The role of lipocalins in airway disease.

The pathogenesis of allergic airway inflammation and disease is complex and still not fully understood. Many cells, factors and mediators are involved in the different aspects of induction, maintenance and persistence of airway inflammation. The heterogeneity and redundancy of this system is one of the main reasons why novel therapeutic targets focusing on the pathogenesis of asthma only hesitantly reach the market and clinical application. Thus, it seems mandatory that we proceed in our efforts to better understand this micro cosmos to succeed in the development of safe and effective drugs for the treatment of more severe and refractory forms of asthma and chronic obstructive pulmonary disease. One of the more recently discovered mediators in the context of airway inflammation are the lipocalins (Lcns). They are a family of proteins that share functional and structural similarities and are involved in the transport of small hydrophobic molecules such as steroids and lipids into the cell. Lcns are found in many different cell types from plants and bacteria through invertebrate cells to cells of vertebrate origin. The purpose of this review is to summarize the role of Lcns in airway diseases, focusing on allergic and infectious inflammation. In particular, we will summarize the present knowledge about Lipocalin 1 and Lipocalin 2, where exciting new discoveries in the recent years have highlighted their role in pulmonary disease and infection. This new class of proteins is another putative candidate for the development of novel drugs against airway inflammation.

Lipocalin2 protects against airway inflammation and hyperresponsiveness in a murine model of allergic airway disease.

BACKGROUND: Allergen-induced bronchial asthma is a chronic airway disease that involves the interplay of various genes with environmental factors triggering different inflammatory pathways. OBJECTIVE: The aim of this study was to identify possible mediators of airway inflammation (AI) in a model of allergic AI via microarray comparisons and to analyse one of these mediators, Lipocalin2 (Lcn2), for its role in a murine model of allergic airway disease. METHODS: Gene microarrays were used to identify genes with at least a twofold increase in gene expression in the lungs of two separate mouse strains with high and low allergic susceptibility, respectively. Validation of mRNA data was obtained by Western blotting, followed by functional analysis of one of the identified genes, Lcn2, in mice with targeted disruption of specific gene expression. Epithelial cell cultures were undertaken to define induction requirements and possible mechanistic basis of the results observed in the Lcn2 knock-out mice. RESULTS: Lcn2 was up-regulated upon allergen sensitization and airway challenges in lung tissues of both mouse strains and retraced on the protein level in bronchoalveolar lavage fluids. Functional relevance was assessed in mice genetically deficient for Lcn2, which showed enhanced airway resistance and increased AI associated with decreased apoptosis of lung inflammatory cells, compared with wild-type controls. Similarly, application of Lcn2-blocking antibodies before airway challenges resulted in increased inflammation and reduced apoptosis. CONCLUSION: These data indicate a protective role for Lcn2 in allergic airway disease, suggesting a pro-apoptotic effect as the underlying mechanism.

Glutathione peroxidase-2 protects from allergen-induced airway inflammation in mice.

The aim of the present study was to identify and validate the biological significance of new genes/proteins involved in the development of allergic airway disease in a murine asthma model. Gene microarrays were used to identify genes with at least a two-fold increase in gene expression in lungs of two separate mouse strains with high and low allergic susceptibility. Validation of mRNA data was obtained by western blotting and immunohistochemistry, followed by functional analysis of one of the identified genes in mice with targeted disruption of specific gene expression. Expression of two antioxidant enzymes, glutathione peroxidase-2 (GPX2) and glutathione S-transferase omega (GSTO) 1-1 was increased in both mouse strains after induction of allergic airway disease and localised in lung epithelial cells. Mice with targeted disruption of the Gpx-2 gene showed significantly enhanced airway inflammation compared to sensitised and challenged wild-type mice. Our data indicate that genes encoding the antioxidants GPX2 and GSTO 1-1 are common inflammatory genes expressed upon induction of allergic airway inflammation, and independently of allergic susceptibility. Furthermore, we provide evidence to illustrate the importance of a single antioxidant enzyme, GPX2, in protection from allergen-induced disease.

Tolerance of ambiguity: text analytic vs self-report measures in two nonclinical groups.

