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International Dairy Journal 22 (2012) 3e14

Contents lists available at SciVerse ScienceDirect

International Dairy Journal


journal homepage: www.elsevier.com/locate/idairyj

Review

Impact of cows milk estrogen on cancer risk


Peter W. Parodi*
Human Nutrition and Health Research, Dairy Australia, Melbourne, Australia

a r t i c l e i n f o a b s t r a c t

Article history: Estrogens have been implicated in cancer at hormone-responsive sites, such as the breast, ovaries,
Received 20 May 2011 endometrium and prostate. Because cows milk contains estrogens, some authors have suggested that its
Received in revised form consumption may contribute to the risk of cancer at these sites. However, these reports do not recognize
12 August 2011
the complex mechanisms these cells possess to regulate estradiol levels. Hormone-responsive and many
Accepted 14 August 2011
peripheral cells contain all the necessary steroidogenic enzymes necessary for in situ synthesis of bioactive
estradiol from abundant androgenic precursors and for inactivation of unwanted estrogens. Estradiol from
dairy products is extensively inactivated in the gastrointestinal tract and only about 5% survives the rst
pass to the liver. Thus daily dairy product intake would supply only about 0.25% of the FAO/WHO upper
acceptable daily intake of estradiol. Available epidemiological evidence does not suggest an association
between dairy product consumption and risk of cancer of the breast, ovaries and endometrium.
2011 Elsevier Ltd. All rights reserved.

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4
2. Estrogens and cancer risk . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4
2.1. Observational studies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4
2.2. Experimental studies and mechanisms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4
3. Serum estrogen levels and the risk of cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .5
3.1. Breast cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
3.2. Endometrial cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
3.3. Ovarian cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
3.4. Prostate cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
4. Relationship between circulating estrogen levels and levels in breast tissue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .5
5. Relationship between urinary estrogens and estrogen metabolites in breast tissue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .6
6. Formation and transformation of tissue estrogen levels in the breast and other estrogen-responsive tissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .6
7. Breast tissue estrogen content: uptake from the circulation versus synthesis in situ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7
8. Estrogen content of cows milk . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .8
8.1. Factors influencing the hormone content of milk . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
9. Absorption and metabolism of estrogens from milk and its products . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .9
10. Physiological influence of various exogenous sources of estrogen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .9
11. Epidemiological evidence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
11.1. Breast cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
11.2. Ovarian cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
11.3. Endometrial cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
11.4. Prostate cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
11.5. Colon cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
12. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .12
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

* Tel.: 617 3359 6110.


E-mail address: peterparodi@uqconnect.net.

0958-6946/$ e see front matter 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.idairyj.2011.08.006
4 P.W. Parodi / International Dairy Journal 22 (2012) 3e14

1. Introduction 2.2. Experimental studies and mechanisms

It is generally accepted that estrogens are involved in the There is ample evidence that administration of estradiol (17b-
development of cancer at hormone-responsive sites, such as the estradiol) to laboratory animals induces tumors and that castration
breast, ovary and endometrium and, in conjunction with testos- and use of antiestrogen drugs inhibits tumor development
terone, the prostate (Bernstein & Ross, 1993; Kaaks, Lukanova, & (Bernstein & Ross, 1993; Liehr, 2000). The mechanisms whereby
Kurzer, 2002; Lukanova & Kaaks, 2005; Risbridger, Bianco, Ellem, estrogens cause cancer have not been established conclusively.
& McPherson, 2003). Even so, the mechanisms whereby estrogens A commonly held hypothesis is that circulating estrogens diffuse
inuence carcinogenesis remain largely unresolved. A number of passively through cellular membranes of estrogen-responsive cells
publications have hypothesized that estrogen present in cows milk and bind to nuclear estrogen receptors (ERs) of which there are two
may be responsible for the development of cancer in estrogen- forms, ERa and ERb. Binding to ERs stimulates transcription of
responsive organs (Farlow, Xu, & Veenstra, 2009; Ganmaa & Sato, genes involved in cell proliferation. Rapidly proliferating cells are
2005; Outwater, Nicholson, & Barnard, 1997; Qin, Wang, Kaneko, susceptible to genetic errors during DNA replication, which if
Hoshi, & Sato, 2004). However, these hypotheses fail to take into uncorrected may ultimately lead to cancer. Estradiol may also
account the complex mechanisms involved in the synthesis and increase proliferation in cells already having mutations due to other
metabolism of estrogens within the cell and the metabolic sources of DNA damage (Santen, 2002; Yager & Davidson, 2006;
processes associated with the absorption of exogenous estrogens. Yue et al., 2003).
These aspects, together with a brief outline of relevant aspects of It has also been proposed that non-receptor-mediated mutagen-
estrogen carcinogenesis, are discussed in this review. esis induced by oxidized estrogen metabolites may contribute to
carcinogenesis (Cavalieri, Devanesan, Bosland, Badawi, & Rogan,
2002a; Cavalieri et al., 2002b; Rogan et al., 2003; Yager & Davidson,
2. Estrogens and cancer risk 2006). Estrogen metabolites are formed by a series of oxidative
biotransformations, both in the liver and in extrahepatic hormone-
2.1. Observational studies responsive organs by the activities of a series of cytochrome P450
enzymes. There are two major mutually exclusive pathways for
The study of breast cancer has received more attention than the transformation of estradiol and estrone. In the rst pathway
cancer at the other hormone-responsive sites. Nevertheless, the the A ring is hydroxylated at position 2 or 4 to produce the
causes of breast cancer are still largely undetermined. Evidence catechol estrogens 2-hydroxyestradiol and 2-hydroxyestrone or 4-
from a number of observational studies suggests that several hydroxyestradiol and 4-hydroxyestrone, respectively. These four
reproductive factors, which indicate an increased exposure to metabolites may be inactivated in the liver and extrahepatic
estrogens, increase breast cancer risk. Early menarche, late meno- estrogen-responsive tissue by glucuronidation and sulfation and
pause, nulliparity and rst pregnancy late in life are associated with particularly in extrahepatic tissues by the enzyme catechol-O-
increased risk. Bilateral oophorectomy, especially at an early age, methyltransferase (COMT), which forms 2- and 4-methoxyestradiols,
reduces risk. On the other hand, oral contraceptive use that and methoxyestrones. In certain conditions catechol estrogens may
prevents ovulation and decreases serum estrogen levels results in be oxidized further to reactive catechol estrogen semiquinones then
a slight increase in risk. In contrast, breast-feeding (particularly for to catechol estrogen-2,3-quinones and catechol estrogen-3,4-
long durations, which also prevents ovulation) is associated with quinones. Protection against the catechol estrogen-quinones occurs
lower risk, but this protection may result from cell differentiation through conjugation with glutathione catalyzed by glutathione-S-
(Bernstein & Ross, 1993; Chodosh et al., 1999; Harris, Lippman, transferases. Another protection mechanism results when catechol
Veronesi, & Willett, 1992; Kuller, 1995). estrogen-quinones are re-cycled to catechol estrogens by the action of
Evidence for a role of estrogens in endometrial cancer is robust quinone reductases (Cavalieri, Frenkel, Liehr, Rogan, & Roy, 2000;
and it is considered that exposure to estrogens, unopposed by Lakhani, Sarkar, Venitz, & Figg, 2003; Yager & Davidson, 2006). The
progesterone, is a major etiological factor. Early menarche and late second major oxidation pathway occurs at the D ring with the
menopause are risk factors, whereas parity is protective. A low level formation of 16a-hydroxyestradiol and 16a-hydroxyestrone. These
of physical activity and obesity increase risk, possibly due to 16a-hydroxyderivatives may be further oxidized to estriol (Clemons &
a reduction in progesterone levels. There is an increased risk of Goss, 2001; Mueck, Seeger, & Lippert, 2002).
endometrial cancer in users of estrogen-only oral contraceptives Estrogenic and genotoxic properties of the various estrogen
and hormone replacement therapy, but no increased risk when metabolites vary from one and other and from their parent estro-
progesterone is included in the preparation (Kaaks et al., 2002; gens. For instance, the 2-hydroxyestrogens bind to ERs with afnity
Pike, Spicer, Dahmoush, & Press, 1993). comparable with that for estradiol, but have limited mitogenic
The cause of ovarian cancer is largely unknown. Observational effect and are considered antiestrogenic and anticarcinogenic
evidence on the role of estrogens is conicting and often open to (Bradlow, Telang, Sepkovic, & Osborne, 1996; Mueck et al., 2002). 2-
alternative interpretation. There is some support for the concept Methoxyestradiol has very low afnity for ERs, but exhibits potent
that continual ovulation may predispose women to development of apoptotic activity against rapidly growing cancer cells, possesses
ovarian cancer. Pregnancy, breast-feeding and oral contraceptive antiangiogenic properties and has been trailed on patients
use are protective, but hormone replacement therapy is associated with breast and prostate cancer (Lakhani et al., 2003). On the other
with increased risk (Lukanova & Kaaks, 2005; Risch, 1998). hand, 16a-hydroxyestrogens that bind to ERs with lower afnity
Although androgens are considered to have an important role in than estradiol are thought to have a carcinogenic inuence (Mueck
prostate cancer development, animal and human cell culture et al., 2002).
studies now suggest a role for estrogens (Bosland, 2004; Harkonen Attention has focused on the estrogen-2,3- and estrogen-3,4-
& Makela, 2004). Recent studies with genetically modied mouse quinones because of their ability to act both as electrophiles and
models show that malignant changes to the prostate gland are oxidants. As electrophiles, catechol quinones can form covalent
dependent on both androgen and estrogen responses, and that adducts with the purine bases adenine and guanine of DNA, these
neither hormone alone can produce an aberrant growth pattern depurinating adducts generate apurinic sites that may lead to
that leads to malignancy (Bosland, 2004; Risbridger et al., 2003). mutations and in turn cancer. As oxidants, catechol quinines redox
P.W. Parodi / International Dairy Journal 22 (2012) 3e14 5

