RESUMO
BACKGROUND: Recent studies have shown that prenatal BPA exposure altered the transcriptome profiles of autism-related genes in the offspring's hippocampus, disrupting hippocampal neuritogenesis and causing male-specific deficits in learning. However, the sex differences in the effects of prenatal BPA exposure on the developing prefrontal cortex, which is another brain region highly implicated in autism spectrum disorder (ASD), have not been investigated. METHODS: We obtained transcriptome data from RNA sequencing analysis of the prefrontal cortex of male and female rat pups prenatally exposed to BPA or control and reanalyzed. BPA-responsive genes associated with cortical development and social behaviors were selected for confirmation by qRT-PCR analysis. Neuritogenesis of primary cells from the prefrontal cortex of pups prenatally exposed to BPA or control was examined. The social behaviors of the pups were assessed using the two-trial and three-chamber tests. The male-specific impact of the downregulation of a selected BPA-responsive gene (i.e., Sema5a) on cortical development in vivo was interrogated using siRNA-mediated knockdown by an in utero electroporation technique. RESULTS: Genes disrupted by prenatal BPA exposure were associated with ASD and showed sex-specific dysregulation. Sema5a and Slc9a9, which were involved in neuritogenesis and social behaviors, were downregulated only in males, while Anxa2 and Junb, which were also linked to neuritogenesis and social behaviors, were suppressed only in females. Neuritogenesis was increased in males and showed a strong inverse correlation with Sema5a and Slc9a9 expression levels, whereas, in the females, neuritogenesis was decreased and correlated with Anxa2 and Junb levels. The siRNA-mediated knockdown of Sema5a in males also impaired cortical development in utero. Consistent with Anxa2 and Junb downregulations, deficits in social novelty were observed only in female offspring but not in males. CONCLUSION: This is the first study to show that prenatal BPA exposure dysregulated the expression of ASD-related genes and functions, including cortical neuritogenesis and development and social behaviors, in a sex-dependent manner. Our findings suggest that, besides the hippocampus, BPA could also exert its adverse effects through sex-specific molecular mechanisms in the offspring's prefrontal cortex, which in turn would lead to sex differences in ASD-related neuropathology and clinical manifestations, which deserves further investigation.
Assuntos
Compostos Benzidrílicos , Fenóis , Córtex Pré-Frontal , Efeitos Tardios da Exposição Pré-Natal , Caracteres Sexuais , Comportamento Social , Animais , Feminino , Córtex Pré-Frontal/efeitos dos fármacos , Córtex Pré-Frontal/metabolismo , Fenóis/toxicidade , Fenóis/efeitos adversos , Masculino , Compostos Benzidrílicos/toxicidade , Gravidez , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Transtorno Autístico/genética , Transtorno Autístico/induzido quimicamente , Ratos Sprague-Dawley , Ratos , Transtorno do Espectro Autista/induzido quimicamente , Transtorno do Espectro Autista/genéticaRESUMO
Alu elements are transposable elements that can influence gene regulation through several mechanisms; nevertheless, it remains unclear whether dysregulation of Alu elements contributes to the neuropathology of autism spectrum disorder (ASD). In this study, we characterized transposable element expression profiles and their sequence characteristics in the prefrontal cortex tissues of ASD and unaffected individuals using RNA-sequencing data. Our results showed that most of the differentially expressed transposable elements belong to the Alu family, with 659 loci of Alu elements corresponding to 456 differentially expressed genes in the prefrontal cortex of ASD individuals. We predicted cis- and trans-regulation of Alu elements to host/distant genes by conducting correlation analyses. The expression level of Alu elements correlated significantly with 133 host genes (cis-regulation, adjusted p < 0.05) associated with ASD as well as the cell survival and cell death of neuronal cells. Transcription factor binding sites in the promoter regions of differentially expressed Alu elements are conserved and associated with autism candidate genes, including RORA. COBRA analyses of postmortem brain tissues showed significant hypomethylation in global methylation analyses of Alu elements in ASD subphenotypes as well as DNA methylation of Alu elements located near the RNF-135 gene (p < 0.05). In addition, we found that neuronal cell density, which was significantly increased (p = 0.042), correlated with the expression of genes associated with Alu elements in the prefrontal cortex of ASD. Finally, we determined a relationship between these findings and the ASD severity (i.e., ADI-R scores) of individuals with ASD. Our findings provide a better understanding of the impact of Alu elements on gene regulation and molecular neuropathology in the brain tissues of ASD individuals, which deserves further investigation.
