RNA Sequencing Reveals the Alteration of the Expression of Novel Genes in Ethanol-Treated Embryoid Bodies

Abstract

Fetal alcohol spectrum disorder is a collective term representing fetal abnormalities associated with maternal alcohol consumption. Prenatal alcohol exposure and related anomalies are well characterized, but the molecular mechanism behind this phenomenon is not well characterized. In this present study, our aim is to profile important genes that regulate cellular development during fetal development. Human embryonic carcinoma cells (NCCIT) are cultured to form embryoid bodies and then treated in the presence and absence of ethanol (50 mM). We employed RNA sequencing to profile differentially expressed genes in the ethanol-treated embryoid bodies from NCCIT vs. EB, NCCIT vs. EB+EtOH and EB vs. EB+EtOH data sets. A total of 632, 205 and 517 differentially expressed genes were identified from NCCIT vs. EB, NCCIT vs. EB+EtOH and EB vs. EB+EtOH, respectively. Functional annotation using bioinformatics tools reveal significant enrichment of differential cellular development and developmental disorders. Furthermore, a group of 42, 15 and 35 transcription factor-encoding genes are screened from all of the differentially expressed genes obtained from NCCIT vs. EB, NCCIT vs. EB+EtOH and EB vs. EB+EtOH, respectively. We validated relative gene expression levels of several transcription factors from these lists by quantitative real-time PCR. We hope that our study substantially contributes to the understanding of the molecular mechanism underlying the pathology of alcohol-mediated anomalies and ease further research.

Maternal alcohol consumption and altered miRNAs in the developing fetus: Context and future perspectives

“…A similar result was previously reported in miR‐9‐2/3 knockout mice, which exhibit fetal growth retardation, microencephaly, and other features that are also common to fetal alcohol spectrum disorder (FASD) (Shibata, Nakao, Kiyonari, Abe, & Aizawa, ). Mandal and his colleagues cultured embryoid bodies exposed to ethanol for 3 and 7 days to profile transcriptomic expressions during early development (Mandal, Halder, et al, ; Mandal, Kim, et al, ). Transcriptomic analysis showed that ethanol dysregulated the expression of the MIR663 and MIRLET7DHG genes after 3 days of ethanol exposure (Mandal, Kim, et al, ).…”

“…Mandal and his colleagues cultured embryoid bodies exposed to ethanol for 3 and 7 days to profile transcriptomic expressions during early development (Mandal, Halder, et al, 2016;Mandal, Kim, et al, 2016). Transcriptomic analysis showed that ethanol dysregulated the expression of the MIR663 and MIRLET7DHG genes after 3 days of ethanol exposure (Mandal, Kim, et al, 2016). Microarray analysis of differentially expressed genes after 7 days of ethanol exposure showed altered expression levels of the MIR9-3, MIR15B, MIR186, MIR19B2, MIR301A, MIR302A, MIR302B and MIRLET7F1 genes (Mandal, Halder, et al, 2016).…”

“…(Mandal, Halder, et al, 2016) Human 50 mM in culture medium Suspension culture (72 hrs) Up-regulation of MIR-663 and MIRLET7DHG mRNAs. (Mandal, Kim, et al, 2016) Human ≥0.21 ounces of absolute alcohol per day (equivalent to ≥3 drinks/wk) ≥2.0 ounces of absolute alcohol per drinking day (equivalent to one bingedrinking)…”

Gestational Alcohol Exposure Altered DNA Methylation Status in the Developing Fetus

“…Alcohol is well known as a teratogenic agent that affects the normal development of a developing embryo. Fetal alcohol spectrum disorder (FASD) is the term commonly used to denote alcohol-related neonatal abnormalities [ 1 , 2 ]. The most severe form of FASD is known as fetal alcohol syndrome (FAS), and manifests as growth retardation, facial abnormalities, and central nervous system (CNS) deficiencies [ 3 , 4 ].…”

“…Many previous studies have described genetic susceptibilities to alcohol-mediated defects. Microarray analysis and next-generation sequencing techniques have found that many developmental genes are altered by in utero alcohol exposure [ 1 , 2 , 3 , 4 , 7 , 8 , 9 , 10 ]. However, the underlying causes of these alcohol-mediated disorders remain unclear.…”