107 research outputs found
Opposite effects of cell differentiation and apoptosis on Ap3A/Ap4A ratio in human cell cultures
No full textAbstractThe biological role of diadenosine oligophosphates (DAOP) remains obscure in spite of numerous attempts to solve this enigma. It is known that Ap3A contrary to Ap4A accumulates in human cultured cells treated with interferons (IFNs) alpha or gamma. Since IFNs are considered as antiproliferative regulators, we assumed that different cell status may be associated with varying intracellular levels of DAOP. Promyelocytic human cell line HL60 induced by phorbol ester (TPA) to differentiate to macrophage-like cells in culture exhibits a profound loss of proliferative potential. Here we have shown a 4–5-fold increase in Ap3A concentration in HL60 cells induced by TPA, similar to the effect of IFN, while the Ap4A concentration remained unchanged. On the contrary, in cells undergoing apoptosis induced by VP16, a topoisomerase II inhibitor, the Ap3A concentration considerably decreased, while the Ap4A concentration increased. These findings combined with earlier results suggest an involvement of the Ap3A/Ap4A ratio in signal transduction pathways controlling the cell status
Cellular Mechanisms of FGF-Stimulated Tissue Repair
No full textGrowth factors belonging to the FGF family play important roles in tissue and organ repair after trauma. In this review, I discuss the regulation by FGFs of the aspects of cellular behavior important for reparative processes. In particular, I focus on the FGF-dependent regulation of cell proliferation, cell stemness, de-differentiation, inflammation, angiogenesis, cell senescence, cell death, and the production of proteases. In addition, I review the available literature on the enhancement of FGF expression and secretion in damaged tissues resulting in the increased FGF supply required for tissue repair
Nonclassically secreted regulators of angiogenesis.
No full textMany secreted polypeptide regulators of angiogenesis are devoid of signal peptides. These proteins are released through nonclassical pathways independent of endoplasmic reticulum and Golgi. In most cases, the nonclassical protein export is induced by stress. It usually serves to stimulate repair or inflammation in damaged tissues. We review the secreted signal peptide-less regulators of angiogenesis and discuss the mechanisms and biological significance of their unconventional export
Carbonic anhydrase IX is a marker of hypoxia and correlates with higher Gleason scores and ISUP grading in prostate cancer
No full textBackground: Carbonic anhydrase IX is a member of α-carbonic anhydrases that is preferentially expressed in solid tumors. It enables bicarbonate transport across the plasma membrane, neutralizing intracellular pH and conferring to cancer cells a survival advantage in hypoxic/acidic microenvironments. Overexpression of carbonic anhydrase IX in cancer tissues is regulated by hypoxia inducible factor 1α - mediated transcription and the enzyme is considered a marker of tumor hypoxia and poor outcome. The role of carbonic anhydrase IX in prostate cancer has not been fully clarified and controversy has arisen on whether this enzyme is overexpressed in hypoxic prostate cancer tissues. Methods: We analyzed the expression of carbonic anhydrase IX and hypoxia inducible factor 1α in two prostate cancer cell lines, LNCaP and PC-3, and in 110 cancer biopsies, by western blotting and immunocyto/histochemistry. Results: In LNCaP and PC-3 cells, carbonic anhydrase IX was mostly cytoplasmic/nuclear, with very limited membrane localization. Nuclear staining became stronger under hypoxia. When we analyzed carbonic anhydrase IX expression in human prostate cancer biopsies, we found that protein staining positively correlated with hypoxia inducible factor 1α and with Gleason pattern and score, as well as with the novel grading system proposed by the International Society of Urological Pathology for prostate cancer. Once more, carbonic anhydrase IX was mainly cytoplasmic in low grade carcinomas, whereas in high grade tumors was strongly expressed in the nucleus of the neoplastic cell. An association between carbonic anhydrase IX expression level and the main clinic-pathological features involved in prostate cancer aggressiveness was identified. Conclusions: There was a statistically significant association between carbonic anhydrase IX and hypoxia inducible factor 1α in prostate cancer tissues, that identifies the enzyme as a reliable marker of tumor hypoxia. In addition, carbonic anhydrase IX expression positively correlated with prostate cancer grading and staging, and with outcome, suggesting that the protein may be an independent prognosticator for the disease. The nuclear translocation of the enzyme in hypoxic cancer cells may epitomize a biological switch of the tumor towards a less favorable phenotype
Mechanisms utilized by growth factors and cytokines in angiogenesis: role of thrombin in the cross-talk between the FGF1 and Notch signaling pathways
No full textTese de Doutoramento na área de Ciências da Saúde (Ramo de Ciências Biológicas e Biomédicas)Angiogenesis, the development of new blood vessels from the existing vasculature, is controlled by different signaling pathways, where diverse cytokines, chemokines and growth factors play important roles. Angiogenesis and blood coagulation are key events of vascular biology. Serine protease, thrombin, which plays a central role in blood coagulation cascade through its ability to cleave fibrinogen and produce fibrin, is also known to be involved in inflammation, wound
healing and tissue remodeling, growth factor activation, embryogenesis, and both normal and aberrant cell growth control. The effects of thrombin are associated with the induction of expression of several growth factors including fibroblast growth factor (FGF) 2, platelet derived growth factor, insulin-like growth factor 1, and vascular endothelial growth factor. In this work, the regulation of expression and release of FGF1, a potent pro-angiogenic factor was investigated in the context of thrombin activity.
