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Influence of pathogenic stimuli on Müller cell transfection by lipoplexes.

European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V (2020-03-17)
Karen Peynshaert, Joke Devoldere, Frederik Philips, Fauve Vergauwe, Stefaan De Smedt, Katrien Remaut
ABSTRACT

Neuroprotection is a mutation-independent therapeutic strategy that seeks to enhance the survival of neuronal cell types through delivery of neuroprotective factors. The Müller cell, a retinal glial cell type appreciated for its unique morphology and neuroprotective functions, could be regarded as an ideal target for this strategy by functioning as a secretion platform within the retina following uptake of a transgene of our choice. In this in vitro study we aimed to investigate the capability of Müller cells to take up a standard liposomal vector (i.e. Lipofectamine 2000) and process its pDNA or mRNA cargo into the reporter GFP protein. By doing so, we found that mRNA outperformed pDNA in Müller cell transfection efficiency. Since neuroprotection is explored as a therapy for diabetic retinopathy and glaucoma, we furthermore examined the Müller cell's lipoplex-induced transfection efficiency and cytotoxicity in stressful conditions linked to these diseases - i.e. hypoxia, hyperglycemia and oxidative stress. Interestingly, Müller cells were able of maintaining high GFP expression regardless of these noxious stimuli. In terms of lipoplex-induced toxicity, hyperglycemia seemed to have a protective effect while hypoxia and oxidative stress led to a slightly higher toxicity. In conclusion, our study indicates that mRNA-lipoplexes have potential in transfecting Müller cells in healthy as well as diseased conditions.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Luperox® TBH70X, tert-Butyl hydroperoxide solution, 70 wt. % in H2O
Sigma-Aldrich
D-(+)-Glucose solution, 100 g/L in H2O, sterile-filtered, BioXtra, suitable for cell culture
Corning® CellBIND® Surface cell culture flasks, CellBIND 75cm2 rectangular canted neck cell culture flask w/ Vent Cap, PS, sterile, 100/cs

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