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1.
Cancer Res ; 43(9): 4198-206, 1983 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-6307506

RESUMEN

Rat liver cytosol contains two protein fractions capable of serving as benzo(a)pyrene (BP) carrier in in vitro oxidation of BP (O. Hanson-Painton, M.J. Griffin, and J. Tang, Biochem. Biophys. Res. Commun., 101: 1364-1371, 1981). The role of the smaller carrier fraction in in vitro BP transport and oxidation was studied. Using Sephadex G-100 chromatography, [14C]BP bound to the carrier was shown to preferentially transfer to the microsomes. In the presence of excess BP in suspension, the carrier protein fraction transferred levels of BP many-fold in excess of its BP-binding capacity. The carrier fraction delivered BP to liposomes prepared from microsomal lipids. However, incubation of fresh liver homogenate with protein-bound BP resulted in the transfer of over 80% of the BP to the microsomes, indicating that BP is transferred selectively. Isolated microsomes containing [14C]BP oxidized bound BP upon the addition of NADPH. Sephadex G-100 chromatography and high-performance liquid chromatography analysis of bound radioactivity indicated that oxidized products of BP were preferentially transferred back to the carrier protein fraction. Thus, the carrier fraction is capable of both transferring BP to the microsomes and accepting oxidized BP from microsomes. The transfer of oxidized BP from microsomes to the carrier fraction was inhibited by an epoxide hydrolase inhibitor, alpha-bromo-4-nitroacetophenone.


Asunto(s)
Aldehído Deshidrogenasa , Benzopirenos/metabolismo , Carcinógenos/metabolismo , Proteínas Portadoras/metabolismo , Hígado/metabolismo , Microsomas Hepáticos/metabolismo , Animales , Benzo(a)pireno , Benzopirenos/farmacología , Proteínas Portadoras/aislamiento & purificación , Cromatografía Líquida de Alta Presión , Citosol/metabolismo , Epóxido Hidrolasas/antagonistas & inhibidores , Liposomas , Masculino , Peso Molecular , Oxidación-Reducción , Ratas , Ratas Endogámicas
2.
Int J Dev Biol ; 36(3): 343-51, 1992 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-1280154

RESUMEN

The highly conserved, intracellular calcium binding protein calmodulin is present in all cells at all times. In addition to this constitutive level, the amount of calmodulin is highly regulated according to the tissue or stage of development. Since there are only a few genes or a single gene for this protein in most species, intricate regulatory elements may be necessary to effect its complex regulation. This report adds new information concerning the gene structure and outlines the developmental and spatial regulation of Drosophila melanogaster calmodulin transcripts. The gene contains five exons, including a 49 bp exon in the 5' untranslated region, and spans over 16 kb. Homologues to this small, 5' noncoding exon have not been found in other calmodulin genes. The combined level of the transcripts is developmentally regulated, and the relative amounts of the two transcript size classes (1.65 kb and 1.9 kb) are differentially regulated during development. Primer extension experiments and RNase protection mapping show that both size classes of Drosophila calmodulin transcripts initiate at the same site but undergo alternative termination within the final exon. The spatial distribution of calmodulin transcripts was examined by in situ hybridization to sections of adults and to developmentally staged whole mount embryos. Calmodulin transcripts are evenly distributed early in embryogenesis. In later stages of embryogenesis, higher levels accumulate in the developing nerve cord and other tissues. Elevated levels of calmodulin transcripts are seen quite distinctly in the adult neural tissues and in the photoreceptor region of the compound eye.


