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Endocrinology, doi:10.1210/en.2007-0770
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Endocrinology Vol. 148, No. 12 6157-6163
Copyright © 2007 by The Endocrine Society

Increased Maternal Nutrition Increases Leptin Expression in Perirenal and Subcutaneous Adipose Tissue in the Postnatal Lamb

B. S. Muhlhausler, J. A. Duffield and I. C. McMillen

Early Origins of Adult Health Research Group (B.S.M., I.C.M.), Sansom Research Institute, School of Pharmacy and Medical Sciences, The University of South Australia, Adelaide 5001, Australia; and Discipline of Physiology (J.A.D.), School of Molecular and Biomedical Science, The University of Adelaide, Adelaide 5005, Australia

Address all correspondence and requests for reprints to: Dr. Beverly Muhlhausler, Early Origins of Adult Health Research Group, Sansom Research Institute, University of South Australia, Adelaide 5000, Australia. E-mail: beverly.muhlhausler{at}unisa.edu.au.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
The present study tested the hypothesis that exposure to an increased level of maternal nutrition before birth results in altered expression of adipogenic, lipogenic, and adipokine genes in adipose tissue in early postnatal life. Pregnant ewes were fed either at or approximately 50% above maintenance energy requirements during late pregnancy, and quantitative RT-PCR was used to measure peroxisome proliferator-activated receptor (PPAR)-{gamma}, lipoprotein lipase (LPL), glycerol-3-phosphate-dehydrogenase (G3PDH), adiponectin, and leptin mRNA expression in perirenal (PAT) and sc adipose tissue (SCAT) in the offspring on postnatal d 30. Relative SCAT mass was higher in lambs of well-fed ewes (40.0 ± 4.0 vs. 22.8 ± 3.3 g/kg, P < 0.05) and was directly related to plasma insulin in the first 24 h after birth and to G3PDH and LPL expression. The expression of leptin mRNA in both the SCAT and PAT depots was higher (P < 0.05) in lambs of well-fed ewes. PPAR{gamma} adiponectin, LPL, and G3PDH mRNA expression were not, however, different between well-fed and control groups in either depot. Relative PPAR{gamma} expression in SCAT was directly related to plasma insulin concentrations in the first 24 h after birth (r2 = 0.23; P < 0.05), and G3PDH and LPL expressions were also positively correlated with PPAR{gamma} expression (r2 = 0.27; P < 0.05). We have demonstrated that exposure to increased prenatal nutrition increases leptin expression at 1 month of age in both PAT and SCAT. The results of this study provide evidence that the nutritional environment before and immediately after birth can influence the development of adipose tissue in early postnatal life.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
IN PREGNANCIES COMPLICATED by maternal diabetes, the fetus is hyperglycemic and hyperinsulinemic, and cord blood leptin concentrations increase in parallel with increased infant adiposity (1, 2). In addition, maternal diabetes mellitus, gestational diabetes, or even mild impairments of glucose tolerance during pregnancy are all risk factors for the development of obesity and type 2 diabetes in the offspring (3, 4, 5). It has been proposed that the programming of the expression of adipogenic and lipogenic genes within the adipocyte may be one mechanism through which exposure to an excess nutrient supply during critical windows of fetal development may result in increased adiposity in adult life (6).

In adults, changes in the patterns of expression of key regulatory and functional genes within adipose tissue are important determinants of fat distribution and accumulation (7). There is considerable evidence that the CCAAT/enhancer-binding protein-{alpha} and peroxisome proliferator-activated receptor (PPAR)-{gamma} serve as pleiotropic transcriptional activators that coordinately induce expression of a suite of adipocyte-specific genes, resulting in the differentiation of adipose cells (8, 9). Furthermore, PPAR{gamma} also regulates adipose tissue mass and the expression of genes involved in signaling of adipose cells to other peripheral tissues. Specifically, PPAR{gamma} ligands act to decrease the expression of leptin and increase the expression of the insulin sensitizing hormone, adiponectin, within adipose tissue (10). They also act to regulate the expression of genes involved in increasing storage of triglycerides within adipose cells, including lipoprotein lipase (LPL) and glycerol-3-phosphate dehydrogenase (G3PDH) (11, 12).

