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International audience; The growing availability of novel interpretation techniques opened the way to the application of deep learning models in the clinical field, including neuroimaging, where their use is still largely underexploited. In this framework, we focus the stratification of Multiple Sclerosis (MS) patients in the Primary Progressive versus the Relapsing-Remitting state of the disease using a 3D Convolutional Neural Network trained on structural MRI data. Within this task, the application of Layer-wise Relevance Propagation visualization allowed detecting the voxels of the input data mostly involved in the classification decision, potentially bringing to light brain regions which might reveal disease state.

Xenorhabdus hominickii ANU1 is known to be an entomopathogenic bacterium symbiotic to nematode Steinernema monticolum. Another bacterial strain X. hominickii DY1 was isolated from a local population of S. monticolum. This bacterial strain X. hominickii DY1 was found to exhibit high insecticidal activities against lepidopteran and coleopteran species after hemocoelic injection. However, these two X. hominickii strains exhibited significant variations in insecticidal activities, with ANU1 strain being more potent than DY1 strain. To clarify their virulence difference, bacterial culture broths of these two strains were compared for secondary metabolite compositions. GC-MS analysis revealed that these two strains had different compositions, including pyrrolopyrazines, piperazines, cyclopeptides, and indoles. Some of these compounds exhibited inhibitory activities against phospholipase A2 to block eicosanoid biosynthesis and induce significant immunosuppression. They also exhibited significant insecticidal activities after oral feeding, with indole derivatives being the most potent. More kinds of indole derivatives were detected in the culture broth of ANU1 strain. To investigate variations in regulation of secondary metabolite production, expression level of leucine-responsive regulatory protein (Lrp), a global transcription factor, was compared. ANU1 strain exhibited significantly lower Lrp expression level than DY1 strain. To assess genetic variations associated with secondary metabolite synthesis, bacterial loci encoding non-ribosomal protein synthase and polyketide synthase (NRPS-PKS) were compared. Three NRPS and four PKS loci were predicted from the genome of X. hominickii. The two bacterial strains exhibited genetic variations (0.12∼0.67%) in amino acid sequences of these NRPS-PKS. Most NRPS-PKS genes exhibited high expression peaks at stationary phase of bacterial growth. However, their expression levels were significantly different between the two strains. These results suggest that differential virulence of the two bacterial strains is caused by the difference in Lrp expression level, leading to difference in the production of indole compounds and other NRPS-PKS-associated secondary metabolites.

A critical issue in executive control is how the nervous system exerts flexibility to inhibit a prepotent response and adapt to sudden changes in the environment. In this study, force measurement was used to capture “partial” unsuccessful trials that are highly relevant in extending the current understanding of motor inhibition processing. Moreover, a modified version of the stop-signal task was used to control and eliminate potential attentional capture effects from the motor inhibition index. The results illustrate that the non-canceled force and force rate increased as a function of stop-signal delay (SSD), offering new objective indices for gauging the dynamic inhibitory process. Motor response (time and force) was a function of delay in the presentation of novel/infrequent stimuli. A larger lateralized readiness potential (LRP) amplitude in go and novel stimuli indicated an influence of the novel stimuli on central motor processing. Moreover, an early N1 component reflects an index of motor inhibition in addition to the N2 component reported in previous studies. Source analysis revealed that the activation of N2 originated from inhibitory control associated areas: the right inferior frontal gyrus (rIFG), pre-motor cortex, and primary motor cortex. Regarding partial responses, LRP and error-related negativity (ERNs) were associated with error correction processes, whereas the N2 component may indicate the functional overlap between inhibition and error correction. In sum, the present study has developed reliable and objective indices of motor inhibition by introducing force, force-rate and electrophysiological measures, further elucidating our understandings of dynamic motor inhibition and error correction.

Aim: To investigate the relationship between PI3K/Akt/NF-κB cellular signal pathway and the expression of P-gp and LRP in multidrug resistance (MDR) cell of nasopharyngeal carcinoma. Method: The PI3K, p-Akt and NF-κB/p65 as the activity of PI3K/Akt/NF-κB were detected by Western blot. The expressions of LRP and P-gp were detected by Western blot and real-time PCR. Result: The RIs of CNE/DDP group to DDP, 5-Fu, VCR, ADR and PTX were 35.04, 18.14, 24.13, 12.00 and 10.18, respectively. The RIs of LY-294002 group were 11.77, 5.83, 3.07, 3.86 and 3.34, and PDTC group were 11.08, 6.55, 7.66, 2.18 and 4.05. The expressions of PI3K, p-Akt and NF-κBp65, LRP and P-gp were increased and mRNA of LRP and P-gp were up-regulated in CNE/DDP. The expression of p-Akt in LY-294002 group was down-regulated. The expression of NF-κB p65 in PDTC group was decreased. The mRNA of LRP and P-gp in LY-294002 group and PDTC group were decreased. Conclusion: MDR of nasopharyngeal carcinoma cell can be regulated by activating PI3K/Akt/NF-κB signal pathway and then increase the expression of P-gp and LRP. The MDR of nasopharyngeal carcinoma cell can be reversed by inhibiting PI3K/Akt/NF-κB signal pathway.

Background An attenuated mutant (designated NY303) of Vibrio vulnificus, which causes serious wound infection and septicemia in humans, was isolated fortuitously from a clinical strain YJ016. This mutant was defective in cytotoxicity, migration on soft agar and virulence in the mouse. The purpose of this study was to map the mutation in this attenuated mutant and further explore how the gene thus identified is involved in virulence. Methods The whole genome sequence of mutant NY303 determined by next-generation sequencing was compared with that of strain YJ016 to map the mutations. By isolating and characterizing the specific gene-knockout mutants, the gene associated with the phenotype of mutant NY303 was identified. This gene encodes a global regulator, Lrp. A mutant, YH01, deficient in Lrp was isolated and examined in vitro, in vivo and ex vivo to find the affected virulence mechanisms. The target genes of Lrp were further identified by comparing the transcriptomes, which were determined by RNA-seq, of strain YJ016 and mutant YH01. The promoters bound by Lrp were identified by genome footprinting-sequencing, and those related with virulence were further examined by electrophoretic mobility shift assay. Results A mutation in lrp was shown to be associated with the reduced cytotoxicity, chemotaxis and virulence of mutant NY303. Mutant YH01 exhibited a phenotype resembling that of mutant NY303, and was defective in colonization in the mouse and growth in mouse serum, but not the antiphagocytosis ability. 596 and 95 genes were down- and up-regulated, respectively, in mutant YH01. Many of the genes involved in secretion of the MARTX cytotoxin, chemotaxis and iron-acquisition were down-regulated in mutant YH01. The lrp gene, which was shown to be negatively autoregulated, and 7 down-regulated virulence-associated genes were bound by Lrp in their promoters. A 14-bp consensus sequence, mkCrTTkwAyTsTG, putatively recognized by Lrp was identified in the promoters of these genes. Conclusions Lrp is a global regulator involved in regulation of cytotoxicity, chemotaxis and iron-acquisition in V. vulnificus. Down-regulation of many of the genes associated with these properties may be responsible, at least partly, for loss of virulence in mutant NY303. Electronic supplementary material The online version of this article (doi:10.1186/s12929-017-0361-9) contains supplementary material, which is available to authorized users.

