Tryptophan Depletion Research Articles

Tryptophan Intake and Tryptophan Losses in Hemodialysis Patients: A Balance Study.

Tryptophan depletion is common in hemodialysis patients. The cause of this depletion remains largely unknown, but reduced nutritional tryptophan intake, losses during dialysis or an increased catabolism due to an inflammatory state are likely contributors. Currently, little is known about tryptophan homeostasis in hemodialysis patients. We assessed dietary tryptophan intake, measured plasma tryptophan during dialysis, and measured the combined urinary and dialysate excretion of tryptophan in 40 hemodialysis patients (66 ± 15 years and 68% male). Patients had low tryptophan concentrations (27 ± 9 µmol/L) before dialysis. Mean dietary tryptophan intake was 4454 ± 1149 µmol/24 h. Mean urinary tryptophan excretion was 15.0 ± 12.3 μmol/24 h, dialysate excretion was 209 ± 67 μmol/24 h and combined excretion was 219 ± 66 µmol/24 h, indicating only 5% of dietary tryptophan intake was excreted. No associations were found between plasma tryptophan concentration and tryptophan intake, plasma kynurenine/tryptophan ratio or inflammatory markers. During dialysis, mean plasma tryptophan concentration increased 16% to 31 ± 8 µmol/L. Intradialytic increase in plasma tryptophan was associated with a lower risk of mortality, independent of age, sex and dialysis vintage (HR: 0.87 [0.76–0.99]; P = 0.04). Tryptophan intake was well above the dietary recommendations and, although tryptophan was removed during dialysis, mean plasma tryptophan increased during dialysis. The cause of this phenomenon is unknown, but it appears to be protective.

Functional consequences of acute tryptophan depletion on raphe nuclei connectivity and network organization in healthy women

Previous research on central nervous serotonin (5-HT) function provided evidence for a substantial involvement of 5-HT in the regulation of brain circuitries associated with cognitive and affective processing. The underlying neural networks comprise core subcortical/cortical regions such as amygdala and medial prefrontal cortex, which are assumed to be modulated amongst others by 5-HT. Beside the use of antidepressants, acute tryptophan depletion (ATD) is a widely accepted technique to manipulate of 5-HT synthesis and its respective metabolites in humans by means of a dietary and non-pharmacological tool. We used a double-blind, randomized, cross-over design with two experimental challenge conditions, i.e. ATD and tryptophan (TRP) supplementation (TRYP+) serving as a control. The aim was to perturb 5-HT synthesis and to detect its impact on brain functional connectivity (FC) of the upper serotonergic raphe nuclei, the amygdala and the ventromedial prefrontal cortex as well as on network organization using resting state fMRI. 30 healthy adult female participants (age: M ​= ​24.5 ​± ​4.4 ​yrs) were included in the final analysis. ATD resulted in a 90% decrease of TRP in the serum relative to baseline. Compared to TRYP ​+ ​for the ATD condition a significantly lower FC of the raphe nucleus to the frontopolar cortex was detected, as well as greater functional coupling between the right amygdala and the ventromedial prefrontal cortex. FC of the raphe nucleus correlated significantly with the magnitude of TRP changes for both challenge conditions (ATD & TRYP+). Network-based statistical analysis using time series from 260 independent anatomical ROIs revealed significantly greater FC after ATD compared to TRYP+ in several brain regions being part of the default-mode (DMN) and the executive-control networks (ECN), but also of salience or visual networks. Finally, we observed an impact of ATD on the rich-club organization in terms of decreased rich-club coefficients compared to TRYP+.In summary we could confirm previous findings that the putative decrease in brain 5-HT synthesis via ATD significantly alters FC of the raphe nuclei as well as of specific subcortical/cortical regions involved in affective, but also in cognitive processes. Moreover, an ATD-effect on the so-called rich-club organization of some nodes with the high degree was demonstrated. This may indicate effects of brain 5-HT on the integration of information flow from several brain networks.

Detection of a High-Turnover Serotonin Circuit in the Mouse Brain Using Mass Spectrometry Imaging.

