RESUMO
Experiments aimed to evaluate the tissue distribution of Mas-related G protein-coupled receptor D (MrgD) revealed the presence of immunoreactivity for the MrgD protein in the rostral insular cortex (rIC), an important area for autonomic and cardiovascular control. To investigate the relevance of this finding, we evaluated the cardiovascular effects produced by the endogenous ligand of MrgD, alamandine, in this brain region. Mean arterial pressure (MAP), heart rate (HR), and renal sympathetic nerve activity (RSNA) were recorded in urethane anesthetized rats. Unilateral microinjection of equimolar doses of alamandine (40 pmol/100 nL), angiotensin-(1-7), angiotensin II, angiotensin A, and Mas/MrgD antagonist d-Pro7-Ang-1-7 (50 pmol/100 nL), Mas antagonist A779 (100 pmol/100 nL), or vehicle (0.9% NaCl) were made in different rats (n = 4-6/group) into rIC. To verify the specificity of the region, a microinjection of alamandine was also performed into intermediate insular cortex (iIC). Microinjection of alamandine in rIC produced an increase in MAP (Δ = 15 ± 2 mmHg), HR (Δ = 36 ± 4 beats/min), and RSNA (Δ = 31 ± 4%), but was without effects at iIC. Strikingly, an equimolar dose of angiotensin-(1-7) at rIC did not produce any change in MAP, HR, and RSNA. Angiotensin II and angiotensin A produced only minor effects. Alamandine effects were not altered by A-779, a Mas antagonist, but were completely blocked by the Mas/MrgD antagonist d-Pro7-Ang-(1-7). Therefore, we have identified a brain region in which alamandine/MrgD receptor but not angiotensin-(1-7)/Mas could be involved in the modulation of cardiovascular-related neuronal activity. This observation also suggests that alamandine might possess unique effects unrelated to angiotensin-(1-7) in the brain.
Assuntos
Angiotensina I/farmacologia , Pressão Arterial/efeitos dos fármacos , Sistema Cardiovascular/inervação , Córtex Cerebral/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , Rim/inervação , Proteínas do Tecido Nervoso/agonistas , Oligopeptídeos/farmacologia , Fragmentos de Peptídeos/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Sistema Nervoso Simpático/efeitos dos fármacos , Animais , Córtex Cerebral/fisiologia , Ligantes , Masculino , Microinjeções , Proteínas do Tecido Nervoso/metabolismo , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas/agonistas , Proteínas Proto-Oncogênicas/metabolismo , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas G/metabolismo , Sistema Nervoso Simpático/fisiologiaRESUMO
GPER-1 is a novel membrane sited G protein-coupled estrogen receptor. Clinical studies have shown that patients suffering an estrogen receptor α (ERα)/GPER-1 positive, breast cancer have a lower survival rate than those who have developed ERα-positive/GPER-1 negative tumors. Moreover, absence of GPER-1 improves the prognosis of patients treated with tamoxifen, the most used selective estrogen receptor modulator to treat ERα-positive breast cancer. MCF-7 breast cancer cells were continuously treated with 1,000 nM tamoxifen for 7 days to investigate its effect on GPER-1 protein expression, cell proliferation and intracellular [Ca2+]i mobilization, a key signaling pathway. Breast cancer cells continuously treated with tamoxifen, exhibited a robust [Ca2+]i mobilization after stimulation with 1,000 nM tamoxifen, a response that was blunted by preincubation of cells with G15, a commercial GPER-1 antagonist. Continuously treated cells also displayed a high [Ca2+]i mobilization in response to a commercial GPER-1 agonist (G1) and to estrogen, in a magnitude that doubled the response observed in untreated cells and was almost completely abolished by G15. Proliferation of cells continuously treated with tamoxifen and stimulated with 2,000 nM tamoxifen, was also higher than that observed in untreated cells in a degree that was approximately 90% attributable to GPER-1. Finally, prolonged tamoxifen treatment did not increase ERα expression, but did overexpress the kinin B1 receptor, another GPCR, which we have previously shown is highly expressed in breast tumors and increases proliferation of breast cancer cells. Although we cannot fully extrapolate the results obtained in vitro to the patients, our results shed some light on the occurrence of drug resistance in breast cancer patients who are ERα/GPER-1 positive, have been treated with tamoxifen and display low survival rate. Overexpression of kinin B1 receptor may explain the increased proliferative response observed in breast tumors under continuous treatment with tamoxifen.
