Review

Kynurenines: Tryptophan’s metabolites in exercise, inflammation, and mental health

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Science  28 Jul 2017:
Vol. 357, Issue 6349, eaaf9794
DOI: 10.1126/science.aaf9794

From stomach ache to depression

Our gut hurts and we feel miserable. Such disparate phenomena are mechanistically connected, but how? Cervenka et al. review the many pathways taken by dietary tryptophan as it is metabolized into kynurenines. These metabolites distribute into homeostatic networks that integrate diverse aspects of mammalian physiology. Depending on physiological context, kynurenines influence health and disease states ranging from intestinal conditions to inflammation to cancer progression. Further, they can mediate the effects of exercise, mood, and neuronal excitability and, ultimately, communicate with the microbiota.

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Structured Abstract

BACKGROUND

The essential amino acid tryptophan is a substrate for the generation of several bioactive compounds with important physiological roles. Only a small fraction of ingested tryptophan is used in anabolic processes, whereas the large majority is metabolized along the kynurenine pathway of tryptophan degradation. This pathway generates a range of metabolites, collectively known as kynurenines, involved in inflammation, immune response, and excitatory neurotransmission. Kynurenines have been linked to several psychiatric and mental health disorders such as depression and schizophrenia. In addition, due to the close relationship between kynurenine metabolism and inflammatory responses, kynurenines are emerging as recognized players in a variety of diseases such as diabetes and cancer. Because the levels of enzymes of the kynurenine pathway in peripheral tissues tend to be much higher than in the brain, their contribution to the kynurenine pathway can have both local and systemic consequences. Due to their characteristics, kynurenine and its metabolites have the right profile to fill the role of mediators of interorgan communication.

ADVANCES

Understanding how the tryptophan-kynurenine pathway is regulated in different tissues, and the diverse biological activities of its metabolites, has become of interest to many areas of science. The bioavailability of tryptophan can be affected by factors that range from gut microbiome composition to systemic inflammatory signals. Gut-resident bacteria can directly absorb tryptophan and thus limit its availability to the host organism. The resulting metabolites can have local effects on both microbiome and host cells and even mediate interspecies communication. In addition, the biochemical fate of absorbed tryptophan will be affected by cross-talk with other nutrients and even by individual fitness, because skeletal muscle has recently been shown to contribute to kynurenine metabolism. With exercise training, skeletal muscle increases the expression of kynurenine aminotransferase enzymes and shifts peripheral kynurenine metabolism toward the production of kynurenic acid. As a consequence, alleviating the accumulation of kynurenine in the central nervous system can positively affect mental health, such as reducing stress-induced depressive symptoms.

The kynurenine pathway is highly regulated in the immune system, where it promotes immunosuppression in response to inflammation or infection. Kynurenine reduces the activity of natural killer cells, dendritic cells, or proliferating T cells, whereas kynurenic acid promotes monocyte extravasation and controls cytokine release. Perturbations in the kynurenine pathway have been linked to several diseases. High kynurenine levels can increase the proliferation and migratory capacity of cancer cells and help tumors escape immune surveillance. Kynurenine metabolites have been proposed as markers of type 2 diabetes and may interfere at some level with either insulin secretion or its action on target cells. Kynurenines can signal through different tissue-specific extra- and intracellular receptors in a network of events that integrates nutritional and environmental cues with individual health and fitness.

OUTLOOK

The modulation of tryptophan-kynurenine metabolism using lifestyle and pharmacological interventions could help prevent and treat several diseases with underlying inflammatory mechanisms, including metabolic, oncologic, and mental health disorders. In this context, and considering the substantial effect that the gut microbiome can have on preabsorptive tryptophan metabolism, it is tempting to envision the use of probiotic-based therapies. The discovery that aerobic exercise training can reduce kynurenine levels in circulation and in the central nervous system could have important implications for the development of future generations of antidepressant medications. This again stresses the many advantages of remaining physically active throughout life. Understanding the multiple levels of control of the kynurenine pathway could help predict susceptibility to disease linked to environmental and dietary signals.

The kynurenine pathway generates tryptophan metabolites with diverse biological activities throughout the body.

Although mainly studied in relation to the brain and mental health, the action of kynurenine metabolites on peripheral tissues might be even more profound. They serve as important mediators of interorgan and interkingdom cross-talk, connecting seemingly diverse processes such as the effects of exercise training and pathologies such as inflammatory diseases, cancer, and depression.

Abstract

Kynurenine metabolites are generated by tryptophan catabolism and regulate biological processes that include host-microbiome signaling, immune cell response, and neuronal excitability. Enzymes of the kynurenine pathway are expressed in different tissues and cell types throughout the body and are regulated by cues, including nutritional and inflammatory signals. As a consequence of this systemic metabolic integration, peripheral inflammation can contribute to accumulation of kynurenine in the brain, which has been associated with depression and schizophrenia. Conversely, kynurenine accumulation can be suppressed by activating kynurenine clearance in exercised skeletal muscle. The effect of exercise training on depression through modulation of the kynurenine pathway highlights an important mechanism of interorgan cross-talk mediated by these metabolites. Here, we discuss peripheral mechanisms of tryptophan-kynurenine metabolism and their effects on inflammatory, metabolic, oncologic, and psychiatric disorders.