Avoidance of ambiguity can be examined by both text analytic and self-report methods. In previous studies using text analytic methods, clinical groups showed a higher avoidance of ambiguity than nonclinical subjects. In nonclinical subjects, however, higher avoidance of ambiguity did not correlate with emotional processes. In these studies, higher avoidance of ambiguity was assessed by a text analytic method (DoTA), which was applied to the Holtzman Inkblot Technique. However, the Inkblot Technique may not activate those cognitive-affective structures in nonclinical subjects required to elicit higher avoidance of ambiguity. Thus, the following discrepant results can be predicted: in nonclinical subjects, DoTA indicators of higher avoidance of ambiguity based on Holtzman Inkblot Technique do not show correlations with self-report measures of higher avoidance of ambiguity such as the Ambiguity Tolerance Questionnaire, the Inventory for the Measurement of Tolerance of Ambiguity (Reis inventory), or self-report measures of related traits of personality such as the Giessen Test. Self-report measures of higher avoidance of ambiguity should show such correlations. Two studies were carried out to test these hypotheses. In Study 1, 80 nonclinical subjects (48 women, M age = 34.5 yr.) were examined using the DoTA text analytic method, the Ambiguity Tolerance Questionnaire-14 and the Giessen Test. In Study 2, 82 nonclinical subjects (43 women, M age = 34.0 yr.) were tested using the Reis inventory. The results obtained in these subjects are consistent with the hypothe-

Depressive symptoms in advanced cancer. Part 1. Assessing depression: the Mood Evaluation Questionnaire.

OBJECTIVE: A prospective cohort study evaluated the performance and acceptability of the cognitive based Mood Evaluation Questionnaire (MEQ) in assessing depressive symptoms over time in patients with advanced cancer. The structured clinical interview for DSM-III-R (SCID) and the single-item interview screening question were also administered. METHOD: Forty-five advanced cancer patients were followed monthly for up to six interviews (108 interviews in total). RESULTS: The MEQ and SCID had moderate agreement (weighted kappa 0.52 over all interviews). At first interview, 26 (58%) patients were depressed using MEQ, seven (16%) of these severely. Attrition rates were high for subsequent interviews. The assessment process was considered positive by 92% patients, and acceptable as a single and repeat measure. Positive responses to MEQ questions about worthlessness, death and self-harm were strong predictors of current depressed mood. CONCLUSION: The MEQ has potential as a useful alternative to SCID in assessing depressive symptoms in a palliative care population, and can be used by nonpsychiatric trained professionals. It is positively received by patients and acceptable as a repeat measure. Most patients appreciate being asked about their mood in depth, including questions around death and self harm.

Depressive symptoms in advanced cancer. Part 2. Depression over time; the role of the palliative care professional.

OBJECTIVE: To survey changes in depressive symptoms over time in patients with advanced cancer using the Mood Evaluation Questionnaire (MEQ), and explore how effective Palliative Care Professionals (PCPs) are at assessing this. METHOD: Forty-five advanced cancer patients were followed monthly for up to six interviews (108 interviews in total). Selected clinical variables were recorded. RESULTS: At first interview, 26 (58%) patients were depressed using MEQ, seven (16%) severely. Attrition rates were high, with 28, 13, nine, seven and six patients available for subsequent interviews. A > 12-point increase in MEQ over sequential interviews was associated with reduced survival (Hazard ratio 3.2, CI 1.2-8.4). PCPs recognized depressed mood, but underdiagnosed severe depression in 9% of patients (CI 3-16%, P = 0.002). A past history of depression was a strong indicator of current depressed mood on the MEQ. CONCLUSION: Change in depressive symptoms over time is related to mortality in this patient cohort, patients with marked deterioration in mood dying earlier. PCPs usually recognize patients' depression but underestimate the severity of depressed mood.

Interactions between Spc2p and other components of the endoplasmic reticulum translocation sites of the yeast Saccharomyces cerevisiae.

In yeast, the endoplasmic reticulum membrane proteins Sec11p and Spc3p are essential for the cleavage of signal peptides of nascent polypeptide chains during their passage through translocation sites. Genetic and biochemical experiments demonstrate that Sec11p and Spc3p are tightly associated with two other proteins, Spc1p and Spc2p, whose functions are largely unknown. Using anti-Spc2p antibodies, we show here that this heterotetrameric complex associates with Sbh1p and Sbh2p, the beta-subunits of the Sec61p complex and the Ssh1p complex, respectively. Depletion of Spc2p decreased the enzymatic activity of the SPC in vitro, led to a loss of Spc1p, and led to a down-regulation of the amount of Sec11p and Spc3p in the endoplasmic reticulum. Moreover, the deletion of Spc2p also decreased the expression level of Sbh2p. These data implicate that Spc2p not only enhances the enzymatic activity of the SPC but also facilitates the interactions between different components of the translocation site.

Mammalian Sec61 is associated with Sec62 and Sec63.

In yeast, efficient protein transport across the endoplasmic reticulum (ER) membrane may occur co-translationally or post-translationally. The latter process is mediated by a membrane protein complex that consists of the Sec61p complex and the Sec62p-Sec63p subcomplex. In contrast, in mammalian cells protein translocation is almost exclusively co-translational. This transport depends on the Sec61 complex, which is homologous to the yeast Sec61p complex and has been identified in mammals as a ribosome-bound pore-forming membrane protein complex. We report here the existence of ribosome-free mammalian Sec61 complexes that associate with two ubiquitous proteins of the ER membrane. According to primary sequence analysis both proteins display homology to the yeast proteins Sec62p and Sec63p and are therefore named Sec62 and Sec63, respectively. The probable function of the mammalian Sec61-Sec62-Sec63 complex is discussed with respect to its abundance in ER membranes, which, in contrast to yeast ER membranes, apparently lack efficient post-translational translocation activity.

