Peripheral vascular disease: who gets it and why? A histomorphological analysis of 261 arterial segments from 58 cases

AIMS: This retrospective study aimed to document and illustrate the histomorphological changes underlying peripheral vascular disease (PVD). More specifically, it aimed to analyse and quantify those changes that lead to lower limb amputations. Histological changes were assessed in relation to various clinical pathologies, and significant correlations were sought thereafter. METHODS: A total of 1305 arterial segments were examined from 58 consecutive patients undergoing a lower limb amputation from January 2002 to December 2003. Serial arterial segments were taken from the femoral, popliteal, anterior tibial, posterior tibial, peroneal, and dorsalis pedis arteries, and the degrees of atherosclerotic stenosis and medial calcification were histologically quantified. RESULTS: Atherosclerosis was associated with severe arterial stenosis. An increased occurrence of severe atherosclerotic narrowing coincided with increasing patient age (p = 0.0166), hypertension (p = 0.0019), and diabetes mellitus (p = 0.0036). The presence of medial calcification was an important pathological feature in patients under 70 years of age (p = 0.0308) and significantly more severe in those with diabetes mellitus (p<0.001). CONCLUSION: Atherosclerosis and medial calcification are significant underlying lesions in diabetic patients undergoing lower limb amputation. Medial calcification can cause significant stiffening of the arterial wall and a reduction in its ability to respond to vasodilator stimuli.     

Generation and characterization of single-chain antibody fragments specific against transmembrane envelope glycoprotein gp46 of maedi-visna virus

A single-chain antibody fragments (scFv) was developed directed against transmembrane envelope glycoprotein gp46 of the virus maedi-visna, by the application of the antibody phage display library. To get specific scFv binders, the library was panned against the biotinylated peptide of 20 amino acids corresponding to the principal immunodominant domain of gp46 protein. The number of positively binding scFvs was evaluated by scFv-phage ELISA, BstN1 fingerprinting and DNA sequencing. The scFvs were expressed in soluble form and purified by immobilized metal affinity chromatography (IMAC) with a yield of 2-2.5 mg/l. Two scFvs have shown to recognize gp46 and gp150 proteins in Western blot analysis. The scFvs also recognized the virus in infected cells as shown by immunofluorescence assay. The affinity of the obtained antibody fragments to gp46 peptide was measured by surface plasmon resonance, and the resulting K(A) was in the 10(6)-10(7)lmol(-1) range. The application of characterized scFvs for expression as intrabodies in intracellular immunization against virus maedi-visna infection and for the diagnosis of this virus is discussed.     

An overview of the <Emphasis Type="Italic">Gyrodactylus</Emphasis> (Monogenea: Gyrodactylidae) species parasitizing African catfishes,and their morphological and molecular diversity

An overview of Gyrodactylus infecting catfishes from the African continent is provided, including new data from Sudan, Senegal, Kenya and Mozambique. Haptoral sclerite morphometry and nuclear ribosomal DNA sequences revealed the presence of eight Gyrodactylus species. On Senegalese Synodontis nigrita, Gyrodactylus synodonti n. sp. and Gyrodactylus nigritae n. sp. are described. These are the first reports of gyrodactylid parasites from mochokid hosts. From the fins of North African catfish Clarias gariepinus collected in Mozambique, Gyrodactylus alekosi n. sp. and Gyrodactylus rysavyi were identified. G. rysavyi was also reported from Kenyan C. gariepinus and Senegalese Clarias anguillaris. From the fins of C. anguillaris studied in Senegal, two more species, Gyrodactylus transvaalensis and Gyrodactylus gelnari n. sp. were recognised. In addition, Gyrodactylus turkanaensis n. sp. from the gills of Kenyan C. gariepinus was described and an undescribed Gyrodactylus sp. was recorded from Sudanese representatives of the same host. Detailed morphometrical and molecular comparisons of the species are presented and discussed. The study highlights the hitherto understudied diversity of viviparous monogenean parasites throughout Africa.     

