrospun poly (ε-caprolactone) (PCL) fibers-on its properties as a tissue engineering scaffold is assessed. Our findings suggest that the resorbable porous silicon component can sensitively accelerate the necessary calcification process in such composites. Calcium phosphate deposition on the scaffolds was measured via in vitro calcification assays both at acellular and cellular levels. Extensive attachment of fibroblasts, human adult mesenchymal stem cells, and mouse stromal cells to the scaffold were observed. Complementary cell differentiation assays and ultrastructural measurements were also carried out; the levels of alkaline phosphatase expression, a specific biomarker for mesenchymal stem cell differentiation, show that the scaffolds have the ability to mediate such processes, and that the location of the Si plays a key role in levels of expression.

Selective fibronectin adsorption against albumin and enhanced stem cell attachment on helium atmospheric pressure glow discharge treated titanium

NASA Astrophysics Data System (ADS)

Han, Inho; Vagaska, Barbora; Joo Park, Bong; Lee, Mi Hee; Jin Lee, Seung; Park, Jong-Chul

2011-06-01

Successful tissue integration of implanted medical devices depends on appropriate initial cellular response. In this study, the effect of helium atmospheric pressure glow discharge (He-APGD) treatment of titanium on selective protein adsorption and the initial attachment processes and focal adhesion formation of osteoprogenitor cells and stem cells were examined. Titanium disks were treated in a self-designed He-APGD system. Initial attachment of MC3T3-E1 mouse pre-osteoblasts and human mesenchymal stem cells (MSCs) was evaluated by MTT assay and plasma membrane staining followed by morphometric analysis. Fibronectin adsorption was investigated by Enzyme-Linked ImmunoSorbant Assay. MSCs cell attachment to treated and non-treated titanium disks coated with different proteins was verified also in serum-free culture. Organization of actin cytoskeleton and focal adhesions was evaluated microscopically. He-APGD treatment effectively modified the titanium surfaces by creating a super-hydrophilic surface, which promoted selectively higher adsorption of fibronectin, a protein of critical importance for cell/biomaterial interaction. In two different types of cells, the He-APGD treatment enhanced the number of attaching cells as well as their attachment area. Moreover, cells had higher organization of actin cytoskeleton and focal adhesions. Faster acceptance of the material by the progenitor cells in the early phases of tissue integration after the implantation may significantly reduce the overall healing time; therefore, titanium treatment with He-APGD seems to be an effective method of surface modification of titanium for improving its tissue inductive properties.

[Metapneumovirus expands the understanding of Paramyxovirus cell fusion--a review].

PubMed

Liu, Xiaoyu; Zhang, Xiaodong; Wei, Yongwei

2014-04-04

For most viruses in Paramyxoviridae, cell fusion requires both attachment protein and fusion protein. The attachment protein is responsible for the binding to its cognate receptors, while the interaction between fusion protein and attachment protein triggers the fusion protein which is responsible for the fusion. However, the Metapneumovirus fusion in Pneumovirinae subfamily displayed different mechanism where the attachment protein is not required. The cell fusion is accomplished by fusion protein alone without the help of the attachment protein. Recent studies indicate that low pH is required for cell fusion promoted by some hMPV strains. The fusion protein of aMPV type A is highly fusogenic, whereas that of type B is low. The original fusion models for Paramyxovirus cannot explain the phenomenon above. The mechanism to regulate the cell fusion of Metapneumovirus is poorly understood. It is becoming a hot spot for the study of cell fusion triggered by Paramyxovirus where it enlarged the traditional scope of Paramyxovirus fusion. In this review, we discuss the new achievements and advances in the understanding of cell fusion triggered by Metapneumovirus.

Directional cell elongation through filopodia-steered lamellipodial extension on patterned silk fibroin films.

PubMed

You, Renchuan; Li, Xiufang; Luo, Zuwei; Qu, Jing; Li, Mingzhong

2015-03-05

Micropatterned biomaterials have been used to direct cell alignment for specific tissue engineering applications. However, the understanding of how cells respond to guidance cues remains limited. Plasticity in protrusion formation has been proposed to enable cells to adapt their motility mode to microenvironment. In this study, the authors investigated the key role of protrusion response in cell guidance on patterned silk fibroin films. The results revealed that the ability to transform between filopodia and small lamellipodia played important roles in directional cell guidance. Filopodia did not show directional extension on patterned substrates prior to spreading, but they transduced topographical cues to the cell to trigger the formation of small lamellipodia along the direction of a microgrooved or parallel nanofiber pattern. The polar lamellipodia formation provided not only a path with directionality, but a driving force for directional cell elongation. Moreover, aligned nanofibers coating provided better mechanical support for the traction of filopodia and lamellipodia, promoting cell attachment, spreading, and migration. This study provides new insight into how cells respond to guidance cues and how filopodia and lamellipodia control cell contact guidance on micropatterned biomaterial surfaces.

A proteinaceous fraction of wheat bran may interfere in the attachment of enterotoxigenic E. coli K88 (F4+) to porcine epithelial cells.

PubMed

González-Ortiz, Gemma; Bronsoms, Sílvia; Quarles Van Ufford, H C; Halkes, S Bart A; Virkola, Ritva; Liskamp, Rob M J; Beukelman, Cees J; Pieters, Roland J; Pérez, José Francisco; Martín-Orúe, Susana María

2014-01-01

Wheat bran (WB) from Triticum aestivum has many beneficial effects on human health. To the best of our knowledge, very little has been published about its ability to prevent pathogenic bacterial adhesion in the intestine. Here, a WB extract was fractionated using different strategies, and the obtained fractions were tested in different in vitro methodologies to evaluate their interference in the attachment of enterotoxigenic Escherichia coli (ETEC) K88 to intestinal porcine epithelial cells (IPEC-J2) with the aim of identifying the putative anti-adhesive molecules. It was found that a proteinaceous compound in the >300-kDa fraction mediates the recognition of ETEC K88 to IPEC-J2. Further fractionation of the >300-kDa sample by size-exclusion chromatography showed several proteins below 90 kDa, suggesting that the target protein belongs to a high-molecular-weight (MW) multi-component protein complex. The identification of some relevant excised bands was performed by mass spectrometry (MS) and mostly revealed the presence of various protease inhibitors (PIs) of low MW: Serpin-Z2B, Class II chitinase, endogenous alpha-amylase/subtilisin inhibitor and alpha-amylase/trypsin inhibitor CM3. Furthermore, an incubation of the WB extract with ETEC K88 allowed for the identification of a 7S storage protein globulin of wheat, Globulin 3 of 66 kDa, which may be one of the most firmly attached WB proteins to ETEC K88 cells. Further studies should be performed to gain an understanding of the molecular recognition of the blocking process that takes place. All gathered information can eventually pave the way for the development of novel anti-adhesion therapeutic agents to prevent bacterial pathogenesis.

Interaction of Francisella noatunensis subsp. orientalis with Oreochromis mossambicus bulbus arteriosus cell line

USGS Publications Warehouse

Soto, Esteban; Yun, Susan; Lewis, J.; Kearney, Michael T.; Hansen, John D.

2017-01-01

Francisella noatunensis subsp. orientalis (Fno) (syn. F. asiatica) is an emergent warmwater fish pathogen and the causative agent of piscine francisellosis. Although Fno causes septicemia and can live extracellularly in infected tilapia (Oreochromis spp.), the early interaction of Fno with vasculature endothelium is unknown. In the present study, we examined the interaction of wild-type Fno (WT) and two Fno knockout [intracellular growth loci C (ΔiglC) and pathogenicity determinant protein A (ΔpdpA)] strains with the endothelial O. mossambicus bulbus arteriosus cell line (TmB) at 25 °C and 30 °C. Similar amounts of WT, ΔiglC, and ΔpdpA attached and were detected intracellularly after 5 h of incubation at both temperatures; however temperature affected attachment and uptake. While significantly greater amounts of Fno (WT, ΔiglC, and ΔpdpA) were detected intracellularly when TmB cells were incubated at 30 °C, bacteria attached to TmBs at greater levels at 25 °C. Only WT Fno was able to replicate intracellularly at 25 °C, which resulted in Fno mediated cytotoxicity and apoptosis at 24 and 72 h post-infection. WT Fno incubated at 30 °C as well as ΔiglC,and ΔpdpA incubated at 25 °C and 30 °C were all defective for survival, replication, and the ability to cause cytotoxicity in TmB. Taken together, these results demonstrate that temperature plays a vital role for Fno intracellular survival, persistence and cytotoxicity.

Stability enhancement of an atomic force microscope for long-term force measurement including cantilever modification for whole cell deformation

NASA Astrophysics Data System (ADS)

Weafer, P. P.; McGarry, J. P.; van Es, M. H.; Kilpatrick, J. I.; Ronan, W.; Nolan, D. R.; Jarvis, S. P.

2012-09-01

Atomic force microscopy (AFM) is widely used in the study of both morphology and mechanical properties of living cells under physiologically relevant conditions. However, quantitative experiments on timescales of minutes to hours are generally limited by thermal drift in the instrument, particularly in the vertical (z) direction. In addition, we demonstrate the necessity to remove all air-liquid interfaces within the system for measurements in liquid environments, which may otherwise result in perturbations in the measured deflection. These effects severely limit the use of AFM as a practical tool for the study of long-term cell behavior, where precise knowledge of the tip-sample distance is a crucial requirement. Here we present a readily implementable, cost effective method of minimizing z-drift and liquid instabilities by utilizing active temperature control combined with a customized fluid cell system. Long-term whole cell mechanical measurements were performed using this stabilized AFM by attaching a large sphere to a cantilever in order to approximate a parallel plate system. An extensive examination of the effects of sphere attachment on AFM data is presented. Profiling of cantilever bending during substrate indentation revealed that the optical lever assumption of free ended cantilevering is inappropriate when sphere constraining occurs, which applies an additional torque to the cantilevers "free" end. Here we present the steps required to accurately determine force-indentation measurements for such a scenario. Combining these readily implementable modifications, we demonstrate the ability to investigate long-term whole cell mechanics by performing strain controlled cyclic deformation of single osteoblasts.

Double-chimera proteins to enhance recruitment of endothelial cells and their progenitor cells.

PubMed

Behjati, M; Kazemi, M; Hashemi, M; Zarkesh-Esfahanai, S H; Bahrami, E; Hashemi-Beni, B; Ahmadi, R

2013-08-20

Enhanced attraction of selective vascular reparative cells is of great importance in order to increase vascular patency after endovascular treatments. We aimed to evaluate efficient attachment of endothelial cells and their progenitors on surfaces coated with mixture of specific antibodies, L-selectin and VE-cadherin, with prohibited platelet attachment. The most efficient conditions for coating of L-selectin-Fc chimera and VE-cadherin-Fc chimera proteins were first determined by protein coating on ELISA plates. The whole processes were repeated on titanium substrates, which are commonly used to coat stents. Endothelial progenitor cells (EPCs) and human umbilical vein endothelial cells (HUVECs) were isolated and characterized by flow cytometry. Cell attachment, growth, proliferation, viability and surface cytotoxicity were evaluated using nuclear staining and MTT assay. Platelet and cell attachment were evaluated using scanning electron microscopy. Optimal concentration of each protein for surface coating was 50 ng/ml. The efficacy of protein coating was both heat and pH independent. Calcium ions had significant impact on simultaneous dual-protein coating (P<0.05). Coating stability data revealed more than one year stability for these coated proteins at 4°C. L-selectin and VE-cadherin (ratio of 50:50) coated surface showed highest EPC and HUVEC attachment, viability and proliferation compared to single protein coated and non-coated titanium surfaces (P<0.05). This double coated surface did not show any cytotoxic effect. Surfaces coated with L-selectin and VE-cadherin are friendly surface for EPC and endothelial cell attachment with less platelet attachment. These desirable factors make the L-selectin and VE-cadherin coated surfaces perfect candidate endovascular device. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Leptospira interrogans Binds to Cadherins

PubMed Central

Evangelista, Karen; Franco, Ricardo; Schwab, Andrew; Coburn, Jenifer

2014-01-01

Leptospirosis, caused by pathogenic species of Leptospira, is the most widespread zoonosis and has emerged as a major public health problem worldwide. The adhesion of pathogenic Leptospira to host cells, and to extracellular matrix (ECM) components, is likely to be necessary for the ability of leptospires to penetrate, disseminate and persist in mammalian host tissues. Previous work demonstrated that pathogenic L. interrogans binds to host cells more efficiently than to ECM. Using two independent screening methods, mass spectrometry and protein arrays, members of the cadherin family were identified as potential L. interrogans receptors on mammalian host surfaces. We focused our investigation on vascular endothelial (VE)-cadherin, which is widely expressed on endothelia and is primarily responsible for endothelial cell-cell adhesion. Monolayers of EA.hy926 and HMEC-1 endothelial cells produce VE-cadherin, bind L. interrogans in vitro, and are disrupted upon incubation with the bacteria, which may reflect the endothelial damage seen in vivo. Dose-dependent and saturable binding of L. interrogans to the purified VE-cadherin receptor was demonstrated and pretreatment of purified receptor or endothelial cells with function-blocking antibody against VE-cadherin significantly inhibited bacterial attachment. The contribution of VE-cadherin to leptospiral adherence to host endothelial cell surfaces is biologically significant because VE-cadherin plays an important role in maintaining the barrier properties of the vasculature. Attachment of L. interrogans to the vasculature via VE-cadherin may result in vascular damage, facilitating the escape of the pathogen from the bloodstream into different tissues during disseminated infection, and may contribute to the hemorrhagic manifestations of leptospirosis. This work is first to describe a mammalian cell surface protein as a receptor for L. interrogans. PMID:24498454

Utilization of cellulosic waste from tequila bagasse and production of polyhydroxyalkanoate (PHA) bioplastics by Saccharophagus degradans.

PubMed

Alva Munoz, Luis Esteban; Riley, Mark R

2008-08-01

Utilization of wastes from agriculture is becoming increasingly important due to concerns of environmental impact. The goals of this work were to evaluate the ability of an unusual organism, Saccharophagus degradans (ATCC 43961), to degrade the major components of plant cell walls and to evaluate the ability of S. degradans to produce polyhydroxyalkanoates (PHAs, also known as bioplastics). S. degradans can readily attach to cellulosic fibers, degrade the cellulose, and utilize this as the primary carbon source. The growth of S. degradans was assessed in minimal media (MM) containing glucose, cellobiose, avicel, and bagasse with all able to support growth. Cells were able to attach to avicel and bagasse fibers; however, growth on these insoluble fibers was much slower and led to a lower maximal biomass production than observed with simple sugars. Lignin in MM alone did not support growth, but did support growth upon addition of glucose, although with an increased adaptation phase. When culture conditions were switched to a nitrogen depleted status, PHA production commences and extends for at least 48 h. At early stationary phase, stained inclusion bodies were visible and two chronologically increasing infrared light absorbance peaks at 1,725 and 1,741 cm(-1) confirmed the presence of PHAs. This work demonstrates for what we believe to be the first time, that a single organism can degrade insoluble cellulose and under similar conditions can produce and accumulate PHA. Additional work is necessary to more fully characterize these capabilities and to optimize the PHA production and purification. (c) 2008 Wiley Periodicals, Inc.

The paths leading from attachment to ageism: a structural equation model approach.

PubMed

Bodner, Ehud; Cohen-Fridel, Sara

2014-01-01

The study introduces a model in which attachment patterns serve as predictors, empathy and fear of death as mediators, and ageism as the predicted variable. Data were collected from young adults (N = 440). Anxious attachment was directly and positively correlated with ageism, and also indirectly and positively by the mediator "fear of death." Avoidant attachment was indirectly and negatively correlated with ageism by the mediator "empathy". It is suggested that interventions for reducing ageist attitudes among younger adults would focus on existential fears, as well as on empathic ability, according to the attachment tendencies of these individuals.

Linking ceragenins to water-treatment membranes to minimize biofouling.

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

Hibbs, Michael R.; Altman, Susan Jeanne; Feng, Yanshu

Ceragenins were used to create biofouling resistant water-treatment membranes. Ceragenins are synthetically produced antimicrobial peptide mimics that display broad-spectrum bactericidal activity. While ceragenins have been used on bio-medical devices, use of ceragenins on water-treatment membranes is novel. Biofouling impacts membrane separation processes for many industrial applications such as desalination, waste-water treatment, oil and gas extraction, and power generation. Biofouling results in a loss of permeate flux and increase in energy use. Creation of biofouling resistant membranes will assist in creation of clean water with lower energy usage and energy with lower water usage. Five methods of attaching three different cerageninmore » molecules were conducted and tested. Biofouling reduction was observed in the majority of the tests, indicating the ceragenins are a viable solution to biofouling on water treatment membranes. Silane direct attachment appears to be the most promising attachment method if a high concentration of CSA-121a is used. Additional refinement of the attachment methods are needed in order to achieve our goal of several log-reduction in biofilm cell density without impacting the membrane flux. Concurrently, biofilm forming bacteria were isolated from source waters relevant for water treatment: wastewater, agricultural drainage, river water, seawater, and brackish groundwater. These isolates can be used for future testing of methods to control biofouling. Once isolated, the ability of the isolates to grow biofilms was tested with high-throughput multiwell methods. Based on these tests, the following species were selected for further testing in tube reactors and CDC reactors: Pseudomonas ssp. (wastewater, agricultural drainage, and Colorado River water), Nocardia coeliaca or Rhodococcus spp. (wastewater), Pseudomonas fluorescens and Hydrogenophaga palleronii (agricultural drainage), Sulfitobacter donghicola, Rhodococcus fascians, Rhodobacter katedanii, and Paracoccus marcusii (seawater), and Sphingopyxis spp. (groundwater). The testing demonstrated the ability of these isolates to be used for biofouling control testing under laboratory conditions. Biofilm forming bacteria were obtained from all the source water samples.« less

Boron nitride nanotube-enhanced osteogenic differentiation of mesenchymal stem cells.

PubMed

Li, Xia; Wang, Xiupeng; Jiang, Xiangfen; Yamaguchi, Maho; Ito, Atsuo; Bando, Yoshio; Golberg, Dmitri

2016-02-01

The interaction between boron nitride nanotubes (BNNTs) layer and mesenchymal stem cells (MSCs) is evaluated for the first time in this study. BNNTs layer supports the attachment and growth of MSCs and exhibits good biocompatibility with MSCs. BNNTs show high protein adsorption ability, promote the proliferation of MSCs and increase the secretion of total protein by MSCs. Especially, BNNTs enhance the alkaline phosphatase (ALP) activity as an early marker of osteoblasts, ALP/total protein and osteocalcin (OCN) as a late marker of osteogenic differentiation, which shows that BNNTs can enhance osteogenesis of MSCs. The release of trace boron and the stress on cells exerted by BNNTs with a fiber structure may account for the enhanced differentiation of MSCs into osteoblasts. Therefore BNNTs are potentially useful for bone regeneration in orthopedic applications. © 2015 Wiley Periodicals, Inc.

Effect of acid tolerance response (ATR) on attachment of Listeria monocytogenes Scott A to stainless steel under extended exposure to acid or/and salt stress and resistance of sessile cells to subsequent strong acid challenge.

PubMed

Chorianopoulos, Nikos; Giaouris, Efstathios; Grigoraki, Ioanna; Skandamis, Panagiotis; Nychas, George-John

2011-02-28

The aim of this study was to investigate the potential effect of adaptive stationary phase acid tolerance response (ATR) of Listeria monocytogenes Scott A cells on their attachment to stainless steel (SS) under low pH or/and high salt conditions and on the subsequent resistance of sessile cells to strong acid challenge. Nonadapted or acid-adapted stationary-phase L. monocytogenes cells were used to inoculate (ca. 10⁸ CFU/ml) Brain Heart (BH) broth (pH 7.4, 0.5% w/v NaCl) in test tubes containing vertically placed SS coupons (used as abiotic substrates for bacterial attachment). Incubation was carried out at 16 °C for up to 15 days, without any nutrient refreshment. L. monocytogenes cells, prepared as described above, were also exposed to low pH (4.5; adjusted with HCl) or/and high salt (5.5% w/v NaCl) stresses, during attachment. On the 5th, 10th and 15th day of incubation, cells attached to SS coupons were detached (through bead vortexing) and enumerated (by agar plating). Results revealed that ATR significantly (p<0.05) affected bacterial attachment, when the latter took place under moderate acidic conditions (pH 4.5, 0.5 or 5.5% w/v NaCl), with the acid-adapted cells adhering slightly more than the nonadapted ones. Regardless of acidity/salinity conditions during attachment, ATR also enhanced the resistance of sessile cells to subsequent lethal acid challenge (exposure to pH 2 for 6 min; pH adjusted with either hydrochloric or lactic acid). The trend observed with viable count data agreed well with conductance measurements, used to indirectly quantify remaining attached bacteria (following the strong acid challenge) via their metabolic activity. To sum, this study demonstrates that acid adaptation of L. monocytogenes cells during their planktonic growth enhances their subsequent attachment to SS under extended exposure (at 16 °C for up to 15 days) to mild acidic conditions (pH 4.5), while it also improves the resistance of sessile cells to extreme acid treatment (pH 2). Therefore, the ATR of bacterial cells should be carefully considered when applying acidic decontamination strategies to eradicate L. monocytogenes attached to food processing equipment. Copyright © 2011 Elsevier B.V. All rights reserved.

Controlling attachment and growth of Listeria monocytogenes in polyvinyl chloride model floor drains using a peroxide chemical, chitosan-arginine, or heat.

PubMed

Berrang, Mark E; Hofacre, Charles L; Frank, Joseph F

2014-12-01

Listeria monocytogenes can colonize a poultry processing plant as a resident in floor drains. Limiting growth and attachment to drain surfaces may help lessen the potential for cross-contamination of product. The objective of this study was to compare a hydrogen peroxide-peroxyacetic acid-based chemical to chitosan-arginine or heat to prevent attachment of or destroy existing L. monocytogenes on the inner surface of model floor drains. L. monocytogenes was introduced to result in about 10(9) planktonic and attached cells within untreated polyvinyl chloride model drain pipes. Treatments (0.13 % peroxide-based sanitizer, 0.1 % chitosan-arginine, or 15 s of hot water at 95 to 100°C) were applied immediately after inoculation or after 24 h of incubation. Following treatment, all pipes were incubated for an additional 24 h; planktonic and attached cells were enumerated by plate count. All treatments significantly (P < 0.05) lowered numbers of planktonic and attached cells recovered. Chitosan-arginine resulted in approximately a 6-log reduction in planktonic cells when applied prior to incubation and a 3-log reduction after the inoculum had a chance to grow. Both heat and peroxide significantly outperformed chitosan-arginine (8- to 9-log reduction) and were equally effective before and after incubation. Heat was the only treatment that eliminated planktonic L. monocytogenes. All treatments were less effective against attached cells. Chitosan-arginine provided about a 4.5-log decrease in attached cells when applied before incubation and no significant decrease when applied after growth. Like with planktonic cells, peroxide-peroxyacetic acid and heat were equally effective before or after incubation, causing decreases ranging from 7 to 8.5 log for attached L. monocytogenes. Applied at the most efficacious time, any of these techniques may lessen the potential for L. monocytogenes to remain as a long-term resident in processing plant floor drains.

Analysis of the site-specific integration system of the Streptomyces aureofaciens phage μ1/6.

PubMed

Farkašovská, Jarmila; Godány, Andrej

2012-03-01

The bacteriophage μ1/6 integrates its DNA into the chromosome of tetracycline producing strains of Streptomyces aureofaciens by a site-specific recombination process. A bioinformatic analysis of the μ1/6 genome revealed that orf5 encodes a putative integrase, a basic protein of 416 amino acids. The μ1/6 integrase was found to belong to the integrase family of site-specific tyrosine recombinases. The phage attachment site (attP) was localized downstream of the int gene. The attachment junctions (attL and attR) were determined, allowing identification of the bacterial attachment site (attB). All attachment sites shared a 46-bp common core sequence within which a site-specific recombination occurs. This core sequence comprises the 3' end of a putative tRNA(Thr) gene (anticodon TGT) which is completely restored in attL after integration of the phage into the host genome. An integration vector containing μ1/6 int-attP region was inserted stably into the S. aureofaciens B96, S. lividans TK24, and S. coelicolor A3. The μ1/6 integrase was shown to be functional in vivo in heterologous Escherichia coli without any other factors encoded by Streptomyces. In vitro recombination assay using purified μ1/6 integrase demonstrated its ability to catalyze integrative recombination in the presence of a crude extract of E. coli cells.

Mechanisms of Ricin Toxin Neutralization Revealed through Engineered Homodimeric and Heterodimeric Camelid Antibodies.

PubMed

Herrera, Cristina; Tremblay, Jacqueline M; Shoemaker, Charles B; Mantis, Nicholas J

2015-11-13

Novel antibody constructs consisting of two or more different camelid heavy-chain only antibodies (VHHs) joined via peptide linkers have proven to have potent toxin-neutralizing activity in vivo against Shiga, botulinum, Clostridium difficile, anthrax, and ricin toxins. However, the mechanisms by which these so-called bispecific VHH heterodimers promote toxin neutralization remain poorly understood. In the current study we produced a new collection of ricin-specific VHH heterodimers, as well as VHH homodimers, and characterized them for their ability neutralize ricin in vitro and in vivo. We demonstrate that the VHH heterodimers, but not homodimers were able to completely protect mice against ricin challenge, even though the two classes of antibodies (heterodimers and homodimers) had virtually identical affinities for ricin holotoxin and similar IC50 values in a Vero cell cytotoxicity assay. The VHH heterodimers did differ from the homodimers in their ability to promote toxin aggregation in solution, as revealed through analytical ultracentrifugation. Moreover, the VHH heterodimers that were most effective at promoting ricin aggregation in solution were also the most effective at blocking ricin attachment to cell surfaces. Collectively, these data suggest that heterodimeric VHH-based neutralizing agents may function through the formation of antibody-toxin complexes that are impaired in their ability to access host cell receptors. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

RNA-seq transcriptional profiling of Herbaspirillum seropedicae colonizing wheat (Triticum aestivum) roots.

PubMed

Pankievicz, V C S; Camilios-Neto, D; Bonato, P; Balsanelli, E; Tadra-Sfeir, M Z; Faoro, H; Chubatsu, L S; Donatti, L; Wajnberg, G; Passetti, F; Monteiro, R A; Pedrosa, F O; Souza, E M

2016-04-01

Herbaspirillum seropedicae is a diazotrophic and endophytic bacterium that associates with economically important grasses promoting plant growth and increasing productivity. To identify genes related to bacterial ability to colonize plants, wheat seedlings growing hydroponically in Hoagland's medium were inoculated with H. seropedicae and incubated for 3 days. Total mRNA from the bacteria present in the root surface and in the plant medium were purified, depleted from rRNA and used for RNA-seq profiling. RT-qPCR analyses were conducted to confirm regulation of selected genes. Comparison of RNA profile of root attached and planktonic bacteria revealed extensive metabolic adaptations to the epiphytic life style. These adaptations include expression of specific adhesins and cell wall re-modeling to attach to the root. Additionally, the metabolism was adapted to the microxic environment and nitrogen-fixation genes were expressed. Polyhydroxybutyrate (PHB) synthesis was activated, and PHB granules were stored as observed by microscopy. Genes related to plant growth promotion, such as auxin production were expressed. Many ABC transporter genes were regulated in the bacteria attached to the roots. The results provide new insights into the adaptation of H. seropedicae to the interaction with the plant.

Regulation of outer kinetochore Ndc80 complex-based microtubule attachments by the central kinetochore Mis12/MIND complex

PubMed Central

Kudalkar, Emily M.; Scarborough, Emily A.; Umbreit, Neil T.; Zelter, Alex; Gestaut, Daniel R.; Riffle, Michael; Johnson, Richard S.; MacCoss, Michael J.; Asbury, Charles L.; Davis, Trisha N.

2015-01-01

Multiple protein subcomplexes of the kinetochore cooperate as a cohesive molecular unit that forms load-bearing microtubule attachments that drive mitotic chromosome movements. There is intriguing evidence suggesting that central kinetochore components influence kinetochore–microtubule attachment, but the mechanism remains unclear. Here, we find that the conserved Mis12/MIND (Mtw1, Nsl1, Nnf1, Dsn1) and Ndc80 (Ndc80, Nuf2, Spc24, Spc25) complexes are connected by an extensive network of contacts, each essential for viability in cells, and collectively able to withstand substantial tensile load. Using a single-molecule approach, we demonstrate that an individual MIND complex enhances the microtubule-binding affinity of a single Ndc80 complex by fourfold. MIND itself does not bind microtubules. Instead, MIND binds Ndc80 complex far from the microtubule-binding domain and confers increased microtubule interaction of the complex. In addition, MIND activation is redundant with the effects of a mutation in Ndc80 that might alter its ability to adopt a folded conformation. Together, our results suggest a previously unidentified mechanism for regulating microtubule binding of an outer kinetochore component by a central kinetochore complex. PMID:26430240

VARIATION IN ACCESSIBLE CELL SURFACE IMMUNOGLOBULIN AMONG ANTIBODY-FORMING CELLS

PubMed Central

Nossal, G. J. V.; Lewis, Heather

1972-01-01

Spleen cells from CBA mice that had been primarily or secondarily immunized with sheep red blood cells were reacted at 0°C with a 125I-labeled polyvalent rabbit anti-mouse globulin reagent. After suitable washing, the cells were placed in a plaque-revealing monolayer and warmed to 37°C. Plaques appeared within 10–20 min. Single plaque-forming cells (PFC) were taken from the middle of plaques, were washed by micromanipulation, and were singly dried on glass slides. The amount of attached antireceptor was assessed by quantitative radioautography. Great variation in "receptor density" was encountered among the 258 single cells studied. However, early, immature PFC in both primary and secondary responses had statistically significantly more receptors than late, mature PFC. On any given day point, no difference was found between IgM- and IgG-forming cells. The results were consistent with the view that cells still able to be driven to further proliferation by antigen retain receptors, and conversely that cells, as they mature, lose both receptors and ability to be influenced by antigen. PMID:4554455

A novel bioactive membrane by cell electrospinning.

PubMed

Chen, Haiping; Liu, Yuanyuan; Hu, Qingxi

2015-11-01

Electrospinning permits fabrication of biodegradable matrices that can resemble the both scale and mechanical behavior of the native extracellular matrix. However, achieving high-cellular density and infiltration of cells within matrices with traditional technique remain challenging and time consuming. The cell electrospinning technique presented in this paper can mitigate the problems associated with these limitations. Cells encapsulated by the material in the cell electrospinning technique survived well and distributed homogenously within the nanofibrous membrane, and their vitality was improved to 133% after being cultured for 28 days. The electrospun nanofibrous membrane has a certain degradation property and favorable cell-membrane interaction that supports the active biocompatibility of the membrane. Its properties are helpful for supporting cell attachment and growth, maintaining phenotypic shape, and secreting an ample amount of extracellular matrix (ECM). This novel membrane may be a potential application within the field of tissue engineering. The ability of cell electrospinning to microintegrate cells into a biodegradable fibrous matrix embodies a novel tissue engineering approach that could be applied to fabricate a high cell density elastic tissue mimetic. Copyright © 2015 Elsevier Inc. All rights reserved.

KSHV cell attachment sites revealed by ultra sensitive tyramide signal amplification (TSA) localize to membrane microdomains that are up-regulated on mitotic cells.

PubMed

Garrigues, H Jacques; Rubinchikova, Yelena E; Rose, Timothy M

2014-03-01

Cell surface structures initiating attachment of Kaposi's sarcoma-associated herpesvirus (KSHV) were characterized using purified hapten-labeled virions visualized by confocal microscopy with a sensitive fluorescent enhancement using tyramide signal amplification (TSA). KSHV attachment sites were present in specific cellular domains, including actin-based filopodia, lamellipodia, ruffled membranes, microvilli and intercellular junctions. Isolated microdomains were identified on the dorsal surface, which were heterogeneous in size with a variable distribution that depended on cellular confluence and cell cycle stage. KSHV binding domains ranged from scarce on interphase cells to dense and continuous on mitotic cells, and quantitation of bound virus revealed a significant increase on mitotic compared to interphase cells. KSHV also bound to a supranuclear domain that was distinct from microdomains in confluent and interphase cells. These results suggest that rearrangement of the cellular membrane during mitosis induces changes in cell surface receptors implicated in the initial attachment stage of KSHV entry. Copyright © 2014 Elsevier Inc. All rights reserved.

Human Macrophages and Dendritic Cells Can Equally Present MART-1 Antigen to CD8+ T Cells after Phagocytosis of Gamma-Irradiated Melanoma Cells

PubMed Central

Barrio, María Marcela; Abes, Riad; Colombo, Marina; Pizzurro, Gabriela; Boix, Charlotte; Roberti, María Paula; Gélizé, Emmanuelle; Rodriguez-Zubieta, Mariana

2012-01-01

Dendritic cells (DC) can achieve cross-presentation of naturally-occurring tumor-associated antigens after phagocytosis and processing of dying tumor cells. They have been used in different clinical settings to vaccinate cancer patients. We have previously used gamma-irradiated MART-1 expressing melanoma cells as a source of antigens to vaccinate melanoma patients by injecting irradiated cells with BCG and GM-CSF or to load immature DC and use them as a vaccine. Other clinical trials have used IFN-gamma activated macrophage killer cells (MAK) to treat cancer patients. However, the clinical use of MAK has been based on their direct tumoricidal activity rather than on their ability to act as antigen-presenting cells to stimulate an adaptive antitumor response. Thus, in the present work, we compared the fate of MART-1 after phagocytosis of gamma-irradiated cells by clinical grade DC or MAK as well as the ability of these cells to cross present MART-1 to CD8+ T cells. Using a high affinity antibody against MART-1, 2A9, which specifically stains melanoma tumors, melanoma cell lines and normal melanocytes, the expression level of MART-1 in melanoma cell lines could be related to their ability to stimulate IFN-gamma production by a MART-1 specific HLA-A*0201-restricted CD8+ T cell clone. Confocal microscopy with Alexa Fluor®647-labelled 2A9 also showed that MART-1 could be detected in tumor cells attached and/or fused to phagocytes and even inside these cells as early as 1 h and up to 24 h or 48 h after initiation of co-cultures between gamma-irradiated melanoma cells and MAK or DC, respectively. Interestingly, MART-1 was cross-presented to MART-1 specific T cells by both MAK and DC co-cultured with melanoma gamma-irradiated cells for different time-points. Thus, naturally occurring MART-1 melanoma antigen can be taken-up from dying melanoma cells into DC or MAK and both cell types can induce specific CD8+ T cell cross-presentation thereafter. PMID:22768350

Protection and attachment of Vibrio cholerae mediated by the toxin-coregulated pilus in the infant mouse model.

PubMed

Krebs, Shelly J; Taylor, Ronald K

2011-10-01

Colonization of the human small intestine by Vibrio cholerae is an essential step in pathogenesis that requires the type IV toxin-coregulated pilus (TCP). To date, three functions of TCP have been characterized: it serves as the CTXΦ receptor, secretes the colonization factor TcpF, and functions in microcolony formation by mediating bacterium-bacterium interactions. Although type IV pili in other pathogenic bacteria have been characterized as playing a major role in attachment to epithelial cells, there are very few studies to suggest that TCP acts as an attachment factor. Taking this into consideration, we investigated the function of TCP in attachment to Caco-2 cells and found that mutants lacking TCP were defective in attachment compared to the wild type. Overexpression of ToxT, the activator of TCP, significantly increased attachment of wild-type V. cholerae to Caco-2 cells. Using field-emission scanning electron microscopy (FESEM), we also observed TCP-mediated attachment to the small intestines of infected infant mice by using antibodies specific to TCP and V. cholerae. Remarkably, we also visualized matrices comprised of TCP appearing to engulf V. cholerae during infection, and we demonstrated that these matrices protected the bacteria from a component of bile, disclosing a possible new role of this pilus in protection of the bacterial cells from antimicrobial agents. This study provides new insights into TCP's function in V. cholerae colonization of the small intestine, describing additional roles in mediating attachment and protection of V. cholerae bacterial cells.

Attachment within life history theory: an evolutionary perspective on individual differences in attachment.

PubMed

Szepsenwol, Ohad; Simpson, Jeffry A

2018-03-15

In this article, we discuss theory and research on how individual differences in adult attachment mediate the adaptive calibration of reproductive strategies, cognitive schemas, and emotional expression and regulation. We first present an integration of attachment theory and life history theory. Then, we discuss how early harsh and/or unpredictable environments may promote insecure attachment by hampering parents' ability to provide sensitive and reliable care to their children. Finally, we discuss how, in the context of harsh and/or unpredictable environments, different types of insecure attachment (i.e. anxiety and avoidance) may promote evolutionary adaptive reproductive strategies, cognitive schemas, and emotional expression and regulation profiles. Copyright © 2018 Elsevier Ltd. All rights reserved.

Morphological Heterogeneity and Attachment of Phaeobacter inhibens.

PubMed

Segev, Einat; Tellez, Adèle; Vlamakis, Hera; Kolter, Roberto

2015-01-01

The Roseobacter clade is a key group of bacteria in the ocean exhibiting diverse metabolic repertoires and a wide range of symbiotic life-styles. Many Roseobacters possess remarkable capabilities of attachment to both biotic and abiotic surfaces. When attached to each other, these bacteria form multi-cellular structures called rosettes. Phaeobacter inhibens, a well-studied Roseobacter, exhibits various cell sizes and morphologies that are either associated with rosettes or occur as single cells. Here we describe the distribution of P. inhibens morphologies and rosettes within a population. We detect an N-acetylglucosamine-containing polysaccharide on the poles of some cells and at the center of all rosettes. We demonstrate that rosettes are formed by the attachment of individual cells at the polysaccharide-containing pole rather than by cell division. Finally, we show that P. inhibens attachment to abiotic surfaces is hindered by the presence of DNA from itself, but not from other bacteria. Taken together, our findings demonstrate that cell adhesiveness is likely to play a significant role in the life cycle of P. inhibens as well as other Roseobacters.

Morphological Heterogeneity and Attachment of Phaeobacter inhibens

PubMed Central

Segev, Einat; Tellez, Adèle; Vlamakis, Hera; Kolter, Roberto

2015-01-01

The Roseobacter clade is a key group of bacteria in the ocean exhibiting diverse metabolic repertoires and a wide range of symbiotic life-styles. Many Roseobacters possess remarkable capabilities of attachment to both biotic and abiotic surfaces. When attached to each other, these bacteria form multi-cellular structures called rosettes. Phaeobacter inhibens, a well-studied Roseobacter, exhibits various cell sizes and morphologies that are either associated with rosettes or occur as single cells. Here we describe the distribution of P. inhibens morphologies and rosettes within a population. We detect an N-acetylglucosamine-containing polysaccharide on the poles of some cells and at the center of all rosettes. We demonstrate that rosettes are formed by the attachment of individual cells at the polysaccharide-containing pole rather than by cell division. Finally, we show that P. inhibens attachment to abiotic surfaces is hindered by the presence of DNA from itself, but not from other bacteria. Taken together, our findings demonstrate that cell adhesiveness is likely to play a significant role in the life cycle of P. inhibens as well as other Roseobacters. PMID:26560130

Stable kinetochore-microtubule attachment is sufficient to silence the spindle assembly checkpoint in human cells.

PubMed

Tauchman, Eric C; Boehm, Frederick J; DeLuca, Jennifer G

2015-12-01

During mitosis, duplicated sister chromatids attach to microtubules emanating from opposing sides of the bipolar spindle through large protein complexes called kinetochores. In the absence of stable kinetochore-microtubule attachments, a cell surveillance mechanism known as the spindle assembly checkpoint (SAC) produces an inhibitory signal that prevents anaphase onset. Precisely how the inhibitory SAC signal is extinguished in response to microtubule attachment remains unresolved. To address this, we induced formation of hyper-stable kinetochore-microtubule attachments in human cells using a non-phosphorylatable version of the protein Hec1, a core component of the attachment machinery. We find that stable attachments are sufficient to silence the SAC in the absence of sister kinetochore bi-orientation and strikingly in the absence of detectable microtubule pulling forces or tension. Furthermore, we find that SAC satisfaction occurs despite the absence of large changes in intra-kinetochore distance, suggesting that substantial kinetochore stretching is not required for quenching the SAC signal.

Surface contact stimulates the just-in-time deployment of bacterial adhesins.

PubMed

Li, Guanglai; Brown, Pamela J B; Tang, Jay X; Xu, Jing; Quardokus, Ellen M; Fuqua, Clay; Brun, Yves V

2012-01-01

The attachment of bacteria to surfaces provides advantages such as increasing nutrient access and resistance to environmental stress. Attachment begins with a reversible phase, often mediated by surface structures such as flagella and pili, followed by a transition to irreversible attachment, typically mediated by polysaccharides. Here we show that the interplay between pili and flagellum rotation stimulates the rapid transition between reversible and polysaccharide-mediated irreversible attachment. We found that reversible attachment of Caulobacter crescentus cells is mediated by motile cells bearing pili and that their contact with a surface results in the rapid pili-dependent arrest of flagellum rotation and concurrent stimulation of polar holdfast adhesive polysaccharide. Similar stimulation of polar adhesin production by surface contact occurs in Asticcacaulis biprosthecum and Agrobacterium tumefaciens. Therefore, single bacterial cells respond to their initial contact with surfaces by triggering just-in-time adhesin production. This mechanism restricts stable attachment to intimate surface interactions, thereby maximizing surface attachment, discouraging non-productive self-adherence, and preventing curing of the adhesive. © 2011 Blackwell Publishing Ltd.

Stable kinetochore–microtubule attachment is sufficient to silence the spindle assembly checkpoint in human cells

PubMed Central

Tauchman, Eric C.; Boehm, Frederick J.; DeLuca, Jennifer G.

2015-01-01

During mitosis, duplicated sister chromatids attach to microtubules emanating from opposing sides of the bipolar spindle through large protein complexes called kinetochores. In the absence of stable kinetochore–microtubule attachments, a cell surveillance mechanism known as the spindle assembly checkpoint (SAC) produces an inhibitory signal that prevents anaphase onset. Precisely how the inhibitory SAC signal is extinguished in response to microtubule attachment remains unresolved. To address this, we induced formation of hyper-stable kinetochore–microtubule attachments in human cells using a non-phosphorylatable version of the protein Hec1, a core component of the attachment machinery. We find that stable attachments are sufficient to silence the SAC in the absence of sister kinetochore bi-orientation and strikingly in the absence of detectable microtubule pulling forces or tension. Furthermore, we find that SAC satisfaction occurs despite the absence of large changes in intra-kinetochore distance, suggesting that substantial kinetochore stretching is not required for quenching the SAC signal. PMID:26620470

Adherence of carp leucocytes to adults and cercariae of the blood fluke Sanguinicola inermis.

PubMed

Richards, D T; Hoole, D; Lewis, J W; Ewens, E; Arme, C

1996-03-01

Live adult and cercarial stages of Sanguinicola inermis Plehn, 1905 (Trematoda:Sanguinicolidae) were maintained in vitro in the presence of carp (Cyprinus carpio L.) leucocytes. Cells and parasites were fixed at intervals from 0.25 to 48 h and examined using light microscopy, SEM and TEM. Within 12 h of exposure, leucocytes were found attached to cercariae although, by 24 h, fewer cells were found attached to postcercarial, juvenile adult stages that had shed their tails. Neutrophils and macrophages were found attached to the damaged tegument of cercarie that had not transformed by 48 h. Few cells were attached to the tegument of adult flukes that were alive when fixed. However, there was extensive tegumental damage and numerous cells were attached to adult flukes that had died before fixation. The results are discussed with reference to parasite survival within the vascular system of the host.

Diesel oil removal by immobilized Pseudoxanthomonas sp. RN402.

PubMed

Nopcharoenkul, Wannarak; Netsakulnee, Parichat; Pinyakong, Onruthai

2013-06-01

Pseudoxanthomonas sp. RN402 was capable of degrading diesel, crude oil, n-tetradecane and n-hexadecane. The RN402 cells were immobilized on the surface of high-density polyethylene plastic pellets at a maximum cell density of 10(8) most probable number (MPN) g(-1) of plastic pellets. The immobilized cells not only showed a higher efficacy of diesel oil removal than free cells but could also degrade higher concentrations of diesel oil. The rate of diesel oil removal by immobilized RN402 cells in liquid culture was 1,050 mg l(-1) day(-1). Moreover, the immobilized cells could maintain high efficacy and viability throughout 70 cycles of bioremedial treatment of diesel-contaminated water. The stability of diesel oil degradation in the immobilized cells resulted from the ability of living RN402 cells to attach to material surfaces by biofilm formation, as was shown by CLSM imaging. These characteristics of the immobilized RN402 cells, including high degradative efficacy, stability and flotation, make them suitable for the purpose of continuous wastewater bioremediation.

Passive and active response of bacteria under mechanical compression

NASA Astrophysics Data System (ADS)

Garces, Renata; Miller, Samantha; Schmidt, Christoph F.; Byophysics Team; Institute of Medical Sciences Collaboration

Bacteria display simple but fascinating cellular structures and geometries. Their shapes are the result of the interplay between osmotic pressure and cell wall construction. Typically, bacteria maintain a high difference of osmotic pressure (on the order of 1 atm) to the environment. This pressure difference (turgor pressure) is supported by the cell envelope, a composite of lipid membranes and a rigid cell wall. The response of the cell envelope to mechanical perturbations such as geometrical confinements is important for the cells survival. Another key property of bacteria is the ability to regulate turgor pressure after abrupt changes of external osmotic conditions. This response relies on the activity of mechanosensitive (MS) channels: membrane proteins that release solutes in response to excessive stress in the cell envelope. We here present experimental data on the mechanical response of the cell envelope and on turgor regulation of bacteria subjected to compressive forces. We indent living cells with micron-sized beads attached to the cantilever of an atomic force microscope (AFM). This approach ensures global deformation of the cell. We show that such mechanical loading is sufficient to gate mechanosensitive channels in isosmotic conditions.

Impact of pre-incubation time of silk fibroin scaffolds in culture medium on cell proliferation and attachment.

PubMed

Amirikia, Mehdi; Shariatzadeh, Seyed Mohammad Ali; Jorsaraei, Seyed Gholam Ali; Soleimani Mehranjani, Malek

2017-12-01

Cell behaviours such as proliferation and attachment can be affected by the length of pre-incubation period of the scaffolds in the culture medium for long term. The aim of this study was to investigate the long term pre-incubation of 3D silk fibroin scaffolds in complete culture medium on cell attachment and proliferation. After the preparation of silk fibroin scaffolds by the technique of freeze drying, the scaffolds were pre-incubated in complete culture medium for 2 d, 6 d or 10 d before apical papilla stem cells (SCAP) seeding. Modifications of the scaffold surface and wettability were examined by FE-SEM and water contact angle, respectively. Results showed a decrease both in roughness and water contact angle as pre-incubation time increases. DNA measurement after 18h and 10 d cell seeding showed a significant increase of DNA concentration which represents better attachment and proliferation with pre-incubation time increase. Qualitative examination, live&dead assay or H&E staining method after 30h and 10 d cell seeding respectively, indicated that pre-incubation of scaffolds has time dependent effect on cell proliferation and attachment. This suggests that improvement of cell attachment and proliferation may be mediated by differences in the amount of wettability (decreased water contact angle) after exposure of scaffold to culture medium for long term which, in turn, causes more protein adsorption in the surface of silk fibroin scaffold (decreased roughness). Copyright © 2017. Published by Elsevier Ltd.

Dimorphism in methane seep-dwelling ecotypes of the largest known bacteria

PubMed Central

Bailey, Jake V; Salman, Verena; Rouse, Gregory W; Schulz-Vogt, Heide N; Levin, Lisa A; Orphan, Victoria J

2011-01-01

We present evidence for a dimorphic life cycle in the vacuolate sulfide-oxidizing bacteria that appears to involve the attachment of a spherical Thiomargarita-like cell to the exteriors of invertebrate integuments and other benthic substrates at methane seeps. The attached cell elongates to produce a stalk-like form before budding off spherical daughter cells resembling free-living Thiomargarita that are abundant in surrounding sulfidic seep sediments. The relationship between the attached parent cell and free-living daughter cell is reminiscent of the dimorphic life modes of the prosthecate Alphaproteobacteria, but on a grand scale, with individual elongate cells reaching nearly a millimeter in length. Abundant growth of attached Thiomargarita-like bacteria on the integuments of gastropods and other seep fauna provides not only a novel ecological niche for these giant bacteria, but also for animals that may benefit from epibiont colonization. PMID:21697959

Expression of adhesion and extracellular matrix genes in human blastocysts upon attachment in a 2D co-culture system.