SummaryMonoamine neurotransmitters are released by specialized neurons regulating behavioral, motor, and cognitive functions. Although the localization of monoaminergic neurons in the brain is well known, the distribution and kinetics of monoamines remain unclear. Here, we generated a murine brain atlas of serotonin (5-HT), dopamine (DA), and norepinephrine (NE) levels using mass spectrometry imaging (MSI). We found several nuclei rich in both 5-HT and a catecholamine (DA or NE) and identified the paraventricular nucleus of the thalamus (PVT), where 5-HT and NE are co-localized. The analysis of 5-HT fluctuations in response to acute tryptophan depletion and infusion of isotope-labeled tryptophan in vivo revealed a close kinetic association between the raphe nuclei, PVT, and amygdala but not the other nuclei. Our findings imply the existence of a highly dynamic 5-HT-mediated raphe to PVT pathway that likely plays a role in the brain monoamine system.

Serum Levels of Tumor Necrosis Factor-α and Loudness Dependence of Auditory Evoked Potentials at Pretreatment and Posttreatment in Patients with Major Depressive Disorder.

Background: Proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α), are associated with the pathophysiology of major depressive disorder (MDD). Several studies have reported that increased TNF-α might be associated with tryptophan depletion, which eventually could result in MDD. However, other studies revealed that TNF-α increased serotonin firing in raphe. Therefore, whether TNF-α increases or decreases serotonin activity remains unclear. Here, we aimed to determine the relationship between serum TNF-α level and central serotonergic activity using the loudness dependence of auditory evoked potentials (LDAEP) and standardized low-resolution brain electromagnetic tomography (sLORETA), as well as to evaluate the effects of antidepressants on TNF-α levels. Methods: LDAEP, serum TNF-α level, and depression severity were measured in 64 MDD outpatients pre and post 3 months of treatment. Results: Pretreatment TNF-α levels were negatively correlated with the pretreatment N1 sLORETA-LDAEP, P2 sLORETA-LDAEP, and N1/P2 sLORETA-LDAEP (p < 0.05). In multiple regression analysis for N1/P2 sLORETA-LDAEP, lower N1/P2 sLORETA-LDAEP was significantly related to higher TNF-α (CE = −0.047, p = 0.017) when all subjects were dichotomized based on the median TNF-α level (7.16 pg/mL) into pretreatment low- and high-TNF-α groups. In addition, the pretreatment Beck Depression Inventory, P2 LDAEP, and N1/P2 sLORETA-LDAEP were greater in the high-TNF-α groups than in the low-TNF-α groups (p < 0.05). Moreover, the posttreatment TNF-α level was significantly decreased compared to the pretreatment TNF-α level (z = −2.581, p = 0.01). However, the posttreatment TNF-α levels were not associated with posttreatment LDAEP. Conclusions: Higher TNF-α level is associated with decreased LDAEP, which could indicate compensatory elevation of central serotonin activity in outpatients with MDD, although this effect disappeared and TNF-α level was reduced after three months of antidepressant treatment.

Host cell depletion of tryptophan by IFNγ-induced Indoleamine 2,3-dioxygenase 1 (IDO1) inhibits lysosomal replication of Coxiella burnetii.

Most intracellular pathogens that reside in a vacuole prevent transit of their compartment to lysosomal organelles. Effector mechanisms induced by the pro-inflammatory cytokine Interferon-gamma (IFNγ) can promote the delivery of pathogen-occupied vacuoles to lysosomes for proteolytic degradation and are therefore important for host defense against intracellular pathogens. The bacterial pathogen Coxiella burnetii is unique in that, transport to the lysosome is essential for replication. The bacterium modulates membrane traffic to create a specialized autophagolysosomal compartment called the Coxiella-containing vacuole (CCV). Importantly, IFNγ signaling inhibits intracellular replication of C. burnetii, raising the question of which IFNγ-activated mechanisms restrict replication of a lysosome-adapted pathogen. To address this question, siRNA was used to silence a panel of IFNγ-induced genes in HeLa cells to identify genes required for restriction of C. burnetii intracellular replication. This screen demonstrated that Indoleamine 2,3-dioxygenase 1 (IDO1) contributes to IFNγ-mediated restriction of C. burnetii. IDO1 is an enzyme that catabolizes cellular tryptophan to kynurenine metabolites thereby reducing tryptophan availability in cells. Cells deficient in IDO1 function were more permissive for C. burnetii replication when treated with IFNγ, and supplementing IFNγ-treated cells with tryptophan enhanced intracellular replication. Additionally, ectopic expression of IDO1 in host cells was sufficient to restrict replication of C. burnetii in the absence of IFNγ signaling. Using differentiated THP1 macrophage-like cells it was determined that IFNγ-activation resulted in IDO1 production, and that supplementation of IFNγ-activated THP1 cells with tryptophan enhanced C. burnetii replication. Thus, this study identifies IDO1 production as a key cell-autonomous defense mechanism that limits infection by C. burnetii, which suggests that peptides derived from hydrolysis of proteins in the CCV do not provide an adequate supply of tryptophan for bacterial replication.

