Original Article

Obesity (2006) 14, 1330–1343; doi: 10.1038/oby.2006.151

The Importance of Catch-up Growth after Early Malnutrition for the Programming of Obesity in Male Rat*

Florence Bieswal*, Marie-Thérèse Ahn*, Brigitte Reusens*, Paul Holvoet†, Martine Raes‡, William D. Rees§ and Claude Remacle*

  1. *Laboratory of Cell Biology, University of Louvain, Louvain-la-Neuve, Belgium
  2. †Center for Experimental Surgery and Anesthesiology, University of Leuven, Leuven, Belgium
  3. ‡Research Unit of Cell Biology, University of Namur, Namur, Belgium
  4. §Rowett Research Institute, Aberdeen, Scotland

Correspondence: C. Remacle Place Croix du Sud 5, Louvain-la-Neuve, Belgium. E-mail: remacle@bani.ucl.ac.be

*The costs of publication of this article were defrayed, in part, by the payment of page charges. This article must, therefore, be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 1 September 2005; Accepted 26 May 2006.

Top

Abstract

Objective: To investigate whether catch-up growth after maternal malnutrition would favor the development of obesity in adulthood.

Research Methods and Procedures: Pregnant rats were submitted to protein or calorie restriction during the course of gestation. During lactation, pups were protein-restricted, normally fed, or overfed [ reduced litter size, control (C) diet] . At weaning, rats were transferred to chow or to a hypercaloric diet (HCD) known to induce obesity. Body weight, food intake, blood parameters, glucose tolerance, adipocyte cellularity, and adipose factors contributing to cardiovascular disease development were measured.

Results: Protein and calorie restriction during gestation led to growth retardation at birth. If malnutrition was prolonged throughout lactation, adult body weight was permanently reduced. However, growth-retarded offspring overfed during the suckling period underwent a rapid catch-up growth and became heavier than the normally fed Cs. Offspring of calorie-restricted rats gained more weight than those of dams fed protein-restricted diet. Feeding an HCD postnatally amplified the effect of calorie restriction, and offspring that underwent catch-up growth became more obese than Cs. The HCD was associated with hyperphagia, hyperglycemia, hyperinsulinemia, glucose intolerance, insulin resistance, and adipocyte hypertrophy. The magnitude of effects varied depending on the type and the timing of early malnutrition. The expression of genes encoding factors implicated in cardiovascular disease was also modulated differently by early malnutrition and adult obesity.

Discussion: Catch-up growth immediately after early malnutrition should be a key point for the programming of obesity.

Keywords:

fetal programming, maternal malnutrition, catch-up growth, hypercaloric diet, cardiovascular disease

Top

Introduction

Low birth weight due to fetal growth retardation is associated with increased risk of developing impaired glucose tolerance, type 2 diabetes, hypertension, and cardiovascular disease in adult life (1). There is particular interest in the observations that the fetal and postnatal environment may predispose an individual to developing obesity (2). Epidemiological studies have shown that people who were exposed to famine during early pregnancy have higher rates of intra-abdominal adiposity than those exposed to undernutrition during late pregnancy (3). Other reports have highlighted the importance of the postnatal environment. In a study of infants, in India, the percentage of body fat, the development of insulin resistance, and central obesity were greatly enhanced when low birth weight was followed by rapid postnatal catch-up growth (4, 5). Because intra-abdominal obesity is known to increase the risk of cardiovascular disease and type 2 diabetes independently of the appearance of generalized obesity, prenatal programming of visceral adipose development may be particularly important in the progression of metabolic disease.

Several animal models of diet restriction have been used to study the early programming of obesity and cardiovascular diseases (6). For example, in a rat model where severe maternal calorie restriction was applied throughout gestation, the offspring developed hyperphagia, obesity, hyperleptinemia, hyperinsulinism, and hypertension in adult life (7). Programming of adult hypertension, independently of the development of obesity, was also observed in rats protein-restricted during fetal life. However, this phenomenon varied with the period of malnutrition and the diet composition (8, 9). The animal studies have, like the human studies, highlighted the importance of the early postnatal environment for adult health. In mice, the offspring of dams fed a low-protein (LP)1 diet during pregnancy and then allowed to catch up by having the nursing dams fed a control (C) diet gained more weight when given free access to a hypercaloric diet (HCD) (10, 11). The catch-up growth immediately after intrauterine calorie restriction had also been suggested to program obesity and orexigenic hormones in offspring fed normally after birth (12). Detrimental effects of early growth restriction on weight, appetite regulation, metabolism, and cardiovascular system are not fully understood but seem dependent on the timing and the type of nutrient restriction and the postnatal environment.

The aim of the present study was to determine the critical time window and the effects of different models of malnutrition for the programming of obesity and its associated metabolic disorders. First, we examined whether protein malnutrition during perinatal life (gestation and lactation) is a risk factor for the development of obesity in adulthood. Second, we aimed to determine whether early catch-up growth, induced by overfeeding during the suckling period, leads to the programming of obesity in fetal growthrestricted offspring. In this latter part, the impact of two types of fetal malnutrition (protein vs. calorie) was compared. We examined also the expression, in adipose tissue, of selected molecules implicated in fibrinolysis [ plasminogen activator inhibitor (PAI-1)] and hypertension [ angiotensinogen (AGT)] , and those exhibiting anti-atherogenic properties (adiponectin).

Top

Research Methods and Procedures

Animals

Wistar rats (Janvier, Le Genest Saint Isle, France) bred in our laboratory were maintained under controlled conditions (25 °C, 12-/12-hour dark/light cycle) with free access to food and water. Virgin females (3 months old and weighing 250 to 350 grams) were caged with male rats overnight, and, after confirmation of mating, dams were housed individually. All animal studies were performed according to the guidelines of the animal ethics committee of the Catholic University of Louvain, Belgium. This study was carried out as two experiments. Apart from diets and experimental groups, materials and methods were common to both experiments.

Diets and Food Intake Measurements

C diet (200 g protein/kg) and isocaloric LP diet (80 g protein/kg) were purchased from Hope Farm (Woerden, Netherlands); the composition and source of the diets were as previously described (13). Standard laboratory chow contained 160 g protein/kg (Carfil Quality, Turnhout, Belgium). The HCD was made according to Ruffin et al. (14) and consisted of an emulsion of 10% sucrose solution mixed with corn oil. Corn oil was added in concentration that was increased progressively to familiarize the rats to the taste of the diet (4% , 2.66 kJ/g diet for 10 days; 8% , 3.72 kJ/g diet for 15 days; 16% , 5.59 kJ/g diet for 15 days; 32% , 8.69 kJ/g diet until the end of the experiment). Tween-80 and lecithin were added to emulsify the lipids in the sucrose solution. The HCD was supplied alongside standard chow diet. In this way, rats fed the HCD had the choice of selecting their normal chow or the liquid HCD. Daily energy intake was controlled by measuring separately the consumption of chow and HCD.

