Chinese Journal of Natural Medicines  2018, Vol. 16Issue (11): 838-845  DOI: 10.3724/SP.J.1009.2018.00838
0

Cite this article as: 

CHEN Juan, YUE Jia, LIU Jiao, LIU Yun, JIAO Kai-Lin, TENG Meng-Ying, HU Chun-Yan, ZHEN Jing, WU Mao-Xuan, ZHOU Ming, LI Zhong, LI Yuan. Salvianolic acids improve liver lipid metabolism in ovariectomized rats via blocking STAT-3/SREBP1 signaling[J]. Chinese Journal of Natural Medicines, 2018, 16(11): 838-845.
[Copy]

Research funding

This work was supported by National Natural Science Foundation of China (Nos. 81573183 and 81673205), the Major Program of Natural Science Research of Jiangsu Higher Education Institutions (No. 15KJA330001), the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD), Gansu Provincial Natural Science Foundation (No. 148RJZA081), and Lanzhou Science and Technology (2014-1-18)

Corresponding author

E-mail:lz-ny@njmu.edu.cn (LI Zhong)
E-mail:liyuan@njmu.edu.cn (LI Yuan)

Article history

Received on: 30-May-2018
Available online: 20 November, 2018
Salvianolic acids improve liver lipid metabolism in ovariectomized rats via blocking STAT-3/SREBP1 signaling
CHEN Juan1 , YUE Jia2 , LIU Jiao1 , LIU Yun1 , JIAO Kai-Lin1 , TENG Meng-Ying1 , HU Chun-Yan1 , ZHEN Jing1 , WU Mao-Xuan1 , ZHOU Ming1 , LI Zhong1 , LI Yuan1     
1 Department of Nutrition and Food Hygiene, School of Public Health, Nanjing Medical University, Nanjing 211166, China;
2 Department of Nutrition and Food Hygiene, School of Public Health, Gansu University of Chinese Medicine, Lanzhou 730000, China
[Abstract]: Postmenopausal women, who have reduced circulating estrogen levels, are more prone to develop obesity and related metabolic diseases than premenopausal women. The absence of safe and effective treatments for postmenopausal obesity has changed the focus to natural products as alternative remedies. Total salvianolic acids (TSA) are the major water-soluble ingredients of Danshen. Salvianolic acid (SA) is the major constituent of the TSA. Salvianolic acids, including TSA and SA, are widely used in traditional Chinese medicine. In the present study, ovariectomized rats and LO2 cells were used to study the effects of salvianolic acids on body weight gain and hepatic steatosis. Salvianolic acids reduced ovariectomy (OVX)-induced body weight gain, attenuated the expressions of hepatic lipogenic genes, such as sterol regulatory element binding protein (SREBP)1, fatty acid synthase (FAS), and stearoyl-CoA desaturase (SCD)1, and decreased the liver triglyceride (TG) and total cholesterol (TC). For the molecular mechanisms, OVX and high glucose-induced phosphorylation of signal transducer and activator of transcription (STAT)-3 was inhibited by salvianolic acids treatment. In LO2 cells, inhibition of STAT-3 by siRNA attenuated the increased expression of SREBP1 and TG induced by high glucose. Salvianolic acids reduced the upregulation of SREBP1 and TG induced by high glucose in LO2 cells. In conclusion, these findings illustrated that salvianolic acids markedly alleviated the lipid metabolism disorders and protected against the postmenopausal obesity. The underlying mechanism was probably associated with the regulation of STAT-3 signaling.
[Key words]: Salvianolic acids     Lipid metabolism     Weight gain     Signal transducer and activator of transcription 3    
Introduction

Salvia miltiorrhiza Bunge, also known as "Danshen", is a shade-growing perennial flowering plant belonging to the family Labiatae. It is an important and widely used medicinal plant in traditional Chinese medicine, which can exert protective effects on the liver, kidneys, and lungs, and used for prevention and treatment of vascular diseases [1-4]. Salvianolic acid B is the major constituent of the salvianolic acids and the most active constituent among the phenolic acids, possessing a wide range of pharmacological effects, such as antioxidant, anti-ischemia-reperfusion and antitumor activities [5-8]. However, knowledge about its role in postmenopausal obesity is still limited.

