Histology and histopathology Vol.16, nº 3 (2001)

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Open Access
A sex-related difference in the hypertrophic versus hyperplastic response of vascular smooth muscle cells to repeated passaging in culture
(Murcia : F. Hernández, 2001) Bkákovál, L.; Pellicciari, C.; Bottone, M.G.; Lisá, V.; Mares, V.
Activation of growth of vascular smooth muscle cells (VSMC) in adults participates in pathogenesis of dysplastic diseases of the vascular system. In this study, we examined the impact of gender of rat donors on the degree of hyperplastic and hypertrophic responses of VSMC in cultures subjected to repeated passaging. The cells were derived from the outgrowth zone of explants of the thoracic aorta and were studied up to passage 45. Under these conditions, the cells undergo repeated growth stimulation by the serum growth factors mimicking some pathological situations in vivo. At lower passages (5-7), the cells from both sex donors did not differ significantly in their doubling time, maximum population density, protein content and ploidy. At higher passages (40-45), we found that the hyperplastic response, monitored by doubling time and BrdU-revealed DNA synthesis, was more intense in VSMC of male origin. In contrast, female-derived cells reacted by more prominent hypertrophic changes. The latter included a relatively higher increase in the volume and protein content of cells. As indicated by the DNA content histograms and chromosome numbers, these cells also showed a higher degree of passage-dependent polyploidization. In addition, the female-derived VSMC were found to be more effective in adhesion to the growth support evidenced by wider spreading and higher resistance of these cells to trypsin-mediated detachment as well as higher expression of some integrin and cytoskeletal molecules. These features could partly account for the slower proliferation and polyploidization of these cells. The results suggest that rat VSMC populations of male and female origin contain cells which are intrinsically different with respect to their capability of reacting to growth stimuli. The lower responsiveness of femalederived cells to growth stimuli may contribute to less frequent formation of hyperplastic vascular lesions in female organisms.
Open Access
Role of nitric oxide in murine cytomegalovirus (MCMV) infection
(Murcia : F. Hernández, 2001) Tanaka, K.; Noda, S.
Cytomegalovirus (CMV) is a typical pathogen of an opportunistic infection. In this review article, various roles of nitric oxide (NO) in murine CMV (MCMV) infections, including acute, persistent and latent infections, are discussed. In the acute phase of MCMV infection, NO plays a protective role against MCMV infection. In contrast, NO has been proven to act as a pathogenic factor in a model of MCMV pneumonitis. In MCMV persistent infection, when MCMV was detected only in the salivary gland, T cells of mice were modified to produce a massive amount of such cytokines as TNF-a and IFN-y upon in vivo stimulation with anti-CD3. These cytokines then induced mRNA for inducible NO synthase (iNOS), thus resulting in the production of a large amount of NO. A histochemical study demonstrated that NO damaged bronchial epithelial cells, and thereby apparently inducing pneumonitis. In the case of a latent infection, when viral DNA was detected in the host in spite of the absence of any infectious particle, NO increased the amount of persistently-infected MCMV-DNA. As a result, NO was found to act as "a double edged sword" in the CMV-host relationship.
Open Access
Heat-induced antigen retrieval of epoxy sections for electron microscopy
(Murcia : F. Hernández, 2001) Brorson, S.H.
The purpose of this manuscript is to review the literature on the use of heat-induced antigen retrieval methods to enhance the immunolabeling of epoxy sections at the electron microscopical level. The history of the development of antigen retrieval by heating formaldehyde fixed paraffin sections in a buffer solution is given in short, and how this technique has been extended to resin sections and in particular epoxy sections is explained. Theories for the mechanism of enhancement of the immunolabeling of epoxy sections by the heat-retrieval method are discussed, and it is finally speculated whether most of the mechanisms for antigen retrieval on epoxy sections in heated buffer solution are essentially the same as for conventional immunoenhancing by deplastizing and etching. The more accelerator used in the processing of the tissue the more intense the immunolabeling of the heated epoxy sections becomes. The intensity of immunolabeling of the epoxy sections increases with the temperature in the heated buffer solution, and the intensity is significantly higher at high autoclave temperatures than at 95 OC, Heat-induced antigen retrieval is also compared with other, conventional techniques for enhancing the immunolabeling of epoxy sections.