Asunto(s)
Calmodulina/genética , Drosophila melanogaster/genética , ARN/análisis , Animales , Secuencia de Bases , Mapeo Cromosómico , Drosophila melanogaster/embriología , Exones , Regulación de la Expresión Génica , Hibridación in Situ , Datos de Secuencia Molecular , Poli A
3.
Neurobiol Aging ; 16(4): 563-9, 1995.
Artículo en Inglés | MEDLINE | ID: mdl-8544906

RESUMEN

Protein kinase C (PKC) is an important intracellular signalling enzyme. Numerous studies have suggested that alterations in this enzyme occur in aging and dementia. The objective of this study was to examine PKC in the cerebral microcirculation in aging and Alzheimer's disease. PKC activity, amount, and isoform distribution were analyzed in microvessels from adult and aged rodents as well as from Alzheimer patients and nondemented elderly controls. PKC activity was lower in Alzheimer vessels than in vessels from control brains, despite the presence of similar levels of PKC enzyme. In contrast, both activity and enzyme levels in young and aged rats were comparable. The beta-isoform was present in both rat and human microvessels and there were no age- or disease-related alterations. The loss in activity in cerebromicrovascular PKC in Alzheimer's suggest that perturbations in phosphorylation signalling cascades may exist at the Alzheimer blood-brain barrier.


Asunto(s)
Enfermedad de Alzheimer/enzimología , Encéfalo/irrigación sanguínea , Proteína Quinasa C/metabolismo , Anciano , Envejecimiento/metabolismo , Animales , Barrera Hematoencefálica , Estudios de Casos y Controles , Citosol/enzimología , Humanos , Immunoblotting , Isoenzimas/metabolismo , Microcirculación/enzimología , Ratas , Ratas Endogámicas F344 , Transducción de Señal , Acetato de Tetradecanoilforbol/metabolismo
4.
Biochem Pharmacol ; 34(10): 1795-800, 1985 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-4004895

RESUMEN

The influence of eleven xenobiotics on the activity and amount of hepatic microsomal epoxide hydrolase was determined. Activity was assayed using three different substrates after rats were fed, throughout 3 weeks, diets containing one of six hepatocarcinogens, viz. 2-acetylaminofluorene, 3'-methyl-4-dimethylaminoazobenzene, 4'-fluoro-4-dimethylaminoazobenzene, thioacetamide, aflatoxin B1 and ethionine. Five hepatocarcinogens induced activity 4- to 10-fold; ethionine was relatively ineffective as an inducer. Two non-carcinogenic analogues of hepatocarcinogens, viz. fluorene and p-aminoazobenzene, caused no appreciable increase in enzyme activity, but phenobarbital, barbital and 1-naphthylisothiocyanate induced activity 2- to 3-fold. All eleven xenobiotics increased the amount of microsomal epoxide hydrolase 2- to 9-fold when examined immunochemically using either a radial diffusion assay or an enzyme-linked immunosorbent assay (ELISA). Serum glutamic oxaloacetic acid transaminase activity was not appreciably elevated by feeding ten of the xenobiotics, suggesting that inductions were not owing to toxicity. Using ELISA, microsomal epoxide hydrolase was detected in post-microsomal (PM) supernatant fractions from control rat liver, thus confirming an earlier report by Gill et al. [Carcinogenesis 3, 1307 (1982)]. The eleven xenobiotics induced the amount of ELISA-detectable antigen in PM supernatant fractions by 3- to 34-fold. Longer centrifugation of PM supernatant fractions yielded a pellet fraction that contained 92 +/- 1.2% of the ELISA-detectable antigen irrespective of the xenobiotic regimen. Relationships between xenobiotic induction of microsomal epoxide hydrolase activity and amount and hepatocarcinogenesis are discussed.


Asunto(s)
Epóxido Hidrolasas/análisis , Neoplasias Hepáticas Experimentales/enzimología , Microsomas Hepáticos/enzimología , Animales , Antígenos/análisis , Aspartato Aminotransferasas/análisis , Carcinógenos , Inducción Enzimática/efectos de los fármacos , Ensayo de Inmunoadsorción Enzimática , Epóxido Hidrolasas/inmunología , Ratas
5.
Anal Biochem ; 192(1): 165-72, 1991 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-1675554

