Morphology of equine allantochorion at the tip of the pregnant horn

The morphology of the equine allantochorion at the tip of the pregnant horn was studied in the membranes of 14 mares. The findings in the allantochorion at the tip of the pregnant horn were of two types; one was growth retardation (hypoplastic villi, tunica adventitia of the vessels resembling embryonal connective tissue and the extended spaces of remnants of the extraembryonic coelom), the other was placental hypoxia or ischaemia (parakeratosis, stratified squamous metaplasia, necrosis of the trophoblasts, thickening of the basement membrane and fibrous hyperplasia of villous stroma). It seems likely that hypoplastic villi are caused by disordered formation of the microcotyledon accompanying the progression of pregnancy.     

Spatiotemporal changes of coxsackievirus and adenovirus receptor in rat hearts during postnatal development and in cultured cardiomyocytes of neonatal rat

Coxsackievirus B is the most common cause of viral myocarditis and is particularly virulent in neonates and children. Adenovirus is also a leading cause of the disease. The determinant of tropism for both viruses is considered to be the expression of coxsackievirus and adenovirus receptor (CAR) in target organs. However, developmental change and physiological localization of CAR in the heart are unknown. We examined expression levels of CAR in rat hearts by quantitative real-time polymerase chain reaction and Western blot analysis and found that CAR decreased gradually during postnatal development, although CAR was detectable, even in adults. Immunohistochemistry revealed CAR on the whole surface of cardiomyocytes in immature rat hearts. In contrast, CAR was detected predominantly on intercalated disks in the adult heart and was accumulated especially at the contact point between the cultured cardiomyocytes, even though they were prepared from the neonatal rat heart. In conclusion, CAR was expressed abundantly on the whole surface of cardiomyocytes in immature rat hearts. Both the expression level and the localization of CAR are possible determinants of the susceptibility to viral myocarditis of neonates and children.     

Effects of repeated subculturing and prolonged storage at room temperature of enterohemorrhagic Escherichia coli O157:H7 on pulsed-field gel electrophoresis profiles

Three clinical strains of enterohemorrhagic Escherichia coli O157:H7 which were subcultured repeatedly or stored at room temperature over a 25-week period showed appreciable variations in their pulsed-field gel electrophoresis fragment patterns. The variations could be explained by a couple of spontaneous genetic events at most and thus did not invalidate the genetic lineage of the strains.     

ACh receptor protein drives primary and memory autoantibody responses in chimeric human-SCID mice

The native antigen that drives the T-helper cells regulating production of muscle acetylcholine receptor (AChR) autoantibodies is unknown. Human T cell lines activated by autoantigens in vitro are of unproven relevance to B cell help. Here we report the functional interaction and unprecedented longevity of AChR-specific human T and B lymphocytes residing in SCID mice. Lymphoid cells from myasthenia gravis (MG) patients and healthy subjects were injected ip. Recombinant human AChR-alpha1-subunit-1-210 was injected after day 75. Human AChR-specific Ig was produced rapidly in MG-SCID mice challenged once. Only 1 of 32 control hu-SCID mice produced AChR-specific Ig. This required multiple immunizations, was initially cross-reactive with Torpedo AChR, and had a slow course. Thus, memory T and B lymphocytes specific for human AChR-alpha1-subunit are readily demonstrable in MG patients, interact to produce autoantibody of the same restricted specificity found in the donor's serum, and are long-lived without exogenous autoantigen challenge. In healthy subjects, AChR-specific lymphocytes are infrequent and exhibit naive response characteristics, including apparent affinity maturation of Ig specificity.     

Rapid detection of Epstein-Barr virus DNA by loop-mediated isothermal amplification method.

BACKGROUND: The loop-mediated isothermal amplification (LAMP) method is a novel technique for the amplification of specific DNA sequences. OBJECTIVES: To establish the LAMP method for amplifying Epstein-Barr virus (EBV) DNA and to examine its reliability for the detection of EBV DNA in clinical specimens. STUDY DESIGN: Sera from 108 patients, who were initially suspected of primary EBV infection, were tested by the EBV LAMP method, and the results were compared with those of the real-time PCR assay. Serological examination was regarded as the standard diagnostic method. RESULTS: To diagnose primary EBV infection, the sensitivity of LAMP was 86.4% and the specificity was 100%. The sensitivity of the real-time PCR assay was 84.1% and the specificity was 98.4%. Longitudinal analysis showed that the detection rate of EBV DNA in serum by the LAMP method decreased with time in accordance with the decrease of the EBV load. EBV DNA could not be detected in serum 40 days after onset of symptoms. CONCLUSIONS: These results indicate that the sensitivity and specificity of the LAMP method are comparable to those of real-time PCR and that detecting EBV DNA in serum by this method is potentially useful for diagnosing primary EBV infection.     

Family-based association study of schizophrenia with 444 markers and analysis of a new susceptibility locus mapped to 11q13.3.

