Home About us Contact
|
Home About us Contact | ||
|
|
||
Strong Expression (strong + expression)
Selected AbstractsRegulation of the Neurofibromatosis 2 gene promoter expression during embryonic developmentDEVELOPMENTAL DYNAMICS, Issue 10 2006Elena M. Akhmametyeva Abstract Mutations in the Neurofibromatosis 2 (NF2) gene are associated with predisposition to vestibular schwannomas, spinal schwannomas, meningiomas, and ependymomas. Presently, how NF2 is expressed during embryonic development and in the tissues affected by neurofibromatosis type 2 (NF2) has not been well defined. To examine NF2 expression in vivo, we generated transgenic mice carrying a 2.4-kb NF2 promoter driving ,-galactosidase (,-gal) with a nuclear localization signal. Whole-mount embryo staining revealed that the NF2 promoter directed ,-gal expression as early as embryonic day E5.5. Strong expression was detected at E6.5 in the embryonic ectoderm containing many mitotic cells. ,-gal staining was also found in parts of embryonic endoderm and mesoderm. The ,-gal staining pattern in the embryonic tissues was corroborated by in situ hybridization analysis of endogenous Nf2 RNA expression. Importantly, we observed strong NF2 promoter activity in the developing brain and in sites containing migrating cells including the neural tube closure, branchial arches, dorsal aorta, and paraaortic splanchnopleura. Furthermore, we noted a transient change of NF2 promoter activity during neural crest cell migration. While little ,-gal activity was detected in premigratory neural crest cells at the dorsal ridge region of the neural fold, significant activity was seen in the neural crest cells already migrating away from the dorsal neural tube. In addition, we detected considerable NF2 promoter activity in various NF2-affected tissues such as acoustic ganglion, trigeminal ganglion, spinal ganglia, optic chiasma, the ependymal cell-containing tela choroidea, and the pigmented epithelium of the retina. The NF2 promoter expression pattern during embryogenesis suggests a specific regulation of the NF2 gene during neural crest cell migration and further supports the role of merlin in cell adhesion, motility, and proliferation during development. Developmental Dynamics 235:2771,2785, 2006. © 2006 Wiley-Liss, Inc. [source] Molecular cloning of several rat ABC transporters including a new ABC transporter, Abcb8, and their expression in rat testisINTERNATIONAL JOURNAL OF ANDROLOGY, Issue 3 2006Nathalie Melaine Summary Several members of the ABC transporter superfamily play an important role in testicular physiology and defence against anticancer drugs. Using a reverse transcription-polymerase chain reaction strategy with degenerate primers and rat testis RNA as template, we have looked for the presence of other members of this superfamily. Of the six partial cDNA found, five corresponded to ABC transporters already known ,Mdr1b, Mrp1, Tapl/Abcb9, Umat/Abcb6 and Sur2/Abcc9, and one presented a strong homology with mouse and human ABCB8. Using a 5, and 3, RACE approach, we cloned the full-length cDNA and found that the predicted protein presented 92% and 80% homology with the mouse and human proteins respectively. Strong expression of rat abcb8 was found in heart, brain and testis when compared with liver, lung and spleen. In the testis, rat abcb8 was expressed both in the somatic Sertoli cells and peritubular cells and in the germline (spermatogonia and pachytene spermatocytes). Furthermore, Umat/Abcb6 was very highly expressed in the testis (high amounts in meiotic pachytene spermatocytes and low amount in post-meiotic early spermatids). In conclusion, we confirm the presence of several ABC transporters in the testis and also provide evidence of the presence of Abcb8 in the organ. [source] Strong expression of IGF1R in pediatric gastrointestinal stromal tumors without IGF1R genomic amplification,INTERNATIONAL JOURNAL OF CANCER, Issue 11 2010Katherine A. Janeway Abstract Wildtype (WT) gastrointestinal stromal tumors (GISTs), lacking mutations in KIT or PDGFRA, represent 85% of GISTs in pediatric patients. Treatment options for pediatric WT GIST are limited. Recently, expression profiling of a limited number of pediatric and adult WT GISTs and more in depth study of a single pediatric WT GIST implicated the insulin-like growth factor 1 receptor (IGF1R) as a potential therapeutic target in pediatric WT GIST. We performed immunoblotting, SNP and FISH studies to determine the extent of expression, biochemical activation and genomic amplification of IGF1R in a larger number of pediatric WT GISTs. Pediatric WT GISTs expressed IGF1R strongly, whereas typical adult KIT mutant GISTs did not. IGF1R gene amplification was not detected in pediatric WT GISTs, and some KIT -mutant GISTs had IGF1R gene deletion due to monosomy 15. Despite the absence of apparent genomic activation mechanisms accounting for overexpression, clinical study of IGF1R-directed therapies in pediatric WT GIST is warranted. [source] Myxoid liposarcoma FUS-DDIT3 fusion oncogene induces C/EBP ,-mediated interleukin 6 expressionINTERNATIONAL JOURNAL OF CANCER, Issue 4 2005Melker Göransson Abstract The myxoid/round cell liposarcoma oncogene FUS-DDIT3 is the result of a translocation derived gene fusion between the splicing factor FUS and DDIT3. In order to investigate the downstream targets of DDIT3, and the transforming effects of the FUS-DDIT3 fusion protein, we have introduced DDIT3-GFP and FUS-DDIT3-GFP constructs into a human fibrosarcoma cell line. The gene expression profiles of stable transfectants were compared to the original fibrosarcoma cell line by microarray analysis. We here report that the NF,B and C/EBP , controlled gene IL6 is upregulated in DDIT3- and FUS-DDIT3-expressing fibrosarcoma cell lines and in myxoid liposarcoma cell lines. Strong expression of the tumor associated multifunctional cytokine interleukin 6 was confirmed both at mRNA and protein level. Knockdown experiments using siRNA against CEBPB transcripts showed that the effect of FUS-DDIT3 on IL6 expression is C/EBP , dependent. Chromatin immunoprecipitation revealed direct interaction between the IL6 promoter and the C/EBP , protein. In addition, the effect of DDIT3 and FUS-DDIT3 on the expression of other acute phase genes was examined using real-time PCR. We demonstrate for the first time that DDIT3 and FUS-DDIT3 show opposite transcriptional regulation of IL8 and suggest that FUS-DDIT3 may affect the synergistic activation of promoters regulated by C/EBP ,,B. © 2005 Wiley-Liss, Inc. [source] Vitamin D receptor distribution in intestines of domesticated sheep Ovis ammon f. ariesJOURNAL OF MORPHOLOGY, Issue 2 2008Katharina Riner