Fecal expression of Escherichia coli lysine decarboxylase (LdcC) is downregulated in E-cadherin negative lobular breast carcinoma.

Breast cancer is characterized by oncobiosis, the abnormal composition of the microbiome in neoplastic diseases. The biosynthetic capacity of the oncobiotic flora in breast cancer is suppressed, as suggested by metagenomic studies. The microbiome synthesizes a set of cytostatic and antimetastatic metabolites that are downregulated in breast cancer, including cadaverine, a microbiome metabolite with cytostatic properties. We set out to assess how the protein expression of constitutive lysine decarboxylase (LdcC), a key enzyme for cadaverine production, changes in the feces of human breast cancer patients (n = 35). We found that the fecal expression of Escherichia coli LdcC is downregulated in lobular cases as compared to invasive carcinoma of no special type (NST) cases. Lobular breast carcinoma is characterized by low or absent expression of E-cadherin. Fecal E. coli LdcC protein expression is downregulated in E-cadherin negative breast cancer cases as compared to positive ones. Receiver operating characteristic (ROC) analysis of LdcC expression in lobular and NST cases revealed that fecal E. coli LdcC protein expression might have predictive values. These data suggest that the oncobiotic transformation of the microbiome indeed leads to the downregulation of the production of cytostatic and antimetastatic metabolites. In E-cadherin negative lobular carcinoma that has a higher potential for metastasis formation, the protein levels of enzymes producing antimetastatic metabolites are downregulated. This finding represents a new route that renders lobular cases permissive for metastasis formation. Furthermore, our findings underline the role of oncobiosis in regulating metastasis formation in breast cancer.

Investigation of surface and filtration properties of TiO2 coated ultrafiltration polyacrylonitrile membranes.

In the present work, the surface and filtration properties of TiO2 coated polyacrylonitrile ultrafiltration membranes were investigated. The membranes were coated using the physical deposition method. The appropriate TiO2 coverage proved to be 0.3 mg/cm2, which formed a hydrophilic cake layer on the membrane surface. The cleanability without chemicals and the retention of the coated membranes was compared to the neat membrane after model oily wastewater filtration. The cleaning sustained of rinsing with distilled water and ultraviolet (UV) irradiation of the fouled membranes. The coated membranes have better antifouling properties; higher flux values during oily water filtration and by the mentioned cleaning process a significantly better flux recovery can be achieved. The amount of the catalyst and the irradiation time are limiting factors to the effectiveness of the cleaning process. The UV irradiation increases the wettability of the fouled membrane surface by degrading the oil layer. The coating, the continuous use, and the cleaning process do not significantly affect the membrane retention expressed in chemical oxygen demand.

Alteration of sarcoplasmic reticulum Ca2+ ATPase expression in lower limb ischemia caused by atherosclerosis obliterans.

Atherosclerosis is a disease caused by a build-up of fatty plaques and cholesterol in the arteries. The lumen of the vessels is obliterated resulting in restricted blood supply to tissues. In ischemic conditions, the cytosolic Ca2+ level of skeletal muscle may increase, indicating the alteration of Ca2+ removal mechanisms. Ca2+ is transported from cytosol into the sarcoplasmic reticulum by Ca2+ ATPase (SERCA), with its 1a isoform expressed in adult, while its 1b isoform in neonatal and regenerating fast-twitch skeletal muscle. To investigate the role of these isoforms in ischemic skeletal muscle, biopsies from musculus biceps femoris of patients who underwent amputation due to atherosclerosis were examined. Samples were removed from the visibly healthy and hypoxia-affected tissue. Significantly increased SERCA1a expression was detected under the ischemic conditions (246 ± 69%; p < 0.05) compared with the healthy tissue. Furthermore, the ratio of SERCA1a-positive fibers was slightly increased (46 ± 4% in healthy tissue and 60 ± 5% in ischemic tissue; p > 0.05), whereas SERCA2a did not change. In addition, in primary cultures derived from hypoxia-affected tissue, the diameter and fusion index of myotubes were significantly increased (30 ± 1.6 µm vs. 41 ± 2.4 µm and 31 ± 4% vs. 45 ± 3%; p < 0.05). We propose that the increased SERCA1a expression indicates the existence and location of compensating mechanisms in ischemic muscle.

Body composition assessment of Crohn's outpatients and comparison with gender- and age-specific multiple matched control pairs.

