Differential regulation of insulin resistance and hypertension by sex hormones in fructose-fed male rats.
ABSTRACT Differences in gender are in part responsible for the development of insulin resistance (IR) and associated hypertension. Currently, it is unclear whether these differences are dictated by gender itself or by the relative changes in plasma estrogen and/or testosterone. We investigated the interrelationships between testosterone and estrogen in the progression of IR and hypertension in vivo in intact and gonadectomized fructose-fed male rats. Treatment with estrogen significantly reduced the testosterone levels in both normal chow-fed and fructose-fed rats. Interestingly, fructose feeding induced a relative increase in estradiol levels, which did not affect IR in both intact and gonadectomized fructose-fed rats. However, increasing the estrogen levels improved insulin sensitivity in both intact and gonadectomized fructose-fed rats. In intact males, fructose feeding increased the blood pressure (140 +/- 2 mmHg), which was prevented by estrogen treatment. However, the blood pressure in the fructose-fed estrogen rats (125 +/- 1 mmHg) was significantly higher than that of normal chow-fed (113 +/- 1 mmHg) and fructose-fed gonadectomized rats. Estrogen treatment did not affect the blood pressure in gonadectomized fructose-fed rats (105 +/- 2 mmHg). These data suggest the existence of a threshold value for estrogen below which insulin sensitivity is unaffected. The development of hypertension in this model is dictated solely by the presence or absence of testosterone. In summary, the development of IR and hypertension is governed not by gender per se but by the interactions of specific sex hormones such as estrogen and testosterone.
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ABSTRACT: Selected steroid compounds (androsterone, epi‐androsterone, dehydro‐epi‐androsterone, testosterone, stigmasterol, β‐sitosterol, estradiol, hydrocortisone, and cholesterol) were separated by adsorption TLC on silica gel 60, not activated, and activated at temperatures of 100°C, 120°C, 150°C, and 200°C, during 15, 30, 60, and 120 min, respectively. The mixture of chloroform and acetone (85∶15, v/v) was used as mobile phase. The lowest RF values of particular substances investigated were obtained on chromatographic plates precoated by silica gel activated at a temperature of 120°C. The time and activation temperature of silica gel influenced the order of substances adsorbed, the values of separation factors: ΔRF, RF and α. None of the chromatographic conditions used allowed for the separation of the following pairs of substances: androsterone‐dehydro‐epi‐androsterone, and testosterone‐estradiol.Journal of Liquid Chromatography & Related Technologies 08/2006; 29(14):2035-2044. · 0.64 Impact Factor
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ABSTRACT: Hyperandrogenism and vascular dysfunction often coexist in women with Polycystic Ovary Syndrome (PCOS). We hypothesized that testosterone compromises cutaneous microvascular dilation in women with PCOS via the endothelin-1 ET-B subtype receptor. To control and isolate testosterone effects on microvascular dilation, we administered a gonadotropin releasing hormone antagonist (GnRHant) for 11 days in obese otherwise healthy women [(Controls, 22.0 (4) y, 36.0 (3.2) kg/m(2))] or women with PCOS [23 (4) y, 35.4 (1.3) kg/m2)], adding testosterone (T, 2.5 mg/day) for days 8-11. Using laser Doppler flowmetry and cutaneous microdialysis, we measured changes in skin microcirculatory responsiveness (ΔCVC) to local heating while perfusing ET-A (BQ-123) and ET-B (BQ-788) receptor antagonists under three experimental conditions: baseline (BL, pre-hormone intervention), GnRHant (day 4 of administration), and T administration. At BL, ET-A receptor inhibition enhanced heat-induced vasodilation in both groups [ΔCVC Control 2.03 (0.65), PCOS 2.10 (0.25), AU/mmHg, P< 0.05]; ET-B receptor inhibition reduced vasodilation in Controls only [ΔCVC 0.98 (0.39), 1.41 (0.45) AU/mmHg for Controls, PCOS] compared to saline [ΔCVC Controls 1.27 (0.48), PCOS 1.31 (0.13) AU/mmHg]. GnRHant enhanced vasodilation in PCOS [saline ΔCVC 1.69 (0.23) AU/mmHg vs. BL, P<0.05), and abolished the ET-A effect in both groups; a response reasserted with T in Controls. ET-B receptor inhibition reduced heat-induced vasodilation in both groups during GnRHant and T [ΔCVC, Controls: 0.95 (0.21) vs. 0.51 (13), PCOS: 1.27 (0.23) vs. 0.84 (0.27); for GnRHant vs. T, P< 0.05)]. These data demonstrate androgen suppression improves microvascular dilation in PCOS via ET-A and ET-B receptors.AJP Endocrinology and Metabolism 08/2013; · 4.51 Impact Factor
