Narrow bandwidth, low-emittance positron beams from a laser-wakefield accelerator.

The rapid progress that plasma wakefield accelerators are experiencing is now posing the question as to whether they could be included in the design of the next generation of high-energy electron-positron colliders. However, the typical structure of the accelerating wakefields presents challenging complications for positron acceleration. Despite seminal proof-of-principle experiments and theoretical proposals, experimental research in plasma-based acceleration of positrons is currently limited by the scarcity of positron beams suitable to seed a plasma accelerator. Here, we report on the first experimental demonstration of a laser-driven source of ultra-relativistic positrons with sufficient spectral and spatial quality to be injected in a plasma accelerator. Our results indicate, in agreement with numerical simulations, selection and transport of positron beamlets containing N e + ≥ 10 5 positrons in a 5% bandwidth around 600 MeV, with femtosecond-scale duration and micron-scale normalised emittance. Particle-in-cell simulations show that positron beams of this kind can be guided and accelerated in a laser-driven plasma accelerator, with favourable scalings to further increase overall charge and energy using PW-scale lasers. The results presented here demonstrate the possibility of performing experimental studies of positron acceleration in a laser-driven wakefield accelerator.

Potential of nonpeptide (ant)agonists to rescue vasopressin V2 receptor mutants for the treatment of X-linked nephrogenic diabetes insipidus.

According to the body's need, water is reabsorbed from the pro-urine that is formed by ultrafiltration in the kidney. This process is regulated by the antidiuretic hormone arginine-vasopressin (AVP), which binds to its type 2 receptor (V2R) in the kidney. Mutations in the gene encoding the V2R often lead to the X-linked inheritable form of nephrogenic diabetes insipidus (NDI), a disorder in which patients are unable to concentrate their urine despite the presence of AVP. Many of these mutations are missense mutations that do not interfere with the intrinsic functionality of V2R, but cause its retention in the endoplasmic reticulum (ER), making it unavailable for AVP binding. Because the current treatments for NDI relieve its symptoms to some extent, but do not cure the disorder, cell-permeable antagonists (pharmacological chaperones) have been successfully used to stabilise the mutant receptors and restore their plasma membrane localisation. Recently, cell-permeable agonists also were shown to rescue ER-retained V2R mutants, leading to increased cAMP levels and translocation of aquaporin-2 to the apical membrane. This makes V2R-specific cell-permeable agonists very promising therapeutics for NDI as a result of misfolded V2R receptors.

Effects of essential fatty acid deficiency on enterohepatic circulation of bile salts in mice.

Essential fatty acid (EFA) deficiency in mice has been associated with increased bile production, which is mainly determined by the enterohepatic circulation (EHC) of bile salts. To establish the mechanism underlying the increased bile production, we characterized in detail the EHC of bile salts in EFA-deficient mice using stable isotope technique, without interrupting the normal EHC. Farnesoid X receptor (FXR) has been proposed as an important regulator of bile salt synthesis and homeostasis. In Fxr(-/-) mice we additionally investigated to what extent alterations in bile production during EFA deficiency were FXR dependent. Furthermore, we tested in differentiating Caco-2 cells the effects of EFA deficiency on expression of FXR-target genes relevant for feedback regulation of bile salt synthesis. EFA deficiency-enhanced bile flow and biliary bile salt secretion were associated with elevated bile salt pool size and synthesis rate (+146 and +42%, respectively, P < 0.05), despite increased ileal bile salt reabsorption (+228%, P < 0.05). Cyp7a1 mRNA expression was unaffected in EFA-deficient mice. However, ileal mRNA expression of Fgf15 (inhibitor of bile salt synthesis) was significantly reduced, in agreement with absent inhibition of the hepatic bile salt synthesis. Bile flow and biliary secretion were enhanced to the same extent in EFA-deficient wild-type and Fxr(-/-) mice, indicating contribution of other factors besides FXR in regulation of EHC during EFA deficiency. In vitro experiments show reduced induction of mRNA expression of relevant genes upon chenodeoxycholic acid and a selective FXR agonist GW4064 stimulation in EFA-deficient Caco-2 cells. In conclusion, our data indicate that EFA deficiency is associated with interrupted negative feedback of bile salt synthesis, possibly because of reduced ileal Fgf15 expression.

Essential fatty acid deficiency in mice impairs lactose digestion.

