Complex regional pain syndrome type I affects brain structure in prefrontal and motor cortex.

The complex regional pain syndrome (CRPS) is a rare but debilitating pain disorder that mostly occurs after injuries to the upper limb. A number of studies indicated altered brain function in CRPS, whereas possible influences on brain structure remain poorly investigated. We acquired structural magnetic resonance imaging data from CRPS type I patients and applied voxel-by-voxel statistics to compare white and gray matter brain segments of CRPS patients with matched controls. Patients and controls were statistically compared in two different ways: First, we applied a 2-sample ttest to compare whole brain white and gray matter structure between patients and controls. Second, we aimed to assess structural alterations specifically of the primary somatosensory (S1) and motor cortex (M1) contralateral to the CRPS affected side. To this end, MRI scans of patients with left-sided CRPS (and matched controls) were horizontally flipped before preprocessing and region-of-interest-based group comparison. The unpaired ttest of the "non-flipped" data revealed that CRPS patients presented increased gray matter density in the dorsomedial prefrontal cortex. The same test applied to the "flipped" data showed further increases in gray matter density, not in the S1, but in the M1 contralateral to the CRPS-affected limb which were inversely related to decreased white matter density of the internal capsule within the ipsilateral brain hemisphere. The gray-white matter interaction between motor cortex and internal capsule suggests compensatory mechanisms within the central motor system possibly due to motor dysfunction. Altered gray matter structure in dorsomedial prefrontal cortex may occur in response to emotional processes such as pain-related suffering or elevated analgesic top-down control.

Two-step chromatographic purification of human plasma alpha(1)-acid glycoprotein: its application to the purification of rare phenotype samples of the protein and their study by chromatography on immobilized metal chelate affinity adsorbent.

Alpha1-Acid glycoprotein (AAG) or orosomucoid was purified to homogeneity from human plasma by a separate two-step method using chromatography on immobilized Cibacron Blue F3G-A to cross-linked agarose and chromatography on hydroxyapatite. The conditions for the pre-purification of AAG by chromatography on immobilized Cibacron Blue F3G-A were first optimized using different buffer systems with different pH values. The overall yield of the combined techniques was 80% and ca. 12 mg of AAG were purified from an initial total amount of ca. 15 mg in a ca. 40 ml sample of human plasma. This method was applied to the purification of AAG samples corresponding to the three main phenotypes of the protein (FI*S/A, F1/A and S/A), from individual human plasma previously phenotyped for AAG. A study by isoelectric focusing with carrier ampholytes showed that the microheterogeneity of the purified F1*S/A, F1/A and S/A AAG samples was similar to that of AAG in the corresponding plasma, thus suggesting that no apparent desialylation of the glycoprotein occurred during the purification steps. This method was also applied to the purification of AAG samples corresponding to rare phenotypes of the protein (F1/A*AD, S/A*X0 and F1/A*C1) and the interactions of these variants with immobilized copper(II) ions were then studied at pH 7, by chromatography on an iminodiacetate Sepharose-Cu(II) gel. It was found that the different variants encoded by the first of the two genes coding for AAG in humans (i.e. the F1 and S variants) interacted non-specifically with the immobilized ligand, whereas those encoded by the second gene of AAG (i.e. the A, AD, X0 and C1 variants) strongly bound to immobilized Cu(II) ions. These results suggested that chromatography on an immobilized affinity Cu(II) adsorbent could be helpful to distinguish between the respective products of the two highly polymorphic genes which code for human AAG.

Protein-protein interactions between adenovirus DNA polymerase and nuclear factor I mediate formation of the DNA replication preinitiation complex.

The in vitro adenovirus (Ad) DNA replication system provides an assay to study the interaction of viral and host replication proteins with the DNA template in the formation of the preinitiation complex. This initiation system requires in addition to the origin DNA sequences 1) Ad DNA polymerase (Pol), 2) Ad preterminal protein (pTP), the covalent acceptor for protein-primed DNA replication, and 3) nuclear factor I (NFI), a host cell protein identical to the CCAAT box-binding transcription factor. The interactions of these proteins were studied by coimmunoprecipitation and Ad origin DNA binding assays. The Ad Pol can bind to origin sequences only in the presence of another protein which can be either pTP or NFI. While NFI alone can bind to its origin recognition sequence, pTP does not specifically recognize DNA unless Ad Pol is present. Thus, protein-protein interactions are necessary for the targetting of either Ad Pol or pTP to the preinitiation complex. DNA footprinting demonstrated that the Ad DNA site recognized by the pTP.Pol complex was within the first 18 bases at the end of the template which constitutes the minimal origin of replication. Mutagenesis studies have defined the Ad Pol interaction site on NFI between amino acids 68-150, which overlaps the DNA binding and replication activation domain of this factor. A putative zinc finger on the Ad Pol has been mutated to a product that fails to bind the Ad origin sequences but still interacts with pTP. These results indicate that both protein-protein and protein-DNA interactions mediate specific recognition of the replication origin by Ad DNA polymerase.

