Browsing by Author "Koning, Tania"

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    Coagulation Factor Xa Promotes Solid Tumor Growth, Experimental Metastasis and Endothelial Cell Activation.
    (2019) Arce, Maximiliano; Pinto, Mauricio P.; Galleguillos, Macarena; Muñoz, Catalina; Lange, Soledad; Ramirez, Carolina; Erices, Rafaela; Gonzalez, Pamela; Velasquez, Ethel; Tempio, Fabián; Lopez, Mercedes N.; Salazar-Onfray, Flavio; Cautivo, Kelly; Kalergis, Alexis M.; Cruz, Sebastián; Lladser, Álvaro; Lobos-González, Lorena; Valenzuela, Guillermo; Olivares, Nixa; Sáez, Claudia; Koning, Tania; Sánchez, Fabiola A.; Fuenzalida, Patricia; Godoy, Alejandro; Contreras Orellana, Pamela; Leyton, Lisette; Lugano, Roberta; Dimberg, Anna; Quest, Andrew F.G.; Owen, Gareth I.
    Hypercoagulable state is linked to cancer progression; however, the precise role of the coagulation cascade is poorly described. Herein, we examined the contribution of a hypercoagulative state through the administration of intravenous Coagulation Factor Xa (FXa), on the growth of solid human tumors and the experimental metastasis of the B16F10 melanoma in mouse models. FXa increased solid tumor volume and lung, liver, kidney and lymph node metastasis of tail-vein injected B16F10 cells. Concentrating on the metastasis model, upon coadministration of the anticoagulant Dalteparin, lung metastasis was significantly reduced, and no metastasis was observed in other organs. FXa did not directly alter proliferation, migration or invasion of cancer cells in vitro. Alternatively, FXa upon endothelial cells promoted cytoskeleton contraction, disrupted membrane VE-Cadherin pattern, heightened endothelial-hyperpermeability, increased inflammatory adhesion molecules and enhanced B16F10 adhesion under flow conditions. Microarray analysis of endothelial cells treated with FXa demonstrated elevated expression of inflammatory transcripts. Accordingly, FXa treatment increased immune cell infiltration in mouse lungs, an effect reduced by dalteparin. Taken together, our results suggest that FXa increases B16F10 metastasis via endothelial cell activation and enhanced cancer cell-endothelium adhesion advocating that the coagulation system is not merely a bystander in the process of cancer metastasis.
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    TNF-α-activated eNOS signaling increases leukocyte adhesion through the S- nitrosylation pathway
    (2021) Aguilar, Gaynor; Cordova, Francisco; Koning, Tania; Sarmiento, Jose; Boric, Mauricio P.; Birukov, Konstantin; Cancino, Jorge; Varas-Godoy, Manuel; Soza, Andrea; Alves, Natascha G.; Mujica, Patricio E.; Duran, Walter N.; Ehrenfeld, Pamela; Sanchez, Fabiola A.
    Nitric oxide ( NO) is a key factor in inflammation. Endothelial nitric oxide synthase (eNOS), whose activity increases after stimulation with proinflammatory cytokines, produces NO in endothelium. NO activates two pathways: 1) soluble guanylate cyclase-protein kinase G and 2) S-nitrosylation (NO-induced modification of free-thiol cysteines in proteins). S-nitrosylation affects phosphorylation, localization, and protein interactions. NO is classically described as a negative regulator of leukocyte adhesion to endothelial cells. However, agonists activating NO production induce a fast leukocyte adhesion, which suggests that NO might positively regulate leukocyte adhesion. We tested the hypothesis that eNOS-induced NO promotes leukocyte adhesion through the S-nitrosylation pathway. We stimulated leukocyte adhesion to endothelium in vitro and in vivo using tumor necrosis factor-alpha (TNF-alpha) as proinflammatory agonist. ICAM-1 changes were evaluated by immunofluorescence, subcellular fractionation, immunoprecipitation, and fluorescence recovery after photobleaching (FRAP). Protein kinase C sigma (PKC sigma) activity and S-nitrosylation were evaluated by Western blot analysis and biotin switch method, respectively. TNF-alpha, at short times of stimulation, activated the eNOS S-nitrosylation pathway and caused leukocyte adhesion to endothelial cells in vivo and in vitro. TNF-alpha-induced NO led to changes in ICAM-1 at the cell surface, which are characteristic of clustering. TNF-alpha-induced NO also produced S-nitrosylation and phosphorylation of PKCf, association of PKCf with ICAM-1, and ICAM-1 phosphorylation. The inhibition of PKCf blocked leukocyte adhesion induced by TNF-alpha. Mass spectrometry analysis of purified PKCf identified cysteine 503 as the only S-nitrosylated residue in the kinase domain of the protein. Our results reveal a new eNOS S-nitrosylation-dependent mechanism that induces leukocyte adhesion and suggests that S-nitrosylation of PKCf may be an important regulatory step in early leukocyte adhesion in inflammation.