Phylogenetic study of the species within the family Streptomycetaceae.

Species of the genus Streptomyces, which constitute the vast majority of taxa within the family Streptomycetaceae, are a predominant component of the microbial population in soils throughout the world and have been the subject of extensive isolation and screening efforts over the years because they are a major source of commercially and medically important secondary metabolites. Taxonomic characterization of Streptomyces strains has been a challenge due to the large number of described species, greater than any other microbial genus, resulting from academic and industrial activities. The methods used for characterization have evolved through several phases over the years from those based largely on morphological observations, to subsequent classifications based on numerical taxonomic analyses of standardized sets of phenotypic characters and, most recently, to the use of molecular phylogenetic analyses of gene sequences. The present phylogenetic study examines almost all described species (615 taxa) within the family Streptomycetaceae based on 16S rRNA gene sequences and illustrates the species diversity within this family, which is observed to contain 130 statistically supported clades, as well as many unsupported and single member clusters. Many of the observed clades are consistent with earlier morphological and numerical taxonomic studies, but it is apparent that insufficient variation is present in the 16S rRNA gene sequence within the species of this family to permit bootstrap-supported resolution of relationships between many of the individual clusters.

Preferential expression of B7.2 (CD86), but not B7.1 (CD80), on B cells induced by CD40/CD40L interaction is essential for anti-DNA autoantibody production in patients with systemic lupus erythematosus.

OBJECTIVE: B7 (CD80/CD86) molecules are over-expressed in patients with SLE. However, it is not clear whether CD80/CD86 molecules are involved in the pathogenic autoantibody production specifically or in the polyclonal antibody production in human SLE. The present study was carried out to characterize B7 molecules on B cells in autoantibody production. METHODS: Expression of costimulatory molecules was analyzed by RT-PCR and two-color immunofluorescence staining. Purified B cells were co-cultured with T cells in the presence of anti-costimulatory molecule antibody. RESULTS: Excessive expression of CD86 and CD80 molecules was evident on freshly isolated B cells in patients with SLE. Normal B cells did not express CD86 molecules spontaneously and expressed it after co-culture with activated T cells. CD86 expression on normal and SLE B cells induced by the activated T cells was inhibited by the addition of anti-CD40L into the cell culture. Furthermore, CD40L expression on T cells upon activation was enhanced in SLE patients. Anti-DNA antibody production by SLE B cells in the presence of activated T cells was markedly inhibited by anti-CD86, but not anti-CD80. Anti-CD86 treatment inhibited polyclonal Ig and anti-SS-A antibody production of SLE B cells, suggesting the preferential involvement of CD86 in polyclonal antibody production. CONCLUSION: SLET T cells express CD40L excessively, and the CD40/CD40L pathway is involved in the CD86 over-expression of SLE B cells; thus T cell abnormality is at least partially involved in B cell hyperactivity. Enhanced CD86 expression of B cells by CD40L is essential for polyclonal antibody production.

Aberrant expression of Fas ligand on anti-DNA autoantibody secreting B lymphocytes in patients with systemic lupus erythematosus: "immune privilege"-like state of the autoreactive B cells.

BACKGROUND: Fas/Fas ligand (FasL) system has been assigned a pivotal role in the development and maintenance of peripheral tolerance, and mice with defects in their Fas/FasL system develop lupus-like symptoms. In this study we examined FasL expression of peripheral blood lymphocytes in patients with systemic lupus erythematosus (SLE). METHODS: We assessed FasL mRNA and protein expression by reverse transcription (RT)-PCR and immunoblotting and immunocytochemical staining, respectively, in patients with SLE. Anti-DNA antibody secreting B cells were purified using biotin labeled DNA and streptavidin-bead. RESULTS: Expression of FasL protein was not or very weakly detected in freshly isolated PBMC in normal individuals. In contrast, freshly isolated SLE PBMC exhibited the enhanced expression of FasL protein without in vitro stimulation. Not only purified T cells but also purified B cells expressed FasL on their cell surface spontaneously. In addition, freshly isolated anti-DNA autoantibody secreting B cells express FasL without in vitro stimulation. CONCLUSION: The results suggest that autoreactive B lymphocytes which aberrantly express FasL may kill Fas+ immunoregulatory T lymphocytes. Thus aberrantly expressed FasL may facilitate escape of the autoreactive B cells from the immune tolerance system, and may contribute to the sustained secretion of autoantibodies in patients with SLE.

