Phthalocyanine-dyed fibers adsorb allergenic proteins.

Phthalocyanine (Pc)-dyed fiber is reported to reduce atopic symptoms in some patients when they use underwear made of the fiber. We investigated the adsorption of allergens on Pc-fiber. Pc-fiber trapped house dust/pollen/food allergens with varied molecular weight and pI. The adsorbed allergens were released in the presence of mild detergent. Pc-fiber did not change the molecular weight or disulfide bonding of the allergens. These observations imply that Pc-fiber is applicable as an "allergen trap" for a wide variety of products.

Clinical diagnosis of MM2-type sporadic Creutzfeldt-Jakob disease.

BACKGROUND: No method for the clinical diagnosis of MM2-type sporadic Creutzfeldt-Jakob disease (sCJD) has been established except for pathologic examination. OBJECTIVE: To identify a reliable marker for the clinical diagnosis of MM2-type sCJD. METHODS: CSF, EEG, and neuroimaging studies were performed in eight patients with MM2-type sCJD confirmed by neuropathologic, genetic, and western blot analyses. RESULTS: The eight cases were pathologically classified into the cortical (n = 2), thalamic (n = 5), and combined (corticothalamic) (n = 1) forms. The cortical form was characterized by late-onset, slowly progressive dementia, cortical hyperintensity signals on diffusion-weighted imaging (DWI) of brain, and elevated levels of CSF 14-3-3 protein. The thalamic form showed various neurologic manifestations including dementia, ataxia, and pyramidal and extrapyramidal signs with onset at various ages and relatively long disease duration. Characteristic EEG and MRI abnormalities were almost absent. However, all four patients examined with cerebral blood flow (CBF) study using SPECT showed reduction of the CBF in the thalamus as well as the cerebral cortex. The combined form had features of both the cortical and the thalamic forms, showing cortical hyperintensity signals on DWI and hypometabolism of the thalamus on [18F]2-fluoro-2-deoxy-d-glucose PET. CONCLUSION: For the clinical diagnosis of MM2-type sporadic Creutzfeldt-Jakob disease, cortical hyperintensity signals on diffusion-weighted MRI are useful for the cortical form and thalamic hypoperfusion or hypometabolism on cerebral blood flow SPECT or [18F]2-fluoro-2-deoxy-d-glucose PET for the thalamic form.

Design, synthesis and biological evaluation of a novel series of potent, orally active adenosine A1 receptor antagonists with high blood-brain barrier permeability.

A novel series of 3-(2-substituted-3-oxo-2,3-dihydropyridazin-6-yl)-2-phenylpyrazolo[1,5-a]pyridines (5-38) were synthesized and evaluated for their in vitro adenosine A1 and A(2A) receptor binding activities, and in vitro metabolism by rat liver in order to search for orally active compounds. Most of the test compounds were potent adenosine A1 receptor antagonists with high A1 selectivity and the A1 affinity and A1 selectivity of carbonyl derivatives (5-11) was particularly high. In particular, compound 7 was an extremely potent and selective adenosine A1 antagonist with high A1 selectivity (Ki=0.026 nM, A(2A)/A1=5400). In terms of metabolic stability, 2-oxopropyl (5), 2-hydroxypropyl (12), N-methylacetamide (16), 2-(piperidin-1-yl)ethyl (28) and 1-methylpiperidin-4-yl (32, FR194921) were the most stable compounds in this series of analogues. Further in vivo evaluation indicated that compounds 5, 13, 17, 28 and 32 were detected in both plasma and brain after oral administration in rats. In particular, 32 displayed good plasma and brain concentrations (dose: 32 mg/kg (n=3); after 30 min, plasma conc.=3390+/-651nM, brain conc.=3670+/-496nM; after 60min, plasma conc.=1580+/-348nM, brain conc.=2143+/-434nM), and a good brain/plasma ratio (1.11+/-0.060 (30min), 1.39+/-0.172 (60min)). As a result, we could show that 32 is a good candidate for an orally active adenosine A1 receptor antagonist with high blood-brain barrier permeability and good bioavailability (Ki=6.6nM, A(2A)/A1=820, BA=60.6+/-4.9% (32 mg/kg)).

