Browsing by Author "Contreras, Pamela"

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    AT2 Receptor Mediated Activation of the Tyrosine Phosphatase PTP1B Blocks Caveolin-1 Enhanced Migration, Invasion and Metastasis of Cancer Cells
    (2019) Martínez-Meza, Samuel; Díaz, Jorge; Sandoval-Bórquez, Alejandra; Valenzuela-Valderrama, Manuel; Díaz-Valdivia, Natalia; Rojas-Celis, Victoria; Contreras, Pamela; Huilcaman, Ricardo; Ocaranza Jeraldino, María Paz; Chiong, Mario; Leyton, Lissette; Lavandero, Sergio; Quest, Andrew F. G.
    The renin-angiotensin receptor AT2R controls systemic blood pressure and is also suggested to modulate metastasis of cancer cells. However, in the latter case, the mechanisms involved downstream of AT2R remain to be defined. We recently described a novel Caveolin-1(CAV1)/Ras-related protein 5A (Rab5)/Ras-related C3 botulinum toxin substrate 1 (Rac1) signaling axis that promotes metastasis in melanoma, colon, and breast cancer cells. Here, we evaluated whether the anti-metastatic effect of AT2R is connected to inhibition of this pathway. We found that murine melanoma B16F10 cells expressed AT2R, while MDA-MB-231 human breast cancer cells did not. AT2R activation blocked migration, transendothelial migration, and metastasis of B16F10(cav-1) cells, and this effect was lost when AT2R was silenced. Additionally, AT2R activation reduced transendothelial migration of A375 human melanoma cells expressing CAV1. The relevance of AT2R was further underscored by showing that overexpression of the AT2R in MDA-MB-231 cells decreased migration. Moreover, AT2R activation increased non-receptor protein tyrosine phosphatase 1B (PTP1B) activity, decreased phosphorylation of CAV1 on tyrosine-14 as well as Rab5/Rac1 activity, and reduced lung metastasis of B16F10(cav-1) cells in C57BL/6 mice. Thus, AT2R activation reduces migration, invasion, and metastasis of cancer cells by PTP1B-mediated CAV1 dephosphorylation and inhibition of the CAV1/Rab5/Rac-1 pathway. In doing so, these observations open up interesting, novel therapeutic opportunities to treat metastatic cancer disease.
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    Protein kinase B (AKT) upregulation and Thy‑1‑αvβ3 integrin‑induced phosphorylation of Connexin43 by activated AKT in astrogliosis
    (2023) Pérez-Núñez, Ramón; Chamorro, Alejandro; González, María F.; Contreras, Pamela; Artigas Barrenechea, Rocío; Corvalán R., Alejandro; van Zundert, Brigitte; Reyes, Christopher; Moya, Pablo R.; Avalos, Ana M.; Schneider, Pascal; Quest, Andrew F. G.; Leyton, Lisette
    Background: In response to brain injury or inflammation, astrocytes undergo hypertrophy, proliferate, and migrate to the damaged zone. These changes, collectively known as "astrogliosis", initially protect the brain; however, astrogliosis can also cause neuronal dysfunction. Additionally, these astrocytes undergo intracellular changes involving alterations in the expression and localization of many proteins, including αvβ3 integrin. Our previous reports indicate that Thy-1, a neuronal glycoprotein, binds to this integrin inducing Connexin43 (Cx43) hemichannel (HC) opening, ATP release, and astrocyte migration. Despite such insight, important links and molecular events leading to astrogliosis remain to be defined. Methods: Using bioinformatics approaches, we analyzed different Gene Expression Omnibus datasets to identify changes occurring in reactive astrocytes as compared to astrocytes from the normal mouse brain. In silico analysis was validated by both qRT-PCR and immunoblotting using reactive astrocyte cultures from the normal rat brain treated with TNF and from the brain of a hSOD1G93A transgenic mouse model. We evaluated the phosphorylation of Cx43 serine residue 373 (S373) by AKT and ATP release as a functional assay for HC opening. In vivo experiments were also performed with an AKT inhibitor (AKTi). Results: The bioinformatics analysis revealed that genes of the PI3K/AKT signaling pathway were among the most significantly altered in reactive astrocytes. mRNA and protein levels of PI3K, AKT, as well as Cx43, were elevated in reactive astrocytes from normal rats and from hSOD1G93A transgenic mice, as compared to controls. In vitro, reactive astrocytes stimulated with Thy-1 responded by activating AKT, which phosphorylated S373Cx43. Increased pS373Cx43 augmented the release of ATP to the extracellular medium and AKTi inhibited these Thy-1-induced responses. Furthermore, in an in vivo model of inflammation (brain damage), AKTi decreased the levels of astrocyte reactivity markers and S373Cx43 phosphorylation. Conclusions: Here, we identify changes in the PI3K/AKT molecular signaling network and show how they participate in astrogliosis by regulating the HC protein Cx43. Moreover, because HC opening and ATP release are important in astrocyte reactivity, the phosphorylation of Cx43 by AKT and the associated increase in ATP release identify a potential therapeutic window of opportunity to limit the adverse effects of astrogliosis.