In vivo histologically equivalent evaluation of gastric mucosal topologic morphology in dogs by using confocal endomicroscopy

Abstract

Background: Confocal endomicroscopy (CEM) is an endoscopic technology permitting in vivo cellular and subcellular imaging. CEM aids real-time clinical assessment and diagnosis of various gastrointestinal diseases in people. CEM allows in vivo characterization of small intestinal mucosal morphology in dogs.

Objective: To determine the feasibility of CEM to evaluate gastric mucosal morphology in dogs and to characterize the appearance in healthy dogs.

Animals: Fourteen clinically healthy research colony dogs.

Methods: Experimental study. Under general anesthesia, dogs underwent standard endoscopic evaluation and CEM of the gastric mucosa. In the initial 6 dogs, fluorescent contrast was provided with the fluorophore acriflavine (0.05% solution), applied topically. Subsequently, 8 dogs were assessed using a combination of fluorescein (10% solution, 15 mg/kg IV), followed by acriflavine administered topically. For each fluorophore, a minimum of 5 sites were assessed.

Results: Confocal endomicroscopy provided high quality in vivo histologically equivalent images of the gastric mucosa, but reduced flexibility of the endoscope tip limited imaging of the cranial stomach in some dogs. Intravenous administration of fluorescein allowed assessment of cellular cytoplasmic and microvasculature features. Topical application of acriflavine preferentially stained cellular nucleic acids, allowing additional evaluation of nuclear morphology. Identification of Helicobacter-like organisms was possible in 13 dogs.

Conclusion and clinical importance: Confocal endomicroscopy provides in vivo images allowing assessment of gastric mucosal morphology during endoscopy, potentially permitting real-time diagnosis of gastrointestinal disease.

Keywords: Acriflavine; Endoscopy; Fluorescein; Gastroenterology; Gastrointestinal.

Figure 1

(A) Pyloric antrum. Confocal endomicroscopy (CEM) image of the mucosal surface of the lower pyloric antrum. Only cellular cytoplasmic features are highlighted. Superficial imaging of the mucosal surface demonstrates the regular mosaic pattern of the epithelial cells. Image collected after intravenous administration of fluorescein. (B) Pyloric antrum. Topical administration of acriflavine results in preferential staining of nuclear contents providing superior visualization of individual cells and enhancing the superficial mosaic pattern. Histologic images of the pyloric antrum, including standard orientations (C) and orientations comparative to those obtained by using CEM (D) are also shown.

Figure 2

Confocal endomicroscopy (CEM) allowed differences in mucosal morphology corresponding to known histologic differences among the various regions of the stomach to be appreciated with either fluorophore. (A) Upper pyloric antrum after intravenous administration of fluorescein. (B) Gastric body after topical administration of acriflavine demonstrating a flattened mucosal architecture with round‐to‐elliptical gastric pit openings separated by wide mucosal folds. Histologic images including standard orientations (C) and orientations comparative to those obtained by using CEM (D) are also shown.

Figure 3

(A) Topical administration of acriflavine in combination with subsurface imaging ensures individual cells within the lamina propria are identifiable. (B, C) The distinction between the surface epithelial cell layer and the lamina propria is seen where the nuclei of individual columnar epithelial cells line up against the basement membrane (black arrow) and are comparative to that seen histologically (B).

Figure 4

Subsurface imaging after administration of intravenous fluorescein allows the distinction between the superficial epithelial layer and the lamina propria (black arrow). The microvascular network can be seen within the lamina propria; however it is indistinct in comparison to that seen in confocal endomicroscopy images of the small intestine (inset).

Figure 5

Organisms consistent with Helicobacter‐like organisms were identified in 13/14 dogs by using confocal endomicroscopy after administration of topical acriflavine and were seen as elongated fluorescent silhouettes displaying a characteristic spiral shape. Organisms were predominantly seen superficially within the gastric mucus, or deeper between gastric folds (inset).

Figure 6

Confocal endomicroscopy identified a number of findings of currently unknown importance. (A) Occasional isolated cells demonstrate distinct differences in fluorescent intensity compared with surrounding cells, cytoplasmic vacuolation, or both (arrow). (B) The distinct mosaic pattern of the superficial epithelial cells is disrupted in this region, with some cells displaying a finely granular appearance to the cytoplasm. (C) In 1 dog, infrequent, but wide areas of finely vacuolated cells were identified. The overall surface architecture was otherwise retained. All images collected after topical acriflavine administration.

Figure 7

(A) Motion artifact. Confocal endomicroscopy image collected after topical administration of acriflavine demonstrating motion artifact affecting the bottom of the image. (B) Food particulate matter. Cellular debris can be seen in the foreground of this image. The sheet of material comprising individual large cell‐like structures is consistent with plant material. Image collected after topical administration of acriflavine. (C) Poor mucosal contact and gas‐bubble artifact. Complete mucosal contact has not been achieved and the mucosal surface is only visualized indistinctly in the bottom left of the image. Large, negatively contrasting, circular artifacts, consistent with gas bubbles are also seen in the upper and lower aspect of the image. Image collected after topical administration of acriflavine. (D) Vibration artifact. Vibration of the optical fiber was also occasionally seen as a subtle, but regular, pattern of parallel lines partially obscuring the width of an image (magnified image inset). Image collected after topical administration of acriflavine.