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Sample records for activation marker expression

  1. Expression of intercellular adhesion molecule-1 on macrophages in vitro as a marker of activation.

    PubMed

    Bernatchez, S F; Atkinson, M R; Parks, P J

    1997-10-01

    Macrophage activation is a major component of wound healing. It also determines the extent of inflammatory reactions and the response of the body to implanted materials. We have previously shown, using an in vitro model, that the extent of spreading of macrophages on different materials is a marker of activation, and that a soluble inducer has a dose-response effect on the secretion of cytokines in the culture medium. This work investigates the expression of three different cell surface markers [macrophages MAC-1, MAC-3 and intercellular adhesion molecule-1 (ICAM-1)] on macrophages in vitro using confocal microscopy and shows that ICAM-1 is also a marker of macrophage activation in this model. We observed increased amounts of ICAM-1 on activated macrophages compared to unactivated macrophages, whereas MAC-1 and MAC-3 were either expressed constitutively or demonstrated no quantitative change in expression after activation under the same experimental conditions. We also tested the expression of ICAM-1 with various concentrations of soluble inducers (lipopolysaccharide, 0.001, 0.01, 0.1, 1 and 10 micrograms ml-1. S-27609, 0.1, 0.25, 0.5, 1, 2 and 3 micrograms ml-1 and on a sheet of polylactic acid alone or in combination with soluble inducers. All doses of soluble inducers induced the expression of ICAM-1 on cells grown in glass chamber slides. The induction was not dose related but seemed to work rather in an on-off manner. There was no effect of material on ICAM-1 expression on the cell surface when no soluble inducer was added. This was similar to cytokine secretion, which was not induced by our material alone. When either lipopolysaccharide or S-27609 was used in combination with the material, there was an increase in the average measured intensity of ICAM-1. In this in vitro model, ICAM-1 staining as measured by confocal microscopy is a marker for macrophage activation. Our results suggest that the extent of macrophage activation as measured by ICAM-1 and by

  2. Expression of T lymphocyte chemoattractants and activation markers in vernal keratoconjunctivitis

    PubMed Central

    El-Asrar, A M Abu; Struyf, S; Al-Kharashi, S A; Missotten, L; Van Damme, J; Geboes, K

    2002-01-01

    Background/aims: T lymphocytes are present in increased numbers in the conjunctiva of patients with vernal keratoconjunctivitis (VKC) and their activation has a central role in the pathogenesis of the chronic allergic inflammatory reactions seen in VKC. The aims of this study were to examine the expression of three recently described potent T lymphocyte chemoattractants, PARC (pulmonary and activation regulated chemokine), macrophage derived chemokine (MDC), and I-309, the MDC receptor CCR4, and T lymphocyte activation markers, CD25, CD26, CD62L, CD71, and CD30, and to correlate them with the counts of CD3+ T lymphocytes in the conjunctiva of patients with VKC. Method: Conjunctival biopsy specimens from 11 patients with active VKC, and eight control subjects were studied by immunohistochemical techniques using a panel of monoclonal and polyclonal antibodies directed against PARC, MDC, I-309, CCR4, CD25, CD26, CD62L, CD71, and CD30. The numbers of positively stained cells were counted. The phenotype of inflammatory cells expressing chemokines was examined by double immunohistochemistry. Results: In the normal conjunctiva, vascular endothelial cells in the upper substantia propria showed weak immunoreactivity for CD26. There was no immunoreactivity for the other antibodies. VKC specimens showed inflammatory cells expressing PARC, MDC, and I-309. The numbers of PARC+ inflammatory cells were higher than the numbers of MDC+ and I-309+ inflammatory cells and the mean values of the three groups differed significantly (17.0 (SD 10.1); 9.5 (9.9), and 4.3 (7.9), respectively, p = 0.0117, ANOVA). The numbers of PARC+ inflammatory cells had the strongest correlation with the numbers of CD3+ T lymphocytes. Few CCR4+ inflammatory cells were observed in only three specimens. Double immunohistochemistry revealed that all inflammatory cells expressing chemokines were CD68+ monocytes/macrophages. The numbers of CD25+ T lymphocytes were higher than the numbers of CD26+, CD62L+, CD71

  3. Activated peripheral lymphocytes with increased expression of cell adhesion molecules and cytotoxic markers are associated with dengue fever disease.

    PubMed

    Azeredo, Elzinandes L; Zagne, Sonia M O; Alvarenga, Allan R; Nogueira, Rita M R; Kubelka, Claire F; de Oliveira-Pinto, Luzia M

    2006-06-01

    The immune mechanisms involved in dengue fever and dengue hemorrhagic/dengue shock syndrome are not well understood. The ex vivo activation status of immune cells during the dengue disease in patients was examined. CD4 and CD8 T cells were reduced during the acute phase. Interestingly, CD8 T cells co-expressing activation marker HLA-DR, Q, P, and cytolytic granule protein-Tia-1 were significantly higher in dengue patients than in controls. Detection of adhesion molecules indicated that in dengue patients the majority of T cells (CD4 and CD8) express the activation/memory phenotype, characterized as CD44HIGH and lack the expression of the naïve cell marker, CD62L LOW. Also, the levels of T cells co-expressing ICAM-1 (CD54), VLA-4, and LFA-1 (CD11a) were significantly increased. CD8 T lymphocytes expressed predominantly low levels of anti-apoptotic molecule Bcl-2 in the acute phase, possibly leading to the exhibition of a phenotype of activated/effector cells. Circulating levels of IL-18, TGF-b1 and sICAM-1 were significantly elevated in dengue patients. Early activation events occur during acute dengue infection which might contribute to viral clearance. Differences in expression of adhesion molecules among CD4 and CD8 T cells might underlie the selective extravasation of these subsets from blood circulation into lymphoid organs and/or tissues. In addition, activated CD8 T cells would be more susceptible to apoptosis as shown by the alteration in Bcl-2 expression. Cytokines such as IL-18, TGF-b1, and sICAM-1 may be contributing by either stimulating or suppressing the adaptative immune response, during dengue infection, thereby perhaps establishing a relationship with disease severity.

  4. Expression of Early Activation Marker CD69 on Peripheral Blood Lymphocytes from Pregnant Women after First Trimester Alloimmunization.

    PubMed

    Krechetova, L V; Vtorushina, V V; Nikolaeva, M A; Golubeva, E L; Van'ko, L V; Saribegova, V A; Tetruashvili, N K

    2016-08-01

    We studied the expression of an early activation marker CD69 in peripheral blood lymphocytes of pregnant women with a history of recurrent pregnancy loss after immunization with paternal lymphocytes. Spontaneous and phytohemagglutinin-stimulated expression of CD69 on the surface of T cells and NK cells isolated from the peripheral blood was analyzed. On gestation week 5-6, the number of T cells expressing CD69 spontaneously and after stimulation was significantly higher in women with miscarriage than in woman with prolonged pregnancy. However, the number of cells with CD56(+) phenotype expressing CD69 did not differ in these groups. No differences were found in the number of cells of all subpopulations expressing CD69 after stimulation on gestation week 12 in woman with full-term current pregnancy and in woman with physiological pregnancy. PMID:27591871

  5. [Analysis of membrane expression of the CD63 human basophil activation marker. Applications to allergologic diagnosis].

    PubMed

    Sainte-Laudy, J; Vallon, C; Guérin, J C

    1994-06-01

    On the basis of the CD 63 bi-modal expression on the membrane of activated basophils, we set up a flow cytometric method for the analysis of human basophils activation by an anti-IgE and anti-CD 63 double labelling. We demonstrated that the statistical characteristics of the percentages of activation obtained by an anti-IgE stimulation allowed the use of this method for pharmacological studies. The percentages of activation were of the same order of magnitude than those obtained by histamine release. CD 63 expression was also observed for a low affinity allergen such as the sulfonyl-HSA conjugate used for sulfites hypersensibility diagnosis, healthy donors being negative. This method, which can be automatized may represent an interesting candidate in the field of hapten hypersensitivity which lacks of reliable diagnostical methods. PMID:7524523

  6. Myofibroblasts are distinguished from activated skin fibroblasts by the expression of AOC3 and other associated markers.

    PubMed

    Hsia, Lin-Ting; Ashley, Neil; Ouaret, Djamila; Wang, Lai Mun; Wilding, Jennifer; Bodmer, Walter F

    2016-04-12

    Pericryptal myofibroblasts in the colon and rectum play an important role in regulating the normal colorectal stem cell niche and facilitating tumor progression. Myofibroblasts previously have been distinguished from normal fibroblasts mostly by the expression of α smooth muscle actin (αSMA). We now have identified AOC3 (amine oxidase, copper containing 3), a surface monoamine oxidase, as a new marker of myofibroblasts by showing that it is the target protein of the myofibroblast-reacting mAb PR2D3. The normal and tumor tissue distribution and the cell line reactivity of AOC3 match that expected for myofibroblasts. We have shown that the surface expression of AOC3 is sensitive to digestion by trypsin and collagenase and that anti-AOC3 antibodies can be used for FACS sorting of myofibroblasts obtained by nonenzymatic procedures. Whole-genome microarray mRNA-expression profiles of myofibroblasts and skin fibroblasts revealed four additional genes that are significantly differentially expressed in these two cell types: NKX2-3 and LRRC17 in myofibroblasts and SHOX2 and TBX5 in skin fibroblasts. TGFβ substantially down-regulated AOC3 expression in myofibroblasts but in skin fibroblasts it dramatically increased the expression of αSMA. A knockdown of NKX2-3 in myofibroblasts caused a decrease of myofibroblast-related gene expression and increased expression of the fibroblast-associated gene SHOX2, suggesting that NKX2-3 is a key mediator for maintaining myofibroblast characteristics. Our results show that colorectal myofibroblasts, as defined by the expression of AOC3, NKX2-3, and other markers, are a distinctly different cell type from TGFβ-activated fibroblasts. PMID:27036009

  7. Myofibroblasts are distinguished from activated skin fibroblasts by the expression of AOC3 and other associated markers

    PubMed Central

    Hsia, Lin-ting; Ashley, Neil; Ouaret, Djamila; Wang, Lai Mun; Wilding, Jennifer; Bodmer, Walter F.

    2016-01-01

    Pericryptal myofibroblasts in the colon and rectum play an important role in regulating the normal colorectal stem cell niche and facilitating tumor progression. Myofibroblasts previously have been distinguished from normal fibroblasts mostly by the expression of α smooth muscle actin (αSMA). We now have identified AOC3 (amine oxidase, copper containing 3), a surface monoamine oxidase, as a new marker of myofibroblasts by showing that it is the target protein of the myofibroblast-reacting mAb PR2D3. The normal and tumor tissue distribution and the cell line reactivity of AOC3 match that expected for myofibroblasts. We have shown that the surface expression of AOC3 is sensitive to digestion by trypsin and collagenase and that anti-AOC3 antibodies can be used for FACS sorting of myofibroblasts obtained by nonenzymatic procedures. Whole-genome microarray mRNA-expression profiles of myofibroblasts and skin fibroblasts revealed four additional genes that are significantly differentially expressed in these two cell types: NKX2-3 and LRRC17 in myofibroblasts and SHOX2 and TBX5 in skin fibroblasts. TGFβ substantially down-regulated AOC3 expression in myofibroblasts but in skin fibroblasts it dramatically increased the expression of αSMA. A knockdown of NKX2-3 in myofibroblasts caused a decrease of myofibroblast-related gene expression and increased expression of the fibroblast-associated gene SHOX2, suggesting that NKX2-3 is a key mediator for maintaining myofibroblast characteristics. Our results show that colorectal myofibroblasts, as defined by the expression of AOC3, NKX2-3, and other markers, are a distinctly different cell type from TGFβ-activated fibroblasts. PMID:27036009

  8. Gli2 protein expression level is a feasible marker of ligand-dependent hedgehog activation in pancreatic neoplasms.

    PubMed

    Sugiyama, Y; Sasajima, J; Mizukami, Y; Koizumi, K; Kawamoto, T; Ono, Y; Karasaki, H; Tanabe, H; Fujiya, M; Kohgo, Y

    2016-06-01

    The hedgehog pathway is known to promote proliferation of pancreatic ductal adenocarcinoma (PDA) and has been shown to restrain tumor progression. To understand how hedgehog causes these effects, we sought to carefully examine protein expression of hedgehog signaling components during different tumor stages. Genetically engineered mice, Pdx1-Cre;LSL-KrasG12D and Pdx1-Cre;LSL-KrasG12D;p53lox/+, were utilized to model distinct phases of tumorigenesis, pancreatic intraepithelial neoplasm (PanIN) and PDA. Human pancreatic specimens of intraductal papillary mucinous neoplasm (IPMN) and PDA were also employed. PanIN and IPMN lesions highly express Sonic Hedgehog, at a level that is slightly higher than that observed in PDA. GLI2 protein is also expressed in both PanIN/IPMN and PDA. Although there was no difference in the nuclear staining, the cytoplasmic GLI2 level in PDA was modest in comparison to that in PanIN/IPMN. Hedgehog interacting protein was strongly expressed in the precursors, whereas the level in PDA was significantly attenuated. There were no differences in expression of Patched1 at early and late stages. Finally, a strong correlation between Sonic Hedgehog and GLI2 staining was found in both human and murine pancreatic tumors. The results indicate that the GLI2 protein level could serve as a feasible marker of ligand-dependent hedgehog activation in pancreatic neoplasms. PMID:27543868

  9. CD24 expression as a marker for predicting clinical outcome and invasive activity in uterine cervical cancer.

    PubMed

    Tanaka, Tomohito; Terai, Yoshito; Kogata, Yuhei; Ashihara, Keisuke; Maeda, Kazuya; Fujiwara, Satoe; Yoo, Saha; Tanaka, Yoshimichi; Tsunetoh, Satoshi; Sasaki, Hiroshi; Kanemura, Masanori; Tanabe, Akiko; Ohmichi, Masahide

    2015-11-01

    CD24, a small heavily glycosylated mucin-like glycosyl-phosphatidylinositol-anchored cell surface protein, plays an important role in the carcinogenesis of various human malignancies. However, its function in cervical cancer remains unclear. The aim of the present study was to evaluate the expression of CD24 clinicopathologically and to analyze its functional behavior biologically in cervical cancer. A total of 117 uterine cervical cancer tumors were immunohistochemically analyzed using a CD24 monoclonal antibody on paraffin blocks. We also examined whether CD24 enhanced the invasive activity or the Akt, ERK, NF-κB and MMP activity in a uterine cervical cancer cell line (CaSki) by a western blot analysis. The patients with enhanced CD24 expression had a higher rate of advanced clinical stage (50 vs. 16.5%, p<0.01), lymph node metastasis (34.6 vs. 14.3%) and lymphovascular involvement (65.4 vs. 20.4%, p=0.01), and a poor overall and disease-free survival (5-year survival rate: 62 vs. 86%, p=0.03). CD24 overexpression in CaSki cells resulted in activation of Cell Signaling proteins, including Akt, ERK, NF-κB and MMP-9. An invasion assay showed that CD24 overexpression in CaSki cells led to increased invasion ability. The CD24 overexpression also increased mRNA expression of Slug but not Snail. Moreover, the CD24 overexpression also decreased expression of E-cadherin and increased N-cadherin protein levels. Increased expression of CD24 may be associated with tumor progression and prognosis in patients with uterine cervical cancer. CD24 expression may therefore be used not only as a prognostic marker in uterine cervical cancer, but also as a target for the development of new therapeutic approaches.

  10. Expression of stemness markers in mouse parthenogenetic-diploid blastocysts is influenced by slight variation of activation protocol adopted.

    PubMed

    Bianchi, Enrica; Geremia, Raffaele; Sette, Claudio

    2010-07-01

    The importance of obtaining stem cells through alternative methods has increased progressively in the recent years due to the potential role that embryonic stem (ES) cells play in the field of regenerative medicine. In this regard, generation of parthenogenetic blastocysts allows the production of ethic-free ES cells without the need to manipulate normal embryos. Our work was aimed at clarifying whether variations in the method adopted to generate diploid parthenogenetic blastocysts could determine differences in the quality of blastocysts produced. In vitro development of mouse oocytes activated with three protocols, using Sr2+ and cytochalasin for different time, was compared with that of in vivo fertilized embryos. We have evaluated the efficiency of blastocyst formation and analysed the expression pattern of the stemness markers OCT4, CDX2, and NANOG. Our results indicate that the yield of diploid parthenogenotes and the segregation of the stemness marker OCT4 in the developing blastocyst are influenced by the parthenogenetic protocol adopted. Particularly, even if all methods tested allowed the production of blastocysts in vitro, the correct segregation of OCT4 occurred only in blastocysts developed from oocytes concomitantly treated for 4 h with Sr2+ and cytochalasin D. Our results indicate that the protocol employed to develop parthenogenetic blastocysts in vitro affects the quality of cells in the inner cell mass.

  11. Expression of stemness markers in mouse parthenogenetic-diploid blastocysts is influenced by slight variation of activation protocol adopted.

    PubMed

    Bianchi, Enrica; Geremia, Raffaele; Sette, Claudio

    2010-07-01

    The importance of obtaining stem cells through alternative methods has increased progressively in the recent years due to the potential role that embryonic stem (ES) cells play in the field of regenerative medicine. In this regard, generation of parthenogenetic blastocysts allows the production of ethic-free ES cells without the need to manipulate normal embryos. Our work was aimed at clarifying whether variations in the method adopted to generate diploid parthenogenetic blastocysts could determine differences in the quality of blastocysts produced. In vitro development of mouse oocytes activated with three protocols, using Sr2+ and cytochalasin for different time, was compared with that of in vivo fertilized embryos. We have evaluated the efficiency of blastocyst formation and analysed the expression pattern of the stemness markers OCT4, CDX2, and NANOG. Our results indicate that the yield of diploid parthenogenotes and the segregation of the stemness marker OCT4 in the developing blastocyst are influenced by the parthenogenetic protocol adopted. Particularly, even if all methods tested allowed the production of blastocysts in vitro, the correct segregation of OCT4 occurred only in blastocysts developed from oocytes concomitantly treated for 4 h with Sr2+ and cytochalasin D. Our results indicate that the protocol employed to develop parthenogenetic blastocysts in vitro affects the quality of cells in the inner cell mass. PMID:20376706

  12. B lymphocytes and B-cell activating factor promote collagen and profibrotic markers expression by dermal fibroblasts in systemic sclerosis

    PubMed Central

    2013-01-01

    Introduction B lymphocytes might play a pathogenic role in dermal fibrosis in systemic sclerosis (SSc). B-cell activating factor (BAFF), a key cytokine for B-cell activation, is increased in the serum and the skin of patients with SSc. However, the ability of B cells directly to stimulate dermal fibroblasts and the role of BAFF are not fully understood. We therefore investigated the involvement of B cells and BAFF in the expression of collagen and profibrotic markers by dermal fibroblasts. Methods Cocultures of blood B cells from healthy blood donors and normal or SSc dermal fibroblasts stimulated with anti-IgM and BAFF were performed. Alpha-SMA, TIMP1, MMP9, COL1A1, COL1A2, and COL3A1 mRNA expression were determined by quantitative RT-PCR. Soluble collagen, BAFF, IL-6, IL-1β, TGF-β1, and CCL2 protein secretion were assessed. Results Coculture of blood B cells and dermal fibroblasts isolated from SSc patients induced IL-6, TGF-β1, CCL2, and collagen secretion, as well as Alpha-SMA, TIMP1, and MMP9 expression in dermal fibroblasts. Transwell assays demonstrated that this induction was dependent on cell-cell contact. Addition of anti-IgM and BAFF to the coculture increased IL-6, CCL2, TGF-β1, and collagen secretion. B cell- and BAFF-induced collagen secretion was highly reduced by anti-TGF-β1 antibodies. Conclusions Our results showed for the first time a direct role of B cells on the production of collagen by dermal fibroblasts, which is further enhanced by BAFF. Thus, these results demonstrate a new pathogenic role of B cells and BAFF in fibrosis and systemic sclerosis. PMID:24289101

  13. Caffedymine from cocoa has COX inhibitory activity suppressing the expression of a platelet activation marker, P-Selectin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Caffedymine (N-caffeoyldopamine) is a clovamide-type of phenylpropenoic acid amide found in plants. Previous studies indicate that caffedymine inhibits P-selectin expression on platelets by increasing cAMP through beta-2 adrenoceptors, but the inhibition was only partially repressed by beta-2 adreno...

  14. Expression of Th1, Th2, lymphocyte trafficking and activation markers on CD4+ T-cells of Hymenoptera allergic subjects and after venom immunotherapy.

    PubMed

    Cabrera, Carmen M; Urra, José M; Alfaya, Teresa; Roca, Federico De La; Feo-Brito, Francisco

    2014-11-01

    Systemic reactions to Hymenoptera stings can be fatal and represent a reduction in the quality of life. The immune mechanisms involved in venom allergic subjects are barely known. Nevertheless, a shift towards a Th1-type response with an increase in IFNγ levels has been observed after venom immunotherapy (VIT). There is currently no information available about the expression of markers on CD4+ T-cells or their involvement in venom allergy, nor following VIT. For this, we have studied the expression of Th1 and Th2-cell markers, homing receptors and activation markers on CD4+ T-cells of subjects who presented systemic allergic reactions, mainly to Polistes dominulus, and after receiving a 4-month conventional VIT protocol. The markers studied were: CD26 (Th1), CD30 (Th2), CXCR4, CXCR3 (Th1), CCR4 (Th2), CD154 (CD40L), CD152 (CTLA-A), and ICOS. We also determined the IL-4 (Th2) and IFNγ (Th1) intracellular cytokine levels in T-cells and carried out a basophil activation test (BAT). Comparing venom allergic subjects with non-allergic healthy controls, we have found up-regulation of CD26, CXCR4, CXCR3, CD154 and ICOS. Conversely, a down-regulation of CD30, CD154 and CD152 occurred upon immune intervention, whereas the remaining markers were not affected. Equally, VIT has been shown to be effective, as evidenced by the decrease of basophil degranulation and increase of IFNγ levels in T-cells after the fourth month of treatment. These new findings highlight the possible application of these surface molecules as markers to distinguish between symptomatic and asymptomatic subjects sensitized to Hymenoptera venom, as well as revealing information about the immune changes associated with VIT.

  15. Sustained activation of DNA damage response in irradiated apoptosis-resistant cells induces reversible senescence associated with mTOR downregulation and expression of stem cell markers

    PubMed Central

    Chitikova, Zhanna V; Gordeev, Serguei A; Bykova, Tatiana V; Zubova, Svetlana G; Pospelov, Valery A; Pospelova, Tatiana V

    2014-01-01

    Cells respond to genotoxic stress by activating the DNA damage response (DDR). When injury is severe or irreparable, cells induce apoptosis or cellular senescence to prevent transmission of the lesions to the daughter cells upon cell division. Resistance to apoptosis is a hallmark of cancer that challenges the efficacy of cancer therapy. In this work, the effects of ionizing radiation on apoptosis-resistant E1A + E1B transformed cells were investigated to ascertain whether the activation of cellular senescence could provide an alternative tumor suppressor mechanism. We show that irradiated cells arrest cell cycle at G2/M phase and resume DNA replication in the absence of cell division followed by formation of giant polyploid cells. Permanent activation of DDR signaling due to impaired DNA repair results in the induction of cellular senescence in E1A + E1B cells. However, irradiated cells bypass senescence and restore the population by dividing cells, which have near normal size and ploidy and do not express senescence markers. Reversion of senescence and appearance of proliferating cells were associated with downregulation of mTOR, activation of autophagy, mitigation of DDR signaling, and expression of stem cell markers. PMID:24626185

  16. Heterogeneity of mouse spleen dendritic cells: in vivo phagocytic activity, expression of macrophage markers, and subpopulation turnover.

    PubMed

    Leenen, P J; Radosević, K; Voerman, J S; Salomon, B; van Rooijen, N; Klatzmann, D; van Ewijk, W

    1998-03-01

    In the normal mouse spleen, two distinct populations of dendritic cells (DC) are present that differ in microanatomical location. The major population of marginal DC is found in the "marginal zone bridging channels" and extends into the red pulp. The interdigitating cells (IDC) are localized in the T cell areas in the white pulp. The aim of the present study was to characterize these two splenic DC populations with regard to their phenotype, in vivo phagocytic function, and turnover. Both marginal DC and IDC are CD11c+ and CD13+, but only IDC are NLDC-145+ and CD8alpha+. Notably, both populations, when freshly isolated, express the macrophage markers F4/80, BM8, and Mac-1. To study the phagocytic capacity of these cells, we employed the macrophage "suicide" technique by injecting liposomes loaded with clodronate i.v. Marginal DC, but not IDC, were eliminated by this treatment. Phagocytosis of DiI-labeled liposomes by DC confirmed this finding. The two DC populations differed significantly with regard to their turnover rates, as studied in a transgenic mouse model of conditional depletion of DC populations with high turnover. In these mice, marginal DC were completely eliminated, but the IDC population remained virtually intact. From these data we conclude that the marginal DC population has a high turnover, in contrast to the IDC population. Taken together, the present results indicate that marginal DC and IDC represent two essentially distinct populations of DC in the mouse spleen. They differ not only in location, but also in phenotype, phagocytic ability, and turnover.

  17. Nonenzymatic Lipid Peroxidation Reprograms Gene Expression and Activates Defense Markers in Arabidopsis Tocopherol-Deficient Mutants[W

    PubMed Central

    Sattler, Scott E.; Mène-Saffrané, Laurent; Farmer, Edward E.; Krischke, Markus; Mueller, Martin J.; DellaPenna, Dean

    2006-01-01

    Tocopherols (vitamin E) are lipophilic antioxidants that are synthesized by all plants and are particularly abundant in seeds. Two tocopherol-deficient mutant loci in Arabidopsis thaliana were used to examine the functions of tocopherols in seedlings: vitamin e1 (vte1), which accumulates the pathway intermediate 2,3-dimethyl-5-phytyl-1,4-benzoquinone (DMPBQ); and vte2, which lacks all tocopherols and pathway intermediates. Only vte2 displayed severe seedling growth defects, which corresponded with massively increased levels of the major classes of nonenzymatic lipid peroxidation products: hydroxy fatty acids, malondialdehyde, and phytoprostanes. In the absence of pathogens, the phytoalexin camalexin accumulated in vte2 seedlings to levels 100-fold higher than in wild-type or vte1 seedlings. Similarly, gene expression profiling in wild-type, vte1, and vte2 seedlings indicated that increased levels of nonenzymatic lipid peroxidation in vte2 corresponded to increased expression of many defense-related genes, which were not induced in vte1. Both biochemical and transcriptional analyses of vte2 seedlings indicate that nonenzymatic lipid peroxidation plays a significant role in modulating plant defense responses. Together, these results establish that tocopherols in wild-type plants or DMPBQ in vte1 plants limit nonenzymatic lipid peroxidation during germination and early seedling development, thereby preventing the inappropriate activation of transcriptional and biochemical defense responses. PMID:17194769

  18. Effects of the interaction of diabetes and iron supplementation on hepatic and pancreatic tissues, oxidative stress markers, and liver peroxisome proliferator-activated receptor-α expression

    PubMed Central

    Silva, Maísa; Bonomo, Larissa de Freitas; Oliveira, Riva de Paula; Geraldo de Lima, Wanderson; Silva, Marcelo Eustáquio; Pedrosa, Maria Lucia

    2011-01-01

    This study evaluated the effects of the interaction of diabetes and a carbonyl iron supplemented on hepatic and pancreatic tissues, oxidative stress markers and liver peroxisome proliferator-activated receptor-α expressions. Hamsters were divided: Control which received a standard AIN 93 diet; Control Iron, composed of control animals that received a diet with 0.83% carbonyl iron; Diabetic, composed of animals that received a injection of streptozotocin (50 mg/kg, intraperitoneal) on day 35; and Diabetic Iron composed of streptozotocin treated animals that received a diet supplemented with carbonyl iron. Diabetes increased the glucose level and reduced triglycerides. Diabetic Iron group showed higher levels of glucose and serum triglycerides as compared to the Diabetic group. Diabetes decreased mRNA levels of peroxisome proliferator-activated receptor-α. Iron attenuated the diabetes induced down regulation of peroxisome proliferator-activated receptor-α mRNA. Moreover, diabetes increased carbonyl protein and decreased glutathione levels and catalase activity, while iron attenuated the increase in levels of carbonyl protein and attenuated the decrease in those of glutathione level and catalase activity. Histological analysis shows that supplementation iron caused an increase in the size of the islets in Control Iron. The results show that iron does not aggravated liver oxidant/antioxidant status and peroxisome proliferator-activated receptor-α expression in diabetic hamsters. PMID:21980225

  19. Increased expression of senescence markers in cystic fibrosis airways.

    PubMed

    Fischer, Bernard M; Wong, Jessica K; Degan, Simone; Kummarapurugu, Apparao B; Zheng, Shuo; Haridass, Prashamsha; Voynow, Judith A

    2013-03-15

    Cystic Fibrosis (CF) is a chronic lung disease characterized by chronic neutrophilic airway inflammation and increased levels of neutrophil elastase (NE) in the airways. We have previously reported that NE treatment triggers cell cycle arrest. Cell cycle arrest can lead to senescence, a complete loss of replicative capacity. Importantly, senescent cells can be proinflammatory and would perpetuate CF chronic inflammation. By immunohistochemistry, we evaluated whether airway sections from CF and control subjects expressed markers of senescence, including p16(INK4a) (p16), a cyclin-dependent kinase inhibitor, phospho-Histone H2A.X (γH2A.X), and phospho-checkpoint 2 kinase (phospho-Chk2), which are also DNA damage response markers. Compared with airway epithelium from control subjects, CF airway epithelium had increased levels of expression of all three senescence markers. We hypothesized that the high load of NE in the CF airway triggers epithelial senescence by upregulating expression of p16, which inhibits cyclin-dependent kinase 4 (CDK4). Normal human bronchial epithelial (NHBE) cells, cultured in air-liquid interface were treated with NE (0, 200, and 500 nM) to induce visible injury. Total cell lysates were collected and evaluated by Western analysis for p16 protein expression and CDK4 kinase activity. NE significantly increased p16 expression and decreased CDK4 kinase activity in NHBE cells. These results support the concept that NE triggers expression of senescence markers in CF airway epithelial cells. PMID:23316069

  20. Activation of inflammatory responses in human U937 macrophages by particulate matter collected from dairy farms: an in vitro expression analysis of pro-inflammatory markers

    PubMed Central

    2012-01-01

    Background The purpose of the present study was to investigate activation of inflammatory markers in human macrophages derived from the U937 cell line after exposure to particulate matter (PM) collected on dairy farms in California and to identify the most potent components of the PM. Methods PM from different dairies were collected and tested to induce an inflammatory response determined by the expression of various pro-inflammatory genes, such as Interleukin (IL)-8, in U937 derived macrophages. Gel shift and luciferase reporter assays were performed to examine the activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and Toll-like-receptor 4 (TLR4). Results Macrophage exposure to PM derived from dairy farms significantly activated expression of pro-inflammatory genes, including IL-8, cyclooxygenase 2 and Tumor necrosis factor-alpha, which are hallmarks of inflammation. Acute phase proteins, such as serum amyloid A and IL-6, were also significantly upregulated in macrophages treated with PM from dairies. Coarse PM fractions demonstrated more pro-inflammatory activity on an equal-dose basis than fine PM. Urban PM collected from the same region as the dairy farms was associated with a lower concentration of endotoxin and produced significantly less IL-8 expression compared to PM collected on the dairy farms. Conclusion The present study provides evidence that the endotoxin components of the particles collected on dairies play a major role in mediating an inflammatory response through activation of TLR4 and NF-κB signaling. PMID:22452745

  1. Expression of Molecular Markers of Angiogenesis, Lymphangiogenesis, and Proliferation Depending on the Stage of Skin Melanoma.

    PubMed

    Bgatova, N P; Lomakin, A I; Fursov, S A; Kachesov, I V; Chepko, S A; Isakova, N B; Borodin, Yu I; Voytsitsky, V E; Konenkov, V I

    2016-08-01

    The expression of molecular markers characterizing activity of the tumor process and metastases (proliferation marker Ki-67, angiogenesis marker CD34, and lymphangiogenesis markers podoplanin and LYVE-1) was assessed by immunohictochemical method in the primary tumor specimens collected during surgery for cutaneous melanoma (40 patients). Proliferative activity of the tumor tissue and volume density of peritumoral blood and lymph vessels increased with increasing tumor malignancy, which could indicate the risk of metastases. PMID:27590758

  2. Early onset of hypersynchronous network activity and expression of a marker of chronic seizures in the Tg2576 mouse model of Alzheimer's disease.

    PubMed

    Bezzina, Charlotte; Verret, Laure; Juan, Cécile; Remaud, Jessica; Halley, Hélène; Rampon, Claire; Dahan, Lionel

    2015-01-01

    Cortical and hippocampal hypersynchrony of neuronal networks seems to be an early event in Alzheimer's disease pathogenesis. Many mouse models of the disease also present neuronal network hypersynchrony, as evidenced by higher susceptibility to pharmacologically-induced seizures, electroencephalographic seizures accompanied by spontaneous interictal spikes and expression of markers of chronic seizures such as neuropeptide Y ectopic expression in mossy fibers. This network hypersynchrony is thought to contribute to memory deficits, but whether it precedes the onset of memory deficits or not in mouse models remains unknown. The earliest memory impairments in the Tg2576 mouse model of Alzheimer's disease have been observed at 3 months of age. We thus assessed network hypersynchrony in Tg2576 and non-transgenic male mice at 1.5, 3 and 6 months of age. As soon as 1.5 months of age, Tg2576 mice presented higher seizure susceptibility to systemic injection of a GABAA receptor antagonist. They also displayed spontaneous interictal spikes on EEG recordings. Some Tg2576 mice presented hippocampal ectopic expression of neuropeptide Y which incidence seems to increase with age among the Tg2576 population. Our data reveal that network hypersynchrony appears very early in Tg2576 mice, before any demonstrated memory impairments. PMID:25768013

  3. Early Onset of Hypersynchronous Network Activity and Expression of a Marker of Chronic Seizures in the Tg2576 Mouse Model of Alzheimer’s Disease

    PubMed Central

    Bezzina, Charlotte; Verret, Laure; Juan, Cécile; Remaud, Jessica; Halley, Hélène

    2015-01-01

    Cortical and hippocampal hypersynchrony of neuronal networks seems to be an early event in Alzheimer’s disease pathogenesis. Many mouse models of the disease also present neuronal network hypersynchrony, as evidenced by higher susceptibility to pharmacologically-induced seizures, electroencephalographic seizures accompanied by spontaneous interictal spikes and expression of markers of chronic seizures such as neuropeptide Y ectopic expression in mossy fibers. This network hypersynchrony is thought to contribute to memory deficits, but whether it precedes the onset of memory deficits or not in mouse models remains unknown. The earliest memory impairments in the Tg2576 mouse model of Alzheimer’s disease have been observed at 3 months of age. We thus assessed network hypersynchrony in Tg2576 and non-transgenic male mice at 1.5, 3 and 6 months of age. As soon as 1.5 months of age, Tg2576 mice presented higher seizure susceptibility to systemic injection of a GABAA receptor antagonist. They also displayed spontaneous interictal spikes on EEG recordings. Some Tg2576 mice presented hippocampal ectopic expression of neuropeptide Y which incidence seems to increase with age among the Tg2576 population. Our data reveal that network hypersynchrony appears very early in Tg2576 mice, before any demonstrated memory impairments. PMID:25768013

  4. Enhanced P-selectin expression on platelet-a marker of platelet activation, in young patients with angiographically proven coronary artery disease.

    PubMed

    George, Reema; Bhatt, Anugya; Narayani, Jayakumari; Thulaseedharan, Jissa Vinoda; Sivadasanpillai, Harikrishnan; Tharakan, Jaganmohan A

    2016-08-01

    P-selectin (CD62p) exposure is an established marker for platelet activation. P-selectin exposure can trigger variety of thrombotic and inflammatory reactions. In patients with coronary artery disease (CAD), platelets are activated, and hence, there is increased P-selectin exposure. The role of P-selectin exposure in patients on treatment with statins and anti-platelets is conflicting. A case-control study was performed to determine P-selectin exposure in consecutively recruited 142 patients (age ≤ 55 years) with angiographically proven CAD on treatment and 92 asymptomatic controls. P-selectin exposure was determined by flow cytometry. Data on conventional risk factors were obtained along with estimation of levels of thrombotic [fibrinogen, lipoprotein (a), tissue plasminogen activator, plasminogen activator inhibitor-1, homocysteine and von Willebrand factor] and anti-thrombotic factors (antithrombin III). The P-selectin exposure was compared among patient groups who had different modes of presentation of CAD and categories of CAD disease severity. The patients were followed up for a period of 26 months. The results indicate that P-selectin exposure was significantly elevated in patients (mean ± SD 9.24 ± 11.81) compared to controls (mean ± SD 1.48 ± 2.85) with p < 0.0001. Similarly, conventional risk factors were significantly elevated in patients. P-selectin exposure showed significant negative correlation with antithrombin III levels. P-selectin exposure was higher in patients who presented with acute coronary syndromes than those who presented with effort angina. Cardiovascular event rate was 6 % on follow-up. The study establishes that thrombotic-inflammatory pathways enhancing P-selectin exposure unrelated to treatment might be activated in patients, while the event rate remained lowered, and hence, treatment strategies should be inclusive to control these factors.

  5. Expression of Lymphatic Markers in the Adult Rat Spinal Cord

    PubMed Central

    Kaser-Eichberger, Alexandra; Schroedl, Falk; Bieler, Lara; Trost, Andrea; Bogner, Barbara; Runge, Christian; Tempfer, Herbert; Zaunmair, Pia; Kreutzer, Christina; Traweger, Andreas; Reitsamer, Herbert A.; Couillard-Despres, Sebastien

    2016-01-01

    Under physiological conditions, lymphatic vessels are thought to be absent from the central nervous system (CNS), although they are widely distributed within the rest of the body. Recent work in the eye, i.e., another organ regarded as alymphatic, revealed numerous cells expressing lymphatic markers. As the latter can be involved in the response to pathological conditions, we addressed the presence of cells expressing lymphatic markers within the spinal cord by immunohistochemistry. Spinal cord of young adult Fisher rats was scrutinized for the co-expression of the lymphatic markers PROX1 and LYVE-1 with the cell type markers Iba1, CD68, PGP9.5, OLIG2. Rat skin served as positive control for the lymphatic markers. PROX1-immunoreactivity was detected in many nuclei throughout the spinal cord white and gray matter. These nuclei showed no association with LYVE-1. Expression of LYVE-1 could only be detected in cells at the spinal cord surface and in cells closely associated with blood vessels. These cells were found to co-express Iba1, a macrophage and microglia marker. Further, double labeling experiments using CD68, another marker found in microglia and macrophages, also displayed co-localization in the Iba1+ cells located at the spinal cord surface and those apposed to blood vessels. On the other hand, PROX1-expressing cells found in the parenchyma were lacking Iba1 or PGP9.5, but a significant fraction of those cells showed co-expression of the oligodendrocyte lineage marker OLIG2. Intriguingly, following spinal cord injury, LYVE-1-expressing cells assembled and reorganized into putative pre-vessel structures. As expected, the rat skin used as positive controls revealed classical lymphatic vessels, displaying PROX1+ nuclei surrounded by LYVE-1-immunoreactivity. Classical lymphatics were not detected in adult rat spinal cord. Nevertheless, numerous cells expressing either LYVE-1 or PROX1 were identified. Based on their localization and overlapping expression with

  6. Expression of Lymphatic Markers in the Adult Rat Spinal Cord.

    PubMed

    Kaser-Eichberger, Alexandra; Schroedl, Falk; Bieler, Lara; Trost, Andrea; Bogner, Barbara; Runge, Christian; Tempfer, Herbert; Zaunmair, Pia; Kreutzer, Christina; Traweger, Andreas; Reitsamer, Herbert A; Couillard-Despres, Sebastien

    2016-01-01

    Under physiological conditions, lymphatic vessels are thought to be absent from the central nervous system (CNS), although they are widely distributed within the rest of the body. Recent work in the eye, i.e., another organ regarded as alymphatic, revealed numerous cells expressing lymphatic markers. As the latter can be involved in the response to pathological conditions, we addressed the presence of cells expressing lymphatic markers within the spinal cord by immunohistochemistry. Spinal cord of young adult Fisher rats was scrutinized for the co-expression of the lymphatic markers PROX1 and LYVE-1 with the cell type markers Iba1, CD68, PGP9.5, OLIG2. Rat skin served as positive control for the lymphatic markers. PROX1-immunoreactivity was detected in many nuclei throughout the spinal cord white and gray matter. These nuclei showed no association with LYVE-1. Expression of LYVE-1 could only be detected in cells at the spinal cord surface and in cells closely associated with blood vessels. These cells were found to co-express Iba1, a macrophage and microglia marker. Further, double labeling experiments using CD68, another marker found in microglia and macrophages, also displayed co-localization in the Iba1+ cells located at the spinal cord surface and those apposed to blood vessels. On the other hand, PROX1-expressing cells found in the parenchyma were lacking Iba1 or PGP9.5, but a significant fraction of those cells showed co-expression of the oligodendrocyte lineage marker OLIG2. Intriguingly, following spinal cord injury, LYVE-1-expressing cells assembled and reorganized into putative pre-vessel structures. As expected, the rat skin used as positive controls revealed classical lymphatic vessels, displaying PROX1+ nuclei surrounded by LYVE-1-immunoreactivity. Classical lymphatics were not detected in adult rat spinal cord. Nevertheless, numerous cells expressing either LYVE-1 or PROX1 were identified. Based on their localization and overlapping expression with

  7. Expression of squamous cell carcinoma markers and adenocarcinoma markers in primary pulmonary neuroendocrine carcinomas.

    PubMed

    Masai, Kyohei; Tsuta, Koji; Kawago, Mitsumasa; Tatsumori, Takahiro; Kinno, Tomoaki; Taniyama, Tomoko; Yoshida, Akihiko; Asamura, Hisao; Tsuda, Hitoshi

    2013-07-01

    Recent clinical trials have revealed that accurate histologic typing of non-small cell lung cancer is essential. Until now, squamous cell carcinoma (SQC) and adenocarcinoma (ADC) markers have not been thoroughly analyzed for pulmonary neuroendocrine carcinomas (NECs). We analyzed the expression of 8 markers [p63, cytokeratin (CK) 5/6, SOX2, CK7, desmocollin 3, thyroid transcription factor-1 (8G7G3/1 and SPT24), and napsin A] in 224 NECs. SOX2 (76.2%) had the greatest expression for NECs. CK5/6 (1.4%), desmocollin 3 (0.5%), and napsin A (0%) were expressed less or not at all in NECs. Although our investigated markers have been reported useful for differentiating between SQC and ADC, some of them were also present in a portion of pulmonary NECs. In our study, CK5/6 and desmocollin 3 were highly specific markers for SQC, and napsin A was highly specific for ADC. These markers are recommended for diagnosis of poorly differentiated non-small cell lung cancer.

  8. Stem Cell-Associated Marker Expression in Canine Hair Follicles.

    PubMed

    Gerhards, Nora M; Sayar, Beyza S; Origgi, Francesco C; Galichet, Arnaud; Müller, Eliane J; Welle, Monika M; Wiener, Dominique J

    2016-03-01

    Functional hair follicle (HF) stem cells (SCs) are crucial to maintain the constant recurring growth of hair. In mice and humans, SC subpopulations with different biomarker expression profiles have been identified in discrete anatomic compartments of the HF. The rare studies investigating canine HF SCs have shown similarities in biomarker expression profiles to that of mouse and human SCs. The aim of our study was to broaden the current repertoire of SC-associated markers and their expression patterns in the dog. We combined analyses on the expression levels of CD34, K15, Sox9, CD200, Nestin, LGR5 and LGR6 in canine skin using RT-qPCR, the corresponding proteins in dog skin lysates, and their expression patterns in canine HFs using immunohistochemistry. Using validated antibodies, we were able to define the location of CD34, Sox9, Keratin15, LGR5 and Nestin in canine HFs and confirm that all tested biomarkers are expressed in canine skin. Our results show similarities between the expression profile of canine, human and mouse HF SC markers. This repertoire of biomarkers will allow us to conduct functional studies and investigate alterations in the canine SC compartment of different diseases, like alopecia or skin cancer with the possibility to extend relevant findings to human patients.

  9. Stem Cell-Associated Marker Expression in Canine Hair Follicles.

    PubMed

    Gerhards, Nora M; Sayar, Beyza S; Origgi, Francesco C; Galichet, Arnaud; Müller, Eliane J; Welle, Monika M; Wiener, Dominique J

    2016-03-01

    Functional hair follicle (HF) stem cells (SCs) are crucial to maintain the constant recurring growth of hair. In mice and humans, SC subpopulations with different biomarker expression profiles have been identified in discrete anatomic compartments of the HF. The rare studies investigating canine HF SCs have shown similarities in biomarker expression profiles to that of mouse and human SCs. The aim of our study was to broaden the current repertoire of SC-associated markers and their expression patterns in the dog. We combined analyses on the expression levels of CD34, K15, Sox9, CD200, Nestin, LGR5 and LGR6 in canine skin using RT-qPCR, the corresponding proteins in dog skin lysates, and their expression patterns in canine HFs using immunohistochemistry. Using validated antibodies, we were able to define the location of CD34, Sox9, Keratin15, LGR5 and Nestin in canine HFs and confirm that all tested biomarkers are expressed in canine skin. Our results show similarities between the expression profile of canine, human and mouse HF SC markers. This repertoire of biomarkers will allow us to conduct functional studies and investigate alterations in the canine SC compartment of different diseases, like alopecia or skin cancer with the possibility to extend relevant findings to human patients. PMID:26739040

  10. Heterogeneous lineage marker expression in naive embryonic stem cells is mostly due to spontaneous differentiation.

    PubMed

    Nair, Gautham; Abranches, Elsa; Guedes, Ana M V; Henrique, Domingos; Raj, Arjun

    2015-08-21

    Populations of cultured mouse embryonic stem cells (ESCs) exhibit a subfraction of cells expressing uncharacteristically low levels of pluripotency markers such as Nanog. Yet, the extent to which individual Nanog-negative cells are differentiated, both from ESCs and from each other, remains unclear. Here, we show the transcriptome of Nanog-negative cells exhibits expression of classes of genes associated with differentiation that are not yet active in cells exposed to differentiation conditions for one day. Long non-coding RNAs, however, exhibit more changes in expression in the one-day-differentiated cells than in Nanog-negative cells. These results are consistent with the concept that Nanog-negative cells may contain subpopulations of both lineage-primed and differentiated cells. Single cell analysis showed that Nanog-negative cells display substantial and coherent heterogeneity in lineage marker expression in progressively nested subsets of cells exhibiting low levels of Nanog, then low levels of Oct4, and then a set of lineage markers, which express intensely in a small subset of these more differentiated cells. Our results suggest that the observed enrichment of lineage-specific marker gene expression in Nanog-negative cells is associated with spontaneous differentiation of a subset of these cells rather than the more random expression that may be associated with reversible lineage priming.

  11. Heterogeneous lineage marker expression in naive embryonic stem cells is mostly due to spontaneous differentiation

    PubMed Central

    Nair, Gautham; Abranches, Elsa; Guedes, Ana M. V.; Henrique, Domingos; Raj, Arjun

    2015-01-01

    Populations of cultured mouse embryonic stem cells (ESCs) exhibit a subfraction of cells expressing uncharacteristically low levels of pluripotency markers such as Nanog. Yet, the extent to which individual Nanog-negative cells are differentiated, both from ESCs and from each other, remains unclear. Here, we show the transcriptome of Nanog-negative cells exhibits expression of classes of genes associated with differentiation that are not yet active in cells exposed to differentiation conditions for one day. Long non-coding RNAs, however, exhibit more changes in expression in the one-day-differentiated cells than in Nanog-negative cells. These results are consistent with the concept that Nanog-negative cells may contain subpopulations of both lineage-primed and differentiated cells. Single cell analysis showed that Nanog-negative cells display substantial and coherent heterogeneity in lineage marker expression in progressively nested subsets of cells exhibiting low levels of Nanog, then low levels of Oct4, and then a set of lineage markers, which express intensely in a small subset of these more differentiated cells. Our results suggest that the observed enrichment of lineage-specific marker gene expression in Nanog-negative cells is associated with spontaneous differentiation of a subset of these cells rather than the more random expression that may be associated with reversible lineage priming. PMID:26292941

  12. Ras protein expression as a marker for breast cancer

    PubMed Central

    CALAF, GLORIA M.; ABARCA-QUINONES, JORGE

    2016-01-01

    Breast cancer, the most common neoplasm in women of all ages, is the leading cause of cancer-related mortality in women worldwide. Markers to help to predict the risk of progression and ultimately provide non-surgical treatment options would be of great benefit. At present, there are no available molecular markers to predict the risk of carcinoma in situ progression to invasive cancer; therefore, all women diagnosed with this type of malignancy must undergo surgery. Breast cancer is a heterogeneous complex disease, and different patients respond differently to different treatments. In breast cancer, analysis using immunohistochemical markers remains an essential component of routine pathological examinations, and plays an import role in the management of the disease by providing diagnostic and prognostic strategies. The aim of the present study was to identify a marker that can be used as a prognostic tool for breast cancer. For this purpose, we firstly used an established breast cancer model. MCF-10F, a spontaneously immortalized breast epithelial cell line was transformed by exposure to estrogen and radiation. MCF-10F cells were exposed to low doses of high linear energy transfer (LET) α particles (150 keV/μm) of radiation, and subsequently cultured in the presence of 17β-estradiol. Three cell lines were used: i) MCF-10F cells as a control; ii) Alpha5 cells, a malignant and tumorigenic cell line; and iii) Tumor2 cells derived from Alpha5 cells injected into nude mice. Secondly, we also used normal, benign and malignant breast specimens obtained from biopsies. The results revealed that the MCF-10F cells were negative for c-Ha-Ras protein expression; however, the Alpha5 and Tumor2 cell lines were positive for c-Ha-Ras protein expression. The malignant breast samples were also strongly positive for c-Ha-Ras expression. The findings of our study indicate that c-Ha-Ras protein expression may be used as a marker to predict the progression of breast cancer; this

  13. Expression Marker-Based Strategy to Improve Beef Quality.

    PubMed

    Cassar-Malek, Isabelle; Picard, Brigitte

    2016-01-01

    For beef cattle research, a main objective is to control concomitantly the development of muscles and the qualities of beef cuts. Beef quality is a complex phenotype that is only detectable after slaughter and is highly variable. The beef industry is in need of tools to estimate beef quality of live cattle or online in abattoirs, with specific attention towards sensory attributes (tenderness, juiciness, flavour, and colour). Identification of relevant genetic and genomic markers is ongoing, especially for tenderness--a top priority quality attribute. In this paper, we describe the steps of an expression marker-based strategy to improve beef sensory quality, from the discovery of biomarkers that identify consistent beef and the biological functions governing beef tenderness to the integration of the knowledge into detection tests for desirable animals. These tools should soon be available for the management of sensory quality in the beef production chain for meeting market's demands and assuring good quality standards. PMID:27066527

  14. Expression Marker-Based Strategy to Improve Beef Quality

    PubMed Central

    Cassar-Malek, Isabelle; Picard, Brigitte

    2016-01-01

    For beef cattle research, a main objective is to control concomitantly the development of muscles and the qualities of beef cuts. Beef quality is a complex phenotype that is only detectable after slaughter and is highly variable. The beef industry is in need of tools to estimate beef quality of live cattle or online in abattoirs, with specific attention towards sensory attributes (tenderness, juiciness, flavour, and colour). Identification of relevant genetic and genomic markers is ongoing, especially for tenderness—a top priority quality attribute. In this paper, we describe the steps of an expression marker-based strategy to improve beef sensory quality, from the discovery of biomarkers that identify consistent beef and the biological functions governing beef tenderness to the integration of the knowledge into detection tests for desirable animals. These tools should soon be available for the management of sensory quality in the beef production chain for meeting market's demands and assuring good quality standards. PMID:27066527

  15. Myoglobin expression in prostate cancer is correlated to androgen receptor expression and markers of tumor hypoxia.

    PubMed

    Meller, Sebastian; Bicker, Anne; Montani, Matteo; Ikenberg, Kristian; Rostamzadeh, Babak; Sailer, Verena; Wild, Peter; Dietrich, Dimo; Uhl, Barbara; Sulser, Tullio; Moch, Holger; Gorr, Thomas A; Stephan, Carsten; Jung, Klaus; Hankeln, Thomas; Kristiansen, Glen

    2014-10-01

    Recent studies identified unexpected expression and transcriptional complexity of the hemoprotein myoglobin (MB) in human breast cancer but its role in prostate cancer is still unclear. Expression of MB was immunohistochemically analyzed in three independent cohorts of radical prostatectomy specimens (n = 409, n = 625, and n = 237). MB expression data were correlated with clinicopathological parameters and molecular parameters of androgen and hypoxia signaling. Expression levels of novel tumor-associated MB transcript variants and the VEGF gene as a hypoxia marker were analyzed using qRT-PCR. Fifty-three percent of the prostate cancer cases were MB positive and significantly correlated with androgen receptor (AR) expression (p < 0.001). The positive correlation with CAIX (p < 0.001) and FASN (p = 0.008) as well as the paralleled increased expression of the tumor-associated MB transcript variants and VEGF suggest that hypoxia participates in MB expression regulation. Analogous to breast cancer, MB expression in prostate cancer is associated with steroid hormone signaling and markers of hypoxia. Further studies must elucidate the novel functional roles of MB in human carcinomas, which probably extend beyond its classic intramuscular function in oxygen storage. PMID:25172328

  16. MAP17 and SGLT1 Protein Expression Levels as Prognostic Markers for Cervical Tumor Patient Survival

    PubMed Central

    Perez, Marco; Praena-Fernandez, Juan M.; Felipe-Abrio, Blanca; Lopez-Garcia, Maria A.; Lucena-Cacace, Antonio; Garcia, Angel; Lleonart, Matilde; Roncador, Guiovanna; Marin, Juan J.; Carnero, Amancio

    2013-01-01

    MAP17 is a membrane-associated protein that is overexpressed in human tumors. Because the expression of MAP17 increases reactive oxygen species (ROS) generation through SGLT1 in cancer cells, in the present work, we investigated whether MAP17 and/or SGLT1 might be markers for the activity of treatments involving oxidative stress, such as cisplatin or radiotherapy. First, we confirmed transcriptional alterations in genes involved in the oxidative stress induced by MAP17 expression in HeLa cervical tumor cells and found that Hela cells expressing MAP17 were more sensitive to therapies that induce ROS than were parental cells. Furthermore, MAP17 increased glucose uptake through SGLT receptors. We then analyzed MAP17 and SGLT1 expression levels in cervical tumors treated with cisplatin plus radiotherapy and correlated the expression levels with patient survival. MAP17 and SGLT1 were expressed in approximately 70% and 50% of cervical tumors of different types, respectively, but they were not expressed in adenoma tumors. Furthermore, there was a significant correlation between MAP17 and SGLT1 expression levels. High levels of either MAP17 or SGLT1 correlated with improved patient survival after treatment. However, the patients with high levels of both MAP17 and SGLT1 survived through the end of this study. Therefore, the combination of high MAP17 and SGLT1 levels is a marker for good prognosis in patients with cervical tumors after cisplatin plus radiotherapy treatment. These results also suggest that the use of MAP17 and SGLT1 markers may identify patients who are likely to exhibit a better response to treatments that boost oxidative stress in other cancer types. PMID:23418532

  17. Expression of cartilage-specific markers in calcified and non-calcified atherosclerotic lesions.

    PubMed

    Aigner, Thomas; Neureiter, Daniel; Câmpean, Valentina; Soder, Stephan; Amann, Kerstin

    2008-01-01

    Recently, molecular mechanisms resembling endochondral ossification were suggested to be important for atherosclerotic vessel calcification. The aim of this study was to investigate in a series of human atherosclerotic (non-diabetic) lesions of the crural arteries the distribution and expression of classical marker genes of the endochondral ossification pathway. Immunostaining for marker proteins S-100 protein and collagen types II and X were performed on atherosclerotic lesions of different grades (according to Stary). Quantitative real-time PCR for human COL1A1, COL2A1, COL10A1, SOX9, and BMP-2 was applied on RNA isolated from atherosclerotic arteries. In most samples, no expression of collagen type II and S-100 protein was found. Exceptionally, S-100 protein and type II collagen expression was observed very focally within advanced atherosclerotic plaques. Type X collagen was not detected in any of the lesions investigated. Overall, in our study we found no evidence that chondrogenic differentiation pathways are generally active in atherosclerotic plaque formation. In particular type X collagen, one important molecule in cartilage calcification, was not expressed in any of the investigated specimens. Occasionally, however, chondrocytic differentiation markers occur within atherosclerotic lesions. This most likely represents a metaplastic event associated, but not causative for atherosclerotic vessel degeneration and calcification. PMID:17335825

  18. Effects of synergistic massage and physical exercise on the expression of angiogenic markers in rat tendons.

    PubMed

    Andrzejewski, Waldemar; Kassolik, Krzysztof; Dziegiel, Piotr; Pula, Bartosz; Ratajczak-Wielgomas, Katarzyna; Jablonska, Karolina; Kurpas, Donata; Halski, Tomasz; Podhorska-Okolow, Marzena

    2014-01-01

    Physical exercise and massage are regarded as key factors in regulating tendon structure. However, information on the mechanism through which massage influences the structure and biology of a tendon is scarce. In this study, we attempted to define the impact of these two activities on rat tendons by using morphological and molecular techniques, determining the expression of VEGF-A, FGF-2, and CD34 in the tendons of rats subjected to 10 weeks of physical exercise (running) with massage of varied duration. The group of rats that was trained and massaged during the entire study was characterized by the highest expression of these markers, compared to the rats subjected to massage before training and to the control group subjected to physical exercises only. The greatest significant differences, compared to the control, were noted in the expression of all the studied markers at mRNA level, and in the case of VEGF-A, at protein level, in the third and fifth weeks of the experiment. The results of this study could point to the synergistic impact of simultaneous massage and physical exercise on the expression of angiogenesis markers in rat tendons.

  19. Tracing functional circuits using c-Fos regulated expression of marker genes targeted to neuronal projections.

    PubMed

    Murphy, Mark; Greferath, Ursula; Nag, Nupur; Nithianantharajah, Jess; Wilson, Yvette M

    2004-01-01

    We have developed novel techniques to trace functionally activated circuits and synaptic plasticity within the brain. We have generated transgenic mice, FTL, which contain a tau-lacZ fusion gene regulated by the promoter for c-fos. Following a particular nervous system stimulation in these mice, only neurons, which are functionally activated, will express LacZ, which is targeted to neuronal processes by the tau protein. In the FTL mice, we found highly inducible expression of lacZ by a range of different stimuli, and successful targeting of expression to neuronal cell bodies, axons and dendrites. To test if a functionally activated circuit could be visualized, the mice were deprived of water, which activates nuclei involved in body fluid homeostasis. LacZ was induced in these nuclei and their projections, allowing the mapping of a neuroendocrine circuit. Further studies have employed these mice in the analysis of neurons and circuits activated in vision, and learning and memory. We have also developed methods to measure markers of synaptic plasticity in the brain, and found significant experience dependent changes in the levels of these markers in different parts of the brain. We believe these techniques will aid in the identification of circuits for many different brain functions, and within those circuits, the locations of synaptic plasticity.

  20. Effect of microgrooves and fibronectin conjugation on the osteoblast marker gene expression and differentiation

    PubMed Central

    2015-01-01

    PURPOSE To determine the effect of fibronectin (FN)-conjugated, microgrooved titanium (Ti) on osteoblast differentiation and gene expression in human bone marrow-derived mesenchymal stem cells (MSCs). MATERIALS AND METHODS Photolithography was used to fabricate the microgrooved Ti, and amine functionalization (silanization) was used to immobilize fibronectin on the titanium surfaces. Osteoblast differentiation and osteoblast marker gene expression were analyzed by means of alkaline phosphatase activity assay, extracellular calcium deposition assay, and quantitative real-time PCR. RESULTS The conjugation of fibronectin on Ti significantly increased osteoblast differentiation in MSCs compared with non-conjugated Ti substrates. On the extracellular calcium deposition assays of MSCs at 21 days, an approximately two-fold increase in calcium concentration was observed on the etched 60-µm-wide/10-µm-deep microgrooved surface with fibronectin (E60/10FN) compared with the same surface without fibronectin (E60/10), and a more than four-fold increase in calcium concentration was observed on E60/10FN compared with the non-etched control (NE0) and etched control (E0) surfaces. Through a series of analyses to determine the expression of osteoblast marker genes, a significant increase in all the marker genes except type I collagen α1 mRNA was seen with E60/10FN more than with any of the other groups, as compared with NE0. CONCLUSION The FN-conjugated, microgrooved Ti substrate can provide an effective surface to promote osteoblast differentiation and osteoblast marker gene expression in MSCs. PMID:26816580

  1. Hexavalent chromium induces expression of mesenchymal and stem cell markers in renal epithelial cells.

    PubMed

    Li, Wei-Jen; Yang, Cheng-Lin; Chow, Kuan-Chih; Kuo, Ting-Wei

    2016-02-01

    Cr(VI) causes severe kidney damage. The patient's renal function could gradually recover by spontaneous kidney regeneration. The molecular effect of Cr(VI) on recovery of kidney cells, however, has not been clearly elucidated. Here we show that Cr(VI) induces expression of mesenchymal and stem cell markers, cell markers, such as paxillin, vimentin, α-SMA, nanog, and CD133 of HK-2 cells. Moreover, Cr(VI) activates epithelial-to-mesenchymal transition (EMT). By revealing that levels of dihydrodiol dehydrogenase were promptly reduced following Cr(VI) challenge, our data suggested that DDH could be involved in a Cr(VI)-related oxidation to generate massive reactive oxygen species and H2 O2 , and to create intracellular hypoxia, which then increased levels of SUMO-1 activating enzyme subunit 2, and sumoylation of eukaryotic elongation factor-2, to mediate the subsequent molecular and cellular responses, e.g., expression of mesenchymal and stem cell markers. Pretreatment with vitamin C reduced Cr(VI)-related cellular effects. However, no evident effect was observed when vitamin C was added following Cr(VI) challenge. PMID:25620490

  2. Rat visceral yolk sac cells: viability and expression of cell markers during maternal diabetes.

    PubMed

    Aires, M B; Santos, J R A; Souza, K S; Farias, P S; Santos, A C V; Fioretto, E T; Maria, D A

    2015-08-01

    The function of the visceral yolk sac (VYS) is critical for embryo organogenesis until final fetal development in rats, and can be affected by conditions such as diabetes. In view of the importance of diabetes during pregnancy for maternal and neonatal health, the objective of this study was to assess fetal weight, VYS cell markers, and viability in female Wistar rats (200-250 g) with induced diabetes (alloxan, 37 mg/kg) on the 8th gestational day (gd 8). At gd 15, rats from control (n=5) and diabetic (n=5) groups were anesthetized and laparotomized to remove the uterine horns for weighing of fetuses and collecting the VYS. Flow cytometry was used for characterizing VYS cells, and for determining mitochondrial activity, cell proliferation, DNA ploidy, cell cycle phases, and caspase-3 activity. Fetal weight was reduced in the diabetic group. Expression of the cell markers CD34, VEGFR1, CD115, CD117, CD14, CCR2, CD90, CD44, STRO-1, OCT3/4, and Nanog was detected in VYS cells in both groups. In the diabetic group, significantly decreased expression of CD34 (P<0.05), CCR2 (P<0.001), and OCT3/4 (P<0.01), and significantly increased expression of CD90 (P<0.05), CD117 (P<0.01), and CD14 (P<0.05) were observed. VYS cells with inactive mitochondria, activated caspase-3, and low proliferation were present in the rats with diabetes. Severe hyperglycemia caused by maternal diabetes had negative effects on pregnancy, VYS cell viability, and the expression of cell markers. PMID:26176314

  3. Inhibition of Fatty Acid Synthase Decreases Expression of Stemness Markers in Glioma Stem Cells

    PubMed Central

    Yasumoto, Yuki; Miyazaki, Hirofumi; Vaidyan, Linda Koshy; Kagawa, Yoshiteru; Ebrahimi, Majid; Yamamoto, Yui; Ogata, Masaki; Katsuyama, Yu; Sadahiro, Hirokazu; Suzuki, Michiyasu; Owada, Yuji

    2016-01-01

    Cellular metabolic changes, especially to lipid metabolism, have recently been recognized as a hallmark of various cancer cells. However, little is known about the significance of cellular lipid metabolism in the regulation of biological activity of glioma stem cells (GSCs). In this study, we examined the expression and role of fatty acid synthase (FASN), a key lipogenic enzyme, in GSCs. In the de novo lipid synthesis assay, GSCs exhibited higher lipogenesis than differentiated non-GSCs. Western blot and immunocytochemical analyses revealed that FASN is strongly expressed in multiple lines of patient-derived GSCs (G144 and Y10), but its expression was markedly reduced upon differentiation. When GSCs were treated with 20 μM cerulenin, a pharmacological inhibitor of FASN, their proliferation and migration were significantly suppressed and de novo lipogenesis decreased. Furthermore, following cerulenin treatment, expression of the GSC markers nestin, Sox2 and fatty acid binding protein (FABP7), markers of GCSs, decreased while that of glial fibrillary acidic protein (GFAP) expression increased. Taken together, our results indicate that FASN plays a pivotal role in the maintenance of GSC stemness, and FASN-mediated de novo lipid biosynthesis is closely associated with tumor growth and invasion in glioblastoma. PMID:26808816

  4. Expression of Stem Cell Markers in Preinvasive Tubal Lesions of Ovarian Carcinoma

    PubMed Central

    Chene, G.; Ouellet, V.; Rahimi, K.; Barres, V.; Meunier, L.; De Ladurantaye, M.; Provencher, D.; Mes-Masson, A. M.

    2015-01-01

    In order to better understand the ovarian serous carcinogenic process with tubal origin, we investigated the expression of stem cell markers in premalignant tubal lesions (serous tubal intraepithelial carcinoma or STIC). We found an increased stem cell marker density in the normal fallopian tube followed by a high CD117 and a low ALDH and CD44 expression in STICs raising the question of the role of the stem cell markers in the serous carcinogenic process. PMID:26504831

  5. Markers

    ERIC Educational Resources Information Center

    Healthy Schools Network, Inc., 2011

    2011-01-01

    Dry erase whiteboards come with toxic dry erase markers and toxic cleaning products. Dry erase markers labeled "nontoxic" are not free of toxic chemicals and can cause health problems. Children are especially vulnerable to environmental health hazards; moreover, schools commonly have problems with indoor air pollution, as they are more densely…

  6. Differentially expressed protein markers in human submandibular and sublingual secretions.

    PubMed

    Hu, Shen; Denny, Patricia; Denny, Paul; Xie, Yongming; Loo, Joseph A; Wolinsky, Lawrence E; Li, Yang; McBride, Jim; Ogorzalek Loo, Rachel R; Navazesh, Mavash; Wong, David T

    2004-11-01

    Proteome analysis of secretions from individual salivary glands is important for understanding the health of the oral cavity and pathogenesis of certain diseases. However, cross-contamination of submandibular (SM) and sublingual (SL) glandular secretions can occur. The close anatomic relationship of the SM and SL ductal orifices can lead to such contamination. Additionally, these glands may share common ducts. To insure the purity of SM/SL secretions for proteomic analysis, it is important to develop unique biomarkers which could be used to verify the integrity of the individual glandular saliva. In this study, a proteomics approach based on mass spectrometry and gel electrophoresis techniques was utilized to identify and verify a set of proteins (cystatin C, calgranulin B and MUC5B mucin), which are differentially expressed in SM/SL secretions. SM/SL fluids were obtained from nine healthy subjects. Cystatin C was found to be an SM-selective protein as it was found in all SM fluids but not detected in two SL fluids. MUC5B mucin and calgranulin B, on the other hand, were found to be SL-selective proteins. All SL samples contained MUC5B mucin, whereas MUC5B mucin was not detected in four SM samples. Eight of the SL samples contained calgranulin B; however, calgranulin B was absent in eight SM samples. This set of protein markers, especially calgranulin B, can be used to determine the purity of SM/SL samples, and therefore identify potential individuals who do not exhibit cross-contaminated SM/SL secretions, an important requirement for subsequent proteome analysis of pure SM and SL secretions.

  7. A not cytotoxic nickel concentration alters the expression of neuronal differentiation markers in NT2 cells.

    PubMed

    Ceci, Claudia; Barbaccia, Maria Luisa; Pistritto, Giuseppa

    2015-03-01

    Nickel, a known occupational/environmental hazard, may cross the placenta and reach appreciable concentrations in various fetal organs, including the brain. The aim of this study was to investigate whether nickel interferes with the process of neuronal differentiation. Following a 4 week treatment with retinoic acid (10μM), the human teratocarcinoma-derived NTera2/D1 cell line (NT2 cells) terminally differentiate into neurons which recapitulate many features of human fetal neurons. The continuous exposure of the differentiating NT2 cells to a not cytotoxic nickel concentration (10μM) increased the expression of specific neuronal differentiation markers such as neural cell adhesion molecule (NCAM) and microtubule associated protein 2 (MAP2). Furthermore, nickel exposure increased the expression of hypoxia-inducible-factor-1α (HIF-1α) and induced the activation of the AKT/PKB kinase pathway, as shown by the increase of P(Ser-9)-GSK-3β, the inactive form of glycogen synthase kinase-3β (GSK-3β). Intriguingly, by the end of the fourth week the expression of tyrosine hydroxylase (TH) protein, a marker of dopaminergic neurons, was lower in nickel-treated than in control cultures. Thus, likely by partially mimicking hypoxic conditions, a not-cytotoxic nickel concentration appears to alter the process of neuronal differentiation and hinder the expression of the dopaminergic neuronal phenotype. Taken together, these results suggest that nickel, by altering normal brain development, may increase susceptibility to neuro-psychopathology later in life.

  8. Ameliorative potential of gingerol: Promising modulation of inflammatory factors and lipid marker enzymes expressions in HFD induced obesity in rats.

    PubMed

    Brahma Naidu, Parim; Uddandrao, V V Sathibabu; Ravindar Naik, Ramavat; Suresh, Pothani; Meriga, Balaji; Begum, Mustapha Shabana; Pandiyan, Rajesh; Saravanan, Ganapathy

    2016-01-01

    Obesity, generally linked to hyperlipidemia, has been occurring of late with distressing alarm and has now become a global phenomenon casting a huge economic burden on the health care system of countries around the world. The present study investigated the effects of gingerol over 30 days on the changes in HFD-induced obese rats in marker enzymes of lipid metabolism such as fatty-acid synthase (FAS), Acetyl CoA Carboxylase (ACC), Carnitine Palmitoyl Transferase-1(CPT-1), HMG co-A Reductase (HMGR), Lecithin Choline Acyl Transferase (LCAT) and Lipoprotein Lipase (LPL) and inflammatory markers (TNF-α and IL-6). The rats were treated orally with gingerol (75 mg kg(-1)) once daily for 30 days with a lorcaserin-treated group (10 mg kg(-1)) included for comparison. Changes in body weight, glucose, insulin resistance and expressions of lipid marker enzymes and inflammatory markers in tissues were observed in experimental rats. The administration of gingerol resulted in a significant reduction in body weight gain, glucose and insulin levels, and insulin resistance, which altered the activity, expressions of lipid marker enzymes and inflammatory markers. It showed that gingerol had significantly altered these parameters when compared with HFD control rats. This study confirms that gingerol prevents HFD-induced hyperlipidemia by modulating the expression of enzymes important to cholesterol metabolism. PMID:26493465

  9. PDGFRβ Is a Novel Marker of Stromal Activation in Oral Squamous Cell Carcinomas

    PubMed Central

    Han, Rong; Haines, Paul; Gallagher, George; Noonan, Vikki; Kukuruzinska, Maria; Monti, Stefano; Trojanowska, Maria

    2016-01-01

    Carcinoma associated fibroblasts (CAFs) form the main constituents of tumor stroma and play an important role in tumor growth and invasion. The presence of CAFs is a strong predictor of poor prognosis of head and neck squamous cell carcinoma. Despite significant progress in determining the role of CAFs in tumor progression, the mechanisms contributing to their activation remain poorly characterized, in part due to fibroblast heterogeneity and the scarcity of reliable fibroblast surface markers. To search for such markers in oral squamous cell carcinoma (OSCC), we applied a novel approach that uses RNA-sequencing data derived from the cancer genome atlas (TCGA). Specifically, our strategy allowed for an unbiased identification of genes whose expression was closely associated with a set of bona fide stroma-specific transcripts, namely the interstitial collagens COL1A1, COL1A2, and COL3A1. Among the top hits were genes involved in cellular matrix remodeling and tumor invasion and migration, including platelet-derived growth factor receptor beta (PDGFRβ), which was found to be the highest-ranking receptor protein genome-wide. Similar analyses performed on ten additional TCGA cancer datasets revealed that other tumor types shared CAF markers with OSCC, including PDGFRβ, which was found to significantly correlate with the reference collagen expression in ten of the 11 cancer types tested. Subsequent immunostaining of OSCC specimens demonstrated that PDGFRβ was abundantly expressed in stromal fibroblasts of all tested cases (12/12), while it was absent in tumor cells, with greater specificity than other known markers such as alpha smooth muscle actin or podoplanin (3/11). Overall, this study identified PDGFRβ as a novel marker of stromal activation in OSCC, and further characterized a list of promising candidate CAF markers that may be relevant to other carcinomas. Our novel approach provides for a fast and accurate method to identify CAF markers without the need for

  10. PDGFRβ Is a Novel Marker of Stromal Activation in Oral Squamous Cell Carcinomas.

    PubMed

    Kartha, Vinay K; Stawski, Lukasz; Han, Rong; Haines, Paul; Gallagher, George; Noonan, Vikki; Kukuruzinska, Maria; Monti, Stefano; Trojanowska, Maria

    2016-01-01

    Carcinoma associated fibroblasts (CAFs) form the main constituents of tumor stroma and play an important role in tumor growth and invasion. The presence of CAFs is a strong predictor of poor prognosis of head and neck squamous cell carcinoma. Despite significant progress in determining the role of CAFs in tumor progression, the mechanisms contributing to their activation remain poorly characterized, in part due to fibroblast heterogeneity and the scarcity of reliable fibroblast surface markers. To search for such markers in oral squamous cell carcinoma (OSCC), we applied a novel approach that uses RNA-sequencing data derived from the cancer genome atlas (TCGA). Specifically, our strategy allowed for an unbiased identification of genes whose expression was closely associated with a set of bona fide stroma-specific transcripts, namely the interstitial collagens COL1A1, COL1A2, and COL3A1. Among the top hits were genes involved in cellular matrix remodeling and tumor invasion and migration, including platelet-derived growth factor receptor beta (PDGFRβ), which was found to be the highest-ranking receptor protein genome-wide. Similar analyses performed on ten additional TCGA cancer datasets revealed that other tumor types shared CAF markers with OSCC, including PDGFRβ, which was found to significantly correlate with the reference collagen expression in ten of the 11 cancer types tested. Subsequent immunostaining of OSCC specimens demonstrated that PDGFRβ was abundantly expressed in stromal fibroblasts of all tested cases (12/12), while it was absent in tumor cells, with greater specificity than other known markers such as alpha smooth muscle actin or podoplanin (3/11). Overall, this study identified PDGFRβ as a novel marker of stromal activation in OSCC, and further characterized a list of promising candidate CAF markers that may be relevant to other carcinomas. Our novel approach provides for a fast and accurate method to identify CAF markers without the need for

  11. Gene expression profiling of craniofacial fibrous dysplasia reveals ADAMTS2 overexpression as a potential marker.

    PubMed

    Zhou, Shang-Hui; Yang, Wen-Jun; Liu, Sheng-Wen; Li, Jiang; Zhang, Chun-Ye; Zhu, Yun; Zhang, Chen-Ping

    2014-01-01

    Fibrous dysplasia (FD) as an abnormal bone growth is one of the common fibro-osseous leasions (FOL) in oral and maxillofacial region, however, its etiology still remains unclear. Here, we performed gene expression profiling of FD using microarray analysis to explore the key molecule events in FD development, and develop potential diagnostic markers or therapeutic targets for FD. We found that 1,881 genes exhibited differential expression with more than two-fold changes in FD compared to normal bone tissues, including 1,200 upregulated genes and 681 downregulated genes. Pathway analysis indicated that obviously activated pathways are Ribosome and ECM-receptor interaction pathways; downregulated pathways are "Hepatitis C" and "cancer" signaling pathways. We further validated the expression of ADAMTS2, one of most differentiated expressed genes, by Immunohistochemistry (IHC) in 40 of FD cases. Results showed that ADAMTS2 was significantly overexpressed in FD tissues, but rarely expressed in normal bone tissues, suggesting that ADAMTS2 could be a potential biomarker for FD. Thus, this study uncovered differentially expressed candidate genes in FD, which provides pilot data for understanding FD pathogenesis, and developing novel biomarkers for diagnosis and targeting of FD.

  12. Analysis of marker expression in porcine cell lines derived from blastocysts produced in vitro and in vivo.

    PubMed

    Vackova, Irena; Novakova, Zora; Krylov, Vladimir; Okada, Konosuke; Kott, Tomas; Fulka, Helena; Motlik, Jan

    2011-10-01

    The present study was designed to extensively characterize cell lines derived from porcine blastocysts by several methodical approaches, including morphological observation, cytogenetic analysis, estimation of alkaline phosphatase activity and detection of specific marker expression at the mRNA/protein level. A comparison was made between the properties of cell lines isolated from in vivo- and in vitro-obtained blastocysts. Our results showed that 57.1% of the in vivo-obtained blastocysts attached to the feeder layer and that 33.3% of them started to grow in a monolayer. The percentage of attached in vitro-produced blastocysts was lower (24.6%), and only 6.9% of them started to grow. Outgrowths from the in vitro-produced blastocysts formed mainly trophectoderm or epithelial-like monolayer, whereas the in vivo-obtained blastocysts formed heterogeneous outgrowths that also contained cells with embryonic stem (ES)-like morphology. Detailed analyses showed that the primary outgrowths with ES-like morphology expressed the pluripotency markers OCT-4 and NANOG and revealed intensive alkaline phosphatase staining, while they did not express markers of differentiation. The majority of passaged cells, including those with ES-like morphology, lacked OCT-4 protein and revealed expression of specific differentiation markers (cytokeratin 18, lamins A/C, transferrin, α-fetoprotein and GATA-4), although they still expressed NANOG and exhibited weak alkaline phosphatase activity. Moreover, these cells spontaneously differentiated into neural, fibroblast or epithelial-like cells, even in the presence of leukaemia inhibitory factor. Our results show that complex analysis of markers of pluripotency as well as differentiation markers is necessary for proper interpretation of data in porcine embryonic stem cell studies. PMID:21685711

  13. Reassessment of Blood Gene Expression Markers for the Prognosis of Relapsing-Remitting Multiple Sclerosis

    PubMed Central

    Hecker, Michael; Paap, Brigitte Katrin; Goertsches, Robert Hermann; Kandulski, Ole; Fatum, Christian; Koczan, Dirk; Hartung, Hans-Peter; Thiesen, Hans-Juergen; Zettl, Uwe Klaus

    2011-01-01

    Despite considerable advances in the treatment of multiple sclerosis, current drugs are only partially effective. Most patients show reduced disease activity with therapy, but still experience relapses, increasing disability, and new brain lesions. Since there are no reliable clinical or biological markers of disease progression, long-term prognosis is difficult to predict for individual patients. We identified 18 studies that suggested genes expressed in blood as predictive biomarkers. We validated the prognostic value of those genes with three different microarray data sets comprising 148 patients in total. Using these data, we tested whether the genes were significantly differentially expressed between patients with good and poor courses of the disease. Poor progression was defined by relapses and/or increase of disability during a two-year follow-up, independent of the administered therapy. Of 110 genes that have been proposed as predictive biomarkers, most could not be confirmed in our analysis. However, the G protein-coupled membrane receptor GPR3 was expressed at significantly lower levels in patients with poor disease progression in all data sets. GPR3 has therefore a high potential to be a biomarker for predicting future disease activity. In addition, we examined the IL17 cytokines and receptors in more detail and propose IL17RC as a new, promising, transcript-based biomarker candidate. Further studies are needed to better understand the roles of these receptors in multiple sclerosis and its treatment and to clarify the utility of GPR3 and IL17RC expression levels in the blood as markers of long-term prognosis. PMID:22216338

  14. Understanding the Mysterious M2 Macrophage through Activation Markers and Effector Mechanisms

    PubMed Central

    Rőszer, Tamás

    2015-01-01

    The alternatively activated or M2 macrophages are immune cells with high phenotypic heterogeneity and are governing functions at the interface of immunity, tissue homeostasis, metabolism, and endocrine signaling. Today the M2 macrophages are identified based on the expression pattern of a set of M2 markers. These markers are transmembrane glycoproteins, scavenger receptors, enzymes, growth factors, hormones, cytokines, and cytokine receptors with diverse and often yet unexplored functions. This review discusses whether these M2 markers can be reliably used to identify M2 macrophages and define their functional subdivisions. Also, it provides an update on the novel signals of the tissue environment and the neuroendocrine system which shape the M2 activation. The possible evolutionary roots of the M2 macrophage functions are also discussed. PMID:26089604

  15. Lymphocyte Activation Markers in Pediatric Kidney Transplant Recipients

    PubMed Central

    Fadel, Fatina I.; Elghoroury, Eman A.; Elshamaa, Manal F.; Bazaraa, Hafez M.; Salah, Doaa M.; Kassem, Neemat M. A.; Ibrahim, Mona H.; El-Saaid, Gamila S.; Nasr, Soha A.; Koura, Hala M.