Tryptophan (Trp) is an essential amino acid critical for protein synthesis, but it also serves as substrate for the generation of several bioactive compounds with important physiological roles. Probably the best-known fate of Trp is its conversion to serotonin (5-hydroxytryptamine), an important neurotransmitter involved in the control of adaptive responses in the central nervous system (CNS) and linked to alterations in mood, anxiety, or cognition (1). Serotonin can be further converted to N-acetylserotonin (NAS) and melatonin, adding control over circadian rhythmicity to the list of biological roles for Trp metabolites (2). However, in mammals, the majority of free Trp is degraded through the kynurenine pathway (KP) (Fig. 1) and generates a range of metabolites involved in inflammation, immune response, and excitatory neurotransmission (3). The final product of the KP is nicotinamide adenine dinucleotide (NAD+), an important cofactor in cellular reactions linked to energy metabolism (4) that is emerging as an attractive therapeutic target for several diseases. Here, we focus on peripheral mechanisms that contribute to Trp-KP metabolism.

Fig. 1 Overview of the kynurenine pathway of tryptophan degradation.

3HAO, 3-hydroxyanthranilic acid oxygenase; NAPRT1, nicotinate phosphoribosyltransferase.

Kynurenine (Kyn) and its metabolites (all with defined chemical identities but often collectively called “kynurenines”) are known for their effects on the CNS and have been linked to several psychiatric and mental health disorders such as depression and schizophrenia (5). The CNS receives about 60% of Kyn from the periphery by transport across the blood-brain barrier (BBB), and the remaining is produced locally. Kyn degradation in the CNS is divided between different cell types, among which astrocytes and microglia play important roles with antagonizing actions (6). Microglia produce quinolinic acid (Quin), an N-methyl-d-aspartate receptor (NMDAR) agonist, whereas astrocytes are equipped to generate kynurenic acid (Kyna), an NMDAR and α7 nicotinic acetylcholine receptor (α7nAChR) antagonist. The levels of these two Kyn metabolites have hence been associated with neuronal excitotoxicity (Quin) or protection (Kyna) and are found to be dysregulated in major depressive disorders and schizophrenia (7).

Like Trp and Kyn, 3-hydroxykynurenine (3-HK) crosses the BBB and contributes to Quin generation in microglia but is also able to exert more direct deleterious effects linked to oxidative stress and apoptosis (8, 9). Defects in Kyn signaling have also been seen in mouse models of neurodegenerative diseases such as Alzheimer’s and Huntington’s (10, 11). The underlying feature of these different pathologies seem to converge on neuroinflammation and associated events, including brain infiltration of circulating immune cells, microglia activation, and high levels of proinflammatory cytokines (12).

The levels of enzymes of the KP in peripheral tissues tend to be much higher than in the brain. For example, macrophages have a 20-fold higher capacity to produce Quin than microglial cells. This is particularly important in situations of macrophage infiltration across the BBB. Immune cells are both important sources and targets for Kyn metabolites as they express high levels of several enzymes of the pathway [e.g., indoleamine 2,3-dioxygenase (IDO) and kynurenine aminotransferases (KATs)] and also of receptors such as G protein–coupled receptor 35 (GPR35). The expression of IDO and KATs allow Trp to be metabolized to Kyna, a GPR35 agonist (13) and a ligand for the transcription factor aryl hydrocarbon receptor (AhR) (similar activity has been shown for Kyn) (14, 15). IDO, together with tryptophan 2,3-dioxygenase (TDO) and AhR are present in some tumor cells, so it has been proposed that Kyn can have a double role in promoting cancer invasion and immune escape. On one hand, activation of cancer cell AhR by Kyn increases the expression of genes that promote cell migration (16, 17). On the other hand, activated immune cell AhR suppresses effector T cells and increases immune tolerance by targeting dendritic and regulatory B cells (18).

The gastrointestinal tract (GIT) has an important role in Trp metabolism. The upper GIT is responsible for the majority of serotonin synthesis (19). It also absorbs and is directly influenced by Kyn metabolites such as Kyna that are present in food and can act locally on GPR35 (20). The lower portion of the GIT is home to substantial numbers of microbiota, which are affected by Trp availability and in turn act on gut mucosal tissues and resident immune population through the production of indole compounds that bind to AhR (21).

Skeletal muscle has recently been added to the list of tissues that contribute to Kyn metabolism (22). This happens in the setting of exercise training and depends on the transcriptional coactivator peroxisome proliferator–activated receptor (PPAR) gamma coactivator-1α1 (PGC-1α1), which enhances KAT gene expression and Kyn to Kyna conversion. This links peripheral and central Kyn metabolism and provides a mechanism for some of the benefits of physical exercise for mental health.