PubMed

Aberkane, A; Essahib, W; Spits, C; De Paepe, C; Sermon, K; Adriaenssens, T; Mackens, S; Tournaye, H; Brosens, J J; Van de, Velde H

2018-05-26

What are the changes in human embryos, in terms of morphology and gene expression, upon attachment to endometrial epithelial cells? Apposition and adhesion of human blastocysts to endometrial epithelial cells are predominantly initiated at the embryonic pole and these steps are associated with changes in expression of adhesion and extracellular matrix (ECM) genes in the embryo. Both human and murine embryos have been co-cultured with Ishikawa cells, although embryonic gene expression associated with attachment has not yet been investigated in an in-vitro implantation model. Vitrified human blastocysts were warmed and co-cultured for up to 48 h with Ishikawa cells, a model cell line for receptive endometrial epithelium. Six-days post fertilisation (6dpf) human embryos were co-cultured with Ishikawa cells for 12 h, 24 h (7dpf) or 48 h (8dpf) and attachment rate and morphological development investigated. Expression of 84 adhesion and ECM genes was analysed by quantitative PCR. Immunofluorescence microscopy was used to assess the expression of three informative genes at the protein level. Data are reported on 115 human embryos. Mann-Whitney U was used for statistical analysis between two groups, with P < 0.05 considered significant. The majority of embryos attached to Ishikawa cells at the level of the polar trophectoderm; 41% of co-cultured embryos were loosely attached after 12 h and 86% firmly attached after 24 h. Outgrowth of hCG-positive embryonic cells at 8dpf indicated differentiation of trophectoderm into invasive syncytiotrophoblast. Gene expression analysis was performed on loosely attached and unattached embryos co-cultured with Ishikawa cells for 12 h. In contrast to unattached embryos, loosely attached embryos expressed THBS1, TNC, COL12A1, CTNND2, ITGA3, ITGAV, and LAMA3 and had significantly higher CD44 and TIMP1 transcript levels (P = 0.014 and P = 0.029, respectively). LAMA3, THBS1 and TNC expression was validated at the protein level in firmly attached 7dpf embryos. Thrombospondin 1 (THBS1) resided in the cytoplasm of embryonic cells whereas laminin subunit alpha 3 (LAMA3) and tenascin C (TNC) were expressed on the cell surface of trophectoderm cells. Incubation with a neutralizing TNC antibody did not affect the rate of embryo attachment or hCG secretion. None. This in-vitro study made use of an endometrial adenocarcinoma cell line to mimic receptive luminal epithelium. Also, the number of embryos was limited. Contamination of recovered embryos with Ishikawa cells was unlikely based on their differential gene expression profiles. Taken together, we provide a 'proof of concept' that initiation of the implantation process coincides with the induction of specific embryonic genes. Genome-wide expression profiling of a larger sample set may provide insights into the molecular embryonic pathways underlying successful or failed implantation. A.A. was supported by a grant from the "Instituut voor Innovatie door Wetenschap en Technologie" (IWT, 121716, Flanders, Belgium). This work was supported by the "Wetenschappelijk Fonds Willy Gepts" (WFWG G142 and G170, Universitair Ziekenhuis Brussel). The authors declare no conflict of interest.

The effect of nicotine and cotinine on human gingival fibroblasts attachment to root surfaces.

PubMed

Esfahrood, Zeinab Rezaei; Zamanian, Amirhosein; Torshabi, Maryam; Abrishami, Maryam

2015-09-01

Different compounds of smoking (e.g., nicotine and cotinine) are risk factors for various diseases such as oral cancer and periodontal diseases. Some studies reported the negative effects of nicotine on cell proliferation and differentiation. The present in vitro study assessed the effects of nicotine and cotinine (long-acting metabolite of nicotine) on the attachment and viability of human gingival fibroblast (HGF) cells to tooth root surfaces. A total of 70 teeth specimens were placed into 48-well culture plates and covered with HGF cell suspension, in complete Dulbecco's modified Eagle's medium culture medium containing 1 nM, 1 μm, 1 mM, and 5 mM of nicotine and cotinine concentrations. Cellular attachment and viability measured using an MTT assay and a scanning electron microscope were used for cell morphological evaluation. After 24 h, low (nanomolar and micromolar) and high concentrations (millimolar) of nicotine and cotinine caused a significant reduction in the initial cell adhesion in comparison with the control group, but no significant difference was observed between the nicotine and the cotinine groups (p<0.05). Dentally attached cells with low concentrations of nicotine and cotinine proliferated 48 h after exposure, the same as the control group. However, dentally attached cells with high concentrations of nicotine and cotinine (especially 5 mM) did not proliferate 24 h after exposure (p<0.05). Low concentrations of nicotine and cotinine caused a reduction in the initial cell adhesion. However, no significant adverse effects on the proliferation of attached cells were seen in the longer period. High concentrations of nicotine and cotinine have adverse effects on the cell adhesion and proliferation of HGF cells.

Surface Oxide Net Charge of a Titanium Alloy ; Modulation of Fibronectin-Activated Attachment and Spreading of Osteogenic Cells

PubMed Central

Rapuano, Bruce E.; MacDonald, Daniel E.

2010-01-01

In the current study, we have altered the surface oxide properties of a Ti6Al4V alloy using heat treatment or radiofrequency glow discharge (RFGD) in order to evaluate the relationship between the physico-chemical and biological properties of the alloy's surface oxide. The effects of surface pretreatments on the attachment of cells from two osteogenic cell lines (MG63 and MC3T3) and a mesenchymal stem cell line (C3H10T1/2) to fibronectin adsorbed to the alloy were measured. Both heat and RFGD pretreatments produced a several-fold increase in the number of cells that attached to fibronectin adsorbed to the alloy (0.001 and 10 nM FN) for each cell line tested. An antibody (HFN7.1) directed against the central integrin binding domain of fibronectin produced a 65-70% inhibition of cell attachment to fibronectin-coated disks, incdicating that cell attachment to the metal discs was dependent on fibronectin binding to cell integrin receptors. Both treatments also accelerated the cell spreading response manifested by extensive flattening and an increase in mean cellular area. The treatment-induced increases in the cell attachment activity of adsorbed fibronectin were correlated with previously demonstrated increases in Ti6Al4V oxide negative net surface charge at physiological pH produced by both heat and RFGD pretreatments. Since neither treatment increased the adsorption mass of fibronectin, these findings suggest that negatively charged surface oxide functional groups in Ti6Al4V can modulate fibronectin's integrin receptor activity by altering the adsorbed protein's conformation. Our results further suggest that negatively charged functional groups in the surface oxide can play a prominent role in the osseointegration of metallic implant materials. PMID:20884181

A nerve guidance conduit with topographical and biochemical cues: potential application using human neural stem cells

NASA Astrophysics Data System (ADS)

Jenkins, Phillip M.; Laughter, Melissa R.; Lee, David J.; Lee, Young M.; Freed, Curt R.; Park, Daewon

2015-06-01

Despite major advances in the pathophysiological understanding of peripheral nerve damage, the treatment of nerve injuries still remains an unmet medical need. Nerve guidance conduits present a promising treatment option by providing a growth-permissive environment that 1) promotes neuronal cell survival and axon growth and 2) directs axonal extension. To this end, we designed an electrospun nerve guidance conduit using a blend of polyurea and poly-caprolactone with both biochemical and topographical cues. Biochemical cues were integrated into the conduit by functionalizing the polyurea with RGD to improve cell attachment. Topographical cues that resemble natural nerve tissue were incorporated by introducing intraluminal microchannels aligned with nanofibers. We determined that electrospinning the polymer solution across a two electrode system with dissolvable sucrose fibers produced a polymer conduit with the appropriate biomimetic properties. Human neural stem cells were cultured on the conduit to evaluate its ability to promote neuronal growth and axonal extension. The nerve guidance conduit was shown to enhance cell survival, migration, and guide neurite extension.

Cellulose promotes extracellular assembly of Clostridium cellulovorans cellulosomes.

PubMed Central

Matano, Y; Park, J S; Goldstein, M A; Doi, R H

1994-01-01

Cellulosome synthesis by Clostridium cellulovorans was investigated by growing the cells in media containing different carbon sources. Supernatant from cells grown with cellobiose contained no cellulosomes and only the free forms of cellulosomal major subunits CbpA, P100, and P70 and the minor subunits with enzymatic activity. Supernatant from cells grown on pebble-milled cellulose and Avicel contained cellulosomes capable of degrading crystalline cellulose. Supernatants from cells grown with cellobiose, pebble-milled cellulose, and Avicel contained about the same amount of carboxymethyl cellulase activity. Although the supernatant from the medium containing cellobiose did not initially contain active cellulosomes, the addition of crystalline cellulose to the cell-free supernatant fraction converted the free major forms to cellulosomes with the ability to degrade crystalline cellulose. The binding of P100 and P70 to crystalline cellulose was dependent on their attachment to the endoglucanase-binding domains of CbpA. These data strongly indicate that crystalline cellulose promotes cellulosome assembly. Images PMID:7961457

What bridges the gap between self-harm and suicidality? The role of forgiveness, resilience and attachment.

PubMed

Nagra, Gurmokh S; Lin, Ashleigh; Upthegrove, Rachel

2016-07-30

Self-harm is the most robust risk for completed suicide. There is a lack of understanding of why some people who self-harm escalate to suicidal behaviour when others do not. Psychological factors such as attachment, self-forgiveness and self-appraisal may be important. To determine whether factors from the Interpersonal Theory and Schematic Appraisals models are useful to identify suicidal behaviour in populations that self-harm. Specifically we investigate whether resilience factors of secure attachment, self-forgiveness and positive self-appraisals significantly influence suicidality in people who self-harm. A cross-sectional online study of 323 participants recruited from self-harm support forum. Validated self-report measures were used to assess appraisals, relationships, self-forgiveness, attachment style, suicidality and self-harm. Emotion coping and support seeking self-appraisals and self-forgiveness were negatively associated with suicidality in participants with a history of self-harm. Dismissing attachment was positively associated with suicidality. The perceived ability to cope with emotions, the perceived ability to gain support and self-forgiveness may protect against suicide in people who self-harm. Conversely the presence of dismissing attachment may increase the risk of suicidality. Findings provide therapeutic targets to reduce risk of suicidality in this high risk group. Copyright © 2016. Published by Elsevier Ireland Ltd.

Sirt1 carboxyl-domain is an ATP-repressible domain that is transferrable to other proteins

PubMed Central

Kang, Hyeog; Oka, Shinichi; Lee, Duck-Yeon; Park, Junhong; Aponte, Angel M.; Jung, Young-Sang; Bitterman, Jacob; Zhai, Peiyong; He, Yi; Kooshapur, Hamed; Ghirlando, Rodolfo; Tjandra, Nico; Lee, Sean B.; Kim, Myung K.; Sadoshima, Junichi; Chung, Jay H.

2017-01-01

Sirt1 is an NAD+-dependent protein deacetylase that regulates many physiological functions, including stress resistance, adipogenesis, cell senescence and energy production. Sirt1 can be activated by energy deprivation, but the mechanism is poorly understood. Here, we report that Sirt1 is negatively regulated by ATP, which binds to the C-terminal domain (CTD) of Sirt1. ATP suppresses Sirt1 activity by impairing the CTD's ability to bind to the deacetylase domain as well as its ability to function as the substrate recruitment site. ATP, but not NAD+, causes a conformational shift to a less compact structure. Mutations that prevent ATP binding increase Sirt1's ability to promote stress resistance and inhibit adipogenesis under high-ATP conditions. Interestingly, the CTD can be attached to other proteins, thereby converting them into energy-regulated proteins. These discoveries provide insight into how extreme energy deprivation can impact Sirt1 activity and underscore the complex nature of Sirt1 structure and regulation. PMID:28504272

Analysis of the synaptotagmin family during reconstituted membrane fusion. Uncovering a class of inhibitory isoforms.

PubMed

Bhalla, Akhil; Chicka, Michael C; Chapman, Edwin R

2008-08-01

Ca(2+)-triggered exocytosis in neurons and neuroendocrine cells is regulated by the Ca(2+)-binding protein synaptotagmin (syt) I. Sixteen additional isoforms of syt have been identified, but little is known concerning their biochemical or functional properties. Here, we assessed the abilities of fourteen syt isoforms to directly regulate SNARE (soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptor)-catalyzed membrane fusion. One group of isoforms stimulated neuronal SNARE-mediated fusion in response to Ca(2+), while another set inhibited SNARE catalyzed fusion in both the absence and presence of Ca(2+). Biochemical analysis revealed a strong correlation between the ability of syt isoforms to bind 1,2-dioleoyl phosphatidylserine (PS) and t-SNAREs in a Ca(2+)-promoted manner with their abilities to enhance fusion, further establishing PS and SNAREs as critical effectors for syt action. The ability of syt I to efficiently stimulate fusion was specific for certain SNARE pairs, suggesting that syts might contribute to the specificity of intracellular membrane fusion reactions. Finally, a subset of inhibitory syts down-regulated the ability of syt I to activate fusion, demonstrating that syt isoforms can modulate the function of each other.

Optimization and inhibition of the adherent ability of Plasmodium falciparum-infected erythrocytes.

PubMed

Smith, H; Crandall, I; Prudhomme, J; Sherman, I W

1992-01-01

The vast majority of the 1-2 million malaria associated deaths that occur each year are due to anemia and cerebral malaria (the attachment of erythrocytes containing mature forms of Plasmodium falciparum to the endothelial cells that line the vascular beds of the brain). A "model system" for the study of cerebral malaria employs amelanotic melanoma cells as the "target" cells in an in vitro cytoadherence assay. Using this model system we determined that the optimum pH for adherence is 6.6 to 6.8, that high concentrations of Ca2+ (50mM) result in increased levels of binding, and that the type of buffer used influences adherence (Bis Tris > MOPS > HEPES > PIPES). We also observed that the ability of infected erythrocytes to cytoadhere varied from (erythrocyte) donor to donor. We have produced murine monoclonal antibodies against P. falciparum-infected red cells which recognize modified forms of human band 3; these inhibit the adherence of infected erythrocytes to melanoma cells in a dose-responsive fashion. Antimalarials (chloroquine, quinacrine, mefloquine, artemisinin), on the other hand, affected adherence in an indirect fashion i.e. since cytoadherence is due, in part, to the presence of knobs on the surface of the infected erythrocyte, and knob formation is dependent on intracellular parasite growth, when plasmodial development is inhibited so is knob production, and consequently adherence is ablated.

In vitro testing of calcium phosphate (HA, TCP, and biphasic HA-TCP) whiskers.

PubMed

Jalota, Sahil; Bhaduri, Sarit B; Tas, A Cuneyt

2006-09-01

Calcium phosphate [single-phase hydroxyapatite (HA, Ca(10)(PO(4))(6)(OH)(2)), single-phase tricalcium phosphate (beta-TCP, Ca(3)(PO(4))(2)), and biphasic HA-TCP] whiskers were formed by using a novel microwave-assisted molten salt mediated process. Aqueous solutions containing NaNO(3), HNO(3), Ca(NO(3))(2) x 4H(2)O, and KH(2)PO(4) (with or without urea) were used as starting reagents. These solutions were irradiated in a household microwave oven for 5 min. As-recovered precursors were then simply stirred in water at room temperature for 1 h to obtain the whiskers of the desired calcium phosphate (CaP) bioceramics. These whiskers were evaluated, respectively, in vitro by (1) soaking those in synthetic body fluid (SBF) solutions at 37 degrees C for one week, and (2) performing cell attachment and total protein assay tests on the neat whiskers by using a mouse osteoblast cell line (7F2). beta-TCP, HA, and HA-TCP biphasic whiskers were all found to possess apatite-inducing ability when soaked in SBF. SBF-soaked whiskers were found to have BET surface areas ranging from 45 to 112 m(2)/g. Although the osteoblast viability and protein concentrations were found to be the highest on the neat HA whiskers, cells were attached and proliferated on all the whiskers.

Attachment style moderates theory of mind abilities in depression.

PubMed

Koelkebeck, Katja; Liedtke, Carla; Kohl, Waldemar; Alferink, Judith; Kret, Mariska Esther

2017-04-15

Being able to understand other people's emotions and intentions is crucial for social interactions and well-being. Deficits in theory of mind (ToM) functioning hamper this ability and have been observed in depression and other neuropsychiatric disorders. However, results of previous research in depression have been inconclusive, possibly due to the presence of comorbid disorders and the disregarding of other modulating factors. Thirty-eight patients with a major depressive disorder (MDD) and forty healthy matched controls were assessed with a ToM task using animated triangles. Results were correlated with attachment styles, empathy abilities and neurocognitive performance. Our findings show that 1) healthy female controls performed significantly stronger on the ToM task than female MDD patients, 2) these performance differences were driven by attachment styles and 3) depression severity did not impact task performance. The pharmacological treatment of the majority of patients might limit the generalizability of this study. Results indicate a gender-specific impact of attachment styles on ToM performance. Future studies should investigate whether impairments in social cognitive tasks pose a risk factor for depression and/or interactional styles or vice versa. Moreover, with regard to remediation programs gender-specific needs should be taken into account. Copyright © 2017 Elsevier B.V. All rights reserved.

Minimum line width of ion beam-modified polystyrene by negative carbon ions for nerve-cell attachment and neurite extension

NASA Astrophysics Data System (ADS)

Sommani, P.; Tsuji, H.; Sato, H.; Kitamura, T.; Hattori, M.; Gotoh, Y.; Ishikawa, J.

2007-04-01

The minimum line width of the negative-ion-modified polystyrene (PS) for guidance and immobilizations of nerve-cell body and neurite extension have been investigated. Carbon negative ions were implanted into PS at fluence of 3 × 1015 ions/cm2 and energy of 5-20 keV through the various triangle apertures of the micro-pattern mask. After in vitro culture of the nerve-like cells of rat adrenal pheochromocytoma (PC12h), results showed that the minimum line widths for a single cell attachment and for neurite extension were 5-7 and 3-5 μm, respectively. While the minimum line width for attachment of cell group with long neurite was about 20 μm. The suitable widths for a large number of cells and for neurite extension were 20 and 5 μm, respectively. Therefore, the guidance for a clear separation of the attachment size of cell body and neurite extension could be achieved by the different modified line widths.

Fluorescence lifetime microscopy for monitoring cell adhesion using metal induced energy transfer

NASA Astrophysics Data System (ADS)

Hwang, Wonsang; Seo, JinWon; Song, Jun ho; Kim, DongEun; Won, YoungJae; Choi, In-Hong; Yoo, Kyung-Hwa; Kim, Dug Young

2018-02-01

A precise control and a reliable monitoring tool for the adhesion properties of a cell are very important in atherosclerosis studies. If endothelial cells in contact with the intracellular membrane are not attached securely, low-density lipoprotein (LDL) particles can enter into the inner membrane. It is therefore necessary to measure conditions under which endothelial cell detachment occurs. When a cell is attached to a metal thin film, the lifetime of a fluorescence probe attached to the membrane of the cell is reduced by the metal-induced energy transfer (MIET). Fluorescence lifetime imaging microscopy (FLIM) is used to monitor the attachment condition of a cell to a metal surface using FRET. However, this requires high numerical aperture (NA) objective lens because axial confocal resolution must be smaller than the cell thickness. This requirement limits the field of view of the measurement specimen. In this study we provides a new method which can measure adhesion properties of endothelial cells even with a low NA objective lens by resolving two lifetime components in FLIM.

Reduction of Fe(III) colloids by Shewanella putrefaciens: A kinetic model

NASA Astrophysics Data System (ADS)

Bonneville, Steeve; Behrends, Thilo; van Cappellen, Philippe; Hyacinthe, Christelle; Röling, Wilfred F. M.

2006-12-01

A kinetic model for the microbial reduction of Fe(III) oxyhydroxide colloids in the presence of excess electron donor is presented. The model assumes a two-step mechanism: (1) attachment of Fe(III) colloids to the cell surface and (2) reduction of Fe(III) centers at the surface of attached colloids. The validity of the model is tested using Shewanella putrefaciens and nanohematite as model dissimilatory iron reducing bacteria and Fe(III) colloidal particles, respectively. Attachment of nanohematite to the bacteria is formally described by a Langmuir isotherm. Initial iron reduction rates are shown to correlate linearly with the relative coverage of the cell surface by nanohematite particles, hence supporting a direct electron transfer from membrane-bound reductases to mineral particles attached to the cells. Using internally consistent parameter values for the maximum attachment capacity of Fe(III) colloids to the cells, Mmax, the attachment constant, KP, and the first-order Fe(III) reduction rate constant, k, the model reproduces the initial reduction rates of a variety of fine-grained Fe(III) oxyhydroxides by S. putrefaciens. The model explains the observed dependency of the apparent Fe(III) half-saturation constant, Km∗, on the solid to cell ratio, and it predicts that initial iron reduction rates exhibit saturation with respect to both the cell density and the abundance of the Fe(III) oxyhydroxide substrate.

A pre-therapeutic coating for medical devices that prevents the attachment of Candida albicans.

PubMed

Vargas-Blanco, Diego; Lynn, Aung; Rosch, Jonah; Noreldin, Rony; Salerni, Anthony; Lambert, Christopher; Rao, Reeta P

2017-05-19

Hospital acquired fungal infections are defined as "never events"-medical errors that should never have happened. Systemic Candida albicans infections results in 30-50% mortality rates. Typically, adhesion to abiotic medical devices and implants initiates such infections. Efficient adhesion initiates formation of aggressive biofilms that are difficult to treat. Therefore, inhibitors of adhesion are important for drug development and likely to have a broad spectrum efficacy against many fungal pathogens. In this study we further the development of a small molecule, Filastatin, capable of preventing C. albicans adhesion. We explored the potential of Filastatin as a pre-therapeutic coating of a diverse range of biomaterials. Filastatin was applied on various biomaterials, specifically bioactive glass (cochlear implants, subcutaneous drug delivery devices and prosthetics); silicone (catheters and other implanted devices) and dental resin (dentures and dental implants). Adhesion to biomaterials was evaluated by direct visualization of wild type C. albicans or a non-adherent mutant edt1 -/- that were stained or fluorescently tagged. Strains grown overnight at 30 °C were harvested, allowed to attach to surfaces for 4 h and washed prior to visualization. The adhesion force of C. albicans cells attached to surfaces treated with Filastatin was measured using Atomic Force Microscopy. Effectiveness of Filastatin was also demonstrated under dynamic conditions using a flow cell bioreactor. The effect of Filastatin under microfluidic flow conditions was quantified using electrochemical impedance spectroscopy. Experiments were typically performed in triplicate. Treatment with Filastatin significantly inhibited the ability of C. albicans to adhere to bioactive glass (by 99.06%), silicone (by 77.27%), and dental resin (by 60.43%). Atomic force microcopy indicated that treatment with Filastatin decreased the adhesion force of C. albicans from 0.23 to 0.017 nN. Electrochemical Impedance Spectroscopy in a microfluidic device that mimic physiological flow conditions in vivo showed lower impedance for C. albicans when treated with Filastatin as compared to untreated control cells, suggesting decreased attachment. The anti-adhesive properties were maintained when Filastatin was included in the preparation of silicone materials. We demonstrate that Filastatin treated medical devices prevented adhesion of Candida, thereby reducing nosocomial infections.

Lack of electricity production by Pelobacter carbinolicus indicates that the capacity for Fe(III) oxide reduction does not necessarily confer electron transfer ability to fuel cell anodes.

PubMed

Richter, Hanno; Lanthier, Martin; Nevin, Kelly P; Lovley, Derek R

2007-08-01

The ability of Pelobacter carbinolicus to oxidize electron donors with electron transfer to the anodes of microbial fuel cells was evaluated because microorganisms closely related to Pelobacter species are generally abundant on the anodes of microbial fuel cells harvesting electricity from aquatic sediments. P. carbinolicus could not produce current in a microbial fuel cell with electron donors which support Fe(III) oxide reduction by this organism. Current was produced using a coculture of P. carbinolicus and Geobacter sulfurreducens with ethanol as the fuel. Ethanol consumption was associated with the transitory accumulation of acetate and hydrogen. G. sulfurreducens alone could not metabolize ethanol, suggesting that P. carbinolicus grew in the fuel cell by converting ethanol to hydrogen and acetate, which G. sulfurreducens oxidized with electron transfer to the anode. Up to 83% of the electrons available in ethanol were recovered as electricity and in the metabolic intermediate acetate. Hydrogen consumption by G. sulfurreducens was important for ethanol metabolism by P. carbinolicus. Confocal microscopy and analysis of 16S rRNA genes revealed that half of the cells growing on the anode surface were P. carbinolicus, but there was a nearly equal number of planktonic cells of P. carbinolicus. In contrast, G. sulfurreducens was primarily attached to the anode. P. carbinolicus represents the first Fe(III) oxide-reducing microorganism found to be unable to produce current in a microbial fuel cell, providing the first suggestion that the mechanisms for extracellular electron transfer to Fe(III) oxides and fuel cell anodes may be different.

Effects of wound dressings on cultured primary keratinocytes.

PubMed

Esteban-Vives, Roger; Young, Matthew T; Ziembicki, Jenny; Corcos, Alain; Gerlach, Jörg C

2016-02-01

Autologous cell-spray grafting of non-cultured epidermal cells is an innovative approach for the treatment of severe second-degree burns. After treatment, wounds are covered with dressings that are widely used in wound care management; however, little is known about the effects of wound dressings on individually isolated cells. The sprayed cells have to actively attach, spread, proliferate, and migrate in the wound for successful re-epithelialization, during the healing process. It is expected that exposure to wound dressing material might interfere with cell survival, attachment, and expansion. Two experiments were performed to determine whether some dressing materials have a negative impact during the early phases of wound healing. In one experiment, freshly isolated cells were seeded and cultured for one week in combination with eight different wound dressings used during burn care. Cells, which were seeded and cultured with samples of Adaptic(®), Xeroform(®), EZ Derm(®), and Mepilex(®) did not attach, nor did they survive during the first week. Mepitel(®), N-Terface(®), Polyskin(®), and Biobrane(®) dressing samples had no negative effect on cell attachment and cell growth when compared to the controls. In a second experiment, the same dressings were exposed to pre-cultured cells in order to exclude the effects of attachment and spreading. The results confirm the above findings. This study could be of interest for establishing skin cell grafting therapies in burn medicine and also for wound care in general. Copyright © 2015 Elsevier Ltd and ISBI. All rights reserved.

The role of particle-to-cell interactions in dictating nanoparticle aided magnetophoretic separation of microalgal cells.

PubMed

Toh, Pey Yi; Ng, Bee Wah; Ahmad, Abdul Latif; Chieh, Derek Chan Juinn; Lim, JitKang

2014-11-07

Successful application of a magnetophoretic separation technique for harvesting biological cells often relies on the need to tag the cells with magnetic nanoparticles. This study investigates the underlying principle behind the attachment of iron oxide nanoparticles (IONPs) onto microalgal cells, Chlorella sp. and Nannochloropsis sp., in both freshwater and seawater, by taking into account the contributions of various colloidal forces involved. The complex interplay between van der Waals (vdW), electrostatic (ES) and Lewis acid-base interactions (AB) in dictating IONP attachment was studied under the framework of extended Derjaguin-Landau-Verwey-Overbeek (XDLVO) analysis. Our results showed that ES interaction plays an important role in determining the net interaction between the Chlorella sp. cells and IONPs in freshwater, while the AB and vdW interactions play a more dominant role in dictating the net particle-to-cell interaction in high ionic strength media (≥100 mM NaCl), such as seawater. XDLVO predicted effective attachment between cells and surface functionalized IONPs (SF-IONPs) with an estimated secondary minimum of -3.12 kT in freshwater. This prediction is in accordance with the experimental observation in which 98.89% of cells can be magnetophoretically separated from freshwater with SF-IONPs. We have observed successful magnetophoretic separation of microalgal cells from freshwater and/or seawater for all the cases as long as XDLVO analysis predicts particle attachment. For both the conditions, no pH adjustment is required for particle-to-cell attachment.

Mechanics of membrane-cytoskeleton attachment in Paramecium

NASA Astrophysics Data System (ADS)

Campillo, C.; Jerber, J.; Fisch, C.; Simoes-Betbeder, M.; Dupuis-Williams, P.; Nassoy, P.; Sykes, C.

2012-12-01

In this paper we assess the role of the protein MKS1 (Meckel syndrome type 1) in the cortical membrane mechanics of the ciliated protist Paramecium. This protein is known to be crucial in the process of cilium formation, and we investigate its putative role in membrane-cytoskeleton attachment. Therefore, we compare cells where the gene coding for MKS1 is silenced to wild-type cells. We found that scanning electron microscopy observation of the cell surface reveals a cup-like structure in wild-type cells that is lost in silenced cells. Since this structure is based on the underlying cytoskeleton, one hypothesis to explain this observation is a disruption of membrane attachment to the cytoskeleton in the absence of MKS1 that should affect plasma membrane mechanics. We test this by probing the mechanics of wild-type and silenced cells by micropipette aspiration. Strikingly, we observe that, at the same aspiration pressure, the membrane of silenced cells is easily aspirated by the micropipette whereas that of wild-type cells enters only at a moderate velocity, an effect that suggests a detachment of the membrane from the underlying cytoskeleton in silenced cells. We quantify this detachment by measuring the deformation of the cell cortex and the rate of cell membrane entry in the micropipette. This study offers a new perspective for the characterization of membrane-cytoskeleton attachment in protists and paves the way for a better understanding of the role of membrane-cortex attachment in cilium formation.

Escherichia coli strains expressing H12 antigens demonstrate an increased ability to attach to abiotic surfaces as compared with E. coli strains expressing H7 antigens.

PubMed

Goulter, Rebecca M; Taran, Elena; Gentle, Ian R; Gobius, Kari S; Dykes, Gary A

2014-07-01

The role of Escherichia coli H antigens in hydrophobicity and attachment to glass, Teflon and stainless steel (SS) surfaces was investigated through construction of fliC knockout mutants in E. coli O157:H7, O1:H7 and O157:H12. Loss of FliC(H12) in E. coli O157:H12 decreased attachment to glass, Teflon and stainless steel surfaces (p<0.05). Complementing E. coli O157:H12 ΔfliC(H12) with cloned wildtype (wt) fliC(H12) restored attachment to wt levels. The loss of FliCH7 in E. coli O157:H7 and O1:H7 did not always alter attachment (p>0.05), but complementation with cloned fliC(H12), as opposed to cloned fliCH7, significantly increased attachment for both strains compared with wt counterparts (p<0.05). Hydrophobicity determined using bacterial adherence to hydrocarbons and contact angle measurements differed with fliC expression but was not correlated to the attachment to materials included in this study. Purified FliC was used to functionalise silicone nitride atomic force microscopy probes, which were used to measure adhesion forces between FliC and substrates. Although no significant difference in adhesion force was observed between FliC(H12) and FliCH7 probes, differences in force curves suggest different mechanism of attachment for FliC(H12) compared with FliCH7. These results indicate that E. coli strains expressing flagellar H12 antigens have an increased ability to attach to certain abiotic surfaces compared with E. coli strains expressing H7 antigens. Copyright © 2014 Elsevier B.V. All rights reserved.

Serratia marcescens Suppresses Host Cellular Immunity via the Production of an Adhesion-inhibitory Factor against Immunosurveillance Cells*

PubMed Central

Ishii, Kenichi; Adachi, Tatsuo; Hamamoto, Hiroshi; Sekimizu, Kazuhisa

2014-01-01

Injection of a culture supernatant of Serratia marcescens into the bloodstream of the silkworm Bombyx mori increased the number of freely circulating immunosurveillance cells (hemocytes). Using a bioassay with live silkworms, serralysin metalloprotease was purified from the culture supernatant and identified as the factor responsible for this activity. Serralysin inhibited the in vitro attachment of both silkworm hemocytes and murine peritoneal macrophages. Incubation of silkworm hemocytes or murine macrophages with serralysin resulted in degradation of the cellular immune factor BmSPH-1 or calreticulin, respectively. Furthermore, serralysin suppressed in vitro phagocytosis of bacteria by hemocytes and in vivo bacterial clearance in silkworms. Disruption of the ser gene in S. marcescens attenuated its host killing ability in silkworms and mice. These findings suggest that serralysin metalloprotease secreted by S. marcescens suppresses cellular immunity by decreasing the adhesive properties of immunosurveillance cells, thereby contributing to bacterial pathogenesis. PMID:24398686

Serratia marcescens suppresses host cellular immunity via the production of an adhesion-inhibitory factor against immunosurveillance cells.

PubMed

Ishii, Kenichi; Adachi, Tatsuo; Hamamoto, Hiroshi; Sekimizu, Kazuhisa

2014-02-28

Injection of a culture supernatant of Serratia marcescens into the bloodstream of the silkworm Bombyx mori increased the number of freely circulating immunosurveillance cells (hemocytes). Using a bioassay with live silkworms, serralysin metalloprotease was purified from the culture supernatant and identified as the factor responsible for this activity. Serralysin inhibited the in vitro attachment of both silkworm hemocytes and murine peritoneal macrophages. Incubation of silkworm hemocytes or murine macrophages with serralysin resulted in degradation of the cellular immune factor BmSPH-1 or calreticulin, respectively. Furthermore, serralysin suppressed in vitro phagocytosis of bacteria by hemocytes and in vivo bacterial clearance in silkworms. Disruption of the ser gene in S. marcescens attenuated its host killing ability in silkworms and mice. These findings suggest that serralysin metalloprotease secreted by S. marcescens suppresses cellular immunity by decreasing the adhesive properties of immunosurveillance cells, thereby contributing to bacterial pathogenesis.

Comparative study on morphologic changes and cell attachment of periodontitis-affected root surfaces following conditioning with CO2 and Er:YAG laser irradiations.

PubMed

Belal, Mahmoud Helmy; Watanabe, Hisashi

2014-10-01

Clinical application of lasers in periodontal therapy has continued to expand in last decades; however there are still some controversies. The present study aimed to compare the conditioning effects of the carbon dioxide (CO2) or erbium-doped: yttrium, aluminum and garnet (Er:YAG) laser on periodontally diseased root surfaces following scaling and root planing (SRP) in terms of the alteration of morphologies as well as the attachment of periodontal ligament cells. Forty-five periodontally affected root specimens were prepared and randomly assigned into three groups: I control (untreated diseased), II. SRP+CO2 laser (pulsed, noncontact mode), and III. SRP+Er:YAG laser (slight contact mode). After treatment, five specimens in each group were used for surface topographic examination. The remaining 10 specimens in each group were incubated with human periodontal ligament cell suspension. All the specimens were finally evaluated by scanning electron microscopy. The control specimens showed the lowest number of cultured cells, mostly in oval shape, with no tightly attached cells. The CO2 lased specimens showed a significant increase in the number of attached cells compared with controls, but demonstrated some major thermal alterations on the surfaces. The Er:YAG lased specimens showed the significantly highest number of attached cells, mostly in flat form, and did not show distinct thermal damage. The present study suggests that compared with the CO2 laser, the Er:YAG laser may constitute a more useful conditioning tool for enhancing periodontal cell attachment to periodontally diseased root surfaces, with fewer undesirable thermal side effects.

Cancer antibody enhanced real time imaging cell probes--a novel theranostic tool using polymer linked carbon nanotubes and quantum dots.

PubMed

Brakmane, Gerda; Madani, Seyed Yazdan; Seifalian, Alexander

2013-06-01

Cancer is a potentially fatal diagnosis, but due to modern medicine there is a potential cure in many of these cases. The rate of treatment success depends on early disease detection and timely, effective delivery of tumour specific treatment. There are many ongoing researches aimed to improve diagnostics or treatment, but the option to use both modalities concomitantly is deficient. In this project we are using the advances in nanotechnology to develop new theranostic tool using single walled carbon nanotubes (SWCNT) and Quantum dots (QDs) for early cancer cell detection, and option to deliver targeted treatment. SWCNTs were refluxed in HNO3/H2SO4 (1:3) at 120ºC for 120 minutes. Functionalised SWCNT was then covalently attached to octa-ammonium polyhedral oligomeric silsesquioxane (POSS), QDs and conjugated with antibodies for targeted cell detection. Fourier transforms infrared spectroscopy (FTIR), Transmission electron microscopy (TEM), UV/NIR analysis, Raman and UV-VIS spectroscopy were used in order to prove the successful conjugation. Toxicology study using alamar blue analysis and DNA assay was conducted in order to choose the best concentration of SWCNT, octa-ammonium-POSS and QDs before commencing the conjugation process. Human colorectal cancer cell line HT29, pancreas cancer cell line PANC-1 and mouse fibroblasts 3T3 were then treated with or without antibody conjugated SWCNT-POSS-QDs (CPQ) compound solution. The cell response was observed under the microscope after 24, 48 and 72 hours. FTIR and Raman spectroscopies confirmed covalent binding of the SWCNTs to Octa-Ammonium-POSS. This was supported by TEM images and photos obtained, which showed well dispersed SWCNTs following its treatment with Octa-Ammonium-POSS compared to pristine SWCNT samples. UV-VIS graphs determined the presence of antibody within the compound. UV/NIR demonstrated QD fluorescence even after attachment of SWCNT-POSS. The cellular behaviour revealed high CPQ-antibody complex affinity towards cancer cells when compared to healthy cell line which internalised the complex only on day three. The pancreas cancer cell line had appearance of lysed pulp after 72 hours of incubation. Colonic cancer cells seemed to regain ability to populate from day three signifying that higher treatment payload is necessary. We have successfully manufactured novel compound consisting of Octa-Ammonium-POSS linked SWCNTs, QDs, and tumour specific antibodies. The complex has proven its potential as cell probing tool, and the attachment of antibodies has shown high affinity to cancer cells rendering this an attractive model for further theranostic developments.

Use of the Adult Attachment Projective Picture System in the formulation of a case of an adolescent refugee with PTSD.

PubMed

Gander, Manuela; Diamond, Diana; Buchheim, Anna; Sevecke, Kathrin

2018-03-16

Forced displacements and their psychosocial consequences in adolescent refugees and their families have received increasing attention in recent years. Although supportive family relations play a key role in buffering the impact of traumatization in adolescents, parental ability to provide such is often subject to extreme pressure. Under conditions of forced dislocation and fear, maladaptive interpersonal strategies in the parent-child relationships may develop, contributing to the onset of psychopathology. We explore new aspects of attachment-related issues for the understanding and treatment of adolescent refugees who have experienced multiple traumas in their childhood. We used a multimethod assessment battery including the Adult Attachment Projective Picture System (AAP), the Structured Clinical Interview, the Youth Self Report and the Wechsler Intelligence Scale in an adolescent boy with post-traumatic stress disorder (PTSD). Our subject was an adolescent refugee from the Middle East who demonstrated an unresolved attachment when confronted with loss and fear. His responses on the AAP evoked aspects of insecure-unresolved attachment, including his belief that it is not safe to trust in attachment figures, his limited access to traumatic attachment experiences, his impaired ability to take concrete actions when dealing with threatening attachment situations and the unintentional role-reversal shed new light on our understanding of his traumatic experiences, family functioning and psychopathological symptoms. Our results demonstrate the utility of the AAP in an adolescent refugee with PTSD by expanding our knowledge of a diverse range of experiences across the interpersonal, cognitive, cultural and developmental contexts that formed the basis for an individualized treatment plan.

Enhanced in vitro biocompatibility of ultrafine-grained biomedical NiTi alloy with microporous surface

NASA Astrophysics Data System (ADS)

Zheng, C. Y.; Nie, F. L.; Zheng, Y. F.; Cheng, Y.; Wei, S. C.; Valiev, R. Z.

2011-08-01

Bulk ultrafine-grained Ni 50.8Ti 49.2 alloy (UFG-NiTi) was successfully fabricated by equal-channel angular pressing (ECAP) technique in the present study, and to further improve its surface biocompatibility, surface modification techniques including sandblasting, acid etching and alkali treatment were employed to produce either irregularly roughened surface or microporous surface or hierarchical porous surface with bioactivity. The effect of the above surface treatments on the surface roughness, wettability, corrosion behavior, ion release, apatite forming ability and cytocompatibility of UFG-NiTi alloy were systematically investigated with the coarse-grained NiTi alloy as control. The pitting corrosion potential ( Epit) was increased from 393 mV (SCE) to 704 mV (SCE) with sandblasting and further increased to 1539 mV (SCE) with following acid etching in HF/HNO 3 solution. All the above surface treatment increased the apatite forming ability of UFG-NiTi in varying degrees when soaked them in simulated body fluid (SBF). Meanwhile, both sandblasting and acid etching could promote the cytocompatibility for osteoblasts: sandblasting enhanced cell attachment and acid etching increased cell proliferation. The different corrosion behavior, apatite forming ability and cellular response of UFG-NiTi after different surface modifications are attributed to the topography and wettability of the resulting surface oxide layer.

In vitro bone formation using muscle-derived cells: a new paradigm for bone tissue engineering using polymer-bone morphogenetic protein matrices.

PubMed

Lu, Helen H; Kofron, Michelle D; El-Amin, Saadiq F; Attawia, Mohammed A; Laurencin, Cato T

2003-06-13

Over 800,000 bone grafting procedures are performed in the United States annually, creating a demand for viable alternatives to autogenous bone, the grafting standard in osseous repair. The objective of this study was to examine the efficacy of a BMP-polymer matrix in inducing the expression of the osteoblastic phenotype and in vitro bone formation by muscle-derived cells. Specifically, we evaluated the ability of bone morphogenetic protein-7 (BMP-7), delivered from a poly(lactide-co-glycolide) (PLAGA) matrix, to induce the differentiation of cells derived from rabbit skeletal muscle into osteoblast-like cells and subsequently form mineralized tissue. Results confirmed that muscle-derived cells attached and proliferated on the PLAGA substrates. BMP-7 released from PLAGA induced the muscle-derived cells to increase bone marker expression and form mineralized cultures. These results demonstrate the efficacy of a BMP-polymer matrix in inducing the expression of the osteoblastic phenotype by muscle-derived cells and present a new paradigm for bone tissue engineering.

Alginate-Encapsulation for the Improved Hypothermic Preservation of Human Adipose-Derived Stem Cells

PubMed Central

Swioklo, Stephen; Constantinescu, Andrei

2016-01-01

Despite considerable progress within the cell therapy industry, unmet bioprocessing and logistical challenges associated with the storage and distribution of cells between sites of manufacture and the clinic exist. We examined whether hypothermic (4°C–23°C) preservation of human adipose-derived stem cells could be improved through their encapsulation in 1.2% calcium alginate. Alginate encapsulation improved the recovery of viable cells after 72 hours of storage. Viable cell recovery was highly temperature-dependent, with an optimum temperature of 15°C. At this temperature, alginate encapsulation preserved the ability for recovered cells to attach to tissue culture plastic on rewarming, further increasing its effect on total cell recovery. On attachment, the cells were phenotypically normal, displayed normal growth kinetics, and maintained their capacity for trilineage differentiation. The number of cells encapsulated (up to 2 × 106 cells per milliliter) did not affect viable cell recovery nor did storage of encapsulated cells in a xeno-free, serum-free,current Good Manufacturing Practice-grade medium. We present a simple, low-cost system capable of enhancing the preservation of human adipose-derived stem cells stored at hypothermic temperatures, while maintaining their normal function. The storage of cells in this manner has great potential for extending the time windows for quality assurance and efficacy testing, distribution between the sites of manufacture and the clinic, and reducing the wastage associated with the limited shelf life of cells stored in their liquid state. Significance Despite considerable advancement in the clinical application of cell-based therapies, major logistical challenges exist throughout the cell therapy supply chain associated with the storage and distribution of cells between the sites of manufacture and the clinic. A simple, low-cost system capable of preserving the viability and functionality of human adipose-derived stem cells (a cell with substantial clinical interest) at hypothermic temperatures (0°C–32°C) is presented. Such a system has considerable potential for extending the shelf life of cell therapy products at multiple stages throughout the cell therapy supply chain. PMID:26826163

Simultaneous dynamic optical and electrical properties of endothelial cell attachment on indium tin oxide bioelectrodes.

PubMed

Choi, Chang K; English, Anthony E; Kihm, Kenneth D; Margraves, Charles H

2007-01-01

This study quantifies the dynamic attachment and spreading of porcine pulmonary artery endothelial cells (PPAECs) on optically thin, indium tin oxide (ITO) biosensors using simultaneous differential interference contrast microscopy (DICM) and electrical microimpedance spectroscopy. A lock-in amplifier circuit monitored the impedance of PPAECs cultivated on the transparent ITO bioelectrodes as a function of frequency between 10 Hz and 100 kHz and as a function of time, while DICM images were simultaneously acquired. A digital image processing algorithm quantified the cell-covered electrode area as a function of time. The results of this study show that the fraction of the cell-covered electrode area is in qualitative agreement with the electrical impedance during the attachment phase following the cell settling on the electrode surface. The possibility of several distinctly different states of electrode coverage and cellular attachment giving rise to similar impedance signals is discussed.

Sphene ceramics for orthopedic coating applications: an in vitro and in vivo study.

PubMed

Ramaswamy, Yogambha; Wu, Chengtie; Dunstan, Colin R; Hewson, Benjamin; Eindorf, Tanja; Anderson, Gail I; Zreiqat, Hala

2009-10-01

The host response to titanium alloy (Ti-6Al-4V) is not always favorable as a fibrous layer may form at the skeletal tissue-device interface, causing aseptic loosening. Recently, sphene (CaTiSiO(5)) ceramics were developed by incorporating Ti in the Ca-Si system, and found to exhibit improved chemical stability. The aim of this study is to evaluate the in vitro response of human osteoblast-like cells, human osteoclasts and human microvascular endothelial cells to sphene ceramics and determine whether coating Ti-6Al-4V implants with sphene enhances anchorage to surrounding bone. The study showed that sphene ceramics support human osteoblast-like cell attachment with organized cytoskeleton structure and express increased mRNA levels of osteoblast-related genes. Sphene ceramics were able to induce the differentiation of monocytes to form functional osteoclasts with the characteristic features of f-actin and alpha(v)beta(3) integrin, and express osteoclast-related genes. Human endothelial cells were also able to attach and express the endothelial cell markers ZO-1 and VE-Cadherin when cultured on sphene ceramics. Histological staining, enzyme histochemistry and immunolabelling were used for identification of mineralized bone and bone remodelling around the coated implants. Ti-6Al-4V implants coated with sphene showed new bone formation and filled the gap between the implants and existing bone in a manner comparable to that of the hydroxyapatite coatings used as control. The new bone was in direct contact with the implants, whereas fibrous tissue formed between the bone and implant with uncoated Ti-6Al-4V. The in vivo assessment of sphene-coated implants supports our in vitro observation and suggests that they have the ability to recruit osteogenic cells, and thus support bone formation around the implants and enhance osseointegration.

Multipolar Spindle Pole Coalescence Is a Major Source of Kinetochore Mis-Attachment and Chromosome Mis-Segregation in Cancer Cells

PubMed Central

Silkworth, William T.; Nardi, Isaac K.; Scholl, Lindsey M.; Cimini, Daniela

2009-01-01

Many cancer cells display a CIN (Chromosome Instability) phenotype, by which they exhibit high rates of chromosome loss or gain at each cell cycle. Over the years, a number of different mechanisms, including mitotic spindle multipolarity, cytokinesis failure, and merotelic kinetochore orientation, have been proposed as causes of CIN. However, a comprehensive theory of how CIN is perpetuated is still lacking. We used CIN colorectal cancer cells as a model system to investigate the possible cellular mechanism(s) underlying CIN. We found that CIN cells frequently assembled multipolar spindles in early mitosis. However, multipolar anaphase cells were very rare, and live-cell experiments showed that almost all CIN cells divided in a bipolar fashion. Moreover, fixed-cell analysis showed high frequencies of merotelically attached lagging chromosomes in bipolar anaphase CIN cells, and higher frequencies of merotelic attachments in multipolar vs. bipolar prometaphases. Finally, we found that multipolar CIN prometaphases typically possessed γ-tubulin at all spindle poles, and that a significant fraction of bipolar metaphase/early anaphase CIN cells possessed more than one centrosome at a single spindle pole. Taken together, our data suggest a model by which merotelic kinetochore attachments can easily be established in multipolar prometaphases. Most of these multipolar prometaphase cells would then bi-polarize before anaphase onset, and the residual merotelic attachments would produce chromosome mis-segregation due to anaphase lagging chromosomes. We propose this spindle pole coalescence mechanism as a major contributor to chromosome instability in cancer cells. PMID:19668340

The role of endothelial cell attachment to elastic fibre molecules in the enhancement of monolayer formation and retention, and the inhibition of smooth muscle cell recruitment.

PubMed

Williamson, Matthew R; Shuttleworth, Adrian; Canfield, Ann E; Black, Richard A; Kielty, Cay M

2007-12-01

The endothelium is an essential modulator of vascular tone and thrombogenicity and a critical barrier between the vessel wall and blood components. In tissue-engineered small-diameter vascular constructs, endothelial cell detachment in flow can lead to thrombosis and graft failure. The subendothelial extracellular matrix provides stable endothelial cell anchorage through interactions with cell surface receptors, and influences the proliferation, migration, and survival of both endothelial cells and smooth muscle cells. We have tested the hypothesis that these desired physiological characteristics can be conferred by surface coatings of natural vascular matrix components, focusing on the elastic fiber molecules, fibrillin-1, fibulin-5 and tropoelastin. On fibrillin-1 or fibulin-5-coated surfaces, endothelial cells exhibited strong integrin-mediated attachment in static conditions (82% and 76% attachment, respectively) and flow conditions (67% and 78% cell retention on fibrillin-1 or fibulin-5, respectively, at 25 dynes/cm2), confluent monolayer formation, and stable functional characteristics. Adhesion to these two molecules also strongly inhibited smooth muscle cell migration to the endothelial monolayer. In contrast, on elastin, endothelial cells attached poorly, did not spread, and had markedly impaired functional properties. Thus, fibrillin-1 and fibulin-5, but not elastin, can be exploited to enhance endothelial stability, and to inhibit SMC migration within vascular graft scaffolds. These findings have important implications for the design of vascular graft scaffolds, the clinical performance of which may be enhanced by exploiting natural cell-matrix biology to regulate cell attachment and function.

