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Biomedical subjects

A M Manning

Publications and source records attributed to A M Manning.

At least 55 records · Page 3Linked to original sources
PubMed1995

Selective inhibition of E-selectin, vascular cell adhesion molecule-1, and intercellular adhesion molecule-1 expression by inhibitors of I kappa B-alpha phosphorylation.

The promoters of the E-selectin, vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) genes contain recognition sequences for the inducible nuclear transcription factor kappa B (NF-kappa B). We demonstrate that the appearance of NF-kappa B DNA-binding activity in the nucleus of TNF-alpha-stimulated HUVEC is associated with the rapid phosphorylation and subsequent degradation of I kappa B-alpha, the cytoplasmic inhibitor of NF-kappa B. Serine protease inhibitors prevented the TNF-alpha-induced accumulation of phosphorylated I kappa B-alpha, and prevented I kappa B-alpha degradation and the appearance of NF-kappa B DNA-binding activity. These inhibitors had no direct effect upon the ability of NF-kappa B to bind its cognate recognition sequences, nor upon the DNA-binding activities of other transcription factors. Inhibition of I kappa B-alpha proteolysis resulted in the inhibition of the cytokine- and LPS-induced transcriptional up-regulation and cell-surface expression of E-selectin, VCAM-1, and ICAM-1. In contrast, the TNF-alpha-induced expression of plasminogen activator inhibitor-1 and the constitutive expression of ICAM-2 were unaffected. These inhibitors also had no effect on cellular viability or rates of RNA or protein synthesis. Inhibitors of other proteases and various protein kinases had no effect on the cytokine-induced expression of these endothelial adhesion molecules. These findings indicate that it is possible, using a single pharmacologic agent, to selectively inhibit the expression of the E-selectin, VCAM-1, and ICAM-1 genes without affecting the constitutive or inducible expression of other genes. Pharmacologic inhibition of I kappa B-alpha proteolysis represents a novel approach to the development of anti-inflammatory therapeutics.

Cells, Cultured↗
PubMed1995

Differential induction of mRNA for ICAM-1 and selectins in hepatocytes, Kupffer cells and endothelial cells during endotoxemia.

Intercellular adhesion molecule-1 (ICAM-1) and selectins (E- and P-selectin) mRNAs were determined in individual liver cell types by Northern blot analysis before and after injection of endotoxin. A constitutive expression of ICAM-1 mRNA was found in endothelial cells and Kupffer cells but not in hepatocytes. All three cell types showed upregulation of ICAM-1 mRNA after endotoxin. No constitutive selectin expression could be detected in any liver cell, but endotoxin induced massive synthesis of E- and P-selectin mRNA in endothelial cells and Kupffer cells. The differential expression of cellular adhesion molecules in the liver is consistent with the involvement of selectins in neutrophil rolling in the vasculature and ICAM-1 in transendothelial migration and adherence to parenchymal cells.

Analysis of Variance↗
PubMed1995

The temporal profiles of ICAM-1 protein and mRNA expression after transient MCA occlusion in the rat.

Leukocytes may contribute to ischemic cell damage. ICAM-1 expression on endothelial cells facilitates the migration of leukocytes into tissue. Therefore, we measured the temporal profiles of ICAM-1 mRNA and protein in rat brain after transient (1 or 2 h) of middle cerebral artery (MCA) occlusion. Male Wistar rats (n = 86) were subjected to 1 or 2 h MCA of occlusion, or 2 h of MCA occlusion followed by reperfusion for a variety of durations ranging from 1 h to 1 week. 10 additional control animals were employed. ICAM-1 mRNA and protein were measured during ischemia and reperfusion, and immunohistochemical methods were used to identify specific cell types expressing ICAM-1. ICAM-1 mRNA was detected 1 h after the onset of ischemia. mRNA maximized at 10 h of reperfusion and persisted out to 1 week of reperfusion. ICAM-1 significantly increased in microvascular endothelial cells at 2 h of reperfusion, maximized at 46 h and persisted out to 1 week of reperfusion (P < 0.05). ICAM-1 mRNA and protein are present in ischemic brain early after the onset of ischemia and reperfusion, respectively. These data provide support for the role of ICAM-1 in mediating leukocyte-endothelial adhesion after transient MCA occlusion in the rat.

