Impact of Repeated Tail Clip and Saphenous Vein Phlebotomy on Rats Used in Toxicology Studies.

Sampling blood for toxicokinetic (TK) evaluation in rodents is typically performed using a satellite group of animals to avoid depleting the blood volume and inducing an additional stressor in the main study animals. This practice does not allow for direct comparison of individual animal toxicity to exposure. These studies evaluated serial collection of twelve, 40-µl blood samples from each rat from either a tail clip or a saphenous vein bleed and its impact on toxicologic parameters over 4- and 14-day periods. The results show the feasibility of successfully collecting TK samples from main study animals, using either of the two techniques. Both procedures were amenable to execution by a single technician using dried blood spot sampling. Any changes observed in the primary markers of erythroid mass between the nonbled control rats and repeat sampled rats were minimal and the range of values often overlapped. This technique would improve the quality of data generated from toxicology studies by allowing a direct comparison of systemic exposure to toxicity while at the same time reducing the number of rats by obviating the need for satellite groups.

Minor compensatory changes in SAGE Mdr1a (P-gp), Bcrp, and Mrp2 knockout rats do not detract from their utility in the study of transporter-mediated pharmacokinetics.

Mdr1a-, Bcrp-, and Mrp2-knockout rats are a more practical species for absorption, distribution, metabolism, and excretion (ADME) studies than murine models and previously demonstrated expected alterations in the pharmacokinetics of various probe substrates. At present, gene expression and pathology changes were systematically studied in the small intestine, liver, kidney, and brain tissue from male SAGE Mdr1a, Bcrp, and Mrp2 knockout rats versus wild-type Sprague-Dawley controls. Gene expression data supported the relevant knockout genotype. As expected, Mrp2 knockout rats were hyperbilirubinemic and exhibited upregulation of hepatic Mrp3. Overall, few alterations were observed within 112 ADME-relevant genes. The two potentially most consequential changes were upregulation of intestinal carboxylesterase in Mdr1a knockouts and catechol-O-methyltransferase in all tissues of Bcrp knockout rats. Previously reported upregulation of hepatic Mdr1b P-glycoprotein in proprietary Wistar Mdr1a knockout rats was not observed in the SAGE counterpart investigated herein. Relative liver and kidney weights were 22-53% higher in all three knockouts, with microscopic increases in hepatocyte size in Mdr1a and Mrp2 knockout rats and glomerular size in Bcrp and Mrp2 knockouts. Increased relative weight of clearing organs is quantitatively consistent with reported increases in the clearance of drugs that are not substrates of the knocked-out transporter. Overall, SAGE knockout rats demonstrated modest compensatory changes, which do not preclude their general application to study transporter-mediated pharmacokinetics. However, until future studies elucidate the magnitude of functional change, caution is warranted in rare instances of extensive metabolism by catechol-O-methyltransferase in Bcrp knockouts and intestinal carboxylesterase in Mdr1a knockout rats, specifically for molecules with free catechol groups and esters subject to gut-wall hydrolysis.

The inhibin B (InhB) response to the testicular toxicants mono-2-ethylhexyl phthalate (MEHP), 1,3 dinitrobenzene (DNB), or carbendazim (CBZ) following short-term repeat dosing in the male rat.

OBJECTIVE: The objective of this study was to evaluate the utility of plasma Inhibin B (InhB) as a biomarker of testicular injury in adult rats following short-term exposure to the known Sertoli cell toxicants mono-2-ethylhexyl phthalate (MEHP), 1,3 dinitrobenzene (DNB), or carbendazim (CBZ). METHODS: Following oral gavage administration of the compounds for 2 or 7 days, the rats were evaluated for clinical signs, body weight, food consumption, organ weights, plasma hormone levels, and gross and microscopic pathology. RESULTS: MEHP, DNB, and CBZ produced a range of testicular toxicity characterized by minimal exfoliation of germ cells as demonstrated by increased cellular debris in the epididymis (MEHP) to more severe and dose/duration responsive Sertoli cell vacuolation, germ cell degeneration, and multinucleated giant cells of germ cell origin (DNB and CBZ). The slight to moderate Sertoli and germinal cell injuries did not correlate with significant changes in plasma InhB levels following 2- or 7-day exposures. However, more severe injury to germinal epithelium following up to 7 days of exposure, but not after 2 days of exposure, correlated with decreased plasma InhB levels and less consistently with increases in plasma follicle stimulating hormone. CONCLUSION: In conclusion, under the conditions of these studies, changes in InhB were not an effective early onset marker of testicular toxicity or an effective marker for slight to moderate levels of acute injury, and only reflected more severe disruption of spermatogenesis. Changes in plasma InhB and follicle stimulating hormone were poorly correlated except in some instances of moderate to marked testicular toxicity.

Mandibular Blastomycosis in a 5-Year-Old Dog.

A case of localized oral mandibular blastomycosis is described in a five-year-old dog. Complete resolution of clinical signs and oral radiographic changes were seen following itraconazole therapy at 5 mg/kg/day for four and a half months. The patient remained free of Blastomyces at the one year follow up based on the Mira Vista Blastomyces urine antigen test by EIA (Enzyme Immunoassay)a. A literature review of localized blastomycosis cases in humans and dogs was performed, available diagnostic tests evaluated, and treatment comparisons made.

Non-Lethal Endotoxin Injection: A Rat Model of Hypercoagulability.

Systemic inflammation co-activates coagulation, which unchecked culminates in a lethal syndrome of multi-organ microvascular thrombosis known as disseminated intravascular coagulation (DIC). We studied an endotoxin-induced inflammatory state in rats to identify biomarkers of hemostatic imbalance favoring hypercoagulability. Intraperitoneal injection of LPS at 15 mg/kg body weight resulted in peripheral leukopenia and widespread neutrophilic sequestration characteristic of an acute systemic inflammatory response. Early indicators of hemostatic pathway activation developed within 4 hours, including increased circulating concentrations of procoagulant extracellular vesicles (EVs), EVs expressing endothelial cell and platelet membrane markers, and high concentration of soluble intercellular adhesion molecule-1 (sICAM-1), plasminogen activator inhibitor-1 (PAI-1), and D-dimers. Inflammation persisted throughout the 48-hour observation period; however, increases were found in a subset of serum microRNA (miRNA) that coincided with gradual resolution of hemostatic protein abnormalities and reduction in EV counts. Dose-adjusted LPS treatment in rats provides a time-course model to develop biomarker profiles reflecting procoagulant imbalance and rebalance under inflammatory conditions.

Plasmodium gallinaceum: clinical progression, recovery, and resistance to disease in chickens infected via mosquito bite.

Historically, in vivo experiments of Plasmodium gallinaceum in chickens have caused high mortality. Perhaps because of this high mortality, it remains to be demonstrated whether recovered birds will resist a second episode of illness when re-exposed to infected mosquitoes. In the current study, groups of 10 chicks were infected with P. gallinaceum via mosquito bite. Parasitemia and anemia were followed by recovery in all birds, although they had persisting, low levels of parasitized erythrocytes (0.007 +/- 0.019%). Twenty-three days after the initial exposure, 10 recovered chicks were rechallenged with P. gallinaceum via mosquito bite; none of them developed clinical or hematological evidence of malaria, in contrast to matched control birds, which all became diseased (P < 0.001). Unlike previous studies, the current experiment had no mortality in chickens infected with P. gallinaceum by mosquito bite. Recovered birds resisted disease from re-exposure to the same organism. The duration and nature of immunity or premunition remain to be determined.