We previously reported that exposure to increased maternal nutrition before birth results in increased sc fat deposition at 30 d of postnatal life in the lamb, a species in which adipose tissue is deposited before birth, as in the human (13). In the sheep, the sc adipose depot develops predominantly in postnatal life, whereas the perirenal adipose depot is deposited before birth (14), and we have recently shown that exposure to maternal overnutrition is associated with increased expression of PPAR{gamma}, LPL, and leptin mRNA in perirenal adipose tissue in the fetal sheep in late gestation (15). Whereas it therefore appears that prenatal overnutrition can increase the expression of genes that regulate adipogenesis and lipogenesis before birth, it is unknown whether such changes in gene expression persist beyond the prenatal period and influence patterns of gene expression and development of the sc adipose depot in postnatal life.

In the present study, we therefore used the lamb as an experimental model to test the hypothesis that exposure to an increased nutrient supply before birth results in increased expression of adipogenic, lipogenic, and adipokine genes (PPAR{gamma}, leptin, adiponectin, LPL, and G3PDH) in the perirenal and sc adipose depots at 30 d of postnatal life.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Animals and feeding regimen
All procedures were approved by the University of Adelaide Animal Ethics Committee. Merino ewes were mated and pregnancy confirmed by ultrasound scanning in early gestation. From 90 d gestation, ewes were individually housed and acclimatized to a control diet consisting of 1 kg Lucerne chaff [85% dry matter, metabolizable energy (ME) content = 8.3 MJ/kg] and 300 g concentrated pellets (89% dry matter, ME content = 11.6 MJ/kg; Ridley Agriproducts Sheep Nut Ration, Murray Bridge, South Australia, Australia). The diet was calculated to provide 100% of the energy requirements for the maintenance of a pregnant ewe bearing a singleton or twin fetuses as appropriate, as specified by the Ministry of Agriculture, Fisheries, and Food (United Kingdom) (16). At 115 d gestation, i.e. before the commencement of the rapid fetal growth phase in late gestation (17), ewes were randomly assigned to either a control (n = 8) or well-fed group (n = 8). Between 115 and 124 d gestation, control ewes were provided with 14.0 ± 0.4 g of Lucerne chaff and 6.5 ± 0.4 g of pelleted concentrate per kilogram body weight, and well-fed ewes were provided with 22.1 ± 0.8 g Lucerne chaff and 10.4 ± 0.7 g pelleted concentrate per kilogram body weight each day. The feed allowance of all ewes was proportionately increased by 15% every 10 d to meet the increasing substrate demands of the growing fetus in late gestation (16). All feed refusals were recorded and weighed daily and used to calculate the ME intake of all ewes as a percentage of their maintenance energy requirements (%MER). Maternal ME intake was significantly higher in well-fed ewes, compared with control ewes, across late gestation (control, 91.9 ± 2.2% MER; well fed, 133.4 ± 3.5% MER, P < 0.01) (13).

Before lambing, food was provided to each ewe in two equal portions each day, one at 0900 h and one at 1500 h, with water provided ad libitum. After lambing, all ewes were provided with the same diet, which consisted of 1 kg Lucerne chaff and 500 g pelleted concentrate once daily. If all feed was consumed before 1500 h, an additional 1 kg of chaff was provided. After birth, each ewe and her lamb(s) were housed in an individual pen in an indoor housing facility, which was maintained at a constant ambient temperature of between 20 and 22 C and a 12-h light, 12-h dark cycle.

Growth rate measures and blood sampling
Lambs were born spontaneously at term (150 ± 3 d gestation). The control group consisted of six males and six females (eight twins, four singletons), and the well-fed group consisted of three males and six females (two twins, seven singletons). The day of birth was designated as d 1. Birth weight (kilograms), crown rump length (centimeters), and abdominal circumference (centimeters) were recorded within 6 h of birth.