Animals↗
PubMed1995

Cloning and comparative sequence analysis of the gene encoding canine intercellular adhesion molecule-1 (ICAM-1).

Canine intercellular adhesion molecule-1 (ICAM-1) plays a primary role in the adherence of canine neutrophils to endothelial cells and in the cytotoxicity of canine neutrophils for adult cardiac myocytes. We have cloned the canine ICAM-1 gene and have analyzed the conservation of ICAM-1 amino acid (aa) sequences in man, chimpanzee, mouse, rat and dog. Canine ICAM-1 displays 61% identity with human ICAM-1. Cys residues critical to the immunoglobulin (Ig) fold structure and four sites of N-linked glycosylation are absolutely conserved in ICAM-1 from all species. Residues in the cytoplasmic tail associated with cytoskeletal alpha-actinin binding are highly conserved, supporting the hypothesis that intracellular attachment is indeed important for ICAM-1 function. Residues critical for human ICAM-1 binding to the beta 2-integrin leukocyte-function-associated antigen 1 (LFA-1) are highly conserved between all species, whereas those residues demonstrated to play an important role in interaction of human ICAM-1 with macrophage activation complex 1 (Mac-1) are not highly conserved. Residues critical for ICAM-1 binding to rhinovirus and malaria-infected red blood cells (IRBC) are not highly conserved.

Amino Acid Sequence↗
PubMed1995

Cloning of a canine cDNA homologous to the human transforming growth factor-beta 1-encoding gene.

A 1369-bp cDNA that encodes a homolog of the human transforming growth factor-beta 1 (TGF-beta 1) has been isolated from canine endothelial cells using a combination of PCR and traditional plaque-screening methods. The deduced 390-amino-acid sequence of the canine TGF-beta 1 precursor has 91-94% identity to those deduced from the previously described human and mouse TGF-beta 1 clones.

Amino Acid Sequence↗
PubMed1995

Transcriptional regulation of endothelial cell adhesion molecules: a dominant role for NF-kappa B.

A growing body of evidence demonstrates that elevated expression of the endothelial cell adhesion molecules E-selectin, VCAM-1 and ICAM-1 at sites of inflammation in vivo is due in whole or part to the upregulation of transcription of their respective genes. Pharmacologic antagonism of transcription from these genes may therefore represent a novel approach to the development of anti-inflammatory therapeutics. This paper reviews our current understanding of nuclear factors which act to regulate the transcriptional activity of the E-selectin, VCAM-1 and ICAM-1 genes, and discusses that evidence which suggests that the nuclear transcription factor NF-kappa B acts as a dominant regulator of transcription from these genes.

Binding Sites↗
PubMed1995

Anti-intercellular adhesion molecule-1 antibody reduces ischemic cell damage after transient but not permanent middle cerebral artery occlusion in the Wistar rat.

BACKGROUND AND PURPOSE: Postischemic cerebral inflammation may contribute to ischemic cell damage. Intercellular adhesion molecule-1 (ICAM-1) is a glycoprotein expressed on endothelial cells that facilitates leukocyte adhesion. We investigated the effect of administration of an anti-ICAM-1 antibody (1A29) on ischemic cell damage after transient (2-hour) or permanent middle cerebral artery (MCA) occlusion in the Wistar rat. METHODS: Groups studied were as follows: (1) transient MCA occlusion: rats were subjected to 2 hours of MCA occlusion, and after 1 hour of reperfusion they were treated with 1A29 (n = 11) or an isotype control antibody (n = 9); and (2) permanent MCA occlusion: rats were treated with 1A29 (n = 9) or an isotype control antibody (n = 7) 2 hours after onset of MCA occlusion. All animals were killed 1 week after onset of ischemia. Brain sections were stained with hematoxylin and eosin for histological evaluation. RESULTS: Significant reductions (P < .05) in both volume (44%) of the ischemic lesion and weight loss were found in animals subjected to transient MCA occlusion and treated with 1A29 compared with vehicle-treated animals. In contrast, in animals subjected to permanent MCA occlusion the lesion and the temporal profile of body weight were not altered by 1A29 administration. CONCLUSIONS: Ischemic cell damage is promoted by postischemic inflammatory response after 2 hours of transient MCA occlusion, and ischemic cell damage is reduced by administration of an anti-ICAM-1 antibody during reperfusion.

