Ubiquitin-dependent protein processing controls radiation-induced apoptosis through the N-end rule pathway

The ubiquitination of nuclear proteins activated in human lymphocytes undergoing radiation-induced apoptosis and the subsequent downstream proteasomal protein processing, shown to be involved in apoptotic death control, may be dependent on an amino-terminal sequence identity of ubiquitin target proteins, the "N-end rule" pathway. Here we report that this selective pathway controls radiation-induced apoptosis and that it is involved in the initiation of this type of cell death. Dipeptide competitors of protein ubiquitination/processing dependent solely on the basic amino-terminal residues (type I) efficiently inhibited the radiation-induced apoptotic death phenotype, indicating that only the substrates of ubiquitination with basic NH2-terminal amino acids are involved in apoptotic death control. This selective inhibition was followed by an early, overall but also target-specific inhibition of ubiquitination and by an activation and stabilization of poly(ADP-ribose) polymerase (PARP) that occurs through inhibition of ubiquitination of its cleaved form (85 kDa). Interestingly, caspases-3 and -7 were not activated following irradiation, further suggesting that PARP cleavage may be regulated by an N-end rule pathway in a caspase-independent manner. These results highly suggest involvement of this subset of the ubiquitin system in the apoptotic death control and in the specific regulation of PARP activity.     

Nucleotides affect neurogenesis and dopaminergic differentiation of mouse fetal midbrain-derived neural precursor cells

The fetal midbrain is a preferred source for isolating and producing dopaminergic neurons for subsequent grafting and replacement of damaged or lost dopaminergic midbrain neurons. We analysed the potential of a variety of nucleotides and of adenosine to support dopaminergic neuron formation from primary mouse fetal midbrain-derived cells, harvested at E10.5 and at E13.5 and subjected to adherent cell culture. In contrast to cells derived at E13.5, cells derived at E10.5 have the potential to produce dopaminergic neurons in culture. These neurons express tyrosine hydroxylase and the dopamine transporter. The fetal ventral midbrain contained mRNA encoding almost all P2X and P2Y receptors, all adenosine receptors as well as the ectonucleotidases nucleoside triphosphate diphosphohydrolase 2 and tissue nonspecific alkaline phosphatase. Essentially, all components of the purinergic signalling pathway were also expressed by the cultured cells. ATP, ADPβS, 2MeSATP, 2ClATP and adenosine increased neuron formation. There was, however, no preference for the formation of dopaminergic neurons—with the exception of 2ClATP that increased the relative contribution of tyrosine hydroxylase-positive neurons. In cells isolated at E13.5 UTP promoted neuron survival but ADPβS and ATPγS essentially eliminated neurons. These data showed that the outcome of nucleotide application was different even though cells isolated at E10.5 and E13.5 expressed very similar receptor mRNA profiles. They suggest that purinergic agonists carry potential for stimulating neurogenesis and enriching the contribution of dopaminergic neurons in vitro. Nucleotide receptor agonists may be of value for contributing to the formation and survival of dopaminergic neurons in vivo.     

Supracondylar fractures of the humerus in children caused by traffic

A sample of 38 patients aged under 14 treated at the Surgery Clinic of the University Hospital Osijek has been used in showing therapeutic guidelines supported by patients' functional status after the completed treatment. The patients suffered from supracondylar fractures caused by traffic accidents and classified into three types according to Gartland. Not a single patient with neurocirculatory disturbances has been registered. The most common treatment was a conservative one while more than half of the patients with fractures from type III underwent surgery. The duration of immobilization approximated three weeks whereas patients' recovery considering their functional status has justified the choice of the treatment. However, a lot can be done in the field of prevention, which would greatly decrease the frequency of such injuries.     

Intercostal catheter analgesia is more efficient vs. intercostal nerve blockade for post-thoracotomy pain relief

