Molecular heterogeneity of D-end products detected by anti-H-2.28 sera

Immunoprecipitation of NP-40 lysates of 125I-labeled lymph-node cells with different anti-H-2 sera and with anti-Qa-2 serum has shown that the BALB/cByA strain (H-2d, Qa-2-negative) expresses, besides H-2Ld, another molecule that is not detectable in the BALB/c-H-2dm2 strain. Electrophoresis in SDS polyacrylamide gels indicated that this molecule, provisionally designated Lq, has an apparent molecular weight of 41000 daltons, in contrast to approximately 49000 daltons for H-2Kd and H-2Ld, and 47000 daltons for H-2Dd molecules. The anti-Qa-2 serum precipitated from the Qa-2-positive strains BALB/cHeA but not from the Qa-2-negative strains BALB/cByA and BALB/c-H-2dm2 a protein that gave a very strong band corresponding to the molecular weight 41000 daltons in the gel electrophoresis. The biochemical characteristics of the Lq molecule are thus more similar to those of Qa-2 than of H-2 antigens.     

Failure of testicular development associated with a rearrangement of 9p24.1 proximal to the SNF2 gene

In 46,XY individuals, testes are determined by the activity of the SRY gene (sex-determining region Y), located on the short arm of the Ychromosome. The other genetic components of the cascade that leads to testis formation are unknown and may be located on the Xchromosome or on the autosomes. Evidence for the existence of several loci associated with failure of male sexual development is indicated by reports of 46,XY gonadal dysgenesis associated with structural abnormalities of the Xchromosome or of autosomes (chromosomes9, 10, 11 and 17). In this report, we describe the investigation of a child presenting with multiple congenital abnormalities, mental retardation and partial testicular failure. The patient had a homogeneous de novo 46,XY,inv dup(9)(pter→p24.1::p21.1 →p23.3::p24.1→qter) chromosome complement. No deletion was found by either cytogenetic or molecular analysis. The SRY gene and DSS region showed no abnormalities. Southern blotting dosage analysis with 9p probes and fluorescent in situ hybridisation data indicated that the distal breakpoint of the duplicated fragment was located at 9p24.1, proximal to the SNF2 gene. We therefore suggest that a gene involved in normal testicular development and/or maintenance is present at this position on chromosome 9. Received: 20 January 1997 / Accepted: 5 November 1997     

Protein binding and stability of norepinephrine in human blood plasma. Involvement of prealbumin, alpha 1-acid glycoprotein and albumin

The binding of norepinephrine (NE) to plasma proteins of fresh human blood obtained from healthy volunteers was studied by ultrafiltration at different NE concentrations and incubation times at 37 degrees C. At 1.7 nM L-[3H]-NE binding was approximately 25%. The binding was rapid and was not influenced by the incubation time. [3H]-NE could be dissociated from its binding sites by acid precipitation and, after HPLC, showed to be unchanged NE. No difference in NE binding was found between plasma collected in EGTA-GSH or heparin solution. There was no degradation of NE when incubated in plasma at 37 degrees C for 10 h, even without the addition of antioxidants. Therefore, in the present study, binding represented interaction of unchanged NE with plasma proteins. The whole plasma binding was saturable over the range of 0.66 nM to 0.59 mM of NE. Scatchard plot of specific binding revealed high-affinity sites with a Kd of 5.4 nM and a Bmax of 3.9 fmoles.mg-1 protein, and low-affinity sites with a Kd of 2.7 microM and a Bmax of 3.3 pmoles.mg-1 protein. Electrophoretic characterization of NE-binding proteins showed that about 60% of bound NE was associated to albumin, and 20% to prealbumin. NE binding to pure human plasma proteins was also studied using ultrafiltration. Scatchard analyses revealed a single class of very high-affinity binding sites for prealbumin (Kd 4.9 nM), a single class of binding sites for alpha 1-acid glycoprotein (Kd 54 microM) and two classes of binding sites for albumin with high (Kd 1.7 microM) and low (Kd 0.8 mM) affinities respectively. The main results obtained in this study - a) reversibility of NE binding, b) stability of free and bound NE in plasma, c) involvement of the prealbumin as a specific binding protein - point out to a specific transport for NE in human blood plasma.     

Accumulation of the labdane diterpene Marrubiin in glandular trichome cells along the ontogeny of Marrubium vulgare plants

The content of the diterpene Marrubiin was assessed by GC-FID in leaves of Marrubium vulgare plants along their ontogeny. Maximum accumulation occurred just before flowering time and in fully expanded leaves. After feeding the plants with radio labeled [³H]-geranyl geranyl diphosphate, up to 70% of the radioactivity was recovered in HPLC-Rt coincidental with authentic Marrubiin, which was also characterized by GC-EIMS, thus confirming that the biosynthesis of Marrubiin proceeds through the 1-deoxy-D-xylulose-5-phosphate pathway. The major accumulation of radioactivity occurred in glandular trichome cells, and the product remained stable throughout.     

