RESUMEN
Epigenetic modifications to DNA and chromatin control oncogenic and tumor-suppressive mechanisms in melanoma. Ezh2, the catalytic component of the Polycomb Repressive Complex 2 (PRC2), which mediates methylation of lysine 27 on histone 3 (H3K27me3), can regulate both melanoma initiation and progression. We previously found that mutant Ezh2Y641F interacts with the immune regulator Stat3 and together they affect anti-tumor immunity. However, given the numerous downstream targets and pathways affected by Ezh2, many mechanisms that determine its oncogenic activity remain largely unexplored. Using genetically engineered mouse models, we further investigated the role of pathways downstream of Ezh2 in melanoma carcinogenesis and identified significant enrichment in several autophagy signatures, along with increased expression of autophagy regulators, such as Atg7. In this study, we investigated the effect of Atg7 on melanoma growth and tumor immunity within the context of a wild-type or Ezh2Y641F epigenetic state. We found that the Atg7 locus is controlled by multiple Ezh2 and Stat3 binding sites, Atg7 expression is dependent on Stat3 expression, and that deletion of Atg7 slows down melanoma cell growth in vivo, but not in vitro. Atg7 deletion also results in increased CD8 + T cells in Ezh2Y641F melanomas and reduced myelosuppressive cell infiltration in the tumor microenvironment, particularly in Ezh2WT melanomas, suggesting a strong immune system contribution in the role of Atg7 in melanoma progression. These findings highlight the complex interplay between genetic mutations, epigenetic regulators, and autophagy in shaping tumor immunity in melanoma.
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Proteína 7 Relacionada con la Autofagia , Proteína Potenciadora del Homólogo Zeste 2 , Factor de Transcripción STAT3 , Animales , Factor de Transcripción STAT3/metabolismo , Ratones , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Proteína Potenciadora del Homólogo Zeste 2/genética , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Melanoma Experimental/inmunología , Melanoma Experimental/patología , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Microambiente Tumoral/inmunología , Ratones Endogámicos C57BL , Regulación Neoplásica de la Expresión Génica , Melanoma/inmunología , Melanoma/metabolismo , Melanoma/genética , Melanoma/patología , Epigénesis Genética , Línea Celular Tumoral , Humanos , Autofagia/inmunología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismoRESUMEN
Aging is a critical risk factor for heart disease, including ischemic heart disease and heart failure. Cellular senescence, characterized by DNA damage, resistance to apoptosis and the senescence-associated secretory phenotype (SASP), occurs in many cell types, including cardiomyocytes. Senescence precipitates the aging process in surrounding cells and the organ through paracrine mechanisms. Generalized autophagy, which degrades cytosolic materials in a non-selective manner, is decreased during aging in the heart. This decrease causes deterioration of cellular quality control mechanisms, facilitates aging and negatively affects lifespan in animals, including mice. Although suppression of generalized autophagy could promote senescence, it remains unclear whether the suppression of autophagy directly stimulates senescence in cardiomyocytes, which, in turn, promotes myocardial dysfunction in the heart. We addressed this question using mouse models with a loss of autophagy function. Suppression of general autophagy in cardiac-specific Atg7 knockout (Atg7cKO) mice caused accumulation of senescent cardiomyocytes. Induction of senescence via downregulation of Atg7 was also observed in chimeric Atg7 cardiac-specific KO mice and cultured cardiomyocytes in vitro, suggesting that the effect of autophagy suppression upon induction of senescence is cell autonomous. ABT-263, a senolytic agent, reduced the number of senescent myocytes and improved cardiac function in Atg7cKO mice. Suppression of autophagy and induction of senescence were also observed in doxorubicin-treated hearts, where reactivation of autophagy alleviated senescence in cardiomyocytes and cardiac dysfunction. These results suggest that suppression of general autophagy directly induces senescence in cardiomyocytes, which in turn promotes cardiac dysfunction.
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Proteína 7 Relacionada con la Autofagia , Autofagia , Senescencia Celular , Ratones Noqueados , Miocitos Cardíacos , Animales , Autofagia/genética , Senescencia Celular/efectos de los fármacos , Senescencia Celular/genética , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Ratones , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Miocardio/metabolismo , Miocardio/patología , Sulfonamidas/farmacología , Doxorrubicina/farmacología , Envejecimiento/metabolismo , Compuestos de AnilinaRESUMEN
Knee osteoarthritis (KOA) is a chronic joint disease that significantly affects the health of the elderly. As an herbal remedy, Gubi decoction (GBD) has been traditionally used for the treatment of osteoarthritis-related syndromes. However, the anti-KOA efficacy and mechanism of GBD remain unclear. This study aimed to experimentally investigate the anti-KOA efficacy and the underlying mechanism of GBD. The medial meniscus (DMM) mice model and IL-1ß-stimulated chondrocytes were, respectively, constructed as in vivo and in vitro models of KOA to evaluate the osteoprotective effect and molecular mechanism of GBD. The UPLC-MS/MS analysis showed that GBD mainly contained pinoresinol diglucoside, rehmannioside D, hesperidin, liquiritin, baohuoside I, glycyrrhizic acid, kaempferol and tangeretin. Animal experiment showed that GBD could alleviate articular cartilage destruction and recover histopathological alterations in DMM mice. In addition, GBD inhibited chondrocyte apoptosis and restored DMM-induced dysregulated autophagy evidenced by the upregulation of ATG7 and LC3 II/LC3 I but decreased P62 level. Mechanistically, METTL3-mediated m6A modification decreased the expression of ATG7 in DMM mice, as it could be significantly attenuated by GBD. METTL3 overexpression significantly counteracted the protective effect of GBD on chondrocyte autophagy. Further research showed that GBD promoted proteasome-mediated ubiquitination degradation of METLL3. Our findings suggest that GBD could act as a protective agent against KOA. The protective effect of GBD may result from its promotion on chondrocyte autophagy by suppressing METTL3-dependent ATG7 m6A methylation.
