Wagner vitreoretinal degeneration with genetic linkage refinement on chromosome 5q13-q14

· Background: It has been previously described that Wagner disease is linked to chromosome 5q13-q14. This study was carried out to describe the ophthalmological aspects and report the results of genetic linkage analysis in a large pedigree affected by Wagner disease. · Methods: Fourty members of one same family agreed to be examined. · Results: Twenty patients presented vitreoretinal degeneration in both eyes without any extra-ocular abnormalities. In young patients, visual acuity was usually normal after correction of frequent mild myopia. Presenile cataracts progressed by the third decade and required removal for visual rehabilitation. The primary disorder involved an abnormal vitreous. A few avascular vitreous bands were usually the only optical feature in the mostly empty vitreous cavity. A circumferential vitreous condensation formed in contact with the retina on many spots. Less common retinal findings included retinal detachment, abnormal retinal pigmentation, progressive atrophy of the RPE simulating choroideremia and lattice degeneration. Genetic analysis revealed a highly significant linkage (lod score >5.0) between the disease and 10 markers of the chromosome 5q13-q14 region. Two recombination events allowed us to refine the linked interval to 20 cM between the D5S650 and D5S618 markers. · Conclusion: Ophthalmological aspects of Wagner’s disease appear to progress with age. Regular ophthalmological examination is important for detecting retinal abnormalities. The gene involved in Wagner’s disease lies in a 20 cM interval on chromosome 5q13-q14. Received: 30 June 1998 Revised version received: 5 October 1998 Accepted: 6 October 1998     

Promotion of cell growth by stimulation of cloned human 5-HT1D receptor sites in transfected C6-glial cells is highly sensitive to intrinsic activity at 5-HT1D receptors

5-Hydroxytryptamine (serotonin, 5-HT), essentially known as a neurotransmitter and vasoactive agent, also functions as a mitogen in various cell types through several different second messenger systems. Stimulation of cloned human 5-HT_1D receptor sites by sumatriptan in stably transfected rat C6-glial/5-HT_1D cells promotes cell growth (Pauwels et al. (1996) Naunyn-Schmiedeberg's Arch Pharmacol 353:144–156). In the present study, the pharmacology of this growth response was investigated using a broad series of 5-HT receptor ligands. The data were compared with the responses obtained by measuring inhibition of forskolin-stimulated cAMP formation. 5-HT (EC₅₀: 25 nM) promoted cell growth of C6-glial/5-HT_1D cells, and this in contrast to the absence of any measurable effect in pcDNA3-plasmid transfected and non-transfected C6-glial cells. The 5-HT effect could be mimicked by the following compounds (EC₅₀ in nM): zolmitriptan (0.41), 2-methyl-4-(5-methyl[1,2,4] oxadiazol-3-yl)biphenyl-4-carboxylic acid [4-methoxy-3-(4-methylpiperazin-1-yl)phenyl]amide (GR 127,935; 0.86), naratriptan (0.92), metergoline (1.9), sumatriptan (2.9), (N,N-dimethyl-2-[5-(1,2,4-triazol-1-ylmethyl)-1H-indol-3-y)]ethylamine (MK-462; 3.0), and R(+)-8-hydroxy-2-(di-n-propylamino)tetralin (R(+)-8-OH-DPAT; 30.7). These EC₅₀-values correspond to the compounds binding affinities at the human 5-HT_1D receptor site and, with the exception of GR 127,935 and metergoline, also to the EC₅₀-values found by measuring over 5 min inhibition of forskolin (100 M)-stimulated cAMP formation. Prolonged exposure of GR 127,935 (3 h) and metergoline (30 min) to cells yielded EC₅₀ values in the cAMP assay more close to those measured in the mitogenic response. The growth response to sumatriptan, 5-HT, GR 127,935 and metergoline was blocked by the apparently silent antagonists methiothepin, ritanserin and ketanserin with potencies similar to blockade of inhibition of stimulated CAMP formation. The 8-OH-DPAT effect also is likely mediated by 5-HT_1D receptors; stereoselectivity was found with its enantiomers at this receptor site and the effect was blocked by ketanserin (1 M) but not by spiperone (1 M). Micromolar concentrations of the 5-HT_1B receptor agonist 3-(1,2,5,6-tetrahydro)-4-pyridil-5-pyrrolo[3, 2-b]pyril-5-one (CP 93,129) and of the 5-HT₂ receptor agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) induced cell growth with a potency that accorded with the affinity of these compounds for the human 5-HT_1D receptor site. These effects were sensitive to ketanserin (1 M) antagonism, but not to blockade by -adrenergic blockers and the 5-HT₂ receptor antagonist 2-anilino-N-[2-(3-chlorophenoxy)-propyl] acetamidine hydroiodide (BW 501-C-67). The findings suggest that 5-HT_1A, 5-HT_1B and 5-HT₂ receptors are not implicated in 5-HT-stimulated C6-glial/5-HT_1D cell growth. In conclusion, human 5-HT_1D receptors are involved in the growth of C6-glial/5-HT_1D cells. This cellular response is highly sensitive to the intrinsic activity of compounds at 5-HT_1D receptors.     