We found that thrombin has the ability to induce FGF1 transcription and redistribution of FGF1 to the inner leaflet of the plasma membrane, resulting in the non-classical (ER/Golgi independent) export of this growth factor with fast kinetics.
FGF1 signaling underlies thrombin mitogenic activity since thrombin does not promote cell proliferation in cells expressing a dominant negative form of FGF receptor 1. In an effort to further define the mechanisms underlying the observed effects of thrombin, we found that both release and expression of FGF1 stimulated by thrombin are dependent on protease activated receptor 1 (PAR1). Additionally thrombin is capable to cleave full-length transmembrane Notch ligand Jagged1 in its extracellular domain and to produce a soluble form of Jagged1 which decreases Notch signaling and induces FGF1 expression and export. Interestingly, we also demonstrated that the long term thrombin treatment can induce FGF1 release from PAR1 knockout cells, most probably as a result of accumulation of soluble Jagged1.
In conclusion, these studies have identified a novel cross-talk bridging thrombin, FGF1 and Notch signaling pathways, which all play important roles in vascular developing and remodeling.O processo de angiogénese consiste no desenvolvimento de novos vasos
sanguíneos a partir de vasos pré-existentes, sendo controlado por diferentes sistemas de sinalização, onde diversas citoquinas, e factores de crescimento têm um papel crucial. Angiogénese e coagulação sanguínea desempenham importantes papéis em biologia vascular. A serina protease trombina, a qual tem um papel principal na cascata de coagulação pela quebra enzimática do fibrinogênio e produção de fibrina, encontra-se também envolvida na inflamação, cicatrização, activação de factores de
crescimento, embriogénese e crescimento celular, quer em condições normais quer em condições de crescimento aberrante. A actividade celular da trombina é devida, em grande parte, à indução de expressão de vários factores de crescimento, tais como o “fibroblast growth factor” (FGF) 2, o “platelet derived growth factor”, “insulin-like
growth factor”, e o “vascular endothelial growht factor”. Neste trabalho, a regulação da expressão e libertação do FGF1, um conhecido e potente factor pró-angiogénico, foi investigada no contexto da actividade da trombina.
Os resultados obtidos demonstraram que a trombina tem a capacidade de induzir a transcrição e redistribuição do FGF1 na porção celular interna da membrana plasmática, resultando na exportação não clássica (por via independente do retículo endoplásmico\Golgi) deste factor pró-angiogénico com rápida cinética. A libertação do FGF1 evidencia a actividade mitótica da trombina em fibroblastos de ratinho, uma
vez que a trombina não é capaz de induzir proliferação celular em células que expressam um mutante do receptor do FGF1 com efeito negativo dominante. Na tentativa de tentar esclarecer este mecanismo, demonstramos que a activação da transcrição e o aparecimento do FGF1 no compartimento extra celular era dependente
do receptor da trombina, denominado por “protease activated receptor 1” (PAR1).
