Acute Oxidative Stress Affects IL-8 and TNF-α Expression in IPEC-J2 Porcine Epithelial Cells Erzsebet Paszti-Gere, 1,3 Edina Csibrik-Nemeth, 1 Krisztina Szeker, 1 Rita Csizinszky, 1 Csaba Jakab, 2 and Peter Galfi 1 Abstract—Reactive oxygen species are implicated in cell and tissue damage in a number of diseases including acute and chronic inflammation of the gut. Effects of H 2 O 2 exposure on non-carcinogenic porcine epithelial cell line, IPEC-J2 cells cultured on collagen-coated membrane inserts were mo- nitored based on transepithelial electrical resistance (TER) change, extent of necrotic cell damage, gene expression of inflammatory cytokines IL-8 and TNF-α. Furthermore, the junction proteins claudin-1 and E-cadherin were also investigated by immunohistochemistry. Peroxide (1mM) increased IL-8 and TNF-α gene expression levels significantly allowing 1 h recovery time without affecting the cellular distribution of junction proteins, TER and cell survival rate. In conclusion, the IPEC-J2 cell line on membrane insert was introduced as a fast and reliable investigation tool for oxidative stimuli-triggered intestinal inflammation and in the future as a screening method for antioxidant and probiotic candidates. KEY WORDS: swine; intestinal epithelial cells; IPEC-J2; H 2 O 2 ; inflammatory cytokines. INTRODUCTION In the cell, a delicately controlled balance, the so-called redox homeostasis exists between the physio- logical production of reactive oxygen species (ROS) such as superoxide anion, hydroxyl radical and H 2 O 2 , on one side, and antioxidant defence mechanisms, on the other, which can be readily tipped by excessive oxidative stress or deficiency of preventive antioxidant network causing inflammation and oxidative tissue injury. It was found that in the colon adenocarcinoma cell lines Caco-2 and T84, H 2 O 2 increased the intestinal epithelial permeability, leading to the disruption of paracellular junctional complexes presumably via pro- tein tyrosine phosphorylation-dependent mechanisms [1]. The transepithelial electrical resistance (TER) of Madin–Darby canine kidney (MDCK) type II epithelial cell line exposed to 5 mM H 2 O 2 was reduced to 23% of the control value and returned to baseline within 6 h [2]. Oxidative stress appears to be an important driving force for an enhanced cytokine production in intestinal epithelial cells causing gut mucosal inflammation. It was found that 2 mM H 2 O 2 and TNF-α induced interleukin- 8 (IL-8) secretion in Caco-2 and HT-29 cells [3]. Under short-term oxidative stress induced by 1 mM H 2 O 2 , IL-8 mRNA expression increased in Caco-2 and in human normal intestinal epithelial cells [4]. Strict regulation of IL-8 expression via externally applied H 2 O 2 was also observed in gastric epithelial MKN28 cells [5], where the mode of action of ROS with putative signalling messenger properties may be in correlation with the redox-sensitive nature of the transcription factor NF-κB[6, 7]. The IPEC-J2 cell line was isolated from the jejunal epithelia of a neonatal, unsuckled piglet [8]. The major advantage of IPEC-J2 cells compared with the most widely used colon carcinoma cell lines Caco-2 and HT-29 is that their glycosylation pattern, proliferation rate and colonisation ability characterize better the in 1 Department of Pharmacology and Toxicology, Faculty of Veterinary Sciences, Szent István University, BudapestIstván u. 2 1078, Hungary 2 Department of Pathology and Forensic Veterinary Medicine, Faculty of Veterinary Sciences, Szent István University, Budapest, Hungary 3 To whom correspondence should be addressed at Department of Pharmacology and Toxicology, Faculty of Veterinary Sciences, Szent István University, BudapestIstván u. 2 1078, Hungary. E-mail: Gere.Erzsebet@aotk.szie.hu 0360-3997/12/0300-0994/0 # 2011 Springer Science+Business Media, LLC Inflammation, Vol. 35, No. 3, June 2012 ( # 2011) DOI: 10.1007/s10753-011-9403-8 994