Postmenopausal status increases cardiovascular risk due to accelerated atherosclerosis progression. Cardiovascular diseases remain the leading cause of mortality and morbidity among postmenopausal women. The cardiovascular risk related to postmenopausal status is predominately due to the rapid decrease of estrogen levels, which are attributed to the indirect protective effect on lipid and glycemic control, and to the direct effect on endothelial function [1
]. The use of hormone replacement therapy (HRT) in cardiovascular prevention failed to meet expectations and it has been recognized that long-term use of hormone therapy may actually increase the risk of cardiovascular disease (CVD) in postmenopausal women, as shown in the Heart and Estrogen/progestin Replacement Study (HERS) trial, which was conducted in older postmenopausal women with established coronary heart disease (CHD) [3
]. The known side effect of HRT—that is, an increased risk of malignant hormone-dependent tumors—also produced a negative impact on the perspectives of such therapy [5
]. The general outcome of several large-scale trials was that neither estrogen nor estrogen/progestin decreased cardiovascular disease [7
]. However, later analysis has shown that HRT started in early postmenopause provides cardiovascular benefit and no harm [8
]. In spite of these findings, the expert opinion says that HRT should not be used for the primary or secondary prevention of CHD; it should be limited to the treatment of menopausal symptoms at the lowest effective dosage over the shortest duration possible, and continued use should be re-evaluated on a periodic basis [10
Phytoestrogens, mainly isoflavonoids, are believed to be an alternative to HRT in postmenopausal women. Phytoestrogens comprise a rather heterogeneous group of natural compounds of plant origin with structures similar to estrogen E2. Three of the most active compounds are coumestans, prenylflavonoids, and isoflavones. The hypothetical effects of phytoestrogens are mediated via estrogen receptors (ERα and ERβ), and the G protein-coupled estrogen receptor (GPER). It is also known that phytoestrogens have high affinity for ERβ, which explains their different action from endogenous estrogens [12
]. Similar to endogenous estrogens, phytoestrogens may provide beneficial effects on cardiovascular system through the effects on the vascular endothelium [13
], vascular smooth muscle cells [14
], intracellular cholesterol metabolism [16
], extracellular matrix synthesis [19
], and vascular inflammation [20
Dietary supplementation with phytoestrogens may inhibit the development of atherosclerotic lesions. It has been demonstrated that phytoestrogens from grapes prevent cholesterol accumulation in blood-derived cultured monocytes from postmenopausal women [17
]. Animal studies support the anti-atherogenic properties of phytoestrogens; for example, genistein inhibited atherogenesis in hypercholesterolemic rabbits mostly via its beneficial effects on endothelial dysfunction [21
]. Resveratrol (stilbene with known estrogen-like activity) exhibited multiple anti-atherogenic effects [22
], including inhibition of intimal hyperplasia [23
] and inhibition of low density lipoprotein (LDL) oxidation [24
]. The results of experimental studies demonstrate that phytoestrogens have a potential in anti-atherosclerotic therapy, because they are able to modulate several mechanisms of atherosclerosis progression.
Previously, in an ex vivo model, we evaluated the anti-atherosclerotic effect of phytoestrogen-rich plants and their combinations [17
]. Based on the results of dose titration studies, qualitative compositions of isoflavonoid-rich anti-atherosclerotic herbal preparation—active ingredients: tannins from grape seeds, green tea leaves, hop cone powder, and garlic powder—was developed. The aim of the present study was to investigate the effect of this isoflavonoid-rich herbal preparation on the progression of subclinical carotid atherosclerosis in healthy postmenopausal women.
The intima-media thickness of common carotid arteries (cIMT) measured by B-mode ultrasound is a widely accepted marker of subclinical atherosclerosis; it is well correlated with the degree of coronary atherosclerosis and is a significant predictor of clinical manifestations of atherosclerosis. This instrumental marker is used in clinical and epidemiological studies to assess the impact of conventional and novel cardiovascular risk factors and treatment regimens on atherosclerosis progression [25
]. Several clinical trials were aimed at the assessment of the effects of HRT or phytoestrogens on cIMT progression in postmenopausal women [28
]. Thus, in this study we have used ultrasound examination of common carotid arteries and cIMT measurement as a tool for quantitative assessment of atherosclerosis, with annual cIMT progression as the endpoint.
