Peculiar Localization Research Articles

Modulation of survival pathways in ovarian carcinoma cell lines resistant to platinum compounds

Because cytotoxic stress elicits various signaling pathways that may be implicated in cell survival or cell death, their alterations may have relevance in the development of platinum-resistant phenotype. Thus, in the present study, we investigated cell response to the epidermal growth factor receptor (EGFR) inhibitor gefitinib of ovarian carcinoma cell lines, including cells selected for resistance to cisplatin (IGROV-1/Pt1) and oxaliplatin (IGROV-1/OHP). Resistant sublines exhibited a marked decrease in sensitivity to gefitinib and resistance to apoptosis. Gefitinib was capable of inhibiting the phosphorylation of EGFR in all the studied cell lines. The Akt and extracellular signal-regulated kinase 1/2 (ERK1/2) kinases, which act downstream of EGFR, were constitutively active in the three cell lines, but phospho-ERK1/2 levels were increased in the two resistant sublines. This feature was associated with reduced sensitivity to the MEK1/2 inhibitor U0126. Pretreatment of resistant cells with U0126 resulted in restoration of sensitivity to gefitinib. Gefitinib was more effective in inhibiting ERK1/2 and Akt phosphorylation in IGROV-1 cells than in IGROV-1/OHP and IGROV-1/Pt1 cells. Phospho-p38 was up-regulated in the resistant sublines, indicating the concomitant activation of distinct mitogen-activated protein kinases. The up-regulation of phospho-p38 was associated with a peculiar localization of EGFR, which, in resistant sublines, was mainly internalized. In conclusion, our results indicate that the development of resistance to platinum drugs is associated with multiple alterations including deregulation of survival pathways activated by EGFR resulting in a reduced cellular response to gefitinib.

Carnitine palmitoyltransferase I in human carcinomas: A novel role in histone deacetylation?

Carnitine palmitoyl transferase I (CPT1) catalyzes the transport of long-chain fatty acids into mitochondria for β-oxidation. A link between CPT1 and apoptosis has been suggested on the basis of several experimental data. Nevertheless, results are contradictory about the effective role of CPT1 in cell survival control and cancer development. Conversely, Fatty acid synthase (FAS) enzyme, required for the synthesis of fatty acids, is found over-expressed in tumors and inhibition of FAS triggers apoptosis in human cancer cells.We have studied the tumor-specific modulation of CPT1 and FAS in human colorectal cancer (n=11) and breast carcinomas (n=24). CPT1 was significantly decreased in the cytoplasm of tumoral samples (p≤0.04), whereas FAS was increased (p≤0.04). A striking CPT1 nuclear localization was evident in the tumors (p≤0.04). In the nuclear environment the protein would modulate the levels of acetyl/acyl-CoA implicated in the regulation of gene transcription. At this purpose, we performed in vitro experiments using epithelial neoplastic (MCF-7, Caco-2, HepG2 cells) and non neoplastic cell lines (MCF-12F) confirming a nuclear localization of CPT1 protein exclusively in neoplastic cells. Moreover histone deacetylase (HDAC) activity showed significantly higher levels in nuclear extracts from neoplastic than from control cells. HDAC1 and CPT1 proteins co-immunoprecipitated in nuclear extracts from MCF-7 cells. The treatment with HDAC inhibitors such as trichostatin A and butyrate significantly decreased nuclear expression of CPT1 and its bond to HDAC1. We also identified the existence of CPT1A mRNA transcript variant 2 in MCF-7, beside to the classic isoform 1.The peculiar localization of CPT1 in the nuclei of human carcinomas and the disclosed functional link between nuclear CPT1 and HDAC1 propose a new role of CPT1 in the histonic acetylation level of tumors.

Oestrogen synthesis, oestrogen metabolism and functional oestrogen receptors in bovine aortic endothelial cells.