A pain after thoracotomy may result in a postoperative hypoventilation and lead to atelectases and pneumonia. This study was aimed to compare two analgesic regimens after posterolateral thoracotomy. 80 patients (40-70 years) undergoing thoracotomy were randomized to intercostal catheter analgesia (group A, n = 40) and intercostal nerve block (group B, n = 40). Patients in group A were given 20 mL of 0.5% bupivacaine injections twice a day by intercostal catheter. Intercostal nerve blockade was performed using 5 mL of 0.5% bupivacaine. Intercostal nerve in thoracotomy wound, nerves below and above thoracotomy wound was also injected. PaO2, PaCO2, FVC, FEV1 and visual analog pain scale (VAS) were obtained preoperatively, 24, 48 and 72 hours after operation. Postoperative complications were recorded at the patient discharge. Differences between groups were calculated using Mann-Whitney, KW test and chi square test. The arterial blood gas analyses did not show statistically significant change in any group and time according to the baseline values. FVC and FEV1 decreased significantly in both groups at first postoperative day according to baseline measurements. Patients in B group had significantly higher FEV1 values in the third postoperative day (73.05 +/- 11.25 in A vs. 83.50 +/- 9.17 in B group, p < 0.05). Intercostal catheter analgesia resulted in significantly lower postoperative VAS scores and reduced opioid requirement as compared to intercostal nerve blockade. No differences in the postoperative complications were observed between groups.     

Trichinella spiralis and breast carcinoma--a case report

Authors report about a patient with recurrent ductal invasive breast carcinoma and trichinosis. The patient underwent mastectomy of the left breast with evacuation of the axilla because of the cancer. Radiation therapy was received. An infestation with Trichinella spiralis was diagnosed two years after The patient was treated with mebendazole. A local recurrence of the tumor was found on the chest wall six years after the surgery. Tumor excision was performed. Histological analysis pointed at a ductal invasive carcinoma with numerous parasites of Trichinella spiralis present within both the muscle and the tumor tissue. The finding of parasites in the tumor tissue witnesses in favor of infestation, and the parasite morphology preserved in the tumor shows at the protective effects of the cysts, i.e. preventing parasite necrosis.     

Environmental Heterogeneity Explains the Genetic Structure of Continental and Mediterranean Populations of Fraxinus angustifolia Vahl

Tree species with wide distributions often exhibit different levels of genetic structuring correlated to their environment. However, understanding how environmental heterogeneity influences genetic variation is difficult because the effects of gene flow, drift and selection are confounded. We investigated the genetic variation and its ecological correlates in a wind-pollinated Mediterranean tree species, Fraxinus angustifolia Vahl, within a recognised glacial refugium in Croatia. We sampled 11 populations from environmentally divergent habitats within the Continental and Mediterranean biogeographical regions. We combined genetic data analyses based on nuclear microsatellite loci, multivariate statistics on environmental data and ecological niche modelling (ENM). We identified a geographic structure with a high genetic diversity and low differentiation in the Continental region, which contrasted with the significantly lower genetic diversity and higher population divergence in the Mediterranean region. The positive and significant correlation between environmental and genetic distances after controlling for geographic distance suggests an important influence of ecological divergence of the sites in shaping genetic variation. The ENM provided support for niche differentiation between the populations from the Continental and Mediterranean regions, suggesting that contemporary populations may represent two divergent ecotypes. Ecotype differentiation was also supported by multivariate environmental and genetic distance analyses. Our results suggest that despite extensive gene flow in continental areas, long-term stability of heterogeneous environments have likely promoted genetic divergence of ashes in this region and can explain the present-day genetic variation patterns of these ancient populations.     

Intact radial and median nerve after open third degree distal fracture of the humerus

A 54 year old man sustained a third degree open fracture at the distal part of the right humerus with massive soft tissue defect involving most of the upper arm. The radial and median nerves were completely bared and exposed by 6 cm for radial and 3 cm for median nerve. The nerves were in continuity, but there was complete rupture of surrounding muscles: biceps, triceps and brachialis. The fracture was stabilized by external fixation method--reinforced by wires. Preoperative and postoperative sensorimotor status of the right hand was good. One year later sensory and motoric status of right hand showed no deficiencies, but flexion and extension in elbow were limited to 100 and 180 degrees respectively. Pronosupination was restricted. This case report is consistent with results of biomechanical studies in vitro confirming high tolerance of radial and median nerve to stretching injury.     