High Archaea diversity in Varvara hot spring, Bulgaria

The phylogeny of the latest recognized domain, Archaea, is still complicated and it is largely based on environmental sequences. A culture independent molecular phylogenetic analysis revealed high Archaea diversity in a terrestrial hot spring, village Varvara, Bulgaria. A total of 35 archaeal operational taxonomic units (OTUs) belonging to three of the classified five Archaea phyla were identified. Most of the sequences were affiliated with the phylum Crenarchaeota (23), grouped in four branches. The rest of the sequences showed highest similarity to the unidentified archaeal clones (9), Euryarchaeota (2), and "Korarchaeota " (1). Eight (23%) of the sequenced 16S rDNAs didn't have known close relatives and represented new and diverse OTUs, four of them forming a new archaeal subgroup without close described sequences or culturable relatives. A sequence affiliated with "Korarchaeota " showed low similarity (90%) to the closest neighbor and both sequences formed unique branch in this phylum. Consequently, the constructed archaeal libraries are characterized by (1) high proportion of OTUs representing uncultivated archaeal phylogroups, (2) the abundance of novel phylotype sequences, (3) the presence of high proportions of Crenarchaeota phylotypes unrelated to cultivated organisms and (4) the presence of a sequence only distantly related to "Korarchaeota " phylum.     

Comprehensive seroprofiling of sixteen B. burgdorferi OspC: Implications for Lyme disease diagnostics design

Early diagnosis of Lyme disease (LD) is critical to successful treatment. However, current serodiagnostic tests do not reliably detect antibodies during early infection. OspC induces a potent early immune response and is also one of the most diverse proteins in the Borrelia proteome. Yet, at least 70% of the amino acid sequence is conserved among all 21 known OspC types. We performed a series of comprehensive seroprofiling studies to select the OspC types that have the most cross-reactive immunodominant epitopes. We found that proteins belonging to seven OspC types detect antibodies from all three infected host species regardless of the OspC genotype of the infecting strain. Although no one OspC type identifies all seropositive human samples, combinations of as few as two OspC proteins identified all patients that had anti-OspC antibodies.     

Typical and atypical neurovascular relations of the trigeminal nerve in the cerebellopontine angle: an anatomical study

The aim of the present study was to anatomically evaluate in adults the neurovascular trigeminal relations in the cerebellopontine angle (CPA), from a morphological and topographical perspective and thus to improve, detail and debate the pre-existing information, with educational and surgical implications. For the present anatomical study we performed bilateral dissections on 20 human adult skull bases, in formalin-fixed cadavers, at the level of the cerebellopontine angle, using the anatomical superior approach; we also studied 20 additional drawn specimens—cerebellum and brainstems, from autopsied cadavers, in order to better document the vasculature at the trigeminal root entry zone (REZ). The most constant but not exclusive neurovascular relations of the trigeminal nerves were those with the superior cerebellar artery (SCA) and the superior petrosal vein (the petrosal vein of Dandy). The regular possibility for the SCA to appear divided into a medial and a lateral branch and these to represent individual trigeminal relations at the level of the pontine cistern or REZ must not be neglected. The petrosal vein tributaries can also represent superior, inferior, or interradicular trigeminal relations. Arterioles emerging from the SCA or the anterior inferior cerebellar artery (AICA) represented trigeminal relations either at the REZ or were coursing between the trigeminal roots. A dissected specimen presented a radicular trigeminal artery emerging from the basilar artery and entering the trigeminal cavum inferior to the nerve. Another specimen presented two bony lamellae superior to the trigeminal nerve at the entrance in the trigeminal cavum—these lamellae were embedded within the lateral border of tentorium cerebelli and the posterior petroclinoid ligament. So we bring here an evidence-based support extremely useful not only for specialists dealing with this area but also for educational purposes. It appears important not only to consider the typical anatomy at this level but also to take into account the atypical and hardly predictable morphologies that may alter the diagnoses and the specific surgical procedures.     