Dietary Groups for Experiment 1

The experimental protocol for the first experiment is shown schematically in Figure 1A. Dams were fed, ad libitum, during gestation and lactation either the C diet (20% , C group) or the isocaloric LP diet (8% LP group, LP1). At birth, litter size was adjusted to eight pups per dam to ensure adequate and standardized nutrition until weaning (male-to-female ratio was 4:4). At weaning (3 weeks), male pups were transferred to standard laboratory chow (16% ), whereas female pups were discarded from the study. At the age of 7 weeks, one-half of the males of each group received, in addition to chow, the HCD (C-HC and LP1-HC groups, n = 8 per group). The other one-half continued with chow only (C and LP1 groups, n = 8 per group). Male rats were maintained with these diets until 42 weeks of age. Body weight and food intake were recorded weekly after weaning. At 42 weeks of age, animals were anesthetized with pentobarbital (75 muL/100 grams body weight; CEVA Santé Animale, Bruxelles, Belgium) and killed by decapitation. Subcutaneous, perirenal, periepididymal, and mesenteric fat pads were removed, weighed, immediately frozen in liquid nitrogen, and stored at - 80 °C.

Figure 1.
Figure 1 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Schematic representation of the dietary groups for Experiments 1 (A) and 2 (B). C, C rats; C-HC, C rats fed with the HCD; LP1, rats submitted to protein restriction during gestation and lactation; LP1-HC, rats submitted to protein restriction during gestation and lactation and fed thereafter with the HCD; LP2, rats submitted to protein restriction during gestation; LP2-HC, rats submitted to protein restriction during gestation and fed thereafter with the HCD; LC, rats submitted to calorie restriction during gestation; LC-HC, rats submitted to calorie restriction during gestation and fed thereafter with the HCD; 20% , diet containing 20% protein; 8% , diet containing 8% protein; 16% , diet containing 16% protein.

Full figure and legend (88K)

Dietary Groups for Experiment 2

The experimental protocol for the second experiment is shown schematically in Figure 1B. During gestation, dams were assigned to one of three nutritional groups: C group, in which dams were fed, ad libitum, the C diet containing 20% protein; LP group (LP2), in which dams received, ad libitum, the isocaloric LP diet containing 8% protein; and the low-calorie (LC) group, in which dams were fed the 20% protein diet with a daily intake reduced to 50% compared with C dams. Food intake and maternal weight were recorded daily from the 1st day of gestation until birth (day before delivery). At birth, pups were weighed and measured, and litter sizes were recorded. Pups from each group were nursed by dams fed the 20% protein diet ad libitum. To favor a catch-up growth, the number of pups was reduced to four per litter in LP2 and LC groups (four males per litter) as opposed to eight in C group (eight males per litter), thereby creating a group of "overfed" rats. At weaning (3 weeks), male offspring were fed either standard laboratory chow only (C, LP2, and LC groups; n = 8 per group) or chow supplemented with the HCD (C-HC, LP2-HC, and LC-HC groups; n = 8 per group). Male rats were maintained on these diets until 38 weeks of age. Body weight and food intake were recorded weekly after weaning. At 38 weeks of age, animals were anesthetized with CO2 and killed by decapitation. Subcutaneous, perirenal, periepididymal, and mesenteric fat pads were removed, weighed, immediately frozen in liquid nitrogen, and stored at - 80 °C.

Blood Sampling and Analysis

Blood from fed animals was collected monthly after weaning. Blood was taken from the tail vein and collected in heparinized tubes chilled on ice. Blood glucose was measured by the glucose oxidase method using glucose Trinder's reagent (Stanbio Laboratory, Boerne, TX). Plasma insulin and triglycerides were determined using ultrasensitive rat insulin enzyme-linked immunosorbent assay (Mercodia, Uppsala, Sweden) and TRF400CH tests (Chema Diagnostica, Jesi, Italy), respectively. In Experiment 2, leptin was measured by enzyme-linked immunosorbent assay test (Quantikine, MOB00; R&D Systems, Lille, France).

Intraperitoneal Glucose Tolerance Test (IPGTT)

An IPGTT was performed on 32-week-old (Experiment 1) or 36-week-old (Experiment 2) rats. After 16 hours of fasting, conscious rats were injected intraperitoneally with 1 gram of glucose/kg body weight (20% glucose solution in NaCl 0.9% ). Blood was collected from the tail vein before injection (time 0) and 15, 30, 60, 120, and 180 minutes after glucose injection. Samples for glucose and insulin determinations were analyzed as described above.

Fat Cell Size

Fat cell size was estimated for retroperitoneal fat tissue isolated from 42-week-old (Experiment 1) and 38-week-old (Experiment 2) male rats. Fat cells were fixed as described by Etherthon et al. (15). Briefly, 100 to 150 mg of retroperitoneal adipose tissue was sliced and rinsed in 37 °C 0.154 M NaCl. Adipose slices were then fixed in 3% osmium tetroxide in collidine-HCl buffer (50 mM, pH 7.4) for 72 to 96 hours. Fixed cells were rinsed in 0.154 M NaCl for 24 hours and liberated from connective tissue by incubation in 8 M urea for 24 to 48 hours. Cell size was determined using the ZEISS KS 400 3.0 software in an Axioskop2 Mot Plus Zeiss microscope (Carl Zeiss GmbH, Jena, Germany).

Carcass Analysis

The eviscerated carcasses from 42-week-old male rats (Experiment 1) were minced, lyophilized, and ground to a fine, uniform powder. Lipids were determined gravimetrically after extraction in petroleum ether (Sigma-Aldrich, St. Louis, MO) using a Soxtec system HT6 extraction unit (Tecator AB, Hoganas, Sweden). Moisture was determined by drying overnight at 105 °C and ash content by overnight combustion at 540 °C to 550 °C. Nitrogen was determined by combustion in an Automated Dumas system (Foss Electric Ltd,. Warrington, Cheshire, United Kingdom).

