DSpace Repository :: Browsing by Author "Retamal, Mauricio A."

Browsing by Author "Retamal, Mauricio A."

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Activation of Intra-nodose Ganglion P2X7 Receptors Elicit Increases in Neuronal Activity
(2023) Alcayaga, Julio; Vera, Jorge; Reyna, Mauricio; Covarrubias, Alejandra; Coddou, Claudio; Díaz, Esteban; Del Rio, Rodrigo; Retamal, Mauricio A.
Vagus nerve innervates several organs including the heart, stomach, and pancreas among others. Somas of sensory neurons that project through the vagal nerve are located in the nodose ganglion. The presence of purinergic receptors has been reported in neurons and satellite glial cells in several sensory ganglia. In the nodose ganglion, calcium depletion-induced increases in neuron activity can be partly reversed by P2X7 blockers applied directly into the ganglion. The later suggest a possible role of P2X7 receptors in the modulation of neuronal activity within this sensory ganglion. We aimed to characterize the response to P2X7 activation in nodose ganglion neurons under physiological conditions. Using an ex vivo preparation for electrophysiological recordings of the neural discharges of nodose ganglion neurons, we found that treatments with ATP induce transient neuronal activity increases. Also, we found a concentration-dependent increase in neural activity in response to Bz-ATP (ED50 = 0.62 mM, a selective P2X7 receptor agonist), with a clear desensitization pattern when applied every ~ 30 s. Electrophysiological recordings from isolated nodose ganglion neurons reveal no differences in the responses to Bz-ATP and ATP. Finally, we showed that the P2X7 receptor was expressed in the rat nodose ganglion, both in neurons and satellite glial cells. Additionally, a P2X7 receptor negative allosteric modulator decreased the duration of Bz-ATP-induced maximal responses without affecting their amplitude. Our results show the presence of functional P2X7 receptors under physiological conditions within the nodose ganglion of the rat, and suggest that ATP modulation of nodose ganglion activity may be in part mediated by the activation of P2X7 receptors.
Acute activation of hemichannels by ethanol leads to Ca2+ -dependent gliotransmitter release in astrocytes
(2024) Gómez, Gonzalo I.; García-Rodríguez, Claudia; Marillán, Jesús E.; Vergara, Sergio A.; Alvear, Tanhia F.; Farias-Pasten, Arantza; Sáez, Juan C.; Retamal, Mauricio A.; Rovegno, Maximiliano; Ortiz, Fernando C.; Orellana, Juan A.
Multiple studies have demonstrated that acute ethanol consumption alters brain function and cognition. Nevertheless, the mechanisms underlying this phenomenon remain poorly understood. Astrocyte-mediated gliotransmission is crucial for hippocampal plasticity, and recently, the opening of hemichannels has been found to play a relevant role in this process. Hemichannels are plasma membrane channels composed of six connexins or seven pannexins, respectively, that oligomerize around a central pore. They serve as ionic and molecular exchange conduits between the cytoplasm and extracellular milieu, allowing the release of various paracrine substances, such as ATP, D-serine, and glutamate, and the entry of ions and other substances, such as Ca2+ and glucose. The persistent and exacerbated opening of hemichannels has been associated with the pathogenesis and progression of several brain diseases for at least three mechanisms.
Assessing Pulmonary Epithelial Damage in Hantavirus Cardiopulmonary Syndrome: Challenging the Predominant Role of Vascular Endothelium through sRAGE as a Potential Biomarker
(2023) Meza, Gabriela; López, René; Vial Cox, María Cecilia; Cortes, Lina; Retamal, Mauricio A.; Delgado, Iris
Hantavirus cardiopulmonary syndrome (HCPS) is a severe respiratory illness primarily associated with microvascular endothelial changes, particularly in the lungs. However, the role of the pulmonary epithelium in HCPS pathogenesis remains unclear. This study explores the potential of soluble Receptors for Advanced Glycation End-products (sRAGE) as a biomarker for assessing pulmonary epithelial damage in severe HCPS, challenging the prevailing view that endothelial dysfunction is the sole driver of this syndrome. We conducted a cross-sectional study on critically ill HCPS patients, categorizing them into mild HCPS, severe HCPS, and negative control groups. Plasma sRAGE levels were measured, revealing significant differences between the severe HCPS group and controls. Our findings suggest that sRAGE holds promise as an indicator of pulmonary epithelial injury in HCPS and may aid in tracking disease progression and guiding therapeutic strategies. This study brings clarity on the importance of investigating the pulmonary epithelium's role in HCPS pathogenesis, offering potential avenues for enhanced diagnostic precision and support in this critical public health concern.