Identification of CRAM, a novel unc-33 gene family protein that associates with CRMP3 and protein-tyrosine kinase(s) in the developing rat brain.

Four members of collapsin response mediator proteins (CRMPs) are thought to be involved in the semaphorin-induced growth cone collapse during neural development. Here we report the identification of a novel CRMP3-associated protein, designated CRAM for CRMP3-associated molecule, that belongs to the unc-33 gene family. The deduced amino acid sequence reveals that the CRAM gene encodes a protein of 563 amino acids, shows 57% identity with dihydropyrimidinase, and shows 50-51% identity with CRMPs. CRAM appears to form a large complex composed of CRMP3 and other unidentified proteins in vivo. Indeed, CRAM physically associates with CRMP3 when co-expressed in COS-7 cells. The expression of CRAM is brain-specific, is high in fetal and neonatal rat brain, and decreases to very low levels in adult brain. Moreover, CRAM expression is up-regulated during neuronal differentiation of embryonal carcinoma P19 and PC12 cells. Finally, immunoprecipitation analysis of rat brain extracts shows that CRAM is co-immunoprecipitated with proteins that contain protein-tyrosine kinase activity. Taken together, our results suggest that CRAM, which interacts with CRMP3 and protein-tyrosine kinase(s), is a new member of an emerging family of molecules that potentially mediate signals involved in the guidance and outgrowth of axons.

Thermal distribution of radio-frequency inductive hyperthermia using an inductive aperture-type applicator: evaluation of the effect of tumour size and depth.

A new radio-frequency (RF) inductive hyperthermia device using an inductive aperture-type applicator (IATA) is proposed. This paper reports the evaluation of the heating characteristics of the IATA using a computer simulation and clinical thermal parameters obtained during treatment of superficial and subsurface-seated tumours. The configuration of the IATA is a one-turn square column-like coil whose bottom plate is set to face the heating target. The IATA has advantages over RF capacitive-type heating, such as: generating less heat in the subcutaneous fat layer; less convergency of electric line of force at the edge of the applicator; and no physical contact with the target lesion. The induced magnetic fields and electrical currents within the heating substance are simulated using computer-assisted design software for electro-magnetic analysis. A total of 40 superficial and subsurface tumours are treated with the IATA. Invasive thermometry is performed continuously for 110 sessions using multi-sensor probes of an optical thermometer. Thermal parameters (Tmax, Tmin, Tave) are assessed based on the tumour size and depth. The treated tumours are categorised into three groups according to tumour depth: group 1 (< 3 cm, n = 28), group 2 (3-5 cm, n = 35) and group 3 (> 5 cm, n = 47). The computer simulation shows that induced electrical currents run without convergency, parallel to the surface of the heating material. All thermal parameters of group 3 are significantly higher than those of group 1 and 2 (p < 0.05), indicating that the larger lesions tend to abtain a higher temperature distribution. In conclusion, RF inductive hyperthermia using the IATA results in effective temperature distributions in superficial and subsurface tumours, with large tumours being most effectively heated.

Mammalian homologue of the Caenorhabditis elegans UNC-76 protein involved in axonal outgrowth is a protein kinase C zeta-interacting protein.

