硫化氢信号分子比例计量荧光探针的构筑与造影应用

As the third gaseous transmitter after nitric oxide (NO) and carbon monoxide (CO),hydrogen sulphide has been proposed to be associated with neurotransmission,singal transduction and regulation of cardiovascular functions, and the related research to understand the biological roles of hydrogen sulphide are attracting considerable attention, which requires the in situ quantitive real-time information of hydrogen sulphide in complicated living systems.Due to the autofluorescence, unknown sensor concentration, and the environment-dependence of fluorescence, the current "turn-on" fluorescent sensors for hydrogen sulphide displaying the hydrogen sulphide-induced emssion enahncement are disable to offer the quantitive temporal/spatial information of hydrogen sulphide in live cells or organism. The ratiometric hydrogen sulphide sensors of hydrogen sulphide-induced emission or excitation shift was proposed to be developed in this proposal to realize the real-time ratiometric imaging of inctracellualr or in vivo hydrogen sulphide homeostasis and offer the quantitive hydrogen sulphide information, profiting from the internal caliberation effect via determining the dual emission ratio between two excitation or emission bands. With the normal ratiometric sensing mechanisms of internal charge transfer (ICT) and F?rster resonance energy transfer (FRET), a ratiometric sensor design strategy via incorporating the H2S sensitive functional group into the ICT or FRET sensor molecules was proposed, and the ratiometric sensing for hydrogen sulphide can be realized by the hydrogen sulphide-triggered reaction of the sensitive group, which alters the ICT or FRET and results in the hydrogen sulphide-induced emission or excitation shift.Among these sensors, those of near infrared（NIR）emission are expected to realize the in vivo hydrogen suiphide imaging in mice.The accomplishement of this project will provides the reliable tool for the quantitive imaging of intracellular/in vivo hydrogen sulphide, which will promote the development of bioinorganic chemistry of hydrogen sulphide as well as the techniques for the diagnose and treatment of related diseases.

硫化氢是继NO和CO之后的第三个气体信号分子，参与神经信号传递、信号转导及心血管功能调节等过程,而这些热门研究工作需要硫化氢的实时原位定量信息。由于荧光的环境依赖性、探针浓度未知和自发荧光干扰等因素，利用现有的增强型硫化氢荧光探针造影无法获得细胞及活体中硫化氢的定量信息。本项目提出通过构建具有硫化氢诱导发射或激发谱带移动的比例计量型荧光探针，利用其双带比例的内校正效应实现硫化氢在细胞或活体水平的实时比例成像，获得硫化氢的实时定量信息。项目提出构建分子内电荷转移(ICT)或荧光共振能量转移(FRET)机制的探针分子，通过硫化氢触发探针中敏感基团的反应诱导分子内ICT和FRET 效应的改变实现对硫化氢的比例响应。构建的近红外比例探针则可望用于裸鼠活体的硫化氢比例成像。项目的实施有望提供硫化氢定量跟踪造影的工具，不仅有利于促进硫化氢生物无机化学的发展，而且可推动对相关疾病诊疗技术的进步。

硫化氢是继NO和CO之后发现的第三个气体信号分子，参与神经信号传递、信号转导及心血管功能调节等过程。对这些过程的阐明需要硫化氢的实时原位定量信息。由于分子荧光的环境依赖性、探针浓度波动和自发荧光干扰等因素，现有增强型硫化氢荧光探针成像无法获得细胞及活体中硫化氢的定量信息。同时由于探针响应时间在20分钟到数小时间，也难以实现硫化氢的实时原位成像。本项目提出通过构建具有硫化氢诱导发射或激发谱带移动的比例计量型荧光探针，利用其双带比例的内校正效应克服增强型探针的不足，实现硫化氢在细胞或活体水平的实时比例成像，获得硫化氢的定量信息；进一步对初步获得的硫化氢比例荧光探通过结构优化提高其硫化氢响应速度和程度，以实现对硫化氢的实时响应和成像。项目一方面基于ICT和FRET机制构建了多个比例计量型硫化氢荧光探针，如基于ICT机制的CouMC系列(CouMC, CM-N’nCm, CM-NCn等)和BDPMC系列比例计量型硫化氢荧光探针，同时还获得了基于FRET机制的硫化氢比例探针NBD-CMC。利用这些探针克服了强度增强型探针难以进行定量检测和造影的缺点，实现溶液对H2S的比例计量响应，同时通过结构优化，获得了多个具有快速响应性能的优秀探针，并实现了特定的亚细胞分布，实现了活细胞内硫化氢的实时比例成像。其中CouMC是第一例具有线粒体靶向功能的比例计量型硫化氢荧光探针，同时也是目前对硫化氢响应最快的有机小分子探针之一，实现了线粒体内外源性硫化氢的比例荧光实时成像。另一方面项目还获得了具有NIR发射性能的比例型硫化氢荧光探针CyBz-1，实现了裸鼠皮下硫化氢的比例荧光成像，初步建立了哺乳动物活体H2S 造影模型。项目的实施提供了生命体系特别是细胞、细胞器以及模式小动物中硫化氢定量实时跟踪成像工具，不仅有利于促进硫化氢生物无机化学的发展，有利于推动对相关疾病诊疗技术的发展。