MYC promotes tryptophan uptake and metabolism by the kynurenine pathway in colon cancer.

Tumors display increased uptake and processing of nutrients to fulfill the demands of rapidly proliferating cancer cells. Seminal studies have shown that the proto-oncogene MYC promotes metabolic reprogramming by altering glutamine uptake and metabolism in cancer cells. How MYC regulates the metabolism of other amino acids in cancer is not fully understood. Using high-performance liquid chromatography (HPLC)-tandem mass spectrometry (LC-MS/MS), we found that MYC increased intracellular levels of tryptophan and tryptophan metabolites in the kynurenine pathway. MYC induced the expression of the tryptophan transporters SLC7A5 and SLC1A5 and the enzyme arylformamidase (AFMID), involved in the conversion of tryptophan into kynurenine. SLC7A5, SLC1A5, and AFMID were elevated in colon cancer cells and tissues, and kynurenine was significantly greater in tumor samples than in the respective adjacent normal tissue from patients with colon cancer. Compared with normal human colonic epithelial cells, colon cancer cells were more sensitive to the depletion of tryptophan. Blocking enzymes in the kynurenine pathway caused preferential death of established colon cancer cells and transformed colonic organoids. We found that only kynurenine and no other tryptophan metabolite promotes the nuclear translocation of the transcription factor aryl hydrocarbon receptor (AHR). Blocking the interaction between AHR and kynurenine with CH223191 reduced the proliferation of colon cancer cells. Therefore, we propose that limiting cellular kynurenine or its downstream targets could present a new strategy to reduce the proliferation of MYC-dependent cancer cells.

Effects of inflammation modulation on tryptophan and kynurenine pathway regulation in treatment resistant bipolar depression

BackgroundAdjunctive immune-modulation can be safe and effective for treatment-resistant bipolar depression (TRBDD), but molecular work is needed to further characterize the safety and efficacy involved in treatment response and the reversal of treatment resistance. Here we profiled the kynurenine pathway (KP) for biomarkers associated with TRBDD and treatment response to celecoxib (CBX)-augmentation. Methods47 TRBDD patients with moderately severe HAMD-17 scores were randomized to receive either escitalopram (ESC) (10 mg twice daily) + CBX (200 mg twice daily), or ESC (10 mg twice daily) + placebo (PBO) (twice daily). Plasma kynurenine pathway (KP) metabolite levels were measured at baseline, week 4, and week 8, and in a healthy control (HC) group of subjects (N = 35) once. ResultsPatients receiving ESC + CBX had 4.278 greater odds of responding (p = 0.021) with NNT=3, and 15.300 greater odds of remitting (p < 0.001) with NNT=2, compared with ESC + PBO patients. Study patients exhibited elevated baseline tryptophan (p < 0.001), low kynurenine/tryptophan (p < 0.001), elevated 3-hydryoxykynurenine/kynurenine (adj-p*<0.001), low kynurenic acid/3-hydroxykynurenine, and low picolinic acid/quinolinic acid (p < 0.001) compared to healthy controls. Treatment responders exhibited tryptophan depletion (p = 0.020) without a concomitant change in kynurenine/tryptophan ratio by week 8 (p = 0.163). ConclusionClinical response to CBX augmentation is not associated with altered neurotoxic or neuroprotective indices within the time frame of this study. TRBDD revealed alterations in neuroprotective and neurotoxic indices, in the context of low kynurenine/tryptophan and high tryptophan. Treatment responders revealed a depletion in tryptophan by week 8, without concomitant kynurenine pathway (KP) activation.

