Serine protease modulation of Dependence Receptors and EMT protein expression.

Expression of the tumour suppressor Deleted in Colorectal Cancer (DCC) and the related protein neogenin is reduced by the mammalian serine protease chymotrypsin or the bacterial serine protease subtilisin, with increased cell migration. The present work examines whether these actions are associated with changes in the expression of cadherins, β-catenin and vimentin, established markers of the Epithelial-Mesenchymal Transition (EMT) which has been linked with cell migration and tumour metastasis. The results confirm the depletion of DCC and neogenin and show that chymotrypsin and subtilisin also reduce expression of β-catenin in acutely prepared tissue sections but not in human mammary adenocarcinoma MCF-7 or MDA-MB-231 cells cultured in normal media, or primary normal human breast cells.

Kynurenine Pathway Activation in Human African Trypanosomiasis.

Background: The kynurenine pathway of tryptophan oxidation is associated with central nervous system (CNS) inflammatory pathways. Inhibition of this pathway ameliorates CNS inflammation in rodent models of the late (meningoencephalitic) stage of human African trypanosomiasis (HAT). In this study, we evaluate whether the kynurenine pathway is activated in clinical HAT and associated with CNS inflammatory responses.

Quinolinic acid induces neuritogenesis in SH-SY5Y neuroblastoma cells independently of NMDA receptor activation.

Glutamate and nicotinamide adenine dinucleotide (NAD ) have been implicated in neuronal development and several types of cancer. The kynurenine pathway of tryptophan metabolism includes quinolinic acid (QA) which is both a selective agonist at N-methyl-D-aspartate (NMDA) receptors and also a precursor for the formation of NAD . The effect of QA on cell survival and differentiation has therefore been examined on SH-SY5Y human neuroblastoma cells.

Selective depletion of tumour suppressors Deleted in Colorectal Cancer (DCC) and neogenin by environmental and endogenous serine proteases: linking diet and cancer.

Background: The related tumour suppressor proteins Deleted in Colorectal Cancer (DCC) and neogenin are absent or weakly expressed in many cancers, whereas their insertion into cells suppresses oncogenic behaviour. Serine proteases influence the initiation and progression of cancers although the mechanisms are unknown.

Methods: The effects of environmental (bacterial subtilisin) and endogenous mammalian (chymotrypsin) serine proteases were examined on protein expression in fresh, normal tissue and human neuroblastoma and mammary adenocarcinoma lines.

Kynurenine pathway metabolism following prenatal KMO inhibition and in Mecp2 mice, using liquid chromatography-tandem mass spectrometry.

To quantify the full range of tryptophan metabolites along the kynurenine pathway, a liquid chromatography - tandem mass spectrometry method was developed and used to analyse brain extracts of rodents treated with the kynurenine-3-mono-oxygenase (KMO) inhibitor Ro61-8048 during pregnancy. There were significant increases in the levels of kynurenine, kynurenic acid, anthranilic acid and 3-hydroxy-kynurenine (3-HK) in the maternal brain after 5 h but not 24 h, while the embryos exhibited high levels of kynurenine, kynurenic acid and anthranilic acid after 5 h which were maintained at 24 h post-treatment. At 24 h there was also a strong trend to an increase in quinolinic acid levels (P = 0.

Dependence receptor involvement in subtilisin-induced long-term depression and in long-term potentiation.

The serine protease subtilisin induces a form of long-term depression (LTD) which is accompanied by a reduced expression of the axo-dendritic guidance molecule Unco-ordinated-5C (Unc-5C). One objective of the present work was to determine whether a loss of Unc-5C function contributed to subtilisin-induced LTD by using Unc-5C antibodies in combination with the pore-forming agents Triton X-100 (0.005%) or streptolysin O in rat hippocampal slices.

Modified neocortical and cerebellar protein expression and morphology in adult rats following prenatal inhibition of the kynurenine pathway.

Inhibition of the kynurenine pathway of tryptophan metabolism during gestation can lead to changes in synaptic transmission, neuronal morphology and plasticity in the rat hippocampus. This suggests a role for the kynurenine pathway in early brain development, probably caused by kynurenine modulation of N-methyl-d-aspartate (NMDA) glutamate receptors which are activated by the tryptophan metabolite quinolinic acid and blocked by kynurenic acid. We have now examined samples of neocortex and cerebellum of adult animals to assess the effects of a prenatally administered kynurenine-3-monoxygenase inhibitor (Ro61-8048) on protein and mRNA expression, dendritic structure and immuno-histochemistry.

Prenatal inhibition of the kynurenine pathway leads to structural changes in the hippocampus of adult rat offspring.

