Background: Cytochrome P450 (CYP450) activity is an important indicator of liver graft function. CYP450 activity is altered by pro-inflammatory cytokines, which are associated with ischemia-reperfusion (I/R) injury during orthotopic liver transplantation (OLT). Treprostinil, an FDA-approved prostacyclin analog, ameliorated cold I/R injury during rat OLT. We hypothesized that treprostinil would improve CYP450 activity in liver graft during cold I/R injury post-OLT.
Methods: OLT was performed in syngeneic male Lewis rats with 18 h graft preservation in cold UW solution. Donor and recipients received treprostinil (100 ng/kg/min) or matching placebo for 24 h before and up to 48 h post-OLT. Liver graft mRNA and protein expression of CYP450 isoforms were analyzed by qRT-PCR and Western blot analysis, respectively. The formation rates of 1-hydroxymidazolam and 6β-hydroxytestosterone, 6-hydroxychlorzoxazone, 2α- and 16α-hydroxytestosterone in liver graft microsomes served as markers for CYP3A, CYP2E1, and CYP2C11 activity, respectively, and were measured by LC-MS.
Results: Treprostinil significantly decreased serum ALT and AST levels at 6-48 h after OLT, compared to placebo. The expressions of TNFα and IFNγ mRNA in the liver graft were significantly inhibited in the treprostinil-treated group at 1 h post-reperfusion. Treprostinil restored CYP2E1 protein expression to that of normal liver and significantly improved CYP3A activity to more than two-fold of placebo early post-OLT.
Conclusions: Treprostinil significantly ameliorated hepatic injury, reduced expression of pro-inflammatory cytokines, and improved CYP450 activity in liver graft early post-OLT. These findings suggest that treprostinil has the potential to serve as a therapeutic option to protect liver graft function against I/R injury during clinical OLT.
Keywords: 1-OH MDZ, 1-hydroxymidazolam; 16α-OH TST, 16α-hydroxytestosterone; 2α-OH TST, 2α-hydroxytestosterone; 6-OH CZN, 6-hydroxychlorzoxazone; 6β-OH TST, 6β-hydroxytestosterone; ALT, alanine aminotransferase; AST, aspartate aminotransferase; AUC, area under the time-concentration curves; CYP450, cytochrome P450; CZN, chlorzoxazone; HPLC-mass spectrometry; I/R, ischemia-reperfusion; IFN-γ, interferon gamma; IL, interleukin; Ischemia-reperfusion injury; MDZ, midazolam; NF-κB, nuclear factor-kappa B; NL, normal liver; OLT, orthotopic liver transplantation; PG, prostaglandin; PGI2, prostacyclin; TNF-α, tumor necrosis factor alpha; TST, testosterone; UW, University of Wisconsin; cytokines; drug metabolism; mRNA, messenger RNA; prostacyclin.