Analysis of the metabolic and oxygen status of 3D neurospheres from iPSCs using FLIM / PLIM methods

A. Kashirina; E. Momotyuk; E. Bukina; E. Dashinimaev; E. Vorotelyak; M. Lukina; V. Dudenkova; A. Gavrina; E. Kryukov; V. Elagin; E. Zagaynova; A. Kashina

doi:10.1117/12.2577638

5 March 2021 Analysis of the metabolic and oxygen status of 3D neurospheres from iPSCs using FLIM / PLIM methods

A. Kashirina, E. Momotyuk, E. Bukina, E. Dashinimaev, E. Vorotelyak, M. Lukina, V. Dudenkova, A. Gavrina, E. Kryukov, V. Elagin, E. Zagaynova, A. Kashina

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Proceedings Volume 11647, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XIX; 1164709 (2021) https://doi.org/10.1117/12.2577638
Event: SPIE BiOS, 2021, Online Only

Abstract

3D models based on cells differentiated from patient-specific induced pluripotent stem cells (iPSCs) are widely used to identify disease phenotypes, to accurately analyze dysfunctions at the level of human tissues and organs, to screen new drugs, which makes them more promising tool for biomedical research tasks than monolayer cultures, which is associated with their proximity to in vivo. The metabolic activity with oxygenation level of cells, assessed by optical imaging methods, can be used as markers of cell viability, proliferative activity and the degree of differentiation in 3D culture conditions. In this paper we used fluorescence and phosphorescence lifetime imaging microscopies (FLIM and PLIM) to study the metabolic status and the oxygenation level of derived from iPSCs neural stem cells (NSC) cultured in 3D condition. An analysis of the fluorescence intensities and FLIM data showed that NSCs in monolayer and at the periphery of large spheroids have more glycolytic phenotypes, NSCs in the center of large spheroids and NSCs grouped into small spheroids have more oxidative state. For determination of the relative oxygen level in spheroids PLIM of BPTDM stained neurospheres was carried out. As it was supposed, oxygen transport in the spheroid depended on it size. In neurospheres with an average size 600 μm O2 distribution is radial, with the lowest concentration in the center. Thus, the metabolic status and oxygenation level of the NSC in the spheroid composition was assessed in a life-time and noninvasive manner.

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A. Kashirina, E. Momotyuk, E. Bukina, E. Dashinimaev, E. Vorotelyak, M. Lukina, V. Dudenkova, A. Gavrina, E. Kryukov, V. Elagin, E. Zagaynova, and A. Kashina "Analysis of the metabolic and oxygen status of 3D neurospheres from iPSCs using FLIM / PLIM methods", Proc. SPIE 11647, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XIX, 1164709 (5 March 2021); https://doi.org/10.1117/12.2577638

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