Mood influences on automatic and controlled semantic priming.

Mood effects on automatic and controlled components of semantic priming were examined. After a neutral or positive mood manipulation, subjects underwent two blocks of a priming task. Automatic and strategic priming processes were operationally differentiated by the assumption that automatic priming remains constant during the task, whereas controlled priming develops slowly during the course of the experiment. In the first block the priming effect was greater for happy than for neutral mood participants. In the second block the priming effect was greater for neutral mood than for happy participants. The results confirm the hypothesis of a mood-dependent processing change: During early trials priming is based on automatic processing facilitated by positive mood. This processing mode is later superseded by controlled processes aided by neutral mood.

The yeast SPC22/23 homolog Spc3p is essential for signal peptidase activity.

In eucaryotic cells signal sequences of secretory and membrane proteins are cleaved by the signal peptidase complex during their transport into the lumen of the endoplasmic reticulum. The signal peptidase complex in yeast consists of four subunits. To date, three of these subunits have been functionally characterized. One of them, the Sec11p, is essential for viability of yeast cells. It shows significant homology to the mammalian SPC18 and SPC21 as well as to bacterial leader peptidases. Two other subunits, Spc1p and Spc2p, have been shown to be homologous to mammalian SPC12 and SPC25, respectively, and are not essential for protein translocation or signal peptide cleavage. We have purified and analyzed the fourth subunit of yeast signal peptidase, Spc3p. The protein is essential for viability of yeast cells. Depletion of SPC3 leads to accumulation of precursors of secretory proteins in vivo and to the loss of the signal peptidase activity in vitro. Therefore, in contrast to the bacterial leader peptidases, yeast signal peptidase requires a second subunit for its function.

Structurally related Spc1p and Spc2p of yeast signal peptidase complex are functionally distinct.

Two subunits of the mammalian signal peptidase complex, SPC12 and SPC25, share similar membrane topologies with the majority of each protein oriented toward the cytoplasm. Such similarities may suggest that these proteins perform redundant functions in signal peptidase activity. In the present study, we addressed this issue through analysis of the yeast homologs to SPC12 and SPC25, Spc1p and Spc2p. We show that both Spc1p and Spc2p are nonessential for signal peptidase activity and growth of yeast cells and that null mutations in the genes encoding Spc1p and Spc2p are synthetically lethal with a conditional mutation affecting Sec11p, an essential subunit of yeast signal peptidase. However, a high copy plasmid encoding Spc1p suppresses the conditional sec11 mutation, whereas the corresponding plasmid encoding Spc2p does not suppress sec11. Moreover, Spc2p, but not Spc1p, is important for signal peptidase activity and cell viability at high temperatures. These results indicate that although both Spc1p and Spc2p are noncatalytic, they are functionally distinct. Evidence is also presented that a double mutant lacking Spc1p and Spc2p grows well relative to wild type yeast cells, indicating that the signal peptidase complex missing at least two of its subunits is sufficient for signal peptidase activity in vivo.

Effects of antisense DNA against the alpha-sarcin stem-loop structure of the ribosomal 23S rRNA.

Antisense DNAs complementary against various sequences of the alpha-sarcin domain (C2646-G2674) of 23S rRNA from Escherichia coli were hybridized to naked 23S rRNA as well as to 70S ribosomes. Saturation levels of up to 0.4 per 70S ribosome were found, the identical fraction was susceptible to the attack of the RNase alpha-sarcin. The hybridization was specific as demonstrated with RNase H digestion, sequencing the resulting fragments and blockage of the action of alpha-sarcin. The RNase alpha-sarcin seems to approach its cleavage site from the 3' half of the loop of the alpha-sarcin domain. Hybridization is efficiently achieved at 37 degrees C and can extend at least into the 3' strand of the stem of the alpha-sarcin domain. However, the inhibition of alpha-sarcin activity is observed at 30 degrees C but not at 37 degrees C. For a significant inhibition of poly(Phe) synthesis the temperature had to be lowered to 25 degrees C. The results imply that the alpha-sarcin domain changes its conformation during protein synthesis and that the conformational changes may include a melting of the stem of the alpha-sarcin domain.

[Reducing ambiguity: semantic statistical studies of "normal" probands, neurotic patients, borderline patients and schizophrenic patients]. / Reduzierung von Ambiguität: sprachstatistische Untersuchungen an "Normalen", Neurotikern, Borderline-Patienten und Schizophrenen.

