The association between HIV-1 Tat and Vif amino acid sequence variation, inflammation and Trp-Kyn metabolism: an exploratory investigation.

BACKGROUND: HIV-1 has well-established mechanisms to disrupt essential pathways in people with HIV, such as inflammation and metabolism. Moreover, diversity of the amino acid sequences in fundamental HIV-1 proteins including Tat and Vif, have been linked to dysregulating these pathways, and subsequently influencing clinical outcomes in people with HIV. However, the relationship between Tat and Vif amino acid sequence variation and specific immune markers and metabolites of the tryptophan-kynurenine (Trp-Kyn) pathway remains unclear. Therefore, this study aimed to investigate the relationship between Tat/Vif amino acid sequence diversity and Trp-Kyn metabolites (quinolinic acid (QUIN), Trp, kynurenic acid (KA), Kyn and Trp/Kyn ratio), as well as specific immune markers (sCD163, suPAR, IL-6, NGAL and hsCRP) in n = 67 South African cART-naïve people with HIV. METHODS: Sanger sequencing was used to determine blood-derived Tat/Vif amino acid sequence diversity. To measure Trp-Kyn metabolites, a LC-MS/MS metabolomics platform was employed using a targeted approach. To measure immune markers, Enzyme-linked immunosorbent assays and the Particle-enhanced turbidimetric assay was used. RESULTS: After adjusting for covariates, sCD163 (p = 0.042) and KA (p = 0.031) were higher in participants with Tat signatures N24 and R57, respectively, and amino acid variation at position 24 (adj R2 = 0.048, ß = -0.416, p = 0.042) and 57 (adj R2 = 0.166, ß = 0.535, p = 0.031) of Tat were associated with sCD163 and KA, respectively. CONCLUSIONS: These preliminary findings suggest that amino acid variation in Tat may have an influence on underlying pathogenic HIV-1 mechanisms and therefore, this line of work merits further investigation.

The correlation of inflammation, tryptophan-kynurenine pathway, and suicide risk in adolescent depression.

Accumulating evidence suggests a role for the tryptophan-kynurenine pathway (TKP) in the psychopathology of major depressive disorder (MDD). Abnormal inflammatory profile and production of TKP neurotoxic metabolites appear more pronounced in MDD with suicidality. Progress in understanding the neurobiology of MDD in adolescents lags significantly behind that in adults due to limited empirical evidence. Aims of this study was to investigate the association between inflammation, TKP, and suicidality in adolescent depression. Seventy-three adolescents with MDD were assessed for serum levels of interleukin (IL)-1ß, IL-6, IL-18, IL-10, tumor necrosis factor-α (TNF-α), tryptophan (TRP), kynurenine (KYN), 3-hydroxykynurenine (3-HK), and kynurenine acid (KA). Correlations between cytokines and TKP measures were examined. Patients were divided into high- (n = 42) and non-high-suicide-risk groups (n = 31), and serum levels of cytokines and TKP metabolites were compared. Significant negative correlations were found between TRP and IL-8 (r = - 0.27, P < 0.05) and IL-10 (r = - 0.23, P < 0.05), while a significant positive correlation was observed between 3-HK and IL-8 (r = 0.39, P < 0.01) in depressed adolescents. The KYN/TPR (index of indoleamine 2,3-dioxygenase, IDO) was positively correlated with IL-1ß (r = 0.34), IL-6 (r = 0.32), IL-10 (r = 0.38) and TNF-α (r = 0.35) levels (P < 0.01); and 3-HK/KYN (index of kynurenine3-monooxidase, KMO) was positively correlated with IL-8 level (r = 0.31, P < 0.01). Depressed adolescents at high suicide risk exhibited significantly higher levels of IL-1ß (Z = 2.726, P < 0.05), IL-10 (Z = 2.444, P < 0.05), and TNF-α (Z = 2.167, P < 0.05) and lower levels of 3-HK (Z = 2.126, P < 0.05) compared to their non-high suicide risk counterparts. Our findings indicated that serum inflammatory cytokines were robustly associated with IDO and KMO activity, along with significantly decreased serum level of TRP, increased level of 3-HK, and higher suicide risk in adolescent depression.

Long-term Treatment with a 5-Alpha-Reductase Inhibitor Alleviates Depression-like Behavior in Obese Male Rats.

