Immunization with glycosylated Kb-binding peptides generates carbohydrate-specific,unrestricted cytotoxic T cells

Cytotoxic T cells (CTL) recognize target proteins as short peptides presented by major histocompatibility complex (MHC) class I restriction elements. However, there is also evidence for peptide-independent T cell receptor (TCR) recognition of target proteins and non-protein structures. How such T cell responses are generated is presently unclear. We generated carbohydrate (CHO)-specific, MHC-unrestricted CTL responses by coupling di- and trisaccharides to K^b- or D^b-binding peptides for direct immunization in mice. Four peptides and three CHO have been analyzed with the CHO either in terminal or central positions on the carrier peptide. With two of these glycopeptides, with galabiose (Galα1-4Gal; Gal₂) bound to a homocysteine (via an ethylene spacer arm) in position 4 or 6 in a vesicular stomatitis virus nucleoprotein-derived peptide (RGYVYQGL binding to K^b), CTL were generated which preferentially killed target cells treated with glycopeptide compared to those treated with the core peptide. Polyclonal CTL were also found to kill target cells expressing the same Gal₂ epitope in a glycolipid. By fractionation of CTL, preliminary data indicate that glycopeptide-specific K^b-restricted CTL and unrestricted CHO-specific CTL belong to different T cell populations with regard to TCR expression. The results demonstrate that hapten-specific unrestricted CTL responses can be generated with MHC class I-binding carrier peptides. Different models that might explain the generation of such responses are discussed.     

Sleep-wake rhythm in an irregular shift system

Sleep in shift work has been studied extensively in regular shift systems but to a lesser degree in irregular shifts. Our main aim was to examine the sleep-wake rhythm in shift combinations ending with the night or the morning shift in two irregular shift systems. Three weeks' sleep/work shift diary data, collected from 126 randomly selected train drivers and 104 traffic controllers, were used in statistical analyses including a linear mixed model and a generalized linear model for repeated measurements. The results showed that the sleep-wake rhythm was significantly affected by the shift combinations. The main sleep period before the first night shift shortened by about 2 h when the morning shift immediately preceded the night shift as compared with the combination containing at least 36 h of free time before the night shift (reference combination). The main sleep period before the night shift was most curtailed between two night shifts, on average by 2.9 and 3.5 h among the drivers and the controllers, respectively, as compared with the reference combination. Afternoon napping increased when the morning or the day shift immediately preceded the night shift, the odds being 4.35-4.84 in comparison with the reference combination. The main sleep period before the morning shift became 0.5 h shorter when the evening shift preceded the morning shift in comparison with the sleep period after a free day. The risk for dozing off during the shift was associated only with the shift length, increasing by 17 and 35% for each working hour in the morning and the night shift, respectively. The results demonstrate advantageous and disadvantageous shift combinations in relation to sleep and make it possible to improve the ergonomy of irregular shift systems.     

Norrie disease caused by a gene deletion allowing carrier detection and prenatal diagnosis

Carrier determination and prenatal diagnosis in Norrie disease (ND) has so far not been reported. We describe a kindred with 4 members affected by ND in which a deletion comprising gene locus DXS7 on the short arm of the X chromosome defined by probe L1.28 causes the disorder. This allowed us to predict via chorion villus biopsy that a male foetus of a carrier woman is unaffected.     

Gene profiling reveals increased expression of uteroglobin and other anti-inflammatory genes in glucocorticoid-treated nasal polyps

BACKGROUND: Treatment with local glucocorticoids (GCs) decreases symptoms and the size of nasal polyps. This might depend on the downregulation of proinflammatory genes, as well as the upregulation of anti-inflammatory genes. OBJECTIVE: We sought to identify GC-regulated anti-inflammatory genes in nasal polyps. METHODS: Affymetrix DNA microarrays were used to analyze the expression of 22,283 genes in 4 nasal polyps before and after local treatment with fluticasone (400 microg/d). Expression of uteroglobin and mammaglobin B was analyzed with real-time PCR in 6 nasal polyps and in nasal biopsy specimens from 6 healthy control subjects. RESULTS: Two hundred three genes had changed in expression in treated polyps, and 139 had known functions: 54 genes were downregulated, and 85 were upregulated. Genes associated with inflammation constituted the largest single functional group. These genes affected key steps in inflammation (eg, immunoglobulin production; antigen processing and presentation; and the chemoattraction and activation of granulocytes, T cells, and B cells). Several proinflammatory genes were downregulated. In contrast, some anti-inflammatory genes were upregulated. The gene that increased most in terms of expression was uteroglobin. This was confirmed with real-time PCR. By contrast, expression of uteroglobin was lower in untreated polyps than in healthy nasal mucosa. Immunohistochemical investigation showed staining of uteroglobin in the epithelium and in seromucous glands in control subjects and in nasal polyps. CONCLUSION: Upregulation of anti-inflammatory genes, such as uteroglobin, might contribute to the effects of local treatment with GCs in nasal polyps.     

