Novel genetic risk markers for ulcerative colitis in the IL2/IL21 region are in epistasis with IL23R and suggest a common genetic background for ulcerative colitis and celiac disease

OBJECTIVES: Recently, a genome-wide association study showed that single-nucleotide polymorphisms (SNPs) in the chromosome 4q27 region containing IL2 and IL21 are associated with celiac disease. Given the increased prevalence of inflammatory bowel disease (IBD) among celiac disease patients, we investigated the possible involvement of these SNPs in IBD. METHODS: Five SNPs strongly associated with celiac disease within the KIAA1109/TENR/IL2/IL21 linkage disequilibrium block on chromosome 4q27 and one coding SNP within the IL21 gene were analyzed in a large German IBD cohort. The study population comprised a total of 2,948 Caucasian individuals, including 1,461 IBD patients (ulcerative colitis (UC): n=514, Crohn's disease (CD): n=947) and 1,487 healthy unrelated controls. RESULTS: Three of the five celiac disease risk markers had a protective effect on UC susceptibility, and this effect remained significant after correcting for multiple testing: rs6840978: P=0.0082, P(corr)=0.049, odds ratio (OR) 0.77, 95% confidence interval (CI) 0.63-0.93; rs6822844: P=0.0028, P(corr)=0.017, OR 0.73, 95% CI 0.59-0.90; rs13119723: P=0.0058, P(corr)=0.035, OR 0.75, 95% CI 0.61-0.92. A haplotype consisting of the six SNPs tested was markedly associated with UC susceptibility (P=0.0025, P(corr)=0.015, OR 0.72, 95% CI 0.58-0.89). Moreover, in UC, epistasis was observed between the IL23R SNP rs1004819 and three SNPs in the KIAA1109/TENR/IL2/IL21 block (rs13151961, rs13119723, and rs6822844). CONCLUSIONS: Similar to other autoimmune diseases such as celiac disease, rheumatoid arthritis, type 1 diabetes, Graves' disease, and psoriatic arthritis, genetic variation in the chromosome 4q27 region predisposes to UC, suggesting a common genetic background for these diseases.

The novel ATP-competitive inhibitor of the MET hepatocyte growth factor receptor EMD1214063 displays inhibitory activity against selected MET-mutated variants

The receptor tyrosine kinase MET is a prime target in clinical oncology due to its aberrant activation and involvement in the pathogenesis of a broad spectrum of malignancies. Similar to other targeted kinases, primary and secondary mutations seem to represent an important resistance mechanism to MET inhibitors. Here, we report the biologic activity of a novel MET inhibitor, EMD1214063, on cells that ectopically express the mutated MET variants M1268T, Y1248H, H1112Y, L1213V, H1112L, V1110I, V1206L, and V1238I. Our results demonstrate a dose-dependent decrease in MET autophosphorylation in response to EMD1214063 in five out of the eight cell lines (IC50 2-43nM). Blockade of MET by EMD1214063 was accompanied by a reduced activation of downstream effectors in cells expressing EMD1214063-sensitive mutants. In all sensitive mutant-expressing lines, EMD1214063 altered cell cycle distribution, primarily with an increase in G1 phase. EMD1214063 strongly influenced MET-driven biological functions, such as cellular morphology, MET-dependent cell motility and anchorage-independent growth. To assess the in vivo efficacy of EMD1214063, we used a xenograft tumor model in immunocompromised mice bearing NIH3T3 cells expressing sensitive and resistant MET mutated variants. Animals were randomized for the treatment with EMD1214063 (50mg/kg/day) or vehicle only. Remarkably, five days of EMD1214063 treatment resulted in a complete regression of the sensitive H1112L-derived tumors, while tumor growth remained unaffected in mice with L1213V tumors and in vehicle-treated animals. Collectively, the current data identifies EMD1214063 as a potent MET small molecule inhibitor with selective activity towards mutated MET variants.

