The Japanese LupusPRO: A cross-cultural validation of an outcome measure for lupus.

Objective This study aimed to validate the Japanese version of the LupusPRO questionnaire for use with systemic lupus erythematosus patients. Methods Participants were 205 lupus patients recruited from three rheumatology centers in Japan. Demographic data were collected and quality of life was assessed using the LupusPRO and the Short Form Health Survey-12. Disease activity was evaluated by physicians using the Systemic Lupus Erythematosus Activity Index. Some participants completed questionnaires 10-14 days after the first survey. Internal consistency reliability, test-retest reliability, content validity and convergent validity were examined, and confirmatory factor analysis was performed. Results Participants' mean age was 47.8 ± 13.6 years. Older participants scored lower on physical quality of life and higher on coping than younger participants. The LupusPRO showed satisfactory test-retest reliability ( n = 111). Test-retest reliability was lower for the mental and social aspects of quality of life, indicating fluctuations in quality of life during the two-week interval. Internal consistency reliability was good and convergent validity with the corresponding domains of the Short Form Health Survey-12 was satisfactory. Confirmatory factor analysis showed a good model fit. Conclusion The Japanese LupusPRO is a reliable and valid measure to evaluate treatment interventions for systemic lupus erythematosus.

TNF-α modulates expression of the circadian clock gene Per2 in rheumatoid synovial cells.

OBJECTIVES: To study the effect of tumour necrosis factor (TNF)-α, responsible for the inflammation and circadian rhythm of rheumatoid arthritis (RA), on the expression of circadian clock genes in primary cultured human rheumatoid synovial cells. METHOD: The expression of circadian clock genes, including circadian locomotor output cycles kaput (Clock), brain and muscle Arnt-like protein-1 (Bmal1), period (Per)1/2, and cryptochrome (Cry)1/2, and the proline and acidic amino acid-rich basic leucine zipper (PAR bZip) genes, a transcriptional activator of Per2, including D site of albumin promoter binding protein (Dbp), hepatic leukaemia factor (Hlf), and thyrotroph embryonic factor (Tef), and a transcriptional repressor of Per2, E4-binding protein 4 (E4bp4), in TNF-α-stimulated synovial cells was determined by real-time polymerase chain reaction (PCR). The D-box motifs in the Per2 promoter were mutated by site-directed mutagenesis, and the promoter activity of the Per2 gene was examined using the luciferase assay. RESULTS: TNF-α enhanced the mRNA expression of Bmal1 and Cry1 but did not affect that of Clock, Per1, or Cry2. However, TNF-α inhibited the mRNA expression of the Per2 gene, as well as Dbp, Hlf, and Tef, but enhanced the mRNA expression of E4bp4. Furthermore, TNF-α inhibited the transcriptional activity of the wild-type Per2 gene in a manner dependent on the D-box 1 and D-box 2 motifs in the Per2 promoter. CONCLUSIONS: TNF-α modulates the expression of the Per2 gene through the D-box binding proteins DBP, HLF, TEF, and E4BP4, in rheumatoid synovial cells, and thereby may contribute to the pathogenesis of RA.

Fatal BK virus pneumonia following stem cell transplantation.

We report the case of a 39-year-old male patient who died of severe BK virus (BKV) pneumonia 168 days after hematopoietic stem cell transplantation (HSCT) for acute lymphoblastic leukemia. After suffering from BKV-associated late-onset hemorrhagic cystitis (HC) with long-term sustained BKV viremia, he died of rapidly progressive pneumonia. On autopsy, numerous viral intranuclear inclusions were seen in his lungs and bladder. An immunohistochemical examination of his lungs was positive for simian virus 40. Based on these pathological results and the high sustained BKV viral load in his blood, we reached a diagnosis of BKV pneumonia. Viral infection can occasionally become life threatening among HSCT recipients. It is widely known that BKV can cause late-onset HC, but BKV-associated pneumonia is rare. Because of its rapid progression and poor prognosis, it is difficult to make an antemortem diagnosis of BKV pneumonia. A treatment strategy for BKV pneumonia also needs to be formulated. Similar to other viral pathogens, BKV can cause pneumonia and the clinician should therefore be aware of it in immunocompromised patients.

Neuroendocrine ductal carcinoma in situ of the breast: cytological features in 32 cases.

