G. Pengo

Comparison of microbiological, histological, and immunomodulatory parameters in response to treatment with either combination therapy with prednisone and metronidazole or probiotic VSL#3 strains in dogs with idiopathic inflammatory bowel disease.

Background Idiopathic inflammatory bowel disease (IBD) is a common chronic enteropathy in dogs. There are no published studies regarding the use of probiotics in the treatment of canine IBD. The objectives were to compare responses to treatment with either combination therapy (prednisone and metronidazole) or probiotic strains (VSL#3) in dogs with IBD. Methodology and Principal Findings Twenty pet dogs with a diagnosis of IBD, ten healthy pet dogs, and archived control intestinal tissues from three euthanized dogs were used in this open label study. Dogs with IBD were randomized to receive either probiotic (D-VSL#3, n = 10) or combination drug therapy (D-CT, n = 10). Dogs were monitored for 60 days (during treatment) and re-evaluated 30 days after completing treatment. The CIBDAI (P<0.001), duodenal histology scores (P<0.001), and CD3+ cells decreased post-treatment in both treatment groups. FoxP3+ cells (p<0.002) increased in the D-VSL#3 group after treatment but not in the D-CT group. TGF-β+ cells increased in both groups after treatment (P = 0.0043) with the magnitude of this increase being significantly greater for dogs in the D-VSL#3 group compared to the D-CT group. Changes in apical junction complex molecules occludin and claudin-2 differed depending on treatment. Faecalibacterium and Turicibacter were significantly decreased in dogs with IBD at T0, with a significant increase in Faecalibacterium abundance observed in the animals treated with VSL#3 strains. Conclusions A protective effect of VSL#3 strains was observed in dogs with IBD, with a significant decrease in clinical and histological scores and a decrease in CD3+ T-cell infiltration. Protection was associated with an enhancement of regulatory T-cell markers (FoxP3+ and TGF-β+), specifically observed in the probiotic-treated group and not in animals receiving combination therapy. A normalization of dysbiosis after long-term therapy was observed in the probiotic group. Larger scale studies are warranted to evaluate the clinical efficacy of VSL#3 in canine IBD.

Immunohistochemical Expression of Ornithine Decarboxylase, Diamine Oxidase, Putrescine, and Spermine in Normal Canine Enterocolic Mucosa, in Chronic Colitis, and in Colorectal Cancer

We compared the immunohistochemical expression of putrescine (PUT), spermine (SPM), ornithine decarboxylase (ODC), and diamine oxidase (DAO) in bioptic samples of canine colonic mucosa with chronic inflammation (i.e., granulomatous colitis and lymphoplasmacytic colitis) or neoplasia. Single and total polyamines levels were significantly higher in neoplastic tissue than in normal samples. Samples with different degrees of inflammation showed a general decrease expression of ODC if compared to controls; SPM was practically not expressed in control samples and very low in samples with chronic-granulomatous inflammation. In carcinomatous samples, the ODC activity was higher with respect to controls and samples with inflammation. This is the first description of polyamines expression in dog colonic mucosa in normal and in different pathological conditions, suggesting that the balance between polyamine degradation and biosynthesis is evidently disengaged during neoplasia.

Severe Gastritis with Double Helicobacter spp. Infection Associated with Barrett's Esophagus in a Cheetah