Animals↗
PubMed1995

Interleukin-8 gene induction in the myocardium after ischemia and reperfusion in vivo.

Neutrophil adhesion and direct cytotoxicity for cardiac myocytes require chemotactic stimulation and are dependent upon CD18-ICAM-1 binding. To characterize the potential role of IL-8 in this interaction, canine IL-8 cDNA was cloned and the mature recombinant protein expressed in Escherichia coli BL21 cells. Recombinant canine IL-8 markedly increased adhesion of neutrophils to isolated canine cardiac myocytes. This adhesion resulted in direct cytotoxicity for cardiac myocytes. Both processes were specifically blocked by antibodies directed against CD18 and IL-8. In vivo, after 1 h of coronary occlusion, IL-8 mRNA was markedly and consistently induced in reperfused segments of myocardium. IL-8 mRNA was not induced in control (normally perfused) myocardial segments. Minimal amounts of IL-8 mRNA were detected after 3 or 4 h of ischemia without reperfusion. Highest levels of induction were evident in the most ischemic myocardial segments. IL-8 mRNA peaked in the first 3 h of reperfusion and persisted at high levels beyond 24 h. IL-8 staining was present in the inflammatory infiltrate near the border between necrotic and viable myocardium, as well as in small veins in the same area. These findings provide the first direct evidence for regulation of IL-8 in ischemic and reperfused canine myocardium and support the hypothesis that IL-8 participates in neutrophil-mediated myocardial injury.

Amino Acid Sequence↗
PubMed1995

Protection of ischemic/reperfused canine myocardium by CL18/6, a monoclonal antibody to adhesion molecule ICAM-1.

OBJECTIVE: A blocking monoclonal antibody to intercellular adhesion molecule-1 (ICAM-1), CL18/6, previously has been demonstrated to inhibit neutrophil attachment to isolated vascular endothelium and cardiomyocytes. Due to the well known participation of ICAM-1 in the inflammatory responses associated with myocardial ischemia/reperfusion injury, we investigated if CL18/6 could attenuate myocardial ischemia/reperfusion injury in vivo. METHODS: Saline (3-5 ml, i.v., n = 6), non-blocking control MAb CL18/1D8 or CL18/6 (both 0.5 mg kg-1, i.v., n = 4) were administered prior to coronary occlusion (1 h) and subsequent reperfusion (5 h) produced by inflation of a coronary balloon angioplasty catheter in isoflurane-anesthetized, closed-chest dogs. Heart rate and arterial pressure were measured, and regional myocardial blood flow (rMBF), and myeloperoxidase activity (MPO) to index local neutrophil sequestration, were determined. Myocardial infarct size (IS) was evaluated using the tetrazolium staining technique and expressed as a percent of area at risk (AR). RESULTS: Changes in heart rate and arterial pressure were insignificant throughout the experiment. rMBF (mean +/- s.e.m.) in the ischemic subendocardium for each treatment group was: Saline (0.07 +/- 0.02 ml min-1 g-1); CL18/1D8 (0.04 +/- 0.02); CL18/6 (0.06 +/- 0.02). IS/AR% was: saline (37 +/- 3%); CL18/1D8 (39 +/- 9%); CL18/6 (15 +/- 4%*); * = significantly different from CL18/1D8 and saline, P < 0.05. MPO assayed from AR immediately adjacent to the infarct was significantly reduced below infarct MPO only in the CL18/6 treated group-36%). CONCLUSIONS: The results indicate that CL18/6 antagonism of ICAM-1 provided cardioprotection associated with reduced neutrophil activity in vulnerable myocardium, and suggest that ICAM-1 mediated neutrophil sequestration in endangered cardiac tissue is an important mechanism of myocardial ischemia/reperfusion injury.