Characterization of a novel picornavirus isolate from a diseased European eel (Anguilla anguilla)

A novel picornavirus was isolated from specimens of a diseased European eel (Anguilla anguilla). This virus induced a cytopathic effect in eel embryonic kidney cells and high mortality in a controlled transmission study using elvers. Eel picornavirus has a genome of 7,496 nucleotides that encodes a polyprotein of 2,259 amino acids. It has a typical picornavirus genome layout, but its low similarity to known viral proteins suggests a novel species in the family Picornaviridae.

Mitochondrial Outer Membrane Proteome of Trypanosoma brucei Reveals Novel Factors Required to Maintain Mitochondrial Morphology

Trypanosoma brucei is a unicellular parasite that causes devastating diseases in humans and animals. It diverged from most other eukaryotes very early in evolution and, as a consequence, has an unusual mitochondrial biology. Moreover, mitochondrial functions and morphology are highly regulated throughout the life cycle of the parasite. The outer mitochondrial membrane defines the boundary of the organelle. Its properties are therefore key for understanding how the cytosol and mitochondria communicate and how the organelle is integrated into the metabolism of the whole cell. We have purified the mitochondrial outer membrane of T. brucei and characterized its proteome using label-free quantitative mass spectrometry for protein abundance profiling in combination with statistical analysis. Our results show that the trypanosomal outer membrane proteome consists of 82 proteins, two-thirds of which have never been associated with mitochondria before. 40 proteins share homology with proteins of known functions. The function of 42 proteins, 33 of which are specific to trypanosomatids, remains unknown. 11 proteins are essential for the disease-causing bloodstream form of T. brucei and therefore may be exploited as novel drug targets. A comparison with the outer membrane proteome of yeast defines a set of 17 common proteins that are likely present in the mitochondrial outer membrane of all eukaryotes. Known factors involved in the regulation of mitochondrial morphology are virtually absent in T. brucei. Interestingly, RNAi-mediated ablation of three outer membrane proteins of unknown function resulted in a collapse of the network-like mitochondrion of procyclic cells and for the first time identified factors that control mitochondrial shape in T. brucei.

Validation of novel 3-dimensional electrocardiographic mapping of atrial tachycardias by invasive mapping and ablation: a multicenter study

OBJECTIVES This study prospectively evaluated the role of a novel 3-dimensional, noninvasive, beat-by-beat mapping system, Electrocardiographic Mapping (ECM), in facilitating the diagnosis of atrial tachycardias (AT). BACKGROUND Conventional 12-lead electrocardiogram, a widely used noninvasive tool in clinical arrhythmia practice, has diagnostic limitations. METHODS Various AT (de novo and post-atrial fibrillation ablation) were mapped using ECM followed by standard-of-care electrophysiological mapping and ablation in 52 patients. The ECM consisted of recording body surface electrograms from a 252-electrode-vest placed on the torso combined with computed tomography-scan-based biatrial anatomy (CardioInsight Inc., Cleveland, Ohio). We evaluated the feasibility of this system in defining the mechanism of AT-macro-re-entrant (perimitral, cavotricuspid isthmus-dependent, and roof-dependent circuits) versus centrifugal (focal-source) activation-and the location of arrhythmia in centrifugal AT. The accuracy of the noninvasive diagnosis and detection of ablation targets was evaluated vis-à-vis subsequent invasive mapping and successful ablation. RESULTS Comparison between ECM and electrophysiological diagnosis could be accomplished in 48 patients (48 AT) but was not possible in 4 patients where the AT mechanism changed to another AT (n = 1), atrial fibrillation (n = 1), or sinus rhythm (n = 2) during the electrophysiological procedure. ECM correctly diagnosed AT mechanisms in 44 of 48 (92%) AT: macro-re-entry in 23 of 27; and focal-onset with centrifugal activation in 21 of 21. The region of interest for focal AT perfectly matched in 21 of 21 (100%) AT. The 2:1 ventricular conduction and low-amplitude P waves challenged the diagnosis of 4 of 27 macro-re-entrant (perimitral) AT that can be overcome by injecting atrioventricular node blockers and signal averaging, respectively. CONCLUSIONS This prospective multicenter series shows a high success rate of ECM in accurately diagnosing the mechanism of AT and the location of focal arrhythmia. Intraprocedural use of the system and its application to atrial fibrillation mapping is under way.