Essential fatty acid (EFA) deficiency in mice induces fat malabsorption. We previously reported indications that the underlying mechanism is located at the level of the intestinal mucosa. We have investigated the effects of EFA deficiency on small intestinal morphology and function. Mice were fed an EFA-deficient or control diet for 8 wk. A 72-h fat balance, the EFA status, and small intestinal histology were determined. Carbohydrate absorptive and digestive capacities were assessed by stable isotope methodology after administration of [U-(13)C]glucose and [1-(13)C]lactose. The mRNA expression and enzyme activity of lactase, and concentrations of the EFA linoleic acid (LA) were measured in small intestinal mucosa. Mice fed the EFA-deficient diet were markedly EFA-deficient with a profound fat malabsorption. EFA deficiency did not affect the histology or proliferative capacity of the small intestine. Blood [13C6]glucose appearance and disappearance were similar in both groups, indicating unaffected monosaccharide absorption. In contrast, blood appearance of [13C]glucose, originating from [1-(13)C]lactose, was delayed in EFA-deficient mice. EFA deficiency profoundly reduced lactase activity (-58%, P<0.01) and mRNA expression (-55%, P<0.01) in mid-small intestine. Both lactase activity and its mRNA expression strongly correlated with mucosal LA concentrations (r=0.77 and 0.79, respectively, P<0.01). EFA deficiency in mice inhibits the capacity to digest lactose but does not affect small intestinal histology. These data underscore the observation that EFA deficiency functionally impairs the small intestine, which in part may be mediated by low LA levels in the enterocytes.

Intestinal capacity to digest and absorb carbohydrates is maintained in a rat model of cholestasis.

Cholestasis is associated with systemic accumulation of bile salts and with deficiency of bile in the intestinal lumen. During the past years bile salts have been identified as signaling molecules that regulate lipid, glucose, and energy metabolism. Bile salts have also been shown to activate signaling routes leading to proliferation, apoptosis, or differentiation. It is unclear, however, whether cholestasis affects the constitution and absorptive capacity of the intestinal epithelium in vivo. We studied small intestinal morphology, proliferation, apoptosis, expression of intestine-specific genes, and carbohydrate absorption in cholestatic (1 wk bile duct ligation), bile-deficient (1 wk bile diversion), and control (sham) rats. Absorptive capacity was assessed by determination of plasma [(2)H]- and [(13)C]glucose concentrations after intraduodenal administration of [(2)H]glucose and naturally enriched [(13)C]sucrose, respectively. Small intestinal morphology, proliferation, apoptosis, and gene expression of intestinal transcription factors (mRNA levels of Cdx-2, Gata-4, and Hnf-1alpha, and Cdx-2 protein levels) were similar in cholestatic, bile-deficient, and control rats. The (unlabeled) blood glucose response after intraduodenal administration was delayed in cholestatic animals, but the absorption over 180 min was quantitatively similar between the groups. Plasma concentrations of [(2)H]glucose and [(13)C]glucose peaked to similar extents in all groups within 7.5 and 30 min, respectively. Absorption of [(2)H]glucose and [(13)C]glucose in plasma was similar in all groups. The present data indicate that neither accumulation of bile salts in the body, nor their intestinal deficiency, two characteristic features of cholestasis, affect rat small intestinal proliferation, differentiation, apoptosis, or its capacity to digest and absorb carbohydrates.

Nutrition for children with cholestatic liver disease.

Cholestatic liver disease (CLD) in children negatively affects nutritional status, growth and development, which all lead to an increased risk of morbidity and mortality. This is illustrated by the fact that the clinical outcome of children with CLD awaiting a liver transplantation is in part predicted by their nutritional status, which is integrated in the pediatric end-stage liver disease model. Preservation of the nutritional status becomes more relevant as the number of patients waiting for liver transplantation increases and the waiting time for a donor organ becomes prolonged. Nutritional strategies are available to optimize feeding of children with CLD. Patients with CLD, however, form a heterogeneous group and the clinical manifestations of their disease vary. This makes a tailor-made approach for these children crucial. Not all aspects of nutrient metabolism and absorption in children with CLD are well understood and studied. Experiments with stable isotope-labeled triglycerides and fatty acids have provided essential information about fat absorption under physiological and cholestatic conditions in animal models and humans. We expect that in the future, tests using other isotope-labeled macronutrients, i.e. carbohydrates and proteins, can be used to further assess nutritional status of children with CLD, thereby creating tailor-made nutritional therapies.

Stem cell factor synergistically enhances thrombopoietin-induced STAT5 signaling in megakaryocyte progenitors through JAK2 and Src kinase.

