Observation of the phosphatidyl ethanolamine amino proton magnetic resonance in phospholipid vesicles: inside/outside ratios and proton transport.

The amino proton resonance of phosphatidyl ethanolamine in sonicated mixed phospholipid vesicles is observed 3.3 ppm downfield from H₂O. Above pH ～ 5 it is broadened beyond detectability as a result of exchange with H₂O protons. In low salt, resonances of amino protons inside the vesicles appear to persist as the pH is raised, while those on the outside disappear. Solvent catalized proton conduction along the surface is proposed, with an effective -NH₂ to -NH₃ transfer rate of about 8 × 10⁵ sec^?1 at 25°C.     

Interleukin-7-aggravated joint inflammation and tissue destruction in collagen-induced arthritis is associated with T-cell and B-cell activation

Introduction

We sought to investigate the capacity of interleukin (IL)-7 to enhance collagen-induced arthritis and to study by what mechanisms this is achieved.

Methods

Mice received multiple injections with IL-7 or phosphate-buffered saline (PBS) as a control. Arthritis severity and incidence were determined by visual examination of the paws. Joint destruction was determined by assessing radiographs and immunohistochemistry of the ankle joints. Total cellularity and numbers of T-cell and B-cell subsets were assessed, as well as ex vivo production of interferon-γ (IFN-γ), IL-17, and IL-4. Proinflammatory mediators were measured in serum with multianalyte profiling.

Results

IL-7 increased arthritis severity and radiology-assessed joint destruction. This was consistent with IL-7-increased intensity of cell infiltrates, bone erosions, and cartilage damage. Splenic CD19⁺ B cells and CD19⁺/GL7⁺ germinal center B cells, as well as CD4 and CD8 numbers, were increased by IL-7. IL-7 expanded memory T cells, associated with increased percentages of IFN-γ-, IL-4-, and IL-17-producing CD4⁺ T cells. On antigen restimulation of draining lymph node cells in vitro IL-7 treatment was found to increase IFN-γ and IL-17 production, whereas IL-4 was reduced. IL-7 also increased concentrations of proinflammatory mediators, indicative of T-cell activation (sCD40L), vascular activation (VCAM-1, VEGF), tissue destruction (fibroblast growth factor-basic (FGF-b), LIF), and chemotaxis (MIP-1γ, MIP-3β, lymphotactin, MDC, and MCP-5).

Conclusions

In arthritic mice, IL-7 causes expansion of T and B cells, associated with increased levels of proinflammatory mediators. IL-7 intensifies arthritis severity and joint destruction, accompanied by increased Th1 and Th17 activity. These data indicate that IL-7 could be an important mediator in arthritic conditions and that targeting IL-7 or its receptor represent novel therapeutic strategies.     

Actions of a novel synthetic natriuretic peptide on hemodynamics and ventricular function in the dog

Dendroaspis natriuretic peptide (DNP) is a recently discovered peptide with structural similarity to known natriuretic peptides. DNP has been shown to possess potent renal actions. Our objectives were to define the acute hemodynamic actions of DNP in normal anesthetized dogs and the acute effects of DNP on left ventricular (LV) function in conscious chronically instrumented dogs. In anesthetized dogs, DNP, but not placebo, decreased mean arterial pressure (141 +/- 6 to 109 +/- 7 mmHg, P < 0.05) and pulmonary capillary wedge pressure (5.8 +/- 0.3 to 3.4 +/- 0.2 mmHg, P < 0.05). Cardiac output decreased and systemic vascular resistance increased with DNP and placebo. DNP-like immunoreactivity and guanosine 3',5'-cyclic monophosphate concentration increased without changes in other natriuretic peptides. In conscious dogs, DNP decreased LV end-systolic pressure (120 +/- 7 to 102 +/- 6 mmHg, P < 0.05) and volume (32 +/- 6 to 28 +/- 6 ml, P < 0.05) and LV end-diastolic volume (38 +/- 5 to 31 +/- 4 ml, P < 0.05) but not arterial elastance. LV end-systolic elastance increased (6.1 +/- 0.7 to 7.4 +/- 0.6 mmHg/ml, P < 0.05), and Tau decreased (31 +/- 2 to 27 +/- 1 ms, P < 0.05). The effects on hemodynamics, LV function, and second messenger generation suggest synthetic DNP may have a role as a cardiac unloading and lusitropic peptide.     

