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The major challenges in pancreatic ductal adenocarcinoma (PDAC) management are local or distant metastasis and limited targeted therapeutics to prevent it. To identify a druggable target in tumor secretome and to explore its therapeutic intervention, we performed a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic analysis of tumors obtained from a patient-derived xenograft model of PDAC. Galectin-3 binding protein (Gal-3BP) is identified as a highly secreted protein, and its overexpression is further validated in multiple PDAC tumors and primary cells. Knockdown and exogenous treatment of Gal-3BP showed that it is required for PDAC cell proliferation, migration, and invasion. Mechanistically, we revealed that Gal-3BP enhances galectin-3-mediated epidermal growth factor receptor signaling, leading to increased cMyc and epithelial-mesenchymal transition. To explore the clinical impact of these findings, two antibody clones were developed, and they profoundly abrogated the metastasis of PDAC cells in vivo. Altogether, our data demonstrate that Gal-3BP is an important therapeutic target in PDAC, and we propose its blockade by antibody as a therapeutic option for suppressing PDAC metastasis.
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Antígenos de Neoplasias , Antineoplásicos Imunológicos , Biomarcadores Tumorais , Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Animais , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Antineoplásicos Imunológicos/imunologia , Antineoplásicos Imunológicos/uso terapêutico , Biomarcadores Tumorais/antagonistas & inibidores , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/secundário , Carcinoma Ductal Pancreático/terapia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Cromatografia Líquida , Transição Epitelial-Mesenquimal , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/terapia , Proteômica , Secretoma , Espectrometria de Massas em Tandem , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The nucleus of the solitary tract (NTS) receives direct viscerosensory vagal afferent input that drives autonomic reflexes, neuroendocrine function and modulates behaviour. A subpopulation of NTS neurons project to the nucleus accumbens (NAc); however, the function of this NTS-NAc pathway remains unknown. A combination of neuroanatomical tracing, slice electrophysiology and fibre photometry was used in mice and/or rats to determine how NTS-NAc neurons fit within the viscerosensory network. NTS-NAc projection neurons are predominantly located in the medial and caudal portions of the NTS with 54 ± 7% (mice) and 65 ± 3% (rat) being TH-positive, representing the A2 NTS cell group. In horizontal brainstem slices, solitary tract (ST) stimulation evoked excitatory post-synaptic currents (EPSCs) in NTS-NAc projection neurons. The majority (75%) received low-jitter, zero-failure EPSCs characteristic of monosynaptic ST afferent input that identifies them as second order to primary sensory neurons. We then examined whether NTS-NAc neurons respond to cholecystokinin (CCK, 20 µg/kg ip) in vivo in both mice and rats. Surprisingly, there was no difference in the number of activated NTS-NAc cells between CCK and saline-treated mice. In rats, just 6% of NTS-NAc cells were recruited by CCK. As NTS TH neurons are the primary source for NAc noradrenaline, we measured noradrenaline release in the NAc and showed that NAc noradrenaline levels declined in response to cue-induced reward retrieval but not foot shock. Combined, these findings suggest that high-fidelity afferent information from viscerosensory afferents reaches the NAc. These signals are likely unrelated to CCK-sensitive vagal afferents but could interact with other sensory and higher order inputs to modulate learned appetitive behaviours.
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OBJECTIVE: Long-term protective effects of menopausal hormone therapy (MHT) at fractures with different doses and components are controversial. We analyzed the effect of MHT on the incidence of spine and femur fractures according to MHT type, age at commencement, duration and dose of hormones in Korean women. METHOD: This retrospective study evaluated propensity score-matched patients with MHT from the Korean National Health Insurance Service database. Among women aged ≥50 years with menopause between 2004 and 2007, spine and femur fracture incidence until 2017 was analyzed in 36,446 women who had received MHT for >1 year. Estrogen-progesterone therapy (EPT), estrogen-only therapy (ET) or tibolone therapy was conducted. RESULTS: EPT significantly lowered the incidence of spine and femur fractures with a conventional dose, but not with a low dose. Tibolone significantly decreased the incidence of spine fractures in women aged 50-59 years when used for >5 years, and the incidence of femur fractures in women older than 60 years when used for >3 years. ET significantly lowered the risk of femur fractures when estradiol was used for >5 years. CONCLUSION: In menopausal women, all MHT including conventional-dose EPT, ET and tibolone tended to lower the incidence of fractures. The effects, however, varied with the type of fracture and type of MHT.
