Fetal Growth Restriction Rat Model Research Articles

Effects of Fetal Growth Restriction on Postnatal Gut Microbiota in a Rat Model.

Fetal growth restriction (FGR) indicates increased risks of lifestyle-related diseases in adulthood. Previous studies showed the association between human gut dysbiosis and various diseases. However, reports examining the relationship between FGR and gut microbiota are scarce. Herein, we hypothesized that FGR may cause gut dysbiosis and analyzed the gut microbiota in a FGR rat model by restricting maternal protein intake during pregnancy. The FGR group was developed by feeding pregnant Sprague Dawley rats a diet containing 7% protein until birth. Control rats were fed 21% protein. Fecal samples of 2-11-week-old pups were collected weekly. DNA was extracted from each sample and subjected to polymerase chain reaction (PCR) amplification and sequencing. Additionally, short-chain fatty acids in the cecum were analyzed at 2 weeks of age, when there were differences in the occupancy of the gut microbiota. Comparative analysis of the gut microbiota showed differences only at 2 weeks of age. Verrucomicrobia was significantly more abundant in the control group ( q < 0.1), whereas pathogenic bacteria, including Enterococcus and Enterobacteriaceae , tended to increase in the FGR group. The abundance of acetic and butyric acid-producing bacteria also differed between groups. Acetic acid in the cecum was considerably decreased in the FGR group, while butyric acid was increased compared to that in the control group. Normalizing the alteration of FGR on postnatal gut microbiota may have beneficial effects for the host, since the FGR group caused gut dysbiosis.

Fetal growth restriction followed by early catch-up growth impairs pancreatic islet morphology in male rats

Fetal growth restriction (FGR), followed by postnatal early catch-up growth, is associated with an increased risk of metabolic dysfunction, including type 2 diabetes in humans. This study aims to determine the effects of FGR and early catch-up growth after birth on the pathogenesis of type 2 diabetes, with particular attention to glucose tolerance, pancreatic islet morphology, and fibrosis, and to elucidate its mechanism using proteomics analysis. The FGR rat model was made by inducing mild intrauterine hypoperfusion using ameroid constrictors (ACs). On day 17 of pregnancy, ACs were affixed to the uterine and ovarian arteries bilaterally, causing a 20.9% reduction in birth weight compared to sham pups. On postnatal day 4 (P4), the pups were assigned to either the good nutrition (GN) groups with 5 pups per dam to ensure postnatal catch-up growth or poor nutrition groups with 15 pups per dam to maintain lower body weight. After weaning, all pups were fed regular chow food ad libitum (P21). Rats in both FGR groups developed glucose intolerance; however, male rats in the FGR good nutrition (FGR-GN) group also developed hypertriglyceridemia and dysmorphic pancreatic islets with fibrosis. A comprehensive and functional analysis of proteins expressed in the pancreas showed that FGR, followed by early catch-up growth, severely aggravated cell adhesion-related protein expression in male offspring. Thus, FGR and early catch-up growth caused pancreatic islet morphological abnormalities and fibrosis associated with the disturbance of cell adhesion-related protein expressions. These changes likely induce glucose intolerance and dyslipidemia in male rats.

Skeletal Muscle Insulin Resistance in a Novel Fetal Growth Restriction Model

The abnormal fetal environment exerts long-term effects on skeletal muscle, and fetal growth restriction (FGR) is associated with insulin resistance in adulthood. In this study, we examined insulin resistance in early adulthood and insulin signaling in skeletal muscle using a novel FGR rat model. Ameroid constrictors (AC) were placed on the bilateral uterine and ovarian arteries of rats on day 17 of gestation; placebo surgery was performed on the control group. We measured body weight at birth, 4, 8, and 12 weeks of age and performed oral glucose tolerance tests at 8 and 12 weeks. Rats were dissected at 12 weeks of age. We examined the mRNA and protein expression of insulin signaling pathway molecules in skeletal muscle. FGR rats had a significantly lower birth weight than control rats (p = 0.002). At 12 weeks of age, the incremental area under the curve of blood glucose was significantly higher, and GLUT4 mRNA and protein expression in soleus muscle was significantly lower in the FGR group than in the control group. Reduced placental blood flow in the AC-attached FGR group caused insulin resistance and altered insulin signaling in skeletal muscles. Therefore, FGR causes skeletal muscle insulin resistance in early adulthood.