Tepekoy, Filiz/0000-0003-1901-3787 WOS:000491949700001 PubMed: 31630211 Wide application of gonadotropin-releasing hormone (GnRH) agonists and antagonists for clinical purposes determines their effects on ovarian signaling pathways. Our study aimed to determine the localization, expression levels of Wnt signaling members in the pubertal and adult mouse ovary and the impact of GnRH antagonist cetrorelix on these signaling members. 0.5 mg/kg of cetrorelix was injected to 3-and 6-week-old mice for 2 weeks. At the end of injection, ovaries from 5 (5Ce)- to 8-week (8Ce)-old mice were embedded in paraffin for immunohistochemistry and homogenized for western blot to compare with control (5C-8C) and sham groups (5S-8S). WNT2 and WNT4 showed higher expression in thecal and stromal cells in adult mouse ovaries and only WNT4 expression was affected by cetrorelix. FZD1 was localized mainly in oocytes of pubertal ovaries and granulosa cells and oocytes of adult ovaries. FZD1 was reduced by cetrorelix in pubertal ovaries. FZD4 was abundantly localized in thecal and stromal cells of all groups and protein level was not affected by cetrorelix. LRP-6 was expressed mainly in oocytes and stromal cells of pubertal, oocytes of adult ovaries and its expression was reduced by cetrorelix in adult ovaries. CTNNB1 intensity in granulosa cells was the lowest in pubertal and the highest in adult ovaries and its expression was decreased by cetrorelix in adult ovaries. Cetrorelix affected the expression of specific members of the Wnt signaling depending on the developmental stage of mice, pointing out its possible interaction with gonadotropins during pubertal and adult stages. Scientific Research Projects Coordination Unit of Akdeniz UniversityAkdeniz University [2012.01.0103.003] This study was supported by the Scientific Research Projects Coordination Unit of Akdeniz University (Project Number: 2012.01.0103.003).

Reward-predictive stimuli can increase an automatic response tendency, which needs to be counteracted by effortful response inhibition when this tendency is inappropriate for the current task. Here we investigated how the human brain implements this dynamic process by adopting a reward-modulated Simon task while acquiring EEG and fMRI data in separate sessions. In the Simon task, a lateral target stimulus triggers an automatic response tendency of the spatially corresponding hand, which needs to be overcome if the activated hand is opposite to what the task requires, thereby delaying the response. We associated high or low reward with different targets, the location of which could be congruent or incongruent with the correct response hand. High-reward targets elicited larger Simon effects than low-reward targets, suggesting an increase in the automatic response tendency induced by the stimulus location. This tendency was accompanied by modulations of the lateralized readiness potential over the motor cortex, and was inhibited soon after if the high-reward targets were incongruent with the correct response hand. Moreover, this process was accompanied by enhanced theta oscillations in medial frontal cortex and enhanced activity in a frontobasal ganglia network. With dynamical causal modeling, we further demonstrated that the connection from presupplementary motor area (pre-SMA) to right inferior frontal cortex (rIFC) played a crucial role in modulating the reward-modulated response inhibition. Our results support a dynamic neural model of reward-induced response activation and inhibition, and shed light on the neural communication between reward and cognitive control in generating adaptive behaviors.

Victims of urban violence are at risk of developing Posttraumatic Stress Disorder (PTSD), one of the most debilitating consequences of violence. Considering that PTSD may be associated with inefficient selection of defensive responses, it is important to understand the relation between motor processing and PTSD. The present study aims to investigate the extent to which the severity of posttraumatic stress symptoms (PTSS) is related to motor preparation against visual threat cues in victims of urban violence. Participants performed a choice reaction time task while ignoring a picture that could be threating or neutral. The EEG indices extracted were the motor-related amplitude asymmetry (MRAA) in the alpha frequency range, and the lateralized readiness potential (LRP). We observed a linear relation between longer LRP latency and a slower reaction time, selectively during threat processing (compared to neutral) in low PTSS, but not in high PTSS participants. Alpha MRAA suppression and the PTSS were also linearly related: the smaller the alpha MRAA suppression in the threat condition relative to neutral, the greater the PTSS. These results provide evidence that threatening cues affect motor processing that is modulated by the severity of PTSS in victims of urban violence.

Cells of the host containing special proteins-transporters, so-called proteins of multiple drug resistance (MDR proteins) can contribute to the drug resistance formation. Goal of the study: to define specific expression of gene and distribution of main MDR proteins (MDR1/Pgp, MRP1, lRP, BCRP) in the pulmonary cells in case of active tuberculosis. Materials and methods. Expression of MDR protein genes was evaluated by RT-PCR for mRNA isolated from the surgical specimen of fibrous cavernous tuberculosis patients. Localization of MDR proteins was performed by immunohistochemical staining and confocal laser microscopy. Main results. Intensity of MDR protein genes expression varies in different zones of tuberculous lesions: MDR1 and BCRP are characterized by the highest level and MRP1 gene is characterized by the minimum level. The level of lRP gene expression depends on the inflammation zone and it is maximum in perifocal zone where protein is detected in the cells of alveolar epithelium and macrophages. High expression of MDR protein genes in various parts of tuberculous lesions witnesses about the potential involvement of these proteins in the development of drug resistance to anti-tuberculosis drugs.

Members of the gut microbiota are thought to experience strong competition for nutrients. However, how such competition shapes their evolutionary dynamics and depends on intra- and interspecies interactions is poorly understood. Here, we test the hypothesis that Escherichia coli evolution in the mouse gut is more predictable across hosts in the absence of interspecies competition than in the presence of other microbial species. In support, we observed that lrp, a gene encoding a global regulator of amino acid metabolism, was repeatedly selected in germ-free mice 2 weeks after mono-colonization by this bacterium. We established that this specific genetic adaptation increased E. coli's ability to compete for amino acids, and analysis of gut metabolites identified serine and threonine as the metabolites preferentially consumed by E. coli in the mono-colonized mouse gut. Preference for serine consumption was further supported by testing a set of mutants that showed loss of advantage of an lrp mutant impaired in serine metabolism in vitro and in vivo. Remarkably, the presence of a single additional member of the microbiota, Blautia coccoides, was sufficient to alter the gut metabolome and, consequently, the evolutionary path of E. coli. In this environment, the fitness advantage of the lrp mutant bacteria is lost, and mutations in genes involved in anaerobic respiration were selected instead, recapitulating the eco-evolutionary context from mice with a complex microbiota. Together, these results highlight the metabolic plasticity and evolutionary versatility of E. coli, tailored to the specific ecology it experiences in the gut. info:eu-repo/semantics/publishedVersion

The human pathogen, Vibrio cholerae, belongs to the 10% of bacteria in which the genome is divided. Each of its two chromosomes, like bacterial chromosomes in general, replicates from a unique origin at fixed times in the cell cycle. Chr1 initiates first, and upon duplication of a site in Chr1, crtS, Chr2 replication initiates. Recent in vivo experiments demonstrate that crtS binds the Chr2-specific initiator RctB and promotes its initiator activity by remodeling it. Compared to the well-defined RctB binding sites in the Chr2 origin, crtS is an order of magnitude longer, suggesting that other factors can bind to it. We developed an in vivo screen to identify additional crtS-binding proteins and identified the global transcription factor, Lrp, as one such protein. Studies in vivo and in vitro indicate that Lrp binds to crtS and facilitates RctB binding to crtS. Chr2 replication is severely defective in the absence of Lrp, indicative of a critical role of the transcription factor in licensing Chr2 replication. Since Lrp responds to stresses such as nutrient limitation, its interaction with RctB presumably sensitizes Chr2 replication to the physiological state of the cell.

