RESEARCH COMMUNICATIONS OF THE 28th ECVIM-CA CONGRESS

Prostaglandin E2 (PGE2) is a prostanoid playing important homeostatic functions, yet is also responsible for regulating pain and inflammation, and has been implicated in the development and progression of various cancers. The deleterious actions of PGE2 are inhibited in varying degrees byNon‐steroidal anti‐inflammatory drugs (NSAIDs); however, administration of these drugs also leads to significant gastrointestinal (GI) side effects. Grapiprant is a new anti‐inflammatory drug from the piprant class that functions as a selective EP4 prostaglandin‐receptor (EP4R) inhibitor and is proposed to be associated with less GI side effects. The aim of this study was to determine the value of canine jejunal tissue as well as a novel in vitro 3‐dimensional model of canine intestinal epithelium (enteroids) for future in vitro and ex‐vivo investigation of possible GI side effects of drugs in the priprant class. Briefly, ten‐centimeter tissue pieces were acquired from the jejunum of 7 healthy dogs which had been euthanized for an unrelated project. Full thickness tissues were fixed in 10% formalin saline, routinely processed and embedded in paraffin. For enteroid culture, minced samples were washed and crypts were enriched, using EDTA chelation, embedded in matrigel, and grown in intestinal stem cell media until full epithelial differentiation (day 7 of culture). RNA in situ hybridization (RNAscopeÒ, ACDBio) was used to evaluate full thickness samples of canine healthy jejunum as well as enteroids derived from the same tissues. To quantify RNAscopeÒ signals in tissue sections, an advanced digital pathology image analysis system (HALO, Indica Labs) was utilized. Data was expressed as copy number for EP4R with that signal compared to the housekeeping target (B‐actin, positive control gene). In 7/7 jejunal samples evaluated, universal positive expression of EP4R was identified in the epithelium as well as in lamina propria immune cells (copy number 0.1 to 2.6 on average per cell). Expression of EP4 in enteroids showed similar expression in the epithelium when compared to epithelial expression in full thickness samples (Wilcoxon signed rank test, p=0.6).