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

doi:10.11958/20221341

天津医药 ›› 2023, Vol. 51 ›› Issue (8): 830-833.doi: 10.11958/20221341

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

王娟(), 赵玉霞, 茹雅维, 汪子涵, 付婷, 李光^△()

天津医科大学基础医学院遗传学系（邮编300070）

收稿日期:2022-09-01 修回日期:2023-02-28 出版日期:2023-08-15 发布日期:2023-08-10
通讯作者: ^△E-mail：lig@tmu.edu.cn
作者简介:王娟（1985），女，讲师，主要从事成骨不全症的治疗方面研究。E-mail：juanwang629@tmu.edu.cn
基金资助:
天津市教委科研计划项目(KJ010168)

Study of effects of metformin on osteogenic differentiation of mesenchymal stem cells and bone tissue regeneration in type I osteogenesis imperfecta mice

WANG Juan(), ZHAO Yuxia, RU Yawei, WANG Zihan, FU Ting, LI Guang^△()

Department of Genetics, School of Basic Medicine, Tianjin Medical University, Tianjin 300070, China

Received:2022-09-01 Revised:2023-02-28 Published:2023-08-15 Online:2023-08-10
Contact: ^△E-mail: lig@tmu.edu.cn

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.

WANG Juan, ZHAO Yuxia, RU Yawei, WANG Zihan, FU Ting, LI Guang. Study of effects of metformin on osteogenic differentiation of mesenchymal stem cells and bone tissue regeneration in type I osteogenesis imperfecta mice[J]. Tianjin Medical Journal, 2023, 51(8): 830-833.

摘要/Abstract

摘要：

目的研究二甲双胍对Ⅰ型成骨不全小鼠（oim）脂肪间充质干细胞（oimADSCs）向成骨细胞分化的影响并初步探讨二甲双胍在oim中的治疗效果。方法从oim中分离培养oimADSCs，通过观察细胞形态，碱性磷酸酶（ALP）染色和油红O染色对其分化能力进行鉴定；将oimADSCs分组进行诱导培养，通过观察ALP染色、检测ALP活性和实时荧光定量聚合酶链反应（qPCR）检测几种成骨分化相关分子的表达情况，分析二甲双胍对oimADSCs向成骨细胞分化的影响；利用微计算机断层扫描技术（Micro-CT）分析股骨的骨骼微观结构，观察二甲双胍在oim模型小鼠中的治疗效果。结果在二甲双胍处理oimADSCs后，ALP染色显著加深、ALP活性显著提高，骨钙素（Bglap）、Ⅰ型胶原（Col1a1）、Runt相关转录因子2（Runx2）以及骨形成蛋白2（BMP2）的mRNA表达水平升高；Micro-CT分析发现oim模型小鼠经过二甲双胍治疗后，股骨骨小梁的骨体积/总体积增加，股骨骨小梁数目增多、厚度增加。结论二甲双胍能够促进oimADSCs向成骨细胞分化，改善oim的骨骼生长情况。

关键词: 成骨不全, 间质干细胞, 二甲双胍, Ⅰ型成骨不全, 脂肪间充质干细胞, 成骨分化

Abstract:

Objective To evaluate the effect of metformin on the osteogenic differentiation of adipose derived mesenchymal stem cells of type I osteogenesis imperfecta mice (oimADSCs), and the therapeutic effect of metformin on bone tissue regeneration in oim mice. Methods The oimADSCs were isolated and cultured. Cell morphology observation, alkaline phosphatase staining and oil red O staining were used to validate the multiple differentiation potential of oimADSCs．After osteogenic induction，ALP staining and ALP activity were compared between groups. qPCR was applied to analyze the expression of osteogenic differentiation associated genes. Micro-CT was used to detect the femoral microstructure and analyze effects of metformin on bone regeneration in oim mice. Results Compared with the control group, ALP staining was more deeper and the ALP activity level was obviously increased in the osteogenesis+metformin group. The transcription levels of Bglap, Col1a1, Runx2 and BMP2 were significantly up-regulated. The micro-CT results revealed that after metformin treatment, BV/TV, trabecular number and Tb.Th of oim mice were increased. Conclusion Metformin can enhance osteogenic differentiation of oimADSCs in vitro and improve bone microstructure of oim mice in vivo．

Key words: osteogenesis imperfecta, mesenchymal stem cells, metformin, type I osteogenesis imperfecta, adipose derived mesenchymal stem cells, osteogenic differentiation

中图分类号:

R681.1

图/表 7

参考文献 22

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基因名称	引物序列（5′→3′）	产物大小（bp）
Bglap	上游：CTGACCTCACAGATGCCAAGCC	191
	下游：CATACTGGTCTGATAGCTCGTCACAAG	191
Col1a1	上游：GACAGGCGAACAAGGTGACAGAG	85
	下游：CAGGAGAACCAGGAGAACCAGGAG	85
Runx2	上游：GCAGCAGCAGCAGCAGGAG	181
	下游：GCACGGAGCACAGGAAGTTGG	181
BMP2	上游：AACACCGTGCGCAGCTTCCATC	142
	下游：CGGAAGATCTGGAGTTCTGCAG	142
GAPDH	上游：CATCACTGCCACCCAGAAGACTG	152
	下游：ATGCCAGTGAGCTTCCCGTTCAG	152

基因名称	引物序列（5′→3′）	产物大小（bp）
Bglap	上游：CTGACCTCACAGATGCCAAGCC	191
	下游：CATACTGGTCTGATAGCTCGTCACAAG	191
Col1a1	上游：GACAGGCGAACAAGGTGACAGAG	85
	下游：CAGGAGAACCAGGAGAACCAGGAG	85
Runx2	上游：GCAGCAGCAGCAGCAGGAG	181
	下游：GCACGGAGCACAGGAAGTTGG	181
BMP2	上游：AACACCGTGCGCAGCTTCCATC	142
	下游：CGGAAGATCTGGAGTTCTGCAG	142
GAPDH	上游：CATCACTGCCACCCAGAAGACTG	152
	下游：ATGCCAGTGAGCTTCCCGTTCAG	152

组别	BMP2	Runx2	Col1a1	Bglap
成骨诱导组	1.10±0.10	0.93±0.12	1.10±0.10	1.06±0.05
实验组	2.53±0.42	1.68±0.04	1.82±0.11	2.01±0.36
t	5.798^＊＊	10.583^＊＊	8.519^＊＊	4.531^＊＊

组别	BMP2	Runx2	Col1a1	Bglap
成骨诱导组	1.10±0.10	0.93±0.12	1.10±0.10	1.06±0.05
实验组	2.53±0.42	1.68±0.04	1.82±0.11	2.01±0.36
t	5.798^＊＊	10.583^＊＊	8.519^＊＊	4.531^＊＊

组别	BV/TV	Tb. N/mm^-1	Tb. Th/mm
对照组	2.06±0.31	0.55±0.05	0.042±0.003
二甲双胍组	6.17±1.00	1.48±0.15	0.051±0.002
t	9.595^＊＊	14.461^＊＊	6.320^＊＊

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

Study of effects of metformin on osteogenic differentiation of mesenchymal stem cells and bone tissue regeneration in type I osteogenesis imperfecta mice

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