[논문]난소 적출 흰쥐 골다공증 모델에서 금은화(金銀花)가 골밀도 증가에 미치는 효과

이성엽; 김민선; 홍수연; 김재현; 김홍식; 이충호; 정혁상; 손영주

doi:10.6116/kjh.2021.36.5.81.

[국내논문] 난소 적출 흰쥐 골다공증 모델에서 금은화(金銀花)가 골밀도 증가에 미치는 효과
Effect of Lonicerae Japonicae Flos on Bone Density in Ovariectomized Rat Model of Osteoporosis 원문보기

大韓本草學會誌 = The Korea journal of herbology, v.36 no.5, 2021년, pp.81 - 91

이성엽 (경희대학교 한의과대학 해부학교실) , 김민선 (경희대학교 한의과대학 해부학교실) , 홍수연 (경희대학교 한의과대학 해부학교실) , 김재현 (경희대학교 한의과대학 해부학교실) , 김홍식 (경희대학교 한의과대학 해부학교실) , 이충호 (경희대학교 한의과대학 해부학교실) , 정혁상 (경희대학교 한의과대학 해부학교실) , 손영주 (경희대학교 한의과대학 해부학교실)

Abstract ▼ AI-Helper

Objectives : Osteoporosis is a systemic skeletal disease that decreases bone density and increases the risk of fractures. Bisphosphonates and SERMs are mainly used to treat osteoporosis, but, long-term use increases the risk of side effects such as jaw bone necrosis and breast cancer. Therefore, it is necessary to develop a therapeutic agent for a natural product with few side effects. Water extract of Lonicerae Japonicae Flos (wLF) was mainly found to have anti-cancer and anti-inflammatory effects. However, the effect of wLF on osteoporosis has not been elucidated. Therefore, this experiment investigated the effect of wLF on osteoclasts, osteoblasts and osteoporosis models. Methods : In order to study the effect of wLF on osteoporosis, the OVX-induced rat model was used for in vivo study. After 8 weeks, we measured body weight, uterine weight, liver weight, femur weight, bone density, trabecular area and tibia ash weight. To determine the effect of wLF on osteoclast differentiation, we measured the number of TRAP-positive cells and TRAP activity. To examine the effect of wLF on the expression of osteoblast-related genes, we measured the mRNA expression of alkaline phosphatase (ALP, Alpl) and osteocalcin (OCN, Bglap2). Results : In vivo experiment, wLF inhibited the reduction of femur weight, trabecular area, bone density and tibia ash weight. In vitro experiment, wLF had no significant effect on osteoclast differentiation. However, wLF increased the mRNA expression of Alpl and Bglap2 in MC3T3-E1 cell. Conclusions : This result suggested that wLF may be used for the treatment and prevention of postmenopausal osteoporosis.

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표/그림 (8)

표 Table 1. PCR primer sequence for RT-PCR analysis.
그림 Fig. 1. The change of body, uterus, liver and Femur weight in OVX-induced rats. (A) Weekly change of body weight after OVX surgery. (B) Body weight gain after OVX-induced surgery. The uterus, liver and femurs were collected and weighed after all animals were sacrificed. (C) Uterus weight and (D) liver (E) femur weights per 100 g body weight. Each data value represents the mean ± standard error of mean (SEM, n=8). ##p<0.01 compared with Sham-group, **p<0.01 compared with OVX-group.
그림 Fig. 2. Effect of wLF on BMD and tibia ash weight in OVX-induced rats. (A) BMD was measured by Archimedes' principle. (B) Tibia ash was weighed after all animals were sacrificed. Each data value represents the mean ± standard error of mean (SEM, n=8). ##p <0.01 compared with sham group and *p<0.05, **p<0.01 compared with OVX-induced group.
그림 Fig. 3. Effect of wLF on trabecular area in OVX-induced rats. Femur sections (A) were stained with H&E staining and trabecular area (B) was measured by Image J software. Each data value represents the mean ± standard error of mean (SEM, n=8). ##p<0.01 compared with Sham group and *p<0.05, **p<0.01 compared with OVX-induced group.
그림 Fig. 4. Effect of wLF on RAW 264.7 cell viability. Cell viability was measured by MTS assay. RAW 264.7 cells were seeded into 96 well plate. After 24 h, cells were treated with various concentrations of wLF (1, 10, 100 and 200 μg/mL) for 24 h and then the cells were treated with MTS solution 20 μl. After 2 h, optical density was measured at 490 nm with an ELISA reader. Data represent the mean ± standard error of mean (SEM) of three independent experiments.
그림 Fig. 5. The effect of wLF on RANKL-induced osteoclast formation in RAW 264.7 cells. (A) TRAP-positive cells were stained with TRAP staining kit, and captured using inverted microscope (‘100). (B) TRAP activity were analyzed with enzyme-Linked immunosorbent assay (ELISA) reader. Data represent the mean ± standard error of mean (SEM) of three independent experiments.
그림 Fig. 6. Effect of wLF on MC3T3-E1 cell viability. Cell viability was measured by MTS assay. MC3T3-E1 cells were seeded into 96 well plate. After 24 h, cells were treated with various concentrations of wLF (10, 20, 40, 60, 80 and 100 μg/mL) for 72 h, and then the cells were treated with MTS solution 20 μL. After 2 h, optical density was measured at 490 nm with an ELISA reader. Data represent the mean ± standard error of mean (SEM) of three independent experiments.
그림 Fig. 7. Effect of wLF on mRNA expression of ALP and osteocalcin in MC3T3-E1 cells. (A) ALP (Alpl) was meseured by RT-PCR. Alpl normalized to Gapdh (Gapdh). (C) OCN (Bglap2) was meseured by RT-PCR. Alpl normalized Gapdh. Data represent the mean ± standard error of mean (SEM) of three independent experiments. *p<0.05 compared with Normal group (non-treated group).

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