Autoantibodies to T cell costimulatory molecules in systemic autoimmune diseases

To determine whether antilymphocyte Abs to T cell costimulatory molecules are generated in patients with autoimmune diseases and, if they exist, to clarify the mechanism of their production and pathological roles, we investigated the presence of autoantibodies to CTLA-4 (CD152), CD28, B7-1 (CD80), and B7-2 (CD86) in serum samples obtained from patients with various autoimmune diseases and from normal subjects using recombinant fusion proteins. In ELISAs, anti-CD28, anti-B7-1, and anti-B7-2 Abs were rarely seen, whereas anti-CTLA-4 Abs were detected in 8.2% of the patients with systemic lupus erythematosus, 18.8% of those with rheumatoid arthritis, 3.1% of those with systemic sclerosis, 31.8% of those with Beh?et's disease, 13.3% of those with Sj?gren's syndrome, and 0% of healthy donors. This reactivity was confirmed by immunoblotting. More importantly, the purified anti-CTLA-4 Abs reacted with CTLA-4 expressed on P815 cells by flow cytometry. In addition, we found at least three epitopes on the CTLA-4 molecule. Furthermore, among the patients with Beh?et's disease, uveitis was seen significantly less frequently in the anti-CTLA-4 Ab-positive patients. Taken collectively, these data indicate that anti-CTLA-4 autoantibodies are generated in systemic autoimmune diseases by an Ag-driven mechanism and may modulate the immune response in vivo by binding to CTLA-4 on T cells.     

Identification and characterization of a new pair of immunoglobulin-like receptors LMIR1 and 2 derived from murine bone marrow-derived mast cells

We have identified and characterized two mouse cDNAs in a mouse antigen-stimulated bone marrow-derived mast cell cDNA library, both of which encode type I transmembrane proteins. The genes were closely mapped in the distal region of mouse chromosome 11 and expressed not only in mast cells but also widely in leukocytes. The extracellular domains of their encoded proteins contain a single variable immunoglobulin (Ig) motif sharing about 90% identity with amino acids, showing that they comprise a pair of molecules and belong to the Ig superfamily. We named these molecules leukocyte mono-Ig-like receptor1 and 2 (LMIR1 and 2). The intracellular domain of LMIR1 contains several immunoreceptor tyrosine-based inhibition motifs (ITIMs). When cross-linked, the intracellular domain was tyrosine phosphorylated and capable of recruiting tyrosine phosphatases, SHP-1 and SHP-2 and inositol polyphosphate 5-phosphatase, SHIP. LMIR2, on the other hand, contains a short cytoplasmic tail and a characteristic transmembrane domain carrying two positively charged amino acids associated with three kinds of immunoreceptor tyrosine-based activation motif (ITAM)-bearing molecules, DAP10, DAP12, and FcRgamma. These findings suggest that a new pair of ITIM/ITAM-bearing receptors, LMIR1 and 2, regulate mast cell-mediated inflammatory responses through yet to be defined ligand(s).     

Simultaneous effect of calcium,magnesium, copper and cobalt ions on sapogenin steroids content in callus cultures of Agave amaniensis

The simultaneous effect of calcium, cobalt, copper and magnesium ions and their interactions on growth and sapogenin steroids accumulation in callus cultures of Agave amaniensis was studied by using a central composite second-order rotatable design. The absence of calcium ions in media increased the sapogenin steroid content, while relatively high concentration of magnesium, cobalt and copper ions simultaneously inhibited the sapogenin steroid formation. This revised version was published online in June 2006 with corrections to the Cover Date.     

Chromosomes of Phagocata kawakatsui and Bdellocephala annandalei from Lake Biwa-ko in Honshú, central Japan

Two lake-dwelling species of paludicolen triclads from Lake Biwa-ko (Honshû, Japan) were studied taxonomically and karyologically. (1) Phagocata kawakatsui Okugawa, 1956, is an epigean species usually inhabiting shallow springs and spring-fed streams in Central Japan. In Lake Biwa-ko, animals were obtained from several bottom stations of the littoral area in the southern part of the northern basin (3–70 m in depth). Chromosome numbers and karyotype: 2x=24 (2m+2sm+2sm+2m+2sm+2m+2sm+2m+2m+2sm+2m+2m). The first pair of metacentric chromosomes is very large in size. (2) Bdellocephala annandalei Ijima et Kaburaki, 1916, an endemic species, is distributed widely in the deep areas of the northern basin (30 to over 100 m in depth). Chromosome numbers and karyotype: 2x=28 (2m+2sm+2sm+2sm+2sm+2m+2m+2m+2m+2m+2m+2m+2m+2m) with the first pair of metacentric chromosomes very long.     

Determination of guanosine tetraphosphate (ppGpp) and adenosine pentaphosphate (pppApp) in various microorganisms by radioimmunoassay

The intracellular levels of ppGpp and pppApp in various microorganisms were determined by radioimmunoassay, with was of more accuracy and convenience than previous ³²P-labeling method. ppGpp was detected in bacteria, actinomycetes, yeasts and fungi. pppApp was found in non-spore forming bacteria such as the genera Pseudomonas and Escherichia, fungi and actinomycetes, but not in yeasts. In conclusion, ppGpp and pppApp are present in various prokaryotes and lower eukaryotes.     

