Creutzfeldt-Jakob Disease with a prion protein gene codon 180 mutation presenting asymmetric cortical high-intensity on magnetic resonance imaging

ABSTRACT. Here we report a genetically confirmed case of Creutzfeldt-Jakob disease with a prion protein gene codon 180 mutation presenting atypical magnetic resonance imaging findings. The present case exhibited an acute onset and lateralized neurologic signs, and progressive cognitive impairment. No myoclonus or periodic synchronous discharges on electroencephalography were observed. Diffusion-weighted images revealed areas of high signal intensity in the right frontal and temporal cortices at onset that extended to the whole cortex and basal ganglia of the right cerebral hemisphere at 3 months. Although the cerebrospinal fluid (CSF) was initially negative for neuron specific enolase, tau protein, 14–3–3 protein, and abnormal prion protein, the CSF was positive for these brain-derived proteins at 3 months after onset.     

Common variants in CDKN2B-AS1 associated with optic-nerve vulnerability of glaucoma identified by genome-wide association studies in Japanese

Background

To date, only a small portion of the genetic variation for primary open-angle glaucoma (POAG), the major type of glaucoma, has been elucidated.

Methods and Principal Findings

We examined our two data sets of the genome-wide association studies (GWAS) derived from a total of 2,219 Japanese subjects. First, we performed a GWAS by analyzing 653,519 autosomal common single-nucleotide polymorphisms (SNPs) in 833 POAG patients and 686 controls. As a result, five variants that passed the Bonferroni correction were identified in CDKN2B-AS1 on chromosome 9p21.3, which was already reported to be a significant locus in the Caucasian population. Moreover, we combined the data set with our previous GWAS data set derived from 411 POAG patients and 289 controls by the Mantel-Haenszel test, and all of the combined variants showed stronger association with POAG (P<5.8×10⁻¹⁰). We then subdivided the case groups into two subtypes based on the value of intraocular pressure (IOP)—POAG with high IOP (high pressure glaucoma, HPG) and that with normal IOP (normal pressure glaucoma, NPG)—and performed the GWAS using the two data sets, as the prevalence of NPG in Japanese is much higher than in Caucasians. The results suggested that the variants from the same CDKN2B-AS1 locus were likely to be significant for NPG patients.

Conclusions and Significance

In this study, we successfully identified POAG-associated variants in the CDKN2B-AS1 locus using a Japanese population, i.e., variants originally reported as being associated with the Caucasian population. Although we cannot rule out that the significance could be due to the differences in sample size between HPG and NPG, the variants could be associated specifically with the vulnerability of the optic nerve to IOP, which is useful for investigating the etiology of glaucoma.     

Cross-talk between integrin α6β4 and insulin-like growth factor-1 receptor (IGF1R) through direct α6β4 binding to IGF1 and subsequent α6β4-IGF1-IGF1R ternary complex formation in anchorage-independent conditions

Integrin αvβ3 plays a role in insulin-like growth factor-1 (IGF1) signaling (integrin-IGF1 receptor (IGF1R) cross-talk). The specifics of the cross-talk are, however, unclear. In a current model, "ligand occupancy" of αvβ3 (i.e. the binding of extracellular matrix proteins) enhances signaling induced by IGF1 binding to IGF1R. We recently reported that IGF1 directly binds to αvβ3 and induces αvβ3-IGF1-IGF1R ternary complex formation. Consistently, the integrin binding-defective IGF1 mutant (R36E/R37E) is defective in inducing ternary complex formation and IGF signaling, but it still binds to IGF1R. Like αvβ3, integrin α6β4 is overexpressed in many cancers and is implicated in cancer progression. Here, we discovered that α6β4 directly bound to IGF1, but not to R36E/R37E. Grafting the β4 sequence WPNSDP (residues 167-172), which corresponds to the specificity loop of β3, to integrin β1 markedly enhanced IGF1 binding to β1, suggesting that the WPNSDP sequence is involved in IGF1 recognition. WT IGF1 induced α6β4-IGF1-IGF1R ternary complex formation, whereas R36E/R37E did not. When cells were attached to matrix, exogenous IGF1 or α6β4 expression had little or no effect on intracellular signaling. When cell-matrix adhesion was reduced (in poly(2-hydroxyethyl methacrylate-coated plates), IGF1 induced intracellular signaling and enhanced cell survival in an α6β4-dependent manner. Also IGF1 enhanced colony formation in soft agar in an α6β4-dependent manner. These results suggest that IGF binding to α6β4 plays a major role in IGF signaling in anchorage-independent conditions, which mimic the in vivo environment, and is a novel therapeutic target.     