Opposite Effects of Voluntary Physical Exercise on β3-Adrenergic Receptors in the White and Brown Adipose Tissue.

Catecholamine effects via β3-adrenergic receptors are important for the metabolism of the adipose tissue. Physical exercise is a core component of antiobesity regimens. We have tested the hypothesis that voluntary wheel running results in enhancement of β3-adrenergic receptor gene expression in the white and brown adipose tissues. The secondary hypothesis is that dietary tryptophan depletion modifies metabolic effects of exercise. Male Sprague-Dawley rats were assigned for sedentary and exercise groups with free access to running wheels for 3 weeks. All animals received normal control diet for 7 days. Both groups were fed either by low tryptophan (0.04%) diet or by control diet (0.2%) for next 2 weeks. The β3-adrenergic receptor mRNA levels in response to running increased in the retroperitoneal and epididymal fat pads. The gene expression of uncoupling protein-1 (UCP-1) was increased in the brown, while unchanged in the white fat tissues. Unlike control animals, the rats fed by low tryptophan diet did not exhibit a reduction of the white adipose tissue mass. Tryptophan depletion resulted in enhanced concentrations of plasma aldosterone and corticosterone, but had no influence on exercise-induced adrenal hypertrophy. No changes in β3-adrenergic receptor and cell proliferation measured by 5-bromo-2'-deoxyuridine incorporation in left heart ventricle were observed. The reduced β3-adrenergic receptor but not enhanced uncoupling protein-1 gene expression supports the hypothesis on hypoactive brown adipose tissue during exercise. Reduction in dietary tryptophan had no major influence on the exercise-induced changes in the metabolic parameters measured.

Effects of Acute Tryptophan Depletion on Repetitive Behavior in Laying Hens.

Repetitive pecking at the feather cover of other birds (FP) is one of the most important welfare problems in domestic birds. It is not only characterized by motor symptoms, but also by an innate vulnerability of the serotonergic system. Moreover, the serotonergic system influences cognitive function. Acute tryptophan depletion (ATD) is a widely used method for studying serotonergic function in mammals and has been recently validated in birds. However, a tryptophan-deficient amino acid mixture has never been tested on groups of birds to impact their social behavior, including repetitive feather pecking, nor has it been given to potentially impact their cognition and motor performance. One hundred and sixty White Leghorn laying hens consisting of two genetic lines divergently selected to perform high (H) or low (L) levels of FP, and an unselected control line (UC), were kept in 10 groups consisting of 4 H, 3 L, and 9 UC genotypes. In a counterbalanced order, half of the groups were first subjected to an ATD treatment, while the other half were first given a balanced control (BC) treatment, and vice versa, after which their feather pecking behavior was observed. The effect of ATD/BC on repetitive pecking, motor performance, and cognition was investigated in a 5-s delayed reward task in an operant chamber with 10 phenotypic feather peckers, 10 recipients of feather pecking, and 10 bystanders (who neither performed nor received feather pecks). ATD given to groups of birds induced gentle, repetitive feather pecking in all genotypes. Following ATD, phenotypic feather peckers performed more poorly during the delayed reward task, as seen by their higher number of repetitive, non-rewarded key, and non-key pecks in the operant chamber. In conclusion, ATD impacted the hens' social behavior by increasing the number of repetitive gentle feather pecks at conspecifics. Furthermore, feather peckers were more likely to peck while waiting for a reward after ATD, suggesting a role for the serotonergic system on cognition in these birds.

