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![Model for the activity of the two strands of the processed pre-miRNA molecules (i.e., the guide strand [miRNA] and passenger strand [miRNA*]). Dicer processing of pre-miRNAs typically results in the formation of a guide stand miRNA that binds to the RNA-induced silencing complex (RISC). This guide strand can be derived from either the 3′- (termed −3p) or 5′- (termed −5p) end of the pre-miRNA. The complementary passenger strand typically is degraded. However, under various conditions, including ethanol exposure, miRNA* strands may be retained or otherwise differentially regulated, resulting the emergence of alternate biological end points.
SOURCE: Guo et al. 2012, Acer 2012, Tal et al. 2012.](profile/Rajesh-Miranda/publication/259209756/figure/fig2/AS:202680711553026@1425334255135/Model-for-the-activity-of-the-two-strands-of-the-processed-pre-miRNA-molecules-ie-the.png)
Model for the activity of the two strands of the processed pre-miRNA molecules (i.e., the guide strand [miRNA] and passenger strand [miRNA*]). Dicer processing of pre-miRNAs typically results in the formation of a guide stand miRNA that binds to the RNA-induced silencing complex (RISC). This guide strand can be derived from either the 3′- (termed −3p) or 5′- (termed −5p) end of the pre-miRNA. The complementary passenger strand typically is degraded. However, under various conditions, including ethanol exposure, miRNA* strands may be retained or otherwise differentially regulated, resulting the emergence of alternate biological end points. SOURCE: Guo et al. 2012, Acer 2012, Tal et al. 2012.
Source publication
MicroRNAs (miRNAs) are members of a large class of non-protein-coding RNA (ncRNA) molecules that represent a significant, but until recently unappreciated, layer of cellular regulation. Assessment of the generation and function of miRNAs suggests that these ncRNAs are vulnerable to interference from genetic, epigenetic, and environmental factors. A...
Citations
... Cluster IV T cell genes-2310001H17Rik, Mapk9, Ets1, Ptcd3, Tvp23b, Tcrg-c4 (Fig. 5B)-were associated with GO terms for vas cular development (SI Appendix, Fig. S13D), leukocyte cell adhesion (SI Appendix, Fig. S13D), and microRNA transcription (SI Appendix, Fig. S13D), processes affected in FASD and OMD (48)(49)(50)(51). ...
Severity of neurobehavioral deficits in children born from adverse pregnancies, such as maternal alcohol consumption and diabetes, does not always correlate with the adversity’s duration and intensity. Therefore, biological signatures for accurate prediction of the severity of neurobehavioral deficits, and robust tools for reliable identification of such biomarkers, have an urgent clinical need. Here, we demonstrate that significant changes in the alternative splicing (AS) pattern of offspring lymphocyte RNA can function as accurate peripheral biomarkers for motor learning deficits in mouse models of prenatal alcohol exposure (PAE) and offspring of mother with diabetes (OMD). An aptly trained deep-learning model identified 29 AS events common to PAE and OMD as superior predictors of motor learning deficits than AS events specific to PAE or OMD. Shapley-value analysis, a game-theory algorithm, deciphered the trained deep-learning model’s learnt associations between its input, AS events, and output, motor learning performance. Shapley values of the deep-learning model’s input identified the relative contribution of the 29 common AS events to the motor learning deficit. Gene ontology and predictive structure–function analyses, using Alphafold2 algorithm, supported existing evidence on the critical roles of these molecules in early brain development and function. The direction of most AS events was opposite in PAE and OMD, potentially from differential expression of RNA binding proteins in PAE and OMD. Altogether, this study posits that AS of lymphocyte RNA is a rich resource, and deep-learning is an effective tool, for discovery of peripheral biomarkers of neurobehavioral deficits in children of diverse adverse pregnancies.
... Cluster IV T cell genes-2310001H17Rik, Mapk9, Ets1, Ptcd3, Tvp23b, Tcrg-c4 (Fig. 5B)-were associated with GO terms for vas cular development (SI Appendix, Fig. S13D), leukocyte cell adhesion (SI Appendix, Fig. S13D), and microRNA transcription (SI Appendix, Fig. S13D), processes affected in FASD and OMD (48)(49)(50)(51). ...
