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. 2009 Jan;37(1):263-9.
doi: 10.1097/CCM.0b013e3181931ed3.

Moderate hypothermia prevents cardiac arrest-mediated suppression of drug metabolism and induction of interleukin-6 in rats

Michael A Tortorici  1 , Ying Mu, Patrick M Kochanek, Wen Xie, Samuel M Poloyac
Affiliations

Moderate hypothermia prevents cardiac arrest-mediated suppression of drug metabolism and induction of interleukin-6 in rats

Michael A Tortorici et al. Crit Care Med. 2009 Jan.

Abstract

Objective: Therapeutic hypothermia is being clinically used to reduce neurologic deficits after cardiac arrest (CA). Patients receiving hypothermia after CA receive a wide-array of medications. During hypothermia, drug metabolism is markedly reduced. Little, however, is known about the impact of hypothermia on drug metabolism after rewarming. The objective of this study was to examine the effect of CA and hypothermia on the functional regulation of two major drug metabolizing cytochrome P450 (CYP) isoforms.

Design: Laboratory investigation.

Setting: University pharmacy school and animal research facility.

Subjects: Thirty-six male Sprague-Dawley rats.

Interventions: Hypothermia was induced via surface cooling in a rat CA model and maintained for 3 hrs. Animals were killed at 5 or 24 hrs and liver was analyzed for hepatic activity and mRNA expression of CYP3A2 and CYP2E1. Plasma interleukin-6 (IL-6) concentrations were determined. The effect of IL-6 on pregnane X receptor-mediated transcription of the rat CYP3A2 promoter was evaluated via luciferase reporter in HepG2 cells.

Measurements and main results: At 24 hrs after CA a decrease in CYP3A2 and CYP2E1 activity was observed, 55.7% +/- 12.8% and 46.8% +/- 29.7% of control, respectively (p < 0.01). CA decreased CYP3A2 mRNA (p < 0.05), but not CYP2E1 mRNA. Expression of other pregnane X receptor target enzymes and transporter genes were similarly down-regulated. CA also produced an approximately ten-fold increase in plasma IL-6. CA-mediated inhibition of CYP3A2 and CYP2E1 was attenuated by hypothermia, as was the increase in IL-6. Furthermore, IL-6 attenuated pregnane X receptor-mediated transcription of the CYP3A2 promoter.

Conclusions: CA produces CYP3A2 down-regulation at 24 hrs, potentially via IL-6 effects on pregnane X receptor-mediated transcription. Also, hypothermia attenuates the CA-mediated down-regulation, thereby normalizing drug metabolism after rewarming.

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Figures

Figure 1
Figure 1
Time-course of rectal temperature beginning 15 min after asphyxial arrest, and ending after 3 hrs of moderate hypothermia plus 1 hr of re-warming. Control and Cardiac Arrest Normothermia rats were maintained at 37°C, and CA Hypothermia rats were maintained at 30°C for 3 hrs followed by 1 hr of re-warming.
Figure 2
Figure 2
6β-hydroxytestosterone formation rate as a measure of CYP3A2 functional activity in liver microsomes from control, CA normothermia, and CA hypothermia treated rats. Panel A shows 6β-hydroxytestosterone formation rate in rats sacrificed at 5 h after injury (1 hr after re-warming). No differences were observed between groups. Panel B shows 6β-hydroxytestosterone formation rate in rats sacrificed at 24 h after injury (20 h after re-warming). A significant decrease in 6β-hydroxytestosterone formation rate in CA normothermia rats was observed compared to both control and CA hypothermia;‡p<0.05.
Figure 3
Figure 3
CYP3A2 mRNA hepatic expression in control, CA normothermia, and CA hypothermia rat liver. Panel A shows CYP3A2 mRNA expression at 5 h after injury. Panel B shows no difference in band density between all 3 groups(n=6). Panel C shows CYP3A2 mRNA expression at 24h after injury. Panel D shows a significant decrease in CYP3A2 relative hepatic expression in the CA normothermia compared to control and CA hypothermia;*p<0.05.
Figure 4
Figure 4
6-hydroxychlorzoxazone formation rate as a measure of CYP2E1 functional activity in liver microsomes from control, CA normothermia, and CA hypothermia treated rats. Panel A shows 6-hydroxychlorzoxazone formation rate at 5 h after injury. No differences were observed between groups. Panel B shows 6-hydroxychlorzoxazone formation rate at 24 h after injury. Activity in CA normothermia was significantly decreased compared to both control and CA hypothermia; ‡p<0.05.
Figure 5
Figure 5
CYP2E1 mRNA hepatic expression in control, CA normothermia, and CA hypothermia rat liver. Panel A shows CYP2E1 mRNA expression at 5 h after injury. Panel B shows no difference between groups(n=6). Panel C shows CYP2E1 mRNA expression at 24 h after injury. Panel D shows CYP2E1 relative hepatic expression in the CA normothermia rats compared to control and CA hypothermia. No significant difference was observed between treatment groups.
Figure 6
Figure 6
Expression levels of PXR target genes were evaluated to determine the effect of CA and hypothermia on drug metabolism and detoxification pathways. Real-time PCR analyses of liver RNA levels were determined from control, CA normothermia, and CA hypothermia groups. The hepatic expression of Oatp2, CYP3A2, CYP 2C2, and UGT1A1 are presented as average with standard division from 4-6 rats per treatment. (*p<0.05,**p<0.01,***p<0.001).
Figure 7
Figure 7
Plasma IL-6 levels in control, CA normothermia, and CA hypothermia after injury. CA normothermia IL-6 plasma concentrations at 60, 70, and 90 min after injury were significantly increased as compared to control or CA hypothermia (***p<0.001). The CA mediated increase in plasma IL-6 was attenuated in the CA hypothermic animals.
Figure 8
Figure 8
Co-transfection of hPXR and pGL3-rCYP3A2 natural promoter treated by human IL6 in HepG2 cells showed suppression of PXR activation. HepG2 cells were cotransfected with 1.4kb natural promoter pGL3-rCYP3A2-Luc reporter and human PXR. Transfected cells were treated with indicated IL6 concentration and/or rifampicin 10mM for 24 h before luciferase assay. Results are shown as —fold induction over vehicle control, which are represented as mean ± standard deviation from hexaplicate assay.
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