Assuntos
Menopausa , Pós-Menopausa , Feminino , Humanos , Incidência , Estudos Retrospectivos , Terapia de Reposição Hormonal , Estrogênios/farmacologia , Progesterona/farmacologia , Terapia de Reposição de EstrogêniosRESUMO
OBJECTIVE: This study aimed to investigate the association of hormone replacement therapy (HRT) use, type, duration and age of commencement with myocardial infarction (MI) and stroke in postmenopausal Korean women. METHODS: This nested case-control study used data from the National Health Insurance Service database to analyze 2017 data from women aged ≥50 years and diagnosed with natural menopause between 2004 and 2007. Among 356,160 eligible women, 36,446 used HRT for ≥1 year and 319,714 did not (controls). These two groups were matched 1:1 for statistical analysis. Type and duration were categorized into three categories. RESULTS: Women who started estrogen-progestogen therapy (EPT) or estrogen therapy (ET) in their 50s, or EPT or tibolone in their ≥60s exhibited a lower stroke risk than controls. MI risk was lower among women who used tibolone - regardless of duration - or EPT or ET for 1-3 years than among controls. Stroke risk was lower with tibolone use for ≥5 years or with EPT or ET use for 1-3 years or ≥5 years than non-users. CONCLUSION: Our study may support the beneficial effect of HRT by showing that Korean postmenopausal women who used HRT at a relatively younger and healthier age had a relative benefit for MI and stroke.
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Terapia de Reposição de Estrogênios , Infarto do Miocárdio , Norpregnenos , Pós-Menopausa , Acidente Vascular Cerebral , Humanos , Feminino , Infarto do Miocárdio/epidemiologia , Pessoa de Meia-Idade , República da Coreia/epidemiologia , Estudos de Casos e Controles , Acidente Vascular Cerebral/epidemiologia , Norpregnenos/efeitos adversos , Terapia de Reposição de Estrogênios/efeitos adversos , Idoso , Fatores Etários , Bases de Dados Factuais , Fatores de Risco , Terapia de Reposição Hormonal/efeitos adversosRESUMO
Decisions to act while pursuing goals in the presence of danger must be made quickly but safely. Premature decisions risk injury or death, whereas postponing decisions risk goal loss. Here we show how mice resolve these competing demands. Using microstructural behavioral analyses, we identified the spatiotemporal dynamics of approach-avoidance decisions under motivational conflict in male mice. Then we used cognitive modeling to show that these dynamics reflect the speeded decision-making mechanisms used by humans and nonhuman primates, with mice trading off decision speed for safety of choice when danger loomed. Using calcium imaging in paraventricular thalamus and optogenetic inhibition of the prelimbic cortex to paraventricular thalamus pathway, we show that this speed-safety trade off occurs because increases in paraventricular thalamus activity increase decision caution, thereby increasing approach-avoid decision times in the presence of danger. Our findings demonstrate that a discrete brain circuit involving the paraventricular thalamus and its prefrontal input adjusts decision caution during motivational conflict, trading off decision speed for decision safety when danger is close. We identify the corticothalamic pathway as central to cognitive control during decision-making under conflict.SIGNIFICANCE STATEMENT Foraging animals balance the need to seek food and energy against the conflicting needs to avoid injury and predation. This competition is fundamental to survival but rarely has a stable, correct solution. Here we show that approach-avoid decisions under motivational conflict involve strategic adjustments in decision caution controlled via a top-down corticothalamic pathway from the prelimbic cortex to the paraventricular thalamus. We identify a novel corticothalamic mechanism for cognitive control that is applicable across a range of motivated behaviors and mark paraventricular thalamus and its prefrontal cortical input as targets to remediate the deficits in decision caution characteristic of unsafe and impulsive choices.