Occurrence of pppApp-synthesizing activity in actinomycetes and isolation of purine nucleotide pyrophosphotransferase

The occurrence of adenosine 5-triphosphate-3-diphosphate-synthesizing activity was detected in five strains of actinomycetes; Streptomyces morookaensis, Streptomyces aspergilloides, Streptomyces hachijoensis, Actinomyces violascens and Streptoverticillium septatum, out of 825 strains of actinomycetes, bacteria, fungi and imperfecti. Purine nucleotide pyrophosphotransferase were extracellularly excreted associating with the cell growth, and were purified partially or to apparent homogeniety from the culture filtrate. The enzymes are a monomeric protein with a molecular weight of 18000–26000 and synthesize adenosine, guanosine and inosine 5-phosphate (mono, di or tri)-3-diphosphate such as pApp, ppApp, pppApp, pGpp, ppGpp, pppGpp and pppIpp by transferring a pyrophosphoryl group from the 5-position of ATP, dATP and pppApp to the 3-position of purine nucleotides in the presence of a divalent cation and in alkaline state.Abbreviations pppApp adenosine 5-triphosphate 3-diphosphate - ppApp adenosine 5-diphosphate 3-diphosphate - pApp adenosine 5-monophosphate 3-diphosphate - pppGpp guanosine 5-triphosphate 3-diphosphate     

Is the nexus necessary for cell-to-cell coupling of smooth muscle?

Summary Electronmicroscopic study of electrically coupled smooth muscles was undertaken to determine the distribution of nexuses in various types of smooth muscle. The study revealed that while nexal structures were commonplace in some types of smooth muscle, they were very rare or absent in others, even though in some cases these cells were only a few nanometers distant from one another. The persistence in thin section of these structures in the main circular muscle of dog intestine after poor fixation, fixation under strain, cell shrinkage, and metabolic damage of various sorts seems to rule out the thesis that they are labile. The absence of nexuses in longitudinal muscle of dog intestine examined both by thin section and by freeze fracture suggests that in this tissue they are absent or very rarein vivo and cannot account for electrical coupling.Nexuses were discernible in thin sections of main circular muscle after a variety of experimental conditions of fixation. Metabolic inhibition orin vitro permanganate fixation partially destroyed nexal contacts. These procedures induced tissue, membrane apposition and an accompanying increase in the number of structures which resemble nexuses at low magnification (nexus-like structures). Nexus-like structures occurred in all smooth muscle fixed byin vitro permanganate associated with apposition of membranes and poor preservation of basement membrane. A technique ofin vitro permanganate fixation was developed which prevented tissue swelling; consequently nexus-like structures were absent in tissues so treated. The suggestion is made that some structures described in the literature as nexuses, following permanganate fixation, may represent nexus-like structures.The balance of evidence suggests that nexuses need not be present for electrical coupling of some smooth muscle cells, in which other types of cell-to-cell contacts must be invoked.     

Ontogenetic Development of the Striatal [3H]Spiperone Binding: Regulation by Sodium and Guanine Nucleotide in Rats

Ontogenetic development of specific [3H]spiperone binding to crude synaptic membranes and its regulation by Na+ and GTP was investigated in the rat striatum. (d)-Butaclamol more effectively inhibited [3H]spiperone binding than (l)-butaclamol. The ratio of inhibitory activity of (d)- and (l)-butaclamol for [3H]spiperone binding was not different between 1-, 7-, and 70-day-old animals but eight- to ninefold lower at 18 days of gestation than during the postnatal period. A Scatchard plot of specific binding indicated the presence of two types of binding: low-affinity (KD = 1.51 nM) and high-affinity (KD = 0.09 nM) binding on day 70. Only one component (KD = 0.075 nM) was observed on days 1 and 7 and both types of binding were found on day 15. Bmax gradually increased with age and reached a peak on day 30, followed by a decline on days 70 and 360. Na+, 100 mM, significantly increased specific binding on days 1, 7, 15, and 70. GTP, 50 microM, completely reversed the Na+-induced decrease in IC50 of apomorphine on both days 15 and 70, but not on day 7. It is suggested that receptors could recognize ligand stereospecificity on day 1. The density in dopamine receptors in the striatum reaches a peak on day 30, followed by a decrease on days 70 and 360. In addition, regulation by Na+ and GTP in agonist binding to dopamine receptors seems to become functional between 1 and 2 weeks after birth.     

Atrial natriuretic peptide and vasopressin in human plasma

Using a specific radioimmunoassay for atrial natriuretic peptide (ANP), plasma immunoreactive ANP was measured in 17 normal subjects and 83 patients with various diseases. Plasma ANP concentration in normal subjects was 14.1 +/- 1.7 pg/ml (mean +/- S.E.). Relatively high plasma ANP concentrations were detected in patients with diabetes mellitus, hyperthyroidism, atrial fibrillation and liver cirrhosis. Plasma ANP concentrations in the patients correlated positively with mean arterial blood pressure and plasma AVP concentrations. Plasma ANP concentrations in the patients also had positive correlations with left atrial dimension and left ventricular diastolic dimension determined by echocardiography. Another positive correlation was observed in the patients between plasma AVP concentrations and mean arterial blood pressure. These results suggest that ANP is a volume regulatory hormone but also that ANP may be involved in the blood pressure regulating system.