Histology and histopathology Vol.24, nº8 (2009)
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- PublicationOpen AccessVaspin and amylin are expressed in human and rat placenta and regulated by nutritional status(Murcia : F. Hernández, 2009) Caminos, Jorge E.; Bravo, Susana B.; Garcés, Maria F.; González, C. Ruth; Cepeda, Libia A.; González, Adriana C.; Nogueiras, Rubén; Gallego, R.; Garcia-Caballero, Tomas; Cordido, Fernando; López, Miguel; Diéguez, C.Amylin (islet amyloid polypeptide) and vaspin (visceral adipose tissue specific serpin) are gut and adipocyte hormones involved in the regulation of body weight homeostasis. The aim of this study was to examine whether amylin and vaspin are expressed in human and rat placenta, as well as their regulation by nutritional status. Our results demonstrate that amylin and vaspin are localized in both human and rat placenta. In the rat term placenta vaspin was demonstrated in the trophoblast of the fetal villi, the labyrinth. Vaspin immunostaining in human placenta was localized in cytotrophoblast and syncytiotrophoblast in the first trimester placentas while in the third trimester vaspin was localized in the syncytiotrophoblast. Regarding amylin, rat placenta of 16 days of gestational age showed an intense immunostaining, mainly localized in the labyrinth. On the other hand, in the human third trimester placenta amylin immunoreactivity was intense in the syncytiotrophoblast of the chorionic villi and in decidual cells. Furthermore, placental amylin and vaspin showed an opposite pattern of expression during pregnancy, with vaspin showing the highest expression level at the end and amylin at the beginning of pregnancy. Finally, food restriction also has contrary effects on their expression, increasing vaspin but decreasing amylin placental mRNA and protein levels. Taken together, our results demonstrate that vaspin and amylin are modulated by energy status in the placenta, which suggests that these proteins may be involved in the regulation of placental metabolic functions.
- PublicationOpen AccessCytoplasmic inclusions of TDP-43 in neurodegenerative diseases: A potential role for caspases(Murcia : F. Hernández, 2009) Rohn, Troy T.TAR DNA-binding protein-43 (TDP-43) proteinopathies are classified based upon the extent of modified TDP-43 inclusions and include a growing number of neurodegenerative diseases including amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration with ubiquitin immunoreactive, tau negative inclusions (FTLD-U) and FTLD with motor neuron disease (FTLD-MND). In addition, TDP-43 inclusions have also been identified in a number of other neurodegenerative disorders including Alzheimer’s disease, corticobasal degeneration, Lewy body related diseases and Pick’s disease. Current understanding suggests that in these diseases, TDP-43 is relocated from the nucleus to the cytoplasm and sequestered into inclusions that contain modified TDP-43. Major modifications of TDP-43 have been identified as being hyperphosphorylation and proteolytic cleavage by caspases. In this review a summary of the major findings regarding the proteolytic modification of TDP-43 will be discussed as well as potential toxic-gain mechanisms these fragments may cause including cytoskeletal disruptions.
- PublicationOpen AccessIn situ detection of APRIL-rich niches for plasma-cell survival and their contribution to B-cell lymphoma development(Murcia : F. Hernández, 2009) Burjanadze, M.; Matthes, T.A proliferation inducing ligand (APRIL) is one of the most recently cloned members of the tumor necrosis factor (TNF) family. Early experiments implicated a pathophysiological role for APRIL in the promotion of solid tumors. Later, identification of APRIL receptors on B lymphocytes indicated a physiological role for APRIL in humoral responses. We have been able to generate antibodies that detect APRIL protein in human tissues. The study of in situ APRIL expression showed that APRIL mainly regulates late stages of B-cell humoral responses. It also provided evidence that APRIL may modulate tumor development in patients, but only for specific B-cell malignancies. Here, we will review to what extent fine characterization of in situ expression adds valuable information on APRIL (patho) physiological functions.