Assuntos
Antineoplásicos Hormonais/administração & dosagem , Neoplasias da Mama/metabolismo , Proliferação de Células/efeitos dos fármacos , Receptores de Estrogênio/biossíntese , Receptores Acoplados a Proteínas G/biossíntese , Tamoxifeno/administração & dosagem , Neoplasias da Mama/patologia , Proliferação de Células/fisiologia , Feminino , Humanos , Células MCF-7 , Receptores Acoplados a Proteínas G/agonistas , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologiaRESUMO
Free fatty acid (FFA) receptors FFA1 and FFA4 are omega-3 molecular targets in metabolic diseases; however, their function in cancer cachexia remains unraveled. We assessed the role of FFA1 and FFA4 receptors in the mouse model of cachexia induced by Lewis lung carcinoma (LLC) cell implantation. Naturally occurring ligands such as α-linolenic acid (ALA) and docosahexaenoic acid (DHA), the synthetic FFA1/FFA4 agonists GW9508 and TUG891, or the selective FFA1 GW1100 or FFA4 AH7614 antagonists were tested. FFA1 and FFA4 expression and other cachexia-related parameters were evaluated. GW9508 and TUG891 decreased tumor weight in LLC-bearing mice. Regarding cachexia-related end points, ALA, DHA, and the preferential FFA1 agonist GW9508 rescued body weight loss. Skeletal muscle mass was reestablished by ALA treatment, but this was not reflected in the fiber cross-sectional areas (CSA) measurement. Otherwise, TUG891, GW1100, or AH7614 reduced the muscle fiber CSA. Treatments with ALA, GW9508, GW1100, or AH7614 restored white adipose tissue (WAT) depletion. As for inflammatory outcomes, ALA improved anemia, whereas GW9508 reduced splenomegaly. Concerning behavioral impairments, ALA and GW9508 rescued locomotor activity, whereas ALA improved motor coordination. Additionally, DHA improved grip strength. Notably, GW9508 restored abnormal brain glucose metabolism in different brain regions. The GW9508 treatment increased leptin levels, without altering uncoupling protein-1 downregulation in visceral fat. LLC-cachectic mice displayed FFA1 upregulation in subcutaneous fat, but not in visceral fat or gastrocnemius muscle, whereas FFA4 was unaltered. Overall, the present study shed new light on FFA1 and FFA4 receptors' role in metabolic disorders, indicating FFA1 receptor agonism as a promising strategy in mitigating cancer cachexia.
Assuntos
Peso Corporal/efeitos dos fármacos , Caquexia/tratamento farmacológico , Carcinoma Pulmonar de Lewis/metabolismo , Ácidos Docosa-Hexaenoicos/uso terapêutico , Receptores Acoplados a Proteínas G/metabolismo , Ácido alfa-Linolênico/uso terapêutico , Animais , Benzoatos/farmacologia , Compostos de Bifenilo/farmacologia , Caquexia/etiologia , Caquexia/metabolismo , Carcinoma Pulmonar de Lewis/complicações , Modelos Animais de Doenças , Ácidos Docosa-Hexaenoicos/farmacologia , Metilaminas/farmacologia , Camundongos , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Transplante de Neoplasias , Fenilpropionatos/farmacologia , Propionatos/farmacologia , Pirimidinas/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Sulfonamidas/farmacologia , Xantenos/farmacologia , Ácido alfa-Linolênico/farmacologiaRESUMO
Elevated plasma concentrations of the ketone body ß-hydroxybutyrate (BHB), an endogenous agonist of the hydroxycarboxylic acid receptor 2 (HCA2), is associated with an increased incidence of inflammatory diseases during lactation in dairy cows. In the early stages of this pathology, an increase in neutrophil recruitment is observed; however, the role of BHB remains elusive. This study characterized the effect of BHB and synthetic agonists of the HCA2 receptor on bovine neutrophil chemotaxis and the signaling pathways involved in this process. We demonstrated that treatment with BHB concentrations between 1.2 and 10 mM and two full selective agonists of the HCA2 receptor, MK-1903 and nicotinic acid, increased bovine neutrophil chemotaxis. We also observed that BHB and HCA2 agonists induced calcium release and phosphorylation of AKT, ERK 1/2 and AMPKα. To evaluate the role of these pathways in bovine neutrophil chemotaxis, we used the pharmacological inhibitors BAPTA-AM, pertussis toxin, U73122, LY294002, U0126 and compound C. Our results suggest that these pathways are required for HCA2 agonist-induced bovine neutrophil chemotaxis in non-physiological condition. Concentrations around 1.4 mM of BHB after calving may exert a chemoattractant effect that is key during the onset of the inflammatory process associated with metabolic disorders in dairy cows.