cycle with their semiquinones, producing the superoxide anion and or luteal estrone levels and the risk of breast cancer (Eliassen &
reactive oxygen species that may damage lipids and DNA (Cavalieri Hankinson, 2008).
et al., 2000; Yager & Davidson, 2006). The proposed ER- and non- Two prospective studies found positive associations between
ER-mediated pathways may act in concert in an additive or urinary estrogen levels and risk of postmenopausal breast cancer
synergistic manner to cause cancer (Yue et al., 2003). However, (Key et al., 1996; Onland-Moret et al., 2003). However, the rele-
under normal physiological conditions deactivation and conjuga- vance of urinary estrogens to functions in target cells is uncertain.
tion processes should ensure that insufcient reactive quinines are Seven prospective studies examined associations between either
produced to cause signicant DNA damage, whereas normal serum or urinary estrogen metabolites and risk of breast cancer in
cellular antioxidants should prevent oxidative damage. High levels both pre- and postmenopausal women. These studies measured
of hydroxy- and methoxyestrogens are found in blood (Eliasssen, only 2-hydroxyestrone and 16a-hydroxyestrone and overall there
Missmer, Tworoger, & Hankinson, 2008) and urine (Eliassen et al., were no statistically signicant associations between these two
2009). Indeed, hydroxy- and methoxyestrogens can represent up metabolites or the ratio of 2-hydroxyestrone: 16a-hydroxyestrone
to 95% of the total estrogens in normal breast tissue (Castagnetta and the risk of breast cancer (Arslan et al., 2009; Eliassen, Missmer,
et al., 2002; Rogan et al., 2003). Overall, no studies have demon- Tworoger, & Hankinson, 2008).
strated that estrogen metabolites contribute to cancer in humans
(Yager & Davidson, 2006). 3.2. Endometrial cancer

3. Serum estrogen levels and the risk of cancer There has been only one large prospective study on circulating
sex hormone levels and the risk of endometrial cancer (Lukanova
Many studies have examined the associations between circu- et al., 2004). This study combined data for postmenopausal
lating estrogen levels and the risk of breast, endometrial, ovarian women from the NYUWHS, ORDET and NSHDS cohorts. Estradiol
and prostate cancer. Because these cancers produce estrogens and estrone levels were strongly associated with endometrial
during their development only studies where blood was drawn cancer risk with RRs of 4.13 (1.76e9.72) and 3.67 (1.71e7.88),
prospectively will be considered here. Estradiol circulates bound to respectively. SHBG was negatively associated with risk of endo-
carrier proteins with around 70% bound to sex hormone-binding metrial cancer, RR 0.46 (0.02e1.05).
globulin (SHBG), which inhibits it bioactivity, and 30% is more
loosely bound to albumin. Only about 1% circulates in the free form 3.3. Ovarian cancer
(Grow, 2002). Eliassen and Hankinson (2008) recently reviewed
seroepidemiology studies of breast, endometrial and ovarian Eliassen and Hankinson (2008) reviewed the few prospective
cancer, and the Endogenous Hormones and Prostate Cancer studies that investigated associations between serum sex hormone
Collaborative Group (2008) conducted a pooled analysis of 18 concentrations and the risk of ovarian cancer. Overall, there is no
prospective studies, which examined the associations between sex convincing evidence for an association between circulating levels of
hormone concentrations and the risk of prostate cancer. estrogens or androgens and the risk of ovarian cancer in pre- or
postmenopausal women.
3.1. Breast cancer
3.4. Prostate cancer
The Endogenous Hormones and Breast Cancer Collaborative
Group (2002) analyzed data from nine prospective studies of 663 The Endogenous Hormones and Prostate Cancer Collaborative
postmenopausal women who developed breast cancer. The relative Group (2008) conducted a collaborative analysis of worldwide
risk (RR) with 95% condence intervals (CI) for the highest data from 18 cohort studies that investigated the association
compared with the lowest quintile of serum estradiol, free estra- between endogenous hormone concentrations and the risk of
diol, estrone and estrone sulfate was 2.00 (1.47e2.71), 2.58 prostate cancer in 3886 men with incident prostate cancer and
(1.76e3.78), 2.19 (1.48e3.22) and 2.00 (1.26e3.16), respectively. For 6438 control subjects. There were no statistical signicant associ-
SHBG, the RR was 0.66 (0.43e1.00). Since this analysis, results for ations between serum concentrations of any androgens or estradiol
combined data from the Swedish Malmo Diet and Cancer Study and the risk of prostate cancer. For estradiol the RR for the highest
(MDCS) and the Northern Sweden Health and Disease Study versus lowest level of estradiol was 0.93 (0.77e1.11) and 0.95
(NSHDS), the New York University Womens Health Study (0.79e1.15) for free estradiol. An investigation of the possibility that
(NYUWHS), the Nurses Health Study (NHS), and the European a combination of estrogens and androgens may be more strongly
Prospective Investigation into Cancer and Nutrition (EPIC), were associated with the risk of prostate cancer than either estrogen or
reported and are reviewed by Eliassen and Hankinson (2008) androgen alone, found no evidence of any interaction.
Results from the Studies of Hormones and Diet in the Etiology of
Breast Tumors (ORDET) were recently reported (Sieri et al., 2009). 4. Relationship between circulating estrogen levels
In all these studies, there was roughly a two-fold increase in risk of and levels in breast tissue
breast cancer with elevated levels of estrogens.
Only a few studies have reported associations between circu- To date, the strongest consistent evidence for a role of circu-
lating estrogens and the risk of premenopausal breast cancer. This lating estrogens in cancer is for postmenopausal breast cancer.
is undoubtedly due to the difculty in obtaining a representative However, circulating hormone levels may not reect hormone
sample for menstruating women. Five small studies found no concentrations in target cells. Even so, epidemiological associations
association between estrogen levels and the risk of breast cancer. cannot be used to ascribe causality and improved knowledge of
Results from two recent large studies indicate that in the EPIC estrogen metabolism at the cellular level is required.
cohort there was no association between estradiol or estrone levels Several studies have compared estrogen levels in normal and
and the risk of breast cancer. On the other hand, in NHS II there was malignant breast tissue with the levels in blood from pre- and
a signicant positive association between breast cancer risk and postmenopausal women (Chetrite, Cortes-Prieto, Philippe, Wright,
estradiol levels in blood collected during the follicular phase, but & Pasqualini, 2000; Geisler, 2003; van Landeghem, Poortman,
not for luteal estradiol. There was no association between follicular Nabuurs, & Thijssen, 1985; Pasqualini et al., 1996; Thijssen, van
6 P.W. Parodi / International Dairy Journal 22 (2012) 3e14