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Motivação , Tálamo , Animais , Tomada de Decisões/fisiologia , Comportamento Impulsivo , Masculino , Camundongos , Córtex Pré-Frontal , RecompensaRESUMO
OBJECTIVE: The identification of allergic rhinitis (AR) in early life is important for the target of intervention. AR is caused by various environmental factors, including house dust mites. We investigated the relationship between the Dermatophagoides farinae (Der f)-IgE and eosinophil in mothers with AR at delivery and the eosinophil levels and AR incidence in children. METHODS: The study participants were 983 mother-child pairs from the COhort for Childhood Origin of Asthma and Allergic Diseases. AR was diagnosed by a doctor at delivery in mother and at 3 years of age in offspring. The association between eosinophil level and AR was assessed using logistic regression analysis. RESULTS: The Der f-IgE level in mother having AR at delivery was associated with the mother's eosinophil level, and the mother's eosinophil level was associated with the child's eosinophil level both at age 1 and 3. The risk of AR at age 3 in children was increased according to increased eosinophil levels in mothers at delivery and in children both aged 1 and 3 years (adjusted odds ratio [aOR] and 95% confidence interval [CI]: 2.57 [1.14-5.78], 2.28 [1.02-5.13], respectively). The risk of childhood AR at the age of 3 is increased when both mothers and children have high eosiniophils (aOR and 95% CI: 2.62 [1.01-6.79], 1.37 [0.98-1.91]). CONCLUSIONS: Der f-IgE in mothers at delivery was related to eosinophil levels in mothers with AR and higher level of eosinophils in both mother and children was associated with the increased risk of AR incidence at the first 3 years of life of children.
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Asma , Rinite Alérgica , Feminino , Humanos , Lactente , Pré-Escolar , Eosinófilos , Incidência , Imunoglobulina E , Rinite Alérgica/epidemiologia , Asma/epidemiologia , Asma/etiologia , Asma/diagnósticoRESUMO
BACKGROUND: Single nucleotide polymorphisms (SNPs) are reportedly associated with repeated abortion. Thus, genetic analysis based on race is the key to developing accurate diagnostic tests. This study analyzed the genetic polymorphisms of recurrent pregnancy loss (RPL) patients among Korean women compared to the controls. METHODS: In 53 women of RPL group and 50 controls, the genetic analysis was performed. The genotype distribution and allele frequency were analyzed statistically for the difference between the two groups. The association between each SNP marker and RPL risk was analyzed. RESULTS: The genotypes of LEPR, endothelial nitric oxide synthase (eNOS), KDR, miR-27a, miR-449b, and tumor necrosis factor-alpha (TNF-α) were analyzed using odds ratio (OR) with 95% confidence intervals (CIs). Only the AG genotype of miR-449b (A>G) polymorphism showed significant association with the risk of RPL when compared to the AA genotype (OR, 2.39). The combination of GG/AG+GG/CA+AA genotypes for eNOS/miR-449b/TNF-α was associated with 7.36-fold higher risk of RPL (OR, 7.36). The GG/AG+GG combination for eNOS/miR-449b showed 2.43-fold higher risk for RPL (OR, 2.43). The combination of AG+GG/CA+AA genotypes for miR-449b/TNF-α showed a significant association with the risk of RPL (OR, 7.60). From the haplotype-based analysis, the G-G-A haplotype of eNOS/miR-449b/TNF-α and the G-A haplotype of miR-449b/TNF-α were associated with increased risk of RPL (OR, 19.31; OR, 22.08, respectively). CONCLUSION: There is a significant association between the risk of RPL and miR-449b/TNF-α combination, and therefore, genetic analysis for specific combined genotypes can be an important screening method for RPL in Korean women.
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Aborto Habitual , MicroRNAs , Gravidez , Humanos , Feminino , Polimorfismo de Nucleotídeo Único , Predisposição Genética para Doença , Fator de Necrose Tumoral alfa/genética , Genótipo , Aborto Habitual/diagnóstico , Aborto Habitual/genética , Biomarcadores , MicroRNAs/genética , República da Coreia , Estudos de Casos e ControlesRESUMO
PURPOSE: This study aimed to investigate the association between metformin usage and the risk of colorectal cancer (CRC) using data from the Korean National Health Insurance Service-National Health Screening Cohort database. METHODS: Data from the NHIS-HEALS cohort between 2002 and 2015 were longitudinally analyzed. Subjects were divided into three groups: metformin non-users with diabetes mellitus (DM), metformin users with DM, and no DM group. CRC was defined using the ICD-10 code (C18.0-C20.0) at the time of admission. Cox proportional hazard regression models were adopted after stepwise adjustment for confounders to investigate the association between metformin usage and colorectal cancer risk. RESULTS: During the follow-up period, of the total 323,430 participants, 2341 (1.33%) of the 175,495 males and 1204 (0.81%) of the 147,935 females were newly diagnosed with CRC. The estimated cumulative incidence of CRC was significantly different among the three groups based on Kaplan-Meier's survival curve (p values < 0.05 in both sexes). Compared with metformin non-users, hazard ratios (95% CIs) of metformin users and the no DM group were 0.66 (0.51-0.85) and 0.72 (0.61-0.85) in males and 0.59 (0.37-0.92) and 0.93 (0.66-1.29) in females, respectively, after being fully adjusted. CONCLUSIONS: Metformin users with diabetes appear to have a significantly lower risk of CRC compared with metformin non-users.
