Crystal structure of human indoleamine 2,3-dioxygenase: catalytic mechanism of O2 incorporation by a heme-containing dioxygenase

Human indoleamine 2,3-dioxygenase (IDO) catalyzes the cleavage of the pyrrol ring of L-Trp and incorporates both atoms of a molecule of oxygen (O2). Here we report on the x-ray crystal structure of human IDO, complexed with the ligand inhibitor 4-phenylimidazole and cyanide. The overall structure of IDO shows two alpha-helical domains with the heme between them. A264 of the flexible loop in the heme distal side is in close proximity to the iron. A mutant analysis shows that none of the polar amino acid residues in the distal heme pocket are essential for activity, suggesting that, unlike the heme-containing monooxygenases (i.e., peroxidase and cytochrome P450), no protein group of IDO is essential in dioxygen activation or proton abstraction. These characteristics of the IDO structure provide support for a reaction mechanism involving the abstraction of a proton from the substrate by iron-bound dioxygen. Inactive mutants (F226A, F227A, and R231A) retain substrate-binding affinity, and an electron density map reveals that 2-(N-cyclohexylamino)ethane sulfonic acid is bound to these residues, mimicking the substrate. These findings suggest that strict shape complementarities between the indole ring of the substrate and the protein side chains are required, not for binding, but, rather, to permit the interaction between the substrate and iron-bound dioxygen in the first step of the reaction. This study provides the structural basis for a heme-containing dioxygenase mechanism, a missing piece in our understanding of heme chemistry.     

Cerebral hemodynamics evaluation by near-infrared time-resolved spectroscopy: correlation with simultaneous positron emission tomography measurements

We compared pharmacologically-perturbed hemodynamic parameters (cerebral blood volume; CBV, and flow; CBF) by acetazolamide administration in six healthy human subjects studied with positron emission tomography (PET) and near-infrared (NIR) time-resolved spectroscopy (TRS) simultaneously to investigate whether NIR-TRS could measure in vivo hemodynamics in the brain tissue quantitatively. Simultaneously with the PET measurements, TRS measurements were performed at the forehead with four different optode spacing from 2 cm to 5 cm. Total hemoglobin and oxygen saturation (SO2) measured by TRS significantly increased after administration of acetazolamide at any optode spacing in all subjects. In PET study, CBV and CBF were estimated in the following three volumes of interest (VOIs) determined on magnetic resonance images, VOI1: scalp and skull, VOI2: gray matter region, VOI3: gray and white matter regions. Acetazolamide treatment elevated CBF and CBV significantly in VOI2 and VOI3 but VOI1. TRS-derived CBV was more strongly correlated with PET-derived counterpart in VOI2 and VOI3 when the optode spacing was above 4 cm, although optical signal from cerebral tissue could be caught with any optode spacing. As to increase of the CBV, 4 cm of optode spacing correlated best with VOI2. To support the result of TRS-PET experiment, we also estimated the contribution ratios of intracerebral tissue to observed absorption change based on diffusion theory. The contribution ratios at 4 cm were estimated as follows: 761 nm: 50%, 791 nm: 72%, 836 nm: 70%. These results demonstrated that NIR-TRS with 4 cm of optode spacing could measure cerebral hemodynamic responses optimally and quantitatively.     

MAP-based kinetic analysis for voxel-by-voxel compartment model estimation: detailed imaging of the cerebral glucose metabolism using FDG

We propose a novel algorithm for voxel-by-voxel compartment model analysis based on a maximum a posteriori (MAP) algorithm. Voxel-by-voxel compartment model analysis can derive functional images of living tissues, but it suffers from high noise statistics in voxel-based PET data and extended calculation times. We initially set up a feature space of the target radiopharmaceutical composed of a measured plasma time activity curve and a set of compartment model parameters, and measured the noise distribution of the PET data. The dynamic PET data were projected onto the feature space, and then clustered using the Mahalanobis distance. Our method was validated using simulation studies, and compared with ROI-based ordinary kinetic analysis for FDG. The parametric images exhibited an acceptable linear relation with the simulations and the ROI-based results, and the calculation time took about 10 min. We therefore concluded that our proposed MAP-based algorithm is practical.     

Pathways for oxidation of high-density lipoprotein in human cardiovascular disease

The cardioprotective effect of high-density lipoprotein (HDL) is thought to involve, in part, the membrane-associated ATP-binding cassette transporter ABCA1, which clears cholesterol from lipid-laden macrophages in the artery wall. If HDL is unable to interact with this transporter because of oxidative damage, cholesterol clearance is impaired. Important insights into the mechanisms that oxidize proteins in the human artery wall have come from the mass spectrometric (MS) detection of oxidized amino acids that result from specific reaction pathways. Recent MS studies indicate that HDL isolated from patients with cardiovascular disease contains elevated levels of 3-chlorotyrosine and 3-nitrotyrosine, which are two characteristic products of myeloperoxidase (MPO), a heme enzyme secreted by macrophages. MPO-dependent chlorination of apolipoprotein A-I (apoA-I), the major HDL protein, impairs its ability to remove excess cellular cholesterol by the ABCA1 pathway. Tandem MS analysis of apoA-I has demonstrated that this loss of activity is associated with methionine oxidation and chlorination of a single tyrosine residue. Analysis of mutated forms of apoA-I has implicated lysine residues in the regiospecific chlorination of tyrosine. It is further suggested that the tyrosine chlorination and methionine oxidation of apoA-I impairs ABCA1 transport activity. The crystal structure of lipid-free apoA-I suggests a potential mechanism for rendering this protein dysfunctional. Collectively, these observations indicate that MPO oxidatively damages HDL in humans and suggest that oxidation of specific amino acid residues in apoA-I may contribute to atherogenesis by impairing cholesterol efflux from macrophages.     

