Studies on the Development of Targeted Gene Delivery Systems for Reversal of Atherosclerosis and obesity

Title
Studies on the Development of Targeted Gene Delivery Systems for Reversal of Atherosclerosis and obesity
Authors
Qurrat Ul Ain
Advisor(s)
Yong-Hee Kim
Issue Date
2017-08
Publisher
한양대학교
Degree
Doctor
Abstract
Endothelial dysfunction combined with inflammation leads to atherosclerosis. Endothelium-specific delivery of therapeutic agents at the cellular level—specifically in vivo—is still a difficult task for proper management of atherosclerosis. We designed a redox-sensitive poly(oligo-L-arginine) (rsPOLA) playing dual roles as an endothelium α-2 adrenoceptors(α-2ARs)-targeted gene carrier and as a substrate for endothelial nitric oxide synthase (eNOS). Overexpression of α-2ARs on atherosclerotic endothelial cells was confirmed and the eNOS/rsPOLA nanoplexes following systemic injection demonstrated to 1) enhance eNOS gene delivery into endothelial cells via α-2ARs/L-arginine specific binding, 2) increase intracellular level of nitric oxide, 3) suppress inflammatory response in endothelium and finally 4) reduce atherosclerotic plaque in a Ldlr-/- atherosclerotic mouse model. Among the tested nanoplexes [eNOS/rsPOLA, eNOS/(poly(oligo-D-arginine), rsPODA) and eNOS/(racemic mixture, rsRM)], eNOS/rsPOLA reduced atherosclerotic inflammation most effectively as we hypothesized. Current treatment strategy provides strong potential for further development of a gene therapeutic system to ameliorate inflammation and progressive atherosclerotic plaques. Obesity is a progressively prevailing in developed societies. In spite of all major progress in the understanding of molecular mechanisms of obesity, currently available commercial anti-obesity drugs have been shown very limited efficacy with severe side effects. Anti-obesity drug development industry is thus concentrating on targeting adipocytes that store excess fat. In this study we used a peptide motif (CKGGRAKDC) that homes to adipocytes, thus named as adipocyte-targeting sequence (ATS). An adipocyte-targeting fusion-oligopeptide gene carrier was developed by conjugating ATS and 9-arginine (ATS-9R). This peptide binds with prohibitin, a vascular marker of adipose tissue. We also constructed a single guide RNA (sgRNA) and complexed it with deactivated Cas9 protein (dCas) to develop a complex to knock-down fatty-acid-binding protein 4 (FABP4) via CRISPR interference technique. Transfection of dCas9+sgFABP4 through ATS-9R caused FABP4 knock down in pre adipocytes and mature adipocytes (>40%). The dCas9+sgFABP4/ATS-9R oligoplexes could be a safe therapeutic approach to regress and treat obesity as well as obesity-induced metabolic syndromes.
URI
http://dcollection.hanyang.ac.kr/jsp/common/DcLoOrgPer.jsp?sItemId=000000102516http://hdl.handle.net/20.500.11754/33396
Appears in Collections:
GRADUATE SCHOOL[S](대학원) > BIOENGINEERING(생명공학과) > Theses (Master)
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