NAFLD is currently recognized as the most common form of chronic liver disease in the United States and in many parts of the world. Some data suggest that Mexican Americans are more likely to have NAFLD and blacks are less likely compared with non-Hispanic whites.1 NAFLD is considered a nonspecific term encompassing several clinicopathologic entities (steatosis alone, steatonecrosis, steato-hepatitis and histologic alcoholic-like hepatitis) that are similar to alcoholic liver diseases2 in the absence of significant alcohol abuse. Simple hepatic steatosis and hepatic steatosis with nonspecific inflammation are believed to have a generally benign course, whereas nonalcoholic steatohepatitis (NASH) can progress to cirrhosis, leading to liver failure and hepatocellular carcinoma (HCC).3,4 Furthermore, it has been proposed that NAFLD is associated with metabolic syndrome and varying degrees of insulin resistance.5-7 Although insulin resistance and hyperinsulinemia are frequently found in obese subjects with NAFLD, both are also noted in lean subjects with fatty liver disease and normal glucose tolerance.

A two-hit theory best describes the progression from simple steatosis to NASH, fibrosis, or cirrhosis. These two hits consist of the accumulation of excessive hepatic fat primarily owing to insulin resistance, and oxidative stress owing to reactive oxygen species (ROS). Mitochondria are the major cellular source of ROS in cases of NASH. Antioxidants such as vitamin E, N-acetylcysteine, betaine, and others may be beneficial in the treatment of NASH.8,9

Interestingly, early descriptions by those who studied NAFLD suggested a female preponderance of the condition. More recent studies, however, including population-based studies, show that NAFLD affects both sexes equally or a higher proportion of men.10,11 In a prospective study, the incidence of NAFLD was higher in men than in women. Age was an independent predictor of the development of NAFLD in Japanese women but not men.12 The differences between the sexes in regards to the influence of age on the incidence and prevalence of NAFLD, are attributed to the putative protective effects of estrogen.

The aim of this study was to investigate the prevalence of NAFLD in premenopausal and postmeno-pausal women and women with PCOS.

Of the 197 patients, 93(47.2%) had NAFLD and 104 (52.8%) did not have NAFLD. There were 90 premenopausal patients (46.2%), 57 postmenopausal patients (29.4%) and 50 PCOS patients (25.4%). The prevalence of NAFLD in premenopausal, postmeno-pausal and PCOS patients was 32.2%, 57.9%, and 62%, respectively.

The comparisons of menopausal status and NA-FLD were statistically significant at the p = 0.05 level by the KW test for the following variables: age, anthropometric variables (waist, waist to hip ratio, BMI, percentage of body fat and diastolic blood pressure), HOMA-IR, liver enzymes (alanine amino-transferase (ALT), with a positive trend for aspartate aminotransferase (AST) and alkaline phosphatase (AP), serum glucose, blood ureic nitrogen (BUN), creatinine, total cholesterol, HDL-C, LDL-C, conjugated bilirubin, and serum estradiol and cortisol concentrations (Table 1).

In general, patients with NAFLD were older, had greater BMI, central obesity and IR. With regard to MS, 11 (22%) patients with PCOS, 7 (7.7%) preme-nopausal and 13 (24.5%) postmenopausal patients fulfilled the criteria of metabolic syndrome (MS). The prevalence of MS was higher in postmenopausal 39%) and PCOS (36%) patients with NAFLD compared with the other groups. (Table 2) Figure 1 shows the predicted probability of NAFLD by meno-pausal status. Premenopausal women show a lower probability of NAFLD, which is significantly different from the probability of the other two groups. MS analysis of weight, height, BMI, waist, waist to hip ratio and percentage of body fat shows an increase in the probability of NAFLD with increasing values. In other words, women with high values of weight, BMI, waist, waist to hip ratio, and percentage of body fat have the highest probability of having NAFLD. We found a similar pattern after correcting for the effect of menopause, although the difference between postmenopausal women and women with PCOS was not significant.

Figure 1.

Relationship between predicted probability of NAFLD and menopause, corrected for MS. Predicted probabilities for NAFLD = with 95% confidence limits AtFAC1_2=-0.

(0.04MB).

With regard to hormonal levels, we observed that premenopausal women had higher serum levels of estradiol and cortisol (Figure 2) and that premeno-pausal women with NAFLD had significantly higher levels of estradiol than the other groups. Postmeno-pausal women with or without NAFLD had no significant difference in cortisol and estradiol levels, but had the lowest serum values of estradiol (but not of cortisol) of all the groups. In the PCOS group, there was a significant difference in the values for estradiol and cortisol between patients with NAFLD and those without the disease, with patients without NAFLD having higher levels of these hormones.

