RESUMEN
ANP32B, a member of the acidic leucine-rich nuclear phosphoprotein 32 kDa (ANP32) family of proteins, is critical for normal development because its constitutive knockout mice are perinatal lethal. It is also shown that ANP32B acts as a tumor-promoting gene in some kinds of cancer such as breast cancer and chronic myelogenous leukemia. Herein, we observe that ANP32B is lowly expressed in B-cell acute lymphoblastic leukemia (B-ALL) patients, which correlates with poor prognosis. Furthermore, we utilized the N-myc or BCR-ABLp190 -induced B-ALL mouse model to investigate the role of ANP32B in B-ALL development. Intriguingly, conditional deletion of Anp32b in hematopoietic cells significantly promotes leukemogenesis in two B-ALL mouse models. Mechanistically, ANP32B interacts with purine rich box-1 (PU.1) and enhances the transcriptional activity of PU.1 in B-ALL cells. Overexpression of PU.1 dramatically suppresses B-ALL progression, and highly expressed PU.1 significantly reverses the accelerated leukemogenesis in Anp32b-deficient mice. Collectively, our findings identify ANP32B as a suppressor gene and provide novel insight into B-ALL pathogenesis.
Asunto(s)
Linfoma de Burkitt , Leucemia Mieloide , Leucemia-Linfoma Linfoblástico de Células Precursoras , Animales , Ratones , Proteínas Nucleares/genética , Ratones Noqueados , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Proteínas de Fusión bcr-abl , Proteínas del Tejido Nervioso/metabolismo , Proteínas de Ciclo Celular/metabolismoRESUMEN
Proper regulation of p53 signaling is critical for the maintenance of hematopoietic stem cells (HSCs) and leukemic stem cells (LSCs). The hematopoietic cell-specific mechanisms regulating p53 activity remain largely unknown. Here, we demonstrate that conditional deletion of acidic leucine-rich nuclear phosphoprotein 32B (ANP32B) in hematopoietic cells impairs repopulation capacity and postinjury regeneration of HSCs. Mechanistically, ANP32B forms a repressive complex with p53 and thus inhibits the transcriptional activity of p53 in hematopoietic cells, and p53 deletion rescues the functional defect in Anp32b-deficient HSCs. Of great interest, ANP32B is highly expressed in leukemic cells from patients with chronic myelogenous leukemia (CML). Anp32b deletion enhances p53 transcriptional activity to impair LSC function in a murine CML model and exhibits synergistic therapeutic effects with tyrosine kinase inhibitors in inhibiting CML propagation. In summary, our findings provide a novel strategy to enhance p53 activity in LSCs by inhibiting ANP32B and identify ANP32B as a potential therapeutic target in treating CML.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Células Madre Neoplásicas/patología , Proteínas del Tejido Nervioso/metabolismo , Proteínas Nucleares/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Proteínas de Ciclo Celular/genética , Células Cultivadas , Regulación Leucémica de la Expresión Génica , Células Madre Hematopoyéticas/metabolismo , Células Madre Hematopoyéticas/patología , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Ratones , Células Madre Neoplásicas/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas Nucleares/genética , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Transgenic animal models with homologous etiology provide a promising way to pursue the neurobiological substrates of the behavioral deficits in autism spectrum disorder (ASD). Gain-of-function mutations of MECP2 cause MECP2 duplication syndrome, a severe neurological disorder with core symptoms of ASD. However, abnormal brain developments underlying the autistic-like behavioral deficits of MECP2 duplication syndrome are rarely investigated. To this end, a human MECP2 duplication (MECP2-DP) rat model was created by the bacterial artificial chromosome transgenic method. Functional and structural magnetic resonance imaging (MRI) with high-field were performed on 16 male MECP2-DP rats and 15 male wildtype rats at postnatal 28 days, 42 days, and 56 days old. Multimodal fusion analyses guided by locomotor-relevant metrics and social novelty time separately were applied to identify abnormal brain networks associated with diverse behavioral deficits induced by MECP2 duplication. Aberrant functional developments of a core network primarily composed of the dorsal medial prefrontal cortex (dmPFC) and retrosplenial cortex (RSP) were detected to associate with diverse behavioral phenotypes in MECP2-DP rats. Altered developments of gray matter volume were detected in the hippocampus and thalamus. We conclude that gain-of-function mutations of MECP2 induce aberrant functional activities in the default-mode-like network and aberrant volumetric changes in the brain, resulting in autistic-like behavioral deficits. Our results gain critical insights into the biomarker of MECP2 duplication syndrome and the neurobiological underpinnings of the behavioral deficits in ASD.