Landeghem, & Poortman, 1986). Even though serum estrogen levels metabolites of the 2- and the 16-hydroxylation pathways were
are up to 50-fold lower in postmenopausal than in premenopausal lower, the latter signicantly lower. The ratio of 2:16 hydroxylation
women, their unconjugated estrogen levels in normal and malig- products was nonsignicantly higher in breast tissue than in urine.
nant breast tissue are similar. Estradiol levels are some 2.5-fold There were large inter-individual variations in tissue to urine ratios
higher in breast cancer tissue than in normal tissue for both pre- of estrogens and their metabolites. It is possible that not all urinary
and postmenopausal women. In breast cancer tissue estradiol estrogens and their derivatives originated from breast tissue.
levels are about 10- to 20-fold higher and estrone levels 2- to 10-
fold higher than their corresponding serum levels. Available 6. Formation and transformation of tissue estrogen levels
studies suggest there is no simple correlation between the levels of in the breast and other estrogen-responsive tissues
estrogens in serum and their level in tissue. These data suggest that
postmenopausal breast tissue has the ability to selectively accu- There is now substantial evidence that normal and malignant
mulate estrogens from the circulation or to synthesize them within breast tissue contains all the necessary steroidogenic enzymes
the tissue. necessary for the synthesis of estradiol from circulating inactive
androgenic precursors (Labrie, 2003; Labrie et al., 2003; Pasqualini,
5. Relationship between urinary estrogens and estrogen 2004; Perel, Wilkins, & Killinger, 1980). Dehydroepiandrosterone
metabolites in breast tissue (DHEA) sulfate (S) is by far the predominant adrenal-derived
circulating precursor hormone in men and women. Its concentra-
Only one study was found that compared the levels of estrogens tion is from 1000 to 10,000 times higher than that of estradiol and
and estrogen metabolites in breast tissue with those in urine. 100e500 times higher than that of testosterone. There is also
In a small study, Taioli et al. (2010) found that total estrogen considerable estrogen sulfate and lesser amounts of androstene-
metabolite levels were similar between breast tissue and urine. dione and androstenediol in serum. Initially, DHEA-S is converted
However, the levels of individual components were quite different to DHEA by the enzyme steroid sulfatase (Fig. 1). DHEA can
between the two sources. The sum of estrone and estradiol was then take part in two pathways. Reduction by members of the
signicantly higher in breast tissue than urine, while the 17b-hydroxysteroid dehydrogenase (17b-HSD) family produces

Fig. 1. Outline of the in situ formation of estrogens from circulating precursors, DHEA-S and estrone sulfate (adapted from Labrie et al., 2003). DHEA, dehydroepiandrosterone;
DHEA-S, DHEA sulfate; 17b-HSD, 17b-hydroxysteroid dehydrogenase; 3b-HSD, 3b-hydroxysteroid dehydrogenase.
P.W. Parodi / International Dairy Journal 22 (2012) 3e14 7

