A synthetic glycosaminoglycan mimetic binds vascular endothelial growth factor and modulates angiogenesis

In a previous study, we showed that in situ injection of glycosaminoglycan mimetics called RGTAs (ReGeneraTing Agents) enhanced neovascularization after skeletal muscular ischemia (Desgranges, P., Barbaud, C., Caruelle, J. P., Barritault, D., and Gautron, J. (1999) FASEB J. 13, 761-766). In the present study, we showed that the RGTA OTR4120 modulated angiogenesis in the chicken embryo chorioallantoic membrane assay, in a dose-dependent manner. We therefore investigated the effect of OTR4120 on one of the most specific angiogenesis-regulating heparin-binding growth factors, vascular endothelial growth factor 165 (VEGF165). OTR4120 showed high affinity binding to VEGF165 (Kd = 2.2 nm), as compared with heparin (Kd = 15 nm), and potentiated the affinity of VEGF165 for VEGF receptor-1 and -2 and for neuropilin-1. In vitro, OTR4120 potentiated VEGF165-induced proliferation and migration of human umbilical vein endothelial cells. In the in vivo Matrigel plug angiogenesis assay, OTR4120 in a concentration as low as 3 ng/ml caused a 6-fold increase in VEGF165-induced angiogenesis. Immunohistochemical staining showed a larger number of well differentiated VEGFR-2-expressing-cells in Matrigel sections of OTR4120-treated plug than in control sections. These findings indicate that OTR4120 enhances the VEGF165-induced angiogenesis and therefore may hold promise for treating disorders characterized by deficient angiogenesis.     

Review: Impact of Helicobacter pylori on Alzheimer's disease: What do we know so far?

Background

Helicobacter pylori has changed radically gastroenterologic world, offering a new concept in patients' management. Over time, more medical data gave rise to diverse distant, extragastric manifestations and interactions of the “new” discovered bacterium. Special interest appeared within the field of neurodegenerative diseases and particularly Alzheimer's disease, as the latter and Helicobacter pylori infection are associated with a large public health burden and Alzheimer's disease ranks as the leading cause of disability. However, the relationship between Helicobacter pylori infection and Alzheimer's disease remains uncertain.

Methods

We performed a narrative review regarding a possible connection between Helicobacter pylori and Alzheimer's disease. All accessible relevant (pre)clinical studies written in English were included. Both affected pathologies were briefly analyzed, and relevant studies are discussed, trying to focus on the possible pathogenetic role of this bacterium in Alzheimer's disease.

Results

Data stemming from both epidemiologic studies and animal experiments seem to be rather encouraging, tending to confirm the hypothesis that Helicobacter pylori infection might influence the course of Alzheimer's disease pleiotropically. Possible main mechanisms may include the bacterium's access to the brain via the oral‐nasal‐olfactory pathway or by circulating monocytes (infected with Helicobacter pylori due to defective autophagy) through disrupted blood‐brain barrier, thereby possibly triggering neurodegeneration.

Conclusions

Current data suggest that Helicobacter pylori infection might influence the pathophysiology of Alzheimer's disease. However, further large‐scale randomized controlled trials are mandatory to clarify a possible favorable effect of Helicobacter pylori eradication on Alzheimer's disease pathophysiology, before the recommendation of short‐term and cost‐effective therapeutic regimens against Helicobacter pylori‐related Alzheimer's disease.     

Gene expression and biochemical characterization of Azotobacter vinelandii cyclophilins and Protein Interaction Studies of the cytoplasmic isoform with dnaK and lpxH

The soil nitrogen-fixing bacterium Azotobacter vinelandii possesses two cyclophilins, comprising putative cytoplasmic and periplasmic isoforms, designated as AvPPIB and AvPPIA, respectively. Both recombinant cyclophilins have been purified and their peptidyl-prolyl cis/trans isomerase activity against Suc-Ala-Xaa-Pro-Phe-pNA synthetic peptides has been characterized. The substrate specificity of both cyclophilins is typical for bacterial cyclophilins, with Suc-Ala-Ala-Pro-Phe-pNA being the most rapidly catalyzed substrate. The cytoplasmic cyclophilin also displays a chaperone function in the citrate synthase thermal aggregation assay. Using real-time quantitative RT-PCR, we demonstrate that AvppiB is expressed under various physiological and growth conditions, mainly upregulated by acetate and downregulated by the stationary growth state, while AvppiA shows a tendency for downregulation under the tested conditions. Further, we identified chaperone protein dnaK and UDP-2, 3-diacylglucosamine hydrolase lpxH as probable interacting partners of AvPPIB and we demonstrate their physical interaction by coexpression studies. An increase in AvPPIB PPIase activity in the presence of AvdnaK and a decrease in the presence of AvlpxH further confirms each interaction. However, the PPIase activity does not seem to be essential for those interactions since AvPPIB active site mutants still interact with dnaK and lpxH, while their minor PPIase activity cannot be modulated by the interaction.     

