Omega-6 and Omega-3 Fatty Acid-Derived Oxylipins from the Lipoxygenase Pathway in Maternal and Umbilical Cord Plasma at Delivery and Their Relationship with Infant Growth.

Omega-3 and omega-6 fatty acids are important for neonatal development and health. One mechanism by which omega-3 and omega-6 fatty acids exert their effects is through their metabolism into oxylipins and specialized pro-resolving mediators. However, the influence of oxylipins on fetal growth is not well understood. Therefore, the objective of this study was to identify oxylipins present in maternal and umbilical cord plasma and investigate their relationship with infant growth. Liquid chromatography-tandem mass spectrometry was used to quantify oxylipin levels in plasma collected at the time of delivery. Spearman's correlations highlighted significant correlations between metabolite levels and infant growth. They were then adjusted for maternal obesity (normal body mass index (BMI: ≤30 kg/m2) vs. obese BMI (>30 kg/m2) and smoking status (never vs. current/former smoker) using linear regression modeling. A p-value < 0.05 was considered statistically significant. Our study demonstrated a diverse panel of oxylipins from the lipoxygenase pathway present at the time of delivery. In addition, both omega-3 and omega-6 oxylipins demonstrated potential influences on the birth length and weight percentiles. The oxylipins present during pregnancy may influence fetal growth and development, suggesting potential metabolites to be used as biomarkers for infant outcomes.

Concentrations of fat-soluble nutrients and blood inflammatory compounds in mother-infant dyads at birth.

BACKGROUND: Perinatal inflammation adversely affects health. Therefore, aims of this IRB-approved study are: (1) compare inflammatory compounds within and between maternal and umbilical cord blood samples at the time of delivery, (2) assess relationships between inflammatory compounds in maternal and cord blood with birth characteristics/outcomes, and (3) assess relationships between blood and placental fat-soluble nutrients with blood levels of individual inflammatory compounds. METHODS: Mother-infant dyads were enrolled (n = 152) for collection of birth data and biological samples of maternal blood, umbilical cord blood, and placental tissue. Nutrient levels included: lutein + zeaxanthin; lycopene; α-, ß-carotene; ß-cryptoxanthin; retinol; α-, γ-, δ-tocopherol. Inflammatory compounds included: tumor necrosis factor-α, superoxide dismutase, interleukins (IL) 1ß, 2, 6, 8, 10. RESULTS: Median inflammatory compound levels were 1.2-2.3 times higher in cord vs. maternal blood, except IL2 (1.3 times lower). Multiple significant correlations existed between maternal vs. infant inflammatory compounds (range of r = 0.22-0.48). While relationships existed with blood nutrient levels, the most significant were identified in placenta where all nutrients (except δ-tocopherol) exhibited relationships with inflammatory compounds. Relationships between anti-inflammatory nutrients and proinflammatory compounds were primarily inverse. CONCLUSION: Inflammation is strongly correlated between mother-infant dyads. Fat-soluble nutrients have relationships with inflammatory compounds, suggesting nutrition is a modifiable factor. IMPACT: Mother and newborn inflammation status are strongly interrelated. Levels of fat-soluble nutrients in blood, but especially placenta, are associated with blood levels of proinflammatory and anti-inflammatory compounds in both mother and newborn infant. As fat-soluble nutrient levels are associated with blood inflammatory compounds, nutrition is a modifiable factor to modulate inflammation and improve perinatal outcomes.

Omega-3 Fatty Acid-Derived Resolvin D2 Regulates Human Placental Vascular Smooth Muscle and Extravillous Trophoblast Activities.

Omega-3 fatty acids are important to pregnancy and neonatal development and health. One mechanism by which omega-3 fatty acids exert their protective effects is through serving as substrates for the generation of specialized pro-resolving lipid mediators (SPM) that potently limit and resolve inflammatory processes. We recently identified that SPM levels are increased in maternal blood at delivery as compared to umbilical cord blood, suggesting the placenta as a potential site of action for maternal SPM. To explore this hypothesis, we obtained human placental samples and stained for the SPM resolvin D2 (RvD2) receptor GPR18 via immunohistochemistry. In so doing, we identified GPR18 expression in placental vascular smooth muscle and extravillous trophoblasts of the placental tissues. Using in vitro culturing, we confirmed expression of GPR18 in these cell types and further identified that stimulation with RvD2 led to significantly altered responsiveness (cytoskeletal changes and pro-inflammatory cytokine production) to lipopolysaccharide inflammatory stimulation in human umbilical artery smooth muscle cells and placental trophoblasts. Taken together, these findings establish a role for SPM actions in human placental tissue.