Asunto(s)
Trastorno del Espectro Autista , Discapacidad Intelectual Ligada al Cromosoma X , Animales , Trastorno del Espectro Autista/diagnóstico por imagen , Trastorno del Espectro Autista/genética , Encéfalo/metabolismo , Mapeo Encefálico/métodos , Humanos , Masculino , Discapacidad Intelectual Ligada al Cromosoma X/genética , Proteína 2 de Unión a Metil-CpG/genética , Proteína 2 de Unión a Metil-CpG/metabolismo , RatasRESUMEN
Vascular network reconstruction plays a pivotal role in the axonal regeneration and nerve function recovery after peripheral nerve injury. Increasing evidence indicates that Schwann cells (SCs) can promote nerve function repair, and the beneficial effects attributed to SCs therapy may exert their therapeutic effects through paracrine mechanisms. Recently, the previous research of our group demonstrated the promising neuroregenerative capacity of Schwann-like cells (SCLCs) derived from differentiated human embryonic stem cell-derived neural stem cells (hESC-NSCs) in vitro. Herein, the effects of SC-like cell conditioned medium (SCLC-CM) on angiogenesis and nerve regeneration were further explored. The assays were performed to show the pro-angiogenic effects of SCLC-CM, such as promoted endothelial cell proliferation, migration and tube formation in vitro. In addition, Sprague-Dawley rats were treated with SCLC-CM after sciatic nerve crush injury, SCLC-CM was conducive for the recovery of sciatic nerve function, which was mainly manifested in the SFI increase, the wet weight ratio of gastrocnemius muscle, as well as the number and thickness of myelin. The SCLC-CM treatment reduced the Evans blue leakage and increased the expression of CD34 microvessels. Furthermore, SCLC-CM upregulated the expressions of p-Akt and p-mTOR in endothelial cells. In conclusion, SCLC-CM promotes angiogenesis and nerve regeneration, it is expected to become a new treatment strategy for peripheral nerve injury.
Asunto(s)
Células Endoteliales , Traumatismos de los Nervios Periféricos , Animales , Medios de Cultivo Condicionados/farmacología , Regeneración Nerviosa , Traumatismos de los Nervios Periféricos/metabolismo , Traumatismos de los Nervios Periféricos/terapia , Ratas , Ratas Sprague-Dawley , Células de Schwann , Nervio CiáticoRESUMEN
RATIONALE: Replication-independent histone turnover has been linked to cis-regulatory chromatin domains in cultured cell lines, but its molecular underpinnings and functional relevance in adult mammalian tissues remain yet to be defined. OBJECTIVE: We investigated regulatory functions of replication-independent histone turnover in chromatin states of postmitotic cardiomyocytes from adult mouse heart. METHODS AND RESULTS: We used H2B-GFP (histone 2B-green fluorescent protein) fusion protein pulse-and-chase approaches to measure histone turnover rate in mouse cardiomyocytes. Surprisingly, we found that the short histone half-life (≈2 weeks) contrasted dramatically with the long lifetime of cardiomyocytes, and rapid histone turnover regions corresponded to cis-regulatory domains of heart genes. Interestingly, recruitment of chromatin modifiers, including Polycomb EED (embryonic ectoderm development), was positively correlated with histone turnover rate at enhancers. Mechanistically, through directly interacting with and engaging the BAF (BRG1 [Brahma-related gene-1]-associated factor) complex for nucleosome exchange for stereotyped histone modifications from the free histone pool, EED augmented histone turnover to restrain enhancer overactivation. CONCLUSIONS: We propose a model in which replication-independent histone turnover reinforces robustness of local chromatin states for adult tissue homeostasis.
Asunto(s)
Ensamble y Desensamble de Cromatina , Epigénesis Genética , Código de Histonas , Histonas/metabolismo , Homeostasis , Miocitos Cardíacos/metabolismo , Animales , Células Cultivadas , ADN Helicasas/metabolismo , Replicación del ADN , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Nucleares/metabolismo , Complejo Represivo Polycomb 2/metabolismo , Factores de Transcripción/metabolismoRESUMEN
RATIONALE: We hypothesized that the differentiation processes of cardiac progenitor cell (CP) from first and second heart fields (FHF and SHF) may undergo the unique instructive gene regulatory networks or signaling pathways, and the precise SHF progression is contingent on the FHF signaling developmental cues. OBJECTIVE: We investigated how the intraorgan communications control sequential building of discrete anatomic regions of the heart at single-cell resolution. METHODS AND RESULTS: By single-cell transcriptomic analysis of Nkx2-5 (NK2 homeobox 5) and Isl1 (ISL LIM homeobox 1) lineages at embryonic day 7.75, embryonic day 8.25, embryonic day 8.75, and embryonic day 9.25, we present a panoramic view of distinct CP differentiation hierarchies. Computational identifications of FHF- and SHF-CP descendants revealed that SHF differentiation toward cardiomyocytes underwent numerous step-like transitions, whereas earlier FHF progressed toward cardiomyocytes in a wave-like manner. Importantly, single-cell pairing analysis demonstrated that SHF-CPs were attracted to and expanded FHF-populated heart tube region through interlineage communications mediated by the chemotactic guidance (MIF [macrophage migration inhibitory factor]-CXCR2 [C-X-C motif chemokine receptor 2]). This finding was verified by pharmacological blockade of this chemotaxis in embryos manifesting limited SHF cell migration and contribution to the growth of the outflow tract and right ventricle but undetectable effects on the left ventricle or heart tube initiation. Genetic loss-of-function assay of Cxcr2 showed that the expression domain of CXCR4 was expanded predominantly at SHF. Furthermore, double knockout of Cxcr2/Cxcr4 exhibited defective SHF development, corroborating the redundant function. Mechanistically, NKX2-5 directly bound the Cxcr2 and Cxcr4 genomic loci and activated their transcription in SHF. CONCLUSIONS: Collectively, we propose a model in which the chemotaxis-mediated intraorgan crosstalk spatiotemporally guides the successive process of positioning SHF-CP and promoting primary heart expansion and patterning upon FHF-derived heart tube initiation.
