Association of insomnia with mania in Lebanese patients with bipolar disorder.

OBJECTIVE: To assess: (1) the association between insomnia experienced at admission, sociodemographic and other patients' characteristics and mania; and (2) the variation of insomnia and mania before and after treatment in bipolar patients with manic episodes (type I). METHODS: Sixty-two patients were interviewed shortly after their admission to the hospital (after 3 to 5 days). The current symptoms experienced by the patients were assessed upon admission and again at discharge from the hospital. RESULTS: A poorer quality of sleep (higher PSQI scores) (Beta=0.590) was significantly associated with higher mania, whereas the intake of SSRIs (Beta=-5.952) and TCAs (Beta=-8.181) was significantly associated with lower mania. Furthermore, highly significant reductions were reported in the PSQI scores (4.96 vs. 2.75, P<0.001), ISI scores (8.30 vs. 3.45, P<0.001) and YMRS scores (8.60 vs. 3.06, P<0.001) between admission to and discharge from the hospital. CONCLUSION: Insomnia in patients with bipolar disorder type I is associated with mania, with a significant reduction of sleep problems seen during a period of approximately 20 days of hospitalization. Further longitudinal studies are needed to confirm the validity of our results and identify the causes. In the meantime, this research recommends a strategy to improve sleeplessness experienced during inter-episode phases may be helpful in preventing manic episodes in BD.

Transcriptomic profiling of trophoblast fusion using BeWo and JEG-3 cell lines.

In human placenta, alteration in trophoblast differentiation has a major impact on placental maintenance and integrity. However, little is known about the mechanisms that control cytotrophoblast fusion. The BeWo cell line is used to study placental function, since it forms syncytium and secretes hormones after treatment with cAMP or forskolin. In contrast, the JEG-3 cell line fails to undergo substantial fusion. Therefore, BeWo and JEG-3 cells were used to identify a set of genes responsible for trophoblast fusion. Cells were treated with forskolin for 48 h to induce fusion. RNA was extracted, hybridised to Affymetrix HuGene ST1.0 arrays and analysed using system biology. Trophoblast differentiation was evaluated by real-time PCR and immunocytochemistry analysis. Moreover, some of the identified genes were validated by real-time PCR and their functional capacity was demonstrated by western blot using phospho-specific antibodies and CRISPR/cas9 knockdown experiments. Our results identified a list of 32 altered genes in fused BeWo cells compared to JEG-3 cells after forskolin treatment. Among these genes, four were validated by RT-PCR, including salt-inducible kinase 1 (SIK1) gene which is specifically upregulated in BeWo cells upon fusion and activated after 2 min with forskolin. Moreover, silencing of SIK1 completely abolished the fusion. Finally, SIK1 was shown to be at the center of many biological and functional processes, suggesting that it might play a role in trophoblast differentiation. In conclusion, this study identified new target genes implicated in trophoblast fusion. More studies are required to investigate the role of these genes in some placental pathology.

Identification and characterization of HIV positive Ethiopian elite controllers in both Africa and Israel.

OBJECTIVES: HIV elite controllers (ECs) are a unique subgroup of HIV-positive patients who are long-term virologically suppressed in the absence of antiretroviral treatment (ART). The prevalence of this subgroup is estimated to be < 1%. Various cohorts of ECs have been described in developed countries, most of which have been demographically heterogeneous. The aim of this study was to identify ECs in two large African cohorts and to estimate their prevalence in a relatively genetically homogenous population. METHODS: We screened two cohorts of HIV-positive Ethiopian patients. The first cohort resided in Mekelle, Ethiopia. The second was comprised of HIV-positive Ethiopian immigrants in Israel. In the Mekelle cohort, ART-naïve subjects with stable CD4 counts were prospectively screened using two measurements of viral load 6 months apart. Subjects were defined as ECs when both measurements were undetectable. In the Israeli cohort, subjects with consistently undetectable viral loads (mean of 17 viral load measurements/patient) and stable CD4 count > 500 cells/µL were defined as ECs. RESULTS: In the Mekelle cohort, 16 of 9515 patients (0.16%) fitted the definition of EC, whereas seven of 1160 (0.6%) in the Israeli cohort were identified as ECs (P = 0.011). CONCLUSIONS: This is the first large-scale screening for HIV-positive ECs to be performed in entirely African cohorts. The overall prevalence of ECs is within the range of that previously described in developing countries. The significant difference in prevalence between the two cohorts of similar genetic background is probably a consequence of selection bias but warrants further investigation into possible environmental factors which may underlie the EC state.

