Tumor necrosis factor alpha interferes with the cell cycle of normal and papillomavirus-immortalized human keratinocytes

We have shown that normal and human papillomavirus (HPV) type 16 immortalized human foreskin keratinocytes are growth inhibited by tumor necrosis factor alpha (TNF-alpha), whereas HPV-18- and SV40-immortalized keratinocytes are resistant to this cytokine (1). In this report, we investigated the expression of mitotic regulatory proteins, such as cyclin A, cyclin B, and p34cdc2. After exposure to TNF-alpha, normal and HPV-16-immortalized cells exhibited a dramatic decrease in the expression of these proteins. In contrast, no alteration in the levels of these proteins was observed after treatment of the resistant cell lines, as well as two HPV-positive cervical carcinoma cell lines. Expression of cyclin E does not seem to be modulated by TNF-alpha in any of the cells tested. On the other hand, cyclin D1, expression is slightly increased in normal keratinocytes and in the HPV-16-immortalized cells, whereas no alteration was observed in the HPV-18-transfected cells. The phosphorylation state of pRb correlated with cell growth; sensitive cells, which accumulate in G0-G1, after exposure to TNF-alpha, exhibited an accumulation of hypophosphorylated pRb, whereas no effect on pRb phosphorylation was observed for HPV-18-immortalized cells. These results clearly correlate with TNF-alpha-induced growth arrest in G0-G1.     

Semiempirical,squeeze flow,and intermolecular diffusion model. II. Model verification using laser microwelding

This article reviews the application of a coupled squeeze flow and intermolecular diffusion model, which was used to predict the quality and size of microwelds in plastics. Weld widths predictions were compared with previously presented experimental results using moving heat source models and temperature fields. The motivation for this work was to develop and verify a model based on fundamental principles that could accurately predict weld size and strength for conventional plastic welding techniques as well as novel techniques such as laser microwelding. It is envisioned that the resulting model could be used to predict proper welding parameters, including laser power and travel speed, to produce welds of varying size. Although insight into weld quality can be derived from this model, it was not the goal of this work to accurately predict weld strength for laser microwelding because of the difficulty in measuring weld strength on the micron scale. However, as reported in Part 1, weld strength for impulse welds were accurately predicted. In this model it was found that variable temperature histories, rather than a single value of maximum weld temperature, allows more accurate modeling of the welding process. In this work (Part 2), microwelds as small as 11 μm in width were produced with transmission infrared welding. In addition, welds over 150‐μm wide were also generated and the model was able to predict the range of weld widths that were found experimentally. It was found that the predictions were in very good agreement with the experimental results. There was some deviation between the experimental data and the model at the extreme parameters and it is believed that this was due to the temperature‐dependent material properties as well as optical aberrations. POLYM. ENG. SCI., 2008. © 2008 Society of Plastics Engineers     

Molecular follow-up of disease progression and interferon therapy in chronic myelocytic leukemia

We previously reported that the abl promoter (Pa) undergoes de novo DNA methylation in the course of chronic myelocytic leukemia (CML). The clinical implications of this finding are the subject of the present study in which samples of CML patients, including a group treated with interferon alpha (IFNalpha) were surveyed. The methylation status of the abl promoter was monitored by polymerase chain reaction (PCR) amplification of the Pa region after digestion with several site-methylation sensitive restriction enzymes. Some 74% of the DNA samples from blood and marrow drawn in the chronic phase were nonmethylated, similar to control samples from non-CML patients. The remaining 26% were partially methylated in the abl Pa region. The latter samples were derived from patients who were indistinguishable from the others on the basis of clinical presentation. Methylated samples were mostly derived from patients known to have a disease of longer duration (26 months v 7.5 months, P = .01). Samples of 30 IFNalpha-treated patients were sequentially analyzed in the course of treatment. Fifteen patients with no evidence of Pa methylation before treatment remained methylation-free. The remainder, who displayed Pa methylation before treatment, reverted to the methylation-free status. The outcome is attributed to IFNalpha therapy, as the Pa methylation status was not reversed in any of the patients treated with hydroxyurea. Methylation of the abl promoter indicates a disease of long-standing, most likely associated with a higher probability of imminent blastic transformation. It appears to predict the outcome of IFNalpha therapy far better than the cytogenetic response.     

