Advances in small-molecule fluorescent probes for the study of apoptosis.

Apoptosis, as type I cell death, is an active death process strictly controlled by multiple genes, and plays a significant role in regulating various activities. Mounting research indicates that the unique modality of cell apoptosis is directly or indirectly related to different diseases including cancer, autoimmune diseases, viral diseases, neurodegenerative diseases, etc. However, the underlying mechanisms of cell apoptosis are complicated and not fully clarified yet, possibly due to the lack of effective chemical tools for the nondestructive and real-time visualization of apoptosis in complex biological systems. In the past 15 years, various small-molecule fluorescent probes (SMFPs) for imaging apoptosis in vitro and in vivo have attracted broad interest in related disease diagnostics and therapeutics. In this review, we aim to highlight the recent developments of SMFPs based on enzyme activity, plasma membranes, reactive oxygen species, reactive sulfur species, microenvironments and others during cell apoptosis. In particular, we generalize the mechanisms commonly used to design SMFPs for studying apoptosis. In addition, we discuss the limitations of reported probes, and emphasize the potential challenges and prospects in the future. We believe that this review will provide a comprehensive summary and challenging direction for the development of SMFPs in apoptosis related fields.

A new mitochondria-targeted fluorescent probe for exogenous and endogenous superoxide anion imaging in living cells and pneumonia tissue.

As a precursor of all reactive oxygen species (ROS), superoxide anions play an important role in organisms. However, excessive superoxide anions can cause various diseases. Thus, it is highly urgent to develop efficient tools for in situ superoxide anion detection. In this work, a novel boric acid-based, mitochondria-targeted fluorescent probe Mito-YX for superoxide anion detection was designed by regulating its intramolecular charge transfer (ICT) effect. The probe exhibited turn-on fluorescence enhancement within 4 min of reaction with the superoxide anion. In addition, Mito-YX also exhibited high selectivity and a low detection limit down to 0.24 µM with good mitochondrial targeting characteristics, which provided a necessary basis for in vivo detection of superoxide anions. What is more, Mito-YX was successfully applied for the in situ monitoring of superoxide anions in living MCF-7 cells, RAW 264.7 cells and a mouse model of lung inflammation stimulated by LPS. This work provided an important and promising tool for rapid in situ diagnosis and research of the progression of pneumonia.

A mitochondria-targeted fluorescent sensor for imaging endogenous peroxynitrite changes in acute lung injury.

Acute lung injury (ALI) is a serious pulmonary inflammatory disease resulting from excessive reactive oxygen species (ROS) which could cause the damage of the alveolar epithelial cells and capillary endothelial cells. Peroxynitrite, as one of short-lived reactive oxygen species, is closely related to the process of ALI. Thus, it is important to monitor the fluctuation of peroxynitrite in living system for understanding the process of ALI. Herein, the novel mitochondria-targeted fluorescent probe BHMT was designed to respond to peroxynitrite and pH with distinct fluorescence properties respectively. The absorption spectrum of the probe BHMT exhibited a notable red shift as the pH value declined from 8.8 to 2.6. Upon reaction with peroxynitrite, BHMT had a significant increase of fluorescence intensity (63-fold) with maintaining a detection limit of only 43.7 nM. Furthermore, BHMT could detect the levels of endogenous peroxynitrite and image the intracellular pH in ratiometric channels utilizing cell imaging. In addition, BHMT was successfully applied to revealing the relationship between the peroxynitrite and the extent of ALI. Thus, these results indicated the probe BHMT could be a potential tool for diagnosing the early stage of ALI and revealed the peroxynitrite was likely to be a crucial therapeutic target in ALI treatment.

Single-Cell RNA Sequencing of Human Corpus Cavernosum Reveals Cellular Heterogeneity Landscapes in Erectile Dysfunction.

Purpose: To assess the diverse cell populations of human corpus cavernosum in patients with severe erectile dysfunction (ED) at the single-cell level. Methods: Penile tissues collected from three patients were subjected to single-cell RNA sequencing using the BD Rhapsody™ platform. Common bioinformatics tools were used to analyze cellular heterogeneity and gene expression profiles from generated raw data, including the packages Seurat, Monocle, and CellPhoneDB. Results: Disease-related heterogeneity of cell types was determined in the cavernous tissue such as endothelial cells (ECs), smooth muscle cells, fibroblasts, and immune cells. Reclustering analysis of ECs identified an arteriole ECs subcluster and another one with gene signatures of fibroblasts. The proportion of fibroblasts was higher than the other cell populations and had the most significant cellular heterogeneity, in which a distinct subcluster co-expressed endothelial markers. The transition trajectory of differentiation from smooth muscle cells into fibroblasts was depicted using the pseudotime analysis, suggesting that the expansion of corpus cavernosum is possibly compromised as a result of fibrosis. Cell-cell communications among ECs, smooth muscle cells, fibroblasts, and macrophages were robust, which indicated that inflammation may also have a crucial role in the development of ED. Conclusions: Our study has demonstrated a comprehensive single-cell atlas of cellular components in human corpus cavernosum of ED, providing in-depth insights into the pathogenesis. Future research is warranted to explore disease-specific alterations for individualized treatment of ED.