Open Access
Scanning electron microscopy and calcification in amelogenesis imperfecta in anterior and posterior human teeth
(Murcia : F. Hernández, 2001) Sánchez-Quevedo, M. C.; Ceballos, G.; García, J. M.; Rodriguez, I. A.; Gómez de Ferraris, M. E.; Campos, Antonio
Teeth fragments from members of a famil? clinically and genetically diagnosed as having amelogenesis imperfecta were studied by scanning electron microscopy and X-ray microprobe analysis to establish the morphological patterns and the quantitative concentration of calcium in the enamel of anterior (canine, incisor) and posterior (premolar and molar) teeth. The prism patterns in the enamel of teeth from both regions were parallel or irregularly decussate, with occasional filamentous prisms accompanied by small, irregularly rounded formations. Prismless enamel showed the R- and P-type patterns. Calcium levels in enamel of amelogenesis imperfecta and control teeth differed significantly between anterior and posterior teeth, indicating that the factors that influence normal mineralization in different regions of the dental arch are not altered in the process of arnelogenesis imperfecta.
Open Access
Development and progression of malignancy in human colon tissues are correlated with expression of specific ca2+-binding S100 proteins
(Murcia : F. Hernández, 2001) Bronckart, Y.; Decaestecker, C.; Nagy, N.; Harper, L.; Schafer, B.W.; Salmon, I.; Pochet, R.; Kiss, R.; Heizman, C.W.
The expression levels of seven different S100 proteins (S100A1, S100A2, S100A3, S100A4, S100A5, S100A6, and S100B) were characterized by immunohistochemistry in the epithelial versus connective tissues of a series of 35 colon specimens, including 6 normal samples, 5 adenomas with low-grade dysplasia, 5 adenomas with high-grade dysplasia, and 19 cancers. The results showed that S100A2, S100A3, and SlOOB proteins could not (or only marginally) be detected in colon tissues. On the other hand, the expression of S100A6 increased in epithelial tissues directly proportional to the increase of malignancy. The percentage of epithelial (or connective tissue) cells expressing S100A4 significantly decreased as the malignancy grade increased. The expression level of SlOOAl proteins was somewhat higher in the connective tissues of normal cases and adenomas with low-grade dysplasia than in adenomas with high-grade dysplasia and cancers. This pattern of expression was not observed in epithelial tissues. While the node-positive cancers did not express S100A1, about half of the node-negative specimens did. The expression levels of S100A5 were similar in different epithelial tissues. However, in the connective tissues the expression levels decreased inversely proportional to the increase in pathological grading of the specimens. Therefore, the present study implicates several S100 proteins as useful tools for histochemical typing of colon cancer malignancy development.
Open Access
Opiomelanins synthesis and properties
(Murcia : F. Hernández, 2001) Rosei, M.A.
Opiomelanins represent a new class of synthetic pigments produced by the tyrosinase-catalyzed oxidation of opioid peptides and other tyrosine aminoterminal peptides. In contrast with dopamelanin, these polymers are fully soluble in hydrophilic media, due to the presence of the peptide moiety. Opiomelanins show paramagnetism as demonstrated by the EPR spectrum identical to that of dopamelanin. The presence of the aminoacidic chain linked to the melaninic moiety, influences the electron transfer properties of opiomelanins i.e. the ability to oxidize NADH. Like dopamelanin Tyr-Gly-melanin exhibits this behaviour whereas leuenkmelanin does not show any oxidizing potential. Opiomelanins UV-Vis spectrum is characterized by an absorption band at 330 nm which disappears upon acid hydrolysis, H202 treatment and under simulated solar illumination. Opiomelanins exhibit a fluorescence emission peaked at 440 and 520 nm if excited at 330 nm. These fluorescence bands are due to the oligomeric units and high molecular weight units, respectively. When opioid peptides are allowed to react with tyrosinase in the presence of an excess of cysteine, cysteinyldopaenkephalins are synthesized. These peptides are furtherly oxidized giving rise to pheoopiomelanins. Reactive oxygen species also are able to oxidize non enzymatically both enkephalins and cysteinyldopaenkephalins, producing the corresponding melanin pigments .