Assuntos
Ácido 3-Hidroxibutírico/farmacologia , Proteínas Quinases Ativadas por AMP/metabolismo , Quimiotaxia , Sistema de Sinalização das MAP Quinases , Neutrófilos/citologia , Neutrófilos/enzimologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Acoplados a Proteínas G/agonistas , Animais , Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Bovinos , Quimiotaxia/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Niacina/farmacologia , Fosforilação/efeitos dos fármacos , Pirazóis/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Fosfolipases Tipo C/metabolismoRESUMO
Hypothalamic adult neurogenesis provides the basis for renewal of neurons involved in the regulation of whole-body energy status. In addition to hormones, cytokines and growth factors, components of the diet, particularly fatty acids, have been shown to stimulate hypothalamic neurogenesis; however, the mechanisms behind this action are unknown. Here, we hypothesized that GPR40 (FFAR1), the receptor for medium and long chain unsaturated fatty acids, could mediate at least part of the neurogenic activity in the hypothalamus. We show that a GPR40 ligand increased hypothalamic cell proliferation and survival in adult mice. In postnatal generated neurospheres, acting in synergy with brain-derived neurotrophic factor (BDNF) and interleukin 6, GPR40 activation increased the expression of doublecortin during the early differentiation phase and of the mature neuronal marker, microtubule-associated protein 2 (MAP2), during the late differentiation phase. In Neuro-2a proliferative cell-line GPR40 activation increased BDNF expression and p38 activation. The chemical inhibition of p38 abolished GPR40 effect in inducing neurogenesis markers in neurospheres, whereas BDNF immunoneutralization inhibited GPR40-induced cell proliferation in the hypothalamus of adult mice. Thus, GPR40 acts through p38 and BDNF to induce hypothalamic neurogenesis. This study provides mechanistic advance in the understating of how a fatty acid receptor regulates adult hypothalamic neurogenesis.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/fisiologia , Hipotálamo/citologia , Hipotálamo/fisiologia , Neurogênese/fisiologia , Receptores Acoplados a Proteínas G/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/fisiologia , Animais , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Hipotálamo/efeitos dos fármacos , Imidazóis/farmacologia , Interleucina-6/fisiologia , Ligantes , Masculino , Metilaminas/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos Neurológicos , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Propionatos/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidoresRESUMO
The peptide angiotensin-(1-7) [Ang (1-7)] and its receptor Mas are involved in controlling arterial pressure and display actions on the nervous system. In a previous study, our laboratory showed that A779 [(peptidyl antagonist of the Ang-(1-7)] treatment had a negative effect following a lesion of the sciatic nerve, possibly by delaying the responses of Schwann cells, resulting in a decreased axonal organization along with a slowed functional return. In the present work, we investigated the central cellular changes after sciatic nerve injury in rodents treated with A779 after two weeks. In the lumbar spinal cords, where the neuronal bodies that make up the sciatic are, the treatment with A779 showed reduced reactivity of astrocytes (p = 0.004, Mann-Whitney U test) and less synaptic density (p = 0.004, Mann-Whitney U test) after injury. Also, the treatment upregulated microglia activity in both sides (p = 0.004, Mann-Whitney U test), ipsilateral and contralateral to the lesion, of the spinal cord. In addition, the Mas expression in spine neurons was increased in response to axotomy especially after two weeks (p = 0.03, Mann-Whitney U test) following the nerve lesion in comparison to earlier stages after injury. Therefore, we can conclude that Ang-(1-7)/Mas axis plays a role during spinal cord recovery after peripheral nerve injury.
