Comparative effects of human-equivalent low, moderate, and high dose oral prednisone intake on autoimmunity and glucocorticoid-related toxicity in a murine model of environmental-triggered lupus.

Autoimmune diseases can be triggered by environmental toxicants such as crystalline silica dust (cSiO2). Here, we characterized the dose-dependent immunomodulation and toxicity of the glucocorticoid (GC) prednisone in a preclinical model that emulates onset and progression of cSiO2-triggered lupus. Two cohorts of 6-wk-old female NZBWF1 mice were fed either control AIN-93G diet or one of three AIN-93G diets containing prednisone at 5, 15, or 50 mg/kg diet which span human equivalent oral doses (HED) currently considered to be low (PL; 5 mg/d HED), moderate (PM; 14 mg/d HED), or high (PH; 46 mg/d HED), respectively. At 8 wk of age, mice were intranasally instilled with either saline vehicle or 1 mg cSiO2 once weekly for 4 wk. The experimental plan was to 1) terminate one cohort of mice (n=8/group) 14 wk after the last cSiO2 instillation for pathology and autoimmunity assessment and 2) to maintain a second cohort (n=9/group) to monitor glomerulonephritis development and survival. Mean blood concentrations of prednisone's principal active metabolite, prednisolone, in mice fed PL, PM, and PH diets were 27, 105, 151 ng/ml, respectively, which are consistent with levels observed in human blood ≤ 12 h after single bolus treatments with equivalent prednisone doses. Results from the first cohort revealed that consumption of PM, but not PL diet, significantly reduced cSiO2-induced pulmonary ectopic lymphoid structure formation, nuclear-specific AAb production, inflammation/autoimmune gene expression in the lung and kidney, splenomegaly, and glomerulonephritis in the kidney. Relative to GC-associated toxicity, PM diet, but not PL diet, elicited muscle wasting, but these diets did not affect bone density or cause glucosuria. Importantly, neither PM nor PL diet improved latency of cSiO2-accelerated death. PH-fed mice in both cohorts displayed robust GC-associated toxicity including body weight loss, reduced muscle mass, and extensive glucosuria 7 wk after the final cSiO2 instillation requiring their early removal from the study. Taken together, our results demonstrate that while moderate doses of prednisone can reduce important pathological endpoints of cSiO2-induced autoimmunity in lupus-prone mice, such as upstream ectopic lymphoid structure formation, these ameliorative effects come with unwanted GC toxicity, and, crucially, none of these three doses extended survival time.

Single oral or intravenous administration of voriconazole achieved recommended therapeutic minimum inhibitory concentrations against Aspergillus in the common raven (Corvus corax).

OBJECTIVE: To determine the pharmacokinetics of voriconazole after single IV or orally administered boluses in common ravens (Corvus corax). ANIMALS: 8 healthy common ravens. PROCEDURES: Voriconazole (5 mg/mL, 10 mg/kg IV) was administered to 8 birds, and then plasma voriconazole concentrations were measured at various time points by high-pressure liquid chromatography with mass spectrometry. Starting 6 months later in a randomized 3-treatment 3-period regimen, birds received a single oral dose of voriconazole suspension (10 mg/mL; 6, 12, and 24 mg/kg PO). The study period was May 2015 to March 2016. RESULTS: Voriconazole (10 mg/kg IV) achieved an initial plasma concentration of 6.31 µg/mL when measured over 21 hours. After oral administration of voriconazole at 6, 12, and 24 mg/kg, the relative bioavailability was 67.5%, 209%, and 183%, respectively. For the 6-mg/kg dose, the maximum plasma concentration was reached at 30 minutes after administration and remained in the therapeutic range of 0.5 to 1 µg/mL for approximately 15 hours. The 12- and 24-mg/kg doses resulted in concentrations in a potentially toxic range. CLINICAL RELEVANCE: Voriconazole was well tolerated. All 4 doses resulted in plasma concentrations of voriconazole > 0.5 µg/mL, which is the minimum inhibitory concentration recommended for pathogenic species of Aspergillus fungi known to affect birds. A single dose of voriconazole administered as 10 mg/kg IV or 6 mg/kg PO resulted in recommended target plasma concentrations. Administration of voriconazole 6 mg/kg PO 2 to 3 times daily may be adequate for treatment without exceeding the toxic range.

Liquid chromatography/tandem mass spectrometric analysis of penicillamine for its pharmacokinetic evaluation in dogs.

Copper storage disease occurs in multiple dog breeds and is one of the most common causes of chronic hepatitis in this species. The disease is caused by hereditary defects in copper metabolism in conjunction with high dietary copper levels. The progressive copper accumulation leads to hepatitis, cirrhosis, and eventually death if left untreated. Copper chelators are critical in modulating the effects of this disease. It is therefore of significant practicality to understand the pharmacokinetic (PK) parameters of chelating agents, particularly since they are oftentimes quite expensive. A liquid chromatography-tandem mass spectrometric (LC/MS/MS) method was developed to measure plasma levels of one of the most common chelators, d-penicillamine. The compound was discovered to exist in two forms, monomeric and dimeric, and various chemical derivatizations were tried to force the compound into one form or the other. Eventually, the simplest approach was individual determination of penicillamine and its dimer, with summation of the two quantities. This enabled determination of canine PK parameters for penicillamine based on comparison of oral and intravenous administration of the drug, including time to maximum drug level (Tmax), concentration at maximum (Cmax), clearance (Cls) and volume of distribution (Vdss). The drug was found to exist predominantly in the dimeric form in plasma, which is incapable of chelating copper owing to lack of free sulfhydryl groups and must therefore provide a storage form of the drug in equilibrium with its monomeric form in vivo. Mechanisms are discussed for the electrospray-induced fragmentation of penicillamine as well as of its dimer.