Dysregulated ion channels and transporters activate endoplasmic reticulum stress in rat kidney of fetal growth restriction

AimsThe mechanisms underlying the fetal origin of renal disease remains unknown. This study aimed to investigate the profiles of ion channel and transporter proteins in the fetal kidney in fetal growth restriction (FGR)rats, and to explore their association with the fetal origin of renal disease. Main methodsAn FGR rat model was developed by administration of a low-protein diet. Then 367 differentially expressed proteins (DEPs) from quantitative proteome analysis were subjected to Ingenuity Pathway Analysis. 22 DEPs associated with ion channels/transporters were evaluated in the fetal kidney. Na+/H+ exchanger1(NHE1) and its downstream unfolded protein response (UPR) pathway were investigated. Furthermore, overexpression of NHE1 were achieved via plasmid transfection to evaluate the potential influence on the UPR pathway and cell apoptosis in human proximal tubular epithelial cell line HK2 cells. Key findingsFindings were as follows: 1) In the FGR fetal kidney, aquaporin 2/4, solute carrier (SLC) 8a1, 33a1, etc. were downregulated, whereas other transporters including SLC 2a1, 4a1, 9a1, 29a3, etc. were upregulated. 2) NHE1 mRNA levels were markedly elevated in the FGR fetus. Further investigation revealed an increase in the UPR pathway regulators. 3) In vitro study showed that NHE1 overexpression in HK2 cells significantly induced expression of the endoplasmic reticulum stress (ERS) regulators and led to a decrease in the anti-apoptotic potential. SignificanceWe speculate that maternal protein malnutrition causes dysregulation of ion channels/transporters in the fetal kidney. Upregulated NHE1 may activate the UPR pathway and induce cell apoptosis thus leading to impairment of kidney function.

Establishment of a Novel Fetal Growth Restriction Model and Development of a Stem-Cell Therapy Using Umbilical Cord-Derived Mesenchymal Stromal Cells.

Fetal growth restriction (FGR) is a major complication of prenatal ischemic/hypoxic exposure and affects 5%–10% of pregnancies. It causes various disorders, including neurodevelopmental disabilities due to chronic hypoxia, circulatory failure, and malnutrition via the placenta, and there is no established treatment. Therefore, the development of treatments is an urgent task. We aimed to develop a new FGR rat model with a gradual restrictive load of uterus/placental blood flow and to evaluate the treatment effect of the administration of umbilical cord-derived mesenchymal stromal cells (UC-MSCs). To create the FGR rat model, we used ameroid constrictors that had titanium on the outer wall and were composed of C-shaped casein with a notch and center hole inside that gradually narrowed upon absorbing water. The ameroid constrictors were attached to bilateral ovarian/uterine arteries on the 17th day of pregnancy to induce chronic mild ischemia, which led to FGR with over 20% bodyweight reduction. After the intravenous administration of 1 × 105 UC-MSCs, we confirmed a significant improvement in the UC-MSC group in a negative geotaxis test at 1 week after birth and a rotarod treadmill test at 5 months old. In the immunobiological evaluation, the total number of neurons counted via the stereological counting method was significantly higher in the UC-MSC group than in the vehicle-treated group. These results indicate that the UC-MSCs exerted a treatment effect for neurological impairment in the FGR rats.

Prenatal Sildenafil Therapy Improves Cardiovascular Function in Fetal Growth Restricted Offspring of Dahl Salt-Sensitive Rats.

Fetal growth restriction (FGR) is associated with increased risk for cardiovascular and renal disorders in later life. Prenatal sildenafil improves birth weight in FGR animal models. Whether sildenafil treatment protects against long-term cardiovascular and renal disease in these offspring is unknown. The aim of this study is to test the hypothesis that prenatal sildenafil ameliorates cardiovascular and renal function in FGR offspring of Dahl salt-sensitive rats. Sildenafil citrate (60 mg/kg per day) or control gel diet (containing 0.3% salt) was administered from gestational day ten until birth. In male and female offspring, the mean arterial pressure was measured by telemetry in 1 subset from week 5 until week twenty. Echocardiographic parameters, glomerular filtration rate, and fractional electrolyte excretion were determined in another subset at week 9. Aortic and mesenteric artery rings were prepared to assess endothelial-dependent (acetylcholine) and -independent (sodium nitroprusside) vasorelaxation (week 10). The rise in mean arterial pressure per week was attenuated in treated versus untreated male offspring. Mesenteric arteries showed an increased endothelium-dependent relaxation and improved endothelium-independent relaxation in treated versus control male offspring. No differences in aortic relaxation, echocardiographic parameters or renal function were observed between groups. Prenatal sildenafil treatment subtly improves cardiovascular but not renal function in the offspring of this FGR rat model. Translationally, in utero treatment could be beneficial for cardiovascular programming in a sex-specific manner; however, caution is warranted since recent human trials have been halted because of potentially deleterious neonatal side effects when treating pregnancies complicated with severe FGR with sildenafil.

