卵巢切除小鼠的骨重塑特征研究

doi:10.11958/20250846

天津医药 ›› 2025, Vol. 53 ›› Issue (6): 566-570.doi: 10.11958/20250846

卵巢切除小鼠的骨重塑特征研究

李林森(), 冯玉梅()

天津医科大学肿瘤医院，国家恶性肿瘤临床医学研究中心，天津市恶性肿瘤临床医学研究中心，天津市肿瘤防治重点实验室（邮编300060）

收稿日期:2025-03-03 修回日期:2025-04-10 出版日期:2025-06-15 发布日期:2025-06-20
通讯作者: ^△E-mail:ymfeng@tmu.edu.cn
作者简介:李林森（1999），男，硕士在读，主要从事生物化学与分子生物学方面研究。E-mail:lol97144245@163.com
基金资助:
天津市医学重点学科（专科）建设项目(TJYXZDXK-009A)

Study on the characteristics of bone remodeling in ovariectomized mice

LI Linsen(), FENG Yumei()

Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer; Tianjin's Clinical Research Center for Cancer; Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China

Received:2025-03-03 Revised:2025-04-10 Published:2025-06-15 Online:2025-06-20
Contact: ^△E-mail: ymfeng@tmu.edu.cn

李林森, 冯玉梅. 卵巢切除小鼠的骨重塑特征研究[J]. 天津医药, 2025, 53(6): 566-570.

LI Linsen, FENG Yumei. Study on the characteristics of bone remodeling in ovariectomized mice[J]. Tianjin Medical Journal, 2025, 53(6): 566-570.

摘要/Abstract

摘要：

目的利用卵巢切除小鼠模型探讨雌激素缺乏对骨重塑特征的影响。方法将10只12周龄雌性C57BL/6J小鼠随机分为假手术（Sham）组和卵巢切除术（OVX）组，每组5只。术后16周分离小鼠子宫，观察形态并统计子宫质量。CT扫描分析小鼠股骨远端，抗酒石酸酸性磷酸酶（TRAP）染色评估股骨组织石蜡切片的破骨细胞数量。采用CD11b磁珠分选小鼠股骨骨髓的骨髓源性间充质干细胞（BM-MSCs）和骨髓来源的巨噬细胞（BMMs）。碱性磷酸酶（ALP）染色评估BM-MSCs成骨分化潜能；反转录-定量PCR检测BM-MSCs成骨分化相关基因（Runx2、Sp7和Spp1）及BMMs破骨细胞分化相关基因（Ctsk、Acp5和Cd40）的表达水平。结果与Sham组相比，OVX组的子宫出现明显萎缩，质量减轻；骨矿物质密度（BMD）、骨体积分数（BV/TV）、骨表面积组织体积比（BS/TV）、骨小梁数量（Tb.N）、骨小梁厚度（Tb.Th）、皮质骨矿物质密度（Ct.BMD）和皮质骨厚度（Ct.Th）明显减少，骨小梁分离度（Tb.Sp）明显增加（P<0.01）；与Sham组相比，OVX组BM-MSCs中Runx2、Sp7和Spp1 mRNA表达水平降低（P<0.01）；诱导后，其成骨分化程度也减低。与Sham组相比，OVX组BMMs中Ctsk、Acp5和Cd40的mRNA表达水平升高，股骨远端骨髓中破骨细胞数量多于Sham组（P<0.01）。结论 OVX导致的雌激素缺乏在减弱成骨分化能力的同时增强了破骨细胞的分化能力，最终引起骨稳态失调，造成骨质疏松。

关键词: 卵巢切除术, 雌激素类, 巨噬细胞, 间质干细胞, 骨重塑

Abstract:

Objective To explore the effect of estrogen deficiency on bone remodeling characteristics in an ovariectomized mouse model. Methods Ten 12-week-old female C57BL/6J mice were randomly divided into the sham operation group and the ovariectomy (OVX) group, with 5 mice in each group. The uteri of mice were isolated at 16 weeks postoperatively. The morphology was observed, and uterine weights were counted. CT scans were performed to analyze the distal femur of the mice, and anti-tartrate acid phosphatase (TRAP) staining was performed to assess the number of osteoclasts in paraffin sections of femoral tissue. Bone marrow-derived mesenchymal stem cells (BM-MSCs) and bone marrow-derived macrophages (BMMs) were sorted from femoral bone marrow of mice using CD11b magnetic beads. Alkaline phosphatase (ALP) staining was performed to assess the osteogenic differentiation potential of BM-MSCs. Reverse transcription-quantitative PCR (RT-qPCR) was performed to detect the expression levels of genes related to osteogenic differentiation of BM-MSCs (Runx2, Sp7, and Spp1) and osteoclast differentiation of BMMs (Ctsk, Acp5, and Cd40). Results Compared with the sham group, the uterus of the OVX group showed significant atrophy and weight loss, and bone mineral density (BMD), bone volume fraction (BV/TV), bone surface area tissue volume ratio (BS/TV), number of trabeculae (Tb.N), trabecular thickness (Tb.Th), cortical bone mineral density (Ct.BMD) and cortical bone thickness (Ct.Th) were significantly reduced, and bone trabecular separation (Tb.Sp) was significantly increased (P<0.01). The mRNA expression levels of Runx2, Sp7 and Spp1 were reduced in BM-MSCs of the OVX group compared with the sham group (P<0.01). The osteogenic differentiation was also diminished after induction. The mRNA expression levels of Ctsk, Acp5 and Cd40 were elevated in the BMMs of the OVX group, and the number of osteoclasts in bone marrow of the distal femur was greater in the OVX group than that in the sham group (P<0.01). Conclusion OVX-induced estrogen deficiency can enhance osteoclast differentiation and attenuate osteogenic differentiation capacity, ultimately causing dysregulation of bone homeostasis and resulting in osteoporosis.

Key words: ovariectomy, estrogens, macrophages, mesenchymal stem cells, bone remodeling

中图分类号:

R332

图/表 9

表1 引物序列

Tab.1 Primer sequences

基因名称	引物序列（5'→3'）	产物大小/bp
Ctsk	上游：GAGGGCCAACTCAAGAAGAA 下游：TTGGAAGGCAGTGGTCATATAG	117
Acp5	上游：CCTGAGATTTGTGGCTGTGG 下游：TCTTGTCGCTGGCATCGTG	179
Cd40	上游：GTCGGCTTCTTCTCCAATCA 下游：CTGACTCGTTCCTTTCTGTAGG	102
Runx2	上游：CTCTGCACCAAGTCCTTTTAATC 下游：AGGAGGGGTAAGACTGGTCATAG	104
Sp7	上游：CTGAGAGAGGAGCAGATCCC 下游：GTGAGCTTCTTCCTGGGTAGG	120
Spp1	上游：ATCTCACCATTCGGATGAGTCT 下游：TGTAGGGACGATTGGAGTGAAA	79
GAPDH	上游：ATTGTCAGCAATGCATCCTG 下游：ATGGACTGTGGTCATGAGCC	102

图1 2组小鼠子宫形态比较

Fig.1 Comparison of uterine morphology between the two groups of mice

图2 术后16周时2组小鼠股骨远端micro-CT扫描图

Fig.2 Micro-CT scans of the distal femur at 16 weeks postoperatively in 2 groups of mice

表2 2组小鼠骨小梁计量学分析（n=5，$\bar{x} \pm s$）

Tab.2 Trabecular metrological analysis of bone in 2 groups of mice

组别	BMD/（mg/cm³）	BV/TV/%	BS/TV/（1/mm）
Sham组	181.00±6.38	30.95±1.72	43.12±3.62
OVX组	132.90±7.49	15.11±2.89	27.77±2.08
t	10.910^＊＊	10.520^＊＊	8.220^＊＊
组别	Tb.N/（1/cm）	Tb.Th/mm	Tb.Sp/cm
Sham组	10.72±0.54	0.053±0.003	0.044±0.004
OVX组	8.24±0.44	0.044±0.002	0.076±0.006
t	7.962^＊＊	6.796^＊＊	9.868^＊＊

表3 2组小鼠皮质骨计量学分析（n=5，$\bar{x} \pm s$）

Tab.3 Cortical osteometric analysis in 2 groups of mice

组别	Ct.BMD/（mg/cm³）	Ct.Th/cm
Sham组	886.80±22.41	0.050±0.001
OVX组	815.10±16.01	0.042±0.003
t	5.816^＊＊	6.558^＊＊