Recovery of gut-associated lymphoid tissue and upper respiratory tract immunity after parenteral nutrition

OBJECTIVE: The authors characterize the recovery of parenteral nutrition-induced changes in gut-associated lymphoid tissue (GALT) and upper respiratory tract immunity with enteral nutrition and provide further information defining the effects of enteral feeding on mucosal immunity. SUMMARY BACKGROUND DATA: The small intestine plays a prominent role in development and maintenance of mucosal immunity, both intestinal and extraintestinal, primarily through immunoglobulin A (IgA)-mediated mechanisms. Prior research has shown that mice fed total parenteral nutrition (TPN) have reduced GALT T and B cells, the cells responsible for IgA production, as well as impaired upper respiratory tract immunity to viral challenge of previously immunized animals. The recovery of TPN-induced changes in GALT and upper respiratory tract immunity after enteral refeeding is studied. METHODS: Male institute of Cancer Research mice received 5 days of TPN followed by 0 to 4 days of chow. Small intestinal GALT was characterized by flow cytometry. In a second experiment, animals were immunized intranasally with moused-adapted influenza virus. Three weeks later, one group received a 5-day course of TPN followed by enteral refeeding for 5 days. A second group received TPN alone. Both groups were challenged with intranasal virus and killed 40 hours postchallenge to determine viral shedding from the upper respiratory tract. RESULTS: Animals fed TPN only had significantly fewer GALT lymphocytes compared with those chow-fed control subjects. Peyer's patch counts increased after a single day of refeeding, returning to normal levels by 48 hours. Lamina propria counts remained significantly depressed after 24 hours of refeeding, but also returned to normal after 48 hours of refeeding. The T-cell and B-cell populations mimicked total cell patterns. Lamina propria CD4+/CD8+ ratio returned to normal only after 72 hours of refeeding. None of the 9 animals refed enterally for 5 days were positive for viral shedding, compared with 8 of 12 matched TPN-fed animals. CONCLUSIONS: Enteral refeeding after TPN is associated with rapid repletion of GALT cellularity, initially within Peyer's patches and subsequently within the lamina propria. Refeeding corrects the impairment of IgA-mediated upper respiratory tract antiviral immunity occurring with TPN administration. This work further enhances the authors' knowledge of the underlying immunologic differences influenced by routes of nutrition.     

"I Buy American": The American Image as Reflected in Israeli Advertising

Analysis of advertisements in Israeli newspapers over the past decade has shown that American symbols have been widely used to market an array of consumer goods. Products made in America, Israel, or other countries are marketed with some sort of American angle by invoking America's values, symbols, landscapes, or lifestyle. Using both qualitative and quantitative methods, we analyzed how central components of the nation-state/national sphere—language, flag, political leaders, borders, landscapes, and sights—are reflected in advertising. We attempt to show how social values metamorphose, shedding their "Israeliness" and taking on an American veneer. Using Israel as an example may provide a test case for the Americanization process in other societies at the turn of the millennium.     

