The isolation of cDNA clones from cucumber (Cucumis sativus L.) floral buds coming from plants differing in sex

In this study, we found flower cDNA clones which may be connected with the development of flower sex in cucumber. Two pairs of nearly-isogenic lines: gynoecious GY3 (FFMMGG) versus hermaphrodite HGY3 (FFmmGG) and monoecious B10 (ffMMGG) versus gynoecious 2gg (ffMMgg) were used for clone isolation. To obtain differentially-expressed clones, we applied the differential screening method. 454 clones from GY3 and 478 from B10 cDNA libraries were isolated. The results of RFLP analysis with 56 cDNA clones showed no clones which cosegregated with sex in cucumber. The 28 cDNA B10 and 33 cDNA GY3 clones isolated using the differential screening method were sequenced. Some of them seem to may play a role in cell differentiation or flower development. Among the 61 identified clones, 14 show high homology to plant proteins, although of unknown function. 11 show high homology to known proteins, and the possible function of some of them is discussed. For 3 clones, no significant similarity was found. The 31 clones displayed high homology to plant cDNA in EST database. The patterns of expression of five differential cDNA clones, 35GY3, 216GY3, 47GY3, 100B10 and 157B10, were analyzed in cucumber flower buds using in situ RT-PCR. The most interesting clone is 35GY3, because of its possible role in the inhibition of the development of male specific elements in the female cucumber flower.     

Circulating VEGF and its soluble receptors sVEGFR-1 and sVEGFR-2 in patients with acute leukemia

Angiogenesis plays an important role in the pathogenesis of acute leukemia, and vascular endothelial growth factor (VEGF) is a crucial, positive regulator of this process. The biological activity of VEGF is mediated by two different receptor tyrosine kinases: VEGFR-2 and VEGFR-1. The soluble form of VEGFR-1 is likely to be a negative regulator of VEGF availability, but the physiological role of sVEGFR-2 is still unclear. The plasma levels of sVEGFR-1 and sVEGFR-2 in patients with acute leukemia have not been investigated. We measured the plasma concentrations of VEGF and its two soluble receptors in 39 AML and 15 ALL patients as well as in the control group, using the ELISA assay. We also correlated the plasma levels of these proteins with disease status and known prognostic factors. The sVEGFR-1 level was significantly higher in patients with AML and ALL than in the healthy subjects (p < 0.002 and p < 0.03 respectively). The sVEGFR-2 level was significantly higher in AML patients compared with the control group (p < 0.03). The VEGF levels in AML and ALL patients and in healthy subjects did not differ significantly. The sVEGFR-1 level was higher in AML patients with > 50% of blasts in the bone marrow (BM), WBC > 20 G/L and elevated LDH level, than in the group with BM blasts < 50% (p < 0.01), WBC < 20 G/L (p < 0.02) and a normal LDH level (p < 0.05). Positive correlations between sVEGFR-1 level and WBC (p < 0.02),% of BM blasts (p < 0.05), the absolute blast count in peripheral blood (ABC) (p < 0.009) and LDH (p < 0.000001) were found. The sVEGFR-1/VEGF ratio (R1) was calculated, and a positive correlation between R1 and ABC in AML (p < 0.03) was determined. A higher (above median) sVEGFR-1/VEGF ratio correlated with a lower CR rate and a shorter survival (p < 0.03 and p = 0.0007 respectively). In conclusion, the plasma concentration of sVEGFR-1 is higher in leukemia patients than in healthy subjects and correlates with tumour burden and poor prognosis. The sVEGFR-1/VEGF ratio may be of greater prognostic value than VEGF alone. Further investigation is recommended to better determine their function.     

Generating social network data using partially described networks: an example informing avian influenza control in the British poultry industry

Background

The United Kingdom (UK) government has been recording the births, deaths, and movements of cattle for the last decade. Despite reservations about the accuracy of these data, they represent a large and valuable body of information about the demographics of the UK cattle herd and its contact structure. In this article, a range of demographic data about UK cattle, and particularly their movements, are presented, as well as yearly trends in the patterns of movements.

Results

A clear seasonal pattern is evident in the number of movements of cattle, as are the reductions in movement volume due to foot and mouth disease outbreaks in 2001 and 2007. The distribution of ages of cattle at their time of death is multimodal, and the impact of the over thirty months rule is marked. Most movements occur between agricultural holdings, markets, and slaughterhouses, and there is a non-random pattern to the types of holdings movements occur between. Most animals move only a short distance and a few times in their life. Most movements between any given pair of holdings only occurred once in the last 10 years, but about a third occurred between 2 and 10 times in that period. There is no clear trend to movement patterns in the UK since 2002.