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Proteína 7 Relacionada con la Autofagia , Autofagia , Condrocitos , Metiltransferasas , Osteoartritis de la Rodilla , Animales , Condrocitos/metabolismo , Condrocitos/efectos de los fármacos , Autofagia/efectos de los fármacos , Osteoartritis de la Rodilla/metabolismo , Osteoartritis de la Rodilla/patología , Osteoartritis de la Rodilla/tratamiento farmacológico , Ratones , Proteína 7 Relacionada con la Autofagia/metabolismo , Proteína 7 Relacionada con la Autofagia/genética , Metiltransferasas/metabolismo , Metilación/efectos de los fármacos , Masculino , Medicamentos Herbarios Chinos/farmacología , Modelos Animales de Enfermedad , Apoptosis/efectos de los fármacos , Ratones Endogámicos C57BL , Adenosina/análogos & derivados , Adenosina/farmacología , Adenosina/metabolismo , Humanos , Cartílago Articular/metabolismo , Cartílago Articular/efectos de los fármacos , Cartílago Articular/patologíaRESUMEN
BACKGROUND: Prostate cancer (PCa) is a leading cause of cancer-related death in men. Understanding the proteomic landscape associated with PCa risk can provide insights into its molecular mechanisms and pave the way for potential therapeutic interventions. METHODS: A proteome-wide Mendelian randomization (MR) analysis was employed to determine associations between genetically predicted protein concentrations in plasma and PCa risk. From an initial list of 4,364 proteins, significant associations were identified and validated. Multiple sensitivity analyses were also conducted to enhance the robustness of our findings. RESULTS: Of the 4,364 genetically predicted proteins, 308 exhibited preliminary associations with PCa risk. After rigorous statistical refinement, genetically predicted concentrations of 14 proteins showed positive associations with PCa risk, with odds ratios spanning from 1.55 (95% CI 1.28-1.87) for ATG4B to 2.67 (95% CI 1.94-3.67) for HCN1. In contrast, genetically predicted concentrations of ATG7, B2M, MSMB, and TMEM108 demonstrated inverse associations with PCa. The replication analysis further substantiated positive associations for MDH1 and LSM1, and a negative one for MSMB with PCa. A meta-analysis harmonizing primary and replication data mirrored these findings. Furthermore, the MVMR analysis pinpointed B2M and MSMB as having significant associations with PCa risk. CONCLUSION: The genetic evidence unveils a refined set of proteins associated with PCa risk. The findings underscore the potential of these proteins as molecular markers or therapeutic targets for PCa, calling for deeper mechanistic studies and exploration into their translational relevance.
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Proteínas Sanguíneas , Análisis de la Aleatorización Mendeliana , Neoplasias de la Próstata , Humanos , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/sangre , Masculino , Proteínas Sanguíneas/genética , Proteína 7 Relacionada con la Autofagia/genética , Predisposición Genética a la Enfermedad , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , Proteómica/métodos , Factores de Riesgo , Proteínas de la Membrana/genética , Proteínas de la Membrana/sangre , Polimorfismo de Nucleótido Simple , Proteínas de Secreción ProstáticaRESUMEN
BACKGROUND: Cellular communication among different types of vascular cells is indispensable for maintaining vascular homeostasis and preventing atherosclerosis. However, the biological mechanism involved in cellular communication among these cells and whether this biological mechanism can be used to treat atherosclerosis remain unknown. We hypothesized that endothelial autophagy mediates the cellular communication in vascular tissue through exosome-mediated delivery of atherosclerosis-related genes. METHODS: Rapamycin and adeno-associated virus carrying Atg7 short hairpin RNA under the Tie (TEK receptor tyrosine kinase) promoter were used to activate and inhibit vascular endothelial autophagy in high-fat diet-fed ApoE-/- mice, respectively. miRNA microarray, in vivo and in vitro experiments, and human vascular tissue were used to explore the effects of endothelial autophagy on endothelial function and atherosclerosis and its molecular mechanisms. Quantitative polymerase chain reaction and miRNA sequencing were performed to determine changes in miRNA expression in exosomes. Immunofluorescence and exosome coculture experiments were conducted to examine the role of endothelial autophagy in regulating the communication between endothelial cells and smooth muscle cells (SMCs) via exosomal miRNA. RESULTS: Endothelial autophagy was inhibited in thoracic aortas of high-fat diet-fed ApoE-/- mice. Furthermore, rapamycin alleviated high-fat diet-induced atherosclerotic burden and endothelial dysfunction, while endothelial-specific Atg7 depletion aggravated the atherosclerotic burden. miRNA microarray, in vivo and in vitro experiments, and human vascular tissue analysis revealed that miR-204-5p was significantly increased in endothelial cells after high-fat diet exposure, which directly targeted Bcl2 to regulate endothelial cell apoptosis. Importantly, endothelial autophagy activation decreased excess miR-204-5p by loading miR-204-5p into multivesicular bodies and secreting it through exosomes. Moreover, exosomal miR-204-5p can effectively transport to SMCs, alleviating SMC calcification by regulating target proteins such as RUNX2 (runt-related transcription factor 2). CONCLUSIONS: Our study revealed the exosomal pathway by which endothelial autophagy protects atherosclerosis: endothelial autophagy activation transfers miR-204-5p from endothelial cells to SMCs via exosomes, both preventing endothelial apoptosis and alleviating SMC calcification. REGISTRATION: URL: https://www.chictr.org.cn/; Unique identifier: ChiCTR2200064155.