Objective Dementia with Lewy bodies is an α-synucleinopathy characterized by neocortical Lewy-related pathology (LRP). We carried out a genome-wide association study (GWAS) on neocortical LRP in a population-based sample of subjects aged 85 or over. Methods LRP was analyzed in 304 subjects in the Vantaa 85+ sample from Southern Finland. The GWAS included 41 cases with midbrain, hippocampal, and neocortical LRP and 177 controls without midbrain and hippocampal LRP. The Medical Research Council Cognitive Function and Ageing Study (CFAS) material was used for replication (51 cases and 131 controls). Results By analyzing 327,010 markers the top signal was obtained at the HLA-DPA1/DPB1 locus (P = 1.29 × 10−7); five other loci on chromosomes 15q14, 2p21, 2q31, 18p11, and 5q23 were associated with neocortical LRP at P

The most important causes of treatment failure in canine lymphoma include intrinsic or acquired drug resistance. Thus, elucidation of molecular mechanisms of drug resistance is essential for the establishment of better treatment alternatives for lymphoma patients. The overexpression of drug transporters is one of the most intensively studied mechanisms of drug resistance in many tumors. In canine lymphoma, it has also been shown that the overexpression of drug efflux pumps such as P-glycoprotein is associated with drug-resistant phenotypes. Canine lymphoma has many pathological similarities to human non-Hodgkin's lymphoma, and they also share similar molecular mechanisms of drug resistance. We have previously demonstrated the association of the overexpression of drug transporters with drug resistance and indicated some molecular mechanisms of the regulation of these transporters' expressions in canine and human lymphoid tumor cells. However, it has also been indicated that other known or novel drug resistance factors should be explored to overcome drug resistance in lymphoma. In this review, we summarize the recent findings on the molecular mechanisms of drug resistance and possible strategies to develop better treatment modalities for canine lymphoma from the comparative aspects with human lymphoid tumors.

N𝜀-lysine acetylation is an abundant and important Post-translational modification in bacteria. We used the bacterial two-hybrid system to screen the genome library of the Salmonella Typhimurium to identify potential proteins involved in acetyltransferase Pat – or deacetylase CobB-mediated acetylation. Then, the in vitro (de)acetylation assays were used to validate the potential targets, such as STM14_1074, NrdF, RhaR. Lrp, a leucine-responsive regulatory protein and global regulator, was shown to interact with Pat. We further demonstrate that Lrp could be acetylated by Pat and deacetylated by NAD+-dependent CobB in vitro. Specifically, the conserved lysine residue 36 (K36) in helix-turn-helix (HTH) DNA-binding domain of Lrp was acetylated. Acetylation of K36 impaired the function of Lrp through altering the affinity with the target promoter. The mutation of K36 in chromosome mimicking acetylation enhanced the transcriptional level of itself and attenuated the mRNA levels of Lrp-regulated genes including fimA, which was confirmed by yeast agglutination assay. These findings demonstrate that the acetylation regulates the DNA-binding activity of Lrp, suggesting that acetylation modification of transcription factors is a conserved regulatory manner to modulate gene expression in bacteria and eukaryotes.

Transhydrogenases catalyse interconversion of the redox cofactors NADH and NADPH, thereby conveying metabolic flexibility to balance catabolic NADPH formation with anabolic or stress-based consumption of NADPH. Escherichia coli is one of the very few microbes that possesses two isoforms: the membrane-bound, proton-translocating transhydrogenase PntAB and the cytosolic, energy-independent transhydrogenase UdhA. Despite their physiological relevance, we have only fragmented information on their regulation and the signals coordinating their counteracting activities. Here we investigated PntAB and UdhA regulation by studying transcriptional responses to environmental and genetic perturbations. By testing pntAB and udhA GFP reporter constructs in the background of WT E. coli and 62 transcription factor mutants during growth on different carbon sources, we show distinct transcriptional regulation of the two transhydrogenase promoters. Surprisingly, transhydrogenase regulation was independent of the actual catabolic overproduction or underproduction of NADPH but responded to nutrient levels and growth rate in a fashion that matches the cellular need for the redox cofactors NADPH and/or NADH. Specifically, the identified transcription factors Lrp, ArgP and Crp link transhydrogenase expression to particular amino acids and intracellular concentrations of cAMP. The overall identified set of regulators establishes a primarily biosynthetic role for PntAB and link UdhA to respiration.

A growing body of evidence implies psychological disturbances in amyotrophic lateral sclerosis (ALS). Specifically, executive dysfunctions occur in up to 50% of ALS patients. The recently shown presence of cytoplasmic aggregates (TDP-43) in ALS patients and in patients with behavioral variants of frontotemporal dementia suggests that these two disease entities form the extremes of a spectrum. The present study aimed at investigating behavioral and electrophysiological indices of conflict processing in patients with ALS. A non-verbal variant of the flanker task demanded two-choice responses to target stimuli that were surrounded by flanker stimuli which either primed the correct response or the alternative response (the latter case representing the conflict situation). Behavioral performance, event-related potentials (ERP), and lateralized readiness potentials (LRP) were analyzed in 21 ALS patients and 20 controls. In addition, relations between these measures and executive dysfunctions were examined. ALS patients performed the flanker task normally, indicating preserved conflict processing. In similar vein, ERP and LRP indices of conflict processing did not differ between groups. However, ALS patients showed enhanced posterior negative ERP waveform deflections, possibly indicating increased modulation of visual processing by frontoparietal networks in ALS. We also found that the presence of executive dysfunctions was associated with more error-prone behavior and enhanced LRP amplitudes in ALS patients, pointing to a prefrontal pathogenesis of executive dysfunctions and to a potential link between prefrontal and motor cortical functional dysregulation in ALS, respectively.

Background Multidrug resistance is one of the major reasons chemotherapy-based treatments failed in hepatitis B virus (HBV) related hepatocellular carcinoma (HCC). Hypoxia is generally associated with tumor chemo-resistance. The aim of the study was to investigate the effect of Arsenic trioxide (As2O3) on the hypoxia-induced chemo-resistance to 5-FU or cisplatin and explored its underlying mechanism in the HBx-HepG2 cells. Methods MTT assay was used to examine the cell viability. Mitochondrial membrane potential (MMP) and cell cycle was examined by flow cytometry. qRT-PCR was employed to observe the mRNA expression level; and western blot assay was used to determine the protein expression level. Results Our results showed that transfection of HBx plasmid established the HBx-HepG2 cells expressing HBx, and the expression of HBx was confirmed by qRT-PCR and western blot. Exposure of HBx-HepG2 cells to hypoxia (5 % O2, 3 % O2, 1 % O2) for 48 h increased the chemo-resistance to 5-fluorouracil (5-FU) (50–1600 µM) and cisplatin (25–800 µM), reduced MMP, and caused the cell cycle arrest at G0/G1 phase in a concentration-dependent manner. Hypoxia also concentration-dependently (5 % O2, 3 % O2, 1 % O2) reduced mRNA expression level of P-glycoprotein (P-gp), multidrug resistance protein (MRP1), lung resistance protein (LRP), and decreased the protein expression level of hypoxia-inducible factor-1α (HIF-1α), P-gp MRP1, and LRP. Following pretreatment with As2O3 at a non-cytotoxic concentration re-sensitized the hypoxia (1 % O2)-induced chemo-resistance to 5-FU and cisplatin in HBx-HepG2 cells. As2O3 pretreatment also prevented MMP reduction and G0/G1 arrest induced by hypoxia. Meanwhile, As2O3 antagonized increase of HIF-1α protein induced by hypoxia, and it also suppresses the increase in expression levels of P-gp, MRP1, and LRP mRNA and proteins. In addition, As2O3 in combination with 5-FU treatment caused up-regulation of DR5, caspase 3, caspase 8, and caspase 9, and down-regulation of BCL-2, but had no effect of DR4. Conclusions Our results may suggest that As2O3 re-sensitizes hypoxia-induced chemo-resistance in HBx-HepG2 via complex pathways, and As2O3 may be a potential agent that given in combination with other anti-drugs for the treatment of HBV related HCC, which is resistant to chemotherapy.

