LPD Offspring Research Articles

Arginase upregulation and eNOS uncoupling contribute to impaired endothelium-dependent vasodilation in a rat model of intrauterine growth restriction.

Individuals born after intrauterine growth restriction (IUGR) are at increased risk of developing cardiovascular diseases in adulthood, notably hypertension (HTN). Alterations in the vascular system, particularly impaired endothelium-dependent vasodilation, may play an important role in long-term effects of IUGR. Whether such vascular dysfunction precedes HTN has not been fully established in individuals born after IUGR. Moreover, the intimate mechanisms of altered endothelium-dependent vasodilation remain incompletely elucidated. We therefore investigated, using a rat model of IUGR, whether impaired endothelium-dependent relaxation precedes the development of HTN and whether key components of the l-arginine-nitric oxide (NO) pathway are involved in its pathogenesis. Pregnant rats were fed with a control (CTRL, 23% casein) or low-protein diet (LPD, 9% casein) to induce IUGR. Systolic blood pressure (SBP) was measured by tail-cuff plethysmography in 5- and 8-wk-old male offspring. Aortic rings were isolated to investigate relaxation to acetylcholine, NO production, endothelial NO synthase (eNOS) protein content, arginase activity, and superoxide anion production. SBP was not different at 5 wk but significantly increased in 8-wk-old offspring of maternal LPD (LP) versus CTRL offspring. In 5-wk-old LP versus CTRL males, endothelium-dependent vasorelaxation was significantly impaired but restored by preincubation with l-arginine or the arginase inhibitor S-(2-boronoethyl)-l-cysteine; NO production was significantly reduced but restored by l-arginine pretreatment; total eNOS protein, dimer-to-monomer ratio, and arginase activity were significantly increased; superoxide anion production was significantly enhanced but normalized by pretreatment with the NO synthase inhibitor Nω-nitro-l-arginine. In this model, IUGR leads to early-impaired endothelium-dependent vasorelaxation, resulting from arginase upregulation and eNOS uncoupling, which precedes the development of HTN.

Accelerated age-related decline in renal and vascular function in female rats following early-life growth restriction.

Many studies report sexual dimorphism in the fetal programming of adult disease. We hypothesized that there would be differences in the age-related decline in renal function between male and female intrauterine growth-restricted rats. Early-life growth restriction was induced in rat offspring by administering a low-protein diet (LPD; 8.7% casein) to dams during pregnancy and lactation. Control dams were fed a normal-protein diet (NPD; 20% casein). Mean arterial pressure (MAP) and renal structure and function were assessed in 32- and 100-wk-old offspring. Mesenteric artery function was examined at 100 wk using myography. At 3 days of age, body weight was ∼24% lower (P < 0.0001) in LPD offspring; this difference was still apparent at 32 wk but not at 100 wk of age. MAP was not different between the male NPD and LPD groups at either age. However, MAP was greater in LPD females compared with NPD females at 100 wk of age (∼10 mmHg; P < 0.001). Glomerular filtration rate declined with age in the NPD male, LPD male and LPD female offspring (∼45%, all P < 0.05), but not in NPD female offspring. Mesenteric arteries in the aged LPD females had reduced sensitivity to nitric oxide donors compared with their NPD counterparts, suggesting that vascular dysfunction may contribute to the increased risk of disease in aged females. In conclusion, females growth-restricted in early life were no longer protected from an age-related decline in renal and arterial function, and this was associated with increased arterial pressure without evidence of renal structural damage.

When early life growth restriction in rats is followed by attenuated postnatal growth: effects on cardiac function in adulthood.

Epidemiological and experimental studies demonstrate that intrauterine growth restriction (IUGR) followed by accelerated postnatal growth leads to increased risk of developing cardiac disease in adulthood. The aim of this study was to examine the effect of early life growth restriction on cardiac structure and function in young adult rats. IUGR was induced in Wistar Kyoto dams through administration of a low protein diet (LPD; 8.7% casein) during pregnancy and lactation; controls received a normal protein diet (NPD; 20% casein). Cardiac function and structure were assessed in female NPD (n = 7) and LPD (n = 7) offspring at 18 weeks of age by echocardiography and pressure-volume techniques, and systolic blood pressure by tail-cuff sphygmomanometry. LPD offspring remained significantly smaller throughout life compared to controls. There were no differences in the levels of systolic blood pressure, left ventricular cardiac dimensions, heart rate, ejection fraction and fractional shortening of the cardiac muscle between the investigated groups. Aortic peak systolic velocity was significantly reduced in the LPD group (P = 0.02). Our findings support the idea that the programming of adult cardiovascular disease can be prevented or delayed in IUGR offspring when postnatal growth trajectory resembles that of in utero.

