Histology and histopathology Vol.16, nº 2 (2001)

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Open Access
The role of gicerin, a novel cell adhesion molecule, in development, regeneration and neoplasia
(Murcia : F. Hernández, 2001) Tsukamoto, Y.; Taira, E.; Miki, N.; Sasaki, F.
Neurite outgrowth factor (NOF) is an extracellular matrix (ECM) protein in the laminin family and its ligand, gicerin, is a novel cell adhesion molecule in the immunoglobulin superfamily. Gicerin has a homophilic adhesive activity as well as a heterotypic manner to NOF. In the nervous systems, gicerin is expressed during developmental stage when neurons migrate or extend neurites to form a neural network. Gicerin promotes neurite extension and migration of embryonic neurons in vitro by its homophilic and heterophilic adhesion activities. Introduction of antigicerin antibody into early developing eyes perturbs the layer formation of neural retina. These data suggest that gicerin participates in the formation of neural tissues. Gicerin is also expressed in other non-neural tissues; in epithelia of trachea, kidney and oviduct, gicerin expression is restricted in the developmental period. In contrast, muscular tissues and endothelial cells express gicerin continuously even after maturation. Interestingly, gicerin re-appears strongly in the regenerating epithelia of trachea, kidney and oviduct, and also anti-gicerin antibody disrupts the healing process of trachea. Furthermore, gicerin and NOF are overexpressed in the chicken nephroblastomas (Wilm's tumor) and oviductal adenocarcinomas. In vitro analyses show that gicerin adhesive activities can promote binding among tumor cells and adhesion of tumor cells to NOF. A polyclonal antibody against gicerin also perturbs the re-attachment of cancer cells onto metastasizing sites. It is clear from these studies that gicerin is a potential effector for pathological tissue formation as well as for normal development.
Open Access
The histochemical profiles of fibre types in porcine skeletal muscle
(Murcia : F. Hernández, 2001) Vázquez, J.Mª; Moreno, F.; Gil Cano, Francisco; Latorre Reviriego, Rafael Manuel; López Albors, Octavio Miguel; Ramírez Zarzosa, Gregorio José
Using a variety of histochemical methods -mATPase staining after alkaline and acid preincubations, NADH-TR and a-MGPDH- we have investigated the fibre types in porcine skeletal muscle. The results reveal that four major fibre types -1, IIA, IIB and II*- can be separated histochemically in Longissimus lumborum muscle of Landrace pigs. The histochemical properties of the muscle fibre type 11* are very similar to that of type IIX described in other mammals. The existence of IIX fibres in pig muscle has been recently demonstrated by molecular biology techniques and our results validate the use of histochemistry (mATPase) as an easy methodology to differentiate the three fast myosins (type 11 fibres) in pig muscle.
Open Access
Decrease in calcitonin and parathyroid hormone mRNA levels and hormone secretion under long-term hypervitaminosis D3 in rats
(Murcia : F. Hernández, 2001) Fernández-Santos, J.M.; Utrilla, J.C.; Conde, E.; Hevia, A.; Loda, M.; Martín Lacave, Inés
In calcium homeostasis, vitamin D3 is a potent serum calcium-raising agent which in vivo regulates both calcitonin (CT) and parathyroid hormone (PTH) gene expression. Serum calcium is the major secretagogue for CT, a hormons product whose biosynthesis is the main biological activity of thyroid Ccells. Taking advantage of this regulatory mechanism, long-term vitamin D3-induced hypercalcemia has been extensively used as a model to produce hyperactivation, hyperplasia and even proliferative lesions of C-cells, supposedly to reduce the sustained high calcium serum concentrations. We have recently demonstrated that CT serum levels did not rise after long-term hypervitaminosis D3. Moreover, C-cells did not have a proliferative response, rather a decrease in CT-producing C-cell number was observed. In order to confirm the inhibitory effect of vitamin D3 on C-cells, Wistar rats were administered vitamin D3 chronically (25,000 IUId) with or without calcium chloride (CaC12). Under these long-term vitaminD -hypercalcemic conditions, calcium, active metaboetes of vitamin 4 , CT and PTH serum concentrations were determined by RIA; CT and PTH mRNA levels were analysed by Northern blot and in situ hybridization; and, finally, the ultrastructure of calciotrophic hormone-producing cells was analysed by electron microscapy. Our results show, that, in rats, long term administration of vitamin D3 results in a decrease in hormone biosynthetic activities of both PTH and CTproducing cells, albeit at different magnitudes. Based upon these results, we conclude that hypervitaminosis D3-based methods do not stimulate C-cell activity and can not be used to induce proliferative lesions of calcitonin-producing cells.