BACKGROUND/OBJECTIVES: Routine clinical care for Crohn's disease (CD) outpatients does not cover the assessment of body composition (BC); although this disease (because of inflammation, surgeries, lack of physical activity and appetite) may have a severe impact on lean body mass. The main aims of this prospective research were to assess the nutritional status of the patients and to compare their data with apparently healthy gender- and age-specific matched control pairs. SUBJECTS/METHODS: Overall, 136 CD patients and 1752 apparently healthy people were involved in the study. All participants were measured by the same bioelectrical impedance analyser. RESULTS: Using body mass index (BMI) and fat-free mass index (FFMI) as the markers of nutritional status, we found low BMI for 21% of the patients and low FFMI for 30% of them. Low BMI values were not gender specific, but substantially more females had low FFMI values. Low BMI was diagnosed in the patients' vs the control group for 21 vs 4% for men and 21 vs 2% for women; whereas low FFMI was diagnosed for 25 vs 5% for men and 36 vs 14% for women. Significant differences were found between patients' and control groups (median BMI: 22.0 vs 25.1 kg/m2, P<0.0001; FFMI: 17.3 vs 18.4 kg/m2, P=0.0044). CONCLUSIONS: This study confirmed the higher prevalence of low FFMI than that of low BMI among the subjects. We recommend implementing the assessment of BC into routine clinical care to diagnose low FFMI and to start intervention in time.

Capacity to improve fine motor skills in Williams syndrome.

BACKGROUND: Individuals with Williams syndrome (WS) are known to have difficulties in carrying out fine motor movements; however, a detailed behavioural profile of WS in this domain is still missing. It is also unknown how great the capacity to improve these skills with focused and extensive practice is. METHOD: We studied initial performance and learning capacity in a sequential finger tapping (FT) task in WS and in typical development. Improvement in the FT task has been shown to be sleep dependent. WS subjects participating in the current study have also participated in earlier polysomnography studies, although not directly related to learning. RESULTS: WS participants presented with great individual variability. In addition to generally poor initial performance, learning capacity was also greatly limited in WS. We found indications that reduced sleep efficiency might contribute to this limitation. CONCLUSIONS: Estimating motor learning capacity and the depth of sleep disorder in a larger sample of WS individuals might reveal important relationships between sleep and learning, and contribute to efficient intervention methods improving skill acquisition in WS.

Occlusion of retinal capillaries caused by glial cell proliferation in chronic ocular inflammation.

The inner blood-retinal barrier is a gliovascular unit in which glial cells surround capillary endothelial cells and regulate retinal capillaries by paracrine interactions. During chronic ocular inflammation, microvascular complications can give rise to vascular proliferative lesions, which compromise visual acuity. This pathologic remodelling caused by proliferating Müller cells determines occlusion of retinal capillaries. The aim of the present study was to identify qualitative and quantitative alterations in the retinal capillaries in patients with post-traumatic chronic ocular inflammation or post-thrombotic vascular glaucoma. Moreover, we investigated the potential role of vascular endothelial growth factor (VEGF) and pro-inflammatory cytokines in retinal inflammation. Our electron microscopy findings demonstrated that during chronic ocular inflammation, thickening of the basement membrane, loss of pericytes and endothelial cells and proliferation of Müller cells occur with irreversible occlusion of retinal capillaries. Angiogenesis takes place as part of a regenerative reaction that results in fibrosis. We believe that VEGF and pro-inflammatory cytokines may be potential therapeutic targets in the treatment of this disease although further studies are required to confirm these findings.

Comparison of proton channel, phagocyte oxidase, and respiratory burst levels between human eosinophil and neutrophil granulocytes.

Robust production of reactive oxygen species (ROS) by phagocyte NADPH oxidase (phox) during the respiratory burst (RB) is a characteristic feature of eosinophil and neutrophil granulocytes. In these cells the voltage-gated proton channel (Hv1) is now considered as an ancillary subunit of the phox needed for intense ROS production. Multiple sources reported that the expression of phox subunits and RB is more intensive in eosinophils than in neutrophils. In most of these studies the eosinophils were not isolated from healthy individuals, and a comparative analysis of Hv1 expression had never been carried out. We performed a systematic comparison of the levels of essential phox subunits, Hv1 expression and ROS producing capacity between eosinophils and neutrophils of healthy individuals. The expression of phox components was similar, whereas the amount of Hv1 was ∼ 10-fold greater in eosinophils. Furthermore, Hv1 expression correlated with Nox2 expression only in eosinophils. Additionally, in confocal microscopy experiments co-accumulation of Hv1 and Nox2 at the cell periphery was observed in resting eosinophils but not in neutrophils. While phorbol-12-myristate-13-acetate-induced peak extracellular ROS release was ∼ 1.7-fold greater in eosinophils, oxygen consumption studies indicated that the maximal intensity of the RB is only ∼ 1.4-fold greater in eosinophils. Our data reinforce that eosinophils, unlike neutrophils, generate ROS predominantly extracellularly. In contrast to previous works we have found that the two granulocyte types display very similar phox subunit expression and RB capacity. The large difference in Hv1 expression suggests that its support to intense ROS production is more important at the cell surface.