Cell membrane permeabilization via connexin hemichannels in living and dying cells
(2010) Sáez, Juan C.; Schalper, Kurt A.; Retamal, Mauricio A.; Orellana, Juan A.; Shoji, Kenji F.; Bennett, Michael V.L
Vertebrate cells that express connexins likely express connexin hemichannels (Cx HCs) at their surface. In diverse cell types, surface Cx HCs can open to serve as a diffusional exchange pathway for ions and small molecules across the cell membrane. Most cells, if not all, also express pannexins that form hemichannels and increase the cell membrane permeability but are not addressed in this review. To date, most characterizations of Cx HCs have utilized cultured cells under resting conditions have and revealed low open probability and unitary conductance close to double that of the corresponding gap junction channels. In addition, the cell membrane permeability through Cx HCs can be markedly affected within seconds to minutes by various changes in the intra and/or extracellular microenvironment (i.e., pH, pCa, redox state, transmembrane voltage and intracellular regulatory proteins) that affect levels, open probability and/or (single channel) permeability of Cx HC. Net increase or decrease in membrane permeability could result from the simultaneous interaction of different mechanisms that affect hemichannels. The permeability of Cx HCs is controlled by complex signaling cascades showing connexin, cell and cell stage dependency. Changes in membrane permeability via hemichannels can have positive consequences in some cells (mainly in healthy cells), whereas in others (mainly in cells affected by acquired and/or genetic diseases) hemichannel activation can be detrimental.
Conexina 46 es un regulador transcripcional en células HeLa
(Universidad del Desarrollo. Facultad de Medicina, 2022) Fernández-Olivares, Ainoa; Retamal, Mauricio A.; Acuña, Rodrigo
Cancer is the second leading cause of death in the world and was responsible for around 10 million deaths in 2020. Metastatic colonies are of greatest interest to clinical oncologists as they are responsible for approximately 90% of cancer-associated deaths. Therefore, understanding the molecular mechanisms underlying tumor progression, local invasion, and metastasis formation represents one of the great challenges in cancer research. Connexins are membrane proteins that form ion channels that participate in cell communication, which is essential for tissues to maintain the ability to grow in a controlled manner and respond to their environment. However, this process is frequently impaired in cancer cells. This is why during the last 50 years, the role of connexins and cell communication has been highlighted in the area of cancer and much of the research effort has been directed to understanding their dysfunction in malignant neoplasms in humans. Within the Connexin family, Cx46 has been proposed as a biomarker of malignancy in breast cancer, it has also been directly correlated with the presence of cancer stem cells in glioblastoma. In addition, it has been positively correlated with invasion, self-renewal capacity and metastasis independently of GJC formation. Each of these processes of transformation towards a malignant phenotype requires the reprogramming of gene transcription. Based on the data collected in the literature, the following hypothesis is proposed, cx46 produces phenotypic changes in HeLa cells through gene regulation networks. For this, a stable cell line of HeLa cells over-expressing human Cx46 was generated. The following objectives were studied in it, 1: To evaluate phenotypic characteristics associated with the expression of Cx46 in HeLa cells by means of proliferation, self-renewal and invasion assays. 2: Determination of the subcellular localization of Cx46 by purification of nuclear proteins and confocal microscopy. 3: Determine the nuclear role of the Cx46 protein through ChIP-seq and RNA-seq studies. The results obtained show that HeLa cells transfected with human Cx46 undergo phenotypic changes such as an increase in their capacity for self-renewal proliferation, migration and invasion. In addition the nuclear presence of the protein, which is found interacting with DNA, could be evidenced. In this context, 77 differentially expressed genes could be found, of which 26 have a Cx46 peak in their promoter region. The expression network analysis determined that those genes that did not have Cx46 in their promoter region were being activated by transcription factors with which Cx46 was interacting. The data obtained in this thesis allow us to propose a new role for Cx46 as a regulator of gene transcription in the HeLa cell line. Based on the number of tumors that overexpress Cx46 with respect to their surrounding healthy tissue it is plausible to propose Cx46 as a tumor biomarker and therefore as a possible therapeutic target, in accordance with the literature.