Lu/BCAM Adhesion Glycoprotein Is a Receptor for Escherichia coli Cytotoxic Necrotizing Factor 1 (CNF1)

The Cytotoxic Necrotizing Factor 1 (CNF1) is a protein toxin which is a major virulence factor of pathogenic Escherichia coli strains. Here, we identified the Lutheran (Lu) adhesion glycoprotein/basal cell adhesion molecule (BCAM) as cellular receptor for CNF1 by co-precipitation of cell surface molecules with tagged toxin. The CNF1-Lu/BCAM interaction was verified by direct protein-protein interaction analysis and competition studies. These studies revealed amino acids 720 to 1014 of CNF1 as the binding site for Lu/BCAM. We suggest two cell interaction sites in CNF1: first the N-terminus, which binds to p37LRP as postulated before. Binding of CNF1 to p37LRP seems to be crucial for the toxin''s action. However, it is not sufficient for the binding of CNF1 to the cell surface. A region directly adjacent to the catalytic domain is a high affinity interaction site for Lu/BCAM. We found Lu/BCAM to be essential for the binding of CNF1 to cells. Cells deficient in Lu/BCAM but expressing p37LRP could not bind labeled CNF1. Therefore, we conclude that LRP and Lu/BCAM are both required for toxin action but with different functions.     

Acromyrmex Leaf-Cutting Ants Have Simple Gut Microbiota with Nitrogen-Fixing Potential

Ants and termites have independently evolved obligate fungus-farming mutualisms, but their gardening procedures are fundamentally different, as the termites predigest their plant substrate whereas the ants deposit it directly on the fungus garden. Fungus-growing termites retained diverse gut microbiota, but bacterial gut communities in fungus-growing leaf-cutting ants have not been investigated, so it is unknown whether and how they are specialized on an exclusively fungal diet. Here we characterized the gut bacterial community of Panamanian Acromyrmex species, which are dominated by only four bacterial taxa: Wolbachia, Rhizobiales, and two Entomoplasmatales taxa. We show that the Entomoplasmatales can be both intracellular and extracellular across different gut tissues, Wolbachia is mainly but not exclusively intracellular, and the Rhizobiales species is strictly extracellular and confined to the gut lumen, where it forms biofilms along the hindgut cuticle supported by an adhesive matrix of polysaccharides. Tetracycline diets eliminated the Entomoplasmatales symbionts but hardly affected Wolbachia and only moderately reduced the Rhizobiales, suggesting that the latter are protected by the biofilm matrix. We show that the Rhizobiales symbiont produces bacterial NifH proteins that have been associated with the fixation of nitrogen, suggesting that these compartmentalized hindgut symbionts alleviate nutritional constraints emanating from an exclusive fungus garden diet reared on a substrate of leaves.     

Profilin 1 is a potential biomarker for bladder cancer aggressiveness

Of the most important clinical needs for bladder cancer (BC) management is the identification of biomarkers for disease aggressiveness. Urine is a "gold mine" for biomarker discovery, nevertheless, with multiple proteins being in low amounts, urine proteomics becomes challenging. In the present study we applied a fractionation strategy of urinary proteins based on the use of immobilized metal affinity chromatography for the discovery of biomarkers for aggressive BC. Urine samples from patients with non invasive (two pools) and invasive (two pools) BC were subjected to immobilized metal affinity chromatography fractionation and eluted proteins analyzed by 1D-SDS-PAGE, band excision and liquid chromatography tandem MS. Among the identified proteins, multiple corresponded to proteins with affinity for metals and/or reported to be phosphorylated and included proteins with demonstrated association with BC such as MMP9, fibrinogen forms, and clusterin. In agreement to the immobilized metal affinity chromatography results, aminopeptidase N, profilin 1, and myeloblastin were further found to be differentially expressed in urine from patients with invasive compared with non invasive BC and benign controls, by Western blot or Elisa analysis, nevertheless exhibiting high interindividual variability. By tissue microarray analysis, profilin 1 was found to have a marked decrease of expression in the epithelial cells of the invasive (T2+) versus high risk non invasive (T1G3) tumors with occasional expression in stroma; importantly, this pattern strongly correlated with poor prognosis and increased mortality. The functional relevance of profilin 1 was investigated in the T24 BC cells where blockage of the protein by the use of antibodies resulted in decreased cell motility with concomitant decrease in actin polymerization. Collectively, our study involves the application of a fractionation method of urinary proteins and as one main result of this analysis reveals the association of profilin 1 with BC paving the way for its further investigation in BC stratification.