The many fates of tryptophan

Humans lack the biochemical pathways to synthesize Trp, which must be acquired from diet with a required daily dose of 3.5 mg per kg of weight (23). The highest concentration of Trp can be found in chocolate, eggs, fish, dairy products, legumes, and meat. Less than 1% of ingested Trp is used for protein synthesis because, under conditions of unaltered nitrogen balance, the demand for protein synthesis is met by protein breakdown (24). The majority of Trp is thus metabolized along one of four known pathways, giving rise to a variety of biologically active compounds (e.g., serotonin, tryptamine, indoles, kynurenines, and NAD+) (25). Trp, together with other neutral amino acids, is transported by large neutral amino acid transporters (LAT) 1 to 4. These are widely distributed throughout the body, and their capacity is sufficient to avoid competition, with the notable exception of the BBB (26). The majority of Trp is imported into the gut, where only a fraction is used, whereas the rest enters portal circulation and undergoes liver metabolism. The remaining Trp, together with its liver degradation products, is distributed to peripheral circulation and transported to tissues such as the brain, heart, and skeletal muscle. Trp not taken up by the upper GIT is metabolized by resident microbiota to indole compounds (27), important interspecies signaling molecules (28).

Trp is the only amino acid transported bound to albumin. However, degradation pathways can only use Trp in its free form, which corresponds to 5 to 10% of total Trp (29). The mode of Trp degradation and concentration of its end products is a function of free Trp concentration, which is in turn readily influenced by nutritional, hormonal, and pharmacological cues. For example, nonesterified fatty acids (NEFA) directly affect Trp availability by displacing it from albumin (30). As a consequence, increasing NEFA levels by, for example, adrenaline or phosphodiesterase inhibitors, increases free Trp. Conversely, antilipolytic agents such as insulin are able to decrease Trp concentration by the same mechanism (25).

The gut microbiota numbers are estimated to outnumber cells in our body by a factor of 10 and tend to increase distally along the intestine (31). Due to their sheer number, they have nonnegligible effects on Trp metabolism. Gut microbiota can directly absorb Trp and thus limit its availability to the host organism. This can be seen in germ-free (GF) mice, which have high circulating Trp levels that normalize postcolonization. Bacterially produced indoles interact with pregnane X receptors (PXRs) and mediate a range of effects, including improved mucosal homeostasis and barrier function, and as such represent a fascinating example of interkingdom communication (32). Moreover, indoles act as hydroxyl radical scavengers, neuroprotectants, and human AhR selective agonists attenuating inflammation (33). Conversely, some bacteria are susceptible to selective serotonin reuptake inhibitors (SSRIs) such as sertraline, fluoxetine, and paroxetine. Initially, serotonin was identified as a player in the peristaltic reflex of the gut and has since been shown to influence colonic morphology, maintenance of enteric mucosa, pellet formation, and propulsive motility. Currently, the role of serotonin in the brain-gut axis through the activity of the microbiota is being avidly explored (21). Thus, the extent of Trp use by bacteria, its dietary supply, and local turnover by the GIT can have far-reaching implications in the development and proper functioning of both the enteric nervous system (ENS) and CNS.

The kynurenine pathway of tryptophan degradation

Decarboxylation of Trp to tryptamine, transamination to indol-3-yl pyruvic acid, and hydroxylation to serotonin are minor Trp degradation pathways. Although serotonin is usually associated with the brain, the majority of its production is localized in the gut. As much as 90% of total serotonin production (stored in secretory granules) comes from enterochromaffin cells and, to a lesser extent, from serotonergic neurons of the ENS (34).

More than 95% of Trp is metabolized along the KP to yield nicotinamide and NAD+ (24) (Fig. 1). The rate-limiting step of the KP is the conversion of Trp to N-formylkynurenine, which is mediated by two spatially segregated members of this pathway. In the liver, the first step in Trp degradation is mediated by TDO, which under normal conditions is responsible for the majority of this conversion (35). TDO is the main determinant of Trp availability to extrahepatic tissues and is inducible by Trp itself, glucocorticoids, and estrogens (36, 37). The extrahepatic branch of the KP is under the control of two IDO enzymes (IDO1 and the more recently discovered IDO2), whose activity is negligible under basal conditions but dramatically inducible by several stimuli, such as inflammatory signals (e.g., interferon-γ). IDOs are mostly active in the immune system and mucosal tissues such as gut (38, 39). Conversely, IDO can be inhibited by elevated levels of Trp, which results in channeling the flux of Trp degradation back to TDO (40). Interestingly, the TDO and IDO genes do not share a common ancestor but are an example of functional convergence (41).

The KP can yield metabolites with neurotoxic and neuroprotective properties, depending on which enzyme tips the conversion scales (5). Under normal conditions, the majority of Kyn is excreted in the urine, so its bioavailability only increases when the flux of Trp down the KP exceeds renal clearance (42). Kyn is usually hydroxylated to 3-HK and then further converted to 3-hydroxyanthranilic acid (3-HAA). 3-HAA is rapidly converted to Quin by the nonenzymatic reaction of an intermediary product and proceeds with conversion to NAD+, a preferred end product of the KP (43). Under specific conditions, picolinic acid (PA) is formed instead. The other branch of the pathway, leading to the production of Kyna and xanthurenic acid (XA) from Kyn is minor under normal conditions but increases under Trp or Kyn loading (Fig. 1) (44). These ratios change dramatically under different Trp loads and are also influenced by vitamin B6 availability (45). Our understanding of how the proportions of different Kyn metabolites change with environmental context is incomplete, and many contradictory results have been reported. Interestingly, this has prompted the development of mathematical models to help us understand metabolite flux through the KP (46).

