RESUMO
BACKGROUND: Uterine adenosarcoma is a rare malignant tumor that accounts for 8% of all uterine sarcomas, and less than 0.2% of all uterine malignancies. However, it is frequently misdiagnosed in clinical examinations, including pathological diagnosis, and imaging studies owing to its rare and non-specific nature, which is further compounded by the lack of specific diagnostic markers. CASE PRESENTATION: We report a case of uterine adenosarcoma for which a comprehensive genomic profiling (CGP) test provided a chance to reach the proper diagnosis. The patient, a woman in her 60s with a history of uterine leiomyoma was diagnosed with an intra-abdominal mass post presentation with abdominal distention and loss of appetite. She was suspected to have gastrointestinal stromal tumor (GIST); the laparotomically excised mass was found to comprise uniform spindle-shaped cells that grew in bundles with a herringbone architecture, and occasional myxomatous stroma. Immunostaining revealed no specific findings, and the tumor was diagnosed as a spindle cell tumor/suspicious adult fibrosarcoma. The tumor relapsed during postoperative follow-up, and showed size reduction with chemotherapy, prior to regrowth. CGP was performed to identify a possible treatment, which resulted in detection of a JAZF1-BCORL1 rearrangement. Since the rearrangement has been reported in uterine sarcomas, we reevaluated specimens of the preceding uterine leiomyoma, which revealed the presence of adenosarcoma components in the corpus uteri. Furthermore, both the uterine adenosarcoma and intra-abdominal mass were partially positive for CD10 and BCOR staining. CONCLUSION: These results led to the conclusive identification of the abdominal tumor as a metastasis of the uterine adenosarcoma. The JAZF1-BCORL1 rearrangement is predominantly associated with uterine stromal sarcomas; thus far, ours is the second report of the same in an adenosarcoma. Adenosarcomas are rare and difficult to diagnose, especially in atypical cases with scarce glandular epithelial components. Identification of rearrangements involving BCOR or BCORL1, will encourage BCOR staining analysis, thereby potentially resulting in better diagnostic outcomes. Given that platinum-based chemotherapy was proposed as the treatment choice for this patient post diagnosis with adenosarcoma, CGP also indirectly contributed to the designing of the best-suited treatment protocol.
Assuntos
Adenossarcoma , Leiomioma , Neoplasias Uterinas , Feminino , Humanos , Adenossarcoma/diagnóstico , Adenossarcoma/genética , Adenossarcoma/patologia , Proteínas Correpressoras , Diagnóstico Diferencial , Proteínas de Ligação a DNA , Genômica , Leiomioma/diagnóstico , Proteínas Repressoras/genética , Neoplasias Uterinas/diagnóstico , Neoplasias Uterinas/genética , Neoplasias Uterinas/patologia , IdosoRESUMO
BACKGROUND: Heart failure (HF) is a common cardiovascular syndrome. Tanshinone IIA (Tan IIA) is a pharmacologically active monomer that exerts a significant cardioprotective effect in the clinic; however, the specific mechanisms are not fully understood. PURPOSE: We mainly investigated the protective effects of Tan IIA on doxorubicin (DOX)-induced HF. METHODS: In an in vitro study, H9C2 and HL-1 cells were cultured and treated with DOX and Tan IIA for 24 h, we investigated the mechanism underlying Tan IIA-mediated protection. In an in vivo study, a model of DOX-induced HF was established in C57BL/6 mice that were divided into the six groups randomly: a control group, a DOX group, DOX groups treated with Tan IIA (DOX+Tan IIA) at dosages of 2.5, 5 and 10 mg/kg/day and DOX groups treated with N-acetylcysteine (NAC) at dosages of 200 mg/kg/day. RESULT: The results demonstrated that Tan IIA significantly increased cell viability and protected against DOX-induced apoptosis. RNA-sequencing showed that the genes expression associated with the apoptotic signaling pathway was altered by Tan IIA. Among the differentially expressed genes, death-domain associated protein (DAXX), which plays an critical role in apoptotic signaling, exhibited increased expression under Tan IIA treatment. In addition, RNA interference was used to silence the expression of DAXX, which abolished Tan IIA-mediated protection against DOX-induced apoptosis; this effect was associated with extracellular signal-regulated protein kinase 1/2 (ERK1/2) and mitogen-activated protein kinase (MEK) expression. In the in vivo study, the echocardiography results revealed that heart function was rescued by Tan IIA, and the histomorphology results showed that Tan IIA prevented myocardial structural alteration and myofibril disruption. Furthermore, Tan IIA induced the expressions of DAXX, p-ERK1/2 and p-MEK. Tan IIA also inhibited apoptosis by suppressing the expression of cleaved caspase-8, p-P38 and cleaved caspase-3. CONCLUSION: Our results provide novel interpretations into the important role of DAXX in DOX-induced cardiotoxicity and show that Tan IIA may be a novel agent strategy for HF treatment via activating the DAXX/MEK/ERK1/2 pathway.
