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Biomedical subjects

M O'Connor

Publications and source records attributed to M O'Connor.

At least 109 records · Page 6Linked to original sources
PubMed1997

Ursodeoxycholate inhibits induction of NOS in human intestinal epithelial cells and in vivo.

Ursodeoxycholate (UDCA) has anti-inflammatory and chemoprotective effects in animal models of inflammatory bowel disease and colon cancer. Because overproduction of nitric oxide (NO) by the inducible isoform of NO synthase (iNOS) is implicated in the pathogenesis of these conditions, we investigated the ability of UDCA to inhibit NO production in transformed human intestinal epithelial (DLD-1) cells. Nitrite/nitrate production was measured by the Griess reaction, enzymatic activity of iNOS was assessed by conversion of L-arginine to L-citrulline, and protein and mRNA were measured by Western and Northern blotting. Dose-dependent inhibition of interleukin-1 beta- and interferon-gamma-stimulated nitrite/nitrate production was observed when cells were preincubated for 6 h with UDCA (0-800 microM), and a substantial inhibition (81 +/- 3.2%) was seen at 500 microM. In cytokine-stimulated cells, UDCA reduced iNOS mRNA, protein, and enzyme activity without exerting cytotoxicity. UDCA had a minimal direct inhibitory effect on iNOS enzyme activity. UDCA pretreatment also reduced the expression of iNOS in the colonic epithelium of rats treated with bacterial lipopolysaccharide. Thus UDCA inhibits the induction of epithelial iNOS in vitro and in vivo, and this effect may contribute to the anti-inflammatory and chemoprotective actions of UDCA.

Adenocarcinoma↗
PubMed1997

Peroxynitrite causes energy depletion and increases permeability via activation of poly (ADP-ribose) synthetase in pulmonary epithelial cells.

Recent studies show that peroxynitrite is a potent trigger of DNA strand breakage, which in turn activates the nuclear repair enzyme poly (ADP-ribose) synthetase (PARS), resulting in a cellular energy deficit. Here we present evidence that treatment of A549 human pulmonary epithelial cells with peroxynitrite (1 mM) results in ADP-ribosylation, NAD+ depletion, inhibition of mitochondrial respiration, and increased epithelial paracellular permeability. The PARS inhibitor 3-aminobenzamide (1 mM) provided a significant, partial protection against the energetic and functional changes. Similarly, inhibition of PARS activity by 3-aminobenzamide reduced the peroxynitrite-induced suppression of mitochondrial respiration in BEAS-2B human bronchial epithelial cells. Thus, PARS activation and energy depletion represents one of the pathways of peroxynitrite-mediated epithelial toxicity. Inhibition of PARS may improve cellular energy homeostasis in pathophysiologic conditions associated with peroxynitrite generation.

Benzamides↗
PubMed1997

Metabolic and structural effects of insulin-like growth factor-I and high-protein diet on dystrophic hamster skeletal muscle.

In muscular dystrophy (MD) there is an imbalance between muscle protein synthesis and protein degradation, which results in a net muscle catabolism, along with muscle wasting and weakness. Using a dystrophic hamster model (BIO 53.58), we examined the chronic (8 weeks) effects of two factors that may enhance muscle protein synthesis and inhibit protein degradation, namely, insulin-like growth factor-I (rhIGF-I) and high-protein diet (HPD). Protein synthesis was determined by measuring the incorporation of 14C phenylalanine into perfused leg muscle, while protein degradation was calculated from the release of tyrosine from the same perfused muscle. Urinary 3-methylhistidine excretion was used as an indicator of myofibrillar degradation. Treatment of dystrophic hamsters with rhIGF-I, HPD, or a combination of the two for 8 weeks resulted in significant decreases in total and myofibrillar degradation when compared with untreated dystrophic animals (P < 0.05) but had minimal effects on protein synthesis. Significant morphologic improvements (P < 0.05), including a normalization and greater uniformity of muscle fibers, were also seen in rhIGF-I- and rhIGF-I + HPD-treated animals. rhIGF-I and HPD were effective in reducing the excessive proteolysis seen in dystrophic muscle, and this reduced proteolysis resulted in improvement of muscle morphology.

