Effect of resistance training on resting metabolic rate and its estimation by a dual-energy X-ray absorptiometry metabolic map.

BACKGROUND/OBJECTIVES: Fat-free mass (FFM) is the major predictor of resting metabolic rate (RMR). As protein supplementation during resistance training may augment gains in FFM, we investigated the effects of resistance training combined with protein supplementation on RMR and whether RMR responses could be estimated by a dual-energy X-ray absorptiometry (DXA) metabolic map. SUBJECTS/METHODS: Healthy adults completed a whole-body periodized resistance training program consisting of 96 workouts (~9 months). Participants were randomly assigned to supplement with whey protein (whey; n=18), soy protein (soy; n=21) or carbohydrate (carb; n=22). RMR was measured using indirect calorimetry (RMR(IC)) and estimated by DXA metabolic mapping (RMR(MM)) pretraining and posttraining. RESULTS: RMR(IC) increased from pretraining to posttraining in the whole cohort (1653±302 to 1726±291 kcal/day, P=0.001) without differences between the groups. Delta RMR(IC) and RMR(MM) (73±158 vs 52±41 kcal/day were not significantly different by t-test (P=0.303), although they were not significantly correlated (r=0.081; P=0.535). Stepwise regression identified 43% of the shared variance in delta RMR(IC) using total serum thyroxine, RMR(IC) and FFM at baseline (P=0.009). CONCLUSIONS: These results indicate that 9 months of resistance training significantly increased RMR ~5% on average, but there was wide variability between individuals, which can be partially accounted for by changes in FFM and thyroid hormones.

Histological findings on fetal striatal grafts in a Huntington's disease patient early after transplantation.

Cell transplantation is a promising therapeutic approach that has the potential to replace damaged host striatal neurons and, thereby, slow down or even reverse clinical signs and symptoms during the otherwise fatal course of Huntington's disease (HD). Open-labeled clinical trials with fetal neural transplantation for HD have demonstrated long-term clinical benefits for HD patients. Here we report a postmortem analysis of an individual with HD 6 months after cell transplantation and demonstrate that cells derived from grafted fetal striatal tissue had developed into graft-derived neurons expressing dopamine-receptor related phosphoprotein (32 kDa) (DARPP-32), neuronal nuclear antigen (NeuN), calretinin and somatostatin. However, a fully mature phenotype, considered by the expression of developmental markers, is not reached by engrafted neurons and not all types of interneurons are being replaced at 6 months, which is the earliest time point human fetal tissue being implanted in a human brain became available for histological analysis. Host-derived tyrosine hydroxylase (TH) fibers had already heavily innervated the transplants and formed synaptic contacts with graft-derived DARPP-32 positive striatal neurons. In parallel, the transplants contained a considerable number of immature neuroepithelial cells (doublecortin+, Sox2+, Prox-1+, ss3-tubulin+) that exhibited a pronounced migration into the surrounding host striatal tissue and considerable mitotic activity. Graft-derived astrocytes could also be found. Interestingly, the immunological host response in the grafted area showed localized increase of immunocompetent host cells within perivascular spaces without deleterious effects on engrafted cells under continuous triple immunosuppressive medication. Thus this study provides for a better understanding of the developmental processes of grafted human fetal striatal neurons in HD and, in addition, has implications for stem cell-based transplantation approaches in the CNS.

Expression and function of cytochrome p450 in brain drug metabolism.