OBJECTIVE: The purpose of this study was to clarify the cytological features of neuroendocrine ductal carcinoma in situ (NE-DCIS) of the breast. METHODS: We analysed the cytopathological findings in 22 fine needle aspiration (FNA) smears and 17 nipple discharge smears obtained from 32 Japanese patients with NE-DCIS. RESULTS: The background of the FNA smears was clear (59%), mucoid (23%), haemorrhagic (14%) or necrotic (5%). Most of the FNA smears (95%) showed high cellularity. Characteristically, NE-DCIS cells were loosely arranged in three-dimensional solid clusters or singly dispersed. Well-developed vascular cores with or without malignant cells were occasionally recognized. The tumour cells were polygonal or spindle-shaped with a fine granular, abundant cytoplasm. Nuclei with finely granular chromatin were round or oval and often eccentrically located (plasmacytoid appearance). Mitotic figures were infrequent. Nuclear grade was estimated to be low in 86%. Most nipple discharge smears had fairly low cellularity with poorly preserved cell clusters in a markedly haemorrhagic background, although two (12%) were extremely cellular with cytological characteristics similar to those of the FNA smears. Pre-operative cytological malignant diagnoses were made in 42% of FNA smears and 0% of nipple discharge smears. Immunohistochemistry for neuroendocrine markers (chromogranin A and synaptophysin) confirmed the neuroendocrine nature of this tumour in adequate cytological specimens. CONCLUSIONS: NE-DCIS has distinctive cytological features and can therefore be diagnosed as a neuroendocrine tumour in most FNAs and some nipple discharge smears by cytological examination employing immunohistochemical techniques. We emphasize that a breast lesion with these features may be in situ and not invasive, and also that there is a risk of under-diagnosis.

Simultaneous Fluorescent Identification of Odontoblasts and Ameloblasts.

The tooth is mainly composed of dentin and enamel. Identification of dentin-producing odontoblasts and enamel-producing ameloblasts using reporter techniques is useful to study tooth development and regeneration with tissue engineering. Ameloblasts express Amelogenin, Ameloblastin, Enamelin, and Amelotin, whereas odontoblasts express Dentin sialophosphoprotein (Dspp) and Dentin matrix protein1 (Dmp1). Although there are several transgenic lines using promoter elements or bacterial artificial chromosomes (BACs) to label odontoblasts and ameloblasts, there is a possibility that the expression patterns vary from the endogenous genes. Here, we established 2 lines of mice where tdTomato was knocked into the second exon of X-chromosomal Amelogenin (Amelx), and green fluorescent protein (GFP) was knocked into the second exon of Dspp. tdTomato and GFP were highly expressed on secretory ameloblasts and secretory and fully differentiated odontoblasts, respectively. In addition, DSPP and AMELX were not produced in the dentin matrix and enamel matrix of DsppGFP/GFP and AmelxtdTomato male mice (as representative of AmelxtdTomato/Y hemizygous male mice), respectively. Moreover, micro-computed tomography analysis of AmelxtdTomato male mice revealed a notable reduction in enamel volume but increased dentin mineral density. DsppGFP/GFP mice had reduced dentin mineral density. To identify odontoblasts and ameloblasts from developing tooth, we examined the expression of mesenchymal cell surface molecules CD90, CD166 and epithelial cell surface molecules CD49f, Epcam1 with fluorescence on odontoblasts and ameloblasts in these mice. We found that GFP+ odontoblasts and tdTomato+ ameloblasts in tooth germ from 0.5-d-old DsppGFP/+ mice and AmelxtdTomato male mice were enriched in CD45-/Ter119-/Epcam1-/CD90+/Integrin α4+cell fractions and CD45-/Ter119-/Epcam1+/CD49f+/CD147+ cell fractions, respectively. By using antibodies against mesenchymal and epithelial cell surface molecules and fluorescence, we can easily distinguish odontoblasts from ameloblasts and isolate each cell for further studies. These mice would serve as useful models for tooth development and regeneration as well as provide concurrent observation for the differentiation processes of odontoblasts and ameloblasts in vivo and in vitro.

Long-term efficacy of Helicobacter pylori eradication in patients with idiopathic thrombocytopenic purpura: 7-year follow-up prospective study.