Abstract The biologically active form of vitamin D, i.e., 1,25-dihydroxycholecalciferol or calcitriol, plays an important role in bone metabolism and calcium homeostasis, which is often disturbed at the onset of lactation in high milk-yielding domestic ruminants. Gene transcription is modulated via vitamin D receptors, but nongenomic effects of vitamin D via membrane receptors have also been described. In the intestines, vitamin D promotes calcium absorption via vitamin D receptors. Vitamin D receptors are of clinical relevance, but have not been systematically assessed within all segments of the intestine in any species. Thus, we present for the first time an immunohistochemical study of the distribution patterns of the vitamin D receptor protein in sheep, which may be the basis for present and future investigations on mineral homeostasis in domestic ruminants. Tissue probes of the intestines were collected from five lambs and five nonlactating and nonpregnant dams, fixed in formalin, embedded in paraffin, and used for the assessment of vitamin D receptor protein. Nuclear vitamin D receptor immunoreaction was scored semiquantitatively and exhibited a segment-specific distribution pattern. Goblet cells always were devoid of any vitamin D receptor immunoreaction. Surface epithelial cells and enterocytes of the crypt openings generally demonstrated only a weak immunoreaction. Basally and/or intermediately located crypt epithelial cells exhibited stronger immunoreactions in duodenum, jejunum, and colon descendens. This basal/intermediate to superficial gradient was most pronounced in the duodenum and less evident in jejunum and colon descendens and not observed in ileum and cecum. There were no age-dependent variations in vitamin D receptor protein expression. Results demonstrate that intestinal vitamin D receptor distribution patterns are segment-specific and strongest immunoreactions correlate with highest intestinal calcium absorptive activities, as reported in literature. Strong expression of vitamin D receptors within the lower half of crypts also suggests a role for calcitriol in epithelial differentiation and cellular homeostasis. J. Morphol., 2008. © 2007 Wiley-Liss, Inc. [source] Candidate's Thesis: Platelet-Activating Factor,Induced Hearing Loss: Mediated by Nitric Oxide?,THE LARYNGOSCOPE, Issue 12 2003Chung-Ku Rhee MD Abstract Objectives/Hypothesis Platelet-activating factor (PAF)in middle ear effusion is thought to induce hearing loss. The purpose of this study is to investigate the role of nitric oxide (NO) in the mechanism of PAF-induced hearing loss by studying the effects of PAF application on the round window membrane (RWM) with and without PAF-antagonist NO-blocker. Study Design Longitudinal study on randomized guinea pigs using PAF to induce hearing loss. Methods Guinea pigs were divided into four groups: PBS, PAF, PAF-antagonist, and L-NAME. The PBS group received phosphate buffered saline (PBS) and the PAF groups received 10, 20, and 40 ,g of PAF soaked into gelfoam and placed on the RWM. PAF-antagonist (WEB 2170) and NOS inhibitor NG-nitro-l-arginine-methylester (L-NAME) were injected intraperitoneally prior to PAF 20 ,g application on the RWM. The following three tests were performed on each animal group: Hearing was tested with an auditory brainstem response (ABR) test over 24 hours. At the end of 24 hours, cochlear hair cells were examined by scanning electron microscopy (SEM) and immunohistochemistry was carried out on the cochlea to test the expression of inducible nitric oxide synthase (iNOS). Results The PAF group developed significant elevation of ABR threshold and cochlear hair cell damage in the SEM group as compared with the PBS control group. The PAF-antagonist (WEB 2170) and the L-NAME groups did not show significant elevation of ABR threshold and cochlear hair cell damage compared with the group administered PAF 20 ,g, but in the PAF-antagonist group, the elevation of ABR threshold was significant compared with that of the PBS control group, whereas it was not significant compared with the PBS group in the L-NAME group. Strong expression of iNOS on cochlea was observed in the PAF group and lighter expression was seen in PBS, WEB 2170, and L-NAME groups. Conclusions This study demonstrated that PAF placed on the RWM induced hearing loss and cochlear hair cell damage. The PAF-antagonists and L-NAME prevented the PAF-induced hearing loss and inhibited iNOS expression in the cochlea. These findings suggest that the PAF-induced hearing loss caused by cochlear hair cell damage may have been mediated by NO. PAF-antagonists and L-NAME may have future therapeutic implications in preventing sensorineural hearing loss associated with chronic otitis media. The results of this study have significant potential clinical application. [source] The prognostic value of inducible nitric oxide synthase in local prostate cancerBJU INTERNATIONAL, Issue 3 2000S.H. Aaltomaa Objective To compare the clinical and histological data from patients with prostate cancer with the results of the immunohistochemical analysis of inducible nitric oxide synthase (iNOS), and thus determine the prognostic value of iNOS. Patients and methods The study included 82 patients (mean age 64.6 years, sd 6.1) with local prostate cancer treated by radical prostatectomy in two Finnish hospitals. Their mean ( sd) follow-up was 3.3 (2.2) years. An immunohistochemical method was used to detect the expression of iNOS in these specimens, and the expression graded according to staining intensity as none, weak or strong. Results There was weak or strong expression of iNOS in 25 (31%) and 56 (68%) of the patients, respectively, and one specimen was negative for iNOS. Strong expression of iNOS was related to high a preoperative prostate specific antigen (PSA) level (P = 0.006) and high pT classification (P < 0.001), but not to nodal status, grade, seminal vesicle or capsular invasion, surgical margin status, perineural infiltration, tumour infiltrating lymphocytes or proliferation rate of cancer cells. A PSA failure was detected in 29 patients but was not predicted by iNOS expression. A Cox multivariate analysis showed that surgical margin positivity, seminal vesicle involvement and number of tumour infiltrating lymphocytes predicted the PSA failure. Conclusion A high expression of iNOS was related to a high pT classification and the preoperative PSA level but not to other established prognostic factors; iNOS expression was not a predictor of PSA failure in patients with local prostate cancer. [source] Direct Stimulation of Adult Neural Stem Cells In Vitro and Neurogenesis In Vivo by Vascular Endothelial Growth FactorBRAIN