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Madina Khubetsova, Sergey Borzenok, Irina Saburina, Natalia Gavrilova, Nastasia Kosheleva, Khadizhat Tonaeva, Patimat Arbukhanova, Natalia Lanevskaya, Dmitry Ostrovsky, Maria Shurygina, Ilia Popov; Neuroprotective potential of 3D culture of human limbal mesenchymal stem cells.. Invest. Ophthalmol. Vis. Sci. 2017;58(8):2570.
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© ARVO (1962-2015); The Authors (2016-present)
It is known that nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) have an antiapoptotic effect and exert the neuroprotection of retinal ganglion cells in glaucoma. However, still it is necessary to solve the question concerning the safe and long-term delivery of neurotrophins. In this trial we studied the ability of 3D-culture of limbal mesenchymal stem cells (MSCs) to secrete NGF and BDNF during in vitro experiment.
MSCs were obtained by culturing the limbal fragments isolated from the cadaveric human eyes. The cell culture were characterized by flow cytometry through the use of specific MSC marker antibodies. In this trial, we carried out the stimulation of the neurotrophic factors secretion using the two-step procedure according to the protocol of O. Sadan (2008).3D-cell spheroids were created using the agarose plates for two comparison groups: I group – intact spheroids, II group – spheroids stimulated on the 7th day of cultivation. The supernatants of cell cultures were collected at different times and research for the content of NFG and BDNF by ELISA.Two-tailed Student’s t-test was used for statistical analysis.
It was established that 3D-cell culture of limbal MSCs is able to secrete NGF and BDNF at least for 21 days. NGF secretion level in the I group remained stable throughout the whole observation period and was on average 11785±389 pg/ml. When evaluating the BDNF content dynamics in the culture liquid the I group had a moderate decrease in the factor level: to 21 days the concentration was 461.59±23.09 pg/ml. The conduction of induction contributed to increase in the NGF and BDNF secretion by 1.8 and 1.6 and was on average 19490±871 and 155.9± 8.2 pg/ml, respectively (p<0.05). Later secretion of the factors significantly decreased and to 21 day it was by 1.8 and 1.4 less compared to the I group that was 6280±223.8 pg/ml, 330 ± 14.1 pg/ml, respectively (p<0.05).
3D cell culture of intact MSCs can be considered as a cell product for safe and long-lasting neuroprotection in treatment of glaucoma optic neuropathy.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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