MO_II_P: Molecular Sciences II. Posters
Minh N. Tran 1,2, Balazs Besztercei 3, Illes Kovacs 4, Xavier Gasull 5, Attila J. Szabo 2,6, Andrea Fekete 1,2, Judit Hodrea 1,2
1. SE - Diabetes Research Group, Semmelweis University, Budapest
2. 1st Department of Pediatrics, Semmelweis University, Budapest
3. Institute of Clinical Experimental Research, Semmelweis University, Budapest
4. Department of Ophthalmology, Semmelweis University, Budapest
5. Department of Biomedicine, Institute of Neurosciences, University of Barcelona, Barcelona
6. ELKH-SE Pediatric and Nephrology Research Group, Semmelweis University, Budapest
Trabecular meshwork (TM) tissue forms the primary egress route of aqueous humor (AH) from the eye. TM maintains normal intraocular pressure (IOP) by regulating AH outflow resistance. Impairing in TM functions affects adversely the outflow pathway which can cause increased IOP and glaucoma.
The neuroprotective effect of sigma-1 receptor (S1R) has been widely investigated in the central nervous system. We showed previously that fluvoxamine (Flu), a specific S1R agonist, is expressed in the kidney and protects against ischemia-reperfusion injury.
To investigate the role of S1R on pro-fibrotic factors-induced cell- proliferation and migration, fibrotic protein levels and actin cytoskeletal rearrangement in primary mouse TM (MsTM) cells and non-glaucomatous human TM cell line (HTM5).
Positive magnetic bead selection was used to isolate primary TM from wild type (WT) and S1R knock-out (KO) mice. HTM5 and MsTM cells were treated for 24h either with 20 ng/mL platelet-derived growth factor (PDGF) or 10 ng/mL transforming growth factor-beta 2 (TGF-β2) combined with 10 µM of Flu. Then Western blot and immunocytochemistry were used to detect and localize S1R in the cells. Cell proliferation was determined by MTT assay, the cell migration by scratch assay, F-actin was visualized by immunostaining and detected with fluorescent microscope. The fibrotic protein levels were detected by immunoblotting.
The magnetic bead-based method was efficient for isolating MsTM cells with minimal microdissection techniques required. S1R is present in both primary MsTM and HTM5 cells and is localize in the endoplasmic reticulum. Cell proliferation, migration and cytoskeletal rearrangement was induced by PDGF and it was prevented by Flu in all assays. The formation of F-actin bundles and actin-clumps that was more pronounced in KO cells. TGF-β2 induced the level of connective tissue growth factor and fibronectin that were also inhibited by Flu.
Flu can reduce the profibrotic factors-induced cytoskeletal rearrangement and fibrotic marker expression that may lead to lower outflow resistance and enhance outflow facility. Our results propose that Flu could be a potential candidate for the development of a novel IOP-lowering drug.
Grants: OTKA- K135398, PD-131637, FK-124491, 2020-4.1.1.-TKP2020-6183069269 (FIKP), 2020-4.1.1.-TKP2020-6183169273.
Semmelweis University, Károly Rácz Doctoral School of Clinical Medicine