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  • Author or Editor: Andreas Zurbriggen x
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Summary

Only a few hybridization experiments have been performed for detection of canine distemper virus (cdv) nucleic acid sequences in tissue cultures and in various tissues. Those published studies used probes derived from tissue culture-adapted cdv, and hybridization signals were not obtained in the CNS tissue, although infective cdv and viral antigen were detectable in this tissue. We developed probes complementary to virulent cdv and were able to detect viral RNA not only in primary brain cell cultures, but also in brain tissues, by use of in situ hybridization. Sensitivity of the test at least equaled that of immunohistochemistry. We applied digoxigenin-labeled, strand-specific RNA probes complementary to the nucleoprotein-coding viral nucleic acid sequence. Our results indicate that to detect cdv nucleic acid sequences in brain tissues, it is essential to use probes derived from the virulent virus.

Free access
in American Journal of Veterinary Research

Abstract

Objective—To determine whether incubation of cruciate ligament cells with acetylsalicylic acid, carprofen, meloxicam, or robenacoxib provides protection against apoptosis induced by sodium nitroprusside (SNP).

Sample—Explants of cranial (CCL) and caudal (CaCL) cruciate ligaments from eight 1-day-old Beagles.

Procedures—Primary cultures of CCL and CaCL cells were created via enzymatic dissociation of cruciate explants. Purified cell cultures were incubated for 2 hours without (controls) or with 1 of 3 concentrations of 1 of 4 NSAIDs (10, 100, or 200 μg of acetylsalicylic acid/mL; 0.1, 1, or 10 μg of carprofen/mL; 0.1, 1, or 10 μg of meloxicam/mL; or 0.1, 1, or 10 μg of robenacoxib/mL) and subsequently incubated for 18 hours with 1 of 3 concentrations of SNP in an attempt to induce mild, moderate, or severe cytotoxic effects. Cell viability and apoptosis were analyzed via a cell proliferation assay and flow cytometry, respectively. Prostaglandin E2 concentrations were measured via an ELISA.

Results—Cytoprotective effects of NSAIDs were dependent on the extent of SNP-induced apoptosis and were greatest in CCL and CaCL cell cultures with moderate SNP-induced cytotoxic effects. Preincubation with an NSAID improved cell viability by 15% to 45% when CCL and CaCL cells were subsequently incubated with SNP. Carprofen (10 μg/mL) had the greatest cytoprotective effects for CCL and CaCL cells. Incubation with NSAIDs resulted in a nonsignificant decrease in PGE2 production from SNP-damaged cells.

Conclusions and Clinical Relevance—Results indicated that carprofen, meloxicam, and robenacoxib may reduce apoptosis in cells originating from canine cruciate ligaments.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To characterize the mucosal IgE network in dogs affected with inflammatory bowel disease (IBD) and compare it with that for healthy dogs.

Animals—9 healthy dogs and 20 dogs with IBD.

Procedure—In situ hybridization of mRNA specific for IgE and interleukin 4 (IL-4) and immunohistochemical analysis for IgE protein and 2 markers of mast cells (ie, tryptase and chymase) were performed on tissue sections obtained from the gastrointestinal (GI) tract and lymph nodes of dogs.

Results—Dogs with IBD had significantly more cells positive for IgE protein and more mast cells in the GI mucosa than healthy dogs. Despite this significant increase in number of cells positive for IgE, cells positive for IgE mRNA were rarely detected in the GI mucosa; most cells positive for IgE mRNA were found in mesenteric lymph nodes. Signal pattern of IL-4 mRNA was similar to that of IgE mRNA.

Conclusions and Clinical Relevance—The increased numbers of cells positive for IgE and mast cells in dogs with IBD suggest hypersensitivity such as hypersensitivity to bacterial or dietary-derived antigens in the intestinal lumen. Future studies need to elucidate whether this represents a cause of inflammation or is a result of the inflammatory process of IBD. (Am J Vet Res 2001;62:211–216)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine whether lymphocyte apoptosis in intestinal mucosae is more common in healthy dogs than dogs with inflammatory bowel disease (IBD) and whether numbers of apoptotic cells increase after successful treatment of affected dogs.

Animals—8 dogs with IBD (IBD dogs) and 8 healthy control dogs.

Procedures—Biopsy specimens of the duodenum and colon were obtained via endoscopy from dogs with IBD before and after 10 weeks of standard treatment and compared with specimens obtained from control dogs. Expression of activated caspase 3 (Casp3), caspase-cleaved fragment p85 from poly-ADP-ribose polymerase (PARP), and B-cell leukemia/lymphoma 2 (Bcl-2) was measured in the duodenal (villous tip and base) and colonic mucosae.

Results—Expression of Casp3 was greater in the duodenal villous tips of control dogs, compared with expression in similar tissues from dogs with IBD before or after treatment. Despite clinical improvement of dogs with IBD, expression of Casp3 did not increase after treatment. Expression of PARP did not differ between groups at any time point. Expression of Bcl-2 was greater at all 3 tissue sites in control dogs, compared with expression at the same sites in dogs with IBD. Furthermore, Bcl-2 expression in duodenal villous tips was higher in dogs with IBD after treatment but was not higher elsewhere. In control dogs, expression patterns for all 3 markers were similar between sites (villous tip > villous base > colon).