- PublicationOpen AccessTumor cell expression of podoplanin correlates with nodal metastasis in esophageal squamous cell carcinoma(Murcia : F. Hernández, 2009) Chuang, Wen-Yu; Yeh, Chi-Ju; Wu, Yi-Chin; Chao, Yin-Kai; Liu, Yun-Hen; Tseng, Chen-Kan; Chang, Hsien-Kun; Liu, Hui-Ping; Hsueh, ChuenPodoplanin is a mucin-like glycoprotein expressed in the lymphatic endothelium. It has been suggested to play a role in lymphangiogenesis, since podoplanin deficient mice were found to have dilated malfunctioning lymphatic vessels and lymphedema. High podoplanin expression in tumor cells was found to correlate with lymph node metastasis and poor clinical outcome in patients with oral squamous cell carcinoma (SCC). However, the prognostic significance of podoplanin expression in esophageal SCC remains unexplored. Herein, we studied podoplanin expression in 59 patients who underwent surgical resection of esophageal SCC, with 43 of them preceded by preoperative concurrent chemoradiotherapy (CCRT). We found that high podoplanin expression strongly correlated with clinical nodal metastasis (cN1; p=0.0063), which was associated with short survival (p=0.012). However, there was no direct association between high podoplanin expression and short survival. We also found that lymphatic vessel invasion in the resected esophagus was strongly associated with pathological nodal metastasis (pN1; p=0.00092). Our results suggest that podoplanin could also play a role in tumor aggressiveness in esophageal SCC, as well as in oral SCC.
- PublicationOpen AccessImproved methodology for the detection and quantification of the acrosome reaction in mouse spermatozoa(Murcia : F. Hernández, 2009) Lybaert, Pascale; Danguy, A.; Leleux, Fabienne; Meuris, Sylvain; Lebrun, PhilippeThis study evaluates the use of two fluorescein-labelled (FITC) plant lectins, Pisum sativum (edible pea) agglutinin (PSA) and Arachis hypogaea (peanut) agglutinin (PNA), in order to determine the most accurate and reliable method to experimentally detect and assess the acrosome reaction in mouse spermatozoa. PNA-FITC labelling was restricted to the acrosome and was not influenced by the fixation procedure; either absolute methanol or paraformaldehyde. In contrast, PSA-FITC not only labelled the acrosome, but also the whole head and the flagellum. This aspect was especially marked after methanol fixation. The cytoplasmic droplet, when present, was also stained by PSA-FITC. Incubation with the calcium ionophore ionomycin induced a concentration and time-dependent increase in the number of acrosome reactions. Compared to spotted preparations, smear samples exhibited a high proportion of spermatozoa with damaged acrosome. In conclusion, PNA-FITC labelling was more accurate than PSA-FITC labelling to detect the acrosome of mouse spermatozoa. The fixation method (methanol vs. paraformaldehyde) had no influence on the staining pattern of PNA-FITC labelling, but spotted preparations are recommended to avoid mechanical damage to the acrosome. Ionophore challenge confirmed the existence of a calcium-dependent acrosome reaction in mouse spermatozoa and validated the use of PNA-FITC to quantify this physiological process. The present study illustrates important methodological considerations which need to be taken into account in order to design a reliable and reproducible protocol for the study of the acrosome reaction.