Cytochrome P450 (CYP, P450) is the collective term for a superfamily of heme-containing membrane proteins responsible for the metabolism of approximately 70 - 80 % of clinically used drugs. Besides the liver and other peripheral organs, P450 isoforms are expressed in glial cells and neurons of the brain. To enlighten their function and significance is a topic of high interest, as most of the neuroactive drugs used in therapy today are not only substrates, but also inducers of brain P450s with far reaching consequences. First of all, brain P450s are regulated differentially from those in liver. The availability of the prosthetic heme group appears to be essential for correct membrane insertion and enzymatic functionality of brain P450s. Furthermore, although not contributing to body's overall drug metabolism, brain P450s fulfil particular functions within specific cell types of the brain. In astrocytes of brain's border lines P450 isoforms are expressed at very high level. They form a metabolic barrier regulating drugs' influx, modulate blood-flow regulation, and act as signalling enzymes in inflammation. In neurons, however, P450s apparently have different function. In specified brain regions such as hypothalamus, hippocampus and striatum they provide signalling molecules like steroids and fatty acids necessary for neuronal outgrowth and maintenance. Induction of these P450s by neuroactive drugs can alter steroid hormone signalling directly in drug target cells, which may cause clinically relevant side effects like reproductive disorders and sexual or mental dysfunction. The understanding of brain P450 function appears to be of major interest in long-term drug mediated therapy of neurological diseases.

Primary CNS lymphoma in immunocompetent patients from 1989 to 2001: a retrospective analysis of 164 cases uniformly diagnosed by stereotactic biopsy.

BACKGROUND: We present outcome data of a cohort of 164 immunocompetent PCNSL patients uniformly diagnosed at a single center for stereotactic neurosurgery, and evaluate the acceptance and impact of combination radiotherapy (RT) and chemotherapy (CHT) with high-dose methotrexate (HD-MTX) over time. METHOD: We assessed choice of treatment and patient survival in a series of 164 PCNSL cases diagnosed from 1989 to 2001, and performed a re-evaluation of histopathology and pre-operative clinical data. FINDINGS: From 1989 to 1993, RT was the predominant therapy, and additional CHT did not improve survival. After 1994, the use of combination CHT/RT increased continuously, consistently contained MTX, and was associated with longer survival than RT only: median survival was 14 months after CHT/RT (2-year survival 35.7%) and 10 months (2-year survival 26.2%) after RT only (not significant). Overall median survival remained poor, increasing from six (1989-1993) to nine months (1994-2001) (p = 0.008). Survival was variable, with a few patients surviving >4 years after diagnosis in the CHT/RT as well as in the RT only group. CONCLUSIONS: Despite considerable improvement of PCNSL therapy, the overall benefit of combined CHT/RT versus RT only was lower than that expected from previous phase II clinical trials. The striking variability of survival in either treatment group may suggest a yet undefined biological heterogeneity of PCNSL, which may also include a more aggressive PCNSL subtype in the group of patients with rapidly progressive disease and not eligible for standard therapy.

Microglial expression of prostaglandin EP3 receptor in excitotoxic lesions in the rat striatum.

Prostaglandins (PG) are produced by the enzymatic activity of cyclooxygenase (COX). PGs and COX have been implicated in the pathophysiology of excitotoxicity and neurodegeneration in the central nervous system (CNS). The PGE2 receptor EP3 is the most abundantly expressed PGE2 receptor subtype in the brain. So far, in the innate rat brain EP3 receptors have been found exclusively in neurons. The aim of this study was to investigate whether EP3 expression in the brain changes under neurodegenerative circumstances such as an acute excitotoxic lesion. Intrastriatal injection of quinolinic acid (QUIN) resulted in a loss of EP3-positive striatal neurons, while simultaneously small glial-shaped EP3-positive cells appeared. Five days after lesioning, 63% of the glial-shaped EP3-positive cells could be identified as ED-1 expressing microglial cells. This percentage increased to 82% after 10 days, suggesting that most of the EP3-positive ED-1-negative cells on day 5 may be microglia which did not yet express ED-1. ED-1-positive microglia also expressed COX-1. These experiments show for the first time that activated microglial cells in excitotoxic lesions express in vivo the PGE2 receptor EP3 and the PGE2 synthesizing enzyme COX-1. Activation of EP3 receptor downregulates cAMP formation and may counteract the upregulation of cAMP formation via EP2 receptors, which has been linked to the anti-inflammatory effects of PGs. This change in EP3-receptor expression in microglia might participate in acute or chronic microglial activation in a variety of brain diseases such as ischemia or Alzheimer's disease (AD). Investigation of the expression of different PGE2 receptor subtypes might promote a better understanding of the pathophysiology of these diseases as well as leading to a modulation of microglial activation by a more specific interference with selective EP receptors than can be achieved by inhibiting global PG synthesis by selective or non-selective COX inhibitors.