Open Access
Cross-talk between RON receptor tyrosine kinase and other transmembrane receptors
(Murcia : F. Hernández, 2001) Danilkovitch-Miagkova, A.; Leonard, E.J.
RON is a transmembrane receptor tyrosine kinase that mediates biological activities of Macrophage Stimulating Protein (MSP). MSP is a multifunctional factor regulating cell adhesion, motility, growth and survival. MSP binding to RON causes receptor tyrosine phosphorylation leading to up-regulation of RON catalytic activity and subsequent activation of downstream signaling molecules. Recent studies show that RON is spatially and functionally associated with other transmembrane molecules including adhesion receptors integrins and cadherins, and cytokine and growth factor receptors IL-3 Bc, EPOR and MET. For example, MSP-induced cell shape change is mediated via RON-activated IL-3 Bc receptor. Activation of integrins causes MSP-independent RON phosphorylation, and the integrin/RON collaboration regulates cell survival. Thus, RON can be activated without MSP by ligand stimulation of RON-associated receptors, and MSP-activated RON can cause ligandindependent activation of RON-associated receptors. As a result of the receptor cross-activation RON-specific pathways become a part of a signal transduction network of other receptors, and conversely signaling pathways activated by other receptors can be used by RON. This receptor collaboration extends the spectrum of cellular responses generated by MSP and by putative ligands of RON-associated receptors. However signaling pathways involved in the receptor cross-talk and underlying activation mechanisms remain to be investigated. The purpose of this review is to summarize data and to discuss a role of cross-talk between RON and other transmembrane receptors.
Open Access
Effect of diabetic state on co-localization of substance P and serotonin in the gut in animal models
(Murcia : F. Hernández, 2001) Spangeus, A.; Forsgren, Sture; El-Salhy, M.
Changes in the numbers of serotonin- and substance P-immunoreactive (IR) cells occur in severa1 animal models of diabetes. It is not known, however, whether these changes are a result of actual ceii loss or are caused by modified gene expression in ceíls showing co-localization of serotonin and substance P. The pattern of mono- and co-expression of serotonin, as well as of substance P, was therefore investigated in gastrointestinal endocrine cells from animal models of human type 1 and type 2 diabetes, namely non-obese diabetic (NOD) and obese diabetic (oblob) mice. Immunocytochemical staining by the avidin-biotin complex method was performed for computerized image analysis of each cell type, and by immunofluorescence double staining to study co-localization. Tissues from antrum, proximal duodenum and dista1 colon were investigated. Co-localization of serotonin- and substance P-IR was found in al1 investigated parts of the gut. In antrum, substance P immunoreactivity was found exclusively in serotonin-IR cells. In both NOD and oblob mice there was a reduced number of substance PIR cells, but an unchanged serotonin-IR cell count, which thus tallies with a shut-off of substance P expression in antral enterochromaffin cells. In duodenum, both diabetes models showed a decreased number of serotonin-IR cells. Furthermore there was a decreased number of substance P-IR cells in the type 2 model. The proportion of serotonin-IR cells showing substance P-immuno-reactivity was decreased in both diabetic models, thus indicating a shut-off of substance P-gene expression. However, this does not fully explain the changes in duodenum, but the diabetic state probably affects the number of mono-expressed cells as well. In colon, no change was found in diabetic mice regarding co-localization of substance P and serotonin. However, pre-diabetic NOD mice showed a decreased proportion of substance P in serotonin-IR cells, which might be explained by the increased number of serotonin-IR cells, combined with an unchanged number of substance P-IR cells. In conclusion, diabetic animal models of both type 1 and type 2 appear to have a combination of decreased expression of substance P in serotonin-IR cells of both antrum and duodenum, as well as a change in the number of mono-expressed cells. The pattern in colon, on the other hand, seems to be unaffected.
