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Interferon-induced transmembrane protein 3 (IFITM3) is a restriction factor that limits viral pathogenesis and exerts poorly understood immunoregulatory functions. Here, using human and mouse models, we demonstrate that IFITM3 promotes MyD88-dependent, TLR-mediated IL-6 production following exposure to cytomegalovirus (CMV). IFITM3 also restricts IL-6 production in response to influenza and SARS-CoV-2. In dendritic cells, IFITM3 binds to the reticulon 4 isoform Nogo-B and promotes its proteasomal degradation. We reveal that Nogo-B mediates TLR-dependent pro-inflammatory cytokine production and promotes viral pathogenesis in vivo, and in the case of TLR2 responses, this process involves alteration of TLR2 cellular localization. Nogo-B deletion abrogates inflammatory cytokine responses and associated disease in virus-infected IFITM3-deficient mice. Thus, we uncover Nogo-B as a driver of viral pathogenesis and highlight an immunoregulatory pathway in which IFITM3 fine-tunes the responsiveness of myeloid cells to viral stimulation.

Peripheral arterial disease (PAD) is the least studied complication of nephrotic syndrome (NS). Risk factors which predispose children with NS to developing PAD include hyperlipidaemia, hypertension and prolonged use of steroids. The development of PAD significantly increases the morbidity and mortality associated with NS as such children are prone to sudden cardiac death. The ankle brachial index (ABI) is a tool that has been proven to have high specificity and sensitivity in detecting PAD even in asymptomatic individuals. We aimed to determine the prevalence of PAD in children with NS and to identify risk factors that can independently predict its development. A comparative cross-sectional study was conducted involving 200 subjects (100 with NS and 100 apparently healthy comparative subjects that were matched for age, sex and socioeconomic class). Systolic blood pressures were measured in all limbs using the pocket Doppler machine (Norton Doppler scan machine). ABI was calculated as a ratio of ankle to arm systolic blood pressure. PAD was defined as ABI less than 0.9. The prevalence of PAD was significantly higher in children with NS than matched comparison group (44.0% vs 6.0%, p < 0.001). Average values of waist and hip circumference were significantly higher in subjects with PAD than those without PAD (61.68± 9.1cm and 67.6± 11.2 cm vs 57.03 ± 8.3cm and 65.60± 12.5cm respectively, p< 0.005). Serum lipids (triglyceride, very low density lipoprotein, total cholesterol and low density lipoprotein) were also significantly higher in subjects with PAD than those without PAD [106.65mg/dl (67.8–136.7) vs 45.72mg/dl (37.7–61.3), 21.33mg/dl (13.6–27.3) vs 9.14mg/dl (7.5–12.3), 164.43mg/dl (136.1–259.6) vs 120.72mg/dl (111.1–142.1) and 93.29mg/dl (63.5–157.3) vs 61.84mg/dl (32.6–83.1), respectively p< 0.05]. Increasing duration since diagnosis of NS, having a steroid resistant NS and increasing cumulative steroid dose were independent predictors of PAD in children with NS; p< 0.05 respectively. With these findings, it is recommended that screening for PAD in children with NS should be done to prevent cardiovascular complications before they arise.

SummaryIntracellular bacterial pathogens inject effector proteins into host cells to hijack diverse cellular processes and promote their survival and proliferation. To systematically map effector-host protein-protein interactions (PPIs) during infection, we generated a library of 32Salmonella entericaserovar Typhimurium (STm) strains expressing chromosomally encoded affinity-tagged effector proteins, and quantified PPIs in macrophages and epithelial cells by Affinity-Purification Quantitative Mass-Spectrometry. Thereby, we identified 25 previously described and 421 novel effector-host PPIs. While effectors converged on the same host cellular processes, most had multiple targets, which often differed between cell types. Using reciprocal co-immunoprecipitations, we validated 13 out of 22 new PPIs. We then used this host-pathogen physical interactome resource to demonstrate that SseJ and SseL collaborate in redirecting cholesterol to theSalmonellaContaining Vacuole (SCV) via NPC1, PipB directly recruits the organelle contact site protein PDZD8 to the SCV, and SteC promotes actin bundling by directly phosphorylating formin-like proteins.

Purpose: Mantle cell lymphoma (MCL) is a fatal subtype of non–Hodgkin lymphoma. SOX11 transcription factor is overexpressed in the majority of nodal MCL. We have previously reported that B cell–specific overexpression of SOX11 promotes MCL pathogenesis via critically increasing BCR signaling in vivo. SOX11 is an attractive target for MCL therapy; however, no small-molecule inhibitor of SOX11 has been identified to date. Although transcription factors are generally considered undruggable, the ability of SOX11 to bind to the minor groove of DNA led us to hypothesize that there may exist cavities at the protein–DNA interface that are amenable to targeting by small molecules. Experimental Design: Using a combination of in silico predictions and experimental validations, we report here the discovery of three structurally related compounds (SOX11i) that bind SOX11, perturb its interaction with DNA, and effect SOX11-specific anti-MCL cytotoxicity. Results: We find mechanistic validation of on-target activity of these SOX11i in the inhibition of BCR signaling and the transcriptional modulation of SOX11 target genes, specifically, in SOX11-expressing MCL cells. One of the three SOX11i exhibits relatively superior in vitro activity and displays cytotoxic synergy with ibrutinib in SOX11-expressing MCL cells. Importantly, this SOX11i induces cytotoxicity specifically in SOX11-positive ibrutinib-resistant MCL patient samples and inhibits Bruton tyrosine kinase phosphorylation in a xenograft mouse model derived from one of these subjects. Conclusions: Taken together, our results provide a foundation for therapeutically targeting SOX11 in MCL by a novel class of small molecules.

The severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is a global threat to human health and has compromised economic stability. In addition to the development of an effective vaccine, it is imperative to understand how SARS‐CoV‐2 hijacks host cellular machineries on a system‐wide scale so that potential host‐directed therapies can be developed. In situ proteome‐wide abundance and thermal stability measurements using thermal proteome profiling (TPP) can inform on global changes in protein activity. Here we adapted TPP to high biosafety conditions amenable to SARS‐CoV‐2 handling. We discovered pronounced temporal alterations in host protein thermostability during infection, which converged on cellular processes including cell cycle, microtubule and RNA splicing regulation. Pharmacological inhibition of host proteins displaying altered thermal stability or abundance during infection suppressed SARS‐CoV‐2 replication. Overall, this work serves as a framework for expanding TPP workflows to globally important human pathogens that require high biosafety containment and provides deeper resolution into the molecular changes induced by SARS‐CoV‐2 infection., Thermal proteome profiling (TPP) measurements of protein state and interaction changes in SARS‐CoV‐2 infection reveal pronounced temporal alterations in host protein thermostability during infection and indicate druggable host targets.

Pioneering work by among others the groups of Hunt and Mann at the start of this century opened up the era of mass spectrometry based phosphoproteomics1-3. Through advances in sample preparation, phosphopeptide enrichment, mass spectrometric detection, peptide sequencing and dedicated database searches this field has matured and it is nowadays possible to monitor thousands of protein phosphorylation events qualitatively and quantitatively4-8. Such phosphoproteomics studies are important as protein phosphorylation is regulating nearly all biological processes and the deregulation of phosphorylation has been associated with the onset of various diseases9-15. Notwithstanding this maturation and broad acceptance of mass spectrometry based phosphoproteomics by the research community, the mass spectrometric (MS) analysis of phosphorylation remains demanding, mainly due to the often severe substoichiometric levels of phosphorylation and the intrinsic lability of the phosphate group, hampering both enrichment and unambiguous sequencing analysis of phosphopeptides. Here we review the increasing knowledge gathered about the mechanisms behind the fragmentation of phosphopeptide ions and describe additionally recent advances made to improve and facilitate the identification and site localization of phosphorylated peptides. Since our earlier related review in 200916 several fragmentation methods have been introduced and successfully applied to phosphopeptides, such as electron capture (ECD), electron transfer induced dissociation(ETD), and hybrid fragmentation techniques such as EThcD (a combination of ETD and HCD) and AI-ETD (a combination of ETD with infrared photo-activation). A plethora of data has been gathered on endogenous Ser, Thr, Tyr phosphorylation, but also on large synthetic phosphopeptide libraries, providing new insight into the mechanisms behind phosphopeptide fragmentation. Moreover, data on phosphopeptides harboring phosphorylated Histidine, Arginine or Lysine have become available, which exhibit some distinct fragmentation behaviors. Through all this acquired knowledge on phosphopeptide fragmentation and site localization mass spectrometry based phosphoproteomics has matured into a respectable tool for the broader life science research community.

Nm23/NME was identified 30 years ago as the first metastatic gene suppressor family. Despite extensive studies, the mechanism of action behind the observed antimetastatic potential of Nm23 has remained largely unresolved. Human Nm23 is present in various isoforms, of which Nm23-H1 and Nm23-H2 are by far the most dominant. Both isoforms are multifunctional enzymes involved in important cellular processes, through their nucleic acid binding ability, their protein-protein interactions and/or their histidine kinase activity. Although Nm23-H1 and Nm23-H2 exhibit 88% sequence homology, they often are considered to have distinct biological functions. Here, we developed an efficient and robust purification protocol to pull-down Nm23 isoforms in their native state. We applied this protocol to purify both overexpressed isoform pure as well as endogenous Nm23 proteins from several human cell lines and mouse brain tissue. Subsequent native mass spectrometry (MS) analysis revealed that all purified Nm23 samples form hexamers, whereby the endogenous protein assembly is primarily present as heterohexamers formed by statistical association of the Nm23-H1 and Nm23-H2 isoforms. Therefore, we conclude that isoform-pure hexameric Nm23 complexes scarcely exist in vivo. We also used native and top-down MS to investigate the histidine autophosphorylation activity of purified Nm23 assemblies. Our data in fine challenge the biological relevance of studying the genes/proteins Nm23-H1 and Nm23-H2 individually.

JLP and centrosomes are essential for the protein transport between the cytoplasm and nucleus induced by external stimuli., JLP belongs to the JIP family whose members serve as scaffolding proteins that link motor proteins and their cargo for intracellular transport. Although JLP is mainly cytoplasmic, it accumulates as a focus in the perinuclear region when stimulated by extracellular stimuli. Focus formation, which changes the nucleus shape and concentrates the nuclear pores, depends on p38MAPK activation and the dynein retrograde motor protein complex. Extracellular stimuli trigger the tethering of PLK1 to the centrosome by JLP, leading to centrosome maturation and microtubule array formation. The centrosome localization domain of JLP is important for the binding of the centrosome and the formation of the JLP focus and the microtubule array. Furthermore, the formation of the JLP focus and the microtubule array is interdependent and important for the transport of NF-κB p65 to the nucleus and its unloading therein. In conclusion, JLP exhibits multiple functions in the nuclear translocation of NF-κB p65.

