Hepatic Cytochrome P450 Profiles in Hemorrhagic Shock Model Rats After Transfusion With Stored Red Blood Cells

Abstract

Red cell transfusions, which deteriorate in quality during storage, triggers several negative biological responses. However, little is known regarding the effects of stored red cell transfusion on cytochrome P450 (P450) profiles. To clarify this issue, we investigated hepatic P450 profiles in hemorrhagic shock model rats after resuscitation with stored packed red cells (PRC). The pharmacokinetics data for P450-metabolizing substrates showed that the clearance of substrates for Cyp1A2 and Cyp3A2 in the stored PRC resuscitation group were decreased compared to sham group. The protein expression, metabolic activity and mRNA expression of the P450 isoforms in the stored PRC resuscitation group were lower than the corresponding values for the sham group. However, these changes would be expected to have weak effects on the in vivo pharmacokinetics of the concomitant drugs based on the criteria stated in the guideline on drug interactions. In contrast, the results of these P450 profiles in the stored PRC and fresh PRC resuscitation group exhibited a similar trend. These results suggest that the stored PRC transfusion has an influence on the hepatic P450 profiles, but is of little clinical significance, not by the deterioration of the quality of red cells but pathophysiological alterations following the hemorrhage and transfusion.

Introduction

The cytochrome P450 (P450) family is a clinically pivotal drug-metabolizing class of enzymes due to fact that half of the top 200 clinically available prescription drugs are mainly metabolized by these enzymes.¹ Among the numerous P450 isoforms, CYP1A, CYP2C, and CYP3A account for over 70% of drugs that are metabolized by the P450 families.1, 2, 3 Therefore, metabolic changes in these P450 isoforms by medications and pathology are attributed to some of the clinical problems associated with the administration of P450-metabolizing drugs, such as unexpected adverse events and insufficient curative effects. In previous studies, Harbrecht et al., reported that the urinary excretion of P450s substrates (mephenytoin, chlorzoxazone, flurbiprofen and dapsone) in severely injured patients who had received transfusions of red cells were increased/decreased compared to that of healthy subjects, indicating that the in vivo P450 metabolic activity in patients with massive hemorrhage was altered by red cell transfusions.⁴ In addition, our group demonstrated that the protein expression of Cyp3A2 in the liver was decreased in hemorrhagic shock model rats that had been transfused with red cells compared to sham rats, resulting in an increase in the blood retention and the prolonged pharmacological effect of midazolam, a substrate for Cyp3A.⁵ These facts indicate that transfusion therapy with red cells is an influential factor for massive hemorrhage patients, in that the profiles for drugs that are metabolized by P450 had been changed. Since patients with massive hemorrhages frequently receive P450-metabolizing drugs to prevent or treat complications resulting from red cell transfusions, accumulating additional evidence regarding P450 profiles after red cell transfusion is of great significance in terms of implementing transfusion therapy with a safe and effective outcome.

In general, suspensions of red cells are stored under refrigerated conditions for periods of up to 3–7 weeks (maximum storage period depends on the country where they are being used) and are distributed from blood centers in a first-in-first-out manner. It has been reported that the deterioration of red cell quality, which is referred to as red cell lesions, progresses with increasing storage time.^6,7 Many clinical trials have shown that these pathophysiological alterations of red cells that are caused by long-term storage are not associated with clinical outcomes based on the results that no significant difference in mortality was observed between transfusion with fresh and stored red cells.8, 9, 10 It should also be noted however that oxidative damage and metabolic impairments by the red cell storage lesions have been reported,⁷ indicating that transfusions with stored red cells can have an impact on a biological system. Therefore, it is possible that the biological responses of P450s after a transfusion with stored red cells are different from that of fresh red cells. However, comparisons in the qualitative and quantitative alterations of P450s when transfusing fresh red cells versus stored red cells are not available.

The aim of this study was to investigate the effect of stored red cell transfusions on P450 profiles in hemorrhagic shock model rats. To achieve this, we prepared packed rat red cells (PRC) that had been stored for 7 days at 4 °C. This storage period for rat red cells is roughly equivalent to human red cells that had been stored for 28 days.¹¹ We then compared the pharmacokinetics of P450-metabolizing substrates, P450 mRNA expression, P450 protein expression and P450 activity, with a focus on Cyp1A2, Cyp2C11 and Cyp3A2, which are homologized to human CYP1A2, CYP2C9 and CYP3A4, respectively, in hemorrhagic shock model rats that had been resuscitated with stored PRC with sham rats and hemorrhagic shock rats that had received transfusions with fresh PRC (no storage period).