androstenediol (17b-HSD types 1,3,5 and 7 are involved in reduc- was signicantly associated with increased risk of recurrence and
tion reactions, whereas 17b-HSD types 2,4 and 6 are involved in worsened prognosis (Suzuki et al., 2003).
oxidation reactions). The enzyme 3b-hydroxysteroid dehydroge- Steroid sulfatase inhibitors blocked the ability of estrogen
nase (3b-HSD) converts both DHEA and androstenediol to andros- sulfatase to stimulate the growth of carcinogen-induced mammary
tenedione and testosterone, respectively, which, in turn, are tumors in ovariectomized rats (Purohit, Woo, Potter, & Reed, 2000).
converted to estrone and estradiol by the aromatase enzyme. The MCF-7 breast cancer cells over-expressing steroid sulfatase injected
DHEA / androstenediol, androstenedione / testosterone and into the anks of ovariectomized nude mice were used to
estrone / estradiol conversions are interconvertable, depending demonstrate the effect of steroid sulfatase inhibitors in reducing
on the oxidative/reductive forms of 17b-HSD available, but the steroid sulfatase activity and tumorogenicity (Foster et al., 2006).
action of aromatase is irreversible. Estrone and estradiol may also A recent initial trial in postmenopausal women with ER metastatic
be formed by the action of steroid sulfatase on estrone sulfate and breast cancer demonstrated that a steroid sulfatase inhibitor almost
estradiol sulfate, respectively. completely blocked steroid sulfatase activity in peripheral blood
Although not studied to the same extent as breast tissue, lymphocytes and tumor tissue (Stanway et al., 2006). A seroepi-
endometrial and ovarian tissue also contain a range of steroido- demiology study by Dorgan et al. (1996) did not detect an associ-
genic enzymes capable of synthesizing estrogens from circulating ation between serum estrone or estrone sulfate levels and the risk
precursors (Ito, Utsunomiya, Yaegashi, & Sasano, 2007; Sasano & of breast cancer. However, the ratio of estrone sulfate to estrone
Harada, 1998). The prostate likewise possesses all the necessary was signicantly inversely associated with risk, suggesting that
enzymes to convert circulating DHEA into estradiol and testos- women who develop breast cancer may be less able to metabolize
terone, as well as 5a-reductase to convert testosterone to the more estrone to its inactive form. How estrogen activation and deacti-
bioactive dihydrotestosterone (Takase et al., 2006). vation mechanisms in cells become unbalanced is unknown at
Normal cells possess mechanisms to ensure estradiol homeo- present.
stasis. ER expression in normal breast epithelial cells is inversely
related to circulating estradiol levels (Khan et al., 1999; Soderqvist, 7. Breast tissue estrogen content: uptake from the circulation
von Schoultz, Tani, & Skoog, 1993). Estrone and estradiol can be versus synthesis in situ
converted to inactive sulfates by the action of steroid sulfo-
transferase and either excreted from the cell or retained for future Levels of estradiol in normal and in malignant breast tissue are
supply of estrone and estradiol. Conjugation of estrone and estra- similar for pre- and postmenopausal women, despite the large
diol by uridine diphosphate-glucuronosyltransferases inactivates differences in serum levels (Thijssen et al., 1986; van Landeghem
and produces the more soluble estrogen glucuronides that are et al., 1985). High estradiol levels in postmenopausal breast tissue
excreted to bile and urine. O-methylation of hydroxyestrogens by may result from enhanced uptake from the circulation or from in
COMT and conjugation of estrogen quinones with glutathione situ aromatization of androgens and from estrogen sulfate via the
facilitates removal of estrogen metabolites from cells. sulfatase/17b-HSD pathway. It is estimated that about 75% of
The expression and level of the various steroidogenic and estrogens in premenopausal women are synthesized in peripheral
metabolizing enzymes in each cell of hormone-responsive tissues tissues and close to 100% after menopause (Labrie, 1991, 2003).
may inuence their estrogen content and potential to transform. However, it is extremely difcult to calculate the relative contri-
For instance, a higher activity of the reducing forms of 17b-HSD butions of uptake from the circulation versus in situ synthesis for an
may lead to higher levels of estradiol. Aromatase is important for individual organ or tissue, such as the breast.
the conversion of androstenedione to estrone and testosterone to A number of lines of indirect evidence suggest a role for in situ
estradiol. Aromatase inhibitors were shown to suppress blood and estrogen synthesis in tumor development that include:
breast cancer tissue estrogen levels in postmenopausal women by
80e95% (Geisler, 2003). However, in both pre- and postmenopausal  The presence of the necessary steroidogenic enzymes in breast
breast cancer patients estrone sulfatase activity was found to be tissue (Labrie, 2003; Pasqualini, 2004).
50e200 times higher than aromatase activity, with activity of both  Levels of DHEA, the androgen precursor of estrogens, are many-
enzymes greater in post- than in premenopausal women fold higher in normal and malignant breast tissue for both pre-
(Pasqualini et al., 1996). Enzyme levels were higher in tumors than and postmenopausal women than in their serum (Labrie, 1991;
in adjacent tissue, which, in turn, were higher than in normal tissue Thijssen et al., 1986).
(Chetrite et al., 2000). Tissue levels of estrone, estradiol and their  ER breast tumor tissue contains estradiol (Edery et al., 1981).
sulfates reected the enzyme activities. It is considered that  There is little correlation between serum estradiol levels and
formation of estradiol from estrone sulfate is far more important breast epithelia proliferation (Khan et al., 1999).
than aromatization in breast tissue (Chetrite et al., 2000; Pasqualini
et al., 1996; Santner, Feil, & Santen, 1984). Yue, Wang, Hamilton, Demers, and Santen (1998) developed an
Several lines of evidence suggest that abnormal regulation of animal model where aromatase (A)- and sham (A)-transfected
the opposing action of estrogen sulfatases and estrogen sulfo- human MCF-7 breast cancer cells were inoculated into ovariecto-
transferases in breast tissue may be responsible for carcinogenesis. mized nude mice, which lack peripheral aromatase activity. With
Falany and Falany (1996) found that estrogen sulfotransferase was the injection of androstenedione and the use of Silastic implants to
present in human normal mammary epithelial cells, but was produce various estradiol levels, it was determined that under
absent in ER and ER breast cancer cell lines. Qian, Deng, and physiological conditions, reecting those in postmenopausal
Song (1998) showed that the estrogen-responsive human MCF-7 women, in situ aromatization made a major contribution to estra-
breast cancer cell line lacks estrogen sulfotransferase activity. diol levels in breast cancer tissue. Previously, this group showed
When this cell line was transfected with estrogen sulfotransferase that rats bearing chemically induced hormone-dependent tumors
cDNA, estrogen-stimulated DNA synthesis and cell proliferation synthesized about 25% of their estrogen content in situ by the
was inhibited (Qian et al., 1998). Estrogen sulfotransferase estrogen sulfatase pathway (Masamura, Santner, & Santen, 1996).
immunoreactivity in human breast carcinoma was signicantly A few studies have determined the contribution that local in situ
associated with a decreased risk of recurrence or improved estrone synthesis made to the estrogen content of normal and
prognosis. On the other hand, steroid sulfatase immunoreactivity malignant breast tissue in postmenopausal women using a double
8 P.W. Parodi / International Dairy Journal 22 (2012) 3e14

