Figure 7. Effect of NO on cellular heme incorporation into CYP 2D6 and 3A4.
HEK293T cells were treated with SA for 48 h followed by transient transfection to initiate CYP 2D6 or 3A4 protein expression. At 24 h post-transfection the cells were treated with 7.5 μM hemin alone or with 125 μM NOC-18 for 3 h before being harvested and supernatants prepared. Some cell cultures were washed to remove NOC-18 and hemin and were then cultured for an additional 3 h prior to harvesting. Panel A, Heme incorporation into the CYP apo-enzymes under the indicated conditions (determined by the spectroscopic method) with values normalized per mg total protein. Panels B and C, Western analysis of CYP protein expression levels in aliquots of equal total protein content for the supernatant samples, and corresponding in-gel heme staining of the CYP protein bands. Results are representative of three trials.