RESUMEN

A physiological assay for measuring surface accessible gamma-glutamyl transpeptidase activity in adherent, living cultures is described. Cell surface transpeptidase activity remained linear throughout a 60-min time course over a wide range of cell densities. In addition, the assay conditions have neither acute nor long-term effects on cell growth potential, cellular morphology, or cell surface transpeptidase activity levels. As a result, cell surface transpeptidase activity can be continually evaluated in the same cultures during proliferation. The assay appears to be specific for cell surface transpeptidase and can be used to study the partitioning of the enzyme between substrate-accessible and substrate-inaccessible pools. This method utilizes an automated microtiter plate reader for the spectrophotometric quantification of small aliquots removed from cultures incubated with the chromogenic substrate L-gamma-glutamyl-p-nitroanilide. The use of a microtiter plate autoreader and the minimal handling of the cells permit a large number of cultures to be assayed with a substantial reduction in the time required to measure surface transpeptidase activity. The assay described is a nondestructive means for studying cell surface-accessible gamma-glutamyl transpeptidase catalytic activity within the microenvironment of the living culture.


Asunto(s)
Membrana Celular/enzimología , gamma-Glutamiltransferasa/metabolismo , Animales , Astrocitos/enzimología , Autoanálisis , Recuento de Células , Membrana Celular/ultraestructura , Transformación Celular Neoplásica , Cinética , Ratas , Espectrofotometría , Especificidad por Sustrato , Células Tumorales Cultivadas
6.
Mol Chem Neuropathol ; 26(3): 259-68, 1995 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-8748928

RESUMEN

Regulation of protein kinase C (PKC)-mediated responses may occur by inhibition of PKC-dependent phosphorylation or by dephosphorylation of targets by specific phosphatases. Mechanisms for the regulation of PKC were examined in isolated cerebral microvessels and compared to those in brain. The data demonstrated that inhibitors of phosphorylation are responsible for the regulation in brain microvessels while dephosphorylation by protein phosphatases accounts for a substantial portion of the regulation of the PKC response in brain. In addition, the inhibitory activity apparently increases with age. These results suggest that the control of PKC may be cell-type specific and developmentally regulated.


Asunto(s)
Circulación Cerebrovascular , Proteína Quinasa C/metabolismo , Envejecimiento/metabolismo , Animales , Encéfalo/enzimología , Capilares/química , Capilares/enzimología , Masculino , Microcirculación/química , Microcirculación/enzimología , Fosfoproteínas Fosfatasas/fisiología , Proteína Quinasa C/antagonistas & inhibidores , Ratas , Ratas Endogámicas F344 , Ratas Sprague-Dawley
7.
Pharm Res ; 17(11): 1345-53, 2000 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-11205726

RESUMEN

Protein kinases and phosphatases are likely targets for the development of therapeutic drugs since they are involved in specific signaling pathways which regulate cell functions such as metabolism, cell cycle progression, cell adhesion, vascular function and angiogenesis. Protein phosphorylation and dephosphorylation serve as molecular switches for modulating these processes and the level and duration of each is a highly regulated process in normal cells. Several compounds that inhibit the activity of tyrosine kinases are being evaluated as cancer therapeutic agents in clinical trials. Diabetes and complications of diabetes also involve deregulated levels of protein kinases. New approaches for regulating kinase gene expression include specific antisense oligonucleotides for inhibiting post-transcriptional processing of the messenger RNA, naturally occurring products and their chemical derivatives to inhibit kinase activity, and monoclonal antibodies to inhibit receptor linked kinases. Inhibition of phosphatases also serves to alter the duration of phosphorylation by kinases. Considerations for development of effective inhibitors include non-specific actions of compounds, cellular uptake, multiple intracellular targets that can dilute the effective cellular concentration of an agent, and tissue specificity. Kinase inhibitors may allow other therapeutic agents additional time to become effective and they may act synergistically with current treatments.