Effect of voluntary attention on auditory processing during REM sleep

STUDY OBJECTIVES: The study investigates whether there is an effect of voluntary attention to external auditory stimuli during rapid eye movement (REM) sleep in humans by measuring event-related potentials (ERPs). DESIGN: Using a 2-tone auditory-discrimination task, a standard 1000-Hz tone and a deviant 2000-Hz tone were presented to participants when awake and during sleep. In the ATTENTIVE condition, participants were requested to detect the deviant stimuli during their sleep whenever possible. In the PASSIVE sleep condition, participants were only exposed to the tones. ERPs were measured during REM sleep and compared between the 2 conditions. SETTING: All experiments were conducted at the sleep laboratory of Hiroshima University. PARTICIPANTS: Twenty healthy university student volunteers. INTERVENTIONS: N/A. Measurements and Results: In the tonic period of REM sleep (the period without REM), P200 and P400 were elicited by deviant stimuli, with scalp distributions maximal at central and occipital sites, respectively. The P400 in REM sleep showed larger amplitudes in the ATTENTIVE condition, whereas the P200 amplitude did not differ between the 2 conditions. No effects on ERPs due to attention were observed during stage 2 sleep. CONCLUSIONS: The instruction to pay attention to external stimuli during REM sleep influenced the late positive potentials. Thus electrophysiologic evidence of voluntary attention during REM sleep has been demonstrated.     

Excessive daytime sleepiness in Parkinson disease: a SPECT study

STUDY OBJECTIVES: The underlying pathologic mechanism of excessive daytime sleepiness (EDS) in Parkinson disease and the relative contributions of brain function to this process are poorly understood. We compared brain perfusion images between patients with Parkinson disease and EDS and those without EDS using n-isopropyl-p-1231 iodoamphetamine single photon emission computed tomography. DESIGN: Clinical study. SETTING: Sumitomo Hospital. PATIENTS: Thirteen patients with Parkinson disease with EDS (EDS group) and 27 patients with Parkinson disease without EDS (no-EDS group) were studied. Whether or not each case had EDS was determined according to the response to the Epworth Sleepiness Scale: patients with an Epworth Sleepiness Scale score > or = 10 were included in the EDS group, and patients with an Epworth Sleepiness Scale score < or = 9 were included in the no-EDS group. MEASUREMENTS AND RESULTS: There were significant hypoperfusions in the left parietal and temporal association cortex in the EDS group. In the multivariable logistic regression model, attention and decreased regional cerebral blood flow of the left parietal association cortex and right caudate and increased regional cerebral blood flow of the right thalamus were the independent and significant factors. CONCLUSIONS: The cortical hypofunction relative to hyperfunction of the brain stem may relate to EDS in Parkinson disease. This is the first imaging study about EDS in Parkinson disease, and further studies are required.     

Mechanism by which Bombyx mori hemocytes recognize microorganisms: direct and indirect recognition systems for PAMPs

Hemocytes play an important role in cellular reactions in the immune system. Although the recognition of pathogens is thought to involve pattern-recognition proteins (PRPs) in insects, the exact mechanisms by which insect hemocytes recognize pathogens are not clear. This study examined the mechanism by which Bombyx mori hemocytes recognize microorganisms and pathogen-associated molecular patterns (PAMPs) using flow cytometry and fluorescence microscopy. Fluorescence-labeled bacterial or fungal cells were observed to bind to hemocytes and this binding was inhibited by adding lipoteichoic acid (LTA) or beta-1,3-glucan. Lipopolysaccharide (LPS) bound to hemocytes directly. These results suggest that hemocytes have a mechanism that recognizes LPS, LTA, and beta-1,3-glucan directly. Previously, we identified two types of C-type lectin (BmLBP and BmMBP) and showed that they recognize a variety of PAMPs leading to the induction of nodule formation. These lectins enhanced hemocyte binding to microorganisms and their direct binding to hemocytes suggests that hemocytes have a mechanism for recognizing microorganisms using lectin receptors.     

A case of anaplastic ductal carcinoma of the pancreas with production of granulocyte-colony stimulating factor

We report a case of anaplastic ductal carcinoma of the pancreas with production of granulocytecolony stimulating factor in a 46-year-old man. He underwent distal pancreatectomy, total gastrectomy with Roux-en-Y reconstruction and splenectomy. He recovered uneventfully postoperatively and was discharged from hospital on the 22nd postoperative day. Histological examination showed anaplastic ductal carcinoma (giant cell type) of the pancreas. However, three months after the operation, multiple liver metastases were demonstrated by CT scan and the peripheral leukocyte count rose to 81,800/microL. Serum granulocyte-colony stimulating factor concentration was 155pg/mL (normal 5.78-27.5 pg/mL). Therefore, we diagnosed granulocyte-colony stimulating factor-producing pancreatic cancer. Anaplastic ductal carcinoma of the pancreas with production of granulocyte-colony stimulating factor is very rare. Tumors with production of granulocyte-colony stimulating factor are highly malignant, and this patient died of hepatic failure four months after the operation.