Does estradiol mediate leptin's effects on adiposity and body weight?

Mary Ann Pelleymounter, Mary Beth Baker, and Michael McCaleb

Department of Neuroscience, Amgen, Thousand Oaks, California 91320


    ABSTRACT
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

The role of estradiol in mediating leptin's effects on body weight was assessed in ovariectomized (OVX) mice before and after the onset of obesity. Ovariectomy did not alter leptin levels before the onset of obesity, and estradiol adminstration (0.05-17 µg/day for 14 days) did not significantly alter leptin levels if they were corrected for the estradiol-induced reduction in body fat. The converse was also true, in that leptin administration (0.4-140 µg/day) did not alter estradiol levels in intact mice. Furthermore, neither estradiol reduction (via ovariectomy) nor addition (via exogenous administration) significantly altered leptin's ability to reduce fat mass. Leptin was equally effective in reducing body weight in lean or obese OVX mice and intact controls. Finally, estradiol did not change the magnitude of leptin's effect on fat mass reduction when it was given in combination with leptin to lean intact or OVX mice. Estradiol may have indirectly affected leptin efficacy, because leptin did not produce as large a change in fat mass at lower doses in lean OVX mice as it did in intact counterparts. Taken together, these data suggested that 1) estradiol does not directly regulate leptin secretion or its effects on fat mass and 2) leptin does not directly regulate estradiol secretion or its effects on fat mass. Leptin and estradiol, however, may interact in an indirect fashion to affect fat utilization.

leptin; estradiol; ovariectomy; efficacy; body fat


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

LEPTIN is a 16-kDa protein that is secreted from adipose tissue in a pulsatile fashion (26, 42, 49). Exogenous administration of leptin results in weight loss, reduced adiposity, and appetite suppression (3, 14, 35). Although leptin level correlates well with body weight and adiposity (6, 9, 15, 22, 27, 29), the association between leptin level and body weight is not the same in both genders. Several groups have found that females have more leptin than males (19, 21, 34, 38, 39), even when 1) both groups are normalized for body fat (19, 21, 34, 38, 39); 2) lean males and females are compared (19); 3) children are used as subjects (17); or 4) venous cord blood samples from newborn males and females are compared (30).

One explanation for the gender difference in leptin levels is that ovarian hormones modulate leptin secretion and, hence, energy balance. The idea that ovarian hormone regulation could be linked to energy balance is an old one. Food deprivation, glucoprivation, and lipoprivation all can affect estrogen receptor number, estrus cycling, and lordosis (46). These manipulations also have negative effects on puberty onset and human fertility (10). Interestingly, leptin can counteract the effects of food deprivation on puberty onset in both males and females (1, 4). Furthermore, leptin levels are reduced in human females with amenorrhea due to anorexia (26). There is also evidence that the leptin receptor is localized on the ovary (5, 48). Finally, estradiol itself can have effects on food intake and body weight that are in the same direction as leptin, in that it causes a transient reduction in food intake and a moderate reduction in body weight in ovariectomized females (2, 33, 44, 46).

It is possible, then, that estrogen could be involved in the mechanism by which leptin regulates adiposity in females. Furthermore, if estradiol and leptin have similar effects on body mass and adiposity, it is possible that estradiol given in combination with leptin, or given as a pretreatment before leptin administration, could act to enhance the effects of leptin on adiposity in females. To test these hypotheses, we 1) assessed the effects of estradiol addition (continuous subcutaneous administration) or estradiol subtraction (ovariectomy) on leptin levels, 2) assessed leptin's ability to reduce body weight and fat mass in lean or obese ovariectomized (OVX) and intact mice, and 3) evaluated the effects of either simultaneous administration or pretreatment with estradiol on leptin's ability to reduce fat mass in lean and obese OVX mice.