Different effects of bortezomib on the expressions of various protein kinase C isoenzymes in T cells of patients with systemic lupus erythematosus and in Jurkat cells.

The effects of proteosome inhibitor Bortezomib (BZ) were studied in vitro for 24 h on the protein kinase C (PKC) profiles, rates of proliferation and apoptosis in Jurkat cells and lymphocytes of 10 patients with systemic lupus erythematosus (SLE) and nine healthy subjects. The expressions of PKC proteins, the rates of proliferation and apoptosis were determined. The effects of BZ were different in the Jurkat and lupus T cells. Whereas BZ elevated the expression of PKC θ, δ and ξ isoenzymes in the Jurkat cells, it was unable to do that in the lupus T cells. BZ induced a dose-dependent increase in the apoptosis of Jurkat cells, while decreased the proliferation. The same effect of BZ was observed on the apoptosis of lymphocytes both in SLE and healthy subjects at concentrations higher than the therapeutic dose. We conclude that BZ treatment in vitro was not able to restore the SLE-specific defect (decrease) in the expression of PKC isoenzymes in the T cells as it was expected. This can be a limiting factor in the positive clinical effects of BZ in lupus.

Atypical sleep architecture and altered EEG spectra in Williams syndrome.

BACKGROUND: Williams syndrome (WS) is a neurodevelopmental genetic disorder characterised by physical abnormalities and a distinctive cognitive profile with intellectual disabilities (IDs) and learning difficulties. METHODS: In our study, nine adolescents and young adults with WS and 9 age- and sex-matched typically developing (TD) participants underwent polysomnography. We examined sleep architecture, leg movements and the electroencephalogram (EEG) spectra of specific frequency bands at different scalp locations. RESULTS: We found an atypical, WS characteristic sleep pattern with decreased sleep time, decreased sleep efficiency, increased wake time after sleep onset, increased non-rapid eye movement percentage, increased slow wave sleep, decreased rapid eye movement sleep percentage, increased number of leg movements and irregular sleep cycles. Patients with WS showed an increased delta and slow wave activity and decreased alpha and sigma activity in the spectral analysis of the EEG. CONCLUSIONS: Sleep maintenance and organisation are significantly affected in WS, while EEG spectra suggest increases in sleep pressure.

Hyperpolarization-activated, cyclic nucleotide-gated, cation non-selective channel subunit expression pattern of guinea-pig spiral ganglion cells.

Although the hyperpolarization-activated non-specific cationic current (I(h)) plays important roles in determining the membrane characteristics of the spiral ganglion cells (SGCs), neither the exact types of the hyperpolarization-activated, cyclic nucleotide-gated, cation non-selective channel (HCN) subunits contributing to the molecular assembly of the relevant channels, nor their distribution pattern presented by the SGCs is known. In the present work immunolabeling and Western blot analysis were performed to describe the presence and distribution of all four known HCN subunits in the guinea-pig spiral ganglion. Besides determining the expression of the HCN1-HCN4 subunits by both type I and type II SGCs, the presence of possible apico-basal gradients in the expression patterns was also sought. The results indicate that both type I and type II SGCs express all four HCN subunits. The intensity of the immunolabeling of the cell surface membrane was generally strong, but it showed pronounced cell-to-cell variability. The Western blot experiments in combination with densitometry revealed that the amount of the HCN1 and HCN3 proteins was more significant in the apical than in the basal third of the guinea-pig cochlea. These findings not only imply potential heteromeric HCN channel formation of the spiral ganglion neurons, but they also offer a possible explanation of the previously reported heterogeneity of I(h) recorded in functional studies.

Density of critical clusters in strips of strongly disordered systems.

We consider two models with disorder-dominated critical points and study the distribution of clusters that are confined in strips and touch one or both boundaries. For the classical random bond Potts model in the large- q limit, we study optimal Fortuin-Kasteleyn clusters using a combinatorial optimization algorithm. For the random transverse-field Ising chain, clusters are defined and calculated through the strong-disorder renormalization group method. The numerically calculated density profiles close to the boundaries are shown to follow scaling predictions. For the random bond Potts model, we have obtained accurate numerical estimates for the critical exponents and demonstrated that the density profiles are well described by conformal formulas.

Expression of neurotransmitters and neurotrophins in neurogenic inflammation of the rat retina.