Novel pseudo-staphylococcal cassette chromosome mec element (ψSCCmec57395) in methicillin-resistant Staphylococcus pseudintermedius CC45

Genetic characterization of methicillin-resistant Staphylococcus pseudintermedius (MRSP) from Thailand and Israel revealed the presence of a predominant atypical clonal lineage which was not typeable by SmaI-PFGE and SCCmec typing. All the atypical isolates (n = 34) belonged to CC45 (30 ST45 and 2 ST179 isolates, 1 ST57 isolate, and 1 ST85 isolate). The isolates originated from healthy and diseased dogs and cats, as well as from the environment of one clinic. Cfr9I-pulsed-field gel electrophoresis (Cfr9I-PFGE) and dru typing permitted the further distinction of CC45 isolates from the two different countries. Microarray analysis identified genes that confer resistance to β-lactams (mecA; blaZ), aminoglycosides [aac(6')-Ie-aph(2')-Ia; aph(3')-III; ant(6)-Ia], macrolides and lincosamides [erm(B)], tetracyclines [tet(M)], trimethoprim [dfr(G)], streptothricin (sat4), and chloramphenicol (catpC221). Fluoroquinolone resistance was attributed to specific amino acid substitutions, i.e., Ser84Leu in GyrA and Ser80Ile and Asp84Asn in GrlA. A novel pseudo-staphylococcal cassette chromosome (ΨSCCmec57395) element was identified in MRSP strain 57395 (sequence type ST45) by whole-genome sequencing. The 12,282-bp ΨSCCmec57395 element contained a class C1 mec gene complex but no ccr genes. In addition to the methicillin resistance gene mecA, ΨSCCmec57395 also carried determinants of resistance to heavy metals, such as arsenic, cadmium, and copper. Bsu36I restriction analysis of the ΨSCCmec57395 element amplified by long-range PCR revealed the presence of ΨSCCmec57395 in the 33 additional isolates of MRSP CC45. The ΨSCCmec57395 element represents a new class of SCCmec and has been identified in MRSP of CC45, which is a predominant clonal lineage in Israel and Thailand.

DNA-based identification of novel bovine casein gene variants

In cattle, at least 39 variants of the 4 casein proteins (α(S1)-, β-, α(S2)- and κ-casein) have been described to date. Many of these variants are known to affect milk-production traits, cheese-processing properties, and the nutritive value of milk. They also provide valuable information for phylogenetic studies. So far, the majority of studies exploring the genetic variability of bovine caseins considered European taurine cattle breeds and were carried out at the protein level by electrophoretic techniques. This only allows the identification of variants that, due to amino acid exchanges, differ in their electric charge, molecular weight, or isoelectric point. In this study, the open reading frames of the casein genes CSN1S1, CSN2, CSN1S2, and CSN3 of 356 animals belonging to 14 taurine and 3 indicine cattle breeds were sequenced. With this approach, we identified 23 alleles, including 5 new DNA sequence variants, with a predicted effect on the protein sequence. The new variants were only found in indicine breeds and in one local Iranian breed, which has been phenotypically classified as a taurine breed. A multidimensional scaling approach based on available SNP chip data, however, revealed an admixture of taurine and indicine populations in this breed as well as in the local Iranian breed Golpayegani. Specific indicine casein alleles were also identified in a few European taurine breeds, indicating the introgression of indicine breeds into these populations. This study shows the existence of substantial undiscovered genetic variability of bovine casein loci, especially in indicine cattle breeds. The identification of new variants is a valuable tool for phylogenetic studies and investigations into the evolution of the milk protein genes.

