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Obesity increases the risk and mortality of breast cancer through dysregulated secretion of proinflammatory cytokines and tumor adipokines that induce an inflammatory breast microenvironment. Resistin is an adipokine secreted by adipocytes, immune cells, and predominantly macrophages, which contributes to cancer progression, but its molecular mechanism in cancer is not completely described. In this study, we analyzed the relationship of resistin on breast cancer prognosis and tumor progression and the effect in vitro of resistin on p38 and ERK1/2 activation in breast cancer cell lines. By bioinformatic analysis, we found that resistin is overexpressed in the basal subtype triple-negative breast cancer and is related to poor prognosis. In addition, we demonstrated a positive correlation between RETN and MAPK3 expression in basal triple-negative breast cancer. Importantly, we found amplifications of the RETN gene in at least 20 % of metastatic samples from patients with breast cancer. Most samples with RETN amplifications metastasized to bone and showed high expression of IL-8 (CXCL8) and IL-6 (IL6). Finally, resistin could be considered a prognostic marker for basal triple-negative breast cancer, and we also proposed the possibility that resistin-induced cell migration involves the activation of MAPK in breast cancer cells.
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Experimental autoimmune encephalomyelitis is a demyelinating disease that causes paralysis in laboratory rats. This condition lacks treatment that reverses damage to the myelin sheaths of neuronal cells. Therefore, in this study, treatment with EPO as a neuroprotective effect was established to evaluate the ERK 1/2 signaling pathway and its participation in the EAE model. EPO was administered in 5000 U/Kg Sprague Dawley rats. U0126 was used as an inhibitor of the ERK 1/2 pathway to demonstrate the possible activation of this pathway in the model. Spinal cord and optic nerve tissues were evaluated using staining techniques such as H&E and the Luxol Fast Blue myelin-specific technique, as well as immunohistochemistry of the ERK 1/2 protein. The EPO-treated groups showed a decrease in cellular sampling in the spinal cord tissues but mainly in the optic nerve, as well as an increase in the expression of the ERK 1/2 protein in both tissues. The findings of this study suggest that EPO treatment reduces cellular death in EAE-induced rats by regulating the ERK pathway.
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Encefalomielitis Autoinmune Experimental , Eritropoyetina , Sistema de Señalización de MAP Quinasas , Fármacos Neuroprotectores , Nervio Óptico , Ratas Sprague-Dawley , Médula Espinal , Animales , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/metabolismo , Encefalomielitis Autoinmune Experimental/patología , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Eritropoyetina/farmacología , Nervio Óptico/efectos de los fármacos , Nervio Óptico/patología , Nervio Óptico/metabolismo , Ratas , Médula Espinal/metabolismo , Médula Espinal/efectos de los fármacos , Médula Espinal/patología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Femenino , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/metabolismoRESUMEN
Ischaemia-reperfusion (IR)-associated acute kidney injury (AKI) is a severe clinical condition that lacks effective pharmacological treatments. Our recent research revealed that pretreatment with the angiotensin II type 2 receptor (AT2R) agonist C21 alleviates kidney damage during IR. Primary cilia are organelles crucial for regulation of epithelial cell homeostasis, which are significantly affected by IR injury. This study aimed to evaluate the impact of AT2R activation on cilia integrity during IR and to identify pathways involved in the nephroprotective effect of C21. Rats were subjected to 40 min of unilateral ischaemia followed by 24 h of reperfusion. Immunofluorescence analysis of the kidneys showed that the nephroprotective effect of C21 was associated with preservation of cilia integrity in tubular cells. AT2R agonists increased α-tubulin acetylation in primary cilia in tubular cells in vivo and in a cell model. Analysis of ERK phosphorylation indicated that AT2R activation led to diminished activation of ERK1/2 in tubular cells. Similar to AT2R agonists, inhibitors of α-tubulin deacetylase HDAC6 or inhibitors of ERK activation ameliorated IR-induced cell death and preserved cilia integrity. Immunofluorescence analysis of tubular cells revealed significant ERK localization at primary cilia and demonstrated that ERK inhibition increased cilia levels of acetylated α-tubulin. Overall, our findings demonstrate that C21 elicits a preconditioning effect that enhances cilia stability in renal tubular cells, thereby preserving their integrity when exposed to IR injury. Furthermore, our results indicate that this effect might be mediated by AT2R-induced inhibition of ERK activation. These findings offer potential insights for the development of pharmacological interventions to mitigate IR-associated AKI. KEY POINTS: The AT2R agonist C21 prevents primary cilia shortening and tubular cell deciliation during renal ischaemia-reperfusion. AT2R activation inhibits ERK1/2 in renal tubular cells. Both AT2R agonists and ERK1/2 inhibitors increase alpha-tubulin acetylation at the primary cilium in tubular cells. AT2R activation, ERK1/2 inhibition or inhibition of alpha-tubulin deacetylation elicit protective effects in tubular cells subjected to ischaemia-reperfusion injury.