    METHODS
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Experiment 1 (lean OVX mice). The importance of the ovary in mediating leptin's effects on body weight was tested in three ways. First, the effects of estradiol elevation were assessed by measuring serum leptin in 36 intact mice treated with 17beta -estradiol pellets that delivered either 0.0, 0.05, 0.5, 1.2, 2.4, or 17 µg/day for a period of 2 wk. The 0.0 (control) pellet contained the solid vehicle only. Second, the impact of ovarian hormone level reduction was tested by measuring leptin levels in PBS-treated OVX mice 4 wk after surgery. Third, the effects of ovariectomy on leptin efficacy were tested by comparing leptin's metabolic effects on OVX vs. intact mice. The comparison was done before the weight gain that eventually occurs in OVX rodents (2, 31, 33). Forty-two OVX and 42 sham-operated C57Bl6J mice (SHAM) (9 wk of age) were obtained from Charles River Laboratories. OVX and SHAM mice were subdivided into seven dose groups, with vehicle controls for both groups. Two weeks after surgery, 1) recombinant methionine murine leptin (r-MetMuLeptin; 0.03-12.6 mg/ml, or 0.4-140 µg/day), 2) a combination of 17beta -estradiol (17 µg/day sc pellets) and leptin (120 µg/day), or 3) PBS was infused subcutaneously by use of an osmotic minipump (Alzet 1007D) that delivered 0.5 µl/h. Leptin, the leptin-estradiol combination, or PBS was administered for 14 days.

Body weight was measured every day in these animals. At the end of the 2-wk period (day 15), blood was taken from the retroorbital sinus under isoflurane anesthesia, and the mice were euthanized. All carcasses were saved for composition analysis.

Experiment 2 (obese OVX mice). In the second experiment, 24 OVX mice were allowed to gain weight and were used as an alternative model of obesity in which to test leptin efficacy after a 4-wk pretreatment with estradiol. Estradiol (EST) or placebo (CON) treatment (17 µg/day sc pellet) began 15.5 wk after OVX or sham surgeries. Four weeks after the onset of hormone therapy, EST and CON groups were further subdivided into either PBS or leptin groups. r-MetMuLeptin (10 mg/ml) or PBS was administered via continuous subcutaneous infusion (Alza osmotic mini-pumps, 0.5 µl/h, 2001D) for 14 days. Body weight was measured daily. Blood was collected 2 days before the onset of leptin or PBS treatment and on the last day of leptin or PBS infusion. Mice were euthanized on the last day of leptin treatment, and carcasses were taken for composition analysis.

Mice from all experimental groups were individually housed in the Amgen vivarium, with the ambient temperature set at 21-23°C and a 12:12-h light-dark cycle in effect (6:30 AM on; 6:30 PM off). All mice were allowed both food and water ad libitum. The procedures described above were approved by the Laboratory Animal Research Committee at Amgen.

Carcass composition analysis. Carcass composition was assessed with the methods of Leshner et al. (24). Water composition was determined by subtraction of carcass weight before and after a 5-day dehydration period. Fat was extracted with ethyl ether and ethyl alcohol from a preweighed portion of the ground, dried carcass, so that percent fat could be calculated from the amount of material remaining after the extraction procedure. Lean mass was defined as the proportion of ground carcass that remained after dehydration and ether extraction.

Serum chemistry. Serum nonesterified free fatty acids (NEFA), cholesterol, beta -hydroxybutyric acid (BHBA), and triglycerides were analyzed using a Hitachi 717 blood chemistry analyzer. Corticosterone was measured using a competitive immunoassay system developed by Boehringer Mannheim Biochemicals (Indianapolis, IN). Leptin levels were measured using a solid phase sandwich enzyme immunoassay, with affinity-purified polyvalent antibody immobilized in microliter wells. Leptin was calculated from standard curves generated for each assay by use of recombinant mouse leptin. The detection limits of the assay were 70 pg/ml (38). Estradiol was measured using RIA technology on serum that was extracted on Sephadex LH20 columns by following the methods described by Goodman (12) and Resko et al. (37).

Statistics. Repeated-measures ANOVA was used for all comparisons between intact and OVX groups over time, with least squares analysis used as the post hoc test for dissecting interaction terms. In the case of single time-point data, 1-, 2-, or 3-way ANOVA was chosen as the analysis method for main effects, with Fisher's least significant difference test used as the post hoc test. Simple regression was used to correlate leptin levels with carcass fat in estradiol-treated mice, and half-maximal effective dose curves were fitted using a third-order sigmoid equation.


    RESULTS
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Experiment 1 (lean OVX mice). Estradiol administration reduced serum leptin in a dose-dependent manner [F(5,29) = 8.01; P < 0.0001] (Fig. 1A), but the reduction correlated significantly with the effects of estradiol on carcass fat (r = 0.84; P < 0.0001), which can be observed in Fig. 1B. Therefore, if leptin level was corrected for carcass fat, there was no longer a significant effect of estradiol on leptin level [F(5,29) = 2.2; P < 0.07]. Ovariectomy, which reduced estradiol levels to approximately one-third of intact levels, did not significantly alter leptin levels 4 wk after surgery [F(1,18) = 0.42; P < 0.52], as is shown in Fig. 1C.