Antidromic stimulation of the rat trigeminal ganglion triggers the release of substance P (SP) and calcitonin gene-related peptide (CGRP) from sensory nerve terminals of the capsaicin sensitive C-fibers. These pro-inflammatory neuropeptides produce a marked hyperemia in the anterior segment of the eye, accompanied by increased intraocular pressure, breakdown of the blood-aqueous barrier and myosis. To assess the effects of neurogenic inflammation on the retina, specifically on the immunostaining of neurotransmitters and neurotrophins, as well as on the expression of neurotrophin receptors in the retina. RT-PCR was also accomplished in control and stimulated animals to confirm the immunohistochemical results. In the electrically stimulated eyes, immunostaining for SP, CGRP, VIP and nNOS demonstrated a marked increase in the RPE/POS (Retinal Pigment Epithelium/Photoreceptor Outer Segments), in the inner and outer granular layers and in the ganglion cells in comparison to the control eyes. CGRP and SP were found increased in stimulated animals and this result has been confirmed by RT- PCR. Changes in neurotrophin immunostaining and in receptor expression were also observed after electric stimulation of trigeminal ganglia. Decrease of BDNF and NT4 in the outer and inner layers and in ganglion cells was particularly marked. In stimulated rat retinas immunostaining and RT-PCR showed a NGF expression increase. Neurotrophin receptors remained substantially unchanged. These studies demonstrated, for the first time, that antidromic stimulation of the trigeminal ganglion and subsequent neurogenic inflammation affect immunostaining of retinal cell neurotransmitter/neuropeptides and neurotrophins as well as the expression of neurotrophin receptors.

Integrity of lateral and feedbackward connections in visual processing in children with pervasive developmental disorder.

Enhanced visual detail processing in subjects with pervasive developmental disorder (PDD) has been related to impairments in feature integration. The functional integrity of two types of neuronal connections involved in visual feature integration, namely horizontal and feedbackward connections, were tested. Sixteen children with PDD and 17 age- and IQ-matched control children (mean age 13.3 years) were included. In a texture segregation task the difference in ERP response to homogeneous and checkered visual stimuli was determined. Additionally, in a contour integration task subjects had to point out a contour consisting of colinearly aligned Gabor signals in backgrounds increasing in noise. Children with PDD showed a normal performance on the contour integration task, suggesting that neurons in the primary visual cortex of children with PDD can effectively integrate the activity of local detectors that process different aspects of the same object information by making use of long-range lateral connections. The amplitude of ERP activity related to texture segregation was also not different between the PDD and control groups, indicating functional visual feedback mechanisms between V1 and higher order areas in subjects with PDD. However, a difference in latency of texture-segmentation related activity between the groups was noted. This effect did not reach significance, which could be due to the small N of the study. Therefore, the data need replication in a study with larger samples before more definitive conclusions can be drawn.

Production of proinflammatory cytokines by syncytiotrophoblasts infected with human cytomegalovirus isolates.

Forty per cent of women with primary cytomegalovirus (CMV) infection during gestation transmit the infection to their fetuses, which may result in abnormalities for the newborn, varying in degree from mild to severe. The factors whereby CMV in the placenta develops into a fulminating infection and spreads to the fetus are not known. In this study the production of proinflammatory cytokines was investigated in syncytiotrophoblast (ST) cultures infected with CMV strains. The interrelationships between the cytokines produced in the ST cultures and the number of nuclei of ST expressing the CMV immediate-early (IE) gene were examined. To resemble a natural infection, clinical CMV isolates and a low multiplicity of infection were used. TNF-alpha and IL-1 beta were not detected in the supernatants of any ST cultures. Similar or increased amounts of IL-6 were found in the CMV-infected cultures. The IL-8-inducing capacities of the CMV strains differed in the ST cultures. The IE gene expression of the virus provided was dependent on the amount of IL-8 produced in the STs. Our observations indicate that certain CMV strains induce high amounts of IL-8, which in turn enhances CMV replication in the placenta, while others can replicate if the IL-8 is provided by a co-infecting agent.

Adenosine inhibits the release of arachidonic acid and its metabolites (AAM) in activated human peripheral mononuclear cells.