Asunto(s)
Quimiotaxis , Células Madre Embrionarias/metabolismo , Regulación del Desarrollo de la Expresión Génica , Corazón/embriología , Proteína Homeótica Nkx-2.5/metabolismo , Transcriptoma , Animales , Linaje de la Célula , Células Cultivadas , Células Madre Embrionarias/citología , Proteína Homeótica Nkx-2.5/genética , Proteínas con Homeodominio LIM/genética , Proteínas con Homeodominio LIM/metabolismo , Ratones , Ratones Endogámicos C57BL , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Receptores de Interleucina-8B/genética , Receptores de Interleucina-8B/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Axon regeneration and functional recovery after peripheral nerve injury remains a clinical challenge. Injury leads to axonal disintegration after which Schwann cells (SCs) and macrophages re-engage in the process of regeneration. At present, biomaterials are regarded as the most promising way to repair peripheral nerve damage. As a natural material, keratin has a wide range of sources and has good biocompatibility and biodegradability. Here, a keratin was extracted from human hair by reducing method and a keratin sponge with porous structure was obtained by further processing. The results suggested that keratin can promote cell adhesion, proliferation, migration as well as the secretion of neurotrophic factors by SCs and the regulation of the expression of macrophage inflammatory cytokines in vitro. We report for the first time that human hair keratin can promote the extension of axon in DRG neurons. The motor deficits caused by a sciatic nerve crush injury were alleviated by keratin sponge dressing in vivo. Thus, keratin has been identified as a valuable biomaterial that can enhance peripheral nerve regeneration.
Asunto(s)
Cabello/química , Queratinas Específicas del Pelo/farmacología , Regeneración Nerviosa/efectos de los fármacos , Nervios Periféricos/efectos de los fármacos , Nervio Ciático/lesiones , Animales , Axones/efectos de los fármacos , Materiales Biocompatibles , Adhesión Celular , Línea Celular , Movimiento Celular , Proliferación Celular , Citocinas/metabolismo , Humanos , Inflamación , Macrófagos/efectos de los fármacos , Masculino , Ratones , Neuronas/metabolismo , Células RAW 264.7 , Ratas , Ratas Sprague-Dawley , Células de Schwann/efectos de los fármacos , Cicatrización de HeridasRESUMEN
Nkx2.5 plays protective roles in cardiac homeostasis and survival in the postnatal hearts. However, the underlying molecular mechanisms that mediate the protective functions of Nkx2.5 remain unknown. Here, we showed that Nkx2.5 was downregulated in response to various stresses and was required for protection against the stress-induced apoptosis of cardiomyocytes. SIRT1, a member of the sirtuin family of proteins, was found to be a direct transcriptional target of Nkx2.5 and was required for the Nkx2.5-mediated protection of cardiomyocytes from doxorubicin (DOX)-induced apoptosis. Moreover, using chromatin immunoprecipitation assays, we found that Nkx2.5 was able to bind to the SIRT1 promoter and that this binding was significantly decreased in DOX-treated mouse hearts. Furthermore, the cardiac-specific overexpression of SIRT1 decreased the DOX-induced apoptosis of cardiomyocytes in SIRT1 transgenic mouse hearts compared with the hearts of their wild-type littermates. These findings demonstrate that SIRT1 acts as a direct transcriptional target of Nkx2.5 that maintains cardiomyocyte homeostasis and survival.
