Distinct characteristics of circulating vascular endothelial growth factor-a and C levels in human subjects

The mechanisms that lead from obesity to atherosclerotic disease are not fully understood. Obesity involves angiogenesis in which vascular endothelial growth factor-A (VEGF-A) plays a key role. On the other hand, vascular endothelial growth factor-C (VEGF-C) plays a pivotal role in lymphangiogenesis. Circulating levels of VEGF-A and VEGF-C are elevated in sera from obese subjects. However, relationships of VEGF-C with atherosclerotic risk factors and atherosclerosis are unknown. We determined circulating levels of VEGF-A and VEGF-C in 423 consecutive subjects not receiving any drugs at the Health Evaluation Center. After adjusting for age and gender, VEGF-A levels were significantly and more strongly correlated with the body mass index (BMI) and waist circumference than VEGF-C. Conversely, VEGF-C levels were significantly and more closely correlated with metabolic (e.g., fasting plasma glucose, hemoglobin A1c, immunoreactive insulin, and the homeostasis model assessment of insulin resistance) and lipid parameters (e.g., triglycerides, total cholesterol (TC), low-density-lipoprotein cholesterol (LDL-C), and non-high-density-lipoprotein cholesterol (non-HDL-C)) than VEGF-A. Stepwise regression analyses revealed that independent determinants of VEGF-A were the BMI and age, whereas strong independent determinants of VEGF-C were age, triglycerides, and non-HDL-C. In apolipoprotein E-deficient mice fed a high-fat-diet (HFD) or normal chow (NC) for 16 weeks, levels of VEGF-A were not significantly different between the two groups. However, levels of VEGF-C were significantly higher in HFD mice with advanced atherosclerosis and marked hypercholesterolemia than NC mice. Furthermore, immunohistochemistry revealed that the expression of VEGF-C in atheromatous plaque of the aortic sinus was significantly intensified by feeding HFD compared to NC, while that of VEGF-A was not. In conclusion, these findings demonstrate that VEGF-C, rather than VEGF-A, is closely related to dyslipidemia and atherosclerosis.     

The E3 ubiquitin ligase activity of Trip12 is essential for mouse embryogenesis

Protein ubiquitination is a post-translational protein modification that regulates many biological conditions. Trip12 is a HECT-type E3 ubiquitin ligase that ubiquitinates ARF and APP-BP1. However, the significance of Trip12 in vivo is largely unknown. Here we show that the ubiquitin ligase activity of Trip12 is indispensable for mouse embryogenesis. A homozygous mutation in Trip12 (Trip12(mt/mt)) that disrupts the ubiquitin ligase activity resulted in embryonic lethality in the middle stage of development. Trip12(mt/mt) embryos exhibited growth arrest and increased expression of the negative cell cycle regulator p16. In contrast, Trip12(mt/mt) ES cells were viable. They had decreased proliferation, but maintained both the undifferentiated state and the ability to differentiate. Trip12(mt/mt) ES cells had increased levels of the BAF57 protein (a component of the SWI/SNF chromatin remodeling complex) and altered gene expression patterns. These data suggest that Trip12 is involved in global gene expression and plays an important role in mouse development.     

Connexin32 is expressed in vascular endothelial cells and participates in gap-junction intercellular communication

Endothelial cells (ECs) play many roles in vascular biology, including control of blood pressure, blood clotting, atherosclerosis, angiogenesis, and inflammation. Gap junctions (GJs) are channel-like assemblies of connexin (Cx) family proteins that connect neighboring cells and modulate and synchronize their intracellular environments by the transfer of intracellular mediators. It has been reported that vascular ECs express Cx37, Cx40, and Cx43, but not Cx32. Here, we showed that Cx32 mRNA and protein are expressed in various cultured human ECs. We confirmed Cx32 expression in blood vessel ECs using wild-type and Cx32 knock-out mice. We observed that dye transfer between cultured ECs through gap junctions is suppressed by an anti-Cx32 monoclonal antibody. These findings suggest that vascular ECs express Cx32, which participates in endothelial gap-junction intercellular communication.     