Dendritic changes in the hippocampal formation of AIDS patients: a quantitative Golgi study.

We have previously shown that in the hippocampal formation of patients with acquired immunodeficiency syndrome (AIDS) there is neuronal atrophy, without cell loss. Because reductions in neuronal size are suggestive of associated neuritic alterations, we decided to study the dendritic trees of the main neuronal populations in the hippocampal formation. Material was obtained in five male AIDS patients and five male controls. After Golgi impregnation, the dendritic arborizations of dentate granule and hilar basket cells, and of CA3 and CA1 pyramidal cells, were hand traced, and their segments classified, counted and measured. We found an impoverishment of the dendritic trees in all neuronal populations in the AIDS group, which was more striking in the hilus and CA3 field. Specifically, hilar neurons had fewer dendritic segments, and reduced branching density and dendritic extent; in CA3 pyramids there was a decrease in the number of terminal segments in the basal trees, and a reduction in the total number of segments, number of medium order terminals, dendritic branching density and dendritic extent in the apical trees. In CA1 pyramids, the terminals were shorter in the apical trees and the dendritic spine density decreased in the basal trees, whereas in granule cells only the dendritic spine density was reduced in AIDS patients. Subtle signs suggestive of dendritic reorganization were observed. These results point to a regional vulnerability of the hippocampal formation to HIV infection, and might contribute to explaining the occurrence of dementia, as a consequence of overall reduction in the hippocampal neuronal receptive surface.

Interactions of solid lipid nanoparticles with model membranes and leukocytes studied by EPR.

Solid lipid nanoparticles (SLN) are colloidal systems which have been proposed for several administration routes. Only limited data are available about the mechanism and rate of interaction of SLN with cells and tissues. The aim of our study was to investigate interactions of SLN with model membranes (liposomes) and cells (leukocytes). SLN dispersions composed of glyceryl tripalmitate, phosphatidylcholine, water, and poloxamer 188 or Tween 20 were prepared by the melt-emulsification process. Spin-labeled phosphatidylcholine (PC(10,3)) and the methylester of doxyl palmitic acid (MeFASL(10,3)) were incorporated into SLN as spin probes (SPs) in order to determine the rate and mechanism of cell interaction by electron paramagnetic resonance (EPR) spectroscopy. Our results indicate that the exchange of SP between SLN and liposomes is much faster for MeFASL(10,3) than for PC(10,3), probably due to the smaller size of the former. In contrast to liposomes, in leukocytes no significant difference in the transfer rates of the two SP was observed after incubation, suggesting that there is an uptake of SLN to leukocytes (endocytosis) although simultaneous SP diffusion is not excluded. The interaction of SLN with leukocytes appears to depend significantly on the stabilizer used. Transfer of PC(10,3) from SLN coated with poloxamer 188 is much faster than from SLN coated with Tween 20.

Predominantly neuronal expression of cytochrome P450 isoforms CYP3A11 and CYP3A13 in mouse brain.

Despite the very small amounts of cytochrome P450 enzymes expressed in different areas and cell populations of the brain as compared with the liver, there is significant evidence for their specific involvement in brain development, function, and plasticity. Nevertheless, the current discussion about occurrence and importance of cerebral cytochrome P450 isoforms is determined by controversial interpretations of their function in general and with respect to single isoforms. Continuing a series of publications about brain P450 isoforms, we now present evidence for the expression of cytochrome P450 3A11 and 3A13 in mouse brain. Immunocytochemical and non-radioactive in situ hybridization studies revealed identical distribution of their proteins and mRNAs throughout the brain especially in neuronal populations, and to some extent in astrocytes. The cerebral expression of these P450 isoforms was confirmed by Western blot and RNAse protection assay analysis. The well-known testosterone-metabolizing capacity and the inducibility of cytochrome P450 3a isoforms by xenobiotics as well as their presence in steroid hormone-sensitive areas and neurons (e.g. hippocampus) clarify the significance of these isoforms for impairment of steroid hormone actions by P450-inducing environmental substances. Therefore, investigation of inducible cerebral P450 isoforms which are able to metabolize xenobiotics as well as steroid hormones might help us to understand neuroendocrine regulation of brain's plasticity.