The menopause is an age-related loss of ovarian functions and a decline in circulating estrogen levels [9]. Compared with premenopausal women, postmenopausal women are more prone to develop obesity and related metabolic diseases such as diabetes, cardiovascular disease, and non-alcoholic fatty liver disease [10-12]. Hormone replacement therapy is the most common and effective treatment for menopausal syndrome. However, the potential risks of breast cancer and endometrial cancer have resulted in much interest in investigating natural products to manage the increase in disease risk caused by the loss of ovarian hormones [13-14].

Signal transducer and activator of transcription (STAT)3 is a transcription factor closely associated with receptor kinases that mediate cellular signaling in response to ligand binding at the receptor. Studies reveal that STAT-3 has the ability to modulate many genes involved with cell growth, survival, and death [15]. Indeed, the STAT-3 signaling pathway is responsible for the regulation of gluconeogenesis and lipid metabolism in the liver in high-fat diet [16-17]. However, the potential molecular mechanisms of STAT-3 involvement in liver lipid metabolism need further study in an ovariectomy (OVX)-induced weight gain model.

Total salvianolic acids (TSA) are the water-soluble ingredients of Danshen. The composition has been revealed as salvianolic acid B, salvianolic acid A, caffeic acid, and other salvianolic acids [18]. TSA are widely used in orally administered herbal medicines and injection [19]. The aims of the present study were to evaluate the difference in effect on OVX-induced body weight gain between TSA and salvianolic acid (SA) and to explore the role of STAT-3 in liver lipid metabolism in OVX-induced weight gain model.

Materials and Methods Reagents

Carboxymethyl cellulose sodium solution and estrogen (E2) were purchased from Sigma-Aldrich (St Louis, MO, USA). TSA and salvianolic acid (SA) were respectively purchased from Jiangsu Langze Medical Technology Co., Ltd. (Nanjing, China) and Shanghai Green Valley Pharmaceutical Co., Ltd. (Shanghai, China). TSA were water-soluble extract of Danshen, mainly contained two components, salvianolic acids B and A, in a ratio of 4 : 1. SA was the drug of salvianolate injection, mainly contains 80% pure salvianolic acid B.

Animals and treatments

This study was approved by Nanjing Medical University Institutional Animal Care and Use Committee (Permit Number: NJMU-IACUC-1403024), and the animals were treated humanely and with regard for alleviation of suffering. Fifty female Sprague-Dawley rats [20] weighing 220 ± 10 g were purchased from Beijing Vitalriver Experimental Animal Co., Ltd. [Certificate No: SCXK (Jing) 2012-0001]. All the animals were housed under controlled conditions (23 ± 2 ℃, 55%-65% humidity, and 12 h/12 h light/dark cycle) with free access to food and tap water. The animals were acclimated for 1 week before experiments and then randomly divided into five groups of ten. All the other groups were subjected to bilateral OVX, except for the sham-operated (SHAM) group, which was subjected to the same surgical procedure, but the ovaries were preserved. One week after the operation, the rats were treated with drugs by oral gavage once daily for 50 days. The groups were SHAM, OVX, OVX + E2, OVX + TSA, and OVX + SA. The drug concentrations were 1 mg·kg-1 of E2 and 50 mg·kg-1 of TSA and SA, respectively. The drugs were dissolved in 0.5% carboxymethyl cellulose sodium solution. The solution volume of drugs given to rats was 10 mL·kg-1.

Cell culture

Human liver cell line LO2 was purchased from the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China. The LO2 cells were cultured in Dulbecco's modified Eagle's medium (5.56 or 25 mmol·L-1 glucose) (Life Technologies/Gibco, Grand Island, NY, USA). The medium contained 10% fetal bovine serum (Life Technologies/Gibco), 100 mg·mL-1 of streptomycin (Gibco) and 100 U·mL-1 of penicillin (Gibco). The LO2 cells were maintained in the presence of 5% CO2 at 37 ℃.