PATHOLOGY, Issue 3 2004Anne Schänzer Hypoxia as well as global and focal ischemia are strong activators of neurogenesis in the adult mammalian central nervous system. Here we show that the hypoxia-inducible vascular endothelial growth factor (VEGF) and its receptor VEGFR-2/Flk-1 are expressed in clonally-derived adult rat neural stem cells in vitro. VEGF stimulated the expansion of neural stem cells whereas blockade of VEGFR-2/Flk-1-kinase activity reduced neural stem cell expansion. VEGF was also infused into the lateral ventricle to study changes in neurogenesis in the ventricle wall, olfactory bulb and hippocampus. Using a low dose (2.4 ng/d) to avoid endothelial proliferation and changes in vascular permeability, VEGF stimulated adult neurogenesis in vivo. After VEGF infusion, we observed reduced apoptosis but unaltered proliferation suggesting a survival promoting effect of VEGF in neural progenitor cells. Strong expression of VEGFR-2/Flk-1 was detected in the ventricle wall adjacent to the choroid plexus, a site of significant VEGF production, which suggests a paracrine function of endogenous VEGF on neural stem cells in vivo. We propose that VEGF acts as a trophic factor for neural stem cells in vitro and for sustained neurogenesis in the adult nervous system. These findings may have implications for the pathogenesis and therapy of neurodegenerative diseases. [source] Role of E-cadherins in development of lymphatic tumor emboliCANCER, Issue 9 2003Anita Gupta M.D. Abstract BACKGROUND E-cadherin (E-cad) is a cell adhesion molecule that is expressed in normal breast tissue. While loss of E-cad expression is a characteristic feature of lobular carcinoma, it also is observed in infiltrating ductal carcinoma (IDC). The presence of peritumoral intralymphatic emboli also is a poor prognostic feature in IDC. Invasive lobular carcinoma rarely is associated with intralymphatic emboli. In the current study, the authors assessed E-cad expression in cases of IDC with and without intralymphatic tumor emboli to examine the potential role played by these molecules in the development of lymphatic emboli. METHODS Fifty patients with high-grade invasive ductal carcinoma,25 with prominent lymphatic invasion (LVI) and intralymphatic tumor emboli and 25 without LVI,were tested for expression of E-cad. For both groups, the intensity and frequency of E-cad expression was evaluated in tumor cells and lymphatic emboli; normal lobules were used as internal controls. RESULTS Membranous expression of E-cad was observed in normal lobules and tumor cells in all patients, with the tumor cells exhibiting varying degrees of loss of expression. In the 25 LVI-positive patients, the majority of tumor cells (including intralymphatic emboli) expressed E-cad with an intensity and distribution similar to what was seen in normal lobules. In the LVI-negative patients, the intensity and the distribution of E-cad staining varied significantly. Tumor cells at the tumor-stroma interface showed a greater frequency and intensity of E-cad expression than did cells in the central region of the tumor. CONCLUSIONS Strong expression of E-cad was observed in LVI-positive patients with high-grade IDC but not in LVI-negative patients. Emboli also exhibited high-intensity expression. These findings, taken in conjunction with the knowledge that intralymphatic tumor emboli in lobular carcinoma (which is E-cad-negative) are rare, suggest that E-cad plays an important role in tumor development and growth within the lymphatics. Cancer 2003;97:2341,7. © 2003 American Cancer Society. DOI 10.1002/cncr.11332 [source] The expression of Scratch genes in the developing and adult brainDEVELOPMENTAL DYNAMICS, Issue 9 2006Faustino Marín Abstract The Scratch genes belong to the Snail superfamily of zinc-finger transcription factors present in the metazoa, represented in mammals by the Scratch1 and Scratch2 genes. We have analyzed the expression of these genes in the brain of mice at developmental stages between 9.5 days-post-coitum to adulthood. Both genes are expressed in the mantle layer of the neuroepithelium at mid-gestational stages in all regions except for the region corresponding to the V2 interneuron column, which lacked Scratch2 transcripts. From perinatal to adult stages, the expression patterns of the two genes differ. Scratch1 remains strongly expressed in almost all brain regions, although it is not found in some ventral structures such as motor nuclei and hypothalamic regions. In contrast, Scratch2 expression progressively diminishes and virtually no expression can be detected in the adult brain. Nevertheless, strong expression of Scratch2 is retained in the postnatal cortical subventricular zone, in the inner part of the cerebellar external granular layer, and in the glial cells of the adult vomeronasal nerve. Developmental Dynamoics 235:2586,2591, 2006. © 2006 Wiley-Liss, Inc. [source] Efficient and selective tumor cell lysis and induction of apoptosis in melanoma cells by a conditional replication-competent CD95L adenovirusEXPERIMENTAL DERMATOLOGY, Issue 8 2010Lothar F. Fecker Please cite this paper as: Efficient and selective tumor cell lysis and induction of apoptosis in melanoma cells by a conditional replication-competent CD95L adenovirus. Experimental Dermatology 2010; 19: e56,e66. Abstract:, The high mortality of melanoma demands the development of new strategies, and gene therapy may be considered provided improvements in efficacy and selectivity. Overexpression of the death ligand CD95L/FasL has been shown in previous studies as highly effective for apoptosis induction in melanoma cells. For efficient and selective targeting of melanoma, a conditional replication-competent adenoviral vector was constructed (Ad5-FFE-02), which drives CD95L expression by a tetracycline-inducible promoter. For restricting its replication to melanoma cells, the adenoviral E1A gene is controlled by a tyrosinase-derived promoter. Furthermore, adenoviral E1B was deleted and a mutated E1A was used to preferentially support replication in tumor cells. Proving its high selectivity and efficiency, strong expression of E1A and doxycycline-dependent induction of CD95L were characteristic for tyrosinase-positive melanoma cells after Ad5-FFE-02 transduction, whereas absent in non-melanoma cell lines. Importantly, Ad5-FFE-02-mediated cell lysis was restricted to melanoma cells, and induction of apoptosis was found only in tyrosinase and CD95 expressing cells. Finally, the combination of adenoviral replication and CD95L-mediated apoptosis resulted in an enhanced repression of melanoma cell growth. This new adenoviral vector may provide a basis for an efficient targeting of