Rapid engulfment of apoptotic cells in the absence of inflammation is required for maintenance of normal tissue homeostasis. The low-density lipoprotein receptor-related protein-1 (LRP/CD91) is a receptor mediating interactions between macrophages and apoptotic cells, but recent reports have challenged the requirement of this surface protein in this process. To explore the role of LRP in the recognition of apoptotic cells, target cells were generated with two distinct inducers of apoptotic cell death, etoposide and actinomycin-D. Jurkat T cells rendered apoptotic with etoposide exposed phosphatidylserine (PtdSer) and triggered engulfment by murine bone marrow-derived macrophages (BMDM), however they failed to suppress lipopolysaccharide-driven inflammatory cytokine secretion or, correspondingly, NFĸB-dependent or TNFα promoter-driven transcriptional activity in transfected RAW264.7 macrophages. In contrast, induction of apoptosis in either Jurkat cells or HeLa epithelial cells with actinomycin-D resulted in diminution of proinflammatory signaling from RAW264.7 cells and BMDM. Treatment of actinomycin-treated Jurkat cells with Q-VD-OPh, an irreversible inhibitor of caspase activity, blocked apoptosis, as assessed by the inhibition of PtdSer exposure; however, the cells maintained anti-inflammatory activity. Anti-inflammatory signaling mediated by actinomycin-treated cells was not affected by a macrophage-specific deletion in LRP. Moreover, the presence of LRP on macrophages did not alter the efficiency of engulfment of apoptotic cells in vitro or in vivo. These data demonstrate that the method of induction of apoptosis of target cells influences subsequent macrophage responsiveness, and that LRP is not required for engulfment of apoptotic cells regardless of the method of induction.

Summary Wnt proteins (Wnts) are palmitoylated and glycosylated ligands that play a central role in the early development of organs and tissues. The discovery that loss-of-function mutations in low density lipoprotein receptor-related protein 5 ( LRP5 ), a Wnt co-receptor, led to low bone mass in humans revealed the possible role of Wnt signaling in the regulation of bone remodeling. Many findings obtained from detailed analyses of mice having mutations of Wnt signaling molecules have confirmed that Wnt signaling has potential roles in bone remodeling in both physiological and pathological conditions. There are two pathways of Wnt signaling: β-catenin-dependent canonical and -independent non-canonical pathways. Wnts act on osteoblast precursor cells and promote their differentiation into mature osteoblasts through the β-catenin-dependent canonical pathway. In addition, Wnts suppress bone resorption by regulating the receptor activator of NF-κB ligand (RANKL)/osteoprotegerin (OPG) ratio through the same pathway in mature osteoblasts. In contrast, recent studies have shown that the activation of the β-catenin-independent non-canonical pathway enhances the RANKL-induced osteoclast formation in mouse macrophage cultures. These results indicate that Wnt-mediated signals are involved in several aspects of bone formation and bone resorption. This review will summarize the current knowledge of the roles of Wnt signaling in bone formation and resorption.

Clinical resistance to chemotherapy in acute myeloid leukemia (AML) is associated with the expression of the multidrug resistance (MDR) proteins P-glycoprotein, encoded by the MDR1/ABCB1 gene, multidrug resistant-related protein (MRP/ABCC1), the lung resistance-related protein (LRP), or major vault protein (MVP), and the breast cancer resistance protein (BCRP/ABCG2). The clinical value of MDR1, MRP1, LRP/MVP, and BCRP messenger RNA (mRNA) expression was prospectively studied in 154 newly diagnosed AML patients >= 60 years who were treated in a multicenter, randomized phase 3 trial. Expression of MDR1 and BCRP showed a negative whereas MRP1 and LRP showed a positive correlation with high white blood cell count (respectively, p

In spite of the excellent temporal resolution of event-related EEG potentials (ERPs), the overlapping potentials evoked by masked and masking stimuli are hard to disentangle. However, when both masked and masking stimuli consist of pairs of relevant and irrelevant stimuli, one left and one right from fixation, with the side of the relevant element varying between pairs, effects of masked and masking stimuli can be distinguished by means of the contralateral preponderance of the potentials evoked by the relevant elements, because the relevant elements may independently change sides in masked and masking stimuli. Based on a reanalysis of data from which only selected contralateral-ipsilateral effects had been previously published, the present contribution will provide a more complete picture of the ERP effects in a masked-priming task. Indeed, effects evoked by masked primes and masking targets heavily overlapped in conventional ERPs and could be disentangled to a certain degree by contralateral-ipsilateral differences. Their major component, the N2pc, is interpreted as indicating preferential processing of stimuli matching the target template, which process can neither be identified with conscious perception nor with shifts of spatial attention. The measurements showed that the triggering of response preparation by the masked stimuli did not depend on their discriminability, and their priming effects on the processing of the following target stimuli were qualitatively different for stimulus identification and for response preparation. These results provide another piece of evidence for the independence of motor-related and perception-related effects of masked stimuli.

Lateralized readiness potential (LRP) and time-frequency domain LRP-type measures, called motor-related amplitude asymmetries (MRAA), in the mu band (9-13 Hz; mu-MRAA) and the beta band (18-26 Hz; beta-MRAA) were used to study the time course of preparation in a task-switching task and a response precuing task. Several dissociations between LRP and mu-MRAA and beta-MRAA were found. Mu-MRAA and beta-MRAA, but not LRP, exhibited an early and strong reversal in cortical lateralization when advance preparation for a switch of response hand was required. LRP, but not mu-MRAA or beta-MRAA, was sensitive to manipulation of the probability that advance preparation of response hand would be useful in a response precuing task. These dissociations replicate earlier findings and suggest that movement-related cortical rhythms and cortical potentials are associated with distinct preparatory component processes that differ in terms of level of abstraction and effort, in line with similar functional distinctions between component processes underlying executive control in task switching. This suggests that a fine-grained analysis of subprocesses involved in motor control may provide important guiding principles for the study and understanding of levels and mechanisms of cognitive control. (c) 2006 Elsevier B.V. All rights reserved.

Overexpression of the yeaS gene encoding a protein belonging to the RhtB transporter family conferred upon cells resistance to glycyl-l-leucine, leucine analogues, several amino acids and their analogues. yeaS overexpression promoted leucine and, to a lesser extent, methionine and histidine accumulation by the respective producing strains. Our results indicate that yeaS encodes an exporter of leucine and some other structurally unrelated amino acids. The expression of yeaS (renamed leuE for “leucine export”) was induced by leucine, l-α-amino-n-butyric acid and, to a lesser extent, by several other amino acids. The global regulator Lrp mediated this induction.

The major vault protein is the main component on multimeric vault particles, that are likely to play an essential role in normal cell physiology and to be associated with multidrug resistance of tumour cells. In order to unravel the function of vaults and their putative contribution to multidrug resistance, specific antibodies are invaluable tools. Until now, only conventional major vault protein-reactive murine monoclonal antibodies have been generated, that are most suitable for immunohistochemical analyses. The phage display method allows for selection of human antibody fragments with potential use in clinical applications. Furthermore, cDNA sequences encoding selected antibody fragments are readily identified, facilitating various molecular targeting approaches. In order to obtain such human Fab fragments recognising major vault protein we used a large non-immunized human Fab fragment phage library. Phages displaying major vault protein-reactive Fabs were obtained through several rounds of selection on major vault protein-coated immunotubes and subsequent amplification in TG1 E coli bacteria. Eventually, one major vault protein-reactive clone was selected and further examined. The anti-major vault protein Fab was found suitable for immunohistochemical and Western blot analysis of tumour cell lines and human tissues. BIAcore analysis showed that the binding affinity of the major vault protein-reactive clone almost equalled that of the murine anti-major vault protein Mabs. The cDNA sequence of this human Fab may be exploited to generate an intrabody for major vault protein-knock out studies. Thus, this human Fab fragment should provide a valuable tool in elucidating the contribution(s) of major vault protein/vaults to normal physiology and cellular drug resistance mechanisms. British Journal of Cancer (2002) 86, 954–962. DOI: 10.1038/sj/bjc/6600159 www.bjcancer.com © 2002 Cancer Research UK

It is well established that, in visual pop-out search, reaction time (RT) performance is influenced by cross-trial repetitions versus changes of target-defining attributes. One instance of this is referred to as “positional priming of pop-out” (pPoP; Maljkovic and Nakayama, 1996). In positional PoP paradigms, the processing of the current target is examined depending on whether it occurs at the previous target or a previous distractor location, relative to a previously empty location (“neutral” baseline), permitting target facilitation and distractor inhibition to be dissociated. The present study combined RT measures with specific sensory- and motor-driven event-related lateralizations to track the time course of four distinct processing levels as a function of the target’s position across consecutive trials. The results showed that, relative to targets at previous target and “neutral” locations, the appearance of a target at a previous distractor location was associated with a delayed build-up of the posterior contralateral negativity wave, indicating that distractor positions are suppressed at early stages of visual processing. By contrast, presentation of a target at a previous target, relative to “neutral” and distractor locations, modulated the elicitation of the subsequent stimulus-locked lateralized readiness potential wave, indicating that post-selective response selection is facilitated if the target occurred at the same position as on the previous trial. Overall, the results of present study provide electrophysiological evidence for the idea that target location priming (RT benefits) does not originate from an enhanced coding of target saliency at repeated (target) locations; instead, they arise (near-) exclusively from processing levels subsequent to focal-attentional target selection.