Connexin-46 Contained in Extracellular Vesicles Enhance Malignancy Features in Breast Cancer Cells
(2020) Acuña, Rodrigo; Varas-Godoy, Manuel; Berthoud, Viviana M.; Alfaro, Iván; Retamal, Mauricio A.
Under normal conditions, almost all cell types communicate with their neighboring cells through gap junction channels (GJC), facilitating cellular and tissue homeostasis. A GJC is formed by the interaction of two hemichannels; each one of these hemichannels in turn is formed by six subunits of transmembrane proteins called connexins (Cx). For many years, it was believed that the loss of GJC-mediated intercellular communication was a hallmark in cancer development. However, nowadays this paradigm is changing. The connexin 46 (Cx46), which is almost exclusively expressed in the eye lens, is upregulated in human breast cancer, and is correlated with tumor growth in a Xenograft mouse model. On the other hand, extracellular vesicles (EVs) have an important role in long-distance communication under physiological conditions. In the last decade, EVs also have been recognized as key players in cancer aggressiveness. The aim of this work was to explore the involvement of Cx46 in EV-mediated intercellular communication. Here, we demonstrated for the first time, that Cx46 is contained in EVs released from breast cancer cells overexpressing Cx46 (EVs-Cx46). This EV-Cx46 facilitates the interaction between EVs and the recipient cell resulting in an increase in their migration and invasion properties. Our results suggest that EV-Cx46 could be a marker of cancer malignancy and open the possibility to consider Cx46 as a new therapeutic target in cancer treatment.
Connexin43 Hemichannels in Satellite Glial Cells, Can They Influence Sensory Neuron Activity?
(2017) Retamal, Mauricio A.; Riquelme, Manuel A; Stehberg, Jimmy; Alcayaga, Julio
In this review article, we summarize the current insight on the role of Connexin- and Pannexin-based channels as modulators of sensory neurons. The somas of sensory neurons are located in sensory ganglia (i.e., trigeminal and nodose ganglia). It is well known that within sensory ganglia, sensory neurons do not form neither electrical nor chemical synapses. One of the reasons for this is that each soma is surrounded by glial cells, known as satellite glial cells (SGCs). Recent evidence shows that connexin43 (Cx43) hemichannels and probably pannexons located at SGCs have an important role in paracrine communication between glial cells and sensory neurons. This communication may be exerted via the release of bioactive molecules from SGCs and their subsequent action on receptors located at the soma of sensory neurons. The glio-neuronal communication seems to be relevant for the establishment of chronic pain, hyperalgesia and pathologies associated with tissue inflammation. Based on the current literature, it is possible to propose that Cx43 hemichannels expressed in SGCs could be a novel pharmacological target for treating chronic pain, which need to be directly evaluated in future studies.
Connexinopathies: a structural and functional glimpse
(2016) García, Isaac E; Prado, Pavel; Pupo, Amaury; Jara, Oscar; Rojas-Gómez, Diana; Mujica, Paula; Flores-Muñoz, Carolina; González-Casanova, Jorge; Soto-Riveros, Carolina; Pinto, Bernardo I; Retamal, Mauricio A.; González, Carlos; Martínez, Agustín D
Mutations in human connexin (Cx) genes have been related to diseases, which we termed connexinopathies. Such hereditary disorders include nonsyndromic or syndromic deafness (Cx26, Cx30), Charcot Marie Tooth disease (Cx32), occulodentodigital dysplasia and cardiopathies (Cx43), and cataracts (Cx46, Cx50). Despite the clinical phenotypes of connexinopathies have been well documented, their pathogenic molecular determinants remain elusive. The purpose of this work is to identify common/uncommon patterns in channels function among Cx mutations linked to human diseases. To this end, we compiled and discussed the effect of mutations associated to Cx26, Cx32, Cx43, and Cx50 over gap junction channels and hemichannels, highlighting the function of the structural channel domains in which mutations are located and their possible role affecting oligomerization, gating and perm/ selectivity processes.
Connexins in Cancer, the Possible Role of Connexin46 as a Cancer Stem Cell-Determining Protein
(2023) León, Isidora; Salgado, María; Novoa, María; Retamal, Mauricio A.
