Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 30
Filter
Add more filters










Publication year range
1.
Allergy ; 74(1): 89-99, 2019 01.
Article in English | MEDLINE | ID: mdl-29900561

ABSTRACT

BACKGROUND: The receptor for advanced glycation endproducts (RAGE) has been implicated as a critical molecule in the pathogenesis of experimental asthma/allergic airway inflammation (AAI). It has been previously shown that RAGE acts both upstream of interleukin-33 (IL-33) release and downstream of IL-33 release via RAGE-dependent IL-33-induced accumulation of type 2 innate lymphoid cells (ILC2s) in the lungs, which perpetuate type 2 inflammation and mucus metaplasia. However, the mechanism by which RAGE mediates downstream IL-33-induced type 2 inflammatory responses is unknown. OBJECTIVE: This study tested the hypothesis that ILC2s are recruited to the lungs via RAGE-dependent vascular cell adhesion molecule 1 (VCAM-1) expression on lung endothelial cells. METHODS: House dust mite extract, Alternaria alternata extract, or rIL-33 was used to induce AAI/VCAM-1 expression in wild-type (WT) and RAGE-knockout (RAGE-KO) mice. Intravenous (i.v.) anti-VCAM-1 or intraperitoneal (i.p.) ß7 blocking antibody administration was used to determine the role of VCAM-1 in IL-33-induced AAI. RESULTS: Enhanced VCAM-1 expression in the lungs by HDM, AA, or rIL-33 exposure was found to be RAGE-dependent. In addition, stimulation of primary mouse lung endothelial cells with IL-33 induced VCAM-1 expression in WT, but not RAGE-KO cells. Administration of VCAM-1 and ß7-integrin blocking antibodies reduced IL-33-induced eosinophilic inflammation, mucus metaplasia, and type 2 inflammatory responses. CONCLUSION: This study demonstrates that allergen- and cytokine-induced VCAM-1 expression is RAGE-dependent and contributes to lung ILC2 accumulation and downstream eosinophilic inflammation, mucus metaplasia, and type 2 inflammatory responses.


Subject(s)
Asthma/pathology , Endothelial Cells/metabolism , Inflammation/chemically induced , Interleukin-33/pharmacology , Receptor for Advanced Glycation End Products/immunology , Vascular Cell Adhesion Molecule-1/metabolism , Allergens/immunology , Animals , Asthma/chemically induced , Cells, Cultured , Cytokines/metabolism , Interleukin-33/immunology , Lung/pathology , Lymphocytes/immunology , Mice
2.
Histopathology ; 49(1): 66-74, 2006 Jul.
Article in English | MEDLINE | ID: mdl-16842247

ABSTRACT

AIMS: Recent studies suggest the importance of oxidant stress in the progression of pulmonary fibrosis. The aim of this study was to investigate extracellular superoxide dismutase (ECSOD), the major antioxidant enzyme of the extracellular matrix of human lung, in biopsy-proven idiopathic pulmonary fibrosis (IPF) related to usual interstitial pneumonia (UIP). METHODS AND RESULTS: Fibrotic areas and fibroblastic foci in UIP lungs were notable for absence of ECSOD by immunohistochemistry. Western blotting showed significantly lowered immunoreactivity of ECSOD in fibrotic compared with non-fibrotic areas of the diseased lung. The only cell type that showed intense ECSOD positivity in UIP was the interstitial mast cell. In order to investigate the mechanism for ECSOD depletion in fibrotic areas, alveolar epithelial cells were exposed to tumour necrosis factor-alpha and transforming growth factor (TGF)-beta1; TGF-beta suggested a trend towards decreased synthesis. Patients with UIP were also assessed to determine whether this disease is associated with a naturally occurring mutation in ECSOD (Arg213Gly) which leads to a loss of tissue binding of ECSOD. No significant differences could be found in the allele or genotype frequencies of this polymorphism between 63 UIP patients and 61 control subjects. CONCLUSION: Overall, consistent with several other antioxidant enzymes, ECSOD is very low in fibrotic areas of UIP, which may further increase the oxidant burden in this disease.


Subject(s)
Lung Diseases, Interstitial/enzymology , Lung Diseases, Interstitial/pathology , Pulmonary Fibrosis/enzymology , Pulmonary Fibrosis/pathology , Superoxide Dismutase/metabolism , Adult , Aged , Aged, 80 and over , Amino Acid Substitution , Base Sequence , Case-Control Studies , Cell Line , DNA/genetics , Female , Genetic Variation , Genotype , Humans , Immunohistochemistry , Lung/enzymology , Lung Diseases, Interstitial/genetics , Male , Middle Aged , Oxidants/metabolism , Pulmonary Fibrosis/genetics , Superoxide Dismutase/genetics
3.
Eur Respir J ; 24(5): 765-71, 2004 Nov.
Article in English | MEDLINE | ID: mdl-15516670