Conclusions and Clinical Relevance—Expression of Casp3 in lymphocytes in duodenal villous tips was significantly reduced in dogs with IBD, compared with expression in healthy dogs, but no increase was detected following successful treatment of IBD. Increased expression of Bcl-2 may be a potential marker of the success of treatment.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To describe the presence and amount of apoptotic ligamentous cells in different areas of partially ruptured canine cranial cruciate ligaments (prCCLs) and to compare these findings with apoptosis of ligamentous cells in totally ruptured cranial cruciate ligaments (trCCLs).

Animals—20 dogs with prCCLs and 14 dogs with trCCLs.

Procedures—Dogs with prCCLs or trCCLs were admitted to the veterinary hospital for stifle joint treatment. Biopsy specimens of the intact area of prCCLs (group A) and the ruptured area of prCCLs (group B) as well as specimens from trCCLs (group C) were harvested during arthroscopy. Caspase-3 and poly (ADP-ribose) polymerase (PARP) detection were used to detect apoptotic ligamentous cells by immunohistochemistry.

Results—No difference was found in the degree of synovitis or osteophytosis between prCCLs and trCCLs. No difference was found in degenerative changes in ligaments between groups A and B. A substantial amount of apoptotic cells could be found in > 90% of all stained slides. A correlation (rs = 0.71) was found between the number of caspase-3-and PARP-positive cells. No significant difference was found in the amount of apoptotic cells among the 3 groups. No significant correlation could be detected between the degree of synovitis and apoptotic cells or osteophyte production and apoptotic cells.

Conclusions and Clinical Relevance—The lack of difference between the 3 groups indicates that apoptosis could be a factor in the internal disease process leading to CCL rupture and is not primarily a consequence of the acute rupture of the ligament.

Full access
in American Journal of Veterinary Research

Abstract

OBJECTIVE To assess the expression of inflammatory cytokines and enzymes in venous whole blood of dogs with impaired renal function attributable to various causes.

ANIMALS 46 dogs with acute kidney injury (AKI), 8 dogs with chronic kidney disease (CKD), and 10 healthy dogs.

PROCEDURES Dogs with AKI and CKD were prospectively enrolled during 2010 if they met inclusion criteria. Demographic and laboratory characteristics were evaluated for each dog, and expression of inflammatory cytokines (interleukin [IL]-1α, IL-1β, IL-8, tumor necrosis factor [TNF]-α, IL-10, and transforming growth factor [TGF]-β) and enzymes (inducible nitric oxide synthase [iNOS] and 5-lipoxygenase [5-LO]) was measured in venous whole blood obtained at initial evaluation.

RESULTS Dogs with impaired renal function had markedly higher expression of the cytokines IL-1α, IL-1β, and TGF-β and the enzyme 5-LO, compared with expression in healthy dogs. Additionally, 17 of 46 AKI dogs (but none of the CKD dogs) had higher IL-8 mRNA expression and 3 of 8 CKD dogs (but only 2/46 AKI dogs) had higher TNF-α expression, compared with results for healthy dogs. No significant difference between renal disease groups was detected for inflammatory markers and laboratory variables, degree of azotemia, or cause of impaired renal function.

CONCLUSIONS AND CLINICAL RELEVANCE In this study, expression of the cytokines IL-1α, IL-1β, and TGF-β and the enzyme 5-LO was clearly increased in dogs with renal disease, which suggested that these markers were part of an inflammatory response in animals with AKI or CKD. (Am J Vet Res 2016;77:218–224)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To evaluate the origin and degree of activity of nitric oxide (NO) and matrix metalloproteinase (MMP) in explants of cranial cruciate ligaments (CCLs) obtained from dogs and cultured with and without inflammatory activators.

Sample Population—Tissue specimens obtained from 7 healthy adult Beagles that were (mean ± SD) 4.5 ± 0.5 years old and weighed 12.5 ± 0.8 kg.

Procedure—The CCLs were harvested immediately after dogs were euthanatized, and specimens were submitted for explant culture. Cultures were stimulated by incubation with a combination of interluekin-1, tumor necrosis factor-α, and lipopolysaccharide, or they were not stimulated. Culture supernatants were examined for production of NO nitrite-nitrate metabolites (NOts) and activity of MMP. Cultured specimens were evaluated by use of immunohistochemical analysis to detect activity of inducible NO synthase (iNOS).

Results—All ligament explants produced measurable amounts of NOts. Stimulated cultures produced significantly more NOts after incubation for 24 and 48 hours, compared with nonstimulated cultures. Production of MMP in supernatants after incubation for 48 hours was significantly higher in stimulated cultures than in nonstimulated cultures. Cells with positive staining for iNOS were detected on all slides. Positively stained cells were predominantly chondroid metaplastic. There was a significant difference in intensity of cell staining between stimulated and nonstimulated cultures.

Conclusion and Clinical Relevance—Explant cultures of intact CCLs obtained from dogs produce iNOS-induced NO. Stimulation of chondroid metaplastic cells in CCL of dogs by use of inflammatory activators can increase production of iNOS, NOts, and MMP. (Am J Vet Res 2002;63:1423–1428)

Full access
in American Journal of Veterinary Research