Focused differential glycan analysis with the platform antibody-assisted lectin profiling for glycan-related biomarker verification

Protein glycosylation is a critical subject attracting increasing attention in the field of proteomics as it is expected to play a key role in the investigation of histological and diagnostic biomarkers. In this context, an enormous number of glycoproteins have now been nominated as disease-related biomarkers. However, there is no appropriate strategy in the current proteome platform to qualify such marker candidate molecules, which relates their specific expression to particular diseases. Here, we present a new practical system for focused differential glycan analysis in terms of antibody-assisted lectin profiling (ALP). In the developed procedure, (i) a target protein is enriched from clinic samples (e.g. tissue extracts, cell supernatants, or sera) by immunoprecipitation with a specific antibody recognizing a core protein moiety; (ii) the target glycoprotein is quantified by immunoblotting using the same antibody used in (i); and (iii) glycosylation difference is analyzed by means of antibody-overlay lectin microarray, an application technique of an emerging glycan profiling microarray. As model glycoproteins having either N-linked or O-linked glycans, prostate-specific antigen or podoplanin, respectively, were subjected to systematic ALP analysis. As a result, specific signals corresponding to the target glycoprotein glycans were obtained at a sub-picomole level with the aid of specific antibodies, whereby disease-specific or tissue-specific glycosylation changes could be observed in a rapid, reproducible, and high-throughput manner. Thus, the established system should provide a powerful pipeline in support of on-going efforts in glyco-biomarker discovery.     

Attention to Eye Contact in the West and East: Autonomic Responses and Evaluative Ratings

Eye contact has a fundamental role in human social interaction. The special appearance of the human eye (i.e., white sclera contrasted with a coloured iris) implies the importance of detecting another person''s face through eye contact. Empirical studies have demonstrated that faces making eye contact are detected quickly and processed preferentially (i.e., the eye contact effect). Such sensitivity to eye contact seems to be innate and universal among humans; however, several studies suggest that cultural norms affect eye contact behaviours. For example, Japanese individuals exhibit less eye contact than do individuals from Western European or North American cultures. However, how culture modulates eye contact behaviour is unclear. The present study investigated cultural differences in autonomic correlates of attentional orienting (i.e., heart rate) and looking time. Additionally, we examined evaluative ratings of eye contact with another real person, displaying an emotionally neutral expression, between participants from Western European (Finnish) and East Asian (Japanese) cultures. Our results showed that eye contact elicited stronger heart rate deceleration responses (i.e., attentional orienting), shorter looking times, and higher ratings of subjective feelings of arousal as compared to averted gaze in both cultures. Instead, cultural differences in the eye contact effect were observed in various evaluative responses regarding the stimulus faces (e.g., facial emotion, approachability etc.). The rating results suggest that individuals from an East Asian culture perceive another''s face as being angrier, unapproachable, and unpleasant when making eye contact as compared to individuals from a Western European culture. The rating results also revealed that gaze direction (direct vs. averted) could influence perceptions about another person''s facial affect and disposition. These results suggest that cultural differences in eye contact behaviour emerge from differential display rules and cultural norms, as opposed to culture affecting eye contact behaviour directly at the physiological level.     

Mineral Metabolism Markers Are Associated with Myocardial Infarction and Hemorrhagic Stroke but Not Ischemic Stroke in Hemodialysis Patients: A Longitudinal Study

Background/Aims

The associations between phosphate, calcium, and intact parathyroid hormone (PTH) levels and composite cardiovascular end points have been studied. This study examined the associations of these markers with myocardial infarction (MI) and stroke separately.

Methods

This is a longitudinal study on 65,849 hemodialysis patients from the Japan Renal Data Registry. Patients with prior events at baseline were excluded. Predictors were phosphate, albumin-corrected calcium, intact PTH, and calcium times phosphate product levels. Outcome was the first episode of MI or stroke during a 1-year observation period. Data were analyzed using multiple logistic regression analyses, adjusted for potential confounders.