Assuntos
Abietanos , Cardiotoxicidade , Sistema de Sinalização das MAP Quinases , Animais , Camundongos , Abietanos/farmacologia , Acetilcisteína/farmacologia , Apoptose , Cardiotoxicidade/tratamento farmacológico , Caspase 3 , Caspase 8 , Proteínas Correpressoras , Doxorrubicina/efeitos adversos , Camundongos Endogâmicos C57BL , Quinases de Proteína Quinase Ativadas por Mitógeno , Chaperonas Moleculares/farmacologia , Miócitos Cardíacos , RNARESUMO
Phospholipids are vital membrane constituents that determine cell functions and interactions with the environment. For bacterial pathogens, rapid adjustment of phospholipid composition to changing conditions during infection can be crucial for growth and survival. Fatty acid synthesis (FASII) regulators are central to this process. This review puts the spotlight on FabT, a MarR-family regulator of FASII characterized in streptococci, enterococci, and lactococci. Roles of FabT in virulence, as reported in mouse and nonhuman primate infection models, will be discussed. We present FabT structure, the FabT regulon, and changes in FabT regulation according to growth conditions. A unique feature of FabT concerns its modulation by an unconventional corepressor, acyl-acyl-carrier protein (ACP). Some bacteria express two ACP proteins, which are distinguished by their interactions with endogenous or exogenous fatty acid sources, one of which causes strong FabT repression. This system seems to allow preferred use of environmental fatty acids, thereby saving energy by limiting futile FASII activity. Control of fabT expression and FabT activity link various metabolic pathways to FASII. The various physiological consequences of FabT loss summarized here suggest that FabT has potential as a narrow range therapeutic target.
Assuntos
Proteína de Transporte de Acila , Proteínas de Bactérias , Ácidos Graxos , Fatores de Transcrição , Proteína de Transporte de Acila/metabolismo , Animais , Bactérias/genética , Bactérias/patogenicidade , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas Correpressoras/metabolismo , Ácidos Graxos/biossíntese , Ácidos Graxos/genética , Regulação Bacteriana da Expressão Gênica , Camundongos , Fosfolipídeos/química , Fosfolipídeos/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Virulência/genéticaRESUMO
Rationale: We recently demonstrated that the 'Metabesity' factor HMG20A regulates islet beta-cell functional maturity and adaptation to physiological stress such as pregnancy and pre-diabetes. HMG20A also dictates central nervous system (CNS) development via inhibition of the LSD1-CoREST complex but its expression pattern and function in adult brain remains unknown. Herein we sought to determine whether HMG20A is expressed in the adult CNS, specifically in hypothalamic astrocytes that are key in glucose homeostasis and whether similar to islets, HMG20A potentiates astrocyte function in response to environmental cues. Methods: HMG20A expression profile was assessed by quantitative PCR (QT-PCR), Western blotting and/or immunofluorescence in: 1) the hypothalamus of mice exposed or not to either a high-fat diet or a high-fat high-sucrose regimen, 2) human blood leukocytes and adipose tissue obtained from healthy or diabetic individuals and 3) primary mouse hypothalamic astrocytes exposed to either high glucose or palmitate. RNA-seq and cell metabolic parameters were performed on astrocytes treated or not with a siHMG20A. Astrocyte-mediated neuronal survival was evaluated using conditioned media from siHMG20A-treated astrocytes. The impact of ORY1001, an inhibitor of the LSD1-CoREST complex, on HMG20A expression, reactive astrogliosis and glucose metabolism was evaluated in vitro and in vivo in high-fat high-sucrose fed mice. Results: We show that Hmg20a is predominantly expressed in hypothalamic astrocytes, the main nutrient-sensing cell type of the brain. HMG20A expression was upregulated in diet-induced obesity and glucose intolerant mice, correlating with increased transcript levels of Gfap and Il1b indicative of inflammation and reactive astrogliosis. Hmg20a transcript levels were also increased in adipose tissue of obese non-diabetic individuals as compared to obese diabetic patients. HMG20A silencing in astrocytes resulted in repression of inflammatory, cholesterol biogenesis and epithelial-to-mesenchymal transition pathways which are hallmarks of reactive astrogliosis. Accordingly, HMG20A depleted astrocytes exhibited reduced mitochondrial bioenergetics and increased susceptibility to apoptosis. Neuron viability was also hindered in HMG20A-depleted astrocyte-derived conditioned media. ORY1001 treatment rescued expression of reactive astrogliosis-linked genes in HMG20A ablated astrocytes while enhancing cell surface area, GFAP intensity and STAT3 expression in healthy astrocytes, mimicking the effect of HMG20A. Furthermore, ORY1001 treatment protected against obesity-associated glucose intolerance in mice correlating with a regression of hypothalamic HMG20A expression, indicative of reactive astrogliosis attenuation with improved health status. Conclusion: HMG20A coordinates the astrocyte polarization state. Under physiological pressure such as obesity and insulin resistance that induces low grade inflammation, HMG20A expression is increased to induce reactive astrogliosis in an attempt to preserve the neuronal network and re-establish glucose homeostasis. Nonetheless, a chronic metabesity state or functional mutations will result in lower levels of HMG20A, failure to promote reactive astrogliosis and increase susceptibility of neurons to stress-induced apoptosis. Such effects could be reversed by ORY1001 treatment both in vitro and in vivo, paving the way for a new therapeutic approach for Type 2 Diabetes Mellitus.