Asunto(s)
Aterosclerosis , Autofagia , Comunicación Celular , Modelos Animales de Enfermedad , Exosomas , Ratones Endogámicos C57BL , Ratones Noqueados para ApoE , MicroARNs , Miocitos del Músculo Liso , MicroARNs/metabolismo , MicroARNs/genética , Exosomas/metabolismo , Exosomas/genética , Animales , Aterosclerosis/patología , Aterosclerosis/genética , Aterosclerosis/metabolismo , Aterosclerosis/prevención & control , Humanos , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Masculino , Ratones , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/patología , Células Endoteliales/metabolismo , Células Endoteliales/patología , Proteína 7 Relacionada con la Autofagia/metabolismo , Proteína 7 Relacionada con la Autofagia/genética , Células Cultivadas , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Placa Aterosclerótica , Enfermedades de la Aorta/patología , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/prevención & control , Enfermedades de la Aorta/metabolismo , Técnicas de Cocultivo , Transducción de Señal , Aorta Torácica/metabolismo , Aorta Torácica/patología , Dieta Alta en GrasaRESUMEN
Autophagy is a self-recycling machinery to maintain cellular homeostasis by degrading harmful materials in the cell. Autophagy-related gene 5 (Atg5) is required for autophagosome maturation. However, the role of Atg5 in tumorigenesis under autophagy deficient conditions remains unclear. This study focused on the autophagy-independent role of Atg5 and the underlying mechanism in tumorigenesis. We demonstrated that knockout of autophagy-related genes including Atg5, Atg7, Atg9, and p62 in mouse embryonic fibroblast (MEF) cells consistently decreased cell proliferation and motility, implying that autophagy is required to maintain diverse cellular functions. An Atg7 knockout MEF (Atg7-/- MEF) cell line representing deprivation of autophagy function was used to clarify the role of Atg5 transgene in tumorigenesis. We found that Atg5-overexpressed Atg7-/-MEF (clone A) showed increased cell proliferation, colony formation, and migration under autophagy deficient conditions. Accordingly, rescuing the autophagy deficiency of clone A by overexpression of Atg7 gene shifts the role of Atg5 from pro-tumor to anti-tumor status, indicating the dual role of Atg5 in tumorigenesis. Notably, the xenograft mouse model showed that clone A of Atg5-overexpressed Atg7-/- MEF cells induced temporal tumor formation, but could not prolong further tumor growth. Finally, biomechanical analysis disclosed increased Wnt5a secretion and p-JNK expression along with decreased ß-catenin expression. In summary, Atg5 functions as a tumor suppressor to protect the cell under normal conditions. In contrast, Atg5 shifts to a pro-tumor status under autophagy deprivation conditions.
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Proteína 5 Relacionada con la Autofagia , Proteína 7 Relacionada con la Autofagia , Autofagia , Carcinogénesis , Proliferación Celular , Animales , Autofagia/genética , Proteína 5 Relacionada con la Autofagia/genética , Proteína 5 Relacionada con la Autofagia/metabolismo , Ratones , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Carcinogénesis/genética , Carcinogénesis/patología , Movimiento Celular/genética , Humanos , Fibroblastos/metabolismo , Ratones NoqueadosRESUMEN
Chronic psychological stress is a critical factor for neurological complications like anxiety disorders, dementia, and depression. Our previous results show that chronic restraint stress causes cognitive deficits and mood dysregulation by inducing autophagic death of adult hippocampal neural stem cells (NSCs). However, it is unknown whether other models of psychological stress also induce autophagic death of adult hippocampal NSCs. Here, we show that chronic unpredictable stress (CUS) for 10 days impaired memory function and increased anxiety in mice. Immunohistochemical staining with SOX2 and KI67 revealed a significant reduction in the number of NSCs in the hippocampus following exposure to CUS. However, these deficits were prevented by NSC-specific, inducible conditional deletion of Atg7. These findings suggest that autophagic death of adult hippocampal NSCs is a critical pathogenic mechanism underlying stress-induced brain disorders.