Evolution of the Pelagonian carbonate platform complex and the adjacent oceanic realm in response to plate tectonic forcing (Late Triassic and Jurassic), Evvoia, Greece

The Late Triassic and Jurassic platform and the oceanic complexes in Evvoia, Greece, share a complementary plate-tectonic evolution. Shallow marine carbonate deposition responded to changing rates of subsidence and uplift, whilst the adjacent ocean underwent spreading, and then convergence, collision and finally obduction over the platform complex. Late Triassic ocean spreading correlated with platform subsidence and the formation of a long-persisting peritidal passive-margin platform. Incipient drowning occurred from the Sinemurian to the late Middle Jurassic. This subsidence correlated with intra-oceanic subduction and plate convergence that led to supra-subduction calc-alkaline magmatism and the formation of a primitive volcanic arc. During the Middle Jurassic, plate collision caused arc uplift above the carbonate compensation depth (CCD) in the oceanic realm, and related thrust-faulting, on the platform, led to sub-aerial exposures. Patch-reefs developed there during the Late Oxfordian to Kimmeridgian. Advanced oceanic nappe-loading caused platform drowning below the CCD during the Tithonian, which is documented by intercalations of reefal turbidites with non-carbonate radiolarites. Radiolarites and bypass-turbidites, consisting of siliciclastic greywacke, terminate the platform succession beneath the emplaced oceanic nappe during late Tithonian to Valanginian time.

Geochemical and H-O-Sr-Nd isotope evidence for magmatic processes and meteoric-water interactions in the basal complex of La Gomera, Canary Islands

The plutonic rocks of the Basal Complex of La Gomera, Canary Islands, Spain, were studied by means of major and trace element contents and by H-O-Sr-Nd isotope compositions in order to distinguish primary magmatic characteristics and late-stage alteration products. Deciphering the effects of alteration allowed us to determine primary, plume-related compositions that indicated D- and (18)O-depletion relative to normal upper mantle, supporting the conclusions of earlier studies on the plutonic rocks of Fuerteventura and La Palma. Late-stage alteration took place during the formation of the intrusive series induced by interaction with meteoric water. Inferred isotopic compositions of the meteoric water indicate that the water infiltrated into the rock edifice at a height of about 1500 m above sea level, suggesting the existence of a subaerial volcano which was active during the intrusive activity and that it has been either distroyed or remain buried by later volcanic and landslide events.

A recombinant rubella virus E1 glycoprotein as a rubella vaccine candidate.

A recombinant rubella virus E1 (rE1) glycoprotein was produced and some of its chemical and immunological features were characterized. Two animal models were then used to establish that the rE1 glycoprotein and rubella virus particles shared antigenic and immunogenic properties. In the first one, sera from rE1 glycoprotein-immunized BALB/c mice neutralized in vitro rubella virus infection. In the second model, severe combined immune deficient (SCID) mice implanted with tonsil fragments from rubella immune donors and immunized with rE1 glycoprotein produced human anti-rubella virus antibodies. Altogether, these results showed that immunization with rE1 glycoprotein elicited neutralizing anti-rubella virus antibodies. This study thus indicated that the rE1 glycoprotein could constitute a non-replicating rubella vaccine.

Role of major histocompatibility complex class II expression by non-hematopoietic cells in autoimmune and inflammatory disorders: facts and fiction.

It is well established that interactions between CD4(+) T cells and major histocompatibility complex class II (MHCII) positive antigen-presenting cells (APCs) of hematopoietic origin play key roles in both the maintenance of tolerance and the initiation and development of autoimmune and inflammatory disorders. In sharp contrast, despite nearly three decades of intensive research, the functional relevance of MHCII expression by non-hematopoietic tissue-resident cells has remained obscure. The widespread assumption that MHCII expression by non-hematopoietic APCs has an impact on autoimmune and inflammatory diseases has in most instances neither been confirmed nor excluded by indisputable in vivo data. Here we review and put into perspective conflicting in vitro and in vivo results on the putative impact of MHCII expression by non-hematopoietic APCs-in both target organs and secondary lymphoid tissues-on the initiation and development of representative autoimmune and inflammatory disorders. Emphasis will be placed on the lacunar status of our knowledge in this field. We also discuss new mouse models-developed on the basis of our understanding of the molecular mechanisms that regulate MHCII expression-that constitute valuable tools for filling the severe gaps in our knowledge on the functions of non-hematopoietic APCs in inflammatory conditions.

Munc18-1 mutations that strongly impair SNARE-complex binding support normal synaptic transmission.

Synaptic transmission depends critically on the Sec1p/Munc18 protein Munc18-1, but it is unclear whether Munc18-1 primarily operates as a integral part of the fusion machinery or has a more upstream role in fusion complex assembly. Here, we show that point mutations in Munc18-1 that interfere with binding to the free Syntaxin1a N-terminus and strongly impair binding to assembled SNARE complexes all support normal docking, priming and fusion of synaptic vesicles, and normal synaptic plasticity in munc18-1 null mutant neurons. These data support a prevailing role of Munc18-1 before/during SNARE-complex assembly, while its continued association to assembled SNARE complexes is dispensable for synaptic transmission.