Assuntos
Angiotensina II/análogos & derivados , Angiotensina I/agonistas , Axotomia , Gliose/tratamento farmacológico , Gliose/patologia , Fragmentos de Peptídeos/agonistas , Fragmentos de Peptídeos/uso terapêutico , Receptores Acoplados a Proteínas G/agonistas , Sinapses/efeitos dos fármacos , Angiotensina II/uso terapêutico , Animais , Astrócitos/efeitos dos fármacos , Imunofluorescência , Masculino , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Medula Espinal/efeitos dos fármacosRESUMO
OBJECTIVE: To investigate if ICI 182,780 (fulvestrant), a selective estrogen receptor alpha/beta (ERα/ERß) antagonist, and G-1, a selective G-protein-coupled receptor (GPER) agonist, can potentially induce autophagy in breast cancer cell lines MCF-7 and SKBr3, and how G-1 affects cell viability. METHODS: Cell viability in MCF-7 and SKBr3 cells was assessed by the MTT assay. To investigate the autophagy flux, MCF-7 cells were transfected with GFP-LC3, a marker of autophagosomes, and analyzed by real-time fluorescence microscopy. MCF-7 and SKBr3 cells were incubated with acridine orange for staining of acidic vesicular organelles and analyzed by flow cytometry as an indicator of autophagy. RESULTS: Regarding cell viability in MCF-7 cells, ICI 182,780 and rapamycin, after 48 hours, led to decreased cell proliferation whereas G-1 did not change viability over the same period. The data showed that neither ICI 182,780 nor G-1 led to increased GFP-LC3 puncta in MCF-7 cells over the 4-hour observation period. The cytometry assay showed that ICI 182,780 led to a higher number of acidic vesicular organelles in MCF-7 cells. G-1, in turn, did not have this effect in any of the cell lines. In contrast, ICI 182,780 and G-1 did not decrease cell viability of SKBr3 cells or induce formation of acidic vesicular organelles, which corresponds to the final step of the autophagy process in this cell line. CONCLUSION: The effect of ICI 182,780 on increasing acidic vesicular organelles in estrogen receptor-positive breast cancer cells appears to be associated with its inhibitory effect on estrogen receptors, and GPER does notseem to be involved. Understanding these mechanisms may guide further investigations of these receptors' involvement in cellular processes of breast cancer resistance.
Assuntos
Autofagia/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Antagonistas do Receptor de Estrogênio/farmacologia , Fulvestranto/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Análise de Variância , Western Blotting , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor beta de Estrogênio/antagonistas & inibidores , Feminino , Citometria de Fluxo/métodos , Humanos , Células MCF-7 , Receptores Acoplados a Proteínas G/análise , Reprodutibilidade dos Testes , Sirolimo/farmacologia , Fatores de Tempo , Transfecção/métodosRESUMO
AIMS: Activation of the angiotensin (Ang)-(1-7)/Mas receptor (R) axis protects from sympathetic overactivity. Endocytic trafficking is an essential process that regulates receptor (R) function and its ultimate cellular responses. We investigated whether the blunted responses to Ang-(1-7) in hypertensive rats are associated to an alteration in MasR trafficking. METHODS AND RESULTS: Brainstem neurons from Wistar-Kyoto (WKY) or spontaneously hypertensive rats (SHRs) were investigated for (i) Ang-(1-7) levels and binding and MasR expression, (ii) Ang-(1-7) responses (arachidonic acid and nitric oxide release and Akt and ERK1/2 phosphorylation), and (iii) MasR trafficking. Ang-(1-7) was determined by radioimmunoassay. MasR expression and functionality were evaluated by western blot and binding assays. MasR trafficking was evaluated by immunofluorescence. Ang-(1-7) treatment induced