Asunto(s)
Proteínas de Homeodominio/fisiología , Miocitos Cardíacos/fisiología , Sirtuina 1/fisiología , Estrés Fisiológico/fisiología , Factores de Transcripción/fisiología , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Supervivencia Celular/fisiología , Células Cultivadas , Doxorrubicina/farmacología , Proteína Homeótica Nkx-2.5 , Homeostasis/fisiología , Ratones , Ratones Transgénicos , Modelos Animales , Miocitos Cardíacos/citología , Miocitos Cardíacos/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Transducción de Señal/fisiología , Sirtuina 1/genética , Regulación hacia Arriba/fisiologíaRESUMEN
RATIONALE: Inactivation of the p66Shc adaptor protein confers resistance to oxidative stress and protects mice from aging-associated vascular diseases. However, there is limited information about the negative regulating mechanisms of p66Shc expression in the vascular system. OBJECTIVE: In this study, we investigated the role of SIRT1, a class III histone deacetylase, in the regulation of p66Shc expression and hyperglycemia-induced endothelial dysfunction. METHODS AND RESULTS: Expressions of p66Shc gene transcript and protein were significantly increased by different kinds of class III histone deacetylase (sirtuin) inhibitors in human umbilical vein endothelial cells and 293A cells. Adenoviral overexpression of SIRT1 inhibited high-glucose-induced p66Shc upregulation in human umbilical vein endothelial cells. Knockdown of SIRT1 increased p66Shc expression and also increased the expression levels of plasminogen activator inhibitor-1 expression, but decreased manganese superoxide dismutase expression in high-glucose conditions. However, knockdown of p66Shc significantly reversed the effects of SIRT1 knockdown. In addition, p66Shc overexpression significantly decreased manganese superoxide dismutase expression and increased plasminogen activator inhibitor-1 expression in high-glucose conditions, which were recovered by SIRT1 overexpression. Moreover, compared to streptozotocin-induced wild-type diabetic mice, endothelium-specific SIRT1 transgenic diabetic mice had decreased p66Shc expression at both the mRNA and the protein levels, improved endothelial function, and reduced accumulation of nitrotyrosine and 8-OHdG (markers of oxidative stress). We further found that SIRT1 was able to bind to the p66Shc promoter (-508 bp to -250 bp), resulting in a decrease in the acetylation of histone H3 bound to the p66Shc promoter region. CONCLUSION: Our findings indicate that repression of p66Shc expression by SIRT1 contributes to the protection of hyperglycemia-induced endothelial dysfunction.
Asunto(s)
Regulación hacia Abajo/genética , Endotelio Vascular/metabolismo , Hiperglucemia/genética , Proteínas Adaptadoras de la Señalización Shc/antagonistas & inhibidores , Sirtuina 1/fisiología , Envejecimiento/genética , Animales , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Endotelio Vascular/patología , Células HEK293 , Humanos , Hiperglucemia/patología , Hiperglucemia/prevención & control , Inmunidad Innata/genética , Masculino , Ratones , Ratones Transgénicos , Estrés Oxidativo/genética , Estabilidad Proteica , Proteínas Adaptadoras de la Señalización Shc/biosíntesis , Proteína Transformadora 1 que Contiene Dominios de Homología 2 de SrcRESUMEN
RATIONALE: Vascular smooth muscle cell (VSMC) proliferation and migration are crucial events involved in the pathophysiology of vascular diseases. Sirtuin 1 (SIRT1), a class III histone deacetylase (HDAC), has been reported to have the function of antiatherosclerosis, but its role in neointima formation remains unknown. OBJECTIVE: The present study was designed to investigate the role of SIRT1 in the regulation of neointima formation and to elucidate the underlying mechanisms. METHODS AND RESULTS: A decrease in SIRT1 expression was observed following carotid artery ligation. smooth muscle cell (SMC)-specific human SIRT1 transgenic (Tg) mice were generated. SIRT1 overexpression substantially inhibited neointima formation after carotid artery ligation or carotid artery wire injury. In the intima of injured carotid arteries, VSMC proliferation (proliferating cell nuclear antigen (PCNA)-positive cells) was significantly reduced. SIRT1 overexpression markedly inhibited VSMC proliferation and migration and induced cell cycle arrest at G1/S transition in vitro. Accordingly, SIRT1 overexpression decreased the induction of cyclin D1 and matrix metalloproteinase-9 (MMP-9) expression by treatment with serum and TNF-α, respectively, whereas RNAi knockdown of SIRT1 resulted in the opposite effect. Decreased cyclin D1 and MMP-9 expression/activity were also observed in injured carotid arteries from SMC-SIRT1 Tg mice. Furthermore, 2 targets of SIRT1, c-Fos and c-Jun, were involved in the downregulation of cyclin D1 and MMP-9 expression. CONCLUSIONS: Our findings demonstrate the inhibitory effect of SIRT1 on the VSMC proliferation and migration that underlie neointima formation and implicate SIRT1 as a potential target for intervention in vascular diseases.