By the yeast two-hybrid screening of a rat brain cDNA library with the regulatory domain of protein kinase C zeta (PKCzeta) as a bait, we have cloned a gene coding for a novel PKCzeta-interacting protein homologous to the Caenorhabditis elegans UNC-76 protein involved in axonal outgrowth and fasciculation. The protein designated FEZ1 (fasciculation and elongation protein zeta-1) consisting of 393 amino acid residues shows a high Asp/Glu content and contains several regions predicted to form amphipathic helices. Northern blot analysis has revealed that FEZ1 mRNA is abundantly expressed in adult rat brain and throughout the developmental stages of mouse embryo. By the yeast two-hybrid assay with various deletion mutants of PKC, FEZ1 was shown to interact with the NH2-terminal variable region (V1) of PKCzeta and weakly with that of PKCepsilon. In the COS-7 cells coexpressing FEZ1 and PKCzeta, FEZ1 was present mainly in the plasma membrane, associating with PKCzeta and being phosphorylated. These results indicate that FEZ1 is a novel substrate of PKCzeta. When the constitutively active mutant of PKCzeta was used, FEZ1 was found in the cytoplasm of COS-7 cells. Upon treatment of the cells with a PKC inhibitor, staurosporin, FEZ1 was translocated from the cytoplasm to the plasma membrane, suggesting that the cytoplasmic translocation of FEZ1 is directly regulated by the PKCzeta activity. Although expression of FEZ1 alone had no effect on PC12 cells, coexpression of FEZ1 and constitutively active PKCzeta stimulated the neuronal differentiation of PC12 cells. Combined with the recent finding that a human FEZ1 protein is able to complement the function of UNC-76 necessary for normal axonal bundling and elongation within axon bundles in the nematode, these results suggest that FEZ1 plays a crucial role in the axon guidance machinery in mammals by interacting with PKCzeta.

Molecular cloning and characterization of RBCK2, a splicing variant of a RBCC family protein, RBCK1.

RBCK1 (RBCC protein interacting with PKC 1) has two coiled-coil regions, a RING finger, a B-box and a B-box-like motif. RBCK2, a cDNA fragment related to RBCK1 was obtained, that lacks the 161-bp sequence of RBCK1 and encodes 260 amino acid residues. The 240-amino acid sequence in the NH2-terminal of RBCK2 is identical with RBCK1 and contains two coiled-coil regions but no other structural motifs, whereas the 20-amino acid sequence in the COOH-terminal is distinct from RBCK1. The analysis of genomic DNA revealed that RBCK1 and RBCK2 are generated from a single gene by alternative splicing. The RBCK1 protein interacted with the RBCK1 and RBCK2 proteins, but the RBCK2 protein did not interact with itself, in vitro. The RBCK2 protein fused with the DNA-binding domain of yeast GAL4 (GAL4DBD) did not show a transcriptional activity, but the RBCK2 protein inhibited the transcriptional activity of the RBCK1 protein fused with GAL4DBD. These results suggest that RBCK2 may inhibit the transcriptional activity of RBCK1 probably through complex formation with RBCK1.

Molecular cloning and characterization of a novel protein kinase C-interacting protein with structural motifs related to RBCC family proteins.

A novel protein kinase C (PKC)-interacting protein was identified by the yeast two-hybrid screening using the regulatory domain of PKC beta I as a bait. The protein contained several structural motifs such as two putative coiled-coil regions, a RING-finger, a B-box, and a B-box-like motif in the order from NH2- to COOH-terminals. The molecular organization of the protein resembles the structure of the RBCC protein family proteins which usually have a RING-finger, a B-box, and a coiled-coil region. Therefore, the protein identified was designated as RBCK1 (RBCC protein interacting with PKC 1). Northern blot analysis showed that RBCK1 gene is expressed ubiquitously among rat tissues. RBCK1 protein associated with PKC beta I and PKC zeta when coexpressed in cultured mammalian cells. By the polymerase chain reaction-assisted DNA-binding site selection and the electrophoretic mobility shift assay, RBCK1 protein was shown to bind to several DNA fragments containing TGG-rich sequences. When the yeast GAL4 DNA-binding domain fused RBCK1 protein was expressed in COS-7 cells harboring the luciferase gene placed under a synthetic promoter containing GAL4-binding sites, the fusion protein showed enhanced transcriptional activity comparing with the GAL4 DNA-binding domain. These results suggest that RBCK1 protein might be a transcription factor that has a role in the signaling pathway through PKC.

Transient ipsilateral blood flow reduction in the thalamus and cerebral cortex after pontine infarction.

Remote cerebral hypoperfusion associated with pontine lesions is rare. We describe a patient who showed transient ipsilateral hypoperfusion in the thalamus and cerebral cortex after a pontine infarct. This resolved within 8 days after onset. Anatomical considerations strongly suggested involvement of the cerebellothalamocortical pathway in this case.