The impact of iron supplementation on the preterm neonatal gut microbiome: A pilot study.

OBJECTIVE: The gastrointestinal microbiome in preterm infants exhibits significant influence on optimal outcomes-with dysbiosis shown to substantially increase the risk of the life-threatening necrotizing enterocolitis. Iron is a vital nutrient especially during the perinatal window of rapid hemoglobin production, tissue growth, and foundational neurodevelopment. However, excess colonic iron exhibits potent oxidation capacity and alters the gut microbiome-potentially facilitating the proliferation of pathological bacterial strains. Breastfed preterm infants routinely receive iron supplementation starting 14 days after delivery and are highly vulnerable to morbidities associated with gastrointestinal dysbiosis. Therefore, we set out to determine if routine iron supplementation alters the preterm gut microbiome. METHODS: After IRB approval, we collected stool specimens from 14 infants born <34 weeks gestation in the first, second, and fourth week of life to assess gut microbiome composition via 16S rRNA sequencing. RESULTS: We observed no significant differences in either phyla or key genera relative abundance between pre- and post-iron timepoints. We observed notable shifts in infant microbiome composition based on season of delivery. CONCLUSION: Though no obvious indication of iron-induced dysbiosis was observed in this unique study in the setting of prematurity, further investigation in a larger sample is warranted to fully understand iron's impact on the gastrointestinal milieu.

Bioactive metabolites of OMEGA-6 and OMEGA-3 fatty acids are associated with inflammatory cytokine concentrations in maternal and infant plasma at the time of delivery.

BACKGROUND & AIMS: Inflammation is necessary for a healthy pregnancy. However, unregulated or excessive inflammation during pregnancy is associated with severe maternal and infant morbidities, such as pre-eclampsia, abnormal infant neurodevelopment, or preterm birth. Inflammation is regulated in part by the bioactive metabolites of omega-6 (n-6) and omega-3 (n-3) fatty acids (FAs). N-6 FAs have been shown to promote pro-inflammatory cytokine environments in adults, while n-3 FAs have been shown to contribute to the resolution of inflammation; however, how these metabolites affect maternal and infant inflammation is still uncertain. The objective of this study was to predict the influence of n-6 and n-3 FA metabolites on inflammatory biomarkers in maternal and umbilical cord plasma at the time of delivery. METHODS: Inflammatory biomarkers (IL-1ß, IL-2, IL-6, IL-8, IL-10, and TNFα) for maternal and umbilical cord plasma samples in 39 maternal-infant dyads were analyzed via multi-analyte bead array. Metabolites of n-6 FAs (arachidonic acid and linoleic acid) and n-3 FAs (eicosapentaenoic acid and docosahexaenoic acid) were assayed via liquid chromatography-mass spectrometry. Linear regression models assessed relationships between maternal and infant inflammatory markers and metabolite plasma concentrations. RESULTS: Increased plasma concentrations of maternal n-6 metabolites were predictive of elevated pro-inflammatory cytokine concentrations in mothers; similarly, higher plasma concentrations of umbilical cord n-6 FA metabolites were predictive of elevated pro-inflammatory cytokine concentrations in infants. Higher plasma concentrations of maternal n-6 FA metabolites were also predictive of elevated pro-inflammatory cytokines in infants, suggesting that maternal n-6 FA status has an intergenerational impact on the inflammatory status of the infant. In contrast, maternal and cord plasma concentrations of n-3 FA metabolites had a mixed effect on inflammatory status in mothers and infants, which may be due to the inadequate maternal dietary intake of n-3 FAs in our study population. CONCLUSIONS: Our results reveal that maternal FA status may have an intergenerational impact on the inflammatory status of the infant. Additional research is needed to identify how dietary interventions that modify maternal FA intake prior to or during pregnancy may impact maternal and infant inflammatory status and associated long-term health outcomes.

Omega-3 Polyunsaturated Fatty Acid Levels in Maternal and Cord Plasma Are Associated with Maternal Socioeconomic Status.

