Statistical shape model-based prediction of tibiofemoral cartilage

Abstract

Computed tomography is used more routinely to design patient-specific instrumentation for knee replacement surgery. Its moderate imaging cost and simplified segmentation reduce design costs compared with magnetic resonance (MR) imaging, but it cannot provide the necessary cartilage information. Our method based on statistical shape modelling proved to be successful in predicting tibiofemoral cartilage in leave-one-out experiments. The obtained accuracy of 0.54?mm for femur and 0.49?mm for tibia outperforms the average cartilage thickness distribution and reported inter-observer MR segmentation variability. These results suggest that shape modelling is able to predict tibiofemoral cartilage with sufficient accuracy to design patient-specific instrumentation.     

Poly-beta-hydroxybutyrate-accumulating bacteria protect gnotobiotic Artemia franciscana from pathogenic Vibrio campbellii

A poly-beta-hydroxybutyrate (PHB)-accumulating enrichment culture was obtained using activated sludge from a polyphosphate-accumulating reactor as inoculum. PHB accumulated by the enrichment culture significantly enhanced the survival of Artemia nauplii, infected with the virulent pathogen Vibrio campbellii LMG 21363. A strain was isolated from the enrichment culture, based on its ability to accumulate PHB, and 16S rRNA gene sequencing of the isolate revealed 99% sequence similarity to Brachymonas denitrificans AS-P1. The isolate, named PHB2, showed good PHB-accumulating activity (up to 32% of the cell dry weight). PHB accumulated by isolate PHB2 was able to protect Artemia completely from the V. campbellii strain. Our data indicate that PHB-accumulating bacteria, such as B. denitrificans PHB2, could be used as an an effective and economically interesting alternative strategy to control infections in aquaculture.     

The area composita of adhering junctions connecting heart muscle cells of vertebrates - III: assembly and disintegration of intercalated disks in rat cardiomyocytes growing in culture

For cell and molecular biological studies of heart formation and function cell cultures of embryonal, neonatal or adult hearts of various vertebrates, notably rat and chicken, have been widely used. As the myocardium-specific cell-cell junctions, the intercalated disks (ID), have recently been found to be particularly sensitive to losses of - or mutations in - certain cytoskeletal proteins, resulting in cardiac damages, we have examined the ID organization in primary cultures of cardiomyocytes obtained from neonatal rats. Using immunofluorescence and immunoelectron microscopy, we have studied the major ID components for up to 2 weeks in culture, paying special attention to spontaneously beating, individual cardiomyocytes and myocardial cell colonies. While our results demonstrate the formation of some ID-like cardiomyocyte-connecting junction arrays, they also reveal a variety of structural disorders such as rather extended, junction-free ID regions, sac-like invaginations and endocytotic blebs as well as accumulations of intracytoplasmic structures suggestive of endocytosed forms of junction-derived vesicles or of junction fragments resembling fascia adhaerens elements. Moreover, we have noticed a novel type of small, obviously plaque-free cytoplasmic vesicles containing one or both of the desmosomal cadherins, desmocollin Dsc2 and desmoglein Dsg2. We conclude that cardiomyocyte cultures are useful model systems for studies of certain aspects of myocardiac differentiation and functions but, on the other hand, show progressive disintegration and deterioration. The potential value of molecular markers and reagents in studies of myocardial pathology as well as in the monitoring of myocardial differentiation of so-called stem cells is discussed.     

Can metabolomics improve tuberculosis diagnostics?

With one third of the global human population suffering from tuberculosis (TB) in 2001, the disease was officially declared a worldwide emergency by the World Health Organization. Since then this disease has grown to epidemic proportions and the recent emergence of drug resistance, the rising incidence of HIV/TB co-infection, and the inability of the currently used vaccination, diagnostic and treatment protocols to control this pandemic, has made TB a research topic inviting urgent attention. The implementation of metabolomics, a relatively new research approach, which is defined as the study of all the small molecular weight compounds or metabolites in a system or sample, using highly sensitive and specific analytical techniques, shows promise in the quest to eradicate this disease. In this context, we describe here the advantages and limitations of the currently available TB diagnostic techniques, and the role that metabolomics has played in the identification of new biomarkers, not only leading to innovative approaches for TB diagnostics, but also to a better understanding of the intra-host changes induced by Mycobacterium tuberculosis infection and active disease.     

Pro-inflammatory mechanisms of muscarinic receptor stimulation in airway smooth muscle

Background

Acetylcholine, the primary parasympathetic neurotransmitter in the airways, plays an important role in bronchoconstriction and mucus production. Recently, it has been shown that acetylcholine, by acting on muscarinic receptors, is also involved in airway inflammation and remodelling. The mechanism(s) by which muscarinic receptors regulate inflammatory responses are, however, still unknown.

Methods

The present study was aimed at characterizing the effect of muscarinic receptor stimulation on cytokine secretion by human airway smooth muscle cells (hASMc) and to dissect the intracellular signalling mechanisms involved. hASMc expressing functional muscarinic M₂ and M₃ receptors were stimulated with the muscarinic receptor agonist methacholine, alone, and in combination with cigarette smoke extract (CSE), TNF-α, PDGF-AB or IL-1β.