Body weight (kilograms), crown rump length (centimeters), and abdominal circumference (centimeters) were recorded each day for the first 5 d of life and at least once every 2 d thereafter until d 30. All measurements were obtained between 0800 and 1100 h. Venous blood samples were collected in chilled tubes between 0800 and 1300 h after a 2-h fast on d 1–5 and every 3 d thereafter until postnatal d 30. The first blood sample was collected within 6 h of birth. All blood samples were centrifuged at 1500 x g for 10 min and plasma separated into aliquots and stored at –20 C.

Postmortem and tissue collection
At 30 d of age, lambs were killed with an overdose of sodium pentobarbitone (Virbac Pty. Ltd., Peakhurst, New South Wales, Australia). A sample of sc adipose tissue from the thoracic region of the carcass was immediately collected and snap frozen in liquid N2. All organs were dissected out and individually weighed. All adipose tissue from the perirenal, omental, pericardial, pelvic, epididymal/parametrial, and axillary fat depots was carefully excised and weighed. One sample of perirenal and sc adipose tissue was fixed in 4% paraformaldehyde for subsequent processing and histological analyses. A second sample from these sites was snap frozen in liquid N2 and stored at –80 C for subsequent determination of gene expressions by quantitative RT-PCR (qRT-PCR).

Plasma glucose, insulin, leptin, and nonesterified fatty acid (NEFA) assays
Plasma glucose concentrations were measured by enzymatic analysis using hexokinase and glucose-6-phosphate dehydrogenase to measure the formation of nicotinamide adenine dinucleotide hydroxide photometrically at 340 nm (COBAS MIRA automated analysis system; Roche Diagnostica, Basel, Switzerland). The sensitivity of the assay was 0.5 mmol/liter, and the intra- and interassay coefficients of variation were both less than 5%. Plasma NEFAs were measured using the Wako NEFA kit (Wako, Osaka, Japan) as previously described (13).

Plasma insulin concentrations were measured using a RIA (rat insulin kit; Linco Research, Inc., St. Charles, MO), which was validated for use with sheep plasma (13). The sensitivity of the assay was 0.01 ng/ml, and the intra- and interassay coefficients of variance were both less than 10%.

Plasma leptin concentrations were measured using a competitive bovine leptin ELISA, which has previously been validated for sheep plasma (18). The sensitivity of the assay was 0.5 ng/ml, and the intra- and interassay coefficients of variation were less than 16%.

RNA extraction
RNA from perirenal and sc adipose tissue (~600 mg) was isolated using Trizol reagent (Invitrogen Australia Pty. Ltd., Mount Waverley, Australia) and chloroform. RNA was treated for genomic DNA contamination using Ambion DNase1, and after enzyme deactivation, the RNA was run through a secondary purification process using the RNeasy minikit (QIAGEN Pty. Ltd. Australia, Doncaster, Australia). The quality and concentration of the RNA was determined by measuring the absorbance at 260 and 280 nm, and RNA integrity was confirmed by agarose gel electrophoresis. cDNA was then synthesized using the purified RNA (5 µg) and SuperScript 3 reverse transcriptase (Invitrogen Australia) and random hexamers.

qRT-PCR
The relative expression of PPAR{gamma}, leptin, adiponectin, LPL, and G3PDH mRNA transcripts was measured by qRT-PCR using the Sybr Green system in an ABI Prism 7000 or 7300 sequence detection system (PE Applied Biosystems, Foster City, CA). All primers have been validated previously for use in sheep tissues (15). For the qRT-PCR measurements, the primer concentrations were consistent for all genes, and the amplification efficiency of all primers was 0.997–0.999. A constant amount of cDNA (1 µl) was used for each qRT-PCR measurement and at least three technical replicates were performed for each gene.

Each qRT-PCR well (10 µl total volume) contained 2.5 µl 2x Sybr Green master mix (Applied Biosystems); 0.25–0.5 µl of each primer, giving a final concentration of 450 or 900 nM; 1.0 µl of molecular grade H2O; and 1.0 µl of a 50 ng/µl dilution of the stock template. The cycling conditions consisted of 40 cycles of 95 C for 15 min and 60 C for 1 min. At the end of each run, dissociation melt curves were obtained.

