Analysis of helicase gene mutations in Japanese Werner’s syndrome patients

The profile of helicase gene mutations was studied in 89 Japanese Werner’s syndrome (WRN) patients by examining the previously described mutations 1– 4 as well as a new mutation found during this study, designated mutation 5. Of 178 chromosomes (89 patients), 89 chromosomes (50%) had mutation 4, 11 (6.2%) chromosomes had mutation 1, and two chromosomes (1.1%) contained mutation 5. Mutations 2 and 3 were not observed in this patient population. The remaining 76 (42.7%) chromosomes had none of these mutations. A significant fraction of all patients (22 total patients, 24.7%) appear to be compound heterozygotes, including those carrying mutations of both types 1 and 4. The genotype analysis of the markers surrounding the WRN helicase gene strongly suggests that most of the chromosomes carrying either mutation 1 or 4 were derived from two single founders. Received: 25 July 1996 / Revised: 20 September 1996     

Highly sensitive analytical method for aluminum movement in soybean root through lumogallion staining

We present highly sensitive aluminum detection method in root using fluorescent lumogallion. Roots treated with 100 μM AlCl₃ including 0.2 mM CaCl₂ (pH 4.5) were stained for 60 min with 10 μM lumogallion fluorescence solution and fluorescence from aluminum complex in root was observed under confocal laser microscope. There was a good correlation between the intensity of fluorescence and aluminum content. When the amount of aluminum lost during each step in staining process was measured, it was found that about 10% of aluminum was lost only at staining stage. Through lumogallion staining method, aluminum accumulation especially at an early stage of aluminum treatment in root was shown. At the beginning (2 hr), aluminum began to be accumulated in root cap. After 4 hr treatment, the aluminum distribution was spread to about 3 mm from root apex in the root cap and outer cortex. When aluminum was found in the outer cortex in 3–5 mm from the root apex, the viability was tended to be decreased in the same area (6 hr). At the same time, aluminum amount in meristem was increased. However the comparison of lumogallion staining method with that of morin, which has been widely used to detect aluminum in root, the sensitivity of lumogallion method was found to be much higher.     

Adaptation to left-right reversed vision rapidly activates ipsilateral visual cortex in humans.

The brain mechanisms of adaptation to visual transposition are of increasing interest, not only for research on sensory-motor coordination, but also for neuropsychological rehabilitation. Sugita [Nature 380 (1996) 523] found that after adaptation to left-right reversed vision for one and a half months, monkey V1 neurons responded to stimuli presented not only in the contralateral visual field, but also in the ipsilateral visual field. To identify the underlying neuronal mechanisms of adaptation to visual transposition, we conducted fMRI and behavioral experiments for which four adult human subjects wore left-right reversing goggles for 35/39 days, and investigated: (1) whether ipsilateral V1 activation can be induced in human adult subjects; (2) if yes, when the ipsilateral activity starts, and what kind of behavioral/psychological changes occur accompanying the ipsilateral activity; (3) whether other visual cortices also show an ipsilateral activity change. The results of behavioral experiments showed that visuomotor coordinative function and internal representation of peripersonal space rapidly adapted to the left-right reversed vision within the first or second week. Accompanying these behavioral changes, we found that both primary (V1) and extrastriate (MT/MST) visual cortex in human adults responded to visual stimuli presented in the ipsilateral visual field. In addition, the ipsilateral activity started much sooner than the one and a half months, which had been expected from the monkey neurophysiological study. The results of the present study serve as physiological evidence of large-scale, cross-hemisphere, cerebral plasticity that exists even in adult human brain.     

Gene expression profiling identifies a set of transcripts that are up-regulated inhuman testicular seminoma.

Seminoma constitutes one subtype of human testicular germ cell tumors and is uniformly composed of cells that are morphologically similar to the primordial germ cells and/or the cells in the carcinoma in situ. We performed a genome-wide exploration of the genes that are specifically up-regulated in seminoma by oligonucleotide-based microarray analysis. This revealed 106 genes that are significantly and consistently up-regulated in the seminomas compared to the adjacent normal tissues of the testes. The microarray data were validated by semi-quantitative RT-PCR analysis. Of the 106 genes, 42 mapped to a small number of specific chromosomal regions, namely, 1q21, 2p23, 6p21-22, 7p14-15, 12pll, 12p13, 12q13-14 and 22q12-13. This list of up-regulated genes may be useful in identifying the causative oncogene(s) and/or the origin of seminoma. Furthermore, immunohistochemical analysis revealed that the seminoma cells specifically expressed the six gene products that were selected randomly from the list. These proteins include CCND2 and DNMT3A and may be useful as molecular pathological markers of seminoma.     

