共担载阿霉素/血红蛋白树枝状介孔硅球的制备及其生物性能研究

doi:10.15541/jim20180076

摘要/Abstract

摘要：

实体瘤中普遍存在乏氧现象, 是导致肿瘤对非手术治疗手段抗拒性增加, 降低药物疗效的重要因素。针对这一问题, 本研究采用简单的两相界面法制备了一种小尺寸(65 nm)、单分散、生物稳定性良好的可共载抗癌药物盐酸阿霉素(DOX)和载氧蛋白血红蛋白(Hb)的树枝状介孔硅纳米颗粒(DMSNs)。利用扫描电子显微镜(SEM)、透射电子显微镜(TEM)、动态光散射仪(DLS)和氮气吸附-脱附仪等对材料进行表征。结果表明, 合成的DMSNs纳米颗粒粒径均一、分散性良好, 具有较大的比表面积(654.52 m²/g)和孔容(1.26 cm³/g)以及两套孔道结构(直径2.7 nm和5.4~6.8 nm)。更重要的是, 树枝状介孔层的孔径仅需改变三乙醇胺(TEA)的用量即可调节。药物释放、流式细胞术、激光共聚焦以及细胞毒性等相关实验结果表明, DMSNs可同时装载DOX与Hb, 且具有较高的药物释放能力(75.6%)和持久的释放性能(48 h)。载入血红蛋白后, 其IC₅₀为20.6 μg/mL, 能够有效提高抗癌药物DOX的细胞致死率。因此, 这种小尺寸的树枝状介孔硅球在药物传输和肿瘤治疗方面具有潜在的应用价值。

关键词: 乏氧, 小尺寸, 树枝状介孔硅纳米颗粒, 药物传输

Abstract:

Tumor hypoxia is a ubiquitous factor to cause the therapeutic resistance of drug or other treatments. To solve this problem, a facile biophase stratification approach was developed to fabricate dendritic mesoporous silica nanoparticles (DMSNs) with small particle size of 65 nm, high dispersity and excellent biostability through simply controlling the oil-water interfacial reaction. The resultant DMSNs were characterized by various techniques such as scanning electron microscope (SEM), transmission electron microscope (TEM), dynamic light scattering (DLS), and N₂ adsorption-desorption analysis, etc. It is verified that DMSNs possess two types of pore size (2.7 nm and 5.4-6.8 nm), high specific surface (654.52 m²/g) and pore volume (1.26 cm³/g), which are capable to co-load DOX and Hb, simultaneously. Moreover, the mesopore size in the dendritic mesoporous layer could be tuned by changing the amounts of triethanolamine (TEA). The testing results from drug release, flow cytometry histograms, confocal laser scanning microscopy, and cell cytotoxicity demonstrate that DMSNs possess high efficiency of drug release (75.6%), durable releasing period (48 h) and significantly enhanced cell lethality (IC₅₀= 20.6 μg/mL). These data demonstrate that such kind of dentritic mesoporous silica naoparticles has great potentials in drug delivery and tumor chemotherapy.

Key words: tumor hypoxia, small size, dendritic mesoporous silica nanoparticles, drug delivery

中图分类号:

TQ174

潘珊, 李永生, 施剑林. 共担载阿霉素/血红蛋白树枝状介孔硅球的制备及其生物性能研究[J]. 无机材料学报, 2018, 33(10): 1097-1102.

PAN Shan, LI Yong-Sheng, SHI Jian-Lin. Facile Synthesis of Dendritic Mesoporous Silica Nanoparticles for Co-loading of Doxorubicin and Hemoglobin[J]. Journal of Inorganic Materials, 2018, 33(10): 1097-1102.