Results

There were 1,048, 651, and 2,089 events of incident MI, hemorrhagic, and ischemic stroke, respectively. Incident MI was associated with phosphate levels ≥6.5 mg/dL (odds ratio 1.49; confidence interval 1.23–1.80) compared with phosphate levels of 4.7–5.4 mg/dL and intact PTH levels>500 pg/mL (1.35; 1.03–1.79) compared with intact PTH levels of 151–300 pg/mL. Higher albumin-corrected calcium level was positively associated with MI (p = 0.04 by trend analysis). Hemorrhagic stroke was associated only with intact PTH levels>500 pg/mL (1.54; 1.10–2.17). Incident ischemic stroke had no association with phosphate, calcium, or intact PTH levels. The association of calcium times phosphate product with outcomes was essentially the same pattern as that of phosphate and outcomes.

Conclusions

MI was associated with phosphate, calcium, and intact PTH levels, whereas hemorrhagic stroke was associated only with intact PTH. Ischemic stroke was not associated with any of them. The potential distinct beneficial effect on MI and stroke by managing bone and mineral disease should be investigated in future studies.     

Predicted expansion of the claudin multigene family

Claudins (Cldn) are essential membrane proteins of tight junctions (TJs), which form the paracellular permselective barrier. They are produced by a multi-gene family of 24 reported members in mouse and human. Based on a comprehensive search combined with phylogenetic analyses, we identified three novel claudins (claudin-25, -26, and -27). Quantitative RT-PCR revealed that the three novel claudins were expressed in a tissue- and/or developmental stage-dependent manner. Claudins-25 and -26, but not claudin-27, were immunofluorescently localized to TJs when exogenously expressed in cultured MDCK and Eph epithelial cell lines. These findings expand the claudin family to include at least 27 members.     

Insulin substrates 1 and 2 are corequired for activation of atypical protein kinase C and Cbl-dependent phosphatidylinositol 3-kinase during insulin action in immortalized brown adipocytes

Phosphatidylinositol 3-kinase (PI3K)-dependent activation of atypical protein kinase C (aPKC) is required for insulin-stimulated glucose transport. Although insulin receptor substrate-1 (IRS-1) and IRS-2, among other factors, activate PI3K, there is little information on the relative roles of IRS-1and IRS-2 during aPKC activation by insulin action in specific cell types. Presently, we have used immortalized brown adipocytes in which either IRS-1 or IRS-2 has been knocked out by recombinant methods to examine IRS-1 and IRS-2 requirements for activation of aPKC. We have also used these adipocytes to see if IRS-1 and IRS-2 are required for activation of Cbl, which is required for insulin-stimulated glucose transport and has been found to function upstream of both PI3K/aPKC and Crk during thiazolidinedione action in 3T3/L1 adipocytes [Miura et al. (2003) Biochemistry 42, 14335]. In brown adipocytes in which either IRS-1 or IRS-2 was knocked out, insulin-induced increases in aPKC activity and glucose transport were markedly diminished. These effects of insulin on aPKC and glucose transport were fully restored by retroviral-mediated expression of IRS-1 or IRS-2 in their respective knockout cells. Knockout of IRS-1 or IRS-2 also inhibited insulin-induced increases in Cbl binding to the p85 subunit of PI3K, which, along with IRS-1/2, may be required for activation of PI3K, aPKC, and glucose transport during insulin action in 3T3/L1 adipocytes. These findings provide evidence that directly links both IRS-1 and IRS-2 to aPKC activation in immortalized brown adipocytes, and further suggest that IRS-1 and IRS-2 are required for the activation of Cbl/PI3K during insulin action in these cells.     

Correlations in one-dimensional fully developed chaos

Correlations in the baker map and the tent map as examples of one-dimensional, fully developed chaos are considered. It is shown, utilizing symbolic dynamical systems derived from these maps, that the vanishing second-order correlation function is not sufficient to guarantee uncorrelatedness. Importance of the higher-order, especially third-order, correlation functions is emphasized for chaotic systems. In search of the quantities that grasp correlational behaviors as a whole in chaotic systems, it is proposed to use the fixed-separation correlation integral, which is a modified quantity of the usual correlation integral devised to calculate the fractal dimension of strange attractors, for these maps. It is shown that the new quantity contains all the even-number orders of autocorrelation function that are commonly considered.