Effect of Biaxial Tensile Strain on Expression of Osteogenic Specificity Markers of Rat Bone Marrow-derived Mesenchymal Stem Cells <i>in Vitro</i>_Journal of Biomedical Engineering

Authors：

ZHANGMengying ¹ , LIJinchuan ² , LIHuiming ¹ , WUGeng ¹ , CAOChengjian ¹ , WUWenchao ³ ,  LILiang ¹

1. Institute of Biomedical Engineering, West China College of Preclinical Medicine and Forensic Medicine, Sichuan University, Chengdu 610041, China;
2. College of Architecture and Environment, Sichuan University, Chengdu 610065, China;
3. Laboratory of Cardiovascular Diseases, West China Hospital, Sichuan University, Chengdu 610041, China;

Corresponding author：

LILiang, Email: lilianghx@163.com

Keywords：

rat bone marrow-derived mesenchymal stem cells; tensile strain; collagen type Ⅰ; Runt-related transcription factor 2; osteocalcin

DOI：

10.7507/1001-5515.20160084

Video：

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The purpose of this study was to investigate the effect of biaxial tensile strain on the osteogenic differentiation of rat bone marrow mesenchymal stem cells (rBMSCs) in vitro. The rBMSCs were isolated from tibia and femur of 4 weeks-old Sprague-Dawley (SD) rats. The rBMSCs were cultured in DMEM-LG complete culture medium and grew to subconfluence in the cell culture device for loading tensile strain. The biaxial tensile strain was applied to the rBMSCs for periods of 2, 4 and 6 hours every day, respectively, lasting 3 days. The amplitude of biaxial tensile strain applied to the rBMSCs were 1%, 2% and 5% respectively, at a frequency of 1 Hz. Unstrained rBMSCs were used as blank control (control group). The rBMSCs cultured with DMEM-LG complete culture medium containing 100 nmol/L β-Estradiol (E2) were used as positive control. The mRNA expression of alkaline phosphatase (ALP), collagen typeⅠ (ColⅠ), Runt-related transcription factor 2 (Runx2) and osteocalcin (OCN) was examined with real-time quantitative PCR and the protein expression of ALP, ColⅠ, Runx2 and OCN was detected with Western blot method. The results showed as follws: (1) The mRNA and protein expression of the ALP, ColⅠ, Runx2, OCN were significantly higher in rBMSCs of the E2 group than those in the control group (P＜0.05). (2) The mRNA and protein expression level of the ALP, Runx2 were higher markedly in the 1% tensile strain groups than those in the control group (P＜0.05), but lower than those in the E2 group (P＜0.05). (3) The mRNA and protein expression level of the ALP, ColⅠ, Runx2, OCN were significantly higher in the 2% tensile strain groups than those in the control group (P＜0.05), and the mRNA and protein expression level of ColⅠ and Runx2 in the group applied with 2% amplitude of tensile strain for 4 h/d was significantly higher than those in E2 group (P＜0.05). (4) The mRNA and protein expression level of the ALP, ColⅠ, Runx2 were significantly higher in the groups applied with 5% amplitude of tensile strain for 2 h/d or for 4 h/d than those in the control group (P＜0.05). In our study, E2 and mechanical stimulation played an important role in the regulation of differentiation of rBMSCs into osteoblasts, and the manner applied with the 2% amplitude of tensile strain for 4 h/d, lasting 3 days was an optimal stimulus for up-regulating the mRNA and protein expression of ALP, ColⅠ, Runx2, OCN of rBMSCs.

