Purchase this article with an account.
Andrei A Kramerov, Pallavi Gangalum, Hui Ding, Julia Y Ljubimova, Alexander V Ljubimov; Novel Nanoconjugates for Gene Therapy Normalization of Cultured Human Diabetic Limbal Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 201657(12):.
Download citation file:
© 2017 Association for Research in Vision and Ophthalmology.
Our purpose was to normalize cultured human diabetic limbal epithelial cells (LEC) that have reduced expression of epithelial stem cell markers and slow wound healing using novel polymalic acid-based nanoconjugates inhibiting MMP-10 and cathepsin F (increased in diabetic corneas) and upregulating c-met (decreased in diabetic corneas).
LEC cultures enriched in stem cells were obtained from postmortem donor eyes. Nanoconjugates on natural-derived polymalic acid scaffold were synthesized as published (1) and contained a targeting antibody to LEC-expressed transferrin receptor (TfR), morpholino antisense oligos (AON) to cathepsin F and to miR-409-3p that targets c-met proto-oncogene, and trileucine endosome escape unit. A separate nanoconjugate contained an AON to MMP-10 and a tracking dye Alexa Fluor 488. Efficiency of target inhibition (proteinases) or upregulation (c-met) was tested with free AON and then with whole nanoconjugates (at 5 mM AON). Healing of scratch wounds was monitored microscopically.
Diabetic LEC from several donors were TfR-positive on western blots justifying the use of respective antibody for cell targeting. LEC internalized nanoconjugates mainly through receptor-mediated endocytosis as revealed by competition with free TfR antibody. By western blot, they were able to reduce the expression of MMP-10 and cathepsin F, and increase the expression of c-met (through inhibition of miR-409). Treatment with nanoconjugates accelerated healing of scratch wounds in LEC cultures with no toxicity observed.
Non-toxic nanoconjugates appear to be a new viable alternative to viral-based gene therapy in normalizing the diabetic limbal epithelial cells and wound healing. 1. Proc Natl Acad Sci USA, 2010;107:18143-18148.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
This PDF is available to Subscribers Only