Online self-reported 24-h dietary recall systems promise increased feasibility of dietary assessment. Comparison against interviewer-led recalls established their convergent validity; however, reliability and criterion-validity information is lacking. The validity of energy intakes (EI) reported using Intake24, an online 24-h recall system, was assessed against concurrent measurement of total energy expenditure (TEE) using doubly labelled water in ninety-eight UK adults (40–65 years). Accuracy and precision of EI were assessed using correlation and Bland–Altman analysis. Test–retest reliability of energy and nutrient intakes was assessed using data from three further UK studies where participants (11–88 years) completed Intake24 at least four times; reliability was assessed using intra-class correlations (ICC). Compared with TEE, participants under-reported EI by 25 % (95 % limits of agreement −73 % to +68 %) in the first recall, 22 % (−61 % to +41 %) for average of first two, and 25 % (−60 % to +28 %) for first three recalls. Correlations between EI and TEE were 0·31 (first), 0·47 (first two) and 0·39 (first three recalls), respectively. ICC for a single recall was 0·35 for EI and ranged from 0·31 for Fe to 0·43 for non-milk extrinsic sugars (NMES). Considering pairs of recalls (first two v. third and fourth recalls), ICC was 0·52 for EI and ranged from 0·37 for fat to 0·63 for NMES. EI reported with Intake24 was moderately correlated with objectively measured TEE and underestimated on average to the same extent as seen with interviewer-led 24-h recalls and estimated weight food diaries. Online 24-h recall systems may offer low-cost, low-burden alternatives for collecting dietary information.

The rapid emergence of antimicrobial resistance is a major threat to human health. Antibiotics modulate a wide range of biological processes in bacteria and as such, the study of bacterial cellular signaling could aid the development of urgently needed new antibiotic agents. Due to the advances in bacterial phosphoproteomics, such a systemwide analysis of bacterial signaling in response to antibiotics has recently become feasible. Here we present a dynamic view of differential protein phosphorylation upon antibiotic treatment and antibiotic resistance. Most strikingly, differential phosphorylation was observed on highly conserved residues of resistance regulating transcription factors, implying a previously unanticipated role of phosphorylation mediated regulation. Using the comprehensive phosphoproteomics data presented here as a resource, future research can now focus on deciphering the precise signaling mechanisms contributing to resistance, eventually leading to alternative strategies to combat antimicrobial resistance.

Here we demonstrate that biomolecular contaminants, such as nucleic acid molecules, can seriously interfere with immobilized metal ion affinity chromatography (IMAC)-based phosphopeptide enrichments. We address and largely solve this issue, developing a robust protocol implementing methanol/chloroform protein precipitation and enzymatic digestion using benzonase, which degrades all forms of DNA and RNA, before IMAC-column loading. This simple procedure resulted in a drastic increase of enrichment sensitivity, enabling the identification of around 17,000 unique phosphopeptides and 12,500 unambiguously localized phosphosites in human cell-lines from a single LC-MS/MS run, constituting a 50% increase when compared with the standard protocol. The improved protocol was also applied to bacterial samples, increasing the number of identified bacterial phosphopeptides even more strikingly, by a factor 10, when compared with the standard protocol. For E. coli we detected around 1300 unambiguously localized phosphosites per LC-MS/MS run. The preparation of these ultra-pure phosphopeptide samples only requires marginal extra costs and sample preparation time and should thus be adoptable by every laboratory active in the field of phosphoproteomics.

c-Myc is a highly pleiotropic transcription factor known to control cell cycle progression, apoptosis, and cellular transformation. Normally, ectopic expression of c-Myc is associated with promoting cell proliferation or triggering cell death via activating p53. However, it is not clear how the levels of c-Myc lead to different cellular responses. Here, we generated a series of stable RPE cell clones expressing c-Myc at different levels, and found that consistent low level of c-Myc induced cellular senescence by activating AP4 in post-confluent RPE cells, while the cells underwent cell death at high level of c-Myc. In addition, high level of c-Myc could override the effect of AP4 on cellular senescence. Further knockdown of AP4 abrogated senescence-like phenotype in cells expressing low level of c-Myc, and accelerated cell death in cells with medium level of c-Myc, indicating that AP4 was required for cellular senescence induced by low level of c-Myc.

c-MYC is an oncogenic transcription factor that is degraded by the proteasome pathway. However, the mechanism that regulates delivery of c-MYC to the proteasome for degradation is not well characterized. Here, the results show that the motor protein complex Kinesin-1 transports c-MYC to the cytoplasm for proteasomal degradation. Inhibition of Kinesin-1 function enhanced ubiquitination of c-MYC and induced aggregation of c-MYC in the cytoplasm. Transport studies showed that the c-MYC aggregates moved from the nucleus to the cytoplasm and KIF5B is responsible for the transport in the cytoplasm. Furthermore, inhibition of the proteasomal degradation process also resulted in an accumulation of c-MYC aggregates in the cytoplasm. Moreover, Kinesin-1 was shown to interact with c-MYC and the proteasome subunit S6a. Inhibition of Kinesin-1 function also reduced c-MYC-dependent transformation activities. Taken together, the results strongly suggest that Kinesin-1 transports c-MYC for proteasomal degradation in the cytoplasm and the proper degradation of c-MYC mediated by Kinesin-1 transport is important for transformation activities of c-MYC. In addition, the results indicate that Kinesin-1 transport mechanism is important for degradation of a number of other proteins as well.