Cancer is a widespread and incurable disease caused by genetic mutations, leading to uncontrolled cell proliferation and metastasis. Connexins (Cx) are transmembrane proteins that facilitate intercellular communication via hemichannels and gap junction channels. Among them, Cx46 is found mostly in the eye lens. However, in pathological conditions, Cx46 has been observed in various types of cancers, such as glioblastoma, melanoma, and breast cancer. It has been demonstrated that elevated Cx46 levels in breast cancer contribute to cellular resistance to hypoxia, and it is an enhancer of cancer aggressiveness supporting a pro-tumoral role. Accordingly, Cx46 is associated with an increase in cancer stem cell phenotype. These cells display radio- and chemoresistance, high proliferative abilities, self-renewal, and differentiation capacities. This review aims to consolidate the knowledge of the relationship between Cx46, its role in forming hemichannels and gap junctions, and its connection with cancer and cancer stem cells.
Cx40 Levels Regulate Hypoxia-Induced Changes in the Migration, Proliferation, and Formation of Gap Junction Plaques in an Extravillous Trophoblast Cell Model
(2024) Rozas Villanueva, María Fernanda; Orellana Villena, Viviana; Alarcón, Rodrigo; Maripillan, Jaime; Martínez, Agustín; Alfaro, Ivan; Retamal, Mauricio A.
Background: Extravillous trophoblasts (EVTs) form stratified columns at the placenta–uterus interface. In the closest part to fetal structures, EVTs have a proliferative phenotype, whereas in the closest part to maternal structures, they present a migratory phenotype. During the placentation process, Connexin 40 (Cx40) participates in both the proliferation and migration of EVTs, which occurs under hypoxia. However, a possible interaction between hypoxia and Cx40 has not yet been established. Methods: We developed two cellular models, one with “low Cx40” (Jeg-3), which reflected the expression of this protein found in migratory EVTs, and one with “high Cx40” (Jeg-3/hCx40), which reflected the expression of this protein in proliferative cells. We analyzed the migration and proliferation of these cells under normoxic and hypoxic conditions for 24 h. Jeg-3 cells under hypoxia increased their migratory capacity over their proliferative capacity. However, in Jeg-3/hCx40, the opposite effect was induced. On the other hand, hypoxia promoted gap junction (GJ) plaque formation between neighboring Jeg-3 cells. Similarly, the activation of a nitro oxide (NO)/cGMP/PKG-dependent pathway induced an increase in GJ-plaque formation in Jeg-3 cells. Conclusions: The expression patterns of Cx40 play a crucial role in shaping the responses of EVTs to hypoxia, thereby influencing their migratory or proliferative phenotype. Simultaneously, hypoxia triggers an increase in Cx40 gap junction (GJ) plaque formation through a pathway dependent on NO
Editorial: Free Fatty Acids as Signaling Molecules: Role of Free Fatty Acid Receptors and CD36
(2022) Puebla, Carlos; Morselli, Eugenia; Khan, Naim Akhtar; Retamal, Mauricio A.
Ion Channels in Inflammatory Processes: What Is Known and What Is Next?
(2016) Retamal, Mauricio A.; V L Bennett, Michael; Pelegrin, Pablo; Fernandez, Ricardo
Inflammation is the primary response of the immune system to infection or tissue injury. Ion channels in different immune cells are very important for inflammatory processes. In this context, connexin (Cx) and pannexin (Panx) cell-cell channels and unapposed hemichannels and P2 receptors play central roles in the initiation and/or progression of inflammation in different tissues. Thus, cellular responses during inflammation can be initiated and/or enhanced by the opening of Cx and/or Panx hemichannels, which in turn allow the release of ATP and other metabolites to the extracellular media
KI04 an Aminoglycosides-Derived Molecule Acts as an Inhibitor of Human Connexin46 Hemichannels Expressed in HeLa Cells
(2023) Chang, Cheng-Wei; Poudyal, Naveena; Peña, Francisca; Verdugo, Daniel; Stehberg, Jimmy; Retamal, Mauricio A.
