Effects of Xenopus laevis mitochondrial single-stranded DNA-binding protein on primer-template binding and 3'-->5' exonuclease activity of DNA polymerase gamma

Mitochondrial DNA (mtDNA) is replicated by DNA polymerase gamma by a strand displacement mechanism involving mitochondrial single-stranded DNA-binding protein (mtSSB). mtSSB stimulates the overall rate of DNA synthesis on singly-primed M13 DNA mainly by stimulating the processivity of DNA synthesis rather than by stimulating primer recognition. We used electrophoretic mobility shift methods to study the effects of mtSSB on primer-template recognition by DNA pol gamma. Preliminary experiments showed that single mtSSB tetramers bind tightly to oligo(dT) single strands containing 32 to 48 residues. An oligonucleotide primer-template was designed with an 18-mer primer annealed to the 3'-portion of a 71-mer template containing 40 dT residues at its 5'-end as a binding site for mtSSB. DNA pol gamma bound to this primer-template either in the absence or presence of mtSSB in complexes that remained intact and enzymatically active following native gel electrophoresis. Association of mtSSB with the 5'-dT40-tail in the 18:71-mer primer-template reduced the binding of DNA polymerase gamma and the efficiency of primer extension. Binding of mtSSB to single-stranded DNA was also observed to block the action of the 3'-->5' exonuclease of DNA polymerase gamma. The size of fragments protected from 3'-->5' exonuclease trimming increases with increasing ionic strength in a manner consistent with the known salt dependence of the binding site size of Escherichia coli SSB.     

Laboratory aid and ultrasonography in the diagnosis of appendicitis in children

The purpose of this study was to evaluate the relationship between the number of yolk sacs and amnionicity in monochorionic twin pregnancies scanned early in the first trimester. We retrospectively reviewed images of all monochorionic twins scanned between 6 and 9.5 weeks' gestation and with pathologic or sonographic confirmation of chorionicity-amnionicity. Each film was reviewed for the number of yolk sacs present, as well as for the gestational age at which the amniotic membrane was first visualized. Twenty monochorionic-diamniotic pregnancies and two monochorionic-monoamniotic pregnancies met the criteria for inclusion in the study. In diamniotic pregnancies scanned at less than 8 weeks' gestation, only the yolk sacs were identified; none of the dividing amniotic membranes were detected. Two yolk sacs were identified in all but one case. In this case, although one yolk sac was seen at 6 weeks, follow-up scanning at 8 weeks revealed two yolk sacs. In each of the monochorionic-monoamniotic twin pregnancies, one yolk sac was seen at 9 weeks and a single amnion encircled both embryos. We conclude that the sonographic identification of two yolk scas in monochorionic twins enables us to make the diagnosis of diamniotic twins early in the first trimester, before the amniotic membrane can be imaged. The presence of one yolk sac should prompt a follow-up ultrasonogram to assign amnionicity definitively.     

Retained common bile duct stones after endoscopic sphincterotomy: temporary and longterm treatment with biliary stenting

Basket extraction after endoscopic sphincterotomy failed to clear the bile ducts immediately in 85 (30%) of 283 consecutive patients with common bile duct stones. Temporary biliary drainage was established by the insertion of a single 7 Fr double pigtail stent before further planned endoscopic attempts at stone removal. In 84 patients (21 male: 63 female, mean age 77 years) this measure relieved biliary obstruction, mean serum bilirubin falling from 101 to 18 umol/l by the time of the second endoscopic retrograde cholangiopancreatography. Six patients died from non-biliary causes with temporary stents in situ. Common bile duct stone extraction was achieved endoscopically in 50 of the remaining 79 patients after a mean of 4.3 months (range 1-12), 34 (68%) requiring only one further procedure. Three patients were referred for biliary surgery. Single stents were also effective for longterm biliary drainage in the remaining 26 elderly patients with unextractable stones. The main biliary complication of stenting was 13 episodes of cholangitis but all except one responded to medical treatment and early stent exchange. If common bile duct stones remain after endoscopic sphincterotomy, a single 7 Fr double pigtail stent is effective and safe for temporary biliary drainage before further endoscopic attempts at duct clearance and for longterm biliary drainage especially in the old and frail.     

Immunoglobulin A levels in southern elephant seal (Mirounga leonina) milk during the suckling period

Immunoglobulin A (IgA) levels in milk samples from southern elephant seals at King George Island, Antarctica are reported. IgA levels were determined throughout the suckling period (approximately 23 days). The IgA concentration in southern elephant seal milk was lower than in other mammals and, unlike most mammalian milk, was not high during early lactation. There was not a definite pattern in IgA levels, which fluctuated within narrow limits throughout the suckling period (mean +/- SD, 30.81 +/- 6.38 mg IgA/100 g milk). If IgG was present, its level was too low to be detected by the method used. This is the first evidence in Southern elephant seal of the possibility of transmission of passive immunity after birth involving secretion of IgA in the milk.     

The relation between rhinitis and allergic asthma. The action of antihistaminics on bronchial hyperreactivity

The intermediate filament nestin is highly expressed in multipotential stem cells of the developing central nervous system (CNS). During neuro- and gliogenesis, nestin is replaced by cell type-specific intermediate filaments, e.g. neurofilaments and glial fibrillary acidic protein (GFAP). In this study, we demonstrate that nestin expression is re-induced in reactive astrocytes in the lesioned adult brain. Following ischaemic and mechanical lesioning, a strong and sustained expression of nestin was noted in GFAP-positive cells surrounding the lesion site. Lesion experiments in transgenic mice carrying the lacZ gene under control of regulatory sequences from the nestin gene suggested that the upregulation of nestin in reactive astrocytes is mediated via the same sequences that control nestin expression during CNS development. These observations and recent data on the co-expression of glial and neuronal marker antigens in reactive astrocytes point to a close relationship between proliferating astrocytes and neuroepithelial precursor cells.     