MC3T3-E1 Cells on Titanium Surfaces with Nanometer Smoothness and Fibronectin Immobilization

PubMed Central

Hayakawa, Tohru; Yoshida, Eiji; Yoshimura, Yoshitaka; Uo, Motohiro; Yoshinari, Masao

2012-01-01

The present study was aimed to evaluate the viability and total protein contents of osteoblast-like cells on the titanium surface with different surface mechanical treatment, namely, nanometer smoothing (Ra: approximately 2.0 nm) and sandblasting (Ra: approximately 1.0 μm), and biochemical treatment, namely, with or without fibronectin immobilization. Fibronectin could be easily immobilized by tresyl chloride-activation technique. MC3T3-E1 cells were seeded on the different titanium surfaces. Cell viability was determined by MTT assay. At 1 day of cell culture, there were no significant differences in cell viability among four different titanium surfaces. At 11 days, sandblasted titanium surface with fibronectin immobilization showed the significantly highest cell viability than other titanium surface. No significant differences existed for total protein contents among four different titanium surfaces at 11 days of cell culture. Scanning electron microscopy observation revealed that smoothness of titanium surface produced more spread cell morphologies, but that fibronectin immobilization did not cause any changes of the morphologies of attached cells. Fibronectin immobilization provided greater amount of the number of attached cells and better arrangement of attached cells. In conclusion, the combination of sandblasting and fibronectin immobilization enhanced the cell viability and fibronectin immobilization providing better arrangements of attached cells. PMID:22675359

Effect of Micro- and Nanoscale Topography on the Adhesion of Bacterial Cells to Solid Surfaces

PubMed Central

Hsu, Lillian C.; Fang, Jean; Borca-Tasciuc, Diana A.; Worobo, Randy W.

2013-01-01

Attachment and biofilm formation by bacterial pathogens on surfaces in natural, industrial, and hospital settings lead to infections and illnesses and even death. Minimizing bacterial attachment to surfaces using controlled topography could reduce the spreading of pathogens and, thus, the incidence of illnesses and subsequent human and financial losses. In this context, the attachment of key microorganisms, including Escherichia coli, Listeria innocua, and Pseudomonas fluorescens, to silica and alumina surfaces with micron and nanoscale topography was investigated. The results suggest that orientation of the attached cells occurs preferentially such as to maximize their contact area with the surface. Moreover, the bacterial cells exhibited different morphologies, including different number and size of cellular appendages, depending on the topographical details of the surface to which they attached. This suggests that bacteria may utilize different mechanisms of attachment in response to surface topography. These results are important for the design of novel microbe-repellant materials. PMID:23416997

Biofunctionalization of a titanium surface with a nano-sawtooth structure regulates the behavior of rat bone marrow mesenchymal stem cells

PubMed Central

Zhang, Wenjie; Li, Zihui; Liu, Yan; Ye, Dongxia; Li, Jinhua; Xu, Lianyi; Wei, Bin; Zhang, Xiuli; Liu, Xuanyong; Jiang, Xinquan

2012-01-01

Background: The topography of an implant surface can serve as a powerful signaling cue for attached cells and can enhance the quality of osseointegration. A series of improved implant surfaces functionalized with nanoscale structures have been fabricated using various methods. Methods: In this study, using an H2O2 process, we fabricated two size-controllable sawtooth-like nanostructures with different dimensions on a titanium surface. The effects of the two nano-sawtooth structures on rat bone marrow mesenchymal stem cells (BMMSCs) were evaluated without the addition of osteoinductive chemical factors. Results: These new surface modifications did not adversely affect cell viability, and rat BMMSCs demonstrated a greater increase in proliferation ability on the surfaces of the nano-sawtooth structures than on a control plate. Furthermore, upregulated expression of osteogenic-related genes and proteins indicated that the nano-sawtooth structures promote osteoblastic differentiation of rat BMMSCs. Importantly, the large nano-sawtooth structure resulted in the greatest cell responses, including increased adhesion, proliferation, and differentiation. Conclusion: The enhanced adhesion, proliferation, and osteogenic differentiation abilities of rat BMMSCs on the nano-sawtooth structures suggest the potential to induce improvements in bone-titanium integration in vivo. Our study reveals the key role played by the nano-sawtooth structures on a titanium surface for the fate of rat BMMSCs and provides insights into the study of stem cell-nanostructure relationships and the related design of improved biomedical implant surfaces. PMID:22927760

Calmodulin enhances ribbon replenishment and shapes filtering of synaptic transmission by cone photoreceptors

PubMed Central

Parmelee, Caitlyn M.; Chen, Minghui; Cork, Karlene M.; Curto, Carina; Thoreson, Wallace B.

2014-01-01

At the first synapse in the vertebrate visual pathway, light-evoked changes in photoreceptor membrane potential alter the rate of glutamate release onto second-order retinal neurons. This process depends on the synaptic ribbon, a specialized structure found at various sensory synapses, to provide a supply of primed vesicles for release. Calcium (Ca2+) accelerates the replenishment of vesicles at cone ribbon synapses, but the mechanisms underlying this acceleration and its functional implications for vision are unknown. We studied vesicle replenishment using paired whole-cell recordings of cones and postsynaptic neurons in tiger salamander retinas and found that it involves two kinetic mechanisms, the faster of which was diminished by calmodulin (CaM) inhibitors. We developed an analytical model that can be applied to both conventional and ribbon synapses and showed that vesicle resupply is limited by a simple time constant, τ = 1/(Dρδs), where D is the vesicle diffusion coefficient, δ is the vesicle diameter, ρ is the vesicle density, and s is the probability of vesicle attachment. The combination of electrophysiological measurements, modeling, and total internal reflection fluorescence microscopy of single synaptic vesicles suggested that CaM speeds replenishment by enhancing vesicle attachment to the ribbon. Using electroretinogram and whole-cell recordings of light responses, we found that enhanced replenishment improves the ability of cone synapses to signal darkness after brief flashes of light and enhances the amplitude of responses to higher-frequency stimuli. By accelerating the resupply of vesicles to the ribbon, CaM extends the temporal range of synaptic transmission, allowing cones to transmit higher-frequency visual information to downstream neurons. Thus, the ability of the visual system to encode time-varying stimuli is shaped by the dynamics of vesicle replenishment at photoreceptor synaptic ribbons. PMID:25311636

Fundamentals in Microalgae Harvesting: From Flocculation to Self-attachment

NASA Astrophysics Data System (ADS)

Cui, Yan

Microalgae are a very promising source of biodiesel and other renewable energy due to their fast grow rates, high lipid contents and tremendous potential for water conservation and CO2 biofixation. However, a bottleneck issue with algae biofuel manufacturing is the lack of cost-effective harvesting methods. This research focuses on the technologies for improved microalgae harvesting to enable commercially viable and environmentally friendly biodiesel production. The first objective of this study was to optimize flocculation of marine microalga Nannochloropsis oculata with metal salts, aluminum sulfate (A.S.) and ferric chloride (F.C.) via response surface methodology. It was found that there existed a positive stoichiometric relationship between the flocculant dose (FD) and the initial biomass concentration (IABC). Optimum flocculation conditions were predicted at IABC of 1.7 g/l, pH 8.3, and FD of 383.5 microM for A.S., and IABC of 2.2 g/l, pH 7.9, and FD of 438.1 microM for F.C., under which the predicted maximum harvested solid concentration of algae were 32.98 and 30.10 g/l by using A.S. and F.C., respectively. The second objective was to investigate the mechanism of microalgae flocculation with metal salts. The theory of Derjaguin, Landau, Verwey and Overbeek (DLVO) was applied to understand the flocculation mechanism of a freshwater alga Scenedesmus dimorphus and a marine alga Nannochloropsis oculata under various pH and aluminum sulphate ionic strengths. Effective flocculation was achieved as a result of charge neutralization and sweep flocculation. When low flocculant dosage (<0.1 mM) was applied, charge neutralization seemed to be predominating and the DLVO predicted flocculation trends were found quantitatively accurate in matching the experimental results. In the case of high flocculant dosage, the DLVO theory failed to explain the results since sweep flocculation was found to predominate at alum dose above 1 mM. Understanding of cell-to-cell interactions of microalgae offered possibilities in design of a novel semi-immobilized algal production and harvesting method, which exploited cell to substrata interactions instead of cell-to-cell interactions. In such method, a solid carrier was used to grow and accumulate algal cells and the cost of harvesting and drying can be simply reduced by easy algae-water separation. In order to enable the envisioned algal attachment, the third objective was to investigate the cell to substrata attachment by a thermodynamic model. Based on the theoretical analysis, when the polar surface energy of the cell is greater than that of water, cellular attachment would be more favorable on materials with higher dispersive surface energy but lower polar surface energy. If the polar surface energy of the cell is smaller than that of water, more cell attachment would be expected on materials that are higher in both dispersive and polar surface energies. The model was also validated its capability in designing, selecting, and matching algal strains and solid carrier materials to enhance cell attachment. The forth objective was to investigate the effect of surface texturing on algal attachment. It was found that surface texturing had a greater effect than surface free energy, by changing the liquid wetting behavior and real contact area. The attachment is preferred when the feature size is close to the diameter of the cell attempting to settle. Larger or smaller feature dimensions have the potential to reduce cellular attachment. The fifth objective was to study the role of carrier materials and their surface roughness in attachment. If the surface chemical composition was similar, the attachment and orientation of algal cells was found to depend on the surface average roughness, wherein rougher surfaces resulted in increased attachment. Whereas, the attachment was strongly related to surface free energy as described by the thermodynamic model if materials were chemically different.

Active metal oxides and polymer hybrids as biomaterials

NASA Astrophysics Data System (ADS)

Jarrell, John D.

Bone anchored prosthetic attachments, like other percutaneous devices, suffer from poor soft tissue integration, seen as chronic inflammation, infection, epithelial downgrowth and regression. We looked at the use of metal oxides as bioactive agents that elicit different bioresponses, ranging from cell attachment, tissue integration and reduction of inflammation to modulation of cell proliferation, morphology and microbe killing. This study presents a novel method for creating titanium oxide and polydimethylsiloxane (PDMS) hybrid coated microplates for high throughput biological, bacterial and photocatalytic screening that overcomes several limitations of using bulk metal samples. Titanium oxide coatings were doped with silver, zinc, vanadium, aluminum, calcium and phosphorous, while PDMS was doped with titanium, vanadium and silver and subjected to hydrothermal heat treatment to determine the influence of chemistry and crystallinity on the viability, proliferation and adhesion of human fibroblasts, keratinocytes and Hela cells. Also explored was the influence of Ag and Zn doping on E. coli proliferation. We determined how titanium concentration in hybrids and silver doping influenced the photocatalytic degradation of methylene blue by coatings. A combined sub/percutaneous, polyurethane device was developed and implanted into the backs of CD hairless rats to investigate how optimized coatings influenced soft tissue integration in vivo. We demonstrate that the bioresponse of cells to coatings is controlled by elemental doping (V & Ag) and that planktonic bacterial growth was greatly reduced or stopped by Ag, but not Zn doping. Hydrothermal heat treatments (65 °C and 121 °C) did not greatly influence cellular bioresponse to coatings. We discovered a range of temperature resistant (up to 400 °C), solid state dispersions with enhanced ability to block full spectrum photon transmission and degrade methylene using medical x-rays, UV, visible and infrared photons. We show that silver doping improved the photoactivity of oxide coatings, but hindered activity of a specific hybrid. Doped titanium oxide and polymer hybrid coatings have potential for improving soft tissue integration of medical implants and wound healing by modulating cell proliferation, attachment, inflammation and providing controlled delivery of bioactive and antimicrobial compounds and photon induced electro-chemical activity.

76 FR 44006 - Information Collection Being Reviewed by the Federal Communications Commission

Federal Register 2010, 2011, 2012, 2013, 2014

2011-07-22

...) Attachment 1--Community Mental Health Center Verification Template; (2) Attachment 2--Invoice Template; (3... community's ability to provide a rapid and coordinated response in the event of a public health crisis.... Title: Universal Service--Rural Health Care Program/Rural Health Care Pilot Program. Form Nos.: FCC...

Ecomorphology of parasite attachment: experiments with feather lice.

PubMed

Bush, Sarah E; Sohn, Edward; Clayton, Dale H

2006-02-01

The host specificity of some parasites can be reinforced by morphological specialization for attachment to mobile hosts. For example, ectoparasites with adaptations for attaching to hosts of a particular size might not be able to remain attached to larger or smaller hosts. This hypothesis is suggested by the positive correlation documented between the body sizes of many parasites and their hosts. We adopted an ecomorphological approach to test the attachment hypothesis. We tested the ability of host-specific feather lice (Phthiraptera: Ischnocera) to attach to 6 novel species of pigeons and doves that vary in size by nearly 2 orders of magnitude. Surprisingly, Rock Pigeon lice (Columbicola columbae) remained attached equally well to all 6 novel host species. We tested the relative importance of 3 factors that could facilitate louse attachment: whole-body insertion, tarsal claw use, and mandible use. Insertion, per se, was not necessary for attachment. However, insertion on coarse feathers of large hosts allowed lice to access feather barbules with their mandibles. Mandible use was a key component of attachment regardless of feather size. Attachment constraints do not appear to reinforce host specificity in this system.

Microscopic Behavior Of Colloidal Particles Under The Effect Of Acoustic Stimulations In The Ultrasonic To Megasonic Range

NASA Astrophysics Data System (ADS)

Abdel-Fattah, Amr I.; Roberts, Peter M.

2006-05-01

It is well known that colloid attachment and detachment at solid surfaces are influenced strongly by physico-chemical conditions controlling electric double layer (EDL) and solvation-layer effects. We present experimental observations demonstrating that, in addition, acoustic waves can produce strong effects on colloid/surface interactions that can alter the behavior of colloid and fluid transport in porous media. Microscopic colloid visualization experiments were performed with polystyrene micro-spheres suspended in water in a parallel-plate glass flow cell. When acoustic energy was applied to the cell at frequencies from 500 kHz to 5 MHz, changes in colloid attachment to and detachment from the glass cell surfaces were observed. Quantitative measurements of acoustically-induced detachment of 300-nm microspheres in 0.1M NaCl solution demonstrated that roughly 30% of the colloids that were attached to the glass cell wall during flow alone could be detached rapidly by applying acoustics at frequencies in the range of 0.7 to 1.2 MHz. The remaining attached colloids could not be detached by acoustics. This implies the existence of both "strong" and "weak" attachment sites at the cell surface. Subsequent re-attachment of colloids with acoustics turned off occurred only at new, previously unoccupied sites. Thus, acoustics appears to accelerate simultaneously both the deactivation of existing weak sites where colloids are already attached, and the activation of new weak sites where future attachments can occur. Our observations indicate that acoustics (and, in general, dynamic stress) can influence colloid-colloid and colloid-surface interactions in ways that could cause significant changes in porous-media permeability and mass transport. This would occur due to either buildup or release of colloids present in the porous matrix.

Attachment of an anti-MUC1 monoclonal antibody to 5-FU loaded BSA nanoparticles for active targeting of breast cancer cells.

PubMed

Kouchakzadeh, Hasan; Shojaosadati, Seyed Abbas; Mohammadnejad, Javad; Paknejad, Malihe; Rasaee, Mohammad Javad

2012-01-01

With PR81 as a murine monoclonal antibody (mAb) that was prepared against the human breast cancer, the MUC1 receptor specific targeting is possible. In this study, PR81-conjugated bovine serum albumin (BSA) nanoparticles loaded with anticancer drug 5-fluorouracil (5-FU) were developed. Enzyme linked immunosorbant assay (ELISA) results showed high immunoreactivity of PR81 mAb conjugated to nanoparticles towards MUC1 related peptide or native cancerous MUC1 and almost no cross-reaction to non-specific proteins. In vitro experiments were performed to determine the ability of this new drug delivery system on overcoming MCF-7 breast cancer cells in comparison with four other systems. The results revealed that these cell-type specific drug loaded nanoparticles could achieve more cell death as compared to when the 5-FU was used with no carriers. Stability studies of produced drug delivery system proved high immunoreactivity of conjugated PR81 even after 11 days of storage in room temperature.

Methods for Maintaining Insect Cell Cultures

PubMed Central

Lynn, Dwight E.

2002-01-01

Insect cell cultures are now commonly used in insect physiology, developmental biology, pathology, and molecular biology. As the field has advanced from methods development to a standard procedure, so has the diversity of scientists using the technique. This paper describes methods that are effective for maintaining various insect cell lines. The procedures are differentiated between loosely or non-attached cell strains, attached cell strains, and strongly adherent cell strains. PMID:15455043

Nitric acid passivation does not affect in vitro biocompatibility of titanium.

PubMed

Faria, Adriana C L; Beloti, Márcio M; Rosa, Adalberto L

2003-01-01

In general, both chemical composition and surface features of implants affect cell response. The aim of this study was to evaluate the effect of titanium (Ti) passivation on the response of rat bone marrow cells, considering cell attachment, cell morphology, cell proliferation, total protein content, alkaline phosphatase (ALP) activity, and bonelike nodule formation. Cells were cultured on both commercially pure titanium (cpTi) and titanium-aluminium-vanadium alloy (Ti-6Al-4V) discs, either passivated or not. For attachment evaluation, cells were cultured for 4 and 24 hours. Cell morphology was evaluated after 4 days. After 7, 14, and 21 days, cell proliferation, total protein content, and ALP activity were evaluated. Bonelike nodule formation was evaluated after 21 days. Data were compared by analysis of variance and the Duncan multiple range test. Cell attachment, cell morphology, cell proliferation, total protein content, ALP activity, and bonelike nodule formation all were unaffected by Ti composition or passivation. Although the protocol for passivation used here could interfere with the pattern of ions released from Ti-6Al-4V and cpTi surfaces, the present study did not show any effect of this surface treatment on in vitro biocompatibility of Ti as evaluated by osteoblast attachment, proliferation, and differentiation.

Novel method for in vitro depletion of T cells by monoclonal antibody-targeted photosensitization.

PubMed

Berki, T; Németh, P

1998-02-01

An immunotargeting method (called photo-immunotargeting) has been developed for selective in vitro cell destruction. The procedure combines the photosensitizing (toxic) effect of light-induced dye-molecules, e.g., hematoporphyrin (HP) and the selective binding ability of monoclonal antibodies (mAb) to cell surface molecules. The photosensitizer HP molecules were covalently attached to monoclonal antibodies (a-Thy-1) recognizing an antigen on the surface of T lymphocytes, and used for T cell destruction. To increase the selectivity of the conventional targeting methods, a physical activation step (local light irradiation) as a second degree of specificity was employed. The HP in conjugated form was sufficient to induce T cell (thymocytes, EL-4 cell line) death after irradiation at 400 nm, at tenfold lower concentration compared to the photosensitizing effect of unbound HP. The selective killing of T lymphocytes (bearing the Thy-1 antigen) in a mixed cell population was demonstrated after a treatment with the phototoxic conjugate and light irradiation. This method can be useful for selective destruction of one population (target cell) in an in vitro heterogeneous cell mixture, e.g., in bone marrow transplants for T cell depletion to avoid graft vs. host reaction.

Electrochemical Characteristics of Cell Cultured Ti-Nb-Zr Alloys After Nano-Crystallized Si-HA Coating.

PubMed

Jeong, Yong-Hoon; Choe, Han-Cheol

2015-01-01

The aim of this study was to investigate the electrochemical characteristics of nano crystallized Si-HA coating on Ti-Nb-Zr alloy after human osteoblast like (HOB) cell attachment. The Ti-Nb-Zr alloy was manufactured with 35 wt.% of Nb and 10 wt.% of Zr by arc melting furnace to appropriate physical properties as biomaterials. The HA and Si-substituted coatings were prepared by electron-beam physical vapor deposition method with 0.5, 0.8 and 1.2 wt.% of Si contents, and nano aging treatment was performed 500 degrees C for 1 h. The characteristics of coating surface were analyzed by field emission scanning electron microscopy, energy dispersive X-ray spectroscopy, and X-ray diffraction, respectively. To evaluate of cell attachment on cell cultured surface, the potentiodynamic test was performed on the surface using HOB cells. The results showed that the Si-HA coating surface showed higher tendency of cell attachment than that of single HA coating, corrosion resistance value was increased by dense of cell attachment.

Electronic platform for real-time multi-parametric analysis of cellular behavior post-exposure to single-walled carbon nanotubes

PubMed Central

Eldawud, Reem; Wagner, Alixandra; Dong, Chenbo; Rojansakul, Yon; Dinu, Cerasela Zoica

2016-01-01

Single-walled carbon nanotubes (SWCNTs) implementation in a variety of biomedical applications from bioimaging, to controlled drug delivery and cellular-directed alignment for muscle myofiber fabrication, has raised awareness of their potential toxicity. Nanotubes structural aspects which resemble asbestos, as well as their ability to induce cyto and genotoxicity upon interaction with biological systems by generating reactive oxygen species or inducing membrane damage, just to name a few, have led to focused efforts aimed to assess associated risks prior their user implementation. In this study, we employed a non-invasive and real-time electric cell impedance sensing (ECIS) platform to monitor behavior of lung epithelial cells upon exposure to a library of SWCNTs with user-defined physicochemical properties. Using the natural sensitivity of the cells, we evaluated SWCNT-induced cellular changes in relation to cell attachment, cell–cell interactions and cell viability respectively. Our methods have the potential to lead to the development of standardized assays for risk assessment of other nanomaterials as well as risk differentiation based on the nanomaterials surface chemistry, purity and agglomeration state. PMID:25913448

Inhibition of experimental ascending urinary tract infection by an epithelial cell-surface receptor analogue

NASA Astrophysics Data System (ADS)

Edén, C. Svanborg; Freter, R.; Hagberg, L.; Hull, R.; Hull, S.; Leffler, H.; Schoolnik, G.

1982-08-01

It has been shown that the establishment of urinary tract infection by Escherichia coli is dependent on attachment of the bacteria to epithelial cells1-4. The attachment involves specific epithelial cell receptors, which have been characterized as glycolipids5-10. Reversible binding to cell-surface mannosides may also be important4,11-13. This suggests an approach to the treatment of infections-that of blocking bacterial attachment with cell membrane receptor analogues. Using E. coli mutants lacking one or other of the two binding specificities (glycolipid and mannose), we show here that glycolipid analogues can block in vitro adhesion and in vivo urinary tract infection.

Integration of multiple cell-matrix interactions into alginate scaffolds for promoting cardiac tissue regeneration.

PubMed

Sapir, Yulia; Kryukov, Olga; Cohen, Smadar

2011-03-01

Cardiac tissue engineering aims to repair damaged myocardial tissues by applying heart patches created in vitro. Herein, we explored the possible role of a combination of two matrix-attached peptides, the adhesion peptide G(4)RGDY and heparin-binding peptide G(4)SPPRRARVTY (HBP) in cardiac tissue regeneration. Neonatal rat cardiac cells were seeded into unmodified, single peptide or double peptide-attached alginate scaffolds, all having the same physical features of porosity, hydrogel forming and matrix stiffness. The cardiac tissue developed in the HBP/RGD-attached scaffolds revealed the best features of a functional muscle tissue, as judged by all studied parameters, i.e., immunostaining of cardiac cell markers, histology, western blot of protein expressions and metabolic activity. By day 7, well-developed myocardial fibers were observed in these cell constructs. At 14 days the HBP/RGD-attached constructs presented an isotropic myofiber arrangement, while no such arrangement was seen in the other constructs. The expression levels of α-actinin, N-cadherin and Connexin-43, showing preservation and an increase in Connexin-43 expression (Cx-43) with time, further supported the formation a contractile muscle tissue in the HBP/RGD-attached scaffolds. Collectively, the attachment of combinatorial peptides representing different signaling in ECM-cell interactions proved to play a key role, contributing to the formation of a functional cardiac muscle tissue, in vitro. Copyright © 2010 Elsevier Ltd. All rights reserved.

The EphA8 Receptor Regulates Integrin Activity through p110γ Phosphatidylinositol-3 Kinase in a Tyrosine Kinase Activity-Independent Manner

PubMed Central

Gu, Changkyu; Park, Soochul

2001-01-01

Recent genetic studies suggest that ephrins may function in a kinase-independent Eph receptor pathway. Here we report that expression of EphA8 in either NIH 3T3 or HEK293 cells enhanced cell adhesion to fibronectin via α5β1- or β3 integrins. Interestingly, a kinase-inactive EphA8 mutant also markedly promoted cell attachment to fibronectin in these cell lines. Using a panel of EphA8 point mutants, we have demonstrated that EphA8 kinase activity does not correlate with its ability to promote cell attachment to fibronectin. Analysis using EphA8 extracellular and intracellular domain mutants has revealed that enhanced cell adhesion is dependent on ephrin A binding to the extracellular domain and the juxtamembrane segment of the cytoplasmic domain of the receptor. EphA8-promoted adhesion was efficiently inhibited by wortmannin, a phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor. Additionally, we found that EphA8 had associated PI 3-kinase activity and that the p110γ isoform of PI 3-kinase is associated with EphA8. In vitro binding experiments revealed that the EphA8 juxtamembrane segment was sufficient for the formation of a stable complex with p110γ. Similar results were obtained in assay using cells stripped of endogenous ephrin A ligands by treatment with preclustered ephrin A5-Fc proteins. In addition, a membrane-targeted lipid kinase-inactive p110γ mutant was demonstrated to stably associate with EphA8 and suppress EphA8-promoted cell adhesion to fibronectin. Taken together, these results suggest the presence of a novel mechanism by which the EphA8 receptor localizes p110γ PI 3-kinase to the plasma membrane in a tyrosine kinase-independent fashion, thereby allowing access to lipid substrates to enable the signals required for integrin-mediated cell adhesion. PMID:11416136

Process-Structure-Property Relationship in Magnesium-Based Biodegradable Alloy for Biomedical Applications

NASA Astrophysics Data System (ADS)

Trivedi, Pramanshu

Magnesium alloys are considered to be the next generation of biomaterials because of their ability to degrade in the physiological environment. We elucidate here the impact of multiaxial forging of Mg-2Zn-2Gd alloy on grain refinement to sub-micron regime and relate the structure to mechanical properties and biological functionality. As-cast and annealed samples were multiaxial forged (MAF) for a total number of two passes with a true strain of 2/pass. Considering that the microstructure governs the biological response of materials, we studied the constituents of the microstructure in conjunction with the mechanical behavior. The antimicrobial behavior in a Mg-2Zn-2Gd alloy with different grain size in the range of 44 microm to 710 nm was studied by seeding. Surface energy and contact angle measurements using goniometer and wettability were assessed with water, SBF, n-Hexane, and DMEM. The structure-property relationship in Mg-2Zn-2Gd alloy to maintaining mechanical integrity during degradation was studied by seeding Escherichia coli ( E. coli). Furthermore, we studied the effect of degradation behavior in the presence and absence of cells. This was followed by the study of bioactivity in terms of phases present on the surface and degradation products in simulated body fluid (SBF). Magnesium coated with apatite using a biomimetic approach were placed in a 24-well culture plate with alpha-MEM media and the degradation behavior was studied in the absence and presence of cells (seeding density: 10,000 cells/cm2). The change in pH was monitored at regular intervals. Cell attachment was studied by seeding the cells for 4h and cell viability was studied by seeding the cells for up to 1, 3, and 7 days. The study underscores that the fine-grained alloys exhibited superior mechanical properties, antimicrobial resistance, and cell attachment. The degradation rate was also least for fine-grained alloy. The higher surface energy of ultrafine-grained Mg-2Zn-2Gd alloy led to the release of more Mg+2 ions at an early stage, which consequently increased the pH of the fluid in the vicinity of the implant, therefore producing an unfavorable environment for the survival of bacteria. This led to damage of bacterial cell walls and reducing their adhesion. Furthermore, a significant degree of apatite formation was an indication of high bioactivity and cell attachment along with controlled degradation in the ultrafine-grained alloy. Thus, the reduction in grain size significantly improved load bearing capacity and biological functionality of Mg-2Zn-2Gd alloy.

Influence of nanoporosity on biological response of sol-gel-derived 70S30C bioactive glass monoliths

NASA Astrophysics Data System (ADS)

Thamma, Ukrit

In the field of bioactive glasses for hard tissue regeneration, the bioactivity of a material is measured by its ability to induce the formation of hydroxyapatite (HA), Ca10(PO4)6(OH)2, under physiological conditions. Due to its close chemical crystallographic resemblance to natural bones, the newly formed HA layer has been shown to be critical for the biological interaction and bonding between the surfaces of bioactive glasses and osteoblast (bone) cells. Since the formation mechanism of HA is dependent on the dissolution behavior of the bioactive glass substrate, the characteristics of HA layer are dominated by the glass composition and structure. By introducing nanoporosity into glass structure, the dissolution rate and HA growth rate on nanoporous sol-gel-derived glasses are drastically enhanced compared to that of non-porous melt-quench glasses with the same composition. While enhanced HA growth on nanoporous glass, compared to non-porous glass, was hypothesized to be associated with greater specific surface area (SSA), other studies argued that growth rate of HA layer on nanoporous glass is dominated by nanopore size distribution, and minimally affected by the bulk SSA of the underlying glass. In order to decouple the influence of nanopore size and SSA on HA formation, we have successfully fabricated homogeneous 70S30C bioactive glass monoliths with different nanopore sizes, yet similar SSA via sol-gel process. After 3-day PBS incubation of 70S30C nanoporous glass monoliths, the presence of hydroxyapatite and Type-B carbonated hydroxyapatite (HA/B-CHA) was confirmed by XPS and FTIR. Here, we report the influence of nanopore size on HA/CHA formation pathway, growth rate, and its microstructure. Due to pore-size limited diffusion of PO43-, two HA/CHA formation pathways were observed: HA/CHA surface deposition and/or HA/CHA incorporation into nanopores. HA/CHA growth rate on the surface of a nanoporous glass monolith is dominated by the pore-size limited transport of Ca2+ ions dissolved from nanoporous glass substrates. Furthermore, with rising overall growth rate controlled by nanopore size, HA/CHA microstructures evolved from needle-like, plate-like, and flower-like, respectively. Furthermore, the levels of initial cell attachment and protein adsorption on HA/CHA microstructures formed on different nanopore sizes were investigated. The initial cell attachment was quantified by measuring the density and average size of attached MC3T3-E1 cells after 2-hour seeding period. The amounts and conformation of adsorbed proteins after 2-hour incubation with HA/CHA were characterized by Western blot and FTIR, respectively. It was shown that the amounts of protein adsorption on various HA/CHA microstructures do not correlate with the initial MC3T3-E1 attachment, while the beta-sheet/alpha-helix ratios in Amide I of bovine albumin serum (BSA) adsorbed on HA/CHA microstructures do correlate to the level of initial cell attachment. This result suggests that the beta-sheet structure in BSA interacts with and activates the RGD sequence of adhesion proteins, such as fibronectin, upon adsorption, thus significantly enhancing the initial attachment of MC3T3-E1 cells. These findings provide new insights that can lead to a more detailed fundamental understanding of protein-surface and protein-protein interactions, which are crucial for the further development of bioactive material.

Guidance: Revised Model CERCLA Section 122(g)(4) De Minimis Contributor CD and AOC and New Model ATP Provisions for De Minimis Settlements

EPA Pesticide Factsheets

Cover memorandum and four attachments containing model language for de minimis settlement consent decrees and/or administrative orders and ability to pay provisions in de minimis settlements for ability to pay and non-ability to pay parties.

Xylella fastidiosa Differentially Accumulates Mineral Elements in Biofilm and Planktonic Cells

PubMed Central

Cobine, Paul A.; Cruz, Luisa F.; Navarrete, Fernando; Duncan, Daniel; Tygart, Melissa; De La Fuente, Leonardo

2013-01-01

Xylella fastidiosa is a bacterial plant pathogen that infects numerous plant hosts. Disease develops when the bacterium colonizes the xylem vessels and forms a biofilm. Inductively coupled plasma optical emission spectroscopy was used to examine the mineral element content of this pathogen in biofilm and planktonic states. Significant accumulations of copper (30-fold), manganese (6-fold), zinc (5-fold), calcium (2-fold) and potassium (2-fold) in the biofilm compared to planktonic cells were observed. Other mineral elements such as sodium, magnesium and iron did not significantly differ between biofilm and planktonic cells. The distribution of mineral elements in the planktonic cells loosely mirrors the media composition; however the unique mineral element distribution in biofilm suggests specific mechanisms of accumulation from the media. A cell-to-surface attachment assay shows that addition of 50 to 100 µM Cu to standard X. fastidiosa media increases biofilm, while higher concentrations (>200 µM) slow cell growth and prevent biofilm formation. Moreover cell-to-surface attachment was blocked by specific chelation of copper. Growth of X. fastidiosa in microfluidic chambers under flow conditions showed that addition of 50 µM Cu to the media accelerated attachment and aggregation, while 400 µM prevented this process. Supplementation of standard media with Mn showed increased biofilm formation and cell-to-cell attachment. In contrast, while the biofilm accumulated Zn, supplementation to the media with this element caused inhibited growth of planktonic cells and impaired biofilm formation. Collectively these data suggest roles for these minerals in attachment and biofilm formation and therefore the virulence of this pathogen. PMID:23349991

Xylella fastidiosa differentially accumulates mineral elements in biofilm and planktonic cells.

PubMed

Cobine, Paul A; Cruz, Luisa F; Navarrete, Fernando; Duncan, Daniel; Tygart, Melissa; De La Fuente, Leonardo

2013-01-01

Xylella fastidiosa is a bacterial plant pathogen that infects numerous plant hosts. Disease develops when the bacterium colonizes the xylem vessels and forms a biofilm. Inductively coupled plasma optical emission spectroscopy was used to examine the mineral element content of this pathogen in biofilm and planktonic states. Significant accumulations of copper (30-fold), manganese (6-fold), zinc (5-fold), calcium (2-fold) and potassium (2-fold) in the biofilm compared to planktonic cells were observed. Other mineral elements such as sodium, magnesium and iron did not significantly differ between biofilm and planktonic cells. The distribution of mineral elements in the planktonic cells loosely mirrors the media composition; however the unique mineral element distribution in biofilm suggests specific mechanisms of accumulation from the media. A cell-to-surface attachment assay shows that addition of 50 to 100 µM Cu to standard X. fastidiosa media increases biofilm, while higher concentrations (>200 µM) slow cell growth and prevent biofilm formation. Moreover cell-to-surface attachment was blocked by specific chelation of copper. Growth of X. fastidiosa in microfluidic chambers under flow conditions showed that addition of 50 µM Cu to the media accelerated attachment and aggregation, while 400 µM prevented this process. Supplementation of standard media with Mn showed increased biofilm formation and cell-to-cell attachment. In contrast, while the biofilm accumulated Zn, supplementation to the media with this element caused inhibited growth of planktonic cells and impaired biofilm formation. Collectively these data suggest roles for these minerals in attachment and biofilm formation and therefore the virulence of this pathogen.

Strong attachment of circadian pacemaker neurons on modified ultrananocrystalline diamond surfaces.

PubMed

Voss, Alexandra; Wei, HongYing; Zhang, Yi; Turner, Stuart; Ceccone, Giacomo; Reithmaier, Johann Peter; Stengl, Monika; Popov, Cyril

2016-07-01

Diamond is a promising material for a number of bio-applications, including the fabrication of platforms for attachment and investigation of neurons and of neuroprostheses, such as retinal implants. In the current work ultrananocrystalline diamond (UNCD) films were deposited by microwave plasma chemical vapor deposition, modified by UV/O3 treatment or NH3 plasma, and comprehensively characterized with respect to their bulk and surface properties, such as crystallinity, topography, composition and chemical bonding nature. The interactions of insect circadian pacemaker neurons with UNCD surfaces with H-, O- and NH2-terminations were investigated with respect to cell density and viability. The fast and strong attachment achieved without application of adhesion proteins allowed for advantageous modification of dispersion protocols for the preparation of primary cell cultures. Centrifugation steps, which are employed for pelletizing dispersed cells to separate them from dispersing enzymes, easily damage neurons. Now centrifugation can be avoided since dispersed neurons quickly and strongly attach to the UNCD surfaces. Enzyme solutions can be easily washed off without losing many of the dispersed cells. No adverse effects on the cell viability and physiological responses were observed as revealed by calcium imaging. Furthermore, the enhanced attachment of the neurons, especially on the modified UNCD surfaces, was especially advantageous for the immunocytochemical procedures with the cell cultures. The cell losses during washing steps were significantly reduced by one order of magnitude in comparison to controls. In addition, the integration of a titanium grid structure under the UNCD films allowed for individual assignment of physiologically characterized neurons to immunocytochemically stained cells. Thus, employing UNCD surfaces free of foreign proteins improves cell culture protocols and immunocytochemistry with cultured cells. The fast and strong attachment of neurons was attributed to a favorable combination of topography, surface chemistry and wettability. Copyright © 2016 Elsevier B.V. All rights reserved.

Phenazines affect biofilm formation by Pseudomonas aeruginosa in similar ways at various scales

PubMed Central

Ramos, Itzel; Dietrich, Lars E. P.; Price-Whelan, Alexa; Newman, Dianne K.

2010-01-01

Pseudomonads produce phenazines, a group of small, redox-active compounds with diverse physiological functions. In this study, we compared the phenotypes of Pseudomonas aeruginosa strain PA14 and a mutant unable to synthesize phenazines in flow cell and colony biofilms quantitatively. Although phenazine production does not impact the ability of PA14 to attach to surfaces, as has been shown for Pseudomonas chlororaphis (Maddula, 2006; Maddula, 2008), it influences swarming motility and the surface-to-volume ratio of mature biofilms. These results indicate that phenazines affect biofilm development across a large range of scales, but in unique ways for different Pseudomonas species. PMID:20123017

Bovine dedifferentiated adipose tissue (DFAT) cells

PubMed Central

Wei, Shengjuan; Du, Min; Jiang, Zhihua; Duarte, Marcio S; Fernyhough-Culver, Melinda; Albrecht, Elke; Will, Katja; Zan, Linsen; Hausman, Gary J; Elabd, Elham M Youssef; Bergen, Werner G; Basu, Urmila; Dodson, Michael V

2013-01-01

Dedifferentiated fat cells (DFAT cells) are derived from lipid-containing (mature) adipocytes, which possess the ability to symmetrically or asymmetrically proliferate, replicate, and redifferentiate/transdifferentiate. Robust cell isolation and downstream culture methods are needed to isolate large numbers of DFAT cells from any (one) adipose depot in order to establish population dynamics and regulation of the cells within and across laboratories. In order to establish more consistent/repeatable methodology here we report on two different methods to establish viable DFAT cell cultures: both traditional cell culture flasks and non-traditional (flat) cell culture plates were used for ceiling culture establishment. Adipocytes (maternal cells of the DFAT cells) were easier to remove from flat culture plates than flasks and the flat plates also allowed cloning rings to be utilized for cell/cell population isolation. While additional aspects of usage of flat-bottomed cell culture plates may yet need to be optimized by definition of optimum bio-coating to enhance cell attachment, utilization of flat plate approaches will allow more efficient study of the dedifferentiation process or the DFAT progeny cells. To extend our preliminary observations, dedifferentiation of Wagyu intramuscular fat (IMF)-derived mature adipocytes and redifferentiation ability of DFAT cells utilizing the aforementioned isolation protocols were examined in traditional basal media/differentiation induction media (DMI) containing adipogenic inducement reagents. In the absence of treatment approximately 10% isolated Wagyu IMF-mature adipocytes dedifferentiated spontaneously and 70% DFAT cells displayed protracted adipogenesis 12 d after confluence in vitro. Lipid-free intracellular vesicles in the cytoplasm (vesicles possessing an intact membrane but with no any observable or stainable lipid inside) were observed during redifferentiation. One to 30% DFAT cells redifferentiated into lipid-assimilating adipocytes in the DMI media, with distinct lipid-droplets in the cytoplasm and with no observable lipid-free vesicles inside. Moreover, a high confluence level promoted the redifferentiation efficiency of DFAT cells. Wagyu IMF dedifferentiated DFAT cells exhibited unique adipogenesis modes in vitro, revealing a useful cell model for studying adipogenesis and lipid metabolism. PMID:23991361

Intervening with Foster Parents to Enhance Biobehavioral Outcomes among Infants and Toddlers

ERIC Educational Resources Information Center

Dozier, Mary; Bick, Johanna; Bernard, Kristin

2011-01-01

Children in foster care face a number of challenges that threaten their ability to form attachment relationships with foster parents and to regulate their behavior and biology. The authors describe the Attachment and Biobehavioral Catch-Up (ABC) intervention, an evidence-based intervention aimed at helping foster children develop trusting…

Evaluations and Orientations in Mother-Child Narratives as a Function of Attachment Security: A Longitudinal Investigation

ERIC Educational Resources Information Center

Newcombe, Rhiannon; Reese, Elaine

2004-01-01

The present study examined the socialisation of children's narrative ability across the preschool period, exploring the association between children's and mothers' narrative style and children's attachment security. Fifty-six children and their mothers engaged in past event memory conversations about everyday shared past experiences when the…

Evaluating the Efficacy of an Attachment-Informed Psychotherapeutic Program for Incarcerated Parents

ERIC Educational Resources Information Center

Kamptner, N. Laura; Teyber, Faith H.; Rockwood, Nicholas J.; Drzewiecki, Dolly

2017-01-01

An attachment-based, psychotherapeutic parent education course was created for incarcerated mothers and fathers to help improve their ability to provide positive parenting and a more stable home environment for their children. The current study assessed the effects of this parenting curriculum on parents' reported tendencies to be abusive, their…

Influence of root exudates and soil on attachment of Pasteuria penetrans to Meloidogyne arenaria

USDA-ARS?s Scientific Manuscript database

Pasteuria penetrans is a parasite of root-knot nematodes (Meloidogyne spp.). Endospores of P. penetrans attach to the cuticle of second-stage juveniles (J2) and subsequently sterilize infected females. When encumbered by large numbers of spores, juveniles are less mobile and their ability to infect ...

Protein and cell micropatterning and its integration with micro/nanoparticles assembly.

PubMed

Yap, F L; Zhang, Y

2007-01-15

Micropatterning of proteins and cells has become very popular over the past decade due to its importance in the development of biosensors, microarrays, tissue engineering and cellular studies. This article reviews the techniques developed for protein and cell micropatterning and its biomedical applications. The prospect of integrating micro and nanoparticles with protein and cell micropatterning is discussed. The micro/nanoparticles are assembled into patterns and form the substrate for proteins and cell attachment. The assembled particles create a micro or nanotopography, depending on the size of the particles employed. The nonplanar structure can increase the surface area for biomolecules attachment and therefore enhance the sensitivity for detection in biosensors. Furthermore, a nanostructured substrate can influence the conformation and functionality of protein attached to it, while cellular response in terms of morphology, adhesion, proliferation, differentiation, etc. can be affected by a surface expressing micro or nanoscale structures. Proteins and cells tend to lose their normal functions upon attachment to substrate. By recognizing the types of topography that are favourable for preserving proteins and cell behaviour, and integrating it with micropattering will lead to the development of functional protein and cell patterns.

Oligosaccharide receptor mimics inhibit Legionella pneumophila attachment to human respiratory epithelial cells.

PubMed

Thomas, Richard J; Brooks, Tim J

2004-02-01

Legionnaire's disease is caused by the intracellular pathogen Legionella pneumophila, presenting as an acute pneumonia. Attachment is the key step during infection, often relying on an interaction between host cell oligosaccharides and bacterial adhesins. Inhibition of this interaction by receptor mimics offers possible novel therapeutic treatments. L. pneumophila attachment to the A549 cell line was significantly reduced by treatment with tunicamycin (73.6%) and sodium metaperiodate (63.7%). This indicates the importance of cell surface oligosaccharide chains in adhesion. A number of putative anti-adhesion compounds inhibited attachment to the A549 and U937 cell lines. The most inhibitory compounds were polymeric saccharides, GalNAcbeta1-4Gal, Galbeta1-4GlcNAc and para-nitrophenol. These compounds inhibited adhesion to a range of human respiratory cell lines, including nasal epithelial, bronchial epithelial and alveolar epithelial cell lines and the human monocytic cell line, U937. Some eukaryotic receptors for L. pneumophila were determined to be the glycolipids, asialo-GM1 and asialo-GM2 that contain the inhibitory saccharide moiety, GalNAcbeta1-4Gal. The identified compounds have the potential to be used as novel treatments for Legionnaire's disease.

Comparison of Two Porcine Collagen Membranes Combined with rhBMP-2 and rhBMP-9 on Osteoblast Behavior In Vitro.

PubMed

Fujioka-Kobayashi, Masako; Schaler, Benoit; Shirakata, Yoshinori; Nakamura, Toshiaki; Noguchi, Kazuyuki; Zhang, Yufeng; Miron, Richard J

To investigate the bone-inducing properties of two types of collagen membranes in combination with recombinant human bone morphogenetic protein (rhBMP)-2 and rhBMP-9 on osteoblast behavior. Porcine pericardium collagen membranes (PPCM) and porcine dermis-derived collagen membranes (PDCM) were coated with either rhBMP-2 or rhBMP-9. The adsorption and release abilities were first investigated via enzyme-linked immunosorbent assay up to 10 days. Moreover, murine bone stromal ST2 cell adhesion, proliferation, and osteoblast differentiation were assessed by MTS assay; real-time polymerase chain reaction for genes encoding runt-related transcription factor 2 (Runx2); alkaline phosphatase (ALP); and osteocalcin, ALP assay, and alizarin red staining. Both rhBMP-2 and rhBMP-9 adsorbed to collagen membranes and were gradually released over time up to 10 days. PPCM showed significantly less cell attachment, whereas PDCM demonstrated comparable cell attachment with the control tissue culture plastic at 8 hours. While both rhBMPs were shown not to affect cell proliferation, collagen membranes combined with rhBMP-9 significantly increased ALP activity at 7 days and ALP mRNA levels at either 3 or 14 days compared with the control tissue culture plastic. Furthermore, rhBMP-9 increased osteocalcin mRNA levels and alizarin red staining at 14 days compared with the control tissue culture plastic. The results from this study suggest that both porcine-derived collagen membranes combined with rhBMP-9 accelerated the osteopromotive potential of ST2 cells. Interestingly, rhBMP-9 demonstrated additional osteogenic differentiation compared with rhBMP-2 and may serve as a suitable growth factor for future clinical use.

Atypical enteropathogenic Escherichia coli that contains functional locus of enterocyte effacement genes can be attaching-and-effacing negative in cultured epithelial cells.

PubMed

Rocha, Sérgio P D; Abe, Cecilia M; Sperandio, Vanessa; Bando, Silvia Y; Elias, Waldir P

2011-05-01

Enteropathogenic Escherichia coli (EPEC) induces a characteristic histopathology on enterocytes known as the attaching-and-effacing (A/E) lesion, which is triggered by proteins encoded by the locus of enterocyte effacement (LEE). EPEC is currently classified as typical EPEC (tEPEC) and atypical EPEC (aEPEC), based on the presence or absence of the EPEC adherence factor plasmid, respectively. Here we analyzed the LEE regions of three aEPEC strains displaying the localized adherence-like (LAL), aggregative adherence (AA), and diffuse adherence (DA) patterns on HEp-2 cells as well as one nonadherent (NA) strain. The adherence characteristics and the ability to induce A/E lesions were investigated with HeLa, Caco-2, T84, and HT29 cells. The adherence patterns and fluorescent actin staining (FAS) assay results were reproducible with all cell lines. The LEE region was structurally intact and functional in all strains regardless of their inability to cause A/E lesions. An EspF(U)-expressing plasmid (pKC471) was introduced into all strains, demonstrating no influence of this protein on either the adherence patterns or the capacity to cause A/E of the adherent strains. However, the NA strain harboring pKC471 expressed the LAL pattern and was able to induce A/E lesions on HeLa cells. Our data indicate that FAS-negative aEPEC strains are potentially able to induce A/E in vivo, emphasizing the concern about this test for the determination of aEPEC virulence. Also, the presence of EspF(U) was sufficient to provide an adherent phenotype for a nonadherent aEPEC strain via the direct or indirect activation of the LEE4 and LEE5 operons.

Change in surface properties of zirconia and initial attachment of osteoblastlike cells with hydrophilic treatment.