Background: Connexins (Cxs) are proteins that help cells to communicate with the extracellular media and with the cytoplasm of neighboring cells. Despite their importance in several human physiological and pathological conditions, their pharmacology is very poor. In the last decade, some molecules derived from aminoglycosides have been developed as inhibitors of Cxs hemichannels. However, these studies have been performed in E. coli, which is a very simple model. Therefore, our main goal is to test whether these molecules have similar effects in mammalian cells. Methods: We transfected HeLa cells with the human Cx46tGFP and characterized the effect of a kanamycin-derived molecule (KI04) on Cx46 hemichannel activity by time-lapse recordings, changes in phosphorylation by Western blot, localization by epifluorescence, and possible binding sites by molecular dynamics (MD). Results: We observed that kanamycin and KI04 were the most potent inhibitors of Cx46 hemichannels among several aminoglycosides, presenting an IC50 close to 10 μM. The inhibitory effect was not associated with changes in Cx46 electrophoretic mobility or its intracellular localization. Interestingly, 5 mM DTT did not reverse KI04 inhibition, but the KI04 effect completely disappeared after washing out KI04 from the recording media. MD analysis revealed two putative binding sites of KI04 in the Cx46 hemichannel. Results: These results demonstrate that KI04 could be used as a Cx46 inhibitor and could help to develop future selective Cx46 inhibitors.
Lack of canonical activities of connexins in highly aggressive human prostate cancer cells
(2024) Asencio, Catalina; Véliz, Loreto; Flores-Faúndes, Emilia; Azócar, Lorena; Echeverría, Carolina E.; Torres-Estay, Verónica; Orellana Villena, Viviana; Ramírez-Santelices, Catalina; Sotomayor, Paula; Cancino, Jorge; Kerr, Bredford; Fernández-Olivares, Ainoa; Retamal, Mauricio A.; Sáez, Juan C.; Godoy, Alejandro S.
Connexins (Cxs) have the ability to form channels that allow the exchange of ions/metabolites between adjacent cells (gap junction channels, GJC) or between the intra- and extra-cellular compartments (hemichannels, HC). Cxs were initially classified as tumor suppressors. However, more recently, it has been shown that Cxs exert anti- and pro-tumorigenic effects depending on the cell and tissue context. In prostate cancer (PCa), the expression and functionality of Cxs remain highly controversial. Here, we analyzed the expression pattern of Cx26, Cx32, Cx37, Cx40, Cx43 and Cx45 in PCa cell lines with increasing levels of tumor aggressiveness (LNCaP < LNCaP-C4-2 < Du-145 < PC-3). In addition, GJ and HC activities were evaluated in the PCa cell lines using dye coupling and dye uptake assays, respectively. Lastly, the cellular localization of Cx26, Cx32, and Cx43 was analyzed in LNCaP and PC-3 cell lines using immunofluorescence analyses. Our results showed a positive association between the mRNA levels of Cx26, Cx37 and Cx45 and the degree of aggressiveness of PCa cells, a negative association in the case of Cx32 and Cx43, and no clear pattern for Cx40. At the protein level, a negative relationship between the expression of Cx26, Cx32 and Cx43 and the degree of aggressiveness of PCa cell lines was observed. No significant differences were observed for the expression of Cx37, Cx40, and Cx45 in PCa cell lines. At the functional level, only LNCaP cells showed moderate GJ activity and LNCaP and LNCaP-C4-2 cells showed HC activity. Immunofluorescence analyses confirmed that the majority of Cx26, Cx32, and Cx43 expression was localized in the cytoplasm of both LNCaP and PC3 cell lines. This data indicated that GJ and HC activities were moderately detected only in the less aggressive PCa cells, which suggest that Cxs expression in highly aggressive PCa cells could be associated to channel-independent roles.
Molecular Interplay Between Non-Coding RNAs and Connexins and Its Possible Role in Cancer
(2025) Perez Moreno, Pablo; Muñoz, Juan; Retamal, Mauricio A.
Non-coding RNAs (ncRNAs) are sequences that do not encode for proteins and play key roles in different cellular processes, including cell proliferation and differentiation. On the other hand, connexins (Cxs) are transmembrane proteins that principally allow intercellular communication. In pathological conditions such as cancer, there is a deregulation in the expression and/or function of ncRNAs and Cxs, which in turn leads to an enhancement in the aggressive phenotype, such as a greater proliferative and invasive capacity. This suggests a plausible interplay between ncRNAs and Cxs. Based on that, this review aims to summarize the current knowledge regarding this relationship and to analyze how it may influence the development of aggressive traits in cancer cells and the clinicopathological features of cancer patients. Finally, we discuss the potential of ncRNAs and Cxs as promising clinical biomarkers for cancer diagnosis, prognosis, and therapeutic targeting. Keywords: cancer; connexins; non-coding RNAs.