The results of this study have demonstrated that mean cIMT progression was slower in asymptomatic postmenopausal women who received isoflavonoid-rich herbal preparation, as compared to women who received the placebo. In addition, the herbal preparation decreased the total cholesterol, LDL-C levels, and suppressed cIMT progression after 12 months of herbal preparation administration. It should be noted that in our study, the baseline LDL-C and total cholesterol level, as well as the prevalence of diabetes in isoflavonoid-rich herbal preparation recipients were higher than in the placebo group; these risk factors of atherosclerosis could suggest more pronounced cIMT and plaque progression. However, we have seen the reverse effect; therefore, it may be expected that the anti-atherosclerotic potency of isoflavonoid-rich herbal preparation may even be underestimated in this study. On the other hand, the reduction in total cholesterol and LDL-C after 12-month follow-up in the isoflavonoid group could be due to regression towards the mean, since they were higher at the baseline.
CVD related to atherosclerotic process is responsible for the majority of deaths in postmenopausal women. The prevention of the lipid accumulation in cells is the key mechanistic factor for inhibition of atherosclerotic plaque formation at the early stages of atherosclerosis progression. Phytoestrogens have the capacity to affect plasma lipid profile, but little is known regarding their effects on atherosclerosis progression. In women undergoing coronary angiography for suspected myocardial ischemia, beneficial association between blood levels of the phytoestrogen daidzein and lipoproteins, particularly lower triglycerides and higher HDL-C levels were previously reported [32
]. The main association of phytoestrogens with lipoprotein levels was incrementally related to diadzein, but not with other lipoprotein modulators. In another clinical study it has been shown that isoflavones induce the reduction of total cholesterol and LDL-C plasma levels without affecting triglycerides or HDL-C levels [33
]. Our results are in line with previous findings; however, in our study blood lipid levels were decreased in both groups: in the placebo group, these changes were significant for total cholesterol and HDL-C; and in the isoflavonoid-rich herbal preparation recipients, for total cholesterol and LDL-C. The ability of phytoestrogens to reduce the accumulation of cholesterol in cells is a possible mechanism to explain the effects on mean cIMT. Previously, we have evaluated the anti-atherogenic effect of phytoestrogen-rich plants using an ex vivo model based on primary cultures of monocytes isolated from the blood of healthy donors. In this model, the ability of human serum to induce accumulation of cholesterol in cultured cells (serum atherogenicity) was measured, as well as the effect of single dose oral administration of plant extract on serum atherogenicity [17
]. Grape seeds extract (100 mg) lowered serum atherogenicity by 71%, 78%, and 81% at 2, 4, and 6 h after oral intake of a single dose. Similar effects were observed for hop cones (250 mg), garlic powder (150 mg), sage leaves (100 mg), green tea leaves (250 mg), sea kelp (500 mg), fucus (250 mg), and carrot (1000 mg). The ability of soya beans extract (35 mg) to lower serum atherogenicity by 28%, 38%, and 30% at 2, 4, and 6 h after a single dose administration, respectively, was demonstrated [17
The main endpoint of the current study was to identify the annual changes in cIMT progression. Several studies demonstrated that cIMT is a significant and independent predictor of cardiovascular events, and allows for non-invasive evaluation of early atherosclerosis progression in asymptomatic patients [36
]. Only a few clinical trials investigated the effect of phytoestrogens on atherosclerosis progression in postmenopausal women. In the recent long-term intervention trial (2.7 years) with soy isoflavones, the inhibition of subclinical atherosclerosis progression evaluated by cIMT was demonstrated. Healthy postmenopausal women were randomized in two groups; the first group who received daily supplement with 25 g soy protein containing 91 mg of isoflavones, and the second group who received a placebo. In both groups, the increment of cIMT was observed. However, in the soy group the cIMT progression was not statistically significant (p
= 0.35), and was 16% lower than in the placebo group. That study has enrolled 350 participants, and the duration was more than two years. The authors suggested that further use of isoflavone-rich dietary supplements would allow achieving the significant difference in the rate of atherosclerosis progression between the two groups [28
In our study, mean cIMT changes in both groups were observed. However, in herbal preparation recipients this increase was negligible, but in the placebo group the increment was significantly higher than in herbal preparation recipients, and accounted for 111 μm, or 13% increase. This fact indicates that in postmenopausal women the rate of carotid atherosclerosis progression is notably high. In the study by Rossi et al. [37
], the mean cIMT progression accounted for 103 μm (range from −250 to 567 μm; IQR from 0 to 200 μm) per year in hypertensive postmenopausal women, and this progression rate is very close to our data. It should be noted that in our study the difference between the two groups in systolic and diastolic blood pressure at the baseline was statistically significant (135/83 versus 127/79 in placebo group and herbal preparation group, respectively), and this fact may partly explain the rather high progression of cIMT in the placebo group, which has not been replicated in any other studies [29
]. On the other hand, Colacurci et al. have demonstrated rather similar cIMT progression rates in non-hypertensive women [31
]. The limitations of our study, such as the duration of the follow-up and rather limited sample size, did not allow defining the proportion of cIMT progression rates in placebo recipients explained by the higher blood pressure.
