Osteoclasts secrete osteopontin into resorption lacunae during bone resorption.

Osteopontin (OPN) is a non-collagenous extracellular sialylated glycoprotein located in bone. It is believed to be one of the key components in osteoclast attachment to bone during resorption. In this study, we characterized OPN and other glycoproteins found in the resorption lacunae to confirm the role of osteoclasts in OPN secretion using electron microscopy and mass spectrometry.

An immunocapture-LC-MS-based assay for serum SPINK1 allows simultaneous quantification and detection of SPINK1 variants.

Pancreatic secretory trypsin inhibitor Kazal type 1 (SPINK1) is a 6420 Da peptide produced by the pancreas, but also by several other tissues and many tumors. Some mutations of the SPINK1 gene, like the one causing amino acid change N34S, have been shown to confer susceptibility to recurrent or chronic pancreatitis. Detection of such variants are therefore of clinical utility.

Extracellular o-linked N-acetylglucosamine is enriched in stem cells derived from human umbilical cord blood.

Stem cells have a unique ability to self-renew and differentiate into diverse cell types. Currently, stem cells from various sources are being explored as a promising new treatment for a variety of human diseases. A diverse set of functional and phenotypical markers are used in the characterization of specific therapeutic stem cell populations.

Extracellular membrane vesicles from umbilical cord blood-derived MSC protect against ischemic acute kidney injury, a feature that is lost after inflammatory conditioning.

Background: Mesenchymal stromal cells (MSC) are shown to have a great therapeutic potential in many immunological disorders. Currently the therapeutic effect of MSCs is considered to be mediated via paracrine interactions with immune cells. Umbilical cord blood is an attractive but still less studied source of MSCs.

Production of a recombinant antibody specific for i blood group antigen, a mesenchymal stem cell marker.

Multipotent mesenchymal stem/stromal cells (MSCs) offer great promise for future regenerative and anti-inflammatory therapies. Panels of functional and phenotypical markers are currently used in characterization of different therapeutic stem cell populations from various sources. The i antigen (linear poly-N-acetyllactosamine) from the Ii blood group system has been suggested as a marker for MSCs derived from umbilical cord blood (UCB).

Phosphorylation regulates FOXC2-mediated transcription in lymphatic endothelial cells.

One of the key mechanisms linking cell signaling and control of gene expression is reversible phosphorylation of transcription factors. FOXC2 is a forkhead transcription factor that is mutated in the human vascular disease lymphedema-distichiasis and plays an essential role in lymphatic vascular development. However, the mechanisms regulating FOXC2 transcriptional activity are not well understood.

Interlaboratory study on differential analysis of protein glycosylation by mass spectrometry: the ABRF glycoprotein research multi-institutional study 2012.

One of the principal goals of glycoprotein research is to correlate glycan structure and function. Such correlation is necessary in order for one to understand the mechanisms whereby glycoprotein structure elaborates the functions of myriad proteins. The accurate comparison of glycoforms and quantification of glycosites are essential steps in this direction.

Transient proteolytic modification of mesenchymal stromal cells increases lung clearance rate and targeting to injured tissue.

Systemic infusion of therapeutic cells would be the most practical and least invasive method of administration in many cellular therapies. One of the main obstacles especially in intravenous delivery of cells is a massive cell retention in the lungs, which impairs homing to the target tissue and may decrease the therapeutic outcome. In this study we showed that an alternative cell detachment of mesenchymal stromal/stem cells (MSCs) with pronase instead of trypsin significantly accelerated the lung clearance of the cells and, importantly, increased their targeting to an area of injury.

Metabolic glycoengineering of mesenchymal stromal cells with N-propanoylmannosamine.

There is an increasing interest in the modification of cell surface glycosylation to improve the properties of therapeutic cells. For example, glycosylation affects the biodistribution of mesenchymal stromal cells (MSCs). Metabolic glycoengineering is an efficient way to modify the cell surface.

Nanoscale reversed-phase liquid chromatography-mass spectrometry of permethylated N-glycans.

Reversed-phase liquid chromatography on the nanoscale coupled to electrospray tandem mass spectrometry was used to analyse a mixture of four commercial glycan standards, and the method was further adapted to N-glycans enzymatically released from alpha-1-acid glycoprotein and immunoglobulin gamma. Glycans were permethylated to enable their separation by reversed-phase chromatography and to facilitate interpretation of fragmentation data. Prior to derivatization of glycans by permethylation, they were reduced to cancel anomerism because, although feasible, it was not desired to separate α- and β-anomers.

Cell surface structures influence lung clearance rate of systemically infused mesenchymal stromal cells.

The promising clinical effects of mesenchymal stromal/stem cells (MSCs) rely especially on paracrine and nonimmunogenic mechanisms. Delivery routes are essential for the efficacy of cell therapy and systemic delivery by infusion is the obvious goal for many forms of MSC therapy. Lung adhesion of MSCs might, however, be a major obstacle yet to overcome.

An optimized isolation of biotinylated cell surface proteins reveals novel players in cancer metastasis.

Details of metastasis, the deadliest aspect of cancer, are unclear. Cell surface proteins play central roles in adhesive contacts between the tumor cell and the stroma during metastasis. We optimized a fast, small-scale isolation of biotinylated cell surface proteins to reveal novel metastasis-associated players from an isogenic pair of human MDA-MB-435 cancer cells with opposite metastatic phenotypes.

Novel data analysis tool for semiquantitative LC-MS-MS2 profiling of N-glycans.