    2015-01-01

    Background and objectives: The role of CD4+CD25+ T regulatory cells (Tregs) in immune tolerance in experimental transplantation is very important but the clinical significance of circulating Tregs in the peripheral blood is undetermined. We evaluated the association between the frequency of T cell activation markers CD25 and CD71 and clinical parameters that may affect the level of these T cell markers. Methods: In 47peditric kidney transplant (KT) recipients and 20 healthy controls, the frequency of T cell activation markers, CD25 and CD71 was measured with flow cytometry after transplantation. Two clinical protocols of induction immunosuppression were used: (1) anti-thymocyte globulin (THYMO) group (n =29) and Basiliximab (BSX) group (n=10). Results: The percentage of circulating CD25 after KT was significantly lower than that in the controls. There is no significant difference between KT and the controls s regard to circulating CD71. The percentage of CD25 was significantly increased in children with acute rejection compared with those without acute rejection. Calcineurin inhibitors (CNIs) decreased the frequency of CD25 but mammalian target rapamycin (mTOR) inhibitor did not. The proportion of CD25 significantly decreased in THYMO group during the first year after transplantation. Conclusion: The frequency of circulating T cell activation marker CD25 in pediatric KT recipients is strongly affected by CNIs, and a high frequency of CD25 is associated with acute rejection during the early posttransplant period. The measurement of T cell activation markers, may become a useful immune monitoring tool after kidney transplantation. PMID:26508906

  16. Conservation of spermatogonial stem cell marker expression in undifferentiated felid spermatogonia.

    PubMed

    Vansandt, Lindsey M; Livesay, Janelle L; Dickson, Melissa Joy; Li, Lei; Pukazhenthi, Budhan S; Keefer, Carol L

    2016-09-01

    Spermatogonial stem cells (SSCs) are distinct in their ability to self-renew, transmit genetic information, and persist throughout the life of an individual. These characteristics make SSCs a useful tool for addressing diverse challenges such as efficient transgenic production in nonrodent, biomedical animal models, or preservation of the male genome for species in which survival of frozen-thawed sperm is low. A requisite first step to access this technology in felids is the establishment of molecular markers. This study was designed to evaluate, in the domestic cat (Felis catus), the expression both in situ and following enrichment in vitro of six genes (GFRA1, GPR125, ZBTB16, POU5F1, THY1, and UCHL1) that had been previously identified as SSC markers in other species. Antibodies for surface markers glial cell line-derived neurotrophic factor family receptor alpha 1, G protein-coupled receptor 125, and thymus cell antigen 1 could not be validated, whereas Western blot analysis of prepubertal, peripubertal, and adult cat testis confirmed protein expression for the intracellular markers ubiquitin carboxy-terminal hydrolase 1, zinc finger and BTB domain-containing protein 16, and POU domain, class 5, transcription factor 1. Colocalization of the markers by immunohistochemistry revealed that several cells within the subpopulation adjacent to the basement membrane of the seminiferous tubules and identified morphologically as spermatogonia, expressed all three intracellular markers. Studies performed on cheetah (Acinonyx jubatus) and Amur leopard (Panthera pardus orientalis) testis exhibited a conserved expression pattern in protein molecular weights, relative abundance, and localization of positive cells within the testis. The expression of the three intracellular SSC marker proteins in domestic and wild cat testes confirms conservation of these markers in felids. Enrichment of marker transcripts after differential plating was also observed. These markers will

  17. Vitamin D receptor expression is linked to potential markers of human thyroid papillary carcinoma.

    PubMed

    Izkhakov, Elena; Somjen, Dalia; Sharon, Orli; Knoll, Esther; Aizic, Asaf; Fliss, Dan M; Limor, Rona; Stern, Naftali

    2016-05-01

    Genes regulated cell-cell and cell-matrix adhesion and degradation of the extracellular matrix (ECM) have been screened as potential markers of malignant thyroid nodules. The mRNA expression levels of two of them, the ECM protein-1 (ECM1) and the type II transmembrane serine protease-4 (TMPRSS4), were shown to be an independent predictor of an existing thyroid carcinoma. The vitamin D receptor (VDR) is expressed in epithelial cells of the normal thyroid gland, as well as in malignant dividing cells, which respond to the active metabolite of vitamin D by decreased proliferative activity in vitro. We evaluated the relationship between mRNA gene expressions of TMPRSS4, ECM1 and VDR in 21 papillary thyroid carcinoma samples and compared it to 21 normal thyroid tissues from the same patients. Gene expression was considered as up- or down-regulated if it varied by more or less than 2-fold in the cancer tissue relative to the normal thyroid tissue (Ca/N) from the same patient. We found an overall significant adjusted correlation between the mRNA expression ratio (ExR) of VDR and that of ECM1 in Ca/N thyroid tissue (R=0.648, P<0.001). There was a high ExR of VDR between Ca/N thyroid tissue from the same patient (3.06±2.9), which also exhibited a high Ca/N ExR of ECM1 and/or of TMPRSS4 (>2, P=0.05).The finding that increased VDR expression in human thyroid cancer cells is often linked to increased ECM1 and/or TPMRSS4 expression warrants further investigation into the potential role of vitamin D analogs in thyroid carcinoma.

  18. Pharmacokinetic and pharmacogenetic predictive markers of irinotecan activity and toxicity.

    PubMed

    Di Paolo, Antonello; Bocci, Guido; Polillo, Marialuisa; Del Re, Marzia; Di Desidero, Teresa; Lastella, Marianna; Danesi, Romano

    2011-12-01

    After the rapid development of new classes of antineoplastic drugs, research activities have focused their efforts to the identification of predictive markers of drug activity and tolerability. Irinotecan (CPT-11) may induce severe toxicities (diarrhea, neutropenia) that limit its clinical use, but the increasing knowledge of its pharmacokinetics offered a potential approach to treatment optimization. Pharmacokinetics, the first area of investigation, has identified markers such as biliary index, the relative extent of conversion and the glucuronidation ratio, which are capable to define the risk for severe adverse effects. Because of the existence of some issues concerning the adoption of pharmacokinetic strategies to optimize CPT-11 dose and schedule, analyses of genetic polymorphisms seemed to offer a more reliable and safer approach for the identification of patients at risk than pharmacokinetics. In this view, the uridine diphosphate glucuronosil transferase isoform 1A1 (UGT1A1) was associated with significant changes in disposition of CPT-11 and its metabolites, and consequently with treatment-induced toxicities. However, the complex pharmacokinetics of irinotecan and the involvement of several enzymes other than UGT (i.e., carboxyl estherases, CYP450 isoforms), and transmembrane transporters (ABCB1, ABCC1, ABCG2, SLCO1B1) make difficult the identification of patients with an optimal sensitivity and specificity, and a large part of variability among patients still remains unexplained. Furthermore, prospective clinical studies that should demonstrate the reliability of those pharmacokinetic and pharmacogenetic markers are still lacking. In the present review, pharmacokinetic and pharmacogenetic markers will be discussed. PMID:21787264

  19. Serum inflammatory mediators as markers of human Lyme disease activity.

    PubMed

    Soloski, Mark J; Crowder, Lauren A; Lahey, Lauren J; Wagner, Catriona A; Robinson, William H; Aucott, John N

    2014-01-01

    Chemokines and cytokines are key signaling molecules that orchestrate the trafficking of immune cells, direct them to sites of tissue injury and inflammation and modulate their states of activation and effector cell function. We have measured, using a multiplex-based approach, the levels of 58 immune mediators and 7 acute phase markers in sera derived from of a cohort of patients diagnosed with acute Lyme disease and matched controls. This analysis identified a cytokine signature associated with the early stages of infection and allowed us to identify two subsets (mediator-high and mediator-low) of acute Lyme patients with distinct cytokine signatures that also differed significantly (p<0.0005) in symptom presentation. In particular, the T cell chemokines CXCL9 (MIG), CXCL10 (IP-10) and CCL19 (MIP3B) were coordinately increased in the mediator-high group and levels of these chemokines could be associated with seroconversion status and elevated liver function tests (p = 0.027 and p = 0.021 respectively). There was also upregulation of acute phase proteins including CRP and serum amyloid A. Consistent with the role of CXCL9/CXCL10 in attracting immune cells to the site of infection, CXCR3+ CD4 T cells are reduced in the blood of early acute Lyme disease (p = 0.01) and the decrease correlates with chemokine levels (p = 0.0375). The levels of CXCL9/10 did not relate to the size or number of skin lesions but elevated levels of serum CXCL9/CXCL10 were associated with elevated liver enzymes levels. Collectively these results indicate that the levels of serum chemokines and the levels of expression of their respective chemokine receptors on T cell subsets may prove to be informative biomarkers for Lyme disease and related to specific disease manifestations.

  20. Flow Cytometric Investigation of Classical and Alternative Platelet Activation Markers

    PubMed Central

    Debreceni, Ildikó Beke; Kappelmayer, János

    2013-01-01

    Platelets show a substantial role in the maintenance of vascular integrity when these cells after a rapid activation adhere to the vessel wall lesion, aggregate with other platelets and leukocytes resulting in an arterial thrombosis. Analysis of in vivo platelet activation at an early time point is crucial in the detection of developing thrombotic events. In addition, the forecast of future complications as well as the evaluation of the efficacy of anti- platelet medication are also essential in a large group of patients. Changes in the levels of platelet receptors or alteration in other surface properties due to intra- and extracellular responses to a stimulus can be measurable primarily by flow cytometry with specific antibodies via the assessment of classical and alternative platelet activation markers. Some of these biomarkers have been already used in routine laboratory settings in many cases, while others still stand in the phase of research applications. Deficiency in platelet receptors is also accessible with this technique for the diagnosis of certain bleeding disorders. We here describe the most important types of platelet activation markers, and give an overview how the levels of these markers are altered in different diseases.

  1. Expression vectors for the use of eukaryotic luciferases as bacterial markers with different colors of luminescence.

    PubMed Central

    Cebolla, A; Vázquez, M E; Palomares, A J

    1995-01-01

    An easy way to identify microorganisms is to provide them with gene markers that confer a unique phenotype. Several genetic constructions were developed to use eukaryotic luciferase genes for bacterial tagging. The firefly and click bettle luciferase genes, luc and lucOR, respectively, were cloned under constitutive control and regulated control from different transcriptional units driven by P1, lambda PR, and Ptrc promoters. Comparison of the expression of each gene in Escherichia coli cells from identical promoters showed that bioluminescence produced by luc could be detected luminometrically in a more sensitive manner. In contrast, luminescence from intact lucOR-expressing cells was much more stable and resistant to high temperatures than that from luc-expressing cells. To analyze the behavior of these constructions in other gram-negative bacteria, gene fusions with luc genes were cloned on broad-host-range vectors. Measurements of light emission from Rhizobium meliloti, Agrobacterium tumefaciens, and Pseudomonas putida cells indicated that both luciferases were poorly expressed from P1 in most bacterial hosts. In contrast, the lambda promoter PR yielded constitutively high levels of luciferase expression in all bacterial species tested. PR activity was not regulated by temperature when the thermosensitive repressor cI857 was present in the bacterial species tested, except for E. coli. In contrast, the regulated lacIq-Ptrc::lucOR fusion expression system behaved in a manner similar to that observed in E. coli cells. After IPTG (isopropyl-beta-D-thiogalactopyranoside) induction, this system produced the highest levels of lucOR expression in all bacterial species tested.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7574604

  2. Co-Expression of Cancer Stem Cell Markers Corresponds to a Pro-Tumorigenic Expression Profile in Pancreatic Adenocarcinoma.

    PubMed

    Skoda, Jan; Hermanova, Marketa; Loja, Tomas; Nemec, Pavel; Neradil, Jakub; Karasek, Petr; Veselska, Renata

    2016-01-01

    Pancreatic ductal adenocarcinoma (PDAC) remains one of the most lethal malignancies. Its dismal prognosis is often attributed to the presence of cancer stem cells (CSCs) that have been identified in PDAC using various markers. However, the co-expression of all of these markers has not yet been evaluated. Furthermore, studies that compare the expression levels of CSC markers in PDAC tumor samples and in cell lines derived directly from those tumors are lacking. Here, we analyzed the expression of putative CSC markers-CD24, CD44, epithelial cell adhesion molecule (EpCAM), CD133, and nestin-by immunofluorescence, flow cytometry and quantitative PCR in 3 PDAC-derived cell lines and by immunohistochemistry in 3 corresponding tumor samples. We showed high expression of the examined CSC markers among all of the cell lines and tumor samples, with the exception of CD24 and CD44, which were enriched under in vitro conditions compared with tumor tissues. The proportions of cells positive for the remaining markers were comparable to those detected in the corresponding tumors. Co-expression analysis using flow cytometry revealed that CD24+/CD44+/EpCAM+/CD133+ cells represented a significant population of the cells (range, 43 to 72%) among the cell lines. The highest proportion of CD24+/CD44+/EpCAM+/CD133+ cells was detected in the cell line derived from the tumor of a patient with the shortest survival. Using gene expression profiling, we further identified the specific pro-tumorigenic expression profile of this cell line compared with the profiles of the other two cell lines. Together, CD24+/CD44+/EpCAM+/CD133+ cells are present in PDAC cell lines derived from primary tumors, and their increased proportion corresponds with a pro-tumorigenic gene expression profile. PMID:27414409

  3. Macrophage Activation Syndrome-Associated Markers in Severe Dengue

    PubMed Central

    Ab-Rahman, Hasliana Azrah; Rahim, Hafiz; AbuBakar, Sazaly; Wong, Pooi-Fong

    2016-01-01

    Hemophagocytosis, a phenomenon of which activated macrophages phagocytosed hematopoietic elements was reportedly observed in severe dengue patients. In the present study, we investigated whether markers of macrophage activation syndrome (MAS) can be used as differential diagnostic markers of severe dengue. Two hundred and eight confirmed dengue patients were recruited for the study. Sandwich ELISA was used to determine serum ferritin, soluble CD163 (sCD163), and soluble CD25 (sCD25) levels. The population of circulating CD163 (mCD163) monocytes was determined using flow cytometry. Receiver operating characteristic (ROC) analysis was plotted to determine the predictive validity of the biomarkers. Serum ferritin and sCD163 were found significantly increased in severe dengue patients compared to dengue fever patients (P = 0.003). A fair area under ROC curves (AUC) at 0.72 with a significant P value of 0.004 was observed for sCD163. sCD25 and mCD163 levels were not significantly different between severe dengue and dengue fever patients. Our findings suggest that in addition to serum ferritin, sCD163 can differentiate severe dengue from that of dengue fever patients. Hence, sCD163 level can be considered for use as a predictive marker for impending severe dengue. PMID:26941578

  4. Expression of Synaptic and Phototransduction Markers During Photoreceptor Development in the Marmoset Monkey Callithrix jacchus

    PubMed Central

    HENDRICKSON, ANITA; TROILO, DAVID; DJAJADI, HIDAYAT; POSSIN, DANIEL; SPRINGER, ALAN

    2009-01-01

    Marmoset photoreceptor development was studied to determine the expression sequence for synaptic, opsin, and phototransduction proteins. All markers appear first in cones within the incipient foveal center or in rods at the foveal edge. Recoverin appears in cones across 70% of the retina at fetal day (Fd) 88, indicating that it is expressed shortly after photoreceptors are generated. Synaptic markers synaptophysin, SV2, glutamate vesicular transporter 1, and CTBP2 label foveal cones at Fd 88 and cones at the retinal edge around birth. Cones and rods have distinctly different patterns of synaptic protein and opsin expression. Synaptic markers are expressed first in cones, with a considerable delay before they appear in rods at the same eccentricity. Cones express synaptic markers 2–3 weeks before they express opsin, but rods express opsin 2–4 weeks before rod synaptic marker labeling is detected. Medium/long-wavelength-selective (M&L) opsin appears in foveal cones and rod opsin in rods around the fovea at Fd 100. Very few cones expressing short-wavelength-selective (S) opsin are found in the Fd 105 fovea. Across peripheral retina, opsin appears first in rods, followed about 1 week later by M&L cone opsin. S cone opsin appears last, and all opsins reach the retinal edge by 1 week after birth. Cone transducin and rod arrestin are expressed concurrently with opsin, but cone arrestin appears slightly later. Marmoset photoreceptor development differs from that in Macaca and humans. It starts relatively late, at 56% gestation, compared with Macaca at 32% gestation. The marmoset opsin expression sequence is also different from that of either Macaca or human. PMID:19003975

  5. Nocodazole treatment decreases expression of pluripotency markers Nanog and Oct4 in human embryonic stem cells.

    PubMed

    Kallas, Ade; Pook, Martin; Maimets, Martti; Zimmermann, Külli; Maimets, Toivo

    2011-01-01

    Nocodazole is a known destabiliser of microtubule dynamics and arrests cell-cycle at the G2/M phase. In the context of the human embryonic stem cell (hESC) it is important to understand how this arrest influences the pluripotency of cells. Here we report for the first time the changes in the expression of transcription markers Nanog and Oct4 as well as SSEA-3 and SSEA-4 in human embryonic cells after their treatment with nocodazole. Multivariate permeabilised-cell flow cytometry was applied for characterising the expression of Nanog and Oct4 during different cell cycle phases. Among untreated hESC we detected Nanog-expressing cells, which also expressed Oct4, SSEA-3 and SSEA-4. We also found another population expressing SSEA-4, but without Nanog, Oct4 and SSEA-3 expression. Nocodazole treatment resulted in a decrease of cell population positive for all four markers Nanog, Oct4, SSEA-3, SSEA-4. Nocodazole-mediated cell-cycle arrest was accompanied by higher rate of apoptosis and upregulation of p53. Twenty-four hours after the release from nocodazole block, the cell cycle of hESC normalised, but no increase in the expression of transcription markers Nanog and Oct4 was detected. In addition, the presence of ROCK-2 inhibitor Y-27632 in the medium had no effect on increasing the expression of pluripotency markers Nanog and Oct4 or decreasing apoptosis or the level of p53. The expression of SSEA-3 and SSEA-4 increased in Nanog-positive cells after wash-out of nocodazole in the presence and in the absence of Y-27632. Our data show that in hESC nocodazole reversible blocks cell cycle, which is accompanied by irreversible loss of expression of pluripotency markers Nanog and Oct4.

  6. Keratins 17 and 19 expression as prognostic markers in oral squamous cell carcinoma.

    PubMed

    Coelho, B A; Peterle, G T; Santos, M; Agostini, L P; Maia, L L; Stur, E; Silva, C V M; Mendes, S O; Almança, C C J; Freitas, F V; Borçoi, A R; Archanjo, A B; Mercante, A M C; Nunes, F D; Carvalho, M B; Tajara, E H; Louro, I D; Silva-Conforti, A M A

    2015-11-25

    Five-year survival rates for oral squamous cell carcinoma (OSCC) are 30% and the mortality rate is 50%. Immunohistochemistry panels are used to evaluate proliferation, vascularization, apoptosis, HPV infection, and keratin expression, which are important markers of malignant progression. Keratins are a family of intermediate filaments predominantly expressed in epithelial cells and have an essential role in mechanical support and cytoskeleton formation, which is essential for the structural integrity and stability of the cell. In this study, we analyzed the expressions of keratins 17 and 19 (K17 and K19) by immunohistochemistry in tumoral and non-tumoral tissues from patients with OSCC. The results show that expression of these keratins is higher in tumor tissues compared to non-tumor tissues. Positive K17 expression correlates with lymph node metastasis and multivariate analysis confirmed this relationship, revealing a 6-fold increase in lymph node metastasis when K17 is expressed. We observed a correlation between K17 expression with disease-free survival and disease-specific death in patients who received surgery and radiotherapy. Multivariate analysis revealed that low expression of K17 was an independent marker for early disease relapse and disease-specific death in patients treated with surgery and radiotherapy, with an approximately 4-fold increased risk when compared to high K17 expression. Our results suggest a potential role for K17 and K19 expression profiles as tumor prognostic markers in OSCC patients.

  7. Co-Expression of Cancer Stem Cell Markers Corresponds to a Pro-Tumorigenic Expression Profile in Pancreatic Adenocarcinoma

    PubMed Central

    Skoda, Jan; Hermanova, Marketa; Loja, Tomas; Nemec, Pavel; Neradil, Jakub; Karasek, Petr; Veselska, Renata

    2016-01-01

    Pancreatic ductal adenocarcinoma (PDAC) remains one of the most lethal malignancies. Its dismal prognosis is often attributed to the presence of cancer stem cells (CSCs) that have been identified in PDAC using various markers. However, the co-expression of all of these markers has not yet been evaluated. Furthermore, studies that compare the expression levels of CSC markers in PDAC tumor samples and in cell lines derived directly from those tumors are lacking. Here, we analyzed the expression of putative CSC markers—CD24, CD44, epithelial cell adhesion molecule (EpCAM), CD133, and nestin—by immunofluorescence, flow cytometry and quantitative PCR in 3 PDAC-derived cell lines and by immunohistochemistry in 3 corresponding tumor samples. We showed high expression of the examined CSC markers among all of the cell lines and tumor samples, with the exception of CD24 and CD44, which were enriched under in vitro conditions compared with tumor tissues. The proportions of cells positive for the remaining markers were comparable to those detected in the corresponding tumors. Co-expression analysis using flow cytometry revealed that CD24+/CD44+/EpCAM+/CD133+ cells represented a significant population of the cells (range, 43 to 72%) among the cell lines. The highest proportion of CD24+/CD44+/EpCAM+/CD133+ cells was detected in the cell line derived from the tumor of a patient with the shortest survival. Using gene expression profiling, we further identified the specific pro-tumorigenic expression profile of this cell line compared with the profiles of the other two cell lines. Together, CD24+/CD44+/EpCAM+/CD133+ cells are present in PDAC cell lines derived from primary tumors, and their increased proportion corresponds with a pro-tumorigenic gene expression profile. PMID:27414409

  8. Ets-1 messenger RNA expression is a novel marker of poor survival in ovarian carcinoma.

    PubMed

    Davidson, B; Reich, R; Goldberg, I; Gotlieb, W H; Kopolovic, J; Berner, A; Ben-Baruch, G; Bryne, M; Nesland, J M

    2001-03-01

    Ets-1 proto-oncogene is a transcription factor involved in several cellular functions, including the activation of several proteases participating in tumor invasion and metastasis. The objective of this study was to analyze the possible correlation between Ets-1 mRNA expression and survival in advanced-stage ovarian carcinomas, studying two patient groups with extremely different disease outcome. Sections from 66 primary ovarian carcinomas and metastatic lesions from 41 patients diagnosed with advanced-stage ovarian carcinoma (International Federation of Gynecologists and Obstetricians stages III and IV) were evaluated for expression of Ets-1 using mRNA in situ hybridization. Patients were divided into long-term (n = 17) and short-term (n = 24) survivors. The mean values for disease-free survival and overall survival were 116 and 133 months for long-term survivors, as compared to 3 and 21 months for short-term survivors, respectively. Expression of Ets-1 mRNA was detected in carcinoma cells and stromal cells in 28 of 66 (42%) and 22 of 66 (33%) lesions, respectively. Ets-1 expression showed an association with mRNA expression of vascular endothelial growth factor (P = 0.001 for carcinoma cells; P = 0.004 for stromal cells), basic fibroblast growth factor (P = 0.049 for carcinoma cells), and membrane type-1 matrix metalloproteinase (P = 0.045), which were previously studied in this patient cohort. Ets-1 mRNA was detected more often in both carcinoma and stromal cells in tumors of short-term survivors (P = 0.038 for carcinoma cells). In univariate survival analysis for all cases, Ets-1 expression in both tumor (P = 0.018) and stroma (P = 0.026) correlated with poor survival. These findings were reproduced in an analysis of primary tumors alone (P = 0.039 for tumor cells; P < 0.001 for stromal cells). Ets-1 mRNA expression in stromal cells retained its predictive power in a multivariate survival analysis in which all molecules studied previously in this patient cohort

  9. Ex vivo expanded SSEA-4+ human limbal stromal cells are multipotent and do not express other embryonic stem cell markers

    PubMed Central

    Hussin, Noor Hamidah; Othman, Ainoon; Umapathy, Thiageswari; Baharuddin, Puteri; Jamal, Rahman; Zakaria, Zubaidah

    2012-01-01

    Purpose The presence of multipotent human limbal stromal cells resembling mesenchymal stromal cells (MSC) provides new insights to the characteristic of these cells and its therapeutic potential. However, little is known about the expression of stage-specific embryonic antigen 4 (SSEA-4) and the embryonic stem cell (ESC)-like properties of these cells. We studied the expression of SSEA-4 surface protein and the various ESC and MSC markers in the ex vivo cultured limbal stromal cells. The phenotypes and multipotent differentiation potential of these cells were also evaluated. Methods Limbal stromal cells were derived from corneoscleral rims. The SSEA-4+ and SSEA-4- limbal stromal cells were sorted by fluorescence-activated cells sorting (FACS). Isolated cells were expanded and reanalyzed for their expression of SSEA-4. Expression of MSC and ESC markers on these cells were also analyzed by FACS. In addition, expression of limbal epithelial and corneal stromal proteins such as ATP-binding cassette sub-family G member 2 (ABCG2), tumour protein p63 (p63), paired box 6 (Pax6), cytokeratin 3 (AE5), cytokeratin 10, and keratocan sulfate were evaluated either by immunofluorecence staining or reverse transcription polymerase chain reaction. Appropriate induction medium was used to differentiate these cells into adipocytes, osteocytes, and chondrocytes. Results Expanded limbal stromal cells expressed the majority of mesenchymal markers. These cells were negative for ABCG2, p63, Pax6, AE-5, and keratocan sulfate. After passaged, a subpopulation of these cells showed low expression of SSEA-4 but were negative for other important ESC surface markers such as Tra-1–60, Tra-1–81, and transcription factors like octamer-binding transcription factor 4 (Oct4), SRY(sex determining region Y)-box 2 (Sox2), and Nanog. Early passaged cells when induced were able to differentiate into adipocytes, osteocytes and chondrocytes. Conclusions The expanded limbal stromal cells showed features

  10. Markers for host-induced gene expression in Trichophyton dermatophytosis.

    PubMed

    Kaufman, Gil; Berdicevsky, Israela; Woodfolk, Judith A; Horwitz, Benjamin A

    2005-10-01

    Dermatophytes are adapted to infect keratinized tissues by their ability to utilize keratin as a nutrient source. Although there have been numerous reports that dermatophytes like Trichophyton sp. secrete proteolytic enzymes, virtually nothing is known about the patterns of gene expression in the host or even when the organisms are cultured on protein substrates in the absence of a host. We characterized the expression of an aminopeptidase gene, the Trichophyton mentagrophytes homolog of the Trichophyton rubrum Tri r 4 gene. The T. rubrum gene was originally isolated based on the ability of the protein encoded by it to induce immediate and delayed-type hypersensitivity in skin tests. T. mentagrophytes Tri m 4 is closely related to Tri r 4 (almost 94% identity at the protein level). Tri m 4 resembles other protease-encoding genes thought to be virulence factors (for example, DPP V of Aspergillus fumigatus). The Tri m 4 protein was detected immunochemically both in fungal extracts and in the culture medium. Expression of the Tri m 4 gene was induced severalfold when T. mentagrophytes was grown on keratin and elastin. Ex vivo, strong induction was observed after culture on blood plasma, but the use of homogenized skin did not result in a significant increase in Tri m 4 transcript levels. In order to identify additional genes encoding putative virulence factors, differential cDNA screening was performed. By this method, a fungal thioredoxin and a cellulase homolog were identified, and both genes were found to be strongly induced by skin extracellular matrix proteins. Induction by superficial (keratin) and deep (elastin) skin elements suggests that the products of these genes may be important in both superficial and deep dermatophytosis, and models for their function are proposed. Upregulation of several newly identified T. mentagrophytes genes on protein substrates suggests that these genes encode proteins which are relevant to the dermatophyte-skin interaction.

  11. Expression and localization of epithelial stem cell and differentiation markers in equine skin, eye and hoof

    PubMed Central

    Linardi, Renata L.; Megee, Susan O.; Mainardi, Sarah R.; Senoo, Makoto; Galantino-Homer, Hannah L.

    2015-01-01

    Background The limited characterization of equine skin, eye and hoof epithelial stem cell (ESC) and differentiation markers impedes the investigation of the physiology and pathophysiology of these tissues. Hypothesis/Objectives To characterize ESC and differentiation marker expression in epithelial tissues of the equine eye, haired skin and hoof capsule. Methods Indirect immunofluorescence microscopy and immunoblotting were utilized to detect expression and tissue localization of keratin (K) isoforms K3, K10, K14, and K124, the transcription factor p63 (a marker of ESCs) and phosphorylated p63 (pp63, a marker of ESC to transit-amplifying (TA) cell transition) in epithelial tissues of the foot (haired skin, hoof coronet and hoof lamellae) and the eye (limbus and cornea). Results K14 expression was restricted to the basal layer of epidermal lamellae, and to basal and adjacent suprabasal layers of the haired skin, coronet and corneal limbus. Coronary and lamellar epidermis was negative for both K3 and K10, which were expressed in the cornea/limbus epithelium and haired skin epidermis, respectively. Variable expression of p63 with relatively low to high levels of phosphorylation was detected in individual basal and suprabasal cells of all epithelial tissues examined. Conclusions This is the first report of the characterization of tissue-specific keratin marker expression and the localization of putative epithelial progenitor cell populations, including ESCs (high p63 expression with low pp63 levels) and TA cells (high expression of both p63 and pp63), in the horse. These results will aid further investigation of epidermal and corneal epithelial biology and regenerative therapies in horses. PMID:25963063

  12. Abnormal bone mineral density and bone turnover marker expression profiles in patients with primary spontaneous pneumothorax

    PubMed Central

    Yu, Lixin; Hou, Shengcai; Hu, Bin; Zhao, Liqiang; Miao, Jinbai; Wang, Yang; Li, Tong; Zhang, Zhenkui; You, Bin; Pang, Baosen; Liang, Yufang; Zhao, Yi; Hao, Wei

    2016-01-01

    Background To examine the bone mineral density (BMD) and the role of bone biomarkers, including bone formation marker procollagen type I aminoterminal propeptide (PINP) and N-terminal midmolecule fragment osteocalcin (N-MID), bone resorption marker b-C-telopeptides of type I collagen (b-CTX) and tartrate-resistant acid phosphatase 5b (TRACP5b) in the pathogenesis of PSP. Methods Eighty-three consecutive primary spontaneous pneumothorax (PSP) patients (PSP group) and 87 healthy individuals (control group) were enrolled in this study. General data, including gender, age, height, weight, and body mass index (BMI), were recorded. Dual-energy X-ray absorptiometry, electrochemiluminescence immunoassay (ECLIA), and ELISA were used to evaluate bone mineral density and expression levels of bone metabolism markers, including PINP, b-CTX, TRACP5b, N-MID, and 25-hydroxyvitamin D (25-OH VD). Results Mean height was significantly greater in the PSP group compared with the control group, whereas weight and BMI were lower. Patients in the PSP group had significantly lower average bone mineral density, which mainly manifested as osteopenia (11/12, 91.7%); however, only one patient (8.3%) developed osteoporosis. Serum overexpression of PINP, b-CTX, TRACP5b, and N-MID were found in PSP patients. Expression of 25-OH VD was low in PSP patients. Bone resorption markers showed positive linear relationships with bone formation markers in all participants; whereas only TRACP5b expression negatively correlated with 25-OH VD. Expression levels of all bone turnover markers negatively correlated with BMI. Regression analysis identified risk factors of PSP as age, height, weight, and TRACP5b and 25-OH VD expression levels; whereas gender and PINP, b-CTX, and N-MID expression levels were not significantly associated with the onset of PSP. Conclusions It had lower bone mineral density in PSP patients. Bone formation marker PINP, N-MID and bone resorption marker b-CTX, TRACP5b were upregulated in

  13. Expression and Activity of Metalloproteinases in Depression

    PubMed Central

    Bobińska, Kinga; Szemraj, Janusz; Czarny, Piotr; Gałecki, Piotr

    2016-01-01

    Background Depression is one of the most common mental disorders and often co-exists with somatic diseases. The most probable cause of comorbidity is a generalized inflammatory process that occurs in both depression and somatic diseases. Matrix metalloproteinases MMPs play a role in modulating inflammation and their impact in many inflammatory diseases has been investigated. The purpose of this study was to evaluate gene expression for selected polymorphisms of MMP-2 (C-735T), MMP-7 (A-181G), and MMP-9 (T-1702A, C1562T), which have been confirmed to participate in development of depression, and TIMP-2 (G-418C, tissue inhibitor of MMP). Activity variability of pro-MMP-2 and pro-MMP-9 was measured in a group of people with depression and a group of healthy individuals. Material/Methods The examined population comprised 142 individuals suffering from depression and 100 individuals who formed a control group (CG). Designations were carried out for MMP-2 (C-735T), MMP-7 (A-181G), MMP-9 (T-1702A, C1562T), and TIMP-2 (G-418C). Results For all examined and tested MMPs and for TIMP-2, gene expression at the mRNA level was higher in patients with depression than in the CG. Similar results were recorded for gene expression at the protein level, while expression on the protein level for TIMP-2 was higher in the CG. Change in activity of MMP-2 and pro-MMP-2 was statistically more significant in the group with depression. The opposite result was recorded for MMP-9 and pro-MMP-9, in which the change in activity was statistically more significant in the CG. Conclusions Changes in MMPs and TIMP expression may be a common element in, or perhaps even a marker for, recurrent depressive disorders and somatic diseases. PMID:27098106

  14. Cells Isolated from Inflamed Periapical Tissue Express Mesenchymal Stem Cell Markers and Are Highly Osteogenic

    PubMed Central

    Liao, James; Al Shahrani, Mohammed; Al-Habib, Mey; Tanaka, Toshinori; Huang, George T.-J.

    2012-01-01

    Introduction We previously reported the presence of mesenchymal stem/progenitor cells (MSCs) in inflamed pulp tissue. Here we asked whether MSCs also exist in inflamed periapical tissues resulting from endodontic infection. The objectives of this study were to detect the expression of MSC markers in periapical inflammatory tissues and to characterize isolated cells from these tissues. Methods Human periapical inflammatory tissues were collected and processed to detect MSC marker expression by immunohistochemistry. Cells were isolated and tested for cell surface marker expression by using flow cytometry and examined for multiple differentiation potential into osteogenic and adipogenic pathways. In vivo formation of mineralized tissues was assessed in a mouse model. Results Immunohistochemistry showed positive staining for MSC markers STRO-1, CD90, and CD146. Isolated cells at passage 0 appeared as typical fibroblastic cells, and a few cells formed colony-forming unit-fibroblasts (CFU-Fs). After passaging, the CFU-F forming ability diminished dramatically, and the population doubling was up to 26. Flow cytometry data showed that these cells at passage 2 expressed low levels of STRO-1 and CD146 and moderate to high levels of CD90, CD73, and CD105. At passage 6, the levels of these markers decreased. When incubated in specific differentiation medium, cells demonstrated a strong osteogenic but weak adipogenic capacity. After in vivo cell transplantation, mineralized tissues formed in immunocompromised mice. Conclusions Human periapical inflammatory tissues expressed MSC markers, suggesting the presence of MSCs. Isolated cells exhibited typical mesenchymal cell immunophenotype with a capacity to form mineralized matrix in vitro and in vivo. PMID:21846537

  15. Myrtucommulone-A treatment decreases pluripotency- and multipotency-associated marker expression in bladder cancer cell line HTB-9.

    PubMed

    Iskender, Banu; Izgi, Kenan; Karaca, Halit; Canatan, Halit

    2015-10-01

    Cancer and stem cells exhibit similar features, including self-renewal, differentiation and immortality. The expression of stem-cell-related genes in cancer cells is demonstrated to be potentially correlated with cancer cell behaviour, affecting both drug response and tumor recurrence. There is an emerging body of evidence that subpopulations of tumors carry a distinct molecular sign and are selectively resistant to chemotherapy. Therefore, it is important to find novel therapeutic agents that could suppress the stem-like features of cancer cells while inhibiting their proliferation. Myrtucommulone-A (MC-A) is an active compound of a nonprenylated acylphloroglucinol isolated from the leaves of myrtle. Here we have investigated the potential of MC-A in inhibiting the expression of self-renewal regulatory factors and cancer stem cell markers in a bladder cancer cell line HTB-9. We used RT-PCR, immunocytochemistry, flow cytometry and western blotting to examine the expression of pluripotency- and multipotency-associated markers with or without treatment with MC-A. Treatment with MC-A not only decreased cancer cell viability and proliferation but also resulted in a decrease in the expression of pluripotency- and multipotency-associated markers such as NANOG, OCT-4, SOX-2, SSEA-4, TRA-1-60, CD90, CD73 and CD44. MC-A treatment was also observed to decrease the sphere-forming ability of HTB-9 cells. In summary, this study provides valuable information on the presence of stem-cell marker expression in HTB-9 cells and our results imply that MC-A could be utilized to target cancer cells with stem-like characteristics. PMID:26054707

  16. ABCG2 regulates self-renewal and stem cell marker expression but not tumorigenicity or radiation resistance of glioma cells.

    PubMed

    Wee, Boyoung; Pietras, Alexander; Ozawa, Tatsuya; Bazzoli, Elena; Podlaha, Ondrej; Antczak, Christophe; Westermark, Bengt; Nelander, Sven; Uhrbom, Lene; Forsberg-Nilsson, Karin; Djaballah, Hakim; Michor, Franziska; Holland, Eric C

    2016-01-01

    Glioma cells with stem cell traits are thought to be responsible for tumor maintenance and therapeutic failure. Such cells can be enriched based on their inherent drug efflux capability mediated by the ABC transporter ABCG2 using the side population assay, and their characteristics include increased self-renewal, high stem cell marker expression and high tumorigenic capacity in vivo. Here, we show that ABCG2 can actively drive expression of stem cell markers and self-renewal in glioma cells. Stem cell markers and self-renewal was enriched in cells with high ABCG2 activity, and could be specifically inhibited by pharmacological and genetic ABCG2 inhibition. Importantly, despite regulating these key characteristics of stem-like tumor cells, ABCG2 activity did not affect radiation resistance or tumorigenicity in vivo. ABCG2 effects were Notch-independent and mediated by diverse mechanisms including the transcription factor Mef. Our data demonstrate that characteristics of tumor stem cells are separable, and highlight ABCG2 as a potential driver of glioma stemness. PMID:27456282

  17. ABCG2 regulates self-renewal and stem cell marker expression but not tumorigenicity or radiation resistance of glioma cells

    PubMed Central

    Wee, Boyoung; Pietras, Alexander; Ozawa, Tatsuya; Bazzoli, Elena; Podlaha, Ondrej; Antczak, Christophe; Westermark, Bengt; Nelander, Sven; Uhrbom, Lene; Forsberg-Nilsson, Karin; Djaballah, Hakim; Michor, Franziska; Holland, Eric C.

    2016-01-01

    Glioma cells with stem cell traits are thought to be responsible for tumor maintenance and therapeutic failure. Such cells can be enriched based on their inherent drug efflux capability mediated by the ABC transporter ABCG2 using the side population assay, and their characteristics include increased self-renewal, high stem cell marker expression and high tumorigenic capacity in vivo. Here, we show that ABCG2 can actively drive expression of stem cell markers and self-renewal in glioma cells. Stem cell markers and self-renewal was enriched in cells with high ABCG2 activity, and could be specifically inhibited by pharmacological and genetic ABCG2 inhibition. Importantly, despite regulating these key characteristics of stem-like tumor cells, ABCG2 activity did not affect radiation resistance or tumorigenicity in vivo. ABCG2 effects were Notch-independent and mediated by diverse mechanisms including the transcription factor Mef. Our data demonstrate that characteristics of tumor stem cells are separable, and highlight ABCG2 as a potential driver of glioma stemness. PMID:27456282

  18. Chondrocyte Culture in Three Dimensional Alginate Sulfate Hydrogels Promotes Proliferation While Maintaining Expression of Chondrogenic Markers

    PubMed Central

    Mhanna, Rami; Kashyap, Aditya; Palazzolo, Gemma; Vallmajo-Martin, Queralt; Becher, Jana; Möller, Stephanie; Schnabelrauch, Matthias

    2014-01-01

    The loss of expression of chondrogenic markers during monolayer expansion remains a stumbling block for cell-based treatment of cartilage lesions. Here, we introduce sulfated alginate hydrogels as a cartilage biomimetic biomaterial that induces cell proliferation while maintaining the chondrogenic phenotype of encapsulated chondrocytes. Hydroxyl groups of alginate were converted to sulfates by incubation with sulfur trioxide–pyridine complex (SO3/pyridine), yielding a sulfated material cross-linkable with calcium chloride. Passage 3 bovine chondrocytes were encapsulated in alginate and alginate sulfate hydrogels for up to 35 days. Cell proliferation was five-fold higher in alginate sulfate compared with alginate (p=0.038). Blocking beta1 integrins in chondrocytes within alginate sulfate hydrogels significantly inhibited proliferation (p=0.002). Sulfated alginate increased the RhoA activity of chondrocytes compared with unmodified alginate, an increase that was blocked by β1 blocking antibodies (p=0.017). Expression and synthesis of type II collagen, type I collagen, and proteoglycan was not significantly affected by the encapsulation material evidenced by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry. Alginate sulfate constructs showed an opaque appearance in culture, whereas the unmodified alginate samples remained translucent. In conclusion, alginate sulfate provides a three dimensional microenvironment that promotes both chondrocyte proliferation and maintenance of the chondrogenic phenotype and represents an important advance for chondrocyte-based cartilage repair therapies providing a material in which cell expansion can be done in situ. PMID:24320935

  19. Differential Expression of Complement Markers in Normal and AMD Transmitochondrial Cybrids

    PubMed Central

    Nashine, Sonali; Chwa, Marilyn; Kazemian, Mina; Thaker, Kunal; Lu, Stephanie; Nesburn, Anthony; Kuppermann, Baruch D.; Kenney, M. Cristina

    2016-01-01

    Purpose Variations in mitochondrial DNA (mtDNA) and abnormalities in the complement pathways have been implicated in the pathogenesis of age-related macular degeneration (AMD). This study was designed to determine the effects of mtDNA from AMD subjects on the complement pathway. Methods Transmitochondrial cybrids were prepared by fusing platelets from AMD and age-matched Normal subjects with Rho0 (lacking mtDNA) human ARPE-19 cells. Quantitative PCR and Western blotting were performed to examine gene and protein expression profiles, respectively, of complement markers in these cybrids. Bioenergetic profiles of Normal and AMD cybrids were examined using the Seahorse XF24 flux analyzer. Results Significant decreases in the gene and protein expression of complement inhibitors, along with significantly higher levels of complement activators, were found in AMD cybrids compared to Older-Normal cybrids. Seahorse flux data demonstrated that the bioenergetic profiles for Older-Normal and Older-AMD cybrid samples were similar to each other but were lower compared to Young-Normal cybrid samples. Conclusion In summary, since all cybrids had identical nuclei and differed only in mtDNA content, the observed changes in components of complement pathways can be attributed to mtDNA variations in the AMD subjects, suggesting that mitochondrial genome and retrograde signaling play critical roles in this disease. Furthermore, the similar bioenergetic profiles of AMD and Older-Normal cybrids indicate that the signaling between mitochondria and nuclei are probably not via a respiratory pathway. PMID:27486856

  20. Immunohistochemical Expression and Clinical Significance of Suggested Stem Cell Markers in Hepatocellular Carcinoma

    PubMed Central

    Sung, Jong Jin; Noh, Sang Jae; Bae, Jun Sang; Park, Ho Sung; Jang, Kyu Yun; Chung, Myoung Ja; Moon, Woo Sung

    2016-01-01

    Background: Increasing evidence has shown that tumor initiation and growth are nourished by a small subpopulation of cancer stem cells (CSCs) within the tumor mass. CSCs are posited to be responsible for tumor maintenance, growth, distant metastasis, and relapse after curative operation. We examined the expression of CSC markers in paraffin-embedded tissue sections of hepatocellular carcinoma (HCC) and correlated the results with clinicopathologic characteristics. Methods: Immunohistochemical staining for the markers believed to be expressed in the CSCs, including epithelial cell adhesion molecule (EpCAM), keratin 19 (K19), CD133, and CD56, was performed in 82 HCC specimens. Results: EpCAM expression was observed in 56% of the HCCs (46/82) and K19 in 6% (5/82). EpCAM expression in HCC significantly correlated with elevated α-fetoprotein level, microvessel invasion of tumor cells, and high histologic grade. In addition, EpCAM expression significantly correlated with K19 expression. The overall survival and relapsefree survival rates in patients with EpCAM-expressing HCC were relatively lower than those in patients with EpCAM-negative HCC. All but two of the 82 HCCs were negative for CD133 and CD56, respectively. Conclusions: Our results suggest that HCCs expressing EpCAM are associated with unfavorable prognostic factors and have a more aggressive clinical course than those not expressing EpCAM. Further, the expression of either CD133 or CD56 in paraffin-embedded HCC tissues appears to be rare. PMID:26581206

  1. Pleomorphic adenoma and adenoid cystic carcinoma: in vitro study of the impact of TGFbeta1 on the expression of integrins and cytoskeleton markers of cell differentiation.

    PubMed

    Lourenço, Silvia Vanessa; Lima, Dirce Mary C

    2007-06-01

    Pleomorphic adenoma (PA) and adenoid cystic carcinoma (ACC) are the commonest benign and malignant salivary gland tumours respectively. Interactions between cells and extracellular matrix of PA and ACC, partially mediated by integrins, are important in their biology. The expression of integrins is regulated by numerous factors, amongst them, transforming growth factor beta1 (TGFbeta1). Our study investigated the effects of TGFbeta1 on the expression of integrin beta subunits in vitro and on the expression of cytoskeletal proteins of cells derived from PA and ACC. The expression of cytoskeletal differentiation markers and integrins was assessed using immunofluorescence. ELISA assays were employed to quantitate the expression integrins and MTT assays evaluated the mitochondrial activity of cells stimulated with TGFbeta1. PA cells showed increased expression of integrins and de novo expression of differentiation markers upon TGFbeta1 stimulation. ACC cells were less responsive to such stimulation. This may reflect important differences in the biological behaviour of benign and malignant cells.

  2. Nuclear YB-1 expression as a negative prognostic marker in nonsmall cell lung cancer.

    PubMed

    Gessner, C; Woischwill, C; Schumacher, A; Liebers, U; Kuhn, H; Stiehl, P; Jürchott, K; Royer, H D; Witt, C; Wolff, G

    2004-01-01

    The human Y-box binding protein, YB-1, is a multifunctional protein that regulates gene expression. Nuclear expression of YB-1 has been associated with chemoresistance and poor prognosis of tumour patients. Representative samples from autopsied material of primary tumours from 77 patients with NSCLC were investigated by immunohistochemistry for subcellular distribution of YB-1 and p53, in order to evaluate the prognostic role of nuclear expression of YB-1. Cytoplasmic YB-1 expression was found in all tumour samples, whereas nuclear expression was only observed in 48%. There was no correlation with histological classification, clinical parameters or tumour size, stage and metastasis status. However, patients with positive nuclear YB-1 expression in tumours showed reduced survival times when compared with patients without nuclear expression. Including information about the histology and mutational status for p53 increased the prognostic value of nuclear YB-1. Patients with nuclear YB-1 expression and p53 mutations had the worst prognosis (median survival 3 months), while best outcome was found in patients with no nuclear YB-1 and wildtype p53 (median survival 15 months). This suggests that the combined analysis of both markers allows a better identification of subgroups with varying prognosis. Nuclear expression of Y-box binding protien seems to be an independent prognostic marker.

  3. Anatomical markers of activity in neuroendocrine systems: are we all 'fos-ed out'?

    PubMed

    Hoffman, G E; Lyo, D

    2002-04-01

    It has now been nearly 15 years since the immediate early gene, c-fos, and its protein product, Fos, were introduced as tools for determining activity changes within neurones of the nervous system. In the ensuing years, this approach was applied to neuroendocrine study with success. With it have come advances in our understanding of which neuroendocrine neurones respond to various stimuli and how other central nervous system components interact with neuroendocrine neurones. Use of combined tract-tracing approaches, as well as double-labelling for Fos and transmitter markers, have added to characterization of neuroendocrine circuits. The delineation of the signal transduction cascades that induce Fos expression has led to establishment of the relationship between neurone firing and Fos expression. Importantly, we can now appreciate that Fos expression is often, but not always, associated with increased neuronal firing and vice versa. There are remaining gaps in our understanding of Fos in the nervous system. To date, knowledge of what Fos does after it is expressed is still limited. The transience of Fos expression after stimulation (especially if the stimulus is persistent) complicates design of experiments to assess the function of Fos and makes Fos of little value as a marker for long-term changes in neurone activity. In this regard, alternative approaches must be sought. Useful alternative approaches employed to date to monitor neuronal changes in activity include examination of (i) signal transduction intermediates (e.g. phosphorylated CREB); (ii) transcriptional/translational intermediates (e.g. heteronuclear RNA, messenger RNA (mRNA), prohormones); and (iii) receptor translocation. Another capitalizes on the fact that many neuroendocrine systems show striking stimulus-transcription coupling in the regulation of their transmitter or its synthetic enzymes. Together, as we move into the 21st Century, the use of multiple approach to study activity within

  4. Ets-1 mRNA expression in effusions of serous ovarian carcinoma patients is a marker of poor outcome.

    PubMed

    Davidson, B; Risberg, B; Goldberg, I; Nesland, J M; Berner, A; Tropé, C G; Kristensen, G B; Bryne, M; Reich, R

    2001-12-01

    Ets-1 proto-oncogene is a transcription factor with a role in the activation of metastasis-associated molecules. We recently found that Ets-1 mRNA expression in solid tumors is a marker of poor prognosis in ovarian carcinoma. The objective of this study was to compare the expression of Ets-1 mRNA in effusions and primary and metastatic tumors of serous ovarian carcinoma patients and to evaluate its prognostic role in effusions. Sections from 67 malignant effusions and 90 primary and metastatic lesions were evaluated for expression of Ets-1 using mRNA in situ hybridization. Expression of Ets-1 mRNA was detected in carcinoma cells in 24 of 67 (36%) effusions. Expression in cancer cells was similar in peritoneal and pleural effusions. In solid lesions Ets-1 expression was detected in both tumor cells and stromal cells in 34 of 90 (38%) lesions. Ets-1 expression in tumor cells showed a strong association with that of stromal cells (p <0.001). Ets-1 expression in effusions showed an association with mRNA expression of basic fibroblast growth factor, previously studied in this patient cohort (p = 0.019). Ets-1 expression in solid lesions showed an association with mRNA expression of vascular endothelial growth factor (p <0.001 for both carcinoma and stromal cells), basic fibroblast growth factor (p = 0.007 for carcinoma cells, p = 0.006 for stromal cells), and interleukin-8 (IL-8) (p = 0.001 for tumor cells). Ets-1 mRNA showed upregulation in metastases when compared with effusion specimens (p = 0.028). In univariate survival analysis Ets-1 expression in carcinoma cells in effusions correlated with poor survival (p = 0.003). Our findings confirm the role of Ets-1 as a novel prognostic marker in advanced-stage ovarian carcinoma and extend it to effusion specimens. The elevated expression in solid metastases supports a central role in tumor progression as well. The association between Ets-1 mRNA expression and the expression of angiogenic genes, documented also in our

  5. The multidrug resistance protein 1: a functionally important activation marker for murine Th1 cells.

    PubMed

    Prechtl, S; Roellinghoff, M; Scheper, R; Cole, S P; Deeley, R G; Lohoff, M

    2000-01-15

    Previously, we described the expression of an energy-dependent pump in resting murine Th2 (but not resting Th1) cells which extruded the fluorescent dye Fluo-3. After stimulation with Ag and APCs, Th1 cells also expressed this pump. Furthermore, expression of the murine multidrug resistance protein 1 (mrp1) correlated with the presence of the pump. In this study, we report that Fluo-3 is indeed transported by murine mrp1 or its human ortholog MRP1, as revealed by transfection of HEK 293 cells with mrp1 or MRP1 cDNA. Like antigenic activation, IL-2 dose-dependently enhanced the Fluo-3-extruding activity in murine Th1 cells. Although TNF-alpha and IL-12 by themselves only weakly enhanced Fluo-3 extrusion, each of them did so in strong synergism with IL-2. An Ab directed against mrp1 was used to quantify the expression of mrp1 protein in T cells at the single-cell level. Like the Fluo-3 pump, mrp1 protein expression was enhanced by IL-2. Immunohistochemical studies using confocal laser microscopy indicated that mrp1 is localized mainly at the plasma membrane. In addition, protein expression of mrp1 was induced in Vbeta8+CD4+ T cells 12 h after in vivo application of Staphylococcal enterotoxin B. Finally, mrp1 was functionally relevant during the activation process of Th1 cells, because T cell activation could be suppressed by exposure of cells to the mrp1 inhibitor MK571. Thus, we present mrp1 as a novel, functionally important activation marker for Th1 cells and short-term in vivo activated CD4+ T cells, whereas its expression seems to be constitutive in Th2 cells.

  6. Differentially expressed genes in human peripheral blood as potential markers for statin response.

    PubMed

    Won, Hong-Hee; Kim, Suk Ran; Bang, Oh Young; Lee, Sang-Chol; Huh, Wooseong; Ko, Jae-Wook; Kim, Hyung-Gun; McLeod, Howard L; O'Connell, Thomas M; Kim, Jong-Won; Lee, Soo-Youn

    2012-02-01

    There is a considerable inter-individual variation in response to statin therapy and one third of patients do not meet their treatment goals. We aimed to identify differentially expressed genes that might be involved in the effects of statin treatment and to suggest potential markers to guide statin therapy. Forty-six healthy Korean subjects received atorvastatin; their whole-genome expression profiles in peripheral blood were analyzed before and after atorvastatin administration in relation with changes in lipid profiles. The expression patterns of the differentially expressed genes were also compared with the data of familial hypercholesterolemia (FH) patients and controls. Pairwise comparison analyses revealed differentially expressed genes involved in diverse biological processes and molecular functions related with immune responses. Atorvastain mainly affected antigen binding, immune or inflammatory response including interleukin pathways. Similar expression patterns of the genes were observed in patients with FH and controls. The Charcol-Leyden crystal (CLC), CCR2, CX3CR1, LRRN3, FOS, LDLR, HLA-DRB1, ERMN, and TCN1 genes were significantly associated with cholesterol levels or statin response. Interestingly, the CLC gene, which was significantly altered by atorvastatin administration and differentially expressed between FH patients and controls, showed much bigger change in high-responsive group than in low-responsive group. We identified differentially expressed genes that might be involved in mechanisms underlying the known pleiotropic effects of atorvastatin, baseline cholesterol levels, and drug response. Our findings suggest CLC as a new candidate marker for statin response, and further validation is needed.

  7. Expression of stem cell markers nanog and PSCA in gastric cancer and its significance

    PubMed Central

    ZHAO, XUANZHONG; WANG, FENG; HOU, MINGXING

    2016-01-01

    The present study aimed to determine the expression of stem cell markers Nanog compared with PSCA in gastric cancer tissues and adjacent normal tissues, and to investigate the association between tumor stem cells and initiation, progression, metastasis, and prognosis of gastric cancer. One hundred chemotherapy- and radiotherapy-naive patients with pathologically confirmed gastric cancer were enrolled from the General Surgery Department and Surgical Oncology Department of the Affiliated Hospital of Inner Mongolia Medical University (Hohhot, P.R. China), between October 2011 and June 2013. Surgically resected specimens of cancer tissues and adjacent normal tissues (>5 cm from the boundary of cancerous component) were collected. The mRNA expression levels of Nanog and PSCA in those tissues was determined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The correlation between the expression of stem cell markers Nanog and PSCA in gastric cancer tissues and clinicopathological factors was analyzed. The qPCR results demonstrated that the relative expression of Nanog was increased in gastric cancer tissues compared with in the adjacent tissues (P<0.05); and relative expression of PSCA was reduced in gastric cancer tissues compared with adjacent tissues (P<0.05). The expression of Nanog and PSCA in gastric cancer tissues was associated with tumor differentiation. The expression of Nanog was increased in poorly-differentiated and undifferentiated tumors compared with moderately- and well-differentiated tumors (P<0.05). The expression of PSCA was reduced in poorly differentiated and undifferentiated tumors compared with moderately- and well-differentiated tumors (P<0.05). However, the expression of Nanog and PSCA was not associated with age, gender, tumor size, TNM stage, depth of invasion, or lymph node metastasis. Therefore, Nanog and PSCA may have potential as molecular markers to reflect the differentiation status of gastric cancer. PMID

  8. Is Interleukin 10 (IL10) Expression in Breast Cancer a Marker of Poor Prognosis?

    PubMed

    Bhattacharjee, Hemanga Kumar; Bansal, Virinder Kumar; Nepal, Bikash; Srivastava, Sandeep; Dinda, Amit K; Misra, Mahesh C

    2016-09-01

    Interleukin 10 (IL10) is a poor prognostic marker in several cancers. Its role in breast cancer is not well elucidated. The present study is designed to see the expression of IL10 in breast cancer tissue and to evaluate its correlation with the established markers of prognosis. Sixty female patients who underwent surgery for breast cancer were enrolled for the study. Immediately after surgery, 2-5 g of tumour tissue and similar volume of peritumoural normal breast tissue were collected for IL10 assay. IL10 expression was assayed by immunohistochemistry. IL10 expressing tumours and IL10 non expressing tumours were compared. Chi square/Fisher exact test and student's t test were used to compare the data. p- valueless than 0.05 was considered as statistically significant. Thirty six patients (60 %) of carcinoma breast showed IL 10 expression in tumour tissue as compared to no IL 10 expression in any peritumouralnormal breast tissue (p < 0.01). IL10 expression had statistically significant correlation with locally advanced disease, tumour grade, HER2 + ve tumours and ER-ve, PR-ve, HER2 + ve breast cancer subtypes (p = 0.001, 0.001, 0.001 and 0.01 respectively). No correlation could be found with patient's age, tumour size, tumour histology and ER and PR status. Correlation of IL10 expressing tumours with several established poor prognostic markers of breast cancer may indicate the possible association of IL10 expression with poor prognosis. Large studies with long term follow up are needed to substantiate the association of IL10 with poor prognosis. PMID:27651693

  9. Resveratrol prevents the expression of glaucoma markers induced by chronic oxidative stress in trabecular meshwork cells

    PubMed Central

    Luna, Coralia; Li, Guorong; Liton, Paloma B; Qiu, Jianming; Epstein, David L.; Challa, Pratap; Gonzalez, Pedro

    2009-01-01

    Elevated intraocular pressure (IOP) constitutes the best characterized risk for primary open-angle glaucoma (POAG). Elevated IOP is believed to result from an increase in aqueous humor outflow resistance at the level of the trabecular meshwork (TM)/ Schlemm's canal (SC). Malfunction of the TM in POAG is associated with the expression of markers for inflammation, cellular senescence, oxidative damage, and decreased cellularity. Current POAG treatments rely on lowering IOP, but there is no therapeutic approach available to delay the loss of function of the TM in POAG patients. We evaluated the effects of chronic administration of the dietary supplement resveratrol on the expression of markers for inflammation, oxidative damage, and cellular senescence in primary TM cells subjected to chronic oxidative stress (40% O2). Resveratrol treatment effectively prevented increased production of intracellular reactive oxygen species (iROS) and inflammatory markers (IL1α, IL6, IL8, and ELAM-1), and reduced expression of the senescence markers sa-β-gal, lipofuscin, and accumulation of carbonylated proteins. Furthermore, resveratrol exerted antiapoptotic effects that were not associated with a decrease in cell proliferation. These results suggest that resveratrol could potentially have a role in preventing the TM tissue abnormalities observed in POAG. PMID:19027816

  10. Prolonged expression of senescence markers in mice exposed to gamma-irradiation

    PubMed Central

    Seol, Min-A; Eom, Hyeon Soo; Kim, Seol-Hwa; Park, Hae-Ran; Jo, Sung-Kee

    2012-01-01

    Although ionizing radiation is known to induce cellular senescence in vitro and in vivo, its long-term in vivo effects are not well defined. In this study, we examined the prolonged expression of senescence markers in mice irradiated with single or fractionated doses. C57BL/6 female mice were exposed to 5 Gy of γ-rays in single or 5, 10, 25 fractions. At 2, 4, and 6 months after irradiation, senescence markers including mitochondrial DNA (mtDNA) common deletion, p21, and senescence-associated β-galactosidase (SA β-gal) were monitored in the lung, liver, and kidney. Increases of mtDNA deletion were detected in the lung, liver, and kidney of irradiated groups. p21 expression and SA β-gal staining were also increased in the irradiated groups compared to the non-irradiated control group. Increases of senescence markers persisted up to 6 months after irradiation. Additionally, the extent of mtDNA deletion and the numbers of SA β-gal positive cells were greater as the number of radiation fractions increased. In conclusion, our results showed that ionizing radiation, especially that delivered in fractions, can cause the persistent upregulation of senescence marker expression in vivo. This should be considered when dealing with chronic normal tissue injuries caused by radiation therapy or radiation accidents. PMID:23271173

  11. Cell cycle markers have different expression and localization patterns in neuron-like PC12 cells and primary hippocampal neurons.

    PubMed

    Negis, Yesim; Unal, Aysegul Yildiz; Korulu, Sirin; Karabay, Arzu

    2011-06-01

    Neuron-like PC12 cells are extensively used in place of neurons in published studies. Aim of this paper has been to compare mRNA and protein expressions of cell cycle markers; cyclinA, B, D, E; Cdk1, 2 and 4; and p27 in post-mitotic primary hippocampal neurons, mitotically active PC12 cells and NGF-differentiated post-mitotic PC12 cells. Contrary to PC12 cells, in neurons, the presence of all these markers was detected only at mRNA level; except for cyclinA, cyclinE and Cdk4, which were detectable also at protein levels. In both NGF-treated PC12 cells and neurons, cyclinE was localized only in the nucleus. In NGF-treated PC12 cells cyclinD and Cdk4 were localized in the nucleus while, in neurons cyclinD expression was not detectable; Cdk4 was localized in the cytoplasm. In neurons, cyclinA was nuclear, whereas in NGF-treated PC12 cells, it was localized in the cell body and along the processes. These results suggest that PC12 cells and primary neurons are different in terms of cell cycle protein expressions and localizations. Thus, it may not be very appropriate to use these cells as neuronal model system in order to understand neuronal physiological activities, upstream of where may lie cell cycle activation triggered events.

  12. Developmental Markers Expressed in Neocortical Layers Are Differentially Exhibited in Olfactory Cortex

    PubMed Central

    Brunjes, Peter C.; Osterberg, Stephen K.

    2015-01-01

    Neurons in the cerebral cortex stratify on the basis of their time of origin, axonal terminations and the molecular identities assigned during early development. Olfactory cortices share many feature with the neocortex, including clear lamination and similar cell types. The present study demonstrates that the markers differentially expressed in the projection neurons of the cerebral cortex are also found in olfactory areas. Three of the four regions examined (pars principalis of the anterior olfactory nucleus: AONpP, anterior and posterior piriform cortices: APC, PPC, and the olfactory tubercle) expressed transcription factors found in deep or superficial neurons in the developing neocortex, though large differences were found between areas. For example, while the AONpP, APC and PPC all broadly expressed the deep cortical marker CTIP2, NOR1 (NR4a3) levels were higher in AONpP and DAARP-32 was more prevalent in the APC and PPC. Similar findings were encountered for superficial cortical markers: all three regions broadly expressed CUX1, but CART was only observed in the APC and PPC. Furthermore, regional variations were observed even within single structures (e.g., NOR1 was found primarily in in the dorsal region of AONpP and CART expression was observed in a discrete band in the middle of layer 2 of both the APC and PPC). Experiments using the mitotic marker EDU verified that the olfactory cortices and neocortex share similar patterns of neuronal production: olfactory cells that express markers found in the deep neocortex are produced earlier than those that express superficial makers. Projection neurons were filled by retrograde tracers injected into the olfactory bulb to see if olfactory neurons with deep and superficial markers had different axonal targets. Unlike the cerebral cortex, no specificity was observed: neurons with each of the transcription factors examined were found to be labelled. Together the results indicate that olfactory cortices are complex

  13. Serological markers of hepatitis B and C in patients with HIV/AIDS and active tuberculosis.

    PubMed

    Araújo-Mariz, Carolline; Lopes, Edmundo Pessoa; Ximenes, Ricardo A A; Lacerda, Heloísa R; Miranda-Filho, Demócrito B; Montarroyos, Ulisses R; Barreto, Silvana; Salustiano, Daniela Medeiros; Albuquerque, Maria Fátima Pessoa Militão

    2016-06-01

    Infection with hepatitis B virus (HBV) and C virus (HCV) are common in patients with HIV/AIDS and tuberculosis (TB). This is a cross-sectional study with patients infected with HIV/AIDS and active TB in Recife, Brazil, aiming to verify the prevalence of markers for HBV: antibody to hepatitis B core antigen (anti-HBc); and HCV: antibody to hepatitis C virus (anti-HCV) by chemiluminescence, and to identify the frequency of associated factors. Data were collected through questionnaires, and blood was drawn from patients for analysis. We used the chi-square test and the Fisher exact test when necessary. We conducted a bivariate logistic regression analysis and the magnitude of the associations was expressed as odds ratio (OR) with a confidence interval of 95%. Among 166 patients studied with HIV/AIDS and active TB, anti-HBc was positive in 61 patients [36.7%; 95%CI (29.4-44.6%)] and anti-HCV in 11[6.6%; 95%CI (3.4-11.5%)]. In the logistic regression analysis, male sex, and age ≥40 years were independent factors associated with the occurrence of anti-HBc. In conclusion, we verified a high frequency of HBV contact marker and a low frequency of HCV markers in patients with HIV/AIDS and TB in Recife.

  14. Expression of epithelial-mesenchymal transition markers at the invasive front of oral squamous cell carcinoma

    PubMed Central

    COSTA, Liana Cristina Melo Carneiro; LEITE, Camila Ferreira; CARDOSO, Sérgio Vitorino; LOYOLA, Adriano Mota; de FARIA, Paulo Rogério; SOUZA, Paulo Eduardo Alencar; HORTA, Martinho Campolina Rebello

    2015-01-01

    Oral squamous cell carcinoma (OSCC) is one of the most common malignances. In epithelial-mesenchymal transition (EMT), epithelial cells switch to mesenchymal-like cells exhibiting high mobility. This migratory phenotype is significant during tumor invasion and metastasis. Objective : The aim of this study is to evaluate the expression of the EMT markers E-cadherin, N-cadherin and vimentin in OSCC. Material and Methods : Immunohistochemical detection of E-cadherin, N-cadherin and vimentin was performed on 20 OSCC samples. Differences in the expression of each protein at the invasive front (IF) and in the central/superficial areas (CSA) of the tumor were assessed. Differences in the expression of each protein at the IF of both histologically high- and low-invasive OSCCs were evaluated. Associations among expression of proteins at the IF were assessed. Correlations between the expression levels of each protein at the IF and the tumor stage and clinical nodal status were also evaluated. Results : Reduced expression of E-cadherin was detected in 15 samples (75%). E-cadherin expression was reduced at the IF when compared to the CSA and in high-invasive tumors when compared to low-invasive tumors. All samples were negative for N-cadherin, even though one sample showed an inconspicuous expression. Positive expression of vimentin was observed in 6 samples (30%). Nevertheless, there was no difference in vimentin expression between the IF and the CSA regions or between the low- and high-invasive tumors. Furthermore, no association was observed among protein expression levels at the IF. Finally, no correlations were observed between each protein’s expression levels and tumor stage or clinical nodal status. Conclusions : Reduced E-cadherin expression at the IF and its association with histological invasiveness suggest that this protein is a noteworthy EMT marker in OSCC. Although vimentin was also detected as an EMT marker, its expression was neither limited to the IF nor was

  15. [Cordyceps sinensis enhances lymphocyte proliferation and CD markers expression in simulated microgravity environment].

    PubMed

    Hao, Tong; Li, Jun-Jie; Du, Zhi-Yan; Duan, Cui-Mi; Wang, Yan-Meng; Wang, Chang-Yong; Song, Jing-Ping; Wang, Lin-Jie; Li, Ying-Hui; Wang, Yan

    2012-10-01

    This study was aimed to explore the effect of cordyceps sinensis enhancing lymphocyte proliferation and surface CD marker expression in simulated microgravity environment. The splenic lymphocytes were separated from mice and cultured in the rotary cell culture system simulated microgravity environment. The cells were treated with different concentration of cordyceps sinensis solution (0, 6.25, 12.5, 25 and 50 µg/ml) for 24, 48 and 72 h respectively, then the cells were harvested, and analyzed for cell proliferation and the expression of cell surface markers (CD4 and CD8). The results showed that under simulated microgravity environment, the lymphocyte proliferation was inhibited. When the concentration of cordyceps sinensis was 25 or 50 µg/ml, the lymphocyte proliferation, CD4 and CD8 expressions all increased, but 50 µg/ml cordyceps sinensis could inhibit the proliferation ability with the time prolonging. It is concluded that the suitable concentration of cordyceps sinensis displayed the ability to enhance the lymphocyte proliferation and CD marker expression in simulated microgravity environment. These results may be valuable for screening drugs which can be potentially against immunosuppression under simulated microgravity. PMID:23114150

  16. Molecular beacon imaging of tumor marker gene expression in pancreatic cancer cells.

    PubMed

    Yang, Lily; Cao, Zehong; Lin, Yiming; Wood, William C; Staley, Charles A

    2005-05-01

    We have developed a fluorescence imaging-based approach to detect expression of tumor marker genes in pancreatic cancer cells using molecular beacons (MBs). MBs are short hairpin oligonucleotide probes that bind to specific oligonucleotide sequences and produce fluorescent signals. MBs targeting transcripts of two tumor marker genes, mutant K-ras and survivin, were synthesized and their specificity in detection of the expression of those genes in pancreatic cancer cells was examined. We found that K-ras MBs differentially bind to mutant K-ras mRNAs, resulting in strong fluorescent signals in pancreatic cancer cells with specific mutant K-ras genes but not in normal cells or cancer cells expressing either wild type or a different mutation of the K-ras gene. Additionally, MBs targeting survivin mRNA produced a bright fluorescent signal specifically in pancreatic cancer cells. We also demonstrated that MBs labeled with different fluorophores could detect survivin and mutant K-ras mRNAs simultaneously in single cancer cells. Furthermore, we showed that survivin and K-ras MBs have a high specificity in identifying cancer cells on frozen sections of pancreatic cancer tissues. In conclusion, molecular beacon-based imaging of expression of tumor marker genes has potential for the development of novel approaches for the detection of pancreatic cancer cells.

  17. Corosolic acid suppresses the expression of inflammatory marker genes in CCL4-induced-hepatotoxic rats.

    PubMed

    Balakrishnan, Aristatile; Al-Assaf, Abdullah Hassan

    2016-07-01

    The aim of the study was to asses the anti-inflammatory effects of corosolic acid on the carbon tetrachloride (CCL4) toxicity in rats. Liver toxicity was induced by administered CCL4 (single dose (1:1 in liquid paraffin) orally at 1.25 ml/kg. Rats were pretreated with CRA for 7 days before made CCL(4) toxicity at 20 mg/kg BW. The mRNA levels of TNF-α, IL-6, iNOS, COX-2 and NF-kB were assayed by reverse transcriptase PCR analysis. The mRNA levels of proinflammatory cytokines such as TNF-α, IL-6, and the inflammatory markers such as iNOS, COX-2 and NF-kB were significantly up regulated in CCl(4) induced rats and treatment with corosolic acid significantly reduced the expression of the above indicators. Our results suggest that the inhibition of TNF-α, IL-6, iNOS, COX-2 and NF-κB by corosolic acid, a potential candidate could possess anti-inflammatory activity besides its hepatoprotective effect in CCl4 liver toxicity in rats. PMID:27393448

  18. Predictive markers of capecitabine sensitivity identified from the expression profile of pyrimidine nucleoside-metabolizing enzymes.

    PubMed

    Yasuno, Hideyuki; Kurasawa, Mitsue; Yanagisawa, Mieko; Sato, Yasuko; Harada, Naoki; Mori, Kazushige

    2013-02-01

    Molecular markers predicting sensitivity to anticancer drugs are important and useful not only for selecting potential responders but also for developing new combinations. In the present study, we analyzed the difference in the sensitivity of xenograft models to capecitabine (Xeloda®), 5'-deoxy-5-fluorouridine (5'-DFUR, doxifluridine, Furtulon®) and 5-FU by comparing the mRNA levels of 12 pyrimidine nucleoside-metabolizing enzymes. Amounts of mRNA in the tumor tissues of 80 xenograft models were determined by real-time RT-PCR and mutual correlations were examined. A clustering analysis revealed that the 12 enzymes were divided into two groups; one group consisted of 8 enzymes, including orotate phosphoribosyl transferase (OPRT), TMP kinase (TMPK) and UMP kinase (UMPK), and was related to the de novo synthesis pathway for nucleotides, with mRNA expression levels showing significant mutual correlation. In the other group, 4 enzymes, including thymidine phosphorylase (TP) and dihydropyrimidine dehydrogenase (DPD), were involved in the salvage/degradation pathway of the nucleotides, and the mRNA levels of this group were dispersed more widely than that of the de novo group. Antitumor activity was assessed in 24 xenograft models for each drug. The antitumor activity of capecitabine and 5'-DFUR correlated significantly with the mRNA levels of TP and with the TP/DPD ratio, whereas the activity of 5-FU correlated significantly with OPRT, TMPK, UMPK and CD. In a stepwise regression analysis, TP and DPD were found to be independent predictive factors of sensitivity to capecitabine and 5'-DFUR, and UMPK was predictive of sensitivity to 5-FU. These results indicate that the predictive factors for sensitivity to capecitabine and 5'-DFUR in xenograft models may be different from those for 5-FU, suggesting that these drugs may have different responders in clinical usage. PMID:23229803

  19. Tumor Vascular Permeability to a Nanoprobe Correlates to Tumor-Specific Expression Levels of Angiogenic Markers

    PubMed Central

    Karathanasis, Efstathios; Chan, Leslie; Karumbaiah, Lohitash; McNeeley, Kathleen; D'Orsi, Carl J.; Annapragada, Ananth V.; Sechopoulos, Ioannis; Bellamkonda, Ravi V.

    2009-01-01

    Background Vascular endothelial growth factor (VEGF) receptor-2 is the major mediator of the mitogenic, angiogenic, and vascular hyperpermeability effects of VEGF on breast tumors. Overexpression of VEGF and VEGF receptor-2 is associated with the degree of pathomorphosis of the tumor tissue and unfavorable prognosis. In this study, we demonstrate that non-invasive quantification of the degree of tumor vascular permeability to a nanoprobe correlates with the VEGF and its receptor levels and tumor growth. Methodology/Principal Findings We designed an imaging nanoprobe and a methodology to detect the intratumoral deposition of a 100 nm-scale nanoprobe using mammography allowing measurement of the tumor vascular permeability in a rat MAT B III breast tumor model. The tumor vascular permeability varied widely among the animals. Notably, the VEGF and VEGF receptor-2 gene expression of the tumors as measured by qRT-PCR displayed a strong correlation to the imaging-based measurements of vascular permeability to the 100 nm-scale nanoprobe. This is in good agreement with the fact that tumors with high angiogenic activity are expected to have more permeable blood vessels resulting in high intratumoral deposition of a nanoscale agent. In addition, we show that higher intratumoral deposition of the nanoprobe as imaged with mammography correlated to a faster tumor growth rate. This data suggest that vascular permeability scales to the tumor growth and that tumor vascular permeability can be a measure of underlying VEGF and VEGF receptor-2 expression in individual tumors. Conclusions/Significance This is the first demonstration, to our knowledge, that quantitative imaging of tumor vascular permeability to a nanoprobe represents a form of a surrogate, functional biomarker of underlying molecular markers of angiogenesis. PMID:19513111

  20. Polyester copolymer scaffolds enhance expression of bone markers in osteoblast-like cells.

    PubMed

    Idris, Shaza Bushra; Arvidson, Kristina; Plikk, Peter; Ibrahim, Salah; Finne-Wistrand, Anna; Albertsson, Ann-Christine; Bolstad, Anne Isine; Mustafa, Kamal

    2010-08-01

    In tissue engineering, the resorbable aliphatic polyester poly(L-lactide) (PLLA) is used as scaffolds in bone regeneration. Copolymers of poly(L-lactide)-co-(epsilon-caprolactone) [poly(LLA-co-CL)] and poly(L-lactide)-co-(1,5-dioxepan-2-one) [poly(LLA-co-DXO)], with superior mechanical properties to PLLA, have been developed to be used as scaffolds, but the influence on the osteogenic potential is unclear. This in vitro study of test scaffolds of poly(LLA-co-CL) and poly(LLA-co-DXO) using PLLA scaffolds as a control demonstrates the attachment and proliferation of human osteoblast-like cells (HOB) as measured by SEM and a methylthiazol tetrazolium (MTT) colorimetric assay, and the progression of HOB osteogenesis for up to 3 weeks; expressed as synthesis of the osteoblast differentiation markers: collagen type 1 (Col 1), alkaline phosphatase, bone sialoprotein, osteocalcin (OC), osteopontin and runt related gene 2 (Runx2). Surface analysis disclosed excellent surface attachment, spread and penetration of the cells into the pores of the test scaffolds compared to the PLLA. MTT results indicated that test scaffolds enhanced the proliferation of HOBs. Cells grown on the test scaffolds demonstrated higher synthesis of Col 1 and OC and also increased bone markers mRNA expression. Compared to scaffolds of PLLA, the poly(LLA-co-CL) and poly(LLA-co-DXO) scaffolds enhanced attachment, proliferation, and expression of osteogenic markers by HOBs in vitro. Therefore, these scaffolds might be appropriate carriers for bone engineering.

  1. Differential expression of Yes-associated protein is correlated with expression of cell cycle markers and pathologic TNM staging in non-small-cell lung carcinoma.

    PubMed

    Kim, Jin Man; Kang, Dong Wook; Long, Liang Zhe; Huang, Song-Mei; Yeo, Min-Kyung; Yi, Eunhee S; Kim, Kyung-Hee

    2011-03-01

    Yes-associated protein, a downstream effector of the Hippo signaling pathway, has been linked to progression of non-small-cell lung carcinoma. The aim of this study was to investigate expression of Yes-associated protein in lung adenocarcinoma and squamous cell carcinoma. Associations of Yes-associated protein expression with clinicopathologic parameters, expression of cell cycle-specific markers, and epidermal growth factor receptor gene amplification were also analyzed. In a univariate analysis of the 66 adenocarcinomas, high nuclear expression of Yes-associated protein was significantly correlated with expression of cyclin A and mitogen-activated protein kinase. Multivariate analysis, including age and sex, showed that cyclin A expression was independently correlated with nuclear expression of Yes-associated protein in adenocarcinomas. Furthermore, high nuclear expression of Yes-associated protein was also a significant predictor of epidermal growth factor receptor gene amplification for adenocarcinoma. For the 102 squamous cell carcinomas, univariate analysis revealed that high cytoplasmic expression of Yes-associated protein was correlated with the low pathologic TNM staging (stage I) and histologic grading. Multivariate analysis, including age and sex, showed that cytoplasmic expression of Yes-associated protein was an independent predictor of low pathologic TNM staging. These results indicate that nuclear overexpression of Yes-associated protein contributes to pulmonary adenocarcinoma growth and that high cytoplasmic expression of Yes-associated protein is an independent predictor of low pathologic TNM staging and histologic grading. The differential effects of Yes-associated protein expression patterns in adenocarcinomas and squamous cell carcinomas suggest that Yes-associated protein may play important roles in different pathways in distinct tumor subtypes. These observations may, therefore, lead to new perspectives on therapeutic targeting of these tumor

  2. Differential expression of monocyte surface markers among TB patients with diabetes co-morbidity.

    PubMed

    Stew, Samuel S; Martinez, Perla J; Schlesinger, Larry S; Restrepo, Blanca I

    2013-12-01

    The expression of monocyte surface markers was compared between tuberculosis patients with and without type 2 diabetes (DM2). DM2 was associated with increased CCR2 expression, which may restrain monocyte traffic to the lung. Other host factors associated with baseline monocyte changes were older age (associated with lower CD11b) and obesity (associated with higher RAGE). Given that DM2 patients are more likely to be older and obese, their monocytes are predicted to be altered in function in ways that affect their interaction with Mycobacterium tuberculosis.

  3. The influence of statin therapy on platelet activity markers in hyperlipidemic patients after ischemic stroke

    PubMed Central

    Chmielewski, Henryk; Kaczorowska, Beata; Przybyła, Monika; Baj, Zbigniew

    2015-01-01

    Introduction Low-density lipoprotein cholesterol (LDL-C) has been reported to increase platelet activation. Reducing the level of LDL-C with statins induces important pleiotropic effects such as platelet inhibition. This association between platelet activity and statin therapy may be clinically important in reducing the risk of ischemic stroke. We investigated the effect of simvastatin therapy on platelet activation markers (platelet CD62P, sP-selectin, and platelet-derived microparticles (PDMPs)) in hyperlipidemic patients after ischemic stroke. Material and methods The study group consisted of 21 hyperlipidemic patients after ischemic stroke confirmed by CT, and 20 healthy subjects served as controls. We assessed the CD62P expression on resting and thrombin-activated blood platelets. CD62P and PDMPs were analyzed by the use of monoclonal antibodies anti-CD61 and anti-CD62 on a flow cytometer. The level of sP-selectin in serum was measured by the ELISA (enzyme-linked immunosorbent assay) method. All markers were re-analyzed after 6 months of treatment with simvastatin (20 mg/day). Results Hyperlipidemic patients presented a significantly higher percentage of CD62+ platelets and higher reactivity to thrombin compared to control subjects. After simvastatin therapy hyperlipidemic patients showed a reduction of the percentage of resting CD62P(+) platelets (p = 0.005) and a reduction of expression and percentage of CD62P(+) platelets after activation by thrombin (median p < 0.05; percentage: p = 0.001). A decrease of sP-selectin levels (p = 0.001) and percentage of PDMPs (p < 0.05) in this group was also observed. Conclusions HMG-CoA reductase inhibitor therapy in stroke patients with hyperlipidemia may be useful not only due to the lipid-lowering effect but also because of a significant role in reduction of platelet activation and reactivity. PMID:25861297

  4. Schlafen-3 decreases cancer stem cell marker expression and autocrine/juxtacrine signaling in FOLFOX-resistant colon cancer cells.

    PubMed

    Oh, Phil-Sun; Patel, Vaishali B; Sanders, Matthew A; Kanwar, Shailender S; Yu, Yingjie; Nautiyal, Jyoti; Patel, Bhaumik B; Majumdar, Adhip P N

    2011-08-01

    We have previously demonstrated that expression of the novel gene schlafen-3 (Slfn-3) correlates with intestinal epithelial cell differentiation (Patel VB, Yu Y, Das JK, Patel BB, Majumdar AP. Biochem Biophys Res Commun 388: 752-756, 2009). The present investigation was undertaken to examine whether Slfn-3 plays a role in regulating differentiation of FOLFOX-resistant (5-fluorouracil + oxaliplatin) colon cancer cells that are highly enriched in cancer stem cells (CSCs). Transfection of Slfn-3 in FOLFOX-resistant colon cancer HCT-116 cells resulted in increase of alkaline phosphatase activity, a marker of intestinal differentiation. Additionally, Slfn-3 transfection resulted in reduction of mRNA and protein levels of the CSC markers CD44, CD133, CD166, and aldehyde dehydrogenase 1 in both FOLFOX-resistant HCT-116 and HT-29 cells. This was accompanied by decreased formation of tumorosphere/colonosphere (an in vitro model of tumor growth) in stem cell medium and inhibition of expression of the chemotherapeutic drug transporter protein ABCG2. Additionally, Slfn-3 transfection of FOLFOX-resistant HCT-116 and HT-29 cells reduced Hoechst 33342 dye exclusion. Finally, Slfn-3 transfection inhibited the expression of transforming growth factor-α in both FOLFOX-resistant colon cancer cells, but stimulated apoptosis in response to additional FOLFOX treatment. In summary, our data demonstrate that Slfn-3 expression inhibits multiple characteristics of CSC-enriched, FOLFOX-resistant colon cancer cells, including induction of differentiation and reduction in tumorosphere/colonosphere formation, drug transporter activity, and autocrine stimulation of proliferation. Thus Slfn-3 expression may render colon CSCs more susceptible to cancer chemotherapeutics.

  5. Expression of T-plastin, FoxP3 and other tumor-associated markers by leukemic T-cells of cutaneous T-cell lymphoma.

    PubMed

    Capriotti, Elisabetta; Vonderheid, Eric C; Thoburn, Christopher J; Wasik, Mariusz A; Bahler, David W; Hess, Allan D

    2008-06-01

    Peripheral blood cells from 28 patients with leukemic cutaneous T-cell lymphoma including 25 patients with Sezary syndrome were evaluated for expression of regulatory T-cell-associated markers (FoxP3, CD25, CTLA-4, neurophilin-1), T-cell activation markers (CD28 and its ligands B7.1 and B7.2) and NK cell-associated markers (NKG2D and its ligands Mic-A and Mic-B) using real-time quantitative polymerase chain reaction. T-plastin served as a positive genetic marker, and its expression correlated to blood tumor burden. More than 90% of samples had transcripts for CD28 and Mic-B, but less than 30% of samples expressed FoxP3, CTLA-4 and CD25. Expression of Mic-B by neoplastic cells could provide another mechanism to inhibit anti-tumor immune responses. FoxP3 expression correlated with a poor prognosis. Although the underlying mechanisms accounting for this correlation remain unclear, the expression of the Foxp3 and CTLA-4 regulatory elements indicates that a subset of leukemic cases displays a regulatory T-cell phenotype.