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Cilios , Receptor de Angiotensina Tipo 2 , Daño por Reperfusión , Animales , Masculino , Ratas , Acetilación , Lesión Renal Aguda/metabolismo , Lesión Renal Aguda/patología , Cilios/metabolismo , Cilios/efectos de los fármacos , Imidazoles , Túbulos Renales/metabolismo , Túbulos Renales/patología , Sistema de Señalización de MAP Quinasas , Ratas Sprague-Dawley , Receptor de Angiotensina Tipo 2/metabolismo , Receptor de Angiotensina Tipo 2/agonistas , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Sulfonamidas , Tiofenos , Tubulina (Proteína)/metabolismoRESUMEN
The endogenous metabolite of estradiol, estradiol 17ß-D-glucuronide (E17G), is considered the main responsible of the intrahepatic cholestasis of pregnancy. E17G alters the activity of canalicular transporters through a signaling pathway-dependent cellular internalization, phenomenon that was attributed to oxidative stress in different cholestatic conditions. However, there are no reports involving oxidative stress in E17G-induced cholestasis, representing this the aim of our work. Using polarized hepatocyte cultures, we showed that antioxidant compounds prevented E17G-induced Mrp2 activity alteration, being this alteration equally prevented by the NADPH oxidase (NOX) inhibitor apocynin. The model antioxidant N-acetyl-cysteine prevented, in isolated and perfused rat livers, E17G-induced impairment of bile flow and Mrp2 activity, thus confirming the participation of reactive oxygen species (ROS) in this cholestasis. In primary cultured hepatocytes, pretreatment with specific inhibitors of ERK1/2 and p38MAPK impeded E17G-induced ROS production; contrarily, NOX inhibition did not affect ERK1/2 and p38MAPK phosphorylation. Both, knockdown of p47phox by siRNA and preincubation with apocynin in sandwich-cultured rat hepatocytes significantly prevented E17G-induced internalization of Mrp2, suggesting a crucial role for NOX in this phenomenon. Concluding, E17G-induced cholestasis is partially mediated by NOX-generated ROS through internalization of canalicular transporters like Mrp2, being ERK1/2 and p38MAPK necessary for NOX activation.
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Estradiol , Hepatocitos , NADPH Oxidasas , Especies Reactivas de Oxígeno , Animales , NADPH Oxidasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Ratas , Hepatocitos/metabolismo , Hepatocitos/efectos de los fármacos , Estradiol/farmacología , Estradiol/metabolismo , Estradiol/análogos & derivados , Femenino , Colestasis/inducido químicamente , Colestasis/metabolismo , Colestasis/patología , Ratas Wistar , Acetofenonas/farmacología , Estrés Oxidativo/efectos de los fármacos , Acetilcisteína/farmacología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células Cultivadas , Antioxidantes/farmacología , Antioxidantes/metabolismo , Colestasis Intrahepática , Complicaciones del Embarazo , Transportadoras de Casetes de Unión a ATPRESUMEN
BACKGROUND: Endotoxemia is a severe and dangerous clinical syndrome that results in elevated morbidity, especially in intensive care units. Neonates are particularly susceptible to endotoxemia due to their immature immune systems. There are few effective treatments for neonatal endotoxemia. One group of compounds with potential in the treatment of neonatal inflammatory diseases such as endotoxemia is the flavonoids, mainly due to their antioxidant and anti-inflammatory properties. Among these, naringenin (NGN) is a citrus flavonoid which has already been reported to have anti-inflammatory, antioxidant, anti-nociceptive and anti-cancer effects. Unfortunately, its clinical application is limited by its low solubility and bioavailability. However, cyclodextrins (CDs) have been widely used to improve the solubility of nonpolar drugs and enhance the bioavailability of these natural products. OBJECTIVE: We, therefore, aimed to investigate the effects of NGN non-complexed and complexed with hydroxypropyl-ß-cyclodextrin (HPßCD) on neonatal endotoxemia injuries in a rodent model and describe the probable molecular mechanisms involved in NGN activities. METHOD: We used exposure to a bacterial lipopolysaccharide (LPS) to induce neonatal endotoxemia in the mice. RESULTS: It was found that NGN (100 mg/kg i.p.) exposure during the neonatal period reduced leukocyte migration and decreased pro-inflammatory cytokine (TNF-α, IL-1ß and IL-6) levels in the lungs, heart, kidneys or cerebral cortex. In addition, NGN upregulated IL-10 production in the lungs and kidneys of neonate mice. The administration of NGN also enhanced antioxidant enzyme catalase and SOD activity, reduced lipid peroxidation and protein carbonylation and increased the reduced sulfhydryl groups in an organ-dependent manner, attenuating the oxidative damage caused by LPS exposure. NGN decreased ERK1/2, p38MAPK and COX-2 activation in the lungs of neonate mice. Moreover, NGN complexed with HPßCD was able to increase the animal survival rate. CONCLUSION: NGN attenuated inflammatory and oxidative damage in the lungs, heart and kidneys caused by neonatal endotoxemia through the MAPK signaling pathways regulation. Our results show that NGN has beneficial effects against neonatal endotoxemia and could be useful in the treatment of neonatal inflammatory injuries.