Omega-3 (n-3) polyunsaturated fatty acids (PUFAs) play a crucial role in fetal growth and neurodevelopment, while omega-6 (n-6) PUFAs have been associated with adverse pregnancy outcomes. Previous studies have demonstrated that socioeconomic status (SES) influences dietary intake of n-3 and n-6 PUFAs, but few studies have evaluated the association between maternal and cord plasma biomarkers of PUFAs and socioeconomic markers. An IRB-approved study enrolled mother-infant pairs (n = 55) at the time of delivery. Maternal and cord plasma PUFA concentrations were analyzed using gas chromatography. Markers of SES were obtained from validated surveys and maternal medical records. Mann-Whitney U tests and linear regression models were utilized for statistical analysis. Maternal eicosapentaenoic acid (EPA) (p = 0.02), cord EPA (p = 0.04), and total cord n-3 PUFA concentrations (p = 0.04) were significantly higher in college-educated mothers vs. mothers with less than a college education after adjustment for relevant confounders. Insurance type and household income were not significantly associated with n-3 or n-6 PUFA plasma concentrations after adjustment. Our findings suggest that mothers with lower educational status may be at risk of lower plasma concentrations of n-3 PUFAs at delivery, which could confer increased susceptibility to adverse pregnancy and birth outcomes.

Something Smells Fishy: How Lipid Mediators Impact the Maternal-Fetal Interface and Neonatal Development.

Normal pregnancy relies on inflammation for implantation, placentation, and parturition, but uncontrolled inflammation can lead to poor maternal and infant outcomes. Maternal diet is one modifiable factor that can impact inflammation. Omega-3 and -6 fatty acids obtained through the diet are metabolized into bioactive compounds that effect inflammation. Recent evidence has shown that the downstream products of omega-3 and -6 fatty acids may influence physiology during pregnancy. In this review, the current knowledge relating to omega-3 and omega-6 metabolites during pregnancy will be summarized.

Retinol and Pro-Vitamin A Carotenoid Nutritional Status during Pregnancy Is Associated with Newborn Hearing Screen Results.

The prenatal period is critical for auditory development; thus, prenatal influences on auditory development may significantly impact long-term hearing ability. While previous studies identified a protective effect of carotenoids on adult hearing, the impact of these nutrients on hearing outcomes in neonates is not well understood. The purpose of this study is to investigate the relationship between maternal and umbilical cord plasma retinol and carotenoid concentrations and abnormal newborn hearing screen (NHS) results. Mother-infant dyads (n = 546) were enrolled at delivery. Plasma samples were analyzed using HPLC and LC-MS/MS. NHS results were obtained from medical records. Statistical analysis utilized Mann-Whitney U tests and logistic regression models, with p ≤ 0.05 considered statistically significant. Abnormal NHS results were observed in 8.5% of infants. Higher median cord retinol (187.4 vs. 162.2 µg/L, p = 0.01), maternal trans-ß-carotene (206.1 vs. 149.4 µg/L, p = 0.02), maternal cis-ß-carotene (15.9 vs. 11.2 µg/L, p = 0.02), and cord trans-ß-carotene (15.5 vs. 8.0 µg/L, p = 0.04) were associated with abnormal NHS. Significant associations between natural log-transformed retinol and ß-carotene concentrations and abnormal NHS results remained after adjustment for smoking status, maternal age, and corrected gestational age. Further studies should investigate if congenital metabolic deficiencies, pesticide contamination of carotenoid-rich foods, maternal hypothyroidism, or other variables mediate this relationship.

Perifacial Lymph Node Metastasis After Treatment of Oropharyngeal Primary Malignancy: A Case Report.

With the rate of oropharyngeal cancer on the rise, appropriate surgical management is an increasingly important consideration. Much debate currently exists regarding the necessary extent of neck dissections when performing curative surgery for primary oropharyngeal malignancies. Here, we present the case of a 64-year-old patient with p16+ T1N1M0 squamous cell carcinoma (SCC) of the right tonsil. Approximately four years following transoral robotic surgery oropharyngectomy and ipsilateral level II-IV right selective neck dissection, metastatic SCC was discovered on fine-needle aspiration biopsy of a right perifacial lymph node (level Ib). The patient then underwent a revision right neck dissection at levels Ia and Ib. Adjuvant immunotherapy was recommended following revision neck dissection. Postoperative imaging and flexible laryngoscopy three months after surgery were not concerning for cervical lymphadenopathy or oropharyngeal lesions. Although rare, physicians must maintain a healthy level of suspicion for recurrence to level Ib in oropharyngeal primary malignancies.