OBJECTIVE: The effects of adenosine (Ado) and subtype-specific activators of adenosine receptors (A(1), A(2A), A(2B) and A(3)) were studied on the release of arachidonic acid (AA) and its metabolites (AAM) from human peripheral mononuclear cells (monocytes). MATERIALS AND METHOD: Adenosine and the selective agonists and antagonists of adenosine receptors were used. (3)H-AA and its metabolites released into the medium were determined by measurement of the total (3)H radioactivity released without separating the AAM. RESULTS: In the cells activated by protein kinase C specific phorbol ester (phorbol 12-myristate 13-acetate) and Ca(2+) ionophore (A23187), adenosine and two subtype-specific receptor agonists, CPA(A(1)) and CGS 21680 (A(2A)) induced concentration-dependent inhibition of the release of AAM, whereas stimulation of A(2B) or A(3) receptors was ineffective. The rank order of potency for the inhibition of AAM release was as follows: CGS 21680 = CPA > adenosine > NECA (in the presence of ZM 24185 and DPCPX as A(2A) and A(1) adenosine receptor antagonists, respectively) = IB-MECA. Adenosine inhibited the release of AAM only at and above the concentration of 100 muM, whereas the inhibitory effect of A(1) and A(2A) receptor specific agonists appeared at a concentration of 10(-7) M. CONCLUSIONS: It can be concluded that adenosine physiologically may not have a significant effect on the AAM release of circulating monocytes, but in pathological conditions, where the local Ado concentrations increases, this nucleoside, through activation of A(2A) and A(1) receptors can exert, at least in part, an antiinflammatory action by decreasing proinflammatory AAM production.

Melanoma cells exhibit strong intracellular TASK-3-specific immunopositivity in both tissue sections and cell culture.

Amplification of the kcnk9 gene and overexpression of the encoded channel protein (TASK-3) seems to be involved in carcinogenesis. In the present work, TASK-3 expression of melanoma cells has been studied. For the investigation of TASK-3-specific immunolabelling, a monoclonal antibody has been developed and applied along with two, commercially available polyclonal antibodies targeting different epitopes of the channel protein. Both primary and metastatic melanoma cells proved to be TASK-3 positive, showing prominent intracellular TASK-3-specific labelling; mostly concentrating around or in the proximity of the nuclei. The immunoreaction was associated with the nuclear envelope, and with the processes of the cells and it was also present in the cell surface membrane. Specificity of the immunolabelling was confirmed by Western blot and transfection experiments. As TASK-3 immunopositivity of benign melanocytes could also be demonstrated, the presence or absence of TASK-3 channels cannot differentiate between malignant and non-malignant melanocytic tumours.

Enhanced spontaneous thrombolysis: a new therapeutic challenge.

Spontaneous thrombolysis is an endogenous protective mechanism against lasting arterial thrombotic occlusion, which is implicated in the pathogenesis of myocardial infarction and acute coronary events. Novel therapies for coronary heart disease (CHD) targeting atherosclerosis and thrombosis, together with cardiovascular prevention programs targeting risk-factors and lifestyle provide evidence that CHD is preventable. Although reduced fibrinolytic activity is a recognized risk-factor for ischemic cardiovascular events, it has so far been neglected. Our knowledge of the fibrinolytic effect of drugs commonly used for CHD such as antiplatelet agents (aspirin, ticlopidine, clopidogrel), anti-diabetic biguanides (phenformin, metformin) or anti-hypertensive drugs is scanty and conflicting. This is mainly due to the lack of a global test of spontaneous thrombolysis, as opposed to fibrinolysis of plasma or whole blood, i.e. the assessment of various activators and inhibitors of the fibrinolytic system. A recently described technique allows the measurement of spontaneous thrombolysis, that is, lysis of an autologous platelet-rich thrombus in the absence of added plasminogen activators. Early results suggest that this test may have significant clinical potential both in identifying those at risk of fatal cardiac events and in finding new therapeutic avenues or lifestyles to improve spontaneous thrombolytic activity.

Immunotoxicological investigation of subacute combined exposure with low doses of Pb, Hg and Cd in rats.

Detectable interactions between NOEL (No Observed Effect Level) doses of Pb, Hg and Cd in general toxicological, hematological, and immune function parameters were investigated. The metals (Pb-acetate, 20 mg/kg; HgCl2, 0.40 mg/kg; CdCl2, 1.61 mg/kg) were combined. First, the rats received the combination Pb + Hg + Cd for 4 weeks per os. Significant difference vs. control was found only in the weight of lung and popliteal lymph node (PLN). The Pb + Hg and Pb + Cd combinations significantly decreased the PLN to 100 g body weight and PLN to brain weight ratio, and Pb+Hg also decreased the relative adrenal weight. After 12 weeks treatment with the same doses, effects on the thymus, kidney, and adrenal weights in the Pb + Hg, and thymus weight in the Pb + Cd, combination were seen. Pb + Cd also affected the white and red blood cell count and hematocrit. Combined with Hg or Cd, NOEL dose Pb showed toxicity, indicating that exposure limits may be inefficient in combined exposure situations.