Evaluation of anti-parvovirus B19 activity in sera by assay using quantitative polymerase chain reaction

Human parvovirus B19 (B19) infects cells of erythroid lineage. Production of neutralizing antibodies (Abs) is indispensable for recovery from B19-related disease state. In this study, we used a convenient method to measure neutralizing activities in human sera by using a real-time quantitative PCR based assay. Erythroid cell line KU812Ep6 was incubated with test sera before infection with B19 virus. The copy number of B19-DNA in cultures was decreased in the presence of the sera from patients who recovered from acute B19 infection, whereas no decrease in B19-DNA was in cultures incubated with sera from healthy volunteers who had no B19 infection. The decrease in B19-DNA copy number was calculated and the inhibition percentage was expressed as neutralizing activity to B19. A clinical study showed that the levels of neutralizing ability were high in patients who recovered soon after acute B19 infection, but were low in some patients with a prolonged clinical course for recovery from B19 infection. This method is simple and convenient compared with methods described previously, showing its usefulness to evaluate the neutralizing activity to B19.     

Endothelium-dependent vasodilation in type II diabetes mellitus

Endothelial dysfunction is a major feature of atherosclerosis and it can also serve as an early atherosclerotic marker. Evaluation and assessment of the endothelial function is important to prevent serious atherosclerotic disease especially myocardial infarction, cerebrovascular disease and renal failure. To evaluate endothelial function we measured endothelium-dependent vasodilation (flow-mediated dilatation: %FMD) of the brachial artery with ultrasound. This method is non-invasive and can be repeatable in order to follow patients individually. Progressive atherosclerosis is often observed in diabetic patients who are not hypertensive. To evaluate the impairment of the endothelial function in type 2 diabetic patients, we examined %FMD in them and compared with hypertensive patients without diabetes and control subjects. We found that type 2 diabetic patients had the same endothelial dysfunction as hypertensive patients without diabetes. %FMD in both diabetic patients and hypertensive patients was lower than in control subjects. Moreover, %FMD of type 2 diabetic patients with hypertension was lower than %FMD of type 2 diabetic patients without hypertension. These finding suggests that endothelial dysfunction develops under the conditions of hypertension and hyperglycemia. Evaluating endothelial function with ultrasound is useful for assessment of atherosclerosis in diabetes.     

Bcl11b is required for differentiation and survival of alphabeta T lymphocytes

The gene Bcl11b, which encodes zinc finger proteins, and its paralog, Bcl11a, are associated with immune-system malignancies. We have generated Bcl11b-deficient mice that show a block at the CD4-CD8- double-negative stage of thymocyte development without any impairment in cells of B- or gammadelta T cell lineages. The Bcl11b-/- thymocytes showed unsuccessful recombination of V(beta) to D(beta) and lacked the pre-T cell receptor (TCR) complex on the cell surface, owing to the absence of Tcrb mRNA expression. In addition, we saw profound apoptosis in the thymus of neonatal Bcl11b-/- mice. These results suggest that Bcl11b is a key regulator of both differentiation and survival during thymocyte development.     

Mutational screening of APP gene in patients with early-onset Alzheimer disease utilizing mismatched PCR-RFLP

To elucidate the frequency of mutations of the β/A4 amyloid protein precursor (APP) gene in early-onset Alzheimer disease, we designed a mismatched PCR-RFLP that can identify all kinds of missense mutations at codon 717 in addition to the seven kinds of known mutations at exon 17. When we screened mutations at exon 17 utilizing this method and the double missense mutations at exon 16 of the APP gene by PCR-RFLP, no cases revealed mutations of the APP gene among 13 familial and 54 sporadic cases, except one family (OS-1) that had previously been reported and used as a positive control of APP717(Val → Ile). Our results support the hypothesis that mutations in the APP gene are not major causes in early-onset Alzheimer disease.     

MEG Characterization of Spontaneous Alpha Rhythm in the Human Brain

The present piece of research studied the spontaneous alpha rhythm of the human brain by combining the use of a whole-cortex neuromagnetometer and Magnetic Resonance Imaging. Single trials of spontaneous brain activity were recorded from ten human subjects asked to rest, with their eyes either closed or open, in relaxed wakefulness. MEG measurements were conducted over a period of one and a half years. The replicability of the results was confirmed for eight subjects out of ten. For three subjects, the alpha rhythm did not show any reductions due to the opening of the eyes. Both field map pattern and location of the estimated source were persistently stationary during each of the bursts of oscillations of the alpha rhythm. Dipoles were concentrated in clusters, indicating the existence of several spatially distributed sources. The calcarine fissure, the parieto-occipital sulcus and the surrounding occipital and parieto-occipital areas were identified as cortical sites of the brain where the alpha rhythm may originate. For four subjects, the majority of the sources were located near or in the calcarine fissure, while for five subjects, they were located near or in the parieto-occipital sulcus and for the remaining subject they were equally divided between the two generation sites.     