RNA Extraction and Real-Time Polymerase Chain Reaction (PCR) Analysis

Total RNA was isolated, as previously described (16), from the perirenal fat tissue of 42-week-old (Experiment 1) and 38-week-old (Experiment 2) rats. The quality of RNA was assessed by electrophoresis in 1% agarose gels stained with ethidium bromide. Samples were stored in 0.1% diethyl pyrocarbonate-treated water at - 80 °C. For first strand cDNA synthesis, 2.5 mug of total RNA was reverse-transcribed in a 45-muL volume using random hexamers (Amersham Bioscience, Piscataway, NJ) as primers and the SuperScript RNase H- reverse transcriptase system (Invitrogen Life Technology, Gaithersburg, MD). Realtime PCR was performed with the ABI Prism 7000 Sequence Detection System instrument and software under standard conditions (Applied Biosystems, Foster City, CA). Gene-specific primers and probes were designed using the Primer Express 2.0 software (Applied Biosystems). AGT and adiponectin were quantified using qPCR Mastermix Plus for SYBR Green I (Eurogentec, Seraing, Belgium). Plasminogen inhibitor-1 and 18S mRNAs were quantified with the Taqman probe system using qPCR Mastermix (Eurogentec). After optimization for each gene, the following primer and probe concentrations were used: AGT, 5'-CTGGGCAAGATGGGTGACA-3' (forward; 400 nM) and 5'-TCCTCGCCTGCTTGGAGTT-3' (reverse; 400 nM); adiponectin, 5'-ACAAGGCCGTTCTCTTCACCTA-3' (forward; 50 nM) and 5'-CCAGATGGAGGAGCATGGA-3' (reverse; 50 nM); 5'-TGGAGAAGCTGGGCATGACT-3' (forward, 800 nM), plasminogen inhibitor-1, 5'-6FAM-ACATCTTCAGCTCAACCCAGGCCGA-TAMRA-3' (probe, 100 nM), and 5'-TCTTGGTCGGAAAGACTTGTGA-3' (reverse, 800 nM); and 18S RNA, 5'-GATCCATTGGAGGGCAAGTCT-3' (forward, 800 nM), 5'-VIC-CCAGCAGCCGCGGTAATTCCAG-TAMRA-3' (probe, 100 nM), and 5'-GCAGCAACTTTAATATACGCTAT-TGG-3' (reverse; 800 nM). The authenticity of the PCR products was verified by melting curve and agarose gel electrophoresis. A standard curve was generated for each gene by a six-point serial dilution of a pool of adipose tissue total RNA from non-treated rats. Each sample was run in duplicate, and the mean value of the duplicates was used to calculate the transcript level. cDNA quantities were normalized to 18S RNA concentrations and then expressed as percentage in relation to the C group.

Statistical Analysis

Experimental results are reported as means plusminus standard error (SE). Data were analyzed by one-way repeated measures ANOVA (body weight, food intake, insulin, glucose, triglycerides, and IPGTT) or by one-way ANOVA (leptin, adipose tissue weights, organs weights, RNA measurements, and carcass analysis) followed by Tukey multiple comparison post hoc test. Kolmogorov-Smirnov two-sample test was performed to analyze data of fat cell size. Data were processed using the Prism Software (GraphPad Software Inc., San Diego, CA). Differences with p < 0.05 were considered significant.

Top

Results

Experiment 1: The Effect of Protein Restriction during Gestation and Lactation for the Programming of Obesity in Adult Life

In Experiment 1, we aimed to determine whether protein restriction during early life (gestation and lactation) favored the development of obesity in adult male rats. Four experimental groups were followed as described in Figure 1A.

Body Weight and Food Intake

As a result of protein restriction during gestation and lactation, body weights of LP1 pups at weaning were lower than those of C pups (67.3 plusminus 2.8 vs. 89.2 plusminus 2.6 grams, respectively, p < 0.001). This growth retardation persisted during adult life regardless of the diet fed after weaning (C vs. LP1, p < 0.01; C-HC vs. LP1-HC p < 0.05; Figure 2A). Hypercaloric nutrition induced a similar degree of obesity in both C-HC and LP1-HC groups (p < 0.001). Total caloric intake was significantly increased in rats supplemented with the HCD (Figure 2B). These animals ingested daily approx1.5 more calories than normal-fed rats (p < 0.001); however, there was no effect of early protein restriction on food intake.

Figure 2.
Figure 2 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

(A) Postnatal growth curves of offspring from Experiment 1 from weaning to 42 weeks of age. (B) Daily energy intake (kilojoules per day) from the time of HCD supplementation (7-week-old rats). (filled square) Daily energy intake (kilojoules per day) coming from the laboratory chow. (square) Daily energy intake (kilojoules per day) coming from the HCD. C, C rats; C-HC, C rats fed with the HCD; LP1, rats submitted to protein restriction during gestation and lactation; LP1-HC, rats submitted to protein restriction during gestation and lactation and fed thereafter with the HCD. Data represent means plusminus SE with n = 8 per group. LP1 vs. C and LP1-HC vs. C-HC, § p < 0.05 and §§ p < 0.01; C-HC vs. C and LP1-HC vs. LP1, *** p < 0.001.

Full figure and legend (56K)

Fat Accumulation and Fat Cell Size

Perirenal, periepididymal, mesenteric, and subcutaneous fat pads were significantly heavier in both C-HC and LP1-HC groups (p < 0.001; Table 1). Total fat in the carcass was also significantly elevated in both groups of rats fed HCD (p < 0.01). However, there was no effect of early malnutrition on either fat accumulation or the distribution in the particular depots. Adipocyte hypertrophy was observed in both C-HC and LP1-HC offspring (p < 0.01; Figure 3). Perirenal adipose tissue from protein-restricted offspring contained smaller adipocytes than that of C animals (p < 0.01); as a result, hypertrophy was more pronounced in LP1-HC rats when compared with the C-HC animals.

Figure 3.
Figure 3 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Distribution of fat cell diameter in perirenal adipose tissue isolated from 42-week-old offspring. Frequency of cell diameter is expressed in percentage for each class of size. Values are means of the size distribution in eight rats (1500 cells per rat). C, C rats; C-HC, C rats fed with the HCD; LP1, rats submitted to protein restriction during gestation and lactation; LP1-HC, rats submitted to protein restriction during gestation and lactation and fed the HCD thereafter. LP1 vs. C, § p < 0.01; C-HC vs. C and LP1-HC vs. LP1, ** p < 0.01.

