Linkage mapping of fifty-eight new rat microsatellite markers

Fifty-eight new anonymous simple sequence repeats (SSR) were generated and mapped to various rat chromosomes. Among them two genes (rat homologs for human cadherin-14 and mouse fibroblast growth factor-related protein) were mapped on Chromosomes (Chrs) 2 and 11 respectively. The majority of markers were generated from a small insert genomic library specific to Chr 11, 13, 14, and 15. Twenty new markers were mapped to Chr 13, which is known to contain a blood pressure quantitative trait locus (QTL). Several approaches to obtain microsatellite markers are described. The protocols and newly generated markers should be useful for ongoing rat genome project.     

Fibroblast growth factor-2 has opposite effects on human breast cancer MCF-7 cell growth depending on the activation level of the mitogen-activated protein kinase pathway

In human breast cancer MCF-7 and MCF-7ras cells, we demonstrated that whereas insulin had a mitogenic effect on both cell lines, fibroblast growth factor-2 (FGF-2) had opposite effects, stimulating MCF-7 and inhibiting MCF-7ras cell proliferation. The inhibitory signal induced by FGF-2 was related to sustained mitogen-activated protein kinase (MAPK) activation in MCF-7ras cells, while transient MAPK activation was associated with MCF-7 cell proliferation. FGF-2 was further used in combination with insulin or cAMP. In MCF-7 cells, insulin and cAMP reversed the mitogenic effect of FGF-2. In MCF-7ras cells, insulin did not modify the inhibitory effect of FGF-2, but cAMP markedly enhanced it. These effects were also associated with an increased level and duration of MAPK activation. PD98056 abolished the effect of FGF-2 on DNA synthesis in both cell lines, demonstrating that the dual effect of FGF-2 on cell proliferation is dependent on the activity of the extracellular-signal-regulated kinase 1 and 2 (ERK1/2) signalling pathway.     

Syndrome of myxomas, spotty skin pigmentation, and endocrine overactivity (Carney complex)

Primary testicular neoplasms are common in dogs, but metastases to the testes are rare. Three dogs had enlargement of the testes and associated structures. Upon histological examination, the enlargements were due to metastatic adenocarcinomas. Further examination identified the gastrointestinal tract as the primary site of the metastatic neoplasms in all three cases. The testicular metastases reflected widespread metastatic disease. When metastatic adenocarcinoma is found in the testes and associated structures in dogs, the gastrointestinal tract should be examined closely for a primary tumor site.     

Cavo-pulmonary anastomosis associated with intracardiac repair of Ebstein anomaly. Value in high-risk patients]

The cyclin-dependent kinase inhibitor p27Kip1 controls cell proliferation in response to normal mitogenic stimuli. We show here that p27Kip1 also safeguards against excessive cell proliferation in specific pathophysiologic settings. We used experimental glomerulonephritis as a paradigm for immune mediated inflammation and ureteral obstruction as a model for non-immune mediated inflammation. Renal function was substantially decreased in nephritic p27-/- mice compared with control mice, and this was associated with increased glomerular cell proliferation, apoptosis and matrix protein accumulation. Tubular epithelial cell proliferation and apoptosis was also increased in p27-/- mice following ureteral obstruction. p27Kip1 may have a general role in protecting cells and tissues from inflammatory injury.     

Antinociceptive analogs of orphanin FQ/nociceptin(1-11)

The presence of pairs of basic amino acids within the sequence of orphanin FQ/nociceptin (OFQ/N) peptide, the endogenous ligand for the ORL1/KOR-3 receptor, has raised the possibility that processing might generate pharmacologically important truncated peptides, including OFQ/N(1-11). OFQ/N(1-11) is pharmacologically active in vivo with a potency comparable to OFQ/N. Several tyrosine-containing analogs of OFQ/N(1-11) have been synthesized and examined for antinociceptive activity. Like OFQ/N(1-11), [Tyr1]OFQ/N(1-11), [Tyr10]OFQ/N(1-11) and [IodoTyr10]OFQ/N(1-11) given supraspinally in mice were antinociceptive in the tailflick assay in mice. The tyrosine analogs showed similar potencies as OFQ/N(1-11) but longer durations of action. This response was readily reversed by the opioid antagonist naloxone despite poor affinities for these analogs at opioid receptors. Another compound, [Tyr11]OFQ/N(1-11) was highly epileptogenic, inducing naloxone-sensitive seizures in greater than 50% of the mice tested at doses comparable to those examined with the other analogs. These results indicate that it is possible to make analgesic OFQ/N(1-11) analogs. The activity of [IodoTyr10]OFQ/N(1-11) suggests that it may prove useful as a radioligand in exploring potential OFQ/N(1-11) binding sites.     