Cloning and nucleotide sequencing of Vero toxin 2 variant genes from Escherichia coli O91:H21 isolated from a patient with the hemolytic uremic syndrome

Cellular DNA extracted from Escherichia coli strain B2F1 (O91:H21) was found to contain two separate DNA sequences that hybridized with a Vero toxin 2 (VT2)-specific gene probe under stringent conditions. These two sequences were cloned and both were shown to encode a variant of Vero toxin 2 (VT2vh). The nucleotide sequences of the operons encoding VT2vh, designated as vtx2ha and vtx2hb, were determined. The two operons were nearly identical (99% overall DNA homology) and both encoded A subunits of 319 amino acid residues and B subunits of 89 amino acid residues, the A and B subunit genes being separated by a stretch of 14 bp. The A and B subunit genes of the vtx2ha operon exhibited 98.6% and 95.5% DNA homology, respectively, with those of the slt-II operon encoding Shiga-like toxin II (or VT2) cloned from a strain from a patient with hemorrhagic colitis, while the A and B subunit genes of the vtx2ha operon showed 94.5% and 82.8% DNA homology, respectively, with those of the slt-IIv operon encoding a SLT-II variant cloned from a strain isolated from a pig with edema disease. The nucleotide sequences of the presumed promoters and presumptive ribosome binding sites in the vtx2ha, vtx2hb, and slt-II, and slt-IIv operons were identical. These results indicate that nucleotide sequences encoding a family of VT2-related toxins are present in various strains of E. coli and that the sequences of the genes for A subunits are better conserved than those of the B subunit genes.     

Benzylpenicillin preparations can evoke a systemic anaphylactic reaction in guinea pigs

All of the five commercially available benzylpenicillin preparations obtained from different sources and a PcG preparation prepared by filtration of a commercial PcG on Sephadex G10 elicited the systemic anaphylactic reactions in guinea pigs which had been immunized with benzylpenicilloyl (BPO)-Ascaris extract conjugate (BPO-As) mixed with aluminum hydroxide gel. These preparations could evoke no such reactions in guinea pigs immunized with BPO-bovine gamma globulin conjugate (BPO-BGG) emulsified with complete Freund's adjuvant. The severity of the systemic anaphylactic reactions correlated significantly with the titers of either 8-day passive cutaneous anaphylactic (8-day PCA) reactions or 4-hr PCA reactions evoked with the same benzylpenicillin preparations. In vitro benzylpenicillin preparation contracted the tracheas of the guinea pigs immunized with BPO-As. These results indicated that the commercially available benzylpenicillin preparations have enough antigenicity to evoke systemic anaphylactic reactions in guinea pigs immunized with BPO-As mixed with aluminum hydroxide gel. Such guinea pigs represent an animal model for investigation of penicillin allergy.     

Blepharophimosis sequence and de novo balanced autosomal translocation [46, XY,t(3;4)(q23;p15.2)]: Possible assignment of the trait to 3q23

We report on a boy with the blepharophimosis sequence and de novo, apparently balanced reciprocal translocation between 3q23 and 4p15.2 [46, XY,t(3;4)(q23;p15.2)de novo]. Possible assignment of this autosomal dominant disorder is discussed. A 3q23 band is a more preferable gene locus of the blepharophi mosis sequence, based on the comparison of clinical manifestations between 4p- and 3q-syndromes.     

Are cerebrovascular factors involved in Alzheimer's disease?

Recent epidemiological studies have shown that vascular risk factors may be involved in Alzheimer's disease (AD) as well as dementia in general. To investigate the relation between a vascular disorder and AD pathology, current criteria are defective because most depend on exclusion of a cerebrovascular disorder. Epidemiological studies have indicated the possibilities that arteriosclerosis, abnormal blood pressure, diabetes mellitus and smoking may be related to the pathogenesis of AD. As for the mechanism that vascular disorders influence AD, it is presumed that amyloid deposition may be caused by a vascular disorder. Alternatively, a vascular event may cause progression of subclinical AD to a clinical stage. Insulin resistance and apolipoprotein E may also be involved in these mechanisms. Our studies show that ischemia-induced the Alzheimer-associated gene presenilin 1 (PS1) and endoplasmic reticulum-stress, generated from a vascular disorder, may unmask clinical AD symptoms caused by presenilin mutation, suggesting that a vascular factor might be involved in the onset of familial AD.     

Colchicine-induced Inhibition of Fat Globule Development in Hepatocytes of Rats Injected with Ethionine

To examine the effect of colchicine on ethionine induced fatty liver, adult female rats were starved overnight and then injected i.p. with 1 g kg ethionine at 11th hour of fasting; then a half of the rats were also injected i.p. with 2.5 mg kg colchicine twice at 3 and 6 h after the single administration of ethionine. Similarly, fasted control rats were injected i.p. with vehicle alone at the above times. All of the rats were sacrificed after a 20 h fast, and the hepatocytes in periportal areas were observed ultra-structurally. In addition, total lipids in the liver tissue were extracted and determined biochemically. Although similar significant increases of triglyceride were observed in the liver tissue of all ethionine-injected rats, the hapatocytes in the group treated with both chemicals had fewer cytoplasmic fat globules (CFG) than those in the group treated with ethionine only. On the other hand, the diameters of markedly increased membrane-bound lipid particles (MLP) in the double treated group were distributed mainly in the range 0.2–0.4 μm, compared with those (0.1-0.2 μm) in the other groups. These findings indicate that colchicine inhibits the development of CFG in ethionine injured hapatocytes. Acta Pathol Jpn 39: 281∼288, 1989.