Etablierung eines Zellkulturmodells zur Erforschung zyklischer Druckschwankungen an retinalen Ganglienzellen

Development of a cell culture model to investigate cyclic pressure fluctuations on retinal ganglion cells

by Matthias Strake
Doctoral thesis
Date of Examination:2025-06-04
Date of issue:2025-05-19
Advisor:PD Dr. Christian van Oterendorp
Referee:Prof. Dr. Tim Gollisch
Referee:Prof. Dr. Margarete Schön
Persistent Address: http://dx.doi.org/10.53846/goediss-11271

 

 

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Abstract

English

Glaucoma, one of the most common causes of irreversible vision loss, leads to the progressive degeneration of retinal ganglion cells (RGCs), with elevated intraocular pressure being the only modifiable risk factor. However, the exact mechanisms by which pressure damages RGCs are not well understood. This study aimed to establish and validate an in vitro cell culture model that simulates the effects of cyclic hydrostatic pressure and shear stress on RGCs in live-cell imaging. A controllable pressure chamber system was developed for mitochondrial imaging of RGCs in primary cell cultures from postnatal rats (day 1–3). Several modifications of the setup were tested, including changes to the surrounding environment using polyacrylamide hydrogels. In the second part of the study, RGCs were exposed to rapid cyclic pressure fluctuations ranging from 0–60 mmHg (one cycle every 8 seconds) for up to 9–16 hours. Rhodamine 123-labeled mitochondria in the longest neurites were tracked via live-cell fluorescence microscopy. Several motility parameters were quantified, such as the proportion of motile versus stationary mitochondria, travel distances and velocity distributions. Subsequently, the fusion and fission behavior of mitochondria was examined, as well as cell survival and neurite outgrowth. RGC identity was confirmed afterwards via anti-βIII-tubulin staining. Contrary to initial expectations, RGCs showed remarkable resilience to cyclic pressure stimuli: no significant differences between cells exposed to pressure and those under atmospheric pressure were found in mitochondrial motility or fusion/fission behavior. Likewise, cell survival and neurite length remained unchanged. These findings suggest that isolated cyclic hydrostatic pressure in the tested range is not sufficient to induce mitochondrial or cellular dysfunction in RGCs.
Keywords: glaucoma; retinal ganglion cell; rgc; mitochondria; hydrostatic pressure