Background: Axillary node status after induction chemotherapy for locally advanced breast cancer has been shown on multivariate analysis to be an independent predictor of relapse. However, it has been postulated that responders to induction chemotherapy with a clinically negative axilla could be spared the burden of lymphadenectomy, because most of them will not show histological nodal invasion. P-glycoprotein expression in the rescue mastectomy specimen has finally been identified as a significant predictor of patient survival. Methods: We studied the expression of the genes encoding multidrug resistance associated protein (MDR1) and lung cancer associated resistance protein (LRP) in formalin-fixed, paraffin-embedded tumor samples from 52 patients treated for locally advanced breast cancer by means of induction chemotherapy followed by rescue mastectomy. P-glycoprotein expression was assessed by means of immunohistochemistry before treatment in 23 cases, and by means of reverse-transcriptase-mediated polymerase chain reaction (RT-PCR) after treatment in 46 (6 failed). LRP expression was detected by means of immunohistochemistry, with the LRP-56 monoclonal antibody, in 31 cases before treatment. Immunohistochemistry for detecting the expression of c-erb-B2, p53, Ki67, estrogen receptor and progesterone receptor are routinely performed in our laboratory in every case, and the results obtained were included in the study. All patients had received between two and six cycles of standard 5-fluorouracil, doxorubicin and cyclophosphamide (FAC) chemotherapy, with two exceptions [one patient received four cycles of a docetaxel-adriamycin combination, and the other four cycles of standard cyclophosphamide-methotrexate-5-fluorouracil (CMF) polychemotherapy]. Response was assessed in accordance with the Response Evaluation Criteria In Solid Tumors (RECIST). By these, 2 patients achieved a complete clinical response, 37 a partial response, and the remaining 13 showed stable disease. This makes a total clinical response rate of 75.0%. None achieved a complete pathological response. Results: MDR1 mRNA expression detected by RT-PCR was associated with the presence of invaded axillary nodes at surgery in 18/22 cases (81.8%), compared with 13/24 (54.2%) in the group with undetectable MDR1 expression. This difference was statistically significant (P < 0.05). LRP expression in more than 20% of tumor cells before any treatment was associated with axillary nodal metastasis after chemotherapy and rescue mastectomy in 17/23 cases, compared with 3/8 in nonexpressors. Again, this difference was highly significant (P < 0.01). LRP expression before treatment and MDR1 mRNA expression after treatment were significantly interrelated (P < 0.001), which might reflect the presence of chemoresistant clones liable to metastasize to the regional nodes. Persistence of previously detected MDR1-positivity after treatment (7/9 compared with 0/2 cases) was significantly associated with axillary node metastasis (P < 0.05). Finally, in a logistic regression multivariate model, histology other than ductal, a Ki67 labeling index of at least 20% and the combination of LRP and MDR1 positivity emerged as independent predictors of axillary node invasion at the time of rescue mastectomy. Conclusion: The expression of different genes involved in resistance to chemotherapy, both before and after treatment with neoadjuvant, is associated with the presence of axillary node invasion at rescue surgery in locally advanced breast cancer. This might reflect the presence of intrinsically resistant clones before any form of therapy, which persist after it, and could be helpful both for prognosis and for the choice of individual treatment.

Multidrug resistance (MDR) and more specifically the expression of P-glycoprotein (Pgp) have been studied extensively in vitro. Unfortunately, it appears that the predictive value of MDR recognized in vitro is mostly an incorrect measure to determine the responsiveness of a particular tumour in the clinic. This misunderstood or overvalued role of MDR might explain the failure of strategies to reverse Pgp function by the use of modulators in solid tumours. To obtain more insight in in vivo drug resistance we investigated a panel of 15 human ovarian cancer xenografts consisting of the most common histological subtypes known in ovarian cancer patients. The response rate to cisplatin, cyclophosphamide and doxorubicin in the xenografts resembled the results of phase II trials with these agents in ovarian cancer patients. This resemblance justifies drug resistance studies in this experimental in vivo human tumour system. We determined the expression levels of MDR 1, MRP 1, LRP and topoisomerase IIα mRNA by the RNase protection assay and the presence of MRP1 and LRP proteins by immunohistochemistry. The S-phase fraction was investigated as a separate parameter by flow cytometry. In none of the 15 ovarian cancer xenografts was MDR 1 expression detectable. The expression levels of MRP 1 and LRP were low to moderate and resembled the presence of the MRP1 and LRP proteins. There was a weak, inverse relationship between the expression levels of LRP and sensitivity to cisplatin and cyclophosphamide (r = –0.44 and –0.45), but not to doxorubicin. The levels of topoisomerase IIα varied among the xenografts (0.73–2.66) and failed to correlate with doxorubicin resistance (r = 0.14). The S-phase fraction, however, showed a relation with the sensitivity to cisplatin (r = 0.66). Among the determinants studied in ovarian cancer in vivo, LRP mRNA and the S-phase fraction were the best predictive factors for drug response and most specifically for the activity of cisplatin. © 2000 Cancer Research Campaign

The processes responsible for the uptake of chylomicron remnants by macrophages were investigated using freshly isolated cells from low density lipoprotein (LDL) receptor, very low density lipoprotein (VLDL) receptor and apolipoprotein E knockout mice. In peritoneal macrophages from normal mice, the metabolism of chylomicron remnants was inhibited 40% by anti-LDL receptor antibody and 60% by a high concentration of receptor-associated protein (RAP). Together they reduced the amount processed by 70%. Digestion of cell proteoglycans decreased remnant degradation by 20% while the addition of acetyl-LDL had no effect. When LDL receptors were absent, the absolute rates of metabolism were less than that of normal cells and were not inhibited by the anti-LDL receptor antibody; the rates, however, were reduced to less than half by RAP. These suggest that the LDL receptor-related protein (LRP) or another LDL receptor family member(s) contributes to chylomicron remnant uptake and becomes the major mechanism of uptake when LDL receptors are absent. In contrast, the VLDL receptor was not involved as its absence did not affect chylomicron remnant metabolism. Similarly, the absence of apoE production did not affect the amount of remnant uptake; however, the proportion that was sensitive to RAP was eliminated. The level of LRP expression was not altered in these cells whereas there was a decrease in LDL receptors. This suggests that the apoE content of chylomicron remnants is sufficient for its recognition by LDL receptors but additional apoE is required for its uptake by the LRP and that there is an up-regulation of a non-LDL receptor family mechanism in apoE deficiency. Together these studies suggest that even in the absence of LDL receptors or apoE secretion, chylomicron remnants could contribute to lipid accumulation in the artery wall during atherogenesis.—Fujioka, Y., A. D. Cooper, and L. G. Fong. Multiple processes are involved in the uptake of chylomicron remnants by mouse peritoneal macrophages. J. Lipid Res. 1998. 39: 2339–2349.