Quantitative real-time polymerase chain reaction (qRT-PCR)

RNA from the cells and liver were isolated with TRIzol reagent (Invitrogen, Carlsbad, CA, USA). For the detection of mRNA, total RNA (1 μg) was transcribed into cDNA using AMV Reverse Transcriptase (Promega, Madison, WI, USA). qRT-PCR was performed using the LightCycler Sequence Detection System (Roche Applied Science, Switzerland) with cycling conditions as follows: 95 ℃ for 300 s, 95 ℃ 10 s and 60 ℃ for 30 s for 40 cycles. Fold changes in expression of each gene were calculated by a comparative threshold cycle (Ct) method with the formula 2-ΔΔCt.

Western blotting analysis

The proteins were quantified using the BCA Kit (Beyotime Biotechnology, Haimen, China). 10% SDS-PAGE was used to separate the proteins, which were transferred onto nitrocellulose membranes (Bio-Rad Laboratories, Hercules, CA, USA). After blocking with 5% non-fat milk, the membranes were incubated with primary antibodies at 4◦C overnight. Antibodies used in the present study were against AKT, p-AKT, STAT-3, and p-STAT-3 (Y705) (Cell Signaling Technology, Boston, MA, USA). The antibodies were diluted 1 : 1000. β-Actin (Beyotime) served as the internal control to eliminate the differences in protein loading. Followed by incubating them with horseradish peroxidase conjugated secondary antibodies (anti-rabbit IgG and anti-mouse IgG) (Beyotime Co. Ltd, the dilution was 1 : 1000) for 1 h at the room temperature. The antigen complexes were detected with enhanced chemiluminescence. For densitometric analyse, the bands on the blots were measured by the Image-Pro-Plus 6.0 software.

Cell transfection

siRNA-con and siRNA-STAT-3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA; http://datasheets.scbt.com/sc-37007.pdf and http://datasheets.scbt.com/sc-270027.pdf). The LO2 cells were transiently transfected with the siRNAs in the presence of Lipofectamine 2000 reagent (Invitrogen) for 12 h.

Measurement of hepatic lipids and total cholesterol

The dissected liver tissues (100 mg) were homogenized in 0.9 mL of 0.86% saline on ice. Residual tissue debris was pelleted by centrifugation at 2500 r·min-1 for 10 min. The supernatant was used to quantify liver triglyceride (TG) and total cholesterol (TC) using the TG and TC Assay Kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China). The LO2 cells were digested by trypsin-EDTA Solution (Beyotime). The prepared LO2 cell suspension was centrifuged at 1000 r·min-1 for 10 min. After washing twice, the cell precipitate was added to 200-300 μL of lysis solution for 30-40 min on ice, then the lysis solution were used for the assay.

Statistical analysis

The data were compared using GraphPad version 6.0 (La Jolla, CA, USA) and were presented as means ± SD. P < 0.05 was considered statistically significant.

Results Effects of salvianolic acids on body weight, body weight gain, and food intake

Bilateral OVX in rats is a well-recognized and widely used animal model of human menopause [21]. We exposed OVX rats to E2, TSA and SA; the initial body weights were not significantly different among all the groups. After 50 days, the OVX group exhibited significantly more weight gain than SHAM group did. In contrast, the weight gain of OVX + E2 group was markedly lower than that of OVX group. Similarly, OVX + TSA and OVX + SA groups also showed less weight gain than OVX group did (Figs. 1A and 1B). There was no difference in OVX-induced body weight gain between TSA and SA groups. The food intake in OVX group was significantly higher than that in SHAM group. Compared with the OVX group, the OVX + E2 group had significantly decreased food intake; however, no apparent difference was found for food intake among OVX, OVX + TSA, and OVX + SA groups (Fig. 1C). These results indicated that salvianolic acids decreased the body weight induced by OVX, not solely dependent on food intake. So, we further determined if the hepatic metabolism was involved in subsequent experiments.