An inhibitor of inducible nitric oxide synthase decreases forearm blood flow in patients with congestive heart failure.

OBJECTIVES: The functional activation of inducible nitric oxide synthase (iNOS) was evaluated as a source of nitric oxide (NO) in the forearm of patients with heart failure. BACKGROUND: Although endogenous NO is normally produced by constitutive NO synthase (cNOS) in patients with congestive heart failure (CHF), expression of iNOS provides an additional source of NO. However, there are no in vivo studies showing functional activation of iNOS in humans. METHODS: A nonselective NOS inhibitor, N(G)-monomethyl-L-arginine (L-NMMA), and a selective inhibitor of iNOS, aminoguanidine, were administered intra-arterially in graded doses into the brachial arteries of 13 patients with CHF and 10 normal control subjects. Forearm blood flow (FBF) was measured simultaneously in the infused and noninfused arms by plethysmography. Arterial and venous plasma concentrations of nitrite/nitrate (NOx) were measured at baseline and at the highest dose of each drug. RESULTS: L-NMMA significantly reduced the FBF ratio between the infused and noninfused arms in both the control and patient groups (35 +/- 12% and 34 +/- 10%, respectively; both p < 0.001). Aminoguanidine at the same concentration significantly reduced the ratio in the patient group (15 +/- 9%, p < 0.01), with no change in the control group. The arterial NOx concentration was not affected by either drug; however, venous NOx concentrations were significantly decreased in both the control and patient groups by L-NMMA (18 +/- 5% and 18 +/- 17%, respectively; both p < 0.05) and in the patient group only by aminoguanidine (7 +/- 6%, p < 0.05). CONCLUSIONS: These findings suggest that NO production in the forearms of patients with CHF is induced partly by iNOS activation, whereas in normal subjects, it can be ascribed to cNOS activation.

Endorphin and enkephalin ameliorate excessive synovial cell functions in patients with rheumatoid arthritis.

OBJECTIVE: To determine whether endorphin (END) and enkephalin (ENK) modulate excessive synovial cell functions in patients with rheumatoid arthritis (RA). METHODS: Effects of leucine-enkephalin (leu-ENK), methionine-enkephalin (met-ENK), and beta-endorphin (END) on proinflammatory cytokine and matrix metalloproteinase (MMP) production by RA synovial cells were analyzed by immunoblotting, and their mRNA expression by reverse transcription-polymerase chain reaction (RT-PCR) using limiting dilution of complementary DNA. Expression of opioid receptors on RA synovial cells was assessed by immunohistochemical staining, radioreceptor assay, and RT-PCR. RESULTS: Leu-ENK, met-ENK, and END inhibited tumor necrosis factor-alpha and interleukin 1beta production at the level of mRNA expression. ENK and END inhibited MMP-9 production and its enzymatic activity by RA synovial cells. The mu-subtype opioid receptor was expressed in the RA synovial lining and sublining cells. Radioreceptor assay suggested expression of high affinity receptor for END on RA synovial cells. The mu-subtype opioid receptor-specific antagonist, naloxone, reversed the inhibitory effect of the opioid peptides. The opioid peptides inhibited nuclear translocation and phosphorylation of the transcription factor, cyclic AMP responsive element binding protein (CREB) in RA synovial cells. CONCLUSION: Leu-ENK, met-ENK, and END inhibited excessive RA synovial cell functions in vitro. The opioid hormones may have not only antinociceptive action, but also antiinflammatory effects on synovitis itself in RA.

Diagnosis of cardiac sarcoidosis and evaluation of the effects of steroid therapy by gadolinium-DTPA-enhanced magnetic resonance imaging.

