Mechanism of prevention and treatment of osteoporosis with traditional Chinese medicine

Dahong Ju, Yong Chai, Yuhe Wang, Zheng Yu, Xiaowei Fu, Hongyan Zhao, Hong Liu, Shaoyun Wang, Meijie Liu*

1 Beijing Key Laboratory of Research of Chinese Medicine on Prevention and Treatment for Major Diseases, Experimental Research Center, China Academy of Chinese Medical Science, Beijing 100700,China

2 Basic Research Center of Syndrome in Traditional Chinese Medicine, Institute of Basic Theory for

Chinese Medicine, China Academy of Chinese Medicine Science, Beijing 100700, China

3 The Second Affiliated Hospital, Shaanxi University of Chinese Medicine, Xianyang 712000, Shaanxi Province, China

Dahong Ju and Yong Chai are the first authors who contributed equally to this article.

Abstract

Key words: osteoporosis; hormones; cytokines; signal pathway

Introduction

Osteoporosis (OP) is a kind of systemic bone diseases characterized by bone mass reduction, bone microstructure destruction, increasing bone brittleness,decreasing bone strength and susceptibility to fracture.According to relevant researches, OP has become one of the world’s five major diseases that seriously endanger human health. The incidence rate is the third highest among chronic diseases. The number of patients with severe complications of fracture is also over 1.6 million[1].There are many risk factors for OP, the most important of which are the aging, gender differences, endocrine system disorders, immune factors, genetic factors, nutritional factors, physical factors and unhealthy living habits[2].

According to the theory of traditional Chinese medicine(TCM), the kidney governs bone and marrow, and the occurrence of OP is closely related to kidney deficiency,and also involves pathogenesis such as spleen deficiency,blood stasis and phlegm coagulation. Therefore, in TCM clinical treatment on OP, tonifying kidney and activating blood circulation play a major role. Li et al[3]and Xie et al[4]found that Bushen Zhuanggu Decoction and Gushen Decoction have ideal therapeutic effects on OP, and they could obviously relieve the clinical symptoms of primary OP patients such as pain, soreness of waist and knees, and increase the bone mineral density (BMD) of the patients.TCM prescriptions for promoting blood circulation and removing blood stasis, such as Qianggu Huoxue Decoction, Fuyuan Huoxue Decoction, Taohong Siwu Decoction, Xuefu Zhuyu Decoction and Shentong Zhuyu Decoction, also have good clinical effects in the treatment of osteoporotic vertebral compression fracture. They can greatly alleviate pain symptoms, adjust relevant serum indexes and significantly increase BMD[5-7].

In order to better understand the therapeutic mechanism of TCM on OP, many scholars have carried out a large number of studies with the help of modern technologies and methods. These studies have demonstrated that the therapeutic effect of TCM on OP was mainly achieved through the following three aspects.

Regulatory Effect on Hormones

Estrogen is the most important hormone closely related to OP. Estrogen can affect the physiological function and activity of osteoblasts and osteoclasts, through estrogen receptor and other ways, it regulates the proliferation,differentiation and apoptosis of osteoblasts and osteoclasts,and then regulates the formation and absorption of bone.Therefore, the regulation of estrogen on bone metabolism plays an important role in the occurrence and development of OP. Through clinical observation and research on postmenopausal OP patients, Lin et al found that the level of estrogen was negatively correlated with the occurrence of OP. Therefore, early detection of estrogen level was of great significance for prevention and treatment of OP in postmenopausal women[8]. Xie et al took ovariectomized rats as postmenopausal OP models, and divided the models into control group and observation group. The observation group was treated with Yigu Decoction on the basis of the control group. They found that the estrogen level in the observation group was significantly increased and bone density was improved after treatment compared with before treatment[9].

