Fetal programming and lactation: modulating gene expression in response to undernutrition during intrauterine life.

BACKGROUND: Adverse environmental conditions during intrauterine life, known as fetal programming, significantly contribute to the development of diseases in adulthood. Fetal programming induced by factors like maternal undernutrition leads to low birth weight and increases the risk of cardiometabolic diseases. METHODS: We studied a rat model of maternal undernutrition during gestation (MUN) to investigate gene expression changes in cardiac tissue using RNA-sequencing of day 0-1 litters. Moreover, we analyzed the impact of lactation at day 21, in MUN model and cross-fostering experiments, on cardiac structure and function assessed by transthoracic echocardiography, and gene expression changes though qPCR. RESULTS: Our analysis identified specific genes with altered expression in MUN rats at birth. Two of them, Agt and Pparg, stand out for being associated with cardiac hypertrophy and fibrosis. At the end of the lactation period, MUN males showed increased expression of Agt and decreased expression of Pparg, correlating with cardiac hypertrophy. Cross-fostering experiments revealed that lactation with control breastmilk mitigated these expression changes reducing cardiac hypertrophy in MUN males. CONCLUSIONS: Our findings highlight the interplay between fetal programming, gene expression, and cardiac hypertrophy suggesting that lactation period is a potential intervention window to mitigate the effects of fetal programming. IMPACT: Heart remodeling involves the alteration of several groups of genes and lactation period plays a key role in establishing gene expression modification caused by fetal programming. We could identify expression changes of relevant genes in cardiac tissue induced by undernutrition during fetal life. We expose the contribution of the lactation period in modulating the expression of Agt and Pparg, relevant genes associated with cardiac hypertrophy. This evidence reveal lactation as a crucial intervention window for preventing or countering fetal programming.

Assessment of Sleep Quality in Spanish Twin Pregnancy: An Observational Single-Center Study.

Women with twin pregnancies experience greater sleep disturbance compared to women with singleton pregnancies. The aims of this study were to explore the sleep quality in women with twin pregnancies and to compare their sleep dimensions with coetaneous single pregnancies. This was an observational study in which women were enrolled at the end of pregnancy in the Obstetric Service of Hospital La Paz (Spain). The women were classified as single (n = 143) or twin pregnancy (n = 62). Pregnant women responded to the Pittsburgh Sleep Quality Index to evaluate sleep quality, latency, duration, efficiency, perturbance, use of medication, and daytime dysfunction. The higher the index, the greater the alteration of sleep quality. Without statistical differences, a poor sleep quality was higher in women with single (66.7%) than women with twin pregnancies (22.8%). The good sleeper slept 6.8 h/day in single pregnancy and 7.3 h/day in twin pregnancy. The sleep perturbation and dysfunctionality were higher in women with twin than single pregnancies. The use of medication to sleep was significantly lower in women with twin than single pregnancies. In women with twin pregnancy, the body weight gain during first trimester had a positive correlation with worse sleep quality and sleep perturbations. Twin pregnancy needed more than 7 h/day to have a high sleep quality, showing greater sleep perturbations and daytime dysfunction than single pregnancies. The control of gestational body weight can improve the sleep quality, disturbances, and duration in twin gestations. Sleep screening during pregnancy would be necessary to handle sleep issues and increase benefits in twin gestational outcomes.

Effects of Zinc on the Right Cardiovascular Circuit in Long-Term Hypobaric Hypoxia in Wistar Rats.

Hypobaric hypoxia under chromic conditions triggers hypoxic pulmonary vasoconstriction (HPV) and right ventricular hypertrophy (RVH). The role of zinc (Zn) under hypoxia is controversial and remains unclear. We evaluated the effect of Zn supplementation in prolonged hypobaric hypoxia on HIF2α/MTF-1/MT/ZIP12/PKCε pathway in the lung and RVH. Wistar rats were exposed to hypobaric hypoxia for 30 days and randomly allocated into three groups: chronic hypoxia (CH); intermittent hypoxia (2 days hypoxia/2 days normoxia; CIH); and normoxia (sea level control; NX). Each group was subdivided (n = 8) to receive either 1% Zn sulfate solution (z) or saline (s) intraperitoneally. Body weight, hemoglobin, and RVH were measured. Zn levels were evaluated in plasma and lung tissue. Additionally, the lipid peroxidation levels, HIF2α/MTF-1/MT/ZIP12/PKCε protein expression and pulmonary artery remodeling were measured in the lung. The CIH and CH groups showed decreased plasma Zn and body weight and increased hemoglobin, RVH, and vascular remodeling; the CH group also showed increased lipid peroxidation. Zn administration under hypobaric hypoxia upregulated the HIF2α/MTF-1/MT/ZIP12/PKCε pathway and increased RVH in the intermittent zinc group. Under intermittent hypobaric hypoxia, Zn dysregulation could participate in RVH development through alterations in the pulmonary HIF2α/MTF1/MT/ZIP12/PKCε pathway.

