Determination of homocysteine levels in endothelial and stromal cell cultures by HPLC

CHED 205

Adrienne Riggi, ariggi@fairmontstate.edu, Gregory Donohoe, gdonohoe@fairmontstate.edu, Zachary Brewer, zbrewer@fairmontstate.edu, Bonnie Freeman, bfreeman@fairmontstate.edu, Mark Flood, mflood@fairmontstate.edu, Sarah Dodson, sdodson@fairmontstate.edu, and Andreas Baur, abaur@fairmontstate.edu. Department of Biology, Chemistry and Geoscience, Fairmont State University, 1201 Locust Ave, Fairmont, WV 26554
An increased blood level of homocysteine (Hcy) has been associated with a higher risk of cardiovascular disease. It was observed that homocysteine has a negative effect on the growth of endothelial cells but a positive effect on the growth of vascular smooth muscle cells. In this study, adherent murine S10 stromal cells and endothelial (HUVEC) cell types were cultured in a 37°C environment with 5% CO2 atmosphere. Cells were exposed to 0 to 500 µM of DL-homocysteine for 48 hours. The homocysteine metabolism was investigated by determining changes in the extra and intracellular Hcy concentration after the initial exposure. Aliquots from the culture medium or intracellular matrix were reduced with tris-(2-carboxyethyl) phosphine, derivatized by SBD-F and analyzed by HPLC (reverse-phase C-18, 0.1 M KH2PO4, pH 2.0, fluorescence detection: excitation 385 nm, emission 515 nm). In addition, cell apoptosis was assessed through DNA laddering assays and changes in cell morphology.
 

Undergraduate Research Poster Session
2:30 PM-4:30 PM, Monday, August 20, 2007 BCEC -- Exhibit Hall - B2, Poster

Division of Chemical Education

The 234th ACS National Meeting, Boston, MA, August 19-23, 2007