Stem cell factor (SCF) has a potent synergistic effect during megakaryopoiesis when administered in combination with the major megakaryocytic cytokine, thrombopoietin (TPO). In this study we analyzed the underlying mechanisms with regard to STAT5 activity. TPO stimulation of MO7e cells resulted in STAT5 transactivation, which could be enhanced 1.6-fold by costimulation with SCF, whereas SCF alone did not induce STAT5 transcriptional activity. This costimulatory effect of SCF was reflected in an increase in TPO-induced STAT5 DNA binding and increased and prolonged STAT5 tyrosine phosphorylation in both MO7e cells and primary human megakaryocyte progenitors. In contrast, serine phosphorylation of STAT5 was constitutive and associated with an inhibitory effect on STAT5 transactivation. Signal transduction pathways that might synergize in TPO-mediated STAT5 transactivation were analyzed using specific pharmacological inhibitors and indicated an essential role for Janus-activated kinase 2 (JAK2) and a partial role for Src-family kinases. Costimulation with SCF was found to increase and prolong tyrosine phosphorylation of JAK2 and the TPO receptor c-mpl. In addition, the Src kinase inhibitor SU6656 partially downregulated the additional effect of SCF costimulation on STAT5 tyrosine phosphorylation. SCF-induced enhancement of JAK2 phosphorylation was not affected by inhibition of Src kinase, suggesting that both JAK2 and Src kinase mediate STAT5 tyrosine phosphorylation. Synergistic activation of JAK2 and Src kinase may thus contribute to the enhanced STAT5 signaling in the presence of TPO and SCF.

Comparison of various primer sets for detection of Toxoplasma gondii by polymerase chain reaction in fetal tissues from naturally aborted foxes.

Tissues from 4 aborted polar foxes (3 samples of brain and 4 samples of liver) were selected for Toxoplasma gondii PCR assay. Positive results of serological tests of mothers and immunofluorescence test (IFT) of fetal organ smears were the criteria of sample selection. Five sets of primers designed from B1 gene and ITS1 sequences of T. gondii were used for detection of the parasite in fetal fox tissues. All used primer sets successfully amplified T. gondii DNA in PCR from organs which were positive by IFT. Single tube nested PCR also showed positive result from a sample negative by IFT, but this product was not confirmed. The studies showed usefullness of PCR for routine diagnosis of toxoplasmosis in carnivores.

Phylogenetic similarity of the canine parvovirus wild-type isolates on the basis of VP1/VP2 gene fragment sequence analysis.

Biological material was taken from dogs with diarrhoea. Faecal samples were taken from within live animals and intestinal tract fragments (i.e. small intestine, and stomach) were taken from dead animals. In total, 18 specimens were investigated from dogs housed alone or in large groups. To test for the presence of the virus, latex (On Site Biotech, Uppsala, Sweden) and direct immunofluorescence tests were performed. At the same time, polymerase chain reaction (PCR) with primers complementary to a conservative region of VP1/VP2 was carried out. The products of amplification were analysed on 2% agarose gel. The purified products were cloned with the Template Generation System (Finnzymes, Espoo, Finland) using a transposition reaction and positive clones were searched using the 'colony screening by PCR' method. The sequencing gave 12 sequences of VP1/VP2 gene fragments that were of high similarity. Among the 12 analysed sequences, six exhibited 88% similarity, four exhibited 100% similarity and two exhibited 71% similarity.

Prognostic value of proliferating cell nuclear antigen in Wilms' tumour in children.

AIMS: To evaluate the prognostic value of index Proliferating Cell Nuclear Antigen (PCNA) in Wilms' tumour in children. METHODS: The study comprised 64 children aged from 2 days to 13 years treated according to the SIOP (Society International of Oncology Paediatric) and accepted by the PPGGL (Polish Paediatric Group for the Treatment of Solid Tumours). The studies were conducted on tumour tissue removed during surgery, fixed in formalin and embedded in paraffin blocks. Sections (4 microns) were evaluated by immunohistochemistry, using the peroxidase method to determine the expression of PCNA in Wilms' tumour cells by primary monoclonal antibody NCL-PCNA from Novocastra. RESULTS: The percentage of immunopositive cells in particular fragments of the tumour ranged from 0--93%, mean 30.5%, median 25.5%. Mean and median values enabled division of children into two groups: Group A, where the percentage of cells staining with anti-PCNA was <30% and Group B, where this percentage was >30%. The expression of PCNA was evaluated in various stages of advancement, various histological types and depending on the course of disease. The studies revealed the correlation between index PCNA and stage of advancement P<0.01, index PCNA and histological type of Wilms' tumour P<0.025. Moreover we observed that deaths were found more frequently in tumours with index PCNA >30%, P<0.001. CONCLUSIONS: PCNA is a useful prognostic factor in Wilms' tumour in children.