AMPK and the Challenge of Treating Hypoxic Pulmonary Hypertension.

Hypoxic pulmonary hypertension (HPH) is characterized by sustained elevation of pulmonary artery pressure produced by vasoconstriction and hyperproliferative remodeling of the pulmonary artery and subsequent right ventricular hypertrophy (RVH). The search for therapeutic targets for cardiovascular pathophysiology has extended in many directions. However, studies focused on mitigating high-altitude pulmonary hypertension (HAPH) have been rare. Because AMP-activated protein kinase (AMPK) is involved in cardiovascular and metabolic pathology, AMPK is often studied as a potential therapeutic target. AMPK is best characterized as a sensor of cellular energy that can also restore cellular metabolic homeostasis. However, AMPK has been implicated in other pathways with vasculoprotective effects. Notably, cellular metabolic stress increases the intracellular ADP/ATP or AMP/ATP ratio, and AMPK activation restores ATP levels by activating energy-producing catabolic pathways and inhibiting energy-consuming anabolic pathways, such as cell growth and proliferation pathways, promoting cardiovascular protection. Thus, AMPK activation plays an important role in antiproliferative, antihypertrophic and antioxidant pathways in the pulmonary artery in HPH. However, AMPK plays contradictory roles in promoting HPH development. This review describes the main findings related to AMPK participation in HPH and its potential as a therapeutic target. It also extrapolates known AMPK functions to discuss the less-studied HAPH context.

Fetal Undernutrition Modifies Vascular RAS Balance Enhancing Oxidative Damage and Contributing to Remodeling.

Fetal stress is known to increase susceptibility to cardiometabolic diseases and hypertension in adult age in a process known as fetal programming. This study investigated the relationship between vascular RAS, oxidative damage and remodeling in fetal programming. Six-month old Sprague-Dawley offspring from mothers that were fed ad libitum (CONTROL) or with 50% intake during the second half of gestation (maternal undernutrition, MUN) were used. qPCR or immunohistochemistry were used to obtain the expression of receptors and enzymes. Plasma levels of carbonyls were measured by spectrophotometry. In mesenteric arteries from MUN rats we detected an upregulation of ACE, ACE2, AT1 receptors and NADPH oxidase, and lower expression of AT2, Mas and MrgD receptors compared to CONTROL. Systolic and diastolic blood pressure and plasma levels of carbonyls were higher in MUN than in CONTROL. Vascular morphology evidenced an increased media/lumen ratio and adventitia/lumen ratio, and more connective tissue in MUN compared to CONTROL. In conclusion, fetal undernutrition indices RAS alterations and oxidative damage which may contribute to the remodeling of mesenteric arteries, and increase the risk of adverse cardiovascular events and hypertension.

Male fetal sex is associated with low maternal plasma anti-inflammatory cytokine profile in the first trimester of healthy pregnancies.