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Hipocampo , Células-Madre Neurales , Estrés Psicológico , Animales , Células-Madre Neurales/metabolismo , Células-Madre Neurales/patología , Hipocampo/patología , Estrés Psicológico/patología , Ratones Endogámicos C57BL , Autofagia/fisiología , Enfermedad Crónica , Proteína 7 Relacionada con la Autofagia/metabolismo , Proteína 7 Relacionada con la Autofagia/genética , Ansiedad/patología , Ansiedad/fisiopatología , Masculino , Células Madre Adultas/patología , Muerte Celular Autofágica , Memoria/fisiología , RatonesRESUMEN
BACKGROUND: Endothelial cell (EC) apoptosis and proliferation of apoptosis-resistant cells is a hallmark of pulmonary hypertension (PH). Yet, why some ECs die and others proliferate and how this contributes to vascular remodeling is unclear. We hypothesized that this differential response may: (1) relate to different EC subsets, namely pulmonary artery (PAECs) versus microvascular ECs (MVECs); (2) be attributable to autophagic activation in both EC subtypes; and (3) cause replacement of MVECs by PAECs with subsequent distal vessel muscularization. METHODS: EC subset responses to chronic hypoxia were assessed by single-cell RNA sequencing of murine lungs. Proliferative versus apoptotic responses, activation, and role of autophagy were assessed in human and rat PAECs and MVECs, and in precision-cut lung slices of wild-type mice or mice with endothelial deficiency in the autophagy-related gene 7 (Atg7EN-KO). Abundance of PAECs versus MVECs in precapillary microvessels was assessed in lung tissue from patients with PH and animal models on the basis of structural or surface markers. RESULTS: In vitro and in vivo, PAECs proliferated in response to hypoxia, whereas MVECs underwent apoptosis. Single-cell RNA sequencing analyses support these findings in that hypoxia induced an antiapoptotic, proliferative phenotype in arterial ECs, whereas capillary ECs showed a propensity for cell death. These distinct responses were prevented in hypoxic Atg7EN-KO mice or after ATG7 silencing, yet replicated by autophagy stimulation. In lung tissue from mice, rats, or patients with PH, the abundance of PAECs in precapillary arterioles was increased, and that of MVECs reduced relative to controls, indicating replacement of microvascular by macrovascular ECs. EC replacement was prevented by genetic or pharmacological inhibition of autophagy in vivo. Conditioned medium from hypoxic PAECs yet not MVECs promoted pulmonary artery smooth muscle cell proliferation and migration in a platelet-derived growth factor-dependent manner. Autophagy inhibition attenuated PH development and distal vessel muscularization in preclinical models. CONCLUSIONS: Autophagic activation by hypoxia induces in parallel PAEC proliferation and MVEC apoptosis. These differential responses cause a progressive replacement of MVECs by PAECs in precapillary pulmonary arterioles, thus providing a macrovascular context that in turn promotes pulmonary artery smooth muscle cell proliferation and migration, ultimately driving distal vessel muscularization and the development of PH.
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Apoptosis , Autofagia , Células Endoteliales , Hipertensión Pulmonar , Arteria Pulmonar , Animales , Humanos , Hipertensión Pulmonar/patología , Hipertensión Pulmonar/fisiopatología , Hipertensión Pulmonar/metabolismo , Hipertensión Pulmonar/genética , Células Endoteliales/metabolismo , Células Endoteliales/patología , Ratones , Arteria Pulmonar/patología , Arteria Pulmonar/metabolismo , Arteria Pulmonar/fisiopatología , Ratas , Proliferación Celular , Masculino , Remodelación Vascular , Ratones Noqueados , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Modelos Animales de Enfermedad , Hipoxia/metabolismo , Hipoxia/patología , Células Cultivadas , Ratones Endogámicos C57BLRESUMEN
Intervertebral disk degeneration (IDD) is a significant cause of low back pain, characterized by excessive senescence and apoptosis of nucleus pulposus cells (NPCs). However, the precise mechanisms behind this senescence and apoptosis remain unclear. This study aimed to investigate the role of T-box transcription factor T (Tbxt) in IDD both in vitro and in vivo, using a hydrogen peroxide (H2O2)-induced NPCs senescence and apoptosis model, as well as a rat acupuncture IDD model. First, the expression of p16 and cleaved-caspase 3 significantly increased in degenerated human NPCs, accompanied by a decrease in Tbxt expression. Knockdown of Tbxt exacerbated senescence and apoptosis in the H2O2-induced NPCs degeneration model. Conversely, upregulation of Tbxt alleviated these effects induced by H2O2. Mechanistically, bioinformatic analysis revealed that the direct downstream target genes of Tbxt were highly enriched in autophagy-related pathways, and overexpression of Tbxt significantly activated autophagy in NPCs. Moreover, the administration of the autophagy inhibitor, 3-methyladenine, impeded the impact of Tbxt on the processes of senescence and apoptosis in NPCs. Further investigation revealed that Tbxt enhances autophagy by facilitating the transcription of ATG7 through its interaction with a specific motif within the promoter region. In conclusion, this study suggests that Tbxt mitigates H2O2-induced senescence and apoptosis of NPCs by activating ATG7-mediated autophagy.NEW & NOTEWORTHY This study investigates the role of Tbxt in IDD. The results demonstrate that knockdown of Tbxt exacerbates H2O2-induced senescence and apoptosis in NPCs and IDD, whereas upregulation of Tbxt significantly protects against IDD both in vivo and in vitro. Mechanistically, in the nucleus, Tbxt enhances the transcription of ATG7, leading to increased expression of ATG7 protein levels. This, in turn, promotes elevated autophagy levels, ultimately alleviating IDD.