Nitric oxide signaling in the retina: What have we learned in two decades?
(2012) Vielma, Alex H.; Retamal, Mauricio A.; Schmachtenberg, Oliver
Two decades after its first detection in the retina, nitric oxide (NO) continues to puzzle visual neuroscientists. While its liberation by photoreceptors remains controversial, recent evidence supports three subtypes of amacrine cells as main sources of NO in the inner retina. NO synthesis was shown to depend on light stimulation, and mounting evidence suggests that NO is a regulator of visual adaptation at different signal processing levels. NO modulates light responses in all retinal neuron classes, and specific ion conductances are activated by NO in rods, cones, bipolar and ganglion cells. Light-dependent gap junction coupling in the inner and outer plexiform layers is also affected by NO. The vast majority of these effects were shown to be mediated by activation of the NO receptor soluble guanylate cyclase and resultant cGMP elevation. This review analyzes the current state of knowledge on physiological NO signaling in the retina.
Potential use of n-3 PUFAs to prevent oxidative stress-derived ototoxicity caused by platinum-based chemotherapy
(2020) Cortés Fuentes, Ignacio A.; Burotto, Mauricio; Retamal, Mauricio A.; Frelinghuysen, Michael; Caglevic, Christian; Gormaz, Juan G.
Platinum-based compounds are widely used for the treatment of different malignancies due to their high effectiveness. Unfortunately, platinum-based treatment may lead to ototoxicity, an often-irreversible side effect without a known effective treatment and prevention plan. Platinum-based compound-related ototoxicity results mainly from the production of toxic levels of reactive oxygen species (ROS) rather than DNA-adduct formation, which has led to test strategies based on direct ROS scavengers to ameliorate hearing loss. However, favorable clinical results have been associated with several complications, including potential interactions with chemotherapy efficacy. To understand the contribution of the different cytotoxic mechanisms of platinum analogues on malignant cells and auditory cells, the particular susceptibility and response of both kinds of cells to molecules that potentially interfere with these mechanisms, is fundamental to develop innovative strategies to prevent ototoxicity without affecting antineoplastic effects. The n-3 long-chain polyunsaturated fatty acids (n-3 PUFAs) have been tried in different clinical settings, including with cancer patients. Nevertheless, their use to decrease cisplatin-induced ototoxicity has not been explored to date. In this hypothesis paper, we address the mechanisms of platinum compounds-derived ototoxicity, focusing on the differences between the effects of these compounds in neoplastic versus auditory cells. We discuss the basis for a strategic use of n-3 PUFAs to potentially protect auditory cells from platinum-derived injury without affecting neoplastic cells and chemotherapy efficacy.
Protocol for the study of Connexin and DNA interactions
(2024) Fernández-Olivares, Ainoa; Retamal, Mauricio A.
