Demonstration of substances of innate immunity in the esophageal epithelium of domesticated mammals. Part I--Methods and evaluation of comparative fixation.

The aim of the investigation was to demonstrate that the esophageal epithelium of domesticated mammals exhibits characteristic features of innate immunity. The esophageal samples used were obtained immediately after euthanization from seven species of domesticated mammals of three nutrition groups (herbivores: horse, goat, cattle; omnivores: pig, dog, laboratory rat; carnivores: cat). The experimental basis was immunohistochemistry, which was evaluated in a qualitative and statistically relevant semi-quantitative manner. The first part of the study analyzed the influence of different fixation media on the immunohistochemical reactivities. Two formalin-based routine fixation solutions (Bouin's solution, Ca-acetate formalin) were compared with the recently introduced formalin-free HOPE® fixative. In this context, we clearly demonstrated a diminished immunoreactivity for Ca-formol fixed samples; satisfactory results were obtained, particularly, from samples fixed in Bouin's solution. The HOPE® fixation method offers a relatively cheap alternative, as the antibody amounts can be reduced. An application in routine diagnostic is not advisable, because of several variable parameters. It can be concluded that immunohistochemical results have always to be evaluated and discussed in close relation to the fixation medium used.

Demonstration of substances of innate immunity in the esophageal epithelium of domesticated mammals: Part II--Defence mechanisms, including species comparison.

The second part of our study deals with a comparative evaluation and discussion of the immunohistochemical results that were obtained. The cryoscanning electron microscopy (cryoSEM) observations confirmed a monolayer colonization of the esophageal surface with bacteria and fungi (yeasts); the latter in particular was prominent in the ruminant species studied. We demonstrated the existence of several innate immune parameters, including pathogen recognition receptors (PRRs), such as Toll-like receptor 2, which was primarily expressed in the stratum basale; however, the presence ß-glucan receptors remained inconclusive. Furthermore, the group of cationic antimicrobial peptides (CAPs) was shown, comprising ß-defensins 2 and 3 and cathelicidin. The less keratinized esophageal epithelium of the carnivorous cat was protected by high amounts of CAPs; whereas the more strongly keratinized epithelium of the herbivorous and omnivorous species with its characteristic layer structure exhibited clearly weaker reactions. Moreover, lysozyme could distinctly be demonstrated in the cells of the esophageal epithelium. It can be concluded that a first line of defence mechanisms of the innate immune system contributes to maintaining a microbial homeostasis on the surface of the esophageal epithelium of domesticated mammals. The results are discussed in comparison to findings from studies on the human esophagus.