Facile Access to Far-Red Fluorescent Probes with Through-Space Charge-Transfer Effects for In Vivo Two-Photon Microscopy of the Mouse Cerebrovascular System.

Much effort has been devoted to the generation of fluorescent probes by synthetic approaches. In this study, we developed a facile strategy to construct far-red fluorescent probes based on through-space charge transfer within complexes of acceptors and donors and their "twist+twist" interactions. Owing to their rare two-photon excitation property, the complexes could be used for in vivo imaging of the mouse cerebrovascular system.

Birth-and-death evolution of ribonuclease 9 genes in Cetartiodactyla.

RNase9 plays a reproductive function and has been recognized as an important member of the ribonuclease (RNase) A superfamily, a gene family that is widely used as a model for molecular evolutionary studies. Here, we identified 178 RNase9 genes from 95 Cetartiodactyla species that represent all four lineages and 21 families of this clade. Unexpectedly, RNase9 experienced an evolutionary scenario of "birth and death" in Ruminantia, and expression analyses showed that duplicated RNase9A and RNase9B genes are expressed in reproductive tissues (epididymis, vas deferens or prostate). This expression pattern combined with the estimate that these genes duplicated during the middle Eocene, a time when Ruminantia become a successful lineage, suggests that the RNase9 gene duplication might have been advantageous for promoting sperm motility and male fertility as an adaptation to climate seasonality changes of this period. In contrast, all RNase9 genes were lost in the Cetacean lineage, which might be associated with their high levels of prostatic lesions and lower reproductive rates as adaptations to a fully aquatic environment and a balance to the demands of ocean resources. This study reveals a complex and intriguing evolutionary history and functional divergence for RNase9 in Cetartiodactyla, providing new insights into the evolution of the RNaseA superfamily and molecular mechanisms for organismal adaptations to the environment.

Long-term monitoring of intravital biological processes using fluorescent protein-assisted NIR-II imaging.

High spatial resolution, low background, and deep tissue penetration have made near-infrared II (NIR-II) fluorescence imaging one of the most critical tools for in vivo observation and measurement. However, the relatively short retention time and potential toxicity of synthetic NIR-II fluorophores limit their long-term application. Here, we report the use of infrared fluorescent proteins (iRFPs) as in vitro and in vivo NIR-II probes permitting prolonged continuous imaging (up to 15 months). As a representative example, iRFP713 is knocked into the mouse genome to generate a transgenic model to allow temporal and/or spatial expression control of the probe. To demonstrate its feasibility in a genuine diagnostic context, we adopt two liver regeneration models and successfully track the process for a week. The performance and monitoring efficacy are comparable to those of µCT and superior to those of indocyanine green dye. We are also able to effectively observe the pancreas, despite its deep location, under both physiological and pathological conditions. These results indicate that the iRFP-assisted NIR-II fluorescence system is suitable for monitoring various tissues and in vivo biological processes, providing a powerful noninvasive long-term imaging platform.