RESUMEN
RB transcriptional corepressor 1 (RB) deletion is the most important genomic factor associated with the prognosis of castration-resistant prostate cancer (CRPC) patients receiving androgen receptor (AR) signaling inhibitor therapy. Loss of RB could support prostate cancer cell growth in a hormone-independent manner, but the underlying mechanism by which RB regulates tumor progression extends far beyond the cell cycle pathway. A previous study indicated that RB inactivates AKT signaling but has no effect on mTOR signaling in cancer cells. Here, we found that the S249/T252 site in RB is key to regulating the transcriptional activity of the tumor-promoting factor TRIM24 in CRPC, as identified through FXXXV mapping. The RB/TRIM24 complex functions through DUSP2, which serves as an intermediate bridge, to activate the mTOR pathway and promote prostate cancer progression. Accordingly, we designed RB-linker-proteolysis-targeting chimera (PROTAC) molecules, which decreased TRIM24 protein levels and inactivated the mTOR signaling pathway, thereby inhibiting prostate cancer. Therefore, this study not only elucidates the novel function of RB but also provides a theoretical basis for the development of new drugs for treating prostate cancer.
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Proteína de Retinoblastoma , Transducción de Señal , Serina-Treonina Quinasas TOR , Animales , Humanos , Masculino , Ratones , Proteínas Portadoras/metabolismo , Línea Celular Tumoral , Proliferación Celular , Ratones Desnudos , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Neoplasias de la Próstata/genética , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/patología , Neoplasias de la Próstata Resistentes a la Castración/genética , Proteína de Retinoblastoma/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Fosfatasa 2 de Especificidad Dual/metabolismoRESUMEN
BACKGROUND: As one of the leading causes of cancer death worldwide, bladder cancer (BCa) ranks 12th in incidence rate. Dual Specific Phosphatase 2 (DUSP2) is a member of the bispecific protein phosphatase subfamily. DUSP2 is closely related to the prognosis of cancer, but the role of DUSP2 in bladder cancer is still unclear. This study aims to explore how DUSP2 affects the prognosis of bladder cancer and clarify the important mechanism in bladder cancer. METHODS: Bioinformatics and experiments have detected the anti-tumor effect of DUSP2. Construct a DUSP2 overexpression cell model, and then use protein blotting experiments to verify the efficiency of transfection. The effects of DUSP2 on proliferation, metastasis, apoptosis, epithelial mesenchymal transition (EMT) and immune invasion of bladder cancer cells were detected in vitro or in vivo. In addition, the mechanism of DUSP2 regulating MEK/ERK through PTPN7 pathway and P38 MAPK inhibiting the progression of bladder cancer was also discussed. RESULTS: The expression of DUSP2 was down regulated in bladder cancer samples and cell lines. The overexpression of DUSP2 inhibits the proliferation, metastasis and immune microenvironment of bladder cancer cells. In addition, we confirmed that DUSP2 regulates MEK/ERK and P38 MAPK through PTPN7 pathway to inhibit the progression of bladder cancer. CONCLUSION: DUSP2 inhibits the progression of bladder cancer by regulating PTPN7. These results suggest that DUSP2/PTPN7/MEK/ERK pathway may become a new therapeutic target for bladder cancer.
Asunto(s)
Sistema de Señalización de MAP Quinasas , Neoplasias de la Vejiga Urinaria , Humanos , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Vejiga Urinaria/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proliferación Celular/genética , Línea Celular Tumoral , Movimiento Celular , Microambiente Tumoral , Proteínas Tirosina Fosfatasas no Receptoras , Fosfatasa 2 de Especificidad Dual/metabolismoRESUMEN
Pancreatic ductal adenocarcinoma (PDAC) is characterized by hypoxic tumor microenvironment (TME), which aids tumor progression, drug resistance, and immune evasion. Dual-specificity phosphatase 2 (DUSP2), a member of the mitogen-activated protein kinase phosphatase family, regulates pancreatic cancer metastasis. However, its role in the hypoxic TME in PDAC remains unknown. We explored the role of DUSP2 by simulating the hypoxic TME. DUSP2 significantly promoted apoptosis in PDAC both in vitro and in vivo, mainly through AKT1 rather than ERK1/2. Mechanistically, DUSP2 competed with AKT1 to bind to casein kinase 2 alpha 1 (CSNK2A1) and inhibited the phosphorylation of AKT1, which plays a crucial role in apoptosis resistance. Interestingly, aberrant activation of AKT1 resulted in an increase in the ubiquitin E3 ligase tripartite motif-containing 21 (TRIM21), which binds to and mediates the ubiquitination-dependent proteasomal degradation of DUSP2. Overall, we identified CSNK2A1 as a novel binding partner of DUSP2 that promotes PDAC apoptosis through CSN2KA1/AKT1 in an ERK1/2-independent manner. Activation of AKT1 also mediated proteasomal degradation of DUSP2 via the AKT1/TRIM21 positive feedback loop. We propose increasing the level of DUSP2 as a potential therapeutic strategy for PDAC.
