Effects of ascorbic acid on trabecular meshwork gene expression and collagen secretion

Area of Science:

• Ocular Biology
• Cellular Biochemistry
• Biotechnology

Background:

• Aqueous humour (AH) is rich in vitamin C (ascorbate), an essential antioxidant.
• Trabecular meshwork (TM) cells, crucial for AH drainage, are exposed to high ascorbate levels in vivo.
• Standard in vitro cell culture often omits ascorbate, potentially altering TM cell behavior.

Purpose of the Study:

• To investigate the molecular changes in TM cells cultured with and without ascorbate.
• To identify specific genes and proteins affected by ascorbate treatment in TM cells.

Main Methods:

• TM cells were cultured in serum-free media with or without 0.1 mM ascorbate for 3 days.
• Protein expression was analyzed using SDS-PAGE and Western blotting, with LC-MS/MS for identification.
• Gene expression was quantified via fluorescent barcode technology and validated by quantitative PCR.

Main Results:

• Ascorbate treatment led to proper modification and secretion of collagen chains.
• Gene expression analysis revealed 76 upregulated and 20 downregulated genes.
• Upregulated genes included those involved in antioxidant stress, collagen synthesis, extracellular matrix remodeling, and signaling pathways like VEGF, TGF-β, and Hedgehog.

Conclusions:

• Ascorbate significantly regulates multiple collagens and 96 genes in TM cells.
• The findings underscore the necessity of including ascorbate in in vitro TM cell culture media.
• Further research into the identified genes' functions can enhance understanding of ascorbate's role in TM cell health.