melanoma. [source] Expression pattern of somatostatin receptor subtypes 1,5 in human skin: an immunohistochemical study of healthy subjects and patients with psoriasis or atopic dermatitisEXPERIMENTAL DERMATOLOGY, Issue 12 2006Lena Hagströmer Abstract:, In psoriasis and atopic dermatitis, the inflammatory events have neurogenic components and the neuropeptides modify the functions of immuno-active cells in the skin. Somatostatin is a neuropeptide with several neuroendocrine and immunomodulating properties and mediates its actions by five distinct subtypes of G-protein-coupled receptors (SSTR1-5). This study describes the distribution of SSTR1,5, analysed with immunohistochemistry, in psoriasis, atopic dermatitis and controls. Normal human skin and lesional skin from patients with psoriasis or atopic dermatitis showed many similarities, but also some differences, as regards SSTR expression. SSTR1,3 were strongly expressed in the epidermis of healthy skin, and in the skin of patients with psoriasis or atopic dermatitis. It is noteworthy that SSTR4 and 5 were strongly expressed in the epidermis of psoriasis patients, but weakly expressed in the epidermis of those with atopic dermatitis and normal skin. The intensity of the staining also varied considerably between the different layers of the epidermis, especially in psoriasis patients. In all cases, the dendritic cells, found mostly in the papillary and upper reticular dermis, showed a strong expression of SSTR1,4, but a weak expression of SSTR5. SSTR1,5 were strongly expressed in the sweat glands in all skin biopsies. Hair follicles and sebaceous glands expressed all five subtypes. Striated muscle fibres showed an intense positive expression of SSTR1,4, but a weak or negative expression of SSTR5. The wide distribution and expression pattern of all five SSTRs in human skin suggest that somatostatin is involved in the interactions between the nervous system and the skin. [source] Activation and deactivation of periventricular white matter phagocytes during postnatal mouse developmentGLIA, Issue 1 2010Mariya Hristova Abstract Brain microglia are related to peripheral macrophages but undergo a highly specific process of regional maturation and differentiation inside the brain. Here, we examined this deactivation and morphological differentiation in cerebral cortex and periventricular subcortical white matter, the main "fountain of microglia" site, during postnatal mouse development, 0,28 days after birth (P0,P28). Only macrophages in subcortical white matter but not cortical microglia exhibited strong expression of typical activation markers alpha5, alpha6, alphaM, alphaX, and beta2 integrin subunits and B7.2 at any postnatal time point studied. White matter phagocyte activation was maximal at P0, decreased linearly over P3 and P7 and disappeared at P10. P7 white matter phagocytes also expressed high levels of IGF1 and MCSF, but not TNFalpha mRNA; this expression disappeared at P14. This process of deactivation followed the presence of ingested phagocytic material but correlated only moderately with ramification, and not with the extent of TUNEL+ death in neighboring cells, their ingestion or microglial proliferation. Intravenous fluosphere labeling revealed postnatal recruitment and transformation of circulating leukocytes into meningeal and perivascular macrophages as well as into ramified cortical microglia, but bypassing the white matter areas. In conclusion, this study describes strong and selective activation of postnatally resident phagocytes in the P0,P7 subcortical white matter, roughly equivalent to mid 3rd trimester human fetal development. This presence of highly active and IGF1- and MCSF-expressing phagocytes in the neighborhood of vulnerable white matter could play an important role in the genesis of or protection against axonal damage in the fetus and premature neonate. © 2009 Wiley-Liss, Inc. [source] Unique epithelial cell production of hepatocyte growth factor/scatter factor by putative precancerous intestinal metaplasias and associated "intestinal-type" biliary cancer chemically induced in rat liverHEPATOLOGY, Issue 6 2000Guan-Hua Lai Recently, we observed that Met, the receptor for hepatocyte growth factor/scatter factor (HGF/SF), is overexpressed in epithelial cells of both early-appearing intestinal metaplastic glands in precancerous hepatic cholangiofibrotic tissue and neoplastic glands in later developed intestinal-type of cholangiocarcinoma originated from the furan rat model of cholangiocarcinogenesis when compared with normal and hyperplastic intrahepatic biliary epithelia. We now show that HGF/SF is also aberrantly expressed in a manner closely paralleling that of its receptor in the neoplastic epithelial cells of furan-induced rat cholangiocarcinomas and in a majority of metaplastic epithelial cells within earlier formed precancerous hepatic cholangiofibrotic tissue. Using in situ hybridization and reverse transcription-polymerase chain reaction (RT-PCR), we further showed specific expression of HGF/SF messenger RNA (mRNA) in a novel rat cholangiocarcinoma epithelial cell line overexpressing Met. This cholangiocarcinoma cell line, termed C611B, was established from tumorigenic cells isolated from a furan-induced transplantable tumor. Moreover, we detected by in situ hybridization strong expression of HGF/SF mRNA transcripts in the cancerous epithelial glands of cholangiocarcinoma developed in recipient rats after in vivo cell transplantation of C611B cells. In contrast, mRNA transcripts and protein immunoreactivity for this cytokine were not detected in hepatocytes and biliary epithelial cells in adult normal rat liver nor in rat hyperplastic intrahepatic biliary epithelium. Our results clearly show that HGF/SF becomes aberrantly expressed in cholangiocarcinoma epithelium and in putative precancerous intestinal metaplastic epithelium induced in the liver of furan-treated rats. [source] Chromogenic in situ hybridization for Her-2/neu-oncogene in breast cancer: comparison of a new dual-colour chromogenic in situ hybridization with immunohistochemistry and fluorescence in situ hybridizationHISTOPATHOLOGY, Issue 6 2009Doris Mayr Aims:, Her-2/neu testing is used as a marker for Herceptin® therapy. The aim was to investigate new dual-colour chromogenic in situ hybridization (CISH), in a large number of breast carcinomas (n = 205) with DNA-specific dual-colour probes (ZytoVision, Bremerhaven, Germany) and to compare the results with immunohistochemistry (n = 205) and fluorescence in situ hybridization (FISH) (n = 129). Methods and results:, Paraffin-embedded tissue of 205 patients was used. After immunohistochemistry with a focus on immunohistochemically