The metalloproteinase ADAM‐12 regulates bronchial epithelial cell proliferation and apoptosis

Abstract. Objectives: The ADAMs (a disintegrin and metalloproteinase) enzymes compose a family of membrane‐bound proteins characterized by their multi‐domain structure and ADAM‐12 expression is elevated in human non‐small cell lung cancers. The aim of this study was to investigate the roles played by ADAM‐12 in critical steps of bronchial cell transformation during carcinogenesis. Materials and methods: To assess the role of ADAM‐12 in tumorigenicity, BEAS‐2B cells were transfected with a plasmid encoding human full‐length ADAM‐12 cDNA, and then the effects of ADAM‐12 overexpression on cell behaviour were explored. Treatment of clones with heparin‐binding epidermal growth factor (EGF)‐like growth factor (HB‐EGF) neutralizing antibodies as well as an EGFR inhibitor allowed the dissection of mechanisms regulating cell proliferation and apoptosis. Results: Overexpression of ADAM‐12 in BEAS‐2B cells promoted cell proliferation. ADAM‐12 overexpressing clones produced higher quantities of HB‐EGF in their culture medium which may rely on membrane‐bound HB‐EGF shedding by ADAM‐12. Targeting HB‐EGF activity with a neutralizing antibody abrogated enhanced cell proliferation in the ADAM‐12 overexpressing clones. In sharp contrast, targeting of amphiregulin, EGF or transforming growth factor‐α failed to influence cell proliferation; moreover, ADAM‐12 transfectants were resistant to etoposide‐induced apoptosis and the use of a neutralizing antibody against HB‐EGF activity restored rates of apoptosis to be similar to controls.Conclusions: ADAM‐12 contributes to enhancing HB‐EGF shedding from plasma membranes leading to increased cell proliferation and reduced apoptosis in this bronchial epithelial cell line.     

Genotype-specific interactions and the trade-off between host and parasite fitness

Background  

Evolution of parasite traits is inextricably linked to their hosts. For instance one common definition of parasite virulence is the reduction in host fitness due to infection. Thus, traits of infection must be viewed in both protagonists and may be under shared genetic and physiological control. We investigated these questions on the oomycete Hyaloperonospora arabidopsis (= parasitica), a natural pathogen of the Brassicaceae Arabidopsis thaliana.     

Regulation of lung endothelin content by the glucocorticosteroid budesonide.

Intratracheal instillation of Sephadex beads induced a long-lasting inflammation in the rat lung as seen by an increase in lung weights. Repeated instillation enhanced this reaction and increased lung endothelin-1 content 3.5 times. Budesonide given s.c. abolished these effects and even reduced basal endothelin-1 content by 72%. The tissue content of the sensory neuropeptide neurokinin A were unaffected by both treatments. Endothelin has been proposed to play a part in the pathogenesis of bronchial asthma. If it is so, the ability of budesonide to reduce endothelin-1 content could thus be added to the list of beneficial effects of glucocorticosteroids in these conditions.     

Artifacts following gold staining of Western-blotted membranes

Protein samples were separated by sodium dodecyl-sulfate-polyacrylamide gel electrophoresis, electrophoretically transferred to nitrocellulose membranes, and stained with colloidal gold. Three artifact bands appeared demonstrating apparent molecular weights of 58,000, 63,000, and 65,000. The appearance of these bands was due to oxidized dithiothreitol used to denature the proteins prior to electrophoresis. The appearance of the artifact bands could be avoided by the addition of excess iodoacetamide to the denatured protein sample and by limiting staining time to 1.5 h at 13 V/cm.     

Toll-like receptor 4 deficiency: Smaller infarcts,but nogain in function

Backgound  

It has been reported that Toll-like receptor 4 (TLR4) deficiency reduces infarct size after myocardial ischemia/reperfusion (MI/R). However, measurement of MI/R injury was limited and did not include cardiac function. In a chronic closed-chest model we assessed whether cardiac function is preserved in TLR4-deficient mice (C3H/HeJ) following MI/R, and whether myocardial and systemic cytokine expression differed compared to wild type (WT).     

An influenza A virus-specific and HLA-DRw8-restricted T cell clone cross-reacting with a transcomplementation product of the HLA-DR2 and DR4 haplotypes

The clone TA10 is a T3+ T4+ T8- proliferative and cytolytic human T cell clone. This clone has been shown to be specific for the hemagglutinin of influenza A Texas virus and restricted by an HLA class II molecule associated with the DRw8-Dw8.1 phenotype. Here we show that TA10 and all of its subclones can also react with eight HLA-DRw8 negative, Epstein-Barr virus (EBV)-transformed cell lines or phytohemagglutinin blasts in the absence of influenza antigens. All of these cell lines are HLA-DR2/DR4 with a classic DR2 long haplotype. The only nonreactive HLA-DR2/DR4 cell line observed bears a DR2 short haplotype. Only heterozygous HLA-DR2/DR4 but not parental DR2 or DR4 EBV-transformed cell lines can be recognized by TA10, indicating that the cross-reacting determinant is a transcomplementation product between HLA-DR2 and HLA-DR4 haplotypes. DR-specific, but not DQ- or DP-specific monoclonal antibodies, inhibit in the proliferation assay and in the chromium release test both the DRw8-Dw8.1-restricted and the anti-DR2/DR4 reactions. These results show that HLA-DR-restricted, anti-viral human T cell clone can evidence cross-reactivity for allospecific class II molecules of the major histocompatibility complex, and human CTL can recognize transcomplementation products of class II HLA genes. In addition, the results suggest that a beta-chain coded for by an HLA-DR gene and associated with an alpha-chain coded for by a still unidentified but possibly HLA-DQ gene constitute this functional transcomplementation product.