In order to investigate the mechanisms by which oestrogenic hormones influence the vascular system, we have studied their metabolism and the functioning of oestrogen receptors in bovine aortic endothelial cells from primo-secondary cultures, a widely studied model of vascular pathophysiology. We have demonstrated the enzymic activity of oestradiol-17 beta-hydroxysteroid dehydrogenase, 17-ketoreductase and aromatase in these cells. Immunocytochemical analyses, using two different monoclonal antibodies that recognize epitopes in the A/B domain of the oestrogen receptor, showed that this molecule has a predominantly cytoplasmic localization even after the addition of oestrogen to the culture medium. We showed that the hormone-receptor complexes were functional by demonstrating their transactivating ability in transfection experiments using the luciferase gene reporter and an oestrogen-responsive element transcriptional enhancer, although the amplitude of the response was in the range of only 140-150%: this was not a consequence of the presence of a specific limiting factor, but instead might be related to the peculiar subcellular localization of the oestrogen receptor.

Unusual breast enlargement in an infant: a case of breast lipoblastoma

We report the first case of lipoblastoma of the breast, diagnosed postoperatively in a 13-month-old infant. We discuss the management of this case, conditioned by peculiar localization, emphasizing the differential diagnosis of other causes of breast enlargement. Surgeons should know this condition and its implications, while dealing with developing breasts.

Glucocorticoid activation of the neuronal nicotinic acetylcholine receptor α7 subunit gene: involvement of transcription factor Egr-1

The α7 subunit is a component of nicotinic acetylcholine receptors expressed in bovine chromaffin cells. The peculiar localization of these receptors at adrenomedullary areas adjacent to the adrenal cortex suggests that factors, probably glucocorticoids, arising from the cortex might diffuse and regulate α7 receptor expression. In reporter gene transfection experiments, dexamethasone increased α7 promoter activity by up to fivefold in a concentration- and time-dependent manner despite the absence of consensus glucocorticoid receptor elements at the α7 promoter. Transcriptional activation induced by glucocorticoids was abolished through simultaneous mutation of at least two of the three sites for the immediate early transcription factor Egr-1, present in the proximal promoter region of the α7 subunit gene. Therefore, glucocorticoids activate the α7 subunit gene through Egr-1 in an indirect way.

Functional significance of a periplasmic Mn-superoxide dismutase from Aeromonas hydrophila.

A better understanding of the role of superoxide dismutases (SODs) from Aeromonas hydrophila and particularly the Mn-SOD which shares a peculiar localization within the bacterial periplasm and is only detected during the stationary phase of growth. A. hydrophila ATCC 7966 can express two distinct SODs: an Fe-SOD and an Mn-SOD. Using insertional mutagenesis, an Mn-SOD-deficient mutant was isolated. After growth of this mutant under conditions leading to the expression of an Mn-SOD, only the Fe-SOD could be detected in nondenaturing PAGE. Study of its response to the oxidative stress showed that the Mn-SOD was not implicated in the protection against intracellular superoxide but defended the bacterial cells against environmental superoxide. By protecting the bacteria against external superoxide, the role of the Mn-SOD from A. hydrophila is equivalent to that of the Cu/Zn-SOD from the well-studied Escherichia coli. The function of this Mn-SOD is in agreement with its periplasmic localization and may confer an advantage on the bacteria such as a virulence factor in cases of pathogenicity.

Involvement of drug-specific T cells in drug-induced interstitial nephritis

Rationale A rare and potentially severe form of drug allergic reaction can lead to interstitial nephritis. Its pathophysiological mechanisms are not well understood. Methods We analyzed drug-specific T cell responses of twelve patients with presumably drug induced interstitial nephritis (most without skin symptoms) with the lymphocyte transformation test and generated drug specific T cell clones (TCC) from peripheral blood and from kidney. Cytokine and chemokine production of the drug-specific TCC was measured by ELISA. Kidney biopsies were analyzed by immunohistochemistry. Results Proliferation assays of PBMCs or TCCs isolated from the twelve patients were performed but only two patients had a proliferative response to the drug. In particular, responses to flucloxacillin of cells generated from a male treated with rifampicin, flucloxacillin and gentamycin and to penicillin of cells from another patient treated with penicillin and vancomycin. The flucloxacillin-reactive cells showed an outgrowth of T cells bearing the T-cell receptor (TCR) Vβ9 and Vβ21.3, which suggests an oligoclonal immune response to the drug-antigen. We subsequently generated flucloxacillin-specific TCCs, all expressing CD4 and αβTCRs. Analysis of cytokines (IL-4, IL-5, IL-10, IFN-γ and TNF-α) revealed a quite heterogeneous secretion pattern, and cytotoxicity was lacking. Immunohistochemical stainings demonstrated an intertubular infiltration of CD4+ and CD8+ T cells. Conclusions Drug-specific T cells may play a decisive role in drug-induced interstitial nephritis. They may be difficult to detect in peripheral blood, as they might recognize a drug metabolite only presented in the kidney. Thus, the peculiar localization in the kidney may make these T cells more harmful.