Conclusions

Despite a substantial number of regulatory interventions during the last decade, movement patterns show no clear trend since 2002. The observed patterns in the repeatability of movements, the types of holdings involved in movements, the distances and frequencies of cattle movements, and the batch sizes involved give an insight into the structure of the UK cattle industry, and could act as the basis for a predictive model of livestock movements in the UK.     

Generation of *OH initiated by interaction of Fe2+ and Cu+ with dioxygen; comparison with the Fenton chemistry

Iron and copper toxicity has been presumed to involve the formation of hydroxyl radical (*OH) from H2O2 in the Fenton reaction. The aim of this study was to verify that Fe2+-O2 and Cu+-O2 chemistry is capable of generating *OH in the quasi physiological environment of Krebs-Henseleit buffer (KH), and to compare the ability of the Fe2+-O2 system and of the Fenton system (Fe2+ + H2O2) to produce *OH. The addition of Fe2+ and Cu+ (0-20 microM) to KH resulted in a concentration-dependent increase in *OH formation, as measured by the salicylate method. While Fe3+ and Cu2+ (0-20 microM) did not result in *OH formation, these ions mediated significant *OH production in the presence of a number of reducing agents. The *OH yield from the reaction mediated by Fe2+ was increased by exogenous Fe3+ and Cu2+ and was prevented by the deoxygenation of the buffer and reduced by superoxide dismutase, catalase, and desferrioxamine. Addition of 1 microM, 5 microM or 10 microM Fe2+ to a range of H2O2 concentrations (the Fenton system) resulted in a H2O2-concentration-dependent rise in *OH formation. For each Fe2+ concentration tested, the *OH yield doubled when the ratio [H2O2]:[Fe2+] was raised from zero to one. In conclusion: (i) Fe2+-O2 and Cu+-O2 chemistry is capable of promoting *OH generation in the environment of oxygenated KH, in the absence of pre-existing superoxide and/or H2O2, and possibly through a mechanism initiated by the metal autoxidation; (ii) The process is enhanced by contaminating Fe3+ and Cu2+; (iii) In the presence of reducing agents also Fe3+ and Cu2+ promote the *OH formation; (iv) Depending on the actual [H2O2]:[Fe2+] ratio, the efficiency of the Fe2+-O2 chemistry to generate *OH is greater than or, at best, equal to that of the Fe2+-driven Fenton reaction.     

Polymorphisms in the glucocorticoid receptor gene that modulate glucocorticoid sensitivity are associated with rheumatoid arthritis

Introduction  

The glucocorticoid receptor (GR) plays an important regulatory role in the immune system. Four polymorphisms in the GR gene are associated with differences in glucocorticoid (GC) sensitivity; the minor alleles of the polymorphisms N363 S and BclI are associated with relative hypersensitivity to GCs, while those of the polymorphisms ER22/23EK and 9β are associated with relative GC resistance. Because differences in GC sensitivity may influence immune effector functions, we examined whether these polymorphisms are associated with the susceptibility to develop Rheumatoid Arthritis (RA) and RA disease severity.     

High serum level of endostatin in multiple myeloma at diagnosis but not in the plateau phase after treatment

We investigated the serum concentration of endostatin in 84 patients with multiple myeloma (MM) and in 13 healthy controls. The level of measured anti-angiogenic agent was correlated with the phase and stage of the disease, and most importantly with clinical and laboratory parameters depicting the disease activity (haemoglobin, creatinine, albumins, calcium, M-component, C-reactive protein, beta2-microglobulin, lactate dehydrogenase, stage of bone disease) as well as serum levels of pro-angiogenic cytokines such as vascular endothelial growth factor, hepatocyte growth factor, fibroblast growth factor and transforming growth factor-beta. The median serum level of endostatin in MM patients was 58 ng/ml and was statistically significantly higher than in the control group (median, 40 ng/ml; p=0.015). MM patients in phase I (at diagnosis) had higher levels of endostatin (median, 69 ng/ml) than those in phase II (plateau phase after treatment) (median, 49 pg/ml; p=0.044). We did not find any statistical correlation between the level of endostatin and stage of MM according to the Durie and Salmon system. The serum concentration of endostatin in MM patients with a normal level of albumins was significantly higher than in others with hypoalbuminaemia (median, 62 ng/ml versus 39 ng/ml; p=0.033). Also, patients with a normal value of lactate dehydrogenase had a higher concentration of endostatin than those with values >425 U/l (median, 70 ng/ml versus 39 ng/ml; p=0.019). We did not show any statistical correlation between the concentration of endostatin and level of haemoglobin, creatinine, calcium, C-reactive protein, beta2-microglobulin and stage of bone disease. We failed to find positive or negative correlations between the level of endostatin and vascular endothelial growth factor, hepatocyte growth factor, fibroblast growth factor and transforming growth factor-beta. The concentration of endostatin did not influence the probability of survival in MM patients in our study. In conclusion, our data indicate that endostatin has a higher level in MM patients than in healthy controls. Highest values were stated in active phases of the disease (at presentation and in progression). Different clinical and laboratory parameters generally do not influence the concentration of endostatin (except albumins and lactate dehydrogenase).     