Animals↗
PubMed1997

CXCR-4 (Fusin), a co-receptor for the type 1 human immunodeficiency virus (HIV-1), is expressed in the human brain in a variety of cell types, including microglia and neurons.

Entry of the type 1 human immunodeficiency virus into most cells requires the presence of the CD4 protein in combination with one of several recently described co-receptors. CXCR-4 (fusin) was the first identified, and it serves as co-receptor for T-cell-line tropic (T-tropic) HIV-1 isolates. To determine the expression of CXCR-4 in the brain, a major target of HIV pathology, we used immunohistochemistry and reverse transcriptase polymerase chain reaction with CXCR-4-specific antibodies and probes. We found that CXCR-4 was expressed in several cell types in brain, but notably in neurons and microglia, a finding that was replicated in tissue culture. The study of the expression of CXCR-4 in the brain, which may be one of many chemokine receptors in the central nervous system, may provide further insight into the interactions between brain cells, pathogens, and the immune system, and help understand the pathogenesis of HIV dementia.

AIDS Dementia Complex↗
PubMed1997

Role of peroxynitrite in the protein oxidation and apoptotic DNA fragmentation in vascular smooth muscle cells stimulated with bacterial lipopolysaccharide and interferon-gamma.

In the present study, we investigated the role of endogenous and exogenous peroxynitrite in the process of DNA fragmentation and protein oxidation in cultured rat aortic smooth muscle cells. Peroxynitrite induced DNA fragmentation over a 24 hr period. The effect of peroxynitrite was unaffected by pretreatment with 3-aminobenzamide, an inhibitor of the nuclear enzyme poly (ADP-ribose) synthetase (PARS). Stimulation of the smooth muscle cells with bacterial lipopolysaccharide and interferon-gamma produced nitric oxide and peroxynitrite, and resulted in a significant degree of apoptotic DNA fragmentation. The nitric oxide synthase inhibitor NG-methyl-L-arginine (3 mM), but not the PARS inhibitor 3-aminobenzamide (1 mM), reduced the DNA fragmentation. Stimulation with bacterial lipopolysaccharide and interferon-gamma also caused a marked oxidation of proteins in the smooth muscle cells, which was inhibited by NG-methyl-L-arginine, as well as by the superoxide dismutase mimetic Mn(III)tetrakis (4-benzoic acid) porphyrin. Based on these data, we propose a role for peroxynitrite-mediated, PARS-independent pathways in the apoptotic process and in the protein oxidation in bacterial lipopolysaccharide and interferon-gamma-stimulated smooth muscle cells.

Animals↗
PubMed1996

Functional Escherichia coli 23S rRNAs containing processed and unprocessed intervening sequences from Salmonella typhimurium.

We have introduced the intervening sequence (IVS) from 23S rRNA of the rrnD operon of Salmonella typhimurium into the equivalent position of Escherichia coli 23S rRNA. Salmonella typhimurium 23S rRNA is fragmented due to the RNase III-dependent removal of the approximately 100 nt stem-loop structure that comprises the IVS. In this study, we have found that insertion of the S. typhimurium IVS into E. coli 23S rRNA causes fragmentation of the RNA but does not affect ribosome function. Cells expressing the fragmented 23S rRNA exhibited wild-type growth rates. Fragmented RNA was found in the actively translating polysome pool and did not alter the sedimentation profile of ribosomal subunits, 70S ribosomes or polysomes. Finally, hybrid 23S rRNA carrying the A2058G mutation conferred high level erythromycin resistance indistinguishable from that of intact 23S rRNA carrying this mutation. These observations indicate that the presence of this IVS and its removal are phenotypically silent. As observed in an RNase III-deficient strain, processing of the IVS was not required for the production of functional ribosomes.

Base Sequence↗
PubMed1996

The influence of base identity and base pairing on the function of the alpha-sarcin loop of 23S rRNA.