E2A protein degradation by the ubiquitin-proteasome system is stage-dependent during muscle differentiation.

The E2A proteins are basic helix-loop-helix transcription factors that regulate proliferation and differentiation in many cell types. In muscle cells, the E2A proteins form heterodimers with muscle regulatory factors such as MyoD, which then bind to DNA and regulate the transcription of target genes essential for muscle differentiation. We now demonstrate that E2A proteins are primarily localized in the nucleus in both C2C12 myoblasts and myotubes, and are degraded by the ubiquitin proteasome system evidenced by stabilization following treatment with the proteasome inhibitor, MG132. During the differentiation from myoblast to myotube, the cellular abundance of E2A proteins is relatively unaltered, despite significant changes (each approximately 5-fold) in the relative rates of protein synthesis and protein degradation via the ubiquitin-proteasome system. The rate of ubiquitin-proteasome-mediated E2A protein degradation depends on the myogenic differentiation state (t 1/2 approximately 2 h in proliferating myoblasts versus t 1/2 > 10 h in differentiated myotubes), and is also associated with cell cycle in non-muscle cells. Our findings reveal an important role for both translational and post-translational regulatory mechanisms in mediating the complex program of muscle differentiation determined by the E2A proteins.

[Veterinary medicine of the GDR in strained relations between technical instructions and political guidelines]. / Die Veterinärmedizin der DDR im Spannungsfeld von Fachauftrag und gesellschaftspolitischer Steuerung.

The development of Veterinary Medicine in the Soviet Occupation Zone in Germany and the former German Democratic Republic (GDR) is sketched in highlights. After the collectivization of agriculture (1960) a centralistically controlled national veterinary system was established. It was suited to the requirements of the industrially organized animal production. The successive classification of socialistic veterinary administration was associated with the following matters: the extension of veterinary subject matters, a vertical division of work with the aid of newly created technical veterinary professions, and a penetration of the profession with political guidelines. As the professional level of the veterinary system in the GDR was relatively high the reflection in retrospective needs to be evaluated in a differentiated way considering the textual and social conditions. In spite of centralism and indoctrination the veterinary system remained professionally autonomous with islands of political independence, which sustained the identity of this profession. The latter formed the base for self renewal of the East German veterinary system at the end of the socialistic area 1989/1990.

The nuclear ubiquitin-proteasome system degrades MyoD.

Many short-lived nuclear proteins are targeted for degradation by the ubiquitin-proteasome pathway. The role of the nucleus in regulating the turnover of these proteins is not well defined, although many components of the ubiquitin-proteasome system are localized in the nucleus. We have used nucleoplasm from highly purified HeLa nuclei to examine the degradation of a physiological substrate of the ubiquitin-proteasome system (MyoD). In vitro studies using inhibitors of the system demonstrate MyoD is degraded via the ubiquitin-proteasome pathway in HeLa nucleoplasm. Purified nucleoplasm in vitro also supports the generation of high molecular mass MyoD-ubiquitin adducts. In addition, in vivo studies, using leptomycin B to inhibit nuclear export, demonstrate that MyoD is degraded in HeLa cells by the nuclear ubiquitin-proteasome system.

Comparison of systematic versus selective screening for methicillin-resistant Staphylococcus aureus carriage in a high-risk dermatology ward.

OBJECTIVE: To compare two strategies for screening methicillin-resistant Staphylococcus aureus (MRSA) carriers in a high-risk dermatology ward: systematic screening of all admitted patients versus selective screening of patients at risk. DESIGN: The two strategies were applied prospectively during two consecutive periods. In period A (8.5 months), only patients transferred from other wards, or with a history of prior hospitalization, or presenting chronic wounds or disease with denuded skin were considered at high risk of MRSA carriage and sampled. In period B (7.5 months), all admitted patients were systematically screened. End-points were the number of patients having a MRSA-positive screening sample on admission during period B and having none of the risk factors used in period A, the rate of imported MRSA cases, and the rate of acquired cases. SETTING: A 1,032-bed university hospital with a 19-bed inpatient dermatology ward, a referral center for toxic epidermal necrolysis and severe extensive dermatoses. PATIENTS: The study included 729 dermatology inpatients (370 in period A and 359 in period B). RESULTS: During period A, screening samples were obtained on admission for 30% of patients (77% of the patients at risk) and identified 25 MRSA carriers. During period B, 90.5% of admitted patients were screened, and 26 MRSA carriers were detected on admission; all of these patients belonged to at least one predefined category at risk for carriage. Overall rates of imported and acquired cases were similar between the two periods (6.8% vs 7.5%, and 2.9% vs 2.4%, respectively). CONCLUSIONS: A screening strategy targeted to patients at risk of harboring MRSA has similar sensitivity and is more cost-effective than a strategy of systematic screening to identify MRSA carriers on admission.

