Development of a novel two dimensional surface plasmon resonance sensor using multiplied beam splitting optics.

A novel two dimensional surface plasmon resonance (SPR) sensor system with a multi-point sensing region is described. The use of multiplied beam splitting optics, as a core technology, permitted multi-point sensing to be achieved. This system was capable of simultaneously measuring nine sensing points. Calibration curves for sucrose obtained on nine sensing points were linear in the range of 0-10% with a correlation factor of 0.996-0.998 with a relative standard deviation of 0.090-4.0%. The detection limits defined as S/N = 3 were 1.98 × 10(-6) - 3.91 × 10(-5) RIU. This sensitivity is comparable to that of conventional SPR sensors.

[A study on diffuse alveolar damage and ubiquitin-positive pneumocytes in the elderly].

AIM: The ubiquitin proteasome system (UPS) is important because homeostasis through proteolysis and ubiquitin (Ub) has been observed to appear in diseases of the central nervous system. However, studies on UPS in relation to pulmonary diseases are few and no other investigators have described Ub-positive cells in the lung. Intracytoplasmic eosinophilic inclusion bodies in pneumocytes have been known to appear in cases of diffuse alveolar damage (DAD). We found that these inclusion bodies in DAD were positive for Ub. In this study, DAD cases in the elderly were studied to clarify the relationship between Ub-positive cells and cellular damage in the lungs. METHODS: Representative lung fields from a total of 26 patients with DAD were studied immunohistochemically, using Ub staining. The severity of DAD was evaluated after each case was scored for hyaline membrane formation and lung injury, respectively. Non-DAD diseases from 19 autopsy cases were studied as controls. The mean age of both groups was 72.1. RESULTS: Variably sized and shaped eosinophilic inclusion bodies were found in 7 cases (26.9%) of the DAD cases (inclusion body group) and all inclusion bodies were positive for Ub and cytokeratin KL-1. Pneumocytes without inclusion bodies were occasionally positive for Ub, with an intracytoplasmic granular pattern, not only in the inclusion body group but also in the non-inclusion body group (4 of DAD cases). These Ub-positive cells (both Ub-positive inclusion bodies and the granular Ub-positive cells) were found to have high rate of hyaline membrane formation and lung injury in the DAD cases. CONCLUSION: Elderly DAD cases had Ub-positive inclusion bodies in pneumocytes and Ub-positive pneumocytes were found with or without the inclusion bodies in DAD. This means that accumulation of ubiquitinated protein including cytokeratin could be recognized as inclusion bodies in pneumocytes. The presence of these Ub-positive cells might be a morphological indicator for evaluation of cellular damage in the patients with DAD.

The phosphoprotein of attenuated measles AIK-C vaccine strain contributes to its temperature-sensitive phenotype.

Measles AIK-C vaccine strain exhibits temperature-sensitivity (ts). To identify the structural proteins, which contribute to the ts property of AIK-C virus, reverse genetics was used. MV-minigenome RNA was replicated at 32.5, 37, and 39 degrees C when the plasmids expressing N, P, and L proteins of the Edmonston strain (the parental strain of AIK-C) were used, whereas the minigenome RNA replicated only at 32.5 degrees C but did not at 37 degrees C and higher temperature when N, P, and L protein expression plasmids of the AIK-C strain were used. A series of minigenome experiments revealed that the amino acid substitution of leucine at position 439 of the P protein by proline (P439-Pro) contributes to the ts phenotype of AIK-C. Four recombinant viruses having various P genes were rescued from the modified AIK-C genome cDNA and ts-characteristics were compared in Vero cells by plaque formation assay. The results showed that the P439-Pro of AIK-C virus played a key role in the ts phenotype, but the other substitutions in the P gene might have an accessory function in the expression of the phenotype.