Open Access
The localization of thrombospondin-1 (TSP-1), cysteine-serine-valine-threoninecysteine- glycine (CSVTCG) TSP receptor, and matrix metalloproteinase-9 (MMP-9) in colorectal cancer
(Murcia : F. Hernández, 2001) Wakiyama, T.; Shinohara, T.; Shirakusa, T.; John, A.S.; Tuszynski, G.P.
Thrombospondin-1 (TSP-1) is a 450 kDa matrix bound glycoprotein involved in tumor invasion, metastasis, and angiogenesis. One of the receptors involved in TSP-1 mediated tumor cell adhesion and metastasis is the cysteine-serine-valine-threoninecysteine- glycine (CSVTCG) receptor. One mechanism of TSP-1 in promoting tumor cell metastasis involves the up-regulation of matrix metalloproteinase-9 (MMP-9) expression, specifically through the CSVTCG TSP-1 receptor. TSP-1 and its CSVTCG receptor has been implicated in tumor progression in a variety of cancers including breast adenocarcinomas, head and neck squamous cell carcinomas, and pancreatic carcinomas. In this study, we examined 99 cases of colorectal cancer by immunohistochemical analysis to investigate 1) the localization of TSP-1 and CSVTCG TSP-1 receptor, 2) the relationship with MMP-9, and 3) the correlation of expression with clinical staging. Strong expression of TSP-1 was observed in the submucosa or the serosa adjacent to the tumor. Positive staining for CSVTCG TSP-1 receptor was observed in tumor cells and microvessels. MMP-9 was also expressed in tumor cells. In addition, staining intensity of CSVTCG TSP-1 receptor was higher in poorly differentiated adenocarcinoma than well or moderately differentiated adenocarcinoma. Tumors in which inflammatory cells stained strongly for CSVTCG TSP-1 receptor correlated with decreased incidence of distant metastasis and angiogenesis. These data were consistent with our previous studies for breast, pancreatic, and head and neck carcinoma. They suggest an important role for TSP-1 and CSVTCG TSP-1 receptor in tumor progression in colorectal cancer.
Open Access
Appearance of vascular endothelial growth factor (VEGF) in femoral head in the growing rat
(Murcia : F. Hernández, 2001) Ichigatani, M.; Saga, T.; Yamaki, K.; Yoshizuka, M.
In this study, we examined the appearance of vascular endothelial growth factor (VEGF) in the femoral head of the growing rat using an immunocytochemical technique. Our results showed VEGF-immunopositive cells existed in the inner region and peripheral region of the femoral head at each developmental stage. In the 19-day-old fetus, immunopositive mesenchymal cells were demonstrated in the peripheral region of the femoral head. At 1 to 10 days after birth, VEGF immunoreactivities were observed in the osteoblasts, osteoclasts, periosteum, perichondrium and cartilage matrix of the femur. At 15 days after birth, VEGF immunoreactive chondrocytes appeared in the apex area of the femoral head. In this stage, the femoral head is still constituted by chondrocytes and no apparent vascular formation has been observed. Thereafter, the immunopositive chondrocytes in the femoral head increased in number. The penetration of capillaries was recognized within the ligament of the femoral head at 60 days after birth. The results indicate that some chondrocytes in the femoral head produce VEGF before the beginning of ossification, and that VEGF may play an important role in the penetration of blood vessels into the femoral head from the ligament of the femoral head.
Open Access
Papillary adenoma of the kidney with mucinous secretion
(Murcia : F. Hernández, 2001) Val-Bernal, José Fernando; Pinto, J.; Gómez-Román, J.J.; Mayorga, M.; Villoria, F.
Although infrequently, mucin secretion has previously been reported in papillary renal cell carcinoma. We here investigate the presence of mucin in a series of 93 renal papillary adenomas in 58 patients. Acid mucin was present in four cases (4.3% of the tumors; 6.9% of the patients), in which basophilic mucin secretion was evident with hematoxylin-eosin. To the best of our knowledge mucin secretion has not been reported in renal papillary adenoma. We describe two different types of mucin secretion: intracytoplasmic and luminal. The secretion was intracellular in numerous scattered tumor cells in two cases, foca1 luminal in one case, and mixed intracellular and luminal in another case. Mucin production, despite its low frequency, can be considered as an additional feature of renal papillary adenoma. Mucin production suggests that renal papillary adenoma and papillary renal cell carcinoma are actually not two independent biological processes, but a continuum of one biological process.