Toxoplasma gondii infection of cats in epizootiological and clinical aspects.

A total of 200 cats from a south-western region of Poland were tested for antibodies to T. gondii. 105 (52.5%) examined cats were seropositive for T. gondii. Positive results were found in 16 (50%) out of 30 homeless cats, 33 (55%) out of 60 cats kept in shelters, and 56 (51.85%) out of 108 pet cats visiting outpatients clinics. 70 of pet cats stayed indoor and never left house, the others (38 cats) lived at house, but frequently abided outdoor. The seroprevalence in the last group was higher than in cats kept indoor, in shelters or homeless cats, however the differences were not statistically significant. Among the cats kept exclusively indoor the statistically significant difference in seropositivity was noted between the cats fed raw meat (69.23%) and the ones eating only commercial feed or cooked meat (19.35%), chi2 = 17.24, P < 001. Males were more frequently infected than females, but the differences were not statistically significant. In cats over 5 years old the percentage of positive results was significantly higher than in the younger ones. The ranges of antibody titres measured by LAT (latex agglutination test) were in agreement with ranges estimated by IFAT (indirect fluorescent antibody test). The LAT titres over 1980 IU/ml and IF IgG titres over 1:4000 were found only in cats, in which clinical toxoplasmosis was confirmed.

The influence of feeding and maintenance system on occurrence of Toxoplasma gondii infections in dogs.

Serum samples of 113 dogs visiting "outpatient clinics", 52 dogs kept in shelters and 35 animals from a military dog training centre were examined for Toxoplasma gondii specific antibodies using a latex agglutination test. Significant differences in seroprevalences were found between dogs from the training centre (8.6% of positive results) and the other populations examined (40.7% of positive seroreagents in animals visiting outpatient clinics and 44.2% in the group from shelters, respectively). Among clinic patients, dogs fed raw meat were significantly more frequently seropositive (65.2%) than those eating only commercial dry feed or cooked meat (25.7%). No statistically significant differences were noted in males compared to females and in pure breed dogs compared to crossbreed dogs. The antibodies were usually found in low titres under 60 IU/ml (69.6% of positive results). High titres (120-480 IU/ml) were detected in 2 of 3 dogs with clinical toxoplasmosis. In these dogs IFAT T. gondii specific IgM were detected and a favourable response to antiprotozoal treatment was observed. All the dogs with medium and high titres were given raw meat. Age and the presence of cats did not seem to have any influence on T. gondii seroprevalence. Neospora caninum specific antibodies in low titres ranging from 1:20-1:320 were found in 7 (9.7%) of 72 T. gondii positive seroreagents.

Prognostic value of p53 expression in Wilms' tumor in children.

BACKGROUND: The aim of this report was to evaluate the prognostic value and clinical correlations of p53 expression in children with Wilms' tumor. MATERIAL AND METHODS: The study comprised 61 children aged from 2 days to 13 years (median 39 months), diagnosed and treated according to SIOP and PPGGL criteria in three centers co-operating with the PPGGL. The studies were conducted on tumor tissue removed during surgery, fixed in formalin and embedded in paraffin blocks. Then 4-micron sections were evaluated by immunohistochemistry, using the peroxidase method to determine the expression of p53 in Wilms' tumor cells by means of primary monoclonal antibody NCL-p53 from Novocastra. RESULTS: The percentage of immunopositive cells in particular fragments of the tumor ranged from 0% to 70% (mean 20.4%, median 16.0%). The mean and median values enabled the children to be divided into two groups: Group A, where the percentage of cells staining with anti-p53 antibody was >20% (23 cases), and Group B, where this percentage did not exceed 20%. The expression of p53 was then evaluated in various stages of advancement and various histological types, depending on the course of the disease. In Group A, tumors at higher stages of advancement stages were more frequent (p<0.05), and showed a higher degree of malignancy (p<0.06; EFS=56.53%). In Group B, lower stages of advancement were more frequent (p<0.05), the degree of malignancy was lower, and the EFS was 81.58%. A discrimination test, however, showed that the determination of p53 expression in Wilms' tumor cells has moderate sensitivity (58.825%), positive prediction (43.47%), and relatively high specificity (70.45%) and negative prediction (81.57%), which means that low indexes of p53 expression have higher prognostic value. CONCLUSIONS: The index of p53 expression is not an independent prognostic factor in Wilms' tumor in children, but this determination may be helpful in identifying high-risk and low-risk patients.