Assuntos
Transtorno do Espectro Autista , Transtorno Autístico , Humanos , Transtorno Autístico/genética , Transtorno do Espectro Autista/genética , Transtorno do Espectro Autista/metabolismo , Elementos Alu/genética , Elementos de DNA Transponíveis , Metilação de DNA , Epigênese Genética , Córtex Pré-Frontal/metabolismoRESUMO
Long interspersed nucleotide element-1 (LINE-1) and Alu elements are retrotransposons whose abilities cause abnormal gene expression and genomic instability. Several studies have focused on DNA methylation profiling of gene regions, but the locus-specific methylation of LINE-1 and Alu elements has not been identified in autism spectrum disorder (ASD). Here we interrogated locus- and family-specific methylation profiles of LINE-1 and Alu elements in ASD whole blood using publicly-available Illumina Infinium 450 K methylation datasets from heterogeneous ASD and ASD variants (Chromodomain Helicase DNA-binding 8 (CHD8) and 16p11.2del). Total DNA methylation of repetitive elements were notably hypomethylated exclusively in ASD with CHD8 variants. Methylation alteration in a family-specific manner including L1P, L1H, HAL, AluJ, and AluS families were observed in the heterogeneous ASD and ASD with CHD8 variants. Moreover, LINE-1 and Alu methylation within target genes is inversely related to the expression level in each ASD variant. The DNA methylation signatures of the LINE-1 and Alu elements in ASD whole blood, as well as their associations with the expression of ASD-related genes, have been identified. If confirmed in future larger studies, these findings may contribute to the identification of epigenomic biomarkers of ASD.
Assuntos
Transtorno do Espectro Autista , Transtorno Autístico , Elementos Alu/genética , Transtorno do Espectro Autista/genética , Transtorno Autístico/genética , Metilação de DNA , Humanos , Elementos Nucleotídeos Longos e Dispersos/genéticaRESUMO
Our recent study has shown that prenatal exposure to bisphenol A (BPA) altered the expression of genes associated with autism spectrum disorder (ASD). In this study, we further investigated the effects of prenatal BPA exposure on ASD-related genes known to regulate neuronal viability, neuritogenesis, and learning/memory, and assessed these functions in the offspring of exposed pregnant rats. We found that prenatal BPA exposure increased neurite length, the number of primary neurites, and the number of neurite branches, but reduced the size of the hippocampal cell body in both sexes of the offspring. However, in utero exposure to BPA decreased the neuronal viability and the neuronal density in the hippocampus and impaired learning/memory only in the male offspring while the females were not affected. Interestingly, the expression of several ASD-related genes (e.g. Mief2, Eif3h, Cux1, and Atp8a1) in the hippocampus were dysregulated and showed a sex-specific correlation with neuronal viability, neuritogenesis, and/or learning/memory. The findings from this study suggest that prenatal BPA exposure disrupts ASD-related genes involved in neuronal viability, neuritogenesis, and learning/memory in a sex-dependent manner, and these genes may play an important role in the risk and the higher prevalence of ASD in males subjected to prenatal BPA exposure.
Assuntos
Transtorno do Espectro Autista/metabolismo , Compostos Benzidrílicos/toxicidade , Regulação da Expressão Gênica/efeitos dos fármacos , Hipocampo/metabolismo , Fenóis/toxicidade , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Caracteres Sexuais , Animais , Transtorno do Espectro Autista/genética , Transtorno do Espectro Autista/patologia , Feminino , Hipocampo/patologia , Masculino , Gravidez , Efeitos Tardios da Exposição Pré-Natal/genética , Efeitos Tardios da Exposição Pré-Natal/patologia , Ratos , Ratos WistarRESUMO
Our recent study revealed that prenatal exposure to bisphenol A (BPA) disrupted the transcriptome profiles of genes in the offspring hippocampus. In addition to genes linked to autism, several genes associated with Alzheimer's disease (AD) were found to be differentially expressed, although the association between BPA-responsive genes and AD-related genes has not been thoroughly investigated. Here, we demonstrated that in utero BPA exposure also disrupted the transcriptome profiles of genes associated with neuroinflammation and AD in the hippocampus. The level of NF-κB protein and its AD-related target gene Bace1 were significantly increased in the offspring hippocampus in a sex-dependent manner. Quantitative RT-PCR analysis also showed an increase in the expression of Tnf gene. Moreover, the reanalysis of transcriptome profiling data from several previously published BPA studies consistently showed that BPA-responsive genes were significantly associated with top AD candidate genes. The findings from this study suggest that maternal BPA exposure may increase AD risk in offspring by dysregulating genes associated with AD neuropathology and inflammation and reveal a possible relationship between AD and autism, which are linked to the same environmental factor. Sex-specific effects of prenatal BPA exposure on the susceptibility of AD deserve further investigation.
