PHOTONIC NANOSENSORS FOR REACTIVE OXYGEN SPECIES AND OXIDATIVE STRESS IN DISEASE

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 中心，不一定是研究者的机构。 活性氧（ROS）是癌症和许多神经退行性疾病的重要潜在原因。然而，连接ROS致癌和这些病理条件的分子机制仍然没有得到充分的理解。这主要是因为我们目前对ROS影响的理解大多来自间接终点测量，如DNA，蛋白质损伤和脂质过氧化。这些测量仅评估ROS的累积效应，并提供有关此类效应的分子机制的有限信息。我们提出了新的基于荧光的光子纳米传感器能够直接和选择性检测活性氧的发展。我们将使用所提出的纳米传感器来监测细胞内ROS浓度的变化和分布直接在癌变过程中。通过将这些变化与来自平行生物测定的关键癌症生物标志物的调节相关联，我们希望进一步阐明个体ROS在致癌作用中的不同作用以及这种作用的分子机制。为了实现这些目标，我们已经合成了几个潜在的活性氧探针，通过引入不同的苯磺酰基到荧光素分子。与传统的ROS探针不同，这些苯磺酰基荧光素衍生物基于非氧化机制检测ROS，从而对不同的ROS产生选择性。通过改变苯磺酰基上取代基的类型和位置，可以调节荧光素衍生物的反应性。这些探针对不同活性氧的选择性是正在进行的研究的主题。本研究的结果将用于进一步辅助设计对单个ROS具有改进特异性的ROS探针。我们还制备了胺修饰的二氧化硅纳米粒子作为ROS探针的纳米平台。为了减小纳米颗粒的尺寸并产生具有更均匀尺寸的纳米颗粒，改变了几个参数。在概念验证实验中，我们成功地将异硫氰酸荧光素（FITC）掺入到胺改性的二氧化硅纳米颗粒上。证明了通过共价连接基于苯磺酰基荧光素的ROS探针获得功能性纳米传感器的可行性。我们计划使用这些初步结果来指导我们合成额外的ROS探针，并进行更广泛的选择性研究。我们还计划在不久的将来使用模型细胞系统来评估纳米传感器的性能。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Reactive oxygen species (ROS) has been implicated as an important underlying cause in cancer and many neurodegenerative diseases. However, the molecular mechanisms that connect ROS to carcinogenesis and these pathological conditions remain insufficiently understood. This is mainly because most of our current understanding of the effects of ROS has come from indirect endpoint measurements such as DNA, protein damage and lipid peroxidation. These measurements only assess the cumulative effects of ROS and provide limited information on the molecular mechanisms of such effects. We proposed the development of novel fluorescence-based photonic nanosensors capable of direct and selective detection of ROS. We will use the proposed nanosensors to monitor changes in intracellular ROS concentration and distribution directly during carcinogenesis. By correlating these changes to the regulation of key cancer biomarker from parallel bioassays, we hope to further elucidate the differential roles of individual ROS in carcinogenesis and the molecular mechanisms of such roles. To achieve these goals, we have synthesized several potential ROS probes by introducing various benzenesulfonyl groups onto the fluorescein molecule. Unlike traditional ROS probes, these benzenesulfonyl-fluorescein derivatives detect ROS based on non-oxidative mechanisms, rendering selectivity toward different ROS. By varying the type and position of the substituent on the benzenesulfonyl group, the reactivity of the fluorescein derivative can be tuned. The selectivity of these probes toward different ROS is the topic of an on-going study. The outcome of this study will be used in further assisting the design of ROS probes with improved specificity toward individual ROS. We have also prepared amine-modified silica nanoparticles as the nano-platform for the ROS probes. Several parameters were varied in an effort to reduce the size of the nanoparticles and produce nanoparticles with more uniform size. In a proof-of-concept experiment, we successfully incorporated fluorescein isothiocyanate (FITC) onto the amine-modified silica nanoparticles. The feasibility of obtaining functional nanosensors through covalent attachment of benzenesulfonyl-fluorescein-based ROS probes was demonstrated. We plan to use these preliminary results to guide our synthesis of additional ROS probes and conduct more extensive selectivity study. We also plan to evaluate the performance of the nanosensors using a model cell system in the near future.