In vitro model suggests oxidative stress involved in keratoconus disease

Area of Science:

• Ophthalmology
• Cell Biology
• Biochemistry

Background:

• Keratoconus (KC) is a corneal disorder leading to thinning and a conical shape, often requiring surgery.
• The specific role of oxidative stress in the pathogenesis of keratoconus remains incompletely understood.
• Understanding cellular differences in oxidative stress is crucial for developing targeted therapies.

Purpose of the Study:

• To investigate and compare oxidative stress levels in human corneal keratocytes (HCKs), human corneal fibroblasts (HCFs), and keratoconus cells (HKCs).
• To analyze the impact of extracellular matrix (ECM) stimulation on oxidative stress markers.
• To explore potential phenotypic differences between HCKs, HCFs, and HKCs.

Main Methods:

• Cell culture in 2D and 3D systems with Vitamin C and TGF-β3 stimulation.
• Analysis of gene expression using qRT-PCR for collagen types and keratocan.
• Metabolomic profiling using LC-MS/MS to quantify over 150 metabolites, including oxidative stress markers.

Main Results:

• HKCs showed reduced collagen I, V, and keratocan, alongside upregulated collagen type III, indicating a myofibroblast phenotype.
• Elevated lactate levels, lactate/malate, and lactate/pyruvate ratios were observed in HKCs.
• Reduced arginine and glutathione/oxidized glutathione ratios were detected in HKCs, consistent with increased oxidative stress.

Conclusions:

• Fibroblasts exhibit higher oxidative stress compared to keratocytes.
• Keratoconus cells (HKCs) display the most significant oxidative stress, supporting a potential myofibroblast phenotype.
• These findings highlight the role of oxidative stress in keratoconus pathogenesis and suggest a distinct cellular phenotype in affected corneas.