Strategies for intra-amniotic administration of fetal therapy in a rabbit model of intrauterine growth restriction.

Intrauterine growth restriction affects up to 10% of all pregnancies, leading to fetal programming with detrimental consequences for lifelong health. However, no therapeutic strategies have so far been effective to ameliorate these consequences. Our previous study has demonstrated that a single dose of nutrients administered into the amniotic cavity, bypassing the often dysfunctional placenta via intra-amniotic administration, improved survival at birth but not birthweight in an intrauterine growth restriction rabbit model. The aim of this study was to further develop an effective strategy for intra-amniotic fetal therapy in an animal model. Intrauterine growth restriction was induced by selective ligation of uteroplacental vessels on one uterine horn of pregnant rabbits at gestational day 25, and fetuses were delivered by cesarean section on GD30. During the five days of intrauterine growth restriction development, three different methods of intra-amniotic administration were used: continuous intra-amniotic infusion by osmotic pump, multiple intra-amniotic injections, and single fetal intraperitoneal injection. Technical feasibility, capability to systematically reach the fetus, and survival and birthweight of the derived offspring were evaluated for each technique. Continuous intra-amniotic infusion by osmotic pump was not feasible owing to the high occurrence of catheter displacement and amnion rupture, while methods using two intra-amniotic injections and one fetal intraperitoneal injection were technically feasible but compromised fetal survival. Taking into account all the numerous factors affecting intra-amniotic fetal therapy in the intrauterine growth restriction rabbit model, we conclude that an optimal therapeutic strategy with low technical failure and positive fetal impact on both survival and birthweight still needs to be found.

A randomized controlled trial investigating the impact of maternal dietary supplementation with pomegranate juice on brain injury in infants with IUGR.

Animal studies have demonstrated the therapeutic potential of polyphenol-rich pomegranate juice. We recently reported altered white matter microstructure and functional connectivity in the infant brain following in utero pomegranate juice exposure in pregnancies with intrauterine growth restriction (IUGR). This double-blind exploratory randomized controlled trial further investigates the impact of maternal pomegranate juice intake on brain structure and injury in a second cohort of IUGR pregnancies diagnosed at 24-34 weeks' gestation. Ninety-nine mothers and their eligible fetuses (n = 103) were recruited from Brigham and Women's Hospital and randomly assigned to 8 oz pomegranate (n = 56) or placebo (n = 47) juice to be consumed daily from enrollment to delivery. A subset of participants underwent fetal echocardiogram after 2 weeks on juice with no evidence of ductal constriction. 57 infants (n = 26 pomegranate, n = 31 placebo) underwent term-equivalent MRI for assessment of brain injury, volumes and white matter diffusion. No significant group differences were found in brain volumes or white matter microstructure; however, infants whose mothers consumed pomegranate juice demonstrated lower risk for brain injury, including any white or cortical grey matter injury compared to placebo. These preliminary findings suggest pomegranate juice may be a safe in utero neuroprotectant in pregnancies with known IUGR warranting continued investigation.Clinical trial registration: NCT04394910, https://clinicaltrials.gov/ct2/show/NCT04394910 , Registered May 20, 2020, initial participant enrollment January 16, 2016.

Dietary supplementation of l-arginine and N-carbamylglutamate enhances duodenal barrier and mitochondrial functions and suppresses duodenal inflammation and mitophagy in suckling lambs suffering from intrauterine-growth-restriction.