Figure 2.

A. Shows serum levels of estradiol in the different menopausal groups, the difference is significative between the groups. B. Shows serum levels of Cortisol in the different menopausal groups; although it does not seem diverse, the difference is significative between the groups.

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When the association between levels of estradiol and cortisol with the presence of NAFLD was analyzed, the results indicated a higher probability of developing NAFLD with decreasing serum levels of these hormones (Figure 3).

Figure 3.

Relationship between predicted probability of NAFLD and estradiol (A) and cortisol (B) serum concentration. Predicted probalities for NAFLD =1 Whit 95% Confidence Limits.

(0.09MB).

NAFLD is a chronic condition in patients who do not have a history of excessive alcohol consumption and is characterized by more than 5% of hepatocytes affected with lipid accumulation. Caldwell, et al., in 1999 suggested a relationship between NAFLD and cryptogenic cirrhosis, and several other studies have shown that NAFLD patients are at higher risk of developing a wide range of progressive liver diseases including hepatocellular carcinoma.15,16

In previous studies, a possible role for estrogens in fatty liver disease has been suggested since Nguyen et al. demonstrated in 2001 that female patients with breast cancer who were receiving treatment with tamoxifen, a nonsteroidal antiestro-genic drug had a higher prevalence of intra-abdomi-nal fat accumulation and incidence of NAFLD.17

As has been reported in many studies,3,4 we observed large differences in BMI and abdominal obesity between the groups, especially between patients with and without NAFLD. Similar differences were observed with biochemical variables, particularly insulin resistance, which is the most common characteristic of subjects with NAFLD. The highest incidence of MS was found in patients with PCOS and NAFLD, whereas none of the patients with PCOS and without NAFLD fulfilled the criteria for metabolic syndrome.18

As mentioned above, it has been suggested that NAFLD is a disease more prevalent in men than in women and more frequent after menopause than it is in premenopausal women, implying that estrogens might have a role in the pathophysiology of fatty liver. Clark, et al., and Fraser, et al., have recently demonstrated in their epidemiological studies that the prevalence is higher in men than in women.10,19 An-gulo et al. in their study of predictors of fibrosis suggested that women with NAFLD had a higher association with fibrosis progression as they aged.20 In this study, the prevalence of NAFLD that we observed was clearly different according to menopausal status. We observed that the postmenopausal women had a higher prevalence of NAFLD than did the pre-menopausal women, but the highest incidence of NA-FLD was present in the patients with PCOS. It has been suggested that several physiological changes occur in postmenopausal women, as a result of the low serum levels of estrogen. One of the most important alterations is fat redistribution and its metabolic consequences including dyslipidemia and glucose intole-rance.21 These physiological and hormonal changes are probably the cause of the alterations of lipid metabolism in postmenopausal women who, as a consequence, have NAFLD. Kojima et al. found in their epidemiological study a higher prevalence of NAFLD in postmenopausal female patients than in men.22 In this study, we found a prevalence of NAFLD of nearly 60% in postmenopausal patients.

PCOS is a common disease and was first described by Stein and Leventhal in 1935. It affects 5-11% of women of reproductive age23 and is recognized as a major factor in alterations of the metabolic, cardiovascular and reproductive systems.24 The most important pathophysiologic feature of PCOS is IR, which is also the main characteristic of MS. Gamba-rin-Gelwan et al. found a prevalence of 55% of NA-FLD in female patients with PCOS,16 furthermore Cerda et al. observed a prevalence of 63% of PCOS patients with biochemical alterations and a prevalence of NAFLD of 41%.18 Moreover Setji et al. described several biochemical alterations, especially in liver enzymes, in patients with PCOS and when those patients underwent liver biopsy all of them had NASH with fibrosis.25 In the present study, we found that women with PCOS had a higher prevalence of NAFLD (62%) than did the other groups. This might have two pathogenic components: firstly, the altered hormonal profile and lower levels of estradiol in these patients and, secondly, the persistent state of hyperinsulinemia and insulin resistance typical of this disease.

Interestingly, we observed higher serum estradiol concentrations in patients without NAFLD, especially in the premenopausal women without NAFLD group, whereas postmenopausal women with NA-

FLD had the lowest cortisol and estradiol levels. Despite these findings, the group with highest prevalence of NAFLD was the PCOS group, which may be related to the two pathogenic components suggested above.