Male fetal sex associates with higher rates of materno-fetal complications. Inflammation and inadequate vasoactive responses are mechanisms implicated in obstetric complications, and alterations in maternal plasma cytokine profile and nitric oxide (NO) metabolites are potential predictive biomarkers. We aimed to assess if these parameters are influenced by fetal sex. A prospective, observational study was carried out in 85 healthy pregnant women with singleton pregnancies in the first trimester of gestation. A blood sample was extracted at the tenth week of gestation. In plasma, we assessed: 1) cytokines (micro-array): pro-inflammatory (IL1α, IL1 ß, IL6, TNFα), anti-inflammatory (IL4, IL10, IL13), and chemoattractant (IL8, MCP1, IFNγ), and 2) NO metabolites (liquid chromatography-tandem mass spectrometry and Griess reaction): L-arginine, ADMA, SDMA, nitrates (NOx). Women with a male fetus (n = 50) exhibited, compared with those with a female (n = 35): higher IL1ß (OR = 1.09 with 95% CI: 0.97-1.28), and lower IL13 (OR = 0.93 with 95% CI: 0.87-0.99), and higher plasma NOx (OR = 1.14 with 95% CI: 1.03-1.31). Our data suggest that fetal sex influences maternal plasma cytokine profile and NO in early pregnancy. Women with a male fetus may have a worse capacity to counteract an inflammatory response. They may have better vasodilator capacity, but in the presence of an oxidative environment, a higher nitrosative damage may occur. These data reinforce the need to include sex as variable in predictive models.

Sex Differences in Placental Protein Expression and Efficiency in a Rat Model of Fetal Programming Induced by Maternal Undernutrition.

Fetal undernutrition programs cardiometabolic diseases, with higher susceptibility in males. The mechanisms implicated are not fully understood and may be related to sex differences in placental adaptation. To evaluate this hypothesis, we investigated placental oxidative balance, vascularization, glucocorticoid barrier, and fetal growth in rats exposed to 50% global nutrient restriction from gestation day 11 (MUN, n = 8) and controls (n = 8). At gestation day 20 (G20), we analyzed maternal, placental, and fetal weights; oxidative damage, antioxidants, corticosterone, and PlGF (placental growth factor, spectrophotometry); and VEGF (vascular endothelial growth factor), 11ß-HSD2, p22phox, XO, SOD1, SOD2, SOD3, catalase, and UCP2 expression (Western blot). Compared with controls, MUN dams exhibited lower weight and plasma proteins and higher corticosterone and catalase without oxidative damage. Control male fetuses were larger than female fetuses. MUN males had higher plasma corticosterone and were smaller than control males, but had similar weight than MUN females. MUN male placenta showed higher XO and lower 11ß-HSD2, VEGF, SOD2, catalase, UCP2, and feto-placental ratio than controls. MUN females had similar feto-placental ratio and plasma corticosterone than controls. Female placenta expressed lower XO, 11ß-HSD2, and SOD3; similar VEGF, SOD1, SOD2, and UCP2; and higher catalase than controls, being 11ß-HSD2 and VEGF higher compared to MUN males. Male placenta has worse adaptation to undernutrition with lower efficiency, associated with oxidative disbalance and reduced vascularization and glucocorticoid barrier. Glucocorticoids and low nutrients may both contribute to programming in MUN males.

Nox2 Upregulation and p38α MAPK Activation in Right Ventricular Hypertrophy of Rats Exposed to Long-Term Chronic Intermittent Hypobaric Hypoxia.

One of the consequences of high altitude (hypobaric hypoxia) exposure is the development of right ventricular hypertrophy (RVH). One particular type of exposure is long-term chronic intermittent hypobaric hypoxia (CIH); the molecular alterations in RVH in this particular condition are less known. Studies show an important role of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex-induced oxidative stress and protein kinase activation in different models of cardiac hypertrophy. The aim was to determine the oxidative level, NADPH oxidase expression and MAPK activation in rats with RVH induced by CIH. Male Wistar rats were randomly subjected to CIH (2 days hypoxia/2 days normoxia; n = 10) and normoxia (NX; n = 10) for 30 days. Hypoxia was simulated with a hypobaric chamber. Measurements in the RV included the following: hypertrophy, Nox2, Nox4, p22phox, LOX-1 and HIF-1α expression, lipid peroxidation and H2O2 concentration, and p38α and Akt activation. All CIH rats developed RVH and showed an upregulation of LOX-1, Nox2 and p22phox and an increase in lipid peroxidation, HIF-1α stabilization and p38α activation. Rats with long-term CIH-induced RVH clearly showed Nox2, p22phox and LOX-1 upregulation and increased lipid peroxidation, HIF-1α stabilization and p38α activation. Therefore, these molecules may be considered new targets in CIH-induced RVH.

Role of fetal nutrient restriction and postnatal catch-up growth on structural and mechanical alterations of rat aorta.