Helicobacter pylori eradication is useful for improvement of a half of patients with idiopathic thrombocytopenic purpura (ITP), but its long-term therapeutic efficacy has not been elucidated. We investigated the long-term efficacy of H. pylori eradication in 30 cases with ITP that were included in our previous study regarding the association between H. pylori infection and ITP. Twenty-one cases were positive and nine cases were negative for H. pylori infection. H. pylori eradication therapy including secondary regimen was successful in 20 cases, half (responder) of whom showed ITP remission 1 month later. Nine responders could be followed up for a long time and did not show re-infection of H. pylori. Eight of nine needed no medication except for eradication therapy. Another case remained in remission for 1 year but thereafter needed a steroid therapy due to the recurrence. Eight nonresponders could be followed up for a long time. All these cases showed a bad clinical course even though they received the other post-treatments including steroid therapy. Three of nine H. pylori-negative cases underwent eradication therapy after obtaining the written informed consent, but none of them showed improvement. Of these three cases, two cases could be followed up. Only one case remained a remission although receiving corticosteroid as a post-treatment. Conditions of H. pylori-negative ITP cases were usually unstable for a long time. H. pylori eradication has a short-term efficacy for about half of H. pylori-positive ITP patients, and the responders to the eradication therapy may receive a long-term clinical benefit without other therapies.

Zinc protoporphyrin in the erythrocytes of patients with lead intoxication and iron deficiency anemia.

The fluorescent porphyrin in the erythrocytes of patients with lead intoxication or with iron deficiency anemia is zinc protoporphyrin that is bound to globin moieties, probably at heme binding sites.

Proton magnetic resonance of transfer RNA.

The temperature dependence of the areas under the proton magnetic resonance spectra of unfractionated yeast transfer RNA in 1.0 molar NaCl is a consequence of salt-induced aggregation and does not constitute a monitor of the melting of secondary molecular structure. Such melting can be observed by following the widths of the resonances in the various regions of the spectra. Peaks attributable to dihydrouracil and the methyl groups of the methylated bases are detected in the spectra of unfractionated transfer RNA and alanine transfer RNA.

Nucleotide sequence of the "denaturable" leucine transfer RNA from yeast.

The nucleotide sequence of " denaturable"leucine acceptor transfer RNA (tRNA(Leu)(3)) from baker's yeast was determined on (32)P-labeled material. The molecule is 85 nucleotides long and can be folded into the "cloverleaf" model for secondary structure. The basis on which the sequence was deduced from the products of complete enzymatic digestion, prior to its unambiguous determination, is presented.

Cholesterol hydroperoxide formation in red cell membranes and photohemolysis in erythropoietic protoporphyria.

3beta-Hydroxy-5alpha-hydroperoxy-Delta(6)-cholestene is produced in protoporphyrin-containing red blood cell ghosts irradiated with approximately 400-nanometer light in the presence of oxygen. Incorporation of this cholesterol photooxidation product into normal red blood cells leads to increased osmotic fragility and eventual hemolysis. These results may be relevant to photohemolysis associated with erythropoietic protoporphyria.

Large cryptic internal sequence repeats in protein structures from Homo sapiens.

Amino acid sequences are known to constantly mutate and diverge unless there is a limiting condition that makes such a change deleterious. However, closer examination of the sequence and structure reveals that a few large, cryptic repeats are nevertheless sequentially conserved. This leads to the question of why only certain repeats are conserved at the sequence level. It would be interesting to find out if these sequences maintain their conservation at the three-dimensional structure level. They can play an active role in protein and nucleotide stability, thus not only ensuring proper functioning but also potentiating malfunction and disease. Therefore,insights into any aspect of the repeats - be it structure, function or evolution - would prove to be of some importance. This study aims to address the relationship between protein sequence and its three-dimensional structure, by examining if large cryptic sequence repeats have the same structure.

Prognostic value of serum surfactant protein D level prior to transplant for the development of bronchiolitis obliterans syndrome and idiopathic pneumonia syndrome following allogeneic hematopoietic stem cell transplantation.

Bronchiolitis obliterans syndrome (BOS) and idiopathic pneumonia syndrome (IPS) cause high mortality and impaired survival after allogeneic hematopoietic stem-cell transplantation (allo-HSCT). Early recognition of patients at high risk of developing BOS/IPS may lead to improving the outcome of allo-HSCT. We retrospectively analyzed serum surfactant protein A, D (SP-A, -D) and Kerbs von Lungren 6 Ag (KL-6) levels before allo-HSCT in 56 patients who survived more than 90 days after allo-HSCT and compared values of these serum markers and other transplant factors in BOS/IPS patients with those in non-BOS/IPS patients. Five patients developed BOS and two developed IPS at a median interval of 303 and 117 days (range, 100-452 and 95-153) from transplantation. As a result of univariate analysis, pretransplant serum SP-D levels but not SP-A, KL-6 in BOS/IPS patients were significantly lower than those in non-BOS/IPS patients (P=0.03). In multivariate analysis, the patients with lower pretransplant serum SP-D level had a trend toward frequent development of BOS/IPS (P=0.08). Constitutive serum SP-D level before allo-HSCT may be a useful, noninvasive predictor for the development of BOS/IPS.