Li et al[10]believed that estrogen regulates osteoblasts and osteoclasts mainly through two pathways. One is that estrogen directly combines with estrogen receptors on osteoblasts and osteoclasts to directly affect those cells.Estrogen receptors can function as transcription factors and regulate gene transcription, thus maintaining the dynamic balance between bone formation and absorption.The other is the paracrine mechanism, estrogen can induce osteoblasts to secrete osteoprotegerin (OPG), and at the same time, it can also rely on non-genomic effect to accelerate the apoptosis of osteoclasts and slow down the apoptosis of osteoblasts. In addition, estrogen can also enhance the function of osteoblasts, inhibit the function of osteoclasts, promote bone formation and reduce bone loss by regulating relevant biochemical indexes of bone metabolism[10]. Xu et al intervened ovariectomized OP rats with estrogen nilestriol. Through the detection of BMD and bone metabolism indexes, it was found that estrogen could significantly increase the content of bone calcium and bone mineral salt in OP rats, increase the level of serum alkaline phosphatase (ALP) and OPG, increase BMD, reduce the protein expression of serum tartrateresistant acid phosphatase-5b (TRAP-5b) and RANKL,inhibit the loss of bone calcium. So the estrogen nilestriol played a therapeutic role against OP[11].

Estrogen receptors α and β dominate an important position in the process of estrogen functioning. Chen found that estrogen could maintain the bone mass of OP mice through its receptor α, slow down the bone mass loss, improve the bone microstructure, and believed that the production of this effect was closely related to the regulation of the expression of glutamate NMDA receptor 2A, which could activate the Runx2 signaling pathway, promote the proliferation and differentiation of osteoblasts and osteoblasts, thus achieving the role of bone protection[12]. Kang found that the G-protein coupled estrogen receptor GPR30/GPER1 played an important role in the physiological process of bone metabolism through the jointin vivoandin vitroexperiments. The role of the receptor would vary with the change of estrogen levels. When the estrogen was at a lower level, it would stimulate the activation of GPR30/GPER1 for “positive feedback” regulation, and improve its own surface. It could directly affect bone tissue, increase bone density, improve microstructure and biomechanical properties of bone,and finally achieve the effect of bone protection. When estrogen level in the body was basically normal, it could inhibit GPR30/GPER1 to “de-inhibit” regulation, indirectly enhance part functions of estrogen receptors α and β, and accordingly improve bone density, microstructure and biomechanical properties of bone, thus playing a role in bone protection[13]. Ma et al used the ovariectomy and modified Huith method to establish OP + osteoarthritis models, and intervened with Zhuanggujie Capsule, detected the estrogen receptor and bone density by immunohistochemistry and dual energy X-ray. The results found that Zhuanggujie Capsule could improve the expression of estrogen receptors α and β of osteoblasts, and increase bone density. So Zhuanggujie Capsule had a certain therapeutic effect for OP combined with arthritis[14].

In addition to estrogen, other hormones involved in bone metabolism would also change. Qu et al found that the combination of Yougui Pill, Liquid Calcium Soft Capsule and Calcitriol Soft Capsule could significantly improve the level of osteocalcin and estrogen, effectively improve the bone density of patients[15]. Ren et al divided postmenopausal OP women into control group and observation group. In the observation group, Lujiao Zhuanggu Capsule was used for intervention on the basis of the control group, and the levels of related reproductive endocrine hormones were detected. Results found that the levels of bone gla-protein (BGP), calcitonin,parathyroid hormone, serum follicle stimulating hormone and other hormones in the observation group were significantly increased after treatment compared with before treatment. At the same time, the BMD of the patients improved significantly, which indicated that Lujiao Zhuanggu Capsule had a good therapeutic effect against postmenopausal OP[16]. Zhang et al used ovariectomized rats as postmenopausal OP models, and used Xianlinggubao Capsule, which was a kind of Chinese medicine for invigorating kidney and strengthening bone, for treatment. Result demonstrated that after the treatment of Xianlinggubao Capsule, the level of BGP in the model rats decreased significantly, the levels of luteinizing hormone and follicle stimulating hormone also decreased, while the level of E2 increased significantly. It showed that Xianlinggubao Capsule had a good effect on the prevention and treatment of postmenopausal OP[17].

