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

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200nm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.

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 process. 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.