ABSTRACT

Oxidant stress is a key mechanism for smoking-induced chronic obstructive pulmonary disease (COPD). Smoking has been shown to upregulate several antioxidant enzymes, with potential effects on the prevention of the disease and/or its progression. Superoxide dismutases (SOD)s are the only enzymes capable of consuming superoxide radicals. The purpose of the present study was to investigate SODs in the lungs of nonsmokers, smokers and COPD patients. Manganese superoxide dismutase (MnSOD), copper zinc SOD (CuZnSOD), and extracellular SOD (ECSOD), were investigated by immunohistochemistry in the airways of 13 nonsmokers, 20 smokers and 22 COPD patients with mild-to-moderate disease. Lung tissue homogenates of three nonsmokers and four smokers were used for Western blot and enzyme activity analysis. The expression of MnSOD was higher in the central bronchial epithelium of smokers with COPD and in the alveolar epithelium of smokers without or with COPD than innonsmokers. Lung MnSOD immunoreactivity, evaluated by Western blotting and specific activity, were 33% and 51% higher, respectively, in smokers than in nonsmokers. No major changes could be observed in lung CuZnSOD or ECSOD immunoreactivities. Manganese superoxide dismutase is elevated in the alveolar epithelium of cigarette smokers, probably due to the increased oxidant burden in smokers' lungs.


Subject(s)
Lung/enzymology , Smoking/metabolism , Superoxide Dismutase/analysis , Aged , Aldehydes/analysis , Blotting, Western , Bronchi/enzymology , Epithelium/enzymology , Female , Humans , Immunohistochemistry , Male , Pulmonary Alveoli/enzymology , Pulmonary Disease, Chronic Obstructive/enzymology , Smoking/adverse effects
4.
Free Radic Biol Med ; 31(10): 1198-207, 2001 Nov 15.
Article in English | MEDLINE | ID: mdl-11705698

ABSTRACT

The antioxidant enzyme extracellular superoxide dismutase (EC-SOD) is highly expressed in the extracellular matrix of lung tissue and is believed to protect the lung from oxidative damage that results in diseases such as pulmonary fibrosis. This study tests the hypothesis that proteolytic removal of the heparin-binding domain of EC-SOD results in clearance of the enzyme from the extracellular matrix of pulmonary tissues and leads to a loss of antioxidant protection. Using a polyclonal antibody to mouse EC-SOD, the immunodistribution of EC-SOD in normal and bleomycin-injured lungs was examined. EC-SOD labeling was strong in the matrix of vessels, airways, and alveolar surfaces and septa in control lungs. At 2 d post-treatment, a slight increase in EC-SOD staining was evident. In contrast, lungs examined 4 or 7 d post-treatment, showed an apparent loss of EC-SOD from the matrix and surface of alveolar septa. Notably, at 7 d post-treatment, the truncated form of EC-SOD was found in the bronchoalveolar lavage fluid of bleomycin-treated mice, suggesting that EC-SOD is being removed from the extracellular matrix through proteolysis. However, loss of EC-SOD through proteolysis did not correlate with a decrease in overall pulmonary EC-SOD activity. The negligible effect on EC-SOD activity may reflect the large influx of intensely staining inflammatory cells at day 7. These results indicate that injuries leading to pulmonary fibrosis have a significant effect on EC-SOD distribution due to proteolytic removal of the heparin-binding domain and may be important in enhancing pulmonary injuries by altering the oxidant/antioxidant balance in alveolar interstitial spaces.


Subject(s)
Lung/enzymology , Pulmonary Fibrosis/enzymology , Superoxide Dismutase/metabolism , Animals , Antioxidants/metabolism , Bleomycin , Bronchoalveolar Lavage Fluid/chemistry , Disease Models, Animal , Extracellular Matrix/enzymology , Heparin/metabolism , Hydrolysis , Immunohistochemistry/methods , Lung/pathology , Mice , Protein Binding , Pulmonary Fibrosis/chemically induced , Pulmonary Fibrosis/pathology
5.
Am J Respir Cell Mol Biol ; 25(2): 164-9, 2001 Aug.
Article in English | MEDLINE | ID: mdl-11509325