图/表 8

图1 DMSNs纳米颗粒的制备示意图

Fig. 1 Schematic illustration for the preparation of DMSNs

图2 MSNs (a)~(b)、DMSNs-0 (c)~(d)、DMSNs-0.3 (e)~(f)、DMSNs-0.6 (g)~(h) 的不同放大倍数的透射电镜照片

Fig. 2 TEM images of MSNs (a, b), DMSNs-0 (c, d), DMSNs-0.3 (e, f), and DMSNs-0.6 (g, h) at different magnification

图3 MSNs、DMSNs-0、DMSNs-0.3和DMSNs-0.6动态光散射粒径分布图

Fig. 3 DLS size distribution curves of MSNs, DMSNs-0, DMSNs- 0.3, and DMSNs-0.6

图4 MSNs\DMSNs-0\DMSNs-0.3和DMSNs-0.6氮气吸附-脱附曲线(a)及孔径分布图(b)

Fig. 4 N2 adsorption-desorption isotherms (a) and corresponding BJH pore size distributions (b) of MSNs, DMSNs-0, DMSNs-0.3, and DMSNs-0.6

表1 MSNs、DMSNs-0、DMSNs-0.3和DMSNs-0.6的孔结构参数比较

Table 1 Pore structural parameters of the synthesized MSNs, DMSNs-0, DMSNs-0.3, and DMSNs-0.6

Sample	Specific surface area/(m²·g^-1)	Pore diameter/nm	Pore volume/ (cm³·g^-1)
MSNs	525.81	2.7	1.02
DMSNs-0	342.86	1.7	0.70
DMSNs-0.3	469.16	2.7 & 5.4	0.83
DMSNs-0.6	654.52	2.7 & 6.8	1.26

图5 DMSNs的稳定性测试(a)、Hb/DOX@DMSN在不同pH (pH=5.4和pH=7.4)PBS溶液里的释放曲线(b)

Fig. 5 Biostability testing of DMSNs (a) and cumulative drug release of Hb/DOX@DMSN in PBS with pH=7.4 and pH=5.4 (b)

图6 Hb/DOX@DMSN与SMMC-7721细胞共培养不同时间(1、4、8和24 h)的流式细胞术结果(a)和激光共聚焦照片(b)

Fig. 6 Flow cytometry histograms (a) and CLSM images (b) of SMMC-7721 cells after being incubated with Hb/DOX@DMSN for 1 h, 4 h, 8 h, and 24 h

图7 SMMC-7721细胞与不同DOX浓度的Hb/DOX@DMSNs、DOX@DMSNs和Free DOX共培养24 h的细胞毒性结果

Fig. 7 Cell viability of SMMC-7721 cells incubated with Hb/DOX@DMSNs, DOX@DMSNs and free DOX with different concentrations of DOX for 24 h