The present study tries to answer three questions: 1.) Do patients with neurotic disorders differ from normals by a stronger tendency to avoid or reduce ambiguity? 2.) Does the tendency to avoid or reduce ambiguity increase with increasing ambiguity of the stimulus? 3.) Does the avoidance or reduction of ambiguity increase with affects of anxiety and hostility? In order to answer these questions, Ertel's dogmatism-dictionary was applied to the answers of 30 normals, 30 patients with neurotic disorders, 30 borderline-patients, 25 acute and 25 chronic schizophrenics in the Holtzman Inkblot Technique (HIT). According to the results, (1) patients with neurotic disorders do not differ from normals by a stronger tendency to avoid or reduce ambiguity. 2.) The tendency to avoid or reduce ambiguity increases with measures of increasing stimulus ambiguity of the HIT cards in all diagnostic groups studied with the exception of chronic schizophrenics. As far as response ambiguity (variability of interpretation) is concerned, only in chronic schizophrenics the tendency to avoid or reduce ambiguity decreases with increasing response ambiguity. 3.) The avoidance or reduction of ambiguity increases with affects of anxiety and/or aggression assessed by HIT-measures in all diagnostic groups studied with the exception of normals and chronic schizophrenics. In both normals and chronic schizophrenics, the reduction of ambiguity decreases significantly with increasing anxiety, in chronic schizophrenics the reduction of ambiguity decreases significantly with an increase of low levels of aggression.

[Cognitive style of schizophrenic patients: ambiguity avoidance and decreased abstract thinking]. / Kognitiver Stil bei Schizophrenen: Ambiguitäts-Vermeidung und Verminderte Abstraktheit.

The hypothesis of specific cognitive dynamics in schizophrenics is derived and tested in this study. This cognitive style should be characterized by a reduction of cognitive ambiguity and could be demonstrated in borderline patients in a previous study. This hypothesis is tested by applying Ertel's (1972) DOTA-dictionary to responses of 25 acute and 25 chronic schizophrenics in the Holtzman Inkblot Technique. Furthermore the level of abstractness of the responses is investigated with the method of Günther and Groeben (1978). According to the results both groups of schizophrenics had a significantly higher "Dogmatismus-Quotient" (Ertel, 1972) than a sample of neurotics, but did not differ from borderline patients. Both acute and chronic schizophrenics used significantly less abstract nouns than borderline patients and neurotics. In contrast to borderline patients the reduction of cognitive ambiguity in schizophrenics is not connected with strong affects of anxiety and hostility.

[Hepatic amebiasis in the Kilimanjaro region. Serodiagnosis on micro-specimens of dried blood and attempts at treatment with tinidazole (fasigyn)]. / Amibiase hépatique dans la région du Kilimanjaro. Séro-diagnostics sur micro-prélévements de sang séché et essais de traitement par le tinidazole (fasigyn)

Amoebic dysentery appears to be rare in the northeast of Tanzania. Hepatic amoebiasis, on the other hand, is apparently widespread since at least 200 cases are seen every year at the Kilimanjaro Christian Medical Centre. This incidence of cases enabled us to carry out trials on the spot with a new imidazole derivative, Tinidazole. Formerly the difficult diagnosis based on clinical symptoms had to be buttressed by radiological evidence and possibly by the result of puncture. Indirect fluorescent antibody tests for the diagnosis of amoebiasis were performed elsewhere on all the patients, using for this purpose microspecimens of dried blood. In 12 cases out of 34 an agglutination test with sensitized latex particles was performed on the spot. This latter test has the practical advantage of being easy to employ. It cannot, however, be considered as a screening test since it is subject to downward and upward errors. The indirect fluorescent antibody test has been found to be constantly and highly positive, certain antibody titres attaining 1/6400. This fully confirms the value of the method even under special working conditions. Seventeen of our 34 patients (2 women and 15 men ranging in age from 20 to 75 years) were treated with 2 g of Tinidazole per day in a single dose for 2 to 3 consecutive days. Puncture to evacuate pus was also performed where abscesses had collected. Tolerance on the whole was good without a single sign of cardiovascular or urinary toxicity. However, paraesthesia of the hands was observed in one case, transitory thrombocytopenia in one other patient, and increased alkaline phosphatases. Minor disorders were also observed in our series of patients: mild vertigo (7 cases), headache (6 cases), and dry mouth (2 cases). After 8 months the therapeutic results were as follows: 12 complete cures out of 17, 2 improvements with final cure probable, 3 partial failures necessitating supplementary treatment with Metronidazole (2.4 g per day for 2 days). These preliminary trials appear to the encouraging and the study is being continued with series compared with cases treated with Emetine or Metronidazole.