Intrauterine Transfer of Polyunsaturated Fatty Acids in Mother-Infant Dyads as Analyzed at Time of Delivery.

Polyunsaturated fatty acids (PUFAs) are essential for fetal development, and intrauterine transfer is the only supply of PUFAs to the fetus. The prevailing theory of gestational nutrient transfer is that certain nutrients (including PUFAs) may have prioritized transport across the placenta. Numerous studies have identified correlations between maternal and infant fatty acid concentrations; however, little is known about what role maternal PUFA status may play in differential intrauterine nutrient transfer. Twenty mother-infant dyads were enrolled at delivery for collection of maternal and umbilical cord blood, and placental tissue samples. Plasma concentrations of PUFAs were assessed using gas chromatography (GC-FID). Intrauterine transfer percentages for each fatty acid were calculated as follows: ((cord blood fatty acid level/maternal blood fatty acid level) × 100). Kruskal-Wallis tests were used to compare transfer percentages between maternal fatty acid tertile groups. A p-value < 0.05 was considered significant. There were statistically significant differences in intrauterine transfer percentages of arachidonic acid (AA) (64% vs. 65% vs. 45%, p = 0.02), eicosapentaenoic acid (EPA) (41% vs. 19% vs. 17%, p = 0.03), and total fatty acids (TFA) (27% vs. 26% vs. 20%, p = 0.05) between maternal plasma fatty acid tertiles. Intrauterine transfer percentages of AA, EPA, and TFA were highest in the lowest tertile of respective maternal fatty acid concentration. These findings may indicate that fatty acid transfer to the fetus is prioritized during gestation even during periods of maternal nutritional inadequacy.

QuPath Digital Immunohistochemical Analysis of Placental Tissue.

BACKGROUND: QuPath is an open-source digital image analyzer notable for its user-friendly design, cross-platform compatibility, and customizable functionality. Since it was first released in 2016, at least 624 publications have reported its use, and it has been applied in a wide spectrum of settings. However, there are currently limited reports of its use in placental tissue. Here, we present the use of QuPath to quantify staining of G-protein coupled receptor 18 (GPR18), the receptor for the pro-resolving lipid mediator Resolvin D2, in placental tissue. METHODS: Whole slide images of vascular smooth muscle (VSM) and extravillous trophoblast (EVT) cells stained for GPR18 were annotated for areas of interest. Visual scoring was performed on these images by trained and in-training pathologists, while QuPath scoring was performed with the methodology described herein. RESULTS: Bland-Altman analyses showed that, for the VSM category, the two methods were comparable across all staining levels. For EVT cells, the high-intensity staining level was comparable across methods, but the medium and low staining levels were not comparable. CONCLUSIONS: Digital image analysis programs offer great potential to revolutionize pathology practice and research by increasing accuracy and decreasing the time and cost of analysis. Careful study is needed to optimize this methodology further.

Sex-Dependent Mechanisms of Glucocorticoid Regulation of the Mouse Hypothalamic Corticotropin-Releasing Hormone Gene.