Open Access
Potential role of a new anti-03 integrin antibody in the development of intimal hyperplasia after vascular surgery: an in vitro smooth muscle cell model
(Murcia : F. Hernández, 2001) Gimeno, M.J.; González, J.; Rodríguez, M.; Corrales, C.; Bellón, J.M.; Buján, J.
The occurrence of intimal hyperplasia after vascular surgery is an ongoing concern in current clinical practice. Arnong the many factors involved in the development of this pathology, platelet adhesion and myointimal proliferation play a major role. Both these processes are mediated by integrins (mainly avP3 integrins). Over the past years, severa1 substances have been designed to delay or inhibit the cell proliferation that leads to hyperplasia and mainly include monoclonal antibodies directed against integrins. The aim of the present study was to evaluate the effects of an antibody denoted P37 (anti B3 integrin) on human smooth muscle cells (SMC) and its role in blocking the 83 subunit. To this end, SMC from human umbilical artery were cultured in the presence or absence of the cell substrate vitronectin (VN) and incubated with P37. After 4 days of treatment, determination was made of cell proliferation and migration. Smooth muscle cells grown on VN showed increased proliferation and migration compared to control VN-free cultures. However, the presence of P37 in the culture medium inhibited proliferation and reduced migration. Combined treatment with VN and P37 led to improved proliferation but VN was unable to reverse the effects on migration observed in the former cultures. Results suggest that in vitro, P37 is capable of blocking human SMC 83 integrins and thus impedes cell proliferation and migration These findings may have clinical implications related to modulation of the development of hyperplasia.
Open Access
Effects of orexins A and B on the secretory and proliferative activity of immature and regenerating rat adrenal glands
(Murcia : F. Hernández, 2001) Malendowicz, L.K.; Jedrzejczak, N.; Belloni, A. S.; Tretjer, M.; Hochol, A.; Nussdorfer, G.G.
Orexins A and B are two hypothalamic peptides, involved in the central regulation of feeding, which act through two receptor subtypes, named OXIR and OX2R. OXIR is selective for orexin-A, and OX2R binds both orexins. We have investigated the effects of three subcutaneous injections of 10 nmollkg body weight of orexins on the secretion and proliferative activity of immature (20-day-old) and regenerating rat adrenal cortex. The presence of both OXIR and OX2R mRNAs has been detected by reverse transcriptionpolymerase chain reaction in adult, immature and regenerating adrenals. Orexin-A increased corticosterone plasma concentration in immature rats, but not in animals with regenerating adrenals. Both orexins raised metaphase index (%o of metaphase-arrested cells) in immature rat adrenals, orexin-B being more effective than orexin-A. In contrast, both orexins equipotently lowered adrenal metaphase index at day 5 (but not day 8) of adrenal regeneration. We conclude that orexins (1) stimulate secretion and proliferative activity of immature rat adrenals, acting through OXIR and OX2R, respectively; and (2) do not affect secretion, but inhibit proliferative activity of regenerating adrenals, mainly via the activation of OX2R.
Open Access
Colonic endocrine cells in rats with chemically induced colon carcinoma
(Murcia : F. Hernández, 2001) Sitohy, B.; El-Salhy, M.
Colonic carcinoma was induced in male Sprague-Dawley rats by injecting them with 1,2- dimethylhydrazine dihydrochloride. Control rats were injected with EDTA solution. Tissue specimens of colon from four groups of animals: (i) rats without tumour, (ii) with dysplasia and lymphoid hyperplasia, (iii) with colonic adenocarcinoma, and (iv) controls, were investigated. The colonic endocrine cells were detected by immunocytochemistry and quantified by computerised image analysis. Peptide YY (PYY)- and serotonin-immunoreactive cells were found in the colon of al1 the groups investigated. There were few somatostatin- or enteroglucagon-immunoreactive cells and no pancreatic polypeptide (PP)-immunoreactive cells in the colon of any of the groups studied. The density of PYY-immunoreactive cells increased significantly in rats with dysplasia and lymphoid hyperplasia and in rats with colon carcinoma. There was no statistically significant difference as regards cell secretory index (CSI) or nuclear area of PYYimmunoreactive cells in any of treated groups examined. Nor was there any statistically significant difference between al1 treated animal groups and controls, as regards cell density, CSI, or nuclear area of serotoninimmunoreactive cells. The present observations in an animal model of human colon carcinoma support the assumption that neuroendocrine peptides in the gut are involved in the carcinogenesis of colorectal carcinoma. However, The nature of the changes in the colonic endocrine cells observed here differed from those in patients with colon carcinoma, possibly due to a difference between the response of young rats to an induced colon carcinoma and a spontaneously developed carcinoma in elderly humans, or due to a species difference.
