The long-term objective of this proposal is to uncover mechanisms responsible for the development of idiosyncratic drug-induced acute liver failure (Id-ALF) in susceptible patients. Id-ALF is the third most common cause of acute liver failure in the United States. Susceptible individuals develop anesthetic Id-ALF following halogenated volatile anesthetics exposure. Serum cytochrome P4502E1 (CYP2E1) autoantibodies detected in anesthetic Id-ALF patients may suggest that self CYP2E1 becomes altered forming hapten- autoantigen complexes capable of eliciting allergic or autoimmune responses. Anesthesiologists exposed to halogenated anesthetics, develop distinct CYP2E1 immunoglobulin subclass-specific autoantibodies from anesthetic Id-ALF patients, suggesting roles for distinct CYP2E1 epitopes in Id-ALF. We have developed an experimental model of immune-mediated drug-induced hepatitis resembling Id-ALF. Hepatitis is induced by immunizing BALB/c mice with a drug hapten-autoantigen, trifluoroacetyl chloride-hepatic S100 (TFA-S100). Splenocytes from TFA-S100-immunized mice proliferate in response to TFA as well as human CYP2E1, and adoptively transfer hepatitis to naive mice suggesting roles for CYP2E1 (self) and TFA (hapten) in Id-ALF. We hypothesize that in experimental, murine, drug-induced hepatitis (1) hepatitis is primarily caused by T cell reactions to critical epitopes of the Id-ALF associated self protein, CYP2E1. (2) hepatitis is also caused by T cell-mediated reactions to other Id-ALF-associated self proteins, such as, 58 kilodalton endoplasmic reticulum protein (ERp58), or autoimmune hepatitis-associated self protein, asialoglycoprotein receptor (ASGPR), (3) autoimmune responses to CYP2E1 induce autoimmune responses to ERp58 and ASGPR.
The specific aims are: (1) a) To identify critical epitopes of CYP2E1 in experimental drug-induced hepatitis using TFA-S100- and TFA-CYP2E1-immunized BALB/c splenocyte proliferative responses to candidate CYP2E1 epitopes, b) to verify critical CYP2E1 epitopes by inducing hepatitis in BALB/c mice by immunization with identified CYP2E1 epitopes haptenated with TFA and c) to verify serum autoantibody responses to TFA-haptenated CYP2E1 epitopes using anesthetic Id-ALF patients and anesthetic-exposed anesthesiologists. (2) To discover principal hepatic self proteins in murine experimental drug-induced hepatitis using splenocyte proliferation assays stimulated with ERp58 and ASGPR self proteins, FACS analysis and adoptive transfer of stimulated cells, and supernatant cytokines. (3) To confirm antigenic spread in TFA-CYP2E1-immunized mice using spleenocyte proliferation assays stimulated with ERp58 and ASGPR and analyzed as in Specific Aim 2. ? ? ? ?