Conservation of kynurenine metabolism throughout evolution

In bacteria, fungi, and plants, the biosynthesis of aromatic amino acids such as Trp is provided by the shikimate pathway. Whereas bacteria spend the majority of their metabolic energy on protein synthesis, plants use this pathway to generate a large variety of secondary metabolites (47). Regarding the conservation of the KP, the TDO enzyme can be found in the majority of bacterial species and in almost all metazoan species but has probably been lost in fungi during the course of evolution (Fig. 2) (48). IDO enzymes have been discovered in vertebrates, and—even though its homologs have been confirmed in other species, such as molluscs, yeasts, or deuterostomian invertebrates—they remain poorly characterized. On the other hand, arthropods and nematodes generally lack IDO enzymes. The analysis of more downstream components of KP shows that vertebrates, yeasts, and some invertebrates can generate NAD+ through the KP, with TDO serving as a main supplier of precursors and IDO having a different role. Insects and invertebrates lack some of the downstream KP enzymes, suggesting that alternate means of NAD+ generation have prevailed throughout evolution (Fig. 2). TDO is regarded as a “high catalytic activity” enzyme in contrast to IDO; however, this seems to be different in fungi where IDO assumed the lost TDO functionality. In vertebrates, IDO1 acquired high affinity for Trp after IDO1/IDO2 divergence. The biological importance of “low-catalytic activity” IDOs remains unclear and controversial, although they have been well conserved throughout evolution. KAT enzymes have also been somewhat conserved during evolution and can be found in prokaryotes, insects, nematodes, and vertebrates. However, some species contain multiple genes (nematodes, 2; humans, 4), which could have arisen during evolution by means of gene duplication. Because the individual KAT enzymes display different tissue profiles, it might be that their functions and localizations have become specialized with time. Plants possess the enzyme tryptophan aminotransferase–related (TAR1) that plays a role in the synthesis of auxin but is also able to transaminate Kyn to Kyna (49).

Fig. 2 Evolutionary conservation of enzymes of the kynurenine pathway.

KYNU, Kynureninase.

Kynurenic acid

Of all the different by-products of the KP, Kyna has been studied most. It was originally discovered in canine urine, but higher concentrations have been measured in the gut (increasing gradually along its length), bile, pancreatic juice of rats and pigs, and, to a lesser extent, in human saliva and synovial and amniotic fluid (20). Its presence in many food products has also been determined. The highest concentrations are found in honeybee products, broccoli, and some potatoes (50). Many medicinal herbs contain high concentrations of Kyna, indicating therapeutic potential for the gastrointestinal system (51). In addition to the Kyna ingested in food or synthesized along the KP in the GIT, the gut microflora possesses the enzyme aspartate aminotransferase, which is analogous to mitochondrial KAT4 and produces Kyna by transamination of Kyn. The action of Kyna on the GIT is several fold. Early reports suggested that Kyna is able to protect the intestinal mucosa in the settings of obstructive jaundice and protect from ethanol- or toxin-induced ulcers (52). In addition, Kyna can also modulate local inflammation, most likely through activation of GPR35, which is highly expressed in the immune cells of the GIT.

Liver control of tryptophan-kynurenine metabolism

Among the many cell types that express KP enzymes, hepatocytes contain all the machinery required for Trp degradation toward any branch of the KP (53). Most important, they are the sole cell type with high TDO activity and thus have a central role in the modulation of systemic Trp levels (35). Because TDO has low affinity for Trp, it remains active even when Trp exceeds the levels required for serotonin and protein synthesis (54). If the requirements for protein synthesis are surpassed, the liver metabolizes excess Trp to NAD+, oxidizing the rest via the glutarate pathway (Fig. 3). If Trp concentration is low, the liver will clear and metabolize circulating Trp to NAD+ for energy demands (53). Interestingly, when Trp requirements are met and liver TDO activity is increased by glucocorticoids, saturation of the pathway will lead to leakage of some of its metabolites, such as Kyn.

Fig. 3 Activity, uptake, and conversion of tryptophan and its metabolites in peripheral tissues during unchallenged conditions.

5-HT, 5-hydroxy tryptamine; ROS, reactive oxygen species.

Of note, factors that are detrimental to mental health, such as stress, social isolation, sleep deprivation, and lack of physical activity, elevate circulating glucocorticoid levels in both humans and nonhuman social mammals (55, 56). This will lead to a feed-forward loop in which liver Kyn metabolism increases the output of KP substrates from the periphery to the CNS. As mentioned before, these compounds can be degraded locally to metabolites with deleterious effects to the CNS (7). Chronically high cortisol levels create a state of glucocorticoid receptor (GR) resistance, which in turn fails to dampen the inflammatory response. The reduction in GR activity leads to suppressed liver TDO expression and a shift to extrahepatic Trp and Kyn metabolism (42). One reason for this shift in Trp degradation could be to promote the immunomodulatory roles of kynurenines and reduce inflammation.

