Effects of non-dextran coated superparamagnetic iron oxide nanoparticles on proliferation of bone marrow mesenchymal stem cells

doi:10.3969/j.issn.2095-4344.2014.16.011

Abstract

Abstract:

BACKGROUND: Currently, the research about effect of non-dextran coated superparamagnetic iron oxide nanoparticles on cell proliferation and cytotoxicity is relatively much less.

OBJECTIVE: To evaluate the effects of 0, 25, 50, 75, 100 mg/L non-dextran coated superparamagnetic iron oxide nanoparticles on the proliferation and cytotoxicity of rat bone marrow mesenchymal stem cells.

METHODS: Culture media containing 0, 25, 50, 75, 100 mg/L non-dextran coated superparamagnetic iron oxide nanoparticles were prepared for culture of bone marrow mesenchymal stem cells. After 24 hours of culture, the cells were confirmed using Prussian blue staining, and cell counting was detected using cell counting kit-8. Meanwhile, lactate dehydrogenase activity in the supernatant and intracellular superoxide dismutase activity were detected.

RESULTS AND CONCLUSION: Loading of non-dextran coated superparamagnetic iron oxide nanoparticles in BMSCs was confirmed by Prussian blue staining. The percentage of cells labeled with non-dextran coated superparamagnetic iron oxide nanoparticles was up to 100% when the cells were incubated with a non-dextran coated superparamagnetic iron oxide nanoparticle solution of 50 mg/L and above, but 25 mg/L was insufficient to label all of the cells. Furthermore, as the concentration of non-dextran coated superparamagnetic iron oxide nanoparticles decreased, the cell proliferation rate decreased gradually. The 25 mg/L group had a minimum cell proliferation rate, but the 25 and 50 mg/L groups showed no statistically significant difference (P > 0.05). Therefore, 50 mg/L is considered as the appropriate concentration of non-dextran coated superparamagnetic iron oxide nanoparticles, under which, the labeling efficiency is higher and the cytotoxicity is lower.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

全文链接：

Key words: biocompatible materials, magnetite nanoparticles, stem cells, cell proliferation, lactate dehydrogenases

CLC Number:

R318

Chen Peng, Zhang Jie, Rong Dong-ming, Han Zhong-yu, Yuan Si-jie, Tian Jing. Effects of non-dextran coated superparamagnetic iron oxide nanoparticles on proliferation of bone marrow mesenchymal stem cells[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(16): 2526-2531.

Figures/Tables 3

2.1 大鼠骨髓间充质干细胞的鉴定与中国科学院苏州纳米技术与纳米仿生研究所合作成功研制出生物性能稳定的、尺寸可控的、单分散的超顺磁性氧化铁纳米颗粒[11]，表面经修饰后其可在生理细胞培养液中保持稳定，颗粒直径约12 nm(图1A)。该颗粒有很强的饱和磁矩，在低温下(6 K)有磁滞，但在室温下表现超顺磁性。采用全骨髓培养法培养法获得第3代的大鼠骨髓间充质干细胞，细胞形态为长梭形及多角形，呈漩涡状排列(图1B)。流失细胞检测结果表明该细胞高表达CD29(99.44%)及CD90(96.40%) (图2)。标记细胞行普鲁士蓝染色后，可见细胞内超顺磁性氧化铁纳米颗粒被染为蓝色，散在分布于细胞核周围(图1C)。实验中观察到，50 mg/L及以上浓度的超顺磁性氧化铁纳米颗粒培养基孵育的细胞，超顺磁性氧化铁纳米颗粒的标记率基本可达100%。25 mg/L组超顺磁性氧化铁纳米颗粒培养基孵育的细胞仍有少量未被染色，说明其未与超顺磁性氧化铁纳米颗粒充分结合。 2.2 CCK-8法测定不同质量浓度无葡聚糖包被超顺磁性氧化铁纳米颗粒标记的骨髓间充质干细胞增殖活性在将无葡聚糖包被超顺磁性氧化铁纳米颗粒标记的骨髓间充质干细胞用于体内实验之前，需寻找到一个合适的浓度，在这个浓度下，骨髓间充质干细胞既能获得最大的标记率又不会影响其增殖及分化。CCK-8实验计算各质量浓度超顺磁性氧化铁纳米颗粒对细胞生长的抑制率，将其绘制成柱状图(图3)，结果显示：随着超顺磁性氧化铁纳米颗粒质量浓度的增高，样本吸光度值逐渐减小，超顺磁性氧化铁纳米颗粒对细胞对细胞的抑制率逐渐增大，25 mg/L组对细胞的抑制率最小，50 mg/L组虽然对细胞生长的抑制率较25 mg/L组为高，但二者在统计学上无明显差别(P=0.076)；25 mg/L组对细胞的标记率尚无法达到100%，而50 mg/L组可对细胞100%标记。"