  6. PDIA3 and PDIA6 gene expression as an aggressiveness marker in primary ductal breast cancer.

    PubMed

    Ramos, F S; Serino, L T R; Carvalho, C M S; Lima, R S; Urban, C A; Cavalli, I J; Ribeiro, E M S F

    2015-01-01

    Changes in the expression of the protein disulfide isomerase genes PDIA3 and PDIA6 may increase endoplasmic reticulum stress, leading to cellular instability and neoplasia. We evaluated the expression of PDIA3 and PDIA6 in invasive ductal carcinomas. Using reverse transcription-quantitative polymerase chain reaction, we compared the mRNA expression level in 45 samples of invasive ductal carcinoma with that in normal breast samples. Increased expression of the PDIA3 gene in carcinomas (P = 0.0009) was observed. In addition, PDIA3 expression was increased in tumors with lymph node metastasis (P = 0.009) and with grade III (P < 0.02). The PDIA6 gene showed higher expression levels in the presence of lymph node metastasis (U = 99.00, P = 0.0476) and lower expression for negative hormone receptors status (P = 0.0351). Our results suggest that alterations in PDIA3/6 expression levels may be involved in the breast carcinogenic process and should be further investigated as a marker of aggressiveness. PMID:26125904

  7. Development of expressed sequence tag and expressed sequence tag–simple sequence repeat marker resources for Musa acuminata

    PubMed Central

    Passos, Marco A. N.; de Oliveira Cruz, Viviane; Emediato, Flavia L.; de Camargo Teixeira, Cristiane; Souza, Manoel T.; Matsumoto, Takashi; Rennó Azevedo, Vânia C.; Ferreira, Claudia F.; Amorim, Edson P.; de Alencar Figueiredo, Lucio Flavio; Martins, Natalia F.; de Jesus Barbosa Cavalcante, Maria; Baurens, Franc-Christophe; da Silva, Orzenil Bonfim; Pappas, Georgios J.; Pignolet, Luc; Abadie, Catherine; Ciampi, Ana Y.; Piffanelli, Pietro; Miller, Robert N. G.

    2012-01-01

    Background and aims Banana (Musa acuminata) is a crop contributing to global food security. Many varieties lack resistance to biotic stresses, due to sterility and narrow genetic background. The objective of this study was to develop an expressed sequence tag (EST) database of transcripts expressed during compatible and incompatible banana–Mycosphaerella fijiensis (Mf) interactions. Black leaf streak disease (BLSD), caused by Mf, is a destructive disease of banana. Microsatellite markers were developed as a resource for crop improvement. Methodology cDNA libraries were constructed from in vitro-infected leaves from BLSD-resistant M. acuminata ssp. burmaniccoides Calcutta 4 (MAC4) and susceptible M. acuminata cv. Cavendish Grande Naine (MACV). Clones were 5′-end Sanger sequenced, ESTs assembled with TGICL and unigenes annotated using BLAST, Blast2GO and InterProScan. Mreps was used to screen for simple sequence repeats (SSRs), with markers evaluated for polymorphism using 20 diploid (AA) M. acuminata accessions contrasting in resistance to Mycosphaerella leaf spot diseases. Principal results A total of 9333 high-quality ESTs were obtained for MAC4 and 3964 for MACV, which assembled into 3995 unigenes. Of these, 2592 displayed homology to genes encoding proteins with known or putative function, and 266 to genes encoding proteins with unknown function. Gene ontology (GO) classification identified 543 GO terms, 2300 unigenes were assigned to EuKaryotic orthologous group categories and 312 mapped to Kyoto Encyclopedia of Genes and Genomes pathways. A total of 624 SSR loci were identified, with trinucleotide repeat motifs the most abundant in MAC4 (54.1 %) and MACV (57.6 %). Polymorphism across M. acuminata accessions was observed with 75 markers. Alleles per polymorphic locus ranged from 2 to 8, totalling 289. The polymorphism information content ranged from 0.08 to 0.81. Conclusions This EST collection offers a resource for studying functional genes, including

  8. Cardiomyocyte marker expression in a human lymphocyte cell line using mouse cardiomyocyte extract.

    PubMed

    Vojdani, Zahra; Tavakolinejad, Sima; Talaei-Khozani, Tahereh; Esmaeilpour, Tahereh; Rasooli, Manuchehr

    2011-03-01

    Cell transplantation shows potential for the treatment of cardiac diseases. Embryonic stem cells, cord blood and mesenchymal stem cells have been suggested as sources for transplantation therapy. Because of some technical limitations with the use of stem cells, transdifferentiation of fully differentiated cells is a potentially useful alternative. We investigated whether human peripheral blood cells could transdifferentiate into cardiomyocyte. Transdifferentiation was induced in a human B lymphocyte cell line (Raji). Cardiomyocyte extract was prepared from adult mouse cardiomyocytes. The cells were treated with 5-aza-2-deoxycytidine and trichostatin A, permeabilized with streptolysin O, and exposed to the mouse cardiomyocyte extract. They were cultured for 10 days, 3 weeks and 4 weeks. Cardiomyocyte markers were detected with immunohistochemistry and flow cytometry. Immunocytochemistry revealed that some cells expressed myosin heavy chain, α-actinin and cardiac troponin T after 3 and 4 weeks. Flow cytometry confirmed these data. In cells exposed to trichostatin A and 5-aza-2-deoxycytidine and permeabilized in the presence of the cardiomyocyte extract, troponin T expression was seen in 3.53% of the cells and 3.11% of them expressed α-actinin. After exposure to the cardiomyocyte extract, some permeabilized cells adhered to the plate loosely; however, the morphology did not change significantly, and they continued to show a rounded shape after 4 weeks. Our treated lymphocytes expressed cardiomyocyte markers. Our results suggest that lymphocytes may be useful in future research as a source of cells for reprogramming procedures.

  9. Transgelin: a potentially useful diagnostic marker differentially expressed in triple-negative and non-triple-negative breast cancers.

    PubMed

    Rao, Deepthi; Kimler, Bruce F; Nothnick, Warren B; Davis, Marilyn K; Fan, Fang; Tawfik, Ossama

    2015-06-01

    Triple negative (TN) (estrogen receptor [ER], progesterone receptor [PR] and HER2-) are highly aggressive, rapidly growing, hormone-unresponsive tumors diagnosed at later stage that affect younger women with shorter overall survival. Most TN tumors are of the basal type. For the remainder, identification of target markers for effective treatment strategies remains a challenge. Transgelin (TGLN) is a 22-kd actin-binding protein of the calponin family. It is one of the earliest markers of smooth muscle differentiation. TGLN has been shown to have important biologic activities including regulating muscle fiber contractility, cell migration, and tumor suppression. We examined TGLN expression in the different molecular subtypes of breast cancer. TGLN expression was examined as a function of tumor size, grade, histologic type, lymph node status, patients' age and overall survival, ER, PR, HER2, and Ki-67 in 101 tumors that included 35 luminal A, 28 luminal B, 4 HER2, and 34 TN types. TGLN positivity (defined as 2+ or 3+) was associated with more aggressive tumors (10% of grade I/II tumors were TGLN+ versus 53% of grade III tumors; P < .001), high Ki-67 count, and low ER and PR expression (P < .001) but not with tumor size, age, or lymph node metastasis. TN (n = 34) tumors were 7.7 times more likely to be TGLN+ than non-TN (n = 67) tumors (77% versus 10%, respectively; P < .001). TGLN may be an excellent diagnostic marker of TN tumors and could be useful in stratification of patients. TGLN may also prove a potential target for future treatment strategies.

  10. Tracking neuronal marker expression inside living differentiating cells using molecular beacons.

    PubMed

    Ilieva, Mirolyuba; Della Vedova, Paolo; Hansen, Ole; Dufva, Martin

    2013-12-19

    Monitoring gene expression is an important tool for elucidating mechanisms of cellular function. In order to monitor gene expression during nerve cell development, molecular beacon (MB) probes targeting markers representing different stages of neuronal differentiation were designed and synthesized as 2'-O-methyl RNA backbone oligonucleotides. MBs were transfected into human mesencephalic cells (LUHMES) using streptolysin-O-based membrane permeabilization. Mathematical modeling, simulations and experiments indicated that MB concentration was equal to the MB in the transfection medium after 10 min transfection. The cells will then each contain about 60,000 MBs. Gene expression was detected at different time points using fluorescence microscopy. Nestin and NeuN mRNA were expressed in approximately 35% of the LUHMES cells grown in growth medium, and in 80-90% of cells after differentiation. MAP2 and tyrosine hydroxylase mRNAs were expressed 2 and 3 days post induction of differentiation, respectively. Oct 4 was not detected with MB in these cells and signal was not increased over time suggesting that MB are generally stable inside the cells. The gene expression changes measured using MBs were confirmed using qRT-PCR. These results suggest that MBs are simple to use sensors inside living cell, and particularly useful for studying dynamic gene expression in heterogeneous cell populations.

  11. Expression of Tgfβ1 and Inflammatory Markers in the 6-hydroxydopamine Mouse Model of Parkinson’s Disease

    PubMed Central

    Haas, Stefan Jean-Pierre; Zhou, Xiaolai; Machado, Venissa; Wree, Andreas; Krieglstein, Kerstin; Spittau, Björn

    2016-01-01

    Parkinson’s disease (PD) is a neurodegenerative disorder that is characterized by loss of midbrain dopaminergic (mDA) neurons in the substantia nigra (SN). Microglia-mediated neuroinflammation has been described as a common hallmark of PD and is believed to further trigger the progression of neurodegenerative events. Injections of 6-hydroxydopamine (6-OHDA) are widely used to induce degeneration of mDA neurons in rodents as an attempt to mimic PD and to study neurodegeneration, neuroinflammation as well as potential therapeutic approaches. In the present study, we addressed microglia and astroglia reactivity in the SN and the caudatoputamen (CPu) after 6-OHDA injections into the medial forebrain bundle (MFB), and further analyzed the temporal and spatial expression patterns of pro-inflammatory and anti-inflammatory markers in this mouse model of PD. We provide evidence that activated microglia as well as neurons in the lesioned SN and CPu express Transforming growth factor β1 (Tgfβ1), which overlaps with the downregulation of pro-inflammatory markers Tnfα, and iNos, and upregulation of anti-inflammatory markers Ym1 and Arg1. Taken together, the data presented in this study suggest an important role for Tgfβ1 as a lesion-associated factor that might be involved in regulating microglia activation states in the 6-OHDA mouse model of PD in order to prevent degeneration of uninjured neurons by microglia-mediated release of neurotoxic factors such as Tnfα and nitric oxide (NO). PMID:26869879

  12. Stem cell marker aldehyde dehydrogenase 1 (ALDH1)-expressing cells are enriched in triple-negative breast cancer.

    PubMed

    Li, Huihui; Ma, Fei; Wang, Haijuan; Lin, Chen; Fan, Ying; Zhang, Xueyan; Qian, Haili; Xu, Binghe

    2013-12-17

    The stem cell marker ALDH1 has been of particular interest to scientists since it has been successfully used as a marker to isolate cancer stem cells from breast cancers. However, little is known, especially in Chinese breast cancer patients, on whether ALDH1 enrichment is prevalent in certain subtypes of breast cancer. In this study, we performed flow cytometry and immunohistochemistry to measure the expression of ALDH1 in 10 breast cancer cell lines and in a set of tissue microarrays consisting of 101 breast cancer tissues from the Chinese population. The 101 breast cancer tissues included 4 cancer subtypes defined on bases of their ER, PR, and HER2 statuses: triple-negative (25 cases), luminal A (33 cases), luminal B (16 cases) and HER2-overexpressing (HER2-OE, 27 cases). We found that ALDH1 was expressed in 25 of the 101 cases of breast cancer tissues. When the analysis was stratified, we found that the expression of ALDH1 varied significantly among the 4 subtypes, with a higher expression in triple-negative breast cancer (TNBC, p=0.003) than in the other 3 subtypes. In a series of breast cancer cell lines, we also confirmed that ALDH1 activity was mainly found in TNBC cell lines compared with non-TNBC ones (15.6% ± 2.45% vs 5.5% ± 2.58%, p=0.026). These data support the concept that the expression of ALDH1 is higher in TNBC than non-TNBC, which may be clinically meaningful for a better understanding of the poor prognosis of TNBC patients.

  13. Dietary methoxychlor exposure modulates splenic natural killer cell activity, antibody-forming cell response and phenotypic marker expression in F0 and F1 generations of Sprague Dawley rats.

    PubMed

    White, K L; Germolec, D R; Booker, C D; Hernendez, D M; McCay, J A; Delclos, K B; Newbold, R R; Weis, C; Guo, T L

    2005-02-14

    Methoxychlor, a chlorinated hydrocarbon pesticide, is a persistent environmental contaminant that has been identified in human reproductive tissues. Methoxychlor has been shown to be estrogenic in both in vivo and in vitro studies. As an endocrine disrupter, it may have the potential to adversely affect endocrine, reproductive, and immune systems in animals. The present study evaluated methoxychlor's immunotoxic potential in F0 (dams) and F1 generations of Sprague Dawley rats exposed to an isoflavone-free diet containing methoxychlor at concentrations of 10, 100, and 1000 ppm. In dams, exposure to methoxychlor from gestation day 7 to postpartum day 51 (65 days total exposure) produced a significant increase in the NK activity (1000 ppm) and the percentages of T cells (1000 ppm), helper T cells (1000 ppm) and macrophages (100 and 1000 ppm). In contrast, a decrease in the numbers of splenocytes and B cells was observed at the 100 and 1000 ppm concentrations. In F1 males, exposure to methoxychlor gestationally, lactationally and through feed from postnatal day 22-64 (78 days total exposure) produced an increase in the spleen IgM antibody-forming cell response to sheep red blood cells (100 and 1000 ppm) and the activity of NK cells (1000 ppm). However, there was a decrease in the terminal body weight (1000 ppm), spleen weight (1000 ppm), thymus weight (100 and 1000 ppm), and the numbers of splenocytes (1000 ppm), B cells (100 and 1000 ppm), cytotoxic T cells (1000 ppm) and NK cells (100 and 1000 ppm). In F1 females, exposure to methoxychlor produced a decrease in the terminal body weight (1000 ppm) and the percentages of cytotoxic T cells (10, 100 and 1000 ppm). These results demonstrate that developmental and adult dietary exposure to methoxychlor modulates immune responses in Sprague Dawley rats. Immunological changes were more pronounced in the F1 generation male rats that were exposed during gestation and postpartum, when compared to the F0 and F1 generation

  14. Neural differentiation potential of human bone marrow-derived mesenchymal stromal cells: misleading marker gene expression

    PubMed Central

    Montzka, Katrin; Lassonczyk, Nina; Tschöke, Beate; Neuss, Sabine; Führmann, Tobias; Franzen, Rachelle; Smeets, Ralf; Brook, Gary A; Wöltje, Michael

    2009-01-01

    Background In contrast to pluripotent embryonic stem cells, adult stem cells have been considered to be multipotent, being somewhat more restricted in their differentiation capacity and only giving rise to cell types related to their tissue of origin. Several studies, however, have reported that bone marrow-derived mesenchymal stromal cells (MSCs) are capable of transdifferentiating to neural cell types, effectively crossing normal lineage restriction boundaries. Such reports have been based on the detection of neural-related proteins by the differentiated MSCs. In order to assess the potential of human adult MSCs to undergo true differentiation to a neural lineage and to determine the degree of homogeneity between donor samples, we have used RT-PCR and immunocytochemistry to investigate the basal expression of a range of neural related mRNAs and proteins in populations of non-differentiated MSCs obtained from 4 donors. Results The expression analysis revealed that several of the commonly used marker genes from other studies like nestin, Enolase2 and microtubule associated protein 1b (MAP1b) are already expressed by undifferentiated human MSCs. Furthermore, mRNA for some of the neural-related transcription factors, e.g. Engrailed-1 and Nurr1 were also strongly expressed. However, several other neural-related mRNAs (e.g. DRD2, enolase2, NFL and MBP) could be identified, but not in all donor samples. Similarly, synaptic vesicle-related mRNA, STX1A could only be detected in 2 of the 4 undifferentiated donor hMSC samples. More significantly, each donor sample revealed a unique expression pattern, demonstrating a significant variation of marker expression. Conclusion The present study highlights the existence of an inter-donor variability of expression of neural-related markers in human MSC samples that has not previously been described. This donor-related heterogeneity might influence the reproducibility of transdifferentiation protocols as well as contributing to the

  15. Expression of IGF-I and Protein Degradation Markers During Hindlimb Unloading and Growth Hormone Administration in Rats

    NASA Astrophysics Data System (ADS)

    Leinsoo, T. A.; Turtikova, O. V.; Shenkman, B. S.

    2013-02-01

    It is known that hindlimb unloading or spaceflight produce atrophy and a number of phenotypic alterations in skeletal muscles. Many of these processes are triggered by the axis growth hormone/insulin-like growth factor I. However growth hormone (GH) and insulin-like growth factor I (IGF-I) expression relationship in rodent models of gravitational unloading is weakly investigated. We supposed the IGF-I is involved in regulation of protein turnover. In this study we examined the IGF-I expression by RT-PCR assay in the rat soleus, tibialis anterior and liver after 3 day of hindlimb suspension with growth hormone administration. Simultaneously were studied expression levels of MuRF-1 and MAFbx/atrogin as a key markers of intracellular proteolysis. We demonstrated that GH administration did not prevent IGF-I expression decreasing under the conditions of simulated weightlessness. It was concluded there are separate mechanisms of action of GH and IGF-I on protein metabolism in skeletal muscles. Gravitational unloading activate proteolysis independently of growth hormone activity.

  16. GFP as a marker for transient gene transfer and expression in Mycoplasma hyorhinis.

    PubMed

    Ishag, Hassan Z A; Liu, Maojun; Yang, Ruosong; Xiong, Qiyan; Feng, Zhixin; Shao, Guoqing

    2016-01-01

    Mycoplasma hyorhinis (M. hyorhinis) is an opportunistic pathogen of pigs and has been shown to transform cell cultures, which has increased the interest of researchers. The green florescence proteins (GFP) gene of Aquorea victoria, proved to be a vital marker to identify transformed cells in mixed populations. Use of GFP to observe gene transfer and expression in M. hyorhinis (strain HUB-1) has not been described. We have constructed a pMD18-O/MHRgfp plasmid containing the p97 gene promoter, origin of replication, tetracycline resistance marker and GFP gene controlled by the p97 gene promoter. The plasmid transformed into M. hyorhinis with a frequency of ~4 × 10(-3) cfu/µg plasmid DNA and could be detected by PCR amplification of the GFP gene from the total DNA of the transformant mycoplasmas. Analysis of a single clone grown on KM2-Agar containing tetracycline, showed a green fluorescence color. Conclusively, this report suggests the usefulness of GFP to monitor transient gene transfer and expression in M. hyorhinis, eventually minimizing screening procedures for gene transfer and expression. PMID:27386255

  17. Expression of Molecular Differentiation Markers Does Not Correlate with Histological Differentiation Grade in Intrahepatic Cholangiocarcinoma

    PubMed Central

    Demarez, Céline; Hubert, Catherine; Sempoux, Christine; Lemaigre, Frédéric P.

    2016-01-01

    The differentiation status of tumor cells, defined by histomorphological criteria, is a prognostic factor for survival of patients affected with intrahepatic cholangiocarcinoma (ICC). To strengthen the value of morphological differentiation criteria, we wished to correlate histopathological differentiation grade with expression of molecular biliary differentiation markers and of microRNAs previously shown to be dysregulated in ICC. We analysed a series of tumors that were histologically classified as well, moderately or poorly differentiated, and investigated the expression of cytokeratin 7, 19 and 903 (CK7, CK19, CK903), SRY-related HMG box transcription factors 4 and 9 (SOX4, SOX9), osteopontin (OPN), Hepatocyte Nuclear Factor-1 beta (HNF1β), Yes-associated protein (YAP), Epithelial cell adhesion molecule (EPCAM), Mucin 1 (MUC1) and N-cadherin (NCAD) by qRT-PCR and immunostaining, and of miR-31, miR-135b, miR-132, miR-200c, miR-221 and miR-222. Unexpectedly, except for subcellular location of SOX9 and OPN, no correlation was found between the expression levels of these molecular markers and histopathological differentiation grade. Therefore, our data point toward necessary caution when investigating the evolution and prognosis of ICC on the basis of cell differentiation criteria. PMID:27280413

  18. Global Gene Expression Profiling Reveals SPINK1 as a Potential Hepatocellular Carcinoma Marker

    PubMed Central

    Kutter, Claudia; Davies, Susan; Alexander, Graeme; Odom, Duncan T.

    2013-01-01

    Background Liver cirrhosis is the most important risk factor for hepatocellular carcinoma (HCC) but the role of liver disease aetiology in cancer development remains under-explored. We investigated global gene expression profiles from HCC arising in different liver diseases to test whether HCC development is driven by expression of common or different genes, which could provide new diagnostic markers or therapeutic targets. Methodology and Principal Findings Global gene expression profiling was performed for 4 normal (control) livers as well as 8 background liver and 7 HCC from 3 patients with hereditary haemochromatosis (HH) undergoing surgery. In order to investigate different disease phenotypes causing HCC, the data were compared with public microarray repositories for gene expression in normal liver, hepatitis C virus (HCV) cirrhosis, HCV-related HCC (HCV-HCC), hepatitis B virus (HBV) cirrhosis and HBV-related HCC (HBV-HCC). Principal component analysis and differential gene expression analysis were carried out using R Bioconductor. Liver disease-specific and shared gene lists were created and genes identified as highly expressed in hereditary haemochromatosis HCC (HH-HCC) were validated using quantitative RT-PCR. Selected genes were investigated further using immunohistochemistry in 86 HCC arising in liver disorders with varied aetiology. Using a 2-fold cut-off, 9 genes were highly expressed in all HCC, 11 in HH-HCC, 270 in HBV-HCC and 9 in HCV-HCC. Six genes identified by microarray as highly expressed in HH-HCC were confirmed by RT qPCR. Serine peptidase inhibitor, Kazal type 1 (SPINK1) mRNA was very highly expressed in HH-HCC (median fold change 2291, p = 0.0072) and was detected by immunohistochemistry in 91% of HH-HCC, 0% of HH-related cirrhotic or dysplastic nodules and 79% of mixed-aetiology HCC. Conclusion HCC, arising from diverse backgrounds, uniformly over-express a small set of genes. SPINK1, a secretory trypsin inhibitor, demonstrated

  19. Post-immunization immunohistochemical expression of Caspase 3 and p53 apoptotic markers in experimental hydatidosis.

    PubMed

    El-Aal, Amany Ahmed Abd; El-Gebaly, Naglaa Saad Mahmoud; Al-Antably, Abeer Said; Hassan, Marwa Adel; El-Dardiry, Marwa Ahmed

    2016-01-01

    The aim of this study was to investigate post-immunization apoptotic changes in experimental hydatidosis, using Caspase 3 and p53 immunohistochemical markers. Two groups of rabbits were immunized with a crude antigen (group 1) or a partially purified antigen (group 2) and were compared to an infected non-immunized control group. More effective immune responses were obtained in group 2 than group 1, signified by fewer and smaller cystic lesions and more severe destructive changes. Normal growth of cysts was attained in the control group, with no expression of apoptotic markers. Significantly higher expression of Caspase 3 and p53 were observed in group 1 compared to group 2, as indicated by OD and area percentage, respectively (Group 1 Caspase 3: 0.89±0.21, 93.5%±6.2; Group 1 p53: 0.46±0.18, 53.26%±11.6; Group 2 Caspase 3: 0.52±0.15, 49.23%±11.7; Group 2 p53: 0.19±0.4, 18.17%±7.3). Vaccine-induced immune responses and cellular damage may underlie the expression of apoptotic markers that appeared to result in a degenerative and atrophic course of action upon immunization. The results of the current study emphasize the importance of immunization for the stimulation of protective immune responses and in preventing mechanisms of evasion to ensure normal cell growth. A cost/benefit control program that implements proper vaccine preparations should be further assessed for complete elimination of severe infections in endemic areas. PMID:27683842

  20. [Expression of AIF and CGRP markers in pineal gland and thymus during aging].

    PubMed

    Lin'kova, N S; Katanugina, A S; Khavinson, V Kh

    2011-01-01

    We investigated the expression of AIF (apoptotic inducing factor) and CGRP (calcitonin gene related peptide) at autopsy material of pineal gland and thymus of people after 60 years old. The expression of AIF and CGRP was identified in both organs, but it did not change with age, which demonstrates the probable safety of functional activity of neuroimmunoendocrine system at aging. We found correlation between expression AIF and CGRP at pineal gland, but the correlation at thymus wasn't found. It is possible that pineal gland can express unidentified signal molecule controlling the expression of AIF and CGRP.

  1. Differential regulation of osteogenic marker gene expression by Wnt-3a in embryonic mesenchymal multipotential progenitor cells.

    PubMed

    Derfoul, Assia; Carlberg, Alyssa L; Tuan, Rocky S; Hall, David J

    2004-06-01

    The Wnt family of secreted glycoproteins plays an integral role in embryonic development and differentiation. To explore the role of Wnt's in one aspect of differentiation, namely osteogenesis, we employed a retroviral gene transfer approach to express Wnt-3a in the multipotent murine embryonic mesenchymal cell line C3H10T1/2. We found that expression of Wnt-3a in these cells had a significant, positive effect on cell growth in serum-containing medium, in that the cells grew to very high densities compared to the control cells. Additionally, apoptosis was markedly inhibited by Wnt-3a. However, when the cells were grown in serum-deficient medium, the Wnt-3a-expressing cells arrested efficiently in G1 phase, indicating that serum growth factors were needed in addition to Wnt-3a for enhanced proliferation. Wnt-3a-expressing cells exhibited high levels of alkaline phosphatase gene expression and enzymatic activity, but did not show any matrix mineralization. Unexpectedly, basal expression of bone sialoprotein, osteocalcin, and osteopontin were markedly inhibited by Wnt-3a, as were other known target genes of Wnt-3a, such as Brachyury, FGF-10, and Cdx1. When Wnt-3a-expressing cells were treated with osteogenic supplements in the presence of BMP-2, alkaline phosphatase gene expression and activity were further elevated. Additionally, BMP-2 was able to reverse the inhibitory effect of Wnt-3a on osteocalcin and osteopontin gene expression. These results indicate that while Wnt-3a represses basal expression of some osteogenic genes, this repression can be partially reversed by BMP-2. Finally, the enhanced gene expression of alkaline phosphatase induced by Wnt-3a could be effectively suppressed by the combined action of dexamethasone and 1,25-dihydroxyvitamin D(3). These data show for the first time that Wnt-3a has an unusual effect on multipotential embryonic cells, in that it enhances cellular proliferation and expression of alkaline phosphatase, while it represses most

  2. Vascular marker expression during the development of various types of gynaecological malignancy.

    PubMed

    Urban, Peter; Rabajdova, Miroslava Bilecova; Varga, Jan; Kruzliak, Peter; Fialkovicova, Viera; Durovcova, Eva; Marekova, Maria

    2014-11-01

    Clinical diagnosis of gynaecological malignancies is usually successful in the advanced stages of the tumour, and this has a major impact on the success of therapy. Therefore, in the last few years, cancer research has tried to identify and characterise new biochemical and molecular markers needed as predictive indicators for the diagnosis of cancer. Our aim has been to search the molecular changes in gene expression of death receptor 6, glycoprotein M6B (Gpm6B) and genes associated with tumours of the female genital system. After isolation of messenger RNA (mRNA), transcription of mRNA into the cDNA was performed. The quantification of gene expression changes was detected using the reverse transcription polymerase chain reaction (RT-PCR) method. Analysis at the protein level was performed using the Western blot method. In both methods, we used actin as a housekeeping gene for normalisation. Numerical quantification of changes in expression and in the level of the specific proteins was evaluated using the Data Syngene program. Significant changes in the levels of protein and mRNA expression were detected, mainly in the death receptor 6 (Dr6) gene of patients suffering from cancer of the corpus and cervix uteri and ovarian cancer, which also corresponded with the level of protein Dr6. At the level of transcription, a significant increase in the expression levels of mRNA for the Gpm6B gene was detected, which led to an increase in corresponding protein in the peripheral blood of patients with gynaecological tumours against the healthy control group. This article could help to find an adequate marker for clinical application that will enable more sensitive detection of the early stages of gynaecological malignancies from the peripheral blood of patients.

  3. Expression pattern of drought stress marker genes in soybean roots under two water deficit systems

    PubMed Central

    Neves-Borges, Anna Cristina; Guimarães-Dias, Fábia; Cruz, Fernanda; Mesquita, Rosilene Oliveira; Nepomuceno, Alexandre Lima; Romano, Eduardo; Loureiro, Marcelo Ehlers; de Fátima Grossi-de-Sá, Maria; Alves-Ferreira, Márcio

    2012-01-01

    The study of tolerance mechanisms for drought stress in soybean is fundamental to the understanding and development of tolerant varieties. Using in silico analysis, four marker genes involved in the classical ABA-dependent and ABA-independent pathways of drought response were identified in the Glycine max genome in the present work. The expression profiles of the marker genes ERD1-like, GmaxRD20A-like, GmaxRD22-like and GmaxRD29B-like were investigated by qPCR in root samples of drought sensitive and tolerant soybean cultivars (BR 16 and Embrapa 48, respectively), submitted to water deficit conditions in hydroponic and pot-based systems. Among the four putative soybean homologs to Arabidopsis genes investigated herein, only GmaxRD29B-like was not regulated by water deficit stress. Distinct expression profiles and different induction levels were observed among the genes, as well as between the two drought-inducing systems. Our results showed contrasting gene expression responses for the GmaxRD20A-like and GmaxRD22-like genes. GmaxRD20A-like was highly induced by continuous drought acclimating conditions, whereas GmaxRD22-like responses decreased after abrupt water deprivation. GmaxERD1-like showed a different expression profile for the cultivars in each system. Conversely, GmaxRD20A-like and GmaxRD22-like genes exhibited similar expression levels in tolerant plants in both systems. PMID:22802707

  4. Differential expression of minimal residual disease markers in peripheral blood and bone marrow samples from high-risk neuroblastoma patients

    PubMed Central

    YAMAMOTO, NOBUYUKI; KOZAKI, AIKO; HARTOMO, TRI BUDI; YANAI, TOMOKO; HASEGAWA, DAIICHIRO; KAWASAKI, KEIICHIRO; KOSAKA, YOSHIYUKI; MATSUO, MASAFUMI; HIRASE, SATOSHI; MORI, TAKESHI; HAYAKAWA, AKIRA; IIJIMA, KAZUMOTO; NISHIO, HISAHIDE; NISHIMURA, NORIYUKI

    2015-01-01

    Neuroblastoma is an aggressive solid tumor that leads to tumor relapse in more than half of high-risk patients. Minimal residual disease (MRD) is primarily responsible for tumor relapses and may be detected in peripheral blood (PB) and bone marrow (BM) samples. To evaluate the disease status and treatment response, a number of MRD detection protocols based on either common or distinct markers for PB and BM samples have been reported. However, the correlation between the expression of MRD markers in PB and BM samples remains elusive in the clinical samples. In the present study, the expression of 11 previously validated MRD markers (CHRNA3, CRMP1, DBH, DCX, DDC, GABRB3, GAP43, ISL1, KIF1A, PHOX2B and TH) was determined in 23 pairs of PB and BM samples collected from seven high-risk neuroblastoma patients at the same time point, and the sample was scored as MRD-positive if one of the MRD markers exceeded the normal range. Although the number of MRD-positive samples was not significantly different between PB and BM samples, the two most sensitive markers for PB samples (CRMP1 and KIF1A) were different from those for BM samples (PHOX2B and DBH). There was no statistically significant correlation between the expression of MRD markers in the PB and BM samples. These results suggest that MRD markers were differentially expressed in PB and BM samples from high-risk neuroblastoma patients. PMID:26722317

  5. Mining and gene ontology based annotation of SSR markers from expressed sequence tags of Humulus lupulus.

    PubMed

    Singh, Swati; Gupta, Sanchita; Mani, Ashutosh; Chaturvedi, Anoop

    2012-01-01

    Humulus lupulus is commonly known as hops, a member of the family moraceae. Currently many projects are underway leading to the accumulation of voluminous genomic and expressed sequence tag sequences in public databases. The genetically characterized domains in these databases are limited due to non-availability of reliable molecular markers. The large data of EST sequences are available in hops. The simple sequence repeat markers extracted from EST data are used as molecular markers for genetic characterization, in the present study. 25,495 EST sequences were examined and assembled to get full-length sequences. Maximum frequency distribution was shown by mononucleotide SSR motifs i.e. 60.44% in contig and 62.16% in singleton where as minimum frequency are observed for hexanucleotide SSR in contig (0.09%) and pentanucleotide SSR in singletons (0.12%). Maximum trinucleotide motifs code for Glutamic acid (GAA) while AT/TA were the most frequent repeat of dinucleotide SSRs. Flanking primer pairs were designed in-silico for the SSR containing sequences. Functional categorization of SSRs containing sequences was done through gene ontology terms like biological process, cellular component and molecular function.

  6. Characterization of genic microsatellite markers derived from expressed sequence tags in Pacific abalone ( Haliotis discus hannai)

    NASA Astrophysics Data System (ADS)

    Li, Qi; Shu, Jing; Zhao, Cui; Liu, Shikai; Kong, Lingfeng; Zheng, Xiaodong

    2010-01-01

    Simple sequence repeat (SSR) markers were developed from the expressed sequence tags (ESTs) of Pacific abalone ( Haliotis discus hannai). Repeat motifs were found in 4.95% of the ESTs at a frequency of one repeat every 10.04 kb of EST sequences, after redundancy elimination. Seventeen polymorphic EST-SSRs were developed. The number of alleles per locus varied from 2-17, with an average of 6.8 alleles per locus. The expected and observed heterozygosities ranged from 0.159 to 0.928 and from 0.132 to 0.922, respectively. Twelve of the 17 loci (70.6%) were successfully amplified in H. diversicolor. Seventeen loci segregated in three families, with three showing the presence of null alleles (17.6%). The adequate level of variability and low frequency of null alleles observed in H. discus hannai, together with the high rate of transportability across Haliotis species, make this set of EST-SSR markers an important tool for comparative mapping, marker-assisted selection, and evolutionary studies, not only in the Pacific abalone, but also in related species.

  7. Differential expression of immune-related markers in breast cancer by molecular phenotypes.

    PubMed

    Choi, Junjeong; Kim, Do Hee; Jung, Woo Hee; Koo, Ja Seung

    2013-01-01

    The purpose of this study is to investigate the relationship between expression of immune-related molecules such as STAT1, CD20, IL-8, IFN-γ, tumor genetic phenotype, and the clinical course of invasive breast cancer. We constructed tissue microarrays from the breast cancers of 727 patients and classified the cases as either luminal A, luminal B, HER-2, or triple negative breast cancer (TNBC) based on standard pathological and clinical classifications using genetic phenotype. Surrogate immunohistochemical stains (STAT1, CD20, IL-8, IFN-γ) and HER-2 FISH were performed on each microarray. Of the 727 patients cases, 303 (41.7 %) were luminal A, 169 (23.2 %) were luminal B, 71 (9.8 %) were HER2+, and 184 (25.3 %) were TNBC. The expression of STAT1 in tumor cells was higher in luminal-type cancers than in HER2+ and TNBC (P < 0.001), and the TNBC-type tumors showed the highest levels of stromal STAT1 expression (P < 0.001), stromal IL-8 expression (P = 0.005), and CD20 index (P < 0.001). Luminal A type tumors showed the lowest expression of these markers. The stromal IL-8 positivity was associated with shorter DFS and OS in ER positive group, HER-2 negative group, and luminal A group (P < 0.05). To conclude, the immune-related molecules, STAT1, IFN-γ, IL-8, and CD20 are differentially expressed and define particular molecular subtypes which correlate with genetically defined types of tumors. High expression of STAT1 in tumor cells is observed in luminal-type tumors, whereas stromal expression of STAT1, stromal IL-8, and IL-8 in tumor cells is the highest in TNBC-type tumors.

  8. LFA-1 expression on exocrine glands as a potential novel marker of malignant disease.

    PubMed

    Futagami-Mizoguchi, E; Yamada, A; Mizoguchi, A; Imai, Y; Yokoyama, M M

    1993-09-01

    The lymphocyte function associated antigen 1 has been found only on leukocytes and lymphoid tissues; however, the expression of lymphocyte function associated antigen 1 on nonhematopoietic cells has not been reported previously. In this study, immunohistochemical expression of lymphocyte function associated antigen 1 was examined on various tissues from 35 patients with malignant diseases and 36 patients with benign diseases including benign tumors. The expression of lymphocyte function associated antigen 1 was found on various exocrine tissues (eg, gastric glands, bronchial epithelium, alveolar epithelium, duodenal glands, bile ducts, pancreatic acini, and salivary glands) uninvolved by tumor in patients with malignant diseases. Localization of lymphocyte function associated antigen 1 was limited to the exocrine glands and differed from tissue-infiltrating leukocytes. The expression of lymphocyte function associated antigen 1 on exocrine tissues was confirmed in all 35 cases of malignant diseases that were examined. These included a wide spectrum of carcinomas and hematopoietic tumors. In contrast, none of the 36 cases with benign diseases examined expressed lymphocyte function associated antigen 1 on their exocrine glands. These results indicate a strong correlation between lymphocyte function associated antigen 1 expression on exocrine glands and malignant disease. The expression of lymphocyte function associated antigen 1 on nonhematopoietic cells was further confirmed in nonhematopoietic cell lines. Two of 19 nonhematopoietic cell lines (MKN45 and PANC-1; exocrine gland cell lines) examined expressed lymphocyte function associated antigen 1 on both cell surface and cytoplasm. These results suggested that immunohistochemically defined lymphocyte function associated antigen 1 molecules on nontumorous exocrine gland cells are a potential marker for the presence of malignant diseases.

  9. Expression of endogenous retroviruses is negatively regulated by the pluripotency marker Rex1/Zfp42

    PubMed Central

    Guallar, D.; Pérez-Palacios, R.; Climent, M.; Martínez-Abadía, I.; Larraga, A.; Fernández-Juan, M.; Vallejo, C.; Muniesa, P.; Schoorlemmer, J.

    2012-01-01

    Rex1/Zfp42 is a Yy1-related zinc-finger protein whose expression is frequently used to identify pluripotent stem cells. We show that depletion of Rex1 levels notably affected self-renewal of mouse embryonic stem (ES) cells in clonal assays, in the absence of evident differences in expression of marker genes for pluripotency or differentiation. By contrast, marked differences in expression of several endogenous retroviral elements (ERVs) were evident upon Rex1 depletion. We demonstrate association of REX1 to specific elements in chromatin-immunoprecipitation assays, most strongly to muERV-L and to a lower extent to IAP and musD elements. Rex1 regulates muERV-L expression in vivo, as we show altered levels upon transient gain-and-loss of Rex1 function in pre-implantation embryos. We also find REX1 can associate with the lysine-demethylase LSD1/KDM1A, suggesting they act in concert. Similar to REX1 binding to retrotransposable elements (REs) in ES cells, we also detected binding of the REX1 related proteins YY1 and YY2 to REs, although the binding preferences of the two proteins were slightly different. Altogether, we show that Rex1 regulates ERV expression in mouse ES cells and during pre-implantation development and suggest that Rex1 and its relatives have evolved as regulators of endogenous retroviral transcription. PMID:22844087

  10. Sexually dimorphic gene expressions in eels: useful markers for early sex assessment in a conservation context

    PubMed Central

    Geffroy, Benjamin; Guilbaud, Florian; Amilhat, Elsa; Beaulaton, Laurent; Vignon, Matthias; Huchet, Emmanuel; Rives, Jacques; Bobe, Julien; Fostier, Alexis; Guiguen, Yann; Bardonnet, Agnès

    2016-01-01

    Environmental sex determination (ESD) has been detected in a range of vertebrate reptile and fish species. Eels are characterized by an ESD that occurs relatively late, since sex cannot be histologically determined before individuals reach 28 cm. Because several eel species are at risk of extinction, assessing sex at the earliest stage is a crucial management issue. Based on preliminary results of RNA sequencing, we targeted genes susceptible to be differentially expressed between ovaries and testis at different stages of development. Using qPCR, we detected testis-specific expressions of dmrt1, amh, gsdf and pre-miR202 and ovary-specific expressions were obtained for zar1, zp3 and foxn5. We showed that gene expressions in the gonad of intersexual eels were quite similar to those of males, supporting the idea that intersexual eels represent a transitional stage towards testicular differentiation. To assess whether these genes would be effective early molecular markers, we sampled juvenile eels in two locations with highly skewed sex ratios. The combined expression of six of these genes allowed the discrimination of groups according to their potential future sex and thus this appears to be a useful tool to estimate sex ratios of undifferentiated juvenile eels. PMID:27658729

  11. Differential Expression of Stem Cell Markers in Ocular Surface Squamous Neoplasia.

    PubMed

    Mishra, Dilip Kumar; Veena, Uppala; Kaliki, Swathi; Kethiri, Abhinav Reddy; Sangwan, Virender S; Ali, Mohammed Hasnat; Naik, Milind N; Singh, Vivek

    2016-01-01

    Ocular Surface Squamous Neoplasm (OSSN) is the neoplasia arising from the conjunctiva, cornea and limbus. OSSN ranges from mild, moderate, severe dysplasia, carcinoma in situ (CIS) to squamous cell carcinoma (SCC). Recent findings on cancer stem cells theory indicate that population of stem-like cell as in neoplasia determines its heterogeneity and complexity leading to varying tumor development of metastatic behavior and recurrence. Cancer stem cell markers are not much explored in the cases of OSSN. In the present study, we aim to evaluate the expression of stem cells using stem cell markers mainly p63, ABCG2, c-KIT (CD117) and CD44 in OSSN tissue, which could have prognostic significance. The present study tries for the first time to explore expression of these stem markers in the cases of OSSN. These cases are subdivided into two groups. One group comprises of carcinoma in situ (n = 6) and the second group comprises of invasive carcinoma (n = 6). The mean age at presentation was 52 years; with 53 years for CIS group and 52 years for SCC group. From each group section from the paraffin block were taken for the IHC staining of p63, c-Kit, ABCG2 and CD44. Our experiments show high expression of P63 and CD44 in the cases of CIN and SCC. Both CIS and SCC displayed positive staining with p63, with more than 80% cells staining positive. However minimal expression of c-kit in both CIN and SCC. But surprisingly we got high expression of ABCG2 in cases of carcinoma in situ as compared to that of invasive squamous cell carcinoma. More than 50% of cells showed CD44 positivity in both CIS and SCC groups. Our results show for the first time that these four stem cells especially the limbal epithelium stem cells play a vital role in the genesis of OSSN but we need to explore more cases before establishing its clinical and biological significance. PMID:27584160

  12. Differential Expression of Stem Cell Markers in Ocular Surface Squamous Neoplasia

    PubMed Central

    Mishra, Dilip Kumar; Veena, Uppala; Kaliki, Swathi; Kethiri, Abhinav Reddy; Sangwan, Virender S.; Ali, Mohammed Hasnat; Naik, Milind N.; Singh, Vivek

    2016-01-01

    Ocular Surface Squamous Neoplasm (OSSN) is the neoplasia arising from the conjunctiva, cornea and limbus. OSSN ranges from mild, moderate, severe dysplasia, carcinoma in situ (CIS) to squamous cell carcinoma (SCC). Recent findings on cancer stem cells theory indicate that population of stem-like cell as in neoplasia determines its heterogeneity and complexity leading to varying tumor development of metastatic behavior and recurrence. Cancer stem cell markers are not much explored in the cases of OSSN. In the present study, we aim to evaluate the expression of stem cells using stem cell markers mainly p63, ABCG2, c-KIT (CD117) and CD44 in OSSN tissue, which could have prognostic significance. The present study tries for the first time to explore expression of these stem markers in the cases of OSSN. These cases are subdivided into two groups. One group comprises of carcinoma in situ (n = 6) and the second group comprises of invasive carcinoma (n = 6). The mean age at presentation was 52 years; with 53 years for CIS group and 52 years for SCC group. From each group section from the paraffin block were taken for the IHC staining of p63, c-Kit, ABCG2 and CD44. Our experiments show high expression of P63 and CD44 in the cases of CIN and SCC. Both CIS and SCC displayed positive staining with p63, with more than 80% cells staining positive. However minimal expression of c-kit in both CIN and SCC. But surprisingly we got high expression of ABCG2 in cases of carcinoma in situ as compared to that of invasive squamous cell carcinoma. More than 50% of cells showed CD44 positivity in both CIS and SCC groups. Our results show for the first time that these four stem cells especially the limbal epithelium stem cells play a vital role in the genesis of OSSN but we need to explore more cases before establishing its clinical and biological significance. PMID:27584160

  13. Following damage, the majority of bone marrow-derived airway cells express an epithelial marker

    PubMed Central

    MacPherson, Heather; Keir, Pamela A; Edwards, Carol J; Webb, Sheila; Dorin, Julia R

    2006-01-01

    Background Adult-derived bone marrow stem cells are capable of reconstituting the haematopoietic system. However there is ongoing debate in the literature as to whether bone marrow derived cells have the ability to populate other tissues and express tissue specific markers. The airway has been an organ of major interest and was one of the first where this was demonstrated. We have previously demonstrated that the mouse airway can be repopulated by side population bone marrow transplanted cells. Here we investigate the frequency and phenotypic nature of these bone marrow derived cells. Methods Female mice were engrafted with male whole bone marrow or side population (SP) cells and subjected to detergent-induced damage after 3 months. Donor cells were identified by Y chromosome fluorescence in situ hybridisation and their phenotype was assessed by immunohistochemistry on the same sections. Slides were visualised by a combination of widefield and deconvolved microscopy and whole cells were analysed on cytospin preparations. Results The frequencies of engraftment of male cells in the airway of mice that show this (9/10), range from 1.0 – 1.6% with whole marrow and 0.6 – 1.5% with SP cells. Undamaged controls have only between 0.1 and 0.2% male cells in the trachea. By widefield microscopy analysis we find 60.2% (53/88) of male donor derived cells express cytokeratins as a marker of epithelial cells. These results were reinforced using deconvolved microscopy and scored by two independent investigators. In addition cytospin analysis of cells dissociated from the damaged trachea of engrafted mice also reveals donor derived Y chromosome positive cells that are immunopositive for cytokeratin. Using cytokeratin and the universal haematopoietic marker CD45 immunohistochemistry, we find the donor derived cells fall into four phenotypic classes. We do not detect cytokeratin positive cells in whole bone marrow using cytokeratin immunostaining and we do not detect any

  14. Inhibition of hypoxia inducible factor-1α downregulates the expression of epithelial to mesenchymal transition early marker proteins without undermining cell survival in hypoxic lens epithelial cells

    PubMed Central

    Neelam, Sudha; Brooks, Morgan M.

    2015-01-01

    Purpose The purpose of this study was to identify potential therapeutic strategies to slow down or prevent the expression of early-onset epithelial to mesenchymal transition (EMT) marker proteins (fibronectin and alpha smooth muscle actin, α-SMA) without sacrificing the synthesis and accumulation of the prosurvival protein vascular endothelial growth factor (VEGF) in cultured virally transformed human lens epithelial (HLE) cells. Methods HLE-B3 cells, maintained in a continuous hypoxic environment (1% oxygen), were treated with SB216763, a specific inhibitor of glycogen synthase kinase-3β (GSK-3β) catalytic activity. Western blot analysis was employed to detect the cytoplasmic and nuclear levels of β-catenin, as well as the total lysate content of fibronectin and α-SMA. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of VEGF in cell culture medium. A hypoxia-inducible factor-1α (HIF-1α) translation inhibitor and an HIF-2α translation inhibitor were independently employed to evaluate the effect of hypoxia inducible factor inhibition on EMT marker protein and VEGF expression. XAV932 was used to assess the suppression of nuclear β-catenin and its downstream effect on EMT marker proteins and VEGF expression. Results SB216763-treated HLE-B3 cells caused marked inhibition of GSK-3β activity prompting a significant increase in the translocation of cytoplasmic β-catenin to the nucleus. The enhancement of nuclear β-catenin looked as if it positively correlated with a significant increase in the basal expression of VEGF as well as increased expression of fibronectin and α-SMA. In conjunction with SB216763, coadministration of an HIF-1α translation inhibitor, but not an HIF-2α translation inhibitor, markedly suppressed the expression of fibronectin and α-SMA without affecting VEGF levels. Treatment with XAV932 significantly reduced the level of nuclear β-catenin, but the levels of neither the EMT marker proteins nor VEGF were changed

  15. Molecular cloning of the lymphocyte activation marker Blast-1.

    PubMed Central

    Staunton, D E; Thorley-Lawson, D A

    1987-01-01

    Blast-1 is an early activation-associated glycoprotein expressed on the surface of human lymphocytes. Here we report the isolation and analysis of a cDNA encoding Blast-1. The translated sequence of the Blast-1 cDNA contains a hydrophobic putative signal peptide and a hydrophobic carboxyl terminus devoid of charged residues. The sequence also contains five N-linked glycosylation sites, the utilization of which was confirmed by the shift in relative mol. wt of Blast-1 upon digestion with N-glycosidase F. The translated sequence reveals that Blast-1 is related to members of the immunoglobulin superfamily, especially to CD4 and MHC class II molecules. The homology to these proteins is greatest in their amino termini where they demonstrate 30-32% identity. This region of Blast-1 also demonstrated 25% identity to a V kappa sequence. Considering conservative amino acid substitutions this homology to CD4, MHC class II and V kappa becomes 60, 49 and 48%, respectively. Images Fig. 1. Fig. 3. Fig. 4. PMID:2828034

  16. Gene expression profile in breast cancer comprising predictive markers for metastatic risk.

    PubMed

    Sirirattanakul, S; Wannakrairot, P; Tencomnao, T; Santiyanont, R

    2015-01-01

    Quantitative multiplex reverse transcriptase-polymerase chain reaction was developed for the simultaneous detection of multiple-gene expression levels of formalin-fixed, paraffin-embedded breast cancer samples. Candidate genes were selected from previous microarray data relevant to breast cancer markers that had the potential to serve as predictive markers for metastatic risk. This multiplex gene set included 11 candidate and 3 housekeeping genes, and the aim was to predict breast cancer progression based on lymph node involvement status. Our study demonstrated that the system generated a good standard curve fit (R(2) = 0.9901-0.9998) correlated with RNA concentration. The multiplex gene expression profile indicated significantly downregulated levels of G protein-coupled receptor kinase interacting ArfGAP 2 (GIT2) and mitochondrial transcription termination factor (MTERF) genes in a lymph node-positive group of patients, with P values of 0.004 and 0.038, respectively. Therefore, this in-house method using multiple genes of interest might be an alternative tool for prediction of breast cancer metastasis. PMID:26400320

  17. Relationship of CD86 surface marker expression and cytotoxicity on dendritic cells exposed to chemical allergen

    SciTech Connect

    Hulette, Ben C.; Ryan, Cindy A.; Gildea, Lucy A.; Gerberick, G. Frank . E-mail: gerberick.gf@pg.com

    2005-12-01

    Human peripheral blood-derived dendritic cells (DC) respond to a variety of chemical allergens by up-regulating expression of the co-stimulatory molecule CD86. It has been postulated that this measure might provide the basis for an in vitro alternative approach for the identification of skin sensitizing chemicals. We recently reported that DC, exposed in culture to the highest non-cytotoxic concentrations of various chemical allergens, displayed marginal up-regulation of membrane CD86 expression; the interpretation being that such changes were insufficiently sensitive for the purposes of hazard identification. For the work presented here, immature DC were derived from human monocytes and treated with the chemical allergens 2,4-dinitrobenzenesulfonic acid (DNBS), nickel sulfate (NiSO{sub 4}), p-phenylenediamine (PPD), Bandrowski's base (BB), hydroquinone (HQ) and propyl gallate (PG) for 48 h at concentrations which induced both no to slight to moderate cytotoxicity. For comparison, DC were treated with the irritants sodium dodecyl sulfate (SDS), benzoic acid (BA), and benzalkonium chloride (BZC) at concentrations resulting in comparable levels of cytotoxicity. CD86 expression, as measured by flow cytometry, was consistently up-regulated (ranging from 162 to 386% control) on DC treated with concentrations of chemical allergens that induced approximately 10-15% cytotoxicity. The irritants BA and BZC did not induce up-regulation of CD86 expression when tested at concentrations that induced similar levels of cytotoxicity. SDS, however, up-regulated CD86 expression to 125-138% of control in 2/4 preparations when tested at concentrations which induced similar toxicity. Our results confirm that chemical allergens up-regulate CD86 expression on blood-derived DC and illustrate further that up-regulation of CD86 surface marker expression is more robust when DC are treated with concentrations of chemical allergen that induce slight to moderate cytotoxicity.

  18. Leptin receptor expression and Gln223Arg polymorphism as prognostic markers in oral and oropharyngeal cancer.

    PubMed

    Rodrigues, P R S; Maia, L L; Santos, M; Peterle, G T; Alves, L U; Takamori, J T; Souza, R P; Barbosa, W M; Mercante, A M C; Nunes, F D; Carvalho, M B; Tajara, E H; Louro, I D; Silva-Conforti, A M A

    2015-11-25

    The leptin gene product is released into the blood stream, passes through the blood-brain barrier, and finds the leptin receptor (LEPR) in the central nervous system. This hormone regulates food intake, hematopoiesis, inflammation, immunity, differentiation, and cell proliferation. The LEPR Gln223Arg polymorphism has been reported to alter receptor function and expression, both of which have been related with prognostics in several tumor types. Furthermore, several studies have shown a relationship between the Gln223Arg polymorphism and tumor development, and its role in oral and oropharyngeal squamous cell carcinoma is now well understood. In this study, 315 DNA samples were used for LEPR Gln223Arg genotyping and 87 primary oral and oropharyngeal squamous cell carcinomas were used for immunohistochemical expression analysis, such that a relationship between these and tumor development and prognosis could be established. Homozygous LEPR Arg223 was found to be associated with a 2-fold reduction in oral and oropharyngeal cancer risk. In contrast, the presence of the Arg223 allele in tumors was associated with worse disease-free and disease-specific survival. Low LEPR expression was found to be an independent risk factor, increasing the risk for lymph node metastasis 4-fold. In conclusion, the Gln223Arg polymorphism and LEPR expression might be valuable markers for oral and oropharyngeal cancer, suggesting that LEPR might serve as a potential target for future therapies.

  19. The expression of retinal cell markers in human retinal pigment epithelial cells and their augmentation by the synthetic retinoid fenretinide

    PubMed Central

    Vugler, Anthony A.; Yu, Lu; Semo, Maayan; Coffey, Pete; Moss, Stephen E.; Greenwood, John

    2011-01-01

    Purpose In several species the retinal pigment epithelium (RPE) has the potential to transdifferentiate into retinal cells to regenerate functional retinal tissue after injury. However, this capacity for regeneration is lost in mammals. The synthetic retinoic acid derivative, fenretinide [N(4-hydroxyphenyl) retinamide], induces a neuronal-like phenotype in the human adult retinal pigment epithelial cell line (ARPE-19). These changes are characterized by the appearance of neural-like processes and the expression of neuronal markers not normally associated with RPE cells. Here we assess whether fenretinide can induce a neuroretinal cell phenotype in ARPE-19 cells, by examining retinal cell marker expression. Methods ARPE-19 cells were treated daily with culture medium containing either 3 μM fenretinide or dimethyl sulfoxide as a control for 7 days. Cells were processed for immunocytochemistry, western blotting, and for analysis by PCR to examine the expression of a panel of RPE, neural, and retinal-associated cellular markers, including classical and non-canonical opsins. Results Treatment with fenretinide for 7 days induced the formation of neuronal-like processes in ARPE-19 cells. Fenretinide induced the expression of the cone long wavelength sensitive opsin (OPN1lw) but not rhodopsin (RHO), while decreasing the expression of RPE cell markers. Many of the neuronal and retinal specific markers examined were expressed in both control and fenretinide treated cells, including those involved in photoreceptor cell development and the multipotency of neural retinal progenitor cells. Interestingly, ARPE-19 cells also expressed both photoreceptor specific and non-specific canonical opsins. Conclusions The expression of retinal-associated markers and loss of RPE cell markers in control ARPE-19 cells suggests that these cells might have dedifferentiated from an RPE cell phenotype under standard culture conditions. The expression of molecules, such as the transcription

  20. [Construction of Fat-1 eukaryotic expression vector of excision markers and the establishment of transgenic sheep cell lines].

    PubMed

    Lima, A; Zhu, Heping; Wang, Ruiyao; Yan, Tao; Su, Xiaohu; Li, Lu; Wang, Bingping; Na, Shunwendoule; Qi, Guichun; Zhou, Huanmin

    2016-02-01

    In order to establish marker-free transgenic cell lines, we cloned Fat-1 gene, attB and Loxp sequences by PCR. Then we inserted these sequences to pN1-EGFP vector and got pEGFP-N1-Fat-1 expression vector. PhiC31 integrase mRNA which was generated by in vitro transcription and a pEGFP-N1-Fat-1 expression vector co-electroporated into sheep fetal fibroblasts, and then we got transgenic cell lines expressing green fluorescence. Prokaryotic expression and purification of Cre recombinant protein was performed. Cre recombinant protein was transducted into stably-transfected cell colonies. We identified cell colonies by sequencing and established marker-free transgenic cell lines and eventually- established marker-free transgenic cell lines which were building more safely basic for producing Fat-1 transgenic animals. PMID:27382771

  1. [Construction of Fat-1 eukaryotic expression vector of excision markers and the establishment of transgenic sheep cell lines].

    PubMed

    Lima, A; Zhu, Heping; Wang, Ruiyao; Yan, Tao; Su, Xiaohu; Li, Lu; Wang, Bingping; Na, Shunwendoule; Qi, Guichun; Zhou, Huanmin

    2016-02-01

    In order to establish marker-free transgenic cell lines, we cloned Fat-1 gene, attB and Loxp sequences by PCR. Then we inserted these sequences to pN1-EGFP vector and got pEGFP-N1-Fat-1 expression vector. PhiC31 integrase mRNA which was generated by in vitro transcription and a pEGFP-N1-Fat-1 expression vector co-electroporated into sheep fetal fibroblasts, and then we got transgenic cell lines expressing green fluorescence. Prokaryotic expression and purification of Cre recombinant protein was performed. Cre recombinant protein was transducted into stably-transfected cell colonies. We identified cell colonies by sequencing and established marker-free transgenic cell lines and eventually- established marker-free transgenic cell lines which were building more safely basic for producing Fat-1 transgenic animals.

  2. Effect of N-acetylcysteine on the early expression of inflammatory markers in the retina and plasma of diabetic rats

    PubMed Central

    Tsai, Gina Y; Cui, Jing Z; Syed, Husnain; Xia, Zhengyuan; Ozerdem, Ugur; McNeill, John H; Matsubara, Joanne A

    2014-01-01

    Purpose The aim of this study is to investigate markers of inflammation and oxidative stress in an early model of diabetic retinopathy, correlate retinal and plasma results and evaluate the influence of treatment by N-acetylcysteine (NAC), a free radical scavenger. Methods Four groups were studied: control (C), streptozotocin (STZ)-induced diabetic rats (D), STZ rats following 8 weeks of NAC (DT), and control rats following 8 weeks of NAC (CT). Plasma levels of free 15-F2t-isoprostane (15-F-2t-IsoP), superoxide dismutase (SOD) and tumour necrosis factor-alpha (TNF-α) were obtained. Primary antibodies against macrophages (ED-1), microglia (Ox-42), pericytes (NG-2), endothelial and perivascular cells (IB-4), haem oxygenase 1 (HO-1) and vascular endothelial growth factor (VEGF) were used. Results Expression of NG-2 was robust in C, CT, DT, and mild in D. The intensity of IB-4 was higher in D and DT compared with the C and CT. Ox-42 and ED-1 expression was higher in the D than in the DT, C or CT. Expression of VEGF and HO-1 was non-specific across the four groups. Plasma levels of 15-F-2t-IsoP and TNF-α were higher in the D as compared with the C, CT and DT. SOD levels were lower in the D when compared with the C, CT and D. Conclusions Macrophage/microglia activation, pericyte loss and endothelial/perivascular cell changes occur early in the pathogenesis of DR. These changes are associated with an increase in plasma markers of oxidative stress and inflammation and are minimized by treatment with NAC. The results suggest that therapies that reduce free radicals will help minimize the early events in diabetic retinopathy in the STZ model. PMID:19723131

  3. The Early Expression of HLA-DR and CD64 Myeloid Markers Is Specifically Compartmentalized in the Blood and Lungs of Patients with Septic Shock

    PubMed Central

    Mikaszewska-Sokolewicz, Małgorzata; Hoser, Grażyna; Zielińska-Borkowska, Urszula

    2016-01-01

    Identification of reliable biomarkers is key to guide targeted therapies in septic patients. Expression monitoring of monocyte HLA-DR and neutrophil CD64 could fulfill the above need. However, it is unknown whether their expression on circulating cells reflects the status of tissue resident cells. We compared expressions of HLA-DR and CD64 markers in the circulation and airways of septic shock patients and evaluated their outcome prognostic value. The expression of CD64 on neutrophils and HLA-DR on monocytes was analyzed in the peripheral blood and mini-bronchoalveolar lavage fluid cells by flow cytometry. Twenty-seven patients with septic shock were enrolled into the study. The fluorescence intensity of HLA-DR on circulating monocytes was 3.5-fold lower than on the pulmonary monocytes (p = 0.01). The expression of CD64 on circulating and airway neutrophils was similar (p = 0.47). Only the expression of CD64 on circulating neutrophils was higher in nonsurvivors versus survivors (2.8-fold; p = 0.031). Pulmonary monocytes display a higher level of HLA-DR activation compared to peripheral blood monocytes but the expression of neutrophil CD64 is similar on lung and circulating cells. Death in septic patients was effectively predicted by neutrophil CD64 but not monocytic HLA-DR. Prognostic value of cellular activation markers in septic shock appears to strongly depend on their level of compartmentalization. PMID:27413252

  4. Identification of Gene-Expression Signatures and Protein Markers for Breast Cancer Grading and Staging

    PubMed Central

    Yao, Fang; Zhang, Chi; Du, Wei; Liu, Chao; Xu, Ying

    2015-01-01

    The grade of a cancer is a measure of the cancer's malignancy level, and the stage of a cancer refers to the size and the extent that the cancer has spread. Here we present a computational method for prediction of gene signatures and blood/urine protein markers for breast cancer grades and stages based on RNA-seq data, which are retrieved from the TCGA breast cancer dataset and cover 111 pairs of disease and matching adjacent noncancerous tissues with pathologists-assigned stages and grades. By applying a differential expression and an SVM-based classification approach, we found that 324 and 227 genes in cancer have their expression levels consistently up-regulated vs. their matching controls in a grade- and stage-dependent manner, respectively. By using these genes, we predicted a 9-gene panel as a gene signature for distinguishing poorly differentiated from moderately and well differentiated breast cancers, and a 19-gene panel as a gene signature for discriminating between the moderately and well differentiated breast cancers. Similarly, a 30-gene panel and a 21-gene panel are predicted as gene signatures for distinguishing advanced stage (stages III-IV) from early stage (stages I-II) cancer samples and for distinguishing stage II from stage I samples, respectively. We expect these gene panels can be used as gene-expression signatures for cancer grade and stage classification. In addition, of the 324 grade-dependent genes, 188 and 66 encode proteins that are predicted to be blood-secretory and urine-excretory, respectively; and of the 227 stage-dependent genes, 123 and 51 encode proteins predicted to be blood-secretory and urine-excretory, respectively. We anticipate that some combinations of these blood and urine proteins could serve as markers for monitoring breast cancer at specific grades and stages through blood and urine tests. PMID:26375396

  5. Long and short photoperiod buds in hybrid aspen share structural development and expression patterns of marker genes

    PubMed Central

    Rinne, Päivi L.H.; Paul, Laju K.; Vahala, Jorma; Ruonala, Raili; Kangasjärvi, Jaakko; van der Schoot, Christiaan

    2015-01-01

    Tree architecture develops over time through the collective activity of apical and axillary meristems. Although the capacity of both meristems to form buds is crucial for perennial life, a comparative analysis is lacking. As shown here for hybrid aspen, axillary meristems engage in an elaborate process of axillary bud (AXB) formation, while apical dominance prevents outgrowth of branches. Development ceased when AXBs had formed an embryonic shoot (ES) with a predictable number of embryonic leaves at the bud maturation point (BMP). Under short days, terminal buds (TBs) formed an ES similar to that of AXBs, and both the TB and young AXBs above the BMP established dormancy. Quantitative PCR and in situ hybridizations showed that this shared ability and structural similarity was reflected at the molecular level. TBs and AXBs similarly regulated expression of meristem-specific and bud/branching-related genes, including CENTRORADIALIS-LIKE1 (CENL1), BRANCHED1 (BRC1), BRC2, and the strigolactone biosynthesis gene MORE AXILLARY BRANCHES1 (MAX1). Below the BMP, AXBs maintained high CENL1 expression at the rib meristem, suggesting that it serves to maintain poise for growth. In support of this, decapitation initiated outgrowth of CENL1-expressing AXBs, but not of dormant AXBs that had switched CENL1 off. This singles out CENL1 as a rib meristem marker for para-dormancy. BRC1 and MAX1 genes, which may counterbalance CENL1, were down-regulated in decapitation-activated AXBs. The results showed that removal of apical dominance shifted AXB gene expression toward that of apices, while developing TBs adopted the expression pattern of para-dormant AXBs. Bud development thus follows a shared developmental pattern at terminal and axillary positions, despite being triggered by short days and apical dominance, respectively. PMID:26248666

  6. Long and short photoperiod buds in hybrid aspen share structural development and expression patterns of marker genes.

    PubMed

    Rinne, Päivi L H; Paul, Laju K; Vahala, Jorma; Ruonala, Raili; Kangasjärvi, Jaakko; van der Schoot, Christiaan

    2015-11-01

    Tree architecture develops over time through the collective activity of apical and axillary meristems. Although the capacity of both meristems to form buds is crucial for perennial life, a comparative analysis is lacking. As shown here for hybrid aspen, axillary meristems engage in an elaborate process of axillary bud (AXB) formation, while apical dominance prevents outgrowth of branches. Development ceased when AXBs had formed an embryonic shoot (ES) with a predictable number of embryonic leaves at the bud maturation point (BMP). Under short days, terminal buds (TBs) formed an ES similar to that of AXBs, and both the TB and young AXBs above the BMP established dormancy. Quantitative PCR and in situ hybridizations showed that this shared ability and structural similarity was reflected at the molecular level. TBs and AXBs similarly regulated expression of meristem-specific and bud/branching-related genes, including CENTRORADIALIS-LIKE1 (CENL1), BRANCHED1 (BRC1), BRC2, and the strigolactone biosynthesis gene MORE AXILLARY BRANCHES1 (MAX1). Below the BMP, AXBs maintained high CENL1 expression at the rib meristem, suggesting that it serves to maintain poise for growth. In support of this, decapitation initiated outgrowth of CENL1-expressing AXBs, but not of dormant AXBs that had switched CENL1 off. This singles out CENL1 as a rib meristem marker for para-dormancy. BRC1 and MAX1 genes, which may counterbalance CENL1, were down-regulated in decapitation-activated AXBs. The results showed that removal of apical dominance shifted AXB gene expression toward that of apices, while developing TBs adopted the expression pattern of para-dormant AXBs. Bud development thus follows a shared developmental pattern at terminal and axillary positions, despite being triggered by short days and apical dominance, respectively.

  7. EEG markers for characterizing anomalous activities of cerebral neurons in NAT (neuronal activity topography) method.

    PubMed

    Musha, Toshimitsu; Matsuzaki, Haruyasu; Kobayashi, Yohei; Okamoto, Yoshiwo; Tanaka, Mieko; Asada, Takashi

    2013-08-01

    A pair of markers, sNAT and vNAT, is derived from the electroencephalogram (EEG) power spectra (PS) recorded for 5 min with 21 electrodes (4-20 Hz) arranged according to the 10-20 standard. These markers form a new diagnosis tool "NAT" aiming at characterizing various brain disorders. Each signal sequence is divided into segments of 0.64 s and its discrete PS consists of eleven frequency components from 4.68 (3 × 1.56) Hz through 20.34 (13 × 1.56) Hz. PS is normalized to its mean and the bias of PS components on each frequency component across the 21 signal channels is reset to zero. The marker sNAT consists of ten frequency components on 21 channels, characterizing neuronal hyperactivity or hypoactivity as compared with NLc (normal controls). The marker vNAT consists of ten ratios between adjacent PS components denoting the over- or undersynchrony of collective neuronal activities as compared with NLc. The likelihood of a test subject to a specified brain disease is defined in terms of the normalized distance to the template NAT state of the disease in the NAT space. Separation of MCI-AD patients (developing AD in 12-18 months) from NLc is made with a false alarm rate of 15%. Locations with neuronal hypoactivity and undersynchrony of AD patients agree with locations of rCBF reduction measured by SPECT. The 2-D diagram composed of the binary likelihoods between ADc and NLc in the two representations of sNAT and vNAT enables tracing the NAT state of a test subject approaching the AD area, and the follow-up of the treatment effects. PMID:23559020

  8. ERBB3 is a marker of a ganglioneuroblastoma/ganglioneuroma-like expression profile in neuroblastic tumours

    PubMed Central

    2013-01-01

    demonstrates the importance of performing unsupervised clustering and subtype discovery of data sets prior to analyses to avoid a mixture of tumour subtypes, which may otherwise give distorted results and lead to incorrect conclusions. The current study identifies ERBB3 as a clear-cut marker of a GNB/GN-like expression profile, and we suggest a 7-gene expression signature (including ERBB3) as a complement to histopathology analysis of neuroblastic tumours. Further studies of ErbB3 and other ErbB family members and their role in neuroblastic differentiation and pathogenesis are warranted. PMID:23835063

  9. The expression profile for the tumour suppressor gene PTEN and associated polymorphic markers

    PubMed Central

    Hamilton, J A; Stewart, L M D; Ajayi, L; Gray, I C; Gray, N E; Roberts, K G; Watson, G J; Kaisary, A V; Snary, D

    2000-01-01

    PTEN, a putative tumour suppressor gene associated with prostate and other cancers, is known to be located within the chromosomal region 10q23.3. Transcription of the PTEN gives rise to multiple mRNA species. Analyses by Northern blots, using cell lines which express PTEN together with cell lines which have lost the PTEN or carry a truncated version of the gene, has allowed us to demonstrate that the pseudogene is not transcribed. In addition, 3′ RACE studies confirmed that the multiple mRNA species arising from the gene probably result from the use of alternative polyadenylation sites. No evidence for tissue- or cell-specific patterns of transcription was found. Analysis by 5′ RACE placed the putative site for the start of transcription around 830 bp upstream of the start codon. A map of the location of the PTEN gene with a series of overlapping YAC, BAC and PACs has been constructed and the relative position of eight microsatellite markers sited. Two known and one novel marker have been positioned within the gene, the others are in flanking regions. The more accurate location of these markers should help in future studies of the extent of gene loss. Several polymorphisms were also identified, all were within introns. Four of the common polymorphisms appear to be linked. In blood, DNA from 200 individuals, including normal, BPH and prostate cancer patients, confirmed this link. Only two samples of 200 did not carry the linked haplotype, both were patients with advanced prostate cancer. It is possible that such rearrangements within PTEN could be evidence of predisposition to prostate cancer in this small number of cases. © 2000 Cancer Research Campaign PMID:10817502

  10. High expression of arachidonate 15-lipoxygenase and proinflammatory markers in human ischemic heart tissue

    SciTech Connect

    Magnusson, Lisa U.; Lundqvist, Annika; Asp, Julia; Synnergren, Jane; Johansson, Cecilia Thalen; Palmqvist, Lars; Jeppsson, Anders; Hulten, Lillemor Mattsson

    2012-07-27

    Highlights: Black-Right-Pointing-Pointer We found a 17-fold upregulation of ALOX15 in the ischemic heart. Black-Right-Pointing-Pointer Incubation of human muscle cells in hypoxia showed a 22-fold upregulation of ALOX15. Black-Right-Pointing-Pointer We observed increased levels of proinflammatory markers in ischemic heart tissue. Black-Right-Pointing-Pointer Suggesting a link between ischemia and inflammation in ischemic heart biopsies. -- Abstract: A common feature of the ischemic heart and atherosclerotic plaques is the presence of hypoxia (insufficient levels of oxygen in the tissue). Hypoxia has pronounced effects on almost every aspect of cell physiology, and the nuclear transcription factor hypoxia inducible factor-1{alpha} (HIF-1{alpha}) regulates adaptive responses to low concentrations of oxygen in mammalian cells. In our recent work, we observed that hypoxia increases the proinflammatory enzyme arachidonate 15-lipoxygenase (ALOX15B) in human carotid plaques. ALOX15 has recently been shown to be present in the human myocardium, but the effect of ischemia on its expression has not been investigated. Here we test the hypothesis that ischemia of the heart leads to increased expression of ALOX15, and found an almost 2-fold increase in HIF-1{alpha} mRNA expression and a 17-fold upregulation of ALOX15 mRNA expression in the ischemic heart biopsies from patients undergoing coronary bypass surgery compared with non ischemic heart tissue. To investigate the effect of low oxygen concentration on ALOX15 we incubated human vascular muscle cells in hypoxia and showed that expression of ALOX15 increased 22-fold compared with cells incubated in normoxic conditions. We also observed increased mRNA levels of proinflammatory markers in ischemic heart tissue compared with non-ischemic controls. In summary, we demonstrate increased ALOX15 in human ischemic heart biopsies. Furthermore we demonstrate that hypoxia increases ALOX15 in human muscle cells. Our results yield

  11. Meninges harbor cells expressing neural precursor markers during development and adulthood.

    PubMed

    Bifari, Francesco; Berton, Valeria; Pino, Annachiara; Kusalo, Marijana; Malpeli, Giorgio; Di Chio, Marzia; Bersan, Emanuela; Amato, Eliana; Scarpa, Aldo; Krampera, Mauro; Fumagalli, Guido; Decimo, Ilaria

    2015-01-01

    Brain and skull developments are tightly synchronized, allowing the cranial bones to dynamically adapt to the brain shape. At the brain-skull interface, meninges produce the trophic signals necessary for normal corticogenesis and bone development. Meninges harbor different cell populations, including cells forming the endosteum of the cranial vault. Recently, we and other groups have described the presence in meninges of a cell population endowed with neural differentiation potential in vitro and, after transplantation, in vivo. However, whether meninges may be a niche for neural progenitor cells during embryonic development and in adulthood remains to be determined. In this work we provide the first description of the distribution of neural precursor markers in rat meninges during development up to adulthood. We conclude that meninges share common properties with the classical neural stem cell niche, as they: (i) are a highly proliferating tissue; (ii) host cells expressing neural precursor markers such as nestin, vimentin, Sox2 and doublecortin; and (iii) are enriched in extracellular matrix components (e.g., fractones) known to bind and concentrate growth factors. This study underlines the importance of meninges as a potential niche for endogenous precursor cells during development and in adulthood.

  12. Connectomic markers of disease expression, genetic risk and resilience in bipolar disorder

    PubMed Central

    Dima, D; Roberts, R E; Frangou, S

    2016-01-01

    Bipolar disorder (BD) is characterized by emotional dysregulation and cognitive deficits associated with abnormal connectivity between subcortical—primarily emotional processing regions—and prefrontal regulatory areas. Given the significant contribution of genetic factors to BD, studies in unaffected first-degree relatives can identify neural mechanisms of genetic risk but also resilience, thus paving the way for preventive interventions. Dynamic causal modeling (DCM) and random-effects Bayesian model selection were used to define and assess connectomic phenotypes linked to facial affect processing and working memory in a demographically matched sample of first-degree relatives carefully selected for resilience (n=25), euthymic patients with BD (n=41) and unrelated healthy controls (n=46). During facial affect processing, patients and relatives showed similarly increased frontolimbic connectivity; resilient relatives, however, evidenced additional adaptive hyperconnectivity within the ventral visual stream. During working memory processing, patients displayed widespread hypoconnectivity within the corresponding network. In contrast, working memory network connectivity in resilient relatives was comparable to that of controls. Our results indicate that frontolimbic dysfunction during affect processing could represent a marker of genetic risk to BD, and diffuse hypoconnectivity within the working memory network a marker of disease expression. The association of hyperconnectivity within the affect-processing network with resilience to BD suggests adaptive plasticity that allows for compensatory changes and encourages further investigation of this phenotype in genetic and early intervention studies. PMID:26731443

  13. Maternal testosterone and placental function: Effect of electroacupuncture on placental expression of angiogenic markers and fetal growth.

    PubMed

    Fornes, Romina; Hu, Min; Maliqueo, Manuel; Kokosar, Milana; Benrick, Anna; Carr, David; Billig, Håkan; Jansson, Thomas; Manni, Luigi; Stener-Victorin, Elisabet

    2016-09-15

    Women with polycystic ovary syndrome (PCOS) have elevated circulating androgens during pregnancy and are at an increased risk of adverse pregnancy outcomes. Here we tested the hypotheses that maternal androgen excess decrease placental and fetal growth, and placental expression of markers of steroidogenesis, angiogenesis and sympathetic activity, and that acupuncture with low-frequency electrical stimulation prevents these changes. Pregnant rats were exposed to vehicle or testosterone on gestational day (GD)15-19. Low-frequency electroacupuncture (EA) or handling, as a control for the EA procedure, was given to control or testosterone exposed dams on GD16-20. On GD21, blood pressure was measured and maternal blood, fetuses and placentas collected. Placental steroid receptor expression and proteins involved in angiogenic, neurotrophic and adrenergic signaling were analyzed. EA did not affect any variables in control rats except maternal serum corticosterone, which was reduced. EA in testosterone exposed dams compared with controls increased systolic pressure by 30%, decreased circulating norepinephrine and corticosterone, fetal and placental weight and placental VEGFR1 and proNGF protein expression, and increased the VEGFA/VEGFR1 ratio, mature NGF (mNGF) and the mNGF/proNGF ratio. In conclusion, low-frequency EA in control animals did not have any negative influence on any of the studied variables. In contrast, EA in pregnant dams exposed to testosterone increased blood pressure and impaired placental growth and function, leading to decreased fetal growth. PMID:27208621

  14. Similarity of markers identified from cancer gene expression studies: observations from GEO.

    PubMed

    Shi, Xingjie; Shen, Shihao; Liu, Jin; Huang, Jian; Zhou, Yong; Ma, Shuangge

    2014-09-01

    Gene expression profiling has been extensively conducted in cancer research. The analysis of multiple independent cancer gene expression datasets may provide additional information and complement single-dataset analysis. In this study, we conduct multi-dataset analysis and are interested in evaluating the similarity of cancer-associated genes identified from different datasets. The first objective of this study is to briefly review some statistical methods that can be used for such evaluation. Both marginal analysis and joint analysis methods are reviewed. The second objective is to apply those methods to 26 Gene Expression Omnibus (GEO) datasets on five types of cancers. Our analysis suggests that for the same cancer, the marker identification results may vary significantly across datasets, and different datasets share few common genes. In addition, datasets on different cancers share few common genes. The shared genetic basis of datasets on the same or different cancers, which has been suggested in the literature, is not observed in the analysis of GEO data.

  15. A rapid, modular and marker-free chloroplast expression system for the green alga Chlamydomonas reinhardtii.

    PubMed

    Bertalan, Ivo; Munder, Matthias C; Weiß, Caroline; Kopf, Judith; Fischer, Dirk; Johanningmeier, Udo

    2015-02-10

    In search of alternative expression platforms heterologous protein production in microalgae has gained increasing importance in the last years. Particularly, the chloroplast of the green alga Chlamydomonas reinhardtii has been adopted to successfully express foreign proteins like vaccines and antibodies. However, when compared with other expression systems, the development of the algal chloroplast to a powerful production platform for recombinant proteins is still in its early stages. In an effort to further improve methods for a reliable and rapid generation of transplastomic Chlamydomonas strains we constructed the key plasmid pMM2 containing the psbA gene and a multiple cloning site for foreign gene insertion. The psbA gene allows a marker-free selection procedure using as a recipient the Fud7 strain of Chlamydomonas, which grows on media containing acetate as a carbon source, but is unable to grow photoautotrophically due to the lack of an intact psbA gene. Biolistic transformation of Fud7 with vectors containing this gene restores photoautotrophic growth and thus permits selection in the light on media without carbon sources and antibiotics. The multiple cloning site with a BsaI recognition sequence allows type IIs restriction enzyme-based modular cloning which rapidly generates new gene constructs without sequences, which could influence the expression and characteristics of the foreign protein. In order to demonstrate the feasibility of this approach, a codon optimized version of the gene for the bacterial protein MPT64 has been integrated into the plastome. Several strains with different promoter/UTR combinations show a stable expression of the HA tagged MPT64 protein in Chlamydomonas chloroplasts.

  16. A rapid, modular and marker-free chloroplast expression system for the green alga Chlamydomonas reinhardtii.

    PubMed

    Bertalan, Ivo; Munder, Matthias C; Weiß, Caroline; Kopf, Judith; Fischer, Dirk; Johanningmeier, Udo

    2015-02-10

    In search of alternative expression platforms heterologous protein production in microalgae has gained increasing importance in the last years. Particularly, the chloroplast of the green alga Chlamydomonas reinhardtii has been adopted to successfully express foreign proteins like vaccines and antibodies. However, when compared with other expression systems, the development of the algal chloroplast to a powerful production platform for recombinant proteins is still in its early stages. In an effort to further improve methods for a reliable and rapid generation of transplastomic Chlamydomonas strains we constructed the key plasmid pMM2 containing the psbA gene and a multiple cloning site for foreign gene insertion. The psbA gene allows a marker-free selection procedure using as a recipient the Fud7 strain of Chlamydomonas, which grows on media containing acetate as a carbon source, but is unable to grow photoautotrophically due to the lack of an intact psbA gene. Biolistic transformation of Fud7 with vectors containing this gene restores photoautotrophic growth and thus permits selection in the light on media without carbon sources and antibiotics. The multiple cloning site with a BsaI recognition sequence allows type IIs restriction enzyme-based modular cloning which rapidly generates new gene constructs without sequences, which could influence the expression and characteristics of the foreign protein. In order to demonstrate the feasibility of this approach, a codon optimized version of the gene for the bacterial protein MPT64 has been integrated into the plastome. Several strains with different promoter/UTR combinations show a stable expression of the HA tagged MPT64 protein in Chlamydomonas chloroplasts. PMID:25554634

  17. Comparative Analysis of Cartilage Marker Gene Expression Patterns during Axolotl and Xenopus Limb Regeneration.

    PubMed

    Mitogawa, Kazumasa; Makanae, Aki; Satoh, Ayano; Satoh, Akira

    2015-01-01

    Axolotls (Ambystoma mexicanum) can completely regenerate lost limbs, whereas Xenopus laevis frogs cannot. During limb regeneration, a blastema is first formed at the amputation plane. It is thought that this regeneration blastema forms a limb by mechanisms similar to those of a developing embryonic limb bud. Furthermore, Xenopus laevis frogs can form a blastema after amputation; however, the blastema results in a terminal cone-shaped cartilaginous structure called a "spike." The causes of this patterning defect in Xenopus frog limb regeneration were explored. We hypothesized that differences in chondrogenesis may underlie the patterning defect. Thus, we focused on chondrogenesis. Chondrogenesis marker genes, type I and type II collagen, were compared in regenerative and nonregenerative environments. There were marked differences between axolotls and Xenopus in the expression pattern of these chondrogenesis-associated genes. The relative deficit in the chondrogenic capacity of Xenopus blastema cells may account for the absence of total limb regenerative capacity.

  18. Development of polymorphic microsatellite markers based on expressed sequence tags in Populus cathayana (Salicaceae).

    PubMed

    Tian, Z Z; Zhang, F Q; Cai, Z Y; Chen, S L

    2016-01-01

    Populus cathayana occupies a large area within the northern, central, and southwestern regions of China, and is considered to be an important reforestation species in western China. In order to investigate the population genetic structure of this species, 10 polymorphic microsatellite loci were identified based on expressed sequence tags from de novo sequencing on the Illumina HiSeq 2000 platform. All microsatellite primers were tested on 48 P. cathayana individuals from four locations on the Qinghai-Tibet Plateau. The observed heterozygosity ranged from 0.000 to 1.000, and the null-allele frequency ranged from 0.000 to 0.904. These microsatellite markers may be a useful tool in genetic studies on P. cathayana and closely related species.