Open Access
Cell adhesion molecules in human osteoblasts: structure and function
(Murcia : F. Hernández, 2001) Bennett, J.H.; Moffatt, S.; Horton, M.
Osteoblasts and bone lining cells form a near continuous layer covering the bone surface and interactions between these cells and the organic matrix of bone are important determinants of osteoblast proliferation and differentiation. In addition, cells of the osteoblast-lineage form functional cornrnunications with each other, with the extra-cellular matrix and with osteocytes through cytoplasmic processes extending through canaliculi in the bone. Together, these cells form a network of putative importance in the regulation of skeletal homeostasis. Cell-cell and cell-matrix interactions are mediated by members of severa1 families of cell adhesion molecules, and knowledge of their interactions will be of fundamental importance in understanding the role of osteoblast in skeletal turnover . Here, the expression pattern of members of the major families of cell adhesion molecules by cells of the osteoblast lineage is reviewed. Special emphasis has been placed on human tissues. In addition, the possibility that cells at progressive stages of the osteoblast lineage have different profiles of cell adhesion molecule expression is explored, and the putative significance of cell-matrix interactions in human skeletal disease briefly discussed.
Open Access
Histological and histochemical observations in the stomach of the Senegal sole, Solea senegalensis
(Murcia : F. Hernández, 2001) Arellano, J.M.; Storch, V.; Sarasquete, C.
An histological and histochemical study was conducted on the stomach of adult Senegal sole, Solea senegalensis specimens. The stomach was made up of four distinct layers: mucosa, lamina propria-submucosa-, muscularis and serosa. Surface epithelial, glandular and rodlet cells were present in the mucosa. Cells of the columnar epithelium contained a basa1 nucleus. Numerous mitochondria, granular endoplasmic reticulum and Golgi apparatus consisting of severa1 parallel cisternae and vesicles were observed in the cytoplasm of these cells. The lysosomes were small, round and dense. The gastric glands were numerous in the pyloric and fundic regions but absent in the cardiac stomach. These glands were formed by two cell-types: light and dark cells. The light cells were characterised by numerous mitochondria, while dark cells had slightly fewer mitochondria and a tubulo-vesicular system. Rodlet cells similar to those observed in other teleostean fish were present among the epithelial cells. Although the epithelial cells of the mucosa contained a weak presence of neutral and acid mucopolysaccharides/mucosubstances, these substances were abundant in the lamina propria-submucosa. Proteins rich in arginine, lysine, cysteine and cystine were rarely present in the mucosa and lamina propriasubmucosa of stomach, while proteins rich in tyrosine were abundant in these layers. Acid phosphatase, and ATP-ase (pH 7.2 and 9.4) activities were detected in the mucosa and lamina propria-submucosa. Alkaline phosphatase activity was not detected.
Open Access
Tumor cell "dead or alive": Caspase and survivin regulate cell death, cell cycle and cell survival
(Murcia : F. Hernández, 2001) Suzuki, A.; Shiraki, K.
Cell death and cell cycle progression are two sides of the same coin, and these two different phenomenons are regulated moderately to maintain the cellular homeostasis. Tumor is one of the disease states produced as a result of the disintegrated regulation and is characterized as cells showing an irreversible progression of cell cycle and a resistance to cell death signaling. Severa1 investigations have been performed for the understanding of cell death or cell cycle, and cell death research has remarkably progressed in these 10 years. Caspase is a nomenclature referring to ICEICED- 3 cysteine proteinase family and plays a central role during cell death. Recently, severa1 investigations raised some possible hypotheses that caspase is also involved in cell cycle regulation. In this issue, therefore, we review the molecular basis of cell death and cell cycle regulated by caspase in tumor, especially hepatocellular carcinoma cells.
Open Access
Expression of VEGF-C and activation of its receptors VEGFR-2 and VEGFR-3 in trophoblast
(Murcia : F. Hernández, 2001) Dunk, C.; Ahmed, A.