Severity of neurobehavioral deficits in children born from adverse pregnancies, such as maternal alcohol consumption and diabetes, does not always correlate with the adversity’s duration and intensity. Therefore, biological signatures for accurate prediction of the severity of neurobehavioral deficits, and robust tools for reliable identification of such biomarkers, have an urgent clinical need. Here, we demonstrate that significant changes in the alternative splicing (AS) pattern of offspring lymphocyte RNA can function as accurate peripheral biomarkers for motor learning deficits in mouse models of prenatal alcohol exposure (PAE) and offspring of mother with diabetes (OMD). An aptly trained deep-learning model identified 29 AS events common to PAE and OMD as superior predictors of motor learning deficits than AS events specific to PAE or OMD. Shapley-value analysis, a game-theory algorithm, deciphered the trained deep-learning model’s learnt associations between its input, AS events, and output, motor learning performance. Shapley values of the deep-learning model’s input identified the relative contribution of the 29 common AS events to the motor learning deficit. Gene ontology and predictive structure–function analyses, using Alphafold2 algorithm, supported existing evidence on the critical roles of these molecules in early brain development and function. The direction of most AS events was opposite in PAE and OMD, potentially from differential expression of RNA binding proteins in PAE and OMD. Altogether, this study posits that AS of lymphocyte RNA is a rich resource, and deep-learning is an effective tool, for discovery of peripheral biomarkers of neurobehavioral deficits in children of diverse adverse pregnancies.
... The mechanism of long-term modulatory effects of PAE on neuroimmune interactions during adulthood remains poorly understood (Sanchez et al., 2019;Noor et al., 2020). Emerging evidence suggests that epigenetic mechanisms involving non-coding RNAs are strong mediators of long-lasting effects of PAE (Savage et al., 2010;Balaraman et al., 2013Balaraman et al., , 2014Balaraman et al., , 2016Tseng et al., 2019;Mahnke et al., 2021). Although several studies have focused on the roles of non-coding RNAs in neuronal function, their roles within the context of PAE-induced neuroinflammatory consequences are not known. ...
Alcohol consumption during pregnancy is associated with Fetal Alcohol Spectrum Disorders (FASD) that results in a continuum of central nervous system (CNS) deficits. Emerging evidence from both preclinical and clinical studies indicate that the biological vulnerability to chronic CNS disease in FASD populations is driven by aberrant neuroimmune actions. Our prior studies suggest that, following minor nerve injury, prenatal alcohol exposure (PAE) is a risk factor for developing adult-onset chronic pathological touch sensitivity or allodynia. Allodynia in PAE rats occurs concurrently with heightened proinflammatory peripheral and spinal glial-immune activation. However, minor nerve-injured control rats remain non-allodynic, and corresponding proinflammatory factors are unaltered. A comprehensive molecular understanding of the mechanism(s) that underlie PAE-induced proinflammatory bias during adulthood remains elusive. Non-coding circular RNAs (circRNAs) are emerging as novel modulators of gene expression. Here, we hypothesized that PAE induces dysregulation of circRNAs that are linked to immune function under basal and nerve-injured conditions during adulthood. Utilizing a microarray platform, we carried out the first systematic profiling of circRNAs in adult PAE rats, prior to and after minor nerve injury. The results demonstrate a unique circRNA profile in adult PAE rats without injury; 18 circRNAs in blood and 32 spinal circRNAs were differentially regulated. Following minor nerve injury, more than 100 differentially regulated spinal circRNAs were observed in allodynic PAE rats. Bioinformatic analysis identified that the parental genes of these circRNAs are linked to the NF-κB complex, a central transcription factor for pain-relevant proinflammatory cytokines. Quantitative real-time PCR was employed to measure levels of selected circRNAs and linear mRNA isoforms. We have validated that circVopp1 was significantly downregulated in blood leukocytes in PAE rats, concurrent with downregulation of Vopp1 mRNA levels. Spinal circVopp1 levels were upregulated in PAE rats, regardless of nerve injury. Additionally, PAE downregulated levels of circItch and circRps6ka3, which are linked to immune regulation. These results demonstrate that PAE exerts long-lasting dysregulation of circRNA expression in blood leukocytes and the spinal cord. Moreover, the spinal circRNA expression profile following peripheral nerve injury is differentially modulated by PAE, potentially contributing to PAE-induced neuroimmune dysregulation.
... Given the role of miRNAs in the regulation of a diverse range of cellular processes such as development and apoptosis [38], neurogenesis and neurodegeneration [39][40][41], as well as their abundance in the CNS [42], they make interesting candidates in the exploration of developmental effects of PAE [43]. There have been multiple investigations of miRNAs in various PAE models, including neural stem cells [44], zebrafish [45] and mice [46,47]. ...
