Effect of chitosan on tobacco mosaic virus（TMV） accumulation,hydrolase activity,and morphological abnormalities of the viral particles in leaves of N. tabacum L. cv. Samsun

The effect of chitosan on the development of infection caused by Tobacco mosaic virus（TMV） in leaves of Nicotiana tabacum L. cv. Samsun has been studied. It was shown that the infectivity and viral coat protein content in leaves inoculated with a mixture of TMV（2 μg/mL） and chitosan（1 mg/mL） were lower in the early period of infection（3 days after inoculation）, by 63% and 66% respectively, than in leaves inoculated with TMV only. Treatment of leaves with chitosan 24 h before inoculation with TMV also caused the antiviral effects, but these were less apparent than when the virus and polysaccharide were applied simultaneously. The inhibitory effects of the agent decreased as the infection progressed. Inoculation of leaves with TMV together with chitosan considerably enhanced the activity of hydrolases（proteases, RNases） in the leaves, in comparison with leaves inoculated with TMV alone. Electron microscope assays of phosphotungstic acid（PTA）-stained suspensions from infected tobacco leaves showed that, in addition to the normal TMV particles（18 nm in diameter, 300 nm long）, these suspensions contained abnormal（swollen, “thin” and “short”） virions. The highest number of abnormal virions was found in suspensions from leaves inoculated with a mixture of TMV and chitosan. Immuno-electron microscopy showed that “thin” virus particles, in contrast to the particles of normal diameter, lost the ability to bind to specific antiserum. It seems that the chitosan-induced activation of hydrolases stimulates the intracellular degradation of TMV particles and hence hydrolase activation may be considered to be one of the polysaccharide-mediated cellular defense mechanisms that limit virus accumulation in cells.     

Modification of phenolic metabolism in soybean hairy roots through down regulation of chalcone synthase or isoflavone synthase

Soybean hairy roots, transformed with the soybean chalcone synthase (CHS6) or isoflavone synthase (IFS2) genes, with dramatically decreased capacity to synthesize isoflavones were produced to determine what effects these changes would have on susceptibility to a fungal pathogen. The isoflavone and coumestrol concentrations were decreased by about 90% in most lines apparently due to gene silencing. The IFS2 transformed lines had very low IFS enzyme activity in microsomal fractions as measured by the conversion of naringenin to genistein. The CHS6 lines with decreased isoflavone concentrations had 5 to 20-fold lower CHS enzyme activities than the appropriate controls. Both IFS2 and CHS transformed lines accumulated higher concentrations of both soluble and cell wall bound phenolic acids compared to controls with higher levels found in the CHS6 lines indicating alterations in the lignin biosynthetic branch of the pathway. Induction of the soybean phytoalexin glyceollin, of which the precursor is the isoflavone daidzein, by the fungal pathogen Fusarium solani f. sp. glycines (FSG) that causes soybean sudden death syndrome (SDS) showed that the low isoflavone transformed lines did not accumulate glyceollin while the control lines did. The (iso)liquritigenin content increased upon FSG induction in the IFS2 transformed roots indicating that the pathway reactions before this point can control isoflavonoid synthesis. The lowest fungal growth rate on hairy roots was found on the FSG partially resistant control roots followed by the SDS sensitive control roots and the low isoflavone transformants. The results indicate the importance of phytoalexin synthesis in root resistance to the pathogen. Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable.     

Characteristics of the active lymph pump in bovine prenodal mesenteric lymphatics

BACKGROUND: The functional characteristics of the bovine mesenteric postnodal lymphatics are well-described. However there are no reports of pumping characteristics of the bovine mesenteric prenodal lymphatics. We propose that the prenodal lymphatics have adapted to the local conditions of lymph flow and are functioning differently than the postnodal vessels. METHODS AND RESULTS: To evaluate pumping in bovine prenodal mesenteric lymphatics, we observed their contractility in response to the changes in transmural pressure and imposed flow. Lymphatics (diameter approximately 460 microm) were isolated, cannulated, and pressurized. Lymphatic diameters were traced from video records; the lymphatic tone index, contraction amplitude and frequency, lymphatic pump indices were calculated. Increasing transmural pressure from 3 to 6 cm H2O produced a strong inotropic response, but did not induce a significant chronotropic response. Pumping reached its maximum at transmural pressures 6-9 cm H2O and was not significantly depressed up to 15 cm H2O, whereas pumping in postnodal lymphatics is typically depressed at transmural pressures higher than 10 cm H2O. Bovine prenodal mesenteric lymphatics also demonstrated very low sensitivity to the increases in imposed flow. CONCLUSIONS: We concluded that the functional heterogeneity exists on the intraregional levels in lymphatic nets.     

