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Simona Nicoara, Cristina Cristian, Cristian Berce, Flaviu Tabaran, Oana Tudoran, Sanda Boca-Farcau, Simion Astilean, Olga Soritau; Nanotechnology for improvement of stem cell therapy of laser damaged retina in an animal model. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4624.
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© ARVO (1962-2015); The Authors (2016-present)
We aimed to develop an animal model of retinal injury and repair by the intravitreal administration of adult mesenchymal stem cells (MSCs) using gold nanoparticles (GNPs) as delivery system for neuronal specific growth factors (NGFs).
Isolated MSCs from the bone marrow of CD1 mice were characterized by immuno-fenotyping and RT-PCR for stem cell markers. We produced GNPs and conjugated them with NGF-beta in order to attain a specific functionality. 20 nm GNPs were chemically synthesized by citrate reduction of gold salt and functionalized with NGF-beta through nonspecific electrostatic interaction between the nanoparticles and the protein. GNPs were characterized by several techniques: transmission electron microscopy, UV-VIS absorption spectroscopy, dynamic light scattering and zeta potential. In vitro differentiation ability of MSCs into retinal progenitors was investigated using a multistep protocol with exploration of the most suitable combination of growth factors and GNPs conjugated with NGF-beta, as well as the most suitable cell substrate. The inducing of the retinal lesions by indirect diode laser photocoagulation in CD1 mice was followed by the intravitreal administration of fluorescent MSCs and GNPs conjugated with NGF. The evaluation of MSC grafting and retinal morphology was performed at 1 and 3 weeks after the procedure, by confocal microscopy and hystopathology.
Isolation and cultivation of MSCs from bone marrow of CD1 mice was possible even without substrate. The substrate supports better the cell proliferation and reduces the percentage of cell death during differentiation. Isolated cells were adult type MSCs. The multistep protocol requires further investigation, mainly in stage 4 where cells might require a richer medium. Growing these cells on substrates that mimic the extracellular matrix (laminin and collagen) supported the viability and cell proliferation. MSCs in CD1 mice replaced the degenerated retinal cells and the association with GNPs as delivery system for NGFs improved the differentiation potential of MSCs.
The administration of cells in the vitreous cavity has an acceptable invasiveness with low risk of infection. GNPs conjugated with NGFs offer a proper microenvironment for MSCs differentiation into retinal cells and is an alternative delivery system to viral vectors.
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