To limit excessive glucocorticoid secretion following hypothalamic-pituitary-adrenal (HPA) axis stimulation, circulating glucocorticoids inhibit corticotropin-releasing hormone (CRH) expression in paraventricular nucleus (PVN) neurons. As HPA function differs between sexes and depends on circulating estradiol (E2) levels in females, we investigated sex/estrous stage-dependent glucocorticoid regulation of PVN Crh. Using NanoString nCounter technology, we first demonstrated that adrenalectomized (ADX'd) diestrous female (low E2), but not male or proestrous female (high E2), mice exhibited a robust decrease in PVN CRH mRNA following 2-day treatment with the glucocorticoid receptor (GR) agonist RU28362. Immunohistochemical analysis of PVN CRH neurons in Crh-IRES-Cre;Ai14 mice, where TdTomato fluorescence permanently tags CRH-expressing neurons, showed similarly abundant co-expression of GR-immunoreactivity in males, diestrous females, and proestrous females. However, we identified sex/estrous stage-related glucocorticoid regulation or expression of GR transcriptional coregulators. Out of 17 coregulator genes examined using nCounter multiplex analysis, mRNAs that were decreased by RU28362 in ADX'd mice in a sex/estrous stage-dependent fashion included: GR (males = diestrous females > proestrous females), signal transducer and activator of transcription 3 (STAT3) (males < diestrous = proestrous), and HDAC1 (males < diestrous > proestrous). Steroid receptor coactivator 3 (SRC-3), nuclear corepressor 1 (NCoR1), heterogeneous nuclear ribonucleoprotein U (hnrnpu), CREB binding protein (CBP) and CREB-regulated transcription coactivator 2 (CRTC2) mRNAs were lower in ADX'd diestrous and proestrous females versus males. Additionally, most PVN CRH neurons co-expressed methylated CpG binding protein 2 (MeCP2)-immunoreactivity in diestrous female and male Crh-IRES-Cre;Ai14 mice. Our findings collectively suggest that GR's sex-dependent regulation of PVN Crh may depend upon differences in the GR transcriptional machinery and an underlying influence of E2 levels in females.

Effect of Maternal Retinol Status at Time of Term Delivery on Retinol Placental Concentration, Intrauterine Transfer Rate, and Newborn Retinol Status.

Retinol (vitamin A) is essential, so the objective of this Institutional Review Board approved study is to evaluate retinol placental concentration, intrauterine transfer, and neonatal status at time of term delivery between cases of maternal retinol adequacy, insufficiency, and deficiency in a United States population. Birth information and biological samples were collected for mother-infant dyads (n = 260). Maternal and umbilical cord blood retinol concentrations (n = 260) were analyzed by HPLC and categorized: deficient (≤0.7 umol/L), insufficient (>0.7-1.05 umol/L), adequate (>1.05 umol/L). Intrauterine transfer rate was calculated: (umbilical cord blood retinol concentration/maternal retinol concentration) × 100. Non-parametric statistics used include Spearman's correlations, Mann-Whitney U, and Kruskal-Wallis tests. p-values <0.05 were statistically significant. Only 51.2% of mothers were retinol adequate, with 38.4% insufficient, 10.4% deficient. Only 1.5% of infants were retinol adequate. Placental concentrations (n = 73) differed between adequate vs. deficient mothers (median 0.13 vs. 0.10 µg/g; p = 0.003). Umbilical cord blood concentrations were similar between deficient, insufficient, and adequate mothers (0.61 vs. 0.55 vs. 0.57 µmol/L; p = 0.35). Intrauterine transfer increased with maternal deficiency (103.4%) and insufficiency (61.2%) compared to adequacy (43.1%), p < 0.0001. Results indicate that intrauterine transfer rate is augmented in cases of maternal retinol inadequacy, leading to similar concentrations in umbilical cord blood at term delivery.

Fat-soluble nutrients and Omega-3 fatty acids as modifiable factors influencing preterm birth risk.

Preterm birth is a leading cause of child morbidity and mortality, so strategies to reduce early birth must remain a priority. One key approach to enhancing birth outcomes is improving maternal dietary intake. Therefore, the purpose of this review is to discuss mechanisms on perinatal status of fat-soluble nutrients (carotenoids, retinol, tocopherols) and omega-3 fatty acids and how they impact risk for preterm birth. Literature review demonstrates that maternal dietary intake and biological (blood and placental tissue) levels of fat-soluble nutrients during pregnancy may provide antioxidative, anti-inflammatory, and immunomodulatory health benefits. Omega-3 fatty acids also promote increased production of specialized pro-resolving mediators, subsequently mediating inflammation resolution. Combined effects of these nutrients support appropriate placental organogenesis and function. Consequently, fat-soluble nutrients and omega-3 fatty acids serve as strong influencers for preterm birth risk. As dietary intake remains a modifiable factor, future intervention would benefit from a focus on optimizing perinatal status of these specific nutrients.

Omega-3 Long-Chain Polyunsaturated Fatty Acids Intake by Ethnicity, Income, and Education Level in the United States: NHANES 2003-2014.