Immunohistochemical localization of several neuropeptides in the adrenal gland of five species belonging to the order of Squamata

Recent studies have shown that several peptides coexist together with catecholamines within chromaffin cells. It has been demonstrated that the same chromaffln cell contains two or more bioactive substances forming several cellular subpopolations. These peptides play an important physiological role within the adrenal gland. Since very few data were available about the localization of peptides in reptiles, the aim of this study was to investigate the presence of different neuropeptides within the adrenal gland of species belonging to different families of this class of vertebrates. In particular, the immunohistochemical distribution of Leu‐enkephalin (Leu‐ENK), neuropeptide Y (NPY), somatostatin (SOM), substance P (SP), and vasoactive intestinal peptide (VIP) was studied in several species belonging to the order of Squamata. The results obtained showed that some of these peptides, e.g. SP, NPY, and VIP were widely distributed in most chromaffin cells, whereas Leu‐ENK was present only in a small number. Considering the distribution of all neuropeptides, there were no fundamental differences among different species, except for the peculiar localization of SOM in the Tarentola mauritanica adrenal gland. The results obtained represent a morphological basis for further studies about the role played by these peptides in the control of reptilian adrenal gland functionality.

Localization of the m5 muscarinic cholinergic receptor in rat circle of Willis and pial arteries

The expression and microanatomical localization of the muscarinic cholinergic m5 receptor subtype was investigated in rat circle of Willis and pial arteries by in situ hybridization, immunoblotting and immunohistochemistry. In situ hybridization histochemistry revealed a strong signal in the endothelium of circle of Willis and pial arteries and a moderate signal in the tunica media of the same arteries, within smooth muscle. Exposure of membranes of arteries to anti-m5 receptor protein antibodies caused the development of a band of approximately 81 kDa. Immunohistochemistry revealed the accumulation of m5 receptor protein immunoreactivity primarily within endothelium of circle of Willis and cerebral arteries and to a lesser extent in the tunica media, within smooth muscle. Medium (external diameter 200–100 μm) and small-sized (external diameter smaller than 100 μm) pial arteries displayed a significantly higher immune staining than large-sized pial arteries or circle of Willis arteries. The above data that are consistent with recent functional studies reporting cholinergic dilation of cerebral blood vessels mediated via a m5 receptor, have shown that both endothelial and muscular components of cerebral arteries synthesize and express a muscarinic m5 receptor. In view of the peculiar localization in cerebral vessels, handling of the muscarinic m5 receptor may be considered as an approach in the treatment of cerebrovascular disease.

Extracellular matrix-regulated p53 expression and nuclear localization in cultured Detroit 562 cells derived from pharyngeal carcinoma.

Tumor suppressor p53 functions as a transcriptional factor that regulates the cell cycle and apoptosis. A mutated p53 gene can result in decreased sequence-specific DNA binding and transcriptional activity of the p53 protein. In this study, we examined the regulatory role of the extracellular matrix components in p53 expression and nuclear localization in a Detroit 562 cell line derived from a pharyngeal carcinoma. When cultured on a polystyrene surface, type I collagen gel, or Matrigel containing basement membrane components, Detroit 562 cells showed a distinct response to extracellular matrix components morphologically. As shown by Western blotting, Detroit 562 cells cultured on Matrigel displayed an increased expression of p53 protein as well as an elevated nuclear p53 level, as compared with the cells cultured on the polystyrene surface or type I collagen gel. When cultured on Matrigel, nuclear p53-positive cells were exclusively localized to the outer surface layer of the cell clusters, whereas most of the inner cells showed no p53 expression. In Detroit 562 cell clusters on Matrigel, proliferative activities, as evaluated by proliferationcell nuclear antigen staining and bromo-deoxyuridine incorporation assay, were evenly distributed; virtually no apoptotic cells, as evaluated by the fluorescence TUNEL assay, were detected in the cell clusters, suggesting that the peculiar localization of nuclear p53-positive cells was not directly related to cell proliferation or apoptosis. These results indicate that p53 expression and its localization in Detroit cells were modulated by extracellular matrix signals, particularly by the basement membrane components in Matrigel.