Vegetation of trampled habitats in the Plitvice Lakes National Park in Croatia

Abstract

The vegetation communities of the trampled habitats in the Plitvice Lakes National Park were studied during 2005 and 2006. With the numerical classification of the relevés, six clusters were obtained, corresponding to the following specific communities: cluster 1 – ass. Matricario–Polygonetum arenastri– the intensively trampled and grazed habitats; cluster 2 – ass. Matricario–Polygonetum arenastri– the only intensively trampled habitats; cluster 3 – ass. Coronopodo–Polygonetum arenastri– very intensively trampled and grazed farmyards; cluster 4 –Trifolium fragiferum community – the trampled surfaces of moist habitats; cluster 5 – ass. Lolietum perennis– the moderately trampled surfaces of roads and yards; cluster 6 – ass. Prunello–Ranunculetum repentis– trampled surfaces of forest roads. The first three clusters belong to the vegetation of intensively trampled habitats of the class Polygono arenastri–Poetea annuae and the last three clusters belong to moderately trampled habitats of the class Molinio–Arrhenatheretea. In the analysis of the ecological factors it is established that separation of the relevés is influenced most greatly by the trampling intensity observed in the field, and of the Ellenberg indicator values, the most important are light, moisture, soil reaction and temperature. Using the analysis of the plant functional traits it is established that therophytes, annuals, biennials, species resistant to trampling and grazing, neophytes and archeophytes prevail in the intensively trampled habitats. In contrast, hemicryptophytes, perennials, and indigenous plant species prevail in the moderately trampled habitats. Representation of the C-S-R plant functional types shows also that trampling as a disturbance factor has a large influence on the differences in the relevés. In the intensively trampled habitats, there is a large proportion of ruderals, while in the moderately trampled habitats there is a large proportion of competitors.     

Candidate gene SNP variation in floodplain populations of pedunculate oak (Quercus robur L.) near the species' southern range margin: Weak differentiation yet distinct associations with water availability

Populations residing near species' low‐latitude range margins (LLMs) often occur in warmer and drier environments than those in the core range. Thus, their genetic composition could be shaped by climatic drivers that differ from those occurring at higher latitudes, resulting in potentially adaptive variants of conservation value. Such variants could facilitate the adaptation of populations from other portions of the geographical range to similar future conditions anticipated under ongoing climate change. However, very few studies have assessed standing genetic variation at potentially adaptive loci in natural LLM populations. We investigated standing genetic variation at single nucleotide polymorphisms (SNPs) located within 117 candidate genes and its links to putative climatic selection pressures across 19 pedunculate oak (Quercus robur L.) populations distributed along a regional climatic gradient near the species' southern range margin in southeastern Europe. These populations are restricted to floodplain forests along large lowland rivers, whose hydric regime is undergoing significant shifts under modern rapid climate change. The populations showed very weak geographical structure, suggesting extensive genetic connectivity and gene flow or shared ancestry. We identified eight (6.2%) positive F_ST‐outlier loci, and genotype–environment association analyses revealed consistent associations between SNP allele frequencies and several climatic variables linked to water availability. A total of 61 associations involving 37 SNPs (28.5%) from 35 annotated genes provided important insights into putative functional mechanisms in our system. Our findings provide empirical support for the role of LLM populations as sources of potentially adaptive variation that could enhance species’ resilience to climate change‐related pressures.     

Natural phosphorylation of CD5 in chronic lymphocytic leukemia B cells and analysis of CD5-regulated genes in a B cell line suggest a role for CD5 in malignant phenotype

Chronic lymphocytic leukemia (CLL) results in the accumulation of B cells, presumably reflecting the selection of malignant cell precursors with Ag combined with complex alterations in protein activity. Repeated BCR stimulation of normal B cells leads to anergy and CD5 expression, both of which are features of CLL. Because CD5 is phosphorylated on tyrosine following BCR engagement and negatively regulates BCR signaling in normal B cells, we investigated its phosphorylation status and found it to be naturally phosphorylated on tyrosine but not on serine residues in CLL samples. To analyze the role of CD5, we established a B cell line in which CD5 is phosphorylated. Gene profiling of vector vs CD5-transfected B cells pointed out gene groups whose expression was enhanced: Apoptosis inhibitors (BCL2), NF-kappaB (RELB, BCL3), Wnt, TGFbeta, VEGF, MAPKs, Stats, cytokines, chemokines (IL-10, IL-10R, IL-2R, CCL-3, CCL-4, and CCR7), TLR-9, and the surface Ags CD52, CD54, CD70, and CD72. Most of these gene groups are strongly expressed in CLL B cells as compared with normal B cells. Unexpectedly, metabolic pathways, namely cholesterol synthesis and adipogenesis, are also enhanced by CD5. Conversely, CD5 inhibited genes involved in RNA splicing and processing, ribosome biogenesis, proteasome, and CD80 and CD86 Ags, whose expression is low in CLL. Comparison of CD5- vs tailless CD5-transfected cells further demonstrated the role of CD5 phosphorylation in the regulation of selected genes. These results support a model where CLL cells are chronically stimulated, leading to CD5 activation and cell survival. In addition to CD5 itself, we point to several CD5-induced genes as potential therapeutic targets.