Infectious CNS disease as a differential diagnosis in systemic rheumatic diseases: three case reports and a review of the literature.

BACKGROUND: Immunosuppressive treatment of rheumatic diseases may be associated with several opportunistic infections of the brain. The differentiation between primary central nervous system (CNS) involvement and CNS infection may be difficult, leading to delayed diagnosis. OBJECTIVE: To differentiate between CNS involvement and CNS infection in systemic rheumatic diseases. METHODS AND RESULTS: Three patients with either longstanding or suspected systemic rheumatic diseases (systemic lupus erythematodes, Wegener's granulomatosis, and cerebral vasculitis) who presented with various neuropsychiatric symptoms are described. All three patients were pretreated with different immunosuppressive drugs (leflunomide, methotrexate, cyclophosphamide) in combination with corticosteroids. Magnetic resonance imaging of the brain was suggestive of infectious disease, which was confirmed by cerebrospinal fluid analysis or stereotactic brain biopsy (progressive multifocal leucoencephalopathy (PML) in two and nocardiosis in one patient). DISCUSSION: More than 20 cases of PML or cerebral nocardiosis in patients receiving corticosteroids and cytotoxic drugs for rheumatic disease have been reported. The clinical aspects of opportunistic CNS infections and the role of brain imaging, cerebrospinal fluid analysis and stereotactic brain biopsy in the differential diagnosis are reviewed.

Reversible coma with raised intracranial pressure: an unusual clinical manifestation of CADASIL.

A 50-year-old woman presented with recurrent episodes of headache, nausea and disturbed consciousness that were fully reversible within a few days. Clinical and radiological findings suggested raised intracranial pressure, which on one occasion was confirmed by intracranial pressure monitoring. Magnetic resonance imaging performed in the asymptomatic interval disclosed a diffuse leukoencephalopathy. Brain biopsy surprisingly revealed the typical vascular changes of CADASIL and subtle endothelial alterations. The white matter showed edematous changes and reactive gliosis. Mutational analysis of the Notch3 gene revealed a previously unreported mutation. We suggest that a transient disturbance of the blood-brain barrier related to the underlying vascular pathology may have caused this unusual presentation of CADASIL.

Continuous interleukin-6 application in vivo via macroencapsulation of interleukin-6-expressing COS-7 cells induces massive gliosis.

The inflammatory cytokine interleukin-6 (IL-6) was found in senile plaques of Alzheimer's patients and might be involved in the pathology of Parkinson's disease and multiple sclerosis. Interestingly, an astocytosis is also found in these neurodegenerative disorders. To evaluate the direct effects of IL-6 in vivo on glial cells, we created a new in vivo model. IL-6 and mock-transfected (control group) COS-7 cells were encapsulated in a poly-acryl-nitril membrane for implantation into the rat striatum. Afterward, the host immune reaction to the membrane without encapsulated cells and the biological action of IL-6-producing capsules was evaluated. Animals with an implanted membrane without cells showed a moderate astrocytosis 5 days after the operation. Furthermore, microglia and T-cells could be detected and after 30 days the astrocytosis decreased to a small layer around the membrane. In comparison to the control group, which received a sham operation, our results demonstrate that the response of glial cells is caused by the mechanical damage of the surgical procedure itself rather than due to the introduced membrane material. In contrast, we found a massive proliferation and activation of astrocytes and microglia after 10 days by IL-6-secreting capsules, indicating that IL-6 is involved in the induction of gliosis. Control animals that received encapsulated mock-transfected COS-7 cells showed only a weak response. These data point to an involvement of IL-6 in the proliferation and activation of glial cells as seen in neurodegenerative disorders.

