[A new methods for determining hydrogen sulfide release in cultured cells]

Area of Science:

• Biochemistry
• Cell Biology
• Analytical Chemistry

Background:

• Hydrogen sulfide (H2S) is a crucial signaling molecule in biological systems.
• Accurate measurement of H2S in living cells is essential for understanding its physiological roles.
• Existing methods for H2S detection in cellular environments can be complex or lack sensitivity.

Purpose of the Study:

• To develop and validate a simple, reliable method for quantifying hydrogen sulfide (H2S) production in cultured living cells.
• To assess the feasibility of this method for distinguishing H2S release between cell lines with different enzyme expressions.

Main Methods:

• A modified cell culture plate lid with a filtration membrane was used to trap H2S released from cells.
• Trapped H2S was converted to zinc sulfide (ZnS) using zinc acetate.
• ZnS was quantified using the methylene blue assay, with H2S production calculated against a standard curve.

Main Results:

• The method successfully measured H2S generation in HepG2 and HUVEC cell lines, revealing significant differences in production rates.
• Inhibition of cystathionine-gamma-lyase (CSE) and cystathionine-beta-synthase (CBS) enzymes demonstrably reduced H2S release in HepG2 cells.
• Cell viability remained above 95%, indicating no significant cytotoxicity associated with the measurement technique.

Conclusions:

• The developed method, utilizing a modified instrument integrated into a cell culture plate lid, is effective for detecting H2S release in living cells.
• This simple and non-cytotoxic approach provides a valuable tool for cellular H2S research.