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. 2020 Feb 1;161(2):bqz044.
doi: 10.1210/endocr/bqz044.

Dysregulation of Hypothalamic Gene Expression and the Oxytocinergic System by Soybean Oil Diets in Male Mice

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Free PMC article

Dysregulation of Hypothalamic Gene Expression and the Oxytocinergic System by Soybean Oil Diets in Male Mice

Poonamjot Deol et al. Endocrinology. .
Free PMC article

Abstract

Soybean oil consumption has increased greatly in the past half-century and is linked to obesity and diabetes. To test the hypothesis that soybean oil diet alters hypothalamic gene expression in conjunction with metabolic phenotype, we performed RNA sequencing analysis using male mice fed isocaloric, high-fat diets based on conventional soybean oil (high in linoleic acid, LA), a genetically modified, low-LA soybean oil (Plenish), and coconut oil (high in saturated fat, containing no LA). The 2 soybean oil diets had similar but nonidentical effects on the hypothalamic transcriptome, whereas the coconut oil diet had a negligible effect compared to a low-fat control diet. Dysregulated genes were associated with inflammation, neuroendocrine, neurochemical, and insulin signaling. Oxt was the only gene with metabolic, inflammation, and neurological relevance upregulated by both soybean oil diets compared to both control diets. Oxytocin immunoreactivity in the supraoptic and paraventricular nuclei of the hypothalamus was reduced, whereas plasma oxytocin and hypothalamic Oxt were increased. These central and peripheral effects of soybean oil diets were correlated with glucose intolerance but not body weight. Alterations in hypothalamic Oxt and plasma oxytocin were not observed in the coconut oil diet enriched in stigmasterol, a phytosterol found in soybean oil. We postulate that neither stigmasterol nor LA is responsible for effects of soybean oil diets on oxytocin and that Oxt messenger RNA levels could be associated with the diabetic state. Given the ubiquitous presence of soybean oil in the American diet, its observed effects on hypothalamic gene expression could have important public health ramifications.

Keywords: Plenish; coconut oil; diabetes; high-fat diet; linoleic acid; oxytocin; stigmasterol.

