Aromatase-deficient (ArKO) mice accumulate excess adipose tissue

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Abstract

Aromatase is the enzyme which catalyses the conversion of C19 steroids into C18 estrogens. We have generated a mouse model wherein the Cyp19 gene, which encodes aromatase, has been disrupted, and hence, the aromatase knockout (ArKO) mouse cannot synthesise endogenous estrogens. We examined the consequences of estrogen deficiency on accumulation of adipose depots in male and female ArKO mice, observing that these animals progressively accrue significantly more intra-abdominal adipose tissue than their wildtype (WT) litter mates, reflected in increased adipocyte volume and number. This increased adiposity was not due to hyperphagia or reduced resting energy expenditure, but was associated with reduced spontaneous physical activity levels, reduced glucose oxidation, and a decrease in lean body mass. Elevated circulating levels of leptin and cholesterol were present in 1-year-old ArKO mice compared to WT controls, as were elevated insulin levels, although blood glucose was unchanged. Associated with these changes, the livers of ArKO animals were characterised by a striking accumulation of lipid droplets. Our findings demonstrate an important role for estrogen in the maintenance of lipid homeostasis in both males and females.

Introduction

Estrogen plays an integral role in lipid homeostasis and adipose tissue deposition and distribution. The sexually dimorphic distribution of adipose tissue in humans has implicated sex steroids in the regulation of adiposity and distribution of fat depots [1], [2]. Thus, whereas premenopausal women tend to have a lower body or gynoid distribution of fat, men and postmenopausal women tend to have an upper body or android distribution of fat [3], [4]. This phenotype is associated with a greater risk of insulin-resistant diabetes, cardiovascular disease and breast cancer [5], [6]. Estrogen insufficiency is thought to be largely responsible for the increase in adiposity during menopause since postmenopausal women who receive estrogen replacement therapy do not display the characteristic abdominal weight gain pattern usually associated with menopause [7], [8]. The role that estrogens play in lipid metabolism in the body is also highlighted by the fact that individuals of both sexes with natural mutations of the gene encoding aromatase, the enzyme responsible for estrogen biosynthesis, develop truncal obesity, insulin resistance, hypercholesterolemia and hypertriglyceridemia [9], [10], [11], [12].

We have recently developed a mouse model of estrogen insufficiency by targeted disruption of the aromatase gene (the ArKO mouse) [13]. Aromatase is encoded by the Cyp19 gene and catalyses the final step in the biosynthesis of C18 estrogens from C19 steroids. Hence, we have further investigated the relationship between estrogen and fat by exploring the phenotype of our estrogen-deficient mouse model. In the course of these studies, we observed that the animals displayed a progressive increase in adiposity as compared to wildtype (WT) litter mates. The aim of the present investigation was to characterise the obese phenotype of these animals in the expectation that this would provide new evidence for the role of estrogens in lipid homeostasis.

Section snippets

Mice

The ArKO mice were generated by disrupting the Cyp19 gene as previously been described [13]. Heterozygous males and females were bred to produce WT and homozygous null offspring. Mice were genotyped by PCR as described [14]. Animals were maintained under SPF conditions and had unlimited access to drinking water and a mouse diet containing 15% of calories as fat, 20% of calories as protein and 65% of calories as carbohydrate (manufactured by Glen Forrest Stockfeeders, Western Australia) [15].

Tissue collection and histology

Body and fat pad masses

Adult female ArKO mice were significantly heavier and had significantly larger gonadal and infra-renal fat pads than WT litter mates from 3 months of age onwards (Table 1). Male ArKO mice also had significantly heavier gonadal fat pads than WT males from 3 months of age and heavier infra-renal fat pads from 4 months of age (Table 1). However, their body weight failed to show a significant increase until 1 year of age. Administration of exogenous 17β-estradiol for 21 days to 7-week-old female

Discussion

The present studies reinforce and extend the concept that estrogens play an important role in the regulation of adiposity as a function of age in both males and females [21]. The data presented here has shown that ArKO mice of both sexes with estrogen insufficiency develop a progressive increase in adiposity, primarily as an accumulation of intra-abdominal fat. This excess fat accretion was associated with an increase in adipocyte volume and number, hyperleptinemia, hyperinsulinemia, and

Acknowledgements

This work is supported by USPHS Grant #R37-AG 08174, Grant #981126 from the NHMRC, and by the Victorian Breast Cancer Research Consortium.

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