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ABSTRACT: Combinations of fructose-and fat-rich diets in experimental animals can model the human metabolic syndrome (MS). In rats, the increase in blood pressure (BP) after diet manipulation is sex related and highly dependent on testosterone secretion. However, the extent of the impact of diet on rodent hypophysial-testicular axis remains undefined. In the present study, rats drink-ing a 10% fructose solution or fed a high-fat (35%) diet for 10weeks had higher plasma levels of luteinizing hormone (LH) and lower plasma levels of testosterone, without significant changes in circulating follicle-stimulating hormone or the weight of most reproductive organs. Diet manipulation brought about a significant increase in body weight, systolic BP, area under the curve (AUC) of glycemia after an intraperitoneal glucose tolerance test (IPGTT), and plasma low-density lipoprotein cholesterol, cholesterol, triglycerides, and uric acid levels. The con-comitant administration of melatonin (25 μg/mL of drink-ing water) normalized the abnormally high LH levels but did not affect the inhibited testosterone secretion found in fructose-or high-fat-fed rats. Rather, melatonin per se inhibited testosterone secretion. Melatonin significantly blunted the body weight and systolic BP increase, the increase in the AUC of glycemia after an IPGTT, and the changes in circulating lipid profile and uric acid found in both MS models. The results are compatible with a pri-mary inhibition of testicular function in diet-induced MS in rats and with the partial effectiveness of melatonin to counteract the metabolic but not the testicular sequelae of rodent MS.Hormone molecular biology and clinical investigation 07/2013; 16(2):101-112.
DIFFERENTIAL REGULATION OF INSULIN RESISTANCE AND
HYPERTENSION BY SEX HORMONES IN
FRUCTOSE-FED MALE RATS
Harish Vasudevan, Hong Xiang and John H. McNeill*
Division of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences,
University of British Columbia, Vancouver, British Columbia, V6T 1Z3,Canada
Short title: Sex hormones in Insulin Resistance and Hypertension
Word count: Title: 14; Abstract 233; Figures: 8
Author for correspondence:
John H. McNeill, PhD, FRSC.
Faculty of Pharmaceutical Sciences
University of British Columbia
2146 East Mall
Vancouver, BC, V6T 1Z3, CANADA
Articles in PresS. Am J Physiol Heart Circ Physiol (June 10, 2005). doi:10.1152/ajpheart.00399.2005
Copyright © 2005 by the American Physiological Society.
Differences in gender are in part responsible for the development of insulin resistance
and associated hypertension. Currently, it is unclear whether these differences are
dictated by gender itself or by the relative changes in plasma estrogen and/or testosterone.
We investigated the interrelationships between testosterone and estrogen in the
progression of insulin resistance and hypertension in vivo in intact and gonadectomized
fructose-fed male rats. Treatment with estrogen significantly reduced the testosterone
levels in both normal chow-fed and fructose-fed rats. Interestingly, fructose feeding
induced a relative increase in estradiol levels, which did not affect insulin resistance in
both intact and gonadectomized fructose-fed rats. However, increasing the estrogen
concentration improved insulin sensitivity in both intact and gonadectomized fructose-fed
rats. In intact males, fructose feeding increased the blood pressure (BP) (F-140±2
mmHg), which was prevented by estrogen (FE). However, the BP in these rats (FE-
125±1 mmHg) was significantly higher than that of normal chow-fed (C-113±1 mmHg)
and fructose-fed gonadectomized rats. Estrogen treatment did not affect the blood
pressure in gonadectomized fructose-fed rats (GFE-105±2 mmHg). These data suggest
the existence of a threshold value for estrogen below which insulin sensitivity is
unaffected. The development of hypertension in this model is dictated solely by the
presence or absence of testosterone. In summary the development of insulin resistance
and hypertension are governed not by gender per se, but by the interactions of specific
sex hormones such as estrogen and testosterone.