Trichloroethylene exposure in mid-pregnancy decreased fetal weight and increased placental markers of oxidative stress in rats

Although epidemiology studies have associated maternal trichloroethylene (TCE) exposure with decreased birth weight and preterm birth, mechanistic explanations for these associations are currently lacking. We hypothesized that TCE targets the placenta with adverse consequences for pregnancy outcomes. Pregnant Wistar rats were exposed orally to vehicle or 480 mg TCE/kg body weight from gestational days (gd) 6–16, and tissues were collected on gd 16. Exposure to TCE significantly decreased average fetal weight without reducing maternal weight. In placenta, TCE significantly increased 8-hydroxy-deoxyguanosine, global 5-hydroxymethylcytosine, and mRNA expression of Tet3, which codes for an enzyme involved in 5-hydroxymethylcytosine formation. Furthermore, glutathione S-transferase activity and immunohistochemical staining were increased in placentas of TCE-exposed rats. The present study provides the first evidence that TCE increases markers of oxidative stress in placenta in a fetal growth restriction rat model, providing new insight into the placenta as a potentially relevant target for TCE-induced adverse pregnancy outcomes.

Akt Activation and Inhibition of Cytochrome C Release: Mechanistic Insights into Leptin-promoted Survival of Type II Alveolar Epithelial Cells.

Fetal growth restriction (FGR) increases the risk of perinatal death, partly due to defects in lung development. Leptin, a polypeptide hormone, is involved in fetal lung development. We previously demonstrated that treatment with exogenous leptin during gestation significantly promotes fetal lung maturity in the rat model of FGR. In this study, to delineate the molecular pathways through which leptin may enhance fetal lung development, we investigated the impact of leptin treatment on the survival of type II alveolar epithelial cells (AECs), essential leptin-responsive cells involved in lung development, in a rat model of FGR. The rat model of FGR was induced in pregnant Sprague-Dawley rats by partial uterine artery and vein ligation. In vivo and in vitro analyses of fetal lung tissues and freshly-isolated cultured AECs, respectively, showed that leptin protects type II AECs from hypoxia-induced apoptosis. Further molecular studies revealed the role of Akt activation in the leptin-mediated promotion of survival of type II AECs. The data also showed that the anti-apoptotic effects of leptin are dependent on phosphoinositol 3-kinase (PI3K) activation, and involve the down-regulation of caspases 3 and 9, upregulation of pro-survival proteins Bcl-2, and p-Bad, and inhibition of the release of cytochrome c from mitochondria. Taken together, our data suggested that leptin enhances the maturity of fetal lungs by mediating the regulation of caspase-3 and -9 during hypoxia-induced apoptosis of type II AECs and provide support for the potential of leptin as a therapeutic agent for promoting lung development in FGR.

Expression of lung surfactant proteins SP-B and SP-C and their modulating factors in fetal lung of FGR rats.

This study investigated the expression of lung surfactant proteins SP-B and SP-C, and their modulating factors TTF-1 and PLAGL2 in the fetal lung of rats with fetal growth restriction (FGR). The rat FGR model was established by prenatal hypoxia in the first stage of pregnancy, 180 rats for experiment served as hypoxia group, and 197 healthy rats served as normal control group. The FGR incidence in hypoxia was compared with that in normal control group. The histological changes in the fetal lung were observed under the light microscope and electronic microscope in two groups. The SP-B, SP-C, TTF-1 and PLAGL2 proteins were determined in the fetal lung of two groups immunohistochemically. The expression levels of SP-B, SP-C, TTF-1 and PLAGL2 protein and mRNA in the fetal lung of two groups were detected by using Western blotting and RT-PCR respectively. The FGR rat model was successfully established by using hypoxia. Pathologically the fetal lung developed slowly, and the expression levels of SP-B, SP-C, TTF-1 and PLAGL2 protein and mRNA in the fetal lung were significantly reduced in hypoxia group as compared with those in normal control group. It was suggested that maternal hypoxia in the first stage of pregnancy could induce FGR, and reduce the expression of SP-B and SP-C, resulting in the disorder of fetal lung development and maturation.

