基于AIE效应的多重刺激响应性聚合物纳米微球的制备及其细胞示踪应用

doi:10.6023/A19060226

化学学报 ›› 2019, Vol. 77 ›› Issue (10): 1036-1044.DOI: 10.6023/A19060226 上一篇下一篇

所属专题：分子探针、纳米生物学与生命分析化学

研究论文

基于AIE效应的多重刺激响应性聚合物纳米微球的制备及其细胞示踪应用

关晓琳^a^*(), 王林^a, 李志飞^a, 刘美娜^a, 王凯龙^a, 林斌^a, 杨学琴^a, 来守军^b, 雷自强^a

^a 西北师范大学生态环境相关高分子材料教育部重点实验室甘肃省高分子材料重点实验室化学化工学院兰州 730070
^b 兰州文理学院化工学院兰州 730000

投稿日期:2019-06-21 发布日期:2019-08-13
通讯作者: 关晓琳 E-mail:guanxiaolin@nwnu.edu.cn
基金资助:
项目受国家自然科学基金(21761032);项目受国家自然科学基金(51363019);生态环境相关高分子材料教育部重点实验室开放基金(KF-18-05)

Preparation of Multi-stimulus Responsive Polymer Nanospheres Based on AIE Effect and Its Cell Tracing Application

Guan, Xiaolin^a^*(), Wang, Lin^a, Li, Zhifei^a, Liu, Meina^a, Wang, Kailong^a, Lin, Bin^a, Yang, Xueqing^a, Lai, Shoujun^b, Lei, Ziqiang^a

^a Key Laboratory of Eco-Environment-Related Polymer Materials Ministry of Education, Key Laboratory of Polymer Materials Ministry of Gansu Province, College of Chemistry and Chemical Engineering, Northwest Normal University, Lanzhou 730070, China
^b School of Chemical Engineering, Lanzhou University of Arts and Science, Lanzhou 730000, China

Received:2019-06-21 Published:2019-08-13
Contact: Guan, Xiaolin E-mail:guanxiaolin@nwnu.edu.cn
Supported by:
Project supported by the National Natural Science Foundation of China(21761032);Project supported by the National Natural Science Foundation of China(51363019);the Key Laboratory of Ecological Environment Related Polymer Materials, Ministry of Education Open Fund(KF-18-05)

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1. 基于AIE效应的多重刺激响应性聚合物纳米微球的制备及其细胞示踪应用.PDF(632KB)

摘要/Abstract

基于AIE分子和智能响应性聚合物构筑的纳米材料,具有优良的AIE发光性能、环境刺激响应性和生物相容性,已在生命科学领域展现出诱人的应用前景. 本研究通过ATRP活性聚合方法, 以合成的TPE-BIB为引发剂, 引发具有多刺激响应特性的N-[2-(二乙氨基)-乙基]丙烯酰胺单体聚合, 成功制备具有温度/pH/CO₂三重响应性的两亲性聚合物: TPE-g-PDEAEAM, 并自组装形成约200 nm的纳米微球. 研究表明: 这种聚合物纳米粒子具有优良的水溶性、单分散性、稳定性及优异的AIE发光特性. 其相转变温度为60 ℃, 溶液荧光对环境温度、pH及CO₂均表现出快速敏感响应性能. 同时, 该纳米粒子表现出低细胞毒性, 能够有效示踪HeLa细胞增殖至11代以上, 有望作为一种活细胞荧光示踪探针材料.