Animals↗
PubMed1995

Antibodies to intercellular adhesion molecule-1 ameliorate the inflammatory response in acetic acid-induced inflammatory bowel disease.

The transendothelial migration of leukocytes in many inflammatory responses is now believed to be dependent on the interaction of leukocyte and endothelial cell-derived adhesion molecules. To examine the role of intercellular adhesion molecule-1 (ICAM-1) in the development of inflammation in a rat model of colitis, we investigated the effects of antibodies to rat ICAM-1 given 24 hrs after inflammation was induced by acetic acid. Antibodies to rat ICAM-1 substantially ameliorated the inflammatory response as indicated by a reduction in gross inflammatory characteristics, tissue/body weight ratio, myeloperoxidase activity and superoxide levels. The results demonstrate that ICAM-1 plays an important role in the development of inflammatory bowel disease in rats. The use of antibodies to ICAM-1 to inhibit the adherence of leukocytes to endothelium, may be of potential therapeutic value in the treatment of inflammatory bowel disease in man.

Acetates↗
PubMed1995

Cytokine-induced upregulation of hepatic intercellular adhesion molecule-1 messenger RNA expression and its role in the pathophysiology of murine endotoxin shock and acute liver failure.

Neutrophil-induced liver injury during endotoxemia is dependent on the adhesion molecule Mac-1 (CD11b/CD18) on neutrophils. The potential involvement of its counterreceptor, intercellular adhesion molecule-1 (ICAM-1), in the pathogenesis was investigated after administration of 100 micrograms/kg Salmonella abortus equi endotoxin (ET) in galactosamine-sensitized mice (Gal). In ET-sensitive mice (C3Heb/FeJ), which generated large amounts of tumor necrosis factor-alpha (TNF-alpha), massive neutrophil infiltration and severe liver injury were observed. In an ET-resistant strain (C3H/HeJ), which did not generate TNF-alpha Gal/ET failed to cause neutrophil accumulation or injury. ICAM-1 messenger RNA (mRNA), negligible in control livers, was selectively induced by Gal/ET in ET-sensitive mice. Intravenous injection of murine TNF-alpha, interleukin-1 alpha (IL-1 alpha) or IL-I beta (13 to 23 micrograms/kg) strongly induced the ICAM-1 message in both strains, showing a comparable capacity for ICAM-1 mRNA synthesis. All cytokines caused similar neutrophil accumulation in the liver; however, only Gal/TNF-alpha also caused upregulation of Mac-1 on circulating neutrophils and liver injury. The anti-murine ICAM-1 monoclonal antibody YN.1 (3 mg/kg) attenuated liver injury in ET-sensitive mice by 67% to 90% compared with isotype-matched control antibody-treated animals but did not reduce neutrophil accumulation in hepatic sinusoids. Our data suggest that the cytokines TNF-alpha and IL-1 are the main mediators responsible for upregulation of ICAM-1 mRNA in the liver during endotoxemia. The upregulation of both adhesion molecules, ICAM-1 and Mac-1, is necessary for a neutrophil-induced liver injury to occur. (ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Intercellular adhesion molecule 1 (ICAM-1) expression and its role in neutrophil-induced ischemia-reperfusion injury in rat liver.

The potential role of intercellular adhesion molecule-1 (ICAM-1) in the pathogenesis of reperfusion injury was investigated in male Fischer rats subjected to 45 min of hepatic ischemia and 24 h of reperfusion. ICAM-1 mRNA levels increased during ischemia in the ischemic liver lobes; however, during reperfusion mRNA levels increased in both the ischemic and nonischemic lobes. Immunohistochemical evaluation indicated ICAM-1 expression only on sinusoidal lining cells in controls; ischemia-reperfusion enhanced ICAM-1 expression in the sinusoids and induced some expression on hepatocytes. The monoclonal anti-ICAM-1 antibody 1A29, but not an immunoglobulin G control antibody, administered at 1 h and 8 h of reperfusion (2 mg/kg) significantly attenuated liver injury as indicated by 51% lower plasma alanine aminotransferase activities and 32-36% less hepatic necrosis at 24 h without affecting reactive oxygen formation by Kupffer cells and hepatic neutrophils. Although 1A29 reduced neutrophil extravasation in a glycogen peritonitis by 60%, the antibody had no significant effect on hepatic neutrophil infiltration during reperfusion. These data suggest that ICAM-1 plays a significant role during the neutrophil-dependent injury phase after hepatic ischemia and reperfusion and therefore blocking this adhesion molecule may have therapeutic potential against postischemic acute liver failure.