Polypeptide vaccines effectively activate human T cells but suffer from poor biological stability, which confines both transport logistics and in vivo therapeutic activity. Synthetic biology has the potential to address these limitations through the generation of highly stable antigenic “mimics” using subunits that do not exist in the natural world. We developed a platform based on D–amino acid combinatorial chemistry and used this platform to reverse engineer a fully artificial CD8+ T cell agonist that mirrored the immunogenicity profile of a native epitope blueprint from influenza virus. This nonnatural peptide was highly stable in human serum and gastric acid, reflecting an intrinsic resistance to physical and enzymatic degradation. In vitro, the synthetic agonist stimulated and expanded an archetypal repertoire of polyfunctional human influenza virus–specific CD8+ T cells. In vivo, specific responses were elicited in naive humanized mice by subcutaneous vaccination, conferring protection from subsequent lethal influenza challenge. Moreover, the synthetic agonist was immunogenic after oral administration. This proof-of-concept study highlights the power of synthetic biology to expand the horizons of vaccine design and therapeutic delivery.

JLP (JNK-associated leucine zipper protein) is a novel scaffolding protein involved in JNK signaling. Although it is known that JLP is highly expressed in brain, the biological function of JLP in neuronal systems remains unknown. Here, we report a novel interaction between JLP and SCG10 (superior cervical ganglia clone 10), which is a microtubule-destabilizing factor that is essential for neurite outgrowth. Inhibition of endogenous JLP expression using small interference RNA methodology strongly enhanced nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells. Our results show that JLP negatively regulates NGF-induced neurite outgrowth by decreasing the level of phosphorylated SCG10. Furthermore, inhibition of JNK phosphorylation by a small molecule inhibitor, SP600125, resulted in inhibition of SCG10 phosphorylation and inhibition of neurite growth. Taken together, our results suggest that JLP negatively regulates NGF-induced neurite outgrowth through a sequestering mechanism that results in an attenuation of NGF-induced SCG10 phosphorylation.

Mitogen-activated protein kinases (MAPKs) regulate critical signaling pathways involved in cell proliferation, differentiation and apoptosis. Recent studies have shown that a novel class of scaffold proteins mediates the structural and functional organization of the three-tier MAPK module. By linking the MAP3K, MAP2K and MAPK into a multienzyme complex, these MAPK-specific scaffold proteins provide an insulated physical conduit through which signals from the respective MAPK can be transmitted to the appropriate spatiotemporal cellular loci. Scaffold proteins play a determinant role in modulating the signaling strength of their cognate MAPK module by regulating the signal amplitude and duration. The scaffold proteins themselves are finely regulated resulting in dynamic intra- and inter-molecular interactions that can modulate the signaling outputs of MAPK modules. This review focuses on defining the diverse mechanisms by which these scaffold proteins interact with their respective MAPK modules and the role of such interactions in the spatiotemporal organization as well as context-specific signaling of the different MAPK modules.

Although recent reports indicated that produce contamination with foodborne pathogens is widespread in Nigeria, the sources and magnitude of microbial contamination of fruits and vegetables on farms and in markets have not been thoroughly identified. To ascertain possible pathways of contamination, the frequency and magnitude of coliform and Escherichia coli contamination of tomatoes produced in northwest Nigeria was assessed on farms and in markets. Eight hundred twenty-six tomato fruit samples and 36 irrigation water samples were collected and assessed for fecal indicator organisms. In addition, the awareness and use of food safety practices by tomato farmers and marketers were determined. Median concentration of coliforms on all field- and market-sourced tomato fruit samples, as well as in irrigation water sources, in Kaduna, Kano, and Katsina states exceeded 1,000 most probable number (MPN) per g. Median E. coli counts from 73 (17%) of 420 field samples and 231 (57%) of 406 market tomato fruit samples exceeded 100 MPN/g. Median E. coli concentrations on tomato fruits were higher (P0.01) in the rainy season (2.45 Log MPN/g), when irrigation was not practiced than in the dry (1.10 Log MPN/g) and early dry (0.92 Log MPN/g) seasons. Eighteen (50%) of 36 irrigation water samples had E. coli counts higher than 126 MPN/100 ml. Median E. coli contamination on market tomato fruit samples (2.66 Log MPN/g) were higher (P0.001) than those from tomatoes collected on farms (0.92 Log MPN/g). Farmers and marketers were generally unaware of the relationship between food safety practices and microbial contamination on fresh produce. Good agricultural practices pertaining to food safety on farms and in local markets were seldom used. Adoption of food safety practices on-farm, during transport, and during marketing could improve the microbial quality of tomatoes available to the public in this region of the world.

Fluorescent quantum dots (semiconductor nanocrystals) have the potential to revolutionize biological imaging, but their use has been limited by difficulties in obtaining quantum dots that are water soluble and biocompatible. The objectives of our research were to develop a methodology for encapsulation of cadnium-selenium (CdSe) quantum dots (QDs) in phospholipid nanoemulsion that mimics the natural lipoprotein core and to study their interactions with cultured non-small cell lung cancer cells (NSCLC). We found that CdSe QDs can be efficiently encapsulated in the phospholipid nanoemulsion. The QD nanoemulsion has a particle size approximately 80 nm and appears physically stable. The QD nanoemulsion interacts well with cells. The intensity of cellular fluorescence imaging increases with the cell incubation time, indicating more QDs were taken up by the cells, respectively. Two types of fluorescence microscopies confirm that QDs are primarily localized in the cytoplasm but not in the nucleus of the cells.