- PublicationOpen AccessExpression of hexokinases and glucose transporters in treated and untreated oesophageal adenocarcinoma(Murcia : F. Hernández, 2009) Fonteyne, Philippe; Casneu, Veerle; Pauwels, Patrick; Van Damme, Nancy; Peeters, Marc; Dierckx, Rudi; Van de Wiele, ChristopheThe aim of this study was to assess the expression pattern of the high glucose affinity glucose transporters GLUT 1, 2, 3, 4, 8 and 9 and of hexokinases I, II and III in newly diagnosed oesophageal adenocarcinoma by means of immunohistochemistry. Twenty patients eligible to undergo primary surgery and 18 patients with incomplete pathological response following induction radio chemotherapy, all suffering from oesophageal adenocarcinoma, were included in the study. The intensity and amount of positive tumour cells in the immuno-reaction (histology score (Hscore)) for GLUT 1, 3, 4, 8 and 9 as well as for hexokinase I, II and III were assessed independently by two experienced observers, blinded to the clinical results. In patients that underwent primary surgery, Hscores of GLUT8 (µ 6.7; sd3.3) and GLUT1 (µ 5.5; sd: 5.3) were significantly higher than Hscores of GLUT9 (µ 2.2; sd 1.5) and GLUT3 (µ 3.2, sd: 2.5). Hscores of hexokinase I (µ : 8.3; sd: 4.3), II (µ 5.5, sd: 4.0) and III (Ì 1.5, sd: 0.7) were all significantly different from each other (p<0.04). In patients that underwent radiochemotherapy prior to surgical tumour resection, µ Hscores were 6.9 (sd: 4.4) for GLUT1, 6.8 (sd: 5.3) for GLUT3, 5.9 (sd: 4.2) for GLUT8, 3.4 for GLUT9 (sd: 2.7) and 2.3 (sd: 3.6) for GLUT 4. Hscores of GLUT1 and GLUT3 were significantly higher than Hscores of GLUT4. Finally, Hscores of patients with radiochemotherapy for GLUT3, hexokinase II and III were significantly higher when compared to patients that underwent primary surgery.
- PublicationOpen AccessBreast carcinoma vascularity, A comparison of manual microvessel count and Chalkley count(Murcia : F. Hernández, 2009) Dhakal, Hari Prasad; Bassarova, A.V.; Naume, Bjørn; Synnestvedt, Marit; Borgen, Elin; Kaaresen, Rolf; Schlichting, Ellen; Wiedswang, Gro; Giercksky, Karl- Erik; Nesland, Jahn M.. Manual counting of microvessels as intratumoral microvessel density (MVD) and Chalkley counting have been used in several studies to assess the prognostic impact of vascularity in invasive breast carcinomas. In our present study, the aim was to evaluate the prognostic value of angiogenesis in invasive breast carcinoma assessed by MVD and Chalkley techniques in the same series of patients. A total of 498 breast carcinoma patients with median follow up time 85 months were evaluated. The tumour vascularity was quantified by both manual microvessel count (MVD) and Chalkley count in CD34 stained breast carcinoma slides by a single investigator blinded to clinical information. Other relevant clinicopathological parameters were noted, including breast cancer related death and both loco-regional and systemic relapse. The patients were stratified by converting MVD and Chalkley counts to categorical variables to assess prognostic impact, and results were compared. High vascular grades using MVD count did not demonstrate any prognostic significance for breast cancer specific survival (BCSS) or distant disease free survival (DDFS) either in whole patient group (BCSS, p=0.517, DDFS, p=0.301) or in non-treated node negative patients (p>0.05). Chalkley count showed prognostic significance for both DDFS and BCSS in whole patient group (p<0.001) and also in untreated node negative patient group (p<0.05). In multivariate analysis, Chalkley count, but not MVD, retained the prognostic value for BCSS (p=0.007) and DDFS (p=0.014). The Chalkley count for assessing angiogenesis in invasive breast carcinomas demonstrated prognostic value. The Chalkley method appears to be the better method in estimating the prognostic impact of vascularity in invasive breast carcinomas.