ABSTRACT

Oxidative stress plays an important role in the development of fibrotic responses in the lung. However, it is not clear whether inhibiting oxidative stress with antioxidants can attenuate fibrotic processes in the lung. The objective of these studies was to test whether the catalytic antioxidant porphyrin manganese (III) tetrakis (4-benzoic acid) porphyrin (MnTBAP) could protect mice against bleomycin-induced lung fibrosis. A 10 mg/kg intraperitoneal dose of MnTBAP was established as safe and had a serum and lung half-life of 9.5 h in mice. Based on this data, four groups of mice were given one dose of bleomycin (3.2 U/kg, intratracheal) or saline and MnTBAP (5 mg/kg, intraperitoneal) or saline twice daily for 14 d. Lung fibrosis was assessed by measuring (1) lung hydroxyproline content as an index of collagen accumulation, (2) airway dysfunction by whole body plethysmography, and (3) histopathology. Bleomycin produced a 20% loss in body weight that was only 10% in the bleomycin/MnTBAP group. Bleomycin produced a twofold increase in hydroxyproline content that was decreased 23% by MnTBAP. Bleomycin produced a twofold increase in airway dysfunction that was also attenuated 30% by MnTBAP. Histopathologic analysis of the lungs of mice treated with bleomycin demonstrated a severe fibrotic response that was attenuated 28% by MnTBAP. Future studies on the oxidant mechanisms that MnTBAP is affecting in this bleomycin model of lung fibrosis may shed light on potential new therapeutic approaches for treating interstitial lung diseases.


Subject(s)
Antioxidants/pharmacology , Bleomycin/antagonists & inhibitors , Bleomycin/toxicity , Metalloporphyrins/pharmacology , Pulmonary Fibrosis/chemically induced , Pulmonary Fibrosis/prevention & control , Animals , Antioxidants/pharmacokinetics , Antioxidants/toxicity , Collagen/metabolism , Hydroxyproline/metabolism , Male , Metalloporphyrins/pharmacokinetics , Metalloporphyrins/toxicity , Mice , Mice, Inbred BALB C , Oxidative Stress , Pulmonary Fibrosis/metabolism , Pulmonary Fibrosis/pathology
6.
J Neurosci ; 20(20): 7631-9, 2000 Oct 15.
Article in English | MEDLINE | ID: mdl-11027223

ABSTRACT

Reactive oxygen species, including superoxide, generally are considered neurotoxic molecules whose effects can be alleviated by antioxidants. Different from this view, we show that scavenging of superoxide with an antioxidant enzyme is associated with deficits in hippocampal long-term potentiation (LTP), a putative neural substrate of memory, and hippocampal-mediated memory function. Using transgenic mice that overexpress extracellular superoxide dismutase (EC-SOD), a superoxide scavenger, we found that LTP was impaired in hippocampal area CA1 despite normal LTP in area CA3. The LTP impairment in area CA1 could be reversed by inhibition of EC-SOD. In addition, we found that EC-SOD transgenic mice exhibited impaired long-term memory of fear conditioning to contextual cues despite exhibiting normal short-term memory of the conditioning experience. These findings strongly suggest that superoxide, rather than being considered exclusively a neurotoxic molecule, should also be considered a signaling molecule necessary for normal neuronal function.


Subject(s)
Association Learning , Extracellular Space/enzymology , Long-Term Potentiation , Memory Disorders/genetics , Superoxide Dismutase/biosynthesis , Animals , Avoidance Learning , Cues , Excitatory Postsynaptic Potentials/physiology , Fear , Heterozygote , Hippocampus/cytology , Hippocampus/metabolism , Hippocampus/physiopathology , Humans , In Vitro Techniques , Long-Term Potentiation/genetics , Male , Memory Disorders/physiopathology , Mice , Mice, Transgenic , Pain Threshold , Patch-Clamp Techniques , Reactive Oxygen Species/metabolism , Signal Transduction , Spatial Behavior , Superoxide Dismutase/genetics , Synaptic Transmission/genetics
7.
Am J Physiol Lung Cell Mol Physiol ; 279(5): L977-84, 2000 Nov.
Article in English | MEDLINE | ID: mdl-11053035

ABSTRACT

Extracellular superoxide dismutase (EC-SOD), the only known enzymatic scavenger of extracellular superoxide, may modulate reactions of nitric oxide (NO) in the lungs by preventing reactions between superoxide and NO. The regulation of EC-SOD has not been examined in developing lungs. We hypothesize that EC-SOD plays a pivotal role in the response to increased oxygen tension and NO in the neonatal lung. This study characterizes rabbit EC-SOD and investigates the developmental regulation of EC-SOD activity, protein expression, and localization. Purified rabbit EC-SOD was found to have several unique biochemical attributes distinct from EC-SOD in other species. Rabbit lung EC-SOD contains predominantly uncleaved subunits that do not form disulfide-linked dimers. The lack of intersubunit disulfide bonds may contribute to the decreased heparin affinity and lower EC-SOD content in rabbit lung. EC-SOD activity in rabbit lungs is low before birth and increases soon after gestation. In addition, the enzyme is localized intracellularly in preterm and term rabbit lungs. Secretion of active EC-SOD into the extracellular compartment increases with age. The changes in EC-SOD localization and activity have implications for the neonatal pulmonary response to oxidative stress and the biological activity of NO at birth.