Oxidative hydrolysis of scoparone by cytochrome p450 CYP2C29 reveals a novel metabolite.

Regioselective 7-demethylation of scoparone is regularly employed as an indicator of phenobarbital-like induction of rat liver cytochrome P450 isoform CYP2B1, e.g., by the antiepileptic drug phenytoin. After induction with phenobarbital and phenytoin, a new reaction sequence catalyzed by Cyp2c29 was identified in mouse liver microsomes. Cyp2c29-dependent 6-demethylation of scoparone resulted in the formation of isoscopoletin, an intermediate which is susceptible to further oxidation. This subsequent oxidation was also catalyzed by Cyp2c29 with a K(m) of 30,31 microM and a V(max) of 3,41 microM/min x microM P450, and resulted in the formation of the new metabolite 3-[4-methoxy-p-(3,6)-benzoquinone]-2-propenoate. This novel metabolite is the product of two consecutive oxidation reactions, proceeding over isoscopoletin to a putative lactone which is accessible to immediate hydrolysis, due to the onium character of the ring oxygen. This opening of the lactone ring corresponds to an oxidative hydrolysis. Differential oxidation of scoparone can be used as a sensitive indicator for distinguishing between different cytochrome P450 isoforms.

Distinct expression patterns and levels of enzymatic activity of matrix metalloproteinases and their inhibitors in primary brain tumors.

Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) have been implicated in the immense invasive potential and neovascularization of primary brain tumors. We investigated the gene expression profiles of MMPs 1, 2, 3, 7, 9, 12, 13, 14, 16 and of TIMPs 1, 2, 3, and 4 in various primary brain tumors (astrocytoma WHO grade I-III, glioblastoma, PNET, ependymoma III and oligoastrocytoma II) using novel RNase protection assay probe sets. In addition, we determined the level and cellular source of gelatinolytic activity and localized gelatinase B and TIMP-1 RNA. Distinct expression patterns of the MMP and TIMP genes were found in the various brain tumors tested. While the WHO grade I and II tumors had MT1/MT3 ratios below 1, the malignant (grade III and IV) tumors had ratios above 1. Strong expression of TIMP-1 RNA was observed in all malignant tumors and in grade I pilocytic astrocytomas and localized to the walls of neovessels. Quantitative analysis of enzymatic activity in the soluble fraction of protein extracts revealed that in most tumors gelatinases remained in the inactive pro-form. In situ zymography revealed net gelatinolytic activity in neurons of normal brain and in tumor cells and vessel walls of all tumors tested. Immunohistochemistry demonstrated that gelatinase B was localized to vessel walls, to neutrophils in areas of hemorrhage, and in glioblastomas to macrophages. Together these data demonstrate that the different primary brain tumors show distinct regulation of MMP and TIMP genes. The localization of the soluble gelatinase B indicates an association with neovascularization, whereas membrane-bound MMPs may account for the invasive potential of the glial tumor cells.

Expression and localization of the CYP2B subfamily predominantly in neurones of rat brain.

Despite the very small amounts of cytochrome P450 (P450, CYP) enzymes expressed in different areas and cell populations of the brain as compared with the liver, there is significant evidence for their specific involvement in brain development, function and plasticity. Nevertheless, the current discussion about occurrence and importance of cerebral cytochrome P450s is determined by inconsistent interpretations of their function in general and with respect to single isoforms. Continuing a series of publications about brain P450 isoforms, we now present evidence for the constitutive expression of CYP2B1 and CYP2B2 mRNAs in rat brain. Immunocytochemical and non-radioactive in situ hybridization studies revealed the same expression pattern throughout the brain predominantly in neuronal populations, but to some extent in astrocytes of corpus callosum and olfactory bulb. The well known testosterone-metabolizing capacity and the presence of CYP2B isoforms shown in steroid hormone-sensitive areas and neurones (e.g. hippocampus) clarify the significance of isoforms like CYP2B1 and CYP2B2 for impairment of steroid hormone actions by P450 inducing environmental substances. We argue that cerebral P450 isoforms which are induced by xenobiotics and are able to metabolize these as well as endogenous substrates help us to understand fundamental aspects of brain's functioning.