  19. Comparative Analysis of Cartilage Marker Gene Expression Patterns during Axolotl and Xenopus Limb Regeneration.

    PubMed

    Mitogawa, Kazumasa; Makanae, Aki; Satoh, Ayano; Satoh, Akira

    2015-01-01

    Axolotls (Ambystoma mexicanum) can completely regenerate lost limbs, whereas Xenopus laevis frogs cannot. During limb regeneration, a blastema is first formed at the amputation plane. It is thought that this regeneration blastema forms a limb by mechanisms similar to those of a developing embryonic limb bud. Furthermore, Xenopus laevis frogs can form a blastema after amputation; however, the blastema results in a terminal cone-shaped cartilaginous structure called a "spike." The causes of this patterning defect in Xenopus frog limb regeneration were explored. We hypothesized that differences in chondrogenesis may underlie the patterning defect. Thus, we focused on chondrogenesis. Chondrogenesis marker genes, type I and type II collagen, were compared in regenerative and nonregenerative environments. There were marked differences between axolotls and Xenopus in the expression pattern of these chondrogenesis-associated genes. The relative deficit in the chondrogenic capacity of Xenopus blastema cells may account for the absence of total limb regenerative capacity. PMID:26186213

  20. Comparative Analysis of Cartilage Marker Gene Expression Patterns during Axolotl and Xenopus Limb Regeneration

    PubMed Central

    Mitogawa, Kazumasa; Makanae, Aki; Satoh, Ayano; Satoh, Akira

    2015-01-01

    Axolotls (Ambystoma mexicanum) can completely regenerate lost limbs, whereas Xenopus laevis frogs cannot. During limb regeneration, a blastema is first formed at the amputation plane. It is thought that this regeneration blastema forms a limb by mechanisms similar to those of a developing embryonic limb bud. Furthermore, Xenopus laevis frogs can form a blastema after amputation; however, the blastema results in a terminal cone-shaped cartilaginous structure called a “spike.” The causes of this patterning defect in Xenopus frog limb regeneration were explored. We hypothesized that differences in chondrogenesis may underlie the patterning defect. Thus, we focused on chondrogenesis. Chondrogenesis marker genes, type I and type II collagen, were compared in regenerative and nonregenerative environments. There were marked differences between axolotls and Xenopus in the expression pattern of these chondrogenesis-associated genes. The relative deficit in the chondrogenic capacity of Xenopus blastema cells may account for the absence of total limb regenerative capacity. PMID:26186213

  1. Development of polymorphic microsatellite markers based on expressed sequence tags in Populus cathayana (Salicaceae).

    PubMed

    Tian, Z Z; Zhang, F Q; Cai, Z Y; Chen, S L

    2016-01-01

    Populus cathayana occupies a large area within the northern, central, and southwestern regions of China, and is considered to be an important reforestation species in western China. In order to investigate the population genetic structure of this species, 10 polymorphic microsatellite loci were identified based on expressed sequence tags from de novo sequencing on the Illumina HiSeq 2000 platform. All microsatellite primers were tested on 48 P. cathayana individuals from four locations on the Qinghai-Tibet Plateau. The observed heterozygosity ranged from 0.000 to 1.000, and the null-allele frequency ranged from 0.000 to 0.904. These microsatellite markers may be a useful tool in genetic studies on P. cathayana and closely related species. PMID:27525845

  2. Increased mRNA expression of peripheral glial cell markers in bipolar disorder: The effect of long-term lithium treatment.

    PubMed

    Ferensztajn-Rochowiak, Ewa; Tarnowski, Maciej; Samochowiec, Jerzy; Michalak, Michal; Ratajczak, Mariusz Z; Rybakowski, Janusz K

    2016-09-01

    Neuroinflammation, with microglial activation as an important element, plays a role in the pathogenesis of bipolar disorder (BD). Also, in mood disorders, pathological changes have been demonstrated in macroglial cells, such as astrocyctes and oligodendrocytes. Postmortem brain studies of BD patients to assess glial cells, such as astrocytes and oligodendrocytes and their markers such as glial fibrillary acidic protein (GFAP), Olig1 and Olig2, produced controversial results. On the other hand, investigation of these markers in the peripheral blood of such patients has not been performed so far. In this study, we examined the mRNA levels of GFAP, Olig1 and Olig2, in the peripheral blood of three groups: 15 BD subjects with a duration of illness at least 10 years (mean 20±9 years) but never treated with lithium, 15 subjects with BD treated continuously with lithium for 8-40 years (mean 16±8 years), and 15 control subjects. The groups were age-and sex-matched. Expression of mRNA markers was measured by real-time quantitative reverse transcription PCR (RQ-PCR). We observed increased mRNA levels of the Olig1 and Olig 2 glial markers studied in the BD patients not taking lithium, compared with the control subjects and increased mRNA level of GFAP, compared with lithium-treated patients. In the lithium-treated BD patients GFAP and Olig1 expression was at similar levels to that in the control group. However, Olig 2 expression was even higher than in the BD patients not taking lithium. The possible mechanisms concerning the higher expression of peripheral mRNA markers in BD patients may involve ongoing inflammatory process, compensatory mechanisms and regenerative responses. The beneficial effect of lithium may be related to its anti-inflammatory properties.

  3. Androgen Receptor Splice Variants Contribute to Prostate Cancer Aggressiveness through Induction of EMT and Expression of Stem Cell Marker Genes

    PubMed Central

    Kong, Dejuan; Sethi, Seema; Li, Yiwei; Chen, Wei; Sakr, Wael A.; Heath, Elisabeth; Sarkar, Fazlul H.

    2014-01-01

    Background The mechanism(s) by which androgen receptor (AR) splice variants contribute to castration-resistant prostate cancer (CRPC) is still lacking. Methods Expressions of Epithelial-to-Mesenchymal Transition (EMT) and stem cell markers were molecularly tested using prostate cancer (PCa) cells transfected with AR and AR3 (also known as AR-V7) plasmids or siRNA, and also cultured cells under androgen deprivation therapy (ADT) condition. Cell migration, clonogenicity, sphere forming capacity was assessed using PCa cells under all experimental conditions and 3, 3′-diindolylmethane (DIM; BR-DIM) treatment. Human PCa samples from BR-DIM untreated or treated patients were also used for assessing the expression of AR3 and stem cell markers. Results Overexpression of AR led to the induction of EMT phenotype, while overexpression of AR3 not only induced EMT but also led to the expression of stem cell signature genes. More importantly, ADT enhanced the expression of AR and AR3 concomitant with up-regulated expression of EMT and stem cell marker genes. Dihydrotestosterone (DHT) treatment decreased the expression of AR and AR3, and reversed the expression of these EMT and stem cell marker genes. BR-DIM administered to PCa patients prior to radical prostatectomy inhibited the expression of cancer stem cell markers consistent with inhibition of self-renewal of PCa cells after BR-DIM treatment. Conclusion AR variants could contribute to PCa progression through induction of EMT and acquisition of stem cell characteristics, which could be attenuated by BR-DIM, suggesting that BR-DIM could become a promising agent for the prevention of CRPC and/or for the treatment of PCa PMID:25307492

  4. Microparticles reveal cell activation during IVF - a possible early marker of a prothrombotic state during the first trimester.

    PubMed

    Olausson, Nina; Mobarrez, Fariborz; Wallen, Håkan; Westerlund, Eli; Hovatta, Outi; Henriksson, Peter

    2016-08-30

    Cell-derived microparticles (MPs) are known to be elevated in a number of diseases related to arterial and venous thromboembolism (VTE), such as acute myocardial infarction, VTE (deep-vein thrombosis and pulmonary embolism) and peripheral arterial disease. IVF-associated pregnancies have previously been shown to be associated with an increased incidence of VTE, mechanisms behind being unknown and sparsely studied. Our objective was to assess cell activation during IVF through analysis of MP levels and phenotype following ovarian stimulation. Thirty-one women undergoing IVF were included and blood samples were collected at down regulation of oestrogen and at high level stimulation with 10- to 100-fold increased endogenous oestrogen levels. MPs were analysed by flow cytometry and phenotyped according to size and protein expression. We found that overall phosphatidylserine positive platelet-, endothelial- and monocyte-derived MPs significantly increased following ovarian stimulation with increased levels of platelet activation markers CD40 ligand and P-selectin. Furthermore, there was an increase in endothelial-derived MPs exposing activation marker E-selectin and monocyte-derived MPs, while neutrophil-derived MPs decreased slightly. In conclusion we found a major increase in MPs and markers indicating cell activation in parallel with the profound oestrogen boost during IVF. To assess whether these changes in MPs are associated with thromboembolic events requires extended longitudinal studies.

  5. Microparticles reveal cell activation during IVF - a possible early marker of a prothrombotic state during the first trimester.

    PubMed

    Olausson, Nina; Mobarrez, Fariborz; Wallen, Håkan; Westerlund, Eli; Hovatta, Outi; Henriksson, Peter

    2016-08-30

    Cell-derived microparticles (MPs) are known to be elevated in a number of diseases related to arterial and venous thromboembolism (VTE), such as acute myocardial infarction, VTE (deep-vein thrombosis and pulmonary embolism) and peripheral arterial disease. IVF-associated pregnancies have previously been shown to be associated with an increased incidence of VTE, mechanisms behind being unknown and sparsely studied. Our objective was to assess cell activation during IVF through analysis of MP levels and phenotype following ovarian stimulation. Thirty-one women undergoing IVF were included and blood samples were collected at down regulation of oestrogen and at high level stimulation with 10- to 100-fold increased endogenous oestrogen levels. MPs were analysed by flow cytometry and phenotyped according to size and protein expression. We found that overall phosphatidylserine positive platelet-, endothelial- and monocyte-derived MPs significantly increased following ovarian stimulation with increased levels of platelet activation markers CD40 ligand and P-selectin. Furthermore, there was an increase in endothelial-derived MPs exposing activation marker E-selectin and monocyte-derived MPs, while neutrophil-derived MPs decreased slightly. In conclusion we found a major increase in MPs and markers indicating cell activation in parallel with the profound oestrogen boost during IVF. To assess whether these changes in MPs are associated with thromboembolic events requires extended longitudinal studies. PMID:27412479

  6. Expression and Prognostic Significance of a Panel of Tissue Hypoxia Markers in Head-and-Neck Squamous Cell Carcinomas

    SciTech Connect

    Le, Quynh-Thu Kong, Christina; Lavori, Phillip W.; O'Byrne, Ken; Erler, Janine T.; Huang Xin; Chen Yijun; Cao Hongbin; Tibshirani, Robert; Denko, Nic; Giaccia, Amato J.; Koong, Albert C.

    2007-09-01

    Purpose: To investigate the expression pattern of hypoxia-induced proteins identified as being involved in malignant progression of head-and-neck squamous cell carcinoma (HNSCC) and to determine their relationship to tumor pO{sub 2} and prognosis. Methods and Materials: We performed immunohistochemical staining of hypoxia-induced proteins (carbonic anhydrase IX [CA IX], BNIP3L, connective tissue growth factor, osteopontin, ephrin A1, hypoxia inducible gene-2, dihydrofolate reductase, galectin-1, I{kappa}B kinase {beta}, and lysyl oxidase) on tumor tissue arrays of 101 HNSCC patients with pretreatment pO{sub 2} measurements. Analysis of variance and Fisher's exact tests were used to evaluate the relationship between marker expression, tumor pO{sub 2}, and CA IX staining. Cox proportional hazard model and log-rank tests were used to determine the relationship between markers and prognosis. Results: Osteopontin expression correlated with tumor pO{sub 2} (Eppendorf measurements) (p = 0.04). However, there was a strong correlation between lysyl oxidase, ephrin A1, and galectin-1 and CA IX staining. These markers also predicted for cancer-specific survival and overall survival on univariate analysis. A hypoxia score of 0-5 was assigned to each patient, on the basis of the presence of strong staining for these markers, whereby a higher score signifies increased marker expression. On multivariate analysis, increasing hypoxia score was an independent prognostic factor for cancer-specific survival (p = 0.015) and was borderline significant for overall survival (p = 0.057) when adjusted for other independent predictors of outcomes (hemoglobin and age). Conclusions: We identified a panel of hypoxia-related tissue markers that correlates with treatment outcomes in HNSCC. Validation of these markers will be needed to determine their utility in identifying patients for hypoxia-targeted therapy.

  7. Brain Region–Specific Alterations in the Gene Expression of Cytokines, Immune Cell Markers and Cholinergic System Components during Peripheral Endotoxin–Induced Inflammation

    PubMed Central

    Silverman, Harold A; Dancho, Meghan; Regnier-Golanov, Angelique; Nasim, Mansoor; Ochani, Mahendar; Olofsson, Peder S; Ahmed, Mohamed; Miller, Edmund J; Chavan, Sangeeta S; Golanov, Eugene; Metz, Christine N; Tracey, Kevin J; Pavlov, Valentin A

    2014-01-01

    Inflammatory conditions characterized by excessive peripheral immune responses are associated with diverse alterations in brain function, and brain-derived neural pathways regulate peripheral inflammation. Important aspects of this bidirectional peripheral immune–brain communication, including the impact of peripheral inflammation on brain region–specific cytokine responses, and brain cholinergic signaling (which plays a role in controlling peripheral cytokine levels), remain unclear. To provide insight, we studied gene expression of cytokines, immune cell markers and brain cholinergic system components in the cortex, cerebellum, brainstem, hippocampus, hypothalamus, striatum and thalamus in mice after an intraperitoneal lipopolysaccharide injection. Endotoxemia was accompanied by elevated serum levels of interleukin (IL)-1β, IL-6 and other cytokines and brain region–specific increases in Il1b (the highest increase, relative to basal level, was in cortex; the lowest increase was in cerebellum) and Il6 (highest increase in cerebellum; lowest increase in striatum) mRNA expression. Gene expression of brain Gfap (astrocyte marker) was also differentially increased. However, Iba1 (microglia marker) mRNA expression was decreased in the cortex, hippocampus and other brain regions in parallel with morphological changes, indicating microglia activation. Brain choline acetyltransferase (Chat ) mRNA expression was decreased in the striatum, acetylcholinesterase (Ache) mRNA expression was decreased in the cortex and increased in the hippocampus, and M1 muscarinic acetylcholine receptor (Chrm1) mRNA expression was decreased in the cortex and the brainstem. These results reveal a previously unrecognized regional specificity in brain immunoregulatory and cholinergic system gene expression in the context of peripheral inflammation and are of interest for designing future antiinflammatory approaches. PMID:25299421

  8. Selectively active markers for solving of the partial occlusion problem in matchmoving and chromakeying workflow

    NASA Astrophysics Data System (ADS)

    Mazurek, Przemysław

    2013-09-01

    Matchmoving (Match Moving) is the process used for the estimation of camera movements for further integration of acquired video image with computer graphics. The estimation of movements is possible using pattern recognition, 2D and 3D tracking algorithms. The main problem for the workflow is the partial occlusion of markers by the actor, because manual rotoscoping is necessary for fixing of the chroma-keyed footage. In the paper, the partial occlusion problem is solved using the invented, selectively active electronic markers. The sensor network with multiple infrared links detects occlusion state (no-occlusion, partial, full) and switch LED's based markers.

  9. Network activity-independent coordinated gene expression program for synapse assembly

    PubMed Central

    Valor, Luis M.; Charlesworth, Paul; Humphreys, Lawrence; Anderson, Chris N. G.; Grant, Seth G. N.

    2007-01-01

    Global biological datasets generated by genomics, transcriptomics, and proteomics provide new approaches to understanding the relationship between the genome and the synapse. Combined transcriptome analysis and multielectrode recordings of neuronal network activity were used in mouse embryonic primary neuronal cultures to examine synapse formation and activity-dependent gene regulation. Evidence for a coordinated gene expression program for assembly of synapses was observed in the expression of 642 genes encoding postsynaptic and plasticity proteins. This synaptogenesis gene expression program preceded protein expression of synapse markers and onset of spiking activity. Continued expression was followed by maturation of morphology and electrical neuronal networks, which was then followed by the expression of activity-dependent genes. Thus, two distinct sequentially active gene expression programs underlie the genomic programs of synapse function. PMID:17360580

  10. Heterochromatin Protein 1 Binding Protein 3 Expression as a Candidate Marker of Intrinsic 5-Fluorouracil Resistance

    PubMed Central

    HADAC, JAMIE N.; MILLER, DEVON D.; GRIMES, IAN C.; CLIPSON, LINDA; NEWTON, MICHAEL A.; SCHELMAN, WILLIAM R.; HALBERG, RICHARD B.

    2016-01-01

    Background Despite receiving post-operative 5-fluorouracil (5-FU)-based chemotherapy, approximately 50% of patients with stage IIIC colon cancer experience recurrence. Currently, no molecular signature can predict response to 5-FU. Materials and Methods Mouse models of colon cancer have been developed and characterized. Individual tumors in these mice can be longitudinally monitored and assessed to identify differences between those that are responsive and those that are resistant to therapy. Gene expression was analyzed in serial biopsies that were collected before and after treatment with 5-FU. Colon tumors had heterogeneous responses to treatment with 5-FU. Microarray analysis of pretreatment biopsies revealed that Hp1bp3, a gene encoding heterochromatin protein 1 binding protein 3, was differentially expressed between sensitive and resistant tumors. Conclusion Using mouse models of human colorectal cancer, Hp1bp3 was identified as a candidate marker of intrinsic 5-FU resistance and may represent a potential biomarker for patient stratification or a target of clinical importance. PMID:26976970

  11. A Rapid Molecular Test for Determining Yersinia pestis Susceptibility to Ciprofloxacin by the Quantification of Differentially Expressed Marker Genes

    PubMed Central

    Steinberger-Levy, Ida; Shifman, Ohad; Zvi, Anat; Ariel, Naomi; Beth-Din, Adi; Israeli, Ofir; Gur, David; Aftalion, Moshe; Maoz, Sharon; Ber, Raphael

    2016-01-01

    Standard antimicrobial susceptibility tests used to determine bacterial susceptibility to antibiotics are growth dependent and time consuming. The long incubation time required for standard tests may render susceptibility results irrelevant, particularly for patients infected with lethal bacteria that are slow growing on agar but progress rapidly in vivo, such as Yersinia pestis. Here, we present an alternative approach for the rapid determination of antimicrobial susceptibility, based on the quantification of the changes in the expression levels of specific marker genes following exposure to growth-inhibiting concentrations of the antibiotic, using Y. pestis and ciprofloxacin as a model. The marker genes were identified by transcriptomic DNA microarray analysis of the virulent Y. pestis Kimberley53 strain after exposure to specific concentrations of ciprofloxacin for various time periods. We identified several marker genes that were induced following exposure to growth-inhibitory concentrations of ciprofloxacin, and we confirmed the marker expression profiles at additional ciprofloxacin concentrations using quantitative RT-PCR. Eleven candidate marker transcripts were identified, of which four mRNA markers were selected for a rapid quantitative RT-PCR susceptibility test that correctly determined the Minimal Inhibitory Concentration (MIC) values and the categories of susceptibility of several Y. pestis strains and isolates harboring various ciprofloxacin MIC values. The novel molecular susceptibility test requires just 2 h of antibiotic exposure in a 7-h overall test time, in contrast to the 24 h of antibiotic exposure required for a standard microdilution test. PMID:27242774

  12. A Rapid Molecular Test for Determining Yersinia pestis Susceptibility to Ciprofloxacin by the Quantification of Differentially Expressed Marker Genes.

    PubMed

    Steinberger-Levy, Ida; Shifman, Ohad; Zvi, Anat; Ariel, Naomi; Beth-Din, Adi; Israeli, Ofir; Gur, David; Aftalion, Moshe; Maoz, Sharon; Ber, Raphael

    2016-01-01

    Standard antimicrobial susceptibility tests used to determine bacterial susceptibility to antibiotics are growth dependent and time consuming. The long incubation time required for standard tests may render susceptibility results irrelevant, particularly for patients infected with lethal bacteria that are slow growing on agar but progress rapidly in vivo, such as Yersinia pestis. Here, we present an alternative approach for the rapid determination of antimicrobial susceptibility, based on the quantification of the changes in the expression levels of specific marker genes following exposure to growth-inhibiting concentrations of the antibiotic, using Y. pestis and ciprofloxacin as a model. The marker genes were identified by transcriptomic DNA microarray analysis of the virulent Y. pestis Kimberley53 strain after exposure to specific concentrations of ciprofloxacin for various time periods. We identified several marker genes that were induced following exposure to growth-inhibitory concentrations of ciprofloxacin, and we confirmed the marker expression profiles at additional ciprofloxacin concentrations using quantitative RT-PCR. Eleven candidate marker transcripts were identified, of which four mRNA markers were selected for a rapid quantitative RT-PCR susceptibility test that correctly determined the Minimal Inhibitory Concentration (MIC) values and the categories of susceptibility of several Y. pestis strains and isolates harboring various ciprofloxacin MIC values. The novel molecular susceptibility test requires just 2 h of antibiotic exposure in a 7-h overall test time, in contrast to the 24 h of antibiotic exposure required for a standard microdilution test. PMID:27242774

  13. Intersex related gene expression profiles in clams Scrobicularia plana: Molecular markers and environmental application.

    PubMed

    Ciocan, Corina M; Cubero-Leon, Elena; Langston, William J; Pope, Nick; Cornelius, Keith; Hill, E M; Alvarez-Munoz, Diana; Indiveri, Paolo; Lerebours, Adelaide; Minier, Christophe; Rotchell, Jeanette M

    2015-06-30

    Intersex, the appearance of female characteristics in male gonads, has been identified in several aquatic species. It is a widespread phenomenon in populations of the bivalve, Scrobicularia plana, from the southwest coast of the U.K. Genes previously identified as differentially expressed (ferritin, testicular haploid expressed gene, THEG, proliferating cell nuclear antigen, PCNA; receptor activated protein kinase C, RACK; cytochrome B, CYB; and cytochrome c oxidase 1, COX1) in intersex clams relative to normal male clams, were selected for characterisation and an environmental survey of the Channel region. Transcripts were significantly differentially expressed at sites with varying intersex incidence and contaminant burdens. Significant correlations between specific gene expressions, key contaminants and sampling locations have been identified, though no single gene was associated with intersex incidence. The results highlight the difficulty in understanding the intersex phenomenon in molluscs where there is still a lack of knowledge on the control of normal reproduction.

  14. Expression Levels of ALA Dehydratase as a Marker of ALA-PDT Efficacy

    NASA Astrophysics Data System (ADS)

    Avital, Schauder; Tamar, Feuerstein; Zvi, Malik

    2010-05-01

    Accelerated synthesis of protoporphyrinIX (PpIX) following ALA pre-treatment followed by light irradiation is the principle of ALA-PDT. Several limiting enzymes were suggested to control PpIX accumulation and PDT efficacy, among them porphobilinogen deaminase (PBGD) and ferrochelatase. Here we reveal the centrality of ALA dehydratase (ALAD) activity in predicting ALA-PDT efficacy. Silencing of ALAD expression and activity was carried out in leukemic cells using shRNA plasmid transfection or Pb2+ intoxication. ALAD activity, porphyrin synthesis and mitochondrial activity were determined versus PDT efficacy. In K562 ALAD-silenced cells, ALAD activity and expression were reduced and as a result, PpIX synthesis was almost abolished. Following ALA treatment and irradiation, ALAD-silenced cells depicted normal mitochondrial activity, in contrast to control and non-silencing transfected cells where accumulated PpIX and irradiation caused ROS formation and mitochondrial damage. Morphological analysis by scanning electron microscopy (SEM) of ALA-PDT treated cells showed no morphological changes in ALAD-silenced cells, while controls exhibited cell deformations and lysis. Annexin V-FITC/PI staining as well as LDH-L leakage testing showed that membrane integrity was undamaged following ALA-PDT in ALAD silenced cells. Pb2+ treatment in MEL cells impaired ALAD activity and reduced PpIX synthesis but to a lesser extent. In conclusion, we show that a dramatic reduction in PpIX accumulation following down regulation of ALAD expression prevents an efficient PDT. Thus, ALAD has a major role in regulating PpIX synthesis and ALA-PDT therapeutic outcome. Monitoring ALAD expression or activity in various tumors may be useful as prognostic tool to predict PDT efficacy.

  15. APOE genotype alters glial activation and loss of synaptic markers in mice.

    PubMed

    Zhu, Yuangui; Nwabuisi-Heath, Evelyn; Dumanis, Sonya B; Tai, Leon M; Yu, Chunjiang; Rebeck, G William; LaDu, Mary Jo

    2012-04-01

    The ε4 allele of the Apolipoprotein E (APOE) gene is the strongest genetic risk factor for late-onset Alzheimer's disease (AD), and affects clinical outcomes of chronic and acute brain damages. The mechanisms by which apoE affect diverse diseases and disorders may involve modulation of the glial response to various types of brain damage. We examined glial activation in a mouse model where each of the human APOE alleles are expressed under the endogenous mouse APOE promoter, as well as in APOE knock-out mice. APOE4 mice displayed increased glial activation in response to intracerebroventricular lipopolysaccharide (LPS) compared to APOE2 and APOE3 mice by several measures. There were higher levels of microglia/macrophage, astrocytes, and invading T-cells after LPS injection in APOE4 mice. APOE4 mice also displayed greater and more prolonged increases of cytokines (IL-1β, IL-6, TNF-α) than APOE2 and APOE3 mice. We found that APOE4 mice had greater synaptic protein loss after LPS injection, as measured by three markers: PSD-95, drebin, and synaptophysin. In all assays, APOE knock-out mice responded similar to APOE4 mice, suggesting that the apoE4 protein may lack anti-inflammatory characteristics of apoE2 and apoE3. Together, these findings demonstrate that APOE4 predisposes to inflammation, which could contribute to its association with Alzheimer's disease and other disorders.

  16. Development and validation of new SSR markers from expressed regions in the garlic genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Limited number of simple sequence repeat (SSR) markers is available for the genome of garlic (Allium sativum L.) although SSR markers have become one of the most preferred marker systems because they are typically co-dominant, reproducible, cross species transferable and highly polymorphic. In this ...

  17. Left insula activation: a marker for language attainment in bilinguals.

    PubMed

    Chee, Michael W L; Soon, Chun Siong; Lee, Hwee Ling; Pallier, Christophe

    2004-10-19

    Several lines of evidence suggest the importance of phonological working memory (PWM) in language acquisition. We investigated the neural correlates of PWM in young adults who were under compelling social pressure to be bilingual. Equal bilinguals had high proficiency in English and Chinese as measured by a standardized examination, whereas unequal bilinguals were proficient in English but not Chinese. Both groups were matched on several measures of nonverbal intelligence and working memory. In-scanner behavioral results did not show between-group differences. Of the regions showing load-dependent increments in activation, the left insula showed greater activation in equal bilinguals. Unequal bilinguals showed greater task-related deactivation in the anterior medial frontal region and greater anterior cingulate activation. Although unequal bilinguals kept apace with equal bilinguals in the simple PWM task, the differential cortical activations suggest that more optimal engagement of PWM in the latter may correlate with better second-language attainment. PMID:15469927

  18. Left insula activation: A marker for language attainment in bilinguals

    PubMed Central

    Chee, Michael W. L.; Soon, Chun Siong; Lee, Hwee Ling; Pallier, Christophe

    2004-01-01

    Several lines of evidence suggest the importance of phonological working memory (PWM) in language acquisition. We investigated the neural correlates of PWM in young adults who were under compelling social pressure to be bilingual. Equal bilinguals had high proficiency in English and Chinese as measured by a standardized examination, whereas unequal bilinguals were proficient in English but not Chinese. Both groups were matched on several measures of nonverbal intelligence and working memory. In-scanner behavioral results did not show between-group differences. Of the regions showing load-dependent increments in activation, the left insula showed greater activation in equal bilinguals. Unequal bilinguals showed greater task-related deactivation in the anterior medial frontal region and greater anterior cingulate activation. Although unequal bilinguals kept apace with equal bilinguals in the simple PWM task, the differential cortical activations suggest that more optimal engagement of PWM in the latter may correlate with better second-language attainment. PMID:15469927

  19. Left insula activation: a marker for language attainment in bilinguals.

    PubMed

    Chee, Michael W L; Soon, Chun Siong; Lee, Hwee Ling; Pallier, Christophe

    2004-10-19

    Several lines of evidence suggest the importance of phonological working memory (PWM) in language acquisition. We investigated the neural correlates of PWM in young adults who were under compelling social pressure to be bilingual. Equal bilinguals had high proficiency in English and Chinese as measured by a standardized examination, whereas unequal bilinguals were proficient in English but not Chinese. Both groups were matched on several measures of nonverbal intelligence and working memory. In-scanner behavioral results did not show between-group differences. Of the regions showing load-dependent increments in activation, the left insula showed greater activation in equal bilinguals. Unequal bilinguals showed greater task-related deactivation in the anterior medial frontal region and greater anterior cingulate activation. Although unequal bilinguals kept apace with equal bilinguals in the simple PWM task, the differential cortical activations suggest that more optimal engagement of PWM in the latter may correlate with better second-language attainment.

  20. Spatial expression of CLAVATA3 in the shoot apical meristem suggests it is not a stem cell marker in soybean.

    PubMed

    Wong, Chui E; Singh, Mohan B; Bhalla, Prem L

    2013-12-01

    CLAVATA3 (CLV3), a stem cell marker in Arabidopsis thaliana, encodes a secreted peptide that maintains the stem cell population within the shoot apical meristem. This work investigated the CLV3 orthologue in a major legume crop, soybean (GmCLV3). Instead of being expressed in the three outermost layers of the meristem as in Arabidopsis, GmCLV3 was expressed deeper in the central zone beneath the fourth layer (L4) of the meristem, overlapping with the expression of soybean WUSCHEL. Subsequent investigation using an alternative stem cell marker (GmLOG1) revealed its expression within layers L2-L4, indicating that GmCLV3 is not a stem cell marker. Overexpression studies of GmCLV3 in Arabidopsis and complementation of clv3-2 mutant suggest similar functional capacity to that of Arabidopsis CLV3. The expression of soybean CLV1, which encodes a receptor for CLV3 in Arabidopsis, was not detectable in the central zone of the meristem via reverse-transcription PCR analysis of amplified RNA from laser-microdissected samples or in situ, implicating a diverged pathway in soybean. This study also reports the novel expression of GmLOG1 in initials of axillary meristem in the boundary region between the SAM and developing leaf primordia, before the expression of GmWUS or GmCLV3, indicating cytokinin as one of the earliest signals in initiating and specifying the stem cell population.

  1. Spatial expression of CLAVATA3 in the shoot apical meristem suggests it is not a stem cell marker in soybean.

    PubMed

    Wong, Chui E; Singh, Mohan B; Bhalla, Prem L

    2013-12-01

    CLAVATA3 (CLV3), a stem cell marker in Arabidopsis thaliana, encodes a secreted peptide that maintains the stem cell population within the shoot apical meristem. This work investigated the CLV3 orthologue in a major legume crop, soybean (GmCLV3). Instead of being expressed in the three outermost layers of the meristem as in Arabidopsis, GmCLV3 was expressed deeper in the central zone beneath the fourth layer (L4) of the meristem, overlapping with the expression of soybean WUSCHEL. Subsequent investigation using an alternative stem cell marker (GmLOG1) revealed its expression within layers L2-L4, indicating that GmCLV3 is not a stem cell marker. Overexpression studies of GmCLV3 in Arabidopsis and complementation of clv3-2 mutant suggest similar functional capacity to that of Arabidopsis CLV3. The expression of soybean CLV1, which encodes a receptor for CLV3 in Arabidopsis, was not detectable in the central zone of the meristem via reverse-transcription PCR analysis of amplified RNA from laser-microdissected samples or in situ, implicating a diverged pathway in soybean. This study also reports the novel expression of GmLOG1 in initials of axillary meristem in the boundary region between the SAM and developing leaf primordia, before the expression of GmWUS or GmCLV3, indicating cytokinin as one of the earliest signals in initiating and specifying the stem cell population. PMID:24179098

  2. [Expression of CD48 as a live marker to distinguish division of hematopoietic stem cells].

    PubMed

    Yang, Xin; Zhang, Yu; Peng, Lu-Yun; Pang, Ya-Kun; Dong, Fang; Ji, Qing; Xu, Jing; Cheng, Tao; Yuan, Wei-Ping; Gao, Ying-Dai

    2014-06-01

    AF488(+) cells was significantly higher than that of AF488(-) cells (P < 0.05). The proliferation ability of AF488(-) cells was also significantly higher than AF488(+) cells (P < 0.05). It is concluded that the expression of CD48 can distinguish cell division of hematopoietic stem cells and can be used as a live marker for the loss of stemness. In comparison with the Numb protein staining, this method can be used in living cells, thus provides greater convenience for subsequent cell culture studies and cell transplantation experiments.

  3. Evaluation of fascin-1 expression as a marker of invasion in urothelial carcinomas

    PubMed Central

    Sharma, Arun; Badwal, Sonia; Dutta, Vibha; Basu, Atoshi

    2014-01-01

    Background Prognostication and therapeutic evaluation of urothelial carcinomas significantly depends on the depth of invasion. The assessment of invasion on routine histopathological sections may be difficult in some cases. Fascin is an actin-bundling protein involved in tumor cell migration with enhanced expression associated with invasive tumors. The data available on fascin-1 expression in urothelial carcinoma however is limited. To characterize fascin-1 expression in urothelial neoplasms and its correlation with invasiveness in urothelial carcinomas. Methods A descriptive study design wherein fascin-1 immunoreactivity was studied in 126 urothelial neoplasms using monoclonal antibody against fascin by immunohistochemistry. 52/126 (41.26%) were low grade carcinomas (48/52 stage pTa and 4/52 stage pT1), 46/126 (36.5%) high grade carcinomas (13/46 stage pTa, 8/46 stage pT1 and 25/46 stage pT2), 02/126 carcinoma-in-situ, 03/126 papilloma, 12/126 papillary urothelial neoplasm of uncertain malignant potential and 11/126 were other variants of urothelial carcinomas. Fascin-1 cytoplasmic immunoreactivity was assessed semiquantitatively in terms of extent, intensity and a combined immunoreactivity score. Correlation between immunoreactivity scores and invasiveness was evaluated using Pearson's chi-square (χ2) and Nonparametric Spearman rho (ρ) correlation coefficient two tailed. Results The scores for intensity, extent and combined immunoreactivity were significantly higher in invasive carcinomas. In addition, strong staining was observed exclusively in invasive carcinomas. None of the pTa tumors demonstrated intense staining, including those categorized as high grade carcinomas. Conclusion Fascin-1 overexpression may be used as a marker in urothelial carcinomas where it is morphologically difficult to determine the status of invasion. PMID:24843202

  4. HER3 Expression Is a Marker of Tumor Progression in Premalignant Lesions of the Gastroesophageal Junction

    PubMed Central

    Zhang, Paul J.; Furth, Emma E.; Ginsberg, Gregory G.; McMillan, Matthew T.; Datta, Jashodeep; Czerniecki, Brian J.; Roses, Robert E.

    2016-01-01

    Overexpression of receptor tyrosine kinases (RTK), including members of the HER family, has prognostic and therapeutic significance in invasive esophagogastric carcinoma. RTK expression in premalignant gastroesophageal lesions has not been extensively explored. Formalin-fixed paraffin-embedded tissue samples of esophageal biopsy specimens from 73 patients with Barrett’s esophagus with either low-grade dysplasia (LGD) (n = 32) or high-grade dysplasia (HGD) (n = 59) were analyzed for HER1, HER2, HER3 and CMET expression by immunohistochemistry (IHC). Immunophenotype was correlated with histologic and clinical features. High-grade dysplasia (HGD) was associated with overexpression of HER1 (20.7% vs. 3.1%, p = 0.023), HER2 (5.3% vs. 0.0%, p = 0.187) and HER3 (47.4% vs. 9.4%, p<0.001) compared to low-grade dysplasia (LGD). There was a significant association of HER2 (20.0% vs. 2.1%, p = 0.022) and HER3 (80.0% vs. 40.4%, p = 0.023) overexpression in HGD lesions associated with foci of invasive carcinoma compared to those without invasive foci. Overexpression of CMET was observed in 42.9% of specimens, was increasingly observed with HGD compared to LGD (58.3% vs. 36.7%, p = 0.200), and was most often co-expressed with HER3 (62.5% of HER3-positive specimens vs. 38.2% of HER3-negative specimens, p = 0.212). In summary, HER3 is frequently overexpressed in high-grade dysplastic lesions of the gastroesophageal junction and may be a marker of invasive progression. These data provide rationale for targeting HER2 and HER3 pathways in an early disease setting to prevent disease progression. PMID:27559738

  5. Hic-5 Regulates Actin Cytoskeletal Reorganization and Expression of Fibrogenic Markers and Myocilin in Trabecular Meshwork Cells

    PubMed Central

    Pattabiraman, Padmanabhan Paranji; Rao, Ponugoti Vasantha

    2015-01-01

    Purpose To explore the role of inducible focal adhesion (FA) protein Hic-5 in actin cytoskeletal reorganization, FA formation, fibrogenic activity, and expression of myocilin in trabecular meshwork (TM) cells. Methods Using primary cultures of human TM (HTM) cells, the effects of various external factors on Hic-5 protein levels, as well as the effects of recombinant Hic-5 and Hic-5 small interfering RNA (siRNA) on actin cytoskeleton, FAs, myocilin, α-smooth muscle actin (αSMA), and collagen-1 were determined by immunofluorescence and immunoblot analyses. Results Hic-5 distributes discretely to the FAs in HTM cells and throughout the TM and Schlemm's canal of the human aqueous humor (AH) outflow pathway. Transforming growth factor-β2 (TGF-β2), endothelin-1, lysophosphatidic acid, hydrogen peroxide, and RhoA significantly increased Hic-5 protein levels in HTM cells in association with reorganization of actin cytoskeleton and FAs. While recombinant Hic-5 induced actin stress fibers, FAs, αv integrin redistribution to the FAs, increased levels of αSMA, collagen-1, and myocilin, Hic-5 siRNA suppressed most of these responses in HTM cells. Hic-5 siRNA also suppressed TGF-β2-induced fibrogenic activity and dexamethasone-induced myocilin expression in HTM cells. Conclusions Taken together, these results reveal that Hic-5, whose levels were increased by various external factors implicated in elevated intraocular pressure, induces actin cytoskeletal reorganization, FAs, expression of fibrogenic markers, and myocilin in HTM cells. These characteristics of Hic-5 in TM cells indicate its importance in regulation of AH outflow through the TM in both normal and glaucomatous eyes. PMID:26313302

  6. Association between the chondrocyte phenotype and the expression of adipokines and their receptors: evidence for a role of leptin but not adiponectin in the expression of cartilage-specific markers.

    PubMed

    Francin, Pierre-Jean; Guillaume, Cécile; Humbert, Anne-Claude; Pottie, Pascale; Netter, Patrick; Mainard, Didier; Presle, Nathalie

    2011-11-01

    Although extensive evidence support the key role of adipokines in cartilage homeostasis, contradictory data have been found for their expression and their effects in chondrocytes. This study was then undertaken to determine whether a phenotypic modulation may affect the expression of adipokines and their receptors in human chondrocytes. The expression of leptin, adiponectin and their receptors, as well as cartilage-specific genes was examined in chondrocytes obtained from patients with osteoarthritis either directly after cells harvest or after culture in monolayer or in alginate beads. The results showed major changes in the gene expression pattern after culture in monolayer with a shift from the adipokines to their receptors. Interestingly, this downregulation of adipokines was associated with a loss of chondrocyte phenotype, and chondrocytes recovered a cartilage-like expression profile of leptin and adiponectin when cultured in a tridimensional chondrocyte phenotype-inducing system, but ceased expressing their receptors. Further experiments clearly showed that leptin but not adiponectin promoted the expression of cartilage-specific markers through mitogen-activated protein kinase, Janus kinase and phosphatidylinositol-3 kinase signaling pathways. In conclusion, our data indicate that any phenotypic modulation could affect chondrocyte responsiveness to leptin or adiponectin, and provide evidence for an important role for leptin in regulating the expression of cartilage-specific markers.

  7. Gene expression markers in circulating tumor cells may predict bone metastasis and response to hormonal treatment in breast cancer.

    PubMed

    Wang, Haiying; Molina, Julian; Jiang, John; Ferber, Matthew; Pruthi, Sandhya; Jatkoe, Timothy; Derecho, Carlo; Rajpurohit, Yashoda; Zheng, Jian; Wang, Yixin

    2013-11-01

    Circulating tumor cells (CTCs) have recently attracted attention due to their potential as prognostic and predictive markers for the clinical management of metastatic breast cancer patients. The isolation of CTCs from patients may enable the molecular characterization of these cells, which may help establish a minimally invasive assay for the prediction of metastasis and further optimization of treatment. Molecular markers of proven clinical value may therefore be useful in predicting disease aggressiveness and response to treatment. In our earlier study, we identified a gene signature in breast cancer that appears to be significantly associated with bone metastasis. Among the genes that constitute this signature, trefoil factor 1 (TFF1) was identified as the most differentially expressed gene associated with bone metastasis. In this study, we investigated 25 candidate gene markers in the CTCs of metastatic breast cancer patients with different metastatic sites. The panel of the 25 markers was investigated in 80 baseline samples (first blood draw of CTCs) and 30 follow-up samples. In addition, 40 healthy blood donors (HBDs) were analyzed as controls. The assay was performed using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) with RNA extracted from CTCs captured by the CellSearch system. Our study indicated that 12 of the genes were uniquely expressed in CTCs and 10 were highly expressed in the CTCs obtained from patients compared to those obtained from HBDs. Among these genes, the expression of keratin 19 was highly correlated with the CTC count. The TFF1 expression in CTCs was a strong predictor of bone metastasis and the patients with a high expression of estrogen receptor β in CTCs exhibited a better response to hormonal treatment. Molecular characterization of these genes in CTCs may provide a better understanding of the mechanism underlying tumor metastasis and identify gene markers in CTCs for predicting disease progression and

  8. Gene expression markers in circulating tumor cells may predict bone metastasis and response to hormonal treatment in breast cancer

    PubMed Central

    WANG, HAIYING; MOLINA, JULIAN; JIANG, JOHN; FERBER, MATTHEW; PRUTHI, SANDHYA; JATKOE, TIMOTHY; DERECHO, CARLO; RAJPUROHIT, YASHODA; ZHENG, JIAN; WANG, YIXIN

    2013-01-01

    Circulating tumor cells (CTCs) have recently attracted attention due to their potential as prognostic and predictive markers for the clinical management of metastatic breast cancer patients. The isolation of CTCs from patients may enable the molecular characterization of these cells, which may help establish a minimally invasive assay for the prediction of metastasis and further optimization of treatment. Molecular markers of proven clinical value may therefore be useful in predicting disease aggressiveness and response to treatment. In our earlier study, we identified a gene signature in breast cancer that appears to be significantly associated with bone metastasis. Among the genes that constitute this signature, trefoil factor 1 (TFF1) was identified as the most differentially expressed gene associated with bone metastasis. In this study, we investigated 25 candidate gene markers in the CTCs of metastatic breast cancer patients with different metastatic sites. The panel of the 25 markers was investigated in 80 baseline samples (first blood draw of CTCs) and 30 follow-up samples. In addition, 40 healthy blood donors (HBDs) were analyzed as controls. The assay was performed using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) with RNA extracted from CTCs captured by the CellSearch system. Our study indicated that 12 of the genes were uniquely expressed in CTCs and 10 were highly expressed in the CTCs obtained from patients compared to those obtained from HBDs. Among these genes, the expression of keratin 19 was highly correlated with the CTC count. The TFF1 expression in CTCs was a strong predictor of bone metastasis and the patients with a high expression of estrogen receptor β in CTCs exhibited a better response to hormonal treatment. Molecular characterization of these genes in CTCs may provide a better understanding of the mechanism underlying tumor metastasis and identify gene markers in CTCs for predicting disease progression and

  9. Up-regulation of stem cell markers by P21-activated kinase 1 contributes to 5-fluorouracil resistance of colorectal cancer.

    PubMed

    Huynh, Nhi; Shulkes, Arthur; Baldwin, Graham; He, Hong

    2016-08-01

    Cancer stem cells (CSC) are tumorigenic and resistant to chemotherapy. In colorectal cancer (CRC), CSCs have been identified by the expression of specific markers, including CD44, Bmi1 and Nanog. Although p21-activated kinase 1 (PAK1), acting downstream of Ras, stimulates Wnt/β-catenin signaling and is known to play an important role in CRC development and progression, the role of PAK1 in the expression of CSC markers has not previously been investigated. The effect of PAK1 over-expression, knockdown or inhibition on the expression or alteration (in the case of CD44) of CSC markers in human CRC cell lines was measured by immunofluorescence and Western blotting. The effect of PAK1 modulation on tumorigenesis, and on resistance to treatment with 5-fluorouracil (5-FU), was measured by sphere formation in vitro and by growth of xenografted tumors in vivo. The results show that PAK1 activity correlated with the expression of CSC markers and the CD44 isoform profile, and with tumor growth both in vitro and in vivo. Furthermore PAK overexpression partially overcame the inhibition of CRC growth by 5-FU, and PAK inhibition was synergistic with 5-FU treatment. Our findings lay the foundation for a combination therapy in which PAK1 inhibitors targeting CSCs may be combined with conventional 5-FU-based chemotherapy for the treatment of CRC.

  10. Phytochelatin synthase activity as a marker of metal pollution.

    PubMed

    Zitka, Ondrej; Krystofova, Olga; Sobrova, Pavlina; Adam, Vojtech; Zehnalek, Josef; Beklova, Miroslava; Kizek, Rene

    2011-08-30

    The synthesis of phytochelatins is catalyzed by γ-Glu-Cys dipeptidyl transpeptidase called phytochelatin synthase (PCS). Aim of this study was to suggest a new tool for determination of phytochelatin synthase activity in the tobacco BY-2 cells treated with different concentrations of the Cd(II). After the optimization steps, an experiment on BY-2 cells exposed to different concentrations of Cd(NO(3))(2) for 3 days was performed. At the end of the experiment, cells were harvested and homogenized. Reduced glutathione and cadmium (II) ions were added to the cell suspension supernatant. These mixtures were incubated at 35°C for 30min and analysed using high performance liquid chromatography coupled with electrochemical detector (HPLC-ED). The results revealed that PCS activity rises markedly with increasing concentration of cadmium (II) ions. The lowest concentration of the toxic metal ions caused almost three fold increase in PCS activity as compared to control samples. The activity of PCS (270fkat) in treated cells was more than seven times higher in comparison to control ones. K(m) for PCS was estimated as 2.3mM. PMID:21715087

  11. Platelet leukocyte aggregates and markers of platelet aggregation, immune activation and disease progression in HIV infected treatment naive asymptomatic individuals.

    PubMed

    Nkambule, Bongani B; Davison, Glenda; Ipp, Hayley

    2015-11-01

    Platelet aggregates play a crucial role in the immune defence mechanism against viruses. Increased levels of lipopolysaccharide have been reported in human immunodeficiency virus (HIV) infected individuals. Platelets are capable of interacting with bacterial LPS and subsequently forming platelet leukocyte aggregates (PLAs). This study aimed at determining the levels of circulating PLAs in treatment naïve HIV infected individuals and correlating them, with markers of immune activation, disease progression and platelet aggregation. Thirty-two HIV negative and 35 HIV positive individuals were recruited from a clinic in the Western Cape. Platelet monocyte and platelet neutrophil aggregates were measured using flow cytometry at baseline and were correlated with markers of platelet activation (CD62P); aggregation (CD36); monocyte and neutrophil activation (CD69); monocyte tissue factor expression (CD142); immune activation (CD38 on T+ cells); D-dimers (a marker of active coagulation); CD4 count and viral load. Platelet monocyte aggregates were also measured post stimulation with lipopolysaccharide. PMA levels were higher in HIV 25.26 (16.16-32.28) versus control 14.12 (8.36-18.83), p = 0.0001. PMAs correlated with %CD38/8 expression (r = 0.54624, p = 0.0155); CD4 count (r = -0.6964, p = 0.0039) viral load (r = 0.633, p < 0.009) and monocyte %CD69 expression (r = 0.757, p = 0.030). In addition the %PMAs correlated with platelet %CD36 (r = 0.606, p = 0.017). The HIV group showed increased levels of %CD62P 5.44 (2.72-11.87) versus control 1.15 (0.19-3.59), p < 0.0001; %CD36 22.53 (10.59-55.15) versus 11.01 (3.69-26.98), p = 0.0312 and tissue factor (CD142) MFI 4.84 (4.01-8.17) versus 1.74 (1.07-9.3), p = 0.0240. We describe increased levels of circulating PMAs which directly correlates with markers of immune activation, disease progression and platelet aggregation in HIV treatment naïve individuals.

  12. Developing expressed sequence tag libraries and the discovery of simple sequence repeat markers for two species of raspberry (Rubus L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: Due to a relatively high level of codominant inheritance and transferability within and among taxonomic groups, simple sequence repeat (SSR) markers are important elements in comparative mapping and delineation of genomic regions associated with traits of economic importance. Expressed S...

  13. Evaluation of anonymous and expressed sequence tag derived polymorphic microsatellite markers in the tobacco budworm Heliothis virescens (Lepidoptera: noctuidae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polymorphic genetic markers were identified and characterized using a partial genomic library of Heliothis virescens enriched for simple sequence repeats (SSR) and nucleotide sequences of expressed sequence tags (EST). Nucleotide sequences of 192 clones from the partial genomic library yielded 147 u...

  14. Molecular Marker Expression Is Highly Heterogeneous in Esophageal Adenocarcinoma and Does Not Predict a Response to Neoadjuvant Therapy.

    PubMed

    Bronson, Nathan W; Diggs, Brian S; Bakis, Gene; Gatter, Kenneth M; Sheppard, Brett C; Hunter, John G; Dolan, James P

    2015-12-01

    A reliable method to identify pathologic complete responders (pCR) or non-responders (NR) to neoadjuvant chemoradiation therapy (NAT) would dramatically improve therapy for esophageal cancer. The purpose of this study is to investigate if a distinct profile of prognostic molecular markers can predict pCR after neoadjuvant therapy. Expression of p53, Her-2/neu, Cox-2, Beta-catenin, E-cadherin, MMP-1, NFkB, and TGF-B was measured by immunohistochemistry in pre-treatment biopsy tissue and graded by an experienced pathologist. A pCR was defined as no evidence of malignancy on final pathology. Molecular profiles comparing responders to non-responders were analyzed using classification and regression tree analysis to investigate response to NAT and overall survival. Nineteen patients were pCRs and 34 were NRs. pCRs were more likely to be alive at follow-up than NRs (p < 0.01). Thirty-seven distinct profiles were identified. Expression of molecular markers was highly heterogeneous between patients and did not correlate with a response to NAT, survival (p = 0.47) or clinical stage (p = 0.39) when evaluated either as individual markers or in combination with other expression patterns. NAT dramatically impacts survival through a mechanism independent of known molecular markers of esophageal cancer, which are expressed in a highly heterogeneous fashion and do not predict response to NAT or survival. PMID:26394876

  15. FAS ligand expression in inflammatory infiltrate lymphoid cells as a prognostic marker in oral squamous cell carcinoma.

    PubMed

    Peterle, G T; Santos, M; Mendes, S O; Carvalho-Neto, P B; Maia, L L; Stur, E; Agostini, L P; Silva, C V M; Trivilin, L O; Nunes, F D; Carvalho, M B; Tajara, E H; Louro, I D; Silva-Conforti, A M A

    2015-09-22

    Currently, the most important prognostic factor in oral squamous cell carcinoma (OSCC) is the presence of regional lymph node metastases, which correlates with a 50% reduction in life expectancy. We have previously observed that expression of hypoxia genes in the tumor inflammatory infiltrate is statistically related to prognosis in OSCC. FAS and FASL expression levels in OSCC have previously been related to patient survival. The present study analyzed the relationship between FASL expression in the inflammatory infiltrate lymphoid cells and clinical variables, tumor histology, and prognosis of OSCC. Strong FASL expression was significantly associated with lymph node metastases (P = 0.035) and disease-specific death (P = 0.014), but multivariate analysis did not confirm FASL expression as an independent death risk factor (OR = 2.78, 95%CI = 0.81-9.55). Disease-free and disease-specific survival were significantly correlated with FASL expression (P = 0.016 and P = 0.005, respectively). Multivariate analysis revealed that strong FASL expression is an independent marker for earlier disease relapse and disease-specific death, with approximately 2.5-fold increased risk compared with weak expression (HR = 2.24, 95%CI = 1.08-4.65 and HR = 2.49, 95%CI = 1.04-5.99, respectively). Our results suggest a potential role for this expression profile as a tumor prognostic marker in OSCC patients.

  16. Bacopa monniera (CDRI-08) Upregulates the Expression of Neuronal and Glial Plasticity Markers in the Brain of Scopolamine Induced Amnesic Mice

    PubMed Central

    Konar, Arpita; Gautam, Akash; Thakur, M. K.

    2015-01-01

    Preclinical studies on animal models have discerned the antiamnesic and memory-enhancing potential of Bacopa monniera (Brahmi) crude extract and standardized extracts. These studies primarily focus on behavioral consequences. However, lack of information on molecular underpinnings has limited the clinical trials of the potent herb in human subjects. In recent years, researchers highlight plasticity markers as molecular correlates of amnesia and being crucial to design therapeutic targets. In the present report, we have investigated the effect of a special extract of B. monniera (CDRI-08) on the expression of key neuronal (BDNF and Arc) and glial (GFAP) plasticity markers in the cerebrum of scopolamine induced amnesic mice. Pre- and postadministration of CDRI-08 ameliorated amnesic effect of scopolamine by decreasing acetyl cholinesterase activity and drastically upregulating the mRNA and protein expression of BDNF, Arc, and GFAP in mouse cerebrum. Interestingly, the plant extract per se elevated BDNF and Arc expression as compared to control but GFAP was unaltered. In conclusion, our findings provide the first molecular evidence for antiamnesic potential of CDRI-08 via enhancement of both neuronal and glial plasticity markers. Further investigations on detailed molecular pathways would encourage therapeutic application of the extract in memory disorders. PMID:26413129

  17. Endoglin regulates cyclooxygenase-2 expression and activity.

    PubMed

    Jerkic, Mirjana; Rivas-Elena, Juan V; Santibanez, Juan F; Prieto, Marta; Rodríguez-Barbero, Alicia; Perez-Barriocanal, Fernando; Pericacho, Miguel; Arévalo, Miguel; Vary, Calvin P H; Letarte, Michelle; Bernabeu, Carmelo; López-Novoa, Jose M

    2006-08-01

    The endoglin heterozygous (Eng(+/-)) mouse, which serves as a model of hereditary hemorrhagic telangiectasia (HHT), was shown to express reduced levels of endothelial NO synthase (eNOS) with impaired activity. Because of intricate changes in vasomotor function in the Eng(+/-) mice and the potential interactions between the NO- and prostaglandin-producing pathways, we assessed the expression and function of cyclooxygenase (COX) isoforms. A specific upregulation of COX-2 in the vascular endothelium and increased urinary excretion of prostaglandin E(2) were observed in the Eng(+/-) mice. Specific COX-2 inhibition with parecoxib transiently increased arterial pressure in Eng(+/-) but not in Eng(+/+) mice. Transfection of endoglin in L6E9 myoblasts, shown previously to stimulate eNOS expression, led to downregulation of COX-2 with no change in COX-1. In addition, COX-2 promoter activity and protein levels were inversely correlated with endoglin levels, in doxycyclin-inducible endothelial cells. Chronic NO synthesis inhibition with N(omega)-nitro-l-arginine methyl ester induced a marked increase in COX-2 only in the normal Eng(+/+) mice. N(omega)-nitro-l-arginine methyl ester also increased COX-2 expression and promoter activity in doxycyclin-inducible endoglin expressing endothelial cells, but not in control cells. The level of COX-2 expression following transforming growth factor-beta1 treatment was less in endoglin than in mock transfected L6E9 myoblasts and was higher in human endothelial cells silenced for endoglin expression. Our results indicate that endoglin is involved in the regulation of COX-2 activity. Furthermore, reduced endoglin levels and associated impaired NO production may be responsible, at least in part, for augmented COX-2 expression and activity in the Eng(+/-) mice. PMID:16840721

  18. Malignant Glioma with Neuronal Marker Expression : A Clinicopathological Study of 18 Cases

    PubMed Central

    Kim, Hong Rye; Lee, Jae Jun; Lee, Jung-Il; Nam, Do Hyun; Suh, Yeon-Lim

    2016-01-01

    Objective Malignant gliomas with neuronal marker expression (MGwNM) are rare and poorly characterized. Increasingly diverse types of MGwNM have been described and these reported cases underscore the dilemmas in the classification and diagnosis of those tumors. The aim of this study is to provide additional insights into MGwNM and present the clinicopathological features of 18 patients. Methods We reviewed the medical records of 18 patients diagnosed as MGwNM at our institute between January 2006 and December 2012. Macroscopic total resection was performed in 11 patients (61%). We evaluated the methylation status of O6-methylguanine-DNA methyltransferase (MGMT) and expression of isocitrate dehydrogenase 1 (IDH-1) in all cases, and deletions of 1p and 19q in available cases. Results The estimated median overall survival was 21.2 months. The median progression-free survival was 6.3 months. Six patients (33%) had MGMT methylation but IDH1 mutation was found in only one patient (6%). Gene analysis for 1p19q performed in nine patients revealed no deletion in six, 19q deletion only in two, and 1p deletion only in one. The extent of resection was significantly correlated with progression free survival on both univariate analysis and multivariate analysis (p=0.002 and p=0.013, respectively). Conclusion In this study, the overall survival of MGwNM was not superior to glioblastoma. The extent of resection has a significant prognostic impact on progression-free survival. Further studies of the prognostic factors related to chemo-radio therapy, similar to studies with glioblastoma, are mandatory to improve survival. PMID:26885285

  19. Sleep Loss Activates Cellular Markers of Inflammation: Sex Differences

    PubMed Central

    Irwin, Michael R.; Carrillo, Carmen; Olmstead, Richard

    2009-01-01

    Sleep disturbance is associated with inflammation and related disorders including cardiovascular disease, arthritis, and diabetes mellitus. Given sex differences in the prevalence of inflammatory disorders with stronger associations in females, this study was undertaken to test the effects of sleep loss on cellular mechanisms that contribute to proinflammatory cytokine activity. In 26 healthy adults (11 females; 15 males), monocyte intracellular proinflammatory cytokine production was repeatedly assessed at 08:00, 12:00, 16:00, 20:00, and 23:00 h during a baseline period and after partial sleep deprivation (awake from 11 PM to 3 AM). In the morning after a night of sleep loss, monocyte production of interleukin 6 and tumor necrosis factor- α differentially changed between the two sexes. Whereas both females and males showed a marked increase in the lipopolysaccharide (LPS) - stimulated production of IL-6 and TNF-α in the morning immediately after PSD, production of these cytokines during the early- and late evening was increased in the females as compared to decreases in the males. Sleep loss induces a functional alteration of monocyte proinflammatory cytokine responses with females showing greater cellular immune activation as compared to changes in males. These results have implications for understanding the role of sleep disturbance in the differential risk profile for inflammatory disorders between the sexes. PMID:19520155

  20. In Silico RAPD Priming Sites in Expressed Sequences and iSCAR Markers for Oil Palm

    PubMed Central

    Premkrishnan, Balakrishnan Vasanthakumari; Arunachalam, Vadivel

    2012-01-01

    RAPD is a simple dominant marker system widely used in biology. Effectiveness of RAPD can be improved by selecting and redesigning primers whose priming sites occur in target sequence(s) of gene or organism at optimum distance. We developed software that uses sequences of random decamer primers and nucleotide sequence(s) as two input files. It locates the priming sites in input sequences and generates output files listing frequency and distance between priming sites. When the priming sites of a single primer occur more than once in a sequence with a distance of 200 to 2000 bp, the software also designs pairs of iSCAR primers. An input of 387 RAPD primers and 42,432 expressed sequences of oil palm are used as test. Wet-lab PCR results from a publication that used the same set of primers were compared with software output on priming sites. In the test sequences of oil palm covering 1.4% of genome, we found that at least 60% the primers chosen using software are sure of giving PCR amplification. We designed 641 iSCAR primers suitable for amplification of oil palm DNA. The software successfully predicted 92% (67 out of 73) of published polymorphic RAPD primers in oil palm. PMID:22474414

  1. Mining of SSR markers from Expressed Sequence Tags of bamboo species

    PubMed Central

    Ramalakshmi, Oviya Iyappan; Piramanayagam, Shanmughavel

    2010-01-01

    With the ever increasing number of Expressed Sequence Tags (ESTs) from various sequencing projects, ESTs have become valuable and first-hand source of in-silico mining of simple sequence repeats (SSR) markers. We examined a total of 3419 EST sequences from three bamboo species, namely, Phyllostachys edulis, Bambusa oldhamii and Dendrocalamus sinicus for the presence of di- to hexa- microsatellites. The frequency of SSR containing ESTs varied from 5.36% in B. oldhamii to 13.05% in P. edulis. No SSRs were found in D. sinicus. Tri-nucleotide repeats (49.34%) were most frequent in P. edulis, while not much comparable difference in repeats was found in B. oldhamii. Flanking primer pairs were also designed in-silico for the sequences containing SSRs and their position on the genome hypothesized using similarity searching. SSRs located in open reading frame (ORF) were given functional annotation using Gene Ontology. Polymorphic SSRs were also detected using new pipeline- polySSR. Polymorphism level was very low (2.43%) and the position of the polymorphic SSRs was determined. The development of SSRs and the study of polymorphism will help in the further study of intra- and inter- gene flow, genetic structure, variability, linkage mapping and evolutionary relationships in bamboo PMID:21364824

  2. Omega-3 Eicosapentaenoic Acid Decreases CD133 Colon Cancer Stem-Like Cell Marker Expression While Increasing Sensitivity to Chemotherapy

    PubMed Central

    De Carlo, Flavia; Witte, Theodore R.; Hardman, W. Elaine; Claudio, Pier Paolo

    2013-01-01

    Colorectal cancer is the third leading cause of cancer-related death in the western world. In vitro and in vivo experiments showed that omega-3 polyunsaturated fatty acids (n-3 PUFAs) can attenuate the proliferation of cancer cells, including colon cancer, and increase the efficacy of various anticancer drugs. However, these studies address the effects of n-3 PUFAs on the bulk of the tumor cells and not on the undifferentiated colon cancer stem-like cells (CSLCs) that are responsible for tumor formation and maintenance. CSLCs have also been linked to the acquisition of chemotherapy resistance and to tumor relapse. Colon CSLCs have been immunophenotyped using several antibodies against cellular markers including CD133, CD44, EpCAM, and ALDH. Anti-CD133 has been used to isolate a population of colon cancer cells that retains stem cells properties (CSLCs) from both established cell lines and primary cell cultures. We demonstrated that the n-3 PUFA, eicosapentaenoic acid (EPA), was actively incorporated into the membrane lipids of COLO 320 DM cells. 25 uM EPA decreased the cell number of the overall population of cancer cells, but not of the CD133 (+) CSLCs. Also, we observed that EPA induced down-regulation of CD133 expression and up-regulation of colonic epithelium differentiation markers, Cytokeratin 20 (CK20) and Mucin 2 (MUC2). Finally, we demonstrated that EPA increased the sensitivity of COLO 320 DM cells (total population) to both standard-of-care chemotherapies (5-Fluorouracil and oxaliplatin), whereas EPA increased the sensitivity of the CD133 (+) CSLCs to only 5-Fluorouracil. PMID:23874993

  3. Evaluation of Myofibroblasts by Expression of Alpha Smooth Muscle Actin: A Marker in Fibrosis, Dysplasia and Carcinoma

    PubMed Central

    Malathi, N.; Narashiman, Sangeetha; Rajan, Sharada T

    2014-01-01

    Objective: Evaluation of Myofibroblasts by studying expression of Alpha smooth muscle actin: A marker of Fibrosis, Dysplasia and Carcinoma. Background: Myofibroblasts are cells that have contractile properties and are involved in inflammation, wound healing, fibrosis and oncogenesis in most of the organs and tissues. They are involved in healing and granulation tissue formation which occur after tissue injuries, also produce inflammatory mediators, growth factors and help in extracellular matrix reorganization by secretion of proteins like collagen, fibronectin, etc. Because of their component, Alpha smooth muscle actin ([alpha]-SMA), they are involved in the contraction of extracellular matrix and aid in tissue contraction. The myofibroblasts disappear by apoptosis after completion of repair, but their persistence causes a dysfunction in the repair mechanism, leading to excessive contraction and extracellular matrix (ECM) secretion and thus, fibrosis. The purpose of this study was to evaluate the presence of myofibroblasts in cases of Oral Submucous fibrosis (OSMF), which consisted of very early, early and moderately advanced OSMF, OSMF with dysplasia and oral squamous cell carcinoma (OSCC), by detecting (alpha)-SMA, which is a specific marker for myofibroblasts. Materials and Methods: The study sample consisted of three groups which comprised of 41 cases of OSMF, 10 cases of OSMF with dysplasia and 11 cases of OSCC. All the cases were subjected to immunohistochemistry by using (alpha)-SMA antibody for detection of myofibroblasts. Results: The presence of myofibroblasts was significantly higher in oral squamous cell carcinomas as compared to that in OSMF with dysplasia and OSMF. A statistical significance was also noted between the staining index and age of the individuals and the staining index and duration of the habit. Conclusion: Myofibroblasts play a role in fibrosis, as was seen in OSMF. Activated myofibroblasts secrete proteolytic enzymes and cause matrix

  4. Expression of KLF4 is a predictive marker for survival in pediatric Burkitt lymphoma.

    PubMed

    Valencia-Hipόlito, Alberto; Hernández-Atenógenes, Miriam; Vega, Gabriel G; Maldonado-Valenzuela, Altagracia; Ramon, Guillermo; Mayani, Héctor; Peña Alonso, Yolanda; Martinez-Maza, Otoniel; Méndez-Tenorio, Alfonso; Huerta-Yepez, Sara; Bonavida, Benjamin; Vega, Mario I

    2014-08-01

    Krüppel-like factor 4 (KLF4) is expressed in a variety of tissues with diverse physiological functions and activities. KLF4 can also function as a tumor suppressor or an oncogene, depending on the cellular context. Its role in hematological malignancies is controversial. This study examined the expression levels of KLF4 by immunohistochemistry in 73 pediatric non-Hodgkin lymphomas (NHLs) in a tissue microarray and also on several B-NHL cell lines. Elevated levels of KLF4 expression were detected in 66% of lymphoma cases and were more frequent in the Burkitt lymphoma (p = 0.05) subtype. There was a significant predictive power for outcome with low KLF4 expression, predicting a favorable overall survival compared to high levels. Multivariate analyses confirmed the association of KLF4 expression with unfavorable overall survival (p < 0.005). These findings were consistent with analyses in existing NHL microarray datasets. The present findings revealed that KLF4 is overexpressed in Burkitt pediatric lymphoma and is a potential biomarker for inferior overall survival. PMID:24067139

  5. Induction of renal senescence marker protein-30 (SMP30) expression by testosterone and its contribution to urinary calcium absorption in male rats.

    PubMed

    Lin, Po-Han; Jian, Cai-Yun; Chou, Jou-Chun; Chen, Chien-Wei; Chen, Chih-Chieh; Soong, Christina; Hu, Sindy; Lieu, Fu-Kong; Wang, Paulus S; Wang, Shyi-Wu

    2016-01-01

    The aim of this study was to investigate the involvement of androgen, mainly testosterone, in the expression of renal senescence marker protein-30 (SMP30) in male rats. We found that the renal SMP30 expression was up-regulated by endogenous testosterone stimulation during puberty. Interestingly, androgen-deficient orchidectomized (ORX) rats exhibited lower SMP30 mRNA and protein expression in the kidney, and that was restored by testosterone propionate (TP) replacement. Abrogation of androgen receptor (AR) activity by co-treatment with flutamide abolished testosterone-induced SMP30 expression in the kidney as well as in the NRK52E cells. However, SMP30 expression was unaltered in the liver of ORX rats. We also showed a positive correlation between renal SMP30 expression and plasma testosterone level during the aging process. TP-induced SMP30 expression in ovariectomized (OVX) rats was observed and was an evidence to explain the gender difference of SMP30 levels. Immunofluorescence assay showed that renal SMP30 was specifically expressed in the proximal tubular segments of the kidney. The urinary Ca(2+) level was increased in both ORX and male aging rats. Taken together, our results indicate a novel role of testosterone in regulating SMP30 expression specifically in the kidney to contribute to urinary calcium absorption. PMID:27553527

  6. Induction of renal senescence marker protein-30 (SMP30) expression by testosterone and its contribution to urinary calcium absorption in male rats

    PubMed Central

    Lin, Po-Han; Jian, Cai-Yun; Chou, Jou-Chun; Chen, Chien-Wei; Chen, Chih-Chieh; Soong, Christina; Hu, Sindy; Lieu, Fu-Kong; Wang, Paulus S.; Wang, Shyi-Wu

    2016-01-01

    The aim of this study was to investigate the involvement of androgen, mainly testosterone, in the expression of renal senescence marker protein-30 (SMP30) in male rats. We found that the renal SMP30 expression was up-regulated by endogenous testosterone stimulation during puberty. Interestingly, androgen-deficient orchidectomized (ORX) rats exhibited lower SMP30 mRNA and protein expression in the kidney, and that was restored by testosterone propionate (TP) replacement. Abrogation of androgen receptor (AR) activity by co-treatment with flutamide abolished testosterone-induced SMP30 expression in the kidney as well as in the NRK52E cells. However, SMP30 expression was unaltered in the liver of ORX rats. We also showed a positive correlation between renal SMP30 expression and plasma testosterone level during the aging process. TP-induced SMP30 expression in ovariectomized (OVX) rats was observed and was an evidence to explain the gender difference of SMP30 levels. Immunofluorescence assay showed that renal SMP30 was specifically expressed in the proximal tubular segments of the kidney. The urinary Ca2+ level was increased in both ORX and male aging rats. Taken together, our results indicate a novel role of testosterone in regulating SMP30 expression specifically in the kidney to contribute to urinary calcium absorption. PMID:27553527

  7. Molecular characterization and expression pattern of a germ cell marker gene dnd in gibel carp (Carassius gibelio).

    PubMed

    Li, Shi-Zhu; Liu, Wei; Li, Zhi; Wang, Yang; Zhou, Li; Yi, Mei-Sheng; Gui, Jian-Fang

    2016-10-10

    As a germ cell marker gene, Dead end (dnd) has been identified and characterized in many vertebrates. Recently, we created a complete germ cell-depleted gonad model by the dnd-specific morpholino-mediated knockdown approach, and revealed sex-biased gene expression alteration through utilizing unisexual gynogenetic superiority in polyploid gibel carp. However, dnd and its expression pattern are still unclear in the gibel carp. In this study, we further analyzed molecular characterization of gibel carp dnd and its dynamic expression pattern during gametogenesis and embryogenesis. Similar to other homologs in vertebrates, gibel carp dnd contains a conserved RRM motif and five other motifs, and is highly evolutionary conserved in genomic organization and neighborhood gene synteny. RT-PCR and Western blot analyses showed its gonad-specific expression intensively in testis and ovary. Section in situ hybridization (SISH) and immunofluorescence localization revealed its dynamic expression pattern specific to oogenic cells and spermatogenetic cells during oogenesis and spermatogenesis. Moreover, its temporal and spatial distribution specific to PGCs were also demonstrated by RT-PCR and whole mount in situ hybridization (WISH) during embryogenesis. Therefore, gibel carp Dnd is a conserved germ cell marker during gametogenesis, and its maternal transcript is also a useful marker for tracing PGC specification and migration. PMID:27418526

  8. Ropinirole regulates emotionality and neuronal activity markers in the limbic forebrain.

    PubMed

    Mavrikaki, Maria; Schintu, Nicoletta; Nomikos, George G; Panagis, George; Svenningsson, Per

    2014-12-01

    Restless legs syndrome (RLS) and Parkinson's disease (PD) are movement disorders usually accompanied by emotional and cognitive deficits. Although D3/D2 receptor agonists are effective against motor and non-motor deficits in RLS and PD, the exact behavioral and neurochemical effects of these drugs are not clearly defined. This study aimed to evaluate the effects of acute ropinirole (0, 0.1, 1 or 10 mg/kg, i.p.), a preferential D3/D2 receptor agonist, on intracranial self-stimulation (ICSS), spontaneous motor activity, anxiety- and depression-like behaviors, spatial reference and working memory in rats as well as on certain markers of neuronal activity, i.e. induction of immediate early genes, such as c-fos and arc, and crucial phosphorylations on GluA1 subunit of α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and NA1, NA2A and NA2B subunits of N-methyl-D-aspartate (NMDA) receptors. Ropinirole decreased ICSS thresholds and induced anxiolytic- and antidepressive-like effects without affecting motor activity or spatial memory. The effects on emotionality were associated with a decrease in p-Ser897-NA1 and an increase in p-Tyr1472-NA2B in the ventral striatum as well as an increased induction of c-fos messenger RNA (mRNA) in the prefrontal cortex (PFC) and decreased expression of arc mRNA in the striatum and the shell of the nucleus accumbens. Our data indicate that ropinirole significantly affects emotionality at doses (1-10 mg/kg, i.p.) that exert no robust effects on locomotion or cognition. The data reinforce the use of D3/D2 receptor agonists in the treatment of RLS and PD patients characterized by emotional deficits and suggest that altered NMDA-mediated neurotransmission in the limbic forebrain may underlie some of ropinirole's therapeutic actions.

  9. Downregulation of lncRNA-MALAT1 Affects Proliferation and the Expression of Stemness Markers in Glioma Stem Cell Line SHG139S.

    PubMed

    Han, Yong; Zhou, Liang; Wu, Tingfeng; Huang, Yulun; Cheng, Zhe; Li, Xuetao; Sun, Ting; Zhou, Youxin; Du, Ziwei

    2016-10-01

    Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is among the most abundant and highly conserved lncRNAs, which has been detected in a wide variety of human tumors, including gastric cancer, gallbladder cancer, and so on. Previous research has showed that MALAT1 can activate LTBP3 gene in mesenchymal stem cells. However, the specific roles of MALAT1 in glioma stem cells (GSCs) remain unclear. In this study, we aimed to identify the effects of MALAT1 on proliferation and the expression of stemness markers on glioma stem cell line SHG139S. Our results showed that downregulation of MALAT1 suppressed the expression of Sox2 and Nestin which are related to stemness, while downregulation of MALAT1 promoted the proliferation in SHG139S. Further research on the underlying mechanism showed that the effects of MALAT1 downregulation on SHG139S were through regulating ERK/MAPk signaling activity. And we also found that downregulation of MALAT1 could activate ERK/MAPK signaling and promoted proliferation in SHG139 cells. These findings show that MALAT1 plays an important role in regulating the expression of stemness markers and proliferation of SHG139S, and provide a new research direction to target the progression of GSCs.

  10. Quantification of cells expressing markers of proliferation and apoptosis in chronic tonsilitis.

    PubMed

    Avramović, V; Petrović, V; Jović, M; Vlahović, P

    2015-10-01

    During chronic tonsillitis, the relationship between proliferation and apoptosis of lymphocytes in tonsillar follicles can be disturbed, which gives rise to attenuation of tonsil immunocompetence and diminishing its contribution in systemic immunity. In this study, we have quantified the cells expressing the markers of proliferation and apoptosis in the follicles of the palatine tonsil. Six tonsils from patients aged 10-29 years with hypertrophic tonsillitis and five tonsils from patients aged 18-22 years with recurrent tonsillitis were studied. The sections of paraffin blocks of tonsillar tissue were stained by the immunohistochemical LSAB/HRP method with the utilisation of antibodies for: Ki-67 antigen-cell marker of proliferation; Bcl-2 and survivin anti-apoptotic factors and Fas/CD95, caspase-3 and Bax pro-apoptotic factors. The size of lymphoid follicles, i.e. mean follicle area and number of lymphoid follicle immunopositive cells per mm2 of a slice area, i.e. numerical areal density were determined by the quantitative image analysis. The localisation of Ki-67, Bcl-2, survivin, Fas/CD95, caspase-3 and Bax- immunopositive cells inside the palatine tonsil was similar in both types of tonsillitis. The number of Ki-67 immunopositive cells was significantly (p < 0.01) larger in the tonsils with hypertrophic tonsillitis (14681.4 ± 1460.5) in comparison to those with recurrent tonsillitis (12491.4 ± 2321.6), although the number of survivin and caspase-3 immunopositive cells was significantly (p < 0.05) larger in recurrent tonsillitis (survivin, 406.9 ± 98.4; caspase-3, 350.4 ± 119.4) when compared to those with hypertrophic tonsillitis (survivin, 117.4 ± 14.5; caspase-3, 210 ± 24). Our results show that the rate of the proliferation and apoptosis of follicular lymphocytes is different in various types of tonsillitis. This suggests that the immunological potential of the palatine tonsil varies in patients with hypertrophic and recurrent tonsillitis, which in

  11. Expressions of Senescence-Associated β-Galactosidase and Senescence Marker Protein-30 are Associated with Lens Epithelial Cell Apoptosis.

    PubMed

    Zhou, Dan; Yin, Dan; Xiao, Fang; Hao, Jie

    2015-11-30

    BACKGROUND To investigate associations of senescence marker protein-30 and senescence-associated β-galactosidase expression with lens epithelial cells apoptosis among Chinese age-related cataract patients. MATERIAL AND METHODS A total of 145 age-related cataract patients (69 cases with nuclear cataract in 91 eyes and 76 cases of cortical cataract with 102 eyes) were enrolled in our study. An annular tear of the central part of anterior lens capsules was performed for each patient. Immunohistochemical staining and real-time PCR were used to detect the protein and mRNA expression levels, and TUNEL was used to assess lens epithelial cells apoptosis. Comparisons of protein expression levels and lens epithelial cells apoptosis were made between the 2 groups. RESULTS The results showed a higher protein expression level of senescence marker protein-30 in surrounding parts of the anterior lens capsule compared with the central part of the anterior lens capsule; however, the positive rate of senescence-associated β-galactosidase was remarkably higher in the central part than in the surrounding part. Compared with cortical cataract patients, nuclear cataract patients had elevated senescence marker protein-30 protein and mRNA expression levels, but had a decreased positive rate of senescence-associated β-galactosidase. TUNEL results showed that the lens epithelial cell apoptosis rate was higher in the central part of the anterior lens capsule than in the surrounding part in both groups. Within either central or surrounding area of anterior lens capsule, cortical cataract patients exhibited a significantly higher lens epithelial cell apoptosis rate in contrast with nuclear cataract patients. CONCLUSIONS Our study results suggest that senescence marker protein-30 and senescence-associated β-galactosidase expressions in both nuclear cataract and cortical cataract patients were associated with lens epithelial cells apoptosis.

  12. Expressions of Senescence-Associated β-Galactosidase and Senescence Marker Protein-30 are Associated with Lens Epithelial Cell Apoptosis

    PubMed Central

    Zhou, Dan; Yin, Dan; Xiao, Fang; Hao, Jie

    2015-01-01

    Background To investigate associations of senescence marker protein-30 and senescence-associated β-galactosidase expression with lens epithelial cells apoptosis among Chinese age-related cataract patients. Material/Methods A total of 145 age-related cataract patients (69 cases with nuclear cataract in 91 eyes and 76 cases of cortical cataract with 102 eyes) were enrolled in our study. An annular tear of the central part of anterior lens capsules was performed for each patient. Immunohistochemical staining and real-time PCR were used to detect the protein and mRNA expression levels, and TUNEL was used to assess lens epithelial cells apoptosis. Comparisons of protein expression levels and lens epithelial cells apoptosis were made between the 2 groups. Results The results showed a higher protein expression level of senescence marker protein-30 in surrounding parts of the anterior lens capsule compared with the central part of the anterior lens capsule; however, the positive rate of senescence-associated β-galactosidase was remarkably higher in the central part than in the surrounding part. Compared with cortical cataract patients, nuclear cataract patients had elevated senescence marker protein-30 protein and mRNA expression levels, but had a decreased positive rate of senescence-associated β-galactosidase. TUNEL results showed that the lens epithelial cell apoptosis rate was higher in the central part of the anterior lens capsule than in the surrounding part in both groups. Within either central or surrounding area of anterior lens capsule, cortical cataract patients exhibited a significantly higher lens epithelial cell apoptosis rate in contrast with nuclear cataract patients. Conclusions Our study results suggest that senescence marker protein-30 and senescence-associated β-galactosidase expressions in both nuclear cataract and cortical cataract patients were associated with lens epithelial cells apoptosis. PMID:26619319

  13. Senescence marker protein 30 (SMP30)/regucalcin (RGN) expression decreases with aging, acute liver injuries and tumors in zebrafish

    SciTech Connect

    Fujisawa, Koichi; Terai, Shuji; Hirose, Yoshikazu; Takami, Taro; Yamamoto, Naoki; Sakaida, Isao

    2011-10-22

    Highlights: {yields} Zebrafish SMP30/RGN mRNA expression decreases with aging. {yields} Decreased expression was observed in liver tumors as compared to the surrounding area. {yields} SMP30/RGN is important for liver proliferation and tumorigenesis. -- Abstract: Senescence marker protein 30 (SMP30)/regucalcin (RGN) is known to be related to aging, hepatocyte proliferation and tumorigenesis. However, expression and function of non-mammalian SMP30/RGN is poorly understood. We found that zebrafish SMP30/RGN mRNA expression decreases with aging, partial hepatectomy and thioacetamide-induced acute liver injury. SMP30/RGN expression was also greatly decreased in a zebrafish liver cell line. In addition, we induced liver tumors in adult zebrafish by administering diethylnitrosamine. Decreased expression was observed in foci, hepatocellular carcinomas, cholangiocellular carcinomas and mixed tumors as compared to the surrounding area. We thus showed the importance of SMP30/RGN in liver proliferation and tumorigenesis.

  14. Regulation of Cardiac Expression of the Diabetic Marker MicroRNA miR-29

    PubMed Central

    Arnold, Nicholas; Koppula, Purushotham Reddy; Gul, Rukhsana; Luck, Christian; Pulakat, Lakshmi

    2014-01-01

    Diabetes mellitus (DM) is an independent risk factor for heart disease and its underlying mechanisms are unclear. Increased expression of diabetic marker miR-29 family miRNAs (miR-29a, b and c) that suppress the pro-survival protein Myeloid Cell Leukemia 1(MCL-1) is reported in pancreatic β-cells in Type 1 DM. Whether an up-regulation of miR-29 family miRNAs and suppression of MCL-1 (dysregulation of miR-29-MCL-1 axis) occurs in diabetic heart is not known. This study tested the hypothesis that insulin regulates cardiac miR-29-MCL-1 axis and its dysregulation correlates with DM progression. In vitro studies with mouse cardiomyocyte HL-1 cells showed that insulin suppressed the expression of miR-29a, b and c and increased MCL-1 mRNA. Conversely, Rapamycin (Rap), a drug implicated in the new onset DM, increased the expression of miR-29a, b and c and suppressed MCL-1 and this effect was reversed by transfection with miR-29 inhibitors. Rap inhibited mammalian target of rapamycin complex 1 (mTORC1) signaling in HL-1 cells. Moreover, inhibition of either mTORC1 substrate S6K1 by PF-4708671, or eIF4E-induced translation by 4E1RCat suppressed MCL-1. We used Zucker diabetic fatty (ZDF) rat, a rodent model for DM, to test whether dysregulation of cardiac miR-29-MCL-1 axis correlates with DM progression. 11-week old ZDF rats exhibited significantly increased body weight, plasma glucose, insulin, cholesterol, triglycerides, body fat, heart weight, and decreased lean muscle mass compared to age-matched lean rats. Rap treatment (1.2 mg/kg/day, from 9-weeks to 15-weeks) significantly reduced plasma insulin, body weight and heart weight, and severely dysregulated cardiac miR-29-MCL1 axis in ZDF rats. Importantly, dysregulation of cardiac miR-29-MCL-1 axis in ZDF rat heart correlated with cardiac structural damage (disorganization or loss of myofibril bundles). We conclude that insulin and mTORC1 regulate cardiac miR-29-MCL-1 axis and its dysregulation caused by reduced insulin

  15. Gene expression profiles in granuloma tissue reveal novel diagnostic markers in sarcoidosis.

    PubMed

    Christophi, George P; Caza, Tiffany; Curtiss, Christopher; Gumber, Divya; Massa, Paul T; Landas, Steve K

    2014-06-01

    Sarcoidosis is an immune-mediated multisystem disease characterized by the formation of non-caseating granulomas. The pathogenesis of sarcoidosis is unclear, with proposed infectious or environmental antigens triggering an aberrant immune response in susceptible hosts. Multiple pro-inflammatory signaling pathways have been implicated in mediating macrophage activation and granuloma formation in sarcoidosis, including IFN-γ/STAT-1, IL-6/STAT-3, and NF-κB. It is difficult to distinguish sarcoidosis from other granulomatous diseases or assess disease severity and treatment response with histopathology alone. Therefore, development of improved diagnostic tools is imperative. Herein, we describe an efficient and reliable technique to classify granulomatous disease through selected gene expression and identify novel genes and cytokine pathways contributing to the pathogenesis of sarcoidosis. We quantified the expression of twenty selected mRNAs extracted from formalin-fixed paraffin embedded (FFPE) tissue (n = 38) of normal lung, suture granulomas, sarcoid granulomas, and fungal granulomas. Utilizing quantitative real-time RT-PCR we analyzed the expression of several genes, including IL-6, COX-2, MCP-1, IFN-γ, T-bet, IRF-1, Nox2, IL-33, and eotaxin-1 and revealed differential regulation between suture, sarcoidosis, and fungal granulomas. This is the first study demonstrating that quantification of target gene expression in FFPE tissue biopsies is a potentially effective diagnostic and research tool in sarcoidosis.