Figure 1 Effects of salvianolic acids on body weight (A), body weight gain (B), and food intake (C). The data are expressed as means ± SEM, n = 7-10 rats/group. SHAM, sham operated; Vehicle, ovariectomized; E2, OVX + estradiol; TSA, OVX + total salvianolic acids; SA, OVX + salvianolic acid. **P < 0.01 vs SHAM, #P < 0.05, ##P < 0.01 vs OVX
Effects of Salvianolic acids on lipid levels of OVX rats

The liver is the first tissue in the body to come in contact with metabolites from ingested diet and an important component in energy balance [15]. The levels of hepatic TG and TC in the OVX group were higher than that in OVX + E2, OVX + TSA, and OVX + SA groups (Figs. 2A and 2B). Lipogenic genes such as hepatic sterol regulatory element binding protein (SREBP)1, fatty acid synthase (FAS), Acetyl-CoA Carboxylase (ACC)1, and stearoyl-CoA desaturase (SCD)1 are related to fat deposition and energy metabolism in the liver after OVX [22-24]. We further determined if OVX had effects on the expressions of lipogenic genes. As shown in Fig. 2C, OVX increased the expression of hepatic SREBP1, but there were marked decreases in hepatic SREBP1 in OVX + E2, OVX + TSA, and OVX + SA groups. Similar results were obtained for the expressions of hepatic FAS and SCD1 (Figs. 2D and 2E). These results indicated that OVX increased the expression of lipogenic genes, which could be inhibited by salvianolic acids, decreasing OVX-induced fat accumulation in liver.

Figure 2 Effects of salvianolic acids on lipid levels of OVX rats. (A) Hepatic TG contents. (B) Hepatic TC contents. (C-E) mRNA of SREBP1, FAS and SCD1 were investigated by qRT-PCR. SHAM, sham operated; Vehicle, ovariectomized; E2, OVX + estradiol; TSA, OVX + total salvianolic acids; SA, OVX + salvianolic acid. *P < 0.05, **P < 0.01 vs SHAM, #P < 0.05, ##P < 0.01 vs OVX. The liver tissue of five rats from each group was mixed as a sample, three independent samples each group
Relationship between STAT-3 and lipogenesis in OVX rats

STAT-3 plays an important role in adipogenesis [25]. Previous research has shown that STAT-3 is involved in lipid synthesis by regulating the expression of SREBP1 in high-fat diet [16]. In the OVX model, it was unclear whether STAT-3 was involved in OVX-induced obesity. We found that salvianolic acids inhibited the OVX-induced phosphorylation of STAT-3 in rats. Protein kinase B (PKB/AKT) signaling is involved in lipid metabolism [26]. In the present study, treatment of the OVX rats with salvianolic acids upregulated the phosphorylation of AKT (Figs. 3A and 3B), indicating a classical activation of AKT signaling. Collectively, the in vivo results revealed that the effects of salvianolic acids on lipid metabolism might be associated with STAT-3/ SREBP1 signaling.

Figure 3 Effects of salvianolic acids on STAT-3 in liver of OVX rats. (A, B) Western blotting and densitometry analysis of expression of p-STAT-3, STAT-3, p-AKT, and AKT. SHAM, sham operated; Vehicle, ovariectomized; E2, OVX + estradiol; TSA, OVX + total salvianolic acids; SA, OVX + salvianolic acid. **P < 0.01 vs SHAM, ##P < 0.01 vs OVX. The liver tissue of five rats from each group was mixed as a sample, three independent samples each group
Effects of Salvianolic acids on lipid levels in LO2 cells