Apolipoprotein (apo) E plays a central role in the transport of lipids among different organs and cell types, whereas α2-macroglobulin (α2M) is responsible for the binding and inactivation of plasma proteases, as well as the transport of various cytokines, growth factors, and hormones. In the present study, evidence is presented for direct binding of apoE with α2M in human plasma, based on the observation that two-dimensional non-denaturing gradient gel electrophoretic separation of plasma resulted in co-migration of apoE with α2M in a complex intermediate in size (18.5 nm in diameter) between low (LDL) and high density lipoproteins (HDL). ApoE associated with α2M could be immunoprecipitated from plasma with anti-human α2M antiserum. Purified apoE, labeled with 125I, bound to native and methylamine-activated α2M (α2M-MA) in vitro in a time- and concentration-dependent manner. ApoE bound to α2M-MA with greater affinity than α2M. The binding of apoE to both α2M and α2M-MA did not depend on the presence of lipid. Ingestion of an oral fat load resulted in a reduction in the amount of apoE associated with α2M. Sphingomyelin vesicles and very low density lipoproteins (VLDL), but not phosphatidylcholine vesicles or HDL3, inhibited the in vitro binding of 125I-labeled apoE3 to α2M and α2M-MA. Binding of 125I-labeled apoE3 was also partially inhibited by an excess of platelet-derived growth factor and β-amyloid protein, but not interferon-γ. Subjects with an apoE 4/4 phenotype had less apoE associated with α2M in plasma than subjects with an apoE 3/3 or 2/2 phenotype, corresponding to reduced in vitro binding of apoE4 with α2M or α2M-MA. Although the functional significance of apoE binding to α2M remains to be determined, the present results demonstrate that: 1) apoE is non-covalently bound to α2M in human plasma, 2) α2M-MA has a greater capacity to bind apoE than α2M, 3) various proteins or lipoproteins known to bind apoE or α2M can potentially affect the interaction of apoE with α2M, and 4) association of apoE with α2M or α2M-MA is dependent on apoE phenotype. —Krimbou, L., M. Tremblay, J. Davignon, and J. S. Cohn. Association of apolipoprotein E with α2-macroglobulin in human plasma. J. Lipid Res. 1998. 39: 2373–2386.

ApoE, ApoE receptors and APP cooperate in the pathogenesis of Alzheimer’s disease. Intriguingly, the ApoE receptor LRP4 and APP are also required for normal formation and function of the neuromuscular junction (NMJ). In this study, we show that APP interacts with LRP4, an obligate co-receptor for muscle-specific tyrosine kinase (MuSK). Agrin, a ligand for LRP4, also binds to APP and co-operatively enhances the interaction of APP with LRP4. In cultured myotubes, APP synergistically increases agrin-induced acetylcholine receptor (AChR) clustering. Deletion of the transmembrane domain of LRP4 (LRP4 ECD) results in growth retardation of the NMJ, and these defects are markedly enhanced in APP−/−;LRP4ECD/ECD mice. Double mutant NMJs are significantly reduced in size and number, resulting in perinatal lethality. Our findings reveal novel roles for APP in regulating neuromuscular synapse formation through hetero-oligomeric interaction with LRP4 and agrin and thereby provide new insights into the molecular mechanisms that govern NMJ formation and maintenance. DOI: http://dx.doi.org/10.7554/eLife.00220.001, eLife digest One of the hallmarks of Alzheimer’s disease is the formation of plaques in the brain by a protein called β-amyloid. This protein is generated by the cleavage of a precursor protein, and mutations in the gene that encodes amyloid precursor protein greatly increase the risk of developing a familial, early-onset form of Alzheimer’s disease in middle age. Individuals with a particular variant of a lipoprotein called ApoE (ApoE4) are also more likely to develop Alzheimer’s disease at a younger age than the rest of the population. Due to its prevalence—approximately 20% of the world’s population are carriers of at least one allele—ApoE4 is the single-most important risk factor for the late-onset form of Alzheimer’s disease. Amyloid precursor protein and the receptors for ApoE—in particular one called LRP4—are also essential for the development of the specialized synapse that forms between motor neurons and muscles. However, the mechanisms by which they, individually or together, contribute to the formation of these neuromuscular junctions are incompletely understood. Now, Choi et al. have shown that amyloid precursor protein and LRP4 interact at the developing neuromuscular junction. A protein called agrin, which is produced by motor neurons and which must bind to LRP4 to induce neuromuscular junction formation, also binds directly to amyloid precursor protein. This latter interaction leads to the formation of a complex between LRP4 and amyloid precursor protein that robustly promotes the formation of the neuromuscular junction. Mutations that remove the part of LRP4 that anchors it to the cell membrane weaken this complex and thus reduce the development of neuromuscular junctions in mice, especially if the animals also lack amyloid precursor protein. These three proteins thus seem to influence the development and maintenance of neuromuscular junctions by regulating the activity of a fourth protein, called MuSK, which is present on the surface of muscle cells. Activation of MuSK by agrin bound to LRP4 promotes the clustering of acetylcholine receptors in the membrane, which is a crucial step in the formation of the neuromuscular junction. Intriguingly, Choi et al. have now shown that amyloid precursor protein can, by interacting directly with LRP4, also activate MuSK even in the absence of agrin, albeit only to a small extent. The work of Choi et al. suggests that the complex formed between agrin, amyloid precursor protein and LRP4 helps to focus the activation of MuSK, and thus the clustering of acetylcholine receptors, to the site of the developing neuromuscular junction. Since all four proteins are also found in the central nervous system, similar processes might well be at work during the development and maintenance of synapses in the brain. Further studies of these interactions, both at the neuromuscular junction and in the brain, should shed new light on both normal synapse formation and the synaptic dysfunction that is seen in Alzheimer’s disease. DOI: http://dx.doi.org/10.7554/eLife.00220.002

Background Microglia are resident brain macrophages that can phagocytose dead, dying or viable neurons, which may be beneficial or detrimental in inflammatory, ischaemic and neurodegenerative brain pathologies. Cell death caused by phagocytosis of an otherwise viable cell is called ‘primary phagocytosis’ or ‘phagoptosis’. Calreticulin (CRT) exposure on the surface of cancer cells can promote their phagocytosis via LRP (low-density lipoprotein receptor-related protein) on macrophages, but it is not known whether this occurs with neurons and microglia. Methods We used primary cultures of cerebellar neurons, astrocytes and microglia to investigate the potential role of CRT/LRP phagocytic signalling in the phagocytosis of viable neurons by microglia stimulated with lipopolysaccharide (LPS) or nanomolar concentrations of amyloid-β peptide1-42 (Aβ). Exposure of CRT on the neuronal surface was investigated using surface biotinylation and western blotting. A phagocytosis assay was also developed using BV2 and PC12 cell lines to investigate CRT/LRP signalling in microglial phagocytosis of apoptotic cells. Results We found that BV2 microglia readily phagocytosed apoptotic PC12 cells, but this was inhibited by a CRT-blocking antibody or LRP-blocking protein (receptor-associated protein: RAP). Activation of primary rat microglia with LPS or Aβ resulted in loss of co-cultured cerebellar granule neurons, and this was blocked by RAP or antibodies against CRT or against LRP, preventing all neuronal loss and death. CRT was present on the surface of viable neurons, and this exposure did not change in inflammatory conditions. CRT antibodies prevented microglia-induced neuronal loss when added to neurons, while LRP antibodies prevented neuronal loss when added to the microglia. Pre-binding of CRT to neurons promoted neuronal loss if activated microglia were added, but pre-binding of CRT to microglia or both cell types prevented microglia-induced neuronal loss. Conclusions CRT exposure on the surface of viable or apoptotic neurons appears to be required for their phagocytosis via LRP receptors on activated microglia, but free CRT can block microglial phagocytosis of neurons by acting on microglia. Phagocytosis of CRT-exposing neurons by microglia can be a direct cause of neuronal death during inflammation, and might therefore contribute to neurodegeneration and be prevented by blocking the CRT/LRP pathway.