PubMed

Watanabe, Hiroaki; Saito, Kensuke; Kokubun, Katsutoshi; Sasaki, Hodaka; Yoshinari, Masao

2012-01-01

The objectives of this study were to characterize change in surface properties of tetragonal zirconia polycrystals (TZP) after hydrophilic treatment, and to determine the effect of such changes on initial attachment of osteoblast-like cells. Roughened surfaces were produced by alumina-blasting and acid-etching. Hydrophilic treatment comprised application of immediately after blasting and acid-etching (Blast/Etch), oxygen plasma (O2-Plasma), ultraviolet light (UV). Specimens stored in air were used as a control. The water contact angle was determined and surface analysis was performed using an X-ray photoelectron spectroscopy. Blast/Etch, O2-Plasma and UV specimens showed superhydrophilicity, and these hydrophilic treatments to TZP elicited a marked decrease in carbon content and an increase in hydroxyl groups. Hydrophilic treatments enhanced initial attachment of osteoblast-like cells and a change in cell morphologies. These results indicate that Blast/Etch, O2-Plasma, or UV treatment has potential in the creation and maintenance of superhydrophilic surfaces and enhancing initial attachment of osteoblast-like cells.

Traumatic events, posttraumatic stress disorder, attachment style, and working alliance in a sample of people with psychosis.

PubMed

Picken, Alicia L; Berry, Katherine; Tarrier, Nicholas; Barrowclough, Christine

2010-10-01

There is a high incidence of trauma and posttraumatic stress disorder (PTSD) in people with a diagnosis of psychosis. Sequelae of trauma may affect the ability to engage in both attachment and therapeutic relationships. This study investigated associations between trauma histories, PTSD, attachment styles, and working alliance in a sample of 110 individuals with psychosis and substance misuse. Anxious attachment was associated with number of interpersonal traumas and PTSD reported, but there were no associations between trauma and alliance. There were discrepancies in number of traumatic events reported by care coordinators and patients. The findings of this study highlight the potential use of attachment theory in working with trauma and PTSD in psychosis.

Incorporating zosteric acid into silicone coatings to achieve its slow release while reducing fresh water bacterial attachment.

PubMed

Barrios, Carlos A; Xu, Qingwei; Cutright, Teresa; Newby, Bi-min Zhang

2005-03-25

Biofouling has posed serious problems in maritime industry including increased fuel consumptions, economic loss from ship-hull maintenances, contamination of drinking water, and serious corrosion for mechanical instruments. Minimizing the attachment of bacteria and formation of biofilm could be advantageous in reducing the early stages of biofouling. Zosteric acid, a natural product present in eelgrass, was found to have ability for preventing the attachment of some bacteria and barnacles. In this study, the antifouling ability of zosteric acid during the early stages of fouling was evaluated using attachment studies of fresh water bacteria. Simultaneously, various methods were sought for incorporating zosteric acid into silicone to prolong the release of the compound. The main results from this study were that zosteric acid exhibited anti-bacterial attachment regardless of whether it dispersed in water or incorporated into a coating. In addition, the release rate of zosteric acid from the incorporated coatings, particularly those where zosteric acid was uniformly dispersed with aggregates size of 4 microm or less, was orders of magnitude slower than those of previous reports. The release results indicate that the service life of our coatings could be far extended even with a small amount of zosteric acid incorporated.

Watch Out for the "Living Dead": Cell-Free Enzymes and Their Fate.

PubMed

Baltar, Federico

2017-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the "gatekeepers" of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell's fate. In contrast, cell-free enzymes belong to a kind of "living dead" realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go "beyond the living things," studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles.

Cell attachment functionality of bioactive conducting polymers for neural interfaces.

PubMed

Green, Rylie A; Lovell, Nigel H; Poole-Warren, Laura A

2009-08-01

Bioactive coatings for neural electrodes that are tailored for cell interactions have the potential to produce superior implants with improved charge transfer capabilities. In this study synthetically produced anionically modified laminin peptides DEDEDYFQRYLI and DCDPGYIGSR were used to dope poly(3,4-ethylenedioxythiophene) (PEDOT) electrodeposited on platinum (Pt) electrodes. Performance of peptide doped films was compared to conventional polymer PEDOT/paratoluene sulfonate (pTS) films using SEM, XPS, cyclic voltammetry, impedance spectroscopy, mechanical hardness and adherence. Bioactivity of incorporated peptides and their affect on cell growth was assessed using a PC12 neurite outgrowth assay. It was demonstrated that large peptide dopants produced softer PEDOT films with a minimal decrease in electrochemical stability, compared to the conventional dopant, pTS. Cell studies revealed that the YFQRYLI ligand retained neurite outgrowth bioactivity when DEDEDYFQRYLI was used as a dopant, but the effect was strongly dependant on initial cell attachment. Alternate peptide dopant, DCDPGYIGSR was found to impart superior cell attachment properties when compared to DEDEDYFQRYLI, but attachment on both peptide doped polymers could be enhanced by coating with whole native laminin.

Evaluation of Blood Cell Attachment on Er:Yag Laser Applied Root Surface Using Scanning Electron Microscopy

PubMed Central

CEKICI, Ali; MADEN, Ilay; YILDIZ, Sercan; SAN, Tangul; ISIK, Gulden

2013-01-01

Background: Periodontal regeneration is dependent on the uninterrupted adhesion, maturation and absorption of fibrin clots to a periodontally compromised root surface. The modification of the root surface with different agents has been used for better fibrin clot formation and blood cell attachment. It is known that Er:YAG laser application on dentin removes the smear layer succesfully. Aim: The aim of this study is to observe blood cell attachment and fibrin network formation following ER:YAG laser irradiation on periodontally compromised root surfaces in comparison to chemical root conditioning techniques in vitro. Materials and methods: 40 dentin blocks prepared from freshly extracted periodontally compromised hopeless teeth. Specimens were divided in 5 groups; those applied with PBS, EDTA, Citric acid and Er:YAG. They were further divided into two groups: those which had received these applications, and the control group. The specimens were evaluated with scanning electron microscope and micrographs were taken. Smear layer and blood cell attachment scoring was performed. Results: In the Er:YAG laser applied group, smear layer were totally removed. In the blood applied specimens, better fibrin clot formation and blood cell attachment were observed in the Er:YAG group. In the group that had been applied with citric acid, the smear layer was also removed. The smear layer could not be fully removed in the EDTA group. Conclusion: Er:YAG laser application on the root dentin seems to form a suitable surface for fibrin clot formation and blood cell attachment. Further clinical studies to support these results are necessitated. PMID:23533017

New York City social workers after 9/11: their attachment, resiliency, and compassion fatigue.

PubMed

Tosone, Carol; Bettmann, Joanna E; Minami, Takuya; Jasperson, Rachael A

2010-01-01

This study examines the relationship between attachment classification, resiliency, and compassion fatigue in New York social workers following 9/11. We used single occasion, quasi-random sampling, surveying 481 social workers living in Manhattan. Hierarchical regression analyses revealed that secure attachment is predictive of the ability to cope with secondary traumatic stress as well as capacity for resilience, explaining approximately 7% of the variance in both compassion fatigue and resiliency. These findings suggest that secure attachment may serve as a source of resilience for social workers, immunizing them from significant compassion fatigue. Such findings have significant implications for clinicians working with traumatized populations.

Novel chitin scaffolds derived from marine sponge Ianthella basta for tissue engineering approaches based on human mesenchymal stromal cells: Biocompatibility and cryopreservation.

PubMed

Mutsenko, Vitalii V; Gryshkov, Oleksandr; Lauterboeck, Lothar; Rogulska, Olena; Tarusin, Dmitriy N; Bazhenov, Vasilii V; Schütz, Kathleen; Brüggemeier, Sophie; Gossla, Elke; Akkineni, Ashwini R; Meißner, Heike; Lode, Anja; Meschke, Stephan; Fromont, Jane; Stelling, Allison L; Tabachnik, Konstantin R; Gelinsky, Michael; Nikulin, Sergey; Rodin, Sergey; Tonevitsky, Alexander G; Petrenko, Alexander Y; Glasmacher, Birgit; Schupp, Peter J; Ehrlich, Hermann

2017-11-01

The extraordinary biocompatibility and mechanical properties of chitinous scaffolds from marine sponges endows these structures with unique properties that render them ideal for diverse biomedical applications. In the present work, a technological route to produce "ready-to-use" tissue-engineered products based on poriferan chitin is comprehensively investigated for the first time. Three key stages included isolation of scaffolds from the marine demosponge Ianthella basta, confirmation of their biocompatibility with human mesenchymal stromal cells, and cryopreservation of the tissue-like structures grown within these scaffolds using a slow cooling protocol. Biocompatibility of the macroporous, flat chitin scaffolds has been confirmed by cell attachment, high cell viability and the ability to differentiate into the adipogenic lineage. The viability of cells cryopreserved on chitin scaffolds was reduced by about 30% as compared to cells cryopreserved in suspension. However, the surviving cells were able to retain their differentiation potential; and this is demonstrated for the adipogenic lineage. The results suggest that chitin from the marine demosponge I. basta is a promising, highly biocompatible biomaterial for stem cell-based tissue-engineering applications. Copyright © 2017 Elsevier B.V. All rights reserved.

The great escape: viral strategies to counter BST-2/tetherin.

PubMed

Douglas, Janet L; Gustin, Jean K; Viswanathan, Kasinath; Mansouri, Mandana; Moses, Ashlee V; Früh, Klaus

2010-05-13

The interferon-induced BST-2 protein has the unique ability to restrict the egress of HIV-1, Kaposi's sarcoma-associated herpesvirus (KSHV), Ebola virus, and other enveloped viruses. The observation that virions remain attached to the surface of BST-2-expressing cells led to the renaming of BST-2 as "tetherin". However, viral proteins such as HIV-1 Vpu, simian immunodeficiency virus Nef, and KSHV K5 counteract BST-2, thereby allowing mature virions to readily escape from infected cells. Since the anti-viral function of BST-2 was discovered, there has been an explosion of research into several aspects of this intriguing interplay between host and virus. This review focuses on recent work addressing the molecular mechanisms involved in BST-2 restriction of viral egress and the species-specific countermeasures employed by various viruses.

Mycoplasma agalactiae Secretion of β-(1→6)-Glucan, a Rare Polysaccharide in Prokaryotes, Is Governed by High-Frequency Phase Variation

PubMed Central

Baranowski, E.; Pau-Roblot, C.; Sagné, E.; Citti, C.

2016-01-01

ABSTRACT Mycoplasmas are minimal, wall-less bacteria but have retained the ability to secrete complex carbohydrate polymers that constitute a glycocalyx. In members of the Mycoplasma mycoides cluster, which are important ruminant pathogens, the glycocalyx includes both cell-attached and cell-free polysaccharides. This report explores the potential secretion of polysaccharides by M. agalactiae, another ruminant pathogen that belongs to a distant phylogenetic group. Comparative genomic analyses showed that M. agalactiae possesses all the genes required for polysaccharide secretion. Notably, a putative synthase gene (gsmA) was identified, by in silico reconstruction of the biosynthetic pathway, that could be involved in both polymerization and export of the carbohydrate polymers. M. agalactiae polysaccharides were then purified in vitro and found to be mainly cell attached, with a linear β-(1→6)-glucopyranose structure [β-(1→6)-glucan]. Secretion of β-(1→6)-glucan was further shown to rely on the presence of a functional gsmA gene, whose expression is subjected to high-frequency phase variation. This event is governed by the spontaneous intraclonal variation in length of a poly(G) tract located in the gsmA coding sequence and was shown to occur in most of the M. agalactiae clinical isolates tested in this study. M. agalactiae susceptibility to serum-killing activity appeared to be dictated by ON/OFF switching of β-(1→6)-glucan secretion, suggesting a role of this phenomenon in survival of the pathogen when it invades the host bloodstream. Finally, β-(1→6)-glucan secretion was not restricted to M. agalactiae but was detected also in M. mycoides subsp. capri PG3T, another pathogen of small ruminants. IMPORTANCE Many if not all bacteria are able to secrete polysaccharides, either attached to the cell surface or exported unbound into the extracellular environment. Both types of polysaccharides can play a role in bacterium-host interactions. Mycoplasmas are no exception despite their poor overall metabolic capacity. We showed here that M. agalactiae secretes a capsular β-(1→6)-glucopyranose thanks to a specific glycosyltransferase with synthase activity. This secretion is governed by high-frequency ON/OFF phase variation that might be crucial in mycoplasma host dissemination, as cell-attached β-(1→6)-glucopyranose increases serum-killing susceptibility. Our results provide functional genetic data about mycoplasmal glycosyltransferases with dual functions, i.e., assembly and export of the sugar polymers across the cell membrane. Furthermore, we demonstrated that nonprotein epitopes can be subjected to surface antigenic variation in mycoplasmas. Finally, the present report contributes to unravel the role of secreted polysaccharides in the virulence and pathogenicity of these peculiar bacteria. PMID:27037120

The role of basal cells in adhesion of columnar epithelium to airway basement membrane.

PubMed

Evans, M J; Plopper, C G

1988-08-01

In this report, we present a new concept of the role of the basal cell in airway epithelium. Previously, the basal cell was thought to be the progenitor cell for the columnar epithelium. However, several studies have shown that this concept may not be correct. The morphologic aspects of the basal cell suggest that it could play a role in adhesion of the columnar epithelium to the basement membrane. Basal cells form attachments with columnar cells (desmosomes) and with the basement membrane (hemidesmosomes). Columnar cells do not form hemidesmosome attachments with the basement membrane. Basal cells could strengthen the adhesion of columnar cells to the basement membrane by forming hemidesmosome attachments to the basement membrane and desmosome attachments with adjacent columnar cells. Incidental evidence from 2 existing publications concerning airway microanatomy support this concept. As columnar cells grow taller, the proportion of the cell surface in contact with the basement membrane becomes progressively smaller, and thus the cell surface area related to adhesion also becomes smaller. It was found that the number of basal cells per millimeter of basement membrane was closely related to the height of the columnar cell epithelium (r = 0.98), but not to the number of columnar cells (r = 0.42). The consistency of the relationship between increased columnar cell height (and thus decreased surface area for adhesion) and the number of basal cells present (r = 0.98) supports the concept that the basal cell plays a role in adhesion of columnar cells to the basement membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

Anti-biofilm formation of a novel stainless steel against Staphylococcus aureus.

PubMed

Nan, Li; Yang, Ke; Ren, Guogang

2015-06-01

Staphylococcus aureus (S. aureus) is a bacterium frequently found proliferating on metal surfaces such as stainless steels used in healthcare and food processing facilities. Past research has shown that a novel Cu-bearing 304 type stainless steel (304CuSS) exhibits excellent antibacterial ability (i.e. against S. aureus) in a short time period (24h.). This work was dedicated to investigate the 304CuSS's inhibition ability towards the S. aureus biofilm formation for an extended period of 7days after incubation. It was found that the antibacterial rate of the 304CuSS against sessile bacterial cells reached over 99.9% in comparison with the 304SS. The thickness and sizes of the biofilms on the 304SS surfaces increased markedly with period of contact, and thus expected higher risk of bio-contamination, indicated by the changes of surface free energy between biofilm and the steel surfaces. The results demonstrated that the 304CuSS exhibited strong inhibition on the growth and adherence of the biofilms. The surface free energy of the 304CuSS after contact with sessile bacterial cells was much lower than that of the 304SS towards the same culture times. The continuously dissolved Cu(2+) ions well demonstrated the dissolution ability of Cu-rich precipitates after exposure to S. aureus solution, from 3.1ppm (2days) to 4.5ppm (7days). For this to occur, a hypothesis mechanism might be established for 304CuSS in which the Cu(2+) ions were released from Cu-rich phases that bond with extracellular polymeric substances (EPS) of the microorganisms. And these inhibited the activities of cell protein/enzymes and effectively prevented planktonic bacterial cells attaching to the 304CuSS metal surface. Copyright © 2015. Published by Elsevier B.V.

Study on performance of magnetic fluorescent nanoparticles as gene carrier and location in pig kidney cells

NASA Astrophysics Data System (ADS)

Wang, Yan; Cui, Haixin; Sun, Changjiao; Du, Wei; Cui, Jinhui; Zhao, Xiang

2013-03-01

We evaluated the performance of green fluorescent magnetic Fe3O4 nanoparticles (NPs) as gene carrier and location in pig kidney cells. When the mass ratio of NPs to green fluorescent protein plasmid DNA reached 1:16 or above, DNA molecules can be combined completely with NPs, which indicates that the NPs have good ability to bind negative DNA. Atomic force microscopy (AFM) experiments were carried out to investigate the binding mechanism between NPs and DNA. AFM images show that individual DNA strands come off of larger pieces of netlike agglomerations and several spherical nanoparticles are attached to each individual DNA strand and interact with each other. The pig kidney cells were labelled with membrane-specific red fluorescent dye 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate and nucleus-specific blue fluorescent dye 4',6-diamidino-2-phenylindole dihydrochloride. We found that green fluorescent nanoparticles can past the cell membrane and spread throughout the interior of the cell. The NPs seem to locate more frequently in the cytoplasm than in the nucleus.

A rhamnose-rich O-antigen mediates adhesion, virulence, and host colonization for the xylem-limited phytopathogen Xylella fastidiosa.

PubMed

Clifford, Jennifer C; Rapicavoli, Jeannette N; Roper, M Caroline

2013-06-01

Xylella fastidiosa is a gram-negative, xylem-limited bacterium that causes a lethal disease of grapevine called Pierce's disease. Lipopolysaccharide (LPS) composes approximately 75% of the outer membrane of gram-negative bacteria and, because it is largely displayed on the cell surface, it mediates interactions between the bacterial cell and its surrounding environment. LPS is composed of a conserved lipid A-core oligosaccharide component and a variable O-antigen portion. By targeting a key O-antigen biosynthetic gene, we demonstrate the contribution of the rhamnose-rich O-antigen to surface attachment, cell-cell aggregation, and biofilm maturation: critical steps for successful infection of the host xylem tissue. Moreover, we have demonstrated that a fully formed O-antigen moiety is an important virulence factor for Pierce's disease development in grape and that depletion of the O-antigen compromises its ability to colonize the host. It has long been speculated that cell-surface polysaccharides play a role in X. fastidiosa virulence and this study confirms that LPS is a major virulence factor for this important agricultural pathogen.

DC-SIGN mediates avian H5N1 influenza virus infection in cis and in trans

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

Wang, S.-F.; Huang, Jason C.; AIDS Prevention and Research Center, School of Medicine, National Yang-Ming University, Taipei 112, Taiwan

2008-09-05

DC-SIGN, a C-type lectin receptor expressed in dendritic cells (DCs), has been identified as a receptor for human immunodeficiency virus type 1, hepatitis C virus, Ebola virus, cytomegalovirus, dengue virus, and the SARS coronavirus. We used H5N1 pseudotyped and reverse-genetics (RG) virus particles to study their ability to bind with DC-SIGN. Electronic microscopy and functional assay results indicate that pseudotyped viruses containing both HA and NA proteins express hemagglutination and are capable of infecting cells expressing {alpha}-2,3-linked sialic acid receptors. Results from a capture assay show that DC-SIGN-expressing cells (including B-THP-1/DC-SIGN and T-THP-1/DC-SIGN) and peripheral blood dendritic cells are capablemore » of transferring H5N1 pseudotyped and RG virus particles to target cells; this action can be blocked by anti-DC-SIGN monoclonal antibodies. In summary, (a) DC-SIGN acts as a capture or attachment molecule for avian H5N1 virus, and (b) DC-SIGN mediates infections in cis and in trans.« less

Attachment and internalization of a Chlamydia trachomatis lymphogranuloma venereum strain by McCoy cells: kinetics of infectivity and effect of lectins and carbohydrates.

PubMed Central

Söderlund, G; Kihlström, E

1983-01-01

The kinetics of attachment and ingestion of Chlamydia trachomatis serotype L1 by monolayers of McCoy cells were studied by using a method that discriminated between attachment and uptake. When about 1% of the McCoy cells was infected, the proteinase K-resistant chlamydial fraction, regarded as ingested chlamydiae, reached a constant value after about 3 h of incubation at 37 degrees C. Uptake of chlamydiae at 4 degrees C could not be demonstrated. The attached and ingested chlamydial fractions were constant over an eightfold increase in chlamydial inoculum. Chitobiose and chitotriose, the di- and trisaccharides of N-acetyl-D-glucosamine, reduced the association of C. trachomatis serotype L1 with McCoy cells. Higher concentrations of chitobiose also selectively inhibited ingestion of chlamydiae. A corresponding effect of chitobiose was also observed on the number of chlamydial inclusions. Wheat germ agglutinin, specific for N-acetyl-D-glucosamine residues, reduced the association of chlamydiae when incubated at 4 degrees C, but not at 37 degrees C. A small inhibiting effect of concanavalin A on association of chlamydiae, but no effect of the corresponding carbohydrates, indicates a nonspecific effect on chlamydial attachment of this lectin. These results suggest that beta 1 leads to 4-linked oligomers of N-acetyl-D-glucosamine are important in the specificity of attachment of C. trachomatis to McCoy cells. PMID:6642670

Expression of the Flp proteins by Haemophilus ducreyi is necessary for virulence in human volunteers.

PubMed

Janowicz, Diane M; Cooney, Sean A; Walsh, Jessica; Baker, Beth; Katz, Barry P; Fortney, Kate R; Zwickl, Beth W; Ellinger, Sheila; Munson, Robert S

2011-09-22

Haemophilus ducreyi, the causative agent of the sexually transmitted disease chancroid, contains a flp (fimbria like protein) operon that encodes proteins predicted to contribute to adherence and pathogenesis. H. ducreyi mutants that lack expression of Flp1 and Flp2 or TadA, which has homology to NTPases of type IV secretion systems, have decreased abilities to attach to and form microcolonies on human foreskin fibroblasts (HFF). A tadA mutant is attenuated in its ability to cause disease in human volunteers and in the temperature dependent rabbit model, but a flp1flp2 mutant is virulent in rabbits. Whether a flp deletion mutant would cause disease in humans is not clear. We constructed 35000HPΔflp1-3, a deletion mutant that lacks expression of all three Flp proteins but has an intact tad secretion system. 35000HPΔflp1-3 was impaired in its ability to form microcolonies and to attach to HFF in vitro when compared to its parent (35000HP). Complementation of the mutant with flp1-3 in trans restored the parental phenotype. To test whether expression of Flp1-3 was necessary for virulence in humans, ten healthy adult volunteers were experimentally infected with a fixed dose of 35000HP (ranging from 54 to 67 CFU) on one arm and three doses of 35000HPΔflp1-3 (ranging from 63 to 961 CFU) on the other arm. The overall papule formation rate for the parent was 80% (95% confidence interval, CI, 55.2%-99.9%) and for the mutant was 70.0% (95% CI, 50.5%-89.5%) (P = 0.52). Mutant papules were significantly smaller (mean, 11.2 mm2) than were parent papules (21.8 mm2) 24 h after inoculation (P = 0.018). The overall pustule formation rates were 46.7% (95% CI 23.7-69.7%) at 30 parent sites and 6.7% (95% CI, 0.1-19.1%) at 30 mutant sites (P = 0.001). These data suggest that production and secretion of the Flp proteins contributes to microcolony formation and attachment to HFF cells in vitro. Expression of flp1-3 is also necessary for H. ducreyi to initiate disease and progress to pustule formation in humans. Future studies will focus on how Flp proteins contribute to microcolony formation and attachment in vivo. © 2011 Janowicz et al; licensee BioMed Central Ltd.

Engineering solutions to improve the removal of fecal indicator bacteria by bioinfiltration systems during intermittent flow of stormwater.

PubMed

Mohanty, Sanjay K; Torkelson, Andrew A; Dodd, Hanna; Nelson, Kara L; Boehm, Alexandria B

2013-10-01

Bioinfiltration systems facilitate the infiltration of urban stormwater into soil and reduce high flow events and flooding. Stormwater carries a myriad of pollutants including fecal indicator bacteria (FIB). Significant knowledge gaps exist about the ability of bioinfiltration systems to remove and retain FIB. The present study investigates the ability of model, simplified bioinfiltration systems containing quartz sand and iron oxide-coated quartz sand (IOCS) to remove two FIB (Enterococcus faecalis and Escherichia coli) suspended in synthetic stormwater with and without natural organic matter (NOM) as well as the potential for accumulated FIB to be remobilized during intermittent flow. The experiments were conducted in two phases: (1) the saturated columns packed with either sand or IOCS were contaminated by injecting stormwater with bacteria followed by injection of sterile stormwater and (2) the contaminated columns were subjected to intermittent infiltration of sterile stormwater preceded by a pause during which columns were either kept saturated or drained by gravity. During intermittent flow, fewer bacteria were released from the saturated column compared to the column drained by gravity: 12% of attached E. coli and 3% of attached Ent. faecalis were mobilized from the drained sand column compared to 3% of attached E. coli and 2% attached Ent. faecalis mobilized from the saturated sand column. Dry and wet cycles introduce moving air-water interfaces that can scour bacteria from grain surfaces. During intermittent flows, less than 0.2% of attached bacteria were mobilized from IOCS, which bound both bacteria irreversibly in the absence of NOM. Addition of NOM, however, increased bacterial mobilization from IOCS: 50% of attached E. coli and 8% of attached Ent. faecalis were released from IOCS columns during draining and rewetting. Results indicate that using geomedia such as IOCS that promote irreversible attachment of bacteria, and maintaining saturated condition, could minimize the mobilization of previous attached bacteria from bioinfiltration systems, although NOM may significantly decrease these benefits.

Enhancement of Cell Attachment to a Substrate Coated with Oral Bacterial Endotoxin by Plasma Fibronectin

DTIC Science & Technology

1983-01-01

root surfaces is unpredict- human gingival fibroblasts (Aleo. De Renzis able (World Workshop in PeriodonticN & Farber 1975). Clearly, if new attachment...1966). of periodontal tissues to a tooth is to be A preliminary characterization of the made possible, therapeutic measures must FIBRONECTIN ENHANCES...CELL ATTACHMENT 155 list he developed it remove. alter. or other- population of bacteria wsithin the gingival wise inactivate the toxic principle of

Use of high concentrations of dimethyl sulfoxide for cryopreservation of HepG2 cells adhered to glass and polydimethylsiloxane matrices.

PubMed

Nagahara, Yukitoshi; Sekine, Hiroaki; Otaki, Mari; Hayashi, Masakazu; Murase, Norio

2016-02-01

Animal cells are generally cryopreserved in cryovials in a cell suspension state containing 5%-10% v/v dimethyl sulfoxide (DMSO) used as a cryoprotective agent. However, cryopreservation of cells in an attached state has not been intensively studied, and the effective freezing solution remains unknown. Here we determined the suitable DMSO concentration for the cryopreservation of human hepatoma HepG2 cells attached to glass and polydimethylsiloxane (PDMS) matrices coated with poly-l-lysine. With the use of the glass matrix, the rate of cell adhesion increased with the DMSO concentration up to 30% v/v in the freezing solution. In contrast, the cell-adhesion rate remained constant in the case of the PDMS matrix irrespective of the DMSO concentration between 10% v/v and 30% v/v. The viability of post-thawed cells attached to glass or PDMS matrix was also investigated. The viability was highest at the DMSO concentration of 20% v/v in the freezing solution. The DMSO concentration of 30% v/v, however, had a cytotoxic effect on the cell viability. Thus, the 20% v/v DMSO concentration was found to be most suitable for the cryopreservation of HepG2 cells in the attached state. This dose is high compared to the DMSO concentration used for the cryopreservation of cells in the suspended state. Copyright © 2015 Elsevier Inc. All rights reserved.

INTEGRIN-MEDIATED CELL ATTACHMENT SHOWS TIME-DEPENDENT UPREGULATION OF GAP JUNCTION COMMUNICATION.

EPA Science Inventory

Integrin-mediated Cell Attachment Shows Time-Dependent Upregulation of Gap Junction
Communication

Rachel Grindstaff and Carl Blackman, National Health & Environmental Effects Research
Laboratory, Office of Research and Development, US EPA, Research Triang...

Delay of Gratification in Two- and Three-Year-Olds: Associations with Attachment, Personality, and Temperament

ERIC Educational Resources Information Center

Mittal, Renu; Russell, Beth S.; Britner, Preston A.; Peake, Philip K.

2013-01-01

Delay of gratification in young children has been linked to long-term behavioral and academic outcomes. This study explored temperament, personality, and child-parent attachment as possible associates of delay ability. The sample consisted of 50 2- and 3-year-old children and their primary caregivers. Two laboratory tasks, the Preschool Strange…

Collagen Self-Assembly on Orthopedic Magnesium Biomaterials Surface and Subsequent Bone Cell Attachment

PubMed Central

Zhao, Nan; Zhu, Donghui

2014-01-01

Magnesium (Mg) biomaterials are a new generation of biodegradable materials and have promising potential for orthopedic applications. After implantation in bone tissues, these materials will directly interact with extracellular matrix (ECM) biomolecules and bone cells. Type I collagen, the major component of bone ECM, forms the architecture scaffold that provides physical support for bone cell attachment. However, it is still unknown how Mg substrate affects collagen assembly on top of it as well as subsequent cell attachment and growth. Here, we studied the effects of collagen monomer concentration, pH, assembly time, and surface roughness of two Mg materials (pure Mg and AZ31) on collagen fibril formation. Results showed that formation of fibrils would not initiate until the monomer concentration reached a certain level depending on the type of Mg material. The thickness of collagen fibril increased with the increase of assembly time. The structures of collagen fibrils formed on semi-rough surfaces of Mg materials have a high similarity to that of native bone collagen. Next, cell attachment and growth after collagen assembly were examined. Materials with rough surface showed higher collagen adsorption but compromised bone cell attachment. Interestingly, surface roughness and collagen structure did not affect cell growth on AZ31 for up to a week. Findings from this work provide some insightful information on Mg-tissue interaction at the interface and guidance for future surface modifications of Mg biomaterials. PMID:25303459

Induction of virulence factors in Giardia duodenalis independent of host attachment

PubMed Central

Emery, Samantha J.; Mirzaei, Mehdi; Vuong, Daniel; Pascovici, Dana; Chick, Joel M.; Lacey, Ernest; Haynes, Paul A.

2016-01-01

Giardia duodenalis is responsible for the majority of parasitic gastroenteritis in humans worldwide. Host-parasite interaction models in vitro provide insights into disease and virulence and help us to understand pathogenesis. Using HT-29 intestinal epithelial cells (IEC) as a model we have demonstrated that initial sensitisation by host secretions reduces proclivity for trophozoite attachment, while inducing virulence factors. Host soluble factors triggered up-regulation of membrane and secreted proteins, including Tenascins, Cathepsin-B precursor, cystatin, and numerous Variant-specific Surface Proteins (VSPs). By comparison, host-cell attached trophozoites up-regulated intracellular pathways for ubiquitination, reactive oxygen species (ROS) detoxification and production of pyridoxal phosphate (PLP). We reason that these results demonstrate early pathogenesis in Giardia involves two independent host-parasite interactions. Motile trophozoites respond to soluble secreted signals, which deter attachment and induce expression of virulence factors. Trophozoites attached to host cells, in contrast, respond by up-regulating intracellular pathways involved in clearance of ROS, thus anticipating the host defence response. PMID:26867958

Dynamic Seeding of Perfusing Human Umbilical Vein Endothelial Cells (HUVECs) onto Dual-Function Cell Adhesion Ligands: Arg-Gly-Asp (RGD)-Streptavidin and Biotinylated Fibronectin

PubMed Central

Anamelechi, Charles C.; Clermont, Edward C.; Novak, Matthew T.; Reichert, William M.

2014-01-01

Surfaces decorated with high affinity ligands can be used to facilitate rapid attachment of endothelial cells; however, standard equilibrium cell detachment studies are poorly suited for assessing these initial adhesion events. Here, a dynamic seeding and cell retention method was used to examine the initial attachment of perfusing human umbilical vein endothelial cells (HUVECs) to bare Teflon-AF substrates, substrates pre-adsorbed with fibronectin alone, or substrates co-pre-adsorbed with two dual-function cell-adhesion ligands: biotinylated fibronectin (bFN) and RGD-streptavidin mutant (RGD-SA). Cell attachment was evaluated as a function of cell trypsinization (integrin digestion), surface protein formulation, and cell perfusion rate. Surfaces co-pre-adsorbed with bFN and RGD-SA showed the highest density of attached cells after 8 min of perfusion and the highest percent retention when subjected to shear flow at 60 dynes/cm2 for 2 min. Surfaces with no ligand treatment showed the lowest cell attachment and retention under flow in all cases. HUVECs trypsinized with mild 0.025% trypsin/ethylenediaminetetraacetic acid (EDTA) showed greater cell adhesion after perfusion and higher percent retention after shear flow than those trypsinized using harsher 0.05% trypsin/EDTA. The preferential affinities of the two dual-function ligands for α5β1 and αvβ3 integrins were also examined by surface plasmon resonance (SPR) spectroscopy. The dynamic cell seeding studies confirmed that the dual-function ligand system promotes HUVEC adhesion and retention at short time points when tested using a perfusion assay. SPR studies showed that the two ligands exhibited equal affinity for both α5β1 and αvβ3 integrins but that the combined ligands bound more total integrins than the two ligands tested separately. PMID:19348476

The Pseudomonas fluorescens SBW25 wrinkly spreader biofilm requires attachment factor, cellulose fibre and LPS interactions to maintain strength and integrity.

PubMed

Spiers, Andrew J; Rainey, Paul B

2005-09-01

The wrinkly spreader (WS) isolate of Pseudomonas fluorescens SBW25 forms a substantial biofilm at the air-liquid interface. The biofilm is composed of an extracellular partially acetylated cellulose-fibre matrix, and previous mutagenesis of WS with mini-Tn5 had identified both the regulatory and cellulose-biosynthetic operons. One uncharacterized WS mutant, WS-5, still expressed cellulose but produced very weak biofilms. In this work, the mini-Tn5 insertion site in WS-5 has been identified as being immediately upstream of the tol-pal operon. Like Tol-Pal mutants of other Gram-negative bacteria, WS-5 showed a "leaky-membrane" phenotype, including the serendipitous ability to utilize sucrose, increased uptake of the hydrophilic dye propidium iodide, and the loss of lipopolysaccharide (LPS) expression. WS-5 cells were altered in relative hydrophobicity, and showed poorer recruitment and maintenance in the biofilm than WS. The WS-5 biofilm was also less sensitive to chemical interference during development. However, growth rate, cellulose expression and attachment were not significantly different between WS and WS-5. Finally, WS-5 biofilms could be partially complemented with WS-4, a biofilm- and attachment-deficient mutant that expressed LPS, resulting in a mixed biofilm with significantly increased strength. These findings show that a major component of the WS air-liquid biofilm strength results from the interactions between LPS and the cellulose matrix of the biofilm--and that in the WS biofilm, cellulose fibres, attachment factor and LPS are required for biofilm development, strength and integrity.

Shape-shifting trypanosomes: Flagellar shortening followed by asymmetric division in Trypanosoma congolense from the tsetse proventriculus.

PubMed

Peacock, Lori; Kay, Christopher; Bailey, Mick; Gibson, Wendy

2018-05-01

Trypanosomatids such as Leishmania and Trypanosoma are digenetic, single-celled, parasitic flagellates that undergo complex life cycles involving morphological and metabolic changes to fit them for survival in different environments within their mammalian and insect hosts. According to current consensus, asymmetric division enables trypanosomatids to achieve the major morphological rearrangements associated with transition between developmental stages. Contrary to this view, here we show that the African trypanosome Trypanosoma congolense, an important livestock pathogen, undergoes extensive cell remodelling, involving shortening of the cell body and flagellum, during its transition from free-swimming proventricular forms to attached epimastigotes in vitro. Shortening of the flagellum was associated with accumulation of PFR1, a major constituent of the paraflagellar rod, in the mid-region of the flagellum where it was attached to the substrate. However, the PFR1 depot was not essential for attachment, as it accumulated several hours after initial attachment of proventricular trypanosomes. Detergent and CaCl2 treatment failed to dislodge attached parasites, demonstrating the robust nature of flagellar attachment to the substrate; the PFR1 depot was also unaffected by these treatments. Division of the remodelled proventricular trypanosome was asymmetric, producing a small daughter cell. Each mother cell went on to produce at least one more daughter cell, while the daughter trypanosomes also proliferated, eventually resulting in a dense culture of epimastigotes. Here, by observing the synchronous development of the homogeneous population of trypanosomes in the tsetse proventriculus, we have been able to examine the transition from proventricular forms to attached epimastigotes in detail in T. congolense. This transition is difficult to observe in vivo as it happens inside the mouthparts of the tsetse fly. In T. brucei, this transition is achieved by asymmetric division of long trypomastigotes in the proventriculus, yielding short epimastigotes, which go on to colonise the salivary glands. Thus, despite their close evolutionary relationship and shared developmental route within the vector, T. brucei and T. congolense have evolved different ways of accomplishing the same developmental transition from proventricular form to attached epimastigote.

The lutheran/basal cell adhesion molecule promotes tumor cell migration by modulating integrin-mediated cell attachment to laminin-511 protein.

PubMed

Kikkawa, Yamato; Ogawa, Takaho; Sudo, Ryo; Yamada, Yuji; Katagiri, Fumihiko; Hozumi, Kentaro; Nomizu, Motoyoshi; Miner, Jeffrey H

2013-10-25

Cell-matrix interactions are critical for tumor cell migration. Lutheran (Lu), also known as basal cell adhesion molecule (B-CAM), competes with integrins for binding to laminin α5, a subunit of LM-511, a major component of basement membranes. Here we show that the preferential binding of Lu/B-CAM to laminin α5 promotes tumor cell migration. The attachment of Lu/B-CAM transfectants to LM-511 was slightly weaker than that of control cells, and this was because Lu/B-CAM disturbed integrin binding to laminin α5. Lu/B-CAM induced a spindle cell shape with pseudopods and promoted cell migration on LM-511. In addition, blocking with an anti-Lu/B-CAM antibody led to a flat cell shape and inhibited migration on LM-511, similar to the effects of an activating integrin β1 antibody. We conclude that tumor cell migration on LM-511 requires that Lu/B-CAM competitively modulates cell attachment through integrins. We suggest that this competitive interaction is involved in a balance between static and migratory cell behaviors.

Poly(lactic- co-glycolic) acid nanoparticles uptake by Vitis vinifera and grapevine-pathogenic fungi

NASA Astrophysics Data System (ADS)

Valletta, Alessio; Chronopoulou, Laura; Palocci, Cleofe; Baldan, Barbara; Donati, Livia; Pasqua, Gabriella

2014-12-01

Poly(lactic- co-glycolic) acid (PLGA)-based NPs are currently considered among the most promising drug carriers, nevertheless their use in plants has never been investigated. In this work, for the first time, we demonstrated the ability of PLGA NPs to cross the plant cell wall and membrane of Vitis vinifera cell cultures and grapevine-pathogenic fungi. By means of fluorescence microscopy, we established that PLGA NPs can enter in grapevine leaf tissues through stomata openings and that they can be absorbed by the roots and transported to the shoot through vascular tissues. TEM analysis on cultured cells showed that NPs ≤ 50 nm could enter cells, while bigger ones remained attached to the cell wall. Viability tests demonstrated that PLGA NPs were not cytotoxic for V. vinifera-cultured cells. The cellular uptake of PLGA NPs by some important grapevine-pathogenic fungi has also been observed, thus suggesting that PLGA NPs could be used to deliver antifungal compounds within fungal cells. Overall the results reported suggest that such NPs may play a key role in future developments of agrobiotechnologies, as it is currently happening in biomedicine.

Holographic lithography for biomedical applications

NASA Astrophysics Data System (ADS)

Stankevicius, E.; Balciunas, E.; Malinauskas, M.; Raciukaitis, G.; Baltriukiene, D.; Bukelskiene, V.

2012-06-01

Fabrication of scaffolds for cell growth with appropriate mechanical characteristics is top-most important for successful creation of tissue. Due to ability of fast fabrication of periodic structures with a different period, the holographic lithography technique is a suitable tool for scaffolds fabrication. The scaffolds fabricated by holographic lithography can be used in various biomedical investigations such as the cellular adhesion, proliferation and viability. These investigations allow selection of the suitable material and geometry of scaffolds which can be used in creation of tissue. Scaffolds fabricated from di-acrylated poly(ethylene glycol) (PEG-DA-258) over a large area by holographic lithography technique are presented in this paper. The PEG-DA scaffolds fabricated by holographic lithography showed good cytocompatibility for rabbit myogenic stem cells. It was observed that adult rabbit muscle-derived myogenic stem cells grew onto PEG-DA scaffolds. They were attached to the pillars and formed cell-cell interactions. It demonstrates that the fabricated structures have potential to be an interconnection channel network for cell-to-cell interactions, flow transport of nutrients and metabolic waste as well as vascular capillary ingrowth. These results are encouraging for further development of holographic lithography by improving its efficiency for microstructuring three-dimensional scaffolds out of biodegradable hydrogels

Haemophilus parasuis serovar 5 Nagasaki strain adheres and invades PK-15 cells.

PubMed

Frandoloso, Rafael; Martínez-Martínez, Sonia; Gutiérrez-Martín, César B; Rodríguez-Ferri, Elías F

2012-01-27

Haemophilus parasuis is the agent responsible for causing Glässer's disease, which is characterized by fibrinous polyserositis, polyarthritis and meningitis in pigs. The purpose of this study was to investigate the in vitro ability of two H. parasuis serovars of different virulence (serovar 5, Nagasaki strain, highly virulent, belonging to serovar 5, and SW114 strain, nonvirulent, belonging to serovar 3) to adhere to and invade porcine kidney epithelial cells (PK-15 line). Nagasaki strain was able to attach at high levels from 60 to 180 min of incubation irrespective of the concentrations compared (10(7)-10(10)CFU), and a substantial increase of surface projections could be seen in PK-15 cells by scanning electron microscopy. This virulent strain was also able to invade effectively these epithelial cells, and the highest invasion capacity was reached at 180 min of infection. On the contrary, nonvirulent SW114 strain hardly adhered to PK-15 cells, and it did not invade these cells, thus suggesting that adherence and invasion of porcine kidney epithelial cells could be a virulence mechanism involved in the lesions caused by H. parasuis Nagasaki strain in this organ. Copyright © 2011 Elsevier B.V. All rights reserved.

Bst1 is required for Candida albicans infecting host via facilitating cell wall anchorage of Glycosylphosphatidyl inositol anchored proteins

PubMed Central

Liu, Wei; Zou, Zui; Huang, Xin; Shen, Hui; He, Li Juan; Chen, Si Min; Li, Li Ping; Yan, Lan; Zhang, Shi Qun; Zhang, Jun Dong; Xu, Zheng; Xu, Guo Tong; An, Mao Mao; Jiang, Yuan Ying

2016-01-01

Glycosylphosphatidyl inositol anchored proteins (GPI-APs) on fungal cell wall are essential for invasive infections. While the function of inositol deacylation of GPI-APs in mammalian cells has been previously characterized the impact of inositol deacylation in fungi and implications to host infection remains largely unexplored. Herein we describe our identification of BST1, an inositol deacylase of GPI-Aps in Candida albicans, was critical for GPI-APs cell wall attachment and host infection. BST1-deficient C. albicans (bst1Δ/Δ) was associated with severely impaired cell wall anchorage of GPI-APs and subsequen unmasked β-(1,3)-glucan. Consistent with the aberrant cell wall structures, bst1Δ/Δ strain did not display an invasive ability and could be recognized more efficiently by host immune systems. Moreover, BST1 null mutants or those expressing Bst1 variants did not display inositol deacylation activity and exhibited severely attenuated virulence and reduced organic colonization in a murine systemic candidiasis model. Thus, Bst1 can facilitate cell wall anchorage of GPI-APs in C. albicans by inositol deacylation, and is critical for host invasion and immune escape. PMID:27708385

Intimate Partner Violence and Children’s Attachment Representations during Middle Childhood

PubMed Central

Gustafsson, Hanna C.; Brown, Geoffrey L.; Mills-Koonce, W. Roger; Cox, Martha. J.

2016-01-01

Despite long-standing hypotheses that intimate partner violence (IPV) may undermine children’s ability to form secure attachment representations, few studies have empirically investigated this association. Particularly lacking is research that examines IPV and attachment during middle childhood, a time when the way that children understand, represent, and process the behavior of others becomes particularly important. Using data from a sample of African American children living in rural, low-income communities (n = 98), the current study sought to address this gap by examining the association between physical IPV occurring early in children’s lives and their attachment security during the first grade. Results indicate that, even after controlling for child- and family-level covariates, physical IPV was associated with a greater likelihood of being rated insecurely attached. This effect was above and beyond the influence of maternal parenting behaviors, demonstrating a unique effect of physical IPV on children’s attachment representations during middle childhood. PMID:28781383

Intimate Partner Violence and Children's Attachment Representations during Middle Childhood.

PubMed

Gustafsson, Hanna C; Brown, Geoffrey L; Mills-Koonce, W Roger; Cox, Martha J

2017-06-01

Despite long-standing hypotheses that intimate partner violence (IPV) may undermine children's ability to form secure attachment representations, few studies have empirically investigated this association. Particularly lacking is research that examines IPV and attachment during middle childhood, a time when the way that children understand, represent, and process the behavior of others becomes particularly important. Using data from a sample of African American children living in rural, low-income communities ( n = 98), the current study sought to address this gap by examining the association between physical IPV occurring early in children's lives and their attachment security during the first grade. Results indicate that, even after controlling for child- and family-level covariates, physical IPV was associated with a greater likelihood of being rated insecurely attached. This effect was above and beyond the influence of maternal parenting behaviors, demonstrating a unique effect of physical IPV on children's attachment representations during middle childhood.

Quantification of Agrobacterium tumefaciens C58 attachment to Arabidopsis thaliana roots.

PubMed

Petrovicheva, Anna; Joyner, Jessica; Muth, Theodore R

2017-10-02

Agrobacterium tumefaciens is the causal agent of crown gall disease and is a vector for DNA transfer in transgenic plants. The transformation process by A. tumefaciens has been widely studied, but the attachment stage has not been well characterized. Most measurements of attachment have used microscopy and colony counting, both of which are labor and time intensive. To reduce the time and effort required to analyze bacteria attaching to plant tissues, we developed a quantitative real-time PCR (qPCR) assay to quantify attached A. tumefaciens using the chvE gene as marker for the presence of the bacteria. The qPCR detection threshold of A. tumefaciens from pure culture was 104 cell equivalents/ml. The A. tumefaciens minimum threshold concentration from root-bound populations was determined to be 105 cell equivalents/ml inoculum to detect attachment above background. The qPCR assay can be used for measuring A. tumefaciens attachment in applications such as testing the effects of mutations on bacterial adhesion molecules or biofilm formation, comparing attachment across various plant species and ecotypes, and detecting mutations in putative attachment receptors expressed in plant roots. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Adhesion of glucosyltransferase phase variants to Streptococcus gordonii bacterium-glucan substrata may involve lipoteichoic acid.

PubMed

Vickerman, M M; Jones, G W

1992-10-01

Growing Streptococcus gordonii Spp+ phase variants, which have normal levels of glucosyltransferase (GTF) activity, use sucrose to promote their accumulation on surfaces by forming a cohesive bacterium-insoluble glucan polymer mass (BPM). Spp- phase variants, which have lower levels of GTF activity, do not form BPMs and do not remain in BPMs formed by Spp+ cells when grown in mixed cultures. To test the hypothesis that segregation of attached Spp+ and unattached Spp- cells was due to differences in adhesiveness, adhesion between washed, [3H]thymidine-labeled cells and preformed BPM substrata was measured. Unexpectedly, the results showed that cells of both phenotypes, as well as GTF-negative cells, attached equally well to preformed BPMs, indicating that attachment to BPMs was independent of cell surface GTF activity. Initial characterization of this binding interaction suggested that a protease-sensitive component on the washed cells may be binding to lipoteichoic acids sequestered in the BPM, since exogenous lipoteichoic acid inhibited adhesion. Surprisingly, the adhesion of both Spp+ and Spp- cells was markedly inhibited in the presence of sucrose, which also released lipoteichoic acid from the BPM. These in vitro findings suggest that, in vivo, sucrose and lipoteichoic acid may modify dental plaque development by enhancing or inhibiting the attachment of additional bacteria.

Adhesion of glucosyltransferase phase variants to Streptococcus gordonii bacterium-glucan substrata may involve lipoteichoic acid.