isotopic technique. Miller et al. (1998) infused 11 patients with Kensinger (2007) measured individual estradiol levels in milk
14
C-labeled estrone and 3H-labeled androstenedione, 18 h before from 206 cows in the Pennsylvania State University dairy herd
breast tissue and peripheral blood were taken for analysis. There using RIA. Mean estradiol levels ranged from undetectable (limit of
was considerable variation between breasts with the contribution quantication 0.7 pg mL1) to 22.9 pg mL1, with a mean value
of in situ synthesis ranging from 0 to 75%. In a subsequent study 1.4  0.2 pg mL1. Twelve samples of milk (4 whole, 5 half-
this group measured the percentage contribution of local skimmed, 3 skimmed) were collected from a French supermarket
estrone synthesis in both normal and tumor tissue from 24 post- and analyzed using GCeMS/MS (Courant et al., 2007). The sum of
menopausal women. Again, there were marked variations between free plus conjugated estradiol ranged from 9.8 to 44.3 pg mL1, with
specimens from different patients and the relative proportion of a mean value of 23.0 pg mL1. For estrone the corresponding values
synthesis to uptake was higher in tumors (0e92%, median value were 75.8e277.5 pg mL1 and 152.8 pg mL1, respectively. Eighty
29%) than in normal tissue (0e58%, median value 15%). There was four percent of estradiol and 96% of estrone were present in the
a positive correlation between uptake in normal and malignant conjugated form. Vicini et al. (2008) conducted the most compre-
tissue (Larionov, Berstein, & Miller, 2002). A small study by Reed hensive survey, with the analysis of 334 samples of retail whole
et al. (1989) using a similar isotopic infusion technique found that milk collected from 48 states in the USA during a three-week
in 3 of 7 subjects there was no or little in situ formation of estrone. period. RIA, with a sensitivity of 0.8 pg mL1, was used for the
For the other 4 subjects in situ estrone synthesis appeared to assay. The mean value for estradiol was 4.97  0.239 pg mL1.
contribute around 85% of total estrone content in tumor tissue Recently, Farlow et al. (2009) measured estrogen levels in single
while the value for normal tissue was 71%. samples of whole, 2% fat and skim milk in the US using LC-MS/MS
Breast tissue can also obtain signicant quantities of estradiol (limit of quantitation 0.2 pg mL1). Levels of free estradiol and
from inltrating macrophages. Macrophages are present in normal estrone were 1.0, 1.8, and 2.4 pg mL1 and 10.3, 6.8 and 3.8 pg mL1,
resting, pregnant, and lactating breast tissue with levels respectively. Corresponding values for total estradiol and estrone
increasing after involution, but probably do not exceed 20% of the were 22.1, 22.2, 20.4 pg mL1 and 85.5, 84.6, 65.0 pg mL1,
total cell population (Ferguson, 1985). A study by Kelly, Davison, respectively.
Bliss, and McGee (1988) found malignant tissue contained the
highest concentrations of macrophages with levels around 50% of 8.1. Factors inuencing the hormone content of milk
the tumor mass. Levels in benign tissue were around one-half this
value. Macrophages contain the necessary steroidogenic enzymes, A number of factors can inuence the hormone content of milk.
including aromatase, to convert the ubiquitous adrenal hormone Both estradiol and estrone levels increase during the estrous cycle,
DHEA into estradiol (Mor et al., 1998; Schmidt, Kreutz, Lofer, reaching peak values at estrus, thereafter decreasing to the end of
Scholmerich, & Straub, 2000). Estrogen production by macro- the cycle (Gyawu & Pope, 1990; Monk, Erb, & Mollett, 1975).
phages is sufcient to stimulate the proliferation of adjacent Hormone levels can also vary with stage of lactation (Gyawu &
epithelial cells (Mor et al., 1998). Pope, 1990). There is considerable inter-individual variation
within a heard and values for individual cows uctuate markedly
8. Estrogen content of cows milk from week to week (Gyawu & Pope, 1990; Henderson, Karanikolas,
Kenealy, & Macmillan, 1994a).
Estrogen levels in cows milk and factors that inuence them The estrogen level in milk increases during pregnancy.
have been inadequately studied. The objective of early studies was Henderson et al. (1994a) measured the levels of estrone sulfate in
to use estrogen levels as a tool to determine estrus and pregnancy. milk from four herds. Levels rose progressively during pregnancy
Early methods that utilized bioassays, colorimetry and spectro- from a mean value of 80e100 pg mL1 at 60e80 d of pregnancy to
uorimetry were crude and had a high limit of detection. Intro- a plateau value of around 1000 pg mL1 at 181e200 d. For non-
duction of radioimmunoassay (RIA) improved sensitivity, but pregnant cows in the herds, estrone sulfate concentration ranged
factors such as loss of hormones during removal of triglycerides, from non-detected to 110 pg mL1 with a mean value of 59 pg mL1
cross reactivity of antibodies with other hormones, incomplete Malekinejad et al. (2006) reported that the level of free estrone in
hydrolysis of hormone conjugates and lack of pure standards still raw milk from non-pregnant cows and cows in the rst, second and
affected results. Recently, improvement in extraction techniques third trimester of gestation was 6.2, 9.2, 57 and 118 pg mL1,
coupled with improved RIA assays and measuring techniques like respectively. Corresponding values for free estradiol were 5.6, 10.2,
liquid chromatography (LC) or gas chromatography tandem mass 20.4 and 21.3 pg mL1, respectively. For total estrogens
spectrometry (GCeMS/MS) have further advanced sensitivity and (free conjugated and for different cows from the above), estrone
specicity. However, these techniques are complex, time levels in the rst, second and third trimester milk were 7.9, 452 and
consuming and expensive, which has limited sample numbers. 1266 pg mL1, and for estradiol 18.6, 51.4 and 51.2 pg mL1,
Only studies that utilized these improved techniques are reviewed respectively.
here. Unfortunately sample sizes for the different types of milk Pape-Zambito et al. (2007) found that mean free estradiol levels
were generally small. of milk from non-pregnant cows, early pregnant cows (1e140 d)
Hartmann, Lacorn, and Steinhart (1998) measured the hormone and mid-pregnant cows (141e210 d) were 1.3, 0.9 and 3.0 pg mL1,
content of German whole milk (sample numbers unknown) using respectively. In a further study (Pape-Zambito, Magliaro, &
GCeMS. The level of total estradiol was less than 20 pg mL1, which Kensinger, 2008) a group of cows was followed throughout preg-
was the limit of detection. Total estrone was present at around nancy. Milk free estrone concentrations averaged 0.6, 7.9 and
130 pg mL1. Four samples of milk from non-pregnant cows of the 27.1 pg mL1 during the rst, second and third trimester, respec-
Utrecht University herd were analyzed by LC-MS/MS. Values for tively. Corresponding values for estradiol were 0.3, 0.9 and
free estrone were 23, 22, 27 and 24 pg mL1, whereas values for 5.0 pg mL1, respectively.
total estrone (free conjugated) were 162, 208, 174, and Free estrogens are lipophilic and Pape-Zambito et al. (2008) and
243 pg mL1. Free estradiol values were 5 pg mL1, not detected Pape-Zambito, Roberts, and Kensinger (2010) found both estrone
(limit of detection 5 pg mL1), 5 pg mL1, and not detected, while and estradiol levels were signicantly correlated with the fat
total estradiol values were 10, 10, 9 and 11 pg mL1 (Malekinejad, content of whole milk. Malekinejad et al. (2006) found that for 0%,
Scherpenisse, & Bergwerff, 2006). Pape-Zambito, Magliaro, and 1.5% and 3.5% fat milk the free estrone content was 8.2, 17.1 and
P.W. Parodi / International Dairy Journal 22 (2012) 3e14 9