表4 2组小鼠BM-MSCs诱导分化前成骨分化标志基因表达水平比较（n=3，$\bar{x} \pm s$）

Tab.4 Comparison of expression levels of osteogenic differentiation marker genes before induction of BM-MSCs between two groups of mice

组别	Runx2	Sp7	Spp1
Sham组	1.00±0.00	1.00±0.00	1.00±0.00
OVX组	0.40±0.02	0.55±0.06	0.34±0.03
t	56.790^＊＊	12.500^＊＊	44.760^＊＊

图3 2组小鼠BM-MSCs成骨诱导后的ALP染色

Fig.3 ALP staining of BM-MSCs after osteogenic induction in 2 groups of mice

表5 2组小鼠BMMs诱导分化前破骨分化标志基因表达水平比较（n=3，$\bar{x} \pm s$）

Tab.5 Comparison of expression levels of osteoclast differentiation marker genes in BMMs before induction of differentiation between two groups of mice

组别	Ctsk	Acp5	Cd40
Sham组	1.00±0.00	1.00±0.00	1.00±0.00
OVX组	4.76±0.07	2.90±0.25	2.39±0.05
t	87.030^＊＊	13.340^＊＊	44.220^＊＊

图4 2组小鼠股骨组织TRAP染色结果（×400）红色箭头示TRAP阳性细胞。

Fig.4 Results of TRAP staining of femoral tissue in two groups of mice (×400)

参考文献 14

[1]	HARVEY N, DENNISON E, COOPER C. Osteoporosis:impact on health and economics[J]. Nat Rev Rheumatol, 2010, 6(2):99-105. doi:10.1038/nrrheum.2009.260.
[2]	EMMANUELLE N E, MARIE-CÉCILE V, FLORENCE T, et al. Critical role of estrogens on bone homeostasis in both male and female:from physiology to medical implications[J]. Int J Mol Sci, 2021, 22(4):1568. doi:10.3390/ijms22041568.
[3]	STREICHER C, HEYNY A, ANDRUKHOVA O, et al. Estrogen regulates bone turnover by targeting RANKL expression in bone lining cells[J]. Sci Rep, 2017, 7(1):6460. doi:10.1038/s41598-017-06614-0.
[4]	WENG Y, WANG H, WU D, et al. A novel lineage of osteoprogenitor cells with dual epithelial and mesenchymal properties govern maxillofacial bone homeostasis and regeneration after MSFL[J]. Cell Res, 2022, 32(9):814-830. doi:10.1038/s41422-022-00687-x.
[5]	YANG C, TAO H, ZHANG H, et al. TET2 regulates osteoclastogenesis by modulating autophagy in OVX-induced bone loss[J]. Autophagy, 2022, 18(12):2817-2829. doi:10.1080/15548627.2022.2048432.
[6]	ANDREW T W, KOEPKE L S, WANG Y, et al. Sexually dimorphic estrogen sensing in skeletal stem cells controls skeletal regeneration[J]. Nat Commun, 2022, 13(1):6491. doi:10.1038/s41467-022-34063-5.
[7]	KOMORI T. Regulation of skeletal development and maintenance by Runx2 and Sp7[J]. Int J Mol Sci, 2024, 25(18):10102. doi:10.3390/ijms251810102.
[8]	KOMORI T. Bone development by Hedgehog and Wnt signaling,Runx2,and Sp7[J]. J Bone Miner Metab, 2025, 43(1):33-38. doi:10.1007/s00774-024-01551-1.
[9]	KUMARI A, KASHYAP D, GARG V K. Osteopontin in cancer[J]. Adv Clin Chem, 2024, 118:87-110. doi:10.1016/bs.acc.2023.11.002.
[10]	PENG Y, SHI K, WANG L, et al. Characterization of Osterix protein stability and physiological role in osteoblast differentiation[J]. PLoS One, 2013, 8(2):e56451. doi:10.1371/journal.pone.0056451.
[11]	CHENG C H, CHEN L R, CHEN K H. Osteoporosis due to hormone imbalance:an overview of the effects of estrogen deficiency and glucocorticoid overuse on bone turnover[J]. Int J Mol Sci, 2022, 23(3):1376. doi:10.3390/ijms23031376.
[12]	BOYCE B F, XING L. Biology of RANK,RANKL,and osteoprotegerin[J]. Arthritis Res Ther, 2007, 9(Suppl 1):S1. doi:10.1186/ar2165.
[13]	BALKAN W, MARTINEZ A F, FERNANDEZ I, et al. Identification of NFAT binding sites that mediate stimulation of cathepsin K promoter activity by RANK ligand[J]. Gene, 2009, 446(2):90-98. doi:10.1016/j.gene.2009.06.013.
[14]	HAYMAN A R. Tartrate-resistant acid phosphatase (TRAP) and the osteoclast/immune cell dichotomy[J]. Autoimmunity, 2008, 41(3):218-223. doi:10.1080/08916930701694667.

