Placement of ureteral stents in three rabbits for the treatment of obstructive ureterolithiasis.

Management of ureteral obstruction with stenting is often associated with a lower rate of complications than ureterotomy in domestic carnivores, but this treatment has not been previously evaluated in rabbits. Three rabbits (7, 6 and 10 years old) were diagnosed with unilateral obstructive ureterolithiasis associated with hydronephrosis and hydroureter on abdominal ultrasound. Decreased overall renal function was confirmed in all three cases. Ureteral stents were placed retrogradely via cystotomy without complication in two cases and anterogradely via nephrostomy in the third case. Survival after stent placement was 30, 3 and 8 months, with encrustation of the stent and re-obstruction occurring 18, 1 and 6 months after stent placement in successive cases. Ureteral stenting can be considered for short-term management of ureterolithiasis in rabbits to improve renal function and maintain quality of life. Ultrasound or radiographic monitoring is recommended to detect encrustation of the stent. Studies comparing ureteral stenting to ureterotomy in rabbits are needed to determine the effectiveness of these techniques.

Blood concentrations of D- and L-lactate in healthy rabbits.

OBJECTIVES: To determine whole blood and serum concentrations of l-lactate and serum concentrations of d-lactate in healthy rabbits and compare three methods of analysis for l-lactate measurement. METHODS: Prospective study using 25 rabbits. Concentrations of whole blood l-lactate were measured using a portable analyser and a blood gas analyser. The remainder of the sample was allowed to clot for centrifugation. Serum was stored at -20°C for determination of l- and d- lactate by high-performance liquid chromatography. RESULTS: d-lactate values by high-performance liquid chromatography were 0 · 17 ± 0 · 08 mmol/L. l-lactate values were 5 · 1 (±2 · 1) mmol/L by high-performance liquid chromatography, 6 · 9 (±2 · 7) mmol/L with the portable analyser and 7 · 1 (±1 · 6) mmol/L with the blood gas analyser. No significant difference (P > 0 · 05) was found between the two analysers. Significant difference existed between serum l-lactate values obtained by high-performance liquid chromatography and the whole blood values obtained with the blood gas analyser (P < 0 · 01) and portable analyser (P < 0 · 05). CLINICAL SIGNIFICANCE: Serum concentrations of d-lactate in healthy rabbits are in the range of those of other mammals. l-lactate values in healthy rabbits are higher compared with other mammals. Good correlation was found between the portable and blood gas analysers for whole blood l-lactate measurement in healthy rabbits.

Further examination of the effects of nitrosylation on Alternaria alternata mycotoxin mutagenicity in vitro.

Previously, Alternaria extract and metabolite mutagenicities+/-nitrosylation were characterized using Ames Salmonella strains TA98 and TA100, which are both reverted at GC sites. To examine other targets for mutation, the metabolites Altertoxin I (ATX I), Altenuene (ALT), Alternariol (AOH), Alternariol monomethyl ether (AME), Tentoxin (TENT), Tenuazonic acid (TA) and Radicinin (RAD) were reexamined+/-nitrosylation, using Ames Salmonella strain TA97, sensitive to frameshift mutations at a run of C's, as well as strains TA102 and TA104, reverted by base pair mutations at AT sites and more sensitive to oxidative damage. ATX I was also assessed for mammalian mutagenicity at the Hprt gene locus in Chinese hamster V79 lung fibroblasts and rat hepatoma H4IIE cells. When tested from 1 to 100 microg/plate without nitrosylation, ATX I was mutagenic in TA102+/-rat liver S9 for activation and weakly mutagenic in TA104+/-S9, demonstrating direct-acting AT base pair mutagenicity. AOH was also directly mutagenic at AT sites in TA102+/-S9 while AME was weakly mutagenic in TA102+/-S9 and TA104+S9. Nitrosylation of ATX I enhanced mutagenicity at AT sites in TA104+/-S9 but produced little change in TA102+/-S9 compared to native ATX I. However, nitrosylated ATX I generated a potent direct-acting frameshift mutagen at C sites in TA97+/-S9. While ATX I was not mutagenic in either V79 cells or H4IIE cells, 5 and 10 microg/ml nitrosylated ATX I produced a doubling of 6-thioguanine resistant V79 colonies and 0.5 and 1 microg/ml were mutagenic to H4IIE cells, becoming toxic at higher concentrations. These results suggest ATX I, AME and AOH induce mutations at AT sites, possibly through oxidative damage, with nitrosylation enhancing ATX I frameshift mutagenicity at runs of C's. Nitrosylated ATX I was also directly mutagenic in mammalian test systems.

Mutagenicity of bisphenol A (4,4'-isopropylidenediphenol) in vitro: effects of nitrosylation.