Although STAT-3 signaling is involved in lipid metabolism, the regulatory mechanism of the whole animal model is complex. We determined if salvianolic acids had effects on phosphorylation of STAT-3, contents of TG, and expression of SREBP1 in high-glucose-treated LO2 cells. The used concentration of TSA and SA in the present study had no toxic effects on LO2 cells (data not shown). In accordance with OVX model, salvianolic acids decreased high-glucose-induced phosphorylation of STAT-3 in LO2 cells (Figs. 4A and 4B). Similar results were found for the expression of SREBP1 and the contents of TG (Figs. 5A and 5B). Furthermore, we verified the relationship between STAT-3 and lipogenic genes, and found that knock down of STAT-3 (Figs. 6A and 6B) significantly attenuated the increased contents of TG and expression of SREBP1 induced by high glucose (Figs. 6C and 6D), but had no effect on FAS (Fig. 6E). These results indicated that salvianolic acids inhibited lipogenesis via STAT-3/SREBP1 signaling.

Figure 4 Effects of salvianolic acids on STAT-3 in LO2 cells. LO2 cells were treated with or without E2 (10 nmol·L-1), TSA (15 μg·L-1), SA (15 μg·L-1), under lipogenic conditions (25 mmol·L-1 glucose) for 48 h. (A) Western blot analysis of p-STAT-3, STAT-3, p-AKT and AKT; (B) densitometry quantification of p-STAT-3 and STAT-3 expression and p-AKT and AKT expression. **P < 0.01 vs 5.56 mmol·L-1 glucose; ##P < 0.01 vs 25 mmol·L-1 glucose. n = 3
Figure 5 Effects of salvianolic acids on lipid metabolism in LO2 cells. LO2 cells were treated with or without E2 (10 nmol·L-1), TSA (15 μg·L-1), SA (15 μg·L-1), under lipogenic conditions (25 mmol·L-1 glucose) for 48 h. (A) mRNA expression of SREBP1. (B) LO2 TG contents. **P < 0.01 vs 5.56 mmol·L-1 glucose, #P < 0.05 vs 25 mmol·L-1 glucose, ##P < 0.01 vs 25 mmol·L-1 glucose. n = 3
Figure 6 Relationship between STAT-3 and SREBP1 in LO2 cells. LO2 cells were transfected by 20 nmol·L-1 NC-siRNA or STAT-3-siRNA for 12 h, followed by exposure to 5.56 or 25 mmol·L-1 glucose for 48 h. (A, B) mRNA expression of STAT-3; (C) LO2 TG contents, (D, E) mRNA expression of SREBP1 and FAS. **P < 0.01 vs si-Con. n = 3
Discussion

The salvianolic acids, a group of plant-derived chemicals, have been identified having numerous biological functions. Several studies have reported the beneficial effects of salvianolic acids in the management of cerebrovascular diseases, hepatitis, diabetes, and diabetic complications [3, 27]. Our current study revealed that salvianolic acids inhibited the OVX-induced body weight gain and lipid accumulation in vivo, and the TG contents in LO2 cells in vitro. For the molecular mechanisms, the STAT-3/SREBP1 signaling pathway was involved. Based on these findings, we suggested that the salvianolic acids played an essential role in lipogenesis in OVX rats and in LO2 cells.

Previous studies have indicated that OVX can induce body weight gain and liver TG accumulation due to estrogen deficiency [28]. Increasing food intake via hypothalamus is responsible for OVX-induced body weight gain. E2 has an anorexigenic functions in OVX rats [29]. In our study, E2 could inhibit the OVX-induced body weight gain and food intake. We also found that salvianolic decreased the body weight gain induced by OVX, but had no effect on food intake. Meanwhile, salvianolic acids inhibited the liver TG contents. These results indicated that salvianolic acids inhibited the OVX-induced body weight gain possibly by improving the hepatic lipid metabolism disorders. There were no differences in the inhibitory effects on OVX-induced body weight gain between TSA and SA, due to that Salvianolic acid B is the major constituent of the TSA. The results indicated that salvianolic acid B in TSA and SA was responsible for inhibiting OVX-induced body weight gain.

