Angiogenic endothelium-specific nestin expression is enhanced by the first intron of the nestin gene

Nestin is a member of intermediate filaments abundantly expressed in neural stem cells and glioblastomas. The nestin gene has four exons and three introns, and neural cell-specific expression is regulated by the second intron. We previously reported that nestin was invariably detected in the tumor endothelium in gliomas even though tumor cells were negative for nestin. In the present study, we further confirmed nestin immunostaining in tumor endothelium of a variety of common cancers, including lung, stomach, colon, and cervical carcinomas. We examined an endothelium-specific regulator using human umbilical vein endothelial cells (HUVECs) and human glioblastoma-derived U251 cells. In a luciferase reporter assay, the first intron plus 5' upstream promoter (5'UP) gave the highest activity, followed by 5'UP, and the second intron plus 5'UP. However, the assay values were much lower by HUVEC extracts than by U251 cell extracts. Although green fluorescent protein expression was positive over all U251 cells under either the first intron, second intron, or ubiquitously active CAG promoter, the fluorescence in HUVECs was limited to a few cells even under the first intron. This difference came from the growth feature of HUVECs which exhibit growth arrest by contact inhibition. We found that the nestin expression was specific to proliferative endothelium, by using proliferation markers in hemangioblastomas and in situ hybridization. Using an endothelial tube formation assay, tyrosine kinase domain-deleted VEGF receptor KDR effectively abolished the tube formation under the first intron. We suggest that the nestin expression in tumor endothelium is enhanced by the first intron.     

Preparation of ceramic microspheres for in situ radiotherapy of deep-seated cancer

Radiotherapy is one of the most effective treatments for cancers. However, external irradiation provides only small doses to deep-seated cancers, and often causes damage to healthy tissues. It has been reported that 20-30 microm diameter 17Y(2)O(3)-19Al(2)O(3)-64SiO(2) (mol%) glass microspheres are useful for the in situ irradiation of cancers. Yttrium-89 (89Y) in this glass can be neutron bombarded to form the beta-emitter 90Y (half-life=64.1h). When injected in the vicinity of the cancer, such activated glass microspheres can provide a large localized dose of beta-radiation. The Y(2)O(3) content of the glass in the microspheres is limited to only 17 mol%. Chemically durable microspheres with a higher Y(2)O(3) content need to be developed. Phosphorus-31 (31P) with 100% natural abundance can also be activated by neutron bombardment to form the beta-emitter 32P (half-life=14.3d). Chemically durable microspheres containing a high phosphorus content are expected to be more effective for cancer treatment. We prepared pure Y(2)O(3) and YPO(4) microspheres using a high-frequency induction thermal plasma melting technique, and investigated the resulting structure and chemical durability. We successfully prepared smooth, highly spherical polycrystalline Y(2)O(3) and YPO(4) microspheres with diameters in the range 20-30 microm. Both the Y(2)O(3) and YPO(4) microspheres showed high chemical durability in saline solutions buffered at pH=6 and 7. These microspheres are expected to be more effective than the conventional glass microspheres for the in situ radiotherapy of cancer.     

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.     

Inhibitory effect of aerosol WP871 on SRS-A mediated bronchoconstriction in the guinea pig in vivo

Slow-reacting substance of anaphylaxis (SRS-A) is an important factor mediating bronchoconstriction in asthma. We developed a guinea pig model for SRS-A mediated bronchoconstriction induced by antigen inhalation. Using this model, we investigated the effect of inhaled WP871, a new anti-allergic drug, on bronchoconstriction. Aerosol WP871 (0.01 and 0.033%) to some extent inhibited the antigen-induced bronchoconstriction in a dose-dependent fashion, but high-dose WP871 (0.1%) inhalation itself produced a non-specific bronchoconstriction. However, aerosol WP871 (0.033%) showed no inhibitory effect on bronchoconstriction caused by direct inhalation of leukotriene C4, a component of SRS-A. These findings indicate that aerosol WP871 does not antagonize SRS-A, but inhibits synthesis and/or release of SRS-A and has some non-specific bronchoconstrictive effect in high concentration.     