Loop mobility in a four-helix-bundle protein: 15N NMR relaxation measurements on human interleukin-4.

15N NOE, T1, and T2 measurements have been carried out on uniformly 15N-labeled human interleukin-4. Analysis of the results in terms of order parameters (S2) shows that although the helical core of this four-helix-bundle protein exists as a well-defined structure with limited conformational flexibility (S2 congruent to 0.9), other regions of the molecule experience substantial fluctuations in the conformation of the main chain (S2 = 0.3-0.8). These regions include both the N- and C-termini and two of the loops joining the helices. The majority of these internal motions are fast compared with the overall rotational correlation time (tau R = 7.6 ns at 35 degrees C) and are localized in regions that are relatively ill-defined in the NMR structures previously determined for this protein [Smith, L. J., Redfield, C., Boyd, J., Lawrence, G. M. P., Edwards, R. G., Smith, R. A. G., & Dobson, C. M. (1992) J. Mol. Biol. 224, 899-904]. Other motions are on a slower time scale and appear to be associated with two of the three disulfide bonds and the beta-sheet region in the protein. The dynamic properties of interleukin-4 in solution have been compared with features of the X-ray structures of other four-helix-bundle proteins. The results suggest that the dynamic properties observed here may be general for this class of proteins and may be significant for the interpretation of both their structural and functional properties.     

Structure of cryptic lambda prophages

When Escherichia coli cells lysogenic for bacteriophage lambda are induced with ultraviolet light, cells carrying cryptic lambda prophages are occasionally found among the apparently cured survivors. The lambda variant crypticogen (lambda crg) carries an insertion of the transposable element IS2, which increases the frequency of cryptic lysogens to about 50% of cured cells: 43 of these cryptic prophages have been characterized. They all contain substitutions that replace the early segment of the prophage genome (from the IS2 to near the cos site) with a duplicate copy of a large segment of the host chromosome. The right end of the substitution always results from recombination between the nin-QSR-cos region of the prophage and the homologous incomplete lambdoid prophage Qsr' at 12.5 minutes in the E. coli chromosome. The left end of the substitution is usually a crossover that recombines the IS2 element in the prophage with an E. coli IS2 at 8.5 minutes, near the lac gene, or with a second IS2 located counterclockwise from leu at 2 minutes, generating duplications of at least 200,000 bases. Five cryptic lysogens derived from cells lysogenic for a reference strain of lambda (which lacks the IS2 present in lambda crg) have been characterized. They contain substitutions whose right termini are generated by a crossover with the Qsr' prophage. The left termini of these substitutions are formed either by a crossover between the lambda exo gene and a short exo-homologous segment of Qsr' (2/5), or by a crossover between sequences to the left of attL and an unmapped distant region of the host chromosome (3/5). The large duplications carried by these cryptic lysogens are stable, unlike tandem duplications, and so may significantly influence the cell's evolutionary potential.     

Genomic organization and chromosomal localization of three members of the UDP-N-acetylgalactosamine: polypeptide N- acetylgalactosaminyltransferase family

A homologous family of UDP- N -acetylgalactosamine: polypeptide N - acetylgalactosaminyltransferases (GalNAc-transferases) initiate O- glycosylation. These transferases share overall amino acid sequence similarities of approximately 45-50%, but segments with higher similarities of approximately 80% are found in the putative catalytic domain. Here we have characterized the genomic organization of the coding regions of three GalNAc-transferase genes and determined their chromosomal localization. The coding regions of GALNT1 , -T2 , and -T3 were found to span 11, 16, and 10 exons, respectively. Several intron/exon boundaries were conserved within the three genes. One conserved boundary was shared in a homologous C. elegans GalNAc- transferase gene. Fluorescence in situ hybridization showed that GALNT1 , -T2 , and -T3 are localized at chromosomes 18q12-q21, 1q41-q42, and 2q24-q31, respectively. These results suggest that the members of the polypeptide GalNAc-transferase family diverged early in evolution from a common ancestral gene through gene duplication.