PubMed Central

Vickerman, M M; Jones, G W

1992-01-01

Growing Streptococcus gordonii Spp+ phase variants, which have normal levels of glucosyltransferase (GTF) activity, use sucrose to promote their accumulation on surfaces by forming a cohesive bacterium-insoluble glucan polymer mass (BPM). Spp- phase variants, which have lower levels of GTF activity, do not form BPMs and do not remain in BPMs formed by Spp+ cells when grown in mixed cultures. To test the hypothesis that segregation of attached Spp+ and unattached Spp- cells was due to differences in adhesiveness, adhesion between washed, [3H]thymidine-labeled cells and preformed BPM substrata was measured. Unexpectedly, the results showed that cells of both phenotypes, as well as GTF-negative cells, attached equally well to preformed BPMs, indicating that attachment to BPMs was independent of cell surface GTF activity. Initial characterization of this binding interaction suggested that a protease-sensitive component on the washed cells may be binding to lipoteichoic acids sequestered in the BPM, since exogenous lipoteichoic acid inhibited adhesion. Surprisingly, the adhesion of both Spp+ and Spp- cells was markedly inhibited in the presence of sucrose, which also released lipoteichoic acid from the BPM. These in vitro findings suggest that, in vivo, sucrose and lipoteichoic acid may modify dental plaque development by enhancing or inhibiting the attachment of additional bacteria. PMID:1398940

Integrin Expression Regulates Neuroblastoma Attachment and Migration1

PubMed Central

Meyer, Amy; van Golen, Cynthia M.; Kim, Bhumsoo; van Golen, Kenneth L.; Feldman, Eva L.

2004-01-01

Abstract Neuroblastoma (NBL) is the most common malignant disease of infancy, and children with bone metastasis have a mortality rate greater than 90%. Two major classes of proteins, integrins and growth factors, regulate the metastatic process. We have previously shown that tumorigenic NBL cells express higher levels of the type I insulin-like growth factor receptor (IGF-IR) and that β1 integrin expression is inversely proportional to tumorigenic potential in NBL. In the current study, we analyze the effect of β1 integrin and IGF-IR on NBL cell attachment and migration. Nontumorigenic S-cells express high levels of β1 integrin, whereas tumorigenic N-cells express little β1 integrin. Alterations in β1 integrin are due to regulation at the protein level, as translation is decreased in N-type cells. Moreover, inhibition of protein synthesis shows that β1 integrin is degraded more slowly in S-type cells (SHEP) than in N-type cells (SH-SY5Y and IMR32). Inhibition of α5β1 integrin prevents SHEP (but not SH-SY5Y or IMR32) cell attachment to fibronectin and increases SHEP cell migration. Increases in IGF-IR decrease β1 integrin expression, and enhance SHEP cell migration, potentially through increased expression of αvβ3. These data suggest that specific classes of integrins in concert with IGF-IR regulate NBL attachment and migration. PMID:15256055

Mechanistic Aspects of Adenovirus Serotype 2 Inactivation with Free Chlorine ▿ †

PubMed Central

Page, Martin A.; Shisler, Joanna L.; Mariñas, Benito J.

2010-01-01

Free chlorine is an effective disinfectant for controlling adenoviruses in drinking water, but little is known about the underlying inactivation mechanisms. The objective of this study was to elucidate the molecular components of adenovirus type 2 (Ad2) targeted by free chlorine during the inactivation process. The effects of free chlorine treatment on several Ad2 molecular components and associated life cycle events were compared to its effect on the ability of adenovirus to complete its life cycle, i.e., viability. Free chlorine treatment of Ad2 virions did not impair their ability to interact with monoclonal antibodies specific for hexon and fiber proteins of the Ad2 capsid, as measured by enzyme-linked immunosorbent assays, nor did it impair their interaction with recombinant, purified Coxsackie-adenovirus receptor (CAR) proteins in vitro. Free chlorine-treated Ad2 virions also retained their ability to bind to CAR receptors on A549 cell monolayers, despite being unable to form plaques, suggesting that free chlorine inactivates Ad2 by inhibiting a postbinding event of the Ad2 life cycle. DNA isolated from Ad2 virions that had been inactivated by free chlorine was able to be amplified by PCR, indicating that genome damage was not the cause of inactivation. However, inactivated Ad2 virions were unable to express E1A viral proteins during infection of A549 host cells, as measured by using immunoblotting. Collectively, these results indicate that free chlorine inactivates adenovirus by damaging proteins that govern life cycle processes occurring after host cell attachment, such as endocytosis, endosomal lysis, or nuclear delivery. PMID:20305026

Investigating the Influence of the Initial Biomass Distribution and Injection Strategies on Biofilm-Mediated Calcite Precipitation in Porous Media

DOE PAGES

Hommel, Johannes; Lauchnor, Ellen; Gerlach, Robin; ...

2015-12-16

Attachment of bacteria in porous media is a complex mixture of processes resulting in the transfer and immobilization of suspended cells onto a solid surface within the porous medium. However, quantifying the rate of attachment is difficult due to the many simultaneous processes possibly involved in attachment, including straining, sorption, and sedimentation, and the difficulties in measuring metabolically active cells attached to porous media. Preliminary experiments confirmed the difficulty associated with measuring active Sporosarcina pasteurii cells attached to porous media. However, attachment is a key process in applications of biofilm-mediated reactions in the subsurface such as microbially induced calcite precipitation.more » Independent of the exact processes involved, attachment determines both the distribution and the initial amount of attached biomass and as such the initial reaction rate. As direct experimental investigations are difficult, this study is limited to a numerical investigation of the effect of various initial biomass distributions and initial amounts of attached biomass. This is performed for various injection strategies, changing the injection rate as well as alternating between continuous and pulsed injections. The results of this study indicate that, for the selected scenarios, both the initial amount and the distribution of attached biomass have minor influence on the Ca 2+ precipitation efficiency as well as the distribution of the precipitates compared to the influence of the injection strategy. The influence of the initial biomass distribution on the resulting final distribution of the precipitated calcite is limited, except for the continuous injection at intermediate injection rate. But even for this injection strategy, the Ca 2+ precipitation efficiency shows no significant dependence on the initial biomass distribution.« less

Investigating the Influence of the Initial Biomass Distribution and Injection Strategies on Biofilm-Mediated Calcite Precipitation in Porous Media

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

Hommel, Johannes; Lauchnor, Ellen; Gerlach, Robin

Attachment of bacteria in porous media is a complex mixture of processes resulting in the transfer and immobilization of suspended cells onto a solid surface within the porous medium. However, quantifying the rate of attachment is difficult due to the many simultaneous processes possibly involved in attachment, including straining, sorption, and sedimentation, and the difficulties in measuring metabolically active cells attached to porous media. Preliminary experiments confirmed the difficulty associated with measuring active Sporosarcina pasteurii cells attached to porous media. However, attachment is a key process in applications of biofilm-mediated reactions in the subsurface such as microbially induced calcite precipitation.more » Independent of the exact processes involved, attachment determines both the distribution and the initial amount of attached biomass and as such the initial reaction rate. As direct experimental investigations are difficult, this study is limited to a numerical investigation of the effect of various initial biomass distributions and initial amounts of attached biomass. This is performed for various injection strategies, changing the injection rate as well as alternating between continuous and pulsed injections. The results of this study indicate that, for the selected scenarios, both the initial amount and the distribution of attached biomass have minor influence on the Ca 2+ precipitation efficiency as well as the distribution of the precipitates compared to the influence of the injection strategy. The influence of the initial biomass distribution on the resulting final distribution of the precipitated calcite is limited, except for the continuous injection at intermediate injection rate. But even for this injection strategy, the Ca 2+ precipitation efficiency shows no significant dependence on the initial biomass distribution.« less

Effects of atmospheric air plasma treatment of graphite and carbon felt electrodes on the anodic current from Shewanella attached cells.

PubMed

Epifanio, Monica; Inguva, Saikumar; Kitching, Michael; Mosnier, Jean-Paul; Marsili, Enrico

2015-12-01

The attachment of electrochemically active microorganisms (EAM) on an electrode is determined by both the chemistry and topography of the electrode surface. Pre-treatment of the electrode surface by atmospheric air plasma introduces hydrophilic functional groups, thereby increasing cell attachment and electroactivity in short-term experiments. In this study, we use graphite and carbon felt electrodes to grow the model EAM Shewanella loihica PV-4 at oxidative potential (0.2 V vs. Ag/AgCl). Cell attachment and electroactivity are measured through electrodynamic methods. Atmospheric air plasma pre-treatment increases cell attachment and current output at graphite electrodes by 25%, while it improves the electroactivity of the carbon felt electrodes by 450%. Air plasma pre-treatment decreased the coulombic efficiency on both carbon felt and graphite electrodes by 60% and 80%, respectively. Microbially produced flavins adsorb preferentially at the graphite electrode, and air plasma pre-treatment results in lower flavin adsorption at both graphite and carbon felt electrodes. Results show that air plasma pre-treatment is a feasible option to increase current output in bioelectrochemical systems. Copyright © 2015 Elsevier B.V. All rights reserved.

Combined copper/zinc attachment to prion protein

NASA Astrophysics Data System (ADS)

Hodak, Miroslav; Bernholc, Jerry

2013-03-01

Misfolding of prion protein (PrP) is responsible for diseases such as ``mad-cow disease'' in cattle and Creutzfeldt-Jacob in humans. Extensive experimental investigation has established that this protein strongly interacts with copper ions, and this ability has been linked to its still unknown function. Attachment of other metal ions (zinc, iron, manganese) have been demonstrated as well, but none of them could outcompete copper. Recent finding, however, indicates that at intermediate concentrations both copper and zinc ions can attach to the PrP at the octarepeat region, which contains high affinity metal binding sites. Based on this evidence, we have performed density functional theory simulations to investigate the combined Cu/Zn attachment. We consider all previously reported binding modes of copper at the octarepeat region and examine a possibility simultaneous Cu/Zn attachment. We find that this can indeed occur for only one of the known binding sites, when copper changes its coordination mode to allow for attachment of zinc ion. The implications of the simultaneous attachment on neural function remain to be explored.

Cable attachment for a radioactive brachytherapy source capsule

DOEpatents

Gross, Ian G; Pierce, Larry A

2006-07-18

In cancer brachytherapy treatment, a small californium-252 neutron source capsule is attached to a guide cable using a modified crimping technique. The guide cable has a solid cylindrical end, and the attachment employs circumferential grooves micromachined in the solid cable end. The attachment was designed and tested, and hardware fabricated for use inside a radioactive hot cell. A welding step typically required in other cable attachments is avoided.

High-Temperature Strain Sensing for Aerospace Applications

NASA Technical Reports Server (NTRS)

Piazza, Anthony; Richards, Lance W.; Hudson, Larry D.

2008-01-01

Thermal protection systems (TPS) and hot structures are utilizing advanced materials that operate at temperatures that exceed abilities to measure structural performance. Robust strain sensors that operate accurately and reliably beyond 1800 F are needed but do not exist. These shortcomings hinder the ability to validate analysis and modeling techniques and hinders the ability to optimize structural designs. This presentation examines high-temperature strain sensing for aerospace applications and, more specifically, seeks to provide strain data for validating finite element models and thermal-structural analyses. Efforts have been made to develop sensor attachment techniques for relevant structural materials at the small test specimen level and to perform laboratory tests to characterize sensor and generate corrections to apply to indicated strains. Areas highlighted in this presentation include sensors, sensor attachment techniques, laboratory evaluation/characterization of strain measurement, and sensor use in large-scale structures.

Watch Out for the “Living Dead”: Cell-Free Enzymes and Their Fate

PubMed Central

Baltar, Federico

2018-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the “gatekeepers” of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell’s fate. In contrast, cell-free enzymes belong to a kind of “living dead” realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go “beyond the living things,” studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles. PMID:29354095

Chk1 and Mps1 jointly regulate correction of merotelic kinetochore attachments.

PubMed

Petsalaki, Eleni; Zachos, George

2013-03-01

If uncorrected, merotelic kinetochore attachments can induce mis-segregated chromosomes in anaphase. We show that checkpoint kinase 1 (Chk1) protects vertebrate cells against merotelic attachments and lagging chromosomes and is required for correction of merotelic attachments during a prolonged metaphase. Decreased Chk1 activity leads to hyper-stable kinetochore microtubules, unstable binding of MCAK, Kif2b and Mps1 to centromeres or kinetochores and reduced phosphorylation of Hec1 by Aurora-B. Phosphorylation of Aurora-B at serine 331 (Ser331) by Chk1 is high in prometaphase and decreases significantly in metaphase cells. We propose that Ser331 phosphorylation is required for optimal localization of MCAK, Kif2b and Mps1 to centromeres or kinetochores and for Hec1 phosphorylation. Furthermore, inhibition of Mps1 activity diminishes initial recruitment of MCAK and Kif2b to centromeres or kinetochores, impairs Hec1 phosphorylation and exacerbates merotelic attachments in Chk1-deficient cells. We propose that Chk1 and Mps1 jointly regulate Aurora-B, MCAK, Kif2b and Hec1 to correct merotelic attachments. These results suggest a role for Chk1 and Mps1 in error correction.

Attached and Unattached Bacterial Communities in a 120-Meter Corehole in an Acidic, Crystalline Rock Aquifer

PubMed Central

Lehman, R. Michael; Roberto, Francisco F.; Earley, Drummond; Bruhn, Debby F.; Brink, Susan E.; O'Connell, Sean P.; Delwiche, Mark E.; Colwell, Frederick S.

2001-01-01

The bacteria colonizing geologic core sections (attached) were contrasted with those found suspended in the groundwater (unattached) by examining the microbiology of 16 depth-paired core and groundwater samples using a suite of culture-independent and culture-dependent analyses. One hundred twenty-two meters was continuously cored from a buried chalcopyrite ore hosted in a biotite-quartz-monzonite porphyry at the Mineral Park Mine near Kingman, Ariz. Every fourth 1.5-m core was acquired using microbiologically defensible methods, and these core sections were aseptically processed for characterization of the attached bacteria. Groundwater samples containing unattached bacteria were collected from the uncased corehole at depth intervals corresponding to the individual cores using an inflatable straddle packer sampler. The groundwater was acidic (pH 2.8 to 5.0), with low levels of dissolved oxygen and high concentrations of sulfate and metals, including ferrous iron. Total numbers of attached cells were less than 105 cells g of core material−1 while unattached cells numbered about 105 cells ml of groundwater−1. Attached and unattached acidophilic heterotrophs were observed throughout the depth profile. In contrast, acidophilic chemolithotrophs were not found attached to the rock but were commonly observed in the groundwater. Attached communities were composed of low numbers (<40 CFU g−1) of neutrophilic heterotrophs that exhibited a high degree of morphologic diversity, while unattached communities contained higher numbers (ca. 103 CFU ml−1) of neutrophilic heterotrophs of limited diversity. Sulfate-reducing bacteria were restricted to the deepest samples of both core and groundwater. 16S ribosomal DNA sequence analysis of attached, acidophilic isolates indicated that organisms closely related to heterotrophic, acidophilic mesophiles such as Acidiphilium organovorum and, surprisingly, to the moderately thermophilic Alicyclobacillus acidocaldarius were present. The results indicate that viable (but possibly inactive) microorganisms were present in the buried ore and that there was substantial distinction in biomass and physiological capabilities between attached and unattached populations. PMID:11319087

PEG attachment to osteoblasts enhances mechanosensitivity.

PubMed

Hamamura, Kazunori; Weng, Yiming; Zhao, Jun; Yokota, Hiroki; Xie, Dong

2008-06-01

Fluid flow induces proliferation and differentiation of osteoblasts, and fibrous structure like a primary cilium on a cell surface contributes to flow sensing and flow-driven gene regulation. We address a question: Does attachment of synthetic polymers on a cell surface enhance mechanosensitivity of osteoblasts? Using MC3T3 osteoblast cells (C4 clone) and a PEG polymer, one of whose termini was covalently linked to a succinimidyl succinate group (functionalized PEG-PEGSS), we examined attachment of PEGSS to osteoblasts and evaluated its effects on the mRNA expression of stress-responsive genes. AFM images exhibited globular PEGSS conformation of approximately 100 nm in size, and SEM images confirmed the attachment of a cluster of pancake-like PEGSS molecules on the osteoblast surface. Compared to control cells incubated with unfunctionalized PEG, real-time PCR revealed that RNA upregulation of c-fos, egr1, ATF3 and Cox2 genes was magnified in the cells incubated with PEGSS. These results support a PEG-induced increase in mechanosensitivity of osteoblasts and indicate that the described approach would be useful to accelerate growth and development of osteoblasts for bone repair and tissue engineering.

The Effect of Ag and Ag+N Ion Implantation on Cell Attachment Properties

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

Urkac, Emel Sokullu; Oztarhan, Ahmet; Gurhan, Ismet Deliloglu

2009-03-10

Implanted biomedical prosthetic devices are intended to perform safely, reliably and effectively in the human body thus the materials used for orthopedic devices should have good biocompatibility. Ultra High Molecular Weight Poly Ethylene (UHMWPE) has been commonly used for total hip joint replacement because of its very good properties. In this work, UHMWPE samples were Ag and Ag+N ion implanted by using the Metal-Vapor Vacuum Arc (MEVVA) ion implantation technique. Samples were implanted with a fluency of 1017 ion/cm2 and extraction voltage of 30 kV. Rutherford Backscattering Spectrometry (RBS) was used for surface studies. RBS showed the presence of Agmore » and N on the surface. Cell attachment properties investigated with model cell lines (L929 mouse fibroblasts) to demonstrate that the effect of Ag and Ag+N ion implantation can favorably influence the surface of UHMWPE for biomedical applications. Scanning electron microscopy (SEM) was used to demonstrate the cell attachment on the surface. Study has shown that Ag+N ion implantation represents more effective cell attachment properties on the UHMWPE surfaces.« less

Child Maltreatment Is Associated with a Reduction of the Oxytocin Receptor in Peripheral Blood Mononuclear Cells

PubMed Central

Krause, Sabrina; Boeck, Christina; Gumpp, Anja M.; Rottler, Edit; Schury, Katharina; Karabatsiakis, Alexander; Buchheim, Anna; Gündel, Harald; Kolassa, Iris-Tatjana; Waller, Christiane

2018-01-01

Background: Child maltreatment (CM) and attachment experiences are closely linked to alterations in the human oxytocin (OXT) system. However, human data about oxytocin receptor (OXTR) protein levels are lacking. Therefore, we investigated oxytocin receptor (OXTR) protein levels in circulating immune cells and related them to circulating levels of OXT in peripheral blood. We hypothesized reduced OXTR protein levels, associated with both, experiences of CM and an insecure attachment representation. Methods: OXTR protein expressions were analyzed by western blot analyses in peripheral blood mononuclear cells (PBMC) and plasma OXT levels were determined by radioimmunoassay (RIA) in 49 mothers. We used the Childhood Trauma Questionnaire (CTQ) to assess adverse childhood experiences. Attachment representations (secure vs. insecure) were classified using the Adult Attachment Projective Picture System (AAP) and levels of anxiety and depression were assessed with the German version of the Hospital Depression and Anxiety scale (HADS-D). Results: CM-affected women showed significantly lower OXTR protein expression with significantly negative correlations between the OXTR protein expression and the CTQ sum score, whereas plasma OXT levels showed no significant differences in association with CM. Lower OXTR protein expression in PBMC were particularly pronounced in the group of insecurely attached mothers compared to the securely attached group. Anxiety levels were significantly higher in CM-affected women. Conclusion: This study demonstrated a significant association between CM and an alteration of OXTR protein expression in human blood cells as a sign for chronic, long-lasting alterations in this attachment-related neurobiological system. PMID:29535656

β1 Integrins Mediate Attachment of Mesenchymal Stem Cells to Cartilage Lesions

PubMed Central

Zwolanek, Daniela; Flicker, Magdalena; Kirstätter, Elisabeth; Zaucke, Frank; van Osch, Gerjo J.V.M.; Erben, Reinhold G.

2015-01-01

Abstract Mesenchymal stem cells (MSC) may have great potential for cell-based therapies of osteoarthritis. However, after injection in the joint, only few cells adhere to defective articular cartilage and contribute to cartilage regeneration. Little is known about the molecular mechanisms of MSC attachment to defective articular cartilage. Here, we developed an ex vivo attachment system, using rat osteochondral explants with artificially created full-thickness cartilage defects in combination with genetically labeled MSC isolated from bone marrow of human placental alkaline phosphatase transgenic rats. Binding of MSC to full-thickness cartilage lesions was improved by serum, but not hyaluronic acid, and was dependent on the presence of divalent cations. Additional in vitro tests showed that rat MSC attach, in a divalent cation-dependent manner, to collagen I, collagen II, and fibronectin, but not to collagen XXII or cartilage oligomeric matrix protein (COMP). RGD peptides partially blocked the adhesion of MSC to fibronectin in vitro and to cartilage lesions ex vivo. Furthermore, the attachment of MSC to collagen I and II in vitro and to cartilage lesions ex vivo was almost completely abolished in the presence of a β1 integrin blocking antibody. In conclusion, our data suggest that attachment of MSC to ex vivo full-thickness cartilage lesions is almost entirely β1 integrin-mediated, whereby both RGD- and collagen-binding integrins are involved. These findings suggest a key role of integrins during MSC attachment to defective cartilage and may pave the way for improved MSC-based therapies in the future. PMID:26309781

Monitoring change in refractive index of cytosol of animal cells on affinity surface under osmotic stimulus for label-free measurement of viability.

PubMed

Park, Jina; Jin, Sung Il; Kim, Hyung Min; Ahn, Junhyoung; Kim, Yeon-Gu; Lee, Eun Gyo; Kim, Min-Gon; Shin, Yong-Beom

2015-02-15

We demonstrated that a metal-clad waveguide (MCW)-based biosensor can be applied to label-free measurements of viability of adherent animal cells with osmotic stimulation in real time. After Chinese hamster ovary (CHO) and human embryonic kidney cell 293 (HEK293) cells were attached to a Concanavalin A (Con A)-modified sensor surface, the magnitudes of cell responses to non-isotonic stimulation were compared between live and dead cells. The live cells exhibited a change in the refractive index (RI) of the cytosol caused by a redistribution of water through the cell membrane, which was induced by the osmotic stimulus, but the dead cells did not. Moreover, the normalized change in the RI measured via the MCW sensor was linearly proportional to the viability of attached cells and the resolution in monitoring cell viability was about 0.079%. Therefore, the viability of attached animal cells can be measured without labels by observing the relative differences in the RI of cytosol in isotonic and non-isotonic buffers. Copyright © 2014 Elsevier B.V. All rights reserved.

Effect of uv light on formation and synthetic capacity of DNA-membrane complexes from T7-infected cells of E. coli

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

Wintermantel, G.

1974-02-01

The action of uv light on the attachment of T7-DNA to the bacterial cell membrane, and the RNA- and DNA-synthesizing activity of the DNA-membrane complex, were investigated. In E. coli H560 cells infected by uv-irradiated T7 phages, the amount of membrane-associated parental /sup 32/P-labeled T7-DNA was determined. To measure the RNA- and DNA-synthesizing activity, the isolated DNA- membrane complexes were incubated with an equimolar mixture of the corresponding four nucleoside triphosphates. The uv-sensitivities of the functions tested were calculated from the slopes of the dose-effect curves. In relation to the uv- sensitivity for plaque-forming ability of the T7bacteriophages they amountmore » to 0.07 ( plus or minus 0.01); 0.15 ( plus or minus 0.01); and 0.56 ( plus or minus 0.04), respectively. (auth)« less

The Candida albicans Hwp2p can complement the lack of filamentation of a Saccharomyces cerevisiae flo11 null strain.

PubMed

Younes, Samer S; Khalaf, Roy A

2013-06-01

The opportunistic fungal pathogen Candida albicans is one of the leading agents of life-threatening infections affecting immunocompromised individuals. Many factors make C. albicans a successful pathogen. These include the ability to switch between yeast and invasive hyphal morphologies in addition to an arsenal of cell wall virulence factors such as lipases, proteases, dismutases and adhesins that promote the attachment to the host, a prerequisite for invasive growth. We have previously characterized Hwp2, a C. albicans cell wall protein which we found necessary for proper oxidative stress, biofilm formation and adhesion to host cells. Baker's yeast Saccharomyces cerevisiae also possesses adhesins that promote aggregation and flocculence. Flo11 is one such adhesin that has sequence similarity to Hwp2. Here we determined that transforming an HWP2 cassette can complement the lack of filamentation of an S. cerevisiae flo11 null strain and impart on S. cerevisiae adhesive properties similar to those of a pathogen.

Click-electron microscopy for imaging metabolically tagged non-protein biomolecules

PubMed Central

Ngo, John T.; Adams, Stephen R.; Deerinck, Thomas J.; Boassa, Daniela; Rodriguez-Rivera, Frances; Palida, Sakina F.; Bertozzi, Carolyn R.; Ellisman, Mark H.; Tsien, Roger Y.

2016-01-01

Electron microscopy (EM) has long been the main technique to image cell structures with nanometer resolution, but has lagged behind light microscopy in the crucial ability to make specific molecules stand out. Here we introduce “Click-EM,” a labeling technique for correlative light microscopy and EM imaging of non-protein biomolecules. In this approach, metabolic labeling substrates containing bioorthogonal functional groups are provided to cells for incorporation into biopolymers by endogenous biosynthetic machinery. The unique chemical functionality of these analogs is exploited for selective attachment of singlet oxygen-generating fluorescent dyes via bioorthogonal “click chemistry” ligations. Illumination of dye-labeled structures generates singlet oxygen to locally catalyze the polymerization of diaminobenzidine into an osmiophilic reaction product that is readily imaged by EM. We describe the application of Click-EM in imaging metabolically tagged DNA, RNA, and lipids in cultured cells and neurons, and highlight its use in tracking peptidoglycan synthesis in the Gram-positive bacterium Listeria monocytogenes. PMID:27110681

Increased Chain Length Promotes Pneumococcal Adherence and Colonization

PubMed Central

Rodriguez, Jesse L.; Dalia, Ankur B.

2012-01-01

Streptococcus pneumoniae is a mucosal pathogen that grows in chains of variable lengths. Short-chain forms are less likely to activate complement, and as a consequence they evade opsonophagocytic clearance more effectively during invasive disease. When grown in human nasal airway surface fluid, pneumococci exhibited both short- and long-chain forms. Here, we determined whether longer chains provide an advantage during colonization when the organism is attached to the epithelial surface. Chain-forming mutants and the parental strain grown under conditions to promote chain formation showed increased adherence to human epithelial cells (A549 cells) in vitro. Additionally, adherence to A549 cells selected for longer chains within the wild-type strain. In vivo in a murine model of colonization, chain-forming mutants outcompeted the parental strain. Together, our results demonstrate that morphological heterogeneity in the pneumococcus may promote colonization of the upper respiratory tract by enhancing the ability of the organism to bind to the epithelial surface. PMID:22825449

Inhibition of Delta-induced Notch signaling using fucose analogs

PubMed Central

Schneider, Michael; Kumar, Vivek; Nordstrøm, Lars Ulrik; Feng, Lei; Takeuchi, Hideyuki; Hao, Huilin; Luca, Vincent C.; Garcia, K. Christopher; Stanley, Pamela; Wu, Peng; Haltiwanger, Robert S.

2017-01-01

Notch is a cell-surface receptor that controls cell fate decisions and is regulated by O-glycans attached to epidermal growth factor-like (EGF) repeats in its extracellular domain. Protein O-fucosyltransferase 1 (Pofut1) modifies EGF repeats with O-fucose and is essential for Notch signaling. Constitutive activation of Notch signaling has been associated with a variety of human malignancies. Therefore, tools for inhibiting Notch activity are being developed as cancer therapeutics. Towards this end, we screened L-fucose analogs for their effects on Notch signaling. Two analogs, 6-alkynyl and 6-alkenyl fucose, were substrates of Pofut1 and were incorporated directly into Notch EGF repeats in cells. Both analogs were potent inhibitors of binding to and activation of Notch1 by Notch ligands Dll1 and Dll4, but not by Jag1. Mutagenesis and modeling studies suggest that incorporation of the analogs into EGF8 of Notch1 markedly reduces the ability of Delta ligands to bind and activate Notch1. PMID:29176671

Chlamydia trachomatis-host cell interactions: role of the chlamydial major outer membrane protein as an adhesin.

PubMed Central

Su, H; Watkins, N G; Zhang, Y X; Caldwell, H D

1990-01-01

The major outer membrane protein (MOMP) of Chlamydia trachomatis is characterized by four symmetrically spaced variable domains (VDs I to IV) whose sequences vary among serotypes. The surface-exposed portions of these VDs contain contiguous sequences that are both serotyping determinants and in vivo target sites for neutralizing antibodies. Previous studies using surface proteolysis of C. trachomatis B implicated VDs II and IV of the MOMP of this serotype in the attachment of chlamydiae to host cells. In this study, we used monoclonal antibodies (MAbs) specific to antigenic determinants located in VDs II and IV of the MOMP of serotype B to further investigate the role of the MOMP in the attachment of chlamydiae to host cells. MABs specific to serotype- and subspecies-specific epitopes located in exposed VDs II and IV, respectively, neutralized chlamydial infectivity for hamster kidney cells by blocking chlamydial attachment. We radioiodinated these MAbs and used them to determine the number and topology of the surface-exposed VDs II and IV epitopes on chlamydial elementary bodies. VDs II and IV each comprised approximately 2.86 x 10(4) negatively charged sites and were in proximity on the chlamydial cell surface. These studies suggest that the MAbs blocked chlamydial attachment by inhibiting electrostatic interactions with host cells. We examined the effects of thermal inactivation on both chlamydial attachment and conformation of the MOMP. Heat-inactivated chlamydiae failed to attach to host cells and exhibited a conformational change in an inaccessible invariant hydrophobic nonapeptide sequence located within VD IV of the MOMPs of C. trachomatis serotypes. These findings suggest that in addition to electrostatic interactions, a common hydrophobic component of the MOMP also contributes to the binding of chlamydiae to host cells. Thus, we propose that the MOMP functions as a chlamydial adhesin by promoting nonspecific (electrostatic and hydrophobic) interactions with host cells. Surface-accessible negatively charged VDs appear to be important in electrostatic binding, while the invariant region of VD IV may provide a subsurface hydrophobic depression which further promotes binding of chlamydiae to host cells through hydrophobic interactions. Images PMID:2318528

Possession attachment predicts cell phone use while driving.

PubMed

Weller, Joshua A; Shackleford, Crystal; Dieckmann, Nathan; Slovic, Paul

2013-04-01

Distracted driving has become an important public health concern. However, little is known about the predictors of this health-risking behavior. One overlooked risk factor for distracted driving is the perceived attachment that one feels toward his or her phone. Prior research has suggested that individuals develop bonds toward objects, and qualitative research suggests that the bond between young drivers and their phones can be strong. It follows that individuals who perceive a strong attachment to their phone would be more likely to use it, even when driving. In a nationally representative sample of young drivers (17-28 years), participants (n = 1,006) completed a survey about driving behaviors and phone use. Risk perception surrounding cell phone use while driving and perceived attachment to one's phone were assessed by administering factor-analytically derived scales that were created as part of a larger project. Attachment toward one's phone predicted the proportion of trips in which a participant reported using their cell phone while driving, beyond that accounted for by risk perception and overall phone use. Further, attachment predicted self-reported distracted driving behaviors, such as the use of social media while driving. Attachment to one's phone may be an important but overlooked risk factor for the engagement of potentially health-risking driving behaviors. Understanding that phone attachment may adversely affect driving behaviors has the potential to inform prevention and intervention efforts designed to reduce distracted driving behaviors, especially in young drivers. 2013 APA, all rights reserved

Postpartum bonding: the impact of maternal depression and adult attachment style.

PubMed

Nonnenmacher, N; Noe, D; Ehrenthal, J C; Reck, C

2016-10-01

Maternal depression poses a risk for the developing mother-infant relationship. Similarly, maternal insecure attachment styles may limit the ability to adequately connect with the newborn during the postpartum period. The aim of this study was to investigate the effect of maternal depression and insecure attachment (insecure and dual/disorganized) on maternal bonding in a sample of n = 34 women with depression according to DSM-IV and n = 59 healthy women. Maternal depression was assessed 3 to 4 months postpartum with the Structured Clinical Interview for DSM-IV Axis I Disorders (SCID-I), bonding with the Postpartum Bonding Questionnaire, and maternal attachment style with the Attachment Style Interview. Women with current and lifetime depression as well as women with dual/disorganized attachment style reported lower bonding. Explorative analysis revealed that depression partially mediated the link between dual/disorganized attachment style and bonding with a medium-sized mediation effect. The combination of maternal depression and dual/disorganized attachment style may pose a special risk constellation for the developing mother-infant bond that should be addressed in prevention and early intervention programs.

Maternal Attachment Representation and Neurophysiological Processing during the Perception of Infants’ Emotional Expressions

PubMed Central

Leyh, Rainer; Heinisch, Christine; Behringer, Johanna; Reiner, Iris; Spangler, Gottfried

2016-01-01

The perception of infant emotions is an integral part of sensitive caregiving within the mother-child relationship, a maternal ability which develops in mothers during their own attachment history. In this study we address the association between maternal attachment representation and brain activity underlying the perception of infant emotions. Event related potentials (ERPs) of 32 primiparous mothers were assessed during a three stimulus oddball task presenting negative, positive and neutral emotion expressions of infants as target, deviant or standard stimuli. Attachment representation was assessed with the Adult Attachment Interview during pregnancy. Securely attached mothers recognized emotions of infants more accurately than insecurely attached mothers. ERPs yielded amplified N170 amplitudes for insecure mothers when focusing on negative infant emotions. Secure mothers showed enlarged P3 amplitudes to target emotion expressions of infants compared to insecure mothers, especially within conditions with frequent negative infant emotions. In these conditions, P3 latencies were prolonged in insecure mothers. In summary, maternal attachment representation was found associated with brain activity during the perception of infant emotions. This further clarifies psychological mechanisms contributing to maternal sensitivity. PMID:26862743

Maternal Attachment Representation and Neurophysiological Processing during the Perception of Infants' Emotional Expressions.

PubMed

Leyh, Rainer; Heinisch, Christine; Behringer, Johanna; Reiner, Iris; Spangler, Gottfried

2016-01-01

The perception of infant emotions is an integral part of sensitive caregiving within the mother-child relationship, a maternal ability which develops in mothers during their own attachment history. In this study we address the association between maternal attachment representation and brain activity underlying the perception of infant emotions. Event related potentials (ERPs) of 32 primiparous mothers were assessed during a three stimulus oddball task presenting negative, positive and neutral emotion expressions of infants as target, deviant or standard stimuli. Attachment representation was assessed with the Adult Attachment Interview during pregnancy. Securely attached mothers recognized emotions of infants more accurately than insecurely attached mothers. ERPs yielded amplified N170 amplitudes for insecure mothers when focusing on negative infant emotions. Secure mothers showed enlarged P3 amplitudes to target emotion expressions of infants compared to insecure mothers, especially within conditions with frequent negative infant emotions. In these conditions, P3 latencies were prolonged in insecure mothers. In summary, maternal attachment representation was found associated with brain activity during the perception of infant emotions. This further clarifies psychological mechanisms contributing to maternal sensitivity.

Quantitative measurements of intercellular adhesion between a macrophage and cancer cells using a cup-attached AFM chip.

PubMed

Kim, Hyonchol; Yamagishi, Ayana; Imaizumi, Miku; Onomura, Yui; Nagasaki, Akira; Miyagi, Yohei; Okada, Tomoko; Nakamura, Chikashi

2017-07-01

Intercellular adhesion between a macrophage and cancer cells was quantitatively measured using atomic force microscopy (AFM). Cup-shaped metal hemispheres were fabricated using polystyrene particles as a template, and a cup was attached to the apex of the AFM cantilever. The cup-attached AFM chip (cup-chip) approached a murine macrophage cell (J774.2), the cell was captured on the inner concave of the cup, and picked up by withdrawing the cup-chip from the substrate. The cell-attached chip was advanced towards a murine breast cancer cell (FP10SC2), and intercellular adhesion between the two cells was quantitatively measured. To compare cell adhesion strength, the work required to separate two adhered cells (separation work) was used as a parameter. Separation work was almost 2-fold larger between a J774.2 cell and FP10SC2 cell than between J774.2 cell and three additional different cancer cells (4T1E, MAT-LyLu, and U-2OS), two FP10SC2 cells, or two J774.2 cells. FP10SC2 was established from 4T1E as a highly metastatic cell line, indicates separation work increased as the malignancy of cancer cells became higher. One possible explanation of the strong adhesion of macrophages to cancer cells observed in this study is that the measurement condition mimicked the microenvironment of tumor-associated macrophages (TAMs) in vivo, and J774.2 cells strongly expressed CD204, which is a marker of TAMs. The results of the present study, which were obtained by measuring cell adhesion strength quantitatively, indicate that the fabricated cup-chip is a useful tool for measuring intercellular adhesion easily and quantitatively. Copyright © 2017 Elsevier B.V. All rights reserved.

Dewetting Based Fabrication of Fibrous Micro-Scaffolds as Potential Injectable Cell Carriers

PubMed Central

Song, Hokyung; Yin, Liya; Chilian, William M.; Newby, Bi-min Zhang

2014-01-01

Although regenerative medicine utilizing tissue scaffolds has made enormous strides in recent years, many constraints still hamper their effectiveness. A limitation of many scaffolds is that they form surface patches, which are not particularly effective for some types of “wounds” that are deep within tissues, e.g., stroke, myocardial infarction. In this study, we reported the generation of fibrous micro-scaffolds feasible for delivering cells by injection into the tissue parenchyma. The micro-scaffolds (widths < 100 μm) were made by dewetting of poly (lactic-coglycolic acid) thin films containing parallel strips, and cells were seeded to form cell/polymer micro-constructs during or post the micro-scaffold fabrication process. Five types of cells including rat induced vascular progenitor cells were assessed for the formation of the micro-constructs. Critical factors in forming fibrous micro-scaffolds via dewetting of polymer thin films were found to be properties of polymers and supporting substrates, temperature, and proteins in the culture medium. Also, the ability of cells to attach to the micro-scaffolds was essential for forming cell/polymer micro-constructs. Both in vitro and in vivo assessments of injecting these micro-scaffolding constructs showed, as compared to free cells, enhanced cell retention at the injected site, which could lead to improved tissue engineering and regeneration. PMID:25579969

5-Fluorouracil may enrich cancer stem cells in canine mammary tumor cells in vitro.

PubMed

Zhou, Bin; Jin, Yipeng; Zhang, Di; Lin, Degui

2018-05-01

Mammary gland carcinomas are the most common neoplasms in women and unsterilized female dogs. Owing to the existence of cancer stem cells (CSCs), chemotherapy is not able to cure these types of diseases completely. A number of studies have demonstrated that CSCs are resistant to chemotherapeutic drugs, but whether canine mammary tumor cells that have acquired resistance to 5-fluorouracil (5-FU) exhibited properties of CSCs remains unknown. The aim of the present study was to investigate whether 5-fluorouracil-resistant canine mammary tumor cells exhibited properties of CSCs. CSCs were analyzed using western blot assays, ultra-low attachment sphere cultures, flow cytometry and migration (wound healing and Transwell) assays. The results indicated that, compared with parental cells, proteins associated with the Wnt/β-catenin signaling pathway and aldehyde dehydrogenase 1 were overexpressed, the number and size of spheres in the 5-FU-resistant cells were increased, the ratio of CD44 + /CD24 -/low cells was increased and the migratory ability was improved in vitro compared with the 5-FU-susceptible cells. In conclusion, stimulation with chemotherapeutic drugs including 5-FU is a good method for increasing the proportion of canine mammary tumor stem cells in vitro , which may provide further understanding of chemotherapeutic methods and CSCs.

TIM-1 Promotes Hepatitis C Virus Cell Attachment and Infection.

PubMed

Wang, Jing; Qiao, Luhua; Hou, Zhouhua; Luo, Guangxiang

2017-01-15

Human TIM and TAM family proteins were recently found to serve as phosphatidylserine (PS) receptors which promote infections by many different viruses, including dengue virus, West Nile virus, Ebola virus, Marburg virus, and Zika virus. In the present study, we provide substantial evidence demonstrating that TIM-1 is important for efficient infection by hepatitis C virus (HCV). The knockdown of TIM-1 expression significantly reduced HCV infection but not HCV RNA replication. Likewise, TIM-1 knockout in Huh-7.5 cells remarkably lowered HCV cell attachment and subsequent HCV infection. More significantly, the impairment of HCV infection in the TIM-1 knockout cells could be restored completely by ectopic expression of TIM-1 but not TIM-3 or TIM-4. Additionally, HCV infection and cell attachment were inhibited by PS but not by phosphatidylcholine (PC), demonstrating that TIM-1-mediated enhancement of HCV infection is PS dependent. The exposure of PS on the HCV envelope was confirmed by immunoprecipitation of HCV particles with a PS-specific monoclonal antibody. Collectively, these findings demonstrate that TIM-1 promotes HCV infection by serving as an attachment receptor for binding to PS exposed on the HCV envelope. TIM family proteins were recently found to enhance infections by many different viruses, including several members of the Flaviviridae family. However, their importance in HCV infection has not previously been examined experimentally. The TIM family proteins include three members in humans: TIM-1, TIM-3, and TIM-4. The findings derived from our studies demonstrate that TIM-1, but not TIM-3 or TIM-4, promotes HCV infection by functioning as an HCV attachment factor. Knockout of the TIM-1 gene resulted in a remarkable reduction of HCV cell attachment and infection. PS-containing liposomes blocked HCV cell attachment and subsequent HCV infection. HCV particles could also be precipitated with a PS-specific monoclonal antibody. These findings suggest that TIM-1 and its binding ligand, PS, may serve as novel targets for antiviral intervention. Copyright © 2017 American Society for Microbiology.

The measurement of place attachment: validity and generalizability of a psychometric approach

Treesearch

Daniel R. Williams; Jerry J. Vaske

2003-01-01

To enhance land managers’ ability to address deeper landscape meanings and place-specific symbolic values in natural resource decision making, this study evaluated the psychometric properties of a place attachment measure designed to capture the extent of emotions and feelings people have for places. Building on previous measurement efforts, this study examined the...

Caregiving Antecedents of Secure Base Script Knowledge: A Comparative Analysis of Young Adult Attachment Representations

ERIC Educational Resources Information Center

Steele, Ryan D.; Waters, Theodore E. A.; Bost, Kelly K.; Vaughn, Brian E.; Truitt, Warren; Waters, Harriet S.; Booth-LaForce, Cathryn; Roisman, Glenn I.

2014-01-01

Based on a subsample (N = 673) of the NICHD Study of Early Child Care and Youth Development (SECCYD) cohort, this article reports data from a follow-up assessment at age 18 years on the antecedents of "secure base script knowledge", as reflected in the ability to generate narratives in which attachment-related difficulties are…

Local adherent technique for transplanting mesenchymal stem cells as a potential treatment of cartilage defect.

PubMed

Koga, Hideyuki; Shimaya, Masayuki; Muneta, Takeshi; Nimura, Akimoto; Morito, Toshiyuki; Hayashi, Masaya; Suzuki, Shiro; Ju, Young-Jin; Mochizuki, Tomoyuki; Sekiya, Ichiro

2008-01-01

Current cell therapy for cartilage regeneration requires invasive procedures, periosteal coverage and scaffold use. We have developed a novel transplantation method with synovial mesenchymal stem cells (MSCs) to adhere to the cartilage defect. For ex vivo analysis in rabbits, the cartilage defect was faced upward, filled with synovial MSC suspension, and held stationary for 2.5 to 15 minutes. The number of attached cells was examined. For in vivo analysis in rabbits, an autologous synovial MSC suspension was placed on the cartilage defect, and the position was maintained for 10 minutes to adhere the cells to the defect. For the control, either the same cell suspension was injected intra-articularly or the defects were left empty. The three groups were compared macroscopically and histologically. For ex vivo analysis in humans, in addition to the similar experiment in rabbits, the expression and effects of neutralizing antibodies for adhesion molecules were examined. Ex vivo analysis in rabbits demonstrated that the number of attached cells increased in a time-dependent manner, and more than 60% of cells attached within 10 minutes. The in vivo study showed that a large number of transplanted synovial MSCs attached to the defect at 1 day, and the cartilage defect improved at 24 weeks. The histological score was consistently better than the scores of the two control groups (same cell suspension injected intra-articularly or defects left empty) at 4, 12, and 24 weeks. Ex vivo analysis in humans provided similar results to those in rabbits. Intercellular adhesion molecule 1-positive cells increased between 1 minute and 10 minutes, and neutralizing antibodies for intercellular adhesion molecule 1, vascular cell adhesion molecule 1 and activated leukocyte-cell adhesion molecule inhibited the attachment. Placing MSC suspension on the cartilage defect for 10 minutes resulted in adherence of >60% of synovial MSCs to the defect, and promoted cartilage regeneration. This adherent method makes it possible to adhere MSCs with low invasion, without periosteal coverage, and without a scaffold.

Cerebral microcirculation shear stress levels determine Neisseria meningitidis attachment sites along the blood–brain barrier

PubMed Central

Mairey, Emilie; Genovesio, Auguste; Donnadieu, Emmanuel; Bernard, Christine; Jaubert, Francis; Pinard, Elisabeth; Seylaz, Jacques; Olivo-Marin, Jean-Christophe; Nassif, Xavier; Duménil, Guillaume

2006-01-01

Neisseria meningitidis is a commensal bacterium of the human nasopharynx. Occasionally, this bacterium reaches the bloodstream and causes meningitis after crossing the blood–brain barrier by an unknown mechanism. An immunohistological study of a meningococcal sepsis case revealed that neisserial adhesion was restricted to capillaries located in low blood flow regions in the infected organs. This study led to the hypothesis that drag forces encountered by the meningococcus in the bloodstream determine its attachment site in vessels. We therefore investigated the ability of N. meningitidis to bind to endothelial cells in the presence of liquid flow mimicking the bloodstream with a laminar flow chamber. Strikingly, average blood flows reported for various organs strongly inhibited initial adhesion. As cerebral microcirculation is known to be highly heterogeneous, cerebral blood velocity was investigated at the level of individual vessels using intravital imaging of rat brain. In agreement with the histological study, shear stress levels compatible with meningococcal adhesion were only observed in capillaries, which exhibited transient reductions in flow. The flow chamber assay revealed that, after initial attachment, bacteria resisted high blood velocities and even multiplied, forming microcolonies resembling those observed in the septicemia case. These results argue that the combined mechanical properties of neisserial adhesion and blood microcirculation target meningococci to transiently underperfused cerebral capillaries and thus determine disease development. PMID:16864659

Magnetic resonance investigation of magnetic-labeled baker's yeast cells

NASA Astrophysics Data System (ADS)

Godoy Morais, J. P. M.; Azevedo, R. B.; Silva, L. P.; Lacava, Z. G. M.; Báo, S. N.; Silva, O.; Pelegrini, F.; Gansau, C.; Buske, N.; Safarik, I.; Safarikova, M.; Morais, P. C.

2004-05-01

In this study, the interaction of DMSA-coated magnetite nanoparticles (5 and 10 nm core-size) with Saccharomyces cerevisae was investigated using magnetic resonance (MR) and transmission electron microscopy (TEM). The TEM micrographs revealed magnetite nanoparticles attached externally to the cell wall. The MR data support the strong interaction among the nanoparticles supported by the cells. A remarkable shift in the resonance field was used as signature of particle attachment to the cell wall.

Effect of modified pectin molecules on the growth of bone cells.

PubMed

Kokkonen, Hanna E; Ilvesaro, Joanna M; Morra, Marco; Schols, Henk A; Tuukkanen, Juha

2007-02-01

The aim of this study was to investigate molecular candidates for bone implant nanocoatings, which could improve biocompatibility of implant materials. Primary rat bone cells and murine preosteoblastic MC3T3-E1 cells were cultured on enzymatically modified hairy regions (MHR-A and MHR-B) of apple pectins. MHRs were covalently attached to tissue culture polystyrene (TCPS) or glass. Uncoated substrata or bone slices were used as controls. Cell attachment, proliferation, and differentiation were investigated with fluorescence and confocal microscopy. Bone cells seem to prefer MHR-B coating to MHR-A coating. On MHR-A samples, the overall numbers as well as proportions of active osteoclasts were diminished compared to those on MHR-B, TCPS, or bone. Focal adhesions indicating attachment of the osteoblastic cells were detected on MHR-B and uncoated controls but not on MHR-A. These results demonstrate the possibility to modify surfaces with pectin nanocoatings.

A role for PVRL4-driven cell–cell interactions in tumorigenesis

PubMed Central

Pavlova, Natalya N; Pallasch, Christian; Elia, Andrew EH; Braun, Christian J; Westbrook, Thomas F; Hemann, Michael; Elledge, Stephen J

2013-01-01

During all stages of tumor progression, cancer cells are subjected to inappropriate extracellular matrix environments and must undergo adaptive changes in order to evade growth constraints associated with the loss of matrix attachment. A gain of function screen for genes that enable proliferation independently of matrix anchorage identified a cell adhesion molecule PVRL4 (poliovirus-receptor-like 4), also known as Nectin-4. PVRL4 promotes anchorage-independence by driving cell-to-cell attachment and matrix-independent integrin β4/SHP-2/c-Src activation. Solid tumors frequently have copy number gains of the PVRL4 locus and some have focal amplifications. We demonstrate that the transformation of breast cancer cells is dependent on PVRL4. Furthermore, growth of orthotopically implanted tumors in vivo is inhibited by blocking PVRL4-driven cell-to-cell attachment with monoclonal antibodies, demonstrating a novel strategy for targeted therapy of cancer. DOI: http://dx.doi.org/10.7554/eLife.00358.001 PMID:23682311

Sub-Domains of Ricin’s B Subunit as Targets of Toxin Neutralizing and Non-Neutralizing Monoclonal Antibodies

PubMed Central

Yermakova, Anastasiya; Vance, David J.; Mantis, Nicholas J.

