Lymphocyte plasma membranes II. Cytochemical localization of 5′-nucleotidase in rat lymphocytes

Cytochemical studies of thymic and splenic lymphocytes from rats showed that 5′-nucleotidase was restricted to the plasma membranes. Isolated plasma membranes contained the highest specific activity of 5′-nucleotidase of any cellular fractions. The results indicate that this enzyme can be used as a plasma membrane marker for lymphocytes.     

Optical analysis of RE3+ (RE = Nd or Er):B2O3–P2O5–CaF2–ZnO–(Li2O/Na2O/K2O) glasses

Calcium boro fluoro zinc phosphate glasses modified using alkali oxide and doped with Nd³⁺ and Er³⁺ ions with the chemical composition of 69.5 (B₂O₃) + 10 (P₂O₅) + 10 (CaF₂) + 5 (ZnO) + 5 (Na₂O/Li₂O/K₂O) + 0.5 (Er₂O₃/Nd₂O₃) were prepared using a conventional melt quenching technique. The results of X-ray diffraction patterns indicated the amorphous nature of all the prepared glasses. The visible–near-infrared red (NIR) absorption spectra of these glasses were analyzed systematically. The NIR emission spectra of Er³⁺ and Nd³⁺:calcium boro fluoro zinc phosphate glasses showed prominent emission bands at 1536 nm (⁴I_13/2→⁴I_15/2) and 1069 nm (⁴F_3/2→⁴I_11/2) respectively with λ_exci = 514.5 nm (Ar⁺ laser) as the excitation source.     

Stability of sheath blight resistance in transgenic ASD16 rice lines expressing a rice <Emphasis Type="Italic">chi11</Emphasis> gene encoding chitinase

Development of transgenic plants by introducing defense genes is one of the strategies to engineer disease resistance. Transgenic ASD16 rice plants harbouring rice chitinase chi11 gene, belonging to a PR-3 group of defense gene conferring sheath blight (Rhizoctonia solani Kuhn) resistance, were used in this study. Three T₂ homozygous lines (ASD16-4-1-1, 5-1-1, and 6-1-1) were identified from seven putative (T₀) transgenic lines expressing chi11 using Western blotting analysis. The inheritance of sheath blight resistance in those lines was studied over generations. The stability of chi11 expression up to T₄ generation in all the three homozygous lines was proved by Western blot and the stability of sheath blight resistance in the homozygous lines was proved up to T₄ generation using detached leaf and intact leaf sheath assays. Among the three homozygous lines tested, ASD16-4-1-1 showed consistent results in all the generations and gave a better protection against the sheath blight pathogen than the other two lines.     

Pre- and post-challenge immuno-haematological changes in Labeo rohita juveniles fed gelatinised or non-gelatinised carbohydrate with n-3 PUFA

The combined effect of dietary carbohydrate type and n-3 PUFA (EPA+DHA) on pre- and post-challenge haemato-immunological responses in Labeo rohita juveniles was studied. Fish were fed for 67days with six different test diets containing either gelatinised (G) or non-gelatinised (NG) corn (43%) with three levels of n-3 PUFA (0.5%, 1.0% and 2.0%). During the pre-challenge period, significantly higher (P<0.05) NBT, serum lysozyme activity, total protein and globulin content was recorded in the NG carbohydrate fed groups. Highest NBT value was recorded in the groups fed with 1.0% n-3 PUFA, whereas the highest serum lysozyme activity (P<0.05) was recorded at either 0.5% or 2.0% n-3 PUFA fed groups in both the pre- and post-challenge period. Feeding of NG corn significantly increased the total leucocyte count, lysozyme activity, A/G ratio and decreased the total erythrocyte count, haemoglobin, serum total protein and globulin content of L. rohita juveniles during the post-challenge period. Similarly, feeding of n-3 PUFA at any level significantly increased the immunological parameters like lysozyme activity or A/G ratio, whereas total leukocyte count increased due to feeding of either 0.5% or 1.0% n-3 PUFA. The NBT and albumin values remained similar in both the pre- and post-challenge period. After challenge with Aeromonas hydrophila, the highest survival was recorded in the NG carbohydrate fed groups, whereas the lowest survival was recorded in the highest level of n-3 PUFA fed group irrespective of dietary carbohydrate type. Thus, a high level of G carbohydrate as well as n-3 PUFA is found to be immunosuppressive in L. rohita juveniles. NG carbohydrate supplemented with 1.0% n-3 PUFA is found to be optimum to enhance the immunity in L. rohita juveniles.     