Asunto(s)
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Fosfatasa 2 de Especificidad Dual/metabolismo , Neoplasias Pancreáticas/patología , Hipoxia , Carcinoma Ductal Pancreático/patología , Apoptosis , Línea Celular Tumoral , Microambiente Tumoral , Proteínas Proto-Oncogénicas c-akt , Neoplasias PancreáticasRESUMEN
Rationale: Acute kidney injury (AKI) is pathologically characterized by renal tubular epithelial cell (RTEC) death and interstitial inflammation, while their pathogenesis remains incompletely understood. Dual-specificity phosphatase 2 (DUSP2) recently emerges as a crucial regulator of cell death and inflammation in a wide range of diseases, but its roles in renal pathophysiology are largely unknown. Methods: The expression of DUSP2 in the kidney was characterized by histological analysis in renal tissues from patients and mice with AKI. The role and mechanism of DUSP2-mediated inhibition of tubular epithelial cell pyroptosis in AKI were evaluated both in vivo and in vitro, and confirmed in RTEC-specific deletion of DUSP2 mice. Results: Here, we show that DUSP2 is enriched in RTECs in the renal tissue of both human and mouse and mainly positions in the nucleus. Further, we reveal that loss-of-DUSP2 in RTECs not only is a common feature of human and murine AKI but also positively contributes to AKI pathogenesis. Especially, RTEC-specific deletion of DUSP2 sensitizes mice to AKI by promoting RTEC pyroptosis and the resultant interstitial inflammation. Mechanistic studies show that gasdermin D (GSDMD), which mediates RTEC pyroptosis, is identified as a transcriptional target of activated STAT1 during AKI, whereas DUSP2 as a nuclear phosphatase deactivates STAT1 to restrict GSDMD-mediated RTEC pyroptosis. Importantly, DUSP2 overexpression in RTECs via adeno-associated virus-mediated gene transfer significantly ameliorates AKI. Conclusion: Our findings demonstrate a hitherto unrecognized role of DUSP2-STAT1 axis in regulating RTEC pyroptosis in AKI, highlighting that DUSP2-STAT1 axis is an attractive therapeutic target for AKI.
Asunto(s)
Lesión Renal Aguda , Piroptosis , Lesión Renal Aguda/metabolismo , Animales , Fosfatasa 2 de Especificidad Dual/metabolismo , Células Epiteliales/metabolismo , Humanos , Inflamación/metabolismo , Ratones , Ratones Endogámicos C57BL , Piroptosis/fisiologíaRESUMEN
BACKGROUND: Distant metastasis remains the leading cause of treatment failure in patients with nasopharyngeal carcinoma (NPC), making it critical to identify efficient therapeutic targets for metastatic NPC. Previous studies have demonstrated that deoxynucleotidyltransferase terminal-interacting protein 1 (DNTTIP1) is associated with the development of various types of cancer. However, its role and mechanism in NPC have not been explored. METHODS: RNA-seq profiling was performed for three pairs of NPC and normal nasopharynx tissues. DNTTIP1 expression in NPC specimens was detected by immunohistochemistry. In vitro and in vivo assays were used to investigate the function of DNTTIP1. The molecular mechanism was determined using RT-qPCR, western blotting, RNA-seq, luciferase reporter assays, ChIP assays, and co-IP assays. FINDINGS: DNTTIP1 was found to be significantly upregulated in NPC tissues. Furthermore, DNTTIP1 promoted NPC growth and metastasis in vitro and in vivo. Upregulation of DNTTIP1 in NPC indicated poor clinical outcomes. Mechanistically, DNTTIP1 suppressed DUSP2 gene expression via recruiting HDAC1 to its promoter and maintaining a deacetylated state of histone H3K27. The downregulation of DUSP2 resulted in aberrant activation of the ERK signaling and elevated MMP2 levels, promoting NPC metastasis. Chidamide, an HDAC inhibitor, was shown to suppress NPC metastasis by regulating the DNTTIP1/HDAC1-DUSP2 axis. INTERPRETATION: Our findings demonstrate that DNTTIP1 not only regulates NPC metastasis but also independently predicts NPC prognosis. Furthermore, targeting DNTTIP1/HDAC1 by Chidamide may benefit NPC patients with metastasis. FUNDING: This work was supported by the National Natural Science Foundation of China (No. 81872464, 82073243).