关键词: AIE, 多重敏感响应, 聚合物纳米微球, 细胞示踪, 荧光成像

In recent years, fluorescent bioimaging technology has great advantages in the fields of life science research and medical diagnosis because of its advantages of fast and effective, high sensitivity, easy realization of multi-channel imaging and economic efficiency. Organic fluorescent dyes have been widely used as biological imaging reagents due to their excellent photoelectric properties, functional modification, adjustable optical properties, and good biocompatibility. However, conventional organic fluorescent molecules cause aggregation-caused quenching (ACQ) due to π-π stacking in the aggregated state, limiting their bioimaging applications in aggregated or high concentrations. Since the discovery of the unique luminescence phenomenon of aggregation-induced emission (AIE), the ACQ phenomenon of traditional fluorescent materials has been eliminated. Stimulating responsive polymer nanoparticles have been widely used in the life sciences due to their combination of nanoparticle and polymer advantages and their ability to respond intelligently with environmental changes. Therefore, nanomaterials with excellent aggregation-induced emission (AIE) property, environmental stimuli responsiveness and biocompatibility based on AIE molecules and smart responsive polymers have shown attractive application prospects in the life sciences. A kind of multi-responsive AIE-active polymer nanospheres, which were composed of tetraphenylethylene (TPE) and stimuli-responsive poly[N]-2-(diethylamino)-ethyl]acrylamide (PDEAEAM), were constructed in this study. Firstly, a multi-stimulation responsive monomer N-[2-(diethylamino)ethyl]acrylamide (DEAEAM) and TPE derivative tetraphenylethene-4-(12-hydroxydodecyl-2-methylpropionyl) (TPE-BIB) with propionyl bromide were synthesized, respectively, and a multi-stimuli-responsive amphiphilic polymer of tetraphenylethene-graft-poly[N-[2-(diethylamino)ethyl]acrylamide] (TPE-g-PDEAEAM) was then successfully synthesized by atom transfer radical polymerization (ATRP) using TPE-BIB as initiator. Lastly, polymer nanospheres TPE-g-PDEAEAM of approximately 200 nm were formed by a self-assembling pro-cess. The results of the performed experiments showed that the LCST of TPE-g-PDEAEAM in aqueous solution is about 60 ℃. Meanwhile, the luminescence change of TPE-g-PDEAEAM at different temperatures from 20 to 66 ℃ was observed. The fluorescence intensity of TPE-g-PDEAEAM firstly decreased with increasing temperature from 20 to 58 ℃, and the fluorescence intensity increased with increasing temperature from 58 to 66 ℃. The phase transfer of PDEAEAM in TPE-g-PDEAEAM may be the reason of luminescence change which may lead to the fluorescent temperature response. Moreover, the fluorescence intensity of TPE-g-PDEAEAM nanospheres in aqueous solution increased with increasing temperature pH. Besides, the fluorescence intensity of TPE-g-PDEAEAM decreased dramatically when the volume of CO₂ increased from 0.0 to 1.2 mL. Therefore, TPE-g-PDEAEAM was a new temperature and pH/CO₂ responsive materials and might be used as multi-functional smart fluorescent sensors. More importantly, the fluorescent signals were significantly strong in HeLa cells after cells were incubated with TPE-g-PDEAEAM for 24 h based on the characteristic of AIE fluorescence and low cytotoxicity. The resultant nanospheres were able to be internalized by the cancer cells and effectively track the HeLa cells for as long as 11 passages. So, the polymer nanomaterial is an ideal living cell fluorescent tracer probe, which is expected to be applied as biosensors, long-term cell traces and medical biomaterials.

Key words: AIE, multiple sensitive response, polymer nanospheres, cell tracing, fluorescence imaging

引用此文

关晓琳, 王林, 李志飞, 刘美娜, 王凯龙, 林斌, 杨学琴, 来守军, 雷自强. 基于AIE效应的多重刺激响应性聚合物纳米微球的制备及其细胞示踪应用[J]. 化学学报, 2019, 77(10): 1036-1044.

Guan, Xiaolin, Wang, Lin, Li, Zhifei, Liu, Meina, Wang, Kailong, Lin, Bin, Yang, Xueqing, Lai, Shoujun, Lei, Ziqiang. Preparation of Multi-stimulus Responsive Polymer Nanospheres Based on AIE Effect and Its Cell Tracing Application[J]. Acta Chimica Sinica, 2019, 77(10): 1036-1044.

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图/表 9

图1. 聚合物TPE-g-PDEAEAM的合成路线

Figure 1. Synthetic route of the polymer TPE-g-PDEAEAM

图2. (a)浓度为2 mg/mL的流体动力学尺寸(插图: 激光照射下TPE-g-PDEAEAM水溶液的丁铎尔效应照片, (b) TPE-g-PDEAEAM的SEM图像

Figure 2. (a) Hydrodynamic size with concentration of 2 mg/mL. Inset: Photograph of TPE-g-PDEAEAM solutions in water under irradiation of laser point. (b) SEM image of TPE-g-PDEAEAM polymer dots

图3. (a)不同浓度的TPE-g-PDEAEAM溶液的荧光光谱(激发波长为337 nm). (b)不同浓度的TPE-g-PDEAEAM溶液的荧光强度的相对变化及在紫外灯照射下(365 nm)的数码相片