Regulatory Effect on Cytokines

Inteleukin-1 (IL-1) is mainly produced by monocyte macrophages, which is the first purified cytokine related to bone resorption. IL-1 can promote bone resorption in vivo and in vitro. IL-6 is a cytokine produced by monocyte phagocytes under the induction of IL-1 and tumor necrosis factor (TNF), and it is a multifunctional bioactive factor.It has been proved that IL-6 is related to bone resorption in vivo and in vitro. Sun et al used ovariectomized rats as OP models to study the pharmacological effect of total flavonoids of Drynaria fortunei (TFA) on OP, and found that total flavonoids of Drynaria fortunei (TFA)could significantly reduce the content of IL-6 and leptin in the serum of model rats, at the same time, it could reduce the expression of β2-adrenergic receptor in bone tissue, which was beneficial to inhibit bone absorption and played a role in treating OP[18]. Zeng et al made OP models by ovariectomy on female rats, and then made OP fracture models by tibial sawing on OP models 90 days later, intervened the model rats with Polygonatum Polysaccharide of different concentrations. It was found that high-concentration Polygonatum Polysaccharide (400 mg/kg) could significantly reduce the expression of IL-1 and IL-6, effectively slow down the apoptosis of osteoblasts and inhibit the activity of osteoclasts and correction of bone loss[19].

TNF-α is a powerful inducer of bone resorption. It mainly acts on the early stage of osteoclast formation. It has the effect of inhibiting bone formation and promoting bone resorption. Its effect depends on the existence of osteoblasts. TGF-β is a group of protein polypeptides with multiple functions, which plays an important role in bone formation and bone reconstruction. TGF-β can regulate osteoblasts and osteoclasts, it can promote the proliferation of osteoblasts and inhibit the proliferation of osteoblasts. Xie et al used ovariectomized rats as OP models, and used Huqian Pill as treatment. They found that Huqian Pill could achieve the purpose of treating OP by increasing the expression of TGF-β1 in renal tissue and increasing bone density[20]. Liu established the OP models by ovariectomy in SD female rats, and applied Jiangu 0 Formula (Jianguling Formula with addition and subtraction) as the experimental drug. They found that Jiangu 0 can reduce the expression of TNF-α and enhance the expression of TGF-β. At the same time, it could act as the estrogen, regulating the metabolism of bone and improve the symptoms of OP[21].

Insulin like growth factor-I (IGF-1) is the most abundant growth factor in bone. It can stimulate mitosis and/or initiate differentiation of all cells involved in bone transformation, especially promote the proliferation and differentiation of osteoblasts, increase the synthesis of type I collagen, the activity of ALP and the production of BGP, and inhibit the degradation of collagen. Shi et al took Kangshu Jiangu Granule as the therapeutic drug and ovariectomized rats as the OP model. After three months of administration, IGF-I in serum was detected. It was found that the Kangshu Jiangu granule could increase the content of IGF-I, inhibit the physiological activity of osteoclasts, regulate bone metabolism, and achieve the purpose of treating OP[22].

Some TCM prescriptions can regulate many kinds of cytokines. For example, Shang et al applied Bushen Prescription (composed of psoralen, Drynaria,Epimedium, Achyranthes bidentata, Cuscuta, etc.) to treat the ovariectomized OP rats. It was found that Bushen Prescription could promote the formation of bone by increasing the expression of TGF-β1, inhibit the absorption of bone by reducing the expression of IL-1 and TNF-α, regulate bone metabolism, and achieve the treatment purpose[23]. Zhang et al found that oral liquid of Boguling could significantly reduce the level of IL-6 and BGP, improve the level of serum leptin, NO and E2, effectively reduce the absorption of bone, and could be used as an effective drug for the treatment of postmenopausal OP[24].

Regulatory Effect on Signal Pathways

Many studies show that the main signaling pathways closely related to OP are: OPG/RANKL/RANK, Wnt/β-Catenin, BMP2/Smad/Runx2. Scholars have explored different signaling pathways and revealed the mechanism of TCM in prevention and treatment of OP from different perspectives.