KEY POINTS: Intrauterine growth restriction (IUGR), induced by maternal undernutrition, leads to impaired aortic development. This is followed by hypertrophic remodelling associated with accelerated growth during lactation. Fetal nutrient restriction is associated with increased aortic compliance at birth and at weaning, but not in adult animals. This mechanical alteration may be related to a decreased perinatal collagen deposition. Aortic elastin scaffolds purified from young male and female IUGR animals also exhibit increased compliance, only maintained in adult IUGR females. These mechanical alterations may be related to differences in elastin deposition and remodelling. Fetal undernutrition induces similar aortic structural and mechanical alterations in young male and female rats. Our data argue against an early mechanical cause for the sex differences in hypertension development induced by maternal undernutrition. However, the larger compliance of elastin in adult IUGR females may contribute to the maintenance of a normal blood pressure level. ABSTRACT: Fetal undernutrition programmes hypertension development, males being more susceptible. Deficient fetal elastogenesis and vascular growth is a possible mechanism. We investigated the role of aortic mechanical alterations in a rat model of hypertension programming, evaluating changes at birth, weaning and adulthood. Dams were fed ad libitum (Control) or 50% of control intake during the second half of gestation (maternal undernutrition, MUN). Offspring aged 3 days, 21 days and 6 months were studied. Blood pressure was evaluated in vivo. In the thoracic aorta we assessed gross structure, mechanical properties (intact and purified elastin), collagen and elastin content and internal elastic lamina (IEL) organization. Only adult MUN males developed hypertension (systolic blood pressure: MUNmales  = 176.6 ± 5.6 mmHg; Controlmales  = 136.1 ± 4.9 mmHg). At birth MUN rats were lighter, with smaller aortic cross-sectional area (MUNmales  = (1.51 ± 0.08) × 105  µm2 , Controlmales  = (2.8 ± 0.04) × 105  µm2 ); during lactation MUN males and females exhibited catch-up growth and aortic hypertrophy (MUNmales  = (14.5 ± 0.5) × 105  µm2 , Controlmales  = (10.4 ± 0.9) × 105  µm2 ), maintained until adulthood. MUN aortas were more compliant until weaning (functional stiffness: MUNmales  = 1.0 ± 0.04; Controlmales  = 1.3 ± 0.03), containing less collagen with larger IEL fenestrae, returning to normal in adulthood. Purified elastin from young MUN offspring was more compliant in both sexes; only MUN adult females maintained larger elastin compliance (slope: MUNfemales  = 24.1 ± 1.9; Controlfemales  = 33.3 ± 2.8). Fetal undernutrition induces deficient aortic development followed by hypertrophic remodelling and larger aortic compliance in the perinatal period, with similar alterations in collagen and elastin in both sexes. The observed alterations argue against an initial mechanical cause for sex differences in hypertension development. However, the maintenance of high elastin compliance in adult females might protect them against blood pressure rise.

Effects of Arachidonic and Docosohexahenoic Acid Supplementation during Gestation in Rats. Implication of Placental Oxidative Stress.

Arachidonic and docosahexaenoic acids (ARA and DHA) are important during pregnancy. However, the effects of dietary supplementation on fetal growth and oxidative stress are inconclusive. We aimed to assess the effect of high ARA and DHA diet during rat gestation on: (1) ARA and DHA availability in plasma and placenta, (2) fetal growth, and (3) placental oxidative stress, analyzing the influence of sex. Experimental diet (ED) was prepared by substituting soybean oil in the control diet (CD) by a fungi/algae-based oil containing ARA and DHA (2:1). Rats were fed with CD or ED during gestation; plasma, placenta, and fetuses were obtained at gestational day 20. DHA, ARA, and their precursors were analyzed in maternal plasma and placenta by gas chromatography/mass spectrophotometry. Fetuses and placentas were weighed, the proportion of fetuses with intrauterine growth restriction (IUGR) determined, and placental lipid and protein oxidation analyzed. ED fetuses exhibited lower body weight compared to CD, being >40% IUGR; fetal weight negatively correlated with maternal plasma ARA, but not DHA. Only ED female placenta exhibited higher lipid and protein oxidation compared to its CD counterparts; lipid peroxidation is negatively associated with fetal weight. In conclusion, high ARA during gestation associates with IUGR, through placental oxidative stress, with females being more susceptible.

A plasma oxidative stress global index in early stages of chronic venous insufficiency.

