Investigation of immune complexes formed by mitochondrial antigens containing a new lipoylated site in sera of primary biliary cholangitis patients.

Primary biliary cholangitis (PBC) is characterized by the presence of serum anti-mitochondrial autoantibodies (AMAs). To date, four antigens among the 2-oxo-acid dehydrogenase complex family, which commonly have lipoyl domains as an epitope, have been identified as AMA-corresponding antigens (AMA-antigens). It has recently been reported that AMAs react more strongly with certain chemically modified mimics than with the native lipoyl domains in AMA-antigens. Moreover, high concentrations of circulating immune complexes (ICs) in PBC patients have been reported. However, the existence of ICs formed by AMAs and their antigens has not been reported to date. We hypothesized that AMAs and their antigens formed ICs in PBC sera, and analyzed sera of PBC and four autoimmune diseases (Sjögren's syndrome, systemic lupus erythematosus, systemic scleroderma, and rheumatoid arthritis) using immune complexome analysis, in which ICs are separated from serum and are identified by nano-liquid chromatography-tandem mass spectrometry. To correctly assign MS/MS spectra to peptide sequences, we used a protein-search algorithm that including lipoylation and certain xenobiotic modifications. We found three AMA-antigens, the E2 subunit of the pyruvate dehydrogenase complex (PDC-E2), the E2 subunit of the 2-oxo-glutarate dehydrogenase complex (OGDC-E2) and dihydrolipoamide dehydrogenase binding protein (E3BP), by detecting peptides containing lipoylation and xenobiotic modifications from PBC sera. Although the lipoylated sites of these peptides were different from the well-known sites, abnormal lipoylation and xenobiotic modification may lead to production of AMAs and the formation ICs. Further investigation of the lipoylated sites, xenobiotic modifications, and IC formation will lead to deepen our understanding of PBC pathogenesis.

Proteomic profile of circulating immune complexes in chronic Chagas disease.

Immune complexes (ICs) are the direct and real-time products of humoral immune responses. The identification of constituent foreign or autoantigens within ICs might bring new insights into the pathology of infectious diseases. We applied immune complexome analysis of plasma to the study of Chagas disease caused by Trypanosoma cruzi. Twenty seropositive plasma samples including cardiac and/or megacolon determinate patients (n = 11) and indeterminate (n = 9) were analysed along with 10 seronegative individuals to characterize the antigens bound to circulating ICs. We identified 39 T. cruzi antigens and 114 human autoantigens specific to patients with Chagas. Among those antigens, two T. cruzi antigens (surface protease GP63, glucose-6-isomerase) and six human autoantigens (CD180 antigen, ceruloplasmin, fibrinogen beta chain, fibrinogen beta chain isoform 2 preprotein, isoform gamma-A of fibrinogen γ-chain, serum paraoxonase) were detected in more than 50% of the patients tested. Human isoform short of complement factor H-related protein 2 and trans-sialidase of T. cruzi were more frequently found in the indeterminate (5/9 for both) compared with in the determinate Chagas (0/11, P = 0·046 for human, 1/11, P = 0·0498 for T. cruzi). The immune complexome could illustrate the difference of immune status between clinical forms of chronic Chagas disease.

Evaluation of lophine derivatives as L-012 (luminol analog)-dependent chemiluminescence enhancers for measuring horseradish peroxidase and H2O2.