参考文献 23

[1]	ZHU W W, DONG Z L, FU T T, et al. Modulation of hypoxia in solid tumor microenvironment with MnO2 nanoparticles to enhance photodynamic therapy. Advanced Functional Materials, 2016, 26(30): 5490-5498.
[2]	CHEN Q, FENG L Z, LIU J J, et al. Intelligent albumin-MnO2 nanoparticles as pH-/H2O2-responsive dissociable nanocarriers to modulate tumor hypoxia for effective combination therapy. Advanced Materials, 2016, 28(33): 7129-7136.
[3]	SONG X J, FENG L Z, LIANG C, et al. Ultrasound triggered tumor oxygenation with oxygen-shuttle nanoperfluorocarbon to overcome hypoxia-associated resistance in cancer therapies. Nano Letters, 2016, 16(10): 6145-6153.
[4]	LUO Z Y, ZHENG M B, ZHAO P F, et al. Self-monitoring artificial red cells with sufficient oxygen supply for enhanced photodynamic therapy. Scientific Reports, 2016, 6(1): 1-11.
[5]	TIAN H, LUO Z, LIU L,et al. Cancer cell membrane-biomimetic oxygen nanocarrier for breaking hypoxia-induced chemoresistance. Advanced Functional Materials, 2017, 27(38): 1703197-1-7.
[6]	CHENG Y, CHENG H, JIANG C, et al. Perfluorocarbon nanoparticles enhance reactive oxygen levels and tumour growth inhibition in photodynamic therapy. Nature Communications, 2015, 6(1): 8785-8793.
[7]	BU H X, XU X, CHEN J M, et al. Synthesis of a hemoglobin- conjugated triblock copolymer for oxygen carrying and specific recognition of cancer cells. RSC Advances, 2017, 7(76): 48166-48175.
[8]	LIU Y H, LIU R T.Spectroscope and molecular model identify the behavior of doxorubicin-SPION binding to bovine hemoglobin.International Journal of Biological Macromolecules, 2015, 79: 564-569.
[9]	HUANG M X, LIU L, WANG S G, et al. Dendritic mesoporous silica nanospheres synthesized by a novel dual-templating micelle system for the preparation of functional nanomaterials. Langmuir, 2017, 33(2): 519-526.
[10]	SHEN D K, YANG J P, LI X M, et al. Biphase stratification approach to three-dimensional dendritic biodegradable mesoporous silica nanospheres. Nano Letters, 2014, 14(2): 923-932.
[11]	KNEŽEVIĆ N Ž, DURAND J O. Large pore mesoporous silica nanomaterials for application in delivery of biomolecules.Nanoscale, 2015, 7(6): 2199-2209.
[12]	SHIN H S, HWANG Y K, HUH S.Facile preparation of ultra-large pore mesoporous silica nanoparticles and their application to the encapsulation of large guest molecules.ACS Applied Materials & Interfaces, 2014, 6(3): 1740-1746.
[13]	XU C, YU M H, NOONAN O, et al. Core-cone structured monodispersed mesoporous silica nanoparticles with ultra-large cavity for protein delivery. Small, 2015, 11(44): 5949-5955.
[14]	DU X, SHI B, LIANG J, et al. Developing functionalized dendrimer- like silica nanoparticles with hierarchical pores as advanced delivery nanocarriers. Advanced Materials, 2013, 25(41): 5981-5985.
[15]	OSAKI F, KANAMORI T, SANDO S, et al. A quantum dot conjugated sugar ball and its cellular uptake. on the size effects of endocytosis in the subviral region. Journal of the American Chemical Society, 2004, 126(21): 6520-6521.
[16]	LU F, WU S H, HUNG Y, et al. Size effect on cell uptake in well-suspended, uniform mesoporous silica nanoparticles. Small, 2009, 5(12): 1408-1413.
[17]	PAN L M, LIU J N, HE Q J, et al. MSN-mediated sequential vascular- to-cell nuclear-targeted drug delivery for efficient tumor regression. Advanced Materials, 2014, 26(39): 6742-6748.
[18]	PAN L M, LIU J N, HE Q J, et al. Overcoming multidrug resistance of cancer cells by direct intranuclear drug delivery using TAT-conjugated mesoporous silica nanoparticles. Biomaterials, 2013, 34(11): 2719-2730.
[19]	PAN L M, HE Q J, LIU J N, et al. Nuclear-targeted drug delivery of TAT peptide-conjugated monodisperse mesoporous silica nanoparticles. Journal of the American Chemical Society, 2012, 134(13): 5722-5725.
[20]	LI N, NIU D C, JIANG Y, et al. Morphology evolution and spatially selective functionalization of hierarchically porous silica nanospheres for improved multidrug delivery. Chemistry of Materials, 2017, 29(24): 10377-10385.
[21]	CHEN Y, CHEN H, GUO L, et al. Hollow/rattle-type mesoporous nanostructures by a structural difference-based selective etching strategy. ACS Nano, 2009, 4(1): 529-539.
[22]	TU J, BOYLE A L, FRIEDRICH H, et al. Mesoporous silica nanoparticles with large pores for the encapsulation and release of proteins. ACS Applied Materials & Interfaces, 2016, 8(47): 32211-32219.
[23]	SINGH R K, KIM T H, MAHAPATRA C, et al. Preparation of self-activated fluorescence mesoporous silica hollow nanoellipsoids for theranostics. Langmuir, 2015, 31(41): 11344-11352.