Full figure and legend (90K)


Blood Parameters

Protein restriction during gestation and lactation had no effect on blood glucose and plasma insulin but reduced the concentration of plasma triglycerides in the offspring (p < 0.05; Figure 4). The development of obesity was associated with hyperglycemia in both C-HC and LP1-HC groups (p < 0.001) and with hyperinsulinemia in C-HC rats (p < 0.05). Plasma triglycerides were reduced in the C-HC group compared with C (p < 0.05). The IPGTT showed that, despite a blunted insulin response (p < 0.01), LP1 offspring presented a rate of glucose disappearance similar to that of Cs, indicating a better sensitivity to insulin. After glucose challenge, C-HC and LP1-HC offspring had higher glucose and insulin concentration compared with chow-fed rats (p < 0.001), revealing glucose intolerance and insulin resistance in obese animals (Figure 5).

Figure 4.
Figure 4 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Blood concentrations of glucose (A), insulin (B), and triglycerides (C). Values are mean concentrations from weaning to 42 weeks of age plusminus SE with n = 8 per group. C, C rats; C-HC, C rats fed with the HCD; LP1, rats submitted to protein restriction during gestation and lactation; LP1-HC, rats submitted to protein restriction during gestation and lactation and fed thereafter with the HCD. LP1 vs. C, § p < 0.05; C-HC vs. C and LP1-HC vs. LP1, * p < 0.05 and *** p < 0.001.

Full figure and legend (42K)

Figure 5.
Figure 5 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Blood glucose concentration (A) and insulin response (B) during IPGTT in 32-week-old rats. Values are means plusminus SE with n = 8 per group. C, C rats; C-HC, C rats fed with the HCD; LP1, rats submitted to protein restriction during gestation and lactation; LP1-HC, rats submitted to protein restriction during gestation and lactation and fed thereafter with the HCD. LP1 vs. C, §§ p < 0.01; C-HC vs. C and LP1-HC vs. LP1, *** p < 0.001.

Full figure and legend (72K)

mRNA Expression in Adipocytes

Protein restriction during early life did not alter AGT and adiponectin mRNA but tended to lower the abundance of PAI-1 mRNA in adult rats (p = 0.088, Figure 6). PAI-1 mRNA expression was significantly higher in C-HC offspring compared with Cs (p < 0.05). The same tendency was observed in LP1-HC rats but to a much lesser extent than in C-HC animals (p = 0.147). In contrast, AGT and adiponectin mRNA abundance tended to be lower in C-HC rats (AGT, p = 0.072; adiponectin, p = 0.078) and was significantly reduced in LP1-HC rats (p < 0.05).

Figure 6.
Figure 6 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Relative mRNA expression of PAI-1, AGT, and adiponectin in retroperitoneal adipose tissue from 42-week-old rats. mRNA expressions were related to 18S mRNA and then expressed relative to the C group. Values are means plusminus SE with n = 8 per group. C, C rats; C-HC, C rats fed with the HCD; LP1, rats submitted to protein restriction during gestation and lactation; LP1-HC, rats submitted to protein restriction during gestation and lactation and fed thereafter with the HCD. C-HC vs. C and LP1-HC vs. LP1, * p < 0.05.

Full figure and legend (42K)

Experiment 2: The Effect of Postnatal Catch-up Growth for the Programming of Obesity in Adult Life

In Experiment 2, we aimed to determine the effect of an early catch-up growth, induced by postnatal overfeeding, on the programming of adult obesity in male rats. Six experimental groups were followed as described in Figure 1B.

Maternal Body Weight Gain, Litter Size, and Offspring Parameters at Birth

Maternal body weight gain during gestation was significantly lower in LC and LP2 dams when compared with Cs (p < 0.001; Table 2). Moreover, dams restricted to 50% caloric intake gained significantly less weight than LP2 dams (p < 0.001). Litter size was similar between groups. Maternal malnutrition resulted in fetal growth retardation reflected by a significantly lower body weight and length in LP2 and LC offspring at birth (p < 0.001).


Body Weight and Food Intake

Because of reduced litter size during lactation, growth-retarded offspring had largely recouped their weight to the weight of Cs at weaning (C, 48.64 plusminus 1.15 grams; LP2, 53.81 plusminus 3.55 grams; and LC, 55.43 plusminus 1.20 grams; p < 0.05). These offspring continued to gain more weight than Cs and at 38 weeks of age were significantly heavier than the C animals (p < 0.001; Figure 7A). The body weight gain for protein and calorie-restricted offspring was similar. Rats supplemented with the HCD gained significantly more weight than their respective Cs and became heavier than the Cs (p < 0.001). Weight gain was more pronounced in LP2-HC and LC-HC offspring compared with C-HC rats (p < 0.001). Moreover, the type of dietary restriction during early life was important. Offspring submitted to caloric restriction during gestation became significantly heavier than protein-restricted offspring (p < 0.001). Animals supplemented with the HCD showed global hyperphagia compared with chow-fed animals (p < 0.001; Figure 7B), but there was no difference in total energy intake according to the early diet. Most of the energy was coming from the HCD (p < 0.001).

Figure 7.
Figure 7 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

(A) Postnatal growth curves of offspring from weaning to 38 weeks. (B) Daily energy intake (kilojoules per day) from the time of HCD supplementation (3-week-old rats). (filled square) Daily energy intake (kilojoules per day) coming from the laboratory chow. (square) Daily energy intake (kilojoules per day) coming from the HCD. Data represent means plusminus SE with n = 8 per group. C, C rats; C-HC, C rats fed with the HCD; LP2, rats submitted to protein restriction during gestation; LP2-HC, rats submitted to protein restriction during gestation and fed thereafter with the HCD; LC, rats submitted to calorie restriction during gestation; LC-HC, rats submitted to calorie restriction during gestation and fed thereafter with the HCD. LP2 vs. C and LC vs. C, §§§ p < 0.001; LP2-HC vs. LC-HC, ††† p < 0.001; HCD vs. chow, *** p < 0.001.

Full figure and legend (69K)

Fat Accumulation and Fat Cell Size

LC offspring stored significantly more fat than Cs (p < 0.05; Table 3). The same tendency was observed in offspring submitted to fetal protein restriction but to a lesser extent (p = 0.06). As expected, the increase in body weight was linked with elevated adipose tissue weight (p < 0.001), and the accumulation of perirenal fat was greater in LC-HC offspring compared with C-HC (p < 0.01) and LP2-HC (p < 0.05) animals. This fat accumulation was already apparent in LC rats (p < 0.05). When related to body weight, the data indicate preferential intra-abdominal fat accumulation in LC and LC-HC offspring. Perirenal adipose tissue from protein and calorie-restricted offspring contained larger adipocytes than that of C animals (p < 0.01; Figure 8). Fat cell diameter varied also with the type of early malnutrition, and LC offspring had larger adipocytes than LP2 rats (p < 0.01). Hypertrophy was linked to an increase in the fat mass in each group. Parallel to hypertrophy, a second population of smaller fat cells (diameter, approx80 to 120 mum) was detected in perirenal adipose tissue of obese rats.

