Strategies for optimum use of superposition diffractogram in scanning electron microscopy.

Characteristics of the superposition diffractogram used for precisely estimating scanning electron microscopy (SEM) resolution are investigated. It is shown that the choice of pixel density to satisfy the sampling theorem, the direction of scanning, the choice of image shift direction, the properties of the specimen, the effect of external disturbances such as vibration and stray magnetic fields, and the effect of the window function required in the Fourier transform, are all factors which must be considered in order to make the superposition diffractogram a practical technique. An additional important improvement required to exploit fully the ability of the superposition diffractogram, which potentially is very high, is a special scanning mode which employs a digital scan generator, and digital image processing technology with autocorrelation functions.

Porcine MCP gene promoter directs high level expression of human DAF (CD55) in transgenic mice.

Porcine membrane cofactor protein (pMCP), a complement regulatory protein, is widely expressed in various tissues. Particularly, it is highly expressed on vascular endothelium. The objective of this study was to investigate whether the pMCP gene promoter can induce efficient expression of a human complement regulatory protein, decay-accelerating factor (DAF; CD55) in transgenic mice. Two fragments of the 5'-flanking region of pMCP gene (0.9 kb and 5.4 kb) connected with human DAF minigene (0.9/hDAF and 5.4/hDAF) were used to produce transgenic mice. The expression of hDAF in heart, liver, kidney, lung, pancreas, brain and testis of the transgenic mice was examined by immunohistochemical analysis. The vascular endothelia and the nerves in all organs examined were intensely stained. The staining pattern in these tissues was similar in all transgenic mice examined regardless of the length of the promoters. The surface expression levels of hDAF on peripheral red blood cells and splenocytes from a mouse carrying 5.4/hDAF hemizygously was twice the level of expression on corresponding human cells. The red blood cells and splenocytes from the transgenic mice exhibited resistance to lysis by human serum in a manner dependent upon expressed hDAF level. The hearts from the transgenic mice functioned for a significantly longer time than those from normal mice under perfusion with human serum in the Langendorff perfusion system. These results demonstrated that the pMCP gene promoter is a good candidate of the regulatory element in the transgene to produce transgenic animals for xenotransplantation.

Molecular cloning of a pig homologue of membrane cofactor protein (CD46).

Organs of transgenic pigs that express human complement regulatory proteins are under assessment as an alternative to transplantation. A major barrier to the transplantation of pig organs is the hyperacute rejection caused by pre-existing antibodies and complement. Pig cells are very susceptible to human complement, presumably because pig cell-surface complement regulatory proteins are inefficient against it. Expression of human complement regulatory proteins, such as decay-accelerating factor and membrane cofactor proteins (MCP or CD46), by means of transgenes would confer resistance to human complement upon pig cells, thereby preventing hyperacute rejection. To express sufficient levels of human complement regulatory proteins at appropriate sites, regulatory elements of genes of pig membrane-bound complement regulatory proteins would be useful. To obtain their cDNAs, we transfected human cells with a pig cDNA library, selected cells by incubation with pig complement and rescued the plasmids. We cloned a cDNA for the pig homologue of MCP, pMCP. The cDNA encoded a predicted protein of 363 amino acids with 42% amino acid identity with human MCP. The pMCP consisted of four short consensus repeats, a Ser/Thr/Pro-rich domain, and transmembrane and cytoplasmic domains. Recombinant soluble pMCP that lacked transmembrane and cytoplasmic domains had factor I cofactor activity in C3b cleavage, indicating that it is functionally, as well as structurally homologous to MCP. FACS analysis with anti-pMCP mAb demonstrated that pMCP is expressed on all blood leukocytes, erythrocytes, and on endothelial and epithelial cell lines.

A beneficial role of bile pigments as an endogenous tissue protector: anti-complement effects of biliverdin and conjugated bilirubin.

Bile pigments possess an anti-complement property and could be involved in tissue protection. In this study, we examined the physiological actions of bile pigments, which had been generally regarded as waste catabolites. Biliverdin inhibited complement cascade reactions in vitro, especially at the C1 step in the classical pathway at low micromolar concentrations. Further, Forssman anaphylaxis in guinea pigs, being closely associated with complement reactions, was inhibited by oral or intravenous administration of biliverdin. Conjugated bilirubin also showed an inhibitory effect on complement-dependent reactions in vitro. From these observations, we propose a hypothesis that the pigments serve as endogenous tissue protectors by multiple lines of mechanisms including antioxidant and anti-complement actions.

[Report of three cases of purple urine bag syndrome which occurred with a combination of both E. coli and M. morganii].

Purple urine bag syndrome is a rare phenomenon in which bags turns purple or blue following catheterization. This phenomenon was reported to occur by Providencia stuartii in the presence of indicanuria. We reported herein three females above 75 years of age having a similar phenomenon. All were bedridden chronically and constipated. In these cases, no growth of P. stuartii was found on repeated urine culture, but Escherichia coli and Morganella morganii were concomitantly isolated during the blue bags. It is suggested that purple urine bag syndrome develops in a combination of E. coli and M. morganii besides P. stuartii.

Model insomnia by methylphenidate and caffeine and use in the evaluation of temazepam.

Experimental sleep disturbances (model insomnia) were produced by the administration of methylphenidate (MPD) 10 mg and caffeine (CAF) 150 mg. The effect of temazepam (TEM), 15 mg or 30 mg, on the model was investigated. All-night polysomnography was performed on 8 normal young male subjects under each of the following 9 conditions: baseline, MPD 10 mg, CAF 150 mg, TEM 15 mg, TEM 30 mg, MPD + TEM 15 mg, MPD + TEM 30 mg, CAF + TEM 15 mg, CAF + TEM 30 mg. A reduction in total sleep time and total amount of stage REM (S-REM) sleep and an increase in the sleep latency and wake time (S-W) were observed in both the MPD and CAF nights. The sleep latency was significantly longer in the CAF night than in the MPD night. Administration of TEM 15 mg or TEM 30 mg alone caused very few modifications in the sleep parameters. These drugs in combination with MPD or CAF resulted in almost complete recovery of the sleep disturbance induced by MPD or CAF. The results indicate that CAF and MPD produced similar models of insomnia except for a greater sleep latency for CAF than for MPD. Both models were useful in the evaluation of hypnotic drugs such as temazepam.

Regional relationship of visual evoked potentials in infants and school-aged children.

VEPs were recorded from the inion, O1, O2, Pz, C3 and C4 in 74 normal children, aged 1--13 years. These VEPs were compared at the various scalp areas using a correlational technique. Wave forms of VEPs were different for different scalp areas; in particular, a sharp distinction was seen in the positive component having a peak latency of about 100 msec. The component was occipital-dominant, showing a maximum amplitude at the inion. Correlations of VEPs from the right and left hemispheres of the central area were high for all ages, while those of the right and left hemispheres of the occipital area were generally lower and showed greater inter-individual variability than in the central area. Correlations between VEPs from the occipital and non-occipital areas (i.e., inion-Pz, inion-C4, O1-C3 and O2-C4), with the exception of inion-Pz, were significantly greater with increasing age.