Results

Muscarinic receptor stimulation induced modest IL-8 secretion by itself, yet augmented IL-8 secretion in combination with CSE, TNF-α or PDGF-AB, but not with IL-1β. Pretreatment with GF109203X, a protein kinase C (PKC) inhibitor, completely normalized the effect of methacholine on CSE-induced IL-8 secretion, whereas PMA, a PKC activator, mimicked the effects of methacholine, inducing IL-8 secretion and augmenting the effects of CSE. Similar inhibition was observed using inhibitors of IκB-kinase-2 (SC514) and MEK1/2 (U0126), both downstream effectors of PKC. Accordingly, western blot analysis revealed that methacholine augmented the degradation of IκBα and the phosphorylation of ERK1/2 in combination with CSE, but not with IL-1β in hASMc.

Conclusions

We conclude that muscarinic receptors facilitate CSE-induced IL-8 secretion by hASMc via PKC dependent activation of IκBα and ERK1/2. This mechanism could be of importance for COPD patients using anticholinergics.     

A lumenal domain-dependent pathway for sorting to intralumenal vesicles of multivesicular endosomes involved in organelle morphogenesis

Cargo partitioning into intralumenal vesicles (ILVs) of multivesicular endosomes underlies such cellular processes as receptor downregulation, viral budding, and biogenesis of lysosome-related organelles such as melanosomes. We show that the melanosomal protein Pmel17 is sorted into ILVs by a mechanism that is dependent upon lumenal determinants and conserved in non-pigment cells. Pmel17 targeting to ILVs does not require its native cytoplasmic domain or cytoplasmic residues targeted by ubiquitylation and, unlike sorting of ubiquitylated cargo, is insensitive to functional inhibition of Hrs and ESCRT complexes. Chimeric protein and deletion analyses indicate that two N-terminal lumenal subdomains are necessary and sufficient for ILV targeting. Pmel17 fibril formation, which occurs during melanosome maturation in melanocytes, requires a third lumenal subdomain and proteolytic processing that itself requires ILV localization. These results establish an Hrs- and perhaps ESCRT-independent pathway of ILV sorting by lumenal determinants and a requirement for ILV sorting in fibril formation.     

Heterotypic complex formation between subunits of microtubule-associated proteins 1A and 1B is due to interaction of conserved domains

The microtubule-associated proteins MAP1A and MAP1B are related but distinct multi-subunit protein complexes that consist of heavy and light chains. The predominant forms of these complexes are homotypic, i.e. they consist of a MAP1A heavy chain associated with MAP1A light chains or a MAP1B heavy chain associated with MAP1B light chains, respectively. In addition, MAP1A and MAP1B can exchange subunits and form heterotypic complexes consisting of a MAP1A heavy chain associated with MAP1B light chains which might play a role in a transition period of neuronal differentiation. Here we extend previous findings by confirming that heterotypic MAP1B heavy chain-MAP1A light chain complexes also exist in the developing murine brain. We show that these complexes form through interaction of homologous domains conserved in heavy and light chains of MAP1A and MAP1B. Likewise, conserved domains of the MAP1A and MAP1B light chains account for formation of light chain heterodimers. By yeast 2-hybrid analysis we located the light chain binding domain on the heavy chain to amino acids 211-508, thereby defining a new functional subdomain.     

Geographical and ecological distribution of marine halacarid genera and species (Acari: Halacaridae)

At the end of 2002, the number of marine halacarid species was 1018, that of genera 51. A single genus, Copidognathus contains 33% of all species (336). Eleven genera are monotypic. Geographical provinces with a large number of species are the tropical western Pacific, temperate northeastern Atlantic, temperate southeastern Pacific, and Mediterranean-Black Sea. Most records of halacarid species are from temperate and tropical areas; 10% of species are known from polar zones. On a generic level, 29 genera are recorded from tropical and temperate but not from polar provinces, five genera are restricted to the tropics, and none to polar regions. The majority (920 species or 90%) of all species live in the upper 200 m. Records of genera with exclusively algivorous or brackish/fresh water species are bound to littoral habitats; all the other genera occur in more than one depth zone. Arenicolous genera, though most abundant in the littoral zone, have representatives in the bathyal. Four marine genera (Copidognathus, Halacarellus, Isobactrus, Lohmannella) have representatives in coastal fresh water, and three genera, Acarothrix, Caspihalacarus and Peregrinacarus, are predominantly inhabitants of diluted brackish and fresh water. None of the free-living halacarid genera of the world's oceans appears to be endemic to one geographical province.     

Rer1p competes with APH-1 for binding to nicastrin and regulates gamma-secretase complex assembly in the early secretory pathway

The gamma-secretase complex, consisting of presenilin, nicastrin, presenilin enhancer-2 (PEN-2), and anterior pharynx defective-1 (APH-1) cleaves type I integral membrane proteins like amyloid precursor protein and Notch in a process of regulated intramembrane proteolysis. The regulatory mechanisms governing the multistep assembly of this "proteasome of the membrane" are unknown. We characterize a new interaction partner of nicastrin, the retrieval receptor Rer1p. Rer1p binds preferentially immature nicastrin via polar residues within its transmembrane domain that are also critical for interaction with APH-1. Absence of APH-1 substantially increased binding of nicastrin to Rer1p, demonstrating the competitive nature of these interactions. Moreover, Rer1p expression levels control the formation of gamma-secretase subcomplexes and, concomitantly, total cellular gamma-secretase activity. We identify Rer1p as a novel limiting factor that negatively regulates gamma-secretase complex assembly by competing with APH-1 during active recycling between the endoplasmic reticulum (ER) and Golgi. We conclude that total cellular gamma-secretase activity is restrained by a secondary ER control system that provides a potential therapeutic value.