Interleukin-4 inhibits the vascular endothelial growth factor- and basic fibroblast growth factor-induced angiogenesis in vitro

The role of interleukin-4 (IL-4) in the inflammatory process has emerged recently. In this study, we investigated the effect of IL-4 on the angiogenic process in an in vitro experimental system. IL-4 significantly inhibited the proliferation of human umbilical vein endothelial cells (HUVEC) that was induced by the vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). VEGF- or bFGF-induced HUVEC chemotaxis was abrogated by the IL-4 treatment. In addition, the formation of tube-like structures by HUVEC in the presence of VEGF or bFGF was also severely down-regulated by IL-4. The inhibitory effects on the critical steps of angiogenesis were not observed with IL-6 that is abundantly found in the inflamed tissue. Our results suggest that IL-4 may play a regulatory role in normal physiology and provide the potential possibility for IL-4 as a therapeutic agent in the intervention of angiogenesis-related diseases.     

洋葱"昌激99-3"的激光诱变选育

采用CO2和He-Ne两种激光的三种剂量,分别辐照两个洋葱品种的湿种子,从He-Ne和CO2激光辐照元谋洋葱变异后代中选育出高产、优质、多抗、适应性较强的红皮洋新品种“昌激99-3”,经专家组实地验收亩产达9186.7Kg,是红皮洋葱经专家正式验收的全国最高产量。     

Molecular diagnosis of Duchenne/Becker muscular dystrophy by polymerase chain reaction and microsatellite analysis

Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are X-linked recessive genetic disorders resulting from mutations in the dystrophin gene. About two-thirds of the affected patients have large deletions or duplications, which occur in the 5' and central region of the gene. The remaining DMD/BMD cases show no deletions, so they cannot be easily identified by current strategies. In these DMD/BMD families, a linkage analysis that involves DNA markers of the flanking and intragenic dystrophin gene are necessary for carrier and prenatal diagnosis. We analyzed eighteen deletion-prone exons of the gene by a polymerase chain reaction (PCR) in order to characterize the molecular defects of the dystrophin gene in Korean DMD/BMD families. We also performed a linkage analysis to assess the usefulness and application of six short tandem repeat markers for molecular diagnosis in the families. We observed a deletion that eliminated the exon 50. Also, a linkage analysis in the families with six short tandem repeat (STR) markers showed heterozygosity at most of the STR markers. The haplotype analysis was useful for detecting the carrier status. This study will be helpful for a molecular diagnosis of DMD/BMD families in the Korean population.     

Development of a new xenoestrogen screening system using fission yeast Schizosaccharomyces pombe

Endocrine disrupters refer to environmental or chemical compounds, which interfere with the endocrine system of organisms. In this study, our aim was to develop a screening method to detect xenoestrogen (an endocrine disrupter that is commonly encountered in our daily life) by using fission yeast Schizosaccharomyces pombe. Although the yeast (the simplest eukaryotic cell) has no endocrine system, estrogen receptors that are created to express in the yeast cell can be activated by estrogen in a similar manner to mammalian cells. First, in order to express the human estrogen receptor (hER) in the yeast cell, we constructed a yeast expression vector that contained hER (pREP42MHN-hER). In the yeast cells that are transformed with the pREP42MHN-hER vector, estrogen receptors could recognize xenoestrogen, which allowed the determination of the presence of xenoestrogen in any given sample. Furthermore, we constructed a yeast strain that contained an estrogen responsive element (ERE) that fused with the Escherichia coli LacZ gene (pERE-LacZ) as a reporter for binding of xenoestrogen with the estrogen receptor. Since this vector system allows determination of the presence and level of xenoestrogen with simple procedures, it is expected that they can serve as an efficient assay system to detect xenoestrogen.     

Regulation of septation and cytokinesis during resumption of cell division requires uvi31+, a UV-inducible gene of fission yeast

uvi31+ is a sequence homolog of Escherichia coli bolA gene in Schizosaccharomyces pombe, identified as a UV-inducible gene. Here, the cellular function of uvi31+ was investigated by null mutant analysis. Deletion of uvi31+ led to a delayed germination of spore and defects in subsequent cell division. However, the uvi31 mutant cell proliferated faster with smaller cell size than the wild-type cell during vegetative growth. In addition, the uvi31 mutant was sensitive to UV-light. It showed a normal cell cycle delay after UV-irradiation but displayed aberrant septum formation and defective cytokinesis when released from the UV damage checkpoint. These results suggest that uvi31+ may be involved in control of cell division, especially during the resumption from cell cycle arrest.     