Bisphenol A (4,4'-isopropylidenediphenol) is a common component of polycarbonate plastics and epoxy resins. Since bisphenol A-containing plastics and resins have found uses in food-contact items, its potential migration into foodstuffs and possible health consequences have been the focus of many recent studies. However, the potential mutagenic activation of bisphenol A by nitrosylation has received little attention. Incubation of bisphenol A with sodium nitrite under acidic conditions produced a yellow-brown product. When nitrosylated bisphenol A was tested in the Ames Salmonella/microsome assay at 100 ng to 1 mg/plate, dose-dependent increases in mutagenicity were found in both TA98 and TA100 Salmonella strains. These results indicated the presence of a direct-acting mutagenic activity causing both frameshift and base pair mutations, respectively. When compared to colony formation in untreated controls, the addition of rat liver S9 for metabolic activation had little influence on revertant colony formation. Unreacted bisphenol A dissolved in DMSO, acidic buffer, or inactivated nitrosylation solution showed negligible mutagenicity. When the nature of the mutagenic changes was examined using the Ames II trade mark Assay, a variety of base pair changes was found including T:A to A:T - S9, G:C to A:T +/- S9,C:G to A:T +/- S9 and C:G to G:C +/- S9. Bisphenol A also induced frameshift mutations at G:C sites. In addition, the presence of electrophiles was shown by the production of an intensely coloured orange-red product upon incubation of nitrosylated bisphenol A with the nucleophile 4-(4'-nitrobenzyl)pyridine. These findings suggest that migration of bisphenol A into nitrite containing foodstuffs, or its ingestion in the presence of nitrite, could lead to the formation of mutagenic compounds.

Examination of Alternaria alternata mutagenicity and effects of nitrosylation using the Ames Salmonella test.

Molds of the genus Alternaria are common food pathogens responsible for the spoilage of fruits, vegetables, grains, and nuts. Although consumption of Alternaria alternata-contaminated foodstuffs has been implicated in an elevated incidence of esophageal carcinogenesis, the mutagenic potencies of several A. alternata toxins seem unable to account for the levels of activity found using crude mycelial extracts. In this study, the mutagenic effects of nitrosylation were examined with the major Alternaria metabolites Altenuene (ALT), Alternariol (AOH), Alternariol Monomethyl Ether (AME), Altertoxin I (ATX I), Tentoxin (TENT), Tenuazonic Acid (TA), and Radicinin (RAD) using the Ames Salmonella strains TA98 and TA100. In the absence of nitrosylation, ATX I was mutagenic when tested from 1 to 100 microg/plate in TA98 with rat liver S9 for activation, while AOH and ATX I were weakly mutagenic +/- S9 in TA100. Incubation with nitrite generally increased mutagenic potencies with ATX I strongly mutagenic +/- S9 in both TA98 and TA100, while ALT, AOH, AME, and RAD responses were enhanced in TA100 + S9. However, subsequent examination of three extracts made from A. alternata culture broth, acetone-washed mycelia, and the acetone washes showed a different mutagenic response with both broth and acetone washes directly mutagenic in TA98 and TA100 but with a reduced response + S9. The acetone-washed mycelial extract was found to have the lowest mutagenic activity of the three extracts tested. Nitrosylation had little effect on the mutagenicity of any of the extracts. Thus, while nitrosylation increases the mutagenicity of ATX I, and to a lesser extent that of several other Alternaria toxins, the results demonstrate that Alternaria produces a major mutagenic activity with a S. typhimurium response different from that found with the purified toxins. Efforts are currently underway to chemically identify this mutagenic species. Published 2001 Wiley-Liss, Inc.

In vitro investigation of toxaphene genotoxicity in S. typhimurium and Chinese hamster V79 lung fibroblasts.

The polychlorinated pesticide toxaphene has been identified as a persistent environmental contaminant and is of particular concern in the Great Lakes and Arctic regions of Canada. Inconsistencies in published in vitro genotoxicology studies have hindered risk assessments of toxaphene exposure. When toxaphene mutagenicity was re-evaluated in the Ames Salmonella/microsome assay at 10-10,000 microg/plate, a dose-dependent increase in His revertants occurred in all five strains of S. typhimurium tested (TA97, TA98, TA100, TA102 and TA104) with higher mutation frequencies observed in the absence of S9 metabolic activation. However, the mutagenic potential of toxaphene was relatively low with concentrations greater than 500 microg/plate required to induce mutation. Toxaphene genotoxicity was also examined in a mammalian system using Chinese hamster V79 lung fibroblasts with metabolic activation provided by human HepG2 hepatoma cells. Genotoxicity of 1-10 microg/ml toxaphene was examined by measuring the frequency of sister chromatid exchange (SCE) and mutation induction at the hypoxanthine guanine phosphoribosyl transferase (HGPRT) gene locus. Although small increases in SCE were observed at toxic concentrations of toxaphene approaching the LD50 (10 microg/ml), they were not found to be statistically significant relative to control. Toxaphene was also unable to induce HGPRT mutagenesis at the concentrations tested. These results show that while toxaphene is a weak, direct-acting mutagen in the Ames Salmonella Test, convincing evidence of dose-dependent SCE induction and mutagenicity at the HGPRT gene locus could not be demonstrated in V79 cells.