Histamine downregulates CD14 expression via H2 receptors on human monocytes

Lipopolysaccharide (LPS) binds to LPS-binding protein (LBP) in plasma and is delivered to the cell surface receptor CD14 on human monocyte. LPS is transferred to the transmembrane signaling receptor toll-like receptor (TLR) 4. In the present study, the effect of histamine on the expression of CD14 on human monocytes was investigated. Histamine concentration- and time-dependently decreased the expression of cell surface CD14, whereas histamine did not decrease mRNA for CD14 nor increase soluble CD14 (sCD14). The inhibitory effects of histamine on CD14 expression were antagonized by H2-receptor antagonist, but not by H1 and H3/H4 antagonist. The effects of selective H2-receptor agonists, 4-methylhistamine and dimaprit, on CD14 expression mimicked that of histamine indicating that histamine regulated CD14 expression through the stimulation of H2-receptors. The pretreatment with histamine partially inhibited the LPS-induced TNF-alpha production in human peripheral blood mononuclear cells (PBMC). Such inhibition might be due to the down-regulation of CD14 expression on monocytes by histamine.     

Genomic structure and eight novel exonic polymorphisms of the human N-cadherin gene

 Analysis of the detailed genomic structure of human N-cadherin revealed that the 16-exon gene is more than 72 kb in length and that it consists of a mosaic of exons. Five repeated cadherin domains, a transmembrane domain, and a cytoplasmic domain are encoded by exons 4 to 13, 13 and 14, and 14 to 16, respectively. A search for molecular variants in the entire coding region in 96 Japanese individuals resulted in the identification of eight sequence polymorphisms including three CCT- or GCC-type trinucleotide repeat polymorphisms adjacent to the initiation codon and five other novel single-nucleoticle polymorphisms (SNPs) in the coding region. Three of the five SNPs accompanied an amino acid substitution: Ala118Thr, Ala826Thr, and Asn845Ser. Knowlege of the fine gene structure and eight novel polymorphisms will be useful for the genetic study of the role of N-cadherin in diseases involving cell adhesion in the brain and in cardiomyocytes. Received: January 23, 2002 / Accepted: March 12, 2002     

Phase stability after aging and its influence on pin-on-disk wear properties of Ce-TZP/Al2O3 nanocomposite and conventional Y-TZP

Recently zirconia/alumina composites have been examined by many researchers as the new generation of bearing materials in total joint replacements. In this study, the phase stability of a Ce-TZP/Al(2)O(3) nanocomposite and conventional Y-TZP after aging, and its influence on wear resistance, were investigated. Very slight phase transformation was observed in both types of ceramics 18 months after the implantation of Ce-TZP/Al(2)O(3) or Y-TZP samples into rabbit tibiae. However, Y-TZP showed marked phase transformation (approximately 80%) after aging in an autoclave (121 degrees C) for 190 h or in physiological saline at 62 degrees C for 18 months, whereas the new composite remained almost resistant to degradation. According to the results of self-pairing pin-on-disk wear tests using ceramic specimens with or without autoclave aging, the wear factor was almost the same between Ce-TZP/Al(2)O(3) samples with and without aging (6.74 +/- 0.36 x 10(-8) and 6.04 +/- 0.95 x 10(-8) mm(3)/Nm, respectively). In contrast, although non-aged Y-TZP had the lowest wear factor (4.88 +/- 0.51 x 10(-8) mm(3)/Nm) of all specimens tested, aged Y-TZP showed 10-fold greater wear than nonaged Y-TZP. The present study suggests that Ce-TZP/Al(2)O(3) nanocomposite has much greater phase stability than Y-TZP, and that its wear properties are not influenced by aging.