Morphology of calretinin-immunoreactive neurons in the superficial layers of hamster superior colliculus after enucleation: lack of co-localization with GABA

We recently reported on the distribution and effects of eye enucleation on the immunoreactivity of calretinin in the superficial layers of the hamster superior colliculus (SC). In the present study, we describe the types of labeled cells and compare this labeling to that of GABA, the major inhibitory neurotransmitter in the central nervous system. An almost complete depletion of calretinin-immunoreactive (IR) fibers in the superficial layers of the contralateral SC was found following unilateral enucleation. On the contralateral SC, many calretinin-IR cells were newly appeared. The majority of the newly-appeared cells had small- to medium-sized round, oval, or vertical fusiform cell bodies. Two-color immunofluorescence revealed that none of these newly-appeared cells were labeled with an antibody to GABA. The present results show that the calretinin-IR cells are unique in the superficial hamster SC when compared to most of the other brain areas, where many calretinin-IR cells are GABAergic interneurons.     

Reprogramming of the activity of the activator/dissociation transposon family during plant regeneration in rice

Many aspects of epigenetic phenomena have been elucidated via studies of transposable elements. An active transposable element frequently loses its ability to mobilize and goes into an inactive state during development. In this study, we describe the cyclic activity of a maize transposable element dissociation (Ds) in rice. In rice genome, Ds undergoes the spontaneous loss of mobility. However, an inactive state of Ds can be changed into an active state during tissue culture. The recovery of mobility accompanies not only changes in the methylation patterns of the terminal region of Ds, but also alteration in the steady state level of the activator (Ac) mRNA that is expressed by a constitutive CaMV 35S promoter. Furthermore, the Ds-reactivation process is not random, but stage-specific during plantlet regeneration. Our findings have expanded previous observations on Ac reactivation in the tissue culture of maize.     

Characterization of mycoplasma arginine deiminase expressed in E. coli and its inhibitory regulation of nitric oxide synthesis

We previously reported that a cytostatic protein that is found in ASC-17D Sertoli cell-conditioned media was Mycoplasma arginine deiminase (ADI), which hydrolyzes L-arginine into L-citrulline and ammonia. Here, we report the over-expression of recombinant ADI (rADI) in E. coli and the down-regulation of lipopolysaccharide (LPS) induced-nitric oxide (NO) production by rADI treatment. We cloned the ADI gene from Mycoplasma arginini genomic DNA by a polymerase chain reaction, and changed five TGA tryptophan codons (stop codon in E. coli) to TGG codons in the coding region by site-directed mutagenesis in order to express in E. coli. The rADI was purified to apparent homogeneity by DEAE-Sepharose and arginine-affinity chromatography. The rADI expressed in E. coli was identified as 45 kDa on SDS-PAGE and 90 kDa on native PAGE, implying that it exists as a dimer like ADI of M. arginini. The Km for arginine of rADI was approximately 370+/-50 microM. Its optimal temperature and pH were 41 degrees C and pH 6.4, respectively, and enzyme activity remained > or = 50% for 5 d at physiological temperature and pH. Treatment of purified rADI suppressed NO production in macrophage-like RAW 264.7 and primary glial cells that were exposed to LPS. Furthermore, an intraperitoneal injection of rADI significantly suppressed the rise of blood nitrite/nitrate levels that were induced by the systemic administration of bacterial endotoxin LPS to mice, resulting in an improvement in their survival rate. These results suggest that the depletion of blood arginine with an arginine-metabolizing enzyme, such as ADI, could suppress excessive production of NO that is caused by inducible NOS (iNOS) during the endotoxemia. Also, rADI may be used as a new approach to control NO-related diseases, such as sepsis.     

A direct repeat of N-type Ca2+ channel alpha1B gene functions as a negative regulatory element in HeLa cells

The expression of the N-type voltage-gated calcium channel alpha1B gene is restricted to neurons by a 5'-upstream region (-3992 to -1788) that contains negative regulatory element(s) that are active in non-neuronal cells. A 39 bp DNA element, which is repeated nine times in a head-to-tail fashion, was found within the same region. To examine whether this direct repeat (DR) may function as a negatively acting cis-regulatory element, several fusion plasmids, DR-110alpha1BLUC (1X), DR-SV40LUC (IX, 2X), in which one or two copies of the DR fragment were subcloned upstream of the homologous and heterologous promoters, were transiently transfected into HeLa and NS20Y cells. The promoter activity of DR-110alpha1BLUC (1X) decreased to approximately 17% of the 110alph(a1B)LUC construct in HeLa cells. The expression of the DR-SV40LUC (1X) and DR-SV40LUC (2X) plasmids was also reduced to 50 to 23% of the levels that were observed in the pGL2-Promoter in the same cells. However, no repression of the DR constructs was observed in NS20Y cells. An electrophoretic mobility shift assay showed that two DR-specific complexes were detected in HeLa cells, but not in NS20Y cells. In addition, Southwestern blotting revealed the presence of approximately 33 and 43 kDa proteins in HeLa cells. Overall, these results suggest that a 39 bp DNA element might act as repressor in non-neuron cells through the specific interactions of the DNA-proteins.