Histological Analysis of Extracranial Carotid Artery Aneurysms

Introduction

Extracranial carotid artery aneurysms (ECAA) are rare but may be accompanied with significant morbidity. Previous studies mostly focused on diagnostic imaging and treatment. In contrast, the pathophysiological mechanisms and natural course of ECAA are largely unknown. Understanding the pathophysiological background may add to prediction of risk for adverse outcome and need for surgical exclusion. The aim of this study was to investigate the histopathological characteristics of ECAA in patients who underwent complete surgical ECAA resection.

Material and Methods

From March 2004 till June 2013, 13 patients were treated with open ECAA repair. During surgery the aneurysm sac was resected and processed for standardized histological analysis. Sections were stained with routine hematoxylin and eosin and special stains to detect elastin, collagen, different types of inflammatory cells, vascular smooth muscle cells and endothelial cells.

Results

Histopathological characterization revealed two distinct categories: dissection (abrupt interruption of the media; n = 3) and degeneration (general loss of elastin fibers in the media; n = 10). In the degenerative samples the elastin fibers in the media were fragmented and were partly absent. Inflammatory cells were observed in the vessel wall of the aneurysms.

Conclusion

Histological analysis in this small sample size revealed dissection and degeneration as the two distinct underlying mechanisms in ECAA formation.     

Improved antigen retrieval in freeze-fracture cytochemistry by evaporation of carbon as first replication layer

The recently developed freeze-fracture replica immunolabeling technique uses sodium dodecyl sulfate to clean replicas obtained from chemically unfixed, rapidly frozen cells by evaporation of platinum as first and carbon as second replication layer. The detergent dissolves remains of cellular material with the exception of components which are in direct contact to the replica film. Membrane lipids and membrane protein complexes of the protoplasmic and the exoplasmic membrane halves remain attached to the replica film and are accessible for cytochemical localization. We immunolabeled the membrane proteins caveolin-1 and connexin 43 in mouse cell lines as well as the membrane attached protein tetrachloroethene reductive dehalogenase (PceA) in bacterial cells at freeze-fracture replicas generated by different evaporation parameters. The labeling experiments for caveolin-1 and the PceA showed that freeze-fracture replication of cellular membranes accomplished with thin platinum layers as well as replication with carbon as first evaporation layer lead in these cases to an improved antigen retrieval, whereas the labeling efficiency of connexin 43 was not affected by different evaporation conditions.     

CRP determination based on a novel magnetic biosensor

The c-reactive protein (CRP) is a very significant human blood marker for inflammatory processes and is routinely determined for many clinical purposes. The widespread and well established detection method for this approximately 115 kDa hepatic protein is the high-sensitivity ELISA assay (hsCRP-ELISA) in blood serum. New approaches in medical CRP diagnosis (e.g. for CVD, inflammatory bowel disease) require rapid quantification in native matrices. A novel CRP determination method based on magnetic detection is described and tested for human blood serum, saliva and urine. The detection principle is based on two different anti-CRP antibodies (monoclonal, IgG) for CRP trapment and labelling. The linear detection range of this immunosensor ranged from 25 ng/ml to 2.5 microg/ml and is therefore much more sensitive than typical hsCRP-ELISA-assays.     

Chlamydophila pneumoniae induces a sustained airway hyperresponsiveness and inflammation in mice

Background

It has been reported that Chlamydophila (C.) pneumoniae is involved in the initiation and promotion of asthma and chronic obstructive pulmonary diseases (COPD). Surprisingly, the effect of C. pneumoniae on airway function has never been investigated.

Methods

In this study, mice were inoculated intranasally with C. pneumoniae (strain AR39) on day 0 and experiments were performed on day 2, 7, 14 and 21.

Results

We found that from day 7, C. pneumoniae infection causes both a sustained airway hyperresponsiveness and an inflammation. Interferon-γ (IFN-γ) and macrophage inflammatory chemokine-2 (MIP-2) levels in bronchoalveolar lavage (BAL)-fluid were increased on all experimental days with exception of day 7 where MIP-2 concentrations dropped to control levels. In contrast, tumor necrosis factor-α (TNF-α) levels were only increased on day 7. From day 7 to 21 epithelial damage and secretory cell hypertrophy was observed. It is suggested that, the inflammatory cells/mediators, the epithelial damage and secretory cell hypertrophy contribute to initiation of airway hyperresponsiveness.

Conclusion

Our study demonstrates for the first time that C. pneumoniae infection can modify bronchial responsiveness. This has clinical implications, since additional changes in airway responsiveness and inflammation-status induced by this bacterium may worsen and/or provoke breathlessness in asthma and COPD.     