Asunto(s)
Inhibidores Enzimáticos/farmacología , Inhibidores de Proteínas Quinasas , Animales , Antineoplásicos/farmacología , Angiopatías Diabéticas/tratamiento farmacológico , Humanos , Proteínas Quinasas/metabolismo , Transducción de Señal/efectos de los fármacos
8.
J Cell Physiol ; 150(1): 104-15, 1992 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-1346140

RESUMEN

A decline in cell surface gamma-glutamyl transpeptidase specific activity was previously observed to be concomitant with C6 glial cell proliferation. To elucidate the underlying factor(s) mediating gamma-glutamyl transpeptidase down-regulation, the effects of C6 cell density and culture conditions on cell surface transpeptidase activity levels were investigated. After 24 h of culture, the transpeptidase specific activities were inversely related to the initial plating densities. The lower-density cultures showed an induction within 24 h of plating. As the cultures proliferated, the specific transpeptidase activities declined to a common low level at post-confluency. The gamma-glutamyl transpeptidase down-regulation was unrelated to cell growth rate and was most pronounced during logarithmic proliferation. Induction and down-regulation of gamma-glutamyl transpeptidase activity at low cell densities were not a result of trypsinization. Supplementation of low-density cultures with conditioned medium, use of matrix-coated wells, or periodic replacement of growth media to prevent conditioning had minor effects on the decline of cell surface activity. Kinetic analysis showed that the Michaelis constants and the reaction mechanism were unaltered by cell density, indicating that down-regulation was not due to allosteric factors or an alteration in enzyme character. A reduction in the maximal velocity of cell surface transpeptidation at higher cell densities suggested that gamma-glutamyl transpeptidase down-regulation is related to the concentration of enzyme at the cell surface. Immunocytochemical localization of gamma-glutamyl transpeptidase demonstrated that gamma-glutamyl transpeptidase antigen levels decrease as C6 cell density increases. These results led us to propose that cell-cell contact stimulates the disappearance of gamma-glutamyl transpeptidase from the surface of cultured C6 glial cells.


Asunto(s)
Astrocitos/enzimología , Membrana Celular/enzimología , gamma-Glutamiltransferasa/metabolismo , Animales , Astrocitos/citología , Astrocitoma , Comunicación Celular , Recuento de Células , División Celular , Medios de Cultivo , Regulación hacia Abajo , Inmunohistoquímica , Cinética , Ratas , Tripsina , Células Tumorales Cultivadas
9.
Kidney Int ; 44(2): 322-30, 1993 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-8377376

RESUMEN

Calbindin-D28k appears in the metanephric kidney during embryogenesis. We studied the temporal appearance and spatial distribution of calbindin-D28k mRNA in the developing kidneys of 12-day fetal through 21-day postnatal mice by in situ hybridization. 35S-UTP-labeled antisense (cRNA) probe to calbindin-D28k mRNA hybridized to the ureteric buds of 12-day embryos, whereas adjacent metanephrogenic tissue was unlabeled. By embryonic day 13, Y-shaped bodies of "advancing" ureteric buds were labeled intensely. In 16-day embryos, ampullae of ureteric buds were located immediately beneath the renal capsule and labeled strongly, in contrast to metanephric tubules and S-shaped bodies. The former were unlabeled and the latter were labeled only at points of contact with the ampullae. Subsequently, the ampullae of the metanephric ureteric buds hybridized with the cRNA probe, and from the 18th embryonic to the 21st postnatal day, this labeling was intense. The cRNA probe did not hybridize with the renal vesicles, proximal tubules, or tubular segments of Henle's loop derived from nephrogenic blastema, but it did label distal nephron segments. By the 21st postnatal day, collecting ducts and ureter no longer were labeled. In conclusion, calbindin-D28k mRNA is present in the developing mouse kidney, and its distribution during nephrogenesis is identical to that of calbindin-D28k per se. Collectively, these findings show that the calbindin-D28k gene is transcribed and its message is translated by the cells of the ureteric bud during the initial stage of renal morphogenesis.