Figure 8.
Figure 8 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Distribution of fat cell diameter in perirenal adipose tissue isolated from 38-week-old rats. Frequency of cell diameter is expressed in percentage for each class of size. Values are means of the size distribution in eight rats (1500 cells per rat). C, C rats; C-HC, C rats fed with the HCD; LP2, rats submitted to protein restriction during gestation; LP2-HC, rats submitted to protein restriction during gestation and fed thereafter with the HCD; LC, rats submitted to calorie restriction during gestation; LC-HC, rats submitted to calorie restriction during gestation and fed thereafter with the HCD. LP2 vs. C and LC vs. C, §§ p < 0.01; LC vs. LP2, †† p < 0.01; HCD vs. chow, ** p < 0.01.

Full figure and legend (65K)


Blood Parameters

Plasma leptin and triglyceride levels were higher in LP2 and LC offspring compared with Cs (Figure 9, C and D). Fat gain due to feeding the HCD resulted in hyperleptinemia (p < 0.001). Plasma triglyceride concentrations were also raised in obese rats, but only C-HC and LP2-HC animals developed significant hypertriglyceridemia (p < 0.05). Plasma triglycerides were positively correlated with homeostasis model assessment index (p < 0.05; Pearson coefficient, 0.2917; data not shown). Blood glucose concentrations were not altered by early malnutrition, but plasma insulin was significantly elevated in LP2 offspring (p < 0.05; Figure 9, A and B). Feeding the HCD led to hyperglycemia and hyperinsulinemia (p < 0.001). IPGTT revealed that the rates of insulin secretion and glucose disappearance were similar in C, LP2, and LC offspring. Obesity was associated with glucose intolerance and insulin resistance (p < 0.001), with insulin resistance being higher in LC-HC offspring (LC-HC vs. C-HC, p < 0.05 and LC-HC vs. LP2-HC, p < 0.001; Figure 10).

Figure 9.
Figure 9 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Blood concentrations of glucose (A), insulin (B), triglycerides (C), and leptin (D). Values are mean concentrations from weaning to 38 weeks of age plusminus SE with n = 8 per group. C, C rats; C-HC, C rats fed with the HCD; LP2, rats submitted to protein restriction during gestation; LP2-HC, rats submitted to protein restriction during gestation and fed thereafter with the HCD; LC, rats submitted to calorie restriction during gestation; LC-HC, rats submitted to calorie restriction during gestation and fed thereafter with the HCD. LP2 vs. C and LC vs. C, § p < 0.05, §§ p < 0.01, and §§§ p < 0.001; LC vs. LP2, † p < 0.05; HCD vs. chow, * p < 0.05 and *** p < 0.001.

Full figure and legend (97K)

Figure 10.
Figure 10 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Blood glucose concentration (A) and insulin response (B) during IPGTT in 36-week-old rats. Values are means plusminus SE with n = 8 per group. C, C rats; C-HC, C rats fed with the HCD; LP2, rats submitted to protein restriction during gestation; LP2-HC, rats submitted to protein restriction during gestation and fed thereafter with the HCD; LC, rats submitted to calorie restriction during gestation; LC-HC, rats submitted to calorie restriction during gestation and fed thereafter with the HCD. LC vs. C, § p < 0.05; LC vs. LP2, † p < 0.05; HCD vs. chow, *** p < 0.001.

Full figure and legend (75K)

mRNA Expression in Adipocytes

The mRNA abundance of PAI-1 (p < 0.01), AGT (p < 0.05), and adiponectin (p < 0.05) was increased in LC offspring compared with Cs (Figure 11). The same tendency was observed in LP2 offspring but to a lesser extent (AGT, p < 0.05). Obesity was associated with increased expression of PAI-1 in C-HC rats (p = 0.0653) and decreased expression of AGT (C-HC, p < 0.05; LP2-HC, p < 0.01; LC-HC, p < 0.01). LP2-HC and LC-HC showed lower adiponectin mRNA content compared with lean rats (p < 0.01).

Figure 11.
Figure 11 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Relative mRNA expression of PAI-1, AGT, and adiponectin from 38-week-old rats. mRNA expressions were related to 18S RNA and then expressed relative to the C group. Values are means plusminus SE with n = 8 per group. C, C rats; C-HC control rats fed with the HCD; LP2, rats submitted to protein restriction during gestation; LP2-HC, rats submitted to protein restriction during gestation and fed thereafter with the HCD; LC, rats submitted to calorie restriction during gestation, LC-HC, rats submitted to calorie restriction during gestation and fed thereafter with the HCD. LP2 vs. C and LC vs. C, § p > 0.05 and §§ p < 0.01; HCD vs. chow, * p < 0.05 and ** p < 0.01.

Full figure and legend (96K)

Top

Discussion

In this study, we showed that early postnatal catch-up growth occurring after fetal malnutrition favors the programming of obesity in adult life. The outcome depends on the type of fetal malnutrition experienced and the degree of catch-up growth. This study shows that both the type of dietary restriction and the time at which malnutrition was applied in early life are critical for programming adult obesity.

Protein restriction during gestation and lactation resulted in growth retardation, which persisted even if a normal diet was given at weaning. In contrast, pups subjected to protein or calorie deficiency throughout gestation and overfed during lactation were able to regain similar weights to the Cs. The accelerated body weight gain continued after weaning so that pups malnourished in gestation went on to become heavier than Cs. This effect was amplified by feeding an HCD after weaning. The observation that catch-up growth programs the development of obesity corroborates recent studies (10, 11, 12). Moreover, calorie restriction was more potent than protein restriction, with the offspring developing intra-abdominal obesity without the challenge of an HCD.