Asunto(s)
Traumatismos de las Arterias Carótidas/metabolismo , Neointima/etiología , Neointima/metabolismo , Sirtuina 1/fisiología , Animales , Traumatismos de las Arterias Carótidas/genética , Traumatismos de las Arterias Carótidas/patología , Células Cultivadas , Humanos , Ligadura , Masculino , Ratones , Ratones Transgénicos , Neointima/patología , Ratas , Ratas Sprague-Dawley , Sirtuina 1/biosíntesisRESUMEN
BACKGROUND: Selectively targeting leukemia stem cells (LSCs) is a promising approach in treating acute myeloid leukemia (AML), for which identification of such therapeutic targets is critical. Increasing lines of evidence indicate that FBXO22 plays a critical role in solid tumor development and therapy response. However, its potential roles in leukemogenesis remain largely unknown. METHODS: We established a mixed lineage leukemia (MLL)-AF9-induced AML model with hematopoietic cell-specific FBXO22 knockout mice to elucidate the role of FBXO22 in AML progression and LSCs regulation, including self-renewal, cell cycle, apoptosis and survival analysis. Immunoprecipitation combined with liquid chromatography-tandem mass spectrometry analysis, Western blotting and rescue experiments were performed to study the mechanisms underlying the oncogenic role of FBXO22. RESULTS: FBXO22 was highly expressed in AML, especially in MLL-rearranged (MLLr) AML. Upon FBXO22 knockdown, human MLLr leukemia cells presented markedly increased apoptosis. Although conditional deletion of Fbxo22 in hematopoietic cells did not significantly affect the function of hematopoietic stem cells, MLL-AF9-induced leukemogenesis was dramatically abrogated upon Fbxo22 deletion, together with remarkably reduced LSCs after serial transplantations. Mechanistically, FBXO22 promoted degradation of BACH1 in MLLr AML cells, and overexpression of BACH1 suppressed MLLr AML progression. In line with this, heterozygous deletion of BACH1 significantly reversed delayed leukemogenesis in Fbxo22-deficient mice. CONCLUSIONS: FBXO22 promotes MLLr AML progression by targeting BACH1 and targeting FBXO22 might be an ideal strategy to eradicate LSCs without influencing normal hematopoiesis.
Asunto(s)
Factores de Transcripción con Cremalleras de Leucina de Carácter Básico , Proteínas F-Box , Leucemia Mieloide Aguda , Receptores Citoplasmáticos y Nucleares , Animales , Humanos , Ratones , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Ciclo Celular , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Células Madre Hematopoyéticas/metabolismo , Leucemia Mieloide Aguda/patología , Ratones Noqueados , Proteína de la Leucemia Mieloide-Linfoide/metabolismo , Células Madre Neoplásicas/patología , Receptores Citoplasmáticos y Nucleares/metabolismoRESUMEN
SIRT1 (Sirtuin type 1), a mammalian orthologue of yeast SIR2 (silent information regulator 2), has been shown to mediate a variety of calorie restriction (CR)-induced physiological events, such as cell fate regulation via deacetylation of the substrate proteins. However, whether SIRT1 deacetylates activator protein-1 (AP-1) to influence its transcriptional activity and target gene expression is still unknown. Here we demonstrate that SIRT1 directly interacts with the basic leucine zipper domains of c-Fos and c-Jun, the major components of AP-1, by which SIRT1 suppressed the transcriptional activity of AP-1. This process requires the deacetylase activity of SIRT1. Notably, SIRT1 reduced the expression of COX-2, a typical AP-1 target gene, and decreased prostaglandin E(2) (PGE(2)) production of peritoneal macrophages (pMPhis). pMPhis with SIRT1 overexpression displayed improved phagocytosis and tumoricidal functions, which are associated with depressed PGE(2). Furthermore, SIRT1 protein level was up-regulated in CR mouse pMPhis, whereas elevated SIRT1 decreased COX-2 expression and improved PGE(2)-related macrophage functions that were reversed following inhibition of SIRT1 deacetylase activity. Thus, our results indicate that SIRT1 may be a mediator of CR-induced macrophage regulation, and its deacetylase activity contributes to the inhibition of AP-1 transcriptional activity and COX-2 expression leading to amelioration of macrophage function.
Asunto(s)
Ciclooxigenasa 2/metabolismo , Regulación de la Expresión Génica , Macrófagos/fisiología , Sirtuina 1/metabolismo , Factor de Transcripción AP-1/metabolismo , Animales , Restricción Calórica , Línea Celular , Ciclooxigenasa 2/genética , Humanos , Leucina Zippers , Macrófagos/citología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Proto-Oncogénicas c-fos/química , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas Proto-Oncogénicas c-jun/química , Proteínas Proto-Oncogénicas c-jun/genética , Proteínas Proto-Oncogénicas c-jun/metabolismo , Distribución Aleatoria , Sirtuina 1/genética , Factor de Transcripción AP-1/genética , Transcripción GenéticaRESUMEN
The paraoxonase (PON) gene cluster consists of the PON1, PON2, and PON3 genes, each of which can individually inhibit atherogenesis. To analyze the functions of the PON gene cluster (PC) in atherogenesis and plaque stability, human PC transgenic (Tg) mice were generated using bacterial artificial chromosome. The high-density lipoprotein from Tg mice exhibited increased paraoxonase activity. When crossed to the ApoE-null background and challenged by high-fat diet, PC Tg/ApoE-null mice formed significantly fewer atherosclerotic lesions. However overexpression of the PC transgene had no additive effect on atherosclerosis compared to the overexpression of the single PON1 or PON3 transgene. Plaques from PC Tg/ApoE-null mice exhibited increased levels of collagen and smooth muscle cells, and reduced levels of macrophages and lipid, compared with those from ApoE-null mice, indicating lesions of PC Tg/ApoE-null mice had characteristics of more stable plaques than those of ApoE-null mice. PC transgene enhanced high-density lipoprotein ability to protect low-density lipoprotein against oxidation in vitro. Serum intercellular adhesion molecule-1 and monocyte chemoattractant protein-1 were also repressed by PC transgene. Proatherogenic reactions of Tg mouse peritoneal macrophages induced by oxidized low-density lipoprotein were inhibited by PC transgene, as indicated by reduced reactive oxygen species generation, inflammation, matrix metalloproteinase-9 expression, and foam cell formation. Our results demonstrate that the PC transgene not only represses atherogenesis but also promotes atherosclerotic plaque stability in vivo. PC may therefore be a useful target for atherosclerosis treatment.