Autoimmune diseases in myelodysplastic syndrome favors patients survival: A case control study and literature review

Background

We conducted a monocentric retrospective study of patients with myelodysplastic syndromes (MDS) and autoimmune or inflammatory disorders (AIMs) and a literature review. We analyzed the association with subgroups of the WHO 2016 MDS classification and patient's survival in a case control study. Risk factors associated with survival were analyzed by uni- and multivariate analysis.

Autonomic Nervous System Function and Essential Hypertension: Individual Response Specificity With and Without Beta-Adrenergic Blockade

The aim of the present research was to study individual response specificity in 22 male patients having essential hypertension (HT) and to compare these patients with age-matched male normotensive controls (NT). Four stimuli, letter identification, mental arithmetic, cold pressor and isometric exercise, were administered while recordings were made of: systolic and diastolic blood pressures, heart rate, respiration, forearm and hand blood flows, and skin conductance level and fluctuations. After each session urine samples were collected and epinephrine and norepinephrine levels were analyzed. Twelve subjects in the HT group were given beta-adrenergic blocking agents and retested 1 to 21 months (X?= 12 months) after the first session. Each response was standardized, using NT as the reference group. Intraclass correlations were computed to evaluate whether HT males reacted with a more consistent hierarchy of responses than did NT. Intraclass correlations were significantly higher among the patients than in the control group, regardless of whether the blood pressure response was included or excluded in the computation of the intraclass correlations. Thus, we conclude that male HT patients show more individual response specificity than NT controls. Beta-adrenergic receptor antagonists reduced levels of cardiovascular activity and attenuated reactivity but did not affect amount of specificity. Thus, intraclass correlations provide unique and useful information, since they are not related to blood pressure reactivity or to urinary catecholamine levels, nor affected by beta-adrenergic blockade.     

Web-based virtual microscopy in teaching and standardizing Gleason grading

Gleason grading forms the basis of prognostic and therapeutic assessment in prostatic carcinoma despite its subjective nature and substantial interobserver variation. The accuracy of Gleason grading can be improved by the use of educational tools such as reference images. However, conventional microscopy images are of limited educational value because it is neither possible to view the sample at different magnifications nor to navigate into different areas of the specimen. This limitation can be overcome by the use of virtual microscopy, which allows viewing entire digitized microscope slides. We created an interactive Web site ( www.webmicroscope.net/gleason ) featuring a comprehensive set of prostatic needle biopsies as virtual slides, which can be viewed with a standard Web browser (Internet Explorer or Netscape). To evaluate the validity of Web-based virtual microscopy for Gleason grading, an experienced uropathologist (TK) scored a series of 62 biopsies from the original glass slides and 6 weeks later from virtual slides on the Web site using an ordinary desktop computer. The intraobserver agreement was excellent, with identical Gleason scores found in 48 of the 62 cases ( kappa = 0.73). The 14 remaining scores differed only by 1 point on the Gleason scale (2-10). The virtual slides were viewed by 2 other uropathologists (PM and HH), with interobserver kappa coefficients ranging from 0.55 to 0.62, which is within the range of previously reported studies using glass slides. The 3 uropathologists' Gleason scores were included as reference scores on the Web site, which now serves as a publicly open platform for self-testing and learning of Gleason grading. We conclude that Web-based virtual microscopy is a promising new tool for teaching and standardizing Gleason grading.     