Placental villous development requires the co-ordinated action of angiogenic factors on both endothelial and trophoblast cells. Like vascular endothelial growth factor (VEGF), VEGF-C increases vascular permeability, stimulates endothelial cell proliferation and migration. In the present study, we investigated the expression of VEGF-C and its receptors VEGFR-3 and VEGFR-2 in normal and intrauterine growth-restricted (IUGR) placenta. Immunolocalisation studies showed that like VEGF and VEGFR-1, VEGF-C, VEGFR-3 and VEGFR-2 co-localised to the syncytiotrophoblast, to cells in the maternal decidua, as well as to the endothelium of the large placental blood vessels. Western blot analysis demonstrated a significant decrease in placental VEGF-C and VEGFR-3 protein expression in severe IUGR as compared to gestationallymatched third trimester pregnancies. Conditioned medium from VEGF-C producing pancreatic carcinoma (Suit-2) and endometrial epithelial (Hec-1B) cell lines caused an increased association of the phosphorylated extracellular signal regulated kinase (ERK) in VEGFR-3 immunoprecipitates from spontaneously transformed first trimester trophoblast cells. VEGF121 caused dosedependant phosphorylation of VEGFR-2 in trophoblast cells as well as stimulating DNA svnthesis. In addition. premixing VEGFl with Yheparin sÚlphate proteoglycan wtentiated trooho8ast oroliferation and the association Lf p h ~ s p hw~ith- t i~e V~E~GF R-2 receptor. VEGF165- mediated DNA synthesis was inhibited by anti-VEGFR- 2 neutralising antibody. The results demonstrate functional VEGFR-2 and VEGFR-3 receptors on trophoblast and suggest that the decreased expression of VEGF-C and VEGFR-3 may contribute to the abnormai villous development observed in IUGR placenta.
Open Access
The hepatocytes of the brown trout (Salmo trutta f. fario):A quantitative study using design-based stereology
(Murcia : F. Hernández, 2001) Rocha, E.; Monteiro, R.A.F.; Oliveira, M.H.; Silva, M.W.
A stereological study was performed on brown trout hepatocytes aiming to disclose whether there are basic gender differences when minimal levels of sex hormones exist, and also to establish a platform for both interspecific comparisons and physiological correlations. We used the so-called "design-based stereology" (with no shape, size or orientation assumptions) and also some new related statistics. Twoyear- old brown trout were collected in April, and the livers were fixed by perfusion. From liver slicing to microscopical field selection, systematic sampling was used. Stereology was applied at light and electron microscopy. Target parameters were the relative and total hepatocyte number, the mean individual hepatocyte volume and surface, and also both relative and total volumes, and surfaces, either of organelles or of cell compartments. Observed variability was usually high, but the precision of estimates was proved to be globaily adequate facing the true biological variation amongst specimens. Females had more hepatocytes per liver ( 1 . 7 9 ~ 1 0v~s. 1 . 1 2~1 0~C) .o nsidering the individual hepatocytes, whereas no gender differences were detected in the cell volume, males had higher values of nuclear volume (199 vs. 151 pm3) and surface (170 vs. 131 pm2), endoplasmic reticulum volume I 1300 vs. 824 pm3), and microvilli volume (82 vs. 54 pm ) and surface (1445 vs. 975 pm2). However, when dealing with quantities per liver, gender differences were found only in the volumes of dense bodies (56 vs. 97 mm3) and of residual cytoplasm (169 vs. 341 mm3) - both volumes were higher in females. Functional implications of data are discussed, namely that females seem to have basic structural traits for coping with the later demands of breeding. Data also support that structural remodelling of hepatocytes occurs after breeding, urging to pursue seasonal studies (namely on lysosomes). We advanced the hypothesis that genders differ in microvilli surface just to maintain an optimal physiological surface-to-volume ratio. Interspecific similarities and differences were disclosed. For example, the number of hepatocytes/cm3 of parenchyma of brown trout was much lower than those reported in rainbow trout, but in both trouts femaies seem to have an higher cell number. In addition, when comparing the size of hepatocytes of brown trout with that from other fish and mammals it was suggested that major interspecific differences exist.