In vitro effects of novel ruthenium complexes in Neospora caninum and Toxoplasma gondii tachyzoites.

Upon the screening of 16 antiproliferative compounds against Toxoplasma gondii and Neospora caninum, two hydrolytically stable ruthenium complexes (compounds 16 and 18) exhibited 50% inhibitory concentrations of 18.7 and 41.1 nM (T. gondii) and 6.7 and 11.3 nM (N. caninum). To achieve parasiticidal activity with compound 16, long-term treatment (22 to 27 days at 80 to 160 nM) was required. Transmission electron microscopy demonstrated the rapid impact on and ultrastructural alterations in both parasites. These preliminary findings suggest that the potential of ruthenium-based compounds should thus be further exploited.

Tumor regression grade of urothelial bladder cancer after neoadjuvant chemotherapy: a novel and successful strategy to predict survival.

Histopathologic tumor regression grades (TRGs) after neoadjuvant chemotherapy predict survival in different cancers. In bladder cancer, corresponding studies have not been conducted. Fifty-six patients with advanced invasive urothelial bladder cancer received neoadjuvant chemotherapy before cystectomy and lymphadenectomy. TRGs were defined as follows: TRG1: complete tumor regression; TRG2: >50% tumor regression; TRG3: 50% or less tumor regression. Separate TRGs were assigned for primary tumors and corresponding lymph nodes. The prognostic impact of these 2 TRGs, the highest (dominant) TRG per patient, and competing tumor features reflecting tumor regression (ypT/ypN stage, maximum diameter of the residual tumor) were determined. Tumor characteristics in initial transurethral resection of the bladder specimens were tested for response prediction. The frequency of TRGs 1, 2, and 3 in the primary tumors were n=16, n=19, and n=21; corresponding data from the lymph nodes were n=31, n=9, and n=16. Interobserver agreement in determination of the TRG was strong (κ=0.8). Univariately, all evaluated parameters were significantly (P≤0.001) related to overall survival; however, the segregation of the Kaplan-Meier curves was best for the dominant TRG. In multivariate analysis, only dominant TRG predicted overall survival independently (P=0.035). In transurethral resection specimens of the chemotherapy-naive bladder cancer, the only tumor feature with significant (P<0.03) predictive value for therapy response was a high proliferation rate. In conclusion, among all parameters reflecting tumor regression, the dominant TRG was the only independent risk factor. A favorable chemotherapy response is associated with a high proliferation rate in the initial chemotherapy-naive bladder cancer. This feature might help personalize neoadjuvant chemotherapy.

A novel family of transporters mediating the transport of glutathione derivatives in plants

Uptake and compartmentation of reduced glutathione (GSH), oxidized glutathione (GSSG), and glutathione conjugates are important for many functions including sulfur transport, resistance against biotic and abiotic stresses, and developmental processes. Complementation of a yeast (Saccharomyces cerevisiae) mutant (hgt1) deficient in glutathione transport was used to characterize a glutathione transporter cDNA (OsGT1) from rice (Oryza sativa). The 2.58-kb full-length cDNA (AF393848, gi 27497095), which was obtained by screening of a cDNA library and 5'-rapid amplification of cDNA ends-polymerase chain reaction, contains an open reading frame encoding a 766-amino acid protein. Complementation of the hgt1 yeast mutant strain with the OsGT1 cDNA restored growth on a medium containing GSH as the sole sulfur source. The strain expressing OsGT1 mediated H-3]GSH uptake, and this uptake was significantly competed not only by unlabeled GSSG and GS conjugates but also by some amino acids and peptides, suggesting a wide substrate specificity. OsGT1 may be involved in the retrieval of GSSG, GS conjugates, and nitrogen-containing peptides from the cell wall.