The synthesis of triglycerides requires the involvement of adipogenic genes [30]. SREBP1, an important nuclear transcription factor, is responsible for the biosynthesis of cholesterols, fatty acids, and TG, favoring the development of hypertriglyceridemia and fatty liver by means of activating the key enzymes of lipogenesis, such as FAS and ACC1 [31]. We found that salvianolic acids suppressed the upregulation of SREBP1, FAS, and SCD1 in OVX rats, suggesting that inhibition of adipogenic genes contributed to suppression of lipid accumulation and thus the improvement of metabolic function.

STAT-3 is an important transcription factor and contributes to various physiological processes [15]. Former studies have confirmed that overexpression of STAT-3 in the liver can increase the levels of atherogenic lipoproteins, likely through alteration of the expression of FAS and ACC involved in lipid metabolism [32]. It has been reported that STAT-3 plays an important role in regulating hepatic insulin resistance induced by palmitate [33]. We found that salvianolic acids inhibited the phosphorylation of STAT-3 induced by OVX. Different with our results, previous studies showed high-fat diet induced obesity owing to inhibition of phosphorylation of STAT-3 [17]. Therefore, we further identified the relationship between lipid metabolism and STAT-3 and explored whether salvianolic acids affected lipid metabolism through STAT-3.

The OVX, due to estrogen deficiency, could increase food intake and much energy storage in body. The high-glucose-treated LO2 cell model in vitro meaning much energy storage in cells, was in accordance with in vivo OVX model. It is reported that high glucose induces adipogenesis and enhances phosphorylation of STAT-3 [34-35]. Previous research has shown that STAT-3 is involved in lipid synthesis by regulating the expression of SREBP1 in high-fat diet [16]. In our study, we found that salvianolic acids inhibited the upregulation of phosphorylation of STAT-3 and SREBP1 and increased contents of TG by high-glucose-treatment in LO2 cells. Furthermore, knock down of STAT-3 significantly attenuated the increased contents of TG and upregulation of SREBP1 induced by high glucose, but had no effect on FAS. Therefore, it demonstrated that phosphorylation of STAT-3 is involved in lipid synthesis. Our in vitro study also confirmed the improvements in lipid metabolism by salvianolic acids in LO2 cells. STAT-3/SREBP1 signaling pathway was involved in salvianolic acids-inhibited synthesis of TG.

In conclusion, our results indicated that STAT-3 played a pivotal role in the regulation of lipid metabolism by salvianolic acids. Salvianolic acids inhibited OVX-induced body weight gain via inactivating STAT-3/SREBP1 signaling. This study provided a new perspective on biochemical and molecular aspects of metabolic complications associated with postmenopausal women.