Subject(s)
Extracellular Space/enzymology , Lung/enzymology , Superoxide Dismutase/metabolism , Aging , Amino Acid Sequence , Animals , Animals, Newborn , Aorta/enzymology , Chromatography, Affinity , Disulfides/analysis , Embryonic and Fetal Development , Isoenzymes/chemistry , Isoenzymes/isolation & purification , Isoenzymes/metabolism , Lung/embryology , Lung/growth & development , Molecular Sequence Data , Peptide Fragments/chemistry , Protein Subunits , Rabbits , Superoxide Dismutase/chemistry , Superoxide Dismutase/isolation & purification
8.
Biochem Biophys Res Commun ; 275(2): 542-8, 2000 Aug 28.
Article in English | MEDLINE | ID: mdl-10964700

ABSTRACT

Extracellular superoxide dismutase (EC-SOD) is the major isozyme of SOD in arteries, but is also abundant in lungs. In particular, mouse lungs contain large amounts of EC-SOD compared to lungs in other mammals. This suggests that EC-SOD may have an amplified function in the mouse lung. This study describes the purification and characterization of mouse EC-SOD as well as its localization in mouse lung. Mouse EC-SOD exists primarily as a homotetramer composed of a pair of dimers linked through disulfide bonds present in the heparin-binding domains of each subunit. In addition, mouse EC-SOD can exist in active multimeric forms. We developed and utilized a polyclonal antibody to mouse EC-SOD to immunolocalize EC-SOD in mouse lung. EC-SOD labeling is strongest in the matrix of vessels, airways, and alveolar septa. This localization suggests that EC-SOD may have important functions in pulmonary biology, perhaps in the modulation of nitric oxide-dependent responses.


Subject(s)
Lung/enzymology , Superoxide Dismutase/isolation & purification , Amino Acid Sequence , Animals , Chromatography, Gel , Electrophoresis, Polyacrylamide Gel , Immunohistochemistry , Mice , Molecular Sequence Data , Superoxide Dismutase/chemistry , Superoxide Dismutase/metabolism
9.
Am J Obstet Gynecol ; 183(1): 199-205, 2000 Jul.
Article in English | MEDLINE | ID: mdl-10920331

ABSTRACT

OBJECTIVE: The aim of this study was to determine placental localization and activity of extracellular superoxide dismutase, a nitric oxide modulator, during early gestation and to correlate these characteristics with fetal vascular development. STUDY DESIGN: First-trimester (n = 10) and second-trimester (n = 10) villi were obtained at elective pregnancy termination. Extracellular superoxide dismutase was localized by means of an immunoperoxidase method. Activity was measured by determining the inhibition of cytochrome c reduction at pH 10 and messenger ribonucleic acid expression by in situ hybridization. RESULTS: Extracellular superoxide dismutase was intracellular within villous trophoblasts until 17 weeks' gestation, when it relocated to the villous extracellular matrix. Activities were similar between first- and second-trimester villi. In situ hybridization confirmed extracellular superoxide dismutase messenger ribonucleic acid within trophoblasts throughout gestation. CONCLUSION: Extracellular superoxide dismutase is produced by trophoblasts early in pregnancy, but it remains intracellular until 17 weeks' gestation, which may be related to fetal vascular development.


Subject(s)
Placenta/enzymology , Superoxide Dismutase/analysis , Blotting, Western , Cytochrome c Group/metabolism , Extracellular Space/enzymology , Female , Gestational Age , Humans , Hydrogen-Ion Concentration , Immunoenzyme Techniques , In Situ Hybridization , Keratins/analysis , Pregnancy , RNA, Messenger/analysis , Trophoblasts/enzymology
10.
J Biol Chem ; 274(21): 14818-22, 1999 May 21.
Article in English | MEDLINE | ID: mdl-10329680

ABSTRACT

Extracellular superoxide dismutase (EC-SOD) is the only known extracellular enzyme designed to scavenge the superoxide anion. The purified enzyme exists in two forms when visualized by reduced SDS-polyacrylamide gel electrophoresis: (i) intact EC-SOD (Trp1-Ala222) containing the C-terminal heparin-binding domain and (ii) cleaved EC-SOD (Trp1-Glu209) without the C-terminal heparin-binding domain. The proteolytic event(s) leading to proteolysis at Glu209-Arg210 and removal of the heparin-binding domain are not known, but may represent an important regulatory mechanism. Removal of the heparin-binding domain affects both the affinity of EC-SOD for and its distribution to the extracellular matrix, in which it is secreted. During the purification of human EC-SOD, the intact/cleaved ratio remains constant, suggesting that proteolytic removal of the heparin-binding domain does not occur during purification (Oury, T. D., Crapo, J. D., Valnickova, Z., and Enghild, J. J. (1996) Biochem. J. 317, 51-57). This was supported by the finding that fresh mouse tissue contains both intact and cleaved EC-SOD. To study other possible mechanisms leading to the formation of cleaved EC-SOD, we examined biosynthesis in cultured rat L2 epithelial-like cells using a pulse-chase protocol. The results of these studies suggest that the heparin-binding domain is removed intracellularly just prior to secretion. In addition, the intact/cleaved EC-SOD ratio appears to be tissue-dependent, implying that the intracellular processing event is regulated in a tissue-specific manner. The existence of this intracellular processing pathway may thus represent a novel regulatory pathway for affecting the distribution and effect of EC-SOD.