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Fig. 1.   Leptin levels are not independently altered by either estradiol administration or ovariectomy. A: leptin levels as a function of estradiol dose in intact lean mice given a range of exogenous 17beta -estradiol doses (0.05-17 µg) in pellet form for 14 days. Administration of exogenous estradiol at these doses resulted in serum estradiol levels of 45.6-513 pg/ml. Graph represents leptin levels on 14th day of estradiol administration. B: serum leptin levels (y-axis) regressed against carcass fat (x-axis) in estradiol-treated intact mice from A (r = 0.84; P < 0.0001). Dose of estradiol is given for each individual subject represented in graph (5-6/dose group). C: leptin (ng/ml) and estradiol (pg/ml) levels in lean ovariectomized (OVX) mice vs. sham control mice 4 wk after surgery. Values are means ± SE; n = 36/surgical group. * P < 0.02 vs. intact group.

Exogenous leptin administration induced a similar magnitude of weight loss in OVX and intact mice, which is suggested by a significant dose effect [F(5,53) = 35.5; P < 0.0001] in the absence of a significant (P < 0.906) dose × surgery interaction. All of these effects can be observed in Fig. 2, A and B. Furthermore, if %maximal weight change was plotted against leptin dose for both OVX and intact mice, neither the slopes nor the half-maximal effective doses were significantly different for the two groups. The half-maximal effective dose for both OVX and intact mice was ~2.8 µg/day, as is illustrated in Fig. 2C. %Maximal weight change was calculated as [(weight change on day 14 for dose group x - weight change for PBS group on day 14)/(weight change on day 14 for high dose group - weight change on day 14 for PBS group)] × 100. 




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Fig. 2.   Weight change from baseline after leptin treatment in intact (A) and OVX mice (B). Leptin was given via subcutaneous osmotic pump at 0.4-140 µg/day (0.02-7 mg · kg-1 · day-1). Baseline weights were 23.5 ± 0.21 g (OVX) and 22.9 ± 0.23 g (sham). Values are means ± SE; n = 5-6/dose group. C: %maximal weight change plotted against leptin dose (µg/day) in sham vs. OVX mice, with a sigmoid equation used for curve-fitting. %Maximal weight change was calculated as [(weight change on day 14 at dose x - weight change on day 14 for PBS group)/(weight change on day 14 at high dose - weight change on day 14 for PBS group)] × 100. Approximate half-maximal effective dose for both OVX and intact mice was 2.8 µg/day.

Carcass fat was lower in leptin-treated mice than in PBS-treated counterparts, as reflected by a significant treatment effect [F(5,118) = 36.9; P < 0.0001] (Table 1). The lowest significant leptin dose for fat reduction in OVX mice was 5.6 µg/day (P < 0.04) and for intact mice was 1.4 µg/day (P < 0.006). There were no significant differences in fat between PBS-treated OVX and intact mice. Leptin-treated intact mice also had significantly less carcass water than their PBS counterparts (P values < 0.0001). There were no significant effects of leptin on lean mass in either group.

                              
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Table 1.   Effects of r-MetMuLeptin on carcass composition in intact vs. OVX C57Bl6J mice

Carcass fat in mice treated with the leptin-estradiol combination was not significantly different from fat mass in mice treated with leptin alone at a much lower dose (20 µg/day) than used in the combination treatment (120 µg/day). However, fat mass in mice treated with the leptin-estradiol combination was lower than that observed in mice treated with leptin alone at the highest (140 µg/day) dose (P < 0.03). Fat mass in mice treated with this highest leptin dose, however, was greater than that of mice treated with a lower dose of leptin. The inconsistency in the high-dose range of the leptin dose-response curve probably reflects the insensitivity of the ether extraction assay when fat mass is so low. Carcass water and lean mass were statistically similar in mice treated with leptin alone and in mice treated with the leptin-estradiol combination.

