The alternative pathway of macrophage activation, via IL-4, has been recently appreciated in many Th2-associated responses, such as parasitic infections and allergic inflammation. However, the role of these cells and their effectors in the broad spectrum of Th2-associated pathological processes is poorly understood. We investigated the role of the enzyme arginase, which is a marker of alternatively activated macrophages and highly upregulated in allergic and parasitic infection. While the role of arginase in the urea cycle is well understood, its role in inflammation is less clear. Based on known downstream mediators, arginase has been speculated to play a role in NO production and subsequent inflammation and regulation of airway tone, as well as proliferation and collagen synthesis via ornithine production. We used arginase I-deficient bone marrow chimeric mice in models of allergic airway inflammation. Use of arginase I BM chimeric mice revealed several novel findings. First, we demonstrate that arginase I expression does not affect bone marrow reconstitution following transfer into lethally irradiated recipients and that arginase is not required for baseline immunity. Second, we demonstrate that BM-derived arginase I is the main source of allergen-induced lung arginase. Third, we found that lung arginase is dispensable for regulation of inflammation, airway tone, fibrosis and cell proliferation during allergic airway inflammation.
Previous studies have suggested that macrophages are the main source of arginase in the allergen challenged mouse lung. However, other cell types, including respiratory epithelial cells have been implicated as potential sources of arginase [
7,
38‐
40]. Furthermore, we and others have shown that both arginase I and II expression is increased in the lungs following allergen challenge [
7]. Thus, it remained possible that arginase II will contribute to lung arginase activity in allergen challenged arginase I BM chimeric mice. Our studies measuring arginase activity in both arginase I BM chimeric and arginase II-deficient mice demonstrated that arginase I, expressed in bone marrow cells, is the main contributor to arginase activity in allergic lung. These data support the notion that bone marrow-derived cells, such as infiltrating inflammatory macrophages, are the main source of the arginase I enzyme, and arginase activity in the allergic lung.
We originally hypothesized that arginase induction has a role in the pathophysiology of allergic airway inflammation. This was based on several findings. First, we and others have shown that arginase expression and function is increased in the lungs of allergen challenged mice and in the lungs and serum of humans with asthma [
7,
41]. However, these studies did not address the specific role of arginase in pathophysiology of the disease. Second, downstream pathways of arginase have been implicated in the regulation of inflammation, collagen deposition, proliferation and airway hyperresponsiveness [
35,
42,
43]; these are all hallmark outcomes in asthma. Specifically, Meurs et al have elegantly demonstrated a role for arginase in contractility of allergen-challenged guinea pig tracheas
ex vivo. They have also demonstrated that the mechanism for this effect includes arginase-mediated decrease of NO production [
44,
45]. These results, obtained in guinea pigs and in reductionist
ex vivo approaches, differ from our data that were obtained in the complex
in vivo environment of allergen challenged mice. We speculate that the difference lies either in the model system (guinea pigs versus mice) or that parallel pathways are invoked
in vivo during allergen challenge, thus making the role of arginase redundant. Third, mouse models of parasite infestation, including Schistosome mansoni, Heligmosomoides polygyrus, Leishmania sp, Toxoplasma gondii and Nipostrongylus brasiliensis [
11‐
16] demonstrated a role for arginase expression in host defense. Interestingly, in at least one of those publications the role of arginase was confined to models of infection by intracellular pathogens; however, arginase did not play a role in a systemic disease despite induction of arginase in both models [
16]. Finally, we have shown a role for arginase in IL-13 induced airway hyperresponsiveness [
46] and others have recently suggested a role for arginase in mouse and guinea pig asthma models using arginase inhibitors [
17‐
19]. Specifically, we have shown that delivery of arginase I siRNA to mice that receive IL-13 i.t. leads to decreased arginase I expression and airway responsiveness. It is important to note that this effect was observed at early time points (12–48 hours following single i.t. installation of IL-13). Thus, we speculate that arginase may play a role in airway responsiveness to a single cytokine at early time points when collateral pathways have not been able to develop; in contrast, in allergen challenge multiple pathways are activated that may contribute to airway responsiveness thus making arginase redundant. The study by Maarsingh et al [
18], demonstrated that inhalation of ABH, an inhibitor of arginase, protects against allergen-induced airway obstruction, hyperresponsiveness and inflammation. We speculate that our results differ from the ones in this study either because of the different species used (mice versus guinea pigs) or because of the different approach (inhibitors versus genetic ablation of arginase). It is possible that the inhibitor has non-specific off-target effects that may be responsible for the observed effects. It is important to note that arginase plays an important role in the urea cycle in the liver; mutations in arginase leads to severe metabolic consequences in humans and mice, including hyperargininemia, hyperammonemia and premature death [
20,
47]. In an effort to confirm our results, we used an inhibitor of arginase, BEC, and found that it leads to systemic effects, including hyperargininemia (data not shown). Since plasma arginine levels can have profound effects on the immune system [
33,
34,
48‐
54], we elected not to pursue studies with inhibitors as we believed we could not distinguish the effects of arginase inhibition locally in the lungs from indirect effects from arginase inhibition in the liver. In contrast to the study by Maarsingh et al where arginase inhibitor decreased inflammation and airway hyperresponsiveness, Ckless et al [
17], found that inhibitor of arginase led to increased inflammation and airway hyperresponsiveness in mouse models of allergic inflammation. Together, these data caution against use of arginase inhibitors in allergic airway inflammation until the effects (specific and off-target) and mechanisms are fully elucidated.
In summary, our data suggest divergent role for arginase I in allergic inflammation compared to parasitic responses. Since arginase is a prominent product of alternatively activated macrophages, which are induced by IL-4 in both allergic and parasitic responses, our data suggests alternatively activated macrophages evolved to combat parasitic infections and are either bystanders in allergic inflammation or have developed other effector molecules for allergic Th2-associated responses.