基于稀土铕离子荧光标记的磷酸钙纳米材料体内分布与代谢研究

doi:10.15541/jim20240504

无机材料学报 ›› 2025, Vol. 40 ›› Issue (7): 754-764.DOI: 10.15541/jim20240504

基于稀土铕离子荧光标记的磷酸钙纳米材料体内分布与代谢研究

汤新丽¹(), 丁自友¹, 陈俊锐¹, 赵刚², 韩颖超¹()

1.武汉理工大学材料复合新技术国家重点实验室, 武汉430070
2.华中科技大学同济医学院协和医院, 急诊外科, 武汉 430022

收稿日期:2024-12-03 修回日期:2025-01-17 出版日期:2025-07-20 网络出版日期:2025-02-13
通讯作者: 韩颖超, 教授. E-mail: hanyingchao@whut.edu.cn
作者简介:汤新丽(1999-), 女, 硕士研究生. E-mail: tangxinli9@163.com
基金资助:
国家自然科学基金(51672206);材料复合新技术国家重点实验室开放基金(2023-KF-9)

In vivo Distribution and Metabolism of Calcium Phosphate Nanomaterials Based on Fluorescent Labeling with Rare Earth Europium Ions

TANG Xinli¹(), DING Ziyou¹, CHEN Junrui¹, ZHAO Gang², HAN Yingchao¹()

1. State Key Laboratory of Advanced Technology for Materials Synthesis and Processing, Wuhan University of Technology, Wuhan 430070, China
2. Department of Emergency Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China

Received:2024-12-03 Revised:2025-01-17 Published:2025-07-20 Online:2025-02-13
Contact: HAN Yingchao, professor. E-mail: hanyingchao@whut.edu.cn
About author:TANG Xinli (1999-), female, Master candidate. E-mail: tangxinli9@163.com
Supported by:
National Natural Science Foundation of China(51672206);State Key Laboratory of Advanced Technology for Materials Synthesis and Processing(2023-KF-9)

摘要/Abstract

摘要：

纳米磷酸钙(nCaP)在药物递送、生物成像、抗菌、促成骨等纳米医学领域具有潜在的应用前景, 但其体内分布与代谢规律尚未完全明确, 有待深入研究。本研究采用稀土铕离子荧光标记法, 以荷瘤裸鼠为模型, 探究了两种尺寸nCaP(纳米点NDs: (2.53±0.63) nm; 纳米颗粒NPs: (107.76±25.37) nm×(17.66±1.63) nm)在肝、脾、肺、肾、肿瘤中的分布与代谢。结果显示, 裸鼠尾静脉注射200 μL质量浓度为1.5 mg/mL的两种nCaP溶液, 4 h后CaP NPs主要分布在肝脏和脾脏中, 分别占比65.70%和29.32%, 在肺脏中占比3.83%, 而在肾脏和肿瘤中仅占比0.84%和0.32%, 表明大尺寸CaP NPs更容易被网状内皮(RES)系统中的吞噬细胞捕获。与CaP NPs相比, CaP NDs在肝脏、脾脏和肺脏中的积聚显著减少了89.40%、87.00%和88.89%, 而在肾脏和肿瘤中的蓄积分别提升了3.67倍和3.06倍, 表明尺寸减小有利于CaP NDs被肾小球滤出并增强其肿瘤靶向能力。CaP NDs在肝、脾、肺中的清除率(CLz)分别是CaP NPs的6.60倍、4.14倍和2.40倍, 是肾脏的42.29%, 表明尺寸减小有助于CaP NDs被肝脏、脾脏和肺脏内的吞噬细胞迅速代谢, 但存在肾小管的重吸收作用。在肿瘤中CaP NDs的CLz值比CaP NPs降低了91.9%, 表明小尺寸CaP NDs具有显著增强的肿瘤靶向和滞留能力。进一步对荷瘤裸鼠初步建立了包含尺寸因素的nCaP生理药代动力学(PBPK)模型, CaP NDs和CaP NPs在肿瘤部位分布的预测拟合度(R²)分别达到0.925和0.827。本研究揭示了nCaP的体内分布与代谢规律, 为其医学应用提供了支撑。