Table 2 shows that exogenous leptin administration did not significantly affect serum estradiol levels. Leptin treatment did, however, induce a significant reduction in serum cholesterol levels in OVX and intact mice at the two highest doses (Ps < 0.01-0.008). The greatest reduction in cholesterol levels was found in intact or OVX mice treated with the leptin-estradiol combination (Ps < 0.01-0.001). Cholesterol levels were generally higher in OVX than in intact mice (P < 0.04). Triglyceride levels, which were higher in intact than OVX mice (P < 0.0001), were dramatically reduced after leptin treatment in intact mice at all doses (Ps < 0.001-0.0001) and at the 20-µg/day dose in OVX mice (P < 0.005). In intact mice, the largest reduction was observed in mice treated with the leptin-estradiol combination (P < 0.0001). Corticosterone levels were significantly elevated in OVX mice (P < 0.008) but were not affected by leptin treatment. BHBA (ketones) were significantly reduced in OVX mice (P < 0.03) and were normalized by leptin treatment to levels observed in intact mice (Ps < 0.01-0.001 vs. OVX PBS). Serum free fatty acids (FFA) were significantly reduced by leptin in intact mice (Ps < 0.04) but were significantly increased in OVX mice (Ps < 0.005-0.0009). Estrus cycle did not appear to be affected by leptin treatment, in that the number of cycles or estrus stage durations across treatment period was not different from that during the pretreatment period (data not shown).

                              
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Table 2.   Effects of r-MetMuLeptin on serum chemistry in intact vs. OVX C57Bl6J mice

Experiment 2 (obese OVX mice). Nineteen to twenty weeks after surgery, OVX mice weighed significantly more (15.3%) than intact controls [F(1,34) = 29.8; P < 0.01 (post hoc P < 0.0001)]. Leptin levels in these OVX mice were also significantly higher (4-fold) than in controls at the above time point [F(1,33) = 26; P < 0.0001 (post hoc P < 0.0001)] (Table 3). After 4 wk of estradiol pretreatment at a high dose (17 µg/day), OVX mice weighed significantly less than placebo-treated counterparts (P < 0.0008), which was reflected by a significant surgery × hormone interaction [F(1,34) = 22.5; P < 0.0001]. In contrast, estradiol-treated intact mice weighed more than placebo controls (P < 0.004). Leptin levels were significantly lower in estradiol-treated mice, regardless of surgical treatment, as is shown by a significant hormone effect [F(1,33) = 47.9; P < 0.0001]. As in experiment 1, leptin levels in estradiol vs. PBS-treated mice correlated with body fat in both OVX and intact mice (r = 0.89; P < 0.0001). All of these effects are shown in Table 3 in columns concerned with baseline weight and baseline leptin levels.

                              
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Table 3.   Effects of r-MetMuLeptin and estradiol pretreatment on carcass composition in obese OVX and lean sham-operated C57Bl6J mice

Leptin significantly reduced body weight in all groups [F(1,27) = 83.1; P < 0.0001], which can be observed in Table 3. Estradiol did not induce any further weight loss during the 5th and 6th wk of treatment in any group, nor did it alter the weight loss of leptin-treated mice. Finally, leptin treatment induced similar weight loss in OVX and intact mice when leptin-induced loss was subtracted from PBS-induced weight loss (OVX = -4.0 g, intact -3.75 g).

As shown in Table 3, final weights were significantly lower overall in leptin-treated mice [F(1,26) = 30.5; P < 0.0001]. OVX mice that had been allowed to gain weight had more carcass fat (4-fold) than intact controls (P < 0.0001), and all leptin-treated mice had significantly less carcass fat than PBS-treated counterparts [F(3,26) = 13.5; P < 0.001]. Furthermore, OVX mice pretreated with estradiol and then given leptin had less body fat than OVX counterparts given leptin without estradiol pretreatment (P < 0.0007). Carcass water was lower in OVX mice than in intact counterparts [F(1,26) = 25.05; P < 0.0001] and was essentially normalized by estradiol treatment [F(1,26) = 56.5; P < 0.0001]. The estradiol-induced normalization in carcass water for OVX mice was not altered by leptin treatment. Lean mass was significantly reduced in intact mice receiving leptin and the estradiol pretreatment [F(1,26) = 13.8; P < 0.001; (post hoc P < 0.005)]. All of these effects can be seen in Table 3.