Low-temperature transport properties ofNd2−xCexCuO4+δ:Metal-insulator crossover in the overdoped regime

The low-temperature in-plane resistivity and magnetoresistance have been measured for underdoped and overdoped single crystals ${\mathrm{Nd}}_{2\ensuremath{-}x}{\mathrm{Ce}}_{x}{\mathrm{CuO}}_{4+\ensuremath{\delta}}$ (x = 0.14, 0.16, 0.18, and 0.20) under magnetic field up to 14 T. We observed a crossover from a metallic to an insulating behavior in the low-temperature normal state for all four samples, which extends the metal-insulator crossover to the overdoped regime. The insulating behavior is marked by a logarithmic temperature dependence of the resistivity, suggesting that a peculiar charge localization is common in all cuprates.

Morphological and clinical characteristics of antrochoanal polyps: comparison with chronic inflammation-associated polyps of the maxillary sinus

Objectives: to compare morphological and clinical features of antrochoanal polyps and chronic inflammation-associated polyps of the maxillary sinus. Study design: histological and scanning electron microscopic examination of ten antrochoanal polyps and ten chronic inflammation-associated polyps of the maxillary sinus; comparison of clinical data in both groups of patients. Methods: following surgical removal, the polyps were halved, the halves being processed for routine light microscopy (formalin fixation, paraffin embedding, HE staining) and scanninng electron microscopy (formaldehyde/glutaraldehyde fixation, critical point drying, gold coating), respectively. Clinical data were retrospectively reviewed, tabulated and compared. Results: the antrochoanal polyps differed from chronic inflammation-associated polyps of the maxillary sinus only in a few minor features; slightly longer duration of the process, lower incidence of maxillary ostial obstruction, higher incidence of frequent headaches, persistent nasal obstruction, presence of cysts in the polyp stroma, thickened basement membrane, lower incidence of squamous cell metaplasia, and higher proportion of migratory cells in nasal smears. In two cases, allergy was diagnosed but it seemed not to influence the polyps, which did not show morphological features typical of allergy-associated (eosinophilic) polyps. Conclusions: In spite of minor differences, antrochoanal polyps can be regarded as chronic inflammation-associated polyps with cystic origin and peculiar localization.

Choroid plexus in the central nervous system: biology and physiopathology.

Choroid plexuses (CPs) are localized in the ventricular system of the brain and form one of the interfaces between the blood and the central nervous system (CNS). They are composed of a tight epithelium responsible for cerebrospinal fluid secretion, which encloses a loose connective core containing permeable capillaries and cells of the lymphoid lineage. In accordance with its peculiar localization between 2 circulating fluid compartments, the CP epithelium is involved in numerous exchange processes that either supply the brain with nutrients and hormones, or clear deleterious compounds and metabolites from the brain. Choroid plexuses also participate in neurohumoral brain modulation and neuroimmune interactions, thereby contributing greatly in maintaining brain homeostasis. Besides these physiological functions, the implication of choroid plexuses in pathological processes is increasingly documented. In this review, we focus on some of the novel aspects of CP functions in relation to brain development, transfer of neuro-humoral information, brain/immune system interactions, brain aging, and cerebral pharmaco-toxicology.

Scale Effects on Exciton Localization and Nonradiative Processes in GaN/AlGaN Quantum Wells

Previous experimental studies have allowed us to observe peculiar localization effects of excitons in GaN/AlGaN quantum wells grown by MBE, as well as efficient nonradiative inter-well carrier transfers. In this work, we use the envelope-function approximation to calculate exciton energies and wave functions. We show that the typical spatial extension of the electron and hole can by itself explain the localization and transfer processes, mainly because, due to its large effective mass, the in-plane extension of the hole is smaller than the average distance between two aluminum atoms in the barriers.

EXOTIC COHERENT STRUCTURES IN THE (2 + 1)-DIMENSIONAL BREAKING-SOLITON EQUATIONS

Hirota's bilinear form of the (2 +1)-dimensional breaking-soliton equations introduced byBogyovlenskii is deduced in a straightforward manner andused to construct wave-type solutions for the fieldvariables. The peculiar localization behavior of thesystem by the generating dromion for the composite fieldvariable qr is also brought out and is generalized to(1, N, 1) dromions.