The abundance of each mRNA transcript was measured and expression relative to that the reference gene, cyclophilin, was calculated using Q-gene qRT-PCR analysis software, which provides a quantitative measure of the relative abundance of a specific transcript in different tissues by the comparative threshold cycle method. The threshold cycle value was taken as the lowest statistically significant (>10 SD) increase in fluorescence above the background signal in an amplification reaction. This procedure takes into account any differences in the amplification efficiencies of the target and reference genes.

Statistical analyses
All data are presented as the mean ± SEM. Multifactorial ANOVA was used to determine the main effects of maternal nutritional treatment, gender, and fetal number and their interaction on birth weight, fat mass, and adipocyte gene expression. There was no effect of fetal number on any measure.

Two-way ANOVA was therefore used to determine the main effects of maternal nutritional treatment (control or well fed) and gender and their interaction on birth weight and fat mass. ANOVA was also used to determine the main effects of maternal nutritional treatment (control or well fed), adipose depot (perirenal or sc) and gender and their interaction on adipocyte gene expression. Where there was no effect of gender, data from male and female lambs were combined for subsequent analyses. Simple linear regression analyses were used to determine relationships between variables. Plasma glucose, insulin, and leptin concentrations across postnatal wk 1–4 were averaged for correlation analyses unless stated otherwise. Partial correlation analysis was used to control for the effects of maternal nutrient intake and mean plasma glucose levels as appropriate. A probability of 5% (P < 0.05) was taken as the level of significance in all analyses.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Increased maternal nutrition and fat cell size in perirenal and sc adipose tissue
There was no effect of maternal overnutrition on the growth rate of lambs over the first month of postnatal life (Fig. 1Go). Relative sc, but not perirenal, fat mass was significantly higher in the well-fed lambs, compared with controls, and relative total fat mass tended (P = 0.06) to be higher in the well-fed lambs (Table 1Go). There was no effect of maternal overnutrition on the relative weight of the perirenal, omental, epidymal/parametrial, or percardial fat depots at 30 d of postnatal age (Table 1Go). The mean size of sc adipose cells, but not perirenal adipose cells, also tended (P = 0.06) to be greater in the well-fed group, compared with controls (control, 1801 ± 195 µm2; well fed, 2405 ± 202 µm2), and the mean adipocyte size in each fat depot was directly related to the relative mass of the corresponding fat depot when data from all lambs were combined (perirenal fat: r2 = 0.59; P < 0.01, n = 21; sc fat: r2 = 0.57; P < 0.01, n = 21). The mean size of adipocytes in the perirenal and sc depots were directly related when the data from all lambs were combined (r2 = 0.35; P < 0.01, n = 21).


Figure 1
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  		FIG. 1. Body weight gain in control (open circles) and well-fed (closed circles) groups in the first month of postnatal life.

 

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  		TABLE 1. The effect of increased maternal nutrition on the mass of sc, perirenal, omental, pericardial, and parmetrial/epidyimal fat and total fat mass at 30 d of postnatal age

 
Metabolic and hormonal regulation of fat deposition
The mean size of perirenal and sc adipocytes were each directly related to insulin concentrations in the first 24 h after birth when data from all lambs were combined (Table 2Go). When data from all lambs were combined, there was a direct relationship between relative sc fat mass and plasma insulin concentrations in the first 24 h after birth (Table 2Go), and this relationship remained significant after controlling for the effects of mean plasma glucose concentrations in early postnatal life. Plasma glucose, but not NEFA or leptin concentrations, on postnatal d 1 were higher in lambs of well-fed ewes. Plasma concentrations of insulin in the first 24 h after birth also tended (P = 0.07) to be higher in the well-fed group (Table 3Go). There were no relationships between relative sc fat mass or size of perirenal and sc adipocytes and insulin concentrations on postnatal d 30. There was also no effect of maternal nutrition on plasma glucose, insulin leptin, or NEFA concentrations on postnatal d 30 (Table 3Go).


