Galectin-1-binding glycoforms of haptoglobin with altered intracellular trafficking, and increase in metastatic breast cancer patients.

Sera from 25 metastatic breast cancer patients and 25 healthy controls were subjected to affinity chromatography using immobilized galectin-1. Serum from the healthy subjects contained on average 1.2 mg per ml (range 0.7-2.2) galectin-1 binding glycoproteins, whereas serum from the breast cancer patients contained on average 2.2 mg/ml (range 0.8-3.9), with a higher average for large primary tumours. The major bound glycoproteins were α-2-macroglobulin, IgM and haptoglobin. Both the IgM and haptoglobin concentrations were similar in cancer compared to control sera, but the percentage bound to galectin-1 was lower for IgM and higher for haptoglobin: about 50% (range 20-80) in cancer sera and about 30% (range 25-50) in healthy sera. Galectin-1 binding and non-binding fractions were separated by affinity chromatography from pooled haptoglobin from healthy sera. The N-glycans of each fraction were analyzed by mass spectrometry, and the structural differences and galectin-1 mutants were used to identify possible galectin-1 binding sites. Galectin-1 binding and non-binding fractions were also analyzed regarding their haptoglobin function. Both were similar in forming complex with haemoglobin and mediate its uptake into alternatively activated macrophages. However, after uptake there was a dramatic difference in intracellular targeting, with the galectin-1 non-binding fraction going to a LAMP-2 positive compartment (lysosomes), while the galectin-1 binding fraction went to larger galectin-1 positive granules. In conclusion, galectin-1 detects a new type of functional biomarker for cancer: a specific type of glycoform of haptoglobin, and possibly other serum glycoproteins, with a different function after uptake into tissue cells.

Bacterial outer membrane vesicles in disease and preventive medicine.

Gram-negative bacteria have the ability to produce outer membrane-derived vesicles (OMVs) that are released into the extracellular milieu. Even though this intriguing phenomenon is well-known since many years, various aspects of bacterial OMVs are not fully described and are still in the process of being characterized in detail. One major reason for this is that depending on the bacterial species and its respective ecological niche, OMVs exhibit an enormous functional diversity. Research of the past years has clearly shown that OMVs of many pathogenic bacteria contribute to the virulence potential by enriching virulence factors and delivering them over long distances, superseding direct bacterial contact with their host. The subsequent interaction of OMVs with the host can occur at different levels regarding the type of immune response or the target cell type and may lead to different outcomes ranging from non-immunogenic activation or a pro-inflammatory response to cytotoxicity. In contrast to being virulence factors, OMVs are used for vaccination purposes in the combat against bacterial pathogens, and recent research thus is focused on to indirectly aim these versatile bacterial weapons against themselves.

Multicomponent Moraxella catarrhalis outer membrane vesicles induce an inflammatory response and are internalized by human epithelial cells.

Moraxella catarrhalis is an emerging human respiratory pathogen in patients with chronic obstructive pulmonary disease (COPD) and in children with acute otitis media. The specific secretion machinery known as outer membrane vesicles (OMVs) is a mechanism by which Gram-negative pathogens interact with host cells during infection. We identified 57 proteins in M. catarrhalis OMVs using a proteomics approach combining two-dimensional SDS-PAGE and MALDI-TOF mass spectrometry analysis. The OMVs contained known surface proteins such as ubiquitous surface proteins (Usp) A1/A2, and Moraxella IgD-binding protein (MID). Most of the proteins are adhesins/virulence factors triggering the immune response, but also aid bacteria to evade the host defence. FITC-stained OMVs bound to lipid raft domains in alveolar epithelial cells and were internalized after interaction with Toll-like receptor 2 (TLR2), suggesting a delivery to the host tissue of a large and complex group of OMV-attributed proteins. Interestingly, OMVs modulated the pro-inflammatory response in epithelial cells, and UspA1-bearing OMVs were found to specifically downregulate the reaction. When mice were exposed to OMVs, a pulmonary inflammation was clearly seen. Our findings indicate that Moraxella OMVs are highly biologically active, transport main bacterial virulence factors and may modulate the epithelial pro-inflammatory response.