Key words: Hyperinsulinemia, fructose hypertensive rat, gender, estrogen and
The metabolic syndrome is comprised of a cluster of cardiovascular risk factors
including abdominal obesity, hypertriglyceridemia, low levels of high-density
lipoprotein, insulin resistance and hypertension (21). A key factor contributing to the
metabolic syndrome is the resistance to insulin (IR) and subsequent hyperinsulinemia,
which is often associated with hypertension in both humans (7, 28) and several animal
models (3, 25). Differences in gender have been shown to influence the progression of
insulin resistance and consequently hypertension (11, 12). Studies using fructose-fed rats
(3, 36) have demonstrated that the degree of insulin resistance developed in males is
greater than that in females (12). As hypertension was observed secondary to insulin
resistance, the changes in blood pressure were dictated by sex-dependent differences in
insulin sensitivity (11). Further, this differential development of insulin resistance and
hypertension was ascribed to the presence of estrogen (10). Clinical evidence supports
the above findings as men and postmenopausal women demonstrate a higher risk of
developing hypertension as compared to premenopausal women (14). Insulin sensitivity
and glucose tolerance are improved in postmenopausal women subsequent to treatment
with estrogen alone or in combination with progesterone (18). Thus experimental
evidence and short-term clinical studies support the role of estrogen in preventing the
development of insulin resistance and hypertension. However, the effects of long-term
hormone replacement therapy on cardiovascular health have been controversial and the
reasons are yet to be fully understood (1, 15, 19, 26, 38).
Taken together, the physiological differences arising due to gender variation
assume a significant role in the development of insulin resistance and hypertension.
However, the effects of estrogen replacement have been studied mainly in
ovariectomized rats. Testosterone is present in females, albeit in lower amounts than men
and, is responsible for governing certain phenotypes associated with males. In vivo, it has
been recently shown that testosterone is essential in male rats for the development of
hypertension secondary to insulin resistance (32). Although the individual contributions
of estrogen and testosterone to the development of insulin resistance and hypertension
have been investigated, little is known as to how they affect insulin sensitivity and blood
pressure in presence of each other. Do estrogen and testosterone complement or
counteract each other in vivo? Furthermore, how does the endogenous residual estrogen
or testosterone in gonadectomized male or female rats respectively, influence insulin
sensitivity and blood pressure?
Testosterone has been strongly implicated in the development of insulin resistance
and hypertension in women with polycystic ovary syndrome (PCOS) (17, 30, 35) or
preeclampsia (22), which is similar to insulin resistance and hypertension observed in
women after menopause. In a recent study by Fortepiani et al. (8), both 18 month-old
ovariectomized spontaneously hypertensive rats (SHR) and intact very old female (post-
estrous) SHR had significantly higher levels of testosterone, as compared to young
estrous rats, which had higher circulating estrogen and lower levels of testosterone
respectively. The mean arterial pressure (MAP) was significantly higher in males and 18-
month OVX rats as compared to control young estrous rats. These results provide an
indication that the unmasking of testosterone-mediated effects following the loss of
estrogen may be responsible for the increased MAP in these rats. On the other hand male
rats, despite having estrogen present in the plasma, are susceptible to developing insulin
resistance independent of testosterone levels. Thus, despite being ascribed opposite roles
in the regulation of blood pressure, no studies to date have attempted to investigate the
interactions and interdependence between testosterone and estrogen in males and females
and how the loss of one hormone influences the actions of the other in normal and insulin
The aim of this study was to determine whether a critical balance between
estrogen and androgen in males is needed for the regulation of insulin sensitivity and
blood pressure. We hypothesized that the balance between testosterone and estrogen in
male rats affects the induction of insulin resistance and subsequent development of
MATERIALS AND METHODS
The fructose hypertensive rat model was used in the studies outlined below. Male
Wistar rats were obtained from Charles River, Montreal, Canada. The rats were cared for
as per the guidelines outlined by the Canadian Council on Animal care (CCAC) and the
American Physiological Society in the Guiding Principles in the Care and Use of
Animals. Subsequent to arriving, the rats were acclimatized in the animal facility at the
Faculty of Pharmaceutical Sciences, University of British Columbia, for one week prior
to treatment. The starch present in normal laboratory rat chow was replaced with excess
fructose, which was obtained commercially as a pre-formulated diet (Teklad Labs,
Madison, WI). Feeding this fructose-enriched diet has been shown in previous studies to
induce insulin resistance and hypertension (9, 20).