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Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas , Línea Celular Tumoral , Movimiento Celular/fisiología , Proliferación Celular , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Fosfatasa 2 de Especificidad Dual/genética , Fosfatasa 2 de Especificidad Dual/metabolismo , Histona Desacetilasa 1/genética , Histona Desacetilasa 1/metabolismo , Humanos , Carcinoma Nasofaríngeo/genética , Carcinoma Nasofaríngeo/metabolismo , Carcinoma Nasofaríngeo/patología , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/patología , Invasividad Neoplásica , Metástasis de la Neoplasia , Transducción de Señal , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer characterized by early dissemination and poor drug response. Therefore, it is an unmet medical need to develop new strategies for treatment. As aberrant activation of ERK due to KRAS activating mutation is a driving force for PDAC, a brake system that can terminate ERK signaling represents an ideal druggable target. Herein, we demonstrate that forced expression of dual specificity phosphatase-2 (DUSP2), a specific ERK phosphatase, abrogated tumor formation and loss of Dusp2 facilitated Kras-driven PDAC progression. We report that a selective HDAC1/2 inhibitor (B390) has multifaceted therapeutic potential in PDAC by restoring the expression and function of DUSP2. In vitro study showed that treatment with B390 inhibited growth and migration abilities of PDAC cells, decreased extracellular vesicle-associated VEGF-C expression, and suppressed lymphatic endothelial cell proliferation. In vivo, B390 not only suppressed tumor growth by increasing tumor cell death, it also inhibited lymphangiogenesis and lymphovascular invasion. Taken together, our data demonstrate that B390 was able to alleviate loss of DUSP2-mediated pathologic processes, which provides the proof-of-concept evidence to demonstrate the potential of using selective HDAC1/2 inhibitors in PDAC treatment and suggests reinstating DUSP2 expression may be a strategy to subside PDAC progression.
Asunto(s)
Fosfatasa 2 de Especificidad Dual/metabolismo , Histona Desacetilasa 1/antagonistas & inhibidores , Histona Desacetilasa 2/antagonistas & inhibidores , Inhibidores de Histona Desacetilasas/farmacología , Linfangiogénesis , Neoplasias Pancreáticas/tratamiento farmacológico , Factor C de Crecimiento Endotelial Vascular/metabolismo , Animales , Apoptosis , Carcinoma Ductal Pancreático/tratamiento farmacológico , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patología , Proliferación Celular , Fosfatasa 2 de Especificidad Dual/genética , Vesículas Extracelulares/metabolismo , Humanos , Ratones , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Células Tumorales Cultivadas , Factor C de Crecimiento Endotelial Vascular/genéticaRESUMEN
To investigate the impact of different anticoagulants and coagulants with autologous platelet-rich plasma (PRP) in order to evaluate the clinical application of PRP standardization. Bone marrow stem cells (BMSCs) were seeded into autologous PRP gel scaffolds with different anticoagulants (EDTA, heparin sodium HS, and sodium citrate SC) as well as control group (the whole blood group). Quality of PRP was evaluated and flow cytometric assay was used to detect the activity of the platelet (CD62p, PAC-1). BMSCs were also seeded into PRP with different coagulants (Thrombin, Collagen-I, ADP) as well as PRP un-activated (negative group) and L-DMEM complete culture without PRP (control group). The effects of different coagulants with PRP on proliferation, osteogenic differentiation of BMSCs were analyzed by methyl thiazolyl tetrazolium assay (MTT), ALP staining, Von Kossa staining, Confocal microscopic observation, RT-PCR and Western Blot at the morphological, cellular and molecular levels. Different anticoagulants (EDTA, HS, and SC) could affect the quality of PRP. EDTA group revealed the best quality and activity (CD62p, PAC-1). With different coagulants (Thrombin, Collagen-I and ADP) in the proliferation of BMSCs, the MTT assay showed that the proliferation of BMSCs was increased in all groups with time. On the sixth day of culture, the cell number of each PRP group was significantly higher than that in the control group (P < 0.05), while the most rapidly increasing was found in Collagen-I group. The cumulative release of growth factor (TGF-ß1, PDGF) at each time point in the PRP gel of the four groups was higher than that in the control group (P < 0.05). Collagen-I was considered as the best PRP coagulant. When thrombin was used as a platelet coagulant, the release of growth factor in PRP was rapid and direct, while the release of growth factor in Collagen-I-activated PRP was sustained and slow, and the total release of ADP-activated PRP growth factors was the lowest. The study demonstrated the similar outcome in osteogenic differentiation. In terms of gene expression and western bolt, the PCR results showed that the expression levels of OCN gene and RUNX2 protein in each PRP group were higher than that in the control group (P < 0.05). Different anticoagulants caused different degrees of lysis and spontaneous activation of platelets, which lead to different quality of PRP. Compared with HS and SC, EDTA could maintain the structural integrity of platelets, reduce their spontaneous activation, and increase the release of PRP growth factors for a longer period of time, thus ensuring the biomass of PRP. In addition, different coagulants also showed different results in the proliferation as well as osteogenic differentiation of BMSCs. Compared with Thrombin and ADP, Collagen-I may be a better choice.