Several studies have reported side effects of finasteride (FIN), such as anxiety/depression in young men. Obesity is also positively associated with anxiety/depression symptoms; however, the impacts of long-term FIN treatment and FIN withdrawal in young obese individuals are still elusive. The present study aimed to investigate the effect of long-term treatment and its withdrawal on anxiety/depression and brain pathologies in lean and obese adult male rats. Forty-eight male Wistar rats were equally divided into two groups and fed either a normal or high-fat diet. At age 13 weeks, rats in each dietary group were divided into three subgroups: 1) the control group receiving drinking water, 2) the long-term treatment group receiving FIN orally at 5 mg/kg/day for 6 weeks, and 3) the withdrawal group receiving FIN orally at 5 mg/kg/day for 2 weeks followed by a 4-week withdrawal period. Anxiety/depression-like behaviors, biochemical analysis, brain inflammation, oxidative stress, neuroactive steroids, brain metabolites, and microglial complexity were tested. The result showed that lean rats treated with long-term FIN and its withdrawal exhibited metabolic disturbances, depressive-like behavior, and both groups showed increased neurotoxic metabolites and reduced microglial complexity. Obesity itself led to metabolic disturbances and brain pathologies, including increased inflammation, oxidative stress, and quinolinic acid, as well as reduced microglial complexity, resulting in increased anxiety- and depression-like behaviors. Interestingly, the long-term FIN treatment group in obese rats showed attenuation of depressive-like behaviors, brain inflammation, and oxidative stress, along with increased brain antioxidants, suggesting the possible benefits of FIN in obese conditions.

A pilot investigation of the association between HIV-1 Vpr amino acid sequence diversity and the tryptophan-kynurenine pathway as a potential mechanism for neurocognitive impairment.

HIV infection compromises both the peripheral and central immune systems due to its pathogenic and neuropathogenic features. The mechanisms driving HIV-1 pathogenesis and neuropathogenesis involve a series of events, including metabolic dysregulation. Furthermore, HIV-subtype-specific variations, particularly alterations in the amino acid sequences of key viral proteins, are known to influence the severity of clinical outcomes in people living with HIV. However, the impact of amino acid sequence variations in specific viral proteins, such as Viral protein R (Vpr), on metabolites within the Tryptophan (Trp)-kynurenine (Kyn) pathway in people living with HIV remains unclear. Our research aimed to explore the relationship between variations in the Vpr amino acid sequence (specifically at positions 22, 41, 45, and 55, as these have been previously linked to neurocognitive function) and peripheral Trp-Kyn metabolites. Additionally, we sought to clarify the systems biology of Vpr sequence variation by examining the link between Trp-Kyn metabolism and peripheral inflammation, as a neuropathogenic mechanism. In this preliminary study, we analyzed a unique cohort of thirty-two (n = 32) South African cART naïve people living with HIV. We employed Sanger sequencing to ascertain blood-derived Vpr amino acid sequence variations and a targeted LC-MS/MS metabolomics platform to assess Trp-Kyn metabolites, such as Trp, Kyn, kynurenic acid (KA), and quinolinic acid (QUIN). Particle-enhanced turbidimetric assay and Enzyme-linked immunosorbent assays were used to measure immune markers, hsCRP, IL-6, suPAR, NGAL and sCD163. After applying Bonferroni corrections (p =.05/3) and adjusting for covariates (age and sex), only the Vpr G41 and A55 groups was nearing significance for higher levels of QUIN compared to the Vpr S41 and T55 groups, respectively (all p =.023). Multiple regression results revealed that Vpr amino acid variations at position 41 (adj R2 = 0.049, ß = 0.505; p =.023), and 55 (adj R2 = 0.126, ß = 0.444; p =.023) displayed significant associations with QUIN after adjusting for age and sex. Lastly, the higher QUIN levels observed in the Vpr G41 group were found to be correlated with suPAR (r =.588, p =.005). These results collectively underscore the importance of specific Vpr amino acid substitutions in influencing QUIN and inflammation (specifically suPAR levels), potentially contributing to our understanding of their roles in the pathogenesis and neuropathogenesis of HIV-1.

Frontostriatal circuitry and the tryptophan kynurenine pathway in major psychiatric disorders.

RATIONALE: An imbalance of the tryptophan kynurenine pathway (KP) commonly occurs in psychiatric disorders, though the neurocognitive and network-level effects of this aberration are unclear. OBJECTIVES: In this study, we examined the connection between dysfunction in the frontostriatal brain circuits, imbalances in the tryptophan kynurenine pathway (KP), and neurocognition in major psychiatric disorders. METHODS: Forty first-episode medication-naive patients with schizophrenia (SCZ), fifty patients with bipolar disorder (BD), fifty patients with major depressive disorder (MDD), and forty-two healthy controls underwent resting-state functional magnetic resonance imaging. Plasma levels of KP metabolites were measured, and neurocognitive function was evaluated. Frontostriatal connectivity and KP metabolites were compared between groups while controlling for demographic and clinical characteristics. Canonical correlation analyses were conducted to explore multidimensional relationships between frontostriatal circuits-KP and KP-cognitive features. RESULTS: Patient groups shared hypoconnectivity between bilateral ventrolateral prefrontal cortex (vlPFC) and left insula, with disorder-specific dysconnectivity in SCZ related to PFC, left dorsal striatum hypoconnectivity. The BD group had higher anthranilic acid and lower xanthurenic acid levels than the other groups. KP metabolites and ratios related to disrupted frontostriatal dysconnectivity in a transdiagnostic manner. The SCZ group and MDD group separately had high-dimensional associations between KP metabolites and cognitive measures. CONCLUSIONS: The findings suggest that KP may influence cognitive performance across psychiatric conditions via frontostriatal dysfunction.