Glutamate receptors for N-methyl-d-aspartate (NMDA) are involved in early brain development. The kynurenine pathway of tryptophan metabolism includes the NMDA receptor agonist quinolinic acid and the antagonist kynurenic acid. We now report that prenatal inhibition of the pathway in rats with 3,4-dimethoxy-N-[4-(3-nitrophenyl)thiazol-2-yl]benzenesulphonamide (Ro61-8048) produces marked changes in hippocampal neuron morphology, spine density and the immunocytochemical localisation of developmental proteins in the offspring at postnatal day 60.

Prenatal activation of maternal TLR3 receptors by viral-mimetic poly(I:C) modifies GluN2B expression in embryos and sonic hedgehog in offspring in the absence of kynurenine pathway activation.

Activation of the immune system during pregnancy is believed to lead to psychiatric and neurological disorders in the offspring, but the molecular changes responsible are unknown. Polyinosinic:polycytidylic acid (poly(I:C)) is a viral-mimetic double-stranded RNA complex which activates Toll-Like-Receptor-3 and can activate the metabolism of tryptophan through the oxidative kynurenine pathway to compounds that modulate activity of glutamate receptors. The aim was to determine whether prenatal administration of poly(I:C) affects the expression of neurodevelopmental proteins in the offspring and whether such effects were mediated via the kynurenine pathway.

Prenatal inhibition of the tryptophan-kynurenine pathway alters synaptic plasticity and protein expression in the rat hippocampus.

Glutamate receptors sensitive to N-methyl-d-aspartate (NMDA) are important in early brain development, influencing cell proliferation and migration, neuritogenesis, axon guidance and synapse formation. The kynurenine pathway of tryptophan metabolism includes an agonist (quinolinic acid) and an antagonist (kynurenic acid) at these receptors. Rats were treated in late gestation with 3,4-dimethoxy-N-[4-(3-nitrophenyl)thiazol-2-yl]-benzene-sulphonamide (Ro61-8048), an inhibitor of kynurenine-3-monoxygenase which diverts kynurenine metabolism to kynurenic acid.

Prenatal activation of Toll-like receptors-3 by administration of the viral mimetic poly(I:C) changes synaptic proteins, N-methyl-D-aspartate receptors and neurogenesis markers in offspring.

Background: There is mounting evidence for a neurodevelopmental basis for disorders such as autism and schizophrenia, in which prenatal or early postnatal events may influence brain development and predispose the young to develop these and related disorders. We have now investigated the effect of a prenatal immune challenge on brain development in the offspring. Pregnant rats were treated with the double-stranded RNA polyinosinic:polycytidylic acid (poly(I:C); 10 mg/kg) which mimics immune activation occurring after activation of Toll-like receptors-3 (TLR3) by viral infection.

Kynurenine pathway inhibition as a therapeutic strategy for neuroprotection.

The oxidative pathway for the metabolism of tryptophan along the kynurenine pathway generates quinolinic acid, an agonist at N-methyl-D-aspartate receptors, as well as kynurenic acid which is an antagonist at glutamate and nicotinic receptors. The pathway has become recognized as a key player in the mechanisms of neuronal damage and neurodegenerative disorders. As a result, manipulation of the pathway, so that the balance between the levels of components of the pathway can be modified, has become an attractive target for the development of pharmacological agents with the potential to treat those disorders.

Molecular changes associated with hippocampal long-lasting depression induced by the serine protease subtilisin-A.

The serine protease subtilisin-A (SubA) induces a form of long-term depression (LTD) of synaptic transmission in the rat hippocampus, and molecular changes associated with SubA-induced LTD (SubA-LTD) were explored by using recordings of evoked postsynaptic potentials and immunoblotting. SubA-LTD was prevented by a selective inhibitor of SubA proteolysis, but the same inhibitor did not affect LTD induced by electrical stimulation or activation of metabotropic glutamate receptors. SubA-LTD was reduced by the protein kinase inhibitors genistein and lavendustin A, although not by inhibitors of p38 mitogen-activated protein kinase, glycogen synthase kinase-3, or protein phosphatases.

Kynurenine metabolism predicts cognitive function in patients following cardiac bypass and thoracic surgery.

Cardiac surgery involving extra-corporeal circulation can lead to cognitive dysfunction. As such surgery is associated with signs of inflammation and pro-inflammatory mediators activate tryptophan oxidation to neuroactive kynurenines which modulate NMDA receptor function and oxidative stress, we have measured blood concentrations of kynurenines and inflammatory markers in 28 patients undergoing coronary arterial graft surgery and, for comparison, 28 patients undergoing non-bypass thoracic surgery. A battery of cognitive tests was completed before and after the operations.

Involvement of kynurenines in Huntington's disease and stroke-induced brain damage.

Several components of the kynurenine pathway of tryptophan metabolism are now recognised to have actions of profound biological importance. These include the ability to modulate the activation of glutamate and nicotinic receptors, to modify the responsiveness of the immune system to inflammation and infection, and to modify the generation and removal of reactive oxygen species. As each of these factors is being recognised increasingly as contributing to major disorders of the central nervous system (CNS), so the potentially fundamental role of the kynurenine pathway in those disorders is presenting a valuable target both for understanding the progress of those disorders and for developing potential drug treatments.