To localize the regulatory elements in the human follicle-stimulating hormone receptor (FSH-R) promoter/enhancer and to determine the role of upstream stimulatingfactors (USFs) in these elements, we transiently transfected constructs of FSH-R promoter/enhancer in pGL3 luciferase reporter plasmids into Chinese hamster ovary cells and the activities were determined by measuring luciferase luminescence of the cell lysates. The 5'-flanking regions of the human FSH-R gene from nt -1485 to -1 with respect to the gene translation start site were amplified by polymerase chain reaction (PCR) and subcloned in pGL3. Deletion mutants were created using PCR or restriction enzyme digestion. Mutation in the E-box sequence from nt -124 to -119 (E-box 3), in the construct from -224 to nt -1 or in the Inr element, which encompasses the transcriptional start site at nt -99, resulted in a substantial reduction in the human FSH-R promoter/enhancer activity. Overexpression of upstream stimulating factor-1 (USF1) suppresses the activity of the human FSH-R promoter/enhancer via Inr and E-box elements. Upstream stimulating factor-2 (USF2) decreases FSH-R promoter/enhancer activity by acting on E-box 3. The results indicate that E-box 3 and the Inr element are important elements of the human FSH-R promoter/ enhancer. USF family members inhibit FSH-R gene activity by acting via these elements. USF1 and USF2 suppress human FSH-R promoter/enhancer activity by acting on E-box 3. USF1 also decreases activity by interacting with the Inr element.

Computed tomography has become an important component in the initial assessment of severely injured patients over the last years. The liberal use coupled with advances in imaging technology often result in incidental findings. In our present investigation, the prevalence incidence of incidental findings of the spine and skull amounted to 58% of all patients with trauma. Degenerative changes were most commonly found, followed by congenital defects and neoplasms. Within the latter, further investigation was necessary in six cases, of which two findings proved to be malignant neoplasms. The high incidence of incidental findings calls for a uniform documentation, handling and clarification of responsibility in treatment and insurance-related competence.Die Computertomographie hat sich in den vergangenen Jahren zu einem festen Bestandteil der Schockraumdiagnostik entwickelt. Neben den traumatisch bedingten Verletzungen ergeben sich dabei oft unfallunabhängige Befunde, die bis dahin unbekannt waren. In der vorliegenden Arbeit wurde ihre Prävalenz im Bereich des Achsenskelettes mit über 58% beziffert. Am häufigsten waren dabei degenerative Veränderungen, gefolgt von angeborene Fehlbildungen und Neubildungen. Letztgenannte waren in sechs Fällen weiter abklärungsbedürftig. Zwei stellten sich als bösartig dar. Durch die hohe Inzidenz von Zufallsbefunden ergibt sich daher die Notwendigkeit einheitlicher Kriterien zur Dokumentation, zum Umgang mit ihnen, sowie zur Klärung der Zuständigkeit auf behandlungs- sowie versicherungstechnischer Ebene.La tomodensitométrie est devenue un examen important dans la salle d'urgence au cours de ces dernières années. Parallèlement aux blessures causées par accidents, d'autres résultats sont apparus par hasard et sans lien direct avec l'accident. Dans ce travail, l'incidence sur le crâne et la colonne vertèbrale se chiffre à 58%. En majorité sont découverts des changements dégénératifs, suivis par des anomalies congénitales et des néoplasies. Six cas ont nécessité d'être clarifiés et deux cas se sont avérés malins. Suite à la répercution de ces découvertes fortuites, il s'est avéré nécessaire d'établir des critères uniformes afin de clarifier la nécessité d'intervention ainsi que pour des questions d'assurance.

v-myc is the viral homolog of c-myc transduced by several acute transforming retroviruses, many of which encode this gene as a Gag-Myc fusion protein. The v-myc oncogene can transform several lineages of mammalian and avian cells either alone or in cooperation with other oncogenes. While the Gag portion of the Gag-Myc fusion protein and the nuclear localization signal each appear to be dispensable for transformation, the N- and C-termini of the Myc sequence have been found to be essential for transformation. All v-myc genes contain point mutations which seem to confer a greater potency to v-myc in the process of transformation, proliferation, and apoptosis. In v-myc-transformed myelomonocytic cells, secondary events occur, such as the expression of colony stimulating factor-1 (CSF-1) which play a critical role in immortalization and subsequent tumor progression. Inhibition of the autocrine loop of CSF-1 was found to induce apoptosis in the immortalized cells. While overexpression of v-Myc blocks terminal differentiation of hematopoietic cells, this is not sufficient to block the differentiation of certain neural and skeletal muscle cells. Recent developments on the effects of v-myc on cell growth, transformation, differentiation and apoptosis are discussed in this review.

The free-running ruby laser has recently been introduced for removal of unwanted hair growth. It is assumed that the mode of action of ruby laser depilation is that of selective photothermolysis of the melanin-rich structures. The present data reflects our results of ruby treatment of 133 patients attending a dermatological laser clinic for hair removal. When success of the laser treatment was defined as greater than 50% hair removal, 59.0% of patients reported successful results after 90 or more days after last treatment. With success defined as greater than 25% hair removal at 90 days, successful treatment was obtained in 75.0%. Also, the patients evaluated the overall result of their treatments. The percentage of patients who were either "very satisfied" or "satisfied" after 90 days was 64.2%. Only a few side-effects were observed. In general, pain was no clinical problem. No significant scarring was observed. A temporary hypopigmentation was experienced by approximately 10%, but only one patient still had hypopigmentation 90 days after treatment. Due to variability of hair density as well as anagen and telogen phase durations in different anatomical locations, firm conclusions regarding the long-term effect still cannot be drawn.