20.0 pg mL1, respectively. Corresponding values for estradiol were These values were due largely to the relatively bio-inactive estrone
10.3, 13.9 and 20.6 pg mL1, respectively. On the other hand, and both estrone and estradiol were predominantly present as
another study found no relationship between total estrogen inactive conjugates. Based on recent studies with sensitive assays
content and the fat content of milk (Courant et al., 2007). This is (Farlow et al., 2009; Malekiunejad et al., 2006; Pape-Zambito et al.,
undoubtedly due to the fact that conjugated estrogens, by far the 2007, 2008, 2010; Vicini et al., 2008), it appears unlikely that free
major component of milk estrogens, are more soluble in aqueous estradiol levels in commercial milk would exceed 5.0 pg mL1.
solution. Pasteurization and homogenization of raw milk do not Thus, consumption of 1.5 L of milk or equivalent dairy products per
appear to affect the estrogen content (Pape-Zambito et al., 2010). day would result in a daily estradiol intake of not more than 7.5 ng,
Ganmaa and Sato (2005), Ganmaa, Wang, Qin, Hoshi, and Sato of which no more than about 0.38 ng would survive rst pass
(2001), Maruyama, Oshima, and Ohyama (2010), Qin et al. (2004) metabolism to the liver. This value must be considered in the
and others have suggested that compared with milk produced 100 context of daily excretion rates for estradiol in humans. In
years ago, modern herd bulk milk contains a larger proportion of premenopausal women the daily production of estradiol varies
milk from cows in the latter half of pregnancy, and thus has elevated between 0.08 and 1.00 mg, depending upon the phase of the
levels of estrogens, which may contribute to reproductive disorders menstrual cycle (Hsueh & Billig, 1995). With the cessation of
and cancer at hormone-responsive sites. However, as noted above, ovarian function after menopause, daily estradiol production
the increased estrogen levels during late pregnancy is mostly due to decreases to around 12 mg, which is mainly derived from estrone,
estrone sulfate, which is largely biologically inactive. In addition, now the predominant estrogen and produced in peripheral tissues
during the past 60e70 years milk production per cow has increased tissue from androstenedione by the action of the aromatase
about 5-fold due to improved genetics for milk production coupled enzyme (Golden & Monroe, 1986). Production rates for estrone
with changes in nutritional management (Capper, Cady, & Bauman, range from 40 mg d1 for slender women to 200 mg d1 for obese
2009). Estradiol concentrations are negatively correlated with milk women (ODell, 1995). In men estradiol production is about
yield (Pape-Zambito et al., 2008). This effect is probably due to the 45 mg d1 (Kaufman & Vermeulen, 2005). Estradiol production in
fact that high feed intake increases liver blood ow and metabolic prepubertal children has not been determined directly, but it is
clearance rate of estradiol by the liver (Sangsritavong, Combs, considered values will be low (Andersson & Skakkebaek, 1999).
Sartori, Armentano, & Wiltbank, 2002). Thus, in the case of premenopausal women, intake of around
Pape-Zambito et al. (2008) estimate that the majority of cows 7.5 ng d1 of estradiol from 1.5 L of milk or equivalent would
producing milk for consumption would have less than 2 pg mL1 of represent merely 0.00076e0.0094% of their daily production and
estradiol. This estimate is based on their studies with herd milk and only 5% of this intake would survive the rst pass of the liver in
milk from pregnant cows and because early lactation cows have a bioactive form. The percentages for postmenopausal women,
high milk yields and farmers generally dry off cows at 60 days prepubertal children and men would be higher.
before the next expected calving. Thus, there is no evidence that the The Joint FAO/WHO Expert Committee on Food Additives (WHO,
levels of estrogen in modern commercial milk are higher than 100 2000) established an acceptable daily intake for estradiol of
years ago. 0e50 ng kg1 of body weight. This gure was based on changes in
several hormone-dependent parameters in postmenopausal
9. Absorption and metabolism of estrogens from women. A safety factor of 10 was used to account for normal
milk and its products variation among individuals, and an additional factor of 10 was
added to protect sensitive individuals. Accordingly, estradiol
Estradiol in consumed dairy products is absorbed from the consumption from dairy products should account for only around
intestine. The intestinal mucosa contains enzymes capable of the 0.25% of the FAO/WHO upper acceptable daily intake value.
transformation of estradiol to estrone and the formation of estrogen Milk contains estrone of which around 90% is estrone sulfate
sulfate and glucuronide conjugates. These metabolites and any free (Henderson, Camberis, Simmons, Starrs, & Hardie, 1994b). Estrone
estradiol pass to the portal circulation and rapidly pass to the liver has low biological activity and estrone sulfate is inactive. It is
where near-complete metabolism to estrone, estrone sulfate (the possible that some of the small quantity of estrone sulfate in milk
major conjugate) and some estrogen glucuronides and hydroxyl- that survives metabolism during rst pass to the liver may be
ated conjugates occurs (Lobo & Cassidenti, 1992; Longcope et al., converted to estrone by sulfatases then to estradiol by 17b-HSD.
1985). Part of the conjugated estrogens pass to the kidneys and However, milk-derived estrones are but a small fraction of a large
are excreted in urine. Most of the remainder is excreted with bile to circulating pool. For instance, for postmenopausal women in the
the intestinal lumen. Here, some of the conjugated estrogens may be Nurses Health study (Hankinson et al., 1995) mean concentrations
deconjugated by bacterial b-glucuronidases, but most of these are of estrone and estrone sulfate were 18- and 98-fold higher than
re-conjugated by intestinal steroid sulfotransferases and glucur- bioavailable estradiol. In addition, there are vastly higher levels of
onosyltransferases and along with the majority of the remaining circulating DHEA-S (Labrie, 2003), and even higher levels in breast
bile-derived estrogen conjugates are re-absorbed and pass to the tissue (Thijssen et al., 1986), which can be bioconverted to estrone
liver (enterohepatic circulation). A portion escapes this enter- and estradiol if required by the cell. Overall, estrogen-sensitive cells
ohepatic circulation and is excreted in the faeces. Any free estradiol possess multiple mechanisms to obtain estradiol for their growth
will be conjugated at the next pass of the liver (Adlercreutz & and maintenance and to inactivate surplus to requirements.
Martin, 1980; Adlercreutz, Martin, Jarvenpaa, & Fotsis, 1979). Over- Coupled with the relatively minute amount of dairy product-
all, it is estimated that as a result of metabolism by the intestinal derived estradiol surviving intestinal absorption and rst pass
mucosa and rst pass to the liver, the bioavailability of oral estradiol liver metabolism, this makes it unlikely that estradiol from this
is only in the range of 2e5% (Dusterberg, Schmidt-Gollwitzer, & source would inuence cancer development.
Humpel, 1985; Kuhnz, Gansau, & Mahler, 1993).
As part of a German market basket survey, Hartmann et al. 10. Physiological inuence of various exogenous
(1998) determined the natural occurrence of steroid hormones in sources of estrogen
food. Based on previously published national nutritional data they
estimated the daily intake of estrogens (estrone plus estradiol) The estrogen content of milk, its contribution to the bodys
from milk products was 0.06 mg for men and 0.05 mg for women. estrogen pool and its physiological signicance may be viewed in
10 P.W. Parodi / International Dairy Journal 22 (2012) 3e14