Independent associations of TOMM40 and APOE variants with body mass index

The TOMM40‐APOE variants are known for their strong, antagonistic associations with Alzheimer's disease and body weight. While a stronger role of the APOE than TOMM40 variants in Alzheimer's disease was suggested, comparative contribution of the TOMM40‐APOE variants in the regulation of body weight remains elusive. We examined additive effects of rs2075650 and rs157580 TOMM40 variants and rs429358 and rs7412 APOE variants coding the ε2/ε3/ε4 polymorphism on body mass index (BMI) in age‐aggregated and age‐stratified cohort‐specific and cohort‐pooled analysis of 27,863 Caucasians aged 20–100 years from seven longitudinal studies. Minor alleles of rs2075650, rs429358, and rs7412 were individually associated with BMI (β = ?1.29, p = 3.97 × 10^?9; β = ?1.38, p = 2.78 × 10^?10; and β = 0.58, p = 3.04 × 10^?2, respectively). Conditional analysis with rs2075650 and rs429358 identified independent BMI‐lowering associations for minor alleles (β = ?0.63, p = 3.99 × 10^?2 and β = ?0.94, p = 2.17 × 10^?3, respectively). Polygenic mega‐analysis identified additive effects of the rs2075650 and rs429358 heterozygotes (β = ?1.68, p = 3.00 × 10^?9), and the strongest BMI‐lowering association for the rs2075650 heterozygous and rs429358 minor allele homozygous carriers (β = ?4.11, p = 2.78 × 10^?3). Conditional analysis with four polymorphisms identified independent BMI‐lowering (rs2075650, rs157580, and rs429358) and BMI‐increasing (rs7412) associations of heterozygous genotypes with BMI. Age‐stratified conditional analysis revealed well‐powered support for a differential and independent association of the rs429358 heterozygote with BMI in younger and older individuals, β = 0.58, 95% confidence interval (CI) = ?1.18, 2.35, p = 5.18 × 10^?1 for 3,068 individuals aged ≤30 years and β = ?4.28, CI = ?5.65, ?2.92, p = 7.71 × 10^?10 for 6,052 individuals aged >80 years. TOMM40 and APOE variants are independently and additively associated with BMI. The APOE ε4‐coding rs429358 polymorphism is associated with BMI in older individuals but not in younger individuals.     

Mesenteric lymph flow in adult and aged rats

The objective of study was to evaluate the aging-associated changes, contractile characteristics of mesenteric lymphatic vessels (MLV), and lymph flow in vivo in male 9- and 24-mo-old Fischer-344 rats. Lymphatic diameter, contraction amplitude, contraction frequency, and fractional pump flow, lymph flow velocity, wall shear stress, and minute active wall shear stress load were determined in MLV in vivo before and after N(ω)-nitro-L-arginine methyl ester hydrochloride (L-NAME) application at 100 μM. The active pumping of the aged rat MLV in vivo was found to be severely depleted, predominantly through the aging-associated decrease in lymphatic contractile frequency. Such changes correlate with enlargement of aged MLV, which experienced much lower minute active shear stress load than adult vessels. At the same time, pumping in aged MLV in vivo may be rapidly increased back to levels of adult vessels predominantly through the increase in contraction frequency induced by nitric oxide (NO) elimination. Findings support the idea that in aged tissues surrounding the aged MLV, the additional source of some yet unlinked lymphatic contraction-stimulatory metabolites is counterbalanced or blocked by NO release. The comparative analysis of the control data obtained from experiments with both adult and aged MLV in vivo and from isolated vessel-based studies clearly demonstrated that ex vivo isolated lymphatic vessels exhibit identical contractile characteristics to lymphatic vessels in vivo.     

