Role of n-3 PUFAs in inflammation via resolvin biosynthesis

The role of n-3 PUFAs has gained more importance these last decades, especially in inflammatory processes because they can display anti-inflammatory properties. Inflammation is a protective response of the body in controlling infection and promoting tissue repair. However, excessive inflammation can cause local tissue damage. This is especially the case for the brain for which the functional consequences of neuroinflammation include alterations in cognition, affect and behavior leading to a negative impact on the quality of life and well-being of patients (Dantzer, 2001, 2008). Hence, limiting the inflammation in the brain is a real strategy for neuroinflammatory disease therapy and treatment. Recent data show that n-3 PUFAs exert anti-inflammatory properties in part through the synthesis of specialized pro-resolving mediators such as resolvins that actively turned off the inflammatory response. This review first outlines basic concepts of neuroinflammation and the role of n-3 PUFAs in this process and then summarizes the biosynthesis, signaling pathways and role of resolvins.


Introduction
The role of essential nutrients in the brain development and neuronal functioning has increased in the last decades. In this regard, polyunsaturated fatty acids (PUFAs), especially n-3 PUFAs have gained importance. They are significant structural components of the phospholipid membranes of brain in which docosahexaenoic acid (DHA; 22:6 n-3) constitutes up to 30% of total fatty acids. They assure the correct environment Correspondence: corinne.joffre@bordeaux.inra.fr for membrane protein function, maintain the fluidity and influence lipid raft formation (Calder, 2010). They also act as signaling molecules or ligands for transcription factors (Norheim et al., 2012). Moreover, they are involved in the cerebral development and in the neuronal structure (Madore et al., 2014). They have the ability to modulate the neurotransmission and the synaptic plasticity (Lafourcade et al., 2011). Of importance in many neurodegenerative diseases, they have immuneregulatory properties (Bazinet and Laye, 2014). One of the possible mechanisms to explain the n-3 PUFAs benefits has recently emerged as their conversion in bioactive lipid mediators such as resolvins. In this review we present an overview of the formation and action of n-3 PUFAs derived anti-inflammatory lipid mediator resolvins.
Microglia are the resident macrophages of the brain, and constitute the first line of immune defense (Ransohoff and Cardona, 2010). They derive from myeloid cells in the periphery and comprise approximately 15% of the cells in the brain (Carson et al., 2006). They are involved in tissue homeostasis control, response to injury and remodeling/repair. Under normal conditions, they are in a surveillance phenotype and constantly monitor the environment (Davalos et al., 2005;Nimmerjahn et al., 2005). Once stimulated by an immune challenge, microglia are capable of acquiring diverse and complex phenotypes as well as performing several macrophagelike functions including inflammatory and anti-inflammatory cytokine production (Biber et al., 2007;Garden and Moller, 2006;Madore et al., 2013). If sustained, microglia activation can aggravate the related injury, leading to neuronal damage that is the basis of a large variety of pathologies (Blais and Rivest, 2003;Laye, 2010;Solito and Sastre, 2012;Woodroofe, 1995;Woodroofe and Cuzner, 1993).
Hence, the identification of mediators limiting the inflammation and/or involved in the resolution of inflammation is of growing interest as it may provide novel targets in brain damage prevention and treatment.

Role of n-PUFAs in inflammation
n-3 PUFAs have been shown to have powerful immunomodulatory effects (Calder, 2001;Labrousse et al., 2012;Laye, 2010;Orr et al., 2013). They are highly concentrated in the central nervous system (CNS) and are necessary for normal brain development and function (Labrousse et al., 2012;Larrieu et al., 2012;Luchtman and Song, 2013;Moranis et al., 2012;Xiao et al., 2005). The dramatic reduction in the dietary supply of n-3 PUFAs in Western societies and the corresponding increase in n-6 PUFAs lead to an imbalanced n-6/n-3 ratio currently estimated at 12-20 in developed countries instead of the recommended ratio of 5 (Simopoulos, 2001). These changes in n-3 PUFAs in the diet lead to modifications in the n-3 PUFA content in the brain. As a result, we have previously demonstrated that low dietary intake of n-3 PUFAs promotes neuroinflammatory responses through the regulation of microglial cell activity and polarization toward a pro-inflammatory phenotype, whereas n-3 PUFA dietary supplementation is rather anti-inflammatory (Delpech et al., 2015c;De Smedt-Peyrusse et al., 2008;Labrousse et al., 2012;Madore et al., 2014;Mingam et al., 2008). Moreover, the central n-3 PUFA increase observed in transgenic Fat-1 mice modulates the brain innate immune system activity, leading to the protection of animals against LPS-induced pro-inflammatory cytokine production and subsequent spatial memory alteration (Delpech et al., 2015a). Hence, a dramatic reduction in the dietary supply of n-3 PUFAs could thus contribute to the sensitization of the brain immune response to further inflammation, and thus to the development of spatial memory disorders.
The mechanisms by which n-3 PUFAs exert their effect are not clearly established. Interestingly, their effect can be mediated via lipid mediators because n-3 PUFAs can act as precursors of specialized pro-resolving mediators (SPM) involved in the anti-inflammation and pro-resolution. The resolution of inflammation is an actively regulated part of the inflammatory response involving the activation of specific molecules and cells that signal the end of inflammation and turn it off.