Subject(s)
Heparin/pharmacokinetics , Protein Splicing , Superoxide Dismutase/biosynthesis , Superoxide Dismutase/metabolism , Animals , Culture Techniques , Extracellular Space/metabolism , Humans , Mice , Rats
11.
Neuroscience ; 88(1): 185-91, 1999 Jan.
Article in English | MEDLINE | ID: mdl-10051199

ABSTRACT

Transgenic mice, which had been transfected with the human extracellular superoxide dismutase gene, causing an approximate five-fold increase in brain parenchymal extracellular superoxide dismutase activity, were used to investigate the role of extracellular superoxide dismutase in ischemic brain injury. Transgenic (n = 21) and wild-type (n = 19) mice underwent 90 min of intraluminal middle cerebral artery occlusion and 24 h of reperfusion. Severity of resultant hemiparesis and cerebral infarct size were measured. Wild-type mice had larger infarcts (cortex: wild type =37+/-14 mm3, transgenic = 27+/-13 mm3, P=0.03; subcortex: wild type = 33+/-14 mm3, transgenic = 23+/-10 mm3, P = 0.02). Neurological scores, however, were similar (P = 0.29). Other mice underwent autoradiographic determination of intra-ischemic cerebral blood flow. The volume of tissue at risk of infarction (defined as volume of tissue where blood flow was <25 ml/100g/min) was similar between groups (cortex: wild type = 51+/-15 mm3, transgenic = 47+/-9 mm3, P=0.65; subcortex: wild type = 39+/-16 mm3, transgenic= 37+/-17 mm3, P=0.81). These results indicate that antioxidant scavenging of free radicals by extracellular superoxide dismutase plays an important role in the histological response to a focal ischemic brain insult.


Subject(s)
Cerebral Infarction/prevention & control , Hemiplegia/prevention & control , Ischemic Attack, Transient/physiopathology , Superoxide Dismutase/genetics , Animals , Blood Pressure , Body Temperature , Cerebral Infarction/pathology , Cerebrovascular Circulation , Circle of Willis/abnormalities , Circle of Willis/anatomy & histology , Functional Laterality , Hematocrit , Hemiplegia/physiopathology , Humans , Immunity, Innate/genetics , Ischemic Attack, Transient/genetics , Ischemic Attack, Transient/pathology , Male , Mice , Mice, Transgenic , Risk Factors , Superoxide Dismutase/metabolism
12.
Head Neck ; 21(1): 72-7, 1999 Jan.
Article in English | MEDLINE | ID: mdl-9890354

ABSTRACT

BACKGROUND: Neuroendocrine tumors of the head and neck region may present problems in diagnosis. Middle ear carcinoid is a rare, recently recognized tumor, which to date has not been reported to metastasize. METHODS: We report the case of a 64-year-old man with a 9-year history of recurrent middle ear neoplasm and ipsilateral cervical lymphadenopathy. A microscopic parathyroid tumor was also identified. The approach to the diagnosis of this unusual combination is presented. RESULTS: The patient had a neuroendocrine tumor metastatic to multiple unilateral cervical lymph nodes, which was morphologically identical to his recurrent middle ear neoplasm. The neoplasm had the morphologic, immunohistochemical, and ultrastructural features of a carcinoid tumor. CONCLUSIONS: This case illustrates that middle ear carcinoids may metastasize. We suggest that immunohistochemical studies be performed on all biopsy specimens from neoplasms of the middle ear, as distinction from the more common paraganglioma may be difficult on morphologic grounds alone.


Subject(s)
Carcinoid Tumor/pathology , Ear Neoplasms/pathology , Ear, Middle , Neoplasm Recurrence, Local/pathology , Humans , Immunoenzyme Techniques , Male , Middle Aged
13.
Brain Res ; 850(1-2): 96-103, 1999 Dec 11.
Article in English | MEDLINE | ID: mdl-10629753

ABSTRACT

Extracellular superoxide dismutase (EC-SOD) is one of three mammalian SOD isozymes. Although there is knowledge of the functional role of EC-SOD in arteries, little is known about the function of EC-SOD in other tissues, including the brain. As a first step toward improving our understanding of EC-SOD in the brain, we studied the localization of EC-SOD in the central nervous system of the adult mouse using immunohistochemistry. We detected EC-SOD staining in a subpopulation of neurons throughout the brain as well as in tanycytes in the mediobasal hypothalamus. Particularly prominent EC-SOD staining was observed in neurons of the hilar region of the hippocampus, the lateral habenular nucleus of the thalamus, and the suprachiasmatic nuclei of the hypothalamus. Substantial numbers of neurons were distributed throughout the striatum and cortex; the morphology and distribution of these cells was similar to neurons previously shown to contain the neuronal isoform of nitric oxide synthase. In contrast to other regions with prominent EC-SOD immunoreactivity, EC-SOD localization in tanycytes occurred in a region lacking a blood-brain barrier. The high levels of EC-SOD present in discrete populations of cells in these regions suggest that EC-SOD plays an important, specialized role in the physiology and/or pathology in the brain.