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  		TABLE 2. Summary of significant relationships between measures of adipose mass, mean adipocyte size, and sc PPAR{gamma} mRNA expression with insulin concentrations in the first 24 h after birth

 

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  		TABLE 3. Plasma concentrations of glucose, insulin, leptin, and NEFA in lambs of control and well-fed ewes on d 1 and 30 of postnatal age

 
Leptin mRNA expression and circulating plasma leptin concentrations
The expression of leptin mRNA in both sc and perirenal adipose tissue was significantly higher in the well-fed group, compared with controls, independent of gender (Fig. 2Go, A and B). The expression of leptin mRNA was higher in the sc, compared with the perirenal, depot independent of the level of maternal nutrition (Fig. 2Go, A and B), and leptin mRNA expression in the sc adipose depot was directly correlated with relative sc fat mass (r2 = 0.38; P < 0.01, n = 21). Leptin mRNA expression in the sc adipose depot was directly related to the mean size of the perirenal adipocytes, but was not related to mean size of the sc adipocytes cells (r2 = 0.33; P < 0.01, n = 21).


Figure 2
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  		FIG. 2. The relative expression of leptin mRNA in perirenal (A) and sc (B) adipose depots in control and well-fed groups at postnatal d 30. Asterisks denote a significant effect of maternal nutrition (P < 0.05).

 
There was a direct relationship between the relative expression of leptin mRNA in sc, but not perirenal, adipose tissue and plasma leptin concentrations on postnatal d 30 (r2 = 0.37; P < 0.01, n = 19). Plasma leptin concentrations on the day of postmortem were directly related to total relative fat mass when data from all lambs were combined (r2 = 0.24; P < 0.04, n = 19).

Expression of adipogenic and lipogenic genes in perirenal and sc adipose tissue
The expression of PPAR{gamma}, adiponectin, and LPL mRNA were significantly higher in the perirenal, compared with the sc, adipose tissue, independent of the level of maternal nutrition (Table 4Go), whereas the expression of G3PDH mRNA was significantly higher in the sc, compared with the perirenal, adipose depot (Table 4Go). There was no significant effect of maternal nutrition or gender on the relative expression of PPAR{gamma}, adiponectin, G3PDH, or LPL mRNA in either the perirenal or sc adipose depot (Table 4Go).


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  		TABLE 4. The effect of increased maternal nutrition on the expression of PPAR{gamma}, G3PDH, adiponectin, and LPL mRNA in the perirenal and sc adipose depots

 
PPAR{gamma} and adiponectin mRNA expression
The relative expression of adiponectin mRNA in the perirenal and sc adipose depots was directly related to the expression of PPAR{gamma} in the same depot when data from all lambs were combined (Fig. 3Go, A and B). The relative expression of adiponectin mRNA in the perirenal adipose tissue was also directly related to mean fractional growth rate across the first 4 wk of postnatal life (r2 = 0.25; P < 0.05, n = 21).


Figure 3
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  		FIG. 3. The relationship between the relative expression of PPAR{gamma} and adiponectin mRNA at 30 d of postnatal age in perirenal (A) and sc (B) adipose tissue. There was a significant positive relationship between the relative expression of PPAR{gamma} and adiponectin mRNA in both the perirenal (r2 = 0.49, P < 0.01) and sc (r2 = 0.23, P < 0.05) adipose tissue when data from all lambs were combined. Open circles, Control; closed circles, well fed.

 
PPAR{gamma}, LPL, and G3PDH mRNA expression and sc fat mass
PPAR{gamma} mRNA expression in sc, but not perirenal, adipose tissue was directly related to plasma insulin concentrations in the first 24 h after birth when data from all lambs were combined (Table 2Go). When the data from all lambs were combined, relative sc fat mass was directly related to either LPL or G3PDH mRNA expression in sc adipose tissue (G3PDH; r2 = 0.19; P < 0.05, n = 21; LPL: r2 = 0.27; P < 0.04, n = 21); however, these relationships were not present in the perirenal adipose depot. In the sc adipose depot, but not the perirenal adipose depot, there was also a direct relationship between the expression of LPL mRNA and mean adipocyte size (r2 = 0.27; P < 0.04, n = 21). These relationships were no longer significant when the effects of insulin concentrations in the first 24 h after birth were controlled for.

