Signaling through Toll-like receptor 3 and Dectin-1 potentiates the capability of human monocyte-derived dendritic cells to promote T-helper 1 and T-helper 17 immune responses

Background aimsRecent studies have shown that the ligation of Toll-like receptor 3 (TLR3) or Dectin-1 on human monocyte-derived dendritic cells (MoDC) elicits their maturation, but with a different outcome on immunomodulation. Therefore the aim of this work was to study the response of MoDC to the combined effect of polyinosinic:polycytydilic acid [Poly (I:C)] and curdlan, selective TLR3 and Dectin-1 agonists, respectively.MethodsImmature MoDC, generated from human monocytes, were treated with Poly (I:C), curdlan or their combination for 2 days. Phenotypic characteristics of MoDC were determined by flow cytometry, and cytokine production was measured by enzyme-linked immunosorbent assay (ELISA) and FlowCytomix, while the stimulatory capability of MoDC was tested using a mixed leukocyte reaction assay.ResultsThe combination of Poly (I:C) and curdlan induced phenotypic maturation of MoDC with the capability to stimulate an alloreactive response. Such treated MoDC up-regulated the production of interleukin (IL)-12, IL-23 and IL-10, compared with the effect of Poly (I:C) alone. Curdlan-treated MoDC stimulated the production of IL-17 by alloreactive CD4 + T cells more strongly than Poly (I:C)-treated MoDC. The opposite effect was observed for interferon(IFN)-γ production. When combined, these agonists primed MoDC to increase further the production of IFN-γ by CD4 + T cells in co-culture, especially those of naive (CD45RA ⁺) phenotype, and IL-17 by memory (CD45RO +) CD4 + T cells.ConclusionsLigation of TLR3 and Dectin-1 receptor up-regulates T-helper (Th) 1 and Th17 immune responses compared with single agonists. These findings may have therapeutic implications for the use of MoDC in immunotherapy.     

High Immune Response Rates and Decreased Frequencies of Regulatory T Cells in Metastatic Renal Cell Carcinoma Patients after Tumor Cell Vaccination

Our previously reported phase I clinical trial with the allogeneic gene–modified tumor cell line RCC-26/CD80/IL-2 showed that vaccination was well tolerated and feasible in metastatic renal cell carcinoma (RCC) patients. Substantial disease stabilization was observed in most patients despite a high tumor burden at study entry. To investigate alterations in immune responses that might contribute to this effect, we performed an extended immune monitoring that included analysis of reactivity against multiple antigens, cytokine/chemokine changes in serum and determination of the frequencies of immune suppressor cell populations, including natural regulatory T cells (nTregs) and myeloid-derived suppressor cell subsets (MDSCs). An overall immune response capacity to virus-derived control peptides was present in 100% of patients before vaccination. Vaccine-induced immune responses to tumor-associated antigens occurred in 75% of patients, demonstrating the potent immune stimulatory capacity of this generic vaccine. Furthermore, some patients reacted to peptide epitopes of antigens not expressed by the vaccine, showing that epitope-spreading occurred in vivo. Frequencies of nTregs and MDSCs were comparable to healthy donors at the beginning of study. A significant decrease of nTregs was detected after vaccination (p = 0.012). High immune response rates, decreased frequencies of nTregs and a mixed T helper 1/T helper 2 (T_H1/T_H2)-like cytokine pattern support the applicability of this RCC generic vaccine for use in combination therapies.     

Identification of naturally processed hepatitis C virus-derived major histocompatibility complex class I ligands

Fine mapping of human cytotoxic T lymphocyte (CTL) responses against hepatitis C virus (HCV) is based on external loading of target cells with synthetic peptides which are either derived from prediction algorithms or from overlapping peptide libraries. These strategies do not address putative host and viral mechanisms which may alter processing as well as presentation of CTL epitopes. Therefore, the aim of this proof-of-concept study was to identify naturally processed HCV-derived major histocompatibility complex (MHC) class I ligands. To this end, continuous human cell lines were engineered to inducibly express HCV proteins and to constitutively express high levels of functional HLA-A2. These cell lines were recognized in an HLA-A2-restricted manner by HCV-specific CTLs. Ligands eluted from HLA-A2 molecules isolated from large-scale cultures of these cell lines were separated by high performance liquid chromatography and further analyzed by electrospray ionization quadrupole time of flight mass spectrometry (MS)/tandem MS. These analyses allowed the identification of two HLA-A2-restricted epitopes derived from HCV nonstructural proteins (NS) 3 and 5B (NS3_1406–1415 and NS5B_2594–2602). In conclusion, we describe a general strategy that may be useful to investigate HCV pathogenesis and may contribute to the development of preventive and therapeutic vaccines in the future.     