Assuntos
Astrócitos/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Gliose/metabolismo , Proteínas de Grupo de Alta Mobilidade/metabolismo , Hipotálamo/metabolismo , Neurônios/metabolismo , Obesidade/metabolismo , Tecido Adiposo/metabolismo , Adulto , Animais , Sobrevivência Celular/efeitos dos fármacos , Proteínas Correpressoras/antagonistas & inibidores , Dieta Hiperlipídica , Proteína Glial Fibrilar Ácida/metabolismo , Glucose/metabolismo , Proteínas de Grupo de Alta Mobilidade/antagonistas & inibidores , Proteínas de Grupo de Alta Mobilidade/genética , Histona Desmetilases/antagonistas & inibidores , Humanos , Interleucina-1beta/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Mitocôndrias/genética , Mitocôndrias/metabolismo , Proteínas do Tecido Nervoso/antagonistas & inibidores , RNA Interferente Pequeno , RNA-SeqRESUMO
The mammalian cochlea cannot regenerate functional hair cells (HCs) spontaneously. Atoh1 overexpression as well as other strategies are unable to generate functional HCs. Here, we simultaneously upregulated the expression of Gfi1, Pou4f3, and Atoh1 in postnatal cochlear supporting cells (SCs) in vivo, which efficiently converted SCs into HCs. The newly regenerated HCs expressed HC markers Myo7a, Calbindin, Parvalbumin, and Ctbp2 and were innervated by neurites. Importantly, many new HCs expressed the mature and terminal marker Prestin or vesicular glutamate transporter 3 (vGlut3), depending on the subtypes of the source SCs. Finally, our patch-clamp analysis showed that the new HCs in the medial region acquired a large K+ current, fired spikes transiently, and exhibited signature refinement of ribbon synapse functions, in close resemblance to native wild-type inner HCs. We demonstrated that co-upregulating Gfi1, Pou4f3, and Atoh1 enhances the efficiency of HC generation and promotes the functional maturation of new HCs.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Ligação a DNA/genética , Células Ciliadas Auditivas/metabolismo , Proteínas de Homeodomínio/genética , Células Labirínticas de Suporte/metabolismo , Organogênese/genética , Fator de Transcrição Brn-3C/genética , Fatores de Transcrição/genética , Potenciais de Ação/fisiologia , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Sistemas de Transporte de Aminoácidos Acídicos/genética , Sistemas de Transporte de Aminoácidos Acídicos/metabolismo , Animais , Animais Recém-Nascidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Calbindinas/genética , Calbindinas/metabolismo , Proteínas Correpressoras/genética , Proteínas Correpressoras/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Células Ciliadas Auditivas/citologia , Proteínas de Homeodomínio/metabolismo , Transporte de Íons , Células Labirínticas de Suporte/citologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Motores Moleculares/genética , Proteínas Motores Moleculares/metabolismo , Miosina VIIa/genética , Miosina VIIa/metabolismo , Neuritos/metabolismo , Neuritos/ultraestrutura , Parvalbuminas/genética , Parvalbuminas/metabolismo , Técnicas de Patch-Clamp , Potássio/metabolismo , Transdução de Sinais , Fator de Transcrição Brn-3C/metabolismo , Fatores de Transcrição/metabolismoRESUMO
FLICE-associated huge protein (FLASH), Yin Yang 1-Associated Protein-Related Protein (YARP) and Nuclear Protein, Ataxia-Telangiectasia Locus (NPAT) localize to discrete nuclear structures called histone locus bodies (HLBs) where they control various steps in histone gene expression. Near the C-terminus, FLASH and YARP contain a highly homologous domain that interacts with the C-terminal region of NPAT. Structural aspects of the FLASH-NPAT and YARP-NPAT complexes and their role in histone gene expression remain largely unknown. In this study, we used multidimensional NMR spectroscopy and in silico modeling to analyze the C-terminal domain in FLASH and YARP in an unbound form and in a complex with the last 31 amino acids of NPAT. Our results demonstrate that FLASH and YARP domains share the same fold of a triple α -helical bundle that resembles the DNA binding domain of Myb transcriptional factors and the SANT domain found in chromatin-modifying and remodeling complexes. The NPAT peptide contains a single α -helix that makes multiple contacts with α -helices I and III of the FLASH and YARP domains. Surprisingly, in spite of sharing a significant amino acid similarity, each domain likely binds NPAT using a unique network of interactions, yielding two distinct complexes. In silico modeling suggests that both complexes are structurally compatible with DNA binding, raising the possibility that they may function in identifying specific sequences within histone gene clusters, hence initiating the assembly of HLBs and regulating histone gene expression during cell cycle progression.