Asunto(s)
Anticoagulantes/farmacología , Coagulantes/farmacología , Plasma Rico en Plaquetas/metabolismo , Animales , Bioensayo , Plaquetas/citología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Linaje de la Célula/efectos de los fármacos , Linaje de la Célula/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Fosfatasa 2 de Especificidad Dual/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Osteocalcina/genética , Osteocalcina/metabolismo , Osteogénesis/efectos de los fármacos , Osteogénesis/genética , Selectina-P/metabolismo , Factor de Crecimiento Derivado de Plaquetas/farmacología , Conejos , Estándares de Referencia , Factor de Crecimiento Transformador beta1/farmacologíaRESUMEN
OBJECTIVES: Reduced platelet (PLT) function during storage has been shown for buffy-coat-derived platelet concentrates (BCP) and apheresis platelet units (AP), while for whole blood (WB) it has not been well studied. The aim of this study was to investigate PLT function in these blood products throughout storage using a novel flow cytometric assay. METHODS: Flow cytometric measurement of agonist-induced platelet aggregation, CD62P expression and PAC-1 binding during storage in BCP, AP (1-9 days at 20°C) and WB (1-21 days at 2-6°C). RESULTS: PLT-aggregation capacity decreased from day 1 to day 7 for almost all product-agonist combinations (P = .004 to P = .029) with aggregation capacity of WB being similar to that of AP and BCP. WB aggregation capacity remained relatively unchanged from day 7 to day 21. For all blood products, the fraction of agonist-induced CD62P-expression remained high and the fraction of PAC-1 binding decreased during storage. WB PLTs underwent only small changes in CD62P expression and PAC-1 binding from day 7 to day 21. CONCLUSION: This study found PLT aggregation in WB stored at 4°C to be as least as good as for BCP and AP stored at 20°C. WB retained significant PLT-aggregation capacity to day 21.
Asunto(s)
Plaquetas/metabolismo , Conservación de la Sangre , Fosfatasa 2 de Especificidad Dual/metabolismo , Selectina-P/metabolismo , Agregación Plaquetaria , Plaquetas/citología , Citometría de Flujo , Humanos , Factores de TiempoRESUMEN
Pituitary adenylate cyclase activating polypeptide (PACAP) is a growth factor for lung cancer cells. PACAP-27 or PACAP-38 binds with high affinity to non-small cell lung cancer (NSCLC) cells, causing elevated cytosolic Ca2+, increased proliferation and increased phosphorylation of extracellular regulated kinase (ERK) and the epidermal growth factor receptor (EGFR). The role of reactive oxygen species (ROS) was investigated in these processes. Addition of PACAP-38 to NCI-H838 or A549 cells increased the tyrosine phosphorylation of the EGFR, HER2 and ERK significantly by 4-, 3-, and 2-fold, respectively. The transactivation of the EGFR and HER2 was inhibited by gefitinib or lapatinib (tyrosine kinase inhibitors), PACAP (6-38) (PAC1 antagonist), N-acetylcysteine (NAC is an anti-oxidant) or dipheyleneiodonium (DPI is an inhibitor of Nox and Duox enzymes). PACAP-38 addition to NSCLC cells increased ROS which was inhibited by PACAP (6-38), NAC or DPI. Nox1, Nox2, Nox3, Nox4, Nox5, Duox1 and Duox2 mRNA was present in many NSCLC cell lines. PACAP-38 stimulated the growth of NSCLC cells whereas PACAP (6-38), gefitinib or DPI inhibited proliferation. The results show that ROS are essential for PAC1 to regulate EGFR and HER2 transactivation as well as proliferation of NSCLC cells.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Fosfatasa 2 de Especificidad Dual/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares/metabolismo , Proteínas de Neoplasias/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Activación Transcripcional , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Fosfatasa 2 de Especificidad Dual/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Proteínas de Neoplasias/genéticaRESUMEN
Dual-specificity phosphatase 2 (DUSP2), a member of nuclear type I DUSP family, abolishes the activation of mitogen-activated protein kinases (MAPKs) and plays critical roles in the immune processes, inflammatory responses, and cancer progression. Currently, whether DUSP2 is involved in pathogenesis of bladder cancer remains unclear. In this study, we demonstrate that the expression level of DUSP2 was predominantly downregulated in bladder cancer tissues and cell lines as compared with that of paired normal tissues and benign urothelial cells. Besides, the expression of DUSP2 was significantly associated with pathological grade (P = .009), AJCC stage (P = .017), and subtype (P = .001) in The Cancer Genome Atlas cohort and mainly related to TNM stage (P = .016) in the tissue microarray cohort. Kaplan-Meier analysis suggested that patients with low DUSP2 expression had a shorter 5-year overall survival (P = .018 in The Cancer Genome Atlas; P = .012 in tissue microarray) and lower recurrence-free survival (P = .008). Cox regression analysis indicated that reduced DUSP2 was an independent high risk factor for survival prognosis in both cohorts. Taken together, our findings for the first time suggested DUSP2 as a progression and prognosis biomarker for bladder cancer. Whether DUSP2 functions as a tumor suppressor in bladder cancer deserves further studies.