Indoleamine 2,3-dioxygenase 2 deficiency associates with autism-like behavior via dopaminergic neuronal dysfunction.

Indoleamine 2,3-dioxygenase 2 (IDO2) is an enzyme of the tryptophan-kynurenine pathway that is constitutively expressed in the brain. To provide insight into the physiological role of IDO2 in the brain, behavioral and neurochemical analyses in IDO2 knockout (KO) mice were performed. IDO2 KO mice showed stereotyped behavior, restricted interest and social deficits, traits that are associated with behavioral endophenotypes of autism spectrum disorder (ASD). IDO2 was colocalized immunohistochemically with tyrosine-hydroxylase-positive cells in dopaminergic neurons. In the striatum and amygdala of IDO2 KO mice, decreased dopamine turnover was associated with increased α-synuclein level. Correspondingly, levels of downstream dopamine D1 receptor signaling molecules such as brain-derived neurotrophic factor and c-Fos positive proteins were decreased. Furthermore, decreased abundance of ramified-type microglia resulted in increased dendritic spine density in the striatum of IDO2 KO mice. Both chemogenetic activation of dopaminergic neurons and treatment with methylphenidate, a dopamine reuptake inhibitor, ameliorated the ASD-like behavior of IDO2 KO mice. Sequencing analysis of exon regions in IDO2 from 309 ASD samples identified a rare canonical splice site variant in one ASD case. These results suggest that the IDO2 gene is, at least in part, a factor closely related to the development of psychiatric disorders.

Investigation of the effects of oxidative stress, inflammation on the pathway of tryptophan/kynurenine in OCD.

OBJECTIVES: Recent studies have shown that the distribution of the tryptophan/kynurenine pathway (KP) plays a role in the development of obsessive-compulsive disorder (OCD). We aimed to reveal the relationship between CYP1A1 rs464903 and aryl hydrocarbon receptor (AhR) rs10249788 associated with the KP and interferon gamma (IFN γ) and oxidative stress in OCD. METHODS: In our study, the serum and DNAs of 150 samples, including 100 OCD patients and 50 controls, were used. The activity of glutathione peroxidase (GSH-Px), and the levels of IFN γ, thiobarbituric acid reactive substances (TBARS), tryptophan, and kynurenine were determined by biochemical methods. AhR rs10249788 and cytochrome P450 family CYP1A1 rs4646903, which interact directly with the KP, were analysed by polymerase chain reaction followed by restriction fragment length polymorphism. P < 0.05 was considered statistically significant. RESULT: There were no significant differences between groups in CYP1A1 rs4646903 and AhR rs10249788 while tryptophan and IFN γ were found to be higher in controls (p < 0.001, for both), and TBARS and indolamine-2,3-dioxygenase were found to be higher in OCD (p < 0.001, for both). There were significant correlations between IFN γ and TBARS and GSH-Px (p = 0.028, p = 0.020, respectively) in the OCD group. CONCLUSIONS: For the first time studied in OCD, it has been shown that IFN γ, tryptophan, oxidative stress parameters, and gene variants of CYP1A1 rs4646903 anAhR rs10249788 are shown effective on the KP.

Identification and characterization of tryptophan-kynurenine pathway-related genes involving lamprey (Lampetra japonica) innate immunity.

The tryptophan-kynurenine (TRP-KYN) pathway is involved in several biological functions, including immunosuppression, inflammatory response, and tumor suppression. Six TRP-KYN pathway-related genes, tryptophan 2,3-dioxygenase (TDO), indoleamine 2,3-dioxygenase 2 (IDO2), aminoadipate aminotransferase (AADAT), glutamate oxaloacetate transaminase 2 (GOT2), kynurenine monooxygenase (KMO), and kynureninase (KYNU) have been identified and cloned from the jawless vertebrate lamprey (Lampetra japonica) to gain insights into their evolution and characterization. Expression distribution showed that the key gene Lj-TDO was highly expressed in the oral gland. Real-time quantitative PCR showed that TRP-KYN pathway-related genes were significantly overexpressed after multi-stimulation. RNA interference showed that Lj-IDO2 knockdown regulated the expression of inflammatory factors. In conclusion, our study successfully clarified the ancestral features and functions of the TRP-KYN pathway, while providing valuable insights into the involvement of this pathway in the immune responses of a jawless vertebrate.