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Neoplasias Colorretais , Diabetes Mellitus Tipo 2 , Diabetes Mellitus , Metformina , Estudos de Coortes , Neoplasias Colorretais/epidemiologia , Diabetes Mellitus/tratamento farmacológico , Diabetes Mellitus/epidemiologia , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/epidemiologia , Feminino , Humanos , Hipoglicemiantes/uso terapêutico , Incidência , Masculino , Metformina/uso terapêutico , Modelos de Riscos Proporcionais , Fatores de RiscoRESUMO
Decision-making often involves motivational conflict because of the competing demands of approach and avoidance for a common resource: behavior. This conflict must be resolved as a necessary precursor for adaptive behavior. Here we show a role for the paraventricular thalamus (PVT) in behavioral control during motivational conflict. We used Pavlovian counterconditioning in male rats to establish a conditioned stimulus (CS) as a signal for reward (or danger) and then transformed the same CS into a signal for danger (or reward). After such training, the CS controls conflicting appetitive and aversive behaviors. To assess PVT involvement in conflict, we injected an adeno-associated virus (AAV) expressing the genetically encoded Ca2+ indicator GCaMP and used fiber photometry to record population PVT Ca2+ signals. We show distinct profiles of responsivity across the anterior-posterior axis of PVT during conflict, including an ordinal relationship between posterior PVT CS responses and behavior strength. To study the causal role of PVT in behavioral control during conflict, we injected AAV expressing the inhibitory hM4Di DREADD and determined the effects of chemogenetic PVT inhibition on behavior. We show that chemogenetic inhibition across the anterior-posterior axis of the PVT, but not anterior or posterior PVT alone, disrupts arbitration between appetitive and aversive behaviors when they are in conflict but has no effect when these behaviors are assessed in isolation. Together, our findings identify PVT as central to behavioral control during motivational conflict.SIGNIFICANCE STATEMENT Animals, including humans, approach attractive stimuli and avoid aversive ones. However, they frequently face conflict when the demands of approach and avoidance are incompatible. Resolution of this conflict is fundamental to adaptive behavior. Here we show a role for the paraventricular thalamus, a nucleus of the dorsal midline thalamus, in the arbitration of appetitive and aversive behavior during motivational conflict.
Assuntos
Condicionamento Operante/fisiologia , Tomada de Decisões/fisiologia , Núcleos da Linha Média do Tálamo/fisiologia , Motivação/fisiologia , Animais , Sinais (Psicologia) , Masculino , Atividade Motora/fisiologia , Ratos , Ratos Sprague-Dawley , RecompensaRESUMO
BACKGROUND: Diabetes mellitus (DM) increases atherosclerotic cardiovascular complications and cancer risks. Stomach cancer is the most common cancer in Korea. Although the survival rate of stomach cancer has improved, the disease burden is still high. METHODS: This retrospective study investigated the association between metformin use and stomach cancer incidence in a Korean population using the National Health Insurance Service-National Health Screening Cohort database. Participants aged 40-80 years old at the baseline period (2002-2003) were enrolled. The study population was categorized into three groups of metformin non-users with DM, metformin users with DM, and individuals without DM (No DM group). RESULTS: A total of 347,895 participants (14,922 metformin non-users, 9891 metformin users, and 323,082 individuals without DM) were included in the final analysis. The median follow-up duration was 12.70 years. The estimated cumulative incidence of stomach cancer was highest in metformin non-users and lowest in the No DM group (men vs. women: 3.75 vs. 1.97% in metformin non-users, 2.91 vs. 1.53% in metformin users, and 2.54 vs. 0.95% in the No DM group). Compared with metformin non-users, the hazard ratios (95% confidence intervals) for stomach cancer incidence of metformin users and the No DM group were 0.710 (0.579-0.870) and 0.879 (0.767-1.006) in men and 0.700 (0.499-0.981) and 0.701 (0.544-0.903) in women, respectively, after full adjustment. CONCLUSIONS: Metformin users with DM in the Korean population were at lower risk of stomach cancer incidence after controlling for potential confounding factors.
