Background:The recently discovered apolipoprotein A5 (ApoA5) is fast gaining attention as a key regulator of serum triglyceride concentrations. An ApoA5 mouse knock-out model produced an approximately fourfold increase in serum triglycerides, whereas a knock-in model with human ApoA5 produced 50 -70% lower concentrations of mouse serum triglycerides. In addition, peroxisome proliferator-activated receptor-␣ agonists, which are used clinically to lower serum triglyceride concentrations, cause increased ApoA5 mRNA expression. Despite these compelling molecular biology data, relatively little is known about ApoA5 protein in human serum. Methods: To better understand circulating concentrations and lipoprotein particle distribution of ApoA5, we expressed the recombinant human ApoA5 protein and raised antibodies against both the NH 2 and COOH termini. Results: Using the above reagents, we demonstrate for the first time that ApoA5 is present in human serum, although at much lower concentrations than other apolipoproteins such as ApoA1. Using a dual-antibody sandwich ELISA that we developed, we observed ApoA5 concentrations in human serum ranging from 24 to 406 g/L compared with ϳ1 g/L for ApoA1. We also

Monocrotaline (MCT) is a toxic pyrrolizidine alkaloid of plant origin. Administration of small doses of MCT or its active metabolite, monocrotaline pyrrole (MCTP), to rats causes delayed and progressive lung injury characterized by pulmonary vascular remodeling, pulmonary hypertension, and compensatory right heart hypertrophy. The lesions induced by MCT(P) administration in rats are similar to those observed in certain chronic pulmonary vascular diseases of people. This review begins with a synopsis of the hemostatic system, emphasizing the role of endothelium since endothelial cell dysfunction likely underlies the pathogenesis of MCT(P)-induced pneumotoxicity. MCT toxicology is discussed, focusing on morphologic, pulmonary mechanical, hemodynamic, and biochemical and molecular alterations that occur after toxicant exposure. Fibrin and platelet thrombosis of the pulmonary microvasculature occurs after administration of MCT(P) to rats, and several investigators have hypothesized that thrombi contribute to the lung injury and pulmonary hypertension. The evidence for involvement of the various components of the hemostatic system in MCT(P)-induced vascular injury and remodeling is reviewed. Current evidence is consistent with involvement of platelets and an altered fibrinolytic system, yet much remains to be learned about specific events and signals in the vascular pathogenesis.

Serum cardiac troponin-I (cTnI) has been validated as a biomarker for cardiotoxicity in numerous animal models; however, owing to sensitivity issues cTnI concentrations in healthy, resting animals used in toxicology studies have not been established. Serum from healthy and isoproterenol hydrochloride (iso)-treated rats, dogs, and monkeys were assayed using the Erenna system. The Erenna cTnI assay provided sensitivity < 1 ng/L across human, rat, dog, and monkey cTnI. Linear responses (R 2 = 0.99) were observed for all species. Precision studies yielded interassay CVs of curve fit quantification from 2% to 4% between 1.6 and 5000 ng/L, and 23% at 0.78 ng/L. Strong correlation (R 2 = 0.99) was obtained between Erenna and Beckman Access cTnI. Concentrations of cTnI in healthy animals ranged from 1 to 9 ng/L. In longitudinal studies of iso-treated animals, the concentrations of cTnI in the control vehicle-treated groups were 10-20 ng/L for rats (N = 10) and predose values of 2-3 ng/L for dogs (N = 3). Measured with the Erenna assay system, cTnI was quantifiable at all time intervals tested in all animals treated with iso. The Erenna system provides sensitive measurement of cTnI in rats, dogs, and monkeys, makes it possible to determine small changes from normal concentrations, and provides cTnI values from small volumes of serum.

ABSTRSCTSerum cardiac troponin-I has been validated as a biomarker for cardiotoxicity in numerous animal models; however, baseline reference ranges for cTnI concentration in a healthy population of laboratory rats, as well as an investigation of biological cTnI variability in rats with respect to time, handling, and placebo dosing methods, have not been reported. In this study, we used an ultrasensitive cTnI immunoassay to quantify hourly concentrations of cTnI in live rats handled under standard laboratory conditions using 15 mL of serum per determination. The baseline reference range (mean 4.94 pg/mL, range 1-15 pg/mL, 99% confidence interval [CI]) of cTnI concentration in rats was consistent with previously reported reference ranges for cTnI in humans (1-12 pg/mL) and with preliminary studies in dogs (1-4 pg/mL) and monkeys (4-5 pg/mL) using the same cTnI assay method. In addition, cTnI concentrations in individual rat serum samples show minimal biological variability over a twenty-four-hour interval when compared to a meaningful reference change value of 193% to 206%. Furthermore, measurements of cTnI concentration were consistent within the reference limits in individual rats over long periods and under three different standard laboratory handling conditions. Thus, using this new method, rats can be followed longitudinally at hourly intervals, and a doubling of cTnI concentration would be significant above biological variability. This is a new paradigm for preclinical testing, which allows transient changes in cTnI concentration to be accurately quantified. This understanding of baseline and biological variability in rats will be fundamental for designing and analyzing future studies that assess potential cardiotoxicity in drug development.