The life cycle of<Emphasis Type="Italic"> Anisakis simplex</Emphasis> in the Norwegian Deep (northern North Sea)

Copepoda (Calanus finmarchicus n=1,722, Paraeuchaeta norvegica n=1,955), Hyperiidae (n=3,019), Euphausiacea (Meganyctiphanes norvegica n=4,780), and the fishes Maurolicus muelleri (n=500) and Pollachius virens (n=33) were collected in the Norwegian Deep (northern North Sea) during summer 2001 to examine the importance of pelagic invertebrates and vertebrates as hosts of Anisakis simplex and their roles in the transfer of this nematode to its final hosts (Cetaceans). Third stage larvae (L3) of A. simplex were found in P. norvegica, M. muelleri and P. virens. The prevalence of A. simplex in dissected P. norvegica was 0.26%, with an intensity of 1. Prevalences in M. muelleri and P. virens were 49.6% and 100.0%, with mean intensities of 1.1–2.6 (total fish length 6.0–7.2) and 193.6, respectively. All specimens of C. finmarchicus and M. norvegica examined were free of anisakid nematode species and no other parasites were detected. P. norvegica, which harboured the third stage larvae, is the obligatory first intermediate host of A. simplex in the investigated area. Though there was no apparent development of larvae in M. muelleri, this fish can be considered as the obligatory second intermediate host of A. simplex in the Norwegian Deep. However, it is unlikely that the larva from P. norvegica can be successfully transmitted into the cetacean or pinniped final hosts, where they reach the adult stage. An additional growth phase and a second intermediate host is the next phase in the life cycle. Larger predators such as P. virens serve as paratenic hosts, accumulating the already infective stage from M. muelleri. The oceanic life cycle of A. simplex in the Norwegian Deep is very different in terms of hosts and proposed life cycle patterns of A. simplex from other regions, involving only a few intermediate hosts. In contrast to earlier suggestions, euphausiids have no importance at all for the successful transmission of A. simplex in the Norwegian Deep. This demonstrates that this nematode is able to select definite host species depending on the locality, apparently having a very low level of host specificity. This could explain the wide range of different hosts that have been recorded for this species, and can be seen as the reason for the success of this parasite in reaching its marine mammal final hosts in an oceanic environment.     

Scaffold protein harmonin (USH1C) provides molecular links between Usher syndrome type 1 and type 2

Usher syndrome (USH) is the most frequent cause of combined deaf-blindness in man. USH is clinically and genetically heterogeneous with at least 11 chromosomal loci assigned to the three USH types (USH1A-G, USH2A-C, USH3A). Although the different USH types exhibit almost the same phenotype in human, the identified USH genes encode for proteins which belong to very different protein classes and families. We and others recently reported that the scaffold protein harmonin (USH1C-gene product) integrates all identified USH1 molecules in a USH1-protein network. Here, we investigated the relationship between the USH2 molecules and this USH1-protein network. We show a molecular interaction between the scaffold protein harmonin (USH1C) and the USH2A protein, VLGR1 (USH2C) and the candidate for USH2B, NBC3. We pinpoint these interactions to interactions between the PDZ1 domain of harmonin and the PDZ-binding motifs at the C-termini of the USH2 proteins and NBC3. We demonstrate that USH2A, VLGR1 and NBC3 are co-expressed with the USH1-protein harmonin in the synaptic terminals of both retinal photoreceptors and inner ear hair cells. In hair cells, these USH proteins are also localized in the signal uptaking stereocilia. Our data indicate that the USH2 proteins and NBC3 are further partners in the supramolecular USH-protein network in the retina and inner ear which shed new light on the function of USH2 proteins and the entire USH-protein network. These findings provide first evidence for a molecular linkage between the pathophysiology in USH1 and USH2. The organization of USH molecules in a mutual 'interactome' related to the disease can explain the common phenotype in USH.     