BACKGROUND: Chronic venous insufficiency (CVI) represents a social and health care problem because it affects working age populations, particularly in jobs requiring orthostasis, has no effective pharmacologic treatment, and requires surgery. Oxidative stress is present in varicose veins, but whether this is reflected in the plasma is controversial. We aimed to quantify plasma oxidative stress biomarkers in the early stages of CVI and calculate a global index of oxidative stress representative of the disease. METHODS: Plasma was obtained from blood samples of nine patients with CEAP C2 stage CVI and 10 healthy controls. Biomarkers related to antioxidant defense systems (total thiols, reduced glutathione, uric acid, total antioxidant capacity, catalase), oxidative damage (malondialdehyde-bound protein, protein carbonyls, advanced oxidation products, and 3-nitrotyrosine), and activity of enzymes producing key free radicals (xanthine oxidase and myeloperoxidase) were assessed. RESULTS: Compared with the controls, CVI patients exhibited decreased catalase activity and thiol levels and increased malondialdehyde-bound protein and protein carbonyls. These parameters were used to calculate the global index of oxidative stress in CVI, which was significantly different between groups. CONCLUSIONS: It is possible to detect significant changes in plasma oxidative stress biomarkers in early stages of CVI and to calculate a global index representative of the oxidative status in an individual. This index, with the appropriate validation in a larger population, could be used for early detection or progression of CVI.

Impaired inhibitory function of presynaptic A1-adenosine receptors in SHR mesenteric arteries.

In hypertension, vascular reactivity alterations have been attributed to numerous factors, including higher sympathetic innervation/adenosine. This study examined the modulation of adenosine receptors on vascular sympathetic nerves and their putative contribution to higher noradrenaline spillover in hypertension. We assessed adenosine receptors distribution in the adventitia through confocal microscopy, histomorphometry, and their regulatory function on electrically-evoked [(3)H]-noradrenaline overflow, using selective agonists/antagonists. We found that: i) A1-adenosine receptor agonist (CPA: 100 nM) inhibited tritium overflow to a lower extent in SHR (25% ± 3%, n = 14) compared to WKY (38% ± 3%, n = 14) mesenteric arteries; ii) A2A-adenosine receptor agonist (CGS 21680: 100 nM) induced a slight increase of tritium overflow that was similar in SHR (22% ± 8%, n = 8) and WKY (24% ± 5%, n = 8) mesenteric arteries; iii) A2B- and A3-adenosine receptors did not alter tritium overflow in either strain; iv) all adenosine receptors were present on mesenteric artery sympathetic nerves and/or some adventitial cells of both strains; and v) A1-adenosine receptor staining fractional area was lower in SHR than in WKY mesenteric arteries. We conclude that there is an impaired inhibitory function of vascular presynaptic A1-adenosine receptors in SHR, likely related to a reduced presence of these receptors on sympathetic innervation, which might lead to higher levels of noradrenaline in the synaptic cleft and contribute to hypertension in this strain.

A simple dot-blot-Sirius red-based assay for collagen quantification.

The assessment of collagen content in tissues is important in biomedical research, since this protein is altered in numerous diseases. Hydroxyproline and Sirius red based assays are the most common methods for collagen quantification. However, these procedures have some pitfalls, such as the requirement of oxygen-free medium or expensive equipment and large sample size or being unsuitable for hydrolyzed collagen, respectively. Our objective was to develop a specific, versatile, and user-friendly quantitative method applicable to small tissue samples and extracts obtained from elastin purification, therefore, suitable for simultaneous quantification of elastin. This method is based on the binding of Sirius red to collagen present in a sample immobilized on a PVDF membrane, as in the dot-blot technique, and quantified by a scanner and image analysis software. Sample loading, Sirius red concentration, temperature and incubation time, type of standard substance, albumin interference, and quantification time are optimized. The method enabled the quantification of (1) intact collagen in several rat tissue homogenates, including small resistance-sized arteries, (2) partially hydrolyzed collagen obtained from NaOH extracts, compatible with elastin purification, and (3) for the detection of differences in collagen content between hypertensive and normotensive rats. We conclude that the developed technique can be widely used since it is versatile (quantifies intact and hydrolyzed collagen), requires small sample volumes, is user-friendly (low-cost, easy to use, minimum toxic materials, and reduced time of test), and is specific (minimal interference with serum albumin).