relation to the inuence of a number of external factors on serum 0.625 mg d1 of conjugated equine estrogens (CEE) had serum
estrogen levels. Many factors have been studied but BMI appears to estrogen levels measured at baseline and after 1 year of therapy
be the most important determinant. Data from the Nurses Health (Edlefsen et al., 2010). Estradiol levels increased from 13.3 to
Study (Hankinson et al., 1995) showed that in postmenopausal 35.5 pg mL1, bioavailable estradiol from 8.9 to 16.1 pg mL1 and
women serum estrogen levels were positively associated with BMI. free estradiol from 0.4 to 0.6 pg mL1. Estrone levels increased from
Mean serum estradiol levels ranged from 4.7 pg mL1 in the lowest 41.4 to 149.9 pg mL1. After a mean of 10.7 years of follow-up in the
quintile of BMI (<21 kg m2) to 10.0 pg mL1 in the highest quintile intervention phase of the Womens Health Initiative randomized
(29 kg m2). For bioavailable estradiol, the corresponding values controlled trial (LaCroix et al., 2011), the hazard ratio for breast
were 0.8 and 3.3 pg mL1. A pooled re-analysis of individual data cancer in postmenopausal women who used 0.625 mg d1 of CEE
from 8 prospective studies showed similar variations (Endogenous for a median of 5.9 years compared to non-users was 0.77
Hormones and Breast Cancer Collaborative Group, 2003). (0.62e0.95) Three other small randomized controlled trials of
Dietary components may inuence intestinal function and the estrogen-only therapy reviewed by Collins, Blake, and Crosignani
bacterial population. Changes in the number of b-glucuronidase- (2005), found no excess risk compared with placebo. Epidemio-
producing bacteria can inuence deconjugation of estrogens, affect logical evidence from the large Nurses Health Study suggested that
the enterohepatic circulation and the level of estrogens excreted in women who had undergone a hysterectomy and used estrogen-
faeces and urine and their level in blood (Goldin & Gorbach, 1994). only therapy for less than 20 years did not have an increased risk
Many studies investigated the role of dietary components such as of developing breast cancer. However, longer duration of usage was
fat, protein and ber, on serum estrogen levels, but most suffered associated with higher risk, primarily for ER and progesterone
from methodological faults, small sample size and lacked controls. receptor positive breast cancers (Chen et al., 2006).
The Nurses Health Study found associations between different Use of oral contraceptives by premenopausal women causes
dietary patterns and serum estrogen levels in postmenopausal a suppression of ovarian function. As a result the elevated levels of
women, but the associations were lost after adjusting for BMI estradiol during mid-cycle and again during the mid-luteal phase
(Fung, Hu, Barbieri, Willett, & Hankinson, 2007). The effect of are suppressed several fold to levels at or below levels found in the
animal products and their nutrient components on circulating rst half of the follicular phase in non-users. A similar suppression
estrogens levels in postmenopausal women was investigated in the occurs for estrone (Carr & Bradshaw, 1998; Gaspard, Dubois, Gillian,
large Melbourne Collaborative Study (Brinkman et al., 2010). None Franchimont, & Duviver, 1984). In addition, in breast tissue, oral
of the food items or nutrients explained more than 1% of the total contraceptive use suppressed estradiol levels 7.8-fold and estrone
variation in concentration of any steroid hormone. levels 2.9-fold compared to non-users (OBrien, Anandjiwala, &
For clinical intervention studies, Carruba et al. (2006) random- Price, 1997). However, epidemiological evidence for women using
ized postmenopausal women to receive their normal diet or oral contraceptives and were subjected to these large reductions in
a traditional Mediterranean diet (intervention). After 6 months, the serum and tissue estrogens does not show a reduced risk for
urinary total estrogen concentration in the intervention group subsequent breast cancer (Hunter et al., 2010; Kahlenborn,
decreased by 40%. This decrease was largely driven by the estrogen Modugno, Potter, & Severs, 2006).
metabolites 2-hydroxy- and 16-ketoestradiol and 17-epiestriol. Around 30 years ago, several observation studies suggested that
Estradiol represented only 1.16% of total estrogens and this level hormonal changes in women might be associated with the risk of
was signicantly higher than the baseline value. However, during colon cancer. However, the results of these studies were not
the study, subjects in the intervention group consumed signi- consistent. During the period 1961e1990 the sex-specic rates for
cantly less calories than at baseline. The intervention group of colon cancer incidence in men increased by 16%, whereas in women
postmenopausal women in the large Womens Health Initiative the incidence declined by 21% (Potter, 1995). This inequality was
randomized controlled dietary modication trial consumed a low- considered to be due to hormone differences. HRT use was
fat diet with increased intake of fruit, vegetables and grains considered a possible determinant of the benecial effect in
compared to their normal diet. Serum estrogen levels were women, and a number of epidemiological studies investigated this
measured in a sub-group of women at baseline and after 1 year of relationship. Hebert-Croteau (1998) conducted a meta-analysis of
intervention. Estradiol concentrations decreased from 7.6 to 11 case-control studies, 7 cohort studies and a randomized
6.7 pg mL1. However, during this period there was a decrease of controlled trial. A summary RR of 0.85 (0.73e0.99) was found
2.4 kg in the weight of women in the intervention group (Prentice between ever versus never users of HRT and the risk of colon
et al., 2006). A change in diet to one low in animal fat and rened cancer. The estimated RR was lower among current and recent
carbohydrates and rich in low-glycemic-index foods, mono- users, RR 0.69 (0.52e0.91) as compared to short-term users, RR
unsaturated and n-3 polyunsaturated fatty acids and phytoes- 0.88 (0.64e1.21). This meta-analysis did not include the 14-year
trogens in a small group of postmenopausal women resulted in follow-up data from the large Nurses Health Study (Grodstein
a fall in serum estradiol levels from 8.62 to 7.07 pg mL1 after 4.5 et al., 1998), which showed current use of postmenopausal HRT
months intervention. During this time, their BMI decreased from was associated with a RR of 0.65 (0.50e0.83). This association was
26.88 to 25.26 kg m2 (Berrino et al., 2001). attenuated to a RR of 0.84 (0.67e1.05) in past users of HRT. A
Based on the limited data available, different dietary patterns subsequent meta-analysis that included the up-dated data from the
may be responsible for a 10e20% change in serum estradiol in Nurses Health Study (Grodstein, Newcomb, & Stampfer, 1999)
postmenopausal women, but about one-half of this effect may be found that women who had ever taken postmenopausal HRT,
due to weight change. Similar changes in serum estradiol levels as compared to never users had a RR of 0.80 (0.74e0.86) for colon
a result of dietary modication in premenopausal women may also cancer and a RR of 0.81 (0.72e0.92) for rectal cancer. Much of the
apply (Goldin et al., 1981). reduction in colorectal cancer was limited to current HRT users (RR,
The effect of oral contraceptive use and hormone replacement 0.66, 0.59e0.74).
therapy (HRT) on serum and tissue estradiol concentrations is also In the Womens Health Initiative Randomized Controlled Trial
informative and illustrates one of the paradoxes associated with the (Rossouw et al., 2002), the HR for participants who received
role of estradiol in cancer development. A sub-group of post- 0.625 mg d1 CEE plus 2.5 mg d1 medroxyprogesterone acetate
menopausal women in the Womens Health Initiative randomized compared with placebo was 0.63 (0.43e0.92). On the other hand, in
controlled clinical trial of estrogen therapy that received the CEE-only arm (LaCroix et al., 2011), the HR for CEE compared
P.W. Parodi / International Dairy Journal 22 (2012) 3e14 11