Assuntos
Proteínas Reguladoras de Apoptose/química , Proteínas de Ligação ao Cálcio/química , Proteínas de Ciclo Celular/química , Proteínas Correpressoras/química , Simulação por Computador , Proteínas de Ligação a DNA/química , Espectroscopia de Ressonância Magnética , Complexos Multiproteicos/química , Humanos , Conformação Proteica em alfa-Hélice , Domínios ProteicosRESUMO
Cochlear synaptopathy, the loss of synaptic connections between inner hair cells (IHCs) and auditory nerve fibers, has been documented in animal models of aging, noise, and ototoxic drug exposure, three common causes of acquired sensorineural hearing loss in humans. In each of these models, synaptopathy begins prior to changes in threshold sensitivity or loss of hair cells; thus, this underlying injury can be hidden behind a normal threshold audiogram. Since cochlear synaptic loss cannot be directly confirmed in living humans, non-invasive assays will be required for diagnosis. In animals with normal auditory thresholds, the amplitude of wave 1 of the auditory brainstem response (ABR) is highly correlated with synapse counts. However, synaptopathy can also co-occur with threshold elevation, complicating the use of the ABR alone as a diagnostic measure. Using an age-graded series of mice and a partial least squares regression approach to model structure-function relationships, this study shows that the combination of a small number of ABR and distortion product otoacoustic emission (DPOAE) measurements can predict synaptic ribbon counts at various cochlear frequencies to within 1-2 synapses per IHC of their true value. In contrast, the model, trained using the age-graded series of mice, overpredicted synapse counts in a small sample of young noise-exposed mice, perhaps due to differences in the underlying pattern of damage between aging and noise-exposed mice. These results provide partial validation of a noninvasive approach to identify synaptic/neuronal loss in humans using ABRs and DPOAEs.
Assuntos
Cóclea/patologia , Doenças Cocleares/diagnóstico , Potenciais Evocados Auditivos do Tronco Encefálico , Perda Auditiva Provocada por Ruído/diagnóstico , Sinapses/patologia , Estimulação Acústica , Fatores Etários , Oxirredutases do Álcool , Animais , Limiar Auditivo , Biomarcadores/metabolismo , Proteínas Correpressoras , Cóclea/metabolismo , Doenças Cocleares/metabolismo , Doenças Cocleares/patologia , Doenças Cocleares/fisiopatologia , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Perda Auditiva Provocada por Ruído/metabolismo , Perda Auditiva Provocada por Ruído/patologia , Perda Auditiva Provocada por Ruído/fisiopatologia , Imuno-Histoquímica , Análise dos Mínimos Quadrados , Aprendizado de Máquina , Masculino , Camundongos Endogâmicos CBA , Ruído , Fosfoproteínas/metabolismo , Valor Preditivo dos Testes , Receptores de AMPA/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Sinapses/metabolismoRESUMO
OBJECTIVE: With the exception of specific metabolic disorders, predictors of response to ketogenic dietary therapies (KDTs) are unknown. We aimed to determine whether common variation across the genome influences the response to KDT for epilepsy. METHODS: We genotyped individuals who were negative for glucose transporter type 1 deficiency syndrome or other metabolic disorders, who received KDT for epilepsy. Genotyping was performed with the Infinium HumanOmniExpressExome Beadchip. Hospital records were used to obtain demographic and clinical data. KDT response (≥50% seizure reduction) at 3-month follow-up was used to dissect out nonresponders and responders. We then performed a genome-wide association study (GWAS) in nonresponders vs responders, using a linear mixed model and correcting for population stratification. Variants with minor allele frequency <0.05 and those that did not pass quality control filtering were excluded. RESULTS: After quality control filtering, the GWAS of 112 nonresponders vs 123 responders revealed an association locus at 6p25.1, 61 kb upstream of CDYL (rs12204701, P = 3.83 × 10-8 , odds ratio [A] = 13.5, 95% confidence interval [CI] 4.07-44.8). Although analysis of regional linkage disequilibrium around rs12204701 did not strengthen the likelihood of CDYL being the candidate gene, additional bioinformatic analyses suggest it is the most likely candidate. SIGNIFICANCE: CDYL deficiency has been shown to disrupt neuronal migration and to influence susceptibility to epilepsy in mice. Further exploration with a larger replication cohort is warranted to clarify whether CDYL is the causal gene underlying the association signal.
Assuntos
Dieta Cetogênica/métodos , Epilepsia Resistente a Medicamentos/dietoterapia , Epilepsia Resistente a Medicamentos/genética , Farmacognosia , Criança , Pré-Escolar , Proteínas Correpressoras , Estudos de Coortes , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Genótipo , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Humanos , Hidroliases , Cooperação Internacional , Modelos Logísticos , Masculino , Polimorfismo de Nucleotídeo Único/genética , Proteínas/genética , Proteínas/metabolismoRESUMO
The study is aimed to construct high quality Salvia miltiorrhiza cDNA library and obtain the SmJAZ8 gene of S. miltiorrhiza by yeast two-hybrid system. In this studyï¼ full-length cDNA was synthesized from rootsï¼ stemsï¼ leavesï¼ flowers and hairy roots of S. miltiorrhiza. The full-length cDNA library was synthesized by SMART method and constructed with DSN homogenization technique. The results showed that the library capacity was 1.45×106ï¼ the recombination rate was 100%ï¼ and the average size of the insert was 500-2 000 bp. The recombinant vector of pDEST-pGADT7-SmJAZ8 was constructed and transformed into Y2HGold strain. The interaction protein was screened by yeast two-hybrid system. The DnaJ protein and UBQ protein were screened by yeast two-hybrid system. This study has successfully constructed a full-length cDNA library of S. miltiorrhizaï¼ and laid the foundation for the follow-up study on functional gene screening and gene function of S. miltiorrhiza.