Factors Associated with Obstetric Violence Implicated in the Development of Postpartum Depression and Post-Traumatic Stress Disorder: A Systematic Review.

Postpartum depression (PPD) and post-traumatic stress disorder (PTSD) continue to be prevalent, and disabling women with mental disorders and obstetric violence (OV) may be a trigger for them, particularly during maternity. We aimed to analyze the association between manifestations of OV with the development of PPD and PTSD during pregnancy, childbirth, and postpartum. This systematic review was based on the PRISMA 2020 statement and explored original articles published between 2012 and 2022. A total of 21 articles were included in the analysis, and bias was assessed by the Effective Public Health Practice Project's Quality Assessment Tool. The highest rate of PPD symptoms appeared in women under 20 years old, multiparous, and with low education levels. The higher PTSD ratio was present in women under 35 years, primiparous, and with secondary studies. The mode of labor (instrumental or C-section) was identified as a major risk factor of PPD, being mediator variables of the informal coercion of health professionals and dissatisfaction with newborn healthcare. Instead, partner support during labor and high satisfaction with healthcare during birth were protective factors. Regarding PTSD, the mode of labor, several perineal tears, and the Kristeller technique were risk factors, and loss of autonomy and coercion modulated PTSD symptomatology. The protective factors for PTSD were respect for the labor plan, adequate communication with health professionals, social support during labor, and the skin-to-skin procedure. This systematic review provides evidence that OV contributes to PPD and PTSD, being important in developing standardized tools to prevent it. This study recommends changes in maternal healthcare policies, such as individualized healthcare assistance, humanized pregnancy protocols, and women's mental health follow-up, and improvements in the methodological quality of future research.

Vascular nitrosative stress in hypertension induced by fetal undernutrition in rats.

Fetal undernutrition predisposes to hypertension development. Since nitric oxide (NO) is a key factor in blood pressure control, we aimed to investigate the role of NO alterations in hypertension induced by fetal undernutrition in rats. Male and female offspring from dams exposed to undernutrition during the second half of gestation (MUN) were studied at 21 days (normotensive) and 6 months of age (hypertension developed only in males). In aorta, we analyzed total and phosphorylated endothelial NO synthase (eNOS, p-eNOS), 3-nitrotyrosine (3-NT), and Nrf2 (Western blot). In plasma we assessed L-arginine, asymmetric and symmetric dimethylarginine (ADMA, SDMA; LC-MS/MS), nitrates (NOx, Griess reaction), carbonyl groups, and lipid peroxidation (spectrophotometry). In iliac arteries, we studied superoxide anion production (DHE staining, confocal microscopy) and vasodilatation to acetylcholine (isometric tension). Twenty-one-day-old MUN offspring did not show alterations in vascular e-NOS or 3NT expression, plasma L-Arg/ADMA ratio, or NOx. Compared to control group, 6-month-old MUN rats showed increased aortic expression of p-eNOS/eNOS and 3-NT, being Nrf2 expression lower, elevated plasma L-arginine/ADMA, NOx and carbonyl levels, increased iliac artery DHE staining and reduced acetylcholine-mediated relaxations. These alterations in MUN rats were sex-dependent, affecting males. However, females showed some signs of endothelial dysfunction. We conclude that increased NO production in the context of a pro-oxidative environment, leads to vascular nitrosative damage and dysfunction, which can participate in hypertension development in MUN males. Females show a better adaptation, but signs of endothelial dysfunction, which can explain hypertension in ageing.

Online pilot intervention to improve nutritional and lifestyle in Spanish breast-feeding women.