Asunto(s)
Apolipoproteínas E/metabolismo , Arildialquilfosfatasa/genética , Aterosclerosis/fisiopatología , Familia de Multigenes/genética , Animales , Apolipoproteínas E/genética , Aterosclerosis/etiología , Aterosclerosis/genética , Quimiocina CCL2/sangre , Grasas de la Dieta/efectos adversos , Modelos Animales de Enfermedad , Humanos , Molécula 1 de Adhesión Intercelular/sangre , Lipoproteínas HDL/metabolismo , Lipoproteínas LDL/metabolismo , Ratones , Ratones Noqueados , Ratones Transgénicos , Oxidación-ReducciónRESUMEN
Angiotensin II (Ang II) stimulates vascular smooth muscle cell (VSMC) hypertrophy as a critical event in the development of vascular diseases such as atherosclerosis. Sirtuin (SIRT) 1, a nicotinamide adenine dinucleotide dependent deacetylase, has been demonstrated to exert protective effects in atherosclerosis by promoting endothelium-dependent vascular relaxation and reducing macrophage foam cell formation, but its role in VSMC hypertrophy remains unknown. In this study, we tried to investigate the effect of SIRT1 on Ang II-induced VSMC hypertrophy. Results showed that adenoviral-mediated over-expression of SIRT1 significantly inhibited Ang II-induced VSMC hypertrophy, while knockdown of SIRT1 by RNAi resulted in an increased [(3)H]-leucine incorporation of VSMC. Accordingly, nicotinamide adenine dinucleotide phosphate oxidase 1 (Nox1) expression induced by Ang II was inhibited by SIRT1 in VSMCs. SIRT1 activator resveratrol decreased, whereas endogenous SIRT1 inhibitor nicotinamide increased Nox1 expression in A7r5 VSMCs. Furthermore, transcription factor GATA-6 was involved in the down-regulation of Nox1 expression by SIRT1. These results provide new insight into SIRT1's anti-atherogenic properties by suppressing Ang II-induced VSMC hypertrophy.
Asunto(s)
Factor de Transcripción GATA6/metabolismo , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , NADH NADPH Oxidorreductasas/metabolismo , Sirtuina 1/metabolismo , Angiotensina II , Animales , Línea Celular , Factor de Transcripción GATA6/genética , Humanos , Hipertrofia/inducido químicamente , Hipertrofia/genética , Hipertrofia/metabolismo , Hipertrofia/patología , Ratones , NADH NADPH Oxidorreductasas/genética , NADPH Oxidasa 1 , Ratas , Sirtuina 1/genéticaRESUMEN
Producing two broods within the same season may be a good strategy by which short-lived species can maximize reproductive success. To produce two clutches in the same breeding season and to ensure offspring quality, choosing a good mate is important for females. Previous studies on double breeding focused on the associated influencing factors, and few studies examined how females choose social mates. Good genes and genetic compatibility are the two main hypotheses of the genetic benefit that females obtain from choosing mates. Uncovering the method used in mate choice for genetic benefits adopted by double-breeding females would provide a better understanding of the life history and rules of female choice. The great tit is an optionally double-breeding species in temperate-latitude populations. Here, we used a dataset for a Chinese population monitored between 2014 and 2016 to test two hypotheses on double-breeding female mate choice. A total of 30.1% of the breeding pairs initiated second breeding attempts, always remating with the same mate. The date of the first egg of the first brood did not affect initiation of a second brood, and female individual heterozygosity slightly influenced initiation of a second breeding. Female great tits choose males with both compatible genes and good genes in double-breeding mating. Double-breeding females prefer males with large breast stripes, high heterozygosity, and lower relatedness, while tarsus length, repertoire size, and individual F are not the main factors considered by females when selecting males for double breeding. The number of offspring of the first clutch did not affect the pairing status of male great tits in double breeding. The genetic quality of offspring from double-breeding pairs was higher than that of those from single-breeding pairs (higher heterozygosity and lower individual F). Taken together, our results showed that double breeding female great tits adopt multiple methods for genetic benefits to choose mates.