with placebo was 1.15 (0.081e1.64). However, the number of colon cannot unequivocally exonerate estrogens in milk and its products
cancer cases during the trial was small. A large case-control study as possible carcinogens.
by Hoffmeister, Raum, Krtschil, Chang-Claude, and Brenner (2009)
found the risk of colorectal cancer decreased in both estrogen-only 11.2. Ovarian cancer
therapy, RR 0.42 (0.23e0.78) and in combination therapy, RR 0.60
(0.41e0.87). Genkinger et al. (2006) conducted a pooled analysis of 12 cohort
The benet of estradiol for colorectal carcinogenesis was studies representing 553,217 women among whom 2132 epithelial
conrmed in animal studies. For instance, Smirnoff, Liel, Gnainsky, ovarian cancer cases were identied. No associations were
Shany, and Schwartz (1999) demonstrated that tumor numbers in observed between intake of specic dairy foods and ovarian cancer
ovariectomized mice induced by dimethylhydrazine, were reduced risk. Subsequently, it was also found that there were no associations
by 72% when treated with estradiol. Reduced cell growth and between intakes of various dairy foods and the risk of ovarian
increased apoptotic activity, in a does-dependent manner, was cancer among participants in the Netherlands Cohort Study on Diet
noted in young adult mouse colonocytes when treated with phys- and Cancer (Mommers, Schouten, Goldbohm, & van den Brandt,
iological levels of estradiol. In addition, estradiol treatment in 2006). Higher intakes of total dairy foods were associated with
ovariectomized mice exhibited a signicant protection against a statistically signicant decreased risk of ovarian cancer in the
preneoplastic lesions (Weige, Allred, & Allred, 2009). Breast Cancer Detection Demonstration Project cohort (Koralek
et al., 2006).
11. Epidemiological evidence
11.3. Endometrial cancer
Because it is not possible to conduct appropriate clinical trials to
The role of dairy products in endometrial cancer has not been
prove unequivocally that the small quantity of estrogens in dairy
studied widely. A systematic literature review and meta-analysis of
products do not inuence cancer development, by default, epide-
the available evidence by Bandera, Kushi, Moore, Gifkins, and
miological studies, despite their limitations, may provide evidence
McCullough (2007) found no support for an association between
on the safety of dairy product consumption.
dairy product consumption and risk of endometrial cancer.

11.1. Breast cancer 11.4. Prostate cancer

Missmer, Smith-Warner, and Spiegelman (2002) conducted The World Cancer Research Fund/American Institute for Cancer
a pooled analysis of 8 cohort studies that provided 351,041 subjects, Research (WCRF/AICR) 2007 report on Food, Nutrition, Physical
7379 of whom were diagnosed with breast cancer. No signicant Activity and the Prevention of Cancer (The World Cancer Research
association was found between dairy product consumption and Fund/American Institute for Cancer Research, 2007) considered
breast cancer risk. A review of 10 cohort and 36 case-control studies that there was limited evidence suggesting that high consumption
by Moorman and Terry (2004) concluded that the epidemiological of milk and dairy products is a cause of prostate cancer. The WCRF/
evidence did not support a strong association between consump- AICR Panel suggested that high calcium intake from dairy products
tion of milk or other dairy products and the risk of breast cancer. In down-regulates the formation of 1,25-dihydroxyvitamin D3 from
later major prospective studies McCullough et al. (2005), from the Vitamin D, thereby increasing cell proliferation in the prostate.
Cancer Prevention Study II, reported that dairy product consump- Also, consumption of milk products increases blood levels of
tion was inversely associated with postmenopausal breast cancer insulin-like growth factor-1, which has been associated with
risk. The European Prospective Investigation into Cancer and increased prostate cancer risk in some studies. The Panel did not
Nutrition (Pala et al., 2009), which recruited cohorts from 10 implicate estrogens in prostate cancer risk.
European countries, did not nd dairy product consumption a risk Since the WCRF/AICR report, numerous prospective studies
factor for breast cancer. have presented results that included sub-group analyses of asso-
Exposure to initiating events during childhood, adolescence and ciations between types of milk and dairy products and stage of
early adulthood, when the mammary gland is attaining adult stage prostate cancer. There were often contradictory results, but overall
morphology, may inuence the risk of breast cancer later in life. the more recent studies do not provide strong support for an
A review of 4 case-control and 3 cohort studies (Parodi, 2005) did increased risk of prostate cancer with dairy product consumption
not nd an association between adolescent dairy product (Parodi, 2009).
consumption and subsequent breast cancer development. A later
large study of participants from the Nurses Health Study and the 11.5. Colon cancer
Nurses Health Study II found that consumption of whole milk as
part of a preschool diet was associated with a slightly decreased Numerous epidemiological studies have demonstrated a nega-
risk of adult breast cancer (Michels, Rosner, Chumlea, Colditz, & tive association between dairy product consumption, particularly
Willett, 2006). Examination of adolescence diet in the Nurses milk, and the risk of colorectal cancer. Huncharek, Muscat, and
Health Study II found no overall association for total milk or total Kupelnick (2009) conducted a meta-analysis of 26,335 cases from
dairy intake and risk of breast cancer; however, a nonsignicant 60 observational studies. The summary RR for high milk and dairy
inverse trend between breast cancer and low-fat milk and low-fat product intake, respectively, on colon cancer risk was 0.78
dairy was noted (Linos, Willett, Cho, & Frazier, 2010). Sixty-ve (0.67e0.92) and 0.84 (0.75e0.95). Milk intake was not associated
year follow-up of the Boyd Orr cohort showed that a family diet with rectal cancer risk and thus in studies where colorectal cancer
rich in dairy products during childhood was not associated with was the end point; summary RR was attenuated to 0.90 (0.81e1.00).
adult breast cancer risk (van der Pols et al., 2007). Epidemiology Calcium in milk is believed to be the principal agent responsible for
suggests that dairy product consumption during various stages of its protective action (Holt, 2008; Pufulete, 2008), although lipid
the life cycle is safe and not associated with a risk of breast cancer. components like conjugated linoleic acid, butyric acid and sphin-
However, because dairy products contain calcium together with gomyelin (Larsson, Bergkvist, & Wolk, 2005; Parodi, 1997) and milk
a range of lipid anticancer agents (Parodi, 2005), such analyses proteins, especially whey proteins (Parodi, 2007) may also
12 P.W. Parodi / International Dairy Journal 22 (2012) 3e14

contribute. Whether the small amount of bioactive estradiol in milk Berrino, F., Bellati, C., Secreto, G., Camerini, E., Pala, V., Panico, S., et al. (2001).
Reducing bioavailable sex hormones through a comprehensive change in diet:
has any impact on colon carcinogenesis is unknown, but epidemi-
the Diet and Androgens (DIANA) Randomized Trial. Cancer Epidemiology,
ology suggests it causes no harm. Biomarkers and Prevention, 10, 25e33.
Bosland, M. C. (2004). Sex steroids and prostate carcinogenesis. Annals of the New
York Academy of Sciences, 1089, 168e176.
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Hydroxyestrone: the good estrogen. Journal of Endocrinology, 150, S259eS265.
Brinkman, M. T., Baglietto, L., Krishnan, K., English, D. R., Severi, G., Morris, H. A.,
Although several lines of evidence suggest a role for estrogens in et al. (2010). Consumption of animal products, their nutrient components and
cancer at hormone-responsive sites, such as the breast, ovaries, postmenopausal circulating steroid hormone concentrations. European Journal
endometrium and the prostate, the mechanisms remain elusive. of Clinical Nutrition, 64, 176e183.
Capper, J. L., Cady, R. A., & Bauman, D. E. (2009). The environmental impact of dairy
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National Cancer Institute Monographs, 27, 75e93.
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Comparison of estrogen concentrations, estrone sulfatase and aromatase
in conjunction with a study of the levels and activity of metabo- activities in normal, and in cancerous, human breast tissues. Journal of Steroid
lizing enzymes require further investigation. Biochemistry and Molecular Biology, 72, 23e27.
Cows milk contains estrogens. Recent high sensitivity assays Chodosh, L. A., DCruz, C. M., Gardner, H. P., Ha, S. I., Marquis, S. T., Rajan, J. V., et al.
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