- PublicationOpen AccessEfficient uptake of mannosylated proteins by a human Schwann cell line(Murcia : F. Hernández, 2009) Baetas-da-Cruz, Wagner; Alves, Lucinéia; Guimarães, Erick V.; Santos-Silva, Alessandra; Pessolani, Maria Cristina V.; Barbosa, Helene S.; Corte-Rea, Suzana; Cavalcante, Leny A.Complex carbohydrate structures are essential molecules of infectious microbes and host cells, and are involved in cell signaling associated with inflammatory and immune responses. The uptake of mannose-tailed glycans is usually carried out by macrophages, dendritic cells (DCs), and other professional phagocytes to trigger MHC class I- and MHC class II-restricted antigen presentation, and to promote T cell effector responses. Since Schwann cells (SCs) have been proposed as immunocompetent cells, we investigated whether a human cell line (ST88-14 cells) could bind mannosylated ligands in a specific manner. The saturation of uptake of mannosylated molecules by ST88-14 cells and the internalization and distribution pathway of these ligands were tested by cytometry and confocal plus electron microscopy, respectively. This uptake showed a dose-dependent increase, the saturation point being reached at high concentrations of mannosyl residues/240mM mannose. Merging of man/BSA-FITC and S100 labeling showed their partial, but, significant colocalization. Ultrastructural analysis of ST88-14 cells after incubation with HRP-colloidal gold, without or with subsequent chasing at 37°C, showed an initial location on the cell surface and temperature- and time-dependent internalization of the probe. Our findings suggest an efficient mannosylated ligand uptake system through putative lectin(s) that may be operational in inflammatory and immune responses.
- PublicationOpen AccessGenetic mouse models for the functional analysis of the perifibrillar components collagen IX, COMP and matrilin-3: Implications for growth cartilage differentiation and endochondral ossification(Murcia : F. Hernández, 2009) Zaucke, Frank; Grässel, SusanneThe mutual interaction of the two supramolecular compartments, the fibrillar and extrafibrillar matrix is a prerequisite for stability and integrity of the cartilage extracellular matrix. The fibrillar periphery, composed of collagen IX, matrilins and cartilage oligomeric matrix protein (COMP) among other components, constitutes the interface which mediates interactions between the two compartments. Mutations in these peripheral macromolecules cause a broad spectrum of skeletal conditions such as pseudoachondroplasia (PSACH) and multiple epiphyseal dysplasia (MED), which severely affect the organization and integrity of the cartilage growth matrix in humans. Transgenic and knockout mouse models for collagen IX, matrilin-3 and COMP and combinations thereof display cartilage abnormalities and pathologies of varying severity. Absence of collagen IX appears to cause the most severe growth plate phenotype with a profoundly disturbed morphological organization affecting size and shape of the long bones. Notably, similar growth plate phenotypes, including irregularities in the proteoglycan content, hypocellular central regions, disorganized proliferation columns with atypically shaped and oriented chondrocytes and alterations in the hypertrophic zone are observed in transgenic mice lacking other macromolecules or carrying mutations therein. These include collagens II and XI, integrin subunits, integrin linked kinase (ILK), HIF-1α, VEGFα and the tumor suppressor PTEN. Notably, mutations in ciliar proteins such as Kif3α, polaris or Smo/Gli severely affect the ability of chondrocytes to move and to become arranged in columns. Absence or mutational changes of a variety of different, non-related cartilage macromolecules apparently cause similar pathologies and abnormalities of the growth cartilage, suggesting a limited number of underlying molecular mechanisms
- PublicationOpen AccessAberrant CCND1 copies and cyclin D1 mRNA expression do not result in the production of functional cyclin D1 protein in anaplastic large cell lymphoma(Murcia : F. Hernández, 2009) Bobos, Mattheos; Kotoula, Vassiliki; Kaloutsi, Vassiliki; Karayannopoulou, Georgia; Papadimitriou, Constantine S.; Kostopoulos, IoannisScattered reports in the literature have shown that Cyclin D1 mRNA and protein may be expressed in anaplastic large cell lymphoma (ALCL). ALCLs are characterized by the presence of ALK translocations. Aberrant Cyclin D1 expression seems to promote proliferation in other types of lymphoma, while a growth promoting CCND1/TACSD1(TROP2) fusion product has also been described in tumors. Herein, we investigated 44 ALCL cases for chromosome 11 and CCND1 status (by FISH), cyclin D1 mRNA expression (by in situ hybridization and RT-PCR) and Cyclin D1 protein (by immunohistochemistry with two different monoclonal antibodies), as well as for the expression of Trop-2/GA733-1 (by immunohistochemistry). Polysomy of CCND1 (11q13) and chromosome 11 was found in 15/38 evaluated cases (39.5%). This change was specific for CD30+ neoplastic cells, as shown by double fluorescent staining. Neoplastic cells in the majority of ALCL expressed cyclin D1 mRNA (29/41 [70.7%]), in association with the presence of ALK translocations (p=0.024) and systemic, rather than cutaneous disease (p=0.021). Remarkably, however, Cyclin D1 protein was not detected in neoplastic cells (0/44 cases), neither were these found positive for Trop-2. In conclusion, aberrant copies of CCND1 / chromosome 11 may be observed in ALCL, probably as a consequence of the reported ploidy changes in these tumors. ALCL may often express cyclin D1 mRNA, which, however, does not result in the production of functional Cyclin D1 protein or Trop-2, suggesting that these proteins do not play a role in the pathogenesis of ALCL.