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Citrus , Endotoxemia , Flavanonas , Ratones , Animales , Flavonoides/uso terapéutico , 2-Hidroxipropil-beta-Ciclodextrina/farmacología , Antioxidantes/farmacología , Antioxidantes/uso terapéutico , Endotoxemia/inducido químicamente , Endotoxemia/tratamiento farmacológico , Lipopolisacáridos/uso terapéutico , Antiinflamatorios/farmacologíaRESUMEN
Fibrosis is one of the main factors that impair the function of many organs. In the heart, fibrosis leads to contractile dysfunction and arrhythmias, which are important in the development of heart failure. Interleukin (IL)-11 is regulated in various heart diseases and has recently been reported to be an important cytokine in fibrosis in this organ. However, this topic has been little explored, and many questions persist. Thus, this systematic review aimed to report on possible IL-11 therapies evaluated in rodent model-induced cardiac fibrosis. Inclusion criteria were experimental in vivo studies that used different rodent models for cardiac fibrosis associated with IL-11 interventions, without year and language restrictions. The search in PubMed, Web of Science, and Embase databases was performed in October 2022. The risk of bias assessment of the studies was based on the guidelines of the SYRCLE tool, and data from the selected articles were also presented in a table as a narrative description. This review was based on eight studies in which five different interventions were used: recombinant human IL-11 (rhIL-11), anti-IL11 (X203), recombinant mouse IL-11 (rmIL-11), lentivirus (LV)-IL-11 + lutein, and anti-IL11RA (X209). Based on the included studies, the results were variable, with IL-11 overexpression inducing cardiac fibrosis, while inhibition protected against this process, preserving the function of this organ. Therefore, IL-11 stands out as a promising therapeutic target for cardiac fibrosis. However, further studies are needed to understand the mechanisms triggered by each treatment, as well as its safety and immunogenicity.
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The angiotensin II type 2 (AT2) receptor has a role in promoting insulin sensitivity. However, the mechanisms underlying the AT2 receptor-induced facilitation of insulin are still not completely understood. Therefore, we investigated whether acute in vivo administration of AT2 receptor agonist compound 21 (C21) could activate insulin signaling molecules in insulin-target tissues. We report that, in male C57BL/6 mice, an acute (5 min, 0.25 mg/kg; i.v.) injection of C21 induces the phosphorylation of Akt and ERK1/2 at activating residues (Ser473 and Thr202/Tyr204, respectively) in both epididymal white adipose tissue (WAT) and heart tissue. In WAT, the extent of phosphorylation (p) of Akt and ERK1/2 induced by C21 was approximately 65% of the level detected after a bolus injection of a dose of insulin known to induce maximal activation of the insulin receptor (IR). In the heart, C21 stimulated p-Akt to a lesser extent than in WAT and stimulated p-ERK1/2 to similar levels to those attained by insulin administration. C21 did not modify p-IR levels in either tissue. We conclude that in vivo injection of the AT2 receptor agonist C21 activates Akt and ERK1/2 through a mechanism that does not involve the IR, indicating the participation of these enzymes in AT2R-mediated signaling.
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Insulinas , Proteínas Proto-Oncogénicas c-akt , Ratones , Animales , Masculino , Fosforilación , Sistema de Señalización de MAP Quinasas , Ratones Endogámicos C57BL , Tejido Adiposo/metabolismo , Receptor de Angiotensina Tipo 2/metabolismoRESUMEN
Corticotropin releasing hormone (CRH) is crucial for basal and stress-initiated reactions in the hypothalamic-pituitary-adrenal axis (HPA) and extrahypothalamic brain circuits, where it acts as a neuromodulator to organize behavioral and humoral responses to stress. We review and describe cellular components and molecular mechanisms involved in CRH system signaling through G protein-coupled receptors (GPCRs) CRHR1 and CRHR2, under the current view of GPCR signaling from the plasma membrane but also from intracellular compartments, which establish the bases of signal resolution in space and time. Focus is placed on latest studies of CRHR1 signaling in physiologically significant contexts of the neurohormone function that disclosed new mechanistic features of cAMP production and ERK1/2 activation. We also introduce in a brief overview the pathophysiological function of the CRH system, underlining the need for a complete characterization of CRHRs signaling to design new and specific therapies for stress-related disorders.