The expansion of fat tissue involves both hypertrophy and hyperplasia of adipocytes. The early stages of life are a period very sensitive to nutritional and hormonal factors, which modulate the multiplication and differentiation of fat cell precursors (17). Recently, we (13) showed that pre-adipocytes derived from protein-restricted offspring similar to those used in Experiment 1 proliferated and differentiated at similar rates in vitro. Nevertheless, the present study indicates that the size distribution of adipocytes and the growth of adipose tissue were sensitive to the type and the timing of early malnutrition. Malnutrition during gestation resulted in adipocyte hypotrophy (18), whereas overfeeding during lactation, which is associated with higher activities of lipogenic enzymes (19, 20), resulted in adipocyte hypertrophy. In parallel to hypertrophy induced by the HCD, a second population of smaller fat cells appeared, suggesting the recruitment and differentiation of new fat cells from the pool of precursors present in adipose tissue. It should be interesting to re-examine in vitro the capacity of preadipocyte proliferation and differentiation to determine whether postnatal catch-up growth may modify the pool of fat cell precursors.

There are conflicting reports on the development of adult hyperphagia after fetal growth retardation (7, 12, 21, 22, 23). The early postnatal period has been identified as the critical developmental step for hypothalamic nuclei involved in the central nervous system regulation of food intake, body weight, and metabolism (24, 25). Both overfeeding and protein restriction during lactation have been postulated to induce a permanent dysregulation of the main hypothalamic centers of food regulation (26, 27, 28). However, we have not observed early programming of appetite in this study, and the results would suggest that the programming of obesity was not a direct consequence of altered appetite regulation.

Vickers et al. (7) indicated that severe fetal undernutrition programmed hyperleptinemia leading to leptin resistance, hyperphagia, and obesity in adult life. All of these changes were amplified by postnatal hypercaloric nutrition. In our study, we showed that the programming of obesity was associated with increased plasma leptin levels in adulthood but without dysregulation in food intake. In all cases, plasma leptin concentrations observed in offspring fed the chow diet after weaning were proportional to fat mass. Therefore, elevated leptin levels appeared as the direct consequence of the higher adipose tissue development rather than impaired leptin action and regulation. In obese rats, persisting hyperphagia in presence of hyperleptinemia suggests that obesity came with central resistance to leptin.

We and others (22, 29, 30) have shown that, depending on the type and the timing of fetal malnutrition, there may be changes in the endocrine pancreas that lead to permanent reduction of beta-cell mass and impairment of insulin secretion. In the present study, we showed that insulin response after a glucose load was blunted in adult offspring that had been protein-restricted during gestation and lactation, whereas it was comparable with that of Cs in offspring malnourished only during fetal life. Despite their blunted insulin response, offspring fed an LP diet during gestation and lactation exhibited normal glucose tolerance. This may be explained by an increased whole-body insulin sensitivity making appropriate the circulating insulin concentration (23, 31, 32, 33, 34). After feeding with an HCD, glucose tolerance was impaired, and the animals became insulin resistant after the development of obesity. The insulin resistance was nevertheless higher in offspring submitted to intrauterine calorie restriction compared with other groups. Because overfeeding during lactation does not seem to exert long-term effects on insulin secretion and glucose tolerance (20), the programming of insulin resistance associated with obesity may be attributed to the calorie restriction during fetal life.

Our results also suggest that lipid metabolism is programmed during lactation. Feeding the dam an LP diet during this period permanently lowered plasma triglyceride concentrations, whereas overfeeding of the offspring raised them. Low plasma triglyceride concentration in offspring submitted to protein restriction during gestation and lactation was also observed by Lucas et al. (35) and should reflect a greater uptake of triglycerides from the circulation. Indeed, in our study, we found that triglyceride plasma levels were positively correlated with insulin sensitivity (homeostasis model assessment index). In addition, the expression of genes involved in the uptake of fatty acid, including very low-density-lipoprotein receptor and low-density lipoprotein-related protein-1, was enhanced in adipose tissue from offspring protein-restricted during gestation and lactation (36). Such alterations favoring fat storage may be advantageous to survival under poor nutrition conditions and are in accordance with the thrifty phenotype hypothesis. Plasma triglyceride levels were increased in the offspring when malnutrition was applied only during fetal life. This may suggest that postnatal overfeeding may jeopardize fetal adaptations to nutrient restriction and, therefore, lead to detrimental effects such as hypertriglyceridemia in adult life. The switch from high-fat diet (maternal milk) to high-carbohydrate diet (laboratory chow) occurring usually at the suckling-weaning transition is a determinant step for insulin sensitivity (37). Weaning rats onto a high-fat diet suppressed the appearance of lipogenic enzymes and led to impaired insulin sensitivity (38). These changes might explain why our C rats weaned onto HCD (high fat and high sucrose) developed hypertriglyceridemia, whereas rats weaned onto chow for 1 month and supplemented with the HCD thereafter did not.

Obesity is a common risk factor for cardiovascular disease and hypertension because adipose tissue secretes several factors that contribute to the development of such pathologies. For example, human studies have shown that PAI-1, an inhibitor of fibrinolysis, is positively correlated with BMI (39) and that secretion of PAI-1 contributes to the elevated serum PAI-1 levels in obesity (40). Our results showed that increased fat accretion was associated with an increased PAI-1 mRNA expression in C animals but not in early growth-restricted rats. Among factors regulating PAI-1 expression in adipocytes, catecholamines, which are known to suppress PAI-1 mRNA and protein release in human adipocytes, are of particular interest (41). Indeed, circulating catecholamine concentration is increased in adult offspring of dams fed a reduced protein diet throughout pregnancy and lactation (42). Moreover, in vitro experiments with isolated adipocytes have suggested that adipocytes from LP offspring are more sensitive to the action of catecholamines (43, 44). Such long-term effects of catecholamine metabolism might contribute to the disturbance of the fibrinolytic system observed in our study. Adiponectin exhibits putative anti-atherogenic properties by inhibiting the adhesion of monocytes to endothelial cells, reducing vascular smooth muscle cell proliferation and migration and suppressing lipid accumulation and class A scavenger receptor expression (45). Adiponectin is down-regulated in human obesity (46). Similarly, we showed that adiponectin expression was reduced in obese rats. However, there was no effect of early malnutrition. On the other hand, in this study, we showed elevated AGT expression in adipose tissue from offspring protein- or calorie-restricted during fetal life and presenting a postnatal catch-up growth. Several animal studies support the hypothesis that hypertension is programmed during fetal life in offspring exposed to intrauterine growth retardation (47). Our results suggest that the programming of elevated adipose AGT expression in offspring growth-restricted during fetal life may be a key step in hypertension development. In addition, because angiotensin II, the product of AGT, acts as a trophic factor implicated in adipocyte differentiation and fat mass enlargement (48, 49), the programming of elevated levels of AGT in adipocytes may contribute to the adipocyte hypertrophy observed in these growth-restricted offspring. It could, therefore, favor the development of obesity.

