Prediction of microRNA–mRNA interaction typically relies on bioinformatic methods, but these methods only suggest the possibility of microRNA binding and may miss important interactions as well as falsely predict others. A major obstacle to the miRNA research has been the lack of experimental procedures for the identification of miRNA–mRNA interactions. Recently, a few studies have attempted to explore experimental methods to isolate and identify miRNA targets or miRNAs targeting a single gene. Here, we developed an more convenient experimental approach for the isolation and identification of miRNAs targeting a single gene by applying short biotinylated DNA anti-sense oligonucleotides mix to enhanced green fluorescent protein (EGFP) mRNA which was fused to target gene mRNA. This method does not require a design of different anti-sense oligonucleotides to any mRNA. This is a simple and an efficient method to potentially identify miRNAs targeting specific gene mRNA combined with chip screen.

Intersectin-2Long (ITSN2L) is a multi-domain protein participating in endocytosis and exocytosis. In this study, RABEP1 was identified as a novel ITSN2L interacting protein using a yeast two-hybrid screen from a human brain cDNA library and this interaction, specifically involving the ITSN2L CC domain and RABEP1 CC3 regions, was further confirmed by in vitro GST (glutathione-S-transferase) pull-down and in vivo co-immunoprecipitation assays. Corroboratively, we observed that these two proteins co-localize in the cytoplasm of mammalian cells. Furthermore, over-expression of ITSN2L promotes RABEP1 degradation and represses RABEP1-enhanced endosome aggregation, indicating that ITSN2L acts as a negative regulator of RABEP1. Finally, we showed that ITSN2L and RABEP1 play opposite roles in regulating endocytosis. Taken together, our results indicate that ITSN2L interacts with RABEP1 and stimulates its degradation in regulation of endocytosis.

Ischemia–reperfusion injury (IRI) is a prevalent pathological process in clinical settings characterized by complex pathogenesis involving the interplay of oxidative stress, inflammation, mitochondrial dysfunction, and diverse cell death pathways. Fundamentally, IRI manifests as a complication arising from reperfusion therapies aimed at restoring blood flow following ischemia. Despite the existence of various therapeutic strategies, the development of effective interventions for IRI remains a significant challenge. Rutin, a low-molecular-weight flavonoid glycoside ubiquitously present in vegetables, fruits, and herbal medicines, exhibits promising therapeutic potential due to its pleiotropic biological activities, including antioxidant, anti-inflammatory, and cytoprotective effects against cell death. This review systematically elucidates the molecular mechanisms underlying the protective effects of rutin against IRI and synthesizes evidence from preclinical studies regarding its diverse modes of action. However, the clinical application of rutin is currently hampered by its relatively low bioavailability. Future research should prioritize the development of innovative pharmaceutical formulations to enhance its bioavailability, thereby fully unlocking its clinical translational value.