2012-01-01

The B subunit (RTB) of ricin toxin is a galactose (Gal)−/N-acetylgalactosamine (GalNac)-specific lectin that mediates attachment, entry, and intracellular trafficking of ricin in host cells. Structurally, RTB consists of two globular domains with identical folding topologies. Domains 1 and 2 are each comprised of three homologous sub-domains (α, β, γ) that likely arose by gene duplication from a primordial carbohydrate recognition domain (CRD), although only sub-domains 1α and 2γ retain functional lectin activity. As part of our ongoing effort to generate a comprehensive B cell epitope map of ricin, we report the characterization of three new RTB-specific monoclonal antibodies (mAbs). All three mAbs, JB4, B/J F9 and C/M A2, were initially identified based on their abilities to neutralize ricin in a Vero cell cytotoxicty assay and to partially (or completely) block ricin attachment to cell surfaces. However, only JB4 proved capable of neutralizing ricin in a macrophage apoptosis assay and in imparting passive immunity to mice in a model of systemic intoxication. Using a combination of techniques, including competitive ELISAs, pepscan analysis, differential reactivity by Western blot, as well as affinity enrichment of phage displayed peptides, we tentatively localized the epitopes recognized by the non-neutralizing mAbs B/J F9 and C/M A2 to sub-domains 2α and 2β, respectively. Furthermore, we propose that the epitope recognized by JB4 is within sub-domain 2γ, adjacent to RTB’s high affinity Gal/GalNAc CRD. These data suggest that recognition of RTB’s sub-domains 1α and 2γ are critical determinants of antibody neutralizing activity and protective immunity to ricin. PMID:22984492

Comparison of methods for quantitating Salmonella enterica Typhimurium and Heidelberg strain attachment to reusable plastic shipping container coupons and preliminary assessment of sanitizer efficacy.

PubMed

Shi, Zhaohao; Baker, Christopher A; Lee, Sang In; Park, Si Hong; Kim, Sun Ae; Ricke, Steven C

2016-09-01

Salmonella serovars, one of the leading contributors to foodborne illness and are especially problematic for foods that are not cooked before consumption, such as fresh produce. The shipping containers that are used to transport and store fresh produce may play a role in cross contamination and subsequent illnesses. However, methods for quantitatively attached cells are somewhat variable. The overall goal of this study was to compare conventional plating with molecular methods for quantitating attached representative strains for Salmonella Typhimurium and Heidelberg on reusable plastic containers (RPC) coupons, respectively. We attached Salmonella enterica serovar Typhimurium ATCC 14028 and serovar Heidelberg SL486 (parent and an antibiotic resistant marker strain) to plastic coupons (2.54 cm(2)) derived from previously used shipping containers by growing for 72 h in tryptic soy broth. The impact of the concentration of sanitizer on log reductions between unsanitized and sanitized coupons was evaluated by exposing attached S. Typhimurium cells to 200 ppm and 200,000 ppm sodium hypochlorite (NaClO). Differences in sanitizer effectiveness between serovars were also evaluated with attached S. Typhimurium compared to attached S. Heidelberg populations after being exposed to 200 ppm peracetic acid (PAA). Treatment with NaClO caused an average of 2.73 ± 0.23 log CFU of S. Typhimurium per coupon removed with treatment at 200 ppm while 3.36 ± 0.54 log CFU were removed at 200,000 ppm. Treatment with PAA caused an average of 2.62 ± 0.15 log CFU removed for S. Typhimurium and 1.41 ± 0.17 log CFU for S. Heidelberg (parent) and 1.61 ± 0.08 log CFU (marker). Lastly, scanning electron microscopy (SEM) was used to visualize cell attachment and coupon surface topography. SEM images showed that remaining attached cell populations were visible even after sanitizer application. Conventional plating and qPCR yielded similar levels of enumerated bacterial populations indicating a high concordance between the two methods. Therefore, qPCR could be used for the rapid quantification of Salmonella attached on RPC.

Prostasomes: Their Characterisation: Implications for Human Reproduction: Prostasomes and Human Reproduction.

PubMed

Ronquist, Gunnar

2015-01-01

The prostate is a principal accessory genital gland that is vital for normal fertility. Epithelial cells lining the prostate acini release in a defined fashion (exocytosis) organellar nanosized structures named prostasomes. They are involved in the protection of sperm cells against immune response in the female reproductive tract by modulating the complement system and by inhibiting monocyte and neutrophil phagocytosis and lymphocyte proliferation. The immunomodulatory function most probably involves small non-coding RNAs present in prostasomes. Prostasomes have also been proposed to regulate the timing of sperm cell capacitation and induction of the acrosome reaction, since they are rich in various transferable bioactive molecules (e.g. receptors and enzymes) that promote the fertilising ability of sperm cells. Antigenicity of sperm cells has been well documented and implicated in involuntary immunological infertility of human couples, and antisperm antibodies (ASA) occur in several body fluids. The propensity of sperm cells to carry attached prostasomes suggests that they are a new category of sperm antigens. Circulating human ASA recognise prostasomes, and among 12 identified prostasomal antigens, prolactin- inducible protein (95 %) and clusterin (85 %) were immunodominant at the expense of the other 10 that were sporadically occurring.

West Nile virus discriminates between DC-SIGN and DC-SIGNR for cellular attachment and infection.

PubMed

Davis, Carl W; Nguyen, Hai-Yen; Hanna, Sheri L; Sánchez, Melissa D; Doms, Robert W; Pierson, Theodore C

2006-02-01

The C-type lectins DC-SIGN and DC-SIGNR bind mannose-rich glycans with high affinity. In vitro, cells expressing these attachment factors efficiently capture, and are infected by, a diverse array of appropriately glycosylated pathogens, including dengue virus. In this study, we investigated whether these lectins could enhance cellular infection by West Nile virus (WNV), a mosquito-borne flavivirus related to dengue virus. We discovered that DC-SIGNR promoted WNV infection much more efficiently than did DC-SIGN, particularly when the virus was grown in human cell types. The presence of a single N-linked glycosylation site on either the prM or E glycoprotein of WNV was sufficient to allow DC-SIGNR-mediated infection, demonstrating that uncleaved prM protein present on a flavivirus virion can influence viral tropism under certain circumstances. Preferential utilization of DC-SIGNR was a specific property conferred by the WNV envelope glycoproteins. Chimeras between DC-SIGN and DC-SIGNR demonstrated that the ability of DC-SIGNR to promote WNV infection maps to its carbohydrate recognition domain. WNV virions and subviral particles bound to DC-SIGNR with much greater affinity than DC-SIGN. We believe this is the first report of a pathogen interacting more efficiently with DC-SIGNR than with DC-SIGN. Our results should lead to the discovery of new mechanisms by which these well-studied lectins discriminate among ligands.

Cytochemical Labeling for Fungal and Host Components in Plant Tissues Inoculated with Fungal Wilt Pathogens

NASA Astrophysics Data System (ADS)

Ouellette, G. B.; Baayen, R. P.; Chamberland, H.; Simard, M.; Rioux, D.; Charest, P. M.

2004-08-01

Antibodies to detect pectin in present investigations attached to distinct fibrils in vessel lumina. In carnation infected with an isolate of Fusarium oxysporum f.sp., labeling of pathogen cells also occurred; in a resistant cultivar (cv.), it was coincident with proximate pectin fibrils and linked to altered fungal walls, which was the opposite in the susceptible cv., indicating that hindrance of pathogen ability to degrade pectin may be related to resistance. Labeling of the fungus in culture was nil, except in media containing pectin, showing that pectin is not native to the pathogen. Labeling of fungal walls for cellulose in elm (inoculated with Ophiostoma novo-ulmi) and carnation also occurred, linked to adsorbed host wall components. The chitin probe often attached to dispersed matter, in vessel lumina, traceable to irregularly labeled fungal cells and host wall degradation products. With an anti-horseradish peroxidase probe, host and fungal walls were equally labeled, and with a glucosidase, differences of labeling between these walls were observed, depending on pH of the test solution. Fungal extracellular matter and filamentous structures, present in fungal walls, predominantly in another elm isolate (Phaeotheca dimorphospora), did not label with any of the probes used. However, in cultures of this fungus, extracellular material labeled, even at a distance from the colony margin, with an anti-fimbriae probe.

Conditional Function of Autoaggregative Protein Cah and Common cah Mutations in Shiga Toxin-Producing Escherichia coli

PubMed Central

Brandl, Maria T.; Kudva, Indira T.; Katani, Robab; Moreau, Matthew R.; Kapur, Vivek

2017-01-01

ABSTRACT Cah is a calcium-binding autotransporter protein involved in autoaggregation and biofilm formation. Although cah is widespread in Shiga toxin-producing Escherichia coli (STEC), we detected mutations in cah at a frequency of 31.3% in this pathogen. In STEC O157:H7 supershedder strain SS17, a large deletion results in a smaller coding sequence, encoding a protein lacking the C-terminal 71 amino acids compared with Cah in STEC O157:H7 strain EDL933. We examined the function of Cah in biofilm formation and host colonization to better understand the selective pressures for cah mutations. EDL933-Cah played a conditional role in biofilm formation in vitro: it enhanced E. coli DH5α biofilm formation on glass surfaces under agitated culture conditions that prevented autoaggregation but inhibited biofilm formation under hydrostatic conditions that facilitated autoaggregation. This function appeared to be strain dependent since Cah-mediated biofilm formation was diminished when an EDL933 cah gene was expressed in SS17. Deletion of cah in EDL933 enhanced bacterial attachment to spinach leaves and altered the adherence pattern of EDL933 to bovine recto-anal junction squamous epithelial (RSE) cells. In contrast, in trans expression of EDL933 cah in SS17 increased its attachment to leaf surfaces, and in DH5α, it enhanced its adherence to RSE cells. Hence, the ecological function of Cah appears to be modulated by environmental conditions and other bacterial strain-specific properties. Considering the prevalence of cah in STEC and its role in attachment and biofilm formation, cah mutations might be selected in ecological niches in which inactivation of Cah would result in an increased fitness in STEC during colonization of plants or animal hosts. IMPORTANCE Shiga toxin-producing Escherichia coli (STEC) harbors genes encoding diverse adhesins, and many of these are known to play an important role in bacterial attachment and host colonization. We demonstrated here that the autotransporter protein Cah confers on E. coli DH5α cells a strong autoaggregative phenotype that is inversely correlated with its ability to form biofilms and plays a strain-specific role in plant and animal colonization by STEC. Although cah is widespread in the STEC population, we detected a mutation rate of 31.3% in cah, which is similar to that reported for rpoS and fimH. The formation of cell aggregates due to increased bacterium-to-bacterium interactions may be disadvantageous to bacterial populations under conditions that favor a planktonic state in STEC. Therefore, a loss-of-function mutation in cah is likely a selective trait in STEC when autoaggregative properties become detrimental to bacterial cells and may contribute to the adaptability of STEC to fluctuating environments. PMID:29054868

Light-activated nanotube-porphyrin conjugates as effective antiviral agents

NASA Astrophysics Data System (ADS)

Banerjee, Indrani; Douaisi, Marc P.; Mondal, Dhananjoy; Kane, Ravi S.

2012-03-01

Porphyrins have been used for photodynamic therapy (PDT) against a wide range of targets like bacteria, viruses and tumor cells. In this work, we report porphyrin-conjugated multi-walled carbon nanotubes (NT-P) as potent antiviral agents. Specifically, we used Protoporphyrin IX (PPIX), which we attached to acid-functionalized multi-walled carbon nanotubes (MWNTs). We decided to use carbon nanotubes as scaffolds because of their ease of recovery from a solution through filtration. In the presence of visible light, NT-P was found to significantly reduce the ability of Influenza A virus to infect mammalian cells. NT-P may be used effectively against influenza viruses with little or no chance of them developing resistance to the treatment. Furthermore, NT-P can be easily recovered through filtration which offers a facile strategy to reuse the active porphyrin moiety to its fullest extent. Thus NT-P conjugates represent a new approach for preparing ex vivo reusable antiviral agents.

Attachment of Asaia bogorensis Originating in Fruit-Flavored Water to Packaging Materials

PubMed Central

Otlewska, Anna; Antolak, Hubert

2014-01-01

The objective of this study was to investigate the adhesion of isolated spoilage bacteria to packaging materials used in the food industry. Microorganisms were isolated from commercial fruit-flavored mineral water in plastic bottles with flocks as a visual defect. The Gram-negative rods were identified using the molecular method through the amplification of a partial region of the 16S rRNA gene. Based on the sequence identity (99.6%) between the spoilage organism and a reference strain deposited in GenBank, the spoilage isolate was identified as Asaia bgorensis. Experiments on bacterial adhesion were conducted using plates made of glass and polystyrene (packaging materials commonly used in the beverage industry). Cell adhesion ability was determined using luminometry, plate count, and the microscopic method. The strain of A. bogorensis was characterized by strong adhesion properties which were dependent on the surface type, with the highest cell adhesion detected on polystyrene. PMID:25295262

Why are people with insecure attachments unhappy? The mediation of meaning in life.

PubMed

Yen, Chih-Long

2014-10-01

This study explores the mediating role of meaning in life with respect to the relationship between adult attachment and psychological well-being. This study proposed that because people with insecure attachments have difficulty in connecting with the external world, this may decrease their ability to attain meaning in life, which may in turn diminish their well-being. Two studies (N = 155 and 234, respectively, with age range from 18 to 31) were conducted. Participants completed self-report measures of attachment, well-being and meaning in life. The results consistently revealed a full mediation effect of meaning in life on the relationship between attachment and well-being. The findings suggested that a deeper sense of meaning in life comes from one's connection with the external world, especially with other people. © 2014 International Union of Psychological Science.

Functionalized coatings by cold spray: An in vitro study of micro- and nanocrystalline hydroxyapatite compared to porous titanium.

PubMed

Vilardell, A M; Cinca, N; Garcia-Giralt, N; Dosta, S; Cano, I G; Nogués, X; Guilemany, J M

2018-06-01

Three different surface treatments on a Ti6Al4V alloy have been in vitro tested for possible application in cementless joint prosthesis. All of them involve the novelty of using the Cold Spray technology for their deposition: (i) an as-sprayed highly rough titanium and, followed by the deposition of a thin hydroxyapatite layer with (ii) microcrystalline or (iii) nanocrystalline structure. Primary human osteoblasts were extracted from knee and seeded onto the three different surfaces. Cell viability was tested by MTS and LIVE/DEAD assays, cell differentiation by alkaline phosphatase (ALP) quantification and cell morphology by Phalloidin staining. All tests were carried out at 1, 7 and 14 days of cell culture. Different cell morphologies between titanium and hydroxyapatite surfaces were exhibited. At 1 day of cell culture, cells on the titanium coating were spread and flattened, expanding the filopodia actin filaments in all directions, while cells on the hydroxyapatite coatings showed round like-shape morphology due to slower attachment. Higher cell viability was detected at all times of cell culture on titanium coating due to a better attachment at 1 day. However, from 7 days of cell culture, cells on hydroxyapatite showed good attachment onto surfaces and highly increased their proliferation, mostly on nanocrystalline, achieving similar cell viability levels than titanium coatings. ALP levels were significantly higher in titanium, in part, because of greatest cell number. Overall, the best cell functional results were obtained on titanium coatings whereas microcrystalline hydroxyapatite presented the worst cellular parameters. However, results indicate that nanocrystalline hydroxyapatite coatings may achieve promising results for the faster cell proliferation once cells are attached on the surface. Copyright © 2018 Elsevier B.V. All rights reserved.

Effects of air polishing and an amino acid buffered hypochlorite solution to dentin surfaces and periodontal ligament cell survival, attachment, and spreading.

PubMed

Schmidlin, Patrick R; Fujioka-Kobayashi, Masako; Mueller, Heinz-Dieter; Sculean, Anton; Lussi, Adrian; Miron, Richard J

2017-06-01

The aim of this study is to examine morphological changes of dentin surfaces following air polishing or amino acid buffered hypochlorite solution application and to assess their influence on periodontal ligament (PDL) cell survival, attachment, and spreading to dentin discs in vitro. Bovine dentin discs were treated with either (i) Classic, (ii) Plus, or (iii) Perio powder (EMS). Furthermore, Perisolv® a hypochlorite solution buffered with various amino acids was investigated. Untreated dentin discs served as controls. Morphological changes to dentin discs were assessed using scanning electron microscopy (SEM). Human PDL cells were seeded onto the respectively treated discs, and samples were then investigated for PDL cell survival, attachment, and spreading using a live/dead assay, adhesion assay, and SEM imaging, respectively. Both control and Perisolv®-rinsed dentin discs demonstrated smooth surfaces at low and high magnifications. The Classic powders demonstrated the thickest coating followed by the Powder Plus. The Perio powder demonstrated marked alterations of dentin discs by revealing the potential to open dentinal tubules even before rinsing. Seeding of PDL cells demonstrated an almost 100 % survival rate on all samples demonstrating very high biocompatibility for all materials. Significantly higher PDL cell numbers were observed on samples treated with the Perio powder and the Perisolv® solution (approximately 40 % more cells; p < 0.05). SEM imaging revealed the potential for PDL cells to attach and spread on all surfaces. The results from the present study demonstrate that cell survival and spreading of PDL cells on root surfaces is possible following either air polishing or application with Perisolv®. Future in vitro and animal testing is necessary to further characterize the beneficial effects of either system in a clinical setting. The use of air polishing or application with Perisolv amino acid buffered hypochlorite solution was effective in treating root surfaces and allowed for near 100 % PDL cell survival, attachment, and spreading onto all root surfaces.

First insights into the pleiotropic role of vrf (yedF), a newly characterized gene of Salmonella Typhimurium.

PubMed

Ballesté-Delpierre, Clara; Fernandez-Orth, Dietmar; Ferrer-Navarro, Mario; Díaz-Peña, Ramón; Odena-Caballol, Antonia; Oliveira, Eliandre; Fàbrega, Anna; Vila, Jordi

2017-11-10

Salmonella possesses virulence determinants that allow replication under extreme conditions and invasion of host cells, causing disease. Here, we examined four putative genes predicted to encode membrane proteins (ydiY, ybdJ, STM1441 and ynaJ) and a putative transcriptional factor (yedF). These genes were identified in a previous study of a S. Typhimurium clinical isolate and its multidrug-resistant counterpart. For STM1441 and yedF a reduced ability to interact with HeLa cells was observed in the knock-out mutants, but an increase in this ability was absent when these genes were overexpressed, except for yedF which phenotype was rescued when yedF was restored. In the absence of yedF, decreased expression was seen for: i) virulence-related genes involved in motility, chemotaxis, attachment and survival inside the host cell; ii) global regulators of the invasion process (hilA, hilC and hilD); and iii) factors involved in LPS biosynthesis. In contrast, an increased expression was observed for anaerobic metabolism genes. We propose yedF is involved in the regulation of Salmonella pathogenesis and contributes to the activation of the virulence machinery. Moreover, we propose that, when oxygen is available, yedF contributes sustained repression of the anaerobic pathway. Therefore, we recommend this gene be named vrf, for virulence-related factor.

Bioactive bredigite coating with improved bonding strength, rapid apatite mineralization and excellent cytocompatibility.

PubMed

Yi, Deliang; Wu, Chengtie; Ma, Bing; Ji, Heng; Zheng, Xuebin; Chang, Jiang

2014-05-01

Previous studies have shown that bredigite (Ca7MgSi4O16) bioceramics possessed excellent biocompatibility, apatite-mineralization ability and mechanical properties. In this paper, the bredigite coating on Ti-6Al-4 V substrate was prepared by plasma spraying technique. The main compositions of the coating were bredigite crystal phase with small parts of amorphous phases. The bonding strength of the coating to Ti-6Al-4 V substrate reached 49.8 MPa, which was significantly higher than that of hydroxyapatite coating and other silicate-based bioceramic coatings prepared by same method. After immersed in simulated body fluid for 2 days, a distinct apatite layer was deposited on the surface of bredigite coating, indicating that the prepared bredigite coating has excellent apatite-mineralization ability. The prepared bredigite coating supported the attachment and proliferation of rabbit bone marrow stem cells. The proliferation level of bone marrow stem cells was significantly higher than that on the hydroxyapatite coating. Our further study showed that the released SiO4 (4-) and Mg(2+) ions from bredigite coating as well as the formed nano-apatite layer on the coating surface might mainly contribute to the improvement of cell proliferation. The results indicated that the bredigite coating may be applied on orthopedic implants due to its excellent bonding strength, apatite mineralization and cytocompatibility.

3D printing of biomaterials with mussel-inspired nanostructures for tumor therapy and tissue regeneration.

PubMed

Ma, Hongshi; Luo, Jian; Sun, Zhe; Xia, Lunguo; Shi, Mengchao; Liu, Mingyao; Chang, Jiang; Wu, Chengtie

2016-12-01

Primary bone cancer brings patients great sufferings. To deal with the bone defects resulted from cancer surgery, biomaterials with good bone-forming ability are necessary to repair bone defects. Meanwhile, in order to prevent possible tumor recurrence, it is essential that the remaining tumor cells around bone defects are completely killed. However, there are few biomaterials with the ability of both cancer therapy and bone regeneration until now. Here, we fabricated a 3D-printed bioceramic scaffold with a uniformly self-assembled Ca-P/polydopamine nanolayer surface. Taking advantage of biocompatibility, biodegradability and the excellent photothermal effect of polydopamine, the bifunctional scaffolds with mussel-inspired nanostructures could be used as a satisfactory and controllable photothermal agent, which effectively induced tumor cell death in vitro, and significantly inhibited tumor growth in mice. In addition, owing to the nanostructured surface, the prepared polydopamine-modified bioceramic scaffolds could support the attachment and proliferation of rabbit bone mesenchymal stem cells (rBMSCs), and significantly promoted the formation of new bone tissues in rabbit bone defects even under photothermal treatment. Therefore, the mussel-inspired nanostructures in 3D-printed bioceramic exhibited a remarkable capability for both cancer therapy and bone regeneration, offering a promising strategy to construct bifunctional biomaterials which could be widely used for therapy of tumor-induced tissue defects. Copyright © 2016 Elsevier Ltd. All rights reserved.

Enhanced Microchip Electrophoresis Separations Combined with Electrochemical Detection Utilizing a Capillary Embedded in Polystyrene.

PubMed

Mehl, Benjamin T; Martin, R Scott

2018-01-07

The ability to use microchip-based electrophoresis for fast, high-throughput separations provides researchers with a tool for close-to real time analysis of biological systems. While PDMS-based electrophoresis devices are popular, the separation efficiency is often an issue due to the hydrophobic nature of PDMS. In this study, a hybrid microfluidic capillary device was fabricated to utilize the positive features of PDMS along with the electrophoretic performance of fused silica. A capillary loop was embedded in a polystyrene base that can be coupled with PDMS microchannels at minimal dead volume interconnects. A method for cleaning out the capillaries after a wet-polishing step was devised through the use of 3D printed syringe attachment. By comparing the separation efficiency of fluorescein and CBI-glycine with both a PDMS-based serpentine device and the embedded capillary loop device, it was shown that the embedded capillary loop device maintained higher theoretical plates for both analytes. A Pd decoupler with a carbon or Pt detection electrode were embedded along with the loop allowing integration of the electrophoretic separation with electrochemical detection. A series of catecholamines were separated to show the ability to resolve similar analytes and detect redox active species. The release of dopamine and norepinephrine from PC 12 cells was also analyzed showing the compatibility of these improved microchip separations with high ionic cell buffers associated with cell culture.

Elevated temperature increases carbon and nitrogen fluxes between phytoplankton and heterotrophic bacteria through physical attachment

DOE PAGES

Arandia-Gorostidi, Nestor; Weber, Peter K.; Alonso-Sáez, Laura; ...

2016-12-06

Quantifying the contribution of marine microorganisms to carbon and nitrogen cycles and their response to predicted ocean warming is one of the main challenges of microbial oceanography. Here we present a single-cell NanoSIMS isotope analysis to quantify C and N uptake by free-living and attached phytoplankton and heterotrophic bacteria, and their response to short-term experimental warming of 4 °C. Elevated temperature increased total C fixation by over 50%, a small but significant fraction of which was transferred to heterotrophs within 12 h. Cell-to-cell attachment doubled the secondary C uptake by heterotrophic bacteria and increased secondary N incorporation by autotrophs bymore » 68%. Warming also increased the abundance of phytoplankton with attached heterotrophs by 80%, and promoted C transfer from phytoplankton to bacteria by 17% and N transfer from bacteria to phytoplankton by 50%. Lastly, our results indicate that phytoplankton-bacteria attachment provides an ecological advantage for nutrient incorporation, suggesting a mutualistic relationship that appears to be enhanced by temperature increases.« less

Elevated temperature increases carbon and nitrogen fluxes between phytoplankton and heterotrophic bacteria through physical attachment

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

Arandia-Gorostidi, Nestor; Weber, Peter K.; Alonso-Sáez, Laura

Quantifying the contribution of marine microorganisms to carbon and nitrogen cycles and their response to predicted ocean warming is one of the main challenges of microbial oceanography. Here we present a single-cell NanoSIMS isotope analysis to quantify C and N uptake by free-living and attached phytoplankton and heterotrophic bacteria, and their response to short-term experimental warming of 4 °C. Elevated temperature increased total C fixation by over 50%, a small but significant fraction of which was transferred to heterotrophs within 12 h. Cell-to-cell attachment doubled the secondary C uptake by heterotrophic bacteria and increased secondary N incorporation by autotrophs bymore » 68%. Warming also increased the abundance of phytoplankton with attached heterotrophs by 80%, and promoted C transfer from phytoplankton to bacteria by 17% and N transfer from bacteria to phytoplankton by 50%. Lastly, our results indicate that phytoplankton-bacteria attachment provides an ecological advantage for nutrient incorporation, suggesting a mutualistic relationship that appears to be enhanced by temperature increases.« less

Deposition and disinfection of Escherichia coli O157:H7 on naturally occurring photoactive materials in a parallel plate chamber†

PubMed Central

Taylor, Alicia A.; Chowdhury, Indranil; Gong, Amy S.; Cwiertny, David M.; Walker, Sharon L.

2014-01-01

Dissolved organic matter in combination with iron oxides has been shown to facilitate photochemical disinfection through the production of reactive oxygen species (ROS) under UV and visible light. However, due to the extremely short lifetime of these radicals, the disinfection effciency is limited by the successful transport of ROS to bacterial surfaces. This study was designed to quantitatively investigate three collector surfaces with various potentials to produce ROS [bare quartz, hematite (α-Fe2O3) coated quartz, and Suwannee River humic acid (SRHA)] and the effects of extracellular polymeric substance (EPS) (full or partial coating) and solution chemistry (ionic strength, IS) on the interactions between bacteria and the ROS-producing substrates. With few exceptions, bacterial deposition studies in a parallel plate (PP) flow chamber have revealed increasing cell adhesion with IS. Furthermore, interactions between collector surfaces and cells can be explained by electrostatic forces, with negatively charged SRHA reducing and positively charged α-Fe2O3 enhancing bacterial deposition significantly. Increased deposition was also observed with full EPS content, indicating the ability of EPS to facilitate interaction between cells and surfaces in the aquatic environment. In complementary disinfection studies conducted with simulated light, viability loss was observed for cells fully coated with EPS when attached to α-Fe2O3 under all IS conditions. Based upon our prior study in which EPS was found to not inhibit hydroxyl radical activity toward bacteria, we proposed that EPS might therefore promote disinfection by facilitating cell attachment to ROS-producing surfaces where higher concentrations of ROS are expected at closer proximities to reactive substrates (e.g., SRHA and α-Fe2O3). Our findings on the mechanism and controlling factors of cell interactions with photoactive substrates provide insight as to the role of ionic strength in photochemical disinfection processes. PMID:24362649

Attachment and spatial organisation of human mesenchymal stem cells on poly(ethylene glycol) hydrogels.

PubMed

Chahal, Aman S; Schweikle, Manuel; Heyward, Catherine A; Tiainen, Hanna

2018-08-01

Strategies that enable hydrogel substrates to support cell attachment typically incorporate either entire extracellular matrix proteins or synthetic peptide fragments such as the RGD (arginine-glycine-aspartic acid) motif. Previous studies have carefully analysed how material characteristics can affect single cell morphologies. However, the influence of substrate stiffness and ligand presentation on the spatial organisation of human mesenchymal stem cells (hMSCs) have not yet been examined. In this study, we assessed how hMSCs organise themselves on soft (E = 7.4-11.2 kPa) and stiff (E = 27.3-36.8 kPa) poly(ethylene glycol) (PEG) hydrogels with varying concentrations of RGD (0.05-2.5 mM). Our results indicate that hMSCs seeded on soft hydrogels clustered with reduced cell attachment and spreading area, irrespective of RGD concentration and isoform. On stiff hydrogels, in contrast, cells spread with high spatial coverage for RGD concentrations of 0.5 mM or higher. In conclusion, we identified that an interplay of hydrogel stiffness and the availability of cell attachment motifs are important factors in regulating hMSC organisation on PEG hydrogels. Understanding how cells initially interact and colonise the surface of this material is a fundamental prerequisite for the design of controlled platforms for tissue engineering and mechanobiology studies. Copyright © 2018 Elsevier Ltd. All rights reserved.

Phenotypic Heterogeneity in Attachment of Marine Bacteria toward Antifouling Copolymers Unraveled by AFM.

PubMed

El-Kirat-Chatel, Sofiane; Puymege, Aurore; Duong, The H; Van Overtvelt, Perrine; Bressy, Christine; Belec, Lénaïk; Dufrêne, Yves F; Molmeret, Maëlle

2017-01-01

Up to recent years, bacterial adhesion has mostly been evaluated at the population level. Single cell level has improved in the past few years allowing a better comprehension of the implication of individual behaviors as compared to the one of a whole community. A new approach using atomic force microscopy (AFM) to measure adhesion forces between a live bacterium attached via a silica microbead to the AFM tipless cantilever and the surface has been recently developed. The objectives of this study is to examine the bacterial adhesion to a surface dedicated to ship hulls at the population and the cellular level to understand to what extent these two levels could be correlated. Adhesion of marine bacteria on inert surfaces are poorly studied in particular when substrata are dedicated to ship hulls. Studying these interactions in this context are worthwhile as they may involve different adhesion behaviors, taking place in salty conditions, using different surfaces than the ones usually utilized in the literacy. FRC (fouling release coatings)-SPC (self-polishing coatings) hybrids antifouling coatings have been used as substrata and are of particular interest for designing environmentally friendly surfaces, combining progressive surface erosion and low adhesion properties. In this study, a hybrid coating has been synthetized and used to study the adhesion of three marine bacteria, displaying different surface characteristics, using microplate assays associated with confocal scanning laser microscopy (CSLM) and AFM. This study shows that the bacterial strain that appeared to have the weakest adhesion and biofilm formation abilities when evaluated at the population level using microplates assays and CSLM, displayed stronger adhesion forces on the same surfaces at the single cell level using AFM. In addition, one of the strains tested which presented a strong ability to adhere and to form biofilm at the population level, displayed a heterogeneous phenotypic behavior at the single cell level. Therefore, these results suggest that the evaluation of adhesion at the population level cannot always be correlated with adhesion forces measured individually by AFM and that some bacteria are prone to phenotypic heterogeneity among their population.

Lipid tethering of breast tumor cells enables real-time imaging of free-floating cell dynamics and drug response

PubMed Central

Whipple, Rebecca A.; Zhang, Peipei; Sooklal, Elisabeth L.; Martin, Stuart S.; Jewell, Christopher M.

2016-01-01

Free-floating tumor cells located in the blood of cancer patients, known as circulating tumor cells (CTCs), have become key targets for studying metastasis. However, effective strategies to study the free-floating behavior of tumor cells in vitro have been a major barrier limiting the understanding of the functional properties of CTCs. Upon extracellular-matrix (ECM) detachment, breast tumor cells form tubulin-based protrusions known as microtentacles (McTNs) that play a role in the aggregation and re-attachment of tumor cells to increase their metastatic efficiency. In this study, we have designed a strategy to spatially immobilize ECM-detached tumor cells while maintaining their free-floating character. We use polyelectrolyte multilayers deposited on microfluidic substrates to prevent tumor cell adhesion and the addition of lipid moieties to tether tumor cells to these surfaces through interactions with the cell membranes. This coating remains optically clear, allowing capture of high-resolution images and videos of McTNs on viable free-floating cells. In addition, we show that tethering allows for the real-time analysis of McTN dynamics on individual tumor cells and in response to tubulin-targeting drugs. The ability to image detached tumor cells can vastly enhance our understanding of CTCs under conditions that better recapitulate the microenvironments they encounter during metastasis. PMID:26871289

Nanoparticle flotation collectors--the influence of particle softness.

PubMed

Yang, Songtao; Razavizadeh, Bi Bi Marzieh; Pelton, Robert; Bruin, Gerard

2013-06-12

The ability of polymeric nanoparticles to promote glass bead and pentlandite (Pn, nickel sulfide mineral) attachment to air bubbles in flotation was measured as a function of the nanoparticle glass transition temperature using six types of nanoparticles based on styrene/N-butylacrylate copolymers. Nanoparticle size, surface charge density, and hydrophobicity were approximately constant over the series. The ability of the nanoparticles to promote air bubble attachment and perform as flotation collectors was significantly greater for softer nanoparticles. We propose that softer nanoparticles were more firmly attached to the glass beads or mineral surface because the softer particles had a greater glass/polymer contact areas and thus stronger overall adhesion. The diameters of the contact areas between polymeric nanoparticles and glass surfaces were estimated with the Young-Laplace equation for soft, liquidlike particles, whereas JKR adhesion theory was applied to the harder polystyrene particles. The diameters of the contact areas were estimated to be more than an order of magnitude greater for the soft particles compared to harder polystyrene particles.

Ability to home in small site-attached coral reef fishes.

PubMed

Booth, D J

2016-08-01

The ability of two common, site-attached coral-reef fishes to return to their home corals after displacement was investigated in a series of field experiments at One Tree Island, southern Great Barrier Reef. The humbug Dascyllus aruanus was displaced up to 250 m, with 42% of individuals returning home, irrespective of body size, displacement, direction (up or across currents) and route complexity, while for the lemon damselfish Pomacentrus moluccensis 35% of individuals returned overall, with 33% from the greatest displacement, 100 m along a reef edge. Given that the home range of both species is <1 m(2) , over their 10+ year life span, the mechanisms and motivations for such homing ability are unclear but it may allow resilience if fishes are displaced by storm events, allowing rapid return to home corals. © 2016 The Fisheries Society of the British Isles.

Rumen Bacterial Degradation of Forage Cell Walls Investigated by Electron Microscopy

PubMed Central

Akin, Danny E.; Amos, Henry E.

1975-01-01

The association of rumen bacteria with specific leaf tissues of the forage grass Kentucky-31 tall fescue (Festuca arundinacea Schreb.) during in vitro degradation was investigated by transmission and scanning electron microscopy. Examination of degraded leaf cross-sections revealed differential rates of tissue degradation in that the cell walls of the mesophyll and pholem were degraded prior to those of the outer bundle sheath and epidermis. Rumen bacteria appeared to degrade the mesophyll, in some cases, and phloem without prior attachment to the plant cell walls. The degradation of bundle sheath and epidermal cell walls appeared to be preceded by attachment of bacteria to the plant cell wall. Ultrastructural features apparently involved in the adhesion of large cocci to plant cells were observed by transmission and scanning electron microscopy. The physical association between plant and rumen bacterial cells during degradation apparently varies with tissue types. Bacterial attachment, by extracellular features in some microorganisms, is required prior to degradation of the more resistant tissues. Images PMID:16350017

How Escherichia coli lands and forms cell clusters on a surface: a new role of surface topography

PubMed Central

Gu, Huan; Chen, Aaron; Song, Xinran; Brasch, Megan E.; Henderson, James H.; Ren, Dacheng

2016-01-01

Bacterial response to surface topography during biofilm formation was studied using 5 μm tall line patterns of poly (dimethylsiloxane) (PDMS). Escherichia coli cells attached on top of protruding line patterns were found to align more perpendicularly to the orientation of line patterns when the pattern narrowed. Consistently, cell cluster formation per unit area on 5 μm wide line patterns was reduced by 14-fold compared to flat PDMS. Contrasting the reduced colony formation, cells attached on narrow patterns were longer and had higher transcriptional activities, suggesting that such unfavorable topography may present a stress to attached cells. Results of mutant studies indicate that flagellar motility is involved in the observed preference in cell orientation on narrow patterns, which was corroborated by the changes in cell rotation pattern before settling on different surface topographies. These findings led to a set of new design principles for creating antifouling topographies, which was validated using 10 μm tall hexagonal patterns. PMID:27412365

Attachment's Links With Adolescents' Social Emotions: The Roles of Negative Emotionality and Emotion Regulation.

PubMed

Murphy, Tia Panfile; Laible, Deborah J; Augustine, Mairin; Robeson, Lindsay

2015-01-01

Recent research has attempted to explain the mechanisms through which parental attachment affects social and emotional outcomes (e.g., Burnette, Taylor, Worthington, & Forsyth, 2007 ; Panfile & Laible, 2012 ). The authors' goal was to examine negative emotionality and emotion regulation as mediators of the associations that attachment has with empathy, forgiveness, guilt, and jealousy. One hundred forty-eight adolescents reported their parental attachment security, general levels of negative emotionality and abilities to regulate emotional responses, and tendencies to feel empathy, forgiveness, guilt, and jealousy. Results revealed that attachment security was associated with higher levels of empathy, forgiveness, and guilt, but lower levels of jealousy. In addition, emotion regulation mediated the links attachment shared with both empathy and guilt, such that higher levels of attachment security were linked with greater levels of emotion regulation, which led to greater levels of empathy and guilt. Alternatively, negative emotionality mediated the links attachment shared with both forgiveness and jealousy, such that higher levels of attachment security were associated with lower levels of negative emotionality, which in turn was linked to lower levels of forgiveness and higher levels of jealousy. This study provides a general picture of how attachment security may play a role in shaping an individual's levels of social emotions.

[Stress in Medical Students: A Cross-Sectional Study on the Relevance of Attachment Style and Structural Integration].

PubMed

Bugaj, Till Johannes; Müksch, Christine; Ehrenthal, Johannes C; Köhl-Hackert, Nadja; Schauenburg, Henning; Huber, Julia; Schmid, Carolin; Erschens, Rebecca; Junne, Florian; Herzog, Wolfgang; Nikendei, Christoph

2016-02-01

From year one of studying medicine an increase of psychological stress is found. The relationship between the occurrence of this stress and attachment patterns or structural personality functions remains unclear. The present study aimed at investigating whether a relationship between the enduring personality variables, attachment style and level of structural integration of the personality, and acute stress experience at the beginning of medical students' studies exists. In this study, all students in the first semester of medicine were invited to participate in a study to identify stress factors via questionnaire (MBI-SS, PSQ, PHQ-9, GAD-7) in the WS 2013/2014. Simultaneously, the predominant attachment style (RQ-2) and structural abilities (OPD-SFK) were evaluated. The study included 293 students (return: 91.3%). Securely attached students experienced significantly less stress than insecurely attached students (p=0.019). Students with a high level of structural integration showed significantly less stress burden (p<0.001) and lower exhaustion- (p<0.001) and cynicism values (p<0.001), while showing a higher experience of self-efficacy (p<0.001). The influence of attachment behavior on stress experience is mediated by the level of the structural integration of the personality. Significant correlations exist between attachment style and the level of structural integration of the personality, and burnout risk as well as stress burden. The level of structural integration of the personality mediates the relationship between the attachment-related "model of self" and stress experience, i. e. a positive "model of self" can have a stress-protective effect when good structural abilities are present. Practical implication: An insecure attachment style and a low level of structural integration may be associated with higher stress experience when transitioning to study. The results suggest that the enduring personality variable attachment style, mediated by the level of structural integration of the personality, leads to higher stress and burnout experience. Affected students could be supported by early preventive measures enabling the sustainable preparation for this transitional period. Longitudinal prospective studies are needed to explore if the assumption is applicable that pre-existing vulnerabilities in school are exacerbated at this transitional stage. © Georg Thieme Verlag KG Stuttgart · New York.

Separation of somatic and germ cells is required to establish primate spermatogonial cultures.

PubMed

Langenstroth, Daniel; Kossack, Nina; Westernströer, Birgit; Wistuba, Joachim; Behr, Rüdiger; Gromoll, Jörg; Schlatt, Stefan

2014-09-01

Can primate spermatogonial cultures be optimized by application of separation steps and well defined culture conditions? We identified the cell fraction which provides the best source for primate spermatogonia when prolonged culture is desired. Man and marmoset show similar characteristics in regard to germ cell development and function. Several protocols for isolation and culture of human testis-derived germline stem cells have been described. Subsequent analysis revealed doubts on the germline origin of these cells and characterized them as mesenchymal stem cells or fibroblasts. Studies using marmosets as preclinical model confirmed that the published isolation protocols did not lead to propagation of germline cells. Testicular cells derived from nine adult marmoset monkeys (Callithrix jacchus) were cultured for 1, 3, 6 and 11 days and consecutively analyzed for the presence of spermatogonia, differentiating germ cells and testicular somatic cells. Testicular tissue of nine adult marmoset monkeys was enzymatically dissociated and subjected to two different cell culture approaches. In the first approach all cells were kept in the same dish (non-separate culture, n = 5). In the second approach the supernatant cells were transferred into a new dish 24 h after seeding and subsequently supernatant and attached cells were cultured separately (separate culture, n = 4). Real-time quantitative PCR and immunofluorescence were used to analyze the expression of reliable germ cell and somatic markers throughout the culture period. Germ cell transplantation assays and subsequent wholemount analyses were performed to functionally evaluate the colonization of spermatogonial cells. This is the first report revealing an efficient isolation and culture of putative marmoset spermatogonial stem cells with colonization ability. Our results indicate that a separation of spermatogonia from testicular somatic cells is a crucial step during cell preparation. We identified the overgrowth of more rapidly expanding somatic cells to be a major problem when establishing spermatogonial cultures. Initiating germ cell cultures from the supernatant and maintaining germ cells in suspension cultures minimized the somatic cell contamination and provided enriched germ cell fractions which displayed after 11 days of culture a significantly higher expression of germ cell markers genes (DDX-4, MAGE A-4; P < 0.05) compared with separately cultured attached cells. Additionally, germ cell transplantation experiments demonstrated a significantly higher absolute number of cells with colonization ability (P < 0.001) in supernatant cells after 11 days of separate culture. This study presents a relevant aspect for the successful setup of spermatogonial cultures but provides limited data regarding the question of whether the long-term maintenance of spermatogonia can be achieved. Transfer of these preclinical data to man may require modifications of the protocol. Spermatogonial cultures from rodents have become important and innovative tools for basic and applied research in reproductive biology and veterinary medicine. It is expected that spermatogonia-based strategies will be transformed into clinical applications for the treatment of male infertility. Our data in the marmoset monkey may be highly relevant to establish spermatogonial cultures of human testes. Funding was provided by the DFG-Research Unit FOR 1041 Germ Cell Potential (SCHL394/11-2) and by the Graduate Program Cell Dynamics and Disease (CEDAD) together with the International Max Planck Research School - Molecular Biomedicine (IMPRS-MBM). The authors declare that there is no conflict of interest. Not applicable. © The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Shape Memory Polymers Containing Higher Acrylate Content Display Increased Endothelial Cell Attachment

PubMed Central

Govindarajan, Tina; Shandas, Robin

2018-01-01

Shape Memory Polymers (SMPs) are smart materials that can recall their shape upon the application of a stimulus, which makes them appealing materials for a variety of applications, especially in biomedical devices. Most prior SMP research has focused on tuning bulk properties; studying surface effects of SMPs may extend the use of these materials to blood-contacting applications, such as cardiovascular stents, where surfaces that support rapid endothelialization have been correlated to stent success. Here, we evaluate endothelial attachment onto the surfaces of a family of SMPs previously developed in our group that have shown promise for biomedical devices. Nine SMP formulations containing varying amounts of tert-Butyl acrylate (tBA) and Poly(ethylene glycol) dimethacrylate (PEGDMA) were analyzed for endothelial cell attachment. Dynamic mechanical analysis (DMA), contact angle studies, and atomic force microscopy (AFM) were used to verify bulk and surface properties of the SMPs. Human umbilical vein endothelial cell (HUVEC) attachment and viability was verified using fluorescent methods. Endothelial cells preferentially attached to SMPs with higher tBA content, which have rougher, more hydrophobic surfaces. HUVECs also displayed an increased metabolic activity on these high tBA SMPs over the course of the study. This class of SMPs may be promising candidates for next generation blood-contacting devices. PMID:29707382

Stylostome organization in feeding Leptotrombidium larvae (Acariformes: Trombiculidae).

PubMed

Shatrov, Andrew B; Takahashi, Mamoru; Noda, Shinichi; Misumi, Hitoko

2014-01-01

The stylostome of larvae of the trombiculids Leptotrombidium scutellare (Nagayo et al.), Leptotrombidium fletcheri (Womersley et Heaslip) and Leptotrombidium deliense (Walch) was studied experimentally at different time intervals after larval attachment using the histological method. The stylostome of these species has the same organization and belongs to the epidermal combined with the mixed type, developing more in width than in length. Neither transverse nor conspicuous longitudinal layers are present within the stylostome walls, which stain predominantly in red with Azan, also showing longitudinal portions with blue staining. Larvae tend to attach closely to each other and scabs, consisting of the hyperkeratotic epidermal layers fusing with migrating inflammatory cells, develop around the attachment sites. The dermis shows inflammatory foci with dilated capillaries and inflammatory cells inserting in the connective tissue layer underneath the stylostome. The feeding cavity, which is moderately expressed, may be found either in the epidermis or in the dermis. It contains inflammatory cells and their debris in the liquefied host tissues. The stylostome length depends on the character of the attachment site (the thicker epidermis or scab the longer the stylostome), and does not directly correspond to the stages of larval feeding. Nevertheless, at the 48-h time interval, nearly all attached larvae are found to be fully fed and their midgut cells are filled with nutritional globules.

Mentalizing Abilities in Preadolescents' and Their Mothers' Autobiographical Narratives

ERIC Educational Resources Information Center

Scopesi, Alda M.; Rosso, Anna Maria; Viterbori, Paola; Panchieri, Erika

2015-01-01

This study aims to investigate the association between the mental state talk of mothers and their preadolescent children, with the hypothesis that an intergenerational transmission of mentalizing abilities may extend beyond early childhood. The participants were 41 mother-preadolescent child nonclinical dyads. The Adult Attachment Interview (AAI)…

The effective photoinduction of Haematococcus pluvialis for accumulating astaxanthin with attached cultivation.

PubMed

Wan, Minxi; Hou, Dongmei; Li, Yuanguang; Fan, Jianhua; Huang, Jianke; Liang, Songtao; Wang, Weiliang; Pan, Ronghua; Wang, Jun; Li, Shulan

2014-07-01

As the optimal source of astaxanthin, Haematococcus pluvialis was cultured for commercial production of astaxanthin through two continuous phases: cell growth and astaxanthin induction. In this study, the efficiency of an attached system for producing astaxanthin from H. pluvialis was investigated and compared to that of the suspended system (bubble column bioreactor) under various conditions. Results showed that this attached system is more suitable for photoinduction of H. pluvialis than the suspended bioreactor. Under the optimal conditions, the astaxanthin productivity of the attached system was 65.8 mg m(-2)d(-1) and 2.4-fold of that in the suspended system. This attached approach also offers other advantages over suspended systems, such as, producing astaxanthin under a wide range of light intensities and temperatures, saving water, ease to harvest cells, resisting contamination. Therefore, the attached approach can be considered an economical, environmentally friendly and highly-efficient technology for producing astaxanthin from H. pluvialis. Copyright © 2014 Elsevier Ltd. All rights reserved.

STAG2 promotes error correction in mitosis by regulating kinetochore-microtubule attachments.

PubMed

Kleyman, Marianna; Kabeche, Lilian; Compton, Duane A

2014-10-01

Mutations in the STAG2 gene are present in ∼20% of tumors from different tissues of origin. STAG2 encodes a subunit of the cohesin complex, and tumors with loss-of-function mutations are usually aneuploid and display elevated frequencies of lagging chromosomes during anaphase. Lagging chromosomes are a hallmark of chromosomal instability (CIN) arising from persistent errors in kinetochore-microtubule (kMT) attachment. To determine whether the loss of STAG2 increases the rate of formation of kMT attachment errors or decreases the rate of their correction, we examined mitosis in STAG2-deficient cells. STAG2 depletion does not impair bipolar spindle formation or delay mitotic progression. Instead, loss of STAG2 permits excessive centromere stretch along with hyperstabilization of kMT attachments. STAG2-deficient cells display mislocalization of Bub1 kinase, Bub3 and the chromosome passenger complex. Importantly, strategically destabilizing kMT attachments in tumor cells harboring STAG2 mutations by overexpression of the microtubule-destabilizing enzymes MCAK (also known as KIF2C) and Kif2B decreased the rate of lagging chromosomes and reduced the rate of chromosome missegregation. These data demonstrate that STAG2 promotes the correction of kMT attachment errors to ensure faithful chromosome segregation during mitosis. © 2014. Published by The Company of Biologists Ltd.

