Research On Effect And Mechanism Of Danshen Extract Or Chuanxiong Extract On Serum Lipid And Bone In Ovariectomized Rats Fed With High Fat Diet

OBJECTIVE AND SIGNIFICANCECardiovascular diseases (CVD) and osteoporosis (OP) are common chronic diseases andaccount for significant morbidity and mortality in the aging population. Accumulatingevidence indicates a pathophysiological link between the two diseases. Due to deficiency offemale hormones (progesterone and estrogen), they frequently coexist in postmenopausalwomen. Postmenopausal women commonly take hormone replacement therapy with estrogenand/or progesterone. Such therapy is really effective in preventing CVD and OP inpostmenopausal women, but it was proven to have various adverse effects. Danshen andchuanxiong are well-known Traditional Chinese Medicinal herbs widely used for thetreatment of CVD, and their major active ingredients have been found to have estrogenic andprogestogenic effects respectively. The aim of this study was to evaluate the effect ofDanshen and Chuanxiong on cardiovascular system and bone system in an ovariectomized(OVX) rat model fed with high fat diet (HFD) for the first time. The study may not only helppostmenopausal women to relieve suffering and save financial burden, but also pave the wayto elucidate the underlying relationship between CVD and OP.METHODS AND RESULTS(1) Danshen herb and Chuanxiong herb were respectively ground into powder andextracted with distilled water at100°C or80%ethanol, and the yield was13.03%and17.08%. The results of HPLC analysis were shown that the Danshen aqueous extract contains3.93±0.10%salvianolic acid B and0.20±0.03%danshensu and Chuanxiong ethanolicextract contains4.76±0.25%ferulic acid and3.00±0.16%tetramethylpyrazine.(2) Forty-nine6-month-old female Sprague-Dawley rats were randomly divided intoseven groups: sham-operated rats with low-fat control diet (LFD)+vehicle, sham-operatedrats with high-fat diet (HFD)+vehicle, OVX rats with LFD+vehicle, OVX rats with HFD+vehicle, OVX rats with HFD+17β-estradiol (1mg/kg/d, PO), and OVX rats with HFD+Danshen aqueous extract (600mg/kg/d, PO), and OVX rats with HFD+Chuanxiongethanolic extract (600mg/kg/d, PO). Rats were orally administered with vehicle, E2or herbextracts and fed with HFD or LFD respectively for12weeks. Results showed that OVX ratsfed with HFD for12weeks had highest body weight gain and lowest liver index, kidneyindex and spleen index. HFD-fed OVX rats treated with E2, Danshen aqueous extract or Chuanxiong ethanolic extract for12weeks were shown to decrease body weight and increaseliver index, kidney index and spleen index.(3) Lipid profile levels in serum, histological examination of livers, lipid content of liver,oxidative/antioxidative activity in liver, vascular activity were detected to evaluate theprotection effect of Danshen aqueous extract or Chuanxiong ethanolic extract oncardiovascular system in HFD-fed OVX rats. The results were shown that abnormal lipidprofiles, fatty liver and impaired vascular endothelium were induced in HFD-fed OVX rats.Comsumption of Danshen aqueous extract or Chuanxiong ethanolic extract improved serumlipid profiles (lower triacylglycerols, total cholesterol and low density lipoprotein cholesterol),reduced ALT and AST activity, prevented formation of fatty liver induce by HFD and OVX,primarily via up-regulating the mRNA and protein expression of LDLR and CYP7A1anddown-regulating the mRNA and protein expression of HMGCR. In addition, they couldincrease endothelial-dependent vasorelaxation and displayed vasoprotection in OVX rats fedwith HFD, primarily by decreasing hepatic MDA levels, increasing hepatic antioxidantenzymes (SOD, CAT, GSH-Px) activities, stimulating nitric oxide (NO) production,up-regulating the mRNA expression of endothelial NO synthase, and down-regulating themRNA expression of tumor necrosis factor alpha, intercellular cell adhesion molecule-1, andvascular cell adhesion molecule-1in the isolated aortas as well as E2.(4) Ca and P levels in serum and urine and bone properties in cancellous bone (proximaltibia and lumbar vertebra2) and cortical bone (midshaft tibia and midshaft femur) bymicro-CT were detected to evaluate the protection effect of Danshen aqueous extract orChuanxiong ethanolic extract on bone system in HFD-fed OVX rats. The results were shownthat HFD and OVX could induce rats to aggravate urinary Ca excretion, decrease bonemineral density (BMD) and change the microstructure of cancellous bone. Treatment withDanshen aqueous extract, Chuanxiong ethanolic extract or E2in HFD-fed OVX rats couldsuppress urinary Ca excretion, increase BMD, trabecular number and bone surface density,decrease trabecular separation, specific bone surface and structure model index and changetrabeculae to parallel plates instead of cylindrical rods, but cancellous bones they mainlyaffected were different. Danshen aqueous extract mainly affected proximal tibia, andChuanxiong ethanolic extract mainly affected lumbar vertebra2, and E2could affect both ofthem.(5) MTS and alkaline phosphatase (ALP) were measured to evaluate the in vitro effect ofDanshen aqueous extract or Chuanxiong ethanolic extract on proliferation and differentiation of rat osteoblast-like UMR106cell. The results were shown that Danshen aqueous extractcould only stimulate differentiation in UMR106cells, but not affect cell proliferation. AndChuanxiong ethanolic extract exhibited to promote differentiation potential as well asproliferative activity in UMR106cells.CONCLUSSION(1) It was exhibited that heavy body weight, abnormal lipid profiles, impaired liver andvascular function, aggravated urinary Ca excretion, decreased BMD and changed themicrostructure of cancellous bone in HFD-fed OVX rats, indicated a successful rat modelmimicking CVD and OP in postmenopausal women.(2) Treatment with Danshen aqueous extract or Chuanxiong ethanolic extract in HFD-fedOVX rats not only prevented CVD, but also displayed anti-osteoporosis for the first time.(3) Effects of Danshen aqueous extract on cancellous bones and on UMR106cell were notexactly same with Chuanxiong ethanolic extract, but they both exhibited to promote celldifferentiation and prevent bone loss in vitro and in vivo, might be potential candidates forprevention and treatment of postmenopausal osteoporosis.