Reproductive and developmental toxicity testing: Examination of the extended one-generation reproductive toxicity study guideline

AbstractAn important aspect of safety assessment of chemicals (industrial and agricultural chemicals and pharmaceuticals) is determining their potential reproductive and developmental toxicity. A number of guidelines have outlined a series of separate reproductive and developmental toxicity studies from fertilization through adulthood and in some cases to second generation. The Extended One-Generation Reproductive Toxicity Study (EOGRTS) is the most recent and comprehensive guideline in this series. EOGRTS design makes toxicity testing progressive, comprehensive, and efficient by assessing key endpoints across multiple life-stages at relevant doses using a minimum number of animals, combining studies/evaluations and proposing tiered-testing approaches based on outcomes. EOGRTS determines toxicity during preconception, development of embryo/fetus and newborn, adolescence, and adults, with specific emphasis on the nervous, immunological, and endocrine systems, EOGRTS also assesses maternal and paternal toxicity. However, EOGRTS guideline is complex, criteria for selecting doses is unclear, and monitoring systemic dose during the course of the study for better interpretation and human relevance is not clear. This paper discusses potential simplification of EOGRTS, suggests procedures for relevant dose selection and monitors systemic dose at multiple life-stages for better interpretation of data and human relevance

Response of a single \u27mega intramuscular dose\u27 of vitamin D on serum 25OHD and parathyroid hormone levels

Objective: To determine the changes produced in serum 25OHD and iPTH levels after 600,000 IU of injection cholecalciferol in volunteers. Study Design: Interventional study. Place and Duration of Study: Section of Chemical Pathology, Department of Pathology and Microbiology, the Aga Khan University Hospital, Karachi, from June 2009 - June 2010. Methodology: Volunteers of either gender aged 18-40 years with known 25OHD, calcium (Ca), creatinine (Cr) and phosphorous (P) levels were included in the study. Subjects on therapy like vitamin D and calcium supplements, corticosteroids or anti-epileptic medicines, primary hyperparathyroidism and hypercalcaemia, with co-morbidity like renal failure, liver disease and history of malabsorption, diarrhea or hyperthyroidism were excluded. All volunteers were given an intramuscular injection of vitamin D3 (cholecalciferol, 600,000 IU). After 8 weeks, serum 25OHD, iPTH, Ca and P levels were determined again. For 25OHD level, cut-off of ≤ 50 nmol/l was defined as deficient, 50-75 nmol/l as insufficient and ≥ 75 as optimal level. Results: Mean 25OHD and iPTH levels were 35.06 ± 16.6 nmol/l and 81.15 ± 76.78 pg/ml respectively at baseline. Seventeen volunteers were 25OHD deficient. Five had high iPTH levels (25%) (mean 156 ± 123.7 pg/ml). 25OHD and iPTH showed a significant inverse correlation at baseline (\u3c 0.01). After 8 weeks of injection vitamin D 25OHD levels became optimal in 6 subjects (35%) [mean 92.9 ± 16.6 nmol/l]. It remained low in 5 volunteers (25%) [mean 41.6 ± 9.6 nmol/l] while insufficient levels were seen in 9 volunteers (40%) [mean 63.3±5.8 nmol/l]. Follow-up mean Ca, P and iPTH were 2.25 mmol/l (± 0.09), 1.1 (± 0.1) and 47.52 pg/ml (± 22.56) respectively. A significant increase in mean 25OHD level was seen at follow-up (p \u3c 0.01), while the change in PTH was insignificant (p=0.05). Conclusion: Single mega-dose of cholecalciferol achieved optimal levels of 25OHD in 35% of subjects after eight weeks of supplementation

Kinetics of Trihalogenated Acetic Acid Metabolism and Isoform Specificity in Liver Microsomes