A novel approach to determining the affinity of protein-carbohydrate interactions employing adherent cancer cells grown on a biosensor surface

The development of biological agents for the treatment of solid tumours is an area of considerable activity. We are pursuing carbohydrate-binding proteins (lectins) in a strategy aimed at targeting cancer-associated changes in glycosylation. To evaluate lectin-cancer cell interactions we developed a novel cell biosensor in which binding events take place at the cell surface, more closely mimicking an in vivo system. Metastatic, SW620, and non-metastatic, SW480, colorectal cancer cells were grown on the surface of a tissue-culture compatible polystyrene coated biosensor chip and housed in a quartz crystal microbalance (QCM) apparatus, the kinetics of binding of a diverse range of lectins was evaluated. The lectin Helix pomatia agglutinin (HPA) has been shown to bind aggressive metastatic cancer and was produced in recombinant form (His- and RFP-tagged). The affinity of HPA was in the nanomolar range to the metastatic SW620 cells but was only in the micromolar range to the non-metastatic SW480. Overall, the dissociation constant (K(D)) of the lectins tested in the new cell biosensor system was an order of magnitude lower (nanomolar range) than has generally been reported with systems such as QCM/SPR. This new cell-biosensor enables molecular interactions to be studied in a more relevant environment. An intrinsic problem with developing new biological therapies is the difficulty in determining the affinity with which proteins will interact with intact cell surfaces. This methodology will be of interest to researchers developing new biological approaches for targeting cell surfaces in a wide range of diseases, including cancer.     

Microarray profiling of lymphocytes in internal diseases with an altered immune response: potential and methodology

Recently it has become possible to investigate expression of all human genes with microarray technique. The authors provide arguments to consider peripheral white blood cells and in particular lymphocytes as a model for the investigation of pathophysiology of asthma, RA, and SLE diseases in which inflammation is a major component. Lymphocytes are an alternative to tissue biopsies that are most often difficult to collect systematically. Lymphocytes express more than 75% of the human genome, and, being an important part of the immune system, they play a central role in the pathogenesis of asthma, RA, and SLE. Here we review alterations of gene expression in lymphocytes and methodological aspects of the microarray technique in these diseases. Lymphocytic genes may become activated because of a general nonspecific versus disease-specific mechanism. The authors suppose that in these diseases microarray profiles of gene expression in lymphocytes can be disease specific, rather than inflammation specific. Some potentials and pitfalls of the array technologies are discussed. Optimal clinical designs aimed to identify disease-specific genes are proposed. Lymphocytes can be explored for research, diagnostic, and possible treatment purposes in these diseases, but their precise value should be clarified in future investigation.     

Elevated minisatellite mutation rate in the post-chernobyl families from ukraine

Germline mutation at eight human minisatellite loci has been studied among families from rural areas of the Kiev and Zhitomir regions of Ukraine, which were heavily contaminated by radionuclides after the Chernobyl accident. The control and exposed groups were composed of families containing children conceived before and after the Chernobyl accident, respectively. The groups were matched by ethnicity, maternal age, parental occupation, and smoking habits, and they differed only slightly by paternal age. A statistically significant 1.6-fold increase in mutation rate was found in the germline of exposed fathers, whereas the maternal germline mutation rate in the exposed families was not elevated. These data, together with the results of our previous analysis of the exposed families from Belarus, suggest that the elevated minisatellite mutation rate can be attributed to post-Chernobyl radioactive exposure. The mechanisms of mutation induction at human minisatellite loci are discussed.     

Carbonate mineralogy of a tropical bryozoan biota and its vulnerability to ocean acidification

Decreasing pH levels in the world’s oceans are widely recognized as a threat to marine life. Bryozoans are among several phyla that produce calcium carbonate skeletons potentially affected by ocean acidification (OA). Depending on species, bryozoan skeletons can consist of calcite, aragonite or have a bimineralic combination of these two minerals. Aragonite is generally more soluble in seawater than calcite, making aragonitic species more vulnerable to OA. Here, for the first time we use Raman spectroscopy to determine the mineral composition of a tropical bryozoan biota. Compared with bryozoan biotas from higher latitudes in which calcite predominates, aragonite was found to occur in a much higher proportion of the 22 cheilostome bryozoan species collected from the shorelines of Penang and Langkawi in Malaysia, where 46% of species are calcitic, 41% aragonitic and 13% bimineralic. All but one of the aragonitic or bimineralic species belong to the ascophorans, whereas calcitic skeletons characterized most of the anascans, many of which are primitive ‘weedy’ malacostegines. These results suggest a relatively high vulnerability of tropical bryozoan faunas to OA, with the weedier taxa likely to be least impacted.