Clostridium perfringens alpha-toxin induces the release of IL-8 through a dual pathway via TrkA in A549 cells

A characteristic feature of gas gangrene with Clostridium perfringens (C. perfringens) is the absence of neutrophils within the infected area and the massive accumulation of neutrophils at the vascular endothelium around the margins of the necrotic region. Intravenous injection of C. perfringens alpha-toxin into mice resulted in the accumulation of neutrophils at the vascular endothelium in lung and liver, and release of GRO/KC, a member of the CXC chemokine family with homology to human interleukin-8 (IL-8). Alpha-toxin triggered activation of signal transduction pathways causing mRNA expression and production of IL-8, which activates migration and binding of neutrophils, in A549 cells. K252a, a tyrosine kinase A (TrkA) inhibitor, and siRNA for TrkA inhibited the toxin-induced phosphorylation of TrkA and production of IL-8. In addition, K252a inhibited the toxin-induced phosphorylation of extracellular regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK). PD98059, an ERK1/2 inhibitor, depressed phosphorylation of ERK1/2 and nuclear translocation of nuclear factor kappa B (NF-κB) p65, but SB203580, a p38 MAPK inhibitor, did not. On the other hand, PD98059 and SB203580 suppressed the toxin-induced production of IL-8. Treatment of the cells with PD98059 resulted in inhibition of IL-8 mRNA expression induced by the toxin and that with SB203580 led to a decrease in the stabilization of IL-8 mRNA. These results suggest that alpha-toxin induces production of IL-8 through the activation of two separate pathways, the ERK1/2/NF-κB and p38 MAPK pathways.     

Analysis of urinary prostacyclin and thromboxane/prostacyclin ratio in patients with rheumatoid arthritis using gas chromatography/selected ion monitoring.

We investigated production of prostacyclin and the urinary ratio of thromboxane and prostacyclin in patients with rheumatoid arthritis. The prostacyclin production level was assessed according to the level of urinary 2,3-dinor-6-keto-prostaglandin F(1 alpha)measuring by gas chromatography/selected ion monitoring. In patients receiving medication, the prostacyclin level was lower and the thromboxane/prostacyclin ratio was greater compare with that of healthy volunteers. The prostacyclin level in patients without medication was approximately 4-fold higher than that of healthy volunteers and 8-fold higher than those of medicated groups. Although the ratio of the group without medication was similar to that of healthy volunteers, the urinary levels of each prostanoid were higher than those of other groups. Then, the ratios of groups receiving steroids were higher than that of other groups owing to high TX level. The present findings demonstrated that endogenous prostacyclin and thromboxane production increased in patients without medication, and prostacyclin production decreased with medication.     

Hydrolysis of fourteen native dextrans by arthrobacter isomaltodextranase and correlation with dextran structure

The isomaltodextranase (EC 3.2.1.94) from Arthrobacter globiformis T6 hydrolysed thirteen dextrans to various extents (11?64% after 13 days) at initially large but gradually decreasing rates. Dextran B-1355 fraction S was, unlike the other dextrans, hydrolysed by the dextranase initially at the lowest rate among the dextrans used, but the rate was maintained for a long period with little decrease, so that the hydrolysis reached as high as 85% after 13 days. Paper chromatography of these dextran digests revealed that this dextranase produces in addition to isomaltose, one or two trisaccharides [isomaltose residues substituted by (1 →2)-, (1→3)-, or (1→4)-α-D-glucopyranosyl groups at the non-reducing D-glucopyranosyl residues] from every dextran used. It is evident that the non-(1→6)-linkages of these trisaccharide products constitute the “anomalous” linkages of the corresponding dextrans. The relative amounts of these trisaccharide products appear to indicate the approxima te relative amounts of a particular linkage among the dextrans, or the relative amounts of two kinds of linkages of each dextran. The kinds and the relative amounts of “anomalous” linkages of some dextrans were established on the basis of the trisaccharides produced by isomaltodextranase.     

Antibody-nucleic acid interactions. Monoclonal antibodies define different antigenic domains in 2',5'-oligoadenylates

To define the epitopes involved in binding anti-oligonucleotide antibodies, several hybridomas producing monoclonal antibodies directed against 2',5'-oligoadenylate were established. A solid-phase enzyme-linked immunoassay that employed microtiter wells coated with Ficoll-2',5'-oligoadenylate conjugates proved useful in screening and characterizing hybridoma supernatants. Control experiments demonstrated that the conjugates were irreversibly adsorbed to polystyrene wells under the conditions employed in the assay. Reactivity of monoclonal antibodies with numerous analogues of 2',5'-oligoadenylate was measured by using a competition assay. Several monoclonal antibodies originating from different mice immunized with the same or different immunogens possessed distinctive fine specificities. At least one 2',5'-phosphodiester bond was important in forming each epitope, suggesting that the ribose phosphate backbone is a critical element in defining an antigenic domain of an oligonucleotide. The purine bases were also important, and modification of the bases had varied effects on the extent of antibody recognition. The length of the oligonucleotide and the nature of the termini were also of some importance. In several instances the modification created by linkage of 2',5'-oligoadenylate to carrier protein also contributed to the determinant. The monoclonal antibody most specific for 2',5'-oligoadenylates was relatively insensitive to ionic strength. In contrast, a monoclonal antibody with a 2',5'-oligopurine specificity appeared to bind 2',5'-oligoadenylate through one ion pair, whereas the binding of a monoclonal antibody with a low degree of base specificity appeared to bind through two ion pairs. The results demonstrated that 2',5'-linked oligoadenylate-protein complexes possess at least three distinct oligonucleotide-related antigenic surfaces that can be recognized with high apparent affinity by monoclonal antibodies. A model for the three epitopes is presented.     

OmpF channel permeability of quinolones and their comparison with β-lactams

The diffusion rates of nalidixic acid, ofloxacin and ofloxacin's two optically active isomers through OmpF channels were measured in proteoliposomes and compared with the rates of beta-lactams. The four quinolones showed high diffusion rates, exceeding that of cephaloridine and being comparable to imipenem. There was no significant difference in diffusion rate between nalidixic acid and ofloxacin, or between the two optically active isomers. The diffusion rates of enoxacin and norfloxacin were also estimated to be higher than many beta-lactams.     

Facile and stereoselective synthesis of 2'-5'-oligothioadenylate by UO2(2+) ion catalyst.

The synthesis and characterization of 2'-5'-oligothioadenylate by UO2(2+) ion catalyst are described. The polymerization of imidazole-activated thioadenylate or thioinosylate yielded oligomers containing mainly 2'-5'-internucleotidyl blond and Rp configuration at phosphorous atom.     

Synthesis and properties of oligodeoxynucleotides containing a C-2 polyamine-bearing deoxyguanosine residue as artificial ribonuclease.

Oligodeoxynucleotides containing a 2-fluoro-2'-deoxyinosine residue substituting normal 2'-deoxyguanosine residue were synthesized. Upon treating with ethanol solution of polyamine, the fluorine atom in the oligomers were readily substituted with the polyamine. The thermal stabilities of the duplexes consisted of the polyamine-bearing oligomers and their cDNAs as well as their RNA cleaving activity were investigated.