On the Biological Importance of the 3-hydroxyanthranilic Acid: Anthranilic Acid Ratio.

Of the major components of the kynurenine pathway for the oxidative metabolism of tryptophan, most attention has focussed on the N-methyl-D-aspartate (NMDA) receptor agonist quinolinic acid, and the glutamate receptor blocker kynurenic acid. However, there is increasing evidence that the redox-active compound 3-hydroxyanthranilic acid may also have potent actions on cell function in the nervous and immune systems, and recent clinical data show marked changes in the levels of this compound, associated with changes in anthranilic acid levels, in patients with a range of neurological and other disorders including osteoporosis, chronic brain injury, Huntington's disease, coronary heart disease, thoracic disease, stroke and depression. In most cases, there is a decrease in 3-hydroxyanthranilic acid levels and an increase in anthranilic acid levels.

Kynurenine metabolites and inflammation markers in depressed patients treated with fluoxetine or counselling.

1. Depression could result from changes in tryptophan availability caused by activation of the kynurenine pathway as a result of inflammation. In the present study, we examined patients newly diagnosed with depression to determine whether kynurenines and related factors change in parallel with improvements in mood.

Inflammatory status and kynurenine metabolism in rheumatoid arthritis treated with melatonin.

Aim: Since melatonin is antioxidant and has some anti-inflammatory actions, we have tested it as adjunctive treatment in patients with rheumatoid arthritis, to determine whether it can improve patients' symptoms.

Methods: A total of 75 patients were allocated randomly to receive melatonin 10 mg at night in addition to ongoing medication, or a placebo of identical appearance. Monthly blood samples were taken and disease severity assessed over 6 months, plasma being analysed for inflammatory indicators [C-reactive protein, erythrocyte sedimentation rate (ESR), neopterin], proinflammatory cytokines [interleukin (IL)-1beta, IL-6, tumour necrosis factor (TNF)-alpha], lipid peroxidation products and the kynurenine pathway metabolites of tryptophan.

Kynurenine pathway metabolism in patients with osteoporosis after 2 years of drug treatment.

1. Metabolism of tryptophan along the oxidative pathway via kynurenine results in the production of quinolinic acid and kynurenic acid, which can act on glutamate receptors in peripheral tissues. We have now measured the concentrations of kynurenine pathway metabolites in the plasma of patients with osteoporosis before treatment with drugs, throughout and after 2 years of treatment with the drugs raloxifene or etidronate.

Adenosine and cytokine levels following treatment of rheumatoid arthritis with dipyridamole.

Adenosine can suppress the release of tumour necrosis factor-alpha (TNF-alpha) from activated monocytes and macrophages, and may contribute to the anti-inflammatory activities of methotrexate and sulphasalazine. Dipyridamole inhibits the cellular uptake and metabolism of adenosine and we have, therefore, examined the effects of dipyridamole in patients with rheumatoid arthritis in an attempt to alleviate their symptoms. Forty patients aged 18-75 years were randomised to receive dipyridamole 400 mg/day or placebo.

Levels of purine, kynurenine and lipid peroxidation products in patients with inflammatory bowel disease.

The factors affecting gut activity in inflammatory bowel disease are unclear, but purines and kynurenines may be involved in the regulation of neuronal activity and therefore gut motility and secretion. We have measured the serum levels of these compounds in patients and in sex- and age-matched controls. Purines and kynurenines were analysed using HPLC.

Kynurenine and neopterin levels in patients with rheumatoid arthritis and osteoporosis during drug treatment.

The kynurenine pathway from tryptophan generates compounds which can act on glutamate receptors in peripheral tissues or modulate free radical activity. We have measured the concentrations of several of these compounds in the plasma of patients with rheumatoid arthritis (RA) and osteoporosis (OP) before treatment with drugs and then at monthly intervals for 6 months during treatment. Kynurenine analysis was performed by HPLC.

Tryptophan metabolites and brain disorders.

Tryptophan is metabolised primarily along the kynurenine pathway, of which two components are now known to have marked effects on neurons in the central nervous system. Quinolinic acid is an agonist at the population of glutamate receptors which are sensitive to N-methyl-D-aspartate (NMDA), and kynurenic acid is an antagonist at several glutamate receptors. Consequently quinolinic acid can act as a neurotoxin while kynurenic acid is neuroprotectant.

Purine, kynurenine, neopterin and lipid peroxidation levels in inflammatory bowel disease.

The kynurenine metabolites of tryptophan may be involved in the regulation of neuronal activity and thus gut motility and secretion. We have now performed a pilot study to measure serum concentrations of purines and kynurenines in patients with mild inflammatory bowel disease, as well as in sex- and age-matched control subjects. For some analyses, the patients were subdivided into subgroups of those with Crohn's disease and those with ulcerative colitis.