Expression of PPAR{gamma} mRNA in the sc adipose depot was directly related to the mean size of perirenal adipocytes but not to the mean size of the sc adipocytes (r2 = 0.23; P < 0.03, n = 21).

In the sc adipose depot there was a direct relationship between the relative expression of PPAR{gamma} mRNA and relative expression of G3PDH mRNA (r2 = 0.27; P < 0.05, n = 21), LPL mRNA (LPL r2 = 0.18; P = 0.05, n = 21), and leptin mRNA (r2 = 0.35; P < 0.05, n = 21) when data from all lambs were combined. In the perirenal adipose depot, PPAR{gamma} mRNA expression was directly related to the expression of G3PDH (r2 = 0.55, P < 0.01, n = 21) but was not related to the expression of LPL or leptin mRNA in this depot.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
In the present study, exposure to increased maternal nutrition resulted in a significant increase in the expression of leptin mRNA in both perirenal and sc adipose tissue after birth. This appeared to be a consequence of increased accumulation of lipid because there was a direct relationship between expression of leptin mRNA abundance and relative sc fat mass. Leptin mRNA expression in the sc depot, but not perirenal depot, was also directly related to circulating plasma leptin concentrations on the day of postmortem, which suggests that the sc adipose depot is the principal source of circulating plasma leptin concentrations in the sheep in early neonatal life, as in the human (19). We have previously shown that in this same group of animals, that the expression of the leptin receptor within the arcuate nucleus of the hypothalamus decreased with increasing fat mass in lambs of well-fed, but not control ewes, and that the lambs of well-fed ewes failed to appropriately down-regulate the expression of the key appetite-inhibiting neuropeptide, cocaine- and amphetamine-regulated transcript, with increasing fat mass, consistent with the development of leptin resistance with increasing adiposity (13). The results of the present study suggest that an increase in leptin expression in adipose tissue in response to maternal overnutrition, which may subsequently be associated with an increased leptin secretion from adipose tissue, may be the primary event that leads to down-regulation of hypothalamic leptin receptors and the development of leptin resistance within hypothalamic neurons.

In the present study, we found that the relative mass of sc fat and the size of perirenal and sc adipocytes were each directly related to plasma insulin concentrations, independent of plasma glucose concentrations in the first 24 h after birth. The accumulation of adipose tissue in the sc depot is also a characteristic feature of infants of pregnancies complicated by maternal diabetes or glucose intolerance, and the extent of this accumulation in early postnatal life can be related directly to plasma glucose and insulin concentrations in the maternal, and hence fetal, circulation during late pregnancy (20, 21, 22).

Whereas there was no increase in PPAR{gamma} expression in sc adipose tissue in the well-fed group, PPAR{gamma} mRNA expression was positively correlated with insulin concentrations in the first 24 h after birth when data from all lambs were combined. Importantly, this relationship appeared to be related to maternal nutritional status because it did not persist when the effects of maternal nutrient intake in late gestation were controlled for. In postnatal life, PPAR{gamma} promotes peripheral insulin sensitivity by up-regulating the expression of genes involved in the uptake of free fatty acids into adipose cells (e.g. LPL) or in the de novo synthesis of triglycerides (e.g. G3PDH), which increases lipid storage and thereby reduces circulating free fatty acid concentrations (11, 23). In the present study, we found that PPAR{gamma} expression in both perirenal and sc adipose tissue was directly related to G3PDH and/or LPL mRNA expression in the respective depot. Furthermore, in the sc adipose tissue, the expression of G3PDH mRNA and LPL mRNA was directly related to relative sc fat mass and cell size. It is therefore possible that the increased expression of PPAR{gamma} in response to higher plasma insulin concentrations in the first 24 h after birth may account, at least in part, for the increased sc fat deposition in lambs of well-fed ewes.

The expression of the insulin-sensitizing adipokine, adiponectin, in perirenal and sc adipose tissue was also directly related to the expression of PPAR{gamma}, from which it would appear that the role of PPAR{gamma} in mediating increases in adiponectin expression, is established early in life (10). The expression of adiponectin in the perirenal adipose depot was directly related to the mean fractional growth rate of lambs across the first 30 d of postnatal life, which may suggest that adiponectin can act to increase the sensitivity of growing tissues to the anabolic actions of insulin (24).