Evidence of altered biochemical composition in the hearts of adult intrauterine growth-restricted rats

Epidemiological studies clearly link intrauterine growth restriction with increased risk of cardiac disease in adulthood. The mechanisms leading to this increased risk are poorly understood; remodeling of the myocardium is implicated. The aim was to determine the effect of early life growth restriction on the biochemical composition of the left ventricular myocardium in adult rats. Wistar Kyoto dams were fed either a low protein diet (LPD; 8.7 % casein) or normal protein diet (NPD; 20 % casein) during pregnancy and lactation; from weaning, the offspring were fed normal rat chow. At 18 weeks of age, the biochemical composition of the hearts of NPD control (n = 9) and LPD intrauterine growth-restricted (n = 7) offspring was analyzed using Fourier Transform Infrared (FTIR) micro-spectroscopy. Body weights at postnatal day 4 were significantly lower and remained lower throughout the experimental period in the LPD offspring compared to controls. FTIR analysis of the infrared absorption spectra across the whole "fingerprint" region (1,800-950 cm(-1)) demonstrated wider variation in absorbance intensity in the LPD group compared to controls. In particular, there were marked differences detected in the protein (1,540 cm(-1)), lipid (1,455 and 1,388 cm(-1)), proteoglycan (1,228 cm(-1)) and carbohydrate (1,038 cm(-1)) bands, indicating increased lipid, proteoglycan and carbohydrate content in the growth-restricted myocardium. In conclusion, changes in the biochemical composition of the myocardium provide a likely mechanism for the increased vulnerability to cardiovascular disease in offspring that were growth restricted in early life.

IUGR in the Absence of Postnatal “Catch-Up” Growth Leads to Improved Whole Body Insulin Sensitivity in Rat Offspring

A suboptimal in utero environment leads to fetal adaptations to ensure short-term survival but in the long-term may lead to disease when the postnatal growth does not reflect that in utero. This study examined the effect of IUGR on whole body insulin sensitivity and metabolic activity in adult rats. Female Wistar-Kyoto rats were fed either a normal protein diet (NPD 20% casein) or a low protein diet (LPD; 8.7% casein) during pregnancy and 2 wk of lactation. In offspring at 32 wk of age, indirect calorimetry and dual energy x-ray absorptiometry (DEXA) were performed to assess metabolic activity and body composition. Insulin sensitivity was assessed using a euglycemic-hyperinsulinemic clamp. At 3 d of age, male and female LPD offspring were 23 and 27% smaller than controls, respectively. They remained significantly smaller throughout the experimental period (∼10% smaller at 32 wk). Importantly, there was increased insulin sensitivity in LPD offspring (47% increase in males and 38% increase in females); pancreatic insulin content was normal. Body composition, O2 consumption, respiratory exchange ratio (RER), and locomotor activity were not different to controls. These findings suggest that in the absence of "catch-up" growth IUGR programs for improved insulin sensitivity.

Elevated vascular resistance and afterload reduce the cardiac output response to dobutamine in early growth-restricted rats in adulthood

Epidemiological studies have linked intra-uterine growth restriction (IUGR) with an increased risk of CVD later in life. The aim of the present study was to examine the effect of maternal protein restriction on cardiac function in adulthood during dobutamine (DOB) stimulation. IUGR was induced in Wistar Kyoto dams through administration of a low-protein diet (LPD; 8.7 % casein) during pregnancy and lactation; the control group received a normal-protein diet (NPD; 20 % casein). At 14 weeks of age, cardiac function was assessed in male and female NPD (eight females and eight males) and LPD offspring (ten females and ten males) by pressure volumetry using an anaesthetised closed-chest approach. We determined mean arterial pressure (MAP), heart rate and left ventricular pressure-volume indices under baseline conditions and DOB stimulation (2 and 4 μg/kg per min). During β-adrenergic activation in LPD offspring, increases in cardiac output (CO, P < 0.018) and stroke volume (SV, P < 0.005) were attenuated in comparison with NPD offspring, while increases in ejection fraction and the maximal rate of ventricular pressure development were not affected. LPD females maintained a smaller end-diastolic volume (P < 0.017). MAP did not differ between the groups and did not change significantly during DOB infusion. Arterial elastance and total peripheral resistance decreased in all rats but remained significantly elevated in LPD offspring (P < 0.015 and < 0.01). Early growth restriction did not affect ventricular contractility but led to an increased afterload and impaired the ability to increase SV and CO during β-adrenergic stimulation.

Effect of Maternal Protein Restriction During Pregnancy and Lactation on the Number of Cardiomyocytes in the Postproliferative Weanling Rat Heart

Maternal protein restriction leads to a reduction in the number of cardiomyocytes in the rat heart at birth. However, in rats, cardiomyocytes continue to proliferate until about 2 weeks after birth. Hence, this study aimed to examine the effect of maternal protein restriction, on the number of cardiomyocytes in the young rat heart at a time point when the cardiomyocytes have ceased proliferating and are terminally differentiated. Female Wistar Kyoto rats were fed either a normal protein diet (NPD; 20% casein) or a low protein diet (LPD; 8.7% casein) during pregnancy and lactation. Offspring (seven males and seven females per group) were perfusion fixed at 4 weeks of age. Heart volume and total cardiomyocyte number were determined using stereological techniques. At 4 weeks of age, body weights in both male and female LPD offspring were significantly reduced compared with NPD controls whereas relative heart volumes were significantly increased in LPD offspring. Total number of cardiomyocytes was not significantly different between groups. In both groups, there was a significant linear correlation between cardiomyocyte number and heart volume. In conclusion, total cardiomyocyte number in the postproliferative rat heart does not appear to be affected by maternal protein restriction per se but is directly related to heart size.