关键词: 纳米磷酸钙, 尺寸效应, 荧光标记, 组织分布, 生理药代动力学模型

Abstract:

Nano-calcium phosphate (nCaP) has potential applications in nanomedicine fields such as drug delivery, bioimaging, antibacterial treatment, and bone formation promotion. However, its distribution and metabolic patterns within the body are not yet fully understood and require further in-depth research. This study employs a rare earth europium ion fluorescence labeling method and uses tumor-bearing mice as a model to investigate the distribution and metabolism of two sizes of nCaP (nanodots NDs: (2.53±0.63) nm; nanoparticles NPs: (107.76±25.37) nm×(17.66±1.63) nm) in the liver, spleen, lung, kidney, and tumor tissue. The results showed that after tail vein injection of 200 μL with a mass concentration of 1.5 mg/mL nCaP into tumor-bearing nude rats for 4 h, CaP NPs were primarily distributed in the liver and spleen, accounting for 65.70% and 29.32%, respectively, with 3.83% in the lung, while only 0.84% and 0.32% in the kidney and tumor. This suggests that larger CaP NPs are more easily captured by phagocytes within the reticuloendothelial system (RES). In contrast, compared to CaP NPs, accumulation of CaP NDs in the liver, spleen, and lung decreased significantly by 89.40%, 87.00%, and 88.89%, respectively, while their accumulation in the kidney and tumor increased by 3.67 and 3.06 times. This indicates that smaller particle size facilitates CaP NDs in glomerular filtration for urinary excretion and enhances their tumor-targeting capability. The clearance rates (CLz) of CaP NDs in the liver, spleen, and lung were 6.60, 4.14, and 2.40 times higher than that of CaP NPs, respectively, and 42.29% in the kidney. This indicates that reduced size of CaP NDs facilitates rapid metabolism by phagocytes in the liver, spleen, and lung but also results in reabsorption in the renal tubules. In tumor, the CLz of CaP NDs decreased by 91.9%, much smaller than that of CaP NPs, suggesting that the smaller CaP NDs exhibit significantly enhanced tumor targeting and retention capability. In the meantime, a physiologically based pharmacokinetic (PBPK) model incorporating particle size factors was preliminarily established for tumor-bearing mice to simulate the distribution of nano-calcium phosphate. The model's predictive fit (R²) for CaP NDs and CaP NPs in tumor sites reached 0.925 and 0.827, respectively. This study provides promising support for understanding in vivo distribution and metabolic patterns of nCaP and applying potential in medicine.

Key words: nano-calcium phosphate, size effect, fluorescent labeling, tissue distribution, physiologically based pharmacokinetic model

中图分类号:

TQ174

汤新丽, 丁自友, 陈俊锐, 赵刚, 韩颖超. 基于稀土铕离子荧光标记的磷酸钙纳米材料体内分布与代谢研究[J]. 无机材料学报, 2025, 40(7): 754-764.

TANG Xinli, DING Ziyou, CHEN Junrui, ZHAO Gang, HAN Yingchao. In vivo Distribution and Metabolism of Calcium Phosphate Nanomaterials Based on Fluorescent Labeling with Rare Earth Europium Ions[J]. Journal of Inorganic Materials, 2025, 40(7): 754-764.