The current work aimed at investigating the effects of the dietary supplementation of N-carbamylglutamate (NCG) or l-arginine (Arg) on the duodenal mitophagy, mitochondrial function, inflammation, and barrier function in suckling lambs suffering from intrauterine-growth-retardation (IUGR). Forty-eight neonate Hu lambs were used in this study: 12 lambs with normal birth weight (NBW: 4.25 ± 0.14 kg) and 36 lambs with IUGR (3.01 ± 0.13 kg). Seven day old lambs were assigned to 4 treatment groups (12 lambs in each group) as follows: control group (CON), IUGR group, IUGR + Arg, and IUGR + NCG. Lambs were fed the experimental diets for 21 days from 7 days to 28 days of age. Compared with IUGR lambs, the Arg or NCG-treated IUGR lambs had a markedly higher duodenal transepithelial electrical resistance (TER) and lower fluorescein isothiocyanate dextran (FD4) (P < 0.05), respectively. The duodenal mitochondrial membrane potential change (ΔΨm), relative mitochondrial DNA (mtDNA) content, adenosine triphosphate (ATP) level, together with the activities of the respiratory complexes I, III, and IV were markedly higher in Arg or NCG-treated IUGR lambs than those in non-supplemented IUGR lambs (P < 0.05). The expressions of the integrity-related proteins (occludin and zonula occludens-1 (ZO-1)), antioxidant- and apoptosis-related proteins (B-cell lymphoma/leukaemia 2 (Bcl2), superoxide dismutase 2 (SOD2), catalase (CAT), and glutathione peroxidase 1 (GPx1)), and the nitric oxide-dependent pathway-related proteins (epithelial NO synthase (eNOS) and inducible NO synthase (iNOS)) were higher in NCG or Arg-supplemented IUGR lambs than those in nontreated IUGR lambs (P < 0.05). The duodenal expressions of the mitophagy-related proteins (microtubule-associated protein light chain 3 (LC3) I, LC3 II, Belin1, PTEN induced putative kinase 1 (PINK1), and Parkin) and the immune function-related proteins (myeloid differentiation factor 88 (MyD88), IL-6, nuclear factor kappa B (p65), toll-like receptor (TLR4) and TNF-α) were reduced (P < 0.05) in NCG or Arg-supplemented IUGR lambs compared with non-supplemented IUGR lambs. These results demonstrated that the dietary supplementation of Arg or NCG enhanced the duodenal barrier function and mitochondrial function, mitigated duodenal inflammation, and suppressed mitophagy in suckling lambs suffering from IUGR.

Update on oxidative stress and inflammation in pregnant women, unborn children (nasciturus), and newborns - Nutritional and dietary effects.

The scientific background of perinatal pathology, regarding both mother and offspring, from the lipidomic perspective, has highlighted the possibility of identifying new, promising clinical markers of oxidative stress and inflammation, closely related to the normal development of unborn and newborn children, together with their application. In this regard, in recent years, significant advances have been achieved, assisted by both newly developed analytical tools and basic knowledge on the biological implications of oxylipins. Hence, in the light of this recent progress, this review aims to provide an update on the relevance of human oxylipins during pregnancy and in the unborn and newborn child, covering two fundamental aspects. Firstly, the evidence from human clinical studies and dietary intervention trials will be used to shed light on the extent to which dietary supplementation can modulate the lipidomic markers of oxidative stress and inflammation in the perinatal state, emphasizing the role of the placenta and metabolic disturbances in the mother and fetus. The second part of this article comprises a review of existing data on specific pathophysiological aspects of human reproduction, in relation to lipidomic markers in pregnant women, unborn children, and newborn children. The information reviewed here evidences the current opportunity to correct reproductive disturbances, in the framework of lipidomics, by fine-tuning dietary interventions.

l-Threonine improves intestinal mucin synthesis and immune function of intrauterine growth-retarded weanling piglets.

OBJECTIVES: The aim of this study was to investigate the effects of dietary l-threonine supplementation on the growth performance, intestinal immune function, mucin synthesis, and goblet cell differentiation in weanling piglets with intrauterine growth retardation (IUGR). METHODS: Eighteen litters of newborn piglets were selected at birth, with one normal birthweight (NBW) and two IUGR piglets in each litter. At weaning, the NBW piglet and one of the IUGR piglets were assigned to groups fed a basal diet (i.e., the NBW-CON and IUGR-CON groups). The other IUGR piglet was assigned to a group fed the basal diet supplemented with 2 g l-threonine per kg of diet (i.e., IUGR-Thr group). Therefore, all piglets were distributed across three groups for a 3-wk feeding trial. RESULTS: Compared with NBW, IUGR decreased growth performance, increased ileal proinflammatory cytokine levels, and reduced ileal mucin 2 (Muc2) content and goblet cell density of weanling piglets. Supplementation of l-threonine increased the feed efficiency of the IUGR-Thr group compared with the IUGR-CON group. The l-threonine-supplemented diet attenuated ileal inflammatory responses of the IUGR-Thr piglets and increased production of Muc2 and secretory immunoglobulin A and density of goblet cells. In addition, L-threonine supplementation downregulated δ-like 1 and hes family bHLH transcription factor 1, whereas growth factor independence 1 and Kruppel-like factor 4 expression levels were upregulated. CONCLUSION: Dietary l-threonine supplementation attenuates inflammatory responses, facilitates Muc2 synthesis, and promotes goblet cell differentiation in the ileum of IUGR piglets.

