Detection of Autophagy-Related Gene Expression by Conjunctival Impression Cytology in Age-Related Macular Degeneration
Chih-Wen Shu1,2,3, Youn-Shen Bee4, Jiunn-Liang Chen4
1Institute of Biopharmaceutical Sciences, National Sun Yat-Sen University, Kaohsiung 80424, Taiwan.
Purpose:
To investigate the association of autophagy-related gene expression with age-related macular degeneration (AMD).
Methods:
Patients with AMD were recruited for analysis by conjunctival impression cytology. mRNA was assessed by real-time polymerase chain reaction (RT-PCR) to evaluate whether the expression of 26 autophagy-related genes (ATGs) was correlated with AMD. Further studies on cell viability and autophagic flux in response to oxidative stress by H2O2 were performed in human retinal pigment epithelial (RPE) cell lines based on the results of impression cytology.
Results:
Both the neovascular AMD (nAMD) and polypoidal choroidal vasculopathy (PCV) groups had significantly higher mRNA levels of gamma-aminobutyric acid receptor-associated protein-like 1 (GABARAPL1) and microtubule-associated proteins 1A/1B light chain 3B (MAP1LC3B) than the control group, but there was no significant difference between these two groups. Age difference existed only in the AMD group. GABARAPL1 and MAP1LC3B mRNA expression increased significantly after acute oxidative stress in adult retinal pigment epithelial (ARPE-19) cells. Cell viability significantly increased and decreased in the cells harboring GABARAPL1 expression vector and silenced with siRNA against GABARAPL1, respectively, during short-term oxidative stress, whereas viability increased in the GABARAPL1-silenced cells after long-term oxidative stress. Silencing GABARAPL1 itself caused a reduction in autophagic flux under both short and long-term oxidative stress.
Conclusion:
Our study showed the possibility of assessing autophagy-related gene expression by conjunctival impression cytology. GABARAPL1 was significantly higher in AMD. Although an in vitro study showed an initial protective effect of autophagy, a cell viability study revealed the possibility of a harmful effect after long-term oxidative injury. The underlying mechanism or critical factors require further investigation.
Insights
Autophagy-related gene expression, including GABARAPL1, is elevated in age-related macular degeneration (AMD). While autophagy may initially protect against oxidative stress, its long-term effects in AMD require further investigation.
Area of Science:
- Ophthalmology
- Molecular Biology
- Genetics
Background:
- Age-related macular degeneration (AMD) is a leading cause of vision loss.
- Autophagy, a cellular degradation process, plays a role in various diseases.
- Understanding the role of autophagy-related genes (ATGs) in AMD is crucial.
Purpose of the Study:
- To investigate the association between autophagy-related gene expression and age-related macular degeneration (AMD).
- To explore the potential of using conjunctival impression cytology for assessing ATG expression in AMD patients.
Main Methods:
- Recruited patients with AMD for conjunctival impression cytology.
- Assessed mRNA levels of 26 autophagy-related genes (ATGs) using RT-PCR.
- Performed in vitro studies on human retinal pigment epithelial (RPE) cells to assess cell viability and autophagic flux under oxidative stress.
Main Results:
- Significantly higher mRNA levels of GABARAPL1 and MAP1LC3B were observed in AMD patients (both neovascular AMD and polypoidal choroidal vasculopathy) compared to controls.
- GABARAPL1 and MAP1LC3B mRNA expression increased in RPE cells following acute oxidative stress.
- GABARAPL1 exhibited an initial protective effect on cell viability during short-term oxidative stress but potentially a harmful effect during long-term oxidative injury.
Conclusions:
- Conjunctival impression cytology can assess autophagy-related gene expression.
- GABARAPL1 expression is significantly elevated in AMD.
- The role of autophagy in AMD is complex, with potential protective and harmful effects depending on the duration of oxidative injury, warranting further mechanistic studies.