  16. Claudin-3 expression in radiation-exposed rat models: A potential marker for radiation-induced intestinal barrier failure

    SciTech Connect

    Shim, Sehwan; Lee, Jong-geol; Bae, Chang-hwan; Lee, Seung Bum; Jang, Won-Suk; Lee, Sun-Joo; Lee, Seung-Sook; Park, Sunhoo

    2015-01-02

    Highlights: • Irradiation increased intestinal bacterial translocation, accompanied by claudin protein expression in rats. • Neurotensin decreased the bacterial translocation and restored claudin-3 expression. • Claudin-3 can be used as a marker in evaluating radiation induced intestinal injury. - Abstract: The molecular events leading to radiation-induced intestinal barrier failure are not well known. The influence of the expression of claudin proteins in the presence and absence of neurotensin was investigated in radiation-exposed rat intestinal epithelium. Wistar rats were randomly divided into control, irradiation, and irradiation + neurotensin groups, and bacterial translocation to the mesenteric lymph node and expression of claudins were determined. Irradiation led to intestinal barrier failure as demonstrated by significant bacterial translocation. In irradiated terminal ilea, expression of claudin-3 and claudin-4 was significantly decreased, and claudin-2 expression was increased. Administration of neurotensin significantly reduced bacterial translocation and restored the structure of the villi as seen by histologic examination. Among the three subtype of claudins, only claudin-3 expression was restored. These results suggest that the therapeutic effect of neurotensin on the disruption of the intestinal barrier is associated with claudin-3 alteration and that claudin-3 could be used as a marker in evaluating radiation-induced intestinal injury.

  17. Expression kinetics of hepatic progenitor markers in cellular models of human liver development recapitulating hepatocyte and biliary cell fate commitment.

    PubMed

    Chaudhari, Pooja; Tian, Lipeng; Deshmukh, Abhijeet; Jang, Yoon-Young

    2016-09-01

    Due to the limitations of research using human embryos and the lack of a biological model of human liver development, the roles of the various markers associated with liver stem or progenitor cell potential in humans are largely speculative, and based on studies utilizing animal models and certain patient tissues. Human pluripotent stem cell-based in vitro multistage hepatic differentiation systems may serve as good surrogate models for mimicking normal human liver development, pathogenesis and injury/regeneration studies. Here, we describe the implications of various liver stem or progenitor cell markers and their bipotency (i.e. hepatocytic- and biliary-epithelial cell differentiation), based on the pluripotent stem cell-derived model of human liver development. Future studies using the human cellular model(s) of liver and biliary development will provide more human relevant biological and/or pathological roles of distinct markers expressed in heterogeneous liver stem/progenitor cell populations.

  18. BMP4 Cooperates with Retinoic Acid to Induce the Expression of Differentiation Markers in Cultured Mouse Spermatogonia

    PubMed Central

    Feng, Yanmin; Feng, Xue; Wang, Xiuxia; Gan, Haiyun; Wang, Lixian; Lin, Xiwen

    2016-01-01

    Spermatogenesis is sustained by the proliferation and differentiation of spermatogonial stem cells (SSCs). However, the molecules controlling these processes remain largely unknown. Here, we developed a simplified high concentration serum-containing system for the culture of mouse SSCs. Analysis of SSCs markers and transplantation results revealed that the cultured spermatogonia retained stem cell characteristics after long-term in vitro propagation. Using this culture system, the expression and function of bone morphogenetic protein 4 (BMP4) were explored. Immunostaining showed that BMP4 was predominantly expressed in germ cells and that its level increased as spermatogenesis progresses. BMP4 receptors BMPR1A and BMPRII were present in spermatogonia, spermatocytes, and round spermatids. Moreover, despite the mRNAs of these two genes being present in mouse Sertoli cells, only BMPRII was detected by using Western blotting assays. While exogenous BMP4 by itself did not induce the expression of Stra8 and c-Kit, two marker genes of differentiating spermatogonia, a significant cooperative effect of BMP4 and retinoic acid (RA) was observed. Moreover, pretreatment of cultured spermatogonia with the BMP4 antagonist Noggin could inhibit RA-induced expression of these two marker genes. In conclusion, BMP4 may exert autocrine effects and act cooperatively with RA to induce the differentiation of spermatogonia in vivo. PMID:27795714

  19. Fos expression in the vestibular brainstem: what one marker can tell us about the network.

    PubMed

    Kaufman, Galen D

    2005-12-01

    Fos inducible transcription factor expression in rodent brains (rats and gerbils) during manipulations of vestibular input is reviewed. Stimuli included centripetal hypergravity, unilateral labyrinth lesion or semicircular canal plugging, rotational axis cross-coupling (Coriolis forces), high and low rotational vestibulo-ocular reflex gain adaptation, translabyrinth galvanic stimulation, pharmacological manipulation, and combinations thereof. Each type of stimulation elicited unique but partially redundant response patterns in the vestibulo-olivo-cerebellar (VOC) network that reflect the origin and interaction of the labyrinth inputs. On the basis of these patterns, a trained observer can predict what the animal experienced during testing; the patterns of VOC Fos expression reveal a trace of recent genomic activity. Based on principal component analysis, VOC network modules associated with lesion recovery, spatial representation and the calibration of gravity, and optokinetic influences are proposed. Probable and possible gene targets of the Fos protein are also reviewed.

  20. Markers of oxidative stress and erythrocyte antioxidant enzyme activity in older men and women with differing physical activity.

    PubMed

    Rowiński, Rafał; Kozakiewicz, Mariusz; Kędziora-Kornatowska, Kornelia; Hübner-Woźniak, Elżbieta; Kędziora, Józef

    2013-11-01

    The aim of the present study was to examine the relationship between markers of oxidative stress and erythrocyte antioxidant enzyme activity and physical activity in older men and women. The present study included 481 participants (233 men and 248 women) in the age group 65-69 years (127 men and 125 women) and in the age group 90 years and over (106 men and 123 women). The classification of respondents by physical activity was based on answers to the question if, in the past 12 months, they engaged in any pastimes which require physical activity. The systemic oxidative stress status was assessed by measuring plasma iso-PGF2α and protein carbonyl concentration as well as erythrocyte antioxidant enzymes activity, i.e., superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR). The concentration of plasma iso-PGF2α and protein carbonyls (CP) was lower in groups of younger men and women compared to the respective older groups. In all examined groups, physical activity resulted in decrease of these oxidative stress markers and simultaneously caused adaptive increase in the erythrocyte SOD activity. Additionally, in active younger men CAT, GPx, and GR activities were higher than in sedentary ones. In conclusion, oxidative stress increase is age-related, but physical activity can reduce oxidative stress markers and induce adaptive increase in the erythrocyte antioxidant enzyme activity, especially SOD, even in old and very old men and women.

  1. Expression of the pituitary stem/progenitor marker GFRα2 in human pituitary adenomas and normal pituitary

    PubMed Central

    Mathioudakis, Nestoras; Sundaresh, Ram; Larsen, Alexandra; Ruff, William; Schiller, Jennifer; Cázares, Hugo Guerrero; Burger, Peter; Salvatori, Roberto; Quiñones-Hinojosa, Alfredo

    2014-01-01

    Purpose Recent studies suggest that adult pituitary stem cells may play a role in pituitary tumorigenesis. We sought to explore whether the Glial cell-line derived neurotrophic factor receptor alpha 2 (GFRα2), a recently described pituitary stem/progenitor marker, might be differentially expressed in pituitary adenomas versus normal pituitary. Methods The expression of GFRα2 and other members of the GFR receptor family (GFRα1, α3, α4) were analyzed using RT-PCR, western blot, and immunohistochemistry in 39 pituitary adenomas, 14 normal pituitary glands obtained at autopsy, and cDNA from 3 normal pituitaries obtained commercially. Results GFRα2 mRNA was ~2.6 fold under-expressed in functioning adenomas (P <0.01) and ~3.5 fold over-expressed in non-functioning adenomas (NFAs) (P <0.05) compared to normal pituitary. Among NFAs, GFRα2 was significantly over-expressed (~5-fold) in the gonadotropinoma subtype only (P<0.05). GFRα2 protein expression appeared to be higher in most NFAs, although there was heterogeneity in protein expression in this group. GFRα2 protein expression appeared consistently lower in functioning adenomas by IHC and western blot. In normal pituitary, GFRα2 was localized in Rathke’s remnant, the putative pituitary stem cell niche, and in corticotropes. Conclusion Our results suggest that the pituitary stem cell marker GFRα2 is under-expressed in functioning adenomas and over-expressed in NFAs, specifically gonadotropinomas. Further studies are required to elucidate whether over-expression of GFRα2 in gonadotropinomas might play a role in pituitary tumorigenesis. PMID:24402129

  2. Expression of p63 and Bcl-2 in Malignant Thyroid Tumors and their Correlation with other Diagnostic Immunocytochemical Markers

    PubMed Central

    Jain, Shyama; Khurana, Nita; Kakar, Arun Kumar

    2016-01-01

    Introduction Bcl-2 is a marker recently studied in thyroid tumours and proposed to have prognostic significance. p63 is expressed in a proportion of papillary thyroid carcinoma cases and may have a role in tumour progression. Aim To study expression of Bcl2 and p63 in thyroid tumours and correlation of Bcl-2 with diagnostic markers including Thyroglobulin, Calcitonin and Carcinoembryonic antigen. Materials and Methods Cytology smears of 35 cases of thyroid cancer were studied over a period of 18 months. In 20 cases histopathology was available. Immunocytochemistry for Bcl-2 and p63 was done, and diagnostic markers were applied as and when required. Results p63 showed focal nuclear expression in 46.1% of papillary thyroid carcinoma cases, and was negative in all other tumours. Bcl-2 was positive in 88.9% of follicular carcinomas, 100% of papillary carcinomas and 83.3% of medullary carcinoma cases, and showed focal weak expression in 40% of Anaplastic Carcinoma (ATC) cases, thereby signifying down regulation (p-value = 0.001). There was significant down regulation of Thyroglobulin (Tg) in ATC vs well differentiated follicular derived tumours (p-value ≤ 0.016). Positive correlation was noted between expression of Bcl-2 and Calcitonin (0.93) and Bcl-2 and Carcinoembryonic Antigen (CEA) (0.89), and weak positive correlation (0.65) between Tg and Bcl-2. Conclusion Bcl-2 is downregulated in anaplastic carcinomas as compared to well differentiated thyroid tumours, and shows correlation with differentiation associated tumour antigens. Thus, loss of Bcl-2 was associated with loss of differentiation in thyroid tumours. Anaplastic carcinoma as such is associated with worse prognosis and loss of Bcl-2 may be partly responsible for the same. p63 is specific but less sensitive marker for PTC. Further studies are required to determine the role of Bcl-2 and p63 in thyroid tumours. PMID:27630849

  3. Transcriptional coactivator undifferentiated embryonic cell transcription factor 1 expressed in spermatogonial stem cells: a putative marker of boar spermatogonia.

    PubMed

    Lee, Won-Young; Lee, Kyung-Hoon; Heo, Young-Tae; Kim, Nam-Hyung; Kim, Jin-Hoi; Kim, Jae-Hwan; Moon, Sung-Hwan; Chung, Hak-Jae; Yoon, Min-Jung; Song, Hyuk

    2014-11-30

    Spermatogenesis is initiated from spermatogonial stem cells (SSCs), which are derived from gonocytes. Although some rodent SSC markers have been investigated, other species- and developmental stage-specific markers of spermatogonia have not been identified. The objective of this study was to characterize the expression of undifferentiated embryonic cell transcription factor 1 (UTF1) gene as a potential marker for spermatogonia and SSCs in the boar testis. In boar testis tissue at pre-pubertal stages (tissues collected at 5, 30, and 60 days of age), UTF1 gene expression was detected in almost all spermatogonia cells that expressed a protein gene product 9.5 (PGP9.5), and immunocytochemical analysis of isolated total testicular cells showed that 91.14% of cells staining for PGP9.5 also stained for UTF1. However, in boar testis tissue at pubertal and post-pubertal stages (tissues collected at 90, 120, 150, and 180 days of age), UTF1 was not detected in all PGP9.5-positive cells in the basement membrane. While some PGP9.5-positive cells stained for UTF1, other cells stained only for PGP9.5 or UTF1. PGP9.5, UTF1, and NANOG was assessed in in vitro cultures of pig SSCs (pSSCs) from testes collected at 5 days of age. The relative amounts of PGP9.5, NANOG, and UTF1 mRNA were greater in pSSC colonies than in testis and muscle tissue. Thus, the UTF1 gene is expressed in PGP9.5-positive spermatogonia cells of pigs at 5 days of age, and its expression is maintained in cultured pSSC colonies, suggesting that UTF1 is a putative marker for early-stage spermatogonia in the pre-pubertal pig testis. These findings will facilitate the study of spermatogenesis and applications in germ cell research.

  4. Expression of p63 and Bcl-2 in Malignant Thyroid Tumors and their Correlation with other Diagnostic Immunocytochemical Markers

    PubMed Central

    Jain, Shyama; Khurana, Nita; Kakar, Arun Kumar

    2016-01-01

    Introduction Bcl-2 is a marker recently studied in thyroid tumours and proposed to have prognostic significance. p63 is expressed in a proportion of papillary thyroid carcinoma cases and may have a role in tumour progression. Aim To study expression of Bcl2 and p63 in thyroid tumours and correlation of Bcl-2 with diagnostic markers including Thyroglobulin, Calcitonin and Carcinoembryonic antigen. Materials and Methods Cytology smears of 35 cases of thyroid cancer were studied over a period of 18 months. In 20 cases histopathology was available. Immunocytochemistry for Bcl-2 and p63 was done, and diagnostic markers were applied as and when required. Results p63 showed focal nuclear expression in 46.1% of papillary thyroid carcinoma cases, and was negative in all other tumours. Bcl-2 was positive in 88.9% of follicular carcinomas, 100% of papillary carcinomas and 83.3% of medullary carcinoma cases, and showed focal weak expression in 40% of Anaplastic Carcinoma (ATC) cases, thereby signifying down regulation (p-value = 0.001). There was significant down regulation of Thyroglobulin (Tg) in ATC vs well differentiated follicular derived tumours (p-value ≤ 0.016). Positive correlation was noted between expression of Bcl-2 and Calcitonin (0.93) and Bcl-2 and Carcinoembryonic Antigen (CEA) (0.89), and weak positive correlation (0.65) between Tg and Bcl-2. Conclusion Bcl-2 is downregulated in anaplastic carcinomas as compared to well differentiated thyroid tumours, and shows correlation with differentiation associated tumour antigens. Thus, loss of Bcl-2 was associated with loss of differentiation in thyroid tumours. Anaplastic carcinoma as such is associated with worse prognosis and loss of Bcl-2 may be partly responsible for the same. p63 is specific but less sensitive marker for PTC. Further studies are required to determine the role of Bcl-2 and p63 in thyroid tumours.

  5. The Calpain Inhibitor MDL28170 Induces the Expression of Apoptotic Markers in Leishmania amazonensis Promastigotes

    PubMed Central

    Marinho, Fernanda A.; Gonçalves, Keyla C. S.; Oliveira, Simone S. C.; Gonçalves, Diego S.; Matteoli, Filipe P.; Seabra, Sergio H.; Oliveira, Ana Carolina S.; Bellio, Maria; Oliveira, Selma S.; Souto-Padrón, Thaïs; d'Avila-Levy, Claudia M.; Santos, André L. S.; Branquinha, Marta H.

    2014-01-01

    Background Human cutaneous leishmaniasis is caused by distinct species, including Leishmania amazonensis. Treatment of cutaneous leishmaniasis is far from satisfactory due to increases in drug resistance and relapses, and toxicity of compounds to the host. As a consequence for this situation, the development of new leishmanicidal drugs and the search of new targets in the parasite biology are important goals. Methodology/Principal Findings In this study, we investigated the mechanism of death pathway induced by the calpain inhibitor MDL28170 on Leishmania amazonensis promastigote forms. The combined use of different techniques was applied to contemplate this goal. MDL28170 treatment with IC50 (15 µM) and two times the IC50 doses induced loss of parasite viability, as verified by resazurin assay, as well as depolarization of the mitochondrial membrane, which was quantified by JC-1 staining. Scanning and transmission electron microscopic images revealed drastic alterations on the parasite morphology, some of them resembling apoptotic-like death, including cell shrinking, surface membrane blebs and altered chromatin condensation pattern. The lipid rearrangement of the plasma membrane was detected by Annexin-V labeling. The inhibitor also induced a significant increase in the proportion of cells in the sub-G0/G1 phase, as quantified by propidium iodide staining, as well as genomic DNA fragmentation, detected by TUNEL assay. In cells treated with MDL28170 at two times the IC50 dose, it was also possible to observe an oligonucleossomal DNA fragmentation by agarose gel electrophoresis. Conclusions/Significance The data presented in the current study suggest that MDL28170 induces apoptotic marker expression in promastigotes of L. amazonensis. Altogether, the results described in the present work not only provide a rationale for further exploration of the mechanism of action of calpain inhibitors against trypanosomatids, but may also widen the investigation of the

  6. Development of expressed sequence tag-simple sequence repeat markers for Chrysanthemum morifolium and closely related species.

    PubMed

    Liu, H; Zhang, Q X; Sun, M; Pan, H T; Kong, Z X

    2015-01-01

    With the development of chrysanthemum breeding in recent years, an increasing number of wild species in genera related to Chrysanthemum were introduced to extend the genetic resources and facilitate the genetic improvement of chrysanthemums via hybridization. However, few simple sequence repeat (SSR) markers are available for marker-assisted breeding and population genetic studies of chrysanthemum and closely related species. Expressed sequence tags (ESTs) in public databases and cross-species transferable markers are considered to be a cost-effective means for developing sequence-based markers. In this study, 25 EST-SSRs were successfully developed from Chrysanthemum EST sequences for Chrysanthemum morifolium and closely related species. In total, 4164 unigene sequences were assembled from 7180 ESTs of chrysanthemum in GenBank, which were subsequently used to screen for the presence of microsatellites with the SSRIT software. The screening criteria were 8, 5, 4, and 3 repeating units for di-, tri-, tetra-, and penta- and higher-order nucleotides, respectively. Moreover, 310 SSR loci from 296 sequences were identified, and 198 primer pairs for SSR amplification were designed with the Primer Premier 5.0 software, of which 25 SSR loci showed polymorphic amplification in 52 species and varieties belonging to Chrysanthemum, Ajania, and Opisthopappus. The application of EST-SSR markers to the identification of intergeneric hybrids between Chrysanthemum and Ajania was demonstrated. Therefore, EST-SSRs can be developed for species that lack gene sequences or ESTs by utilizing ESTs of closely related species. PMID:26214436

  7. Markers of activated T cells on synovial fluid lymphocytes in rheumatoid arthritis.

    PubMed

    Mathieu, A

    1979-01-31

    Membrane markers of activated T lymphocytes of synovial fluid of two groups of patients with various forms of arthritis were studied. The first group (group A) concerns patients affected by rheumatoid arthritis (RA), and the other (group B) includes those affected by not immunologically-mediated arthropathies as osteoarthrosis, crystal synovitis, post-traumatic arthritis. Some other arthropathies included in a third group (group C) have been considered separately. Both the receptor for human group O Rh negative erythrocytes (H rosettes forming cells) and the receptor able to bind at 37 degrees C sheep red blood cells (stable-E-rosette forming cells) respectively were used as markers for the identification of activated T lymphocytes. The results show a marked increase of activated T cells in group A in comparison to group B. So the possible causes of this lymphocyte activation in rheumatoid patients are suggested.

  8. Intratumoral expression of cyclooxygenase-2 (COX-2) is a negative prognostic marker for patients with cutaneous melanoma.

    PubMed

    Kuźbicki, Łukasz; Lange, Dariusz; Stanek-Widera, Agata; Chwirot, Barbara W

    2016-10-01

    Because of the well-known heterogeneity of melanomas, prognosis of the disease is often difficult to assess even for lesions classified in similar stages. The aim of this study was to assess the usefulness of COX-2 as a melanoma prognostic marker and to establish an optimum algorithm for analysis of COX-2 expression levels in lesions of interest. Expression of COX-2 was detected immunohistochemically in standard sections of formalin-fixed paraffin-embedded tissue samples of 85 primary melanomas, 36 lymph node metastases, and five skin metastases including 39 cases of paired primary and metastatic lesions obtained from the same patient. Enhanced expression of COX-2 in primary melanomas is an indicator of poorer prognosis. A significant correlation was found between high expression of COX-2 in primary lesions and shorter survival. The enhancement of COX-2 expression is also positively correlated with other prognostic factors such as tumor thickness and infiltration level, ulceration, high mitotic index, more invasive histologic type, vertical growth phase, and lymph node metastasis. On the whole, the results suggest that intratumoral expression of COX-2 is a strong negative prognostic marker for patients with melanoma. Moreover, our work shows that a simple and objective immunohistochemical scoring algorithm involving the determination of only a percentage fraction of positively stained cells is sufficient to obtain the prognostic information.

  9. Radiation Dose-effects on Cell Cycle, Apoptosis, and Marker Expression of Ataxia Telangiectasia-Heterozygous Human Breast Epithelial Cells

    NASA Technical Reports Server (NTRS)

    Cruz, A.; Bors, K.; Jansen, H.; Richmond, R.

    2003-01-01

    Ataxia-telangiectasia (A-T) is a radiation-sensitive genetic condition. AT-heterozygous human mammary epithelial cells (HMEC) were irradiated using a Cs137 source in order to compare cell cycle, apoptosis, and marker expression responses across 3 radiation doses. No differences in cell cycle and apoptosis were found with any of the radiation doses used (30, 60, and 90 rads) compared with the unirradiated control (0 rad). At the same doses, however, differences were found in marker expression, such as keratin 18 (kl8), keratin 14 (k14), insulin-like growth factor I receptor (IGF-IR), and connexin 43 (cx43). This may indicate that radiation sensitivity in the heterozygous state may be initiated through signal transduction responses.

  10. Elevated expression of hepatic proliferative markers during early hepatocarcinogenesis in hepatitis-B virus transgenic mice lacking mdr1a-encoded P-glycoprotein.

    PubMed

    Bao, J J; Lee, B P; Stephens, L C; Sahin, A A; Van, N T; Johnston, D A; Ou, C N; Kuo, M T

    2000-10-01

    Recent studies have shown that expression levels of the multidrug resistance gene MDR1, which encodes the drug transporter P-glycoprotein, correlate with prognostic outcomes of certain tumor types. These findings suggest that expression of MDR1 may affect tumor behaviors. To address this issue further, we investigated the expression of mdr1a, a human MDR1 homolog, on the development of hepatocellular carcinoma in a transgenic mouse model carrying the liver-targeted expression of human hepatitis-B virus (HBV) surface antigen. The pathogenetic program was compared in HBV mice carrying either mdr1a(+/+) or mdr1a(-/-). We found that the expressions of proliferative activity markers, Ki67 nuclear antigen, and proliferating cell nuclear antigen were elevated in mdr1a(-/-) mice younger than 10 wk in comparison with those in the same age group of wild-type animals. Replication in the hepatic population as determined by bromodeoxyuridine incorporation tended to support observation that mdr1a(-/-) mice exhibited elevated labeling indices in this age group. Moreover, histologic staining and flow-cytometric analysis showed that the mdr1a(-/-) animals exhibited a higher cell population with polyploidy than did the mdr1a(+/+) counterparts of the same age. However, no significant differences in the expression of the liver-injury markers serum alanine transaminase and aspartate transaminase were observed. Although our results showed that absence of mdr1a expression is correlated with modest enhanced proliferative characteristics in the livers at stage before the development of hepatocellular carcinoma, the overall life spans between these two strains of mice were not significantly different. The implication of these findings to the role of P-glycoprotein in tumor development and cancer chemotherapy is discussed.

  11. Physical activity and bone turnover markers: a cross-sectional and a longitudinal study.

    PubMed

    Adami, Silvano; Gatti, Davide; Viapiana, Ombretta; Fiore, Carmelo Erio; Nuti, Ranuccio; Luisetto, Giovanni; Ponte, Marco; Rossini, Maurizio

    2008-12-01

    Strenuous physical activity in young individuals has an important effect on both bone mass and bone turnover but the effect of moderate physical activity in adults remains uncertain. In a large cohort (N = 530) of healthy premenopausal women, bone formation markers (osteocalcin and N-terminal propeptide of type 1 procollagen [P1NP]), but not serum C-telopeptide of type 1 collagen (sCTX), were found to be significantly associated with the level of physical activity, and this association remained significant after adjusting the data (ANCOVA) by age and body mass index. Mean spine and hip bone mineral density (BMD) values were positively associated with physical activity but this was statistically significant (P = 0.050) only for adjusted values of spine BMD. Twenty-four healthy sedentary premenopausal women, subscribing to participate in a community exercise program lasting a month, and 18 age-matched controls were included in the longitudinal study. Serum osteocalcin and P1NP, but not sCTX, rose significantly, by ca. 25%, only in the active group after a month of exercise. The changes in the two bone formation markers remained statistically significant for values adjusted for body weight, which fell significantly in the exercise group. In conclusion, both the cross-sectional and the longitudinal parts of our study demonstrate that even minor changes in physical activity are associated with a clear effect on bone formation markers.

  12. T lymphocyte surface expression of exhaustion markers as biomarkers of the efficacy of chemotherapy for tuberculosis

    PubMed Central

    Henao-Tamayo, Marcela; Irwin, Scott M.; Shang, Shaobin; Ordway, Diane; Orme, Ian M.

    2011-01-01

    Summary Predictive biomarkers illustrating the effectiveness of chemotherapeutic regimens for tuberculosis still remain elusive. To date, most are predicated on assays using sputum or serum; as a result, if not predictive, treatment failure in patients may not be evident for some time. We report here the results of a simple screening study in which T cell surface markers were examined in mice infected with Mycobacterium tuberculosis and then treated with drugs. These studies identified certain markers, the exhaustion markers PD-1 and TIM-3, as well as the marker KLRG-1, particularly on CD8 T cells, that changed in concert with reduction of the bacterial load in the lungs. While there is no guarantee these changes would also be seen on T cells in the blood, this approach should be further investigated. PMID:21530406

  13. Micro- and minisatellite-expressed sequence tag (EST) markers discriminate between populations of Rhipicephalus appendiculatus.

    PubMed

    Kanduma, Esther G; Mwacharo, Joram M; Sunter, Jack D; Nzuki, Inosters; Mwaura, Stephen; Kinyanjui, Peter W; Kibe, Michael; Heyne, Heloise; Hanotte, Olivier; Skilton, Robert A; Bishop, Richard P

    2012-06-01

    Biological differences, including vector competence for the protozoan parasite Theileria parva have been reported among populations of Rhipicephalus appendiculatus (Acari: Ixodidae) from different geographic regions. However, the genetic diversity and population structure of this important tick vector remain unknown due to the absence of appropriate genetic markers. Here, we describe the development and evaluation of a panel of EST micro- and minisatellite markers to characterize the genetic diversity within and between populations of R. appendiculatus and other rhipicephaline species. Sixty-six micro- and minisatellite markers were identified through analysis of the R. appendiculatus Gene Index (RaGI) EST database and selected bacterial artificial chromosome (BAC) sequences. These were used to genotype 979 individual ticks from 10 field populations, 10 laboratory-bred stocks, and 5 additional Rhipicephalus species. Twenty-nine markers were polymorphic and therefore informative for genetic studies while 6 were monomorphic. Primers designed from the remaining 31 loci did not reliably generate amplicons. The 29 polymorphic markers discriminated populations of R. appendiculatus and also 4 other Rhipicephalus species, but not R. zambeziensis. The percentage Principal Component Analysis (PCA) implemented using Multiple Co-inertia Analysis (MCoA) clustered populations of R. appendiculatus into 2 groups. Individual markers however differed in their ability to generate the reference typology using the MCoA approach. This indicates that different panels of markers may be required for different applications. The 29 informative polymorphic micro- and minisatellite markers are the first available tools for the analysis of the phylogeography and population genetics of R. appendiculatus. PMID:22789728

  14. Differential gene expression and apoptosis markers in presymptomatic scrapie affected sheep.

    PubMed

    Hedman, Carlos; Lyahyai, Jaber; Filali, Hicham; Marín, Belén; Serrano, Carmen; Monleón, Eva; Moreno, Bernardino; Zaragoza, Pilar; Badiola, Juan José; Martín-Burriel, Inmaculada; Bolea, Rosa

    2012-09-14

    Neuronal loss is one of the characteristics of scrapie neuropathology. Previous analysis of brains from sheep naturally infected with scrapie that were in a terminal stage did not detect a clear induction of apoptosis, although molecular changes were evidenced. As neuronal death could be occurring early in scrapie, we developed a neuropathological and gene expression study of sheep infected with scrapie in a presymptomatic stage. The histopathology, immunolabelling of PrP(Sc), Bax and activated caspase-3, and the analysis of the expression of 7 genes involved in the regulation of the mitochondrial pathway of apoptosis were investigated in the following 4 central nervous system areas: medulla oblongata, diencephalon, frontal cortex and cerebellum. Moreover, TUNEL and NeuN immunolabelling was performed in the medulla oblongata. The PrP(Sc) immunolabelling in the four areas, as well as a neuropil spongiform change, were more evident in the terminal stage than in presymptomatic animals. Cytoplasmic Bax immunostaining was observed in the presymptomatic medulla oblongata. In contrast to symptomatic animals, the immunostaining was not extended to the hypothalamus, indicating the progression of Bax induction during the course of the disease. Although neither caspase-3 immunostaining nor the TUNEL technique detected neurons with apoptosis, NeuN-immunolabelled cell counting determined that presymptomatic animals have already suffered neuronal loss in a lower or equal degree than symptomatic animals. Finally, the gene expression profiles indicated that the mitochondrial pathway of apoptosis was activated with higher intensity in presymptomatic animals than in symptomatic sheep and confirmed the implication of genes such as BAX or AIF in the disease.

  15. [Optimization of in vitro culture conditions for human amniotic epithelial cells and expression of stem cell markers].

    PubMed

    Chen, You-Yi; Lu, Yan; Wang, Ke; Wang, Yan; Wu, Dong-Ying; Liu, Bin; Yang, Ying; Lü, Shuang-Hong

    2011-04-01

    This study was purposed to optimize the culture conditions of the human amniotic epithelium cells (hAEC) in vitro, and detect the expression of hAEC pluripotent markers. Amnion tissues were separated from the underlying chorion through the spongy layer immediately after elective cesarean section of healthy pregnancy women at term. After the subsequent exposure to trypsin digestion, hAEC were cultured in DMEM with different supplements. The growth and proliferation potential of hAEC was evaluated, and the expression of cultured hAEC pluripotent markers was detected by using flow cytometry and immunohistochemistry methods. The results indicated that when being cultured in the mediums similar to that of embryonic stem cell culture supplemented with 10 ng/ml EGF, the hAEC grew better and the time for passage was shortened. In addition, compared to other culture conditions, under this condition, the cells could be passaged up to 5 times as much without obvious morphological changes, and the pluripotent marker SSEA-4 was detected in the cultured cells by flow cytometry. Meanwhile, the detection of immunofluorescence showed the expression of vimentin in cultured hAEC was strengthened as compared with primary cells. It is concluded that the culture condition similar to that for embryonic stem cells supplemented with EGF facilitates the proliferation and passage of hAEC in vitro.

  16. Association between Mutation and Expression of TP53 as a Potential Prognostic Marker of Triple-Negative Breast Cancer

    PubMed Central

    Kim, Ji-Yeon; Park, Kyunghee; Jung, Hae Hyun; Lee, Eunjin; Cho, Eun Yoon; Lee, Kwang Hee; Bae, Soo Youn; Lee, Se Kyung; Kim, Seok Won; Lee, Jeong Eon; Nam, Seok Jin; Ahn, Jin Seok; Im, Young-Hyuck; Park, Yeon Hee

    2016-01-01

    Purpose TP53, the most frequently mutated gene in breast cancer, is more frequently altered in HER2-enriched and basal-like breast cancer. However, no studies have clarified the role of TP53 status as a prognostic and predictive marker of triple-negative breast cancer (TNBC). Materials and Methods We performed p53 immunohistochemistry (IHC), nCounter mRNA expression assay, and DNA sequencing to determine the relationship between TP53 alteration and clinical outcomes of TNBC patients. Results Seventy-seven of 174 TNBC patients were found to harbor a TP53 mutation. Patients with missense mutations showed high protein expression in contrast to patients with deletion mutations (positivity of IHC: wild type vs. missense vs. deletion mutation, 53.6% vs. 89.8% vs. 25.0%, respectively; p < 0.001). TP53 mRNA expression was influenced by mutation status (mRNA expression [median]: wild type vs. missense vs. deletion mutation, 207.36± 132.73 vs. 339.61±143.21 vs. 99.53±99.57, respectively; p < 0.001). According to survival analysis, neither class of mutation nor protein or mRNA expression status had any impact on patient prognosis. In subgroup analysis, low mRNA expression was associated with poor prognosis in patients with a TP53 missense mutation (5-year distant recurrence-free survival [5Y DRFS]: low vs. high, 50.0% vs. 87.8%; p=0.009), while high mRNA expression with a TP53 deletion mutation indicated poor prognosis (5Y DRFS: low vs. high, 91.7% vs. 75.0%; p=0.316). Conclusion Association between TP53 mutation and expression indicates a potential prognostic marker of TNBC; hence both DNA sequencing and mRNA expression analysis may be required to predict the prognosis of TNBC patients. PMID:26910472

  17. Gene expression profiling of metaplastic lineages identifies CDH17 as a prognostic marker in early-stage gastric cancer

    PubMed Central

    Lee, Hyuk-Joon; Nam, Ki Taek; Park, Heae Surng; Kim, Min A; LaFleur, Bonnie J.; Aburatani, Hiroyuki; Yang, Han-Kwang; Kim, Woo Ho; Goldenring, James R.

    2010-01-01

    Background & Aims Intestinal metaplasia (IM) and spasmolytic polypeptide-expressing metaplasia (SPEM) are precursors to gastric carcinogenesis. We sought to identify molecular biomarkers of gastric metaplasias and gastric cancer by gene expression profiling of metaplastic lesions from patients. Methods cDNA microarray analysis was performed on IM and SPEM cells isolated from patient samples using laser capture microdissection. Up-regulated transcripts in metaplstic lesions were confirmed by immunostaining analysis in IM, SPEM, and gastric cancer tissues. Proteins that were highly expressed specifically in gastric cancer tissues were analyzed for their association with survival in a test set (n=450) and a validation set (n=502) of samples from gastric cancer patients. Results Compared to normal chief cells, 858 genes were differentially expressed in IM or SPEM samples. Immunostaining was detected for 12 proteins, including 3 new markers of IM (ACE2, LGALS4, AKR1B10) and 3 of SPEM (OLFM4, LYZ, DPCR1). Of 13 proteins expressed in IM or SPEM, 8 were expressed by 17%–50% of human gastric cancer tissues (MUC13, OLFM4, CDH17, KRT20, MUC5AC, LGALS4, AKR1B10, REG4). Expression of CDH17 or MUC13 correlated with patient survival in the test and a validation sets. Multivariate analysis showed that CDH17 was an independent prognostic factor in patients with stage I or node-negative disease. Conclusion We identified several novel biomarkers for IM, SPEM, and gastric cancer using gene expression profiling of human metaplastic lesions. Expression of CDH17 and MUC13 was upregulated in gastric cancer tissues. CDH17 is a promising prognostic marker for early-stage gastric cancer. PMID:20398667

  18. Impact of physical activity, cardiorespiratory fitness, and exercise training on markers of inflammation.

    PubMed

    Lavie, Carl J; Church, Timothy S; Milani, Richard V; Earnest, Conrad P

    2011-01-01

    Physical activity and exercise training (ET) enhance overall cardiorespiratory fitness (ie, fitness), thus producing many benefits in the primary and secondary prevention of cardiovascular diseases. Substantial evidence also indicates that acute and chronic inflammation is involved in the development and progression of atherosclerosis and major cardiovascular events. The most commonly utilized marker of inflammation is C-reactive protein (CRP). In this review, we discuss the importance of inflammation, especially CRP, as a cardiovascular risk marker by reviewing an abundant cross-sectional and clinical intervention literature providing evidence that physical activity, enhanced fitness, and ET are inversely associated with CRP and that being overweight or obese is directly related with inflammation/CRP. Although we discuss the controversy regarding whether or not ET reduces CRP independent of weight loss, clearly physical activity, improved fitness, and ET are associated with reductions in inflammation and overall cardiovascular risk in both primary and secondary prevention.

  19. Imaging of moving fiducial markers during radiotherapy using a fast, efficient active pixel sensor based EPID

    SciTech Connect

    Osmond, John P. F.; Zin, Hafiz M.; Harris, Emma J.; Lupica, Giovanni; Allinson, Nigel M.; Evans, Philip M.

    2011-11-15

    Purpose: The purpose of this work was to investigate the use of an experimental complementary metal-oxide-semiconductor (CMOS) active pixel sensor (APS) for tracking of moving fiducial markers during radiotherapy. Methods: The APS has an active area of 5.4 x 5.4 cm and maximum full frame read-out rate of 20 frame s{sup -1}, with the option to read out a region-of-interest (ROI) at an increased rate. It was coupled to a 4 mm thick ZnWO4 scintillator which provided a quantum efficiency (QE) of 8% for a 6 MV x-ray treatment beam. The APS was compared with a standard iViewGT flat panel amorphous Silicon (a-Si) electronic portal imaging device (EPID), with a QE of 0.34% and a frame-rate of 2.5 frame s{sup -1}. To investigate the ability of the two systems to image markers, four gold cylinders of length 8 mm and diameter 0.8, 1.2, 1.6, and 2 mm were placed on a motion-platform. Images of the stationary markers were acquired using the APS at a frame-rate of 20 frame s{sup -1}, and a dose-rate of 143 MU min{sup -1} to avoid saturation. EPID images were acquired at the maximum frame-rate of 2.5 frame s{sup -1}, and a reduced dose-rate of 19 MU min{sup -1} to provide a similar dose per frame to the APS. Signal-to-noise ratio (SNR) of the background signal and contrast-to-noise ratio (CNR) of the marker signal relative to the background were evaluated for both imagers at doses of 0.125 to 2 MU. Results: Image quality and marker visibility was found to be greater in the APS with SNR {approx}5 times greater than in the EPID and CNR up to an order of magnitude greater for all four markers. To investigate the ability to image and track moving markers the motion-platform was moved to simulate a breathing cycle with period 6 s, amplitude 20 mm and maximum speed 13.2 mm s{sup -1}. At the minimum integration time of 50 ms a tracking algorithm applied to the APS data found all four markers with a success rate of {>=}92% and positional error {<=}90 {mu}m. At an integration time of 400

  20. Generation and evaluation of a chimeric classical swine fever virus expressing a visible marker gene.

    PubMed

    Li, Yongfeng; Wang, Xiao; Sun, Yuan; Li, Lian-Feng; Zhang, Lingkai; Li, Su; Luo, Yuzi; Qiu, Hua-Ji

    2016-03-01

    Classical swine fever virus (CSFV) is a noncytopathogenic virus, and the incorporation of an enhanced green fluorescent protein (EGFP) tag into the viral genome provides a means of direct monitoring of viral infection without immunostaining. It is well established that the 3' untranslated region (3'-UTR) of the CSFV plays an important role in viral RNA replication. Although CSFV carrying a reporter gene and chimeric CSFV have been generated and evaluated, a chimeric CSFV with a visible marker has not yet been reported. Here, we generated and evaluated a chimeric virus containing the EGFP tag and the 3'-UTR from vaccine strain HCLV (C-strain) in the genetic background of the highly virulent CSFV Shimen strain. The chimeric marker CSFV was fluorescent and had an approximately 100-fold lower viral titer, lower replication level of viral genome, and weaker fluorescence intensity than the recombinant CSFV with only the EGFP tag or the parental virus. Furthermore, the marker chimera was avirulent and displayed no viremia in inoculated pigs, which were completely protected from lethal CSFV challenge as early as 15 days post-inoculation. The chimeric marker virus was visible in vitro and attenuated in vitro and in vivo, which suggests that CSFV can be engineered to produce attenuated variants with a visible marker to facilitate in vitro studies of CSFV infection and replication and to develop of novel vaccines against CSF. PMID:26614259

  1. Vitamin D compounds reduce mammosphere formation and decrease expression of putative stem cell markers in breast cancer.

    PubMed

    Wahler, Joseph; So, Jae Young; Cheng, Larry C; Maehr, Hubert; Uskokovic, Milan; Suh, Nanjoo

    2015-04-01

    Breast cancer stem cells (BCSCs) are a subset of tumor cells that are believed to be the cells responsible for the establishment and maintenance of tumors. Moreover, BCSCs are suggested to be the main cause of progression to metastasis and recurrence of cancer because of their tumor-initiating abilities and resistance to conventional therapies. Ductal carcinoma in situ (DCIS) is an early precursor in breast carcinogenesis which progresses to invasive ductal carcinoma (IDC). We have previously reported that a vitamin D compound, BXL0124, inhibits the progression of DCIS to IDC. In the present study we sought to determine whether this effect was mediated through an influence on BCSCs. In MCF10DCIS cells treated with vitamin D compounds (1α25(OH)2D3 or BXL0124), the breast cancer stem cell-like population, identified by the CD44(+)/CD24(-/low) and CD49f(+)/CD24(-/low) subpopulations, was reduced. To determine the effects of vitamin D compounds on cancer stem cell activity, the MCF10DCIS mammosphere cell culture system, which enriches for mammary progenitor cells and putative BCSCs, was utilized. Untreated MCF10DCIS mammospheres showed a disorganized and irregular shape. When MCF10DCIS cells were treated with 1α25(OH)2D3 or BXL0124, the mammospheres that formed exhibited a more organized, symmetrical and circular shape, similar to the appearance of spheres formed by the non-malignant, normal mammary epithelial cell line, MCF10A. The mammosphere forming efficiency (MFE) was significantly decreased upon treatment with 1α25(OH)2D3 or BXL0124, indicating that these compounds have an inhibitory effect on mammosphere development. Treatment with 1α25(OH)2D3 or BXL0124 repressed markers associated with the stem cell-like phenotype, such as CD44, CD49f, c-Notch1, and pNFκB. Furthermore, 1α25(OH)2D3 and BXL0124 reduced the expression of pluripotency markers, OCT4 and KLF-4 in mammospheres. This study suggests that vitamin D compounds repress the breast cancer stem cell

  2. Polyphenol Oxidase Activity Expression in Ralstonia solanacearum

    PubMed Central

    Hernández-Romero, Diana; Solano, Francisco; Sanchez-Amat, Antonio

    2005-01-01

    Sequencing of the genome of Ralstonia solanacearum revealed several genes that putatively code for polyphenol oxidases (PPOs). To study the actual expression of these genes, we looked for and detected all kinds of PPO activities, including laccase, cresolase, and catechol oxidase activities, in cellular extracts of this microorganism. The conditions for the PPO assays were optimized for the phenolic substrate, pH, and sodium dodecyl sulfate concentration used. It was demonstrated that three different PPOs are expressed. The genes coding for the enzymes were unambiguously correlated with the enzymatic activities detected by generation of null mutations in the genes by using insertional mutagenesis with a suicide plasmid and estimating the changes in the levels of enzymatic activities compared to the levels in the wild-type strain. The protein encoded by the RSp1530 locus is a multicopper protein with laccase activity. Two other genes, RSc0337 and RSc1501, code for nonblue copper proteins exhibiting homology to tyrosinases. The product of RSc0337 has strong tyrosine hydroxylase activity, and it has been shown that this enzyme is involved in melanin synthesis by R. solanacearum. The product of the RSc1501 gene is an enzyme that shows a clear preference for oxidation of o-diphenols. Preliminary characterization of the mutants obtained indicated that PPOs expressed by R. solanacearum may participate in resistance to phenolic compounds since the mutants exhibited higher sensitivity to l-tyrosine than the wild-type strain. These results suggest a possible role in the pathogenic process to avoid plant resistance mechanisms involving the participation of phenolic compounds. PMID:16269713

  3. Sequential hepatogenic transdifferentiation of adipose tissue-derived stem cells: relevance of different extracellular signaling molecules, transcription factors involved, and expression of new key marker genes.

    PubMed

    Bonora-Centelles, A; Jover, R; Mirabet, V; Lahoz, A; Carbonell, F; Castell, J V; Gómez-Lechón, M J

    2009-01-01

    Adipose tissue contains a mesenchymal stem cell (MSC) population known as adipose-derived stem cells (ASCs) capable of differentiating into different cell types. Our aim was to induce hepatic transdifferentiation of ASCs by sequential exposure to several combinations of cytokines, growth factors, and hormones. The most efficient hepatogenic protocol includes fibroblastic growth factors (FGF) 2 and 4 and epidermal growth factor (EGF) (step 1), hepatocyte growth factor (HGF), FGF2, FGF4, and nicotinamide (Nic) (step 2), and oncostatin M (OSM), dexamethasone (Dex), and insulin-tranferrin-selenium (step 3). This protocol activated transcription factors [GATA6, Hex, CCAAT/enhancer binding protein alpha and beta (CEBPalpha and beta), peroxisome proliferator-activated receptor-gamma, coactivator 1 alpha (PGC1alpha), and hepatocyte nuclear factor 4 alpha (HNF4alpha)], which promoted a characteristic hepatic phenotype, as assessed by new informative markers for the step-by-step hepatic transdifferentiation of hMSC [early markers: albumin (ALB), alpha-2-macroglobuline (alpha2M), complement protein C3 (C3), and selenoprotein P1 (SEPP1); late markers: cytochrome P450 3A4 (CYP3A4), apolipoprotein E (APOE), acyl-CoA synthetase long-chain family member 1 (ACSL1), and angiotensin II receptor, type 1 (AGTR1)]. The loss of adipose adult stem cell phenotype was detected by losing expression of Thy1 and inhibitor of DNA binding 3 (Id3). The reexpression of phosphoenolpyruvate corboxykinase (PEPCK), apolipoprotein C3 (APOCIII), aldolase B (ALDOB), and cytochrome P450 1A2 (CYP1A2) was achieved by transduction with a recombinant adenovirus for HNF4alpha and finally hepatic functionality was also assessed by analyzing specific biochemical markers. We conclude that ASCs could represent an alternative tool in clinical therapy for liver dysfunction and regenerative medicine.

  4. Association of expression levels of pluripotency/stem cell markers with the differentiation outcome of Wharton's jelly mesenchymal stem cells into insulin producing cells.

    PubMed

    Kassem, Dina H; Kamal, Mohamed M; El-Kholy, Abd El-Latif G; El-Mesallamy, Hala O

    2016-08-01

    Recently, there has been much attention towards generation of insulin producing cells (IPCs) from stem cells, especially from Wharton's jelly mesenchymal stem cells (WJ-MSCs). However, generation of mature IPCs remains a challenge. Assessment of generation of IPCs was usually done by examining β-cell markers, however, assessment of pluripotency/stem cell markers drew less attention. Therefore, the purpose of this study was to investigate the levels of pluripotency/stem cell markers during differentiation of WJ-MSCs into IPCs and the association of these levels with differentiation outcomes. WJ-MSCs were isolated, characterized then induced to differentiate into IPCs using three different protocols namely A, B and C. Differentiated IPCs were assessed by the expression of pluripotency/stem cell markers, together with β-cell markers using qRT-PCR, and functionally by measuring glucose stimulated insulin secretion. Differentiated cells from protocol A showed lowest expression of pluripotency/stem cell markers and relatively best GSIS. However, protocol B showed concomitant expression of pluripotency/stem cell and β-cell markers with relatively less insulin secretion as compared to protocol A. Protocol C failed to generate glucose-responsive IPCs. In conclusion, sustained expression of pluripotency/stem cell markers could be associated with the incomplete differentiation of WJ-MSCs into IPCs. A novel finding for which further investigations are warranted.

  5. PPARy and GLUT-4 expression as developmental regulators/markers for preadipocyte differentiation into an adipocyte

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this document, we have integrated knowledge about two major cellular markers found in cells of the adipocyte lineage. The first factor is PPARy, which has been identified as an important adipogenic regulator. PPARy plays an important role in converting adipofibroblasts, fibroblasts or preadipocyt...

  6. GLOBAL EXPRESSION PROFILING AS A ROOL TO DEVELOP MOLECULAR MARKERS LINKED TO HERBICIDE STRESS IN ARABIDOPSIS

    EPA Science Inventory

    Herbicide drift (unintentional physical movement from target to off-target plants) is a cause of crop loss in US. Low-dose, high-potency herbicides that have short environmental persistence times constrain efforts to develop or identify metabolite or biochemical markers of exposu...

  7. Markers for Persistent Specific Expressive Language Delay in 3-4-Year-Olds

    ERIC Educational Resources Information Center

    Everitt, Andrea; Hannaford, Philip; Conti-Ramsden, Gina

    2013-01-01

    Background: Identifying 3-4-year-olds who are most at risk of persisting language difficulties, and possibly specific language impairment (SLI), is difficult due to the natural variation of language in young children. In older children, markers for SLI have been identified that differentiate between children with and without SLI. It is not known…

  8. The role of the graft endothelium in transplant rejection: evidence that endothelial activation may serve as a clinical marker for the development of chronic rejection.

    PubMed

    Denton, M D; Davis, S F; Baum, M A; Melter, M; Reinders, M E; Exeni, A; Samsonov, D V; Fang, J; Ganz, P; Briscoe, D M

    2000-11-01

    In this review, we discuss the role of the allograft endothelium in the recruitment and activation of leukocytes during acute and chronic rejection. We discuss associations among endothelial activation responses, the expression of adhesion molecules, chemokines and chemokine receptors, and rejection; and we propose that endothelial vascular cellular adhesion molecule-1 (VCAM-1) may be used as a surrogate marker of acute rejection and allograft vasculopathy. In addition, we describe potential mechanistic interpretations of persistent endothelial cell (EC) expression of major histocompatibility complex (MHC) class II molecules in allorecognition. The graft endothelium may provide an antigen-specific signal to transmigrating, previously activated, T cells and may induce B7 expression on locally transmigrating leukocytes to promote costimulation. Taken together, these functions of the EC provide it with a potent regulatory role in rejection and in the maintenance of T-cell activation via the direct and/or the indirect pathways of allorecognition.

  9. Development, characterization and cross species amplification of polymorphic microsatellite markers from expressed sequence tags of turmeric (Curcuma longa L.).

    PubMed

    Siju, S; Dhanya, K; Syamkumar, S; Sasikumar, B; Sheeja, T E; Bhat, A I; Parthasarathy, V A

    2010-02-01

    Expressed sequence tags (ESTs) from turmeric (Curcuma longa L.) were used for the screening of type and frequency of Class I (hypervariable) simple sequence repeats (SSRs). A total of 231 microsatellite repeats were detected from 12,593 EST sequences of turmeric after redundancy elimination. The average density of Class I SSRs accounts to one SSR per 17.96 kb of EST. Mononucleotides were the most abundant class of microsatellite repeat in turmeric ESTs followed by trinucleotides. A robust set of 17 polymorphic EST-SSRs were developed and used for evaluating 20 turmeric accessions. The number of alleles detected ranged from 3 to 8 per loci. The developed markers were also evaluated in 13 related species of C. longa confirming high rate (100%) of cross species transferability. The polymorphic microsatellite markers generated from this study could be used for genetic diversity analysis and resolving the taxonomic confusion prevailing in the genus.

  10. Eosinophils in the blood of hematopoietic stem cell transplanted patients are activated and have different molecular marker profiles in acute and chronic graft-versus-host disease

    PubMed Central

    Cromvik, Julia; Johnsson, Marianne; Vaht, Krista; Johansson, Jan-Erik; Wennerås, Christine

    2014-01-01

    While increased numbers of eosinophils may be detected in patients with graft-versus-host disease (GVHD) following hematopoietic stem cell transplantation, it is not known if eosinophils play a role in GVHD. The aims of this study were to determine: whether eosinophils are activated during GVHD; whether the patterns of activation are similar in acute and chronic GVHD; and the ways in which systemic corticosteroids affect eosinophils. Transplanted patients (n = 35) were investigated for eosinophil numbers and the expression levels of 16 eosinophilic cell surface markers using flow cytometry; all the eosinophil data were analyzed by the multivariate method OPLS-DA. Different patterns of molecule expression were observed on the eosinophils from patients with acute, chronic, and no GVHD, respectively. The molecules that provided the best discrimination between acute and chronic GVHD were: the activation marker CD9; adhesion molecules CD11c and CD18; chemokine receptor CCR3; and prostaglandin receptor CRTH2. Patients with acute or chronic GVHD who received systemic corticosteroid treatment showed down-regulation of the cell surface markers on their eosinophils, whereas corticosteroid treatment had no effect on the eosinophil phenotype in the patients without GVHD. In summary, eosinophils are activated in GVHD, display different activation profiles in acute and chronic GVHD, and are highly responsive to systemic corticosteroids. PMID:25400930

  11. Mean platelet volume as an inflammation marker in active pulmonary tuberculosis

    PubMed Central

    2014-01-01

    Background The mean platelet volume (MPV) reflects the size of platelets. It has been shown to be inversely correlated with level of the inflammation in some chronic inflammatory diseases. This prospective study aims to show the usability of MPV as an inflammation marker in patients with active pulmonary tuberculosis (PTB) by comparison with healthy controls. In addition, its relationships with other inflammatory markers such as C-reactive protein (CRP) and the erythrocyte sedimentation rate (ESR) as well as with the radiological extent of disease were examined. Methods This study included 82 patients with active PTB and 95 healthy subjects (control group). Whole blood counts, CRP level, and ESR were compared between the two groups. In the PTB group, the relationships between the radiological extent of disease and the MPV and other inflammation markers were investigated. Results The MPV was 7.74 ± 1.33/μL in the PTB group and 8.20 ± 1.13/μL in the control group (p = 0.005). The blood platelet count, CRP level, and ESR were significantly higher in the active PTB group than in the control group (p < 0.0001). In the PTB group, CRP levels (r = 0.26, p = 0.003) and ESR (r = 0.39, p = 0.003), but not MPV (p = 0.80), were significantly correlated with the radiologic extent of the disease. Conclusions The MPV was lower in patients with PTB than in healthy controls, however, the difference was limited. The MPV does not reflect the severity of the disease. The use of MPV as an inflammation marker and a negative acute-phase reactant in PTB does not seem to be reliable. PMID:24581084

  12. Trefoil factor 3: a novel serum marker identified by gene expression profiling in high-grade endometrial carcinomas.

    PubMed

    Bignotti, E; Ravaggi, A; Tassi, R A; Calza, S; Rossi, E; Falchetti, M; Romani, C; Bandiera, E; Odicino, F E; Pecorelli, S; Santin, A D

    2008-09-01

    This study identifies the genetic fingerprint of poorly differentiated endometrioid endometrial carcinomas (G3-EEC) and analyses the potential utility of trefoil factor 3 (TFF3) as novel serum marker in G3-EEC. Affymetrix microarrays were used to identify the gene expression patterns of 19 snap-frozen G3-EEC and 15 normal endometrium (NE) biopsies. Quantitative real-time PCR (qRT-PCR) and immunohistochemistry were used to validate TFF3 expression. Finally, TFF3 serum levels were determined by ELISA in 25 G3-EEC patients, 42 healthy controls, and in 13 endometrial hyperplasia patients. Hierarchical cluster analysis showed TFF3 as the top differentially expressed gene between 363 upregulated genes in G3-EEC, when compared with NE. Trefoil factor 3 gene expression levels analysed by qRT-PCR significantly correlated with Affymetrix results (P<0.001; rs=0.85). By immunohistochemistry, TFF3 protein was significatively more expressed in EEC compared with NE (P<0.01), with cytoplasmatic positivity in 79% G3-EEC and 18% NE. Patients harbouring G3-EECs had significantly higher TFF3 serum concentration by ELISA when compared with healthy patients (P<0.001) or patients harbouring endometrial hyperplasia (P=0.012). In conclusion, TFF3 is highly expressed at gene and protein level in G3-EEC. Further investigations on a wider set of samples are warranted to validate TFF3 as a novel serum marker for early detection and/or monitoring of G3-EEC patients.

  13. Spectroscopic detection of fluorescent protein marker gene activity in genetically modified plants

    NASA Astrophysics Data System (ADS)

    Liew, O. W.; Chong, Jenny P. C.; Asundi, Anand K.

    2005-04-01

    This work focuses on developing a portable fibre optic fluorescence analyser for rapid identification of genetically modified plants tagged with a fluorescent marker gene. Independent transgenic tobacco plant lines expressing the enhanced green fluorescence protein (EGFP) gene were regenerated following Agrobacterium-mediated gene transfer. Molecular characterisation of these plant lines was carried out at the DNA level by PCR screening to confirm their transgenic status. Conventional transgene expression analysis was then carried out at the RNA level by RT-PCR and at the protein level by Western blotting using anti-GFP rabbit antiserum. The amount of plant-expressed EGFP on a Western blot was quantified against known amounts of purified EGFP by scanning densitometry. The expression level of EGFP in transformed plants was found to range from 0.1 - 0.6% of total extractable protein. A comparison between conventional western analysis of transformants and direct spectroscopic quantification using the fibre optic fluorescence analyser was made. The results showed that spectroscopic measurements of fluorescence emission from strong EGFP expressors correlated positively with Western blot data. However, the fluorescence analyser was also able to identify weakly expressing plant transformants below the detection limit of colorimetric Western blotting.

  14. Gene expression profiling of serrated polyps identifies annexin A10 as a marker of a sessile serrated adenoma/polyp.

    PubMed

    Gonzalo, David Hernandez; Lai, Keith K; Shadrach, Bonnie; Goldblum, John R; Bennett, Ana E; Downs-Kelly, Erinn; Liu, Xiuli; Henricks, Walter; Patil, Deepa T; Carver, Paula; Na, Jie; Gopalan, Banu; Rybicki, Lisa; Pai, Rish K

    2013-08-01

    Sessile serrated adenomas/polyps (SSA/Ps) are precursors of colon cancer, particularly those that exhibit microsatellite instability. Distinguishing SSA/Ps from the related, but innocuous, microvesicular hyperplastic polyp (MVHP) can be challenging. In this study seven gastrointestinal pathologists reviewed 109 serrated polyps and identified 60 polyps with histological consensus. Microarray analysis was performed on six distal consensus MVHPs < 9 mm, six proximal consensus SSA/Ps > 9 mm, and six normal colon biopsies (three proximal, three distal). Comparative gene expression analysis confirmed the close relationship between SSA/Ps and MVHPs as there was overlapping expression of many genes. However, the gene expression profile in SSA/Ps had stronger and more numerous associations with cancer-related genes compared with MVHPs. Three genes (TFF2, FABP6, and ANXA10) were identified as candidates whose expression can differentiate SSA/Ps from MVHPs, and the differences in expression were confirmed by quantitative RT-PCR. As ANXA10 showed the most promise in differentiating these polyps, the expression of ANXA10 was evaluated by immunohistochemistry in consensus SSA/Ps (n = 26), MVHPs (n = 21), and normal colon (n = 9). Immunohistochemical expression of ANXA10 was not identified in separate samples of normal colon or in the normal colonic epithelium adjacent to the serrated polyps. Consistent with the microarray and quantitative RT-PCR experiments, immunohistochemical expression of ANXA10 was markedly increased in SSA/Ps compared to MVHPs (p < 0.0001). An ANXA10 score ≥ 3 has a sensitivity of 73% and a specificity of 95% in the diagnosis of an SSA/P. In conclusion, we show that SSA/Ps and MVHPs have significant overlap in gene expression, but also important differences, particularly in cancer-related pathways. Expression of ANXA10 may be a potential marker of the serrated pathway to colon cancer.

  15. Transcriptional expression levels and biochemical markers of oxidative stress in the earthworm Eisenia andrei after exposure to 2,4-dichlorophenoxyacetic acid (2,4-D).

    PubMed

    Hattab, Sabrine; Boughattas, Iteb; Boussetta, Hamadi; Viarengo, Aldo; Banni, Mohamed; Sforzini, Susanna

    2015-12-01

    This study investigated the stress response of earthworms (Eisenia andrei) to exposure to a commonly used herbicide, 2,4 dichloro-phenoxy-acetic acid (2,4-D). We evaluated both stress biomarkers and the transcriptional expression levels and activity of three enzymes involved in oxidative stress responses. Earthworms were exposed to three sublethal concentration of 2,4-D (3.5, 7, and 14 mg kg(-1)) for 7 and 14 days. Exposure to 7 and 14 mg kg(-1) 2,4-D significantly reduced both worm body weight and lysosomal membrane stability (LMS); the latter is a sensitive stress biomarker in coelomocytes. Exposure to 2,4-D caused a pronounced increase in the accumulation of malonedialdehyde (MDA), a marker of oxidative stress, and significantly increased the activity of the antioxidant enzymes catalase (CAT), superoxide dismutase (SOD),and glutathione-S-transferase (GST). Compared to expression in controls, the expression levels of the sod, cat, and gst genes increased in worms exposed to all three 2,4-D doses for 7 days. However, after 14 days of exposure, only the expression of the gst gene remained higher than controls. These data provide new insights into the cytotoxicity of 2,4-D in the earthworm E. andrei and should be carefully considered in view of the biological effects of herbicides in soils organisms.

  16. Mesoderm patterning and morphogenesis in the polychaete Alitta virens (Spiralia, Annelida): Expression of mesodermal markers Twist, Mox, Evx and functional role for MAP kinase signaling.

    PubMed

    Kozin, Vitaly V; Filimonova, Daria A; Kupriashova, Ekaterina E; Kostyuchenko, Roman P

    2016-05-01

    Mesoderm represents the evolutionary youngest germ layer and forms numerous novel tissues in bilaterian animals. Despite the established conservation of the gene regulatory networks that drive mesoderm differentiation (e.g. myogenesis), mechanisms of mesoderm specification are highly variable in distant model species. Thus, broader phylogenetic sampling is required to reveal common features of mesoderm formation across bilaterians. Here we focus on a representative of Spiralia, the marine annelid Alitta virens, whose mesoderm development is still poorly investigated on the molecular level. We characterize three novel early mesodermal markers for A. virens - Twist, Mox, and Evx - which are differentially expressed within the mesodermal lineages. The Twist mRNA is ubiquitously distributed in the fertilized egg and exhibits specific expression in endomesodermal- and ectomesodermal-founder cells at gastrulation. Twist is expressed around the blastopore and later in a segmental metameric pattern. We consider this expression to be ancestral, and in support of the enterocoelic hypothesis of mesoderm evolution. We also revealed an early pattern of the MAPK activation in A. virens that is different from the previously reported pattern in spiralians. Inhibition of the MAPK pathway by U0126 disrupts the metameric Twist and Mox expression, indicating an early requirement of the MAPK cascade for proper morphogenesis of endomesodermal tissues. PMID:27000638

  17. BLV-infected lymphocytes exhibit two patterns of expression as determined by Ig and CD5 markers.

    PubMed

    Meirom, R; Brenner, J; Trainin, Z

    1993-03-01

    Lymphocytes were defined by their cell surface markers, Ig and CD5 in three groups of cows naturally infected with bovine leucosis virus (BLV). Lymphocytes were enumerated and groups were designated BLV seropositive with persistent lymphocytosis (BLV + PL +), BLV seropositive without persistent lymphocytosis (BLV + PL-) and BLV negative. The competence of peripheral blood mononuclear cells (PBMC) from the tested cows to express these two markers was determined by the double staining immunofluorescence procedure. Cows which developed persistent lymphocytosis (PL) as a result of BLV infection consequently underwent massive proliferation of B lymphocytes which express both Ig and CD5 antigens. In contrast, cows which were defined as BLV positive and PL negative showed a remarkable decrease of CD5 + Ig-, CD5- Ig+ and CD5+ Ig+ cells and also in the total number of lymphocytes. We suggest that BLV infection affects bovine lymphocytes through two different pathways of expression which might be related to the genetic properties of the target cells. PMID:7682745

  18. Characterization and Expression of Senescence Marker in Prolonged Passages of Rat Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Ridzuan, Noridzzaida; Al Abbar, Akram; Yip, Wai Kien; Maqbool, Maryam

    2016-01-01

    The present study is aimed at optimizing the in vitro culture protocol for generation of rat bone marrow- (BM-) derived mesenchymal stem cells (MSCs) and characterizing the culture-mediated cellular senescence. The initial phase of generation and characterization was conducted using the adherent cells from Sprague Dawley (SD) rat's BM via morphological analysis, growth kinetics, colony forming unit capacity, immunophenotyping, and mesodermal lineage differentiation. Mesenchymal stem cells were successfully generated and characterized as delineated by the expressions of CD90.1, CD44H, CD29, and CD71 and lack of CD11b/c and CD45 markers. Upon induction, rBM-MSCs differentiated into osteocytes and adipocytes and expressed osteocytes and adipocytes genes. However, a decline in cell growth was observed at passage 4 onwards and it was further deciphered through apoptosis, cell cycle, and senescence assays. Despite the enhanced cell viability at later passages (P4-5), the expression of senescence marker, β-galactosidase, was significantly increased at passage 5. Furthermore, the cell cycle analysis has confirmed the in vitro culture-mediated cellular senescence where cells were arrested at the G0/G1 phase of cell cycle. Although the currently optimized protocols had successfully yielded rBM-MSCs, the culture-mediated cellular senescence limits the growth of rBM-MSCs and its potential use in rat-based MSC research. PMID:27579045

  19. Enhanced expression of stem cell markers and drug resistance in sphere-forming non-small cell lung cancer cells

    PubMed Central

    Sun, Feng-Feng; Hu, Yong-He; Xiong, Lv-Ping; Tu, Xiao-Yun; Zhao, Ji-Hua; Chen, Sheng-Song; Song, Juan; Ye, Xiao-Qun

    2015-01-01

    There is growing evidence suggesting that cancer stem cells (CSCs) are playing critical roles in tumor progression, metastasis and drug resistance. However, the role of CSCs in non-small cell lung cancer (NSCLC) remains elusive. In this study, we enriched for stem-like cells from tumor spheres derived from NSCLC cell line A549 cultured in serum-free medium. Our results showed that sphere-derived cells expressed various stem cell markers such as CD44, CD133, Sox2 and Oct4. Compared with the corresponding cells in monolayer cultures, sphere-derived cells showed marked morphologic changes and increased expression of the stem cell markers CD133. Furthermore, we found that sphere-derived cells exhibited increased proliferation, cell-cycle progression as well as drug-resistant properties as compared to A549 adherent cells. Consistently, expression of several drug resistance proteins, including lung resistance-related protein (LRP), glutathion-S-transferase-π (GST-π) and multidrug resistance proteins-1 (MRP1) were all significantly enhanced in sphere-derived cells. These results indicate the enrichment of CSCs in sphere cultures and support their role in regulating drug resistance in NSCLC. PMID:26261505

  20. Relationship between physical activity and markers of oxidative stress in independent community-living elderly individuals.

    PubMed

    Fraile-Bermúdez, A B; Kortajarena, M; Zarrazquin, I; Maquibar, A; Yanguas, J J; Sánchez-Fernández, C E; Gil, J; Irazusta, A; Ruiz-Litago, F

    2015-10-01

    The aim of the present study was to examine the relationship between objective data of physical activity and markers of oxidative stress in older men and women. Participants were old adults, aged≥60years (61 women and 34 men) who were all capable of performing basic daily activities by themselves and lived on their own. To describe physical activity we used objective data measured by accelerometers which record active and sedentary periods during everyday life for five days. Determination of oxidative stress was conducted from three perspectives: determination plasma total antioxidant status (TAS), plasma antioxidant enzyme activities, i.e., glutathione peroxidase (GPx), catalase (CAT) and superoxide dismutase (SOD), and membrane lipid peroxidation (TBARS). In the group of women, those who met physical activity recommendations (WR) had lower level of TAS. In addition, the moderate to vigorous physical activity (MVPA) was negatively correlated with TAS. Simultaneously, MVPA was correlated with increase in the GPx antioxidant enzyme activity, and the counts per minute were positively correlated with CAT activity. In the group of men, the cpm and the MVPA were negatively correlated with lipid peroxidation while lifestyle physical activity was positively correlated with CAT activity. These findings suggest that MVPA in the elderly although it is related to a decrease in the TAS in women, induces adaptive increase in antioxidant enzyme activity and decreases lipid peroxidation in both women and men. These results suggest that at this time of life, it is not only the amount of physical activity performed that is important but also its intensity.

  1. Vigna unguiculata modulates cholesterol induced cardiac markers, genotoxicity and gene expressions profile in an experimental rabbit model.

    PubMed

    Janeesh, P A; Abraham, Annie

    2013-04-25

    Vigna unguiculata (VU) leaves are edible and used as a leafy vegetable in cuisine from traditional times in India. This study was designed to investigate the cardioprotective effect of VU in cholesterol fed rabbits. The animals were randomly divided into 4 groups of 6 animals each and the experimental period was 3 months. Group I-ND [normal diet 40 g feed], Group II-ND + FVU [flavanoid fraction of Vigna unguiculata (150 mg kg (-1) per body weight)], Group III-ND + CH [cholesterol (400 mg)] and Group IV-ND + CH (400 mg) +FVU (150 mg kg(-1) per body weight). After the experimental period, animals were sacrificed and the various parameters, such as cardiac markers, toxicity parameters, genotoxicity and gene expression, were investigated. Cholesterol feeding causes a significant increase in the levels of cardiac marker enzymes, namely lactate dehydrogenase (LDH) and creatine phospokinase (CPK), atherogenic index, toxicity parameters like serum glutamate oxaloacetate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) were elevated. Antioxidant enzyme levels were decreased, lipid peroxidation products in heart tissue and inflammatory markers, namely cyclooxygenase (COX2) and lipooxygenase (LOX15) in peripheral blood monocytes (PBMCs), were significantly increased. A genotoxicity study using a Comet assay and gene expression by reverse transcriptase-polymerase chain reaction (RT-PCR) of transforming growth factor-b1 (TGF-b1) and heme oxygenase-1 (HO-1) from heart tissue showed an altered expression in the disease group. The supplementation of the flavonoid fraction of Vigna unguiculata leaves (FVU) in the CH + FVU group caused the reversal of the above parameters and cardiotoxicity to near normal when compared with the CH group and FVU. This study revealed the cardioprotective nature of Vigna unguiculata in preventing cardiovascular diseases and this effect is attributed to the presence of antioxidants and the antihyperlipidemic properties of the

  2. Expression of 20 alpha-hydroxysteroid dehydrogenase activity in human lymphoid and non lymphoid cells.

    PubMed Central

    Carbone, A; Piantelli, M; Musiani, P; Larocca, L M; Revoltella, R P; Ranelletti, F O

    1986-01-01

    Expression of 20-alpha-hydroxysteroid dehydrogenase (20 alpha-SDH), a putative T cell marker in the murine system, has been examined in human haematopoietic cells. Higher levels of enzymatic activity were expressed by human peripheral blood mononuclear cells (PBMC) in comparison with thymocytes. When PBMC were fractionated into T and non T cell subsets, the greatest values of 20 alpha-SDH activity were associated with the non T cell population. Cells from bone marrow exhibited lower levels of 20 alpha-SDH than PBMC and thymocytes. Both myeloid and lymphoid leukaemic cells expressed significant 20 alpha-SDH activity. In addition our data in U-937 and CM-S human cell lines indicate that cells belonging to the myelomonocytic precursor compartment constitutively expressed 20 alpha-SDH activity. Furthermore, this activity was less expressed when these cells were induced to monocyte-macrophage differentiation. In conclusion, our data in the human system indicate that 20 alpha-SDH should not be viewed as a lymphoid lineage-restricted marker enzyme. PMID:3485485

  3. Expression of Excision Repair Cross-Complementation Group 1 as Predictive Marker for Nasopharyngeal Cancer Treated With Concurrent Chemoradiotherapy

    SciTech Connect

    Sun, Jong-Mu; Ahn, Myung-Ju; Park, Min Jae; Lee, Hui-Young; Ahn, Jin Seok; Lee, Seungkoo; Kang, Gu; Han, Joungho; Son, Young-Ik; Baek, Chung-Hwan; Ahn, Yong Chan; Park, Keunchil

    2011-07-01

    Purpose: Cisplatin-based concurrent chemoradiotherapy is the standard treatment of nasopharyngeal cancer. The expression of excision repair cross-complementation group 1 (ERCC1) has been reported to be associated with resistance to platinum-based chemotherapy. We evaluated whether ERCC1 expression could predict the treatment response and survival outcome of patients with locally advanced nasopharyngeal cancer who were treated with cisplatin-based concurrent chemoradiotherapy. Methods and Materials: Immunohistochemistry was used to examine the expression of ERCC1 in nasopharyngeal tumor tissue. Patients were categorized into either a resistant or sensitive group depending on their treatment response outcome. A total of 77 patients were assessed in the present study. Results: The resistant and sensitive groups included 25 and 52 patients, respectively. ERCC1 expression was positive in the tumor tissue for 39 of the 77 patients (51%). Significantly more ERCC1-negative tumors were in the sensitive group than in the resistant group (p = .035). In terms of survival outcome, univariate analysis determined that patients with ERCC1-negative tumors had longer disease-free survival (p = .076) and overall survival (p = .013) than patients with ERCC1-positive tumors. Multivariate analysis determined that negative ERCC expression in tumors was an independent predictor for prolonged overall survival (hazard ratio, 0.14; 95% confidence interval, 0.03-0.71). Conclusion: These results suggest that ERCC1 expression might be a useful predictive marker in patients with locally advanced nasopharyngeal cancer who are under consideration for cisplatin-based concurrent chemoradiotherapy.

  4. Identification of histological markers for malignant glioma by genome-wide expression analysis: dynein, alpha-PIX and sorcin.

    PubMed

    Yokota, Takashi; Kouno, Jun; Adachi, Koji; Takahashi, Hiroshi; Teramoto, Akira; Matsumoto, Koshi; Sugisaki, Yuichi; Onda, Masamitsu; Tsunoda, Tatsuhiko

    2006-01-01

    Glioblastoma multiforme (GBM), the most malignant class of glial neoplasm (grade IV in WHO criteria), carries the worst clinical prognosis among primary brain tumors in adults. To identify a set of genes involved in the tumorigenesis of GBM, we evaluated expression profiles of GBM tissues from 11 patients using a cDNA microarray representing 25,344 human genes. By comparing the profiles with those of normal brain tissue, we identified a number of differentially expressed genes: 54 with increased expression and 45 with reduced expression in GBMs. Semi-quantitative RT-PCR experiments with 6 of those genes confirmed higher expression of DNCH2, ARHGEF6, NPM1 and SRI and lower expression of NRGN and TM4SF2 in GBM tumors. Immunohistochemical staining for 3 of the respective gene products, dynein (product of DNCH2), alpha-PIX (product of ARHGEF6), and sorcin (product of SRI) indicated that this technique might be useful for histological grading of glial tumors. To establish criteria for this diagnostic approach, we scored glial tumor tissues of different histological grades according to the staining results; the scores were significantly higher in anaplastic astrocytomas and GBMs than in diffuse astrocytomas or normal brain tissues. These findings indicated that levels of these three proteins might serve as histological markers for malignant glioma classification. PMID:16320026

  5. Prognostic and Predictive Value of Baseline and Posttreatment Molecular Marker Expression in Locally Advanced Rectal Cancer Treated With Neoadjuvant Chemoradiotherapy

    SciTech Connect

    Bertolini, Federica . E-mail: bertolini.federica@policlinico.mo.it; Bengala, Carmelo; Losi, Luisa; Pagano, Maria; Iachetta, Francesco; Dealis, Cristina; Jovic, Gordana; Depenni, Roberta; Zironi, Sandra; Falchi, Anna Maria; Luppi, Gabriele; Conte, Pier Franco

    2007-08-01

    Purpose: To evaluate expression of a panel of molecular markers, including p53, p21, MLH1, MSH2, MIB-1, thymidylate synthase, epidermal growth factor receptor (EGFR), and tissue vascular endothelial growth factor (VEGF), before and after treatment in patients treated with neoadjuvant chemoradiotherapy for locally advanced rectal cancer, to correlate the constitutive profile and dynamics of expression with pathologic response and outcome. Methods and Materials: Expression of biomarkers was evaluated by immunohistochemistry in tumor samples from 91 patients with clinical Stage II and III rectal cancer treated with preoperative pelvic radiotherapy (50 Gy) plus concurrent 5-fluorouracil by continuous intravenous infusion. Results: A pathologic complete remission was observed in 14 patients (15.4%). Patients with MLH1-positive tumors had a higher pathologic complete response rate (24.3% vs. 9.4%; p = 0.055). Low expression of constitutive p21, absence of EGFR expression after chemoradiotherapy, and high Dworak's tumor regression grade (TRG) were significantly associated with improved disease-free survival and overall survival. A high MIB-1 value after chemoradiotherapy was significantly associated with worse overall survival. Multivariate analysis confirmed the prognostic value of constitutive p21 expression as well as EGFR expression and MIB-1 value after chemoradiotherapy among patients not achieving TRG 3-4. Conclusions: In our study, we observed the independent prognostic value of EGFR expression after chemoradiotherapy on disease-free survival. Moreover, our study suggests that a constitutive high p21 expression and a high MIB-1 value after neoadjuvant chemoradiotherapy treatment could predict worse outcome in locally advanced rectal cancer.

  6. Effect of different degrees of impaired glucose metabolism on the expression of inflammatory markers in monocytes of patients with atherosclerosis.

    PubMed

    Bernal-Lopez, M R; Llorente-Cortes, V; Calleja, F; Lopez-Carmona, D; Mayas, M D; Gomez-Huelgas, R; Badimon, L; Tinahones, F J

    2013-08-01

    Inflammatory markers are elevated in type 2 diabetic patients (DP) and may predict the development of type 2 diabetes. Our aims were to analyze differences in the expression of inflammatory and immunological molecules between DP and healthy subjects and to investigate whether glycemic control might prevent the overexpression of inflammatory markers in DP. Twenty-two DP with advanced atherosclerosis and eight healthy blood donors were included. DP were classified as well (HbA1c ≤ 6.5) or poorly controlled (HbA1c > 6.5). In "in vitro" studies, monocytes were exposed to low (5.5 mM) or high glucose (26 mM) concentrations in the absence or presence of insulin. Expression profiling of 14 inflammatory genes was analyzed using TLDA analysis. "In vivo" results show that monocytes from DP had increased levels of monocyte chemoattractant protein (MCP-1) and interleukin 6 (IL6) and lower levels of Toll-like receptor 2 (TLR2) mRNA than healthy subjects. Well-controlled DP had lower levels of IL-6 than poorly controlled DP, suggesting that glycemic control may prevent IL6 mRNA alterations associated with diabetes. "In vitro" results demonstrate that glucose directly and significantly induced MCP-1 and IL6 and reduced TLR2 mRNA expression. Insulin at high dose (100 IU/ml) dramatically enhanced the upregulatory effects of glucose on MCP-1 and IL-6 and reduced per se TLR2 mRNA expression. MCP-1, IL-6 and TLR2 are key inflammatory players altered in monocytes from type 2 DP. Both hyperinsulinemia and hyperglycemia contribute to alter the expression of these genes. The glycemic control only significantly prevented IL6 overexpression in this group of patients.

  7. An integrated study of heart pain and behavior in freely moving rats (using fos as a marker for neuronal activation).

    PubMed

    Albutaihi, Ibrahim A M; DeJongste, Mike J L; Ter Horst, Gert J

    2004-01-01

    The awareness in specific brain centers of angina pectoris most often results from ischemic episodes in the heart. These ischemic episodes induce the release of a collage of chemicals that activate chemosensitive and mechanoreceptive receptors in the heart, which in turn excite receptors of the sympathetic afferent pathways. Ascending pain signals from these fibers result in the activation of the brain centers which are involved in the perception and integration of cardiac pain. Cytochemical studies of the nervous system provide the opportunity to identify these areas at the cellular level. In the present investigation, cardiac nociception was studied in the brains and the spinal cords of rats, using Fos protein as a marker of neuronal activation, following the application of pain-inducing chemicals to the heart. Induction of myocardial pain in conscious rats was achieved by infusion of bradykinin (0.5 microg) or capsaicin (5 microg) into the pericardial sac. During pain stimulation, the rats demonstrated pain behavior, in conjunction with alterations in heart rate and blood pressure. The cerebral Fos expression pattern was studied 2 h after pain stimulation. In contrast to the control group, increased Fos expression was found following the use of both capsaicin and bradykinin in a variety of areas of the brain. Bradykinin, but not capsaicin, induced Fos expression in the upper thoracic and upper cervical spinal cord; these segments are the sites where cardiac sympathetic fibers terminate. This finding suggests that these two chemicals use two different pathways, and provides extra evidence for the role of the vagus nerve in the transmission of cardiac nociception. Different cerebral areas showed an increase in the c-fos activity following pericardial application of pain-inducing chemicals. The role of these cerebral areas in the integration of cardiac pain is discussed in relation to the identified pathways which transmit cardiac pain. PMID:15305089

  8. Aberrant expression of T cell and B cell markers in myelocyte/monocyte/histiocyte-derived lymphoma and leukemia cells. Is the infrequent expression of T/B cell markers sufficient to establish a lymphoid origin for Hodgkin's Reed-Sternberg cells?

    PubMed Central

    Hsu, S. M.; Hsu, P. L.