Open Access
Ultrastructural observations on the microvasculature in advanced gastric carcinomas
(Murcia : F. Hernández, 2001) Caruso, R.A.; Speciale, G.; Inferrera, A.; Rigolis, L.; Inferrera, C.
The ultrastructural features associated with vascular permeability in 9 cases of advanced gastric carcinomas were studied, and compared with that of control non-neoplastic mucosa. Tumour rnicrovasculature showed features in common with those of control mucosa, including complete basa1 lamina, welldeveloped interendothelial junctions, fenestrations and caveolae. Some tumour blood vessels showed endothelial cell swelling accompained by luminal narrowing and perivascular fibrosis. In 2 out of 9 cases, there were endothelial attenuation with numerous fenestrations and vesiculo-vacuolar organelles. The vesiculo-vacuolar organelle is a recently described cytoplasmic structure found in the endothelial cells lining turnour microvessels and normal venules and which provides an important pathway for extravasation of circulating macromolecules. Our ultrastructural data suggest that advanced gastric carcinomas share with experimental tumour models in vivo only some morphologic features associated with hyperpermeability including fenestration, endothelial attenuation and vesiculo-vacuolar organelles. The implications of perivascular fibrosis on the delivery of immune cells to gastric carcinomas are discussed.
Open Access
Neurotransmitters, neuromodulators, and neurotrophin receptors in the gut of pantex, a hybrid sparid fish (Pagrus major x Dentex dentex). Localizations in the enteric nervous and endocrine systems
(Murcia : F. Hernández, 2001) Radaelli, G.; Domeneghini, C.; Arrighi, S.; Castaldo, L.; Lucini, C.
The gut of Pantex, a sparid hybrid fish (Pagrus major x Dentex dentex) with a great potential importance for the Italian aquaculture, was histochemically and immunohistochemically investigated in order to evidence components of the intramural nervous and diffuse endocrine systems. The general structural aspects of the intramural nervous system were shown by the Nissl-thionin staining. As in most other fish, it was only organized in the myenteric plexus. Acetylcholinesterase (AChE) activity was observed in both nerve cell bodies and terminals al1 along the gut. The NADPH-diaphorase reactivity too, possibly linked to the synthesis and release of nitric oxide, was present in nerve cell bodies and nerve terminals of the oesophagus, stomach and intestine. In addition, the intramural nervous system was shown to contain Trk (tyrosinekinase) receptors for neurotrophin, as evidenced by Trk A-, Trk B- and Trk C-like immunoreactivities, thus suggesting an involvement of neurotrophin in the function of this system. Trk B- and Trk C-like immunoreactivities were detected in epithelial endocrine cells, too. The additional presence of serotonin- and metenkephalin- like immunoreactivities in numerous endocrine cells in the epithelial layers of the stomach and intestine was showed.
Open Access
Acute toxicity of anionic surfactants sodium dodecyl sulphate (SDS) and linear alkylbenzene sulphonate (LAS) on the fertilizing
(Murcia : F. Hernández, 2001) Rosety, M.A.; Ordóñez Muñoz, F.J.; Rosety-Rodriguez, M.; Rosety, J.M.; Rosety, I.; Carrasco, C.; Ribelles, A.
In the present work we have evaluated and compared the acute toxicity of two anionic surfactants, Sodium Dodecyl Sulphate (SDS) and Linear Alkylbenzene Sulphonate (LAS) on the fertilizing capability of gilthead Sparus aurata L. sperm. The criterion used to judge exposure effectiveness was fertilization success. Spawned eggs and sperms were collected from adult giltheads. Sperms were dosed separately with different concentrations of SDS and LAS for 60 minutes. After this period, sperms and eggs were combined for 20 minutes during which fertilization took place. Finally, the number of fertilized eggs were counted and recorded to estimate the percentage of fertilization. Exposure to SDS and LAS concentrations of 0.3, 0.6, 1.5, 3 and 6 mg/L for 60 minutes caused a significant inhibitory effect on fertilization success in gilthead Sparus aurata L.. In addition, the EC50 value for gilthead fertilization after sperm exposure was found to be 2.8 mg/L and in the case of LAS it was of 2.6 The comparison of the results from SDS and LAS shows that the latter has a stronger negative effect on sperm viability than SDS.
