3'-phosphodiesterase activity of human apurinic/apyrimidinic endonuclease at DNA double-strand break ends

In order to assess the possible role of human apurinic/apyrimidinic endonuclease (Ape) in double-strand break repair, the substrate specificity of this enzyme was investigated using short DNA duplexes and partial duplexes, each having a single 3'-phosphoglycolate terminus. Phosphoglycolate removal by Ape was detected as a shift in mobility of 5'-end-labeled DNA strands on polyacrylamide sequencing gels, and was quantified by phosphorimaging. Recombinant Ape efficiently removed phosphoglycolates from the 3'-terminus of an internal 1 base gap in a 38mer duplex, but acted more slowly on 3'-phosphoglycolates at a 19 base-recessed 3'-terminus, at an internal nick with no missing bases, and at a double-strand break end with either blunt or 2 base-recessed 3'-termini. There was no detectable activity of Ape toward 3'-phosphoglycolates on 1 or 2 base protruding single-stranded 3'-overhangs. The results suggest that both a single-base internal gap, and duplex DNA on each side of the gap are important binding/recognition determinants for Ape. While Ape may play a role in repair of terminally blocked double-strand breaks, there must also be additional factors involved in removal of at least some damaged 3'-termini, particularly those on 3'-overhangs.     

Search for hepatitis C virus extrahepatic replication sites in patients with acquired immunodeficiency syndrome: specific detection of negative-strand viral RNA in various tissues

Systemically administered interleukin-1 (IL-1) has been shown to preferentially bind to IL-1 receptors (IL-1Rs) in inflammation. Using radiolabeled IL-1alpha and molecular methods to assess gene expression for these receptors, the in vivo behavior of these receptors was investigated in a number of experimental inflammatory conditions. The uptake of 125I-labeled IL-1alpha in inflammatory foci significantly correlated with the mRNA expression for the type I and type II IL-1Rs (P < .05). Type II IL-1R mRNA showed a greater increase in expression than type I IL-1R mRNA. In neutropenic mice, inflammatory lesions, which are devoid of granulocytes, significantly lower 125I-labeled IL-1alpha uptake (P < .001), and type II IL-1R mRNA expression (P < .005) was found. Thus, there is strong up-regulation of IL-1Rs at sites of focal inflammation. Of interest, this mainly involved the type II IL-1R on granulocytes, which is not involved in signal transduction.     

Strategic hospital alliances: impact on financial performance

Acute care hospitals have increasingly been forming local strategic hospital alliances (SHAs), which consume considerable resources in forming and may affect the competitiveness of provider markets. This research shows that SHAs and market factors, which have been perceived to be threats to hospitals, are related to hospitals' financial performance. Among the findings are that SHA members have higher net revenues but that they are not more effective at cost control. Nor do the higher net revenues result in higher cash flow. However, increasing SHA penetration in a market is related to lower net revenues per case. In addition, the penetration of private health maintenance organizations in markets is associated with lower revenues and expenses.     

Modulation of adenosine-induced cAMP accumulation via metabotropic glutamate receptors in chick optic tectum

Changes on cyclic adenosine monophosphate (cAMP) levels in response to adenosine and glutamate and the subtype of glutamate receptors involved in this interaction were studied in slices of optic tectum from 3-day-old chicks. cAMP accumulation mediated by adenosine (100 microM) was abolished by 8-phenyltheophylline (15 microM). Glutamate and the glutamatergic agonists kainate or trans-D, L-1-aminocyclopentane-1,3-dicarboxylic acid (trans-ACPD) did not evoke cAMP accumulation. Glutamate blocked the adenosine response in a dose-dependent manner. At 100 microM, glutamate did not inhibit the effect of adenosine. The 1 mM and 10 mM doses of glutamate inhibited adenosine-induced cAMP accumulation by 55% and 100%, respectively. When glutamatergic antagonists were used, this inhibitory effect was not affected by 200 microM 6,7-dihydroxy-2,3,dinitroquinoxaline (DNQX), an ionotropic antagonist, and was partially antagonized by 1 mM (RS)-alpha-methyl-4-carboxyphenylglycine [(RS)M-CPG], a metabotropic antagonist, while 1 mM L-2-amino-3-phosphonopropionate (L-AP3) alone, another metabotropic antagonist, presented the same inhibitory effect of glutamate. Kainate (10 mM) and trans-ACPD (100 microM and 1 mM) partially blocked the adenosine response. This study indicates the involvement of metabotropic glutamate receptors in adenylate cyclase inhibition induced by glutamate and its agonists trans-ACPD and kainate.     

