Abstract
Introduction Glutathione deficiency and chronic bacterial inflammation exacerbates the oxidative stress damage to airways in cystic fibrosis. Improvements to current antioxidant therapeutic strategies are needed. Dietary supplement, γ-glutamylcysteine (GGC), the immediate precursor to glutathione, rapidly boosts cellular glutathione levels following a single dose in healthy individuals. Efficacy of GGC against Pseudomonas aeruginosa derived lipopolysaccharide (LPS), a prominent factor in mediating both bacterial virulence and host responses, in CF remains unassessed.
Methods Primary F508del/F508del mucociliary differentiated bronchial and nasal epithelial cells were created to model LPS-induced oxidative stress and inflammation of CF. The proteomic signature of GGC treated cells was resolved by qLC-MS/MS. Parameters including cell redox state (glutathione, ROS), anti-inflammatory mediators (IL-8, IDO-1) and cellular health (membrane integrity, stress granule formation and cell viability) were assayed.
Results Proteomic analysis identified perturbation of several pathways related to cellular respiration and stress responses upon LPS challenge. Most of these were resolved when cells were treated with GGC. While GGC did not resolve LPS-induced IL-8 and IDO-1 activity, it effectively attenuated LPS-induced ROS and stress granule formation, while significantly increasing intracellular glutathione levels and improving epithelial cell barrier integrity. Moreover, we compared the effect of GGC with thiols NAC and glutathione on cell viability. GGC was the only thiol that increased cell viability; protecting cells against LPS induced cell death. Both therapeutic and prophylactic treatments were successful.
Conclusion Together, these findings indicate that GGC has therapeutic potential for treatment and prevention of oxidative stress related damage to airways in Cystic Fibrosis.
Competing Interest Statement
The authors have declared no competing interest.
Abbreviations
- ABC
- ATP-binding cassette
- ALI
- Air-liquid interface
- BALF
- Bronchoalveolar lavage fluid
- BEGM
- Bronchial epithelial growth medium
- CFTR
- Cystic fibrosis transmembrane regulator
- CREC
- Conditionally reprogrammed epithelial cell
- ELF
- Epithelial lining fluid
- ELISA
- Enzyme linked immunosorbent assay
- GGC
- γ-glutamylcysteine
- GLRX
- Glutaredoxin
- GPx
- Glutathione peroxidase
- GSH
- Glutathione
- HAEC
- Human airway epithelial cell
- HBEC
- Human bronchial epithelial cell
- HNEC
- Human nasal epithelial cell
- HPLC
- High performance liquid chromatography
- IPA
- Ingenuity Pathway Analysis
- LC-MS/MS
- Liquid chromatography tandem mass spectrometry
- LPS
- Lipopolysaccharide
- NAC
- N-acetylcysteine
- PIC
- Pre-initiation complex
- ROS
- Reactive oxygen species
- SG
- Stress granule
- SOD
- Superoxide dismutase