Among all kinds of signaling pathways, OPG-RANKLRANK pathway is the most important regulatory pathway,which plays a decisive role in the generation, development,maturation and activation of osteoclasts, and is the final link of many factors regulating bone metabolism. The process of bone reconstruction is regulated by this signal pathway[25]. Wu used Zuogui Pill as a therapeutic drug to intervene the ovariectomized OP rats. After ovariectomy,because of iron ion deposition, iron overload phenomenon occurred, which acted OPG/RANKL signal pathway, and the expression of RANKL increased significantly, while the expression of OPG reduced, finally the osteoclast activity was enhanced and bone mass was significantly reduced.Under the treatment of Zuogui Pill, the bone loss had been significantly improved[26].

Many experiments show that Wnt/β-Catenin signaling pathway plays an important role in bone formation.Wnts can directly affect the process of differentiation from pluripotent precursor cells to osteoblasts. The stable expression of Wnt1 and Wnt3α can promote the proliferation of C3H10T1/2 cell line and induce the activity of ALP, which is the early marker of osteoblast differentiation. This suggests that Wnts can promote the growth and early differentiation of osteoblasts. In addition, Wnt1 and Wnt10b can inhibit the differentiation of adipocytes or preadipocytes. Overexpression of Wnt3α in C3H10T1/2 can inhibit the expression of PPARγ2, a fatty cell marker, and promote its differentiation into osteoblasts. Zhou et al also confirmed that osteoblasts can regulate the differentiation direction of mesenchymal progenitor cells through Wnt signaling pathway[27]. Liu made the medicated serum of Guben Zenggu Capsule(composed of Radix rehmanniae, Epimedium, Cistanche,Codonopsis, Astragalus and Angelica), and then used different concentrations of the medicated serum of Guben Zenggu Capsule to interfere with the cultivation of P3 generation bone marrow mesenchymal stem cells. It was found that the medicated serum of Guben Zenggu Capsule could promote the differentiation of bone marrow mesenchymal stem cells into osteoblasts and improve the gene expression of BGP and Runx2, which are closely related to the osteogenesis. They also found that the serum containing Guben Zenggu Capsule could act on Wnt/β-Catenin signal pathway, thus promoting the mRNA expression of β-Catenin and Runx2, and improving bone formation[28]. Shi et al performed an ovariectomy on female rats to prepare OP models, they used type I collagen of sika deer antler for treatment, detected the expression of related proteins in Wnt/β-Catenin signal pathway by using RT-PCR technology, and found that the expression of β-Catenin and low density lipoprotein receptor-related protein 5 (LRP-5) mRNA could be significantly increased by this collagen, which meant that Wnt/β-Catenin signal pathway was activated, so type I collagen of sika deer antler could enhance the BMD and prevent OP[29].

BMP2 can promote the osteogenesis of the non-osteoblasts,and is considered to be an accelerant of bone formation.Zhang et al used ovariectomized rats to prepare OP model,and applied Bushen Huatan Prescription (composed of Psoralen, Epimedium, Cuscuta, Trichosanthes, Monascus,hawthorn) for intervention treatment. They detected the content of BMP2, fetal globulin, matrix Gla protein, Runx2 in serum and their expression level in bone tissue, and found that Bushen Huatan Prescription could significantly improve BMP2, fetal globulin and the content of Runx2 in serum and bone tissue, as well as the content of matrix Gla protein in serum, which indicated that Bushen Huatan Prescription could act on BMP2/Smad/Runx2 signal pathway, to prevent OP and increase BMD[30].

To sum up, OP is a chronic disease with a high incidence all over the world. In order to understand OP in a comprehensive, multi-level and wide field, TCM scholars have conducted a lot of clinical and experimental researches, and have a certain understanding of the pathogenesis and prevention mechanism of OP, but it is not enough. TCM scholars should give full play to the advantages of diagnosis and treatment of TCM, constantly improve the quality and efficacy of TCM, at the same time,learn and improve the advanced experimental technology,and carry forward the advantages of TCM, in order to have a more in-depth and comprehensive understanding of the pathogenesis of OP, and to find a more effective treatment method to prevent OP.