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Apoptosis , Proteína 7 Relacionada con la Autofagia , Autofagia , Senescencia Celular , Degeneración del Disco Intervertebral , Núcleo Pulposo , Ratas Sprague-Dawley , Núcleo Pulposo/metabolismo , Núcleo Pulposo/patología , Autofagia/efectos de los fármacos , Degeneración del Disco Intervertebral/patología , Degeneración del Disco Intervertebral/metabolismo , Degeneración del Disco Intervertebral/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Proteína 7 Relacionada con la Autofagia/genética , Animales , Senescencia Celular/efectos de los fármacos , Humanos , Ratas , Masculino , Femenino , Adulto , Persona de Mediana Edad , Peróxido de Hidrógeno/toxicidad , Peróxido de Hidrógeno/farmacología , Peróxido de Hidrógeno/metabolismo , Células CultivadasRESUMEN
BACKGROUND: Myocardial infarction (MI) is an acute condition in which the heart muscle dies due to the lack of blood supply. Previous research has suggested that autophagy and angiogenesis play vital roles in the prevention of heart failure after MI, and miR-106a is considered to be an important regulatory factor in MI. But the specific mechanism remains unknown. In this study, using cultured venous endothelial cells and a rat model of MI, we aimed to identify the potential target genes of miR-106a and discover the mechanisms of inhibiting autophagy and angiogenesis. METHODS: We first explored the biological functions of miR-106a on autophagy and angiogenesis on endothelial cells. Then we identified ATG7, which was the downstream target gene of miR-106a. The expression of miR-106a and ATG7 was investigated in the rat model of MI. RESULTS: We found that miR-106a inhibits the proliferation, cell cycle, autophagy and angiogenesis, but promoted the apoptosis of vein endothelial cells. Moreover, ATG7 was identified as the target of miR-106a, and ATG7 rescued the inhibition of autophagy and angiogenesis by miR-106a. The expression of miR-106a in the rat model of MI was decreased but the expression of ATG7 was increased in the infarction areas. CONCLUSION: Our results indicate that miR-106a may inhibit autophagy and angiogenesis by targeting ATG7. This mechanism may be a potential therapeutic treatment for MI.
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Proteína 7 Relacionada con la Autofagia , Autofagia , MicroARNs , Infarto del Miocardio , Animales , MicroARNs/genética , MicroARNs/metabolismo , Infarto del Miocardio/genética , Infarto del Miocardio/metabolismo , Infarto del Miocardio/patología , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Ratas , Masculino , Neovascularización Fisiológica , Ratas Sprague-Dawley , Humanos , Células Endoteliales de la Vena Umbilical Humana , Proliferación Celular , Células Endoteliales/metabolismo , Apoptosis , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Modelos Animales de Enfermedad , Células Cultivadas , AngiogénesisRESUMEN
Macrophages (Mφ) autophagy is a pivotal contributor to inflammation-related diseases. However, the mechanistic details of its direct role in acute kidney injury (AKI) were unclear. Here, we show that Mφ promote AKI progression via crosstalk with tubular epithelial cells (TECs), and autophagy of Mφ was activated and then inhibited in cisplatin-induced AKI mice. Mφ-specific depletion of ATG7 (Atg7Δmye) aggravated kidney injury in AKI mice, which was associated with tubulointerstitial inflammation. Moreover, Mφ-derived exosomes from Atg7Δmye mice impaired TEC mitochondria in vitro, which may be attributable to miR-195a-5p enrichment in exosomes and its interaction with SIRT3 in TECs. Consistently, either miR-195a-5p inhibition or SIRT3 overexpression improved mitochondrial bioenergetics and renal function in vivo. Finally, adoptive transfer of Mφ from AKI mice to Mφ-depleted mice promotes the kidney injury response to cisplatin, which is alleviated when Mφ autophagy is activated with trehalose. We conclude that exosomal miR-195a-5p mediate the communication between autophagy-deficient Mφ and TECs, leading to impaired mitochondrial biogenetic in TECs and subsequent exacerbation of kidney injury in AKI mice via miR-195a-5p-SIRT3 axis.
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Lesión Renal Aguda , Autofagia , Cisplatino , Macrófagos , MicroARNs , Mitocondrias , Sirtuina 3 , Animales , Humanos , Masculino , Ratones , Lesión Renal Aguda/metabolismo , Lesión Renal Aguda/genética , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/patología , Autofagia/efectos de los fármacos , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Cisplatino/farmacología , Modelos Animales de Enfermedad , Células Epiteliales/metabolismo , Células Epiteliales/efectos de los fármacos , Exosomas/metabolismo , Riñón/patología , Riñón/metabolismo , Túbulos Renales/patología , Túbulos Renales/metabolismo , Macrófagos/metabolismo , Macrófagos/efectos de los fármacos , Ratones Endogámicos C57BL , MicroARNs/genética , MicroARNs/metabolismo , Mitocondrias/metabolismo , Mitocondrias/efectos de los fármacos , Sirtuina 3/metabolismo , Sirtuina 3/genética , Trehalosa/farmacologíaRESUMEN
Circular RNA (circRNA) has emerged as a pivotal regulatory factor in cancer biology, yet its exact role in cervical cancer remains incompletely understood. In this study, we investigated the functional role of circCUL3 in cervical cancer and explored its potential as a therapeutic target. Functional gain and loss experiments were conducted in Hela and Siha cell lines to elucidate the biological functions of circCUL3 in cervical cancer. The results revealed that circCUL3 overexpression significantly enhanced cell viability, migration, and invasion while suppressing apoptosis, while circCUL3 knockout displayed the opposite effects. Mechanistically, we identified hsa-miR-223-3p as a target of circCUL3, with its expression being negatively regulated by circCUL3. Furthermore, we discovered that circCUL3 could sequester miR-223-3p, leading to the upregulation of ATG7 expression, and this was linked to the regulation of autophagy in cervical cancer cells. In vivo validation using a xenograft mouse model further supported our in vitro findings. Notably, we found that chloroquine (CQ), an autophagy inhibitor, restored miR-223-3p expression and counteracted the oncogenic effect of circCUL3 overexpression. In conclusion, circCUL3 potentially contributes to the malignant progression of cervical cancer by acting as a sponge for miR-223-3p, resulting in the upregulation of ATG7 and the activation of autophagy.
