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PubMed2000

Prevalence of HEp-2 cell-adherent Escherichia coli and characterisation of enteroaggregative E. coli and chain-like adherent E. coli isolated from children with and without diarrhoea, in Londrina, Brazil.

A total of 919 Escherichia coli isolates from 125 children with diarrhoea (cases) and 98 controls were assayed for adherence to HEp-2 cells. Localised adherence was found only in isolates from cases. Diffuse, aggregative (AA), chain-like adherence (CLA) and variants of the AA pattern were found in both cases and controls. The AA isolates were tested for gene sequences associated with enteroaggregative E. coli (EAEC). Only 25% of the isolates hybridised with the EAEC probe, and the aafA, astA and pet gene sequences were found in 7.9%, 44.7% and 7.9% of the isolates, respectively. The aggA gene was not found, although 7.9% were positive for aggC. The CLA isolates reacted with the EAEC probe (55.6%), and the aggC, astA and pet gene sequences were found in 66.7%, 33.3% and 11.1%, respectively. The aggR (55.6%), aspU (55.6%), shf (33.3%) and she (22.2%) genes were also found in CLA isolates.

Adhesins, Escherichia coli↗
PubMed1990

Oligonucleotide probe for detection of the enteropathogenic Escherichia coli (EPEC) adherence factor of localized adherent EPEC.

The enteropathogenic Escherichia coli adherence factor (EAF) probe detects isolates of enteropathogenic E. coli that exhibit localized adherence to HEp-2 cells. A 21-base oligonucleotide probe was constructed on the basis of a sequence from within the 1-kb EAF probe and was shown to have greater sensitivity and specificity than the EAF fragment probe in detecting localized adherent E. coli.

Adhesins, Escherichia coli↗
PubMed2001

Flow around cells adhered to a microvessel wall II: comparison to flow around adherent cells in channel flow.

The flow around adherent cells in a parallel-plate channel and that in a circular cylindrical tube are numerically analyzed, and their effects on the adherent cells are compared. The cells are modeled as rigid spherical particles and they are assumed to be attached to a wall of a 2D channel uniformly in a square array, or a wall of a circular tube regularly in a line along the tube axis. It is found that, when the size ratios of the particle-to-channel height and the particle-to-tube diameter are smaller than approximately 0.2, the distributions of the shear stress and the pressure exerted on the surface of an adherent particle as well as the drag force and torque acting on it compare favorably in the 2D channel flow and tube flow. As the size ratios increase from 0.2, the differences between the 2D channel and the tube increase drastically, especially when separation distances between neighboring particles are large.

Cell Adhesion↗
PubMed1998

IL-2 induces T cell adherence to extracellular matrix: inhibition of adherence and migration by IL-2 peptides generated by leukocyte elastase.

Migration of inflammatory cells requires cell adhesion and their subsequent detachment from the extracellular matrix (ECM). Leukocyte activation and migration must be terminated to stop inflammation. Here, we report that IL-2 enhances human T cell adherence to laminin, collagen type IV, and fibronectin (FN). In contrast, neutrophil elastase, an enzyme activated during inflammation, degrades IL-2 to yield IL-2 fractions that inhibit IL-2-induced T cell adhesion to FN. The amino acid composition of two of these IL-2 fractions, which appear to block T cell adherence to FN, were analyzed, and three peptides were consequently synthesized. The three peptides IVL, RMLT, and EFLNRWIT, but not the corresponding inversely synthesized peptides, inhibited T cell adhesion to FN induced by a variety of activators: IL-2, IL-7, macrophage inflammatory protein (MIP)-1beta, and PMA, as well as anti-CD3 and anti-beta1 integrin-activating mAb. Moreover, these IL-2 peptides inhibited T cell chemotaxis via FN-coated membranes induced by IL-2 and MIP-1beta. Inhibition of T cell adherence and migration apparently involves abrogation of the rearrangement of the T cell actin cytoskeleton. Thus, the migrating immune cells, the cytokines, and the ECM can create a functional relationship in which both inflammation-inducing signals and inhibitory molecules of immune responses can coexist; the enzymatic products of IL-2 may serve as natural feedback inhibitors of inflammation.

Actins↗
PubMed1984

Amplification of lymph node cell tube leukocyte adherence inhibition (LAI) reactivity by leukocyte adherence inhibition factor (LAIF).