8-Amino-5-chloro-7-phenylpyrido[3,4-d]pyridazine-1,4(2H,3H)dione (L-012) was recently synthesized as a new chemiluminescence (CL) probe; the light intensity and the sensitivity of L-012 are higher than those of other CL probes such as luminol. Previously, our group developed four lophine-based CL enhancers of the horseradish peroxidase (HRP)-catalyzed CL oxidation of luminol, namely 2-(4-hydroxyphenyl)-4,5-diphenylimidazole (HDI), 2-(4-hydroxyphenyl)-4,5-di(2-pyridyl)imidazole (HPI), 4-(4,5-diphenyl-1H-imidazol-2-yl)phenylboronic acid (DPA), and 4-[4,5-di(2-pyridyl)-1H-imidazol-2-yl]phenylboronic acid (DPPA), and showed that DPPA was suitable for the photographic detection of HRP. In this study, we replaced luminol with L-012 and evaluated these as L-012-dependent CL enhancers. In addition, to detect HRP and/or H2O2 with higher sensitivity, each detection condition for the L-012-HRP-H2O2 enhanced CL was optimized. All the derivatives enhanced the L-012-dependent CL as well as luminol CL; HPI generated the highest enhanced luminescence. Under optimized conditions for HRP detection, the detection limit of HRP was 0.08 fmol. By contrast, the detection limit of HRP with the enhanced L-012-dependent CL using 4-iodophenol, which is a common enhancer of luminol CL, was 1.1 fmol. With regard to H2O2 detection, the detection limits for enhanced CL with HPI and 4-iodophenol were 0.29 and 1.5 pmol, respectively. Therefore, it is demonstrated that HPI is the most superior L-012-dependent CL enhancer.

Assessment of air pollution in Nagasaki City: determination of polycyclic aromatic hydrocarbons and their nitrated derivatives, and some metals.

Air pollutants such as polycyclic aromatic hydrocarbons (PAHs), their nitrated derivatives (NPAHs), and some metals on airborne particles in Nagasaki city, Japan were determined over a period of 12 months by high-performance liquid chromatography with chemiluminescence, fluorescence and flameless atomic absorption spectrometry. The average concentrations (range) were 18.24 (4.07-41.54) ng/m3 for total PAHs, 0.91 (0.23-4.10) pg/m3 for NPAHs, 7.95 (1.71-16.31) ng/m3 for Pb, 11.56 (3.35-24.96) ng/m3 for Mn and 3.79 (0.97-14.71) ng/m3 for Ni (n = 136). The toxic equivalency factors adjusted concentration of total PAHs determined in Nagasaki city area was 2.33 ng/m3. Concentrations of total PAHs and NPAHs in winter were higher than those in summer. In a weekly variations study, total PAHs and NPAHs concentrations, as well as traffic volume showed a similar tendency with all values higher during weekdays and lower at the weekend. The correlation coefficients between total PAHs or NPAHs and traffic volume were 0.781 and 0.818, respectively. These results suggested that one of the main sources for NPAHs and PAHs in a city area might be motor vehicles.

Activity of the rat pontomedullary reticular neurons related to rhythmical jaw movements.

Electrical stimulation of the cerebral peduncle or oral mechanical or chemical stimulation induced rhythmical jaw movements (RJM) in the anesthetized rat. Extracellular recording was made from the pontomedullary reticular neurons in relation to RJM. The RJM-related activity was classified to a tonic, a transient and a phase-dependent rhythmical activities (116, 92 and 45 neurons, respectively). We found that the neurons showing the phase-dependent activities discharged exclusively or almost exclusively during RJM without responding to at least one of three kinds of RJM-inducing stimulation. Fourteen neurons were activated by all kinds of RJM-inducing stimulation and majority of them showed weak or no change in activity during RJM, although some showed the transient or the rhythmical activity during RJM. We propose that the phase-dependent rhythmical activity is necessary to generate RJM since any stimulation-induced RJM was reversibly blocked by microinjection of lidocaine or glutamate receptor blocking agents into the brainstem site concentrated by this type of neurons. No other type of activity was observed exclusively during RJM.

Primary round cell liposarcoma of the omentum. A case report.

Liposarcomas are the most common of the malignant soft tissue tumors arising in the mediastinum and retroperitoneum. We have treated patients for round cell liposarcoma, an unusual histologic subtype, which is reported to be histologically mistaken for Ewing's sarcoma, lymphoma and other round cell neoplasms. The successful resection of a 2,300-g round cell liposarcoma of the omentum was recently performed at our hospital. Ultrastructural observations of this tumor are characterized by massive proliferation of the mitochondria in addition to the presence of lipid vacuoles in tumor cells. To our knowledge, no description of substantial mitochondrial proliferation in round cell liposarcoma has appeared previously in the literature; therefore, this is the first case report of histopathologic, cytopathologic and electron microscopic observations on this extremely rare tumor.