Three-photon-excited luminescence from unsymmetrical cyanostilbene aggregates: morphology tuning and targeted bioimaging.

PubMed

Mandal, Amal Kumar; Sreejith, Sivaramapanicker; He, Tingchao; Maji, Swarup Kumar; Wang, Xiao-Jun; Ong, Shi Li; Joseph, James; Sun, Handong; Zhao, Yanli

2015-05-26

We report an experimental observation of aggregation-induced enhanced luminescence upon three-photon excitation in aggregates formed from a class of unsymmetrical cyanostilbene derivatives. Changing side chains (-CH3, -C6H13, -C7H15O3, and folic acid) attached to the cyanostilbene core leads to instantaneous formation of aggregates with sizes ranging from micrometer to nanometer scale in aqueous conditions. The crystal structure of a derivative with a methyl side chain reveals the planarization in the unsymmetrical cyanostilbene core, causing luminescence from corresponding aggregates upon three-photon excitation. Furthermore, folic acid attached cyanostilbene forms well-dispersed spherical nanoaggregates that show a high three-photon cross-section of 6.0 × 10(-80) cm(6) s(2) photon(-2) and high luminescence quantum yield in water. In order to demonstrate the targeted bioimaging capability of the nanoaggregates, three cell lines (HEK293 healthy cell line, MCF7 cancerous cell line, and HeLa cancerous cell line) were employed for the investigations on the basis of their different folate receptor expression level. Two kinds of nanoaggregates with and without the folic acid targeting ligand were chosen for three-photon bioimaging studies. The cell viability of three types of cells incubated with high concentration of nanoaggregates still remained above 70% after 24 h. It was observed that the nanoaggregates without the folic acid unit could not undergo the endocytosis by both healthy and cancerous cell lines. No obvious endocytosis of folic acid attached nanoaggregates was observed from the HEK293 and MCF7 cell lines having a low expression of the folate receptor. Interestingly, a significant amount of endocytosis and internalization of folic acid attached nanoaggregates was observed from HeLa cells with a high expression of the folate receptor under three-photon excitation, indicating targeted bioimaging of folic acid attached nanoaggregates to the cancer cell line. This study presents a paradigm of using organic nanoaggregates for targeted three-photon bioimaging.

Control of proliferation and osteogenic differentiation of human dental-pulp-derived stem cells by distinct surface structures.

PubMed

Kolind, K; Kraft, D; Bøggild, T; Duch, M; Lovmand, J; Pedersen, F S; Bindslev, D A; Bünger, C E; Foss, M; Besenbacher, F

2014-02-01

The ability to control the behavior of stem cells provides crucial benefits, for example, in tissue engineering and toxicity/drug screening, which utilize the stem cell's capacity to engineer new tissues for regenerative purposes and the testing of new drugs in vitro. Recently, surface topography has been shown to influence stem cell differentiation; however, general trends are often difficult to establish due to differences in length scales, surface chemistries and detailed surface topographies. Here we apply a highly versatile screening approach to analyze the interplay of surface topographical parameters on cell attachment, morphology, proliferation and osteogenic differentiation of human mesenchymal dental-pulp-derived stem cells (DPSCs) cultured with and without osteogenic differentiation factors in the medium (ODM). Increasing the inter-pillar gap size from 1 to 6 μm for surfaces with small pillar sizes of 1 and 2 μm resulted in decreased proliferation and in more elongated cells with long pseudopodial protrusions. The same alterations of pillar topography, up to an inter-pillar gap size of 4 μm, also resulted in enhanced mineralization of DPSCs cultured without ODM, while no significant trend was observed for DPSCs cultured with ODM. Generally, cells cultured without ODM had a larger deposition of osteogenic markers on structured surfaces relative to the unstructured surfaces than what was found when culturing with ODM. We conclude that the topographical design of biomaterials can be optimized for the regulation of DPSC differentiation and speculate that the inclusion of ODM alters the ability of the cells to sense surface topographical cues. These results are essential in order to transfer the use of this highly proliferative, easily accessible stem cell into the clinic for use in cell therapy and regenerative medicine. Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Biofilm initiation and growth of Pseudomonas aeruginosa on 316L stainless steel in low gravity in orbital space flight

NASA Astrophysics Data System (ADS)

Todd, Paul; Pierson, Duane L.; Allen, Britt; Silverstein, JoAnn

The formation of biofilms by water microorganisms such as Pseudomonas aeruginosa in spacecraft water systems has been a matter of concern for long-duration space flight. Crewed spacecraft plumbing includes internal surfaces made of 316L stainless steel. Experiments were therefore undertaken to compare the ability of P. aeruginosa to grow in suspension, attach to stainless steel and to grow on stainless steel in low gravity on the space shuttle. Four categories of cultures were studied during two space shuttle flights (STS-69 and STS-77). Cultures on the ground were held in static horizontal or vertical cylindrical containers or were tumbled on a clinostat and activated under conditions identical to those for the flown cultures. The containers used on the ground and in flight were BioServe Space Technologies’ Fluid Processing Apparatus (FPA), an open-ended test tube with rubber septa that allows robotic addition of bacteria to culture media to initiate experiments and the addition of fixative to conclude experiments. Planktonic growth was monitored by spectrophotometry, and biofilms were characterized quantitatively by epifluorescence and scanning electron microscopy. In these experiments it was found that: (1) Planktonic growth in flown cultures was more extensive than in static cultures, as seen repeatedly in the history of space microbiology, and closely resembled the growth of tumbled cultures. (2) Conversely, the attachment of cells in flown cultures was as much as 8 times that in tumbled cultures but not significantly different from that in static horizontal and vertical cultures, consistent with the notion that flowing fluid reduces microbial attachment. (3) The final surface coverage in 8 days was the same for flown and static cultures but less by a factor of 15 in tumbled cultures, where coverage declined during the preceding 4 days. It is concluded that cell attachment to 316L stainless steel in the low gravity of orbital space flight is similar to that found in stagnant cultures at 1 x g. Research was supported by NASA contract NAGW-1197 to the University of Colorado.

Transfer of Fas (CD95) protein from the cell surface to the surface of polystyrene beads coated with anti-Fas antibody clone CH-11

PubMed Central

Sawai, H.; Domae, N.

2010-01-01

Mouse monoclonal anti-Fas (CD95) antibody clone CH-11 has been widely used in research on apoptosis. CH-11 has the ability to bind to Fas protein on cell surface and induce apoptosis. Here, we used polystyrene beads coated with CH-11 to investigate the role of lipid rafts in Fas-mediated apoptosis in SKW6.4 cells. Unexpectedly, by treatment of the cells with CH-11-coated beads Fas protein was detached from cell surface and transferred to the surface of CH-11-coated beads. Western blot analysis showed that Fas protein containing both extracellular and intracellular domains was attached to the beads. Fas protein was not transferred from the cells to the surface of the beads coated with other anti-Fas antibodies or Fas ligand. Similar phenomenon was observed in Jurkat T cells. Furthermore, CH-11-induced apoptosis was suppressed by pretreatment with CH-11-coated beads in Jurkat cells. These results suggest that CH-11 might possess distinct properties on Fas protein compared with other anti-Fas antibodies or Fas ligand, and also suggest that caution should be needed to use polystyrene beads coated with antibodies such as CH-11. PMID:20353915

Specialized activities and expression differences for Clostridium thermocellum biofilm and planktonic cells.

PubMed

Dumitrache, Alexandru; Klingeman, Dawn M; Natzke, Jace; Rodriguez, Miguel; Giannone, Richard J; Hettich, Robert L; Davison, Brian H; Brown, Steven D

2017-02-27

Clostridium (Ruminiclostridium) thermocellum is a model organism for its ability to deconstruct plant biomass and convert the cellulose into ethanol. The bacterium forms biofilms adherent to lignocellulosic feedstocks in a continuous cell-monolayer in order to efficiently break down and uptake cellulose hydrolysates. We developed a novel bioreactor design to generate separate sessile and planktonic cell populations for omics studies. Sessile cells had significantly greater expression of genes involved in catabolism of carbohydrates by glycolysis and pyruvate fermentation, ATP generation by proton gradient, the anabolism of proteins and lipids and cellular functions critical for cell division consistent with substrate replete conditions. Planktonic cells had notably higher gene expression for flagellar motility and chemotaxis, cellulosomal cellulases and anchoring scaffoldins, and a range of stress induced homeostasis mechanisms such as oxidative stress protection by antioxidants and flavoprotein co-factors, methionine repair, Fe-S cluster assembly and repair in redox proteins, cell growth control through tRNA thiolation, recovery of damaged DNA by nucleotide excision repair and removal of terminal proteins by proteases. This study demonstrates that microbial attachment to cellulose substrate produces widespread gene expression changes for critical functions of this organism and provides physiological insights for two cells populations relevant for engineering of industrially-ready phenotypes.

A Review of Cell Adhesion Studies for Biomedical and Biological Applications.

PubMed

Khalili, Amelia Ahmad; Ahmad, Mohd Ridzuan

2015-08-05

Cell adhesion is essential in cell communication and regulation, and is of fundamental importance in the development and maintenance of tissues. The mechanical interactions between a cell and its extracellular matrix (ECM) can influence and control cell behavior and function. The essential function of cell adhesion has created tremendous interests in developing methods for measuring and studying cell adhesion properties. The study of cell adhesion could be categorized into cell adhesion attachment and detachment events. The study of cell adhesion has been widely explored via both events for many important purposes in cellular biology, biomedical, and engineering fields. Cell adhesion attachment and detachment events could be further grouped into the cell population and single cell approach. Various techniques to measure cell adhesion have been applied to many fields of study in order to gain understanding of cell signaling pathways, biomaterial studies for implantable sensors, artificial bone and tooth replacement, the development of tissue-on-a-chip and organ-on-a-chip in tissue engineering, the effects of biochemical treatments and environmental stimuli to the cell adhesion, the potential of drug treatments, cancer metastasis study, and the determination of the adhesion properties of normal and cancerous cells. This review discussed the overview of the available methods to study cell adhesion through attachment and detachment events.

A Review of Cell Adhesion Studies for Biomedical and Biological Applications

PubMed Central

Ahmad Khalili, Amelia; Ahmad, Mohd Ridzuan

2015-01-01

Cell adhesion is essential in cell communication and regulation, and is of fundamental importance in the development and maintenance of tissues. The mechanical interactions between a cell and its extracellular matrix (ECM) can influence and control cell behavior and function. The essential function of cell adhesion has created tremendous interests in developing methods for measuring and studying cell adhesion properties. The study of cell adhesion could be categorized into cell adhesion attachment and detachment events. The study of cell adhesion has been widely explored via both events for many important purposes in cellular biology, biomedical, and engineering fields. Cell adhesion attachment and detachment events could be further grouped into the cell population and single cell approach. Various techniques to measure cell adhesion have been applied to many fields of study in order to gain understanding of cell signaling pathways, biomaterial studies for implantable sensors, artificial bone and tooth replacement, the development of tissue-on-a-chip and organ-on-a-chip in tissue engineering, the effects of biochemical treatments and environmental stimuli to the cell adhesion, the potential of drug treatments, cancer metastasis study, and the determination of the adhesion properties of normal and cancerous cells. This review discussed the overview of the available methods to study cell adhesion through attachment and detachment events. PMID:26251901

Miniature spectrally selective dosimeter

NASA Technical Reports Server (NTRS)

Adams, R. R.; Macconochie, I. O.; Poole, B. D., Jr. (Inventor)

1980-01-01

A miniature spectrally selective dosimeter capable of measuring selected bandwidths of radiation exposure on small mobile areas is described. This is achieved by the combination of photovoltaic detectors, electrochemical integrators (E-cells) and filters in a small compact case which can be easily attached in close proximity to and substantially parallel to the surface being measured. In one embodiment two photovoltaic detectors, two E-cells, and three filters are packaged in a small case with attaching means consisting of a safety pin. In another embodiment, two detectors, one E-cell, three filters are packaged in a small case with attaching means consisting of a clip to clip over a side piece of an eye glass frame.

Unresolved trauma in mothers: intergenerational effects and the role of reorganization

PubMed Central

Iyengar, Udita; Kim, Sohye; Martinez, Sheila; Fonagy, Peter; Strathearn, Lane

2014-01-01

A mother's unresolved trauma may interfere with her ability to sensitively respond to her infant, thus affecting the development of attachment in her own child, and potentially contributing to the intergenerational transmission of trauma. One novel construct within the Dynamic Maturational Model of Attachment and Adaptation (DMM) coding of the Adult Attachment Interview (AAI) is “reorganization,” a process whereby speakers are actively changing their understanding of past and present experiences and moving toward attachment security. We conducted a study of mothers with unresolved trauma, exploring their own attachment classification, attachment outcomes of their children, and the potential effects of reorganization on child attachment. Forty-seven first-time mothers participated in the AAI during pregnancy, and returned with their child at 11 months to assess child attachment using the Strange Situation Procedure. Mothers with and without unresolved trauma were compared. We found that mothers with unresolved trauma had insecure attachment themselves and were more likely to have infants with insecure attachment. However, the one exception was that all of the mothers with unresolved trauma who were reorganizing toward secure attachment had infants with secure attachment. These preliminary findings suggest that mothers who are reorganizing may be able to more sensitively respond to their child's cues, contributing to the development of secure attachment. While our results need to be replicated in a larger cohort, this study is the first to explore the construct of reorganization and its potential relationship with child attachment. If confirmed in future studies, it may provide clinical insight into the intergenerational transmission of insecure attachment within the context of unresolved trauma. PMID:25225490

UV laser-ablated surface textures as potential regulator of cellular response.

PubMed

Chandra, Prafulla; Lai, Karen; Sung, Hak-Joon; Murthy, N Sanjeeva; Kohn, Joachim

2010-06-01

Textured surfaces obtained by UV laser ablation of poly(ethylene terephthalate) films were used to study the effect of shape and spacing of surface features on cellular response. Two distinct patterns, cones and ripples with spacing from 2 to 25 μm, were produced. Surface features with different shapes and spacings were produced by varying pulse repetition rate, laser fluence, and exposure time. The effects of the surface texture parameters, i.e., shape and spacing, on cell attachment, proliferation, and morphology of neonatal human dermal fibroblasts and mouse fibroblasts were studied. Cell attachment was the highest in the regions with cones at ∼4 μm spacing. As feature spacing increased, cell spreading decreased, and the fibroblasts became more circular, indicating a stress-mediated cell shrinkage. This study shows that UV laser ablation is a useful alternative to lithographic techniques to produce surface patterns for controlling cell attachment and growth on biomaterial surfaces.

Bioluminescence Truth Data Measurement and Signature Detection

DTIC Science & Technology

2006-01-01

bioluminescence activity and related forcing factors. Kilroy sensors are shown attached to pilings with the senor system below water and the cell phone based...communications module attached to the top of the piling. A cell phone tower represents communication of data to shore. Also shown are distributed...installation are located based on GPS coordinates telemetered by the cell phone module. Icons point in direction of most recently measured flow and

Inhibition of Ovarian Cancer Chemoresistance and Metastasis with Antagonists of Hyaluronan-CD44-CD147 Interactions

DTIC Science & Technology

2014-07-01

attached but most leucocytes did not. The attached cells were washed and used for FACS analysis as described previously (Slomiany et al., 2009b...SKOV3 cells, Hey-A8-MDR cells and ascites-derived cells were subject to FACS analysis in the same manner, as follows. We first separated live cells...Cancer 11, 254-267. Zucker, S., Hymowitz, M., Rollo, E. E., Mann, R., Conner, C. E., Cao, J., Foda , H. D., Tompkins, D. C. and Toole, B. P. (2001

Enhancing cell migration in shape-memory alginate-collagen composite scaffolds: In vitro and ex vivo assessment for intervertebral disc repair.

PubMed

Guillaume, Olivier; Naqvi, Syeda Masooma; Lennon, Kerri; Buckley, Conor Timothy

2015-04-01

Lower lumbar disc disorders pose a significant problem in an aging society with substantial socioeconomic consequences. Both inner tissue (nucleus pulposus) and outer tissue (annulus fibrosus) of the intervertebral disc are affected by such debilitating disorders and can lead to disc herniation and lower back pain. In this study, we developed an alginate-collagen composite porous scaffold with shape-memory properties to fill defects occurring in annulus fibrosus tissue of degenerated intervertebral discs, which has the potential to be administered using minimal invasive surgery. In the first part of this work, we assessed how collagen incorporation on preformed alginate scaffolds influences the physical properties of the final composite scaffold. We also evaluated the ability of annulus fibrosus cells to attach, migrate, and proliferate on the composite alginate-collagen scaffolds compared to control scaffolds (alginate only). In vitro experiments, performed in intervertebral disc-like microenvironmental conditions (low glucose and low oxygen concentrations), revealed that for alginate only scaffolds, annulus fibrosus cells agglomerated in clusters with limited infiltration and migration capacity. In comparison, for alginate-collagen scaffolds, annulus fibrosus cells readily attached and colonized constructs, while preserving their typical fibroblastic-like cell morphology with spreading behavior and intense cytoskeleton expression. In a second part of this study, we investigated the effects of alginate-collagen scaffold when seeded with bone marrow derived mesenchymal stem cells. In vitro, we observed that alginate-collagen porous scaffolds supported cell proliferation and extracellular matrix deposition (collagen type I), with secretion amplified by the local release of transforming growth factor-β3. In addition, when cultured in ex vivo organ defect model, alginate-collagen scaffolds maintained viability of transplanted mesenchymal stem cells for up to 5 weeks. Taken together, these findings illustrate the advantages of incorporating collagen as a means to enhance cell migration and proliferation in porous scaffolds which could be used to augment tissue repair strategies. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

Hybrid ZnPc@TiO2 nanostructures for targeted photodynamic therapy, bioimaging and doxorubicin delivery.

PubMed

Flak, Dorota; Yate, Luis; Nowaczyk, Grzegorz; Jurga, Stefan

2017-09-01

In this study ZnPc@TiO 2 hybrid nanostructures, both nanoparticles and nanotubes, as potential photosensitizers for the photodynamic therapy, fluorescent bioimaging agents, as well as anti-cancer drug nanocarriers, were prepared via zinc phthalocyanine (ZnPc) deposition on TiO 2 . In order to provide the selectivity of prepared hybrid nanostructures towards cancer cells they were modified with folic acid molecules (FA). The efficient attachment of both ZnPc and FA molecules was confirmed with dynamic light scattering (DLS), zeta potential measurements and X-ray photoelectron spectroscopy (XPS). It was presented that ZnPc and FA attachment has a strong effect on fluorescence emission properties of TiO 2 nanostructures, which can be further used for their simultaneous visualization upon cellular uptake. ZnPc@TiO 2 and FA/ZnPc@TiO 2 hybrid nanotubes were then employed as doxorubicin nanocarriers. It was demonstrated that doxorubicin can be easily loaded on these hybrid nanostructures via an electrostatic interaction and then released. In vitro cytotoxicity and photo-cytotoxic activity studies showed that prepared hybrid nanostructures were selectively targeting to cancer cells. Doxorubicin loaded hybrid nanostructures were significantly more cytotoxic than un-loaded ones and their cytotoxic effect was even more severe upon irradiation. The cellular uptake of prepared hybrid nanostructures and their localization in cells was monitored in vitro in 2D cell culture and tumor-like 3D multicellular culture environment with fluorescent confocal microscopy. These hybrid nanostructures preferentially penetrated into human cervical cancer cells (HeLa) than into normal fibroblasts (MSU-1.1) and were mainly localized within the cell cytoplasm. HeLa cells spheroids were also efficiently labelled by prepared hybrid nanostructures. Fluorescent imaging of Hela cells treated with doxorubicin loaded hybrid nanostructures showed that doxorubicin was effectively delivered into cells, released and evenly distributed in the cytoplasm. In conclusion, prepared hybrid nanostructures exhibit high potential as selective bioimaging agents next to their photodynamic activity and drug delivery ability. Copyright © 2017 Elsevier B.V. All rights reserved.

Caregiving Antecedents of Secure Base Script Knowledge: A Comparative Analysis of Young Adult Attachment Representations

PubMed Central

Steele, Ryan D.; Waters, Theodore E. A.; Bost, Kelly K.; Vaughn, Brian E.; Truitt, Warren; Waters, Harriet S.; Booth-LaForce, Cathryn; Roisman, Glenn I.

2015-01-01

Based on a sub-sample (N = 673) of the NICHD Study of Early Child Care and Youth Development (SECCYD) cohort, this paper reports data from a follow-up assessment at age 18 years on the antecedents of secure base script knowledge, as reflected in the ability to generate narratives in which attachment-related difficulties are recognized, competent help is provided, and the problem is resolved. Secure base script knowledge was (a) modestly to moderately correlated with more well established assessments of adult attachment, (b) associated with mother-child attachment in the first three years of life and with observations of maternal and paternal sensitivity from childhood to adolescence, and (c) partially accounted for associations previously documented in the SECCYD cohort between early caregiving experiences and Adult Attachment Interview states of mind (Booth-LaForce & Roisman, 2014) as well as self-reported attachment styles (Fraley, Roisman, Booth-LaForce, Owen, & Holland, 2013). PMID:25264703

The Laminin Receptor Is a Cellular Attachment Receptor for Classical Swine Fever Virus

PubMed Central

Chen, Jianing; He, Wen-Rui; Shen, Liang; Dong, Hong; Yu, Jiahui; Wang, Xiao; Yu, Shaoxiong; Li, Yongfeng; Li, Su; Luo, Yuzi; Sun, Yuan

2015-01-01

ABSTRACT Classical swine fever virus (CSFV) is the causative agent of classical swine fever (CSF), a highly contagious, economically important viral disease in many countries. The Erns and E2 envelope glycoproteins are responsible for the binding to and entry into the host cell by CSFV. To date, only one cellular receptor, heparan sulfate (HS), has been identified as being involved in CSFV attachment. HS is also present on the surface of various cells that are nonpermissive to CSFV. Hence, there must be another receptor(s) that has been unidentified to date. In this study, we used a set of small interfering RNAs (siRNAs) against a number of porcine cell membrane protein genes to screen cellular proteins involved in CSFV infection. This approach resulted in the identification of several proteins, and of these, the laminin receptor (LamR) has been demonstrated to be a cellular receptor for several viruses. Confocal analysis showed that LamR is colocalized with CSFV virions on the membrane, and a coimmunoprecipitation assay indicated that LamR interacts with the CSFV Erns protein. In inhibition assays, anti-LamR antibodies, soluble laminin, or LamR protein significantly inhibited CSFV infection in a dose-dependent manner. Transduction of PK-15 cells with a recombinant lentivirus expressing LamR yielded higher viral titers. Moreover, an attachment assay demonstrated that LamR functions during virus attachment. We also demonstrate that LamR acts as an alternative attachment receptor, especially in SK6 cells. These results indicate that LamR is a cellular attachment receptor for CSFV. IMPORTANCE Classical swine fever virus (CSFV) is the causative agent of classical swine fever (CSF), an economically important viral disease affecting the pig industry in many countries. To date, only heparan sulfate (HS) has been identified to be an attachment receptor for CSFV. Here, using RNA interference screening with small interfering RNAs (siRNAs) against a number of porcine membrane protein genes, we identified the laminin receptor (LamR) to be another attachment receptor. We demonstrate the involvement of LamR together with HS in virus attachment, and we elucidate the relationship between LamR and HS. LamR also serves as an attachment receptor for many viral pathogens, including dengue virus, a fatal human flavivirus. The study will help to enhance our understanding of the life cycle of flaviviruses and the development of antiviral strategies for flaviviruses. PMID:25694590

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. © 2011 Taylor & Francis

Immobilisation of hydroxyapatite-collagen on polydopamine grafted stainless steel 316L: Coating adhesion and in vitro cells evaluation.

PubMed

Tapsir, Zafirah; Jamaludin, Farah H; Pingguan-Murphy, Belinda; Saidin, Syafiqah

2018-02-01

The utilisation of hydroxyapatite and collagen as bioactive coating materials could enhance cells attachment, proliferation and osseointegration. However, most methods to form crystal hydroxyapatite coating do not allow the incorporation of polymer/organic compound due to production phase of high sintering temperature. In this study, a polydopamine film was used as an intermediate layer to immobilise hydroxyapatite-collagen without the introduction of high sintering temperature. The surface roughness, coating adhesion, bioactivity and osteoblast attachment on the hydroxyapatite-collagen coating were assessed as these properties remains unknown on the polydopamine grafted film. The coating was developed by grafting stainless steel 316L disks with a polydopamine film. Collagen type I fibres were then immobilised on the grafted film, followed by the biomineralisation of hydroxyapatite. The surface roughness and coating adhesion analyses were later performed by using AFM instrument. An Alamar Blue assay was used to determine the cytotoxicity of the coating, while an alkaline phosphatase activity test was conducted to evaluate the osteogenic differentiation of human fetal osteoblasts on the coating. Finally, the morphology of cells attachment on the coating was visualised under FESEM. The highest RMS roughness and coating adhesion were observed on the hydroxyapatite-collagen coating (hydroxyapatite-coll-dopa). The hydroxyapatite-coll-dopa coating was non-toxic to the osteoblast cells with greater cells proliferation, greater level of alkaline phosphate production and more cells attachment. These results indicate that the immobilisation of hydroxyapatite and collagen using an intermediate polydopamine is identical to enhance coating adhesion, osteoblast cells attachment, proliferation and differentiation, and thus could be implemented as a coating material on orthopaedic and dental implants.

Potential for Aedes albopictus and Ochlerotatus j. japonicus to Change the Field Ecology of Arboviruses of Human Health Importance in the Mid-Atlantic Region of the United States

DTIC Science & Technology

2001-10-16

attachment, the viral attachment proteins on the surface of the virion bind to receptors on the microvillar membrane of the mosquito s midgut ...Penetration refers to the process through which the virions enter the midgut cells; arboviruses enter cell through receptor -mediated endocytosis. During...inactivation of the virus by digestive enzymes in the lumen of the midgut , and the absence or reduced number of cellular receptor sites for virus attachment

An innovative approach to attached cultivation of Chlorella vulgaris using different materials.

PubMed

Jafari, Negar; Shafiee Alavijeh, Razieh; Abdolahnejad, Ali; Farrokhzadeh, Hossein; Amin, Mohammad Mehdi; Ebrahimi, Afshin

2018-05-10

This article investigates the innovative attached cultivation of Chlorella vulgaris (C. vulgaris) using different materials as an alternative to high capital techniques of harvesting such as centrifugation, flocculation, and filtration. A simple attached algal cultivation system was proposed that was equipped by 10 submerged supporting materials which can harvest algal cells, efficiently. The effect of operational parameters such as light intensity, the rate of aeration, and auto-harvesting time was investigated. A chip, durable, and abundant cellulosic material (Kaldnes carriers covered by kenafs, KCCKs) was proposed for auto-harvesting C. vulgaris cells. The results revealed that optimum aeration rate, light intensity, and auto-harvesting of microalgal cells were 3.6 vvm, 10,548 W/m 2 , and 12 days, respectively. Six of these KCCKs had the highest biofilm formation percent up to 33%. In this condition, the rate of cell growth increased to 0.6 mg/cm 2 . Therefore, this system can be used for appropriate auto-harvesting of microalgae in the attached growth systems. C. vulgaris biomass composition is valuable for biodiesel, bioethanol, and animal protein production.

Synthesis and characterization of gold nanotube/nanowire-polyurethane composite based on castor oil and polyethylene glycol.

PubMed

Ganji, Yasaman; Kasra, Mehran; Salahshour Kordestani, Soheila; Bagheri Hariri, Mohiedin

2014-09-01

Gold nanotubes/nanowires (GNT/NW) were synthesized by using the template-assisted electrodeposition technique and mixed with castor oil-polyethylene glycol based polyurethane (PU) to fabricate porous composite scaffolds for biomedical application. 100 and 50 ppm of GNT/NW were used to synthesize composites. The composite scaffolds were characterized by Fourier transform infrared spectroscopy, dynamic mechanical thermal analysis, differential scanning calorimetry, and scanning electron microscopy. Cell attachment on polyurethane-GNT/NW composites was investigated using fat-derived mesenchymal stem cells. Addition of 50 or 100 ppm GNT/NW had significant effects on thermal, mechanical, and cell attachment of polyurethane. Higher crosslink density and better cell attachment and proliferation were observed in polyurethane containing 50 ppm GNT/NW. The results revealed that GNT/NW formed hydrogen bonding with the polyurethane matrix and improved the thermomechanical properties of nanocomposites. Compared with pure PU, better cellular attachment on polyurethane-GNT/NW composites was observed resulting from the improved surface properties of composites. Copyright © 2014 Elsevier B.V. All rights reserved.

The C-type Lectin Langerin Functions as a Receptor for Attachment and Infectious Entry of Influenza A Virus

PubMed Central

Ng, Wy Ching; Londrigan, Sarah L.; Nasr, Najla; Cunningham, Anthony L.; Turville, Stuart; Brooks, Andrew G.

2015-01-01

ABSTRACT It is well established that influenza A virus (IAV) attachment to and infection of epithelial cells is dependent on sialic acid (SIA) at the cell surface, although the specific receptors that mediate IAV entry have not been defined and multiple receptors may exist. Lec2 Chinese hamster ovary (CHO) cells are SIA deficient and resistant to IAV infection. Here we demonstrate that the expression of the C-type lectin receptor langerin in Lec2 cells (Lec2-Lg) rendered them permissive to IAV infection, as measured by replication of the viral genome, transcription of viral mRNA, and synthesis of viral proteins. Unlike SIA-dependent infection of parental CHO cells, IAV attachment and infection of Lec2-Lg cells was mediated via lectin-mediated recognition of mannose-rich glycans expressed by the viral hemagglutinin glycoprotein. Lec2 cells expressing endocytosis-defective langerin bound IAV efficiently but remained resistant to IAV infection, confirming that internalization via langerin was essential for infectious entry. Langerin-mediated infection of Lec2-Lg cells was pH and dynamin dependent, occurred via clathrin- and caveolin-mediated endocytic pathways, and utilized early (Rab5+) but not late (Rab7+) endosomes. This study is the first to demonstrate that langerin represents an authentic receptor that binds and internalizes IAV to facilitate infection. Moreover, it describes a unique experimental system to probe specific pathways and compartments involved in infectious entry following recognition of IAV by a single cell surface receptor. IMPORTANCE On the surface of host cells, sialic acid (SIA) functions as the major attachment factor for influenza A viruses (IAV). However, few studies have identified specific transmembrane receptors that bind and internalize IAV to facilitate infection. Here we identify human langerin as a transmembrane glycoprotein that can act as an attachment factor and a bone fide endocytic receptor for IAV infection. Expression of langerin by an SIA-deficient cell line resistant to IAV rendered cells permissive to infection. As langerin represented the sole receptor for IAV infection in this system, we have defined the pathways and compartments involved in infectious entry of IAV into cells following recognition by langerin. PMID:26468543

[Emotional experience of early traumatic experiences and relationship-specific attachment styles in alcohol-dependent patients].

PubMed

Hebein, Ursula; Quantschnig, Bettina; Andreas, Sylke

2018-03-01

Emotional experience of early traumatic experiences and relationship-specific attachment styles in alcohol-dependent patients Objectives: The present investigation studied the differences in emotional experiences among alcohol-dependent patients in inpatient psychotherapy, taking into account their traumatic experiences and relationship-specific attachment styles. Three standardized instruments were used on this clinical sample of 70 patients in inpatient psychotherapy: (1) Scales for Experiencing Emotions (SEE), (2) Childhood Trauma Questionnaire (CTQ), (3) Relationship-Specific Attachment Scales for Adults (RASA). The results showed that traumatic experiences in childhood and adolescence as well as relationship-specific attachment styles were indeed associated with deficits in emotional experience and regulation. Especially emotional abuse was associated with deficits in emotional experience. An insecure attachment style was associated with a lower level of acceptance of emotions, a lower level of self-control, and dysfunctional emotion regulation. In particular, the attachment style to the mother seems to be important. These results suggest that a distinction of alcohol-addicted patients, with respect to attachment styles and traumatic experiences in childhood and adolescent, seems to be important in order to define specific interventions to improve the ability to regulate emotions.

Long-term effects on adult attachment in German occupation children born after World War II in comparison with a birth-cohort-matched representative sample of the German general population.

PubMed

Kaiser, Marie; Kuwert, Philipp; Braehler, Elmar; Glaesmer, Heide

2018-02-01

Children born of war are a phenomenon of every conflict. At the end of World War II and thereafter, approximately 400,000 children were fathered by foreign soldiers and born to local women in Germany. Quantitative research on psychosocial consequences of growing up as German occupation child (GOC) has been missing so far. This study examines adult attachment and its association with current depression in GOC (N = 146) using self-report instruments: Adult Attachment Scale, Patient Health Questionnaire. Data were compared to a birth-cohort-matched representative sample of the German population (BCMS; N = 786). GOC differ in both attachment dimensions (less comfortable with closeness/intimacy, lowered ability to depend on others) and adult attachment (more dismissive and fearful) compared to BCMS. Insecure adult attachment is associated with current depression. GOC grew up under difficult circumstances (e.g. poverty, adverse events, and stigmatization). Even decades later they display more insecure attachment in current relationships. Findings underline the complex and long-term impact of their developmental conditions on attachment and current mental health.

Counselors' attachment anxiety and avoidance and the congruence in clients' and therapists' working alliance ratings.

PubMed

Kivlighan, Dennis M; Marmarosh, Cheri L

2018-07-01

To determine how counselors' attachment anxiety and avoidance related to congruence between counselors' and clients' Working alliance (WA) ratings. Congruence strength was defined as the regression coefficient for clients' WA ratings predicting counselors' WA ratings. Directional bias was defined as the difference in level between counselors' and clients' WA ratings. Twenty-seven graduate student counselors completed an attachment measure and they and their 64 clients completed a measure of WA early in therapy. The truth-and-bias analysis was adapted to analyze the data. As hypothesized counselors' WA ratings were significantly and positively related to clients' WA ratings. Also as hypothesized, counselors' WA ratings were significantly lower than their clients' WA ratings (directional bias). Increasing counselor attachment anxiety was related to increasing negative directional bias; as counselors' attachment anxiety increased the difference between counselors and clients WA ratings became more negative. There was a significant interaction between counselor attachment anxiety and congruence strength in predicting counselor WA ratings. There was a stronger relationship between client WA ratings and counselor WA ratings for counselors low versus high in attachment anxiety. Counselors' attachment anxiety is realted to their ability to accurately percieve their clients' WA.

Functionalized magnetic-fluorescent hybrid nanoparticles for cell labelling.

PubMed

Lou, Lei; Yu, Ke; Zhang, Zhengli; Li, Bo; Zhu, Jianzhong; Wang, Yiting; Huang, Rong; Zhu, Ziqiang

2011-05-01

A facile method of synthesizing 60 nm magnetic-fluorescent core-shell bifunctional nanocomposites with the ability to label cells is presented. Hydrophobic trioctylphosphine oxide (TOPO)-capped CdSe@ZnS quantum dots (QDs) were assembled on polyethyleneimine (PEI)-coated Fe(3)O(4) nanoparticles (MNP). Polyethyleneimine was utilized for the realization of multifunction, including attaching 4 nm TOPO capped CdSe@ZnS quantum dots onto magnetite particles, altering the surface properties of quantum dots from hydrophobic to hydrophilic as well as preventing the formation of large aggregates. Results show that these water-soluble hybrid nanocomposites exhibit good colloidal stability and retain good magnetic and fluorescent properties. Because TOPO-capped QDs are assembled instead of their water-soluble equivalents, the nanocomposites are still highly luminescent with no shift in the PL peak position and present long-term fluorescence stability. Moreover, TAT peptide (GRKKRRQRRRPQ) functionalized hybrid nanoparticles were also studied due to their combined magnetic enrichment and optical detection for cell separation and rapid cell labelling. A cell viability assay revealed good biocompatibility of these hybrid nanoparticles. The potential application of the new magnetic-fluorescent nanocomposites in biological and medicine is demonstrated. © The Royal Society of Chemistry 2011

Calcium pectate chemistry causes growth to be stored in Chara corallina: a test of the pectate cycle.

PubMed

Proseus, Timothy E; Boyer, John S

2008-08-01

Calcium pectate chemistry was reported to control the growth rate of cells of Chara corallina, and required turgor pressure (P) to do so. Accordingly, this chemistry should account for other aspects of growth, particularly the ability of plants to compensate for brief exposure to low P, that is, to 'store' growth. Live Chara cells or isolated walls were attached to a pressure probe, and P was varied. Low P caused growth to be inhibited in live cells, but when P returned to normal (0.5 MPa), a flush of growth completely compensated for that lost at low P for as long as 23-53 min. This growth storage was absent in isolated walls, mature cells and live cells exposed to cold, indicating that the cytoplasm delivered a metabolically derived growth factor needing P for its action. Because the cytoplasm delivered pectate needing P for its action, pectate was supplied to isolated walls at low P as though the cytoplasm had done so. Growth was stored while otherwise none occurred. It was concluded that a P-dependent cycle of calcium pectate chemistry not only controlled growth rate and new wall deposition, but also accounted for stored growth.

Mechanisms of erosion of atherosclerotic plaques.

PubMed

Quillard, Thibaut; Franck, Grégory; Mawson, Thomas; Folco, Eduardo; Libby, Peter

2017-10-01

The present review explores the mechanisms of superficial intimal erosion, a common cause of thrombotic complications of atherosclerosis. Human coronary artery atheroma that give rise to thrombosis because of erosion differ diametrically from those associated with fibrous cap rupture. Eroded lesions characteristically contain few inflammatory cells, abundant extracellular matrix, and neutrophil extracellular traps (NETs). Innate immune mechanisms such as engagement of Toll-like receptor 2 (TLR2) on cultured endothelial cells can impair their viability, attachment, and ability to recover a wound. Hyaluronan fragments may serve as endogenous TLR2 ligands. Mouse experiments demonstrate that flow disturbance in arteries with neointimas tailored to resemble features of human eroded plaques disturbs endothelial cell barrier function, impairs endothelial cell viability, recruits neutrophils, and provokes endothelial cells desquamation, NET formation, and thrombosis in a TLR2-dependent manner. Mechanisms of erosion have received much less attention than those that provoke plaque rupture. Intensive statin treatment changes the characteristic of plaques that render them less susceptible to rupture. Thus, erosion may contribute importantly to the current residual burden of risk. Understanding the mechanisms of erosion may inform the development and deployment of novel therapies to combat the remaining atherothrombotic risk in the statin era.

Characterization of a 3rd Generation Lentiviral Vector Pseudotyped With Nipah Virus Envelope Proteins For Endothelial Cell Transduction

PubMed Central

Witting, Scott R.; Vallanda, Priya; Gamble, Aisha L.

2013-01-01

Lentiviruses are becoming progressively more popular as gene therapy vectors due to their ability to integrate into quiescent cells and recent clinical trial successes. Directing these vectors to specific cell types and limiting off-target transduction in vivo remains a challenge. Replacing the viral envelope proteins responsible for cellular binding, or pseudotyping, remains a common method to improve lentiviral targeting. Here, we describe the development of a high titer, 3rd generation lentiviral vector pseudotyped with Nipah virus fusion protein (NiV-F) and attachment protein (NiV-G). Critical to high titers was truncation of the cytoplasmic domains of both NiV-F and NiV-G. As known targets of wild-type Nipah virus, primary endothelial cells are shown to be effectively transduced by the Nipah pseudotype. In contrast, human CD34+ hematopoietic progenitors were not significantly transduced. Additionally, the Nipah pseudotype has increased stability in human serum compared to VSV pseudotyped lentivirus. These findings suggest that the use of Nipah virus envelope proteins in 3rd generation lentiviral vectors would be a valuable tool for gene delivery targeted to endothelial cells. PMID:23698741

Characterization of a third generation lentiviral vector pseudotyped with Nipah virus envelope proteins for endothelial cell transduction.

PubMed

Witting, S R; Vallanda, P; Gamble, A L

2013-10-01

Lentiviruses are becoming progressively more popular as gene therapy vectors due to their ability to integrate into quiescent cells and recent clinical trial successes. Directing these vectors to specific cell types and limiting off-target transduction in vivo remains a challenge. Replacing the viral envelope proteins responsible for cellular binding, or pseudotyping, remains a common method to improve lentiviral targeting. Here, we describe the development of a high titer, third generation lentiviral vector pseudotyped with Nipah virus fusion protein (NiV-F) and attachment protein (NiV-G). Critical to high titers was truncation of the cytoplasmic domains of both NiV-F and NiV-G. As known targets of wild-type Nipah virus, primary endothelial cells are shown to be effectively transduced by the Nipah pseudotype. In contrast, human CD34+ hematopoietic progenitors were not significantly transduced. Additionally, the Nipah pseudotype has increased stability in human serum compared with vesicular stomatitis virus pseudotyped lentivirus. These findings suggest that the use of Nipah virus envelope proteins in third generation lentiviral vectors would be a valuable tool for gene delivery targeted to endothelial cells.

Surface properties of Streptococcus phocae strains isolated from diseased Atlantic salmon, Salmo salar L.

PubMed

González-Contreras, A; Magariños, B; Godoy, M; Irgang, R; Toranzo, A E; Avendaño-Herrera, R

2011-03-01

Streptococcus phocae is an emerging pathogen for Chilean Atlantic salmon, Salmo salar, but the factors determining its virulence are not yet elucidated. In this work, cell surface-related properties such as hydrophobicity and haemagglutination, adhesion to mucus and cell lines, capsule detection, survival and biofilm formation in skin mucus and serum resistance of the isolates responsible for outbreaks in Atlantic salmon and seals were examined. Adhesion to hydrocarbons and the results of salt aggregation tests indicated most of the S. phocae were strongly hydrophobic. All isolates exhibited a similar ability to attach to the Chinook salmon embryo (CHSE) cells line, but were not able to enter CHSE cells. Haemagglutination was not detected. Our data clearly indicate that S. phocae can resist the killing activity of mucus and serum and proliferate in them, which could be associated with the presence of a capsular layer around the cells. Pathogenicity studies using seal and fish isolates demonstrated mortality or pathological signs in fish injected only with the Atlantic salmon isolate. No mortalities or histopathological alterations were observed in fish injected with extracellular products. © 2011 Blackwell Publishing Ltd.

Phenotypic Heterogeneity and the Evolution of Bacterial Life Cycles.

PubMed

van Gestel, Jordi; Nowak, Martin A

2016-02-01

Most bacteria live in colonies, where they often express different cell types. The ecological significance of these cell types and their evolutionary origin are often unknown. Here, we study the evolution of cell differentiation in the context of surface colonization. We particularly focus on the evolution of a 'sticky' cell type that is required for surface attachment, but is costly to express. The sticky cells not only facilitate their own attachment, but also that of non-sticky cells. Using individual-based simulations, we show that surface colonization rapidly evolves and in most cases leads to phenotypic heterogeneity, in which sticky and non-sticky cells occur side by side on the surface. In the presence of regulation, cell differentiation leads to a remarkable set of bacterial life cycles, in which cells alternate between living in the liquid and living on the surface. The dominant life stage is formed by the surface-attached colony that shows many complex features: colonies reproduce via fission and by producing migratory propagules; cells inside the colony divide labour; and colonies can produce filaments to facilitate expansion. Overall, our model illustrates how the evolution of an adhesive cell type goes hand in hand with the evolution of complex bacterial life cycles.

Attachment defect in mouse fibroblasts (L cells) persistently infected with Chlamydia psittaci.

PubMed Central

Moulder, J W; Levy, N J; Zeichner, S L; Lee, C K

1981-01-01

Almost all the cells in populations of mouse fibroblasts (L cells) persistently infected with the 6BC strain of Chlamydia psittaci were immune to superinfection with high multiplicities of C. psittaci, whether or not the L cells contained visible chlamydial inclusions. As ascertained by experiments with 14C-labeled C. psittaci, immunity to superinfection resulted from the failure of added chlamydiae to attach to persistently infected host cells. However, when exogenous C. psittaci was introduced into persistently infected L cells by centrifuging the inoculum onto host cell monolayers or by pretreating the monolayers with diethylaminoethyl-dextran, these chlamydiae produced expected numbers of infectious progeny. Persistently infected L cells were associated in an unknown way with a C. psittaci population that entered the host cells only with the aid of centrifugation or pretreatment with diethylaminoethyl-dextran. Inclusion-free, persistently infected L cells appeared to present at least two separate hindrances to chlamydial activity: blockage of the attachment of exogenous elementary bodies to persistently infected host cells and prevention of the initiation of chlamydial multiplication by means of a normal developmental cycle in the absence of added C. psittaci. Images PMID:7298188

Differentiation of anchoring junctions in tracheal basal cells in the growing rat.

PubMed

Evans, M J; Cox, R A; Burke, A S; Moller, P C

1992-02-01

A function of airway basal cells is to attach ciliated and nonciliated columnar cells to the basal lamina. The significance of the basal cell in attachment is related to the height of the columnar epithelium. In taller epithelia, basal cells are more numerous and differentiated with respect to anchoring junctional adhesion mechanisms (desmosomes, hemidesmosomes, and the cytoskeleton) than in shorter epithelia. In this study, we determined if basal cell anchoring junctional adhesion mechanisms differentiated during growth of the airway. Tracheas from five 3-day-old, five 30-day-old, and five 90-day-old rats were prepared for electron microscopy and morphometrically studied by standard techniques. The circumference of the trachea increased from 2.5 +/- 0.2 to 7.5 +/- 0.4 mm during growth. The height of the columnar cell increased from 13.4 +/- 1.5 to 24.6 +/- 3.9 microns, and the number of basal cells per millimeter increased from 3.2 +/- 0.7 to 9.6 +/- 1.8 during growth. The number of desmosomes per basal cell profile increased significantly from 1.5 +/- 0.1 to 2.1 +/- 0.1, as did keratin filament volume density from 0.046 +/- 0.05 to 0.098 +/- 0.032. The amount of hemidesmosome attachment per basal cell did not increase significantly during growth of the airway. These data demonstrate that as tracheas grow in circumference, the columnar cells increase in height, basal cells increase in number, and anchoring junctional adhesion mechanisms differentiate in the basal cells. These changes are closely related to the height of the epithelium and result in maintaining a constant amount of attachment between the columnar epithelium and the basal lamina as the epithelium increases in height.

Reducing background noise in near-infrared medical imaging: Routes to activated fluorescing

NASA Astrophysics Data System (ADS)

Burdette, Mary K.; Bandera, Yuriy; Powell, Rhonda R.; Bruce, Terri F.; Foulger, Stephen H.

2016-03-01

Activated fluorescence was achieved for nanoparticle based systems. One particulate system consisting of a poly(propargyl acrylate) (PA) core with covalently attached derivatized fluorescein and modified bovine serum albumin covalently conjugated to a cyanine 3 derivative was initially nonfluorescent. Upon trypsin addition and subsequent proteolytic digestion, Förster resonance energy transfer (FRET) was induced. The other particulate system consisted of a PA core with covalently attached azide modified BSA, which was covalently attached to a silicon phthalocyanine derivative (PA/BSA/akSiPc600). Both systems were biocompatible. To investigate activated fluorescence with the PA/BSA/akSiPc600 system in cancer cells, human non-small cell lung cancer cells (A549 cell line) were used as a model system. The PA/BSA/akSiPc600 system was incubated with the cells at varying time points in an effort to see a fluorescence increase over time as the cells uptake the particles and as they digest the BSA, most probably, via endocytosis. It was seen, through live cell scanning confocal microscopy, that the fluorescence was activated in the cell.

Influence of various superhydrophilic treatments of titanium on the initial attachment, proliferation, and differentiation of osteoblast-like cells.

PubMed

Yamamura, Keisuke; Miura, Tadashi; Kou, I; Muramatsu, Takashi; Furusawa, Masahiro; Yoshinari, Masao

2015-01-01

The purpose of this study was to investigate the influence of superhydrophilic treatments of titanium on the behavior of osteoblastlike cells. Superhydrophilic specimens were prepared with sandblast and acid-etching (DW), oxygen plasma (Plasma) and ultraviolet light (UV), and were stored in distilled water for 3 days immediately after these treatments. Specimens stored in air for 3 weeks were used as a control Air group. Initial cell attachment, proliferation, alkaline phosphatase activity, and osteocalcin secretion of mouse osteoblast-like cells MC3T3-E1 were enhanced more on superhydrophilic groups than were Air specimens. On confocal laser scanning microscope images of cell morphology, the expression of actin filaments was observed on the superhydrophilic groups, whereas relatively little actin filament expression was seen on the Air surfaces on all culture periods. These results indicate that DW, Plasma, or UV treatment has potential for the creation and maintenance of superhydrophilic surfaces and the enhancement of the initial attachment, proliferation, and differentiation of osteoblast-like cells.