SSR181507, a dopamine D(2) receptor antagonist and 5-HT(1A) receptor agonist. I: Neurochemical and electrophysiological profile.

SSR181507 ((3-exo)-8-benzoyl-N-[[(2S)7-chloro-2,3-dihydro-1,4-benzodioxin-1-yl]methyl]-8-azabicyclo[3.2.1]octane-3-methanamine monohydrochloride) is a novel tropanemethanamine benzodioxane derivative that possesses high and selective affinities for D2-like and 5-HT(1A) receptors (K(I)=0.8, 0.2, and 0.2 nM for human D(2), D(3), and 5-HT(1A), respectively). In vivo, SSR181507 inhibited [(3)H]raclopride binding to D(2) receptors in the rat (ID(50)=0.9 and 1 mg/kg, i.p. in limbic system and striatum, respectively). It displayed D(2) antagonist and 5-HT(1A) agonist properties in the same concentration range in vitro (IC(50)=5.3 nM and EC(50)=2.3 nM, respectively, in the GTPgammaS model) and in the same dose range in vivo (ED(50)=1.6 and 0.7 mg/kg, i.p. on striatal DA and 5-HT synthesis, respectively, and 0.03-0.3 mg/kg, i.v. on dorsal raphe nucleus firing rate). It selectively enhanced Fos immunoreactivity in mesocorticolimbic areas as compared to the striatum. This regional selectivity was confirmed in electrophysiological studies where SSR181507, given acutely (0.1-3 mg/kg, i.p.) or chronically (3 mg/kg, i.p., o.d., 22 days), increased or decreased, respectively, the number of spontaneous active DA cells in the ventral tegmental area, but not in the substantia nigra. Moreover, SSR181507 increased both basal and phasic DA efflux (as assessed by microdialysis and electrochemistry) in the medial prefrontal cortex and nucleus accumbens, but not in the striatum. This study shows that the combination of D(2) receptor antagonism and 5-HT(1A) agonism, in the same dose range, confers on SSR181507 a unique neurochemical and electrophysiological profile and suggests the potential of this compound for the treatment of the main dimensions of schizophrenia.     

Resolution of isoforms of mutalysin II, the metalloproteinase from bushmaster snake venom

Mutalysin II, a zinc endopeptidase possessing direct-acting fibrinolytic activity has been previously purified from bushmaster (Lachesis muta muta) snake venom. We now report a method to isolate two isoforms of natural mutalysin II (mut IIa and mut IIb) using chromatographies on Sephacryl S-200, CM Sepharose CL 6B and Sephadex G-50. The two proteins are monomeric non-glycosylated proteinases with similar molecular masses of 23 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Tryptic peptide mapping of the two native enzymes suggested a large degree of structural similarity. Both isoforms showed high similarity in all enzymatic properties using fibrinogen, fibrin and dimethylcasein as substrates. Thus, the specific fibrinolytic activity was estimated as 12±1.04 and 11.5±1.02 U/μg for mut IIa and mut IIb, respectively. The antigenic cross-reactivity of both isoforms was examined using rabbit hyperimmune serum or immunoglobulin G anti-mut IIa assays on immunodiffusion microscope slides, indirect enzyme-linked immunoabsorbent assay and western blots. From these experiments it was concluded that the two metalloproteinases mut IIa and mut IIb share identical antigenic structures. Since the stability of mutalysin II is dependent upon the presence of zinc, we examined the EDTA sensitivity of the isoforms of mutalysin II. Thus, the IC₅₀ values (concentration of EDTA to produce 50% inhibition of dimethylcasein hydrolysis) for mut IIa is 180 μM and 165 μM for mut IIb.     

