Background: It has been suggested that replicative senescence might be involved in the pathophysiology of age-related diseases.
Aim: To study the process of senescence in trabecular meshwork (TM) cells.
Methods: Porcine TM tissues were obtained and placed in primary cultures with Dulbecco’s modified Eagle’s medium/Ham’s F-12 medium. After 2–3 weeks, migrated and proliferated TM cells were trypsinised and cultured in serial passages, and identified with fluorescein-labelled low-density lipoprotein (DiI-Ac-LDL), a marker of TM cells. Staining for senescence-related β-galactosidase activity was performed at population doubling level (PDL) 2, 8 and 16 at pH 6. Terminal restriction fragment (TRF) length was examined by Southern blot analysis using a 32P-labelled telomere-specific sequence (TTAGGG)3 at each PDL.
Results: DiI-Ac-LDL staining revealed that most (nearly 100%) of the cells in the culture were TM cells, which were flattened in shape and positive for senescence-related β-galactosidase staining at PDL 16. Reduction of TRF length as a function of population doubling was also shown.
Conclusions: TM cells exhibited characteristics of senescence at PDL 16 in vitro. The results demonstrated that cellular senescence may be related to the pathophysiology of primary open-angle glaucoma.
- PDL, population doubling level
- POAG, primary open-angle glaucoma
- LDL, low-density lipoprotein
- TM, trabecular meshwork
- TRF, terminal restriction fragment
- RPE cells, retinal pigment epithelial cells
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Published Online First 10 January 2007
Competing interests: None.