It should be noted that the time since menopause is of the essence when studying atherosclerosis progression and medical intervention. There exists an opinion that timely HRT may offer protection against CVD, whereas in older women there may be cardiovascular harm associated with HRT use [8
]. Hodis et al. have recently demonstrated that anti-atherosclerotic effects of HRT on cIMT progression differed between early and late postmenopause. Oral estradiol therapy was associated with less progression of subclinical atherosclerosis measured as cIMT dynamics than was placebo when therapy was initiated within six years after menopause, but not when it was initiated ten or more years after menopause [29
]. Thus, estradiol was shown to be effective in reducing cIMT progression. On the other hand, in the Kronos Early Estrogen Prevention Study (KEEPS) performed in more than 700 healthy women aged 42 to 59 within three years after menopause, the carotid ultrasound studies showed similar rates of progression of cIMT in all three treatment groups (0.45 mg a day of Premarin—an oral conjugated equine estrogen (o-CEE)—or 50 µg a day of transdermal estradiol via a Climara patch, or placebo) over the four years of study. However, these changes were reported to be generally small; therefore, slow cIMT progression limited the statistical power to detect any differences among the groups [30
]. According to the results of our study, isoflavonoid-rich herbal preparations may provide a direct anti-atherosclerotic effects, but no direct comparisons with estrogens were performed. In general, it may be proposed that anti-atherosclerotic action of drugs should be realized via prevention of intracellular cholesterol accumulation in vascular wall cells, but it is unclear if estrogens may possess the same mechanistic effect at the cellular level. In our previous studies on prevention of intracellular cholesterol accumulation, modified LDL were used to induce intracellular lipid deposition, and the effects of drugs or chemical compounds mainly related to LDL binding, uptake, internalization, and metabolism in cells were in focus. In contrast, Wang et al. explored the alternative way of preventing foam cell formation via cholesterol efflux modulation. They have demonstrated that 17β-estradiol promotes cholesterol efflux from vascular smooth muscle cells and reduces foam cell formation via ERβ- and liver X receptor (LXR) α-dependent upregulation of ABCA1 and ABCG1 [38
]. Another mechanism of atherosclerosis prevention may be related to anti-inflammatory effects, and it was shown that estradiol can regulate monocyte chemotactic protein-1 (MCP-1) in human coronary artery smooth muscle cells [39
], increase prostacyclin synthesis in cells from atherosclerotic lesions [40
], impair endothelial function in postmenopausal women [41
], transform growth factor activity [42
], and attenuate atherogenesis via selective estrogen receptor beta modulator 8β-VE2 [43
]. On the other hand, anti-inflammatory effects of phytoestrogens are also known [44
]. Therefore, anti-atherogenic effects of both estrogens and isoflavonoids are not limited to the inhibition of direct accumulation of cholesterol in cells only.
Finally, the findings of our study are in line with the results obtained from the study aimed to evaluate the effect of selective estrogen receptor modulator Raloxifene on atherosclerosis progression in postmenopausal clinically healthy women. In a prospective study enrolling 155 postmenopausal women without clinical manifestations of CVD, study participants were randomized in two groups, receiving Raloxifene 60 mg daily or placebo for 18 months. The cIMT progression for 18 months was 11.2 μm in Raloxifene group versus 85.7 μm in the placebo group (p
< 0.004). Thus, the lower risk of cIMT progression was demonstrated in Raloxifene recipients (odds ratio = 0.41; 0.32–0.70 at a 95% confidence interval) [31
Nevertheless, our study has certain limitations. The main one is the duration of the follow-up, only 12 months. Long-term effects of isoflavonoid-rich dietary supplement Karinat need to be further studied in order to evaluate its effects on main cardiovascular risk factors and long-term outcomes, such as myocardial infarction and stroke [48
]. Indeed, longer observation may help to better understand the effects of isoflavonoid-rich herbal preparations on main outcomes of CVD, but in this study that was not the primary endpoint. The second notable limitation is a rather small sample size. To interpret the results from our study, the limited number of enrolled subjects needs to be taken into consideration, as it may lead to confounding results, despite randomization. Finally, it should be noted that the effect of isoflavonoids or other estrogen-like molecules on cardiovascular health may be realized more through endothelial function/dysfunction. In our study we have evaluated only the effects on lipids, and the effect of treatment on the arterial wall that reflects atherosclerotic profile. It should be expedient to study the effects of isoflavonoid-rich herbal medications also on endothelial function using, for example, flow-mediated dilatation.