Development of tolerance to 2-butoxyethanol-induced hemolytic anemia and studies to elucidate the underlying mechanisms.

Early work demonstrated that a single administration of 2-butoxyethanol (BE) causes acute hemolytic anemia in rats. Current studies were undertaken to investigate the effect of repetitive daily dosing of BE on the hematologic parameters of male F344 rats. Treatment of rats with BE daily (125 mg/kg/day) for 1 to 3 consecutive days resulted in a time-dependent increase in the hemolysis of erythrocytes. However, when daily treatment with BE continued beyond 3 days, the number of erythrocytes began to rebound and approached pretreatment levels within 12 days despite continued daily exposure, suggesting development of tolerance to the hemolytic effect of BE. In vivo and in vitro studies were designed to investigate the underlying mechanism(s) of tolerance to the hematotoxicity of BE. Rats were treated with 125 mg BE/kg/day for 3 days followed by a 7-day recovery. At the end of this recovery period, rats were challenged with a single 125 or 250 mg BE/kg dose and the hematologic profiles were assessed at 2, 8, and 24 hr later. A significant decline in the sensitivity of BE-pretreated/recovered rats compared to vehicle-pretreated rats was observed. Further, in vitro incubation of blood obtained from BE-pretreated/recovered with the hematotoxic metabolite of BE, 2-butoxyacetic acid (BAA), revealed that erythrocytes obtained from these rats were significantly less sensitive to BAA than those obtained from normal rats. These studies suggested that tolerance is due, at least in part, to the lesser sensitivity of young erythrocytes formed during the regeneration process. In another study, rats were rendered anemic by bleeding followed by a 7-day recovery. BE administration to bled/recovered rats demonstrated that these rats were less sensitive than rats which were not subjected to bleeding. In vitro incubation of blood obtained from the bled/recovered animals with BAA demonstrated that erythrocytes were significantly less sensitive to BAA than those obtained from control rats. This further confirmed that young erythrocytes, formed during the regeneration process, were less sensitive to BAA than older erythrocytes. Current data also suggested that it is unlikely that tolerance is caused by modification of BE metabolism in rats repetitively exposed to this chemical. In conclusion, chronic exposure to BE would be expected to result in tolerance to BE-induced hemolytic anemia. The mechanisms responsible are likely related to the fact that older cells are more susceptible to BE and BAA and that hemolysis of these cells during the initial exposure followed by their replacement with less susceptible younger cells may account for tolerance development.