Possible function of astrocyte cytochrome P450 in control of xenobiotic phenytoin in the brain: in vitro studies on murine astrocyte primary cultures.

[4-(14)C]Phenytoin underwent a rapid cellular uptake by diffusion within 5 min when applied in a concentration of 10 microM to mouse brain astrocyte cultures. Subsequently, a slow linear increase of intracellular radioactivity indicated metabolic trapping of the drug, with final concentrations reaching 144 pmol phenytoin/mg protein in the astrocytes. Phenytoin levels from 1 to 10 microM decreased cell viability by 15%. The action of cytochrome P450 present in astrocytes in concentrations of 16-17 pmol P450/mg protein could explain these slight cytotoxic effects by generating intermediate metabolites of phenytoin. In contrast, concentrations of 50 microM strongly inhibited cell proliferation. A Cyp2c29 immunorelated P450 isoform was expressed in nearly all astrocytes in culture. Intracellular [4-(14)C]phenytoin was degraded to its major metabolites dihydrodiol, p-HPPH, and m-HPPH through a P450-dependent reaction with a specific activity of 0.66 pmol/min x mg protein, or 0.12 pmol/min x mg protein as measured in cell homogenates. These data underscore the importance of astrocytes as brain cells active in the detoxification of foreign substrates, but also in their toxification due to reactive metabolites generated during these metabolic processes. After diffusionary influx of drugs and other xenobiotics, the astrocyte P450 monooxygenases perform an essential role in the mediation of toxicity most frequently encountered in highly vulnerable neurons.

Identification, induction and localization of cytochrome P450s of the 3A-subfamily in mouse brain.

Several cytochrome P450 subfamilies are inducible by specific exogenous compounds like the antiepileptic drug phenytoin. Some of these P450 enzymes are involved in the metabolism of gonadal hormones also contributing to neuronal differentiation. CYP3A enzymes have the capacity to catalyze the hydroxylation of testosterone and a wide variety of therapeutic agents, but little is known about the expression and potential function of this subfamily in mouse brain. Here, we report the identification of mouse CYP3A isoforms, their induction and localization in mouse brain. Western blot analysis with anti-CYP3A1 antibodies revealed the phenytoin-inducible expression of CYP3A in brain microsomes, and also a constitutive expression of members of this subfamily in brain mitochondria. Using RT-PCR with a consensus primer pair for known mouse liver CYP3A-isoforms we could demonstrate the expression of CYP3A11 and 3A13 mRNA in mouse brain. Finally, using double immunofluorescence labeling we analyzed the histoanatomical distribution of CYP3A throughout the brain with confocal laser scanning microscopy. We found strong immunoreactivity in neurons of hippocampus and hypothalamic areas which are sensitive to steroid hormones. CYP3A immunoreactivity was apparent also in neurons of the cerebellum, the thalamus and the olfactory bulb. Non-neuronal expression of CYP3A could be found in some astrocyte populations and in vascular as well as ventricular border lines. The presence of CYP3A predominantly in neurons but also in cells contributing to the blood-brain and blood-liquor barrier suggests important roles of this subfamily in mediation of steroid hormone action in mouse brain as well as in preventing the brain from potentially cytotoxic compounds.

Regulation of matrix metalloproteinases and their inhibitor genes in lipopolysaccharide-induced endotoxemia in mice.