The alpha-sarcin loop of large subunit rRNAs is one of the sites of interaction of elongation factors with the ribosome, and the target of the cytotoxins alpha-sarcin and ricin. Using a genetic selection for increased frameshifting in a reporter gene, we have isolated a C --> U mutation at position 2666 in the alpha-sarcin loop. In the NMR-derived structure of the loop, bases equivalent to 2666 and 2654 are paired via a non-canonical base pairing interaction. Each of the three base substitutions at C2666 and A2654 was constructed by site-directed mutagenesis of a plasmid borne copy of the rrnB operon of Escherichia coli. Only the C2666 --> U and A2654 --> G mutations that resulted in the formation of canonical A-U and C-G base pairs respectively, increased the levels of stop codon readthrough and frameshifting. The effects of different base pair combinations at positions 2666 and 2654 on ribosome function were then tested by constructing and analyzing all possible base combinations at these sites. All A --> G base substitution mutations at position 2654 and C --> U substitutions at position 2666 increased the levels of translational errors. However, these effects were greatest when G2654 and U2666 had the potential to engage in standard Watson-Crick base pairing interactions. These data indicate that base identity as well as base pairing interactions are important for the function of this essential component of the large subunit rRNA.

Base Composition↗
PubMed1996

DNA strand breakage, activation of poly (ADP-ribose) synthetase, and cellular energy depletion are involved in the cytotoxicity of macrophages and smooth muscle cells exposed to peroxynitrite.

The free radicals nitric oxide and superoxide anion react to form peroxynitrite (ONOO-), a highly toxic oxidant species. In vivo formation of ONOO- has been demonstrated in shock and inflammation. Herein we provide evidence that cytotoxicity in cells exposed to ONOO- is mediated by DNA strand breakage and the subsequent activation of the DNA repair enzyme poly(ADP ribose) synthetase (PARS). Exposure to ONOO- (100 microM to 1 mM) inhibited mitochondrial respiration in cultured J774 macrophages and in rat aortic smooth muscle cells. The loss of cellular respiration was rapid, peaking 1-3 h after ONOO- exposure, and reversible, with recovery after a period of 6-24 h. The inhibition of mitochondrial respiration was paralleled by a dose-dependent increase in DNA strand breakage, reaching its maximum at 20-30 min after exposure to ONOO-. We observed a dose-dependent increase in the activity of PARS in cells exposed to ONOO-. Inhibitors of PARS such as 3-aminobenzamide (1 mM) prevented the inhibition of cellular respiration in cells exposed to ONOO-. Activation of PARS by ONOO--mediated DNA strand breakage resulted in a significant decrease in intracellular energy stores, as reflected by a decline of intracellular NAD+ and ATP content. 3-Aminobenzamide prevented the loss of NAD+ and ATP in cells exposed to ONOO-. In contrast, impairment of cellular respiration by the addition of the nitric oxide donors S-nitroso-N-acetyl-DL-penicillamine or diethyltriamine nitric oxide complex, was not associated with the development of DNA strand breaks, in concentrations up to 1 mM, and was largely refractory to PARS inhibition. Our results suggest that DNA damage and activation of PARS, an energy-consuming futile repair cycle, play a central role in ONOO--mediated cellular injury.

Adenosine Triphosphate↗
PubMed1996

HIV-1 infection of cultured human adult oligodendrocytes.

The mechanism through which HIV-1 causes HIV dementia (HIVD) is not well understood. Myelin pallor is a common pathological finding in HIVD and could be explained by a direct infection of oligodendrocytes or interaction with HIV-1 gp 120. To determine if oligodendrocytes could be infected by HIV-1, we purified oligodendrocytes from adult human brain tissues obtained from temporal lobe resections. These cells were exposed to HIV-1 and infectivity was assayed by detection of p24gag antigen, PCR amplification, and cocultivation with CD4+ cells. These results indicate that HIV-1(IIIB and BaL) and one of four primary isolates tested can infect oligodendrocytes, resulting in the production of infectious virus. Furthermore, in an experiment that mimics a potential in vivo scenario, infected microglia were able to transmit virus to oligodendrocytes in a trans-well culture system. These experiments indicate that oligodendrocyte infection should be considered in studying the pathophysiology of HIVD.

Adult↗
PubMed1996

Peroxynitrite-mediated DNA strand breakage activates poly-adenosine diphosphate ribosyl synthetase and causes cellular energy depletion in macrophages stimulated with bacterial lipopolysaccharide.