Pre-birth origins of allergy and asthma.

Allergy is a chronic disease that can develop as early as infancy, suggesting that early life factors are important in its aetiology. Variable associations between size at birth, a crude marker of the fetal environment, and allergy have been reported in humans and require comprehensive review. Associations between birth weight and allergy are however confounded in humans, and we and others have therefore begun exploring the effects of early life events on allergy in experimental models. In particular, we are using ovine models to investigate whether and how a restricted environment before birth protects against allergy, whether methyl donor availability contributes to allergic protection in IUGR, and why maternal asthma during pregnancy is associated with increased risks of allergic disease in children. We found that experimental intrauterine growth restriction (IUGR) in sheep reduced cutaneous responses to antigens in progeny, despite normal or elevated IgE responses. Furthermore, maternal methyl donor supplementation in late pregnancy partially reversed effects of experimental IUGR, consistent with the proposal that epigenetic pathways underlie some but not all effects of IUGR on allergic susceptibility. Ovine experimental allergic asthma with exacerbations reduces relative fetal size in late gestation, with some changes in immune populations in fetal thymus suggestive of increased activation. Maternal allergic asthma in mice also predisposes progeny to allergy development. In conclusion, these findings in experimental models provide direct evidence that a perturbed environment before birth alters immune system development and postnatal function, and provide opportunities to investigate underlying mechanisms and develop and evaluate interventions.

Dietary interventions for fetal growth restriction - therapeutic potential of dietary nitrate supplementation in pregnancy.

Fetal growth restriction (FGR) affects around 5% of pregnancies and is associated with significant short- and long-term adverse outcomes. A number of factors can increase the risk of FGR, one of which is poor maternal diet. In terms of pathology, both clinically and in many experimental models of FGR, impaired uteroplacental vascular function is implicated, leading to a reduction in the delivery of oxygen and nutrients to the developing fetus. Whilst mechanisms underpinning impaired uteroplacental vascular function are not fully understood, interventions aimed at enhancing nitric oxide (NO) bioavailability remain a key area of interest in obstetric research. In addition to endogenous NO production from the amino acid l-arginine, via nitric oxide synthase (NOS) enzymes, research in recent years has established that significant NO can be derived from dietary nitrate, via the 'alternative NO pathway'. Dietary nitrate, abundant in green leafy vegetables and beetroot, can increase NO bioactivity, conferring beneficial effects on cardiovascular function and blood flow. Given the beneficial effects of dietary nitrate supplementation to date in non-pregnant humans and animals, current investigations aim to assess the therapeutic potential of this approach in pregnancy to enhance NO bioactivity, improve uteroplacental vascular function and increase fetal growth.

Coenzyme Q10 prevents hepatic fibrosis, inflammation, and oxidative stress in a male rat model of poor maternal nutrition and accelerated postnatal growth.