Adiposity and plasma concentrations of kynurenine pathway metabolites and traditional markers of inflammation.

AIM: The kynurenine pathway is increasingly recognised to play a role in inflammation and disease. We assessed the cross-sectional and longitudinal associations of adiposity measures (body mass index, waist-hip ratio, waist circumference and fat mass ratio) with plasma concentrations of kynurenine pathway metabolites and traditional markers of inflammation. METHODS: We used data from 970 Melbourne Collaborative Cohort Study participants who had plasma markers measured at baseline (median age 59 years) and follow-up (median age 70 years). Linear regression was used to assess cross-sectional and longitudinal associations between four adiposity measures and concentrations of i) nine kynurenine pathway metabolites; ii) two derived markers; iii) eight traditional inflammatory markers. RESULTS: Cross-sectionally, most kynurenine metabolites were strongly associated with adiposity measures at both time points; associations were generally stronger than for most inflammation markers except CRP (e.g. body mass index at baseline, quinolinic acid (per S.D. ß = 0.30, 95%CI: 0.24-0.36, P = 10-21), kynurenine (ß = 0.25, 95%CI: 0.19-0.31, P = 10-16) and CRP (ß = 0.31, 95%CI: 0.25-0.37, P = 10-24), and remained largely unchanged after adjustment for confounders. Longitudinally, changes in adiposity measures over approximately a decade were positively associated with changes in kynurenine metabolite concentrations (in particular for 3-hydroxyanthranilic acid, kynurenine and quinolinic acid), and more strongly so than for other markers of inflammation, including CRP. CONCLUSIONS: In middle-aged and older adults, plasma concentrations of kynurenine metabolites are strongly associated with adiposity, both cross-sectionally and longitudinally. Our study demonstrates that kynurenine metabolites may be valuable markers to monitor the adverse consequences of obesity.

Minocycline, a classic antibiotic, exerts psychotropic effects by normalizing microglial neuroinflammation-evoked tryptophan-kynurenine pathway dysregulation in chronically stressed male mice.

The dysregulation of tryptophan-kynurenine pathway (TKP) is extensively involved in the pathophysiology of Alzheimer's disease, depression, and neurodegenerative disorders. Minocycline, a classic antibiotic, may exert psychotropic effects associated with the modulation of TKP. In this study, we examined the effects of minocycline in improving behaviour and modulating TKP components in chronically stressed male mice. Following repeated treatment with 22.5 mg/kg and 45 mg/kg minocycline for 27 days, the stressed mice particularly with higher dose displayed significant improvement on cognitive impairment, depression- and anxiety-like behaviour. Minocycline suppressed stress-induced overexpression of pro-inflammatory cytokines and restored anti-inflammatory cytokines. Chronic stress dramatically suppressed blood and prefrontal cortical levels of the primary substrate tryptophan (TRP), the neuroprotective metabolite kynurenic acid (KYNA), and KYNA/KYN ratio, but increased the intermediate kynurenine (KYN), 3-hydroxykynurenine (3-HK), KYN/TRP ratio, and the neurotoxic metabolite quinolinic acid (QUIN). Minocycline partially or completely reversed changes in these components. Minocycline also inhibited stress-induced overexpression of QUIN-related enzymes, indoleamine 2, 3-dioxygenase 1(iDO-1), kynureninase (KYNU), kynurenine 3-monooxygenase (KMO), 3-hydroxyanthranilate 3,4-dioxygenase (3-HAO), but rescued the decreased expression of kynurenine aminotransferase (KAT) in brain regions. Behavioral improvements were correlated with multiple TKP metabolites and enzymes. These results suggest that the psychotropic effects of minocycline are mainly associated with the restoration of biodistribution of the primary substrate in the brain and normalization of neuroinflammation-evoked TKP dysregulation.

The prevalence and the effect of interferon -γ in the comorbidity of rheumatoid arthritis and depression.