The inducible isoform of nitric oxide (NO) synthase produces large quantities of NO, a cytotoxic free radical. Recent studies show that treatment with exogenous NO produces DNA strand breaks, activating the nuclear repair enzyme poly(ADP)ribosyltransferase (PARS), which results in ADP ribosylation, NAD+ consumption, and exhaustion of intracellular energy stores. Here we have characterized the cytotoxic effect of endogenous NO and peroxynitrite, a reactive oxidant formed from NO and superoxide. Immunostimulation of J774.2 macrophages with endotoxin resulted in the generation of superoxide (within 1 h) and NO (after 8 h). NO production paralleled an increase in peroxynitrite formation and DNA strand breakage, and a decrease in intracellular NAD+ content and mitochondrial respiration. Inhibition of NO synthase by NG-methyl-L-arginine or S-methyl-isothiourea or inhibition of PARS activity by 3-aminobenzamide or nicotinamide prevented the decrease in mitochondrial respiration and the depletion of NAD+. A similar pattern of free radical formation and cytotoxicity was observed in peritoneal macrophages from endotoxemic rats (formation of NO, superoxide, peroxynitrite, and DNA strand breaks). In vivo treatment with 3-aminobenzamide preserved mitochondrial respiration, NAD+, and ATP. Our data suggest that inflammatory cell injury involved DNA strand breakage and PARS, triggering an energy-consuming, futile repair cycle leading to cellular energy depletion. The active species responsible for the development of DNA strand breaks is peroxynitrite, rather than NO, since exogenous peroxynitrite, but not NO, induces DNA strand breaks. Inhibition of PARS may improve cellular energy homeostasis in patho-physiologic conditions associated with peroxynitrite generation.

Adenosine Diphosphate Ribose↗
PubMed1996

Absence of neglect for mental representations during the intracarotid amobarbital procedure.

BACKGROUND: The unilateral suppression of right-hemisphere electrophysiological activity during the intracarotid amobarbital procedure results in left neglect on visuomotor search tasks. OBJECTIVE: To investigate whether right-hemisphere inactivation during the intracarotid amobarbital procedure also results in left neglect of the mental representation of a remembered scene. DESIGN: Before the intracarotid amobarbital procedure, 16 subjects were asked to draw a room from their homes. Following the right-hemisphere injection, subjects wee tested for the presence of left neglect of extrapersonal space using a Random Letter Cancellation Test. To test for neglect of the mental representation of a remembered scene, they were then asked to report items from the room they had drawn. RESULTS: While 10 of the 16 subjects showed left neglect on the Random Letter Cancellation Test, only one subject showed left neglect of the mental representation. The other 15 subjects recalled approximately equal numbers of items from the left and the right hemispaces. CONCLUSIONS: These results replicate previous findings of left neglect of extra personal space during the intracarotid amobarbital procedure and fail to show concurrent neglect for mental representations. They provide further evidence for the multiplicity and dissociability of the behavioral manifestations of neglect.

Amobarbital↗
PubMed1996

Heterogeneity in cellular and cytokine profiles from multiple samples of tissue surrounding revised hip prostheses.

Previous studies have attempted to define the biologic properties of the bone-implant interface using a single specimen harvested from the periprosthetic tissues. The purpose of this study was to examine the heterogeneity in cellular and cytokine profiles of multiple samples taken from the tissues surrounding revised hip prostheses. Clinical and radiographic data for nine patients undergoing surgical revision was gathered prospectively. Three tissue samples were taken systematically from the acetabular and/or femoral bed. Morphologic characteristics of the tissues were assessed using hematoxylin and eosin-stained sections. Immunohistochemical staining was performed using monoclonal antibodies to identify macrophages (EMB11 and CD68); activated macrophages (Leu M3); total T lymphocytes (Leu 4 and T11); T-helper lymphocytes (Leu 3A and CD4); cytotoxic/suppressor T lymphocytes (Leu 2A and CD3); and fibroblasts (5B5). In situ hybridization was used to identify the mRNA for specific proteins: interleukin (IL)1 alpha and -beta, IL-2, IL-6, transforming growth factor beta, tumor necrosis factor alpha (TNF alpha), platelet-derived growth factor alpha (PDGF alpha), and interferon gamma. A quantitative assessment was performed for each section by calculating the percentage of positively staining cells using a light microscope and grid-counting technique. A random effect analysis of variance was calculated to determine both the variance between samples within each patient and the variance between different patients. Standard deviations contributed by sampling variance and patient variance were calculated and an F test was applied. Tissue samples taken from different regions of the bone-prosthesis interface showed marked heterogeneity in cellular and cytokine profiles. Critical F values indicating a statistically significant degree of variance between different tissue samples were exceeded for macrophages, cytotoxic/suppressor T lymphocytes, and T-helper lymphocytes. The cytokine profile was significantly different for IL-2, PDGF alpha, and TNF alpha. This tissue heterogeneity may be due to different mechanical and biologic environments along the bone-prosthesis interface. Thus, caution must be exercised in defining the biologic properties of the tissue surrounding revised prostheses according to a single biopsy.