Assuntos
Proteínas Correpressoras/genética , Biblioteca Gênica , Proteínas de Plantas/genética , Salvia miltiorrhiza/genética , Técnicas do Sistema de Duplo-Híbrido , DNA ComplementarRESUMO
We studied the role of the synaptic ribbon for sound encoding at the synapses between inner hair cells (IHCs) and spiral ganglion neurons (SGNs) in mice lacking RIBEYE (RBEKO/KO). Electron and immunofluorescence microscopy revealed a lack of synaptic ribbons and an assembly of several small active zones (AZs) at each synaptic contact. Spontaneous and sound-evoked firing rates of SGNs and their compound action potential were reduced, indicating impaired transmission at ribbonless IHC-SGN synapses. The temporal precision of sound encoding was impaired and the recovery of SGN-firing from adaptation indicated slowed synaptic vesicle (SV) replenishment. Activation of Ca2+-channels was shifted to more depolarized potentials and exocytosis was reduced for weak depolarizations. Presynaptic Ca2+-signals showed a broader spread, compatible with the altered Ca2+-channel clustering observed by super-resolution immunofluorescence microscopy. We postulate that RIBEYE disruption is partially compensated by multi-AZ organization. The remaining synaptic deficit indicates ribbon function in SV-replenishment and Ca2+-channel regulation.
Assuntos
Proteínas de Ligação a DNA/deficiência , Células Ciliadas Auditivas Internas/fisiologia , Audição , Fosfoproteínas/deficiência , Gânglio Espiral da Cóclea/citologia , Sinapses/fisiologia , Estimulação Acústica , Oxirredutases do Álcool , Animais , Proteínas Correpressoras , Camundongos , Camundongos Knockout , Microscopia Eletrônica , Microscopia de Fluorescência , Sinapses/ultraestruturaRESUMO
A major factor determining aluminium (Al) sensitivity in higher plants is the binding of Al to root cell walls. The Al binding capacity of cell walls is closely linked to the extent of pectin methylesterification, as the presence of methyl groups attached to the pectin backbone reduces the net negative charge of this polymer and hence limits Al binding. Despite recent progress in understanding the molecular basis of Al resistance in a wide range of plants, it is not well understood how the methylation status of pectin is mediated in response to Al stress. Here we show in Arabidopsis that mutants lacking the gene LEUNIG_HOMOLOG (LUH), a member of the Groucho-like family of transcriptional co-repressor, are less sensitive to Al-mediated repression of root growth. This phenotype is correlated with increased levels of methylated pectin in the cell walls of luh roots as well as altered expression of cell wall-related genes. Among the LUH-repressed genes, PECTIN METHYLESTERASE46 (PME46) was identified as reducing Al binding to cell walls and hence alleviating Al-induced root growth inhibition by decreasing PME enzyme activity. seuss-like2 (slk2) mutants responded to Al in a similar way as luh mutants suggesting that a LUH-SLK2 complex represses the expression of PME46. The data are integrated into a model in which it is proposed that PME46 is a major inhibitor of pectin methylesterase activity within root cell walls.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Hidrolases de Éster Carboxílico/metabolismo , Parede Celular/metabolismo , Proteínas Correpressoras/metabolismo , Pectinas/metabolismo , Raízes de Plantas/metabolismo , Proteínas Repressoras/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Hidrolases de Éster Carboxílico/genética , Proteínas Correpressoras/genética , Regulação da Expressão Gênica de Plantas , Raízes de Plantas/genética , Plantas Geneticamente Modificadas , Proteínas Repressoras/genéticaRESUMO
Atopic dermatitis (AD) is a common pruritic chronic skin disease. AD pathogenesis remains elusive, but may involve complex interplays among skin barrier dysfunction, Th2 inflammation, and pruritus. Current treatments for AD are still limited to symptomatic therapies. We previously showed that HR-1 hairless mice fed a special diet (HR-AD) develop AD-like symptoms; however, the ingredient(s) causing dermatitis remain unclear. In this study, we examined whether the deficiency of certain polyunsaturated fatty acids (PUFAs) was involved in the diet-induced AD pathogenesis. In the serum of HR-AD-fed mice, levels of linoleic acid (LA, 18:2n-6) and α-linolenic acid (ALA, 18:3n-3), as well as their metabolites, were markedly decreased. HR-AD-induced AD symptoms were significantly ameliorated by LA supplementation, and to a lesser extent by ALA supplementation. In addition, LA metabolites, such as γ-linolenic acid and arachidonic acid, had effects similar to those of LA. Further, using semi-purified custom diets, we attempted to reproduce HR-AD-induced AD symptoms. Unexpectedly, a deficiency in unsaturated fatty acids (UFAs) alone caused mild symptoms. However, several modifications of fat and carbohydrate components in the diet revealed that dietary deficiencies of UFA and cornstarch were required to fully induce severe AD symptoms. Furthermore, we examined the influence of genetic background on the development of diet-induced AD and found that a hypomorphic mutation in the hairless gene Hr, encoding a nuclear receptor (NR) corepressor, was essential for the complete development of diet-induced pruritic atopic skin. Thus, our findings suggest that certain PUFAs and NRs are new, potential therapeutic targets for treating AD.