    DISCUSSION
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Exogenous estradiol did not significantly affect serum leptin if the data were corrected for the fat loss induced by estradiol. Estradiol's lipolytic effects and the association of leptin level with fat mass are both well-documented phenomena (6, 7-9, 15, 22, 27, 29, 45). Therefore, it is unlikely that estradiol directly affects leptin secretion. Consistent with this, lean OVX mice had leptin levels that were similar to intact controls, suggesting that a reduction in estradiol also did not alter leptin levels. Our ovariectomy data, however, are not in agreement with data showing a reduction in leptin levels (41) and ob gene expression (41, 47) in rats 2-8 wk after surgery, despite the fact that fat mass is typically increased during this time period in rats (31). The reason for the inconsistency is unclear. There is some disagreement in the human subject literature, as well, regarding the relationship between estradiol level and leptin level. Havel et al. (18), for example, showed that leptin levels were not different in pre- and postmenopausal women if the levels were matched for fat mass, and that hormone replacement therapy did not alter leptin levels in postmenopausal women. Shimizu et al. (41) and Rosenbaum et al. (38), however, concluded by use of correlational methods that leptin levels were reduced in postmenopausal women. Finally, Messinis et al. (32) reported that leptin levels increased along with estradiol levels during the second half of the follicular phase of the menstrual cycle. Therefore, the relationship between estradiol and leptin levels appears to be a complex one, which may be dependent on factors such as the subjects' metabolic status and age.

The finding that exogenous leptin administration induced similar weight loss in OVX and intact mice further supports the hypothesis that estradiol does not directly regulate leptin production or functional activity. In addition, combined leptin and estradiol treatment did not reliably reduce fat mass more than leptin alone at a dose that was much lower than that used in the combination treatment. Again, the above evidence suggests that the two hormones do not act directly through a shared receptor or signaling pathway. Taken together, these data suggest that neither the ovary nor estradiol or estradiol receptors (by inference) are required for leptin-induced weight loss.

Ovarian hormones may induce a metabolic state that influences leptin's effects on fat mass, however, because leptin-induced fat loss was significantly attenuated in OVX mice compared with intact counterparts. For example, ovariectomy could have caused metabolic or neuroendocrine changes that indirectly affected leptin-induced fat loss before obvious weight gain. Cholesterol, triglycerides, BHBA, and corticosterone were indeed altered by ovariectomy, in that cholesterol and corticosterone were increased, whereas triglycerides and BHBA were reduced. All of these serum chemistry changes are consistent with previous observations in OVX animals (11, 20, 28).

Whether the observed increase in cholesterol levels or reduction in triglycerides would affect leptin's ability to reduce fat mass is unclear. Leptin has been shown to reduce carcass fat in animals with elevated cholesterol (16, 25, 36), but there are no data that address the effects of leptin in animals with reduced serum triglycerides, as observed in OVX mice.

It does not appear that OVX mice are incapable of responding to the effects of leptin on fat mobilization, because serum NEFA level was significantly increased in OVX mice, even at very low doses of leptin. Perhaps the intracellular FFA oxidation response to stress or energy deprivation is somehow impaired in OVX animals. In view of several reports suggesting that leptin reduces carcass fat by increasing intracellular FFA oxidation (40, 50), it is possible that leptin's effects on FFA oxidation could be blunted in an organism with an impaired FFA oxidation response to stress or fuel deprivation.

Leptin administration also reduced fat mass and body weight in OVX mice that were allowed to become obese. Nineteen to twenty weeks after surgery, OVX mice weighed 15% more than intact (sham) controls, an observation that is in agreement with earlier reports (2, 31, 43). These mice had 3-4 times more carcass fat and 3-4 times higher leptin levels than intact controls, which is consistent with the idea that leptin correlates very closely with body fat (6, 9, 15, 22, 27, 29). Two weeks of continuously infused leptin reduced body fat by ~40% in OVX mice. Fat mass appeared to be reduced to a greater degree in intact counterparts, in that it was reduced to undetectable levels. Again, this could suggest that leptin and estradiol may interact in an indirect fashion, with both hormones eventually affecting fat utilization through a common final pathway (as was suggested for the lean OVX mice). Such an interpretation is complicated for this obese set of OVX mice by the fact that animals with increased fat mass do not respond to low-dose leptin with the same magnitude of fat loss as do lean counterparts (3, 13, 36), which is why it was important to assess leptin efficacy in OVX mice that had not yet become obese.