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Proteína 7 Relacionada con la Autofagia , Autofagia , Regulación Neoplásica de la Expresión Génica , MicroARNs , ARN Circular , Regulación hacia Arriba , Neoplasias del Cuello Uterino , Animales , Femenino , Humanos , Ratones , Apoptosis/genética , Autofagia/genética , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Progresión de la Enfermedad , Células HeLa , Ratones Desnudos , MicroARNs/genética , MicroARNs/metabolismo , ARN Circular/genética , ARN Circular/metabolismo , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patología , Neoplasias del Cuello Uterino/metabolismoRESUMEN
Rac1 is a key regulator of the cytoskeleton and neuronal plasticity, and is known to play a critical role in psychological and cognitive brain disorders. To elucidate the engram specific Rac1 signaling in fear memory, a doxycycline (Dox)-dependent robust activity marking (RAM) system was used to label dorsal dentate gyrus (DG) engram cells in mice during contextual fear conditioning. Rac1 mRNA and protein levels in DG engram cells were peaked at 24 h (day 1) after fear conditioning and were more abundant in the fear engram cells than in the non-engram cells. Optogenetic activation of Rac1 in a temporal manner in DG engram cells before memory retrieval decreased the freezing level in the fear context. Optogenetic activation of Rac1 increased autophagy protein 7 (ATG7) expression in the DG engram cells and activated DG microglia. Microglia-specific transcriptomics and fluorescence in situ hybridization revealed that overexpression of ATG7 in the fear engram cells upregulated the mRNA of Toll-like receptor TLR2/4 in DG microglia. Knockdown of microglial TLR2/4 rescued fear memory destabilization induced by ATG7 overexpression or Rac1 activation in DG engram cells. These results indicate that Rac1-driven communications between engram cells and microglia contributes to contextual fear memory destabilization, and is mediated by ATG7 and TLR2/4, and suggest a novel mechanistic framework for the cytoskeletal regulator in fear memory interference.
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Giro Dentado , Miedo , Hipocampo , Memoria , Microglía , Optogenética , Proteína de Unión al GTP rac1 , Animales , Miedo/fisiología , Ratones , Proteína de Unión al GTP rac1/metabolismo , Memoria/fisiología , Microglía/metabolismo , Hipocampo/metabolismo , Giro Dentado/metabolismo , Masculino , Ratones Endogámicos C57BL , Proteína 7 Relacionada con la Autofagia/metabolismo , Proteína 7 Relacionada con la Autofagia/genética , Neuropéptidos/metabolismo , Plasticidad Neuronal/fisiología , Receptor Toll-Like 2/metabolismo , Receptor Toll-Like 2/genética , Receptor Toll-Like 4/metabolismo , Condicionamiento Clásico/fisiologíaRESUMEN
PURPOSE: In this study, the effect of thymoquinone (TQ) on CP-induced spermatogenesis defects in mice has been investigated. METHODS: Sperm parameters, serum testosterone concentration, histology, Bax/Bcl-2 ratio, and expression of autophagy-related biomarkers have been assessed. Total antioxidant capacity (TAC), total oxidant status (TOS), and oxidative stress index (OSI) in testicular tissue were examined for the evaluation of oxidative stress levels. RESULTS: CP has induced histological changes and significantly increased the Bax/Bcl-2 ratio, decreased testosterone concentration, testicular weight, and sperm quality. CP induced oxidative stress by elevating OSI in the testicular tissue (p < 0.05). Expression of the autophagy-inducer genes (ATG7, ATG5, and Beclin-1) and ratio of LC3B/LC3A proteins were significantly decreased, while mTOR expression was increased in the CP group. TQ pretreatment dose-dependently decreased the Bax/Bcl-2 ratio and mTOR gene expression while increasing the expression of ATG5 and ATG7 genes, LC3B/LC3A ratio, and Beclin-1 proteins. TQ could also dose-dependently reverse the histology, testosterone level, and sperm quality of the CP-intoxicated mice. CONCLUSIONS: These findings show that TQ pretreatment can enhance sperm production by inducing autophagy and reducing apoptosis and oxidative stress in the CP-intoxicated mouse testicles.
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Apoptosis , Autofagia , Benzoquinonas , Cisplatino , Estrés Oxidativo , Espermatozoides , Testículo , Masculino , Animales , Estrés Oxidativo/efectos de los fármacos , Ratones , Autofagia/efectos de los fármacos , Testículo/efectos de los fármacos , Testículo/metabolismo , Testículo/patología , Apoptosis/efectos de los fármacos , Benzoquinonas/farmacología , Cisplatino/efectos adversos , Cisplatino/farmacología , Espermatozoides/efectos de los fármacos , Espermatozoides/metabolismo , Espermatozoides/patología , Testosterona/sangre , Espermatogénesis/efectos de los fármacos , Antioxidantes/farmacología , Antioxidantes/metabolismo , Beclina-1/genética , Beclina-1/metabolismo , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismoRESUMEN
PURPOSE: To explore the mechanism underlying autophagy disruption in gingival epithelial cells (GECs) in diabetic individuals. METHODS AND MATERIALS: Bone marrow-derived macrophages (BMDMs) and GECs were extracted from C57/bl and db/db mice, the exosomes (Exo) were isolated from BMDMs. qRTâPCR and Western blotting were performed to analyse gene expression. The AnimalTFDB database was used to identify relevant transcription factors, and miRNA sequencing was utilised to identify relevant miRNAs with the aid of the TargetScan/miRDB/miRWalk databases. A dual-luciferase assay was conducted to verify intermolecular targeting relationships. RESULTS: Similar to BMDMs, BMDM-derived Exos disrupted autophagy and exerted proinflammatory effects in GEC cocultures, and ATG7 may play a vital role. AnimalTFDB database analysis and dual-luciferase assays indicated that NR5A2 is the most relevant transcription factor that regulates Atg7 expression. SiRNA-NR5A2 transfection blocked autophagy in GECs and exacerbated inflammation, whereas NR5A2 upregulation restored ATG7 expression and ameliorated ExoDM-mediated inflammation. MiRNA sequencing, with TargetScan/miRDB/miRWalk analyses and dual-luciferase assays, confirmed that miR-381-3p is the most relevant miRNA that targets NR5A2. MiR-381-3p mimic transfection blocked autophagy in GECs and exacerbated inflammation, while miR-381-3p inhibitor transfection restored ATG7 expression and attenuated ExoDM-mediated inflammation. CONCLUSION: BMDM-derived Exos, which carry miR-381-3p, inhibit NR5A2 and disrupt autophagy in GECs, increasing periodontal inflammation in diabetes.
