Método Integrado

Arginine and glutamine, among other essential amino acids, have a relevant role in growth and immunity because they are important regulators in different metabolic pathways (Wu et al., 2007). Under hypermetabolic states (e.g. inflammation, endotoxin infection, etc.) dietary supplementation with those amino acids helped to preserve intestinal mucosa structure and function, and to support normal immune response (Duggan et al., 2002; Ziegler et al., 2003). This has stimulated the interest and promoted research to use these amino acids in farm animals aiming to enhance health and production, especially during stressful periods as weaning. Weaning stress, associated with villous atrophy and immunosuppression, also increased the metabolism of arginine and glutamine in the intestinal epithelial cells of pigs (Wu et al., 1994; Flynn and Wu, 1997) and enhanced its requirements. Reduced feed intake during the early weaning period may exacerbate these deficiencies leading to bacterial translocation, gut atrophy, mucosal shedding and weight loss. Supplementation with arginine and glutamine appear to reverse these effects by enhancing total gut weight and preventing villous atrophy (Ewtushik et al., 2000). Inflammation and infection change the nutritional requirements, particularly of proteins and amino acids. In these circumstances, the organism may redirect the flux of nutrients to the tissues implicated in inflammatory and immune responses at the cost of those needed for growth (Le Floc’h and Séve, 2000).

40 Experimental results suggest that the profile of amino acids required for the immune system differs substantially from those for growth (Reeds et al., 1994; Klasing and Calvert, 2000).

Arginine plays a key role in the metabolism of amino acids via the urea cycle, enabling the disposal of the nitrogen excess from the amino acids that cannot be used for anabolic purposes. Moreover, increase cell size and protein synthesis (Naomoto et al., 2005), it is essential for the release of growth hormones (Wu et al., 2007), and it is the precursor for the nitric oxide which is catabolized by the nitric oxide synthase. Nitric oxide is a key regulator of the immunity response, and for polyamines synthesis which are essential in tissues that are actively synthetizing proteins, but also act as a mediator in the histological development of enterocytes (Piva et al., 2002). The inducible isoform of the nitric oxide synthase (iNOS), is produced in macrophages, hepatocytes, in the vascular smooth muscle cells and in the endothelia in response to endotoxins, cytokines and other inflammatory factors (Tapiero et al., 2002), that could improve the intestinal health state. In humans, there is clinical evidence suggesting that arginine participates in the regulation of inflammation and enhances the immune response of patients suffering injury, surgical trauma, malnutrition or sepsis (Suchner et al., 2002). In piglets, dietary supplementation with arginine improves their immune status (Tan et al., 2007). A positive effect of dietary arginine supplementation against Eimeria spp. infection has also been reported in chicks (Allen, 1999; Kidd et al., 2001). Several authors observed that oral supplementation with 150-200 mg/kg/day of arginine and 150-200 mg/kg/day of glutamine together resulted in certain beneficial additive effects (Kul et al., 2009; Zhou et al., 2012). Particularly, growing rabbits fed with diets supplemented with 1% of glutamine and 0.5% of arginine tended to improve growth performance, reducing the presence of Clostridium spp. and Helicobacter ssp. in the caecum and in the ileum (Chamorro et al., 2010). Ex vivo experiments suggest that giving arginine and glutamine together decrease the production of pro-inflammatory cytokines (Lecleire et al., 2008).

Otherwise, there are metabolic and age-depending differences between species amino acids requirements especially for arginine. In this way, previous works in young rabbits showed that arginine requirements were particularly high compared to adult animals (Adamson and Fisher, 1976), and higher than in other mammalian species including human, pig and rat (Adamson and Fisher, 1973).

41 Glutamine plays an important role in the metabolism of the intestinal mucosa, because it is necessary for the synthesis of glycoproteins that are secreted by the intestinal mucosal cells (Wu et al., 2001; Wang et al., 2006), maintaining intestinal barrier integrity and functionality (Wu, 1998). In this way, the dietary supplementation with 1% of glutamine enhanced jejunal villi length during the first week after weaning in early-weaned piglets (Wu et al., 1996). Furthermore, in apparent healthy rabbits, there are a trend to reduce lesions caused by developing oocyst (Eimeria spp.) in the villi of the jejunum (Chamorro et al., 2010). Moreover, several studies have demonstrated beneficial effects of supplying glutamine on preventing the bacterial translocation in diverse experimental models of challenged animals by reducing intestinal permeability (Chun et al., 1997; White et al., 2005). In this context, glutamine provides nitrogen for the synthesis of amino sugars, playing a critical role on intestinal mucins synthesis and hence in the maintenance of the passive barrier that limits bacterial colonization of intestinal mucosa (Khan et al., 1999) and on the maintenance of the tight junctions (Panigrahi et al., 1997;

Li et al 2004). Glutamine also is absorbed by the small intestine, representing the major energy source for enterocytes (Wu et al., 1998), being essential for proliferation of cells (Rhoads et al., 1997; Curi et al., 2005) including intraepithelial lymphocytes and macrophages (Wu, 1996; Yoo et al., 1997) and for the synthesis of other non-essential amino acids as nucleotides. Furthermore, the role of glutamine as an immunomodulatory and anti-inflammatory is well-recognized (Van der Hulst et al., 1997; Newsholme et al., 1999). All the diverse cells of the immunity system including monocytes, macrophages, lymphocytes and neutrophils appear to be dependent on glutamine for their functionality (Newsholme et al., 1999), especially during disease. Moreover, glutamine reduced the pro-inflammatory response, reducing the expression of IL-6 and IL-8 and increasing the IL-10 anti-inflammatory response in the gut (Coeffier et al., 2001, 2003). Glutamine is also essential in the glutathione synthesis, which is the most abundant antioxidant in the small intestine (Wu et al., 2004b) and increased the expression of genes that prevent oxidative stress (Wang et al., 2008a). In growing rabbit’s supplementation with 1%

glutamine, decreased fattening mortality and promoted a modification in the intestinal microbiota decreasing the frequency of Helicobacter spp. in the ileum and caecum and Clostridium spp. in the ileum (Chamorro et al., 2010). The supplementation of glutamine has also proved to be effective in calves using an Escherichia coli model (Brooks et al., 1997) and in broiler challenged with Eimeria maxima (Yi et al., 2005). However, there are some studies showing no effects of glutamine supplementation in other disease

42 models (Naylor et al., 1987; Drackey et al., 2006). The effectiveness of glutamine may depend on whether the digestive insult challenges the immune system or not.

Figure 3. Possible mechanisms responsible for the beneficial effect of glutamine on intestinal barrier function and growth. Abbreviations: F-6-P, fructose-6-phosphate; Gln6-P, N-acetilglucosamine-6-phosphate. The symbol (+) denotes activation. (source: Wu et al., 2007)