Cytokine-induced neutrophil chemoattractant (CINC), a chemotactic molecule of the interleukin (IL)-8 family, is known to be induced in the rat in response to tumor necrosis factor (TNF), IL-1, and lipopolysaccharide (LPS). Intratracheal injection of endotoxin (LPS) is shown to cause CINC mRNA expression in pulmonary tissue, peaking after 2 h, and CINC protein expression in bronchoalveolar lavage (BAL) fluid, peaking after 2-4 h. Intratracheal injection of synthetic CINC causes acute inflammation that is abrogated by coinjection of antiserum to purified natural rat CINC. Intratracheal injection of antiserum to CINC inhibits intratracheal LPS- and IL-1-induced neutrophil emigration into BAL fluid by approximately 60-70%. Despite the anti-inflammatory activity of anti-CINC antiserum, TNF is elevated in the lavage fluid of rats receiving anti-CINC, suggesting that CINC may act in a negative feedback loop to downregulate TNF expression. Intratracheal injection of antiserum to CINC combined with intravenous injection of anti-E-selectin antibody inhibits intratracheal LPS- and IL-1-induced neutrophil emigration into BAL fluid by approximately 75-85%. CINC-mediated chemotactic activity and E-selectin-mediated adherence of neutrophils to endothelium contribute significantly to the pathogenesis of LPS-initiated acute inflammation.

The initiation of graft-vs-host disease (GVHD) after stem cell transplantation is dependent on direct Ag presentation by host APCs, whereas the effect of donor APC populations is unclear. We studied the role of indirect Ag presentation in allogenic T cell responses by adding populations of cytokine-expanded donor APC to hemopoietic grafts that would otherwise induce lethal GVHD. Progenipoietin-1 (a synthetic G-CSF/Flt-3 ligand molecule) and G-CSF expanded myeloid dendritic cells (DC), plasmacytoid DC, and a novel granulocyte-monocyte precursor population (GM) that differentiate into class II+,CD80/CD86+,CD40− APC during GVHD. Whereas addition of plasmacytoid and myeloid donor DC augmented GVHD, GM cells promoted transplant tolerance by MHC class II-restricted generation of IL-10-secreting, Ag-specific regulatory T cells. Importantly, although GM cells abrogated GVHD, graft-vs-leukemia effects were preserved. Thus, a population of cytokine-expanded GM precursors function as regulatory APCs, suggesting that G-CSF derivatives may have application in disorders characterized by a loss of self-tolerance.

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Electrospray and tandem mass spectrometry are used to characterize underivatized oligosaccharides that have been digested from asparagine side chains of glycoproteins. Oligosaccharides that contain sialic acids were detected with the best sensitivity in the negative-ion detection mode whereas those that do not contain sialic acid were detected with the best sensitivity in the positive-ion detection mode. The positive-ion abundances of oligosaccharides were greatly enhanced in electrospray mass spectra by adding 10 mM sodium acetate or ammonium acetate to the sample solvent. Tandem mass spectrometry was used to determine primary structural features of the oligosaccharides. Methodology that has been developed on branched high-mannose, hybrid, and complex carbohydrate standards was applied to a mixture of oligosaccharides that were digested with N-glycanase from the glycoprotein, ovalbumin. The composition and relative abundances of individual oligosaccharides obtained from the electrospray mass spectrum compare favorably to those obtained by anion-exchange chromatography/pulsed amperometric detection and by gel permeation chromatography of the oligosaccharides after radiolabelling the reducing end of the carbohydrates. The oligosaccharide content of ovalbumin was independently determined from the heterogeneity observed in the electrospray mass spectrum of the intact 44-kDa glycoprotein. Comparison of the oligosaccharide compositions determined before and after enzymatic digestion shows a selective digestion of high-mannose and low molecular weight oligosaccharides by N-glycanase.