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Autofagia , Células Epiteliales , Exosomas , Encía , Macrófagos , Ratones Endogámicos C57BL , MicroARNs , Animales , Masculino , Ratones , Autofagia/genética , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Células Epiteliales/metabolismo , Exosomas/metabolismo , Encía/citología , Encía/metabolismo , Macrófagos/metabolismo , MicroARNs/genética , MicroARNs/metabolismoRESUMEN
OBJECTIVE: Maternal obesity affects 39.7% of reproductive-age women in the United States. Emerging research has suggested that in utero exposure to maternal obesity is associated with adverse neurodevelopmental outcomes, but knowledge of underlying mechanisms in human samples is lacking. METHODS: A matched case-control study was performed in women with singleton fetuses who were undergoing elective pregnancy termination at gestational ages 15 to 21 weeks. Maternal adiponectin levels from plasma were measured using ELISA kits. RNA was extracted from fetal brain tissue using RNeasy Mini Kit (QIAGEN). mRNA expression from ADIPOR1, ADIPOR2, MTOR, ATG5, ATG7, BECN1, and MAP1LC3B was quantified through the ΔΔCt method and using GAPDH as a housekeeping gene. RESULTS: We have identified transcription patterns associated with inhibition of autophagy in male fetal brain tissue exposed to maternal obesity (↑MTOR, ↓ATG5, ↓ATG7, and ↓MAP1LC3B), with female fetuses demonstrating either no change in transcription or nonsignificant changes associated with increased autophagy. There was significant downregulation of the autophagy-associated gene BECN1 in both male and female individuals who were exposed to obesity in utero. CONCLUSIONS: We present novel evidence suggesting that in utero exposure to maternal obesity in humans may significantly affect neurodevelopment, especially in male fetuses, through alterations in normal autophagy molecular mechanisms and with adiponectin as a potential mediator.
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Adiponectina , Autofagia , Beclina-1 , Encéfalo , Proteínas Asociadas a Microtúbulos , Obesidad Materna , Serina-Treonina Quinasas TOR , Humanos , Femenino , Embarazo , Masculino , Estudios de Casos y Controles , Obesidad Materna/metabolismo , Encéfalo/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Adiponectina/metabolismo , Adiponectina/sangre , Beclina-1/metabolismo , Adulto , Proteínas Asociadas a Microtúbulos/metabolismo , Proteína 5 Relacionada con la Autofagia/metabolismo , Proteína 5 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Receptores de Adiponectina/metabolismo , Receptores de Adiponectina/genética , Feto/metabolismo , ARN Mensajero/metabolismo , Factores Sexuales , Edad Gestacional , Regulación hacia Abajo , Obesidad/metabolismoRESUMEN
BACKGROUND: The efficacy of anti-PD-1 therapy is primarily hindered by the limited T-cell immune response rate and immune evasion capacity of tumor cells. Autophagy-related protein 7 (ATG7) plays an important role in autophagy and it has been linked to cancer. However, the role of ATG7 in the effect of immune checkpoint blockade (ICB) treatment on high microsatellite instability (MSI-H)/mismatch repair deficiency (dMMR) CRC is still poorly understood. METHODS: In this study, patients from the cancer genome altas (TCGA) COAD/READ cohorts were used to investigate the biological mechanism driving ATG7 development. Several assays were conducted including the colony formation, cell viability, qRT-PCR, western blot, immunofluorescence, flow cytometry, ELISA, immunohistochemistry staining and in vivo tumorigenicity tests. RESULTS: We found that ATG7 plays a crucial role in MSI-H CRC. Its knockdown decreased tumor growth and caused an infiltration of CD8+ T effector cells in vivo. ATG7 inhibition restored surface major histocompatibility complex I (MHC-I) levels, causing improved antigen presentation and anti-tumor T cell response by activating reactive oxygen species (ROS)/NF-κB pathway. Meanwhile, ATG7 inhibition also suppressed cholesterol accumulation and augmentation of anti-tumor immune responses. Combining ATG7 inhibition and statins improved the therapeutic benefit of anti-PD-1 in MSI-H CRC. Importantly, CRC patients with high expression of both ATG7 and recombinant 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) experienced worse prognosis compared to those with low ATG7 and HMGCR expression. CONCLUSIONS: Inhibition of ATG7 leads to upregulation of MHC-I expression, augments immune response and suppresses cholesterol accumulation. These findings demonstrate that ATG7 inhibition has therapeutic potential and application of statins can increase the sensitivity to immune checkpoint inhibitors.
