Impaired calcium mobilization and differential tyrosine phosphorylation in intestinal intraepithelial lymphocytes.

Intestinal intraepithelial lymphocytes (iIEL) exhibit a unique activation state characterized by the expression of activation markers and effector functions, but a minimal response to mitogenic signals in vitro. To further characterize this activation status, iIEL were compared with splenic T cells for two key activation signals, calcium mobilization and tyrosine phosphorylation. Calcium mobilization was impaired in iIEL treated with the calcium ionophores ionomycin or A23187, thapsigargin, or by CD3-cross-linking. The calcium mobilization defect is shared by mature and embryonic iIEL. Anti-phosphotyrosine Western blot analysis revealed that the iIEL are able to respond to T-cell receptor (TCR)-mediated signals by tyrosine phosphorylation, although the patterns of phosphorylation differ from those seen in splenic T cells. We conclude that iIEL are unable to mobilize calcium in vitro, which may be due to modulation of TCR-mediated signal transduction pathways by the microenvironment of the intestinal epithelium and/or caused by the standard isolation procedure used to prepare iIEL, which must be considered in future in vitro studies of iIEL function.

Acute experimentally induced aflatoxicosis in the weanling pony.

Nineteen weanling ponies and 1 adult pony were given a single oral dose of aflatoxin B1 (AFB1). Dosages were: 0, 0.5, 1, 2, 4, 5, 6, and 7.4 mg of AFB1/kg of body weight. Vital signs were monitored, and whole blood and serum collected for analysis of serum enzymes, prothrombin time, blood cell counts, and serum urea nitrogen. Ponies that died were examined for gross lesions, and tissues were collected for histopathologic examination and analysis of AFB1 and AFM1 residues. Two of the 4 ponies given the 2 mg/kg dose and all ponies given the larger dosages died within 76 hours. Clinical signs included increased rectal temperature, faster heart and respiratory rates, abdominal straining, bloody feces, and tetanic convulsions. At necropsy, ponies that died of acute aflatoxicosis showed visceral petechiae and hepatic focal lesions. Histopathologic changes included severe hepatic necrosis, vacuolation, and bile duct hyperplasia. Aflatoxins B1 and M1 were recovered from liver, kidney, skeletal muscle, and gastrointestinal contents. One other pony given the 2 mg/kg dose died 32 days after dosing, and 1 control pony died after 70 days. Continuous elevations in prothrombin time and serum aspartate aminotransferase, alanine aminotransferase, and gamma-glutamyl transpeptidase levels were observed in ponies dosed at 4 mg/kg or more. Significant (P less than 0.05) elevations in these values, which peaked 2 to 3 days after dosing, were seen in ponies given the 2 mg/kg dose. This group also had significant increases over controls in PCV and hemoglobin concentration 5 days after dosing.

Effects of aflatoxins in young ponies.

Sixteen clinically normal, healthy ponies were randomly assigned to 4 groups and given aflatoxin B1 in doses of 0.045, 0.030, 0.015, and 0 (control) mg/kg of body weight per day for 21 days (or total doses of 0.945, 0.630, 0.315, and 0 mg/kg). The animals were allowed to recover for 3 months and then were reassigned to 4 treatment groups such that each group during the 2nd trial included a pony from each of the groups of the 1st trial. The animals in the new groups were intubated and were given aflatoxin in doses of 0.4, 0.2, 0.1, and 0 (control) mg/kg/day for 5 days ( or total doses of 2.0, 1.0, 0.5, and 0 mg/kg). Venous blood samples were drawn every other day to monitor for toxicosis; examinations were made for RBC and WBC counts, hemoglobin concentration, PCV, serum urea nitrogen, prothrombin time, and serum concentrations of aspartate aminotransferase, iditol dehydrogenase, alkaline phosphatase, albumin, gamma-glutamyl transferase, and arginase. There were no significant differences between treatment groups and controls (given no aflatoxin) in the toxicologic values examined for during the 1st trial. During the 2nd experiment, 2 of the ponies in the large-dose treatment gorup (2.0 mg/kg) demonstrated increased serum enzyme activities. These animals had been in the large-dose (0.945 mg/kg) and median-dose (0.63 mg/kg) groups during the 1st trial. Arginase, iditol dehydrogenase, and gamma-glutamyl transpeptidase activities became increased on the 4th day of treatment and continued to increase until the 6th day of the experiment (1 day after treatment was terminated). These enzymes approached control group values at 10 days after cessation of treatment. These increases were indicative of hepatocellular toxicity. It was concluded that the possibility of equine aflatoxicosis exists although ponies given high quality rations appear to be less susceptible than some other species. Prior exposure to aflatoxins may predispose to clinical toxicity on subsequent exposure, despite lack of expression of clinical signs.

Plasma concentration of iditol dehydrogenase (sorbitol dehydrogenase) in ponies treated with aflatoxin B1.

Twelve clinically normal Shetland ponies were allocated to one of four treatment groups. Aflatoxin B1 was administered at the dosage level of 2 mg/kg of body weight to group A, 1 mg/kg to group B, and 0.5 mg/kg to group C; a placebo was given to group D (controls). Plasma samples were assayed at 4-hour intervals for iditol dehydrogenase (ID) (sorbitol dehydrogenase) concentrations as an indicator of hepatic damage. One of the ponies in group A died 68 hours after dosing; another pony in group A died 76 hours after dosing. All other animals survived the experiment. The means of peak ID values were as follows: group A, 1514.0 IU/l (P < 0.05); group B, 192.6 IU/L (P < 0.05); group C, 8.5 IU/L (P < 0.05); and group D (controls), 2.7 IU/L. A square root transformation analysis of the postdosing response of ID values in relation to time demonstrated that the mean of group A became significantly higher (P < 0.05) than did the mean of the control group at 5 to 6 hours. The mean of group B at 12 to 16 hours and the mean of group C at 20 to 24 hours also were significantly higher (P < 0.05) than was the mean of the control group.