Involvement of MAP3K8 and miR-17-5p in Poor Virologic Response to Interferon-Based Combination Therapy for Chronic Hepatitis C

Despite advances in chronic hepatitis C treatment, a proportion of patients respond poorly to treatment. This study aimed to explore hepatic mRNA and microRNA signatures involved in hepatitis C treatment resistance. Global hepatic mRNA and microRNA expression profiles were compared using microarray data between treatment responses. Quantitative real-time polymerase chain reaction validated the gene signatures from 130 patients who were infected with hepatitis C virus genotype 1b and treated with pegylated interferon-alpha and ribavirin combination therapy. The correlation between mRNA and microRNA was evaluated using in silico analysis and in vitro siRNA and microRNA inhibition/overexpression experiments. Multivariate regression analysis identified that the independent variables IL28B SNP rs8099917, hsa-miR-122-5p, hsa-miR-17-5p, and MAP3K8 were significantly associated with a poor virologic response. MAP3K8 and miR-17-5p expression were inversely correlated with treatment response. Furthermore, miR-17-5p repressed HCV production by targeting MAP3K8. Collectively, the data suggest that several molecules and the inverse correlation between mRNA and microRNA contributed to a host genetic refractory hepatitis C treatment response.     

Surface components of shigellae involved in adhesion and haemagglutination

Strains of Shigella species were studied for their ability to adhere and agglutinate mammalian erythrocytes. Shigella dysenteriae and Sh. flexneri exhibited haemagglutinating (HA) properties when cultured in Casamino Acids-Yeast Extract (CYE) broth in the presence of 1 mmol 1^-1 calcium chloride, but other shigellae did not show this property under the same culture conditions. Repeated subcultivation of Sh. boydii, Sh. sonnei and HA negative strains of Sh. dysenteriae and Sh. flexneri in CYE broth medium induced adhesive and haemagglutinating properties that were inhibited by sodium periodate. HA activities of Shigella spp. were also inhibited by N -acetylneuraminic acid, α₁-glycoprotein and fetuin, but not by protease. Electron microscopy of Sh. dysenteriae 1, Sh. flexneri 2a, Sh. boydii 12 and Sh. sonnei 1 grown in CYE broth showed the presence of an extracellular slime layer that promoted agglutination of erythrocytes. The slime layer extracted from the cell surface of Shigella spp. showed HA properties, whereas lipopolysaccharide (LPS) obtained from the same strains, except Sh. dysenteriae 1, did not agglutinate erythrocytes. This evidence suggests that the cell surface haemagglutinin is a loosely bound slime layer which is expressed in CYE broth medium.     

Geographical distribution of Polycelis (Seidlia) auriculata in Japan

Polycelis (Seidlia) auriculata is endemic to mountain districts of Japan, from the central part of Honshû to the area of the Daisetsu Mts of Hokkaidô. In northern Japan, it sometimes occurs in cold-water biotopes of lowland areas. The progenitor of P. auriculata appears to have been the oldest immigrant into northern Japan among the Japanese Polycelis species, entering through a northern route as a preglacial faunal element. P. auriculata now shows a discontinious distribution in northern Japan. By virtue of its geographical and vertical distribution, ecological niche, variation in anatomy of the copulatory apparatus, and cytodemes, this species appears to be in the process of transformation.     