Open Access
lmmunohistochemical expression of the p53, mdm2, p21/Waf-1, Rb, p16, Ki67, Cyclin DI, Cyclin A and Cyclin B1 proteins and apoptotic index in T-cell lymphomas
(Murcia : F. Hernández, 2001) Kanavaros, Panagiotis; Bai, M.; Stefanaki, K.; Rontogianni, D.; Zioga, E.; Gorgoulis, V.; Agnantis, N.J.; Poussias, G.
Fifty-seven cases of T-cell lymphomas (TCL) including 5 lymphoblastic (T-LBL) and 52 peripheral TCL (PTCL) were analyzed by immunohistochemistry for the expression of p53, mdm2, p21, Rb, cyclin DI, cyclin A, cyclin B1, and Ki67IMIB1 proteins and 39/52 PTCL were also analyzed for the expression of p16 protein and for the presence of apoptotic cells by the TUNEL method. The aim was to search for abnormal immunoprofiles of p53 and Rb growth control pathways and to determine the proliferative activity and the apoptotic index of TCL. Abnormal overexpression of p53, p21 and mdm2, in comparison to normal lymph nodes, was found in 12/57,10/57 and 2/57 cases of TCL, respectively. Abnormal loss of Rb and p16 expression was found in 1/57 and 2/39 cases, respectively, whereas abnormal overexpression of cyclin D1 was not detected in any of the 57 cases. Our data revealed entity-related p53/p21/mdm2 phenotypes. Indeed, most nodal and cutaneous CD30+ anaplastic large cell lymphomas (ALCL) showed concomitant overexpression of p53 and p21 proteins (718 cases), and mdm2 was overexpressed in 2 p53-positive nodal ALCL. In contrast, overexpression of p53 was found in 3/17 cases of nodal peripheral TCL unspecified (PTCL-UC) and 217 non- ALCL cutaneous pleomorphic TCL. Overexpression of p21 protein was detected in 213 p53-positive PTCLUC and in 112 p53-positive non-ALCL cutaneous pleomorphic TCL. Finally, al1 the remaining 25 cases of TCL did not show p53 and p21 overexpression. Overali, the p53+/p21+ phenotype in 10157 TCL suggests wildtype p53 capable of inducing p21 expression. The highest apoptotic index (Al) was found in ALCL and a positive correlation between apoptotic index and Ki67 index (p<0.001) was detected. Ki67, cyclin A and cyclin Omrint requests to: Dr. Maria Bai, Medical School, Department of Pathology. University of loannina, 451 10, loannina, Hellas, Greece. Fax: 0651 -46209 #These authors contributed equally to the work. B1 expression was found in al1 57 TCL and on the basis of the combined use of these 3 variables, 3 groups of proliferative activity,.could be determined: a) high in ALCL and T-LBL, b) low in mycosis fungoides (MF) and y6 hepatosplenic TCL, and c) intermediate in the remaining TCL entities. The proliferative activity in the 12 p53 overexpressing cases was higher in comparison to the 45 p53-negative cases. Ki67 expresion in more than 25% of tumour cells showed significant correlation with p53 overexpression (p<0.001). Rb expression tended to be parallel to Ki67, cyclin A and cyclin B1 expression in al1 but one case of nodal PTCL-UC which displayed loss of RB expression. Interestingly, this case was p53-negative, whereas the p53-positive cases were Rb-positive. These findings suggest that different pathogenetic routes may function in some TCL, involving either the p53 or, less frequently, the Rb pathways.
Open Access
Beta-catenin expression pattern in small cell lung cancer: correlation with clinical and evolutive features
(Murcia : F. Hernández, 2001) Rodríguez-Salas, N.; Palacios, J.; Castro, J.de; Moreno, G.; González-Barón, M.; Gamallo, C.
B-catenin expression in small cell lung carcinomas (SCLC) was investigated by immunohistochemical method using antibodies against B-catenin. 50 pre-treatment biopsies were examined and the relationship between B-catenin expression and the patients' relevant clinical characteristics, response to chemotherapy, time to relapse or progression, and overall survival, were analyzed. 0-catenin expression exhibited different intensity within each sample, predominantly localized in the cytoplasm, and no sample showed nuclear expression. There was cytoplasmic hyperexpression in 14 cases, hypoexpression in 15 cases, and normal expression in 21 cases. We did not find any association between B-catenin expression and clinical data. Our results show, however, correlation between Bcatenin cytoplasmic hyperexpression with a shorter time to progression (p=0.0437) as well as with a shorter overall survival @=0.0253). B-catenin hyperexpression could have prognostic significance in SCLC.