BACKGROUND: Depression is the most common comorbidities associated with rheumatoid arthritis (RA). We aimed to explore the mechanism of association between RA and depression. METHODS: 120 subjects were enrolled and depression was diagnosed and assessed using DSM-5 and 24-item version of Hamilton Depression Scale. Pain intensity and joint function in patients with RA were assessed using the visual analog scale (VAS) and health assessment questionnaire (HAQ). Serum levels of interferon-gamma (IFN-γ), indoleamine 2,3-dioxygenase (IDO), kynurenine (KYN), tryptophan (TRP), and quinolinic acid (QUIN)were detected. In animal experiments, K/BxN mice with RA-like phenotype was used and depressive behavior was observed. The protein expression level of N-methyl -D- aspartate receptor 2B (NR2B) in the hippocampus was detected. RESULTS: In this study, 36.67 % of patients with RA also had depression. The working status, month family income, tender joint count, the VAS and HAQ score were the main factors influencing the depression in RA patients. HAQ score was found to be an independent risk factor for depression in RA. Serum IDO, IFN-γ, KYN were increased and TRP contents were decreased in RA group. K/BxN mice with RA-like phenotype showed depressive behavior. However, injection of IFN-γ neutralizing antibody could inhibit kynurenine pathway and reverse the depressive behavior in mice. The levels of QUIN in the neurotoxic metabolic pathway were increased and N-methyl -D- aspartate receptors (NMDAR) were activated, which may be the mechanism behind the onset of depression. CONCLUSIONS: From clinical and preclinical aspects, the occurrence of depression in RA was explored and the related mechanism was revealed.

Tryptophan metabolism can modulate immunologic tolerance in primitive vertebrate lamprey via IDO-kynurenine-AHR pathway.

Tryptophan is mainly degraded through kynurenine pathway (KP) in vertebrates which is closely related to the nerve and depression, while the studies on immunity is still limited. This study aims to explore the functions of tryptophan in the innate immunity of primitive vertebrate lamprey. MTT (3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide) assay showed that tryptophan had no obvious effect on cell viability. Tryptophan was transported into leukocytes and degraded via the KP after tryptophan supplement. Tryptophan treatment (T1x and T2x) failed to alter the total antioxidant capacity regardless of stimulation and exposure time. Real-time quantitative PCR and western blotting results revealed that tryptophan was not only able to reduce the expression of pro-inflammatory factors Lj-TNF-α, Lj-IL1ß and Lj-NF-κB, but also to upregulate the expression of anti-inflammatory factor Lj-TGF-ß independent of stimulation and time. In addition, tryptophan can exert immune tolerance function by inhibiting TLR-MyD88 and promoting (Indoleamine 2, 3-Dioxygenase) IDO-kynurenine-AHR (aryl hydrocarbon receptor) pathways. This study provides a new understanding for tryptophan-kynurenine metabolism and mechanism of immune tolerance function in primitive vertebrate lamprey.

Relationship between the tryptophan-kynurenine pathway and painful physical symptoms in patients with major depressive disorder.

OBJECTIVES: To investigate the relationship between the tryptophan-kynurenine (TRP-KYN) pathway and painful physical symptoms (PPS) in major depressive disorder (MDD). METHODS: Eighty-four patients with MDD (40 patients with PPS and 44 without PPS) and forty-six healthy controls (HC) were recruited. The serum levels of tryptophan (TRP), kynurenine(KYN), kynurenic acid (KA), quinolinic acid (QA), 3-hydroxy-kynurenine (3-HK), serotonin (5-HT) were measured using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Depression, anxiety and pain were assessed using Hamilton Depression Scale (HAMD), Hamilton Anxiety Scale (HAMA) and Short-form McGill pain questionnaire (SFMPQ) respectively. RESULTS: Patients in the MDD group exhibited significantly lower KA and 5-HT levels than HC, whereas MDD patients with PPS showed higher KYN and QA levels, and a higher KYN/TRP ratio than those without. There was a positive correlation between the scores of SFMPQ and QA levels and a negative correlation between the scores of SFMPQ and TRP levels or KA/QA ratios in MDD patients with PPS group. Stepwise multiple regression analysis showed that the KYN/TRP ratios, the KA/QA ratios, and the HAMD scores were significant predictor factors for SFMPQ scores. CONCLUSIONS: These results demonstrated that the TRP-KYN pathway may play a role in the pathophysiology of pain in patients with major depressive disorder, suggesting that further studies of this pathway as a potential biomarker or therapeutic target are required.

Differential Levels of Tryptophan-Kynurenine Pathway Metabolites in the Hippocampus, Anterior Temporal Lobe, and Neocortex in an Animal Model of Temporal Lobe Epilepsy.