uncertain cases, Her-2/neu amplification using dual-colour CISH (ZytoVision®) was analysed. Validation by FISH was performed. The results were: immunohistochemistry, 27.8% with strong expression, 53.7% with uncertain overexpression and 18.5% with no expression; FISH, 25.6% amplified and 74.4% negative; CISH, 35.6% amplified, 62.9% negative and 1.5% not evaluable. Comparison of immunohistochemistry with CISH: CISH negative in 100% with immunohistochemistry 0/1+, amplified in 82.5% with immunohistochemistry 3+; 5.9% contradictory results: 4.4% immunohistochemistry 3+ and negative by CISH, 1.5% negative in immunohistochemistry but amplified by CISH; FISH (129 cases), 8.5% contradictory results to immunohistochemistry, 6.2% immunohistochemistry 3+ and negative by FISH, 2.3% negative by immunohistochemistry and amplified by FISH; comparison of CISH and FISH, 94.6% same results, 3.9% different ones, 1.6% CISH not analysable. Conclusions:, CISH, using dual-colour probes (ZytoVision®) is as good as FISH for Her-2/neu analysis. The few discrepant results are likely to be caused by polysomy or tumour heterogeneity. [source] Stromal cells promote bone invasion by suppressing bone formation in ameloblastomaHISTOPATHOLOGY, Issue 4 2008G S A Sathi Aims:, To study the stromal variation and role of stromal,tumour cell interaction in impaired bone formation as well as enhanced bone resorption in ameloblastoma. Methods and results:, Four types of stroma were observed histologically; fibrous, desmoplastic, myxoid and myxoid with hyalinization. Osteoblast and osteoclast were counted using haematoxylin and eosin sections and immunohistochemistry with CD68. After histomorphometric analysis, only fibrous and myxoid types of stroma were distinctly identified. Secreted frizzled-related peptide (sFRP)-2, transforming growth factor-beta 1 and receptor activator of nuclear factor-,B ligand (RANKL) revealed strong expression in myxoid type compared with the normal stroma. Bone morphogenetic protein (BMP)-2 was negative in myxoid type, but positive in normal stroma. Fibrous-type stroma showed weak expression of all antigens except RANKL compared with myxoid type. Conclusions:, The results suggest that stroma does not act only in bone resorption, but also in the suppression of new bone formation. sFRP-2 is the main factor for impaired bone formation. The expression of markers related to osteoclastogenesis and suppression of osteoblast formation is higher in myxoid-type than in fibrous-type stroma. Tumour cells create a favourable environment for impaired bone formation by secreting sFRP-2 as well as bone resorption by secreting RANKL and interleukin-6. [source] Validation of novel promoter sequences derived from two endogenous ubiquitin genes in transgenic Aedes aegyptiINSECT MOLECULAR BIOLOGY, Issue 4 2010M. A. E. Anderson Abstract To date, only a limited number of promoter sequences have been described to drive transgene expression in the disease vector Aedes aegypti. We sought to increase this repertoire by characterizing the ability of upstream sequences derived from the Ae. aegypti UbL40 and polyubiquitin genes to drive the expression of marker proteins. Both genomic fragments were able to drive robust expression of luciferase in cultured mosquito cells. Following Mos1-transformation, the UbL40 promoter drove strong expression of a fluorescent marker in early larvae and in ovaries, while the polyubiquitin promoter drove robust EGFP expression in all stages of development, including constitutive expression throughout the midgut. These promoter fragments provide two new expression profiles for future Ae. aegypti genetic experiments. [source] Immunolocalization of bone extracellular matrix proteins (type I collagen, osteonectin and bone sialoprotein) in human dental pulp and cultured pulp cellsINTERNATIONAL ENDODONTIC JOURNAL, Issue 6 2003J. M. Q. Garcia Abstract Aim, To simultaneously analyse the expression of type I collagen, osteonectin and bone sialoprotein (BSP) in human dental pulp of different ages. Methodology, Cultured dental pulp fibroblasts (FP1 cell line), pulps from dental germs with incomplete root formation (n = 4) and pulps of erupted teeth with total root formation (n = 4) were used. Bone proteins were searched by immunohistochemistry and immunofluorescence using polyclonal antibodies and compared among the three groups assessed. Results, Immunohistochemistry detected the three proteins in dental pulp tissue, as it labelled extracellular matrix, predentine and odontoblasts. The BSP label was weaker, when compared to both type I collagen and osteonectin. The presence of type I collagen was more evident in pulps from erupted teeth, when compared to germ dental pulps. On the other hand, a strong expression of osteonectin in germ dental pulps was observed. Conclusions, Regardless of the degree of maturation, dental pulps present type I collagen, osteonectin and BSP in the extracellular matrix (ECM) and in the odontoblastic layer. Thus, the results suggest that these proteins are related to the production and mineralization of dentine. [source] Mammalian target of rapamycin is activated in human gastric cancer and serves as a target for therapy in an experimental modelINTERNATIONAL JOURNAL OF CANCER, Issue 8 2007Sven A. Lang Abstract The mammalian target of rapamycin (mTOR) has become an interesting target for cancer therapy through its influence on oncogenic signals, which involve phosphatidylinositol-3-kinase and hypoxia-inducible factor-1, (HIF-1,). Since mTOR is an upstream regulator of HIF-1,, a key mediator of gastric cancer growth and angiogenesis, we investigated mTOR activation in human gastric adenocarcinoma specimens and determined whether rapamycin could inhibit gastric cancer growth in mice. Expression of phospho-mTOR was assessed by immunohistochemical analyses of human tissues. For in vitro studies, human gastric cancer cell lines were used to determine S6K1, 4E-BP-1 and HIF-1, activation and cancer cell motility upon rapamycin treatment. Effects of rapamycin on tumor growth and angiogenesis in vivo were assessed in both a subcutaneous tumor model and in an experimental model with orthotopically grown tumors. Mice received either rapamycin (0.5 mg/kg/day or 1.5 mg/kg/day) or diluent per intra-peritoneal injections. In addition, antiangiogenic effects were monitored in vivo using a dorsal-skin-fold chamber model. Immunohistochemical analyses showed strong expression of phospho-mTOR in 60% of intestinal- and 64% of diffuse-type human gastric adenocarcinomas. In vitro, rapamycin-treatment effectively blocked S6K1, 4E-BP-1 and HIF-1, activation, and significantly impaired tumor cell migration. In