Hepatic copper and other trace mineral concentrations in dogs with hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common primary liver tumor in dogs. Abnormalities in hepatic copper, iron, zinc, and selenium concentrations increase risk for HCC development in other species, but trace mineral concentrations have not been evaluated in dogs with HCC. OBJECTIVES: To investigate hepatic trace mineral concentrations in dogs with HCC. ANIMALS: Archived liver specimens from 85 dogs with HCC and 85 control dogs. METHODS: Retrospective case-control study. A histopathology database was searched to identify dogs with HCC (test population) and an age-matched control population. Demographic information was retrieved, and H&E and rhodanine stained slides were reviewed for all cases. Copper, iron, zinc, and selenium concentrations were determined in noncancerous liver tissues (test and control population) and in HCC tissues (test population) using inductively coupled plasma mass spectrometry. RESULTS: Hepatic copper concentrations (non-neoplastic hepatic tissue) were greater in test population dogs (median, IQR; 294.9 µg/g, 233.5-475.9 µg/g) than in control dogs (202.8 µg/g, 135.0-295.3 µg/g; P < .001). Hepatic zinc concentrations in test (132.1 µg/g,108.6-163.2 µg/g) and control dogs (151.5 µg/g, 117.1-184.5 µg/g) also were different (P = .03). Within test population dogs, all trace mineral concentrations were decreased in the HCC tissue as compared to the non-neoplastic hepatic tissue (all P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE: Hepatic copper accumulation and other abnormalities in hepatic trace mineral concentrations could be involved in the pathogenesis of HCC in some dogs.

Evidence of accumulation and elimination of inorganic contaminants from the lachrymal salt glands of leatherback sea turtles (Dermochelys coriacea).

Plasma osmolalities of marine vertebrates are generally lower than the surrounding medium; therefore, marine organisms must cope with the osmoregulatory challenges of life in a salty environment. The salt glands serve to maintain osmotic and ionic homeostasis in a number of lower marine vertebrates. One marine reptile, the leatherback sea turtle (Dermochelys coriacea), ingests excessive amounts of salts due to their diet of gelatinous zooplankton. Outside of the normal osmoregulatory function of the salt gland, little research has been conducted on contaminant accumulation and excretion in this organ. Here, we established arsenic, cadmium, lead, mercury, and selenium concentrations in red blood cells (RBCs) and salt gland secretions (SGSs) of nesting leatherbacks. We also collected salt glands from different life stage classes of dead stranded leatherbacks from the western Atlantic Ocean to determine if inorganic contaminants accumulate in this organ. Using non-metric multidimensional scaling and regression analyses, we determined that RBC and SGS inorganic contaminant concentrations were not correlated. Additionally, RBCs showed significantly higher concentrations of these contaminants in comparison to SGSs, likely due to the affinity of inorganic contaminants for the heme group of RBCs. Lastly, we found that salt gland cadmium and mercury concentrations tended to increase with increasing curved carapace length (CCL) in stranded leatherbacks. Our results indicate that different physiological mechanisms determine the distribution of inorganic contaminants in blood and SGSs. Increases in salt gland contaminant concentrations with increasing CCL suggest this organ as a potential target for accumulation.

Toxic elements and associations with hematology, plasma biochemistry, and protein electrophoresis in nesting loggerhead sea turtles (Caretta caretta) from Casey Key, Florida.

Toxic elements (arsenic, cadmium, lead, mercury, selenium, thallium) are a group of contaminants that are known to elicit developmental, reproductive, general health, and immune system effects in reptiles, even at low concentrations. Reptiles, including marine turtles, are susceptible to accumulation of toxic elements due to their long life span, low metabolic rate, and highly efficient conversion of prey into biomass. The objectives of this study were to (1) document concentrations of arsenic, cadmium, lead, mercury, selenium, and thallium in whole blood and keratin from nesting loggerhead sea turtles (Caretta caretta) from Casey Key, Florida and document correlations thereof and (2) correlate whole blood toxic element concentrations to various hematological and plasma biochemistry analytes. Baselines for various hematological and plasma analytes and toxic elements in whole blood and keratin (i.e., scute) in nesting loggerheads are documented. Various correlations between the toxic elements and hematological and plasma biochemistry analytes were identified; however, the most intriguing were negative correlations between arsenic, cadmium, lead, and selenium with and α- and γ-globulins. Although various extrinsic and intrinsic variables such as dietary and feeding changes in nesting loggerheads need to be considered, this finding may suggest a link to altered humoral immunity. This study documents a suite of health variables of nesting loggerheads in correlation to contaminants and identifies the potential of toxic elements to impact the overall health of nesting turtles, thus presenting important implications for the conservation and management of this species.