an increase in nitric oxide and arachidonic acid release and ERK1/2 and Akt phosphorylation in WKY neurons but did not have an effect in SHR neurons. Although SHR neurons showed greater MasR expression, Ang-(1-7)-elicited responses were substantially diminished presumably due to decreased Ang-(1-7) endogenous levels concomitant with impaired binding to its receptor. Through immunocolocalization studies, we evidenced that upon Ang-(1-7) stimulation MasRs were internalized through clathrin-coated pits and caveolae into early endosomes and slowly recycled back to the plasma membrane. However, the fraction of internalized MasRs into early endosomes was larger and the fraction of MasRs recycled back to the plasma membrane was smaller in SHR than in WKY neurons. Surprisingly, in SHR neurons but not in WKY neurons, Ang-(1-7) induced MasR translocation to the nucleus. Nuclear MasR expression and Ang-(1-7) levels were significantly greater in the nuclei of Ang-(1-7)-stimulated SHR neurons, indicating that the MasR is translocated with its ligand bound to it. CONCLUSION: MasRs display differential trafficking in brainstem neurons from SHRs, which may contribute to the impaired responses to Ang-(1-7).
Assuntos
Angiotensina I/farmacologia , Tronco Encefálico/efeitos dos fármacos , Hipertensão/metabolismo , Neurônios/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Proteínas Proto-Oncogênicas/agonistas , Receptores Acoplados a Proteínas G/agonistas , Transporte Ativo do Núcleo Celular , Animais , Animais Recém-Nascidos , Ácido Araquidônico/metabolismo , Tronco Encefálico/metabolismo , Tronco Encefálico/fisiopatologia , Células Cultivadas , Modelos Animais de Doenças , Endocitose , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Hipertensão/fisiopatologia , Ligantes , Neurônios/metabolismo , Óxido Nítrico/metabolismo , Fosforilação , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptores Acoplados a Proteínas G/metabolismoRESUMO
It is known that spontaneously hypertensive rats (SHR) present a marked encephalopathy, targeting vulnerable regions such as the hippocampus. Abnormalities of the hippocampus of SHR include decreased neurogenesis in the dentate gyrus (DG), partial loss of neurons in the hilus of the DG, micro and astrogliosis and inflammation. It is also known that 17ß-estradiol (E2) exert neuroprotective effects and prevent hippocampal abnormalities of SHR. The effects of E2 may involve a variety of mechanisms, including intracellular receptors of the ERα and ERß subtypes or membrane-located receptors, such as the G protein-coupled estradiol receptor (GPER). We have now investigated the protective role of GPER in SHR employing its synthetic agonist G1. To accomplish this objective, 5 month-old male SHR received 150 µg/day of G1 during 2 weeks. At the end of this period, we analyzed neuronal progenitors by staining for doublecortin (DCX), and counted the number of glial fibrillary acidic protein (GFAP)-labeled astrocytes and Iba1-stained microglial cells by computerized image analysis. We found that G1 activation of GPER increased DCX+ cells in the DG and reduced GFAP+ astrogliosis and Iba1+ microgliosis in the CA1 region of hippocampus. We also found that the high expression of proinflammatory makers IL1ß and cyclooxygenase 2 (COX2) of SHR was decreased after G1 treatment, which correlated with a change of microglia phenotype from the activated to a resting morphology. Additionally, G1 treatment increased the anti-inflammatory factor TGFß in SHR hippocampus. Altogether, our results suggest that activation of GPER plays a neuroprotective role on the encephalopathy of SHR, an outcome resembling E2 effects but avoiding secondary effects of the natural hormone.