Although there are many recognized health benefits for the consumption of omega-3 (n-3) long-chain polyunsaturated fatty acids (LCPUFA), intake in the United States remains below recommended amounts. This analysis was designed to provide an updated assessment of fish and n-3 LCPUFA intake (eicosapentaenoic (EPA), docosahexaenoic acid (DHA), and EPA+DHA) in the United States adult population, based on education, income, and race/ethnicity, using data from the 2003-2014 National Health and Nutrition Examination Survey (NHANES) (n = 44,585). Over this survey period, participants with less education and lower income had significantly lower n-3 LCPUFA intakes and fish intakes (p < 0.001 for all between group comparisons). N-3 LCPUFA intake differed significantly according to ethnicity (p < 0.001), with the highest intake of n-3 LCPUFA and fish in individuals in the "Other" category (including Asian Americans). Supplement use increased EPA + DHA intake, but only 7.4% of individuals consistently took supplements. Overall, n-3 LCPUFA intake in this study population was low, but our findings indicate that individuals with lower educational attainment and income are at even higher risk of lower n-3 LCPUFA and fish intake.

Omega-3 Fatty Acid Intake by Age, Gender, and Pregnancy Status in the United States: National Health and Nutrition Examination Survey 2003⁻2014.

Despite the importance of n-3 fatty acids for health, intakes remain below recommended levels. The objective of this study was to provide an updated assessment of fish and n-3 fatty acid intake (i.e., eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and EPA+DHA) in the United States using the 2003⁻2014 National Health and Nutrition Examination Survey (NHANES) data (n = 45,347)). Over this survey period, toddlers, children, and adolescents (aged 1⁻19) had significantly lower n-3 fatty acid intake (p < 0.001) compared to adults and seniors, which remained significant after adjusting for caloric intake. Females demonstrated lower n-3 fatty acid intake than males (p < 0.001), with adult and senior women having significantly lower intakes compared to men in the same age categories (p < 0.001) after adjustment for energy intake. Women also consumed less fish than men (5.8 versus 6.1 servings/month, p < 0.001). The estimated intakes of n-3 fatty acids in pregnant women did not differ from non-pregnant women (p = 0.6 for EPA+DHA), although pregnant women reported consuming less high n-3 fatty acid-containing fish than non-pregnant women (1.8 versus 2.6 servings/month, p < 0.001). Our findings indicate that subgroups of the population may be at higher risk of n-3 fatty acid intakes below recommended levels.

Distribution and chemical composition of estrogen receptor ß neurons in the paraventricular nucleus of the female and male mouse hypothalamus.

Activation of estrogen receptor beta (ERß)-expressing neurons regulates the mammalian stress response via the hypothalamic-pituitary-adrenal (HPA) axis. These neurons densely populate the paraventricular nucleus of the hypothalamus (PVN). Recent research has revealed striking differences between rat and mouse PVN cytochemistry, but careful exploration of PVN ERß neurons in mice has been hindered by a lack of specific ERß antisera. Therefore, we used male and female transgenic mice expressing EGFP under the control of the mouse ERß promoter (ERß-EGFP) to examine the chemical architecture of PVN ERß cells. Using immunohistochemistry, we found that 90% of ERß-immunoreactivity (-ir) colocalized with EGFP. Cellular colocalization of EGFP with neuropeptides, transcription modulators, and neuronal tracers was examined throughout the PVN. ERß-EGFP cells expressed oxytocin more abundantly in the rostral (71 ± 3%) than caudal (33 ± 8%) PVN. Arginine vasopressin colocalized with EGFP more often in females (18 ± 3%) than males (4 ± 1%). Moreover, estrogen receptor α-ir colocalized with ERß-EGFP at low levels (15 ± 3%). Using a corticotropin releasing hormone-cre driver X tdTomato reporter mouse, we found a moderate colocalization with ERß-ir (48 ± 16%) in the middle PVN. Peripheral injection of fluorogold revealed that the rostral PVN ERß-EGFP cells are neuroendocrine neurons whereas non-neuroendocrine (presumably pre-autonomic) ERß-EGFP neurons predominated in the posterior PVN. These data demonstrate chemoarchitectural differences in ERß neurons of the mouse PVN that are different from that previously described for the rat, thus, elucidating potential neuronal pathways involved in the regulation of the HPA axis in mice.