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Sistema Hipotálamo-Hipofisario , Sistema Hipófiso-Suprarrenal , Humanos , Sistema Hipotálamo-Hipofisario/metabolismo , Sistema Hipófiso-Suprarrenal/metabolismo , Hormona Liberadora de Corticotropina/metabolismo , Sistema Nervioso Central/metabolismo , EndocitosisRESUMEN
Purinergic signaling has been implicated in many biological functions, including development. In this study, we investigate the functions of extracellular adenosine and adenosine receptors using a mouse embryonic stem cell (ESC) line and morula stages isolated from mouse embryos. Feeder-free mouse ESC was investigated in the absence and presence of the leukemia inhibitory factor (LIF), configuring undifferentiated cells and cells undergoing spontaneous differentiation. High alkaline phosphatase (ALPL) and low CD73 levels resulting in low adenosine (eADO) levels were characteristic for pluripotent cells in the presence of the LIF, while LIF deprivation resulted in augmented adenosine levels and reduced pluripotency marker expression, which indicated differentiation. Tracing ESC proliferation by BrdU labeling revealed that the inhibition of ALPL by levamisole resulted in a decrease in proliferation due to less eADO accumulation. Furthermore, caffeine and levamisole treatment, inhibiting adenosine receptor and eADO accumulation, respectively, reduced ESC migration, similar to that observed in the absence of the LIF. Pharmacological approaches of selective adenosine receptor subtype inhibition triggered specific adenosine receptor activities, thus triggering calcium or MAP kinase pathways leading to differentiation. In line with the in vitro data, mouse embryos at the morula stage were sensitive to treatments with A1 and A3 receptor antagonists, leading to the conclusion that A1 receptor and A3 receptor inhibition impairs proliferation and self-renewal and triggers inappropriate differentiation, respectively. The findings herein define the functions of eADO signaling in early development with implications for developmental disorders, in which adenosine receptors or ectonucleotidase dysfunctions are involved, and which could lead to malformations and miscarriages, due to exposure to caffeine.
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INTRODUCTION: Calotropis procera (Aiton) Dryand (Apocynaceae) is an herb that has been commonly used in folk medicine to treat various diseases for more than 1500 years. AIMS: Our goal was to investigate the anti-metastatic effects of phenolics extracted from C. procera (CphE) against 4T1 breast cancer cells and in BALB/c mice. METHODS: 4T1 cells were treated with CphE and quercetin (positive control) at concentrations that inhibited cell viability by 50% (IC50). Levels of reactive oxygen species (ROS), wound healing, and protein expressions were determined following standard protocols. For the in vivo pilot study, the syngeneic BALB/c mouse model was used. 4T1 cells were injected into mammary fat pads. Tumors were allowed to grow for 9 days before gavage treatment with CphE (150 mg GAE/kg/day) or PBS (controls) for one week. Excised tumors, liver, and lungs were analyzed for gene and protein expression and histology. RESULTS: In vitro results showed that CphE suppressed cell viability through apoptosis induction, via caspase-3 cleavage and total PARP reduction. CphE also scavenged ROS and suppressed Akt, mTOR, ERK1/2, CREB, and Src activation contributing to cell motility inhibition. CphE reduced IR, PTEN, TSC2, p70S6, and RPS6, protein levels, which are proteins involved in the PI3K/Akt/mTOR pathway, suggesting this pathway as CphE primary target. In vivo results showed downregulation of ERK1/2 activation by phosphorylation in tumor tissues, accompanied by angiogenesis reduction in tumor and lung tissues. A reduction of Cenpf mRNA levels in liver and lung tissues strongly suggested anti-invasive cancer activity of CphE. CONCLUSION: CphE inhibited 4T1 cell signal pathways that play a key role in cell growth and invasion. The potential for in vitro results to be translated in vivo was confirmed. A complete animal study is a guarantee to confirm the CphE anticancer and antimetastatic activity in vivo.