BACKGROUND: Cardiac involvement is an important prognostic factor in patients with sarcoidosis. In this study, we evaluated the usefulness of gadolinium-DTPA (diethylene triamine pentaacetic acid)-enhanced magnetic resonance imaging (Gd-MRI) for diagnosing cardiac sarcoidosis and evaluating the effects of steroid therapy. METHODS: Sixteen patients with sarcoidosis diagnosed by histology or by Japanese Ministry of Health and Welfare criteria for cardiac sarcoidosis underwent Gd-MRI with a 1.5-Tesla superconducting magnet system using a T1-weighted spin-echo sequence. RESULTS: Gd-MRI showed localized enhancement of signal intensity, indicating interstitial edema, in the left ventricle in 8 of the 16 patients. Two patients with enhancement also had thinning of the left ventricular septal wall. After 1 month of prednisolone therapy (60 mg every other day or 30 to 40 mg every day), the localized high-intensity signals were markedly diminished in all 8 patients. CONCLUSIONS: Images of the heart obtained by Gd-MRI may reflect active inflammation with interstitial edema in patients with sarcoidosis. Gd-MRI may be a useful noninvasive method for early detection of cardiac sarcoidosis and for evaluating the effects of steroid therapy.

Atrial fibrillation impairs endothelial function of forearm vessels in humans.

BACKGROUND: Although there have been many studies on the effects of atrial fibrillation (AF) on cardiac function, few studies have been done on its effects on endothelial function. The present study was designed to examine the effects of AF on endothelial function in human subjects. METHODS AND RESULTS: Changes in forearm blood flow (FBF) induced by acetylcholine and nitroglycerin were measured by using plethysmography in 14 patients with lone AF, 13 patients with AF and underlying heart disease, and 12 normal control subjects. In the patients, these measurements were repeated after cardioversion. Although baseline FBF was the same in the 3 groups, acetylcholine-induced increases in FBF were significantly smaller in both patient groups than in the control group, and FBF increases were particularly depressed in AF patients with underlying heart disease. After restoration of sinus rhythm by cardioversion, FBF response to the highest dose of acetylcholine increased by 46% in patients with lone AF (n = 10) and by 90% in AF patients with underlying heart disease (n = 11). Nitroglycerin-induced vasodilatation was the same in all 3 groups and was not affected by cardioversion. CONCLUSIONS: These findings suggest that endothelium-dependent vasodilatation is impaired by AF and improves after sinus rhythm is restored.

Analysis of IL-12 receptor beta 2 chain expression of circulating T lymphocytes in patients with atopic asthma.

Th2 cell predominance relative to Th1 cells contributes to pathological immune responses in patients with atopic asthma. IL-12 is a key cytokine in the induction of Th1 cells, and downregulation of IL-12 production is reported in these patients. However, IL-12 receptor expression of their T lymphocytes has not been clarified. In this study, expression of IL-12 receptor beta 2 on T cells and secretion of cytokines which affect IL-12 receptor beta 2 expression by their PBMC were examined. We found that IL-12 receptor beta 2 expression of the T cells is reduced. This is partly due to the diminished production of IL-12 and enhanced secretion of IL-4 by their PBMC. IL-18 production is not significantly modulated in these patients. Furthermore, intrinsic defects of the CD4(+) T cells, which reduce their IL-12 receptor beta 2 expression in response to IL-12 and/or IL-18 stimulation, are evident and are importantly involved in the Th1/Th2 imbalance of patients with atopic asthma.

Pyloricidins, novel anti-helicobacterpylori antibiotics produced by Bacillus sp. I. Taxonomy, fermentation and biological activity.

Novel anti-Helicobacter pylori antibiotics, pyloricidins A, A1, A2, B, C and D, were discovered in the culture broth of two bacilli strains. Pyloricidins selectively inhibited the growth of H. pylori. Pyloricidin B was efficacious in the treatment of gastric infection caused by H. pylori in Mongolian gerbils and may be promising for cure of H. pylori infection as a single agent.