RESUMEN
Wnt signaling regulates cortical and hippocampal development. In a previous study we found that a particular Wnt receptor, Frizzled9 (Fzd9), was selectively expressed in both the developing and adult hippocampus. Taking advantage of the specificity of this promoter, we generated a transgenic cre mouse line using the putative control elements of the Fzd9 gene. In the Fzd9-cre mice, Cre is mainly detected in the developing cortex and hippocampus and is confined to the CA fields and dentate gyrus in adults. Furthermore, by crossing the Fzd9-cre mouse with the ROSA26 reporter line, we examined the activity of Cre and found that it has very high recombination efficiency. Thus, this mouse line will likely prove to be a useful tool for studying cortical and hippocampal development via activation or inactivation of interesting genes.
Asunto(s)
Corteza Cerebral/embriología , Embrión de Mamíferos/embriología , Hipocampo/embriología , Integrasas/metabolismo , Animales , Corteza Cerebral/metabolismo , Embrión de Mamíferos/metabolismo , Receptores Frizzled , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Hipocampo/metabolismo , Inmunohistoquímica , Integrasas/genética , Operón Lac/genética , Ratones , Ratones Transgénicos , Proteínas Asociadas a Microtúbulos/metabolismo , Regiones Promotoras Genéticas/genética , Proteínas/genética , Proteínas/metabolismo , ARN no Traducido , Receptores de Neurotransmisores/genética , Factores de Tiempo , beta-Galactosidasa/metabolismoRESUMEN
The proinflammatory cytokine TNF-alpha plays an important role in stimulating inflammatory responses of vascular smooth muscle cells (VSMCs). The anti-inflammatory function of Sirtuin 1 (SIRT1), a NAD-dependent class III histone/protein deacetylase, has been well documented, but how SIRT1 is regulated under inflammatory conditions is largely unknown. In the present research, we showed that levels of SIRT1 mRNA and protein expression increased in TNF-alpha-treated VSMCs. Overexpression of the p65/RelA subunit of NF-kappaB, a TNF-alpha-activated inflammatory transcription factor, in A7r5 cells, upregulated SIRT1 mRNA and protein expression as well as SIRT1 promoter activity, while knockdown of endogenous p65/RelA expression by RNAi not only led to a decrease in SIRT1's basal protein expression and promoter activity, but almost abolished the TNF-alpha-induced elevation of SIRT1 protein expression and SIRT1 promoter activity. Furthermore, using promoter deletion analysis and chromatin immunoprecipitation assays, we found that p65/RelA bound to the SIRT1 promoter at a consensus NF-kappaB binding site. Our study indicates that p65/RelA mediates the TNF-alpha-induced elevated expression of SIRT1 in VSMCs, shedding new light on the regulation of SIRT1 under inflammatory conditions.
Asunto(s)
Inflamación/metabolismo , Músculo Liso Vascular/metabolismo , Miocitos del Músculo Liso/metabolismo , Sirtuina 1/biosíntesis , Factor de Transcripción ReIA/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Humanos , Inflamación/genética , Músculo Liso Vascular/efectos de los fármacos , Miocitos del Músculo Liso/efectos de los fármacos , Biosíntesis de Proteínas , ARN Mensajero/biosíntesis , Sirtuina 1/genética , Factor de Transcripción ReIA/genética , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
Chronic persistent inflammation is thought to impede axon regeneration and cause demyelinating disease also with neuropathic pain, leading to more severe dysfunction after peripheral nerve injury. Increasing evidence indicates that neural stem cells (NSCs) have immunomodulatory effects, and previous studies have shown that many of the beneficial effects attributed to stem cell therapy may exert their therapeutic effects through paracrine mechanisms. In this research, the repairing effect of NSC-conditioned medium (NSC-CM) on sciatic nerve injury and its mechanism of repair were further explored. The present research showed that NSC-CM promoted histopathological and functional recovery after crush injury in rats, and what counts is that NSC-CM inhibited the inflammation of sciatic nerve in the late stage of injury. NSC-CM significantly downregulated the infiltration of proinflammatory factors [tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), and IL-1ß] as well as decreased the CD68 inflammatory macrophages infiltrating in the sciatic nerve. In addition, to study the effect of NSC-CM on the inflammatory state of macrophages in vitro, lipopolysaccharide (LPS) was used to induce the proinflammation of macrophages. The results showed that NSC-CM decreased the expression of macrophage proinflammatory-related proteins (IL-6, IL-1ß, TNF-α, inducible nitric oxide synthase) induced by LPS. The activation of Sirt-1 signaling in macrophages effectively countered the proinflammation induced by LPS in the presence of NSC-CM. Using Sirt-1-specific inhibitor EX527 partially weakened the anti-inflammatory effect of NSC-CM. Altogether, this study demonstrated for the first time that NSC-CM promotes functional recovery after sciatic nerve crush injury in vivo and also inhibits the inflammation in activated macrophages by activating Sirt-1 signaling pathway in vitro.