Glutamate-receptor-mediated hyperexcitability contributes to seizure generation in temporal lobe epilepsy (TLE). Tryptophan-kynurenine pathway (TKP) metabolites regulate glutamate receptor activity under physiological conditions. This study was designed to investigate alterations in the levels of TKP metabolites and the differential regulation of glutamatergic activity by TKP metabolites in the hippocampus, anterior temporal lobe (ATL), and neocortex samples of a lithium-pilocarpine rat model of TLE. We observed that levels of tryptophan were reduced in the hippocampus and ATL samples but unaltered in the neocortex samples. The levels of kynurenic acid were reduced in the hippocampus samples and unaltered in the ATL and neocortex samples of the TLE rats. The levels of kynurenine were unaltered in all three regions of the TLE rats. The magnitude of reduction in these metabolites in all regions was unaltered in the TLE rats. The frequency and amplitude of spontaneous excitatory postsynaptic currents were enhanced in hippocampus ATL samples but not in the neocortex samples of the TLE rats. The exogenous application of kynurenic acid inhibited glutamatergic activity in the slice preparations of all these regions in both the control and the TLE rats. However, the magnitude of reduction in the frequency of kynurenic acid was higher in the hippocampus (18.44 ± 2.6% in control vs. 30.02 ± 1.5 in TLE rats) and ATL (16.31 ± 0.91% in control vs. 29.82 ± 3.08% in TLE rats) samples of the TLE rats. These findings suggest the differential regulation of glutamatergic activity by TKP metabolites in the hippocampus, ATL, and neocortex of TLE rats.

Decreased cerebrospinal fluid kynurenic acid in epileptic spasms: A biomarker of response to corticosteroids.

BACKGROUND: Epileptic (previously infantile) spasms is the most common epileptic encephalopathy occurring during infancy and is frequently associated with abnormal neurodevelopmental outcomes. Epileptic spasms have a diverse range of known (genetic, structural) and unknown aetiologies. High dose corticosteroid treatment for 4 weeks often induces remission of spasms, although the mechanism of action of corticosteroid is unclear. Animal models of epileptic spasms have shown decreased brain kynurenic acid, which is increased after treatment with the ketogenic diet. We quantified kynurenine pathway metabolites in the cerebrospinal fluid (CSF) of infants with epileptic spasms and explored clinical correlations. METHODS: A panel of nine metabolites in the kynurenine pathway (tryptophan, kynurenine, kynurenic acid, 3-hydroxykynurenine, xanthurenic acid, anthranilic acid, 3-hydroxyanthranilic acid, quinolinic acid, and picolinic acid) were measured using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). CSF collected from paediatric patients less than 3 years of age with epileptic spasms (n=34, 19 males, mean age 0.85, median 0.6, range 0.3-3 yrs) were compared with other epilepsy syndromes (n=26, 9 males, mean age 1.44, median 1.45, range 0.3-3 yrs), other non-inflammatory neurological diseases (OND) (n=29, 18 males, mean age 1.47, median 1.6, range 0.1-2.9 yrs) and inflammatory neurological controls (n=12, 4 males, mean age 1.80, median 1.80, range 0.8-2.5 yrs). FINDINGS: There was a statistically significant decrease of CSF kynurenic acid in patients with epileptic spasms compared to OND (p<0.0001). In addition, the kynurenic acid/kynurenine (KYNA/KYN) ratio was lower in the epileptic spasms subgroup compared to OND (p<0.0001). Epileptic spasms patients who were steroid responders or partial steroid responders had lower KYNA/KYN ratio compared to patients who were refractory to steroids (p<0.005, p<0.05 respectively). INTERPRETATION: This study demonstrates decreased CSF kynurenic acid and KYNA/KYN in epileptic spasms, which may also represent a biomarker for steroid responsiveness. Given the anti-inflammatory and neuroprotective properties of kynurenic acid, further therapeutics able to increase kynurenic acid should be explored. FUNDING: Financial support for the study was granted by Dale NHMRC Investigator grant APP1193648, Petre Foundation, Cerebral Palsy Alliance and Department of Biochemistry at the Children's Hospital at Westmead. Prof Guillemin is funded by NHMRC Investigator grant APP1176660 and Macquarie University.

Can kynurenine pathway be considered as a next-generation therapeutic target for Parkinson's disease? An update information.

By far, no revolutionary breakthrough in the treatment of Parkinson's disease (PD) was found. It is indeed a knotty problem to select a satisfactory strategy for treating some patients with advanced stage PD. Development of novel therapeutic targets against PD has been an urgent task faced by global PD researchers. Targets in the tryptophan-kynurenine pathway (KP) were then considered. Metabolites in the KP are liposoluble. Some neurotoxic metabolites, including 3-hydroxykynurenine and its downstream 3-hydroxyanthranilic acid and quinolinic acid, are mainly produced peripherally. They can easily cross the blood-brain barrier (BBB) and exert their neurotoxic effects in the central neuron system (CNS), which is considered as a potential pathophysiological mechanism of neurodegenerative diseases. Hence, agents against the targets in the KP have two characteristics: (1) being independent from the dopaminergic system and (2) being seldom affected by the BBB. Inspiringly, one agent, namely, the inhibitor of indoleamine 2,3-dioxygenase 1, has been currently reported to present satisfactory efficacy comparable to levodopa, implying that the KP might be a potential novel target for PD. This review collected and summarized the updated information regarding the association of the KP with PD, which is helpful for understanding the clinical value of the KP in the PD scenario.