Figure 3. (a) Fluorescence spectra of aqueous solutions of TPE-g-PDEAEAM at different concentrations (ex=337 nm). (b) Relative changes in fluorescence intensity of different concentrations of TPE-g-PDEAEAM aqueous solution and digital photographs under UV light (365 nm)

图4. (a)在水/THF混合溶剂中TPE-g-PDEAEAM溶液的荧光光谱(激发波长为337 nm). (b) TPE-g-PDEAEAM在水/THF混合溶剂荧光强度的相对变化及其在紫外灯下(365 nm)的数码相片

Figure 4. (a) Fluorescence spectra of TPE-g-PDEAEAM in a mixed solvent of water and THF (λex=337 nm). (b) The relative change in fluorescence intensity of TPE-g-PDEAEAMTPE in a water/THF mixed solvent and its digital photograph under UV light (365 nm)

图5. TPE-g-PDEAEAM在浓度为2 g/L的变温紫外. 插图: TPE-PDEAEAM溶液在25 ℃(左)和66 ℃(右)下在可见光的照片

Figure 5. TPE-g-PDEAEAM polymer quantum dots in a variable temperature ultraviolet (UV) concentration of 2 g/L. Inset: Photograph of visible light in TPE-PDEAEAM solution at 25 ℃ (left) and 65 ℃ (right)

图6. (a) TPE-g-PDEAEAM在温度从20 ℃升高到66 ℃的水溶液中的荧光光谱. [TPE-g-PDEAEAM]=2.0 g/L. λex=337 nm. 插图: TPE-g-PDEAEAM溶液在25 ℃(左)和65 ℃(右)下在可见光和紫外灯下的照片. (b)不同温度下TPE-g-PDEAEAM溶液中的相对荧光强度 (I/I0)的变化

Figure 6. (a) Fluorescent spectra of the TPE-g-PDEAEAM in aqueous solution with temperature increasing from 20 to 66 ℃. [TPE-g- PDEAEAM]=2.0 g/L. λex=337 nm. Inset: Photo of TPE-g-PDEAEAM solution under 25 ℃ (left) and 65 ℃ (right) under visible and UV light. (b) Change of relative fluorescent intensity (I/I0) of TPE-g-PDEAEAM in aqueous solution under different temperature

图7. (a) TPE-g-PDEAEAM在不同pH溶液中的荧光光谱. (b) 不同pH下TPE-g-PDEAEAM溶液中的相对荧光强度(I/I0)的变化. [TPE-g-PDEAEAM]=2.0 g/L

Figure 7. (a) Fluorescent spectra of the TPE-g-PDEAEAM in different pH solution. (b) Change of relative fluorescent intensity (I/I0) of TPE-g-PDEAEAM in aqueous solution under different pH. [TPE-g-PDEAEAM]=2.0 g/L

图8. (a) TPE-g-PDEAEAM在不同的CO2体积溶液的荧光光谱. 插图: 通入二氧化碳前后在紫外灯下(365 nm) 2 mg/mL的TPE-g- PDEAEAM水溶液的数码相片. (b) 相对荧光强度的线性(I/I0)与CO2体积的关系

Figure 8. (a) Fluorescence spectra of TPE-g-PDEAEAM in different CO2 volume solutions. Inset: Digital photo of a 2 mg/mL TPE-g- PDEAEAM aqueous solution under UV light (365 nm) before and after carbon dioxide. (b) The relationship between the linearity of relative fluorescence intensity (I/I0) and the volume of CO2

图9. (a)用不同浓度的TPE-g-PDEAEAM处理48 h后HeLa细胞毒性测试; (b) TPE-g-PDEAEAM染色HeLa细胞不同代数的共聚焦显微镜图像, 比例尺是25 μm

Figure 9. (a) Cell viabilities of HeLa cells treated with different concentrations of TPE-g-PDEAEAM for 48 h by MTT assay. (b) Confocal microscopy images of HeLa cells TPE-g-PDEAEAM staining (100 μg/mL) in different algebras. The scale bar is 25 μm

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基于AIE效应的多重刺激响应性聚合物纳米微球的制备及其细胞示踪应用

Preparation of Multi-stimulus Responsive Polymer Nanospheres Based on AIE Effect and Its Cell Tracing Application

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