vivo, rapamycin-treatment led to significant inhibition of subcutaneous tumor growth, decreased CD31-positive vessel area and reduced tumor cell proliferation. Similar significant results were obtained in an orthotopic model of gastric cancer. In the dorsal-skin-fold chamber model, rapamycin-treatment significantly inhibited tumor vascularization in vivo. In conclusion, mTOR is frequently activated in human gastric cancer and represents a promising new molecular target for therapy. © 2007 Wiley-Liss, Inc. [source] What is a role of haeme oxygenase-1 in psoriasis?INTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 2 2007Current concepts of pathogenesis Summary The skin is constantly exposed to endogenous and environmental pro-oxidant agents, which lead to harmful generation of reactive oxygen species (ROS). Healthy skin, being a potential target for oxidative stress, is equipped with a large number of defence mechanisms including antioxidant systems. This protection can be corrupted by an imbalance between ROS and antioxidants with pathological level of oxidants prevailing. There is a great body of evidence indicating that some inflammatory skin diseases, such as psoriasis, are mediated by oxidative stress. Keratinocytes of normal skin, the primary target for pro-oxidant agents, show strong expression of ROS-detoxifying enzymes. In addition, normal keratinocytes express haeme oxygenase (HO), an enzyme which might be involved in the protection of cells against oxidative stress. HO (inducible HO-1, constitutive HO-2 and HO-3) is the rate-limiting enzyme in haeme catabolism, which leads to the generation of biliverdin, iron, and carbon monoxide. HO-1 is a stress-responsive protein whose expression is induced by various oxidative agents. HO-1 is known for its cytoprotective, antioxidant and anti-inflammatory properties. Interestingly, a strong overexpression of HO-1 was observed in psoriatic skin. However, the role of HO-1 in psoriasis remains unclear. In this review, we will discuss some current concepts concerning pathogenesis of psoriasis and the contribution of HO-1 in skin inflammation to show the relationships between HO-1, ROS and cytokine network in psoriatic skin. We will try to answer a question whether enhanced HO-1 expression in keratinocytes results in beneficial or detrimental effect on the development and severity of psoriatic lesions. [source] Gap junctional communication in human osteoclasts in vitro and in vivoJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 6a 2008A. F. Schilling Abstract Bone-forming cells are known to be coupled by gap junctions, formed primarily by connexin43 (Cx43). The role of Cx43 in osteoclasts has so far only been studied in rodents, where Cx43 is important for fusion of mononuclear precursors to osteoclasts. Given the potential importance for human diseases with pathologically altered osteoclasts, we asked whether a similar influence of Cx43 can also be observed in osteoclasts of human origin. For this purpose, Cx43 mRNA expression was studied in a time course experiment of human osteoclast differentiation by RT-PCR. Localization of Cx43 in these cells was determined by immunohistochemistry and confocal microscopy. For the assessment of the effect of gap junction inhibition on cell fusion, gap junctions were blocked with heptanol during differentiation of the cells and the cells were then evaluated for multinuclearity. Paraffin sections of healthy bone and bone from patients with Paget's disease and giant cell tumour of the bone were used to study Cx43 expression in vivo. We found mRNA and protein expression of Cx43 in fully differentiated osteoclasts as well as in precursor cells. This expression decreased in the course of differentiation. Consistently, we found a lower expression of Cx43 in osteoclasts than in bone marrow precursor cells in the histology of healthy human bone. Blockade of gap junctional communication by heptanol led to a dose-dependent decrease in multinuclearity, suggesting that gap junctional communication precedes cell fusion of human osteoclasts. Indeed, we found a particularly strong expression of Cx43 in the giant osteoclasts of patients with Paget's disease and giant cell tumour of the bone. These results show that gap junctional communication is important for fusion of human mononuclear precursor cells to osteoclasts and that gap junctional Cx43 might play a role in the regulation of size and multinuclearity of human osteoclasts in vivo. [source] Expression of cytokeratin subtypes in intraepidermal malignancies: a guide for differentiationJOURNAL OF CUTANEOUS PATHOLOGY, Issue 8 2006Figen Aslan Background:, Among intraepidermal malignancies of epithelial origin, Bowen's disease, bowenoid actinic keratosis (BAK), intraepidermal malignant eccrine poroma (MEP), and Paget's disease may pose diagnostic difficulties. Methods:, Histologic features and immunohistochemical profiles of 24 cases of Bowen's disease, 21 cases of BAK, 18 cases of intraepidermal MEP, and 11 cases of Paget's disease were analyzed. Results:, Using multivariate logistic regression test, multinuclear giant cells and solar degeneration were found to be the only histologic parameters of diagnostic help. On the other hand, a widespread positive reaction for CK 5/8, CK 7, CK 19, and negative reaction for CK 10, was a helpful feature in the differentiation of Paget's disease from Bowen's disease and BAK. The widespread and strong expression of CK 10 was seen in almost all cases of Bowen's disease in contrast to BAK. The widespread expression of CK 5/8 and CK 7, and negative reaction for CK 10, was in favor of Paget's disease, compared to intraepidermal MEP. On the other hand, widespread expression of CK 19 was a common finding in intraepidermal MEP, in contrast to Bowen's disease. Conclusion:, An immunohistochemical panel may provide significant hints on the differentiation of common intraepidermal malignancies, especially in problematic cases. [source] Enhanced type I interferon signaling and recruitment of chemokine receptor CXCR3-expressing lymphocytes into the skin following treatment with the TLR7-agonist imiquimodJOURNAL OF CUTANEOUS PATHOLOGY, Issue 4 2005Joerg Wenzel Introduction:, Imiquimod (AldaraÔ) is an immune response modifier approved for the topical treatment of external genital and perianal warts which can mediate regression of several cutaneous malignancies [basal cell carcinoma (BCC), Bowen's disease, actinic keratosis, and metastasis of malignant melanoma]. Recently, it was discovered that imiquimod acts through the toll-like receptor (TLR) 7. We hypothesize that TLR7-signaling strongly induces the production of interferon (IFN) ,, which is able to enhance