Asunto(s)
Feto/metabolismo , Riñón/embriología , Riñón/metabolismo , Proteína G de Unión al Calcio S100/genética , Animales , Animales Recién Nacidos , Calbindina 1 , Calbindinas , Desarrollo Embrionario y Fetal , Femenino , Feto/citología , Riñón/crecimiento & desarrollo , Masculino , Ratones , Ratones Endogámicos , Proteína G de Unión al Calcio S100/química , Distribución Tisular
10.
Mol Chem Neuropathol ; 20(3): 245-61, 1993 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-8172627

RESUMEN

Activation of protein kinase C is a key event in the transduction of receptor-mediated extracellular signals. Little is known about the role of protein kinase C in the microcirculation of the brain. In this study, we examined protein kinase C in isolated cerebral microvessels. A technique for partial purification of protein kinase C from microvessels was employed, using Q-Sepharose batch adsorption and single-step salt elution in microfuge tubes. This procedure greatly reduced variability and increased protein kinase C specific activity in both the cytosolic and particulate fractions by nearly 50-fold. The identity of the enzyme was confirmed by its inhibition by staurosporine and bisindolylmaleimide and by its translocation in response to phorbol ester. The level of protein kinase C was assessed by [3H]phorbol ester binding and the endogenous substrates evaluated by in vitro phosphorylation studies. Finally, western blot analysis of protein kinase C isoforms indicated that the beta-isoform was present in both cytosolic and particulate fractions. The alpha-isoform was present at low levels in the cytosolic fraction, whereas the gamma-isoform was not detected.


Asunto(s)
Encéfalo/enzimología , Isoenzimas/metabolismo , Proteína Quinasa C/metabolismo , Secuencia de Aminoácidos , Animales , Western Blotting , Calcio/metabolismo , Capilares/enzimología , Citosol/enzimología , Técnicas In Vitro , Isoenzimas/antagonistas & inhibidores , Isoenzimas/aislamiento & purificación , Masculino , Datos de Secuencia Molecular , Forbol 12,13-Dibutirato/metabolismo , Fosfolípidos/metabolismo , Fosfoproteínas/metabolismo , Fosforilación , Proteína Quinasa C/antagonistas & inhibidores , Proteína Quinasa C/aislamiento & purificación , Ratas , Ratas Sprague-Dawley
11.
J Immunol ; 148(2): 457-65, 1992 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-1309557

RESUMEN

To test the hypothesis that mesothelial cells play a role in regulating inflammatory responses within the pleural space, we examined neutrophil chemotactic activity released by cytokine-stimulated mesothelial cells. Human mesothelial cells were isolated from patients with transudative pleural effusions and cultured. The purity of the cell population was assessed by morphologic, immunocytochemical, and biochemical characteristics. Confluent fourth passage mesothelial cell plates were exposed to varying concentrations of the recombinant human cytokines IL-1 alpha, TNF-alpha, or IFN-gamma, or Escherichia coli endotoxin (LPS). Polymorphonuclear neutrophil (PMN) chemotactic activity in the conditioned media was measured in microchemotaxis chambers. Although none of the cytokines demonstrated inherent chemotactic activity, each stimulated mesothelial cells to produce PMN chemotactic activity in a dose-dependent manner. TNF-alpha stimulated the release of the greatest quantity, whereas stimulation with IFN-gamma and IL-1 alpha resulted in the release of lesser but still significant quantities of PMN chemotactic activity. By contrast, LPS did not increase the basal level of chemotactic activity produced by the cells. The cytokine-induced chemotactic activity was proteinaceous, required de novo synthesis, and had a predominant m.w. of 10,000. Significant quantities of immunoreactive neutrophil-activating peptide-1 (NAP-1)/IL-8 were detected in mesothelial cell supernatants after stimulation with each of the cytokines. The neutrophil chemotactic activity of supernatants from mesothelial cells stimulated with either IL-1 alpha or IFN-gamma was completely neutralized with rabbit anti-human NAP-1/IL-8 polyclonal antiserum. The same antiserum neutralized the majority, but not all, of the neutrophil chemotactic activity in supernatants from TNF-stimulated mesothelial cells. Stimulated mesothelial cells also expressed an inducible mRNA transcript that hybridized with a specific oligonucleotide probe for human NAP-1/IL-8. These observations provide a mechanism whereby mesothelial cells could respond to inflammatory stimuli in the underlying lung and regulate inflammatory responses within the pleural space.