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Calotropis , Neoplasias , Animales , Apoptosis , Calotropis/genética , Calotropis/metabolismo , Caspasa 3/metabolismo , Línea Celular Tumoral , Ratones , Ratones Endogámicos BALB C , Fosfatidilinositol 3-Quinasas/metabolismo , Proyectos Piloto , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Quercetina/farmacología , ARN Mensajero , Especies Reactivas de Oxígeno , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
BACKGROUND: Betahistine is a clinical medication for the treatment of benign paroxysmal positional vertigo (BPPV). Otolin, a secreted glycoprotein with a C-terminal globular domain homologous to the immune complement C1q, has been identified as a biomarker for BPPV. However, the role of complement C1q/TNF-related proteins (CTRPs) with a C-terminal globular domain in BPPV is unclear, so we explored the change of CTRPs in betahistine treated BPPV. METHODS: We treated BPPV patients with Betahistine (12 mg/time, 3 times/day) for 4 weeks and observed the clinical efficacy and the expression of CTRP family members in BPPV patients. Then, we constructed a vertigo mice model of vestibular dysfunction with gentamicin (150 mg/Kg) and a BPPV model of Slc26a4loop/loop mutant mice. Adenoviral vectors for CTRP expression vector and small interfering RNA were injected via the intratympanic injection into mice and detected the expression of CTRP family members, phosphorylation levels of ERK and AKT and the expression of PPARγ. In addition, we treated mice of vestibular dysfunction with Betahistine (10 mg/Kg) and/or ERK inhibitor of SCH772984 (12 mg/Kg) and/or and PPARγ antagonist GW9662 (1 mg/Kg) for 15 days, and evaluated the accuracy of air righting reflex, the time of contact righting reflex and the scores of head tilt and swimming behavior. RESULTS: After treatment with Betahistine, the residual dizziness duration and the score of the evaluation were reduced, and the expression of CTRP1, 3, 6, 9 and 12 were significantly increased in BPPV patients. We also found that Betahistine improved the accuracy of air righting reflex, reduced the time of contact righting reflex and the scores of head tilt and swimming behavior in gentamicin-treated mice and Slc26a4loop/loop mutant mice. The expression levels of CTRP1, 3, 6, 9 and 12, phosphorylation levels of ERK and AKT, and PPARγ expression were significantly increased, and the scores of head tilt and swimming behavior were decreased in vestibular dysfunction mice with overexpression of CTRPs. Silencing CTRPs has the opposite effect. SCH772984 reversed the effect of Betahistine in mice with vestibular dysfunction. CONCLUSION: Betahistine alleviates BPPV through inducing production of multiple CTRP family members and activating the ERK1/2-AKT/PPARy pathway.
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Vértigo Posicional Paroxístico Benigno , Betahistina , Animales , Vértigo Posicional Paroxístico Benigno/tratamiento farmacológico , Betahistina/farmacología , Betahistina/uso terapéutico , Mareo/tratamiento farmacológico , Humanos , Sistema de Señalización de MAP Quinasas , Ratones , PPAR gamma , Proteínas Proto-Oncogénicas c-aktRESUMEN
Leishmania spp. infection outcomes are dependent on both host and parasite factors. Manipulation of host signaling pathways involved in the generation of immune responses is thought to be one of the most common mechanisms used by parasites for persistence within the host. Considering the diversity of pathologies caused by different Leishmania spp., it is plausible that significant differences may exist in the mechanisms of host cell manipulation by each parasite species, which may have implications when developing new vaccine or treatment strategies. Here we show that in L. braziliensis-infection in BALB/c mice, a model of resistance, activation of ERK1/2 coincides with the peak of inflammatory responses and resolution of tissue parasitism. In contrast, in the susceptibility model of L. amazonensis-infection, an early silent phase of infection is observed, detected solely by quantification of parasite loads. At this early stage, only basal levels of P-ERK1/2 are observed. Later, after a brief shutdown of ERK1/2 phosphorylation, disease progression is observed and is associated with increased inflammation, lesion size and tissue parasitism. Moreover, the short-term down-regulation of ERK1/2 activation affected significantly downstream inflammatory pathways and adaptive T cell responses. Administration of U0126, a MEK/ERK inhibitor, confirmed this phenomenon, since bigger lesions and higher parasite loads were seen in infected mice that received U0126. To investigate how kinetics of ERK1/2 activation could affect the disease progression, U0126 was administered to L. amazonensis-infected animals earlier than the P-ERK1/2 switch off time-point. This intervention resulted in anticipation of the same effects on inflammatory responses and susceptibility phenotype seen in the natural course of infection. Additionally, in vitro inhibition of ERK1/2 affected the phagocytosis of L. amazonensis by BMDMs. Collectively, our findings reveal distinct temporal patterns of activation of inflammatory responses in L. braziliensis and L. amazonensis in the same animal background and a pivotal role for a brief and specific shutdown of ERK1/2 activation at late stages of L. amazonensis infection. Since activation of inflammatory responses is a crucial aspect for the control of infectious processes, these findings may be important for the search of new and specific strategies of vaccines and treatment for tegumentary leishmaniasis.