Assuntos
Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Hipocampo/anormalidades , Hipocampo/patologia , Encefalopatia Hipertensiva/metabolismo , Inflamação/metabolismo , Neurogênese , Receptores Acoplados a Proteínas G/metabolismo , Animais , Astrócitos/metabolismo , Proteína Duplacortina , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/agonistas , Receptor beta de Estrogênio/genética , Proteína Glial Fibrilar Ácida , Encefalopatia Hipertensiva/tratamento farmacológico , Masculino , Microglia/metabolismo , Quinolinas/farmacologia , Quinolinas/uso terapêutico , Ratos , Ratos Endogâmicos SHR , Receptores de Estradiol/agonistas , Receptores de Estradiol/metabolismo , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/genéticaRESUMO
ABSTRACT Objective To investigate if ICI 182,780 (fulvestrant), a selective estrogen receptor alpha/beta (ERα/ERβ) antagonist, and G-1, a selective G-protein-coupled receptor (GPER) agonist, can potentially induce autophagy in breast cancer cell lines MCF-7 and SKBr3, and how G-1 affects cell viability. Methods Cell viability in MCF-7 and SKBr3 cells was assessed by the MTT assay. To investigate the autophagy flux, MCF-7 cells were transfected with GFP-LC3, a marker of autophagosomes, and analyzed by real-time fluorescence microscopy. MCF-7 and SKBr3 cells were incubated with acridine orange for staining of acidic vesicular organelles and analyzed by flow cytometry as an indicator of autophagy. Results Regarding cell viability in MCF-7 cells, ICI 182,780 and rapamycin, after 48 hours, led to decreased cell proliferation whereas G-1 did not change viability over the same period. The data showed that neither ICI 182,780 nor G-1 led to increased GFP-LC3 puncta in MCF-7 cells over the 4-hour observation period. The cytometry assay showed that ICI 182,780 led to a higher number of acidic vesicular organelles in MCF-7 cells. G-1, in turn, did not have this effect in any of the cell lines. In contrast, ICI 182,780 and G-1 did not decrease cell viability of SKBr3 cells or induce formation of acidic vesicular organelles, which corresponds to the final step of the autophagy process in this cell line. Conclusion The effect of ICI 182,780 on increasing acidic vesicular organelles in estrogen receptor-positive breast cancer cells appears to be associated with its inhibitory effect on estrogen receptors, and GPER does notseem to be involved. Understanding these mechanisms may guide further investigations of these receptors' involvement in cellular processes of breast cancer resistance.
RESUMO Objetivo Avaliar o efeito dos compostos ICI 182,780 (fulvestranto), um antagonista seletivo dos receptores de estrógeno alfa/beta (REα/REβ), e do G-1, um agonista seletivo de receptores de estrógeno acoplados a proteínas-G (GPER), na possível indução de autofagia em linhagens de câncer de mama MCF-7 e SKBr3, bem como o efeito de G-1 na viabilidade celular. Métodos A viabilidade celular de células MCF-7 e SKBr3 foi avaliada pelo ensaio com MTT. Para investigar a indução da autofagia, células MCF-7 foram transfectadas com GFP-LC3, um marcador de autofagossomos, e analisadas por microscopia de fluorescência em tempo real. As células MCF-7 e SKBr3 foram incubadas com o indicador de compartimentos ácidos laranja de acridina e analisadas por citometria de fluxo como indicativo para autofagia. Resultados Em células MCF-7, o ICI 182,780 e rapamicina após 48 horas levaram à diminuição da viabilidade celular, enquanto o G-1 não alterou a viabilidade no mesmo período de tratamento. Nem o ICI 182,780 e nem o G-1 induziram aumento na pontuação de GFP-LC3 em células MCF-7 até 4 horas. Já os ensaios de citometria de fluxo demonstraram que ICI 182,780 levou ao aumento de compartimentos ácidos em células MCF-7. O G-1 não aumentou estes parâmetros em ambas as linhagens. Por outro lado, ICI 182,780 e G-1 não induziram à redução da viabilidade em células SKBr3 e nem à formação de compartimentos ácidos, como etapa final do processo autofágico. Conclusão O aumento de compartimentos ácidos pelo ICI 182,780 em células de câncer de mama positivas para receptores de estrógeno parece estar associado com seu efeito inibidor de receptores de estrógeno, mas sem o envolvimento de GPER. A compreensão desses mecanismos pode direcionar estudos sobre o envolvimento dos receptores nos processos celulares de resistência do câncer de mama.