To the Editor, Gastric infection secondary to Helicobacter spp. is frequent in both humans and many animal species. Therefore, the better understanding of these infections in animals may be also useful as a model for human diseases, and to increase knowledge on potentially transmissible conditions. For instance, recent genomic studies suggest that several species of large felids may have acquired Helicobacter infections through predation on early humans [1]. Although gastritis is rare in wild cheetahs despite the presence of abundant spiral bacteria, worldwide the majority of captive cheetahs (Acinonyx jubatus) have a progressive gastritis associated with a specific Helicobacter infection (Helicobacter acinonyx), and such infection represents a major cause of death or the reason for euthanasia [2]. There are many studies on Helicobacter-induced gastritis in cheetahs; however, to the best of our knowledge, there are no studies of esophagitis related to chronic vomiting due to gastritis in such species. A 7-year-old male cheetah hosted at Falconara zoo underwent upper endoscopy due a history of vomiting after food intake since the age of 3 months, vomiting that in the last months increased to an episode two hours after every meal, with deterioration of the clinical conditions. Endoscopy revealed hiatal hernia (about 4 cm diameter) and severe esophagitis of the lower esophagus associated with slightly raised and fragile mucosa well demarcated from the remaining mucosa and extending upward for about 4 cm starting from the esophago-gastric junction. The stomach revealed diffuse erythema. Multiple biopsies were taken from both the esophagus and the gastric mucosa. Three lm thick sections were stained with hematoxylin and eosin, periodic acid-Schiff–alcian blue (pH 2.5), and Giemsa. Barrett’s esophagus was diagnosed by identifying columnar epithelium in specimens taken more than 3 cm above the cardia. The activity of the inflammatory process was evaluated by grouping the specimens into three categories: inactive chronic inflammation, with no polymorphonuclear leukocytes (PMNs) in the lamina propria; active chronic inflammation when PMNs were present in the lamina propria with intra-epithelial infiltration; and erosion-ulceration in terms of the fibrinoleukocytic exudate observed. Helicobacter-like organisms (HLOs) were identified from their morphological characteristics using Giemsa stain at a magnification of x 400 (concentration) and x1000 (morphology). Then, the presence of spiral-shaped bacteria was also confirmed by immunohistochemistry, using a polyclonal antibody (rabbit polyclonal anti-H. pylori – DAKO, Denmark). The degree of infection was determined as mild moderate or severe by using a similar visual analog scale for canine gastric biopsy specimens as proposed by Happonen et al. [3]. Esophageal histology was characterized by hyperplasia of the epithelial basal layer. As a result of the hyperplasia, the deeper and thicker papillae occupied more than two-thirds of the mucosal layer. Some of the squamous epithelium was replaced by metaplastic epithelium (Fig. 1A) with abundance of goblet columnar cells that were periodic acid-Schiff (PAS)-positive, and columnar cells with a brush border and few cytoplasmic vacuoli similar to small intestinal epithelium (Fig. 1B). These cells were organized in rough, pseudo-villous structures lacking absorptive capacity (i.e., prismatic absorptive cells were absent, indicative of incomplete intestinal metaplasia), and they contained areas of cardial gastric epithelium characterized by atrophic mucous glands. The presence of spiral-shaped bacteria in metaplastic glandular cardial gastric epithelium was evidenced by immunohistochemistry (Fig. 1B, insert). A moderate inflammatory infiltrate of lymphocytes, plasma cells, and rare PMNs was observed in the mucosa, sometimes penetrating among pavement cells. Rare fundic glands with parietal cells were also seen. Inflammatory cells (i.e., neutrophils, lymphocytes, plasma cells) were also present among the epithelial cells. An intense inflammatory infiltrate of lymphocytes, plasma cells, and rare macrophages was present in the underlying stroma (Fig. 1A). According to Terio et al. [4], the stomach displayed diffuse severe gastritis, with large numbers of inflammatory cells in both the superficial and deep regions of the lamina propria, as well as abundant intra-epithelial lymphocytes. Inflammatory cells consisted predominately of lymphocytes and plasma cells with variable numbers of large globule leukocytes. Disruption of normal glandular structure, loss of parietal cells (many of which were sloughed and within gland lumens), and necrosis were also present. Intraglandular

Evidence of Anti-Gliadin and Transglutaminase Antibodies in Sera of Dogs Affected by Lymphoplasmacytic Enteritis

Celiac disease is a disorder that affects the small intestine of genetically predisposed individuals and is developed by the presence of the gliadin fraction of wheat gluten. This disease is characterized by a high seric titre of antibodies against cereal components (anti-gliadin antibodies) and anti-endomysium antibodies (Fasano and Catassi, 2001). It has recently been demonstrated that the modification of gliadin by tissue transglutaminase (tTG) and the formation of gliadin-tTG complexes is important in the immunophatogenesis of the disorder (Dieterich et al., 1998). In human medicine the diagnosis of celiac disease takes advantage of the presence of specific antibodies in the serum of patients (anti-gliadin antibodies, AAG; anti-endomysium antibodies, EmA; anti-transglutaminase antibodies, anti-tTG). The evaluation of such antibodies is useful in selection of patients that have a high probability to be affected by celiac disease, in order to perform an intestinal biopsy. In the dog lymphoplasmacytic enteritis is a significant problem, because it is correlated to weight loss, decrease in performance, diarrhoea and vomiting. In many cases strong plasmacytic infiltration can induce lymphagiectasis, and consequently hypoproteinemia. The aim of this work was to set up a method to evaluate the presence of AAG and anti-tTG antibodies in symptomatic dogs with a clinical diagnosis of lymphoplasmacytic enteritis. For this purpose, one commercial preparation of gliadin and one of transglutaminase were loaded on polyacrylamide gel electrophoresis in the presence of sodium-dodecyl sulphate (SDS-PAGE), blotted on a nitrocellulose membrane and utilized as antigens to search for AAG and anti-tTG antibodies. The sera of clinically and histologically healthy dogs were used as negative controls, while a preparation of anti-gliadin and anti-transglutaminase antibodies was used as a positive control. The obtained data indicated the presence of a high prevalence of AAG and anti-tTG antibodies in the sera of dogs with lymphoplasmacytic enteritis. Therefore, this test, once it has been tested on greater numbers of animals, may be used to evaluate the presence of an intestinal pathology and to establish an appropriate diet for the dog.