Liang Wenxuan, Chen Defu, Guan Honghua, Park Hyeon-Cheol, Li Kaiyan, Li Ang, Li Ming-Jun, Gannot Israel, Li Xingde
School of Biomedical Engineering, Suzhou Institute for Advanced Research, University of Science and Technology of China, Suzhou, Jiangsu 215000, China; School of Physical Sciences, University of Science and Technology of China, Hefei, Anhui 230026, China.
Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland 21205, United States.
ACS Photonics. 2022 Dec 21;9(12):4017-4029. doi: 10.1021/acsphotonics.2c01493. Epub 2022 Dec 9.
NADH intensity and fluorescence lifetime characteristics have proved valuable intrinsic biomarkers for profiling the cellular metabolic status of living biological tissues. To fully leverage the potential of NADH fluorescence lifetime imaging microscopy (FLIM) in (pre)clinical studies and translational applications, a compact and flexible endomicroscopic embodiment is essential. Herein we present our newly developed two-photon fluorescence (2PF) lifetime imaging endomicroscope (2p-FLeM) that features an about 2 mm diameter, subcellular resolution, and excellent emission photon utilization efficiency and can extract NADH lifetime parameters of living tissues and organs reliably using a safe excitation power (~30 mW) and moderate pixel dwelling time (≤10 s). In vivo experiments showed that the 2p-FLeM system was capable of tracking NADH lifetime dynamics of cultured cancer cells and subcutaneous mouse tumor models subject to induced apoptosis, and of a functioning mouse kidney undergoing acute ischemia-reperfusion perturbation. The complementary structural and metabolic information afforded by the 2p-FLeM system promises functional histological imaging of label-free internal organs in vivo and in situ for practical clinical diagnosis and therapeutics applications.
NADH强度和荧光寿命特征已被证明是用于描绘活生物组织细胞代谢状态的有价值的内在生物标志物。为了在（临床前）研究和转化应用中充分发挥NADH荧光寿命成像显微镜（FLIM）的潜力，紧凑且灵活的内镜实施方案至关重要。在此，我们展示了我们新开发的双光子荧光（2PF）寿命成像内镜（2p-FLeM），其特点是直径约2毫米、具有亚细胞分辨率、出色的发射光子利用效率，并且能够使用安全的激发功率（约30毫瓦）和适度的像素驻留时间（≤10秒）可靠地提取活组织和器官的NADH寿命参数。体内实验表明，2p-FLeM系统能够追踪培养的癌细胞和皮下小鼠肿瘤模型在诱导凋亡时的NADH寿命动态，以及经历急性缺血再灌注扰动的正常运作的小鼠肾脏的NADH寿命动态。2p-FLeM系统提供的互补结构和代谢信息有望实现体内和原位对无标记内部器官的功能组织学成像，用于实际临床诊断和治疗应用。