BACKGROUND: It is well established that low birth weight and accelerated postnatal growth increase the risk of liver dysfunction in later life. However, molecular mechanisms underlying such developmental programming are not well characterized, and potential intervention strategies are poorly defined. OBJECTIVES: We tested the hypotheses that poor maternal nutrition and accelerated postnatal growth would lead to increased hepatic fibrosis (a pathological marker of liver dysfunction) and that postnatal supplementation with the antioxidant coenzyme Q10 (CoQ10) would prevent this programmed phenotype. DESIGN: A rat model of maternal protein restriction was used to generate low-birth-weight offspring that underwent accelerated postnatal growth (termed "recuperated"). These were compared with control rats. Offspring were weaned onto standard feed pellets with or without dietary CoQ10 (1 mg/kg body weight per day) supplementation. At 12 mo, hepatic fibrosis, indexes of inflammation, oxidative stress, and insulin signaling were measured by histology, Western blot, ELISA, and reverse transcriptase-polymerase chain reaction. RESULTS: Hepatic collagen deposition (diameter of deposit) was greater in recuperated offspring (mean ± SEM: 12 ± 2 µm) than in controls (5 ± 0.5 µm) (P < 0.001). This was associated with greater inflammation (interleukin 6: 38% ± 24% increase; P < 0.05; tumor necrosis factor α: 64% ± 24% increase; P < 0.05), lipid peroxidation (4-hydroxynonenal, measured by ELISA: 0.30 ± 0.02 compared with 0.19 ± 0.05 µg/mL per µg protein; P < 0.05), and hyperinsulinemia (P < 0.05). CoQ10 supplementation increased (P < 0.01) hepatic CoQ10 concentrations and ameliorated liver fibrosis (P < 0.001), inflammation (P < 0.001), some measures of oxidative stress (P < 0.001), and hyperinsulinemia (P < 0.01). CONCLUSIONS: Suboptimal in utero nutrition combined with accelerated postnatal catch-up growth caused more hepatic fibrosis in adulthood, which was associated with higher indexes of oxidative stress and inflammation and hyperinsulinemia. CoQ10 supplementation prevented liver fibrosis accompanied by downregulation of oxidative stress, inflammation, and hyperinsulinemia.

Fish oil improves lipid profile in juvenile rats with intrauterine growth retardation by altering the transcriptional expression of lipid-related hepatic genes.

OBJECTIVE: To determine whether maternal intrauterine undernutrition and post-weaning fish oil intake influence lipid profile in juvenile offspring, and explore the possible mechanisms at transcriptional levels. METHODS: After weaning, 32 control offspring and 24 intrauterine growth retardation (IUGR) offspring were randomly allocated to standard chow or fish oil diet. At 10 weeks, fasting plasma glucose, triglycerides, total cholesterol and expressions of related hepatic genes were examined. RESULTS: IUGR offspring without catch-up growth tended to develop hyperglycemia, dyslipidemia and hepatic steatosis. Down-regulation of CPT-1 and LDLR at transcriptional levels were found in IUGR offspring. Early short-term fish oil intervention reversed these unfavorable changes in juvenile rats with IUGR. The mechanisms might be mediated by decreased expression of ACC-1, increased expression of CPT-1, LDLR and ABCG5. CONCLUSION: These data suggest that IUGR offspring already present lipid abnormality in juvenile stage, and early short-term fish oil consumption is beneficial to prevent these unfavorable changes.

Dietary Tributyrin Supplementation Attenuates Insulin Resistance and Abnormal Lipid Metabolism in Suckling Piglets with Intrauterine Growth Retardation.

Intrauterine growth retardation (IUGR) is associated with insulin resistance and lipid disorder. Tributyrin (TB), a pro-drug of butyrate, can attenuate dysfunctions in body metabolism. In this study, we investigated the effects of TB supplementation on insulin resistance and lipid metabolism in neonatal piglets with IUGR. Eight neonatal piglets with normal birth weight (NBW) and 16 neonatal piglets with IUGR were selected, weaned on the 7th day, and fed basic milk diets (NBW and IUGR groups) or basic milk diets supplemented with 0.1% tributyrin (IT group, IUGR piglets) until day 21 (n = 8). Relative parameters for lipid metabolism and mRNA expression were measured. Piglets with IUGR showed higher (P < 0.05) concentrations of insulin in the serum, higher (P < 0.05) HOMA-IR and total cholesterol, triglycerides (TG), non-esterified fatty acid (NEFA) in the liver, and lower (P < 0.05) enzyme activities (hepatic lipase [HL], lipoprotein lipase [LPL], total lipase [TL]) and concentration of glycogen in the liver than the NBW group. TB supplementation decreased (P < 0.05) the concentrations of insulin, HOMA-IR, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol in the serum, and the concentrations of TG and NEFA in the liver, and increased (P < 0.05) enzyme activities (HL, LPL, and TL) and concentration of glycogen in the liver of the IT group. The mRNA expression for insulin signal transduction pathway and hepatic lipogenic pathway (including transcription factors and nuclear factors) was significantly (P < 0.05) affected in the liver by IUGR, which was efficiently (P < 0.05) attenuated by diets supplemented with TB. TB supplementation has therapeutic potential for attenuating insulin resistance and abnormal lipid metabolism in IUGR piglets by increasing enzyme activities and upregulating mRNA expression, leading to an early improvement in the metabolic efficiency of IUGR piglets.