An imbalance between matrix metalloproteinases (MMPs) and inhibitors of MMPs (TIMPs) may contribute to tissue destruction that is found in various inflammatory disorders. To determine in an in vivo experimental setting whether the inflammatory reaction in the course of lipopolysaccharide (LPS)-induced endotoxemia causes an altered balance in the MMP/TIMP system, we analyzed the expression of a number of MMP and TIMP genes as well as MMP enzymatic activity in the liver, kidney, spleen, and brain at various time points after systemic injection of different doses of LPS in mice. Injection of sublethal doses of LPS led to an organ- and time-specific pattern of up-regulation of several MMP genes and the TIMP-1 gene in the liver, spleen, and kidney, whereas in the brain only TIMP-1 was induced. Injection of a lethal dose of LPS caused similar but more prolonged expression of these MMP genes as well as the induction of additional MMP genes in all organs. In LPS-treated mice in situ hybridization revealed collagenase 3 gene induction in cells resembling macrophages whereas TIMP-1 RNA was detected predominantly in parenchymal cells. Finally, gelatin zymography revealed increased gelatinolytic activity in all organs after LPS treatment. These observations highlight a dramatic shift in favor of increased expression of the MMP genes over the TIMP genes during LPS-induced endotoxemia, and suggest that MMPs may contribute to the development of organ damage in endotoxemia.

AIDS does not alter the total number of neurons in the hippocampal formation but induces cell atrophy: a stereological study.

Although cognitive dysfunction is a common finding in patients with acquired immunodeficiency syndrome (AIDS) its pathogenesis remains controversial. Given the involvement of the hippocampal formation in the processing of cognitive information and the scarcity of quantitative studies in this brain region, we have examined, using stereological methods, the hippocampal formations of AIDS patients. The study was performed in ten AIDS patients and ten age-matched controls. All cases were male. The Principle of Cavalieri was applied to estimate the volume of the layers of the dentate gyrus and of the CA3 and CA1 hippocampal fields. The fractionator and the nucleator were used as estimators of the total number, and mean somatic and nuclear volumes of the neurons in the cell-containing layers of all hippocampal subdivisions. No cell death was detected in AIDS patients but the global volume of their hippocampal formations was significantly decreased due to the reduced volume of its layers, mainly the cell-containing layers. Furthermore, the somatic and nuclear volumes of the neurons in the hippocampal formation were significantly decreased in AIDS patients. No correlation was found between the estimates obtained and the presence or absence of neurological involvement. Our results show that neurons in the hippocampal formation of AIDS patients display marked morphological changes, despite the maintenance of their total number. These alterations are likely to lead to dysfunction of the hippocampal circuitries and, thus, might contribute to explaining the dementia features which occur in this condition.

Cytochrome P450 catalyzed nitric oxide synthesis: a theoretical study.

Similar to nitric oxide synthase (NOS) cytochrome P450 isoforms (e.g. 3A and 4E) can produce nitric oxide from arginine. Although the active site of both proteins contains a protoporphyrin IX unit having an axial cystein ligand, their effectiveness in the synthesis of NO differs significantly. Now the molecular basis of this functional difference was investigated. A homology model for cytochrome P450 3A4 was refined and compared to the X-ray structure of iNOS. We found the active site of iNOS to be more readily accessible for the substrate than that of P450. Docking calculations were performed using the Monte Carlo conformational analysis technique on all internal and external degrees of freedom of arginine and active site residues as well. The lowest energy conformation of the cytochrome P450 3A4-substrate complex was compared to the high resolution X-ray structure of the iNOS-arginine complex. Comparison of substrate orientations revealed that arginine binds in a similar conformation in both enzymes. In contrast to iNOS we found, however, that in P450 partially negative propionate side chains of protoporphyrin IX are located on the opposite side of the heme plane. As a result of this and the absence of other negatively charged residues the distal (substrate binding) side of P450 should be less negative than that of NOS and therefore its affinity toward the partially positive arginine is reduced. Comparison of molecular electrostatic potentials calculated within the active site of the proteins supports this proposal. Reduced affinity in combination with limited substrate access might be responsible for the less effective NO synthesis of cytochrome P450 observed experimentally.