INTRODUCTION: breastfeeding women often cannot adequately follow dietary and healthy habits recommendations. In addition, after labor, their care is usually focused on the newborn and the mother´s health may be neglected. The Mediterranean Diet is the standard of healthy eating, with the Healthy Food Pyramid (HFP) being its graphical representation. OBJECTIVE: the aim of this study was to determine whether a nutritional and lifestyle online intervention may improve HFP adherence in breastfeeding women. METHODS: a total of 181 breastfeeding women in the first sixth months postpartum were enrolled in a non-randomized interventional pilot study. These women answered three questionnaires: sociodemographic and for adherence to Healthy Food Pyramid (APQ), before and after an online intervention for 13 weeks, providing information about nutrition and healthy habits. The APQ ranges from 0 to 10, with higher scores corresponding to greater adherence to HFP. The intervention consisted of lifestyle and nutritional information, which was provided through e-mail. RESULTS: a low adherence to HFP was found, which was improved after 13 weeks of intervention. The online intervention significantly increased the scores for physical activity, grain, seeds and legumes consumption, olive oil use, dairy products, and ani-mal proteins, as well as the HFP adherence global score. The adjusted models showed that the online intervention was associated with adherence to the HFP and physical activity. CONCLUSION: we conclude that an online intervention for breastfeeding women had an impact on their lifestyle, improving nutritional and healthy habits, and can be a useful tool to monitor their health status. Given the importance of this stage for women and their newborns, this is an aspect postnatal healthcare professionals should consider.

Evaluation of Parameters Which Influence Voluntary Ingestion of Supplements in Rats.

Drug safety and efficacy studies frequently use oral gavage, but repetitive usage may cause problems. Administration through voluntary ingestion represents an opportunity for refinement. We aimed to develop a protocol for voluntary ingestion of gelatin-based supplements in rats, assessing the influence of age, sex, fasting (4 h), and additives (vanilla, VF; sucralose, S), and to test it in lactating dams. Three-week-old and 5-month-old Sprague-Dawley rats were placed individually in an empty cage containing a gelatin cube and trained daily (5 days/week), recording the day the whole cube was consumed (latency). Rats trained prior to gestation were offered a gelatin containing 250 mg/kg cocoa shell extract (CSE) during lactation. Rats that did not eat the cube after 8 training days were considered non-habituated, with a proportion similar in young males (7.1%), young females (11.1%), and adult females (10.3%), but significantly higher in adult males (39.3%). Excluding non-habituated rats, latency was 2-3 days, without differences between young and adult rats (p = 0.657) or between males and females (p = 0.189). VF or VF + S in the gelatin did not modify latency, while fasting significantly reduced it in females (p = 0.007) but not in males (p = 0.501). During lactation, trained females ate the CSE-gelatin within 1-5 min without litter problems. Conclusions: Acceptance of a gelatin-based supplement is negatively influenced by male sex, facilitated by fasting, and not modified by additives. Training is remembered after 2 months and does not interfere with lactation. Gelatin-based voluntary ingestion is suitable to administer drugs that need to pass through the digestive system, ensuring adequate dosage, and is important to detect non-habituated rats prior to the study. The current protocol may be implemented by training the rats in their own cage.

Cocoa Shell Extract Reduces Blood Pressure in Aged Hypertensive Rats via the Cardiovascular Upregulation of Endothelial Nitric Oxide Synthase and Nuclear Factor (Erythroid-Derived 2)-like 2 Protein Expression.

Cocoa shell is a by-product of cocoa manufacturing. We obtained an aqueous extract (CSE) rich in polyphenols and methylxanthines with antioxidant and vasodilatory properties. We aimed to evaluate the effects of CSE supplementation in aged hypertensive rats on blood pressure and the mechanism implicated. Eighteen-month-old male and female rats exposed to undernutrition during the fetal period who developed hypertension, with a milder form in females, were used (MUN rats). Systolic blood pressure (SBP; tail-cuff plethysmography) and a blood sample were obtained before (basal) and after CSE supplementation (250 mg/kg; 2 weeks, 5 days/week). Plasma SOD, catalase activity, GSH, carbonyls, and lipid peroxidation were assessed (spectrophotometry). In hearts and aortas from supplemented and non-supplemented age-matched rats, we evaluated the protein expression of SOD-2, catalase, HO-1, UCP-2, total and phosphorylated Nrf2 and e-NOS (Western blot), and aorta media thickness (confocal microscopy). MUN males had higher SBP compared with females, which was reduced via CSE supplementation with a significant difference for group, sex, and interaction effect. After supplementation with plasma, GSH, but not catalase or SOD, was elevated in males and females. Compared with non-supplemented rats, CSE-supplemented males and females exhibited increased aorta e-NOS and Nrf2 protein expression and cardiac phosphorylated-Nrf2, without changes in SOD-2, catalase, HO-1, or UCP-2 in cardiovascular tissues or aorta remodeling. In conclusion, CSE supplementation induces antihypertensive actions related to the upregulation of e-NOS and Nrf2 expression and GSH elevation and a possible direct antioxidant effect of CSE bioactive components. Two weeks of supplementation may be insufficient to increase antioxidant enzyme expression.

