Full length article A co culture approach show that polyamine turnover is affected during inflammation in Atlantic salmon immune and liver cells and that arginine and LPS exerts opposite effects on p38MAPK signaling Elisabeth Holen a, * , Marit Espe a , Synne M. Andersen a , Richard Taylor b , Anders Aksnes b , Zebasil Mengesha a, c , Pedro Araujo a a National Institute of Nutrition and Seafood Research (NIFES), P.B. 2029 Nordnes, 5817 Bergen, Norway b EWOS Innovation AS, N-4335 Dirdal, Bergen, Norway c Department of Industrial Chemistry, Bahir Dar University, P.B. 79, Bahir Dar, Ethiopia article info Article history: Received 28 October 2013 Received in revised form 3 February 2014 Accepted 9 February 2014 Available online 21 February 2014 Keywords: Atlantic salmon head kidney cells Salmon liver cells Co culture Inflammation Arginine abstract This study assess which pathways and molecular processes are affected by exposing salmon head kidney cells or liver cells to arginine supplementation above the established requirements for growth support. In addition to the conventional mono cultures of liver and head kidney cells, co cultures of the two cell types were included in the experimental set up. Responses due to elevated levels of arginine were measured during inflammatory (lipopolysaccharide/LPS) and non -inflammatory conditions. LPS up regulated the genes involved in polyamine turnover; ODC (ornithine decarboxylase), SSAT (spermidine/ spermine-N1-acetyltransferase) and SAMdc (S-adenosyl methionine decarboxylase) in head kidney cells when co cultured with liver cells. Regardless of treatment, liver cells in co culture up regulated ODC and down regulated SSAT when compared to liver mono cultures. This suggests that polyamines have anti- inflammatory properties and that both salmon liver cells and immune cells seem to be involved in this process. The transcription of C/EBP b/CCAAT, increased during inflammation in all cultures except for liver mono cultures. The observed up regulation of this gene may be linked to glucose transport due to the highly variable glucose concentrations found in the cell media. PPARa transcription was also increased in liver cells when receiving signals from head kidney cells. Gene transcription of Interleukin 1b (IL-1b), Interleukin-8 (IL-8), cyclooxygenase 2 (COX2) and CD83 were elevated during LPS treatment in all the head kidney cell cultures while arginine supplementation reduced IL-1b and IL-8 transcription in liver cells co cultured with head kidney cells. This is probably connected to p38MAPK signaling as arginine seem to affect p38MAPK signaling contrary to the LPS induced p38MAPK signaling, suggesting anti-inflammatory effects of arginine/arginine metabolites. This paper shows that co culturing these two cell types reveals the connection between metabolism and inflammation, suggesting different pathways and candidate biomarkers to be further explored. Ó 2014 Elsevier Ltd. All rights reserved. 1. Introduction Opposite to mammalian species, arginine is considered indis- pensable in fish as the urea cycle is too low to fulfill the re- quirements [1]. When farmed fish are offered diets in which the traditional fish meal is replaced with high inclusion of alternative plant protein ingredients, the amino acid composition may change if not properly blended and supplemented with crystalline amino acids [2]. Also the nitrogen-containing metabolites may be absent or present in too low concentration to support the metabolism and growth [3,4]. In addition, the salmon’s nutritional needs under normal conditions may be inadequate under infectious conditions, exposure to environmental pollutants and other stressful condi- tions. Arginine is the precursor of polyamines (PA), nitrogen oxide (NO), creatine and urea. The enzyme arginase converts arginine to urea and ornithine. Ornithine is the precursor for PA converted by the enzyme ornithine decarboxylase (ODC), the rate limiting enzyme for PA synthesis. Both PA and NO have potential to affect oxidation status through their turnover and degradation [5e7]. PA synthesis requires amino propyl groups which are delivered by * Corresponding author. Tel.: þ47 95241633; fax: þ47 55905299. E-mail address: eho@nifes.no (E. Holen). Contents lists available at ScienceDirect Fish & Shellfish Immunology journal homepage: www.elsevier.com/locate/fsi http://dx.doi.org/10.1016/j.fsi.2014.02.004 1050-4648/Ó 2014 Elsevier Ltd. All rights reserved. Fish & Shellfish Immunology 37 (2014) 286e298