Immune system

In immune cells, as in most extrahepatic tissues, the KP is initiated by IDO. This enzyme is ubiquitously expressed and has affinity for substrates other than Trp, including 5-hydroxytryptophan and serotonin. IDO is highly regulated in the immune system, where its expression is increased by interferon-γ (IFN-γ), tumor necrosis factor α (TNFα), and pathogenic infections (57, 58). IDO-mediated Trp catabolism in the host microenvironment surrounding parasites, viruses, and bacteria was seen as a way to curb their proliferation (59). However, immune cells can also contribute to Trp degradation during nonpathogenic inflammation, indicating that IDO has a broader spectrum of activity on immune cell regulation (60). An unrestrained immune response would be detrimental, so cells have developed metabolic pathways to control immune activation (61). Thus, IDO activity is stimulated by type 1 or proinflammatory cytokines (62) and inhibited by type 2 or anti-inflammatory cytokines (63). Trp degradation by IDO has emerged as a rate-limiting step for metabolic immune regulation, according to two proposed mechanisms: first, by the generation of Trp metabolites with immune activity, such as Kyn and Kyna (64); second, by triggering an amino acid–sensing signal in cells undergoing Trp depletion (65). Initial observations showed that Kyn metabolites, in particular Kyn itself, suppress the activity of natural killer cells (NKT) (66) and antigen-presenting cells (APC) such as dendritic cells (DC), monocytes, and macrophages in mice (67, 68). Furthermore, Kyn blocks T cell proliferation and induces T cell death (69), and IDO-mediated Kyn production in DC leads to the proliferation of regulatory T cells (Tregs) (67, 70). These effects are, at least in part, mediated by Kyn activation of AhR (Fig. 3) (15, 71), a ligand-activated transcription factor involved in xenobiotic response to foreign substances. It is expressed in cells of both innate and adaptive immune systems, and it has been shown to have anti-inflammatory activity in mice (72, 73). Given the Kyn-AhR–mediated decrease in immune surveillance, regulating the Kyn pathway has become an attractive target for cancer therapy. Interestingly, the Kyn-AhR axis has been postulated to constitute one of the links between chronic inflammation and tumor progression (15). Similarly, Kyna was also found to activate AhR (14), but whether this activation leads to similar immune regulation remains unclear.

Kyna has been recently shown to also be a ligand for GPR35. This receptor is expressed in human CD14+ monocytes, T cells, neutrophils, DCs, eosinophils, basophils and invariant NKT (iNKT) cells (13, 74). Of those, circulating monocytes display the highest expression of GPR35, and its interaction with Kyna has been shown to promote monocyte extravasation (75). It was later confirmed that Kyna-GPR35 interaction reduces the inflammatory response induced by lipopolysaccharide (LPS) stimulation in monocytes and macrophages (76) and controls cytokine release in human iNKT cells (74). Taken together, the effects of Kyna on immune cell activation might represent a direct anti-inflammatory mechanism that further reinforces the immunosuppressant function of Trp catabolism. Other metabolites of the KP, such as 3-HAA and Quin, have been shown to induce apoptosis of type 1 T helper (TH1) cells, while promoting proliferation of type 2 T helper (TH2) cells (67). This immune shift would favor cell survival against the deleterious effect of uncontrolled immune activation.

Inflammatory conditions are characterized by high levels of cellular stress and energy use, often accompanied by increased rates of DNA damage. In macrophages, as in the liver, oxidation of Trp through the KP can replenish NAD+ levels to meet energy requirements (77). In addition, NAD+ is used by poly (ADP-ribose) polymerase (PARP) in DNA repair mechanisms (78). Trp catabolism in immune cells is therefore a negative feedback mechanism that suppresses ongoing inflammatory response. However, situations such as chronic low-grade inflammation can lead to a robust elevation of circulating Kyn levels, which will also increase in the CNS. The discovery that skeletal muscle can contribute to the KP (22) is highlighted by the fact that sedentary lifestyles can lead to chronic low-grade inflammation in this organ. This adds a new regulatory node to Kyn metabolism in the periphery.

Skeletal muscle

The effect of skeletal muscle and exercise on nutrient metabolism has been appreciated for a long time. Aerobic exercise training elevates the levels of PGC-1α1 in skeletal muscle of mice and humans. PGC-1α1 is a transcriptional coactivator important for adaptive responses in many tissues (79), most notably in skeletal muscle. When activated, PGC-1α1, together with PPARα/δ, increases skeletal muscle expression of KAT enzymes and shifts peripheral Kyn metabolism toward the production of Kyna. Under these conditions, Kyna does not cross the BBB, so reducing Kyn levels in the brain can reduce stress-mediated effects underlying depressive symptoms. In fact, mice with transgenic expression of PGC-1α1 in skeletal muscle (and therefore higher skeletal muscle KAT levels) are resilient to developing depressive-like behavior caused by elevated Kyn levels resulting from stress or exogenous administration (Fig. 4) (22). This pathway has been shown to be active in both mouse and human muscle (22, 80). Modulating PGC-1α1-PPAR activation in skeletal muscle could become a new therapeutic intervention to regulate Trp-Kyn metabolism. In particular, this would preserve the immunosuppressant action of kynurenines via Kyna and decrease the neurotoxic effects associated with Kyn during chronic inflammatory states.