Adicionalmente, a serina protease trombina foi capaz de clivar o Jagged1 na sua porção extra celular levando à produção uma forma solúvel deste ligando, a qual é capaz de inibir a activação do mecanismo de sinalização do Notch1 e ao mesmo tempo levar à activação da transcrição e exportação do FGF1. Curiosamente, demonstramos também que a incubação com trombina, por longos períodos de tempo, em células que não expressam PAR1 conduz a libertação do FGF1, mais
provavelmente devido à acumulação da forma solúvel do Jagged1.
Em conclusão, estes estudos permitiram no contexto de remodelação vascular
identificar uma nova via que interliga as vias de sinalização da trombina, FGF1 e Notch
Regulation of DNA Synthesis Investigated in Heterokaryons of Dividing and Nondividing Cells
No full textDifferential effects of phorbol ester on signaling and gene expression in human leukemia cells
No full textNotch3 is activated by chronic hypoxia and contributes to the progression of human prostate cancer
No full textProstate cancer (PC) is still the second cause of cancer-related death among men. Although patients with metastatic presentation have an ominous outcome, the vast majority of PCs are diagnosed at an early stage. Nonetheless, even among patients with clinically localized disease the outcome may vary considerably. Other than androgen sensitivity, little is known about which other signaling pathways are deranged in aggressive, localized cancers. The elucidation of such pathways may help to develop innovative therapies aimed at specific molecular targets. We report that in a hormone-sensitive PC cell line, LNCaP, Notch3 was activated by hypoxia and sustained cell proliferation and colony formation in soft agar. Hypoxia also modulated cellular cholesterol content and the number and size of lipid rafts, causing a coalescence of small rafts into bigger clusters; under this experimental condition, Notch3 migrated from the non-raft into the raft compartment where it colocalized with the γ-secretase complex. We also looked at human PC biopsies and found that expression of Notch3 positively correlated with Gleason score and with expression of carbonic anhydrase IX, a marker of hypoxia. In conclusion, hypoxia triggers the activation of Notch3, which, in turn, sustains proliferation of PC cells. Notch3 pathway represents a promising target for adjuvant therapy in patients with PC. Copyright © 2013 UICC
Protein-phospholipid interactions in nonclassical protein secretion: problem and methods of study.
No full textExtracellular proteins devoid of signal peptides use nonclassical secretion mechanisms for their export. These mechanisms are independent of the endoplasmic reticulum and Golgi. Some nonclassically released proteins, particularly fibroblast growth factors (FGF) 1 and 2, are exported as a result of their direct translocation through the cell membrane. This process requires specific interactions of released proteins with membrane phospholipids. In this review written by a cell biologist, a structural biologist and two membrane engineers, we discuss the following subjects: (i) Phenomenon of nonclassical protein release and its biological significance; (ii) Composition of the FGF1 multiprotein release complex (MRC); (iii) The relationship between FGF1 export and acidic phospholipid externalization; (iv) Interactions of FGF1 MRC components with acidic phospholipids; (v) Methods to study the transmembrane translocation of proteins; (vi) Membrane models to study nonclassical protein release
Sustained Inhibition of Proliferative Response After Transient FGF Stimulation Is Mediated by Interleukin 1 Signaling.
No full textTransient FGF stimulation of various cell types results in FGF memory--a sustained blockage of efficient proliferative response to FGF and other growth factors. FGF memory establishment requires HDAC activity, indicating its epigenetic character. FGF treatment stimulates proinflammatory NFκB signaling, which is also critical for FGF memory formation. The search for FGF-induced mediators of FGF memory revealed that FGF stimulates HDAC-dependent expression of the inflammatory cytokine IL1α. Similarly to FGF, transient cell treatment with recombinant IL1α inhibits the proliferative response to further FGF and EGF stimulation, but does not prevent FGF receptor-mediated signaling. Interestingly, like cells pretreated with FGF1, cells pretreated with IL1α exhibit enhanced restructuring of actin cytoskeleton and increased migration in response to FGF stimulation. IRAP, a specific inhibitor of IL 1 receptor, and a neutralizing anti-IL1α antibody prevent the formation of FGF memory and rescue an efficient proliferative response to FGF restimulation. A similar effect results following treatment with the anti-inflammatory agents aspirin and dexamethasone. Thus, FGF memory is mediated by proinflammatory IL1 signaling. It may play a role in the limitation of proliferative response to tissue damage and prevention of wound-induced hyperplasia
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