In vitro effects of the Cimicifuga racemosa extract BNO 1055
Introduction
The controversial discussion about adverse effects and increased risks of estrogen/progestin replacement therapy (HRT) and/or estrogen replacement therapy (ERT) has focussed the interest on alternative therapies like phytoestrogens [1]. Estrogen deficiency after menopause causes numerous changes in estrogen receptive tissues like the brain, the bone, the urogenital tract and the cardiovascular system, which affects quality of life and may result in diseases like osteoporosis [2], [3].
As an alternative treatment of climacteric symptoms, owing to estrogen deficiency, CR extracts have been used since decades. Efficacy of CR, particularly concerning neurovegetative and psychic symptoms, has been proven in clinical studies [4], [5], [6] (Wuttke et al., this volume). Hot flushes, the most prominent neurovegetative symptom, are closely linked to the occurrence of LH pulses, owing to estrogen deficiency in menopausal women or after ovariectomy [7]. In both situations, estrogen deficiency causes hyperactivity of neurons, which regulate the pulsatile release of GnRH from the hypothalamus. The GnRH pulses induce the episodic release of LH from the pituitary gland. The neuronal network in the hypothalamus, which causes rhythmic GnRH release, is called GnRH pulse generator [8]. The overactivation of the GnRH pulse generator in climacteric women additionally causes co-activation of other hypothalamic neurons, which regulate body temperature and thereby vasodilatation of skin vessels. This is experienced as hot flushes [9]. Estradiol diminishes the overactivity of the GnRH pulse generator and reduces hot flushes [10].
There is also evidence that hot flushes may be successfully treated with dopaminergic drugs [11]. This correlates with the observation that the activity of the GnRH pulse generator is also modulated by dopaminergic drugs [12]. Besides an effect on hot flushes, dopaminergic drugs reduce body temperature [13]. Recently, it was demonstrated that yet unknown compounds in CR extract reduced the number of hot flush equivalents in ovariectomized rats. This effect was inhibited by the co-administration of the dopamine receptor types 2 and 3 antagonist sulpiride [14]. Therefore, it is assumed that the neurovegetative symptoms in climacteric women are ameliorated by estrogenic compounds contained in CR extract, which may act via dopaminergic mechanisms. In addition, the extracts may contain substances which per se act on dopaminergic receptors.
Prerequisite of an estrogenic action is the binding of the ligand to the estrogen receptor (ER). Currently, two subtypes of the estrogen receptor ERα and ERβ, are known, which are encoded by two distinct genes. These receptors are transcription factors with distinct functional domains involved in DNA- and ligand-binding and transcriptional regulation [15]. While the endogenous ligand 17β-estradiol does not exert preferential binding to either subtype of ER, evidence accumulates that some phytoestrogens may have a higher binding affinity to the ERβ [16].
To examine whether compounds of CR extract BNO 1055 bind to either receptor, ER ligand-binding assays (ER-LBA) were performed with ER preparations obtained from uterine tissue or with human recombinant ERα and ERβ. To investigate whether CR extract BNO 1055 exerts dopaminergic activity, dopamine ligand-binding assays (DA-LBA) with human recombinant D2-receptors were conducted.
Section snippets
Chemicals
The tracers 125I-estradiol and 125I-sulpiride were supplied by NEN (Dreieich, Germany). Recombinant human ERα and ERβ were obtained from Panvera (Madison, USA), and recombinant human D2-receptors were obtained from Biotrend (Cologne, Germany). All other chemicals were purchased from Sigma (Deisenhofen, Germany).
Cimicifuga racemosa extract BNO 1055
The CR extract BNO 1055 was used to determine the binding of its compounds to ERs, in either porcine or uterine human endometrial cytosol or to recombinant human ERα, ERβ or human
Results
In the initial step of the investigations, the binding of compounds of the CR extract BNO 1055 to a cytosolic ER preparation, prepared from porcine uteri, was examined. As shown in Fig. 1, there is a clear competition of the radioactive-labeled estradiol with compounds of the CR extract BNO 1055, which shows that this extract contains phytoestrogens. The porcine cytosol preparation was also used for monitoring of chromatographic separation of the extract (see below).
The available data, which
Discussion
Previous investigations with an ethanolic CR extract in ovariectomized rats revealed an estrogenic effect in the hypothalamo/pituitary unit. A suppression of pituitary LH secretion has been shown. No increase of uterine weight was observed [19]. This effect was attributed to unknown phytoestrogens of the CR extract. In accordance with this assumption are the results of the present in vitro investigations. The displacement of estradiol from binding sites in human endometrium cytosol preparation
Acknowledgements
This work is in part funded by an EU grant ((E)UROESTROGEN(E)ES #QLK-6-CT-2000-00565).
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