Enzyme-Activated Molecular MRI for Specific Delineation of the Ischemic Penumbra in acute ischemic stroke.

Abstract

Ischemic stroke, the second leading cause of death and a primary source of severe disability in adults worldwide, is associated with high morbidity and mortality rates that correlate closely with the extent of neuronal damage. A critical strategy for improving patient outcomes is the early identification and rescue of the ischemic penumbra, a region at risk but potentially salvageable. However, accurately identifying and visualizing this penumbra poses a significant challenge in treatment and is crucial for predicting patient prognosis. Conventional magnetic resonance imaging (MRI) techniques often fail to delineate viable tissue effectively. Cleaved Caspase-3 (c-Casp3), a key executor of apoptosis, is specifically and highly expressed within the penumbra, making it an ideal molecular target for precise imaging. To address this challenge, we designed and constructed a multifunctional, enzyme-responsive MRI nanoprobe (FGAPT) for the molecular imaging of the ischemic penumbra. This probe capitalizes on the elevated expression of activated Caspase-3 during ischemic neuronal apoptosis, facilitating its specific accumulation and activation at the lesion site. As a result, there is a significant enhancement of the T1-weighted imaging signal, allowing for accurate delineation of the penumbra. This study establishes an imaging strategy for the precise identification of the ischemic penumbra, thereby presenting a paradigm for guiding individualized thrombolytic and neuroprotective interventions in ischemic stroke. The proposed methodology lays a robust imaging foundation for transitioning stroke management from a generalized therapeutic approach to personalized treatment optimization. STATEMENT OF SIGNIFICANCE: The precise delineation of the ischemic penumbra is critical for guiding therapy in acute ischemic stroke, yet remains a challenge for conventional imaging. To address this, we developed an enzyme-responsive magnetic resonance imaging (MRI) nanoprobe, FGAPT, for the molecular visualization of this salvageable tissue. This smart probe is engineered around a MRET mechanism, linking a superparamagnetic quencher (Fe₃O₄) and a paramagnetic enhancer (Gd-DOTA) with a peptide sequence specifically cleaved by activated Caspase-3-a key executor of apoptosis upregulated in the penumbra. Surface conjugation of a brain-targeting aptamer ensures blood-brain barrier penetration. In a rodent stroke model, the probe achieved high-contrast, specific T1-signal enhancement exclusively within the penumbra, as confirmed by spatial colocalization with histologically verified apoptotic cells. This Caspase-3-activated imaging strategy enables accurate differentiation between the infarct core and the ischemic penumbra. By directly visualizing a pivotal molecular determinant of cellular fate, our work provides a novel tool to advance stroke management from a rigid "time window" paradigm toward a precision "tissue window" approach.