In conclusion, maternal malnutrition at defined time windows has long-term effects on weight gain and adipocyte metabolism. Lactation is a critical period for the programming of obesity. Indeed, overfeeding immediately after fetal growth retardation induces a catch-up growth and leads to the programming of obesity at adulthood. In contrast, permanent weight deficit is observed if malnutrition is applied throughout gestation and lactation. The metabolic abnormalities resulting from early programming differ according to the type of fetal malnutrition.

Top

Notes

1 Nonstandard abbreviations: LP, low protein; LC, low calorie; C, control; HCD; hypercaloric diet; PAI-1, plasminogen activator inhibitor-1; AGT, angiotensinogen; IPGTT, intraperitoneal glucose tolerance test; PCR, polymerase chain reaction; SE, standard error.

Top

References

  1. Godfrey, K. M., Barker, DJ. (2001) Fetal programming and adult health. Public Health Nutr. 4: 611–624. | PubMed | ChemPort |
  2. Oken, E., Gillman, MW. (2003) Fetal origins of obesity. Obes Res. 11: 496–506. | PubMed | ISI |
  3. Ravelli, A. C. J., van der Meulen, J. H. P., Osmond, C., Barker, D. J. P., Bleker, OP. (1999) Obesity at the age of 50 years in men and women exposed to famine prenatally. Am J Clin Nutr. 70: 811–816. | PubMed | ISI | ChemPort |
  4. Yajnik, C. S., Fall, C. H. D., Coyaji, K. J., et al (2003) Neonatal anthropometry: the thin-fat Indian baby: The Pune maternal nutrition study. Int J Obes Relat Metab Disord. 27: 173–180. | Article | PubMed | ChemPort |
  5. Bavdekar, A., Yajnik, C. S., Fall, C. H., et al (1999) Insulin resistance syndrome in 8-year-old Indian children: small at birth, big at 8 years, or both? Diabetes 48: 2422–2429. | Article | PubMed | ISI | ChemPort |
  6. Remacle, C., Bieswal, F., Reusens, B. (2004) Programming of obesity and cardiovascular disease. Int J Obes Relat Metab Disord. 28: S46–S53. | Article | PubMed |
  7. Vickers, M. H., Breier, B. H., Cutfield, W. S., Hofman, P. L., Gluckman, PD. (2000) Fetal origins of hyperphagia, obesity, and hypertension and postnatal amplification by hypercaloric nutrition. Am J Physiol Endocrinol Metab. 279: E83–E87. | PubMed | ISI | ChemPort |
  8. Kwong, W. Y., Wild, A. E., Roberts, P., Willis, A. C., Fleming, TP. (2000) Maternal undernutrition during the preimplantation period of rat development causes blastocyst abnormalities and programming of postnatal hypertension. Development 127: 4195–4202. | PubMed | ISI | ChemPort |
  9. Langley-Evans, SC. (2000) Critical differences between two low protein diet protocols in the programming of hypertension in the rat. Int J Food Sci Nutr. 51: 11–17.
  10. Ozanne, S. E., Lewis, R., Jennings, B. J., Hales, CN. (2004) Early programming of weight gain in mice prevents the induction of obesity by a highly palatable diet. Clin Sci. 106: 141–145. | Article | PubMed | ISI | ChemPort |
  11. Ozanne, S. E., Hales, CN. (2004) Lifespan: catch-up growth and obesity in male mice. Nature 427: 411–412. | Article | PubMed | ISI | ChemPort |
  12. Desai, M., Gayle, D., Babu, J., Ross, MG. (2005) Programmed obesity in intrauterine growth-restricted newborns: modulation by newborn nutrition. Am J Physiol Regul Integr Comp Physiol. 288: R91–R96.
  13. Bieswal, F., Hay, S. M., McKinnon, C., et al (2004) Prenatal protein restriction does not affect the proliferation and differentiation of rat preadipocytes. J Nutr. 134: 1493–1499.
  14. Ruffin, M. P., Adage, T., Kuipers, F., Strubbe, J. H., Scheurink, A. J., van Dijk, G. (2004) Feeding and temperature responses to intravenous leptin infusion are differential predictors of obesity in rats. Am J Physiol Regul Integr Comp Physiol. 286: R756–R763. | PubMed | ISI | ChemPort |
  15. Etherton, T. D., Thompson, E. H., Allen, CE. (1977) Improved techniques for studies of adipocyte cellularity and metabolism. J Lipid Res. 18: 552–557.
  16. Chomczynski, P., Sacchi, N. (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 162: 156–159. | Article | PubMed | ISI | ChemPort |
  17. Ailhaud, G., Guesnet, P. (2004) Fatty acid composition of fats is an early determinant of childhood obesity: a short review and an opinion. Obes Rev. 5: 21–26. | Article | PubMed | ChemPort |
  18. Tsujikawa, M., Kimura, S. (1980) Effect of early feeding on cellularity of rat adipose tissue. J Nutr Sci Vitaminol. 26: 475–482.
  19. Dugail, I., Quignard-Boulange, A., Dupuy, F. (1986) Role of adipocyte precursors in the onset of obesity induced by overfeeding in suckling rats. J Nutr. 116: 524–535.
  20. Balonan, L. C., Sheng, HP. (2000) Perinatal feedings adversely affect lipogenic activities but not glucose handling in adult rats. Pediatr Res. 48: 668–673. | PubMed |
  21. Jones, A. P., Simson, E. L., Friedman, MI. (1984) Gestational undernutrition and the development of obesity in rats. J Nutr. 114: 1484–1492.
  22. Dahri, S., Snoeck, A., Reusens-Billen, B., Remacle, C., Hoet, JJ. (1991) Islet function in offspring of mothers on low protein diet during gestation. Diabetes 40: 115–120. | PubMed | ISI | ChemPort |
  23. Petry, C. J., Ozanne, S. E., Wang, C. L., Hales, CN. (1997) Early protein restriction and obesity independently induce hypertension in 1-year-old rats. Clin Sci. 93: 147–152. | PubMed | ISI | ChemPort |
  24. Bouret, S. G., Draper, S. J., Simerly, RB. (2004) Trophic action of leptin on hypothalamic neurons that regulate feeding. Science 304: 108–110. | Article | PubMed | ISI | ChemPort |