Facile synthesis and evaluation of C-functionalized benzyl-1-oxa-4,7,10-triazacyclododecane-N,N',N″-triacetic acid as chelating agent for ¹¹¹In-labeled polypeptides

A 12-membered polyazamacrocycle, 1-oxa-4,7,10-triazacyclododecane-N,N′,N″-triacetic acid (ODTA), has been reported to provide an indium chelate of net neutral charge with thermodynamic stability higher than 1,4,7,10-tetraazacyclododecane-N,N′,N″,N?-tetraacetic acid (DOTA). However, neither synthetic procedure for a C-functionalized ODTA (C-ODTA) nor its chelating ability with a trace amount of radioactive indium-111 (¹¹¹In) has been elucidated. We herein present a facile synthetic procedure for C-ODTA, and estimated its ability as a chelating agent for radiolabeling peptides and proteins with ¹¹¹In. The synthetic procedure involves the synthesis of a linear precursor using a para-substituted phenylalanine derivative as a starting material. The following intramolecular cyclization reaction was best performed (>73% yield) when Boc-protected linear compound and the condensation reagent, HATU, were simultaneously added to the reaction vessel at the same flow rate. The cyclic compound was then reduced with BH₃ and alkylated with tert-butyl bromoacetate. The synthetic procedure was straightforward and some optimization would be required. However, most of the intermediate compounds were obtained easily in good yields, suggesting that the present synthetic procedure would be useful to synthesize C-ODTA derivatives. The intramolecular cyclization reaction might also be applicable to synthesize polyazamacrocycles of different ring sizes and cyclic peptides. In ¹¹¹In radiolabeling reactions, C-ODTA provided ¹¹¹In chelates in higher radiochemical yields at low ligand concentrations when compared with C-DOTA. The ¹¹¹In-labeled C-ODTA remained unchanged in the presence of apo-transferrin. The biodistribution studies also showed that the ¹¹¹In-labeled compound was mainly excreted into urine as intact. These findings indicate that C-ODTA would be useful to prepare ¹¹¹In-labeled peptides of high specific activities in high radiochemical yields.     

The reticulon and DP1/Yop1p proteins form immobile oligomers in the tubular endoplasmic reticulum

We recently identified a class of membrane proteins, the reticulons and DP1/Yop1p, which shape the tubular endoplasmic reticulum (ER) in yeast and mammalian cells. These proteins are highly enriched in the tubular portions of the ER and virtually excluded from other regions. To understand how they promote tubule formation, we characterized their behavior in cellular membranes and addressed how their localization in the ER is determined. Using fluorescence recovery after photobleaching, we found that yeast Rtn1p and Yop1p are less mobile in the membrane than normal ER proteins. Sucrose gradient centrifugation and cross-linking analyses show that they form oligomers. Mutants of yeast Rtn1p, which no longer localize exclusively to the tubular ER or are even totally inactive in inducing ER tubules, are more mobile and oligomerize less extensively. The mammalian reticulons and DP1 are also relatively immobile and can form oligomers. The conserved reticulon homology domain that includes the two membrane-embedded segments is sufficient for the localization of the reticulons to the tubular ER, as well as for their diffusional immobility and oligomerization. Finally, ATP depletion in both yeast and mammalian cells further decreases the mobilities of the reticulons and DP1. We propose that oligomerization of the reticulons and DP1/Yop1p is important for both their localization to the tubular domains of the ER and for their ability to form tubules.     

Immunomodulatory strategies prevent the development of autoimmune emphysema

Background

The presence of anti-endothelial cell antibodies and pathogenic T cells may reflect an autoimmune component in the pathogenesis of emphysema. Whether immune modulatory strategies can protect against the development of emphysema is not known.

Methods

Sprague Dawley rats were immunized with human umbilical vein endothelial cells (HUVEC) to induce autoimmune emphysema and treated with intrathymic HUVEC-injection and pristane. Measurements of alveolar airspace enlargement, cytokine levels, immuno histochemical, western blot analysis, and T cell repertoire of the lung tissue were performed.