References
[1]
Xing HC, Li LJ, Xu KJ, et al. Effects of Salvia miltiorrhiza on intestinal microflora in rats with ischemia/reperfusion liver injury[J]. HPPD Int, 2005, 4(2): 274-280.
[2]
Chen CG, Wang YP. Magnesium lithospermate B ameliorates renal cortical microperfusion in rats[J]. Acta Pharmacol Sin, 2006, 27(2): 217-222. DOI:10.1111/aphs.2006.27.issue-2
[3]
Lei XL, Chiou GC. Studies on cardiovascular actions of Salvia miltiorrhiza[J]. Am J Chin Med, 1986, 14(1-2): 26-32.
[4]
Zhou X, Chan K, Yeung JH. Herb-drug interactions with Danshen (Salvia miltiorrhiza): a review on the role of cytochrome P450 enzymes[J]. Drug Metabol Drug Interact, 2012, 27(1): 9-18.
[5]
He X, Shen Q. Salvianolic acid B promotes bone formation by increasing activity of alkaline phosphatase in a rat tibia fracture model: a pilot study[J]. BMC Complement Altern Med, 2014, 14: 493. DOI:10.1186/1472-6882-14-493
[6]
Zhou X, Cheung CM, Yang JM, et al. Danshen (Salvia miltiorrhiza) water extract inhibits paracetamol-induced toxicity in primary rat hepatocytes via reducing CYP2E1 activity and oxidative stress[J]. J Pharm Pharmacol, 2015, 67(7): 980-989. DOI:10.1111/jphp.2015.67.issue-7
[7]
Han JY, Fan JY, Horie Y, et al. Ameliorating effects of compounds derived from Salvia miltiorrhiza root extract on microcirculatory disturbance and target organ injury by ischemia and reperfusion[J]. Pharmacol Ther, 2008, 117(2): 280-295. DOI:10.1016/j.pharmthera.2007.09.008
[8]
Ma L, Guan YQ, Du ZD. Salvianolic acid B down-regulates matrix metalloproteinase-9 activity and expression in tumor necrosis factor-alpha-induced human coronary artery endothelial cells[J]. Chin Med J, 2015, 128(19): 2658-2663. DOI:10.4103/0366-6999.166037
[9]
Boulware MI, Kent BA, Frick KM. The impact of age-related ovarian hormone loss on cognitive and neural function[J]. Curr Top Behav Neurosci, 2012, 10: 165-184.
[10]
Carr MC. The emergence of the metabolic syndrome with menopause[J]. J Clin Endocrinol Metab, 2003, 88(6): 2404-2411. DOI:10.1210/jc.2003-030242
[11]
Kaaja RJ. Metabolic syndrome and the menopause[J]. Menopause Int, 2008, 14(1): 21-25. DOI:10.1258/mi.2007.007032
[12]
Zhu LR, Zou F, Yang YJ, et al. Estrogens prevent metabolic dysfunctions induced by circadian disruptions in female mice[J]. Endocrinology, 2015, 156(6): 2114-2123. DOI:10.1210/en.2014-1922
[13]
North American Menopause Society. The 2012 hormone therapy position statement of: The North American Menopause Society[J]. Menopause, 2012, 19(3): 257-271. DOI:10.1097/gme.0b013e31824b970a
[14]
Al-Safi ZA, Santoro N. Menopausal hormone therapy and menopausal symptoms[J]. Fertil Steril, 2014, 101(4): 905-915. DOI:10.1016/j.fertnstert.2014.02.032
[15]
Miller CN, Della-Fera MA, Baile CA. The mediation of hepatic lipogenesis through estrogens[J]. Postdoc J, 2013, 1(5): 27-38.
[16]
Zeng Z, He W, Jia Z, et al. Lycopene improves insulin sensitivity through inhibition of STAT-3/Srebp-1c-mediated lipid accumulation and inflammation in mice fed a High-Fat Diet[J]. Exp Clin Endocrinol Diabetes, 2017, 125(9): 610-617. DOI:10.1055/s-0043-101919
[17]
Inoue H, Ogawa W, Ozaki M, et al. Role of STAT-3 in regulation of hepatic gluconeogenic genes and carbohydrate metabolism in vivo[J]. Nat Med, 2004, 10(2): 168-174. DOI:10.1038/nm980
[18]
Han JY, Miura S, Akiba Y, et al. Chronic ethanol consumption exacerbates microcirculatory damage in rat mesentery after reperfusion[J]. Am J Physiol Gastrointest Liver Physiol, 2001, 280(5): G939-948. DOI:10.1152/ajpgi.2001.280.5.G939
[19]