In the first study, forty rats at 6 weeks of age and having intact testes were used.
In the second study, two-thirds of the total animals underwent gonadectomy at 5 weeks of
age. The rats were shipped at the age of 6 weeks. The animals were cared for as stated
In the first study, male rats with intact testes were divided into four experimental
groups; normal chow-fed control (C), control rats treated with estrogen (CE), fructose-fed
(F) and fructose-fed estrogen treated (FE). Prior to introduction of fructose, the basal
blood pressure was measured by the non-invasive tail cuff method as explained below. In
addition, changes in body weight and food intake were estimated. On the day of starting
the fructose diet, rats were fasted for 5 hours and blood was collected from the tail vein.
The rat chow in groups F and FE was replaced with the diet containing fructose. All the
rats were allowed ad libitum access to food and water throughout the 6-7 weeks of
treatment. Estrogen treatment was carried out by implantation of estrogen pellets (0.5 mg
for 60 day release) (Innovative Research of America, Sarasota Fl). Under light halothane
anaesthesia, the pellets were subcutaneously implanted using a trochar under aseptic
conditions. The implantation was performed on the same day as the initiation of fructose
feeding. The food consumption of the rats was assessed once a week, along with body
weights. Owing to the unexplained death of 2 rats in the fructose-fed group treated with
estrogen, plasma blood glucose was measured in the surviving rats during the subsequent
2 weeks using AccucheckTM glucose strips (Roche).
In the second study, sham-operated and gonadectomized male Wistar rats were
initially divided into 4 experimental groups namely, sham-operated chow-fed control (C;
n=5), sham-operated fructose-fed (F; n=5), gonadectomized normal chow-fed (G; n=4)
and gonadectomized-fructose-fed (GF; n=16). Following 6 weeks of feeding fructose,
one-half of the GF rats were subcutaneously implanted with a 0.5 mg estrogen pellet (60
day release, Innovative research of America, Sarasota Fl) to form the GFE group (5th
experimental group). All implantations were carried out under light halothane anesthesia.
One rat died due to halothane, but the procedure was successful in the remaining animals.
Thus the final ‘n’ in GFE was 7. 17β-estradiol levels in the plasma were estimated at 3
and 10 days following implantation. Blood pressure was measured in all the 5 groups, i.e.
C, F, G, GF and GFE, 2 weeks subsequent to implantation, i.e. at the end of study week
8. Subsequently, changes in insulin sensitivity were assessed as explained below. The
animals were euthanized and blood was collected for measuring plasma testosterone and
Measurement of blood pressure and assessment of insulin resistance/sensitivity
Systolic blood pressure was measured in conscious rats using the indirect non-
invasive tail-cuff method as previously described (4, 10). An oral glucose tolerance test
(OGTT) was performed in the week following blood pressure measurement. After the
oral glucose challenge, insulin sensitivity was estimated using the formula of Matsuda
and DeFronzo using 100 as constant: ISI = 100/Sq. rt. [(fasting glucose x fasting insulin)
x (mean glucose x mean insulin)] (24). Among all methods used, the values obtained by
oral glucose tolerance test (OGTT) offered the best correlation with values from the
euglycemic hyperinsulinemic clamp (24).
Blood samples were collected from the tail vein at week 0 for estimation of
plasma glucose, insulin, testosterone and 17β-estradiol. Upon termination, blood was
collected by cardiac puncture into plastic centrifuge tubes containing 2% ethylene
diaminotetraacetic acid (EDTA) and 0.04M indomethacin and centrifuged at 4500 rpm
for 25 minutes at 4oC. Plasma was aspirated and stored at -80oC for measuring
testosterone, 17β-estradiol and triglycerides.
Plasma glucose was estimated using a Beckman Glucose Analyzer II, while
triglycerides were measured colourimetrically using a commercially available kit. Plasma
insulin was measured using commercially available radioimmunoassay (RIA) kits from
Linco Diagnostics Inc, USA, while 17β-estradiol and testosterone were measured using
RIA kits from MP Biomedicals, USA.
Chemicals and Reagents
All chemicals unless otherwise mentioned were of reagent grade and purchased
from Sigma, Mo.