Low protein diet fed exclusively during mouse oocyte maturation leads to behavioural and cardiovascular abnormalities in offspring

Early embryonic development is known to be susceptible to maternal undernutrition, leading to a disease-related postnatal phenotype. To determine whether this sensitivity extended into oocyte development, we examined the effect of maternal normal protein diet (18% casein; NPD) or isocaloric low protein diet (9% casein; LPD) restricted to one ovulatory cycle (3.5 days) prior to natural mating in female MF-1 mice. After mating, all females received NPD for the remainder of gestation and all offspring were litter size adjusted and fed standard chow. No difference in gestation length, litter size, sex ratio or postnatal growth was observed between treatments. Maternal LPD did, however, induce abnormal anxiety-related behaviour in open field activities in male and female offspring (P < 0.05). Maternal LPD offspring also exhibited elevated systolic blood pressure (SBP) in males at 9 and 15 weeks and in both sexes at 21 weeks (P < 0.05). Male LPD offspring hypertension was accompanied by attenuated arterial responsiveness in vitro to vasodilators acetylcholine and isoprenaline (P < 0.05). LPD female offspring adult kidneys were also smaller, but had increased nephron numbers (P < 0.05). Moreover, the relationship between SBP and kidney or heart size or nephron number was altered by diet treatment (P < 0.05). These data demonstrate the sensitivity of mouse maturing oocytes in vivo to maternal protein undernutrition and identify both behavioural and cardiovascular postnatal outcomes, indicative of adult disease. These outcomes probably derive from a direct effect of protein restriction, although indirect stress mechanisms may also be contributory. Similar and distinct postnatal outcomes were observed here compared with maternal LPD treatment during post-fertilization preimplantation development which may reflect the relative contribution of the paternal genome.

Effect of Maternal Protein Restriction in Rats on Cardiac Fibrosis and Capillarization in Adulthood

This study examines the effect of maternal protein restriction in rats on levels of cardiac fibrosis, myocardial capillarization, and media:lumen ratio of intramyocardial arteries in adult offspring. Female Wistar Kyoto rats were fed either a normal protein diet (NPD; 20% casein) or a low-protein diet (LPD; 8.7% casein) during pregnancy and lactation. Female offspring (seven per group) were weaned at 4 wk of age and grown to adulthood. At 24 wk of age, the offspring were perfusion fixed. Cardiac fibrosis and media:lumen ratio of intramyocardial arterioles was assessed using image analysis and cardiac capillarization was stereologically investigated. Body weights at 2 and 24 wk of age were significantly reduced (31% and 8%, respectively) in the LPD offspring; however, heart size was not different at 24 wk. Importantly by adulthood, there was a significant 15% increase in left ventricular interstitial fibrosis in LPD offspring. There were no differences in levels of perivascular fibrosis, myocardial capillarization, or in the media:lumen ratio of intramyocardial arteries between groups. Because cardiac fibrosis is associated with impaired cardiac contractility and arrhythmia, our results suggest that induction of interstitial fibrosis may contribute to the increased cardiac disease in adult subjects who were exposed to an adverse intrauterine environment.

A developmental nephron deficit in rats is associated with increased susceptibility to a secondary renal injury due to advanced glycation end-products

The aim of this study was to investigate the effects of a secondary renal insult, due to chronic infusion of AGEs on renal function, and on early pathological markers in rats with a developmental nephron deficit. Female Wistar-Kyoto rats were fed a low-protein diet (LPD; 8.7% casein) or a normal-protein diet (NPD; 20% casein) during pregnancy and lactation. Nephron number was estimated in 4-week-old female offspring. Male offspring were allowed to grow to 20 weeks of age, when AGEs derived from BSA (AGE-BSA) or BSA was infused subcutaneously (20 mg kg(-1) day(-1)) for 4 weeks. At 24 weeks, blood pressure, renal function and circulating and renal AGEs were assessed. Real-time PCR was used to investigate early molecular markers of renal pathology. As expected, maternal protein restriction led to reduced nephron endowment in LPD offspring. This alone did not affect blood pressure or lead to hyperfiltration in adulthood. However, when coupled with the secondary renal insult, the expression of the genes encoding transforming growth factor-beta(1) and procollagen III was significantly upregulated in the kidneys. In addition, there was renal accumulation of AGEs in LPD offspring, and this was exacerbated by AGE infusion. Our results demonstrate that the adult kidney with a reduced nephron endowment is more vulnerable to secondary renal insult from AGE-BSA. Since AGE formation is markedly elevated with hyperglycaemia, our findings suggest that a developmental or acquired deficit may render the kidney susceptible to diabetic renal disease.