Abstract
Purpose :
Bone marrow-derived activated monocytes and progenitor cells migrate to the retina in response to inflammation and neovascularization. Hypoxia-inducible factor-1α (HIF-1α) has been shown to contribute to the pathogenesis of neovascularization. However, contribution of monocyte-derived macrophages to neovascularization is largely unknown. We describe here the synthesis of a new hybrid nanoparticle for targeted delivery and gene silencing in activated monocytes that are associated with pathological neovascularization.
Methods :
Single-cell RNAseq data analysis were performed to characterize HIF-1α mRNA expression in activated monocytes in mouse oxygen-induced retinopathy (OIR). HIF-1α targeted AS-shRNA-lipids were synthesized by conjugating diacyl-lipids to anti-sense short hairpin RNA with an anti-sense sequence complimentary to HIF-1α mRNA. The short hairpin RNA compounds were stabilized using 2’-O-methyl-protected ribonucleotides allowing in vivo delivery of AS-shRNA-lipid to the activated monocytes that are associated with neovascularization.
Results :
HIF-1α mRNA expression was associated with MRC-1 positive activated monocytes. In addition, HIF-1α mRNA was induced in monocytes by exposing the cells to hypoxia, and the expression was inhibited by AS-HIF-1α-shRNA-lipid. CD14 mRNA expression was significantly elevated in monocytes that were exposed to hypoxia, suggesting monocyte activation. AS-HIF-1α-shRNA-lipid could significantly inhibit this activation. In addition, significant reduction of neovascularization was achieved in mouse OIR after intraperitoneal injection of AS-shRNA-lipids.
Conclusions :
Inflammation and hypoxia could activate monocyte functions and inhibition of HIF-1α mRNA could recover these cells from activation. In summary, we have developed a novel method for targeted delivery and inhibit HIF-1α mRNA in activated monocytes in living ocular tissues using diacyl-lipid-conjugates of antisense short hairpin RNA (AS-shRNA-lipid) designed to target HIF-1α mRNA. These conjugates are readily internalized by activated monocytes that are associated with neovascularization in the living retina. These findings may provide a framework for a novel strategy to inhibit retinal neovascularization.
Keywords: targeted delivery, activated monocytes, gene therapy, short-hairpin RNA, neovascularization.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.