Assuntos
Doença de Alzheimer/induzido quimicamente , Doença de Alzheimer/genética , Compostos Benzidrílicos/agonistas , Fenóis/agonistas , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Transcriptoma/efeitos dos fármacos , Animais , Transtorno Autístico/induzido quimicamente , Transtorno Autístico/genética , Feminino , Perfilação da Expressão Gênica/métodos , Hipocampo/efeitos dos fármacos , Inflamação/induzido quimicamente , Inflamação/genética , Masculino , Exposição Materna , NF-kappa B/genética , Gravidez , Efeitos Tardios da Exposição Pré-Natal/genética , Ratos , Ratos WistarRESUMO
BACKGROUND: The exact cause and mechanisms underlying the pathobiology of autism spectrum disorder (ASD) remain unclear. Dysregulation of long interspersed element-1 (LINE-1) has been reported in the brains of ASD-like mutant mice and ASD brain tissues. However, the role and methylation of LINE-1 in individuals with ASD remain unclear. In this study, we aimed to investigate whether LINE-1 insertion is associated with differentially expressed genes (DEGs) and to assess LINE-1 methylation in ASD. METHODS: To identify DEGs associated with LINE-1 in ASD, we reanalyzed previously published transcriptome profiles and overlapped them with the list of LINE-1-containing genes from the TranspoGene database. An Ingenuity Pathway Analysis (IPA) of DEGs associated with LINE-1 insertion was conducted. DNA methylation of LINE-1 was assessed via combined bisulfite restriction analysis (COBRA) of lymphoblastoid cell lines from ASD individuals and unaffected individuals, and the methylation levels were correlated with the expression levels of LINE-1 and two LINE-1-inserted DEGs, C1orf27 and ARMC8. RESULTS: We found that LINE-1 insertion was significantly associated with DEGs in ASD. The IPA showed that LINE-1-inserted DEGs were associated with ASD-related mechanisms, including sex hormone receptor signaling and axon guidance signaling. Moreover, we observed that the LINE-1 methylation level was significantly reduced in lymphoblastoid cell lines from ASD individuals with severe language impairment and was inversely correlated with the transcript level. The methylation level of LINE-1 was also correlated with the expression of the LINE-1-inserted DEG C1orf27 but not ARMC8. CONCLUSIONS: In ASD individuals with severe language impairment, LINE-1 methylation was reduced and correlated with the expression levels of LINE-1 and the LINE-1-inserted DEG C1orf27. Our findings highlight the association of LINE-1 with DEGs in ASD blood samples and warrant further investigation. The molecular mechanisms of LINE-1 and the effects of its methylation in ASD pathobiology deserve further study.
Assuntos
Transtorno do Espectro Autista/metabolismo , Metilação de DNA , Elementos Nucleotídeos Longos e Dispersos , Transcriptoma , Proteínas do Domínio Armadillo/metabolismo , Transtorno do Espectro Autista/genética , Linhagem Celular , Epigênese Genética , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , HumanosRESUMO
Background: Alu elements are a group of repetitive elements that can influence gene expression through CpG residues and transcription factor binding. Altered gene expression and methylation profiles have been reported in various tissues and cell lines from individuals with autism spectrum disorder (ASD). However, the role of Alu elements in ASD remains unclear. We thus investigated whether Alu elements are associated with altered gene expression profiles in ASD. Methods: We obtained five blood-based gene expression profiles from the Gene Expression Omnibus database and human Alu-inserted gene lists from the TranspoGene database. Differentially expressed genes (DEGs) in ASD were identified from each study and overlapped with the human Alu-inserted genes. The biological functions and networks of Alu-inserted DEGs were then predicted by Ingenuity Pathway Analysis (IPA). A combined bisulfite restriction analysis of lymphoblastoid cell lines (LCLs) derived from 36 ASD and 20 sex- and age-matched unaffected individuals was performed to assess the global DNA methylation levels within Alu elements, and the Alu expression levels were determined by quantitative RT-PCR. Results: In ASD blood or blood-derived cells, 320 Alu-inserted genes were reproducibly differentially expressed. Biological function and pathway analysis showed that these genes were significantly associated with neurodevelopmental disorders and neurological functions involved in ASD etiology. Interestingly, estrogen receptor and androgen signaling pathways implicated in the sex bias of ASD, as well as IL-6 signaling and neuroinflammation signaling pathways, were also highlighted. Alu methylation was not significantly different between the ASD and sex- and age-matched control groups. However, significantly altered Alu methylation patterns were observed in ASD cases sub-grouped based on Autism Diagnostic Interview-Revised scores compared with matched controls. Quantitative RT-PCR analysis of Alu expression also showed significant differences between ASD subgroups. Interestingly, Alu expression was correlated with methylation status in one phenotypic ASD subgroup. Conclusion: Alu methylation and expression were altered in LCLs from ASD subgroups. Our findings highlight the association of Alu elements with gene dysregulation in ASD blood samples and warrant further investigation. Moreover, the classification of ASD individuals into subgroups based on phenotypes may be beneficial and could provide insights into the still unknown etiology and the underlying mechanisms of ASD.