Open Access
Caspase inhibition, a potential therapeutic strategy in neurological diseases
(Murcia : F. Hernández, 2001) Rideout, H.J.; Stefanis, L.
Caspases are intracellular proteases that participate in apoptotic pathways in mammalian cells, including neurons. Here we review evidence that caspase inhibition, through pharmacological or molecular means, rnay inhibit neuronal cell death in a number of in vitro and in vivo models of neurological disease. It has recently become clear that, at least in most cell culture models, caspase inhibition offers only transient protection, and that a caspase-independent death eventually occurs. This rnay be due to irreversible caspase-independent alterations at the leve1 of the mitochondria. Despite concerns that targeting caspases alone rnay prove insufficient to truly reverse the effects of various death stimuli, in vivo studies indicate that caspase inhibition promotes survival and functional outcome in a variety of neurological disease models. In addition, studies of human post-mortem material suggest that caspases are activated in certain human neurological diseases. Caspase inhibition rnay therefore provide a novel strategy for the treatment of such disorders. Caspas'es, through the generation of toxic fragments of critica1 protein substrates, rnay also be involved in earlier steps of neuronal dysfunction, such as protein aggregation in Huntington's and Alzheimer's disease, and therefore caspase inhibition rnay be of additional value in the treatment of these particular disorders.
Open Access
Gene therapy strategies in neurodegenerative diseases
(Murcia : F. Hernández, 2001) Giménez y Ribotta, M.
Treatment of neurodegenerative diseases by classical pharmacotherapy is restricted by blood-brain barrier which prevents access to the brain of potentially therapeutic molecules. Recent progress in the knowledge of pathophysiological molecular processes, and in the development of molecular biotechnology have opened the way to new therapeutic interventions for these disorders. This chapter reviews the most recent gene therapy strategies using experimental models for neurodegenerative diseases.
Open Access
Ultrastructural study of ovine pulmonary pasteurellosis, involvement of neutrophils and macrophages
(Murcia : F. Hernández, 2001) Gázquez, A.; Redondo, E.; Martínez, S.; Gómez, L.
Pasteurellosis is a common infectious disease characterised by fibrinous pneumonia and involving neutrophils and macrophages. This study aimed to determine the timing and extent of the pathogenic involvement of these cell elements in lesions induced in experimentally-infected lambs. A concentration of approximately 3 x 1 0 ~ba cterialml. was inoculated into 30 two-month-old disease-free Merino lambs. Five lambs were assigned to each of five experimental batches, slaughtered on days 1, 3,7, 11 and 15 following intratracheal inoculation, and to one control batch inoculated with a sterile solution. One control animal was slaughtered at the same time as each experimental batch. More characteristic lesions occur in bronchioles, peribronchial tissue and alveoli and are characterised by fibrinous processes. From the start of the experiment, epithelial-cell disruption and loss of microvilli were apparent; cell debris, desquamate cells and bacterial elements were observed in bronchiolar lumina, embedded in a fibrillar granular material. Alveolar structures displayed fewer neutrophils and macrophages, containing phagocytic vacuoles. Laminar bodies were apparent in type 11 pneumocytes. The interseptal area contained similar cell types, as well as abundant edema. In the course of the experiment, macrophage numbers increased in al1 the areas involved, with signs of intense phagocytic activity. The final phase of the experiment was characterised by a mild interseptal infiltrate and by clear alveolar lumina.
Open Access
The immunohistochemical expression of stress-response protein (srp) 60 in human brain tumours: Relationship of srp 60 to the other five srps, proliferating cell nuclear antigen and p53 protein
(Murcia : F. Hernández, 2001) Kato, S.; Kato, Massuo J.; Hirano, A.; Takikawa, M.; Ohama, E.