Animals↗
PubMed1995

NF-kappa B is activated during acute inflammation in vivo in association with elevated endothelial cell adhesion molecule gene expression and leukocyte recruitment.

Leukocytes accumulate at sites of inflammation in response to the induced expression of endothelial cell adhesion molecules. The nuclear transcription factor kappa B (NF-kappa B) plays a critical role in the cytokine-induced expression of these genes in cultured endothelium. We examined the relationship between NF-kappa B activation and endothelial cell adhesion molecule gene expression in vivo during the initiation of acute inflammation. Nuclear NF-kappa B DNA-binding activity was rapidly increased within lung and heart tissues of rats administered endotoxin, consistent with the translocation of NF-kappa B complexes from the cytoplasm to the nucleus. This NF-kappa B was composed of p50 and p65 subunits, and could bind NF-kappa B elements in the E-selectin promoter. NF-kappa B activation was maximal within 30 min and persisted for at least 3 hr after endotoxin treatment. NF-kappa B activation preceded the transcriptional activation of the P-selectin, E-selectin, VCAM-1, and ICAM-1 genes. In the lung, increased expression of P-selectin and ICAM-1 protein was detected immunohistochemically. These molecular events were temporally associated with the sequestration of leukocytes and the development of pulmonary inflammation. NF-kappa B activation is therefore an early event in the initiation of acute inflammation in vivo. This molecular pathway may be of consequence in the pathogenesis of acute inflammatory disease.

Animals↗
PubMed1994

Cloning, sequence comparison and in vivo expression of the gene encoding rat P-selectin.

We have cloned the cDNA encoding rat P-selectin (Psel) and have examined the regulation of Psel expression in vivo. Sequence analysis of the complete Psel cDNA demonstrated significant nucleotide and amino-acid identity with human and mouse Psel. Similar to mouse Psel, the rat sequence lacks the equivalent of human complement regulatory protein-like repeat 2 (CR2). Seven potential N-linked glycosylation sites are conserved between the three species, suggesting that carbohydrate modification may play an important role in Psel function. To examine expression of Psel in vivo, levels of Psel mRNA were examined in several different tissues after systemic administration of lipopolysaccharide (LPS). Psel mRNA was undetectable in tissues of vehicle-treated animals. By 3 after LPS administration, Psel mRNA levels were elevated in all tissues examined, the highest levels being seen in the lung. Significant increases in Psel mRNA were also seen in the heart, thymus, spleen and kidney. By 24 h after LPS, mRNA levels for Psel remained elevated in the lung, heart, kidney, thymus and small intestine. Psel mRNA was not detectable in total RNA isolated from purified rat platelets, suggesting that the increased levels of Psel mRNA were the result of upregulation of endothelial gene expression. In addition, only minimal levels of platelet factor 4 mRNA (PF4), used as a platelet-specific marker, were observed in the tissues studied. These data demonstrate that part of the response to acute inflammation in vivo includes the rapid increase in endothelial Psel expression.

Amino Acid Sequence↗
PubMed1994

Interdomain linkage in the polymeric hemoglobin molecule of Artemia.