The carboxyl-terminal domain of receptor-associated protein facilitates proper folding and trafficking of the very low density lipoprotein receptor by interaction with the three amino-terminal ligand-binding repeats of the receptor.

The 39-kDa receptor-associated protein (RAP) is a specialized antagonist that inhibits all known ligand interactions with receptors that belong to the low density lipoprotein (LDL) receptor gene family. Recent studies have demonstrated a role for RAP as a molecular chaperone for the LDL receptor-related protein during receptor folding and trafficking within the early secretory pathway. In the present study, we investigated a potential role for RAP as a chaperone for the very low density lipoprotein (VLDL) receptor, another member of the LDL receptor gene family. Using intracellular cross-linking techniques, we found that RAP is associated with newly synthesized VLDL receptor. In the absence of RAP co-expression, newly synthesized VLDL receptor exhibited slower trafficking along the early secretory pathway, most likely due to misfolding of the receptor. The role of RAP in the folding of the VLDL receptor was further studied using an anchor-free, soluble VLDL receptor. Metabolic pulse-chase labeling experiments showed that while only 3% of the soluble VLDL receptor was folded and secreted in the absence of RAP co-expression, over 50% of the soluble receptor was secreted in the presence of RAP co-expression. The functions of RAP in VLDL receptor folding and trafficking were mediated by its carboxyl-terminal repeat but not by the amino-terminal and central repeats. Using truncated VLDL receptor constructs, we identified the RAP-binding site within the first three ligand-binding repeats of the VLDL receptor. Thus, our present study demonstrates that RAP serves as a folding and trafficking chaperone for the VLDL receptor via interactions of its carboxyl-terminal repeat with the three amino-terminal ligand-binding repeats of the VLDL receptor.

[Pilotropic T-cell lymphoma without mucinosis: 5 new cases]. / Lymphomes T pilotropes sans mucinose: 5 nouveaux cas.

BACKGROUND: Pilotropic cutaneous T-cell lymphomas without mucinosis are rare, with 27 cases previously reported. Diagnosis and classification may be difficult. The clinical course and histopathological and immunohistochemical findings in 5 patients are described. PATIENTS AND METHODS: Patients were selected from the register of the French Study Group for cutaneous lymphomas. The criteria for inclusion were clinical pilofollicular manifestations and histological features of pilotropic T-cell lymphoma without mucinosis. RESULTS: Five patients were selected. The most frequent clinical manifestations were follicular keratosis, alopecia and follicular papules. Typical lesions of mycosis fongoides were present for several years in 3 patients, and lymphomatoid papulosis preexisted in one patient. Histopathological analysis showed an infiltrate composed of CD3+ and CD4+ atypical lymphocytes involving the follicular epithelium with alteration of the hair follicle walls. Epidermotropism was associated with pilotropism and situated near the follicular lesions or farther apart. Alcian blue stains results were negative in all specimens. PCR studies showed the presence of a T-cell clone in the skin lesions in all cases. COMMENTS: Diagnosis of pilotropic cutaneous T cell lymphomas without mucinosis may be difficult in case of discrete epidermotropism, minimal infiltrate or involvement of the follicular epithelium. Pilotropism could define a particular variant of T-cell lymphomas.

Identification of a region within the ubiquitin-activating enzyme required for nuclear targeting and phosphorylation.

The ubiquitin-activating enzyme exists as two isoforms: E1a, localized predominantly in the nucleus, and E1b, localized in the cytoplasm. Previously we generated hemagglutinin (HA) epitope-tagged cDNA constructs, HA1-E1 (epitope tag placed after the first methionine) and HA2-E1 (epitope tag placed after the second methionine) (Handley-Gearhart, P. M., Stephen, A. G., Trausch-Azar, J. S., Ciechanover, A., and Schwartz, A. L. (1994) J. Biol. Chem. 269, 33171-33178), which represent the native isoforms. HA1-E1 is exclusively nuclear, whereas HA2-E1 is found predominantly in the cytoplasm. Using high resolution isoelectric focusing and SDS-polyacrylamide gel electrophoresis, we confirm that these epitope-tagged constructs HA1-E1 and HA2-E1 represent the two isoforms E1a and E1b. HA1-E1/E1a exists as one non-phosphorylated and four phosphorylated forms, and HA2-E1/E1b exists as one predominant non-phosphorylated form and two minor phosphorylated forms. We demonstrate that the first 11 amino acids are essential for phosphorylation and exclusive nuclear localization of HA1-E1. Within this region are four serine residues and a putative nuclear localization sequence (NLS; 5PLSKKRR). Removal of these four serine residues reduced phosphorylation levels by 60% but had no effect on nuclear localization of HA1-E1. Each serine residue was independently mutated to an alanine and analyzed by two-dimensional electrophoresis; only serine 4 was phosphorylated. Disruption of the basic amino acids within the NLS resulted in loss of exclusive nuclear localization and a 90-95% decrease in the phosphorylation of HA1-E1. This putative NLS was able to confer nuclear import on a non-nuclear protein in digitonin-permeabilized cells in a temperature- and ATP-dependent manner. Thus the predominant requirement for efficient phosphorylation of HA1-E1/E1a is a functional NLS, suggesting that E1a may be phosphorylated within the nucleus.