Effects of different doses of apomorphine on the glutamate decarboxylase activity of the substantia nigra and the medial basal hypothalamus

The activity of gamma-aminobutyric acid (GABA) synthesizing enzyme glutamic acid decarboxylase (GAD, EC 4.1.1.15) was assayed in the rat substantia nigra (SN) and medial basal hypothalamus (MBH) following systemic injection of different doses of the dopamine receptor agonist apomorphine. In SN, the highest dose of apomorphine (1000 micrograms/kg) causes an increase of the GAD activity whilst an opposite effect is observed with the lowest dose (35 micrograms/kg). Results obtained in SN are in accordance with previous neurochemical and behavioural data suggesting an opposite action of high (500 micrograms/kg) and low doses (100 micrograms/kg) of apomorphine in nigro-striatal system, probably due to the existence of two classes of dopamine receptors, i.e. classical postsynaptic dopamine receptors and presynaptic inhibitory dopamine autoreceptors. In MBH, the evidence for similar effects of low and high doses of apomorphine (the decrease of GAD activity) may suggest that, as already reported, at this level only one class of dopamine receptors is present.     

Determination of the structure of exochelin MN, the extracellular siderophore from Mycobacterium neoaurum

BACKGROUND: Siderophores are compounds produced by bacteria to acquire iron. Exochelin MN, the extracellular siderophore from Mycobacterium neoaurum, is of particular interest because it has been shown to transport iron into M. leprae, which is responsible for the disease leprosy. Exochelins from other species cannot mediate iron transport in M. leprae, suggesting a specific uptake mechanism involving exochelin MN. We set out to determine the structure of exochelin MN and identify the features of the molecule that may account for this specificity. RESULTS: The structure of exochelin MN was elucidated by a combination of techniques including nuclear magnetic resonance, mass spectrometry, derivatization and gas chromatography. Exochelin MN is a peptide, containing the unusual amino acid beta-hydroxyhistidine and an unusual N-methyl group. The peptide coordinates iron(III) octahedrally using its two cis-hydroxamate groups plus the hydroxyl and imidazole nitrogen of the beta-hydroxyhistidine. The three-dimensional structure of the hexadentate exochelin/gallium complex was deduced from NMR data. CONCLUSIONS: Exochelin MN has some structural features in common with other siderophores, but has a unique three-dimensional structure, which is presumably important for its specific activity in M. leprae. Exochelin MN may be a target for drug design in the fight against infection with this pathogen.     

Femorodistal vein grafts: the utility of graft surveillance criteria

PURPOSE: This retrospective review of femorodistal vein grafts was analyzed to determine the usefulness of various graft surveillance criteria. METHOD: The surveillance schedule involved evaluations at 1 month, every 3 months the first year, and then every 6 months. Salvage intervention or graft occlusion occurring within the next follow-up interval defined surveillance end points. One hundred two grafts (329 surveillance visits) had an ankle/brachial index (ABI). A duplex scanning-determined midgraft peak systolic flow velocity (PSFV) was available for 81 grafts (262 visits). Forty-eight grafts (137 visits) had both a PSFV and entire graft duplex scanning (EGDS) to determine stenosis greater than 50%, whereas 40 grafts (91 visits) had simultaneous ABI and EGDS. RESULTS: When a greater than 15% decrease in ABI denoted an abnormal surveillance study result, a positive predictive value (PPV) of 24.3% and negative predictive value of 94.5% were noted. Similarly, a PSFV cutoff of less than 35 cm/sec demonstrated values of 26.3% and 94.2%, respectively. When an EGDS of greater than 50% stenosis or a PSFV of less than 35 cm/sec were the cutoff criteria, the PPV was 36.7% and negative predictive value 99.1%, whereas characterizing abnormal results further with ABI (> 15% decreases) increased the PPV to 83.3%. CONCLUSION: The combination of an EGDS, midgraft PSFV, and ABI provides optimal follow-up for our patients with a femorodistal vein graft.     

AGN 191976: a novel thromboxane A2-mimetic with ocular hypotensive properties

The possible subdivision of thromboxane A2-sensitive (TP) receptors is currently a controversial subject. We report herein on a novel thromboxane A2 mimetic, AGN 191976, which has almost identical pharmacological activity to the well-characterized prostaglandin H2/thromboxane A2 (PGH2/TxA2) mimetic U-46619, but its effects on intraocular pressure are quite distinct from U-46619. Prostanoid receptor activity was determined in vitro using different smooth muscle assays and platelets. Intraocular pressure was measured tonometrically in ocular normotensive Beagle dogs and Cynomolgus monkeys. Conjunctival microvascular permeability was determined in guinea pigs. Despite closely resembling U-46619 as a potent and selective TP receptor agonist, AGN 191976 was a potent ocular hypotensive in dogs and monkeys whereas U-46619 did not lower IOP in either species. The ocular hypotensive effect of AGN 191976 in dogs was attenuated by pretreatment with the TP receptor antagonist SQ 29548. Thus, the ocular hypotensive effects of AGN 191976 are consistent with TP receptor stimulation. Both TxA2-mimetics caused plasma leakage in the guinea pig conjunctiva. The disparate activities of U-46619 and AGN 191976 in our studies suggest the existence of heterogeneous populations of TP-receptors in the eye.