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Inmunidad Celular , Leishmania mexicana/inmunología , Leishmaniasis/inmunología , Leishmaniasis/metabolismo , Leishmaniasis/parasitología , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Animales , Biomarcadores , Citocinas/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Interacciones Huésped-Patógeno/inmunología , Mediadores de Inflamación/metabolismo , Leishmaniasis/patología , Ratones , Carga de Parásitos , Fagocitosis/inmunología , Fosforilación , Transducción de SeñalRESUMEN
Insulin-like growth factor-1 (IGF-1) plays an important role in function and development of the mammary gland. However, high levels of IGF-1 has been associated with an increased risk of breast cancer development. Epithelial-mesenchymal transition (EMT) is a process where epithelial cells lose their epithelial characteristics and acquire a mesenchymal phenotype, which is considered one of the most important mechanisms in cancer initiation and promotion of metastasis. Extracellular vesicles (EVs) are released into the extracellular space by different cell types, which mediate intercellular communication and play an important role in different physiological and pathological processes, such as cancer. In this study, we demonstrate that EVs from MDA-MB-231 breast cancer cells stimulated with IGF-1 (IGF-1 EVs) decrease the levels of E-cadherin, increase the expression of vimentin and N-cadherin and stimulate the secretion of metalloproteinase-9 in mammary non-tumorigenic epithelial cells MCF10A. IGF-1 EVs also induce the expression of Snail1, Twist1 and Sip1, which are transcription factors involved in EMT. Moreover, IGF-1 EVs induce activation of ERK1/2, Akt1 and Akt2, migration and invasion. In summary, we demonstrate, for the first time, that IGF-1 EVs induce an EMT process in mammary non-tumorigenic epithelial cells MCF10A.
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BACKGROUND: Betahistine is a clinical medication for the treatment of benign paroxysmal positional vertigo (BPPV). Otolin, a secreted glycoprotein with a C-terminal globular domain homologous to the immune complement C1q, has been identified as a biomarker for BPPV. However, the role of complement C1q/TNF-related proteins (CTRPs) with a C-terminal globular domain in BPPV is unclear, so we explored the change of CTRPs in betahistine treated BPPV. METHODS: We treated BPPV patients with Betahistine (12 mg/time, 3 times/day) for 4 weeks and observed the clinical efficacy and the expression of CTRP family members in BPPV patients. Then, we constructed a vertigo mice model of vestibular dysfunction with gentamicin (150 mg/Kg) and a BPPV model of Slc26a4loop/loop mutant mice. Adenoviral vectors for CTRP expression vector and small interfering RNA were injected via the intratympanic injection into mice and detected the expression of CTRP family members, phosphorylation levels of ERK and AKT and the expression of PPARγ. In addition, we treated mice of vestibular dysfunction with Betahistine (10 mg/Kg) and/or ERK inhibitor of SCH772984 (12 mg/Kg) and/or and PPARγ antagonist GW9662 (1 mg/Kg) for 15 days, and evaluated the accuracy of air righting reflex, the time of contact righting reflex and the scores of head tilt and swimming behavior. RESULTS: After treatment with Betahistine, the residual dizziness duration and the score of the evaluation were reduced, and the expression of CTRP1, 3, 6, 9 and 12 were significantly increased in BPPV patients. We also found that Betahistine improved the accuracy of air righting reflex, reduced the time of contact righting reflex and the scores of head tilt and swimming behavior in gentamicin-treated mice and Slc26a4loop/loop mutant mice. The expression levels of CTRP1, 3, 6, 9 and 12, phosphorylation levels of ERK and AKT, and PPARγ expression were significantly increased, and the scores of head tilt and swimming behavior were decreased in vestibular dysfunction mice with overexpression of CTRPs. Silencing CTRPs has the opposite effect. SCH772984 reversed the effect of Betahistine in mice with vestibular dysfunction. CONCLUSION: Betahistine alleviates BPPV through inducing production of multiple CTRP family members and activating the ERK1/2-AKT/PPARy pathway.