Subject(s)
Brain/enzymology , Extracellular Space/enzymology , Superoxide Dismutase/metabolism , Animals , Antibody Specificity , Cerebral Cortex/cytology , Cerebral Cortex/enzymology , Diencephalon/enzymology , Hippocampus/enzymology , Immunoenzyme Techniques , Immunohistochemistry , Male , Mice , Mice, Inbred C3H , Mice, Inbred C57BL , Neostriatum/enzymology , Subcellular Fractions/enzymology
14.
Hum Pathol ; 29(12): 1382-92, 1998 Dec.
Article in English | MEDLINE | ID: mdl-9865823

ABSTRACT

The distinction of malignant mesothelioma from tumors metastatic to the serosal membranes can often be made based on the results of histochemical or immunohistochemical studies. However, in some cases, these techniques are inadequate to make a firm diagnosis. In these instances, electron microscopic studies with the observation of a constellation of characteristic ultrastructural findings may permit an unequivocal diagnosis of mesothelioma.


Subject(s)
Mesothelioma/diagnosis , Mesothelioma/ultrastructure , Microscopy, Electron , Adenocarcinoma/diagnosis , Adenocarcinoma/metabolism , Cytoplasmic Granules/metabolism , Cytoplasmic Granules/ultrastructure , Diagnosis, Differential , Humans , Hyaluronic Acid/metabolism , Immunohistochemistry , Intercellular Junctions/ultrastructure , Intermediate Filaments/ultrastructure , Mesothelioma/metabolism , Microvilli/ultrastructure , Sarcoma/diagnosis , Sarcoma/metabolism , Sarcoma/ultrastructure
15.
Mol Vis ; 4: 15, 1998 Sep 08.
Article in English | MEDLINE | ID: mdl-9736767

ABSTRACT

PURPOSE: To examine the molecular structure and ultrastructural distribution of a novel amine oxidase in human ciliary body. METHODS: Human ciliary bodies were solubilized with a nonionic detergent. The solubilized material was subjected to affinity chromatography with 2B4.14.1, a monoclonal antibody which recognizes a family of ciliary body glycoproteins. Proteins eluted from the affinity column were further separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Peptides produced from a 2B4.14. 1-reactive protein with an approximate molecular weight of 100 kDa were analyzed by Edman degradation. The protein thus identified was further examined by Western blotting and immunoelectron microscopy with anti-peptide antisera. RESULTS: Peptide sequences from the 100 kDa ciliary body protein were identical to the predicted protein sequence of an amine oxidase identified recently in a human placental cDNA library. The identity of the ciliary body protein was confirmed by Western blotting with rabbit antiserum generated against the predicted carboxy-terminal peptide of human placenta amine oxidase. Western blotting under nonreducing conditions and following glycosidase digestion indicated that the native enzyme is a disulfide-linked homodimer with multiple N-linked oligosaccharide side chains. By immunoelectron microscopy, the ciliary body amine oxidase was localized to the plasma membranes of inner epithelial cells. CONCLUSIONS: Human placenta amine oxidase is present on the plasma membranes of ciliary body inner epithelial cells. This finding provides a potential explanation for amine oxidase enzyme activity detected in previous studies of anterior segment tissues. Though the functional role of human placenta amine oxidase in the eye is unclear, it may contribute to the production of H2O2 in aqueous humor.


Subject(s)
Amine Oxidase (Copper-Containing)/chemistry , Amine Oxidase (Copper-Containing)/metabolism , Ciliary Body/enzymology , Monoamine Oxidase/chemistry , Monoamine Oxidase/metabolism , Aged , Aged, 80 and over , Amino Acid Sequence , Blotting, Western , Ciliary Body/chemistry , Copper , Electrophoresis, Polyacrylamide Gel , Epithelium/enzymology , Humans , Metalloproteins/chemistry , Metalloproteins/metabolism , Microscopy, Immunoelectron , Middle Aged
16.
Placenta ; 19(5-6): 417-22, 1998.
Article in English | MEDLINE | ID: mdl-9699963