Effects of a new platelet glycoprotein IIb/IIIa antagonist, SR121566, on platelet activation, platelet-leukocyte interaction and thrombin generation

The effects of SR121566, a new inhibitor of the glycoprotein (GP) IIb/IIIa complex on platelet activation and platelet-leukocyte interactions, as well as on thrombin generation were investigated. SR121566 dose-dependently inhibited adenosine diphosphate (ADP)-induced platelet fibrinogen binding determined either by flow cytometry analysis (IC50=50 nmol/l) or by measuring the binding of 125I-fibrinogen to activated human gel-filtered platelets (IC50=20 nmol/l). Consistent with its inhibitory effects on platelet fibrinogen binding, SR121566 demonstrated a dose-dependent inhibition of collagen-, ADP- or thrombin-induced platelet aggregation with IC50 values ranging between 20 and 60 nmol/l. SR121566, even tested at high concentrations, did not significantly affect ADP-induced platelet-leukocyte aggregate formation. The GPIIb/IIIa antagonist strongly inhibited thrombin generation in both native clotting blood and recalcified whole blood, suggesting that SR121566, by interfering with the platelet-activation events involved in facilitating thrombin generation, may also function as an anticoagulant, an effect which may contribute to its antithrombotic properties in humans.     

Starvation selection restores elastase and rhamnolipid production in a Pseudomonas aeruginosa quorum-sensing mutant

The las quorum-sensing system of Pseudomonas aeruginosa controls the expression of elastase and rhamnolipid. We report that starvation can select a mutant producing these virulence factors in spite of a lasR deletion. Expression of the autoinducer synthase gene rhlI was increased in this suppressor mutant, suggesting compensation by the rhl system. These data show that P. aeruginosa can restore elastase and rhamnolipid production in the absence of a functional las quorum-sensing system.     

Dual control by divalent cations and mitogenic cytokines of alpha 4 beta 1 and alpha 5 beta 1 integrin avidity expressed by human hemopoietic cells

Beta-1 integrins have essential functions in hemopoietic and immune systems by controlling phenomenons such as cell homing and cell activation. The function alpha 4 beta 1 and alpha 5 beta 1 integrins is regulated by divalent cations and, as demonstrated more recently, by mitogenic cytokines which activate them by "inside-out" mechanisms. Using the adhesive interaction of a cytokine-dependent human hemopoietic cell line to immobilized fibronectin, we have analyzed the requirements in divalent cations Mn2+, Mg2+ and Ca2+ for alpha 4 beta 1 and alpha 5 beta 1 activation by "inside-out" mechanisms triggered by cytokines such as granulocyte-macrophage colony stimulating factor or KIT ligand, or by external conformational constraints with the function-activating anti-beta 1 integrin monoclonal antibody 8A2. The intrinsic difference between these two modes of beta 1 integrin activation was revealed by their different requirements in divalent cations. We found that in the absence of any divalent cations, alpha 4 beta 1 and alpha 5 beta 1 were non-functional even after further stimulation by cytokines or 8A2. However, whilst either Ca2+, Mg2+ or Mn2+ were able to restore adhesive functions of alpha 4 beta 1 and alpha 5 beta 1 when activated by 8A2, only Mg2+ and Mn2+ were able to support activation of alpha 4 beta 1 and alpha 5 beta 1 by cytokines. Furthermore, high concentrations of Ca2+ exceeding 20 mM dramatically inhibited cell adhesion to fibronectin induced by Mn2+ and cytokines but not by 8A2. On the contrary, in the presence of both Ca2+ and Mg2+, Mn2+ had an additive effect on the activation of alpha 4 beta 1 and alpha 5 beta 1 by mitogenic cytokines. The presence of the absence of these divalent cations did not inhibit early tyrosine phosphorylation induced by the binding of KIT ligand to its tyrosine-kinase receptor KIT. Therefore, we propose that in hemopoietic cells, Ca2+, Mg2+ and Mn2+ may modulate in vivo alpha 4 beta 1 and alpha 5 beta 1 regulation by mitogenic cytokines, a phenomenon involved in the regulation of hemopoietic progenitor cell homing within the bone marrow.