MicroRNAs (miRNAs) are short-length non-protein-coding RNA sequences that post-transcriptionally regulate gene expression in a broad range of cellular processes including neuro- development and have previously been implicated in fetal alcohol spectrum disorders (FASD). In this study, we use our vervet monkey model of FASD to follow up on a prior multivariate (developmental age × ethanol exposure) mRNA analysis (GSE173516) to explore the possibility that the global mRNA downregulation we observed in that study could be related to miRNA expression and function. We report here a predominance of upregulated and differentially expressed miRNAs. Further, the 24 most upregulated miRNAs were significantly correlated with their predicted targets (Target Scan 7.2). We then explored the relationship between these 24 miRNAs and the fold changes observed in their paired mRNA targets using two prediction platforms (Target Scan 7.2 and miRwalk 3.0). Compared to a list of non-differentially expressed miRNAs from our dataset, the 24 upregulated and differentially expressed miRNAs had a greater impact on the fold changes of their corresponding mRNA targets across both platforms. Taken together, this evidence raises the possibility that ethanol-induced upregulation of specific miRNAs might contribute functionally to the general downregulation of mRNAs observed by multiple investigators in response to prenatal alcohol exposure.
... Martinez et al. [56] showed that chronic ethanol exposure over the course of 55 days elevated miR-9-3p in the serum of rats. Balaraman proposed that the ratio between these two mature microRNAs is important in the regulation of neuronal differentiation and in the development of cancer [57]. Both microRNAs impact the differentiation of neural stem cells through the co-regulation of a transcription factor, REST (RE1 silencing transcription factor/neuronrestrictive silencer factor). ...
... Both microRNAs impact the differentiation of neural stem cells through the co-regulation of a transcription factor, REST (RE1 silencing transcription factor/neuronrestrictive silencer factor). miR-9-5p targets REST directly, while miR-9-3p regulates the expression of coREST, a cofactor of REST [42,57]. Therefore, miR-9-5p and miR-9-3p working in tandem can create various combinations of REST:coREST, thus influencing neuronal differentiation [58,59]. ...
microRNA-9 (miR-9) is one of the most abundant microRNAs in the mammalian brain, essential for its development and normal function. In neurons, it regulates the expression of several key molecules, ranging from ion channels to enzymes, to transcription factors broadly affecting the expression of many genes. The neuronal effects of alcohol, one of the most abused drugs in the world, seem to be at least partially dependent on regulating the expression of miR-9. We previously observed that molecular mechanisms of the development of alcohol tolerance are miR-9 dependent. Since a critical feature of alcohol action is temporal exposure to the drug, we decided to better understand the time dependence of alcohol regulation of miR-9 biogenesis and expression. We measured the effect of intoxicating concentration of alcohol (20 mM ethanol) on the expression of all major elements of miR-9 biogenesis: three pri-precursors (pri-mir-9-1, pri-mir-9-2, pri-mir-9-3), three pre-precursors (pre-mir-9-1, pre-mir-9-2, pre-mir-9-3), and two mature microRNAs: miR-9-5p and miR-9-3p, using digital PCR and RT-qPCR, and murine primary medium spiny neurons (MSN) cultures. We subjected the neurons to alcohol based on an exposure/withdrawal matrix of different exposure times (from 15 min to 24 h) followed by different withdrawal times (from 0 h to 24 h). We observed that a short exposure increased mature miR-9-5p expression, which was followed by a gradual decrease and subsequent increase of the expression, returning to pre-exposure levels within 24 h. Temporal changes of miR-9-3p expression were complementing miR-9-5p changes. Interestingly, an extended, continuous presence of the drug caused a similar pattern. These results suggest the presence of the adaptive mechanisms of miR-9 expression in the presence and absence of alcohol. Measurement of miR-9 pre- and pri-precursors showed further that the primary effect of alcohol on miR-9 is through the mir-9-2 precursor pathway with a smaller contribution of mir-9-1 and mir-9-3 precursors. Our results provide new insight into the adaptive mechanisms of neurons to alcohol exposure. It would be of interest to determine next which microRNA-based mechanisms are involved in a transition from the acute, intoxicating effects of alcohol to the chronic, addictive effects of the drug.
... Chronic intermittent ethanol exposure on mouse cerebral cortical neurons during the fetal developmental stage also increased expression of several miRNAs [25]. MicroRNA dysregulation is also associated with disease etiology of FASD [26]. In addition, several studies also reported that alterations of miRNA expression, induced by stress, affect the homeostatic functions of the hypothalamic pituitary adrenal (HPA) axis [27][28][29]. ...