Toxoplasma Actin Is Required for Efficient Host Cell Invasion.

PubMed

Drewry, Lisa L; Sibley, L David

2015-06-16

Apicomplexan parasites actively invade host cells using a mechanism predicted to be powered by a parasite actin-dependent myosin motor. In the model apicomplexan Toxoplasma gondii, inducible knockout of the actin gene, ACT1, was recently demonstrated to limit but not completely abolish invasion. This observation has led to the provocative suggestion that T. gondii possesses alternative, ACT1-independent invasion pathways. Here, we dissected the residual invasive ability of Δact1 parasites. Surprisingly, we were able to detect residual ACT1 protein in inducible Δact1 parasites as long as 5 days after ACT1 deletion. We further found that the longer Δact1 parasites were propagated after ACT1 deletion, the more severe an invasion defect was observed. Both findings are consistent with the quantity of residual ACT1 retained in Δact1 parasites being responsible for their invasive ability. Furthermore, invasion by the Δact1 parasites was also sensitive to the actin polymerization inhibitor cytochalasin D. Finally, there was no clear defect in attachment to host cells or moving junction formation by Δact1 parasites. However, Δact1 parasites often exhibited delayed entry into host cells, suggesting a defect specific to the penetration stage of invasion. Overall, our results support a model where residual ACT1 protein retained in inducible Δact1 parasites facilitates their limited invasive ability and confirm that parasite actin is essential for efficient penetration into host cells during invasion. The prevailing model for apicomplexan invasion has recently been suggested to require major revision, based on studies where core components of the invasion machinery were genetically disrupted using a Cre-Lox-based inducible knockout system. For the myosin component of the motor thought to power invasion, an alternative parasite myosin was recently demonstrated to functionally compensate for loss of the primary myosin involved in invasion. Here, we highlight a second mechanism that can account for the surprising ability of parasites to invade after genetic disruption of core invasion machinery. Specifically, residual actin protein present in inducible knockout parasites appears able to support their limited invasion of host cells. Our results have important implications for the interpretation of the apicomplexan invasion model and also highlight significant considerations when analyzing the phenotypes of inducible knockout parasites generated using Cre-Lox technology. Copyright © 2015 Drewry and Sibley.

Final Report: Rational Design of Anode Surface Chemistry in Microbial Fuel Cells for Improved Exoelectrogen Attachment and Electron Transfer

DTIC Science & Technology

2015-12-21

SECURITY CLASSIFICATION OF: The overall goal of this project is to determine how electrode surface chemistry can be rationally designed to decrease...2015 Approved for Public Release; Distribution Unlimited Final Report: Rational Design of Anode Surface Chemistry in Microbial Fuel Cells for...ABSTRACT Final Report: Rational Design of Anode Surface Chemistry in Microbial Fuel Cells for Improved Exoelectrogen Attachment and Electron Transfer

Low childhood subjective social status and telomere length in adulthood: The role of attachment orientations.

PubMed

Murdock, Kyle W; Seiler, Annina; Chirinos, Diana A; Garcini, Luz M; Acebo, Sally L; Cohen, Sheldon; Fagundes, Christopher P

2018-04-01

Low subjective social status (SSS) in childhood places one at greater risk of a number of health problems in adulthood. Theoretical and empirical evidence indicates that exposure to supportive parenting may buffer the negative effects of low childhood SSS on adult health. Given the importance of supportive caregivers and close others for the development of attachment orientations throughout the lifespan, attachment theory may be important for understanding why some individuals are resilient to the negative effects of low childhood SSS on adult health while others are not. We examined if attachment anxiety and attachment avoidance altered the association between childhood subjective social status (SSS) and length of telomeres in white blood cells in adulthood. Shorter telomere length is associated with increased risk of age-related diseases including cancer, type 2 diabetes, and cardiovascular disease. Participants (N = 128) completed self-report measures of childhood SSS and attachment orientations, as well as a blood draw. We found that among those with low childhood SSS, low attachment anxiety was associated with longer telomere length in white blood cells in comparison to high attachment anxiety controlling for participant age, sex, race, body mass index, and adult SSS. Among those with high childhood SSS, low attachment anxiety was associated with a slight decrease in telomere length. Attachment avoidance was unrelated to length of telomeres. Such findings provide further evidence for the role that close relationships may have on buffering SSS related health disparities. © 2018 Wiley Periodicals, Inc.

SERS-fluorescence joint spectral encoded magnetic nanoprobes for multiplex cancer cell separation.

PubMed

Wang, Zhuyuan; Zong, Shenfei; Chen, Hui; Wang, Chunlei; Xu, Shuhong; Cui, Yiping

2014-11-01

A new kind of cancer cell separation method is demonstrated, using surface-enhanced Raman scattering (SERS) and fluorescence dual-encoded magnetic nanoprobes. The designed nanoprobes can realize SERS-fluorescence joint spectral encoding (SFJSE) and greatly improve the multiplexing ability. The nanoprobes have four main components, that is, the magnetic core, SERS generator, fluorescent agent, and targeting antibody. These components are assembled with a multi-layered structure to form the nanoprobes. Specifically, silica-coated magnetic nanobeads (MBs) are used as the inner core. Au core-Ag shell nanorods (Au@Ag NRs) are employed as the SERS generators and attached on the silica-coated MBs. After burying these Au@Ag NRs with another silica layer, CdTe quantum dots (QDs), that is, the fluorescent agent, are anchored onto the silica layer. Finally, antibodies are covalently linked to CdTe QDs. SFJSE is fulfilled by using different Raman molecules and QDs with different emission wavelengths. By utilizing four human cancer cell lines and one normal cell line as the model cells, the nanoprobes can specifically and simultaneously separate target cancer cells from the normal ones. This SFJSE-based method greatly facilitates the multiplex, rapid, and accurate cancer cell separation, and has a prosperous potential in high-throughput analysis and cancer diagnosis. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The susceptibility of Staphylococcus aureus CIP 65.8 and Pseudomonas aeruginosa ATCC 9721 cells to the bactericidal action of nanostructured Calopteryx haemorrhoidalis damselfly wing surfaces.

PubMed

Truong, Vi Khanh; Geeganagamage, Nipuni Mahanamanam; Baulin, Vladimir A; Vongsvivut, Jitraporn; Tobin, Mark J; Luque, Pere; Crawford, Russell J; Ivanova, Elena P

2017-06-01

Nanostructured insect wing surfaces have been reported to possess the ability to resist bacterial colonization through the mechanical rupture of bacterial cells coming into contact with the surface. In this work, the susceptibility of physiologically young, mature and old Staphylococcus aureus CIP 65.8 and Pseudomonas aeruginosa ATCC 9721 bacterial cells, to the action of the bactericidal nano-pattern of damselfly Calopteryx haemorrhoidalis wing surfaces, was investigated. The results were obtained using several surface characterization techniques including optical profilometry, scanning electron microscopy, synchrotron-sourced Fourier transform infrared microspectroscopy, water contact angle measurements and antibacterial assays. The data indicated that the attachment propensity of physiologically young S. aureus CIP 65.8 T and mature P. aeruginosa ATCC 9721 bacterial cells was greater than that of the cells at other stages of growth. Both the S. aureus CIP 65.8 T and P. aeruginosa ATCC 9721 cells, grown at the early (1 h) and late stationary phase (24 h), were found to be most susceptible to the action of the wings, with up to 89.7 and 61.3% as well as 97.9 and 97.1% dead cells resulting from contact with the wing surface, respectively.

Gold Nano Popcorn Attached SWCNT Hybrid Nanomaterial for Targeted Diagnosis and Photothermal Therapy of Human Breast Cancer Cells

PubMed Central

Beqa, Lule; Fan, Zhen; Singh, Anant Kumar; Senapati, Dulal; Ray, Paresh Chandra

2011-01-01

Breast cancer presents greatest challenge in health care in today’s world. The key to ultimately successful treatment of breast cancer disease is an early and accurate diagnosis. Current breast cancer treatments are often associated with severe side effects. Driven by the need, we report the design of novel hybrid nanomaterial using gold nano popcorn-attached single wall carbon nanotube for targeted diagnosis and selective photothermal treatment. Targeted SK-BR-3 human breast cancer cell sensing have been performed in 10 cancer cells/mL level, using surface enhanced Raman scattering of single walls carbon nanotube’s D and G bands. Our data show that S6 aptamer attached hybrid nanomaterial based SERS assay is highly sensitive to targeted human breast cancer SK-BR-3 cell line and it will be able to distinguish it from other non targeted MDA-MB breast cancer cell line and HaCaT normal skin cell line. Our results also show that 10 minutes of photothermal therapy treatment by 1.5 W/cm2 power, 785 nm laser is enough to kill cancer cells very effectively using S6 aptamer attached hybrid nanomaterials. Possible mechanisms for targeted sensing and operating principle for highly efficient photothermal therapy have been discussed. Our experimental results reported here open up a new possibility for using aptamers modified hybrid nanomaterial for reliable diagnosis and targeted therapy of cancer cell lines quickly. PMID:21842867

Gold nano-popcorn attached SWCNT hybrid nanomaterial for targeted diagnosis and photothermal therapy of human breast cancer cells.

PubMed

Beqa, Lule; Fan, Zhen; Singh, Anant Kumar; Senapati, Dulal; Ray, Paresh Chandra

2011-09-01

Breast cancer presents greatest challenge in health care in today's world. The key to ultimately successful treatment of breast cancer disease is an early and accurate diagnosis. Current breast cancer treatments are often associated with severe side effects. Driven by the need, we report the design of novel hybrid nanomaterial using gold nano popcorn-attached single wall carbon nanotube for targeted diagnosis and selective photothermal treatment. Targeted SK-BR-3 human breast cancer cell sensing have been performed in 10 cancer cells/mL level, using surface enhanced Raman scattering of single walls carbon nanotube's D and G bands. Our data show that S6 aptamer attached hybrid nanomaterial based SERS assay is highly sensitive to targeted human breast cancer SK-BR-3 cell line and it will be able to distinguish it from other non targeted MDA-MB breast cancer cell line and HaCaT normal skin cell line. Our results also show that 10 min of photothermal therapy treatment by 1.5 W/cm(2) power, 785 nm laser is enough to kill cancer cells very effectively using S6 aptamer attached hybrid nanomaterials. Possible mechanisms for targeted sensing and operating principle for highly efficient photothermal therapy have been discussed. Our experimental results reported here open up a new possibility for using aptamers modified hybrid nanomaterial for reliable diagnosis and targeted therapy of cancer cell lines quickly.

Reconsidering English Grammar Teaching for Improving Non-English Majors' English Writing Ability

ERIC Educational Resources Information Center

Shen, Yuru

2012-01-01

With the globalization of world economy, English learners' writing ability has been attached less and less importance. As a result, many college students in China, especially the non-English majors, cannot express themselves effectively in written English. They make various kinds of mistakes, mostly grammar mistakes, such as writing sentence…

Assembling high activity phosphotriesterase composites using hybrid nanoparticle peptide-DNA scaffolded architectures

NASA Astrophysics Data System (ADS)

Breger, Joyce C.; Buckhout-White, Susan; Walper, Scott A.; Oh, Eunkeu; Susumu, Kimihiro; Ancona, Mario G.; Medintz, Igor L.

2017-06-01

Nanoparticle (NP) display potentially offers a new way to both stabilize and, in many cases, enhance enzyme activity over that seen for native protein in solution. However, the large, globular and sometimes multimeric nature of many enzymes limits their ability to attach directly to the surface of NPs, especially when the latter are colloidally stabilized with bulky PEGylated ligands. Engineering extended protein linkers into the enzymes to achieve direct attachment through the PEG surface often detrimentally alters the enzymes catalytic ability. Here, we demonstrate an alternate, hybrid biomaterials-based approach to achieving directed enzyme assembly on PEGylated NPs. We self-assemble a unique architecture consisting of a central semiconductor quantum dot (QD) scaffold displaying controlled ratios of extended peptide-DNA linkers which penetrate through the PEG surface to directly couple enzymes to the QD surface. As a test case, we utilize phosphotriesterase (PTE), an enzyme of bio-defense interest due to its ability to hydrolyze organophosphate nerve agents. Moreover, this unique approach still allows PTE to maintain enhanced activity while also suggesting the ability of DNA to enhance enzyme activity in and of itself.

Deformation simulation of cells seeded on a collagen-GAG scaffold in a flow perfusion bioreactor using a sequential 3D CFD-elastostatics model.

PubMed

Jungreuthmayer, C; Jaasma, M J; Al-Munajjed, A A; Zanghellini, J; Kelly, D J; O'Brien, F J

2009-05-01

Tissue-engineered bone shows promise in meeting the huge demand for bone grafts caused by up to 4 million bone replacement procedures per year, worldwide. State-of-the-art bone tissue engineering strategies use flow perfusion bioreactors to apply biophysical stimuli to cells seeded on scaffolds and to grow tissue suitable for implantation into the patient's body. The aim of this study was to quantify the deformation of cells seeded on a collagen-GAG scaffold which was perfused by culture medium inside a flow perfusion bioreactor. Using a microCT scan of an unseeded collagen-GAG scaffold, a sequential 3D CFD-deformation model was developed. The wall shear stress and the hydrostatic wall pressure acting on the cells were computed through the use of a CFD simulation and fed into a linear elastostatics model in order to calculate the deformation of the cells. The model used numerically seeded cells of two common morphologies where cells are either attached flatly on the scaffold wall or bridging two struts of the scaffold. Our study showed that the displacement of the cells is primarily determined by the cell morphology. Although cells of both attachment profiles were subjected to the same mechanical load, cells bridging two struts experienced a deformation up to 500 times higher than cells only attached to one strut. As the scaffold's pore size determines both the mechanical load and the type of attachment, the design of an optimal scaffold must take into account the interplay of these two features and requires a design process that optimizes both parameters at the same time.

An adaptor role for cytoplasmic Sam68 in modulating Src activity during cell polarization.

PubMed

Huot, Marc-Etienne; Brown, Claire M; Lamarche-Vane, Nathalie; Richard, Stéphane

2009-04-01

The Src-associated substrate during mitosis with a molecular mass of 68 kDa (Sam68) is predominantly nuclear and is known to associate with proteins containing the Src homology 3 (SH3) and SH2 domains. Although Sam68 is a Src substrate, little is known about the signaling pathway that link them. Src is known to be activated transiently after cell spreading, where it modulates the activity of small Rho GTPases. Herein we report that Sam68-deficient cells exhibit loss of cell polarity and cell migration. Interestingly, Sam68-deficient cells exhibited sustained Src activity after cell attachment, resulting in the constitutive tyrosine phosphorylation and activation of p190RhoGAP and its association with p120rasGAP. Consistently, we observed that Sam68-deficient cells exhibited deregulated RhoA and Rac1 activity. By using total internal reflection fluorescence microscopy, we observed Sam68 near the plasma membrane after cell attachment coinciding with phosphorylation of its C-terminal tyrosines and association with Csk. These findings show that Sam68 localizes near the plasma membrane during cell attachment and serves as an adaptor protein to modulate Src activity for proper signaling to small Rho GTPases.

[Microtubules suppress blebbing and stimulate lamellae extension in spreading fibroblasts].

PubMed

Tvorogova, A V; Vorob'ev, I A

2012-01-01

We compared spreading of Vero fibroblasts when microtubules were depolymerized or stabilized. After initial attachment cells start blebbing that continues for different time and abruptly transfers into spreading. After spreading initiation, most cells spread in an anisotropic manner through stochastic formation of lamellipodia. A second mode was rapid, isotropic spreading via formation of circular lamellum that occurs in 15% of cells. The rate of spreading was maximal at the beginning and decreased during the first hour according to logarithmic law. After 60 min many cells formed stable efges and started migrating on the substrate. However, cell area slowly continued to increase. Actin bundles are formed 20 min after cell attachment and they first run along cell boundary. This system disassembles within 20-40 min and is substituted with stress fibers crossing the cell. In the isotropically spread cells no actin bunbles are seen. Microtubules in the spreading cells enter into large blebs and all nascent lamella and later form radial array. When MTs has been depolymerized or stabilized blebbing started before cells attached to the substrate and continue much longer than in control cells. In both cases the initial rate of spreading decrease several fold, and remains constant for many hours. After 24 h the mean area occupied by cells with altered MT system was the same as in control. Alteration of MT system had moderate effect on actin system--formation of actin cables started at the same time as in control (within 20 min upon cell attachment), however, they grew even in cells undergoing prolonged blebbing. Actin cables running along cell margin were similar to tat in control cells, but they did not disappear up to 1 h. When stabilized, microtubules form chaotic array: they do not enter blebs and in spread cells run parallel to the cell margin at a distance of 3-5 microm. We conclude that dynamic microtubules speed up completion of blebbing and promote early stages of fibroblasts spreading.

Connexin-43 upregulation in micrometastases and tumor vasculature and its role in tumor cell attachment to pulmonary endothelium

PubMed Central

Elzarrad, M Khair; Haroon, Abu; Willecke, Klaus; Dobrowolski, Radoslaw; Gillespie, Mark N; Al-Mehdi, Abu-Bakr

2008-01-01

Background The modulation of gap junctional communication between tumor cells and between tumor and vascular endothelial cells during tumorigenesis and metastasis is complex. The notion of a role for loss of gap junctional intercellular communication in tumorigenesis and metastasis has been controversial. While some of the stages of tumorigenesis and metastasis, such as uncontrolled cell division and cellular detachment, would necessitate the loss of intercellular junctions, other stages, such as intravasation, endothelial attachment, and vascularization, likely require increased cell-cell contact. We hypothesized that, in this multi-stage scheme, connexin-43 is centrally involved as a cell adhesion molecule mediating metastatic tumor attachment to the pulmonary endothelium. Methods Tumor cell attachment to pulmonary vasculature, tumor growth, and connexin-43 expression was studied in metastatic lung tumor sections obtained after tail-vein injection into nude mice of syngeneic breast cancer cell lines, overexpressing wild type connexin-43 or dominant-negatively mutated connexin-43 proteins. High-resolution immunofluorescence microscopy and Western blot analysis was performed using a connexin-43 monoclonal antibody. Calcein Orange Red AM dye transfer by fluorescence imaging was used to evaluate the gap junction function. Results Adhesion of breast cancer cells to the pulmonary endothelium increased with cancer cells overexpressing connexin-43 and markedly decreased with cells expressing dominant-negative connexin-43. Upregulation of connexin-43 was observed in tumor cell-endothelial cell contact areas in vitro and in vivo, and in areas of intratumor blood vessels and in micrometastatic foci. Conclusion Connexin-43 facilitates metastatic 'homing' by increasing adhesion of cancer cells to the lung endothelial cells. The marked upregulation of connexin-43 in tumor cell-endothelial cell contact areas, whether in preexisting 'homing' vessels or in newly formed tumor vessels, suggests that connexin-43 can serve as a potential marker of micrometastases and tumor vasculature and that it may play a role in the early incorporation of endothelial cells into small tumors as seeds for vasculogenesis. PMID:18647409

Connexin-43 upregulation in micrometastases and tumor vasculature and its role in tumor cell attachment to pulmonary endothelium.

PubMed

Elzarrad, M Khair; Haroon, Abu; Willecke, Klaus; Dobrowolski, Radoslaw; Gillespie, Mark N; Al-Mehdi, Abu-Bakr

2008-07-22

The modulation of gap junctional communication between tumor cells and between tumor and vascular endothelial cells during tumorigenesis and metastasis is complex. The notion of a role for loss of gap junctional intercellular communication in tumorigenesis and metastasis has been controversial. While some of the stages of tumorigenesis and metastasis, such as uncontrolled cell division and cellular detachment, would necessitate the loss of intercellular junctions, other stages, such as intravasation, endothelial attachment, and vascularization, likely require increased cell-cell contact. We hypothesized that, in this multi-stage scheme, connexin-43 is centrally involved as a cell adhesion molecule mediating metastatic tumor attachment to the pulmonary endothelium. Tumor cell attachment to pulmonary vasculature, tumor growth, and connexin-43 expression was studied in metastatic lung tumor sections obtained after tail-vein injection into nude mice of syngeneic breast cancer cell lines, overexpressing wild type connexin-43 or dominant-negatively mutated connexin-43 proteins. High-resolution immunofluorescence microscopy and Western blot analysis was performed using a connexin-43 monoclonal antibody. Calcein Orange Red AM dye transfer by fluorescence imaging was used to evaluate the gap junction function. Adhesion of breast cancer cells to the pulmonary endothelium increased with cancer cells overexpressing connexin-43 and markedly decreased with cells expressing dominant-negative connexin-43. Upregulation of connexin-43 was observed in tumor cell-endothelial cell contact areas in vitro and in vivo, and in areas of intratumor blood vessels and in micrometastatic foci. Connexin-43 facilitates metastatic 'homing' by increasing adhesion of cancer cells to the lung endothelial cells. The marked upregulation of connexin-43 in tumor cell-endothelial cell contact areas, whether in preexisting 'homing' vessels or in newly formed tumor vessels, suggests that connexin-43 can serve as a potential marker of micrometastases and tumor vasculature and that it may play a role in the early incorporation of endothelial cells into small tumors as seeds for vasculogenesis.

Megakaryocyte Polyploidization and Proplatelet Formation in Low-Attachment Conditions.

PubMed

Schlinker, Alaina C; Duncan, Mark T; DeLuca, Teresa A; Whitehead, David C; Miller, William M

2016-07-15

In vitro -derived platelets (PLTs), which could provide an alternative source of PLTs for patient transfusions, are formed from polyploid megakaryocytes (MKs) that extend long cytoplasmic projections, termed proplatelets (proPLTs). In this study, we compared polyploidization and proPLT formation (PPF) of MKs cultured on surfaces that either promote or inhibit protein adsorption and subsequent cell adhesion. A megakaryoblastic cell line exhibited increased polyploidization and arrested PPF on a low-attachment surface. Primary human MKs also showed low levels of PPF on the same surface, but no difference in ploidy. Importantly, both cell types exhibited accelerated PPF after transfer to a surface that supports attachment, suggesting that pre-culture on a non-adhesive surface may facilitate synchronization of PPF and PLT generation in culture.

An investigation of the psychological characteristics of stalkers: empathy, problem-solving, attachment and borderline personality features.

PubMed

Lewis, S F; Fremouw, W J; Del Ben, K; Farr, C

2001-01-01

This study examined the psychological characteristics of a sample of self-reported stalkers in comparison with a control group, on measures of empathy, problem-solving skills, attachment, and borderline personality features. Stalkers were identified by their endorsement of specific behavioral items, consistent with a widely adopted definition of stalking, denoting behaviors that: (a) are repeatedly directed toward an identified target; (b) are intrusive and unwanted; and (c) evoke fear in the victim. Stalkers scored significantly higher than controls on measures of insecure attachment and borderline personality features, suggesting that the stalking group demonstrates a general pattern of inadequate interpersonal attachment, has limited abilities to form and maintain appropriate relationships, is emotionally labile and unstable, and experiences ambivalence regarding their interpersonal relationships. Treatment implications are discussed herein.

Dewetting based fabrication of fibrous micro-scaffolds as potential injectable cell carriers.

PubMed

Song, Hokyung; Yin, Liya; Chilian, William M; Zhang Newby, Bi-Min

2015-03-01

Although regenerative medicine utilizing tissue scaffolds has made enormous strides in recent years, many constraints still hamper their effectiveness. A limitation of many scaffolds is that they form surface patches, which are not particularly effective for some types of "wounds" that are deep within tissues, e.g., stroke and myocardial infarction. In this study, we reported the generation of fibrous micro-scaffolds feasible for delivering cells by injection into the tissue parenchyma. The micro-scaffolds (widths<100μm) were made by dewetting of poly(lactic-co-glycolic acid) thin films containing parallel strips, and cells were seeded to form cell/polymer micro-constructs during or post the micro-scaffold fabrication process. Five types of cells including rat induced vascular progenitor cells were assessed for the formation of the micro-constructs. Critical factors in forming fibrous micro-scaffolds via dewetting of polymer thin films were found to be properties of polymers and supporting substrates, temperature, and proteins in the culture medium. Also, the ability of cells to attach to the micro-scaffolds was essential in forming cell/polymer micro-constructs. Both in vitro and in vivo assessments of injecting these micro-scaffolding constructs showed, as compared to free cells, enhanced cell retention at the injected site, which could lead to improved tissue engineering and regeneration. Copyright © 2014 Elsevier B.V. All rights reserved.

Differentiation of Mesenchymal Stem Cells from Human Umbilical Cord Tissue into Odontoblast-Like Cells Using the Conditioned Medium of Tooth Germ Cells In Vitro

PubMed Central

Li, Tian Xia; Yuan, Jie; Chen, Yan; Pan, Li Jie; Song, Chun; Bi, Liang Jia; Jiao, Xiao Hui

2013-01-01

The easily accessible mesenchymal stem cells in the Wharton's jelly of human umbilical cord tissue (hUCMSCs) have excellent proliferation and differentiation potential, but it remains unclear whether hUCMSCs can differentiate into odontoblasts. In this study, mesenchymal stem cells were isolated from the Wharton's jelly of human umbilical cord tissue using the simple method of tissue blocks culture attachment. UCMSC surface marker expression was then evaluated for the isolated cells using flow cytometry. The third-passage hUCMSCs induced by conditioned medium from developing tooth germ cells (TGC-CM) displayed high alkaline phosphatase (ALP) levels (P < 0.001), an enhanced ability to proliferate (P < 0.05), and the presence of mineralized nodules. These effects were not observed in cells treated with regular medium. After induction of hUCMSCs, the results of reverse transcriptional polymerase chain reaction (PCR) indicated that the dentin sialophosphoprotein (DSPP) and dentin matrix protein 1 (DMP1) genes were significantly tested. Additionally, dentin sialoprotein (DSP) and DMP1 demonstrated significant levels of staining in an immunofluorescence analysis. In contrast, the control cells failed to display the characteristics of odontoblasts. Taken together, these results suggest that hUCMSCs can be induced to differentiate into odontoblast-like cells with TGC-CM and provide a novel strategy for tooth regeneration research. PMID:23762828

Efficient gene transfer into nondividing cells by adeno-associated virus-based vectors.

PubMed Central

Podsakoff, G; Wong, K K; Chatterjee, S

1994-01-01

Gene transfer vectors based on adeno-associated virus (AAV) are emerging as highly promising for use in human gene therapy by virtue of their characteristics of wide host range, high transduction efficiencies, and lack of cytopathogenicity. To better define the biology of AAV-mediated gene transfer, we tested the ability of an AAV vector to efficiently introduce transgenes into nonproliferating cell populations. Cells were induced into a nonproliferative state by treatment with the DNA synthesis inhibitors fluorodeoxyuridine and aphidicolin or by contact inhibition induced by confluence and serum starvation. Cells in logarithmic growth or DNA synthesis arrest were transduced with vCWR:beta gal, an AAV-based vector encoding beta-galactosidase under Rous sarcoma virus long terminal repeat promoter control. Under each condition tested, vCWR:beta Gal expression in nondividing cells was at least equivalent to that in actively proliferating cells, suggesting that mechanisms for virus attachment, nuclear transport, virion uncoating, and perhaps some limited second-strand synthesis of AAV vectors were present in nondividing cells. Southern hybridization analysis of vector sequences from cells transduced while in DNA synthetic arrest and expanded after release of the block confirmed ultimate integration of the vector genome into cellular chromosomal DNA. These findings may provide the basis for the use of AAV-based vectors for gene transfer into quiescent cell populations such as totipotent hematopoietic stem cells. Images PMID:8057446

Efficient gene transfer into nondividing cells by adeno-associated virus-based vectors.

PubMed

Podsakoff, G; Wong, K K; Chatterjee, S

1994-09-01

Gene transfer vectors based on adeno-associated virus (AAV) are emerging as highly promising for use in human gene therapy by virtue of their characteristics of wide host range, high transduction efficiencies, and lack of cytopathogenicity. To better define the biology of AAV-mediated gene transfer, we tested the ability of an AAV vector to efficiently introduce transgenes into nonproliferating cell populations. Cells were induced into a nonproliferative state by treatment with the DNA synthesis inhibitors fluorodeoxyuridine and aphidicolin or by contact inhibition induced by confluence and serum starvation. Cells in logarithmic growth or DNA synthesis arrest were transduced with vCWR:beta gal, an AAV-based vector encoding beta-galactosidase under Rous sarcoma virus long terminal repeat promoter control. Under each condition tested, vCWR:beta Gal expression in nondividing cells was at least equivalent to that in actively proliferating cells, suggesting that mechanisms for virus attachment, nuclear transport, virion uncoating, and perhaps some limited second-strand synthesis of AAV vectors were present in nondividing cells. Southern hybridization analysis of vector sequences from cells transduced while in DNA synthetic arrest and expanded after release of the block confirmed ultimate integration of the vector genome into cellular chromosomal DNA. These findings may provide the basis for the use of AAV-based vectors for gene transfer into quiescent cell populations such as totipotent hematopoietic stem cells.

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

PubMed Central

2011-01-01

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

Establishment and Characterization of Immortalized Human Amniotic Epithelial Cells

PubMed Central

Zhou, Kaixuan; Koike, Chika; Yoshida, Toshiko; Okabe, Motonori; Fathy, Moustafa; Kyo, Satoru; Kiyono, Tohru; Saito, Shigeru

2013-01-01

Abstract Human amniotic epithelial cells (HAEs) have a low immunogenic profile and possess potent immunosuppressive properties. HAEs also have several characteristics similar to stem cells, and they are discarded after parturition. Thus, they could potentially be used in cell therapy with fewer ethical problems. HAEs have a short life, so our aim is to establish and characterize immortalized human amniotic epithelial cells (iHAEs). HAEs were introduced with viral oncogenes E6/E7 and with human telomerase reverse transcriptase (hTERT) to create iHAEs. These iHAEs have proliferated around 200 population doublings (PDs) for at least 12 months. High expression of stem cell markers (Oct 3/4, Nanog, Sox2, Klf4) and epithelial markers (CK5, CK18) were detected by immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). These iHAEs were expanded in ultra-low-attachment dishes to form spheroids similarly to epithelial stem/precursor cells. High expression of mesenchymal (CD44, CD73, CD90, CD105) and somatic (CD24, CD29, CD271, Nestin) stem cell markers was detected by flow cytometry. The iHAEs showed adipogenic, osteogenic, neuronal, and cardiac differentiation abilities. In conclusion, the immortalization of HAEs with the characteristics of stem cells has been established, allowing these iHAEs to become useful for cell therapy and regenerative medicine. PMID:23298399

Vibrio cholerae use pili and flagella synergistically to effect motility switching and conditional surface attachment

NASA Astrophysics Data System (ADS)

Utada, Andrew S.; Bennett, Rachel R.; Fong, Jiunn C. N.; Gibiansky, Maxsim L.; Yildiz, Fitnat H.; Golestanian, Ramin; Wong, Gerard C. L.

2014-09-01

We show that Vibrio cholerae, the causative agent of cholera, use their flagella and mannose-sensitive hemagglutinin (MSHA) type IV pili synergistically to switch between two complementary motility states that together facilitate surface selection and attachment. Flagellar rotation counter-rotates the cell body, causing MSHA pili to have periodic mechanical contact with the surface for surface-skimming cells. Using tracking algorithms at 5 ms resolution we observe two motility behaviours: ‘roaming', characterized by meandering trajectories, and ‘orbiting’, characterized by repetitive high-curvature orbits. We develop a hydrodynamic model showing that these phenotypes result from a nonlinear relationship between trajectory shape and frictional forces between pili and the surface: strong pili-surface interactions generate orbiting motion, increasing the local bacterial loiter time. Time-lapse imaging reveals how only orbiting mode cells can attach irreversibly and form microcolonies. These observations suggest that MSHA pili are crucial for surface selection, irreversible attachment, and ultimately microcolony formation.

Synthesis and antioxidant properties of caffeic acid corn bran arabinoxylan esters.

PubMed

Li, Y; Zhu, Y; Liang, R; Yang, C

2017-08-01

As a potential of bioactive additives, corn bran arabinoxylan (CAX) was prominent in its probiotic benefits and immuno-enhancing activities. To improve the antioxidant ability of CAX, naturally occurring caffeic acid (CA) was covalently attached to CAX by esterification to generate caffeic acid corn bran arabinoxylan esters (CA-CAX) with various degrees of substitution (DS). The structure of CA-CAX was analysed by NMR, and the DS was determined by HPLC. The antioxidant activity of CA-CAX was investigated on scavenging the 2,2'-diphenyl-1-picrylhydrazyl (DPPH) radical and autoxidazing methyl linoleate, and an aqueous linolenic acid dispersion was also used as an in vitro test system to examine the inhibition effect of CA-CAX on the lipid peroxidation level after UV exposure. The ability of CA-CAX to prevent H 2 O 2 -induced oxidative damage of cell was studied by reducing MDA levels and increasing SOD and GSH-Px activities in human hepatocarcinoma (HepG2) cell. The results certificated that CA was linked to CAX successfully with various DS. Compared with CAX, the antioxidant capacity of CA-CAX was improved significantly and enhanced with the increasing concentration and DS. © 2017 Society of Cosmetic Scientists and the Société Française de Cosmétologie.

Developing a Continuous Bioprocessing Approach to Stromal Cell Manufacture.

PubMed

Miotto, Martina; Gouveia, Ricardo; Abidin, Fadhilah Zainal; Figueiredo, Francisco; Connon, Che J

2017-11-29

To this day, the concept of continuous bioprocessing has been applied mostly to the manufacture of molecular biologics such as proteins, growth factors, and secondary metabolites with biopharmaceutical uses. The present work now sets to explore the potential application of continuous bioprocess methods to source large numbers of human adherent cells with potential therapeutic value. To this purpose, we developed a smart multifunctional surface coating capable of controlling the attachment, proliferation, and subsequent self-detachment of human corneal stromal cells. This system allowed the maintenance of cell cultures under steady-state growth conditions, where self-detaching cells were continuously replenished by the proliferation of those remaining attached. This facilitated a closed, continuous bioprocessing platform with recovery of approximately 1% of the total adherent cells per hour, a yield rate that was maintained for 1 month. Moreover, both attached and self-detached cells were shown to retain their original phenotype. Together, these results represent the proof-of-concept for a new high-throughput, high-standard, and low-cost biomanufacturing strategy with multiple potentials and important downstream applications.

Carbon nanotube-coating accelerated cell adhesion and proliferation on poly (L-lactide)

NASA Astrophysics Data System (ADS)

Hirata, Eri; Akasaka, Tsukasa; Uo, Motohiro; Takita, Hiroko; Watari, Fumio; Yokoyama, Atsuro

2012-12-01

The surface of a polylactic acid (PLLA) was coated multiwalled carbon nanotubes (MWCNTs) in order to improve the surface properties. In addition, its surface characteristics and cell culturing properties were examined. Whole surface of PLLA was homogeneously covered by MWCNTs maintained a unique tubular structure. MWCNT-coated PLLA showed remarkable higher wettability than uncoated PLLA. Human osteosarcoma cell line (Saos2) adhered well on the CNT-coated PLLA whereas there are few cells attached on the uncoated PLLA at 2 h after seeding. The number of the cells on uncoated PLLA was still smaller than on the MWCNT-coated PLLA at 1 and 3 days. Moreover, The DNA content in the cells attached to the MWCNT-coated PLLA was significantly higher than that on the uncoated PLLA (p < 0.05) at 1 and 3 days. There was no significant difference between the scaffolds for ALP activity normalized by DNA content at both term (p > 0.1). Therefore MWCNT-coating on PLLA improved the surface wettability and initial cell attachment at early stage.

Filamentation and spatiotemporal distribution of extracellular polymeric substances: role on X.fastidiosa single cell adhesion and biofilm formation (Conference Presentation)

NASA Astrophysics Data System (ADS)

Janissen, Richard; Murillo, Duber M.; Niza, Barbara; Sahoo, Prasana K.; Monteiro, Moniellen P.; César, Carlos L.; Carvalho, Hernandes F.; de Souza, Alessandra A.; Cotta, Monica A.

2016-04-01

Biofilms can be defined as a community of microorganisms attached to a surface, living embedded in a self- produced matrix of hydrated extracellular polymeric substances (EPS) which comprises most of the biofilm mass. We have recently used an extensive pool of microscopy techniques (confocal fluorescence, electron and scanning probe microscopies) at the micro and nanoscales in order to create a detailed temporal observation of Xylella fastidiosa biofilm formation, using both wild type strain and Green Fluorescent Protein (GFP)-modified cells of this citrus phytopathogen. We have identified three different EPS compositions, as well as their spatial and temporal distribution from single cell to mature biofilm formation stages. In the initial adhesion stage, soluble-EPS (S-EPS) accumulates at cell polar regions and forms a surface layer which facilitates irreversible cell attachment and cell cluster formation. These small clusters are subsequently connected by filamentous cells; further S-EPS surface coverage facilitates cell attachment and form filaments, leading to a floating framework of mature biofilms. The important role of EPS in X.fastidiosa biology was further investigated by imunolabelling experiments to detect the distribution of XadA1 adhesin, which is expressed in early stages of biofilm formation and released in outer membrane vesicles. This protein is located mainly in S-EPS covered areas, as well as on the filaments, indicating a molecular pathway to the enhanced cell attachment previously observed. These results suggest that S-EPS may thus represent an important target for disease control, slow plant colonization by the bacteria, keeping the plant more productive in the field.

Type XVII collagen (BP180) can function as a cell-matrix adhesion molecule via binding to laminin 332

PubMed Central

Van den Bergh, F.; Eliason, S.L.; Giudice, G.J.

2010-01-01

Collagen XVII (COL17) is a transmembrane glycoprotein that is expressed on the basal surface of basal epidermal keratinocytes. Previous observations have led to the hypothesis that an interaction between COL17 and laminin 332, an extracellular matrix protein, contributes to the attachment of the basal keratinocyte to the basement membrane. In order to isolate and manipulate COL17 interactions with ECM components, we induced COL17 expression in two cells lines, SK-MEL1 and K562, that exhibit little or no capacity to attach to our test substrates, including laminin 332, types I and IV collagen, and fibronectin. Cells expressing high levels of COL17 preferentially adhered to a laminin 332 matrix, and, to a lesser extent, type IV collagen, while showing little or no binding to type I collagen or fibronectin. A quantitative analysis of cell adhesive forces revealed that, compared with COL17-negative cells, COL17-positive cells required over 7-fold greater force to achieve 50% detachment from a laminin 332 substrate. When a cell preparation (either K562 or SK-MEL1) with heterogeneous COL17 expression levels was allowed to attach to a laminin 332 matrix, the COL17-positive and COL17-negative cells differentially sorted to the bound and unbound cell fractions, respectively. COL17-dependent attachment to laminin 332 could be reduced or abolished by siRNA-mediated knockdown of COL17 expression or by adding to the assay wells specific antibodies against COL17 or laminin 332. These findings provide strong support for the hypothesis that cell surface COL17 can interact with laminin 332 and, together, participate in the adherence of a cell to the extracellular matrix. PMID:21034821

Natural zwitterionic l-Carnitine as efficient cryoprotectant for solvent-free cell cryopreservation.

PubMed

Zhai, Hongwen; Yang, Jing; Zhang, Jiamin; Pan, Chao; Cai, Nana; Zhu, Yingnan; Zhang, Lei

2017-07-15

Organic solvents, such as dimethyl sulfoxide (DMSO) and glycerol, have been commonly used as cryoprotectants (CPAs) in cell cryopreservation. However, their cytotoxicity and need of complex freezing protocols have impeded their applications especially in clinical cell therapy and regenerative medicine. Trehalose has been explored as a natural CPA to cryopreserve cells, but its poor cell permeability frequently results in low cryopreservation efficacy. In this work, we presented that a natural zwitterionic molecule-l-carnitine-could serve as a promising CPA for solvent-free cryopreservation. We demonstrated that l-carnitine possessed strong ability to depress water freezing point, and with ultrarapid freezing protocol, we studied the post-thaw survival efficiency of four cell lines (GLC-82 cells, MCF-7 cells, NIH-3T3 cells and Sheep Red Blood Cells) using l-carnitine without addition of any organic solvents. At the optimum l-carnitine concentration, all four cell lines could achieve above 80% survival efficiency, compared with the significantly lower efficiency using organic CPAs and trehalose. After cryopreservation, the recovered cell behaviors including cell attachment and proliferation were found to be similar to the normal cells, indicating that the cell functionalities were not affected. Moreover, l-carnitine showed no observable cytotoxicity, which was superior to the organic CPAs. This work offered an attractive alternative to traditional CPAs and held great promise to revolutionize current cryopreservation technologies, to benefit the patients in various cell-based clinical applications. Copyright © 2017 Elsevier Inc. All rights reserved.

Mechanisms of Bacterial (Serratia marcescens) Attachment to, Migration along, and Killing of Fungal Hyphae.

PubMed

Hover, Tal; Maya, Tal; Ron, Sapir; Sandovsky, Hani; Shadkchan, Yana; Kijner, Nitzan; Mitiagin, Yulia; Fichtman, Boris; Harel, Amnon; Shanks, Robert M Q; Bruna, Roberto E; García-Véscovi, Eleonora; Osherov, Nir

2016-05-01

We have found a remarkable capacity for the ubiquitous Gram-negative rod bacterium Serratia marcescens to migrate along and kill the mycelia of zygomycete molds. This migration was restricted to zygomycete molds and several basidiomycete species. No migration was seen on any molds of the phylum Ascomycota. S. marcescens migration did not require fungal viability or surrounding growth medium, as bacteria migrated along aerial hyphae as well.S. marcescens did not exhibit growth tropism toward zygomycete mycelium. Bacterial migration along hyphae proceeded only when the hyphae grew into the bacterial colony. S. marcescens cells initially migrated along the hyphae, forming attached microcolonies that grew and coalesced to generate a biofilm that covered and killed the mycelium. Flagellum-defective strains of S. marcescens were able to migrate along zygomycete hyphae, although they were significantly slower than the wild-type strain and were delayed in fungal killing. Bacterial attachment to the mycelium does not necessitate type 1 fimbrial adhesion, since mutants defective in this adhesin migrated equally well as or faster than the wild-type strain. Killing does not depend on the secretion of S. marcescens chitinases, as mutants in which all three chitinase genes were deleted retained wild-type killing abilities. A better understanding of the mechanisms by which S. marcescens binds to, spreads on, and kills fungal hyphae might serve as an excellent model system for such interactions in general; fungal killing could be employed in agricultural fungal biocontrol. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Mechanisms of Bacterial (Serratia marcescens) Attachment to, Migration along, and Killing of Fungal Hyphae

PubMed Central

Hover, Tal; Maya, Tal; Ron, Sapir; Sandovsky, Hani; Shadkchan, Yana; Kijner, Nitzan; Mitiagin, Yulia; Fichtman, Boris; Harel, Amnon; Shanks, Robert M. Q.; Bruna, Roberto E.; García-Véscovi, Eleonora

2016-01-01

We have found a remarkable capacity for the ubiquitous Gram-negative rod bacterium Serratia marcescens to migrate along and kill the mycelia of zygomycete molds. This migration was restricted to zygomycete molds and several basidiomycete species. No migration was seen on any molds of the phylum Ascomycota. S. marcescens migration did not require fungal viability or surrounding growth medium, as bacteria migrated along aerial hyphae as well. S. marcescens did not exhibit growth tropism toward zygomycete mycelium. Bacterial migration along hyphae proceeded only when the hyphae grew into the bacterial colony. S. marcescens cells initially migrated along the hyphae, forming attached microcolonies that grew and coalesced to generate a biofilm that covered and killed the mycelium. Flagellum-defective strains of S. marcescens were able to migrate along zygomycete hyphae, although they were significantly slower than the wild-type strain and were delayed in fungal killing. Bacterial attachment to the mycelium does not necessitate type 1 fimbrial adhesion, since mutants defective in this adhesin migrated equally well as or faster than the wild-type strain. Killing does not depend on the secretion of S. marcescens chitinases, as mutants in which all three chitinase genes were deleted retained wild-type killing abilities. A better understanding of the mechanisms by which S. marcescens binds to, spreads on, and kills fungal hyphae might serve as an excellent model system for such interactions in general; fungal killing could be employed in agricultural fungal biocontrol. PMID:26896140

Potential for Aedes albopictus and Ochlerotatus j. japonicus to Change the Field Ecology of Arboviruses of Human Health Importance in the Mid-Atlantic Region of the United States

DTIC Science & Technology

2001-10-16

in the midgut epithelial cells. Step 3 is release 17 of virus from the midgut epithelial cell into the hemolymph. Step 4 is infection of the...proteins, maturation, and budding. During attachment, the viral attachment proteins on the surface of the virion bind to receptors on the microvillar...membrane of the mosquito s midgut . Penetration refers to the process through which the virions enter the midgut cells; arboviruses enter cell

The Macrophage Galactose-Type Lectin Can Function as an Attachment and Entry Receptor for Influenza Virus

PubMed Central

Ng, Wy Ching; Liong, Stella; Tate, Michelle D.; Irimura, Tatsuro; Denda-Nagai, Kaori; Brooks, Andrew G.; Londrigan, Sarah L.

2014-01-01

Specific protein receptors that mediate internalization and entry of influenza A virus (IAV) have not been identified for any cell type. Sialic acid (SIA), the primary attachment factor for IAV hemagglutinin, is expressed by numerous cell surface glycoproteins and glycolipids, confounding efforts to identify specific receptors involved in virus infection. Lec1 Chinese hamster ovary (CHO) epithelial cells express cell surface SIA and bind IAV yet are largely resistant to infection. Here, we demonstrate that expression of the murine macrophage galactose-type lectin 1 (MGL1) by Lec1 cells enhanced Ca2+-dependent IAV binding and restored permissivity to infection. Lec1 cells expressing MGL1 were infected in the presence or absence of cell surface SIA, indicating that MGL1 can act as a primary receptor or as a coreceptor with SIA. Lec1 cells expressing endocytosis-deficient MGL1 mediated Ca2+-dependent IAV binding but were less sensitive to IAV infection, indicating that direct internalization via MGL1 can result in cellular infection. Together, these studies identify MGL1 as a cell surface glycoprotein that can act as an authentic receptor for both attachment and infectious entry of IAV. PMID:24257596

Physical and metabolic requirements for early interaction of poliovirus and human rhinovirus with HeLa cells.

PubMed Central

Lonberg-Holm, K; Whiteley, N M

1976-01-01

Attachment, ""tight binding'' and eclipse of radioactive poliovirus 2 (P2) and human rhinovirus 2 (HRV 2) were investigated. The activation energy for attachment of both HRV2 and P2 was about 13 kcal/mol. HRV2 differed from P2 in two respects: the Arrhenius plot for attachment of HRV2 showed a break at 15 to 19 degrees C when the cells were first treated several hours at 0 degrees C, and attachment of HRV2 was inhibited by treatment of cells with metabolic poisons able to reduce cellular ATP by more than 90%. Tight binding was determined by isolation of a specific P2-membrane complex or by loss of EDTA dissociability of HRV2. Tight binding of both viruses was slowed by 0.01 M iodoacetamide but not by 0.02 M F-; F- plus 0.002 M CN- slowed tight binding of HRV2 but not of P2. Eclipse, the irreversible alteration of parental virions, was detected by isolation of cell-associated subviral particles or by loss of cell-associated infectious virus. Eclipse of both viruses is slowed by iodoacetamide or F-. It seems likely that the early steps of infection with picornaviruses may be sensitive to alterations in the cell membrane produced by metabolic inhibitors or by treatment at low temperature. PMID:184301

Correlation of open cell-attached and excised patch clamp techniques.

PubMed

Filipovic, D; Hayslett, J P

1995-11-01

The excised patch clamp configuration provides a unique technique for some types of single channel analyses, but maintenance of stable, long-lasting preparations may be confounded by rundown and/or rapid loss of seal. Studies were performed on the amiloride-sensitive Na+ channel, located on the apical surface of A6 cells, to determine whether the nystatin-induced open cell-attached patch could serve as an alternative configuration. Compared to excised inside-out patches, stable preparations were achieved more readily with the open cell-attached patch (9% vs. 56% of attempts). In both preparations, the current voltage (I-V) relation was linear, current amplitudes were equal at opposite equivalent clamped voltages, and Erev was zero in symmetrical Na+ solutions, indicating similar Na+ activities on the cytosolic and external surfaces of the patch. Moreover, there was no evidence that nystatin altered channel activity in the patch because slope conductance (3-4pS) and Erev (75 mV), when the bath was perfused with a high K:low Na solution (ENa = 80 mV), were nearly equal in both patch configurations. Our results therefore indicate that the nystatin-induced open cell-attached patch can serve as an alternative approach to the excised inside-out patch when experiments require modulation of univalent ions in the cytosol.