Open Access
Reactive oxygen species (ROS)-generating oxidases in the normal rabbit cornea and their involvement in the corneal damage evoked by UVB rays
(Murcia : F. Hernández, 2001) Cejkova, J.; Stipek, S.; Crkovska, J.; Ardan, T.; Midelfart, A.
The comeas of albino rabbits were irradiated (5 min exposure once a day) with UVB rays (312 nm) for 4 days (shorter procedure) or 8 days (longer procedure). The eyes were examined microbiologically and only the corneas of sterile eyes or eyes with nonpathogenic microbes were employed. Histochemically, the activities of reactive oxygen species (ROS)- generating oxidases (xanthine oxidase, D-amino acid oxidase and a-hydroxy acid oxidase) were exarnined in cryostat sections of the whole corneas. Biochemically, the activity of xanthine oxidoreductase/xanthine oxidase was investigated in the scraped comeal epithelium. UVB rays significantly changed enzyme activities in the corneas. In comparison to the normal cornea, where of ROS-generating oxidases only xanthine oxidase showed significant activity in the corneal epithelium and endothelium, D-amino acid oxidase was very low and ahydroxy acid oxidase could not be detected at all, in the cornea repeatedly irradiated with UVB rays, increased activities of xanthine oxidase and D-amino acid oxidase were observed in al1 comeal layers. Only after the longer procedure the xanthine oxidase and D-amino acid oxidase activities were decreased in the thinned epithelium in parallel with its morphological disturbances. Further results show that the xanthine oxidaselxanthine oxidoreductase ratio increased in the epithelium together with the repeated irradiation with UVB rays. This might suggest that xanthine dehydrogenase is converted to xanthine oxidase. However, in comparison to the normal corneal epithelium, the total amount of xanthine oxidoredutase was decreased in the irradiated epithelium. It is presumed that xanthine oxidoreductase might be released extracellularly (into tears) or the enzyme molecules were denatured due to UVB rays (particulary after the longer procedure). Comparative histochemical and biochemical findings suggest that reactive oxygen species-generating oxidases (xanthine oxidase, D-amino acid oxidase) contribute to the comeal damage evoked by UVB rays.
Open Access
A human in vitro granuloma model for the investigation of multinucleated giant cell and granuloma formation
(Murcia : F. Hernández, 2001) Seitzer, U.; Haas, H.; Gerdes, J.
A method for the in vitro generation of granulomas and its use in the analysis of the human granulomatous response is summarized. As a target for the cellular response L3 larvae of Nippostrongylus brasiliensis are coincubated with human mononuclear blood cells, and within seven to fourteen days the development of blood monocytes to mature macrophages and to epithelioid cells and multinucleated giant cells (MGC) as typical constituents of granulomas clustered around the nematode is observed. The following review describes the uses and applications of this model for phenotyping, functional, formation and modulating studies of granulomas and MGCs, taking into account its unique features compared to other in vitro models. With respect to MGC formation, procedures are described and examples are given which allow the phenotyping of these cells using immunofluorescence and immunohistological techniques. In addition, the potential of this model for illuminating functional aspects of MGC is described applying an isolation protocol for MGC and a subsequent reverse-transcriptase polymerase chain reaction method for the analysis of single cells. Moreover, the significance and relevance of using this granuloma model is discussed in the follow up analysis of in vivo findings of interleukin-6 expression in MGC of granulomas of patients with sarcoidosis. These in vivo results implicated a role for interleukin-6 in granuloma and MGC development. The in vitro granuloma model was used to investigate potential modulatory effects of this cytokine by analysing the cell numbers and the number of MGC per in vitro granuloma, the size of the MGC formed, the fusion index and the morphology of the in vitro granuloma. The results demonstrated significant modulatory effects of interleukin-6 on the cell number per in vitro granuloma and on the morphology of the cells involved. Conceivably, elevated interleukin-6 levels may modulate granuloma formation with respect to the number of cells involved and in influencing distinct cell populations involved in granuloma formation.