    1989-01-01

    Most Hodgkin's mononuclear cells and Reed-Sternberg (H-RS) cells are characterized by the expression of the antigen CD30, but not of T or B cell markers. A few H-RS cells, however, may express a limited number of T or B cell markers. Whether this expression is sufficient to allow the conclusion that H-RS cells are derived from T and/or B cells has been debated vigorously. The present study examined whether CD30 and aberrant T and B cell markers are expressed in cell lines that are well documented as being derived from the granulocyte/monocyte/histiocyte lineage. These cells included HL-60, KG-1, ML-1, THP-1, and U-937. Four other cell lines derived from patients with leukemias/lymphomas of monocytic or granulocytic origins also were studied. These cells included BV173, CML-Brown, CTV-2, and SU-DHL-1. If aberrant expression is detected, by analogy one may expect that rare T or B cell marker expression may occur in H-RS cells, because abundant evidence has indicated that H-RS cells may be related to cells in histiocyte lineage. In all nine of the cell lines studied, it was confirmed that numerous monocyte/granulocyte markers were expressed. The marker expression was enhanced after cells were induced to differentiate with phorbol ester (TPA) and tumor necrosis factor (TNF). It was noted that several T and B cell markers also were expressed by these cells. Unlike the expression of monocyte/granulocyte markers, the expression of T or B cell markers was not affected, or only minimally affected, by treatment of the cells with TPA or TNF. Five of the cell lines (BV173, CML-Brown, CTV-2, SU-DHL-1, and THP-1) were shown to be CD30-positive. In CTV-2 and BV173, the expression of CD30 was greatly increased after induction with phorbol ester or TNF. Based on these studies, the following conclusions were reached: 1) The expression of aberrant B or T cell markers is not an uncommon finding in granulocyte/monocyte/histiocyte-related neoplastic cells. 2) The expression of

  9. The pan-deacetylase inhibitor panobinostat modulates the expression of epithelial-mesenchymal transition markers in hepatocellular carcinoma models

    PubMed Central

    DI FAZIO, PIETRO; MONTALBANO, ROBERTA; QUINT, KARL; ALINGER, BEATE; KEMMERLING, RALF; KIESSLICH, TOBIAS; OCKER, MATTHIAS; NEUREITER, DANIEL

    2013-01-01

    Deacetylase inhibitors (DACis) represent a novel therapeutic option for human cancers by classically affecting proliferation or apoptosis. Since transdifferentiation and dedifferentiation play a key role in carcinogenesis, we investigated the epigenetic influence on the molecular differentiation status in human hepatocellular carcinoma (HCC) models. Markers of differentiation, including cytokeratin (Ck) 7, Ck8, Ck18, Ck19, Ck20, vimentin, sonic hedgehog homolog (SHH), smoothened (Smo), patched (Ptc), glioma-associated oncogene homolog 1 (Gli1), CD133, octamer-binding transcription factor 4 (Oct4) and β-catenin, were examined in the human HCC cell lines HepG2 and Hep3B in vitro and in vivo (xenograft model) using quantitative real-time PCR and immunohistochemistry following treatment with the pan-DACi panobinostat (LBH589). Compared to untreated controls, treated HepG2 xenografts, and to a lesser extent cell lines, demonstrated a significant increase of differentiation markers Ck7 and Ck19 (classical cholangiocellular type) and Ck8 and Ck18 (classical HCC type), and a decreased level of dedifferentiation markers vimentin (mesenchymal) and SHH/Ptc (embryonic), paralleled with a more membranous expression of β-catenin. These findings were dose-dependently correlated with tumor size, necrosis rate, microvessel density and mitosis/Ki-67-associated proliferation rate. Our results demonstrate that the differentiation status of human HCC cells is influenced by the pan-DACi panobinostat, indicating that this treatment may influence the epithelial-mesenchymal transition (EMT) status related to metastasis and aggressiveness. PMID:23255907

  10. Long-term restricted feeding alters circadian expression and reduces the level of inflammatory and disease markers.

    PubMed

    Sherman, Hadas; Frumin, Idan; Gutman, Roee; Chapnik, Nava; Lorentz, Axel; Meylan, Jenny; le Coutre, Johannes; Froy, Oren

    2011-12-01

    The circadian clock in peripheral tissues can be entrained by restricted feeding (RF), a regimen that restricts the duration of food availability with no calorie restriction (CR). However, it is not known whether RF can delay the occurrence of age-associated changes similar to CR. We measured circadian expression of clock genes, disease marker genes, metabolic factors and inflammatory and allergy markers in mouse serum, liver, jejunum and white adipose tissue (WAT) after long-term RF of 4 months. We found that circadian rhythmicity is more robust and is phase advanced in most of the genes and proteins tested under RF. In addition, average daily levels of some disease and inflammatory markers were reduced under RF, including liver Il-6 mRNA, tumour necrosis factor (TNF)-α and nuclear factor κB (NF-κB) protein; jejunum Arginase, Afp, Gadd45β, Il-1α and Il-1β mRNA, and interleukin (IL)-6 and TNF-α protein and WAT Il-6, Il-1β, Tnfα and Nfκb mRNA. In contrast, the anti-inflammatory cytokine Il-10 mRNA increased in the liver and jejunum. Our results suggest that RF may share some benefits with those of CR. As RF is a less harsh regimen to follow than CR, the data suggest it could be proposed for individuals seeking to improve their health.

  11. Characterization of the "sporadically lurking HAP1-immunoreactive (SLH) cells" in the hippocampus, with special reference to the expression of steroid receptors, GABA, and progenitor cell markers.

    PubMed

    Islam, Md N; Fujinaga, R; Yanai, A; Jahan, M R; Takeshita, Y; Kokubu, K; Shinoda, K

    2012-05-17

    Huntingtin-associated protein 1 (HAP1) is a neural huntingtin interactor that is widely expressed as a core molecule of the stigmoid body (a neurocytoplasmic inclusion) in the limbic and hypothalamic regions and has putative protective functions against some neurodegenerative diseases (HAP1 protection hypothesis). Although HAP1 has been reported to be intimately associated with several steroid receptors, HAP1-immunoreactive (HAP1-ir) cells remain to be identified in the hippocampus, which is one of the major steroidal targets. In this study, we determined the distribution of hippocampal HAP1-ir cells in light and fluorescence microscopy and characterized their morphological relationships with steroid receptors, markers of adult neurogenesis, and the GABAergic system in adult male and female Wistar rats. HAP1-ir cells, which were sporadically distributed particularly in the subgranular zone (SGZ) of the dentate gyrus and in the interface between the stratum lacunosum-moleculare and stratum radiatum of Ammon's horn, were identified as the "sporadically lurking HAP1-ir (SLH)" cells. The SLH cells showed no clear association with neural progenitor/proliferating or migrating cell markers of adult neurogenesis, such as Ki-67, proliferating cell nuclear antigen, doublecortin, and glial fibrillary acidic protein in the SGZ, whereas all the SLH cells expressed a neuronal specific nuclear protein (NeuN). More than 90% of the SLH cells expressed nuclear estrogen receptor (ER) α but neither ERβ nor the androgen receptor, whereas glucocorticoid receptor was differently stained in the SLH cells depending on the antibodies. More than 60% of them exhibited GABA immunoreactivity in the SGZ, suggestive of basket cells, but they were distinct from the ones expressing cholecystokinin or parvalbumin. We conclude that SLH cells, which should be stable against apoptosis due to putative HAP1 protectivity, might be involved in estrogen-dependent maturation, remodeling and activation of

  12. Alveolar epithelial cells in idiopathic pulmonary fibrosis display upregulation of TRAIL, DR4 and DR5 expression with simultaneous preferential over-expression of pro-apoptotic marker p53

    PubMed Central

    Akram, Khondoker M; Lomas, Nicola J; Forsyth, Nicholas R; Spiteri, Monica A

    2014-01-01

    Idiopathic pulmonary fibrosis (IPF) is a progressive, debilitating, and fatal lung disease of unknown aetiology with no current cure. The pathogenesis of IPF remains unclear but repeated alveolar epithelial cell (AEC) injuries and subsequent apoptosis are believed to be among the initiating/ongoing triggers. However, the precise mechanism of apoptotic induction is hitherto elusive. In this study, we investigated expression of a panel of pro-apoptotic and cell cycle regulatory proteins in 21 IPF and 19 control lung tissue samples. We reveal significant upregulation of the apoptosis-inducing ligand TRAIL and its cognate receptors DR4 and DR5 in AEC within active lesions of IPF lungs. This upregulation was accompanied by pro-apoptotic protein p53 overexpression. In contrast, myofibroblasts within the fibroblastic foci of IPF lungs exhibited high TRAIL, DR4 and DR5 expression but negligible p53 expression. Similarly, p53 expression was absent or negligible in IPF and control alveolar macrophages and lymphocytes. No significant differences in TRAIL expression were noted in these cell types between IPF and control lungs. However, DR4 and DR5 upregulation was detected in IPF alveolar macrophages and lymphocytes. The marker of cellular senescence p21WAF1 was upregulated within affected AEC in IPF lungs. Cell cycle regulatory proteins Cyclin D1 and SOCS3 were significantly enhanced in AEC within the remodelled fibrotic areas of IPF lungs but expression was negligible in myofibroblasts. Taken together these findings suggest that, within the remodelled fibrotic areas of IPF, AEC can display markers associated with proliferation, senescence, and apoptotosis, where TRAIL could drive the apoptotic response. Clear understanding of disease processes and identification of therapeutic targets will direct us to develop effective therapies for IPF. PMID:24551275

  13. Thrombin-Mediated Platelet Activation of Lysed Whole Blood and Platelet-Rich Plasma: A Comparison Between Platelet Activation Markers and Ultrastructural Alterations.

    PubMed

    Augustine, Tanya N; van der Spuy, Wendy J; Kaberry, Lindsay L; Shayi, Millicent

    2016-06-01

    Platelet ultrastructural alterations representing spurious activation have been identified in pathological conditions. A limitation of platelet studies is that sample preparation may lead to artifactual activation processes which may confound results, impacting the use of scanning electron microscopy as a supplemental diagnostic tool. We used scanning electron microscopy and flow cytometry to analyze platelet activation in platelet-rich plasma (PRP) and whole blood (WB) samples. PRP generated using a single high g force centrifugation, and WB samples treated with a red blood cell lysis buffer, were exposed to increasing concentrations of the agonist thrombin. Platelets in lysed WB samples responded to thrombin by elevating the activation marker CD62p definitively, with corresponding ultrastructural changes indicating activation. Conversely, CD62p expression in PRP preparations remained static. Ultrastructural analysis revealed fully activated platelets even under low concentration thrombin stimulation, with considerable fibrin deposition. It is proposed that the method for PRP production induced premature platelet activation, preventable by using an inhibitor of platelet aggregation and fibrin polymerization. Nevertheless, our results show a definitive correspondence between flow cytometry and scanning electron microscopy in platelet activation studies, highlighting the potential of the latter technique as a supplemental diagnostic tool. PMID:27329313

  14. p27Kip1 expression as a prognostic marker for squamous cell carcinoma of the head and neck

    PubMed Central

    DE ALMEIDA, MIGUEL REIS; PÉREZ-SAYÁNS, MARIO; SUÁREZ-PEÑARANDA, JOSÉ MANUEL; SOMOZA-MARTÍN, JOSÉ MANUEL; GARCÍA-GARCÍA, ABEL

    2015-01-01

    Regulation of the cell cycle is essential for carcinogenesis. The cell cycle is controlled by cyclin-dependent kinases (CDKs), which are upregulated by cyclins and downregulated by CDK inhibitors (CDKIs). Decreased p27Kip1 expression has been associated with survival rate, tumor size, histological differentiation and the presence of lymph node metastasis in patients with various types of cancer. The aim of the current study is to provide a literature review on the association between p27Kip1 expression and the clinical and pathological aspects of head and neck squamous cell carcinoma (HNSCC), and the expression of other CDKIs of the Cip/Kip family and cyclins. Throughout the literature, different methodologies were used to determine the immunohistochemical expression of p27Kip1; thus, results concerning p27Kip1 expression in HNSCC vary widely. However, it has now been confirmed that p27Kip1 is underexpressed in SCC cells. p27 may be a promising marker for determining the prognosis of HNSCC, despite the marked variability of the results obtained. An association between p27 expression and survival rate, time to recurrence and tumor stage has been observed. Based on the information currently available, it is premature to recommend the analysis of p27Kip1 expression in guiding HNSCC treatment planning. However, although relatively unstudied, the correlation between p27Kip1 expression and other tumor suppressor genes may turn out to be important in determining the prognosis of HNSCC. Further prospective studies utilizing standardized laboratory methodologies and statistics that facilitate meta-analyses are required to confirm this proposal. PMID:26722226

  15. Inhibition of FGF Signalling Pathway Augments the Expression of Pluripotency and Trophoblast Lineage Marker Genes in Porcine Parthenogenetic Blastocyst.

    PubMed

    Li, L Y; Li, M M; Yang, S F; Zhang, J; Li, Z; Zhang, H; Zhu, L; Zhu, X; Verma, V; Liu, Q; Shi, D; Huang, B

    2016-10-01

    The consistent failure to isolate bona fide pluripotent cell lines from livestock indicates that the underlying mechanisms of early lineage specification are poorly defined. Unlike other species, the contrivances of segregation have been comprehensively studied in the mouse. In mouse, FGF/MAPK signalling pathway dictates the segregation of hypoblast (primitive endoderm). However, it is not evident whether this mechanism is also conserved in livestock. Here, in this study, we examined the roles of FGF/MAP kinase signalling pathways in porcine parthenogenetic embryos during the early development. Porcine parthenogenetic embryos were cultured in the medium addition with FGFR inhibitor BGJ398 (10 μm) or DEMOS. Pluripotency- and lineage-related gene expressions in the early porcine embryos were determined. Compared to control, total cell numbers on day 7 were significantly higher (55 ± 5.96 vs 47 ± 1.97, p < 0.05) in embryos cultured in the presence of BGJ398, but had no significant effect on the rate of blastocyst development (47% vs 44%, p > 0.05). Nonetheless, BGJ398 treatment significantly augmented the expression of pluripotency and trophoblast marker genes (SOX2, OCT4, KLF4 and CDX2), but did not significantly change the expression of NANOG and hypoblast marker gene (GATA4). Furthermore, the addition of FGF signalling agonist (FGF2) during the embryo development significantly decreased the expression of pluripotency and trophoblast marker genes (SOX2, NANOG, KLF4 and CDX2), but no significant effect on the expression of OCT4 and GATA4 was observed. Here, we exhibit that inhibition of FGF signalling could improve the quality of the porcine embryo and escalate the chance to capture pluripotency. Besides, it also promotes the trophoblast development of porcine parthenogenetic embryo. In addition, the data suggested that FGF signalling pathway is dispensable for the segregation of hypoblast and epiblast lineages in porcine embryo during the early

  16. WT1 expression in salivary gland pleomorphic adenomas: a reliable marker of the neoplastic myoepithelium.

    PubMed

    Langman, Gerald; Andrews, Claire L; Weissferdt, Annikka

    2011-02-01

    Pleomorphic adenoma is a benign salivary gland neoplasm with a diverse morphology. This is considered to be a function of the neoplastic myoepithelium, which shows histological and immunophenotypical variability. Wilms' tumor 1 gene (WT1) protein, involved in bidirectional mesenchymal-epithelial transition, has been detected by reverse transcription PCR in salivary gland tumors showing myoepithelial-epithelial differentiation. The aim of this study was to investigate the immunoreactivity of WT1 in pleomorphic adenomas and to compare the pattern of staining with p63 and calponin, two reliable markers of myoepithelial cells. A total of 31 cases of pleomorphic adenoma were selected. The myoepithelium was classified as myoepithelial-like (juxtatubular and spindled), modified myoepithelium (myxoid, chondroid and plasmacytoid) and transformed myoepithelium (solid epithelioid, squamous and basaloid cribriform). Immunohistochemistry for WT1, p63 and calponin was assessed in each myoepithelial component, as well as in nonneoplastic myoepithelial cells and inner tubular epithelial cells. There was no immunostaining of tubular epithelial cells by any of the markers. In contrast to p63 and calponin, WT1 did not react with normal myoepithelial cells. Cytoplasmic WT1 staining was present in all pleomorphic adenomas, and in 29 cases (94%), >50% of neoplastic myoepithelial cells were highlighted. p63 and calponin stained the myoepithelium in 30 tumors. In comparison, 50% of cells were positive in 21 (68%) and 9 (29%) cases of p63 and calponin, respectively. Staining with WT1 showed less variability across the spectrum of myoepithelial differentiation with the difference most marked in the transformed myoepithelium. WT1 is a sensitive marker of the neoplastic myoepithelial cell in pleomorphic adenomas. The role of this protein in influencing the mesenchymal-epithelial state of cells suggests that WT1 and the myoepithelial cell have an important role in the histogenesis of

  17. Selective Inducible Nitric Oxide Synthase Inhibitor Reversed Zinc Chloride-Induced Spatial Memory Impairment via Increasing Cholinergic Marker Expression.

    PubMed

    Tabrizian, Kaveh; Azami, Kian; Belaran, Maryam; Soodi, Maliheh; Abdi, Khosrou; Fanoudi, Sahar; Sanati, Mehdi; Mottaghi Dastjerdi, Negar; Soltany Rezaee-Rad, Mohammad; Sharifzadeh, Mohammad

    2016-10-01

    Zinc, an essential micronutrient and biochemical element of the human body, plays structural, catalytic, and regulatory roles in numerous physiological functions. In the current study, the effects of a pretraining oral administration of zinc chloride (10, 25, and 50 mg/kg) for 14 consecutive days and post-training bilateral intra-hippocampal infusion of 1400W as a selective inducible nitric oxide synthase (iNOS) inhibitor (10, 50, and 100 μM/side), alone and in combination, on the spatial memory retention in Morris water maze (MWM) were investigated. Animals were trained for 4 days and tested 48 h after completion of training. Also, the molecular effects of these compounds on the expression of choline acetyltransferase (ChAT), as a cholinergic marker in the CA1 region of the hippocampus and medial septal area (MSA), were evaluated. Behavioral and molecular findings of this study showed that a 2-week oral administration of zinc chloride (50 mg/kg) impaired spatial memory retention in MWM and decreased ChAT expression. Immunohistochemical analysis of post-training bilateral intra-hippocampal infusion of 1400W revealed a significant increase in ChAT immunoreactivity. Furthermore, post-training bilateral intra-hippocampal infusion of 1400W into the CA1 region of the hippocampus reversed zinc chloride-induced spatial memory impairment in MWM and significantly increased ChAT expression in comparison with zinc chloride-treated animals. Taken together, these results emphasize the role of selective iNOS inhibitors in reversing zinc chloride-induced spatial memory deficits via modulation of cholinergic marker expression.

  18. Expression of Phenotypic Astrocyte Marker Is Increased in a Transgenic Mouse Model of Alzheimer's Disease versus Age-Matched Controls: A Presymptomatic Stage Study

    PubMed Central

    Doméné, Aurélie; Cavanagh, Chelsea; Page, Guylène; Bodard, Sylvie; Klein, Christophe; Delarasse, Cécile; Chalon, Sylvie

    2016-01-01

    Recent mouse studies of the presymptomatic stage of Alzheimer's disease (AD) have suggested that proinflammatory changes, such as glial activation and cytokine induction, may occur already at this early stage through unknown mechanisms. Because TNFα contributes to increased Aβ production from the Aβ precursor protein (APP), we assessed a putative correlation between APP/Aβ and TNFα during the presymptomatic stage as well as early astrocyte activation in the hippocampus of 3-month-old APPswe/PS1dE9 mice. While Western blots revealed significant APP expression, Aβ was not detectable by Western blot or ELISA attesting that 3-month-old, APPswe/PS1dE9 mice are at a presymptomatic stage of AD-like pathology. Western blots were also used to show increased GFAP expression in transgenic mice that positively correlated with both TNFα and APP, which were also mutually correlated. Subregional immunohistochemical quantification of phenotypic (GFAP) and functional (TSPO) markers of astrocyte activation indicated a selective and significant increase in GFAP-immunoreactive (IR) cells in the dentate gyrus of APPswe/PS1dE9 mice. Our data suggest that subtle morphological and phenotypic alterations, compatible with the engagement of astrocyte along the activation pathway, occur in the hippocampus already at the presymptomatic stage of AD. PMID:27672476

  19. Expression of Phenotypic Astrocyte Marker Is Increased in a Transgenic Mouse Model of Alzheimer's Disease versus Age-Matched Controls: A Presymptomatic Stage Study

    PubMed Central

    Doméné, Aurélie; Cavanagh, Chelsea; Page, Guylène; Bodard, Sylvie; Klein, Christophe; Delarasse, Cécile; Chalon, Sylvie

    2016-01-01

    Recent mouse studies of the presymptomatic stage of Alzheimer's disease (AD) have suggested that proinflammatory changes, such as glial activation and cytokine induction, may occur already at this early stage through unknown mechanisms. Because TNFα contributes to increased Aβ production from the Aβ precursor protein (APP), we assessed a putative correlation between APP/Aβ and TNFα during the presymptomatic stage as well as early astrocyte activation in the hippocampus of 3-month-old APPswe/PS1dE9 mice. While Western blots revealed significant APP expression, Aβ was not detectable by Western blot or ELISA attesting that 3-month-old, APPswe/PS1dE9 mice are at a presymptomatic stage of AD-like pathology. Western blots were also used to show increased GFAP expression in transgenic mice that positively correlated with both TNFα and APP, which were also mutually correlated. Subregional immunohistochemical quantification of phenotypic (GFAP) and functional (TSPO) markers of astrocyte activation indicated a selective and significant increase in GFAP-immunoreactive (IR) cells in the dentate gyrus of APPswe/PS1dE9 mice. Our data suggest that subtle morphological and phenotypic alterations, compatible with the engagement of astrocyte along the activation pathway, occur in the hippocampus already at the presymptomatic stage of AD.

  20. Expression of Phenotypic Astrocyte Marker Is Increased in a Transgenic Mouse Model of Alzheimer's Disease versus Age-Matched Controls: A Presymptomatic Stage Study.

    PubMed

    Doméné, Aurélie; Cavanagh, Chelsea; Page, Guylène; Bodard, Sylvie; Klein, Christophe; Delarasse, Cécile; Chalon, Sylvie; Krantic, Slavica

    2016-01-01

    Recent mouse studies of the presymptomatic stage of Alzheimer's disease (AD) have suggested that proinflammatory changes, such as glial activation and cytokine induction, may occur already at this early stage through unknown mechanisms. Because TNFα contributes to increased Aβ production from the Aβ precursor protein (APP), we assessed a putative correlation between APP/Aβ and TNFα during the presymptomatic stage as well as early astrocyte activation in the hippocampus of 3-month-old APPswe/PS1dE9 mice. While Western blots revealed significant APP expression, Aβ was not detectable by Western blot or ELISA attesting that 3-month-old, APPswe/PS1dE9 mice are at a presymptomatic stage of AD-like pathology. Western blots were also used to show increased GFAP expression in transgenic mice that positively correlated with both TNFα and APP, which were also mutually correlated. Subregional immunohistochemical quantification of phenotypic (GFAP) and functional (TSPO) markers of astrocyte activation indicated a selective and significant increase in GFAP-immunoreactive (IR) cells in the dentate gyrus of APPswe/PS1dE9 mice. Our data suggest that subtle morphological and phenotypic alterations, compatible with the engagement of astrocyte along the activation pathway, occur in the hippocampus already at the presymptomatic stage of AD. PMID:27672476

  1. Expression of survivin mRNA in gallbladder cancer: a diagnostic and prognostic marker?

    PubMed

    Nigam, Jaya; Chandra, Abhijit; Kazmi, Hasan Raza; Parmar, Devendra; Singh, Devendra; Gupta, Vishal; M, Noushif

    2014-09-01

    Survivin, an inhibitor of apoptosis, has been shown to be expressed in various malignancies. However, its role in gallbladder cancer (GBC) has not been evaluated yet. We investigated its expression in peripheral blood of patients with gallbladder diseases (gallstone disease (GSD), n = 30; GBC, n = 39) and compared with healthy controls (n = 25). Survivin expression was correlated with clinicopathological parameters, diagnosis, and prognosis of patients with GBC. Expression of survivin messenger RNA (mRNA) in blood was evaluated by real-time PCR. Significantly higher (P < 0.0001) expression of survivin mRNA was observed in GBC (2.2-fold) and GSD (1.52-fold) as compared to control. In GBC, increased survivin expression was significantly associated with higher tumor stage (stage III vs. stage II; P < 0.0001) and tumor differentiation (poor and moderate vs. well differentiated; P < 0.0001). No significant correlation was observed with any of the other clinicopathological parameters (age, gender, and presence or absence of gallstones) studied. Cutoff value of survivin mRNA relative quantification (RQ) was 1.08, with a sensitivity of 98.55 % and specificity of 100 % for the diseased group (GSD or GBC). RQ value of 1.71 differentiated GBC from GSD with a sensitivity of 89.74 % and specificity of 100 %. Increased expression of survivin was associated with a shorter median overall survival (12 vs. 18 months) in GBC patients. Differential expression of survivin in GBC suggests its possible role and association with poor prognosis. Expression of survivin in peripheral blood could be useful both in the diagnosis and prognosis of GBC.

  2. Removal of selectable marker gene from fibroblast cells in transgenic cloned cattle by transient expression of Cre recombinase and subsequent effects on recloned embryo development.

    PubMed

    Wang, S; Sun, X; Ding, F; Zhang, K; Zhao, R; Li, S; Li, R; Tang, B; Zhang, L; Liu, Y; Li, J; Gao, F; Wang, H; Wang, L; Dai, Y; Li, N

    2009-09-01

    Introduction of selectable marker genes to transgenic animals could create an inconvenience to further research and may exaggerate public concerns regarding biological safety. The objective of the current study was to excise loxP flanked neo(R) in transgenic cloned cattle by transient expression of Cre recombinase. Green fluorescent protein gene (GFP) was incorporated to monitor Cre expression; therefore, Cre-expressed cells could be selected indirectly by fluorescence-activated cell sorting (FACS). The neo(R) was removed and Cre expressed transiently in GFP-positive colonies; excision of neo(R) was confirmed by single-blastocyst PCR in recloned blastocysts, with neo(R)-free fibroblast cells as donors. There was no difference (P>0.05) in rates of cleavage (76.0% vs. 68.8%) or blastocyst formation (56.6% vs. 52.9%) between recloned embryos with neo(R)-free or neo(R)-included donors. The differential staining of recloned blastocysts were similar (P >0.05) in terms of total cell number (124 vs. 122) and the ratio of ICM (Inner Cell Mass) to the total cell number (38.1% vs. 38.2%). Furthermore, pregnancy and calving rates were not different (P>0.05) from those of the control. In conclusion, we successfully excised neo(R) from transgenic cloned cattle; the manipulation did not affect the developmental competence of recloned preimplantation embryos. This approach should benefit bioreactor and transgenic research in livestock.

  3. CD107a as a marker of activation in chicken cytotoxic T cells.

    PubMed

    Wattrang, Eva; Dalgaard, Tina S; Norup, Liselotte R; Kjærup, Rikke B; Lundén, Anna; Juul-Madsen, Helle R

    2015-04-01

    The study aimed to evaluate cell surface mobilisation of CD107a as a general activation marker on chicken cytotoxic T cells (CTL). Experiments comprised establishment of an in vitro model for activation-induced CD107a mobilisation and design of a marker panel for the detection of CD107a mobilisation on chicken CTL isolated from different tissues. Moreover, CD107a mobilisation was analysed on CTL isolated from airways of infectious bronchitis virus (IBV)-infected birds direct ex vivo and upon in vitro stimulation. Results showed that phorbol 12-myristate 13-acetate (PMA) in combination with ionomycin was a consistent inducer of CD107a cell surface mobilisation on chicken CTL in a 4h cell culture model. In chickens experimentally infected with IBV, higher frequencies of CTL isolated from respiratory tissues were positive for CD107a on the cell surface compared to those from uninfected control chickens indicating in vivo activation. Moreover, upon in vitro PMA+ ionomycin stimulation, higher proportions of CTL isolated from the airways of IBV-infected chickens showed CD107a mobilisation compared to those from uninfected control chickens. Monitoring of CD107a cell surface mobilisation may thus be a useful tool for studies of chicken CTL cytolytic potential both in vivo and in vitro.

  4. Regulation of Aicda expression and AID activity.

    PubMed

    Zan, Hong; Casali, Paolo

    2013-03-01

    Activation-induced cytidine deaminase (AID) is expressed in a B cell differentiation stage-specific fashion and is essential for immunoglobulin (Ig) gene class switch DNA recombination (CSR) and somatic hypermutation (SHM). CSR and SHM play a central role in the maturation of antibody and autoantibody responses. AID displays a mutagenic activity by catalyzing targeted deamination of deoxycytidine (dC) residues in DNA resulting in dU:dG mismatches, which are processed into point-mutations in SHM or double-strand breaks (DSBs) in CSR. Although AID specifically targets the Ig gene loci (IgH, Igκ and Igλ), it can also home into a wide array of non-Ig genes in B-and non-B-cell backgrounds. Aberrant expression of AID is associated with multiple diseases such as allergy, inflammation, autoimmunity and cancer. In autoimmune systemic lupus erythematosus, dysregulated AID expression underpins increased CSR, SHM and autoantibody production. As a potent mutator, AID is under stringent transcriptional, post-transcriptional and post-translational regulation. AID is also regulated in its targeting and enzymatic function. In resting naïve or memory B cells, AID transcripts and protein are undetectable. These, however, are readily and significantly up-regulated in B cells induced to undergo CSR and/or SHM. Transcription factors, such as HoxC4 and NF-κB, which are up-regulated in a B cell lineage-and/or differentiation stage-specific manner, regulate the induction of AID. HoxC4 induces AID expression by directly binding to the AID gene promoter through an evolutionarily conserved 5'-ATTT-3' motif. HoxC4 is induced by the same stimuli that induce AID and CSR. It is further up-regulated by estrogen through three estrogen responsive elements in its promoter region. The targeting of AID to switch (S) regions is mediated by 14-3-3 adaptor proteins, which specifically bind to 5'-AGCT-3' repeats that are exist at high frequency in S region cores. Like HoxC4, 14-3-3 adaptors are induced

  5. Strain magnitude-dependent calcific marker expression in valvular and vascular cells.

    PubMed

    Ferdous, Zannatul; Jo, Hanjoong; Nerem, Robert M

    2013-01-01

    Aortic valve disease and atherosclerosis tend to coexist in most patients with cardiovascular disease; however, the causes and mechanisms of disease development in heart valves are still not clearly understood. To understand the contributions of the magnitude of cyclic strain (5% hypotension, 10% physiological, and 15% hypertension) in calcification, we used a model system of tissue-engineered collagen gels containing human aortic smooth muscle cells and human aortic valvular interstitial cells, both isolated from noncalcific heart transplant tissue. The compacted collagen gels were cultured in osteogenic media for 3 weeks in a custom-designed bioreactor and all assessments were performed at the end of the culture period. The major finding of this study is that bone morphogenic protein (BMP)-2 and BMP-4 and transforming growth factor-β1 mRNA expression significantly changed in response to the magnitude of applied strain in valvular cells, while the lowest expression was observed for the representative physiological strain. On the other hand, mRNA expression in vascular cells did not vary in response to the magnitude of strain. Regarding BMP-2 and BMP-4 protein expression determined by immunostaining, trends were similar to mRNA expression in vascular and valvular cells, where only valvular cells showed a varied protein expression depending on the magnitude of the strain applied. Our results suggest that cellular differences exist between vascular and valvular cells in their response to altered levels of cyclic strain during calcification. PMID:23548742

  6. Gene expression screening of salivary gland neoplasms: molecular markers of potential histogenetic and clinical significance.

    PubMed

    Maruya, Shin-Ichiro; Kim, Hyung-Woo; Weber, Randal S; Lee, Jack J; Kies, Merril; Luna, Mario A; Batsakis, John G; El-Naggar, Adel K

    2004-08-01

    Salivary gland neoplasms comprise phenotypically and biologically diverse lesions of uncertain histogenesis. The molecular events associated with their development and clinicopathological heterogeneity remain unknown. To reveal these events, we performed microarray expression analysis using a nylon-filter membrane platform on 18 primary lesions representing the most common benign and malignant types. Our study identified a small set of genes that are differentially altered between normal salivary gland tissues and benign and malignant tumors. Of the 5000 genes arrayed, 136 genes were differentially expressed by normal tissue, benign tumors, and various malignant neoplasms. Hierarchical clustering analysis differentiated between adenoid cystic carcinomas (ACCs) and other malignant subtypes. Non-ACC specimens manifested overlapping patterns of gene expression within and between tumors. Most of the differentially expressed genes share functional similarities with members of the adhesion, proliferation, and signal transduction pathways. Our study identified: 1) a set of genes that differentiate normal tissue from tumor specimens, 2) genes that differentiate pleomorphic adenoma and ACCs from other malignant salivary gland neoplasms, and 3) different patterns of expression between ACCs arising from major and minor salivary gland sites. The differentially expressed genes provide new information on potential genetic events of biological significance in future studies of salivary gland tumorigenesis.

  7. Pyrophosphate Stimulates Differentiation, Matrix Gene Expression and Alkaline Phosphatase Activity in Osteoblasts

    PubMed Central

    Pujari-Palmer, Michael; Pujari-Palmer, Shiuli; Lu, Xi; Lind, Thomas; Melhus, Håkan; Engstrand, Thomas; Karlsson-Ott, Marjam; Engqvist, Hakan

    2016-01-01

    Pyrophosphate is a potent mitogen, capable of stimulating proliferation in multiple cell types, and a critical participant in bone mineralization. Pyrophosphate can also affect the resorption rate and bioactivity of orthopedic ceramics. The present study investigated whether calcium pyrophosphate affected proliferation, differentiation and gene expression in early (MC3T3 pre-osteoblast) and late stage (SAOS-2 osteosarcoma) osteoblasts. Pyrophosphate stimulated peak alkaline phosphatase activity by 50% and 150% at 100μM and 0.1μM in MC3T3, and by 40% in SAOS-2. The expression of differentiation markers collagen 1 (COL1), alkaline phosphatase (ALP), osteopontin (OPN), and osteocalcin (OCN) were increased by an average of 1.5, 2, 2 and 3 fold, by high concentrations of sodium pyrophosphate (100μM) after 7 days of exposure in MC3T3. COX-2 and ANK expression did not differ significantly from controls in either treatment group. Though both high and low concentrations of pyrophosphate stimulate ALP activity, only high concentrations (100μM) stimulated osteogenic gene expression. Pyrophosphate did not affect proliferation in either cell type. The results of this study confirm that chronic exposure to pyrophosphate exerts a physiological effect upon osteoblast differentiation and ALP activity, specifically by stimulating osteoblast differentiation markers and extracellular matrix gene expression. PMID:27701417

  8. Neuropsin Expression Correlates with Dendritic Marker MAP2c Level in Different Brain Regions of Aging Mice.

    PubMed

    Konar, Arpita; Thakur, M K

    2015-01-01

    Neuropsin (NP) is a serine protease, implicated in synaptic plasticity and memory acquisition through cleavage of synaptic adhesion molecule, L1CAM. However, NP has not been explored during brain aging that entails drastic deterioration of plasticity and memory with selective regional vulnerability. Therefore, we have analysed the expression of NP and correlated with its function via analysis of endogenous cleavage of L1CAM and level of dendritic marker MAP2c in different regions of the aging mouse brain. While NP expression gradually decreased in the cerebral cortex during aging, it showed a sharp rise in both olfactory bulb and hippocampus in adult and thereafter declined in old age. NP expression was moderate in young medulla, but undetectable in midbrain and cerebellum. It was positively correlated with L1CAM cleavage and MAP2c level in different brain regions during aging. Taken together, our study shows age-dependent regional variation in NP expression and its positive correlation with MAP2c level, suggesting the involvement of NP in MAP2c mediated alterations in dendritic morphology during aging. PMID:24965600

  9. IGF2 expression is a marker for paraganglionic/SIF cell differentiation in neuroblastoma.

    PubMed Central

    Hedborg, F.; Ohlsson, R.; Sandstedt, B.; Grimelius, L.; Hoehner, J. C.; Pählman, S.

    1995-01-01

    Neuroblastoma is a childhood tumor of the sympathetic nervous system. Observations in the Beckwith-Wiedemann syndrome suggest that sympathetic embryonal cells with an abundant expression of the insulin-like growth factor 2 gene (IGF2) may be involved in the genesis of low-malignant infant neuroblastomas. We have therefore compared the cell type-specific IGF2 expression of the human sympathetic nervous system during early development with that of neuroblastoma. An abundant expression in normal sympathetic tissue was specific to extra-adrenal chromaffin cells, ie, paraganglia and small intensely fluorescent (SIF) cells, whereas sympathetic neuronal cells were IGF2-negative. A subpopulation of neuroblastomas expressed IGF2, which correlated with an early age at diagnosis, an extra-adrenal tumor origin, and severe hemodynamic signs of catecholamine secretion. Histologically IGF2-expressing tumors displayed a lobular growth pattern, and expression was restricted to the most mature and least proliferative cells. Typically, these cells were morphologically and histochemically similar to paraganglia/SIF cells and formed distinct ring-like zones in the center of the lobules around a core of apoptosis-like tumor cells. The similarities found between IGF2-expressing neuroblastoma cells and paraganglia/SIF cells in terms of histological features, anatomical origin, and age-dependent growth suggest a paraganglionic/SIF cell lineage of most infant tumors and also of extra-adrenal tumors diagnosed after infancy. Furthermore, since paraganglia/SIF cells undergo postnatal involution, the same cellular mechanism may be responsible for spontaneous regression in infant neuroblastoma. Images Figure 2 Figure 3 p839-a Figure 4 PMID:7717451

  10. Exposure to Diesel Exhaust Particle Extracts (DEPe) Impairs Some Polarization Markers and Functions of Human Macrophages through Activation of AhR and Nrf2

    PubMed Central

    Jaguin, Marie; Fardel, Olivier; Lecureur, Valérie

    2015-01-01

    Macrophages (MΦ), well-known to play an important role in immune response, also respond to environmental toxic chemicals such as diesel exhaust particles (DEP). Potential effects of DEPs towards MΦ polarization, a key hall-mark of MΦ physiology, remain however poorly documented. This study was therefore designed to evaluate the effects of a reference DEP extract (DEPe) on human MΦ polarization. Human blood monocytes-derived MΦ were incubated with IFNγ+LPS or IL-4 to obtain M1 and M2 subtypes, respectively; a 24 h exposure of polarizing MΦ to 10 μg/ml DEPe was found to impair expression of some macrophagic M1 and M2 markers, without however overall inhibition of M1 and M2 polarization processes. Notably, DEPe treatment increased the secretion of the M1 marker IL-8 and the M2 marker IL-10 in both MΦ subtypes, whereas it reduced lipopolysaccharide-induced IL-6 and IL-12p40 secretion in M1 MΦ. In M2 MΦ, DEPe exposure led to a reduction of CD200R expression and of CCL17, CCL18 and CCL22 secretion, associated with a lower chemotaxis of CCR4-positive cells. DEPe activated the Nrf2 and AhR pathways and induced expression of their reference target genes such as Hmox-1 and cytochrome P-4501B1 in M1 and M2 MΦ. Nrf2 or AhR silencing through RNA interference prevented DEPe-related down-regulation of IL-6. AhR silencing also inhibited the down-secretion of IL-12p40 and CCL18 in M1- and M2-DEPe-exposed MΦ, respectively. DEPs are therefore likely to alter expression of some M1 and M2 markers in an AhR- and Nrf2-dependent manner; such regulations may contribute to deleterious immune effects of atmospheric DEP. PMID:25710172

  11. Large-scale production and evaluation of marker-free indica rice IR64 expressing phytoferritin genes.

    PubMed

    Oliva, Norman; Chadha-Mohanty, Prabhjit; Poletti, Susanna; Abrigo, Editha; Atienza, Genelou; Torrizo, Lina; Garcia, Ruby; Dueñas, Conrado; Poncio, Mar Aristeo; Balindong, Jeanette; Manzanilla, Marina; Montecillo, Florencia; Zaidem, Maricris; Barry, Gerard; Hervé, Philippe; Shou, Huxia; Slamet-Loedin, Inez H

    2014-01-01

    Biofortification of rice (Oryza sativa L.) using a transgenic approach to increase the amount of iron in the grain is proposed as a low-cost, reliable, and sustainable solution to help developing countries combat anemia. In this study, we generated and evaluated a large number of rice or soybean ferritin over-accumulators in rice mega-variety IR64, including marker-free events, by introducing soybean or rice ferritin genes into the endosperm for product development. Accumulation of the protein was confirmed by ELISA, in situ immunological detection, and Western blotting. As much as a 37- and 19-fold increase in the expression of ferritin gene in single and co-transformed plants, respectively, and a 3.4-fold increase in Fe content in the grain over the IR64 wild type was achieved using this approach. Agronomic characteristics of a total of 1,860 progenies from 58 IR64 single independent transgenic events and 768 progenies from 27 marker-free transgenic events were evaluated and most trait characteristics did not show a penalty. Grain quality evaluation of high-Fe IR64 transgenic events showed quality similar to that of the wild-type IR64. To understand the effect of transgenes on iron homeostasis, transcript analysis was conducted on a subset of genes involved in iron uptake and loading. Gene expression of the exogenous ferritin gene in grain correlates with protein accumulation and iron concentration. The expression of NAS2 and NAS3 metal transporters increased during the grain milky stage. PMID:24482599

  12. The Expression of c-Myb Correlates with the Levels of Rhabdomyosarcoma-specific Marker Myogenin

    PubMed Central

    Kaspar, Petr; Zikova, Martina; Bartunek, Petr; Sterba, Jaroslav; Strnad, Hynek; Kren, Leos; Sedlacek, Radislav

    2015-01-01

    The transcription factor c-Myb is required for modulation of progenitor cells in several tissues, including skeletal muscle and its upregulation is observed in many human malignancies. Rhabdomyosarcomas (RMS) are a heterogeneous group of mesodermal tumors with features of developing skeletal muscle. Several miRNAs are downregulated in RMS, including miR-150, a negative regulator of c-Myb expression. Using the C2C12 myoblast cell line, a cellular model of skeletal muscle differentiation, we showed that miR-150 controls c-Myb expression mainly at the level of translation. We hypothesized that a similar mechanism of c-Myb regulation operates in RMS tumors. We examined expression of c-Myb by immunohistochemistry and revealed c-Myb positivity in alveolar and embryonal tumors, the two most common subgroups of RMS. Furthermore, we showed direct correlation between c-Myb production and myogenin expression. Interestingly, high myogenin levels indicate poor prognosis in RMS patients. c-Myb could, therefore, contribute to the tumor phenotype by executing its inhibitory role in skeletal muscle differentiation. We also showed that c-Myb protein is abundant in migratory C2C12 myoblasts and its ectopic expression potentiates cell motility. In summary, our results implicate that metastatic properties of some RMS subtypes might be linked to c-Myb function. PMID:26462877

  13. Endosialin expression in soft tissue sarcoma as a potential marker of undifferentiated mesenchymal cells

    PubMed Central

    Thway, Khin; Robertson, David; Jones, Robin L; Selfe, Joanna; Shipley, Janet; Fisher, Cyril; Isacke, Clare M

    2016-01-01

    Background: Soft tissue sarcomas are a group of neoplasms with differentiation towards mesenchymal tissue, many of which are aggressive and chemotherapy resistant. Histology and immunoprofiles often overlap with neoplasms of other lineages, and establishing an accurate histopathological diagnosis is crucial for correct management, and therapeutic stratification. The endosialin cell surface glycoprotein is predominantly expressed by stromal fibroblasts and pericytes in epithelial neoplasms; however, tumour cell expression has been reported in small series of sarcomas. Methods: We assessed endosialin expression by immunohistochemistry in a large set of 514 human soft tissue sarcomas. Results: Tumour cell endosialin expression was seen in 89% of undifferentiated pleomorphic sarcomas (104/117), 77% adult fibrosarcomas/spindle cell sarcomas (20/26), 62% synovial sarcomas (37/60), 51% leiomyosarcomas (94/185) and 31% rhabdomyosarcomas (39/126). Conclusions: Endosialin immunohistochemistry has potential to distinguish undifferentiated and poorly differentiated sarcomas from other poorly differentiated, non-mesenchymal neoplasms. A Phase II trial randomising patients with advanced sarcomas to receive chemotherapy with/without an endosialin therapeutic antibody has recently completed enrolment. Endosialin expression could be used to select patients for such clinical trials. Based on our results, patients with undifferentiated pleomorphic sarcoma may be particularly suitable for such a therapeutic approach. PMID:27434038

  14. Robust expression of heterologous genes by selection marker fusion system in improved Chlamydomonas strains.

    PubMed

    Kong, Fantao; Yamasaki, Tomohito; Kurniasih, Sari Dewi; Hou, Liyuan; Li, Xiaobo; Ivanova, Nina; Okada, Shigeru; Ohama, Takeshi

    2015-09-01

    Chlamydomonas is a very attractive candidate plant cell factory. However, its main drawback is the difficulty to find the transformants that robustly express heterologous genes randomly inserted in the nuclear genome. We previously showed that domestic squalene synthase (SQS) gene of Chlamydomonas was much more efficiently overexpressed in a mutant strain [UV-mediated mutant (UVM) 4] than in wild type. In this study, we evaluated the possibility of a new mutant strain, met1, which contains a tag in the maintenance type methyltransferase gene that is expected to play a key role in the maintenance of transcriptional gene silencing. The versatile usefulness of the UVM4 strain to express heterologous genes was also analyzed. We failed to overexpress CrSSL3 cDNA, which is the codon-adjusted squalene synthase-like gene originated from Botryococcus braunii, using the common expression cassette in the wild-type CC-1690 and UVM4 strains. However, we succeeded in isolating western blot-positive transformants through the combinational use of the UVM4 strain and ble2A expression system of which expression cassette bears a fused ORF of the target gene and the antibiotic resistance gene ble via the foot-and-mouth disease virus (FMDV) self-cleaving 2A sequence. It is noteworthy that even with this system, huge deviations in the accumulated protein levels were still observed among the UVM4 transformants. PMID:25660568

  15. Impact of physical activity on ovarian reserve markers in normal, overweight and obese reproductive age women.

    PubMed

    Surekha, T; Himabindu, Y; Sriharibabu, M; Pandey, Anil Kumar

    2014-01-01

    Physical inactivity is a leading risk factor for overweight and obesity in the society. Prevalence of overweight and obesity in the reproductive age group women not only affects maternal health but also the health of the off spring. Infertility is a common problem in India affecting 13-19 million people at any given time. Even though it is not life threatening, infertility causes intense mental agony and trauma that can only be best described by infertile couples themselves. Infertility is more common in overweight and obese individuals compared to normal weight individuals. Decreasing ovarian reserve is an important factor for infertility in women. This study examined the impact of physical activity on ovarian reserve markers in normal, overweight and obese reproductive age women. The observations made in this study reveal that physical activity improves ovarian reserve markers in all reproductive age women but this improvement is more distinct and statistically significant in overweight and obese women compared to normal weight women. PMID:25509968

  16. Prospective active marker motion correction improves statistical power in BOLD fMRI.

    PubMed

    Muraskin, Jordan; Ooi, Melvyn B; Goldman, Robin I; Krueger, Sascha; Thomas, William J; Sajda, Paul; Brown, Truman R

    2013-03-01

    Group level statistical maps of blood oxygenation level dependent (BOLD) signals acquired using functional magnetic resonance imaging (fMRI) have become a basic measurement for much of systems, cognitive and social neuroscience. A challenge in making inferences from these statistical maps is the noise and potential confounds that arise from the head motion that occurs within and between acquisition volumes. This motion results in the scan plane being misaligned during acquisition, ultimately leading to reduced statistical power when maps are constructed at the group level. In most cases, an attempt is made to correct for this motion through the use of retrospective analysis methods. In this paper, we use a prospective active marker motion correction (PRAMMO) system that uses radio frequency markers for real-time tracking of motion, enabling on-line slice plane correction. We show that the statistical power of the activation maps is substantially increased using PRAMMO compared to conventional retrospective correction. Analysis of our results indicates that the PRAMMO acquisition reduces the variance without decreasing the signal component of the BOLD (beta). Using PRAMMO could thus improve the overall statistical power of fMRI based BOLD measurements, leading to stronger inferences of the nature of processing in the human brain.

  17. Parietal Epithelial Cell Activation Marker in Early Recurrence of FSGS in the Transplant

    PubMed Central

    Fatima, Huma; Moeller, Marcus J.; Smeets, Bart; Yang, Hai-Chun; D’Agati, Vivette D.; Alpers, Charles E.

    2012-01-01

    Summary Background and objectives Podocyte loss is key in glomerulosclerosis. Activated parietal epithelial cells are proposed to contribute to pathogenesis of glomerulosclerosis and may serve as stem cells that can transition to podocytes. CD44 is a marker for activated parietal epithelial cells. This study investigated whether activated parietal epithelial cells are increased in early recurrent FSGS in transplant compared with minimal change disease. Design, setting, participants, & measurements CD44 staining in renal allograft biopsies from 12 patients with recurrent FSGS was performed and compared with native kidneys with minimal change disease or FSGS and normal control native and transplant kidneys without FSGS. CD44+ epithelial cells along Bowman’s capsule in the parietal epithelial cell location and over the glomerular tuft in the visceral epithelial cell location were assessed. Results Cases with early recurrent FSGS manifesting only foot process effacement showed significantly increased CD44+ visceral epithelial cells involving 29.0% versus 2.6% of glomeruli in minimal change disease and 0% in non-FSGS transplants. Parietal location CD44 positivity also was numerically increased in recurrent FSGS. In later transplant biopsies, glomeruli with segmental lesions had more CD44+ visceral epithelial cells than glomeruli without lesions. Conclusions Parietal epithelial cell activation marker is significantly increased in evolving FSGS versus minimal change disease, and this increase may distinguish early FSGS from minimal change disease. Whether parietal epithelial cell activation contributes to pathogenesis of sclerosis in idiopathic FSGS or is a regenerative/repair response to replace injured podocytes awaits additional study. PMID:22917699

  18. TSPAN33 is a novel marker of activated and malignant B cells

    PubMed Central

    Luu, Van Phi; Hevezi, Peter; Vences-Catalan, Felipe; Maravillas-Montero, Jose Luis; White, Clayton Alexander; Casali, Paolo; Llorente, Luis; Jakez-Ocampo, Juan; Lima, Guadalupe; Vilches-Cisneros, Natalia; Flores-Gutiérrez, Juan Pablo; Santos-Argumedo, Leopoldo; Zlotnik, Albert

    2014-01-01

    We have identified Tspan33 as a gene encoding a transmembrane protein exhibiting a restricted expression pattern including expression in activated B cells. TSPAN33 is a member of the tetraspanin family. TSPAN33 is not expressed in resting B cells, but is strongly induced in primary human B cells following activation. Human 2E2 cells, a Burkitt’s lymphoma-derived B cell model of activation and differentiation, also upregulate TSPAN33 upon activation. TSPAN33 is expressed in several lymphomas including Hodgkin’s and Diffuse large B Cell Lymphoma. TSPAN33 is also expressed in some autoimmune diseases where B cells participate in the pathology, including rheumatoid arthritis patients, systemic lupus erythematosus (SLE), and in spleen B cells from MRL/Faslpr/lpr mice (a mouse model of SLE). We conclude that TSPAN33 may be used as a diagnostic biomarker or as a target for therapeutic antibodies for treatment of certain B cell lymphomas or autoimmune diseases. PMID:24211713

  19. Cloning and Expression of CD19, a Human B-Cell Marker in NIH-3T3 Cell Line

    PubMed Central

    Abbasi-Kenarsari, Hajar; Shafaghat, Farzaneh; Baradaran, Behzad; Movassaghpour, Ali Akbar; Shanehbandi, Dariush; Kazemi, Tohid

    2015-01-01

    Background CD19 is a pan B cell marker that is recognized as an attractive target for antibody-based therapy of B-cell disorders including autoimmune disease and hematological malignancies. The object of this study was to stably express the human CD19 antigen in the murine NIH-3T3 cell line aimed to be used as an immunogen in our future study. Methods Total RNA was extracted from Raji cells in which high expression of CD19 was confirmed by flow cytometry. Synthesized cDNA was used for CD19 gene amplification by conventional PCR method using Pfu DNA polymerase. PCR product was ligated to pGEM-T Easy vector and ligation mixture was transformed to DH5α competent bacteria. After blue/white selection, one positive white colony was subjected to plasmid extraction and direct sequencing. Then, CD19 cDNA was sub-cloned into pCMV6-Neo expression vector by double digestion using KpnI and HindIII enzymes. NIH-3T3 mouse fibroblast cell line was subsequently transfected by the construct using Jet-PEI transfection reagent. After 48 hours, surface expression of CD19 was confirmed by flow cytometry and stably transfected cells were selected by G418 antibiotic. Results Amplification of CD19 cDNA gave rise to 1701 bp amplicon confirmed by alignment to reference sequence in NCBI database. Flow cytometric analysis showed successful transient and stable expression of CD19 on NIH-3T3 cells (29 and 93%, respectively). Conclusion Stable cell surface expression of human CD19 antigen in a murine NIH-3T3 cell line may develop a proper immunogene which raises specific anti-CD19 antibody production in the mice immunized sera. PMID:25926951

  20. Down-regulation of tumor endothelial marker 8 suppresses cell proliferation mediated by ERK1/2 activity

    PubMed Central

    Cao, Chuangjie; Wang, Zhuo; Huang, Leilei; Bai, Lihong; Wang, Yuefeng; Liang, Yingjie; Dou, Chengyun; Wang, Liantang

    2016-01-01

    Tumor endothelial marker 8 (TEM8) was recently suggested as a putative anti-tumor target in several types of human cancer based on its selective overexpression in tumor versus normal endothelial cells. The objective of this study was to detect the potential functions of TEM8 in osteosarcoma. Overall, TEM8 was mainly located in cytoplasm and was up-regulated in osteosarcoma compared to benign bone lesions and adjacent non tumor tissue (ANT). High TEM8 expression group had a significant lower overall survival rate than that in the low TEM8 expression group. TEM8 knock-down by siRNA or shRNA results in significant reduction of osteosarcoma cell growth and proliferation both in vitro and in vivo. Ablation of TEM8 led to increasing of p21 and p27 and suppression of cyclin D1 mediated by Erk1/2 activity. These findings suggest that down-regulation of TEM8 play an important role in the inhibition of tumorigenesis and development of osteosarcoma. PMID:26996335

  1. Zingerone suppresses liver inflammation induced by antibiotic mediated endotoxemia through down regulating hepatic mRNA expression of inflammatory markers in Pseudomonas aeruginosa peritonitis mouse model.

    PubMed

    Kumar, Lokender; Chhibber, Sanjay; Harjai, Kusum

    2014-01-01

    Antibiotic-induced endotoxin release is associated with high mortality rate even when appropriate antibiotics are used for the treatment of severe infections in intensive care units. Since liver is involved in systemic clearance and detoxification of endotoxin hence it becomes a primary target organ for endotoxin mediated inflammation. Currently available anti-inflammatory drugs give rise to serious side effects. Hence, there is an urgent need for safe and effective anti-inflammatory therapy. It is likely that anti-inflammatory phytochemicals and neutraceutical agents may have the potential to reduce the endotoxin mediated inflammation and complications associated with endotoxin release. Keeping this in mind, the present study was planned to evaluate the hepatoprotective potential of zingerone (active compound of zingiber officinale) against liver inflammation induced by antibiotic mediated endotoxemia. The selected antibiotics capable of releasing high content of endotoxin were employed for their in vivo efficacy in P.aeruginosa peritonitis model. Released endotoxin induced inflammation and zingerone as co-anti-inflammatory therapy significantly reduced inflammatory response. Improved liver histology and reduced inflammatory markers MDA, RNI, MPO, tissue damage markers (AST, ALT, ALP) and inflammatory cytokines (MIP-2, IL-6 and TNF-α) were indicative of therapeutic potential of zingerone. The mechanism of action of zingerone may be related to significant inhibition of the mRNA expression of inflammatory markers (TLR4, RelA, NF-kB2, TNF- α, iNOS, COX-2) indicating that zingerone interferes with cell signalling pathway and suppresses hyper expression of cell signaling molecules of inflammatory pathway. Zingerone therapy significantly protected liver from endotoxin induced inflammatory damage by down regulating biochemical as well as molecular markers of inflammation. In conclusion, this study provides evidence that zingerone is a potent anti

  2. Loss of H3K27me3 Expression Is a Highly Sensitive Marker for Sporadic and Radiation-induced MPNST

    PubMed Central

    Prieto-Granada, Carlos N.; Wiesner, Thomas; Messina, Jane L.; Jungbluth, Achim A.; Chi, Ping; Antonescu, Cristina R.

    2016-01-01

    Most malignant peripheral nerve sheath tumors (MPNSTs) exhibit combined inactivation of NF1, CDKN2A, and polycomb repressive complex 2 component genes (Embryonic Ectoderm Development [EED] and Suppressor of Zeste 12 [SUZ12]). Mutations in EED and SUZ12 induce loss of trimethylation at lysine 27 of histone 3 (H3K27me3), with subsequent aberrant transcriptional activation of polycomb repressive complex 2–repressed homeobox master regulators. These findings prompted us to investigate the performance of an anti-H3K27me3 monoclonal antibody clone C36B11 as an immunohistochemical marker for MPNSTs. We assessed the C36B11 reactivity pattern in a pathologically and genetically well-characterized cohort of 68 MPNSTs, spanning various clinical presentations, such as type 1 neurofibromatosis (NF1), radiotherapy, and sporadic MPNSTs. We found that 69% (n=47) of all MPNSTs demonstrated loss of H3K27me3 expression, with 42 (61%) showing complete loss and 5 (7%) showing partial loss, whereas 31% (n = 21) retained H3K27me3 expression. Among the NF1-related high-grade MPNSTs, 60% demonstrated loss of expression. In contrast, the majority of both sporadic (95%) and radiotherapy-related (91%) MPNSTs showed loss of H3K27me3 expression. Two of the 3 low-grade MPNSTs and all neurofibromas showed retained expression. Furthermore, all 5 epithelioid MPNSTs retained H3K27me3 labeling. The specificity of H3K27me3 loss as a marker for MPNSTs was studied by testing a large spectrum of lesions included in MPNST differential diagnosis, such as spindle/desmoplastic melanomas, synovial sarcomas, myoepithelial tumors, and other mesenchymal neoplasms, all of which retained expression of H3K27me3. We conclude that immunohistochemical analysis of H3K27me3 has good sensitivity and robust specificity for the diagnosis of MPNST, particularly outside of NF1 clinical history, which represents the most challenging diagnostic setting. PMID:26645727

  3. Determination of protein markers in human serum: Analysis of protein expression in toxic oil syndrome studies.

    PubMed

    Quero, Carmen; Colomé, Nuria; Prieto, Maria Rosario; Carrascal, Montserrat; Posada, Manuel; Gelpí, Emilio; Abian, Joaquin

    2004-02-01

    Toxic oil syndrome (TOS) is a disease that appeared in Spain in 1981. It affected more than 20 000 people and produced over 300 deaths in the first 2 years. In this paper, a prospective study on the differences in gene expression in sera between a control versus a TOS-affected population, both originally exposed to the toxic oil, is presented. Differential protein expression was analyzed by two-dimensional electrophoresis (2-DE). Several problems related with serum analysis by 2-DE were addressed in order to improve protein detection in the gel images. Three new commercial systems for albumin depletion were tested to optimize the detection of minor proteins that can be obscured by the presence of a few families of high abundance proteins (albumin, immunoglobulins). Other factors, such as the use of nonionic reductants or the presence of thiourea in the gels, were also tested. From these optimized images, a group of 329 major gel spots was located, matched and compared in serum samples. Thirty-five of these protein spots were found to be under- or overexpressed in TOS patients (> three-fold increase or decrease). Proteins in the differential spots were identified by matrix-assisted laser desorption/ionization-time of flight peptide map fingerprinting and database search. Several haptoglobin isoforms were found to be differentially expressed, showing expression phenotypes that could be related with TOS affection. Haptoglobin phenotypes have been previously reported to have important biological and clinical consequences and have been described as risk factors for several diseases.

  4. Expressing Uncertainty in Computer-Mediated Discourse: Language as a Marker of Intellectual Work

    ERIC Educational Resources Information Center

    Jordan, Michelle E.; Schallert, Diane L.; Park, Yangjoo; Lee, SoonAh; Chiang, Yueh-hui Vanessa; Cheng, An-Chih Janne; Song, Kwangok; Chu, Hsiang-Ning Rebecca; Kim, Taehee; Lee, Haekyung

    2012-01-01

    Learning and dialogue may naturally engender feelings and expressions of uncertainty for a variety of reasons and purposes. Yet, little research has examined how patterns of linguistic uncertainty are enacted and changed over time as students reciprocally influence one another and the dialogical system they are creating. This study describes the…

  5. Critical role for BRCA1 expression as a marker of chemosensitivity response and prognosis.

    PubMed

    De Luca, Paola; De Siervi, Adriana

    2016-01-01

    Chemotherapy is still the leader option for cancer treatment. Nevertheless some patients develop chemotherapy resistance. One major research goal is to identify the critical genes involved in chemotherapy response to predict the best therapy option for patients. Germline mutations in the BReast Cancer susceptibility gene (BRCA1) are associated to increased risk of developing breast, ovarian and other types of cancers. However, due to harmful BRCA1 gene mutations are relatively rare in the general population, nowadays most researchers focused on BRCA1 expression downregulation and/or epigenetic inactivation in sporadic tumors as a prognosis tool for chemotherapy response in patients. Chemotherapy response can be dramatically different depending on BRCA1 expression status, tumor type and drug. Hence, the chemotherapy response could be dissimilar in breast, ovarian, uterine, prostate, esophageal, gastric and lung cancers. Additionally, differential BRCA1 expression in sporadic tumors shows different response to DNA-damaging agents, mitotic inhibitors or PARP inhibitors. In this review we will examine the response to different chemotherapy agents in several cancer types depending on BRCA1 expression status.

  6. Hypoxia Increases the Expression of Stem-Cell Markers and Promotes Clonogenicity in Glioblastoma Neurospheres

    PubMed Central

    Bar, Eli E.; Lin, Alex; Mahairaki, Vasiliki; Matsui, William; Eberhart, Charles G.

    2010-01-01

    Hypoxia promotes the expansion of non-neoplastic stem and precursor cell populations in the normal brain, and is common in malignant brain tumors. We examined the effects of hypoxia on stem-like cells in glioblastoma (GBM). When GBM-derived neurosphere cultures are grown in 1% oxygen, hypoxia-inducible factor 1α (HIF1α) protein levels increase dramatically, and mRNA encoding other hypoxic response genes, such as those encoding hypoxia-inducible gene-2, lysyl oxidase, and vascular endothelial growth factor, are induced over 10-fold. Hypoxia increases the stem-like side population over fivefold, and the percentage of cells expressing CD133 threefold or more. Notch pathway ligands and targets are also induced. The rise in the stem-like fraction in GBM following hypoxia is paralleled by a twofold increase in clonogenicity. We believe HIF1α plays a causal role in these changes, as when oxygen-stable HIF1α is expressed in normoxic glioma cells CD133 is induced. We used digoxin, which has been shown to lower HIF protein levels in vitro and in vivo, to inhibit the hypoxic response. Digoxin suppressed HIF1α protein expression, HIF1α downstream targets, and slowed tumor growth in vivo. In addition, pretreatment with digoxin reduced GBM flank xenograft engraftment of hypoxic GBM cells, and daily intraperitoneal injections of digoxin were able to significantly inhibit the growth of established subcutaneous glioblastoma xenografts, and suppressed expression of vascular endothelial growth factor. PMID:20671264

  7. Changes in urinary amino acids excretion in relationship with muscle activity markers over a professional cycling stage race: in search of fatigue markers.

    PubMed

    Corsetti, Roberto; Barassi, Alessandra; Perego, Silvia; Sansoni, Veronica; Rossi, Alessandra; Damele, Clara Anna Linda; Melzi D'Eril, Gianlodovico; Banfi, Giuseppe; Lombardi, Giovanni

    2016-01-01

    The aim of this study was to identify the relationship between metabolic effort, muscular damage/activity indices, and urinary amino acids profile over the course of a strenuous prolonged endurance activity, as a cycling stage race is, in order to identify possible fatigue markers. Nine professional cyclists belonging to a single team, competing in the Giro d'Italia cycling stage race, were anthropometrically characterized and sampled for blood and urine the day before the race started, and on days 12 and 23 of the race. Diet was kept the same over the race, and power output and energy expenditure were recorded. Sera were assayed for muscle markers (lactate dehydrogenase, aspartate aminotransferase, and creatine kinase activities, and blood urea nitrogen), and creatinine, all corrected for plasma volume changes. Urines were profiled for amino acid concentrations, normalized on creatinine excretion. Renal function, in terms of glomerular filtration rate, was monitored by MDRD equation corrected on body surface area. Creatine kinase activity and blood urea were increased during the race as did serum creatinine while kidney function remained stable. Among the amino acids, taurine, glycine, cysteine, leucine, carnosine, 1-methyl histidine, and 3-methyl histidine showed a net decreased, while homocysteine was increased. Taurine and the dipeptide carnosine (β-alanyl-L-histidine) were significantly correlated with the muscle activity markers and the indices of effort. In conclusion, the metabolic profile is modified strikingly due to the effort. Urinary taurine and carnosine seem useful tools to evaluate the muscle damage and possibly the fatigue status on a long-term basis. PMID:26306846

  8. Changes in urinary amino acids excretion in relationship with muscle activity markers over a professional cycling stage race: in search of fatigue markers.

    PubMed

    Corsetti, Roberto; Barassi, Alessandra; Perego, Silvia; Sansoni, Veronica; Rossi, Alessandra; Damele, Clara Anna Linda; Melzi D'Eril, Gianlodovico; Banfi, Giuseppe; Lombardi, Giovanni

    2016-01-01

    The aim of this study was to identify the relationship between metabolic effort, muscular damage/activity indices, and urinary amino acids profile over the course of a strenuous prolonged endurance activity, as a cycling stage race is, in order to identify possible fatigue markers. Nine professional cyclists belonging to a single team, competing in the Giro d'Italia cycling stage race, were anthropometrically characterized and sampled for blood and urine the day before the race started, and on days 12 and 23 of the race. Diet was kept the same over the race, and power output and energy expenditure were recorded. Sera were assayed for muscle markers (lactate dehydrogenase, aspartate aminotransferase, and creatine kinase activities, and blood urea nitrogen), and creatinine, all corrected for plasma volume changes. Urines were profiled for amino acid concentrations, normalized on creatinine excretion. Renal function, in terms of glomerular filtration rate, was monitored by MDRD equation corrected on body surface area. Creatine kinase activity and blood urea were increased during the race as did serum creatinine while kidney function remained stable. Among the amino acids, taurine, glycine, cysteine, leucine, carnosine, 1-methyl histidine, and 3-methyl histidine showed a net decreased, while homocysteine was increased. Taurine and the dipeptide carnosine (β-alanyl-L-histidine) were significantly correlated with the muscle activity markers and the indices of effort. In conclusion, the metabolic profile is modified strikingly due to the effort. Urinary taurine and carnosine seem useful tools to evaluate the muscle damage and possibly the fatigue status on a long-term basis.

  9. RNA Sequencing of Sessile Serrated Colon Polyps Identifies Differentially Expressed Genes and Immunohistochemical Markers

    PubMed Central

    Delker, Don A.; Pop, Stelian; Neklason, Deborah W.; Bronner, Mary P.; Burt, Randall W.; Hagedorn, Curt H.

    2014-01-01

    Background Sessile serrated adenomas/polyps (SSA/Ps) may account for 20–30% of colon cancers. Although large SSA/Ps are generally recognized phenotypically, small (<1 cm) or dysplastic SSA/Ps are difficult to differentiate from hyperplastic or small adenomatous polyps by endoscopy and histopathology. Our aim was to define the comprehensive gene expression phenotype of SSA/Ps to better define this cancer precursor. Results RNA sequencing was performed on 5′ capped RNA from seven SSA/Ps collected from patients with the serrated polyposis syndrome (SPS) versus eight controls. Highly expressed genes were analyzed by qPCR in additional SSA/Ps, adenomas and controls. The cellular localization and level of gene products were examined by immunohistochemistry in syndromic and sporadic SSA/Ps, adenomatous and hyperplastic polyps and controls. We identified 1,294 differentially expressed annotated genes, with 106 increased ≥10-fold, in SSA/Ps compared to controls. Comparing these genes with an array dataset for adenomatous polyps identified 30 protein coding genes uniquely expressed ≥10-fold in SSA/Ps. Biological pathways altered in SSA/Ps included mucosal integrity, cell adhesion, and cell development. Marked increased expression of MUC17, the cell junction protein genes VSIG1 and GJB5, and the antiapoptotic gene REG4 were found in SSA/Ps, relative to controls and adenomas, were verified by qPCR analysis of additional SSA/Ps (n = 21) and adenomas (n = 10). Immunohistochemical staining of syndromic (n≥11) and sporadic SSA/Ps (n≥17), adenomatous (n≥13) and hyperplastic (n≥10) polyps plus controls (n≥16) identified unique expression patterns for VSIG1 and MUC17 in SSA/Ps. Conclusion A subset of genes and pathways are uniquely increased in SSA/Ps, compared to adenomatous polyps, thus supporting the concept that cancer develops by different pathways in these phenotypically distinct polyps with markedly different gene expression profiles. Immunostaining

  10. Expression of Estrogen-Related Gene Markers in Breast Cancer Tissue Predicts Aromatase Inhibitor Responsiveness

    PubMed Central

    Moy, Irene; Lin, Zhihong; Rademaker, Alfred W.; Reierstad, Scott; Khan, Seema A.; Bulun, Serdar E.

    2013-01-01

    Aromatase inhibitors (AIs) are the most effective class of drugs in the endocrine treatment of breast cancer, with an approximate 50% treatment response rate. Our objective was to determine whether intratumoral expression levels of estrogen-related genes are predictive of AI responsiveness in postmenopausal women with breast cancer. Primary breast carcinomas were obtained from 112 women who received AI therapy after failing adjuvant tamoxifen therapy and developing recurrent breast cancer. Tumor ERα and PR protein expression were analyzed by immunohistochemistry (IHC). Messenger RNA (mRNA) levels of 5 estrogen-related genes–AKR1C3, aromatase, ERα, and 2 estradiol/ERα target genes, BRCA1 and PR–were measured by real-time PCR. Tumor protein and mRNA levels were compared with breast cancer progression rates to determine predictive accuracy. Responsiveness to AI therapy–defined as the combined complete response, partial response, and stable disease rates for at least 6 months–was 51%; rates were 56% in ERα-IHC-positive and 14% in ERα-IHC-negative tumors. Levels of ERα, PR, or BRCA1 mRNA were independently predictive for responsiveness to AI. In cross-validated analyses, a combined measurement of tumor ERα and PR mRNA levels yielded a more superior specificity (36%) and identical sensitivity (96%) to the current clinical practice (ERα/PR-IHC). In patients with ERα/PR-IHC-negative tumors, analysis of mRNA expression revealed either non-significant trends or statistically significant positive predictive values for AI responsiveness. In conclusion, expression levels of estrogen-related mRNAs are predictive for AI responsiveness in postmenopausal women with breast cancer, and mRNA expression analysis may improve patient selection. PMID:24223121

  11. Long-term Exposure to Air Pollution and Markers of Inflammation, Coagulation, and Endothelial Activation

    PubMed Central

    Hajat, Anjum; Allison, Matthew; Diez-Roux, Ana V.; Jenny, Nancy Swords; Jorgensen, Neal W.; Szpiro, Adam A.; Vedal, Sverre; Kaufman, Joel D.

    2015-01-01

    Background Air pollution is associated with cardiovascular disease, and systemic inflammation may mediate this effect. We assessed associations between long- and short-term concentrations of air pollution and markers of inflammation, coagulation, and endothelial activation. Methods We studied participants from the Multi-Ethnic Study of Atherosclerosis from 2000 to 2012 with repeat measures of serum C-reactive protein (CRP), interleukin-6 (IL-6), fibrinogen, D-dimer, soluble E-selectin, and soluble Intercellular Adhesion Molecule-1. Annual average concentrations of ambient fine particulate matter (PM2.5), individual-level ambient PM2.5 (integrating indoor concentrations and time–location data), oxides of nitrogen (NOx), nitrogen dioxide (NO2), and black carbon were evaluated. Short-term concentrations of PM2.5 reflected the day of blood draw, day prior, and averages of prior 2-, 3-, 4-, and 5-day periods. Random-effects models were used for long-term exposures and fixed effects for short-term exposures. The sample size was between 9,000 and 10,000 observations for CRP, IL-6, fibrinogen, and D-dimer; approximately 2,100 for E-selectin; and 3,300 for soluble Intercellular Adhesion Molecule-1. Results After controlling for confounders, 5 µg/m3 increase in long-term ambient PM2.5 was associated with 6% higher IL-6 (95% confidence interval = 2%, 9%), and 40 parts per billion increase in long-term NOx was associated with 7% (95% confidence interval = 2%, 13%) higher level of D-dimer. PM2.5 measured at day of blood draw was associated with CRP, fibrinogen, and E-selectin. There were no other positive associations between blood markers and short- or long-term air pollution. Conclusions These data are consistent with the hypothesis that long-term exposure to air pollution is related to some markers of inflammation and fibrinolysis. PMID:25710246

  12. CNMS: The preferred genic markers for comparative genomic, molecular phylogenetic, functional genetic diversity and differential gene regulatory expression analyses in chickpea.

    PubMed

    Bajaj, Deepak; Das, Shouvik; Parida, Swarup K

    2015-09-01

    The intra/inter-genomic comparative mapping-based phylogenetic footprinting identified 5 paralogous and 656 orthologous genome-wide CNMS markers in the upstream sequences of chickpea genes. These CNMS markers revealed a high-degree of gene-based syntenic relationship between chickpea and Medicago genomes while minimum between chickpea and Vitis genomes. The time of divergence and duplication estimated using CNMS markers highlight the expected phylogenetic relationships between chickpea and six dicot (legume) species as well as occurrence of ancient genome (approximately 53 Mya) with small-scale recent segmental (approximately 10 Mya) duplication events in chickpea. A wider level of functional molecular diversity (14 to 88 percent) and admixed population genetic structure was detected among desi, kabuli and wild genotypes by genic CNMS markers at a genome-wide scale suggesting their utility in large-scale genetic analysis in chickpea. The subfunctionalization at the cis-regulatory element region and TFBS (transcription factor binding site) motif levels in the upstream sequences of CNMS marker-associated orthologous genes than the paralogues was predominant. Functional constraint might have considerable effect on these CNMScontaining regulatory elements controlling consistent orthologous gene expression in dicots. A rapid subfunctionalization based on diverge differential expression of paralogous CNMS marker-associated genes particularly those that underwent recent small-scale segmental duplication events in chickpea was apparent. The differential regulation of expression and subfunctionalization potential of Ultra CNMS marker-associated genes suggest their utility in deciphering the complex gene regulatory function as well as identification and targeted mapping of potential genes/QTLs governing vital agronomic traits in chickpea. The gene-based CNMS markers with desirable inherent genetic attributes like higher degree of comparative genome mapping, functional

  13. Gene Expression Profiling Supports the Neural Crest Origin of Adult Rodent Carotid Body Stem Cells and Identifies CD10 as a Marker for Mesectoderm-Committed Progenitors.

    PubMed

    Navarro-Guerrero, Elena; Platero-Luengo, Aida; Linares-Clemente, Pedro; Cases, Ildefonso; López-Barneo, José; Pardal, Ricardo

    2016-06-01

    Neural stem cells (NSCs) are promising tools for understanding nervous system plasticity and repair, but their use is hampered by the lack of markers suitable for their prospective isolation and characterization. The carotid body (CB) contains a population of peripheral NSCs, which support organ growth during acclimatization to hypoxia. We have set up CB neurosphere (NS) cultures enriched in differentiated neuronal (glomus) cells versus undifferentiated progenitors to investigate molecular hallmarks of cell classes within the CB stem cell (CBSC) niche. Microarray gene expression analysis in NS is compatible with CBSCs being neural crest derived-multipotent progenitor cells able to sustain CB growth upon exposure to hypoxia. Moreover, we have identified CD10 as a marker suitable for isolation of a population of CB mesectoderm-committed progenitor cells. CD10 + cells are resting in normoxia, and during hypoxia they are activated to proliferate and to eventually complete maturation into mesectodermal cells, thus participating in the angiogenesis necessary for CB growth. Our results shed light into the molecular and cellular mechanisms involved in CBSC fate choice, favoring a potential use of these cells for cell therapy. Stem Cells 2016;34:1637-1650.

  14. Decreased expression of stem cell markers by simvastatin in 7,12-dimethylbenz(a)anthracene (DMBA)-induced breast cancer.

    PubMed

    Rennó, André Lisboa; Alves-Júnior, Marcos José; Rocha, Rafael Malagoli; De Souza, Philipi Coutinho; de Souza, Valéria Barbosa; Jampietro, Juliano; Vassallo, José; Hyslop, Stephen; Anhê, Gabriel Forato; de Moraes Schenka, Natália Guimarães; Soares, Fernando Augusto; Schenka, André Almeida

    2015-04-01

    Simvastatin, a competitive inhibitor of HMG-CoA reductase widely used in the treatment and prevention of hyperlipidemia-related diseases, has recently been associated to in vitro anticancer stem cell (CSC) actions. However, these effects have not been confirmed in vivo. To assess in vivo anti-CSC effects of simvastatin, female Sprague-Dawley rats with 7,12-dimethyl-benz(a)anthracene (DMBA)-induced mammary cancer and control animals were treated for 14 days with either simvastatin (20 or 40 mg/kg/day) or soybean oil (N = 60). Tumors and normal breast tissues were removed for pathologic examination and immunodetection of CSC markers. At 40 mg/kg/day, simvastatin significantly reduced tumor growth and the expression of most CSC markers. The reduction in tumor growth (80%) could not be explained solely by the decrease in CSCs, since the latter accounted for less than 10% of the neoplasia (differentiated cancer cells were also affected). Stem cells in normal, nonneoplastic breast tissues were not affected by simvastatin. Simvastatin was also associated with a significant decrease in proliferative activity but no increase in cell death. In conclusion, this is the first study to confirm simvastatin anti-CSC actions in vivo, further demonstrating that this effect is specific for neoplastic cells, but not restricted to CSCs, and most likely due to inhibition of cell proliferation.

  15. Melanoma-specific marker expression in skin biopsy tissues as a tool to facilitate melanoma diagnosis.

    PubMed

    Alexandrescu, Doru T; Kauffman, C Lisa; Jatkoe, Timothy A; Hartmann, Dan P; Vener, Tatiana; Wang, Haiying; Derecho, Carlo; Rajpurohit, Yashoda; Wang, Yixin; Palma, John F

    2010-07-01

    Diagnosis of cutaneous melanoma requires accurate differentiation of true malignant tumors from highly atypical lesions, which lack the capacity to develop uncontrolled proliferation and to metastasize. We used melanoma markers from previous work to differentiate benign and atypical lesions from melanoma using paraffin-embedded tissue. This critical step in diagnosis generates the most uncertainty and discrepancy between dermatopathologists. A total of 193 biopsy tissues were selected: 47 melanomas, 48 benign nevi, and 98 atypical/suspicious, including 48 atypical nevi and 50 melanomas as later assigned by expert dermatopathologists. Performance for SILV, GDF15, and L1CAM normalized to TYR in unequivocal melanoma versus benign nevi resulted in an area under the curve (AUC) of 0.94, 0.67, and 0.5, respectively. SILV also differentiated atypical cases classified as melanoma from atypical nevi with an AUC=0.74. Furthermore, SILV showed a significant difference between suspicious melanoma and each suspicious atypia group: melanoma versus severe atypia and melanoma versus moderate atypia had P-values of 0.0077 and 0.0009, respectively. SILV showed clear discrimination between melanoma and benign unequivocal cases as well as between different atypia subgroups in the group of suspicious samples. The role and potential utility of this molecular assay as an adjunct to the morphological diagnosis of melanoma are discussed.

  16. Application of an electric DNA-chip for the expression analysis of bioprocess-relevant marker genes of Bacillus subtilis.

    PubMed

    Jürgen, Britta; Barken, Kim Bundvig; Tobisch, Steffen; Pioch, Daniel; Wümpelmann, Mogens; Hecker, Michael; Schweder, Thomas

    2005-11-01

    The knowledge of critical process-relevant genes can be used for an improved control of bioprocesses. So far bioprocess-relevant marker genes can be analyzed by established expression analysis methods only off-line. In this study, an alternative approach for a potential at-line monitoring of gene expression during bioprocesses is suggested. This approach is based on the measurement of specific mRNAs on an electric DNA-chip in connection with a magnetic bead-based sandwich hybridization. In order to allow an at-line measurement of specific mRNAs an improved method for a fast and partially automated isolation of high quality-RNA samples was developed. The expression analysis of the electric DNA-chip was compared with optical DNA micro arrays and the real time RT-PCR for three selected process-relevant genes of Bacillus subtilis. We demonstrate that the mRNA analysis by means of the electric DNA-chip gives similar results compared to the micro array analysis and the real time RT-PCR technique.

  17. Co-expression of autophagic markers following photodynamic therapy in SW620 human colon adenocarcinoma cells

    PubMed Central

    Ziółkowska, Barbara; Woźniak, Marta; Ziółkowski, Piotr

    2016-01-01

    Photodynamic therapy (PDT) is a minimally invasive cancer treatment. It involves the combination of a photosensitizer and light of a specific wavelength to generate singlet oxygen and other reactive oxygen species that lead to tumor cell death. Autophagy is one of the pathways that tumor cells undergo during photodamage and it is common in photodynamic therapy. The aim of this study was to examine the effect of in vitro PDT on the expression of autophagy-related proteins, autophagy related 7 (Atg7), light chain 3 (LC3) and Beclin-1. Human SW620 colon carcinoma cells were treated with 5-aminolevulinic acid (ALA)-based PDT at a dose of 3 mM. The irradiation was performed using 4.5 J/cm2 total light and a fluence rate of 60 mW/cm2. Autophagy was evaluated by immunocytochemistry using specific antibodies to Atg7, Beclin-1 and LC3. The evaluation was repeated at several time points (0, 4, 8 and 24 h) following irradiation. The induction of autophagy was observed directly following the 5-ALA-mediated PDT procedure with the strongest expression of autophagy-related proteins at 4 and 8 h after irradiation as demonstrated using immunocytochemistry. It was characterized by significantly increased expression of Beclin-1, Atg7 and LC3. To the best of our knowledge this is the first study to analyze Beclin-1, Atg7 and LC3 expression in a PDT-related experiment. This study enhances the understanding of the role of autophagy in PDT, which may contribute to better and more effective tumor responses to this therapy. PMID:27485939

  18. Development and characterization of 1,827 expressed sequence tag-derived simple sequence repeat markers for ramie (Boehmeria nivea L. Gaud).

    PubMed

    Liu, Touming; Zhu, Siyuan; Fu, Lili; Tang, Qingming; Yu, Yongting; Chen, Ping; Luan, Mingbao; Wang, Changbiao; Tang, Shouwei

    2013-01-01

    Ramie (Boehmeria nivea L. Gaud) is one of the most important natural fiber crops, and improvement of fiber yield and quality is the main goal in efforts to breed superior cultivars. However, efforts aimed at enhancing the understanding of ramie genetics and developing more effective breeding strategies have been hampered by the shortage of simple sequence repeat (SSR) markers. In our previous study, we had assembled de novo 43,990 expressed sequence tags (ESTs). In the present study, we searched these previously assembled ESTs for SSRs and identified 1,685 ESTs (3.83%) containing 1,878 SSRs. Next, we designed 1,827 primer pairs complementary to regions flanking these SSRs, and these regions were designated as SSR markers. Among these markers, dinucleotide and trinucleotide repeat motifs were the most abundant types (36.4% and 36.3%, respectively), whereas tetranucleotide, pentanucleotide, and hexanucleotide motifs represented <10% of the markers. The motif AG/CT was the most abundant, accounting for 28.74% of the markers. One hundred EST-SSR markers (97 SSRs located in genes encoding transcription factors and 3 SSRs in genes encoding cellulose synthases) were amplified using polymerase chain reaction for detecting 24 ramie varieties. Of these 100 markers, 98 markers were successfully amplified and 81 markers were polymorphic, with 2-6 alleles among the 24 varieties. Analysis of the genetic diversity of all 24 varieties revealed similarity coefficients that ranged from 0.51 to 0.80. The EST-SSRs developed in this study represent the first large-scale development of SSR markers for ramie. These SSR markers could be used for development of genetic and physical maps, quantitative trait loci mapping, genetic diversity studies, association mapping, and cultivar fingerprinting.