- PublicationOpen AccessExpression and distribution of GABAergic system in rat knee joint synovial membrane(Murcia : F. Hernández, 2009) Tamura, Shigenori; Watanabe, M.; Kanbara, Kiyoto; Yanagawa, Tetsuji; Watanabe, K.; Otsuki, Yoshinori; Kinoshita, MitsuoThe GABAergic system, found in the adult mammalian brain and composed of γ-aminobutyric acid (GABA), GABA synthesizing enzyme glutamate decarboxylase (GAD) and GABA receptors, is also located in many peripheral nonneuronal tissues. Studies suggest that synovial membranes possess GABA, and that GABA participates in the control of the inflammatory response in rheumatoid arthritis (RA). However, no studies on the GABAergic system in synovial membranes have been done so far. Therefore, expression and distribution of the GABAergic system in the synovial membrane of the normal rat knee joint were investigated by reverse transcription-polymerase chain reaction (RT-PCR) analyses and immunohistochemistry. Results of RT-PCR analysis showed that mRNA encoding the GAD65 and GABAB receptor subunits necessary for the assembly of functional receptors, R1 and R2, are expressed in the synovial membrane. GAD and GABAB receptor subunits were localized in macrophage-like A cells of the synovial membrane. Macrophage-like A cells of the synovial membrane have a GABA production system and GABAB receptors, and GABA seems to play functional roles in the synovial membrane.
- PublicationOpen AccessExpression of eight genes of nuclear factor-kappa B pathway in multiple myeloma using bone marrow aspirates obtained at diagnosis(Murcia : F. Hernández, 2009) Sampaio Almeida, Manuella S.; Vettore, André L.; Yamamoto, M.; Chauffaille, Maria de Lourdes L.F.; Zago, Marco Antônio; Colleoni, Gisele W. B.Purpose: To evaluate the expression of NF- κB pathway genes in total bone marrow samples obtained from MM at diagnosis using real-time quantitative PCR and to evaluate its possible correlation with disease clinical features and survival. Material and methods: Expression of eight genes related to NF-κB pathway (NFKB1, IKB, RANK, RANKL, OPG, IL6, VCAM1 and ICAM1) were studied in 53 bone marrow samples from newly diagnosed MM patients and in seven normal controls, using the Taqman system. Genes were considered overexpressed when tumor expression level was at least four times higher than that observed in normal samples. Results: The percentages of overexpression of the eight genes were: NFKB1 0%, IKB 22.6%, RANK 15.1%, RANKL 31.3%, OPG 7.5%, IL6 39.6%, VCAM1 10% and ICAM1 26%. We found association between IL6 expression level and International Staging System (ISS) (p=0.01), meaning that MM patients with high ISS scores have more chance of overexpression of IL6. The mean value of ICAM1 relative expression was also associated with the ISS score (p=0.02). Regarding OS, cases with IL6 overexpression present worse evolution than cases with IL6 normal expression (p=0.04). Conclusion: We demonstrated that total bone marrow aspirates can be used as a source of material for gene expression studies in MM. In this context, we confirmed that IL6 overexpression was significantly associated with worse survival and we described that it is associated with high ISS scores. Also, ICAM1 was overexpressed in 26% of cases and its level was associated with ISS scores.