Citation： ZHANGMengying, LIJinchuan, LIHuiming, WUGeng, CAOChengjian, WUWenchao, LILiang. Effect of Biaxial Tensile Strain on Expression of Osteogenic Specificity Markers of Rat Bone Marrow-derived Mesenchymal Stem Cells in Vitro. Journal of Biomedical Engineering, 2016, 33(3): 499-505,519. doi: 10.7507/1001-5515.20160084 Copy

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5.	LI L, YAO X L, HE X L, et al. Role of mechanical strain and estrogen in modulating osteogenic differentiation of mesenchymal stem cells (MSCs) from normal and ovariectomized rats[J]. Cell Mol Biol (Noisy-le-grand), 2013, (Suppl 59):L1889-L1893.
6.	严中琴, 杨刚,崔燎,等.电刺激促骨髓间充质干细胞向心肌样细胞分化[J].生物医学工程学杂志,2013,30(3):556-561.
7.	况薇, 唐敏,何学令,等.周期性拉伸应变对大鼠骨髓间充质干细胞向心肌样细胞分化的影响[J].生物医学工程学杂志,2014,31(3):596-600.
8.	DEAL C, GIDEON J. Recombinant human PTH 1-34(Forteo):an anabolic drug for osteoporosis[J]. Cleve Clin J Med, 2003, 70(7):585-586, 589-590, 592-594.
9.	Drugs for prevention and treatment of postmenopausal osteoporosis[J]. Treat Guidel Med Lett, 2005, 3(38):69-74.
10.	FAN Jinzhu, YANG Liu, MENG Guo-lin, et al. Estrogen improves the proliferation and differentiation of hBMSCs derived from postmenopausal osteoporosis through notch signaling pathway[J]. Mol Cell Biochem, 2014, 392(1-2):85-93.
11.	JAALOUK D E, LAMMERDING J. Mechanotransduction gone awry[J]. Nat Rev Mol Cell Biol, 2009, 10(1):63-73.
12.	BOERCKEL J D, DUPONT K M, KOLAMBKAR Y M, et al. In vivo model for evaluating the effects of mechanical stimulation on tissue-engineered bone repair[J]. J Biomech Eng, 2009, 131(8):084502.
13.	PALOMARES K T, GLEASON R E, MASON Z D, et al. Mechanical stimulation alters tissue differentiation and molecular expression during bone healing[J]. J Orthop Res, 2009, 27(9):1123-1132.
14.	YAMAZAKI S, MIZUMOTO T, NASU A, et al. Regulation of osteogenetic differentiation of mesenchymal stem cells by two axial rotational culture[J]. J Artif Organs, 2011, 14(4):310-317.
15.	LUU Y K, OZCIVICI E, CAPILLA E, et al. Development of diet-induced fatty liver disease in the aging mouse is suppressed by brief daily exposure to low-magnitude mechanical signals[J]. Int J Obes (Lond), 2010, 34(2):401-405.
16.	OZCIVICI E, LUU Y K, ADLER B, et al. Mechanical signals as anabolic agents in bone[J]. Nat Rev Rheumatol, 2010, 6(1):50-59.
17.	YANG Xingmei, GONG Ping, LIN Yunfeng, et al. Cyclic tensile stretch modulates osteogenic differentiation of adipose-derived stem cells via the BMP-2 pathway[J]. Arch Med Sci, 2010, 6(2):152-159.
18.	VICO L, HINSENKAMP M, JONES D, et al. Osteobiology, strain, and microgravity. Part Ⅱ:studies at the tissue level[J]. Calcif Tissue Int, 2001, 68(1):1-10.
19.	LI Runguang, LIANG Liang, DOU Yonggang, et al. Mechanical strain regulates osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells[J]. Biomed Res Int, 2015, 2015:873251.

