Near-infrared fluorescent probe-mediated high-throughput discovery of a natural β-galactosidase inhibitor for senescence-associated atherosclerosis therapy.

Atherosclerosis (AS), a disease associated with aging, significantly compromises the health and quality of life of patients. β-galactosidase (β-gal), a key cell senescence biomarker, shows significantly increased activity in senescent cells. Upregulated β-gal in senescent cells amplifies a proinflammatory senescence-associated secretory phenotype (SASP), driving cellular dysfunction, monocyte recruitment, foam cell formation, and extracellular matrix remodeling, thereby promoting atherogenesis. Inhibiting β-gal activity is a potential strategy for delaying AS and other aging-related diseases. Herein, we designed and synthesized a novel near-infrared fluorescent probe activated by β-gal (NFP-gal). This probe exhibited high sensitivity and selectivity with low cytotoxicity and biotoxicity under the tested conditions. We used this fluorescence-guided screening method to evaluate the β-gal inhibitory activity of 58 traditional Chinese medicines (TCMs) and identified Arnebia euchroma (AE) as a strong candidate. Through targeted separation and identification of complex components in AE using ultra-high-performance liquid chromatography with quadrupole orbitrap high-resolution mass spectrometry (UHPLC-Q-orbitrap-HRMS), we identified β,β-dimethyl-acryl-alkannin (DMAA) as a novel β-gal inhibitor. The pharmacodynamic effects of this compound were validated through in vitro cellular imaging using senescent cell and foam cell models, real-time in vivo fluorescence tracking in zebrafish models of AS, and histopathological analysis using apolipoprotein E-deficient (ApoE -/-) mice with high-fat diet (HFD)-induced AS. The findings confirmed the therapeutic potential of DMAA in mitigating senescence-associated vascular dysfunction. This study comprehensively evaluated the use of β-gal-based probes and natural compounds to suppress aging-related processes, highlighting their therapeutic implications for AS and other aging-associated diseases.