Asunto(s)
Regulación hacia Abajo , Fosfatasa 2 de Especificidad Dual/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias de la Vejiga Urinaria/metabolismo , Vejiga Urinaria/metabolismo , Anciano , Línea Celular Tumoral , Progresión de la Enfermedad , Fosfatasa 2 de Especificidad Dual/genética , Femenino , Humanos , Masculino , Persona de Mediana Edad , Clasificación del Tumor , Estadificación de Neoplasias , Pronóstico , Supervivencia sin Progresión , Tasa de Supervivencia , Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/mortalidad , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
BACKGROUND Ovarian cancer commonly presents at a late stage and is associated with poor prognosis. The most common histological subtype is serous ovarian carcinoma. Dual-specificity phosphatase 2 (DUSP2) is a protein phosphatase and substrate for mitogen-activated protein kinases (MAPKs) with increased expression levels in malignancy. This study aimed to evaluate the expression of DUSP2 in tumor tissues from patients with serous ovarian carcinoma and the association with tumor grade, stage, and patient survival and to investigate the effects of DUSP2 expression in SKOV3 and OVCAR3 cells in vitro. MATERIAL AND METHODS Tumor tissue and adjacent normal ovarian tissue from 127 patients with histologically confirmed serous ovarian carcinoma underwent quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry to measure DUSP2 mRNA and protein expression, respectively. Tumor grade, stage, and clinicopathological data underwent correlation analysis with DUSP2 expression, and survival data were assessed with Kaplan-Meier and Cox regression analysis. The effects of DUSP2 expression on the proliferation and migration of SKOV3 and OVCAR3 cells were evaluated. RESULTS Immunohistochemistry showed that DUSP2 was down-regulated in serous ovarian carcinoma tissues compared with adjacent ovarian tissues, and was significantly correlated with tumor stage. Survival analysis showed that DUSP2 expression was an independent risk factor for patient survival. DUSP2 expression in SKOV3 and OVCAR3 cells in vitro suppressed cell proliferation and migration. CONCLUSIONS Down-regulation of DUSP2 expression in serous ovarian carcinoma was an independent risk factor for patient survival, and its expression in SKOV3 and OVCAR3 cells inhibited cell proliferation and migration in vitro.
Asunto(s)
Fosfatasa 2 de Especificidad Dual/metabolismo , Neoplasias Quísticas, Mucinosas y Serosas/enzimología , Neoplasias Ováricas/enzimología , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Regulación hacia Abajo/genética , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Flavonoides/farmacología , Regulación Neoplásica de la Expresión Génica , Humanos , Persona de Mediana Edad , Análisis Multivariante , Neoplasias Quísticas, Mucinosas y Serosas/genética , Neoplasias Quísticas, Mucinosas y Serosas/patología , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Pronóstico , Análisis de SupervivenciaRESUMEN
BACKGROUND: We evaluated the effect of surfactant on platelet function as a potential contributing mechanism to the pathogenesis of pulmonary hemorrhage (PHEM) in term and preterm infants. METHODS: Cord blood samples were collected from neonates following delivery. Complete blood count and platelet function were measured using a cone and platelet analyzer (CPA). Increasing surfactant concentrations were added to platelets in vitro, and the adhesion molecule P-selectin and the monoclonal antibody PAC-1 were evaluated following platelet activation by flow cytometry. RESULTS: Forty-one infants (11 preterm and 30 term) were studied. CPA revealed a significant decrease in the average size of the aggregates and in platelet adhesion when surfactant was added. In term infants, the addition of surfactant to native platelets yielded an increased binding capacity of PAC-1 but did not affect P-selectin expression. In preterm infants, platelet activation with adenosine diphosphate in the presence of a high surfactant concentration (0.5 mg/mL) resulted in increased PAC-1 binding and no change in P-selectin expression. CONCLUSIONS: The platelets of preterm infants are less active (hyporesponsive) than those of term infants, both in their native state as well as after stimulation with various agonists. Surfactant may play an important role in treating PHEM in preterm infants.
Asunto(s)
Hemorragia/tratamiento farmacológico , Enfermedades Pulmonares/tratamiento farmacológico , Tensoactivos/uso terapéutico , Plaquetas/citología , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Fosfatasa 2 de Especificidad Dual/química , Fosfatasa 2 de Especificidad Dual/metabolismo , Hemorragia/diagnóstico , Humanos , Recién Nacido , Recien Nacido Prematuro , Enfermedades Pulmonares/diagnóstico , Selectina-P/metabolismo , Unión ProteicaRESUMEN
BACKGROUND 5-Fluorouracil (5-FU)-based chemotherapy is a conventional therapeutic approach for the treatment of patients with colorectal cancer (CRC). However, development of 5-FU resistance frequently occurs. We explored a potential method for regulating the sensitivity to 5-FU-based chemotherapy in CRC patients. MATERIAL AND METHODS Cell viability was determined by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Gene expression levels were detected by real-time quantitative polymerase chain reaction (RT-qPCR). Protein expression levels were evaluated by Western blot. TargetScan was used for the prediction of binding sites for miRNA in mRNAs. The interaction between mRNA 3'UTR and miRNA was verified by dual luciferase reporter assay. Tissue samples were obtained from 33 CRC patients who received surgery at Xingtai People's Hospital. RESULTS miR-106a level was associated with 5-FU sensitivity in CRC cells. Overexpression of miR-106a reduced 5-FU sensitivity of HCT116 and SW620 cells, and antagonist of miR-106a sensitized HCT116 and SW620 towards 5-FU. miR-106a overexpression decreased dual-specificity phosphatases 2 (DUSP2) expression at mRNA and protein levels in HCT116 and SW620 cells. Through downregulation of DUSP2, miR-106a elevation increased COX-2 expression and stemness-maintenance genes (SOX2 and OCT4). Furthermore, we predicted that miR-106a directly binds to 3'UTR of DUSP2 mRNA, which was confirmed by dual luciferase assay. Silencing of DUSP2 reversed elevated 5-FU sensitivity induced by miR-106a antagonist in HCT116 cells. A negative correlation was discovered between miR-106a and DUSP2 in tumor samples of CRC patients. CONCLUSIONS miR-106a plays an important role in mediating response to 5-FU-based chemotherapy in CRC and could serve as a potential target for CRC patients.