Objective To reveal the mechanism of Zuogui Pill ( ) in treatment of glucocorticoid-induced osteoporosis from the angle of the Wnt signal transduction pathway and to provide further experimental evidence for expounding the scientific connotation of “the kidney dominating the bones” in TCM. Methods Forty-two male Wistar rats were selected and randomly divided into three groups, control group (n=12), model group (n=15) and Zuogui Pill group (n=15). Form the beginning, The rats were injected dexamethasone for eight weeks to make the model of osteoporosis, and the Zuogui Pill were administered intragastrically to the rats of Zuogui Pill group for eight weeks. The relative morphological parameters were measured in the undecalcified tibial slices. And the protein expression levels of Wnt1, LRP-5 and β-catenin in rat tibial osteoblasts (OB) and bone marrow stromal cells (BMC) were detected by immunohistochemistry. Results Compared with the control group, TBV% and TFS% decreased significantly, while TRS% increased significantly, and the protein expression of Wnt1, LRP-5 and β-catenin in OB and BMC decreased significantly in the model group. And compared with the model group, TBV% and TFS% increased significantly, and expression levels of Wnt1, LRP-5 and β-catenin proteins increased significantly in the Zuogui pill group. Conclusion Zuogui Pill can prevent and treat glucocorticoid-induced osteoporosis in rats by up-regulating the expression of the key signal molecules Wnt1, LRP-5 and β-catenin in Wnt signal transduction pathway.

Highlights ► The pathophysiological link between NAFLD and hepatic insulin resistance is unknown. ► We studied the effect of postprandial lipoproteins on hepatic insulin sensitivity. ► Postprandial lipoproteins cause liver steatosis and hepatic insulin resistance. ► We characterize the underlying molecular mechanisms. ► Postprandial lipoproteins are a link between NAFLD and hepatic insulin resistance., Non-alcoholic fatty liver disease (NAFLD) is associated with hepatic insulin resistance with the molecular basis of this association being not well understood. Here we studied the effect of hepatic triglyceride accumulation induced by postprandial triglyceride-rich lipoproteins (TGRL) on hepatic insulin sensitivity in HepG2 cells. Incubation of HepG2 cells with purified TGRL particles induced hepatocellular triglyceride accumulation paralleled by diminished insulin-stimulated glycogen content and glycogen synthase activity. Accordingly, insulin-induced inhibition of glycogen synthase phosphorylation as well as insulin-induced GSK-3 and AKT phosphorylation were reduced by TGRL. The effects of TGRL were dependent on the presence of apolipoproteins and more pronounced for denser TGRL. Moreover, TGRL effects required the presence of heparan sulfate-proteoglycans on the cell membrane and lipase activity but were independent of the cellular uptake of TGRL particles by receptors of the LDL receptor family. We suggest postprandial lipemia to be an important factor in the pathogenesis of NAFLD.

Wnts are known to bind and activate multiple membrane receptors/coreceptors and to regulate dopaminergic (DA) neuron development and ventral midbrain (VM) morphogenesis. The low density lipoprotein receptor–related protein (Lrp6) is a Wnt co-receptor, yet it remains unclear whether Lrp6 is required for DA neuron development or VM morphogenesis. Lrp6 is expressed ubiquitously in the developing VM. In this study, we show that Lrp6 2/2 mice exhibit normal patterning, proliferation and cell death in the VM, but display a delay in the onset of DA precursor differentiation. A transient 50% reduction in tyrosine hydroxylase–positive DA neurons and in the expression of DA markers such as Nurr1 and Pitx3, as well as a defect in midbrain morphogenesis was detected in the mutant embryos at embryonic day 11.5. Our results, therefore, suggest a role for Lrp6 in the onset of DA neuron development in the VM as well as a role in midbrain morphogenesis. Developmental Dynamics 239:211–221, 2010. V C 2009 Wiley-Liss, Inc.

Background:Molecular mechanism of differentiation in lipogenic tumor is still unknown in detail. Low-density lipoprotein receptor-related protein (LRP) and peroxisome proliferator-activated receptor gamma (PPAR-γ), representative regulatory molecules of lipogenic differentiation, have been reported today as multi-functional molecules and to modulate tumorigenesis in various kind of cancer. To date, diagnostic and therapeutic significance of the expression of these molecules in lipogenic tumors are not defined.Methods:The immunohistochemical expression status of LRP and PPAR-γ in various grades of 54 lipogenic tumors was analyzed. Correlation between the expression levels and the differentiation of the tumors was confirmed. Forstatistical analyses, the Kruskal-Wallis test, the Steel-Dwass test and the Mann–Whitney U test were used.Results:LRP and PPAR-γ expression was detected in 50 (92.6%) and 44 (81.5%) cases, respectively. The expression level in LRP was significantly higher in cases with well differentiated liposarcoma, pleomorphic liposarcoma and dedifferentiated liposarcoma than in lipoma. Compared with lipoma or well differentiated liposarcoma, significant elevation in expression level of PPAR-γ was confirmed in myxoid liposarcoma, pleomorphic liposarcoma, dedifferentiated liposarcoma and the differentiated area of dedifferentiated liposarcoma.Conclusion:The up-regulation of LRP and PPAR-γ in higher grade cases, i.e. less differentiated tumors than in low grade cases was shown, suggesting the candidate role of these molecules as tumor progression modulators rather than regulatory molecules of differentiation in lipogenic tumors.

Zhi Jiang1,6, Bao-An Chen1,6, Guo-Hua Xia1, Qiang Wu2, Yu Zhang1, Tie-Yan Hong1, Wei Zhang1, Jian Cheng1, Feng Gao1, Li-Jie Liu3, Xiao-Mao Li4, Xue-Mei Wang51Department of Hematology, the Affiliated Zhongda Hospital of Southeast University, Nanjing, China; 2The Jiangsu Province Cancer Hospital, Nanjing, China; 3Institutions of Physiology, Southeast University, Nanjing, China; 4Department of Physics, University of Saarland, Saarbruechen, Germany; 5National Key Lab of Bioelectronics (Chien-Shiung Wu Laboratory), Southeast University, Nanjing, China; 6These authors have contributed equally to this workAbstract: To explore whether the magnetic nanoparticles of Fe3O4 (MNPs-Fe3O4) loaded with cisplatin can reverse the diaminedichloro platinum (DDP) resistance to multidrug resistance of ovarian carcinoma cells and to investigate its mechanisms. The SKOV3/DDP cells were divided into DDP treatment (DDP group), MNPs-Fe3O4 treatment (MNPs-Fe3O4 group), DDP + MNPs-Fe3O4 treatment (DDP + MNPs-Fe3O4 group), and control group. After incubation with those conjugates for 48 h, the cytotoxic effects were measured by MTT assay. Apoptosis and the intracellular DDP concentration were investigated by flow cytometry and inductively coupled plasma atomic emission spectroscopy, respectively. The expression of apoptosis associated gene Bcl-2 mRNA was detected by reverse transcription polymerase chain reaction and the expressions of MDR1, lung resistance-related protein (LRP), and P-glycoprotein (P-gp) genes were studied by Western blot. Our results indicated that the 50% inhibition concentration (IC50) of the MNPs-Fe3O4 loaded with DDP was 17.4 µmol/ l, while the IC50 was 39.31 µmol/l in DDP groups (p< 0.05); Apoptosis rates of SKOV3/DDP cells increased more than those of DDP groups. Accumulation of intracellular cisplatin in DDP + MNPs-Fe3O4 groups was higher than those in DDP groups (p< 0.05). Moreover, the expression of Bcl-2 mRNA and the protein expressions of MDR1, LRP, and P-gp were decreased when compared with those of DDP groups, respectively. Our results suggest that MNPs-Fe3O4 can reverse the DDP resistance to the ovarian carcinoma cell. The effects may be associated with over-expression of MDR1, LRP, P-gp, and Bcl-2, which can increase the intracellular platinum accumulation and induce the cell apoptosis.Keywords: magnetic nanoparticles of Fe3O4 , multidrug resistance reversal, SKOV3/DDP, MDR1, LRP, P-gp, Bcl-2