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Humanos , Animales , Ratones , Betahistina/uso terapéutico , Betahistina/farmacología , Vértigo Posicional Paroxístico Benigno/tratamiento farmacológico , Sistema de Señalización de MAP Quinasas , PPAR gamma , Mareo/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-aktRESUMEN
The cardiac circadian clock is responsible for the modulation of different myocardial processes, and its dysregulation has been linked to disease development. How this clock machinery is regulated in the heart remains an open question. Because noradrenaline (NE) can act as a zeitgeber in cardiomyocytes, we tested the hypothesis that adrenergic signaling resets cardiac clock gene expression in vivo. In its anti-phase with Clock and Bmal1, cardiac Per1 abundance increased during the dark phase, concurrent with the rise in heart rate and preceded by an increase in NE levels. Sympathetic denervation altered Bmal1 and Clock amplitude, while Per1 was affected in both amplitude and oscillatory pattern. We next treated mice with a ß-adrenergic receptor (ß-AR) blocker. Strikingly, the ß-AR blockade during the day suppressed the nocturnal increase in Per1 mRNA, without altering Clock or Bmal1. In contrast, activating ß-AR with isoproterenol (ISO) promoted an increase in Per1 expression, demonstrating its responsiveness to adrenergic input. Inhibitors of ERK1/2 and CREB attenuated ISO-induced Per1 expression. Upstream of ERK1/2, PI3Kγ mediated ISO induction of Per1 transcription, while activation of ß2-AR, but not ß1-AR induced increases in ERK1/2 phosphorylation and Per1 expression. Consistent with the ß2-induction of Per1 mRNA, ISO failed to activate ERK1/2 and elevate Per1 in the heart of ß2-AR-/- mice, whereas a ß2-AR antagonist attenuated the nocturnal rise in Per1 expression. Our study established a link between NE/ß2-AR signaling and Per1 oscillation via the PI3Ky-ERK1/2-CREB pathway, providing a new framework for understanding the physiological mechanism involved in resetting cardiac clock genes.
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Regulación de la Expresión Génica , Sistema de Señalización de MAP Quinasas , Miocardio/metabolismo , Proteínas Circadianas Period/biosíntesis , Receptores Adrenérgicos beta 2/metabolismo , Factores de Transcripción ARNTL/biosíntesis , Factores de Transcripción ARNTL/genética , Antagonistas de Receptores Adrenérgicos beta 2/farmacología , Animales , Proteínas CLOCK/biosíntesis , Isoproterenol/farmacología , Masculino , Ratones , Ratones Noqueados , Proteínas Circadianas Period/genética , Receptores Adrenérgicos beta 2/genéticaRESUMEN
Ligand-independent epidermal growth factor receptor (EGFR) endocytosis is inducible by a variety of stress conditions converging upon p38 kinase. A less known pathway involves phosphatidic acid (PA) signaling toward the activation of type 4 phosphodiesterases (PDE4) that decrease cAMP levels and protein kinase A (PKA) activity. This PA/PDE4/PKA pathway is triggered with propranolol used to inhibit PA hydrolysis and induces clathrin-dependent and clathrin-independent endocytosis, followed by reversible accumulation of EGFR in recycling endosomes. Here we give further evidence of this signaling pathway using biosensors of PA, cAMP, and PKA in live cells and then show that it activates p38 and ERK1/2 downstream the PKA inhibition. Clathrin-silencing and IN/SUR experiments involved the activity of p38 in the clathrin-dependent route, while ERK1/2 mediates clathrin-independent EGFR endocytosis. The PA/PDE4/PKA pathway selectively increases the EGFR endocytic rate without affecting LDLR and TfR constitute endocytosis. This selectiveness is probably because of EGFR phosphorylation, as detected in Th1046/1047 and Ser669 residues. The EGFR accumulates at perinuclear recycling endosomes colocalizing with TfR, fluorescent transferrin, and Rab11, while a small proportion distributes to Alix-endosomes. A non-selective recycling arrest includes LDLR and TfR in a reversible manner. The PA/PDE4/PKA pathway involving both p38 and ERK1/2 expands the possibilities of EGFR transmodulation and interference in cancer.
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Sistema de Señalización de MAP Quinasas , Ácidos Fosfatidicos , Clatrina/metabolismo , Endocitosis/fisiología , Receptores ErbB/metabolismo , Ligandos , Ácidos Fosfatidicos/metabolismo , Fosforilación , Transducción de SeñalRESUMEN
Oxidative stress is a key factor contributing to the development of diabetes complications. Glutathione S-transferases (GSTs) protect against products of oxidative stress by conjugating glutathione to electrophilic substrates, producing compounds that are generally less reactive and more soluble. The expression and activity of GSTs during diabetes have been extensively studied, but little is known about regulation mechanisms of Pi-class GST (GSTP). The aim of the present study was to evaluate how GSTP is regulated in a Streptozotocin (STZ)-induced murine diabetes model. GST activity and GSTP expression were determined in adult male mice diabetized with STZ. Specificity protein 1 (Sp1) expression and O-glycosylation, as well as the role of AP-1 members Jun and Fos in the regulation of GSTP expression, were also assessed. The results showed that GST total activity and GSTP mRNA and protein levels were decreased in the diabetic liver, and returned to normal values after insulin administration. The insulin-mimetic drug vanadate was also able to restore GST activity, but failed to recover GSTP mRNA/protein levels. In diabetic animals, O-glycosylated Sp1 levels were increased, whereas, in insulin-treated animals, glycosylation values were similar to those of controls. After vanadate administration, Sp1 expression levels and glycosylation were lower than those of controls. Our results suggest that hyperglycemia could lead to the observed increase in Sp1 O-glycosylation, which would, in turn, lead to a decrease in the expression of Sp1-dependent GSTP in the liver of diabetic mice.