Assuntos
Dermatite Atópica/etiologia , Dieta/efeitos adversos , Ácido Linoleico/deficiência , Animais , Proteínas Correpressoras/genética , Dermatite Atópica/genética , Dermatite Atópica/terapia , Modelos Animais de Doenças , Ácidos Graxos Insaturados/deficiência , Humanos , Ácido Linoleico/administração & dosagem , Ácido Linoleico/sangue , Camundongos Pelados , Terapia de Alvo Molecular , Mutação , Amido/efeitos adversos , Fatores de Transcrição/genéticaRESUMO
Dysregulation of histone methylation has emerged as a major driver of neurodevelopmental disorders including intellectual disabilities and autism spectrum disorders. Histone methyl writer and eraser enzymes generally act within multisubunit complexes rather than in isolation. However, it remains largely elusive how such complexes cooperate to achieve the precise spatiotemporal gene expression in the developing brain. Histone H3K4 methylation (H3K4me) is a chromatin signature associated with active gene-regulatory elements. We review a body of literature that supports a model in which the RAI1-containing H3K4me writer complex counterbalances the LSD1-containing H3K4me eraser complex to ensure normal brain development. This model predicts H3K4me as the nexus of previously unrelated neurodevelopmental disorders.
Assuntos
Encéfalo/metabolismo , Histonas/metabolismo , Anormalidades Múltiplas/genética , Animais , Transtornos Cromossômicos/genética , Duplicação Cromossômica/genética , Ritmo Circadiano/genética , Proteínas Correpressoras/genética , Expressão Gênica , Proteínas de Grupo de Alta Mobilidade/genética , Histona Desmetilases/genética , Histona-Lisina N-Metiltransferase/genética , Humanos , Metilação , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas do Tecido Nervoso/genética , Síndrome de Smith-Magenis/genética , Transativadores , Fatores de Transcrição/genéticaRESUMO
OBJECTIVE: Curcumin (capable of inhibiting angiogenic growth of human umbilical vein endothelial cells [HUVECs]), can be employed in vitro as a model of pathogenesis of corneal neovascularization (CRNV). The aim of this study was to explore regulatory mechanisms of microRNA (miR) levels after curcumin treatment. MATERIALS AND METHODS: Expression profiles of miRs in curcumin-treated HUVECs were investigated by miR microassay. Specific mimics and inhibitors of miR-1275 or miR-1246 were transfected into HUVECs. Then, their target genes, vascular endothelial growth factor B (VEGFB) and nuclear transcription factor kappa B acting protein (NKAP) were detected by quantitative real-time PCR, Western blotting assay or immunofluorescence assay. Cell proliferation and cell cycle parameters were measured with the help of CCK-8 assay and flow cytometry. RESULTS: MiR-1275 and miR-1246 expression levels were up-regulated by curcumin. Administration of the specific mimics and inhibitors of the two miRs led to significant changes in expression of VEGFB and NKAP as well as the indicators related to angiogenesis. Anti-angiogenic effect of curcumin depended on expression patterns of the two miRs in that inhibition of either miR interfered with the effect of curcumin. Furthermore, overexpression of NKAP interrupted effects of curcumin on the cells. CONCLUSION: Collectively, our findings demonstrate that curcumin inhibited HUVEC proliferation by up-regulation of miR-1275 and miR-1246.
Assuntos
Inibidores da Angiogênese/farmacologia , Neovascularização da Córnea/tratamento farmacológico , Curcumina/farmacologia , MicroRNAs/genética , Neovascularização Patológica/tratamento farmacológico , Regulação para Cima/efeitos dos fármacos , Proteínas Correpressoras/genética , Neovascularização da Córnea/genética , Neovascularização da Córnea/patologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Neovascularização Patológica/genética , Neovascularização Patológica/patologia , Proteínas Nucleares/genética , Proteínas RepressorasRESUMO
We recently found that the product of the AES gene functions as a metastasis suppressor of colorectal cancer (CRC) in both humans and mice. Expression of amino-terminal enhancer of split (AES) protein is significantly decreased in liver metastatic lesions compared with primary colon tumors. To investigate its downregulation mechanism in metastases, we searched for transcriptional regulators of AES in human CRC and found that its expression is reduced mainly by transcriptional dysregulation and, in some cases, by additional haploidization of its coding gene. The AES promoter-enhancer is in a typical CpG island, and contains a Yin-Yang transcription factor recognition sequence (YY element). In human epithelial cells of normal colon and primary tumors, transcription factor YY2, a member of the YY family, binds directly to the YY element, and stimulates expression of AES. In a transplantation mouse model of liver metastases, however, expression of Yy2 (and therefore of Aes) is downregulated. In human CRC metastases to the liver, the levels of AES protein are correlated with those of YY2. In addition, we noticed copy-number reduction for the AES coding gene in chromosome 19p13.3 in 12% (5/42) of human CRC cell lines. We excluded other mechanisms such as point or indel mutations in the coding or regulatory regions of the AES gene, CpG methylation in the AES promoter enhancer, expression of microRNAs, and chromatin histone modifications. These results indicate that Aes may belong to a novel family of metastasis suppressors with a CpG-island promoter enhancer, and it is regulated transcriptionally.