Th1-mediated cellular antiviral and antitumor immunity. Patients and methods:, In the present study we analyzed the expression of MxA, a protein specifically induced by type I IFNs during topical imiquimod treatment in several patients suffering from different cutaneous malignancies (BCC, cutaneous metastasis of melanoma, and breast cancer), and characterized the inflammatory infiltrate, along with the expression of chemokine receptor CXCR3, by immunohistochemistry. Results:, Treatment with the TLR7-agonist imiquimod induced a significant lesional lymphocytic inflammation, associated with strong expression of MxA, indicating the induction of type I IFN signaling. The extent of lesional MxA staining closely correlated with the number of infiltrating T lymphocytes and the expression of the chemokine receptor CXCR3, characteristic for Th1-biased immune responses. Discussion:, Our in vivo results suggest an important role for TLR7-induced production of type I IFN, which links innate and adaptive immunity and promotes specific Th1-biased cellular immune response capable of eliminating cutaneous malignancies. MxA appears to be a valuable parameter to demonstrate IFN-type I expression in imiquimod therapy. [source] Cyclo-oxygenase-2 inhibitors suppress epithelial cell kinetics and delay gastric wound healing in ratsJOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY, Issue 7 2000Wei-Hao Sun Abstract Background and Aims: The present study examined the effects of NS-398, a specific cyclo-oxygenase-2 inhibitor, on gastric mucosal cell kinetics and gastric wound healing following acid-induced injury. Methods: Male Sprague-Dawley rats were fasted for 24 h and then 0.6 mol/L hydrochloric acid (HCl; 1 mL) was administered into the stomach; NS-398 or indomethacin was administered to the animals 10 min after the acid. Levels of constitutive cyclo-oxygenase (COX-1) and mitogen-inducible cyclo-oxygenase (COX-2) in the gastric mucosa were analysed using western blotting and immunohistochemical staining. The grade of the lesion was assessed using planimetry and histological examination, including immunohistochemistry for proliferating cell nuclear antigen (PCNA). Results: Although there was strong expression of COX-1, there was minimal expression of COX-2 in the gastric mucosa. Expression of COX-2 was enhanced mainly in surface epithelial cells and neck cells following HCl administration. Gastric mucosal ulcers and erosions healed within 48 h, during which time the proliferative zone expanded in the control animals. Indomethacin and NS-398 suppressed the expansion of the proliferative zone and delayed the healing of the gastric injury. Conclusion: The present study demonstrated that cyclo-oxygenase-2 inhibitors delay gastric wound healing by suppressing expansion of the mucosal proliferative zone. These results provide evidence that cyclo-oxygenase-2 has an important role in gastric mucosal regeneration. [source] Developmental change and function of chondroitin sulfate deposited around cerebellar Purkinje cellsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 2 2005Yumiko Shimazaki Abstract Chondroitin sulfate is a long sulfated polysaccharide with enormous structural heterogeneity that binds with various proteins, such as growth factors, in a structure-dependent manner. In this study, we analyzed the expression of chondroitin sulfate in the postnatally developing cerebellar cortex by using three monoclonal antibodies against chondroitin sulfate, MO-225, 2H6, and CS-56, which recognize different structural domains in this polysaccharide. During the first postnatal week, the patterns of immunohistochemical staining made by these antibodies were quite similar, and the molecular layer, the granule cell layer, and Bergmann glial fibers in the external granular layer were densely stained. After postnatal day 12 (P12), the expression of 2H6 epitopes was down-regulated in the molecular layer, and the expression of CS-56 epitopes in this layer was also reduced after P16. On the other hand, the strong expression of MO-225 epitopes, GlcA(2S),1,3GalNAc(6S) (D unit)-containing structures, remained until adulthood. These chondroitin sulfate epitopes were observed around Purkinje cells, including cell soma and dendrites. Detailed immunohistochemical analysis suggested that chondroitin sulfate was deposited between Purkinje cell surfaces and the processes of Bergmann glia. Furthermore, the amount of pleiotrophin, a heparin-binding growth factor, in the cultured cerebellar slices was remarkably diminished after treatment with chondroitinase ABC or D unit-rich chondroitin sulfate. With the previous findings that pleiotrophin binds to D unit-rich chondroitin sulfate, we suggest that the D-type structure is important for the signaling of pleiotrophin, which plays roles in Purkinje cell,Bergmann glia interaction, and that the structural changes of chondroitin sulfate regulate this signaling pathway. © 2005 Wiley-Liss, Inc. [source] Direct cell,cell interactions control apoptosis and oligodendrocyte marker expression of neuroepithelial cellsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 3 2001J.P. Hugnot Abstract During brain development, the neuroepithelium generates neurons and glial cells. Proliferation and differentiation of neuroepithelial cells are controlled by a complex combination of secreted factors and more intrinsic or local mechanisms, such as lateral inhibition and asymmetric division. To obtain further insights into the signals governing neuroepithelial cell fate, we used the immortomouse to derive conditionally immortalised cell lines from mouse E10 neuroepithelium. We isolated a nestin-positive basic fibroblast growth factor (bFGF)-responsive cell line (SVE10-23) which mostly differentiate into astrocytes when cocultured with primary cortical cells. We found that, by simply lowering the cell density, SVE10-23 cells embarked on oligodendrocytic differentiation as indicated by the strong expression of galactocerebroside C and 2,3,-cyclic nucleotide 3,-phosphodiesterase. Apoptosis accompanied the differentiation, and all cells died within 1 week. We present here evidence that direct interactions between cells are the main mechanism regulating this oligodendrocytic differentiation. We demonstrate that SVE10-23 cells contact or proximity inhibit their differentiation, prevent apoptosis, and promote their proliferation. Similarly, others nestin-positive precursor cell lines and nonimmortalised bFGF-grown E10 cells were found to spontaneously differentiate at low density, thus generalising the idea that neural precursor fate is regulated by direct cell,cell interactions. The SVE10-23 cell line provides a valuable tool