Onset of keratin 17 expression coincides with the definition of major epithelial lineages during skin development.

The type I keratin 17 (K17) shows a peculiar localization in human epithelial appendages including hair follicles, which undergo a growth cycle throughout adult life. Additionally K17 is induced, along with K6 and K16, early after acute injury to human skin. To gain further insights into its potential function(s), we cloned the mouse K17 gene and investigated its expression during skin development. Synthesis of K17 protein first occurs in a subset of epithelial cells within the single-layered, undifferentiated ectoderm of embryonic day 10.5 mouse fetuses. In the ensuing 48 h, K17-expressing cells give rise to placodes, the precursors of ectoderm-derived appendages (hair, glands, and tooth), and to periderm. During early development, there is a spatial correspondence in the distribution of K17 and that of lymphoid-enhancer factor (lef-1), a DNA-bending protein involved in inductive epithelial-mesenchymal interactions. We demonstrate that ectopic lef-1 expression induces K17 protein in the skin of adult transgenic mice. The pattern of K17 gene expression during development has direct implications for the morphogenesis of skin epithelia, and points to the existence of a molecular relationship between development and wound repair.

Fourier transform IR and Raman spectroscopy and structure of carbohydrates

To determine the role of the skeletal base configuration of carbohydrate molecules, a comparative study was made, using IR and Raman spectroscopy, of theoretical calculations of the vibrational spectra of a series of carbohydrates differing in the configuration of CO (CH) bonds in various positions of the pyranose ring. The normal vibrations of carbohydrate molecules have, with few exceptions, close or coinciding frequencies; however, they differ greatly in the shape and contribution to the potential energy distribution (PED) of individual groups and bonds. Despite the cooperative character of the vibrations, each compound is characterized by a specific set of frequencies with a prevailing contribution to the PED of particular CO and CC bonds of the molecule. It is concluded that vibrations have a peculiar localization and that steric factors play an important role in the vibrational spectra of carbohydrates.

Mitochondria buffer Ca2+ entry but not intracellular Ca2+ release in mouse DRG neurones.

Mitochondrial Ca2+ pool was investigated in dorsal root ganglion neurones acutely isolated from 4-5 week old mice. Under the resting conditions application of mitochondrial protonophore CCCP triggered a slight increase in [Ca2+]i. In contrast, application of CCCP during the plateau phase of KCl-induced [Ca2+]i transient triggered large elevation in [Ca2+]i. In the presence of CCCP the amplitude of depolarization-induced [Ca2+]i elevation significantly increased, whereas the plateau phase of [Ca2+]i transient disappeared. On the contrary, CCCP failed to modulate [Ca2+]i transients resulting from caffeine-induced Ca2+ release. We concluded that mitochondrial Ca2+ pool distinguished the origin of Ca2+ ions entering the cytoplasm: it effectively buffers Ca2+ entering the cell through plasmalemmal channels but fails to regulate [Ca2+]i transients resulting from Ca2+ release from ER Ca2+ stores possibly due to peculiar localization of mitochondria far away from the Ca2+ release sites.

Underestimation of ovarian pathology: a review

Ovarian cancer is the fourth cause of death due to cancer in America and North Europe 1. In only one-third of cases is ovarian cancer diagnosed when still localized, with a survival rate after 5 years of 60-70%, while in most cases the disease is diagnosed in an advanced stage, with a prognosis quod vitam after 5 years of less than 10% 2. Ovarian masses have considerable legal risks, due to lack of specific symptomatology and to their peculiar localization. Thus, patients with an ovarian mass have to be always suspected as having a malignant disease, although histological examination is positive in only 1520% of cases 3.

Exotic coherent structures in the (2+1) dimensional long dispersive wave equation

In this paper, we investigate the integrability aspects of the (2+1) dimensional coupled long dispersive wave (2LDW) equation introduced recently by Chakravarty, Kent, and Newman and establish its Painlevé (P-) property. We then deduce its bilinear form from the P analysis and use it to construct wave type solutions for the field variables. We then identify line solitons for the composite field variable “qr” which eventually helps to bring out the peculiar localization behavior of the system by generating localized structures (dromions) for the composite field from out of only one ghost soliton driving the boundary. We have then extended this analysis to multidromion solutions.