Fig. 4 Activity, uptake, and conversion of tryptophan and its metabolites in peripheral tissues during disease states and challenges to homeostasis.

Skeletal muscle KP is likely to be affected by other amino acid metabolic pathways. For example, during physical exercise, skeletal muscle can oxidize branched-chain amino acids (BCAA) and Trp for energy (81). Muscle fibers contain all the necessary transporters/carriers for amino acid clearance; however, circulating BCAA compete with Trp and kynurenines for the same transporters (82). Moreover, BCAA inhibit some of the enzymes of the KP (83), especially KATs (84), which indicates that fuel availability (in particular, BCAA) can affect skeletal muscle Trp metabolism and Kyn clearance.

Kynurenine metabolites and disease

Trp metabolism is most widely known and studied in relation to disorders of the nervous system. Its effect on stress-related depression, schizophrenia, and Alzheimer’s and Parkinson’s diseases have been comprehensively reviewed elsewhere (85). The following sections summarize recent advances in our understanding of how Kyn metabolism is dysregulated in peripheral tissue dysfunction. However, it is important to remember that the majority of defects of Trp metabolism in peripheral organs can also have a strong effect on the CNS, resulting in complications such as anxiety and depression.

Irritable bowel syndrome and disease

Two main diseases of the GIT are associated with Trp metabolism: irritable bowel syndrome (IBS) and irritable bowel disease (IBD). IBS is characterized by abdominal pain together with altered bowel habits and affects a considerable portion of the adult population (15 to 20%) (86). Increase in serum-free Trp has been documented in IBS patients. The etiology of IBS has been connected to abnormal serotonergic neuronal signaling during development and also to alterations in serotonin production and signaling in enterochromaffin cells (87, 88). One of the hallmarks of the disease, visceral hypersensitivity, is thought to occur as a result of the sensitization of afferent neurons and to compromised epithelial integrity, which in turn makes it possible for intraluminal compounds to cross the gut wall barrier (89).

By contrast, IBD is a relapsing inflammatory condition with complex etiology that affects 1 in 500 individuals, peaking around the age of 20. The etiology of IBD lies at the intersection of dysbiosis of microbiota, host immunity, and genetic predisposition, with anxiety and depression as common comorbidities (90). In this context, it is not unexpected that aberrant Trp metabolism is a common denominator of these complications. IBD patients have increased plasma levels of Kyn and Kyna, probably as a result of increased IDO expression. IBD is also connected to microbiota homeostasis, and IBD patients also have increased risk of colorectal cancer (91). Recently, caspase recruitment domain–containing protein 9 (CARD9) has been found to be an IBD susceptibility gene. CARD9 encodes a host adaptor protein critical for immune responses against microorganisms. Microbiota derived from CARD9 knockout (KO) mice have compromised Trp to indole conversion and cannot activate AhR. Furthermore, bacteria from CARD9 KO animals are sufficient to induce colitis in wild-type germ-free animals. Colonization of CARD9 KO animals with bacteria capable of catabolizing Trp alleviates the disease (92). Due to the tight regulation between microbiota and host responses, individual contributions will prove challenging to distinguish. Interestingly, plant-derived indole compounds have been used in traditional medicine to treat IBD, which lends support to the importance of the interactions of kynurenines with AhR and their actions on the immune system (93).

Pancreatitis

One of the less-studied diseases with connection to Trp metabolism is acute pancreatitis (AP). It is a severe sterile inflammation of the pancreas connected to gut dysfunction that can lead to multiorgan failure with very high mortality rates (94). Plasma Kyn of AP patients seems to originate in the gut-associated lymphoid tissue (GALT), and its levels correlate with magnitude of injury and systemic inflammatory burden (95). Kynurenine-3-monooxidase (KMO) is central to the pathogenesis of pancreatitis, and its genetic ablation or pharmacological inhibition conveys protection from deleterious effects. The pathology of AP is mediated by 3-HK transported in the mesenteric lymph to other organs such as the lungs, where it causes near total cell death by oxidative stress, apoptosis, and pathological protein cross-linking (96). Decreasing 3-HK production from Kyn by diverting it to generating Kyna inhibits LPS-induced TNFα secretion and leads to increased survival in rodent models of acute pancreatitis (96). Melatonin, another Trp metabolite, has emerged as a treatment option of AP by reducing oxidative stress and protecting from inflammation (97).