Inhibition of ammonium assimilation restores elongation of seminal rice roots repressed by high levels of exogenous ammonium

Elongation of seminal and lateral roots of rice seedlings was markedly inhibited by high ammonium levels in growth medium. However, high exogenous nitrate concentrations had little inhibitory effect on root growth. The objective of this study was to elucidate the relationship between inhibition of rice root growth induced by high ammonium conditions and ammonium assimilation in the seedlings. Activity of glutamine synthetase (GS) was kept at a low level in the seminal roots of the seedlings grown under high nitrate levels. In contrast, high ammonium levels significantly enhanced the GS activity in the roots, so that Gln abundantly accumulated in the shoots. These results indicate that ammonium assimilation may be activated in the seminal roots under high ammonium conditions. Application of methionine sulfoximine (MSO), an inhibitor of GS, relieved the repression of the seminal root elongation induced by high ammonium concentrations. However, the elongation of lateral roots remained inhibited even under the same condition. Furthermore, MSO drastically increased ammonium level and remarkably decreased Gln level in the shoots grown under high ammonium conditions. These results show that, for rice seedlings, an assimilatory product of ammonium, and not ammonium itself, may serve as an endogenous indicator of the nitrogen status involved in the inhibition of seminal root elongation induced by high levels of exogenous ammonium.     

Functional dissection of human and mouse POT1 proteins

The single-stranded telomeric DNA binding protein POT1 protects mammalian chromosome ends from the ATR-dependent DNA damage response, regulates telomerase-mediated telomere extension, and limits 5'-end resection at telomere termini. Whereas most mammals have a single POT1 gene, mice have two POT1 proteins that are functionally distinct. POT1a represses the DNA damage response, and POT1b controls 5'-end resection. In contrast, as we report here, POT1a and POT1b do not differ in their ability to repress telomere recombination. By swapping domains, we show that the DNA binding domain of POT1a specifies its ability to repress the DNA damage response. However, no differences were detected in the in vitro DNA binding features of POT1a and POT1b. In contrast to the repression of ATR signaling by POT1a, the ability of POT1b to control 5'-end resection was found to require two regions in the C terminus, one corresponding to the TPP1 binding domain and a second representing a new domain located between amino acids (aa) 300 and 350. Interestingly, the DNA binding domain of human POT1 can replace that of POT1a to repress ATR signaling, and the POT1b region from aa 300 to 350 required for the regulation of the telomere terminus is functionally conserved in human POT1. Thus, human POT1 combines the features of POT1a and POT1b.     

Mannose-specific lectin from the mushroom Hygrophorus russula

A lectin was purified from the mushroom Hygrophorus russula by affinity chromatography on a Sephadex G-50 column and BioAssist S cation exchange chromatography and designated H. russula lectin (HRL). The results of sodium dodecyl sulfate-polyaclylamidegel electrophoresis, gel filtration and matrix-assisted laser desorption ionization time-of-flight mass spectrometry of HRL indicated that it was composed of four identical 18.5?kDa subunits with no S-S linkage. Isoelectric focusing of the lectin showed bands near pI 6.40. The complete sequence of 175 amino acid residues was determined by amino acid sequencing of intact or enzyme-digested HRL. The sequence showed homology with Grifola frondosa lectin. The cDNA of HRL was cloned from RNA extracted from the mushroom. The open reading frame of the cDNA consisted of 528?bp encoding 176 amino acids. In hemagglutination inhibition assay, α1-6 mannobiose was the strongest inhibitor and isomaltose, Glcα1-6Glc, was the second strongest one, among mono- and oligosaccharides tested. Frontal affinity chromatography indicated that HRL had the highest affinity for Manα1-6(Manα1-3)Manβ1-4GlcNAcβ1-4GlcNAc, and non-reducing terminal Manα1-6 was essential for the binding of HRL to carbohydrate chains. The sugar-binding specificity of HRL was also analyzed by using BIAcore. The result from the analysis exhibited positive correlations with that of the hemagglutination inhibition assay. All the results suggested that HRL recognized the α1-6 linkage of mannose and glucose, especially the Manα1-6 bond. HRL showed a mitogenic activity against spleen lymph cells of an F344 rat. Furthermore, an enzyme-linked immunosorbent assay showed strong binding of HRL to human immunodeficiency virus type-1 gp120.