Introduction:
Early life ethanol exposure is known to program hypothalamic proopiomelanocortin (POMC) neurons to express a reduced level of POMC and its control of stress axis functions throughout the life span. In this study we tested whether miRNAs contribute to the ethanol-induced suppression of Pomc gene expression during the developmental period.
Methods:
In in vivo studies, POMC-EGFP male mice were fed with 2.5 g/kg ethanol using milk formula (AF) or pair-fed isocaloric milk formula (PF) or left in the litter (AD) during postnatal day (PND)-2-6. In in vitro studies, mHypoA-POMC/GFP cells were treated with ethanol (50 mM) for a 24 h period. Hypothalamic tissues or cell extracts were used for measurement of miRNAs and POMC mRNA.
Results:
Determination of genome-wide microRNA expression profile identified 40 miRNAs significantly altered in hypothalamic tissues of AF mice. In-silico analysis further identified miRNA-383, 384 and 488 have putative binding sites at the POMC 3'UTR. However, only miR-383 and miR-384 are identified to be responsive to ethanol. Administration of miR-383 or 384 inhibitor oligos suppressed ethanol-stimulated miR-383 or 384 expression and restored Pomc mRNA and protein expression in AF mice. mHypoA-POMC/GFP cells when treated with ethanol showed elevated levels of miR-383 and miR-384 and reduced level of Pomc mRNA. Treatment with miR-383 or 384 mimic oligos reduced the level of Pomc mRNA, while treatment with miR-383 or 384 inhibitor oligos increased the level of Pomc mRNA. Reporter assay further confirms the binding specificity of miR-383 and miR-384 to Pomc 3'UTR.
Conclusion:
These data suggest that miR-383 and miR-384 suppress Pomc gene expression and may contribute to the ethanol-induced alteration of the stress axis functions.
... Cluster IV T cell genes-2310001H17Rik, Mapk9, Ets1, Ptcd3, Tvp23b, Tcrg-c4 (Fig. 5B)-were associated with GO terms for vas cular development (SI Appendix, Fig. S13D), leukocyte cell adhesion (SI Appendix, Fig. S13D), and microRNA transcription (SI Appendix, Fig. S13D), processes affected in FASD and OMD (48)(49)(50)(51). ...
Introduction:
Primary aldosteronism (PA) is a common disease. Especially in unilateral PA (UPA), the risk of cardiovascular disease is high and proper localization is important. Adrenal vein sampling (AVS) is commonly used to localize PA, but its availability is limited. Therefore, it is important to predict the unilateral or bilateral PA and to choose the appropriate cases for AVS or watchful observation.
Aim:
The purpose of this study is to develop a model using machine learning to predict bilateral or unilateral PA to extract cases for AVS or watchful observation.
Methods:
We retrospectively analyzed 154 patients diagnosed with PA and who underwent AVS at our hospital between January 2010 and June 2021. Based on machine learning, we determined predictors of PA subtypes diagnosis from the results of blood and loading tests.
Results:
The accuracy of the machine learning was 88% and the top predictors of the UPA were plasma aldosterone concentration after the saline infusion test, aldosterone to renin ratio after the captopril challenge test, serum potassium and aldosterone-to-renin ratio. By using these factors, the accuracy, sensitivity, specificity and the area under the curve (AUC) were 91%, 70%, 99% and 0.91, respectively. Furthermore, we examined the surgical outcomes of UPA and found that the group diagnosed as unilateral by the predictors showed improvement in clinical findings, while the group diagnosed as bilateral by the predictors showed no improvement.
Conclusion:
Our predictive model based on machine learning can support to choose the performance of adrenal vein sampling or watchful observation.
... Interestingly, early studies into genome wide methylation patterns due to PAE in mice revealed global de-methylation [48]; however, more recently, it has been shown that this pattern is more consistent with demethylation and hyper-methylation depending on the region and the gene in question in both mice [49,50], as well as in humans [51]. With respect to miRNA, there have been several groups that have already examined the effect of alcohol on miRNA expression in various animal models of FASD (as reviewed in [52]). ...
Background
Fetal alcohol spectrum disorders (FASD) are common, yet preventable developmental disorders that stem from prenatal exposure to alcohol. This exposure leads to a wide array of behavioural and physical problems with a complex and poorly defined biological basis.
Molecular investigations to date predominantly use rodent animal models, but because of genetic, developmental and social behavioral similarity, primate models are more relevant. We previously reported reduced cortical and hippocampal neuron levels in an Old World monkey ( Chlorocebus sabaeus ) model with ethanol exposure targeted to the period of rapid synaptogenesis and report here an initial molecular study of this model. The goal of this study was to evaluate mRNA expression of the hippocampus at two different behavioural stages (5 months, 2 years) corresponding to human infancy and early childhood.