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Neoplasias Encefálicas , Neoplasias Colorrectales , Inhibidores de Hidroximetilglutaril-CoA Reductasas , Síndromes Neoplásicos Hereditarios , Humanos , Proteína 7 Relacionada con la Autofagia/genética , Colesterol , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Reparación de la Incompatibilidad de ADN , Inhibidores de Hidroximetilglutaril-CoA Reductasas/uso terapéutico , Inhibidores de Puntos de Control Inmunológico/farmacología , Inmunidad , Inestabilidad de MicrosatélitesRESUMEN
Obesity stands as a significant risk factor for type 2 diabetes, hyperlipidemia, and cardiovascular diseases, intertwining increased inflammation and decreased adipogenesis with metabolic disorders. Studies have highlighted the correlation between Caspase-1 and inflammation in obesity, elucidating its essential role in the biological functions of adipose tissue. However, the impact of Caspase-1 on adipogenesis and the underlying mechanisms remain largely elusive. In our study, we observed a positive correlation between Caspase-1 expression and obesity and its association with adipogenesis. In vivo experiments revealed that, under normal diet conditions, Caspase-1 deficiency improved glucose homeostasis, stimulated subcutaneous adipose tissue expansion, and enhanced adipogenesis. Furthermore, our findings indicate that Caspase-1 deficiency promotes the expression of autophagy-related proteins and inhibits autophagy with 3-MA or CQ blocked Caspase-1 deficiency-induced adipogenesis in vitro. Notably, Caspase-1 deficiency promotes adipogenesis via Atg7-mediated autophagy activation. In addition, Caspase-1 deficiency resisted against high-fat diet-induced obesity and glucose intolerance. Our study proposes the downregulation of Caspase-1 as a promising strategy for mitigating obesity and its associated metabolic disorders.
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Adipogénesis , Proteína 7 Relacionada con la Autofagia , Autofagia , Caspasa 1 , Inflamación , Obesidad , Adipogénesis/genética , Animales , Proteína 7 Relacionada con la Autofagia/genética , Proteína 7 Relacionada con la Autofagia/metabolismo , Ratones , Caspasa 1/metabolismo , Caspasa 1/genética , Caspasa 1/deficiencia , Obesidad/metabolismo , Obesidad/patología , Obesidad/genética , Inflamación/metabolismo , Inflamación/patología , Inflamación/genética , Masculino , Dieta Alta en Grasa/efectos adversos , Ratones Endogámicos C57BL , Células 3T3-L1 , Ratones NoqueadosRESUMEN
Autophagy is a key step involved in many unicellular eukaryotic diseases, including leishmaniasis, for cellular remodelling and differentiation during parasite's lifecycle. Lipids play a significant role in the infection process that begins with Leishmania major invading host cells. MicroRNAs (miRNAs), a family of small, 22-24 nucleotide noncoding regulatory RNAs, target mRNAs to modify gene expression and, subsequently, proteome output may have a regulatory role in altering the host cell processes. We observed miR-146a-3p expression increases in a time-dependent manner post Leishmania major infection. Transfecting miR-146a-3p mimic increases the expression of ATG7, an autophagy gene that encodes an E1-like enzyme in two ubiquitin-like conjugation systems required for autophagosome progression. HPGD (15-hydroxyprostaglandin dehydrogenase) operates as an enzyme, converting prostaglandin to its non-active form. Microarray data and western studies reveal that miR-146a-3p targets and inhibits HPGD, thereby increasing prostaglandin activity in lipid droplets. Herein, our research focuses on miR-146a-3p, which boosts ATG7 expression while reducing HPGD post Leishmania major infections helping us comprehend the intricate network of microRNA, autophagy, and lipid metabolism in leishmaniasis.
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Autofagia , Leishmania major , Leishmaniasis Cutánea , Metabolismo de los Lípidos , MicroARNs , MicroARNs/metabolismo , MicroARNs/genética , Leishmania major/genética , Leishmania major/fisiología , Leishmania major/metabolismo , Leishmaniasis Cutánea/parasitología , Animales , Ratones , Proteína 7 Relacionada con la Autofagia/metabolismo , Proteína 7 Relacionada con la Autofagia/genética , Ratones Endogámicos BALB C , Macrófagos/parasitología , Macrófagos/metabolismo , Humanos , Transfección , Western BlottingRESUMEN
Sunlight exposure results in an inflammatory reaction of the skin commonly known as sunburn, which increases skin cancer risk. In particular, the ultraviolet B (UVB) component of sunlight induces inflammasome activation in keratinocytes to instigate the cutaneous inflammatory responses. Here, we explore the intracellular machinery that maintains skin homeostasis by suppressing UVB-induced inflammasome activation in human keratinocytes. We found that pharmacological inhibition of autophagy promoted UVB-induced NLRP3 inflammasome activation. Unexpectedly, however, gene silencing of Atg5 or Atg7, which are critical for conventional autophagy, had no effect, whereas gene silencing of Beclin1, which is essential not only for conventional autophagy but also for Atg5/Atg7-independent alternative autophagy, promoted UVB-induced inflammasome activation, indicating an involvement of alternative autophagy. We found that damaged mitochondria were highly accumulated in UVB-irradiated keratinocytes when alternative autophagy was inhibited, and they appear to be recognized by NLRP3. Overall, our findings indicate that alternative autophagy, rather than conventional autophagy, suppresses UVB-induced NLRP3 inflammasome activation through the clearance of damaged mitochondria in human keratinocytes and illustrate a previously unknown involvement of alternative autophagy in inflammation. Alternative autophagy may be a new therapeutic target for sunburn and associated cutaneous disorders.