Requirement of cysteine-rich repeats of the Fas receptor for binding by the Fas ligand

The Fas receptor is a member of a family of cell death receptors, including tumor necrosis factor receptor I (TNFR I), death receptor 3 and 4 (DR3 and DR4), and cytopathic avian receptor 1 (CAR1). The Fas receptor is composed of several discrete domains, including three cysteine-rich domains (CRDs), a transmembrane domain, and an intracellular domain responsible for transmitting an apoptotic signal. While the mechanism of Fas-mediated cell death has become elucidated, the requirements for Fas ligand binding to the receptor have not been fully defined. Using a series of chimeric Fc-receptor fusion proteins between the human Fas receptor and TNFR I, each cysteine-rich domain of Fas was found to be required for interaction with the Fas ligand. Interestingly, TNFR I CRD1 could partially substitute for the Fas CRD1. The importance of this domain was underscored by the analysis of a Fas extracellular mutation (C66R), which resulted in a complete loss of ligand binding. This mutation was cloned from a human patient suffering from Canale-Smith syndrome, which is characterized by autoimmunity resembling that observed in the lpr and lprcg mice. The localization of essential ligand binding domains in the Fas receptor correlated exactly with the ability of the Fas receptor fusion proteins to prevent cell death mediated by the Fas ligand.     

RAFTK, a novel member of the focal adhesion kinase family, is phosphorylated and associates with signaling molecules upon activation of mature T lymphocytes

The related adhesion focal tyrosine kinase (RAFTK), a recently discovered member of the focal adhesion kinase family, has previously been reported to participate in signal transduction in neuronal cells, megakaryocytes, and B lymphocytes. We have found that RAFTK is constitutively expressed in human T cells and is rapidly phosphorylated upon the activation of the T cell receptor (TCR). This activation also results in an increase in the autophosphorylation and kinase activity of RAFTK. After its stimulation, there was an increase in the association of the src cytoplasmic tyrosine kinase Fyn and the adapter protein Grb2. This association was mediated through the SH2 domains of Fyn and Grb2. RAFTK also co-immunoprecipitates with the SH2 domain of Lck and with the cytoskeletal protein paxillin through its COOH-terminal proline-rich domain. The tyrosine phosphorylation of RAFTK after T cell receptor-mediated stimulation was reduced by the pretreatment of cells with cytochalasin D, suggesting the role of the cytoskeleton in this process. These observations indicate that RAFTK participates in T cell receptor signaling and may act to link signals from the cell surface to the cytoskeleton and thereby affect the host immune response.     

Identification of an interaction between the m-band protein skelemin and beta-integrin subunits. Colocalization of a skelemin-like protein with beta1- and beta3-integrins in non-muscle cells

Signaling across integrins is regulated by interaction of these receptors with cytoskeletal proteins and signaling molecules. To identify molecules interacting with the cytoplasmic domain of the beta3-integrin subunit (glycoprotein IIIa), a placental cDNA library was screened in the yeast two-hybrid system. Two identical clones coding for a 96-amino acid sequence were identified. This sequence was 100% identical to a sequence in skelemin, a protein identified previously in skeletal muscle. Skelemin is a member of a superfamily of cytoskeletal proteins that contain fibronectin-type III-like motifs and immunoglobulin C2-like motifs and that regulate the organization of myosin filaments in muscle. The amino acid residues in the isolated clones encompassed C2 motifs 4 and 5 of skelemin. A recombinant skelemin protein consisting of C2 motifs 3-7 interacted with beta1- and beta3-integrin cytoplasmic domains expressed as glutathione S-transferase (GST) fusion proteins, but not with GST-beta2-integrin cytoplasmic tail or GST alone. The skelemin-binding region was in the membrane proximal cytoplasmic domains of the integrins. Full-length skelemin interacted with integrin in intact cells as demonstrated by the colocalization of hemagglutinin-tagged skelemin in Chinese hamster ovary (CHO) cells containing alphaIIbbeta3-integrin and by the finding that microinjection of C2 motif 4 of skelemin into C2C12 mouse myoblast cells caused spread cells to round up. A skelemin-like protein was detected in CHO cells, endothelial cells, and platelets, and this protein colocalized with beta1- and beta3-integrins in CHO cells. This study suggests the presence of a skelemin-like protein in non-muscle cells and provides evidence that it may be involved in linking integrins to the cytoskeleton.