Development of a translational inflammation panel for the quantification of cerebrospinal fluid Pterin, Tryptophan-Kynurenine and Nitric oxide pathway metabolites.

BACKGROUND: Neuroinflammatory diseases such as encephalitis, meningitis, multiple sclerosis and other neurological diseases with inflammatory components, have demonstrated a need for diagnostic biomarkers to define treatable and reversible neuroinflammation. The development and clinical validation of a targeted translational inflammation panel using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) could provide early diagnosis, rapid treatment and insights into neuroinflammatory mechanisms. METHODS: An inflammation panel of 13 metabolites (neopterin, tryptophan, kynurenine, kynurenic acid, 3-hydroxykynurenine, xanthurenic acid, anthranilic acid, 3-hydroxyanthranilic acid, quinolinic acid, picolinic acid, arginine, citrulline and methylhistamine) was measured based on a simple precipitation and filtration method using minimal CSF volume. The chromatographic separation was achieved using the Acquity UPLC BEH C18 column in combination with a gradient elution within a 12-min time frame. Acute encephalitis (n=10; myelin oligodendrocyte glycoprotein encephalitis n=3, anti-N-methyl-D-aspartate encephalitis n=2, acute disseminated encephalomyelitis n=2, herpes simplex encephalitis n=1, enteroviral encephalitis n=1) and frequency-matched non-inflammatory neurological disease controls (n=10) were examined. FINDINGS: The method exhibited good sensitivity as the limits of quantification ranged between 0.75 and 3.00 ng mL-1, good linearity (r2 > 0.99), acceptable matrix effects (<± 19.4%) and high recoveries (89.8-109.1 %). There were no interferences observed from common endogenous CSF metabolites, no carryover and concordance with well-established clinical methods. The accuracy and precision for all analytes were within tolerances, at <± 15 mean relative error and < 15 % coefficient of variation respectively. All analytes in matrix-matched pooled human CSF calibrators and human CSF extracts were stable for 24 h after extraction and two freeze-thaw cycles. INTERPRETATION: The method was successfully applied to a pilot study investigating acute brain inflammation case-control groups. Statistical discrimination between encephalitis (n=10) and control groups (n=10) was achieved using orthogonal partial least squares discriminant analysis and heatmap cluster analysis. Statistical analysis of the measured metabolites identified significant alterations of seven metabolites in the tryptophan-kynurenine pathway (tryptophan, kynurenine, kynurenic acid, 3-hydroxykynurenine, anthranilic acid, 3-hydroxyanthranilic acid, quinolinic acid), arginine and neopterin in presence of acute neuroinflammation. Furthermore, elevated ratios of CSF kynurenine/tryptophan ratio, quinolinic acid/kynurenic acid and anthranilic acid/3-hydroxyanthranilic acid provided strong discriminative power for neuroinflammatory conditions. Studies of large groups of neurological diseases are required to explore the sensitivity and specificity of the inflammation panel. FUNDING: Financial support for the study was granted by Dale NHMRC Investigator grant APP1193648, Petre Foundation, Cerebral Palsy Alliance and Department of Biochemistry at the Children's Hospital at Westmead.

[Quantitative analysis of tryptophan and its metabolites in urine by ultra performance liquid chromatography-tandem mass spectrometry].

