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101.
Afonso Rocha Martins Almeida Sílvia Arroz-Madeira Diogo Fonseca-Pereira Hélder Ribeiro Reena Lasrado Vassilis Pachnis Henrique Veiga-Fernandes 《PloS one》2012,7(12)
Identification of thymocyte regulators is a central issue in T cell biology. Interestingly, growing evidence indicates that common key molecules control neuronal and immune cell functions. The neurotrophic factor receptor RET mediates critical functions in foetal hematopoietic subsets, thus raising the possibility that RET-related molecules may also control T cell development. We show that Ret, Gfra1 and Gfra2 are abundantly expressed by foetal and adult immature DN thymocytes. Despite the developmentally regulated expression of these genes, analysis of foetal thymi from Gfra1, Gfra2 or Ret deficient embryos revealed that these molecules are dispensable for foetal T cell development. Furthermore, analysis of RET gain of function and Ret conditional knockout mice showed that RET is also unnecessary for adult thymopoiesis. Finally, competitive thymic reconstitution assays indicated that Ret deficient thymocytes maintained their differentiation fitness even in stringent developmental conditions. Thus, our data demonstrate that RET/GFRα signals are dispensable for thymic T cell development in vivo, indicating that pharmacological targeting of RET signalling in tumours is not likely to result in T cell production failure. 相似文献
102.
103.
Papoutsi Z Kassi E Papaevangeliou D Pratsinis H Zoumpourlis V Halabalaki M Mitakou S Kalofoutis A Moutsatsou P 《Steroids》2004,69(11-12):727-734
We have isolated from the plant Onobrychis ebenoides three novel arylobenzofurans with binding affinity for the estrogen receptor. In this study, we evaluated these arylobenzofurans, namely ebenfuran I, ebenfuran II and ebenfuran III for their potential selective estrogen receptor modulator (SERM)-like properties. We examined their ability, (1) to induce the insulin growth factor binding protein-3 (IGFBP-3) in MCF-7 breast cancer cells, (2) to stimulate differentiation and mineralization of osteoblastic cell culture by histochemical staining for alkaline phosphatase, Alizarin Red-S staining and calcium levels in the supernatants and (3) to inhibit cell proliferation of cervical adenocarcinoma (Hela) cells by use of the MTT assay. An estrogen receptor mediated effect was investigated by carrying out chloramphenicol acetyl transferase (CAT) assay on transient MCF-7 transfectants. Estradiol and the "pure" antiestrogen ICI 182780 were included to serve as control samples of the estrogenic and antiestrogenic effect respectively. Our data reveal that ebenfuran II is a highly potent SERM, exhibiting antiestrogenic activity in breast cancer cells via the estrogen receptor, estrogenic effect on osteoblasts and no stimulatory effect on cervix adenocarcinoma cells. In conclusion, our study is the first to demonstrate that plant derived arylobenzofurans show a SERM profile and may be considered for the prevention and treatment of diseases such as breast cancer, cervical cancer and osteoporosis. 相似文献
104.
Ikonomopoulos J Gazouli M Pavlik I Bartos M Zacharatos P Xylouri E Papalambros E Gorgoulis V 《Journal of microbiological methods》2004,56(3):315-321
Mycobacterium avium subsp. paratuberculosis (MAP) can cause a very serious, often-fatal disease, namely paratuberculosis, in several animal species, especially ruminants. Recently, it has also been implicated in the pathogenesis of Infectious Bowel Disease of man. The aim of this study was to develop a molecular method for the routine detection and identification of MAP, from tissue samples of animal origin. The proposed assay would have to combine optimum performance and cost, with high reproducibility. To this goal, three laboratories in Greece and the Czech Republic undertook different parts of a study that involved evaluation of DNA extraction procedures, and PCR assays, for MAP detection. For DNA extraction we used one in-house, and one commercial method, and for the PCR we assessed a number of different assays, starting with the evaluation of primer specificity with an extended GenBank database search. Based on these results, we chose to assess a one-tube nested, 2 two-tube nested, and a single PCR assay, targeted to different genomic regions of the IS900 element. These four methods were applied on positive and negative control samples, consisted of pure bacterial cultures and formalin-fixed paraffin-embedded (FFPE) tissue samples collected from cattle with paratuberculosis and chickens with M. avium subsp. avium infection. Based on the criteria of reliability and cost, the procedure that performed better was the one-tube nested PCR assay combined with the in-house DNA extraction method. The agreement of the results obtained by the three collaborating laboratories indicates the reliability of the proposed assay even under different laboratory conditions. 相似文献
105.