  19. A Balanced Tissue Composition Reveals New Metabolic and Gene Expression Markers in Prostate Cancer.

    PubMed

    Tessem, May-Britt; Bertilsson, Helena; Angelsen, Anders; Bathen, Tone F; Drabløs, Finn; Rye, Morten Beck

    2016-01-01

    Molecular analysis of patient tissue samples is essential to characterize the in vivo variability in human cancers which are not accessible in cell-lines or animal models. This applies particularly to studies of tumor metabolism. The challenge is, however, the complex mixture of various tissue types within each sample, such as benign epithelium, stroma and cancer tissue, which can introduce systematic biases when cancers are compared to normal samples. In this study we apply a simple strategy to remove such biases using sample selections where the average content of stroma tissue is balanced between the sample groups. The strategy is applied to a prostate cancer patient cohort where data from MR spectroscopy and gene expression have been collected from and integrated on the exact same tissue samples. We reveal in vivo changes in cancer-relevant metabolic pathways which are otherwise hidden in the data due to tissue confounding. In particular, lowered levels of putrescine are connected to increased expression of SRM, reduced levels of citrate are attributed to upregulation of genes promoting fatty acid synthesis, and increased succinate levels coincide with reduced expression of SUCLA2 and SDHD. In addition, the strategy also highlights important metabolic differences between the stroma, epithelium and prostate cancer. These results show that important in vivo metabolic features of cancer can be revealed from patient data only if the heterogeneous tissue composition is properly accounted for in the analysis. PMID:27100877

  20. A Balanced Tissue Composition Reveals New Metabolic and Gene Expression Markers in Prostate Cancer

    PubMed Central

    Tessem, May-Britt; Bertilsson, Helena; Angelsen, Anders; Bathen, Tone F.; Drabløs, Finn; Rye, Morten Beck

    2016-01-01

    Molecular analysis of patient tissue samples is essential to characterize the in vivo variability in human cancers which are not accessible in cell-lines or animal models. This applies particularly to studies of tumor metabolism. The challenge is, however, the complex mixture of various tissue types within each sample, such as benign epithelium, stroma and cancer tissue, which can introduce systematic biases when cancers are compared to normal samples. In this study we apply a simple strategy to remove such biases using sample selections where the average content of stroma tissue is balanced between the sample groups. The strategy is applied to a prostate cancer patient cohort where data from MR spectroscopy and gene expression have been collected from and integrated on the exact same tissue samples. We reveal in vivo changes in cancer-relevant metabolic pathways which are otherwise hidden in the data due to tissue confounding. In particular, lowered levels of putrescine are connected to increased expression of SRM, reduced levels of citrate are attributed to upregulation of genes promoting fatty acid synthesis, and increased succinate levels coincide with reduced expression of SUCLA2 and SDHD. In addition, the strategy also highlights important metabolic differences between the stroma, epithelium and prostate cancer. These results show that important in vivo metabolic features of cancer can be revealed from patient data only if the heterogeneous tissue composition is properly accounted for in the analysis. PMID:27100877

  1. Gene Expression Changes in Phosphorus Deficient Potato (Solanum tuberosum L.) Leaves and the Potential for Diagnostic Gene Expression Markers

    PubMed Central

    Hammond, John P.; Broadley, Martin R.; Bowen, Helen C.; Spracklen, William P.; Hayden, Rory M.; White, Philip J.

    2011-01-01

    Background There are compelling economic and environmental reasons to reduce our reliance on inorganic phosphate (Pi) fertilisers. Better management of Pi fertiliser applications is one option to improve the efficiency of Pi fertiliser use, whilst maintaining crop yields. Application rates of Pi fertilisers are traditionally determined from analyses of soil or plant tissues. Alternatively, diagnostic genes with altered expression under Pi limiting conditions that suggest a physiological requirement for Pi fertilisation, could be used to manage Pifertiliser applications, and might be more precise than indirect measurements of soil or tissue samples. Results We grew potato (Solanum tuberosum L.) plants hydroponically, under glasshouse conditions, to control their nutrient status accurately. Samples of total leaf RNA taken periodically after Pi was removed from the nutrient solution were labelled and hybridised to potato oligonucleotide arrays. A total of 1,659 genes were significantly differentially expressed following Pi withdrawal. These included genes that encode proteins involved in lipid, protein, and carbohydrate metabolism, characteristic of Pi deficient leaves and included potential novel roles for genes encoding patatin like proteins in potatoes. The array data were analysed using a support vector machine algorithm to identify groups of genes that could predict the Pi status of the crop. These groups of diagnostic genes were tested using field grown potatoes that had either been fertilised or unfertilised. A group of 200 genes could correctly predict the Pi status of field grown potatoes. Conclusions This paper provides a proof-of-concept demonstration for using microarrays and class prediction tools to predict the Pi status of a field grown potato crop. There is potential to develop this technology for other biotic and abiotic stresses in field grown crops. Ultimately, a better understanding of crop stresses may improve our management of the crop, improving

  2. Telomerase activity, estrogen receptors (α, β), Bcl-2 expression in human breast cancer and treatment response

    PubMed Central

    Murillo-Ortiz, Blanca; Astudillo-De la Vega, Horacio; Castillo-Medina, Sebastian; Malacara, JM; Benitez-Bribiesca, Luis

    2006-01-01

    Background The mechanism for maintaining telomere integrity is controlled by telomerase, a ribonucleoprotein enzyme that specifically restores telomere sequences, lost during replication by means of an intrinsic RNA component as a template for polymerization. Among the telomerase subunits, hTERT (human telomerase reverse transcriptase) is expressed concomitantly with the activation of telomerase. The role of estrogens and their receptors in the transcriptional regulation of hTERT has been demonstrated. The current study determines the possible association between telomerase activity, the expression of both molecular forms of estrogen receptor (ERα and ERβ) and the protein bcl-2, and their relative associations with clinical parameters. Methods Tissue samples from 44 patients with breast cancer were used to assess telomerase activity using the TRAP method and the expression of ERα, ERβ and bcl-2 by means of immunocytochemical techniques. Results Telomerase activity was detected in 59% of the 44 breast tumors examined. Telomerase activity ranged from 0 to 49.93 units of total product generated (TPG). A correlation was found between telomerase activity and differentiation grade (p = 0.03). The only significant independent marker of response to treatment was clinical stage. We found differences between the frequency of expression of ERα (88%) and ERβ (36%) (p = 0.007); bcl-2 was expressed in 79.5% of invasive breast carcinomas. We also found a significant correlation between low levels of telomerase activity and a lack of ERβ expression (p = 0.03). Conclusion Lower telomerase activity was found among tumors that did not express estrogen receptor beta. This is the first published study demonstrating that the absence of expression of ERβ is associated with low levels of telomerase activity. PMID:16911782

  3. Expression of Putative Stem Cell Marker, Hepatocyte Nuclear Factor 4 Alpha, in Mammary Gland of Water Buffalo.

    PubMed

    Choudhary, Ratan K; Choudhary, Shanti; Kaur, Harmanjot; Pathak, Devendra

    2016-01-01

    Buffaloes account for more than 56% of total milk production in India. Cyclic remodeling of mammary glands of human, mice, cow, sheep, and goat is determined by mammary stem cells. It is logical to assume that buffalo mammary gland will have mammary stem/progenitor cells. Thus far, no report exists on identification of buffalo mammary stem cells. Hepatocyte nuclear factor 4 alpha (HNF4A) is a candidate marker for hepatic progenitor cells and has recently been suggested as a marker of bovine mammary stem/progenitor cells. We hypothesized that ( 1 ) HNF4A identifies putative buffalo mammary stem/progenitor cells and ( 2 ) the number of HNF4A-positive cells increases during mastitis. Sixteen buffalo mammary samples were collected from a local slaughterhouse. Hematoxylin and eosin staining were performed on 5-micron thick sections and on the basis of gross examination and histomorphology of the mammary glands, physiological stages of the animals were estimated as non-lactating (n = 4), mastitis (n = 9), and prepubertal (n = 3). In total, 24048 cells were counted (5-10 microscopic fields/animal; n = 16 animals) of which, 40% cells were mammary epithelial cells (MEC) and 60% cells were the stromal cells. The percentage of MEC in non-lactating animals was higher compared to mastitic animals (47.3% vs. 37.3%), which was likely due to loss of MEC in mastitis. HNF4A staining was observed in nuclei of MEC of ducts, alveoli, and stromal cells. Basal location and low frequency of HNF4A-positive MEC (ranges from 0.4-4.5%) were consistent with stem cell characteristics. Preliminary study showed coexpression of HNF4A with MSI1 (a mammary stem cell marker in sheep), suggesting HNF4A was likely to be a putative mammary stem/progenitor cell marker in buffalo. HNF4A-positive MEC (basal and luminal; light and dark stained) tended to be higher in non-lactating than the mastitic animals (8.73 ± 1.71% vs. 4.29 ± 1.19%; P = 0.07). The first hypothesis that HNF4A identify

  4. Expression of Putative Stem Cell Marker, Hepatocyte Nuclear Factor 4 Alpha, in Mammary Gland of Water Buffalo.

    PubMed

    Choudhary, Ratan K; Choudhary, Shanti; Kaur, Harmanjot; Pathak, Devendra

    2016-01-01

    Buffaloes account for more than 56% of total milk production in India. Cyclic remodeling of mammary glands of human, mice, cow, sheep, and goat is determined by mammary stem cells. It is logical to assume that buffalo mammary gland will have mammary stem/progenitor cells. Thus far, no report exists on identification of buffalo mammary stem cells. Hepatocyte nuclear factor 4 alpha (HNF4A) is a candidate marker for hepatic progenitor cells and has recently been suggested as a marker of bovine mammary stem/progenitor cells. We hypothesized that ( 1 ) HNF4A identifies putative buffalo mammary stem/progenitor cells and ( 2 ) the number of HNF4A-positive cells increases during mastitis. Sixteen buffalo mammary samples were collected from a local slaughterhouse. Hematoxylin and eosin staining were performed on 5-micron thick sections and on the basis of gross examination and histomorphology of the mammary glands, physiological stages of the animals were estimated as non-lactating (n = 4), mastitis (n = 9), and prepubertal (n = 3). In total, 24048 cells were counted (5-10 microscopic fields/animal; n = 16 animals) of which, 40% cells were mammary epithelial cells (MEC) and 60% cells were the stromal cells. The percentage of MEC in non-lactating animals was higher compared to mastitic animals (47.3% vs. 37.3%), which was likely due to loss of MEC in mastitis. HNF4A staining was observed in nuclei of MEC of ducts, alveoli, and stromal cells. Basal location and low frequency of HNF4A-positive MEC (ranges from 0.4-4.5%) were consistent with stem cell characteristics. Preliminary study showed coexpression of HNF4A with MSI1 (a mammary stem cell marker in sheep), suggesting HNF4A was likely to be a putative mammary stem/progenitor cell marker in buffalo. HNF4A-positive MEC (basal and luminal; light and dark stained) tended to be higher in non-lactating than the mastitic animals (8.73 ± 1.71% vs. 4.29 ± 1.19%; P = 0.07). The first hypothesis that HNF4A identify

  5. Evaluation of Potential Clinical Surrogate Markers of a Trauma Induced Alteration of Clotting Factor Activities

    PubMed Central

    Payas, Arzu; Schoeneberg, Carsten; Wegner, Alexander; Kauther, Max Daniel; Lendemans, Sven

    2016-01-01

    Objective. The aim of this study was to identify routinely available clinical surrogate markers for potential clotting factor alterations following multiple trauma. Methods. In 68 patients admitted directly from the scene of the accident, all soluble clotting factors were analyzed and clinical data was collected prospectively. Ten healthy subjects served as control group. Results. Patients showed reduced activities of clotting factors II, V, VII, and X and calcium levels (all P < 0.0001 to 0.01). Levels of hemoglobin and base deficit correlated moderately to highly with the activities of a number of clotting factors. Nonsurvivors and patients who needed preclinical intubation or hemostatic therapy showed significantly reduced factor activities at admission. In contrast, factor VIII activity was markedly elevated after injury in general (P < 0.0001), but reduced in nonsurvivors (P < 0.05). Conclusions. Multiple trauma causes an early reduction of the activities of nearly all soluble clotting factors in general. Initial hemoglobin and, with certain qualifications, base deficit levels demonstrated a potential value in detecting those underlying clotting factor deficiencies. Nevertheless, their role as triggers of a hemostatic therapy as well as the observed response of factor VIII to multiple trauma and also its potential prognostic value needs further evaluation. PMID:27433474

  6. Increased serum β2-microglobulin is associated with clinical and immunological markers of disease activity in systemic lupus erythematosus patients.

    PubMed

    Hermansen, M-L F; Hummelshøj, L; Lundsgaard, D; Hornum, L; Keller, P; Fleckner, J; Fox, B; Poulsen, L K; Jacobsen, S

    2012-09-01

    The objective of this study was to explore the relationship between serum levels of β2-microglobulin (β2MG), which some studies suggest reflect disease activity in systemic lupus erythematosus (SLE), and various clinical and immunological markers of disease activity in SLE. Twenty-six SLE patients and 10 healthy controls were included. Disease activity was assessed by: SLEDAI, 24 hr-proteinuria, circulating levels of complement C3, anti-double-stranded DNA (anti-dsDNA), β2MG and various pro-inflammatory and anti-inflammatory cytokines (IL-6, IL-8, IL-10, IL-18) measured with a multiplex assay, IFN-α assessed with a reporter gene assay, and a combined expression score of 12 IFN-α inducible genes in peripheral blood mononuclear cells. Median serum levels of β2MG were significantly higher in SLE patients vs controls (2.8 mg/L, range: 1.1-21.6 and 1.2 mg/L, range: 0.9-1.7, respectively, p < 0.001). β2MG was correlated with SLEDAI score (R = 0.68, p < 0.001), 24 hr-proteinuria (R = 0.64, p < 0.001), and complement C3 (R = -0.52, p = 0.007). The cytokines were significantly correlated with β2MG: IL-6 (R = 0.45, p = 0.02), IL-8 (R = 0.75, p < 0.001), IL-10 (R = 0.67, p < 0.001) and IL-18 (R = 0.71, p < 0.001) as were serum IFN-α (R = 0.45, p = 0.02) and the IFN-α inducible gene-score (R = 0.51, p = 0.01). The results support that β2MG may serve as a marker of disease activity in SLE. The correlations with the measured cytokines indicate that increased β2MG in SLE reflects immunological activity.

  7. Characterizing proton-activated materials to develop PET-mediated proton range verification markers

    NASA Astrophysics Data System (ADS)

    Cho, Jongmin; Ibbott, Geoffrey S.; Kerr, Matthew D.; Amos, Richard A.; Stingo, Francesco C.; Marom, Edith M.; Truong, Mylene T.; Palacio, Diana M.; Betancourt, Sonia L.; Erasmus, Jeremy J.; DeGroot, Patricia M.; Carter, Brett W.; Gladish, Gregory W.; Sabloff, Bradley S.; Benveniste, Marcelo F.; Godoy, Myrna C.; Patil, Shekhar; Sorensen, James; Mawlawi, Osama R.

    2016-06-01

    Conventional proton beam range verification using positron emission tomography (PET) relies on tissue activation alone and therefore requires particle therapy PET whose installation can represent a large financial burden for many centers. Previously, we showed the feasibility of developing patient implantable markers using high proton cross-section materials (18O, Cu, and 68Zn) for in vivo proton range verification using conventional PET scanners. In this technical note, we characterize those materials to test their usability in more clinically relevant conditions. Two phantoms made of low-density balsa wood (~0.1 g cm‑3) and beef (~1.0 g cm‑3) were embedded with Cu or 68Zn foils of several volumes (10–50 mm3). The metal foils were positioned at several depths in the dose fall-off region, which had been determined from our previous study. The phantoms were then irradiated with different proton doses (1–5 Gy). After irradiation, the phantoms with the embedded foils were moved to a diagnostic PET scanner and imaged. The acquired data were reconstructed with 20–40 min of scan time using various delay times (30–150 min) to determine the maximum contrast-to-noise ratio. The resultant PET/computed tomography (CT) fusion images of the activated foils were then examined and the foils’ PET signal strength/visibility was scored on a 5 point scale by 13 radiologists experienced in nuclear medicine. For both phantoms, the visibility of activated foils increased in proportion to the foil volume, dose, and PET scan time. A linear model was constructed with visibility scores as the response variable and all other factors (marker material, phantom material, dose, and PET scan time) as covariates. Using the linear model, volumes of foils that provided adequate visibility (score 3) were determined for each dose and PET scan time. The foil volumes that were determined will be used as a guideline in developing practical implantable markers.

  8. Characterizing proton-activated materials to develop PET-mediated proton range verification markers.

    PubMed

    Cho, Jongmin; Ibbott, Geoffrey S; Kerr, Matthew D; Amos, Richard A; Stingo, Francesco C; Marom, Edith M; Truong, Mylene T; Palacio, Diana M; Betancourt, Sonia L; Erasmus, Jeremy J; DeGroot, Patricia M; Carter, Brett W; Gladish, Gregory W; Sabloff, Bradley S; Benveniste, Marcelo F; Godoy, Myrna C; Patil, Shekhar; Sorensen, James; Mawlawi, Osama R

    2016-06-01

    Conventional proton beam range verification using positron emission tomography (PET) relies on tissue activation alone and therefore requires particle therapy PET whose installation can represent a large financial burden for many centers. Previously, we showed the feasibility of developing patient implantable markers using high proton cross-section materials ((18)O, Cu, and (68)Zn) for in vivo proton range verification using conventional PET scanners. In this technical note, we characterize those materials to test their usability in more clinically relevant conditions. Two phantoms made of low-density balsa wood (~0.1 g cm(-3)) and beef (~1.0 g cm(-3)) were embedded with Cu or (68)Zn foils of several volumes (10-50 mm(3)). The metal foils were positioned at several depths in the dose fall-off region, which had been determined from our previous study. The phantoms were then irradiated with different proton doses (1-5 Gy). After irradiation, the phantoms with the embedded foils were moved to a diagnostic PET scanner and imaged. The acquired data were reconstructed with 20-40 min of scan time using various delay times (30-150 min) to determine the maximum contrast-to-noise ratio. The resultant PET/computed tomography (CT) fusion images of the activated foils were then examined and the foils' PET signal strength/visibility was scored on a 5 point scale by 13 radiologists experienced in nuclear medicine. For both phantoms, the visibility of activated foils increased in proportion to the foil volume, dose, and PET scan time. A linear model was constructed with visibility scores as the response variable and all other factors (marker material, phantom material, dose, and PET scan time) as covariates. Using the linear model, volumes of foils that provided adequate visibility (score 3) were determined for each dose and PET scan time. The foil volumes that were determined will be used as a guideline in developing practical implantable markers. PMID:27203621

  9. Characterizing proton-activated materials to develop PET-mediated proton range verification markers

    NASA Astrophysics Data System (ADS)

    Cho, Jongmin; Ibbott, Geoffrey S.; Kerr, Matthew D.; Amos, Richard A.; Stingo, Francesco C.; Marom, Edith M.; Truong, Mylene T.; Palacio, Diana M.; Betancourt, Sonia L.; Erasmus, Jeremy J.; DeGroot, Patricia M.; Carter, Brett W.; Gladish, Gregory W.; Sabloff, Bradley S.; Benveniste, Marcelo F.; Godoy, Myrna C.; Patil, Shekhar; Sorensen, James; Mawlawi, Osama R.

    2016-06-01

    Conventional proton beam range verification using positron emission tomography (PET) relies on tissue activation alone and therefore requires particle therapy PET whose installation can represent a large financial burden for many centers. Previously, we showed the feasibility of developing patient implantable markers using high proton cross-section materials (18O, Cu, and 68Zn) for in vivo proton range verification using conventional PET scanners. In this technical note, we characterize those materials to test their usability in more clinically relevant conditions. Two phantoms made of low-density balsa wood (~0.1 g cm-3) and beef (~1.0 g cm-3) were embedded with Cu or 68Zn foils of several volumes (10-50 mm3). The metal foils were positioned at several depths in the dose fall-off region, which had been determined from our previous study. The phantoms were then irradiated with different proton doses (1-5 Gy). After irradiation, the phantoms with the embedded foils were moved to a diagnostic PET scanner and imaged. The acquired data were reconstructed with 20-40 min of scan time using various delay times (30-150 min) to determine the maximum contrast-to-noise ratio. The resultant PET/computed tomography (CT) fusion images of the activated foils were then examined and the foils’ PET signal strength/visibility was scored on a 5 point scale by 13 radiologists experienced in nuclear medicine. For both phantoms, the visibility of activated foils increased in proportion to the foil volume, dose, and PET scan time. A linear model was constructed with visibility scores as the response variable and all other factors (marker material, phantom material, dose, and PET scan time) as covariates. Using the linear model, volumes of foils that provided adequate visibility (score 3) were determined for each dose and PET scan time. The foil volumes that were determined will be used as a guideline in developing practical implantable markers.

  10. MicroRNA 141 Expression Is a Potential Prognostic Marker of Biliary Tract Cancers

    PubMed Central

    Kim, Jaihwan; Ryu, Ji Kon; Lee, Sang Hyub; Kim, Yong-Tae

    2016-01-01

    Background/Aims In recent years, a large number of micro-ribonucleic acids (miRNAs) have been identified as putative prognostic biomarkers for solid cancers because of their role in controlling the expression of oncogenes and tumor suppressor genes. The aim of this study was to verify the utility of miRNA 141 as a prognostic biomarker of biliary tract cancers. Methods From June 2010 to June 2012, common bile duct cancer tissue samples and matched noncancerous tissues from the ampulla of Vater were obtained from patients with biliary tract cancer undergoing endoscopic retrograde cholangiopancreatography. Using quantitative real-time polymerase chain reaction assays, we measured the mean relative expression levels of miRNA 141 in both groups of tissues. Overexpression of miRNA 141 was defined as a greater than 2-fold increase in expression levels as determined by the 2−ΔΔCt method. Results In a cohort of 38 patients with biliary tract cancers (seven gallbladder, 13 hilar, and 18 distal bile duct cancers), 26 patients (68.4%) were male, and the median age was 69.5 (52 to 85) years. Nineteen patients (50%) had undergone R0 resection procedures, including three Whipple operations, seven pylorus-preserving pancreaticoduodenectomies, six bile duct resections, and three extended lobectomies. Among the patients who had undergone R0 resection, the overexpression of miRNA 141 was significantly associated with shorter disease-free survival and a greater risk of angiolymphatic invasion. Among the patients who did not undergo R0 resection, miRNA 141 overexpression was significantly associated with reduced overall survival. Conclusions Overexpression of miRNA 141 is an indicator of a poor prognosis in patients with biliary tract cancer, suggesting that miRNA 141 may be a valuable prognostic biomarker of this disease. PMID:27172928

  11. Changes of neural markers expression during late neurogenic differentiation of human adipose-derived stem cells

    PubMed Central

    Razavi, Shahnaz; Khosravizadeh, Zahra; Bahramian, Hamid; Kazemi, Mohammad

    2015-01-01

    Background: Different studies have been done to obtain sufficient number of neural cells for treatment of neurodegenerative diseases, spinal cord, and traumatic brain injury because neural stem cells are limited in central nerves system. Recently, several studies have shown that adipose-derived stem cells (ADSCs) are the appropriate source of multipotent stem cells. Furthermore, these cells are found in large quantities. The aim of this study was an assessment of proliferation and potential of neurogenic differentiation of ADSCs with passing time. Materials and Methods: Neurosphere formation was used for neural induction in isolated human ADSCs (hADSCs). The rate of proliferation was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and potential of neural differentiation of induced hADSCs was evaluated by immunocytochemical and real-time reverse transcription polymerase chain reaction analysis after 10 and 14 days post-induction. Results: The rate of proliferation of induced hADSCs increased after 14 days while the expression of nestin, glial fibrillary acidic protein, and microtubule-associated protein 2 was decreased with passing time during neurogenic differentiation. Conclusion: These findings showed that the proliferation of induced cells increased with passing time, but in early neurogenic differentiation of hADSCs, neural expression was higher than late of differentiation. Thus, using of induced cells in early differentiation may be suggested for in vivo application. PMID:26605238

  12. Serum Copper as a Marker of Disease Activity in Rheumatoid Arthritis

    PubMed Central

    Chakraborty, Montosh; Changkakati, Rita

    2015-01-01

    Introduction Copper is an important trace element for normal growth and development of the body. It is also essential for maturation of collagen tissues. The purpose of the study was to estimate the serum copper levels in rheumatoid arthritis patients and to see its association with the various parameters of disease activity. Materials and Methods The study was carried out among 50 diagnosed rheumatoid arthritis patients (25 each of active disease & remission patients) and 50 age and sex matched controls. Fasting blood sample was collected for estimation of serum copper, haemoglobin level and ESR in the subjects. Results Mean serum copper level in the case group was found to be significantly higher than that of the control group (p-value<0.001). This increase of copper level was more in active disease than those with remission (p-value < 0.0001). A significant positive correlation was found between serum copper level and ESR, serum copper level and morning stiffness and a negative correlation was found between serum copper level and haemoglobin level in rheumatoid arthritis patients. Conclusion In rheumatoid arthritis patients, serum copper level may be used as an additional biochemical marker for estimation of disease activity. PMID:26816881

  13. Antimicrobial Activity of Euplotin C, the Sesquiterpene Taxonomic Marker from the Marine Ciliate Euplotes crassus

    PubMed Central

    Savoia, Dianella; Avanzini, Claudio; Allice, Tiziano; Callone, Emanuela; Guella, Graziano; Dini, Fernando

    2004-01-01

    Strains of the marine ciliate protist Euplotes crassus produce exclusive terpenoids called euplotins that play an ecological role. Among these derivatives, euplotin C is the main of four secondary metabolites isolated from cultures of this protozoon and represents the sesquiterpene taxonomic marker from E. crassus. Because different terpenoid metabolites of plant origin showed a certain antimicrobial activity, we assessed the compound euplotin C, purified by high-pressure liquid chromatography and solubilized in two solubility enhancers, against the protozoa Leishmania major and Leishmani infantum, the fungus Candida albicans, and nine strains of gram-positive and gram-negative microorganisms. An activity of euplotin C against Leishmania promastigotes was demonstrated (50% lethal doses were 4.6 or 8.1 μg/ml depending on the agent used to solubilize the compound), while the effect was less evident on Candida and nearly absent on bacteria. A nonsignificant cytotoxicity (50% lethal dose, >200 μg/ml) against the J774 cell line was observed. A leishmanicidal activity was also shown by the living, euplotin-producing cells of E. crassus cultured together with promastigotes; this activity increased with time from 10 min to 6 h of incubation. This study provides an initial rationale for the evaluation of euplotin C and other similar natural products as alternative or possibly synergistic compounds for current antiprotozoon chemotherapeutics. PMID:15388442

  14. Expression of AKR1C3 and CNN3 as markers for detection of lymph node metastases in colorectal cancer.

    PubMed

    Nakarai, Chiaki; Osawa, Kayo; Akiyama, Minami; Matsubara, Nagahide; Ikeuchi, Hiroki; Yamano, Tomoki; Hirota, Seiichi; Tomita, Naohiro; Usami, Makoto; Kido, Yoshiaki

    2015-08-01

    The aim of the study was to identify a set of discriminating genes that could be used for the prediction of Lymph node (LN) metastasis in human colorectal cancer (CRC), and for this, we compared the whole genome profiles of two CRC cell lines (the primary cell line SW480 and its LN metastatic variant, SW620) and identified eight genes [S100 calcium-binding protein P; aldo-keto reductase family 1(AKR1), member B1 (aldose reductase; AKR1B1); AKR1, member C3 (AKR1C3); calponin 3, acidic; metastasis associated in colon cancer 1; hemoglobin, epsilon 1; trefoil factor 3; and FGGY carbohydrate kinase domain containing]. These genes were examined by quantitative RT-PCR in tissues and LNs in 14 CRC patients and 11 control patients. The level of AKR1C3 mRNA expression was significantly different between the Dukes' stage A, B, and C groups and the control group (p < 0.05, p < 0.001, and p < 0.001) and was also significantly different between Dukes' stage C and A or B groups (p < 0.05 and p < 0.001, respectively). The expression of CNN3 was significantly different between the Dukes' stage C and B or control groups (p < 0.001 and p < 0.01, respectively). There were significant correlations between the expression levels of AKR1C3 and CNN3. AKR1C3 and CNN3 expressions are more accurate and suitable markers for the diagnosis of LN metastasis than the other six genes examined in this study.

  15. Expression of KIT (CD117) in neoplasms of the head and neck: an ancillary marker for adenoid cystic carcinoma.

    PubMed

    Mino, M; Pilch, B Z; Faquin, W C

    2003-12-01

    Adenoid cystic carcinoma is an indolent salivary gland malignancy that is associated with a poor long-term prognosis. The distinction of adenoid cystic carcinoma from other head and neck neoplasms can occasionally be problematic, particularly in small biopsies. Recent studies suggest that KIT (CD117) might be useful as an ancillary marker for adenoid cystic carcinoma; however, the expression of KIT in other benign and malignant head and neck neoplasms, including those that might mimic adenoid cystic carcinoma, has not been well studied. Here we use two different antibodies against KIT to evaluate its expression in a series of 66 adenoid cystic carcinomas compared with its expression in 98 other neoplasms of the head and neck. Overall, 94% (n = 62) of adenoid cystic carcinomas from various anatomic sites and of various histologic subtypes were positive for at least one of the KIT antibodies, and 77% (n = 50) of adenoid cystic carcinoma cases were positive for both antibodies. This contrasted with only 8% (n = 8) of other head and neck neoplasms that were positive for both KIT antibodies (P <.001). It was of note that certain neoplasms, including pleomorphic adenoma, basal cell adenoma, polymorphous low-grade adenocarcinoma, and basal cell carcinoma, that can show histologic overlap with adenoid cystic carcinoma had significantly less KIT immunoreactivity than did adenoid cystic carcinoma (P <.001). In contrast, KIT expression did not reliably distinguish adenoid cystic carcinoma from basal cell adenocarcinoma and basaloid squamous carcinoma (P >.05). The overall sensitivity of the two KIT antibodies for adenoid cystic carcinoma was 82-89%, and the specificity was 87-88%. The findings in this study support the potential use of KIT immunoexpression for distinguishing adenoid cystic carcinoma from many other benign and malignant head and neck neoplasms.

  16. Glucose-induced serum- and glucocorticoid-regulated kinase activation in oncofetal fibronectin expression

    SciTech Connect

    Khan, Zia A.; Barbin, Yousef P.; Farhangkhoee, Hana; Beier, Norbert; Scholz, Wolfgang; Chakrabarti, Subrata . E-mail: subrata.chakrabarti@fmd.uwo.ca

    2005-04-01

    Preferential expression of oncofetal extra domain-B fibronectin (EDB{sup +} FN), a proposed angiogenic marker, has been shown in proliferative diabetic retinopathy. High levels of glucose also increase EDB{sup +} FN expression in endothelial cells (ECs) via transforming growth factor-{beta}1 (TGF-{beta}1) and endothelin-1 (ET-1). The present study was aimed at elucidating the role of serum- and glucocorticoid-regulated kinase (SGK-1) in glucose-induced EDB{sup +} FN expression. Using human macro- and microvascular ECs, we show that high levels of glucose, TGF-{beta}1, and ET-1 increase the EDB{sup +} FN expression via SGK-1 alteration at the mRNA, protein, and activity levels. Inhibition of TGF-{beta}1 and ET-1 prevented glucose-induced SGK-1 activation and the EDB{sup +} FN expression. Furthermore, using siRNA-mediated SGK-1 gene silencing, we show that glucose-induced EDB{sup +} FN expression can be completely prevented. These findings provide first evidence of glucose-induced SGK-1 activation in altered EDB{sup +} FN expression and provide novel avenues for therapeutic modalities.

  17. Identification of novel markers of alternative activation and potential endogenous PPARγ ligand production mechanisms in human IL-4 stimulated differentiating macrophages.

    PubMed

    Czimmerer, Zsolt; Varga, Tamas; Poliska, Szilard; Nemet, Istvan; Szanto, Attila; Nagy, Laszlo

    2012-12-01

    We analyzed global gene expression profiles of IL-4 induced alternatively activated as well as IFNγ+TNFα stimulated classically activated human monocyte derived macrophages and identified novel IL-4 regulated alternative activation marker genes including MS4A4A, SLA, CD180, and ENPP2. Transcription factor prediction analysis of IL-4 regulated genes suggested that the regulated genes are involved in a complex regulation of lipid metabolism, defense against cell metabolism derived reactive oxygen species, and basal expression of inflammation linked genes. Both an in silico transcription activation prediction as well as experimental data suggested the presence of alternative macrophage activation specific endogenous PPARγ ligand producing mechanisms. We found the induction of three enzymes whose activity can potentially generate endogenous PPARγ ligands in an IL-4 dependent manner. These are MAOA, ENPP2, and ALOX15 producing 5-methoxy-indole acetate, lysophosphatidic acid (LPA) and 13-hydroxyoctadienoic acid (13-HODE), and/or 15-hydroxyeicosatetraenoic acid (15-HETE), respectively. Our data suggest that global gene expression profiling, combined with computational transcription activity prediction, can lead to identification of transcriptional networks that underpin cellular subtype specification.

  18. Expression of MMP-1/PAR-1 and patterns of invasion in oral squamous cell carcinoma as potential prognostic markers

    PubMed Central

    Fan, Hai-Xia; Chen, Yan; Ni, Bo-Xiong; Wang, Shan; Sun, Miao; Chen, Dong; Zheng, Jin-Hua

    2015-01-01

    Background Matrix metalloproteinase (MMP)-1 degrades type I collagen of the extracellular matrix and also activates protease activated receptor (PAR)-1 to induce angiogenesis. The aims of this study were to evaluate microvessel density (MVD) and the expression of PAR-1 and MMP-1 in oral squamous cell carcinoma (SCC) specimens with different patterns of invasion (POI) and to evaluate their association with clinical outcomes. Methods Seventy-four surgically obtained oral SCC samples were classified by POI according to hematoxylin-eosin staining. MVD and the localization and intensity of PAR-1 and MMP-1 expression were detected by immunohistochemistry. Results Of the 74 oral SCC samples, 18, 5, 34, and 17 showed type I, II, III, and IV POI, respectively. MVD and expression levels of MMP-1 and PAR-1 differed between POI types I–II and POI types III–IV. Patients with low tumor expression of MMP-1 and PAR-1 and low MVD had a longer survival time than those with high tumor expression of MMP-1 and PAR-1. Moreover, the survival time of patients with POI types III–IV was shorter than that of patients with POI types I–II. Conclusion POI combined with expression levels of MMP-1 and PAR-1 may be a valuable tool for assessing the clinical prognosis of patients with oral SCC. PMID:26170698

  19. Differential expression of embryonic epicardial progenitor markers and localization of cardiac fibrosis in adult ischemic injury and hypertensive heart disease.

    PubMed

    Braitsch, Caitlin M; Kanisicak, Onur; van Berlo, Jop H; Molkentin, Jeffery D; Yutzey, Katherine E

    2013-12-01

    During embryonic heart development, the transcription factors Tcf21, Wt1, and Tbx18 regulate activation and differentiation of epicardium-derived cells, including fibroblast lineages. Expression of these epicardial progenitor factors and localization of cardiac fibrosis were examined in mouse models of cardiovascular disease and in human diseased hearts. Following ischemic injury in mice, epicardial fibrosis is apparent in the thickened layer of subepicardial cells that express Wt1, Tbx18, and Tcf21. Perivascular fibrosis with predominant expression of Tcf21, but not Wt1 or Tbx18, occurs in mouse models of pressure overload or hypertensive heart disease, but not following ischemic injury. Areas of interstitial fibrosis in ischemic and hypertensive hearts actively express Tcf21, Wt1, and Tbx18. In all areas of fibrosis, cells that express epicardial progenitor factors are distinct from CD45-positive immune cells. In human diseased hearts, differential expression of Tcf21, Wt1, and Tbx18 also is detected with epicardial, perivascular, and interstitial fibrosis, indicating conservation of reactivated developmental mechanisms in cardiac fibrosis in mice and humans. Together, these data provide evidence for distinct fibrogenic mechanisms that include Tcf21, separate from Wt1 and Tbx18, in different fibroblast populations in response to specific types of cardiac injury.

  20. Development and Validation of Single Nucleotide Polymorphism (SNP) Markers from an Expressed Sequence Tag (EST) Database in Olive Flounder (Paralichthys olivaceus).

    PubMed

    Kim, Jung Eun; Lee, Young Mee; Lee, Jeong-Ho; Noh, Jae Koo; Kim, Hyun Chul; Park, Choul-Ji; Park, Jong-Won; Kim, Kyung-Kil

    2014-12-01

    To successful molecular breeding, identification and functional characterization of breeding related genes and development of molecular breeding techniques using DNA markers are essential. Although the development of a useful marker is difficult in the aspect of time, cost and effort, many markers are being developed to be used in molecular breeding and developed markers have been used in many fields. Single nucleotide polymorphisms (SNPs) markers were widely used for genomic research and breeding, but has hardly been validated for screening functional genes in olive flounder. We identified single nucleotide polymorphisms (SNPs) from expressed sequence tag (EST) database in olive flounder; out of a total 4,327 ESTs, 693 contigs and 514 SNPs were detected in total EST, and these substitutions include 297 transitions and 217 transversions. As a result, 144 SNP markers were developed on the basis of 514 SNP to selection of useful gene region, and then applied to each of eight wild and culture olive flounder (total 16 samples). In our experimental result, only 32 markers had detected polymorphism in sample, also identified 21 transitions and 11 transversions, whereas indel was not detected in polymorphic SNPs. Heterozygosity of wild and cultured olive flounder using the 32 SNP markers is 0.34 and 0.29, respectively. In conclusion, we identified SNP and polymorphism in olive flounder using newly designed marker, it supports that developed markers are suitable for SNP detection and diversity analysis in olive flounder. The outcome of this study can be basic data for researches for immunity gene and characteristic with SNP.

  1. The relationship between clinicopathological features and expression of epithelial and mesenchymal markers in spontaneous canine mammary gland tumors.

    PubMed

    Yoshida, Kota; Yoshida, Saori; Choisunirachon, Nan; Saito, Tomochika; Matsumoto, Kaori; Saeki, Kohei; Mochizuki, Manabu; Nishimura, Ryohei; Sasaki, Nobuo; Nakagawa, Takayuki

    2014-10-01

    It is known that epithelial mesenchymal transition (EMT) contributes to the acquisition of malignant property in human cancers. However, the role of EMT in canine tumors remains to be elucidated. To evaluate the correlation between expression levels of protein markers involved in EMT and clinicopathological characteristics in canine mammary gland tumors, immunohistochemistry using antibodies against ZO-1, E-cadherin, vimentin, N-cadherin and fibronectin was performed on 119 clinical tissue samples. Consequently, loss of ZO-1 and E-cadherin, and gain of vimentin and N-cadherin were more frequently observed in malignant tumors than in benign tumors. However, there was no correlation among expression of these molecules. Univariate and multivariate analysis identified that loss of E-cadherin independently had a low one-year survival rate (adjusted odds ratio: 2.3, P=0.02). These results suggested that EMT might relate to acquisition of malignancy, and additionally, E-cadherin was strongly correlated with malignant behavior in canine mammary gland tumors.

  2. Identification of potential transcriptomic markers in developing ankylosing spondylitis: a meta-analysis of gene expression profiles.

    PubMed

    Fang, Fang; Pan, Jian; Xu, Lixiao; Li, Gang; Wang, Jian

    2015-01-01

    The goal of this study was to identify potential transcriptomic markers in developing ankylosing spondylitis by a meta-analysis of multiple public microarray datasets. Using the INMEX (integrative meta-analysis of expression data) program, we performed the meta-analysis to identify consistently differentially expressed (DE) genes in ankylosing spondylitis and further performed functional interpretation (gene ontology analysis and pathway analysis) of the DE genes identified in the meta-analysis. Three microarray datasets (26 cases and 29 controls in total) were collected for meta-analysis. 905 consistently DE genes were identified in ankylosing spondylitis, among which 482 genes were upregulated and 423 genes were downregulated. The upregulated gene with the smallest combined rank product (RP) was GNG11 (combined RP=299.64). The downregulated gene with the smallest combined RP was S100P (combined RP=335.94). In the gene ontology (GO) analysis, the most significantly enriched GO term was "immune system process" (P=3.46×10(-26)). The most significant pathway identified in the pathway analysis was antigen processing and presentation (P=8.40×10(-5)). The consistently DE genes in ankylosing spondylitis and biological pathways associated with those DE genes identified provide valuable information for studying the pathophysiology of ankylosing spondylitis.

  3. Hepatic cancer stem cell marker granulin-epithelin precursor and β-catenin expression associate with recurrence in hepatocellular carcinoma

    PubMed Central

    Cheung, Phyllis F.Y.; Cheung, Tan To; Yip, Chi Wai; Ng, Linda W.C.; Fung, Sze Wai; Lo, Chung Mau; Fan, Sheung Tat; Cheung, Siu Tim

    2016-01-01

    Granulin-epithelin precursor (GEP) has been demonstrated to confer enhanced cancer stem-like cell properties in hepatocellular carcinoma (HCC) cell line models in our previous studies. Here, we aimed to examine the GEP-expressing cells in relation to the stem cell related molecules and stem-like cell properties in the prospective HCC clinical cohort. GEP protein levels were significantly higher in HCCs than the paralleled non-tumor liver tissues, and associated with venous infiltration. GEPhigh cells isolated from clinical HCC samples exhibited higher levels of stem cell marker CD133, pluripotency-associated signaling molecules β-catenin, Oct4, SOX2, Nanog, and chemodrug transporter ABCB5. In addition, GEPhigh cells possessed preferential ability to form colonies and spheroids, and enhanced in vivo tumor-initiating ability while their xenografts were able to be serially subpassaged into secondary mouse recipients. Expression levels of GEP and pluripotency-associated genes were further examined in the retrospective HCC cohort and demonstrated significant correlation of GEP with β-catenin. Notably, HCC patients with high GEP and β-catenin levels demonstrated poor recurrence-free survival. In summary, GEP-positive HCC cells directly isolated from clinical specimens showed β-catenin elevation and cancer stem-like cell properties. PMID:26942873

  4. Stable RNA markers for identification of blood and saliva stains revealed from whole genome expression analysis of time-wise degraded samples.

    PubMed

    Zubakov, Dmitry; Hanekamp, Eline; Kokshoorn, Mieke; van Ijcken, Wilfred; Kayser, Manfred

    2008-03-01

    Human body fluids such as blood and saliva represent the most common source of biological material found at a crime scene. Reliable tissue identification in forensic science can reveal significant insights into crime scene reconstruction and can thus contribute toward solving crimes. Limitations of existing presumptive tests for body fluid identification in forensics, which are usually based on chemoluminescence or protein analysis, are expected to be overcome by RNA-based methods, provided that stable RNA markers with tissue-specific expression patterns are available. To generate sets of stable RNA markers for reliable identification of blood and saliva stains we (1) performed whole-genome gene expression analyses on a series of time-wise degraded blood and saliva stain samples using the Affymetrix U133 plus2 GeneChip, (2) consulted expression databases to obtain additional information on tissue specificity, and (3) confirmed expression patterns of the most promising candidate genes by quantitative real-time polymerase chain reaction including additional forensically relevant tissues such as semen and vaginal secretion. Overall, we identified nine stable mRNA markers for blood and five stable mRNA markers for saliva detection showing tissue-specific expression signals in stains aged up to 180 days of age, expectedly older. Although, all of the markers were able to differentiate blood/saliva from semen samples, none of them could differentiate vaginal secretion because of the complex nature of vaginal secretion and the biological similarity of buccal and vaginal mucosa. We propose the use of these 14 stable mRNA markers for identification of blood and saliva stains in future forensic practice. PMID:17579879

  5. Effect of Cryopreserved Amniotic Membrane Orientation on the Expression of Limbal Mesenchymal and Epithelial Stem Cell Markers in Prolonged Limbal Explant Cultures

    PubMed Central

    Lužnik, Zala; Hawlina, Marko; Maličev, Elvira; Bertolin, Marina; Kopitar, Andreja Nataša; Ihan, Alojz; Ferrari, Stefano; Schollmayer, Petra

    2016-01-01

    Purpose To evaluate the effect of prolonged limbal explants cultured without any scaffolds or on amniotic membrane (AM) on the viability, proliferation and differentiation potential of putative phenotypically defined cultured limbal mesenchymal (LMSC) and epithelial stem cells (LESC). Methods Limbal explants were cultivated on cryopreserved intact AM or plastic plates using medium supplemented with only human serum. AM was positioned with either the epithelial or stromal side up. The outgrowing cells were immunophenotyped for the co-expression of mesenchymal stem cell markers (CD73/CD90/CD105 positive and CD45 negative), proliferation and putative progenitor markers (CXCR4, CD117), epithelial markers and antigen presenting cell markers (CD80, CD83, CD86) by flow cytometry. Immunohistochemistry on limbal cultures cultivated on AM was carried out with antibodies against pan-cytokeratin, p63, Ki67. Results Morphological and immunostaining analyses revealed two distinct stem cell population types, which could be identified over prolonged culturing time periods. Expression of LMSC markers and CXCR4 was significantly higher (p < 0.05) in cultures cultivated without AM. However, no statistically significant difference was observed in CD117 expression. The cells cultivated on AM retained an epithelial cell structure, which was further confirmed by histology examination. Histology revealed limbal epithelial growth and p63, Ki67 positive cells on both sides of AM. Conclusion Limbal cells cultivated on AM exhibited a lower expression profile of LMSC and CXCR4 markers as limbal cells cultivated on plastic culture plates. However, CD117 expression was similar. Histology confirmed limbal epithelial cell growth on both sides of AM, with no morphological differences, or positivity of cells for p63 and Ki67. PMID:27723792

  6. Stable RNA markers for identification of blood and saliva stains revealed from whole genome expression analysis of time-wise degraded samples.

    PubMed

    Zubakov, Dmitry; Hanekamp, Eline; Kokshoorn, Mieke; van Ijcken, Wilfred; Kayser, Manfred

    2008-03-01

    Human body fluids such as blood and saliva represent the most common source of biological material found at a crime scene. Reliable tissue identification in forensic science can reveal significant insights into crime scene reconstruction and can thus contribute toward solving crimes. Limitations of existing presumptive tests for body fluid identification in forensics, which are usually based on chemoluminescence or protein analysis, are expected to be overcome by RNA-based methods, provided that stable RNA markers with tissue-specific expression patterns are available. To generate sets of stable RNA markers for reliable identification of blood and saliva stains we (1) performed whole-genome gene expression analyses on a series of time-wise degraded blood and saliva stain samples using the Affymetrix U133 plus2 GeneChip, (2) consulted expression databases to obtain additional information on tissue specificity, and (3) confirmed expression patterns of the most promising candidate genes by quantitative real-time polymerase chain reaction including additional forensically relevant tissues such as semen and vaginal secretion. Overall, we identified nine stable mRNA markers for blood and five stable mRNA markers for saliva detection showing tissue-specific expression signals in stains aged up to 180 days of age, expectedly older. Although, all of the markers were able to differentiate blood/saliva from semen samples, none of them could differentiate vaginal secretion because of the complex nature of vaginal secretion and the biological similarity of buccal and vaginal mucosa. We propose the use of these 14 stable mRNA markers for identification of blood and saliva stains in future forensic practice.

  7. Study of the cytotoxic activity of Styrax camporum extract and its chemical markers, egonol and homoegonol.

    PubMed

    de Oliveira, Pollyanna Francielli; Damasceno, Jaqueline Lopes; Bertanha, Camila Spereta; Araújo, Alba Regina Barbosa; Pauletti, Patrícia Mendonça; Tavares, Denise Crispim

    2016-08-01

    The benzofuran lignans egonol and homoegonol are found in all species of the genus Styrax. Since natural products are important sources of new anticancer drugs, this study evaluated the cytotoxic activity of a hydroalcoholic extract of the stems of S. camporum (SCHE) and their chemical markers, egonol (EG) and homoegonol (HE), against different tumor cell lines (B16F10, MCF-7, HeLa, HepG2, and MO59J). A normal human cell line (GM07492A) was included. Cytotoxic activity was evaluated at different treatment times (24, 48 and 72 h) using the XTT assay. More effective results were observed after 72 h of treatment. The lowest IC50 values were found for the HepG2 cell line, ranging from 11.2 to 55.0 µg/mL. The combination of EG and HE exerted higher cytotoxic activity than SCHE or treatment with either lignan alone, with the lowest IC50 (13.31 µg/mL) being observed for the MCF-7 line. Furthermore, treatment with these lignans was significantly more cytotoxic for some tumor cell lines compared to the normal cell line, GM07492A, indicating selectivity. These results suggest that these lignans may be used to treat cancer without affecting normal cells.

  8. The Role of Power Doppler Ultrasonography as Disease Activity Marker in Rheumatoid Arthritis

    PubMed Central

    Bhasin, Shaloo; Cheung, Peter P.

    2015-01-01

    Structural damage in rheumatoid arthritis (RA) occurs early if inflammation is not treated promptly. Treatment targeted to reduce inflammation, in particular, that of synovial inflammation in the joints (synovitis), has been recommended as standard treat-to-target recommendations by rheumatologists. The goal is to achieve disease remission (i.e., no disease activity). Several accepted remission criteria have not always equated to the complete absence of true inflammation. Over the last decade, musculoskeletal ultrasonography has been demonstrated to detect subclinical synovitis not appreciated by routine clinical or laboratory assessments, with the Power Doppler modality allowing clinicians to more readily appreciate true inflammation. Thus, targeting therapy to Power Doppler activity may provide superior outcomes compared with treating to clinical targets alone, making it an attractive marker of disease activity in RA. However, more validation on its true benefits such as its benefits to patients in regard to patient related outcomes and issues with standardized training in acquisition and interpretation of power Doppler findings are required. PMID:26063952

  9. Neuronal activity topography parameters as a marker for differentiating vascular cognitive impairment in carotid stenosis.

    PubMed

    Shibata, Takashi; Musha, Toshimitu; Kubo, Michiya; Horie, Yukio; Asahi, Takashi; Kuwayama, Naoya; Kuroda, Satoshi; Hayashi, Karin; Kobayashi, Yohei; Tanaka, Mieko; Matsuzaki, Haruyasu; Asada, Takashi

    2014-10-01

    Previously, we reported on the differentiation between patients with Alzheimer disease and normal controls using a quantitative electroencephalographic technique called neuronal activity topography (NAT). In this technique, cerebral neuronal activities are characterized by the signal intensity and coherence (sNAT and vNAT, respectively). In the present study, we examined 47 patients with vascular cognitive impairment in carotid stenosis and 52 normal controls. All subjects underwent electroencephalography in a resting state with closed eyes for 5 minutes. Electroencephalographic markers of the differential likelihood, that is, the sensitivity-versus-specificity characteristics, sL(x:VCI-NLc) and vL(x:VCI-NLc), were assessed with neuronal activity topography and were compared between the 2 groups. sL(x:VCI-NLc) and vL(x:VCI-NLc) crossed each other at a cutoff value of the differential likelihood. Separation of the patients and controls was made with a sensitivity of 92% and 88%, as well as a false-positive rate of 8% and 12% for sL(x:VCI-NLc) and vL(x:VCI-NLc), respectively. Using sNAT, we accurately differentiated 92% patients with vascular cognitive impairment. We recommend that sNAT, rather than vNAT, should be used in detecting vascular cognitive impaired patients. PMID:25174560

  10. Subchronic exposure to leachate activates key markers linked with neurological disorder in Wistar male rat.

    PubMed

    Akintunde, J K; Oboh, G

    2015-12-01

    The linking of various environmental chemicals exposure to neurodegenerative disorders is current. This study was undertaken to elucidate the toxic effects and the underlying biochemical mechanism of leachate obtained from Elewi Odo municipal battery recycling site (EOMABRL) using key markers of neuronal damage in rat via an oral route. Analysis of the concentrations of heavy metals showed that lead, cadmium, nickel, chromium, manganese, and iron were higher than the acceptable limits set by the regulatory authority-the World Health Organization. Whereas, copper, zinc, and cobalt were lower than permissible limits. EOMABRL was administered at 0, 20, 40, 60, 80, and 100% concentrations to adult male rats for 60 days. An in vitro study was also carried out in the cerebellum to assess cholinesterase biochemistry assays. Following exposure, brain was collected to determine the antioxidant status. EOMABRL administration significantly increased superoxide dismutase (SOD) and catalase (CAT) activities, and a sequential decrease in reduced glutathione (GSH) level with a concomitant increase in the accumulation of hydrogen peroxide (H2O2) and malondialdehyde (MDA) level was observed, when compared with the control. The treated rat had a significant (P < 0.05) increase in the activities of acetycholinesterase (AChE) and butyrylcholinesterase (BuChE). Taken together, these findings conclude that some possible mechanisms by which EOMABRL elicits neuronal disorder in male rat could be through the activation of AChE and BuChE and induction of oxidative stress with necrosis of neuronal cells. PMID:26362636

  11. Exploiting expressed sequence tag databases for the development and characterization of gene-derived simple sequence repeat markers in the opium poppy (Papaver somniferum L.) for forensic applications.

    PubMed

    Lee, Eun Jung; Jin, Gang Nam; Lee, Kyung Lyong; Han, Myun Soo; Lee, Yang Han; Yang, Moon Sik

    2011-09-01

    Simple sequence repeat (SSR) markers in the opium poppy (Papaver somniferum L.) were identified from an expressed sequence tag (EST) database comprised of 20,340 sequences. In total, 2780 SSR-containing sequences were identified. The most frequent microsatellite had an AT/TA motif (37%). Twenty-two opium poppy EST-SSR markers were presently developed and polymorphisms of six markers (psom 2, 4, 12, 13, 17, and 22) were utilized in 135 individuals under narcotic control investigation. An average of three alleles per locus (range: 2-5 alleles) with a mean heterozygosity of 0.167 was detected. Six loci identified 29 unique profiles in 135 individuals. The EST-SSR markers exhibited small degrees of genetic differentiation (fixation index = 0.727, p < 0.001). Other variable markers will be needed to facilitate the forensic identification of the opium poppy for future cases. To determine the potential for cross-species amplification, six markers were tested in five Papaver genera species and two Eschscholzia genera. The psom 4 and psom 17 primer pair was transferable. This is the first study to report SSR markers of the opium poppy.

  12. Altered expression of glial and synaptic markers in the anterior hippocampus of behaviorally depressed female monkeys.

    PubMed

    Willard, Stephanie L; Hemby, Scott E; Register, Thomas C; McIntosh, Scot; Shively, Carol A

    2014-03-20

    The anterior hippocampus is associated with emotional functioning and hippocampal volume is reduced in depression. We reported reduced neuropil volume and number of glia in the dentate gyrus (DG) and cornu ammonis (CA)1 of the anterior hippocampus in behaviorally depressed adult female cynomolgus macaques. To determine the biochemical correlates of morphometric and behavioral differences between behaviorally depressed and nondepressed adult female monkeys, glial and synaptic transcripts and protein levels were assessed in the DG, CA3 and CA1 of the anterior hippocampus. Glial fibrillary acidic protein (GFAP) was increased whereas spinophilin and postsynaptic density (PSD)-95 protein were decreased in the CA1 of depressed monkeys. GFAP was reciprocally related to spinophilin and PSD-95 protein in the CA1. Gene expression of GFAP paralleled the protein changes observed in the CA1 and was inversely related to serum estradiol levels in depressed monkeys. These results suggest that behavioral depression in female primates is accompanied by astrocytic and synaptic protein alterations in the CA1. Moreover, these findings indicate a potential role for estrogen in modulating astrocyte-mediated impairments in synaptic plasticity.

  13. Altered Expression of Glial and Synaptic Markers in the Anterior Hippocampus of Behaviorally Depressed Female Monkeys

    PubMed Central

    Willard, Stephanie L.; Hemby, Scott E.; Register, Thomas C.; McIntosh, Scot; Shively, Carol A.

    2014-01-01

    The anterior hippocampus is associated with emotional functioning and hippocampal volume is reduced in depression. We reported reduced neuropil volume and number of glia in the dentate gyrus (DG) and cornu ammonis (CA)1 of the anterior hippocampus in behaviorally depressed adult female cynomolgus macaques. To determine the biochemical correlates of morphometric and behavioral differences between behaviorally depressed and nondepressed adult female monkeys, glial and synaptic transcripts and protein levels were assessed in the DG, CA3 and CA1 of the anterior hippocampus. Glial fibrillary acidic protein (GFAP) was increased whereas spinophilin and postsynaptic density (PSD)-95 protein were decreased in the CA1 of depressed monkeys. GFAP was reciprocally related to spinophilin and PSD-95 protein in the CA1. Gene expression of GFAP paralleled the protein changes observed in the CA1 and was inversely related to serum estradiol levels in depressed monkeys. These results suggest that behavioral depression in female primates is accompanied by astrocytic and synaptic protein alterations in the CA1. Moreover, these findings indicate a potential role for estrogen in modulating astrocyte-mediated impairments in synaptic plasticity. PMID:24440617

  14. Arginase activity - a marker of disease status in patients with visceral leishmaniasis in ethiopia.

    PubMed

    Abebe, Tamrat; Takele, Yegnasew; Weldegebreal, Teklu; Cloke, Tom; Closs, Ellen; Corset, Camille; Hailu, Asrat; Hailu, Workagegnehu; Sisay, Yifru; Corware, Karina; Corset, Margaux; Modolell, Manuel; Munder, Markus; Tacchini-Cottier, Fabienne; Müller, Ingrid; Kropf, Pascale

    2013-01-01

    The underlying mechanisms resulting in the profound immune suppression characteristic of human visceral leishmaniasis (VL) are not fully understood. Here, we tested the hypothesis that arginase, an enzyme associated with immunosuppression, is higher in patients with VL and contributes to impaired T cell responses. We recruited patients with VL before and after treatment and healthy controls and measured the arginase metabolism in the blood of these individuals. Our results show that arginase activity is significantly higher in the blood of patients with active VL as compared to controls. These high levels of arginase decline considerably once the patients are successfully treated. We identified the phenotype of arginase-expressing cells among PBMCs as neutrophils and show that their frequency was increased in PBMCs of patients before treatment; this coincides with reduced levels of L-arginine in the plasma and decreased expression levels of CD3ζ in T cells. PMID:23556019

  15. Identifying differentially expressed genes under heat stress and developing molecular markers in orchardgrass (Dactylis glomerata L.) through transcriptome analysis.

    PubMed

    Huang, L K; Yan, H D; Zhao, X X; Zhang, X Q; Wang, J; Frazier, T; Yin, G; Huang, X; Yan, D F; Zang, W J; Ma, X; Peng, Y; Yan, Y H; Liu, W

    2015-11-01

    Orchardgrass (Dactylis glomerata L.) is a long-lived, cool-season forage grass that is commonly used for hay production. Despite its economic importance, orchardgrass genome remains relatively unexplored. In this study, we used Illumina RNA sequencing to identify gene-associated molecular markers, including simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs), as well as heat stress-induced differentially expressed genes (DEGs) in two orchardgrass genotypes, 'Baoxing' (heat resistant) and '01998' (heat susceptible). Approximately 163 million high-quality trimmed reads were generated from 207 million raw reads using the Illumina HiSeq 2000 platform. A total of 126,846 unigenes were obtained after de novo assembly of the trimmed reads, and 40,078 unigenes were identified as coding sequences (CDSs). Based on the assembled unigenes, 669,300 high-quality SNPs, including 416,099 transitions and 257,736 transversions, were contained in 75,875 unigenes. In addition, a total of 8475 microsatellites were detected in 7764 unigenes. When placed under heat stress, the total number of DEGs in 'Baoxing' (3527) was higher than in '01998' (2649), indicating that in comparison with heat-susceptible '01998', heat-resistant 'Baoxing' seems to have more unigenes that respond to heat stress. The high-throughput transcriptome sequencing of orchardgrass under heat stress provides useful information for gene identification and for the development of SNP and SSR molecular markers. The comparison of DEGs under different periods of heat stress allowed us to identify a wealth of candidate DEGs that can be further analysed in order to determine the genetic mechanisms underlying heat tolerance in orchardgrass.

  16. Expression of Cell Competition Markers at the Interface between p53 Signature and Normal Epithelium in the Human Fallopian Tube

    PubMed Central

    Kito, Masahiko; Maeda, Daichi; Kudo-Asabe, Yukitsugu; Sato, Naoki; Shih, Ie-Ming; Wang, Tian-Li; Tanaka, Masamitsu; Terada, Yukihiro; Goto, Akiteru

    2016-01-01

    There is a growing body of evidence regarding cell competition between normal and mutant mammalian cells, which suggest that it may play a defensive role in the early phase of carcinogenesis. In vitro study in the past has shown that overexpression of vimentin in normal epithelial cells at the contact surface with transformed cells is essential for the cell competition involved in epithelial defense against cancer. In this study, we attempted to examine cell competition in human tissue in vivo by investigating surgically resected human fallopian tubes that contain p53 signatures and serous tubal intraepithelial lesions (STILs), a linear expansion of p53-immunopositive/TP53 mutant tubal epithelial cells that are considered as precursors of pelvic high grade serous carcinoma. Immunofluorescence double staining for p53 and the cell competition marker vimentin was performed in 21 sections of human fallopian tube tissue containing 17 p53 signatures and 4 STILs. The intensities of vimentin expression at the interface between p53-positive cells at the end of the p53 signature/STIL and adjacent p53-negative normal tubal epithelial cells were compared with the background tubal epithelium. As a result, the average vimentin intensity at the interfaces relative to the background intensity was 1.076 (95% CI, 0.9412 – 1.211 for p53 signature and 0.9790 (95% CI, 0.7206 – 1.237) for STIL. Thus, it can be concluded that overexpression of the cell competition marker vimentin are not observed in human tissue with TP53 alterations. PMID:27258067

  17. Markers of thrombogenesis are activated in unmedicated patients with acute psychosis: a matched case control study

    PubMed Central

    2011-01-01

    Background Antipsychotic treatment has been repeatedly found to be associated with an increased risk for venous thromboembolism in schizophrenia. The extent to which the propensity for venous thromboembolism is linked to antipsychotic medication alone or psychosis itself is unclear. The objective of this study was to determine whether markers of thrombogenesis are increased in psychotic patients who have not yet been treated with antipsychotic medication. Methods We investigated the plasma levels of markers indicating activation of coagulation (D-dimers and Factor VIII) and platelets (soluble P-selectin, sP-selectin) in an antipsychotic-naive group of fourteen men and eleven women with acute psychosis (age 29.1 ± 8.3 years, body mass index 23.6 ± 4.7), and twenty-five healthy volunteers were matched for age, gender and body mass index. Results D-dimers (median 0.38 versus 0.19 mg/l, mean 1.12 ± 2.38 versus 0.28 ± 0.3 mg/l; P = 0.003) and sP-selectin (median 204.1 versus 112.4 ng/ml, mean 209.9 ± 124 versus 124.1 ± 32; P = 0.0005) plasma levels were significantly increased in the group of patients with acute psychosis as compared with healthy volunteers. We found a trend (median 148% versus 110%, mean 160 ± 72.5 versus 123 ± 62.5; P = 0.062) of increased plasma levels of factor VIII in psychotic patients as compared with healthy volunteers. Conclusions The results suggest that at least a part of venous thromboembolic events in patients with acute psychosis may be induced by pathogenic mechanisms related to psychosis rather than by antipsychotic treatment. Finding an exact cause for venous thromboembolism in psychotic patients is necessary for its effective treatment and prevention. PMID:21199572

  18. Blood acylpeptide hydrolase activity is a sensitive marker for exposure to some organophosphate toxicants.

    PubMed

    Quistad, Gary B; Klintenberg, Rebecka; Casida, John E

    2005-08-01

    Acylpeptide hydrolase (APH) unblocks N-acetyl peptides. It is a major serine hydrolase in rat blood, brain, and liver detected by derivatization with (3)H-diisopropyl fluorophosphate (DFP) or a biotinylated fluorophosphonate. Although APH does not appear to be a primary target of acute poisoning by organophosphorus (OP) compounds, the inhibitor specificity of this secondary target is largely unknown. This study fills the gap and emphasizes blood APH as a potential marker of OP exposure. The most potent in vitro inhibitors for human erythrocyte and mouse brain APH are DFP (IC(50) 11-17 nM), chlorpyrifos oxon (IC(50) 21-71 nM), dichlorvos (IC(50) 230-560 nM), naled (IC(50) 370-870 nM), and their analogs with modified alkyl substituents. (3)H-diisopropyl fluorophosphate is a potent inhibitor of mouse blood and brain APH in vivo (ED(50) 0.09-0.2 mg/kg and 0.02-0.03 mg/l for ip and vapor exposure, respectively). Mouse blood and brain APH and blood butyrylcholinesterase (BChE) are of similar sensitivity to DFP in vitro and in vivo (ip and vapor exposure), but APH inhibition is much more persistent in vivo (still >80% inhibition after 4 days). The inhibitory potency of OP pesticides in vivo in mice varies from APH selective (dichlorvos, naled, and trichlorfon), to APH and BChE selective (profenofos and tribufos), to ChE selective or nonselective (many commercial insecticides). Sarin administered ip at a lethal dose to guinea pigs inhibits blood acetylcholinesterase and BChE completely but erythrocyte APH only partially. Blood APH activity is therefore a sensitive marker for exposure to some but not all OP pesticides and chemical warfare agents. PMID:15888665

  19. Gene expression fingerprint of uterine serous papillary carcinoma: identification of novel molecular markers for uterine serous cancer diagnosis and therapy

    PubMed Central

    Santin, A D; Zhan, F; Cane', S; Bellone, S; Palmieri, M; Thomas, M; Burnett, A; Roman, J J; Cannon, M J; Shaughnessy, J; Pecorelli, S

    2005-01-01

    Uterine serous papillary cancer (USPC) represents a rare but highly aggressive variant of endometrial cancer, the most common gynecologic tumour in women. We used oligonucleotide microarrays that interrogate the expression of some 10 000 known genes to profile 10 highly purified primary USPC cultures and five normal endometrial cells (NEC). We report that unsupervised analysis of mRNA fingerprints readily distinguished USPC from normal endometrial epithelial cells and identified 139 and 390 genes that exhibited >5-fold upregulation and downregulation, respectively, in primary USPC when compared to NEC. Many of the genes upregulated in USPC were found to represent adhesion molecules, secreted proteins and oncogenes, such as L1 cell adhesion molecule, claudin-3 and claudin-4, kallikrein 6 (protease M) and kallikrein 10 (NES1), interleukin-6 and c-erbB2. Downregulated genes in USPC included SEMACAP3, ras homolog gene family, member I (ARHI), and differentially downregulated in ovarian carcinoma gene 1. Quantitative RT–PCR was used to validate differences in gene expression between USPC and NEC for several of these genes. Owing to its potential as a novel therapeutic marker, expression of the high-affinity epithelial receptor for Clostridium perfringens enterotoxin (CPE) claudin-4 was further validated through immunohistochemical analysis of formalin-fixed paraffin-embedded specimens from which the primary USPC cultures were obtained, as well as an independent set of archival USPC specimens. Finally, the sensitivity of primary USPC to the administration of scalar doses of CPE in vitro was also demonstrated. Our results highlight the novel molecular features of USPC and provide a foundation for the development of new type-specific therapies against this highly aggressive variant of endometrial cancer. PMID:15785748

  20. Expression level of chromodomain Y (CDY): potential marker for prediction of sperm recovery in non-obstructive azoospermia

    PubMed Central

    Heydarian, Neda; Favaedi, Raha; Sadighi Gilani, Mohammad Ali; Shahhoseini, Maryam

    2016-01-01

    Background: The availability of testis specific genes will be of help in choosing the most promising biomarkers for the detection of testicular sperm retrieval in patients with non-obstructive azoospermia (NOA). Testis specific chromodomain protein Y 1 (CDY1) is a histone acetyltransferase which concentrates in the round spermatid nucleus, where histone hyperacetylation occurs and causes the replacement of histones by the sperm-specific DNA packaging proteins, TNPs and PRMs. Objective: The aim was to evaluate CDY1 gene as a marker for predicting of successful sperm retrieval in NOA patients. Materials and Methods: This research was conducted on 29 patients with NOA who had undergone testicular sperm extraction (TESE) procedure. NOA patients were subdivided into patients with successful sperm retrieval (NOA+, n=12) and patients with unsuccessful sperm retrieval (NOA-, n=17). Relative expression of CDY1 gene and chromatin incorporation of CDY1 protein were measured by quantitative real-time polymerase chain reaction (qRT-PCR) and ELISA assay, respectively. Results: Quantification of mRNA relative expression and incorporation of CDY1 protein in chromatin showed significant lower expressions and protein levels of CDY1 in testis tissues of NOA- in comparison to NOA+ group. Conclusion: The findings in this study demonstrated a correlation between the low levels of CDY1 function and unsuccessful sperm recovery in the testicular tissues of NOA- compared to NOA+ patients. Therefore, it can be reasonable to consider CDY1 as a potential biomarker for predicting the presence of spermatozoa, although the claim needs more samples to be confirmed. PMID:27525321

  1. The expression of renal Epstein-Barr virus markers in patients with lupus nephritis

    PubMed Central

    YU, XIAO-XIA; YAO, CUI-WEI; TAO, JING-LI; YANG, CHEN; LUO, MIAN-NA; LI, SHANG-MEI; LIU, HUA-FENG

    2014-01-01

    The aim of this study was to investigate the role of renal Epstein-Barr virus (EBV) infection in the pathogenesis of lupus nephritis (LN). A total of 58 renal tissue samples from patients with LN, seven normal renal tissue samples from patients with non-glomerular hematuria and 37 renal tissue samples from patients with minimal change nephropathy were collected. The expression of EBV-latent membrane protein-1 (EBV-LMP1) and EBV-encoded RNA 1 (EBER-1) in the renal tissue was examined by immunohistochemistry (IHC) and in situ hybridization (ISH), respectively. The sera levels of anti-nuclear antibody as well as antibodies to extractable nuclear antigen in patients with LN were also measured. An equivalence test showed that the results from the IHC and the ISH analyses had strong agreement. The positive rates of renal EBER-1 and EBV-LMP1 in the LN patients were significantly higher than those of the normal and minimal change nephropathy patients (P<0.001), while no significant difference was identified between those of the normal and minimal change nephropathy groups (P>0.05). The positive rates of EBV-LMP1 and EBER-1 in the renal tissues of patients with LN were not determined to be significantly different between the relapse (immunosuppressant-treated) and initial onset (non-treated) patients, between the patients with and without concurrent infection, and among the patients with different age ranges (P>0.05). The proportion of LN patients positive for anti-Sm antibody was significantly higher in the renal EBV-positive group than in the EBV-negative group (P<0.05), while the proportions of LN patients positive for the other autoantibodies that were examined were not identified to be significantly different between these two groups (P>0.05). The present study shows that renal EBV infection may contribute to the pathogenesis of LN by inducing anti-Sm antibody production. PMID:24940399

  2. Specific marker of feigned memory impairment: The activation of left superior frontal gyrus.

    PubMed

    Chen, Zi-Xiang; Xue, Li; Liang, Chun-Yu; Wang, Li-Li; Mei, Wei; Zhang, Qiang; Zhao, Hu

    2015-11-01

    Faking memory impairment means normal people complain lots of memory problems without organic damage in forensic assessments. Using alternative forced-choice paradigm, containing digital or autobiographical information, previous neuroimaging studies have indicated that faking memory impairment could cause the activation in the prefrontal and parietal regions, and might involve a fronto-parietal-subcortical circuit. However, it is still unclear whether different memory types have influence on faking or not. Since different memory types, such as long-term memory (LTM) and short-term memory (STM), were found supported by different brain areas, we hypothesized that feigned STM or LTM impairment had distinct neural activation mapping. Besides that, some common neural correlates may act as the general characteristic of feigned memory impairment. To verify this hypothesis, the functional magnetic resonance imaging (fMRI) combined with an alternative word forced-choice paradigm were used in this study. A total of 10 right-handed participants, in this study, had to perform both STW and LTM tasks respectively under answering correctly, answering randomly and feigned memory impairment conditions. Our results indicated that the activation of the left superior frontal gyrus and the left medial frontal gyrus was associated with feigned LTM impairment, whereas the left superior frontal gyrus, the left precuneus and the right anterior cingulate cortex (ACC) were highly activated while feigning STM impairment. Furthermore, an overlapping was found in the left superior frontal gyrus, and it suggested that the activity of the left superior frontal gyrus might be acting as a specific marker of feigned memory impairment. PMID:26479324

  3. Xanthurenic Acid Activates mGlu2/3 Metabotropic Glutamate Receptors and is a Potential Trait Marker for Schizophrenia.

    PubMed

    Fazio, Francesco; Lionetto, Luana; Curto, Martina; Iacovelli, Luisa; Cavallari, Michele; Zappulla, Cristina; Ulivieri, Martina; Napoletano, Flavia; Capi, Matilde; Corigliano, Valentina; Scaccianoce, Sergio; Caruso, Alessandra; Miele, Jessica; De Fusco, Antonio; Di Menna, Luisa; Comparelli, Anna; De Carolis, Antonella; Gradini, Roberto; Nisticò, Robert; De Blasi, Antonio; Girardi, Paolo; Bruno, Valeria; Battaglia, Giuseppe; Nicoletti, Ferdinando; Simmaco, Maurizio

    2015-12-08

    The kynurenine pathway of tryptophan metabolism has been implicated in the pathophysiology of psychiatric disorders, including schizophrenia. We report here that the kynurenine metabolite, xanturenic acid (XA), interacts with, and activates mGlu2 and mGlu3 metabotropic glutamate receptors in heterologous expression systems. However, the molecular nature of this interaction is unknown, and our data cannot exclude that XA acts primarily on other targets, such as the vesicular glutamate transporter, in the CNS. Systemic administration of XA in mice produced antipsychotic-like effects in the MK-801-induced model of hyperactivity. This effect required the presence of mGlu2 receptors and was abrogated by the preferential mGlu2/3 receptor antagonist, LY341495. Because the mGlu2 receptor is a potential drug target in the treatment of schizophrenia, we decided to measure serum levels of XA and other kynurenine metabolites in patients affected by schizophrenia. Serum XA levels were largely reduced in a large cohort of patients affected by schizophrenia, and, in patients with first-episode schizophrenia, levels remained low after 12 months of antipsychotic medication. As opposed to other kynurenine metabolites, XA levels were also significantly reduced in first-degree relatives of patients affected by schizophrenia. We suggest that lowered serum XA levels might represent a novel trait marker for schizophrenia.

  4. Xanthurenic Acid Activates mGlu2/3 Metabotropic Glutamate Receptors and is a Potential Trait Marker for Schizophrenia.

    PubMed

    Fazio, Francesco; Lionetto, Luana; Curto, Martina; Iacovelli, Luisa; Cavallari, Michele; Zappulla, Cristina; Ulivieri, Martina; Napoletano, Flavia; Capi, Matilde; Corigliano, Valentina; Scaccianoce, Sergio; Caruso, Alessandra; Miele, Jessica; De Fusco, Antonio; Di Menna, Luisa; Comparelli, Anna; De Carolis, Antonella; Gradini, Roberto; Nisticò, Robert; De Blasi, Antonio; Girardi, Paolo; Bruno, Valeria; Battaglia, Giuseppe; Nicoletti, Ferdinando; Simmaco, Maurizio

    2015-01-01

    The kynurenine pathway of tryptophan metabolism has been implicated in the pathophysiology of psychiatric disorders, including schizophrenia. We report here that the kynurenine metabolite, xanturenic acid (XA), interacts with, and activates mGlu2 and mGlu3 metabotropic glutamate receptors in heterologous expression systems. However, the molecular nature of this interaction is unknown, and our data cannot exclude that XA acts primarily on other targets, such as the vesicular glutamate transporter, in the CNS. Systemic administration of XA in mice produced antipsychotic-like effects in the MK-801-induced model of hyperactivity. This effect required the presence of mGlu2 receptors and was abrogated by the preferential mGlu2/3 receptor antagonist, LY341495. Because the mGlu2 receptor is a potential drug target in the treatment of schizophrenia, we decided to measure serum levels of XA and other kynurenine metabolites in patients affected by schizophrenia. Serum XA levels were largely reduced in a large cohort of patients affected by schizophrenia, and, in patients with first-episode schizophrenia, levels remained low after 12 months of antipsychotic medication. As opposed to other kynurenine metabolites, XA levels were also significantly reduced in first-degree relatives of patients affected by schizophrenia. We suggest that lowered serum XA levels might represent a novel trait marker for schizophrenia. PMID:26643205

  5. Xanthurenic Acid Activates mGlu2/3 Metabotropic Glutamate Receptors and is a Potential Trait Marker for Schizophrenia

    PubMed Central

    Fazio, Francesco; Lionetto, Luana; Curto, Martina; Iacovelli, Luisa; Cavallari, Michele; Zappulla, Cristina; Ulivieri, Martina; Napoletano, Flavia; Capi, Matilde; Corigliano, Valentina; Scaccianoce, Sergio; Caruso, Alessandra; Miele, Jessica; De Fusco, Antonio; Di Menna, Luisa; Comparelli, Anna; De Carolis, Antonella; Gradini, Roberto; Nisticò, Robert; De Blasi, Antonio; Girardi, Paolo; Bruno, Valeria; Battaglia, Giuseppe; Nicoletti, Ferdinando; Simmaco, Maurizio

    2015-01-01

    The kynurenine pathway of tryptophan metabolism has been implicated in the pathophysiology of psychiatric disorders, including schizophrenia. We report here that the kynurenine metabolite, xanturenic acid (XA), interacts with, and activates mGlu2 and mGlu3 metabotropic glutamate receptors in heterologous expression systems. However, the molecular nature of this interaction is unknown, and our data cannot exclude that XA acts primarily on other targets, such as the vesicular glutamate transporter, in the CNS. Systemic administration of XA in mice produced antipsychotic-like effects in the MK-801-induced model of hyperactivity. This effect required the presence of mGlu2 receptors and was abrogated by the preferential mGlu2/3 receptor antagonist, LY341495. Because the mGlu2 receptor is a potential drug target in the treatment of schizophrenia, we decided to measure serum levels of XA and other kynurenine metabolites in patients affected by schizophrenia. Serum XA levels were largely reduced in a large cohort of patients affected by schizophrenia, and, in patients with first-episode schizophrenia, levels remained low after 12 months of antipsychotic medication. As opposed to other kynurenine metabolites, XA levels were also significantly reduced in first-degree relatives of patients affected by schizophrenia. We suggest that lowered serum XA levels might represent a novel trait marker for schizophrenia. PMID:26643205

  6. Altered expression of oligodendrocyte and neuronal marker genes predicts the clinical onset of autoimmune encephalomyelitis and indicates the effectiveness of multiple sclerosis-directed therapeutics.

    PubMed

    Evangelidou, Maria; Karamita, Maria; Vamvakas, Sotiris-Spyros; Szymkowski, David E; Probert, Lesley

    2014-05-01

    Experimental autoimmune encephalomyelitis (EAE) is a valuable model for studying immunopathology in multiple sclerosis (MS) and for exploring the interface between autoimmune responses and CNS tissue that ultimately leads to lesion development. In this study, we measured gene expression in mouse spinal cord during myelin oligodendrocyte gp35-55 peptide-induced EAE, using quantitative RT-PCR, to identify gene markers that monitor individual hallmark pathological processes. We defined a small panel of genes whose longitudinal expression patterns provided insight into the timing, interrelationships, and mechanisms of individual disease processes and the efficacy of therapeutics for the treatment of MS. Earliest transcriptional changes were upregulation of Il17a and sharp downregulation of neuronal and oligodendrocyte marker genes preceding clinical disease onset, whereas neuroinflammatory markers progressively increased as symptoms and tissue lesions developed. EAE-induced gene-expression changes were not altered in mice deficient in IKKβ in cells of the myeloid lineage compared with controls, but the administration of a selective inhibitor of soluble TNF to mice from the day of immunization delayed changes in the expression of innate inflammation, myelin, and neuron markers from the presymptomatic phase. Proof of principle that the gene panel shows drug screening potential was obtained using a well-established MS therapeutic, glatiramer acetate. Prophylactic treatment of mice with glatiramer acetate normalized gene marker expression, and this correlated with the level of therapeutic success. These results show that neurons and oligodendrocytes are highly sensitive to CNS-directed autoimmunity before the development of clinical symptoms and immunopathology and reveal a role for soluble TNF in mediating the earliest changes in gene expression.

  7. Ethanol Activation of PKA Mediates Single-Minded 2 Expression in Neuronal Cells.

    PubMed

    Wang, Xiaolan; Yang, Zhihua; Sun, Yinan; Zhou, Hanjing; Chu, Guangpin; Zhang, Jing; Meng, Xianfang

    2015-12-01

    Prenatal ethanol exposure can cause extensive apoptotic neurodegeneration throughout the developing central nervous system (CNS), which results in cognitive deficits and memory decline. However, the underlying mechanisms need further study. Single-minded 2 (Sim2), a transcriptional repressor, is reportedly involved in diseases that impair learning and memory, such as Down syndrome (DS) and Alzheimer's disease. It is still unknown whether Sim2 is involved in regulating ethanol-mediated neuronal injury that might ultimately lead to neuronal dysfunction and subsequent learning and memory deficits. To study the effects of ethanol on Sim2 expression and neuronal injury, we used animal models and cell culture experiments. Our results indicated that in SH-SY5Y cells, ethanol exposure increased Sim2 expression and levels of cleaved caspase 3, which is a marker for cells undergoing apoptosis. Silencing Sim2 expression attenuated caspase 3 activation and cellular apoptosis. We also found that protein kinase A (PKA) activation induced Sim2 expression, as did ethanol. Inhibiting the PKA signaling pathway with H-89 decreased Sim2 expression and cleavage of caspase 3 that was induced by ethanol in vivo and in vitro. We further found that PKA regulated Sim2 expression at the transcriptional level. These results demonstrate that ethanol leads to increased Sim2 expression via the PKA pathway, ultimately resulting in apoptotic cell death.

  8. FOXA1 expression affects the proliferation activity of luminal breast cancer stem cell populations.

    PubMed

    Tachi, Kana; Shiraishi, Akira; Bando, Hiroko; Yamashita, Toshiharu; Tsuboi, Ikki; Kato, Toshiki; Hara, Hisato; Ohneda, Osamu

    2016-03-01

    The expression of estrogen receptor is the key in most breast cancers (BC) and binding of estrogen receptor to the genome correlates to Forkhead protein (FOXA1) expression. We herein assessed the correlation between the cancer stem cell (CSC) population and FOXA1 expression in luminal BC. We established luminal BC cells derived from metastatic pleural effusion and analyzed the potency of CSC and related factors with established luminal BC cell lines. We also confirmed that mammosphere cultures have an increased aldehyde dehydrogenase-positive population, which is one of the CSC markers, compared with adherent culture cells. Using a quantitative PCR analysis, we found that mammosphere forming cells showed a higher expression of FOXA1 and stemness-related genes compared with adherent culture cells. Furthermore, the growth activity and colony-forming activity of 4-hydroxytamoxifen-treated BC cells were inhibited in a mammosphere assay. Interestingly, 4-hydroxytamoxifen-resistant cells had significantly increased FOXA1 gene expression levels. Finally, we established short hairpin RNA of FOXA1 (shFOXA1) MCF-7 cells and investigated the relationship between self-renewal potential and FOXA1 expression. As a result, we found no significant difference in the number of mammospheres but decreased colony formation in shFOXA1 MCF-7 cells compared with control. These results suggest that the expression of FOXA1 appears to be involved in the proliferation of immature BC cells rather than the induction of stemness-related genes and self-renewal potency of CSCs.

  9. Diversity analysis in Cannabis sativa based on large-scale development of expressed sequence tag-derived simple sequence repeat markers.