Family-based linkage disequilibrium (LD) mapping has been suggested as a powerful and practical alternative to linkage analysis. We have performed a genome-wide LD survey of susceptibility loci for schizophrenia in a Japanese population. We first typed 119 schizophrenic pedigrees (357 individuals) using 444 microsatellite markers, and analyzed the data using the pedigree disequilibrium test. This analysis revealed 14 markers demonstrating significant transmission distortion. To corroborate these findings, the statistical methods were changed to the extended transmission disequilibrium test (ETDT), using 80 independent complete trios (schizophrenic proband and both parents), with 68 derived from initial pedigrees and 12 newly recruited trios. ETDT supported two markers for continued association, D11S987 on 11q13.3 (P = 0.00009) and D16S423 on 16p13.3 (P = 0.002). We scrutinized the most significant genomic locus on 11q11-13 by adding 26 new markers for analysis. Results of three-marker haplotype analysis in the region showed evidence of association with schizophrenia (most significant haplotype P = 0.0005, global P = 0.022). Although the present study may have missed other potential genomic intervals because of the sparse mapping density, we hope that it has identified promising anchor points for further studies to identify risk-conferring genes for schizophrenia in the Japanese population. In addition, we provide useful information on genomic LD structures in Japanese populations, which can be used for LD mapping of complex diseases.     

Apoptosis-inducing protein derived from hepatocyte selectively induces apoptosis in lymphocytes

The liver is where lymphocytes undergo activation-induced cell death (AICD) at the resolution phase of an immune response, which is crucial for homeostasis of the immune system and prevention of autoimmunity. Exploring the machinery of AICD in the liver, we found that a primary culture supernatant of murine hepatocytes had an antiproliferative effect on antigen-stimulated T clone and T lymphoma cells. Biological study showed that the antiproliferation was due to induction of apoptosis in a caspase-dependent manner. The apoptosis-inducing potential was sensitive to trypsin, heat (> 70 degrees ) and acid (< pH 5) treatment but could not be neutralized by anti-tumour necrosis factor-alpha, anti-Fas ligand, or anti-transforming growth factor-beta antibodies. Biochemical study of the isolated and purified apoptosis-inducing component from the supernatant showed that it was a protein with a molecular mass of about 68,000-70,000. It induced apoptotic change in murine T and B cells, and to a lesser degree, in human lymphoid cells, but not in macrophages. Biochemical and biological characteristics distinguish this protein from others that have been reported to induce apoptosis of lymphocytes. The identification of an apoptosis-inducing protein derived from murine hepatocytes, which selectively induces apoptosis in lymphocytes, suggests one possible mechanism for immune suppression in the liver.     

Astaxanthin limits exercise-induced skeletal and cardiac muscle damage in mice

Dietary antioxidants may attenuate oxidative damage from strenuous exercise in various tissues. Beneficial effects of the antioxidant astaxanthin have been demonstrated in vitro, but not yet in vivo. We investigated the effect of dietary supplementation with astaxanthin on oxidative damage induced by strenuous exercise in mouse gastrocnemius and heart. C57BL/6 mice (7 weeks old) were divided into groups: rested control, intense exercise, and exercise with astaxanthin supplementation. After 3 weeks of exercise acclimation, both exercise groups ran on a treadmill at 28 m/min until exhaustion. Exercise-increased 4-hydroxy-2-nonenal-modified protein and 8-hydroxy-2'-deoxyguanosine in gastrocnemius and heart were blunted in the astaxanthin group. Increases in plasma creatine kinase activity, and in myeloperoxidase activity in gastrocnemius and heart, also were lessened by astaxanthin. Astaxanthin showed accumulation in gastrocnemius and heart from the 3 week supplementation. Astaxanthin can attenuate exercise-induced damage in mouse skeletal muscle and heart, including an associated neutrophil infiltration that induces further damage.     

Comparative studies of dose-response curves for recessive lethal mutations induced by ethylnitrosourea in spermatogonia and in spermatozoa of Drosophila melanogaster

Induction of recessive lethal mutations by N-ethyl-N-nitrosourea (ENU) was studied for the second chromosome of spermatogonia and spermatozoa in Drosophila melanogaster. ENU (0.03, 0.3, and 1.0 mM) was given to flies by dissolving it in feeding sucrose solution. Since flies are known to show increased avoidance of the feeding solution with increase in ENU concentration, the absorbed doses of ENU by flies were estimated from the previously determined empirical data for relation of ENU concentration in the feeding solution and the intake volumes of the solution by flies as measured indirectly via intake radioactivities of 3H-labeled sucrose added to the solution. When plotted against absorbed doses of ENU, the observed frequencies of recessive lethals showed a linear relationship for induction in spermatozoa but a sigmoidal relationship for induction in spermatogonia. These results suggest that in spermatogonia ENU-induced mutational damage is more repairable in a lower dose range of ENU. Mosaic lethal mutations were induced by ENU but not in spermatogonia.