Neuronal expression of alphaB crystallin in cerebral infarction.

alphaB crystallin (alphaBC) is one of the heat shock proteins that are induced under stressful conditions. In normal brains, alphaBC is present in oligodendrocytes and astrocytes, but not in neurons. Neuronal alphaBC expression in the central nervous system under pathological conditions has been investigated in several previous studies, most of which dealt with various neurodegenerative diseases with and without ballooned neurons. Neuronal expression of alphaBC has seldom been studied in cerebral infarction (CI), and the frequency of alphaBC-positive neurons in the various stages of CI is unknown. To investigate this issue, we examined 48 autopsy brains of patients with CI, and found neuronal expression of alphaBC in 68.8% of the cases. We found three types of alphaBC-positive neurons: normal morphological, convex, and ballooned neurons. Although alphaBC-positive neurons were present in the every stage of CI, they were more frequent later than 10 days after the onset of CI, and the frequency of alphaBC-positive ballooned neurons was particularly increased in the later stages of CI. This may indicate that morphologically normal neurons gradually swelled up through convex neurons, finally forming ballooned neurons. Previous studies indicated that alphaBC might have a cytoprotective function as a molecular chaperone, and we speculate that alphaBC is expressed in neurons subjected to ischemic stress, and exerts a cytoprotective effect on the neurons.

Ubiquitin-positive inclusions in ependymal cells.

Ubiquitin-positive inclusions (UbIs) have not been well studied in ependymal cells. Since we detected such UbIs in the central canals of the medulla and spinal cord while investigating UbIs in neurodegenerative diseases, we studied UbIs in the entire ependymal system of 42 patients with various neurological diseases and of 10 non-neurological controls. UbIs were located in the cytoplasm of the ependymal cells, and were round to oval in shape, measuring 4-11 microm in diameter. The UbIs were non-argyrophilic and undetectable by hematoxylin and eosin staining, but mildly reactive to periodic acid-Schiff staining with and without digestion. The UbIs were variably immunoreactive for anti-epithelial membrane antigen (EMA) antibody, but did not react with several other antibodies. The co-existence of ubiquitin and EMA was confirmed by confocal laser microscopy. Throughout the ependymal system, UbIs were variably found in ependymal cells as well as in subependymal cells. There was no significant difference in the overall incidence of either ependymal or subependymal UbIs between the patients with neurological diseases and controls. However, ependymal UbIs in the central canal were more frequent in the neurological disease patients than in controls, although there was no disease specificity. This is the first comprehensive report to show common occurrence of UbIs in the ependymal cells of adult human brains.

Clinical presentation of subacute sclerosing panencephalitis in Papua New Guinea.

Eighty-three children presented at Goroka Base Hospital in the Eastern Highlands Province (EHP) of Papua New Guinea over a period of 3 years and 9 months between February 1997 and November 2000 were confirmed to have subacute sclerosing panencephalitis (SSPE). Confirmation of the diagnosis was based on the demonstration of high titres of measles antibodies in the cerebrospinal fluid and/or serum in association with clinical features supportive of SSPE, including characteristic electroencephalographic changes and amplification of measles virus genome by reverse transcriptase polymerase chain reaction in some cases. The mean cerebrospinal fluid and serum enzyme immunoassay antibody levels among the SSPE patients were 38 250 and 860 580, respectively. The mean age of onset of SSPE was 7.9 +/- 2.6 years and ranged between 2 and 14 years. The overall male to female ratio was 1.2:1 and 1.4:1 for EHP.

Molecular analysis of measles virus genome derived from SSPE and acute measles patients in Papua, New Guinea.

A very high annual incidence of 56 per million population below the age of 20 years for subacute sclerosing panencephalitis (SSPE) has been reported from Papua New Guinea (PNG). In a more recent study, we have confirmed this unusual high incidence for Eastern Highlands Province (EHP) of PNG. In the study, it was observed that the vaccination rate among SSPE patients registered at Goroka Base General Hospital (GBGH) in EHP was higher than that of other infants in the province in recent years. To identify the measles virus (MV) responsible for SSPE in EHP, sequence analysis of hypervariable region of the N gene was performed from 13 MV genomes: 2 amplified from clinical specimens of SSPE patients and 11 from acute measles patients. In 2 cases among the 11 with acute measles, nucleotide sequence of the entire H gene derived from isolated viruses was determined. Both nucleotide sequence and phylogenetic tree analyses showed that the amplified MV cDNAs were closely related to one another and belonged to the D3 genotype though they were different from any previously reported MV sequences. No genome sequences of vaccine strains were detected. These findings suggest that the MV strains prevailing in the highlands of PNG belong to genotype D3 of the MV and this wild-type MV rather than the vaccine strains was likely to be responsible for SSPE in these patients.