Spontaneous apoptosis of Reed-Sternberg and Hodgkin cells; clinical and pathological implications in patients with Hodgkin's disease.

The experiments were designed to study correlation between frequency of apoptosis of Reed-Sternberg/Hodgkin (R-S/H) cells, EBV infection of these cells, expression of the key proteins involved in regulation of apoptosis and cell cycle in R-S/H cells, the patients' pretreatment markers and the clinical outcome. One hundred and ten Hodgkin's disease (HD) patients were studies, of which 69 obtained complete remission (CR) after first-line treatment and 41 did not respond. The time of follow-up was from 18 to 242, median 69.7, months. Apoptosis was evaluated by TUNEL technique (TdT-mediated dUTP nick end labeling) and the presence of EBV-latent membrane protein 1 as well as expression of Bcl-2, tumor suppressor p53, p21WAF1, MDM-2, Rb1, PCNA, p27KIP1 and caspase-3, was detected immunocytochemically on paraffin-embedded lymph node specimens obtained at diagnosis. Positive TUNEL reaction was found in 43 patients with apoptotic index (AI) in this group varying between 10% and 60%. In the remaining 57 patients AI of R-S/H cells was below 10%. In 62 patients the cells surrounding R-S/H cells were also TUNEL-positive; their frequency was variable. The expression of LMP1 protein on R-S/H cells was found in 38 patients, without any correlation with the presence or frequency of apoptosis. No significant difference was seen between the AI and both clinical stage and histological type of the disease. However, the mean AI in non-responding patients was significantly higher than in CR group (p=0.015); the high frequency of apoptosis was also negatively correlated with the progression free survival time (p=0.031) and the overall survival (p=0.042). The expression of PCNA, p21WAF1, p53 protein and caspase-3 also showed positive correlation with frequency of apoptosis (p=0.011, p=0.036, and p=0.001, respectively). On the other hand, no statistically confirmed correlation was found between AI and expression of bcl-2, MDM-2, Rb1, and p27KIP1 on R-S/H cells. These data provide evidence that tumor cells in HD undergo spontaneous apoptosis regardless of EBV infection. High pretreatment AI correlates with poor response to the treatment, and may be considered as a potential negative prognostic factor in HD.

Hepatitis C virus infection in hospitalized children.

AIM: The aim of the study was the estimation of the incidence of HCV infection and the analysis of risk factors for the infection in hospitalized children. MATERIAL AND METHODS: Retrospective analysis of the data of 1263 patients treated in hospital wards of Chair and Department of Gastroenterology and Paediatric Diseases from July 1995 to March 1998; all the patients had hepatitis C virus antibodies determined. Hepatitis C virus antibodies were assessed with the use of enzyme immunoassay. All the positive results were confirmed by means of Lia Tek HCV or PCR HCV RNA method. History data regarding frequency of previous hospitalizations, operations, blood transfusions and invasive diagnostic procedures patients had undergone were included in analysis. RESULTS: HCV infection was diagnosed in 47 children which accounted for 3.7% of the group enrolled in the study. The majority, that is 96.2% of seropositive children had the history of hospitalization, while 73.2% of them underwent intervention procedures disrupting tissue continuity during previous hospitalizations. Physical examination analysis indicates that all the patients with antibodies against HCV could have acquired the infection through parenteral transmission. The data indicating another route of transmission were not obtained (e.g. familial transmission). CONCLUSIONS: 1. Serological tests for HCV infection should be performed on routine basis in frequently hospitalized children. 2. HCV infection resulting from iatrogenic transmission can be suspected in the majority of seropositive children.

[Chronic hepatitis C in a 12-year-old girl coinfected with HGV]. / Przewlekle zapalenie watroby typu C wspólistniejace z zakazeniem HGV u 12-letniej dziewczynki.

In this study we present a 12-year-old girl with chronic C hepatitis coinfected with HGV, in which severe life-threatening side effects from alfa interferon therapy occurred after 3 months of injection and required definite IFN withdrawal. It seems, that infection HGV may predispose patients with chronic C hepatitis treated with alpha interferon to this severe side effect.