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Neoplasias Colorrectales/patología , Fosfatasa 2 de Especificidad Dual/metabolismo , MicroARNs/fisiología , Regiones no Traducidas 3'/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , China , Regulación hacia Abajo/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Fosfatasa 2 de Especificidad Dual/genética , Fluorouracilo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HCT116 , Humanos , MicroARNs/genética , ARN Mensajero/genéticaRESUMEN
Metastasis remains one of the most intractable challenges in pancreatic ductal adenocarcinoma (PDAC) biology, and epithelial-to-mesenchymal transition (EMT) is essential to the epithelium-originated solid tumor metastasis cascade. Emerging evidence demonstrates that aberrant miRNA expression is involved in pancreatic cancer progression. We found that miR-361-3p was associated with an advanced stage of PDAC and poor prognosis. Hence, the effect of miR-361-3p on metastasis of PDAC cells was evaluated using Transwell assay and wound healing assay in vitro as well as orthotopic and liver metastasis pancreatic cancer models in vivo. Overexpression of miR-361-3p promoted pancreatic cancer cell migration and invasion in vitro, and miR-361-3p-elevated PDAC cells were prone to generating metastatic nodules in vivo. However, miR-361-3p showed no significant effect on the proliferation of PDAC cells in vivo or in vitro. Further study demonstrated that miR-361-3p could enhance EMT and ERK pathway activation, and ERK inhibitor could attenuate miR-361-3p-induced EMT. Luciferase assays, qPCR, and western blot and Ago2 co-immunoprecipitation were performed to identify the direct target of miR-361-3p. Mechanistic investigations identified DUSP2 as a direct target of miR-361-3p, and DUSP2 was revealed to be involved in miR-361-3p-induced EMT by directly leading to the inactivation of the ERK pathway. Moreover, we found that miR-361-3p-induced EMT was dependent on Ago2, the core component of RNA-induced silencing complex, while enforced expression of Ago2 enhanced the miR-361-3p-induced effect by promoting interference efficacy and specificity rather than regulating miR-361-3p stability and biogenesis. Thus, this study revealed that miR-361-3p functions as an oncomiR for promoting metastasis and identified the miR-361-3p/DUSP2/ERK axis as a novel EMT axis dependent on Ago2 in PDAC.
Asunto(s)
Carcinoma Ductal Pancreático/patología , Fosfatasa 2 de Especificidad Dual/genética , MicroARNs/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Neoplasias Pancreáticas/patología , Regiones no Traducidas 3' , Animales , Antagomirs/metabolismo , Proteínas Argonautas/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/mortalidad , Línea Celular Tumoral , Movimiento Celular , Fosfatasa 2 de Especificidad Dual/química , Fosfatasa 2 de Especificidad Dual/metabolismo , Transición Epitelial-Mesenquimal , Humanos , Ratones , Ratones Desnudos , MicroARNs/antagonistas & inhibidores , MicroARNs/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/mortalidad , Estabilidad del ARN , Tasa de SupervivenciaRESUMEN
Alzheimer's disease (AD) is one of the most common causes of dementia. AD pathogenesis has been hypothesized to involve cholinergic deficits, amyloid-beta protein (Aß) deposition, tau protein hyperphosphorylation, and chronic neuroinflammation. Many single-target drugs have gone through the various stages of pre-clinical and clinical development in an effort to cure AD; however, the current clinically approved drugs have only limited effects on the disease progression. With the accumulation of unsuccessful clinical trials using single-target drugs, multi-target directed ligand (MTDL) drug development is becoming more common. MTDLs incorporate two or more pharmacophores into a single drug molecule. This approach can alleviate side effects and lead to a better pharmacokinetic profile of the MTDL compared to two or more separate drugs representing respective single pharmacophores. This review discusses cathepsin B (CatB), dual specificity phosphatase 2 (DUSP2), and monoglycerol lipase (MAGL) as targets for MTDLs aimed at slowing down the neuroinflammatory component of neurodegenerative diseases. CatB, DUSP2 and MAGL inhibitors show promising preclinical anti-inflammatory effects in vivo and in vitro. Incorporating pharmacophores that inhibit these targets into MTDLs represents a promising avenue towards effective suppression of neuroinflammation associated with AD.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Inflamación/tratamiento farmacológico , Péptidos beta-Amiloides/metabolismo , Animales , Antiinflamatorios/química , Catepsina B/metabolismo , Dinoprostona/metabolismo , Progresión de la Enfermedad , Fosfatasa 2 de Especificidad Dual/metabolismo , Humanos , Interleucina-1beta/metabolismo , Ligandos , Lipasa/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Enfermedades Neurodegenerativas/tratamiento farmacológico , Óxido Nítrico/química , Fosforilación , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Growing evidence has revealed that the transcription factor basic leucine zipper transcription factor ATF-like 2 (BATF2) has unique transcriptional activities, including regulating cytokines via TLR signals in macrophages, which affect mortality due to infection and cancer. On the basis of genome-wide analyses using the chromatin immunoprecipitation-sequencing