Abstract 2187: Tryptophan Metabolism Plays a Central Role in Immunosuppression

Abstract Tryptophan metabolism plays a central role in immunosuppression through the local depletion of tryptophan with the concomitant production and accumulation of kynurenine, both of which are immunosuppressive. Indoleamine 2,3-dioxygenase 1 (IDO1), a heme-dependent enzyme, catalyzes the initial and rate-limiting step of the kynurenine pathway. Tumor cells selectively upregulate IDO1 as an immune-evasion mechanism either through intrinsic expression of IDO1, or in response to IFN-γ, a cytokine secreted by immune cells during an active immune response. In addition to the immuno-suppressive role of IDO1 and kynurenine, we investigated their additional roles in tumor cells. Here we describe an NMR-based readout assay for IDO1 activity, in which we are able to trace the extra- and intracellular destination of the single tryptophan carbon groups. This assay allowed us to detect contribution of the tryptophan catabolism to purine synthesis in tumor cells, and suggested further roles of tryptophan catabolism in tumors, apart from kynurenine production, that contribute to the modulation of tumor growth. We detected the incorporation of labeled tryptophan carbon units in intracellular purine pools. Our highly potent, orally available, and CNS penetrant small molecule inhibitor of IDO1 (LY3381916) inhibits this incorporation of tryptophan-derived one carbon units into purines in vitro and in vivo. This activity leads to pre-clinical efficacy and may be an added advantage of inhibiting IDO1 in cancer. Citation Format: Sandaruwan Geeganage, Lillian Sams, James Henry, Frank Dorsey, Kenneth Roth, Alexander Nikolayev, Karim Benhadji, Raymond Gilmour, Ana Cerezo, Sandra Peregrina, Gloria Martínez-del Hoyo, Ramón Campos-Olivas, Juan Manuel Funes, Laura Diezma, Susana Velasco-Miguel. Tryptophan Metabolism Plays a Central Role in Immunosuppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2187.

Dissociable Effects of Tryptophan Supplementation on Negative Feedback Sensitivity and Reversal Learning.

Serotonin has been shown to modulate probabilistic reversal learning (PRL) and negative feedback sensitivity (NFS) in both animal and human studies. Whilst these two measures are tightly coupled, some studies have suggested that these may be mediated by independent mechanisms; the former, representing perseveration and cognitive flexibility, and the latter measuring the ability to maintain a response set (win-stay) at the expense of lose-shift behavior when occasional misleading feedback has been presented. Here, we tested this hypothesis in 44 healthy participants who were administered tryptophan (22 placebo, 22 tryptophan), a precursor to serotonin. We found a dissociable effect of tryptophan supplementation on PRL/NFS. Specifically, tryptophan administration increased NFS compared to the placebo group but had no effect on PRL. We discuss these findings in relation to dosages and with a particular focus on the acute tryptophan depletion (ATD) procedures.

Role of Tryptophan in Microbiota-Induced Depressive-Like Behavior: Evidence From Tryptophan Depletion Study.

During the past decade, there has been a substantial rise in the knowledge about the effects of gut microbiota on host physiology and behavior, including depressive behavior. Initial studies determined that gut microbiota can regulate host tryptophan levels, which is a main serotonin precursor. A dysfunctional serotonergic system is considered to be one of the main factors contributing to the development of depression. Therefore, we hypothesized that regulation of brain tryptophan and serotonin can explain, at least partly, the effects of microbiota on depressive behavior. To test this hypothesis, we examined depressive-like behavior and brain levels of serotonin and tryptophan, of germ free (GF) and specific-pathogen free (SPF) mice under basal conditions, or after acute tryptophan depletion (ATD) procedure, which is a method to decrease tryptophan and serotonin levels in the brain. In basal conditions, GF mice exhibited less depressive-like behavior in sucrose preference, tail-suspension and forced swim tests, compared to SPF mice. In addition, in mice that were not subjected to ATD, GF mice displayed higher levels of tryptophan, serotonin and 5-hydroxyindoleacetic acid (the main degradation product of serotonin) in medial prefrontal cortex (mPFC) and hippocampus (HIPPO), compared to SPF mice. Interestingly, ATD increased depressive-like behavior of GF, but not of SPF mice. These behavioral changes were accompanied by a stronger reduction of tryptophan, serotonin and 5-hydroxyindoleacetic acid in mPFC and HIPPO in GF mice after ATD, when compared to SPF mice. Therefore, the serotonergic system of GF mice is more vulnerable to the acute challenge of tryptophan reduction, and GF mice after tryptophan reduction behave more similarly to SPF mice. These data provide functional evidence that microbiota affects depression-like behavior through influencing brain tryptophan accessibility and the serotonergic system.