Assuntos
Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Ilhas de CpG/genética , Metástase Neoplásica/genética , Regiões Promotoras Genéticas/genética , Proteínas Repressoras/genética , Fatores de Transcrição/metabolismo , Animais , Linhagem Celular Tumoral , Proteínas Correpressoras , Metilação de DNA/genética , Regulação para Baixo , Genes Supressores de Tumor , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Camundongos , Mutação/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Elementos de Resposta/genética , Fatores de Transcrição/genética , Transcrição Gênica/genética , Fator de Transcrição YY1/metabolismoRESUMO
UNLABELLED: Noise-induced hearing loss (NIHL) is a major unresolved public health problem. Here, we investigate pathomechanisms of sensory hair cell death and suggest a novel target for protective intervention. Cellular survival depends upon maintenance of energy homeostasis, largely by AMP-activated protein kinase (AMPK). In response to a noise exposure in CBA/J mice, the levels of phosphorylated AMPKα increased in hair cells in a noise intensity-dependent manner. Inhibition of AMPK via siRNA or the pharmacological inhibitor compound C attenuated noise-induced loss of outer hair cells (OHCs) and synaptic ribbons, and preserved auditory function. Additionally, noise exposure increased the activity of the upstream AMPK kinase liver kinase B1 (LKB1) in cochlear tissues. The inhibition of LKB1 by siRNA attenuated the noise-increased phosphorylation of AMPKα in OHCs, reduced the loss of inner hair cell synaptic ribbons and OHCs, and protected against NIHL. These results indicate that noise exposure induces hair cell death and synaptopathy by activating AMPK via LKB1-mediated pathways. Targeting these pathways may provide a novel route to prevent NIHL. SIGNIFICANCE STATEMENT: Our results demonstrate for the first time that the activation of AMP-activated protein kinase (AMPK) α in sensory hair cells is noise intensity dependent and contributes to noise-induced hearing loss by mediating the loss of inner hair cell synaptic ribbons and outer hair cells. Noise induces the phosphorylation of AMPKα1 by liver kinase B1 (LKB1), triggered by changes in intracellular ATP levels. The inhibition of AMPK activation by silencing AMPK or LKB1, or with the pharmacological inhibitor compound C, reduced outer hair cell and synaptic ribbon loss as well as noise-induced hearing loss. This study provides new insights into mechanisms of noise-induced hearing loss and suggests novel interventions for the prevention of the loss of sensory hair cells and cochlear synaptopathy.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Cóclea/patologia , Células Ciliadas Auditivas/patologia , Perda Auditiva Provocada por Ruído/etiologia , Perda Auditiva Provocada por Ruído/patologia , Ruído/efeitos adversos , Estimulação Acústica , Oxirredutases do Álcool , Animais , Morte Celular/efeitos dos fármacos , Proteínas Correpressoras , Cóclea/efeitos dos fármacos , Proteínas de Ligação a DNA/metabolismo , Relação Dose-Resposta a Droga , Vias de Administração de Medicamentos , Inibidores Enzimáticos/uso terapêutico , Potenciais Evocados Auditivos do Tronco Encefálico/efeitos dos fármacos , Potenciais Evocados Auditivos do Tronco Encefálico/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Células Ciliadas Auditivas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos CBA , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Pirazóis/uso terapêutico , Pirimidinas/uso terapêutico , RNA Interferente Pequeno/uso terapêutico , Receptores de AMPA/metabolismo , Sinapses/efeitos dos fármacos , Sinapses/fisiologiaRESUMO
In principle, the inhibition of candidate gain-of-function genes defined through genomic analyses of large patient cohorts offers an attractive therapeutic strategy. In this study, we focused on changes in expression of CD24, a well-validated clinical biomarker of poor prognosis and a driver of tumor growth and metastasis, as a benchmark to assess functional relevance. Through this approach, we identified GON4L as a regulator of CD24 from screening a pooled shRNA library of 176 candidate gain-of-function genes. GON4L depletion reduced CD24 expression in human bladder cancer cells and blocked cell proliferation in vitro and tumor xenograft growth in vivo Mechanistically, GON4L interacted with transcription factor YY1, promoting its association with the androgen receptor to drive CD24 expression and cell growth. In clinical bladder cancer specimens, expression of GON4L, YY1, and CD24 was elevated compared with normal bladder urothelium. This pathway is biologically relevant in other cancer types as well, where CD24 and the androgen receptor are clinically prognostic, given that silencing of GON4L and YY1 suppressed CD24 expression and growth of human lung, prostate, and breast cancer cells. Overall, our results define GON4L as a novel driver of cancer growth, offering new biomarker and therapeutic opportunities. Cancer Res; 76(17); 5175-85. ©2016 AACR.