References

[1] Ouyang HW,Cao T,Zou XH,et al.Mesenchymal stem cell sheets revitalize nonviable dense grafts: implications for repair of large-bone and tendon defects. Transplantation. 2006;82(2):170-174.
[2] He X,Chen Y,Wang K,et al.Selective separation of proteins with pH-dependent magnetic nanoadsorbents. Nanotechnology.2007;18:365604.
[3] Buyukhatipoglu K,Jo W,Sun W,et al.The role of printing parameters and scaffold biopolymer properties in the efficacy of a new hybrid nano-bioprinting system. Biofabrication. 2009; 1(3):035003.
[4] Leem G,Zhang S,Jamison AC,et al.Light-induced covalent immobilization of monolayers of magnetic nanoparticles on hydrogen-terminated silicon.ACS Appl Mater Interfaces.2010; 2(10):2789-2796.
[5] Kanczler JM,Sura HS,Magnay J,et al.Controlled differentiation of human bone marrow stromal cells using magnetic nanoparticle technology.Tissue Eng Part A.2010; 16(10):3241-3250.
[6] Huang DM,Hsiao JK,Chen YC,et al.The promotion of human mesenchymal stem cell proliferation by superparamagnetic iron oxide nanoparticles.Biomaterials. 2009;30(22):3645-3651.
[7] Lellouche JP,Periman N,Joseph A,et al.New magnetically responsive polydicarbazole magnetite nanoparticles.Chem Commun.2004;(5):560-561.
[8] 梁文英,莫润阳,郑敏.SPIO磁性标记细胞技术研究进展[J].青海师范大学学报:自然科学版,2010,26(1):21-24.
[9] Rath SN,Nooeaid P,Arkudas A,et al.Adipose- and bone marrow-derived mesenchymal stem cells display different osteogenic differentiation patterns in 3D bioactive glass-based scaffolds.J Tissue Eng Regen Med.2013. doi: 10.1002/term.1849.
[10] Zeng G,Wang G,Guan F,et al.Human amniotic membrane-derived mesenchymal stem cells labeled with superparamagnetic iron oxide nanoparticles: the effect on neuron-like differentiation in vitro.Mol Cell Biochem.2011; 357(1-2): 331-341.
[11] Jiang J,Gu HW,Shao H,et al.Bifunctional Fe3O4–Ag Heterodimer Nanoparticles for Two-Photon Fluorescence Imaging and Magnetic Manipulation.Adv Mater.2008;20(23):4403-4407.
[12] Yan SG,Zhang J,Tu Q,et al.Transcription factor and bone marrow stromal cells in osseointegration of dental implants.Eur Cell Mater.2013;26:263-270; discussion 270-271.
[13] Li M,Li S,Yu L,et al.Bone mesenchymal stem cells contributed to the neointimal formation after arterial injury.PLoS One.2013;8(12):e82743. doi: 10.1371/journal.pone.0082743.
[14] Ritfeld GJ,Rauck BM,Novosat TL,et al.The effect of a polyurethane-based reverse thermal gel on bone marrow stromal cell transplant survival and spinal cord repair. Biomaterials. 2014;35(6):1924-1931.
[15] Zheng L,Chu J,Shi Y,et al.Bone marrow-derived stem cells ameliorate hepatic fibrosis by down-regulating interleukin-17.Cell Biosci.2013;3(1):46.
[16] Hauger O,Frost EE,vanHeeswijk R,et al.MR evaluation of the glo-merular homing ofmagnetically labeled mesenchymal stem cells in a ratmodel ofnephropathy.Radiology.2006; 238(1):200-210.
[17] Sun S,Chen G,Xu M,et al.Differentiation and migration of bone marrow mesenchymal stem cells transplanted through the spleen in rats with portal hypertension.PLoS One. 2013;8(12):e83523.
[18] Wu W,Chen B,Cheng J,et al.Biocompatibility of Fe3O4/DNR magnetic nanoparticles in the treatment of hematologic malignancies.Int J Nanomedicine.2010;5:1079-1084.
[19] Wang J,Chen Y,Chen B,et al.Pharmacokinetic parameters and tissue distribution of magnetic Fe3O4 nanoparticles in mice.Int J Nanomedicine.2010;21:861-866.
[20] Daldrup-Link HE,Rudelius M,Oostendorp RAJ,et al.Targeting of hematopoietic progenitor cells with MR contrast agents. Radiology.2003;228(3):760-767.
[21] Arbab AS,Yocum GT,Kalish H,et al.Efficient magnetic cell labeling with protamine surface complexed to ferumoxides for cellular MRI.Blood.2004;104(4):1217-1223.
[22] Bulte JW,Douglas T,Witwer B,et al. Magnetodendrimers allow endosomal magnetic labeling and in vivo tracking of stem cells.Nat Biotechnol.2001;19(11):1141-1147.
[23] Sosnovik DE,Nahrendorf M,Weissleder R.Magnetic nanoparticles for MR imaging: agents, techniques and cardiovascular applications. Basic Res Cardiol. 2008; 103(2): 122-130.
[24] Barrett T,Kobayashi H,Brechbiel M,et al.Macromolecular MRI contrast agents for imaging tumor angiogenesis.Eur J Radiol. 2006;60(3):353-366.
[25] 唐大宗,夏黎明.对比剂的发展史[J].放射学实践,2007, 22(6): 631-633.
[26] van Buul GM,Farrell E,Kops N,et al.Ferumoxides protamine sulfate is more effective than ferucarbotran for cell labeling: implications for clinically applicable cell tracking using MRI.Contrast Media Mol Imaging. 2009;4(5):230-236．
[27] Jing XH,Yang L,Duan XJ,et al.In vivo MR imaging tracking of magnetic iron oxide nanoparticle labeled, engineered, autologous bone marrow mesenchymal stem cells following intra-articular injection.J Bone Spine.2008;75:432-438．
[28] Wang Y,Wang Y,Wang L,et al. Preparation and evaluation of magnetic nanoparticles for cell labeling.J Nanosci Nanotechnol. 2011;11(5):3749-3756.
[29] Yuan F,Dong P,Wang X,et al.Toxicological effects of cigarette smoke on Ana-1 macrophages in vitro.Exp Toxicol Pathol. 2013;65(7-8):1011-1018.
[30] El-Bahr SM.Curcumin regulates gene expression of insulin like growth factor, B-cell CLL/lymphoma 2 and antioxidant enzymes in streptozotocin induced diabetic rats. BMC Complement Altern Med.2013;13(1):368.