ABSTRACT

Maintenance of low vascular tone within the placenta is mediated by nitric oxide (NO). The half-life of NO is very short, as superoxide anion (O2-) rapidly inactivates NO to form peroxynitrite. Superoxide dismutases compete with NO for O2-. By scavenging O2-, superoxide dismutase promotes the vasodilatory action of NO. Extracellular superoxide dismutase (EC-SOD) is present in high concentrations within the extracellular matrix of systemic arteries and has been proposed to mediate vascular smooth muscle tone by increasing NO bioavailability. The localization and activity of EC-SOD within the human placenta has not been determined. Placental EC-SOD may be involved in placental vascular tone, and abnormal activity may lead to pre-eclampsia secondary to increased O2--mediated inactivation of NO. To investigate this possibility, the activity and localization of human placental EC-SOD was determined in normal women, and then compared to pre-eclamptic women. Placental EC-SOD localized within the villous extracellular matrix around arterioles, and there were no differences in distribution between normal and pre-eclamptic women. There were no differences in placental EC-SOD activity between normal and pre-eclamptic subjects in either center (33.7+/-4.1 versus 33.1+/-2.5, P=0.6), or peripheral (34.3+/-5.6 versus 34.0+/-3.5, P=0.9) samples. EC-SOD localization around villous vessels suggests that EC-SOD serves potentially to protect the fetal vasculature from O2-, in both normal and pre-eclamptic pregnancies. Placental EC-SOD distribution and activity is not different between pre-eclamptic and normal women, suggesting that EC-SOD is not involved in the vascular changes seen in pre-eclampsia.


Subject(s)
Extracellular Space/enzymology , Placenta/enzymology , Superoxide Dismutase/metabolism , Adult , Arterioles/enzymology , Blotting, Western , Chorionic Villi/blood supply , Chorionic Villi/enzymology , Extracellular Matrix/enzymology , Female , Humans , Immunoenzyme Techniques , Pregnancy
17.
J Histochem Cytochem ; 46(8): 887-94, 1998 Aug.
Article in English | MEDLINE | ID: mdl-9671439

ABSTRACT

In this study we demonstrate that, in addition to blood, alpha1-microglobulin (alpha1m) is present in most tissues, including liver, heart, eye, kidney, lung, pancreas, and skeletal muscle. Western blotting of perfused and homogenized rat tissue supernatants revealed alpha1m in its free, monomeric form and in high molecular weight forms, corresponding to the complexes fibronectin-alpha1m and alpha1-inhibitor-3-alpha1m, which have previously been identified in plasma. The liver also contained a series of alpha1m isoforms with apparent molecular masses between 40 and 50 kD. These bands did not react with anti-inter-alpha-inhibitor antibodies, indicating that they do not represent the alpha1m-bikunin precursor protein. Similarly, the heart contained a 45-kD alpha1m band and the kidney a 50-kD alpha1m band. None of these alpha1m isoforms was present in plasma. Immunohistochemical analysis of human tissue demonstrated granular intracellular labeling of alpha1m in hepatocytes and in the proximal epithelial cells of the kidney. In addition, alpha1m immunoreactivity was detected in the interstitial connective tissue of heart and lung and in the adventitia of blood vessels as well as on cell surfaces of cardiocytes. alpha1m mRNA was found in the liver and pancreas by polymerase chain reaction, suggesting that the protein found in other tissues is transported via the bloodstream from the production sites in liver and pancreas. The results of this study indicate that in addition to its role in plasma, alpha1m may have important functions in the interstitium of several tissues. (J Histochem Cytochem 46:887-893, 1998)


Subject(s)
Glycoproteins/metabolism , Membrane Glycoproteins , Trypsin Inhibitor, Kunitz Soybean , Animals , Blotting, Western , Electrophoresis, Polyacrylamide Gel , Glycoproteins/blood , Humans , Immunohistochemistry , Organ Specificity , Protein Isoforms/blood , Protein Isoforms/metabolism , Rats , Rats, Sprague-Dawley , Reverse Transcriptase Polymerase Chain Reaction
18.
Behav Genet ; 28(5): 381-90, 1998 Sep.
Article in English | MEDLINE | ID: mdl-9926619

ABSTRACT

Extracellular superoxide dismutase (EC-SOD) controls the availability of extracellular superoxide (O2.-), which is important for a variety of physiological pathways, including the primary means of inactivating nitric oxide (NO). The role of EC-SOD in neurobehavioral function has been until now unexplored. In the current studies, the phenotypic expression of genotypic alterations of EC-SOD production in mice were characterized for spatial learning and memory. Dramatic impairments in spatial learning in the win-shift 8-arm radial maze were seen in both EC-SOD knockout mice and EC-SOD overexpressing mice. The EC-SOD overexpressing mice were further characterized as having significant deficits in a repeated acquisition task in the radial-arm maze, which permitted the dissociation of long and short-term learning. Long-term learning was significantly impared by EC-SOD overexpression, whereas short-term learning was not significantly affected by EC-SOD overexpression. No systems have been shown to be importantly involved in learning and memory. This may be important in the current studies because EC-SOD has primary control over the inactivation of NO. We found that EC-SOD overexpressing mice were resistant to the cognitive effects of L-NAME (NG-nitro-L-arginine methyl ester hydrochloride), an NO synthase inhibitor. Decreased NO catabolism in these mice may have served to counter the effects of NOS inhibition by L-NAME. The current finding that EC-SOD levels that were either higher or lower than controls impaired learning demonstrates that the proper control of brain extracellular O2.- may be more vital than merely reduction of brain extracellular O2.- in maintaining adequate learning function.