Tubulin, the major structural component of microtubules, is a target for the development of anticancer agents. A series of (Z)-1-Aryl-3-arylamino-2-propen-1-one (10) were synthesized and evaluated for anti-proliferative activity in cell based assay. The most active compound (Z)-1-(2- bromo-3,4,5-trimethoxyphenyl)-3-(3-hydroxy-4-methoxyphenylamino)-prop-2-en-1-one (10ae) was tested in 20 tumor cell lines including multidrug resistant phenotype and was found to induce apoptosis in all these cell lines with similar GI50 values. Flow cytometry studies showed that 10ae arrested the cells in G2/M phase of cell cycle. In addition to G2/M block, these compounds caused microtubule stabilization like paclitaxel and induced apoptosis via activation of the caspase family. The observations made in this investigation demonstrate that (Z)-1-Aryl-3- arylamino-2-propen-1-one (10) represents a new class of microtubule – stabilizing agents.

Receptor internalization is a common mechanism underlying surface receptor down-regulation (and thus receptor signaling) upon its engagement with the cognate ligand. Tight regulation of surface CD40 expression is critical in regulating different functional properties of dendritic cell (DC). Engagement of CD40 on mature DC and the cognate CD40 ligand on T cell activates c-Jun N-terminal MAPK, p38 and ERK1/2 MAPK pathways in mature DC. JNK-associated leucine zipper protein (JLP) is a scaffolding protein that interacted with p38 and JNK. The molecular mechanism underlying CD40 internalization and its physiological impact on DC functions remained unclear. Here we reported that the engagement of CD40 on the LPS-activated DC down-regulated the surface expression of CD40. We examined the role of the JLP protein in DC differentiation, and in the regulation of DC function(s) in vitro. In contrast to the abundant JLP expression observed in immortal cell lines, primary immature DC expressed low levels of the JLP proteins. The induction of the JLP protein expression was observed in the LPS-mature DC that were activated by CD40 ligation, and also in the poly I:C stimulated DC. JLP-silenced DC was impaired in regulating CD40 surface expression upon LPS stimulation and CD40 induced receptor internalization. Such aberrant change in the regulation of surface CD40 expression was associated with an augmented capacity of the JLP-silenced DC in IL-12 production. Collectively, our data identified a novel role of a scaffolding protein JLP in the regulation of surface CD40 expression and fine-tuning of DC function.

Nursing educators are frequently confronted with challenges that bring about innovation and transition to new ways of transferring knowledge in their home environments. These challenges are magnified when approached from an international perspective. Optimal implementation of knowledge transfer incorporates choosing models that promote local initiatives in line with increasingly decentralized educational structures. These decentralized models are a means to foster ongoing participation for both educators and students in their own professional development. Innovative education stems from creativity in approaching the need with formats and activities to meet a specific challenge. This experimental study builds upon previous study by the authors which was conducted in March, 2009, based upon the qualitative open focus forum at each of the five nursing programs. Overwhelmingly, the Cameroonian nursing students expressed a keen desire to study the HIV infected pregnant woman and the feeding options of the newborn. The study team developed the train-the-trainer program which was delivered at the University of Buea in the Southwest region of Cameroon in March, 2011. TTT is particularly effective for reaching large audiences and also permits a degree of sustainability such that the Cameroonian students will be trainers for subsequent cohorts of their peers. This study continues to strengthen the collaborative endeavors between the two nursing schools; the University of Buea (UB) and Goldfarb School of Nursing (GSON) at Barnes Jewish College in Saint Louis, Missouri, USA. The final aim of the intervention was the initiation of collaborative relationships between the faculty members of the two educational organizations.Key words: Workshop, HIV, AIDS, capacity building, education, intervention, Cameroon

Anchorage of segmental replacement prostheses in diaphyseal bone remains a challenge in lower limb reconstructions. We developed and studied a new prosthesis design that features an intramedullary anchorage system for which finite element analysis predicted favorable bone remodeling. We retrospectively analyzed the cases of all patients who underwent implantation of the new stem. Their data were prospectively collected. Twenty-four patients (25 prosthetic reconstructions using diaphyseal fixation of the prosthesis) had 18 primary implantations and 7 revision cases. At a mean follow- up of 61 months, TESS (Toronto Extremity Salvage Score) and MSTS (Musculoskeletal Tumor Society Rating Scale score) were 80% and 65% that of a normal extremity, respectively. SF-36 (36-Item Short-Form Health Survey) Mental and Physical scores were 54 and 44 points, respectively. Minimum follow-up was 31 months (mean, 61 months; range, 31-107 months). Radiographic evaluation (1991 International Symposium of Limb Salvage [ISOLS] Radiological Implant Evaluation System) revealed 65% excellent and 35% good bone remodeling around the implant as a whole, 65% excellent and 35% good results for the anchorage proper, and 70% excellent and 30% good findings for lucencies at the bone-metal interface. Two patients (1 traumatic event) developed a loose stem. The results support the expectations as shown by finite element analysis - that the risk for loosening is reduced and that favorable bone remodeling occurs around the stem over time.