technique, we found that dual-specificity phosphatase 2 (Dusp2) had a significantly lower acetyl-histone status in Batf2-/- bone marrow-derived macrophages (BMDMs) compared with wild-type (WT) BMDMs. The phosphatase DUSP2 has been reported to play a critical role in inflammatory responses. Therefore, we evaluated the BATF2 transcriptional activities on the Dusp2 promoter. We found that the DUSP2 and IL-12 p40 expression levels were significantly lower in Batf2-/- BMDMs than in WT controls following their stimulation with TLR7 ligands. Further in vitro studies revealed that phospho-STAT3 was up-regulated and NF-κB p50/p65 were down-regulated in Batf2-/- BMDMs compared with their levels in WT controls. Additionally, Th1 immunity was impaired in Batf2-/- mice following their stimulation with TLR7 ligands. We also found that BATF2 interacts with NF-κB p65 and promotes DUSP2 expression through the NF-κB-binding site in the Dusp2 promoter at -203 to -121. Collectively, our findings suggest that BATF2 activates DUSP2 gene expression and up-regulates NF-κB activity via phospho-STAT3 dephosphorylation.
Asunto(s)
Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Fosfatasa 2 de Especificidad Dual/genética , FN-kappa B/metabolismo , Factor de Transcripción STAT3/metabolismo , Regulación hacia Arriba , Animales , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/deficiencia , Fosfatasa 2 de Especificidad Dual/metabolismo , Perfilación de la Expresión Génica , Ratones , Ratones Noqueados , Fosforilación , Células RAW 264.7RESUMEN
Shikonin is a naphthoquinone isolated from the dried root of Lithospermum erythrorhizon, an herb used in Chinese medicine. Although several studies have indicated that shikonin exhibits antitumor activity in breast cancer, the mechanism of action remains unclear. In the present study, we performed transcriptome analysis using RNA-seq and explored the mechanism of action of shikonin in regulating the growth of different types of breast cancer cells. The IC50 of shikonin on MCF-7, SKBR-3 and MDA-MB-231 cells were 10.3 µΜ, 15.0 µΜ, 15.0 µΜ respectively. Our results also demonstrated that shikonin arrests the progression of cell cycle and induces apoptosis in MDA-MB-231 cells. Using RNA-seq transcriptome analysis, we found 38 common genes that significantly express in different types of breast cancer cells under shikonin treatment. In particular, our results indicated that shikonin induces the expression of dual specificity phosphatase (DUSP)-1 and DUSP2 in both RNA and protein levels. In addition, shikonin also inhibits the phosphorylation of JNK and p38, the downstream signaling molecules of DUSP1 and DUSP2. Therefore, our results suggest that shikonin induces the expression of DUSP1 and DUSP2 which consequently switches off JNK and p38 MAPK pathways and causes cell cycle arrest and apoptosis in breast cancer cells.
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Neoplasias de la Mama/genética , Naftoquinonas/farmacología , Transcriptoma/efectos de los fármacos , Apoptosis/efectos de los fármacos , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Fosfatasa 1 de Especificidad Dual/metabolismo , Fosfatasa 2 de Especificidad Dual/metabolismo , Perfilación de la Expresión Génica , Humanos , Lithospermum/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células MCF-7 , Naftoquinonas/metabolismo , ARN/metabolismo , Transducción de Señal/efectos de los fármacos , Transcriptoma/genéticaRESUMEN
αIIbß3, the major platelet integrin, plays a central role in hemostasis and thrombosis. Upon platelet activation, conformation of αIIbß3 changes and allows fibrinogen binding and, subsequently, platelet aggregation. It was previously shown that a lipid-modified platelet permeable peptide, which corresponds to the intracellular acidic membrane distal sequence 1000LEEDDEEGE1008 of αIIb (pal-K-LEEDDEEGE or pal-K-1000-1008), inhibits thrombin-induced human platelet aggregation, by inhibiting talin association with the integrin. YMESRADR, a peptide corresponding to the extracellular sequence 313-320 of αIIb, is also a potent platelet aggregation inhibitor by mimicking the effect of a clasp between the head domains of αIIb and ß3. The aim of the present study was to investigate the synergistic effect of the intra- and extracellular- peptide inhibitors on platelet aggregation, as well as on the phosphorylation of two signaling proteins, focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK). Platelet preincubation with Pal-K-LEEDDEGE followed by YMESRADR showed a synergistic inhibitory activity on platelet aggregation. Platelet incubation with threshold inhibitory concentrations of both peptides provoked almost the total inhibition of aggregation, PAC-1 binding, and fibrinogen binding, but not P-selectin exposure on activated platelets' surface. Like RGDS peptide, this mixture inhibits FAK phosphorylation whose phosphorylation is well known to be consecutive to the aggregation (postoccupancy events). However, in contrast to RGDS peptide that enhances ERK phosphorylation and activation, the mixture of threshold inhibitory concentrations of Pal-K-LEEDDEEGE and YMESRADR inhibits ERK phosphorylation. We suggest that the use of the intracellular in combination with the extracellular peptide inhibitor, acting with a non-RGD-like mechanism, may provide an alternative way to antagonize integrin αIIbß3 activation.