This investigation examines the immunologic basis for specific antigen-induced tube leukocyte adherence inhibition (LAI) reactivity of draining lymph node cells (LNC) from dogs with canine transmissible venereal sarcoma (CTVS). CTVS regressor LNC, macrophage-depleted LNC, and enriched T lymphocyte fractions, but not enriched B lymphocyte fractions, were specifically reactive to CTVS antigen extract in direct tube LAI. In addition, regressor LNC amplified tube LAI responses by generating supernatants with leukocyte adherence inhibition factor (LAIF) activity for normal dog indicator LNC and enriched peripheral blood mononuclear cells (PBMC) in an indirect tube LAI assay. However, macrophage-depleted LNC and enriched T lymphocyte fractions failed to generate supernatants with LAIF activity, suggesting that macrophage accessory cells play a central role in the amplification of tube LAI. Interestingly, CTVS regressor peripheral blood leukocytes (PBL) and PBMC, which were specifically reactive in direct tube LAI, also failed to generate supernatants with LAIF activity. These findings demonstrate a distinction between LAIF-mediated amplification and direct tube LAI reactivity, and suggest that leukocyte populations with differing cellular proportions and from different immunologic compartments may participate in tube LAI via different mechanisms.

Animals↗
PubMed1980

Specific immunodiagnosis of hepatoma by tube leukocyte adherence inhibition assay and a modified method of repeated tube leukocyte adherence inhibition assay.

Tube leukocyte adherence inhibition (LAI) assays were performed with normal liver extract as nonspecific antigen and with hepatoma extract as specific antigen in patients with hepatoma. Titration experiments revealed that the optimal extract concentration was 400 micrograms/ml when expressing the results in terms of a nonadherence index. The results of tube LAI assays were positive in 26 of 40 cases (65%) of hepatoma. The results were negative in all cases of other liver diseases and other cancers. The tube LAI assay was repeated after discarding the non-adherent cells in the initial tube LAI assay with normal liver extract. The nonadherence index of the repeated tube LAI assay we devised was significantly higher than that of the original tube LAI assay (p less than 0.001) in patients with hepatoma. Ten of 12 patients with hepatoma in whom the results of the original tube LAI assay were negative showed positive results in the repeated tube LAI assay. The present study suggests that the problem of false-negative results in tube LAI assay can be solved by repeating the tube LAI assay.

Antigens, Neoplasm↗
PubMed1981

Macrophage adherence inhibition assay: measurements of adherent cells by the TV image analysis system Densitron.

A procedure is described to automate the cell counting step in the indirect macrophage adherence inhibition assay using the TV image analysis system Densitron. Comparison of the results of direct visual counting and of Densitron measurements of the adherent cells in the haemocytometer chamber indicates the qualification on principle of Densitron to substitute the time consuming and exerting visual counting procedure.

Animals↗
PubMed1990

Differences in c-myc and pvt-1 amplification in SEWA sarcoma sublines selected for adherent or non-adherent growth.

Conversion of solid sarcomas and carcinomas into ascites tumors depends on the in vivo selection of phenotypically altered tumor cell variants that can grow in the dissociated form. Once selected, they retain this property even after prolonged s.c. growth as solid tumors. From an s.c.-passaged subline of an ascites-converted murine sarcoma (SEWA-AS12), we were able to separate cells adapted to the ascites form of growth from cells that can only grow in the solid form on the basis of their differential adherence to plastic. Both c-myc and pvt-1 were amplified approximately 63- to 77-fold in the nonadherent subline (SEWA-AS12-NA), but only 5- to 8-fold in the adherent subline (SEWA-AS12-ADH). This suggests that c-myc and/or pvt-1 amplification may provide a selective advantage to cells that can grow in the dissociated form.

Animals↗
PubMed1996

Determination of LDL- and scavenger-receptor activity in adherent and non-adherent cultured cells with a new single-step fluorometric assay.

Lipoproteins labeled with 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) are widely used to visualize LDL-and scavenger-receptor activity in cultured cells. The purpose of this study was to evaluate a new single-step fluorometric assay with high sensitivity for the quantitative determination of the LDL- or scavenger-receptor activity in adherent and non-adherent cells. We used an aqueous solution of 1 g/l SDS dissolved in 0.1 M NaOH to lyse the cells after incubation with DiI-LDL or DiI-acetylated LDL. This allows for the first time the determination of fluorescence intensity and cell protein in the same sample without prior lipid extraction of the fluorochrome. Fluorescence of the cell lysates was determined in microtiter plates with excitation-emission set at 520 and 580 nm, respectively. This rapid method demonstrates high specificity for determining the LDL- and scavenger-receptor activity in cultured cells (e.g., human skin fibroblasts from patients with and without familial hypercholesterolemia; human U-937 monocyte and murine P388 D1 macrophage cell lines). The validity of our fluorescence assay is demonstrated by comparison of cellular uptake and metabolism of lipoproteins labeled with both, DiI and 125iodine. The rapidity and accuracy of this assay allows its routine application for studying receptor-mediated lipoprotein uptake in various cell types.