Proteolytic cleavage of bovine herpesvirus 1 (BHV-1) glycoprotein gB is not necessary for its function in BHV-1 or pseudorabies virus.

Glycoprotein B homologs represent the most highly conserved group of herpesvirus glycoproteins. They exist in oligomeric forms based on a dimeric structure. Despite the high degree of sequence and structural conservation, differences in posttranslational processing are observed. Whereas gB of herpes simplex virus is not proteolytically processed after oligomerization, most other gB homologs are cleaved by a cellular protease into subunits that remain linked via disulfide bonds. Proteolytic cleavage is common for activation of viral fusion proteins, and it has been shown that herpesvirus gB homologs are essential for membrane fusion events during infection, e.g., virus penetration and direct viral cell-to-cell spread. To analyze the importance of proteolytic cleavage for the function of gB homologs, we isolated a mutant bovine herpesvirus 1 (BHV-1) expressing a BHV-1 gB that is no longer proteolytically processed because of a deletion of the proteolytic cleavage site and analyzed its phenotype in cell culture. We showed previously that BHV-1 gB can functionally substitute for the homologous glycoprotein in pseudorabies virus (PrV), based on the isolation of a PrV gB-negative PrV recombinant that expresses BHV-1 gB (A. Kopp and T. C. Mettenleiter, J. Virol, 66:2754-2762, 1992). Therefore, we also isolated a mutant PrV lacking PrV gB but expressing a noncleavable BHV-1 gB. Our results show that cleavage of BHV-1 gB is not essential for its function in either a BHV-1 or a PrV background. Compared with the PrV recombinant expressing cleavable BHV-1 gB, deletion of the cleavage site in the recombinant PrV did not detectably alter the viral phenotype, as analyzed by plaque assays, one-step growth kinetics, and penetration kinetics. In the BHV-1 mutant, the uncleaved BHV-1 gB was functionally equivalent to the wild-type protein with regard to penetration and showed only slightly delayed one-step growth kinetics compared with parental wild-type BHV-1. However, the resulting plaques were significantly smaller, indicating a role for proteolytic cleavage of BHV-1 gB in cell-to-cell spread of BHV-1.     

Significant role of estrogen and progesterone receptor sequence variants in gallbladder cancer predisposition: a multi-analytical strategy

Background

Carcinoma of gallbladder (GBC) is an aggressive malignancy. The higher incidence of gallbladder cancer in women has been partly attributed to hormonal factors. Therefore the present study was designed to explore the role of genetic variants in estrogen (ESR1, ESR2) and progesterone (PGR) receptors in conferring risk of gallbladder cancer.

Materials and Methods

The present case-control study recruited total of 860 subjects, including 410 GBC patients, 230 gallstone patients and 220 controls. We examined the associations of 6 selected polymorphisms in three genes: ESR1 (rs2234693, rs9340799, rs1801132), ESR2 (rs1271572, rs1256049) and PGR (rs1042838) with GBC risk. Genotyping for all the polymorphisms was done using PCR-RFLP. Multifactor dimensionality reduction and classification and regression tree approaches were combined with logistic regression to discover high-order gene-gene interactions in hormonal pathway.