International audience; AIM: To evaluate our practice following Clinical Practice Guidelines (CPG) of the French College of Obstetricians and Gynecologists (CNGOF) in 2005 advocating a policy of restrictive episiotomy and to show that a significative decrease in the rate of episiotomy does not increase the number of third and fourth degree perineal tears. MATERIAL AND METHODS: A retrospective study of episiotomies and third/fourth degree perineal tears of the year 2003 (before the CPG) was compared with the year 2007 (after the CPG). We analyzed the indications of episiotomies and compared the rate of episiotomies and severe perineal tears between the two periods. RESULTS: In 2003, the rate of episiotomies was 18.8% (upon 1755 vaginal deliveries). We observed 16 (9 per thousand) third-degree perineal tears, five of which was associated with episiotomies; and two (1 per thousand) fourth-degree perineal tears. In 2007, the rate of episiotomies was 3.4% (upon 1940 vaginal deliveries). There were eight (4 per thousand) third-degree and four (2 per thousand) fourth-degree perineal tears. The two periods of study were similar in terms of age, parity, gestational age, birthweight, rate of spontaneous deliveries, breech and instrumental deliveries. There were a difference regarding deliveries in the occipitoposterior position (5.8% vs 13.8% ; p=0.02). No significant difference was found between the rates of third degree (9 per thousand vs 4 per thousand ; p=0.059) and fourth degree perineal tears (1 per thousand vs 2 per thousand ; p=0.487). However, there was a significant decrease in the rate of episiotomies between the two periods (18.8% vs 3.4% ; p

Chemokine receptors control several fundamental cellular processes in both hematopoietic and structural cells, including directed cell movement, i.e., chemotaxis, cell differentiation, and proliferation. We have previously demonstrated that CXCR3, the chemokine receptor expressed by Th1/Tc1 inflammatory cells present in the lung, is also expressed by human airway epithelial cells. In airway epithelial cells, activation of CXCR3 induces airway epithelial cell movement and proliferation, processes that underlie lung repair. The present study examined the expression and function of CXCR3 in human alveolar type II pneumocytes, whose destruction causes emphysema. CXCR3 was present in human fetal and adult type II pneumocytes as assessed by immunocytochemistry, immunohistochemistry, and Western blotting. CXCR3-A and -B splice variant mRNA was present constitutively in cultured type II cells, but levels of CXCR3-B greatly exceeded CXCR3-A mRNA. In cultured type II cells, I-TAC, IP-10, and Mig induced chemotaxis. Overexpression of CXCR3-A in the A549 pneumocyte cell line produced robust chemotactic responses to I-TAC and IP-10. In contrast, I-TAC did not induce chemotactic responses in CXCR3-B and mock-transfected cells. Finally, I-TAC increased cytosolic Ca2+and activated the extracellular signal-regulated kinase, p38, and phosphatidylinositol 3-kinase (PI 3-kinase)/protein kinase B kinases only in CXCR3-A-transfected cells. These data indicate that the CXCR3 receptor is expressed by human type II pneumocytes, and the CXCR3-A splice variant mediates chemotactic responses possibly through Ca2+activation of both mitogen-activated protein kinase and PI 3-kinase signaling pathways. Expression of CXCR3 in alveolar epithelial cells may be important in pneumocyte repair from injury.

Stem cells represent a unique population of cells with self-renewal capacity. However, the molecular control of self-renewal and differentiation of stem cells has remained enigmatic. Here, we show that short-type PB-cadherin (STPB-C) promoted self-renewal of spermatogonial stem cells (SSCs) via activating Janus kinase/signal transducer and activator of transcription (JAK-STAT) and phosphoinositide-3 kinase (PI3-K)/Akt, and blocking transforming growth factor (TGF)-beta1 signaling. These data were obtained with varied approaches, including the use of RNA interference (RNAi), SSC cultures infected by STPB-C retroviral vector, bromodeoxyuridine (BrdU) incorporation assay, and other techniques. These findings have important implications for germ cell biology and create the possibility of using SSCs for biotechnology and medicine. They are also critical in understanding tissue homeostasis, the aging process, tumor formation and degenerative diseases.

International audience; Extracellular signals are transduced into cells through mitogen-activated protein kinases (MAPKs), which are activated by their upstream kinases. Recently, families of scaffolding proteins have been identified to tether specific combinations of these kinases along specific signaling pathways. Here we describe a protein, JLP (c-Jun NH2-terminal kinase-associated leucine zipper protein), which acts as a scaffolding protein to bring together Max and c-Myc along with JNK (c-Jun NH2-terminal kinase) and p38MAPK, as well as their upstream kinases MKK4 (MAPK kinase 4) and MEKK3 (MAPK kinase kinase 3). Thus, JLP defines a family of scaffolding proteins that bring MAPKs and their target transcription factors together for the execution of specific signaling pathways.