Artemia has evolved the longest known concatenation of hemoglobin domains, the alpha subunit containing nine domains and the beta subunit having a similar size. Translation of the cDNA sequence of the alpha subunit reveals eight regions of inter-domain polypeptide linking together the nine heme-binding domains, together with partially analogous sequences preceding the first domain and following the last. Analysis of the structural possibilities of the linker sequences suggests how the domains may be organized in the subunit. The interdomain linker sequences were 14%-64% identical (62%-91% similar by Dayhoff substitution matrix) and approximately 14 residues in length including a consensus -Val-Asp-Pro-Val-Thr-Gly-Leu-. The linker composition resembled that of the 11 amino acid pre-A leader sequence of Petromyzon marinus (lamprey) hemoglobin V, the structure of which is known. Prediction of structure from the Artemia linker sequences indicated a nonhelical, turn-associated linker which could be modeled to the Petromyzon leader. Measurements confirmed that such a structure could support the packing of nine Artemia domains into a polymeric subunit of annular shape, two of which subunits (which can be similar or dissimilar) comprise the physiological molecule.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Attenuation of oxidant-induced lung injury by 21-aminosteroids (lazaroids): correlation with the mRNA expression for E-selectin, P-selectin, ICAM-1, and VCAM-1.

We compared the effects of treatment with methylprednisolone or the 21-aminosteroids, U-74389 and U-74006F (Tirilizad mesylate), on hyperoxic lung injury and the associated expression of mRNA for several adhesion molecules in rats. Inhalation of > 95% oxygen for up to 72 hr in Sprague-Dawley rats produced a marked increase in lung weight and an accumulation of fluid in the thorax when compared with air-breathing controls. Hyperoxia also induced a marked neutrophil-rich influx of inflammatory cells into the bronchial lumen as measured by bronchoalveolar lavage. Neutrophil numbers in bronchoalveolar lavage fluid peaked after 60 hr of exposure to s 95% oxygen; this was associated with a marked upregulation of mRNA for the adhesion molecules P-selectin and E-selectin but not VCAM-1. mRNA for ICAM-1 was constitutively expressed at high levels in both air-breathing controls and in the lungs of rats exposed to high concentrations of oxygen. Pretreatment with the 21-aminosteroids reduced hyperoxic lung damage and improved survival times in animals exposed to > 95% oxygen. However, treatment with methylprednisolone significantly decreased survival times. Treatment with U-74389 did not significantly (p > 0.05) inhibit the BAL neutrophilia and did not significantly (p > 0.05) reduce hyperoxia-induced increases in mRNA expression for P-selectin and E-selectin. The inhibition of hyperoxic lung damage coupled with improved survival seen in treated animals suggests that 21-aminosteroids may provide valuable treatments for pulmonary disorders in which oxidant damage has been implicated.

Animals↗
PubMed1994

Transfection and expression of mutant p53 protein does not alter the in vivo or in vitro growth characteristics of the AA/C1 human adenoma derived cell line, including sensitivity to transforming growth factor-beta 1.

Mutation of the p53 gene is thought to be a late event in human colorectal carcinogenesis, involved in the malignant conversion of the adenoma to the carcinoma. One of the questions that we hoped to address was whether, in vivo, a single mutational event in one p53 gene is sufficient to confer a significant growth advantage on a colonic epithelial cell. Such a growth advantage could result either from an increase in growth rate and/or loss of response to inhibitory growth signals naturally present in the colonic crypt. We therefore introduced the pC53-SCX3 143 (Val-Ala) p53 mutation into a non tumorigenic adenoma derived cell line, AA/C1, which contained a truncating APC mutation, activating K-ras mutation but was wild-type for the p53 protein. High levels of mutant p53 protein were detected in the pC53-SCX3 transfected AA/C1 cell lines but was found not to affect either the in vitro (colony forming efficiency, anchorage independence) or in vivo (tumorigenicity in nude mice) growth, when compared to vector control or the parental AA/C1 cell line. In addition, to test whether the cells become less sensitive to inhibitory growth factors, the response of the cell lines to the naturally occurring growth inhibitor TGF beta was also investigated. Even though TGF beta had previously been implicated in the control of growth of intestinal epithelium, expression of the mutant p53 protein did not affect the sensitivity of the parental AA/C1 cell line to TGF beta. Under the experimental conditions tested expression of the 143 (Val-Ala) p53 protein was unable to affect the in vitro or in vivo growth characteristics of the adenoma derived AA/C1 cell line. When compared to other studies, these results suggest that the genetic background of the individual recipient cell may greatly influence the effect of expression of a particular p53 mutation.

Adenoma↗