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Complicações do Diabetes/epidemiologia , Diabetes Mellitus/tratamento farmacológico , Hipoglicemiantes/uso terapêutico , Metformina/uso terapêutico , Neoplasias Gástricas/epidemiologia , Adulto , Bases de Dados Factuais , Complicações do Diabetes/prevenção & controle , Detecção Precoce de Câncer/estatística & dados numéricos , Feminino , Humanos , Incidência , Masculino , Programas Nacionais de Saúde , República da Coreia/epidemiologia , Estudos Retrospectivos , Fatores de Risco , Neoplasias Gástricas/etiologia , Neoplasias Gástricas/prevenção & controleRESUMO
BACKGROUND AND AIMS: Cancer is the number one cause of death in Korea. This study aimed to investigate if statin use in cancer survivors was inversely associated with all-cause mortality. METHODS AND RESULTS: Data from the 2002 to 2015 National Health Insurance Service-National Health Screening Cohort (NHIS-HEALS) were used. The Kaplan-Meier estimator was used to estimate the survival function according to statin usage. Cox proportional hazards regression models were adopted after stepwise adjustment for potential confounders to calculate hazard ratios (HRs) and 95% confidence intervals (CIs) for all-cause mortality. The median follow-up duration was 10.0 years. Statin users had a higher percentage of diabetes and hypertension in both sexes. Survival rates of statin users were higher than non-users (p-values <0.001 in men and 0.021 in women). Compared to non-users, the HRs (95% CIs) of statin users for all-cause mortality were 0.327 (0.194-0.553) in men and 0.287 (0.148-0.560) in women after adjustment for potential confounding factors. CONCLUSIONS: Statin users in cancer survivors had higher survival rate than non-users in both sexes.
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Sobreviventes de Câncer , Doenças Cardiovasculares/prevenção & controle , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Neoplasias/terapia , Adulto , Idoso , Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/mortalidade , Causas de Morte , Bases de Dados Factuais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Programas Nacionais de Saúde , Neoplasias/diagnóstico , Neoplasias/mortalidade , Prognóstico , Fatores de Proteção , República da Coreia/epidemiologia , Medição de Risco , Fatores de Risco , Fatores de TempoRESUMO
BACKGROUND AND AIM: Several studies have reported the preventive effect of metformin on cancer development. This study aimed to investigate the relationship between use of metformin and risk of cancer in Koreans. METHODS AND RESULTS: This study was designed retrospectively using the National Health Insurance Service-National Health Screening Cohort conducted between 2002 and 2015. 40 to 69-year-old subjects who received a health screening examination from 2002 to 2003 were enrolled. Hazard ratios (HRs) and 95% confidence intervals (CIs) for cancer were estimated in a multivariate Cox proportional regression analysis. A total of 323,430 subjects was enrolled (301,905 individuals without diabetes [No DM], 8643 diabetic patients with metformin treatment [metformin users], and 12,882 diabetic patients without metformin treatment [metformin non-users]). The median follow-up period was 12.7 years. Cumulative incidence of overall cancer was 7.9% (7.7, 10.3, and 11.1% in No DM, metformin users and non-users, respectively). Compared to metformin non-users, the fully adjusted HRs (95% CIs) of metformin users and No DM for overall cancer incidence were 0.73 (0.66-0.81) and 0.75 (0.64-0.88), respectively, in men and 0.83 (0.78-0.89) and 0.81 (0.72-0.92) in women. CONCLUSIONS: Diabetic patients receiving metformin treatment, and individuals without diabetes were at lower risk for cancer incidence than diabetic patients without metformin treatment.