Increased proportion of CD56bright natural killer cells in active and inactive systemic lupus erythematosus

Natural killer (NK) cells belong to the innate immune system but can also affect adaptive immune reactions. This immune regulatory function is often ascribed to the CD56(bright) subpopulation of NK cells that is prevalent in secondary lymphoid tissues and has potent cytokine-producing ability. The NK cells have been described as affecting autoimmune disease and stimulating B-cell production of antibodies, but their role in systemic lupus erythematosus (SLE) pathology has not been extensively studied. We have studied NK cells in SLE, a B-cell-driven systemic autoimmune disease, and phenotypically characterized peripheral blood NK cells in comparison to NK cells from patients with immunoglobulin A nephritis, rheumatoid arthritis and healthy individuals. We have found an increased proportion of CD56(bright) NK cells in SLE, regardless of disease activity. We detected a somewhat increased expression of the activating receptor NKp46/CD335 on NK cells from SLE patients, although neither the percentage of NK cells of all lymphocytes nor the expression of other NK receptors analysed (LIR-1/CD85j, CD94, NKG2C/CD159c, NKG2D/CD314, NKp30/CD337, NKp44/CD336, CD69) differed between patient groups. We show that type I interferon, a proinflammatory cytokine known to be abundant in SLE, can cause increases of CD56(bright) NK cells in vitro. We confirmed that serum levels of interferon-alpha were increased in active, but not in inactive, disease in the SLE patient group. In conclusion, we found an increased proportion of CD56(bright) NK cells in the blood of SLE patients, although it remains to be examined whether and how this relates to the disease process.     

High-Dimensional Single-Cell Mapping of Central Nervous System Immune Cells Reveals Distinct Myeloid Subsets in Health,Aging, and Disease

1. Download high-res image (303KB)
2. Download full-size image

     

Subnuclear partitioning of rRNA genes between the nucleolus and nucleoplasm reflects alternative epiallelic states

Eukaryotes can have thousands of 45S ribosomal RNA (rRNA) genes, many of which are silenced during development. Using fluorescence-activated sorting techniques, we show that active rRNA genes in Arabidopsis thaliana are present within sorted nucleoli, whereas silenced rRNA genes are excluded. DNA methyltransferase (met1), histone deacetylase (hda6), or chromatin assembly (caf1) mutants that disrupt silencing abrogate this nucleoplasmic–nucleolar partitioning. Bisulfite sequencing data indicate that active nucleolar rRNA genes are nearly completely demethylated at promoter CGs, whereas silenced genes are nearly fully methylated. Collectively, the data reveal that rRNA genes occupy distinct but changeable nuclear territories according to their epigenetic state.     

Probing the biofunctionality of biotinylated hyaluronan and chondroitin sulfate by hyaluronidase degradation and aggrecan interaction

Molecular interactions involving glycosaminoglycans (GAGs) are important for biological processes in the extracellular matrix (ECM) and at cell surfaces, and also in biotechnological applications. Enzymes in the ECM constantly modulate the molecular structure and the amount of GAGs in our tissues. Specifically, the changeable sulfation patterns of many GAGs are expected to be important in interactions with proteins. Biotinylation is a convenient method for immobilizing molecules to surfaces. When studying interactions at the molecular, cell and tissue level, the native properties of the immobilized molecule, i.e. its biofunctionality, need to be retained upon immobilization. Here, the GAGs hyaluronan (HA) and chondroitin sulfate (CS), and synthetically sulfated derivatives of the two, were immobilized using biotin-streptavidin binding. The degree of biotinylation and the placement of biotin groups (end-on/side-on) were varied. The introduction of biotin groups could have unwanted effects on the studied molecule, but this aspect that is not always straightforward to evaluate. Hyaluronidase, an enzyme that degrades HA and CS in the ECM, was investigated as a probe to evaluate the biofunctionality of the immobilized GAGs, using both quartz crystal microbalance and high-performance liquid chromatography. Our results showed that end-on biotinylated HA was efficiently degraded by hyaluronidase, whereas already a low degree of side-on biotinylation destroyed the degrading ability of the enzyme. Synthetically introduced sulfate groups also had this effect. Hence hyaluronidase degradation is a cheap and easy way to investigate how molecular function is influenced by the introduced functional groups. Binding experiments with the proteoglycan aggrecan emphasized the influence of protein size and surface orientation of the GAGs for in-depth studies of GAG behavior.