Adult↗
PubMed1996

Usefulness and limitations of serum and urine lysozyme levels in the classification of acute myeloid leukemia: an analysis of 208 cases.

The revised French-American-British (FAB) classification system for acute myeloid leukemia (AML) recommends the determination of serum lysozyme (SL) or urine lysozyme (UL) levels as an aid in distinguishing acute myeloblastic leukemia with maturation (FAB M2) from acute myelomonocytic leukemia (M4). We reviewed retrospectively 208 cases of adult leukemia in which SL and/or UL were obtained. Elevated lysozyme levels were not found in any of the M0, M3, or M7 cases, but were increased (false positive) in three (14%) M1 cases, 18 (19%) M2 cases and one (20%) M6 case. Although a UL value in excess of 3x normal was found in most cases of AML M4 and M5, only five (11%) M4 cases and three (20%) M5 cases had SL elevations of this magnitude. Lysozyme levels need to be interpreted in conjunction with other parameters for FAB classification.

Aged↗
PubMed1996

Methylnaltrexone prevents morphine-induced delay in oral-cecal transit time without affecting analgesia: a double-blind randomized placebo-controlled trial.

Methylnaltrexone is a quaternary opioid antagonist with limited ability to cross the blood-brain barrier and the potential to antagonize the peripherally mediated effects of opioids. The effectiveness of methylnaltrexone in preventing morphine-induced changes in gastrointestinal motility and transit without affecting analgesia was evaluated in humans. Twelve healthy volunteers were given intravenous placebo, placebo plus 0.05 mg/kg morphine, or 0.45 mg/kg methylnaltrexone plus 0.05 mg/kg morphine. Oral-cecal transit time was assessed by the pulmonary hydrogen measurement technique, and analgesia was measured with use of the cold-pressor test. Morphine significantly increased oral-cecal transit time from 104.6 +/- 31.1 minutes (mean +/- SD) to 163.3 +/- 39.8 minutes (p < 0.01). Methylnaltrexone prevented 97% of morphine-induced increase in oral-cecal transit time (106.3 +/- 39.8 minutes; not significant compared with baseline; p < 0.01 compared with morphine alone). Methylnaltrexone did not affect the analgesic effect of morphine on both pain intensity and pain bothersomeness ratings. At a higher dose of morphine (0.1 mg/kg), our preliminary results indicated that 0.45 mg/kg methylnaltrexone also prevented the morphine-induced delay in oral-cecal transit time, with no effect on analgesia. Methylnaltrexone may be a useful adjunct to opioids for the relief of opioid-induced constipation.

Adolescent↗
PubMed1996

Recovery of memory and executive function following anterior communicating artery aneurysm rupture.

We studied the recovery of memory and executive function in 10 patients following anterior communicating artery aneurysm (ACoA) rupture and repair. Patients were tested at 2 consecutive points in time following surgery (approximately at 2 and 3 months). At the first testing, the patients divided into 2 groups based on the severity of impairment on executive measures. Both groups had severe anterograde amnesia, but only patients with severe executive impairments had retrograde amnesia with a temporal gradient. At second testing, both groups had persistent severe anterograde amnesia. The dysexecutive group showed significant improvement in executive deficits and in retrograde amnesia, with attenuation of the temporal gradient. Patients with more severe executive impairments had more extensive bilateral frontal lesions than other patients. These results suggest that the cognitive profile following ACoA rupture changes with time. Time postonset following aneurysm rupture and lesion site are both critical for defining the neuropsychological profile, and determining the underlying cognitive mechanisms in this neurological disorder.

Adult↗