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Plaquetas/efectos de los fármacos , Plaquetas/fisiología , Péptidos/farmacología , Activación Plaquetaria/efectos de los fármacos , Agregación Plaquetaria/efectos de los fármacos , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/química , Dominios y Motivos de Interacción de Proteínas , Secuencia de Aminoácidos , Sinergismo Farmacológico , Fosfatasa 2 de Especificidad Dual/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Citometría de Flujo , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Humanos , Selectina-P/metabolismo , Fosforilación/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/farmacología , Unión ProteicaRESUMEN
Dual-specificity MAP kinase (MAPK) phosphatases (DUSPs) are well-established negative modulators in regulating MAPK signaling in mammalian cells and tissues. Our previous studies have shown the involvement of DUSP6 in regulating innate immunity in Japanese flounder Paralichthys olivaceus. In order to gain a better understanding of the role of DUSPs in fish innate immunity, in the present study we identified and characterized three additional DUSP genes including DUSP1, 2 and 5 in P. olivaceus. The three Japanese flounder DUSP proteins share common domain structures composed of a conserved N-terminal Rhodanase/CDC25 domain and a C-terminal catalytic phosphatase domain, while they show only less than 26% sequence identities, indicating that they may have different substrate selectivity. In addition, mRNA transcripts of all the three DUSP genes are detected in all examined Japanese flounder tissues; however, DUSP1 is dominantly expressed in spleen while DUSP2 and 5 are primarily expressed in skin. Furthermore, all the three DUSP genes are constitutively expressed in the Japanese flounder head kidney macrophages (HKMs) and peripheral blood leucocytes (PBLs) with unequal distribution patterns. Moreover, all the three DUSPs gene expression was induced differently in response to the LPS and double-stranded RNA mimic poly(I:C) stimulations both in the Japanese flounder HKMs and PBLs, suggesting an association of DUSPs with TLR signaling in fish. Taken together, the co-expression of various DUSPs members together with their different responses to the immune challenges indicate that the DUSP members may operate coordinately in regulating the MAPK-dependent immune responses in the Japanese flounder.
Asunto(s)
Fosfatasas de Especificidad Dual/genética , Proteínas de Peces/genética , Peces Planos/genética , Peces Planos/inmunología , Regulación de la Expresión Génica/inmunología , Inmunidad Innata/genética , Secuencia de Aminoácidos , Animales , Técnicas de Cultivo de Célula , Fosfatasa 1 de Especificidad Dual/química , Fosfatasa 1 de Especificidad Dual/genética , Fosfatasa 1 de Especificidad Dual/inmunología , Fosfatasa 1 de Especificidad Dual/metabolismo , Fosfatasa 2 de Especificidad Dual/química , Fosfatasa 2 de Especificidad Dual/genética , Fosfatasa 2 de Especificidad Dual/inmunología , Fosfatasa 2 de Especificidad Dual/metabolismo , Fosfatasas de Especificidad Dual/química , Fosfatasas de Especificidad Dual/inmunología , Fosfatasas de Especificidad Dual/metabolismo , Proteínas de Peces/química , Proteínas de Peces/inmunología , Regulación de la Expresión Génica/efectos de los fármacos , Lipopolisacáridos/farmacología , Filogenia , Poli I-C/farmacología , Alineación de Secuencia/veterinariaRESUMEN
Cancer stem-like cells (CSC) evolve to overcome the pressures of reduced oxygen, nutrients or chemically induced cell death, but the mechanisms driving this evolution are incompletely understood. Here, we report that hypoxia-mediated downregulation of the dual specificity phosphatase 2 (DUSP2) is critical for the accumulation of CSC in colorectal cancer. Reduced expression of DUSP2 led to overproduction of COX-2-derived prostaglandin E2, which promoted cancer stemness via the EP2/EP4 signaling pathways. Genetic and pharmacological inhibition of PGE2 biosynthesis or signal transduction ameliorated loss-of-DUSP2-induced tumor growth and cancer stemness. Genome-wide profile analysis revealed that genes regulated by DUSP2 were similar to those controlled by histone deacetylase. Indeed, treatment with novel histone deacetylase inhibitors abolished hypoxia-induced DUSP2 downregulation, COX-2 overexpression, cancer stemness, tumor growth, and drug resistance. Our findings illuminate mechanisms of cancer stemness and suggest new cancer therapy regimens. Cancer Res; 77(16); 4305-16. ©2017 AACR.