A homeo domain protein reveals the metameric nature of the developing chick hindbrain

The segmented embryonic hindbrain of vertebrates develops by sequential constriction of the neural tube into eight metameric units known as rhombomeres. The cellular and molecular basis of this segmentation process is largely unknown. Using an antibody, we analyzed the expression pattern of the chick homeo domain-containing protein Ghox-lab in the developing chick hindbrain. At the neural plate stage, prior to the appearance of rhombomeres, Ghox-lab is expressed within a single domain that extends anteriorly up to the site where rhombomere 4 will later form. After rhombomere 4 has appeared and as hindbrain segmentation progresses, the level of Ghox-lab protein increases significantly within the fourth rhombomere. This intensification, accompanied by the elimination of Ghox-lab protein in rhombomeres 5 and 6, eventually results in the formation of a distinct island of expression in rhombomere 4. All cells in the newly formed rhombomere 4 express Ghox-lab, except for the cells of the floor plate. In addition, neural crest cells migrating from the fourth rhombomere are also Ghox-lab-positive. These data raise the possibility that Ghox-lab protein might be one of the factors involved in the specification of the metameric pattern of the vertebrate hindbrain.     

Use of O-antigen gene cluster-specific PCRs for the identification and O-genotyping of Yersinia pseudotuberculosis and Yersinia pestis

Yersinia pestis is a very recently evolved clone of Yersinia pseudotuberculosis serotype O:1b. This close relationship causes potential difficulties in DNA-based diagnostic methods. Analysis of the O-antigen gene clusters in these two organisms identified two regions that were used to specifically identify Y. pestis-Y. pseudotuberculosis as a group or Y. pestis alone. Both PCR assays were found to be 100% specific when tested on a large collection of Yersinia species and other Enterobacteriaceae. Furthermore, advantage was taken of the different setups of the O-antigen gene clusters of the 21 known Y. pseudotuberculosis serotypes to develop a multiplex PCR assay to replace the conventional serotyping method of Y. pseudotuberculosis by O-genotyping. The multiplex PCR assay contained nine sets of specific PCRs in a single tube and when used on Y. pseudotuberculosis reference strains allowed the distinction of 14 individual serotypes and two duplex serotypes (O:4a-O:8 and O:12-O:13). Serotype O:7, O:9, and O:10 strains required additional PCRs for O-genotyping. Once applied to Y. pseudotuberculosis strains of various origins, a very good correlation between classical serotypes and O-genotypes was observed, although some discrepancies were found. O-genotyping also proved useful to correct misidentification of some strains and to type Y. pseudotuberculosis isolates that had lost the expression of the O-antigen. The PCR-based O-genotyping can easily be applied in conventional laboratories, without the need for tedious preparation of a large set of specific antisera.     

Role of YadA, Ail, and Lipopolysaccharide in Serum Resistance of Yersinia enterocolitica Serotype O:3

Complement attack is a host strategy leading to elimination of pathogens. Yersinia enterocolitica expresses several potential complement resistance factors: the outer membrane proteins YadA and Ail as well as lipopolysaccharide (LPS). To study the contribution of these factors to the survival of Y. enterocolitica serotype O:3 in nonimmune human serum, we constructed 23 mutant strains of Y. enterocolitica O:3 expressing different combinations of YadA, Ail, LPS O antigen, and LPS outer core. Survival of bacteria was analyzed in normal serum (with functional classical, lectin, and alternative complement activation pathways) and EGTA-Mg-treated serum (only alternative pathway functional). Kinetic killing tests revealed that the most potent single-serum resistance factor needed for long-term survival was YadA; Ail was also indispensable, but it provided short-term survival and delayed the bacterial killing. On the contrary, the LPS O antigen and outer core, when in combination with YadA, Ail, or both, had a minor and often negative effect on serum resistance. Bacteria in the exponential phase of growth were more resistant to serum killing than stationary-phase bacteria. After exposing bacteria to EGTA-Mg-treated serum, O antigen could prevent deposition of covalently bound C3b on bacteria at 3 min of incubation, even as a single factor. At later time points (15 and 30 min) it had to be accompanied by YadA, Ail, and outer core. In normal serum, the bacteria were less resistant to C3b deposition. However, no direct correlation between the C3 deposition pattern and bacterial resistance was observed.