1. 李良,陈孟诗,邓力,等.力学刺激对大鼠成骨细胞Integrins α_2、β_1、β_3表达和细胞增殖、合成功能的影响[J].生物医学工程学杂志,2003,20(2):187-192.
2. EHRLICH P J, LANYON L E. Mechanical strain and bone cell function:a review[J]. Osteoporos Int, 2002, 13(9):688-700.
3. KHAYAT G, ROSENZWEIG D H, QUINN T M. Low frequency mechanical stimulation inhibits adipogenic differentiation of C3H10T1/2 mesenchymal stem cells[J]. Differentiation, 2012, 83(4):179-184.
4. QI M C, HU J, ZOU S J, et al. Mechanical strain induces osteogenic differentiation:Cbfa1 and Ets-1 expression in stretched rat mesenchymal stem cells[J]. Int J Oral Maxillofac Surg, 2008, 37(5):453-458.
5. LI L, YAO X L, HE X L, et al. Role of mechanical strain and estrogen in modulating osteogenic differentiation of mesenchymal stem cells (MSCs) from normal and ovariectomized rats[J]. Cell Mol Biol (Noisy-le-grand), 2013, (Suppl 59):L1889-L1893.
6. 严中琴, 杨刚,崔燎,等.电刺激促骨髓间充质干细胞向心肌样细胞分化[J].生物医学工程学杂志,2013,30(3):556-561.
7. 况薇, 唐敏,何学令,等.周期性拉伸应变对大鼠骨髓间充质干细胞向心肌样细胞分化的影响[J].生物医学工程学杂志,2014,31(3):596-600.
8. DEAL C, GIDEON J. Recombinant human PTH 1-34(Forteo):an anabolic drug for osteoporosis[J]. Cleve Clin J Med, 2003, 70(7):585-586, 589-590, 592-594.
9. Drugs for prevention and treatment of postmenopausal osteoporosis[J]. Treat Guidel Med Lett, 2005, 3(38):69-74.
10. FAN Jinzhu, YANG Liu, MENG Guo-lin, et al. Estrogen improves the proliferation and differentiation of hBMSCs derived from postmenopausal osteoporosis through notch signaling pathway[J]. Mol Cell Biochem, 2014, 392(1-2):85-93.
11. JAALOUK D E, LAMMERDING J. Mechanotransduction gone awry[J]. Nat Rev Mol Cell Biol, 2009, 10(1):63-73.
12. BOERCKEL J D, DUPONT K M, KOLAMBKAR Y M, et al. In vivo model for evaluating the effects of mechanical stimulation on tissue-engineered bone repair[J]. J Biomech Eng, 2009, 131(8):084502.
13. PALOMARES K T, GLEASON R E, MASON Z D, et al. Mechanical stimulation alters tissue differentiation and molecular expression during bone healing[J]. J Orthop Res, 2009, 27(9):1123-1132.
14. YAMAZAKI S, MIZUMOTO T, NASU A, et al. Regulation of osteogenetic differentiation of mesenchymal stem cells by two axial rotational culture[J]. J Artif Organs, 2011, 14(4):310-317.
15. LUU Y K, OZCIVICI E, CAPILLA E, et al. Development of diet-induced fatty liver disease in the aging mouse is suppressed by brief daily exposure to low-magnitude mechanical signals[J]. Int J Obes (Lond), 2010, 34(2):401-405.
16. OZCIVICI E, LUU Y K, ADLER B, et al. Mechanical signals as anabolic agents in bone[J]. Nat Rev Rheumatol, 2010, 6(1):50-59.
17. YANG Xingmei, GONG Ping, LIN Yunfeng, et al. Cyclic tensile stretch modulates osteogenic differentiation of adipose-derived stem cells via the BMP-2 pathway[J]. Arch Med Sci, 2010, 6(2):152-159.
18. VICO L, HINSENKAMP M, JONES D, et al. Osteobiology, strain, and microgravity. Part Ⅱ:studies at the tissue level[J]. Calcif Tissue Int, 2001, 68(1):1-10.
19. LI Runguang, LIANG Liang, DOU Yonggang, et al. Mechanical strain regulates osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells[J]. Biomed Res Int, 2015, 2015:873251.

Journal of Biomedical Engineering

Effect of Biaxial Tensile Strain on Expression of Osteogenic Specificity Markers of Rat Bone Marrow-derived Mesenchymal Stem Cells in Vitro

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