All data were analyzed using one-way analysis of variance (ANOVA). Data
involving multiple time points were subject to the GLM ANOVA using the NCSS 2000
statistical software. For all the results, Newman Keuls test was used as post hoc test. The
value of P<0.05 was taken as the level of significance. All results are reported as mean ±
Physical appearance, body weight and food intake
Two animals from the fructose-fed estrogen treated group (FE) died during the
course of study 1 due to hypoglycemia. The remaining rats in FE, although hypoglycemic
at 2 weeks, improved over subsequent weeks of the study (data not shown). We observed
a significant decrease in the testicle size of male rats subsequent to treatment with
estrogen. At termination, the estrogen-implanted male rats (CE and FE) had significantly
lower body weights compared to the non-implanted groups (Table 1a). This was observed
from the second week following estrogen implantation, until the end of the study. The
animals' weights stabilized after 3 weeks, although they were significantly lower than C
and F throughout the study. There was no difference between the body weights of normal
chow and fructose-fed groups. Thus fructose feeding per se did not affect the changes in
weights. Estrogen implanted rats consumed less food as compared to the untreated groups
In the gonadectomized rats treated with estrogen (GFE), a lower weight gain was
observed compared to the other groups upon termination at the end of 3 weeks (Table 1b)
following estrogen implantation (study week 9). Further estrogen induced a weight loss
in the GF rats when compared with values prior to treatment.
Plasma triglyceride levels
Seven weeks of fructose feeding elevated the plasma triglyceride (TG) levels (F; 2
± 0.4 vs. C; 0.6 ± 0.1 mmol). Treatment with estrogen did not reduce TG levels in the
fructose-fed rats (FE; 1.6 ± 0.2 mmol respectively). Control estrogen treated rats (CE; 0.5
± 0.1 mmol) did not show any change in triglyceride levels compared to untreated
Plasma 17β β-estradiol and testosterone levels
Plasma 17-β-estradiol levels were elevated in both CE and FE compared to C and
F respectively (Table 2). Correspondingly, testosterone levels dropped significantly in the
estrogen-treated groups at termination. Curiously, although fructose feeding did not alter
the testosterone levels in F as compared to C, estradiol levels in F were higher than C.
However, this difference in estrogen did not prevent the development of insulin
resistance in these rats. The testosterone to estradiol ratio was significantly higher in the
untreated animals compared to the estrogen-implanted groups. Estrogen reduced the
testosterone to estradiol ratio in CE and FE respectively (Table 2).
Prior to fructose feeding, gonadectomized rats had significantly higher plasma
estradiol levels (87.2 ± 3.9 pg/ml) than the sham-operated males (66.7 ± 3.3 pg/ml). At
termination, the F and GF groups had higher plasma estradiol compared to C and G
respectively. As shown in Table 3, there was a marked release of estradiol from the pellet
on the 3rd day following estrogen implantation. 17β-estradiol values were greater than
3000 pg/ml at the end of 3 days. After ten days of treatment, although estradiol levels
were significantly higher than the other groups, there was a significant decrease
compared to the values on day 3. Estradiol was the highest in the estrogen-implanted
group (GFE) compared to other groups until the rats were terminated.
We did not detect any changes in testosterone in both the sham-operated groups
(C and F) used in the study (data not shown). However in the untreated and estrogen-
treated gonadectomized rats, testosterone levels were undetectable.
Assessment of insulin resistance/sensitivity
Rats fed with fructose for 6-7 weeks (F) rats developed hyperinsulinemia within
10 minutes of ingestion of glucose, which was sustained for 60 minutes (figure 1a).
Glucose levels were elevated in all the groups 10 minutes following dosing. However,
there was no significant difference among the groups with respect to body glucose
disposal pattern over the entire 90 minutes of observation (figure 1b). Estrogen prevented
hyperinsulinemia in the fructose-fed rats (FE) and maintained insulin levels at par with
controls. The insulin sensitivity index (ISI) was significantly lower in the fructose-fed
group compared to the other groups, indicating the presence of insulin resistance in these
animals. Estrogen treatment improved the insulin sensitivity in fructose fed rats as
indicated by elevated insulin sensitivity index (figure 1c).