Results

The immunomodulatory strategies protected lungs against cell death as demonstrated by reduced numbers of TUNEL and active caspase-3 positive cells and reduced levels of active caspase-3, when compared with lungs from HUVEC-immunized rats. Immunomodulatory strategies also suppressed anti-endothelial antibody production and preserved CNTF, IL-1alpha and VEGF levels. The immune deviation effects of the intrathymic HUVEC-injection were associated with an expansion of CD4+CD25+Foxp3+ regulatory T cells. Pristane treatment decreased the proportion of T cells expressing receptor beta-chain, Vβ16.1 in the lung tissue.

Conclusions

Our data demonstrate that interventions classically employed to induce central T cell tolerance (thymic inoculation of antigen) or to activate innate immune responses (pristane treatment) can prevent the development of autoimmune emphysema.     

NADPH dependent activation of microsomal glutathione transferase 1

Microsomal glutathione transferase 1 (MGST1) can become activated up to 30-fold by several mechanisms in vitro (e.g. covalent modification by reactive electrophiles such as N-ethylmaleimide (NEM)). Activation has also been observed in vivo during oxidative stress. It has been noted that an NADPH generating system (g.s.) can activate MGST1 (up to 2-fold) in microsomal incubations, but the mechanism was unclear. We show here that NADPH g.s treatment impaired N-ethylmaleimide activation, indicating a shared target (identified as cysteine-49 in the latter case). Furthermore, NADPH activation was prevented by sulfhydryl compounds (glutathione and dithiothreitol). A well established candidate for activation would be oxidative stress, however we could exclude that oxidation mediated by cytochrome P450 2E1 (or flavine monooxygenase) was responsible for activation under a defined set of experimental conditions since superoxide or hydrogen peroxide alone did not activate the enzyme (in microsomes prepared by our routine procedure). Actually, the ability of MGST1 to become activated by hydrogen peroxide is critically dependent on the microsome preparation method (which influences hydrogen peroxide decomposition rate as shown here), explaining variable results in the literature. NADPH g.s. dependent activation of MGST1 could instead be explained, at least partly, by a direct effect observed also with purified enzyme (up to 1.4-fold activation). This activation was inhibited by sulfhydryl compounds and thus displays the same characteristics as that of the microsomal system. Whereas NADPH, and also ATP, activated purified MGST1, several nucleotide analogues did not, demonstrating specificity. It is thus an intriguing possibility that MGST1 function could be modulated by ligands (as well as reactive oxygen species) during oxidative stress when sulfhydryls are depleted.     

Spring migration dynamics and sex-specific patterns in stopover strategy in the Wood Sandpiper Tringa glareola

Due to being a virtually monomorphic wader species, migration dynamics and sex-related migration patterns in the Wood Sandpiper (Tringa glareola) have rarely been investigated. We captured spring migrants at an important stopover site in northeastern Austria. Birds were individually color-marked, and sex was determined by an analysis of DNA from tail feather material. Among temporary residents (birds seen again after day of capture), males migrated on average 3 days earlier than females. However, since sexes did not differ in fat score, the length of stay and the proportion of transients (birds not seen again after day of capture) and temporary residents, we suggest that males and females adopt similar migration strategies in the spring. The large number of transients captured as well as shorter stopover durations in later temporary residents indicate that Wood Sandpipers minimize time at this stage of their northbound migration. Temporary residents earlier in the season exhibited lower fat stores than later ones. Nevertheless, since the fat stores of transients and temporary residents were similar even after the progress of the season had been accounted for, we assume that Wood Sandpipers may afford to exhibit individual flexibility in migration strategy and the use of stopover sites, especially early in the season. This variability may be a necessary adaptation to cope with possible varying environmental conditions at dynamic and unpredictable inland stopover sites. After having reached North Mediterranean regions, mean body mass of spring migrants gradually increases during successive stopovers, indicating that Wood Sandpipers follow a ‘hopping’ migration strategy. This emphasizes the high conservation value of even small artificial mudflat pools as important stepping stones in order to maintain a continuous network of wetland habitats for this continental migrant.     

Design, synthesis and structure-activity relationship studies of novel indazole analogues as DNA gyrase inhibitors with Gram-positive antibacterial activity

In this study, we report the design, synthesis and structure-activity relationships of novel indazole derivatives as DNA gyrase inhibitors with Gram-positive antibacterial activity. Our results show that selected compounds from this series exhibit potent antibacterial activity against Gram-positive bacteria including multi-drug resistant strains that is methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecalis (VRE).