Investigation of three new mouse mammary tumor cell lines as models for transforming growth factor (TGF)-β and Neu pathway signaling studies: identification of a novel model for TGF-β-induced epithelial-to-mesenchymal transition

Introduction  

This report describes the isolation and characterization of three new murine mammary epithelial cell lines derived from mammary tumors from MMTV (mouse mammary tumor virus)/activated Neu + TβRII-AS (transforming growth factor [TGF]-β type II receptor antisense RNA) bigenic mice (BRI-JM01 and BRI-JM05 cell lines) and MMTV/activated Neu transgenic mice (BRI-JM04 cell line).     

Outcome of treatment in adults with acute lymphoblastic leukemia: analysis of the LALA-94 trial.

PURPOSE: We analyzed the benefits of a risk-adapted postremission strategy in adult lymphoblastic leukemia (ALL), and re-evaluated stem-cell transplantation (SCT) for high-risk ALL. PATIENTS AND METHODS: A total of 922 adult patients entered onto the trial according to risk groups: standard-risk ALL (group 1), high-risk ALL (group 2), Philadelphia chromosome-positive ALL (group 3), and CNS-positive ALL (group 4). All received a standard four-drug/4-week induction course. Patients from group 1 who achieved a complete remission (CR) after one course of induction therapy were randomly assigned between intensive and less intensive postremission chemotherapy, whereas those who achieved CR after salvage therapy were then included in group 2. Patients in groups 2, 3, and 4 with an HLA-identical sibling were assigned to allogeneic SCT. In groups 3 and 4, autologous SCT was offered to all other patients, whereas in group 2 they were randomly assigned between chemotherapy and autologous SCT. RESULTS: Overall, 771 patients achieved CR (84%). Median disease-free survival (DFS) was 17.5 months, with 3-year DFS at 37%. In group 1, the 3-year DFS rate was 41%, with no difference between arms of postremission randomization. In groups 2 and 4, the 3-year DFS rates were 38% and 44%, respectively. In group 2, autologous SCT and chemotherapy resulted in comparable median DFS. Patients with an HLA-matched sibling (groups 2 and 4) had improved DFS. Three-year DFS was 24% in group 3. CONCLUSION: Allogeneic SCT improved DFS in high-risk ALL in the first CR. Autologous SCT did not confer a significant benefit over chemotherapy for high-risk ALL.     

Albumin-protamine-oligonucleotide nanoparticles as a new antisense delivery system. Part 1: physicochemical characterization.

