An ex vivo readout for evaluation of dendritic cell-induced autologous cytotoxic T lymphocyte responses against esophageal cancer

Esophageal cancer is a highly malignant disease that despite surgery and adjuvant therapies has an extremely poor outcome. Dendritic cell (DC) immunotherapy as a novel promising strategy could be an alternative for treating this malignancy. Effective DC-mediated immune responses can be achieved by raising cytotoxic T lymphocyte (CTL) response against multiple antigens through loading DCs with total tumor RNA. However, the efficacy of this strategy first needs to be evaluated in a pre-clinical setting. The aim of the study was to set up an ex vivo autologous human readout assay for assessing the effects of DC-mediated cytotoxic responses, using total tumor RNA as an antigen load. Biopsy specimens of seven esophageal cancer patients were used to establish primary cultures of normal and cancer cells and to obtain autologous RNA for loading DCs. Mature DCs loaded with either normal or tumor RNA were obtained and subsequently used to raise various lymphocytes populations. Apoptosis levels of the autologous cultures were measured before and after incubating the cultures with the different lymphocytes populations. The mean apoptosis levels in the tumor cell cultures, induced by lymphocytes instructed by DCs loaded with tumor RNA, significantly increased with 15.6% ±2.9 SEM (range 3.4–24.5%, t-test, P < 0.05). Incubation of the normal cultures with the lymphocytes populations showed a mean non-significant increase in apoptosis of 0.4% ±3.4 SEM (range −13.9 to 9.8%, t-test, P = 0.7). Here, we introduce a practical, patient-specific autologous readout assay for pre-clinical testing of DC-mediated cytotoxic responses. Additionally, we demonstrated that the use of autologous tumor RNA as a strategy for raising cytotoxic responses against multiple tumor antigens could be effective for treating esophageal cancer. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Dual Roles of Capsular Extracellular Polymeric Substances in Photocatalytic Inactivation of Escherichia coli: Comparison of E. coli BW25113 and Isogenic Mutants

The dual roles of capsular extracellular polymeric substances (EPS) in the photocatalytic inactivation of bacteria were demonstrated in a TiO₂-UVA system, by comparing wild-type Escherichia coli strain BW25113 and isogenic mutants with upregulated and downregulated production of capsular EPS. In a partition system in which direct contact between bacterial cells and TiO₂ particles was inhibited, an increase in the amount of EPS was associated with increased bacterial resistance to photocatalytic inactivation. In contrast, when bacterial cells were in direct contact with TiO₂ particles, an increase in the amount of capsular EPS decreased cell viability during photocatalytic treatment. Taken together, these results suggest that although capsular EPS can protect bacterial cells by consuming photogenerated reactive species, it also facilitates photocatalytic inactivation of bacteria by promoting the adhesion of TiO₂ particles to the cell surface. Fluorescence microscopy and scanning electron microscopy analyses further confirmed that high capsular EPS density led to more TiO₂ particles attaching to cells and forming bacterium-TiO₂ aggregates. Calculations of interaction energy, represented by extended Derjaguin-Landau-Verwey-Overbeek (XDLVO) potential, suggested that the presence of capsular EPS enhances the attachment of TiO₂ particles to bacterial cells via acid-base interactions. Consideration of these mechanisms is critical for understanding bacterium-nanoparticle interactions and the photocatalytic inactivation of bacteria.     

Regulation of protein tyrosine phosphatase 1B by sumoylation

Protein-tyrosine phosphatase 1B (PTP1B) is an ubiquitously expressed enzyme that negatively regulates growth-factor signalling and cell proliferation by binding to and dephosphorylating key receptor tyrosine kinases, such as the insulin receptor. It is unclear how the activity of PTP1B is regulated. Using a yeast two-hybrid assay, a protein inhibitor of activated STAT1 (PIAS1) was isolated as a PTP1B-interacting protein. Here, we show that PIAS1, which functions as a small ubiquitin-like modifier (SUMO) E3 ligase, associates with PTP1B in mammalian fibroblasts and catalyses sumoylation of PTP1B. Sumoylation of PTP1B reduces its catalytic activity and inhibits the negative effect of PTP1B on insulin receptor signalling and on transformation by the oncogene v-crk. Insulin-stimulated sumoylation of endogenous PTP1B results in a transient downregulation of the enzyme; this event does not occur when the endogenous enzyme is replaced with a sumoylation-resistant mutant of PTP1B. These results suggest that sumoylation, which has been implicated primarily in processes in the nucleus and nuclear pore, also modulates a key enzyme-substrate signalling complex that regulates metabolism and cell proliferation.     

Effects of permanent magnetic fields on in vitro growth of Cymbidium and Spathiphyllum shoots

Magnetic fields affect biological systems. However, this is the first study on the effects of permanent magnetic fields (MFs) on the micropropagation of two ornamental plants, Spathiphyllum cv. i.e ‘Merry’ and Cymbidium Music Hour ‘Maria’. Cymbidium and Spathiphyllum shoots cultured in the ‘Miracle Pack’? culture system were exposed to MFs of different intensities, polarities, and duration of exposure. The results show that by increasing intensity from 5 × 10⁻⁶ Tesla (T) as the geo-magnetic field to 0.1, 0.15, and 0.2 T negatively influenced height and fresh mass of roots of Cymbidium plants (except for 0.1 T–S and 0.2 T–N treatments), but had no significant effect on other plantlet parameters. Long-term exposure (1, 2, or 3 mo) of Cymbidium shoots to 0.15 T–MFs negatively influenced plant height, positively affected the number of leaves (with the exception of 0.15 T–S—1 mo), and had no clear effect on other parameters compared to the control. MFs (0.1, 0.15, and 0.2 T), regardless of their polarity, increased chlorophyll content (SPAD value) and the number of leaves, but slightly decreased the dry mass of Spathiphyllum shoots. Different exposure duration to 0.15 T (i.e., 2, 4, or 8 wk) had no significant influence on Spathiphyllum plantlet development other than increasing the SPAD value. These two ornamentals could serve as model systems to study plant development, space production, yield maximization, and the development of new morphotypes essential for the floricultural market.