This study analyzed the expression of stressresponse (heat-shock) protein 60 (srp 60) in a series of 158 human brain tumours. Immunohistochemical procedures were employed; cells of the human cervical cancer line HeLa S3 exposed to hyperosmolar stress served as positive controls. Deposits of reaction products were found in the cytoplasm. Approximately half of the glioblastomas multiforme (17/31), breast carcinoma metastases (6/10), and lung carcinoma metastases (5111) as well as about one-third of the astrocytomas (5113) and meningiomas (8123) had tumour cells that expressed srp 60. A positive reaction for srp 60 was also seen in some medulloblastomas (2/16), primitive neuroectodermal tumours (PNETs) (2/11), schwannomas (2121), and pituitary adenomas (2/7), but no positive reactions were observed with oligodendrogliomas and ependymomas. Compared with srp 60-negative tumours, srp 60-positive tumours coexpressed one or more stress-related proteins, among which srp 90, srp 72, srp 27, alphaB-crystallin and ubiquitin occurred with higher frequencies; a high correlation between srp 60 and the other five srps (0.88 - 0.97, pe0.01, Pearson correlation coefficient) was observed in srp 60-positive tumours. In contrast, the correlation coefficient in srp 60-negative tumours was not significant (-0.26 - 0.71). There was a tendency for the proliferating cell nuclear antigen (PCNA)-labeling index to be higher in glioblastomas, astrocytornas, medulloblastomas, PNETs, and breast and lung carcinoma metastases that expressed srp 60 than in those that did not. No significant immunohistochemical reactions of srp 60, PCNA and p53 protein were seen with sections of normal brain tissues. We conclude that primary and metastatic tumours of the brain produce srp 60 and that srp 60 in certain brain tumour cells may coexpress the other five srps. In addition, srp 60 expression might depend, in part, on proliferating potential.
Open Access
Immunohistochemical profile of galectin-8 expression in benign and malignant tumors of epithelial, mesenchymatous and adipous origins, and of the nervous system
(Murcia : F. Hernández, 2001) Danguy, A.; Rorive, S.; Decaestecker, C.; Bronckart, Y.; Kaltner, H.; Hadari, Y.R.; Goren, R.; Zich, Y.; Petein, M.; Salmon, I.; Gabius, H.J.; Kiss, R.
This study aims to investigate whether the immunohistochemical expression of galectin-8 could be used as a diagnostic marker in tumor tissues of various histogenetic origins including specimens from epithelial (n=145), mesenchymatous (n=16), adipous (n=10) and central and peripheral nervous system (n=25) tissue, and 4 mesotheliomas. Immunohistochemical reactions were carried out with a polyclonal anti-galectin-8 antibody and histological slides from tissues derived from the files of the Laboratory of Anatomopathology of University Erasmus Hospital, Brussels. Formalin-fixed paraffinembedded tissues of 45 normal cases as well as 41 benign and 114 malignant tumors were studied. Marked decreases in immunohistochemical galectin-8 expression were obsewed in colon (p=0.001), pancreas (p=0.007), liver (p=0.0008), skin (p=0.002) and larynx (p=0.02) tissue when comparing malignant tissue to normal tissue andlor benign tumors. The reverse relationship was observed for breast tissue (p=0.007). No statistically significant differences (p>0.05) were detected when comparing normal tissue andlor benign to malignant tumors in lung, bladder, kidney, prostate and stomach tissue. Significant galectin-8 expression was also measured in non-epithelial tissue including tumors of the central and peripheral nervous system as well as in skeletal muscle and mesotheliomas. Immunohistochemical monitoring of galectin-8 thus reveals an organtype- dependent regulation of expression upon malignant transformation of various tissue types of epithelial origin. This observation will prompt further studies to 0ffprin.int requests to: Roberl Kiss, Ph.D., Laboratory of Histopathology, Faculty of Medicine - Free University of Brussels, 808 route de Lennik - 1070 Brussels, Belgiurn. Fax: 322 555 62 85. e-rnail: rkiss@ulb.ac.be delineate any relationship with prognosis
Open Access
Widespread cellular distribution of aldehyde oxidase in human tissues found by immunohistochemistry staining
(Murcia : F. Hernández, 2001) Moriwaki, Y.; Yamamoto, T.; Takahashi, S.; Tsutsumi, Z.; Hada, T.