[1]	钟玉梅, 何流, 张敏, 周海燕. 艾灸调控巨噬细胞不同表型极化治疗RA滑膜炎症的疗效[J]. 天津医药, 2025, 53(3): 236-241.
[2]	范慧娟, 刘涛涛, 李楠, 夏时海. 间充质干细胞治疗急性胰腺炎的机制及研究进展[J]. 天津医药, 2025, 53(3): 331-336.
[3]	马莉莉, 李子沐, 王亮, 许彭, 李秀梅. 间充质干细胞外泌体对食管癌ECA109细胞生物学行为的影响[J]. 天津医药, 2025, 53(2): 113-117.
[4]	张亚伟, 齐赛卿, 申志国, 李君朝. 注射用丹参多酚酸盐联合替罗非班治疗急性脑梗死的疗效观察[J]. 天津医药, 2025, 53(2): 203-207.
[5]	郝成飞, 陈国杉, 张西波, 戚峰. 肿瘤相关巨噬细胞的研究进展[J]. 天津医药, 2025, 53(1): 108-112.
[6]	贾维宁, 鲍亚玲, 雷慧, 殷晓宁. 夏枯草提取物对脓毒症小鼠炎症反应和腹腔巨噬细胞的影响[J]. 天津医药, 2024, 52(9): 930-935.
[7]	周丽芸, 王岩, 董本超, 杨培川, 申佳慧, 马剑雄, 马信龙. 干细胞机械敏感性对抗骨质疏松症研究进展[J]. 天津医药, 2024, 52(8): 877-881.
[8]	钟家帅, 冯玉梅. 小鼠骨髓和脂肪间充质干细胞定向分化能力的比较研究[J]. 天津医药, 2024, 52(2): 129-135.
[9]	陈俊杰, 许雪梅, 李岩, 傅金英. 八珍汤合白术附子汤对多囊卵巢综合征大鼠糖代谢及雌激素水平的调节作用[J]. 天津医药, 2024, 52(2): 154-160.
[10]	王鑫瑶, 杨惠, 李冰冰. 间充质干细胞在子宫内膜异位症中的研究进展[J]. 天津医药, 2024, 52(2): 215-219.
[11]	张睿, 陈思思, 王彤丹, 于珮. KLF4通过促进自噬减轻高糖浓度下巨噬细胞胆固醇沉积[J]. 天津医药, 2024, 52(10): 1014-1019.
[12]	侯永波, 余骏马, 朱海娟. 外泌体在心肌梗死治疗中的研究新进展[J]. 天津医药, 2023, 51(9): 1016-1019.
[13]	李宏军, 钱亮, 邓新超, 余庭, 吴岩, 吴红丽. 舒筋活血胶囊通过JAK2/STAT3通路缓解大鼠膝骨关节炎的机制研究[J]. 天津医药, 2023, 51(9): 961-967.
[14]	王娟, 赵玉霞, 茹雅维, 汪子涵, 付婷, 李光. 二甲双胍对Ⅰ型成骨不全小鼠间充质干细胞成骨分化的影响及疗效探讨[J]. 天津医药, 2023, 51(8): 830-833.
[15]	杨雨涛, 王袁, 谢嘉欣, 付霞霏. 人骨髓间充质干细胞来源外泌体分离、鉴定及卵巢摄取[J]. 天津医药, 2023, 51(5): 468-472.

卵巢切除小鼠的骨重塑特征研究

Study on the characteristics of bone remodeling in ovariectomized mice

引用本文

RichHTML

PDF (PC)

可视化

摘要/Abstract

使用本文

图/表 9

参考文献 14

相关文章 15

编辑推荐

Metrics

本文评价