In vitro hemocompatibility of self-assembled monolayers displaying various functional groups

Self-assembled monolayers (SAMs) of alkanethiols with various terminating groups (-OH, -CH3, -COOH) and binary mixtures of these alkanethiols were studied with respect to their hemocompatibility in vitro by means of freshly taken human whole blood. The set of smooth monomolecular films with graded surface characteristics was applied to scrutinize hypotheses on the impact of surface chemical-physical properties on distinct blood activation cascades, i.e. to analyze -OH surface groups vs. complement activation, acidic surface sites vs. contact activation/coagulation and surface hydrophobicity vs. thrombogenicity. Blood and model surfaces were analyzed after incubation for the related hemocompatibility parameters. Our results show that the adhesion of leukocytes is abolished on a -CH3 surface and greatly enhanced on surfaces with -OH groups. The opposite was detected for the adhesion of platelets. A strong correlation between the activation of the complement system and the adhesion of leukocytes with the content of -OH groups could be observed. The contact activation for hydrophilic surfaces was found to scale with the amount of acidic surface sites. However, the coagulation and platelet activation did not simply correlate with any surface property and were therefore concluded to be determined by a superposition of contact activation and platelet adhesion.     

Metabolic functions of duplicate genes in Saccharomyces cerevisiae

The roles of duplicate genes and their contribution to the phenomenon of enzyme dispensability are a central issue in molecular and genome evolution. A comprehensive classification of the mechanisms that may have led to their preservation, however, is currently lacking. In a systems biology approach, we classify here back-up, regulatory, and gene dosage functions for the 105 duplicate gene families of Saccharomyces cerevisiae metabolism. The key tool was the reconciled genome-scale metabolic model iLL672, which was based on the older iFF708. Computational predictions of all metabolic gene knockouts were validated with the experimentally determined phenotypes of the entire singleton yeast library of 4658 mutants under five environmental conditions. iLL672 correctly identified 96%-98% and 73%-80% of the viable and lethal singleton phenotypes, respectively. Functional roles for each duplicate family were identified by integrating the iLL672-predicted in silico duplicate knockout phenotypes, genome-scale carbon-flux distributions, singleton mutant phenotypes, and network topology analysis. The results provide no evidence for a particular dominant function that maintains duplicate genes in the genome. In particular, the back-up function is not favored by evolutionary selection because duplicates do not occur more frequently in essential reactions than singleton genes. Instead of a prevailing role, multigene-encoded enzymes cover different functions. Thus, at least for metabolism, persistence of the paralog fraction in the genome can be better explained with an array of different, often overlapping functional roles.     

Retinoids induce Fas(CD95) ligand cell surface expression via RARgamma and nur77 in T cells

Cells from the CD4+ murine T hybridoma line IP-12-7 enter the apoptotic suicide program via the Fas ligand (FasL)/Fas-mediated pathway upon TCR stimulation. This stimulus regulates the sensitization of the Fas death pathway and the cell surface appearance of preformed FasL. The apoptosis is dependent on new mRNA and protein synthesis and involves up-regulation of nur77.Two groups of nuclear receptors for retinoic acids (RA) have been identified: retinoic acid receptors (RAR) and retinoid X receptors. IP-12-7 cells express RARalpha and RARgamma. Here we show that,in the IP-12-7 T cells, RA also induced the expression and DNA binding of nur77, and the cell surface appearance of FasL. The induction was mediated via RARgamma. Despite the induced expression of cell surface FasL, only two structurally related RARgamma-selective compounds, CD437 and CD2325, initiated apoptosis in these cells. The lack of apoptosis induction by natural RA was related to the inability of RARgamma to sensitize the Fas death-pathway. Cell surface FasL, however, was able to induce cell death in Fas-bearing target cells. Natural RA also induced the expression of FasL in phytohemagglutinin-activated peripheral murine T cells. It is proposed that therapeutically administered RA might induce apoptosis in Fas-sensitive cells via induction of FasL expression in activated Tcells.     

Evaluation of a resorbable, in situ setting bone substitute in a sheep model

The gold standard for bone substitution is the autologous bone graft, but because of its limited supply and the associated morbidity, the search for synthetic alternatives is necessary. A new in situ setting tricalcium phosphate cement was implanted in a trepanation defect (9.4 mm diameter, 10 mm depth) in the distal femoral epiphysis of sheep. Empty cavities and autologous bone graft were used as controls. Histologic and histomorphometric examinations were carried out after 12 weeks. Nearly 90% of the implanted cement was resorbed and replaced by ingrown bone with close contact between surrounding bone, new bone, and remaining cement particles. The amount of bone in the defect area was significantly higher in defects filled with cement relative to defects filled with autologous bone graft (mean 27 vs. 21%, 95% confidence intervals 23 to 31 and 18 to 23, p = 0.026). In conclusion, this new in situ setting cement is bioactive, resorbable, and osteoconductive. It will be useful as an alternative to autologous bone graft to fill stable defects.     