图/表 8

图1 荷瘤裸鼠的PBPK模型

Fig. 1 PBPK model of tumor-bearing mice PCs indicate phagocytic cells; TCs indicate tumor cells

图2 CaP NDs和CaP NPs的表征

Fig. 2 Characterizations of CaP NDs and CaP NPs (a-d) TEM images of CaP NDs (a), Eu:CaP NDs (b), CaP NPs (c), and Eu:CaP NPs (d); (e, f) Particle size statistical distributions of CaP NDs (e) and Eu:CaP NDs (f); (g-j) Length (g, i) and width (h, j) of CaP NPs (g, h) and Eu:CaP NPs (i, j); (k) DLS particle size distribution; (l) XRD patterns; (m) FT-IR spectra; (n) Europium ion release curves of Eu:CaP NDs and Eu:CaP NPs

表1 Eu:CaP NDs和Eu:CaP NPs中元素含量及摩尔比

Table 1 Elemental content and molar ratio of Eu:CaP NDs and Eu:CaP NPs

Sample	Value	Ca/(mg·L^-1)	P/(mg·L^-1)	Eu/(mg·L^-1)	(Eu/(Ca+Eu))/%	(Ca+Eu)/P
Eu:CaP NDs	Actual	8.72	4.53	2.82	7.86	1.61
Eu:CaP NDs	Theoretical	—	—	—	8.00	1.67
Eu:CaP NPs	Actual	22.50	11.85	7.23	7.82	1.59
Eu:CaP NPs	Theoretical	—	—	—	8.00	1.67

图3 CaP NDs和CaP NPs的生物安全性评价

Fig. 3 Biosafety evaluation of CaP NDs and CaP NPs (a) Cell viability; (b) Hemolysis rate; (c, d) Blood biochemistry analysis (ALT indicates alanine transaminase, AST indicates aspartate aminotransferase, CR indicates creatinine, and BUN indicates blood urea nitrogen); (e) H&E staining images of heart, liver, spleen, lung, and kidney organs in different groups with scale bar indicating 100 μm; (f) CLSM images of BMSC co-cultured with CaP NPs and CaP NDs for 24 h, respectively. Red: CaP NPs or CaP NDs fluorescence image excited at 458 nm. Blue: Cell nuclei stained with DAPI. Scale bar in (f) is 50 μm. Colorful figures are available on website

图4 注射不同时间后CaP NDs和CaP NPs在不同组织中的分布

Fig. 4 Biodistributions of CaP NDs and CaP NPs in different tissues after injection for different durations Colorful figures are available on website

图5 CaP NDs和CaP NPs在不同组织中的代谢

Fig. 5 CaP NDs and CaP NPs metabolic patterns in different tissues (a-e) Biodistributions of CaP NDs and CaP NPs in liver (a), spleen (b), lung (c), kidney (d), and tumor (e); (f) Clearance rate of CaP NDs and CaP NPs in different tissues

表2 CaP NDs和CaP NPs荷瘤小鼠各脏器和肿瘤部位的主要药动学参数

Table 2 Pharmacokinetic parameters of CaP NDs and CaP NPs in organ and tumor site of tumor-bearing mice

Sample	Parameter	Liver	Spleen	Lung	Kidney	Tumor
CaP NDs	AUC(0-t)/(mg·h·L^-1)	157.78	71.35	19.34	70.61	20.31
CaP NDs	CLz/(L·h^-1·kg^-1)	0.033	0.091	0.235	0.144	0.146
CaP NPs	AUC(0-t)/(mg·h·L^-1)	1347.46	578.05	66.74	19.85	6.07
CaP NPs	CLz/(L·h^-1·kg^-1)	0.005	0.022	0.098	0.341	1.796

图6 肿瘤部位PBPK模型的建立与优化

Fig. 6 Establishment and optimization of PBPK model for tumor bearing sites (a, b) PBPK model of CaP NDs (a) and CaP NPs (b) in the tumor; (c, d) Linear regression results between logarithmically transformed measured and simulated concentrations of CaP NDs (c) and CaP NPs (d) with R2 indicating the correlation coefficient; (e) Prediction of the distribution of nCaP with different sizes in tumor

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基于稀土铕离子荧光标记的磷酸钙纳米材料体内分布与代谢研究

In vivo Distribution and Metabolism of Calcium Phosphate Nanomaterials Based on Fluorescent Labeling with Rare Earth Europium Ions

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