Cascaded bio-responsive delivery of eNOS gene and ZNF₅₈₀ gene to collaboratively treat hindlimb ischemia <i>via</i> pro-angiogenesis and anti-inflammation

Abstract

Gene therapy is a promising strategy for treating ischemic disease by solving the dual dilemma of ischemia and inflammation. However, its development remains limited by inefficient gene transfection. Hence, we propose a "dual genes + all-adaptive carrier" idea. We have innovatively co-delivered eNOS gene and the ZNF₅₈₀ gene encoding its transcription factor to enhance the efficiency of eNOS expression. The overexpressed ZNF₅₈₀ protein significantly promotes angiogenesis via regulating the transcription of multiple genes. This implies a potential synergistic effect of eNOS and ZNF₅₈₀ genes in anti-ischemic therapy. Additionally, we have designed an all-adaptive gene carrier with cascaded bio-responsive functions based on the characteristic bio-signals of the ischemic site (including extracellular excessive matrix metalloproteinase-2, the endo/lysosomal pH gradient and high cytoplasmic glutathione level). This carrier can sequentially overcome transfection bottlenecks and achieve high transfection. Excitingly, this cascaded bio-responsive delivery strategy remarkably enhanced blood perfusion, accelerated angiogenesis and alleviated inflammation in critical limb ischemia (CLI) mice, which was attributed to the combined effects of pro-angiogenic ZNF₅₈₀ expression and synergistically produced eNOS expression. Thereby, we believe that the co-delivery of eNOS and ZNF₅₈₀ genes assisted by a cascaded bio-responsive carrier is a powerful strategy to treat CLI.