Animals↗
PubMed1990

Necessity for interaction between adherent and non-adherent rat spleen cells for the generation of a suppressor factor of NK activation.

Whole rat spleen cell populations in culture release constitutively a factor which suppresses the NK activation in response to interleukin-2. Culture supernatants derived from adherent (AD) and non-adherent (NAD) cell preparations obtained from rat spleen cells, did not have a suppressor activity comparable to that present in the culture supernatants of whole spleen cells. When reconstituted mixtures of the AD and NAD cells were cultured, high levels of suppressor activity could again be demonstrated in the culture supernatants, indicating that some kind of co-operation between AD and NAD cells was needed for efficient generation of the suppressor. Non-suppressive culture supernatants derived from NAD cells became suppressive when AD cells were cultured in them. On the other hand, relatively little suppressor activity was generated when NAD cells were cultured in non-suppressive culture supernatants from AD cells. Our results seem to indicate that NAD cells may release some agent which is not itself a suppressor but either (a) induces the generation of suppressor activity from AD cells, or (b) is "processed" into a suppressor agent by AD cells.

Animals↗
PubMed1999

Comparison of cytoskeletal structures in glass-adherent and polyethylene terephthalate (PET) membrane-adherent human pulmonary fibroblasts.

In order to use microporous membranes as a culture substratum for later air/liquid exposures with human pulmonary fibroblasts, we were interested to find out whether the behaviour of cells grown on PET membranes differs from those cultured under conventional conditions. We therefore compared cytoskeletal structures of membrane-adherent and glass-adherent cells. There were no differences found between the cytoskeleton of cells grown on membranes or on glass. They attach equally to both of the substrates and develop the same cytoskeletal structures. We also investigated the proliferative activity of the fibroblasts on PET-cell culture inserts by determination of Pico Green intercalation into DNA and turnover of a tetrazolium salt (WST) by mitochondrial activity. In summary, the fibroblasts were found to show characteristic growth tendencies when cultivated on PET membranes.

Cell Adhesion↗
PubMed2002

Phenotype modulation in non-adherent and adherent sublines of Walker carcinosarcoma cells: the role of cell-substratum contacts and microtubules in controlling cell shape, locomotion and cytoskeletal structure.

We characterised two sublines of Walker carcinosarcoma cells generated by epigenetic changes. Subline 1 cells were mostly polarised and made no or only non-adhesive cell-substratum contacts. Subline 2 cells were spread, adhesive and mainly non-polar. Subline 1 cells migrate in a non-adhesive mode which is very efficient but operates only in a 3D environment, whereas subline 2 cells migrate in an adhesive mode, which is less efficient but works on 2D and 3D substrata. Nocodazole had little or no effect on shape, polarity and locomotion of subline 1 cells. In glass-adherent subline 2 cells, 10(-6)M nocodazole increased the proportion of polarised cells migrating in an adhesive mode and decreased adhesion to the substratum, whereas 10(-5)M nocodazole further reduced the contacts and the cells reverted to a non-adhesive mode of locomotion. When non-polar subline 2 cells were detached mechanically or by nocodazole, they became polarised and morphologically indistinguishable from non-adherent subline 1 cells. On more adhesive plastic substrata, subline 2 cells produced heterogeneous responses to nocodazole including loss of polarity. The phenotypes of Walker carcinosarcoma sublines have similarities with a broad range of cell types ranging from leucocytes to fibroblast-like cells, suggesting that these phenotypic differences can be controlled by the adhesive and contractile state rather than the cell type. Adhesion modulates contractility (isometric or isotonic contraction) and vice versa and this determines morphology (shape, F-actin, myosin and alpha-actinin), locomotion and responses to microtubule-disassembly. The model may be applied to analyse the mechanisms controlling the phenotype of cells in general.

Actinin↗
PubMed1996

Effect of adherent Lactobacillus spp. on in vitro adherence of salmonellae to the intestinal epithelial cells of chicken.