The dynamics of cardiac contraction in dogs with alloxan diabetes

Development of alloxan diabetes was accompanied in dogs by regular changes in the phasic structure of the cardiac contractions. Direction of the changes in the indices of phasic analysis in the groups of dogs with an average (up to 144.3 +/- 12.5 mg%) and marked (up to 258.0 +/- 22.8 mg%) increase in the blood sugar level was the same, but their expression increased with the elevation of hyperglycemia. In marked hyperglycemia there was established an increase (in comparison with normal values) of the index of myocardial tension, a shortening of the period of ejection, a decrease of the mechanical coefficient and of the intrasystolic index, this coinciding with the character of changes of the corresponding indices in the patients suffering from diabetes mellitus, depending on the severity of the disease. The data obtained pointed to the significant influence of metabolic changes (assessed by hyperglycemia level) on the contractile function of the heart.     

Characterization of vestibular potentials evoked by linear acceleration pulses in the chinchilla

The objective of this study was to improve vestibular evoked potentials as a qualitative parameter for vestibular function in small laboratory animals. Linear upward acceleration pulses (up to 8 g within 1 ms) were applied to the head of anesthetized chinchillas. Electrophysiologic responses recorded by a chronically implanted electrode within the facial nerve canal consisted of an initial negative potential, labeled N1, within the first millisecond following the onset of acceleration. This potential was followed by a series of positive and negative potentials found to be highly labile to acoustic masking. The initial negative potential was only minimally sensitive to acoustic masking and persisted following surgical cochlear ablation, but completely disappeared following administration of potassium chloride into the inner ear. Recorded from the contralateral ear, N1 was unaffected by these procedures. Amplitudes of N1 decreased with attenuating stimulus intensity (1.45 microV/dB), whereby N1 latencies slightly increased (-0.015 ms/dB). These data, when coupled with the ability to completely abolish N1 with potassium intoxication while the contralateral ear remained intact, indicate that this potential represents electrophysiologic activity resulting from activation of the ipsilateral vestibular labyrinth.     

Variation in cytoskeletal assembly during spreading of progressively subcultivated human embryo fibroblasts (IMR-90)

The cytoskeletons of early and late passage IMR-90 human diploid fibroblasts have been directly imaged in replicas of Triton X-100 extracted cells during spreading following reseeding. All cells from both young and sensescent cultures exhibit a cytoskeletal network of actin microfilaments, intermediate (10 nm) filaments, microtubules, and interconnecting thin filaments (6-8 nm in diameter) which do not interact with heavy meromyosin. Early passage cells assemble linear aggregates of actin filaments within 1 h of spreading. By 4 h of incubation, these bundles establish a structural bond with the cell membrane which results in resistance by the plasmalemma to detergent extraction at these sites. Furthermore, these membrane regions are associated with developing stress fibers of well-spread cells. In contrast, late passage cells exhibit slower spreading which correlates with a retarded assembly of actin bundles. In addition, by 8 h of spreading, cells of older cultures do not exhibit the regions of membrane-actin interaction which impart detergent resistance to the plasmalemma. We conclude that the ability to reassemble actin-actin and actin-membrane association during cell spreading is reduced with increased serial subcultivation of cells.