Abstract 1358 Poster Session IV, Tuesday, 5/4 (poster 359)

Heme oxygenase (HO) catalyzes the rate-limiting step in the predominant pathway for heme degradation leading to the equimolar production of Fe(II), carbon monoxide (CO), biliverdin, and, ultimately, bilirubin. We are developing a hemolytic mouse (Balb-C) model for studying in vivo efficacy of metalloporphyrin inhibitors of HO to suppress bilirubin production, which is measured by total body CO excretion rate (VeCO) with a flow-through chamber system. The VeCO of each mouse in a 50-mL chamber, supplied with 30 mL air/min, was monitored by measuring chamber exit air for CO by gas chromatography. The basal VeCO was established before mice (20.9 ± 3.1 g) were injected via the tail vein with 30-µmol heme arginate/kg. At various times during the VeCO measurement period, animals were sacrificed for determination of hepatic and splenic HO activity by the gas chromatographic CO method. We found that the VeCO of untreated animals (58 ± 10 µL CO/hr/kg) did not change during the 6-h monitoring period. However, the VeCO of heme-treated animals started to increase 15 ± 5 min after injection, and after 78 ± 15 min, reached a peak VeCO of 202 ± 27 µL CO/hr/kg. The VeCO returned to baseline after 6h. During this period, splenic HO activity did not change significantly from baseline (555 ± 82 µmol CO/hr/mg FW tissue). In contrast, the basal hepatic HO activity (141 ± 29 µmol CO/hr/mg FW tissue) began to increase approximately 1.5h after heme injection, reached a plateau at 2.5 ± 1h of 719 ± 75 µmol CO/hr/mg FW tissue, then slowly diminished from 6h to 12d when 138% of baseline HO activity, or 19% of peak HO activity, remained. We conclude that the mouse is an attractive hemolytic model for the study of HO inhibitors because it displays a relatively rapid and reproducible VeCO profile in comparison to the rat model. Furthermore, we have improved the HO activity assay so that the activity in organs weighing as little as 50 mg can be easily determined. We demonstrate here for the first time that hepatic HO activity upregulation occurs quickly and shortly (1.5h) after heme injection and remained at a high level for days, whereas the injected heme was metabolized rather quickly. In contrast, splenic HO activity was not increased because this organ apparently is maximally active.