Apoptosis Associated Tyrosine Kinase (AATYK), a novel protein recently isolated from differentiating 32D mouse myeloid cells, contains a putative tyrosine kinase domain and several binding motifs for src homology 2 (SH-2) and src homology 3 (SH-3) domain containing proteins. We observed that AATYK is expressed in different regions of the brain. Although it might play a role in normal nervous system development by modulating apoptosis, little is known regarding its function in the brain or its intracellular localization and kinase activity. Recognizing its homology with Insulin like growth factor-I (IGF-I) receptor (IGF-IR) and the critical role of IGF-I in neuronal survival, we hypothesized that AATYK plays an important role in neuronal differentiation/apoptosis. To test this hypothesis, we transfected the human adrenergic neuroblastoma (NB):SH-SY5Y cells with AATYK cDNA under a tetracycline-repressible promoter and established stable cell lines that readily express AATYK on removal of tetracycline. AATYK immunoprecipitated from these cell lysates is an active kinase. Indirect immunofluorescent staining of the clones revealed AATYK to be localized in the cytoplasm. By itself, AATYK overexpression for short duration (2–3 days) did not induce differentiation in the stable SH-SY5Y clones. On the other hand, overexpression for longer periods (7–8 days) per se, significantly (P

Ultrasonography of a 26-week-old fetus suggested the presence of a goiter. Amniocentesis was performed at 26 and 38 weeks gestation to evaluate thyroid function. An elevated amniotic fluid TSH level was found on both occasions, though amniotic fluid T4, T3, and rT3 levels were similar to those of euthyroid fetuses. The diagnosis of goitrous hypothyroidism was confirmed at birth. Therapy with thyroid hormone was instituted on the first day of life. Growth and development of the infant have been normal during the first year of his life. We suggest that the measurement of amniotic fluid TSH can be useful in the diagnosis of intrauterine primary hypothyroidism.

The sonogram of a patient with an intrauterine gestation revealed a partial posterior placenta praevia and a hydatidiform mole within the praevia portion of the placenta.

Exome sequencing has identified the causes of several Mendelian diseases, although it has rarely been used in a clinical setting to diagnose the genetic cause of an idiopathic disorder in a single patient. We performed exome sequencing on a pedigree with several members affected with attention deficit/hyperactivity disorder (ADHD), in an effort to identify candidate variants predisposing to this complex disease. While we did identify some rare variants that might predispose to ADHD, we have not yet proven the causality for any of them. However, over the course of the study, one subject was discovered to have idiopathic hemolytic anemia (IHA), which was suspected to be genetic in origin. Analysis of this subject's exome readily identified two rare non-synonymous mutations in PKLR gene as the most likely cause of the IHA, although these two mutations had not been documented before in a single individual. We further confirmed the deficiency by functional biochemical testing, consistent with a diagnosis of red blood cell pyruvate kinase deficiency. Our study implies that exome and genome sequencing will certainly reveal additional rare variation causative for even well-studied classical Mendelian diseases, while also revealing variants that might play a role in complex diseases. Furthermore, our study has clinical and ethical implications for exome and genome sequencing in a research setting; how to handle unrelated findings of clinical significance, in the context of originally planned complex disease research, remains a largely uncharted area for clinicians and researchers.

CD31, a trans-homophilic inhibitory receptor expressed on both T- and B-lymphocytes, drives the mutual detachment of interacting leukocytes. Intriguingly, T cell CD31 molecules relocate to the immunological synapse (IS), where the T and B cells establish a stable interaction. Here, we show that intact CD31 molecules, which are able to drive an inhibitory signal, are concentrated at the periphery of the IS but are excluded from the center of the IS. At this site, were the cells establish the closest contact, the CD31 molecules are cleaved, and most of the extracellular portion of the protein, including the trans-homophilic binding sites, is shed from the cell surface. T cells lacking CD31 trans-homophilic binding sites easily establish stable interactions with B cells; at the opposite, CD31 signaling agonists inhibit T/B IS formation as well as the ensuing helper T cell activation and function. Confocal microscopy and flow cytometry analysis of experimental T/B IS shows that the T cell inhibitory effects of CD31 agonists depend on SHP-2 signaling, which reduces the phosphorylation of ZAP70. The analysis of synovial tissue biopsies from patients affected by rheumatoid arthritis showed that T cell CD31 molecules are excluded from the center of the T/B cell synapses in vivo. Interestingly, the administration of CD31 agonists in vivo significantly attenuated the development of the clinical signs of collagen-induced arthritis in DBA1/J mice. Altogether, our data indicate that the T cell co-inhibitory receptor CD31 prevents the formation of functional T/B immunological synapses and that therapeutic strategies aimed at sustaining CD31 signaling will attenuate the development of autoimmune responses in vivo.

Background— The atheromodulating activity of B cells during the development of atherosclerosis is well documented, but the mechanisms by which these cells are regulated have not been investigated. Methods and Results— Here, we analyzed the contribution of Qa-1–restricted CD8 + regulatory T cells to the control of the T follicular helper–germinal center B-cell axis during atherogenesis. Genetic disruption of CD8 + regulatory T cell function in atherosclerosis-prone apolipoprotein E knockout mice resulted in overactivation of this axis in secondary lymphoid organs, led to the increased development of tertiary lymphoid organs in the aorta, and enhanced disease development. In contrast, restoring control of the T follicular helper–germinal center B-cell axis by blocking the ICOS-ICOSL pathway reduced the development of atherosclerosis and the formation of tertiary lymphoid organs. Moreover, analyses of human atherosclerotic aneurysmal arteries by flow cytometry, gene expression analysis, and immunofluorescence confirmed the presence of T follicular helper cells within tertiary lymphoid organs. Conclusions— This study is the first to demonstrate that the T follicular helper–germinal center B-cell axis is proatherogenic and that CD8 + regulatory T cells control the germinal center reaction in both secondary and tertiary lymphoid organs. Therefore, disrupting this axis represents an innovative therapeutic approach.