Subject(s)
Extracellular Space/enzymology , Genotype , Maze Learning/physiology , Superoxide Dismutase/genetics , Animals , Brain/enzymology , Female , Gene Expression Regulation, Enzymologic/physiology , Humans , Male , Mental Recall/physiology , Mice , Mice, Knockout , Mice, Transgenic , Nitric Oxide Synthase/physiology , Pregnancy , Retention, Psychology/physiology
19.
Am J Respir Cell Mol Biol ; 17(4): 393-403, 1997 Oct.
Article in English | MEDLINE | ID: mdl-9376114

ABSTRACT

Extracellular superoxide dismutase (EC-SOD) is the major extracellular antioxidant enzyme. We have determined the primary structure of mouse EC-SOD by characterization of complementary DNA (cDNA) clones and by amino-acid sequence analysis of purified protein. cDNA sequence analysis indicates that mouse EC-SOD is synthesized as a 251-amino-acid precursor protein with a predicted molecular weight of 27,400 D. Amino-terminal micro sequence analysis of purified mature mouse lung EC-SOD demonstrated the sequence to begin with SSFDLADRLDPV-. These results indicate that EC-SOD as initially synthesized contains a 24-amino-acid precursor peptide, and that the mature protein is 227 amino acids in length. Computer algorithms that predict the most likely site of cotranslational signal peptidase cleavage suggest that processing will occur between amino acids 18 and 19 or 20 and 21, which implies that EC-SOD may be initially synthesized as a pre-pro-protein. Like human EC-SOD, mature mouse EC-SOD is glycosylated. The full-length mouse EC-SOD cDNA is 1,834 base pairs long and is 82% (79% for protein) identical to rat EC-SOD, but only 60% (60% for protein) identical to human EC-SOD. The mouse EC-SOD gene locus (Sod3) was mapped by interspecific backcross haplotype analysis as being 0.9 +/- 0.9 centimorgans distal to the Qdpr locus on mouse Chromosome 5, a position suggesting that the human homologue of EC-SOD will map close to the human QDPR locus (4p15.3). Of nine tissues examined by Northern blot analysis, those of the kidney and lung are by far the major tissues that express EC-SOD messenger RNA. Using in situ hybridization in the mouse lung, we demonstrate EC-SOD gene expression to be highly localized to alveolar Type II epithelial cells. These data suggest that alveolar Type II cells play a central role in mediating EC-SOD antioxidant function in the lung.


Subject(s)
Lung/enzymology , Superoxide Dismutase , Amino Acid Sequence , Animals , Base Sequence , Chromosome Mapping , Extracellular Space/enzymology , Gene Expression Regulation, Enzymologic , Humans , In Situ Hybridization , Lung/cytology , Mice , Molecular Sequence Data , Organ Specificity , Rats , Sequence Alignment , Superoxide Dismutase/chemistry , Superoxide Dismutase/genetics , Superoxide Dismutase/metabolism
20.
Anat Pathol ; 2: 147-63, 1997.
Article in English | MEDLINE | ID: mdl-9575374

ABSTRACT

About 8% of our cases of mesothelioma occur in women, with a median age of 59 years. Our percentage is lower than other series reported in the literature because of the large number of occupationally exposed men referred to our laboratory. Tumor arose in the pleura in 86% of the women in our study, and the majority were epithelial. Pleural plaques were found in half of the women for which this information was available, and asbestosis was found in only 16%. A history of exposure to asbestos was identified in three quarters of the women, more than half of whom were household contacts of asbestos workers. Occupational exposure to asbestos was identified in only 19% of patients. An elevated tissue asbestos burden was noted in 70% of women from whom lung tissue was available for analysis. The main fiber type identified was amosite, followed by tremolite and chrysotile. These findings and those from other countries suggest a need for reassessment of the background rate of mesothelioma in industrialized nations.


Subject(s)
Asbestosis/pathology , Mesothelioma/pathology , Peritoneal Neoplasms/pathology , Peritoneum/pathology , Pleura/pathology , Pleural Neoplasms/pathology , Adult , Aged , Aged, 80 and over , Asbestos, Amosite/analysis , Asbestos, Amphibole/analysis , Asbestos, Serpentine/analysis , Female , Humans , Male , Middle Aged , Mineral Fibers/analysis
SELECTION OF CITATIONS
SEARCH DETAIL
...