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Diabetes Mellitus/tratamento farmacológico , Hipoglicemiantes/administração & dosagem , Metformina/administração & dosagem , Neoplasias/prevenção & controle , Adulto , Idoso , Bases de Dados Factuais , Diabetes Mellitus/diagnóstico , Diabetes Mellitus/epidemiologia , Feminino , Humanos , Hipoglicemiantes/efeitos adversos , Incidência , Masculino , Metformina/efeitos adversos , Pessoa de Meia-Idade , Neoplasias/diagnóstico , Neoplasias/epidemiologia , Fatores de Proteção , República da Coreia/epidemiologia , Estudos Retrospectivos , Medição de Risco , Fatores de Risco , Fatores de Tempo , Resultado do TratamentoRESUMO
Foraging animals balance the need to seek food and energy against the accompanying dangers of injury and predation. To do so, they rely on learning systems encoding reward and danger. Whereas much is known about these separate learning systems, little is known about how they interact to shape and guide behavior. Here we show a key role for the rat paraventricular nucleus of the thalamus (PVT), a nucleus of the dorsal midline thalamus, in this interaction. First, we show behavioral competition between reward and danger: the opportunity to seek food reward negatively modulates expression of species-typical defensive behavior. Then, using a chemogenetic approach expressing the inhibitory hM4Di designer receptor exclusively activated by a designer drug in PVT neurons, we show that the PVT is central to this behavioral competition. Chemogenetic PVT silencing biases behavior toward either defense or reward depending on the experimental conditions, but does not consistently favor expression of one over the other. This bias could not be attributed to changes in fear memory retrieval, learned safety, or memory interference. Rather, our results demonstrate that the PVT is essential for balancing conflicting behavioral tendencies toward danger and reward, enabling adaptive responding under this basic selection pressure.SIGNIFICANCE STATEMENT Among the most basic survival problems faced by animals is balancing the need to seek food and energy against the accompanying dangers of injury and predation. Although much is known about the brain mechanisms that underpin learning about reward and danger, little is known about how these interact to solve basic survival problems. Here we show competition between defensive (to avoid predatory detection) and approach (to obtain food) behavior. We show that the paraventricular thalamus, a nucleus of the dorsal midline thalamus, is integral to this behavioral competition. The paraventricular thalamus balances the competing behavioral demands of danger and reward, enabling adaptive responding under this selection pressure.
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Aprendizagem da Esquiva/fisiologia , Tomada de Decisões/fisiologia , Mecanismos de Defesa , Núcleos da Linha Média do Tálamo/fisiologia , Rede Nervosa/fisiologia , Recompensa , Animais , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
mRNA alternative polyadenylation (APA) plays a critical role in post-transcriptional gene control and is highly regulated during development and disease. However, the regulatory mechanisms and functional consequences of APA remain poorly understood. Here, we show that an mRNA 3' processing factor, Fip1, is essential for embryonic stem cell (ESC) self-renewal and somatic cell reprogramming. Fip1 promotes stem cell maintenance, in part, by activating the ESC-specific APA profiles to ensure the optimal expression of a specific set of genes, including critical self-renewal factors. Fip1 expression and the Fip1-dependent APA program change during ESC differentiation and are restored to an ESC-like state during somatic reprogramming. Mechanistically, we provide evidence that the specificity of Fip1-mediated APA regulation depends on multiple factors, including Fip1-RNA interactions and the distance between APA sites. Together, our data highlight the role for post-transcriptional control in stem cell self-renewal, provide mechanistic insight on APA regulation in development, and establish an important function for APA in cell fate specification.
