Bernhard Widner

Neopterin as a marker for immune system activation.

Increased amounts of neopterin are produced by human monocytes/macrophages upon stimulation with the cytokine interferon-y. Therefore, measurement of neopterin concentrations in body fluids like serum, cerebrospinal fluid or urine provides information about activation of T helper cell 1 derived cellular immune activation. Increased neopterin production is found in infections by viruses including human immunodeficiency virus (HIV), infections by intracellular living bacteria and parasites, autoimmune diseases, malignant tumor diseases and in allograft rejection episodes. But also in neurological and in cardiovascular diseases cellular immune activation indicated by increased neopterin production, is found. Major diagnostic applications of neopterin measurements are, e.g. monitoring of allograft recipients to recognize immunological complications early. Neopterin production provides prognostic information in patients with malignant tumor diseases and in HIV-infected individuals, high levels being associated with poorer survival expectations. Neopterin measurements are also useful to monitor therapy in patients with autoimmune disorders and in individuals with HIV infection. Screening of neopterin concentrations in blood donations allows to detect acute infections in a non-specific way and improves safety of blood transfusions. As high neopterin production is associated with increased production of reactive oxygen species and with low serum concentrations of antioxidants like alpha-tocopherol, neopterin can also be regarded as a marker of reactive oxygen species formed by the activated cellular immune system. Therefore, by neopterin measurements not only the extent of cellular immune activation but also the extent of oxidative stress can be estimated.

Tryptophan degradation and immune activation in Alzheimer's disease.

Summary. Alzheimers disease (AD) is likely associated with systemic immune activation. During immune response, interferon-gamma stimu-lates indoleamine 2,3-dioxygenase (IDO) converting tryptophan to N-formylkynurenine followed by kynurenine in an ensuing step. Thus, IDO activity is estimated by the kynurenine per tryptophan quotient (Kyn/Trp). In 21 patients suffering from AD, in 20 controls of similar age, and in 49 blood donors we measured serum tryptophan and kynurenine concentrations by HPLC. Lower tryptophan concentrations were found in elderly control subjects compared to blood donors (62.1 vs. 73.0 μM, p < 0.005). Tryptophan concentrations tended to be still lower in AD patients (54.4 μM, p = 0.07) compared to elderly controls. Enhanced tryptophan degradation in patients was reflected by significantly increased Kyn/Trp (46.1 vs. 34.1 in elderly controls, p < 0.05). Correlations were found in patients between Kyn/Trp and concentrations of soluble immune markers in serum, i.e., neopterin, interleukin-2 receptor and tumor necrosis factor receptor (all p < 0.001). Increased Kyn/Trp was associated with reduced cognitive performance. Tryptophan degradation due to immune activation may exert impact on the pathogenesis of AD.

Neopterin production, tryptophan degradation, and mental depression—What is the link?

The cytokine interferon-gamma stimulates human monocytes/macrophages to release large amounts of neopterin. Increased neopterin concentrations in body fluids of patients are observed during diseases with activated cellular (=TH1-type) immune response such as allograft rejection, virus infections, autoimmune disorders, or malignant tumors but also in neurodegenerative diseases or during pregnancy. In various cells interferon-gamma induces indoleamine 2,3-dioxygenase (IDO) which degrades tryptophan via the kynurenine pathway. Therefore like increased neopterin formation, enhanced tryptophan degradation is observed in diseases concomitant with cellular immune activation. Disturbed metabolism of tryptophan affects biosynthesis of neurotransmitter 5-hydroxytryptamine (serotonin), and it appears to be associated with an increased susceptibility for depression. In fact, enhanced neopterin concentrations together with increased degradation of tryptophan and low serum levels of tryptophan correlate with neuropsychiatric abnormalities like cognitive decline and depressive symptoms especially in long-lasting and chronic diseases. Activation of IDO could represent an important link between the immunological network and the pathogenesis of depression.

Serum tryptophan decrease correlates with immune activation and impaired quality of life in colorectal cancer