Sequence analysis of the UL39, UL38, and UL37 homologues of bovine herpesvirus 1 and expression studies of UL40 and UL39, the subunits of ribonucleotide reductase.

We sequenced the region of the bovine herpesvirus type 1 (BHV-1) genome corresponding to map units 0.172-0.230 (7964 bp), representing the UL39, UL38, and UL37 homologues of herpes simplex virus which encode the large subunit of ribonucleotide reductase (RR) and components of the viral capsid and the tegument, respectively. To discriminate between two potential initiator AUGs of the UL39 gene, the 5' end of the mRNA was mapped by S1 nuclease protection assays. Comparison of the amino acid sequences of the three BHV-1 proteins with analogous polypeptides from several other herpesviruses revealed significant levels of homology. We also compared the expression kinetics of the large (R1, UL39) versus the small (R2, UL40) RR subunits during the course of in vitro BHV-1 infection by Western blotting using specifically developed and calibrated antisera. Our results show that the R1 protein was synthesized earlier than its R2 counterpart. Moreover, the R1 protein accumulated to a higher level than the R2 protein even though the R2 transcript was in greater abundance than the R1 mRNA. This is discussed with regard to the translational efficiency of their transcripts.

Evaluation of cytotoxicity and genotoxicity of okadaic acid, a non-phorbol ester type tumour promoter, in V79 Chinese hamster lung cells.

Okadaic acid, a specific phosphatase inhibitor and non-phorbol ester type tumour promoter, was examined for cytotoxic and genotoxic properties in a hepatocyte-mediated assay with V79 Chinese hamster lung fibroblasts. Genetic endpoints measured were: mutation to 6-thioguanine resistance at the hgprt gene locus and induction of sister chromatid exchange (SCE). The cytotoxicity of okadaic acid to V79 cells was determined in the absence of hepatocytes at concentrations from 0 to 30 ng/ml medium. Okadaic acid decreased colony size at 20 ng/ml, reduced colony formation by 50% (LC(50)) at 24 ng/ml, and was lethal to 100% of the cells at concentrations above 30 ng/ml. Exposure of V79 cells to okadaic acid for as little as 24 hr without hepatocytes invoked morphological changes that were concentration related. By comparison, okadaic acid was less able to invoke changes in morphology of V79 cells when co-cultivated with rat hepatocytes, but did alter hepatocyte morphology. In genotoxicity tests, mutation frequencies were increased signficantly by okadaic acid only in the absence of hepatocytes and only at the highest tested concentration (20 ng/ml medium). In contrast, SCE frequencies were not affected when okadaic acid was tested alone or with hepatocyte activation at concentrations as high as 20 ng/ml. It was concluded that, within the limits of the test system used, okadaic acid was strongly cytotoxic and may function as a direct-acting mutagen, but was otherwise without cytogenetic activity towards V79 cells.

Gonadotropins induce accumulation of insulin-like growth factor I mRNA in pig granulosa cells in vitro.

Pig granulosa cells have been shown to synthesize insulin-like growth factor (IGF) I peptide in vitro, and this expression is regulated by gonadotropins via the cAMP pathway. By hybridizing an IGF I cDNA probe with total RNA isolated from pig granulosa cells cultured in vitro, we show that these cells contain two IGF I transcripts of about 0.9 kb and 9 kb in size. Treatment of the cells with gonadotropins (follicle-stimulating hormone, luteinizing hormone) or cAMP agonists (dibutyryl-cAMP, forskolin) induces an accumulation of the transcripts which can be abolished by transcriptional inhibitors, but not by translational inhibitors. We thus provide new evidence that pig granulosa cells are a site of IGF I synthesis, and we conclude that (1) gonadotropins increase IGF I mRNA levels; (2) the accumulation of IGF I mRNA results from an increased transcription; (3) the stimulation of IGF I gene transcription does not require ongoing protein synthesis; (4) these effects of follicle-stimulating hormone can be mimicked by cAMP agonists.

Effect of polychlorinated biphenyls on several endocrine reproductive parameters in the female rhesus monkey.

Female rhesus monkeys (Macaca mulatta) ingested gelatin capsules containing daily doses of 0 (control), 5, 20, 40, or 80 micrograms of Aroclor 1254/kg/day (PCB) which was dissolved in corn oil plus glycerol. After approximately two years of dosing and when the monkeys were near an adipose tissue PCBs equilibrium, each dose group of 16 animals was randomly divided into two test groups. Daily blood samples from both groups were acquired for estrogen and progesterone analysis during one menstrual cycle. Test group 1 was sampled during February-March and test group 2 during August-September. Serum estrogen and progesterone concentrations in monkeys dosed with PCBs were equivalent to control values with the exception of the luteal phase progesterone levels in the 20 and 80 micrograms/kg/day dosed monkeys in test group 1. There was no difference in menstrual cycle length between control and treated monkeys for the month sampled, however, menses duration was marginally longer in the 80 micrograms/kg/day dose group. There were no apparent treatment related differences in the incidence of anovulatory cycles nor on the temporal relationship between the estrogen peak and menses onset, menses end or the progesterone peak.