    PubMed

    Gao, Chunsheng; Xin, Pengfei; Cheng, Chaohua; Tang, Qing; Chen, Ping; Wang, Changbiao; Zang, Gonggu; Zhao, Lining

    2014-01-01

    Cannabis sativa L. is an important economic plant for the production of food, fiber, oils, and intoxicants. However, lack of sufficient simple sequence repeat (SSR) markers has limited the development of cannabis genetic research. Here, large-scale development of expressed sequence tag simple sequence repeat (EST-SSR) markers was performed to obtain more informative genetic markers, and to assess genetic diversity in cannabis (Cannabis sativa L.). Based on the cannabis transcriptome, 4,577 SSRs were identified from 3,624 ESTs. From there, a total of 3,442 complementary primer pairs were designed as SSR markers. Among these markers, trinucleotide repeat motifs (50.99%) were the most abundant, followed by hexanucleotide (25.13%), dinucleotide (16.34%), tetranucloetide (3.8%), and pentanucleotide (3.74%) repeat motifs, respectively. The AAG/CTT trinucleotide repeat (17.96%) was the most abundant motif detected in the SSRs. One hundred and seventeen EST-SSR markers were randomly selected to evaluate primer quality in 24 cannabis varieties. Among these 117 markers, 108 (92.31%) were successfully amplified and 87 (74.36%) were polymorphic. Forty-five polymorphic primer pairs were selected to evaluate genetic diversity and relatedness among the 115 cannabis genotypes. The results showed that 115 varieties could be divided into 4 groups primarily based on geography: Northern China, Europe, Central China, and Southern China. Moreover, the coefficient of similarity when comparing cannabis from Northern China with the European group cannabis was higher than that when comparing with cannabis from the other two groups, owing to a similar climate. This study outlines the first large-scale development of SSR markers for cannabis. These data may serve as a foundation for the development of genetic linkage, quantitative trait loci mapping, and marker-assisted breeding of cannabis.

  10. Diversity analysis in Cannabis sativa based on large-scale development of expressed sequence tag-derived simple sequence repeat markers.

    PubMed

    Gao, Chunsheng; Xin, Pengfei; Cheng, Chaohua; Tang, Qing; Chen, Ping; Wang, Changbiao; Zang, Gonggu; Zhao, Lining

    2014-01-01

    Cannabis sativa L. is an important economic plant for the production of food, fiber, oils, and intoxicants. However, lack of sufficient simple sequence repeat (SSR) markers has limited the development of cannabis genetic research. Here, large-scale development of expressed sequence tag simple sequence repeat (EST-SSR) markers was performed to obtain more informative genetic markers, and to assess genetic diversity in cannabis (Cannabis sativa L.). Based on the cannabis transcriptome, 4,577 SSRs were identified from 3,624 ESTs. From there, a total of 3,442 complementary primer pairs were designed as SSR markers. Among these markers, trinucleotide repeat motifs (50.99%) were the most abundant, followed by hexanucleotide (25.13%), dinucleotide (16.34%), tetranucloetide (3.8%), and pentanucleotide (3.74%) repeat motifs, respectively. The AAG/CTT trinucleotide repeat (17.96%) was the most abundant motif detected in the SSRs. One hundred and seventeen EST-SSR markers were randomly selected to evaluate primer quality in 24 cannabis varieties. Among these 117 markers, 108 (92.31%) were successfully amplified and 87 (74.36%) were polymorphic. Forty-five polymorphic primer pairs were selected to evaluate genetic diversity and relatedness among the 115 cannabis genotypes. The results showed that 115 varieties could be divided into 4 groups primarily based on geography: Northern China, Europe, Central China, and Southern China. Moreover, the coefficient of similarity when comparing cannabis from Northern China with the European group cannabis was higher than that when comparing with cannabis from the other two groups, owing to a similar climate. This study outlines the first large-scale development of SSR markers for cannabis. These data may serve as a foundation for the development of genetic linkage, quantitative trait loci mapping, and marker-assisted breeding of cannabis. PMID:25329551

  11. Procoagulant activity may be a marker of the malignant phenotype in experimental prostate cancer.

    PubMed Central

    Adamson, A. S.; Luckert, P.; Pollard, M.; Snell, M. E.; Amirkhosravi, M.; Francis, J. L.

    1994-01-01

    Using a one-stage kinetic chromogenic assay, we studied the procoagulant activity (PCA) of prostatic tissue in an experimental model of prostate cancer in the rat. PCA was present in homogenates of rat prostate glands containing either benign or malignant tumours. The procoagulant activated factor X directly and was provisionally characterised as a tissue factor-factor VIIa complex. There was no significant differences in PCA between control rats and rats exposed to carcinogens that did not develop tumour. Levels in rats that developed tumours were significantly higher (P < 0.01) than all other groups and there was a positive correlation between tumour weight and PCA (r = 0.85, P < 0.001). Furthermore, prostatic PCA levels were higher in the metastasis (P < 0.02). We conclude that PCA reflects the malignant phenotype in this animals, the PCA of the primary tumour was compared with that of the corresponding secondary deposit and levels were higher in the metastasis (P < 0.02). We conclude that PCA reflects the malignant phenotype in this model of experimental prostate cancer and suggest that this parameter is worth evaluating as a potential tumour marker in the human disease. PMID:8297726

  12. Cancer procoagulant: a factor X activator, tumor marker and growth factor from malignant tissue.

    PubMed

    Gordon, S G; Mielicki, W P

    1997-03-01

    Hemostatic abnormalities associated with malignant disease led to the search for and discovery of a proteolytic enzyme that activated factor X in the blood coagulation cascade. It was named cancer procoagulant (CP). CP is a cysteine proteinase that is found in malignant and fetal (human amnion-chorion) tissue; it has not been found in normally differentiated tissue. It is a calcium-dependent, Mn2+ stimulated enzyme that has enhanced activity and inhibition in a reduced environment. This review presents a complete compilation and discussion of the known chemical and enzymatic characteristics of CP as well as many purification and assay procedures. Several unique properties of these procedures are described. Some problems and controversies are highlighted in each of the sections. An immunoassay for CP as a tumor marker and some of its potential applications in the diagnosis and monitoring of cancer are reviewed. Some therapeutic implications of CP are noted in light of the observation that antibodies to CP block the metastatic seeding of lung colonies in vivo and diminish the viability of tumor cells in vitro. Finally, comments about the relationship between tissue factor and CP in the malignant cells are provided.

  13. A population of human brain cells expressing phenotypic markers of more than one lineage can be induced in vitro to differentiate into mesenchymal cells

    SciTech Connect

    Rieske, Piotr; Augelli, Brian J.; Stawski, Robert; Gaughan, John; Azizi, S. Ausim; Krynska, Barbara

    2009-02-01

    Proliferating astrocytic cells from germinal, as well as mature areas of brain parenchyma, have the characteristics of neural stem/progenitor cells and are capable of generating both neurons and glia. We previously reported that primary fetal human brain cells, designated as Normal Human Astrocytes (NHA), expressed, in addition to GFAP, Vimentin and Nestin, low levels of {beta}III-Tubulin, an early neuronal marker, and differentiated into neurons and astrocytes in vitro. Here, we showed that primary NHA cells co-express low levels of mesenchymal markers Fibronectin and Collagen-1 in culture. These cells transitioned into mesenchymal-like cells when cultured in adherent conditions in serum containing media. The mesenchymal-like derivatives of these cells were characterized based on their morphological changes, high expression of Vimentin and extracellular matrix (ECM) proteins, Collagen-1 and Fibronectin, and decline of neural markers. When incubated in osteogenic and adipogenic induction media, the mesenchymal-like cells differentiated into osteoblasts and adipocytes. Furthermore, NHA cells express markers of neural crest cells, SOX-10 and p75. These data support the idea of ectoderm-derived mesenchymal lineages. These findings suggest that a population of primitive fetal brain cells with neural/neural crest/mesenchymal phenotype, resembles the remarkable phenotypic plasticity of neural crest cells, and differentiates into adipocytes and osteocytes under the influence of environmental factors.

  14. Genome-wide characterization and selection of expressed sequence tag simple sequence repeat primers for optimized marker distribution and reliability in peach

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Expressed sequence tag (EST) simple sequence repeats (SSRs) in Prunus were mined, and flanking primers designed and used for genome-wide characterization and selection of primers to optimize marker distribution and reliability. A total of 12,618 contigs were assembled from 84,727 ESTs, along with 34...

  15. Modulating Effect of Enicostemma littorale on the Expression Pattern of Apoptotic, Cell Proliferative, Inflammatory and Angiogenic Markers During 7, 12-Dimethylbenz (a) Anthracene Induced Hamster Buccal Pouch Carcinogenesis.

    PubMed

    Manoharan, Shanmugam; Rajasekaran, Duraisamy; Prabhakar, Murugaraj Manoj; Karthikeyan, Sekar; Manimaran, Asokan

    2015-01-01

    Enicostemma littorale leaves are traditionally used for the treatment of several diseases, including inflammation and cancer. This study has taken effort to explore the antitumor initiating potential of E. littorale leaves (ElELet) by analyzing the expression pattern of apoptotic (p53, Bcl-2 and Bcl-2 associated X-protein), cell-proliferative (cyclin D1 and proliferating cell nuclear antigen), angiogenic (vascular endothelial growth factor), invasive (matrix metalloproteinase-2 and 9), and inflammatory (NF-κB and cyclooxygenase-2) markers during 7, 12-dimethylbenz (a) anthracene (DMBA) induced hamster buccal pouch carcinogenesis. Oral tumors were induced in the buccal pouches of hamsters using the potent site and organ specific carcinogen, DMBA. DMBA application 3 times a week for 14 weeks resulted in tumor formation in the buccal pouches. Hundred percent tumor formations with dysregulation in the expression pattern of apoptotic, cell proliferative, inflammatory, angiogenic, and invasive markers were observed in the buccal pouches of hamsters treated with DMBA alone. ElELet at a dose of 250 mg/kg body weight orally to DMBA treated hamsters significantly prevented the tumor formation as well as corrected the abnormalities in the expression pattern of above mentioned molecular markers. ElELet thus modulated the expression pattern of all the above mentioned molecular markers in favor of the suppression of cell proliferation occurring in DMBA induced hamster buccal pouch carcinogenesis. PMID:26862274

  16. Modulating Effect of Enicostemma littorale on the Expression Pattern of Apoptotic, Cell Proliferative, Inflammatory and Angiogenic Markers During 7, 12-Dimethylbenz (a) Anthracene Induced Hamster Buccal Pouch Carcinogenesis

    PubMed Central

    Manoharan, Shanmugam; Rajasekaran, Duraisamy; Prabhakar, Murugaraj Manoj; Karthikeyan, Sekar; Manimaran, Asokan

    2015-01-01

    Enicostemma littorale leaves are traditionally used for the treatment of several diseases, including inflammation and cancer. This study has taken effort to explore the antitumor initiating potential of E. littorale leaves (ElELet) by analyzing the expression pattern of apoptotic (p53, Bcl-2 and Bcl-2 associated X-protein), cell-proliferative (cyclin D1 and proliferating cell nuclear antigen), angiogenic (vascular endothelial growth factor), invasive (matrix metalloproteinase-2 and 9), and inflammatory (NF-κB and cyclooxygenase-2) markers during 7, 12-dimethylbenz (a) anthracene (DMBA) induced hamster buccal pouch carcinogenesis. Oral tumors were induced in the buccal pouches of hamsters using the potent site and organ specific carcinogen, DMBA. DMBA application 3 times a week for 14 weeks resulted in tumor formation in the buccal pouches. Hundred percent tumor formations with dysregulation in the expression pattern of apoptotic, cell proliferative, inflammatory, angiogenic, and invasive markers were observed in the buccal pouches of hamsters treated with DMBA alone. ElELet at a dose of 250 mg/kg body weight orally to DMBA treated hamsters significantly prevented the tumor formation as well as corrected the abnormalities in the expression pattern of above mentioned molecular markers. ElELet thus modulated the expression pattern of all the above mentioned molecular markers in favor of the suppression of cell proliferation occurring in DMBA induced hamster buccal pouch carcinogenesis. PMID:26862274

  17. BACE1 activity in cerebrospinal fluid and its relation to markers of AD pathology.

    PubMed

    Mulder, Sandra D; van der Flier, Wiesje M; Verheijen, Jan H; Mulder, Cees; Scheltens, Philip; Blankenstein, Marinus A; Hack, C Erik; Veerhuis, Robert

    2010-01-01

    Several studies have shown that reduced amyloid-beta 1-42 (Abeta(42)) and increased tau levels in cerebrospinal fluid (CSF) reflect increased Alzheimer's disease (AD) pathology in the brain. beta-site APP cleaving enzyme (BACE1) is thought to be the major beta-secretase involved in Abeta production in the brain, and therefore we investigated the relation between BACE1 activity and CSF markers Abeta(40), Abeta(42), total tau (t-tau), and tau phosphorylated at threonine 181 (p-tau) in CSF of control (n=12), mild cognitive impairment (n=18), and AD (n=17) subjects. Patients were classified according to their Abeta(42), t-tau, and p-tau CSF biomarker levels, with either an AD-like biomarker profile (two or three biomarkers abnormal: Abeta(42) < 495 pg/ml in combination with t-tau > 356 pg/ml, and/or p-tau > 54 pg/ml) or a normal biomarker profile (activity levels, compared to patients with a normal biomarker profile (20 pg/ml and 16 pg/ml respectively; p=0.01), when controlled for age and gender. In the whole sample, BACE1 activity correlated with CSF levels of Abeta(40), t-tau, and p-tau (r=0.38, r=0.63, and r=0.65; all p< 0.05), but not with Abeta(42). These data suggest that increased BACE1 activity in CSF relates to AD pathology in the brain.

  18. BACE1 activity in cerebrospinal fluid and its relation to markers of AD pathology.

    PubMed

    Mulder, Sandra D; van der Flier, Wiesje M; Verheijen, Jan H; Mulder, Cees; Scheltens, Philip; Blankenstein, Marinus A; Hack, C Erik; Veerhuis, Robert

    2010-01-01

    Several studies have shown that reduced amyloid-beta 1-42 (Abeta(42)) and increased tau levels in cerebrospinal fluid (CSF) reflect increased Alzheimer's disease (AD) pathology in the brain. beta-site APP cleaving enzyme (BACE1) is thought to be the major beta-secretase involved in Abeta production in the brain, and therefore we investigated the relation between BACE1 activity and CSF markers Abeta(40), Abeta(42), total tau (t-tau), and tau phosphorylated at threonine 181 (p-tau) in CSF of control (n=12), mild cognitive impairment (n=18), and AD (n=17) subjects. Patients were classified according to their Abeta(42), t-tau, and p-tau CSF biomarker levels, with either an AD-like biomarker profile (two or three biomarkers abnormal: Abeta(42) < 495 pg/ml in combination with t-tau > 356 pg/ml, and/or p-tau > 54 pg/ml) or a normal biomarker profile (activity levels, compared to patients with a normal biomarker profile (20 pg/ml and 16 pg/ml respectively; p=0.01), when controlled for age and gender. In the whole sample, BACE1 activity correlated with CSF levels of Abeta(40), t-tau, and p-tau (r=0.38, r=0.63, and r=0.65; all p< 0.05), but not with Abeta(42). These data suggest that increased BACE1 activity in CSF relates to AD pathology in the brain. PMID:20164582

  19. Tissue factor activity. A marker of alveolar macrophage maturation in rabbits. Effects of granulomatous pneumonitis.

    PubMed Central

    Rothberger, H; McGee, M P; Lee, T K

    1984-01-01

    amounts of tissue factor activity due to the presence of large numbers of mature alveolar macrophage forms that had high levels of the procoagulant. Thus, tissue factor activity in alveolar macrophages is a marker of cellular maturation in vivo and in vitro. Increased amounts of this initiator of the extrinsic clotting pathway, as found in alveolar macrophage populations from animals with granulomatous pneumonitis induced by BCG hypersensitivity, suggest that alveolar macrophage tissue factor may contribute to the pathology of immune lung diseases. PMID:6373826

  20. Matriptase Expression and Zymogen Activation in Human Pilosebaceous Unit

    PubMed Central

    Wu, Bai-Yao; Lee, Shiao-Pieng; Hsiao, Hui-Chung; Chiu, Han; Chen, Chi-Yung; Yeo, Yee Hui; Lee, Herng-Sheng; Chen, Ya-Wen; Kaul, Malvika; Kataoka, Hiroaki; Johnson, Michael D.; Lin, Chen-Yong

    2014-01-01

    Studies of human genetic disorders and mouse models reveal the important roles of matriptase in hair growth. Here, we investigate matriptase expression and zymogen activation in hair follicles. We show: 1) layer-dependent distribution patterns, with much higher matriptase expression in cells of the outer root sheath and matrix cells of the hair bulb than in cells of the inner root sheath; 2) cycle-dependent expression patterns, with matriptase expressed in the anagen and catagen phases of the hair lifecycle, but not in the telogen phase; 3) reduced expression of the matriptase inhibitor, HAI-1, in the catagen phase, suggesting increased proteolytic activity in this phase; and 4) definitive matriptase zymogen activation patterns, with the highest matriptase activation observed in matrix cells and outer root sheath cells in the isthmus/bulge region. In sebaceous glands, matriptase is highly expressed in basal and ductal cells, with much lower expression in the differentiated, lipid-filled cells of the interior. We also show that matriptase potently activates hepatocyte growth factor (HGF) in vitro, and that the HGF receptor, c-Met, is co-expressed in those cells that express activated matriptase. Our observations suggest that the matriptase-HGF-c-MET pathway has the potential to be engaged, primarily in proliferative cells rather than terminally differentiated epithelial cells of the human pilosebaceous unit. PMID:24004857

  1. GPR30 Promotes Prostate Stromal Cell Activation via Suppression of ERα Expression and Its Downstream Signaling Pathway.

    PubMed

    Jia, Bona; Gao, Yu; Li, Mingming; Shi, Jiandang; Peng, Yanfei; Du, Xiaoling; Klocker, Helmut; Sampson, Natalie; Shen, Yongmei; Liu, Mengyang; Zhang, Ju

    2016-08-01

    Cancer-associated fibroblasts (CAFs) play a vital role in malignant transformation and progression of prostate cancer (PCa), and accumulating evidence suggests an enhancing effect of estrogens on PCa. The present study aimed to investigate the possible origin of prostate CAFs and the effects of estrogen receptors, G protein-coupled receptor 30 (GPR30) and estrogen receptor (ER)-α, on stromal cell activation. High expression of fibroblast activation protein (FAP), CD44, and nonmuscle myosin heavy chain B (SMemb) accompanied by low expression of smooth muscle differentiation markers was found in the stromal cells of PCa tissues and in cultured human prostate CAFs. Additionally, SMemb expression, which is coupled to cell phenotype switching and proliferation, was coexpressed with FAP, a marker of activated stromal cells, and with the stem cell marker CD44 in the stromal cells of PCa tissue. Prostate CAFs showed high GPR30 and low ERα expression. Moreover, GPR30 was coexpressed with FAP, CD44, and SMemb. Furthermore, the study demonstrated that the overexpression of GPR30 or the knockdown of ERα in prostate stromal cells induced the up-regulation of FAP, CD44, Smemb, and the down-regulation of smooth muscle markers. The conditioned medium from these cells promoted the proliferation and migration of LNCaP and PC3 PCa cells. GPR30 knockdown or ERα overexpression showed opposite effects. Finally, we present a novel mechanism whereby GPR30 limits ERα expression via inhibition of the cAMP/protein kinase A signaling pathway. These results suggest that stem-like cells within the stroma are a possible source of prostate CAFs and that the negative regulation of ERα expression by GPR30 is centrally involved in prostate stromal cell activation. PMID:27163843

  2. Production of marker-free transgenic Jatropha curcas expressing hybrid Bacillus thuringiensis δ-endotoxin Cry1Ab/1Ac for resistance to larvae of tortrix moth (Archips micaceanus)

    PubMed Central

    2014-01-01

    Background The potential biofuel plant Jatropha curcas L. is affected by larvae of Archips micaceanus (Walker), a moth of the family Tortricidae. The hybrid Bacillus thuringiensis (Bt) δ-endotoxin protein Cry1Ab/1Ac confers resistance to lepidopteran insects in transgenic rice. Results Here, we report the production of a marker-free transgenic line of J. curcas (L10) expressing Cry1Ab/1Ac using Agrobacterium-mediated transformation and a chemically regulated, Cre/loxP-mediated DNA recombination system. L10 carries a single copy of marker-free T-DNA that contains the Cry1Ab/1Ac gene under the control of a maize phosphoenolpyruvate carboxylase gene promoter (P Pepc :Cry1Ab/1Ac:T Nos ). The P Pepc :Cry1Ab/1Ac:T Nos gene was highly expressed in leaves of L10 plants. Insecticidal bioassays using leaf explants of L10 resulted in 80-100% mortality of larvae of A. micaceanus at 4 days after infestation. Conclusion The results demonstrate that the hybrid Bt δ-endotoxin protein Cry1Ab/1Ac expressed in Jatropha curcas displays strong insecticidal activity to A. micaceanus. The marker-free transgenic J. curcas line L10 can be used for breeding of insect resistance to A. micaceanus. PMID:24808924

  3. HPV16 E2 is an immediate early marker of viral infection, preceding E7 expression in precursor structures of cervical carcinoma.

    PubMed

    Xue, Yuezhen; Bellanger, Sophie; Zhang, Wenying; Lim, Diana; Low, Jeffrey; Lunny, Declan; Thierry, Françoise

    2010-07-01

    The viral E2 gene product plays a crucial role in the human papillomavirus (HPV) vegetative cycle by regulating both transcription and replication of the viral genome. E2 is a transcriptional repressor of the E6 and E7 viral oncogenes for HPV types 16 and 18, which are involved in cervical cancers. Using new polyclonal antibodies against the HPV16 E2 protein, we showed that E2 is expressed at various precursor stages of cervical carcinoma by immunohistochemistry on paraffin-embedded clinical samples. E2 was found to be highly expressed in the nuclei and cytoplasm of cells forming the intermediate and upper layers of cervical intraepithelial neoplasia (CIN). We could show that the expressions of E2 and p16(INK4a) (surrogate marker for oncogenic E7 expression) were exclusive in most of the cases, thus implying that E2 is not expressed together with high levels of E7. Moreover, we found that E2 is expressed in a subset of columnar cells adjacent to the CIN. We could show that expression of E2 is topologically distinct from the proliferation markers p63 and Ki67, whereas it coincides with the expression of cytokeratin K13, a marker of squamous cell differentiation. Expression of E2 also topologically coincides with episomal amplification of viral genomes in the upper layers of CIN1. These in vivo data thus validate previous assumptions of the crucial role of E2 in the early steps of HPV infection and of its negative link with expression of the viral E6 and E7 oncogenes.

  4. Influence of inositol hexaphosphate on the expression of selected proliferation markers in IL-1β-stimulated intestinal epithelial cells.

    PubMed

    Kapral, Małgorzata; Sośnicki, Stanisław; Wawszczyk, Joanna; Węglarz, Ludmiła

    2014-01-01

    The aim of the present study was to examine the influence of IP6, a naturally occurring phytochem- ical, on the expression of genes coding for proliferation markers, i.e., cyclin D1 (CCND1) and histone H3 in IL-1β-stimulated intestinal cancer cell line Caco-2. Quantification of genes expression was carried out using real time RT-QPCR technique in Caco-2 cells after treatment with IL-1β, 1 and 2.5 mM of IP6 for 3, 6 and 12 h. In separate cultures, cells were incubated with IL-1β for the indicated times. The untreated Caco-2 cells were used as the control. In a time course experiment, stimulation of cells with IL-1β only resulted in an overex- pression of both CCND1 and histone H3 mRNAs as compared with control. IP6 had no influence on IL-1β-stimulated CCND1 expression for 3 and 6 h. After 12 h, statistically significant decrease in CCND1 mRNA was observed in cells exposed to IL-1β and IP6 (1 and 2.5 mM) in relation to cells treated with IL-1β only. The levels of H3 mRNA in IL-1β-stimulated cells and cells treated with IL-1β and IP6 revealed no statistically significant differences after 3 h. IP6 at 1 and 2.5 mM enhanced IL1β-stimulated transcription of H3 gene after 6 h. Subsequently (12 h), the combination of IP6 and IL-1β decreased H3 mRNA level compared to IL1β-treated cells. In conclusion, pro-inflammatory cytokine IL-1β up-regulates CCND1 and histone H3 mRNAs expression in Caco-2 cells. These results suggest that the ability of IP6 to inhibit colon cancer cells proliferation may be mediated through downregulation of genes encoding cyclin D1 and histone H3 at the mRNA level. PMID:25745771

  5. Glucocorticoids Alter CRTC-CREB Signaling in Muscle Cells: Impact on PGC-1α Expression and Atrophy Markers

    PubMed Central

    Rahnert, Jill A.; Zheng, Bin; Hudson, Matthew B.; Woodworth-Hobbs, Myra E.; Price, S. Russ

    2016-01-01

    Muscle wasting associated with chronic diseases has been linked to decreased expression of PGC-1α and overexpression of PGC-1α counters muscle loss. CREB, in conjunction with the CREB-regulated transcription coactivator (CRTC2), is a positive modulator of PGC-1α transcription. We previously reported that PGC-1α expression is decreased in skeletal muscle of diabetic rats despite a high level of CREB phosphorylation (i.e., activation), suggesting that CRTC2-CREB signaling may be dysregulated. In this study, the relationship between CREB/CRTC signaling and PGC-1α expression was examined in L6 myotubes treated with dexamethasone (Dex, 48h) to induce atrophy. Dex decreased PGC-1α mRNA and protein as well as the levels of CRTC1 and CRTC2 in the nucleus. Dex also altered the nuclear levels of two known regulators of CRTC2 localization; the amount of calcinuerin catalytic A subunit (CnA) was decreased whereas SIK was increased. To assess PGC-1α transcription, muscle cells were transfected with a PGC-1α luciferase reporter plasmid (PGC-1α-Luc). Dex suppressed PGC-1α luciferase activity while both isobutylmethylxanthine (IBMX) and over-expression of CRTC1 or CRTC2 increased PGC-1α-Luc activity. Mutation of the CRE binding site from PGC-1α-Luc reporter attenuated the responses to both IBMX and the CRTC proteins. Consistent with the reporter gene results, overexpression of CRTC2 produced an increase in CRTC2 in the nucleus and in PGC-1α mRNA and PGC-1α protein. Overexpression of CRTC2 was not sufficient to prevent the decrease in PGC-1α mRNA or protein by Dex. In summary, these data suggest that attenuated CREB/CRTC signaling contributes to the decrease in PGC-1α expression during atrophy. PMID:27404111

  6. Glucocorticoids Alter CRTC-CREB Signaling in Muscle Cells: Impact on PGC-1α Expression and Atrophy Markers.

    PubMed

    Rahnert, Jill A; Zheng, Bin; Hudson, Matthew B; Woodworth-Hobbs, Myra E; Price, S Russ

    2016-01-01

    Muscle wasting associated with chronic diseases has been linked to decreased expression of PGC-1α and overexpression of PGC-1α counters muscle loss. CREB, in conjunction with the CREB-regulated transcription coactivator (CRTC2), is a positive modulator of PGC-1α transcription. We previously reported that PGC-1α expression is decreased in skeletal muscle of diabetic rats despite a high level of CREB phosphorylation (i.e., activation), suggesting that CRTC2-CREB signaling may be dysregulated. In this study, the relationship between CREB/CRTC signaling and PGC-1α expression was examined in L6 myotubes treated with dexamethasone (Dex, 48h) to induce atrophy. Dex decreased PGC-1α mRNA and protein as well as the levels of CRTC1 and CRTC2 in the nucleus. Dex also altered the nuclear levels of two known regulators of CRTC2 localization; the amount of calcinuerin catalytic A subunit (CnA) was decreased whereas SIK was increased. To assess PGC-1α transcription, muscle cells were transfected with a PGC-1α luciferase reporter plasmid (PGC-1α-Luc). Dex suppressed PGC-1α luciferase activity while both isobutylmethylxanthine (IBMX) and over-expression of CRTC1 or CRTC2 increased PGC-1α-Luc activity. Mutation of the CRE binding site from PGC-1α-Luc reporter attenuated the responses to both IBMX and the CRTC proteins. Consistent with the reporter gene results, overexpression of CRTC2 produced an increase in CRTC2 in the nucleus and in PGC-1α mRNA and PGC-1α protein. Overexpression of CRTC2 was not sufficient to prevent the decrease in PGC-1α mRNA or protein by Dex. In summary, these data suggest that attenuated CREB/CRTC signaling contributes to the decrease in PGC-1α expression during atrophy.

  7. Decrease in drug accumulation and in tumour aggressiveness marker expression in a fenretinide-induced resistant ovarianumour cell line

    PubMed Central

    Appierto, V; Cavadini, E; Pergolizzi, R; Cleris, L; Lotan, R; Canevari, S; Formelli, F

    2001-01-01

    We investigated whether the efficacy of fenretinide (HPR) against ovarian tumours may be limited by induction of resistance. The human ovarian carcinoma cell line A2780, which is sensitive to a pharmacologically achievable HPR concentration (IC 50= 1 μM), became 10-fold more resistant after exposure to increasing HPR concentrations. The cells (A2780/HPR) did not show cross-resistance to the synthetic retinoid 6-[3-adamantyl-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) and were not sensitive, similarly to the parent line, to all- trans -retinoic acid, 13- cis -retinoic acid or N-(4-methoxyphenyl)retinamide. A2780/HPR cells showed, compared to parental cells, a 3-fold reduction in colony-forming ability in agar. The development of HPR resistance was associated with a marked increase in retinoic acid receptor β (RARβ) mRNA and protein levels, which decreased, together with drug resistance, after drug removal. The expression of cell surface molecules associated with tumour progression including HER-2, laminin receptor and β1 integrin was markedly reduced. The increase in the levels of reactive oxygen species is not involved in HPR-resistance because it was similar in parental and resistant cells. Conversely differences in pharmacokinetics may account for resistance because, in A2780/HPR cells, intracellular peak drug levels were 2 times lower than in A2780 cells and an as yet unidentified polar metabolite was present. These data suggest that acquired resistance to HPR is associated with changes in marker expression, suggestive of a more differentiated status and may be explained, at least in part, by reduced drug accumulation and increased metabolism. © 2001 Cancer Research Campaign http://www.bjcancer.com PMID:11384104

  8. Expression of mesenchymal stem cell marker CD90 on dermal sheath cells of the anagen hair follicle in canine species

    PubMed Central

    Mercati, F.; Pascucci, L.; Ceccarelli, P.; Dall’Aglio, C.; Pedini, V.; Gargiulo, A.M.

    2009-01-01

    The dermal sheath (DS) of the hair follicle is comprised by fibroblast-like cells and extends along the follicular epithelium, from the bulb up to the infundibulum. From this structure, cells with stem characteristics were isolated: they have a mesenchymal origin and express CD90 protein, a typical marker of mesenchymal stem cells. It is not yet really clear in which region of hair follicle these cells are located but some experimental evidence suggests that dermal stem cells are localized prevalently in the lower part of the anagen hair follicle. As there are no data available regarding DS stem cells in dog species, we carried out a morphological analysis of the hair follicle DS and performed both an immunohistochemical and an immunocytochemical investigation to identify CD90+ cells. We immunohistochemically evidenced a clear and abundant positivity to CD90 protein in the DS cells located in the lower part of anagen hair follicle. The positive cells showed a typical fibroblast-like morphology. They were flat and elongated and inserted among bundles of collagen fibres.The whole structure formed a close and continuous sleeve around the anagen hair follicle. Our immunocytochemical study allowed us to localize CD90 protein at the cytoplasmic membrane level.

  9. Development and characterization of novel expressed sequence tag-derived simple sequence repeat markers in Hevea brasiliensis (rubber tree).

    PubMed

    An, Z W; Li, Y C; Zhai, Q L; Xie, L L; Zhao, Y H; Huang, H S

    2013-11-22

    Cultivated clones of Hevea brasiliensis have a narrow genetic base. In order to broaden the genetic base, it is first necessary to investigate the genetic diversity of wild populations. Expressed sequence tag-simple sequence repeat (EST-SSR) markers were developed to investigate the genetic diversity of Hevea populations. Four hundred and thirty microsatellites were identified and 148 primers were designed to amplify the loci. Twenty-nine primer pairs were synthesized and evaluated for their ability to detect genetic polymorphisms among 40 wild accessions of H. brasiliensis. Twenty-one of the 29 loci were polymorphic. The number of alleles per locus in the 40 accessions ranged from 2 to 7. H(O) and H(E) at each locus ranged from 0.0000 to 0.9000 and from 0.0000 to 0.8704, respectively. All 21 loci could amplify in H. brasiliensis, H. pauciflora, H. nitida, H. spruceana, and H. camargoana. The EST-SSR primers developed herein can be used in genetic diversity and structure studies in H. brasiliensis.

  10. Identification and characterization of 43 microsatellite markers derived from expressed sequence tags of the sea cucumber ( Apostichopus japonicus)

    NASA Astrophysics Data System (ADS)

    Jiang, Qun; Li, Qi; Yu, Hong; Kong, Lingfeng

    2011-06-01

    The sea cucumber Apostichopus japonicus is a commercially and ecologically important species in China. A total of 3056 potential unigenes were generated after assembling 7597 A. japonicus expressed sequence tags (ESTs) downloaded from Gen-Bank. Two hundred and fifty microsatellite-containing ESTs (8.18%) and 299 simple sequence repeats (SSRs) were detected. The average density of SSRs was 1 per 7.403 kb of EST after redundancy elimination. Di-nucleotide repeat motifs appeared to be the most abundant type with a percentage of 69.90%. Of the 126 primer pairs designed, 90 amplified the expected products and 43 showed polymorphism in 30 individuals tested. The number of alleles per locus ranged from 2 to 26 with an average of 7.0 alleles, and the observed and expected heterozygosities varied from 0.067 to 1.000 and from 0.066 to 0.959, respectively. These new EST-derived microsatellite markers would provide sufficient polymorphism for population genetic studies and genome mapping of this sea cucumber species.

  11. Evaluation of periodontitis treatment effects on carotid intima-media thickness and expression of laboratory markers related to atherosclerosis.

    PubMed

    Toregeani, Jeferson Freitas; Nassar, Carlos Augusto; Nassar, Patrícia Oehlmeyer; Toregeani, Krischina Mendes; Gonzatto, Geyssi Karolyne; Vendrame, Rafael; Castilhos, Jussimar Scheffer; Rotta, Larissa Sokol; Reinheimer, Andréia Carpenedo; Longoni, Anelise; Barcella, Mariana Waterkemper Andrade

    2016-01-01

    The objective of this research was to evaluate the treatment of periodontal disease and its effects on carotid intima-media thickness (CIMT) and expression of laboratory markers related to atherosclerosis. Twenty-three healthy patients (group 1) and 21 patients with moderate to severe periodontitis (group 2) underwent evaluation of clinical periodontal parameters. The patients were submitted to CIMT measurements and laboratory evaluations at the start of the study (0 months), 6 months, and 12 months. All patients received oral hygiene instruction; patients in group 2 also underwent supragingival and subgingival scaling and root planing. A statistically significant improvement in clinical periodontal parameters occurred in both groups (P < 0.05). Improvements were more evident between the first and second evaluations and were greater in group 2. Both groups experienced a statistically significant decrease in CIMT in the first 6 months (P < 0.05). Treatments--oral hygiene instruction in group 1 and instruction plus mechanical periodontal instrumentation in group 2--were effective in improving clinical periodontal parameters of both groups and promoting reduction in CIMT at 6 months. PMID:26742169

  12. Blue light-induced inflammatory marker expression in the retinal pigment epithelium-choroid of mice and the protective effect of a yellow intraocular lens material in vivo.

    PubMed

    Narimatsu, Toshio; Negishi, Kazuno; Miyake, Seiji; Hirasawa, Manabu; Osada, Hideto; Kurihara, Toshihide; Tsubota, Kazuo; Ozawa, Yoko

    2015-03-01

    Oxidative stress in the retinal pigment epithelium (RPE) is a well-accepted pathogenic change in vision-threatening diseases such as age-related macular degeneration. One source of oxidative stress is excessive light exposure, which causes excessive activation of the visual cycle. Because short wavelength light (blue light) has more energy, it is reported to be more harmful to photoreceptor cells than the other wavelengths of light. However, the biological effect of blue light in the RPE of living animals and the protective effect of a yellow intraocular lens (IOL) material that blocks blue light is still obscure. Therefore, we compared the pathogenic effect in the RPE-choroid complexes of mice exposed to light in a box made of a clear or a yellow IOL material. We measured the level of reactive oxygen species (ROS) using 2', 7'-dichlorodihydrofluorescein diacetate, the mRNA levels of inflammatory cytokines and a macrophage marker by real-time polymerase chain reaction, and the protein level of monocyte chemotactic protein-1 (MCP-1) by ELISA. The ROS level after light exposure was suppressed in the RPE-choroids of light-exposed mice in the yellow IOL material box. In parallel, all the inflammatory cytokines that we measured and a macrophage marker were also suppressed in the RPE-choroids of light-exposed mice in the yellow IOL material box. Therefore, a yellow IOL material suppressed, and thus blue light exacerbated, the increase in the ROS level and inflammatory cytokine expression as well as macrophage recruitment in the RPE-choroid in vivo after light exposure.

  13. Quantitative real-time RT-PCR assessment of spinal microglial and astrocytic activation markers in a rat model of neuropathic pain.

    PubMed

    Tanga, F Y; Raghavendra, V; DeLeo, J A

    2004-01-01

    Activated spinal glial cells have been strongly implicated in the development and maintenance of persistent pain states following a variety of stimuli including traumatic nerve injury. The present study was conducted to characterize the time course of surface markers indicative of microglial and astrocytic activation at the transcriptional level following an L5 nerve transection that results in behavioral hypersensitivity. Male Sprague-Dawley rats were divided into a normal group, a sham surgery group with an L5 spinal nerve exposure and an L5 spinal nerve transected group. Mechanical allodynia (heightened response to a non-noxious stimulus) of the ipsilateral hind paw was assessed throughout the study. Spinal lumbar mRNA levels of glial fibrillary acidic protein (GFAP), integrin alpha M (ITGAM), toll-like receptor 4 (TLR4) and cluster determinant 14 (CD14) were assayed using real-time reverse transcription polymerase chain reaction (RT-PCR) at 4 h, 1, 4, 7, 14 and 28 days post surgery. The spinal lumbar mRNA expression of ITGAM, TLR4, and CD14 was upregulated at 4 h post surgery, CD14 peaked 4 days after spinal nerve transection while ITGAM and TLR4 continued to increase until day 14 and returned to almost normal levels by postoperative day 28. In contrast, spinal GFAP mRNA did not significantly increase until postoperative day 4 and then continued to increase over the duration of the study. Our optimized real-time RT-PCR method was highly sensitive, specific and reproducible at a wide dynamic range. This study demonstrates that peripheral nerve injury induces an early spinal microglial activation that precedes astrocytic activation using mRNA for surface marker expression; the delayed but sustained expression of mRNA coding for GFAP implicates astrocytes in the maintenance phase of persistent pain states. In summary, these data demonstrate a distinct spinal glial response following nerve injury using real-time RT-PCR. PMID:15145554

  14. Identification of a β-galactosidase transgene that provides a live-cell marker of transcriptional activity in growing oocytes and embryos.

    PubMed

    Edwards, Nicole; Farookhi, Riaz; Clarke, Hugh J

    2015-07-01

    Identifying the events and molecular mechanisms that regulate oocyte growth has emerged as a key objective of research in human fertility, fuelled by evidence from human and animal studies indicating that disease and environmental factors can act on oocytes to affect the health of the resulting individual and by efforts to grow oocytes in vitro to enable fertility preservation of cancer survivors. Techniques that monitor the development of growing oocytes would be valuable tools to assess the progression of growth under different conditions. Most methods used to assess oocytes grown in vitro are indirect, however, relying on characteristics of the somatic compartment of the follicle, or compromise the oocyte, preventing its subsequent culture or fertilization. We investigated the utility of T-cell factor/lymphoid enhancer-binding factor (TCF/Lef)-LacZ transgene expression as a predictor of global transcriptional activity in oocytes and early embryos. Using a fluorescent β-galactosidase substrate combined with live-cell imaging, we show that TCF/Lef-LacZ transgene expression is detectable in growing oocytes, lost in fully grown oocytes and resumes in late two-cell embryos. Transgene expression is likely regulated by a Wnt-independent mechanism. Using chromatin analysis, LacZ expression and methods to monitor and inhibit transcription, we show that TCF/Lef-LacZ expression mirrors transcriptional activity in oocytes and preimplantation embryos. Oocytes and preimplantation embryos that undergo live-cell imaging for TCF/Lef-LacZ expression are able to continue development in vitro. TCF/Lef-LacZ reporter expression in living oocytes and early embryos is thus a sensitive and faithful marker of transcriptional activity that can be used to monitor and optimize conditions for oocyte growth.

  15. Tyrosine Hydroxylase Phosphorylation in Catecholaminergic Brain Regions: A Marker of Activation following Acute Hypotension and Glucoprivation

    PubMed Central

    Damanhuri, Hanafi A.; Burke, Peter G. R.; Ong, Lin K.; Bobrovskaya, Larisa; Dickson, Phillip W.; Dunkley, Peter R.; Goodchild, Ann K.

    2012-01-01

    The expression of c-Fos defines brain regions activated by the stressors hypotension and glucoprivation however, whether this identifies all brain sites involved is unknown. Furthermore, the neurochemicals that delineate these regions, or are utilized in them when responding to these stressors remain undefined. Conscious rats were subjected to hypotension, glucoprivation or vehicle for 30, 60 or 120 min and changes in the phosphorylation of serine residues 19, 31 and 40 in the biosynthetic enzyme, tyrosine hydroxylase (TH), the activity of TH and/or, the expression of c-Fos were determined, in up to ten brain regions simultaneously that contain catecholaminergic cell bodies and/or terminals: A1, A2, caudal C1, rostral C1, A6, A8/9, A10, nucleus accumbens, dorsal striatum and medial prefrontal cortex. Glucoprivation evoked phosphorylation changes in A1, caudal C1, rostral C1 and nucleus accumbens whereas hypotension evoked changes A1, caudal C1, rostral C1, A6, A8/9, A10 and medial prefrontal cortex 30 min post stimulus whereas few changes were evident at 60 min. Although increases in pSer19, indicative of depolarization, were seen in sites where c-Fos was evoked, phosphorylation changes were a sensitive measure of activation in A8/9 and A10 regions that did not express c-Fos and in the prefrontal cortex that contains only catecholaminergic terminals. Specific patterns of serine residue phosphorylation were detected, dependent upon the stimulus and brain region, suggesting activation of distinct signaling cascades. Hypotension evoked a reduction in phosphorylation in A1 suggestive of reduced kinase activity. TH activity was increased, indicating synthesis of TH, in regions where pSer31 alone was increased (prefrontal cortex) or in conjunction with pSer40 (caudal C1). Thus, changes in phosphorylation of serine residues in TH provide a highly sensitive measure of activity, cellular signaling and catecholamine utilization in catecholaminergic brain regions, in the

  16. Connexin 50 Expression in Ependymal Stem Progenitor Cells after Spinal Cord Injury Activation.

    PubMed

    Rodriguez-Jimenez, Francisco Javier; Alastrue-Agudo, Ana; Stojkovic, Miodrag; Erceg, Slaven; Moreno-Manzano, Victoria

    2015-01-01

    Ion channels included in the family of Connexins (Cx) help to control cell proliferation and differentiation of neuronal progenitors. Here we explored the role of Connexin 50 (Cx50) in cell fate modulation of adult spinal cord derived neural precursors located in the ependymal canal (epSPC). epSPC from non-injured animals showed high expression levels of Cx50 compared to epSPC from animals with spinal cord injury (SCI) (epSPCi). When epSPC or epSPCi were induced to spontaneously differentiate in vitro we found that Cx50 favors glial cell fate, since higher expression levels, endogenous or by over-expression of Cx50, augmented the expression of the astrocyte marker GFAP and impaired the neuronal marker Tuj1. Cx50 was found in both the cytoplasm and nucleus of glial cells, astrocytes and oligodendrocyte-derived cells. Similar expression patterns were found in primary cultures of mature astrocytes. In addition, opposite expression profile for nuclear Cx50 was observed when epSPC and activated epSPCi were conducted to differentiate into mature oligodendrocytes, suggesting a different role for this ion channel in spinal cord beyond cell-to-cell communication. In vivo detection of Cx50 by immunohistochemistry showed a defined location in gray matter in non-injured tissues and at the epicenter of the injury after SCI. epSPCi transplantation, which accelerates locomotion regeneration by a neuroprotective effect after acute SCI is associated with a lower signal of Cx50 within the injured area, suggesting a minor or detrimental contribution of this ion channel in spinal cord regeneration by activated epSPCi. PMID:26561800

  17. Plasminogen activator inhibitor-1 as an early potential diagnostic marker for Alzheimer's disease.

    PubMed

    Oh, Jaeho; Lee, Hye-Ja; Song, Ji-Hyun; Park, Sang Ick; Kim, Hyunyoung

    2014-12-01

    Alzheimer's disease (AD) is the most common cause of dementia in individuals over 65 years old. However, to date, no useful early diagnostic markers for AD have been discovered. We examined the utility of plasminogen activator inhibitor-1 (PAI-1) as a potential biomarker for AD in subjects with mild cognitive impairment (MCI) or AD, as well as in nondemented healthy controls. Plasma PAI-1 levels were measured by enzyme-linked immunosorbent assays (ELISAs) in samples collected from 76 patients with MCI, 74 patients with AD, and 76 healthy controls. Our results show that plasma PAI-1 levels gradually increased as dementia progressed. The mean levels of plasma PAI-1 in patients with MCI and AD patients were significantly higher than those of in healthy controls. Consistently, neuropsychological examination (e.g., MMSE, CDR) also demonstrated significant correlations between the plasma PAI-1 levels and cognitive function. In conclusion, the level of plasma PAI-1 is a potential biomarker for the early detection and diagnosis of AD.

  18. ADENOSINE DEAMINASE ACTIVITY AND SERUM C-REACTIVE PROTEIN AS PROGNOSTIC MARKERS OF CHAGAS DISEASE SEVERITY.

    PubMed

    Bravo-Tobar, Iván Darío; Nello-Pérez, Carlota; Fernández, Alí; Mogollón, Nora; Pérez, Mary Carmen; Verde, Juan; Concepción, Juan Luis; Rodriguez-Bonfante, Claudina; Bonfante-Cabarcas, Rafael

    2015-01-01

    Chagas disease is a public health problem worldwide. The availability of diagnostic tools to predict the development of chronic Chagas cardiomyopathy is crucial to reduce morbidity and mortality. Here we analyze the prognostic value of adenosine deaminase serum activity (ADA) and C-reactive protein serum levels (CRP) in chagasic individuals. One hundred and ten individuals, 28 healthy and 82 chagasic patients were divided according to disease severity in phase I (n = 35), II (n = 29), and III (n = 18). A complete medical history, 12-lead electrocardiogram, chest X-ray, and M-mode echocardiogram were performed on each individual. Diagnosis of Chagas disease was confirmed by ELISA and MABA using recombinant antigens; ADA was determined spectrophotometrically and CRP by ELISA. The results have shown that CRP and ADA increased linearly in relation to disease phase, CRP being significantly higher in phase III and ADA at all phases. Also, CRP and ADA were positively correlated with echocardiographic parameters of cardiac remodeling and with electrocardiographic abnormalities, and negatively with ejection fraction. CRP and ADA were higher in patients with cardiothoracic index ≥ 50%, while ADA was higher in patients with ventricular repolarization disturbances. Finally, CRP was positively correlated with ADA. In conclusion, ADA and CRP are prognostic markers of cardiac dysfunction and remodeling in Chagas disease.

  19. ADENOSINE DEAMINASE ACTIVITY AND SERUM C-REACTIVE PROTEIN AS PROGNOSTIC MARKERS OF CHAGAS DISEASE SEVERITY

    PubMed Central

    BRAVO-TOBAR, Iván Darío; NELLO-PÉREZ, Carlota; FERNÁNDEZ, Alí; MOGOLLÓN, Nora; PÉREZ, Mary Carmen; VERDE, Juan; CONCEPCIÓN, Juan Luis; RODRIGUEZ-BONFANTE, Claudina; BONFANTE-CABARCAS, Rafael

    2015-01-01

    SUMMARY Chagas disease is a public health problem worldwide. The availability of diagnostic tools to predict the development of chronic Chagas cardiomyopathy is crucial to reduce morbidity and mortality. Here we analyze the prognostic value of adenosine deaminase serum activity (ADA) and C-reactive protein serum levels (CRP) in chagasic individuals. One hundred and ten individuals, 28 healthy and 82 chagasic patients were divided according to disease severity in phase I (n = 35), II (n = 29), and III (n = 18). A complete medical history, 12-lead electrocardiogram, chest X-ray, and M-mode echocardiogram were performed on each individual. Diagnosis of Chagas disease was confirmed by ELISA and MABA using recombinant antigens; ADA was determined spectrophotometrically and CRP by ELISA. The results have shown that CRP and ADA increased linearly in relation to disease phase, CRP being significantly higher in phase III and ADA at all phases. Also, CRP and ADA were positively correlated with echocardiographic parameters of cardiac remodeling and with electrocardiographic abnormalities, and negatively with ejection fraction. CRP and ADA were higher in patients with cardiothoracic index ≥ 50%, while ADA was higher in patients with ventricular repolarization disturbances. Finally, CRP was positively correlated with ADA. In conclusion, ADA and CRP are prognostic markers of cardiac dysfunction and remodeling in Chagas disease. PMID:26603224

  20. Mining functional relationships in feature subspaces from gene expression profiles and drug activity profiles.

    PubMed

    Bao, Lei; Guo, Tao; Sun, Zhirong

    2002-04-10

    In an effort to determine putative functional relationships between gene expression patterns and drug activity patterns of 60 human cancer cell lines, a novel method was developed to discover local associations within cell line subsets. The association of drug-gene pairs is an explorative way of discovering gene markers that predict clinical tumor sensitivity to therapy. Nine drug-gene networks were discovered, as well as dozens of gene-gene and drug-drug networks. Three drug-gene networks with well studied members were discussed and the literature shows that hypothetical functional relationships exist. Therefore, this method enables the gathering of new information beyond global associations.

  1. Osteosarcoma is characterised by reduced expression of markers of osteoclastogenesis and antigen presentation compared with normal bone

    PubMed Central

    Endo-Munoz, L; Cumming, A; Sommerville, S; Dickinson, I; Saunders, N A

    2010-01-01

    Background: Osteosarcoma (OS) is the most common primary bone tumour in children and adolescents. Patients who respond poorly to chemotherapy have a higher risk of metastatic disease and 5-year survival rates of only 10–20%. Therefore, identifying molecular targets that are specific for OS, or more specifically, metastatic OS, will be critical to the development of new treatment strategies to improve patient outcomes. Methods: We performed a transcriptomic analysis of chemo-naive OS biopsies and non-malignant bone biopsies to identify differentially expressed genes specific to OS, which could provide insight into OS biology and chemoresistance. Results: Statistical analysis of the OS transcriptomes found differential expression of several metallothionein family members, as well as deregulation of genes involved in antigen presentation. Tumours also exhibited significantly increased expression of ID1 and profound down-regulation of S100A8, highlighting their potential as therapeutic targets for OS. Finally, we found a significant correlation between OS and impaired osteoclastogenesis and antigen-presenting activity. The reduced osteoclastogenesis and antigen-presenting activity were more profound in the chemoresistant OS samples. Conclusion: Our results indicate that OS displays gene signatures consistent with decreased antigen-presenting activity, enhanced chemoresistance, and impaired osteoclastogenesis. Moreover, these alterations are more pronounced in chemoresistant OS tumour samples. PMID:20551950

  2. Constitutive expression of the embryonic stem cell marker OCT4 in bovine somatic donor cells influences blastocysts rate and quality after nucleus transfer.

    PubMed

    Rodríguez-Alvarez, Lleretny; Manriquez, Jose; Velasquez, Alejandra; Castro, Fidel Ovidio

    2013-10-01

    Nuclear transfer (NT) is associated with epigenetic reprogramming of donor cells. Expression of certain genes in these cells might facilitate their expression in the NT embryo. This research was aimed to investigate the effect of constitutive expression of OCT4 in bovine somatic cells used for NT on the developmental potential of derived cloned embryos as well as in the expression of pluripotency markers in the Day-7 resulting embryos. Cloned blastocysts were generated from five cell lines that expressed OCT4. Pools of blastocysts were screened to detect OCT4, SOX2, and NANOG by qPCR. In vitro-fertilized time-matched blastocysts were used as controls. The development potential was assessed on the basis of blastocysts rate; grading and total cell counts at Day 7. OCT4 expression in the cell lines positively correlates with blastocysts rate (r = 0.92; p = 0.02), number of grade I blastocysts (r = 0.96; p = 0.01), and total cell number (r = 0.98; p = 0.002). The high expression of OCT4 in the cell line did not improve the final outcome of cloning. Somatic expression of OCT4 lead to increased expression of OCT4 and SOX2 in cloned grade I blastocysts; however, there was a bigger variability in OCT4 and SOX2 (p = 0.03; p = 0.02) expression in the embryos generated from cells expressing highest levels of OCT4. Probably the higher variability in OCT4 expression in cloned embryos is due to incorrect reprogramming and incapability of the oocyte to correct for higher OCT4 levels. For that reason, we concluded that OCT4 expression in somatic cells is not a good prognosis marker for selecting cell lines. PMID:23846396

  3. C-fos expression in the pons and medulla of the cat during carbachol-induced active sleep.

    PubMed

    Yamuy, J; Mancillas, J R; Morales, F R; Chase, M H

    1993-06-01

    Microinjection of carbachol into the rostral pontine tegmentum of the cat induces a state that is comparable to naturally occurring active (REM, rapid eye movement) sleep. We sought to determine, during this pharmacologically induced behavioral state, which we refer to as active sleep-carbachol, the distribution of activated neuron within the pons and medulla using c-fos immunocytochemistry as a functional marker. Compared with control cats, which were injected with saline, active sleep-carbachol cats exhibited higher numbers of c-fos-expressing neurons in (1) the medial and portions of the lateral reticular formation of the pons and medulla, (2) nuclei in the dorsolateral rostral pons, (3) various raphe nuclei, including the dorsal, central superior, magnus, pallidus, and obscurus, (4) the medial and lateral vestibular, prepositus hypoglossi, and intercalatus nuclei, and (5) the abducens nuclei. On the other hand, the mean number of c-fos-expressing neurons found in the masseter, facial, and hypoglossal nuclei was lower in carbachol-injected than in control cats. The data indicate that c-fos expression can be employed as a marker of state-dependent neuronal activity. The specific sites in which there were greater numbers of c-fos-expressing neurons during active sleep-carbachol are discussed in relation to the state of active sleep, as well as the functional role that these sites play in generating the various physiological patterns of activity that occur during this state. PMID:8501533

  4. The Transcription Factor MIST1 Is a Novel Human Gastric Chief Cell Marker Whose Expression Is Lost in Metaplasia, Dysplasia, and Carcinoma

    PubMed Central

    Lennerz, Jochen K. M.; Kim, Seok-Hyung; Oates, Edward L.; Huh, Won Jae; Doherty, Jason M.; Tian, Xiaolin; Bredemeyer, Andrew J.; Goldenring, James R.; Lauwers, Gregory Y.; Shin, Young-Kee; Mills, Jason C.

    2010-01-01

    The lack of reliable molecular markers for normal differentiated epithelial cells limits understanding of human gastric carcinogenesis. Recognized precursor lesions for gastric adenocarcinoma are intestinal metaplasia and spasmolytic polypeptide expressing metaplasia (SPEM), defined here by ectopic CDX2 and TFF2 expression, respectively. In mice, expression of the bHLH transcription factor MIST1, normally restricted to mature chief cells, is down-regulated as chief cells undergo experimentally induced metaplasia. Here, we show MIST1 expression is also a specific marker of human chief cells. SPEM, with and without MIST1, is present in human lesions and, akin to murine data, likely represents transitional (TFF2+/MIST1+ = “hybrid”-SPEM) and established (TFF2+/MIST1− = SPEM) stages. Co-visualization of MIST1 and CDX2 shows similar progressive loss of MIST1 with a transitional, CDX2+/MIST1− hybrid-intestinal metaplasia stage. Interinstitutional analysis and comparison of findings in tissue microarrays, resection specimens, and biopsies (n > 400 samples), comprising the entire spectrum of recognized stages of gastric carcinogenesis, confirm MIST1 expression is restricted to the chief cell compartment in normal oxyntic mucosa, rare in established metaplastic lesions, and lost in intraepithelial neoplasia/dysplasia and carcinoma of various types with the exception of rare chief cell carcinoma (∼1%). Our findings implicate MIST1 as a reliable marker of mature, healthy chief cells, and we provide the first evidence that metaplasia in humans arises at least in part from the chief cell lineage. PMID:20709804

  5. The dynamic expression of extraembryonic marker genes in the beetle Tribolium castaneum reveals the complexity of serosa and amnion formation in a short germ insect.

    PubMed

    Sharma, Rahul; Beermann, Anke; Schröder, Reinhard

    2013-12-01

    Most insect embryos develop with two distinct extraembryonic membranes, the serosa and the amnion. In the insect beetle Tribolium the early origin of the serosa within the anterior blastoderm is well established but the origin of the amnion is still debated. It is not known whether this tissue develops from a blastodermal precursor or originates de novo later from embryonic tissue during embryogenesis. We undertook an in-depth analysis of the spatio-temporal expression pattern profile of important extraembryonic membrane marker genes with emphasis on early blastoderm development in Tribolium. The amnion marker iroquois (Tc-iro) was found co-expressed with the serosa marker zerknüllt1 (Tc-zen1) during early blastoderm formation in an anterior cap domain. This domain later resolved into two adjacent domains that likely represent the precursors of the serosa and the amnion. In addition, we found the hindsight ortholog in Tribolium (Tc-hnt) to be a serosa-specific marker. Surprisingly, decapentaplegic (Tc-dpp) expression was not seen as a symmetric cap domain but detected asymmetrically first along the DV- and later also along the AP-axis. Moreover, we found a previously undescribed domain of phosphorylated MAD (pMAD) protein in anterior ventral serosal cells. This is the first study showing that the anterior-lateral part of the amnion originates from the anterior blastoderm while the precursor of the dorsal amnion develops later de novo from a dorsal-posterior region within the differentiated blastoderm.

  6. Global microRNA expression profiling uncovers molecular markers for classification and prognosis in aggressive B-cell lymphoma.

    PubMed

    Iqbal, Javeed; Shen, Yulei; Huang, Xin; Liu, Yanyan; Wake, Laura; Liu, Cuiling; Deffenbacher, Karen; Lachel, Cynthia M; Wang, Chao; Rohr, Joseph; Guo, Shuangping; Smith, Lynette M; Wright, George; Bhagavathi, Sharathkumar; Dybkaer, Karen; Fu, Kai; Greiner, Timothy C; Vose, Julie M; Jaffe, Elaine; Rimsza, Lisa; Rosenwald, Andreas; Ott, German; Delabie, Jan; Campo, Elias; Braziel, Rita M; Cook, James R; Tubbs, Raymond R; Armitage, James O; Weisenburger, Dennis D; Staudt, Louis M; Gascoyne, Randy D; McKeithan, Timothy W; Chan, Wing C

    2015-02-12

    We studied the global microRNA (miRNA) expression in diffuse large B-cell lymphoma (DLBCL; n = 79), Burkitt lymphoma (BL; n = 36), primary mediastinal B-cell lymphoma (PMBL; n = 12), B-cell lines (n = 11), and normal subsets of naïve B cells, centroblasts (CBs), and peripheral blood B cells along with their corresponding gene expression profiles (GEPs). The normal B-cell subsets have well-defined miRNA signatures. The CB miRNA signature was significantly associated with germinal center B-cell (GCB)-DLBCL compared with activated B-cell (ABC)-DLBCL (P = .002). We identified a 27-miRNA signature that included v-myc avian myelomatosis viral oncogene homolog (MYC) targets and enabled the differentiation of BL from DLBCL, a distinction comparable with the "gold standard" GEP-defined diagnosis. Distinct miRNA signatures were identified for DLBCL subgroups, including GCB-DLBCL, activated B-cell (ABC)-DLBCL, and PMBL. Interestingly, most of the unclassifiable-DLBCL by GEP showed a strong similarity to the ABC-DLBCL by miRNA expression profiling. Consistent results for BL and DLBCL subgroup classification were observed in formalin-fixed, paraffin-embedded tissue, making such tests practical for clinical use. We also identified predictive miRNA biomarker signatures in DLBCL, including high expression of miR-155, which is significantly associated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) treatment failure. This finding was further supported by the observation that high expression of miR-155 sensitizes cells to v-akt murine thymoma viral oncogene homolog-1 inhibitors in vitro, suggesting a novel treatment option for resistant DLBCL. PMID:25498913

  7. Global microRNA expression profiling uncovers molecular markers for classification and prognosis in aggressive B-cell lymphoma

    PubMed Central

    Shen, Yulei; Huang, Xin; Liu, Yanyan; Wake, Laura; Liu, Cuiling; Deffenbacher, Karen; Lachel, Cynthia M.; Wang, Chao; Rohr, Joseph; Guo, Shuangping; Smith, Lynette M.; Wright, George; Bhagavathi, Sharathkumar; Dybkaer, Karen; Fu, Kai; Greiner, Timothy C.; Vose, Julie M.; Jaffe, Elaine; Rimsza, Lisa; Rosenwald, Andreas; Ott, German; Delabie, Jan; Campo, Elias; Braziel, Rita M.; Cook, James R.; Tubbs, Raymond R.; Armitage, James O.; Weisenburger, Dennis D.; Staudt, Louis M.; Gascoyne, Randy D.; McKeithan, Timothy W.; Chan, Wing C.

    2015-01-01

    We studied the global microRNA (miRNA) expression in diffuse large B-cell lymphoma (DLBCL; n = 79), Burkitt lymphoma (BL; n = 36), primary mediastinal B-cell lymphoma (PMBL; n = 12), B-cell lines (n = 11), and normal subsets of naïve B cells, centroblasts (CBs), and peripheral blood B cells along with their corresponding gene expression profiles (GEPs). The normal B-cell subsets have well-defined miRNA signatures. The CB miRNA signature was significantly associated with germinal center B-cell (GCB)–DLBCL compared with activated B-cell (ABC)–DLBCL (P = .002). We identified a 27-miRNA signature that included v-myc avian myelomatosis viral oncogene homolog (MYC) targets and enabled the differentiation of BL from DLBCL, a distinction comparable with the “gold standard” GEP-defined diagnosis. Distinct miRNA signatures were identified for DLBCL subgroups, including GCB-DLBCL, activated B-cell (ABC)-DLBCL, and PMBL. Interestingly, most of the unclassifiable-DLBCL by GEP showed a strong similarity to the ABC-DLBCL by miRNA expression profiling. Consistent results for BL and DLBCL subgroup classification were observed in formalin-fixed, paraffin-embedded tissue, making such tests practical for clinical use. We also identified predictive miRNA biomarker signatures in DLBCL, including high expression of miR-155, which is significantly associated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) treatment failure. This finding was further supported by the observation that high expression of miR-155 sensitizes cells to v-akt murine thymoma viral oncogene homolog-1 inhibitors in vitro, suggesting a novel treatment option for resistant DLBCL. PMID:25498913

  8. [THE COMBINATION OF EXPRESSION OF MARKERS CR1 AND CR2 (CD35/CD21) IN DIAGNOSTIC OF B-CELL LYMPHOPROLIFERATIVE DISEASES].

    PubMed

    Chibisova, O N; Burtsev, D V; Galstian, K M; Lugovskaia, G I

    2015-04-01

    The study was carried out to analyze rate of expression of antigens CD35 and CD21 in norm and under different forms of B-cell lymphoproliferative diseases. The level of average intensity of fluorescence of antigens CD35, CD21 and CD200 is compared for different groups of patients with B-cell lymphoproliferative diseases. The established patterns of expression of antigens CD35 and CD21 under B-cell lymphoproliferative diseases permit considering expression of the given markers as a characteristic of differential diagnostic. PMID:26189291

  9. Stromal cells in phyllodes tumors of the breast are enriched for EZH2 and stem cell marker expression.

    PubMed

    Zhang, Yanhong; Liss, Adam L; Chung, Eugene; Pierce, Lori J; Kleer, Celina G

    2016-07-01

    Phyllodes tumors (PTs) of the breast are fibroepithelial neoplasms with stromal hypercellularity, which is the basis for their classification as benign, borderline, and malignant. The histologic diagnosis of PTs is often difficult, and the pathological features may not always predict clinical behavior. The pathobiology of PT remains poorly understood. Enhancer of Zeste 2 (EZH2) epigenetically regulates cell-type identity, cellular differentiation, and breast cancer stem cells. EZH2 exerts oncogenic functions in breast cancer and is associated with metastasis. We hypothesized that in PTs, EZH2 and the stem cell marker ALDH1 may be expressed in stromal cells and may be associated with their degree of differentiation. Forty PTs were histologically characterized at our institution following the World Health Organization criteria. We investigated the expression of EZH2 and ALDH1 by immunohistochemistry and recorded as percentage of positive epithelial and stromal cells. EZH2 was positive when over 10 % of cells exhibited nuclear staining; ALDH1 was positive when over 5 % of cells had cytoplasmic staining. Of the 40 PTs, 24 (60 %) were histologically benign, 8 (20 %) borderline, and 8 (20 %) malignant. Stromal EZH2 was significantly associated with the diagnosis of malignant PT, as it was detected in 1 of 24 (4 %) benign, 3 of 8 (37.5 %) borderline, and 5 of 8 (62.5 %) malignant tumors. Stromal EZH2 was significantly associated with stromal overgrowth (p = 0.01), atypia (p = 0.01), hypercellularity (p = 0.01), and mitoses (p = 0.02), all features of malignant PT. Stromal EZH2 and ALDH1 were significantly associated with grade of PT (p = 0.01 and p < 0.05 respectively). In conclusion, EZH2 and ALDH1 expression in the stroma of PT may mark malignant progression and may be helpful to distinguish histologically benign from borderline and malignant tumors in challenging cases. Our study also suggests that PTs contain mesenchymal stem cells, shedding light

  10. Patterns of activity expressed by juvenile horseshoe crabs.

    PubMed

    Dubofsky, E A; Simpson, S D; Chabot, Christopher C; Watson, Winsor H

    2013-09-01

    Adult American horseshoe crabs, Limulus polyphemus, possess endogenous circadian and circatidal clocks controlling visual sensitivity and locomotion, respectively. The goal of this study was to determine the types of activity rhythms expressed by juvenile horseshoe crabs (n = 24) when exposed to a 14:10 light/dark cycle (LD) for 10 days, followed by 10 days of constant darkness (DD). Horseshoe crab activity was recorded with a digital time-lapse video system that used an infrared-sensitive camera so animals could be monitored at night. In LD, 15 animals expressed daily patterns of activity, 6 displayed a circatidal pattern, and the remaining 3 were arrhythmic. Of the 15 animals with daily patterns of locomotion, 7 had a significant preference (P < 0.05) for diurnal activity and 3 for nocturnal activity; the remainder did not express a significant preference for day or night activity. In DD, 13 horseshoe crabs expressed circatidal rhythms and 8 maintained a pattern of about 24 h. Although these results suggest the presence of a circadian clock influencing circatidal patterns of locomotion, these apparent circadian rhythms may actually represent the expression of just one of the two bouts of activity driven by the putative circalunidian clocks that control their tidal rhythms. Overall, these results indicate that, like adults, juvenile horseshoe crabs express both daily and tidal patterns of activity and that at least one, and maybe both, of these patterns is driven by endogenous clocks. PMID:24088795

  11. Patterns of activity expressed by juvenile horseshoe crabs.

    PubMed

    Dubofsky, E A; Simpson, S D; Chabot, Christopher C; Watson, Winsor H

    2013-09-01

    Adult American horseshoe crabs, Limulus polyphemus, possess endogenous circadian and circatidal clocks controlling visual sensitivity and locomotion, respectively. The goal of this study was to determine the types of activity rhythms expressed by juvenile horseshoe crabs (n = 24) when exposed to a 14:10 light/dark cycle (LD) for 10 days, followed by 10 days of constant darkness (DD). Horseshoe crab activity was recorded with a digital time-lapse video system that used an infrared-sensitive camera so animals could be monitored at night. In LD, 15 animals expressed daily patterns of activity, 6 displayed a circatidal pattern, and the remaining 3 were arrhythmic. Of the 15 animals with daily patterns of locomotion, 7 had a significant preference (P < 0.05) for diurnal activity and 3 for nocturnal activity; the remainder did not express a significant preference for day or night activity. In DD, 13 horseshoe crabs expressed circatidal rhythms and 8 maintained a pattern of about 24 h. Although these results suggest the presence of a circadian clock influencing circatidal patterns of locomotion, these apparent circadian rhythms may actually represent the expression of just one of the two bouts of activity driven by the putative circalunidian clocks that control their tidal rhythms. Overall, these results indicate that, like adults, juvenile horseshoe crabs express both daily and tidal patterns of activity and that at least one, and maybe both, of these patterns is driven by endogenous clocks.

  12. Smooth Muscle-Like Cells Generated from Human Mesenchymal Stromal Cells Display Marker Gene Expression and Electrophysiological Competence Comparable to Bladder Smooth Muscle Cells

    PubMed Central

    Brun, Juliane; Lutz, Katrin A.; Neumayer, Katharina M. H.; Klein, Gerd; Seeger, Tanja; Uynuk-Ool, Tatiana; Wörgötter, Katharina; Schmid, Sandra; Kraushaar, Udo; Guenther, Elke; Rolauffs, Bernd; Aicher, Wilhelm K.; Hart, Melanie L.

    2015-01-01

    The use of mesenchymal stromal cells (MSCs) differentiated toward a smooth muscle cell (SMC) phenotype may provide an alternative for investigators interested in regenerating urinary tract organs such as the bladder where autologous smooth muscle cells cannot be used or are unavailable. In this study we measured the effects of good manufacturing practice (GMP)-compliant expansion followed by myogenic differentiation of human MSCs on the expression of a range of contractile (from early to late) myogenic markers in relation to the electrophysiological parameters to assess the functional role of the differentiated MSCs and found that differentiation of MSCs associated with electrophysiological competence comparable to bladder SMCs. Within 1–2 weeks of myogenic differentiation, differentiating MSCs significantly expressed alpha smooth muscle actin (αSMA; ACTA2), transgelin (TAGLN), calponin (CNN1), and smooth muscle myosin heavy chain (SM-MHC; MYH11) according to qRT-PCR and/or immunofluorescence and Western blot. Voltage-gated Na+ current levels also increased within the same time period following myogenic differentiation. In contrast to undifferentiated MSCs, differentiated MSCs and bladder SMCs exhibited elevated cytosolic Ca2+ transients in response to K+-induced depolarization and contracted in response to K+ indicating functional maturation of differentiated MSCs. Depolarization was suppressed by Cd2+, an inhibitor of voltage-gated Ca2+-channels. The expression of Na+-channels was pharmacologically identified as the Nav1.4 subtype, while the K+ and Ca2+ ion channels were identified by gene expression of KCNMA1, CACNA1C and CACNA1H which encode for the large conductance Ca2+-activated K+ channel BKCa channels, Cav1.2 L-type Ca2+ channels and Cav3.2 T-type Ca2+ channels, respectively. This protocol may be used to differentiate adult MSCs into smooth muscle-like cells with an intermediate-to-late SMC contractile phenotype exhibiting voltage-gated ion channel

  13. Comparative Analysis of Immune Activation Markers of CD8+ T Cells in Lymph Nodes of Different Origins in SIV-Infected Chinese Rhesus Macaques

    PubMed Central

    Liu, Jinbiao; Xiao, Qianhao; Zhou, Runhong; Wang, Yong; Xian, Qiaoyang; Ma, Tongcui; Zhuang, Ke; Zhou, Li; Guo, Deyin; Wang, Xu; Ho, Wen-Zhe; Li, Jieliang

    2016-01-01

    Altered T-cell homeostasis, such as expansion of CD8+ T cells to the secondary lymphatic compartments, has been suggested as a mechanism of HIV/simian immunodeficiency virus (SIV)-pathogenesis. However, the role of immune activation of CD8+ T cells in the CD4/CD8 turnover and viral replication in these tissues is not completely understood. In this study, we compared the expression of immune activation markers (CD69 and HLA-DR) on CD8+ T cells in the peripheral blood and lymph nodes (LNs) of SIV-infected/uninfected Chinese rhesus macaques. SIV-infected macaques had significantly higher percentages of CD8+CD69+ and CD8+HLA-DR+ T cells in all these anatomical compartments than uninfected macaques. LNs that located close to the gastrointestinal (GI) tract (colon, mesenteric, and iliac LNs) of SIV-infected macaques had profoundly lower numbers of CD4+ T cells, but no significant difference in expression of activation marker (CD8+CD69+ and CD8+HLA-DR+) as compared with the peripheral lymphatic tissues (axillary and inguinal LNs). The CD4/CD8 ratios were negatively correlated with the activation of CD8+ T cells in the overall LNs, with further associations with CD8+HLA-DR+ in GI LNs while CD8+CD69+ in peripheral LNs. These observations demonstrate that the increase of CD8+ T cell activation is a contributing factor for the decline of CD4/CD8 ratios in GI system. PMID:27708644

  14. Isomorphisms between psychological processes and neural mechanisms: from stimulus elements to genetic markers of activity.

    PubMed

    Fanselow, Michael S; Zelikowsky, Moriel; Perusini, Jennifer; Barrera, Vanessa Rodriguez; Hersman, Sarah

    2014-02-01

    Traditional learning theory has developed models that can accurately predict and describe the course of learned behavior. These "psychological process" models rely on hypothetical constructs that are usually thought to be not directly measurable or manipulable. Recently, and mostly in parallel, the neural mechanisms underlying learning have been fairly well elucidated. The argument in this essay is that we can successfully uncover isomorphisms between process and mechanism and that this effort will help advance our theories about both processes and mechanisms. We start with a brief review of error-correction circuits as a successful example. Then we turn to the concept of stimulus elements, where the conditional stimulus is hypothesized to be constructed of a multitude of elements only some of which are sampled during any given experience. We discuss such elements with respect to how they explain acquisition of associative strength as an incremental process. Then we propose that for fear conditioning, stimulus elements and basolateral amygdala projection neurons are isomorphic and that the activational state of these "elements" can be monitored by the expression of the mRNA for activity-regulated cytoskeletal protein (ARC). Finally we apply these ideas to analyze recent data examining ARC expression during contextual fear conditioning and find that there are indeed many similarities between stimulus elements and amygdala neurons. The data also suggest some revisions in the conceptualization of how the population of stimulus elements is sampled from.

  15. Bone marrow-derived and peritoneal macrophages have different inflammatory response to oxLDL and M1/M2 marker expression – implications for atherosclerosis research

    PubMed Central

    Bisgaard, Line S.; Mogensen, Christina K.; Rosendahl, Alexander; Cucak, Helena; Nielsen, Lars Bo; Rasmussen, Salka E.; Pedersen, Tanja X.

    2016-01-01

    Macrophages are heterogeneous and can polarize into specific subsets, e.g. pro-inflammatory M1-like and re-modelling M2-like macrophages. To determine if peritoneal macrophages (PEMs) or bone marrow derived macrophages (BMDMs) resembled aortic macrophages from ApoE−/− mice, their M1/M2 phenotype, inflammatory status, and lipid metabolism signatures were compared. oxLDL accumulation was similar in PEMs and BMDMs. On protein expression level, BMDMs showed an M2-like CD206highCD11clow profile, while cholesterol loading led to enhanced CD11c expression and reduced MCP-1 secretion. In contrast, PEMs expressed low levels of CD206 and CD11c, and responded to cholesterol loading by increasing CD11c expression and MCP-1 secretion. mRNA expression of M1/M2 markers was higher in PEMS than BMDMs, while lipid metabolism genes were similarly expressed. Whole aorta flow cytometry showed an accumulation of M2-like CD206highCD11clow macrophages in advanced versus early atherosclerotic disease in ApoE−/− mice. In isolated lesions, mRNA levels of the M2 markers Socs2, CD206, Retnla, and IL4 were downregulated with increasing disease severity. Likewise, mRNA expression of lipid metabolism genes (SREBP2, ACSL1, SRB1, DGAT1, and cpt1a) was decreased in advanced versus early lesions. In conclusion, PEMs and BMDMs are phenotypically distinct and differ from macrophages in lesions with respect to expression of M1/M2 markers and lipid metabolism genes. PMID:27734926

  16. Robust, synergistic regulation of human gene expression using TALE activators.

    PubMed

    Maeder, Morgan L; Linder, Samantha J; Reyon, Deepak; Angstman, James F; Fu, Yanfang; Sander, Jeffry D; Joung, J Keith

    2013-03-01

    Artificial activators designed using transcription activator-like effector (TALE) technology have broad utility, but previous studies suggest that these monomeric proteins often exhibit low activities. Here we demonstrate that TALE activators can robustly function individually or in synergistic combinations to increase expression of endogenous human genes over wide dynamic ranges. These findings will encourage applications of TALE activators for research and therapy, and guide design of monomeric TALE-based fusion proteins.

  17. Carcinogen-induced trans activation of gene expression

    SciTech Connect

    Kleinberger, T.; Flint, Y.B.; Blank, M.; Etkin, S.; Lavi, S.

    1988-03-01

    The authors report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later.

  18. Carcinogen-induced trans activation of gene expression.

    PubMed Central

    Kleinberger, T; Flint, Y B; Blank, M; Etkin, S; Lavi, S

    1988-01-01

    We report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later. Images PMID:2835673

  19. Epidermal Cells Expressing Putative Cell Markers in Nonglabrous Skin Existing in Direct Proximity with the Distal End of the Arrector Pili Muscle

    PubMed Central

    Rufaut, N. W.; Jones, L.; Sinclair, R.

    2016-01-01

    Inconsistent with the view that epidermal stem cells reside randomly spread along the basal layer of the epidermal rete ridges, we found that epidermal cells expressing stem cell markers in nonglabrous skin exist in direct connection with the distal end of the arrector pili muscle. The epidermal cells that express stem cell markers consist of a subpopulation of basal keratinocytes located in a niche at the lowermost portion of the rete ridges at the distal arrector pili muscle attachment site. Keratinocytes in the epidermal stem cell niche express K15, MCSP, and α6 integrin. α5 integrin marks the distal end of the APM colocalized with basal keratinocytes expressing stem cell markers located in a well-protected and nourished environment at the lowermost point of the epidermis; these cells are hypothesized to participate directly in epidermal renewal and homeostasis and also indirectly in wound healing through communication with the hair follicle bulge epithelial stem cell population through the APM. Our findings, plus a reevaluation of the literature, support the hierarchical model of interfollicular epidermal stem cell units of Fitzpatrick. This new view provides insights into epidermal control and the possible involvement of epidermal stem cells in nonmelanoma skin carcinogenesis. PMID:27375744

  20. Expression of the carcinoma markers: the sialylated Lewis A and X carbohydrate antigens in normal laryngeal surface epithelium and submucosal glands from old humans.

    PubMed

    Kirkeby, Svend; Moe, Dennis

    2013-03-01

    Aberrant surface expression of the carbohydrate ABH and Lewis antigens are often used as markers for the diagnosis of cancer, but while the distribution of these histo-blood group antigens is relatively well-described in tissues and organs from young and middle-aged humans little is known of their expression in old age. The objective for this study was to estimate if the Lewis A and X antigens together with their sialylated modifications, are expressed in sections of normal laryngeal tissue from old humans. Antibodies directed against the tumor markers Sialyl Lewis A and Sialyl Lewis X showed positive reaction in the surface epithelia from normal larynx autopsies obtained from people aged 77-90 years. The sialylated and non-sialylated Lewis A antigens were more frequently expressed in the pseudostratified epithelium than in squamous surface epithelium. Both the sialylated and the non-sialylated carbohydrates were stained in the submucosal glands in all the autopsies. In conclusion, visualization of Lewis tumor markers in the larynx should be interpreted with great care, as they may be present in normal laryngeal epithelial cells from old humans.

  1. Cytosine deaminase as a negative selectable marker for the microalgal chloroplast: a strategy for the isolation of nuclear mutations that affect chloroplast gene expression.

    PubMed

    Young, Rosanna E B; Purton, Saul

    2014-12-01

    Negative selectable markers are useful tools for forward-genetic screens aimed at identifying trans-acting factors that are required for expression of specific genes. Transgenic lines harbouring the marker fused to a gene element, such as a promoter, may be mutagenized to isolate loss-of-function mutants able to survive under selection. Such a strategy allows the molecular dissection of factors that are essential for expression of the gene. Expression of individual chloroplast genes in plants and algae typically requires one or more nuclear-encoded factors that act at the post-transcriptional level, often through interaction with the 5' UTR of the mRNA. To study such nuclear control further, we have developed the Escherichia coli cytosine deaminase gene codA as a conditional negative selectable marker for use in the model green alga Chlamydomonas reinhardtii. We show that a codon-optimized variant of codA with three amino acid substitutions confers sensitivity to 5-fluorocytosine (5-FC) when expressed in the chloroplast under the control of endogenous promoter/5' UTR elements from the photosynthetic genes psaA or petA. UV mutagenesis of the psaA transgenic line allowed recovery of 5-FC-resistant, photosynthetically deficient lines harbouring mutations in the nuclear gene for the factor TAA1 that is required for psaA translation. Similarly, the petA line was used to isolate mutants of the petA mRNA stability factor MCA1 and the translation factor TCA1. The codA marker may be used to identify critical residues in known nuclear factors and to aid the discovery of additional factors required for expression of chloroplast genes. PMID:25234691

  2. Breast cancer statistics and markers.

    PubMed

    Donepudi, Mallika Siva; Kondapalli, Kasturi; Amos, Seelam Jeevan; Venkanteshan, Pavithra

    2014-01-01

    Breast cancer is one of the familiar diseases in women. Incidence and mortality due to cancer, particularly breast cancer has been increasing for last 50 years, even though there is a lacuna in the diagnosis of breast cancer at early stages. According to World Health Organization (WHO) 2012 reports, breast cancer is the leading cause of death in women, accounting 23% of all cancer deaths. In Asia, one in every three women faces the risk of breast cancer in their lifetime as per reports of WHO 2012. Here, the review is been focused on different breast cancer markers, that is, tissue markers (hormone receptors, human epidermal growth factor-2, urokinase plasminogen activator, plasminogen activator inhibitor, p53 and cathepsin D), genetic markers (BRAC1 and 2 and gene expression microarray technique, etc.), and serum markers (CA 15.3, BR 27.29, MCA, CA 549, carcinoembryonic antigen, oncoproteins, and cytokeratins) used in present diagnosis, but none of the mentioned markers can diagnose breast cancer at an early stage. There is a disquieting need for the identification of best diagnosing marker, which can be able to diagnose even in early stage of breast carcinogenesis.

  3. Simple sequence repeat marker development from bacterial artificial chromosome end sequences and expressed sequence tags of flax (Linum usitatissimum L.).

    PubMed

    Cloutier, Sylvie; Miranda, Evelyn; Ward, Kerry; Radovanovic, Natasa; Reimer, Elsa; Walichnowski, Andrzej; Datla, Raju; Rowland, Gordon; Duguid, Scott; Ragupathy, Raja

    2012-08-01

    Flax is an important oilseed crop in North America and is mostly grown as a fibre crop in Europe. As a self-pollinated diploid with a small estimated genome size of ~370 Mb, flax is well suited for fast progress in genomics. In the last few years, important genetic resources have been developed for this crop. Here, we describe the assessment and comparative analyses of 1,506 putative simple sequence repeats (SSRs) of which, 1,164 were derived from BAC-end sequences (BESs) and 342 from expressed sequence tags (ESTs). The SSRs were assessed on a panel of 16 flax accessions with 673 (58 %) and 145 (42 %) primer pairs being polymorphic in the BESs and ESTs, respectively. With 818 novel polymorphic SSR primer pairs reported in this study, the repertoire of available SSRs in flax has more than doubled from the combined total of 508 of all previous reports. Among nu