with which to study further the molecular components implicated in this mode of regulation. J. Neurosci. Res. 65:195,207, 2001. © 2001 Wiley-Liss, Inc. [source] ,-TUBULIN GENE OF PORPHYRA PURPUREA ( RHODOPHYTA),JOURNAL OF PHYCOLOGY, Issue 5 2002Ron M. MacKay The life cycle of the marine red alga Porphyra purpurea (Roth) C. Agardh includes a shell-boring filamentous sporophyte and a leafy gametophyte. A single intronless gene for the microtubule protein ,-tubulin was discovered by molecular cloning of P. purpurea cDNA and genomic DNA. This gene, named TubB1, encodes a ,-tubulin with a divergent amino acid sequence, showing 74% identity with the conserved ,-tubulin of Chlamydomonas reinhardtii P. A. Dangeard. Southern hybridization analysis of nuclear DNA confirmed that P. purpurea has a single TubB1 gene. Transcripts (1.8 kb) of TubB1 are present in the sporophyte and gametophyte. Codon bias indicates strong expression of TubB1. The divergent nature of the TubB1 genes suggests that the absence of axonemal structures has allowed substantial genetic drift in red algal ,-tubulin genes. [source] Enhanced B7 Costimulatory Molecule Expression In Inflammatory Human Sural Nerve BiopsiesJOURNAL OF THE PERIPHERAL NERVOUS SYSTEM, Issue 1 2001R Kiefer Objectives-To define the role of the costimulatory molecules B7-1 and B7-2 in inflammatory disorders of the peripheral nervous system. B7 molecules are essential for effective antigen presentation and may determine the differentiation of T cells into a Th-1 or Th-2 phenotype, thus modulating immune response and disease course. Methods-Forty nine sural nerve biopsies from patients with neuroborreliosis, Guillain-Barre syndrome (GBS), chronic inflammatory demyelinating polyneuropathy (CIDP), CIDP variants and hereditary neuropathies, and those with no detectable abnormality were investigated. The expression of B7-1 and B7-2 mRNA and protein was investigated by polymerase chain reaction (PCR) and immunocytochemistry. Results-B7-1 mRNA was strongly upregulated in both cases of neuroborreliosis, in two cases of GBS and one case of variant CIDP. Moderate to low levels were detected in the remaining GBS and CIDP biopsies and were rarely found in a noninflammatory control group consisting of hereditary neuropathy and normal nerves. At the immunocytochemical level, strong expression of B7-1 protein was found in both neuroborreliosis cases, and moderate or low expression in six of eight GBS cases and seven of 17 CIDP cases investigated, whereas only one of five non-inflammatory control nerves showed staining, which was very weak. In neuroborreliosis, B7-1 protein was found very pronounced in epineurial infiltrates, whereas in CBS and CIDP, labelling was predominantly endoneurial and localised to putative macrophages. B7-2 mRNA and protein were expressed only at low levels in neuroborreliosis and selected autoimmune neuropathy cases, and were essentially absent from noninflammatory controls. Conclusions-B7 molecules are expressed in the peripheral nervous system and regulated during disease, and their presence in macrophages underlines the putative function of endoneurial macrophages as local antigen presenting cells in the immunopathology of peripheral nerve. B7-1 rather than B7-2 is preferentially upregulated, possibly promoting the induction of a Th-1-type T cell response within the nerve. [source] Plasma levels and skin-eosinophil-expression of vascular endothelial growth factor in patients with chronic urticariaALLERGY, Issue 11 2009A. Tedeschi Background:, Although chronic urticaria (CU) is often regarded as autoimmune in nature, only less than 50% of sera from CU patients contain histamine-releasing autoantibodies. This suggests that other factors may contribute to its pathogenesis. We evaluated the possible involvement of vascular endothelial growth factor (VEGF), one of the major mediators of vascular permeability, in CU. Methods:, Eighty consecutive adult patients with CU and 53 healthy subjects were studied. VEGF and prothrombin fragment F1+2 were measured by enzyme immunoassays. Autologous plasma skin test (APST) was performed in CU patients and, in six of them, skin biopsy specimens were taken from wheals to evaluate the immunohistochemical expression of VEGF and eosinophil cationic protein (ECP). Results:, Plasma VEGF concentrations were higher in CU patients (8.00 ± 0.90 pmol/l) than in controls (0.54 ± 0.08 pmol/l) (P = 0.0001) and tended to parallel both the severity of CU and to correlate with F1+2 levels. APST was positive in 85.1% of patients. VEGF concentration was significantly higher in APST-positive than in APST-negative patients (P = 0.0003). Immunohistochemically, all specimens from patients with CU showed a strong expression of VEGF (P = 0.002) that colocalized with ECP, a classic eosinophil marker. Conclusions:, VEGF plasma levels are elevated in CU and parallel the disease severity. This supports a possible role of this molecule in CU pathophysiology. Eosinophils are the main cellular source of VEGF in CU lesional skin. [source] Studies on the mechanism of resistance to Bipolaris sorokiniana in the barley lesion mimic mutant bst1MOLECULAR PLANT PATHOLOGY, Issue 5 2009MATTIAS PERSSON SUMMARY The Bipolaris sorokiniana tolerant 1 (bst1) barley mutant is derived from fast neutron-irradiated seeds of wild-type Bowman(Rph3). The induced mutation was genetically localized to a position on chromosome 5HL distal to the centromere using amplified fragment length polymorphism markers. In addition, the defence responses and related gene expression in the bst1 mutant after fungal challenge were compared with those occurring in wild-type plants. Hydrogen peroxide generation, determined by 3,3-diaminobenzidine staining, revealed a clearly reduced level of bst1, compared with the wild-type, during the entire experimental time: 8,120 h post-inoculation (hpi). At 48 hpi, the wild-type samples displayed twice as much fungal mass and three times greater H2O2 production than bst1. At the same time, staining of B. sorokiniana showed less fungal growth in the spontaneous lesions of bst1 compared with the wild-type. Monitoring of defence-related genes at 48 hpi demonstrated strong expression of PR-1a, PR-2, PR-5 and PR-10 in bst1. A gene coding for a unique oxidoreductase enzyme, designated as HCP1, was expressed at much higher levels in inoculated leaves of the bst1 mutant than in those of the wild-type plant. Taken together, the results suggest that the defence to B. sorokiniana largely relies on salicylic acid-responsive pathogenesis-related (PR) genes, as well as selected reactive oxygen species and unknown HCP1 -associated factors. [source] | ||||||||||||||||||||||||||||