OBJECTIVE To investigate the effects of triamcinolone acetonide (TA) and methylprednisolone acetate (MPA) on the viability of resident cells within the fibrocartilage on the dorsal surface of the deep digital flexor tendon (FC-DDFT) and fibrocartilage on the flexor surface of the navicular bone (FC-NB) of horses. SAMPLE 12 to 14 explants of FC-DDFT and of FC-NB from grossly normal forelimbs of 5 cadavers of horses aged 9 to 15 years without evidence of musculoskeletal disease. PROCEDURES Explants were incubated with culture medium (control) or TA-supplemented (0.6 or 6 mg/mL) or MPA-supplemented (0.5 or 5 mg/mL) medium for 6 or 24 hours. Explant metabolic activity and percentage of dead cells were assessed with a resazurin-based assay and live-dead cell staining, respectively, at each time point. Drug effects were assessed relative to findings for the respective control group. RESULTS Application of TA (at both concentrations) did not significantly change the cell viability of FC-DDFT explants. For FC-NB explants, TA at 6 mg/mL significantly reduced the metabolic activity and increased the percentage of dead cells at both time points. With either MPA concentration, FC-DDFT and FC-NB explants had reduced metabolic activity and an increased percentage of dead cells at 24 hours, whereas only MPA at 5 mg/mL was cytotoxic at the 6-hour time point. CONCLUSIONS AND CLINICAL RELEVANCE In ex vivo explants, TA was less cytotoxic to equine FC-DDFT and FC-NB cells, compared with MPA. Further work is warranted to characterize the drugs' transcriptional and translational effects as well as investigate their cytotoxicity at lower concentrations.

OBJECTIVE To compare pregnancy-associated glycoprotein 1 (PAG1) concentrations in maternal (jugular vein) and fetal (uterine vein) circulations and amniotic fluid samples between pregnant ewes that were and were not experimentally infected with bovine viral diarrhea virus (BVDV). ANIMALS 11 healthy pregnant yearling ewes. PROCEDURES Before study initiation, all ewes were naïve to BVDV and confirmed pregnant by transabdominal ultrasonography at approximately 60 days of gestation. At 65 days of gestation, ewes were intranasally inoculated with a noncytopathic BVDV type 1b strain (concentration, 107 TCID50/mL; 2 mL/nostril; n = 6) or an equal volume of BVDV-free viral culture medium (control; 5). A blood sample was collected for measurement of PAG1 concentration before inoculation. At 80 days of gestation, each ewe was anesthetized and underwent an ovariohysterectomy. While sheep were anesthetized, blood samples from the jugular and uterine veins and an amniotic fluid sample were collected for measurement of PAG1 concentration. Fetal tissues underwent real-time PCR analysis for BVDV RNA, and placental specimens underwent histologic evaluation and immunohistochemical staining for BVDV antigen. RESULTS At 80 days of gestation, BVDV RNA in fetal tissues and mild placentitis were detected in 5 of 6 BVDV-inoculated ewes. Mean PAG1 concentrations in the maternal and fetal circulations of BVDV-inoculated ewes were significantly less than those in control ewes. Mean amniotic fluid PAG1 concentration did not differ significantly between the 2 groups. CONCLUSIONS AND CLINICAL RELEVANCE Concentration of PAG1 in the maternal circulation may be a useful biomarker for determining placental health in sheep after viral infection of the reproductive tract.

OBJECTIVE To develop a technique for isolation and culture of canine insulinoma cells and assess expression of cellular hexokinases (glucokinase and hexokinase I) and expression and secretion of insulin from these cells in vitro. SAMPLE Pancreatic insulinomas and normal pancreatic tissue from 4 and 3 dogs, respectively. PROCEDURES Tissues were collected by surgical excision or at necropsy. Insulinoma cells from 2 dogs were cultured for up to 10 weeks with standard techniques; insulin synthesis in vitro was confirmed by immunohistochemical analysis of freshly prepared slides of cultured cells, and insulin secretion was assessed by measurement of insulin concentrations in culture medium with an ultrasensitive mouse insulin ELISA. Expression of cellular hexokinases in insulinomas and adjacent normal (nontumor) pancreatic tissue from the same dog (n = 3) was examined by quantitative reverse transcriptase PCR assay. RESULTS Insulinoma cells survived for up to 10 weeks but did not proliferate in culture. Insulin was detected in isolated cells and secreted into culture medium for up to 10 weeks. Both cellular hexokinases were expressed; glucokinase appeared to be overexpressed in insulinomas, compared with normal pancreatic tissue from the same dogs. CONCLUSIONS AND CLINICAL RELEVANCE Canine insulinomas expressed hexokinases responsible for glucose responsiveness. Insulinoma cells were successfully maintained in short-term culture; cultured cells remained functional for 10 weeks as evidenced by cellular insulin content and had detectable secretion of insulin into the culture medium for ≥ 5 weeks. Apparent glucokinase overexpression by insulinomas suggested a possible mechanism underlying excessive insulin release by these tumors.