Assuntos
Antígeno CD24/metabolismo , Carcinoma de Células de Transição/patologia , Receptores Androgênicos/metabolismo , Fatores de Transcrição/metabolismo , Neoplasias da Bexiga Urinária/patologia , Fator de Transcrição YY1/metabolismo , Animais , Western Blotting , Carcinoma de Células de Transição/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Proteínas Correpressoras , Proteínas de Ligação a DNA , Feminino , Técnicas de Silenciamento de Genes , Xenoenxertos , Humanos , Imunoprecipitação , Masculino , Camundongos , Camundongos Nus , Neoplasias/metabolismo , Neoplasias/patologia , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase , Neoplasias da Bexiga Urinária/metabolismoRESUMO
Functional maturation of afferent synaptic connections to inner hair cells (IHCs) involves pruning of excess synapses formed during development, as well as the strengthening and survival of the retained synapses. These events take place during the thyroid hormone (TH)-critical period of cochlear development, which is in the perinatal period for mice and in the third trimester for humans. Here, we used the hypothyroid Snell dwarf mouse (Pit1(dw)) as a model to study the role of TH in afferent type I synaptic refinement and functional maturation. We observed defects in afferent synaptic pruning and delays in calcium channel clustering in the IHCs of Pit1(dw) mice. Nevertheless, calcium currents and capacitance reached near normal levels in Pit1(dw) IHCs by the age of onset of hearing, despite the excess number of retained synapses. We restored normal synaptic pruning in Pit1(dw) IHCs by supplementing with TH from postnatal day (P)3 to P8, establishing this window as being critical for TH action on this process. Afferent terminals of older Pit1(dw) IHCs showed evidence of excitotoxic damage accompanied by a concomitant reduction in the levels of the glial glutamate transporter, GLAST. Our results indicate that a lack of TH during a critical period of inner ear development causes defects in pruning and long-term homeostatic maintenance of afferent synapses.
Assuntos
Cóclea/crescimento & desenvolvimento , Células Ciliadas Auditivas Internas/fisiologia , Células Ciliadas Auditivas Internas/ultraestrutura , Sinapses/fisiologia , Sinapses/ultraestrutura , Tri-Iodotironina/fisiologia , Oxirredutases do Álcool , Animais , Canais de Cálcio Tipo L/metabolismo , Proteínas Correpressoras , Cóclea/efeitos dos fármacos , Cóclea/ultraestrutura , Proteínas de Ligação a DNA/metabolismo , Transportador 1 de Aminoácido Excitatório/metabolismo , Células Ciliadas Auditivas Internas/efeitos dos fármacos , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/metabolismo , Fosfoproteínas/metabolismo , Sinapses/efeitos dos fármacos , Fator de Transcrição Pit-1/genética , Tri-Iodotironina/administração & dosagemRESUMO
Recent studies have reported that noise exposure at relatively low intensities can cause temporary threshold shifts (TTS) in hearing. However, the mechanism underlying the TTS is still on debate. Here, we report that an acoustic stimulation (100 dB SPL, white noise) induced TTS in mice, with the maximal ABR threshold elevations seen on the 4(th) day after noise exposure. On the other hand, there were no significant morphological changes in the cochlea. Further, there were paralleled changes of pre-synaptic ribbons in both the number and postsynaptic density (PSDs) during this noise exposure. The numbers of presynaptic ribbon, postsynaptic density (PSDs), and colocalized puncta correlated with the shifts of ABR thresholds. Moreover, a complete recovery of ABR thresholds and synaptic puncta was seen on the 14(th) day after the noise stimulations. Thus, our study may indicate that noise exposure can cause a decline in cochlear ribbon synapses and result in consequent hearing loss. The reduction of synaptic puncta appears reversible and may contribute to hearing restoration in mice after noise exposure.
Assuntos
Fadiga Auditiva , Células Ciliadas Auditivas Internas , Perda Auditiva Provocada por Ruído/fisiopatologia , Plasticidade Neuronal , Sinapses , Estimulação Acústica , Oxirredutases do Álcool , Animais , Proteínas Correpressoras , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Células Ciliadas Auditivas Internas/metabolismo , Células Ciliadas Auditivas Internas/ultraestrutura , Perda Auditiva Provocada por Ruído/etiologia , Perda Auditiva Provocada por Ruído/metabolismo , Perda Auditiva Provocada por Ruído/patologia , Camundongos Endogâmicos C57BL , Microscopia Eletrônica de Varredura , Ruído , Emissões Otoacústicas Espontâneas , Fosfoproteínas/metabolismo , Recuperação de Função Fisiológica , Sinapses/metabolismo , Sinapses/ultraestrutura , Fatores de TempoRESUMO
Hearing loss among the elderly correlates with diminished social, mental, and physical health. Age-related cochlear cell death does occur, but growing anatomical evidence suggests that synaptic rearrangements on sensory hair cells also contribute to auditory functional decline. Here we present voltage-clamp recordings from inner hair cells of the C57BL/6J mouse model of age-related hearing loss, which reveal that cholinergic synaptic inputs re-emerge during aging. These efferents are functionally inhibitory, using the same ionic mechanisms as do efferent contacts present transiently before the developmental onset of hearing. The strength of efferent inhibition of inner hair cells increases with hearing threshold elevation. These data indicate that the aged cochlea regains features of the developing cochlea and that efferent inhibition of the primary receptors of the auditory system re-emerges with hearing impairment. SIGNIFICANCE STATEMENT: Synaptic changes in the auditory periphery are increasingly recognized as important factors in hearing loss. To date, anatomical work has described the loss of afferent contacts from cochlear hair cells. However, relatively little is known about the efferent innervation of the cochlea during hearing loss. We performed intracellular recordings from mouse inner hair cells across the lifespan and show that efferent innervation of inner hair cells arises in parallel with the loss of afferent contacts and elevated hearing threshold during aging. These efferent neurons inhibit inner hair cells, raising the possibility that they play a role in the progression of age-related hearing loss.