Tryptophan (Trp), also known as α-amino ß-indolepropionic acid, is an essential amino acid, which is involved in various physiological processes. Studies have shown that tumors, infectious diseases, and neurological diseases are accompanied by Trp-related metabolic disorders. Understanding the excretion of Trp and its metabolites in normal individuals is of great significance for treating Trp-related diseases and monitoring the health. A rapid quantitative method was developed based on ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Further, this method was applied to the simultaneous determination of Trp and its metabolites, including kynurenine (Kyn), kynurenic acid (KA), 3-hydroxykynurenine (3-OH-Kyn), 3-hydroxyanthranilic acid (3-OH-AA), xanthurenic acid (XA), 5-hydroxytryptamine (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA). The excretion and amount of target compounds in random urine samples collected from healthy participants were studied using this method. Urine samples were collected from healthy male volunteers (between 20-22 years old) without any diet and exercise restrictions. Urine samples were collected between 11∶00-13∶00 daily for 10 d. Thereafter, the urine samples were diluted, centrifuged, and subjected to pre-column derivatization with dansyl chloride (DNS-Cl). Caffeic acid (CA) was used as the internal control. Later, the derivatives were detected using triple quadrupole mass spectrometry with electron pray ionization (ESI) in positive and multi reaction monitoring (MRM) modes. The samples were separated using a Thermo C18 column (50 mm×3 mm, 2.7 µm) with 0.1% aqueous formic acid aqueous solution and methanol as mobile phases at a flow rate of 0.2 mL/min. The three most abundant ions for each derivative were selected for downstream analysis, and the internal control was used for quantification. The polarity and molecular weight of the compounds were found to be altered effectively after DNS-Cl derivatization treatment. The dansyl group effectively altered the polarities of the derivatives, such that their retention behaviors in the reverse elution system were similar and they were well separated. The interference due to impurities was effectively eliminated using the MRM mode. The results showed significant linear correlation, since the correlation coefficients were greater than 0.9740. The recoveries were between 93.24%-107.65%, and the LODs were 0.005-0.5 ng/mL for the eight compounds. Trp prototype and the seven target metabolites, including 3-OH-Kyn, 3-OH-AA, XA, Kyn, KA, 5-HIAA, and 5-HT generated through Trp-5-HT and Trp-Kyn pathways were detected in the urine samples. These results indicated that Trp was excreted in a prototypic form or after being metabolized. The level of the target compounds in random urine samples of individuals were 0.99-3.72 (3-OH-Kyn), 2.51-21.11 (3-OH-AA), 0.25-1.12 (XA), 0.15-1.53 (Kyn), 0.24-2.58 (KA), 0-0.31 (5-HT), and 2.2-17.94 (5-HIAA) µg/mL. For the same individual, in the state of physical health, the fluctuations of Trp and its metabolites in urine were large. Due to these large fluctuations in the absolute content, the difference between individuals was not significant. The data generated using 70 urine samples revealed that the amount of excreted Trp being metabolized was 124%-268% of prototype, which further indicated that the excretion after metabolism was the major underlying mechanism. Upon comparing the levels of metabolites in the Trp-5-HT and Trp-Kyn pathways, the results indicated that the levels of 3-OH-AA and 3-OH-Kyn generated upon Trp degradation through the Kyn pathway was higher than those of the other products. Trp was degraded via Kyn pathway to produce 3-OH-AA, which was the main metabolite of Trp found to be present in the body. This manuscript detected the levels of Trp and its metabolites, as well as summarized the characteristics of excretion using random urine samples, which could provide valuable information for clinical practice.

Postpartum corticosterone and fluoxetine shift the tryptophan-kynurenine pathway in dams.

Perinatal depression (PND) affects 15% of mothers. Selective serotonin reuptake inhibitors (SSRIs) are currently the first-line of treatment for PND but are not always efficacious. Previously, we found significant reductions in plasma tryptophan concentrations and higher hippocampal proinflammatory cytokine, IL-1ß levels, due to maternal SSRI treatment. Both inflammation and tryptophan-kynurenine metabolic pathway (TKP) are associated with SSRI efficacy in individuals with major depressive disorder (MDD). TKP is divided into neuroprotective and neurotoxic pathways. Higher metabolite concentrations of the neurotoxic pathway are associated with depression onset and implicated in SSRI efficacy. Metabolites in TKP were investigated in a rodent model of de novo postpartum depression (PPD) given treatment with the SSRI, fluoxetine (FLX). Dams were administered corticosterone (CORT) (40 mg/kg, s.c.), and treated with the SSRI, fluoxetine (FLX) (10 mg/kg, s.c.), during the postpartum for 22 days after parturition. Plasma TKP metabolite concentrations were quantified on the last day of treatment. Maternal postpartum CORT increased neurotoxic metabolites and co-enzyme/cofactors in dams (3-hydroxykynurenine, 3-hydroxyanthranilic acid, vitamin B2, flavin adenine dinucleotide). The combination of both CORT and FLX shifted the neuroprotective-to-neurotoxic ratio towards neurotoxicity. Postpartum FLX decreased plasma xanthurenic acid concentrations. Together, our data indicate higher neurotoxic TKP expression due to maternal postpartum CORT treatment, similar to clinical presentation of MDD. Moreover, maternal FLX treatment showed limited efficacy to influence TKP metabolites, which may correspond to its limited efficacy to treat depressive-like endophenotypes in the postpartum. Overall suggesting changes in TKP may be used as a biomarker of de novo PPD and antidepressant efficacy and targeting this pathway may serve as a potential therapeutic target.