The compatibility of individual herbs in Erxian Decoction, an anti-menopausal herbal formula, for treating osteoporosis

Abstract

BACKGROUND: Premenopausal osteoporosis is one of the syndromes in postmenopausal women, which is associated with the estrogen deficient state (1, 2). Estrogen deficiency would lead to increased receptor activator of nuclear factor kappa-B ligand (RANKL) which binds to RANKL receptor of osteoclast precursor triggering osteoclastogenesis (3-5). Estrogen deficieny will also suppress the expression of osteoprotegerin (OPG), which can bind and inhibit the action of RANKL (6). Risk of bone fracture increase when the balance between bone formation and bone resorption is tipped (7). Erxian Decoction (EXD) has been used for relieving menopausal symptoms for more than fifty years. It is composed of six herbs namely Rhizome curculiginis, Herba epimedii, Radix Morindae officinalis, Rhizome anemarrhenae, Cortex phellodendri, and Radix Angelicae sinenisis. It is reported that EXD can stimulate ovarian estrogen production (8) and also possesses anti-osteoporotic properties (9). However, the compatibility and the role of different herbs in EXD formula in treating osteoporosis have not been reported. The effects of EXD and its component herbs on proliferation of human fetal osteoblast cell (hFOB 1.19), murine macrophage-like osteoclast precusor (RAW 264.7), OPG secretion, and the molecular signaling involved in osteoclastogenesis have been investigated in this study. RESULTS AND DISCUSSION: Our results showed that EXD could significantly stimulate proliferation of hFOB 1.19 while inhibit proliferation of RAW264.7. The inhibitory effect on RAW 264.7 was most prominent in cells treated with Herba epimedii alone, and this inhibitory effect was not observed in cell treated with EXD without Herba epimedii. This indicates that the anti-osteoporotic efficacy is contributed largely by Herba epimedii in term of regulating the proliferation of osteoclast precursor. The stimulatory effect of hFOB 1.19 was most prominent in cell treated with Radix Morindae officinalis, and such stimulatory effect was also eliminated in EXD without Radix Morindae officinalis. Thus the stimulation of osteoblast proliferation by EXD was due to the presence of Radix Morindae officinalis. The measure of osteoblast secreted OPG has also revealed that EXD can significantly increase OPG production from hFOB 1.19 cells. The increased in OPG was only observed in EXD, suggesting that different individual herbs exerted mutual effect in stimulating OPG production by EXD, thus inhibiting the osteoclastogenic effect of RANKL. In the differentiation assay, in which EXD could significantly suppress the differentiation of RAW 264.7 cells into osteoclasts, as signified by the lower number of multi-nucleated cells detected by mature osteoclast marker tartrate-resistant acid phosphatase (TRAP) staining method. The inhibitory effect of osteoclast differentiation was mainly mediated by Herba epimedii, as the inhibitory effect was less prominent in treatment of EXD without Herba epimedii. CONCLUSION: Our results have demonstrated the potent anti-osteoporotic effect of EXD in vitro. The anti-osteoporotic effects of EXD could due to the anti-proliferation of osteoclast precursor cells and stimulatory effect of osteoblast cells. These effects are contributed by different herbs in the EXD formula, in which the effect on osteoclast was mainly contributed by Herba epimedii and the effect on osteoblast was mainly due to Radix Morindae officinalis. The six component herbs could also exert stimulatory effect on osteoblast OPG production in the EXD formula but not in single herbs alone. EXD could also inhibit differentiation of RAW 264.7 into mature osteoclast which was also mainly mediated by Herba epimedii. Our results have demonstrated the anti-osteoporotic effect of EXD and its component herbs, and suggested the possible interaction and compatibility of component herbs in an anti-menopausal Chinese medicine formulation.