Asunto(s)
Quimiotaxis de Leucocito/fisiología , Citocinas/farmacología , Interleucina-8/biosíntesis , Pleura/metabolismo , Secuencia de Bases , Separación Celular , Complemento C5a/biosíntesis , Epitelio/metabolismo , Humanos , Interferón gamma/farmacología , Interleucina-1/farmacología , Interleucina-8/genética , Interleucina-8/inmunología , Leucotrieno B4/biosíntesis , Lipopolisacáridos , Datos de Secuencia Molecular , Peso Molecular , Pleura/efectos de los fármacos , ARN Mensajero/análisis , Factor de Necrosis Tumoral alfa/farmacología
12.
Acta Obstet Gynecol Scand ; 73(5): 429-31, 1994 May.
Artículo en Inglés | MEDLINE | ID: mdl-8009978

RESUMEN

We report the case of a 50 year old woman with metastatic breast carcinoma refractory to chemotherapy who died of candidal septicemia after autologous bone marrow transplantation. Although there was no apparent active cytomegalovirus (CMV) infection (negative cultures and serology for active infection), autopsy revealed histologic evidence of CMV inclusions limited to both ovaries. DNA in situ hybridization was performed on multiple organs, and additional foci of infection in one fallopian tube and the adrenal glands were detected. Previous reports of isolated CMV oophoritis may represent sampling error. An ascending route of infection is suggested. Tubo-ovarian changes due to CMV infection may occur more frequently than suspected; they are difficult to diagnose because even actively CMV infected cells may not be detected by routine histology alone, and because, after the active infection 'heals', no evidence of the virus can be found on histologic examination.


Asunto(s)
Enfermedades de las Glándulas Suprarrenales/patología , Trasplante de Médula Ósea/efectos adversos , Neoplasias de la Mama/complicaciones , Candidiasis/complicaciones , Infecciones por Citomegalovirus/patología , ADN Viral , Enfermedades de las Trompas Uterinas/patología , Fungemia/complicaciones , Hibridación in Situ/métodos , Enfermedades del Ovario/patología , Enfermedades de las Glándulas Suprarrenales/complicaciones , Neoplasias de la Mama/terapia , Infecciones por Citomegalovirus/complicaciones , Enfermedades de las Trompas Uterinas/complicaciones , Femenino , Humanos , Persona de Mediana Edad , Enfermedades del Ovario/complicaciones
13.
Nucleic Acids Res ; 15(8): 3335-48, 1987 Apr 24.
Artículo en Inglés | MEDLINE | ID: mdl-3106931

RESUMEN

We have isolated and characterized cDNA and genomic clones representing the calmodulin gene of Drosophila melanogaster. As demonstrated by genomic blots and by reconstruction experiments, the calmodulin gene is represented once in the Drosophila genome. In situ hybridization of cloned probes to the polytene chromosomes of third instar larvae permitted the localization of the gene to region 49A on the left arm of the second chromosome. Two transcripts of 1.65 and 1.9 kb are produced from this gene. The accumulation of calmodulin message was measured at several stages of Drosophila development. The results of these experiments suggest developmental regulation of the gene. Three intervening sequences interrupt the protein coding nucleotides and two of these are located within calmodulin functional domains. The DNA sequence encoding the protein is presented; the derived amino acid sequence is compared to that of other species. The structural similarities of the Drosophila calmodulin gene to calmodulin genes of other species and to other calcium binding protein genes are discussed.


Asunto(s)
Calmodulina/genética , Drosophila melanogaster/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Mapeo Cromosómico , ADN/análisis , ADN Recombinante/análisis , Regulación de la Expresión Génica , Genes , Intrones , ARN Mensajero/análisis
14.
Mod Pathol ; 5(3): 277-82, 1992 May.
Artículo en Inglés | MEDLINE | ID: mdl-1323103