Despite recent technical advances in glycan analysis, the rapidly growing field of glycomics still lacks methods that are high throughput and robust, and yet allow detailed and reliable identification of different glycans. LC-MS-MS(2) methods have a large potential for glycan analysis as they enable separation and identification of different glycans, including structural isomers. The major drawback is the complexity of the data with different charge states and adduct combinations.

Comparative metaproteomics and diversity analysis of human intestinal microbiota testifies for its temporal stability and expression of core functions.

The human intestinal tract is colonized by microbial communities that show a subject-specific composition and a high-level temporal stability in healthy adults. To determine whether this is reflected at the functional level, we compared the faecal metaproteomes of healthy subjects over time using a novel high-throughput approach based on denaturing polyacrylamide gel electrophoresis and liquid chromatography-tandem mass spectrometry. The developed robust metaproteomics workflow and identification pipeline was used to study the composition and temporal stability of the intestinal metaproteome using faecal samples collected from 3 healthy subjects over a period of six to twelve months.

The i blood group antigen as a marker for umbilical cord blood-derived mesenchymal stem cells.

Multipotent mesenchymal stem cells (MSCs) offer great promise for future regenerative and anti-inflammatory therapies. However, there is a lack of methods to quickly and efficiently isolate, characterize, and ex vivo expand desired cell populations for therapeutic purposes. Single markers to identify cell populations have not been characterized; instead, all characterizations rely on panels of functional and phenotypical properties.

Mitochondrial function and energy metabolism in umbilical cord blood- and bone marrow-derived mesenchymal stem cells.

Human mesenchymal stem cells (hMSCs) are an attractive choice for a variety of cellular therapies. hMSCs can be isolated from many different tissues and possess unique mitochondrial properties that can be used to determine their differentiation potential. Mitochondrial properties may possibly be used as a quality measure of hMSC-based products.

The binding specificity of the marker antibodies Tra-1-60 and Tra-1-81 reveals a novel pluripotency-associated type 1 lactosamine epitope.

The expression of the epitopes recognized by the monoclonal antibodies Tra-1-60 and Tra-1-81 is routinely used to assess the pluripotency status of human embryonic stem cells (hESCs) and induced pluripotent stem (iPS) cells. Although it is known that the epitopes recognized by Tra-1-60 and Tra-1-81 are carbohydrates, the exact molecular identity of these epitopes has been unclear. Glycan array analysis with more than 500 oligosaccharide structures revealed specific binding of Tra-1-60 and Tra-1-81 to two molecules containing terminal type 1 lactosamine: Galβ1-3GlcNAcβ1-3Galβ1-4GlcNAc and Galβ1-3GlcNAcβ1-3Galβ1-4GlcNAcβ1-6(Galβ1-3GlcNAcβ1-3)Galβ1-4Glc.

Are globoseries glycosphingolipids SSEA-3 and -4 markers for stem cells derived from human umbilical cord blood?

Umbilical cord blood (UCB) is an efficient and valuable source of hematopoietic stem cells (HSCs) for transplantation. In addition to HSCs it harbours low amounts of mesenchymal stem cells (MSCs). No single marker to identify cord blood-derived stem cells, or to indicate their multipotent phenotype, has been characterized so far.

Proteomic analysis of pancreatic secretory trypsin inhibitor/tumor-associated trypsin inhibitor from urine of patients with pancreatitis or prostate cancer.

The development of proteomic methods, especially mass spectrometry, has brought new possibilities to tumor marker research. Pancreatic secretory trypsin inhibitor (PSTI), a common known biomarker for various malignancies, occurs on genetic variants that we are able to detect at the protein level with proteomic techniques using immunoaffinity capture prior to liquid chromatography-mass spectrometry (LC-MS). We also show that PSTI can be detected in urine from cancer patients using a two-step peptide enrichment technique and LC-MS.

Human CMP-N-acetylneuraminic acid hydroxylase is a novel stem cell marker linked to stem cell-specific mechanisms.

Human stem cells contain substantial amounts of the xenoantigen N-glycolylneuraminic acid (Neu5Gc), although the levels of Neu5Gc are low or undetectable in human body fluids and most other human tissues. The lack of Neu5Gc in human tissues has been previously explained by the loss of hydroxylase activity of the human CMP-N-acetylneuraminic acid hydroxylase (CMAH) protein caused by a genetic error in the human Cmah gene. We thus wanted to investigate whether the human redundant Cmah gene could still function in stem cell-specific processes.

Glycomics of bone marrow-derived mesenchymal stem cells can be used to evaluate their cellular differentiation stage.

Human mesenchymal stem cells (MSCs) are adult multipotent progenitor cells. They hold an enormous therapeutic potential, but at the moment there is little information on the properties of MSCs, including their surface structures. In the present study, we analyzed the mesenchymal stem cell glycome by using mass spectrometric profiling as well as a panel of glycan binding proteins.

Structural characterization and anti-angiogenic properties of prostate-specific antigen isoforms in seminal fluid.

Background: The prostate produces high levels of prostate-specific antigen (PSA), which has been shown to exert anti-angiogenic properties and thus might slow down prostate tumor growth. It has been suggested that the protease activity of PSA is not needed for its anti-angiogenic function. We have previously shown that seminal fluid contains both active and inactive, internally cleaved forms of PSA.

Mass spectrometric detection of tyrosine sulfation in human pancreatic trypsinogens, but not in tumor-associated trypsinogen.

Trypsinogen-1 and -2 are well-characterized enzymes that are expressed in the pancreas and also in several other tissues. Many cancers produce trypsinogen isoenzymes that differ from the pancreatic ones with respect to substrate specificity and isoelectric point. These tumor-associated trypsinogens play a pivotal role in cancer progression and metastasis.