Results

On comparing the genotype frequency distribution in gallstone and GBC patients with that of healthy subjects, the homozygous variant genotypes of ESR1-397TT (rs2234693) polymorphism showed significant risk for developing gallstone [odds ratio: OR = 2.9] and GBC [OR = 1.8] respectively. Detailed haplotypes analysis suggested that ESR1 T _rs2234693G _rs9340799C _rs1801132 have significant association in conferring risk for both gallstones [OR = 2.2] and GBC [OR = 3.0]. However, the variant-containing genotypes (DI+II) of PGR (rs1042838) showed low risk in both GBC [OR = 0.4] and gallstone patients [OR = 0.4].On performing the MDR analysis, ESR1 IVS1-397C>T, ESR1 IVS1-351A>G, and ESR2-789 A>C yielded the highest testing accuracy of 0.634. These results were further supported by the CART analysis which revealed that individuals with the combined genotypes of ESR1-397 CT or TT, ESR1-351 AG or GG and ESR2 -789 AA had the highest risk for GBC [OR  = 3.9].

Conclusion

Using multi-analytical approaches, our study showed important role of ESR1 IVS1-397C>T, ESR1 IVS1-351A>G, and ESR2-789 A>C variants in GBC susceptibility and the risk appears to be mediated through gallstone dependent pathway.     

Traits and selection strategies to improve root systems and water uptake in water-limited wheat crops

Wheat yields globally will depend increasingly on good management to conserve rainfall and new varieties that use water efficiently for grain production. Here we propose an approach for developing new varieties to make better use of deep stored water. We focus on water-limited wheat production in the summer-dominant rainfall regions of India and Australia, but the approach is generally applicable to other environments and root-based constraints. Use of stored deep water is valuable because it is more predictable than variable in-season rainfall and can be measured prior to sowing. Further, this moisture is converted into grain with twice the efficiently of in-season rainfall since it is taken up later in crop growth during the grain-filling period when the roots reach deeper layers. We propose that wheat varieties with a deeper root system, a redistribution of branch root density from the surface to depth, and with greater radial hydraulic conductivity at depth would have higher yields in rainfed systems where crops rely on deep water for grain fill. Developing selection systems for mature root system traits is challenging as there are limited high-throughput phenotyping methods for roots in the field, and there is a risk that traits selected in the lab on young plants will not translate into mature root system traits in the field. We give an example of a breeding programme that combines laboratory and field phenotyping with proof of concept evaluation of the trait at the beginning of the selection programme. This would greatly enhance confidence in a high-throughput laboratory or field screen, and avoid investment in screens without yield value. This approach requires careful selection of field sites and years that allow expression of deep roots and increased yield. It also requires careful selection and crossing of germplasm to allow comparison of root expression among genotypes that are similar for other traits, especially flowering time and disease and toxicity resistances. Such a programme with field and laboratory evaluation at the outset will speed up delivery of varieties with improved root systems for higher yield.     

Oxidant-antioxidant imbalance in the erythrocytes of sporadic amyotrophic lateral sclerosis patients correlates with the progression of disease

Free radicals are implicated in numerous disease processes including motor neuron degeneration (MND). Antioxidant defense enzymes: superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSHPx), glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G-6-PDH) in the erythrocytes are capable of detoxifying reactive oxygen species produced endogenously or exogenously. In the present study, the extent of lipid peroxidation (LPO) and antioxidant defenses were evaluated in the erythrocytes of 20 sporadic amyotrophic lateral sclerosis (ALS) patients and 20 controls. We observed that lipid peroxidation in the erythrocytes of amyotrophic lateral sclerosis patients significantly increased with respect to controls (P<0.001). On the other hand, catalase activity was found to be significantly lower (P<0.001). The activities of glucose-6-phosphate dehydrogenase, glutathione reductase and glutathione levels were also found to be significantly reduced in ALS patients compared to healthy subjects (P<0.001, P<0.01 and P<0.01, respectively). It was further observed that lipid peroxidation started to increase and catalase, glutathione reductase, glucose-6-phosphate dehydrogenase enzyme activities and glutathione levels started to decrease as amyotrophic lateral sclerosis progressed from 6 to 24 months, suggesting a correlation between these parameters and duration of amyotrophic lateral sclerosis. This study confirms the involvement of oxidative stress during the progression of amyotrophic lateral sclerosis and the need to develop specific peripheral biomarkers.