Medium-chain TAG attenuate hepatic oxidative damage in intra-uterine growth-retarded weanling piglets by improving the metabolic efficiency of the glutathione redox cycle.

The present study investigated the effects of medium-chain TAG (MCT) on hepatic oxidative damage in weanling piglets with intra-uterine growth retardation (IUGR). At weaning (mean 21 (SD 1·06) d of age), twenty-four IUGR piglets and twenty-four normal-birth weight (NBW) piglets were selected according to their birth weight (BW; IUGR: mean 0·95 (SD 0·04) kg; NBW: mean 1·58 (SD 0·04) kg) and weight at the time of weaning (IUGR: mean 5·26 (SD 0·15) kg; NBW: mean 6·98 (SD 0·19) kg) and fed either a soyabean oil (SO) diet (containing 5% SO) or a MCT diet (containing 1% SO and 4% MCT) for 28 d. IUGR piglets exhibited poor (P<0·05) growth performance, lower (P<0·05) metabolic efficiency of hepatic glutathione (GSH) redox cycle, and increased (P<0·05) levels of reactive oxygen species, apoptosis and necrosis in hepatocytes compared with NBW piglets. The MCT diet increased (P<0·05) the average daily gain and feed efficiency of piglets during the first 4 weeks after weaning. Furthermore, MCT diet-fed piglets had a higher (P<0·05) GSH:oxidised glutathione ratio and increased (P<0·05) activities of glucose-6-phosphate dehydrogenase (G6PD) and GSH reductase. The expression of G6PD was up-regulated (P<0·05) by the MCT diet irrespective of BW. Moreover, malondialdehyde concentrations in the liver and apoptosis and necrosis levels in hepatocytes were decreased (P<0·05) by the MCT diet irrespective of BW. These results indicate that MCT might have auxiliary therapeutic potential to attenuate hepatic oxidative damage in IUGR offspring during early life, thus leading to an improvement in the metabolic efficiency of the hepatic GSH redox cycle.

Dietary melatonin supplementation alters uteroplacental amino acid flux during intrauterine growth restriction in ewes.

Dietary melatonin supplementation during mid- to late-gestation increased umbilical artery blood flow and caused disproportionate fetal growth. This melatonin-induced increase in umbilical artery blood flow may alter nutrient availability to the fetus, which may lead to alterations in fetal size. The objectives of the current experiment were to determine amino acid (AA) and glucose concentrations as well as AA and glucose flux across the uteroplacenta using a mid- to late-gestation model of intrauterine growth restriction supplemented with dietary melatonin as a 2 × 2 factorial design. At day 50 of gestation, 32 ewes were supplemented with 5 mg of melatonin (MEL) or no melatonin (CON) and were allocated to receive 100% (adequate; ADQ) or 60% (restricted; RES) of nutrient requirements. On day 130 of gestation, uterine and umbilical blood flows were determined via Doppler ultrasonography during a non-survival surgery. Blood samples were collected under general anesthesia from the maternal saphenous artery, gravid uterine vein, umbilical artery, and umbilical vein for AA analysis and glucose. Total α-AA concentrations in maternal artery and gravid uterine vein were decreased (P < 0.05) in RES v. ADQ fed ewes. Maternal arterial - venous difference in total α-AA was increased (P ⩽ 0.01) in RES v. ADQ fed ewes, while total uterine α-AA flux was not different (P > 0.40) across all treatment groups. Fetal venous - arterial difference in total α-AA as well as uteroplacental flux of total α-AA were decreased (P < 0.05) in CON-RES v. CON-ADQ, and similar (P > 0.20) in MEL-RES v. CON-ADQ. Maternal concentrations and uterine flux of branched-chain AA (BCAA) were not different across all treatment groups; however, fetal uptake of BCAA was decreased (P < 0.05) in CON-RES v. CON-ADQ, and similar (P > 0.20) in MEL-RES v. CON-ADQ. Uterine uptake of glucose was not different (P ⩾ 0.08) across all treatment groups, while uteroplacental uptake of glucose was increased (P ⩽ 0.05) in RES v. ADQ ewes. In conclusion, maternal nutrient restriction increased maternal arterial - venous difference in total α-AA, while total uterine α-AA flux was unaffected by maternal nutrient restriction. Melatonin supplementation did not impact maternal serum concentrations or uterine flux of glucose or AA; however, melatonin did improve fetal BCAA uptake during maternal nutrient restriction.