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The maintenance of homeostasis in living systems requires the elimination of unwanted cells which is performed, among other mechanisms, by type I cell death or apoptosis. This type of programmed cell death involves several morphological changes such as cytoplasm shrinkage, chromatin condensation (pyknosis), nuclear fragmentation (karyorrhexis), and plasma membrane blebbing that culminate with the formation of apoptotic bodies. In addition to the maintenance of homeostasis, apoptosis also represents an important defense mechanism for cells against intracellular microorganisms. In counterpart, diverse intracellular pathogens have developed a wide array of strategies to evade apoptosis and persist inside cells. These strategies include the manipulation of signaling pathways involved in the inhibition of apoptosis where mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K) play a key role. Leishmania is an intracellular protozoan parasite that causes a wide spectrum of diseases known as leishmaniasis. This parasite displays different strategies, including apoptosis inhibition, to down-regulate host cell defense mechanisms in order to perpetuate infection.
RESUMEN
The combined allergic rhinitis and asthma syndrome (CARAS) is a chronic airway inflammation of allergic individuals, with a type 2 immune response. Pharmacotherapy is based on drugs with relevant side effects. Thus, the goal of this study was to evaluate the synthetic alkaloid, MHTP in the experimental model of CARAS. Therefore, BALB/c mice were ovalbumin (OVA) -sensitized and -challenged and treated with MHTP by intranasal or oral routes. Treated animals showed a decrease (p < 0.05) of sneezing, nasal rubbings, and histamine nasal hyperactivity. Besides, MHTP presented binding energy and favorable interaction for adequate anchoring in the histamine H1 receptor. MHTP treatment inhibited the eosinophil migration into the nasal (NALF) and the bronchoalveolar (BALF) fluids. Histological analysis showed that the alkaloid decreased the inflammatory cells in the subepithelial and perivascular regions of nasal tissue and in the peribronchiolar and perivascular regions of lung tissue. The MHTP treatment also reduced the pulmonary hyperactivity by decreasing the smooth muscle layer hypertrophy and the collagen fiber deposition in the extracellular matrix. The immunomodulatory effect of the alkaloid was due to the decrease of cytokines like IL-5 and IL-17A (type 2 and 3), TSLP (epithelial), and the immunoregulatory cytokine, TGF-ß. These MHTP effects on granulocytes were dependent on the p38/ERK1/2 MAP kinase signaling pathway axis. Indeed, the synthetic alkaloid reduced the frequency of activation of both kinases independent of the NF-κB (p65) pathway indicating that the molecule shut down the intracellular transduction signals underlie the cytokine gene transcription.
Asunto(s)
Antiasmáticos/uso terapéutico , Asma/tratamiento farmacológico , Rinitis Alérgica/tratamiento farmacológico , Tetrahidroisoquinolinas/uso terapéutico , Alérgenos/inmunología , Animales , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Sistema de Señalización de MAP Quinasas , Ratones , Ratones Endogámicos BALB C , Simulación del Acoplamiento Molecular , Ovalbúmina/inmunología , Receptores Histamínicos H1/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
During pregnancy, Toxoplasma gondii can triggers serious manifestations and potentially affect the fetal development. In this scenario, differences in susceptibility of trophoblast cells to T. gondii infection might be evaluated in order to establish new therapeutic approaches capable of interfering in the control of fetal infection by T. gondii. This study aimed to evaluate the susceptibility of cytotrophoblast, syncytiotrophoblast and extravillous trophoblast cells to T. gondii infection. Our data demonstrate that HTR-8/SVneo cells (extravillous trophoblast cells) present higher susceptibility to T. gondii infection when compared to syncytiotrophoblast and cytotrophoblast cells, whereas syncytiotrophoblast was the cell type more resistant to the parasite infection. Also, cytotrophoblast and syncytiotrophoblast cells produced significantly more IL-6 than HTR-8/SVneo cells. On the other hand, HTR-8/SVneo cells showed higher ERK1/2 phosphorylation than cytotrophoblast and syncytiotrophoblast cells. ERK1/2 inhibition reduced T. gondii infection and increased IL-6 production in HTR-8/SVneo cells. Thus, it is plausible to conclude that the greater susceptibility of HTR-8/SVneo cells to infection by T. gondii is related to a higher ERK1/2 phosphorylation and lower levels of IL-6 in these cells compared to other cells, suggesting that these mediators may be important to favor the parasite infection in this type of trophoblastic population.