Cancer-related indoleamine (2,3)-dioxygenase up-regulation by interferon-γ might influence quality of life by depleting serum tryptophan. We correlated serum tryptophan levels with immune activation and quality of life in patients with colorectal liver metastases. Venous blood was sampled from patients with primary colorectal cancer and from patients with metachronous colorectal liver metastases who completed quality of life and psychological questionnaires. Serum tryptophan, kynurenine, neopterin, interleukin 2 soluble receptor α (IL-2 sRα), soluble tumour necrosis factor receptor I (sTNF RI), interleukin 6, and C-reactive protein were measured. Liver metastasis volume was estimated by computerised tomography, and survival from blood sampling was noted. Sixty-six patients with colorectal cancer were studied (39 males; median age 66 years) of whom 25 had colorectal liver metastases only (17 males; median age 62 years; median liver metastasis volume 208 ml; median survival 234 days). Reduced serum tryptophan was significantly associated with Rotterdam Symptom Checklist physical symptom (r=−0.51, P=0.01) and Sickness Impact Profile (r=−0.42, P=0.04) scores, and correlated with increased serum neopterin (r=−0.36, P=0.003), IL-2 sRα (r=−0.51, P=0.01) and sTNF RI (r=−0.45, P=0.02) levels. Stepwise regression analyses suggested that serum tryptophan was an independent predictor of Rotterdam Symptom Checklist physical symptom (regression coefficient −20.78, P=0.01) and Sickness Impact Profile (regression coefficient −109.09, P=0.04) scores. The results supported a role for interferon-γ-mediated serum tryptophan decrease in cancer-induced quality of life deterioration.

Modulation of neopterin formation and tryptophan degradation by Th1- and Th2-derived cytokines in human monocytic cells

In order to examine the regulatory effects of major Th1‐derived cytokines, such as IL‐12, and Th2 cytokines, IL‐4 and IL‐10, on the formation of neopterin and degradation of tryptophan, two metabolic pathways induced by interferon‐gamma (IFN‐γ) in human monocytes/macrophages, we investigated the human monocytic cell line THP‐1, primary human macrophages, and peripheral blood mononuclear cells (PBMC). Neopterin formation and tryptophan degradation were induced similarly by IFN‐γ in all three cell types investigated, but the effects of interleukins were different between THP‐1, primary macrophages and PBMC. In PBMC, but not in THP‐1 cells and primary macrophages, IL‐12 was found to be additive to the effects of IFN‐γ to superinduce neopterin formation and tryptophan degradation. IL‐4 and IL‐10 reduced the effects of IFN‐γ on monocytic cells, and both cytokines were additively antagonistic to IFN‐γ in PBMC and THP‐1 cells. Finally, on preincubation, but not on addition of IL‐12, the effects of IL‐4 and IL‐10 on PBMC could be abrogated, whereas no such effect was seen in THP‐1 cells. The results show that IL‐12 up‐regulates neopterin formation and tryptophan degradation by inducing additional IFN‐γ production by Th1 cells, while a direct effect of IL‐12 on monocytes/macrophages appears to be absent. Similarly, IL‐4 and IL‐10 inhibit neopterin production and tryptophan degradation in PBMC by down‐regulating Th1‐type cytokine production and possibly also via direct deactivation of IFN‐γ effects towards monocytes/macrophages. The results clearly show how Th1 cell‐mediated immunity may be up‐ or down‐regulated by endogenous cytokine production.

Monocyte-derived dendritic cells release neopterin

Increased neopterin concentrations in body fluids are found in diseases associated with activated, cell‐mediated immunity including infections, autoimmune diseases, and certain malignancies. Monocytes/macrophages are known to secrete large amounts of neopterin upon stimulation with interferon‐γ (IFN‐γ). Ontogenetically, the major part of dendritic cells (DC) belongs to the myeloid lineage. Therefore, we investigated whether cultured monocyte‐derived DC can elaborate neopterin. Cells were treated with cytokines in the presence or absence of monocyte‐conditioned medium as a maturation stimulus. DC secreted an average 3.5 nmol/l neopterin. In response to IFN‐γ, cells significantly increased their output of neopterin. In distinction to monocytes/macrophages, neopterin production in DC was highly sensitive to IFN‐α and IFN‐β. Further, lipopolysaccharides (LPS) enhanced neopterin synthesis, whereas tumor necrosis factor α, interleukin (IL)‐1β, IL‐2, IL‐10, and IL‐18 were ineffective. Simultaneously, tryptophan degradation by induction of indoleamine (2,3)‐dioxygenase (IDO) was tested in stimulated cells. Our results showed that IFN‐γ as well as LPS are inducers of IDO in DC.

Hyperhomocysteinemia in dementia

Summary. Hyperhomocysteinemia is a strong risk factor for atherosclerotic vascular disease, and elevated serum homocysteine is correlated with vitamin B deficiency. In this pilot study, significantly elevated homocysteine levels were found in patients with Alzheimers disease as well as in patients with vascular dementia, probably indicating similar pathophysiological pathways. We found significant correlations between low folic acid concentrations as well as high homocysteine concentrations and cognitive decline. Supplementation with folic acid may be an inexpensive way to reduce elevated homocysteine levels in demented patients.