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Regulação da Expressão Gênica no Desenvolvimento , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Processamento Pós-Transcricional do RNA , RNA Mensageiro/metabolismo , Células-Tronco/fisiologia , Animais , Camundongos , Modelos Biológicos , PoliadenilaçãoRESUMO
mRNA 3' processing is dynamically regulated spatially and temporally. However, the underlying mechanisms remain poorly understood. CstF64τ is a paralog of the general mRNA 3' processing factor, CstF64, and has been implicated in mediating testis-specific mRNA alternative polyadenylation (APA). However, the functions of CstF64τ in mRNA 3' processing have not been systematically investigated. We carried out a comprehensive characterization of CstF64τ and compared its properties to those of CstF64. In contrast to previous reports, we found that both CstF64 and CstF64τ are widely expressed in mammalian tissues, and their protein levels display tissue-specific variations. We further demonstrated that CstF64 and CstF64τ have highly similar RNA-binding specificities both in vitro and in vivo. CstF64 and CstF64τ modulate one another's expression and play overlapping as well as distinct roles in regulating global APA profiles. Interestingly, protein interactome analyses revealed key differences between CstF64 and CstF64τ, including their interactions with another mRNA 3' processing factor, symplekin. Together, our study of CstF64 and CstF64τ revealed both functional overlap and specificity of these two important mRNA 3' processing factors and provided new insights into the regulatory mechanisms of mRNA 3' processing.
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Poliadenilação , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/fisiologia , Animais , Fator Estimulador de Clivagem , Sequência Consenso , Expressão Gênica , Perfilação da Expressão Gênica , Células HeLa , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Células NIH 3T3 , Proteínas Nucleares/metabolismo , Especificidade de Órgãos , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Mapeamento de Interação de Proteínas , RNA Mensageiro/genética , Proteínas de Ligação a RNA/químicaRESUMO
BACKGROUND: The hypocholesterolemic effects of lactic acid bacteria and kimchi have been demonstrated previously. However, the kimchi fermentation process still relies on naturally present microorganisms. To obtain functional kimchi with consistent quality, we validated the capacity of Leuconostoc kimchii GJ2 as a starter culture to control kimchi fermentation. Moreover, cholesterol-lowering effects of starter kimchi as a health-promoting product were explored. RESULTS: Bacteriocin production by Lc. kimchii GJ2 was highly enhanced in the presence of 5% Lactobacillus sakei NJ1 cell fractions. When kimchi was fermented with bacteriocin-enhanced Lc. kimchii GJ2, Lc. kimchii GJ2 became overwhelmingly predominant (98.3%) at the end of fermentation and maintained its dominance (up to 82%) for 84 days. Growing as well as dead cells of Lc. kimchii GJ2 showed high cholesterol assimilation (in vitro). Rats were fed a high-fat and high-cholesterol diet supplemented with starter kimchi. The results showed that feeding of starter kimchi significantly reduced serum total cholesterol, triglyceride and low-density lipoprotein cholesterol levels. Additionally, atherogenic index, cardiac risk factor and triglyceride and total cholesterol levels in liver and epididymal adipose tissue decreased significantly in rats fed starter kimchi. CONCLUSION: Kimchi fermented with Lc. kimchii GJ2 as a starter culture has efficient cholesterol-lowering effects.
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Anticolesterolemiantes/uso terapêutico , Brassica , Colesterol/metabolismo , Fermentação , Hipercolesterolemia/dietoterapia , Leuconostoc/metabolismo , Verduras/microbiologia , Tecido Adiposo/metabolismo , Animais , Anticolesterolemiantes/metabolismo , Bacteriocinas/metabolismo , Reatores Biológicos , Dieta Hiperlipídica , Microbiologia de Alimentos , Hipercolesterolemia/etiologia , Hipercolesterolemia/metabolismo , Lactobacillus , Lipoproteínas LDL/sangue , Fígado/metabolismo , Masculino , Ratos Sprague-Dawley , Triglicerídeos/metabolismo , Verduras/metabolismoRESUMO
Neuropeptide Y (NPY) plays a crucial role in controlling energy homeostasis and feeding behaviour. The role of NPY neurons located in the arcuate nucleus of the hypothalamus (Arc) in responding to homeostatic signals has been the focus of much investigation, but most studies have used AgRP promoter-driven models, which do not fully encompass Arc NPY neurons. To directly investigate NPY-expressing versus AgRP-expressing Arc neurons function, we utilised chemogenetic techniques in NPY-Cre and AgRP-Cre animals to activate Arc NPY or AgRP neurons in the presence of food and food-related stimuli. Our findings suggest that chemogenetic activation of the broader population of Arc NPY neurons, including AgRP-positive and AgRP-negative NPY neurons, has equivalent effects on feeding behaviour as activation of Arc AgRP neurons. Our results demonstrate that these Arc NPY neurons respond specifically to caloric signals and do not respond to non-caloric signals, in line with what has been observed in AgRP neurons. Activating Arc NPY neurons significantly increases food consumption and influences macronutrient selection to prefer fat intake.
