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. 2017 Dec 20;17(1):545.
doi: 10.1186/s12906-017-2047-y.

Effects of Salvia miltiorrhiza extract with supplemental liquefied calcium on osteoporosis in calcium-deficient ovariectomized mice

Bongkyun Park  1 , Hae Seong Song  1 , Jeong Eun Kwon  1 , Se Min Cho  1 , Seon-A Jang  1 , Mi Yeon Kim  2 , Se Chan Kang  3
Affiliations

Effects of Salvia miltiorrhiza extract with supplemental liquefied calcium on osteoporosis in calcium-deficient ovariectomized mice

Bongkyun Park et al. BMC Complement Altern Med. .

Abstract

Background: Extracts from Salvia miltiorrhiza Bunge have been used in traditional Asian medicine to treat coronary heart disease, chronic renal failure, atherosclerosis, myocardial infraction, angina pectoris, myocardial ischemia, dysmenorrheal, neurasthenic insomnia, liver fibrosis and cirrhosis. The aim of the study was to investigate the anti-RANK signal effect of the combination of S.miltiorrhiza Bunge (SME) and liquefied calcium (LCa) supplement with ovariectomized (OVX-SML) mice, a osteoporosis animal model. Results were compared to 17β-estradiol (E2) treatment.

Methods: A total of 70 female ICR strain mice (7 weeks) were randomly divided into 10 groups with 7 mice in each group as follows: (1) sham-operated control mice (sham) received daily oral phosphate-buffered-saline (PBS) of equal volumes through oral administration. (2) OVX mice received a daily oral administration of PBS (OVX). (3) OVX mice treated daily with 50 mg/kg b.w./ day of SME (4) with 100 mg/kg b.w./day of SME or (5) with 200 mg/kg b.w./day of SME via oral administration. (6) OVX mice treated daily with 50 mg/kg b.w./day of SML (7) with 100 mg/kg b.w./day of SML or (8) with 200 mg/kg b.w./day of SML via oral administration. (9) OVX mice treated daily with 10 ml/kg b.w./day of LCa (10) OVX mice received i.p. injections of 17β-estradiol (E2) (0.1 mg/kg b.w./day) three times per week for 12 weeks.

Results: micro-CT analysis revealed that oral administration of SML inhibited tibial bone loss, sustained trabecular bone state, and ameliorated bone biochemical markers. In addition, SML administration compared to SEM and LCa reduced serum levels of RANKL, osteocalcin and BALP through increased serum levels of OPG and E2 in OVX mice. SML also had more beneficial effects on protection of estrogen-dependent bone loss through blocking expression of TRAF6 and NFTAc1 and produces cathepsin K and calcitonin receptor to develop osteoclast differentiation.

Conclusion: These data suggest that S. miltiorrhiza Bunge combined with liquefied calcium supplement has an inhibitory activity in OVX mice. This result implies the possibility of a pharmacological intervention specifically directed toward a disease such as osteoporosis where decreased bone strength increases the risk of a broken bone .

Keywords: Liquefied calcium supplement; OPG; Ovariectomized mice; RANKL; Salvia miltiorrhiza Bunge.

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Conflict of interest statement

Authors’ information

BP is a research professor in Department of Oriental Medicine Biotechnology, Kyung Hee University.

HSS, JEK, SMC and SAJ are a graduate student in Department of Oriental Medicine Biotechnology, Kyung Hee University and is supervised by SCK.

MYK is a senior researcher in KMF corp.

SCK is a professor in in Department of Oriental Medicine Biotechnology, Kyung Hee University

Ethics approval

The Experimental Animal Facility and protocols were approved by the Institutional Animal Care and Use Committee of Kyung Hee University.

Consent for publication

Not applicable

Competing interests

The authors declare that they have no competing interests.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1
Fig. 1
Effects of SML on body weight and uterus weight in OVX mice. The uterus weights were collected at the conclusion of the study. Values are given as mean ± S.E.M. of three independent experiments (n = 7) for all groups. Means unlike letters in a column with differ significantly (p < 0.05)
Fig. 2
Fig. 2
Effects of SML on the structural characteristics of cortical bone. a Three-dimensional micro-CT images of cortical bone after 12 weeks in OVX mice. The cortical bone parameters were measured by a three-dimensional micro-CT analysis program. The measured cortical bone parameters were b bone volume density (BV, mm3), c mean polar moment of inertia (MMI, mm4), d cross-section thickness (Cs.Th, mm), and e bone mineral density (BMD, g/cm3). Values are given as the mean ± S.E.M. of three independent experiments (n = 7) for all groups. Means unlike letters in a column with differ significantly (p < 0.05)
Fig. 3
Fig. 3
Effects of SML on the mechanical characteristics of trabecular bone. a Three-dimensional micro-CT images of trabecular bone after 12 weeks in OVX mice. The trabecular bone parameters were monitored by a three-dimensional micro-CT analysis program. The measured trabecular bone parameters were b bone volume fraction (BV/TV), c trabecular thickness (Tb.Th), d trabecular separation (Tb.Sp), e trabecular number (Tb.N), f trabecular bone pattern factor (Tb.Pf), g specific bone surface (BS/BV), h structure model index (SMI), and i bone mineral density (BMD). Values are given as mean ± S.E.M. of three independent experiments (n = 7) for all groups. Means unlike letters in a column with differ significantly (p < 0.05)
Fig. 4
Fig. 4
Effects of SML on serum biochemical markers in OVX mice. a RANKL and b OPG were determined by murine RANKL and OPG ELISA assays using serum, and c the RANKL/OPG ratio was calculated. d E2, e osteocalcin, and f BALP were determined by murine ELISA assays. Data are the mean ± S.E.M. of three independent experiments (n = 7) for all groups. Means unlike letters in a column with differ significantly (p < 0.05)
Fig. 5
Fig. 5
Effects of SML on the expression of cathepsin K and calcitonin receptor (CalcR) in OVX mice. RNA was extracted from the bone marrow cells of OVX mice, and a the mRNA levels of cathepsin K and b calcitonin receptor were assessed by real-time PCR assay. β-actin was considered an internal control. Data are the mean ± S.E.M. of three independent experiments (n = 7) for all groups. Means unlike letters in a column with differ significantly (p < 0.05)
Fig. 6
Fig. 6
Effects of SML on the expression of TRAF6 and NFATc1 in OVX mice. RNA was extracted from the bone marrow cells of OVX mice, and a the mRNA levels of TRAF6 and b NFATc1 were assessed by real-time PCR assay. β-actin was considered an internal control. Data are the mean ± S.E.M. of three independent experiments (n = 7) for all groups. Means unlike letters in a column with differ significantly (p < 0.05)
Fig. 7
Fig. 7
Schematic diagram showing the inhibitory effects of SML on OVX-induced osteoporosis via regulation of RANK/RANKL/OPG signaling pathways
See this image and copyright information in PMC

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