Fetal growth restriction rat model induced by low protein diet during pregnancy

Objective To explore the impacts of 75％ low-protein diet intake during gestation on fetal growth restriction (FGR) rat model establishment.Methods Thirty-eight pregnant Sprague-Dawley rats were included into the study.At first,five pregnant rats were fed with sufficient normal diet with protein content of 22％.Their daily food consumption was recorded and taken as the basis to determine daily feed consumption of 75％ low-protein group (protein content 9.2％).In order to ensure that each group finally had at least ten pregnant rats to deliver,there were 11 rats assigned to the control group (pregnant rats fed with sufficient normal diet,protein content was 22％),13 to the low-protein group (pregnant rats fed with low protein diet,protein content was 9.2％,but the food consumption was the same as control group) and 14 to the 75％ lowprotein group (pregnant rats fed with low-protein diet,protein content was 9.2％,the food consumption was 75％ of the control group).All female rats were fed with sufficient normal diet after delivery.The body weight,overall weight gain during gestation,the mortality rate and the non-delivery rate of pregnant rats were compared.The third day's newborn weight after birth,FGR incidence and the mortality rate within three days after birth of newborns were also compared.One way analysis of variance,LSD-t test,independent sample t-test and Chisquare test were used as statistical methods.Results (1) The body weight of pregnant rats:There was no significant difference in body weight among the three groups at gestational day 0,3 and 6.On day 9,body weight of 75％ low-protein group [(271.9±8.4) g] and low-protein group [(274.1 ±7.8) g] were significantly lower than that of the control group [(287.2± 18.7) g] (t=2.514 and 2.170,both P＜0.05),but there was no significant difference between the former two groups.On Day 12,body weight of 75％ low-protein group [(275.7 ± 10.7) g] and low protein group [(285.1 ± 12.5) g] were significantly lower than that of the control group [(306.4±29.7) g] (t=3.262 and 2.218,both P＜0.05),and the difference between the former two groups was also statistically significant (t=2.098,P＜0.05).Before delivery,body weight of 75％ low-protein group,low protein group and control group were (300.4±14.1) g,(317.0±16.3) g and (372.9±19.1) g,respectively with statisticall significance (F=64.219,P＜0.05).The overall weight gain during pregnancy for 75％low-protein group,low-protein group and control group was (61.6± 19.8) g,(81.8±21.6) g and (139.3± 12.0) g,respectively.The difference among the three groups was statistically significant (F=55.863,P＜0.05).(2) The mortality rates of pregnant rats for 75％ low-protein group,low-protein group and control group were 3/14,2/13 and 1/11 respectively without significant difference (P＞0.05).Neither was the non-delivery rate within 30 days (embryonic resorption) for the three groups (1/14,1/13,0/11,P＞0.05).(3) The numbers of pups were 101 in 75％ low-protein group,104 in low-protein group and 107 in control group.The newborn mortality rate within three days after birth was 28.7％ (29/101) in 75％ tow-protein group and 23.0％ (24/104)in low-protein group,with were significantly higher than that of the control group (7.5％,8/107) (x2=16.022and 9.976,both P＜0.05),but there was no significant difference between groups.The third day's newborn weight after birth for 75％ low-protein group,low-protein group and control group were (6.3 ±0.8) g,(6.9±0.9) g and (8.1 ±0.9) g,the difference was statistically significant (F=90.602,P＜0.05).FGR incidence for 75％ low-protein group was 55.6％ (40/72),which was significantly higher than that of the low-protein group (28.8％,23/80) and the control group (5.0％,5/99) (x2=11.220,54.834 and 18.833 all P＜0.05).Conclusion 75％ low-protein diet feeding during pregnancy is an ideal method to induce FGR rat model with high FGR incidence,whereas and low mortality rates of pregnant rats,the fetuses and newborns. Key words: Diet, protein-restricted; Fetal growth retardation; Disease models, animal