Cancer

Cancer cells have a multifaceted relationship with altered Trp metabolism. Several tumor types show increased Trp uptake [as evidenced by α-[11C]-methyl-l-tryptophan (AMT)–positron emission tomography (PET) scanning of human patients], which in turn correlates with poor disease prognosis (98). Although the reason for this is not fully understood, tumor cells might need high Trp levels to fuel an ever-increasing demand for protein synthesis. On the other hand, Trp starvation will induce general control nonderepressible 2 (GCN2) kinase that inhibits G1 to S transition, inducing cell cycle arrest (99). To prevent this, tumors might increase their Trp supply by up-regulating LAT1 expression. However, LAT1 has been shown to work in a bidirectional manner, exchanging glutamate for Trp. To counteract this shortcoming, it has been shown that proliferating tumor cells, but not resting T cells, up-regulate expression of the glutamate transporter by the activating transcription factor 4 (ATF4) pathway after sensing Trp unavailability (100, 101). Moreover, tumors show enhanced IDO expression, with downstream metabolites, such as Kyn, being able to activate beta-catenin signaling, leading to increased colon cancer proliferation in mice (102). IDO expression in mouse models of ovarian cancer, melanoma, and renal cell carcinoma correlates with increased angiogenesis (103). TDO, on the other hand, has been predominantly connected to the escape from the immune system surveillance and increased migratory capacity. Kyn generated in tumors might be subsequently released into the surrounding milieu, where it can affect a variety of immune cell populations by binding to AhR (Fig. 4). Moreover, it can have autocrine effects and stimulate invasiveness in an AhR-dependent manner (104). Rapid Trp usage and its subsequent local depletion results in proliferative block of T cells (99). Of note, certain types of macrophages can suppress T cell proliferation in the same manner (105). Production of NAD+ pathway intermediates can also induce apoptosis in a variety of immune cells (67).

NAD+ is an important cofactor involved in genome stability, stress tolerance, and metabolism (4). Probably for a combination of all those reasons, tumor cells have broadly altered NAD+ use and production. It has been postulated that, in tumor cells, most NAD+ comes from salvage pathways. However, they also possess the ability to shift to de novo production using Trp as a source. This becomes relevant in the setting of NAD+-depleting anticancer drugs, irradiation, or induction of oxidative stress by alkylating agents. For example, it has been shown that reducing Trp uptake leads to rapid NAD+ depletion by PARP, resulting in apoptotic cell death of lung cancer cells mediated by NAD(P)H quinone dehydrogenase 1 (NQO1) (106). On the other hand, tumors elevate expression of quinolinate phosphoribosyltransferase (QPRT), which protects them from oxidative stress by converting Quin to NAD+. Increased QPRT expression is generally associated with poor disease prognosis in humans (107). At the time of writing, several modulators of the KP or analogs of Trp metabolites are undergoing clinical trials for cancer treatment (108).

Diabetes

The main links between diabetes and Trp metabolism are inflammation and immune suppression. Additionally, increased production of serotonin has been implicated in the pathogenesis of diabetes (109). Glucocorticoid-mediated insulin resistance elevates the synthesis of serotonin in the liver and adipose tissue in a tryptophan hydroxylase 1 (Tph1)–dependent manner. Serotonin binds to mechanistic target of rapamycin (mTOR), increasing liver lipogenesis and impairing insulin signaling in adipocytes. Accordingly, inhibition of serotonin degradation by monoamine oxidase A (MAOA) exacerbates the effects (110). Compounding the problem, serotonin has well-documented effects on brain-mediated control of appetite. Despite its anorexigenic effects, serotonin transporter (SERT) KO animals are obese and, conversely, Tph1 and 2 KO mice lose body weight (111). Several studies have highlighted the role of serotonin in the regulation of white and brown adipose tissue energy storage and expenditure. Serotonin can increase fat accumulation in humans and rodents, and the activation of its receptors in hypertrophied fat cells induces adiponectin production. Conversely, Tph1 KO mice have significantly lower weight, improved glycemic control, enhanced energy expenditure, and lower adiposity when on high-fat diets (112).

In the Torii rat model of spontaneous diabetes, decreases in Trp and Kyn production were identified as biomarkers of a prediabetic state. Morbidly obese patients have lower circulating levels of Trp and higher Kyn/Trp ratios (113). Chronic stress and low-grade inflammation are major risk factors in prediabetes to diabetes transition. They can skew the balance of Trp metabolism toward Kyn, 3-HK, and Kyna, both by activating TDO/IDO and by reducing the availability of pyridoxal-5-phosphate, a necessary cofactor for many KP enzymes. This diverts the system away from NAD+ production and instead generates a compendium of molecules with substantial biological effects (114, 115). Diabetic patients show increased levels of XA and Kyna in urine, which have been consequently suggested as biomarkers for type 2 diabetes mellitus (T2DM) (116, 117). Moreover, Trp metabolites inhibit both proinsulin synthesis and glucose- and leucine-induced insulin release from rat pancreatic islets, and XA in particular binds to circulating insulin and prevents its action on target cells (118). Recently, Kyn-AhR signaling in mice has been suggested to play a role in the etiology of obesity stimulated by transforming growth factor–β1 and Toll-like receptor 2/4 signaling pathways (119).