Developing a flow cytometry-based quantitative ido assay to measure immune potency of mesenchymal stromal cells product for phase i clinical trial

Background & Aim Establishment of a potency assay in the manufacturing of clinical grade mesenchymal stromal cells (MSCs) has been a challenge due to the issues of variability in starting cellular material, complex mechanisms of action, and lack of reference cells or materials. Indoleamine 2, 3-dioxygenase (IDO) is one of the most important factors for human MSC-mediated immunosuppression. IDO is expressed in IFN-γ-licensed MSC, but not in resting MSC in vitro. This intracellular enzyme catalyzes tryptophan degradation toward kynurenine and depletion of tryptophan suppresses T cells proliferation. Methods, Results & Conclusion In this study, we have developed a flow cytometry-based IDO assay for quantitatively measuring the IDO expression level per cell, the IPBV (Intracellular Protein Binding Value) of the cells represents the relative quantity of IDO expressed in the MSC. IPBV is defined as the number of anti-IDO fluorescent antibody molecules bound to intracellular IDO and standardized by fluorescence intensity of beads. Two kinds of in-house quality control reference cells were running in parallel with product to ensure that the assay is performing as expected and the level of IDO expression. Validation of the quantitative IDO assay, including accuracy, precision, specificity, linearity, range and robustness, were presented in this study. Moreover, we presented the application of the quantitative IDO assay for analysis of MSC product stability, comparability, and compatibility in the manufacturing of clinical grade MSC-based product- RegStem® ASC. In this study, we demonstrated the flow cytometry-based quantitative MSC IDO assay correlates with MSC's inhibition of autologous T cells proliferation, as well as the correlation with preliminary clinical effects in the phase I autologous MSC-based clinical trial for the treatment of Osteoarthritis. As the results, we believe that our novel flow cytometry-based quantitative IDO assay can be a potential potency assay candidate for advanced phase clinical trial of MSCs products.

Indoleamine 2,3-dioxygenase during acute toxoplasmosis and the hyporesponsiveness of splenocytes

Abstract Toxoplasma gondii (T. gondii) is an obligate intracellular parasite of medical and veterinary importance. Major mediators of defense to a T. gondii infection are interferon-gamma induced effector mechanisms, e.g. inducible nitric oxide synthase, immunity related GTPases and indoleamine 2,3-dioxygenase (IDO). Suppression of T cell responses – that diminish after several weeks post infection – is observed during acute toxoplasmosis as well as in other infectious diseases. IDO catalyzes the rate limiting step of the kynurenine pathway by utilizing tryptophan. Furthermore, IDO activity has been described to possess antimicrobial and immunoregulatory properties, but its role during acute toxoplasmosis and the altered T cell responsiveness is obscure. To investigate this part, WT (C57BL/6) and IDO-deficient mice were infected intraperitoneally with T. gondii ME49 tachyzoites. Parasite burden, IDO expression as well as tryptophan and kynurenine concentrations were analyzed. During murine T. gondii infection tryptophan depletion and kynurenine detection was clearly measureable. Since the role of IDO in the T cell hyporesponsiveness during acute toxoplasmosis is incompletely understood, primed and unprimed splenocytes were isolated from infected and naïve mice, respectively. Proliferative responses of the splenocytes to ex vivo mitogen stimulation were measured by detection of incorporated 3H-thymidine. Cell sorting and subsequent cell mixing experiments as well as supernatant transfer experiments were used to break down the mode of action. The observed hyporesponsiveness was independent of IDO. We suggest that the hyporesponsiveness is influenced by nitric oxide synthase and regulatory T cells.

Co-treatment with interferon-\u03b3 and 1-methyl tryptophan ameliorates cardiac fibrosis through cardiac myofibroblasts apoptosis