Fetal growth restriction alters hippocampal 17-beta estradiol and estrogen receptor alpha levels in the newborn male rat

Fetal growth restriction (FGR) is associated with impaired neurodevelopmental outcomes in affected newborns. The pathogenesis of FGR-associated neurodevelopmental impairment implicates abnormal hippocampal function. The steroid hormone estrogen and its receptor, estrogen receptor alpha (ERα), are involved in the normal programming of hippocampal development and structure. However, the impact of FGR on hippocampal estrogen and hippocampal ERα is not well characterized. We hypothesized that FGR will reduce hippocampal and serum levels of 17-beta estradiol and its receptor, ERα, in the newborn rat hippocampus. We further hypothesize that FGR will reduce hippocampal ERα levels in a region-specific manner. To test our hypotheses, we used the well characterized rat model of FGR induced by uteroplacental-insufficiency in the pregnant Sprague-Dawley rat. Hippocampi and serum were obtained from FGR and control day 0 rat pups and examined for hippocampal 17-beta estradiol, serum 17-beta estradiol, and ERα mRNA and protein levels. Immunohistochemistry was performed to examine region-specific ERα staining. FGR decreased hippocampal 17-beta estradiol levels in the hippocampi of male newborn rats but not females. Serum 17-beta estradiol levels were not affected by FGR in either gender. FGR decreased hippocampal ERα mRNA levels in males but not females. Hippocampal ERα protein levels by Western blotting were not affected by FGR. However, FGR decreased apparent ERα staining in the cornu ammonis (CA)1, CA3, and dentate gyrus regions in the hippocampi of male newborn rats but not females. We conclude that FGR affects the programming of hippocampal estrogen and hippocampal ERα levels in the newborn rat in a gender-specific manner.

Impairments of spatial learning and memory in rat offspring with fetal growth restriction

Objective Throughout the world, fetal growth restriction(FGR) is one of the most severe complications occurring during pregnancy. It is subsequently associated with neurologic abnormalities in chldren. Our aim was to investigate the spatial learning and memory ability of rat offspring born with FGR. Methods A rat model of FGR was constructed using the method of passive smoking. Spatial learning and memory were studied in rat offspring born with FGR by assessing the animals' performance using the Morris water maze task. Results At 1- and 2-months of age, both female and male offspring rats showed impairment of performance, while at 4 months of age, only female rats showed impaired performance. The FGR offspring spent a longer time swimming and used inefficient strategies( P < 0.05, respectively). However, there were no significant maze performance FGR effects in the 4 month old male rats. In all groups of FGR offspring, irrespective of age or sex, the time spent in the platform quadrant by the rat was significantly less than that in the control group( P < 0.05). Conclusion The Morris water maze performance decreased in rat offspring born with FGR. It is suggested that FGR can cause impairments of spatial learning and memory in young animals.

Effect of behavior training on learning, memory and the expression of NR2B, GluR1 in hippocampus of rats offspring with fetal growth restriction

Objective To study effects of behavior training on learning, memory and the expression of NR2B, GluR1 in hippocampus of rat' s offspring with fetal growth restriction(FGR). Methods The rat model of FGR was established by passive smoking method. The rats offspring were divided into the FGR group and the control group, then randomly divided into the trained and untrained group, respectively. Morris water maze test was proceeded on postnatal month(PM2/4) as a behavior training method, then the learning-memory of rats was detected through dark-avoidance and step-down tests. The expressions of NR2B and GluR1 subunits in hippocampal CA1 and CA3 areas were detected by immunohistochemical method. Results In the dark-avoidance and step-down tests, the performance record of rats with FGR was worse than that of control rats, and the behavior-trained rats was better than the untrained rats, when the FGR model and training factors were analyzed singly. The model factor and training factor had significant interaction( P < 0.05). The expressions of NR2B and GluR1 subunits in hippocampal CA1 and CA3 areas of rats with FGR reduced. In contrast, the expressions of GluR1 and NR2B subunits in CA1 area of behavior-trained rats increased, when the FGR model and training factors were analyzed singly. Conclusion These findings suggested that the effect of behavior training on the expressions of NR2B and GluR1 subunits in CA1 area should be the mechanistic basis for the training-induced improvement in learning-memory abilities.