  25. Bouret, S. G., Simerly, RB. (2004) Minireview: leptin and development of hypothalamic feeding circuits. Endocrinology 145: 2621–2626. | Article | PubMed | ISI | ChemPort |
  26. Plagemann, A., Harder, T., Rake, A., et al (1999) Observations on the orexigenic hypothalamic neuropeptide Y-system in neonatally overfed weanling rats. J Neuroendocrinol. 11: 541–546. | Article | PubMed | ChemPort |
  27. Plagemann, A., Harder, T., Rake, A., et al (1999) Perinatal elevation of hypothalamic insulin, acquired malformation of hypothalamic galaninergic neurons, and syndrome x-like alterations in adulthood of neonatally overfed rats. Brain Res. 836: 146–155. | Article | PubMed | ISI | ChemPort |
  28. Plagemann, A., Harder, T., Rake, A., Melchior, K., Rohde, W., Dorner, G. (2000) Hypothalamic nuclei are malformed in weanling offspring of low protein malnourished rat dams. J Nutr. 130: 2582–2589. | PubMed | ISI | ChemPort |
  29. Garofano, A., Czernichow, P., Breant, B. (1999) Effect of ageing on beta-cell mass and function in rats malnourished during the perinatal period. Diabetologia 42: 711–718. | PubMed |
  30. Merezak, S., Reusens, B., Renard, A., et al (2004) Effect of maternal low protein diet and taurine on the vulnerability of adult Wistar rat islets to cytokines. Diabetologia 47: 669–675.
  31. Hales, C. N., Desai, M., Ozanne, S. E., Crowther, NJ. (1996) Fishing in the stream of diabetes: from measuring insulin to the control of fetal organogenesis. Biochem Soc Trans. 24: 341–350. | PubMed |
  32. Ozanne, S. E., Wang, C. L., Coleman, N., Smith, GD. (1996) Altered muscle insulin sensitivity in the male offspring of protein-malnourished rats. Am J Physiol. 271: E1128–E1134.
  33. Ozanne, S. E., Nave, B. T., Wang, C. L., Shepherd, P. R., Prins, J., Smith, GD. (1997) Poor fetal nutrition causes long-term changes in expression of insulin signaling components in adipocytes. Am J Physiol. 273: E46–E51. | PubMed |
  34. Petry, C. J., Dorling, M. W., Pawlak, D. B., Ozanne, S. E., Hales, CN. (2001) Diabetes in old male offspring of rat dams fed a reduced protein diet. Int J Exp Diabetes Res. 2: 139–143.
  35. Lucas, A., Baker, B. A., Desai, M., Hales, CN. (1996) Nutrition in pregnant or lactating rats programs lipid metabolism in the offspring. Br J Nutr. 76: 605–612. | Article | PubMed | ChemPort |
  36. Guan, H., Arany, E., van Beek, J. P., et al (2005) Adipose tissue gene expression profiling reveals distinct molecular pathways that define visceral adiposity in the offspring of maternal protein restricted rats. Am J Physiol Endocrinol Metab. 288: E663–E73A. | Article | PubMed | ISI | ChemPort |
  37. Girard, J., Perdereau, D., Foufelle, F., Prip-Buus, C., Ferre, P. (1994) Regulation of lipogenic enzyme gene expression by nutrients and hormones. FASEB J. 8: 36–42.
  38. Issad, T., Coupe, C., Pastor-Anglada, M., Ferre, P., Girard, J. (1988) Development of insulin-sensitivity at weaning in the rat: role of the nutritional transition. Biochem J. 251: 685–690.
  39. Mertens, I., Van Gaal, LF. (2002) Obesity, haemostasis and the fibrinolytic system. Obes Rev. 3: 85–101. | Article | PubMed | ChemPort |
  40. Skurk, T., Hauner, H. (2004) Obesity and impaired fibrinolysis: role of adipose production of plasminogen activator inhibitor-1. Int J Obes Relat Metab Disord. 28: 1357–1364. | Article | PubMed | ChemPort |
  41. Gottschling-Zeller, H., Aprath, I., Skurk, T., Hauner, H. (2000) Beta-Adrenoceptor agonists and other cAMP elevating agents suppress PAI-1 production of human adipocytes in primary culture. Horm Metab Res. 32: 509–514. | PubMed |
  42. Petry, C. J., Dorling, M. W., Wang, C. L., Pawlak, D. B., Ozanne, SE. (2000) Catecholamine levels and receptor expression in low protein rat offspring. Diabet Med. 17: 848–853.
  43. Ozanne, S. E., Wang, C. L., Dorling, M. W., Petry, CJ. (1999) Dissection of the metabolic actions of insulin in adipocytes fromearly growth-retarded male rats. J Endocrinol. 162: 313–319. | PubMed |
  44. Ozanne, S. E., Dorling, M. W., Wang, C. L., Petry, CJ. (2000) Depot-specific effects of early growth retardation on adipocyte insulin action. Horm Metab Res. 32: 71–75.
  45. Goldstein, B. J., Scalia, R. (2004) Adiponectin: a novel adipokine linking adipocytes and vascular function. J Clin Endocrinol Metab. 89: 2563–2568. | Article | PubMed | ISI | ChemPort |
  46. Matsuda, M., Shimomura, I., Sata, M., et al (2002) Role of adiponectin in preventing vascular stenosis: the missing link of adipo-vascular axis. J Biol Chem. 277: 37487–37491. | Article | PubMed | ISI | ChemPort |
  47. Langley-Evans, S. C., Langley-Evans, A. J., Marchand, MC. (2003) Nutritional programming of blood pressure and renal morphology. Arch Physiol Biochem. 111: 8–16. | Article | PubMed | ChemPort |
  48. Saint-Marc, P., Kozak, L. P., Ailhaud, G., Darimont, C., Negrel, R. (2001) Angiotensin II as a trophic factor of white adipose tissue: stimulation of adipose cell formation. Endocrinology 142: 487–492. | Article | PubMed | ChemPort |
  49. Ailhaud, G. (2002) Autocrine/paracrine effectors of adipogenesis. Ann Endocrinol. 63: 83–85.
Top

Acknowledgments

This work was supported by The Parthenon Trust, London, United Kingdom (University College London), by the National Foundation for Scientific Research (Belgium), by the European Union (Grant QLTR 200-00083), and by the Federal Scientific Policy of Belgium. W.D.R. was supported by the Scottish Executive Environment and Rural Affairs Department as part of the core funding to the Rowett Research Institute.

Extra navigation

.
ADVERTISEMENT