Aldehyde oxidase (EC 1.2.3.1) is a xenobiotic metabolizing enzyme that catalyzes a variety of organic aldehydes and N-heterocyclic compounds. However, its precise pathophysiological function in humans, other than its xenobiotic metabolism, remains unknown. In order to gain a better understanding of the role of this enzyme, it is important to know its exact localization in human tissues. In this study, we investigated the distribution of aldehyde oxidase at the cellular leve1 in a variety of human tissues by immunohistochemistry. The enzyme was found to be widespread in respiratory, digestive, urogenital, and endocrine tissues, though we also observed a cellspecific localization in the various tissues studied. In the respiratory system, it was particularly abundant in epithelial cells from the trachea and bronchium, as well as alveolar cells. In the digestive system, aldehyde oxidase was observed in surface epithelia of the small and large intestines, in addition to hepatic cells. Furthermore, the proximal, distal, and collecting tubules of the kidney were immunostained with various intensities, while glomerulus tissues were not. In epididymus and prostate tissues, staining was observed in the ductuli epididymidis and glandular epithelia. Moreover, the adrenal gland, cortex, and notably the zona reticularis, showed strong immunostaining. This prevalent tissue distribution of aldehyde oxidase in humans suggests some additional pathophysiological functions besides xenobiotic metabolism. Accordingly, some possible roles are discussed.
Open Access
Visualization of bioavailable liposomal doxorubicin using a non-perturbing confocal imaging technique
(Murcia : F. Hernández, 2001) Krishna, R.; Ghiu, G.; Mayer, L.D.
Commonly employed tissue processing techniques can significantly alter tissue drug distribution patterns for liposomal encapsulated drugs by virtue of drug leakage via loss of membrane integrity. We report here a method that has been developed to determine the fluorescence of bioavailable doxorubicin (DOX) in tissues after administration of liposomal DOX formulations. A non-perturbing confocal fluorescence microscopy (CFM) technique with image processing analysis was used with unprocessed fresh tissues. This method takes advantage of the fact that considerable quenching occurs when DOX is within liposomes, leading to the selective visualization of the fluorescence due to DOX released from liposomes. We demonstrate that fresh tissue confocal imaging can be applied to provide detailed drug distribution information with improved accuracy and is a superior method for analyzing tissue distribution of liposome entrapped fluorescent agents
Open Access
F9 embryocarcinoma cells, a cell autonomous model to study the functional selectivity of RARs and RXRs in retinoid signaling
(Murcia : F. Hernández, 2001) Rochette-Egly, C.; Chambon, P.
Mouse F9 embryocarcinoma (EC) cells constitute a well established cell-autonomous model system for investigating retinoid signaling in vitro as, depending on culture conditions, retinoic acid (RA) can induce their differentiation into either primitive, parietal or visceral extraembryonic endoderm-like cells. These RA-induced differentiations are accompanied by decreases in proliferation rates, modifications of expression of subsets of RA-target genes, and induction of apoptosis. To elucidate the roles played by the multiple retinoid receptors (RARs and RXRs) in response to RA treatments, F9 EC cells lacking one or severa1 RARs or RXRs were engineered through homologous recombination. Mutated RARs and/or RXRs were then reexpressed in given RAR or RXR null backgrounds. WT and mutant cells were also treated with different combinations of ligands selective for RXRs and/or for each of the three RAR isotypes. These studies lead to the conclusion that most RA-induced events (e.g. primitive and visceral differentiation, growth arrest, apoptosis and activation of expression of a number of genes) are transduced by RARy/RXRa heterodimers, whereas some other events (e.g. parietal differentiation) are mediated by RARa/RXRa heterodimers. They also demonstrate that both AF-1 and AF-2 activation functions of RARs and RXRs, as well as their phosphorylation, are differentially required in these RA-induced events. In RARy/RXRa heterodimers, the phosphorylation of RARy is necessary for triggering primitive differentiation, while that of RXRa is required for growth arrest. On the other hand, phosphorylation of RARa is necessary for parietal differentiation. Thus, retinoid receptors are sophisticated signal integrators that transduce not only the effects of their cognate ligands, but also those of ligands that bind to membrane receptors.