Introduction.
Chronic hepatitis C (CHC) is the primary cause of death in patients with end-stage liver disease [1]. Its prognosis depends on the accumulation of fibrosis over time due to various mechanisms of tissue damage caused by viral infection, ultimately leading to the development of cirrhosis and related complications. The development of hepatic fibrosis is most directly correlated with necro-inflammation, and several other host and viral factors have been associated with the rate of fibrosis progression [2], including older age, alcohol consumption, duration of infection, viral co-infections, steatosis, insulin resistance, and vitamin D deficiency [3].

Studies of large cohorts of patients with CHC have also found that high levels of estrogens (as observed during pregnancy) [4] are associated with decreased inflammatory activity in HCV women and that the progression of fibrosis in CHC is twice as rapid in men as in women [5], [6]. This difference has been attributed to the protective role of estrogens and has been supported by experimental and clinical data. Experimentally, estrogens were shown to have a relevant fibrosuppressive role in a rat model of dimethylnitrosamine- or pig-serum-induced liver fibrosis [7], [8]. Clinically, Di Martino et al. [9] and Codes et al. [10] showed that the progression of fibrosis increased significantly in women after menopause, whereas prolonged periods of hormone replacement therapy (HRT) were able to maintain this progression at a level similar to that in premenopausal women. In women, menarche initiates a long period of estrogen exposure that begins to decline in the premenopausal period, often leading to undetectable estrogen levels in early and late menopause. In women with CHC, this reduction in estrogen levels is accompanied by consensual fluctuation in the levels of pro-inflammatory [11], [12] and anti-inflammatory [12] cytokines that could interfere with the course of necro-inflammation and the progression of fibrosis. Much less is known about this process in men: pro-inflammatory cytokines do not fluctuate in age subgroups corresponding to female reproductive stages [11], but it remains unknown whether a relationship exists with changes in sex hormone balance in men.

Thus, the aim of our study was to evaluate the effects of differential hormonal exposure on the severity of fibrosis in men and women: we therefore compared women with CHC in different reproductive phases (reproductive age, premenopause, early menopause, late menopause) with four age-matched groups of men and investigated the correlations between fibrosis and the respective Estradiol and Testostosterone levels.

Abstract
Introduction
Chronic hepatitis C is the main cause of death in patients with end-stage liver disease. Prognosis depends on the increase of fibrosis, whose progression is twice as rapid in men as in women. Aim of the study was to evaluate the effects of reproductive stage on fibrosis severity in women and to compare these findings with age-matched men.

Materials and Methods
A retrospective study of 710 consecutive patients with biopsy-proven chronic hepatitis C was conducted, using data from a clinical database of two tertiary Italian care centers. Four age-matched groups of men served as controls. Data about demographics, biochemistry, liver biopsy and ultrasonography were analyzed. Contributing factors were assessed by multivariate logistic regression analysis.

Results
Liver fibrosis was more advanced in the early menopausal than in the fully reproductive (P<0.0001) or premenopausal (P = 0.042) group. Late menopausal women had higher liver fibrosis compared with the other groups (fully reproductive, P<0.0001; premenopausal, P = <0.0001; early menopausal, P = 0.052). Multivariate analyses showed that male sex was independently associated with more severe fibrosis in the groups corresponding to premenopausal (P = 0.048) and early menopausal (P = 0.004) but not late menopausal pairs. In women, estradiol/testosterone ratio decreased markedly in early (vs. reproductive age: P = 0.002 and vs. premenopausal: P<0.0001) and late menopause (vs. reproductive age: P = 0.001; vs. premenopausal: P<0.0001). In men age-matched with menopausal women, estradiol/testosterone ratio instead increased (reproductive age group vs. early: P = 0.002 and vs. late M: P = 0.001).

Conclusions
The severity of fibrosis in women worsens in parallel with increasing estrogen deprivation and estradiol/testosterone ratio decrease. Our data provide evidence why fibrosis progression is discontinuous in women and more linear and severe in men, in whom aging-associated estradiol/testosterone ratio increase occurs too late to noticeably influence the inflammatory process leading to fibrosis.