Role of resolvins in inflammation
Recent data emphasize the importance of SPM generated from PUFAs. These compounds are key regulators and mediators of inflammation. They were identified using a lipidometabolomic system approach to analyze the cellular and molecular components of exudates during inflammation. They are active at nanomolar range unlike their precursors that act at micromolar concentrations (Claria et al., 2011). They act locally and may be rapidly inactivated by further metabolism via enzymatic pathways (Arita et al., 2005;Seki et al., 2009). They have the ability to regulate the progress of inflammatory response and activate the resolution of inflammation in a number of cell types and models of inflammation. To date, only a few DHA-derived mediators, including 17S-hydroxy-DHA (17-HDHA), neuroprotectin D1 (NPD1), resolvin D5 (RvD5), 14-HDHA and maresin 1 (MaR1), have been identified within brain tissue (Orr et al., 2013;Serhan, 2014). In patients, RvD1 was measured in plasma and macrophages (Fiala et al., 2015;Wang et al., 2015a). As resolvins have been mostly studied on peripheral cells, we focused on these compounds.

Actions of RvD1 and RvE1 at the periphery
The anti-inflammatory activities of RvD1 and E1 have been reported both in vitro and in vivo mostly on peripheral cells. Their pro-resolving effects are widely described in macrophages in rodent models of inflammation (for reviews: Claria et al., 2011;Fredman and Serhan, 2011;Lee and Surh, 2012;Recchiuti, 2013;Seki et al., 2009;Serhan, 2014).

Actions of resolvins in the central nervous system
Very few studies described the role of resolvins in the central nervous system, in particular in microglia cells. RvD1 and its receptor were detected in the cerebrospinal fluid of control and Alzheimer patients (Wang et al., 2015b). The importance of the resolution pathway in maintaining normal cognition is suggested by the highlighted positive correlation between Mini-Mental State of Examination (MMSE) and the levels of RvD1 in the cerebrospinal fluid, suggesting that resolution can inhibit Alzheimer disease-related cognitive decline. Other studies published data reporting that a supplementation in n-3 PUFAs in patients with minor cognitive impairment increases RvD1 in macrophages (Fiala et al., 2015) and in vitro RvD1 with vitamin D promotes Aβ-phagocytosis in isolated Alzheimer's patient macrophages (Mizwicki et al., 2013). A study of Harrison et al. (2015) demonstrates that RvE1, administered intraperitoneally for consecutive days, decreases the traumatic brain injury-induced activation of microglia. RvE1 increases the proportion of ramified microglia and decreases the proportion of rod microglia in the sensory cortex. Moreover, RvE1 significantly alters the inflammatory profile of microglia (Harrison et al., 2015).

Mechanisms of actions of RvD1 and RvE1
The mechanisms by which RvD1 acts are not yet clearly established. It was shown that RvD1 acts via its receptor ALX/fpr2 to regulate specific miRNAs that are key regulators for resolution of inflammation (Bartel, 2009;Recchiuti, 2013). miRNA are small ∼23 nt endogenous RNA that can play important gene regulatory roles by pairing to the mRNA of protein coding genes to direct their posttranscriptional repression. miRNAs has recently emerged as a major class of gene expression regulators linked to most biological functions including immune regulation (Ceppi et al., 2009;O'Neill et al., 2011;Recchiuti et al., 2011;Recchiuti and Serhan, 2012). miRNAs in macrophages downregulate the mRNA translation of key inflammatory cytokines . miR-155, miR-21 and miR-146 have been central in much miRNA research due to their expression levels following LPS-induced inflammation (Quinn and O'Neill, 2011). Ceppi et al. (2009) reported that both miR-155 and miR-146 are up-regulated upon LPS stimulation in human primary dendritic cells (Ceppi et al., 2009). miR-155 targets the proteins involved in the activation of NFκB, thus controlling tissue damage due to inflammation (Faraoni et al., 2009). It is characterized as a common target of a broad range of inflammatory mediators (O'Connell et al., 2007). miR-146 is D104, page 3 of 6 Dossier C. Joffre et al.: OCL 2016, 23(1) D104 involved as a negative regulator fine tuning the immune response (Quinn and O'Neill, 2011). These miRNAs play a key role in modulating the IL-1 and IL-6 pathways. miR-21 is also involved as a central player in the inflammatory response (Quinn and O'Neill, 2011). miR-21 plays a key role in the resolution of inflammation and in negatively regulating the proinflammatory response in particular in macrophages (Sheedy and O'Neill, 2008). Resolvins have been shown to regulate specific miR-target genes involved in inflammation and resolution . These include miR-21, miR-146, miR-208 and miR-219, which represent a new class of pro-resolving miRNAs.
Results from Serhan and coworkers help to identify the possible pathways and lead to a hypothetical scheme for RvE1/ChemR23 dependent signaling in human macrophages Oh et al., 2011;Ohira et al., 2010). RvE1 regulates phosphorylation of Akt and ribosomal protein rS6 via RvE1-specific interaction with ChemR23 on both human ChemR23-transfected CHO cells and human macrophages enhancing phagocytosis (Ohira et al., 2010). A decrease in p42 and p44 MAP kinase phosphorylation, induced by a bacteria, is also observed when neutrophils in culture are pretreated 15 min before challenge bacteria with 100 ng/ml RvE1 (Herrera et al., 2015).

Conclusion
More studies are needed to understand the actions of resolvins in the central nervous system. Indeed, resolvins are promising therapeutic compounds: these mediators are of natural origin and are active at very low concentrations (nM) as compared to their precursors (µM) (Ariel and Serhan, 2007;Bannenberg and Serhan, 2010). Resolvins administered orally to mice reduce acute inflammation and accelerate or initiate resolution (Recchiuti et al., 2014). These results highlight the possibility to exploit the beneficial effect of RvD1 in Human. Resolvins open novel strategies for the treatment of inflammatory diseases.