RESUMEN

Although it has been suggested that cytomegalovirus (CMV) infection of the kidney might facilitate the development of human immunodeficiency virus-associated nephropathy (HIVAN) or other morphologic renal changes in patients with AIDS, no systematic study has been performed on kidneys from AIDS patients. We examined 75 autopsy kidneys, two renal biopsy specimens, and a nephrectomy specimen from 78 HIV-infected patients (five with HIVAN) for the presence of CMV. Immunocytochemistry (ICC) utilizing a monoclonal antibody against the late antigen of CMV and in situ hybridization (ISH) with a biotinylated DNA probe for CMV sequences were used. The detection system for both ICC and ISH was streptavidin-conjugated alkaline phosphatase with Fast Red TR chromogen. CMV was detected in only 10 of the 78 kidneys examined (12.8%): eight by both methods, one by ISH only, and another by ICC only. All 10 positive kidneys were obtained from autopsies of patients with AIDS. The average number of positive cells (in approximately 15 x 10 mm sections) was 22 with ICC and 10 with ISH. Glomerular intracapillary cells (possibly endothelial cells) were the most commonly stained, followed by positive cells in the interstitium and peritubular capillaries. Relatively few tubular epithelial cells were stained. The majority of positive cells by either ICC or ISH did not show nuclear or cytoplasmic inclusions; however, only two of the 10 positive kidneys did not contain cells with typical Cowdry type-A intranuclear CMV inclusions. The most frequent pathologic finding in the kidneys positive for CMV by either ICC or ISH was acute tubular necrosis (in six of 10, 60%).(ABSTRACT TRUNCATED AT 250 WORDS)


Asunto(s)
Nefropatía Asociada a SIDA/microbiología , Síndrome de Inmunodeficiencia Adquirida/complicaciones , Infecciones por Citomegalovirus/microbiología , Citomegalovirus/aislamiento & purificación , Nefropatía Asociada a SIDA/patología , Infecciones por Citomegalovirus/patología , Sondas de ADN , Humanos , Inmunohistoquímica , Hibridación de Ácido Nucleico
15.
Mod Pathol ; 5(3): 283-91, 1992 May.
Artículo en Inglés | MEDLINE | ID: mdl-1379713

RESUMEN

A number of studies have suggested that HIV infection can be detected in a variety of routinely fixed archival tissues using antibodies to various viral proteins. In order to study this immunocytochemical approach, paraffin sections were examined with a large panel of commercially available monoclonal antibodies against the various HIV proteins (5 antibodies to p24, 1 to p17, 1 to gp41, and 1 to gp120) using a streptavidin-biotin method. A polyclonal antibody against p24 was also tested. Formalin-fixed, paraffin-embedded HIV infected CEM E5 T cells were used as positive controls. Tissues from AIDS patients included 31 kidneys, 8 lymph nodes, 2 spleens and 3 brains. Non-AIDS tissues examined were 6 renal biopsies with focal segmental glomerulosclerosis, 5 with interstitial nephritis, 6 reactive lymph nodes, and a brain with encephalitis, all from patients not known to be at high risk for HIV infection. Additional negative controls included: 1) replacement of primary antibody with a hybridoma derived mouse monoclonal IgG1 standard, 2) omission of the primary antibody, and 3) sections of formalin-fixed paraffin-embedded CEM E5 T cells cultures not infected with HIV. Competition experiments with excess recombinant p24 protein were also performed. False positive staining with the IgG1 standard or with the antibodies to HIV proteins was frequently seen in tissues with pathologic findings (inflammation, hyalin degeneration), particularly following protein digestion. Protein digestion also had a major impact on specific staining. Digestion with proteinase K abolished specific staining for the core proteins of the virus (p17, p24) on the positive control sections.(ABSTRACT TRUNCATED AT 250 WORDS)


Asunto(s)
Nefropatía Asociada a SIDA/microbiología , Proteína p24 del Núcleo del VIH/inmunología , VIH/aislamiento & purificación , Riñón/microbiología , Adhesión del Tejido , Fijación del Tejido , Anciano , Anticuerpos Monoclonales , Niño , Preescolar , Humanos , Inmunohistoquímica/métodos , Riñón/patología , Persona de Mediana Edad , Coloración y Etiquetado
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