Association of maternal body composition and diet on breast milk hormones and neonatal growth during the first month of lactation.

Introduction: Preterm birth is associated with altered growth patterns and an increased risk of cardiometabolic diseases, with breast milk (BM) being a counteracting factor. Preterm infants also show alterations in adipokines and gut hormones influencing appetite and metabolism. Since these hormones are present in BM, it is possible that their levels may equilibrate deficiencies improving infant growth. We aimed to assess 1) the BM levels of ghrelin, resistin, leptin, insulin, peptide YY, and the gastrointestinal peptide in women with preterm and term labor; 2) the relationship between BM hormones and neonatal growth; and 3) the influence of maternal body composition and diet on these BM hormones. Methods: BM from 48 women (30 term and 18 preterm labor) was collected at days 7, 14, and 28 of lactation. Maternal body composition was evaluated by bioimpedance, and neonate anthropometric parameters were collected from medical records. The maternal dietary pattern was assessed by a 72-h dietary recall at days 7 and 28 of lactation. BM hormones were analyzed by the U-Plex Ultra-sensitive method. Data were analyzed using linear regression models. BM from women with preterm labor had lower ghrelin levels, with the other hormones being significantly higher compared to women with term delivery. Results: In premature infants, growth was positively associated with BM ghrelin, while, in term infants, it was positively associated with insulin and negatively with peptide YY. In the first week of lactation, women with preterm labor had higher body fat compared to women with term labor. In this group, ghrelin levels were positively associated with maternal body fat and with fiber and protein intake. In women with term labor, no associations between anthropometric parameters and BM hormones were found, and fiber intake was negatively associated with peptide YY. Discussion: Preterm labor is a factor influencing the levels of BM adipokines and gut hormones, with BM ghrelin being a relevant hormone for premature infant growth. Since ghrelin is lower in BM from women with preterm labor and the levels are associated with maternal fat storage and some dietary components, our data support the importance to monitor diet and body composition in women who gave birth prematurely to improve the BM hormonal status.

Effect of Supplementation with Coffee and Cocoa By-Products to Ameliorate Metabolic Syndrome Alterations Induced by High-Fat Diet in Female Mice.

Coffee and cocoa manufacturing produces large amounts of waste. Generated by-products contain bioactive compounds with antioxidant and anti-inflammatory properties, suitable for treating metabolic syndrome (MetS). We aimed to compare the efficacy of aqueous extracts and flours from coffee pulp (CfPulp-E, CfPulp-F) and cocoa shell (CcShell-E, CcShell-F) to ameliorate MetS alterations induced by a high-fat diet (HFD). Bioactive component content was assessed by HPLC/MS. C57BL/6 female mice were fed for 6 weeks with HFD followed by 6 weeks with HFD plus supplementation with one of the ingredients (500 mg/kg/day, 5 days/week), and compared to non-supplemented HFD and Control group fed with regular chow. Body weight, adipocyte size and browning (Mitotracker, confocal microscopy), plasma glycemia (basal, glucose tolerance test-area under the curve, GTT-AUC), lipid profile, and leptin were compared between groups. Cocoa shell ingredients had mainly caffeine, theobromine, protocatechuic acid, and flavan-3-ols. Coffee pulp showed a high content in caffeine, protocatechuic, and chlorogenic acids. Compared to Control mice, HFD group showed alterations in all parameters. Compared to HFD, CcShell-F significantly reduced adipocyte size, increased browning and high-density lipoprotein cholesterol (HDL), and normalized basal glycemia, while CcShell-E only increased HDL. Both coffee pulp ingredients normalized adipocyte size, basal glycemia, and GTT-AUC. Additionally, CfPulp-E improved hyperleptinemia, reduced triglycerides, and slowed weight gain, and CfPulp-F increased HDL. In conclusion, coffee pulp ingredients showed a better efficacy against MetS, likely due to the synergic effect of caffeine, protocatechuic, and chlorogenic acids. Since coffee pulp is already approved as a food ingredient, this by-product could be used in humans to treat obesity-related MetS alterations.