S. M. Koehler1, S. Welak2, K. Fredrich1, M. Christensen1, M. Knezevich1, D. Gourlay1 1Children’s Hospital Of Wisconsin,Pediatric Surgery,Milwaukee, WISCONSIN, USA 2Children’s Hospital Of Wisconsin,Neonatology,Milwaukee, WISCONSIN, USA
Introduction: Necrotizing enterocolitis (NEC) is the most common inflammatory intestinal disease in premature neonates and a significant cause of infant mortality. The purpose of this study is to compare levels of intestinal inflammation in neonates with NEC to those with congenital bowel malformations. We hypothesize that NEC will have high levels of inflammation at the involved portion of the bowel, but areas remote from the disease will be uninflamed similar to intestine resected for congenital malformations.
Methods: This is an IRB approved prospective study involving discarded tissue from neonates undergoing intestinal resection for NEC, history of NEC, or congenital anomalies. Formalin-fixed sections were utilized to access for histological grade of injury. Levels mRNA expression of interleukin 6 (IL6), interleukin 1 beta (IL1B), toll-like receptor 4 (TLR4), tissue necrosis factor alpha (TNFα), tissue nonspecific alkaline phosphatase (TNAP), (intestinal alkaline phosphatase (IAP) and NADPH oxidase 1 (NOX1) were evaluated via quantified PCR. Statistical analysis was performed using a Student’s t-test and a p-value of <0.05 was considered significant.
Results: Fifteen samples were collected from eight patients at nine operations. Four samples were considered diseased and were from patients with acute NEC near the site of perforation. The remaining samples were considered healthy. Three were from patients with acute NEC at a site remote from the perforation, three from patients with a history of NEC undergoing stoma reversal, and five from patients with congenital bowel malformations. When comparing mRNA expression of inflammatory and anti-inflammatory markers between intestinal samples from active NEC with the remaining samples, there were significantly higher levels of TNAP (12205 vs. 1270, p=0.009), IL6 (86654 v 24807, p=0.044) and IL1B (133568 vs 14009, p=0.047). Interestingly, some “healthy” samples were highly inflamed and contained ulcerated mucosa. When these tissue samples were compared to those samples with less evidence of histological damage, regardless of diagnosis, there were significantly higher levels of TNAP (10538 vs 1010, p=0.019), IL6 (81377 vs 21260, p=0.036) and TLR4 (12265 vs 4795, p=0.008).
Conclusions: This preliminary data confirms that there are high levels of inflammation in active NEC despite histological grade. The level of inflammation is similar to samples with mucosal ulceration. While both groups showed elevation in TNAP and IL6, they did vary in expression of TLR4 and IL1B. However, the receptor for IL1B shares a signaling domain with toll-like receptors thus this may simply represent an alternative signaling pathway. Furthermore, while TNAP is not typically considered an inflammatory enzyme, its expression is increased during periods of intestinal inflammation. Further studies will be necessary to confirm these findings and further evaluate the relative activity of TNAP and IAP.