Connexins (Cxs) are transmembrane proteins which form hemichannels and gap junction channels at the plasma membrane. These channels allow the exchange of ions and molecules between the intra- and extracellular space and between cytoplasm of adjacent cells, respectively. The channel function of Cx assemblies has been extensively studied; however, “noncanonical” functions have emerged in the last few decades and have capture the attentions of many researchers, including the role of some Cxs as gene modulators or transcription factors. In this chapter, we describe a protocol to study the interaction of Cx46 with DNA in HeLa cells. These methods can facilitate understanding the role of Cxs in physiological processes and pathological mechanisms, including, for example, the contribution of Cx46 in maintaining stemness of glioma cancer stem cells
Regulation of gap junction channels and hemichannels by phosphorylation and redox changes: a revision
(2016) Pogoda, Kiristin; Kameritsch, Petra; Retamal, Mauricio A.; Vega, José L
Post-translational modifications of connexins play an important role in the regulation of gap junction and hemichannel permeability. The prerequisite for the formation of functional gap junction channels is the assembly of connexin proteins into hemichannels and their insertion into the membrane. Hemichannels can affect cellular processes by enabling the passage of signaling molecules between the intracellular and extracellular space. For the intercellular communication hemichannels from one cell have to dock to its counterparts on the opposing membrane of an adjacent cell to allow the transmission of signals via gap junctions from one cell to the other. The controlled opening of hemichannels and gating properties of complete gap junctions can be regulated via post-translational modifications of connexins. Not only channel gating, but also connexin trafficking and assembly into hemichannels can be affected by post-translational changes. Recent investigations have shown that connexins can be modified by phosphorylation/dephosphorylation, redox-related changes including effects of nitric oxide (NO), hydrogen sulfide (H2S) or carbon monoxide (CO), acetylation, methylation or ubiquitination. Most of the connexin isoforms are known to be phosphorylated, e.g. Cx43, one of the most studied connexin at all, has 21 reported phosphorylation sites. In this review, we provide an overview about the current knowledge and relevant research of responsible kinases, connexin phosphorylation sites and reported effects on gap junction and hemichannel regulation. Regarding the effects of oxidants we discuss the role of NO in different cell types and tissues and recent studies about modifications of connexins by CO and H2S.
Rol de la conexina 46 en la estabilización de HIF-1α y la agresividad tumoral de la línea de melanoma SK-MEL-2
(Universidad del Desarrollo. Facultad de Medicina, 2021) Orellana Villena, Viviana Paulina; Retamal, Mauricio A.; Godoy Sánchez, Alejandro
El melanoma es el cáncer de piel más agresivo, cuya tasa de incidencia se encuentra en constante aumento por sobre otros tipos de tumores sólidos. Recientemente la conexina 46 (Cx46) fue descrita en este tipo de cáncer, pero se desconoce su rol en la biología de este. En otros tipos de cáncer como son el cáncer de mama y glioblastoma la Cx46 está asociada con la metástasis y el fenotipo cancer stem cells. Por otro lado, se ha sugerido que la Cx46 actúa como un factor protector frente a la muerte celular inducida por hipoxia. Interesantemente, la hipoxia activa al factor de transcripción HIF-1α el cual está relacionado con el desarrollo de tumores de mal pronóstico. La expresión de Cx46 y HIF-1α ha sido descrita de forma independiente en melanoma, y en este sentido, nuestra investigación tiene como foco evaluar la posible interrelación entre estas dos proteínas, y su posible efecto sobre la progresión tumoral en la línea celular de melanoma SK-MEL-2. Para este estudio, se estableció un modelo celular con reducción de la expresión de Cx46, en el cual se observó un aumento en la estabilización de HIF-1α mediado por la reducción de los niveles de las proteínas PHD2 y pVHL cuyo rol es degradar a HIF-1α en normoxia. Además, se observó que la reducción de Cx46 impacta negativamente sobre las características que favorecen la progresión tumoral, tales como la proliferación, migración y crecimiento de esferas tumorales. Este comportamiento menos agresivo, se explica por el estudio de proteómica el cual demuestra que la vía de señalización RAS se encuentra disminuida, específicamente por una reducción de las proteínas RAS y NFκβ y otras vías representadas por STAT3, CSPG4 e ICAM1. Todas estas relacionadas con favorecer la sobrevida celular mediante la evasión de la apoptosis, y el aumento de la proliferación y la metástasis. Finalmente, se realizó un estudio preliminar en modelo in vivo en ratones NSG, en donde se observó que la reducción de Cx46 inhibe el crecimiento tumoral al igual que en el estudio in vitro. Y junto con esto, pudimos observar a nivel de proteína, que los tumores reducidos en Cx46 presentan la diminución de los niveles de AKT, RAS y NFκβ, confirmando los datos de proteómica. Los datos obtenidos en esta tesis nos permiten proponer un mecanismo en el cual la Cx46 actúa aumentando los niveles de RAS y proteínas rio abajo (AKT y NFκβ). Lo que aumenta las características de proliferación, migración y crecimiento de esferas tumorales, asociadas al fenotipo CSCs. Lo que orientado a la aplicación clínica o traslacional nos permite pensar que la Cx46 es un target interesante de desarrollar con el fin de disminuir la agresividad tumoral del melanoma.

Browsing by Author "Retamal, Mauricio A."

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