Intracellular imaging and therapeutic application of liver fibrosis via a microRNA-on self-stacking amplification...

Liver fibrosis is a major global health issue with limited treatment options. Activation of hepatic stellate cells (HSCs) induces collagen deposition, which is a core step in the progression of liver fibrosis. MicroRNA221-3p (miR221-3p) is a key profibrotic regulator of the activation of hepatic stellate cells. Sensitive and selective imaging and regulation of miR221-3p within complex cellular environments are essential for both diagnosis and therapy, yet remains challenging.

To address this, we engineered a miR-on Self-Assembly Cascade (SAC) amplification strategy for miR221-3p visualization and regulation. The system consists of a DNA initiator strand (Hc) and four hairpin strands (H1-H4). MiR221-3p binds to and opens Hc, triggering a self-sustained circular hairpin reporter reaction. The resulting replicas serve as feedback to drive subsequent cycles, achieving exponential signal amplification. This design enables highly sensitive and selective fluorescence imaging of miR221-3p in living cells. Additionally, as a functional demonstration, the SAC system also induced inactivation of HSCs. Mechanistic studies revealed that the SAC strategy disrupts the miR221-3p/Suppressor Of Cytokine Signaling 1 (SOCS1) axis, thereby suppressing Signal Transducer and Activator of Transcription 3 (STAT3) phosphorylation, thus alleviating liver fibrosis. In vivo studies further demonstrate the sustained administration of SAC significantly reduced extracellular matrix accumulation and ameliorated liver fibrosis in a CCl₄-induced mouse model.

Owing to its self-sustained signal amplification, the SAC strategy achieves both sensitive miR221-3p imaging and effective fibrosis suppression. This work provides a robust analytical platform for miRNA-based diagnostics and holds promise for precision medicine in fibrotic and miRNA-related disorders.