The gonadectomized fructose-fed rats (8 weeks), which were treated with
estrogen for 2 weeks did not become insulin resistant, as they had a higher ISI compared
to F and GF, indicating normal insulin sensitivity (figure 2c). Following 2 weeks of
treatment, estrogen reduced plasma insulin in the gonadectomized fructose-fed rats
(GFE) to levels comparable with C and G groups (figure 2a; see inset). Further, glucose
levels in GFE were significantly lower than the remaining groups (figure 2b).
In the first study, after 6 weeks of fructose feeding and estrogen treatment,
systolic blood pressure was higher in the fructose-fed rats (F; 140±2 mm Hg) compared
control (C; 113 ± 1 mm Hg). Blood pressure was also higher in fructose-fed animals
treated with estrogen (FE) (125±1 mm Hg). However, FE was significantly lower as
compared to F. Blood pressure was unchanged in both the control groups (C and CE)
after 6 weeks (figure 3).
In the second study, following 2 weeks of estrogen treatment, at 8 weeks post
fructose feeding, the fructose-fed sham-operated rats (F) had higher blood pressure as
compared to normal chow-fed controls (C). Blood pressure was unaffected in the
gonadectomized fructose-fed rats regardless of the presence (GFE) or absence (GF) of
estrogen (figure 4).
Insulin resistance (IR) is the initial stage of the metabolic syndrome, which is
suggested to result from complex interactions between genetic and environmental factors.
Feeding rats a high fructose diet induces insulin resistance, hyperinsulinemia,
hypertriglyceridemia and hypertension (11). In humans (14) as well as rodents (29),
females are less susceptible to developing hypertension as compared to males. A similar
profile was observed in the induction of insulin resistance in Wistar rats, where females
fed with fructose did not develop insulin resistance and hyperinsulinemia in comparison
with age-matched males (10, 11). The protective effects of estrogen (10, 11) and
permissive effects of testosterone (32) in the development of hypertension secondary to
insulin resistance have been demonstrated in separates studies. However, little is known
regarding the effects of hormonal balance in vivo on the development of insulin
resistance and hypertension in either of the sexes. In the present set of studies, we have
shown for the first time that it is not merely the differences in gender per se but the
interactions between estrogen and testosterone, which influence the development of
insulin resistance and hypertension in either sex. In males the absolute presence or
absence of testosterone is responsible for the changes in blood pressure secondary to
insulin resistance. Secondly, in males, estrogen may be required to attain a threshold level
in order to observe its preventive effects on insulin resistance and hypertension.
Effects of sex hormone balance on the induction of insulin resistance in male rats
Fructose-fed male rats with intact testes developed insulin resistance after 6
weeks of fructose feeding, as indicated by hyperinsulinemia and lower insulin sensitivity
index (ISI) compared to control (C). This result agrees with the previous reports (9, 36).
Estrogen elevated the insulin sensitivity in male fructose-fed rats (figure 1c). This is
supported by results from our laboratory (33), which indicate a similar effect of estrogen
on insulin sensitivity in ovariectomized rats. Estrogen treatment resulted in a lower food
intake and weight gain in male rats (Table 1a). Treatment with estrogen significantly
reduced the testicle size in male rats, suggesting decreased testicular activity. In a study
by Valigora et al, chronic estrone treatment in male SHR reduced plasma testosterone
levels from 1.2 ng/ml to 0.1-0.2 ng/ml (34). A similar profile was observed in our studies
wherein 17β-estradiol levels were elevated while testosterone levels fell subsequent to
estrogen treatment (Table 2). Interestingly, plasma estradiol levels were higher in both
intact and gonadectomized fructose-fed male rats, which were untreated (F & GF) as
compared to normal chow-fed rats (C & G respectively) (Tables 2 & 3). Thus fructose-
feeding seems to retard the fall in estradiol levels, although the resulting higher level of
estradiol was still unable to prevent insulin resistance secondary to fructose feeding
(Table 3). This finding seems to contradict the currently held belief that estrogen
enhances insulin sensitivity. To date, we have not found any studies, which have
investigated the effects of diet-induced insulin resistance on changes in plasma estradiol
levels. While it may be speculated that the fructose-enriched diet affects estradiol
synthesis in males, we have no data concerning this issue. Additional studies are needed
to look at the changes in the levels and actions of estradiol under states of excess
carbohydrate feeding. Secondly, potential insulin resistance-induced changes in the
“bioavailability” of estrogen in males need to be determined. The levels of circulating
estradiol in male rats (Tables 2 & 3 respectively), although in the same range as that