Trp supplementation in rats, on the other hand, suppresses hyperglycemia and increases energy expenditure and insulin secretion when administered together with glucose (120). It also leads to decreased glucose absorption from the intestine and increased glucose uptake to adipocytes (Fig. 4). Interestingly, oral Trp is more effective than intraperitoneal administration. Unlike serotonin, tryptamine has been shown to increase insulin-stimulated glucose uptake into adipocytes. In rats with hereditary T2DM, consumption of Trp-rich chow at a young age protects beta cells from exhaustion in old animals. However, since Trp conversion to tryptamine is very low, a substantial increase in dietary Trp is needed to increase the biological availability of tryptamine (120). How this influences the activity of other Trp degradation pathways and the concentration of their biologically active products has not been investigated.

During the course of many of the aforementioned diseases and in the process of aging, the intracellular levels of Trp and NAD+ can fall if the KP is stressed by inflammation or imbalance between catabolic and anabolic substrates. However, de novo synthesis from Trp is not a very efficient way to boost NAD+ levels, because it requires very high Trp to saturate other branches of the catabolic pathway (4). Nevertheless, NAD+ boosting strategies seem to be successful in restoring mitochondrial and stem cell function (121) and in ameliorating diseases such as muscular dystrophy and diabetes (122, 123) or even prolonging life span (124). Our understanding of age-related effects of the KP is still largely incomplete. However, there is a general notion that alterations of the KP can bring about oxidative stress, immune response decline, or inflammation. Lower levels of Trp and TDO activity have been reported in the brain of aged rodents; however, contrary to expectations, the amount of downstream KP metabolites is increased (25). This could be in part due to the activity of IDO, initiated by proinflammatory cytokines. Interestingly, depletion or inactivation of TDO in Caenorhabditis elegans or Drosophila melanogaster increases life span (125, 126).

Microbiome, mycobiome, and virome

In the GIT, there is a complex interkingdom regulatory network and cross-talk occurring between the host, the microbiome, and the mycobiome. Fungal and bacterial commensals coexist in a complex milieu, and their interactions can have far-reaching implications for pathogenicity (127). To deal with fungal infection, immune responses need to be regulated in a way that limits tissue damage and preserves the commensals. Overactive inflammation primes the gut for fungal colonization, which leads to further inflammation and propagation of this vicious cycle. Bacteria produce indole derivatives, which have been shown to activate interleukin-22 (IL-22)–producing innate lymphoid cells (ILC3) in mice. In turn, IL-22 can increase mucosal protection by driving the production of antimicrobial peptides (18, 33). IL-22 also seems to have an indirect effect by mediating survival of mixed microbial populations, which prevents colonization by opportunistic pathogens (128). Tolerogenic DCs convert Trp to Kyn, promoting immune tolerance through expansion of Treg cells in humans (Fig. 4) (129). AhR activation by Trp metabolites during development is required for proper formation of innate lymphoid cells. Of note, fungi possess a constitutively active IDO, although its importance in gut homeostasis has not been sufficiently addressed. In addition to inhibiting the growth of parasites and bacteria, Trp depletion is also associated with antiviral properties. Serotonin depletion by the KP plays a role in neuroinflammation in conditions associated with chronic viral infections [e.g., human immunodeficiency virus (HIV)] (130). HIV, for example, is able to alter mucosal permeability in patients, facilitating microbe invasion leading to potentiated systemic immune activation (131). Enhanced Trp to Kyn metabolism during infection contributes to both immune suppression and to the loss of memory T cells (132).

Conclusions and future perspectives

As the role of Kyn metabolites continues to be explored in different physiologic and disease settings, the relevance of these compounds as important integrators of environmental, metabolic, and immune system signals continues to emerge. In this context, understanding the importance of the gut microbiome for controlling Trp availability and Kyn metabolism could be crucial to better understanding interindividual variability in interpreting nutritional cues. In addition, recruiting skeletal muscle through exercise training to enhance Kyn clearance and improve mental health could have additional, still unknown, consequences. In this context, it is tempting to speculate that situations where skeletal muscle oxidative capacity and PGC-1α1 are reduced (such as with aging or metabolic disease) could negatively affect the KP. Interestingly, there are several known PPAR agonists, which could be explored as potential therapeutic agents to activate the KP in skeletal muscle. This could have applications not only in stress-induced depression but also in other situations where reducing the Kyn burden would be beneficial—for example, as cytostatic drugs, where the benefit would be several fold. Inhibition of the KP would allow for tumor cells that escape immune surveillance to be properly recognized. Additionally, the same intervention could interfere with cell proliferation, angiogenesis, and metastatic potential and at the same time deprive the tumor of energy by reducing NAD+ production. It may seem that inhibition of a single pathway addresses the majority of cancer progression hallmarks.

As the interest in Kyn metabolites grows, it becomes clear that where you find them conditions their biological activity. While exploring their role in the brain and processes affecting mental health, kynurenines are being rediscovered in peripheral tissues where they induce local and systemic adaptations in both health and disease. We expect the tales of these metabolites to grow in the years to come.

References and Notes

  1. Acknowledgments: The authors acknowledge members of the Ruas laboratory for critical reading of the manuscript and funding from the Swedish Research Council, the Novo Nordisk Foundation (Denmark), Karolinska Institutet, the Lars Hierta Memorial Foundation, the Strategic Research Program (SRP) in Diabetes, and the SRP in Regenerative Medicine at Karolinska Institutet.
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