[Foreign bodies of the respiratory tract in children. A retrospective study of 100 cases]. / Corps étrangers des voies respiratoires chez l'enfant. Etude rétrospective de 100 cas.

Foreign body inhalation is a universal problem, most common cause of death from domestic accident in children aged five years and less. Over 15 years, one hundred children aged less than six years were evaluated in Hotel Dieu de France; findings are comparable to previous data, with one avoidable death; boys are chiefly concerned (64%); mean age is 22.5 months; circumstances are often hazy (65%); vegetables are prominently responsible (90%) especially peanuts and pistachios (48%); foreign bodies are seldom radiopaque (1%); autumnal predominance is noted. Inhalation is not reported in 25% of cases; immediate risk is subglottic impaction; the child survives if the foreign body is expelled one way or another. The most frequent site is the bronchial system (87%) with a slight right predominance (49%); symptoms include: dyspnea, persistent cough, and, in case of bronchial obstruction: wheezing and asymmetry of breath sounds; 15% of children are free of symptoms. Delay before hospital care is long (21.5 days), mostly because diagnosis is misread particularly in case of bronchial foreign body; pulmonary distension is a frequent finding (45%). In case of asphyxia, first aid resuscitation is performed immediately: in fact it is rarely useful, sometimes harmful. Extraction is mandatory with the stiff bronchoscope; otherwise, bronchopulmonary infection and destruction is the usual outcome ... (25%). Management is revisited, and prevention is recalled.

The ubiquitin-activating enzyme E1 is phosphorylated and localized to the nucleus in a cell cycle-dependent manner.

The ubiquitin-activating enzyme E1 exists as two isoforms, E1a (117 kDa) and E1b (110 kDa). E1a is phosphorylated, whereas E1b is not. In the present study we have demonstrated the cell cycle dependence of E1a phosphorylation: a 2-fold increase in the specific phosphorylation of E1a in G2 compared with the basal level of phosphorylation in the other stages of the cell cycle. Two-dimensional gel electrophoresis resolved E1 into the two isoforms E1a and E1b; E1a resolved further as three phosphorylated forms and one nonphosphorylated form, while E1b resolved as one nonphosphorylated form. E1a is found predominantly in the phosphorylated forms. However, the distribution of E1a among these different phosphorylated forms was not cell cycle-dependent. We next evaluated the enzymatic activity of E1 as well as its subcellular localization throughout the cell cycle. 32P-Pyrophosphate exchange activity of E1 did not vary along the cell cycle; however, the amount of ubiquitin-protein conjugates decreased by 50% in G2. Nuclear and cytosolic fractionation of cells revealed the nuclear to cytosolic ratio of phosphorylated E1a was 3-fold greater in G2 compared with the other stages of the cell cycle. Finally, purified nuclear extracts supported E1-dependent ubiquitin conjugation of exogenous substrates as did purified cytosol. However, in nuclear extracts but not in cytosol the amount of E1 activity was rate-limiting. Thus we establish nuclear E1-dependent protein ubiquitination and propose that an increase in phosphorylation of E1a in G2 functions to increase the import and/or retention of E1a in the nucleus and may modulate nuclear protein ubiquitination.

Rescue of the complex temperature-sensitive phenotype of Chinese hamster ovary E36ts20 cells by expression of the human ubiquitin-activating enzyme cDNA.