Lamprou I Tsakas S Theodorou GL Karakantza M Lampropoulou M Marmaras VJ 《Biochimica et biophysica acta》2005,1744(1):1-10
In response to LPS/E. coli treatment, extracellular signal-regulated kinase (ERK) is activated in medfly hemocytes. To explore the molecular mechanisms underlying LPS/E. coli/latex beads endo- and phagocytosis, we studied the signalling pathways leading to p38 and c-jun N-terminal kinase (JNK) activation. JNK and p38-like proteins were initially identified within medfly hemocytes. Flow cytometry analysis revealed that mitogen-activated protein kinases (MAPK) are required for phagocytosis. Inhibition of specific MAPK signalling pathways, with manumycin A, toxin A, cytochalasin D and latrunculin A, revealed activation of p38 via Ras/Rho/actin remodelling pathway and activation of JNK that was independent of actin cytoskeleton reorganization. ERK and p38 pathways, but not JNK, appeared to be involved in LPS-dependent hemocyte secretion, whereas all MAPK subfamilies seemed to participate in E. coli-dependent secretion. In addition, flow cytometry experiments in hemocytes showed that the LPS/E. coli-induced release was a prerequisite for LPS/E. coli uptake, whereas latex bead phagocytosis did not depend on hemocyte secretion. This is a novel aspect, as in mammalian monocytes/macrophages LPS/E. coli-triggered release has not been yet correlated with phagocytosis. It is of interest that these data suggest distinct mechanisms for the phagocytosis of E. coli and latex beads in medfly hemocytes. 相似文献
106.
Linehan JD Kolios G Valatas V Robertson DA Westwick J 《American journal of physiology. Gastrointestinal and liver physiology》2005,288(2):G261-G267
Nitric oxide (NO) production is increased in the human colonic mucosa in intestinal inflammation. We examined the effect of corticosteroids and the role of mononuclear cells in this production. Colonic biopsies from patients with ulcerative colitis and normal controls were cultured with either budesonide or prednisolone in the presence of proinflammatory cytokines. Human mixed mononuclear cells (MMCs) were cocultured with HT-29 cells stimulated with IFN-gamma and LPS in the presence or absence of corticosteroids. Nitrite production was measured in supernatants by a modification of the Griess reaction, and inducible NO synthase (iNOS) mRNA expression was studied in colonic tissue by RT-PCR. Both steroids significantly suppressed the nitrite production and iNOS mRNA expression in inflamed colonic biopsies from ulcerative colitis patients and in cytokine-stimulated normal colonic biopsies but not in cytokine-stimulated HT-29 cells. Nitrite production by HT-29 cells was significantly increased (P < 0.01) in cocultures with MMCs stimulated with IFN-gamma and LPS. The presence of either prednisolone or budesonide significantly (P < 0.01) suppressed nitrite production from cocultures of HT-29 cells and MMCs but not from cultures of HT-29 cells stimulated with conditioned media from activated MMCs. Interestingly, stimulation of HT-29 with conditioned media from MMCs pretreated with steroids before stimulation with LPS and IFN-gamma induced a significantly (P < 0.01) lower nitrite production. These results suggest that the inhibitory effect of corticosteroids on the NO production in the intestinal inflammation might be via the inhibition of MMC-produced mediators responsible for NO production by colonic epithelial cells. 相似文献
107.
Paschalis V Koutedakis Y Jamurtas AZ Mougios V Baltzopoulos V 《Journal of strength and conditioning research / National Strength & Conditioning Association》2005,19(1):184-188
We examined differences in muscle damage and muscle performance perturbations in relation to the same volumes of high (HI) and low intensity (LI) of eccentric exercise. Untrained young healthy men (n = 12) underwent 2 isokinetic quadriceps eccentric exercise sessions, 1 on each randomly selected leg, separated by a 2-week interval. In the first session subjects performed HI exercise (i.e., 12 sets of 10 maximal voluntary efforts). In the second session, volunteers were subjected to continuous exercise of LI (50% of peak torque) until the total work done was approximately equal to that generated during HI. Muscle damage (serum creatine kinase concentration [CK], delayed onset of muscle soreness, and range of motion) and muscle performance (eccentric [EPT] and isometric peak torque [IPT]) indicators were assessed pre-exercise and 24, 48, 72, and 96 hours postexercise. Compared to baseline data, changes in muscle damage indicators were significantly different (p < 0.05) at almost all postexercise time points in both conditions. However, apart from the significant elevation of CK at 24 hours after HI (p < 0.05), no other significant differences were observed between the 2 exercise conditions (p > 0.05). The main finding in relation to muscle performance was that decrements following HI exercise were significantly greater (p < 0.05) compared to LI. Compared with baseline data, the EPT values following HI and LI exercise were as follows: 24 hours, 72.1% vs. 92%; 48 hours, 81.9% vs. 94.8%; 72 hours, 77.7% vs. 100.6%; 96 hours, 86.8% vs. 107.9%. The corresponding data for IPT were as follows: 24 hours, 86.4% vs. 102.8%; 48 hours, 84.2% vs. 107%; 72 hours, 84.8% vs. 109.2%; 96 hours, 86.8% vs. 114.4%. These results indicate that matching volumes of HI and LI eccentric exercise have similar effects on muscle damage, but HI has a more prominent effect on muscle performance. 相似文献
108.