Cardiac remodeling characterized by cardiac fibrosis is a pathologic process occurring after acute myocardial infarction. Fibrosis can be ameliorated by interferon-gamma (IFN-γ), which is a soluble cytokine showing various effects such as anti-fibrosis, apoptosis, anti-proliferation, immunomodulation, and anti-viral activities. However, the role of IFN-γ in cardiac myofibroblasts is not well established. Therefore, we investigated the anti-fibrotic effects of IFN-γ in human cardiac myofibroblasts (hCMs) in vitro and whether indoleamine 2,3-dioxygenase (IDO), induced by IFN-γ and resulting in cell cycle arrest, plays an important role in regulating the biological activity of hCMs. After IFN-γ treatment, cell signaling pathways and DNA contents were analyzed to assess the biological activity of IFN-γ in hCMs. In addition, an IDO inhibitor (1-methyl tryptophan; 1-MT) was used to assess whether IDO plays a key role in regulating hCMs. IFN-γ significantly inhibited hCM proliferation, and IFN-γ-induced IDO expression caused cell cycle arrest in G0/G1 through tryptophan depletion. Moreover, IFN-γ treatment gradually suppressed the expression of α-smooth muscle actin. When IDO activity was inhibited by 1-MT, marked apoptosis was observed in hCMs through the induction of interferon regulatory factor, Fas, and Fas ligand. Our results suggest that IFN-γ plays key roles in anti-proliferative and anti-fibrotic activities in hCMs and further induces apoptosis via IDO inhibition. In conclusion, co-treatment with IFN-γ and 1-MT can ameliorate fibrosis in cardiac myofibroblasts through apoptosis.

Measuring endogenous changes in serotonergic neurotransmission with [11C]Cimbi-36 positron emission tomography in humans

Developing positron emission tomography (PET) radioligands for the detection of endogenous serotonin release will enable the investigation of serotonergic deficits in many neuropsychiatric disorders. The present study investigates how acute challenges that aim to increase or decrease cerebral serotonin levels affect binding of the serotonin 2A receptor (5-HT2AR) agonist radioligand [11C]Cimbi-36. In a randomized, double-blind, placebo-controlled, three-arm design, 23 healthy volunteers were PET scanned twice with [11C]Cimbi-36: at baseline and following double-blind assignment to one of three interventions (1) infusion of the selective serotonin reuptake inhibitor (SSRI) citalopram preceded by oral dosing of the 5-HT1AR antagonist pindolol, (n = 8) (2) acute tryptophan depletion (ATD) (n = 7) and (3) placebo (n = 8). Two-sample t-tests revealed no significant group differences in percent change of neocortical [11C]Cimbi-36 binding from baseline to intervention between placebo and citalopram/pindolol (p = 0.4) or between placebo and ATD (p = 0.5). Notably, there was a significantly larger within-group variation in 5-HT2AR binding after intervention with citalopram/pindolol, as compared with placebo (p = 0.007). These findings suggest that neither ATD nor a combination of citalopram and pindolol elicit acute unidirectional changes in serotonin levels sufficient to be detected with [11C]Cimbi-36 PET in neocortex. We suggest that the large interindividual variation in 5-HT2AR binding after citalopram/pindolol reflects that after an acute SSRI intervention, individuals respond substantially different in terms of their brain serotonin levels. Our observation has a potential impact for the understanding of patient responses to SSRI.

The association between plasma cholesterol and the effect of tryptophan depletion on heart rate variability.

Studies on the cholesterol-serotonin hypothesis and its link to mood disorders are scarce. In addition, little is known about the association between cholesterol and the effects of tryptophan depletion (TD). The aim of the present study was to investigate the association between plasma cholesterol and changes in heart rate variability (HRV), an important marker of depression and anxiety, after TD. The plasma cholesterol levels of 28 healthy participants were noted, and their HRVs were measured by spectrum analysis. TD was carried out on testing day, and participants provided blood samples just before and 5 hours for tryptophan level after TD. HRV was measured again after TD. An association was found between plasma cholesterol levels and the change in HRV. Decreased high frequency HRV was marginally associated with lower levels of high-density lipoprotein cholesterol, while increased low frequency HRV was significantly associated with lower levels of total and low-density lipoprotein cholesterol. Our findings indicate that low cholesterol levels may play parts of role in the mechanism of the deactivation of parasympathetic, and activation of sympathetic, functions induced by altered serotonergic function.

Exploring common changes after acute mental stress and acute tryptophan depletion: Resting-state fMRI studies