Induction of CTL in vivo by major histocompatibility complex class I-peptide complexes covalently associated on the cell surface.

The identification of endogenously produced antigenic peptides presented by MHC class I molecules has opened the way to peptide-based strategies for CTL induction in vivo. Here we demonstrate that the induction in vivo of CTL directed against naturally processed antigens can be triggered by injection of syngeneic cells expressing covalent major histocompatibility complex class I-peptide complexes. In the model system used, the induction of HLA-Cw3 specific cytotoxic T lymphocytes (CTL) in mice by cell surface-associated, covalent H-2Kd (Kd)-Cw3 peptide complexes was investigated. The Kd-restricted Cw3 peptide 170-179 (RYLKNGKETL), which mimics the major natural epitope recognized by Cw3-specific CTL in H-2d mice, was converted to a photoreactive derivative by replacing Arg-170 with N-beta-(4-azidosalicyloyl)-L-2,3-diaminopropionic acid. This peptide derivative was equivalent to the parental Cw3 peptide in terms of binding to Kd molecules and recognition by Cw3-specific CTL clones and could be cross-linked efficiently and selectively to Kd molecules on the surface of Con A-stimulated spleen cells from H-2d mice. Photocross-linking prevented the rapid dissociation of Kd-peptide derivative complexes that takes place under physiological conditions. Cultures of spleen cells or peritoneal exudate cells from mice inoculated i.p. with peptide-pulsed and photocross-linked cells developed a strong CTL response following antigenic stimulation in vitro. The cultured cells efficiently lysed not only target cells sensitized with the Cw3 170-179 peptide but also target cells transfected with the Cw3 gene. Moreover, their TCR preferentially expressed V beta 10 and J alpha pHDS58 segments as well as conserved junctional sequences, as has been observed previously in Cw3-specific CTL responses. In contrast, no Cw3-specific CTL response could be obtained in cultures derived from mice injected with Con A-stimulated spleen cells pulsed with the peptide derivative without photocross-linking.

Regions of mouse mammary tumor virus superantigen involved in interaction with the major histocompatibility complex class II I-A molecule.

To study the major histocompatibility complex class II I-E dependence of mouse mammary tumor virus (MMTV) superantigens, we constructed hybrids between the I-E-dependent MMTV(GR) and the I-E-independent mtv-7 superantigens and tested them in vivo. Our results suggest that, although the C-terminal third mediates I-A interaction, additional binding sites are located elsewhere in the superantigen.

Selectivity in the binding of psychotropic drugs to the variants of alpha-1 acid glycoprotein.

The S- and F-forms of alpha-1 acid glycoprotein (AAG) variants have been isolated by isoelectric focusing with immobilines from commercially available AAG. In equilibrium dialysis experiments using a multicompartmental system, a higher affinity for various basic drugs has been found with S- in comparison with F-AAG: Amitriptyline, nortriptyline, imipramine, desipramine, trimipramine, methadone, thioridazine, clomipramine, desmethylclomipramine, and maprotiline. The selectivity (binding to S- vs. F-AAG) is the most pronounced for methadone and the lowest for thioridazine, while it is absent for the acidic drug mephenytoin.

Metal concentrations in soils around the copper smelter and surrounding industrial complex of Port Kembla, NSW, Australia

Anthropogenic emissions of metals from sources such as smelters are an international problem, but there is limited published information on emissions from Australian smelters. The objective of this study was to investigate the regional distribution of heavy metals in soils in the vicinity of the industrial complex of Port Kembla, NSW, Australia, which comprises a copper smelter, steelworks and associated industries. Soil samples (n=25) were collected at the depths of 0-5 and 5-20 cm, air dried and sieved to < 2 mm. Aqua regia extractable amounts of As, Cr, Cu, Ph and Zn were analysed by inductively coupled plasma mass spectrometry (lCP-MS) and inductively coupled plasma atomic emission spectrometry (ICP-AES). Outliers were identified from background levels by statistical methods. Mean background levels at a depth of 0-5 cm were estimated at 3.2 mg/kg As, 12 mg/kg Cr, 49 mg/kg Cu, 20 mg/kg Ph and 42 mg/kg Zn. Outliers for elevated As and Cu values were mainly present within 4 km from the Port Kembla industrial complex, but high Ph at two sites and high Zn concentrations were found at six sites up to 23 km from Port Kembla. Chromium concentrations were not anomalous close to the industrial complex. There was no significant difference of metal concentrations at depths of 0-5 and 5-20 cm, except for Ph and Zn. Copper and As concentrations in the soils are probably related to the concentrations in the parent rock. From this investigation, the extent of the contamination emanating from the Port Kembla industrial complex is limited to 1-13 km, but most likely <4 km, depending on the element; the contamination at the greater distance may not originate from the industrial complex. (C) 2003 Elsevier B.V. All rights reserved.