Nutritional management and growth hormone treatment of preterm infants born small for gestational age.

In severe cases of intrauterine growth retardation, elective preterm delivery may provide the possibility for nutritional intervention to prevent some of the long-term consequences of the catabolic condition in utero. Neonatal nutritional management is aimed at providing a high protein intake of up to 4 g/kg/day in order to obtain the rapid increase in protein that is seen in normally growing infants during the early postnatal period. Unfortunately, due to impaired production of urea, high plasma levels of ammonia, which may be rate limiting with respect to an optimal gain in protein, are often observed in the preterm infant born small for gestational age (SGA). In an attempt to stimulate protein synthesis in preterm infants born SGA, growth hormone (GH) treatment was given to seven such infants during the early postnatal period. The infants received daily subcutaneous injections of GH (1.0 IU/kg/day) from postnatal day 7 until a body weight of 2000 g was reached (postnatal week 7-8). A further seven preterm infants born SGA were studied as controls. GH treatment had no significant effects on growth, body composition, net protein gain and glucose metabolism. Furthermore, plasma levels of insulin-like growth factor I (IGF-1) and IGF-binding protein-3 revealed a normal developmental increase and were not significantly altered by GH treatment. These results may be explained by a relative GH insensitivity or resistance during this period of early preterm life.

[Post-conceptional age. Effect on protein requirements of premature infants]. / Das postkonzeptionelle Alter. Einfluss auf den Proteinbedarf Frühgeborener.

In 12 preterm infants (mean gestational age: 28.9 weeks; mean birthweight: 1230 g) (Group 1) the serum concentrations of urea and alpha-amino-nitrogen were studied during the 32nd and between the 36th and 37th week of post-conceptional age. In 9 preterm infants with a mean gestational age of 34.6 weeks (mean birthweight: 1860 g) (Group 2) the same parameters were estimated during the 36th or 37th week of postconceptional age. In all male infants (group 1: n = 6; group 2: n = 5) the excretion of nitrogen in urine and stools were measured during a 48 h balance period. During the study all infants were fed with human milk enriched with 3 g of a bovine human milk fortifier (Eoprotin, Milupa AG, Germany) per 100 ml human milk (mean protein intake: 3.4 g/kg.day; mean caloric intake: 132.5 kcal/kg.day). Despite the similar protein and energy intakes on all study days the serum concentrations of urea and alpha-amino-nitrogen were significantly higher during the 36th or 37th postconceptional week if compared to those found during the 32nd postconceptional week. This was accompanied by elevated renal excretion of total nitrogen due to an increased excretion of urea in the older infants. The nitrogen excretions in stools were low during both study periods. During the period 36 and 37 weeks of postconceptional age no differences could be found for all parameters studied between the both groups of different gestational ages. The data of the present study suggest that in preterm infants with postconceptional age of 37 weeks or more protein intakes of 3.4 g/kg.day and more may exceed the requirements of these infants. Thus, supplementation of human milk with protein is not necessary after the 37th week of postconceptional age if feeding volumes of 170 ml/kg.day or more are tolerated.