Absolute calibration of imaging plate for GeV electrons.

An imaging plate has been used as a useful detector of energetic electrons in laser electron acceleration and laser fusion studies. The absolute sensitivity of an imaging plate was calibrated at 1 GeV electron energy using the injector Linac of SPring-8. The sensitivity curve obtained up to 100 MeV in a previous study was extended successfully to GeV range.

Erythropoietic protoporphyria and lead intoxication: the molecular basis for difference in cutaneous photosensitivity. II. Different binding of erythrocyte protoporphyrin to hemoglobin.

Acidic solvents extract the same porphyrin-protoporphyrin-from the erythrocytes of patients with either erythropoietic protoporphyria or lead intoxication. However, extractable protoporphyrin disappears rapidly, both in vivo and in vitro, from erythrocytes in erythropoietic protoporphyria but slowly, if at all, in lead intoxication. Consistent with these observations, fluorescence spectroscopy revealed that the intracellular state of the erythrocyte protoporphyrin is different in the two diseases. Spectrofluorometric measurements coupled with fractionations and biochemical syntheses showed that in erythropoietic protoporphyria the protoporphyrin is bound as the free base to hemoglobin molecules at sites other than the heme binding sites. In lead intoxication the fluorescent porphyrin is also bound to hemoglobin but is present as zinc protoporphyrin. The data suggest that the zinc protoporphyrin is bound at heme binding sites. Acidic extraction solvents remove the chelated zinc, but zinc protoporphyrin may be extracted intact from erythrocytes with acetone, ethanol, or the detergent Ammonyx-LO.

Predictors of poor sleep quality in patients with systemic lupus erythematosus.

Sleep problems are common in patients with systemic lupus erythematosus (SLE). This study aimed to examine the following: (1) predictors of sleep quality and (2) fluctuations in sleep quality in patients with SLE. Patients with SLE were recruited from three rheumatology centers in Japan. We collected demographic and clinical data and data on sleep quality as measured by the Pittsburgh Sleep Quality Index (PSQI), the Medical Outcome Study Short Form-12, and the Lupus Patient Reported Outcome Tool (LupusPRO). Fluctuations in sleep quality were examined by administering the PSQI a second time after a 2-week interval. We used multiple linear regression analysis to predict sleep quality. Of 205 patients who completed the survey, 62.9% showed poor sleep quality. The largest fluctuation in sleep quality was for "waking in the middle of the night or early morning." "LupusPRO pain/vitality" was a major predictor of poor sleep. The other significant predictors were mostly LupusPRO subscales and clinical variables and SF-12 subscales were mostly non-predictive. The majority of the participants had poor sleep quality. A lupus-specific QoL scale is important for understanding poor sleep quality in SLE patients. Symptom management appeared to play a key role in improving sleep quality.

Activation of human neutrophil by cytokine-activated endothelial cells.

Cytokine activation of vascular endothelial cells renders the hyperadhesiveness for neutrophils. During the processes of inflammation and atherosclerosis, the production of reactive oxygen species by neutrophils contributes to endothelial cell (EC) damage and injury. However, the precise mechanisms for neutrophil activation by ECs remain unknown. Thus, we investigated what kinds of pathophysiological factors synthesized by inflammatory cytokine-activated ECs potentiated the activity of neutrophil functions. The magnitude of O(2)(-) release from neutrophils, which is one of pivotal neutrophil functions, was measured as an indicator potentiated by activated ECs. Neutrophils release massive amounts of O(2)(-) on coculture with activated ECs. Anti-granulocyte-macrophage colony-stimulating factor (GM-CSF) antibody (Ab) or specific platelet-activating factor (PAF)-receptor antagonist suppressed the O(2)(-) release from neutrophils on coculture with the activated ECs by 50% to 70%. The supernatants from activated ECs also induced O(2)(-) release by neutrophils. This stimulatory effect of activated EC supernatants on O(2)(-) release by neutrophils was abolished by anti-GM-CSF Ab or by PAF-receptor antagonist. As we previously reported, we demonstrated the expression of GM-CSF mRNA by Northern blotting and protein synthesis of GM-CSF by ELISA on tumor necrosis factor as well as interleukin-1-activated ECs. Although phosphorylation of mitogen-activated protein kinases was observed in ECs stimulated by tumor necrosis factor and interleukin-1, treatment of ECs with PD98059 (MEK1 inhibitor) and SB203580 (p38 mitogen-activated protein kinase inhibitor) in the presence of the cytokine failed to attenuate the stimulatory effect of activated ECs on neutrophil activation. We found that activated ECs regulated neutrophil function on coculture. We show here for the first time, to our knowledge, that the collaboration between GM-CSF and PAF synthesized by activated ECs markedly potentiated neutrophil activation.