RESUMO
Alternative polyadenylation (APA) of mRNAs has emerged as an important mechanism for post-transcriptional gene regulation in higher eukaryotes. Although microarrays have recently been used to characterize APA globally, they have a number of serious limitations that prevents comprehensive and highly quantitative analysis. To better characterize APA and its regulation, we have developed a deep sequencing-based method called Poly(A) Site Sequencing (PAS-Seq) for quantitatively profiling RNA polyadenylation at the transcriptome level. PAS-Seq not only accurately and comprehensively identifies poly(A) junctions in mRNAs and noncoding RNAs, but also provides quantitative information on the relative abundance of polyadenylated RNAs. PAS-Seq analyses of human and mouse transcriptomes showed that 40%-50% of all expressed genes produce alternatively polyadenylated mRNAs. Furthermore, our study detected evolutionarily conserved polyadenylation of histone mRNAs and revealed novel features of mitochondrial RNA polyadenylation. Finally, PAS-Seq analyses of mouse embryonic stem (ES) cells, neural stem/progenitor (NSP) cells, and neurons not only identified more poly(A) sites than what was found in the entire mouse EST database, but also detected significant changes in the global APA profile that lead to lengthening of 3' untranslated regions (UTR) in many mRNAs during stem cell differentiation. Together, our PAS-Seq analyses revealed a complex landscape of RNA polyadenylation in mammalian cells and the dynamic regulation of APA during stem cell differentiation.
Assuntos
Sequenciamento de Nucleotídeos em Larga Escala/métodos , Poliadenilação , RNA Mensageiro/química , Análise de Sequência de RNA/métodos , Animais , Células-Tronco Embrionárias/metabolismo , Perfilação da Expressão Gênica , Células HeLa , Histonas/química , Humanos , Camundongos , Células-Tronco Neurais/metabolismo , Neurônios/metabolismo , RNA Mensageiro/genéticaRESUMO
Pre-mRNA splicing is carried out by the spliceosome, which identifies exons and removes intervening introns. In vertebrates, most splice sites are initially recognized by the spliceosome across the exon, because most exons are small and surrounded by large introns. This gene architecture predicts that efficient exon recognition depends largely on the strength of the flanking 3' and 5' splice sites. However, it is unknown if the 3' or the 5' splice site dominates the exon recognition process. Here, we test the 3' and 5' splice site contributions towards efficient exon recognition by systematically replacing the splice sites of an internal exon with sequences of different splice site strengths. We show that the presence of an optimal splice site does not guarantee exon inclusion and that the best predictor for exon recognition is the sum of both splice site scores. Using a genome-wide approach, we demonstrate that the combined 3' and 5' splice site strengths of internal exons provide a much more significant separator between constitutive and alternative exons than either the 3' or the 5' splice site strength alone.
Assuntos
Processamento Alternativo , Éxons , Sítios de Splice de RNA , Células HeLa , HumanosRESUMO
Phenotypic markers that denote different developmental stages of thymocytes are important for understanding T cell development in the thymus. Here, we show that CD1b is a critical discriminator of thymocyte maturation stage in cynomolgus monkeys. CD1b was expressed by immature thymocytes prior to ß-selection, and its expression decreased as cells became fully mature in the thymus. MHC-I expression was lowest at the CD3loCD1b+ immature double-positive (DP) stage, while the ratio of CD1d:MHC-I expression was significantly higher at this stage than at other developmental stages. PLZF was expressed by < 0.2% of thymocytes; most PLZF+ thymocytes were CD3-/loCD1b+ immature DP thymocytes with the potential to produce IL-4. EOMES+ thymocytes, which accounted for > 2% of total thymocytes, were mostly CD3+CD1b- mature thymocytes and predominantly of the CD8 single-positive (SP) lineage. An unconventional CD8+ T cell subset expressing the NKG2AC+CXCR3+ innate-like T cell marker was identified within the EOMES+ CD8 SP lineage; these cells exhibited a memory phenotype. Taken together, these findings show that CD1b is a valuable discriminatory marker of thymocyte development. The data presented herein can be used to characterize the features of PLZF- and EOMES-associated unconventional T cells in the thymus.