Single strains of Lactobacillus acidophilus and Lact. fermentum, isolated from chicken intestine, were used to study in vitro interactions with Salmonella enteritidis, Salm. pullorum or Salm. typhimurium in an ileal epithelial cell (IEC) radioactive assay. Exclusion, competition and displacement phenomena were investigated by respectively incubating (a) lactobacilli and IEC together, prior to addition of salmonellae, (b) lactobacilli, IEC and salmonellae together, and (c) salmonellae and IEC, followed by the lactobacilli. Lactobacilli were selected for study because of their strong ability to adhere to IEC and poor aggregation with salmonellae. The results demonstrated that Lact. acidophilus significantly reduced (P < 0.05) the attachment of Salm. pullorum to IEC in the tests for exclusion and competition, but not in the displacement tests. Lactobacillus fermentum was found to have some ability to reduce the attachment of Salm. typhimurium to IEC under the conditions of exclusion (P < 0.08), competition (P < 0.09), but not displacement. However, both Lact. acidophilus and Lact. fermentum were unable to reduce the adherence of Salm. enteritidis to IEC under any of the conditions.

Animals↗
PubMed1987

Adherence of cells to myelin basic protein. I. Adherence of red and white blood cells from patients with multiple sclerosis to myelin basic protein.

Red blood cells (RBC) and white blood cells (WBC) of patients with multiple sclerosis (MS) show decreased adherence to myelin basic protein (MBP) immobilized on plastic surfaces compared to the binding of cells from patients with other neurological diseases (OND), or such other autoimmune diseases as psoriasis (PS), and to that of healthy controls (HC). No similar phenomenon occurred to basic and non-basic type proteins other than MBP, for example, to histone (HIS), lysozyme (LYS) and ovalbumin (OVA). Thus, decreased adherence of RBC and WBC in MS patients to MBP appears to be a unique feature of the disease if compared with OND or PS.

Cell Adhesion↗
PubMed1992

Aggregative adherence fimbriae I of enteroaggregative Escherichia coli mediate adherence to HEp-2 cells and hemagglutination of human erythrocytes.

Strains of enteroaggregative Escherichia coli (EAggEC) have been implicated in several studies as important agents of persistent diarrhea among infants in the developing world. We have previously shown that the aggregative adherence (AA) property of EAggEC is associated with the presence of a 60-MDa plasmid which confers AA when introduced into E. coli HB101. Here, we report the cloning of the AA determinant from EAggEC strain 17-2 into the 21.5-kb cosmid vector pCVD301. TnphoA mutagenesis of the AA cosmid clone pJPN31 implicated an AA region of approximately 12 kb. Transmission electron microscopy of HB101 (pJPN31) revealed the presence of bundle-forming fimbriae, which were absent in AA- TnphoA insertion mutants. The presence of these fimbriae, AA, and hemagglutination (HA) of human erythrocytes were all concurrently lost by single-insertion mutations. A 14-kDa protein was seen on polyacrylamide gel electrophoresis and Western blotting (immunoblotting) of surface shear preparations from fimbriated clones. Twelve of nineteen volunteers fed EAggEC 17-2 developed rises in antibodies to the 14-kDa protein as determined by Western blot. We have termed the cloned bundle-forming fimbriae aggregative adherence fimbriae I (AAF/I); positivity with a previously described EAggEC probe and human erythrocyte HA appear to correlate with the presence of AAF/I.

Antibodies, Bacterial↗
PubMed1996

Phospholipase C and perfringolysin O from Clostridium perfringens upregulate endothelial cell-leukocyte adherence molecule 1 and intercellular leukocyte adherence molecule 1 expression and induce interleukin-8 synthesis in cultured human umbilical vein endothelial cells.

Clostridium perfringens phospholipase C (PLC) and perfringolysin O (PFO) differentially induced human umbilical vein endothelial cell expression and synthesis of endothelial cell-leukocyte adherence molecule-1 (ELAM-1), intracellular leukocyte adherence molecule-1 (ICAM-1), and interleukin-8 (IL-8). PLC strongly induced expression of ELAM-1, ICAM-1, and IL-8, while PFO stimulated early ICAM-1 expression but did not promote ELAM-1 expression or IL-8 synthesis. PLC caused human umbilical vein endothelial cells to assume a fibroblastoid morphology, whereas PFO, in high concentrations or after prolonged low-dose toxin exposure, caused cell death. The toxin-induced expression of proadhesive and activational proteins and direct cytopathic effects may contribute to the leukostasis, vascular compromise, and capillary leak characteristics of C. perfringens gas gangrene.

Bacterial Toxins↗
PubMed1996

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

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

Actinomyces↗