Molecular cloning and characterization of a new member of the RAC protein kinase family: association of the pleckstrin homology domain of three types of RAC protein kinase with protein kinase C subspecies and beta gamma subunits of G proteins.

cDNA clones encoding the third member of the RAC protein kinase family, termed RAC-PK gamma, were isolated from a rat brain cDNA library. The deduced amino acid sequence of RAC-PK gamma was highly related to those of previously identified family members, RAC-PK alpha and beta, that have a pleckstrin homology domain and a protein-serine/threonine kinase catalytic domain at the amino- and carboxyl-terminal regions, respectively. Northern blot analysis indicated that RAC-PK gamma was expressed abundantly in brain and testis. Specific activities of RAC-PK alpha, beta, and gamma purified from transfected COS-7 cells were similar when measured by using myelin basic protein as a phosphate acceptor. Analysis using fusion proteins of glutathione S-transferase revealed that the pleckstrin homology domain of the three subtypes of RAC-PK associate with both protein kinase C subspecies and beta gamma subunits of G proteins. These results suggest that the pleckstrin homology domains of RAC protein kinase family could associate more than one protein to regulate the activity and/or intracellular distribution of this enzyme family by different ways.

Molecular cloning of rat RAC protein kinase alpha and beta and their association with protein kinase C zeta.

cDNAs of RAC protein kinases (RAC-PK) alpha and beta were cloned from a rat testis cDNA library. The predicted open reading frames encode 480 and 481 amino acids of RAC-PK alpha and beta, respectively, and the rat RAC-PK alpha and beta have sequences conserved among different mammalian species such as the pleckstrin homology domain at their amino-terminal region and the protein-serine/threonine kinase catalytic domain at their carboxyl-terminal region. RNA blot analysis showed wide distribution of two RAC-PK in rat tissues. Immunoprecipitation analysis revealed that RAC-PK alpha and beta associate with protein kinase C zeta through the pleckstrin homology domain in vitro, suggesting the interaction between RAC-PK and protein kinase C.

Novel subtype of human angiotensin II type 1 receptor: analysis of signal transduction mechanism in transfected Chinese hamster ovary cells.

We examined the intracellular signal transduction of two subtypes of human angiotensin II type 1 receptor (AT1aR and AT1bR) by means of the stable expression of each receptor cDNA in Chinese hamster ovary cells. Both receptors showed a rapid stimulation of phosphatidylinositol hydrolysis, a transient increase of intracellular Ca2+ concentration and an inhibitory action on the forskolin-induced cyclic AMP formation. Interestingly, at high AII concentrations (> 1 microM), these AT1bR-mediated responses were inhibited, whereas the AT1aR-mediated responses were not. Thus, the two AT1Rs are considered to be coupled to the same signal transduction cascades, but to be regulated differently on the post-translational level (presumably desensitization).

[Korsakoff's syndrome as a prominent feature of bilateral paramedian thalamic infarction--a case report].

We reported the first Japanese case of bilateral paramedian thalamic infarction associated with prominent Korsakoff's syndrome. 53-year-old man suffered from semicoma on the morning of September 16th, 1988. After recovery of consciousness disturbance, neurological examination revealed vertical eye gaze palsy, areflexia of lower extremities, apathy with hypersomnia and amnesia. Amnesia was accompanied with prominent confabulation, disorientation and lack of insight into his own disability. While X ray-CT revealed only ambiguous low density area in the bilateral thalamus, MRI of horizontal section by short spin echo revealed symmetrical low signal area restricted in the paramedian area of bilateral thalamus, and that of coronal section revealed characteristic butterfly-shaped lesion. Left BAG revealed that both posterior thalamoperforating arteries showed type 3 variation of Percheron's classification which arisen from artery arcade bridging between both side of interpeduncular segment of posterior cerebral artery. He showed gradual improvement in apathy with hypersomnia and disorientation but not in Korsakoff's syndrome nor ophthalmoplegia.