Leptin was as effective in reducing total body weight in OVX mice as it was in intact controls, as demonstrated by similar net weight loss after 14 days of treatment for the two groups. Leptin, then, reduced total body weight more effectively in OVX-induced obesity than reported in diet-induced obese AKR mice or in obese female CD1 mice, where lower doses of leptin did not produce as great a magnitude of weight loss as in lean controls (3, 13, 36). It should be emphasized, however, that our OVX mice were not as "obese" as the diet-induced AKR mice or the obese female CD1 mice (in terms of body weight) that were used as subjects in the cited studies, and low-dose leptin causes a greater reduction in body weight in lean animals than in obese counterparts (13, 36).

Four weeks of estradiol treatment also reduced body weight in obese OVX mice, a result that is in agreement with previous work (2, 43, 45). After the 4-wk treatment period, OVX mice treated with estradiol were only slightly heavier than untreated, intact controls, and they and intact mice had very similar levels of fat and serum leptin. Subsequent leptin treatment along with continued estradiol, however, reduced all of these variables even further, to the point where body weight was lower than in intact mice or OVX mice treated with estradiol alone, and carcass fat was reduced to undetectable levels, as it was in intact mice treated with leptin. This result is not particularly surprising, because estradiol pretreatment reduced body fat and leptin levels, rendering a "lean" phenotype to these OVX mice, as can be inferred from the comparison of baseline weight for OVX/EST/PBS groups with that of either SHAM/CON/PBS or SHAM/EST/PBS groups in Table 3. As we have mentioned, lean animals are more sensitive than obese counterparts to leptin's effects on body weight at low to moderate dose ranges (13, 36). This, along with the fact that fat loss was not increased by simultaneous administration of leptin and estradiol in lean OVX mice, suggests that leptin was not additive to estradiol pretreatment in obese OVX mice.

A similar pattern of effects on fat mass were observed in intact mice pretreated with estradiol and given leptin, although total body weight was actually increased with estradiol adminstration in intact mice and was normalized with subsequent leptin treatment. The effect of estradiol on body weight in intact mice was probably due to its effects on carcass water, because water was also increased by chronic estradiol treatment and normalized by leptin.

All of the above addresses whether estradiol could either directly or indirectly affect leptin secretion or function. Some of our data also address the converse: that is, whether leptin acts by affecting estradiol production. The data in this study demonstrated that leptin administration did not significantly affect serum estradiol levels, which is in agreement with data showing that leptin did not alter basal granulosa cell number or production of estradiol in vitro (43, 48), despite the fact that leptin receptors have been localized on ovary tissue (5). Perhaps these receptors act to mediate leptin's inhibitory effects on estradiol production in states of metabolic stress, as would be the case when insulin is elevated (43).

Taken together, our results suggest that leptin does not directly affect estradiol production, because leptin administration does not alter estradiol levels. In addition, estradiol does not directly affect leptin production or its effects on fat mass reduction, because 1) exogenous estradiol does not alter leptin levels independently of its effects on fat mass, 2) the effects of leptin on body weight are the same in OVX and intact mice, and 3) the effects of combined estradiol and leptin treatment on fat mass are not consistently different from the effects of leptin alone. In addition, the ovary is not a necessary component of leptin's effects on metabolism, because OVX mice show leptin-induced weight loss and fat reduction, whether the mice are lean or obese. Ovarian hormones, however, may have indirect effects on these variables, since both lean and obese OVX mice show a moderate attenuation of leptin-induced fat loss. Therefore, whereas estradiol may indirectly affect leptin efficacy or secretion, it is unlikely that estradiol acts directly on leptin receptors or its signaling pathway to reduce fat mass.


    ACKNOWLEDGEMENTS

We are indebted to Larry Ross (Amgen Pathology Department) for measurement of the serum chemistry variables, to Margery Nicolson and Jason Moore (Amgen Neuroscience) for measurement of serum leptin, and to David Hess (Oregon Regional Primate Center) for measurement of serum estradiol.


    FOOTNOTES

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

Address for correspondence and reprint requests: M. A. Pelleymounter, Dept. of Neuroscience, Neurocrine Biosciences, 10555 Science Center Dr., San Diego, CA 92130 (E-mail: Mpelleymounter{at}neurocrine.com).

Received 16 November 1998; accepted in final form 4 February 1999.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
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