Interferon gama induced Tryptophan Degradation Neuropsychiatric and Immunological Consequences

Tryptophan is a constituent of proteins and in parallel it represents a source for mainly two pivotal biochemical pathways: the generation of 5-hydroxytryptamine (serotonin), and the formation of kynurenine by the enzymes tryptophan pyrrolase (TP) and indoleamine 2,3-dioxygenase (IDO). IDO is induced by interferon-gamma (IFN-gamma) in a broad variety of cells. Therefore, enhanced tryptophan degradation is observed in diseases and disorders concomitant with cellular immune activation, e.g. infectious diseases, autoimmune diseases, malignant diseases as well as in pregnancy. IFN-gamma-derived tryptophan degradation may represent an effector mechanism within in the comprehensive network of immune stimulation. In addition, the cytostatic and, respectively, antiproliferative properties on e.g., T-lymphocytes may contribute to the immunomodulatory function of IFN-gamma. However, especially in states of persistent immune activation increased tryptophan catabolism leads to the depletion of free serum tryptophan and to the accumulation of neuroactive kynurenine metabolites. As a consequence, serotonergic functions may be affected, and the neurotoxic properties of kynurenine derivatives may lead to neuronal disorders evoking neurological/psychiatric symptoms. This notion provides a basis for the better understanding of mood disorders and related syptoms in chronic diseases. Moreover, IDO could represent a link between the immunological network and neuroendocrine functions with far reaching consequences regarding to the psychological status of patients.

Increased neopterin production and tryptophan degradation in advanced Parkinson's disease.

Summary. Large amounts of neopterin are produced by interferon-(IFN)-γ-stimulated human monocytes/macrophages, and increased neopterin concentrations indicate cellular immune activation. In parallel, IFN-γ induces indoleamine 2,3-dioxygenase which degrades l_-tryptophan to kynurenine. Increased tryptophan degradation rates are indicated by an increased kynurenine/tryptophan ratio (kyn/trp-ratio), reflecting immune system activation, too. In 22 patients with Parkinsons disease (PD) and in 11 age-matched controls, serum and cerebrospinal fluid (CSF) neopterin concentrations were measured by ELISA. Tryptophan and kynurenine concentrations were determined by HPLC. Neopterin concentrations and kyn/trp-ratios were increased both in serum and CSF of patients as compared to controls. Serum tryptophan was lower in patients. Patients with the highest disease activity presented with highest degree of immune activation. Significant correlations existed between neopterin concentrations and kyn/trp-ratios in serum and CSF. Increased formation of neopterin and enhanced degradation of tryptophan suggest activated cell-mediated immune response in a subgroup of patients with advanced Parkinsons disease.

Enhanced Tryptophan Degradation in Systemic Lupus Erythematosus

In vitro and in vivo, tryptophan degradation was found to be associated with T cell functional loss and tolerance induction. In systemic lupus erythematosus (SLE) besides the Th2-type cytokine interleukin-10, Th1-type cytokines including interferon-gamma (IFN-gamma) are expressed especially during exacerbation of the disease. IFN-gamma stimulates the enzyme indoleamine (2,3)-dioxygenase (IDO) converting tryptophan to the metabolite kynurenine which in macrophages is subsequently degraded to other, partly neurotoxic compounds like quinolinic acid, and finally to nicrotinamides. We measured kynurenine and tryptophan concentrations in the sera of 55 SLE patients. In these patients, the concentrations of tryptophan (median, interquartile range: 53.9, 45.7-64.1 microM) were lower (p < 0.0001), and the kynurenine concentrations (2.45, 1.75-3.40 microM) were increased (p < 0.0005) compared to healthy blood donors (70.0, 63.8-80.6; 1.80, 1.45-2.27 microM, respectively). Also the kynurenine per tryptophan quotients (K/T), which allow to estimate IDO activity, were significantly higher in patients than in normals (0.043, 0.033-0.062 vs. 0.027, 0.021-0.030; p < 0.0001), indicating enhanced IDO-induced tryptophan degradation in SLE. There was no significant relationship between tryptophan, kynurenine and the SLEDAI, and also the correlation of K/T with SLEDAI was rather weak (rs = 0.243, p < 0.05). Higher K/T was found in patients presenting with serositis (p = 0.01), decrease of complement (c3, c4; p < 0.01) and blood count change (anemia, leucopenia, lymphopenia; p = 0.032) than in patients without such disease manifestations. The significant correlation found between K/T and neopterin (rs = 0.808, p < 0.001), a marker of immune activation, points to a role of immune activation to be responsible for tryptophan degradation in SLE patients.