Figures

Figure 1.
Figure 1.
Study design and 3-dimensional principal components analysis (PCA) showing differential effects of dietary fat on hypothalamic gene expression. A, Workflow showing the 2 cohorts of mice used, different diets, time on diets, and various analyses performed. B, PCA of RNA-sequencing data from hypothalami of male mice fed Viv chow, CO, SO + CO or PL + CO for 24 weeks showing 3 biological replicates for each diet. As indicated, the 2 soybean oil diets, SO + CO and PL + CO, can be grouped together as well as Viv chow and CO. CO, coconut oil diet; EIA, enzyme immunoassay; GTT, glucose tolerance test; IHC, immunohistochemistry; RT-qPCR, reverse transcriptase-quantitative polymerase chain reaction; Viv chow, vivarium chow (low-fat control). CO diets enriched in conventional soybean oil (SO + CO), genetically modified Plenish (PL + CO), and stigmasterol (ST + CO).
Figure 2.
Figure 2.
Soybean oil affects the hypothalamic transcriptome. A, Heat map of differential hypothalamic gene expression (RNA sequencing [RNA-seq]) between mice fed vivarium (Viv) chow, CO, SO + CO, and PL + CO diets for 24 weeks. Color bar indicates level of gene expression relative to the whole data set. Top, genes upregulated in Viv chow vs the 3 high-fat diets. Bottom, genes downregulated in Viv chow. Arrows, genes mentioned in text. Asterisk, Kcng1, the only gene significantly different between SO + CO and PL + CO. B, Gene ontology showing categories of molecular function affected by diet. C, Absolute expression levels from RNA-seq of indicated diets. DEG, differentially expressed genes; RPKM, reads per kilobase per million. Statistically different from #CO, *Viv chow, @PL + CO. CO diets enriched in conventional soybean oil (SO + CO), genetically modified Plenish (PL + CO), and stigmasterol (ST + CO).
Figure 3.
Figure 3.
Comparison of differentially expressed hypothalamic genes in different diet groups. A, Venn diagram of number of genes dysregulated in left, SO + CO, and right, PL + CO vs CO and Viv chow diets (≥1.5-fold, P and Padj ≤.05). B, Absolute expression levels from RNA sequencing (RNA-seq) of genes that are most dysregulated between SO + CO, PL + CO, or both soybean oil diets vs CO. Statistically different from #CO, *Viv chow. CO diets enriched in conventional soybean oil (SO + CO), genetically modified Plenish (PL + CO), and stigmasterol (ST + CO).
Figure 4.
Figure 4.
Biological pathways associated with hypothalamic genes dysregulated by soybean oil (SO)-rich high-fat diets. A, Venn diagram showing number of genes dysregulated (≥ 1.5-fold, P and Padj ≤ .05) either exclusively or commonly by SO + CO and PL + CO vs CO and gene ontology for biological pathways altered in the comparisons shown. B, Absolute expression levels from RNA sequencing of dysregulated genes (≥ 1.5 fold, P and Padj ≤ .05) in the most prominent pathways. Significantly different from #CO, *Viv chow. CO diets enriched in conventional soybean oil (SO + CO), genetically modified Plenish (PL + CO), and stigmasterol (ST + CO).
Figure 5.
Figure 5.
Oxt is the only gene dysregulated in both soybean oil diets (vs CO and Viv chow) that is associated with neurological, metabolic, and inflammation disease categories. A, Venn diagram showing overlap in hypothalamic genes from RNA sequencing (RNA-seq) related to neurological diseases vs metabolic diseases and inflammation dysregulated in SO + CO and PL + CO vs CO (Tables 5 and 6). B, Absolute expression levels from RNA-seq data of 6 of the 9 common genes. Significantly different from #CO, *Viv chow. C, Venn diagram showing overlap in genes related to metabolic diseases (obesity, diabetes, and lipid metabolism), inflammation, and neurological diseases that are dysregulated in the hypothalamus of both soybean oil diets vs CO diet. Oxytocin (Oxt, red font) is the only gene common to all 3 disease categories and is significantly different both from CO and Viv chow. CO diets enriched in conventional soybean oil (SO + CO), genetically modified Plenish (PL + CO), and stigmasterol (ST + CO).
Figure 6.
Figure 6.
Obesogenic and diabetogenic effects of soybean oil (SO) diets with high or low linoleic acid (LA) is not mimicked by stigmasterol. A, Average weekly body weights of male C57BL/6N mice started on the indicated diets at weaning (N = 6-12/diet) as in Fig. 1A. All diets were isocaloric with 40 kcal% total fat except Viv chow, which had 13.4 kcal% fat. B, Glycemia (mg/dL) measured during a glucose tolerance test (GTT) depicted as area under the curve after 16 weeks on diet (N = 6–12/diet). C, Average weekly food consumption of mice on various diets measured on a per-cage basis and normalized to the number of mice per cage. Food was changed and measured twice weekly; values were combined to generate the weekly average. Consumption of Viv chow was highest because this diet has the fewest calories per gram (N = 6-12 mice [3-4 cages]/diet). @Statistical difference between PL + CO and ST + CO at 12 weeks on diet. Statistically different from #CO, %ST + CO, &all others using one-way analysis of variance followed by Tukey post hoc analysis for B or two-way ANOVA followed by Tukey post-hoc analysis for A and C.
Figure 7.
Figure 7.
Dietary soybean oil (SO) dysregulates hypothalamic gene expression and plasma levels of oxytocin. C57BL/6N male mice fed Viv chow, CO, SO + CO, PL + CO, or ST + CO diets for 17 to 28 weeks. A, Transcript levels of Oxt, expressed relative to β-actin (Actb) measured in hypothalamic homogenates using reverse transcriptase-quantitative polymerase chain reaction and expressed relative to levels in CO using the Pfaffl method (N = 3-6/diet). B, Peripheral oxytocin peptide levels measured via EIA, (N = 3-10/diet). Statistically different from: #CO; @@@PL + CO; ^SO + CO and PL + CO (single symbols, P < .05; triple symbols P < .001) using 1-way analysis of variance (ANOVA) followed by Tukey post hoc analysis for A and one-way ANOVA followed by Bonferroni post hoc analysis for B.
Figure 8.
Figure 8.
Dietary soybean oil decreases oxytocin immunoreactivity in the magnocellular neuroendocrine nuclei of the hypothalamus. Oxytocin-neurophysin immunoreactivity in micrographs of hypothalamic tissue sections from perfused brains of male mice fed Viv chow (Viv), CO, SO + CO, PL + CO, or ST + CO diets for 17 to 28 weeks. Reduced immunofluorescence was observed in SO + CO, PL + CO, and ST + CO compared to CO and Viv chow in the large magnocellular neuroendocrine of the SON and PVN and smaller parvocellular neurons of the PVN. 3V, third ventricle; arrow, magnocellular neuroendocrine cells; arrowhead, parvocellular neurons; asterisk, immunolabeled axonal projections; dashed line, PVN boundary in SO + CO and ST + CO; OX, optic chiasm; PVN, paraventricular nucleus; SON, supraoptic nucleus. Calibration bar = 100 μm.

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