High-resolution structure of the conger eel galectin, congerin I, in lactose-liganded and ligand-free forms: emergence of a new structure class by accelerated evolution.

BACKGROUND: Congerin I is a member of the galectin (animal beta-galactoside-binding lectin) family and is found in the skin mucus of conger eel. The galectin family proteins perform a variety of biological activities. Because of its histological localization and activity against marine bacteria and starfish embryos, congerin I is thought to take part in the eels' biological defense system against parasites. RESULTS: The crystal structure of congerin I has been determined in both lactose-liganded and ligand-free forms to 1. 5 A and 1.6 A resolution, respectively. The protein is a homodimer of 15 kDa subunits. Congerin I has a beta-sheet topology that is markedly different from those of known relatives. One of the beta-strands is exchanged between two identical subunits. This strand swap might increase the dimer stability. Of the known galectin complexes, congerin I forms the most extensive interaction with lactose molecules. Most of these interactions are substituted by similar interactions with water molecules, including a pi-electron hydrogen bond, in the ligand-free form. This observation indicates an increased affinity of congerin I for the ligand. CONCLUSIONS: The genes for congerin I and an isoform, congerin II, are known to have evolved under positive selection pressure. The strand swap and the modification in the carbohydrate-binding site might enhance the cross-linking activity, and should be the most apparent consequence of positive selection. The protein has been adapted to functioning in skin mucus that is in direct contact with surrounding environments by an enhancement in cross-linking activity. The structure of congerin I demonstrates the emergence of a new structure class by accelerated evolution under selection pressure.

Iterative ACORN as a high throughput tool in structural genomics.

High throughput macromolecular structure determination is very essential in structural genomics as the available number of sequence information far exceeds the number of available 3D structures. ACORN, a freely available resource in the CCP4 suite of programs is a comprehensive and efficient program for phasing in the determination of protein structures, when atomic resolution data are available. ACORN with the automatic model-building program ARP/wARP and refinement program REFMAC is a suitable combination for the high throughput structural genomics. ACORN can also be run with secondary structural elements like helices and sheets as inputs with high resolution data. In situations, where ACORN phasing is not sufficient for building the protein model, the fragments (incomplete model/dummy atoms) can again be used as a starting input. Iterative ACORN is proved to work efficiently in the subsequent model building stages in congerin (PDB-ID: lis3) and catalase (PDB-ID: 1gwe) for which models are available.

Hormonal control of alpha-fetoprotein secretion in human hepatoma cell lines proliferating in chemically defined medium.

The regulation of alpha-fetoprotein (AFP) secretion and growth rate by various hormones in established human hepatoma (HuH-7, PLC/PRF/5, huH-1, huH-4, and KIM-1/c-4) and hepatoblastoma (HUH-6 Clone 5) cell lines was studied. These 6 cell lines replicated continuously in a chemically defined medium and secreted 84 ng (HuH-7) to 23 pg (huH-4) AFP per 24 h per 1 X 10(4) cells into the culture medium. The addition of insulin increased the growth rate of all examined cell lines and partially inhibited the AFP secretion in those cell lines except KIM-1/c-4, while the addition of dexamethasone inhibited the growth and stimulated the AFP secretion in all of the cell lines. The addition of 3,3',5-triiodothyronine inhibited the growth of all cell lines; however, different effects on the AFP secretion were observed depending on the cell lines used. Obviously, the AFP secretion was unrelated to the change in growth rate. When dexamethasone and N6-O2-dibutyryl cyclic AMP were added together, the AFP secretion was further stimulated. On the other hand, when dexamethasone and insulin were added simultaneously, the dexamethasone-mediated stimulation of AFP secretions was diminished. The data indicated that the regulatory mechanisms of AFP secretion by the hormones in the established human hepatoma and hepatoblastoma cell lines cannot be deduced according to the results of one cell line.