RESUMO
Heart valve disease affects up to 30% of the population and has been shown to have origins during embryonic development. Valvulogenesis begins with formation of endocardial cushions in the atrioventricular canal and outflow tract regions. Subsequently, endocardial cushions remodel, elongate and progressively form mature valve structures composed of a highly organized connective tissue that provides the necessary biomechanical function throughout life. While endocardial cushion formation has been well studied, the processes required for valve remodeling are less well understood. The transcription factor Scleraxis (Scx) is detected in mouse valves from E15.5 during initial stages of remodeling, and expression remains high until birth when formation of the highly organized mature structure is complete. Heart valves from Scx-/- mice are abnormally thick and develop fibrotic phenotypes similar to human disease by juvenile stages. These phenotypes begin around E15.5 and are associated with defects in connective tissue organization and valve interstitial cell differentiation. In order to understand the etiology of this phenotype, we analyzed the transcriptome of remodeling valves isolated from E15.5 Scx-/- embryos using RNA-seq. From this, we have identified a profile of protein and non-protein mRNAs that are dependent on Scx function and using bioinformatics we can predict the molecular functions and biological processes affected by these genes. These include processes and functions associated with gene regulation (methyltransferase activity, DNA binding, Notch signaling), vitamin A metabolism (retinoic acid biosynthesis) and cellular development (cell morphology, cell assembly and organization). In addition, several mRNAs are affected by alternative splicing events in the absence of Scx, suggesting additional roles in post-transcriptional modification. In summary, our findings have identified transcriptome profiles from abnormal heart valves isolated from E15.5 Scx-/- embryos that could be used in the future to understand mechanisms of heart valve disease in the human population.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Doenças das Valvas Cardíacas/embriologia , Valvas Cardíacas/embriologia , RNA Mensageiro/biossíntese , Análise de Sequência de RNA , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Doenças das Valvas Cardíacas/genética , Doenças das Valvas Cardíacas/patologia , Valvas Cardíacas/patologia , Humanos , Camundongos , Camundongos Knockout , RNA Mensageiro/genéticaRESUMO
Hypoplastic Left Heart Syndrome (HLHS) is a congenital defect characterized by underdevelopment of the left ventricle and pathological compensation of the right ventricle. If untreated, HLHS is invariably lethal due to the extensive increase in right ventricular workload and eventual failure. Despite the clinical significance, little is known about the molecular pathobiological state of HLHS. Splicing of mRNA transcripts is an important regulatory mechanism of gene expression. Tissue specific alterations of this process have been associated with several cardiac diseases, however, transcriptional signature profiles related to HLHS are unknown. In this study, we performed genome-wide exon array analysis to determine differentially expressed genes and alternatively spliced transcripts in the right ventricle (RV) of six neonates with HLHS, compared to the RV and left ventricle (LV) from non-diseased control subjects. In HLHS, over 180 genes were differentially expressed and 1800 were differentially spliced, leading to changes in a variety of biological processes involving cell metabolism, cytoskeleton, and cell adherence. Additional hierarchical clustering analysis revealed that differential gene expression and mRNA splicing patterns identified in HLHS are unique compared to non-diseased tissue. Our findings suggest that gene expression and mRNA splicing are broadly dysregulated in the RV myocardium of HLHS neonates. In addition, our analysis identified transcriptome profiles representative of molecular biomarkers of HLHS that could be used in the future for diagnostic and prognostic stratification to improve patient outcome.
Assuntos
Processamento Alternativo/genética , Perfilação da Expressão Gênica , Síndrome do Coração Esquerdo Hipoplásico/genética , Síndrome do Coração Esquerdo Hipoplásico/patologia , Miocárdio/metabolismo , Estudos de Casos e Controles , Análise por Conglomerados , Feminino , Idade Gestacional , Humanos , Síndrome do Coração Esquerdo Hipoplásico/metabolismo , Recém-Nascido , Masculino , Análise em Microsséries , Miocárdio/patologia , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estudos de Validação como AssuntoRESUMO
Retinal ganglion cells apoptosis is linked to matrix metalloproteinase 9 (MMP-9) controlled changes of extracellular matrix. Abnormal expression of MMP-9 is associated with glaucomatous alterations. Thus, the knowledge of MMP-9 regulation is important for the understanding the pathogenesis of glaucoma. Here, we investigated the role of 3'-untranslated regions (3'-UTR) and microRNAs in MMP-9 regulation. We used in vitro mutagenesis and Luc reporter system to identify regulatory elements in the 3'-UTR of MMP-9. microRNAs were analyzed by qRT-PCR, and their role was investigated with inhibitors and mimics. We identified targets for miRNAs in 3'-UTR of MMP-9 involved in the regulation of MMP-9 expression. We then isolated miRNAs from the optic nerve A7 astrocytes and 293 T cells and confirmed the role of mi340 in the regulation using specific inhibitors and mimics. The results obtained show a new miRNA-mediated mechanism of MMP-9 expression regulation.
RESUMO
The presence of workers that forgo reproduction and care for their siblings is a defining feature of eusociality and a major challenge for evolutionary theory. It has been proposed that worker behavior evolved from maternal care behavior. We explored this idea by studying gene expression in the primitively eusocial wasp Polistes metricus. Because little genomic information existed for this species, we used 454 sequencing to generate 391,157 brain complementary DNA reads, resulting in robust hits to 3017 genes from the honey bee genome, from which we identified and assayed orthologs of 32 honey bee behaviorally related genes. Wasp brain gene expression in workers was more similar to that in foundresses, which show maternal care, than to that in queens and gynes, which do not. Insulin-related genes were among the differentially regulated genes, suggesting that the evolution of eusociality involved major nutritional and reproductive pathways.