Use of quantitative real-time PCR to determine the local inflammatory response in the intestinal mucosa and muscularis of horses undergoing small intestinal resection.

ABSTRACT

BACKGROUND: Studies in rodents and humans have demonstrated that intestinal manipulation or surgical trauma initiates an inflammatory response in the intestine which results in leucocyte recruitment to the muscularis externa causing smooth muscle dysfunction. OBJECTIVES: To examine the intestinal inflammatory response in horses undergoing colic surgery by measuring relative differential gene expression in intestinal tissues harvested from surgical colic cases and control horses. STUDY DESIGN: Prospective case-control study. METHODS: Mucosa and muscularis externa were harvested from healthy margins of resected small intestine from horses undergoing colic surgery (n = 12) and from intestine derived from control horses euthanised for reasons unrelated to the gastrointestinal tract (n = 6). Tissue was analysed for genes encoding proteins involved in the inflammatory response interleukin (IL) 6 and IL1ß, C-C motif chemokine ligand 2 (CCL2), tumour necrosis factor (TNF), prostaglandin-endoperoxide synthase 2 (PTGS2) and indoleamine 2,3-dioxygenase (IDO1). Relative expression of these genes was compared between the two groups. Further analysis was applied to the colic cases to determine whether the magnitude of relative gene expression was associated with the subsequent development of post-operative reflux (POR). RESULTS: Samples obtained from colic cases had increased relative expression of IL1ß, IL6, CCL2 and TNF in the mucosa and muscularis externa when compared with the control group. There was no difference in relative gene expression between proximal and distal resection margins and no association between duration of colic, age, resection length, short-term survival and the presence of pre-operative reflux and the relative expression of the genes of interest. Horses that developed POR had significantly greater relative gene expression of TNF in the mucosa compared with horses that did not develop POR. MAIN LIMITATIONS: Small sample size per group and variation within the colic cases. CONCLUSIONS: These preliminary data support an upregulation of inflammatory genes in the intestine of horses undergoing colic surgery.