Si YY, Li N, Tong L, et al. Bioactive minor components of the total salvianolic acids injection prepared from Salvia miltiorrhiza Bge[J]. Bioorg Med Chem Lett, 2016, 26(1): 82-86. DOI:10.1016/j.bmcl.2015.11.028
[20]
Sun Y, Yu Q, Shen Q, et al. Black cohosh ameliorates metabolic disorders in female ovariectomized rats[J]. Rejuvenation Res, 2016, 19(3): 204-214. DOI:10.1089/rej.2015.1724
[21]
Hoegh-Andersen P, Tanko LB, Andersen TL, et al. Ovariectomized rats as a model of postmenopausal osteoarthritis: validation and application[J]. Arthritis Res Ther, 2004, 6(2): R169-180. DOI:10.1186/ar1152
[22]
Paquette A, Wang D, Jankowski M, et al. Effects of ovariectomy on PPAR alpha, SREBP-1c, and SCD-1 gene expression in the rat liver[J]. Menopause, 2008, 15(6): 1169-1175. DOI:10.1097/gme.0b013e31817b8159
[23]
Rogers NH, Perfield JW, Strissel KJ, et al. Reduced energy expenditure and increased inflammation are early events in the development of ovariectomy-induced obesity[J]. Endocrinology, 2009, 150(5): 2161-2168. DOI:10.1210/en.2008-1405
[24]
Domingos MM, Rodrigues MF, Stotzer US, et al. Resistance training restores the gene expression of molecules related to fat oxidation and lipogenesis in the liver of ovariectomized rats[J]. Eur J Appl Physiol, 2012, 112(4): 1437-1444. DOI:10.1007/s00421-011-2098-6
[25]
Zhang KM, Guo W, Yang Y, et al. JAK2/STAT-3 pathway is involved in the early stage of adipogenesis through regulating C/EBPbeta transcription[J]. J Cell Biochem, 2011, 112(2): 488-497. DOI:10.1002/jcb.22936
[26]
Leavens KF, Easton RM, Shulman GI, et al. Akt2 is required for hepatic lipid accumulation in models of insulin resistance[J]. Cell Metab, 2009, 10(5): 405-418. DOI:10.1016/j.cmet.2009.10.004
[27]
Huang MQ, Xie YL, Chen LD, et al. Antidiabetic effect of the total polyphenolic acids fraction from Salvia miltiorrhiza Bunge in diabetic rats[J]. Phytother Res, 2012, 26(6): 944-948. DOI:10.1002/ptr.v26.6
[28]
Camara C, Zhou LY, Ma Y, et al. Effect of ovariectomy on serum adiponectin levels and visceral fat in rats[J]. J Huazhong Univ Sci Technol Med Sci, 2014, 34(6): 825-829. DOI:10.1007/s11596-014-1360-7
[29]
Roepke TA. Oestrogen modulates hypothalamic control of energy homeostasis through multiple mechanisms[J]. J Neuroendocrinol, 2009, 21(2): 141-150. DOI:10.1111/jne.2009.21.issue-2
[30]
Panneerselvam S, Packirisamy RM, Bobby Z, et al. Soy isoflavones (Glycine max) ameliorate hypertriglyceridemia and hepatic steatosis in high fat-fed ovariectomized Wistar rats (an experimental model of postmenopausal obesity)[J]. J Nutr Biochem, 2016, 38: 57-69. DOI:10.1016/j.jnutbio.2016.08.007
[31]
Grefhorst A, Elzinga BM, Voshol PJ, et al. Stimulation of lipogenesis by pharmacological activation of the liver X receptor leads to production of large, triglyceride-rich very low density lipoprotein particles[J]. J Biol Chem, 2002, 277(37): 34182-34190. DOI:10.1074/jbc.M204887200
[32]
Kinoshita S, Ogawa W, Okamoto Y, et al. Role of hepatic STAT-3 in the regulation of lipid metabolism[J]. Kobe J Med Sci, 2008, 54(4): E200-208.
[33]
Xia NN, Tang ZQ, Wang CF, et al. PCBP2 regulates hepatic insulin sensitivity via HIF-1alpha and STAT-3 pathway in HepG2 cells[J]. Biochem Biophys Res Commun, 2015, 463(1-2): 116-122. DOI:10.1016/j.bbrc.2015.04.150
[34]
Guillet-Deniau I, Pichard AL, Kone A, et al. Glucose induces de novo lipogenesis in rat muscle satellite cells through a sterol-regulatory-element-binding-protein-1c-dependent pathway[J]. J Cell Sci, 2004, 117: 1937-1944. DOI:10.1242/jcs.01069
[35]
Saengboonmee C, Seubwai W, Pairojkul C, et al. High glucose enhances progression of cholangiocarcinoma cells via STAT-3 activation[J]. Sci Rep, 2016, 6: 18995. DOI:10.1038/srep18995