目的 探究Noggin-siRNA的胞内转染对间充质干细胞(MSCs)成骨分化的影响。方法 利用转染试剂将Noggin-siRNA转导入MSCs中,细胞流式检测转染后MSCs的细胞表型,MTT检测转染后MSCs的增殖活性并绘制细胞生长曲线,碱性磷酸酶(ALP)染色、实时荧光定量PCR(QRT-PCR)、Western Blotting检测转染后的MSCs的成骨分化。结果 通过与对照组比较,实验组细胞的细胞表型和生长曲线没有明显改变,ALP染色阳性细胞比率及成骨相关基因与蛋白表达量明显提高。结论 Noggin-siRNA的胞内转染对MSCs成骨分化的促进作用是安全的、可靠的、高效的。
Abstract
Objective To investigate whether delivery of noggin-siRNA has a strong osteogenic effect on MSCs.Methods Noggin-siRNA was transferred into MSC with a transfection reagent. The cell phenotype and proliferation of MSCs were tested by flow cytometry and MTT assay, respectively. The osteogenesis of MSCs was tested by ALP staining, QRT-PCR and Western Blotting, respectively.Results Compared with the control group, the cell phenotype and growth curve of MSC did not change significantly while the proportion of ALP positive cells and the expression of osteogenesis related genes and proteins were significantly increased.Conclusions The delivery of Noggin-siRNA can enhance osteogenesis of MSCs safely, reliably and efficiently.
关键词
Noggin-siRNA /
MSCs /
胞内转染 /
成骨诱导
Key words
Noggin-siRNA /
MSCs /
delivery /
osteogenesis
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参考文献
[1] Fan J,Park H,Tan S,et al. Enhanced osteogenesis of adipose derived stem cells with Noggin suppression and delivery of BMP-2[J]. PLoS One,2013,8(8):e72474.
[2] Nkenke E,Weisbach V,Winckler E,et al. Morbidity of harvesting of bone grafts from the iliac crest for preprosthetic augmentation procedures: a prospective study[J]. Int J Oral Maxillofac Surg,2004,33(2):157-163.
[3] Schultheiss J,Seebach C,Henrich D,et al. Mesenchymal stem cell (MSC) and endothelial progenitor cell (EPC) growth and adhesion in six different bone graft substitutes[J]. Eur J Trauma Emerg Surg,2011,37(6):635-644.
[4] Hao W,Dong J,Jiang M,et al. Enhanced bone formation in large segmental radial defects by combining adipose-derived stem cells expressing bone morphogenetic protein 2 with nHA/RHLC/PLA scaffold[J]. Int Orthop,2010,34(8):1341-1349.
[5] Cui Z K,Fan J,Kim S,et al. Deliveryof siRNA via cationic Sterosomes to enhance osteogenic differentiation of mesenchymal stem cells[J]. J Control Release,2015,217:42-52.
[6] Lu C H,Chang Y H,Lin S Y,et al. Recent progresses in gene delivery-based bone tissue engineering[J]. Biotechnol Adv,2013,31(8):1695-1706.
[7] Cheema S K,Chen E,Shea L D,et al. Regulation and guidance of cell behavior for tissue regeneration via the siRNA mechanism[J]. Wound Repair Regen,2007,15(3):286-295.
[8] Choi B,Cui Z K,Kim S,et al. Glutamine-chitosan modified calcium phosphate nanoparticles for efficient siRNA delivery and osteogenic differentiation[J]. HHS Public Access,2015,3(31):6448-6455.
[9] Kilgore M L,Morrisey M A,Becker D J,et al. Health care expenditures associated with skeletal fractures among medicare beneficiaries,1999-2005[J]. J Bone Miner Res,2009,24(12):2050-2055.
[10] Evans C H,Ghivizzani S C,Robbins P D. Progress and prospects: genetic treatments for disorders of bones and joints[J]. Gene Ther,2009,16(8):944-952.
[11] Evans C H,Ghivizzani S C,Robbins P D. Orthopedic gene therapy in 2008[J]. Mol Ther,2009,17(2):231-244.
[12] Zhang Y,Satterlee A,Huang L. In vivo gene delivery by nonviral vectors:overcoming hurdles?[J]. Mol Ther,2012,20(7):1298-1304.
[13] Vannucci L,Lai M,Chiuppesi F,et al. Viral vectors: a look back and ahead on gene transfer technology[J]. New Microbiol,2013,36(1):1-22.
[14] 张铭杰,张庆文,何 伟,等. 成人骨髓间充质干细胞体外培养、鉴定与成骨诱导[J]. 中国组织工程研究,2013,17(45):7947-7953.
[15] Yan J,Zhang C,Zhao Y,et al. Non-viral oligonucleotide antimiR-138 delivery to mesenchymal stem cell sheets and the effect on osteogenesis[J]. Biomaterials,2014,35(27):7734-7749.
[16] Yau W W,Rujitanaroj P O,Lam L,et al. Directing stem cell fate by controlled RNA interference[J]. Biomaterials,2012,33(9):2608-2628.
[17] Poggio P, Sainger R, Branchetti E, et al. Noggin attenuates the osteogenic activation of human valve interstitial cells in aortic valve sclerosis[J]. Cardiovascular Res,2013,98(3):402-410.
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