Petrou P Pavlakis E Dalezios Y Galanopoulos VK Chalepakis G 《The Journal of biological chemistry》2005,280(11):10350-10356
Fras1 is a putative extracellular matrix protein that has been implicated in the structural adhesion of embryonic epidermis to dermis. Moreover, mutations in Fras1/FRAS1 have been associated with the mouse blebbed phenotype and the human rare genetic disorder Fraser syndrome, respectively. Here we report the mapping of Fras1 within the extracellular space and evaluate the effects of Fras1 deficiency on lung development in the mouse. Expression of Fras1 was detected in the mesothelial cells of the visceral pleura and in the conducting airway epithelia. Immunogold histochemistry identified Fras1 as a component of the extracellular matrix localized below the lamina densa of epithelial basement membranes in the embryonic lung. Embryos homozygous for a targeted mutation of Fras1 exhibited fused pulmonary lobes resulting from incomplete separation during development as well as a profound disarrangement of blood capillaries in the terminal air sacs. We demonstrate that loss of Fras1 causes alterations in the molecular composition of basement membranes, concomitant with local disruptions of epithelial-endothelial contacts and extravasation of erythrocytes into the embryonic respiratory lumen. Thus, our findings identify Fras1 as an important structural component of the sub-lamina densa of basement membranes required for lobar septation and the organization of blood capillaries in the peripheral lung. 相似文献
109.
Vassilis Bitsikas Kirsi Riento Jonathan D. Howe Nicholas P. Barry Benjamin J. Nichols 《PloS one》2014,9(1)
Flotillin 1 and flotillin 2 associate in the plasma membrane to form microdomains that have roles in cell signaling, regulation of cell-cell contacts, membrane-cytoskeletal interactions, and endocytosis. They are thought to be involved in the trafficking and hence processing of the Amyloid Precursor Protein, APP. In this study we set out to obtain in vivo confirmation of a link between flotillins and cleavage of APP to release amyloidogenic Aβ peptide, and to generate tools that would allow us to ask whether flotillins are functionally redundant. We used a mouse model for Aβ-dependent cerebral amyloidosis, APPPS1 mice, combined with deletion of either flotillin 1 singly, or flotillin 1 and flotillin 2 together. There was a small but significant reduction in Aβ levels, and the abundance of congo-red stained plaques, in brains of 12 week old mice lacking flotillin 1. A similar reduction in Aβ levels was observed in the flotillin 1-/-, flotillin 2-/- double knockouts. We did not observe large effects on the clustering or endocytosis of APP in flotillin 1-/- mouse embryonic fibroblasts. We conclude that flotillins are likely to play some role in APP trafficking or processing, but the relevant cellular mechanisms require more investigation. The availability of flotillin 1-/-, flotillin 2-/- mice, which have no overt phenotypes, will facilitate research into flotillin function in vivo. 相似文献
110.
Georgiadou M Notas G Xidakis C Drygiannakis I Sfakianaki O Klironomos S Valatas V Kouroumalis E 《Journal of receptor and signal transduction research》2011,31(4):291-298
Introduction: Somatostatin is a mediator of immune functions and has been used as an antineoplastic agent in animal models and human neoplasias. We have demonstrated that Octreotide inhibits only LPS induced secretion of proinflammatory cytokines including TNFa by Kupffer cells (KC). We, therefore, tested the hypothesis that somatostatin modulates the expression of tumor necrosis factor alpha (TNF?) receptors and apoptosis of KC. Methods: Rat KC were isolated by centrifugal elutriation. TNFR1 and TNFR2 expression was studied by RT-PCR, quantitative PCR, Western Blot and immunofluorescence before and after Octreotide pre-incubation. Apoptosis was assessed by quantitative measurement of cytoplasmic histone-associated DNA fragments. TNFa mRNA expression was assessed by semiquantitative PCR and TNFa was measured in cell supernatants by ELISA. Results: TNFR1 and TNFR2 mRNA are constitutively expressed in KC. Octreotide incubation increased both receptors expression with a peak at 6?h and return to basal levels at 24?h. TNFR1 was mostly influenced. However, only increase in TNFR2 protein was identified, whereas a band at 90 kD was present instead of a band at 55 kD as expected for TNFR1. TNF? mRNA expression was inhibited by Octreotide and a significant inhibition was observed at 48?h. TNF had no effect on KC apoptosis, whereas Octreotide significantly increased their apoptosis, and this effect was not influenced by co-incubation with TNFa. Conclusion: TNFR1 and TNFR2 are constitutively expressed in KC and their expression is strongly increased by somatostatin. Moreover, somatostatin increases KC apoptosis. These findings may in part explain the antineoplasmatic effect of somatostatin. 相似文献