Age-related changes in the articular cartilage of human sacroiliac joint

 Iliac and sacral articular cartilage of 25 human sacroiliac joints (1–93 years) are examined by light microscopy and immunohistochemistry in order to gain further insight into the nature and progress of degenerative changes appearing during aging. These changes can already be seen in younger adults as compared to cartilage degeneration known in other diarthrodial joints. Structural differences between sacral and iliac cartilage can already be observed in the infant: the sacral auricular facet is covered with a hyaline articular cartilage, reaching 4 mm in thickness in the adult and staining intensely blue with alcian blue at pH1. Iliac cartilage of the newborn is composed of a dense fibrillar network of thick collagen bundles, crossing each other at approximately right angles. A faint staining with alcian blue suggests a low content of acidic glycosaminoglycans. In the adult, iliac cartilage becomes hyaline and its maximal thickness reaches 1–2 mm. Both articular facets exhibit morphological changes during aging that are more pronounced in the iliac cartilage and resemble osteoarthritic degeneration; the staining pattern of the extracellular matrix becomes inhomogenous, chondrocytes are arranged in clusters and the articular surface develops superficial irregularities and fissures. Sometimes fibrous tissue fills up these defects. Nevertheless, large areas of iliac cartilage remain hyaline in nature. Sacral articular cartilage often remains largely unaltered until old age. The sacral subchondral bone plate is usually thin and shows spongiosa trabeculae inserted at right angles, suggesting a perpendicular load on the articular facet. Iliac subchondral spongiosa shows no definite alignment and joins the thickened subchondral bone plate in an oblique direction. The iliac cartilage therefore seems to be stressed predominantly by shearing forces, arising from the changing monopodal support of the pelvis during locomotion. The subchondral bone plate on both the iliac and sacral auricular facet is penetrated by blood vessels that come into close contact with the overlying articular cartilage. These vessels may contribute to the high incidence of rheumatoid and inflammatory diseases in the human sacroiliac joint. Immunolabelling with an antibody against type II collagen reveals a diminished immunoreactivity in the upper half of adult sacral cartilage and only a faint and irregular labelling in the iliac cartilage. Type I collagen can be detected in a superficial layer on the sacral articular surface and around chondrocyte clusters in iliac cartilage, as in dedifferentiating chondrocytes during the development of osteoarthritis. Accepted: 22 April 1998     

Self-organization of a liquid crystalline methacrylate-monofunctionalized crown-ether compound in low-shrinkage acrylate mixtures

Crystallization and supramolecular aggregation of 1,4,7,10,13-pentaoxacyclopentadecane-2-ylmethyl 3,4-bis[4-(dodecyl-1-oxy)benzyloxy]-5-(11-methacryloyloxyundecyl-1-oxy)benzoate ( 1 ) and its complexes with sodium triflate are described in solutions of the methacrylate monomers. Formation of a gel was observed covering a rather wide concentration range in the pseudo-binary phase diagram. In the low concentration regime and at low temperatures, gel formation by elongated crystals of 1 was observed. It was demonstrated that the formation of a crystalline network and the shape of the crystals strongly depend on the cooling rate.     

Expression of Matrix Metalloproteinases and Their Tissue Inhibitors in Human Brain Tumors

In this study, we investigated the expression patterns of 15 matrix metalloproteinases (MMPs) and three tissue inhibitors of metalloproteinase in gliomas, medulloblastomas, and normal brain tissue. By Northern blot analysis we found increased levels of mRNAs encoding for gelatinase A, gelatinase B, two membrane-type MMPs (mt1- and mt2-MMP), and tissue inhibitors of metalloproteinase-1 in glioblastomas and medulloblastomas. We observed a significant increase of mt1-MMP, gelatinase A, gelatinase B, and tissue inhibitors of metalloproteinase-1 in glioblastomas as compared with low-grade astrocytomas, anaplastic astrocytomas, and normal brain. In medulloblastomas, the expression of mt1-MMP, mt2-MMP, and gelatinase A were also increased, but to a lesser extent than that observed in glioblastomas. These data were confirmed at the protein level by immunostaining analysis. Moreover, substrate gel electrophoresis showed that the activated forms of gelatinases A and B were present in glioblastomas and medulloblastomas. These results suggest that increased expression of mt1-MMP/gelatinase A is closely related to the malignant progression observed in gliomas. Furthermore, the present study demonstrates, to our knowledge for the first time, that medulloblastomas express high levels of MMP.