The ubiquitin conjugation system is a multi-step pathway in which ubiquitin is activated and conjugated to acceptor proteins, one function of which is to target acceptor proteins for rapid degradation within the cell. The conjugation system is involved in many aspects of cellular functions, including the cell cycle. Several cell-cycle arrest mutant cell lines have been characterized and appear to harbour a mutant ubiquitin-activating enzyme, E1, as their primary defect. One such cell line is ts20, which is derived from Chinese hamster ovary E36 cells. This cell line has been used to characterize some of the potential functions of the ubiquitin conjugation system in vivo, such as its involvement in the maturation of autophagic vacuoles. The present study describes the complete rescue of the complex ts20 phenotype following the expression of the cDNA for human E1. Stable transfectants expressing the human E1 cDNA in the CMVneo expression vector were measured for ubiquitin-conjugation activity, protein degradation and growth in culture at the nonpermissive temperature. This rescue confirms that the phenotype observed in the ts20 cells is due to a defect in the E1 enzyme. Thus, the ts20 cell line will serve as a useful tool to delineate the functions of the ubiquitin system in vivo.

Human ubiquitin-activating enzyme, E1. Indication of potential nuclear and cytoplasmic subpopulations using epitope-tagged cDNA constructs.

The ubiquitin-activating enzyme E1 catalyzes the first step in the ubiquitin conjugation pathway. Previously, we have cloned and sequenced the cDNA for human E1. Expression of the E1 cDNA in the ts20 cell line, which harbors a thermolabile E1, abrogated the phenotypic defects associated with this line. However, little is known of the cell biology of the E1 protein or the nature of the E1 doublet. Thus, we constructed epitope-tagged E1 cDNAs in which the HA monoclonal antibody epitope tag sequence (from influenza hemagglutinin and recognized by the 12CA5 monoclonal antibody) was fused to the amino terminus of E1. Because the amino-terminal amino acid sequence of E1 is unknown, three constructs were made in which the HA tag was placed at each of the first three ATGs in the open reading frame (HA-1E1, HA-2E1, and HA-3E1). Western analysis of HeLa cells transfected with the constructs revealed that HA-1E1 closely comigrated with the upper band of the E1 doublet, and HA-2E1 comigrated with the lower band of the E1 doublet; HA-3E1 appeared smaller than either of the E1 bands. Metabolic labeling with 32P and immunoprecipitation with anti-HA antibody revealed that only the HA-1E1 protein product is phosphorylated; polyclonal anti-E1 antibody showed that only the upper band of the endogenous E1 doublet is phosphorylated. Each of the constructs was able to rescue the mutant phenotype of the ts20 cell line. Immunofluorescence studies showed that HA-2E1 and HA-3E1 were distributed in the cytoplasm with both negative and positive nuclei. This pattern of distribution has also been observed when immunostaining with a monoclonal antibody to E1 (1C5). However, the staining pattern associated with a polyclonal anti-E1 antibody (JJJ) is characterized by positive staining cytoplasm and nuclei in all cells. The HA-1E1 construct exhibited apparently exclusive nuclear distribution in HeLa cells. The difference between the staining patterns of the polyclonal and monoclonal anti-E1 antibodies can be explained by the existence of two subpopulations of E1: one cytoplasmic and partially nuclear, and one that is nuclear. Deletion of a small region at the amino terminus of the HA-1E1, including the basic sequence KKRR, transformed its immunostaining pattern to that observed with HA-2E1.

Nuclear localization of the ubiquitin-activating enzyme, E1, is cell-cycle-dependent.

The mechanisms that regulate ubiquitin-mediated degradation of proteins such as the mitotic cyclins at defined stages of the cell cycle are poorly understood. The initial step in the conjugation of ubiquitin to substrate proteins involves the activation of ubiquitin by the ubiquitin-activating enzyme, E1. Previously we have described the subcellular localization of this enzyme to both nuclear and cytoplasmic compartments. In the present study, we have used the 1C5 anti-E1 monoclonal antibody in immunofluorescent-microscopy and subcellular-fractionation techniques to examine the distribution of E1 during the HeLa cell cycle. E1 is both cytoskeletal and nuclear during the G1-phase. As the cells progress into S-phase, E1 is exclusively cytoskeletal and has a perinuclear distribution. During G2-phase, E1 reappears in the nucleus before breakdown of the nuclear envelope. In mitotic cells, E1 localizes to both the mitotic spindle and the cytosol, but is absent from the chromosomes. Immunoblot analysis reveals multiple forms of E1 in HeLa whole cell extract. This heterogeneity is not a result of polyubiquitination and may represent inactive pools of E1. Only the characteristic E1 doublet is able to activate ubiquitin. Cell-fractionation studies reveal a differential distribution of specific E1 isoforms throughout the cell cycle. Therefore we propose that the subcellular localization of E1 may play a role in regulating cell-cycle-dependent conjugation of ubiquitin to target proteins.