Intestinal Cytokine mRNA Expression in Canine Inflammatory Bowel Disease: A Meta-Analysis with Critical Appraisal

Data implicating mucosal cytokines in the pathogenesis of canine inflammatory bowel disease (IBD) are limited. The aims of the present study were to report new findings of intestinal cytokine expression in dogs with IBD and to compare these data with previous studies through meta-analysis. Cytokine mRNA abundance in intestinal biopsies collected prospectively was evaluated by using a semiquantitative RT-PCR technique. For meta-analysis, an electronic database search revealed 3 clinical trials, all of which were nonrandomized (type III) case series. Prospective analysis showed that the intestines of healthy dogs and those with IBD express numerous cytokines and that a proinflammatory expression profile is not a feature of small or large-intestinal IBD. The meta-analysis data included 158 dogs characterized as healthy (n = 45), diarrheic nonIBD dogs (n = 6), nonresponders (n = 2), small-intestinal IBD (n = 41), colonic IBD (n = 25), and chronic enteropathy (n = 39). German shepherd dogs were overrepresented in 3 of the 4 studies. Healthy dogs showed mRNA expression for most cytokines including IL2, IL4, IL5, IL10, IL12, IFNγ, TNFα, and TGFβ. Only IL12 mRNA expression was increased consistently in small-intestinal IBD, whereas IBD colitis lacked consistent patterns of expression. In summary, dogs with IBD fail to express a predominant Th1- or Th2 cytokine bias in inflamed mucosa. Heterogeneity of results among these studies might be explained by numerous factors including the method of mRNA quantification, stage of disease, and demographic differences in study populations.Abbreviations: GAPDH, glyceraldehyde 3-phosphate dehydrogenase; IBD, inflammatory bowel disease; LP, lymphocytic–plasmacyticIdiopathic inflammatory bowel disease (IBD) in dogs is a chronic immune-mediated disorder empirically defined by clinical, histologic, and therapeutic features.^{18,26,27,29,45} Evidence suggests that intestinal inflammation in IBD results from altered interaction between the resident microflora and mucosa in a susceptible host.^48,53 Aggressive host immune responses directed against commensal bacteria play a central role in the pathogenesis of chronic mucosal inflammation. The concept of impaired immunoregulation in canine IBD is supported by observations of increased numbers of immunoglobulin-containing cells and T cells in inflamed tissues,^16,28,30,49 upregulated mucosal and luminal expression of nitric oxide metabolites,^20,29 and altered serum concentrations of select acute phase proteins, such as C-reactive protein, in diseased dogs.³¹ C-reactive protein is a marker of inflammation and tissue injury and is produced by the liver in response to stimulation by IL6, IL1β, and TNFα.^14,51Cytokines play a key role in the modulation of the mucosal immune system of humans. To maintain gut homeostasis, the normal mucosal immune system balances a network of inflammatory mediators, including proinflammatory, antiinflammatory, and regulatory cytokines.⁴⁷ Cytokines are synthesized rapidly and secreted on stimulation and induce the production of adhesion molecules and other inflammatory mediators including reactive oxygen species, nitric oxide metabolites, and lipid products such as prostaglandins, leukotrienes, and platelet-activating factor. Cytokine-producing cells induce, amplify, prolong, and mediate intestinal mucosal injury.¹³ Disturbances in the balance of proinflammatory (Th1/Th17-derived) and immunoregulatory (Th2/Tr1-derived) cytokines occur in humans with IBD as well as numerous animal models of intestinal inflammation.^16,39,44Data evaluating the role of CD4⁺ T cells and mucosal cytokines in the pathogenesis of canine IBD are limited. Lymphocytes expressing CD4⁺ are either increased¹⁶ or decreased²⁸ in dogs having small-intestinal IBD, whereas mucosal CD4⁺ T cells are increased in dogs with IBD colitis.^30,49 A recent study¹⁵ described a balance between proinflammatory and antiinflammatory cytokine mRNA expression in dogs with small-intestinal enteropathies but included only 4 dogs diagnosed with IBD. A separate investigation⁴⁴ evaluating mucosal cytokine mRNA expression in dogs with lymphocytic–plasmacytic colitis reported upregulated expression of proinflammatory cytokines IL2 and TNFα. Yet another study⁴¹ reported no difference in cytokine expression in the duodenal mucosa of dogs with or without chronic diarrhea; however, the dogs of this report were not subdivided in terms of response to therapy as having idiopathic IBD, antibiotic-responsive diarrhea, or food-responsive enteropathy. Because of these varied observations, it is unclear which cytokines, if any, control or enhance the local immune response of canine IBD because 1) few dogs with IBD have been evaluated, 2) German shepherd dogs with enteropathies were over-represented in most studies, 3) various measures of cytokine mRNA expression were used, and (4) distinctly variable patterns of cytokine expression were present among these earlier studies.The objectives of this study were to 1) assess cytokine mRNA expression in the intestinal mucosa of dogs diagnosed with small- and large-intestinal IBD by using an RT-PCR technique and 2) compare these data with previously published data to determine the putative role of cytokine expression in the pathogenesis of canine IBD and other forms of chronic enteropathy through meta-analysis of combined findings.