Structural and functional analysis of hypothetical proteins in mouse hippocampus from two-dimensional gel electrophoresis

Protein profiling in five individual mouse strains showed strain-specific expression of three hypothetical proteins (HPs). As functional and structural assignment of HPs were based on predictions and low identity to known structures, HPs were identified by MALDI-TOF/TOF, and their proposed tentative function was determined by enzyme assays. Three identified HPs were extracted from gels and renatured, and pyridoxal phosphate phosphatase, inorganic pyrophosphate phosphatase, and antioxidant activities were revealed, findings in agreement with functional predictions.     

The CORVET tethering complex interacts with the yeast Rab5 homolog Vps21 and is involved in endo-lysosomal biogenesis

The dynamic equilibrium between vesicle fission and fusion at Golgi, endosome, and vacuole/lysosome is critical for the maintenance of organelle identity. It depends, among others, on Rab GTPases and tethering factors, whose function and regulation are still unclear. We now show that transport among Golgi, endosome, and vacuole is controlled by two homologous tethering complexes, the previously identified HOPS complex at the vacuole and a novel endosomal tethering (CORVET) complex, which interacts with the Rab GTPase Vps21. Both complexes share the four class C Vps proteins: Vps11, Vps16, Vps18, and Vps33. The HOPS complex, in addition, contains Vps41/Vam2 and Vam6, whereas the CORVET complex has the Vps41 homolog Vps8 and the (h)Vam6 homolog Vps3. Strikingly, the CORVET and HOPS complexes can interconvert; we identify two additional intermediate complexes, both consisting of the class C core bound to Vam6-Vps8 or Vps3-Vps41. Our data suggest that modular assembled tethering complexes define organelle biogenesis in the endocytic pathway.     

Mass spectrometric studies on mouse hippocampal synapsins Ia, IIa, and IIb and identification of a novel phosphorylation site at serine-546

Synapsins are key phosphoproteins in the mammalian brain, and structural research on synapsins is still holding center stage. Proteins were extracted from hippocampal tissue and separated on two-dimensional gel electrophoresis (2-DE), and the spots were analyzed by MALDI-TOF-TOF and nano-LC-ESI-MS/MS. Synapsins Ia, IIa, and IIb were unambiguously identified and represented by 15 individual spots on 2-DE. Several serine phosphorylation sites were confirmed, and a novel phosphorylation site was observed at Ser-546 in synapsin IIa in all gels analyzed.     

Nitric oxide and oxygen radical attack on GDP-dissociation inhibitor 2 (GDI-2) in spinal cord injury of the rat

Protein oxidation and nitration have been described during spinal cord injury (SCI) in animal models. Herein, mass spectrometry unambiguously identified GDP-dissociation inhibitor-2 (GDI-2) in SCI with post-translational modifications of 3-aminotyrosine (8 h post-injury) and an acrolein adduct of GDI-2 (72 h post-injury). On the basis of mass spectrometry evidence, we conclude that lipid-peroxidation and protein nitration do take place on an important signalling protein that may be prevented by specific experimental therapeutic interventions.     

Pharmacokinetics of poly(hydroxyethyl-l-asparagine)-coated liposomes is superior over that of PEG-coated liposomes at low lipid dose and upon repeated administration

‘Stealth’ liposomes with a poly(ethylene glycol) (PEG) coating are frequently studied for drug delivery and diagnostic purposes because of their prolonged blood circulation kinetics. However, several recent reports have demonstrated that PEG-liposomes are rapidly cleared at single low lipid doses (< 1 μmol/kg) and upon repeated administration (time interval between the injections 5 days-4 weeks). Recently, poly(amino acid)-based stealth liposome coatings have been developed as alternative to the PEG-coating. In this study, the pharmacokinetic behavior of liposomes coated with the poly(amino acid) poly(hydroxyethyl-l-asparagine) (PHEA) was evaluated at low lipid doses and upon repeated administration in rats. Blood circulation times and hepatosplenic localization of PHEA-liposomes were assessed after intravenous injection. When administered at a dose of 0.25 μmol/kg or less, PHEA-liposomes showed significantly longer blood circulation times than PEG-liposomes. A second dose of PHEA-liposomes 1 week after the first injection was less rapidly cleared from the circulation than a second dose of PEG-liposomes. Although the mechanisms behind these observations are still not clear yet, the use of PHEA-liposomes appears beneficial when single low lipid doses and/or repeated dosing schedules are being applied.