Many Escherichia coli K-12 strains display an intrinsic resistance to the peptide antibiotic microcin J25. In this study, we present results showing that the leucine-responsive regulatory protein, Lrp, is involved in this phenotype by acting as a positive regulator of YojI, a chromosomally encoded efflux pump which expels microcin out of cells. Exogenous leucine antagonizes the effect of Lrp, leading to a diminished expression of the pump and an increased susceptibility to microcin J25. © 2009, American Society for Microbiology. All Rights Reserved. Fil: Socias, Sergio Benjamin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentina Fil: Vincent, Paula Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentina Fil: Salomon, Raul Armando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentina

Visual stimuli undetected by normal subjects as a result of masking procedures can nonetheless activate response preparation in motor areas and yield a motor response. An unanswered question is whether the same holds for undetected subliminal stimuli that are not responded to. To answer this question, in this study normal subjects were tested on a simple visual reaction time task with stimuli above, at, or below the psychophysical threshold while the lateralized readiness potential (LRP), i.e. an electrophysiological correlate of premotor activation in the primary motor cortex, was computed. We found a reliable LRP not only for suprathreshold stimuli but also for subthreshold stimuli to which subjects did not respond. The main thrust of this study is that it provides evidence that activation of the motor cortex occurs even with subthreshold visual stimuli and without an overt response.

The present study investigates the event related potential (ERP) components associated with the random version of the Sustained Attention to Response Task (SART). The random SART is a Go/No-Go task in which the No-Go target appears unpredictably and rarely. In the present experiment, the EEG was recorded from 58 electrodes with mastoids as reference. As expected, the N2 and P3 were larger in the No-Go trials compared to the Go trials, with the P3 more frontal for the No-Go trials compared to the Go trials. The functional role of the N2 and P3 in the No-Go trials is discussed comparing our results to the fixed SART results obtained by Dockree et al. [Dockree P. M., Kelly S. P., Robertson I. H., Reilly R. B., & Foxe J. J. (2005). Neurophysiological markers of alert responding during goal-directed behaviour: A high-density electrical mapping study. NeuroImage, 27, 587-601]. In addition, the lateralized readiness potential (LRP) was computed. The absence of an LRP in the No-Go trials suggests that a central inhibitory mechanism intervenes to prevent the preparation and execution of a dominant motor response.

The task-switching paradigm provides an opportunity to study whether oscillatory relations in neuronal activity are involved in switching between and maintaining task sets. The EEG of subjects performing an alternating runs [Rogers, R.D., Monsell, S., 1995. Costs of a predictable switch between simple cognitive tasks. Journal of Experimental Psychology: General 124, 207-231] task-switching task was analyzed using event-related potentials, the lateralized readiness potential, instantaneous amplitude and the phase-locking value [Lachaux, J.P., Rodriguez, E., Martinirie, J., Varela, F.J., 1999. Measuring phase synchrony in brain signals. Human Brain Mapping 8, 194-208]. The two tasks differed in the relevant modality (visual versus auditory) and the hand with which responses were to be given. The mixture model [de Jong, R., 2000. An intention driven account of residual switch costs. In: Monsell, S., Driver, J. (Eds.), Attention and Performance XVII: Cognitive Control. MIT Press, Cambridge] was used to assign pre-stimulus switch probabilities to switch trials based on reaction time; these probabilities were used to create a fast-slow distinction between trials on both switch and hold trials. Results showed both time- and time-frequency-domain effects, during the intervals preceding stimuli, of switching versus maintenance, response speed of the upcoming stimulus, and response hand. Of potential importance for task-switching theory were interactions between reaction time by switch-hold trial type that were found for a frontal slow negative potential and the lateralized readiness potential during the response-stimulus interval, indicating that effective preparation for switch trials involves different anticipatory activity than for hold trials. Theta-band oscillatory activity during the pre-stimulus period was found to be higher when subsequent reaction times were shorter, but this response speed effect did not interact with trial type. The response hand of the upcoming task was associated with lateralization of pre-stimulus mu- and beta-band amplitude and, specifically for switch trials, beta-band phase locking. (c) 2005 Elsevier B.V. All rights reserved.

The biological behaviour of different histological types and grades of soft tissue sarcomas (STS) varies. This might result in a differing sensitivity to cytotoxic drugs. Cross-resistance to functionally and structurally distinct natural-product drugs, known as multidrug resistance (MDR), is associated with the overexpression of P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1) and lung resistance-related protein (LRP). The purpose of this study was to evaluate the expression of P-gp, MRP1 and LRP in STS according to their histological type and grade. In 141 chemotherapy-naive STS patients, the expression of the three MDR proteins was detected by immunohistochemistry. Nine histological types were documented. These were 19% grade 1, 34% grade 2 and 47% grade 3 tumours. Expression of P-gp and LRP was observed more frequently than the expression of MRP1 (P

Using random Tn10 insertion mutagenesis, we isolated an Escherichia coli mutant strain affected in the regulation of lysU, the gene encoding the inducible form of lysyl-tRNA synthetase. The transposon giving rise to the altered expression of lysU was found inserted within lrp. The latter gene codes for the leucine-responsive regulatory protein (Lrp) which mediates a global response of the bacterium to leucine. An involvement of Lrp in the regulation of lysU was searched for by using a lysU-lacZ operon fusion. The following conclusions were reached: (i) inactivation of lrp causes an increased activity of the lysU promoter, whatever the growth conditions assayed, (ii) insertion of a wild-type lrp gene into a multi-copy plasmid significantly reduces lysU expression, and (iii) sensitivity of the lysU promoter to the presence of leucine in the growth medium is abolished in the lrp context.

Aims/hypothesis Our recent studies suggest that activation of the wingless-type MMTV integration site (WNT) pathway plays pathogenic roles in diabetic retinopathy and age-related macular degeneration. Here we investigated the causative role of oxidative stress in retinal WNT pathway activation in an experimental model of diabetes. Methods Cultured retinal pigment epithelial cells and retinal capillary endothelial cells were treated with a lipid peroxidation product, 4-hydroxynonenal (HNE), and an antioxidant, N-acetyl-cysteine (NAC). In vivo, rats with streptozotocin-induced diabetes were treated by NAC for 8 weeks. Activation of the canonical WNT pathway was measured by TOPFLASH assay and by western blot analysis of WNT pathway components and a WNT target gene, Ctgf. Oxidative stress in the retina was evaluated by immunostaining of HNE and 3-nitrotyrosine. Results Levels of phosphorylated and total LDL receptor-related protein (LRP)6, and cytosolic β-catenin, as well as transcriptional activity of T cell factor (TCF)/β-catenin were significantly increased by HNE. The production of connective tissue growth factor (CTGF) was also upregulated by HNE. NAC blocked the WNT pathway activation induced by HNE. Furthermore, LRP6 stability was increased by HNE and decreased by NAC. Retinal levels of HNE and 3-nitrotyrosine were significantly increased in diabetic rats, compared with those in non-diabetic rats. In the same diabetic rat retinas, levels of LRP6, cytosolic β-catenin and CTGF were significantly increased. NAC treatment reduced HNE and 3-nitrotyrosine levels and attenuated the upregulation of LRP6, β-catenin and CTGF in diabetic rat retina. Conclusions/interpretation Lipid peroxidation products activate the canonical WNT pathway through oxidative stress, which plays an important role in the development of retinal diseases.