Dual-Locked Probe with Activatable Sonoafterglow Luminescence for Precise Imaging of MET-Induced Liver Injury

Metformin (MET) is currently the first-line treatment for type 2 diabetes mellitus (T2DM). However, overdose and long-term use of MET may induce a serious liver injury. What's worse, diagnosis of MET-induced liver injury remains challenging in clinic. Although several probes have been reported for imaging MET-induced liver injury utilizing upregulated hepatic H₂S as a biomarker, they are still at risk of nonspecific activation in complex physiological environments and rely on light excitation with limited imaging depth. Herein, we rationally designed and developed a dual-locked probe, DPA-H₂S, for precise imaging of MET-induced liver injury by H₂S-activated sonoafterglow luminescence. DPA-H₂S is a small molecule consisting of a sonosensitizer protoporphyrin IX (PpIX) and an afterglow substrate that is dual-locked with a H₂S-responsive 2,4-dinitrobenzene group and a ¹O₂-responsive electron-rich double bond. When employing DPA-H₂S for imaging of MET-induced liver injury <i>in vivo</i>, since the PpIX moiety can produce ¹O₂ <i>in situ</i> at the liver site under focused ultrasound (US) irradiation, the two locks of DPA-H₂S can be specifically activated by the highly upregulated H₂S at the liver injury sites and the <i>in situ</i> generated ¹O₂, respectively. Thus, the sonoafterglow signal of DPA-H₂S is significantly turned on, enabling precise imaging of the MET-induced liver injury. <i>In vitro</i> results showed that, through H₂S-activated sonoafterglow luminescence, DPA-H₂S was capable of imaging H₂S with good sensitivity and high selectivity and realized deep tissue imaging (∼20 mm, signal-to-background ratio (SBR) = 3.4). Furthermore, we successfully applied DPA-H₂S for precise <i>in vivo</i> imaging of MET-induced liver injury. We anticipate that our dual-locked probe, DPA-H₂S, may serve as a promising tool in assisting the diagnosis of MET-induced liver injury in clinics and informing the clinical utilization of MET in the near future.