CO2 fixation and chlorophyll a fluorescence in leaves of Ramonda serbica during a dehydration-rehydration cycle

Changes in CO(2) photo-assimilation and PSII photochemical efficiency in Ramonda serbica leaves during a dehydration-rehydration cycle were examined. The rate of CO(2) photo-assimilation was greatly reduced during dehydration, but recovery was complete with rehydration when the relative water content of leaves reached values similar to those of well-hydrated, control leaves. The results showed that the response of R. serbica leaves to severe water stress involves two different mechanisms. In the first, CO(2) assimilation is limited by stomata closure that creates an excess proton concentration in the lumen and activates non-photochemical quenching. This plays an important role in the mechanism of photoprotection by dissipation of excitation energy. When dehydration became severe and leaf RWC reached very low values, the electron transport rate (ETR) decreased markedly, while the capacity for regulatory mechanisms such as q(NP) (non-photochemical quenching) was greatly reduced. For severely dehydrated leaves of R. serbica, it appears that reactive oxygen species (ROS) formation is better prevented by mechanisms that quench chlorophyll triplet formation via lutein.     

Effect of dietary zinc on the levels and distribution of fatty acids and vitamin A in blood plasma chylomicrons

The aim of this work was to explore the effects of a low- and high-zinc diet and vitamin Aon the distribution of fatty acids in chylomicrons. Mongolian Gerbils were fed a basal diet (for 3 wk) containing 8 or 38 mg zinc/kg of feed (low-zinc group [termed LZ group] and saturated zinc group [termed SZ group], respectively). The following day, the animals were given sunflower oil containing 50 nmol vitamin A. The results showed that the concentration of zinc in blood plasma was similar in both groups. The amount of plasma chylomicrons was lower in the LZ group than in the SZ group (p < 0.001). The concentration of retinol in blood plasma was lower in the LZ group than in the SZ group (p < 0.01). However, the results demonstrated an increase in the blood plasma retinol concentration in the LZ group compared to the SZ group when calculated per milligram of plasma chylomicrons (p < 0.01). In plasma chylomicrons, fatty acids corresponding to 16:0, 16:1, 17:0, 17:1, 18:0, 18:1, 18:2, 18:3, 20:0, 21:0, and 20:4 were detected. The fatty acid distribution was similar in both groups. There was no major difference in the concentration of fatty acids in plasma chylomicrons between both experimental groups, except for 20:4 (a lower amount was found in the SZ group). Our results show that dietary zinc influences both the amount of chylomicrons in blood plasma and the concentrations of retinol and arachidonic acid in chylomicrons.     

Anatomical Injury Induced by the Eriophyid Mite Aceria anthocoptes on the Leaves of Cirsium arvense

Anatomical injury of the leaves of the invasive species, Cirsium arvense (L.) Scop., caused by the eriophyid mite Aceria anthocoptes (Nal.), which is the only eriophyid mite that has been recorded on C. arvense worldwide, is described. The injury induced by the mite feeding on the leaves of C. arvense results in visible russeting and bronzing of the leaves. Other conspicuous deformations are folding and distortion of the leaf blade and curling of leaf edge, as well as gradual drying of leaves. The anatomical injury of the mature leaves of field-collected plants was limited to the epidermis of the lower leaf surface. However, on young leaves of experimentally infested plants, rust mite injuries extend to epidermal cells on both leaf surfaces and to those of deeper mesophyll layers. On these leaves, lesions on the lower leaf surface even affected the phloem of the vascular bundles. Leaf damage induced by A. anthocoptes is discussed with regard to the mite’s potential as a biological control agent of C. arvense.     

Impact of Carboxymethylcellulose on Morphology and Antibiotic Production by Streptomyces hygroscopicus

A chemically defined media consisting of carboxymethylcellulose (CMC) was developed to maximize the production of antibiotics, hexaene H-85 and azalomycine, by Streptomyces hygroscopicus CH-7. The production of antibiotics by filamentous organisms is often dependent on the morphology and size distribution of the pellet population within the culture. By adding the polymer to the fermentation medium, the growth was changed from a single large glob to small reproducible pellets, and wall growth was diminished to a minimum. Maximum concentrations of hexaene H-85 (146.7 mg/dm(3)) and azalomycine (188.6 mg/dm(3)) were reached at 3.0% and 1.0% (w/v) CMC, respectively.     

Absorbed fractions for electrons and beta particles in sensitive regions of human respiratory tract

The absorbed fractions (AF) of electrons in sensitive layers of human respiratory tract were calculated in this paper. For that purpose the source code for simulation package PENELOPE, based on Monte Carlo method, was developed. The human respiratory tract was modeled according to ICRP66 publication, where AF of electrons was calculated using EGS4 simulation software. Some approximations used in ICRP66 were corrected in this work, and new values of AF for radon progeny are given. Minimal energy (EABS) that electron can have during transport through material is 1 keV in ICRP66, while it is set as low as 100 eV in the presented work. Lowering value of EABS gives more accurate results for AF when initial energy of electrons is below 50 keV. To represent tissue, water is used in ICRP66, while in this work epithelia tissue is used.