What possible mechanisms could explain the lower prevalence of NAFLD in premenopausal women? Interestingly, an animal study showed that hepatic steatosis becomes evident spontaneously in aromatase deficient mice, which lack the ability to produce estrogen and are impaired with respect to hepatocellular fatty acid β-oxidation. Estradiol replacement reduces hepatic steatosis and restores the impairment in mitochondrial and peroxisomal fatty acid β-oxidation to the wild-type level.26

Estradiol is a strong endogenous antioxidant that suppresses liver fibrosis in animal models and attenuates induction of oxidation-reduction sensitive transcription factors, hepatocyte apoptosis and stellate cell activation by avoiding ROS generation in cultures.27-30

One of the most important activities of estradiol is its effect on serum lipid concentration that results from estrogen-mediated effects on the hepatic expression of apoprotein genes. These effects decrease the concentration of LDL and increase the concentration of triglycerides, total cholesterol and HDL.31 We propose that another mechanism is the antioxi-dant effect that estradiol seems to have in several studies, which may be due to changes mediated by estrogen receptors in the expression of genes that regulate the production of superoxide.31,32 In hepatic steatosis, there is an increase in hepatocyte lipope-roxidation, which has as its consequence the activation of hepatic stellate cells. These are the principal target of inflammatory and oxidative stimulation and produce extracellular matrix components that cause triglyceride accumulation in hepatocytes and finally fibrosis.3

On the other hand when alcohol is involved, in vivo studies had concluded that the sensitivity of rat liver to alcohol-induced injury is directly related to estrogen, which increases endotoxin in the blood and CD14 expression in the liver, leading to increased TNF-alpha production.33 In studies of rats chronically treated with ethanol, toremefene, an antiestrogen agent, showed a partial protection of liver injury through significantly alleviated both ethanol induction of the pro-oxidant enzyme CYP2E1 and ethanol reduction of the oxidant-pro-tective enzyme Se-glutathione peroxidase. In humans, Kotoh et al., 2007 demonstrated that estrogen may exacerbate nonalcoholic steatohepati-tis in women with insulin resistance.34

As in the case of estradiol, here we found that plasma cortisol levels were higher in premenopause women, suggesting an unexpected protector effect of this hormone against NAFLD, interestingly cortisol metabolism may be altered in individuals with a metabolic profile such as the one present in patients with NAFLD.35

Some limitations of our study are, firstly, the fact that it is a cross-sectional study. For that reason, the patients were not followed up and we were not able to see the progression of the liver disease. There needs to be further research, such as a cohort study that could follow a group of patients from premenopause through time to determine es-tradiol and cortisol levels and the progression of fatty liver disease. This monitoring should also be performed in a group of PCOS patients from the diagnosis of this endocrinal pathology. Another important limitation of our study is that the fatty liver disease was diagnosed by hepatic ultrasound, whereas it has been demonstrated that liver biopsy is the gold standard for the diagnosis of NAFLD. Another limitation is the fact that the value of ga-mma-glutamyl transpeptidase (GGT), a very important feature that must be determined in all patients with liver disease, was not measured in all patients.

These results suggest that NAFLD is more prevalent in postmenopausal and women with PCOS than those premenopausal ones. The estrogens may have a protective effect of against NAFLD in women. However, future studies should focus on the role of estrogen in other populations and the mechanisms that explain this association between the NAFLD and estrogens are needed.

This study was supported by Medica Sur Clinic & Foundation.

The authors have no financial arrangements with a company whose product figures significantly in the submitted.

None.

The authors have nothing to disclose.

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  NAFLD: Nonalcoholic fatty liver disease.

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  PCOS: Polycystic ovary syndrome.

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  BMI: Body mass index.

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  HOMA-IR: Homeostasis model assessment of insulin resistance.

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  NASH: Nonalcoholic steatohepatitis.

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  HCC: Hepatocellular carcinoma.

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  ROS: Reactive oxygen species.

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  LDL: Low density lipoprotein.

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  HDL: High density lipoprotein.

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  ALT: Alanine aminotransferase.

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  AST: Aspartate aminotransferase.

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  AP: Alkaline phosphatase.

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  BUN: Blood ureic nitrogen.

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  SD: Standard deviations.

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  KW: Kruskal Wallis test.

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  MS: Metabolic syndrome.