Perirenal and sc fat depots: evidence for cross talk in early postnatal life
In the current study, we found that the relative expression of PPAR{gamma}, adiponectin, and LPL mRNA, genes associated with adipogenesis and insulin sensitivity, were higher in perirenal adipose depot, whereas the expression of G3PDH and leptin mRNA, genes associated more with lipid storage, were higher in sc adipose depot. These differential patterns of gene expression in the visceral and sc fat depots are consistent with previous findings in adult humans and rodents (7, 25). This suggests that the distinct physiological roles of the perirenal and sc adipose cells in relation to the regulation of whole-body energy balance and insulin sensitivity are already present in early postnatal life.

We found that, in contrast to sc adipose tissue, there was no relationship between G3PDH mRNA expression and the mean size of perirenal adipocytes or the relative mass of perirenal adipose tissue. In the adult human and rodent, perirenal adipocytes have higher rates of lipolysis in the basal state, greater sensitivity to cathecholamine-induced lipolysis, and reduced sensitivity to the antilipolytic activity of insulin, compared with sc adipocytes (7, 25). One possible explanation, therefore, is that higher lipolytic or thermogenic activity within the perirenal adipose tissue during early postnatal life may counteract the G3PDH-mediated lipogenesis in this depot. An alternate possibility may be that the activation of uncoupling protein 1 in perirenal adipose tissue, which occurs immediately after delivery in the sheep, may act to limit the capacity for accumulation of lipid in this depot in early postnatal life (26).

An intriguing finding of the present study was that the expression of PPAR{gamma} and leptin mRNA in the sc adipose depot was related directly to the mean size of the perirenal, but not sc, adipocytes. We recently demonstrated that exposure to increased prenatal nutrition is associated with a significant increase in the expression of genes that regulate adipogenesis (PPAR{gamma}) and lipogenesis (LPL) in the perirenal adipose tissue before birth (15). Whereas the perirenal adipose depot develops before birth, the sc adipose depot develops predominantly in postnatal life (14), and the findings of the present study may therefore provide evidence of cross talk between the perirenal and sc adipose depots during in the early postnatal period. One possible explanation, and one that clearly warrants further study, is that paracrine/endocrine signals from perirenal adipose tissue may have the potential to regulate gene expression in sc adipocytes and therefore dictate the progression of sc fat deposition in the immediate postnatal period.

Summary
In summary, we have demonstrated that exposure to increased maternal nutrition in late gestation results in increased sc fat deposition in lambs in early postnatal life, which may be a consequence of an increased accumulation of lipid within individual sc adipocytes. Furthermore, we have reported that expression of leptin mRNA in perirenal and sc adipose tissue at 1 month of postnatal age is increased in lambs exposed to prenatal overnutrition. We have also presented evidence that may suggest that changes within the perirenal adipose depot, which develops before birth, may have the potential to regulate gene expression and therefore development of the sc adipose depot, which develops predominantly in postnatal life (14). The results of the present study provide important new insights into the mechanisms through which exposure to an increased nutrient supply before birth results in an increased accumulation of adipose stores, and ultimately obesity, in postnatal life.


   Acknowledgments
 
We gratefully acknowledge the support of Laura O’Carroll and Animal Technician staff of the University of Adelaide with assistance with animal protocols.


   Footnotes
 
This work was supported by a National Health and Medical Research Council Program Grant Award (to I.C.M.).

Disclosure Statement: The authors have nothing to disclose.

First Published Online September 20, 2007

Abbreviations: G3PDH, Glycerol-3-phosphate dehydrogenase; LPL, lipoprotein lipase; ME, metabolizable energy; %MER, maintenance energy requirements; NEFA, nonesterified fatty acid; PPAR, peroxisome proliferator-activated receptor; qRT-PCR, quantitative RT-PCR.

Received June 11, 2007.

Accepted for publication September 12, 2007.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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