An intragenic distribution bias of DNA uptake sequences in <Emphasis Type="Italic">Pasteurellaceae</Emphasis> and <Emphasis Type="Italic">Neisseriae</Emphasis>

   

Most sequenced strains from Pasteurellaceae and Neisseriae contain hundreds to thousands of uptake sequence (US) motifs in their genome, which are associated with natural competence for DNA uptake. The mechanism of their recognition is still unclear, and I searched for intragenic location patterns of these motifs for clues about their distribution. In all cases, one orientation of the US has a higher occurrence in the reading frame, and in all Pasteurellaceae, the US and the reverse complement motifs are biased towards the gene termini. These findings could help design experimental set-ups to study preferential DNA uptake, thereby further unravelling the phenomenon of natural competence.     

Mathematical modelling of the influence of heat shock proteins on cancer invasion of tissue

Tumour cell invasion is crucial for cancer metastasis, which is the main cause of cancer mortality. An important group of proteins involved in cancer invasion are the Heat Shock Proteins (HSPs). According to experimental data, inhibition of one of these proteins, Hsp90, slows down cancer cells while they are invading tissue, but does not affect the synthesis of matrix metalloproteinases (MMP2 and MMP9), which are very important for cancer metastasis, acting as extracellular matrix (ECM) degrading enzymes. To test different biological hypotheses regarding how precisely Hsp90 influences tumour invasion, in this paper we use a model of solid tumour growth which accounts for the interactions between Hsp90 dynamics and the migration of cancer cells and, alternatively, between Hsp90 dynamics and the synthesis of matrix degrading enzymes (MDEs). The model consists of a system of reaction-diffusion-taxis partial differential equations describing interactions between cancer cells, MDE, and the host tissue (ECM). Using numerical simulations we investigate the effects of the administration of Hsp90 inhibitors on the dynamics of tumour invasion. Alternative mechanisms of reduction of cancer invasiveness result in different simulated patterns of the invading tumour cells. Therefore, predictions of the model suggest experiments which might be performed to develop a deeper understanding of the tumour invasion process.     

Combined effects of singlet oxygen and hydroxyl radical in photodynamic therapy with photostable bacteriochlorins: evidence from intracellular fluorescence and increased photodynamic efficacy in vitro

Sulfonamides of halogenated bacteriochlorins bearing Cl or F substituents in the ortho positions of the phenyl rings have adequate properties for photodynamic therapy, including strong absorption in the near-infrared (λ(max) ≈ 750 nm, ε ≈ 10(5) M(-1) cm(-1)), controlled photodecomposition, large cellular uptake, intracellular localization in the endoplasmic reticulum, low cytotoxicity, and high phototoxicity against A549 and S91 cells. The roles of type I and type II photochemical processes are assessed by singlet oxygen luminescence and intracellular hydroxyl radical detection. Phototoxicity of halogenated sulfonamide bacteriochlorins does not correlate with singlet oxygen quantum yields and must be mediated both by electron transfer (superoxide ion, hydroxyl radicals) and by energy transfer (singlet oxygen). The photodynamic efficacy is enhanced when cellular death is induced by both singlet oxygen and hydroxyl radicals.     

Engagement of the OX-40 receptor in vivo enhances antitumor immunity

The OX-40 receptor (OX-40R), a member of the TNFR family, is primarily expressed on activated CD4+ T lymphocytes. Engagement of the OX-40R, with either OX-40 ligand (OX-40L) or an Ab agonist, delivers a strong costimulatory signal to effector T cells. OX-40R+ T cells isolated from inflammatory lesions in the CNS of animals with experimental autoimmune encephalomyelitis are the cells that respond to autoantigen (myelin basic protein) in vivo. We identified OX-40R+ T cells within primary tumors and tumor-invaded lymph nodes of patients with cancer and hypothesized that they are the tumor-Ag-specific T cells. Therefore, we investigated whether engagement of the OX-40R in vivo during tumor priming would enhance a tumor-specific T cell response. Injection of OX-40L:Ig or anti-OX-40R in vivo during tumor priming resulted in a significant improvement in the percentage of tumor-free survivors (20-55%) in four different murine tumors derived from four separate tissues. This anti-OX-40R effect was dose dependent and accentuated tumor-specific T cell memory. The data suggest that engagement of the OX-40R in vivo augments tumor-specific priming by stimulating/expanding the natural repertoire of the host's tumor-specific CD4+ T cells. The identification of OX-40R+ T cells clustered around human tumor cells in vivo suggests that engagement of the OX-40R may be a practical approach for expanding tumor-reactive T cells and thereby a method to improve tumor immunotherapy in patients with cancer.