In this paper, a ternary system of albumin-protamine-oligonucleotide nanoparticles (AlPrO-NP) recently developed by Vogel et al. [V. Vogel, D. Lochmann, J. Weyermann, G. Mayer, C. Tziatzios, J.A. van den Broek, W. Haase, D. Wouters, U.S. Schubert, J. Kreuter, A. Zimmer, D. Schubert, Oligonucleotide-protamine-albumin nanoparticles: preparation, physical properties and intracellular processing, J. Controlled Rel. (in press)] which could serve as a potential drug delivery system for antisense oligonucleotides. Former studies of binary protamine-oligonucleotide nanoparticles showed two main disadvantages: (i) aggregation of the particles within a few minutes in the presence of salt; (ii) low intracellular dissociation between protamine and oligonucleotide, especially phosphorothioates. To overcome these problems, human serum albumin (HSA) as a non-toxic, biodegradable macromolecule was introduced as protective colloid. The assembly process of AlPrO-NP was investigated by small angle X-ray scattering (SAXS), fluorescence correlation spectroscopy (FCS), photon correlation spectroscopy (PCS) measurements and scanning electron microscopy (SEM). 'Initial complexes' of HSA and protamine sulphate with a mean hydrodynamic diameter (dh) of about 10-14 nm were found. After adding oligonucleotides (unmodified, phosphorothioate DNA and small interfering RNA), nanoparticles (NPs) were assembled in water and in isotonic media with a dh in a range of 230-320 nm for most preparations. The chemical composition of the particles was investigated by high performance liquid chromatography and fluorescence spectrometry. The whole amount of oligonucleotides (30 microg) was entrapped into the particles at a 1:2 mass ratio (oligonucleotide/protamine). Approximately 7-10% (w/w) of the HSA was bound to the particles. The surface charge of the particles ranged from about +12 to -60 mV depending on the protamine concentration and the ionic conditions. The size and the molecular weight of the components, initial complexes and two model NP preparations were calculated from FCS data. These data verified the PCS, SEM and SAXS measurements.     

Assessing the need to update prevention guidelines: a comparison of two methods.

BACKGROUND: An important concern for developers of clinical practice guidelines is how best to determine when guidelines require updating to ensure they remain current and evidence based. Because of the high costs associated with updating guidelines, recent attention has focused on approaches that can reliably assess the extent of updating required. Recently, Shekelle and colleagues proposed a model of limited literature searches with modest expert involvement as a way to reduce the cost and time requirements for assessing whether a guideline needs updating. METHODS: The main objective of this study was to compare the Shekelle et al. assessment model (review approach) and a conventional process using typical systematic review methods (traditional approach) in terms of comprehensiveness and effort. We modeled the review approach on that by Shekelle and colleagues but refined it iteratively over three phases to achieve greater efficiency. Using both methods independently, we assessed the need to update six topics from the 1996 Guide to Clinical Preventive Services from the US Preventive Services Task Force. Main outcomes included completeness of study identification, importance of missed studies and the effort involved. RESULTS: Although the review approach identified fewer eligible studies than the traditional approach, none of the studies missed was rated as important by task force members acting as liaisons to the project with respect to whether the topic required an update. On average, the review approach produced substantially fewer citations to review than the traditional approach. The effort involved and potential time saving depended largely on the scope of the topic. CONCLUSIONS: The revised review approach provides an efficient and acceptable method for judging whether a guideline requires updating.     

Prevalence of nosocomial infections in Swiss children's hospitals.

OBJECTIVE: To acquire data on pediatric nosocomial infections (NIs), which are associated with substantial morbidity and mortality and for which data are scarce. DESIGN: Prevalence survey and evaluation of a new comorbidity index. SETTING: Seven Swiss pediatric hospitals. PATIENTS: Those hospitalized for at least 24 hours in a medical, surgical, intensive care, or intermediate care ward. RESULTS: Thirty-five NIs were observed among 520 patients (6.7%; range per hospital, 1.4% to 11.8%). Bacteremia was most frequent (2.5 per 100 patients), followed by urinary tract infection (1.3 per 100 patients) and surgical-site infection (1.1 per 100 patients; 3.2 per 100 patients undergoing surgery). The median duration until the onset of infection was 19 days. Independent risk factors for NI were age between 1 and 12 months, a comorbidity score of 2 or greater, and a urinary catheter. Among surgical patients, an American Society of Anesthesiologists (ASA) score of 2 or greater was associated with any type of NI (P = .03). Enterobacteriaceae were the most frequent cause of NI, followed by coagulase-negative staphylococci; viruses were rarely the cause. CONCLUSIONS: This national prevalence survey yielded valuable information about the rate and risk factors of pediatric NI. A new comorbidity score showed promising performance. ASA score may be a predictor of NI. The season in which a prevalence survey is conducted must be considered, as this determines whether seasonal viral infections are observed. Periodic prevalence surveys are a simple and cost-effective method for assessing NI and comparing rates among pediatric hospitals.