Asunto(s)
Medios de Cultivo Condicionados/farmacología , Inflamación/patología , Macrófagos/patología , Células-Madre Neurales/metabolismo , Recuperación de la Función , Nervio Ciático/lesiones , Transducción de Señal , Sirtuina 1/metabolismo , Animales , Axones/efectos de los fármacos , Axones/patología , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Mediadores de Inflamación/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , FN-kappa B/metabolismo , Regeneración Nerviosa/efectos de los fármacos , Fosforilación/efectos de los fármacos , Ratas Sprague-Dawley , Recuperación de la Función/efectos de los fármacos , Remielinización/efectos de los fármacos , Nervio Ciático/efectos de los fármacos , Nervio Ciático/patología , Nervio Ciático/fisiopatología , Transducción de Señal/efectos de los fármacosRESUMEN
OBJECTIVE: To study the morphological behavior, the expression of MCP-1 and PDGF-B of distal anastomotic neointima after expanded polytetrafluoroethylene (ePTFE) bypass grafting. And to identify the effects of Batroxobin (BX) on distal anastomotic intimal hyperplasia. METHODS: 12 adult mongrel dogs underwent end to side bypass grafting in left common carotid artery using ePTFE vascular graft (6mm in diameter, 5cm in length) were divided randomly into two groups. All 12 bypass grafts were removed 28 days after grafting and distal anastomotic parts were obtained. Using HE, Masson, IHC, RT-PCR and western blot, we analyzed the thickness, relative content of ECM, SMC PCNA index, expression of MCP-1 and PDGF-B in distal anastomotic neointima. And we compared the differences between the two groups. RESULTS: 28 day after grafting, the neointima was formed by SMC and ECM and was covered by epithelial cells. After using BX, the thickness of neointima was reduced [( 381.3 +/- 144.7) microm versus (213.8 +/- 29.0) microm, P < 0.05] and the relative content of ECM were significantly decreased (P = 0.006). The PCNA index (P = 0.109) and PDGF-B level (P = 0.055 by RT-PCR and P = 0.337 by WB) between the two groups had no significant difference. MCP-1 level was markedly decreased (P = 0.025 by RT-PCR and P = 0.016 by WB). CONCLUSIONS: Using BX after ePTFE grafting could effectively reduce the relative content of ECM, reduce the expression of MCP-1 and decrease the thickness of neointima in the early time.
Asunto(s)
Batroxobina/farmacología , Arteria Carótida Común/cirugía , Estomas Quirúrgicos/patología , Túnica Íntima/efectos de los fármacos , Túnica Íntima/patología , Anastomosis Quirúrgica , Animales , Prótesis Vascular , Quimiocina CCL2/metabolismo , Perros , Femenino , Hiperplasia , Masculino , Politetrafluoroetileno , Proteínas Proto-Oncogénicas c-sis/metabolismo , Procedimientos Quirúrgicos VascularesRESUMEN
AIMS: Hazardous environmental and genetic factors can damage endothelial cells to induce atherosclerotic vascular disease. Recent studies suggest that class III deacetylase SIRT1 may promote cell survival via novel antioxidative mechanisms. The current study tested the hypothesis that SIRT1, specifically overexpressed in the endothelium, is atheroprotective. METHODS AND RESULTS: Human umbilical vein endothelial cells (HUVECs) were used to study the effects of oxidized low-density lipoprotein (LDL) on SIRT1 expression. Endothelial cell-specific SIRT1 transgenic (SIRT1-Tg) mice were used to study the effects of SIRT1 on aortic vascular tone. SIRT1-Tg mice were crossed with apolipoprotein E null (apoE(-/-)) mice to obtain SIRT1-Tg/apoE(-/-) mice for the analysis of atherogenesis in the presence of endothelial overexpression of SIRT1. SIRT1 expression in HUVECs was increased by the treatment with oxidative LDL. Adenoviral-mediated overexpression of SIRT1 was protective of apoptosis of HUVECs. Calorie restriction increased, whereas high-fat diet decreased, the SIRT1 expression in mouse aortas. In SIRT1-Tg mice, high fat-induced impairment in endothelium-dependent vasorelaxation was improved compared with that of wild-type littermates. This was accompanied by an upregualtion of aortic endothelial nitric oxide synthase expression in the SIRT1-Tg mice. The SIRT1-Tg/apoE(-/-) mice had less atherosclerotic lesions compared with apoE(-/-) controls, without affecting blood lipids and glucose levels. CONCLUSION: These results suggest that endothelium-specific SIRT1 overexpression likely suppresses atherogenesis via improving endothelial cell survival and function.