Possible role of VEGF in the progression of kidney disease in streptozotocin (STZ)-induced diabetic rats: effects of an ACE inhibitor and an angiotensin II receptor antagonist.

Two endothelium-derived factors, endothelin (ET), a vasoconstrictor, and vascular endothelial growth factor (VEGF), an angiogenic factor are thought to be involved in the pathogenesis of diabetic vascular complications. The aim of this study was to determine the effects of an angiotensin II type I (AT-1) receptor antagonist and an ACE inhibitor on the pathogenesis of VEGF and ET-1-mediated kidney disease in STZ-induced diabetic rats. Two days after STZ administration, diabetic rats were treated for 8 weeks with enalapril maleate, an ACE inhibitor, candesartan cilexetil, an AT-1 receptor antagonist, or saline. Urinary albumin and N-acetyl beta-D glucosaminidase (NAG) excretion as well as the VEGF protein content in the kidney were all found to be elevated in diabetic rats. Administration of enalapril maleate or candesartan cilexetil decreased the level of microalbuminuria and NAG excretion in diabetic rats. Administration of enalapril maleate also suppressed the elevated renal VEGF protein content in these animals while candesartan cilexetil treatment had no effect. Serum ET-1 and VEGF levels were unchanged by these treatments. These data support a role for AT-1 receptor antagonists and ACE inhibitors in the prevention of diabetic nephropathy, and suggest that the former may work by reducing renal VEGF levels.     

Rapid extraction of high-quality genomic DNA from Porphyra yezoensis (Bangiales, Rhodophyta)

We developed a simple, rapid and stable method for extraction of high molecular weight DNA from the marine red alga Porphyra yezoensis Ueda using both guanidium treatment and QIAGEN? kit (Funakoshi, Tokyo, Japan). The method does not require expensive equipment and complex steps. The DNA yield averaged 1.5 μg 100 mg^?1 of Porphyra tissue and the A260/A280 and A230/A260 ratios of the DNA were approximately 1.8 and 0.4, respectively. It was of sufficient quality to be used for not only polymerase chain reactions but also other DNA manipulation techniques such as restriction digestion and construction of genomic libraries.     

Hematological characteristics of rats spontaneously developing eosinophilia.

Hematological and genetic characteristics of newly found eosinophilic rats were studied. Hematologically, high blood eosinophil counts started at 6 weeks of age. Almost all 10-week-old rats had eosinophilia with individual counts above 500/microliter and 5 to 100 times the normal level. Proliferating eosinophils had normal morphology. An increase in lymphocyte counts was observed at 5 weeks of age, one week earlier than the onset of eosinophilosis. In bone marrow, proliferation of eosinophils was also observed at 8 weeks of age and thereafter progressed, suggesting a role in the pathogenesis of eosinophilia in this rat. The results of genetic cross experiments revealed the disease to be hereditary. The spontaneously eosinophilic rat therefore warrants attention as a model for studying the underlying mechanisms of human and animal eosinophilia.     

The plant aspartic proteinase-specific polypeptide insert is not directly related to the activity of oryzasin 1.

Many plant aspartic proteinases (APs) are different from animal and microbial APs in that they contain a polypeptide insert, approximately 100 amino acids in length, in the C-terminal region. To interpret the significance of this insert, we constructed an expression system for rice AP oryzasin 1 by linking a pro-oryzasin 1 downstream of glutathione S-transferase (GST). GST-proOS1 expressed the highest degree of hemoglobin-hydrolytic activity when treated at pH 3.3 and incubated for 24 h at room temperature. We carried out a similar experiment using an insert-lacking proOS1 mutant, GST-DeltaproOS1, as the fusion protein, and found it to show similar activity. This result indicates that the insert is not involved in the production of AP activity. We then investigated the autolysis of the two proteins by Western blot analysis. GST-proOS1 was autolyzed into 67- and 64-kDa fragments, while GST-DeltaproOS1 autolyzed to 54- and 52-kDa products. GST-DeltaproOS1 clearly produced two molecular species early in the autolytic process, and not later than 3 h from the start, but no such clear result was observed in the case of GST-proOS1. This suggests that, although the presence of the plant AP-specific insert does not influence the enzyme activity by itself, it apparently has an effect on the autolysis of OS1.