Open Access
Transcriptional regulation of the bcl-x gene encoding the anti-apoptotic Bcl-xL protein by Ets, Rel/NFKB STAT and AP1 transcription factor families
(Murcia : F. Hernández, 2001) Sevilla, L.; Zaldumbide, A.; Pognonec, P.; Boulukos, K.E.
Transcription factors play an essential role in determining the fate of a cell by affecting the expression of target genes involved in proliferation, in differentiation and in programmed cell death. Under certain conditions, some of these factors are capable of deregulating expression of genes involved in the cell cycle andlor in programmed cell death resulting in uncontrolled proliferation of the cell. The focus of this review is on the transcriptional regulation of the bcl-x gene encoding the anti-apoptotic Bcl-xL protein. Since 1999, severa1 papers have implicated members of the Ets, R~~/NFKSBT, N and AP-1 families as transcription factors regulating bcl-x expression. A specific emphasis of these different transcription factor families on bcl-x regulation in hematopoietic cells is discussed
Open Access
Immunocytohistochemical characterization of pituitary cells of the bluefin tuna, Thunnus thynnus L
(Murcia : F. Hernández, 2001) Rodriguez-Gomez, F.J.; Rendón-Unceta, M.C.; Piñuela, C.; Muñoz-Cueto, J.A.; Sarasquete, C.
In this paper we report the first complete mapping of the pituitary in a tuna species. The various different adenohypophysis cell types of the bluefin tuna, Thunnus thynnus L. have been identified and located using different antisera against mammalian and piscine hormones and various histochemical techniques: PAS, Alcian Blue pH 2.5 and lectins -ConA and WGA- (Neutral and Acidic Glycoproteins); Bromophenol Blue (Proteins) and Tioglycollate-Ferric-Ferricianide-Fe111 (- S-S- groups). Prolactin (PRL) and adrenocorticotrophic (ACTH) cells were located in the rostral pars distalis (RPD) of the pituitary, while the proximal pars distalis (PPD) displayed gonadotrophic (GTH), thyrotrophic (TSH), somatotrophic (GH) and also a few PRL cells. Moreover, somatolactin (SL) and melanotrophic (MSH) cells were identified inside the pars intermedia (PI). Interestingly, some SL-immunoreactive fibers were dso detected in the neurohypophysis. Some GTH cells were also located on the exterior surface of the PI. Glycoproteins containing mannose (Man) andlor glucose (Glc); N-acetyl-glucosamine (GlcNAc) andlor sialic acid sugar residues, as well as -S-S- groups, were observed in GTH, TSH and SL cells. The Bromophenol Blue technique stained amphiphilic SL, acidophilic GH cells and weakly ACTH cells. GH and ACTH cells were unreactive to PAS, Alcian Blue, Tioglycollate-Ferric- Ferricianide-Fe111 and lectin (Con A and WGA) techniques. Finally, PAS reaction was positive in amphiphilic SL cells, which were PbH unreactive, while MSH and ACTH cells were stained with PbH technique
Open Access
The dynamics of cellular injury: transformation into neuronal and vascular protection
(Murcia : F. Hernández, 2001) Maiese, K.
Despite the immediate event, such as cerebral trauma, cardiac arrest, or stroke that may result in neuronal or vascular injury, specific cellular signal transduction pathways in the central nervous system ultirnately influence the extent of cellular injury. Yet, it is a cascade of mechanisms, rather than a single cellular pathway, which determine cellular survival during toxic insults. Although neuronal injury associated with several disease entities, such as Alzheimer's disease, Parkinson's disease, and cerebrovascular disease was initially believed to be irreversible, it has become increasingly evident that either acute or chronic modulation of the cellular and molecular environment within the brain can prevent or even reverse cellular injury. In order to develop rational, efficacious, and safe therapy against neurodegenerative disorders, it becomes vital to elucidate the cellular and molecular mechanisms that control neuronai and vascular injury. These include the pathways of free radical injury, the independent mechanisms of programmed cell death, and the downstream signal transduction pathways of endonuclease activation, intracellular pH, cysteine proteases, the cell cycle, and tyrosine phosphatase activity. Employing the knowledge gained from investigations into these pathways will hopefully further efforts to successfully develop effective treatments against central nervous system disorders.