Methods
Offspring of alcohol-preferring or control dams drank a maximum of 3.5 g ethanol per kg body weight or calorically matched sucrose solution 4 days per week during the last 2 months of gestation. Total mRNA expression was measured with the Affymetrix GeneChip Rhesus Macaque Genome Array in a 2 × 2 study design that interrogated two independent variables, age at sacrifice, and alcohol consumption during gestation.
Results and discussion
Statistical analysis identified a preferential downregulation of expression when interrogating the factor ‘alcohol’ with a balanced effect of upregulation vs. downregulation for the independent variable ‘age’. Functional exploration of both independent variables shows that the alcohol consumption factor generates broad functional annotation clusters that likely implicate a role for epigenetics in the observed differential expression, while the variable age reliably produced functional annotation clusters predominantly related to development. Furthermore, our data reveals a novel connection between EFNB1 and the FASDs; this is highly plausible both due to the role of EFNB1 in neuronal development as well as its central role in craniofrontal nasal syndrome (CFNS). Fold changes for key genes were subsequently confirmed via q RT-PCR.
Conclusion
Prenatal alcohol exposure leads to global downregulation in mRNA expression. The cellular interference model of EFNB1 provides a potential clue regarding how genetically susceptible individuals may develop the phenotypic triad generally associated with classic fetal alcohol syndrome.
... MicroRNAs (miRNAs) are a class of small non-protein-coding RNAs, known to repress protein translation, that are alcohol-sensitive and can regulate the effects of alcohol on fetal brain development in animal models [53][54][55][56]. Because miRNAs are key players in stem cell self-renewal, cell cycle regulation, apoptosis, and development, dysregulation of miRNAs by PAE can have significant toxic and teratogenic effects on a developing fetus [57][58][59][60]. Understanding the interaction between PAE and miRNAs, and the regulatory role ethanolsensitive miRNAs can have on an individual, may help us to better understand the ethanolassociated toxicology and teratology. ...
Prenatal alcohol exposure (PAE) can have immediate and long-lasting toxic and teratogenic effects on an individual’s development and health. As a toxicant, alcohol can lead to a variety of physical and neurological anomalies in the fetus that can lead to behavioral and other impairments which may last a lifetime. Recent studies have focused on identifying mechanisms that mediate the immediate teratogenic effects of alcohol on fetal development and mechanisms that facilitate the persistent toxic effects of alcohol on health and predisposition to disease later in life. This review focuses on the contribution of epigenetic modifications and intercellular transporters like extracellular vesicles to the toxicity of PAE and to immediate and long-term consequences on an individual’s health and risk of disease.
... Furthermore, alterations of miRNA expression patterns in response to developmental alcohol exposure have been reported. It has been suggested that miRNAs are vulnerable to alcohol exposure and may contribute to the detrimental effects on fetal development [60][61][62]. Potential fetal tissue origin of these AF miRNAs altered by FAE was demonstrated by the coherent changes in miRNA level in total fetal embryo tissue (Fig 4A). ...
Alcohol (ethanol, EtOH) consumption during pregnancy can result in fetal alcohol spectrum disorders (FASDs), which are characterized by prenatal and postnatal growth restriction and craniofacial dysmorphology. Recently, cell-derived extracellular vesicles, including exosomes and microvesicles containing several species of RNAs (exRNAs), have emerged as a mechanism of cell-to-cell communication. However, EtOH’s effects on the biogenesis and function of non-coding exRNAs during fetal development have not been explored. Therefore, we studied the effects of maternal EtOH exposure on the composition of exosomal RNAs in the amniotic fluid (AF) using rat fetal alcohol exposure (FAE) model. Through RNA-Seq analysis we identified and verified AF exosomal miRNAs with differential expression levels specifically associated with maternal EtOH exposure. Uptake of purified FAE AF exosomes by rBMSCs resulted in significant alteration of molecular markers associated with osteogenic differentiation of rBMSCs. We also determined putative functional roles for AF exosomal miRNAs (miR-199a-3p, miR-214-3p and let-7g) that are dysregulated by FAE in osteogenic differentiation of rBMSCs. Our results demonstrate that FAE alters AF exosomal miRNAs and that exosomal transfer of dysregulated miRNAs has significant molecular effects on stem cell regulation and differentiation. Our results further suggest the usefulness of assessing molecular alterations in AF exRNAs to study the mechanisms of FAE teratogenesis that should be further investigated by using an in vivo model.