Stress and low serotonin levels are important biological factors in depression and anxiety etiologies. Although studies indicate that low serotonin levels, stress, and other factors may interact in depression/anxiety psychopathology, few studies have investigated the potentially shared neural substrates. We conducted resting-state fMRI scans pre- and post-stress task, and under control and tryptophan depletion condition, to explore the common changes induced by acute mental stress (AMS) and acute tryptophan depletion (ATD). The present study targeted regions within core brain networks – default mode network, salience network, executive control network, and emotion network – reported altered in AMS and ATD, and used regional homogeneity (ReHo) and functional connectivity (FC) analyses to explore their overlapped effects. We additionally examined the relationships among core neural networks – operationalized as an index of resource allocation bias that quantifies the shift from internal to external modes of processing. We found both manipulations induced increased ReHo of the amygdala and decreased ReHo of the posterior cingulate cortex (PCC). The PCC-amygdala FC was negatively correlated with the change of negative affect, whereas the right dorsolateral prefrontal cortex and right anterior insula FC was positively associated with anxiety level. In addition, we found that a greater shift to an external mode was correlated with higher anxiety level under both conditions. Common changes induced by acute mental stress and acute tryptophan depletion confirmed our hypothesis that AMS and ATD induce changes in common neural pathways, which in turn might mark vulnerability to depression and anxiety.

Understandings of Immunomodulatory Pathways of Indoleamine 2,3‐dioxygenase (IDO) in Bladder Cancer Cells

Bladder cancer (BC) is the fourth most common cancer between men and the ninth in women, with a high recurrence rate and progression after conventional treatments. A growing body of evidence suggests that immunomodulatory molecules protect bladder tumors against immune surveillance by triggering signaling pathways that modulate locally the immune system. One important molecule is indoleamine 2,3‐dioxygenase (IDO), which promotes tryptophan breakdown, activating sensitive pathways such as GCN2 in immune cells. In addition, tryptophan breakdown leads to the production of kynurenine compounds, which activate aryl hydrocarbon receptor (AHR), promoting anergy and apoptosis on T cells and Natural killer cells. Although these IDO‐mediated pathways are well recognized in immune cells, they were not yet analyzed in bladder cancer cells. The use of IDO inhibitors arises as a promise anti‐tumor strategy, however, they have divergent effects depending on the activated pathway. Here, we analyzed the expression of IDO and the activation of its pathways in non‐invasive muscle and in invasive muscle bladder cancer cells, and the effect of two IDO inhibitors on these pathways.RT4 and T24 cells were maintained in RPMI medium and incubated for 48h with the following stimuli: IFN‐γ (1000U/mL), Methyl‐D‐tryptophan (MT) (500μM), Incb024360 analog (INCB) (1μM). Control cells were maintained in RPMI medium. After the stimuli, RNA was extracted and qRT‐PCR for IDO, CHOP (a marker of GCN2 activation), CYP1A1 (an AHR activation marker), and AHR was performed.T24 cells constitutively express higher levels of IDO when compared to RT4 cells (relative expression (RE) of 1.0±0.1 in RT4 vs. 73.8±17.0 in T24), accompanied by high expression of CHOP (RE of 1.0±1.2 in RT4 vs. 10.5±4.5 in T24) and CYP1A1 (RE 1.0±0.0 in RT4 vs. 28.2±1.5 in T24), with no differences in AHr expression. When stimulated with IFN‐γ, both RT4 and T24 cells showed increased IDO expression, CHOP and CYP1A1 (p&lt;0.001). While INCB significantly reduced the IDO‐mediated pathways (CHOP and CYP1A1 expression), MT treatment increased CYP1A1 expression (p&lt;0.05).IDO‐regulated pathways are differentiated in invasive and noninvasive bladder cancer cells. As such, adjuvant treatments to modulate IDO are indeed promising. The use of certain inhibitors, however, such as MT, while may inhibit tryptophan depletion, also lead to activation of AHR, promoting kynurenine‐like effects. The use of non AHR binding compounds, such as Incb024360 analogs are promising then proposed as an alternative and safer approach for regulating IDO‐mediated events without nonspecific bindings.Support or Funding InformationFundação de Amparo a Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Expression of Indoleamine 2,3-dioxygenase in erlotinib-treated patients with advanced pancreatic cancer: Translational results from a multi-center, randomized phase III trial

Background: Indoleamine 2,3-dioxygenase 1 (IDO1) is a key enzyme that catalyzes the first and rate-limiting step in the conversion of tryptophan to kynurenine. Depletion of tryptophan and accumulation of kynurenine and its downstream metabolites inhibit effector T cells and favor a regulatory T cell phenotype. Preliminary phase I/II results suggest, that IDO1 might be a promising target in pancreatic ductal adenocarcinoma (PDAC).