This study determined the metabolism of 3 drinking water disinfection by-products (halogenated acetic acids [HAAs]), bromodichloroacetic acid (BDCAA), chlorodibromoacetic acid (CDBAA), and tribromoacetic acid (TBAA), using rat, mouse, human liver microsomes, and recombinant P450. Metabolism proceeded by reductive debromination forming a di-HAA, the highest under nitrogen \u3e\u3e2% oxygen \u3e atmospheric headspaces. V(max) for the loss of tri-HAA was 4 to 5 times higher under nitrogen than atmospheric headspace. Intrinsic metabolic clearance was TBAA\u3eCDBAA\u3e\u3eBDCAA. At the high substrate concentrations, tri-HAA consumption rate was 2 to 3 times higher than the formation of di-HAA. Liberation of Br(-) from TBAA corresponded to the expected amount produced after DBAA formation, indicating retention of Br(-) by additional metabolite/metabolites. Subsequent experiments with CDBAA detected negligible formation of chlorodibromomethane (CDBM) and failed to account for the missing tri-HAA. Carbon monoxide and especially diphenyleneiodonium ([DPI] P450 reductase inhibitor) blocked CDBAA metabolism. Other chemical inhibitors were only partially able to block CDBAA metabolism. Most effective were inhibitors of CYP 2E1 and CYP 3A4. Immunoinhibition studies using human liver microsomes and anti-human CYP 2E1 antibodies were successful in reducing CDBAA metabolism. However, CDBAA metabolism in wild-type (WT) and CYP 2E1 knockout (KO) mouse liver microsomes was similar, suggesting significant interspecies differences in CYP isoform in tri-HAA metabolism. Additional assessment of CYP isoform involvement was complicated by the finding that recombinantly expressed rat and human P450 reductase was able to metabolize CDBAA, which may be a contributing factor in interspecies differences in tri-HAA metabolism

Rethinking toxicity testing: Influence of aging on the outcome of long-term toxicity testing and possible remediation

Traditionally, toxicity testing is conducted at fixed dose rates (i.e., mg/kg/day) without considering life-changing events, e.g., stress, sickness, aging- and/or pregnancy-related changes in physical, physiological and biochemical parameters. In humans, life-changing events may cause systemic dose non-proportionality requiring modulation of drug dosage; similar changes occur in animals altering systemic dose during chronic/carcinogenic testing leading to late-occurring effects in some studies. For example, propylene monomethyl ether, an industrial chemical, initially induced sedation in rats and mice with recovery upon induction of hepatic CYPs after ~1 week. Sedation reappeared in rats but not in mice after ~12 months of exposure due to decreased CYP activity in rats, elderly mice were able to maintain slightly higher CYP activity avoiding recurrence of sedation. The systemic dose of two pharmaceuticals (doxazosin and brimonidine tartrate) increased up to 6-fold in ≥12-month old rats with no toxicity. In a rat reproductive toxicity study, systemic dose of 2,4-D, an herbicide, rapidly increased due to increased consumption of 2,4-D-fortified diet during pregnancy, lactation and neonatal growth, requiring adjustment to maintain the targeted systemic dose. Ideally, toxicological studies should be based on systemic dose with the option of modulating external dose rates to maintain the targeted systemic dose. Systemic dose can easily be monitored in selected core study animals at desired intervals considering recent developments in sampling and analysis at a fraction of the overall cost of a study

Role of environmental toxicants in the development of hypertensive and cardiovascular diseases

The incidence of hypertension with diabetes mellitus (DM) as a co-morbid condition is on the rise worldwide. In 2000, an estimated 972 million adults had hypertension, which is predicted to grow to 1.56 billion by 2025. Hypertension often leads to diabetes mellitus that strongly puts the patients at an increased risk of cardiovascular, kidney, and/or atherosclerotic diseases. Hypertension has been identified as a major risk factor for the development of diabetes; patients with hypertension are at two-to-three-fold higher risk of developing diabetes than patients with normal blood pressure (BP). Causes for the increase in hypertension and diabetes are not well understood, environmental factors (e.g., exposure to environmental toxicants like heavy metals, organic solvents, pesticides, alcohol, and urban lifestyle) have been postulated as one of the reasons contributing to hypertension and cardiovascular diseases (CVD). The mechanism of action(s) of these toxicants in developing hypertension and CVDs is not well defined. Research studies have linked hypertension with the chronic consumption of alcohol and exposure to metals like lead, mercury, and arsenic have also been linked to hypertension and CVD. Workers chronically exposed to styrene have a higher incidence of CVD. Recent studies have demonstrated that exposure to particulate matter (PM) in diesel exhaust and urban air contributes to increased CVD and mortality. In this review, we have imparted the role of environmental toxicants such as heavy metals, organic pollutants, PM, alcohol, and some drugs in hypertension and CVD along with possible mechanisms and limitations in extrapolating animal data to human